血清羧甲基半胱氨酸(CMCys) AccQ-Tag测定法

Determination of carboxymethylcystein in serum with high performance liquid chromatography

目的 建立血清羧甲基半胱氨酸 (CMCys)含量的高效液相色谱测定法。方法 采用 6 氨基喹啉基 N 羟基琥珀酰亚胺基氨基甲酸酯 (ACQ)为衍生剂 ,与CMCys柱前定量衍生后 ,以WatersHPLC分析仪 ,AQC TagTMC18氨基酸分析柱 ,14 0mmol/L乙酸钠 (pH =5 8)为流动相A ,乙腈为流动相B ,水为流动相C ,进行梯度洗脱 ,荧光检测 (λex=2 5 0nm ,λem=395nm) ,CMCys和其它 18种氨基酸组分在 6 3min内分析完成。结果 本方法测定CMCys在 10～6 0 0 μmol/L范围内线性关系良好 ,相关系数 0 9978,批内和批间CV %分别为 3 7%～ 4 9%和 4 5 %～ 5 8% ,平均回收率为 91%～ 114 %。结论 本方法有操作简单、稳定性好、敏感性和准确性高等特点 。

Objective To establish a method to determine serum carboxymethylcystein(CMCys)with high performance liquid chromatography.Methods CMCys reacts with derivative agent,6aminoquinolyhydroxysuccinimidyl carbamate(AQC),quantitatively before added to an analytical column.The derivative was eluded in gradient using Waters high performance liquid chromatography apparatus with an analytical column of AQCTag TM C18 amino acid,the flowing phase A of 140 mmol/L sodium acetate(pH=58),the flowing phase B of acetonitrile and the flowing phase of water,and then determined by a fluorescence(λ ex =250nm,λ em =395nm)detector.Analysis for CMCys and other 18 amino acids components could be finished in 63 minutes.Results Determination of serum CMCys concentrations within 10 to 600μmol/L with this method showed a good linear pattern,with coefficient of correlation of 0997 8,coefficients of variation of 32%～49% within the same batch of specimens and 45%～58% between different batches of specimens,respectively,and with a rate of recovery of 91%～114%.Conclusion This method of determination showed characteristics of simple performance,stable for analysis and higher sensitivity and specificity,and suitable for serum CMCys determination.