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The effect of α-Tocopherol (α-Toc) on the reduction of metmyoglobin (MetMb) was investigated using a liver extract containing cytochrome b(5) (LE) prepared from bovine liver in a Triton X-100 system at 37 °C and at pH 7.2, 6.2, and 5.6. The combination of α-Toc and LE reduced MetMb at pH 5.6 and pH 6.2 (p0.05), but not at pH 7.2. When LE was added, substitution of α-Toc with glutathione (0.15μM) resulted in 13% MetMb reduction, a rate similar to α-Toc plus LE (pH 5.6). The addition of NADH and NADPH with LE at pH 5.6 resulted in MetMb reduction of 85% and 25%, respectively, relative to controls. α-Toc-mediated reduction at pH 5.6 was further investigated using a purified bovine liver cytochrome b(5) (Cyt b(5)). α-Toc effectively reduced Cyt b(5), as indicated by an increased absorbance at 424 nm. The combination of α-Toc plus purified Cyt b(5) resulted in a 15% reduction in MetMb at pH 5.6 (p0.05) relative to controls. These results suggested a potential reaction whereby α-Toc maintains OxyMb via enhancement of Cyt b(5) mediated reduction of MetMb. |
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