Analysis of crude heparin by 1H NMR, capillary electrophoresis, and strong-anion-exchange-HPLC for contamination by over sulfated chondroitin sulfate
Analysis of crude heparin by 1H NMR, capillary electrophoresis, and strong-anion-exchange-HPLC for contamination by over sulfated chondroitin sulfate
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DOI：
10.1016/j.jpba.2009.10.017
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发表时间：
2010-03-11
影响因子：
3.4
通讯作者：
Buhse, Lucinda F.
中科院分区：
文献类型：
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作者：
Keire, David A.;Trehy, Michael L.;Buhse, Lucinda F.
我们之前发表了一种强阴离子交换-高效液相色谱（SAX-HPLC）方法，用于检测肝素钠活性药物成分（API）中硫酸软骨素（OSCS）的污染物。虽然原料药已被处理以去除杂质，但粗肝素含有不溶性物质、硫酸软骨素、硫酸乙酰肝素和可能干扰OSCS回收和测量的蛋白质。我们研究了500 MHz H-1 NMR、毛细管电泳（CE）和SAX-HPLC，以定量肝素粗品中的OSCS。使用我们的标准API方案对OSCS加标的粗肝素样品进行分析;我们观察到NMR方法的LOD和LOQ重量百分比分别为0.1%和0.3%，而SAX-HPLC方法的LOD和LOQ重量百分比分别为0.03%和0.09%。CE数据不适用于粗肝素中OSCS的定量测量。我们开发了一种改进的HPLC样品制备方案，使用用2.5 M NaCl溶液以100 mg/mL水平溶解的粗品。该SAX-HPLC方法得到的重量百分比LOD为0.02%，LOQ为0.07%，并且具有比API所用方案更好的性能特征。由爱思唯尔公司出版
We previously published a strong-anion-exchange-high performance liquid chromatography (SAX-HPLC) method for the detection of the contaminant over sulfated chondroitin sulfate (OSCS) in heparin sodium active pharmaceutical ingredient (API). While APIs have been processed to remove impurities, crude heparins contain insoluble material, chondroitin sulfates, heparan sulfate, and proteins that may interfere with the recovery and measurement of OSCS. We examined 500 MHz H-1 NMR, capillary electrophoresis (CE), and SAX-HPLC to quantify OSCS in crude heparin. Using our standard API protocol on OSCS spiked crude heparin samples; we observed a weight percent LOD and LOQ for the NMR approach of 0.1% and 0.3%, respectively, while the SAX-HPLC method gave values of 0.03% and 0.09%, respectively. CE data was not amenable to quantitative measurement of OSCS in crude heparin. We developed a modified HPLC sample preparation protocol using crude dissolved at the 100 mg/mL level with a 2.5 M NaCl solution. This SAX-HPLC approach gave a weight percent LOD of 0.02% and a LOQ of 0.07% and had better performance characteristics than that of the protocol used for APIs. Published by Elsevier B.V.