harman and norharman

Harman and norharman, two neuroactive β-carbolines, are present in several plants and in thermally processed foods. They exhibited a wide spectrum of biological and pharmacological effects, including antioxidant, neuroprotective, and anti-inflammatory effects. In this article, we review the progress of recent research on the presence of these compounds in food, as well as their various biological and neuroactive properties. Our findings strongly suggest that some foods, especially coffee, can act as a rich source of β-carbolines, which may possibly be associated with a reduced risk for serious neurodegenerative diseases, such as Parkinson's and Alzheimer's.

Topics: Animals; Brain; Brain Chemistry; Carbolines; Essential Tremor; Food; Food Handling; Harmine; Humans; Neurodegenerative Diseases; Neuroprotective Agents; Oxidative Stress; Parkinson Disease; Plant Extracts

Inhibition of monoamine oxidase is one way to treat depression and anxiety. The information now available on the pharmacokinetics of flavonoids and of the components of tobacco prompted an exploration of whether a healthy diet (with or without smoking) provides active compounds in amounts sufficient to partially inhibit monoamine oxidase. A literature search was used to identify dietary monoamine oxidase inhibitors, the levels of these compounds in foods, the pharmacokinetics of the absorption and distribution, and tissue levels observed. An estimated daily intake and the expected tissue concentrations were compared with the measured efficacies of the compounds as inhibitors of monoamine oxidases. Norharman, harman and quercetin dietary presence, pharmacokinetics, and tissue levels were consistent with significant levels reaching neuronal monoamine oxidase from the diet or smoking; 1,2,3,4-tetrahydroisoquinoline, eugenol, 1-piperoylpiperidine, and coumarin were not. Quercetin was equipotent with norharman as a monoamine oxidase A inhibitor and its metabolite, isorhamnetin, also inhibits. Total quercetin was the highest of the compounds in the sample diet. Although bioavailability was variable depending on the source, a healthy diet contains amounts of quercetin that might give sufficient amounts in brain to induce, by monoamine oxidase A inhibition, a small decrease in neurotransmitter breakdown.

Topics: Animals; Anxiety Disorders; Carbolines; Depressive Disorder; Harmine; Humans; Monoamine Oxidase; Monoamine Oxidase Inhibitors; Quercetin; Serotonin

The aromatic beta-carbolines norharman and harman have been implicated in a number of human diseases including Parkinson's disease, tremor, addiction and cancer. It has been shown that these compounds are normal body constituents formed endogenously but external sources have been identified. Here, we summarise literature data on levels of norharman and harman in fried meat and fish, meat extracts, alcoholic drinks, and coffee brews. Other sources include edible and medicinal plants but tobacco smoke has been identified as a major source. Exposure levels from these different dietary sources are estimated to a maximum of 4 microg norharman per kg body weight (bw) per day and 1 microg harman per kg bw per day. Exposure via tobacco smoke depends on smoking habits and type of cigarettes but can be estimated to 1.1 microg/kg bw for norharman and 0.6 microg/kg bw for harman per package of cigarettes smoked. Studies on toxicokinetics indicate that inhalative exposure leads to a rapid increase in plasma levels and high bioavailability of norharman and harman. Oral bioavailability is lower but there are indications that sublingual absorption may increase dietary uptake of beta-carbolines. Endogenous formation can be estimated to be 50-100 ng/kg bw per day for norharman and about 20 ng/kg bw per day for harman but these rates may increase with high intake of precursors. Biomarker studies on plasma levels of beta-carbolines reported on elevated levels of norharman, harman or both in diseased patients, alcoholics and following tobacco smoking or consumption of beta-carboline-containing food. Cigarette smoking has been identified as major influence but dietary exposure may contribute to exposure.

Topics: Biological Availability; Biomarkers; Carbolines; Harmine; Humans

Specific laboratory tests can be used to identify patients who are alcohol-dependent. The laboratory values of a number of biological 'markers', including carbohydrate-deficient transferrin, are often elevated in cases of chronic and acute alcohol abuse. Trait markers reflect a predisposition for alcoholism; state markers reflect actual alcohol consumption. It has been suggested that state markers can be subdivided into screening and relapse markers, and even further subdivided into pre-relapse markers, i.e. craving markers. We hypothesize that methanol metabolism and the presence of condensation products in the blood may serve as state and pre-relapse markers for alcoholism. Since the sensitivities and specificities of laboratory screening tests vary, and an absolute marker for alcoholism has yet to be identified, research in the area of biological markers for alcoholism should continue.

Topics: Alanine Transaminase; Alcoholism; Aspartate Aminotransferases; Biomarkers; Carbolines; Erythrocyte Indices; Harmine; Humans; Sensitivity and Specificity; Transferrin

The mutagenic and co-mutagenic properties of harman, norharman and of some of their pharmacologically important derivatives are reviewed. These compounds do not behave as true mutagens, but rather interact, directly or indirectly with DNA, leading to various consequences. This unusual behaviour is most probably related to the particular structure of the chemical nucleus common to all beta-carbolines which confers to the different derivatives the property to interact with various macromolecules and enzymatic systems. These interactions are compiled and discussed in this review. The alterations, by beta-carbolines, of some important enzymatic systems, e.g. cytochrome P-450, have been clearly demonstrated, yet many discrepancies and contradictions exist so that an interpretation of the results and the definition of some common mechanism appears premature. Since beta-carbolines are widely distributed in tissues and since they may modify and increase genotoxic and toxic consequences of other compounds, these interactions need to be clarified.

Topics: Animals; Carbolines; DNA; Drug Interactions; Harmine; Humans; Mutagenicity Tests; Mutagens

Topics: Animals; Breast Neoplasms; Carbolines; Carcinogens, Environmental; Cattle; Colonic Neoplasms; Cooking; Food; Harmine; Head and Neck Neoplasms; Humans; Imidazoles; Male; Meat; Mutagens; Neoplasms; Nitrates; Nitrites; Nitrosamines; Prostatic Neoplasms

Epidemiological studies suggest that smoking reduces the risk for Parkinson's disease. It has been hypothesized that inhibition of monoamineoxidase contributes to this action. The present study examined the contribution of the beta-carbolines norharman, an inhibitor of monoamineoxidase B, and harman, an inhibitor of monoamineoxidase A, which are present in high concentrations in tobacco smoke to the protective action. Nineteen active smokers and five nonsmokers smoked one and two cigarettes. The levels of norharman and harman increased in plasma from smokers and nonsmokers. Ex vivo saturation kinetic experiments revealed that the baseline affinity constant of monoamineoxidase in platelets from smokers was higher than that of nonsmokers in contrast to the maximum turnover rate, which did not differ. Acute smoking affected the monoamineoxidase in nonsmokers only. It is discussed that inhibition of both isoforms of monoamineoxidase is necessary for the neuroprotection and that both norharman and harman play an important role.

Topics: Adult; Blood Platelets; Carbolines; Harmine; Humans; Kinetics; Male; Monoamine Oxidase; Monoamine Oxidase Inhibitors; Smoking; Time Factors

Topics: Amines; Animals; Antioxidants; Carbolines; Carcinogens; Chromatography, High Pressure Liquid; Garlic; Harmine; Hot Temperature; Humans; Meat; Meat Products; Mutagens; Onions; Swine

Unusual cascade transformation was developed involving microwave assisted electrocyclic cyclization of aci (alkylideneazinic acid) forms of nitrovinylindoles acting as heterotrienes. Subsequent one-pot reduction allowed for efficient access to β-carbolines, including several natural products, alkaloids norharmane, harmane and eudistomin N.

Topics: Alkaloids; Biological Products; Carbolines; Cyclization; Harmine

The β-carbolines, mainly including harman and norharman, are a group of naturally occurring, plant-derived alkaloids, and are also considered as nonpolar heterocyclic aromatic amines. Sesame seed oils contain a high level of β-carbolines (harman and norharman). In China, sesame seed oil blends are one of the most popular types of vegetable oils blends, which can be used as cooking oils or frying oils. Thus, it is meaningful to investigate the degradation of β-carbolines (harman and norharman) in sesame seed oil blends as frying oils during heating. In this work, the loss of harman and norharman in different types of sesame seed oil blends have been investigated. The results showed that the degradation of harman and norharman were dependent both on the type of oil blends, heating temperature and time. Harman and norharman were more degraded during heating (150 °C, 180 °C) in oleic acid-rich oil blends compared to polyunsaturated acid-rich oil blends. Mechanistic investigation suggested that the reduction in harman and norharman in oil blends during heating was mainly due to the oxidative degradation reaction between β-carbolines and lipid oxidation products. Therefore, the contents of β-carbolines (harman and norharman) in sesame seed oil blends when used as frying oils and heated can be decreased with prolonged cooking time.

Topics: Alkaloids; Carbolines; Harmine; Heating; Oxidation-Reduction; Plant Oils

Zebrafish provide a valuable emerging complementary model for neurobehavioral research. They offer a powerful way to screen for the potential therapeutic effects of neuroactive drugs. A variety of behavioral tests for zebrafish have been developed and validated for assessing neurobehavioral function. The novel tank diving test is a straightforward, reproducible way of measuring anxiety-like behavior in zebrafish. When introduced into a novel tank, zebrafish normally dive to the bottom of the tank and then gradually explore the higher levels of the water column as time progresses. Buspirone is an effective anxiolytic drug in humans, which has been found, with acute administration, to reduce this anxiety-like response in zebrafish. The current study used the zebrafish model to evaluate the potential anxiolytic effects of alkaloids, commonly found in Solanaceae plants, with known neuropharmacology relevant to mood regulation. In line with previous findings, acute treatment with anxiolytic positive controls buspirone and the plant alkaloid nicotine reduced the anxiety-like diving response in the zebrafish novel tank diving test. Further, both buspirone and nicotine continued to produce anxiolytic-like effects in zebrafish after 5 days of exposure. In the same treatment paradigm, the effects of five other alkaloids-cotinine, anatabine, anabasine, harmane, and norharmane-were investigated. Cotinine, the major metabolite of nicotine, also caused anxiolytic-like effects, albeit at a dose higher than the effective dose of nicotine. Nicotine's anxiolytic-like effect was not shared by the other nicotinic alkaloids, anabasine and anatabine, or by the naturally present monoamine oxidase inhibitors harmane and norharmane. We conclude that nicotine uniquely induces anxiolytic-like effects after acute and subchronic treatment in zebrafish. The zebrafish model with the novel tank diving test could be a useful complement to rodent models for screening candidate compounds for anxiolytic effects in nonclinical studies.

Topics: Alkaloids; Anabasine; Animals; Anti-Anxiety Agents; Anxiety; Behavior, Animal; Buspirone; Carbolines; Cotinine; Disease Models, Animal; Female; Harmine; Humans; Male; Nicotine; Pyridines; Solanaceae; Zebrafish

The beta-carbolines norharman and harman, two heterocyclic aromatic amines with potential mutagenicity, have been determined in vegetable oils. Identification and analysis were carried out by ultra-performance liquid chromatography-triple quadrupole tandem mass spectrometry (UPLC-MS/MS). In 88 samples analysed, the concentrations of norharman and harman were < LOD to 336.22 ng/g and < LOD to 505.14 ng/g, respectively. A high variability of norharman and harman levels among different oil types was observed. Sesame-, flaxseed-, sunflower seed-, peanut- and rapeseed oils were most contaminated. Both

Topics: Arachis; Brassica napus; Carbolines; China; Chromatography, High Pressure Liquid; Cooking; Diet; Flax; Food Contamination; Harmine; Helianthus; Hot Temperature; Humans; Mutagens; Plant Oils; Seeds; Tandem Mass Spectrometry

Topics: Acrylamide; Amines; Asparagine; Carbolines; Chromatography, Gas; Chromatography, High Pressure Liquid; Glucose; Glycation End Products, Advanced; Harmine; Hot Temperature; Lysine; Maillard Reaction; Models, Biological; Pyruvaldehyde

Some studies have ascribed a protective effect against neurodegenerative diseases to the β-carbolines harman (H) and norharman (NH), which occur mostly in coffee and coffee substitutes. We determined the concentrations of β-carbolines and undesirable compounds (such as acrylamide) in roasted coffee substitute ingredients and found that chicory coffee was optimal. Two in vivo experiments were conducted with seventeen-month-old male Sprague Dawley rats fed a diet with the addition of pure carboline standards in the first stage, and chicory in the second. We observed an increase in the level of H and NH in blood plasma, as well as higher activity of animals in the battery behavioral test, particularly in the second stage. The results of in vitro studies-particularly the level of the expression in brain tissue of genes associated with aging processes and neurodegenerative diseases-clearly show the benefits of a diet rich in β-carbolines.

Topics: Animals; Brain; Carbolines; Cichorium intybus; Coffee; Gene Expression Regulation; Harmine; Male; Neurodegenerative Diseases; Rats; Rats, Sprague-Dawley

Identifying novel constituents that contribute to tobacco addiction is essential for developing more effective treatments and informing FDA regulation of tobacco products. While preclinical data indicate that monoamine oxidase (MAO) inhibitors can have abuse liability or potentiate the addiction-related effects of nicotine, most of these studies have used clinical MAO inhibitors (e.g., tranylcypromine) that are not present in cigarette smoke. The primary goal of this study was to evaluate the abuse potential of the β-carbolines harmane, norharmane, and harmine - MAO inhibitors that are found in cigarette smoke - in an intracranial self-simulation (ICSS) model in rats. A secondary goal was to evaluate the ability of norharmane to influence nicotine's acute effects on ICSS. None of the β-carbolines lowered ICSS thresholds at any dose studied when administered alone, suggesting a lack of abuse liability. Rather, all three β-carbolines produced dose-dependent elevations in ICSS thresholds, indicating aversive/anhedonic effects. Harmane and harmine also elevated ICSS response latencies, suggesting a disruption of motor function, albeit with reduced potency compared to their ICSS threshold-elevating effects. Norharmane (2.5 mg/kg) modestly attenuated the effects of nicotine on ICSS thresholds. Our findings indicate that these β-carbolines produced only aversive/anhedonic effects in an ICSS model when administered alone, and that norharmane unexpectedly attenuated nicotines acute effects on ICSS. Future work evaluating the addiction-related effects of nicotine combined with these and other MAO inhibitors present in smoke may be useful for understanding the role of MAO inhibition in tobacco addiction and informing FDA tobacco regulation.

