Covalently bound conjugates of albumin and heparin: Synthesis, fractionation and characterization

Abstract

Covalently bound conjugates of human serum albumin and heparin were prepared as compounds which could improve the blood-compatibility of polymer surfaces either by preadsorption or by covalent coupling of the conjugates onto blood contacting surfaces. The conjugates (10–16 weight % of heparin) were obtained by a condensation reaction between albumin and heparin using 1-ethyl-3-(dimethylaminopropyl)-carbodiimide. Unreacted albumin and heparin were removed by diethylaminoethyl (DEAE)-cellulose and Cibacron Blue Sepharose chromatography respectively. The activity of the heparin component incorporated in the albumin-heparin conjugates (Ac) was compared with that of the heparin used for the synthesis of the conjugates (Anat) by thrombin time, inhibition of Factor Xa and the activated partial thromboplastin time (APTT) assays. The Ac/Anat ratio for the above assays was as follows': Thrombin time 1.25, Factor Xa inhibition 0.5. and APTT 0.5. Gel filtration chromatography showed broad-molecular weight distributions. The conjugates were fractionated using immobilized antithrombin III (ATIII). High ATIII and low ATIII affinity conjugate fractions showed the same behavior as ATIII fractionated heparin with respect to thrombin times and Factor Xa inhibition.