细脚拟青霉代谢产物清除自由基和抑制单胺氧化酶活性的研究

用二苯代苦味肼基自由基(DPPH)-TLC法和酶标仪法对一株细脚拟青霉RCEF0394发酵液甲醇提取物的清除自由基活性进行了定性和定量测定,发现该提取物具有较强的清除自由基活性,在浓度为5.0mg/mL,于37℃下保温10min时,它对0.4mg/mL的DPPH自由基的清除率可达75.4%。以大鼠肝脏线粒体单胺氧化酶为靶标的体外实验发现供试细脚拟青霉发酵液有较强的抑制单胺氧化酶活性,且其活性和浓度呈量效关系。该发酵液冻干品对单胺氧化酶的半数抑制浓度IC50为118.1μg/mL。其三氯甲烷提取物对单胺氧化酶的抑制活性明显强于发酵液冻干品,表明该抑制剂可能为极性较低的化合物。进一步的分型试验表明该三氯甲烷提取物对A型单胺氧化酶呈混合抑制,对B型呈竞争性抑制,其Km值分别为0.44mg/L,0.34mg/L。     

一种白僵菌中MAO抑制剂的分离纯化和结构鉴定

本研究对前期筛选出的一株具有较强的单胺氧化酶(MAO)抑制活性的白僵菌菌株Ba02进行了液体培养;通过不同提取剂的提取效果比较,发现乙酸乙酯能较好地提取出该发酵液中单胺氧化酶抑制剂。通过活性指导下的色谱分离,从乙酸乙酯提取物中得到了一种深红色粉末状化合物。活性测定结果显示该化合物在15μg.mL-1时对MAO-A和MAO-B的抑制率分别为97.50%和95.34%。MS和NMR的鉴定结果表明该化合物为卵孢菌素(Oosporein)。虽然该化合物是一已知化合物,但其对单胺氧化酶的抑制活性尚属首次发现。     

单胺氧化酶抑制剂在临床方面的应用

单胺氧化酶(monoamine oxidase,MAO)是人体内天然存在的一种酶,催化单胺类物质氧化脱氨反应的酶。人体内含有两种单胺氧化酶:单胺氧化酶A和单胺氧化酶B。单胺氧化酶A主要分布在儿茶酚胺能神经元中;单胺氧化酶B主要分布在5-羟色胺能神经元、组胺能神经元和神经胶质细胞中,这两种亚型都均可以使单胺类神经递质失活。而单胺氧化酶抑制剂则能够通过抑制单胺氧化酶的对单胺类物质的氧化活性,从而达到减轻或者消除由各种原因引起的单胺类物质减少或单胺氧化酶活性过高导致的疾病。本文主要总结了近几年单胺氧化酶抑制剂在临床上用于治疗帕金森病、抑郁症和幽门螺旋杆菌方面的最新进展。     

单胺氧化酶抑制剂在临床方面的应用

单胺氧化酶（monoamine oxidase, MAO）是人体内天然存在的一种酶,催化单胺类物质氧化脱氨反应的酶。人体内含有两种单胺氧化酶：单胺氧化酶A 和单胺氧化酶B。单胺氧化酶A 主要分布在儿茶酚胺能神经元中;单胺氧化酶B 主要分布在5- 羟色胺能神经元、组胺能神经元和神经胶质细胞中,这两种亚型都均可以使单胺类神经递质失活。而单胺氧化酶抑制剂则能够通过抑制单胺氧化酶的对单胺类物质的氧化活性,从而达到减轻或者消除由各种原因引起的单胺类物质减少或单胺氧化酶活性过高导致的疾病。本文主要总结了近几年单胺氧化酶抑制剂在临床上用于治疗帕金森病、抑郁症和幽门螺旋杆菌方面的最新进展。     

不同菌株白僵菌代谢产物对大鼠肝线粒体单胺氧化酶的抑制作用研究

对8株白僵菌[Beauveriasp.]代谢产物的抑制单胺氧化酶的活性进行测定,发现菌株Ba02和Ba05的乙酸乙酯提取物在终浓度为55μg/mL时,对单胺氧化酶A型(MAO＿＿A)有较强的抑制活性,其抑制率分别为95.9%和83.4%;Ba02及Ba03菌株脂溶性成分在终浓度为55μg/mL时对单胺氧化酶B型(MAO＿B)有较强的抑制活性,其抑制率分别为94.8%和84.1%。浓度和抑制活性关系研究表明在一定浓度范围内Ba02菌株脂溶性成分对MAO的抑制活性呈量效关系,经计算得出其对MAO＿A、MAO＿B抑制的IC50分别为18.3μg/mL、28.2μg/mL;抑制特征曲线表明Ba02菌株脂溶性成分对MAO＿A呈竞争型抑制,对MAO＿B为混合型抑制,Km值分别为0.47×10-5mol/L,0.11×10-5mol/L。     

