wogonoside and baicalin

Glucuronidation is one of the major metabolic pathways for flavonoids. However, quantification of flavonoid glucuronides in biological samples, especially in the bile, is sometimes challenging due to signal suppression by bile acids. The purpose of this study is to establish a robust LC-MS/MS method for directly measuring flavonoid glucuronides in bile and blood. Wogonoside (wogonin-7-O-glucuronide), baicalin (baicalein-7-O-glucuronide) and apigenin-7-O-glucuronide were used as the model compounds and taurocholic acid (T-CA) were used as the model bile acid to establish the method. Bile samples were processed using solid phase extraction (SPE) and blood samples were prepared using protein precipitation method. The analytes were separated on a Resteck HPLC (50 mm × 2.1 mm ID, 1.7 μm) column using acetonitrile and 0.1% formic acid in water as the mobile phases. The mass analysis was performed in an AB Sciex 5500 Qtrap mass spectrometer via multiple reaction monitoring (MRM) in the positive mode. The results showed that the linear range of the above three analytes were 10 nM-5000 nM in the bile and 1.56 nM-4000 nM in the blood, respectively. The recoveries of three glucuronides were >85% and the matrix effects were <20% at low, medium and high concentrations in the bile and the blood. The results also showed that >90% of these bile acids were removed by the selected SPE procedure to facilitate glucuronide analysis. The validated method was successfully applied to a portal vein infusion study using rats to quantify baicalin, wogonoside, and apigenin-glucuronide in bile and blood samples.

Topics: Animals; Apigenin; Bile; Chromatography, High Pressure Liquid; Flavanones; Flavonoids; Glucosides; Male; Portal Vein; Rats; Rats, Wistar; Tandem Mass Spectrometry

An efficient and target-oriented pH-peak-focusing countercurrent chromatographic method was established for large-scale separation of baicalin and wogonoside from the crude exact of traditional Chinese medicinal herb Scutellaria baicalensis Georgi. An optimized two-phase solvent system composed of n-butanol-ethyl acetate-methanol-water (1:4:0.5:5, v/v) was selected. Trifluoroacetic acid (10 mmol/L) was added to the upper organic phase, used as the stationary phase. One liter of the aqueous lower phase was used as the mobile phase for 0-350 min, and then 10 mmol/L ammonia was added to remaining 1 L of the aqueous lower phase and used as the mobile phase for 350-600 min. In total, 493.2 mg of baicalin with 98.6% purity and 88.6 mg of wogonoside with 98.9% purity were obtained from 1.0 g of crude exact of S. baicalensis by countercurrent chromatography in a single run. The acid dissociation constant (pKa) and oil-water partition coefficient values of two components were measured to better understand the mechanism of separation. Results showed that pH-peak-focusing countercurrent chromatography with a polar solvent system added with trifluoroacetic acid could be an efficient method for large-scale isolation of organic acids, which are difficult to separate with conventional countercurrent chromatography due to their poor solubility in non-polar solvents.

Topics: Countercurrent Distribution; Flavanones; Flavonoids; Glucosides; Hydrogen-Ion Concentration; Medicine, Chinese Traditional; Plant Extracts; Scutellaria baicalensis

Topics: Chemistry Techniques, Analytical; Countercurrent Distribution; Flavanones; Flavonoids; Glucosides; Hydrogen-Ion Concentration; Scutellaria baicalensis; Solvents

Topics: Cluster Analysis; Discriminant Analysis; Flavanones; Flavonoids; Glucosides; Least-Squares Analysis; Principal Component Analysis; Scutellaria baicalensis; Xylem

The increasing demand of Chinese materia medica could not be supplied by wild resource, and the cultivated medicinal materials become popular, which led to decreased quality of many medicinal materials due to the difference of the circumstance between the wild and the cultivated. How to improve quality becomes key points of Chinese medicine resource. The leaves of Scutellaria baicalensis were sprayed with H₂O₂, the activities of superoxide dismutase (SOD) and catalase (CAT) changed little, but there had been a marked decrease of peroxidase (POD) and ascorbic oxidase (APX), which showed that the antioxidase system declined. Meanwhile, H₂O₂, as enhanced the expression of phenylalnine ammonialyase (PAL) and β-glucuronidase (GUS) as well as activity of PAL, promoted the biosynthesis and biotransformation of flavonoids. At the day 2 after treated, H₂O₂ of 0.004 μmol·L⁻¹ the contents of the baicalin and the wogonoside decreased slightly, but the contents of the baicalein and the wogonin increased significantly, the baicalein from 0.094% to 0.324%, the wogonin from 0.060% to 0.110%, i. e. increased 246% and 83.3%, respectively.

Topics: Ascorbate Oxidase; Catalase; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Glucuronidase; Hydrogen Peroxide; Peroxidase; Phenylalanine Ammonia-Lyase; Scutellaria baicalensis; Secondary Metabolism; Superoxide Dismutase

Scutellariae Radix (SR) and its bioactive flavones elicit a variety of effects in the brain. However, the brain uptake of individual SR flavones and its relationship to the elicited effects after SR administration remain unknown. Moreover, previous studies seldom measured pharmacokinetic and pharmacodynamic outcomes simultaneously. In the current study, the brain uptake of six major SR flavones and the anxiolytic behavior following oral administration of a SR extract at two clinically relevant doses (600 and 1200 mg/kg twice daily) were simultaneously investigated in mice (n = 18 per group). Brain and plasma concentrations of the flavones were measured by LC-MS/MS, while the anxiolytic effect was evaluated using the elevated plus maze. To further investigate the mechanism behind the differential brain uptake of the six SR flavones, these flavones were separately administered to mice at an equivalent molar oral dose (n = 6). The brain tissue bindings of the SR flavones were also measured with the in vitro brain slice method. Our results indicated that all six SR flavones including three aglycons (baicalein, wogonin, and oroxylin A) and three glucuronides (baicalin, wogonoside, and oroxyloside) could pass through the blood-brain barrier, with brain concentrations ranging from 7.9 to 224.0 pmol/g. It provided novel evidence that oroxylin A had the highest brain uptake among the six SR flavones regardless of its limited content in SR extract, in which 3.6-3.9% of the administered oroxylin A dose was present in the brain 6 h postdosing and with a brain-to-plasma ratio of 0.42-0.46. Although SR extract contains flavones that are positive modulators of the benzodiazepine binding site of GABA

Topics: Animals; Anti-Anxiety Agents; Brain; Flavanones; Flavones; Flavonoids; Glucosides; Male; Maze Learning; Mice; Mice, Inbred ICR; Scutellaria baicalensis; Tandem Mass Spectrometry

Baicalin and wogonoside are two of the most abundant flavonoid glycosides in the root of Scutellaria baicalensis Georgi, which is a widely used peroral herbal medicine with anticancer, antiviral, antibacterial and anti-inflammatory properties. In the present study, the effects of intestinal microecology on the metabolism and pharmacokinetics of orally administered baicalin and wogonoside were investigated by UPLC-QTOF/MS measurement of the difference in metabolites between normal and antibiotic-pretreated rats. In the antibiotic-pretreated rats, the plasma concentration-time profile and pharmacokinetic parameters of the two flavonoid glycosides and their relevant aglycone forms were significantly changed compared with those in normal rats. Further, hydrolysis and glucuronidated metabolites were not detected in the cecum contents and urine samples from antibiotic-pretreated rats. These results suggested that intestinal microbiota may play a key role in the pharmacokinetics and metabolism of peroral baicalin and wogonoside. According to our findings, it is recommended that the root of S. baicalensis should not be co-administered with antibiotics in clinical use.

Topics: Animals; Cecum; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Gastrointestinal Microbiome; Glucosides; Male; Mass Spectrometry; Plant Roots; Rats; Rats, Sprague-Dawley; Scutellaria baicalensis

This paper is aimed to study plant biomass and active compounds of Scutellaria baicalensis germchit in different five stages (from germination to transplant). The length of shoot and root, the diameter and the weight of root were determined. HPLC method was used to determine the content of active compounds (baicalin, scutellarin, wogonoside, baicalein, wogonin). According to the results, various biological indicators increased with the germination of seedling. However, the drying rate of the root declined to 27.96% from 32.90%. The contents of scutellarin and baicalein increased firstly, and then decreased. The maximums of them were 3.22,3.89 mg•g⁻¹ while the data of shoot/root was 0.35. The maximums of the contents of baicalin and wogonoside were 107.39,16.11 mg•g⁻¹ while the data of shoot/root was 0.23 and 0.06. The contents of wogonin gradually increased to the maximum of 0.88 mg•g⁻¹ while the data of shoot/root was 0.50. In conclusion, the contents of baicalin, scutellarin, wogonoside, baicalein and wogonin reached or approached the maximum at germination stage while the data of shoot/root was 0.35. The rate of shoot and root can be used as a judging index of active compounds for S. baicalensis germchit.

