海洋鱼类血清游离皮质醇液质分析研究

Determination of Free Cortisol in Marine Fish Sera Samples by Liquid Chromatography-Mass Spectrometry

血清样品中加入地塞米松作为定量内标,用氯仿提取,氮气吹干,V(甲醇):V(水)=1:3混合溶液定容后用超高效液相色谱-四极杆-飞行时间质谱(UPLC-Q-TOF-MS)对游离皮质醇进行分析。采用电喷雾电离源负离子模式,分别取m/z407.2和437.2作为皮质醇和内标的定量目标离子进行检测。结果表明,皮质醇与内标在BEHC18色谱柱上得到良好的分离,方法在0～200μg/L浓度范围内线性相关系数为0.9993。仪器检出限为1.64pg。方法平均回收率达到93.3%～98.1%,相对标准偏差小于10%。对采自浙江宁波几批大黄鱼(Preudosciaena Crocea)和鲈鱼(Lateolabrax Japonicus)血清进行测定,游离皮质醇含量在2.11～75.84μg/L范围内。与三重四极质谱仪(Q-Q-Q-MS)进行比较,仪器检测灵敏度要优于三重四极质谱仪;在0～2000μg/L。浓度范围内线性相关不及三重四极质谱仪的分析结果,但在0～200μg/L浓度范围内相关性相近。

The free cortisol in marine fish serum was extracted with chloroform,dried under nitrogen gas,solved in 1∶3 methanol/H2O（V/V） and analyzed by ultra performance liquid chromatography-quadrupole-time of flight-mass spectrometry（UPLC-Q-TOF-MS）,with dexamethasone as internal standard.The mass spectrometry was performed in reflective time-of-flight using electron spraying ionization in negative mode,with m/z 407.2 for cortisol and 437.2 for dexamethasone,respectively.The results show that efficient separation of cortisol and dexamethasone has been achieved on BEH C18 column.The linearity was superior in the range of 0 to 200 μg/L,with correlation coefficient of 0.9993 and low detection limit of 1.64 pg.The average recovery rate was from 93.3% to 98.1%,with RSD less than 10%.Using this method,free corsitol in yellow croaker and Japanese sea bass collected in Ningbo China,have been determined with content ranged from 2.1 to 75.8 μg/L.Comparative analysis indicates that Q-TOF-MS has more efficient analytical sensitivity than quadrupole-quadrapde-quadrupole-mass spectrometry（Q-Q-Q-MS）,and with the similar superior quantitative linearity in the range of 0-200 μg/L,though better for Q-Q-Q-MS in the range of 0-2000 μg/L.