Clement B, Schultze-Mosgau M H, Wohlers H
Pharmazeutisches Institut, Christian-Albrechts Universität, Kiel, F.R.G.
Biochem Pharmacol. 1993 Dec 14;46(12):2249-67. doi: 10.1016/0006-2952(93)90616-5.
The microsomal N-hydroxylation of the strongly basic guanidinium group (debrisoquine) to N-hydroxyguanidine (N-hydroxydebrisoquine) and the retroreduction of the N-hydroxyguanidine are demonstrated for the first time. The reduction of the N-hydroxyguanidine by liver homogenates and hepatocytes is catalysed by a microsomal NADH-dependent system that is strongly inhibited by hydroxylamine or N-methylhydroxylamine. In the presence of these alternate substrates for the reductase the microsomal catalysed N-hydroxylation of debrisoquine is readily characterized. The oxidation was inhibited by antibodies against NADPH cytochrome P450 reductase and the role of the P450 monooxygenase was further verified by studies with partially purified and purified P450 2C3 reconstituted systems. The transformation of N-hydroxydebrisoquine to the corresponding urea derivative was also detected in in vitro experiments with microsomal fractions and enriched P450 fractions as well as with flavin-containing monooxygenase (FMO). Experiments with catalase, superoxide dismutase and H2O2 have shown that the H2O2 or O2-, respectively, formed from the respective enzyme and the substrate, apparently participated in the reaction. Whereas the N-hydroxylation of the guanidine involves the usual monooxygenase activity of cytochrome P450 the resultant N-hydroxyguanidine decouples monooxygenases (cytochrome P450, FMO) and the H2O2 and, above all, O2- thus formed transform the N-hydroxyguanidine further to the corresponding urea derivative. The possibility for the N-hydroxylation of non-physiological guanidines to N-hydroxyguanidines and subsequent oxidative conversion to the respective urea is comparable to the physiological transformation of arginine to citrulline via N-hydroxyarginine with the liberation of nitric oxide (endothelial derived relaxing factor) and could, therefore, contribute to the efficacy of drugs containing guanidine and similar functional groups.
首次证明了强碱性胍基（地布喹）的微粒体N-羟基化生成N-羟基胍（N-羟基地布喹）以及N-羟基胍的逆向还原。肝匀浆和肝细胞对N-羟基胍的还原由微粒体NADH依赖性系统催化，该系统受到羟胺或N-甲基羟胺的强烈抑制。在存在这些还原酶的替代底物的情况下，微粒体催化的地布喹N-羟基化很容易被表征。该氧化反应被抗NADPH细胞色素P450还原酶的抗体抑制，并且通过使用部分纯化和纯化的P450 2C3重组系统的研究进一步证实了P450单加氧酶的作用。在使用微粒体组分和富集的P450组分以及含黄素单加氧酶（FMO）的体外实验中，也检测到了N-羟基地布喹向相应尿素衍生物的转化。用过氧化氢酶、超氧化物歧化酶和H2O2进行的实验表明，分别由各自的酶和底物形成的H2O2或O2-显然参与了反应。胍基的N-羟基化涉及细胞色素P450通常的单加氧酶活性，而生成的N-羟基胍使单加氧酶（细胞色素P450、FMO）解偶联，由此形成的H2O2，尤其是O2-进一步将N-羟基胍转化为相应的尿素衍生物。非生理性胍类向N-羟基胍类的N-羟基化以及随后氧化转化为相应尿素的可能性与精氨酸通过N-羟基精氨酸向瓜氨酸的生理性转化相当，同时释放一氧化氮（内皮源性舒张因子），因此可能有助于含胍基和类似官能团药物的疗效。
