DNFB柱前衍生化RP-HPLC法测定氨肽素的氨基酸含量

Determination of the content of amino acid by pre-column derivatization and RP-HPLC

目的用二硝基氟苯 (DNFB)柱前衍生化RP HPLC法测定氨肽素中氨基酸的含量。方法采用二硝基氟苯为衍生试剂进行衍生 ,ODS(KromasilC18,4 6mm× 2 5 0mm ,5 μm)色谱柱分离 ,以乙腈 0 0 5mol·L-1乙酸钠水溶液为流动相 ,梯度洗脱 ,3 60nm检测 ,在 3 0min内 ,测定氨肽素的氨基酸组成与含量。结果线性范围 (总氨基酸 1 4种 )为 0 1 5～ 0 75 g·L-1;回收率为 94 8%～ 1 0 3 5 % ;RSD为0 2 6%～ 1 47% (n =5 )。结论DNFB柱前衍生化RP

Objective To develop a pre-column derivatization procedure for the determination of the content of amino acid in the sample by reversed-phase high performance liquid chromatography.Method DNFB was used as a derivatization reagent and separation of DNP-amino acid derivatives were performed on an ODS column. The signals were detected at 360 nm with the UV detector. Result Under the obove conditions all the 14 amino acids in the standard solutions were separated to baseline in 30 min.Conclusion The method is applied to analyzing amino acid composition of ampeptide and good results are obtained.