ABSTRACT
ß-Carbolines (ßCs) are bioactive indole alkaloids found in foods and in vivo. This work describes the identification, formation, and occurrence in foods of the ßC with a furan moiety flazin (1-[5-(hydroxymethyl)furan-2-yl]-9H-pyrido[3,4-b]indole-3-carboxylic acid). Flazin was formed by the reaction of l-tryptophan with 3-deoxyglucosone but not with 5-hydroxymethylfurfural. Its formation was favored in acidic conditions and heating (70-110 °C). The proposed mechanism of formation occurs through the formation of intermediates 3,4-dihydro-ß-carboline-3-carboxylic acid (imines), followed by the oxidation to CâO in the carbohydrate chain and aromatization to ßC ring with subsequent dehydration steps and cyclization to afford the furan moiety. Flazin is generated in the reactions of tryptophan with carbohydrates. Its formation from fructose was higher than from glucose, whereas sucrose gave flazin under acidic conditions and heating owing to hydrolysis. Flazin was identified in foods by HPLC-MS, and its content was determined by HPLC-fluorescence. It occurred in numerous processed foods, such as tomato products, including crushed tomato puree, fried tomato, ketchup, tomato juices, and jams, but also in soy sauce, beer, balsamic vinegar, fruit juices, dried fruits, fried onions, and honey. Their concentrations ranged from not detected to 22.3 µg/mL, with the highest mean levels found in tomato concentrate (13.9 µg/g) and soy sauce (9.4 µg/mL). Flazin was formed during the heating process, as shown in fresh tomato juice and crushed tomatoes. These results indicate that flazin is widely present in foods and is daily uptaken in the diet.
Subject(s)
Carbolines , Tryptophan , Carbohydrates , FuransABSTRACT
The AT-rich mitochondrial DNA (kDNA) of trypanosomatid parasites is a target of DNA minor groove binders. We report the synthesis, antiprotozoal screening, and SAR studies of three series of analogues of the known antiprotozoal kDNA binder 2-((4-(4-((4,5-dihydro-1H-imidazol-3-ium-2-yl)amino)benzamido)phenyl)amino)-4,5-dihydro-1H-imidazol-3-ium (1a). Bis(2-aminoimidazolines) (1) and bis(2-aminobenzimidazoles) (2) showed micromolar range activity against Trypanosoma brucei, whereas bisarylimidamides (3) were submicromolar inhibitors of T. brucei, Trypanosoma cruzi, and Leishmania donovani. None of the compounds showed relevant activity against the urogenital, nonkinetoplastid parasite Trichomonas vaginalis. We show that series 1 and 3 bind strongly and selectively to the minor groove of AT DNA, whereas series 2 also binds by intercalation. The measured pKa indicated different ionization states at pH 7.4, which correlated with the DNA binding affinities (ΔTm) for series 2 and 3. Compound 3a, which was active and selective against the three parasites and displayed adequate metabolic stability, is a fine candidate for in vivo studies.
Subject(s)
Antiprotozoal Agents , Benzamides , Leishmania donovani , Parasites , Trypanosoma brucei brucei , Trypanosoma cruzi , Animals , Antiprotozoal Agents/chemistry , DNA/metabolism , DNA, Kinetoplast/metabolism , Imidazoles/chemistry , Imidazoles/pharmacology , Leishmania donovani/metabolism , Parasites/drug effects , Parasites/metabolism , Benzamides/chemistry , Benzamides/pharmacologyABSTRACT
ß-Carbolines are naturally occurring bioactive alkaloids found in foods and in vivo. This research reports the identification, characterization, mechanism of formation, and occurrence of perlolyrine (1-(5-(hydroxymethyl)furan-2-yl)-9H-pyrido[3,4-b]indole), a ß-carboline with a furan moiety. Perlolyrine did not arise from l-tryptophan and hydroxymethylfurfural but from the reaction of l-tryptophan with 3-deoxyglucosone, an intermediate of carbohydrate degradation. The mechanism of formation occurs through 3,4-dihydro-ß-carboline-3-carboxylic acid intermediates (imines), followed by the oxidation of C1'-OH to ketoimine and oxidative decarboxylation at C-3, along with dehydration and cyclization to afford the ß-carboline with a furan moiety. The formation of perlolyrine was favored in acidic conditions and temperatures in the range of 70-110 °C. Perlolyrine occurred in the reactions of tryptophan with carbohydrates. The formation rate from fructose was much higher than from glucose. Sucrose also gave perlolyrine under acidic conditions and heating. Perlolyrine was identified in many foods by HPLC-MS and analyzed by HPLC-fluorescence. It occurred in many processed foods such as tomato products including tomato puree, fried tomato, ketchups, tomato juices, and jams but also in soy sauce, beer, balsamic vinegar, fruit juices, dried fruits, fried onion, and honey. The concentrations ranged from an undetected amount to 3.5 µg/g with the highest average levels found in tomato concentrate (1.9 µg/g) and soy sauce (1.5 µg/mL). The results show that perlolyrine formed during the heating process of foods. It is concluded that perlolyrine is widely present in foods and it is daily ingested in the diet.
