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. 1965 Oct;90(4):1026–1031. doi: 10.1128/jb.90.4.1026-1031.1965

Microbiological Method for the Determination of l-Tryptophan

Oldrich K Sebek 1
PMCID: PMC315771  PMID: 5847792

Abstract

Sebek, Oldrich K. (The Upjohn Company, Kalamazoo, Mich.). Microbiological method for the determination of l-tryptophan. J. Bacteriol. 90:1026–1031. 1965.—The ability of Chrombacterium violaceum to utilize l-tryptophan for the synthesis of a purple pigment, violacein, served as a basis for the development of a quantitative estimation of this amino acid. The method consists of suspending washed colorless cells of the organism in an agar layer, placing a paper disc impregnated with a tryptophan solution on top of the layer, and allowing the system to incubate. As tryptophan diffuses into the agar, it is converted into violacein, and appears as a zone of striking purple color. Since the diameter of the zone is a function of the amount of tryptophan applied, the amino acid can be quantitatively estimated within the range of 10 to 320 μg per sample with 5.6% standard deviation. The method is fairly specific for free tryptophan, since only indole, indole-3-pyruvic acid, and, to a small degree, anthranilic acid interfere. Other amino acids, tissue homogenates, tryptophan in peptide linkage, or compounds related to this amino acid do not affect its determination. The bacterium does not utilize tryptophan for the synthesis of cellular material unless its growth has been initiated by another substrate.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. BARRNETT R. J., SELIGMAN A. M. Histochemical demonstration of esterases by production of indigo. Science. 1951 Nov 30;114(2970):579–582. doi: 10.1126/science.114.2970.579. [DOI] [PubMed] [Google Scholar]
  2. DELEY J. EFFECT OF MUTATION ON DNA-COMPOSITION OF SOME BACTERIA. Antonie Van Leeuwenhoek. 1964;30:281–288. doi: 10.1007/BF02046734. [DOI] [PubMed] [Google Scholar]
  3. DEMOSS R. D., EVANS N. R. Incorporation of C14-labeled substrates into violacein. J Bacteriol. 1960 May;79:729–733. doi: 10.1128/jb.79.5.729-733.1960. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. DEMOSS R. D., EVANS N. R. Physiological aspects of violacein biosynthesis in nonproliferating cells. J Bacteriol. 1959 Oct;78:583–588. doi: 10.1128/jb.78.4.583-588.1959. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. DEMOSS R. D., HAPPEL M. E. Nutritional requirements of Chromobacterium violaceum. J Bacteriol. 1959 Feb;77(2):137–141. doi: 10.1128/jb.77.2.137-141.1959. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. DICKMAN S. R., CROCKETT A. L. Reactions of xanthydrol. IV. Determination of tryptophan in blood plasma and in proteins. J Biol Chem. 1956 Jun;220(2):957–965. [PubMed] [Google Scholar]
  7. FRANK L. H., DEMOSS R. D. Specific enzymic method for the estimation of L-tryptophan. Arch Biochem Biophys. 1957 Apr;67(2):387–397. doi: 10.1016/0003-9861(57)90293-x. [DOI] [PubMed] [Google Scholar]
  8. HOLT S. J. A new principle for the histochemical localization of hydrolytic enzymes. Nature. 1952 Feb 16;169(4294):271–273. doi: 10.1038/169271a0. [DOI] [PubMed] [Google Scholar]
  9. INGLIS A. S., LEAVIN I. H. STUDIES IN THE DETERMINATION OF TRYPTOPHAN. MODIFIED FISCHL PROCEDURE. Anal Biochem. 1964 Jan;7:10–17. doi: 10.1016/0003-2697(64)90114-9. [DOI] [PubMed] [Google Scholar]
  10. Lugg J. W. On the use of mercuric salts and nitrous acid in the colorimetric determination of tyrosine and tryptophan present in solution. Biochem J. 1937 Aug;31(8):1422–1433. doi: 10.1042/bj0311422. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. MITOMA C., WEISSBACH H., UDENFRIEND S. 5-Hydroxytryptophan formation and tryptophan metabolism in Chromobacterium violaceum. Arch Biochem Biophys. 1956 Jul;63(1):122–130. doi: 10.1016/0003-9861(56)90016-9. [DOI] [PubMed] [Google Scholar]
  12. MRSKOS A., TOVAREK J. Note on a new color reaction for tryptophan and on determination of serum cholesterol. Arch Biochem Biophys. 1960 Nov;91:152–152. doi: 10.1016/0003-9861(60)90469-0. [DOI] [PubMed] [Google Scholar]
  13. SAVCHUCK W. B., MERRIMAN M. E., LOCKHART W. L. ASSAY OF VITAMINS AND AMINO ACIDS WITH CULTURED TISSUE CELLS AND ANTIMETABOLITES. Appl Microbiol. 1964 Jul;12:316–320. doi: 10.1128/am.12.4.316-320.1964. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. SCOTT T. A. An enzymic method for the estimation of L-tryptophan. Biochem J. 1961 Sep;80:462–464. doi: 10.1042/bj0800462. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. SEBEK O. K., JAGER H. Divergent pathways of indole metabolism in Chromobacterium violaceum. Nature. 1962 Nov 24;196:793–795. doi: 10.1038/196793a0. [DOI] [PubMed] [Google Scholar]
  16. SNEATH P. H. Cultural and biochemical characteristics of the genus Chromobacterium. J Gen Microbiol. 1956 Aug;15(1):70–98. doi: 10.1099/00221287-15-1-70. [DOI] [PubMed] [Google Scholar]

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