ASCORBATE-2-SULFATE SULFOHYDROLASE IN FISH AND MAMMAL - COMPARATIVE CHARACTERIZATION AND POSSIBLE INVOLVEMENT IN ASCORBATE METABOLISM

1 . The new assay conditions were determined for crude and purified enzyme ascorbate-2-sulfate sulfohydrolase from liver tissues of two fish species and bovine. 2. The major departure from the existing indirect method, based on reduction of 2,6-dichlorophenolindophenol (DCIP) by released ascorbic acid and change from pink-blue to a colorless molecule, takes into account the shift of maximum absorbance of DCIP from 516 nm at pH 5.14 to 600 nm at pH 6.5. 3. The direct method is based on colorimetric assay of liberated ascorbic acid including correction for interfering substances. The optimum pH for both fish ascorbate sulfatases was 5.5. 4. The K(m) for bovine ascorbate sulfatase was confirmed to be approximately 7 mM at 37-degrees-C. 5. Partly purified ascorbate-sulfate sulfohydrolase has a K(m) value in rainbow trout of 0.4 mM and it changes very little in the range of water temperatures characteristic for this stenothermic fish species. 6. In eurythermic chub, the K(m) values increased from 1.2 to 4.3 mM with rising temperatures.