高效液相色谱法测定缩宫素注射剂含量及其与生物效价测定法比较和应用

目的: 针对缩宫素注射液现行标准中采用生物效价测定法进行含量控制的情况,建立缩宫素注射剂含量的HPLC测定法并进行方法学验证,通过对8个生产企业的414批缩宫素注射剂样品含量的测定,了解HPLC法与生物效价测定法结果的相关性,并对样品长期稳定性进行考察。方法: 采用 Waters XBridge C₁₈(250 mm×4.6 mm,5 μm)色谱柱,以0.1 mol·L^-1磷酸盐缓 冲液作为流动相A,以水-乙腈(1:1)作为流动相B,梯度洗脱,流速1.0 mL·min^-1,检测波长220 nm,柱温40 ℃。结果: 在 本文色谱条件下,缩宫素主峰和杂质峰分离良好;缩宫素浓度在0.36~23.04 IU·mL^-1范围内与峰面积呈良好线性关系,r=0.999 9;平均回收率为99.1%。414批缩宫素注射剂中,缩宫素含量为54.1%~129.5%;HPLC含量测定结果与生物效价测定结果相关性良好,Pearson相关系数为0.683;储存2年后,8批缩宫素注射剂样品HPLC平均含量下降4.5%。结论: 该法专属性强,灵敏、准确,可作为缩宫素注射剂的质量控制方法以替代生物效价测定法。现行标准中生物效价测定法,存在动物个体差异和人为测定误差,难以有效控制药品质量,有必要将生物效价测定法修订为HPLC方法,以稳定控制药品质量。缩宫素注射剂储存2年后,主药可能发生部分降解导致含量降低,需注意储存期。

Objective: To develop a selective gradient liquid chromatographic method for the determination of oxytocin in the context of application of biological titer measurement for quality control according to the current standard of oxytocin injection,and apply the HPLC method to analyze 414 batches of commercial oxytocin injection samples from 8 production enterprises so as to investigate the correlation of HPLC assay and bioassay and to test long term stability. Methods: A Waters XBridge C₁₈ column(250 mm×4.6 mm,5 μm)was adopted.The mobile phases consisted of 0.1 mol·L^-1 dihydrogen phosphate solution(phase A,adjusted to pH 6.0)and water – acetonitrile(phase B,1:1)with gradient elution at a flow rate of 1.0 mL·min^-1.UV detection was performed at 220 nm,and the column temperature was set at 40 ℃. Results: Oxytocin and its related substances were successfully separated by the established method.The method showed good resolution and linearity between peak area and oxytocin concentration at the range of 0.36 -23.04 IU·mL^-1 with r=0.999 9.The average recovery was 99.1%.In 414 batches of commercial oxytocin injection samples,the total content of oxytocin ranged from 54.1% to 129.5%,the results correlated well to the bioassay method with a Pearson correlation coefficient of 0.683.After 2-year storage,the content of 8 batches of oxytocin injection decreased by 4.5%. Conclusion: The method is proved to be specific,sensitive and accurate and can be used to control the quality of oxytocin injection for substitution of bioassay.The research suggests that the bioassay method in the existing standard has deficiencies due to individual differences and error,which needs to be revised to HPLC method to control and stabilize the quality of drugs.The basic drug may have partial degradation after storage of two years,so attention should be paid to the storage period.

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