顶空气相色谱法测定猪纤维蛋白原中的乙醇残留量

摘要

目的 建立顶空气相色谱法测定猪纤维蛋白原中的乙醇残留量,并对方法进行验证和初步应用.方法 采用Agilent DB-624毛细管柱,以高纯氮气为载气,顶空进样,由氢火焰离子化检测器检测信号,通过内标法计算乙醇含量.确定标准曲线线性范围以及检测限和定量限,并对方法的专属性、准确性、重复性、稳定性、耐用性进行验证.应用该方法对3批猪纤维蛋白原样品中的乙醇残留量进行检测.结果 空白基质、供试品、对照(乙醇)及内标物(正丙醇)在保留时间处相互均无干扰,且对照和内标物峰形良好.标准曲线的线性范围为5.390～80.850 μg/ml.检测限为1.617ug/ml,定量限为5.390 μg/ml.低、中、高3个质量浓度乙醇的总平均加标回收率为(102.84±3.57)％,总相对标准偏差为3.00％;重复检测6次的相对标准偏差为2.17％.对照品和供试品在不同时间点与起始时间的乙醇含量之比为99.74％～105.26％.方法参数微小变动前后测得样品中的乙醇含量之比为94.00％～100.18％.3批猪纤维蛋白原样品中的乙醇残留量分别为35、36、54 μg/ml,均满足中国药典2015年版三部的限量要求(不高于250 μg/ml).结论 本法专属性强,准确性、重复性、稳定性和耐用性均良好,可用于猪纤维蛋白原中的乙醇残留量检测.%Objective To develop a headspace gas chromatography method to determine the residual ethanol in porcine fibrinogen,and to perform method validation and application.Methods Using Agilent DB-624 capillary column with highly pure nitrogen as carrier gas,through headspace sampling,the ethanol content was calculated by internal standard method with signals detected by hydrogen flame ionization detector.The linear range,limit of detection,and limit of quantitation were determined.The specificity,accuracy,repeatability,stability,and durability were verified.The method was applied to measure residual ethanol in 3 porcine fibrinogen batches.Results The blank matrix,test sample,reference (ethanol),and internal standard (n-propanol) had no interference with each other at retention time.The reference and internal standard had good peak shape.The linear range was 5.390-80.850 μg/ml.The limit of detection was 1.617 μg/ml and the limit of quantitation was 5.390 μg/ml.The overall average recovery of low,medium,and high levels was (102.84 ± 3.57)％ and the overall relative standard deviation (RSD) was 3.00％.RSD value of the repeatability was 2.17％.The ethanol content ratios of reference and test sample at different time points to starting point were 99.74％-105.26％.The ethanol content ratios of samples determined by the method before to after minor modification were 94.00％-100.18％.The residual ethanol values (35,36,54 μg/ml) of 3 porcine fibrinogen batches were all well within the limit set by Chinese pharmacopoeia (volume Ⅲ,2015 edition) (≤250 μg/ml).Conclusion This method,with good specificity,accuracy,reproducibility,stability,and durability,is suitable for determination of residual ethanol in porcine fibrinogen.