ESTIMATION 



chemical methods. E. E. Snell and L. D. Wright ^^ measured the 

 amount of lactic acid produced by Lactobacillus arabinosus 17-5 on a 

 synthetic mediimi, containing all known growth factors except nico- 

 tinic acid and to which the test solution had been added, and com- 

 pared it with the amount produced on the basal medium containing 

 known amounts of added nicotinic acid. The nicotinic acid contents 

 of blood, milk, urine, plant and animal extracts were estimated in this 

 way with a fair degree of accuracy ; as little as 0-05 jug. of nicotinic 

 acid could be measured. 



The method of Snell and Wright failed to give a linear standard 

 curve with concentrations in excess of 0-2 /u,g. per 10 ml., but this 

 objection was overcome by Krehl et al.^^ who, in addition to other 

 modifications, increased the concentration of glucose and buffer 

 (sodiimi acetate) in the basal medium, whilst H. Isbell ^* advocated 

 the addition of ^-aminobenzoic acid to the medium. The medium was 

 still further modified by E. C. Barton- Wright, ^^ whose method has 

 been adopted as a standard procedure by most workers in this country. 

 The medium consists of acid-hydrolysed casein, the preparation of 

 which must be carefully carried out if satisfactory results are to be 

 obtained, DL- tryptophan and L-cystine, glucose, sodirnn acetate and 

 xylose, calcium pantothenate, pyridoxine, riboflavine, ^-amino- 

 benzoic acid and biotin, adenine, guanine, uracil and xanthine, and the 

 usual inorganic salts. Barton-Wright recommends that extracts be 

 prepared in the same way as for riboflavine assays (page 158), although 

 the growth of L. arabinosus is not stimulated by starch or free fatty 

 acids ; hydrolysis is effected by autoclaving with N-hydrochloric acid 

 at 15 lb. pressure for fifteen to twenty minutes. A similar method was 

 made official in the U.S.A.^^ 



Sarett et al}"^ proposed to use for the assay of nicotinic acid with 

 L. arabinosus a basal medium consisting of yeast extract, peptone, 

 and liver extract treated with Lloyd's reagent to remove nicotinic 

 acid. 



A method for the microbiological estimation of nicotinamide in 

 presence of nicotinic acid by means of L. arabinosus was devised by 

 Atkin et al.,^^ who carried out two assays, one before and one after 

 the conversion of nicotinamide, by treatment with bromine and 

 potassium hydroxide, into 3-aminopyridine which has no growth- 

 stimulating activity. The difference in the amount of lactic acid 

 produced in the two tests was proportional to the amount of nicotin- 

 amide present. 



Assay methods based on the response of L. arabinosus have been 

 most commonly employed, but other organisms have been used. 

 P. Fildes ^^ found that Proteus vulgaris required nicotinic acid, and 

 A. Lwoff and A. Querido^® used it for the estimation of nicotinic 



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