重组人白介素-15理化对照品的分析鉴定

Evaluation of Recombinant Human Interleukin-15 Reference Standard

目的为了对产品质量进行有效的控制,根据《中国药典》要求,对重组人白介素^(-1)5理化对照品进行分析鉴定。方法参照2010年版《中国药典》三部,测定该样品的生物学活性、蛋白含量、纯度、等电点等指标;通过测定N-末端氨基酸序列、质谱相对分子质量与液质肽图,确证其一级结构。结果该样品的比活性为1.03×107IU·mg^(-1);蛋白含量为(0.879±0.065)mg·m L^(-1);非还原SDS-PAGE纯度为100.0%;分子排阻色谱纯度为100.0%;等电点测定结果为5.2;以上结果均符合质量标准要求。实测质谱相对分子质量(12 900.80)与理论值(12 900.71)一致;氨基酸序列与理论一致,氨基酸覆盖率100%;二硫键连接方式为Cys36-Cys86、Cys43-Cys89,与文献报道相符。结论该理化对照品质量合格、氨基酸序列正确,可用于白介素^(-1)5的常规质量控制。

OBJECTIVE To evaluate the reference standard of recombinant human interleukin-15 （ IL-15 ） for the effective quality control of IL-15 products according to the requirements of Chinese Pharmacopeia. METHODS The biological activity, concentration, purity, and isoelectric point of IL-15 were tested according to Chinese Pharmacopeia （volume m, 2010 edition）. The primary structure was confirmed by analyzing the N-terminal amino acid sequence and relative molecular mass and peptide mass mapping. RE- SULTS The measured biological activity of IL-15 was 1.03×107IU·mg-1, the content was （0.879±0.065）mg·m L-1 , the pu- rities tested by SDS-PAGE and SEC-HPLC were all 100% , and the isoelectric point was 5.2, which all met the criteria specified in the quality standard. The observed relative molecular mass, 12 900. 80, was consistent with theoretical value （ 12 900. 71 ）. Its amino acid sequence was verified with coverage of 100%. The disulfide bonds were identified to to be Cys36-Cys86/CyM3-Cys89, which was in accordance with the published papers. CONCLUSION This reference standard, which is qualified and has correct amino acid sequence, can be used for the routine quality control of IL-15.