固相萃取偶联液相色谱质谱联用法测定鹿血中睾丸酮的含量  

Determination of testosterone in deer blood by solid phase extraction coupled with liquid chromatography-mass spectrometry

作　　者：胡春兰 陈媛媛 王春民 任贵奇 HU Chun-lan;CHEN Yuan-yuan;WANG Chun-min;REN Gui-qi(Jingfukang Pharmaceutical Group Co.Ltd.,Hebei Province,Chengde 067000,China;Hebei Traditional Chinese Medicine New Auxiliary Material Technology Innovation Center,Hebei Province,Chengde 067000,China)

机构地区：[1]颈复康药业集团有限公司,河北承德067000 [2]河北省中药新辅料技术创新中心,河北承德067000

出　　处：《中国当代医药》2021年第25期23-26,共4页China Modern Medicine

摘　　要：目的建立固相萃取偶联液相色谱质谱联用法测定鹿血中睾丸酮含量的方法,用于鹿血的质量评价。方法首先对样品进行前处理,其次在液相色谱条件和质谱条件下,通过定量子离子(109.06)对鹿血中睾丸酮进行含量测定。样品前处理:乙腈沉淀鹿血中的蛋白,乙酸乙酯初次提取睾丸酮,所得溶液加至活化的SAX(最大上样量为500 mg,体积为3 mL)固相萃取柱,所得液经氮气吹至近干,正己烷萃取3次,再加至活化的Florisil(最大上样量为500 mg,体积为3 mL)固相萃取柱,弃滤液,乙酸乙酯-正己烷(60∶40,V/V)洗脱柱子,氮气吹干。50%甲醇溶解,过0.22μm微孔滤膜后即得供试品溶液。高效液相色谱条件:色谱柱为shim-Pack XR-ODS(2.0 mm×75 mm,2.2μm);流动相为甲醇-0.1%甲酸-水,梯度洗脱;柱温为40℃;流速为0.3 mL/min;进样量为20μL。质谱条件:去簇电压(DP)为60.0 V;碰撞能(CE)为10.0 eV。结果在1.5165×10^(-4)~3.033×10^(-1)μg/mL线性范围下,定量子离子109.06的R=0.9995。平均回收率为66.62%(n=6),RSD=3.44%。该方法测定4个批次鹿血中睾丸酮的含量范围为3.30×10^(-4)~3.48×10^(-4)μg/mL,RSD均<3%。结论本文建立了固相萃取偶联液相色谱质谱联用法测定鹿血中睾丸酮含量的方法,其线性范围和回收率符合法规要求,可以为鹿血的质量评价提供参考。Objective To establish a method for the determination of testosterone in deer blood by solid phase extraction coupled with liquid chromatography-mass spectrometry for the quality control of deer blood.Methods Firstly,the samples were pretreated,and then the testosterone content in deer blood was determined by using fixed quantum ion(109.06)under the conditions of liquid chromatography and mass spectrometry.Build sample preparation:acetonitrile precipitation deer blood protein,ethyl acetate extracted first testosterone,the solution to the activation of SAX(Maximum loading volume was 500 mg,volume was 3 mL)solid phase extraction column,the liquid obtained by nitrogen blow to nearly dry,N-hexane extraction,three times to add to the activated Florisil(maximum loading volume was 500 mg,volume was 3 mL)solid phase extraction column,abandon the filtrate,ethyl acetate,N-hexane(60∶40,V/V)elution pillars,nitrogen blow dry.The solution was dissolved in 50%methanol and passed through 0.22μm microporous filtration membrane.High performance liquid chromatography condition:shim-Pack XR-ODS column(2.0 mm×75 mm,2.2μm).The mobile phase consisted of methanol-0.1%formic acid-water and gradient elution.Column temperature was 40℃.Flow rate was 0.3 mL/min.Sample size was 20μL.The mass spectrum condition:de-clustering voltage(DP)was 60.0 V,collision energy(CE)was 10.0 eV.Results In the linear range of 1.5165×10^(-4) to 3.033×10^(-1)μg/mL,the R of the fixed quantum ion 109.06 was 0.9995.The average recovery was 66.62%(n=6),RSD was 3.44%.The content of testosterone in four batches of deer blood was 3.30×10^(-4) to 3.48×10^(-4)μg/mL,with RSD<3%.Conclusion The method for the determination of testosterone in deer blood by solid phase extraction coupled with liquid chromatography-mass spectrometry was established.The linear range and recovery rate of solid phase extraction coupled with liquid chromatography-mass spectrometry are in line with the requirements of laws and regulations,and could provide reference for the quality eva

关 键 词：液相色谱质谱联用法 固相萃取工艺 鹿血 睾丸酮 含量测定 

分 类 号：R927.1[医药卫生—药学]