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标题：Simultaneous Determination of 25-Hydroxyvitamin D2 and 25-Hydroxyvitamin D3 in Human Plasma by Liquid Chromatography-Tandem Mass Spectrometry Employing Derivatization with a Cookson-Type Reagent
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作者：Tatsuya HIGASHI ; Daisuke AWADA ; Kazutake SHIMADA 等
期刊名称：Biological and Pharmaceutical Bulletin
印刷版ISSN：0918-6158
电子版ISSN：1347-5215
出版年度：2001
卷号：24
期号：7
页码：738-743
DOI：10.1248/bpb.24.738
出版社：The Pharmaceutical Society of Japan
摘要：A rapid and highly sensitive LC-MS-MS method for simultaneous determination of 25-hydroxyvitamin D₂ [25(OH)D₂] and 25-hydroxyvitamin D₃ [25(OH)D₃] in human plasma has been developed using derivatization with a Cookson-type reagent, 4-[2-(6,7-dimethoxy-4-methyl-3-oxo-3,4-dihydroquinoxalyl)ethyl]-1,2,4-triazoline-3,5-dione (DMEQTAD). The derivatization with DMEQTAD significantly improved the ionization efficiencies of 25(OH)D₂ and 25(OH)D₃ with detection limits of 20 and 12.5 fmol (8 and 5 pg) per injection, respectively. The method employed two steps of solvent extraction but did not require chromatographic purifications for sample pretreatment. The determination was carried out by mass chromatography of the protonated molecular ions formed by atmospheric pressure chemical ionization operating in the positive-ion mode after the derivatization, and 25-hydroxyvitamin D₄ was used as an internal standard. The intra-assay coefficients of variation were below 4.02 and 3.24% for 25(OH)D₂ and 25(OH)D₃, respectively, and the analytical recoveries of both compounds were quantitative. Assay linearity was obtained in the range of 0.05—1 ng per tube and the determination limit was 3 ng/ml for a 20 μl plasma aliquot, for each compound. The developed method was applied to plasma samples obtained from volunteers, two of whom had received vitamin D₂ supplementation, and gave satisfactory results.
关键词：25-hydroxyvitamin D;LC-MS-MS;Cookson-type reagent;derivatization;human plasma