strychnine and brucine

Nicotine, a lipid-soluble alkaloid obtained from the dried leaves of Nicotiana, is most frequently encountered in tobacco products for smoking, chewing or sniffing as well as in a limited number of pesticides. Though nicotine is one of the most toxic drugs of abuse, it has rarely led to fatalities. Sudden death can be caused by cardiovascular arrest, respiratory muscle paralysis and/or central respiratory failure. A 42-year-old man was found dead by his wife. He was lying on the floor, next to a box containing many empty bottles of beer and vodka. Some labeled chemical bottles found at the scene contained various substances, including nicotine and brucine. Gross examination of the organs at autopsy revealed no specific findings. The toxicological examination failed to disclose any lethal toxic agents other than a high concentration of nicotine and its primary metabolite cotinine in femoral venous blood (2.2 microg/mL). Blood alcohol was determined to be 2.1 g/L in femoral venous blood. Only a paucity of fatal cases of nicotine poisoning has been reported in the literature so far.

Topics: Adult; Analgesics; Central Nervous System Depressants; Cotinine; Ethanol; Forensic Toxicology; Ganglionic Stimulants; Gas Chromatography-Mass Spectrometry; Gastrointestinal Contents; Humans; Male; Nicotine; Strychnine

Alcohol Denat. is the generic term used by the cosmetics industry to describe denatured alcohol. Alcohol Denat. and various specially denatured (SD) alcohols are used as cosmetic ingredients in a wide variety of products. Many denaturants have been previously considered, on an individual basis, as cosmetic ingredients by the Cosmetic Ingredient Review (CIR) Expert Panel, whereas others, including Brucine and Brucine Sulfate, Denatonium Benzoate, and Quassin, have not previously been evaluated. Quassin is a bitter alkaloid obtained from the wood of Quassia amara. Quassin has been used as an insect antifeedant and insecticide and several studies demonstrate its effectiveness. At oral doses up to 1000 mg/kg using rats, Quassin was not toxic in acute and short-term tests, but some reversible piloerection, decrease in motor activity, and a partial loss of righting reflex were found in mice at 500 mg/kg. At 1000 mg/kg given intraperitoneally (i.p.), all mice died within 24 h of receiving treatment. In a cytotoxicity test with brine shrimp, 1 mg/ml of Quassin did not possess any cytotoxic or antiplasmodial activity. Quassin administered to rat Leydig cells in vitro at concentrations of 5-25 ng/ml inhibited both the basal and luteinizing hormone (LH)-stimulated testosterone secretion in a dose-related fashion. Quassin at doses up to 2.0 g/kg in drinking water using rats produced no significant effect on the body weights, but the mean weights of the testes, seminal vesicles, and epididymides were significantly reduced, and the weights of the anterior pituitary glands were significantly increased. The sperm counts and levels of LH, follicle-stimulating hormone (FSH), and testosterone were significantly lower in groups treated with Quassin. Brucine is a derivative of 2-hydroxystrychnine. Swiss-Webster mice given Brucine base, 30 ml/kg, had an acute oral LD(50) of 150 mg/kg, with central nervous system depression followed by convulsions and seizures in some cases. In those animals that died, respiratory arrest was the cause. The acute i.p. LD(50) for 15 ml/kg of Brucine base was 62.0 mg/kg, with central nervous system depression prior to the onset of convulsions, just as with oral Brucine. The acute intravenous (i.v.) LD(50) was 12.0 mg/kg. Brucine was nonmutagenic in an Ames assay at levels up to 6666 mu g/plate, with and without metabolic activation. In a repeat-insult patch test, for a hair care product containing 47% SD Alcohol 40 (95%), it was reported that Bruc

Topics: Adjuvants, Pharmaceutic; Alcohols; Animals; Consumer Product Safety; Cosmetics; Dose-Response Relationship, Drug; Humans; Quassins; Quaternary Ammonium Compounds; Risk Assessment; Strychnine; Toxicity Tests

The ternary allosteric model predicts the possibility of discovering molecules with novel and highly subtype-selective modes of action. This approach has been applied to muscarinic receptors. The alkaloid brucine is capable of selectively enhancing by an allosteric mechanism the effects of low but not high concentrations of acetylcholine at only the m1 subtype of muscarinic receptors. A simple derivative of brucine, N-chloromethylbrucine, enhances acetylcholine actions selectively at only m3 receptors. In addition it binds to, but does not affect, the properties of m4 receptors, thereby demonstrating neutral cooperativity and an 'absolute' selectivity of action at m3 receptors over m4 receptors. Brucine N-oxide enhances acetylcholine binding at m3 and m4 receptors and is neutral at m1 and m5 receptors. These findings allow the possibility of developing muscarinic agents that have a novel and highly targeted mode of action; they may act only on a single muscarinic receptor subtype which is functioning sub-optimally and therefore be of use therapeutically in the early stages of Alzheimer's Disease.

Topics: Acetylcholine; Allosteric Regulation; Animals; Humans; Receptor, Muscarinic M1; Receptor, Muscarinic M3; Receptor, Muscarinic M4; Receptors, Muscarinic; Strychnine

To assess the safety of individual medication of Guo's Ma Qian Decoction on the basis of effective treatment of fluorosis of bone with Guo's therapy.. One hundred and fourteen cases of moderate fluorosis of bone were randomly divided into a treatment group (n = 60) and a control group (n = 54) between December 2007 and August 2009 by using the block randomized method and a central random system. At the same time of basic treatment, the patients in the treatment group were orally administrated with Guo's Ma Qian Decoction. The initial dose of Ma Qian Zi (Semen Strychni) was 0.4 g and increased by 0.05 g every two days, with the doses of other drugs unchanged, until the patient had "nux vomica response". For the patients with no "nux vomica response", the dosage was continued to increase and the maximum dosage was not more than 1.2 g/day. The control group was treated with decoction placebo. The changes of strychnine and brucine contents before and after processing and after decoction of Ma Qian Zi (Semen Strychni) were determined with reversed-phase high-performance liquid chromatography, which were controlled within ranges stipulated in the Pharmacopeia; Adverse events were analyzed; Blood strychnine and brucine contents in 10 cases who had taken the drugs were determined.. 1) Strychnine (2.125%) and brucine (1.425%) contents before processing of Ma Qian Zi and 1.88% and 1.31% after processing all conformed with the standards of strychnine (1.2-2.2%) and brucine (no less than 0.8%) stipulated in the Pharmacopeia. When the maximum dosage of Ma Qian Zi was 1.2 g/day, strychnine in the decoction was 11.17 mg and brucine was 7.44 mg, which all conformed with the maximum limited amount (strychnine 13.32 and brucine no less than 4.8 mg) stipulated in the Pharmacopeia. 2) Eight cases had "nux vomica response" in the treatment group and one case in the control group, with a significant difference between the two groups (P < 0.05). 3) Altogether 18 cases had adverse events, with an incidence rate of 15.38% (8 cases) in the treatment group and 18.52% (10 cases) in the control group, with no difference between the two groups (P > 0.05); Among them, 10 cases (8.77%) with the adverse event were not related with therapeutic drugs, with an incidence rate of 6.67% (4 cases) in the treatment and 11.11% (6 cases) in the control group, with no significant difference between the two groups (P > 0.05). Seven cases had suspicious relative adverse events, the risk in the treatment group was 0.658 times of the control group, with no significant difference (P > 0.05), and one case had the toxic reaction of nux-vomica seed. 4) Strychnine and brucine were unable to be detected in the blood in all points of time in the 10 cases who had taken the drugs, indicating that plasma strychnine and brucine contents were lower than the minimum detectable amount (10 ng), and accumulation of strychnine and brucine were not found in blood of the patient during and after administration for 8 weeks.. The individual medication of Ma Qian Zi (Semen Strychni) in the Guo's therapy has a better safety.

Topics: Adult; Drugs, Chinese Herbal; Female; Fluorosis, Dental; Humans; Male; Middle Aged; Strychnine

Brucine (BRU) and brucine N-oxide (BNO) are prominent, bioactive, and toxic alkaloids in crude and processed Semen Strychni. Studies have demonstrated that BRU and BNO possess comprehensive pharmacological activities, such as anti-inflammatory and analgesic. In this context, a comparative study of BRU and BNO was performed by combination analysis of in silico ADMET prediction, in vivo toxicity evaluation, and potential action mechanism exploration. ADMET prediction showed that BRU and BNO might induce liver injury, and BRU may have a stronger hepatoxic effect. The prediction was experimentally verified using the zebrafish model. The BRU-induced hepatotoxicity of zebrafish larvae had a dose-response relationship. The mechanism of BRU-induced hepatotoxicity might relate to phosphorylation, kinase activity, and signal transduction. By comparison, signal transduction and gap junctions might involve BNO-induced hepatotoxicity. Our results provided a better understanding of BRU- and BNO-induced hepatotoxicity. We also built a foundation to elucidate the material base of the hepatotoxicity of traditional Chinese medicine Semen Strychni.

Topics: Animals; Chemical and Drug Induced Liver Injury; Drugs, Chinese Herbal; Strychnine; Zebrafish

Strychnine is a natural product that, through isolation, structural elucidation and synthetic efforts, shaped the field of organic chemistry. Currently, strychnine is used as a pesticide to control rodents

Topics: Biosynthetic Pathways; Metabolic Engineering; Nicotiana; Strychnine

Assessment of neural activity in the specific brain area is critical for understanding the circuit mechanisms underlying altered brain function and behaviors. A number of immediate early genes (IEGs) that are rapidly transcribed in neuronal cells in response to synaptic activity have been used as markers for neuronal activity. However, protein detection of IEGs requires translation, and the amount of newly synthesized gene product is usually insufficient to detect using western blotting, limiting their utility in western blot analysis of brain tissues for comparison of basal activity between control and genetically modified animals. Here, we show that the phosphorylation status of eukaryotic elongation factor-2 (eEF2) rapidly changes in response to synaptic and neural activities. Intraperitoneal injections of the GABA A receptor (GABA

Topics: Animals; Brain; CA3 Region, Hippocampal; Eukaryotic Initiation Factor-2; Genes, Reporter; Mice; Muscimol; Nerve Tissue Proteins; Phosphorylation; Picrotoxin; Prosencephalon; Protein Processing, Post-Translational; Pyramidal Cells; Quinoxalines; Restraint, Physical; Stress, Physiological; Strychnine

Brucine, one of the natural medications obtained from Nux vomica seeds, is used as an anti-inflammatory drug. Several investigations were performed to overcome its drawbacks, which will affect significantly its pharmaceutical formulation. The goal of the current investigation was to design, optimize, and evaluate the anti-inflammatory performance of BRU ethosomal gel. Brucineethosomal formulations were prepared using thin film hydration method and optimized by central composite design approach using three independent variables (lecithin concentration, cholesterol concentration, and ethanol percentage) and three response variables (vesicular size, encapsulation efficiency, and skin permeation). The optimized formulation was examined for its stability and then incorporated into HPMC gel to get BRU ethosomal gel. The obtained BRU-loaded ethosomal gel was evaluated for its physical properties, in vitro release, and ex vivo permeation and skin irritation. Finally, carrageenan-induced rat hind paw edema test was adopted for the anti-inflammatory activity. The developed BRU ethosomal gel exhibited good physical characteristics comparable with the conventional developed BRU gel. In vitro release of BRU from ethosomal gel was effectively extended for 6 h. Permeation of BRU from ethosomes was significantly higher than all formulations (p < 0.05), since it recorded steady state transdermal flux value 0.548 ± 0.03 μg/cm

Topics: Administration, Cutaneous; Animals; Anti-Inflammatory Agents; Lecithins; Liposomes; Prospective Studies; Rats; Skin; Skin Absorption; Strychnine

Myocardial infarction (MI) is the most extensive manifestations of cardiovascular disease (CVD), associated with prolonged supply and demand blood oxygen imbalance to the heart muscle. The treatment of MI includes several conventional medicines which are beta-blockers and calcium antagonists. Though, these were reported to be either not efficient or associated with life threatening adverse effects. Brucine, the main alkaloid bioactive compound from Strychnos nux-vomica seeds, offers unique compatibility advantages in inflammatory diseases associated clinical practices. Thus, the present investigation was projected to explore the activity of brucine towards MI provoked by isoprenaline hydrochloride (ISO) in rats. The cardioprotective properties of brucine were evaluated via detecting the infarct size, serum cardiac marker enzymes (CK, CK-MB, cTnT, and cTnI), endogenous antioxidants (CAT, SOD, GPx), and lipid peroxidation (TBARS and LOOH), inflammatory mediators (NF-κB, TNF-α and IL-6) and histopathological analysis. The results demonstrated, brucine effectively restored the infarct size by increasing the endogenous antioxidants and decreasing the status of TBARS and LOOH, marker enzymes and ameliorated the histopathological injuries. Brucine's cardioprotective effect might be associated with TNF-α, IL-6 signaling molecules activation, revealing its pharmacological actions.

Topics: Animals; Disease Models, Animal; Isoproterenol; Myocardium; Rats; Strychnine

We aimed to determine the circadian responses of mice to Semen Strychni and to investigate the role of pharmacokinetics in generating chronotoxicity.. Total extract of Semen Strychni was administered by oral gavage to wild-type (WT) and Bmal1-/- (a circadian clock-deficient model) mice at different circadian time points for toxicity (including survival) and pharmacokinetic characterization. Nephrotoxicity and neurotoxicity were evaluated by measuring plasma creatinine and creatine kinase BB (CK-BB), respectively. Drug metabolism and transport assays were performed using liver/intestine microsomes and everted gut sacs, respectively.. Semen Strychni nephrotoxicity and neurotoxicity as well as animal survival displayed significant circadian rhythms (the highest level of toxicity was observed at ZT18 and the lowest level at ZT2 to ZT6). According to pharmacokinetic experiments, herb dosing at ZT18 generated higher plasma concentrations (and systemic exposure) of strychnine and brucine (two toxic constituents) compared with ZT6 dosing. This was accompanied by reduced formation of both dihydroxystrychnine and strychnine glucuronide (two strychnine metabolites) at ZT18. Bmal1 ablation sensitized mice to Semen Strychni-induced toxicity (with increased levels of plasma creatinine and CK-BB) and abolished the time dependency of toxicity. Metabolism of Semen Strychni (strychnine and brucine) in the liver and intestine microsomes of WT mice was more extensive at ZT6 than at ZT18. These time differences in hepatic and intestinal metabolism were lost in Bmal1-/- mice. Additionally, the intestinal efflux transport of Semen Strychni (strychnine and brucine) was more extensive at ZT6 than ZT18 in WT mice. However, the time-varying transport difference was abolished in Bmal1-/- mice.. Circadian responses of mice to Semen Strychni are associated with time-varying efflux transport and metabolism regulated by the circadian clock (Bmal1). Our findings may have implications for optimizing phytotherapy with Semen Strychni via timed delivery.

Topics: Animals; ARNTL Transcription Factors; Biological Transport; Circadian Clocks; Circadian Rhythm; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Microsomes; Neurotoxicity Syndromes; Plant Extracts; Strychnine; Strychnos nux-vomica; Time Factors

Brucine (BRU) is a natural product derived from nux-vomica seeds. It is commonly used as an anti-inflammatory and anti-nociceptive drug to relieve arthritis and traumatic pain. Nevertheless, its use is significantly limited by its low aqueous solubility, as well as the gastrointestinal problems and systemic toxicity that may occur following oral administration. The goal of this study, therefore, was to formulate and evaluate a nanoemulgel formulation of BRU for enhanced topical anti-inflammatory and anti-nociceptive activities. Different formulations were developed (BRU gel, emulgel and nanoemulgel) using 1% w/w NaCMC as a gelling agent. The formulated preparations were assessed for their physical appearance, spreadability, viscosity, particle size, in vitro drug release and ex vivo permeation studies. In addition, the carrageenan-induced rat hind paw edema method was adopted to scrutinize the anti-inflammatory activity, while the hot plate method and acetic acid-induced writhing test were used to assess the anti-nociceptive activity of different formulations in male BALB/c mice. The formulated BRU-loaded preparations showed good physical characteristics. Cumulative drug release from BRU-loaded nanoemulgel was remarkably higher than that of the other formulations. Ex vivo drug permeation of the nanoemulgel formulation across rat skin showed enhanced drug permeation and higher transdermal flux as compared to BRU-loaded gel or emulgel. Most importantly, the carrageenan-induced rat hind paw edema model verified the efficient anti-inflammatory potential of BRU-loaded nanoemulgel. In addition, BRU-loaded nanoemulgel exhibited significant protective effects against thermal stimulation in the hot plate test and remarkably inhibited acetic acid-induced abdominal writhing in mice. Furthermore, a skin irritation test indicated that BRU-loaded nanoemulgel elicited neither edema nor erythema upon application to rat skin. Collectively, our results suggest that myrrh oil-based nanoemulgel might represent a promising delivery vehicle for potentiating the anti-inflammatory and anti-nociceptive actions of brucine.

Topics: Administration, Cutaneous; Animals; Anti-Inflammatory Agents; Edema; Male; Mice; Mice, Inbred BALB C; Rats; Skin Absorption; Strychnine

Brucine are the main constituents of Strychnos nux-vomica. Earlier reports have determined brucine shows anti-inflammatory, analgesic and excellent anti-tumor drug. Even though its anticervical cancer cells remains not clearly evaluated. So that, we hypothesized the anti-cervical cancer activity of brucine against the cervical (ME-180) cells. Brucine inhibited the inflammation, cell proliferation and promoted rate of apoptotic cell death ad reduced the mitochondrial potential, which is evidenced by respective (AO/EB, Rh-123, and PI) staining. Furthermore ELISA and real time PCR reaction determined that brucine were down regulated inflammatory (TNF-α, NF-kB, IL-6 & COX-2) cell proliferation (Cyclin D1) and apoptotic marker Bax, caspase-3, PI3K (phosphoinosital 3 kinase), AKT, mTOR (mammalian target of rapamycin) and over expression Bcl-2, associated death promoter. These findings were confirmed and finally suggested that brucine inhibited inflammation, cell proliferation and promoted the apoptosis through the down-regulation of PI3K/AKT/mTOR pathway. Taken together, these data were exhibited brucine as a good therapeutic agents for the prevention of anticancer cervical cancer drugs.

Topics: Apoptosis; Cell Line, Tumor; Cell Proliferation; Female; Humans; Inflammation; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Signal Transduction; Strychnine; TOR Serine-Threonine Kinases; Uterine Cervical Neoplasms

Ferroptotic cell death is characterized by iron-dependent lipid peroxidation that is initiated by ferrous iron and H

Topics: Activating Transcription Factor 3; Amino Acid Transport System y+; Animals; Antineoplastic Agents; Catalase; Cell Line, Tumor; Endoplasmic Reticulum Stress; Ferroptosis; Humans; Hydrogen Peroxide; Iron; Mice, Inbred BALB C; Mice, Nude; NADPH Oxidase 4; Neoplasms; Strychnine; Superoxide Dismutase-1; Up-Regulation; Xenograft Model Antitumor Assays

Natural products have been extensively used for treating a wide variety of disorders. In recent times, Brucine (BRU) as one of the natural medications extracted from seeds of nux vomica, was investigated for its anticancer activity. As far as we know, this is the first study on BRU anticancer activity against skin cancer. Thus, the rational of this work was implemented to develop, optimize and characterize the anticancer activity of BRU loaded ethosomal gel. Basically, thin film hydration method was used to formulate BRU ethosomal preparations, by means of Central composite design (CCD), which were operated to construct (3

Topics: Administration, Cutaneous; Animals; Ethanol; Gels; Male; Phospholipids; Rats; Rats, Wistar; Skin; Skin Absorption; Skin Neoplasms; Strychnine

A simple, rapid, cost-effective and green analytical method is developed based on ultrasound-assisted dispersive liquid-liquid microextraction (US-DLLME) coupled to thin-layer chromatography (TLC)-image analysis for the simultaneous determination of two major alkaloids of Strychnos nux-vomica L i.e., strychnine and brucine. The method is composed of three steps, namely (i) US-DLLME by injecting a mixture of 100-μL chloroform (extraction solvent) and 1-mL methanol (disperser solvent) in 5 mL of aqueous sample, followed by ultrasonication and centrifugation, (ii) TLC of 20 μL of sedimented phase with methanol: ammonia (100:1.5, v/v) as the mobile phase and visualization under ultraviolet radiation (254 nm) and (iii) photography of TLC plate and quantification of spots by image analysis using freely available imageJ software (National Institute of Health, Bethesda, MD, USA). The limit of detection and limit of quantification for both alkaloids were found to be in the range of 0.12-0.15 and 0.36-0.48 μg/spot, respectively. The method was found to be linear in the range of 0.5-5 μg/spot with correlation coefficient (R2) of 0.995 and 0.997 for strychnine and brucine, respectively. The developed method was successfully applied for the determination of strychnine and brucine in Ayurvedic formulations and blood samples. The method does not require any sophisticated instrument and handling skills and can be adopted for rapid analysis of strychnine and brucine in forensic toxicological laboratories.

Topics: Chromatography, Thin Layer; Cost-Benefit Analysis; Humans; Image Processing, Computer-Assisted; Limit of Detection; Liquid Phase Microextraction; Medicine, Ayurvedic; Reproducibility of Results; Strychnine; Strychnos nux-vomica; Tablets; Ultrasonics; Ultraviolet Rays

To investigate the antagonistic effect of the extract of Baizhu (Rhizoma Atractylodis Macrocephalae) (RAM) on the intestinal absorption of brucine and strychnine in Strychnos nux-vomica (NUX) and propose the mechanism of these effects.. The apparent permeability value (Papp) and absorption rate constant (Ka) were chosen as indices. The everted intestinal sac model and in situ single-pass intestinal perfusion model were used to study the effects of the RAM extract on the absorption of brucine and strychnine. To confirm the results, the brucine and strychnine concentrations in hepatic portal venous blood were determined. Western blotting was used to study P-glycoprotein (P-gp) expression in the Caco-2 cell line.. Papp and Ka of brucine and strychnine were significantly increased in the presence of a P-gp inhibitor, but no significant increase was noted in the presence of a tight junction regulator. The RAM extract inhibited the absorption of brucine and strychnine and enhanced P-gp expression.. The primary absorption mechanism for brucine and strychnine is passive transport, which is affected by P-gp.

Topics: Animals; Atractylodes; Caco-2 Cells; Cell Line, Tumor; Drugs, Chinese Herbal; Humans; Intestinal Absorption; Male; Rats; Rats, Sprague-Dawley; Rhizome; Strychnine; Strychnos nux-vomica

Autophagy is an important tightly controlled cellular process that regulates cellular homeostasis and is involved in deciding cell fate such as cell survival and death. The role of autophagy in many intracellular signaling pathways explains its interaction with other different types of cell death, including apoptosis and immunogenic cell death (ICD). The reports showed the complex and intriguing relationship existing between autophagy and immune system signaling pathways. However, the role of autophagy in ICD remains to be clearly elucidated. In this study, we demonstrated that Brucine, a clinically-used small molecule in traditional Chinese medicine, elicited autophagy inhibition. Brucine also triggered cell stress and induced features of ICD, including calreticulin (CRT) exposure and high-mobility group box 1 (HMGB1) release in MDA-MB-231 and CT26 cancer cells. Brucine impaired autolysosomal degradation and exerted a feedback regulation of ERK1/2-mTOR-p70S6K signaling cascade. Brucine-elicited ICD was confirmed by the rejection of CT26 tumor cells, implanted in the mice after vaccination with Brucine-treated CT26 cells. The impaired autophagy contributed to Brucine-induced ICD, as knock-down of Atg5 significantly reduced Brucine-elicited CRT exposure and HMGB1 release. Our results revealed Brucine as a novel autophagy regulator, ICD inducer and hitherto undocumented role of autophagy in ICD. Thus, these results imply the importance of Brucine in cancer immunotherapy. Therefore, Brucine may be used as an ICD inducer and improve its application in cancer treatment with minimized toxicity.

Topics: Animals; Autophagy; Autophagy-Related Protein 5; Calreticulin; Cell Death; Cell Line, Tumor; Drugs, Chinese Herbal; Gene Knockdown Techniques; HMGB1 Protein; Humans; Immunotherapy; Lysosomes; MAP Kinase Signaling System; Mice; Neoplasms; Phytotherapy; Strychnine

The detoxification effects of licorice are believed to be related to its pharmacokinetic (PK) interference. This paper aimed to evaluate the effects of licorice water extracts (LWE) on the pharmacokinetics of brucine. Rats were administered brucine and/or LWE. The pharmacokinetic behavior of brucine and bioactive components of licorice were quantified by HPLC-MS/MS. P-glycoprotein (P-gp) inhibitor verapamil, real time PCR, vesicular transport assay and everted gut sacs were employed to investigate its possible mechanism. We found LWE reduced the Cmax and AUC of oral brucine in a dose-dependent way. In contrast, the AUC values of intraperitoneal brucine showed no significant difference between LWE treated and untreated rats, which indicating the intestinal absorption of brucine was influenced by LWE. We found that high dose of LWE activated the transport activity of P-gp in vesicular transport assay, while the mRNA level of P-gp in the intestinal was not affected by licorice. Moreover, high dose of LWE decreased the intestinal absorption of brucine in the everted gut sacs model, which could over turned by verapamil. These results suggested that a single high dose of LWE could impair the intestine absorption of brucine, and its potential mechanism may be mediated by P-gp in intestine.

Topics: Administration, Oral; Animals; ATP Binding Cassette Transporter, Subfamily B, Member 1; Glycyrrhiza; Herb-Drug Interactions; Injections, Intraperitoneal; Intestinal Absorption; Intestinal Mucosa; Male; Plant Extracts; Rats, Sprague-Dawley; Strychnine

Licorice, one of the most widely used medicinal herbs in East Asia, has effects such as anti-inflammation, antioxidant, and detoxifying. This study aimed to evaluate the protective effect of licorice on brucine-induced nephrotoxicity. Sprague Dawley rats were administered with brucine intraperitoneally for 7 consecutive days with or without treatment with licorice. The content of blood urea nitrogen and creatinine in serum, the activities of superoxide dismutase and content of glutathione, malonaldehyde in kidney tissue were detected. Hematoxylin-eosin staining was employed to observe the histopathological changes of kidney. The expression and phosphorylation levels of protein were evaluated by Western blotting and immunohistochemical analysis. The results illustrated that treatment with licorice extracts (LE) significantly protected against the brucine-induced nephrotoxicity by reducing the content of blood urea nitrogen and serum creatinine, attenuating pathologic damage. The unbalance of oxidative stress was repaired by LE via increasing the level of glutathione, promoting the activities of superoxide dismutase and decreasing the content of malonaldehyde. In addition, LE overturned the influence of brucine on apoptosis-related protein and signal transducer and activator of transcription-3 (STAT3) activation. Taken together, these data demonstrate that licorice may attenuate brucine-induced nephrotoxicity via inactivation of oxidative stress and mitochondrial-mediated apoptosis pathway. More importantly, the renoprotective effects may be mediated, at least partly, by preventing the activation of STAT3 protein.

Topics: Acute Kidney Injury; Animals; Apoptosis; Blood Urea Nitrogen; Creatinine; Disease Models, Animal; Gene Expression Regulation; Glutathione; Glycyrrhiza; Injections, Intraperitoneal; Male; Malondialdehyde; Mitochondria; Plant Extracts; Random Allocation; Rats; Rats, Sprague-Dawley; STAT3 Transcription Factor; Strychnine; Superoxide Dismutase

The present study was aimed to investigate the dose dependent chemopreventive activity of brucine against 7, 12-dimethylbenz (a) anthracene induced mammary gland tumorigenesis in rats. The mammary tumor was induced by a single dose of DMBA (25 mg/rat) injected subcutaneously near the mammary gland. We observed reduced body weight and increase in tumor incidence, the total number of tumors, and tumor volume in DMBA alone injected rats and also observed decreased antioxidant status (SOD, CAT, GPX, and GSH) and increased lipid peroxidation (TBARS and LOOH) in plasma and mammary tissues. Increased levels of CYP450, Cyt-b5 and decreased levels of phase II (GST and GR) biotransformation enzymes were noticed in the liver and mammary tissues. Further, increased levels of lipid profile (TC, TG, PL, and FFA) and lipoprotein (LDL and VLDL) were noticed. Whereas, decrease in the levels of HDL in plasma and decreased levels of PL and FFA in mammary tissues were observed. Oral administration of brucine in different doses (2, 4 and 8 mg/kg bw) inhibited the tumor incidence and restored the levels of biochemical markers near to normal in dose responsive manner. Biochemical findings are supported by histopathological studies. The results suggest that brucine at a dose of 8 mg/kg bw shows more significant chemopreventive activity in DMBA-induced mammary carcinogenesis.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Anticarcinogenic Agents; Antioxidants; Chemoprevention; Dose-Response Relationship, Drug; Female; Lipid Metabolism; Mammary Glands, Animal; Mammary Neoplasms, Experimental; Oxidative Stress; Rats, Sprague-Dawley; Strychnine

Strychnos nux-vomica L. (Loganiaceae) is grown extensively in South Asian. The dried seed of this plant, nux vomica, has been clinically used in Chinese medicine for relieving rheumatic pain, reducing swelling and treating cancer. Brucine, the second abundant alkaloid constituent of nux vomica, shows excellent clinical therapeutic effect, especially in relieving pain, but mechanism of brucine in relieving pain is still unclear.. Explore the analgesic effect of brucine, reveal the molecular mechanism of brucine analgesia.. Antinociceptive effects of brucine were assessed in acute and chronic pain mice model. Electrophysiological experiments were used to evaluate the effects of brucine on neuronal activity and sodium channel function.. In acute pain models, brucine significantly inhibits response induced by nociceptive heat and mechanical stimulation. Furthermore, thermal hypersensitivity and mechanical allodynia were also alleviated by brucine treatment in a chronic constriction injury (CCI) mouse model. Sodium channel plays a crucial role in neuropathic pain. Electrophysiological results show that brucine inhibits the excitability of DRG neurons directly, the number of action potential (AP) was significantly reduced after brucine treatment, and this kind of inhibition is due to brucine inhibits both tetrodotoxin-sensitive (TTXs) and tetrodotoxin-resistant (TTXr) sodium channel.. Taken together, brucine is a novel drug candidate in treating acute and chronic pain diseases, which might be attributed to inhibition the excitability of sodium channel directly.

