rHPLC定量分析血浆Hcy方法的比较研究  

Study on Comparison of Methods for Analyzing Plasma Homocysteine Quantitatively by rHPLC

作　　者：范新萍[1] 刘德敏[1] 石建党[1] 汤新之[1] 

机构地区：[1]天津医科大学代谢病医院,300070

出　　处：《天津医药》2003年第9期589-592,共4页Tianjin Medical Journal

基　　金：天津市高等学校科技发展基金项目(项目编号 :991017)

摘　　要：目的 :建立一种高效简单的血浆同型半胱氨酸(Hcy)浓度的定量分析法。方法 :以N -乙酰半胱氨酸为内标 ,SBD -F为衍生剂 ,分别使用硼氢化钠和3 -丁基膦为还原剂及磷酸盐和乙酸盐为流动相 ,应用反相高效液相色谱 -荧光检测器(rHPLC -FD) ,在测定Hcy 标准品的基础上 ,测定63例健康人血浆Hcy 浓度。结果 :所有色谱图均有较好分离效果。应用硼氢化钠为还原剂和磷酸盐为流动相的方法检测Hcy 浓度 ,其最小检测限为1.0μmol/L ,批间差异为4.48 % ,批内差异为4.20 % ,回收率为(98.62±5.05) %。测得健康成人血浆平均Hcy 浓度为(10.11±1.91)μmol/L。同份样品用硼氢化钠还原比用3 -丁基膦还原反应快 ,但色谱图杂峰多 ,用磷酸盐为流动相洗脱时间略长 ,但对色谱柱损害小。结论 :本方法适用于临床实验室对Hcy 的常规定量测定。3 -丁基膦更适合为Hcy 的还原剂 ,使用磷酸盐为流动相可延长色谱柱的使用寿命。Objective:To develop an efficient and simple method for determinating quantificationally total homocysteine(tHcy)concentration.Methods:Using sodium borohydride(NaBH 4 )and tri-n-butylphosphine as reductant and phosphate or acetate as mobile phase respectively,the tHcy concentrations in63healthy indiˉviduals plasma were analyzed by reverse-phase high performance liquid chromatography-fluorescence detecˉtion(rHPLC-FD)with the help of the internal standard N-acetyl-Cysteine and the derivatizing reagent SBD-F.Results:All of the chromatograms showed that the peaks were well separated.When the sample was reduced with NaBH 4 and eluted with phosphate in the experiment,the lower limit of detection was1.0μmol/L,the intra-assay was4.48%,the inter-assay was4.20%,and the recovery was(98.62±5.05)%.The mean tHcy level was(10.11±1.91)μmol/L in healthy persons.The same samples with NaBH 4 reduction reacted rapidly but more unknown peaks were observed.Using phosphate as mobile phase elutes slowly but the damages to column were more slightly.Conclusion:As required in clinical and research laboratory,this method is apˉplicable to routine quantitative analysis of tHcy in plasma.Tri-n-butylphosphine is suitable as reductant and the expiration of column is prolonged utilizing phosphate as mobile phase.

关 键 词：血浆 同型半胱氨酸 rHPLC 定量分析 比较研究 稳定性 同型半胱氨酸血症 

分 类 号：R589.3[医药卫生—内分泌]