Inhibition of monoamine oxidase by selected C5- and C6-substituted isatin analogues

Previous studies have shown that (E)-5-styrylisatin and (E)-6-styrylisatin are reversible inhibitors of human monoamine oxidase (MAO) A and B. Both homologues are reported to exhibit selective binding to the MAO-B isoform with (E)-5-styrylisatin being the most potent inhibitor. To further investigate these structure-activity relationships (SAR), in the present study, additional C5- and C6-substituted isatin analogues were synthesized and evaluated as inhibitors of recombinant human MAO-A and MAO-B. With the exception of 5-phenylisatin, all of the analogues examined were selective MAO-B inhibitors. The C5-substituted isatins exhibited higher binding affinities to MAO-B than the corresponding C6-substituted homologues. The most potent MAO-B inhibitor, 5-(4-phenylbutyl)isatin, exhibited an IC₅₀ value of 0.66 nM, approximately 13-fold more potent than (E)-5-styrylisatin and 18,500-fold more potent than isatin. The most potent MAO-A inhibitor was found to be 5-phenylisatin with an IC₅₀ value of 562 nM. The results document that substitution at C5 with a variety of substituents is a general strategy for enhancing the MAO-B inhibition potency of isatin. Possible binding orientations of selected isatin analogues within the active site cavities of MAO-A and MAO-B are proposed.     

Thio- and aminocaffeine analogues as inhibitors of human monoamine oxidase

In a recent study it was shown that 8-benzyloxycaffeine analogues act as potent reversible inhibitors of human monoamine oxidase (MAO) A and B. Although the benzyloxy side chain appears to be particularly favorable for enhancing the MAO inhibition potency of caffeine, a variety of other C8 oxy substituents of caffeine also lead to potent MAO inhibition. In an attempt to discover additional C8 substituents of caffeine that lead to potent MAO inhibition and to explore the importance of the ether oxygen for the MAO inhibition properties of C8 oxy-substituted caffeines, a series of 8-sulfanyl- and 8-aminocaffeine analogues were synthesized and their human MAO-A and -B inhibition potencies were compared to those of the 8-oxycaffeines. The results document that the sulfanylcaffeine analogues are reversible competitive MAO-B inhibitors with potencies comparable to those of the oxycaffeines. The most potent inhibitor, 8-{[(4-bromophenyl)methyl]sulfanyl}caffeine, exhibited an IC₅₀ value of 0.167 μM towards MAO-B. While the sulfanylcaffeine analogues also exhibit affinities for MAO-A, they display in general a high degree of MAO-B selectivity. The aminocaffeine analogues, in contrast, proved to be weak MAO inhibitors with a number of analogues exhibiting no binding to the MAO-A and -B isozymes. The results of this study are discussed with reference to possible binding orientations of selected caffeine analogues within the active site cavities of MAO-A and -B. MAO-B selective sulfanylcaffeine derived inhibitors may act as lead compounds for the design of antiparkinsonian therapies.     

The Novel Neuropsychotropic Agent Milacemide Is a Specific Enzyme-Activated Inhibitor of Brain Monoamine Oxidase B

The novel neuropsychotropic agent milacemide hydrochloride (2-n-pentylaminoacetamide HCl) is a highly selective substrate of the B form of monoamine oxidase (EC 1.4.3.4; MAO). Under the in vitro conditions used in the present study, milacemide acts as an enzyme-activated, partially reversible inhibitor of MAO-B. A reversible inhibition of MAO-A activity is also observed at high concentrations. The inhibitory activity of milacemide is significantly greater for MAO-B. In vivo, after single or repeated oral administration, a specific inhibition of MAO-B is apparent in brain and liver, with a lack of inhibition of the MAO-A activity. In contrast to the irreversible inhibitory action of L-deprenyl, the recovery of MAO-B activity in vivo after milacemide administration is significantly faster, a result suggesting that it is a partially reversible inhibitor. The selective inhibitory effect of milacemide for MAO-B in vivo is confirmed by its potentiation of phenylethylamine-induced stereotyped behavior, whereas vasopressor responses to tyramine were not affected. These observations suggest that milacemide could enhance dopaminergic activity in the brain and could be used as therapy for Parkinson's disease in association with L-3,4-dihydroxyphenylalanine.     