Topics: Action Potentials; Analgesics; Animals; Behavior, Animal; Cells, Cultured; Ganglia, Spinal; Male; Mice, Inbred C57BL; Neuralgia; Neurons; Sodium Channels; Strychnine

Vasculogenic mimicry (VM) with the pattern of endothelial independent tubular structure formation lined by aggressive tumor cells mimics regular tumor blood vessels to ensure robust blood supply and correlates with the proliferation, invasion, metastasis, and poor prognosis of malignant tumors, which was demonstrated to be a major obstacle for resistance to antiangiogenesis therapy. Therefore, it is urgent to discover methods to abrogate the VM formation of tumors, which possesses important practical significance for improving tumor therapy. Brucine is a traditional medicinal herb extracted from seeds of Strychnos nux-vomica L. (Loganiaceae) exhibiting antitumor activity in a variety of cancer models. In the present study, the effect of brucine on vasculogenic mimicry and the related mechanism are to be investigated. We demonstrated that, in a triple-negative breast cancer cell line MDA-MB-231, brucine induced a dose-dependent inhibitory effect on cell proliferation along with apoptosis induction at higher concentrations. The further study showed that brucine inhibited cell migration and invasion with a dose-dependent manner. Our results for the first time indicated that brucine could disrupt F-actin cytoskeleton and microtubule structure, thereby impairing hallmarks of aggressive tumors, like migration, invasion, and holding a possibility of suppressing vasculogenic mimicry. Hence, the inhibitory effect of brucine on vasculogenic mimicry was further verified. The results illustrated that brucine significantly suppressed vasculogenic mimicry tube formation with a dose-dependent effect indicated by the change of the number of tubules, intersections, and mean length of tubules. The in-depth molecular mechanism of vasculogenic mimicry suppression induced by brucine was finally suggested. It was demonstrated that brucine inhibited vasculogenic mimicry which might be through the downregulation of erythropoietin-producing hepatocellular carcinoma-A2 and matrix metalloproteinase-2 and metalloproteinase-9.

Topics: Apoptosis; Cell Line, Tumor; Cell Movement; Cell Proliferation; Female; Gene Expression Regulation, Neoplastic; Humans; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Neoplasm Invasiveness; Neovascularization, Pathologic; Strychnine; Strychnos nux-vomica; Triple Negative Breast Neoplasms

Brucine and Strychnine are alkaloids isolated from the seeds of Strychnos nux vomica L., which have long been used as a traditional medicine for the treatment of tumor. However, the effect of Brucine and Strychnine on colorectal cancer (CRC) and the underlying molecular mechanism remain unclear. In the present study, Brucine and Strychnine displayed profound inhibitory effects on the growth of human colon cancer cells. The results of flow cytometric analysis demonstrated that the two alkaloids induced cellular apoptosis. Moreover, the growth of DLD1 xenografted tumors in nude mice was significantly suppressed in the Brucine or Strychnine treated group. Mechanistically, the Wnt/β-catenin is involved in this phenomenon, which is characterized by significantly increased expression of DKK1 and APC, whereas decreased expression of β-catenin, c-Myc, and p-LRP6 in CRC cells as well as tumor tissues. Collectively, Brucine and Strychnine have targeted inhibition for colon cancer proliferation both in vitro and in vivo, and it is valuable for future exploitation and utilization as an antitumor agent of CRC.

Topics: Alkaloids; Animals; Colonic Neoplasms; Humans; Mice; Mice, Nude; Strychnine; Strychnos nux-vomica; Wnt Signaling Pathway

Brucine is one of the main bioactive and toxic constituents of the herb drug Semen Strychni. Here we aimed to determine dosing time-dependent hepatotoxicity of brucine, and to investigate the role of metabolism in generation of brucine chronotoxicity. Brucine was administered to wild-type or Npas2

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Basic Helix-Loop-Helix Transcription Factors; Chemical and Drug Induced Liver Injury; Circadian Rhythm; Cytochrome P-450 CYP3A; Drug Chronotherapy; HEK293 Cells; Humans; Liver; Male; Membrane Proteins; Mice, Inbred C57BL; Mice, Knockout; Microsomes, Liver; Nerve Tissue Proteins; Photoperiod; Strychnine

Semen Strychni is known for its treatment of rheumatic arthritis with a low therapeutic index. Liquorice contributes a lot in herb detoxification according to the traditional Chinese medicine theory. A simple, rapid, and sensitive liquid chromatography-mass spectrometric method (LC-MS) was developed and validated for simultaneous determination of main bioactive ingredients in liquorice and Semen Strychni in rat plasma. Using moclobemide and cyproterone acetate as the internal standards, the analytes were pretreated via protein precipitation with methanol. An Ultimate AQ-C18 column (3.0 μm, 3.0 × 100 mm) was employed for chromatographic separation, combining with gradient elution. The mobile phase consisted of 0.07% formic acid and 0.12% ammonium acetate in aqueous phase (A) and acetonitrile in organic phase (B). The elution program was as follows: 0-0.5 min, 20% B; 0.5-1 min, 20-60% B; 1-7 min, 60-85% B; and 7-7.5 min, returned to 20% B, then continued to 12 min. Selected reaction monitoring was performed in both positive and negative ESI. Positive mode was adopted for detection of strychnine, brucine, and moclobemide, while negative mode was used for glycyrrhizic acid, glycyrrhetinic acid, liquiritigenin, isoliquiritigenin, liquiritin, and cyproterone acetate. The method was validated for specificity, linearity, matrix effect, recovery, precision, accuracy, and stability. The results show that this method is sensitive, accurate and robust for biological matrix analysis. Moreover, the proposed method was applied to a pharmacokinetic study in Sprague-Dawley rats for investigating the mechanism of which liquorice detoxifies Semen Strychni.

Topics: Animals; Chromatography, Liquid; Flavanones; Glucosides; Glycyrrhiza; Glycyrrhizic Acid; Plasma; Rats; Reproducibility of Results; Semen; Strychnine

To examine the effect of brucine on the migration, invasion, adhesion and expressions of epithelial-to-mesenchymal transition (EMT) markers and matrix metalloproteinases (MMPs) in the highly metastatic breast cancer cell lines MDA-MB-231 and Hs578-T.. MDA-MB-231 and Hs578-T cells were divided to 4 groups: the control group (0.1% DMSO), and 25, 50 and 100 μmol/L brucine groups. The cell viability was determined using a CellTiter-Glo® luminescent cell viability. The scratch wound healing assay and tanswell migration assay were used to determine the migration ability of these cells treated by different concentrations of brucine. The proliferation rate, invasive potential and adhesive ability were respectively performed by colony formation assay, transwell invasion assay and adhension assay. The protein and mRNA expressions of EMT biomarkers, MMP-2 and MMP-9 were investigated by real-time reverse transcription polymerase chain reaction and Western blot.. Compared with the control group, brucine had little effect on cell viability or proliferation (P>0.05), but led to a dose-dependent decrease on migration, invasion, adhension of MDA-MB-231 and Hs578-T cells (P<0.01). Furthermore, brucine increased the protein and mRNA levels of EMT markers such as E-cadherin and β-catenin in MDA-MB-231 and Hs578-T cells, and decreased the protein and mRNA levels of mesenychmal markers such as vimentin and fibronectin, as well as the expressions of MMP-2 and MMP-9 (all P<0.01).. Brucine inhibited triple negative breast cancer cells metastasis potentially through EMT reversion and MMP-2 and MMP-9 inhibition.

Topics: Biomarkers, Tumor; Breast Neoplasms; Cell Adhesion; Cell Line, Tumor; Cell Movement; Cell Survival; Collagen; Drug Combinations; Epithelial-Mesenchymal Transition; Female; Humans; Laminin; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Neoplasm Invasiveness; Neoplasm Proteins; Proteoglycans; RNA, Messenger; Strychnine

Topics: Administration, Oral; Animals; Chromatography, Liquid; Drugs, Chinese Herbal; Female; Linear Models; Male; Rats; Rats, Sprague-Dawley; Reproducibility of Results; Sensitivity and Specificity; Strychnine; Tandem Mass Spectrometry

An enhanced cleanup efficiency hydroxy functionalized-magnetic graphene oxide (EH-Mag-GO) fully covered porous nano-titania as coating has been designed and synthesized. It has been evaluated in PRiME (process, robustness, improvements, matrix effects, ease of use) pass-through cleanup procedure for human plasma prior to analysis of strychnine and brucine by liquid chromatography-tandem quadrupole mass spectrometry (LC-MS/MS). Comparing with the magnetic carboxyl-graphene (Mag-CG), EH-Mag-GO is much more effective for the removal of matrix effect resulted from blood phospholipids. Under optimal conditions, the results show higher cleanup efficiency of EH-Mag-GO with recoveries in the range of 89.4%-118%. The limits of quantification (LOQs) for strychnine and brucine are 0.088 μg/L and 0.092 μg/L, respectively. Especially, the EH-Mag-GO is also evaluated for reuse (20 times) without much sacrifice of the cleanup efficiency. Validation results on linearity, specificity, accuracy and precision, as well as on the application to analysis of strychnine and brucine in six cases of suspected semen strychni poisoning demonstrate the applicability to clinical studies.

Topics: Chromatography, Liquid; Graphite; Humans; Magnetic Phenomena; Molecular Conformation; Oxides; Particle Size; Strychnine; Surface Properties; Tandem Mass Spectrometry

Strychnos alkaloids (SAs) are the main toxic constituents in Semen Strychni, a traditional Chinese medicine, which is known for its fatal neurotoxicity. Hence, the present study was carried out to evaluate the neurotoxicity induced by SAs and the pre-protective effects of the total glucosides of Paeoniae Radix Alba (TGP). An SA brain damage model was firstly established. The neurotoxicity induced by SAs and the pre-protective effects of TGP were confirmed by physical and behavioral testing, biochemical assay, and histological examination. Then, a liquid chromatography-tandem mass spectrometry method was developed and validated to investigate the time-course change and distribution of strychnine and brucine (two main SAs) in the brain after oral SA administration with or without TGP pretreatment. Biochemical analysis results indicated that TGP could ameliorate the oxidative stress status caused by SAs. Time-course change and distribution studies demonstrated that strychnine and brucine were rapidly absorbed into the brain, peaked early at 0.5 h, and were mainly located in the hippocampus and cerebellum. TGP showed a pre-protective effect against neurotoxicity by reducing the absorption of toxic alkaloids into the brain. These findings could provide beneficial information in facilitating future studies of Semen Strychni neurotoxicity and developing herbal medicines to alleviate neurotoxicity in the clinic.

Topics: Administration, Oral; Animals; Antioxidants; Behavior, Animal; Brain; Chromatography, High Pressure Liquid; Glucosides; Male; Motor Activity; Neuroprotective Agents; Neurotoxicity Syndromes; Oxidative Stress; Paeonia; Permeability; Phytotherapy; Plant Extracts; Plants, Medicinal; Rats, Sprague-Dawley; Strychnine; Strychnos; Tandem Mass Spectrometry; Time Factors; Tissue Distribution

Bi qi capsule (BQC) is a traditional Chinese medicine prescription that is clinically used for the treatment of rheumatoid arthritis. Strychnine and brucine, as two typical kinds of alkaloids, are the primary active and neurotoxic constituents of BQC. In this study, a sensitive and reliable rapid resolution liquid chromatography-tandem mass spectrometry (RRLC-MS/MS) quantitative method was used to determine the concentrations of brucine and strychnine in rat brain and blood dialysates. The blood-brain barrier (BBB) penetration of free brucine and strychnine and their pharmacokinetic characteristics were investigated by the validated RRLC-MS/MS method coupled with in vivo microdialysis for the first time. The dialysate brain-blood AUC ratios of brucine were 0.098, 0.44 and 0.40 respectively at 0.4, 0.8 and 1.6 g kg

Topics: Animals; Brain Chemistry; Chromatography, Liquid; Drugs, Chinese Herbal; Linear Models; Male; Microdialysis; Rats; Rats, Sprague-Dawley; Reproducibility of Results; Sensitivity and Specificity; Strychnine; Tandem Mass Spectrometry

Colorectal cancer remains the third most common malignancies and migration is one of the main factors for its high mortality rate. Brucine, a natural plant alkaloid, has been proved to possess a variety of pharmacological functions including anti-tumor activities.. The aim of this study was to investigate the inhibitory effect of brucine on the colorectal cancer and the underlying mechanism.. In this study, colony formation assay and transwell assay were used to investigate the effect of brucine on LoVo cells viability and migration. Immunofluorescence assay, western blot assay and Gelatin zymography assay were used to study the mechanism of brucine. Xenograft model in nude mice was induced to investigate the in vivo effect of brucine on LoVo cells.. Brucine could significantly decrease the viability, inhibit the colony formation and induce the apoptosis of LoVo cells. Brucine could also suppress the migration of LoVo cells in a dose-dependent manner. Western blot analysis elucidated that the inhibition of migration was associated with the decreasing expression of matrix metalloproteinases including MMP2, MMP3 and MMP9. Moreover, we found that treatment of brucine could downregulate the expression of Frizzled-8, Wnt5a, APC and GSNK1A1, and increase the expression of AXIN1. Meanwhile, brucine also decreased the phosphorylation level of LRP5/6 and GSK3β, and increased the level of p-β-catenin. Xenografted model in nude mice study also revealed that oral administration of brucine could inhibit the growth and migration of LoVo cells by activating the expression of AXIN1 and p-β-catenin.. Brucine could suppress the migration of the colorectal cancer in vitro and in vivo and the effect was associated with the inhibition of the Wnt/β-catenin signaling pathway.

Topics: Animals; Apoptosis; Axin Protein; beta Catenin; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cell Survival; Colorectal Neoplasms; Down-Regulation; Glycogen Synthase Kinase 3 beta; Humans; Male; Matrix Metalloproteinases; Mice; Mice, Nude; Strychnine; Wnt Signaling Pathway; Xenograft Model Antitumor Assays

Nux vomica has been effectively used in Traditional Chinese Medicine. The processing of Nux vomica is necessary to reduce toxicity before it can be used in clinical practice. However, the mechanism for processing detoxification is unclear. hERG channels have been subjected to a routine test for compound cardiac toxicity in the drug development process. Therefore, we examined the effects and mechanisms of strychnine and brucine, two main ingredients of Nux vomica, and their N-oxides on hERG channels. Strychnine and brucine exhibited concentration-dependent inhibition of hERG channels with IC

Topics: Alkaloids; Cell Line; HEK293 Cells; Humans; Medicine, Chinese Traditional; Nitrogen; Oxygen; Potassium Channels; Sodium; Structure-Activity Relationship; Strychnine; Strychnos nux-vomica; Transcriptional Regulator ERG

The seeds of

Topics: Administration, Cutaneous; Analgesics; Animals; Anti-Inflammatory Agents; Arthritis, Experimental; Cell Line; Cell Proliferation; Cell Survival; Cells, Cultured; Dinoprostone; Disease Models, Animal; Drug Delivery Systems; Edema; Formaldehyde; Gels; Humans; Macrophages; Male; Mice; Pain; Phytotherapy; Rats, Wistar; Strychnine; Strychnos nux-vomica; Synoviocytes

A neoteric approach for the electrochemical analysis of brucine in biological environment has been developed. The glassy carbon electrode modified with single walled carbon nanotubes and nafion composite film is delineated for the first time to determine brucine employing square wave voltammetry. The quantification of brucine at physiological pH 7.2 manifests remarkable performance at the developed biosensor. The effect of several operational parameters has been studied in the present investigation. Under optimized conditions, a dynamic linear range from 1nM to 8µM with a high sensitivity of 340.8µAµM

Topics: Biosensing Techniques; Carcinoma, Hepatocellular; Cell Line, Tumor; Electrochemical Techniques; Humans; Liver Neoplasms; Nanotubes, Carbon; Strychnine

In this study, a novel NGR (Asn-Gly-Arg) peptide-modified liposomal brucine was prepared by using spray-drying method. The surface morphology of the liposomes, encapsulation efficiency and particle size were investigated. The data showed that the addition of NGR did not produce any significant influence on brucine liposomes in terms of particle size or zeta potential. In addition, after 3 months of storage, no dramatic change such as visible aggregation, drug content changes or precipitation in the appearance of NGR-brucine liposomes occurred. The in vitro release results indicated that the release of brucine from NGR liposomes was similar to that of liposomes, demonstrating that the NGR modification did not affect brucine release. The in vitro drug-release kinetic model of NGR-brucine liposomes fitted well with the Weibull's equation. In vivo, NGR-brucine liposomes could significantly extend the bioavailability of brucine; however, there was no significant difference observed in the pharmacokinetic parameters between liposomes and NGR liposomes after intravenous administration. Antitumor activity results showed that NGR-modified liposomes exhibited less toxicity and much higher efficacy in HepG2-bearing mice compared with non-modified liposomes. The enhanced antitumor activity might have occurred because brucine was specifically recognized by NGR receptor on the surface of tumor cells, which enhanced the intracellular uptake of drugs.

Topics: Animals; Antineoplastic Agents; Biological Availability; Drug Carriers; Drug Liberation; Hep G2 Cells; Humans; Liposomes; Mice; Oligopeptides; Particle Size; Rats, Sprague-Dawley; Strychnine; Xenograft Model Antitumor Assays

In this study, we have aimed to analyze the phytochemical composition of this plant and the concentration of strychnine and brucine. The identification of bioactive compounds was done by GC-MS with NIST Library. Strychnine and Brucine were quantified using HPLC. Twenty one medicinal bio active compounds were identified from the Strychnos nux-vomica leaf ethanolic extract. Strychnine is showing 28.43% purity and brucine was not detected in GCMS analysis. Quantified the concentration of strychnine (0.6 mg in 500mg of extract) and brucine (1.6 mg in 500mg of extract) was done by HPLC against Strychnine and Brucine standard. These compounds are having natural properties of Anti-inflammatory, Hypocholesterole, Cancer preventive, Hepatoprotective, Antimicrobial, Antioxidant, Cardio protective, Antiaging, Antialzheimeran, Antidermatitic, Immunostimulant, Anthepatotoxic, biosynthesis of steroid hormones, Nematicide, Antiandrogenic, 5-alpha reductase inhibitor, antipsychotic, analgesic, apoptotic effect, antidepressant, antidote for snake poisoning and diabetic activity.

Topics: Chromatography, High Pressure Liquid; Gas Chromatography-Mass Spectrometry; Plant Extracts; Plant Leaves; Strychnine; Strychnos nux-vomica

Brucine is the active component in traditional Chinese medicine "Ma-Qian-Zi" (Strychnos nux-vomica Linn), with capabilities of analgesic, anti-inflammatory, anti-tumor and so on. It is crucial how to break through the impact of cuticle skin which reduces the penetration of drugs to improve drug transmission rate. The aim of this study is to improve the local drug concentration by using ultrasound. We used fresh porcine skin to study the effects of ultrasound on the transdermal absorption of brucine under the influence of various acoustic parameters, including frequency, amplitude and irradiation time. The transdermal conditions of yellow-green fluorescent nanoparticles and brucine in skin samples were observed by laser confocal microscopy and ultraviolet spectrophotometry. The results show that under ultrasonic conditions, the permeability of the skin to the fluorescent label and brucine (e.g., the depth and concentration of penetration) is increased compared to its passive diffusion permeability. The best ultrasound penetration can make the penetration depth of more than 110 microns, fluorescent nanoparticles and brucine concentration increased to 2-3 times. This work will provide supportive data on how the brucine is better used for transdermal drug delivery (TDD).

Topics: Administration, Cutaneous; Animals; Anti-Inflammatory Agents; Drug Carriers; Fluorescent Dyes; Nanoparticles; Skin Absorption; Strychnine; Strychnos nux-vomica; Swine; Ultrasonic Therapy

To examine the effects of brucine on the invasion, migration and bone resorption of receptor activator of nuclear factor-kappa B ligand (RANKL)-induced osteoclastogenesis.. The osteoclastogenesis model was builded by co-culturing human breast tumor MDA-MB-231 and mouse RAW264.7 macrophages cells. RANKL (50 ng/mL) and macrophage-colony stimulating factor (50 ng/mL) were added to this system, followed by treatment with brucine (0.02, 0.04 and 0.08 mmol/L), or 10 μmol/L zoledronic acid as positive control. The migration and bone resorption were measured by transwell assay and in vitro bone resorption assay. The protein expressions of Jagged1 and Notch1 were investigated by Western blot. The expressions of transforming growth factor-β1 (TGF-β1), nuclear factor-kappa B (NF-κB) and Hes1 were determined by enzyme-linked immunosorbent assay.. Compared with the model group, brucine led to a dose-dependent decrease on migration of MDA-MB-231 cells, inhibited RANKL-induced osteoclastogenesis and bone resorption of RAW264.7 cells (P<0.01). Furthermore, brucine decreased the protein levels of Jagged1 and Notch1 in MDA-MB-231 cells and RAW264.7 cells co-cultured system as well as the expressions of TGF-β1, NF-κB and Hes1 (P<0.05 or P<0.01).. Brucine may inhibit osteoclastogenesis by suppressing Jagged1/Notch1 signaling pathways.

Topics: Animals; Bone Neoplasms; Breast Neoplasms; Cell Differentiation; Cells, Cultured; Female; Humans; Jagged-1 Protein; Macrophages; Mice; Osteoclasts; Receptor, Notch1; Signal Transduction; Strychnine

To prepare the liposomes and transfersomes of brucine, characterize their pharmaceutical properties, and compare their in vitro transdermal permeation properties. The liposomes and transfersomes of brucine were prepared by ammonium sulfate gradient method to investigate their pharmaceutical properties such as the particle size, encapsulation efficiency and deformation. The transdermal permeation properties in vitro of liposome and transfersomes from different prescriptions were compared by using modified Franz-diffustion cells with rat skin as the transdermal barrier. The results showed that the particle size of liposomes and transfersomes for brucine ranged from 100 nm to 150 nm, with even distribution for particle size. As compared with the soybean phosphatidylcholine (SPC) transfersomes, the encapsulation efficiency of complex phospholipid transfersomes was significantly improved. The deformation index of complex phospholipid transfersomes in brucine was 2.09 times and 1.76 times as much as SPC liposomes and SPC transfersomes respectively. The steady state flux of complex phospholipid transfersomes was 3.43 times and 1.41 times as much as SPC liposomes and SPC transfersomes. The steady state flux of the physical mixture of brucine and blank complex phospholipid transfersomes was 2.20 times as much as brucine solution. The concentration of complex phospholipid had effect on transdermal permeation of blank transfersomes. In conclusion, as compared with liposomes, the permeation behavior of transfersomes was significantly improved; complex phospholipid technology can improve the membrane phase behavior of transfersomes, and further improve the deformation index and transdermal flux of transfersomes; in addition, blank transfersomes have promoting effect on transdermal absorption of brucine.

Topics: Administration, Cutaneous; Animals; Drug Carriers; Liposomes; Particle Size; Rats; Skin; Skin Absorption; Strychnine

We designed two novel polymer materials N-glycyrrhetinic acid-polyethylene glycol-chitosan derivatives (NGPC) and N-quaternary ammonium-chitosan derivatives (NQC). We prepared three kinds of drug loaded chitosan nanoparticles (brucine/NGPC-NPs, brucine/NQC-NPs, brucine/MNPs) by ionic crosslinking method with brucine as a model drug and chitosan nanoparticles（brucine/NGPC-NPs, brucine/NQC-NPs） as the reference formulation. Using high content analysis, flow cytometry, immunofluorescence, transmission electron microscopy and other advanced technology, we tested the effect of 20 μg·mL(-1) concentration of brucine solution and brucine/ chitosan nanoparticles（brucine/CTS-NPs） in hepatocarcinoma (HEpG2) cells and evaluated the apoptosis induced by the treatment. The results suggested that brucine-CTS/NPs had a strongest activity in killing tumor cells, and increased the total cell apoptosis rate with a significant formation of "crescent-shaped" body, swelling mitochondria, mitochondria cristae missing, decreased mitochondrial membrane potential and release of cytochrome C. The activity was enhanced by multifunctional nanocomposite particles that increased the cumulative amount of drug in the mitochondria for the anti-tumor effect.

Topics: Apoptosis; Carcinoma, Hepatocellular; Chitosan; Drug Carriers; Glycyrrhetinic Acid; Hep G2 Cells; Humans; Liver Neoplasms; Membrane Potential, Mitochondrial; Microscopy, Electron, Transmission; Nanoparticles; Polymers; Strychnine

To isolate and identify the chemical constituents from the seeds of Strychnos nux-vomica.. Chromatographic separation techniques such as silica gel chromatography,ODS chromatography and Sephadex LH-20 chromatography were used for the isolation and purification. The structures of the chemical constituents were identified on the basis of mass spectrometry,NMR spectroscopy and so on.. 16 compounds were isolated and their structures were identified as: α-amyrin( 1), vomicine( 2), stearic acid( 3), β-sitosterol( 4),vanillin( 5), ethyl gallate( 6),methyl gallate( 7),novacine( 8),strychnine( 9), daucosterol( 10),brucine chloromethochloride( 11),loganic acid( 12),strychnine chloromethochloride( 13),brucine( 14),geniposide( 15) and loganin( 16).. Compounds 3,6,7 and 15 are isolated from this genus for the first time.

Topics: Iridoids; Seeds; Strychnine; Strychnos nux-vomica

A rapid, simple and sensitive UHPLC-MS/MS method was developed and validated for the simultaneous determination of brucine, strychnine and brucine N-oxide in rat plasma using huperzine A as an internal standard (IS) after protein precipitation with methanol. The analytes were separated on a Purospher® STAR RP18 UHPLC column (2 µm, 2.1 × 100 mm) by gradient elution using a mobile phase composed of methanol and water (containing 0.1% formic acid) at a flow rate of 0.3 mL/min. Brucine, strychnine, brucine N-oxide and IS were detected in positive ion multiple reaction monitoring mode by means of an electrospray ionization interface (m/z 395.2 → 324.1, m/z 335.2 → 184.1, m/z 411.2 → 394.2, m/z 243.1 → 226.1). The calibration curve was linear over the range of 1-500 ng/mL for brucine and strychnine and 0.2-50 ng/mL for brucine N-oxide. The intra- and inter-day precisions of these analytes were all within 15% and the accuracy ranged from 85 to 115%. The stability experiment indicated that the plasma samples at three concentration levels were stable under different conditions. The developed method was successfully applied for the first time to pharmacokinetic studies of brucine, strychnine and brucine N-oxide following a single oral and intravenous administration of modified total alkaloid fraction in rats. Copyright © 2016 John Wiley & Sons, Ltd.

Topics: Animals; Chromatography, High Pressure Liquid; Cyclic N-Oxides; Limit of Detection; Rats; Reproducibility of Results; Strychnine; Tandem Mass Spectrometry

A rapid, specific and sensitive liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the simultaneous determination of strychnine, brucine, strychnine N-oxide and brucine N-oxide in rat plasma. Plasma samples were pretreated via simple protein precipitation with methanol and ephedrine hydrochloride was used as internal standard. Chromatographic separation was carried out on an ZORBAX Eclipse XDB-C18 column (2.1×150mm, 3.5μm) by gradient elution with methanol and 10mM ammonium acetate (adjusted to pH 4.0 with formic acid). The quantification of the analytes was performed by mass spectrometry with TurboIonSpray ionization (ESI) inlet in the positive ion multiple reaction monitoring (MRM) mode. The results showed that the calibration curve was linear in the concentration range of 0.510∼306.3ngmL(-1) for strychnine, brucine and 0.102∼306.0ngmL(-1) for strychnine N-oxide and brucine N-oxide, respectively. The intra- and inter-day precisions were less than 14.9%, and the accuracy ranged from 89.4 to 113% at three QC levels for the 4 analytes. The validated method was successfully applied to the pharmacokinetic study of strychnine, brucine, strychnine N-oxide and brucine N-oxide in rat plasma after oral administration of each monomer and the total alkaloids from Semen Strychni. After single oral administration of the total alkaloids from Semen Strychni at 4 dose levels, Cmax, AUC0-t of strychnine and brucine increased and were proportional to the oral doses. In comparative pharmacokinetics studies, no significant difference was found between each monomer and the total strychnos alkaloids on the pharmacokinetic parameters such as Cmax and AUC. Mean Cmax and AUC of strychnine and brucine were slight increased in the monomer groups in comparison to the total strychnos alkaloids groups, which suggested that some other alkaloids in the Semen Strychni may decrease the absorption of strychnine and brucine in body.

