Preliminary Study On The Extraction And Modification Porcine Hemoglobin

Stroma-free porcine hemoglobin is separated and purified from fresh pig blood by centrifugation and filtration with 0.45 um to 0.22um films. SDS-PAGE analysis of purified porcine hemoglobin shows only one band. It is proved that the dissociation of porcine hemoglobin from tetramer to dimer is taken place. We have determined the oxygen pressure corresponding to the half-saturation of stroma free hemoglobin (P50) and the Hill coefficient (n), and also investigated the dependence of allosteric cofactor(lHP) upon oxygen affinity of pHb.The pHb modified by DBBF shows the special character to resist the dissociation of tetramer in comparison with nature pHb. According to the analysis in SDS-PAGE, the DBBF modified derivatives are intramolecular crosslinked protein. DBBF is utilized to modify oxyhemoglobin and deoxyhemoglobin respectively. The derivatives show different results. We also investigated the affection of the mass of modifier and the modified time to pHb modified by DBBF. The oxygen affinity of pHb modified by DBBF is also affected by allosteric cofactor(IHP).Increasing the total molecular weight of hemoglobin through PEG coupling has been proved to be a good choice for prolonging vascular retention time. As a long, liner, hydrophilic molecule, PEG exerts significant influence on the oxygen-carrying properties of pHb conjugates. PEG-hemoglobin's conjugates show a wide range of different properties that are attributed to the size of PEGs amounts of bound PEGs, and the presence of allosteric cofactors. Combination of three procedures , DBBF intramolecular crosslinking, allosteric cofactors treating and activated PEG conjugating, result in a stable tetramer pHb derivative with a large molecular weight and well oxygen-carrying capacity.