Human monoamine oxidase is inhibited by tobacco smoke: beta-carboline alkaloids act as potent and reversible inhibitors

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. Recently, positron emission tomography imaging has shown that smokers have a much lower activity of peripheral and brain MAO-A (30%) and -B (40%) isozymes compared to non-smokers. This MAO inhibition results from a pharmacological effect of smoke, but little is known about its mechanism. Working with mainstream smoke collected from commercial cigarettes we confirmed that cigarette smoke is a potent inhibitor of human MAO-A and -B isozymes. MAO inhibition was partly reversible, competitive for MAO-A, and a mixed-type inhibition for MAO-B. Two beta-carboline alkaloids, norharman (beta-carboline) and harman (1-methyl-beta-carboline), were identified by GC-MS, quantified, and isolated from the mainstream smoke by solid phase extraction and HPLC. Kinetics analysis revealed that beta-carbolines from cigarette smoke were competitive, reversible, and potent inhibitors of MAO enzymes. Norharman was an inhibitor of MAO-A (K(i)=1.2+/-0.18 microM) and MAO-B (K(i)=1.12+/-0.19 microM), and harman of MAO-A (K(i)=55.54+/-5.3nM). Beta-carboline alkaloids are psychopharmacologically active compounds that may occur endogenously in human tissues, including the brain. These results suggest that beta-carboline alkaloids from cigarette smoke acting as potent reversible inhibitors of MAO enzymes may contribute to the MAO-reduced activity produced by tobacco smoke in smokers. The presence of MAO inhibitors in smoke like beta-carbolines and others may help us to understand some of the purported neuropharmacological effects associated with smoking.     

Effect of cadmium and zinc ethanolamine complexes on rat brain monoamine oxidase-B activity in vitro

Monoamine oxidase-B (MAO-B) from rat brain was inhibited strongly by the prepared cadmium and zinc ethanolamine complexes obtained from their sulphate and chloride salts. The inhibition of MAO-B by these complexes was time-dependent and fully reversible after dilution and sedimentation. In vitro, the cadmium ethanolamine complexes were more potent at inhibiting MAO-B than the zinc complexes. The inhibitory effect of these complexes follow the order: TEA>DEA>MEA, due to the alkyl residues and steric effect properties. The inhibition of MAO-B by cadmium and zinc ethanolamine complexes was a noncompetitive type. The K(i) values were calculated. The influence of the complexes on the activity of MAO-B was rather evaluated. It decreased the MAO-B activity. The IC(50) values of the two potent cadmium and zinc triethanolamine complexes on MAO-B were evaluated indicating that the complexes were tightly binding, but reversible inhibitors for MAO-B. In general, these systems may be used for preventing some neurodegenerative diseases.     

Inhibition of monoamine oxidase by 8-benzyloxycaffeine analogues

Based on recent reports that several (E)-8-styrylcaffeinyl analogues are potent reversible inhibitors of monoamine oxidase B (MAO-B), a series of 8-benzyloxycaffeinyl analogues were synthesized and evaluated as inhibitors of baboon liver MAO-B and recombinant human MAO-A and -B. The 8-benzyloxycaffeinyl analogues were found to inhibit reversibly both MAO isoforms with enzyme–inhibitor dissociation constants (K_i values) ranging from 0.14 to 1.30 μM for the inhibition of human MAO-A, and 0.023–0.59 μM for the inhibition of human MAO-B. The most potent MAO-A inhibitor was 8-(3-methylbenzyloxy)caffeine while 8-(3-bromobenzyloxy)caffeine was the most potent MAO-B inhibitor. The analogues inhibited human and baboon MAO-B with similar potencies. A quantitative structure–activity relationship (QSAR) study indicated that the MAO-B inhibition potencies of the 8-benzyloxycaffeinyl analogues are dependent on the Hansch lipophilicity (π) and Hammett electronic (σ) constants of the substituents at C-3 of the benzyloxy ring. Electron-withdrawing substituents with a high degree of lipophilicity enhance inhibition potency. These results are discussed with reference to possible binding orientations of the inhibitors within the active site cavities of MAO-A and -B.     

