During the determination of molecular weight calibration curve of thymosin by size exclu-sion chromatography (SEC), the acetonitrile ratio in mobile phase performanced significant influence on not only the retention time of four proteins(e. g. ribonuclease A, human insulin, thymosin al and somatostatin ) , but also the linearity of molecular weight calibration curve. The best linearity was ob-tained at the ratio of acetonitrile by 75% , with a calibration equation of y = - 3. 138 6x + 21. 724 and a linear correlation coefficient r2) of 0. 988 5. In that condition, the theoretical plate numbers of four proteins were between 45 783 and 63 345, tailing factors were between 0. 96 and 1.18, and res-olutions were between 3. 52 and 8. 82. The established LC method to determine molecular weight cal-ibration curve of thymosin by SEC was proved to have excellent separation and good linearity. Thus it can be used to detect the high molecular weight substance in thymosin preparation.%在体积排阻色谱(SEC)法测定胸腺肽分子量校准曲线过程中,流动相中乙腈的比例对核糖核酸酶A、人胰岛素、胸腺肽α1和生长激素释放抑制因子4种蛋白的保留时间有重要影响,进而影响校准曲线的线性关系.当乙腈比例为75％时,胸腺肽分子量校准曲线线性最好,此时分子量校准方程为y=-3.138 6x+21.724,线性相关系数r2=0.988 5.4种蛋白的理论塔板数在45783 ～63345之间,拖尾因子在0.96～1.18之间,分离度在3.52～8.82之间.SEC法测定胸腺肽分子量校准曲线的液相色谱条件对4种蛋白的分离效果优异,分子量校准曲线线性良好,可用于胸腺肽制剂中高分子量物质的检测.
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