单胺氧化酶

单胺氧化酶（monoamine oxidase,MAO）是生物体内一种十分重要的酶,它在大脑和周围神经组织中催化一些生物体产生的胺,氧化脱氨产生过氧化氢（H₂O₂）.单胺氧化酶A和B基因的克隆清楚地证明了这些酶是由不同的多肽组成的.单胺氧化酶A和B的基因定位于X染色体（Xp11.23）,都由15个外显子组成,而且它们的内含子-外显子组织是完全一致的.这些事实表明单胺氧化酶A和B的基因很可能从同一个祖先进化而来.单胺氧化酶A和B具有不同的底物和抑制剂专一性,在生物神经递质代谢和行为方面具有不同的作用.     

肼基单胺氧化酶抑制剂活性与电子结构构效关系的计算分析

采用量子化学方法,在DFT/B3LYP/6-31G*基组水平上对肼基单胺氧化酶抑制剂进行了几何构型优化和电子结构计算.根据计算结果,分析了肼基单胺氧化酶抑制剂的抑制活性与电子结构的构效关系,结果表明,肼基单胺氧化酶抑制剂衍生物的活性与最低空轨道的能量E_LUMO与最高占据轨道的能量E_HOMO的差值、分子偶极矩和苯环上5位碳原子电荷密度有显著相关性.     

单胺氧化酶的抑制作为杀虫脒对澳氰菊酯的增效作用

杀虫脒对拟除虫菊酯具有增效作用, 其与溴氰菊酯的共毒系数高达890.3, 增效显著.当用溴氰菊酯单独处理美洲蜚蠊时, 血淋巴内酪胺增加125%, 章鱼胺增加108%;当杀虫脒与溴氰菊酯(1:1)混用处理美洲蜚蠊时, 引起酪胺及章鱼胺增加的量更多, 其分别增加925%及500%, 杀虫脒对蜚蠊体内的单胺氧化酶(MAO)有显著的抑制作用.由此推断, 杀虫脒抑制单胺氧化酶后, 引起酪胺及章鱼胺的积累(不能进行氧化脱氨所致), 造成不正常的生理效应, 这应是杀虫脒对溴氰菊酯的增效机制.     

糖尿病和氨基脲敏感性胺氧化酶活性

氨基脲敏感性胺氧化酶（SSAO）分布广泛,可催化伯胺脱氨基生成醛、过氧化氢和氨.糖尿病、肥胖症等患者血浆SSAO活性增加.SSAO介导的毒性醛类产物增加可导致血管内皮损伤,从而促进动脉粥样硬化和多种糖尿病并发症的发生发展.SSAO对于脂肪组织的内稳态具有重要的调节作用,在脂肪组织的能量平衡中起到重要的作用.脂肪组织中SSAO活性的增加可能是由于甲胺或氨基丙酮等SSAO作用底物增加而对其产生了上调作用.抑制SSAO活性对于糖尿病可能具有治疗价值.     

靛红生物活性研究进展

靛红是一种重要的天然产物,广泛分布于动植物和人体内,具有多种生物活性,在生物体内起着重要的作用.本文对靛红在动物和人体内作用于神经系统、单胺氧化酶、利钠肽以及其抗肿瘤、抗衰老等方面的活性的研究进展进行了综述.     

一株具有抑制单胺氧化酶作用的干酪乳杆菌筛选

【目的】通过体外模型从健康人体粪便内分离筛选出具有抑制单胺氧化酶(MAO)活性的乳酸菌,为今后乳酸菌体内抗衰老的研究提供参考。【方法】采用单胺氧化酶体外抑制模型对乳酸菌的发酵上清及无细胞提取物进行了筛选,并对筛选出的样品进行了两种指标的测定,即样品的剂量效应,以及样品与酶的预保温时间对酶活抑制率的影响;同时利用膜分离技术对不同分子量范围的样品进行了MAO的抑制测定。以筛选出的菌株JH-23为目的菌,通过16S rDNA序列分析及API细菌鉴定系统对菌株进行鉴定。【结果】筛选出的菌株JH-23无细胞提取物对MAO的抑制率达到33.7%。样品经冻干后,在反应浓度为16 mg/mL时抑制率达到53.2%,且MAO抑制率随预保温时间的增加而上升,在30 min之后抑制效果趋于平稳;粗样品经48 h透析后,透析液中的MAO抑制率较透析前明显升高。菌株JH-23的鉴定结果显示其属于干酪乳杆菌。【结论】开发了一种以单胺氧化酶作为靶位酶的新式体外筛选模型,该模型方便快捷且灵敏性高,对之后的抗衰老体内研究有所帮助。筛选出的干酪乳杆菌JH-23细胞裂解物对MAO有抑制作用,其中起到MAO抑制作用的主要是细胞内的小分子类物质。     

