Abstract
The improved thiochrome method using benzenesulfonyl chloride as thiamine inactivating agent was proposed by O.Bessey (J.Biol.Chem.195,453,1952). We checked this method and one drop of benzenesulfonyl chloride was proved sufficient to decompose 50 μg thiamine completely in alkaline solution. A sample solution (I) containing thiamine can be converted into thiochrome by alkaline ferricyanide solution, while another solution (II) containing the same amount of thiamine, after alkaline benzenesulfonyl chloride treatment does not produce thiochrome on ferricyanideoxidation. Excess of ferricyanide is both decolorized by alcoholic H_2O_2 solution, then a thiochrome standard solution (0.1 mg% in 0.01N HCl) is poured into the latter solution (II) drop by drop until its fluorescence becomes the same strength as the former (I) by comparing them under ultraviolet ray. Thus thiamine can be estimated by the simplified method using thiochrome water solution without isobutanal extraction. Crystals of mp. 113-114℃ were isolated as the reaction product of thiamine and benzenesulfonyl chloride, as we published it this journal 13.