Subject(s)
Alkaloids , Antineoplastic Agents , Tryptophan , Carbolines , Furans , FruitABSTRACT
Monoamine oxidase (MAO) oxidizes neurotransmitters and xenobiotic amines, including vasopressor and neurotoxic amines such as the MPTP neurotoxin. Its inhibitors are useful as antidepressants and neuroprotectants. This work shows that diluted soy sauce (1/3) and soy sauce extracts inhibited human MAO-A and -B isozymes in vitro, which were measured with a chromatographic assay to avoid interferences, and it suggests the presence of MAO inhibitors. Chromatographic and spectrometric studies showed the occurrence of the ß-carboline alkaloids harman and norharman in soy sauce extracts inhibiting MAO-A. Harman was isolated from soy sauce, and it was a potent and competitive inhibitor of MAO-A (0.4 µM, 44 % inhibition). The concentrations of harman and norharman were determined in commercial soy sauces, reaching 243 and 52 µg/L, respectively. Subsequently, the alkaloids 1,2,3,4-tetrahydro-ß-carboline-3-carboxylic acid (THCA) and 1-methyl-1,2,3,4-tetrahydro-ß-carboline-3-carboxylic acid (MTCA) were identified and analyzed in soy sauces reaching concentrations of 69 and 448 mg/L, respectively. The results show that MTCA was a precursor of harman under oxidative and heating conditions, and soy sauces increased the amount of harman under those conditions. This work shows that soy sauce contains bioactive ß-carbolines and constitutes a dietary source of MAO-A and -B inhibitors.
Subject(s)
Alkaloids , Soy Foods , Humans , Carbolines/pharmacology , Carbolines/analysis , Monoamine Oxidase , Monoamine Oxidase Inhibitors/pharmacology , Monoamine Oxidase Inhibitors/chemistry , Alkaloids/pharmacology , Alkaloids/analysis , Plant Extracts/pharmacology , AminesABSTRACT
Monoamine oxidase (MAO) enzymes (MAO A and B) catalyze the oxidative deamination of biogenic amines, neurotransmitters, and xenobiotic amines and contribute to the regulation of the content of these active substances in mammalian organisms. The oxidation of biogenic amines by MAO produces hydrogen peroxide (H2O2) and aldehydes that represent risk factors for oxidative injury. The inhibitors of MAO are useful as antidepressants and neuroprotective agents. Usually, the assays of MAO determine amine deamination products or measure the H2O2 released by using direct spectrophotometric or fluorimetric methods. Direct methods are more prone to interferences and can afford inaccurate results. Those limitations can be avoided by using chromatographic techniques. This work describes a chromatographic method to assay MAO A and MAO B activity by using kynuramine as a nonselective substrate and the subsequent analysis of 4-hydroxyquinoline by RP-HPLC-DAD-fluorescence and mass spectrometry (MS). Alternatively, the assay uses the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxin as a substrate of MAO that is oxidized (bioactivated) to neurotoxic pyridinium cations which are analyzed by HPLC. These methods are applied to assess the inhibition of MAO by bioactive ß-carboline alkaloids occurring in foods, plants, and biological systems.