Topics: Biomass; Chromatography, High Pressure Liquid; Flavanones; Flavonoids; Germination; Glucosides; Plant Extracts; Plant Roots; Scutellaria baicalensis; Seedlings

A simple and accurate high-performance liquid chromatography-photodiode array (HPLC-PDA) detection method has been developed and validated for simultaneous determination of nine components-liquiritin, coptisine, baicalin, palmatine, berberine, wogonoside, baicalein, glycyrrhizin and wogonin-in the traditional Korean formula, Banhasasim-tang decoction. A Gemini C18 analytical column was used to separate the nine constituents and kept at 40°C by gradient elution with 0.1% (v/v) trifluoroacetic acid in distilled water (A) and acetonitrile (B) as mobile phases. The flow rate was 1.0 mL/min and the injection volume was 10 µL. The PDA detection wavelengths were set at 254, 275 and 350 nm. Calibration curves of all compounds showed good linearity with coefficients of determination ≥0.9998 within the test ranges. The limits of detection and quantification of all compounds were in the range 0.01-0.09 and 0.03-0.30 µg/mL, respectively. All recoveries of the nine marker compounds ranged from 98.65 to 103.22% with relative standard deviation (RSD) values <1.25%. The RSDs of intraday and interday precision were <1.13 and 1.83%, respectively. The concentrations of the nine marker constituents were 0.19-41.09 mg/g.

Topics: Acetonitriles; Berberine; Berberine Alkaloids; Chromatography, High Pressure Liquid; Flavanones; Flavonoids; Glucosides; Glycyrrhizic Acid; Humans; Limit of Detection; Medicine, Korean Traditional; Observer Variation; Pinellia; Plants, Medicinal; Reproducibility of Results; Solvents; Trifluoroacetic Acid; Water

Scutellariae Radix (SR) has been extensively prescribed in folk medicines due to its notable beneficial activities. The flavonoid glucuronides baicalin (BG), wogonoside (WG), oroxylin A 7-O-β-d-glucuronide (OG) and their aglycones baicalein, wogonin and oroxylin A, are the main components of the herb. So far, majority of previous studies failed to report the pharmacokinetics and none offered an explanation for the systemic exposures of these six flavonoids when the herbal extract was orally administered. In this study, when a SR extract was orally dosed to rats (800mg/kg, equivalent to BG 324.80, WG 124.00, OG 43.04, baicalein 25.36, wogonin 24.40, and oroxylin A 5.79mg/kg), all six flavonoids were detectable throughout the experimental period (48h) using an LC-MS/MS method with the Cmax and AUC0-48h of the glucuronides 10-130 times that of respective aglycones. As the lowest among the three glucuronides in the herb, OG was the most abundant in vivo, while the systemic exposure of wogonin was the highest amongst the three aglycones. The dose-normalized AUC0-48h descended in orders of OG/oroxylin A, WG/wogonin and BG/baicalein. Two di-conjugates of baicalein (BG glucuronide and BG glucoside), two BG isomers (minor BM1 and major BM2), and one WG isomer (wogonin 5-O-glucuronide) were detected in rat plasma. Semi-quantitation of the isomers with peak area data revealed that the AUPs (area under peak area ratio-time curves) of BG isomers were ∼3 times that of BG, yet the AUP of wogonin 5-O-glucuronide was only one seventh of WG. BM2, tentatively assigned as baicalein 6-O-glucuronide, was formed from both microbial isomerization of BG and hepatic glucuronidation of baicalein. Wogonin 5-O-glucuronide was only formed in hepatic glucuronidation of wogonin. Demethylated wogonin was observed in gut bacteria, offering an optional origin of BM1 apart from baicalein glucuronidation. Microbial isomerization of BG and extensive hepatic glucuronidation of baicalein to form BM2as well as a poorer intestinal permeability of baicalein (Papp×10(-6)cm/s) should account for the lower systemic exposures of BG and baicalein. Faster microbial hydrolysis of WG, high intestinal permeability (Papp×10(-5)cm/s) and less hepatic glucuronidation of wogonin explain the relatively high systemic exposure of wogonin. Sole microbial deglycosylation of OG, high intestinal permeability (Papp×10(-5)cm/s) and extensive hepatic glucuronidation of oroxylin A supported the highest systemic exposure

Topics: Administration, Oral; Animals; Caco-2 Cells; Chromatography, High Pressure Liquid; Flavanones; Flavones; Flavonoids; Glucosides; Glucuronides; Humans; Male; Plant Extracts; Rats; Rats, Sprague-Dawley; Scutellaria baicalensis; Tandem Mass Spectrometry

Chlorophyll-type contaminants are commonly encountered in the isolation and determination of flavones of plant aerial plant parts. Heme is also a difficult background substance in whole blood analysis. Both chlorophyll and heme are porphyrin type compounds. In this study, a rapid method for isolating flavones with 5-hydroxyl or ortho-hydroxyl groups from biological samples was developed based on the different solubilities of porphyrin-metal and flavone-metal complexes. It is important that other background substances, e.g., proteins and lipids, are also removed from flavones without an additional processing. The recoveries of scutellarin, baicalin, baicalein, wogonoside and wogonin, which are the primary constituents of Scutellaria baicalensis (skullcaps) were 99.65% ± 1.02%, 98.98% ± 0.73%, 99.65% ± 0.03%, 97.59% ± 0.09% and 95.19% ± 0.47%, respectively. As a sample pretreatment procedure, this method was coupled to high-performance liquid chromatography (HPLC) with good separation, sensitivity and linearity and was applied to determine the flavone content in different aerial parts of S. baicalensis and in dried blood spot samples.

Topics: Apigenin; Chromatography, High Pressure Liquid; Dried Blood Spot Testing; Flavanones; Flavones; Flavonoids; Glucosides; Glucuronates; Humans; Plant Components, Aerial; Plant Extracts; Scutellaria baicalensis; Zinc Acetate

We evaluated the antioxidant properties of four main flavones from Scutellaria baicalensis: baicalein, wogonin and their glucuronides - baicalin and wogonoside. We used three in vitro assays: free radical scavenging with 2,2'-diphenylpicrylhydrazyl radical, transition metal ions reducing power by phosphomolybdenum assay and inhibition of the hydroxyl radical-induced peroxidation of linoleic acid assay. All flavones have antioxidant capacity, which differs depending on the structure and mechanisms of activity. In all tests, only baicalein - the aglycone with three adjacent hydroxyl groups - exhibited consistent antioxidant effect. Wogonin protected linoleic acid against oxidation. Baicalin displayed less potent antioxidant properties whereas wogonoside did not have significant antioxidant activity.

Topics: Antioxidants; Flavanones; Flavones; Flavonoids; Free Radical Scavengers; Glucosides; Molecular Structure; Plant Extracts; Plant Roots; Scutellaria baicalensis

A novel free radical reaction combined with high-performance LC-photodiode array-ESI-MS/MS screening method was developed for the detection and identification of natural antioxidants from whole Scutellariae. Six compounds and whole Scutellariae extracts were found to possess a potential antioxidant capacity, and their free radical-scavenging activities were investigated in detail. The six compounds were identified as baicalin, baicalein, scutellarin, scutellarein, wogonoside and chrystin-7-glucoronide. The present study reveals that the radical-scavenging capacities of the whole Scutellariae extracts are as follows: hydroxyl radical > superoxide radical > peroxy radical. Wogonoside showed the strongest capability for scavenging hydroxyl radical. Baicalein not only showed the strongest capability for scavenging superoxide radical but also showed capability for lipid radical; Scutellarein (Peak 5) exhibited the highest reactivity in the lipid peroxidation processes. Based on these studies, the current paper accomplishes the evaluation of activities of some important anti-radical substances extracted from the radix, stem and leaf of the whole Scutellariae.

