Abstract
1. Symmetrical disulfides, such as O-benzoylthiamine disulfide (BTDS) or thiamine disulfide (TDS) are reduced to free thiamine when it is allowed to react with heat- or urea-denatured egg or serum albumin under certain conditions, whereby a protein-thiamine complex is partially formed. The complex can be separated by deproteinization with metaphosphoric acid, which can also stop the reducing reaction of protein-SH. The complex thus separated can easily be converted into free thiamine by reduction with thiosulfate, followed by determination by thiochrome method. Applying this method, reactions of BTDS or its related compounds with protein-SH were examined and the following results were obtained.
2. Egg albumin in native state scarcely reacts with the thiamine derivatves. Further, denatured protein, whose SH group had been oxidized by iodine or blocked by P-chloromercuribenzoate, loses its reactivity, scarcely producing the complex.
3. Serum albumin reacts both in native and denatured states, producing a complex.
4. The complex formation with denaturedprotein takes place only with symmetrical disulfides. Asymmetric thiamine derivatives such as thiamine propyl disulfide, produce only free thiamine. It was also suggested that the same complex was formed by a reaction of thiamine or O-benzoylthiamine (OBT) with oxidized protein.
5. In the reaction of BTDS with heat-denatured protein, the optimun pH of the reaction, the effects of the protein amount, the concentration of added BTDS, temperature and time were investigated and the relationship between complex formation and production of free thiamine was clarified. In any case, complex formation was proportional to the amount of free thiamine formation, increasing with the progress of the reaction.
6. The protein-thiamine complex is, therefore, considered to be due to chemical binding of the thiamine derivatives with protein-SH as protein-S-S-thiamine.