Subject(s)
Alkaloids , Neuroprotective Agents , 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine , Aldehydes , Alkaloids/analysis , Animals , Carbolines/chemistry , Carbolines/pharmacology , Cations , Chromatography, High Pressure Liquid/methods , Hydrogen Peroxide , Kynuramine , Mammals , Mass Spectrometry , Monoamine Oxidase , Monoamine Oxidase Inhibitors/chemistry , Monoamine Oxidase Inhibitors/pharmacology , Neuroprotective Agents/pharmacology , Neurotoxins/analysis , XenobioticsABSTRACT
ß-Carbolines (ßCs) are naturally occurring bioactive alkaloids, whereas α-dicarbonyl compounds are reactive substances generated in foods and in vivo. In this work, l-tryptophan reacted with α-dicarbonyl compounds affording new ß-carbolines. Glyoxal afforded 1-hydroxymethyl-ß-carboline (HME-ßC) and its 3-carboxylic acid, and methylglyoxal afforded 1-(1-hydroxyethyl)-ß-carboline (HET-ßC) and its 3-carboxylic acid. 3-Deoxyglucosone afforded 1-(1,3,4,5-tetrahydroxypent-1-yl)-ß-carboline isomers (1a/b), 1-(1,4,5-trihydroxypent-1-yl)-ß-carboline (2), and 1-(1,5-dihydroxypent-3-en-1-yl)-ß-carboline (3). The formation of these ßCs increased under acidic conditions and with increasing temperature. A mechanism is proposed explaining the conversion of a carbonyl into a hydroxy group based on tautomerism and cyclization to the dihydro-ßC-3-COOH intermediates, which were isolated and gave the ßCs. These α-dicarbonyl-derived ßCs occurred in model reactions of l-tryptophan with fructose or glucose incubated under heating and can be considered as advanced glycation end products (AGEs). They were also present in foods and formed during heating processes. HET-ßC appeared in processed foods, reaching up to 309 ng/g, with the highest amount found in dried tomato, fried onion, toasted bread, and Manuka honey. HME-ßC was only detected in some foods with lower amounts than HET-ßC. HET-ßC appeared in foods as a racemic mixture of enantiomers suggesting the same mechanism of formation as the synthetized product. α-Dicarbonyl-derived ßCs (HET-ßC, HME-ßC, and 1a/b-3) occur in foods and food processing and, therefore, they are ingested during diet.
Subject(s)
Alkaloids , Tryptophan , Carbolines , Carboxylic AcidsABSTRACT
ß-Carbolines are naturally occurring bioactive alkaloids. In this work, carbohydrate-derived ß-carbolines (ßCs), 1-(1,3,4,5-tetrahydroxypent-1-yl)-ß-carboline isomers (1a/b), 1-(1,4,5-trihydroxypent-1-yl)-ß-carboline (2), 1-(1,5-dihydroxypent-3-en-1-yl)-ß-carboline (3), and 1-(1,2,3,4,5-pentahydroxypent-1-yl)-ß-carboline (4) were identified and analyzed in commercial foods. The concentrations of ßCs 1-4 in foods ranged from undetectable to 11.4 µg/g levels, suggesting their intake in the diet. Processed foods contained higher amounts than fresh or unprocessed foods, and the highest content was found in processed tomato and fruit products, sauces, and baked foods. ßCs 1-3 were formed in foods during heating, and 1a/b were the main compounds. The formation of carbohydrate-derived ßCs was studied in model reactions of tryptophan and carbohydrates. They formed in reactions of tryptophan with glucose under acidic conditions at temperatures higher than 80 °C. The formation of 1a/b was favored, but 2-3 increased at high temperatures. Noticeably, the ßCs 1-3 formed in the reactions of tryptophan with fructose or sucrose, and the formation from fructose was much higher than from glucose. Thus, fructose was the main carbohydrate involved in the formation of 1-3, whereas sucrose gave these ßCs after acid hydrolysis. It is shown for the first time that the mechanism of formation of ßCs 1-3 occurs from the sugar intermediate 3-deoxyglucosone that reacts with tryptophan affording these carbohydrate-derived ßCs. A mechanism of reaction to give ßCs 1-3 is proposed that relies on the tautomerism (keto-enediol or enamine-imine) of intermediates involved in the reaction. Carbohydrate ßCs 1-4 were assessed as inhibitors of monoamine oxidase (MAO), as antioxidants, and for their interaction with DNA. They were not good inhibitors of MAO-A or -B, were poor antioxidants, and did not appreciably interact with DNA.
Subject(s)
Alkaloids , Fructose , Carbohydrates , Carbolines , Deoxyglucose/analogs & derivativesABSTRACT
We report the discovery of new 4-hydroxyphenyl phosphonium salt derivatives active in the submicromolar range (EC50 from 0.04 to 0.28 µM, SI > 10) against the protozoan parasite Leishmania donovani. The pharmacokinetics and in vivo oral efficacy of compound 1 [(16-(2,4-dihydroxyphenyl)-16-oxohexadecyl)triphenylphosphonium bromide] in a mouse model of visceral leishmaniasis were established. Compound 1 reduced the parasite load in spleen (98.9%) and liver (95.3%) of infected mice after an oral dosage of four daily doses of 1.5 mg/kg. Mode of action studies showed that compound 1 diffuses across the plasma membrane, as designed, and targets the mitochondrion of Leishmania parasites. Disruption of the energetic metabolism, with a decrease of intracellular ATP levels as well as mitochondrial depolarization together with a significant reactive oxygen species production, contributes to the leishmanicidal effect of 1. Importantly, this compound was equally effective against antimonials and miltefosine-resistant clinical isolates of Leishmania infantum, indicating its potential as antileishmanial lead.