Topics: Animals; Behavior, Addictive; Brain; Carbolines; Female; Harmine; Male; Monoamine Oxidase Inhibitors; Motor Activity; Nicotiana; Nicotine; Rats; Rats, Sprague-Dawley; Reinforcement, Psychology; Self Stimulation; Smoke

Heterocyclic amines (HCAs) are foodborne carcinogens for which their formation is highly dependent on cooking conditions. HCAs have been commonly quantified in food items prepared with simple procedures. This approach is suitable for elucidating HCAs' formation, but it only partially reflects the contamination in consumed food. In the current investigation, the generation of HCAs has been investigated in fried beef items prepared with elaborated cooking recipes, and their occurrence has been compared with control beef fried without the addition of ingredients other than oil. The food recipes that included a variety of food ingredients had lower yields of mutagenic HCAs (≥47% reduction, with individual HCA levels ranging between 0.01 and 2.22 ng/g) with respect to the control beef. In contrast, the co-mutagens norharman and harman were formed generally at greater levels (up to three times the contamination in the control fried beef) in the items prepared including a greater variety of ingredients.

Topics: Amines; Animals; Carbolines; Cattle; Cooking; Food Contamination; Harmine; Heterocyclic Compounds; Mutagens; Red Meat

Heterocyclic aromatic amines (HAAs) are process-induced food contaminants with high mutagenic and/or carcinogenic potential. Although the human gut microbiota is known to affect the metabolism of dietary constituents, its impact on HAA metabolism and toxicity has been little studied. Here, the glycerol-dependent metabolism of seven foodborne HAAs (AαC, Trp-P-1, harman, norharman, PhIP, MeIQx, and MeIQ) by the human fecal microbiota is investigated.. As analyzed by HPLC-DAD/FLD, the extent of conversion is strongly dependent on glycerol supplementation and HAA structure. AαC (60-100%) and the 2-aminoimidazoazarenes (up to 58%) are especially prone to microbial conversion. Based on high-resolution MS and/or NMR spectroscopy data, 70 fecal metabolites are identified in total, mainly formed by chemical reactions with one or two molecules of microbially derived reuterin. Moreover, it has been demonstrated that the human fecal microbiota can further transform reuterin adducts by reduction and/or hydroxylation reactions. Upon isolation, some reuterin-induced HAA metabolites appear to be partially unstable, complicating structural identification.. The formation of microbial metabolites needs to be incorporated into risk assessment considerations for HAAs in human health. In this study, several HAA metabolites, mainly reuterin-dependent, are identified in vitro, providing the basis for future human studies investigating microbial HAA metabolism.

Topics: Adult; Amines; Animals; Carbolines; Feces; Female; Food Contamination; Gastrointestinal Microbiome; Glyceraldehyde; Harmine; Heterocyclic Compounds, Fused-Ring; Humans; Male; Microsomes, Liver; Middle Aged; Propane; Quinolines; Quinoxalines; Rats, Wistar

An untargeted metabolomic method based on UPLC-QTOF were used to investigate the differences in coffee brewed by boiled, pour-over and cold-brew methods here. Distinctive separation among the three groups could be seen from principal component analysis and hierarchical clustering analysis. Analysis of variance, fold change and orthogonal projection to latent structures discriminant mode were conducted to find the characteristic potential markers, subsequently, nine potential markers were putatively identified using general chemical databases, and five of them were further confirmed by acquisition of reference standards. This work provides an efficient way for discrimination of coffee brewed by different methods. Interestingly, the result of this work also suggested that the contents of two selected markers, norharman and harman, were higher in the pour-over and boiled methods, compared to the cold-brew method. This content difference were further verified by the quantitative analysis data of commercial coffee samples.

Topics: Biomarkers; Carbolines; Chromatography, High Pressure Liquid; Cluster Analysis; Coffee; Cooking; Food Analysis; Harmine; Mass Spectrometry; Metabolomics; Principal Component Analysis

Heterocyclic amines (HCAs) are primarily produced during high temperature meat cooking. These compounds have been intensively investigated as mutagens and carcinogens. However, converging data suggest that HCAs may also be neurotoxic and potentially relevant to neurodegenerative diseases such as Parkinson's disease (PD). The identification of new potential etiological factors is important because most PD cases are sporadic. Our group previously showed that 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) was selectively neurotoxic to dopaminergic neurons. However, PhIP is one of many HCAs, a class of compounds that exhibits wide structural variability. The goal of this study was to determine the neurotoxicity of the most prevalent and best studied HCAs from three subclasses: aminoimidazoaazarenes (AIA), α-carbolines, and β-carbolines. Using E17 rat primary midbrain cultures, we tested dopaminergic and non-dopaminergic neurotoxicity elicited by the following compounds: 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 2-amino-3,8-dimethylmidazo[4,5-f]quinoxaline (MeIQx), 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx), PhIP, 1-methyl-9H-pyrido[3,4-b]indole (harmane), 9H-pyrido[3,4-b]indole (norharmane) and 2-amino-9H-pyrido[2,3-b]indole (AαC) at concentrations ranging from 100 nM-5 μM. All tested HCAs were selectively neurotoxic, though the dose required to elicit selective loss of dopaminergic neurons or decreases in dopaminergic neurite length was compound specific. Non-dopaminergic neurons were unaffected at all tested doses. The sensitivity (determined by threshold dose required to elicit selective neurotoxicity) appears to be unrelated to published mutagenic potency. Both AIA and α/β-carbolines produced oxidative damage, which was magnified in dopaminergic neurons vs. non-dopaminergic neurons as further evidence of selective neurotoxicity. These studies are expected to prompt clinical and mechanistic studies on the potential role of HCA exposure in PD.

Topics: Amines; Animals; Carbolines; Dopaminergic Neurons; Dose-Response Relationship, Drug; Harmine; Heterocyclic Compounds, 3-Ring; Mesencephalon; Molecular Structure; Nerve Degeneration; Neurites; Neurons; Primary Cell Culture; Quinolines; Quinoxalines; Rats

Data from National Health and Nutrition Examination Survey for US population aged ≥ 6 years for 2013-2014 were used to analyze data for four heterocyclic aromatic amines (HCAA), namely 2-amino-9H-pyrido[2,3-b]indole (AαC), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhlP), harman, and norharman. Data were analyzed separately for children aged 6-11 years (N = 416), adolescents aged 12-19 years (N = 475), adults aged 20-64 years (N = 1913), and seniors aged ≥ 65 years (N = 458). Adult males had lower concentrations of AαC and harman than adult females (1.44 vs. 2.22 pg/mL for AαC, p < 0.01 and 136.8 vs. 163.2 pg/mL for harman, p = 0.04). Racial/ethnic differences were observed in the adjusted concentrations of HCAAs. For adults, adjusted concentrations of HCAAs were lower for non-Hispanic Asians and Hispanics as compared to non-Hispanic blacks and whites. For example for AαC, the adjusted concentrations for non-Hispanic Asians, Hispanics, non-Hispanic blacks and whites were 1.16, 2.00, 2.37, and 2.16 pg/mL respectively. Adjusted concentrations of AαC were found to be lower among nonsmokers as compared to smokers for adolescents (0.34 vs. 1.32 pg/mL, p < 0.01), adults (0.40 vs. 7.91 pg/mL, p < 0.01), and seniors (0.30 vs. 4.29 pg/mL, p < 0.01). For both harman and norharman, adult nonsmokers had lower adjusted concentrations than smokers (125.7 vs. 177.6 pg/mL, p < 0.01 for harman, 296.1 vs. 421.6 pg/mL, p < 001, for norharman). Exposure to environmental tobacco smoke was found to be associated with higher concentrations of AαC among adolescents (p = 0.01) and adults (p = 0.01) and for harman (p = 0.01) and norharman (p = 0.01) among seniors. In conclusion, concentrations of selected HCAAs can be several fold higher among smokers as compared to nonsmokers and gender as well as race/ethnicity also affect the observed concentrations of HCAA.

Topics: Adolescent; Adult; Aged; Air Pollution, Indoor; Carbolines; Child; Environmental Exposure; Female; Harmine; Humans; Imidazoles; Male; Middle Aged; Nutrition Surveys; Racial Groups; Tobacco Smoke Pollution; United States; Young Adult

Naturally occurring entomopathogenic fungi such as Conidiobolus coronatus are important regulatory factors of insect populations. GC-MS analysis of fungal cell-free filtrates showed that C. coronatus synthesizes two β- carboline alkaloids: harman and norharman. Significantly higher levels of both alkaloids are produced by C. coronatus in minimal postincubation medium than in rich medium. The beta-carboline alkaloids may have an effect on the nervous system of insects and their behavior. Harman and norharman were applied to Galleria mellonella larvae (a parasite of honeybees) either topically or mixed with food. Larvae received alkaloids in three concentrations: 750, 1000 or 1250 ppm. The effect on the survival and further development of larvae was examined. Both harman and norharman delayed pupation and adult eclosion, and inhibit total monoamine oxidase activity. In addition, they increased the serotonin concentration and decreased the monoamine oxidase A level in the heads of the moths. It is likely that the alkaloids were metabolized by the insects, as their effect wore off 24 hours after topical application. This is the first study to show that C. coronatus produces alkaloids. Its aim was to identify the actions of β-carboline alkaloids on insect development and serotonin-regulating enzymes. Knowledge of the potential role of harman and norharman in the process of fungal infection might lead to the development of more effective and environmentally-friendly means of controlling insect pests.

Topics: Animals; Carbolines; Conidiobolus; Gas Chromatography-Mass Spectrometry; Gene Expression Regulation, Developmental; Harmine; Insect Proteins; Larva; Monoamine Oxidase; Moths; Serotonin; Solid Phase Extraction

Eudistomin U is a member of the β-carboline class of heterocyclic amine-containing molecules that are capable of binding to DNA. The structure of eudistomin U is unique since it contains an indole ring at the 1-position of the pyridine ring. While simple β-carbolines are reported to intercalate DNA, an examination of the mode of binding of eudistomin U has been lacking. We report preliminary spectroscopic (UV-Vis, thermal denaturation, CD) and calorimetric (DSC) data on the binding of eudistomin U to DNA, which suggest that eudistomin U binds weakly according to a mechanism that is more complicated than other members of its class.

Topics: Calorimetry, Differential Scanning; Carbolines; Circular Dichroism; DNA; Spectrophotometry, Ultraviolet; Structure-Activity Relationship

The naturally occurring β-carboline, harmane, has been implicated in various physiological and psychological conditions. Some of these effects are attributed to its interaction with monoaminergic systems. Previous literature indicates that certain β-carbolines including harmane modulate central monoamine levels partly through monoamine oxidase (MAO) inhibition. However, this is not always the case and thus additional mechanisms may be involved. This study set to assess the potential modulatory role of harmane on the basal or K(+) stimulated release of preloaded radiolabelled noradrenaline (NA), dopamine (DA) and serotonin (5-HT) in rat brain cortex in vitro in the presence of the MAO inhibitor pargyline. Harmane displayed an overt elevation in K(+) -evoked [(3)H]5-HT release; whilst little and no effect was reported with [(3)H]DA and [(3)H]NA respectively. The effect of harmane on [(3)H]5-HT efflux was partially compensated in K(+)-free medium. Further analyses demonstrated that removal of Ca(2+) ions and addition of 1.2mM EGTA did not alter the action of harmane on [(3)H]5-HT release from rat brain cortex. The precise mechanism of action however remains unclear but is unlikely to reflect an involvement of MAO inhibition. The current finding aids our understanding on the modulatory action of harmane on monoamine levels and could potentially be of therapeutic use in psychiatric conditions such as depression and anxiety.

Topics: Animals; Calcium; Calcium Chelating Agents; Carbolines; Cerebral Cortex; Dopamine; Dose-Response Relationship, Drug; Egtazic Acid; Harmine; Ions; Male; Monoamine Oxidase Inhibitors; Norepinephrine; Pargyline; Potassium; Rats, Wistar; Serotonin; Serotonin Agents; Synaptic Transmission; Tissue Culture Techniques

The β-carboline compounds norharman and harman exhibit neuroactive activity in the human body. Chicory coffee has proved to be a source of β-carboline compounds. This study assessed the norharman and harman contents of traditional and novel raw materials for the production of chicory coffee, as well as in samples of chicory coffee with novel additives. The highest content of the β-carbolines among the traditional raw materials was recorded in roasted sugar beet (2.26 μg/g), while roasting the chicory caused a 25-fold increase in the content of norharman in this raw material (from 0.05 to 1.25 μg/g). In novel raw materials not subjected to the action of high temperature, β-carboline was not detected. Among the roasted novel raw materials, the highest contents of harman and norharman were found in artichokes. High harman levels were also recorded in roasted chokeberry.

Topics: Beverages; Carbolines; Cichorium intybus; Coffee; Harmine; Hot Temperature

Iodobenzene diacetate was employed as a mild and efficient reagent for one-pot oxidative decarboxylation of tetrahydro-β-carboline acids and dehydrogenation of tetrahydro-β-carbolines to access the corresponding aromatic β-carbolines. To the best of our knowledge this is the first synthesis of β-carbolines via a one-pot oxidative decarboxylation at ambient temperature. The utility of this protocol has been demonstrated in the synthesis of β-carboline alkaloids norharmane (2o), harmane (2p), eudistomin U (9) and eudistomin I (12).

Topics: Acetates; Carbolines; Decarboxylation; Harmine; Iodobenzenes; Molecular Structure; Oxidation-Reduction

β-Carbolines are indole alkaloids that occur in plants, foods, and endogenously in mammals and humans, and which exhibit potent biological, psychopharmacological and toxicological activities. They form from naturally-occurring tetrahydro-β-carboline alkaloids arising from tryptophan by still unknown way and mechanism. Results in this research show that heme peroxidases catalyzed the oxidation of tetrahydro-β-carbolines (i.e. 1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid and 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid) into aromatic β-carbolines (i.e. norharman and harman, respectively). This oxidation followed a typical catalytic cycle of peroxidases through redox intermediates I, II, and ferric enzyme. Both, plant peroxidases (horseradish peroxidase, HRP) and mammalian peroxidases (myeloperoxidase, MPO and lactoperoxidase, LPO) catalyzed the oxidation in an efficient manner as determined by kinetic parameters (VMAX and KM). Oxidation of tetrahydro-β-carbolines was inhibited by peroxidase inhibitors such as sodium azide, ascorbic acid, hydroxylamine and excess of H2O2. The formation of aromatic β-carbolines by heme peroxidases can help to explain the presence and activity of these compounds in biological systems.

Topics: Ascorbic Acid; Carbolines; Enzyme Inhibitors; Harmine; Heme; Horseradish Peroxidase; Hydroxylamine; Kinetics; Lactoperoxidase; Oxidation-Reduction; Peroxidase; Peroxidases; Sodium Azide; Tryptophan

The simultaneous determination of harman and norharman using second derivative synchronous fluorescence method has been developed based on their natural fluorescence. Due to their similar molecular structures, it is difficult to determine them simultaneously in the mixture using conventional fluorimetry. Overlapping of fluorescence spectra was resolved by using a constant second derivative synchronous fluorimetry. The derivative synchronous spectrum, maintaining a constant difference of Δλ=150 nm between emission and excitation for both the compounds, has been selected for the analysis. The range of application is between 0.182 and 18.2 μg/mL (correlation coefficient, R=0.9982) for harman and between 0.504 and 16.8 μg/mL (correlation coefficient, R=0.9962) for norharman. The recovery ranges of 98.5-101.1% for harman and 97.5-99.1% for norharman from their synthetic mixture was reported. The detection limits are 0.016 μg/mL and 0.038 μg/mL for harman and norharman, respectively. Similarly, the quantitation limit of the two compounds was found to be 0.049 and 0.109 μg/mL, respectively. The method was applied to the simultaneous determination of both compounds in coffee samples with satisfactory results.

Topics: Carbolines; Coffee; Harmine; Limit of Detection; Neurotoxins; Spectrometry, Fluorescence

Heterocyclic amines (HAs) are formed as Maillard reaction products in the crust of meat products during heating processes. Two typical pizza toppings--salami and cooked ham--were analyzed for the presence of HAs after baking frozen pizzas at top and bottom temperatures of 250 and 230 °C, respectively. After baking pizza slices for 12 min, MeIQx (2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline; 0.2 ng/g), 4,8-DiMeIQx (2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline; 0.5 ng/g), PhIP (2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine; 0.2 ng/g), norharman (4.5 ng/g), and harman (2.5 ng/g) were found in the ham toppings, whereas only the comutagenic norharman (107.4 ng/g) and harman (11.4 ng/g) were found in the salami toppings. The content of MeIQx and 4,8-DiMeIQx in ham increased from 0.3 to 1.8 ng/g and 0.8 to 1.6 ng/g, respectively, when the recommended baking time was increased from 15 min (manufacturer's specification) to 18 min at 230 °C. MeIQx was formed in salami when the heating time was extended to 18 min. Moreover, higher concentrations of PhIP in salami or ham slices were found when baking temperatures were 250 °C rather than 230 °C (baking time of 12 min). However, sensory tests showed that panelists preferred longer-baked pizzas due to an increased crispiness. Thus, results show that a substantial formation of HAs may occur in pizza toppings such as ham and salami, with ham being particularly susceptible when compared to salami. Formation of HAs increases with increasing baking time and temperature. The occurrence of the cupping of ham or salami slices during baking may also increase the formation of HAs.

Topics: Amines; Animals; Carbolines; Chromatography, High Pressure Liquid; Color; Consumer Behavior; Cooking; Freezing; Harmine; Hot Temperature; Humans; Imidazoles; Maillard Reaction; Meat Products; Quinoxalines; Swine; Taste

Harmane and norharmane are representative members of the large group of natural β-carboline alkaloids featured with diverse pharmacological activities. In blood, these agents are transported by human serum albumin (HSA) which has a profound impact on the pharmacokinetic and pharmacodynamic properties of many therapeutic drugs and xenobiotics. By combination of various spectroscopic methods, the present contribution is aimed to elucidate how nonesterified fatty acids (FAs), the primary endogenous ligands of HSA, affect the binding properties of harmane and norharmane. Analysis of induced circular dichroism (CD) and fluorescence spectroscopic data indicates the inclusion of the neutral form of both molecules into the binding pocket of subdomain IIIA, which hosts two FA binding sites, too. The induced CD and UV absorption spectra of harmane and norharmane exhibit peculiar changes upon addition of FAs, suggesting the formation of ternary complexes in which the lipid ligands significantly alter the binding mode of the alkaloids via cooperative allosteric mechanism. To our knowledge, it is the first instance of the demonstration of drug-FA cobinding at site IIIA. In line with these results, molecular docking calculations showed two distinct binding positions of norharmane within subdomain IIIA. The profound increase in the affinity constants of β-carbolines estimated in the presence of FAs predicts that the unbound, pharmacologically active serum fraction of these compounds strongly depends on the actual lipid binding profile of HSA.

Topics: Alkaloids; Binding Sites; Carbolines; Fatty Acids; Harmine; Humans; Hydrogen-Ion Concentration; Ligands; Protein Binding; Serum Albumin

Heterocyclic amines (HAs) are an important class of food mutagens and carcinogens produced in meat cooked at high temperature. In the present study, the effects of various cooking methods: boiling, microwave cooking, charcoal-grilling, roasting, deep-frying and pan-frying on the formation of HAs in duck breast were studied. The various HAs formed during cooking were isolated by solid-phase extraction and analysed by HPLC. Results showed that both the varieties and contents of HAs and the cooking loss of duck breast increase along with increasing cooking temperature and time. Pan-fried duck breasts contained the highest amount of total HAs, followed by charcoal-grilling, deep-frying, roasting, microwave cooking and boiling. 9H-pyrido[3,4-b]indole (norharman) and 1-methyl-9H-pyrido[3,4-b]indole (harman) were detected in all of the cooked duck meat, with levels in the range of 0.1-33 ng g⁻¹. 2-Amino-1-methyl-6-phenylimidazo[4,5-f]pyridine (PhIP) was formed easily in duck meat cooked by pan-frying and charcoal-grilling in the range of 0.9-17.8 ng g⁻¹. 2-Amino-3-methylimidazo[4,5-f]quinoline (IQ) was identified in duck meat cooked by charcoal-grilling and pan-frying, in the range of 0.4-4.2 ng g⁻¹. 2-Amino-3,8-dimethyl-imidazo[4,5-f]quinoxaline (MeIQx) was detected in amounts below 4.5 ng g⁻¹ in duck meat cooked by charcoal-grilling, roasting, deep-frying and pan-frying. The other HAs were detected in amounts below 10 ng g⁻¹. Colour development increased with cooking temperature, but no correlation with HAs' content was observed.

Topics: Amines; Animals; Carbolines; China; Chromatography, High Pressure Liquid; Cooking; Ducks; Food Contamination; Harmine; Heterocyclic Compounds; Hot Temperature; Imidazoles; Maillard Reaction; Meat; Microwaves; Mutagens; Quinolines; Quinoxalines; Solid Phase Extraction; Time Factors

Harmane, harmine and norharmane are β-carboline compounds which have been referred to as inverse agonists of benzodiazepine receptors. The effect of these compounds on apomorphine-induced licking behavior was studied in rats. Subcutaneous (s.c.) injection of apomorphine (0.5 mg/kg) induced licking. The licking behavior was counted with a hand counter and recorded for a period of 75 min by direct observation. Intraperitoneal (i.p.) injections of harmane (1.25-5 mg/kg), harmine (2.5-10 mg/kg) and norharmane (1.25-5 mg/kg) significantly reduced the licking behavior. In rats pretreated with reserpine (5 mg/kg, i.p., 18 h before the test), the effects of harmane (4 mg/kg, i.p.), harmine (7.8 mg/kg, i.p.) and norharmane (2.5 mg/kg, i.p.) were unchanged. When flumazenil (2 mg/kg, i.p.) was administered 20 min before apomorphine, it was able to antagonize the effects of harmane, harmine and norharmane. It was concluded that the β-carbolines harmane, harmine and norharmane reduce the licking behavior via an inverse agonistic mechanism located in the benzodiazepine receptors.

Topics: Animals; Apomorphine; Carbolines; Corpus Striatum; Dopamine Agonists; Drug Interactions; Flumazenil; Harmine; Male; Rats; Rats, Wistar; Receptors, GABA-A; Reserpine; Stereotyped Behavior

A rapid and simple procedure for the direct screening of urine samples is described. The method involves microextraction in a packed sorbent (MEPS) that is on-line coupled to a capillary liquid chromatograph with fluorimetric detection. The overall arrangement works as a screening/confirmatory system for monitoring non-polar heterocyclic aromatic amines (HAAs) in urine samples. This configuration allows the selective retention of HAAs from urine on a C(18) MEPS cartridge integrated in the needle of a micro-well plate autosampler. Retained HAAs were eluted with methanol/water (90:10, v/v) and directly injected into the fluorimetric detector. This screening method provides a yes/no binary response that may require confirmation. The samples for which the concentration of HAAs was close to or above the established threshold limit (30 ng mL(-1)) were subjected to capillary liquid chromatography (CLC) for confirmation purposes. A mobile phase of acetonitrile and triethylamine (25 mM) at pH 2.5, through a gradient of composition at a flow rate of 20 μL min(-1), resulted in good separations between the analytes in less than 11 min. This confirmation method allowed the determination of the analytes in the 10-100 ng mL(-1) range for harmane and norharmane and from 20 to 200 ng mL(-1) for 3-amino-1,4-dimethyl-5H-pyrido-[4,3-b] indole (Trp-P-1), 3-amino-1-methyl-5H-pyrido-[4,3-b] indole (Trp-P-2), 2-amino-9H-pyrido-[2,3-b] indole (AαC) and 2-amino-3-methyl-9H-pyrido-[2,3-b] indole (MeAαC), with relative standard deviation (RSD) values between 2.12% and 3.73%, and limits of detection between 1.6 and 5.6 ng mL(-1) for all the HAAs.

Topics: Carbolines; Chromatography, Liquid; Fluorescence; Harmine; Heterocyclic Compounds; Humans; Molecular Structure; Time Factors

The beta-carbolines 1-methyl-9H-pyrido [3,4-b]indole and 9H-pyrido[3,4b]indole have been implicated as having causative roles in a number of human diseases, such as Parkinson's disease and cancer. As they can be formed during the heating of protein-rich food, a number of analytical methodologies have been proposed for their detection and quantification in foodstuff. For this purpose, LC-MS and LC-MS/MS have emerged as the most specific analytical methods, and the quantification is based on the occurrence of unusual ions, such as [M+H-(H(*))]+ and [M+H-2H]+. In this study, we have investigated the formation of these ions by accurate-mass electrospray MS/MS and demonstrated that these ions are formed from gas-phase ion-molecule reactions between water vapor present in the collision cell and the protonated molecule of 1-methyl-9H-pyrido [3,4-b]indole and 9H-pyrido[3,4b]indole. Although this reaction has been previously described for heterocyclic amine ions, it has been overlooked in the most of recent LC-MS and LC-MS/MS studies, and no complete data of the fragmentation are reported. Our results demonstrate that additional attention should be given with respect to eliminating water vapor residues in the mass spectrometer when analysis of beta-carbolines is performed, as this residue may affect the reliability in the results of quantification.

Topics: Carbolines; Food Analysis; Food Handling; Food Technology; Harmine; Hot Temperature; Meat; Mutagens; Reproducibility of Results; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry; Water

It has been suggested that the beta-carbolines harmane and norharmane may be involved in the pathophysiology of Parkinson's disease, psychosis and addiction, but the mechanisms of these possible effects remain to be elucidated. In the present study, the effects of the two compounds were examined by using in vivo extracellular recordings of ventral tegmental dopamine neurons. The effects of harmane (2mg/kg) and norharmane (2mg/kg), were compared to those of nicotine (11microg/kg), of cotinine (0.5mg/kg), of the monoamine-oxidase-A inhibitor befloxatone (0.12mg/kg), and of the monoamine-oxidase-B inhibitor selegiline (0.5mg/kg). The effects of harmane were also tested after pre-treatment with the nicotine receptor antagonist mecamylamine. The results show that all substances, except befloxatone, activate the firing and/or burst activity of dopamine neurons. The increase in firing rate produced by harmane was approximately 18 times greater than that produced by nicotine. Such powerful excitation of dopamine neurons by harmane may in part explain its involvement in neurotoxicity, psychosis and addiction. The absence of effect of befloxatone supports the hypothesis that the effect of harmane is not related to its monoamine-oxidase-A inhibitory properties. Mecamylamine inhibited by approximately 80% the activity of harmane, indicating that the activating effect of harmane on dopamine neurons involves several mechanisms, among which activation of nicotinic receptors likely has a prominent importance. The results of the present study support the hypothesis that harmane could be a tobacco (or smoke) component other than nicotine involved in tobacco dependence.

Topics: Animals; Carbolines; Dopamine; Dose-Response Relationship, Drug; Electrophysiological Phenomena; Harmine; Male; Neurons; Nucleus Accumbens; Rats; Rats, Sprague-Dawley; Ventral Tegmental Area

The thermal stability of several commonly used crystalline matrix-assisted ultraviolet laser desorption/ionization mass spectrometry (UV-MALDI-MS) matrices, 2,5-dihydroxybenzoic acid (gentisic acid; GA), 2,4,6-trihydroxyacetophenone (THA), alpha-cyano-4-hydroxycinnamic acid (CHC), 3,5-dimethoxy-4-hydroxycinnamic acid (sinapinic acid; SA), 9H-pirido[3,4-b]indole (nor-harmane; nor-Ho), 1-methyl-9H-pirido[3,4-b]indole (harmane; Ho), perchlorate of nor-harmanonium ([nor-Ho+H]+) and perchlorate of harmanonium ([Ho+H]+) was studied by heating them at their melting point and characterizing the remaining material by using different MS techniques [electron ionization mass spectrometry (EI-MS), ultraviolet laserdesorption/ionization-time-of-flight-mass spectrometry (UV-LDI-TOF-MS) and electrospray ionization-time-of-flight-mass spectrometry (ESI-TOF-MS)] as well as by thin layer chromatography analysis (TLC), electronic spectroscopy (UV-absorption, fluorescence emission and excitation spectroscopy) and 1H nuclear magnetic resonance spectroscopy (1H-NMR). In general, all compounds, except for CHC and SA, remained unchanged after fusion. CHC showed loss of CO2, yielding the trans-/cis-4-hydroxyphenylacrilonitrile mixture. This mixture was unambiguously characterized by MS and 1H-NMR spectroscopy, and its sublimation capability was demonstrated. These results explain the well-known cluster formation, fading (vanishing) and further recovering of CHC when used as a matrix in UV-MALDI-MS. Commercial SA (SA 98%; trans-SA/cis-SA 5:1) showed mainly cis- to-trans thermal isomerization and, with very poor yield, loss of CO2, yielding (3',5'-dimethoxy-4'-hydroxyphenyl)-1-ethene as the decarboxilated product. These thermal conversions would not drastically affect its behavior as a UV-MALDI matrix as happens in the case of CHC. Complementary studies of the photochemical stability of these matrices in solid state were also conducted.

Topics: Acetophenones; Carbolines; Coumaric Acids; Gentisates; Harmine; Hot Temperature; Magnetic Resonance Spectroscopy; Phase Transition; Photochemistry; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

beta-Carbolines are potent modulators of GABA type A receptors and they have recently been shown to inhibit glycine receptors in a subunit-specific manner. The present study screened four structurally similar beta-carbolines, 1,2,3,4-tetrahydronorharmane, norharmane, harmane and 6-methoxyharmalan, at recombinantly expressed alpha1, alpha1beta, alpha2 and alpha3 glycine receptors with the aims of identifying structural elements of both the receptor and the compounds that are important for binding and subunit specificity. The four compounds exhibited only weak subunit specificity, rendering them unsuitable as pharmacological probes. Because they displayed competitive antagonist activity, we investigated the roles of known glycine binding residues in coordinating the four compounds. The structural similarity of the compounds, coupled with the differential effects of C-loop mutations (T204A, F207Y) on compound potency, implied direct interactions between variable beta-carboline groups and mutated residues. Mutant cycle analysis employing harmane and norharmane revealed a strong pairwise interaction between the harmane methyl group and the C-loop in the region T204 and F207. These results which define the orientation of the bound beta-carbolines were supported by molecular docking simulations. The information may also be relevant to understanding the mechanism beta-carboline of binding to GABA type A receptors where they are potent pharmacological probes.

Topics: Binding Sites; Carbolines; Cell Line; Harmine; Humans; Mutagenesis, Site-Directed; Mutation; Protein Structure, Secondary; Protein Subunits; Receptors, Glycine; Recombinant Proteins

Norharman and harman are naturally occurring beta-carboline alkaloids exhibiting a wide range of biological, psychopharmacological, and toxicological actions. They occur in foods and tobacco smoke and also appear endogenously in humans. In this research, metabolic and kinetic studies with cytochrome P450 enzymes and human liver microsomes showed that beta-carbolines were efficiently oxidized to several ring-hydroxylated and N-oxidation products that were subsequently identified and quantified. 6-Hydroxy- beta-carboline (6-hydroxynorharman and 6-hydroxyharman) was a major metabolite efficiently produced (high kcat and low Km) by P450 1A2 and 1A1 and to a minor extent by P450 2D6, 2C19 and 2E1. 3-Hydroxy-beta-carboline (3-hydroxynorharman and 3-hydroxyharman), another major metabolite, was specifically produced by P450 1A2 and 1A1, whereas beta-carboline-N(2)-oxide (harman-2-oxide and norharman-2-oxide) was produced by P450 2E1. The same pattern of metabolism was confirmed for human liver microsomes. Oxidative metabolism for harman was slightly higher than norharman, but norharman showed lower Km values. The oxidation of beta-carbolines is a detoxication route performed mainly by P450 1A2 and 1A1, with the participation of P450 2D6, 2C19, and 2E1, as additional contributors. Then, individual variations in the levels and activity of these P450s may influence biotransformation of beta-carboline alkaloids and their ultimate biological effects. beta-Carbolines were previously reported as comutagens and/or inhibitors of mutagens activated by P450 1A enzymes such as heterocyclic amines and polycyclic hydrocarbons. Results in this work show that beta-carbolines are good ligands and substrates for P450 1A2/1A1, contributing to the explanation of some of their toxicological effects.

Topics: Carbolines; Cytochrome P-450 CYP1A1; Cytochrome P-450 CYP1A2; Cytochrome P-450 Enzyme System; Harmine; Humans; Hydroxylation; Microsomes, Liver; Oxidation-Reduction

The presence of cyclodextrins (CDs) in the mobile phase alters the chromatographic equilibria and induces a secondary chemical equilibrium associated to the chromatographic separation by HPLC. In this study the influence of the presence of CDs in the mobile phase as chemically modified beta-CDs, i.e. 2,3-di-O-methyl-beta-cyclodextrin (DMbeta-CD) and 2,3,6-tri-O-methyl-beta-cyclodextrin (TMbeta-CD) on the separation of the alkaloids norharmane, harmane and harmine is described. These beta-carboline alkaloids are chemically and structurally related and their quantitation by RP-HPLC is of interest due to their biological and pharmacological properties. Two stationary phases (methyl-, C(1) and octadecyl-, C(18)) were employed, with methanol:buffered aqueous solution and ethanol:buffered aqueous solution as mobile phases. The role of tert-butyl alcohol as a mobile phase modifier and also as an inclusion complex stabiliser was also considered. The concentrations of DMbeta-CD and TMbeta-CD vary from 0 to 17 mM. The presence of increasing amounts of CDs in the mobile phase reduces the retention factor. The changes observed in the retention factor allow the determination of the alkaloid/CD apparent association constants, whose magnitude is influenced by the chemical and structural properties of the guest molecules but also by the composition of the mobile phase. Assuming a 1:1 stoichiometry for the inclusion complexes, the apparent association constants obtained were higher for norharmane and for both DMbeta-CD and TMbeta-CD. The strength of the complexes is higher for DMbeta-CD than for TMbeta-CD and this behaviour can be explained considering the steric problems associated to the permethylated-beta-CDs. Besides significant differences in the magnitude of the apparent association constants were observed for the two stationary phases employed and thus can be related to the adsorption of CDs on the stationary phase. A significant reduction in the proportion of organic solvent in the mobile phase (50%) without a decrease in the selectivity or resolution of the separation is a favourable consequence of the presence of the CDs in the mobile phase.

Topics: beta-Cyclodextrins; Carbolines; Chromatography, High Pressure Liquid; Harmine; Indicators and Reagents; Reference Standards; Solvents; Spectrometry, Fluorescence; Spectrophotometry, Ultraviolet

The effects of harman and norharman on dopamine biosynthesis and L-DOPA-induced cytotoxicity in PC12 cells were investigated. Harman and norharman at a concentration of 20 microM and 100 microM showed 49.4% and 49.5% inhibition of dopamine content for 48 h, respectively. The IC50 values of harman and norharman were 21.2 microM and 103.3 microM. Dopamine content, tyrosine hydroxylase (TH) activity and TH mRNA levels were decreased during the first 6 h, maintained for up to 48 h and then gradually recovered at 72 h after exposure to 20 microM harman and 100 microM norharman. Under the same conditions, the intracellular cyclic AMP levels and Ca2+ concentrations were also decreased by harman and norharman. In addition, harman and norharman at concentrations higher than 80 microM and 150 microM caused cytotoxicity at 48 h in PC12 cells. Non-cytotoxic ranges of 10-30 microM harman and 50-150 microM norharman inhibited L-DOPA (20-50 microM)-induced increases in dopamine content at 48 h. Harman at 20-150 microM and norharman at 100-300 microM also enhanced L-DOPA (20-100 microM)-induced cytotoxicity at 48 h with an apoptotic process. These results suggest that harman and norharman inhibit dopamine biosynthesis by reducing TH activity and enhance L-DOPA-induced cytotoxicity in PC12 cells.

Topics: Animals; Antiparkinson Agents; Apoptosis; Aromatic-L-Amino-Acid Decarboxylases; Blotting, Northern; Calcium; Carbolines; Cell Survival; Cyclic AMP; Dopamine; Flow Cytometry; Harmine; In Situ Nick-End Labeling; Levodopa; Membrane Potentials; Mitochondrial Membranes; PC12 Cells; Rats; RNA; Tetrazolium Salts; Thiazoles; Tyrosine 3-Monooxygenase

A recently developed HPTLC/UV-FLD method was compared to the routinely used HPLC/UV-FLD method for the quantification of heterocyclic aromatic amines (HAA) formed at trace levels during the heating process of meat. For formation of these process contaminants under normal cooking conditions, beef patties were fried in a double-contact grill at 230 degrees C for five different frying times and extracted by solid-phase extraction. The HAAs most frequently found, that is, 2-amino-1-methyl-6-phenylimidazo[4,5- b]pyridine (PhIP), 2-amino-3,8-dimethylimidazo[4,5- f]quinoxaline (MeIQx), 2-amino-3,4,8-trimethylimidazo[4,5- f]quinoxaline (4,8-DiMeIQx), 9 H-pyrido[3,4- b]indole (norharman), and 1-methyl-9 H-pyrido[3,4- b]indole (harman), were quantified by two chromatographic methods, which were orthogonal to each other (normal versus reversed phase system). Both methods showed a similar performance and good correlation of the results ( R (2) between 0.8875 and 0.9751). The comparison of running costs and run time in routine analysis proved HPTLC/UV-FLD to be more economical (factor of 3) and faster (factor of 4) due to its capability of parallel chromatography. The HAA findings calculated by standard addition increased with the heating time from <1 to 33 microg/kg related to 3-6 min of frying time. The precision (RSD) was between 7 and 49% (HPTLC) and between 5 and 38% (HPLC) at these very low HAA levels formed.

Topics: Amines; Animals; Carbolines; Carcinogens; Cattle; Chromatography, High Pressure Liquid; Chromatography, Thin Layer; Harmine; Heterocyclic Compounds; Hot Temperature; Imidazoles; Meat; Mutagens; Quinoxalines

Espresso coffee (EC) brews were analyzed for beta-carboline [norharman (NH) and harman (H)] contents, by RP-HPLC with fluorescence detection. The influence of the coffee species (arabica or robusta), the roast degree, and the brew length was studied. The results show that the content of NH and H in EC is dependent primarily on the coffee species, followed by brew length. The roast degree has only a minor influence on the final content of NH and H in EC. When compared with other coffee brews, EC has an amount of these beta-carbolines (in micrograms per liter) similar to that of mocha coffee, both being more concentrated than filter and press-pot coffees. Therefore, the consumer's preferences will determine the amount of NH and H ingested daily. For the caffeinated 30 mL of EC, the arabica coffees contain about 4.08 microg of NH and 1.54 microg of H. Commercial blends (usually with a maximum of 30% robusta) range from the cited arabica values to 10.37 microg of NH and 4.35 microg of H.

Topics: Carbolines; Coffea; Coffee; Food Handling; Harmine; Hot Temperature; Mutagens; Neurotoxins; Pressure; Seeds; Species Specificity; Time Factors

Monoamine oxidase (MAO) is a mitochondrial outer-membrane flavoenzyme involved in brain and peripheral oxidative catabolism of neurotransmitters and xenobiotic amines, including neurotoxic amines, and a well-known target for antidepressant and neuroprotective drugs. Recent epidemiological studies have consistently shown that coffee drinkers have an apparently lower incidence of Parkinson's disease (PD), suggesting that coffee might somehow act as a purported neuroprotectant. In this paper, "ready to drink" coffee brews exhibited inhibitory properties on recombinant human MAO A and B isozymes catalyzing the oxidative deamination of kynuramine, suggesting that coffee contains compounds acting as MAO inhibitors. MAO inhibition was reversible and competitive for MAO A and MAO B. Subsequently, the pyrido-indole (beta-carboline) alkaloids, norharman and harman, were identified and isolated from MAO-inhibiting coffee, and were good inhibitors on MAO A (harman and norharman) and MAO B (norharman) isozymes. beta-carbolines isolated from ready-to-drink coffee were competitive and reversible inhibitors and appeared up to 210 microg/L, confirming that coffee is the most important exogenous source of these alkaloids in addition to cigarette smoking. Inhibition of MAO enzymes by coffee and the presence of MAO inhibitors that are also neuroactive, such as beta-carbolines and eventually others, might play a role in the neuroactive actions including a purported neuroprotection associated with coffee consumption.

Topics: Carbolines; Chromatography, High Pressure Liquid; Coffee; Harmine; Humans; Isoenzymes; Monoamine Oxidase; Monoamine Oxidase Inhibitors; Neuroprotective Agents; Recombinant Proteins; Spectrometry, Mass, Electrospray Ionization

The purpose of the present study was to determine the effects of harmane, norharmane and harmine on the immobility time in the mouse forced swim test (FST) - an animal model of depression. After 30 min of the beta-carbolines injections, mice were placed individually in a vertical glass cylinder (height, 25 cm; diameter, 12 cm) containing water about 15 cm deep at 22+/-1 degrees C and forced to swim. Treatment of animals with harmane (5-15 mg/kg, i.p.), norharmane (2.5-10 mg/kg, i.p.) and harmine (5-15 mg/kg, i.p.) reduced dose-dependently the time of immobility. Their antidepressant-like effects were not affected by pretreatment with reserpine at the dose of 5 mg/kg, i.p., 18 h before the test, which did not modify the immobility time. Conversely, when flumazenil (5 mg/kg, i.p.) was administered 30 min before the test, it was able to antagonize completely the antidepressant-like effects of harmane, norharmane and harmine. It was concluded that harmane, norharmane and harmine reduce the immobility time in this test, suggesting an antidepressant-like effect, via an inverse-agonistic mechanism located in the benzodiazepine receptors.

Topics: Animals; Antidepressive Agents; Behavior, Animal; Carbolines; Dose-Response Relationship, Drug; Drug Interactions; Flumazenil; GABA Modulators; Harmine; Male; Mice; Monoamine Oxidase Inhibitors; Motor Activity; Reserpine; Swimming; Sympatholytics

N-nitrosocompounds, which induce cancers in various organs, may be formed endogenously with intake of amino compounds such as secondary amines and sodium nitrite (NaNO(2)) in combination. The present study was performed to investigate whether three amino compounds, 1-methyl-9H-pyrido[3,4-b]indole (harman), 9H-pyrido[3,4-b]indole (norharman) and 2-amino-1,3,4-triazole (amitrole), might be converted in vivo to compounds capable of promoting hepatocarcinogenesis when given with NaNO(2). However, in an 8-week model, no modifying potential was evident in terms of numbers and areas of putative preneoplastic glutathione S-transferase placental form (GST-P)-positive foci in any of the groups receiving paired treatments. These results demonstrate that combinations of harman, norharman and amitrole with NaNO(2) lack promoting effects for liver carcinogenesis in our medium-term bioassay system.