Topics: Animals; Chromatography, Liquid; Loganiaceae; Rats; Strychnine; Tandem Mass Spectrometry

The c-myb gene is a potential therapeutic target for human tumors and leukemias. Active ingredients from natural products may be used as drugs in chemotherapy for human cancers. Here, electrospray ionization mass spectrometry (ESI-MS) was used to probe the formation and recognition of the G-quadruplex structure from the G-rich sequence that is found in the c-myb gene promoter, 5'-GGGCTGGGCTGGGCGGGG-3'. The aim of our study is to evaluate a potential binder for the c-myb gene from natural products, and thereby to modulate c-myb gene expression.. ESI-MS, as an effective method, was utilized not only to characterize the formation of the G-quadruplex in the c-myb oncogene, but also as a tool to probe the binding characteristics of alkaloid molecules with the target G-quadruplex DNA.. ESI-MS results with the support of circular dichroism (CD) spectra demonstrated the formation of an intramolecular parallel-stranded G-quadruplex in the c-myb oncogene promoter. A screening of six alkaloid molecules showed that brucine (P1) had a strong binding affinity to the c-myb G-quadruplex DNA. It is notable that P1 can bind selectively to the c-myb G-quadruplex with respect to duplex DNAs, as well as to G-quadruplexes in other types of gene sequences. According to ESI-MS results, in which the stability was tested by capillary heating and collision-induced dissociation, the binding of P1 could thermally stabilize the c-myb G-quadruplex DNA.. In this work, brucine (P1), an alkaloid molecule, has been found to bind to the intramolecular parallel G-quadruplex in the c-myb oncogene promoter with high affinity and selectivity, and could thermally stabilize the c-myb G-quadruplex DNA, indicating that the binding of P1 has the potential to modulate c-myb gene expression. Copyright © 2015 John Wiley & Sons, Ltd.

Topics: Binding Sites; G-Quadruplexes; Humans; Kinetics; Promoter Regions, Genetic; Proto-Oncogene Mas; Proto-Oncogene Proteins c-myb; Spectrometry, Mass, Electrospray Ionization; Strychnine

A sensitive and efficient mixed cloud point extraction combined with high-performance liquid chromatography was developed for the simultaneous separation and determination of four alkaloids (strychnine, strychnine N-oxide, brucine, and brucine N-oxide) in plasma after the oral administration of processed semen strychni extract. Tergitol TMN-6 and cetyl-trimethyl ammonium bromide were chosen as the mixed surfactants, and ultrasound was employed to enhance the extraction efficiency. Some important parameters affecting the mixed cloud point extraction efficiency, such as the content of Tergitol TMN-6 and cetyl-trimethyl ammonium bromide, pH, salt effect, extraction temperature, and ultrasound time were studied and optimized. Under optimum conditions, the linear range of four alkaloids was from 1.0 to 1000 ng/mL. All correlation coefficients of the calibration curves were higher than 0.9993. The intraday and interday precision were below 8.65% and the limits of detection for the four alkaloids were less than 1.0 ng/mL (S/N = 3).

Topics: Administration, Oral; Chromatography, High Pressure Liquid; Cyclic N-Oxides; Drugs, Chinese Herbal; Humans; Plant Extracts; Strychnine; Ultrasonic Waves

This study was designed to investigate the combination effects of brucine and gemcitabine, each with anticancer properties, in MCF-7 human breast cancer cells in culture. With regard to cell viability, effects of both the drugs and their combinations were inversely proportional to dose and time. For various proportional drug combinations studied, combination effects were analysed using CompuSyn software. The analyses revealed synergistic and/or additive effects regarding cell viability, anchorage-independent growth and cell migration. Combination analyses exhibited diversified impacts of the type of combination treatment, namely pretreatment with either drug followed by exposure to the other, or treatment with both drugs at the same time. Compared with untreated cells, combination treatment of asynchronised MCF-7 cells resulted in 17.2 × decrease in G2 phase, increasing G1 (2.1 × ) and S (1.5 × ) phase cells in cell cycle analysis. Brucine, either individually or in combination, but not gemcitabine, inhibited NF-kB subunit (p65) expression in MCF-7 cells.

Topics: Antimetabolites, Antineoplastic; Breast Neoplasms; Cell Cycle; Deoxycytidine; Drug Therapy, Combination; Female; Gemcitabine; Humans; MCF-7 Cells; Strychnine

Alkaloids constitute a large family of natural products possessing diverse biological properties. Their unique and complex structures have inspired numerous innovations in synthetic chemistry. In the realm of late-stage C-H functionalization, alkaloids remain a significant challenge due to the presence of the basic amine and a variety of other functional groups. Herein we report the first examples of dirhodium(II)-catalysed intermolecular C-H insertion into complex natural products containing nucleophilic tertiary amines to generate a C-C bond. The application to a diverse range of alkaloids and drug molecules demonstrates remarkable chemoselectivity and predictable regioselectivity. The capacity for late-stage diversification is highlighted in the catalyst-controlled selective functionalizations of the alkaloid brucine. The remarkable selectivity observed, particularly for site-specific C-H insertion at N-methyl functionalities, offers utility in a range of applications where efficient installation of synthetic handles on complex alkaloids is desired.

Topics: Alkaloids; Amines; Biological Products; Carbon; Catalysis; Chemistry, Pharmaceutical; Drug Design; Hydrogen; Magnetic Resonance Spectroscopy; Rhodium; Strychnine

Enantio- and diastereodivergent approaches to pyrrolidines are described by using catalyst- and substrate-controlled reaction pathways. A concerted endo-selective [3 + 2]-cycloaddition pathway is developed for the reaction of methyl imino ester, whereas endo-pyrrolidines with an opposite absolute stereochemical outcome are prepared by using the stepwise reaction pathway of tert-butyl imino ester. The development of catalyst- and substrate-controlled stereodivergent approaches highlights the inherent substrate-catalyst interactions in the [3 + 2]-cycloaddition reactions of metalated azomethine ylides.

Topics: Azo Compounds; Catalysis; Copper; Molecular Structure; Pyrrolidines; Stereoisomerism; Strychnine; Thiosemicarbazones

The cytotoxicities of the two alkaloids strychnine and brucine from the seed of Strychnos nux-vomica and their interaction with DNA were investigated. A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrasolium bromide (MTT) assay was used to examine the growth inhibitory effects of these alkaloids on Vero cells after 24, 48 and 72h of incubation. The cytotoxicities of strychnine and brucine were found to be time- and concentration-dependent. Strychnine was determined to be more toxic to Vero cells than brucine. At the same time, the interactions of strychnine and brucine with DNA were investigated using neutral red (NR) dye as a probe by UV-vis spectroscopy, fluorescence spectroscopy, and an examination of the ionic strength effect, and the effects of alkaloids on DNA melting were also examined. The results indicated that a DNA-brucine mixture but not a DNA-strychnine mixture could be extracted from Vero cells after treatment with brucine and strychnine, respectively. Brucine competitively intercalated into the DNA double-helix causing fluorescence quenching of the DNA-NR system. UV absorption spectroscopy and the melting temperature (Tm) curve also provided evidence that brucine interacted with DNA through intercalation. Furthermore, the results of the ionic strength effect experiment suggested that electrostatic interactions between brucine and phosphate groups in the DNA backbone might also play an important role in the binding of brucine to DNA.

Topics: Animals; Chlorocebus aethiops; Cytotoxins; DNA; Drugs, Chinese Herbal; Nucleic Acid Denaturation; Osmolar Concentration; Seeds; Strychnine; Strychnos nux-vomica; Vero Cells

The overwhelming majority of drugs exert their pharmacological effects after reaching their target sites of action, however, these target sites are mainly located in the cytosol or intracellular organelles. Consequently, delivering drugs to the specific organelle is the key to achieve maximum therapeutic effects and minimum side-effects. In the work reported here, we designed, synthesized, and evaluated a novel mitochondrial-targeted multifunctional nanoparticles (MNPs) based on chitosan derivatives according to the physiological environment of the tumor and the requirement of mitochondrial targeting drug delivery. The intelligent chitosan nanoparticles possess various functions such as stealth, hepatocyte targeting, multistage pH-response, lysosomal escape and mitochondrial targeting, which lead to targeted drug release after the progressively shedding of functional groups, thus realize the efficient intracellular delivery and mitochondrial localization, inhibit the growth of tumor, elevate the antitumor efficacy, and reduce the toxicity of anticancer drugs. It provides a safe and efficient nanocarrier platform for mitochondria targeting anticancer drug delivery.

Topics: Animals; Antineoplastic Agents; Apoptosis; Chitosan; Chromatography, High Pressure Liquid; Cytoplasm; Cytosol; Drug Carriers; Drug Delivery Systems; Drug Liberation; Hep G2 Cells; Hepatocytes; Humans; Hydrogen-Ion Concentration; Lysosomes; Male; Mice; Mice, Inbred ICR; Microscopy, Confocal; Mitochondria; Nanoparticles; Nanotechnology; Neoplasm Transplantation; Neoplasms; Rats; Rats, Wistar; Schiff Bases; Strychnine

To provide insights into the mechanism for the attenuate-synergistic effect of Zuota to Tibetan medicine Renqing Mangjue, a contrasted study was carried out on the pharmacokinetics of brucine and strychnine in mice plasm, which are active and toxicant ingredient in the Tibetan medicine Renqing Mangjue. LC-MS/MS was used to detect simultaneously the concentrations of brucine and strychnine in mice plasm at-different time intervals after administration parallelly and randomly, and the pharmacokinetic software Kinetica 5. 0 was selected to non-compartmental analysis (NCA) for data, and statistical analysis software SPSS 19. 0 was used for significance test on the pharmacokinetic parameters. A reliable LC-MS/MS method was established for the determination of brucine and strychnine in blood plasma, which are consistent with the requirements of the preclinical pharmacokinetic study confirmed by the methodology. The linear concentration ranges of brucine and strychnine were 0.301-104.4 µg · L(-1) (r = 0.999 5) and 0.305-106 µg · L(-1) (r = 0.999 7), respectively; The intra-day and inter-day variable coefficients were both less than 10.0% with good precision; The average extraction recoveries of brucine and strychnine were 116.23% and 112.82%, and RSD were 3.2% and 2.3% separately;The average matrix effects of brucine and strychnine were 122.48% and 116.36%, and RSD were 7.7% and 4.4%, respectively. The pharmacokinetic results showed that AUCtot of brucine and strychnine in Zuota group were both increased remarkably (P < 0.05), and the Cmax of brucine in Zuota group was about 5.25-fold higher than that of brucine in non-Zuota group (P < 0.05). The Tmax of brucine and strychnine reduced to one-eighth and one-quarter respectively compared with those in Non-Zuota group. In addition, the eliminations of brucine and strychnine in vivo were accelerated after the compatibility of Zuota. A significant difference (P < 0.05) occurred at the MRT0-t, of brucine, while the MRT0-∞ and Lz of strychnine were statistically significant upon the inspection level α = 0.1. It was found that the absorption degree of brucine and strychnine in Zuota group increased in the range of the safe dose (or concentration), while their elimination rates were accelerated, which may be one of the mechanisms for attenuate-synergistic effect of Zuota to Tibetan medicine Renqing Mangjue.

Topics: Animals; Female; Male; Medicine, Tibetan Traditional; Mice; Strychnine

To compare the pharmacokinetic differences of brucine in rats after different administration methods of brucine liposome.. To determine brucine in rat plasma at different points in time by HPLC after oral administration, intramuscular injection, subcutaneous injection and intravenous injection of brucine liposome, respectively. The pharmacokinetic parameters were calculated and analyzed by DAS 3.0.. Compared with other groups, AUC(0 --> t) of subcutaneous injection were higher, C(max) were lower and MRT(0 --> 1), were significantly improved. The pharmacokinetics parameters and absolute bioavailability of brucine show that bioavailability in rats after different administration methods of brucine liposome is subcutaneous injection > intramuscular injection > oral administration.

Topics: Administration, Oral; Animals; Biological Availability; Chromatography, High Pressure Liquid; Injections, Intramuscular; Injections, Intravenous; Injections, Subcutaneous; Liposomes; Rats; Rats, Sprague-Dawley; Strychnine

The objective of this study is to test the hypothesis that the phase transition temperature (T(m)), the main property of liposomes, can be easily controlled by changing the molar ratio of hydrogenated soy phosphatidylcholine (HSPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphacholine (DPPC) after drug encapsulation.. Brucine, an antitumor alkaloid, was encapsulated into the liposomes with different HSPC/DPPC compositions. The T(m)s of the brucine-loaded liposomes (BLs) were determined by differential scanning calorimetry (DSC). Then the physicochemical properties and pharmacokinetics of the BLs with different HSPC/DPPC compositions were investigated and compared.. The results of DSC revealed that HSPC and DPPC can combine into one phase. The findings of molecular modeling study suggested that HSPC interacts with DPPC via electrostatic interaction. The molar ratio of HSPC/DPPC influenced the sizes of BLs but had little effect on the entrapment efficiency (EE). The stability of BLs was improved with the increase of the HSPC ratios, especially with the presence of plasma. Following i.v. administration, it was found that AUC values of BLs in vivo were directly related to the HSPC/DPPC ratios of BLs, namely the T(m)s of BLs.. The behavior of liposomes, especially in vivo pharmacokinetic behavior, can be controlled by the modification of T(m).. The characterization of BLs in vitro and in vivo had demonstrated that the Tm could be flexibly modified for liposomes composed of both HSPC and DPPC. Using HSPC/DPPC composition may be an efficient strategy to control the T(m), thus control the in vivo pharmacokinetic behavior, of BLs.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Animals; Drug Evaluation, Preclinical; Glycine max; Hydrogenation; Liposomes; Male; Phosphatidylcholines; Random Allocation; Rats; Rats, Sprague-Dawley; Strychnine

Semen Strychni, a known toxic drug in Chinese pharmacopoeia, is notable for its therapeutic effects on local muscle and joint pain. However, oral administration can be risky. Topically administered drugs accumulate in the topical muscles and knee joints without any major increase in plasma levels; only non-protein-bound drugs in the biological fluids of target tissues are effective for therapeutic effects. A sensitive and rapid ultra performance liquid chromatography - mass spectrometry (UPLC-MS) method coupled with a microdialysis technique was developed to determine the non-protein-bound strychnine (Str) and brucine (Bru) in rabbit muscle and synovial fluid microdialysate. The UPLC separation was carried out using a 1.7μm BEH C18 column (50 mm × 2.1 mm) with a mobile phase consisting of methanol: water (29.5:70.5, v/v) with 0.1% formic acid and 20 mM ammonium acetate in water. The method was validated at concentrations ranging from 0.58 ng/ml to 467.20 ng/ml for Str and from 0.42 ng/ml to 422.40 ng/ml for Bru. Intra-day and inter-day accuracy ranged from 99.1% to 103.2% for Str and from 95.8% to 108.8% for Bru with intra-day and inter-day precision within 9.7%. The proposed method was successfully applied to determine non-protein-bound Str and Bru, and the analysates concentration remained stable in rabbit muscle and synovial fluid after topical application of total Strychnos alkaloid patches, which indicated that total Strychnos alkaloid patches could substitute for the traditional oral administration of Semen Strychni.

Topics: Administration, Topical; Analgesics; Animals; Calibration; Chromatography, High Pressure Liquid; Male; Muscles; Rabbits; Seeds; Strychnine; Strychnos; Synovial Fluid; Tandem Mass Spectrometry; Transdermal Patch

To increase the intra-articular (IA) retention time of osteoarthritis drugs in the synovial cavity and slow down the burst release of microspheres (MPs), we prepared a novel drug delivery system named nanoparticles-in-microspheres (NiMs). The system was constructed by dispersing the brucine-loaded nanoparticle, which was prepared by an emulsification method in the MPs. The NiMs were characterized by scanning electron microscope, Fourier transform infrared spectra and differential scanning calorimetry. After investigating the biocompatibility with synovium of NiMs in rats, the pharmacokinetics was studied and FX-imaging was used to visualize the transmission of nanoparticles after IA administration in rats. From the results, we know that the NiMs were spherical, there was no chemical bond between the drug and the polymer, and the drug was dispersed in the polymer in an amorphous form. Compared with MPs (41%), the burst release of NiMs could be slowed down to 9%. After that, the drug was released from NiMs by diffusion. The results of FX imaging in rats showed that the NiMs could stay in the articular cavity for over 11 d. The studies of pharmacokinetics revealed that the NiMs could slow down the burst release and improve retention in vivo. This study demonstrates the feasibility of using NiMs to slow down the burst release and increase the retention of therapeutic agents in articular joints.

Topics: Animals; Drug Carriers; Drug Delivery Systems; Injections, Intra-Articular; Male; Microspheres; Nanoparticles; Osteoarthritis; Particle Size; Rats; Rats, Sprague-Dawley; Strychnine; Synovial Fluid; Synovial Membrane

Brucine is a widely prescribed glycine antagonist, but a complete understanding of its metabolic pathway is still lacking. The present work represents the first investigation of in vivo metabolism of brucine in rats using LC-ESI-ion trap-TOF-MS.. A total of 12 Phase I and five Phase II metabolites were tentatively identified. Brucine can be metabolized by hydrolysis, demethylation and methoxylation, in addition to diverse oxidations in a Phase I manner followed by glucuronidation in Phase II metabolism. Both the renal and biliary routes were observed for the excretion of brucine and its metabolites.. Our results update the metabolism and disposition data on brucine, which provides basic information for better understanding of the pharmacological and toxicological activities of brucine-containing medicines.

Topics: Alkaloids; Animals; Bile; Biotransformation; Chromatography, High Pressure Liquid; Glucuronidase; Hydrolysis; Hydroxylation; Indoles; Metabolic Networks and Pathways; Oxidation-Reduction; Rats; Spectrometry, Mass, Electrospray Ionization; Strychnine

An on-line sample preconcentration method by two-step stacking i.e., sweeping and micelle to solvent stacking, in capillary zone electrophoresis (CZE) has been developed for the determination of strychnine and brucine in traditional Chinese herbal medicines. After experimental optimizations, the best separation was achieved by using 75 mM phosphate buffer (pH 2.5) with 30% methanol (v/v). Compared with normal CZE injection, 51- and 38-fold improvement in concentration sensitivity was achieved for strychnine and brucine, respectively. The calibration curve was linear in the range of 0.1-5.0 μg mL(-1) for both strychnine and brucine, with the correlation coefficients of 0.9998 and 0.9997, respectively. The limits of detection (S/N=3) for both alkaloids were 0.01 μg mL(-1). The inter-day (n=8) and intra-day (n=5) reproducibilities expressed as the relative standard deviations for corrected peak area were less than 9.5%. The method was applied to determine strychnine and brucine in two Chinese herbal medicines, with recoveries ranging from 94.2% to 105.4%. The results indicated that the method is simple, rapid, reliable, and can be applied to determine strychnos alkaloids in traditional Chinese herbal medicines.

Topics: Calibration; Drugs, Chinese Herbal; Electrophoresis, Capillary; Equipment Design; Limit of Detection; Micelles; Strychnine

Recently, the renal injury caused by Semen strychni and its major toxic constituents, strychnine and brucine, was reported in many clinical cases. Hence, this study was conducted to investigate the renal injury induced by Semen Strychni and the protective effects of Radix Glycyrrhizae and Rhizoma Ligustici. The protective mechanisms were related to the comparative toxicokinetics of strychnine and brucine. Serum and urine uric acid and creatinine were used as renal function markers to evaluate the condition of kidney, and renal injury was directly reflected by histopathological changes. Compared with rats in blank group and protective herb groups, rats in Semen Strychni high-dose group showed significant differences in the results of renal function markers, and various glomerular and tubular degenerations were found in the histopathological study. The decreased AUC (only strychnine) and Cmax, the increased Tmax by Radix Glycyrrhizae and the decreased T1/2 by Radix Glycyrrhizae and Rhizoma Ligustici were found in model groups. Results indicated that high dose of Semen Strychni might induce renal injury. Radix Glycyrrhizae and Rhizoma Ligustici might work together and have effects on the elimination of strychnine and brucine. The protective effects of Radix Glycyrrhizae might also be explained by the slow absorption of the alkaloids.

Topics: Animals; Biomarkers; Creatinine; Dose-Response Relationship, Drug; Drugs, Chinese Herbal; Glycyrrhiza; Herb-Drug Interactions; Kidney; Kidney Diseases; Male; Plant Extracts; Rats; Rats, Sprague-Dawley; Rhizome; Strychnine; Strychnos; Toxicokinetics; Uric Acid

The aim of the study was to investigate the possibility of improving the therapeutic efficacy of the total alkaloid fraction (TAF) extracted from processed nux vomica by reducing the strychnine contents. Most strychnine was removed from TAF to obtain the modified total alkaloid fraction (MTAF). The toxicity and pharmacokinetics of TAF and MTAF were further investigated and compared besides their antitumor, analgesic and anti-inflammatory activities. The results showed that the ratios of brucine to strychnine were 1:2.05 and 2.2:1 for TAF and MTAF, respectively, and the toxicity of TAF was about 3.17-fold higher than that of MTAF. Compared to brucine alone, the elimination of brucine was found to be inhibited by other alkaloids in TAF or MTAF except strychnine. Significantly increased pharmacological activities when administered by the oral route were obtained with MTAF in comparison to TAF and nux vomica powder (NVP). In summary, MTAF might replace NVP and TAF in the clinical application of Chinese medicine to obtain much higher efficacy.

Topics: Analgesics; Animals; Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Agents, Phytogenic; Area Under Curve; Cell Line, Tumor; Cell Survival; Half-Life; Humans; Lethal Dose 50; Mice; Plant Extracts; Seeds; Strychnine; Strychnos nux-vomica

A novel brucine imprinted polymer was prepared on multi-walled carbon nanotubes by reversible addition-fragmentation chain transfer (RAFT) precipitation polymerization. The polymer was further grafted with hydrophilic poly(glycerol monomethacrylate) brushes to improve its water-compatibility. The obtained molecularly imprinted material showed enhanced accessibility to brucine and improved selective recognition property in water medium. When the material was supported on an ionic liquid functionalized graphene coated glassy carbon electrode for the electrochemical determination of brucine, the resulting electrochemical sensor presented good analytical performance. Under the optimized conditions, the peak current was linear to brucine concentration in the ranges of 0.006-0.6 μM and 0.6-5.0 μM with sensitivities of 15.3 μA/μMmm(2) and 5.4 μA/μM mm(2), respectively; the detection limit was 2 nM (S/N=3). The sensor was successfully applied to the determination of brucine in practical samples and the recovery for the standards added was 94-104%.

Topics: Conductometry; Equipment Design; Equipment Failure Analysis; Fractional Precipitation; Hydrophobic and Hydrophilic Interactions; Methylmethacrylates; Molecular Imprinting; Nanotubes, Carbon; Reproducibility of Results; Sensitivity and Specificity; Solutions; Strychnine; Surface Properties; Water

The density functional theoretical (DFT) computations were performed at the B3LYP/6-311G++(d, p) level to calculate the equilibrium geometry, vibrational wave numbers, intensities, and various other molecular properties of brucine and strychnine, which were found in satisfactory agreement with the experimental data. The out-of-phase stretching modes of aromatic rings and carbonyl stretching modes in combination with CH stretching modes at stereogenic centers generate VCD signals, which are remarkably efficient configuration markers for these chiral molecular systems. NBOs analysis reveals that the large values of second order perturbation energy (47.24kcal/mol for brucine and 46.93kcal/mol for strychnine) confirms strong hyperconjugative interaction between the orbital containing the lone pair of electron of nitrogen and the neighboring CO antibonding orbital. The molecular electrostatic potential map of strychnine molecule, with no polar groups other than the lone keto group, shows less polarization, which accounts for its lower susceptibility towards electrophilic attack as compared to brucine.

Topics: Analgesics; Convulsants; Models, Molecular; Quantum Theory; Spectrophotometry, Infrared; Spectrophotometry, Ultraviolet; Static Electricity; Strychnine; Strychnos; Thermodynamics

Brucine (BRU) extracted from the seeds of Strychnos nux-vomica L is glycine receptor antagonist. We hypothesize that BRU may modify alcohol consumption by acting at glycine receptors, and evaluated the pharmacodynamic profiles and adverse effects of BRU in rat models of alcohol abuse.. Alcohol-preferring Fawn-Hooded (FH/Wjd) rats were administered BRU (10, 20 or 30 mg/kg, sc). The effects of BRU on alcohol consumption were examined in ethanol 2-bottle-choice drinking paradigm, ethanol/sucrose operant self-administration paradigm and 5-d ethanol deprivation test. In addition, open field test was used to assess the general locomotor activity of FH/Wjd rats, and conditioned place preference (CPP) was conducted to assess conditioned reinforcing effect.. In ethanol 2-bottle-choice drinking paradigm, treatment with BRU for 10 consecutive days dose-dependently decreased the ethanol intake associated with a compensatory increase of water intake, but unchanged the daily total fluid intake and body weight. In ethanol/sucrose operant self-administration paradigms, BRU (30 mg/kg) administered before each testing session significantly decreased the number of lever presses for ethanol and the ethanol intake, without affecting the number of sucrose (10%) responses, total sucrose intake, and the number of lever presses for water. Acute treatment with BRU (30 mg/kg) completely suppressed the deprivation-induced elevation of ethanol consumption. Treatment with BRU (10, 20, and 30 mg/kg) did not alter locomotion of FH/Wjd rats, nor did it produce place preference or aversion.. BRU selectively decreases ethanol consumption with minimal adverse effects. Therefore, BRU may represent a new pharmacotherapy for alcoholism.

Topics: Alcohol Drinking; Alcoholism; Animals; Ethanol; Male; Motor Activity; Rats; Rats, Inbred Strains; Receptors, Glycine; Strychnine; Strychnos nux-vomica

An ionic liquid-based electromembrane extraction (IL-EME) method was presented, and its performance was compared with 2-ethylnitrobenzene (ENB) based EME for the determination of strychnine and brucine in human urine. For the two methods, the fundamental extraction parameters such as supported liquid membrane, voltage, extraction time, pH values of sample solution and acceptor solution, temperature and salting-out effect were separately optimized. IL-EME provided 96- and 122-fold enrichment factors for strychnine and brucine, respectively, which were better than those obtained in EME (83- and 86-fold, respectively). The calibration curves were linear over the ranges of 20-720 μg L(-1) for strychnine and 20-640 μg L(-1) for brucine with the correlation coefficients higher than 0.9950. The repeatability of EME and IL-EME were evaluated by five parallel experiments giving the relative standard deviations of 5.12-6.98%. As the results indicated, compared with ENB based EME, the proposed IL-EME is more reliable and could provide better extraction performance for the determination of strychnine and brucine in human urine.

Topics: Calibration; Humans; Ionic Liquids; Membranes, Artificial; Reproducibility of Results; Solvents; Strychnine; Urinalysis

It has been proven that Cyclin D1 and Cyclin E are the important positive regulators of cell cycle, they are closely related to the tumor proliferation. The aim of this study is to explore the relationship between Brucine and the proliferation in human lung cancer cell line PC-9, and the effect of it on the expression of Cyclin D1 and Cyclin E.. PC-9 cells were divided to 4 groups: the normal control group, the DMSO control group (2‰), the 150 μM Brucine group, and the 300 μM Brucine group. The proliferation rate of PC-9 cells was determined by The CellTiter-Glo Luminescent Cell Viability Assay and Colony Formation assay. The change of cell cycle was detected by Flow cytome try. Expressions of cell cycle regulators Cyclin D1, Cyclin E mRNA were determined by qRT-PCR. Protein expression of cell cycle regulators Cyclin D1, Cyclin E were determined by Western blot.. Compared with the control, Brucine remarkably inhibited the proliferation of PC-9 cells in a dose- and time-dependent manner (P<0.01); Flow cytome try showed that Brucine blocked the cell cycle of PC-9 cells at G0/G1, and the differences were statistically significant (P<0.01); qRT-PCR showed that the expression of Cyclin D1, Cyclin E mRNA were down-regulated; Western blot showed that the protein expression of Cyclin D1, Cyclin E were down-regulated.. Brucine can inhibit the proliferation of human lung cancer cell line PC-9 mainly by blocking the cell cycle at G0/G1 via down-regulating the expression of Cyclin D1, Cyclin E.. 背景与目的 已有的研究表明：Cyclin D1和Cyclin E是细胞周期中重要的正性调控因子，其高表达与肿瘤的增殖密切相关。本研究旨在探讨马钱子碱（Brucine）对人肺癌细胞株PC-9增殖的影响，及其与Cyclin D1和Cyclin E表达的影响。方法 将PC-9细胞分为4组：空白对照组、DMSO对照组（2‰）、150 μM Brucine组、300 μM Brucine组。CellTiter-Glo发光法、平板克隆形成实验观察该药对PC-9细胞增殖的影响，流式细胞仪检测细胞周期，qRT-PCR检测细胞周期相关基因Cyclin D1、Cyclin E mRNA的表达，Western blot检测细胞周期相关基因Cyclin D1、Cyclin E蛋白的表达。结果 与对照组比较，CellTiter-Glo发光法、平板克隆形成实验结果显示：Brucine可以抑制人肺癌细胞株PC-9的增殖，并呈时间-剂量依赖性（P<0.01）；流式结果显示对细胞周期的影响主要是阻滞PC-9细胞于G0/G1期；qRT-PCR结果显示Cyclin D1、Cyclin E mRNA的表达下调；Western blot结果显示Brucine使Cyclin D1、Cyclin E的表达降低。结论 Brucine能明显抑制人肺癌细胞株PC-9的增殖，机制主要与其通过下调Cyclin D1、Cyclin E表达，进而阻滞细胞周期有关。

Topics: Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cyclin D1; Cyclin E; Down-Regulation; Drugs, Chinese Herbal; Gene Expression Regulation, Neoplastic; Growth Inhibitors; Humans; Lung Neoplasms; Strychnine

This study was aimed to investigate the inducing-apoptosis effect of brucine on human monocytic leukemia cell line THP-1 cells and its possible mechanism. The inhibition effect of brucine on growth of THP-1 cells was measured by CCK-8 method. Morphological changes of THP-1 cells treated with brucine was detected by acridine orange/ethidium bromide (AO/EB)double staining. Annexin-V/PI double labeling method was used to assay the apoptosis rate of THP-1 cells. The effect of brucine on THP-1 cell cycle distribution was detected by PI single staining. RT-PCR was used to detect the expression of BCL-2 and BAX. The results showed that the brucine could inhibit the THP-1 cell growth in concentration and time-dependent manners at the range of 50 to 400 µg/ml. The cells stained with AO/EB revealed that the brucine induced the nuclear chromatin condensation. After the THP-1 cells were treated with brucine of 400µg/ml for 48 hours, most nucleic were stained as orange-red, and condensed, displaying the late apoptotic cell morphology. Annexin-V/PI detection showed that brucine could induce apoptosis of THP-1 cells in a concentration-dependent manner. Compared with the control group, more cells in brucine-treated group were arrested at G0/G1 phase in a concentration-dependent manner. RT-PCR detection revealed that the expression of BCL-2 was down-regulated strikingly and BAX was up-regulated. It is concluded that brucine can efficiently inhibit cell growth and block THP-1 cells in G0/G1 phase. The mechanism of THP-1 cell apoptosis induced by brucine may be related to the inhibition of BCL-2 and activation of BAX.