Subject(s)
Antiprotozoal Agents/chemistry , Antiprotozoal Agents/therapeutic use , Leishmaniasis, Visceral/drug therapy , Animals , Antiprotozoal Agents/chemical synthesis , DNA Fragmentation , Drug Discovery , Drug Resistance , Female , Leishmania donovani/drug effects , Leishmania infantum/drug effects , Macrophages/parasitology , Mice , Mice, Inbred BALB C , Parasite Load , Reactive Oxygen Species , Structure-Activity RelationshipABSTRACT
Malaria caused by Plasmodium affects millions people worldwide. Plasmodium consumes hemoglobin during its intraerythrocytic stage leaving toxic heme. Parasite detoxifies free heme through formation of hemozoin (ß-hematin) pigment. Proteolysis of hemoglobin and formation of hemozoin are two main targets for antimalarial drugs. Quinoline antimarial drugs and analogs (ß-carbolines or nitroindazoles) were studied as inhibitors of ß-hematin formation. The most potent inhibitors were quinacrine, chloroquine, and amodiaquine followed by quinidine, mefloquine and quinine whereas 8-hydroxyquinoline and ß-carbolines had no effect. Compounds that inhibited ß-hematin increased free hemin that promoted peroxidative reactions as determined with TMB and ABTS substrates. Hemin-catalyzed peroxidative reactions were potentiated in presence of proteins (i.e. globin or BSA) while antioxidants and peroxidase inhibitors decreased peroxidation. Free hemin increased by chloroquine action promoted oxidative reactions resulting in inhibition of proteolysis by three cysteine proteases: papain, ficin and cathepsin B. Glutathione reversed inhibition of proteolysis. These results show that active quinolines inhibit hemozoin and increase free hemin which in presence of H2O2 that abounds in parasite digestive vacuole catalyzes peroxidative reactions and inhibition of cysteine proteases. This work suggests a link between the action of quinoline drugs with biochemical processes of peroxidation and inhibition of proteolysis.
Subject(s)
Antimalarials/pharmacology , Cysteine Proteases/metabolism , Hemeproteins/antagonists & inhibitors , Hemin/metabolism , Plasmodium/drug effects , Quinolines/pharmacology , Oxidation-Reduction , ProteolysisABSTRACT
Monoamine oxidase (MAO) catalyzes the oxidative deamination of amines and neurotransmitters and is involved in mood disorders, depression, oxidative stress, and adverse pharmacological reactions. This work studies the inhibition of human MAO-A by Hypericum perforatum, Peganum harmala, and Lepidium meyenii, which are reported to improve and affect mood and mental conditions. Subsequently, the antioxidant activity associated with the inhibition of MAO is determined in plant extracts for the first time. H. perforatum inhibited human MAO-A, and extracts from flowers gave the highest inhibition (IC50 of 63.6 µg/mL). Plant extracts were analyzed by HPLC-DAD-MS and contained pseudohypericin, hypericin, hyperforin, adhyperforin, hyperfirin, and flavonoids. Hyperforin did not inhibit human MAO-A and hypericin was a poor inhibitor of this isoenzyme. Quercetin and flavonoids significantly contributed to MAO-A inhibition. P. harmala seed extracts highly inhibited MAO-A (IC50 of 49.9 µg/L), being a thousand times more potent than H. perforatum extracts owing to its content of ß-carboline alkaloids (harmaline and harmine). L. meyenii root (maca) extracts did not inhibit MAO-A. These plants may exert protective actions related to antioxidant effects. Results in this work show that P. harmala and H. perforatum extracts exhibit antioxidant activity associated with the inhibition of MAO (i.e., lower production of H2O2).
