目的： 建立复方牛胎肝提取物片高效液相色谱（HPLC）指纹图谱,结合L-02细胞/CCK-8测定法,为复方牛胎肝提取物片的质量评价提供一种新方法。方法： 选择3家生产企业生产的9批复方牛胎肝提取物片样品。采用 HPLC 法,以 0.1% 甲酸为流动相 A,0.1% 甲酸甲醇溶液为流动相 B,进行梯度洗脱;流速 1.0 mL·min-1;检测波长 260 nm;柱温30℃;进样量 20 μL。用中药色谱指纹图谱相似度评价系统（2012 版）建立复方牛胎肝提取物片的指纹图谱,进行相似度分析,并用L-02 细胞 /CCK-8 测定法对产品进行活性测定。结果： 建立了复方牛胎肝提取物片的 HPLC 指纹图谱,相似度计算结果在 0.963~1.000 范围内,3 家企业生产工艺稳定,同企业产品批间一致性良好。L-02 细胞 /CCK-8 测定法检测 3 家企业 9 批复方牛胎肝提取物片的活性,产品对细胞的修复率在 49%~147%范围内,不同企业产品活性存在较大差异。结论： 所建立的指纹图谱方法专属性强,高效快速,可用于考察复方牛胎肝提取物片的批间一致性。采用 L-02细胞/CCK-8测定法测定产品对细胞的修复率,明显提升了指标的科学性,改善现标准的指标钝化的现状,为复方牛胎肝提取物片的质量评价提供参考。
Objective： To establish an HPLC fingerprint of compound embryonic bovine liver extract tablets, and to provide a new method for the quality control of compound embryonic bovine liver extract tablets using L-02 cells/CCK-8 method. Methods： A total of 9 batches of compound embryonic bovine liver extract tablets supplied by 3 pharmaceutical manufacturers were applied in this study. HPLC analysis was performed with 0.1% formic acid solution（A）and 0.1% formic acid methanol solution（B）as the mobile phase at a flow rate of 1.0 mL·min-1. The detection wavelength was 260 nm. The column temperature was 30 ℃ . The injection volume was 20 μL. The fingerprint common pattern and similarity calculation were established using traditional Chinese medicine chromatographic fingerprint similarity evaluation system（version 2012）. The L-02 cells/CCK-8 method was established to perform activity determination analysis. Results： An HPLC fingerprint of compound embryonic bovine liver extract tablets was established. The similarities ranged from 0.963 to 1.000. The production processes of these 3 pharmaceutical manufacturers were stable. The results indicated components in each batch of compound embryonic bovine liver extract tablets were consistent for each manufacturer. However,there were differences among the 9 different batches in the activities as determined by L-02 cells/CCK-8 method. As showed in the results,the cell repair rate were from 49% to 147%. There were significant differences in productions in the 3 pharmaceutical manufacturers. Conclusion： The established fingerprint method is specific,efficient and rapid,which could be used as a reference for the inter batch consistency of compound embryonic bovine liver extract tablets. The L-02 cells/CCK-8 method used to measure the cell repair rate significantly improved the scientificity and index passivation,which provided a reference for the quality evaluation of compound embryonic bovine liver extract tablets.
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