用BOC－Leu－Ser－Thr－Arg－pNA建立前激肽释放酶活性测定法

BOC-Leu-Ser-Thr-Arg-pNA,A Substrate Suitable for the Functional Determination of Human Plasma Prekallikrein

作者发现发色底物ＢＯＣ－Ｌｅｕ－Ｓｅｒ－Ｔｈｒ－Ａｒｇ－ｐＮＡ对激肽释放酶具有良好的敏感性和特异性。它受激肽释放酶作用的Ｋｍ和Ｖｍａｘ。分别为２３５ｐｍｏｌ·Ｌ－１和３３７ｎｍｏｌ（ＰＮＡ）·Ｓ－１·Ｕ－１（ｋＫ）；抑制试验显示，该底物对ＡＴ－Ⅲ和ＬＢＴＩ不敏感，用作激肽释放酶测定的回收率为９７．２％～１０２．ｌ％，ＣＶ值２．３％～４．６％。用它建立了血浆激肽释放酶活性测定法，标准曲线线性甚佳，线性范围６．２５％～３００．０％（激肽释放酶活性）．测得健康人、妊娠妇女、肝功能衰竭病人和晚期癌症病人的血浆前激肽释放酶活性分别为１１３．９％，１４５．７％，４０．０％和９０．０％。作者认为，发色底物ＢＯＣ－Ｌｅｒ－Ｓｅｒ－Ｔｈｒ－Ａｒｇ－ｐＮＡ是ＰＫ（ＫＫ）活性测定的良好底物，建立的ＰＫ活性测定法简便、快速、敏感、可靠，实用价值大。

OC-Leu-Ser-Thr-Arg-pNA, an intended substrate for protein C, was identified as asubstrate of kallikrein which is suitable for the determination of the human plasmaprekallikrein activity. The enzyme's Kmand Vmax of this substrate were calculated to be235μmol / L and 337 nmols(PNA-released) per second per unit of kallikrein respectively,which were proved to be superior to those of ChromozymPk, a well-known substrate ofkallikrein。 The amidolysis of the kaolin activated pooled human normal plasma toward it wasindepedndnt of coagulatory factor Ⅱ,Ⅹ,Ⅸ, protein C except prekallikrein, but was depen-dent on certain extent affected by factor Ⅻ.The splitting of this substrate by kallikrein wasinhibited slightly by limabean trypsin inhibitor and even less by antithrombin-Ⅲ. With thissubstrate, the recovery rates of kallikrein and kaolin activated plasma of near to l00% wereyielded and the low CV values ranging from 2.3% to .6% of intra-and inter-assay were ob-tained. Subsequently, a sensitive, specific, reliable and simple method of the functional humanplasma prekallikrein was developed by using this substrate。