Combined characterization of bovine polyhemoglobin microcapsules by UV–Vis absorption spectroscopy and cyclic voltammetry

Polyhemoglobin produced from pure bovine hemoglobin by reaction with PEG bis(N-succynimidil succinate) as a cross-linking agent was encapsulated in gelatin and dehydrated by freeze–drying. Free carboxyhemoglobin and polyhemoglobin microcapsules were characterized by UV–Vis spectroscopy in the absorption range 450–650 nm and cyclic voltammetry in the voltage range from −0.8 to 0.6 mV to evaluate the ability to break the bond with carbon monoxide and to study the carrier’s affinity for oxygen, respectively. SEM used to observe the shape of cross-linked gelatin-polyhemoglobin microparticles showed a regular distribution of globular shapes, with mean size of ~750 nm, which was ascribed to gelatin. Atomic absorption spectroscopy was also performed to detect iron presence in microparticles. Cyclic voltammetry using an Ag–AgCl electrode highlighted characteristic peaks at around −0.6 mV that were attributed to reversible oxygen bonding with iron in oxy-polyhemoglobin structure. These results suggest this technique as a powerful, direct and alternative method to evaluate the extent of hemoglobin oxygenation.