Topics: Amitrole; Animals; Carbolines; Harmine; Liver Neoplasms, Experimental; Male; Rats; Rats, Inbred F344; Sodium Nitrite; Weight Gain

Reversed phase ion-pair chromatography (RP-IPC) of seven heterocyclic aromatic amines encompassing quinoline (IQ, MeIQ), quinoxaline (MeIQx), pyridine (PhIP) and carboline derivatives (AalphaC, Harman, Norharman) was carried out with formate as counter ion in an aqueous eluent with acetonitrile as organic modifier. TSKgel ODS-80TS was used as the stationary phase. With the aim of acquiring a better insight into the mutual influence of ion-pair reagent and the organic modifier upon solute retention, the study was performed by using an experimental design approach able to evidencing the effect of the simultaneous variation of the two factors. A model for the chromatographic behavior of the amines is proposed that includes classical ion-pair mechanism involving formate in the case of MeIQx, PhIP, Harman and Norharman. A competitive ion-exchange mechanism was hypothesized to govern retention of quinoline compounds, whereas electrostatic interactions and hydrogen bond formation with the silanols of the stationary phase were judged to be responsible for the retention of AalphaC. Further, the chromatographic behavior of the analytes using the formic acid-ammonium formate buffer in the mobile phase was compared with that observed using acetic acid-ammonium acetate buffer. The method based on the use of RP IPC with tandem mass spectrometry when the eluent contained formate buffer at pH 2.8 exhibited higher detectability with respect to that achieved using the acetate buffer.

Topics: Acetonitriles; Amines; Buffers; Carbolines; Chromatography, Liquid; Formates; Harmine; Heterocyclic Compounds; Hydrogen-Ion Concentration; Imidazoles; Quinolines; Quinoxalines; Sensitivity and Specificity; Spectrometry, Mass, Electrospray Ionization

Harmane and norharmane (two beta-carbolines) are tobacco components or products. The effects of harmane and norharmane on serotonergic raphe neurons remain unknown. Harmane and norharmane are inhibitors of the monoamine oxidases A (MAO-A) and B (MAO-B), respectively.. To study the effects of harmane, norharmane, befloxatone (MAOI-A), and selegiline (MAOI-B) on the firing of serotonergic neurons. To compare the effects of these compounds to those of nicotine (whose inhibitory action on serotonergic neurons has been previously described). The effects of cotinine, a metabolite of nicotine known to interact with serotonergic systems, are also tested.. In vivo electrophysiological recordings of serotonergic dorsal raphe neurons in the anaesthetized rat.. Nicotine, harmane, and befloxatone inhibited serotonergic dorsal raphe neurons. The other compounds had no effects. The inhibitory effect of harmane (rapid and long-lasting inhibition) differed from that of nicotine (short and rapidly reversed inhibition) and from that of befloxatone (slow, progressive, and long-lasting inhibition). The inhibitory effects of harmane and befloxatone were reversed by the 5-HT1A antagonist WAY 100 635. Pretreatment of animals with p-chlorophenylalanine abolished the inhibitory effect of befloxatone, but not that of harmane.. Nicotine, harmane, and befloxatone inhibit the activity of raphe serotonergic neurons. Therefore, at least two tobacco compounds, nicotine and harmane, inhibit the activity of serotonergic neurons. The mechanism by which harmane inhibits serotonergic dorsal raphe neurons is likely unrelated to a MAO-A inhibitory effect.

Topics: Action Potentials; Analysis of Variance; Animals; Carbolines; Citalopram; Dose-Response Relationship, Drug; Drug Interactions; Enzyme Inhibitors; Harmine; Male; Neurons; Neurotoxins; Nicotine; Nicotinic Agonists; Oxazoles; Phenethylamines; Piperazines; Pyridines; Raphe Nuclei; Rats; Rats, Sprague-Dawley; Selective Serotonin Reuptake Inhibitors; Selegiline; Serotonin; Serotonin Antagonists

Norharman and harman are two heterocyclic beta-carboline (9H-pyrido[3,4-b]indole) alkaloids with biological and potential toxicological activity that appear in foodstuffs and environmental sources. To assess the occurrence and distribution of these compounds and to estimate the exposure levels based on the detected amounts, numerous samples of foodstuffs and cigarette smoke were analysed by solid-phase extraction and high-performance liquid chromatography-fluorescence. The levels found of beta-carbolines were highly variable. Low processed foodstuffs (i.e. milk, yoghurt, uncooked meats and fish) did not contain norharman and harman above the detection limit. Others, however, contained relatively high concentrations (at the tens of ng g(-1) or microg l(-1) level) depending on the processing conditions as, for example, 'well-done' cooked meat and fish. The highest amounts of norharman and harman were found in brewed coffee (29-207 microg l(-1)), sauces (soy sauce and Tabasco, among others; 4-252 microg l(-1)), 'well done' cooked meat and fish (57-160 ng g(-1)), toasted bread (42-160 ng g(-1)), and fermented alcoholic beverages (n.d.-41 mug l(-1)). beta-Carbolines also occurred in a high amount in the mainstream of cigarette smoke (207-2780 ng/cigarette), which is an important contributor to daily exposure to these compounds. Based on these results, it is concluded that the daily exposure to beta-carbolines in humans might be from tens to hundreds of micrograms, with cigarette smoke, coffee, certain seasonings, cooked foods and alcoholic beverages, in this order, being the major contributors. Many other foodstuffs might also contribute with minor amounts of norharman and harman. Foods and tobacco smoke might be potential contributors to the reported endogenous presence of beta-carbolines in humans.

Topics: Carbolines; Chromatography, High Pressure Liquid; Environmental Exposure; Food Analysis; Food Contamination; Food Handling; Harmine; Mutagens; Neurotoxins; Nicotiana; Smoke

Norharman and harman, two heterocyclic beta-carboline alkaloids with biological activity, were found in brewed coffee. Identification and analysis were carried out by HPLC-MS and RP-HPLC-fluorescence, respectively. All tested samples of brewed coffee including ground coffee, decaffeinated coffee, instant coffee and espresso contained both norharman and harman in variable amounts. Norharman was the major beta-carboline alkaloid in brewed coffee at levels up to 9.34 microg g(-1) in instant ground coffee compared with harman, which had levels up to 1.67 microg g(-1). The two beta-carbolines appeared to be formed during roasting of the coffee beans. It is concluded that drinking coffee is a major exogenous dietary source of these bioactive beta-carboline alkaloids previously reported as mild psychoactive compounds in animal studies and in vitro co-mutagens. These results support our previous conclusion that foods containing beta-carbolines are an important exogenous source of these alkaloids in humans.

Topics: Alkaloids; Carbolines; Chromatography, High Pressure Liquid; Coffee; Food Contamination; Harmine; Mass Spectrometry

Tetrahydro-beta-carboline alkaloids that occur in foods such as wine, seasonings, vinegar and fruit products juices, jams) acted as good radical scavengers (hydrogen- or electron donating) in the ABTS (2,2'-Azinobis-(3-ethylbenzothiazoline-6-sulfonic acid)) assay, and therefore, they could contribute to the beneficial antioxidant capacity attributed to foods. In contrast, the fully aromatic beta-carbolines norharman and harman did not show any radical scavenger activity in the same assay. During the reaction with ABTS.+ radical cation, tetrahydro-beta-carboline-3-carboxylic acid such as 1-methyl-1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid (MTCA) and 1-methyl-1,2,3,4-tetrahydro-beta-carboline-1,3-dicarboxylic acid (MTCA-COOH) were converted to harman, whereas 1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid (THCA) and 1,2,3,4-tetrahydro-beta-carboline-1,3-dicarboxylic acid (THCA-COOH) afforded norharman. These results suggest that food and naturally-occurring tetrahydro-beta-carboline alkaloids if accumulated in tissues, as reported elsewhere, might exhibit antioxidant activity.

Topics: Antioxidants; Benzothiazoles; Carbolines; Cations; Chromatography, High Pressure Liquid; Dose-Response Relationship, Drug; Food Analysis; Free Radicals; Gas Chromatography-Mass Spectrometry; Harmine; Indicators and Reagents; Models, Chemical; Sulfonic Acids; Time Factors

Harmane, harmaline and norharmane are beta-carboline related compounds which have been proposed to be endogenous ligands for imidazoline receptors. The effect of these compounds on the activity of locus coeruleus (LC) neurons was studied by extracellular recordings techniques. Intracerebroventricular administration of harmane and harmaline increased the firing rate of LC neurons. Systemic administration of efaroxan, a mixed alpha(2)-adrenoceptor/I(1)-imidazoline antagonist or vagotomy failed to modify the harmane effect. Furthermore, local applications of harmane and harmaline increased the firing rate of LC neurons in a dose-related manner. Finally, intravenous administration of norharmane also increased the activity of LC neurons. Our results demonstrate that beta-carbolines stimulate LC neuron activity and indicate that this stimulation occurs directly in the LC by a mechanism independent of I(1)- and I(2)-imidazoline receptors.

Topics: Anesthesia; Animals; Carbolines; Electrophysiology; Harmaline; Harmine; Imidazoline Receptors; Locus Coeruleus; Male; Neurons; Neurotoxins; Rats; Rats, Sprague-Dawley; Receptors, Drug; Vagotomy

Motivated by the identification of numerous novel tetrahydro-beta-carboline-carboxylic acids in food samples, we studied the reactions of tetrahydro-beta-carbolines in the presence of nitrosating agents. The anticipated formation of nitroso derivatives from unsubstituted tetrahydro-beta-carbolines, and from tetrahydro-beta-carboline-3-carboxylic acids was indicated by HPLC-MS/MS analysis and validated by the characteristic product ion spectra of the respective nitroso compounds. In addition, oxidative decarboxylation resulted in formation of the corresponding dihydro-beta-carbolines, and in the generation of the beta-carbolines harman or norharman. Subsequently, we studied the reactivity of tetrahydro-beta-carboline-1-carboxylic acids derived from the Pictet-Spengler condensation of indole amines with alpha-oxo acids. Again, in the presence of nitrosating agents the rapid disappearance of the starting material was obvious, but no nitroso derivatives could be observed. Instead, further HPLC-MS/MS studies demonstrated that dihydro-beta-carbolines were the major products of tetrahydro-beta-carboline-1-carboxylic acids. Finally, we demonstrated that freshly isolated nitroso-precursors spontaneously decomposed to yield harman alkaloids. In conclusion, we revealed that nitroso-tetrahydro-beta-carbolines can represent intermediates involved in the generation of beta-carbolines, and we established a novel pathway for the formation of harman alkaloids from nutritional tetrahydro-beta-carbolines.

Topics: Carbolines; Chromatography, High Pressure Liquid; Food Analysis; Harmine; Kinetics; Mass Spectrometry; Molecular Conformation; Nitrites

Harman and norharman are widely distributed in the environment and consequently contaminate in domestic waste-water. It has been reported that they have co-mutagenic activity in the presence of non- mutagenic aromatic amines such as aniline and o-toluidine with S9 mix. When these beta-carbolines were treated with sodium hypochiorite under mild conditions, chlorinated derivatives were produced. Among them, 6-chloroharman and 6-chloronorharman showed much more potent co-mutagenic activities than harman and norharman in the presence of o-toluidine toward Salmonella typhimurium TA98 with S9 mix. These results suggest that the chlorination of harman and norharman occurs during disinfection at the sewage plant to produce potent co-mutagens that contaminate river water.

Topics: Biotransformation; Carbolines; Harmine; Mutagenicity Tests; Mutagens; Salmonella typhimurium; Sodium Hypochlorite

The presence of tetrahydro-beta-carbolines and beta-carbolines was studied in raw, cooked and smoked fish and meat. 1,2,3,4-Tetrahydro-beta-carboline-3-carboxylic acid (THCA) usually was the major beta-carboline found, whereas 1-methyl-1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid (MTCA) appeared in smoked and 'well done' cooked samples. THCA was detected in raw fish (nd-2.52 micrograms/g), cooked fish (nd-6.43 micrograms/g), cooked meats (nd-0.036 microgram/g), smoked fish (0.19-0.67 microgram/g) and smoked meats (0.02-1.1 micrograms/g). Smoked and cooked samples contained higher amounts of THCA and MTCA than raw products. Deep cooking of fish and meat increased both THCA and MTCA, and this was accompanied by the formation of more beta-carbolines, norharman and harman. The tetrahydro-beta-carbolines THCA and MTCA were chemical precursors of the co-mutagens norharman and harman during cooking. These and previous results confirm that foods are an important source of beta-carbolines in humans.

Topics: Animals; Carbolines; Cattle; Chromatography, High Pressure Liquid; Food Handling; Food Preservation; Harmine; Meat; Mutagens; Seafood; Swine

Co-mutagenic beta-carbolines, such as norharman and harman, were quantified in mainstream and sidestream smoke condensates of six Japanese brands of cigarettes, and also in 13 kinds of cooked foods, using a combination of blue cotton treatment and HPLC. Norharman and harman were detected in all the cigarette smoke condensate samples. Their levels in the mainstream smoke case were 900-4240 ng per cigarette for norharman, and 360-2240 ng for harman, and in sidestream smoke, 4130-8990 ng for norharman and 2100-3000 ng for harman. These beta-carbolines were also found to be present in all the cooked food samples, at levels of 2.39-795 ng for norharman and 0.62-377 ng for harman per gram of cooked food. The observed concentrations are much higher than those found for mutagenic and carcinogenic heterocyclic amines (HCAs), suggesting that humans are exposed to norharman and harman in daily life to a larger extent than to HCAs.

Topics: Carbolines; Food; Harmine; Humans; Mutagens; Tobacco Smoke Pollution

Heat processing of muscle foods gives rise to the formation of mutagenic and carcinogenic heterocyclic amines, often at ng/g levels. A gas chromatographic-mass spectrometric (GC-MS) technique was introduced for the analysis of nonpolar heterocyclic amines in common cooked meats, pan residues, and meat extracts after solid-phase extraction. The mutagenic heterocyclic amines 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1), 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2), 2-amino-9H-pyrido[2,3-b]indole (A alpha C) and 2-amino-3-methyl-9H-pyrido[2,3-b]indole (MeA alpha C) were identified in several samples in amounts up to 8 ng/g. Also the comutagenic substances 1-methyl-9H-pyrido [3,4-b]indole (harman) and 9H-pyrido[3,4-b]indole (norharman) were detected in the samples in amounts up to almost 200 ng/g. The GC-MS method can be applied without derivatisation of the sample. The technique offers high chromatographic efficiency, yielding detection limits for pure references in the range 0.1-2 ng per injection.

