Bovine Hemoglobin Anaerobically Reacted with Divinyl Sulfone: a Potential Source for Hypothermic Oxygen Carriers

Abstract

The bifunctional reagent divinyl sulfone was anaerobically reacted with bovine hemoglobin to give a noncrosslinked intramolecularly-modified new derivative (HbBv-DVS). By employing a high molar ratio of divinyl sulfone to HbBv-DVS, it was possible to effect anaerobic intermolecular crosslinkage. The polymerized material (Poly HbBv-DVS) was shown to consist of a mixture of modified intermolecularly-crosslinked hemoglobins characterized by molecular masses ranging from 130 to-500 kDa. Some functional properties of HbBv-DVS and Poly HbBv-DVS have been evaluated in vitro and in vivo. Viscosities of HbBv-DVS solutions at temperatures as low as 15°C and concentrations up to 14.0 g/dl were proved to be much lower than that of normal human blood at 37°C (-4 cp). Poly HbBv-DVS (14.0 g/dl) was iso-oncotic (COP=23 mm Hg) with plasma, and had viscosities of 3.37 and 4.57 cp at 37 and 15°C, respectively. The clearance of Poly HbBv-DVS from the circulation was significantly delayed (T_1/2=270 min), compared with those of HbBv and HbBv-DVS (T_1/2=80 and 100 min, respectively). The P₅₀ values were substantially increased (P₅₀=52 and 61 mm Hg at 37°C, 0.15 M Cl and pH 7.4 for HbBv-DVS and Poly HbBv-DVS, respectively). Due to their right-shifted oxygen equilibrium curves, these derivatives still yielded P₅₀ values of about 20 mm Hg at the low temperature of 15°C, as compared with only 7 mm Hg for native bovine hemoglobin. These properties make HbBv-DVS and Poly HbBv-DVS potential new candidates for low-temperature organ perfusion.