Topics: Apoptosis; bcl-2-Associated X Protein; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Humans; Proto-Oncogene Proteins c-bcl-2; Strychnine

Thermosensitive liposomes (TSL) in combination with local hyperthermia (HT) represent a promising tool for tumor specific drug delivery. The objective of the study was to investigate the influence of phase transition temperature (Tm) on the properties of TSL. High temperature triggered TSL (HTSL), low temperature triggered TSL (LTSL) and non-TSL (NTSL) were prepared and temperature sensitive release properties were extensively compared in different media. Mouse plasma was determined to have similar effect on the release profiles compared to human plasma, in which complete release were obtained at 38 °C and 40 °C for LTSL and HTSL, respectively. The temperature at which complete release achieved was found to be obviously lower than Tm. Brucine, an antitumor alkaloid, was encapsulated into different TSLs. After HT treatment, the viabilities of SMMC 7721 cells were determined to be 21.3±3.8% and 16.8±3.3% for 127 μM brucine LTSL and HTSL, respectively. Treating the tumor-bearing mice with LTSL, HTSL and NTSL led to significantly increased brucine uptake in the heated tumor site compared to the brucine solution group by 2.30, 3.80 and 2.26-fold, respectively. The results of this study suggested that Tm of TSL should be increased to obtain improved drug delivery efficiency to tumor.

Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Doxorubicin; Drug Delivery Systems; Drug Liberation; Fluoresceins; Humans; Liposomes; Mice; Mice, Inbred ICR; Phase Transition; Phosphatidylcholines; Phosphatidylethanolamines; Polyethylene Glycols; Strychnine; Temperature; Tissue Distribution; Transition Temperature

Glioblastoma multiforme (GBM) is one of the most common glial cell tumors and has drawn more and more attention in the clinic in recent years. Brucine has been reported to significantly suppress gastric cancer, lung cancer, and prostate cancer growth in vivo by inducing cell apoptosis. Here, the effects of brucine on U251 human glioma cell growth were investigated in vitro by cell proliferation assay, FACs, and qPCR in a xenograft tumor model. Treatment with brucine reduced the expression of BCL-2 and cyclooxygenase-2 (COX-2), while upregulated BAX expression in U251 human glioma cells resulted in reduced glioma cell survival rate and inhibited the growth of xenograft tumors. We concluded that brucine has a suppressive effect on U251 human glioma cells in vitro and in vivo, which could help in understanding the role of brucine in glioma cells and guiding drug use in the clinic.

Topics: Animals; Brain Neoplasms; Cell Line, Tumor; Cell Proliferation; Dose-Response Relationship, Drug; Female; Glioblastoma; Growth Inhibitors; Humans; Medicine, Chinese Traditional; Mice; Mice, Nude; Strychnine; Xenograft Model Antitumor Assays

Before the design of brucine-containing transdermal formulations, the pharmacodynamics and pharmacokinetics of brucine following transdermal administration should be evaluated. In this study, the effect of addition of ethanol on solubility of bruicne was investigated and 20% ethanol was added into PBS to obtain 10mg/mL brucine solution. Then three transdermal doses (10, 20 and 40 mg/kg) were administered to mice to evaluate pharmacological activity. It had been demonstrated that brucine possessed analgesic and anti-inflammatory activity in a dose-dependent manner. Cytotoxicities of brucine against various tumor cells including skin tumor cell were also compared in vitro. Brucine was found to possess antitumor activity in a concentration and time-dependent manner and gastrointestinal tumor cells seemed to be more sensitive to brucine. Then in vitro skin permeation behavior and in vivo pharmacokinetics following transdermal administration were further investigated. The cumulative amounts of brucine across mouse skin in vitro were found to be higher than 90%. The absolute bioavailability of brucine was determined to be 40.83%. And compared with intravenous administration, MRT and T1/2 values were increased about 8~12-fold by transdermal route. Moreover, fluctuations of drug levels were found to be significantly decreased in tissues, especially in brain. Finally, no dermal toxicity of brucine was observed. The results of this study indicated that transdermal administration might be beneficial for the sustained efficacy and reduced toxicity of brucine.

Topics: Administration, Cutaneous; Animals; Antineoplastic Agents, Phytogenic; Biological Availability; Cell Line, Tumor; Female; Gastrointestinal Neoplasms; Guinea Pigs; Humans; Male; Mice; Mice, Inbred ICR; Phytotherapy; Plant Extracts; Skin; Solubility; Strychnine; Strychnos nux-vomica

Brucine is an alkaloid from nux vomica, has been shown various pharmacological actions. To study the possible anti-cancer mechanisms on LoVo cells, effects of Brucine on cell viability, cell cycle and apoptosis were investigated. The results showed that Brucine revealed strong growth inhibitory effect on LoVo cells, and caused LoVo cell shrinkage and membrane blobbing, induced cellular and DNA morphological changes. Cell cycle and apoptosis analysis documented that Brucine could change cell cycle and induce cell apoptosis. Brucine-mediated cell cycle arrest in G1 phase was associated with a marked increase of protein levels of CCND1 and decrease in CCNB1, cyclin E and CDC2. In addition, Brucine dose-dependently caused LoVo cells apoptosis evidenced by Annexin V/PI staining Brucine-induced apoptosis was mediated via up-regulation of Bax and down-regulation of Bcl-2. Furthermore, proteins Erk1/2, p38 and Akt phosphorylation were down regulated by Brucine in a dose-dependent manner. In summary, this paper indicates Brucine is effective against LoVo cells proliferation, and promotes LoVo cells death via apoptosis. These results reveal functional interplay among a series of pathway that are deregulated in cancer and suggest that their simultaneous targeting by Brucine could result in efficacious inhibition on cancer cells.

Topics: Apoptosis; Blotting, Western; Cell Death; Cell Line; Cell Line, Tumor; G1 Phase; Humans; Membrane Potential, Mitochondrial; Strychnine

Brucine, one of the main active ingredients in semen Strychni, has been included in many oral prescriptions of traditional Chinese medicine. In this study, we investigated the in vitro metabolism of brucine by human liver microsomes (HLMs) and the metabolic interactions of brucine with the substrates of cytochrome P450 (CYP450). Brucine was incubated with HLMs or CYP3A4 and then analysed by Liquid chromatography/mass spectrometry. The Km and Vmax values for HLMs were 30.53±3.14μM and 0.08±0.0029nmol/mg protein/min, respectively, while the corresponding values for CYP3A4 were 20.12±3.05μM and 6.40±0.21nmol/nmol P450/min. CYP3A4 may be the major enzyme responsible for brucine metabolism in HLMs, other human isoforms of CYP showed minimal or no effect on brucine metabolism. The inhibitory action of brucine was observed in CYP3A4 for the 1'-hydroxylation of midazolam, with inhibitory concentration 50 (IC50) of 8.4-fold higher than specific inhibitors in HLMs. Furthermore, brucine significantly inhibited the CYP3A4-catalyzed midazolam 1'-hydroxylation (Ki=2.14μM) at a concentration lower than 10μM, but no obvious inhibitory effects were observed on other CYP substrates (IC50>50μM). These results suggest that brucine has the potential to interact with a wide range of xenobiotics and endogenous chemicals especially CYP3A4 substrates.

Topics: Biological Assay; Cells, Cultured; Cytochrome P-450 CYP3A; Drugs, Chinese Herbal; Enzyme Activation; Enzyme Inhibitors; Humans; Hydroxylation; Inhibitory Concentration 50; Kinetics; Microsomes, Liver; Midazolam; Strychnine

The HPLC method for determining plasma concentration of brucine was optimized during the study on the effect of the extraction reagent, the extraction frequency and the volume of extraction solvent on the extraction recovery of brucine. The optimum sample treatment method was obtained in the study. Specifically, ammonia water was added, 4 mL extraction solvent (N-hexane-methylene chloride-isopropyl alcohol 65:30:5) were adopted to extract brucine for twice. The method to determine plasma concentration of brucine was applied in pharmacokinetic study to compare pharmacokinetic properties of intravenous injection (5 mg x kg(-1)) and transdermal administration (40 mg x kg(-1)) of brucine aqueous alkali. The results showed that both pharmacokinetic parameters of brucine after intravenous injection and transdermal administration were in conformity with the two-compartment model. After transdermal administration, the absolute bioavailability was calculated to be 18.72%. The optimized HPLC method can satisfy the demands of the pharmacokinetic study on brucine.

Topics: Animals; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Male; Rats; Rats, Sprague-Dawley; Strychnine

Angiogenesis plays an important role in colon cancer development. This study aimed to demonstrate the effect of brucine on tumour angiogenesis and its mechanism of action. The anti-angiogenic effect was evaluated on the chicken chorioallantoic membrane (CAM) model and tube formation. The mechanism was demonstrated through detecting mRNA and protein expressions of VEGFR2 (KDR), PKCα, PLCγ and Raf1 by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot (WB), as well as expressions of VEGF and PKCβ and mTOR by ELISA and WB. The results showed that brucine significantly reduced angiogenesis of CAM and tube formation, inhibited the VEGF secretion and mTOR expression in LoVo cell and down-regulated the mRNA and phosphorylation protein expressions of KDR, PKCα, PLCγ and Raf1. In addition, the effects of brucine on KDR kinase activity, viability of LoVo cell and gene knockdown cell were detected with the Lance™ assay, WST-1 assay and instantaneous siRNA. Compared to that of normal LoVo cells, the inhibition on proliferation of knockdown cells by brucine decreased significantly. These results suggest that brucine could inhibit angiogenesis and be a useful therapeutic candidate for colon cancer intervention.

Topics: Base Sequence; Cell Division; Cell Line, Tumor; Colonic Neoplasms; DNA Primers; Humans; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Small Interfering; Signal Transduction; Strychnine; Vascular Endothelial Growth Factor Receptor-2

Brucine is an alkaloid derived from the seeds of Strychnos nux-vomica Linn. which have long been used as a traditional medicine for the treatment of hepatocellular carcinoma (HCC) in China. HCC prognosis can be greatly influenced by metastasis. There has thus far been little research into brucine as a source of anti-metastasis activity against HCC. In this study, we revealed that brucine dramatically repressed HepG2 and SMMC-7721 HCC cell migration with few cytotoxic effects. Hypoxia inducible factor 1 (HIF-1) is a key transcription factor mediating cell migration and invasion. Brucine suppressed HIF-1-dependent luciferase activity in HepG2 cells. The transcriptions of four known HIF-1 target genes involved in HCC metastasis, i.e., fibronectin, matrix metallopeptidase 2, lysyl oxidase, and cathepsin D, were also attenuated after brucine treatment. Experiments in vivo showed that an intraperitoneal injection of 5 and 15 mg/kg of brucine resulted in dose-dependent decreases in the lung metastasis of H22 ascitic hepatoma cells. Moreover, a dosage of brucine at 15 mg/kg exhibited very low toxic effects to tumor-bearing mice. Consistently, brucine downregulated expression levels of HIF-1 responsive genes in vivo. Our current study demonstrated the capacity of brucine in suppressing HCC cell migration in vitro and lung metastasis in vivo. The inhibition of the HIF-1 pathway is implicated in the anti-metastasis activity of brucine.

Topics: Animals; Animals, Outbred Strains; Antineoplastic Agents, Phytogenic; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Movement; China; Dose-Response Relationship, Drug; Ethnopharmacology; Gene Expression Regulation, Neoplastic; Humans; Hypoxia-Inducible Factor 1; Liver Neoplasms; Lung Neoplasms; Male; Mice; Neoplasm Proteins; Random Allocation; Seeds; Strychnine; Strychnos nux-vomica; Xenograft Model Antitumor Assays

We evaluated the effects of brucine on N-nitrosodiethylamine (DENA)-induced hepatocarcinogenesis in rats. Initiation of hepatocarcinogenesis was done by intraperitoneal injection of diethylnitrosamine (DENA) followed by promotion with phenobarbital. The rats were exposed to dietary brucine for 4 weeks prior to initiation, and the treatment was continued for 22 consecutive weeks. Brucine decreased the incidence, total number, multiplicity, size and volume of preneoplastic hepatic nodules in a dose-dependent manner. Administration of DENA induced hepatocellular carcinoma (HCC), as evidenced by changes in histopathological architecture, increased activity of cytochrome P450, decreased activity of glutathione Stransferase (GST) as well as decreased antioxidant status, enhanced lipid peroxidation, increased liver marker enzymes. Western blot analysis showed decreased expression of cyclin D1 and Bcl-2 with activation of caspase-3 and increased expression of Bax. Immunohistochemical demonstrated the decreased expression of the PCNA and VEGF. These results indicate that brucine prevents lipid peroxidation and hepatic cell damage and also protects the antioxidant system in DENA-induced hepatocarcinogenesis.

Topics: Alkylating Agents; Animals; Anticarcinogenic Agents; Apoptosis; bcl-2-Associated X Protein; Caspase 3; Cyclin D1; Diethylnitrosamine; Female; Gene Expression Regulation; Immunohistochemistry; Lipid Peroxidation; Liver Neoplasms, Experimental; Phenobarbital; Proliferating Cell Nuclear Antigen; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Strychnine; Vascular Endothelial Growth Factor A

To evaluate the potential of hyaluronic acid (HA)-coated bovine serum albumin nanoparticles (BSANPs) as a novel chondrocyte-targeting drug-delivery nanomedicine.. The HA-BSANPs were characterized by dynamic light scattering, transmission electron microscopy, differential scanning calorimetry, and X-ray diffraction. Fluorescence imaging was used to visualize the distribution of nanoparticles after intra-articular injection. The chondrocyte-targeting efficiency and cellular uptake mechanism of HA-BSANPs were investigated using endocytic inhibitors.. HA-BSANPs were successfully prepared with HA coating the surface and amorphous drug in the core. Compared with BSANPs, HA-BSANPs exhibited improved uptake by chondrocytes through a receptor-mediated active uptake mechanism. The endocytosis process of BSANPs and HA-BSANPs involved clathrin-mediated endocytosis, caveolae-mediated endocytosis, and macropinocytosis. No apparent thickening or hyperplasia of the synovium was observed in either BSANPs or HA-BSANPs. The HA-BSANPs could reside in the articular cavity of rats for more than 14 days, which was significantly longer than BSANPs.. HA-BSANPs are a promising carrier for articular-related diseases due to elongated articular residence and improved chondrocytic accumulation.

Topics: Adjuvants, Immunologic; Animals; Cells, Cultured; Chondrocytes; Hyaluronic Acid; Injections, Intra-Articular; Joints; Male; Nanocapsules; Rabbits; Rats, Sprague-Dawley; Serum Albumin, Bovine; Strychnine; Tissue Distribution

A mixed matrix membrane (MMM), based on carbon nanotubes (CNTs) and hollow fiber (HF), was prepared and combined with solid phase microextraction (SPME) mode to determine strychnine and brucine in urine. This MMM was prepared by dispersing CNTs in water via surfactant assistance, and then immobilizing CNTs into the pores of HF by capillary forces and sonification. The prepared carbon nanotubes reinforced hollow fiber (CNTs-HF) was subsequently wetted by a few microliters of organic solvent (1-octanol), and then applied to extract the target analytes in direct immersion sampling mode. After extraction, analytes were desorbed via ultrasonic-assisted effect, and then detected via high-performance liquid chromatography (HPLC). To achieve the highest extraction efficiency, main extraction parameters such as the type and amount of surfactant, the diameter and doping level of CNTs, extraction time, desorption condition, pH value, stirring rate and volume of the donor phase were optimized. Under the optimum extraction conditions, the method showed good linearity ranges with correlation coefficients higher than 0.9990, good repeatability and batch-to-batch reproducibility with relative standard deviations (RSDs) less than 6% and 5% for strychnine and brucine, respectively, and low limits of detection (0.7 and 0.9 µg L(-1) for strychnine and brucine, respectively). The recoveries were in the range of 83.81-116.14% at three spiked levels. The developed method was successfully applied to real urine sample with mean relative recoveries of 94.28% and 91.30% for strychnine and brucine, respectively. The developed method shows comparable results against reference methods and is a simple, green, and cost-effective microextraction technique.

Topics: 1-Octanol; Chromatography, High Pressure Liquid; Humans; Hydrogen-Ion Concentration; Limit of Detection; Membranes, Artificial; Nanotubes, Carbon; Reproducibility of Results; Solid Phase Microextraction; Strychnine

The aim of this study was to investigate the mechanism of the apoptotic effect of brucine on human multiple myeloma (MM) cells. U266 cells (5x104) were plated in the presence or absence of brucine (0, 0.05, 0.1, 0.2 and 0.4 mg/ml) in 96‑well culture plates for 24‑72 h. The anti-proliferative response to brucine was assessed by MTT assay. Analysis of the cell cycle of U266 cells treated with or without brucine was performed using flow cytometry. The expression change of c-Jun following treatment with brucine or brucine plus the c-Jun N-terminal kinase (JNK)-specific inhibitor SP600125 was detected using RT-PCR. Brucine appeared to have an effect on apoptosis in a dose- and time‑dependent manner. Cell cycle analysis using flow cytometry revealed the accumulation of cells at the sub-G0/G1 phase. The apoptotic rates were 4.137, 10.55, 12.31, 27.67 and 29.67% (0, 0.05, 0.1, 0.2, 0.4 mg/ml brucine, respectively; P<0.01). The gray scale values were 0.7961 ± 0.007 and 0.4683 ± 0.003 (mRNA expression of c-Jun of U266 cells with or without SP600125, respectively). Concentrations of ≤ 0.4 mg/ml brucine induced apoptosis in U266 cells. Thus, brucine‑induced apoptosis in U266 cells occurs via the JNK signaling pathway and phosphorylation of c-Jun.

Topics: Adjuvants, Immunologic; Anthracenes; Apoptosis; Caspase 3; Cell Line, Tumor; G1 Phase Cell Cycle Checkpoints; Humans; JNK Mitogen-Activated Protein Kinases; Multiple Myeloma; Phosphorylation; Signal Transduction; Strychnine

In this report, the in vitro metabolism of Strychnos alkaloids was investigated using liquid chromatography/high-resolution mass spectrometry for the first time. Strychnine and brucine were selected as model compounds to determine the universal biotransformations of the Strychnos alkaloids in rat liver microsomes. The incubation mixtures were separated by a bidentate-C18 column, and then analyzed by on-line ion trap/time-of-flight mass spectrometry. With the assistance of mass defect filtering technique, full-scan accurate mass datasets were processed for the discovery of the related metabolites. The structural elucidations of these metabolites were achieved by comparing the changes in accurate molecular masses, calculating chemical component using Formula Predictor software and defining sites of biotransformation based upon accurate MS(n) spectral information. As a result, 31 metabolites were identified, of which 26 metabolites were reported for the first time. These biotransformations included hydroxylation, N-oxidation, epoxidation, methylation, dehydrogenation, de-methoxylation, O-demethylation, as well as hydrolysis reactions.

Topics: Alkaloids; Animals; Biotransformation; Chromatography, High Pressure Liquid; Male; Microsomes, Liver; Rats; Rats, Sprague-Dawley; Spectrometry, Mass, Electrospray Ionization; Strychnine; Strychnos

In this study, we investigated the mechanism of brucine in tumor angiogenesis. We found that brucine inhibits VEGF-induced cell proliferation, chemotactic motility, and the formation of capillary-like structures in HUVECs in a dose-dependent manner. Brucine suppresses VEGF- induced p-VEGFR2 kinase activity and inhibits neovascularization in vivo. Brucine inhibits the downstream protein kinases of VEGFR2, including Src, FAK, ERK, AKT and mTOR. And further downregulates levels of VEGF, NO, IL-6, IL-8, TNF-α and IFN-γ in HUVECs. Taken together, our study suggests that brucine potently suppresses angiogenesis by targeting VEGFR2 activation and may be a viable drug candidate in anti-angiogenesis and anti-cancer therapies.

Topics: Angiogenesis Inhibitors; Animals; Carcinoma, Ehrlich Tumor; Cell Proliferation; Cell Survival; Cells, Cultured; Chemotaxis; Cytokines; Dose-Response Relationship, Drug; Female; Gene Expression Regulation; Human Umbilical Vein Endothelial Cells; Humans; Male; Mice; Neovascularization, Physiologic; Nitric Oxide; Phosphorylation; Plants, Medicinal; Protein Kinase Inhibitors; Rats; Rats, Sprague-Dawley; RNA, Messenger; Signal Transduction; Strychnine; Strychnos nux-vomica; Time Factors; Tissue Culture Techniques; Tumor Burden; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2

Strychnos nux vomica Linn.(Loganaceae) commonly known as Nux vomica (Kupeelu), is a poisonous plant and its seeds are used widely in Ayurvedic system of medicine since time immemorial. Ayurveda advocates that nux vomica seeds are to be administered in therapeutics only after going through certain purificatory measures (Shodhana). There are more than six media: cow's urine (Go mutra), cow's milk (Go dugdha), cow's ghee (Go ghrita), Kanji (thin gruel), castor oil (Eranda taila) and fresh ginger juice (Ardraka swarasa) etc., which have been reported in different classical texts of Ayurveda for proper processing of nux vomica seeds. In this study, an attempt has been made to purify the seeds by using three different methods as described in ancient treatise by using cow's urine and cow's milk as media alone and together. This study revealed that all the methods studied reduced the toxicity of strychnine and brucine contents in comparison to the raw seeds as determined by HPTLC. Out of these three methods maximum reduction in strychnine and brucine contents was found when the seeds were purified by keeping them in cow's urine for seven days followed by boiling in cow's milk for three hrs.

Topics: Animals; Cattle; Chromatography, High Pressure Liquid; Drug Compounding; History, Ancient; Medicine, Ayurvedic; Milk; Plant Preparations; Seeds; Strychnine; Strychnos; Urine

The features of brucine (BC) binding to two blood proteins, bovine hemoglobin (BHb), and bovine serum albumin (BSA), were investigated via fluorescence, circular dichroism and UV/Vis absorption spectroscopy. The results revealed that BC caused the fluorescence quenching of blood proteins by the formation of BC-protein complex. The corresponding thermodynamic parameters were measured at different temperatures. The process of binding BC molecule on protein was a spontaneous molecular interaction procedure in which entropy increased and Gibbs free energy decreased. Hydrophobic and electrostatic interactions play a major role in stabilizing the complex. The molecular docking has been employed to explore the binding site of the BC in BHb and BSA on the Autodock 4.2. The distances r between BC and protein were calculated to be 4.93 and 5.08 nm for BHb, and BSA, respectively. The effect of BC on the conformation of blood proteins was analyzed using CD, synchronous fluorescence and three-dimensional fluorescence spectra.

Topics: Animals; Binding Sites; Blood Proteins; Cattle; Circular Dichroism; Energy Transfer; Hemoglobins; Hydrogen-Ion Concentration; Kinetics; Protein Structure, Secondary; Serum Albumin, Bovine; Spectrometry, Fluorescence; Spectrophotometry, Ultraviolet; Strychnine; Thermodynamics

Hepatocellular carcinoma is difficult to diagnose early, and most patients are already in the late stages of the disease when they are admitted to hospital. The total 5-year survival rate is less than 5%. Recent studies have showed that brucine has a good anti-tumor effect, but high toxicity, poor water solubility, short half-life, narrow therapeutic window, and a toxic dose that is close to the therapeutic dose, which all limit its clinical application. This study evaluated the effects of brucine immuno-nanoparticles (BIN) on hepatocellular carcinoma.. Anionic polymerization, chemical modification technology, and phacoemulsification technology were used to prepare a carboxylated polyethylene glycol-polylactic acid copolymer carrier material. Chemical coupling technology was utilized to develop antihuman AFP McAb-polyethylene glycol-polylactic acid copolymer BIN. The size, shape, zeta potential, drug loading, encapsulation efficiency, and release of these immune-nanoparticles were studied in vitro. The targeting, and growth, invasion, and metastasis inhibitory effects of this treatment on liver cancer SMMC-7721 cells were tested.. BIN were of uniform size with an average particle size of 249 ± 77 nm and zeta potential of -18.7 ± 4.19 mV. The encapsulation efficiency was 76.0% ± 2.3% and the drug load was 5.6% ± 0.2%. Complete uptake and even distribution around the liver cancer cell membrane were observed.. BIN had even size distribution, was stable, and had a slow-releasing effect. BIN targeted the cell membrane of the liver cancer cell SMMC-7721 and significantly inhibited the growth, adhesion, invasion, and metastasis of SMMC-7721 cells. As a novel drug carrier system, BIN are a potentially promising targeting treatment for liver cancer.

Topics: Antibodies, Monoclonal; Antineoplastic Agents; Carcinoma, Hepatocellular; Cell Adhesion; Cell Line, Tumor; Cell Movement; Cell Proliferation; Humans; Immunotoxins; Liver Neoplasms; Nanoparticles; Particle Size; Strychnine

A liquid chromatography-electrospray ionization-ion trap mass spectrometry (LC-ESI-ITMS) method was developed for the simultaneous analysis of strychnine, brucine and their major metabolites. Strychnine and brucine were individually incubated with rat liver S9 fraction. The incubation samples were pooled together and analyzed with LC-ESI-ITMS in positive ion and full-scan detection mode. The calibration curves of strychnine and brucine in rat liver showed good linearity in ranges of 0.020 to 8.0 µg/mL for strychnine and 0.020 to 8.5 µg/mL for brucine. The limits of detections were both 0.008 µg/mL and the recoveries were 88.3 and 83.2% for strychnine and brucine, respectively. Two metabolites were identified as strychnine N-oxide and brucine N-oxide by comparing the molecular mass, retention time, full-scan mass spectra, tandem MS and MS(3) spectra with those of strychnine and brucine. The developed method provided high sensitivity and selectivity for the determination of poisonous alkaloids and their major metabolites and can be applied in the determination of samples in forensic and clinically toxicological cases.

Topics: Analgesics; Animals; Cyclic N-Oxides; Forensic Toxicology; Limit of Detection; Liver; Male; Poisons; Rats; Rats, Sprague-Dawley; Ribosomal Protein S9; Ribosomal Proteins; Spectrometry, Mass, Electrospray Ionization; Strychnine; Substance Abuse Detection; Tandem Mass Spectrometry

The aim of this study was to explore the effects of brucine on bone metabolism in multiple myeloma (MM) and to compare brucine and bortezomib regarding the effects on MM in vitro. The half maximal inhibitory concentration (IC50) values of brucine and bortezomib in the MM cell line U266 were detected by MTT assay. In addition, the expression of alkaline phosphatase (ALP), osteocalcin (OC), osteoprotegerin (OPG) and osteoprotegerin ligand (also termed receptor activator of nuclear factor κB ligand) (RANKL) at mRNA levels were measured by RT-PCR. IC50 of bortezomib in the U266 cell line at 48 h was 22.4 nmol/l, and that of brucine was 0.16 nmol/l. Compared with osteoblasts incubated with MM cell supernatant alone, the mRNA levels of ALP, OC and OPG in osteoblasts co-treated with brucine and MM cell supernatant were higher (p<0.05), while the mRNA expression of RANKL was lower, and the ranges of the changes were all larger than those of the group treated with bortezomib (P<0.05). Brucine exerts effects on bone metabolism in multiple myeloma through the regulation of osteoclasts by osteoblasts.

