柱前衍生-高效液相色谱法测定大熊猫血清中18种游离氨基酸

Determination of 18 free amino acids in giant panda serum by high performance liquid chromatography after derivatization with 2,4-dinitrochlorobenzene

建立了大熊猫血清中18种游离氨基酸的反相高效液相色谱分析方法。血清样品经HPO3沉淀蛋白后,在弱碱性条件（pH 9.0）下游离氨基酸与2,4-二硝基氯苯在90℃水浴中避光反应1.5 h,生成具有紫外吸收的衍生物,用反相C18柱分离,紫外检测器检测,内标法定量。18种氨基酸的工作曲线的相关系数范围为0.9951～0.9999;相对标准偏差为1.4%～4.7%;检出限为0.20～7.72μg/mL;加标回收率为65.9%～118%。

A method for the determination of 18 free amino acids in giant panda serum by pre-colunm derivatizationhigh performance liquid chromatography was developed. After removing protein in serum with HPO3, the free amino acids in supernatant were derivatizated with 2,4-dinitrochlorobenzene in alkalescent media （pH 9.0） in a water bath of 90 ℃. Then the derivatives were separated by Cis column and detected by UV detector, and the amount of amino acids was quantified with internal standard method. The correlation coefficients of regression equations for the 18 free amino acids were between 0. 9951 to 0. 9999; the range of RSD were 1.39 % - 4.67 % ; the range of detection limit were- 0.20- 7.72μg/mL; and the recoveries were 65.9% ～118%.