Topics: Amines; Animals; Carbolines; Carcinogens; Cooking; Gas Chromatography-Mass Spectrometry; Harmine; Heterocyclic Compounds; Hot Temperature; Meat; Meat Products; Mutagens; Reindeer; Surface Properties; Swine

Topics: Animals; Carbolines; Drug Synergism; Harmine; Mutagenicity Tests; Mutagens; Salmonella typhimurium

Naturally occurring beta-carbolines are lipophilic compounds which show psychotropic and physiological effects in mammals. They bind to distinct high-affinity binding sites in various mammalian tissues. However, the mechanism by which the beta-carbolines affect transmembrane signal transduction processes is still unknown. Since beta-carbolines are cationic-amphiphilic substances and since such substances are known to activate heterotrimeric regulatory guanine nucleotide binding proteins (G-proteins) in a receptor-independent manner, we put forward the hypothesis that beta-carbolines act directly on G-proteins. Therefore, we investigated the ability of beta-carbolines to stimulate high-affinity GTP hydrolysis in membranes of dibutyryl-cAMP differentiated HL-60 cells and of the purified bovine G-protein, transducin (TD). The beta-carbolines norharman and harman, stimulated the GTPase in HL-60 membranes with an EC50 of 410 microM and 450 microM, respectively, and a maximum effect at 1 mM each. Norharman and harman stimulated the GTPase of TD with an EC50 of 60 microM and 300 microM, and a maximum at 1 mM for both compounds. The stimulatory effect of norharman in HL-60 membranes was pertussis toxin-sensitive. Structure/activity characteristics of the beta-carbolines showed a specificity of norharman to stimulate the GTPase of TD, because norharman activated GTP hydrolysis in HL-60 membranes approximately 7 times less potently than that of TD. Norharman was a five-fold more potent activator of TD than tetrahydronorharman. Hydroxylation of the beta-carboline molecule in position 6 led to a loss of GTPase-activating properties. Our data suggest that naturally occurring beta-carbolines are a novel class of receptor-independent G-protein activating substances. This mechanism could contribute to their diverse biological effects.

Topics: Carbolines; Cell Membrane; Enzyme Activation; GTP Phosphohydrolases; GTP-Binding Proteins; Harmine; HL-60 Cells; Humans; Intercellular Signaling Peptides and Proteins; Peptides; Structure-Activity Relationship; Transducin; Wasp Venoms

beta-Carbolines, harman (1-methyl-9H-pyrido[3,4-b]indole) and norharman (9H-pyrido[3,4-b]indole) and gamma-carbolines, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) and 3-amino-4-methyl-5H-pyrido[4,3-b]indole (Trp-P-2), are present in cooked foods and cigarette smoke. We studied the effects of these heterocyclic amines on the activity of DNA topoisomerases. Trp-P-1 and Trp-P-2 inhibited topoisomerase I (topo I) activity with ED50 values of 1.48 and 1.55 micrograms/ml, respectively, in a relaxation assay. Harman and norharman inhibited topo I activity but with much higher ED50 values, 23.8 and 34.4 micrograms/ml, respectively. Trp-P-1 and Trp-P-2 also inhibited topoisomerase II (topo II) activity at about 50 micrograms/ml, in a decatenation assay. Harman and norharman showed a much lower inhibitory effect on topo II activity. None of these compounds stabilized the cleavable complex mediated by topo II. Trp-P-1 and Trp-P-2 intercalated into DNA at concentrations inhibitory to topoisomerases. We considered that the intercalation with DNA and the inhibition of DNA topoisomerases by heterocyclic amines might be partly related to their inhibition of DNA excision repair and their enhancing effect on UV- or chemically induced mutagenic activity.

Topics: Breast Neoplasms; Carbolines; Carcinoma, Small Cell; Cell Line; Enzyme Inhibitors; Female; Harmine; Humans; Kinetics; Mutagens; Topoisomerase I Inhibitors; Topoisomerase II Inhibitors; Tumor Cells, Cultured

In the search for mechanisms specific for alcoholism, it has become evident that beta-carbolines (BCs; e.g., harman and norharman) are compounds that may act on brain reward systems, thereby mediating an increase in voluntary ethanol (ETOH) drinking in animals. This study was undertaken to analyze relationships between these compounds and clinical variables (e.g., family history, personality data, and affect) in alcoholics and to trace the time course of blood concentrations in subjects abstaining from alcohol for at least 6 months. Nonalcoholics were investigated during sober and ETOH-loading conditions (1 g ETOH/kg body weight). Levels of harman were elevated in the chronically intoxicated alcoholics and correlated with the scores on the self-rating depression (SDS) and the self-rating anxiety (SAS) scales. The group of alcoholics with at least one alcoholic parent had higher levels than the group without such a history. Levels remained elevated for 6 months. Norharman levels were only slightly elevated on the day of admission. They were correlated to high harm avoidance and SDS scores. A family history of alcoholism and the severity of alcoholism as assessed by the number of ICD-10 criteria fulfilled were correlated with norharman levels. Long-term observation revealed elevated levels of norharman after 3 months of abstinence, but not after 6 months. The association of harman levels with anxiety and depression demonstrated in the present study suggests that alcoholics with high harman levels use alcoholic beverages as self-medication in an attempt to overcome possible anxiogenic/depressiogenic actions of harman. Norharman levels are less strongly associated with these mood states, but significantly correlated to harm avoidance tendencies. It has been suggested that the activity of the indolergic neurons is relatively high in individuals with a high harm avoidance score. Biosynthesis of norharman might be stimulated under these conditions (tryptamine serves as precursor).

Topics: Adult; Alcoholism; Anxiety; Carbolines; Depression; Ethanol; Harmine; Humans; Male; Middle Aged; Motivation; Self Medication

Electrospray ionization mass spectrometry was applied to the study of the amines IQ, Trp-P-1, Trp-P-2, PhIP and A alpha C and the co-mutagens harman and norharman. The results obtained on a triple quadrupole mass spectrometer equipped with a pneumatically assisted electrospray source are reported. The chromatographic conditions were optimized with a reversed-phase column (1 mm I.D.) using acetonitrile-5 mM ammonium acetate (pH 6.7) (50:50) as the mobile phase at a flow-rate of 50 microliters min-1. Different parameters influencing the mass spectra were investigated. For these compounds [M + H]+ in the positive-ion mode and also some fragments produced through collisionally activated decomposition in the interface were observed. Detection limits of 5.4-44 pg were obtained for standard solutions of these amines. Analysis of a meat extract was performed by HPLC-MS using single-ion monitoring after a solid-phase extraction clean-up.

Topics: Amines; Animals; Calibration; Carbolines; Cattle; Chromatography, High Pressure Liquid; Harmine; Heterocyclic Compounds; Mass Spectrometry; Meat; Mutagens

Animal experiments suggest that endogenous substances that could result from the interaction between neurotransmitters (dopamine and indoleamines) and ethanol and its metabolite acetaldehyde might be involved in the pathogenesis and maintenance of alcohol dependence. Therefore, aromatic beta-carbolines (norharman and harman) were investigated repeatedly in 24-hr urine of 13 male severe alcoholics without any psychiatric comorbidity during a controlled inpatient abstention program of up to 8 weeks. Harman excretion was approximately 2-fold above levels in control subjects, with a steady decline after 3 weeks of abstinence and lower levels in patients with a longer duration of alcohol dependence. Severity of withdrawal symptoms and actual feelings of anxiety/depression were negatively associated with urinary harman excretion. Positive associations could be established with daily ethanol consumption the month before admission and the score on the scale "reward dependence" according to Cloninger's Tridimensional Personality Questionnaire. Moreover, patients without alcohol-dependent first-degree relatives and higher "reward dependence" exhibited an increased excretion of harman. Therefore, harman levels might characterize a distinct subgroup of alcoholic patients, who in part resemble the so-called type l alcoholics of Cloninger. However, this awaits further study in a larger number of individuals. In contrast, norharman excretion was elevated up to 6-fold, compared with nonalcoholics over 6 to 8 weeks of controlled abstention. No correlations to demographic or clinical variables could be observed. Therefore, increased norharman levels might be proposed as a "residual marker" or a trait variable. Whether the observed changes are specific markers of at least certain aspects of alcoholism or dependence remain to be elucidated.

Topics: Adult; Alcohol Withdrawal Delirium; Alcoholism; Biomarkers; Carbolines; Harmine; Humans; Male; Middle Aged; Motivation; Patient Admission; Personality Inventory; Treatment Outcome

Several beta-carbolines, including harman, induce voluntary ethanol intake in rats. It is not clear yet which mechanisms cause these effects. One possibility is the stimulation of the mesolimbic reward system. In vivo microdialysis was used to investigate the effects of acute injections of harman (1-methyl-beta-carboline) on extraneuronal concentrations of dopamine and 5-hydroxytryptamine in the nucleus accumbens, which in part the mesolimbic reward system. Administration of harman (2.27 mumol/kg, intraperitoneal application) elicited an increase of the dopamine efflux by 72% which returned to basal levels after approximately 300 min. In contrast, administration of an intermediate dose of harman (13.65 mumol/kg, intraperitoneal application) caused a significant decrease in efflux, to 76% of basal levels. Still higher doses included again an increased extracellular dopamine concentration. This change was statistically significant in only a subgroup rats, possibly because individual animals reacted differently to the high doses. Extracellular 5-hydroxytryptamine in the nucleus accumbens was increased during the first 2 h after the administration of high doses (40.94 and 81.93 mumol/kg, intraperitoneal application). These findings indicate that harman affects the activity of mesolimbic dopaminergic neurons following a U-shaped dose-response relationship.

Topics: Animals; Carbolines; Dopamine; Harmine; Male; Microdialysis; Neurotoxins; Nucleus Accumbens; Rats; Rats, Wistar; Serotonin

Both physical rehabilitation and the course of the alcoholism improve after orthotopic liver transplantation (OLT) in patients with end-stage alcoholic liver cirrhosis. In the present study including 17 alcoholics and 14 nonalcoholics, after OLT, three of the alcoholic patients resumed their pre-OLT alcohol drinking habits, 4 consumed alcohol occasionally, 10 remained abstinent over the observation period of 13 to 36 months. The laboratory parameters before OLT did not discriminate alcoholics from nonalcoholic patients. Furthermore, the blood levels of two so-called alcogens (harman and norharman) were determined to investigate whether they discriminate between the two groups. Alcogens are natural compounds that are presumed to induce alcohol abuse in predisposed individuals. Both alcogens measured were elevated in plasma from nonalcoholics and alcoholics before OLT, suggesting a disturbance in inactivation in end-stage liver disease. Following OLT, the alcogens normalized but in the alcoholics this process was slower with respect to harman. The present exploratory study suggests that the normalized metabolic capacity of the liver after OLT causes a normalization of the levels of alcogens, for which harman and norharman are representative. These changes could contribute to the observed benefit to the outcome in alcoholics with respect to the alcohol dependence.

Topics: Adult; Alcoholism; Analysis of Variance; Carbolines; Harmine; Humans; Liver Diseases, Alcoholic; Liver Function Tests; Liver Transplantation; Male; Middle Aged; Neurotoxins; Time Factors

Death of dopaminergic neurons in Parkinson's disease (PD) may partially be caused by synthesis and accumulation of endogenous and exogenous toxins. Because of structural similarity to MPTP, beta-carbolines, like norharman and harman, have been proposed as putative neurotoxins. In vivo they may easily be formed by cyclization of indoleamines with e.g. aldehydes. For further elucidation of the role of beta-carbolines in neurodegenerative disorders harman and norharman levels in cerebrospinal fluid (CSF) were measured in 14 patients with PD and compared to an age- and sex-matched control group (n = 14). CSF levels of norharman and harman in PD were significantly higher compared to controls. These results may suggest a possible role of harman and norharman or its N-methylated carbolinium ions in the pathophysiological processes initiating PD. However the origin of increased levels of these beta-carbolines remains unclear. On the one hand one may speculate, that unknown metabolic processes induce the increased synthesis of harman and norharman in PD. On the other hand a possible impact of exogenous sources may also be possible.

Topics: Adult; Aged; Carbolines; Case-Control Studies; Female; Harmine; Humans; Male; Middle Aged; Neurotoxins; Parkinson Disease

The prevalence of chronic alcoholism in patients with carcinomas of the upper digestive tract exceeds 60%. The patient's history and laboratory markers, preoperatively, are often not sensitive or specific enough to detect alcohol-dependent patients, preoperatively, who are at risk of developing alcohol withdrawal syndrome (AWS) during their postoperative intensive care unit (ICU) stay. Previously, it was found that plasma norharman was elevated in chronic alcoholics, suggesting marker characteristics for chronic ethanol misuse and possibly alcohol dependence. We investigated whether beta-carbolines (i.e., harman and norharman) were different between chronic alcoholics and nonalcoholics with carcinoma, and how the levels change in alcohol-dependent patients during their hospital stay. Ninety-seven patients with oral, pharyngeal, laryngeal, or esophageal carcinomas were evaluated regarding their drinking habits. Sixty patients were transferred to the ICU following tumor resection. Chronic alcoholics met the DSM-III-R and ICD-10 criteria for alcohol dependence or chronic alcohol abuse/harmful use. The daily ethanol intake in chronic alcoholics was > or = 60 g. Blood samples were collected on admission to the hospital, preoperatively, on admission to the ICU and on days 2, 4, and 7 in the ICU. Harman and norharman were determined by HPLC. Elevated norharman was found in chronic alcoholics on admission to the hospital, whereas harman did not differ between groups. On admission, the area under the receiver operating characteristics curve was significantly larger for carbohydrate-deficient transferrin and preoperatively for norharman. The preoperative norharman levels were significantly correlated with the period of mechanical ventilation and the length of ICU stay. Postoperatively, norharman decreased in all patients, except a group of 11 alcohol-dependent patients who developed AWS during their ICU stay. The finding that elevated norharman levels were found in chronic alcoholics on admission to the hospital and preoperatively supports the view of a specific marker for alcoholism. Preoperative norharman was superior to carbohydrate-deficient transferrin and was associated with a prolonged ICU stay and a prolonged period of mechanical ventilation. Further studies are required to determine whether norharman aids in the preoperative diagnosis of chronic alcohol misuse with respect to the prevention of postoperative complications.

Topics: Adult; Aged; Aged, 80 and over; Alcoholism; Biomarkers; Carbolines; Critical Care; Esophageal Neoplasms; Harmine; Humans; Male; Middle Aged; Otorhinolaryngologic Neoplasms; Postoperative Complications; Risk Factors

Several lines of evidence suggest that endogenous and exogenous toxins may play a major role in the pathogenesis of Parkinson's disease (PD). In the brain aromatic beta-carbolines, like harman or norharman, may be formed by cyclization of indoleamines. Because of the structural similarity to MPTP, beta-carbolines have been proposed as endogenous toxins. For further elucidation of the role of beta-carbolines in neurodegenerative disorders, harman and norharman plasma levels were measured in 36 patients with PD and compared to an age- and sex-matched control group. Plasma levels of norharman in PD were significantly higher compared to the control group. Harman in the plasma of Parkinsonian patients was also elevated compared to the controls, but this difference was not significant. On the one hand these results may suggest a possible role of beta-carbolines in the pathophysiological processes initiating PD. But on the other hand one may speculate that elevated levels of norharman and harman are due to an endogenous upregulation, caused by unknown metabolic processes to reduce oxidative stress by inhibiting e.g. monoaminooxidases in neurons.