Topics: Chromatography, Liquid; Flavanones; Flavonoids; Free Radical Scavengers; Free Radicals; Glucosides; Plant Extracts; Plant Leaves; Plant Stems; Scutellaria baicalensis; Tandem Mass Spectrometry

We determined the in vitro antioxidant activity of methanolic extracts from the shoots and roots of Scutellaria species (S. altissima and S. alpina) against the action of strong oxidants: hydrogen peroxide (H2O2) and H2O2+Fe(2+) (donor of hydroxyl radicals) on plasma proteins and lipids. Lipid peroxidation in human plasma was measured by the level of thiobarbituric acid reactive species (TBARS). Protein oxidation was measured by quantitation of thiol group. We observed that the extracts (5-50 µg ml(-1)) containing phenolic compounds from both Scutellaria species distinctly reduced oxidation of lipids and proteins in human plasma treated with H2O2. These results also indicated that the extracts have a protective effect against oxidative damage to the human plasma lipids and proteins by induced hydroxyl radical. The main components of the plant materials analysed were flavonoids, present as aglycones (luteolin) or glycosides (cynaroside, baicalin, wogonoside). In all of the extracts, the phenylethanoid verbascoside was also found. The properties of the tested plant extracts were also compared with the action of a well characterised commercial antioxidative polyphenolic extract from the berries of Aronia melanocarpa (Aronox(®)). The comparative studies indicated that the analysed plant extracts were comparable to or even more effective in reducing the oxidation processes than the A. melanocarpa extract. The present study suggests that natural extracts from S. altissima and S. alpina have antioxidant activities and, therefore, may be beneficial in the prevention of diseases related to oxidant stress, such as cancer, cardiovascular, and inflammatory diseases.

Topics: Antioxidants; Blood; Flavanones; Flavonoids; Glucosides; Humans; Hydrogen Peroxide; Lipid Peroxidation; Molecular Structure; Oxidative Stress; Phenols; Plant Extracts; Plant Roots; Plant Shoots; Scutellaria

This experiment aimed to explore and research the process of preparing baicalein and wogonin through liquid fermentation with Bacillus natto. Active enzymes of produced by B. natto was used for the biological transformation of baclin and wogonoside, in order to increase the content of the haicalein and wogonin in the scutellaria. With the content of the baicalein and wogonin as evaluating indexes, the effects of carbon source, nitrogen source, the types and suitable concentration of inorganic salt, medium pH, granularities of medical materials, liquid volume in flask, shaking speed, liquid-to-solid ratio, fermentation time on the fermentation process were studied. The optimal process conditions for liquid fermentation of scutellaria were 1.0% of peptone, 0.05% of NaCl, pH at 6, the granularities of medical materials of the scutellaria screened through 40-mesh sifter, 33% of liquid, shaker incubator speed at 200 r x min(-1), liquid-to-solid ratio of 5:1, temperature at 37 degrees C, fermentation for 6 days, baclin's conversion rate at 97.6% and wogonoside's conversion rate at 97% in the scutellaria. According to the verification test, the process was stable and feasible, and could provide data reference for the industrial production.

Topics: Bacillus subtilis; Biotransformation; Fermentation; Flavanones; Flavonoids; Glucosides; Soy Foods

Pharmacochemistry and integrated pharmacokinetics of six alkaloids (groenlandicine, berberine, palmatine, epiberberine, jatrorrhizine, and columbamine) after oral administration of Huang-Lian-Jie-Du-Tang (HLJDT) decoction were investigated in this paper. The method of plasma pharmacochemistry was applied to predict the potential bioactive components in HLJDT decoction. Based on the accurate molecular weight, 10 components including 2 flavonoids (baicalin and wogonoside), 1 iridoid glycoside (geniposide), and 7 alkaloids (above-mentioned 6 alkaloids and coptisine) were structurally identified. Then, integrated pharmacokinetics of the alkaloids in Sprague-Dawley rats after oral administration of HLJDT decoction was investigated by HPLC method. The results showed that the pharmacokinetic behaviors of the alkaloids were different although their chemical structures were similar. This study developed a method to predict the potential bioactive components in HLJDT decoction and research the pharmacokinetic behaviors of the potential bioactive components.

Topics: Administration, Oral; Alkaloids; Animals; Berberine; Berberine Alkaloids; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Iridoids; Molecular Structure; Rats; Rats, Sprague-Dawley

This study aims to identify and quantify the six major bioactive flavones of the traditional Chinese medicine Scutellariae Radix (RS), including baicalein, baicalin, wogonin, wogonoside, oroxylin A and oroxyloside in rat after oral administration of a standardized RS extract. A novel, sensitive and selective method for simultaneous determination of these six analytes in rat brain and plasma using solid phase extraction-liquid chromatography-tandem mass spectrometry (SPE-LC/MS/MS) was developed and fully validated. The lower limits of quantification (LLOQs) for the six RS flavones in brain tissue were 0.02nmol/g. The LLOQs in plasma were 0.005nmol/ml for B, W and OA, 0.025nmol/ml for WG and OAG, and 0.1875nmol/ml for BG. The current study provides novel evidence of the presence of all the tested RS flavones and an isoform of BG (BG', probably baicalein-6-O-glucuronide) in the rat brain after oral administration of RS extract, suggesting their ability to permeate through the blood-brain barrier. The method was also successfully applied to the pharmacokinetic study of all these analytes in plasma after oral administration of RS extract (300mg/kg) to Sprague-Dawley rats. The developed assay method provides a useful tool for both preclinical and clinical investigations on the disposition of RS flavones in brain and plasma.

Topics: Animals; Brain; Chromatography, Liquid; Flavanones; Flavones; Flavonoids; Glucosides; Glucuronides; Male; Rats; Solid Phase Extraction; Tandem Mass Spectrometry

Huanglian Jiedu decoction (HLJDD) is used traditionally in China for the treatment of diabetes mellitus in clinical practice, which has been proved to be effective. The purpose of this study was to investigate the pharmacokinetic characteristics (especially the area under the curve, AUC) of baicalin and wogonoside in type 2 diabetic rats after oral administration of HLJDD extract and to explore its possible mechanism.. HLJDD extract and Radix scutellariae extract were prepared and the contents of baicalin and wogonoside contained in two extracts were assayed with high performance liquid chromatography (HPLC). Type 2 diabetic rats were induced by high fat diet and intraperitoneal injection of streptozotocin. Pharmacokinetics of baicalin and wogonoside in type 2 diabetic and normal control rats after oral administration of HLJDD extract or Radix scutellariae extract were investigated. Pharmacokinetics of baicalin in type 2 diabetic and normal rats after oral administration of pure baicalin was also investigated.. The pharmacokinetic parameters (especially AUCs) of baicalin and wogonoside in type 2 diabetic rats after oral administration of HLJDD extract were remarkably different from those in normal rats. And the alterations of the AUCs of baicalin and wogonoside in type 2 diabetic rats after oral administration of Radix scutellariae extract were similar to those after oral administration of HLJDD extract. Moreover, the increase of the AUC of baicalin in type 2 diabetic rats after oral administration of pure baicalin was similar to that after oral administration of HLJDD extract or Radix scutellariae extract.. The pharmacokinetic behaviors of baicalin and wogonoside (especially the systemic exposure [AUCs] of baicalin and wogonoside) were significantly altered in type 2 diabetic rats after orally administrated HLJDD extract. And the increased AUCs of baicalin and wogonoside in type 2 diabetic rats after oral administration of HLJDD extract resulted from neither the effects of other herbs contained in HLJDD nor the effects of other components contained in Radix scutellariae. It might result from the effects of the pathological status of type 2 diabetes mellitus.

Topics: Administration, Oral; Animals; Area Under Curve; Chromatography, High Pressure Liquid; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 2; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Hypolipidemic Agents; Male; Medicine, Chinese Traditional; Plant Extracts; Rats; Rats, Sprague-Dawley; Scutellaria baicalensis

The pharmacodynamic (PD) and pharmacokinetic (PK) properties of Huangqin Tang (HQT) were investigated in yeast-induced febrile rats. Blood sample and rectal temperature data of the rats were collected at different times after single oral administration of HQT at 20 g x kg(-1). The plasma concentrations of paeoniflorin, baicalin, wogonoside, baicalein, wogonin, oroxylin A, glycyrrhizic acid and glycyrrhetinic acid were quantified by a sensitive liquid chromatography-tandem mass spectrometric (LC-MS) method. The blood concentrations of PGE2, 1L-1β and TNF-α were detected by radioimmunoassay (RIA). All pharmacokinetic parameters were processed by non-compartmental analysis using WinNonlin software. The potential relationship between the mean concentration of eight constituents and the antifebrile efficacy was investigated by calculating Pearson correlation coefficients. It was found that HQT had significant antifebrile efficacy in yeast-induced febrile rats, but had no effect to normal rats. The antifebrile effect of HQT can be attributed to the inhibition of PGE2, 1L-1β and TNF-α. The constituents (baicalin, wogonoside, baicalein, wogonin, oroxylin A, glycyrrhizic acid and glycyrrhetinic acid) in febrile rats had delayed absorption and elimination, a longer residence time in the body, and higher C(max) and AUC than those in normal rats. Febrile condition could affect the pharmacokinetic behaviour of HQT in vivo; the flavonoids with the same backbone showed the similar fate in the body; baicalein and wogonin had a strong positive correlation (R > 0.66, P ≤ 0.02) with the antifebrile efficacy determined. Together, these constituents demonstrated different pharmacokinetic properties in the febrile body.

