title: Kinetics of immobilized heparinase in human blood.
abstract: Immobilized enzyme reactors can form the basis of useful blood detoxification systems. One such reactor was developed for heparin neutralization by immobilized heparinase. In this article, reactor kinetics were studied under clinically relevant conditions. Heparin neutralization was assessed in vitro in whole human blood using (a) a well-mixed batch reactor, and (b) an oscillating, continuous-flow reactor. The kinetics of heparin neutralization in human blood were first order over the entire range of heparin and enzyme concentrations and particle fractions tested. The kinetic rate was not sensitive to physiological variations in the concentration of antithrombin, a heparin binding protein in blood. Enzyme activity did not decrease significantly over the 2 hour test period. Kinetic control of the system with minimal intraparticle diffusional limitations was suggested by the Thiele moduli (0.11-0.67) and effectiveness factors (0.98 +/- 0.01). The ratio kcat/Km obtained in batch studies was 0.0028 +/- 0.0008 cm3/microgram-min. A continuous-flow oscillating reactor within a closed recirculation loop performed as a single well mixed batch reactor; there was a short mixing time of recirculating blood when compared to reaction time. A model based on this mixing pattern and the kinetics obtained in independent batch studies accurately predicted heparin neutralization profiles observed in the continuous-flow system.
authors.affiliation: Division of Health Sciences & Technology, Massachusetts Institute of Technology, Cambridge 02139.
authors.forename: L E
authors.initials: LE
authors.lastname: Freed
authors.forename: G V
authors.initials: GV
authors.lastname: Vunjak-Novakovic
authors.forename: H
authors.initials: H
authors.lastname: Bernstein
authors.forename: C L
authors.initials: CL
authors.lastname: Cooney
authors.forename: R
authors.initials: R
authors.lastname: Langer
mesh_terms: D000991:Antithrombins / Q000737:chemistry / Q000493:pharmacokinetics; D004800:Enzymes, Immobilized / Q000493:pharmacokinetics*; D004867:Equipment Design / Q000592:standards; D005069:Evaluation Studies as Topic; D015199:Extracorporeal Membrane Oxygenation / Q000295:instrumentation*; D019763:Heparin Lyase; D006801:Humans; D016018:Least-Squares Analysis; D009991:Oscillometry / Q000295:instrumentation / Q000592:standards*; D011133:Polysaccharide-Lyases / Q000097:blood / Q000737:chemistry / Q000493:pharmacokinetics*; D011237:Predictive Value of Tests; D012212:Rheology; D013997:Time Factors
chemical_list: D000991:Antithrombins; D004800:Enzymes, Immobilized; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase