头发中内源性氢化可的松的高效液相色谱/串联质谱检测

Detection of endogenous cortisol in human hair by high performance liquid chromatography tandem mass spectrometry

基于高效液相色谱/串联质谱(HPLC/MS/MS)建立了一种检测头发中内源性氢化可的松含量的方法,并与高效液相色谱/荧光法(HPLC/FLU)的测定结果进行比较.头发样品(50 mg)经过清洗与研磨、甲醇提取和C18固相萃取后,在电喷雾(ESI)负离子模式和多反应监测(MRM)方式下进行定性,以外标法进行定量.方法的定量限为2 pg/mg,工作曲线的线性范围为2～100 pg/mg;在空白头发添加50 pg/mg外标时,方法的回收率为(99.4±9.1)%,日内精密度为7.6%,日间精密度为8.3%.由于前处理提取方法、基质效应和定量方法不同,所以HPLC/MS/MS法的测定结果明显低于HPLC/FLU法的测定结果.尽管如此,皮尔逊相关性分析发现2种方法测定的结果具有显著相关性(r=0.749,p<0.01).

A new method for detection of endogenous cortisol in human hair was developed based on high performance liquid chromatography tandem mass spectrometry（HPLC/MS/MS）,and the results detected was compared with that determined by high performance liquid chromatography with fluorescence detector（HPLC/FLU）.Hair sample（50 mg） was incubated in methanol after being washed and pulverized,and then the extraction was purified by C18 solid phase extraction column.Cortisol identification was performed by electro-spray ionization（ESI） in negative mode using multiple reaction monitoring（MRM） and the quantification was performed using external standard method.The limit of quantification is 2 pg/mg,and the linear range is 2 to 100 pg/mg.The recovery of cortisol spiked at 50 pg/mg is（99.4±9.1） %,intra-day precision is 7.6 %,inter-day precision is 8.3 %.Because of the differences in hair pretreatment method,cortisol extraction method,matrix effect and quantitative method,cortisol concentration detected by HPLC/MS/MS is significantly lower than that determined by HPLC/FLU.However,Pearson correlation analysis showed that there is a significant linear correlation（r=0.749,p〈0.01） between hair cortisol contents analyzed by these two methods.