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After proteolytic digestion, aqueous extraction, and derivatization with diethyl pyrocarbonate or ethyl chloroformate, HPLC-inductively coupled plasma (ICP)-MS, GC-atomic emission detection (AED), and GC-MS analysis of high-selenium yeast stored at room temperature for more than 10 years showed selenomethionine as the major Se product along with substantial amounts of selenomethionine selenoxide hydrate and the previously unreported selenoamino acid having a Se-S bond, S-(methylseleno)cysteine. The identity of the latter compound was confirmed by synthesis. The natural product was shown to be different from a synthetic sample of the isomeric compound Se-(methylthio)selenocysteine. Selenium-specific NMR spectroscopic methods were developed to directly analyze the aqueous extracts of the hydrolyzed selenized yeast without derivatization or separation. Selenomethionine and S-(methylseleno)cysteine were identified by 77Se-1H HMQC-TOCSY experiments.