反相HPLC同时测定人血清视黄醇、α-与γ-生育酚及-β胡萝卜素

Simultaneous Determination of Retinol(VA),Alpha-tocopherol(α-VE), Gamma-tocopherol(γ-VE),and Beta-carotene(BC)in Human Serum by Reversed-Phase HPLC

本文采用国产NucleosilC_(18)层析柱与岛津产其它部件组成HPLC系统,通过内标法(以国产α-醋酸生育酚为内标)测定人血清视黄醇、α-生育酚、γ-生育酚及β-胡萝卜素含量。血清100μl经正己烷提取,氮气挥干,溶于100μl流动相(甲醇:乙腈:氯仿为25:60:15),进样50μl,以1.5ml/min速度洗脱,一次完全洗脱需13min。根据待测组份与内标峰面积比定量。视黄醇、α-生育酚、γ-生育酚及β-胡萝卜素的最低检出浓度分别是0.05、2.06、0.10及0.16μg/ml血清;梯度浓度平均回收率分别是93.83±3.05％,101.51±4.52％,96.42±1.28%及99.11±6.31％。日内变异系数小于4％,日间小于5%。本法已用于实验及人群研究中上述维生素的测定。

A specific assay for several vitamins by reversed phase HPLC was introduced. The system separates VA, α-V_E, γ-V_E, and BC in human serum using a Nucleosil C18, 5μ particle size column with a mobile phase of methanol-acetonitrile-chloroform (25:60:15)at 1.5ml/min and a Shimadzu SPD-MIA detector. With alpha tocopheryl acetate as an internal standard, curves were developed for each compound on the basis of peak area ratios. The recoveries for V_A, α-V_E, γ VE, and BC were 93.83±3.05％, 101.51±44.52％, 96.42±1.28％, and 99.12±6.32％ respectively. The coefficient of variation was less than 4％ within a run or 5％ between runs. The lowest deter minations for V_A, α_V_E, γ_V_E, and BC were 0.05, 2.06, 0.10, and 0.16 μg/ml serum respectively. The method required 100μl serum and 13min of elution time per sample, and proved useful in experimental and epidemiological research.