Hydrolytic enzyme substrates. I. Chemical synthesis and characterization

The synthesis and characterization of a number of new phosphate, sulfate and acetate esters of 3-(p-nitrophenoxy)-1,2-propanediol (PNG); 3-(2,4-dinitrophenoxy)-1,2-propanediol (DNG); 4-(p-nitrophenoxy)-1,2-butanediol (PNB) and 4-(2,4-dinitrophenoxy)-1,2-butanediol (DNB) are described. These esters were prepared to serve as substrates for their corresponding hydrolytic enzymes. The assay system used to measure enzyme hydrolysis requires periodate oxidation of the diol formed after hydrolysis of the ester. Base treatment of the resulting aldehyde yields either p-nitrophenolate ion or the 2,4-dinitrophenolate ion depending upon the substrate. In the presence of high concentrations of methylamine and excess periodate the oxidation and elimination reactions can be carried out simultaneously at pH 7.5. The reactions leading to these results are described.