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目的建立同时分析尿中4种内源性雄激素(睾酮和雄烯二酮)和孕激素(孕酮和21α-羟化孕酮)的超高效液相色谱串联三重四极杆质谱法。方法取1.0 ml尿样用β-葡萄糖醛酸酶水解,C18固相萃取小柱净化,用C18超高效液相色谱柱和含10 mmol/L乙酸铵的甲醇和水流动相梯度洗脱分离后,采用串联四极杆质谱电喷雾阳离子多反应监测模式分析,内标标准曲线法定量。结果在0.1 ng/ml~100 ng/ml浓度范围内,4种内源性激素呈现良好的线性关系,相关系数为0.992~0.9995。以3倍基线噪声所对应的浓度为检出限时,4种内源性激素检出限在0.06 ng/ml~0.12 ng/ml之间。在1 ng/ml和5 ng/ml的加标水平时,4种内源性激素加标回收率平均值为99.4%~111.6%,相对标准偏差在3.2%~14.7%之间。该方法成功用于实际尿样中4种内源性激素的分析。结论该方法快速,灵敏,准确,完全满足尿中上述4种内源性激素的分析要求。
OBJECTIVE To establish a simultaneous determination of 4 endogenous androgens (testosterone and androstenedione) and progestins (progesterone and 21α-hydroxy progesterone) in urine by tandem triple-quadrupole mass spectrometry. Methods Urine samples were extracted with β-glucuronidase (1.0 ml) and cleaned up by C18 solid phase extraction (SPE). The separation was performed on a C18 ultra-high performance liquid chromatography column and a mobile phase of methanol and water containing 10 mmol / L ammonium acetate , Using tandem quadrupole mass spectrometry electrospray cation multi-reaction monitoring mode analysis, internal standard curve method. Results The four endogenous hormones showed a good linearity within the range of 0.1 ng / ml ~ 100 ng / ml with the correlation coefficient of 0.992 ~ 0.9995. The limit of detection of four endogenous hormones was between 0.06 ng / ml and 0.12 ng / ml when the concentration corresponding to 3 times the baseline noise was the detection limit. At the spiked levels of 1 ng / ml and 5 ng / ml, the average recoveries of four endogenous hormones were between 99.4% and 111.6% with relative standard deviations between 3.2% and 14.7%. The method was successfully used for the analysis of four endogenous hormones in real urine samples. Conclusion The method is rapid, sensitive and accurate and fully meets the analytical requirements of the above four endogenous hormones in urine.