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目的建立高效液相色谱-串联质谱(LC-MS/MS)法,用于测定原发性肝癌患者血清中7种甾类激素的含量。方法血清样品采用乙酸乙酯液-液萃取,经丹磺酰氯衍生化后测定。色谱柱为Agilent Poroshell 120EC-C18柱(2.1mm×150mm,2.7μm),流动相为乙腈-0.1%(V∶V)甲酸水溶液,梯度洗脱,流速为0.3mL/min,柱温为35℃,质谱检测采用动态多反应离子监测模式(MRM)。结果 7种甾类激素(雌二醇、雌三醇、雌酮、氢化可的松、睾酮、雄烯二酮和黄体酮)在8min内达到基线分离。在相应浓度范围内线性关系良好,相关系数均>0.99,日内及日间精密度RSD均<15%,提取回收率为80%~119%,基质效应为85%~112%,样品在放置6h后处理、处理后室温放置24h、3个冻融循环以及-80℃保存30d等测定过程中均稳定性良好。结论该方法操作简便、结果准确、灵敏度高、重现性好,适用于原发性肝癌患者体内甾类激素水平检测。
Objective To establish a high performance liquid chromatography-tandem mass spectrometry (LC-MS / MS) method for the determination of seven steroid hormones in serum of patients with primary liver cancer. Methods Serum samples were extracted by liquid-liquid ethyl acetate and derivatized with dansyl chloride. The column was Agilent Poroshell 120EC-C18 column (2.1 mm × 150 mm, 2.7 μm) with a mobile phase of acetonitrile-0.1% (V: V) formic acid in water with a gradient of 0.3 mL / min and a column temperature of 35 ° C , MS detection using dynamic multi-reactive ion monitoring mode (MRM). Results Seven steroid hormones (estradiol, estriol, estrone, hydrocortisone, testosterone, androstenedione and progesterone) were baseline resolved within 8 min. The correlation coefficient was> 0.99. The RSD of intra-day and inter-day precision was less than 15%, the extraction recovery was 80% ~ 119% and matrix effect was 85% ~ 112% After treatment, the samples were stored at room temperature for 24h, after three freeze-thaw cycles and at -80 ℃ for 30 days, the stability was good. Conclusion The method is simple, accurate, sensitive and reproducible. It is suitable for the detection of steroid hormone in patients with primary liver cancer.