Studies on extracts of cockroach foregut employing enzymes inactivating cholinergic and adrenergic materials

Extracts of foregut were treated with cholinesterase and monoamine oxidase. The activity of the extracts was either potentiated or reduced depending on the level of cardioacceleratory activity in the foregut extracts. Low extract activity was potentiated and high extract activity reduced by monoamine oxidase. The reverse was true for cholinesterase. Ingluvial ganglion extracts contained cardioacceleratory activity which when low was potentiated and when high reduced by atropine blocking. These findings provide additional evidence for the presence of cholinergic and adrenergic materials in extracts of insect foregut.     

Isolation and characterization of a monoamine oxidase B selective inhibitor from tobacco smoke

It is well established that tobacco smokers have reduced levels of monoamine oxidase activities both in the brain and peripheral organs. Furthermore, extensive evidence suggests that smokers are less prone to develop Parkinson's disease. These facts, plus the observation that inhibition of monoamine oxidase B protects against the parkinsonian inducing effects of the nigrostriatal neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, have prompted studies to identify monoamine oxidase inhibitors in the tobacco plant and tobacco cigarette smoke. Our previous efforts on cured tobacco leaf extracts have led to the characterization of 2,3,6-trimethyl-1,4-naphthoquinone, a non-selective monoamine oxidase inhibitor, and farnesylacetone, a selective monoamine oxidase B inhibitor. We now have extended these studies to tobacco smoke constituents. Fractionation of the smoke extracts has confirmed and extended the qualitative results of an earlier report [J. Korean Soc. Tob. Sci.1997, 19, 136] demonstrating the inhibitory activity of the terpene trans,trans-farnesol on rat brain MAO-B. In the present study, K(i) values for the inhibition of human, baboon, monkey, dog, rat, and mouse liver MAO-B have been determined. Noteworthy is the absence of inhibitory effects on human placental MAO-A and beef liver MAO-B. A limited structure-activity relationship study of analogs of trans,trans-farnesol is reported. Although the health hazards associated with the use of tobacco products preclude any therapeutic opportunities linked to smoking, these results suggest the possibility of identifying novel structures of compounds that could lead to the development of neuroprotective agents.     

Early changes in mitochondrial monoamine oxidase activity of rat liver during 2-acetylaminofluorene feeding

The activities of mitochondrial type A and B monoamine oxidase were determined in the liver of rats fed a diet containing 2-acetylaminofluorene (AAF). Three days after the initiation of AAF-feeding, there was a significant decrease of type B monoamine oxidase activity without affect on type A enzyme. The decreased activity of type B monoamine oxidase, which reached a minimum after three weeks, was sustained for as long as AAF-feeding was continued. Sex-related difference in response to AAF was seen in the rat with respect to the onset and the intensity of the decreased type B monoamine oxidase activity, male rats being more sensitive to the carcinogen than female rats. In contrast to the in vivo effect, AAF showed a potent inhibitory effect on type A monoamine oxidase, rather than on type B enzyme, when added in vitro. The pI₅₀ values were estimated to be 7.5 against type A monoamine oxidase and 4.1 against type B enzyme, respectively. The in vitro inhibition of both types of monoamine oxidase by AAF was competitive. The K_i values for AAF were calculated to be 9.51 · 10^?9 M for type A monoamine oxidase and 1.30 · 10^?5 M for type B enzyme, respectively. In accordance with the potent inhibitory effect of AAF on type A monoamine oxidase in vitro, a single administration of the carcinogen, at a dose of 50 mg/kg, resulted in a marked and temporal decrease of the enzyme activity in the mitochondria of male rat liver. Recovery of the decreased type B monoamine oxidase activity was slow, and the enzyme activity did not return to control levels, even if rats were fed the basal diet for 2 or 4 weeks after the cessation of AAF-feeding.     

