第七批细菌内毒素国家标准品原料制备方法的研究

Research of Preparing Methods for the 7th Bacterical Endotoxin National Standard's Material

目的进行第七批细菌内毒素国家标准品原料制备方法的研究。方法采用4种方法制备内毒素,对得到的内毒素原料进行理化分析后,选取其中最好的制备方法,再采用正交法对其实验条件进行优化。结果最终方法确定为:复苏、培养和收获大肠杆菌;热酚水法粗提;超速离心(4℃,105000×g,4h)纯化;冻干得到原料。结论确立了第七批细菌内毒素国家标准品原料的制备方法。

OBJECTIVE To make research of preparing methods for the 7th bacterical endotoxin national standard＇s material. METHODS Bacterical endotoxin was extracted by four methods. Physical and chemical analysis on material endotoxin was carried out. Among these methods, the optimum method were screened and optimized by orthogonal method on its experiment condition. Then the preparing method was ascertained. RESULTS The optimum method was decided： Escherichia coli O11B4 was cultured and harvested. Crude endotoxin material was extracted by hot phenol-water, after sufficiently dialysis to remove all traces of phenol, the supernatant was purified by ultracentrifugal at 105000×g at 4℃ for 4 h. Final the sediment was frozen and dried to be bacterial endotoxin national standard＇s material. CONCLUSION The preparing method for the 7th bacterial endotoxin national standard＇s material was established. KEY WORDS： endotoxin; bacterical endotoxin national standard; hot phenol-water method