Subject(s)
Antidepressive Agents/pharmacology , Antioxidants/pharmacology , Monoamine Oxidase Inhibitors/pharmacology , Monoamine Oxidase/metabolism , Plant Extracts/pharmacology , Alkaloids/pharmacology , Anthracenes , Flavonoids/pharmacology , Humans , Hypericum/chemistry , Lepidium/chemistry , Peganum/chemistry , Perylene/analogs & derivatives , Perylene/pharmacology , Phloroglucinol/analogs & derivatives , Phloroglucinol/pharmacology , Terpenes/pharmacology , Triterpenes/pharmacologyABSTRACT
African trypanosomiasis is a neglected parasitic disease that is still of great public health relevance, and a severe impediment to agriculture in endemic areas. The pathogens possess certain unique metabolic features that can be exploited for the development of new drugs. Notably, they rely on an essential, mitochondrially-localized enzyme, Trypanosome Alternative Oxidase (TAO) for their energy metabolism, which is absent in the mammalian hosts and therefore an attractive target for the design of safe drugs. In this study, we cloned, expressed and purified the physiologically relevant form of TAO, which lacks the N-terminal 25 amino acid mitochondrial targeting sequence (ΔMTS-TAO). A new class of 32 cationic and non-cationic 4-hydroxybenzoate and 4-alkoxybenzaldehyde inhibitors was designed and synthesized, enabling the first structure-activity relationship studies on ΔMTS-TAO. Remarkably, we obtained compounds with enzyme inhibition values (IC50) as low as 2â¯nM, which were efficacious against wild type and multidrug-resistant strains of T. brucei and T. congolense. The inhibitors 13, 15, 16, 19, and 30, designed with a mitochondrion-targeting lipophilic cation tail, displayed trypanocidal potencies comparable to the reference drugs pentamidine and diminazene, and showed no cross-resistance with the critical diamidine and melaminophenyl arsenical classes of trypanocides. The cationic inhibitors 15, 16, 19, 20, and 30 were also much more selective (900 - 344,000) over human cells than the non-targeted neutral derivatives (selectivity >8-fold). A preliminary in vivo study showed that modest doses of 15 and 16 reduced parasitaemia of mice infected with T. b. rhodesiense (STIB900). These compounds represent a promising new class of potent and selective hits against African trypanosomes.
Subject(s)
Benzaldehydes/pharmacology , Mitochondrial Proteins/antagonists & inhibitors , Oxidoreductases/antagonists & inhibitors , Parabens/pharmacology , Plant Proteins/antagonists & inhibitors , Trypanocidal Agents/pharmacology , Trypanosoma brucei brucei/drug effects , Trypanosoma congolense/drug effects , Trypanosoma/drug effects , Benzaldehydes/chemical synthesis , Benzaldehydes/chemistry , Cations/chemistry , Cations/pharmacology , Dose-Response Relationship, Drug , Mitochondrial Proteins/metabolism , Molecular Structure , Oxidoreductases/metabolism , Parabens/chemical synthesis , Parabens/chemistry , Parasitic Sensitivity Tests , Plant Proteins/metabolism , Structure-Activity Relationship , Trypanocidal Agents/chemical synthesis , Trypanocidal Agents/chemistry , Trypanosoma/enzymologyABSTRACT
Monoamine oxidase (MAO) enzymes catalyze the oxidative deamination of biogenic amines and neurotransmitters and produce ammonia, aldehydes, and hydrogen peroxide which is involved in oxidative processes. Inhibitors of MAO-A and -B isozymes are useful as antidepressants and neuroprotectants. The assays of MAO usually measure amine oxidation products or hydrogen peroxide by spectrophotometric techniques. Those assays are often compromised by interfering compounds resulting in poor results. This research describes a new method that combines in the same assay the oxidative deamination of kynuramine to 4-hydroxyquinoline analyzed by HPLC-DAD with the oxidation of tetramethylbenzidine (TMB) (or Amplex Rex) by horseradish peroxidase (HRP) in presence of hydrogen peroxide. The new method was applied to study the inhibition of human MAO-A and -B by bioactive compounds including ß-carboline alkaloids and flavonoids occurring in foods and plants. As determined by HPLC-DAD, ß-carbolines, methylene blue, kaempferol and clorgyline inhibited MAO-A and methylene blue, 5-nitroindazole, norharman and deprenyl inhibited MAO-B, and all of them inhibited the oxidation of TMB in the same extent. The flavonoids catechin and cyanidin were not inhibitors of MAO by HPLC-DAD but highly inhibited the oxidation of TMB (or Amplex Red) by peroxidase whereas quercetin and resveratrol were moderate inhibitors of MAO-A by HPLC-DAD, but inhibited the peroxidase assay in a higher level. For some phenolic compounds, using the peroxidase-coupled assay to measure MAO activity led to mistaken results. The new method permits to discern between true inhibitors of MAO from those that are antioxidants and which interfere with peroxidase assays but do not inhibit MAO. For true inhibitors of MAO, inhibition as determined by HPLC-DAD correlated well with inhibition of the oxidation of TMB and this approach can be used to assess the in vitro antioxidant activity (less hydrogen peroxide production) resulting from MAO inhibition.
