不同配方的冻干保护剂对人凝血因子Ⅷ制品干热法灭活猪细小病毒效果的影响  

Effect of formula of stabilizer on inactivation of porcine parvovirus in human coagulation factor Ⅷ by terminal dry heat treatment

作　　者：李策生[1] 邹莉[1] 李陶敬[1] 胡勇[1] 邢延涛[1] 彭焱[1] 周雁翔[1] 李娟[1] 

机构地区：[1]武汉生物制品研究所有限责任公司血液制剂研究室,湖北武汉430060

出　　处：《中国生物制品学杂志》2013年第12期1772-1775,共4页Chinese Journal of Biologicals

基　　金：国家863计划(2012AA021904)

摘　　要：目的研究不同配方的冻干保护剂对人凝血因子Ⅷ制品干热法灭活非脂包膜指示病毒猪细小病毒(porcine parvovirus,PPV)效果的影响。方法取人凝血因子Ⅷ原液,分别加入A、B、C 3种配方(氯化钠浓度为C<A<B)缓冲液,经超滤透析制备人凝血因子Ⅷ半成品后,加入>6 LgTCID50/ml的指示病毒PPV,进行冻干、干热法灭活(100℃30 min),检测残余PPV滴度;用筛选出的最佳配方制备3批人凝血因子Ⅷ半成品,进行冻干和干热法灭活,检测残余PPV滴度,验证PPV灭活效果;按照《中国药典》三部(2010版)人凝血因子Ⅷ的成品检测要求对人凝血因子Ⅷ效价、外观、水分、复溶时间和复溶后外观进行检测。结果在100℃30 min条件下,可有效灭活配方A制备的人凝血因子Ⅷ半成品中的指示病毒PPV,确定配方A为最佳冻干保护剂配方;3批凝血因子Ⅷ经干热灭活后,可使残余PPV滴度下降(4.08±0.02)LgTCID50/0.1 ml,且灭活过程对人凝血因子Ⅷ的生物学活性无明显影响,步骤活性回收率为(91.1±2)%。结论已成功研制出1种冻干保护剂配方,应用干热灭活法对人凝血因子Ⅷ制品中非脂包膜病毒PPV具有较好的灭活效果,该方法与有机溶剂/表面活性剂(solvent/detergent,S/D)法联合去除/灭活病毒,可有效提高人凝血因子Ⅷ产品的安全性。Objective To investigate the effect stabilizer of various formula on inactivation of porcine parvovirus （PPV） in human coagulation factor Ⅷ （FⅧ） by dry heat sterilization. Methods Bulks of FVI were subjected to ultrafihraion and dialysis using buffers of formula A, B and C （sodium chloride concentration： C 〈 A 〈 B）. The obtained final bulks were added with PPV as an indicator, at a titer of more than 6 LgTCID50/ml, then lyophilized, inactivated by terminal dry heat treatment, and determined for residual PPV titer to evaluate the inactivation effect. The final product of FⅧ was observed for appearance and determined for titer, moisture, reconstitution time and appearance after reconstitution according to the requirements in Chinese Pharmacopoeia （Volume Ⅲ, 2010 edition）. Results The PPV in FⅧ containing stabilizer of formula A was inactivated effectively by heating at 100 ℃ for 30 min, indicating formula A as the optimal formula. The residual PPV titers in three batches of FⅧ decreased by （4. 08 ± 0. 02） LgTCID50/0. 1 ml after dry heat sterilization. The inactivation process showed no significant influence on biological activity of FⅧ, expressed as a recovery rate of （91. 1± 2 ）%. Conclusion A fo-nula of stabilizer was successfully developed, with which the PPV in FVl was effectively inactivated by terminal dry heat treatment. The method combined with solvent/detergent（S/D） treatment may enhance the safety of FⅧ by removal / inactivation of viruses.

关 键 词：冷冻干燥 人凝血因子Ⅷ 猪细小病毒 灭活 

分 类 号：R554.1[医药卫生—血液循环系统疾病] R392.32[医药卫生—内科学]