Kawasaki T, Maeda M, Tsuji A
J Chromatogr. 1983 Feb 11;272(2):261-8. doi: 10.1016/s0378-4347(00)86128-9.
A high-performance liquid chromatographic method with fluorescence detection is described for the determination of bile acids and their conjugates. After enzymatic conversion of bile acids to 3-oxo-bile acids using the immobilized 3 alpha-hydroxysteroid dehydrogenase reactor column, 3-oxo-bile acids were extracted with a Sep-Pak C18 cartridge, labelled with dansyl hydrazine and then separated by high-performance liquid chromatography on a reversed-phase column. The eluate is monitored by a fluorophotometer at 365 nm (excitation) and 520 nm (emission). Reactions proceed quickly under mild conditions to give fluorescent derivatives. Linearity of the fluorescence intensity (peak height) with the amounts of various bile acids and their conjugates was obtained above 0.5-1.0 pmol. The method is sensitive, reliable and useful for the simultaneous determination of bile acids in biological samples.
本文描述了一种采用荧光检测的高效液相色谱法来测定胆汁酸及其共轭物。使用固定化的3α-羟基类固醇脱氢酶反应器柱将胆汁酸酶促转化为3-氧代胆汁酸后，用Sep-Pak C18柱提取3-氧代胆汁酸，用丹磺酰肼标记，然后在反相柱上通过高效液相色谱进行分离。用荧光光度计在365nm（激发）和520nm（发射）处监测洗脱液。反应在温和条件下快速进行，生成荧光衍生物。在0.5 - 1.0 pmol以上，各种胆汁酸及其共轭物的量与荧光强度（峰高）呈线性关系。该方法灵敏、可靠，可用于同时测定生物样品中的胆汁酸。
