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N'-Methylnicotinamide has been shown to be one of the principal excretion products derived from the metabolism ofnicotinic acid in man, the dog and the rat (Najjar, Scott & Holt, 1943; Huff & Perlzweig, 1943 a, b; Ellinger & Coulson, 1943). Various methods have been worked out for its estimation employing adsorption on to, and elution from, an active surface, followed by extraction into a butanol phase. The fluorescence in the butanol layer, that is the F2 fluorescence, has then been measured (Huff & Perlzweig, 1943c; Coulson, Ellinger & Holden, 1944; Najjar, 1944). Huff, Perlzweig & Tilden (1945), and Huff & Perlzweig (1947) have published a new method for the estimation of Nl-methylnicotinamide, involving the formation of a fluorescence in the presence of acetone in a single aqueous phase. A fluorescence was produced if coenzymes were similarly treated and this method has been used for their estimation in blood (Levitas, Robinson, Rosen, Huff & Perlzweig, 1947) and animal tissues (Robinson, Levitas, Rosen & Perlzweig, 1947). The present paper deals with studies of the optimum conditions for the production of fluorescence ofsome Nl-substituted pyridine compounds. A modified procedure has been devised using methyl ethyl ketone instead of acetone, and employing different blanks. The fluorescence due to Nl-methylnicotinamide has been differentiated from that ofthe phosphopyridine nucleotides and Nl-methylnicotinuric acid betaine. The interference due to indole has been eliminated.

The fluorimetric estimation of N1-methylnicotinamide and its differentiation from coenzyme I.