An immobilized microbial heparinase for blood deheparinization

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 complications. 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 development 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 purification 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 successful testing of a fluidized bed reactor containing immobilized heparinase in the removal of clinically used quantities of heparin from both human blood in vitro and canine 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.