To know the amount of oxygen absorbed in the autoxidation of myoglobin (Mb) to metmyoglobin (MMb) is needed for elucidating the mechanism of, and devising the prevention of, fish meat discoloration during freezing storage. So, experiments were carried out to confirm whether or not the oxygen absorption of 2.5±0.3 moles per mole of horse Mb
1) is also true for fish Mb's.
Crystalline MMb's of two species of tuna, whale, and horse, were dissolved in 0.1M phosphate buffer, pH 6.7, added with small amounts of sodium hydrosulfite, and shaken in air to oxidize excess hydrosulfite. The oxymyoglobin (MbO
2) solution thus obtained was subjected to manometric analysis at 30°C using a WARBURG apparatus. The amount of MMb formed during the manometry was estimated by the authors' method
5). According to the results, the amount of oxygen absorbed per mole of MMb formed roughly resembled to that reported by GEORGE
et al., varying widely as pointed out by them (Table 1). The low reproducibility was assumed to be attributable to the presence of substances interfering in manometry such as decomposition products of hydrosulfite which are able to absorb oxygen, although not to reduce MMb. This assumption was just supported by an experiment on sodium bisulfite, one of such decomposition products (Fig. 1). Therefore, the subsequent experiments were performed on MbO
2 solutions, from which the decomposition products of hydrosulfite were previously removed by dialysis before manometry. Thus the absorption amount of oxygen was found to be only about 0.34 mole per mole of Mb, irrespective of the species exam ?? ned, and the results were fairly reproducible (Table 2).
And some model experiments were performed to clear up whether large amounts of oxygen are absorbed or not by globin moiety of Mb in the autoxidation, as suggested by GEORGE
et at.
1) None of several amino acids tested absorbed oxygen in appreciable amounts (Fig. 2). In addition, it was also proved that free sulfhydryl groups were not oxidized during manometric experiments in the case of tuna (Table 3). These results seem to indicate slight, if any, involvement of protein moiety in Mb autoxidation.
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