摘要目的 建立检测黄热病减毒活疫苗中乳糖和山梨糖醇含量的高效液相色谱法(high performance liquid chromatography,HPLC),并验证该法.方法 建立的HPLC的色谱条件确定为:离子排斥色谱柱Aminex?HPX-87H(300 mm×7.8 mm),流动相0.004 mol/L硫酸、流速0.8 ml/min、柱温50℃,进样量20μl.检测该法的系统适用性,并验证该法的线性、准确度、精密度、专属性、稳定性和耐用性.结果 该法的乳糖与山梨糖醇峰间的分离度为6.68,乳糖和山梨糖醇标准曲线的线性良好,线性的决定系数分别为0.99998和0.99987.该法的准确度良好,乳糖和山梨糖醇的回收率分别为100.2％ ～100.4％ 和101.0％ ～101.7％,均符合规定的要求.6次重复检测的乳糖和山梨糖醇结果相对标准偏差分别为0.00％ 和0.08％;2名分析员于不同时间检测12次的乳糖和山梨糖醇结果相对标准偏差分别为0.30％ 和0.99％.该法检测乳糖和山梨糖醇质量浓度的定量限分别为0.05 mg/ml和0.06 mg/ml.结论 建立的HPLC可用于检测黄热病减毒活疫苗中的乳糖和山梨糖醇含量.

abstractsObjective To develop and validate high performance liquid chromatography (HPLC ) for determination of lactose and sorbitol contents in finished products of live attenuated yellow fever vaccine .Methods The chromatographic conditions of developed HPLC were chosen as following :ion exclusion chromatographic column Aminex ? HPX-87H (300 mm × 7 .8 mm) ,mobile phase 0 .004 mol/L sulfuric acid ,flow rate 0 .8 ml/min ,column temperature 50 ℃ ,and sampling volume 20 μl .The system suitability was detected ,and the linearity ,accuracy ,precision ,specificity ,stability and robustness of this method were validated .Results The resolution between lactose and sorbitol peaks was 6 .68 .The standard curves of lactose and sorbitol had good linearity .The linear coefficient of determination was 0 .99998 for lactose and 0 .99987 for sorbitol .This method had good accuracy .The recovery rates of lactose and sorbitol were 100 .2％-100 .4％ and 101 .0％-101 .7％ , respectively , both meeting requirement .The relative standard deviation (RSD) of 6 repeated testing results was 0 .00％ for lactose and 0 .08％ for sorbitol .The RSD of 12 testing results by 2 persons at different time was 0 .30％ for lactose and 0 .99％ for sorbitol .The limit of quantitation of this method was 0 .05 mg/ml for lactose and 0 .06 mg/ml for sorbitol .Conclusion The developed HPLC can be used to detect lactose and sorbitol contents in live attenuated yellow fever vaccine .