Oxidation of Ascorbic Acid by Enzyme Preparations from Orange

Many of the flavor compounds in citrus fruit are synthesized by oxidation-reduction reactions mediated by enzymes in the fruit. The oxidant in some of these reactions is the coenzyme, NADP. One possible way to regenerate this coenzyme to the active oxidized form is through the enzymic oxidation-reduction reactions involving glutathione and ascorbic acid. A system which degrades ascorbic acid in preparations from mature fruit without the con sumption of O2 has been found. This atypical system is sensitive to a number of compounds which inhibit typical ascorbic acid oxidase as well as several which do not. Typical ascorbic acid oxidase prepared from the peel of green oranges is easily distinguishable from the ma ture fruit system on the basis of chemical and physical properties. A number of factors point to the enzymic nature of this system: 1) the system is heat labile, being completely inactivated by two min utes at 100°C; 2) the reaction is stereospecific, oxidizing L-ascorbic acid approximately eight times faster than D-ascorbic acid; 3) the system exhibits an optimum reaction pH; and 4) ex tensive dialysis does not reduce the activity more than 20%. Results suggest that an inhibi tory reaction product is quickly formed which prevents further ascorbic acid oxidation. The existence of other enzymes which could function in the oxidation of alcohols and alde hydes utilizing oxidized glutathione and dehydroascorbic acid was demonstrated. These en zymes, glutathione reductase and dehydroascorbic acid reductase, are linked to a number of dehydrogenases via the oxidation of coenzymes. Both enzymes were present in mature fruit at somewhat higher concentrations than in green peel. Ascorbic acid oxidase activity of the prepa ration is destroyed at pH 4.0, which is slightly lOne of the laboratories of the Southern Utilization Re search and Development Division, Agricultural Research Service, U. S. Department of Agriculture. References to specific commercial products do not con stitute endorsement. higher than the pH of commercially extracted juice. In addition activity has been observed only in preparations from carefully neutralized juice, and not from conventionally extracted juice. Thus, the enzyme would not be expected to contribute to destruction of ascorbic acid in