高效液相色谱-质谱法测定大鼠血浆中硫酸酯型脱氢表雄酮和孕烯醇酮

HPLC/MS Determination of Dehydroisoandrosterone 3-sulfate and Pregnenolone Sulfate in Rat Plasma

目的：建立同时测定SD大鼠血浆中脱氢表雄酮硫酸酯（DHEAS）和孕烯醇酮硫酸酯（PREGS）的高效液相色谱-质谱法。方法：选择雌酮硫酸酯（ES）作为内标。SD大鼠的血浆使用乙腈沉淀除去蛋白后，经固相萃取，水解，衍生化后，以高效液相色谱-质谱测定脱氢表雄酮硫酸酯和孕烯醇酮硫酸酯的浓度，采用大气压化学离子源（HPLC／APCI-MS），选择离子检测方式。结果：血浆中脱氢表雄酮硫酸酯和孕烯醇酮硫酸酯线性范围分别为：0、050～2．00ng，0．030～2．00ng，相关系数分别为0．9990和0．9979。低、中、高浓度血样的提取回收率在66．8％～82．3％之间，相对回收率为：98．4％～111．6％，rt内、rt间变异均小于15％。正常SD大鼠的血浆脱氢表雄酮硫酸酯和孕烯醇酮硫酸酯分别为0．070±0．020ng／ml、0．13±0．03lng／ml。结论：本实验方法具有灵敏度高、准确度好及变异较小的特点，适合测定SD大鼠血浆中脱氢表雄酮硫酸酯和孕烯酮硫酸酯的含量。

Objective ：To establish a method for simultaneous determination of dehydroisoandrosterone 3-sulfate（DHEAS）and pregnenolone sulfate （PREGS）in rat plasma by HPLC/MS. Method：The plasma samples,added estrogen sulfate （ES）as internal standard,are deposited with acetonitrile （1 ： 2）,extracted by Oasis HLB extraction cartridge,after hydrolysis and derivation,the concentration of DHEAS and PREGS are determined by liquid chromatography/Negative atmosphere pressure chemical ionization-mass spectrometry. Results：The linear ranges of DHEAS and PREGS were 0. 050- 2.00ng （r= 0. 9990）, 0. 03- 2.0ng （r = 0. 9979）,respectively. The extraction recoveries of plasma samples with low,middle and high concentrition were all between 66.8% -82.3%. The method recoveries were 98. 4% - 111.6% ,The RSD of intra-day and inter-day were less than 15 %. The concentrition of DHEAS and PREGS in normal SD rats were 0. 070±0. 020ng/ml, 0. 13±0. 031ng/ml ,respectively. Conclusion：The method is sensitive,accurate,reproducible and could be used for determination of concentration of DHEAS and PREGS in rat＇s plasma.