Physicochemical studies of fractionated bovine heparin: I. Some dilute solution properties

Abstract

Commercially prepared bovine heparin was fractionated by sequential elution from an ECTEOLA column into four discrete fractions and by solvent elution with an ethanol-water mixture into 12 fractions. Equilibrium centrifugation of the fractions in 0.5 m NaCl at pH 2.5 and interference optics revealed wide distribution of molecular weights. The fractions varied in biological activity and intrinsic viscosity. Chemical differences between fractions were minimal while the biological activity for the low molecular weight fractions were uniformly low. Inactivation with HCl, under mild conditions where biological activity was reduced fivefold, was not accompanied by reduction in intrinsic viscosity or molecular weight. A linear intrinsic viscosity-molecular weight relationship for the fractions prepared by alcohol elution can be described by the functions [ η] = 1.75 × 10 −5 M w 0.98 in 0.1 m NaCl, pH 6.5, and [ η] = 7.85 × 10 −5 M w 0.80 in 0.5 m NaCl, pH 2.5. An analogous relation for the sedimentation constant determined in 0.5 m NaCl at pH 2.5 was linear and described by s 0 = 1.2 × 10 −1 M w 0.44. The data suggest that heparin in solution under the conditions studied exhibits substantial flexibility.