Kawai M, Yamaguchi N, Nasu M
Graduate School of Pharmaceutical Sciences, Osaka University, Japan.
J Appl Microbiol. 1999 Mar;86(3):496-504. doi: 10.1046/j.1365-2672.1999.00689.x.
Physiologically active bacteria in purified water used in the manufacturing process of pharmaceutical products were enumerated in situ. Bacteria with growth potential were enumerated using the micro-colony technique and direct viable counting (DVC), followed by 24 h of incubation in 100-fold diluted SCDB (Soybean Casein Digest Broth) at 30 degrees C. Respiring and esterase-active bacteria were detected by fluorescent staining with 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) and 6-carboxyfluorescein diacetate (6CFDA), respectively. A large number of bacteria in purified water retained physiological activity, while most could not form colonies on conventional media. The techniques applied in this study enabled bacteria to be counted within 24 h so results could be available within one working day. These rapid and convenient techniques should be useful for the systematic monitoring of bacteria in water used for pharmaceutical manufacturing.
对药品生产过程中使用的纯水中的生理活性细菌进行了原位计数。利用微菌落技术和直接活菌计数（DVC）对具有生长潜力的细菌进行计数，然后在30℃下于100倍稀释的大豆酪蛋白消化肉汤（SCDB）中培养24小时。分别用5-氰基-2,3-二苯基四氮唑氯化物（CTC）和6-羧基荧光素二乙酸酯（6CFDA）进行荧光染色，检测呼吸活性细菌和酯酶活性细菌。纯水中的大量细菌保持着生理活性，而大多数细菌在传统培养基上无法形成菌落。本研究中应用的技术能够在24小时内对细菌进行计数，因此可以在一个工作日内获得结果。这些快速便捷的技术对于系统监测药品生产用水中的细菌应该是有用的。
