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BIOSYNTHETIC RESEARCH OF FETAL BILE ACID AND ANALYSIS OF CONGENITAL DISORDER OF BILE ACID BIOSYNTHESIS

胎儿胆汁酸生物合成研究及先天性胆汁酸生物合成障碍分析

基本信息

批准号：
07672323
负责人：
KUROSAWA Takao
金额：
$ 1.47万
依托单位：
HEALTH SCIENCES UNIVERSITY OF HOKKAIDO
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

To clarify the biosynthetic pathway of fetal bile acids and the substrate specificity of beta-oxidation enzymes system were investigated, and the analytic application of the determination of bile acid intermediates in urine with congenital disorder of bile acid biosynthesis was performed to establish a convenient diagnosis for Zellweger syndrome The intermediates of bile acid biosynthesis (C_<27>-bile acids) were synthesized by the nwely developed synthetic method. The stereoisomers of the intermediates and the C_<27>-bile acid analogues containing the precursor for fetal bile acids were also synthesized to make clear the substrate and stereoselectivity of related enzymes(beta-oxidation enzymes). Using the above synthetic standard, the quantitative and simultaneous analytical methods for bile acid intermediates were developed by GC/MS and HPLC.The incubation of synthetic samples with rat liver homogenate was tired and the products were quantitatively determined by the above analytical methods. The results indicated that fetal bile acids (1beta-and 6alpha-hydroxylated cholic acids) are biologically synthesized through the beta-oxidation pathway. The substrate and stereoselectivity of the related enzymes were also investigated and the results showed the substrate specificity conccrning to the numbers of hydroxyl groups and high stereoselectivity for the formation of intermediates. The analytical method were applied to the analysis of bile acids in urine of a patient of Zellweger syndrome. The quantitative analysis clearly showed the deficiency of beta-oxidation system for bile acid biosynthesis. And the C_<27>-precursor of fetal bile acids were also determined.

为了阐明胎儿胆汁酸的生物合成途径和β-氧化酶系统的底物专一性，并对先天性胆汁酸生物合成障碍患者尿中胆汁酸中间体的测定进行了分析应用，以方便地诊断Zellweger综合征。还合成了中间体的立体异构体和含有胎儿胆汁酸前体的胆汁酸类似物，以明确相关酶(β-氧化酶)的底物和立体选择性。利用上述合成标准，建立了胆汁酸中间体的GC/MS和HPLC同时定量分析方法。人工合成样品与大鼠肝匀浆孵育疲倦，用上述分析方法对产物进行定量测定。结果表明，胎儿胆汁酸(1β-和6α-羟基胆酸)是通过β-氧化途径生物合成的。对相关酶的底物和立体选择性进行了研究，结果表明，底物专一性与羟基数目有关，对中间体的形成有较高的立体选择性。将该分析方法应用于Zellweger综合征患者尿液中胆汁酸的分析。定量分析清楚地显示了胆汁酸生物合成的β-氧化系统的不足。并测定了胎儿胆汁酸C_(27)-前体。