Oxidation of Tetrahydropteridines and Tetrahydrofolates

Abstract

The autoxidation of tetrahydropteridines in aqueous media has been studied. Particular attention has been paid to tetrahydrofolic acid, which is readily oxidised in solution by air. At pH's 3, 7, 10 and 13 the products were p-aminobenzoylglutamic acid, 2-amino-4-hydroxypteridine, xanthopterin, folic acid and dihydrofolic acid. These were identified by t.l.c.,mass spectra, u.v. spectra and fluorescence spectra and estimated using fluorescence spectra. Although 2-amino-4-hydroxypteridine and xanthopterin accounted for 50 - 60% of the tetrahydrofolic acid used, their ratio was pH dependent. In the presence of aqueous ammonia 2, 6-diamino-4-hydroxypteridine was formed. A mechanism involving hydroperoxide intermediates has been proposed for these reactions. Dihydrofolic acid was oxidised to give folic acid and 2-amino-4-hydroxy-6-formyl-7, 8-dihydropteridine, the latter compound being further oxidised to xanthopterin. A buffer effect on the rate of oxidation was observed. The formation of hydrogen peroxide was shown to occur during the oxidation of dihydrofolic acid. Oxidations of tetrahydropteridines were followed using time-based u.v. spectral studies. The effects of pH, buffer and deaeration of the solution were observed using this technique. Each compound showed the most rapid and marked spectral changes when in phosphate buffer, pH 7. The results are explained and discussed on the basis of hydroperoxide involvement. Previous work has postulated the formation of a quinonoid dihydropteridine intermediate from the tetrahydropteridine by a reversible step, which irreversibly changes to the dihydropteridine. The oxidation of tetrahydropteridines by 2, 6-dichlorophenolindophenol and the appearance of a transient peak at max 303nm. when oxidised by ferricyanide have been attributed to such an intermediate. This evidence has been examined more fully and the results obtained are not in accord with the quinonoid dihydropteridine scheme. All the evidence supporting such a scheme is considered and a better interpretation, on the basis of a scheme involving hydroperoxide intermediates, is discussed in detail.