Studies on Models for Tetrahydrofolic Acid. III. Hydrolytic Interconversions of the Tetrahydroquinoxaline Analogs at the Formate Level of Oxidation

Hydrolysis of the formamidinium salts IV–VI was studied as a function of pH and buffer concentration as a model for nonenzymic interconversion of formate-carrying tetrahydrofolic acid. Rates for the p-ethoxycarbonylformamidinium salt hydrolysis determined above pH 6 exhibited first-order kinetics with marked buffer catalysis by all buffers studied. Rates measured at pH ≤6 exhibited nonlinear kinetics due to subsequent isomerization of the initially formed N¹⁰-formyl product to the more stable N¹-formyl compound. These data are rationalized by one kinetic scheme in which all three species arise from a common intermediate or a collection of protonically related intermediates, the step for production of the N¹-formyl being kinetically significant only at pH ≤6. Each buffer used exhibited a unique catalytic facility with respect to relative rates of formation of the two N-formyl products. Equilibrium constants relating the p-ethoxycarbonylformamidinium salt to each N-formyl product were determined and found to be in quantitative agreement with those of the natural cofactor. The close resemblance between the kinetic and thermodynamic dispositions of the model and folate compounds supports the validity of the model and permits comment on the biological implications.