Synthesis and evaluation of 2-substituted 4(3H)-quinazolinone thioether derivatives as monoamine oxidase inhibitors

In the present study, a series of fourteen 2-mercapto-4(3H)-quinazolinone derivatives was synthesised and evaluated as potential inhibitors of the human monoamine oxidase (MAO) enzymes. Quinazolinone is the oxidised form of quinazoline, and although this class has not yet been extensively explored as MAO inhibitors, it has been shown to possess a wide variety of biological activities. Among the quinazolinone derivatives investigated, seven compounds (IC₅₀?<?1?µM) proved to be potent and specific MAO-B inhibitors, with the most potent inhibitor, 2-[(3-iodobenzyl)thio]quinazolin-4(3H)-one, exhibiting an IC₅₀ value of 0.142?μM. Further investigation showed that this inhibitor is a reversible and competitive inhibitor of MAO-B with a K_i value of 0.068?µM. None of the test compounds were MAO-A inhibitors. Analysis of the structure-activity relationships (SARs) for MAO-B inhibition shows that substitution on the C2 position of quinazolinone with a benzylthio moiety bearing a Cl, Br or I on the meta position yields the most potent inhibitors of the series. In contrast, substitution with the unsubstituted benzylthio moiety (IC₅₀?=?3.03?µM) resulted in significantly weaker inhibition activity towards MAO-B. This study suggests that quinazolinones are promising leads for the development of selective MAO-B inhibitors which may be used for the treatment of neurodegenerative disorders such as Parkinson’s disease.     

Positional scanning of natural product hispidol’s ring-B: discovery of highly selective human monoamine oxidase-B inhibitor analogues downregulating neuroinflammation for management of neurodegenerative diseases

Multifunctional molecules might offer better treatment of complex multifactorial neurological diseases. Monoaminergic pathways dysregulation and neuroinflammation are common convergence points in diverse neurodegenerative and neuropsychiatric disorders. Aiming to target these diseases, polypharmacological agents modulating both monoaminergic pathways and neuroinflammatory were addressed. A library of analogues of the natural product hispidol was prepared and evaluated for inhibition of monoamine oxidases (MAOs) isoforms. Several molecules emerged as selective potential MAO B inhibitors. The most promising compounds were further evaluated in vitro for their impact on microglia viability, induced production of proinflammatory mediators and MAO-B inhibition mechanism. Amongst tested compounds, 1p was a safe potent competitive reversible MAO-B inhibitor and inhibitor of microglial production of neuroinflammatory mediators; NO and PGE₂. In-silico study provided insights into molecular basis of the observed selective MAO B inhibition. This study presents compound 1p as a promising lead compound for management of neurodegenerative disease.     

Selected chromone derivatives as inhibitors of monoamine oxidase

A previous study has shown that a series of C6-benzyloxy substituted chromones exhibit high binding affinities for human monoamine oxidase (MAO) B. In an attempt to discover additional chromones with potent and selective MAO-B inhibitory potencies and to further examine the structure-activity relationships of MAO-B inhibition by chromones, the series was expanded with homologues containing polar functional groups on C3 of the chromone ring. The results demonstrate that 6-[(3-bromobenzyl)oxy]chromones containing acidic and aldehydic functional groups on C3 act as potent reversible MAO-B inhibitors with IC(50) values of 2.8 and 3.7nM, respectively. Interestingly, a 2-hydroxy-2,3-dihydro-1-benzopyran-4-one derivative as well as open-ring 2-acetylphenol analogues of the chromones also were potent MAO-B inhibitors with IC(50) values ranging from 4 to 11nM. Chromone derivatives containing the benzyloxy substituent on C5 of the chromone ring, however, exhibit MAO-B inhibition potencies that are several orders of magnitude weaker. High potency inhibitors of MAO-B may find application in the therapy of neurodegenerative disorders such as Parkinson's disease.     

Inhibition of monoamine oxidase B by selected benzimidazole and caffeine analogues

We have recently reported that a series of (E)-8-styrylcaffeines and (E)-2-styrylbenzimidazoles are moderate to very potent competitive inhibitors of monoamine oxidase B (MAO-B). The most potent member of the series was found to be (E)-8-(3-chlorostyryl)caffeine (CSC) with an enzyme-inhibitor dissociation constant (K(i) value) of 128 nM. In the present study, we have prepared additional caffeine and benzimidazole analogues in an attempt to identify compounds with improved MAO-B inhibition potency while still acting reversibly. The most potent inhibitor among the caffeine analogues was (E)-8-(3,4-dichlorostyryl)caffeine with a K(i) value of 36 nM, approximately 3.5 times more potent than CSC. The most potent inhibitor among the benzimidazole analogues was (E)-2-(4-trifluoromethylstyryl)-1-methylbenzimidazole with a K(i) value of 430 nM. An SAR analysis indicated that the potency of MAO-B inhibition by (E)-2-styryl-1-methylbenzimidazole analogues depended upon the Taft steric parameter (E(s)) of the substituents attached to C-4 of the styryl phenyl ring. Substituents with a large degree of steric hindrance appear to enhance inhibition potency. The proposal that potent MAO-B inhibition by (E)-8-styrylcaffeines and (E)-2-styrylbenzimidazoles can be explained by a mode of binding that involves traversing both the entrance and substrate cavities was supported by the finding that 1-methylbenzimidazole only weakly inhibited MAO-B with a K(i) value of 2084 microM. Without the styryl side chain, 1-methylbenzimidazole is not expected to be able to bind simultaneously to both the entrance and substrate cavities.     