Topics: Alkaline Phosphatase; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Bone and Bones; Boronic Acids; Bortezomib; Cell Differentiation; Cell Line, Tumor; Culture Media; Humans; Inhibitory Concentration 50; Multiple Myeloma; Osteoblasts; Osteocalcin; Osteoclasts; Osteoprotegerin; Polymerase Chain Reaction; Pyrazines; RANK Ligand; RNA, Messenger; RNA, Neoplasm; Stem Cells; Strychnine

A sensitive liquid chromatography-tandem mass spectrometric (LC-MS/MS) method has been developed and validated for the determination of brucine and strychnine in rat plasma.. Samples were extracted by ethyl acetate-n-butanol (7: 3). Chromatographic separation was operated on ZORBAX XDB-C18 column with gradient elution of acetonitrile-methanol-water (0.05% acetic acid and 10 nmol x L(-1) ammonium formate contained), followed by LC-MS/MS in positive electrospray ionization. Quantification was carried out on multiple reaction monitoring (MRM) of the transition m/z 395.2/324.2, m/z 335.2/184.2 and m/z 199.1/171.1 for brucine, strychnine and tacrine (internal standard), respectively.. The method was linear in the range of 0.195-100 and 0.07840 microg x L(-1) for brucine and strychnine, with coefficient correlation 0.994 and 0.996 respectively. The recoveries of extraction were 78.9% - 102.4% for brucine and 95.2% - 106.1% for strychnine. Precision, accuracy, stability and matrix effect of the analytes met the requirement. The method was applied to a pharmacokinetic study of brucine and strychnine after cutaneous administration of Semen Strychni niosome gel. The C(max) were (26.20 +/- 5.81) and (12.50 +/- 3.00) microg x L(-1) while the AUC(0-infinity), were (193.75 +/- 39.43) and (98.25 +/- 28.54) microg x h x L(-1) of the two components.. We conclude that the niosomes may reduce the systemic exposures and prolong the local release of brucine and strychnine.

Topics: Administration, Cutaneous; Analgesics; Animals; Chromatography, Liquid; Convulsants; Female; Gels; Liposomes; Male; Plants, Medicinal; Rats; Rats, Sprague-Dawley; Seeds; Specific Pathogen-Free Organisms; Strychnine; Strychnos nux-vomica; Tandem Mass Spectrometry

Brucine is a central agonist that can pass through the blood-brain barrier (BBB). The goal of this study is to examine whether brucine is one of the substrates of the drug transporter P-glycoprotein (P-gp) and to examine the effects of P-gp on the brucine transport at the in vitro BBB model. The P-gp ATPase assay was utilized to investigate the in vitro affinity of P-gp to brucine. Results suggested that K(m) of brucine (11.4 μmol/l) was smaller than the positive control, verapamil (16.4 μmol/l). In this study, we developed an in vitro BBB model, comprising a co-culture of primary rat brain microvessel endothelial cells and astrocytes for the transport study. The validated model was correct and available. Transendothelial electrical resistance reached (283.78 ± 18.85) Ω cm(2). The model displayed limited permeability to fluorescein sodium and [(125)I]albumin, with the apparent permeability coefficient Papp of (10.36 ± 0.86) × 10(-6) cm/s and (6.00 ± 0.78) × 10(-6)cm/s, respectively. The quantity of the bidirectional transport of brucine was determined by ultra-performance liquid chromatography-tandem mass spectrometry. In the absence of verapamil, the transport of brucine from basolateral compartment to apical compartment (BL-AP) was higher than from AP to BL at low, middle, and high concentrations (P<0.05). The excretion rate was 1.32, 1.56, and 1.54, respectively. However, following exposure to verapamil, the excretion rate at three different concentrations was decreased (P<0.05). All the results suggest that P-gp prevented brucine from passing through the in vitro BBB model.

Topics: Animals; Astrocytes; ATP Binding Cassette Transporter, Subfamily B, Member 1; Biological Transport; Blood-Brain Barrier; Endothelial Cells; Rats; Rats, Sprague-Dawley; Reproducibility of Results; Rhodamine 123; Strychnine

To study the molecular mechanism of cyclooxygenase-2 (COX-2), one of effective ingredient of brucine, in inducing non-small cell lung cancer cell apoptosis.. COX-2 promoter, transcription factor deletion mutants and COX-2 mRNA 3'-UTR-containing report plasmids were transfected with Renillia to non-small cell lung cancer A549 cell, in order to detect the activity of report gene luciferase and minimum cis-acting element of COX-2 promoter inhibited by brucine. The influence of brucine on IkappaB phosphorylation and the nuclear translocation of p65 were detected by immunoblotting assay.. Brucine significantly suppressed LPS-induced COX-2 promoter activation, but revealed minor impact on COX-2 mRNA stability. NF-kappaB in the vicinity of COX-2 promoter-262 was an important cis-acting element of brucine for inhibiting the activity of COX-2 promoter. Brucine was found to inhibit the phosphorylation of IkappaBalpha as well as the nuclear translocation of p65.. Brucine can improve A549 cells apoptosis by inhibiting the activity of NF-kappaB and the subsequent COX-2 gene expression.

Topics: Biological Transport; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cyclooxygenase 2; Humans; NF-kappa B; Phosphorylation; Promoter Regions, Genetic; RNA Stability; Strychnine

The aim of this study was to develop a sustained release converse thermosensitive hydrogel for intra-articular injection using chitosan-glycerol-borax as matrix, its physical properties and biocompatibility were investigated. Taking gelation time and gelation condition as index, the influence of concentration of chitosan, ratio of chitosan to glycerol, pH on physical properties of hydrogel were investigated. And then the in vitro drug release, rheological properties and biocompatibility were studied. The thermosensitive hydrogel flows easily at room temperature and turns to gelation at body temperature, which can certainly prolong the release of drug and has good biocompatibility.

Topics: Analgesics; Animals; Chitosan; Delayed-Action Preparations; Drug Compounding; Hydrogels; Hydrogen-Ion Concentration; Inflammation; Injections, Intra-Articular; Knee Joint; Male; Materials Testing; Plants, Medicinal; Rats; Rats, Sprague-Dawley; Rheology; Seeds; Strychnine; Strychnos nux-vomica; Surface Properties; Synaptic Membranes; Temperature

To study the effects of brucine on vascular endothelial growth factor (VEGF) expression and microvessel density (MVD) in a nude mouse model of bone metastasis due to breast cancer, and to assess the possible antitumor mechanism of brucine.. A syringe needle was used to directly inject 0.2 mL monoplast suspension (with 2×10(5) human breast cancer cells contained) into the bony femoral cortex of the right hind leg for modeling. Twenty-five nude mice were randomized into five groups and administered with an intraperitoneal injection of saline or drug for 8 consecutive days: model group (0.2 mL normal saline), low-dose brucine group (1.73 mg·kg(-1)), medium-dose brucine group (3.45 mg·kg(-1)), high-dose brucine group (6.90 mg·kg(-1)), and thalidomide group (200 mg·kg(-1)). Diet and activity were recorded, and the tumors were harvested 5 weeks later. The percentage of VEGF-positive cells was determined with hematoxylin and eosin staining and immunohistochemical staining, and MVD expression was determined by optical microscopy.. The VEGF expressions in brucine- or thalidomide-treated mice were significantly reduced as compared with mice in the model group (P <0.01). There were no significant difference between the high-dose brucine group and the thalidomide group (P >0.05). Significant difference was between the high- and low-dose brucine group P<0.05). Further, VEGF expression was significantly increased in the low- and medium-dose brucine groups compared with the thalidomide group (P <0.05). The MVD values in the three brucine and thalidomide groups were significantly lower than that in the model group (P <0.01). The MVD values in the medium- and high-dose brucine groups were not significantly different from those in the thalidomide group (P >0.05), while the MVD value showed a significant increase in the low-dose group compared with the thalidomide group (P <0.05).. Brucine could inhibit the growth of breast cancer to bone metastases, possibly by inhibiting tumor angiogenesis.

Topics: Animals; Bone Neoplasms; Breast Neoplasms; Cell Line, Tumor; Disease Models, Animal; Female; Humans; Immunohistochemistry; Mice; Mice, Inbred BALB C; Mice, Nude; Microvessels; Strychnine; Vascular Endothelial Growth Factor A; Xenograft Model Antitumor Assays

Brucine was encapsulated into stealth liposomes using the ammonium sulfate gradient method to improve therapeutic index.. Four brucine stealth liposomal formulations were prepared, which were made from different phosphatidylcholines (PCs) with different phase transition temperatures (T(m)). The PCs used were soy phosphatidylcholine (SPC), dipalmitoyl phosphatidylcholine (DPPC), hydrogenated soy phosphatidylcholine (HSPC), and distearoyl phosphatidylcholine (DSPC). The stabilities, pharmacokinetics, and toxicities of these liposomal formulations were evaluated and compared.. Size, zeta potential, and entrapment efficiency of brucine-loaded stealth liposomes (BSL) were not influenced by PC composition. In vitro release studies revealed that drug release rate increased with decreased T(m) of PCs, especially with the presence of rat plasma. After intravenous administration, the area under the curve (AUC) values of BSL-SPC, BSL-DPPC, BSL-HSPC, and BSL-DSPC in plasma were 7.71, 9.24, 53.83, and 56.83-fold as large as that of free brucine, respectively. The LD(50) values of brucine solution, BSL-SPC, BSL-DPPC, BSL-HSPC, and BSL-DSPC following intravenous injection were 13.17, 37.30, 37.69, 51.18, and 52.86 mg/kg, respectively. It was found in calcein retention experiments that the order of calcein retention in rat plasma was SPC < DPPC << HSPC < DSPC stealth liposomes.. PC composition could exert significant influence on the stabilities, pharmacokinetics, and toxicities of brucine-loaded stealth liposomes. DSPC or HSPC with T(m) above 50°C should be used to prepare the stealth liposomal formulation for the intravenous delivery of brucine. However, it was found in the present paper that the pharmacokinetics and toxicity of BSL were not influenced by the PC composition when the T(m) of the PC was in the range of -20°C to 41°C.

Topics: Animals; Area Under Curve; Cell Membrane Permeability; Chromatography, High Pressure Liquid; Lethal Dose 50; Liposomes; Male; Mice; Phosphatidylcholines; Rats; Rats, Sprague-Dawley; Strychnine; Temperature; Toxicity Tests, Acute

Intra-articular drug delivery system could directly deliver a drug to an affected joint and offer the possibility of reaching high drug concentrations at the site of action with limited systemic toxicity. However, depending on their chemical structure, some active compounds were rapidly cleared from the joint, thus requiring numerous injections, which could cause infection or joint disability. To control the release behavior for prolonged time periods, a novel biologically based drug delivery vehicle was designed for intra-articular using microsphere/thermally responsive hydrogel combination system in this paper. And brucine was the test drug. The system was constructed by dispersing the brucine microspheres which was prepared by using a spray-drying method in a thermally responsive biopolymer hydrogel contained with chitosan-glycerol-borax. The microspheres were spherical as evidenced by the scanning electron microscopy (SEM) photographs. And the entrapment rate was 98.60% w/w with an average size range of 0.9-4.5 μm. Fourier transforms infrared (FT-IR) spectroscopy and X-ray diffraction (XRD) revealed the absence of drug-polymer interaction and amorphous nature of an entrapped drug. From the in vitro drug release study we could see that there was a burst release of microsphere, which was obviously retarded when dispersed in hydrogel. And the studies of biocompatibility with synovium showed that no apparent thickening or hyperplasia of the synovium, a small quality of phlogocyte imbibitions was observed. The results of FX imaging in rats showed that by intra-articular injection the BMH could stay in articular for over 7 days were consistent with our in vitro release. And the results of pharmacodynamics revealed the BMH could benefit OA joint by suppressing the levels of TNF-α and IL-1β, protect the damaged joint from degradation. The novel microsphere/thermoresponsive hydrogel combination system could be a promising treatment option for OA and RA. In conclusion, the system appears to be generally biocompatible with synovium and could control the drug release for several days; hence it might be suitable for the development of treatment strategies for rheumatic diseases.

Topics: Analgesics; Animals; Chitosan; Hydrogels; Injections, Intra-Articular; Interleukin-1beta; Male; Microspheres; Osteoarthritis; Particle Size; Rabbits; Rats; Rats, Sprague-Dawley; Spectroscopy, Fourier Transform Infrared; Strychnine; Surface Properties; Synovial Fluid; Temperature; Tumor Necrosis Factor-alpha; X-Ray Diffraction

The latent fingermarks developed with iodine fumes are not permanent due to the sublimation of iodine. In this paper a new method has been proposed to fix the latent fingermarks developed with iodine fumes. Latent fingermarks were developed on both porous as well as non-porous substrates which were subsequently fixed by treating them with brucine based reagent. The fingermarks fixed with this method were not only permanent but also without any background coloration. Adsorption of the brucine over the iodine has been found to be a possible reason for permanency of iodine developed fingermarks. This research has successfully demonstrated the first use of the brucine solution for fixing latent fingermarks on porous and as well as on non porous substrates.

Topics: Dermatoglyphics; Forensic Medicine; Humans; Indicators and Reagents; Iodine; Strychnine; Surface Properties; Volatilization

To study different in vivo pharmacokinetic regularity of brucine, total alkaloids of scorched sand-prepared Strychni Semen products and Strychni Semen pulveratum in rats, and probe into mutual impact between single component and compound.. Rats in each group were orally administered with brucine, total alkaloids of scorched sand-prepared Strychni Semen products and Strychni Semen pulveratum suspension. The in vivo plasma concentrations of brucine in rats were determined by HPLC. A compartment model was made for the blood drug concentration-time curve using 3P97 software package and the pharmacokinetic parameters of each group were calculated and compared.. The in vivo metabolic process of brucine in rats complied with the two-compartment model, weight W = 1/C2. The results of variance analysis showed that among three existing forms of brucine with same dosage, the brucine solution group and the total alkaloids group of scorched sand-prepared Strychni Semen products showed significant differences in C(max), MRT (P < 0.05); and the brucine solution group and the Strychni Semen pulveratum suspension group showed significant differences in C(max), AUC(0-t), and AUC(0-infinity), in which the latter displayed minimum C(max), AUC(0-t) and AUC(0-infinity).. The total alkaloids group of scorched sand-prepared Strychni Semen products showed a relatively longer retention time of effective components of brucine in plasma, while the Strychni Semen pulveratum suspension group showed a lower bioavailability.

Topics: Animals; Male; Medicine, Chinese Traditional; Plants, Medicinal; Rats; Rats, Sprague-Dawley; Strychnine

To establish a method for determination of strychnine and brucine in formaldehyde fixed tissue by LC-MS/MS analysis.. The samples were pretreated with solid phase extraction using SCX cartridges and separated on SB-C18 column with mobile phase 0.1% formic acid : 0.1% formic acid-acetonitrile (75:25). Electrospray ionization (ESI) source was utilized and operated in positive ion mode. Multiple reactions monitoring (MRM) mode was applied. External standard method was applied for quantitation.. The chromatographic separation of strychnine and brucine in formaldehyde fixed nephritic and hepatic tissues resulted successfully. The standard curve was linear in the range of 0.002-2.0 microg/g for strychnine and brucine in formaldehyde fixed tissues, and the correlation coefficient was more than 0.996. The limits of detection (LOD) of strychnine and brucine in nephritic tissues were 0.06ng/g and 0.03 ng/g, respectively. The LOD of both chemicals were 0.3 ng/g in hepatic tissues. The extraction recovery rate was more than 74.5%. The precision of intra-day and inter-day were both less than 8.2%.. Strychnine and brucine can be sensitive to be determined in formaldehyde fixed tissue by LC-MS/MS analysis. It can be applied in the forensic toxicological analysis.

Topics: Chromatography, Liquid; Forensic Toxicology; Formaldehyde; Formates; Kidney; Limit of Detection; Liver; Mass Spectrometry; Molecular Structure; Reproducibility of Results; Spectrometry, Mass, Electrospray Ionization; Strychnine; Tandem Mass Spectrometry; Tissue Distribution

To investigate the effect of dose on pharmacokinetic properties of brucine hydrogel patch.. The plasma concentration of brucine was determined by HPLC. Brucine hydrogel patch was prepared and its pharmaceutical characterization was investigated. After transdermal administration of different dose brucine hydrogel patch; Plasma concentration versus time profiles were determined and pharmacokinetic parameters were calculated by DAS program.. The pharmaceutical properties of brucine hydrogel patch were satisfactory. The AUC0-1 values were 7.24 +/- 0.61, 16.02 +/- 2.34 and 54.84 +/- 26.59 microg x h/mL after administration of 30, 60 and 180 mg/kg brucine hydrogel patch, respectively. The corresponding C(max) values were 0.73 +/- 0.23, 1.45 +/- 0.28 and 4.59 +/- 1.85 microg/mL, respectively. And the corresponding T(max) values were 8.67 +/- 2.07, 11.67 +/- 2.66 and 8.33 +/- 2.65 h, respectively.. The pharmacokinetic properties of brucine do not vary with the dose of brucine hydrogel patch.

Topics: Animals; Area Under Curve; Carboxymethylcellulose Sodium; Chromatography, High Pressure Liquid; Female; Hydrogels; Male; Povidone; Random Allocation; Rats; Rats, Sprague-Dawley; Strychnine; Strychnos nux-vomica; Transdermal Patch

A sensitive method for identifying and quantifying brucine by means of liquid chromatography-tandem mass spectrometry is presented in this article. Based on a solid-phase extraction for human serum, the validation indicated limits of detection and quantification of 0.12 and 0.23 ng/mL, respectively. In one case of lethal suicidal brucine monointoxication, brucine concentrations of 1.51 μg/mL, 1.69 μg/mL, 9.94 μg/mL, 16.4 μg/g, 0.99 μg/g, 0.75 μg/g, and 1.95 mg/g were determined in femoral blood, urine, bile collected from the gallbladder, liver tissue, cerebellum, cerebrum, and stomach contents, respectively.

Topics: Diagnosis; Fatal Outcome; Female; Humans; Limit of Detection; Male; Middle Aged; Poisoning; Poisons; Solid Phase Extraction; Strychnine; Suicide

This study was aimed to explore the influence of brucine on the early differentiation of osteoblasts and the metabolic pathway of osteoclast in multiple myeloma (MM) and to compare the effects of brucine and bortezomib on MM. The half inhibitory concentration (IC(50)) of brucine and bortezomib on MM cell line U266 was determined by MTT method; the mRNA levels of alkaline phosphatase (ALP), osteocalcin (OC), osteoprotegerin (OPG) and osteoprotegerin ligand (RANKL) were detected by RT-PCR after the supernatant of cultured U266 cells was added into the culture system for inducing the differentiation of osteoblast line MC3T3-E1 and culturing. The results showed that the IC(50)of bortezomib and brucine on U266 cells for 48 hours were 22.4 nmol/L and 0.16 mg/ml respectively. As compared with osteoblasts treated by supernatant of cultured MM cells alone, the mRNA levels of ALP, OC and OPG in osteoblasts treated by brucine combined with supernatant of cultured MM cells were enhanced (p < 0.05), while the RANKL mRNA level was lowered (p < 0.05), moreover the enhanced and lowered degree also was large (p < 0.05). It is concluded that the influence of brucine on metabolism of osteoblasts and osteoclasts in MM may be realized through the regulation of osteoclasts by osteoblasts. The therapeutic efficacy of brucine on MM is superior to bortezomib.

Topics: Animals; Cell Line; Cell Line, Tumor; Humans; Inhibitory Concentration 50; Mice; Multiple Myeloma; Osteoblasts; Osteoclasts; Strychnine

The aim of this study was to investigate the effect of brucine on secretion of TNF-α, IFN-γ, IL-4 and proliferation of T lymphocytes in patients with aplastic anemia (AA), and to explore its mechanism. Peripheral blood T lymphocytes from 10 patients with AA and 10 healthy volunteers were isolated, purified and cultured. T lymphocytes from the patients were divided into 0, 100, 200 and 400 µg/ml brucine-treated groups. T lymphocytes from healthy volunteers were used as control group. After being cultured for 72 hours, the levels of TNF-α, IFN-γ, IL-4 in the supernatant of cultured T lymphocytes from AA patients were detected by ELISA, and the proliferation of T lymphocytes from AA patients was detected by MTT. The results showed that compared with the normal control group, the levels of TNF-α and IFN-γ in the culture supernatant significantly increased, and IL-4 was significantly decreased. The levels of TNF-α and IFN-γ in the culture supernatant of brucine treated groups were lower, and were dependent on the concentration of brucine. However, the levels of IL-4 were found to be not obviously changed. The inhibition rate of T lymphocytes in 100, 200 and 400µg/ml brucine-treated groups were (13.61 ± 4.31)%, (14.28 ± 4.31)% and (15.12 ± 4.56)% respectively, among which the differences were not statistically significant. It is concluded that the brucine can reduce the levels of TNF-α and IFN-γ through inhibiting the proliferation of T lymphocytes in AA patients, which provides experimental basis for therapy of AA patients.

Topics: Adolescent; Adult; Anemia, Aplastic; Case-Control Studies; Cell Proliferation; Cells, Cultured; Child; Female; Humans; Interferon-gamma; Interleukin-4; Male; Middle Aged; Strychnine; T-Lymphocytes; Tumor Necrosis Factor-alpha; Young Adult

A simple and low-cost HPLC method with UV absorbance detection was developed and validated to simultaneously determine strychnine and brucine, the most abundant alkaloids in the processed Semen Strychni, in rat tissues (kidney, liver, spleen, lung, heart, stomach, small intestine, brain and plasma). The tissue samples were treated with a simple liquid-liquid extraction prior to HPLC. The LOQs were in the range of 0.039-0.050 µg/ml for different tissue or plasma samples. The extraction recoveries varied from 71.63 to 98.79%. The linear range was 0.05-2 µg/ml with correlation coefficient of over 0.991. The intra- and inter-day precision was less than 15%. Then the method was used to measure the tissue distribution of strychnine and brucine after intravenous administration of 1 mg/kg crude alkaloids fraction (CAF) extracted from the processed Semen Strychni. The results revealed that strychnine and brucine possessed similar tissue distribution characterization. The highest level was observed in kidney, while the lowest level was found in brain. It was indicated that kidney might be the primary excretion organ of prototype strychnine and brucine. It was also deduced that strychnine and brucine had difficulty in crossing the blood-brain barrier. Furthermore, no long-term accumulation of strychnine and brucine was found in rat tissues.

Topics: Animals; Brain; Chromatography, High Pressure Liquid; Kidney; Liver; Male; Plants, Medicinal; Rats; Rats, Sprague-Dawley; Strychnine; Tissue Distribution

Brucine (BRU), a natural plant alkaloid is reported to possess cytotoxic and antiproliferative activities. In this study we aimed to investigate its in vitro and in vivo antitumor and antiangiogenic effects.. Cell proliferation and viability was assessed using microculture tetrazolium tests (MTT). As predictive markers we determined intracellular levels of vascular endothelial growth factor (VEGF), Interleukin-12 (IL-12), tumor necrosis factor (TNF-α), caspase-3, -8 and -9 by ELISA and enzymatic activity assays. In addition, anti-VEGF neutralization effect was evaluated to assess whether it could result in augmented anticancer efficacy than the single agent. Antitumor activity was evaluated against Ehrlich ascites and solid tumor models. 15×10(6) EAC cells were implanted intraperitoneally (i.p., ascites tumor) and subcutaneous (s.c., solid tumor) in Swiss albino mice. Mice with established tumors received brucine i.p. at 12.5, 25, and 50mg/kg for 14days in ascites tumor and 50mg/kg in solid tumor for 30days. Tumor volume, cell viability, angiogenic, anti-angiogenic, anti-inflammatory factors and antioxidant parameters were determined. Immunohistochemistry analysis for VEGF and CD-31 was also performed.. BRU produced time and dose-dependent inhibition of MCF-7 in vitro and EAC tumors in vivo. The anti-angiogenic effects were accompanied with decreased VEGF and TNF-α and increased IL-12 expression. BRU reduced peritoneal angiogenesis and microvessel density in vivo.. Our findings suggest that BRU possesses antitumor and anti-angiogenic activities in vitro and in vivo. The above results showed that BRU can be used as a potential anticancer agent as an antimetastatic and anti-angiogenic agent.

Topics: Animals; Antineoplastic Agents; Antioxidants; bcl-2-Associated X Protein; Blood Cell Count; Carcinoma, Ehrlich Tumor; Caspases; Cell Line, Tumor; Dose-Response Relationship, Drug; Enzyme Activation; Female; Humans; Immunohistochemistry; Interleukin-12; Mice; Neoplasm Transplantation; Neovascularization, Pathologic; Oxidative Stress; Strychnine; Survival Analysis; Tetrazolium Salts; Thiazoles; Tumor Necrosis Factor-alpha; Vascular Endothelial Growth Factor A

To evaluate the pharmacokinetics and tissue distribution of liposomal brucine (LB) for dermal application.. Pharmacokinetics and tissue distribution were studied by in vivo animal testing. High performance liquid chromatography (HPLC) was used to detect the concentration of brucine in rats' skin, plasma and various tissues.. After dermal administration, LB was absorbed rapidly in the skin and could be detected after 0.5 hours. After 36 hours, levels were too low to be detected. In plasma, levels were also too low to be detected after 36 hours. The concentration of LB reached 50% of the maximum in all tissues except the brain, peaking after 1.5 hours but still detectable after 12 hours.. The concentration of LB was high in skin at the application site. LB was quickly absorbed into tissues through the blood circulation and widely distributed throughout the whole body. There was no obvious toxicity and LB did not readily accumulate in tissues and organs. It showed local potency but low overall systemic toxicity.

Topics: Administration, Cutaneous; Animals; Chromatography, High Pressure Liquid; Female; Half-Life; Liposomes; Male; Rats; Rats, Sprague-Dawley; Skin; Strychnine; Tissue Distribution

To investigate the apoptosis-induction effect of brucine on human chronic myeloid leukemia cell line K562 cells, K562 cells were exposed to various dosages of brucine. MTT method was used to assayed the growth inhibition effect of brucine on K562 cells. The apoptosis of K562 cells was detected by acridine orange/ethidium bromide (AO/EB) double staining, Annexin-V/PI double labeling method and DNA agarose gel electrophoresis. The results showed that brucine could remarkably inhibit the K562 cell growth in a concentration-dependent and time-dependent manners at the range of 50 to 400 µg/ml, and its most significant inhibition was observed at 400 µg/ml for 72 hours and the inhibition rate was 94.0%. Staining of cells with AO-EB revealed that brucine induced nuclear chromatin condensation. After the K562 cells were treated with the brucine of 400 µg/ml for 72 hours, the most of the nucleus were orange stained and condensation-like or bead-like showing apoptotic morphology. The K562 cells treated with brucine of different concentrations (50, 100, 200, 400, 800 µg/ml) for 72 hours, Annexin-V/PI detection showed brucine could induce apoptosis of K562 cells, and apoptosis rate increased gradually with increasing concentration of drugs. The K562 cells treated with brucine of 400 µg/ml for 72 hours displayed typical ladder strap in DNA gel electrophoresis. It is concluded that brucine can efficiently inhibit cell growth and induce apoptosis of K562 cells with dose-dependent manner in concentrations of 50 - 400 µg/ml.

Topics: Apoptosis; Cell Proliferation; Humans; K562 Cells; Strychnine

To compare the pharmaceutical properties and the anti-tumor activities of three kinds of stealth liposomes prepared with different phospholipid composition containing brucine.. Stealth liposomes with different phospholipids composition, such as soybean phosphatidycholine (SPC), hydrogenated soybean phosphatidylcholine (HSPC) and the complex of SPC and HSPC, were prepared by ammonium sulfate transmembrane gradient method. Pharmaceutical properties such as shape, encapsulation efficiency and size of three stealth liposomes were compared intensively. Anti-tumor activity of SPC, HSPC and novel stealth liposomes composed of both SPC and HSPC were compared by established mouse liver cancer H22 model. Meanwhile, the mice body weight and immune organ weight were also compared.. The encapsulation efficiency of novel, SPC and HSPC stealth liposomes were 77.7%, 64.8% and 74.8%, respectively. The mean diameters of them were less than 100 nm. The tumor inhibition rate of novel, HSPC and SPC stealth liposomes were 57.88%, 49.15%, 23.37%, respectively. The mice body weight, thymus gland index of three stealth liposomes group and spleen index of novel stealth liposomes group had no significant difference with the negative group while SPC and HSPC stealth liposomes group increased the spleen index.. Phospholipids composition is the key factor which determines the antitumor activity of brucine-loaded stealth liposomes.

Topics: Animals; Antineoplastic Agents; Body Weight; Cell Line, Tumor; Cell Proliferation; Humans; Liposomes; Mice; Particle Size; Phospholipids; Strychnine

Abstract: The activities of four CYP450 enzymes (CYP3A, 1A2, 2El and 2C) and the mRNA expression levels of CYP1A2, 2El, 2Cll and 3A1 in rat liver were determined after Wistar rats were orally administered with brucine (BR) at three dosage levels (3, 15 and 60 mg.kg-1 per day) and the high dose of BR combined with glycyrrhetinic acid (GA, 25 mg.kg-1 per day) or liquiritin (LQ, 20 mg.kg-1 per day) for 7 consecutive days. Compared with the control, brucine caused 24.5% and 34.6% decrease of CYP3A-associated testosterone 6beta-hydroxylation (6betaTesto-OH) and CYP2C-associated tolbutamide hydroxylation (Tol-OH), respectively, and 146.1% increase of CYP2El-associated para-nitrophenol hydroxylation (PNP-OH) at the high dose level. On the other hand, (BR+GA) caused 51.4% and 33.5% decrease, respectively, of CYP2El-associated PNP-OH and CYP1A2-associated ethoxyresorufin-O-de-ethylation (EROD) as compared with the high dose of BR group. Meanwhile, (BR+LQ) caused 41.1% decrease of CYP2El-associated PNP-OH and 37.7% increase of CYP2C-associated Tol-OH. The results indicated that the co-administration of BR with GA or LQ had effect on mRNA expression and activities of the CYP450 enzymes mentioned above to some extent, and the in vivo antagonism of LQ on BR-induced CYPs adverse effects and the in vivo inhibitory action of GA on CYP2E1 and 1A2 might play an important role in the detoxification of Radix Glycyrrhizae against Strychnos nux-vomica L.