Functional groups of mitochondrial monoamine oxidase]

Functional groups of mitochondrial monoamine oxidase critical for monoamine oxidase activity were investigated by chemical modification of highly purified monoamine oxidase preparations from pig liver by specific inhibitors. The substrate and inhibitory properties of synthesized derivatives of beta-phenylethylamine, containing various acylating and alkylating groups in the p-position of the benzene ring, were studied. It was shown that 4-carbmethoxy-beta-phenylethylamine (I) is readily deaminated by monoamine oxidase, whereas 4-O-acetyl-beta-phenylethylamine (II) is not affected by the enzyme. 4-O-acetyl-beta-phenylethylamine (II) and 4-ethyl-O-chloroacethyl phenol (III) inhibit deamination of tyramine, 4-amino-beta-phenylethylamine, beta-phenylethylamine, 4-chloro-beta-phenylethylamine and serotonin in different degrees. The kinetic studies demonstrated that this inhibition is probably due to the acylating properties of the compounds obtained. Selectivity in inhibition may be accounted for by acylation of the group of monoamine oxidase which is located in the nearest proximity to the nucleophynoamine oxidase which is located in the nearest proximity to the nucleophylic site of monoamine oxidase active centre important for binding of tyramine. This group is neither the imidazole group of histidyl, nor the SH-group of cysteinyl residues of monoamine oxidase protein molecule. Its nature is discussed in the light of the data obtained.     

Kinetic mechanism of monoamine oxidase A

Steady-state kinetic data for monoamine oxidase A in crude extracts suggest an exclusively ping-pong mechanism, in contrast to those for monoamine oxidase B, which indicate alternate mechanisms involving either a binary or ternary complex. In this study, with use of purified monoamine oxidase A, steady-state data for the inhibition by D-amphetamine of the oxidation of primary amines indicate the possibility of a ternary complex mechanism for monoamine oxidase A also. Stopped-flow studies demonstrate that the rate of reoxidation of reduced enzyme is enhanced by substrates but not by the product, 1-methyl-4-phenylpyridinium. Thus, for the A enzyme, the ternary complex with substrate, but not product, is reoxidized at a faster rate than the free, reduced enzyme. For both the A and B forms of monoamine oxidase, the mechanism is determined by competition between alternate pathways on the basis of the relative rate constants and dissociation constants.     

On rat liver mitochondrial monoamine oxidase activity and lipids

Following earlier observations on the retention of 5-hydroxytryptamine oxidizing activity by a purified preparation of monoamine oxidase from rat liver mitochondria, this fraction has been obtained in a water-soluble form by Triton X-100 gradient gel filtration and DEAE-Bio-Gel A chromatography. The soluble fraction appears to depend on Triton X-100 and phospholipids for its activity. The results seem to implicate membrane lipid components in the expression of rat liver mitochondrial monoamine oxidase activity.     

The distribution of monoamine oxidase and α-glycerophosphate dehydrogenase in pig brain

Activities of the enzymes monoamine oxidase (EC 1.4.3.4), alpha-glycerophosphate dehydrogenase (EC 1.1.99.5) and cytochrome oxidase (EC 1.9.3.1) were determined in homogenates and in the mitochondrial fraction prepared from individual regions of pig brain. The variation in the activity of alpha-glycerophosphate dehydrogenase paralleled that of cytochrome oxidase, but this was not the case with monoamine oxidase. The differences in the activities of the enzymes among homogenates of the various regions of the brain persisted in mitochondria prepared from these homogenates. The purification of these three enzymes paralleled each other when mitochondria were prepared, suggesting that the three enzymes are bound to the same particles.     

On preparative separation of brain mitochondrial monoamine oxidases.

A method was developed for solubilization from bovine brain stem mitochondrial fraction of monoamine oxidases deminating biogenic amines. Preparative separation of the monoamine oxidases, possessing different substrate specificities, was achieved by column chromatography on a biospecific adsorbent AH-Sepharose 4 B. The enzyme preparations thus obtained did not contain any detectable by disc-electrophoresis of isoelectrofocusing in polyacrylamide gels proteins which were devoid of the monoamine oxidase activity.     

Isolation of pure,catalytically active human liver monoamine oxidase B: Antibody complex

Monoamine oxidase B was purified from human liver mitochondria using a monoclonal antibody, MAO B-1C2, which recognizes monoamine oxidase B but not A. Triton X-100 extracts of mitochondria were incubated with purified MAO B-1C2 (IgG₁), and the catalytically active enzyme:antibody complex was isolated by affinity chromatography on Protein A-Sepharose. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the complex revealed the presence of four polypeptide bands (monoamine oxidase B, 57,900 dalton; antibody heavy chain, 52,200 dalton; and two light chains, 29,400 and 27,700 dalton), and indicated a 1:1 stoichiometric ratio of enzyme to antibody. This method gave 154-fold purification of the enzyme from mitochondria.