This work examines the physical properties of bovine hemoglobin (BHb) chemically cross-linked to a pH responsive polymer (poly(acrylamide)) with the goal of taking advantage of the polymer's pH sensitivity to generate low-P-50 oxygen carriers for application in physiological conditions characterized by deviations from normal pH. BHb-hydrogel-based oxygen carriers encapsulating 10-16 g/dL BHb displayed P(50)s < 10 mmHg when encapsulated inside of liposomes (i.e. Hb-LGs) and < 15 mmHg in the absence of a lipid bilayer (i.e. Hb-NHPs), when synthesized at pHs less than normal physiological pH. The results of this work suggest that synthesis of Hb-LGs/Hb-NHPs under different pH conditions affect the ionization of BHb and the flexibility of the polymer chains to which BHb is chemically cross-linked. The degree of BHb cross-linking to the polymer matrix and the chain flexibility of the polymer influences the oxygen affinity and cooperativity of the oxygen carrier by influencing how easy it is for the Hb molecule to change conformations between the R and T states. The magnitude of the zeta potential of Hb-LGs and Hb-NHPs was shown to be within the range of stored red blood cells and within the range of limited flocculation. Taken together, this work describes the preparation and characterization of oxygen carriers with increased oxygen affinities compared to those of red blood cells.
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