Topics: Aged; Aged, 80 and over; Carbolines; Case-Control Studies; Female; Harmine; Humans; Male; Middle Aged; Neurotoxins; Parkinson Disease

Harman (1-methyl-beta-carboline) displaces [3H]pargyline in vitro from high affinity binding sites on membranes from cerebral cortex, provided that experimental conditions are chosen under which [3H]pargyline labels selectively monoamine oxidase type A. Norharman (beta-carboline) is a much weaker displacing compound. It is well known that the type A enzyme can be blocked irreversibly in vivo by treatment of rats with clorgyline. Under these conditions no specific binding of [3H]harman and [3H]pargyline to monoamine oxidase type A was detected in brain, whereas the specific binding was reduced to 5% in liver tissue. The in vitro and ex vivo experiments suggest that there is a specific binding site for harman on monoamine oxidase type A, thereby extending earlier in vitro findings. It has been postulated that harman operates as a natural inhibitor of monoamine oxidase type A in mammals. The present study demonstrates that harman and norharman occur in rat brain, blood plasma, heart, kidney and liver. It further shows that pretreatment with clorgyline induces a time-dependent increase in the blood plasma levels of harman, suggesting the displacement of harman from the enzyme in tissue with its subsequent delivery into the blood. These findings strongly support the hypothesis based on in vitro experiments, that harman binds reversibly to the active site of monoamine oxidase type A in vivo. Dietary sources for mammalian harman play probably only a minor role, because the concentrations in beer and wine as well as other foodstuffs are too low to contribute substantially to endogenous levels of harman.

Topics: Animals; Beer; Binding Sites; Brain; Carbolines; Harmine; Kidney; Liver; Male; Monoamine Oxidase; Monoamine Oxidase Inhibitors; Myocardium; Pargyline; Rats; Rats, Wistar; Tissue Distribution; Wine

The in vitro binding of the naturally occurring beta-carbolines harman and norharman in their tritium-labelled forms to cell membranes from the rat brain and liver and from bovine adrenal medulla was investigated. Displacement of the specific [3H]harman binding in bovine adrenal medulla and rat liver by several beta-carbolines and monoamine oxidase (MAO) inhibitors revealed the pharmacological profile of a single, high-affinity binding site (KD 4.92 +/- 0.43 nmol/l, Bmax 8.47 +/- 0.17 pmol/mg protein; adrenal medulla) which corresponded to the active site of MAO type A (MAO-A). Similar characteristics have previously been found for brain tissue from rat, marmoset and pig. In order to determine the temperature dependence of the [3H]harman binding, the KD and Bmax values for rat cerebral cortex were calculated from the results of saturation experiments at 5 temperatures (range: 0 degree C-37 degrees C). Whereas the Bmax values under all conditions were approximately 4 pmol/mg protein, the KD values, with increasing temperature, ranged from approximately 3 nmol/l to 30 nmol/l. The calculated linear van't Hoff plot (-ln KD against 1/T) suggested an enthalpy-driven binding of [3H]harman to MAO-A. At least three different [3H]norharman-binding sites were detected. In the rat forebrain, approximately 85% of the specific binding (at about 2 nmol/l of [3H]norharman) can be attributed to a MAO binding site of type B: the binding is displaceable, in nmol/l concentrations by the potent and selective MAO-B inhibitors MDL 72,974 A, R(-)-deprenyl and pargyline and, in mumol/l concentrations, by S(+)-deprenyl and the potent and selective MAO-A inhibitors clorgyline, harmine, harman, harmaline, brofaromine 5-F-alpha-methyltryptamine. After suppression of the MAO binding sites with 1 mumol/l clorgyline and 1 mumol/l R(-)-deprenyl, a second binding site was found. However, the binding at this site was biphasically displaceable by harman and norharman (Hill-slopes about 0.5 and 0.6, curvilinear Rosenthal plots) suggesting the presence of negative co-operativity or of two binding sites (states). A similar clorgyline/R(-)-deprenyl resistant single (Hill-slopes of displacement by norharman, harman and 6-hydroxy-beta-carboline about unity; linear Rosenthal plots) high affinity binding sites (KD 7.5 +/- 2 nmol/l, Bmax 130+/- 30 fmol/mg protein) was found in bovine adrenal medullary cell membranes. A third quite different clorgyline/R(-)-deprenyl resistant high-affinity (KD approx

Topics: Adrenal Medulla; Animals; Brain; Carbolines; Cattle; Harmine; In Vitro Techniques; Liver; Male; Rats; Rats, Wistar; Subcellular Fractions; Temperature

Norharman (beta-carboline, a so-called mammalian alkaloid) is identified as a high-affinity type II ligand for two steroidogenic cytochromes P450, viz. CYP11 in rat adrenal mitochondria and CYP17 in rat testicular microsomes. Progesterone binding to CYP17 is competitively inhibited, with Ki = 2.6 microM norharman, whereas harman, tetrahydronorharman and tetrahydroharman are nearly ineffective. The potential role of norharman as an endogenous modulator of steroid hormone biosynthesis and as a basic drug for development of more specific cytochrome P450 inhibitors is emphasized.

Topics: Adrenal Glands; Animals; Binding, Competitive; Carbolines; Harmine; Male; Microsomes; Mitochondria; Progesterone; Rats; Spectrophotometry, Ultraviolet; Steroid 11-beta-Hydroxylase; Steroid 17-alpha-Hydroxylase; Testis

Endogenous substances resulting from interactions between alcohol and possibly opioid metabolites and neurotransmitters (dopamine, indolamines) are mediators of the pathochemical process towards dependence. Beta-carbolines are increased in alcoholics and--according to our own results--in heroin-addicts. Still unclear is the impact of other psychopathological disturbances like states of anxiety or depression; unclear is also, if it has to be interpreted as state, trait or residual marker of the dependence syndrome.

Topics: Adult; Carbolines; Female; Harmine; Heroin Dependence; Humans; Male; Methadone; Substance Withdrawal Syndrome

Harman and norharman are two beta-carboline derivatives known to be present in certain foods and are formed during pyrolysis of amino-acids. Their effects on the metabolism of aflatoxin B1 and aminopyrine by 3-methylcholanthrene and phenobarbital-induced rat liver microsomes were studied. Both harman and norharman markedly inhibited the metabolism of aflatoxin B1 to its hydroxylated derivative, aflatoxin M1. However, only norharman showed an inhibitory effect on aminopyrine N-demethylation; harman had no effect. Harman and norharman inhibited aflatoxin B1 binding to DNA, mediated by hepatic microsomes in vitro.

Topics: Aflatoxin B1; Aminopyrine N-Demethylase; Animals; Carbolines; Harmine; Male; Methylcholanthrene; Microsomes, Liver; Phenobarbital; Rats; Rats, Wistar

The modifying potential of two nephrotoxic agents, harman and norharman, on N-ethyl-N-hydroxyethylnitrosamine (EHEN)-induced renal and hepatic carcinogenesis was investigated in male F344/DuCrj rats. Animals were given 0.1% EHEN in their drinking water for the first 2 weeks as an initiator. Subsequently, starting 3 weeks from the commencement, they were fed diet containing these compounds at concentrations of 1000, 500 or 0 ppm until week 26, and then killed for light microscopic examination. The mean numbers of renal tubular cell hyperplasias/cm2 and those of tumors/cm2 in rats given harman and norharman at 1000 ppm after initiation, but not at 500 ppm, were significantly increased as compared to the control values. However, neither compound modified liver carcinogenesis. It is concluded that harman and norharman show enhancing effects on rat kidney carcinogenesis, when ingested at dose levels which cause renal tubular damage.

Topics: Animals; Carbolines; Carcinogens; Carcinoma, Renal Cell; Diethylnitrosamine; Harmine; Kidney Neoplasms; Male; Precancerous Conditions; Rats; Rats, Inbred F344

Fresh, mature, ungrazed Tribulus terrestris plant material was subjected to a standard alkaloid extraction procedure. The extract was fractionated by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC). Two major alkaloid fractions were demonstrated. These fractions were identified by means of TLC, ultraviolet spectrofluorimetry (UVS) and HPLC, as the beta-carboline indoleamines harmane and norharmane. The extractable alkaloid content was determined to be 44 mg/kg dry matter. Synthetic harmane and norharmane were administered subcutaneously to sheep at a dose rate of 54 mg/kg. Both compounds caused similar nervous effects. The main effect observed was limb paresis, which in some sheep was body side blased. The clinical signs observed in the experimental sheep were consistent with those described for naturally occurring cases of Tribulus terrestris staggers. It was proposed that harmane and norharmane accumulate in tryptamine-associated neurones of the central nervous system, during months of tribulus ingestion, and gradually interact irreversibly with a specific neuronal gene DNA sequence.

Topics: Animals; Carbolines; Chromatography, High Pressure Liquid; Chromatography, Thin Layer; Female; Harmine; Locomotion; Plant Extracts; Plant Poisoning; Plants, Edible; Sheep; Sheep Diseases; Spectrophotometry, Ultraviolet

The renal toxicity of harman and norharman, administered for 2 or 4 weeks at dietary levels of 1,000, 500, or 0 parts per million (ppm), was investigated in 6-week-old male F344/DuCrj rats. Although rats fed 1,000 ppm harman or norharman, but not the 500 ppm level, demonstrated marked body weight retardation from 1 week to termination, no mortalities occurred. Marked elevation of water consumption was evident in rats given harman or norharman at 1,000 ppm, but not at 500 ppm, together with large increases in urine of low specific gravity. Urinary lysosomal enzymes (N-acetyl-beta-D-glucosaminidase, NAG, and lactate dehydrogenase, LDH) and sugar levels were increased, and the brush border enzymes (gamma-glutamyl transpeptidase, GGT, and alkaline phosphatase, ALP) decreased. Furthermore, serum biochemistry revealed clear elevation of parameters indicating renal toxicity in these rats. Histopathologically, rats fed 1,000 ppm harman or norharman, but not 500 ppm, demonstrated focal toxic renal degenerative/necrotic and regenerative lesions in proximal, distal, and collecting tubules. These changes were associated with a clearly increased labeling index (LI) of the nuclei of renal tubular epithelial cells on immunohistochemical staining for 5-bromo-2'-deoxyuridine (BrdU). Chemical specific crystal formation within tubular lumina was evident in rats fed 1,000 ppm, but not 500 ppm, this being considered the cause of the renal tubular lesions. It was concluded that harman and norharman exert renal toxicity at the dietary level of 1,000 ppm, but not 500 ppm, in male F344 rats.

Topics: Acetylglucosamine; Animals; Bromodeoxyuridine; Carbolines; DNA; Drinking; Harmine; Kidney Diseases; Kidney Tubules; L-Lactate Dehydrogenase; Male; Microvilli; Organ Size; Phagocytosis; Rats; Rats, Inbred F344; Urination

Based on the hypothesis that condensation products of neurotransmitters with aldehydes are involved in the pathogenesis of alcoholism, aromatic beta-carbolines (norharman and harman) were measured in the blood plasma of alcoholics and nonalcoholics. The identity of the extracted compounds was confirmed by various elution conditions of the high performance liquid chromatography (HPLC), newly developed radioreceptor assays, and the mass spectrum of norharman. The levels of norharman and harman in nonalcoholics were unchanged after a load with ethanol (1 g/kg body weight). The norharman levels of the alcoholics were significantly higher than that of the nonalcoholic controls (99.5 +/- 26.6 pg/ml vs. 26.9 +/- 10.7 pg/ml; p less than 0.001) and did not change significantly during a 3-week detoxication period. In the subgroup of alcoholics with delirium or hallucinosis, a slight increase of norharman during detoxication could be detected while in alcoholics with vegetative withdrawal symptoms norharman levels dropped slightly over time (p = 0.07). No difference was found with respect to harman between nonalcoholics and alcoholics. These results suggest disturbed regulatory processes in the formation and/or metabolism of norharman in alcoholics. Further investigations are needed to reveal a possible marker function of norharman in alcoholic patients.

Topics: Adolescent; Adult; Alcoholism; Carbolines; Chromatography, High Pressure Liquid; Ethanol; Female; Follow-Up Studies; Harmine; Humans; Male; Middle Aged; Psychiatric Status Rating Scales; Psychopathology; Substance Withdrawal Syndrome

A high-performance liquid chromatographic method combined with fluorimetric detection is described for the determination of beta-carboline (norharman) and 1-methyl-beta-carboline (harman). The analysis of foodstuffs for the identification of beta-carbolines is facilitated by clean-up samples using Bond Elut PRS cartridges. Recoveries were excellent. Further, a high-performance liquid chromatographic-mass spectrometric method was also developed for their identification. The concentration of beta-carboline among the foodstuffs and alcoholic beverages varied greatly. Also, norharman and harman were observed in uncooked foodstuffs, whereas acetaldehyde was found in most fermented food. The toxicological implication of beta-carbolines in foodstuffs is discussed.

Topics: Acetaldehyde; Alcoholic Beverages; Carbolines; Chromatography, High Pressure Liquid; Ethanol; Food Analysis; Harmine; Ions; Mass Spectrometry

The modifying effects of harman or norharman on liver carcinogenesis were investigated in male F344/DuCrj rats initially treated with N-nitrosodiethylamine (DEN). Two weeks after a single dose of DEN (200 mg/kg, intraperitoneally), rats were given harman or norharman at dietary levels of 1000 and 200 parts per million (ppm), or sodium phenobarbital (PB) at 500 ppm as a positive control, for 6 wk. At wk 3 following DEN administration, all animals were subjected to partial hepatectomy. Marked retardation of body weight gain was observed in rats treated with harman or norharman at 1000 ppm, but not at 200 ppm. Increased relative kidney but not liver weights were associated with harman or norharman treatment, especially in the higher dose groups. Although no toxicity-related hepatocyte lesions were found, severe renal toxic tubular lesions and regeneration were evident. Neither harman nor norharman significantly increased the numbers or areas of glutathione S-transferase placental form (GST-P) positive foci observed after DEN initiation, in clear contrast to PB. The results thus demonstrated that harman and norharman are nontoxic for the liver and lack modifying potential for liver carcinogenesis in our medium-term bioassay system.