Topics: Animals; Aryl Hydrocarbon Hydroxylases; Cytochrome P-450 CYP1A1; Cytochrome P-450 CYP1A2; Cytochrome P-450 CYP2E1; Cytochrome P-450 CYP3A; Cytochrome P-450 Enzyme System; Cytochrome P450 Family 2; Flavanones; Gene Expression Regulation, Enzymologic; Glucosides; Glycyrrhetinic Acid; Hydroxylation; Liver; Male; Nitrophenols; Plants, Medicinal; Rats; Rats, Wistar; RNA, Messenger; Steroid 16-alpha-Hydroxylase; Steroid Hydroxylases; Strychnine; Strychnos nux-vomica; Tolbutamide

To compare the pharmacokinetic characteristics of brucine following intravenous administration of liposomes, containing total alkaloids from seed of Strychnos nux-vomica, to rats with different phospholipids composition.. Liposomes containing the total alkaloids were prepared by the method of ammonium sulfate transmembrane gradients and stealth liposome technique. The contents of total alkaloids and brucine in liposomes were determined and compared after free drug being removed. After intravenous administration of total alkaloids solution or liposomes with different composition, plasma samples were drawn at predetermined time points and the concentrations of brucine were determined by a validated method of HPLC. Pharmacokinetic analysis was performed by 3P97 program.. The ratios of brucine to total alkaloids in liposomes hardly varied with phospholipids composition. Compared with SPC liposome, AUC of brucine was increased 13.3-fold and apparent volume of distribution was decreased to only 3.6% following intravenous administration of HSPC liposome. In addition, besides that AUC of brucine was slightly increased, most pharmacokinetic parameters were not significantly changed after administration of the novel liposome compared with those of SPC liposome.. Phospholipids composition has a significant influence on the pharmacokinetics of brucine after intravenous administration of liposomes containing total alkaloids from seed of S. nux-vomica.

Topics: Alkaloids; Animals; Drugs, Chinese Herbal; Infusions, Intravenous; Liposomes; Male; Models, Animal; Rats; Rats, Sprague-Dawley; Seeds; Strychnine; Strychnos nux-vomica

A selective, simple and efficient method-ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was developed for determination of two toxic alkaloids, namely strychnine and brucine in mice plasma. The UPLC separation was carried out using a 1.7 μm BEH C(18) column (50 mm × 2.1 mm) with a mobile phase consisting of methanol:0.1% formic acid (25:75, v/v), hence providing high efficiency, high resolution and excellent peak shape for the analytes and internal standard. The method was validated over the range of 2.48-496.4 ng/ml for strychnine and 2.64-528 ng/ml for brucine, respectively. Intra- and inter-day accuracy ranged from 95.0% to 107.9% for strychnine, 93.4% to 103.3% for brucine, and the precisions were within 13.8%. The extraction recoveries of both the two alkaloids exceed 81.9%. With a simple and minor sample preparation procedure and short run-time (<3 min), the proposed method was applicable for the pharmacokinetic and toxicological analysis of strychnine and brucine in vivo.

Topics: Animals; Chromatography, High Pressure Liquid; Drug Stability; Linear Models; Male; Mice; Reproducibility of Results; Sensitivity and Specificity; Strychnine; Tandem Mass Spectrometry

The toxicity depending on both dose and administration route is the major obstacle to the development of brucine, a bioactive alkaloid from Semen Strychni. In this study, the apparent partition coefficient and plasma protein binding extent of brucine were determined. In addition, the dose-dependency of the pharmacokinetics of brucine was investigated. Three intravenous (2.5, 5 and 10 mg/kg) and three oral (10, 20 and 40 mg/kg) doses were administered to rats. After intravenous administration, the systemic clearance was reduced and AUC was nonlinearly increased as a function of dose. Upon oral administration, brucine was rapidly absorbed (T(max)<0.5h), which was consistent with previously reported high Caco-2 P(app) values. The increase in AUC was proportional to the increase in dose. The oral bioavailability (F) did not vary with the dose (F=40.31%, 47.15% and 43.02% for 10, 20, 40 mg/kg doses, respectively). However, the dose-proportionality was not observed with C(max). The values of C(max)/Dose were calculated to be 92.92±45.83, 55.73±24.01 and 36.29±22.44 μg/L for 10, 20 and 40 mg/kg, respectively. The results of dose-dependent pharmacokinetic behavior under different administration routes may account for the significantly different toxicities of brucine between intravenous and oral administration.

Topics: Administration, Oral; Animals; Area Under Curve; Biological Availability; Caco-2 Cells; Dose-Response Relationship, Drug; Humans; Injections, Intravenous; Intestinal Absorption; Male; Plant Extracts; Protein Binding; Rats; Rats, Sprague-Dawley; Seeds; Strychnine; Strychnos

A simple, rapid, sensitive and low-cost method using capillary electrophoresis (CE) coupled with field-amplified sample stacking (FASS) has been developed and validated for the simultaneous determination of strychnine and brucine residues in human urine. Before sample loading, a water plug (3.5 kPa, 3 s) was injected to contain sample cations and to permit FASS. Electrokinetic injection at a voltage (20 kV, 25 s) was then used to introduce cations. Separation was performed using 20 mM acetate buffer (pH 3.8) with an applied voltage of 20 kV. The calibration curves were linear over a range of 8.00-2.56 infinity 10(2) ng/mL (r = 0.9995) for strychnine and 10.0-3.20 x 10(2) ng/mL (r = 0.9999) for brucine. Extraction recoveries in urine were greater than 79.6 and 82.8% for strychnine and brucine, respectively, with an RSD of less than 4.9%. The detection limits (signal-to-noise ratio 3) for strychnine and brucine were 2.00 and 2.50 ng/mL, respectively. A urine sample from one healthy female volunteer (26 years old, 50 kg) was pretreated and analyzed. Strychnine and brucine levels in urine could be detected 24 h after administration. On these grounds, this method was feasible for application to preliminary screening of trace levels of abused drugs for both doping control and forensic analysis.

Topics: Adult; Buffers; Calibration; Capillary Electrochromatography; Electrochemistry; Female; Forensic Medicine; Humans; Hydrogen-Ion Concentration; Poisons; Reference Standards; Reproducibility of Results; Solutions; Spectrophotometry, Ultraviolet; Strychnine; Strychnos nux-vomica

Brucine, the major active alkaloid constituent extracted from traditional Chinese herbal medicine Nux vomica, had been found to possess remarkable antitumor, analgesic, and anti-inflammatory activities. In this study, we attempted to encapsulate brucine into liposomes to improve its therapeutic effects. The entrapment efficiency (EE) and the stability of liposomes are two key factors associated with the therapeutic effects of liposomal drugs. We developed a novel liposome-based brucine formulation that was composed of soybean phosphatidylcholine (SPC) and hydrogenated soybean phosphatidylcholine (HSPC).. The liposomes with different phospholipid composition were characterized for their EE, vesicle size, drug release profile, and leakage in vitro.. The molar ratio of HSPC/SPC = 1:9 was determined as the optimum ratio. Compared with conventional liposomes composed of only SPC or HSPC, EE of the brucine-loaded novel liposomes was increased markedly, especially at high drug/lipid molar ratios. The results of drug release showed that the novel liposomes were more stable than the conventional SPC liposomes in the presence of fetal calf serum. In addition, the results of the leakage experiments revealed that the novel liposomes also had better stability in phosphate buffer solution (PBS) with respect to drug retention. Although the conventional HSPC liposomes is more stable than the novel liposomes, the novel liposomes composed of 10% HSPC and 90% SPC may still have promising application potential because HSPC is much more expensive than SPC.. Taken together, efficient encapsulation of brucine into the novel liposomes, their improved stability, and the price of phospholipids indicate that the novel liposomes may act as promising carriers for active alkaloids such as brucine.

Topics: Ammonium Sulfate; Analgesics, Non-Narcotic; Animals; Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Agents; Chemical Phenomena; Drug Carriers; Glycine max; Hydrogenation; Lethal Dose 50; Liposomes; Male; Mice; Mice, Inbred ICR; Particle Size; Phosphatidylcholines; Phospholipids; Seeds; Serum; Strychnine

Saccharomyces cerevisiae is a tractable yeast species for expression and coupling of heterologous G protein-coupled receptors with the endogenous pheromone response pathway. Although this platform has been used for ligand screening, no studies have probed its ability to predict novel pharmacology and functional selectivity of allosteric ligands. As a proof of concept, we expressed a rat M(3) muscarinic acetylcholine receptor (mAChR) bearing a mutation (K(7.32)E) recently identified to confer positive cooperativity between acetylcholine and the allosteric modulator brucine in various strains of S. cerevisiae, each expressing a different human Galpha/yeast Gpa1 protein chimera, and probed for G protein-biased allosteric modulation. Subsequent assays performed in this system revealed that brucine was a partial allosteric agonist and positive modulator of carbachol when coupled to Gpa1/G(q) proteins, a positive modulator (no agonism) when coupled to Gpa1/G(12) proteins, and a neutral modulator when coupled to Gpa1/G(i) proteins. It is noteworthy that these results were validated at the human M(3)K(7.32)E mAChR expressed in a mammalian (Chinese hamster ovary) cell background by determination of calcium mobilization and membrane ruffling as surrogate measures of G(q) and G(12) protein activation, respectively. Furthermore, the combination of this functionally selective allosteric modulator with G protein-biased yeast screens allowed us to ascribe a potential G protein candidate (G(12)) as a key mediator for allosteric modulation of M(3)K(7.32)E mAChR-mediated ERK1/2 phosphorylation, which was confirmed by small interfering RNA knockdown experiments. These results highlight how the yeast platform can be used to identify functional selectivity of allosteric ligands and to facilitate dissection of convergent signaling pathways.

Topics: Allosteric Regulation; Animals; Base Sequence; Carbachol; CHO Cells; Cricetinae; Cricetulus; DNA Primers; Humans; Radioligand Assay; Rats; Receptor, Muscarinic M3; Saccharomyces cerevisiae; Strychnine

Two porphyrin-brucine quaternary ammonium salts were immobilized on gold nanoparticles and their suitability for both in vitro and in vivo photodynamic therapy (PDT) was assayed using the basaloid squamous cell carcinoma PE/CA-PJ34 cell line. In vitro PDT experiments revealed that the gold nanoparticle-bound conjugates were less effective than unbound conjugates in killing cells. However, the same conjugates were more effective in reducing tumor size in vivo, with complete tumor regression observed.

Topics: Alkylation; Animals; Biological Transport; Cell Death; Cell Line, Tumor; Gold; Humans; Intracellular Space; Metal Nanoparticles; Mice; Neoplasms; Photochemotherapy; Porphyrins; Solvents; Strychnine

Salt of brucine hydrogen maleate pentahydrate was synthesized and grown as a single crystal by slow evaporation solution growth technique. The cell parameters of the grown crystal were calculated from powder XRD. The presence of the functional groups and the nature of the vibrations were identified in vibrational studies. The decomposition character of the title material was studied by recording TGA/DTA. The way of promotion of electron from ground state to higher energy state was premeditated by recording UV-VIS-NIR spectrum also the mechanical behaviour was deliberated in hardness measurement.

Topics: Chemical Phenomena; Crystallization; Hardness; Hydroxyl Radical; Kinetics; Maleates; Optical Phenomena; Organic Chemicals; Spectroscopy, Fourier Transform Infrared; Spectrum Analysis, Raman; Strychnine; Temperature; Thermogravimetry; Vibration; X-Ray Diffraction

Multiple myeloma (MM) is an incurable hematological malignancy with high incidence in the elderly. The currently used chemotherapeutic drugs show severe side effects, dose-limiting toxicity and development of resistance. In search of novel plant derived anti-cancer agents, Strychnos nux-vomica L. (SN) root extract was screened using the human MM-cell line, RPMI 8226. SN-extract exhibited anti-proliferative activity in a dose and time dependent manner. The morphological assessment of SN-extract treated cells showed significant features associated with apoptosis. Cell cycle analysis using flow cytometry of cells stained with propidium iodide revealed accumulation of cells at sub-G(0)/G(1) phase. In addition, disruption of mitochondrial membrane potential and subsequent leakage of mitochondrial cytochrome c was observed in SN-extract treated myeloma cells. The anti-proliferative and cytotoxic activity could be due to the alkaloids strychnine and brucine, which have been identified by LC-mass spectral analysis of the SN-extract in comparison to the reference standards analyzed under identical conditions.

Topics: Antineoplastic Agents, Phytogenic; Cell Cycle; Cell Line, Tumor; Cell Nucleus; Cell Proliferation; Cell Survival; Cytochromes c; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Flow Cytometry; Humans; Membrane Potential, Mitochondrial; Mitochondria; Multiple Myeloma; Plant Extracts; Plant Roots; Strychnine; Strychnos nux-vomica

To reduce the toxicity and enhance the therapeutic efficacy of brucine, a traditional Chinese medicine for relieving arthritic and traumatic pain, in this study, a novel brucine-loaded liposomal hydrogel (BLH) formulation, suitable for topical application, was developed. Spherical liposomes composed of lecithin and cholesterol, with brucine, was prepared by a modified ethanol-dripping method. High percentage (over 80%) of encapsulated brucine in liposomes was obtained. Topical liposomal hydrogel formulations were prepared by further incorporation of the prepared liposomes into structured carbopol 940 hydrogels with the concentration of carbopol 1.0%, the ratio of glycerol to carbopol 8:1 and the brucine content 0.1%. The liposomal hydrogel formulations provided an obvious promotion for skin permeation of bruicne while for the free brucine in hydrogels (BH), there was no detectable drug permeation through the skin. The safety evaluation showed that the prepared BLH were no irritation to both the broken and integrity skin. Pharmacodynamic evaluation revealed that the BLH showed a better therapeutic efficacy than that of the BH. So, it can be concluded that the BLH developed here could represent a safe, effective and promising transdermal formulation for local treatment of analgesic and anti-inflammatory disease.

Topics: Administration, Topical; Analgesics; Animals; Chemistry, Pharmaceutical; Dosage Forms; Drug Evaluation, Preclinical; Female; Hydrogel, Polyethylene Glycol Dimethacrylate; Liposomes; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Rabbits; Skin; Strychnine

A three-phase microchip was developed for the rapid and efficient small-scale purification of alkaloids from plant extracts. As part of the development of such a three-phase microchip, first a two-phase microchip with two channels (3.2 cm and 9.3 cm) was used to study the extraction efficiency of strychnine nitrate and strychnine at various flow rates. Strychnine was extracted from a basic aqueous phase to a chloroform phase (extraction) or strychnine was extracted from a chloroform phase into an acidic aqueous phase (back extraction). Subsequently, the "simultaneous extraction and back extraction" of strychnine was carried out in a three-phase microchip. The experimental extraction rate and yield were compared with model data. At a residence time of 25 sec, 79.5% of strychnine was extracted into the acidic aqueous phase using the three-phase microchip. In general, a good correlation was found between experimental results and model data for both two- and three-phase extractions. Finally, the three-phase microchip was employed in the purification of alkaloids (strychnine and brucine) from Strychnos seed extracts.

Topics: Alkaloids; Chloroform; Microfluidic Analytical Techniques; Models, Chemical; Nanotechnology; Plant Extracts; Solvents; Spectrometry, Mass, Electrospray Ionization; Strychnine; Time Factors

Brucine is the predominant alkaloid present in the bark of the tree Strychnos nux vomica and is a weaker alkaloid when compared to strychnine. However, its toxicological property is akin to strychnine. We report a rare case of brucine poisoning complicated by acute renal failure and rhabdomyolysis. A 24-year-old male presented with a history of consumption of a decoction made from the bark of the Strychnos nux vomica tree. Soon after, he developed widespread muscle spasms and convulsions, which were promptly treated. On the fifth day of admission, he developed features of rhabdomyolysis and acute renal failure. Investigations revealed elevated creatine phosphokinase levels and elevated blood urea and serum creatinine. The patient was managed with hemodialysis and recovered gradually. There are many reports of strychnine poisoning producing rhabdomyolysis and renal failure. In this case report, attention is drawn to the fact that brucine, although a weaker alkaloid, can also produce life threatening complications like rhabdomyolysis and acute renal failure.

Topics: Acute Kidney Injury; Creatine Kinase; Humans; Male; Plant Extracts; Poisoning; Poisons; Renal Dialysis; Rhabdomyolysis; Seizures; Strychnine; Strychnos nux-vomica; Treatment Outcome; Young Adult

To prepare and evaluate brucine-loaded polylacticacid nanoparticles (Bru-PLA-NPs).. The Bru-PLA-NPs were prepared by solvent diffusion method. The physical, chemical properties and in vitro release behavior of the prepared Bru-PLA-NPs were evaluated, respectively.. The mean particle size of the prepared Bru-PLA-NPs was 95 nm with polydispersity index of 0.362. The zeta potential was -15.68 mV. The mean loading and entrapment efficiency of Bru were 7% and 37%, respectively. Compared with Bru solution, an obvious sustained release behavior of Bru from Bru-PLA-NPs was observed in the in vitro release experiment.. The Bru-PLA-NPs prepared by solvent diffusion method exhibit small particle size, high Bru-loading efficiency, and obvious sustained release in vitro

Topics: Drug Carriers; Drug Delivery Systems; Drugs, Chinese Herbal; Kinetics; Lactic Acid; Nanoparticles; Particle Size; Polyesters; Polymers; Strychnine

To establish the fingerprint of Semen Strychni and the processed by HPLC.. The HPLC Method was used, the chromatgraphic conditions were as follows: Chromatographic column: Lichrospher C18 (250 mm x 4.6 mm, 5 microm), mobile phase: solvent A consisted of water-ethanoic acid-triethylamine (100: 0.2: 0.2), solvent B contained acetonitril (gradient elution), column temperature: 30 degrees C, flow rate: 1.0 mL/min, detection wavelength: 254 nm.. The similar degree of 10 batch samples buiding sharing mode of crude drug and processed product was above 0.9. 18 common peaks in chromatograms were separated from 10 batches of Semen Strychni samples and 21 common peaks in chromatograms were separated from 10 batches of the processed.. This study establishes the HPLC fingerprint commonmode, and researches the difference between Semen Strychni and the processed in HPLC fingerprint.

Topics: Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Pharmacognosy; Plants, Medicinal; Quality Control; Reproducibility of Results; Seeds; Strychnine; Strychnos

To study the transdermal absorption characteristics of brucine in vitro.. The oil-water partition coefficient of brucine was measured. Drug permeation tests were performed through excised rat skin in improved Franz diffusion cell. Brucine concentration in samples was determined by HPLC.. The oil-water partition coefficient of brucine was between 4.05 and 5.02 at different temperatures. The permeation rate of 0.5 and 1 mg/ml brucine solution were (1.83 +/- 0.85) and (3.74 +/- 1.54) microg/cm2 x h, respectively. The cumulative permeation ratio in 24 hours were (51.30 +/- 18.51)% and (50.01 +/- 12. 80)%, respectively.. The research provides experimental datas for the design of transdermal delivery system of brucine.

Topics: Administration, Cutaneous; Animals; Chromatography, High Pressure Liquid; In Vitro Techniques; Male; Permeability; Pharmaceutical Solutions; Plants, Medicinal; Rats; Rats, Sprague-Dawley; Skin; Skin Absorption; Solubility; Strychnine; Strychnos nux-vomica

A novel analytical method was developed and validated for the rapid and simultaneous analysis of five toxic alkaloids: Brucine, Strychnine, Ephedrine, Aconitine and Colchicine, in blood and urine using high-performance liquid chromatography-electrospray ionization tandem mass spectrometry in the multiple reaction monitoring (HPLC-ESI-MRM) mode. The linear range was 0.05-50.0 ng mL(-1) for Brucine, 0.1-50.0 ng mL(-1) for Strychnine and Ephedrine, 0.01-10.0 ng mL(-1) for Aconitine and Colchicine. The limits of quantification for Brucine, Strychnine, Ephedrine, Aconitine and Colchicine were found to be 0.03, 0.05, 0.20, 0.05, 0.01 ng mL(-1), respectively. The average extraction recoveries in urine ranged from 96.0 to 114.0% and in whole blood were 94.0 to 113.0%. The intra-day and inter-day RSDs were less than 8.3 and 10.6%, respectively. The five alkaloids could be well separated within 7 min in a single run. The established method should be suitable for the determination of trace alkaloids in body fluids.

Topics: Aconitine; Alkaloids; Chromatography, High Pressure Liquid; Colchicine; Drug Stability; Ephedrine; Linear Models; Reproducibility of Results; Sensitivity and Specificity; Solid Phase Extraction; Spectrometry, Mass, Electrospray Ionization; Strychnine; Tandem Mass Spectrometry

To prepare brucine stealth liposomes and compare the in vitro characteristics with brucine conventional liposomes.. Brucine stealth liposomes and conventional liposomes were both prepared by ammonium sulfate transmembrane gradients. The encapsulation efficiency, particle size, in vitro release profiles and stability were compared respectively.. The encapsulation efficiency of brucine stealth liposomes and conventional liposomes were (80.7 +/- 0.5)%, and (80.5 +/- 0.3)%, respectively. The mean paricle sizes were 103.5 nm and 169. 4 nm, respectively. Whether rat plasma was added or not, the release rate and degree of brucine stealth liposomes were significantly lower than those of conventional liposomes. Brucine stealth liposomes were more stable than conventional liposomes.. As the antitumor durg delivery system, the in vitro characteristics of brucine stealth liposomes are more satisfactory than the corresponding conventional liposomes.

Topics: Animals; Drug Stability; Drugs, Chinese Herbal; Liposomes; Particle Size; Rats; Strychnine

Liposomal Brucine (LB) with high encapsulation efficiency (72%) and small particle diameter (mean particle diameter, 54 nm) was prepared by ethanol-dripping method. The safety and pharmacodynamic action of LB, a new transdermal preparation, were investigated in details with the use of white rabbits, guinea-pigs and mice, respectively. The tests revealed that LB had no acute toxicity to integral and broken skin, and had no allergic effects on skin. In writhing test, the analgesic effect of LB was higher than that of free brucine. The anti-inflammatory activity of LB was significantly higher than that of free brucine (P<0.01). Meanwhile, LB exhibited a better dose-response manner and a longer duration of analgesic effects. In conclusion, LB could reduce the toxicity of brucine, enhance the analgesic and antiinflammatory effects of brucine, and achieve its sustained-release.

Topics: Administration, Cutaneous; Analgesics; Animals; Anti-Inflammatory Agents; Female; Guinea Pigs; Liposomes; Male; Mice; Rabbits; Skin; Strychnine; Toxicity Tests, Acute

To study the influencing factors in preparation of brucine liposomes by ammonium sulfate transmembrane gradients.. The brucine liposomes were separated by Sephadex G-50, and the influence of various factors on the entrapment efficiencies were investigated.. The entrapment efficiency was enhanced by increased ammonium sulfate concentration, ethanol volume and PC concentration.. Burcine liposomes prepared by ammonium sulfate transmembrance gradients can get a high entrapment efficiency, the main influencing factors were ammonium sulfate concentration, ethanol volume and PC concentration.

Topics: Ammonium Sulfate; Area Under Curve; Drug Carriers; Drug Stability; Ethanol; Hydrogen-Ion Concentration; Liposomes; Phospholipids; Plants, Medicinal; Quality Control; Strychnine; Strychnos; Technology, Pharmaceutical; Temperature

The concept of supramolecular chirality has assumed increasing importance in association with the development of supramolecular chemistry over the last two decades. In chiral crystals, 2 1 helical molecular assemblies are frequently observed as key motifs. Helical handedness of the 2 1 assemblies, however, has not been determined from the mathematical or crystallographical viewpoints. In this context, we have proposed two new concepts, three-axial chirality and tilt chirality. On the basis of the concepts, we describe supramolecular chirality and determine the handedness of 2 1 assemblies that are composed of relatively complicated molecules with multiple stereogenic centers such as brucine, bile acids, and cinchona alkaloids as well as those of simple molecules.

Topics: Alkaloids; Bile Acids and Salts; Cinchona Alkaloids; Crystallization; Macromolecular Substances; Models, Molecular; Stereoisomerism; Steroids; Strychnine

A new method for the enrichment of Strychnos alkaloids in biological samples via liquid-phase microextraction (LPME) based on porous polypropylene hollow fibers combined with on-line sweeping in micellar electrokinetic chromatography (MEKC) was developed. Strychnos alkaloids were first extracted from urine sample which was adjusted to alkaline conditions (0.5 mol l(-1) NaOH). The unionized analytes were subsequently extracted into 1-octanol impregnated in the pores of hollow fibers, and then into an acidic acceptor solution (100 mmol l(-1) H3PO4) inside the hollow fiber. The extract was analyzed directly by on-line sweeping in MEKC. In the method, the compound berberine was used as the internal standard (I.S.) for the improvement of the experimental reproducibility. The calibration curve was linear over a range of 20-200 ng ml(-1) for both strychnine and brucine in human urine sample, with a correlation coefficient of 0.996 and 0.997, respectively. The detection limits (S/N=3:1) for strychnine and brucine were 1 and 2 ng ml(-1), respectively. The LPME-sweeping method has been successfully applied to the analysis of strychnine and brucine in real urine sample, indicating that LPME-sweeping-MEKC is a promising combination for analysis of basic drugs present at low levels in some biological matrices.

Topics: Calibration; Chromatography, Micellar Electrokinetic Capillary; Humans; Reference Standards; Reproducibility of Results; Sensitivity and Specificity; Strychnine; Strychnos

To study the cytotoxicity of four alkaloids: brucine, strychnine, brucine N-oxide and isostrychnine from nux vomica on SMMC 7721 cells and their possible mechanisms, MET assay was used to examine the growth inhibitory effects of these alkaloids. Brucine revealed the strongest growth inhibitory effect on SMMC-7721 cells. Furthermore, as directly observed under an inverted microscope, fluorescent microscope and transmission electronic microscope, brucine caused SMMC-7721 cell shrinkage, membrane blobbing, formation of apoptotic body as well as nucleus condensation, all of which are typical characteristics of apoptotic programmed cell death. In addition, brucine dose-dependently caused SMMC-7721 cells apoptosis via formation of subdipolid DNA and phosphatidylserine externalization, as evidenced by flow cytometry analysis. The brucine-induced apoptosis was partially attributed to the activation of caspase 3 as well as cyclooxygenase 2 inhibition, since neither caspase 3 specific inhibitor, z-DEVD-fmk nor was exogenous addition of prostaglandin E(2) able to completely abrogate the brucine-induced SMMC 7721 cell apoptosis. In sum, this paper indicate that the major alkaloids present in the seed of Strychnos nux-vomica are effective against SMMC-7721 cells proliferation, among which brucine proceeds SMMC-7721 cells death via apoptosis, probably through the participation of caspase 3 and cyclooxygenase 2.

Topics: Antineoplastic Agents; Apoptosis; Carcinoma, Hepatocellular; Caspase 3; Cell Line, Tumor; Cell Proliferation; Cyclic N-Oxides; Cyclooxygenase 2; Dose-Response Relationship, Drug; Enzyme Activation; Flow Cytometry; Humans; Liver Neoplasms; Seeds; Strychnine; Strychnos nux-vomica

The interaction between brucine and bovine serum albumin (BSA) was investigated using fluorescence spectroscopy (FS) and ultraviolet spectroscopy (UV). The experimental results showed that the brucine quenches the inner fluorescence by forming a brucine-BSA complex. It was found that both static quenching and non-radiation energy transfer were the main reasons for the fluorescence quenching. The apparent binding constants (K(A)) between brucine and BSA were 6. 3 x 10(3) (27 degrees C) and 7.7 x 10(3) (37 degrees C), and the binding sites (n) were 0.94 (27 degrees C) and 0.97 (37 degrees C). According to the Förster theory of non-radiation energy transfer, the binding distances (r) were also obtained. The process of binding was a spontaneous molecular interaction in which entropy increased and Gibbs free energy decreased, indicating that the interaction between brucine and BSA was driven mainly by hydrophobic force.

Topics: Animals; Cattle; Hydrophobic and Hydrophilic Interactions; Serum Albumin, Bovine; Spectrometry, Fluorescence; Spectrophotometry, Ultraviolet; Strychnine

We employed CE to identify mixtures of the toxic alkaloids lappaconitine, bullatine A, atropine sulfate, atropine methobromide, scopolamine hydrobromide, anisodamine hydrobromide, brucine, strychnine, quinine sulfate, and chloroquine in human blood and urine, using procaine hydrochloride as an internal standard. The separation employed a fused-silica capillary of 75 microm id x 60 cm length (effective length: 50.2 cm) and a buffer containing 100 mM phosphate and 5% ACN (pH 4.0). The sample was injected in a pressure mode and the separation was performed at a voltage of 16 kV and a temperature of 25 degrees C. The compounds were detected by UV absorbance at wavelengths of 195 and 235 nm. All the ten alkaloids were separated within 16 min. The method was validated with regard to precision (RSD), accuracy, sensitivity, linear range, LOD, and LOQ. In blood and urine samples, the detection limits were 5-40 ng/mL and linear calibration curves were obtained over the range of 0.02-10 microg/mL. The precision of intra- and interday measurements was less than 15%. Electrophoretic peaks could be identified either by the relative migration time or by their UV spectrum.

