HPLC法测定血中25-羟基维生素D_3的含量

Determination of 25-OH-D_3 concentration in human serum by HPLC

目的:建立HPLC法测定血中25-羟基维生素D_3的含量,以评价维生素D的营养状况。方法:血清经无水乙醇沉淀蛋白后,以10%二氯甲烷-正己烷提取．进行 HPLC法分析。色谱柱:Hypersil ODS 5 μm,250 × 4．0 mm;流动相:乙腈:水(80:20);流速:1ml/min;检测波长:265 nm;柱温:30℃。结果;25-羟基维生素D_3在80－800 ng/ml浓度范围内线性良好(r=0.9992)。提取回收率大于 84%,方法回收率大于 94%;日内、日间相对标准偏差均小于 5%,以本法测定了 30例体检儿童血清25-羟基维生素D_3浓度。结论:本法操作简便易行,可用于血中25-羟基维生素D_3的测定。

Objective: To establish a HPLC assay for the determination of 25-hydroxyvitamin D3 (25-OH-D3) in human serumand evaluate the nutritional value of vitamin D. Methods: Extracted with dichloromethane/n-hexane (1: 9, v/v) after being depro-teinizated with absolute alcohol, the serum sample was determined by HPLC, 25-OH-Ds was separated on Hypersil ODS 5 u,m,250 ×4. 0 mm column and detected at 265 nm by using acetonitrite/water (80: 20, v/v) as mobile phase when the column temperature was 30℃. Results: The ranges of calibration curves were 80 ~ 800 ng/ml (r =0. 9992). The extracted recovery was more than 84 % , andthe method recovery was more than 94 %. RSD of intra - day and inter - day were less than 5 % . 30 serum samples of children havebeen determined. Conclusion: The method is simple and sensitive. It is suitable for the determination of 25-OH-D3 concentration in