Abstract
Thiamine anhydride (TA) was isolated from the alkaline solution of thiamine with benzenesulfonyl chloride (BSC) after its overnight standing at room temperature but thiamine disulfide(TDS) (I) and its diphenylsulfonate (II), instead of TA, were obtained by the same procedure under ice-cooling. Bessey's method of estimating thiamine by alkaline BSC-treatment also depends on the same inactivation of thiamine. We checked whether formation of TDS or its disulfonate is possible during the estimation procedure by the following ways : (1) Thiamine in 0.1〜l000mg% solutions was inactivated by addition of BSC and N NaOH completely ; no thiamine was detected by thiochrome method. After the inactivation, TDS was transformed into thiamine by means of cysteine-treatment at pH 5.5 and determined by cyanogen bromide oxidation ; 2,1,5,5 and 9% of the Initial thiamine contents were found as TDS respectively to 0.1,1,10,100 and 1000 mg % concentration of thiamine. If the inactivation by BSC was carried out under ice-cooling, formation of TDS was increased to 3,5.5,9,12 and 16 % at the corresponding concentrations. (2) Thiamine-phenyl-sulfonate (III) was converted into TA easily and completely by making its solution more alkaline than pH 10 ; no TDS was proved by means of cysteine treatment. On the other hand TDS diphenylsulfonate was transformed into TA slowly and incompletely by making its solution even more alkaline than pH 12. Thus it was proved that thiamine was transformed into TA throgh thiamine-phenylsulfonate (thiol-form) (IV) by the alkaline BSC-inactivation and formation of TDS was less than 2% at dilute concentration such as used in the estimation procedure.
