用于测定7-乙氧基香豆素和伞形花内酯的自动化高效液相色谱分析法。

Automated high-performance liquid chromatographic assay for the determination of 7-ethoxycoumarin and umbelliferone.

作者信息

Evans R R, Relling M V

机构信息

Pharmaceutical Division, St. Jude Children's Research Hospital, Memphis, TN 38101.

出版信息

J Chromatogr. 1992 Jul 1;578(1):141-5. doi: 10.1016/0378-4347(92)80237-k.

DOI:10.1016/0378-4347(92)80237-k

PMID:1400780

Abstract

An improved high-performance liquid chromatographic assay is presented for the determination of 7-ethoxycoumarin O-deethylase activity. Following a 30-min microsomal incubation, 7-ethoxycoumarin, 4-methylumbelliferone (internal standard), and the metabolite umbelliferone were extracted with chloroform. Separation was achieved with an isocratic mobile phase using a microBondapak phenyl (300 mm x 3.9 mm I.D.) analytical column. The effluent was monitored by fluorescence detection with an excitation wavelength of 360 nm and an emission wavelength of 470 nm. The intra- and inter-assay coefficients of variation were 10 and 6%, respectively. A detection limit of 0.07 micrograms/ml was achieved, making this method suitable for characterizing P-450 activity of human livers.

摘要

本文介绍了一种改进的高效液相色谱法，用于测定7-乙氧基香豆素O-脱乙基酶的活性。在进行30分钟的微粒体孵育后，用氯仿提取7-乙氧基香豆素、4-甲基伞形酮（内标）和代谢产物伞形酮。使用微Bondapak苯基（300 mm x 3.9 mm内径）分析柱，采用等度流动相实现分离。通过荧光检测对流出物进行监测，激发波长为360 nm，发射波长为470 nm。批内和批间变异系数分别为10%和6%。检测限达到0.07微克/毫升，使该方法适用于表征人肝脏的P-450活性。