Topics: Alkaloids; Animals; Body Weight; Carbolines; Cocarcinogenesis; Diethylnitrosamine; Glutathione Transferase; Harmine; Liver; Male; Rats; Rats, Inbred F344

The beta-carbolines harmane, norharmane, tetrahydronorharmane, harmine, harmaline and harmol were administered to sheep to assess their effects on upper motor neurone function. Harmane at a dose rate of 54 mg/kg induced hypomotility, head tremors, pelvic limb paresis, hypermetria and a wide based stance. A range of similar effects were observed with norharmane at the same dose rate. Tetrahydronorharmane at a dose rate of 54 mg/kg induced hypermotility followed by hypomotility, asymmetrical pelvic limb paresis, hypermetria, a wide based stance, and stereotyped eating behaviour. Harmine and harmaline at 6 mg/kg induced mild head and body tremors, and at 18 mg/kg induced hypomotility, intense head and body tremors, pelvic limb paresis, crossing over of limbs, neck extension and head swaying. Harmol was not effective at 54 mg/kg by either the subcutaneous or intraperitoneal routes, but at an intravenous dose of 27 mg/kg it induced hypermotility followed by hypomotility, body tremors, limb paresis, muscle asynergy, a wide based stance and jumping behaviour. Harmane, tetrahydronorharmane, harmaline and harmol were convulsive in some sheep at high dose rates.

Topics: Alkaloids; Animals; Carbolines; Female; Harmaline; Harmine; Locomotion; Motor Neurons; Plants, Toxic; Sheep

Erythrosine (diNa, tetraiodofluorescein) was nonmutagenic to the Ames/Salmonella typhimurium strains TA97a, TA98, TA100, TA102, and TA104, to a concentration of 2 mg/plate. No mutative intermediates were detected on metabolism by rat caecal cell-free extracts or rat liver S9 mixture; or on incubation with the comutagens, harman and norharman (+/- S9). Instead, an unexpected dose-dependent suppression in spontaneous reversion frequencies was observed (maximum approximately equal to 35% decrease). Erythrosine was antimutagenic to benzo[a]pyrene, but it did not decrease the mutagenicity of the other adduct-forming mutagen, 4-nitroquinoline N-oxide. The food dye was strongly antimutagenic to the bifunctional alkylating agent, mitomycin C, though it did not exhibit a similar effect on the mutagenicity of the corresponding monofunctional agent, methyl methanesulphonate. It partially depressed the mutagenic potentials of sodium azide. The antimutagenic effect of erythrosine on an intercalating agent, ethidium bromide, was discernible only at the highest dose (2 mg/plate). These results have been interpreted in terms of a genointeractive role of erythrosine. Erythrosine produced differential toxic effects in repair-deficient (TA97a, TA98, TA100) and repair-proficient (TA102, TA104) Salmonella tester strains; survival of the repair-deficient strains was found to be decreased. Photoinduced potentiation of erythrosine toxicity was observed, although light irradiation in the presence of erythrosine did not modify the reversion frequencies of the tester strains. The evidence strongly suggests that erythrosine, which exhibits nonmutagenicity in the Ames/Salmonella test, can interact with DNA repair enzymes and/or with DNA.

Topics: 4-Nitroquinoline-1-oxide; Animals; Azides; Benzo(a)pyrene; Carbolines; Cecum; DNA Damage; DNA Repair; Drug Interactions; Erythrosine; Ethidium; Fluoresceins; Food Coloring Agents; Harmine; Male; Methyl Methanesulfonate; Microsomes, Liver; Mitomycin; Mitomycins; Mutation; Rats; Rats, Inbred Strains; Salmonella typhimurium; Sodium Azide

Covalent modifications of DNA in various tissues of mice with harman or norharman were analyzed by 32P-postlabeling assay. Administration of 0.1% harman to mice in their diet for 4 weeks resulted in DNA adducts in the liver and kidney. No specific DNA adduct was detected in other tissues, such as the glandular stomach, large intestine and brain. Similar treatment of mice with norharman resulted in DNA adducts in the kidney, glandular stomach and large intestine, but not in the liver or brain. These results suggests the in vivo genotoxicities of harman and norharman.

Topics: Alkaloids; Animals; Carbolines; DNA; DNA Damage; Gastric Mucosa; Harmine; Intestine, Large; Kidney; Liver; Mice; Tissue Distribution

A method using high-performance liquid chromatography with fluorescence detection was developed for the determination of beta-carboline compounds norharman, harman, norharmol, and harmol in lung. Aqueous derivatization with acetic anhydride was used to facilitate the isolation and separation of the phenolic compounds and to reduce the fluorescence background of the biological samples. Harman was identified and quantitated in rat lung (1.88 +/- 0.55 ng/g) using this method and its identity confirmed by means of gas chromatography-negative-ion chemical ionization mass spectrometry.

Topics: Animals; Carbolines; Chromatography, High Pressure Liquid; Fluorobenzenes; Gas Chromatography-Mass Spectrometry; Harmine; Male; Rats; Rats, Inbred Strains; Spectrometry, Fluorescence

Norharman and harman, beta-carboline derivatives with comutagenic activity in Salmonella typhimurium, were examined for their activity to induce SOS responses in S. typhimurium using the umu-test and mutations in Escherichia coli. The inducibility of the umuC gene by norharman and harman was assayed by measuring the levels of beta-galactosidase activity in tester cells harbouring the umuC'-'lacZ fusion gene on a plasmid. In the umu-test, both norharman and harman weakly induced umuC gene expression at 25-100 and 50-150 micrograms/ml, respectively. In the mutation test using reversion from trpE9777 to Trp+, harman was relatively more potent than norharman in inducing the mutations. These results indicate that norharman and harman induce SOS responses as well as reversion of trpE9777 frameshift mutation in bacteria.

Topics: Alkaloids; Carbolines; DNA Repair; Harmine; Mutagenicity Tests; Mutagens; Mutation; Salmonella typhimurium; SOS Response, Genetics

Monolayers of rat hepatocytes metabolize 0.25 mM 2-acetylaminofluorene (AAF) to various ether-extractable, water-soluble as well as covalently bound products. The major ether-extractable metabolite formed is 2-aminofluorene (AF), followed by 7-OH-AAF and 9-OH-AAF. Pretreatment of rats with the inducer Aroclor 1254 (PCB) increased the metabolism of AAF and caused an increased DNA repair synthesis in hepatocytes exposed to AAF or AF. With N-OH-AAF, a decreased genotoxic response in PCB-treated cells compared to control cells was seen. The addition of harman and norharman decreased the metabolism of AAF to ether-extractable metabolites, water-soluble metabolites and metabolites covalently bound to macromolecules. In contrast, the DNA-repair synthesis caused by the same concentrations of AAF was increased by harman. One explanation for this apparent discrepancy could be that the aromatic amines changed the metabolism of harman and norharman in such a way that these compounds were converted into genotoxic metabolites.

Topics: 2-Acetylaminofluorene; Alkaloids; Animals; Carbolines; Cells, Cultured; DNA; DNA Repair; Drug Synergism; Harmine; Liver; Male; Mutagens; Rats; Rats, Inbred Strains

Synthetic 3-aminoharman and 3-aminonorharman (amino-beta-carbolines) caused slight but definite induction of sister-chromatid exchanges (SCEs) in human lymphoblastoid cells NL3 and Chinese hamster cells CHO-K1. These amino-beta-carbolines are ranked between 2-amino-alpha-carboline and 2-amino-6-methyl-9a-aza-delta-carboline (Glu-P-2) and much lower than 3-amino-gamma-carbolines (Trp-P-1 and 2) in inductive activity. 1-Amino-beta-carboline, harman and norharman had very weak, if any, SCE-inducer activity. Norharman had a synergistic effect with aromatic amines such as Trp-P-2 and aniline on SCE induction, while 3-aminoharman suppressed SCE induction by more potent inducers such as Trp-P-2 and benzo[a]pyrene.

Topics: Animals; Carbolines; Cell Line; Cricetinae; Cricetulus; Crossing Over, Genetic; Female; Harmine; Indoles; Ovary; Sister Chromatid Exchange; Structure-Activity Relationship

Topics: Alkaloids; Animals; Biotransformation; Carbolines; Cell Line; Cell Survival; Cricetinae; Cricetulus; Diphtheria Toxin; Drug Resistance; Female; Harmine; Microsomes, Liver; Mutagenicity Tests; Mutagens; Mutation; Ovary

Acetylcholinesterase (acetylcholine acetylhydrolase, EC 3.1.1.7) activity and muscarinic receptor binding of homogenates from several brain structures were inhibited by beta-carbolines. The inhibition was of the noncompetitive type in the case of the enzyme and of the mixed type in the case of the receptor binding. This effect was most strongly manifested by pyridoindoles(harmane, norharmane), i.e., carbolines containing an aromatic C ring than by the corresponding piperidoindoles (tetrahydroharmane, tetrahydronorharmane), i.e., those with a reduced C ring. The activity of choline acetyltransferase (acetyl-CoA:choline O-acetyltransferase, EC 2.3.1.6) was not altered. These data are further evidence of the interactions between indoleamine derivatives and the cholinergic system. The results are discussed in terms of their possible biological significance.

Topics: Acetylcholinesterase; Alkaloids; Animals; Brain; Carbolines; Cattle; Choline O-Acetyltransferase; Erythrocytes; Harmaline; Harmine; Indoles; Kinetics; Male; Rats; Rats, Inbred Strains; Receptors, Cholinergic; Receptors, Muscarinic; Structure-Activity Relationship

The enhancing or decreasing effect of harman or norharman on benzo[a]pyrene mutagenesis depends mainly on the metabolizing system (S9) used. When mammalian activation was performed by using liver homogenates from mice induced by 3-methylcholanthrene, phenobarbital or Aroclor 1254, then the mutagenicity of benzo[a]pyrene in Salmonella typhimurium TA98 decreased upon addition of harman or norharman. In the presence of rat-liver homogenate induced by Aroclor 1154, however, harman enhanced the number of benzo[a]pyrene revertants, whereas norharman did not show any significant alteration of the benzo[a]pyrene mutagenicity. Further tests carried out with phenobarbital-induced mouse-liver homogenate showed that, within certain limits, neither variations of the amount of S9 extract nor the amount of the cofactors nor the relative proportions of the test substances had any influence on the antagonistic effect of harman or norharman on the benzo[a]pyrene mutagenesis.

Topics: Alkaloids; Animals; Benzopyrenes; Biotransformation; Carbolines; Dose-Response Relationship, Drug; Harmine; In Vitro Techniques; Liver; Mice; Mutagenicity Tests; Mutagens; Rats; Salmonella typhimurium

Topics: Alkaloids; Aniline Compounds; Aniline Hydroxylase; Animals; Carbolines; Harmine; Male; Microsomes, Liver; p-Dimethylaminoazobenzene; Rats

The beta-carbolines harmane and norharmane competitively inhibit [3H]flunitrazepam ([3H]FNZ) binding to deoxycholate-solubilized benzodiazepine receptors from calf cerebral cortex, with Ki in the micromolar range [3H]Propyl-beta-carboline-3-carboxylate ([3H]PrCC) binds to the soluble receptors with an affinity similar to its binding to particulate receptors (0.41 nM vs 0.48 nM, respectively). The component that binds [3H]PrCC displays a sedimentation profile on sucrose gradient centrifugation similar to that of [3H]FNZ binding component (sedimentation coefficient about 11S).

Topics: Alkaloids; Animals; Benzodiazepines; Carbolines; Cattle; Cerebral Cortex; Deoxycholic Acid; Flunitrazepam; Harmine; Receptors, Drug; Solubility; Statistics as Topic; Tritium

Harman, but not norharman, induced sister-chromatid exchanges in human peripheral lymphocytes in vitro. Transcription of isolated DNA in vitro was inhibited by both compounds, harman being more effective than norharman. A filter-binding assay with isolated DNA showed that harman binds more effectively to the DNA than norharman.

Topics: Alkaloids; Carbolines; Chromosome Aberrations; Crossing Over, Genetic; DNA; Harmine; Humans; Lymphocytes; Sister Chromatid Exchange; Structure-Activity Relationship; Transcription, Genetic; Triaziquone

Norharman enhanced the known mutagenicity of 2-acetylaminofluorene derivatives in the Salmonella test system. The mutagenicities of 2-acetylaminofluorene, 2-aminofluorene, and N-hydroxy-2-acetylaminofluorene were enhanced by norharman only when rat liver microsomal enzymes were added, whereas the mutagenicity of N-acetoxy-2-acetylaminofluorene was increased in the absence of microsomal enzymes. Harman also increased mutagenesis, althouth less so than norharman.

Topics: 2-Acetylaminofluorene; Alkaloids; Carbolines; Drug Synergism; Fluorenes; Harmine; Microsomes, Liver; Mutagens; Salmonella typhimurium

Nontoxic concentrations of harman and norharman were tested in cultured Chinese hamster cells for their effects on DNA repair and mutagenesis. The following effects of harman were observed: (a) the survival of ultraviolet light- or X-ray-damaged cells was reduced; (b) the ultraviolet light-induced unscheduled DNA synthesis was slightly inhibited; and (c) the frequency of spontaneous or ultraviolet light-induced ouabain-resistant (ouar) or 6-thioguanine-resistant (6-TGr) mutations was reduced. Furthermore, the effect of harman on survival and mutagenesis was greater than that of norharman and was detected primarily in treatments in which cells were exposed to harman immediately following ultraviolet light irradiation. Our data clearly indicate that harman decreases the capacity to repair DNA damage and fix mutations in Chinese hamster cells, possibly because of the intercalation properties of this compound.

Topics: Alkaloids; Carbolines; Cell Survival; Cells, Cultured; DNA Repair; Harmine; Mutation; Tryptophan; Ultraviolet Rays

The interactions of norharman (9H-pyrido [3,4-b] indole) and harman (1-methyl-9H-pyrido [3,4-b] indole) with DNA were studied. DNA caused remarkable fluorescence quenching and change in the absorption spectra of the dyes. Scatchard plots obtained by optical titration gave Kd values of 2.2 X 10(-5)M and 7.7 X 10(-6)M, and apparent numbers of binding sites of 0.13/base and 0.12/base for norharman and harman, respectively. Agarose gel electrophoresis of circular DNA, closed in the presence or absence of norharman revealed that the dye intercalates DNA, thereby causing 17 +/- 3 degrees unwinding of the double helix.

Topics: Alkaloids; Binding Sites; Carbolines; Coliphages; DNA; DNA, Circular; DNA, Superhelical; DNA, Viral; Harmine; Nucleic Acid Conformation; Spectrometry, Fluorescence; Spectrum Analysis

Topics: Alkaloids; Aryl Hydrocarbon Hydroxylases; Benzopyrenes; Carbolines; Cells, Cultured; DNA; Enzyme Induction; Harmine; Liver; Mutagens