Topics: Aconitine; Alkaloids; Atropine; Atropine Derivatives; Electrophoresis, Capillary; Scopolamine; Solanaceous Alkaloids; Strychnine

The structure of a third pseudopolymorphic hydrate of brucine, brucine-water (1/2) [systematic name: 2,3-dimethoxystrychnidin-10-one-water (1/2)], C(23)H(26)N(2)O(4).2H(2)O, has been determined at 130 K. The asymmetric unit comprises two independent brucine molecules and four water molecules of solvation. The four water molecules form uncommon cyclic hydrogen-bonded homomolecular R(4)(4)(8) tetramer rings, which then form primary hydrogen-bonded chain substructures extending down the 2(1) screw axis in the unit cell. The two brucine molecules are linked peripherally to these substructures by either single O-H...N(brucine) or asymmetric three-centre O-H...O(brucine) hydrogen bonds.

Topics: Anions; Crystallography, X-Ray; Hydrogen; Hydrogen Bonding; Indicators and Reagents; Models, Chemical; Models, Molecular; Molecular Conformation; Molecular Structure; Protons; Strychnine; Temperature; Water

To prepare a novel transdermal preparation of liposomal brucine (LB) and investigate its pharmaceutical/pharmacodynamic characterization.. LB was prepared by a modified ethanol-dripping method. Its drug encapsulation efficiency (EE), particle size, in vitro release, and skin permeation were studied. Furthermore, a safety evaluation and pharmacodynamic analysis of LB, including acute dermal toxicity, skin irritation, and analgesic and anti-inflammatory effects were investigated.. the EE of LB was 72% and the mean particle size of the liposomes was 55.4 nm. The in vitro release profile indicated that less than 68% of the encapsulated brucine was released in 10 h. A skin permeation study showed that compared with the free brucine, LB exhibited higher cumulative drug permeation through the skin and lower drug accumulation in skin tissue, indicative of an obvious promotion of skin permeation with liposomal encapsulation. The acute dermal LD50 of LB was greater than 100 mg/kg (brucine content) and skin irritation tests revealed that LB had no irritation to both integrity and broken skin. A pharmacodynamic evaluation of LB was performed by xylene-induced mouse ear edema test and acetic acid-induced writhing test at the dosage of 1.5, 3, and 6 mg/kg, respectively. The results showed that anti-inflammatory activities and analgesic effects of brucine encapsulated were significantly higher than that of the free brucine (P<0.01). Moreover, LB maintained a remarkably longer antiinflammatory and analgesic duration.. It can be proposed that LB prepared here could represent a safe, effective and promising transdermal formulation for analgesic and anti-inflammatory effects.

Topics: Administration, Cutaneous; Analgesics; Animals; Anti-Inflammatory Agents; Drug Delivery Systems; Female; Liposomes; Male; Rabbits; Skin; Strychnine

A method has been developed for rapidly separating and detecting strychnine and brucine using a poly(dimethysiloxane) (PDMS) microchip and electrochemical (EC) detection. PDMS microchannels dynamically modified by Brij35 are shown to be more efficient than native ones. The two analytes are well separated within 90 s in 70 mmol/L acetate buffer (pH 5.5) containing 0.01% (v/v) Brij35. Detection limits were found to be 1.0 micromol/L for strychnine and 0.2 micromol/L for brucine at S/N = 3. The method was used to determine trace strychnine and brucine in rat serum, and the results obtained correlate well with those obtained via high-performance liquid chromatography (HPLC).

Topics: Animals; Buffers; Dimethylpolysiloxanes; Electrochemistry; Hydrogen-Ion Concentration; Microarray Analysis; Molecular Structure; Polyethylene Glycols; Rats; Reproducibility of Results; Strychnine; Time Factors

An easy, rapid method for simultaneous determination of strychnine and brucine in Strychnos nux-vomica L. and its preparation was developed by nonaqueous capillary electrophoresis (NACE) without pretreatment for the first time. Optimum separation was achieved with a fused-silica capillary column (50 cmx75 microm i.d.) and a running buffer containing 30 mM ammonium acetate, 1.0% acetic acid and 15% acetonitrile (ACN) in methanol medium. The applied voltage was 30.0 kV. The analytes were detected by UV at 214 nm. The effects of concentration of ammonium acetate, acetic acid and organic modifier on electrophoretic behavior of the analytes were studied. The established method with sophoridine as internal standard was linear in the range of 5-1000 mg/mL for both strychnine and brucine. The extracts of Strychnos nux-vomica and its preparation could be directly injected for determination with recoveries ranging from 94.5 to 104%.

Topics: Acetates; Acetonitriles; Buffers; Central Nervous System Stimulants; Drugs, Chinese Herbal; Electrophoresis, Capillary; Hydrogen-Ion Concentration; Methanol; Reproducibility of Results; Solvents; Spectrophotometry, Ultraviolet; Strychnine; Strychnos nux-vomica; Time Factors

To screen the anti-tumor effects of the four alkaloids: brucine, strychnine, brucine N-oxide and isostrychnine from the seed of Strychnos nux-vomica, MTT assay was used to examine the growth inhibitory effects of these alkaloids on human hepatoma cell line (HepG2). Brucine, strychnine and isostrychnine revealed significant inhibitory effects against HepG2 cell proliferation, whereas brucine N-oxide didn't have such an effect. In addition, brucine caused HepG2 cell shrinkage, membrane blebbing, apoptotic body formation, all of which are typical characteristics of apoptotic programmed cell death. The results of flow cytometric analysis demonstrated that brucine caused dose-dependent apoptosis of HepG2 cells through cell cycle arrest at G0/G1 phase, thus preventing cells entering S or G2/M phase. Immunoblot results revealed that brucine significantly decreased the protein expression level of cyclooxygenase-2, whereas increased the expression caspase-3 as well as the caspase-3-like protease activity in HepG2 cells, suggesting the involvement of cyclooxygenase-2 and caspase-3 in the pro-apoptotic effects exerted by brucine. Therefore, this paper indicate that the major alkaloids present in the seed of Strychnos nux-vomica are effective against HepG2 cells proliferation, among which brucine proceed HepG2 cells death via apoptosis, probably through the participation of caspase-3 and cyclooxygenase-2.

Topics: Alkaloids; Antineoplastic Agents; Apoptosis; Carcinoma, Hepatocellular; Caspase 3; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cell Shape; Cyclic N-Oxides; Cyclooxygenase 2; Dose-Response Relationship, Drug; Humans; Liver Neoplasms; Seeds; Strychnine; Strychnos nux-vomica

In an attempt to dissect the mechanism of Strychnos nux-vomica, a commonly used Chinese folk medicine in the therapy of liver cancer, the cytotoxic effects of four alkaloids in Strychnos nux-vomica, brucine, brucine N-oxide, strychnine, and isostrychnine, on human hepatoma cells (HepG2) were screened by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrasolium bromide (MTT) assay. Brucine, among the four alkaloids, exhibited the strongest toxic effect, the mechanism of which was found to cause HepG2 cell apoptosis, since brucine caused HepG2 cell shrinkage, the formation of apoptotic bodies, DNA fragmentation, cell cycle arrest, as well as phosphatidylserine externalization, all of which are typical characteristics of apoptotic programmed cell death. Brucine-induced HepG2 cell apoptosis was caspase dependent, with caspase-3 activated by caspase-9. Brucine also caused the proteolytic processing of caspase-9. In addition, brucine caused depolarization of the mitochondrial membrane of HepG2 cells, the inhibition of which by cyclosporine A completely abrogated the activation of casapses and release of cytochrome c in brucine-treated HepG2 cells. These findings suggested a pivotal role of mitochondrial membrane depolarization in HepG2 cell apoptosis elicited by brucine. Furthermore, brucine induced a rapid and sustained elevation of intracellular [Ca2+], which compromised the mitochondrial membrane potential and triggered the process of HepG2 cell apoptosis. Finally, Bcl-2 was found to predominately control the whole event of cell apoptosis induced by brucine. The elevation of [Ca2+]i caused by brucine was also suppressed by overexpression of Bcl-2 protein in HepG2 cells. From the facts given above, Ca2+ and Bcl-2 mediated mitochondrial pathway were found to be involved in brucine-induced HepG2 cell apoptosis.

Topics: Apoptosis; Calcium; Carcinoma, Hepatocellular; Caspase 3; Caspases; Cell Line, Tumor; Cyclic N-Oxides; Cytochromes c; Enzyme Activation; Humans; Liver Neoplasms; Mitochondria, Liver; Proto-Oncogene Proteins c-bcl-2; Seeds; Strychnine; Strychnos nux-vomica

The fragmentations of four strychnos alkaloids have been investigated by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICR-MS) in the positive ion mode. Experiments using multi-stage tandem mass spectrometry (ESI-FT-ICR-MSn) allowed us to obtain precise elemental compositions of product ions at high mass resolution. The experimental data demonstrated that the nitrogen bridge and the coordinated oxygen atom on the nitrogen bridge in the alkaloid compounds were the active sites in the MS2 fragmentations. The loss of CH3 or the OCH3 group in those alkaloids, which have an OCH3 substituent, was the dominant fragmentation mode in the MS3 fragmentations. Logical fragmentation schemes for strychnos alkaloids have been proposed and these should be useful for the identification of these compounds.

Topics: Alkaloids; Cyclotrons; Fourier Analysis; Spectrometry, Mass, Electrospray Ionization; Strychnine; Strychnos; Strychnos nux-vomica; Tandem Mass Spectrometry

Strychnine and brucine from the plant Strychnos nux vomica have been shown to have interesting pharmacological effects on several neurotransmitter receptors, including some members of the superfamily of ligand-gated ion channels. In this study, we have characterised the pharmacological properties of tertiary and quaternary analogues as well as bisquaternary dimers of strychnine and brucine at human alpha1 and alpha1beta glycine receptors and at a chimera consisting of the amino-terminal domain of the alpha7 nicotinic receptor (containing the orthosteric ligand binding site) and the ion channel domain of the 5-HT3A serotonin receptor. Although the majority of the analogues displayed significantly increased Ki values at the glycine receptors compared to strychnine and brucine, a few retained the high antagonist potencies of the parent compounds. However, mirroring the pharmacological profiles of strychnine and brucine, none of the analogues displayed significant selectivity between the alpha1 and alpha1beta subtypes. The structure-activity relationships for the compounds at the alpha7/5-HT3 chimera were significantly different from those at the glycine receptors. Most strikingly, quaternization of strychnine and brucine with substituents possessing different steric and electronic properties completely eliminated the activity at the glycine receptors, whereas binding affinity to the alpha7/5-HT3 chimera was retained for the majority of the quaternary analogues. This study provides an insight into the structure-activity relationships for strychnine and brucine analogues at these ligand-gated ion channels.

Topics: alpha7 Nicotinic Acetylcholine Receptor; Cell Line; Dimerization; Humans; Ion Channel Gating; Ligands; Membrane Potentials; Quaternary Ammonium Compounds; Radioligand Assay; Receptors, Glycine; Receptors, Nicotinic; Receptors, Serotonin, 5-HT3; Recombinant Fusion Proteins; Structure-Activity Relationship; Strychnine

The structures of two brucinium (2,3-dimethoxy-10-oxostrychnidinium) salts of the alpha-hydroxy acids L-malic acid and L-tartaric acid, namely brucinium hydrogen (S)-malate pentahydrate, C23H27N2O4+.C4H5O5-.5H2O, (I), and anhydrous brucinium hydrogen (2R,3R)-tartrate, C23H27N2O4+.C4H5O6-,(II), have been determined at 130 K. Compound (I) has two brucinium cations, two hydrogen malate anions and ten water molecules of solvation in the asymmetric unit, and forms an extensively hydrogen-bonded three-dimensional framework structure. In compound (II), the brucinium cations form the common undulating brucine sheet substructures, which accommodate parallel chains of head-to-tail hydrogen-bonded tartrate anion species in the interstitial cavities.

Topics: Crystallography, X-Ray; Hydrogen; Hydrogen Bonding; Malates; Models, Molecular; Molecular Structure; Salts; Strychnine; Tartrates

Diffusion ordered spectroscopy (DOSY) is a powerful two-dimensional NMR method to study molecular translation in various systems. The diffusion coefficients are usually retrieved, at each frequency, from a fit procedure on the experimental data, considering a unique coefficient for each molecule or mixture. However, the fit can be improved if one regards the decaying curve as a multiexponential function and the diffusion coefficient as a distribution. This work presents a computer code based on the Hopfield neural network to invert the data. One small-molecule binary mixture with close diffusion coefficients is treated with this approach, demonstrating the effectiveness of the method.

Topics: Bridged Bicyclo Compounds; Diffusion; Neural Networks, Computer; Nuclear Magnetic Resonance, Biomolecular; Propionates; Strychnine

Two quality control methods of nuxvomica were established for mutual. By selecting the appropriate measuring wavelength or wavelength range, the contents of strychnine and brucine in nuxvomica were determined without any preliminary separation by a new rapid spectrophotometry and a multi-wavelength linear regression spectrophotometry and with a computer program. The linear range of strychnine measured was 8.0-30.0 microg x mL(-1) (r = 0.999 9); The linear range of brucine measured was 7.0-31.2 microg x mL(-1) (r = 0.999 4). The average recoveries and relative standard deviations of strychnine and brucine were 98.18%-99.82%, 0.56%-1.54% and 100.5%-100.6%, 0.57%-0.62%, respectively. The methods are simple, rapid and reproducible, the interference of two components with each other may be eliminated and the methods are appropriate for quality control of nuxvomica.

Topics: Drugs, Chinese Herbal; Quality Control; Spectrophotometry; Strychnine; Strychnos nux-vomica

To establish a new method for determination of strychnine alkaloid in biological fluids based on molecularly imprinted polymers.. A strychnine molecularly imprinted monolithic column was prepared by in-situ molecularly imprinted technique. The polymer was filled to a 1cm column, and a method was developed to concentrate and determine strychnine alkaloids in biological fluids.. the limit of detection of the method was 4.9 ng, and the recoveries were more than 92%. The relative standard deviations were smaller than 6.59%. The linear correlation coefficients of standard curves were 0.999 1 and 0.9966 respectively. This method was applied to concentrate and determine strychnine in plasma and urine of poisoned rabbit.. The new method could concentrate and simultaneously determine strychnine alkaloids in biological fluids, and it was applied to forensic toxicological analysis.

Topics: Alkaloids; Animals; Chromatography, High Pressure Liquid; Humans; Male; Polymers; Rabbits; Sensitivity and Specificity; Strychnine

The human glycine receptor subtypes alpha1beta and alpha2 have been expressed stably in HEK293 cells, and the functional characteristics of the receptors have been characterised in the FLIPR Membrane Potential Assay. The pharmacological properties obtained for nine standard ligands at the two receptors in this assay were found to be in good agreement with those from electrophysiology studies of the receptors expressed in Xenopus oocytes or mammalian cell lines. Hence, this high throughput screening assay will be of great use in future pharmacological studies of glycine receptors, particular in the search for novel compound structures acting at them.

Topics: Androstanes; Azasteroids; beta-Alanine; Cell Line; Dose-Response Relationship, Drug; Fluorescence; Glycine; Glycine Agents; Humans; Membrane Potentials; Patch-Clamp Techniques; Picrotoxin; Receptors, Glycine; Strychnine; Taurine; Transfection

A capillary zone electrophoresis (CZE) method has been developed for investigating the physicochemical characteristics of five Strychnos alkaloids in Strychnos nux-vomica L. Firstly, the dissociation constants of the five Strychnos alkaloids were determined, based on the relation between the effective mobility of the solutes and the buffer pH. The mathematical relationship was strictly deduced from the fundamental electrophoretic theory and the dissociation equilibrium. Secondly, an equation describing the relation between the migration time of alkaloids of similar structure and their molecular weights was developed and used to predict the migration order and to calculate the electrosomotic velocity. The results predicted by the theory agreed with those from experiments.

Topics: Alkaloids; Electrophoresis, Capillary; Hydrogen-Ion Concentration; Matrines; Molecular Structure; Molecular Weight; Quinolizines; Sophora; Strychnine; Strychnos nux-vomica

A portable chip-CE system with potential gradient detection (PGD) was developed and applied to the determinations of alkali metals and alkaloids. The separation efficiency appeared to be satisfactory and nonaqueous capillary electrophoresis (NACE) proved to be applicable to PGD or conductivity detection. The power supplies, separation and detection were built on a device of 3 kg in weight. A branch channel near the end of the separation channel was designed to perform PGD and make the application of relatively high field strength possible. The study is the first report on the application of PGD on the microchip platform. The design of the chip-CE system shows several advantages, such as simplicity, miniaturization and wide applicability.

Topics: Alkaloids; Electrophoresis, Capillary; Metals, Alkali; Miniaturization; Strychnine

A sensitive method for the identification and quantitation of the toxic alkaloids strychnine and brucine from postmortem specimens has been established. After solid-phase extraction using Oasis MCX cartridges the extracts were analyzed by high-performance liquid chromatography with diode-array detection. The limit of detection was 0.5 ng/mL blood for strychnine and brucine, and the limit of quantitation was 5 ng/mL blood for strychnine and brucine. The method was applied for the analysis of blood and gastric contents of a 34-year-old female who died after ingestion of a packet of herbal medicine powder containing the seeds of Strychnos nux-vomica L. Strychnine and brucine were detected in all the samples. The concentration in our case is consistent with that in previous reports.

Topics: Adult; Chromatography, High Pressure Liquid; Fatal Outcome; Female; Forensic Medicine; Humans; Poisons; Seeds; Strychnine; Strychnos nux-vomica

Topics: Carbon Isotopes; Cholesterol Esters; Magnetic Resonance Spectroscopy; Molecular Structure; Spin Labels; Strychnine; Tyrosine

Following a recent description of fluorescence resonance energy transfer between enhanced green fluorescent protein (EGFP)-fused human muscarinic M1 receptors and Bodipy-labeled pirenzepine, we synthesized seven fluorescent derivatives of this antagonist in order to further characterize ligand-receptor interactions. These compounds carry Bodipy [558/568], Rhodamine Red-X [560/580], or Fluorolink Cy3 [550/570] fluorophores connected to pirenzepine through various linkers. All molecules reversibly bind with high affinity to M1 receptors (radioligand and energy transfer binding experiments) provided that the linker contains more than six atoms. The energy transfer efficiency exhibits modest variations among ligands, indicating that the distance separating EGFP from the fluorophores remains almost constant. This also supports the notion that the fluorophores may bind to the receptor protein. Kinetic analyses reveal that the dissociation of two Bodipy derivatives (10 or 12 atom long linkers) is sensitive to the presence of the allosteric modulator brucine, while that of all other molecules (15-24 atom long linkers) is not. The data favor the idea that these analogues might interact with both the acetylcholine and the brucine binding domains.

Topics: Allosteric Regulation; Binding Sites; Binding, Competitive; Boron Compounds; Fluorescence Resonance Energy Transfer; Fluorescent Dyes; Green Fluorescent Proteins; Humans; Kinetics; Ligands; Luminescent Proteins; Pirenzepine; Radioligand Assay; Receptor, Muscarinic M1; Recombinant Fusion Proteins; Rhodamines; Structure-Activity Relationship; Strychnine

The enantiomeric resolution of (+/-)-ibuprofen into its enantiomers was achieved by TLC on silica gel plate using optically pure (-)-brucine as a chiral selector and acetonitrile-methanol (5:1, v/v) as the solvent system. Spots were located in an iodine chamber. The detection limit was 4.9 microg. The effect of concentration of the chiral selector, temperature and pH on resolution has been studied.

Topics: Acetonitriles; Anti-Inflammatory Agents, Non-Steroidal; Chromatography, Thin Layer; Hydrogen-Ion Concentration; Ibuprofen; Indicators and Reagents; Methanol; Solvents; Stereoisomerism; Strychnine; Temperature

4-Amino-2-mercaptopyrimidine self-assembled monolayer (AMP SAMs/Au) was prepared on a gold electrode. The AMP SAMs/Au was characterized by using attenuated total reflection-fourier transform infrared (ATR-FTIR) and A.C. Impedance. The electrochemical behavior of brucine on AMP SAMs/Au was studied by cyclic voltammetry (CV) and square wave adsorptive stripping voltammetry (SWASV). The modified electrode showed an excellent electrocatalytic activity for the redox of brucine. The catalytic current increased linearly with the concentration of brucine in the range of 4.0 x 10(-7) to 2.0 x 10(-4) mol l(-1) by square wave voltammetry response. The detection limit was 6.0 x 10(-8) mol l(-1).

Topics: Electrochemistry; Electrodes; Ferrocyanides; Gold; Molecular Structure; Oxidation-Reduction; Pyrimidines; Sensitivity and Specificity; Spectroscopy, Fourier Transform Infrared; Strychnine; Sulfhydryl Compounds

A new capillary electrophoresis procedure with field-enhanced stacking concentration for the analysis of strychnine and brucine is established. After optimization of the separation and concentration conditions, the two alkaloids can be separated within 5 min and quantified with high sensitivity (The detection limits were 1.0 ng mL(-1) for strychnine and 1.4 ng mL(-1) for brucine). The method was useful for qualitative and quantitative analysis of strychnine and brucine in Strychnos nux-vomica L with recovery of 105.1% for strychnine and 98.4% for brucine.

Topics: Drugs, Chinese Herbal; Electrophoresis, Capillary; Hydrogen-Ion Concentration; Magnoliopsida; Sensitivity and Specificity; Strychnine; Strychnos nux-vomica

Bisquaternary dimers of strychnine and brucine were synthesized and their allosteric effect on muscarinic acetylcholine M(2) receptors was examined. The compounds retarded the dissociation of the antagonist [(3)H]N-methylscopolamine ([(3)H]NMS) from porcine cardiac cholinoceptors. This action indicated ternary complex formation. All compounds exhibited higher affinity to the allosteric site of [(3)H]NMS-occupied M(2) receptors than the monomeric strychnine and brucine, while the positive cooperativity with NMS was fully maintained. SAR studies revealed the unchanged strychnine ring as an important structural feature for high allosteric potency.

Topics: Allosteric Regulation; Animals; Dimerization; Molecular Conformation; Muscarinic Antagonists; N-Methylscopolamine; Radioligand Assay; Receptor, Muscarinic M2; Structure-Activity Relationship; Strychnine; Swine

A modified and highly sensitive spectrophotometric method for the determination of nitrate in trace quantities in environmental samples is described. The method is based on the reaction of nitrate ion with brucine and 3-methyl-2-benzothiazolinone hydrazone hydrochloride (MBTH) in sulfuric acid medium to yield a violet-colored product which is stable for over two days. The optimum photometric range for the determination of nitrate is 0.04-0.16 microg cm(-3) and the Sandell's sensitivity being 0.000279 microg cm(-2). The proposed method is applied to various water samples and the results indicate that the reaction is highly sensitive than the original brucine method.

Topics: Benzothiazoles; Fresh Water; Hydrazones; Indicators and Reagents; Nitrates; Rain; Sewage; Spectrophotometry, Ultraviolet; Strychnine; Thiazoles; Water Pollutants, Chemical; Water Supply

To further understand the purpose of the traditional processing method of the seeds of Strychnos nux-vomica L. (Loganiaceae) as well as analgesic and anti-inflammatory activities of brucine and brucine N-oxide extracted from this medicinal plant, various pain and inflammatory models were employed in the present study to investigate their pharmacological profiles. Both brucine and brucine N-oxide revealed significant protective effects against thermic and chemical stimuli in hot-plate test and writhing test. However, on different phases they exerted analgesic activities in formalin test. Brucine N-oxide showed stronger inhibitory effect than brucine in carrageenan-induced rat paw edema, both of them significantly inhibited the release of prostaglandin E2 in inflammatory tissue, reduced acetic acid-induced vascular permeability and the content of 6-keto-PGF1a in Freund's complete adjuvant (FCA) induced arthritis rat's blood plasma. In addition, brucine and brucine N-oxide were shown to reduce the content of 5-hydroxytryptamine (5-HT) in FCA-induced arthritis rat's blood plasma, while increase the content of 5-hydroxytryindole-3-acetic acid (5-HIAA) accordingly. These results suggest that central and peripheral mechanism are involved in the pain modulation and anti-inflammation effects of brucine and brucine N-oxide, biochemical mechanisms of brucine and brucine N-oxide are different even though they are similar in chemical structure.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Experimental; Capillary Permeability; Cyclic N-Oxides; Disease Models, Animal; Edema; Female; Male; Mice; Mice, Inbred ICR; Pain; Plant Extracts; Rats; Rats, Sprague-Dawley; Seeds; Serotonin; Strychnine; Strychnos nux-vomica

To study the pharmacokinetic process about the concentration in rat plasma of the alkaloids from processed seeds of Strychnos nux-vomica with RP-HPLC method.. Hypersil BDS C18 column was used and the mobile phase consisted of acetonitrile-water at the flow rate of 0.8 mL.min-1. The UV detection wave length was 254 nm.. The concentration-time data of strychnine, brucine, strychnine N-oxide and brucine N-oxide were all in accordance with an open two-compartment model after i.v. alkaloids. Their parameters were as follows: T1/2 alpha were (8 +/- 5), (4 +/- 3), (6.2 +/- 1.7) and (3.0 +/- 0.8) min, T1/2 beta were (262 +/- 125), (416 +/- 131), (285 +/- 50) and (342 +/- 141) min, CL were (17 +/- 4), (21 +/- 12), (1.9 +/- 1.8) and (2.8 +/- 1.1) mL.min-1, Vc were (1.4 +/- 0.5), (1.7 +/- 1.1), (0.24 +/- 0.16) and (0.23 +/- 0.06) L.kg-1, Vd were (6.0 +/- 1.2), (12 +/- 7), (0.8 +/- 0.6) and (1.5 +/- 0.6) L.kg-1, AUC were (57,578 +/- 25,578), (35,240 +/- 15,616), (93,088 +/- 22,375) and (177,712 +/- 120,110) h.microgram.L-1, respectively.. The method is a good reference for pharmacokinetics in human bodies.

Topics: Alkaloids; Animals; Cyclic N-Oxides; Drugs, Chinese Herbal; Female; Hot Temperature; Male; Plants, Medicinal; Rats; Rats, Sprague-Dawley; Seeds; Strychnine; Strychnos nux-vomica; Technology, Pharmaceutical

A quantitative analysis using (1)H-NMR (Q-NMR) has been developed for the determination of strychnine and brucine in Strychnos nux-vomica seeds and stems. The advantages of the method are that no reference alkaloids are needed for calibration curves, the quantification could be directly realized on a crude extract, strychnine and brucine could easily be distinguished, an overall profile of the preparation (including non alkaloid compounds) could be directly obtained, and a very significant time-gain could be achieved, in comparison to conventional HPLC methods, for instance.

Topics: Chromatography, High Pressure Liquid; Humans; Magnetic Resonance Spectroscopy; Phytotherapy; Plant Stems; Seeds; Strychnine; Strychnos nux-vomica

This paper reports a HPLC method for determinition of strychnine and brucine in Semen Strychni and its processed products of Jiangxi method and innovated methed.. SiO2 was used as the stationary phase, n-hexane-dichloromethane-methanol-ammonia(47.5:47.5:5:0.35) as the mobile phase, with detection wavelength of 254 nm.. The contents of strychnine and brucine in the processed products of Jiangxi are lower.. This method is accurate, simple and reliable.

Topics: Chromatography, High Pressure Liquid; Hot Temperature; Plants, Medicinal; Seeds; Strychnine; Strychnos nux-vomica; Technology, Pharmaceutical

The asymmetric coupling of various phenol or aniline derivatives with bulky aryllead triacetates was thoroughly investigated using optically active amines, including strychnine and brucine. We found that conformationally restricted tertiary amines, as well as lithium aryloxides and molecular sieves, are essential for accelerating the rate of phenol coupling. Consequently, the reaction can be carried out at a low temperature (-40 to -20 degrees C) and gives a high degree of diastereo- and enantioselectivity. In contrast to the effectiveness of lithiation in phenol coupling, magnesation of anilines was a critical technique for aniline coupling with aryllead triacetates. Using these coupling methods, a diverse set of di-, tri, and polyaryl compounds with axial chirality can be easily obtained, and these should be useful for the construction of a variety of aryl-aryl frameworks involved in metal ligands, natural products, and artificial helical polymers.