Topics: Administration, Oral; Animals; Area Under Curve; Chromatography, Liquid; Dinoprostone; Drugs, Chinese Herbal; Fever; Flavanones; Flavonoids; Glucosides; Interleukin-1beta; Mass Spectrometry; Monoterpenes; Rats; Tumor Necrosis Factor-alpha

Scutellaria baicalensis Georgi (Labiatae) is one of the most commonly used medicinal herbs, especially in traditional Chinese medicine. However, compared to many pharmacological studies of this botanical, much less attention has been paid to the quality control of the herb's pretreatment prior to extract preparation, an issue that may affect therapeutic outcomes.. The current study was designed to evaluate whether different pretreatment conditions change the contents of the four major flavonoids in the herb, i.e., two glycosides (baicalin and wogonoside) and two aglycones (baicalein and wogonin).. A high-performance liquid chromatography assay was used to quantify the contents of these four flavonoids. The composition changes of four flavonoids by different pretreatment conditions, including solvent, treatment time, temperature, pH value and herb/solvent ratio were evaluated.. After selection of the first order time-curve kinetics, our data showed that at 50 °C, 1:5 herb/water (in w/v) ratio and pH 6.67 yielded an optimal conversion rate from flavonoid glycosides to their aglycones. In this optimized condition, the contents of baicalin and wogonoside were decreased to 1/70 and 1/13, while baicalein and wogonin were increased 3.5- and 3.1-fold, respectively, compared to untreated herb.. The markedly variable conversion rates by different pretreatment conditions complicated the quality control of this herb, mainly due to the high amount of endogenous enzymes of S. baicalensis. Optimal pretreatment conditions observed in this study could be used obtain the highest level of desired constituents to achieve better pharmacological effects.

Topics: Chemical Fractionation; Chromatography, High Pressure Liquid; Drug Stability; Flavanones; Flavonoids; Glucosides; Hydrogen-Ion Concentration; Kinetics; Phytotherapy; Plant Extracts; Plants, Medicinal; Quality Control; Scutellaria baicalensis; Solvents; Temperature

Scutellaria baicalensis has been used as a traditional herbal medicine for bronchitis, hepatitis, and allergic diseases. The root of Scutellaria baicalensis contains active flavonoid components, including baicalin, baicalein, wogonoside, and wogonin, which have pharmaceutical properties. In the present study, the antiallergic properties of a standardized aqueous extract of S. baicalensis were evaluated, and the skin toxicity of its dermal application was also determined. The in vivo and in vitro assays were performed by using the β-hexosaminidase assay in rat basophilic leukemia cells (RBL-2H3) and cutaneous skin reaction in BALB/c mice, respectively. In addition, the acute dermal irritation/corrosion test was carried out in New Zealand white rabbits, and the skin sensitization test was conducted by Buhler's method in Hartley guinea pigs to estimate the safety of the standardized aqueous extract of S. baicalensis for topical application. β-Hexosaminidase release in RBL-2H3 was markedly decreased following treatment with the standardized aqueous extract of S. baicalensis. It also ameliorated antigen-induced ear swelling compared with the control group in BALB/c mice. In the toxicological studies, it did not induce any dermal irritation/corrosion in rabbits or skin sensitization in guinea pigs. Although still limited, these results concerning the toxicological effects of S. baicalensis could be an initial step toward the topical application of S. baicalensis extracts on hypersensitive skin.

Topics: Administration, Topical; Animals; Anti-Allergic Agents; beta-N-Acetylhexosaminidases; Cell Line, Tumor; Flavanones; Flavonoids; Glucosides; Guinea Pigs; Hypersensitivity; Male; Mice; Mice, Inbred BALB C; Plant Extracts; Plant Roots; Plants, Medicinal; Rabbits; Rats; Scutellaria baicalensis; Skin

Scutellaria baicalensis (SB), a traditional herb with high pharmacological value, contains more than 10% flavone by weight. To improve the biological activity of flavones in SB, we aimed to enhance the bioconversion of baicalin (BG) to baicalein (B) and wogonoside (WG) to wogonin (W) in SB during fermentation using beta-glucuronidase produced from Lactobacillus brevis RO1. After activation, L. brevis RO1 was cultured in milk containing SB root extract with various carbon or nitrogen sources at 37°C for 72 h. During fermentation, the growth patterns of L. brevis RO1 and changes in the flavone content were assessed using thin-layer chromatography and high-performance liquid chromatography. After 72 h of fermentation, the concentrations of B and W in the control group increased by only 0.15 and 0.12 mM, respectively, whereas they increased by 0.57 and 0.24 mM in the fish peptone group. The production of B and W was enhanced by the addition of 0.4% fish peptone, which not only improved the growth of L. brevis RO1 (p < 0.001) but also enhanced the bioconversion of flavones. In conclusion, the bioconversion of flavones in SB may provide a potential application for the enhancement of the functional components in SB.

Topics: Animals; Biotransformation; Carbon; Chromatography, High Pressure Liquid; Chromatography, Thin Layer; Fermentation; Flavanones; Flavonoids; Glucosides; Glucuronidase; Levilactobacillus brevis; Milk; Nitrogen; Plant Extracts; Scutellaria baicalensis; Temperature

The current study aims to investigate the pharmacokinetic properties of Huangqin Tang on different oral doses. An LC-MS method for simultaneous determination of flavonoids and terpenoids in rat plasma was developed and validated. Plasma samples were treated with hydrochloric acid (containing 1% ascorbic acid), precipitated with acetonitrile, separated on a Zorbax SB-C18 column, detected by single quadruple mass spectrometry with an electrospray ionization interface, and quantified using selected ion monitoring mode. All pharmacokinetic parameters were processed by non-compartmental analysis using WinNonlin software. The results of specificity, linearity, intra-day and inter-day precisions, accuracy, and stability for LC-MS assay were suitable for the quantification of paeoniflorin, baicalin, wogonoside, baicalein, wogonin, oroxylin A, glycyrrhizic acid and glycyrrhetinic acid in rat plasma. The concentration-time profiles of baicalin, wogonoside, baicalein, wogonin, oroxylin A and glycyrrhizic acid showed double-peak phenomenon after Huangqin Tang was orally administered at 40 g x kg(-1) dose; all eight constituents in rat plasma showed good dose-exposure relationship within the dosage of 10-40 g x kg(-1); although plasma concentrations were different, the flavonoids with the same backbone showed the similar fate in the body with the corresponding dosage. In conclusion, the LC-MS assay was successfully applied for the pharmacokinetic study of multi-constituents of Huangqin Tang with different doses. Additionally, these constituents demonstrated good pharmacokinetic properties in the body.

Topics: Administration, Oral; Animals; Chromatography, Liquid; Dose-Response Relationship, Drug; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Glycyrrhetinic Acid; Glycyrrhizic Acid; Male; Monoterpenes; Pentacyclic Triterpenes; Rats; Rats, Wistar; Spectrometry, Mass, Electrospray Ionization

Previous phytochemical studies showed that the major flavonoids in Scutellaria baicalensis are baicalin, baicalein, wogonoside and wogonin. The two glycosides (baicalin and wogonoside) can be transformed into their aglycons (baicalein and wogonin), which possess positive anticancer potential. In this study, we used glycosidase to catalyze flavonoids in S. baicalensis to enhance the herb's anticancer activities. Our HPLC data showed that, using the optimized conditions obtained in our experiments (20 U/g of cellulase, 50˚C, pH 4.8 and treatment for 8 h), there was a marked transformation from the two glycosides to their aglycons. The anticancer activity was subsequently evaluated using a series of S. baicalensis extracts in which variable lengths of glycosidase treatment time were used. Combining analytical and bioassay results, we observed that the higher the aglycon content, the stronger the antiproliferation effects. Compared to the untransformed control, 8 h of glycosidase catalyzing significantly increased antiproliferative activity on human colorectal and breast cancer cells, and its cancer cell growth inhibition is, in part, mediated by cell cycle arrest at the S-phase and induction of apoptosis. Data from this study suggest that using glycosidase to catalyze S. baicalensis offers a promising approach to increase its anticancer activity.

