Quantitative PCR and Disaccharide Profiling to Characterize the Animal Origin of Low-Molecular-Weight Heparins
Quantitative PCR and Disaccharide Profiling to Characterize the Animal Origin of Low-Molecular-Weight Heparins
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DOI：
10.1177/1076029608320831
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发表时间：
2009-02-01
影响因子：
2.9
通讯作者：
Anger, Pascal
中科院分区：
文献类型：
--
作者：
Houiste, Celine;Auguste, Cecile;Anger, Pascal
低分子量肝素（LMWH）广泛用于治疗血栓形成和急性冠脉综合征。它们是通过猪肠肝素的酶解或化学解聚而获得的。依诺肝素钠是一种广泛使用的低分子肝素，具有独特且可重复的寡糖特征，这是由起始材料的来源和严格控制的制造工艺决定的。尽管确实存在其他依诺肝素样 LMWH，但尚未确定具体的释放标准，包括用于生产的粗肝素的来源。定量聚合酶链式反应方法已开发出来，可确保猪源粗肝素的纯度，牛的 DNA 检测限低至 1 ppm，羊污染物的 DNA 检测限低至 10 ppm。该方法通常用作依诺肝素钠生产过程中的放行验收标准。此外，当该过程去除 DNA 时，可以使用其他分析技术来评估任何污染。彻底解聚后的二糖分析可以确定至少 10% 牛或 20% 羊材料的存在；多变量分析对于执行数据分析很有用。与新技术的可用性一致，这些方法应该被要求作为用于生产 LMWH 的粗肝素的验收标准，以确保其安全性、质量和免疫学特征。
Low-molecular-weight heparins (LMWHs) are widely used in the management of thrombosis and acute coronary syndromes. They are obtained by the enzymatic or chemical depolymerization of porcine intestinal heparin. Enoxaparin sodium, a widely used LMWH, has a unique and reproducible oligosaccharide profile which is determined by the origin of the starting material and a tightly controlled manufacturing process. Although other enoxaparin-like LMWHs do exist, specific release criteria including the origin of the crude heparin utilized for their production, have not been established. A quantitative polymerase chain reaction method has been developed to ensure the purity of the porcine origin of crude heparin, with a DNA detection limit as low as 1 ppm for bovine, or 10 ppm for ovine contaminants. This method is routinely used as the release acceptance criterion during enoxaparin sodium manufacturing. Furthermore, when the process removes DNA, other analytical techniques can be used to assess any contamination. Disaccharide profiling after exhaustive depolymerization can determine the presence of at least 10% bovine or 20% ovine material; multivariate analysis is useful to perform the data analysis. Consistent with the availability of newer technology, these methods should be required as acceptance criteria for crude heparins used in the manufacture of LMWHs to ensure their safety, quality, and immunologic profile.