Subject(s)
Chromatography, High Pressure Liquid/methods , Monoamine Oxidase Inhibitors/metabolism , Monoamine Oxidase/analysis , Monoamine Oxidase/metabolism , Peroxidase/metabolism , Antioxidants/metabolism , Carbolines , Flavonoids , Humans , Kynuramine/analysis , Kynuramine/metabolism , Oxidation-Reduction , Plant Extracts/analysis , Plant Extracts/metabolismABSTRACT
Endogenous and dietary nitrite produces reactive nitrogen species (RNS) that react with DNA causing mutations. The nitrosation of 2'-deoxyguanosine (dGuo) and DNA with nitrite was studied under different conditions, and the reaction and degradation products identified and analysed by HPLC-DAD-MS. Nitrosative deamination of dGuo produced xanthine along with 2'-deoxyxanthosine whereas DNA afforded xanthine. Formation of xanthine increased with nitrite concentration and in low pH such as that of stomach. Xanthine was measured as a marker of nitrosation of dGuo and DNA, and it was subsequently used to study the antinitrosating activity of ß-carboline alkaloids, and selected antioxidants. Food-occurring tetrahydro-ß-carbolines (THßCs) decreased nitrosative deamination of dGuo and DNA under conditions simulating the stomach. Antinitrosating activity was also evidenced for flavonoids (catechin, quercetin) and indole (melatonin) antioxidants. Among THßCs the most active antinitrosating compounds were 1,2,3,4-tetrahydro-ß-carboline-3-carboxylic acids (THßC-3-COOHs) that reacted with nitrite to give N-nitroso derivatives as main products along with 3,4-dihydro-ß-carboline-3-carboxylic acids and aromatic ß-carbolines (norharman and harman). Antinitrosating activity of THßCs correlated well with the formation of N-nitroso-THßC-3-COOHs. These N-nitroso derivatives were stable at pH 7 but degraded in acid conditions affording nitrosating species.
Subject(s)
Alkaloids/pharmacology , Antioxidants/pharmacology , Carbolines/pharmacology , DNA/chemistry , Deoxyguanosine/chemistry , Nitrites/chemistry , Alkaloids/chemistry , Antioxidants/chemistry , Carbolines/chemistry , DNA/genetics , Nitrites/toxicity , Nitrosation/drug effectsABSTRACT
Peganum harmala L. is a medicinal plant from the Mediterranean region and Asia currently used for recreative psychoactive purposes (Ayahuasca analogue), and increasingly involved in toxic cases. Its psychopharmacological and toxicological properties are attributed to quinazoline and ß-carboline alkaloids. In this work three major quinazoline alkaloids were isolated from P. harmala extracts and characterized as peganine (vasicine), deoxypeganine (deoxyvasicine) and a novel compound identified by HPLC-DAD-MS and NMR as peganine ß-d-glucopyranosyl-(1 â 6)-ß-d-glucopyranoside (peganine glycoside). Peganine appeared in flowers and leaves in high levels; high amounts of deoxypeganine and peganine were found in immature and green fruits whereas peganine and peganine glycoside accumulated in high amount in dry seeds reaching up to 1 and 3.9% (w/w), respectively. Roots and stems contained low amount of quinazolines. Seeds extracts containing both quinazoline and ß-carboline alkaloids potently inhibited human monoamine oxidase (MAO)-A. However, quinazoline alkaloids did not contribute to MAO inhibition that was due to ß-carbolines, suggesting that MAO-related psychoactive or toxic actions do not arise from quinazolines. Quinazoline alkaloids were poor radical scavengers in the ABTS assay whereas seed extracts had good activity. Quinazoline alkaloids are known to exert bronchodilator and abortifacient actions, and could contribute to such effects reported in P. harmala.