Topics: Apoptosis; Breast Neoplasms; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Colonic Neoplasms; Female; Flavanones; Flavonoids; Glucosides; Glycoside Hydrolases; Humans; Phytotherapy; Plant Extracts; Scutellaria baicalensis

This paper aims to establish a new method of calibration and positioning in quantitative analysis of multicomponents by single marker (QAMS), using Shuanghuanglian oral liquid as the research object. Establishing relative correction factors with reference chlorogenic acid to other 11 active components (neochlorogenic acid, cryptochlorogenic acid, cafferic acid, forsythoside A, scutellarin, isochlorogenic acid B, isochlorogenic acid A, isochlorogenic acid C, baicalin and phillyrin wogonoside) in Shuanghuanglian oral liquid by 3 correction methods (multipoint correction, slope correction and quantitative factor correction). At the same time chromatographic peak was positioned by linear regression method. Only one standard uas used to determine the content of 12 components in Shuanghuanglian oral liquid, in stead of needing too many reference substance in quality control. The results showed that within the linear ranges, no significant differences were found in the quantitative results of 12 active constituents in 3 batches of Shuanghuanglian oral liquid determined by 3 correction methods and external standard method (ESM) or standard curve method (SCM). And this method is simpler and quicker than literature methods. The results were accurate and reliable, and had good reproducibility. While the positioning chromatographic peaks by linear regression method was more accurate than relative retention time in literature. The slope and the quantitative factor correction controlling the quality of Chinese traditional medicine is feasible and accurate.

Topics: Apigenin; Calibration; Chlorogenic Acid; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Glucuronates; Glycosides; Reproducibility of Results

The aim is to study the intestinal absorption of different combinations of active compounds out of Gegenqinlian decoction. Rat single pass intestinal perfusion model with jugular vein cannulated was used. Samples were obtained continuously from the outlet perfusate and the mesenteric vein. The levels of puerarin, daidzin, liquilitin, baicalin, wogonoside, jatrorrhizine, berberine and palmatine were determined by LC-MS/MS and their permeability coefficients were calculated. The results showed that Glycyrrhiza could promote the absorption of the active ingredients in Pueraria which is the monarch herb; meanwhile, Pueraria also played a role in promoting the absorption of liquilitin. Based on the Gegenqinlian decoction and the different combinations experiments, the results concerning the absorption of baicalin and wogonoside were as follows. For baicalin, Pueraria and Glycyrrhiza could promote its absorption and the effect of Pueraria was more obvious. For wogonoside, Pueraria could also promote its absorption, while Glycyrrhiza played a opposite role. Pueraria and Glycyrrhiza both played a part in promoting the absorption of jateorhizine, berberine and palmatine, the effective compounds in Coptis.

Topics: Animals; Berberine; Berberine Alkaloids; Coptis; Drug Combinations; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Glycyrrhiza; Intestinal Absorption; Intestinal Mucosa; Isoflavones; Male; Plant Roots; Plants, Medicinal; Pueraria; Rats; Rats, Sprague-Dawley; Rhizome; Scutellaria baicalensis

Scutellariae Radix, the dried root of Scutellaria baicalensis Georgi, has been widely used in Asian countries for the treatment of dermatitis, diarrhoea, inflammatory disease and hepatic disease. A simple, sensitive and precise reversed-phase liquid chromatographic method with isocratic elution was developed to simultaneously determine four bioactive compounds in Scutellariae Radix: baicalein, baicalin, wogonin and wogonoside. Chromatographic analysis was performed on a YMC Pack Pro C(8) column (150 × 4.6 mm(2), 3 µm), with a mobile phase of 0.1% formic acid : acetonitrile (70 : 30, v/v) at a flow rate of 1.0 mL min(-1), and UV detection at 280 nm. Linear behaviour was observed over the investigated concentration range (0.25-10 µg mL(-1)) for all analytes, with a correlation coefficient of >0.997. The intra- and inter-day precisions were <8.07%, and accuracies were 92.3-102.9%. This method was successfully applied for the analysis of marker compounds for the quality control of Scutellariae Radix extract.

Topics: Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Limit of Detection; Quality Control; Reproducibility of Results; Scutellaria baicalensis

Reversed phase high performance liquid chromatography with diode array detector was employed for simultaneous determination of five main index components and specific chromatograms analysis in Xiaochaihu granules with a linear gradient elution of acetonitrile-water (containing 0.1% phosphoric acid) as mobile phase. The results showed that five main index components (baicalin, baicalein, wogonoside, wogonin, enoxolone) were separated well under the analytical condition. The linear ranges of five components were 0.518 - 16.576, 0.069 - 2.197, 0.167 - 5.333, 0.009 - 0.297 and 0.006 - 0.270 mg x g(-1), respectively. The correlation coefficients were 0.999 9, and the average recoveries ranged from 95% to 105%. Twelve common peaks were selected as the specific chromatograms of Xiaochaihu granules with baicalin as the reference peak. There were good similarities between the reference and the ten batches of samples. The similarity coefficients were no less than 0.9. The analytical method established is highly sensitive with strong specificity and it can be used efficiently in the quality control of Xiaochaihu granules.

Topics: Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Glycyrrhetinic Acid; Quality Control; Sensitivity and Specificity

This work is to study the baicalin and its three analogs, baicalin, wogonoside, and wogonin, on the protective effect of neuron from oxygen-glucose deprivation (OGD) and toll-like receptor 2 (TLR2) expression in OGD damage. The results showed that baicalin and its three analogs did protect neurons from OGD damage and downregulated protein level of TLR2. D-Glucopyranosiduronic acid on site 7 in the structure played a core of cytotoxicity of these flavonoid analogs. The methoxyl group on carbon 8 of the structure had the relation with TLR2 protein expression, as well as the anti-inflammation. In addition, we detected caspase3 and antioxidation capability, to investigate the effect of four analogs on cell apoptosis and total antioxidation competence in OGD model.

Topics: Animals; Antioxidants; Caspase 3; Cell Survival; Flavanones; Flavonoids; Glucose; Glucosides; Immunity, Innate; Neurons; Oxygen; PC12 Cells; Rats; Toll-Like Receptor 2; Tumor Necrosis Factor-alpha

Hairy root cultures of Scutellaria lateriflora were established using Agrobacterium rhizogenes A4 and produced acteoside (18.5 mg g(-1) dry wt), baicalin (14.5 mg g(-1) dry wt) and wogonoside (12 mg g(-1) dry wt). Yeast extract (50 μg ml(-1)) increased acteoside production 1.4-fold and flavone production 1.7-fold after 7 and 14 days of elicitation. Addition of Pectobacterium carotovorum lysate in the stationary phase of the hairy root culture stimulated only the accumulation of wogonin to 30 mg g(-1) dry wt. The production of wogonin in hairy roots could be associated with its role as a phytolaexin.

Topics: Agrobacterium; Culture Media; Flavanones; Flavonoids; Glucosides; Pectobacterium carotovorum; Phenols; Plant Roots; Scutellaria; Time Factors; Transformation, Genetic

It is well-known that baicalin-berberine complex (1) precipitates in the water decoction of numerous Chinese Medicinal formulae containing Radix Scutellariae and Rhizoma Coptidis or Cortex Phellodendri. In the current study, ionic interaction between the carboxylate ion of baicalin and the quaternary ammonium ion of berberine was revealed to be responsible for the formation of 1 and wogonoside-berberine (2) by using FAB-MS and NMR titration experiments. In addition, nuclear Overhauser effect spectroscopy (NOESY) correlations observed in 1 and 2 suggested quite different conformation of the two complexes, which was further supported by the fact that the [α](D) of the canadine obtained by reduction of 1 is of an opposite sign to that obtained from 2. Partition coefficients (n-octanol/water) determination demonstrated 12-20 times larger partition coefficient of each complex (1, 2) than that of each single compound (baicalin, wogonoside, and berberine), indicating the significant role of the formation of the complex in the bioavailability enhancement of these pharmacologically active constituents.

Topics: Berberine; Coordination Complexes; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Magnetic Resonance Spectroscopy; Molecular Conformation; Octanols; Water

A liquid chromatography tandem mass spectrometry (LC-MS/MS) method was developed for the simultaneous determination of baicalin, wogonoside, baicalein, wogonin, oroxylin A and chrysin in rat plasma, using naringin as an internal standard. After acidifying with HCl, plasma samples were pretreated by liquid-liquid extraction with acetone. Chromatographic separation was accomplished on a Hypersil Gold-C(18) analytical column (2.1×150 mm, 5 μm) utilizing a gradient elution profile and a mobile phase consisting of (A) 0.1% formic acid in water and (B) acetonitrile. Detection was performed by multiple reaction monitoring (MRM) mode using electrospray ionization in the positive ion mode. All analytes showed good linearity over the investigated concentration range (r>0.9900). The lower limit of quantification was 0.5 ng/ml for baicalin, wogonoside, wogonin and oroxylin A, and 1.0 ng/ml for baicalein and chrysin. Intra-day and inter-day precisions (RSD%) were less than 15% and accuracy (RE%) ranged from -6.7% to 5.8%. The validated method was successfully applied to investigate the pharmacokinetics of the major flavonoids of Radix scutellariae extract after oral administration to rats.

Topics: Administration, Oral; Animals; Area Under Curve; Chromatography, Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Half-Life; Limit of Detection; Male; Metabolic Clearance Rate; Plant Extracts; Rats; Rats, Wistar; Reference Standards; Reproducibility of Results; Scutellaria baicalensis; Sensitivity and Specificity; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry

In this study, resin adsorption as a means to separate and purify baicalin and wogonoside from extracts of Scutellaria baicalensis was investigated. Among the ten tested resins, the non-polar resin HPD-100 offered the best adsorption and desorption properties. Langmuir and Freundlich isotherms were used to describe the interactions between solutes and resin at different temperatures, and the equilibrium experimental data were well fitted to the two isotherms. Column packed with HPD-100 resin was used to perform dynamic adsorption and desorption tests to optimize the separation process. After one round treatment with HPD-100 resin, the contents of baicalin and wogonoside were 3.6-fold and 12.0-fold increased with recovery yields of 85.7% and 65.6%, respectively. In addition, a laboratory preparative-scale separation was carried out under the final conditions. The results showed that the preparative separation of baicalin and wogonoside can be easily and efficiently achieved via adsorption and desorption on HPD-100 resin. The developed method is a promising basis for large-scale preparation of baicalin and wogonoside from S. baicalensis extracts.

Topics: Adsorption; Chromatography, High Pressure Liquid; Ethanol; Flavanones; Flavonoids; Glucosides; Hydrogen-Ion Concentration; Kinetics; Plant Extracts; Plant Roots; Scutellaria baicalensis; Temperature

To study the transportation of Xiaochaihu Tang in Caco-2 cell model.. The safety concentration of Xiaochaihu Tang in Caco-2 cells was determined by MTT assay. Then the Caco-2 cell model was used to investigate the bi-directional transportation of Xiaochaihu Tang. The multicomponents of Xiaochaihu Tang and the influence of time were measured by high performance liquid chromatography (HPLC).. The P(app) values of wogonoside and wogonin were (1.23 +/- 0.09) x 10(-6), (1.07 +/- 0.89) x 10(-5) cm x s(-1) from the AP side to BL side, and (2.12 +/- 0.19) x 10(-6) and (7.12 +/- 1.02) x 10(-6) cm x s(-1) from the BL side to AP side, respectively. The P(appAP --> BL)/P(app BL --> AP) ratio of wogonoside and wogonin were 0.58 and 1.49, respectively. Baicalin, baicalein and glycyrrhizic acid could not permeate the Caco-2 cell model.. The transportation of wogonoside and wogonin in Caco-2 cell model may be a passive transportation.

Topics: Biological Transport, Active; Caco-2 Cells; Cell Line, Tumor; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Glycyrrhizic Acid; Humans

To establish a HPLC-PDA method for the determination of baicalin, wogonoside, baicalein, wogonin and glycyrrhizic acid in Xiaochaihu Tang.. A Symmetry Shield RP18 (4.6 mm x 250 mm, 5.0 microm) was used with a mobile phase of acetonitrile-0.01% H3PO4 in gradient elution. The detection wavelength was 251 nm,the flow rate was 0.45 mL x min(-1) and the column temperature was maintained at 30 degrees C.. The accuracy, precision, sensitivity, specificity and linearity of this method met the requirements. The contents of the five effective fractions were determined simultaneously.. The method is rapid,simple and accurate and it can be suitable for the determination of baicalin, wogonoside, baicalein, wogonin and glycyrrhizic acid in Xiaochaihu Tang simultaneously.

Topics: Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Glycyrrhizic Acid; Indicators and Reagents; Medicine, Chinese Traditional; Rehmannia; Spectrophotometry, Ultraviolet; Technology, Pharmaceutical

The aim of this study was to compare the pharmacokinetics of baicalin and wogonoside in rats following oral administration of Xiaochaihu Tang (Minor Radix Bupleuri Decoction) and Radix scutellariae extract. Thus, a specific LC-MS method was developed and validated for the determination of these flavonoids in the plasma of rats after oral administration Xiaochaihu Tang and Radix scutellariae extract. Chromatographic separation was performed on a Zorbax SB C(18) column (150mmx4.6mm, i.d.: 5microm) with 0.1% formic acid in water and acetonitrile by linear gradient elution. Baicalin, wogonoside and carbamazepine (internal standard, I.S.) were detected in select-ion-monitoring (SIM) mode with a positive electrospray ionization (ESI) interface. The following ions: m/z 447 for baicalin, m/z 461 for wogonoside and m/z 237 for the I.S. were used for quantitative determination. The calibration curves were linear over the concentration ranges from 0.1231 to 6.156microg mL(-1) for baicalin and 0.08832 to 4.416microg mL(-1) for wogonoside. The lower limit of detection (LLOD) based on a signal-to-noise ratio of 2 was 0.06155microg mL(-1) for baicalin and 0.04416microg mL(-1) for wogonoside. Intra-day and inter-day precisions (RSD%) were within 10% and accuracy (RE%) ranged from -6.4 to 4.4%. The extraction recovery at three QC concentrations ranged from 74.7 to 86.0% for baicalin and from 71.3 to 83.7% for wogonoside. The plasma concentrations of baicalin and wogonoside in rats at designated time periods after oral administration were successfully determined using the validated method, pharmacokinetic parameters were estimated by a non-compartment model. Following oral administration of Xiaochaihu Tang and Radix scutellariae extract, the t(1/2) of baicalin was 3.60+/-0.90 and 5.64+/-1.67, the C(max1) was 1.64+/-0.99 and 5.66+/-2.02, the t(max1) was 0.13+/-0.05 and 0.20+/-0.07, the C(max2) was 2.43+/-0.46 and 3.18+/-1.66, and the t(max2) were 6.40+/-1.67 and 5.66+/-2.02, respectively. Following oral administration of Xiaochaihu Tang and Radix scutellariae extract, the t(1/2) of wogonoside was 4.97+/-1.68 and 7.71+/-1.55, the C(max1) was 1.39+/-0.83 and 1.45+/-0.37, the t(max1) was 0.21+/-0.20 and 0.17+/-0.01, the C(max2) was 1.90+/-0.55 and 1.42+/-0.70, and the t(max2) was 5.60+/-1.67 and 5.20+/-1.79, respectively. A significant difference (p<0.05) was observed for t(1/2), and the elimination of baicalin and wogonoside in Xiaochaihu Tang was increased.

Topics: Animals; Chromatography, Liquid; Drug Stability; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Linear Models; Mass Spectrometry; Rats; Reproducibility of Results; Scutellaria baicalensis; Sensitivity and Specificity

A rapid, sensitive, and specific method by high-performance liquid chromatography (HPLC) coupled to diode-array detection (DAD) and tandem mass spectrometry (MS) techniques was developed for the identification of absorbed constituents and their metabolites in rats after the oral administration of a Chai-Huang decoction (CHD), which consists of Bupleurum chinense and Scutellaria baicalensis in the proportion 1 : 1 (w/w). By comparing their retention times and MS data with those of authentic compounds and published data, a total of 14 compounds were identified in the CHD samples. In addition, eleven and seven compounds were characterized in the urine and serum samples of the rats, respectively. The results indicated that the main absorbed constituents were chrysin-6-C-arabinosyl-8-C-glucoside, chrysin-6-C-glucosyl-8-C-arabinoside, baicalin, wogonin-5-O-glucoside, oroxylin A-7-O-glucuronide, wogonoside, saikosaponin A, saikosaponin C, saikosaponin D, baicalein, and wogonin. These compounds might be responsible for the curative effects of the CHD. The findings demonstrated that the proposed method could be used to rapidly and simultaneously analyze and screen the multiple absorbed bioactive constituents in a formula of traditional Chinese medicines (TCM). This is very important not only for the pharmaceutical discovery process and the quality control of crude drugs but also to explain the mechanisms of action of TCM.

