Abstract
DETERMINATION of individual amino-acids in a chromatographed mixture by measuring nitrogen content was discouraged by Martin and Mittelmann1, who reported interference from the solvents used. The method has now been re-investigated. Chromatograms were prepared using Whatman No. 1 paper and the apparatus described by Datta, Dent and Harris2; the solvents were (a) n-butanol-acetic acid3, and (b) a mixture of phenol and water (4 : 1) in the presence of acetic acid. The papers were dried overnight at room temperature, then sprayed with Moore and Stein4 reagent diluted with methyl alcohol to contain 0.1 gm. ninhydrin/100 ml. The colours obtained with this reagent were found to be more stable than those given by a solution of ninhydrin in n-butanol, and there was less vertical streaking of the spots on the paper. The papers were heated, the spots outlined with pencil and identified by comparison with a series of similarly prepared ‘maps’. Demarcation of the spots is helped by the fact that adjacent spots often give slightly different colours with ninhydrin. The identified spot and a piece of paper of the same area were cut from the same sheet, the latter for a blank determination.
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References
Martin, A. J. P., and Mittelmann, R., Biochem. J., 43, 353 (1948).
Datta, S. P., Dent, C. E., and Harris, H., Science, 112, 621 (1950).
Partridge, S. M., Biochem. J., 42, 238 (1948).
Moore, S., and Stein, W. H., J. Biol. Chem., 176, 367 (1948).
Fowden, L., and Penney, J. R., Nature, 165, 846 (1950).
Chibnall, A. C., Rees, M. W., and Williams, E. F., Biochem. J., 37, 354 (1943).
Awapara, J., Arch. Biochem., 19, 172 (1948).
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KLATZKIN, C. Quantitative Determination of Amino-acids Separated by Paper Partition Chromatography. Nature 169, 422 (1952). https://doi.org/10.1038/169422a0
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DOI: https://doi.org/10.1038/169422a0
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