异硫氰酸苯酯柱前衍生化反相高效液相色谱法同时测定18种氨基酸

Simultaneous Determination of 18 Amino Acids by Reversed-Phase High Performance Liquid Chromatography with Precolumn Phenylisothiocyanate Derivatization

建立了一种利用反相高效液相色谱同时测定 18种氨基酸的方法。以正亮氨酸为内标物 ,异硫氰酸苯酯为柱前衍生剂 ,用C18色谱柱在柱温 38℃下采用二元梯度洗脱 ,于 2 5 4nm波长处检测。氨基酸质量浓度在 3 5mg/L～ 5 5 6mg/L时 ,其峰面积与内标物峰面积的比值和氨基酸的质量浓度的线性相关系数 ,除胱氨酸 (0 96 2 )外均大于 0 99;18种氨基酸的加标回收率在 96 0 %～ 10 2 .4 %。信噪比为 2时 ,亮氨酸最低检测限为 0 5mg/L。应用该方法对小牛血去蛋白注射液中的游离氨基酸含量进行了测定 。

A reversed phase high performance liquid chromatographic method for simultaneous determination of 18 amino acids is described. The sample was mixed with norleucine as the internal standard, then derivatized with phenylisothiocyanate (PITC) and analyzed on a Kromasil C 18 column at 38 ℃, using gradient elution with detection at 254 nm. The correlation coefficients between the ratios of peak area of amino acid to that of the internal standard and the amino acid concentrations were above 0 99, except for cysteine(Cys)(0 962). The recoveries of amino acids added were 96 0%-102 4%. Detection limit of leucine was 0 5 mg/L. The method was applied to analyze free amino acids in deproteinized calf blood injection and good results were obtained.