Subject(s)
Alkaloids/chemistry , Alkaloids/pharmacology , Peganum/chemistry , Quinazolines/chemistry , Quinazolines/pharmacology , Alkaloids/analysis , Antioxidants/chemistry , Antioxidants/pharmacology , Chromatography, High Pressure Liquid , Disaccharides/analysis , Disaccharides/pharmacology , Drug Evaluation, Preclinical/methods , Magnetic Resonance Spectroscopy , Molecular Structure , Monoamine Oxidase Inhibitors/chemistry , Monoamine Oxidase Inhibitors/pharmacology , Plants, Medicinal/chemistry , Quinazolines/analysis , Spectrometry, Mass, Electrospray IonizationABSTRACT
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and 1-methyl-4-phenylpyridinium cation (MPP+) are selective dopaminergic neurotoxins producing Parkinsonism. MPTP is activated by monoamine oxidase-B (MAO-B) to MPP+ that inhibits mitochondrial function. Molecules resembling MPTP which afford pyridinium cations are also neurotoxins. The herbicide paraquat (a bipyridinium dication) and the naturally-occurring ß-carboline and isoquinoline alkaloids are structural analogues of MPTP/MPP+. Paraquat generates reactive oxygen species (ROS) producing neurotoxicity by a mechanism that differs from MPTP/MPP+. Human exposure to PQ is increasingly associated with neurodegeneration. Tetrahydro-ß-carbolines (THßCs), ß-carbolines (ßCs) and tetrahydroisoquinolines (TIQs) are bioactive compounds occurring in foods and the human body. They are not MPTP-like toxins and do not appear to induce neurotoxicity at normal levels of exposure. Among TIQs, endogenous dopamine-derived TIQs (i.e. salsolinol) and 1-benzyl-TIQ are toxic through ROS generation. In contrast, ß-carbolinium (ßC+s) and isoquinolinium cations (IQ+s) are neurotoxicants resembling MPP+ although they are less potent and selective. ßC+s and IQ+s have been detected in the human brain but their toxicological significance remains unknown. THßCs/ßCs and TIQs are activated to toxic cations by N-methyltransferases (NMT) and/or heme peroxidases and are metabolized by cytochrome P450 enzymes. Remarkably, recent findings suggest, instead, that ßCs and TIQs are neuroprotectants and neurorestorative, raising the interest of these molecules.
Subject(s)
Alkaloids/pharmacology , Brain/drug effects , Neurotoxins/pharmacology , Pyridines/pharmacology , Pyridinium Compounds/pharmacology , Cations , HumansABSTRACT
Treatment of late-stage sleeping sickness requires drugs that can cross the blood-brain barrier (BBB) to reach the parasites located in the brain. We report here the synthesis and evaluation of four new N-hydroxy and 12 new N-alkoxy derivatives of bisimidazoline leads as potential agents for the treatment of late-stage sleeping sickness. These compounds, which have reduced basicity compared to the parent leads (i.e., are less ionized at physiological pH), were evaluated in vitro against Trypanosoma brucei rhodesiense and in vivo in murine models of first- and second-stage sleeping sickness. Resistance profile, physicochemical parameters, in vitro BBB permeability, and microsomal stability also were determined. The N-hydroxy imidazoline analogues were the most effective in vivo, with 4-((1-hydroxy-4,5-dihydro-1H-imidazol-2-yl)amino)-N-(4-((1-hydroxy-4,5-dihydro-1H-imidazol-2-yl)amino)phenyl)benzamide (14d) showing 100% cures in the first-stage disease, while 15d, 16d, and 17d appeared to slightly improve survival. In addition, 14d showed weak activity in the chronic model of central nervous system infection in mice. No evidence of reduction of this compound with hepatic microsomes and mitochondria was found in vitro, suggesting that N-hydroxy imidazolines are metabolically stable and have intrinsic activity against T. brucei. In contrast to its unsubstituted parent compound, the uptake of 14d in T. brucei was independent of known drug transporters (i.e., T. brucei AT1/P2 and HAPT), indicating a lower predisposition to cross-resistance with other diamidines and arsenical drugs. Hence, the N-hydroxy bisimidazolines (14d in particular) represent a new class of promising antitrypanosomal agents.