Topics: Administration, Oral; Animals; Bupleurum; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Glucuronides; Medicine, Chinese Traditional; Oleanolic Acid; Rats; Saponins; Scutellaria baicalensis; Spectrometry, Mass, Electrospray Ionization

A specific ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) method was developed for the simultaneous determination of puerarin, daidzein, baicalin, wogonoside and liquiritin in rat plasma. Chromatographic separation was performed on a C(18) column packed with 1.7 microm particles by a linear gradient elution. The analytes and carbamazepine (internal standard, I.S.) were monitored in a selected-ion reaction (SIR) mode with a positive electrospray ionization (ESI) interface by the following ions: m/z 417.2 for puerarin, m/z 255.2 for daidzein, m/z 271.0 for baicalin, m/z 461.0 for wogonoside, m/z 441.0 for liquiritin and m/z 237.2 for carbamazepine (I.S.), respectively. The calibration curves of these analytes were linear over the concentration ranges from 0.00254-1.02 microg mL(-1) to 0.0102-10.2 microg mL(-1). Within-batch and between-batch precisions (RSD%) were all within 15% and accuracy (RE%) ranged from -10% to 10%. The extraction recoveries were on average 79.8% for puerarin, 90.8% for daidzein, 74.4% for baicalin, 70.2% for wogonoside and 84.7% for liquiritin. The validated method was successfully applied to investigate the pharmacokinetics of five bioactive compounds of GegenQinlian decoction (GQD) in rats.

Topics: Animals; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Isoflavones; Mass Spectrometry; Rats; Sensitivity and Specificity

Radix Scutellariae (RS) and Coptis Chinensis (CC) are the most popular components in traditional Chinese medicine prescriptions. Flavonoids are the main effective ingredients in RS and berberine is the main effective ingredient in CC. The aim of this study was to determine the influence of CC on the pharmacokinetics of flavonoids following the administration of RS in rats and to investigate the effects of CC on the pharmacokinetic mechanism. Rats were administered RS or RS+CC by intragastric gavage (i.g.). Plasma concentrations of baicalin (baicalein 7-glucuronide) and wogonoside (wogonin 7-glucuronide) were measured by HPLC. Pharmacokinetic parameters were calculated from the plasma concentration time curve. The effect of CC on the metabolism of flavonoids in RS by rat intestinal flora was studied using rat fecal suspensions after ig administration of CC. The effect of berberine on the bi-directional transport of baicalein and baicalin was studied in Caco-2 monolayer models. The effect of CC on the metabolism of baicalein and baicalin by the liver was studied using rat liver microsomes after i.g. administration of CC. Compared to RS alone, RS+CC decreased AUC and C(max) of baicalin and wogonoside. CC decreased the hydrolyzation of baicalin and wogonoside by intestinal flora. Berberine decreased the amount of baicalein transported to the basolateral side and the amount of baicalin which appeared at the basolateral side. The results showed that CC may decrease the bioavailability of baicalin and wogonoside in RS and the mechanism was related to CC decreasing the transport of flavonoid aglycones from the mucosa side to the serosal side and the hydrolyzation of flavonoids by inhibiting intestinal flora.

Topics: Administration, Oral; Animals; Coptis; Drugs, Chinese Herbal; Female; Flavanones; Flavonoids; Glucosides; Glucuronides; Intestinal Mucosa; Male; Plant Extracts; Rats; Rats, Sprague-Dawley; Scutellaria baicalensis

The beta-glucuronidase (GUS) gene from Lactobacillus brevis RO1 was cloned and expressed in Escherichia coli GMS407. The GUS gene was composed of 1812 bp, encoding a 603-amino-acid protein belonging to the glycosyl hydrolase family 2 with three conserved domains. The amino acid similarity was higher than 70% with the beta-glucuronidases of various microorganisms, yet less than 58% with the beta-glucuronidase of L. gasseri ADH. Overexpression and purification of the GUS was performed in beta-glucuronidase-deficient E. coli GMS407. The purified GUS protein was 71 kDa and showed 1284 U/mg of specific activity at optimum condition of pH 5.0 and 37 degrees C. At 37 degrees C, the GUS remained stable for 80 min at pH values ranging from 5.0 to 8.0. The purified enzyme exhibited a half-life of 1 h at 60 degrees C and more than 2 h at 50 degrees C. When the purified GUS was applied to transform baicalin and wogonoside into their corresponding aglycones, 150 microM of baicalin and 125 microM of wogonoside were completely transformed into baicalein and wogonin, respectively, within 3 h.

Topics: Amino Acid Sequence; Cloning, Molecular; Enzyme Stability; Escherichia coli; Flavanones; Flavonoids; Glucosides; Glucuronidase; Hydrogen-Ion Concentration; Levilactobacillus brevis; Molecular Sequence Data; Molecular Weight; Sequence Homology, Amino Acid; Substrate Specificity; Temperature; Time Factors

To develop a quantitative method for simultaneously determining four flavones in Radix Scutellariae, by using one chemical standard substance.. The relative correction factors (RCF) of four flavones were determined by HPLC-DAD. Within the linear ranges, the values of RCF at 274 nm of wogonoside, baicalein, and wogonin to baicalin, were 0.7467, 0.5638, 0.4351, respectively. And the contents of baicalin in 27 samples of Scutellaria baicalensis were authentically determined by the external standard method, and the contents of the three other flavones were calculated according to their RCF. The contents of these four flavones in the all samples were determined with the external standard method.. The RCF had a good reproducibility (RSD 0.66%-2.83%). No significant differences between the quantitative results by the above two methods were observed.. It is a fast, convenient, and accurate method to determine multi-components especially when some authentic standard substances were unavailable. It can be used as a method to control the quality of Radix Scutellariae.

Topics: Chromatography, High Pressure Liquid; Flavanones; Flavones; Flavonoids; Glucosides; Plant Extracts; Scutellaria baicalensis

Scutellaria baicalensis was collected from four wild and four cultivated populations from different locations in China. Forty-two samples were analyzed using random amplification of polymorphic DNA (RAPD) techniques for genetic profiling, and high performance liquid chromatography (HPLC) techniques to determine the flavonoid content. The selected 23 RAPD primers yielded a total of 838 clear and reproducible bands of which 237 were found to be polymorphic. The wild population exhibited higher polymorphism than that of the cultivated population. The dendrogram generated by the UPGMA method via Nei's genetic distance revealed three distinct genotypes from the cultivated populations and several branches from the wild populations. The contents of baicalin and wogonoside in dried roots of the samples ranged from (w/w) 8.63 to 17.84%, and from 1.99 to 4.21%, respectively, whereas their aglycones, baicalein and wogonin, were within the range of only 0.04-0.23%. The total content of the four flavonoids varied from 9.45 to 26.24%. Comparatively, the cultivated populations contained much higher levels of baicalin and total flavonoids than those in the wild populations. The results from genetic characterization and phytochemical analysis demonstrated that the quality variation of this drug was mainly determined by extrinsic environmental or agricultural factors, rather than by genetic differences. Our findings can be used for the commercial production and germplasm management of this medicinal plant.

Topics: Chromatography, High Pressure Liquid; Flavanones; Flavonoids; Genetic Variation; Glucosides; Random Amplified Polymorphic DNA Technique; Scutellaria baicalensis

Three flavonoids of pharmaceutical importance-baicalein, baicalin, and wogonoside-were isolated from a Chinese medicinal plant Scutellaria baicalensis Georgi and studied by 13C NMR in solution and solid state. Two-dimensional (2D) NMR spectroscopy in the liquid phase and dipolar dephasing (DD) experiments in magic-angle spinning (MAS) spectra enabled the assignment of 13C resonances. The cross-polarization (CP) time constants T(CH) and relaxation times T(H) (1rho) were obtained from the variable-contact time experiments. The principal elements of the 13C chemical shift tensor were determined in the spectra recorded under slow sample spinning (2 kHz) using phase-adjusted spinning sideband (PASS)-2D NMR technique, and were verified by density functional theory gauge-independent atomic orbital (DFT GIAO) calculations of shielding constants. Analysis of the 13C delta(ii) and comparison with shielding parameters calculated for different conformers of compounds 1-3 enabled the selection of the most reliable geometry in the solid phase. In all three compounds, an intramolecular hydrogen bond C5--OH...=C4 is formed; the existence of baicalein and baicalin with 'anticlockwise' orientation of OH groups is more probable.

