Quantitation of impurities in biological products

总结：本项目的目的是开发和验证生物制品中可能影响产品安全性或有效性的化学杂质和残留生产制剂的方法。 其中包括：1）在疫苗中用作灭活剂或结合剂的残留戊二醛，2）花粉变应原提取物和变应原源材料中的残留四氯乙烯，3）粗变应原提取物和变应原斑贴试验材料的非活性成分，以及4）免疫球蛋白产品中的抗病毒剂inactine。 目前正在进行的工作或过去一年进行的工作包括： 第一章 花粉源材料变应原提取物加工中使用的四氯乙烯测定方法的开发和验证。开发并验证了一种用于过敏性提取物中四氯乙烯的吹扫捕集气相色谱法，并获得了最终产品代表性采样的数据。 通过气相色谱/质谱法测定过敏源材料中的同一化合物正在验证过程中，已收集了花粉源材料代表性采样的中期数据。 （二） 建立了一种测定戊二醛的高效液相色谱法，包括对硝基苄基羟胺柱前衍生和紫外检测，验证研究正在进行中。 3)对免疫球蛋白中Inactine的HPLC离子交换柱后衍生荧光法进行了改进。 为了使制造商的程序合格，进行了长时间的尝试，该程序需要进行大量修改，才能在所需的十亿分之一的限度下成功检测。 4)最近获得的离子阱HPLC/质谱仪将与气相色谱/质谱联用沿着用于开发各种产品中杂质筛选以及过敏原提取物和过敏原斑贴试验产品中低分子量化合物表征的通用程序。

Summary: The objectives of this project are to develop and validate methodologies for chemical impurities and residual manufacturing agents in biological products whose presence may affect product safety or efficacy. These include: 1) residual glutaraldehyde used as inactivating or conjugating agent in vaccines, 2) residual tetrachloroethylene in pollen allergenic extracts and allergenic source materials, 3) inactive components of crude allergenic extracts and allergen patch test materials and 4) an antiviral agent, inactine, in immune globulin products. Current work in progress, or that performed in the past year includes: 1) The development and validation of methods for the determination of tetrachloroethylene, used in the processing of pollen source materials for allergenic extracts. A purge-and-trap gas chromatographic method for tetrachloroethylene in allergenic extracts has been developed and validated, data has been obtained for a representative sampling of final products. The determination of this same compound in allergenic source materials by gas chromatography/mass spectrometry is in the process of validation, interim data on a representative sampling of pollen source materials have been collected. 2) An HPLC method for the determination of glutaraldehyde, involving pre-column derivatization with p-nitrobenzyl-hydroxylamine and UV detection has been developed, validatation studies are in progress. 3) An HPLC ion-exchange and post-column derivatization/fluourescence procedure for the determination of inactine in immune globulins was transfered and modified. A lengthy attempt was made to qualify a manufacturer's procedure, which required extensive modification before successful detection at the required limit of 1 part-per-billion was accomplished. 4) A recently acquired ion-trap HPLC/mass spectrometer will be used along with gas chromatography/mass spectrometry to develop a general procedure for impurities screening in a variety of products as well as the characterization of low-molecular weight compounds in allergenic extracts and allergen patch test products.