An immobilized microbial heparinase for blood deheparinization

Abstract

A new medical application of an immobilized microbial enzyme is described. Extracorporeal devices require systemic heparin administration to prevent thrombus formation; however, the use of heparin often leads to serious hemorrhagic complica tions. Heparinase isolated from Flavobacterium has been immobilized and used in a fluidized bed reactor to eliminate heparin from blood passing through an extracorporeal circuit both in vitro and in vivo. This paper discusses the stepwise de velopment of this heparinase reactor including: (1) improvements in the fermentation resulting in an inexpensive large-scale source of heparinase without the addition of the previously required inducer, heparin; (2) the use of batch processes to adapt previous purification schemes to large-scale heparinase production and the subsequent purifica tion of heparinase to a single SDS-PAGE banding protein; (3) the immobilization of heparinase with a 91% activity recovery and good stability, (4) the design and suc cessful testing of a fluidized bed reactor containing immobilized heparinase in the re moval of clinically used quantities of heparin from both human blood in vitro and ca nine blood in vivo; and (5) the initiation of animal studies focusing on the toxicology of heparinase-derived heparin degradation products and the short and long term effects of exposure to these products and to heparinase.