中药提取物淫羊藿苷作为骨质疏松症抑制剂的试验研究

Study on the Inhibitor of Osteoporosis by Icariin Extracted from Herbal Medicine

利用组织蛋白酶K降解I型胶原蛋白的特性,从中草药中筛选出组织蛋白酶K选择性抑制剂以用于人与宠物骨质疏松的治疗.试验使用真核酵母表达的人源组织蛋白酶K,用AKTA蛋白纯化系统进行纯化,获得较纯的组织蛋白酶K,再利用荧光酶标仪测出其酶活性,将酶活力单位定义为37℃每10min释放1nmol/LAMC的酶量为一个活力单位,得出组织蛋白酶的活性为每微克蛋白115.82U;用I型胶原蛋白做为组织蛋白酶K的水解底物,从中药淫羊藿中筛选获得了一个组织蛋白酶K抑制剂成分,即淫羊藿苷.淫羊藿苷可以抑制组织蛋白酶对胶原蛋白的降解,从而达到保护骨质吸收的功效,因此,淫羊藿苷可以用于制备预防和治疗由组织蛋白酶K破坏胶原蛋白引起的骨质疏松、关节炎等疾病的预防和治疗药物.

In order to the discovery the mechanism of the cleavage of triple-helical collagens by Cathepsin K （Cat K）, discover Cat K coUagenase inhibitors from Chinese herbal extract, find highly effective, novel and low cost drugs for the treatment of increasingly musculoskeletal diseases such as osteoporosis and arthritis in human and pet animals. First, the recombinant Cat K will be expressed as the proenzyme in Pichia pastoris; Second, use an AKTA protein purification system to purify the precursor of Cat K, and to prepare the high pure enzyme; Third, use a fluorescence microplate reader to measure the protein activity, the enzyme activity unit is defined as the substrates release lnmol/LAMC with 10mins in37℃, The activity of Cathepsin K is 115.82u per microgram protein; Finally, take Type I collagen as the substrate to screen Cat K collagenase inhibitors from Chinese herbal extracts. We obtain an inhibitor, Icraiin. The Icraiin can prevent the degradation of collagen by Cathepsin K, so as to achieve the role of bone protection for osteoporosis.