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The fate of radiolabeled cyanide added to washed non-proliferating suspensions of the cyanogenic organism Chromobacterium violaceum was studied. Within 6 h of addition, over 90% of the added cyanide had been metabolized to other compounds, except when l-glutamate alone was included in the suspension medium; in this case only 50% of the added cyanide had been metabolized after 6h incubation. Less than 2% of radioactivity was incorporated into cell material; the rest remained in the suspension medium. The only identified compound into which radioactivity accumulated was β-cyanoalanine. Formation of β-cyanoalanine was stimulated by inclusion of l-glutamate and either O-acetyl-l-serine or l-serine in the incubation medium. Under optimal conditions, approximately 50% of added radiolabel accumulated in β-cyanoalanine. Compounds capable of supplying carbon for growth of C. violaceum were able to substitute, to some extent, for l-glutamate in stimulating β-cyanoalanine synthesis. Both O-acetyl-l-serine and l-serine inhibited β-cyanoalanine formation when included at concentrations in excess of 3 mm. l-Cysteine, l-threonine and l-alanine were unable to substitute for O-acetyl-l-serine or l-serine, and l-cysteine inhibited β-cyanoalanine synthesis. l-Methionine inhibited β-cyanoalanine formation by cells resuspended in cyanide, l-glutamate and O-acetyl-l-serine or l-serine. The effect of growth conditions on the ability of resuspended cells to synthesize β-cyanoalanine was studied.