KINETICS OF ENZYMATIC REDUCTION OF METMYOGLOBIN IN RELATION TO OXYGEN ACTIVATION IN MEAT-PRODUCTS

Enzymatic reduction of metmyoglobin by partially purified cytochrome b5 reductase and cytochrome b5 from beef liver and NADH was investigated in model systems by absorbance measurements; and the influence on myoglobin-induced oxidation of linoleic acid was investigated by measuring oxygen consumption of emulsified linoleic acid solutions. The effect of the enzyme system was further investigated for meat products in order to elucidate the coupling between redox processes of meat pigments and lipids during freezer storage of minced beef. The rate of enzymatic reduction of metmyoglobin to myoglobin was strongly dependent on pH, ionic strength and temperature. For excess of NADH, the energy of activation was pH-dependent and approximated 60 kJ . mol-1, and was assigned to the electron transfer between reduced cytochrome b5 and metmyoglobin. The pH-dependence indicated parallel reactions of two protein forms in acid/base equilibrium with each other; a pK(a) = 6.0 suggests the involvement of an imidazol moiety, for which the protonization facilitates the electron transfer. The reaction centres of the proteins are oppositely charged, as concluded from a decrease in rate with increasing ionic strength. For conditions of high enzyme activity, metmyoglobin was reduced to myoglobin but not further oxygenated to oxymyoglobin, indicating oxygen consumption as the result of a direct reduction of O2 by cytochrome b5. For intermediate enzyme activity (25-degrees-C, pH 6.3 and 5.8), the liver extract and NADH were capable of decreasing the metmyoglobin-induced oxygen consumption rate of linoleic acid emulsions by a factor of 10, showing that the iron(III) form of myoglobin is the active oxidation catalyst. However, for meat products, addition of the liver extract and NADH did not improve the colour stability as monitored by tristimulus colorimetric measurements but significantly increased lipid oxidation, as indicated by an increase in thiobarbituric-acid-reactive substances.