Abstract
Oxidation of thiamine by BrCN-NaOH method gave two clearly distinct fluorescent products when developed through a gel filtration chromatography and also when developed by two solvent systems in a thin layer chromatography. Further, a third fluorescent oxidation product was detected in a small amount when a large amount (70mg) of thiamine was similarly oxidized and developed by the gel filtration chromatography. Oxidation of thiamine by K_3Fe(CN)_6-NaOH method gave a single fluorescent product, which was identical with an authentic sample of thiochrome. The two major oxidation products obtained by BrCN-NaOH method (Thc I and Thc II) were crystallized and compared with the standard sample of thiochrome in mixed melting point test, IR spectra, thin layer chromatography, NMR spectra, Mass spectra, UV absorption spectra and fluorescence spectra. It was concluded from those data that Thc I is the same compound as the authentic thiochrome and Thc II is a different product. These two oxidation products showed, however, nearly the same UV absorption spectra and fluorescence spectra. From the spectrophotometric and fluorophotometric properties of Thc I and Thc II, we conclude that the BrCN-NaOH method is still one of the most valuable assay method of thiamine.