Sulfanylphthalonitrile analogues as selective and potent inhibitors of monoamine oxidase B

It has recently been reported that nitrile containing compounds frequently act as potent monoamine oxidase B (MAO-B) inhibitors. Modelling studies suggest that this high potency inhibition may rely, at least in part, on polar interactions between nitrile functional groups and polar moieties within the MAO-B substrate cavity. In an attempt to identify potent and selective inhibitors of MAO-B and to contribute to the known structure–activity relationships of MAO inhibition by nitrile containing compounds, the present study examined the MAO inhibitory properties of series of novel sulfanylphthalonitriles and sulfanylbenzonitriles. The results document that the evaluated compounds are potent and selective MAO-B inhibitors with most homologues possessing IC₅₀ values in the nanomolar range. In general, the sulfanylphthalonitriles exhibited higher binding affinities for MAO-B than the corresponding sulfanylbenzonitrile homologues. Among the compounds evaluated, 4-[(4-bromobenzyl)sulfanyl]phthalonitrile is a particularly promising inhibitor since it displayed a high degree of selectivity (8720-fold) for MAO-B over MAO-A, and potent MAO-B inhibition (IC₅₀ = 0.025 μM). Based on these observations, this structure may serve as a lead for the development of therapies for neurodegenerative disorders such as Parkinson’s disease.     

Synthesis and evaluation of biaryl derivatives for structural characterization of selective monoamine oxidase B inhibitors toward Parkinson’s disease therapy

Benzyloxyphenyl moiety is a common structure of highly potent, selective and reversible inhibitors of monoamine oxidase B (MAO-B), safinamide and sembragiline. We synthesized 4-(benzyloxy)phenyl and biphenyl-4-yl derivatives including halogen substituents on the terminal aryl unit. In addition, we modified the carbon linker between amine group and the biaryl linked unit. Among synthesized compounds, 12c exhibited the most potent and selective MAO-B inhibitory effect (hMAO-B IC₅₀: 8.9?nM; >10,000-fold selectivity over MAO-A) as a competitive inhibitor. In addition, 12c showed greater MAO-B inhibitory activity and selectivity compared to well-known MAO-B inhibitors such as selegiline, safinamide and sembragiline. In the MPTP-induced mouse model of Parkinson’s disease (PD), 12c significantly protected the tyrosine hydroxylase (TH)-immunopositive DAergic neurons and attenuated the PD-associated behavioral deficits. This study suggests characteristic structures as a MAO-B inhibitor that may provide a good insight for the development of therapeutic agents for PD.     

Inhibition of monoamine oxidase by 8-phenoxymethylcaffeine derivatives

A recent study has reported that a series of 8-benzyloxycaffeines are potent and reversible inhibitors of both human monoamine oxidase (MAO) isoforms, MAO-A and -B. In an attempt to discover additional caffeine derivatives with potent MAO inhibitory activities, and to contribute to the known structure-activity relationships of MAO inhibition by caffeine derived compounds, the present study investigates the MAO inhibitory potencies of series of 8-phenoxymethylcaffeine and 8-[(phenylsulfanyl)methyl]caffeine derivatives. The results document that the 8-phenoxymethylcaffeine derivatives act as potent reversible inhibitors of MAO-B, with IC(50) values ranging from 0.148 to 5.78 μM. In contrast, the 8-[(phenylsulfanyl)methyl]caffeine derivatives were found to be weak inhibitors of MAO-B, with IC(50) values ranging from 4.05 to 124 μM. Neither the 8-phenoxymethylcaffeine nor the 8-[(phenylsulfanyl)methyl]caffeine derivatives exhibited high binding affinities for MAO-A. While less potent than the 8-benzyloxycaffeines as MAO-B inhibitors, this study concludes that 8-phenoxymethylcaffeines may act as useful leads for the design of MAO-B selective inhibitors. Such compounds may find application in the therapy of neurodegenerative disorders such as Parkinson's disease. Using molecular docking experiments, this study also proposes possible binding orientations of selected caffeine derivatives in the active sites of MAO-A and -B.     