Topics: Amines; Aniline Compounds; Molecular Conformation; Organometallic Compounds; Phenols; Polycyclic Aromatic Hydrocarbons; Stereoisomerism; Strychnine

The plant alkaloid brucine is an analogue of strychnine and is known to be an allosteric modulator at cloned M(1) muscarinic receptors. The functional effects of brucine were examined on the M(1) muscarinic receptors in the rabbit isolated vas deferens preparation. Brucine (10-100 microM) enhanced the effects of the muscarinic agonist McN-A-343 at presynaptic M(1) muscarinic receptors in the rabbit isolated vas deferens preparation, but only when brucine was added prior to McN-A-343. This effect is indicative of a positive allosteric action. It was poorly reversed on washing. Brucine did not affect the responses to the mamba venom muscarinic toxins MT2 and MT4, which are also allosteric activators in this preparation. Brucine (10-100 microM) caused a significant decrease in the twitch response to electrical stimulation in the rabbit vas deferens preparation, which was not antagonised by 100 nM pirenzepine (an M(1) muscarinic antagonist). Brucine and MT4, but not MT2, caused significant decreases (p<0.05) in the responses to noradrenaline in the rabbit vas deferens preparation. Responses to ATP and KCl were not affected. In radioligand binding assays, brucine displaced the alpha(1)-adrenoceptor ligand prazosin from its specific binding sites in membranes made from rat cerebral cortex and rat vas deferens. The apparent K(i) values were 150 and 3.4 microM in the cortical and vas deferens membranes, respectively. The positive allosterism found with brucine at cloned M(1) receptors seems to be mirrored at native M(1) receptors. However, the unexpected blocking effects at alpha(1)-adrenoceptors indicates that more selective ligands than brucine are required as starting points for the design of specific enhancers of the activity of M(1) receptors with therapeutic potential.

Topics: (4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethylammonium Chloride; Allosteric Regulation; Animals; Male; Prazosin; Rabbits; Rats; Rats, Sprague-Dawley; Receptor, Muscarinic M1; Receptors, Adrenergic, alpha-1; Receptors, Muscarinic; Strychnine; Vas Deferens

Allosteric enhancement of the affinity of muscarinic receptors for their ligands offers a new way to influence cholinergic neurotransmission. The structure of the allosteric binding domain(s) and the features of agonists, antagonists and modulators which determine the occurrence of either positive or negative cooperativity require clarification. We tested interactions between allosteric modulators alcuronium, strychnine and brucine and eight antagonists at muscarinic receptors expressed in CHO cells. In experiments with unlabeled antagonists, all three modulators enhanced the affinity for 4-diphenylacetoxy-N-dimethylpiperidinium (4-DAMP) at the M2 receptors, and strychnine did so also at the M4 receptors. Positive interactions were also observed between alcuronium and L-hyoscyamine (M2) and scopolamine (M2), between strychnine and butylscopolamine (M4), L-hyoscyamine (M2 and M4) and scopolamine (M4), and between brucine and scopolamine (M2). Positive effects of alcuronium, strychnine and brucine on the affinity of the M2 receptors for 4-DAMP have been confirmed by direct measurements of the binding of [3H]-4-DAMP. A comparison of molecular models of several antagonists which are esters revealed that antagonists in which the distance between the N and the carboxyl C atoms corresponds to five chemical bonds are more likely to display positive cooperativity with alcuronium at the M2 receptors than the antagonists in which the N-carboxyl C distance corresponds to four chemical bonds.

Topics: Alcuronium; Allosteric Regulation; Animals; CHO Cells; Cricetinae; Muscarinic Antagonists; N-Methylscopolamine; Piperidines; Radioligand Assay; Strychnine; Tritium

An HPLC method for the determination of brucine in seed dressing agents was investigated. The sample solution was separated on a Spherisorb C18 column(4.6 mm i.d. x 200 mm, 5 microns) with CH3OH-H2O-CTAB (88:12:0.04, m/m) as the mobile phase and detected at 254 nm. Calibration curve of brucine was obtained for the concentration range of 0.01 g/L-0.25 g/L. The linear regression equation was A = 11,485,814 rho + 21,229, r = 0.9997. The RSD was 1.2%(n = 11). The results show that this method is simple, specific and accurate.

Topics: Chromatography, High Pressure Liquid; Pesticides; Strychnine

A capillary zone electrophoresis method was developed for the separation and determination of strychnine and brucine in Strychnos nux-vomica L. and its preparation. The factors that could affect the separation were studied, such as the types and concentrations of electrolytes, pH, ionic strength and organic modifier. The optimum running buffer was 20 mmol/L of ammonium acetate containing 0.2 mol/L of glacial acetic acid (pH 3.64). The applied voltage was 25 kV and the wavelength of the UV detector was set at 214 nm. The established method with dopamine hydrochloride as internal standard was linear in the range of 5-100 microg/mL for both strychnine and brucine. The recovery was 102.96% for strychnine and 98.56% for brucine. The extracts of Strychnos nux-vomica and its preparation could be directly injected for analysis.

Topics: Buffers; Electrophoresis, Capillary; Hydrogen-Ion Concentration; Magnoliopsida; Strychnine

The affinity and allosteric properties of 22 quaternary derivatives of strychnine and brucine at the m1-m4 subtypes of muscarinic receptors have been analyzed and compared. The subtype selectivity, in terms of affinity, was in general m2 > m4 > m1 > m3. The highest affinities were found for N-benzyl, N-2-naphthylmethyl, and N-4-biphenylylmethyl strychnine (13, 14, and 18, respectively). All the strychnine and brucine derivatives were positively cooperative with the antagonist, N-methylscopolamine, at m2 receptors and, in the case of the strychnine analogues, were positively cooperative with N-methylscopolamine at least at one other subtype. The strychnine analogues were negatively cooperative with the neurotransmitter, acetylcholine, at all subtypes whereas brucine and five of the six derivatives examined were positively cooperative with acetylcholine at one or more subtypes (m1-m5) and exhibited different patterns of subtype selectivity. The ability to generate subtype-selective allosteric enhancers of acetylcholine binding and function may be of use in the development of drugs for the treatment of Alzheimer's disease.

Topics: Allosteric Regulation; Animals; CHO Cells; Cricetinae; Humans; Muscarinic Antagonists; N-Methylscopolamine; Receptors, Muscarinic; Structure-Activity Relationship; Strychnine

We previously demonstrated that brucine and some analogues allosterically enhance the affinity of ACh at muscarinic receptor subtypes M1, M3 or M4. Here we describe allosteric effects at human M1-M4 receptors of four stereoisomers of a pentacyclic structure containing features of the ring structure of brucine. All compounds inhibited 3H-NMS dissociation almost completely at all subtypes with slopes of 1, with similar affinity values at the 3H-NMS-occupied receptor to those estimated from equilibrium assays, consistent with the ternary complex allosteric model. Compound 1a showed positive cooperativity with H-NMS and small negative or neutral cooperativity with ACh at all subtypes. Its stereoisomer, 1b, showed strong negative cooperativity with both 3H-NMS and ACh across the subtypes. Compound 2a was positive with 3H-NMS at M2 and M4 receptors, neutral at M3 and negative at M1 receptors; it was negatively cooperative with ACh at all subtypes. Its stereoisomer, 2b, was neutral with 3H-NMS at M1 receptors and positive at the other subtypes; 2b was negatively cooperative with ACh at M1, M3 and M4 receptors but showed 3-fold positive cooperativity with ACh at M2 receptors. This latter result was confirmed with further 3H-NMS and 3H-ACh radioligand binding assays and with functional assays of ACh-stimulated 35S-GTPgammaS binding. These results provide the first well characterised instance of a positive enhancer of ACh at M2 receptors, and illustrate the difficulty of predicting such an effect.

Topics: Acetylcholine; Allosteric Regulation; Allosteric Site; Animals; Cell Membrane; CHO Cells; Cricetinae; Guanosine 5'-O-(3-Thiotriphosphate); Guanosine Triphosphate; Humans; Indoles; Kinetics; Muscarinic Agonists; Muscarinic Antagonists; N-Methylscopolamine; Propylbenzilylcholine Mustard; Receptors, Muscarinic; Stereoisomerism; Strychnine; Thermodynamics

In radioligand binding studies, it has been reported that brucine, N-chloromethyl brucine, and brucine N-oxide increased the affinity of acetylcholine for M1, M3, and M4 muscarinic receptors, respectively, in a manner consistent with the predictions of the ternary complex allosteric model. We now demonstrate an equivalent ability of these three allosteric agents to modulate the actions of acetylcholine in functional studies in membranes and in whole cells. The enhancing actions of brucine and brucine N-oxide on acetylcholine (ACh) potency at M1 and M4 receptors respectively have been confirmed in guanosine-5'-O-(3-[35S]thio)triphosphate, GTPase, cAMP, and intracellular Ca2+ mobilization assays of function. In general, neither the basal nor the maximally stimulated response to ACh is affected. The subtype-selective allosteric effects of N-chloromethyl brucine on M2 and M3 receptors were shown to be qualitatively and quantitatively the same in guanosine-5'-O-(3-[35S]thio)triphosphate functional assays, in terms of both its affinity and cooperativity with ACh, as those found in binding assays. Neutral cooperativity of N-chloromethyl brucine with ACh on M4 receptor function was also observed, thereby demonstrating its "absolute subtype selectivity": a lack of action at any concentration at M4 receptors and an action at M2 and M3 receptors. The enhancing action of N-chloromethyl brucine on neurogenically released ACh binding at M3 receptors was also detected in whole tissue as an increased contraction of the isolated guinea pig ileum to submaximal electrical stimulation. In conclusion, these functional studies confirm that brucine analogs are allosteric enhancers of ACh affinity at certain muscarinic receptor subtypes.

Topics: Acetylcholine; Allosteric Regulation; Animals; Cell Membrane; CHO Cells; Cricetinae; Cyclic N-Oxides; Guinea Pigs; Humans; Male; Receptor, Muscarinic M1; Receptor, Muscarinic M3; Receptor, Muscarinic M4; Receptors, Muscarinic; Strychnine

Direct enantioseparation of (+/-)-ibuprofen and (+/-)-flurbiprofen was achieved by two-dimensional thin-layer chromatography on silica gel plates impregnated with optically pure (-)-brucine as chiral selector. The solvent systems that were successful in resolving both the compounds were acetonitrile-methanol (16:3 v/v) for the first dimension and acetonitrile-methanol-water (16:3:0.4, v/v) for the second dimension. Iodine vapour was used for detection; the detection limit for both (+/-)-ibuprofen and (+/-)-flurbiprofen was 0.1 microgram.

Topics: Chromatography, Thin Layer; Flurbiprofen; Ibuprofen; Stereoisomerism; Strychnine

We studied the interactions of strychnine, brucine, and three of the N-substituted analogues of brucine with [3H]N-methylscopolamine (NMS) and unlabeled acetylcholine at m1-m5 muscarinic receptors using equilibrium and nonequilibrium radioligand binding studies. The results were consistent with a ternary allosteric model in which both the primary and allosteric ligands bind simultaneously to the receptor and modify the affinities of each other. The compounds had Kd values in the submillimolar range, inhibited [3H]NMS dissociation, and showed various patterns of positive, neutral, and negative cooperativity with [3H]NMS and acetylcholine, but there was no predictive relationship between the effects. Acetylcholine affinity was increased approximately 2-fold by brucine at m1 receptors, approximately 3-fold by N-chloromethyl brucine at m3 receptors, and approximately 1.5-fold by brucine-N-oxide at m4 receptors. The existence of neutral cooperativity, in which the compound bound to the receptor but did not modify the affinity of acetylcholine, provides the opportunity for a novel form of drug selectivity that we refer to as absolute subtype selectivity: an agent showing positive or negative cooperativity with the endogenous ligand at one receptor subtype and neutral cooperativity at the other subtypes would exert functional effects at only the one subtype, regardless of the concentration of agent or its affinities for the subtypes. Our results demonstrate the potential for developing allosteric enhancers of acetylcholine affinity at individual subtypes of muscarinic receptor and suggest that minor modification of a compound showing positive, neutral, or low negative cooperativity with acetylcholine may yield compounds with various patterns of cooperativity across the receptor subtypes.

Topics: Acetylcholine; Allosteric Regulation; Animals; CHO Cells; Cricetinae; N-Methylscopolamine; Radioligand Assay; Receptors, Muscarinic; Strychnine

1. Radioligand binding experiments indicate that the affinity of muscarinic receptors for their agonists may be enhanced by allosteric modulators. We have now investigated if brucine can enhance the inhibitory effects of muscarinic receptor agonists on the electrically evoked release of [3H]acetylcholine ([3H]ACh) from superfused slices of rat striatum. 2. The evoked release of [3H]ACh was inhibited by all agonists tested (i.e., furmethide, oxotremorine-M, bethanechol and oxotremorine). 3. Brucine enhanced the inhibitory effects of furmethide, oxotremorine-M and bethanechol on the evoked [3H]ACh release without altering the inhibitory effect of oxotremorine. 4. Alcuronium was applied for comparison and found to diminish the inhibitory effect of furmethide on the evoked [3H]ACh release. 5. The results demonstrate that it is possible both to enhance and diminish the functional effects of muscarinic receptor agonists by allosteric modulators. 6. The direction of the observed effects of brucine and alcuronium on [3H]ACh release fully agrees with the effects of these modulators on the affinities of human M4 receptors for furmethide, oxotremorine-M, bethanechol and oxotremorine, as described by Jakubik et al. (1997). This supports the view that the presynaptic muscarinic receptors responsible for the autoinhibition of ACh release in rat striatum belong to the M4 muscarinic receptor subtype.

Topics: Acetylcholine; Alcuronium; Animals; Bethanechol; Corpus Striatum; Humans; In Vitro Techniques; Male; Muscarinic Agonists; Oxotremorine; Quaternary Ammonium Compounds; Rats; Rats, Wistar; Strychnine; Tritium

To examine the cytotoxicities of 6 crude Strychnos alkaloid fractions from the seeds of Strychnos nux-vomica unprocessed or processed with various traditional processing methods and 13 pure Strychnos alkaloids from the fractions.. Using cell culture, their inhibitory effects on Vero cell growth-inhibition assay, and host cell DNA synthesis by [3H]thymidine ([3H]TdR) uptake assay.. The IC50 of processed seeds were 155% and 212% of unprocessed ones in cell growth-inhibition assay and in [3H]TdR uptake assay, respectively. The IC50 of 13 compounds were 0.45-0.80 mmol.L-1 and 0.50-12 mmol.L-1, respectively. The processing method with sand bath exhibited a wide safety margin compared with other traditional processing methods or no processing. The isomers of Strychnos alkaloids and their N-oxides showed much lower cytotoxicities among these alkaloids. Isobrucine N-oxide showed the lowest cytotoxicity. The contents of isomers and N-oxides of Strychnos alkaloids were the highest in the sand processing.. Processing of nux vomica plays a critical role in its toxicity.

Topics: Alkaloids; Animals; Cell Division; Chlorocebus aethiops; DNA; Drugs, Chinese Herbal; Hot Temperature; Magnoliopsida; Strychnine; Technology, Pharmaceutical; Vero Cells

Determination has been made on the contents of strychnine, brucine and ephedrin in different processed products of Strychnos nux-vomica. The acute toxicity, analgesic and antiphlogistic actions of these products have also been detected. The result shows that the product processed with Ephedra sinica can reduce toxicity and promote curative effect. Among the different processing methods the preparation with Ephedra and Liqorice root and the preparation with Ephedra and alcohol appear better and thus useful in practical application.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Drugs, Chinese Herbal; Ephedrine; Female; Hot Temperature; Lethal Dose 50; Magnoliopsida; Male; Mice; Plants, Medicinal; Random Allocation; Strychnine; Technology, Pharmaceutical

In mice, aversion to the bitter acetylated sugar sucrose octaacetate (SOA) is determined by a single genetic locus with three alleles. SWR/J (SW) inbred mice are SOA tasters: They avoid many compounds characterized as bitter-tasting by humans, at concentrations to which C3HeB/FeJ (C3:SOA demitasters) mice are less sensitive. C3.SW-Soa(a) congenic taster mice contain the taster allele transposed to a 99% C3 bitter-insensitive genetic background. SW, C3, C3.SW-Soa(a) congenic taster, and C3.SW demitaster mice were behaviorally tested with a series of 48-h two-bottle preference tests to determine the influence of the Soa(a) taster allele on sensitivity to a variety of bitter-tasting compounds. Soa allelic variation had a major effect on sensitivity to 0.003-1.0 mM SOA and several concentrations of the bitter-tasting alkaloids brucine, strychnine, and quinine. Effects were also found for 0.1 mM denatonium and 1 mM propylthiouracil. For caffeine, cycloheximide, thiamine, and two nonbitter compounds (NaCl and calcium hydroxide), the SW mice avoided lower concentrations than the other strains, but this avoidance was not due to the Soa(a) allele because both the C3 inbred and C3.SW-Soa(a) congenics were less sensitive. These results suggest the Soa gene product influences sensitivity to a subset of bitter-tasting compounds.

Topics: Alleles; Animals; Female; Food Preferences; Genes; Glycine Agents; Male; Mice; Mice, Inbred Strains; Quaternary Ammonium Compounds; Strychnine; Taste

To evaluate the effect of brucine (Bru) i.p. at analgesic doses on the nonspecific immune responses in normal and cyclophosphamide (Cyc)-treated mice.. The clearance of charcoal particles, the immune organ weights, the white blood cell counts in peripheral blood, the phagocytosis to neutral red (NR) of PMO and its IL-1 production in vitro were tested.. In normal mice, Bru slightly enhanced the clearance of charcoal particles, the phagocytosis of PMO, IL-1 production, the immune organ weights and the WBC counts (P > 0.05), whereas in Cyc-induced subnormal immunity model mice, Bru greatly enhanced these nonspecific immune responses (P < 0.05 or P < 0.01). The effects of Bru were most marked i.p. at 10 mg.kg-1 in vivo or 0.1-10 mg.L-1 in vitro.. Bru i.p. at an analgesic dosage has dose- and function-dependent immunoregulatory effects.

Topics: Adjuvants, Immunologic; Analgesics; Animals; Charcoal; Cyclophosphamide; Female; Immunocompromised Host; Interleukin-1; Leukocyte Count; Macrophages, Peritoneal; Male; Metabolic Clearance Rate; Mice; Mice, Inbred BALB C; Phagocytosis; Strychnine

This paper describes the determination of strychnine and brucine in the seeds, root, stem and leaves of Strychnos species by HPLC. The analytical column used was ZY110 YNG-C18. The mobile phase was KH2PO4(0.01 mol.L-1)--MeOH(73:27), pH2.5, regulated by 10% H3PO4. Flow rate was 1.0 ml.min-1. The detection wavelength was 264 nm. The linear ranges of strychmine and brucine were 0.18-7.26 micrograms and 0.11-4.32 micrograms, respectively. The recoveries of strychnine and brucine were 98.27% and 98.04%, respectively. The analytical results showed that the contents of strychnine and brucine in samples showed great difference between different species. The contents of strychnine in the seeds of Strychnos wallichiana and S. ignatii were 5.6% and 3.9%, respectively. These results show that the two Strychnos species may be developed as the resources of strychnine.

Topics: Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Magnoliopsida; Plants, Medicinal; Strychnine

We have indicated that strychnine and brucine induce concurrently an unknown P450 (2B-UP) which cross-reacts with anti-CYP2B1 antibody (Fujisaki et al., J. Pharmacol. Exp. Ther., 268, 1024-1031, 1994). We purified 2B-UP from brucine-treated rats and characterized it in this study. The purification was achieved by solubilization with sodium cholate followed by successive chromatographic steps with omega-aminooctyl-Sepharose 4B, DEAE-Sephacel and hydroxyapatite. The minimum molecular weight of purified 2B-UP was calculated to be 48000 by sodium dodecyl sulfate-gel electrophoresis. This preparation showed no Soret peak in the ultraviolet absorption spectrum indicating absence of heme. The amino terminal sequence of 2B-UP up to the 10th residue was consistent with the deduced amino acid sequence of CYP2B3 cDNA, but did not agree with the sequences of CYP2B1/2. The result strongly suggests that 2B-UP is CYP2B3. Thus, we indicated here that Strychnos alkaloid, brucine, is a potent inducer of the CYP2B3 or the closely related P450.

Topics: Amino Acid Sequence; Animals; Aryl Hydrocarbon Hydroxylases; Cytochrome P-450 Enzyme System; Cytochrome P450 Family 2; Enzyme Induction; Molecular Sequence Data; Rats; Strychnine

A flow-injection technique involving on-line catalytic digestion and spectrophotometric detection has been developed for the determination of iodine in urine. After urine samples are digested by KMnO4-K2Cr2O7-H2SO4 solution, the iodine in the urine catalyzes the reaction of As(III) with Ce(IV). The remaining Ce(IV) is then reacted with brucine and the product is detected with a spectrophotometer at 480 nm. With this technique, we obtained a detection limit for urinary iodine of 0.039 mumol/L, and the linear range was 0.039-7.88 mumol/L with a CV < 3%. Analytical recovery ranged between 92% and 104% (mean 99%). The sampling frequency of the flow-injection technique was 70/h. We applied the method to measure the iodine concentration in a freeze-dried urine reference sample and in collected urine samples, and compared the results with those obtained by the accepted alkaline ashing technique. The proposed technique has the advantages of being simple, rapid, precise, accurate, and sensitive. It can be used to assess iodine-deficient populations as well as those receiving treatment.

Topics: Arsenic; Catalysis; Cesium; Drug Stability; Female; Flow Injection Analysis; Freeze Drying; Humans; Iodine; Male; Manganese; Quality Control; Sensitivity and Specificity; Solutions; Spectrophotometry; Strychnine

A procedure for quantitative estimation of strychnine and brucine in the extracts of Strychnos nux-vomica seeds by capillary zone electrophoresis (CZE) was developed. The buffer solution used was 10mM phosphate buffer-MeOH (9:1), pH 2.5. The linear calibration range was 0.01-0.15 mg/ml. This method is useful for the qualitative and quantitative determination of strychnine and brucine in plant drug samples, as well as in human plasma.

Topics: Drugs, Chinese Herbal; Electrophoresis, Capillary; Feasibility Studies; Humans; Seeds; Strychnine

We used molecular modeling techniques to examine six reported antagonists of glycine with varying Ki values against strychnine. We found the data suggest two groups operating with different mechanisms. In group 1 (strychnine, brucine, Pitrazepin, and bicuculline methobromide) the antagonist contains two or three sites that can electrostatically bind to the three comparable groups of opposite charge in the recognition site where the natural neurotransmitter binds, thus opening the chloride channel. In addition, when in this position, the antagonist is able to also block the now opened chloride channel with a different portion of its structure. In many cases, this involves an interaction between a carbonyl group on the antagonist and the guanidinium group of arginine which is part of the polypeptide segment of the outer mouth of the chloride channel (Grenningloh et al., Nature 330:25-26, 1987). In group 2 (R5135 and 1,5-diphenyl-3,7-diazaadamantan-9-ol) the antagonist contains charged sites but when one of these molecules attaches to the recognition site, the chloride channel is not opened. In addition, R5135 contains a carbonyl group which attaches to arginine as pointed out in the text, whereas 1,5-diphenyl-3,7-diazaadamantan-9-ol contains a phenyl group that can block the channel.

Topics: Animals; Binding, Competitive; Dibenzazepines; GABA Antagonists; Glycine; Models, Molecular; Molecular Structure; Receptors, Glycine; Strychnine

The purpose of this work was to define the pharmacology of an intestinal epithelial [3H]strychnine binding site. Strychnine, brucine, verapamil and desmethoxyverapamil bind to small intestinal mucosal homogenates with nanomolar affinity at a site not related to the strychnine receptor, which is in the spinal cord. The antidiarrheal agents, fluperamide and loperamide, and several alkaloids have an order of magnitude lower affinity. Agents that bind to cytochrome P450IID6 also displace [3H]strychnine binding, which implies that the binding site may have some properties similar to the catalytic site of this cytochrome P450 enzyme.

Topics: Animals; ATP Binding Cassette Transporter, Subfamily B, Member 1; Binding Sites; Carrier Proteins; Cystic Fibrosis Transmembrane Conductance Regulator; Intestine, Small; Male; Membrane Proteins; Rats; Rats, Sprague-Dawley; Strychnine

Inbred and congenic strains exhibited several patterns of relative sensitivity to bitter tastants in 48-h, two-bottle preference tests. With segregation analyses of descendents of crosses between contrasting strains, these patterns suggested at least three genetic loci influencing bitter perception. The extensively characterized Soa (sucrose octaacetate) locus underlies one pattern. Variation at this locus had pleiotropic effects on avoidance of other acetylated sugars, plus such structurally dissimilar bitter tastants as brucine, denatonium benzoate, and quinine sulfate. Unlike SOA, however, sensitivity to quinine sulfate was polygenically determined, and produced a second characteristic pattern. At least one, possibly several, additional unlinked loci contributed to quinine differences. Phenylthiocarbamide (PTC) aversion differences exemplified a third pattern. Segregation consistent with monogenic control of PTC aversion has been reported, and within segregating populations PTC aversion did not covary with SOA or quinine sulfate avoidance. Variants of the three major patterns may be useful for analysis of specific mechanisms. While both showed the SOA pattern, strychnine differences were markedly smaller than brucine (dimethoxystrychnine) differences. Likewise, a hop extract containing primarily iso-alpha acids (e.g., isohumulone) produced an SOA-like pattern, while an extract with nonisomerized alpha-acids (e.g., humulone) did not.

Topics: Animals; Chromosome Mapping; Cyclohexenes; Cyclopentanes; Dose-Response Relationship, Drug; Female; Food Preferences; Male; Mice; Mice, Inbred Strains; Phenotype; Phenylthiourea; Quaternary Ammonium Compounds; Species Specificity; Strychnine; Sucrose; Synaptic Transmission; Taste; Taste Buds; Taste Threshold; Terpenes

The contents of strychnine, brucine, isostrychnine and isobrucine in different processed products of Strychnos nux-vomica were determined by TLC-densitometry. The relationship of the contents of strychnos alkaloids with processing methods was studied.

Topics: Chromatography, Thin Layer; Densitometry; Drugs, Chinese Herbal; Hot Temperature; Methods; Strychnine; Technology, Pharmaceutical

Topics: Strychnine; Strychnos nux-vomica

Topics: Aminobutyrates; gamma-Aminobutyric Acid; Pharmacology; Rats; Research; Seizures; Strychnine; Synapses; Toxicology

Topics: Alkaloids; Arsenicals; Cerium; Colorimetry; Iodine; Research; Strychnine; Temperature; Thyroid Function Tests; Zinc

Topics: Alkaloids; Chemistry, Pharmaceutical; Chromatography; Chromatography, Thin Layer; Pharmacy; Radiometry; Research; Strychnine

Topics: Alkaloids; Aniline Compounds; Anisoles; Humans; In Vitro Techniques; Liver; Metabolism; Microsomes; Research; Strychnine; Strychnos nux-vomica

Topics: Alkylation; Chromatography; Feces; Glucuronates; Metabolism; Rabbits; Research; Spectrophotometry; Strychnine; Urine

Topics: Alkaloids; Chromatography; Chromatography, Thin Layer; Research; Strychnine; Toxicology

The apparatus and technique used in the preparation and observation of explants of brain tissue capable of producing spontaneous potentials in vitro are described. The magnitude and pattern of spontaneous potentials from explants of telencephalon of 15 day chick embryos (measured using external bare platinum electrodes) and some aspects of their "normal" behavior during 12 days in vitro are also described. No change was noted in these potentials with change of amplifiers, recorders, or electrodes. The response of the potentials to change in temperature and proportionate composition of the atmosphere around the explant was such as to suggest that the potentials arise as a result of a living process. The changes brought about by the administration of anesthetics, strychnine, brucine, and barbiturates were those that might be anticipated in a normal functional activity of the central nervous system. It is concluded that these potentials are a true physiological phenomenon and arise from living cells of the central nervous system.

Topics: Animals; Brain; Central Nervous System; Chick Embryo; In Vitro Techniques; Strychnine; Telencephalon

Topics: Alkaloids; Strophanthins; Strychnine

Topics: Strophanthins; Strychnine; Strychnos nux-vomica

Topics: Electrophoresis, Paper; Strychnine; Strychnos nux-vomica

Topics: Humans; Strophanthins; Strychnine; Strychnos nux-vomica; Telencephalon

Topics: Alkaloids; Strychnine; Strychnos nux-vomica

Topics: Alkaloids; Strychnine

Topics: Chromatography, Paper; Strychnine; Strychnos nux-vomica

Topics: Humans; Pharmaceutical Solutions; Seeds; Strychnine; Strychnos nux-vomica

Topics: Alkaloids; Atropine; Cocaine; Electrophoresis; Morphine Derivatives; Purines; Strychnine

Topics: Chromatography, Paper; Strychnine; Strychnos nux-vomica

Topics: Ammonium Compounds; Colorimetry; Indicators and Reagents; Seeds; Strychnine; Strychnos; Strychnos nux-vomica; Thiocyanates

Topics: Body Fluids; Feces; Humans; Nitrates; Strychnine

Topics: Alkaloids; Humans; Morphine; Spectrophotometry; Strychnine; Viscera

Topics: Alkaloids; Biological Assay; Humans; Spectrophotometry; Strychnine; Viola

Topics: Cadmium; Climate; Humans; Nephelometry and Turbidimetry; Strychnine

Topics: Benzene; Crystallization; Solvents; Strychnine; Thermodynamics

Topics: Humans; Strychnine

Topics: Humans; Strychnine

Topics: Humans; Strychnine

Topics: Strychnine

Topics: Humans; Strychnine

Topics: Humans; Strychnine

Topics: Humans; Strychnine

Topics: Strychnine

Topics: Humans; Strychnine

Topics: Acids; Humans; Strychnine