Farjam A, de Jong G J, Frei R W, Brinkman U A, Haasnoot W, Hamers A R, Schilt R, Huf F A
Department of Analytical Chemistry, Free University, Amsterdam, The Netherlands.
J Chromatogr. 1988 Oct 28;452:419-33. doi: 10.1016/s0021-9673(01)81465-5.
A liquid chromatographic column-switching system for automated sample pretreatment and determination of the anabolic hormone beta-19-nortestosterone (beta-19-NT) and its metabolite alpha-19-nortestosterone (19-norepitestosterone) in calf urine is described. The system consists of an immunoaffinity pre-column (immuno pre-column) packed with Sepharose-immobilized polyclonal antibodies against beta-19-NT, a second pre-column packed with C18 bonded silica and an analytical C18 column. Urine (25 ml) is directly loaded on the immuno pre-column, where the analytes of interest are trapped by the immobilized antibodies. Next the analytes are desorbed selectively with a solution containing an excess of the cross-reacting steroid hormone norgestrel and transferred, via the second pre-column, to the analytical column. The recovery of beta-19-NT in spiked urine samples was over 95%. The detection limit was 50 ng/l for a 25-ml urine injection. The system showed no loss of analytical performance over a 6-month period, during which about 100 samples were analysed with the same immuno pre-column. The general applicability of this sample pretreatment method is discussed.
本文描述了一种液相色谱柱切换系统，用于自动样品预处理以及测定小牛尿液中的合成代谢激素β-19-去甲睾酮（β-19-NT）及其代谢物α-19-去甲睾酮（19-去甲表睾酮）。该系统由一个免疫亲和预柱（免疫预柱）、一个填充有C18键合硅胶的第二预柱和一个分析C18柱组成，其中免疫预柱填充有固定在琼脂糖上的抗β-19-NT多克隆抗体。将25 ml尿液直接加载到免疫预柱上，目标分析物被固定化抗体捕获。接下来，用含有过量交叉反应甾体激素炔诺孕酮的溶液选择性地解吸分析物，并通过第二预柱将其转移到分析柱上。加标尿液样品中β-19-NT的回收率超过95%。对于25 ml尿液进样，检测限为50 ng/l。该系统在6个月内分析性能没有损失，在此期间使用同一免疫预柱分析了约100个样品。讨论了这种样品预处理方法的一般适用性。