Selected C7-substituted chromone derivatives as monoamine oxidase inhibitors

A series of C7-substituted chromone (1-benzopyran-4-one) derivatives were synthesized and evaluated as inhibitors of recombinant human monoamine oxidase (MAO) A and B. The chromones are structurally related to a series of C7-functionalized coumarin (1-benzopyran-2-one) derivatives which has been reported to act as potent MAO inhibitors. The results of the current study document that the chromones are highly potent reversible inhibitors of MAO-B with IC₅₀ values ranging from 0.008 to 0.370 μM. While the chromone derivatives also exhibit affinities for MAO-A, with IC₅₀ values ranging from 0.495 to 8.03 μM, they are selective for the MAO-B isoform. Structure–activity relationships (SAR) show that 7-benzyloxy substitution of chromone is suitable for MAO-B inhibition with tolerance for a variety of substituents and substitution patterns on the benzyloxy ring. It may be concluded that 7-benzyloxychromones are appropriate lead compounds for the design of reversible and selective MAO-B inhibitors. With the aid of modeling studies, potential binding orientations and interactions of selected chromone derivatives in the MAO-A and -B active sites are examined.     

Pyrazoline-based mycobactin analogues as MAO-inhibitors

3,5-Diaryl carbothioamide pyrazolines designed as mycobactin analogs (mycobacterial siderophore) were reported to be potent antitubercular agents under iron limiting condition in our earlier study. Clinical complications of newly introduced antibiotic Linezolid, due its MAO inhibitory activity, prompted us to evaluate our compounds for their MAO-inhibitory activity against rat liver MAO-A and MAO-B as pyrazolines were reported to be antidepressants and MAO inhibitors. The present study carried out with this pilot library of 32 compounds will provide us with necessary information for designing antitubercular molecules with reduced MAO-inhibitory activity and also help us in identifying a selective MAO-B inhibitor which has potential clinical utility in neurodegenerative disorders. Thirty-two compounds analyzed has shown spectrum of activity from selective to nonselective against two isoforms of rat liver MAO-A and MAO-B and also as competitive, reversible to non-competitive, irreversible. It is also interesting to note that anti-tubercular compound 11, 14 and 16 were also found to be selective inhibitors of rat liver MAO-B. Docking studies with human MAO shows that compound 11 interacts with the catalytic site of both the isoforms, suggesting compound 11 as nonselective inhibitor of human MAO isoforms.     

Selective inhibition of monoamine oxidase A by purpurin,an anthraquinone

Monoamine oxidase (MAO) catalyzes the oxidation of monoamines that act as neurotransmitters. During a target-based screening of natural products using two isoforms of recombinant human MAO-A and MAO-B, purpurin (an anthraquinone derivative) was found to potently and selectively inhibit MAO-A, with an IC₅₀ value of 2.50 μM, and not to inhibit MAO-B. Alizarin (also an anthraquinone) inhibited MAO-A less potently with an IC₅₀ value of 30.1 μM. Furthermore, purpurin was a reversible and competitive inhibitor of MAO-A with a K_i value of 0.422 μM. A comparison of their chemical structures suggested the 4-hydroxy group of purpurin might play an important role in its inhibition of MAO-A. Molecular docking simulation showed that the binding affinity of purpurin for MAO-A (?40.0 kcal/mol) was higher than its affinity for MAO-B (?33.9 kcal/mol), and that Ile 207 and Gly 443 of MAO-A were key residues for hydrogen bonding with purpurin. The findings of this study suggest purpurin is a potent, selective, reversible inhibitor of MAO-A, and that it be considered a new potential lead compound for development of novel reversible inhibitors of MAO-A (RIMAs).     

Inhibition of human monoamine oxidase A and B by 5-phenoxy 8-aminoquinoline analogs

8-Aminoquinolines (8-AQs) are important class of anti-infective therapeutics. 5-Phenoxy 8-aminoquinoline analogs have shown improved metabolic stability compared to primaquine. In view or predictive role of monoamine oxidases (MAO) in metabolism of 8-aminoquinolines the 5-phenoxy analogs were evaluated in vitro for the inhibition of recombinant human MAO-A and MAO-B. The analogs were several folds more potent inhibitors of MAO-A and MAO-B compared to primaquine, the parent drug, with selectivity for MAO-B. 5-(4-Trifluoromethylphenoxy)-4-methylprimaquine (6) Inhibited MAO-B with IC(50) value of 150 nM (626-fold more potent than primaquine). These results will have important implications in optimizing metabolic stability of 8-AQs to improve therapeutic value and also indicate scope for development of 8-AQs as selective MAO inhibitors.     

