【作者】 王艳萍；

【导师】 洪敏；

【作者基本信息】 吉林大学 ， 生物化学与分子生物学， 2006， 硕士

【摘要】 本实验研究课题组经过数年的研究,从我国沿海海洋生物中提取了一种新的蛋白酶,称为N-V蛋白酶。随着药理、药代和酶学特征等研究的展开,该酶制剂有可能应用于临床,因此,迫切需要对这一新的蛋白酶的质量标准进行制定。本研究报告了N-V蛋白酶制剂的部分质量标准。本文应用高效液相色谱法进行N-V蛋白酶的液相分析研究,先后使用了C18、C4反相柱和凝胶过滤柱进行分析。检测波长为214nm时,凝胶过滤分析得到了三个峰,分离度较好。用PAGE电泳的方法检测这三个峰,并切割两个主要条带,胶内用胰酶消化后,用质谱分析发现肽指纹图谱中有近50%的片段相同。用凝胶过滤柱分析N-V酶的方法快速、高效,可作为N-V蛋白酶制剂生产的检测标准。以往的研究发现,该酶的生物来源体内存在一种毒素,即沙蚕毒素。本文采用了薄层层析法检测,检测限为0.5μg/mL,在制剂中未检测到该毒素,方法快速稳定,检测限较好,可作为生产检测中该相关物质的检测标准。更多还原

【Abstract】 Cardial and cerebral vascular thrombosis disease is a threat against human’s health in modern time, it will lead to sudden death and nerves functional disturbance. The principal method of clinical treatment is to cut down fibrinogen in the blood and to dissolve internal clot at earlier period as soon as possible. It is the important topic to study and research this kind of drug in the medicine domain. N-V protease is a kind of protease abstracted from some marine life, N-V protease have the function of hydrolyzing plasminogen and dissolving internal clot. Using high performance liquid chromatography method to detect protease, and using high performance thin-layer chromatography method to detect the related substances NTX.1 Reverse phase high performance liquid chromatography (RP-HPLC)Use the Agilent poroshell 300SB-C18 (21×75mm) which suited for reversed-phase separation of protein and other biomacromolecules produce by the Agilent company recently. The retention time of the main peak is 0.42min and the response value is 0.020AU. The retention time is short他that is concern with the length of the column is 75mm.This columns length is half of common. It is also have been find out that, retention time did not alter obviously with the increasing of organic phase in the mobile phase, we can conclude that the main peak is not a protein peak. The elution time is更多还原