The purposes of this study were to determine: 1) how changes in oxygen affinity induced by different chemical modifications of hemoglobin affect rates of autooxidation and 2) the degree to which mono- or bivalent crosslinkers prevent precipitation of oxidation products. We studied human stroma-free Hb bivalently crosslinked between alpha globin chains (ααHb) or between beta chains (ββHb) with bis(3,5-dibromosalicyl)fumarate and between beta chains with 2-nor-2-formylpyridoxal 5′-phosphate (HbNFPLP). We also studied the 64KD fractions of Hb reacted with a crosslinker derived from peroxidate oxidized raffinose under oxy- (Hb[R]) or deoxy- (Hb[T]) conditions. Monovalent derivatives included Hb with mono(3,5-dibromosalicyl)fumarate bound to the beta chains (HbFMDA) and Hb in which alpha and beta chains were reacted with 2-isothiocyanato-benzene-1,4-disulfonic acid (HbICB) or 4-isothiocyanatobenzene sulfonic acid (HbICBS). Anion-induced autooxidations of Hbs were carried out with azide in PO4 buffer in a rapid scanning spectrophotometer. Pseudo-first-order rate constants (K′1) were calculated for each Hb and compared with those of unmodified HbAo. P50s (torr) were measured in BisTris (pH 7.4, 25°C) and in PO4 buffer (pH 7.2, 27°C). Decreased Hb oxygen affinities were associated with increased rates of autooxidation. Menadione-induced oxidation in the presence of 2-propanol caused precipitation of HbAo within 35-45 minutes, whereas precipitation of the bivalently crosslinked Hbs occurred much later (> 24 hrs). Precipitation of the monovalent Hb derivatives occurred within 1-2 hrs. This work provides further evidence that modifying Hb to enhance oxygen off-loading in tissues will increase intrinsic rates of methemoglobin formation. In addition, greater stabilization of the oxidized tetramers in solution is achieved with bivalent as opposed to monovalent crosslinkers.
|Original language||English (US)|
|Number of pages||1|
|Journal||Biomaterials, Artificial Cells, and Immobilization Biotechnology|
|State||Published - Dec 1 1991|
|Event||8th World Congress of the International Society for Artificial Organs in conjunction with the 4th International Symposium on Blood Substitutes - Montreal, Que, Can|
Duration: Aug 19 1991 → Aug 23 1991
ASJC Scopus subject areas