【摘要】 本文报告了采用高效液相色谱法反相梯度洗脱,邻苯二甲醛和β-巯基乙醇柱前衍生化,萤光检测分析血浆游离氨基酸。实验采用线性洗脱,在50分钟内可同时测定22种氨基酸。血浆样品的预处理简单,衍生化反应的时间仅需1分30秒,血浆样品的实际进样量少于1微升.本测定方法的精确度高,各个氨基酸保留时间的变异系数平均为0.33％±0.26％(SD),峰面积的变异系数平均为1.72％±1.25％(SD),各个氨基酸的浓度在10～300毫微克/微升的范围中,线性关系的相关系数平均为0.996±0.007(SD).准确性亦好,各个氨基酸的回收率平均为96.8％±4.5％(SD).实验中还比较和讨论了梯度型式和洗脱体积,不同固定相和色谱柱温度,流动相中缓冲溶液的pH值和离子浓度以及衍生化反应的时间和温度等因素对分析结果的影响。更多还原

【Abstract】 Amino acids are usually determined by an ami-no acid analyser, the whole procedure consisting of separation of atnino acid mixtures by classical ion-exchange chromatography, post-column deri-vatization of ninhydrin and UV detection, is quite time consuming, especially in the determination of amino acids in physiological fluids. So we have successfully tried to analyse free amino acids in plasma with reversed-phase high performance liquid chromatograhy, using gradient elution, o-phthalaldehyde / 2-mercaptoethanol pre-column derivatization and fluorescence detection. The present method is able to determine twenty two plasma amino acids at one run within fifty minutes. The apparent advantages of this method are \ simple preparation, short period of derivatization, Small sample size, high precision and accuracy, and high recorvery of each amino acid(96.8%?.5%). We have examined experimentally the factors that could influence chromatographic separation, and disclosed suitable conditions for determination, such as program and volume of elution, property and temperature of the column, pH and ion concentration of buffer solutioa in the mobile phase, and the time and temperature of derivatization reaction.更多还原