Inhibition of type A monoamine oxidase by coptisine in mouse brain

The inhibitory effects of coptisine, a protoberberine isoquinoline alkaloid, on type A and type B monoamine oxidase (MAO-A and MAO-B) activities in mouse brain were investigated. Coptisine showed an inhibitory effect on MAO-A activity in a concentration-dependent manner using a substrate kynuramine, but coptisine did not inhibit MAO-B activity. Coptisine exhibited 54.3% inhibition of MAO-A activity at 2 microM. The values of Km and Vmax of MAO-A were 151.9 +/- 0.6 microM and 0.40 +/- 0.03 nmol/min/mg protein, respectively (n=5). Coptisine competitively inhibited MAO-A activity with kynuramine. The Ki value of coptisine was 3.3 microM. The inhibition of MAO-A by coptisine was found to be reversible by dialysis of the incubation mixture. These results suggest that coptisine is a potent reversible inhibitor of MAO-A, and that coptisine functions to regulate the catecholamine content.     

PF 9601N [N-(2-propynyl)-2-(5-benzyloxy-indolyl) methylamine], a new MAO-B inhibitor,attenuates MPTP-induced depletion of striatal dopamine levels in C57/BL6 mice

Monoamine oxidase isoform B (MAO-B) is involved in Parkinson's disease (PD) induced by the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine toxin (MPTP) in human and non-human-primate. MAO-B inhibitors, such as L-deprenyl have shown to prevent against MPTP-toxicity in different species, and it has been used in Parkinson therapy, however, the fact that it is metabolized to (-)-methamphetamine and (-)-amphetamine highlights the need to find out new MAO-B inhibitors without a structural amphetaminic moiety. In this context we herein report, for the first time, anywhere a novel non-amphetamine-like MAO-B inhibitor, PF 9601N, N-(2-propynyl)-2-(5-benzyloxy-indolyl) methylamine. This attenuates the MPTP-induced striatal dopamine depletion in young-adult and adult-old C57/BL mice, using different schedules of administration, and which behave "ex vivo" as a slightly more potent and selective MAO-B inhibitor than L-deprenyl, assayed for comparative purposes in the same experimental conditions. The MAO-B ID(50) values were calculated from the total MAO-B activity measured against [14C] phenylethylamine (22 microM) as substrate, at each inhibitor concentration. The MAO-B ID(50) values resulted to be 381 and 577 nmol/kg for PF 9601N and L-deprenyl, respectively. The intraperitoneally (i.p.) co-administration to young-adult C57/BL6 mice of MPTP (30 mg/kg), with different concentrations of PF 9601N or L-deprenyl (29.5-0.357 micromol/kg) showed a dose-dependent protective effect against striatal dopamine depletion, measuring the dopamine contents and its metabolites by HPLC. The ED(50) value proved to be 3.07 micromol/kg without any significant differences between either MAO-B inhibitor. Nevertheless, lower doses of PF 9601N (1.5 micromol/kg) were necessary to get almost total protection, without any change in the DOPAC and HVA content, when administered 2 h before MPTP (30 mg/kg), whereas partial protection (45%) against dopamine depletion was observed in the case of L-deprenyl. In both cases, MAO-B inhibition was a necessary condition in order to observe the protective effect. When adult-old (8-10 months) C57/BL6 mice were used, MPTP (25 mg/kg) administration induced 25 days later, an irreversible dopamine depletion. In these conditions, chronic administration with 0.15 micromol/kg of PF 9601N, before the toxin, every 24 h for 10 days, rendered almost total protection of dopamine depletion, whereas L-deprenyl yielded only 50% protection of the dopamine content, assayed in the same conditions. It is worth remarking, that in both cases MAO-B was not affected. From these results, it can be concluded that PF 9601N attenuates MPTP neurotoxicity "in vivo" better than L-deprenyl through different mechanisms, with special relevance to the protective effect, independent of MAO-B inhibition, observed in the irreversibly MPTP-lesioned adult-old mice. Therefore, this novel non-amphetamine MAO-B inhibitor could be potentially effective in PD therapy.