GASTRIC CARCINOGENESIS - 2-CHLORO-4-METHYLTHIOBUTANOIC ACID, A NOVEL MUTAGEN IN SALTED, PICKLED SANMA-HIRAKI FISH, OR SIMILARLY TREATED METHIONINE

The customary salting and pickling of fish in high risk gastric cancer regions were modeled to explore the relevant causative chemicals. The fish Sanma hiraki was treated with sodium chloride and sodium nitrite at pH 3. Previously, it had been found that an extract of the treated fish was mutagenic in Salmonella typhimurium TA 1535 without S9 and a lso that it induced glandular stomach cancer upon gavage to rats. We n ow demonstrate that the mutagenicity was enhanced by preincubation of the raw meat for several days before salt-nitrite treatment. HPLC tech niques showed that three mutagens were present in the fish extract. On e of the mutagens was found to be stable over the pH range of 1.0-9.0. This mutagen was purified by silica gel solid phase extraction, follo wed by a series of reverse phase HPLC steps, and was characterized by low and high resolution MS, NMR, and FT-IR. While N-nitroso compounds were generally believed to be associated with gastric carcinogenesis, it was unexpectedly found that the mutagen has the novel structure 2-c hloro-4-methylthiobutanoic acid (CMBA). Based on the structure, it see med likely that methionine might be the precursor, and this was, indee d, proven. Both salt and nitrite are essential factors for forming thi s mutagen. The yield of CMBA was linear for chloride concentrations fr om 0 to 800 mM NaCl. Of 20 amino acids reacted with nitrite and chlori de at pH 3, only methionine generated a mutagen for S. typhimurium TA 1535. Tryptophan gave a product mutagenic in S. typhimurium TA 100 and TA 98, but not TA 1535, and in the case of tyrosine, the mutagen was active only for TA 100. These results suggest an important role for sa lt in gastric carcinogenesis and provide new approaches for exploring the formation of mutagens/carcinogens for specific target organs.