重组人甲状旁腺激素(1-84)的质量研究

Quality research of the recombinant human parathyroid hormone(1-84)

目的:对重组人甲状旁腺激素rh-PTH(1-84)进行质量研究,为其生产工艺的进一步优化提供依据。方法:采用RP-HPLC和非还原型SDS-PAGE测定纯度;还原型SDS-PAGE测定相对分子质量;Western blot鉴定并进行N-末端氨基酸序列测定;考马斯亮兰法测定蛋白含量;ELISA法测定效价。结果:rh-PTH(1-84)经RP-HPLC和SDS-PAGE分析,其纯度分别为81%,85%。其相对分子质量为9480,N-末端15个氨基酸序列为SVSEIQLMHNLGKHL,均与理论值一致。蛋白含量为标示量的87%,效价在标示量的80%～120%。结论:重组人甲状旁腺激素rh-PTH(1-84)质量基本符合规定,但其纯度偏低,生产工艺有待进一步优化。

Objective： The quality research of recombinant human parathyroid hormone rh - PTH （ 1 - 84） was con- ducted, which could help improve the manufacture process of rh-PTH （1 - 84）. Methods： The purity of rh - PTH （ 1 - 84） was determined by RP - HPLC and non - reduced SDS - PAGE methods. The relative molecular mass was determined by reduced SDS - PAGE. Western blot and N - terminal sequence of the rh - PTH （ 1 - 84） were ana- lyzed. The protein content was assayed by bradford method, and the potency was tested by ELISA. Results：The puri- ty of the rh- PTH（ 1 -84） was 81% by RP- HPLC and 85% by SDS -PAGE,with the relative molecular mass 9480. The determined N - terminal sequence was SVSEIQLMHNLGKHL, which was identical to the theoretical se- quence. The protein content and potency were 87% and 80% -120% respectively compared to the labeled content. Conclusion ：The quality of the rh -PTH（ 1 -84 ） was consistent with the quality standard ,but the purity was some- what low, so the manufacture process was recommended improvement further.