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目的:考察降纤酶的质量现状及存在问题。方法:对国家食品药品监督管理总局(CFDA)网站数据库上检索到的有降纤酶生产批文的生产企业进行问卷调查,对部分生产企业进行现场调研,针对上述两个环节中发现的问题,对降纤酶原液、中间体、半成品、成品开展了降纤酶原料(液)蛋白质序列分析等一系列实验室探索性研究,提升其安全性、有效性、稳定性和可控性。结果:目前市场上流通的大部分样品的纯度均较低,其中尖吻蝮蛇降纤酶的主要成分为蕲蛇酶,白眉蝮蛇降纤酶的主要成分为NCBI蛋白数据库中编号为gi|143681919|(sp|P85109.1|)的蛋白;降纤酶效价测定影响因素中Tris的浓度影响最大;加入大分子保护剂可以改善样品中的辅料右旋糖酐对效价测定结果的影响。效价测定中两种仪器法(磁珠法和光学法)测定的结果差距较大,均比目测法低,目前尚不适用于降纤酶效价的测定。热稳定性表明注射用降纤酶热稳定性较好,降纤酶注射液的热稳定性较差。本研究采用浓缩一倍的Tris缓冲液稀释纤原,并加入适当的保护剂,建立了降纤酶半成品效价测定的方法。结论:目前降纤酶的质量有待提高。两种蛇毒来源的降纤酶结构、纯度等差别较大,现行的标准项目设置不全面;缺少专属性较强目前的鉴别;异常毒性、效价测定可操作性差等,不能有效地控制降纤酶的质量。有必要将尖吻蝮蛇和白眉蝮蛇来源的降纤酶作为两个品种管理,同时加强降纤酶的标准化研究。
Objective: To investigate the quality of defibrase and its existing problems. Methods: A questionnaire survey was conducted among the manufacturing enterprises with defibrase production approval retrieved from the database of CFDA website. Some production enterprises were investigated on site. According to the problems found in the above two links, Fibrinogen, intermediates, semi-finished products and finished products have carried out a series of laboratory exploratory studies on the protein sequence analysis of defibrase (raw material) to improve their safety, effectiveness, stability and controllability. Results: The purity of most samples circulating in the market was low at present. The main component of Agkistrodon halysii defibrase was Acutobin, the main component of Agkistrodon halysii defibrase was NCI protein database named gi | 143681919 | (sp | P85109.1 |) protein; defibrase titer in the determination of the most influential factors Tris concentration; addition of a large molecule protective agent can improve the sample dextran accessories on the titer results. The determination of titer between the two instrument method (magnetic bead method and optical method) the results of the larger gap, both lower than the visual method, is not currently suitable for the determination of defibrase titer. Thermal stability shows that defibrase for injection has better thermal stability and defibrase is less stable. In this study, the use of concentrated double dilution of Tris buffer fibrillar fibroin, and adding appropriate protective agent, established defibrase semi-finished product titer method. Conclusion: At present, the quality of defibrase is to be improved. Two kinds of snake venom defibrase source from the structure, purity and other large differences, the current standard project settings are not comprehensive; the lack of specificity of the current identification; abnormal toxicity, potency determination operability is poor, can not effectively control the defibrillators Enzyme quality. It is necessary to manage the defibrillators from Agkistrodon acutus and Agkistrodon acutus as two breeds and to standardize defibrase.