Topics: Flavanones; Flavonoids; Glucosides; Magnetic Resonance Spectroscopy; Models, Molecular; Molecular Structure; Plant Roots; Quantum Theory; Scutellaria baicalensis

Huang-Lian-Jie-Du-Tang (HLJDT) is an important "heat-clearing" multiherb remedy of traditional Chinese medicine, and Radix scutellariae (Scutellaria baicalensis Georgi, Labiatae) is a key ingredient herb in it. Baicalin and wogonoside are two main effective ingredients enriched in Radix scutellariae. In the present study, pharmacokinetic differences of baicalin following oral administration of pure baicalin, Radix scutellariae extract, baicalin co-administrated with extract of the other three herbs of HLJDT and HLJDT were investigated in male S.D. rats with approximately the same dose of 200 mg/kg baicalin. The pharmacokinetic comparison of wogonoside was conducted only in Radix scutellariae extract and HLJDT. Plasma concentrations of baicalin and wogonoside were determined using HPLC method. Unpaired Student's t-test was used for statistical comparison. The results indicated that baicalin and wogonoside demonstrated bimodal phenomenon in the plasma profile. Some ingredients in the other three herbs of HLJDT, not in Radix scutellariae itself, had pharmacokinetic interaction with baicalin and wogonoside and hence decreased their systematic exposure level (p<0.01). The absorption site of baicalin was preliminary evaluated in rat using in situ absorption in stomach and different intestinal segments and results revealed the existence of double-site absorption of baicalin. The first absorption site was in upper intestinal, probably via directly absorption of baicalin; while the second absorption site was in colon in the form of aglygon.

Topics: Administration, Oral; Animals; Area Under Curve; Biological Transport; Chromatography, High Pressure Liquid; Drug Interactions; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Intestinal Absorption; Intestinal Mucosa; Male; Plant Extracts; Rats; Rats, Sprague-Dawley; Scutellaria baicalensis

To study the pharmacokinetics of flavonoids from Xiexin decoction in rats. SD rats were given a single ig dose of Xiexin decoction 12 g x kg(-1), plasma and urine were collected before and after dosing. Flavonoids components in plasma and urine were measured by HPLC. Pharmacokinetic parameters were determined from the plasma concentration-time data and urinary excretion-time data with the DAS software package. Baicalin was incubated with the rat renal homogenate to investigate its metabolism in vitro. After oral administration of Xiexin decoction baicalin and wogonoside were quickly absorbed and exhibited double peak phenomena in their plasma concentrations. The first peaks in plasma concentrations of baicalin and wogonoside reached Cmax1 of (10 +/- 8) and (1.5 +/- 0.5) mg x L(-1) at Tmax1 of (0.27 +/- 0.09) and (0.17 +/- 0.00) h, while the second peaks reached Cmax2 of (3. 9 0. 5) and (0. 74 +/- 0.11) mg x L(-1) at Tmax2 of (7.6 +/- 2.6) and (16.0 +/- 0.0) h, respectively. The T(1/2) of baicalin and wogonoside were (7 +/- 3) and (6.4 +/- 2.1) h, AUC(0-infinity) were (57 +/- 12) and (15 +/- 3) mg x h x L(-1), respectively. After oral administration of Xiexin decoction, not only baicalin and wogonoside but also baicalein and wogonin can be detected in the urine. The amounts of baicalin, wogonoside, baicalein and wogonin excreted from urine during 0-72 h were (1.4 +/- 0.3), (3.4 +/- 1.3), (2.2 +/- 0.97), (10 +/- 4)% of dose given in rats, respectively. The excretion T(1/2) of the four flavonoids were (6.9 +/- 2.1), (9 +/- 4) , (8.2 +/- 2.0) and (7.2 +/- 1.8) h, respectively. Baicalin was metabolized into baicalein in the rat renal homogenate in vitro, and the kinetic parameters were measured as Vmax = 702 nmol x min(-1) x g(-1) (protein) and Km=135 micromol x L(-1). After oral administration of Xiexin decoction, flavonoids can be absorbed quickly. Only a small quantity of baicalin, wogonoside, baicalein and wogonin were excreted from urine. Baicalin may be metabolized into baicalein in the rat kidney.

Topics: Administration, Oral; Animals; Area Under Curve; Chromatography, High Pressure Liquid; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Kidney; Male; Rats; Rats, Sprague-Dawley

To compare and analyze the effects of the contents of the active components in Radix Scutellariae by different processing and extracting methods.. The raw and processed Radix Scutellariae were used, and the concentration of baicalin, wogonoside, baicalein, wogonin were determined by HPLC after five extracting techniques. HPLC method was performed on methanol-acetonitrile-water-acetic acid (18: 25: 57: 0.2) as the mobile phase at 30 C; The chromatographic column was Agilent Zorbax Extend-C18 (4.6 mm x 250 mm, 5 microm); The flow rate was 0.8 mL min(-1), and the detection wavelength was 275 nm.. the ratios of baicalein and wogonin in raw material were 5.41%, 1.30% respectively by enzymatic extraction, which is higher than other extracting methods by raw material; The ratios of baicalin and wogonoside in raw material were 10.11% and 3.55% by ethanol of 60%, which is higher than other extracting methods; Different extracting methods have no evident effects on processed materials, enzymatic extraction is the best. The ratio of baicalin, wogonoside and baicalein, wogonin is 10.63%, 3.60% and 1.54%, 0.59%.. Different methods have evident extracting effects on the four active components with raw material, but have no evident effects with processed material. According to different active components most suitable extracting method should be adopted.

Topics: Chromatography, High Pressure Liquid; Flavanones; Flavonoids; Glucosides; Hot Temperature; Plant Roots; Plants, Medicinal; Reproducibility of Results; Scutellaria baicalensis; Technology, Pharmaceutical

A preparative high-speed counter-current chromatography (HSCCC) method for isolation and purification of baicalin and wogonoside from the Chinese medicinal plant Scutellaria baicalensis Georgi (Huang-qin in Chinese) was successfully established by using ethyl acetate-methanol-1% acetic acid water (5:0.5:5, v/v) as the two-phase solvent system. The upper phase of ethyl acetate-methanol-1% acetic acid water (5:0.5:5, v/v) was used as the stationary phase of HSCCC. Baicalin (58.1 mg) and wogonoside (17.0mg) with the purity of 99.2 and 99.0%, respectively, were separated successfully in one-step separation from 120 mg of crude sample from S. baicalensi, Georgi. The structures of baicalin and wogonoside were identified by 1H NMR and 13C NMR.

Topics: Chromatography, High Pressure Liquid; Countercurrent Distribution; Flavanones; Flavonoids; Glucosides; Magnetic Resonance Spectroscopy; Plant Roots; Scutellaria baicalensis; Solvents

To study the process in vivo of the components in the precipitation of Xiexin decoction.. The components in the rats' urine samples after oral administration of baicalin, single herb decoction of Radix Scutellariae or the precipitation of Xiexin decoction were analyzed by LC-MS-MS method; and the absorption of baicalin in different samples were calculated by intestinal in situ loop method.. The urinary excretion amounts of baicalin in three samples were not obviously different, whereas the time reach elimination-peak of baicalin in them had significant difference. The absorption of baicalin in the precipitation was obviously greater than that in the single herb decoction and single baicalin. We found that wogonoside and 7-O-glucuronide chrysin were the metabolites presenting in the rat urine samples after oral administration of baicalin by LC-MS-MS, which had never been reported.. The resident time in the body of baicalin in the precipitation is prolonged, compared with that in the single herb decoction and single baicalin. The metabolites may be the potential biological components in vivo of baicalin in the precipitation.

Topics: Administration, Oral; Animals; Chemical Precipitation; Coptis; Drug Combinations; Drugs, Chinese Herbal; Flavanones; Flavonoids; Glucosides; Male; Plants, Medicinal; Rats; Rats, Wistar; Rheum; Scutellaria baicalensis

The four flavonoids baicalin, baicalein, wogonoside, and wogonin have been isolated from Scutellaria baicalensis (Lamiaceae) root extract as main bioactive components. We evaluated their antimutagenic activity as potential chemopreventive natural substances using the Ames test. Free radical scavenging properties have also been measured with the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. Both crude extract and isolated flavone glycosides and aglycones have antimutagenic and free radical scavenging properties. The lipophilic flavone baicalein has been confirmed to be the most powerful antimutagen among the compounds tested.

Topics: Antimutagenic Agents; Antioxidants; Biphenyl Compounds; Flavanones; Flavonoids; Free Radical Scavengers; Glucosides; Hydrazines; Mutagenicity Tests; Picrates; Plant Extracts; Plant Roots; Scutellaria baicalensis