Summary
Glyoxylic acid vapour is a most powerful reagent for the fluorescence histochemical visualization of biogenic monoamines. In the present investigation the mechanisms of fluorophore formation in the glyoxylic acid reaction has been studied in detail for tryptamine in histochemical models and in freeze-dried tissue, utilizing microspectrofluorometric, Chromatographic, and mass spectrometric techniques in combination with isotope measurements.
The glyoxylic acid-tryptamine reaction proceeds through an initial Pictet-Spengler type cyclization to 1,2,3,4-tetrahydro-β-carboline-1-carboxylic acid, followed by two alternative fluorophore forming reactions yielding 3,4-dihydro-β-carboline, or the 2-carboxymethyl-3,4-dihydro-β-carbolinium and 2-methyl-3,4-dihydro-β-carbolinium salts, which are all strongly fluorescent. It is shown that the yield of fluorophores is considerably higher in the glyoxylic acid vapour reaction than in the formaldehyde vapour reaction of the standard Falck-Hillarp method, and that this higher efficiency of glyoxylic acid is due to the most favourable catalysing properties of the carboxylic group of the glyoxylic acid molecule.
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Björklund, A., Lindvall, O. & Svensson, LÅ. Mechanisms of fluorophore formation in the histochemical glyoxylic acid method for monoamines. