文章摘要
黄和飞,吕小波.液相色谱串联质谱法测定人体血清中的类固醇激素[J].药学与临床研究,2022,30(4):300~304
液相色谱串联质谱法测定人体血清中的类固醇激素
Determination of Steroid Hormones in Human Serum by Liquid Chromatography Tandem Mass Spectrometry
投稿时间:2022-02-26  修订日期:2022-08-23
DOI:
中文关键词: 类固醇激素  血清  高效液相色谱串联质谱
英文关键词: Steroid hormone  Serum  HPLC-MS/MS
基金项目:
作者单位E-mail
黄和飞 昆明和合医学检验所有限公司 464511258@qq.com 
吕小波 昆明和合医学检验所有限公司 237883996@qq.com 
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中文摘要:
      目的:建立一种能同时测定血清中6种类固醇激素[包括孕酮(P)、17-羟基孕酮(17-OHP)、皮质醇(COL)、去氢表雄酮(DHEA)、双氢睾酮(DHT)、硫酸去氢表雄酮(DHEAS)]的快速、灵敏的高效液相色谱串联质谱(HPLC-MS/MS)方法。方法:血清样本经甲基叔丁基醚液液萃取,高速离心后上清液氮气吹干,加入复溶液后上机检测。采用安捷伦C18(100 mm × 2.1 mm,1.8 μm)色谱柱进行分离,水相A为纯水,有机相B为纯甲醇,质谱采用电喷雾电离(ESI)模式进行离子化,多反应监测模式(MRM)扫描,同位素稀释法进行定量分析。结果:血清中6个类固醇激素检测限(LOD,以信噪比(S/N)≥3为标准)为0.03~1.25 ng·mL-1;定量限(LOQ,以信噪比(S/N)≥10为标准)为0.05~2.50 ng·mL-1;在0.05~4000 ng·mL-1范围内线性关系良好,相关系数(r)≥0.995 0;低浓度样本日内精密度为2.4%~6.2%;中浓度样本日内精密度为2.3%~5.7%;高浓度样本日内精密度为1.3%~5.4%。低浓度样本日间精密度为0.4%~2.6%;中浓度样本日间精密度为0.3%~3.9%;高浓度样本日间精密度为0.2%~4.7%。加标回收率为86%~106%。结论:建立一种能同时测定血清中6个类固醇激素的HPLC-MS/MS方法。该方法简单、快速、灵敏度高、特异性强,可在6.5 min之内完成分离和检测,适用于人体血清中多种类固醇激素含量的定量分析。
英文摘要:
      Objective: To establish a rapid and sensitive HPLC tandem mass spectrometry (HPLC-MS/MS) method for simultaneous determination of six steroid hormones in serum, including progesterone (P), 17-hydroxyprogesterone (17-OHP), cortisol (COL), dehydroepiandrosterone (DHEA), dihydrotestosterone (DHT) and dehydroepiandrosterone sulfate (DHEAS). Methods: The Serum samples were extracted with methyl tert-butyl ether, centrifuged at high speed, dried with liquid nitrogen, redissolved and tested. An Agilent C18 (100 mm × 2.1 mm, 1.8 μm) column was used for separation. Water (A) and methanol (B) were applied for gradient elution. An Electrospray ionization (ESI) mode was used for ionization, multiple reaction monitoring mode (MRM) scanning was used for quantitative analysis by an isotope internal standard method. Results: The HPLC-MS/MS method separated and detected the 6 steroid hormones in serum within 6.5 min. The limits of detection (LOD) of the 6 steroid hormones in serum were 0.03-1.25 ng·mL-1, based on S/N≥3. The limits of quantitation (LOQ, based on signal-to-noise ratio (S/N)≥10) ranged from 0.05-2.50 ng·mL-1. The linearity was good in the range of 0.05-4000 ng·mL-1, and the correlation coefficient (r) was ≥0.995 0. The intra-day precision of low, medium and high concentration samples were 2.4%-6.2%, 2.3%-5.7% and 1.3%-5.4%, and the inter-day precision of low, medium and high concentration samples were 0.4%-2.6%, 0.3%-3.9% and 0.2%-4.7%, respectively. The spiked recoveries were 86%-109% (acceptable standard was 80%-120%). Conclusion: The established HPLC-MS/MS method is simple, rapid, sensitive and specific, and is suitable for the quantitative analysis of various steroid hormones in human serum.
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