Iwata T, Yamaguchi M, Nakamura M
Faculty of Pharmaceutical Sciences, Fukuoka University, Japan.
J Chromatogr. 1987 Oct 9;421(1):43-50. doi: 10.1016/0378-4347(87)80377-8.
A simple and highly sensitive high-performance liquid chromatographic method for the determination of cholesterol and cholestanol in human serum is described. After extraction of serum with n-hexane, these compounds and 1-eicosanol (internal standard) are converted into the corresponding fluorescent carbamic esters by treatment with 3,4-dihydro-6,7-dimethoxy-4-methyl-3-oxo-quinoxaline-2-carbonyl azide in benzene. The derivatives are separated within 32 min on a reversed-phase column (YMC Pack C8) with acetonitrile-methanol-water (81:9:10, v/v/v) as eluent and detected fluorimetrically. The detection limits are 2 pg (5 fmol) and 3 pg (7 fmol) for cholesterol and cholestanol, respectively, at a signal-to-noise ratio of 2 in a 20-microliters injection volume. This sensitivity permits precise determination of cholesterol and cholestanol in 1-5 microliters of normal human serum.
本文描述了一种用于测定人血清中胆固醇和胆甾烷醇的简单且高灵敏度的高效液相色谱法。用正己烷萃取血清后，这些化合物和1-二十烷醇（内标）在苯中用3,4-二氢-6,7-二甲氧基-4-甲基-3-氧代喹喔啉-2-羰基叠氮化物处理，转化为相应的荧光氨基甲酸酯。衍生物在反相柱（YMC Pack C8）上，以乙腈-甲醇-水（81:9:10，v/v/v）为洗脱剂，在32分钟内分离，并进行荧光检测。在20微升进样体积下，信噪比为2时，胆固醇和胆甾烷醇的检测限分别为2皮克（5飞摩尔）和3皮克（7飞摩尔）。这种灵敏度允许精确测定1 - 5微升正常人血清中的胆固醇和胆甾烷醇。
