254 PYRIDOXINE AND RELATED COMPOUNDS 



under many conditions'^' '^; under others, pyridoxal and pyridoxamine 

 may show shghtly less activity than pyridoxine. The three forms show very 

 different activities, however, for many microorganisms.'^ All lactic acid 

 bacteria, for example, are unable to utilize pyridoxine for growth; most use 

 pyridoxal and pyridoxamine, some only pyridoxal. "* These organisms, 

 therefore, are not useful for determining the total vitamin Be content of 

 natural materials. For many yeasts and molds, however, all three forms 

 show equal activity;'^ conseciuently, these organisms are most useful for 

 assay purposes. 



Among these latter organisms, assay methods utilizing Neurospora sito- 

 phila pyridoxineless'^ ■ '^' '^ and Saccharomyces carlsbergensis 4228''^ ■ ^"'^^ 

 have been studied most thoroughly. Both yield excellent results; the latter 

 procedure, however, is simpler, faster, and more convenient. GroA th of the 

 test organism in the vitamin Be-free medium increases with the vitamin Be 

 concentration in the range from to about 0.04 y per 10 ml. of medium. 

 Pure vitamin Be (any of the forms may be used as standard) and samples 

 to supply the vitamin at several levels within this range are added to indi- 

 vidual 1 in. by 8 in. tubes containing 4 ml. of medium. Eac h tube is then 

 diluted to 9 ml., plugged or otherwise ( overed, and sterilized by heating in 

 flowing steam at 100° for 10 minutes. After cooling, 1 ml. of inoculum 

 suspension (0.3 mg. of moist yeast) is sdded, and the tubes are shaken 

 mechanically for 16 to 18 hours. Yeist growth is then determined turbidi- 

 metrically. The responses to various levels of the pure vitamin are plotted 

 to yield a standard curve, from which the vitamin Be ( ontent of the sample 

 aliquots can be determined by interpolation. Details of the procedure have 

 been treated elsewhere.'^' *'" 



If it is desired to determine pyridoxal plus pyridoxamine, but not pyri- 

 doxine, Streptococcus faecalis may be used as the assay organism.''* • ^^- ®^ 

 Similarly, use of Lactohacillus casei as the assay organism permits the spe- 

 cific estimation of pyridoxal.''' ■ ^''-' "'* Differential assays of this type reveal 

 that most of the vitamin Be present in yeast, meats, glandular organs, etc., 

 is present as pyridoxal and pyridoxamine, with only traces (if any) of 



S6 E. E. Snell and A. N. R:uinefeld, J. Hiui. Chnn. 157, 475 (l!)4o). 



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Interscience Publishers, Ncvv ^'ork, 1951. 

 «iL. Atkin, A. S. Schultz, W. L. Willi;ims, and ('. N. Froy, /«</. E,,,/. ('hn,i. Annl. 



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