A method for the simultaneous determination of pyrithiamine and thiamine in biological materials.

Abstract

A more precise and sensitive method for the separate determination of Pyrithiamine (PyT) and thiamine in biological materials was described. The method was based on the different behavior of these two compounds toward the oxidation with cyanogen bromide or potassium ferricyanide, and on the different characteristics of the fluorescence spectra of the oxidation products.Thiamine and PyT in biological materials were extracted by homogenizing and heating in an acidic medium, followed by deproteinization with metaphosphoric acid, adsorption on permutit and elution with hot KCI-HCI. For the determination of PyT, an aliquot of the eluate was oxidized with alkaline ferricyanide (1% in 10% NaOH) and the oxidation product, after extraction into isobutanol, was determined fluorometrically using a spectrofluorometer with an excitation at 430mμ and emission at 460mμ. In the case of the thiamine determination, the same eluate was oxidized with cyanogen bromide and the isobutanol extracted thiochrome was determined at 375mμ exciting and 420mμ emitting wavelengths. The lowest limit of detection for PyT was less than 0.3mμmole, and that for thiamine was less than 0.03mμmole per assay.