分光光度法标定酵母超滤液浓度及其在脑膜炎球菌培养基优化过程中的应用

Optimization of Fermentation Medium for Neisseria meningitidis with Quantified Ultrafiltered Yeast Extract Solution by Spectrophotometry

目的:建立检测酵母超滤液浓度的方法,并探索适合A、C群脑膜炎球菌生长的无动物源性培养基中酵母超滤液的浓度。方法:通过全波长扫描及不同波长吸光度值的比较确定检测酵母提取物浓度的最佳波长;用3000 Da超滤膜超滤酵母提取物溶液,制备酵母超滤液并检测其性质;观察脑膜炎球菌在含不同批次酵母超滤液的半综合培养基中的生长情况,验证不同批次标化的酵母超滤液的稳定性;配制含不同浓度酵母超滤液的半综合液体培养基,观察A、C群脑膜炎球菌的生长情况。结果:通过全波长扫描及不同波长吸光度值的比较,确定了检测酵母提取物浓度的最佳波长为405 nm;采用3000 Da超滤膜超滤后的酵母超滤液不与CTAB产生沉淀,通过检测其D_(405nm)值,可以确定酵母超滤液的浓度;从生长情况考虑,A、C群脑膜炎球菌半综合培养基中最优酵母超滤液浓度分别为4、2 mg/L。结论:建立了制备酵母超滤液及分光光度法标定酵母超滤液浓度的方法,确定了培养A、C群脑膜炎球菌半综合培养基中最优酵母超滤液浓度。

Objective： To develop a method for determination of ultrafiltered yeast extract（UYE） content, and find the UYE in animal-free medium suitable for group A and group C Neisseria meningitidis. Methods： The optimum absorbance wavelength of yeast extract solution was determined by comparing different wavelength absorbance values out of full wavelength scanning. Yeast extracts（YE） were fractionated by uhrafiltration（UF） with 3000 Da molecular weight cutoff membranes to prepare UYE, and then the properties of UYE were tested, the stability of different batches of UYE was validated by observing the growth of N.meningitidis in the semi-synthetic medium containing different batches of UYE. The growth of A group and C group N.meningitidis was observed in the prepared semi-synthetic liquid medium containing different concentrations of UYE. Results： The optimum detection wavelength for YE concentration was 405 nm. There was no precipitant when CTAB was added into the UYE. The UYE could be determined at 405 nm. The best concentration of UYE was 4 g/L for group A N.meningitidis, and 2 g/L for group C N.meningitidis. Conclusion： We have developed the UYE preparation and UYE quantification spectrophotometry method, and also determined the best concentration of UYE in the semi-synthetic medium for group A and group C N.meningitidis.