Subject(s)
Trypanocidal Agents/therapeutic use , Trypanosoma brucei brucei/pathogenicity , Trypanosoma brucei rhodesiense/pathogenicity , Trypanosomiasis, African/drug therapy , Animals , Disease Models, Animal , Female , Imidazolines/therapeutic use , Mice , Trypanosoma brucei brucei/drug effects , Trypanosoma brucei rhodesiense/drug effectsABSTRACT
ß-Carbolines are bioactive pyridoindole alkaloids occurring in foods, plants and the human body. Their activity as hydroxyl radical (OH) scavengers is reported here by using three different methods: deoxyribose degradation, hydroxylation of benzoate and hydroxylation of 2'-deoxyguanosine to give 8-hydroxy-2'-deoxyguanosine (8-OHdG) as assessed by RP-HPLC (MS). Fenton reactions (Fe(2+)/Fe(3+) plus H2O2) were used for OH generation, and the radical increased in the presence of ascorbic acid or 6-hydroxydopamine as pro-oxidants. ß-Carbolines were scavengers of OH in the three assays and in the presence of pro-oxidants. Tetrahydro-ß-carboline-3-carboxylic acids were active against the hydroxylation of 2'-deoxyguanosine. ß-Carbolines reacted with hydroxyl radicals (OH) affording hydroxy-ß-carbolines, whereas tetrahydro-ß-carbolines gave oxidative and degradation products. On the basis of IC50 and reaction rates (k), ß-carbolines (norharman and harman), and tetrahydro-ß-carbolines (tetrahydro-ß-carboline, 1-methyltetrahydro-ß-carboline and pinoline) were good OH radical scavengers and their activity was comparable to that of the indole, melatonin, which is an effective hydroxyl radical scavenger and antioxidant.
Subject(s)
Alkaloids/pharmacology , Carbolines/pharmacology , Hydroxyl Radical/pharmacology , 8-Hydroxy-2'-Deoxyguanosine , Alkaloids/chemistry , Antioxidants/chemistry , Antioxidants/pharmacology , Ascorbic Acid/chemistry , Carbolines/chemistry , Chromatography, High Pressure Liquid , Deoxyguanosine/analogs & derivatives , Deoxyguanosine/analysis , Hydrogen Peroxide/chemistry , Hydroxyl Radical/chemistry , Melatonin/chemistry , Melatonin/pharmacology , Oxidative Stress/drug effects , Oxidopamine/chemistryABSTRACT
ß-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-ß-carboline alkaloids arising from tryptophan by still unknown way and mechanism. Results in this research show that heme peroxidases catalyzed the oxidation of tetrahydro-ß-carbolines (i.e. 1,2,3,4-tetrahydro-ß-carboline-3-carboxylic acid and 1-methyl-1,2,3,4-tetrahydro-ß-carboline-3-carboxylic acid) into aromatic ß-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-ß-carbolines was inhibited by peroxidase inhibitors such as sodium azide, ascorbic acid, hydroxylamine and excess of H2O2. The formation of aromatic ß-carbolines by heme peroxidases can help to explain the presence and activity of these compounds in biological systems.
Subject(s)
Carbolines/chemistry , Harmine/analogs & derivatives , Peroxidases/chemistry , Tryptophan/chemistry , Ascorbic Acid/pharmacology , Enzyme Inhibitors/pharmacology , Harmine/chemistry , Heme , Horseradish Peroxidase/chemistry , Hydroxylamine/pharmacology , Kinetics , Lactoperoxidase/chemistry , Oxidation-Reduction , Peroxidase/chemistry , Peroxidases/antagonists & inhibitors , Sodium Azide/pharmacologyABSTRACT
5-(2-Aminopropyl)indole (5-IT) is a psychoactive compound that has recently been associated with several fatal and non-fatal intoxications in a number of European countries. There are indications that acute effects may include symptoms of monoaminergic (e.g. serotonin) toxicity and one mechanism involved in the increase of serotonin levels includes the inhibition of monoamine oxidase. This study investigated the effect of 5-IT on human MAO-A and -B isozymes using kynuramine as the substrate. Substrate conversion to 4-hydroxyquinoline was monitored by high-performance liquid chromatography coupled to diode array detection. This method was employed to determine the extent of MAO inhibition (IC50 and Ki ) and it was found that 5-IT was a selective, competitive and reversible inhibitor of MAO-A. 5-IT revealed a relatively potent ability to inhibit MAO-A (IC50 =1.6 µM and Ki =0.25 µM) while MAO-B inhibition was not observed (0-500 µM 5-IT). Under identical experimental conditions, other established inhibitors of MAO-A and antidepressants provided the following IC50 values: clorgyline 16 nM, harmaline 20 nM, toloxatone 6.7 µM and moclobemide >500 µM. These data indicated that 5-IT was less potent than clorgyline and harmaline but more potent than toloxatone and moclobemide under the in-vitro conditions studied. The inhibition of MAO-A suggests that 5-IT by itself or in combination with other substances may be able to potentiate serotonergic/monoaminergic effects and further studies are needed to clarify its relevance to the adverse effects reported for 5-IT.