NICOTINIC ACID (NIACIN) 



Estimation of N^-Methylnicotinamide 



As already stated, ingested nicotinamide is partially converted 

 into N^-methylnicotinamide, which can be estimated by conversion 

 into a fluorescent substance by treatment with alkali. 



J. W. Huff and W. A. Perlzweig *'' used adsorption on Permutit at 

 pH 4 to separate the substance from impurities, and eluted it from the 

 adsorbate by means of potassium chloride solution. It was then 

 extracted into butanol, aqueous alkali was added and the fluorescence 

 evaluated. In estimating N^-methylnicotinamide in urine, the pre- 

 liminary adsorption on zeolite was unnecessary. Coulson et al.^^ 

 adsorbed on Decalso, eluted with potassium chloride solution and then 

 extracted the eluate with sec. butanol after making alkaline. V. A. 

 Najjar *^ treated the solution with active charcoal, adsorbed the 

 N^-methylnicotinamide on Permutit and eluted with potassium 

 chloride solution ; as in the foregoing method, the eluate was then 

 made alkaline, extracted with butanol, and the fluorescence of the 

 solution was evaluated in a fluorimeter. M. Hochberg et al.^^ also 

 adsorbed on a zeolite. 



Huff et al.^^ improved the method by treating an aqueous alkaline 

 solution with acetone instead of extracting into isobutanol. This 

 produced a greenish-blue fluorescence, the intensity of which was pro- 

 portional to the concentration of N^-methylnicotinamide. E. Kodicek ^^ 

 used methyl ethyl ketone in place of acetone, but subsequently found 

 that satisfactory results were obtained in the absence of a solvent. 



Estimation of Ni-Methyl-6-pyridone-3-carboxylamide 



The amount of N^-methyl-b-pyridone-s-carboxylamide in urine 

 was estimated by clarifying the urine with lead acetate, saturating the 

 filtrate with potassium carbonate and extracting with ether, evaporat- 

 ing the extract to dryness, dissolving the residue in acetone and 

 measuring the fluorescence produced on addition of a small amount of 

 concentrated potassium hydroxide solution. Alternatively the pyri- 

 done was estimated spectrophotometrically in an aqueous solution of 

 the residue left after evaporation of the ethereal extract. ^i" A third 

 method was to adsorb the pyridone on Lloyd's reagent, elute with 

 chloroform, nitrate and measure the coloured product in alkaline 

 solution. ^1* 



iViicrobiological Methods of Assay 



An entirely different principle, depending on the response of micro- 

 organisms to nicotinic acid, has been used by a number of workers 

 who claim that such methods are more expeditious and accurate than 



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