ASSAY METHODS 57 



is used, and the ^-alanine effect has been cancelled out by using more 

 asparagin in the medium; the results appear to be satisfactory. 93 It has 

 the advantage of speed (16 hours) over the titration procedure using 

 lactic acid bacteria, but has the disadvantage that, as described, the 

 cultures have to be continuously shaken. 



Lactic acid bacteria 94 have been used most extensively for pantothenic 

 acid assay following the similar methods of Pennington et al. 90 and 

 Strong et al. 95 The organism L. casei has been used successfully in the 

 assay of a great many types of materials. 96-106 Fatty substances inter- 

 fere, as in the case of riboflavin assay, 52 but this difficulty can often be 

 cared for by careful filtration at pH 4.5 or preliminary fat extraction, as 

 in the case of the riboflavin assay. Inhibiting substances produced by 

 the clarase digestion of yeasts and other materials are reported to inter- 

 fere with the determination by causing a downward drift of values with 

 increasing test dosage. 103 Such substances would be most disturbing if 

 present in low-potency material ; but yeast, which was the worst offender, 

 is, of course, a relatively rich source. Thompson, Cunningham and Snell 106 

 found in the assay of canned foods that there was often a downward 

 drift in the assay values with increasing dosage which, however, tended 

 to reach a definite level. Presumably in such cases the effect is that of 

 extraneous stimulating substances which are effective at low dosage levels. 

 Neal and Strong, 104 by suitable supplements to the basal medium, elimi- 

 nated such drifts which, however, appear not to prevent obtaining satis- 

 factory assay values, provided the higher dosage values (lower assay 

 values) are accepted as correct. 106 



While L. casei has been used extensively as a test organism for pan- 

 tothenic acid assay, it has the disadvantage that its growth is greatly 

 affected by fatty substances and by unknowns which must be introduced 

 into the basal medium in the form of crude extracts. Hoag, Sarett and 

 Cheldelin 107 and Skeggs and Wright 108 have developed assay methods 

 using L. arabinosus 17-5. In one laboratory, the Pennington et al. 90 

 medium was modified, principally by using norite-treated autolyzed yeast 

 and rice bran concentrate in the basal medium in place of the alkali- 

 treated yeast extract. In the other, the medium of Snell and Wright 7S 

 for nicotinic acid assay was modified by altering the amounts of some of 

 the constituents, by introducing xanthine and p-aminobenzoic acid, and 

 by substituting nicotinic acid for pantothenic acid. For this organism 

 the effects of fatty substances are less marked, but are enough to intro- 

 duce substantial errors. Hoag et al. advocate a preliminary ether extrac- 

 tion for samples of high fat content. Skeggs and Wright introduced oleic 

 acid, which they found to be stimulative, into the basal medium. If this 

 is not done, they advocate a preliminary ether extraction. In both 



