高效阴离子交换色谱法测定人凝血因子Ⅷ中枸橼酸离子的含量  

Determination of citrate ion content in human coagulation factor Ⅷ by high performance anion exchange chromatography

作　　者：张伟[1] 任连杰[1] 张彤[1] 韩春霞 

机构地区：[1]北京市药品检验所,北京100035 [2]赛默飞世尔科技(中国)有限公司,北京100007

出　　处：《中国生物制品学杂志》2014年第1期125-128,共4页Chinese Journal of Biologicals

摘　　要：目的采用高效阴离子交换色谱法测定人凝血因子Ⅷ中枸橼酸离子的含量。方法高效阴离子交换色谱法色谱条件：色谱柱：IonPacASll—HC离子交换柱（4mm×250mm）；流动相：20mmol／L的NaOH溶液，等度洗脱20min，流速：1．0ml／min；抑制器：ASRS4mm阴离子抑制器，抑制电流120mA；检测器：电导检测器，检测池温度30℃，进样量：25μl。对建立的方法进行线性范围、检测限的确定以及准确性、精密性验证，并进行初步应用。结果枸橼酸离子浓度在0．1～2．0μg／ml范围内，对照品色谱峰面积与枸橼酸离子浓度呈良好的线性关系，r=0．9990，按信噪比（S／N）为3确定方法的检测限为0．01μg／ml；准确性试验枸橼酸离子的总平均回收率为97．80％，RSD为0．69％；1份加样人凝血因子Ⅷ溶液连续测定6次峰面积的RSD为0．08％；高效阴离子交换色谱法、比色法及阳离子交换色谱法测定3批人凝血因子Ⅷ中枸橼酸离子含量的结果基本一致，均符合《中国药典》三部（2010版）规定。结论高效阴离子交换色谱法可用于测定人凝血因子Ⅷ中枸橼酸离子的含量，且该方法操作简便、快速，准确性、精密性高。Objective To determine the citrate ion content in human coagulation factor Ⅷ （FⅧ） by high performance anion exchange chromatography （HPAEC）. Methods The condition for HPAEC was as follows： chromatographic col- umn： IonPac AS11-HC（4 mm × 250 mm）; mobile phase： 20 mmol/L sodium hydroxide solution; time for isocratic elu- tion： 20 min; flow rate： 1. 0 ml/min; inhibitor： ASRS 4 mm anion inhibitor; inhibition current： 120 mA; detector： electrical conductivity detector; detector cell temperature： 30 ℃; sample loading： 25 pal. The developed method was de- termined for linear range and detection limit, verified for accuracy and precision, and applied preliminarily. Results The chromatographic peak area of reference showed good linear relationship to citrate ion content at a range of 0. 1 2. 0 μg/ml （r = 0. 999 0）. Serving the signal-to-noise （S/N） ratio as 3, the detection limit of the developed method was 0. 01 μg/ml. The total mean recovery of citrate ion in accuracy test was 97. 80%, with a RSD of 0. 69%. The RSD of 6 test results of one sample of FⅧ was 0. 08%. The determination results of three batches of FⅧ by HPAEC, colorimetry and cation exchange chromatography were basically in agreement, which met the requirements in Chinese Phar- macopoeia（Volume Ⅲ, 2010 edition）. Conclusion HPAEC may be used for determination of citrate ion content in FⅧ, which was simple and rapid, with high accuracy and precision.

关 键 词：阴离子交换色谱法 凝血因子Ⅷ 枸橼酸 含量测定 

分 类 号：O657.75[理学—分析化学]