Development of a rapid analytical method for the determination of sulfated bile acids in urine based on flow-injection spectrophotometry by using immobilized enzyme technology.

DOI NDL Digital Collections Web Site Web Site 12 References
  • GAO Xiu-Feng
    Department of Medical Test, College of Medicine, University of Beihua
  • LI Yong-Sheng
    Department of Applied Chemistry, Northeast China Institute of Electrical Power Engineering
  • KARUBE Isao
    Research Center for Advanced Science and Technology, University of Tokyo

Bibliographic Information

Other Title
  • フローインジェクション／固定化酵素分光光度法による尿中硫酸抱合胆汁酸の測定法の開発
  • 技術論文 フローインジェクション/固定化酵素分光光度法による尿中硫酸抱合胆汁酸の測定法の開発
  • ギジュツ ロンブン フロー インジェクション コテイカ コウソ ブンコウ コウドホウ ニ ヨル ニョウ チュウ リュウサン ホウゴウ タンジュウサン ノ ソクテイホウ ノ カイハツ

Search this article

Abstract

A simple assay of sulfated bile acid (SBA) in urine using flow-injection(FI)-spectrophotometry with immobilized enzyme reactors is proposed. The system consists of an injection valve, a switch valve, two immobilized enzymes reactors and a UV-VIS detector with a flow cell. The multi-step enzymatic reactions occur when an injected sample containing SBA passes through the immobilized enzyme reactors. First, SBA will desulfate under the catalysis of immobilized bile acid sulphate sulfatase (BSS) to form 3β-hydroxyl bile acids; the produced 3β-hydroxyl bile acid reacts with nicotinamide adenine dinucleotide (NAD+) under the catalysis of co-immobilized 3β-hydroxylsteroid dehydrogenase (3β-HSD), and is converted to the 3-ketosteroid. Meanwhile, β-NAD+ is converted to reduced nicotinamide adenine dinucleotide (NADH). Then, by the catalysis of immobilized diaphorase, NADH reacts with a novel reagent, called “water soluble tetrazolium blue” (WST-5), to generate a blue diformazan dye, which is detected at 550 nm. By using FI-spectrophotometry manifold and optimized conditions, we obtained a linear response for 1∼75 μM glycolithocholate sulfate (GLCA-S) with a correlation coefficient of 0.999 and an analytical rate of 15 samples per hour. The RSD was less than 1%. The recoveries (91∼108%) of GLCA-S added into urine were satisfactory and the assay correlativity well with the manual UBASTEC kit. Therefore, it will be applicable for urine tests on patients suffering from hepatobiliary disease.<br>

Journal

  • BUNSEKI KAGAKU

    BUNSEKI KAGAKU 52 (3), 195-200, 2003

    The Japan Society for Analytical Chemistry

References(12)*help

See more

Keywords

Details 詳細情報について

Report a problem

Back to top