双滤膜过滤方法的建立及在卡波姆眼用凝胶无菌检查试验中的应用

目的：建立卡波姆眼用制剂的无菌检查方法，探索双滤膜过滤方法运用于药品无菌检查试验的可行性。方法：用0.1%的蛋白胨水溶液稀释供试品，在供试品溶液中加入适量的10%氯化钙溶液，使卡波姆凝胶生成沉淀。采用孔径30~50 μm和不大于0.45 μm 2种无菌滤膜过滤供试品溶液，分别截留沉淀物和微生物，对2种滤膜进行适量冲洗后，置适宜的培养基中进行培养。结果：6种验证用菌株，枯草芽孢杆菌、大肠埃希菌、金黄色葡萄球菌、生孢梭菌、白色念珠菌、黑曲霉在接种供试品的培养基中生长良好。采用双滤膜过滤的方法，可以有效削除供试品的抑菌活性，避免卡波姆凝胶堵塞滤膜，实现对容器中全部内容物进行检验，最大程度降低实验过程对此类特殊供试品中微生物的损伤或损失。结论：双滤膜过滤方法实验过程符合中国药典2010年版无菌检查法相关规定，试验结果有效，可用于卡波姆眼用制剂的无菌检查试验。

Objective: To establish a sterility test method for carbomer eye drops and explore the feasibility of two-step membrane filtration method in sterility test for drugs. Methods: Samples were first diluted with 0.1% peptone solution,and then the carbomer gel was precipitated by adding moderate portions of 10% calcium chloride solution.Precipitation and microorganisms were intercepted by two sterile membrane filters,30-50 μm and ≤ 0.45 μm in pore size,respectively,and then the two membrane filters were moderately washed and incubated in suitable media for at least 14 d. Results: Clearly visible growth of the six test microorganisms,Bacillus subtilis,Escherichia coli,Staphylococcus aureus,Clostridium sporogenes,Candida albicans,Aspergillus niger was obtained.Antibacterial activity of the samples was effectively eliminated by using the two membranes filtration method,carbomer gel blockage was avoided,detection of all the contents was achieved,and influence of microorganisms in special samples was reduced to a great extent. Conclusion: The two-step membrane filtration method meets the sterility test requirements stipulated by the Chinese pharmacopoeia (2010 edition);the test results are valid and the method can be used for sterility test of carbomer eye drops.