Effect of Alkaloids Isolated from Phyllodium Pulchellum on Monoamine Levels and Monoamine Oxidase Activity in Rat Brain
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Hindawi Publishing Corporation Evidence-Based Complementary and Alternative Medicine Volume 2016, Article ID 6826175, 6 pages http://dx.doi.org/10.1155/2016/6826175 Research Article Effect of Alkaloids Isolated from Phyllodium pulchellum on Monoamine Levels and Monoamine Oxidase Activity in Rat Brain Lu Cai,1 Chao Wang,1 Xiao-kui Huo,1 Pei-pei Dong,1 Bao-jing Zhang,1 Hou-Li Zhang,1 Shan-shan Huang,1 Bo Zhang,2 Sheng-ming Yu,3 Ming Zhong,3 and Xiao-Chi Ma1 1 College of Pharmacy, Dalian Medical University, Dalian 116044, China 2Department of Neurosurgery, The Second Affiliated Hospital, Dalian Medical University, Dalian 116044, China 3Institute of Nationality Medicine in Guangxi, Nanning 530001, China Correspondence should be addressed to Hou-Li Zhang; [email protected] and Bo Zhang; [email protected] Received 30 September 2015; Revised 28 February 2016; Accepted 24 March 2016 Academic Editor: I-Min Liu Copyright © 2016 Lu Cai et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Phyllodium pulchellum (P. pulchellum) is a folk medicine with a significant number of bioactivities. The aim of this study was to investigate the effects displayed by alkaloids fractions, isolated from the roots of P. pulchellum, on neurotransmitters monoamine levelsandonmonoamineoxidase(MAO)activity.Six alkaloids, which had indolealkylamine or -carboline skeleton, were obtained by chromatographic technologies and identified by spectroscopic methods such as NMR and MS. After treatment with alkaloids of P. pulchellum, the reduction of DA levels (54.55%) and 5-HT levels (35.01%) in rat brain was observed by HPLC-FLD. The effect of alkaloids on the monoamines metabolism was mainly related to MAO inhibition, characterized by IC50 values of 37.35 ± 6.41 and 126.53 ± 5.39 g/mL for MAO-A and MAO-B, respectively. The acute toxicity indicated that P. pulchellum extract was nontoxic. 1. Introduction to interact with diverse neuronal and molecular targets [6]. IAA drugs were 5-hydroxytryptamine (5-HT or serotonin) Phyllodium pulchellum (P. pu l chel lum ) Desv. (Leguminosae) analogs that mainly act on a variety of 5-HT receptors, is a folk medicine distributed in the southern parts of serotonin transporter, or even MAO enzyme that were highly China, with a significant number of bioactivities especially favorable molecular targets for treating depression, anxiety, for the central nervous system (CNS). P. pu l chel lum exhib- schizophrenia, and other psychiatric disturbances [7, 8]. ited hypothermia and mild analgesic effects on rheumatoid Additionally, -carbolines were naturally occurring alkaloids arthritis. Recently, many investigations suggested that the that exhibited a wide range of psychopharmacological effects ethanol extract of P. pu l chel lum had significant bioactivity due to their binding to benzodiazepine, imidazoline, sero- in vivo system of liver fibrosis [1, 2]. P. pu l chel lum is widely tonin, and opiate receptors as well as monoamine oxidase used in traditional medicine with no literature evidence (MAO) activity inhibition [9, 10]. substantiating its safety. To optimize their safe use, it would Dopamine (DA) and serotonin (5-HT) play a major role be urgent to study the safety and the chemical foundation. as neurotransmitters in the control and regulation of the Previous research had shown that chemical constituents central nervous system. DA is one of the most important of P. pu l chel lum included alkaloids, alcohols, and phenols. In excitatory neurotransmitters, involved in a variety of behav- total alkaloids part, indolealkylamine (IAA) and -carboline iors and brain functions, such as motor activity, cognition, type alkaloids were its main constituents [3–5]. To the emotion, positive reinforcement, food intake, and endocrine best of our knowledge, these alkaloids are regarded as the regulation [11]. Changes on DA transmission are associated promising plant-derived compounds to treat CNS illnesses, with Parkinson’s disease and schizophrenia [12]. 5-HT, as a duetotheiruniquecomplexnitrogen-containingstructures conventional neurotransmitter, is involved in the regulation 2 Evidence-Based Complementary and Alternative Medicine Table 1: Effects of P. pulchellum alkaloids administered p.o. in mice. Dose (mg/kg) Obituary/total animals Mortality (%) Mortality latency (h) LD50 (mg/kg) 6000 0/10 0 — 8000 2/10 20 >72, <120 11000 5/10 50 >24, <48 11300 15000 7/10 70 >24, <48 20000 10/10 100 >2, <24 1,000 of mood, sleep, memory, learning, and sexual behavior as 2 WVL: 210 nm a crucial fine-tuner of normal and pathological processes 875 [13]. Alterations in 5-HT transmission are related to some 750 neurological and psychiatric illness including migraine, hal- 625 lucinations, anxiety, and depression [14]. 500 1 Monoamine oxidases (MAOs) are mitochondrial bound 375 (mAU) isoenzymes which catalyze the oxidative deamination of 250 monoamine neurotransmitters, including 5-HT, histamine, 6 125 5 and catecholamines (dopamine, noradrenaline, and 3 4 adrenaline). MAO is classified into two typesA ( and B), −100 according to their sensitivity towards specificity substrates 0.3 5.0 10.0 15.0 20.0 25.0 30.0 35.0 40.0 45.0 50.0 and inhibitors. MAO-A shows a higher affinity for 5-HT (min) and noradrenaline and is selectively inhibited by clorgyline, Figure 1: Chromatogram of total alkaloids of Phyllodium pulchellum whereas MAO-B preferentially deaminates phenylethylamine with six alkaloids (1–6) indicated (210 nm). and benzylamine and is selectively inhibited by l-deprenyl or pargyline. Dopamine is oxidized by both forms of the enzyme in most species [15]. MAO-A inhibitors have proven to be effective in the pharmacological treatment of depression, and during the later stages of the experiment. The histological further developments have provided reversible inhibitors analysis showed an absence of alterations in all organs of MAO-A, which offer antidepressant activity without examined (results were not shown). the serious side effects of the earlier inhibitors. On the P. pu l chel lum is widely used traditionally in the southern other hand, selective inhibitors of MAO-B have found a parts of China with no literature evidence substantiating therapeutic role in the treatment of Parkinson’s disease [16]. its safety, so it is necessary to evaluate the toxicity of this Considering the presence of indolealkylamine and - medicinal herb. In the present study, the LD50 of P. pu l chel lum carboline alkaloids compounds in P. pu l chel lum and the extract was 11300 mg/kg, based on the classification of Loomis previous effects described for these alkaloids on the CNS,it and Hayes [17], namely, that substances with LD50 between becomes relevant to investigate the effects of fractions of P. 5000 and 15000 mg/kg bodyweight are regarded as being pulchellum on monoamines metabolites and MAO activity. practically nontoxic. However, some mild adverse effects such In order to give vital guidance to uses and further develop- as dizziness, trembling, crouching, and sluggishness were ments of P. pu l chel lum , the investigation regarding chemical observed, and the effect was reversible within 30 min and constituents of P. pu l chel lum alkaloid and its influences of vanished after 1 hr. monoamine levels and monoamine oxidases (MAO-A and -B) was carried out in the present paper. 2.2. Chemical Constituents. The total alkaloids were obtained as the CHCl3 extracted materials from the hydrochloric acid 2. Results and Discussion water extract of the roots of P. pu l chel lum with the content of 0.12%. It was also analyzed by HPLC subjected to a RP ∘ 2.1. Acute Toxicity. The results of the acute toxicity were C18 column at 30 CwithDADdetection.Themobilephase shown in Table 1. There was a regular dose-dependent was comprised of water (solvent A) and acetonitrile (solvent increase in mortality and decrease in mortality latency in B) both acidified with 0.03%3 CF COOH using a gradient both sexes of mice after the administration of P. pu l chel lum manner: 5% B– 35% B for 90 min, at 0.8 mL/min. HPLC extract. The first mouse died between 72 and 120 hafter chromatograms of total alkaloids were recorded with UV injection of the 8000 mg/kg dose of the extract, and the detection at 210 nm, as shown in Figure 1. maximum frequency of death occurred at 20000 mg/kg. The Totally, six alkaloids were obtained by various chroma- no-observed–adverse-effect (NOAEL) dose for the extract tography techniques (Figure 2). They were elucidated as N,N- was6000mg/kg,themaximumtolerateddose(MTD:highest dimethyltryptamine (1), 5-methoxy-N,N-dimethyltrypta- dose at which the mice recovered completely) was assumed mine (2), N-methyltetrahydrocarboline (3), 7-methoxy- to be between 6000 mg and 8000 mg/kg, and the single N-methyltetrahydrocarboline (4), tryptamine (5), and N- dose LD50 was 11300 mg/kg (95% confidence limit: 9762– methyl-3-indoylmethanamine (6)fromtheirspectroscopic 13075 mg/kg). The symptom of weight loss was observed data upon comparisons with values reported in the literature Evidence-Based Complementary and Alternative Medicine 3 9 9 8 8 11 11 4 N 4 N 6 5 4 7 3 H3CO 3 5 4a 4a 6a 5a 10 5 10 2 3 N 2 8 10 6 1 6 1 1 9a 2a 2 7a N 7a N 9 N 7 H 7 H H 12 3 9 9 8 8 NH H3CO 6 5 4 4 NH 4 3 2 3 7 6a 5a 5 4a 5 4a N 8 10 2 2 1 6 1 6 1 9a 2a 2 9 N 7a N 7a N H 7 H 7 H 45 6 Figure 2: The alkaloids 1–6 isolated from the roots of P. pulchellum. Table 2: Effects on monoamine neurotransmitters and their metab- In serotonin system, there was a significant reduction olites of the rat brain after administration of P.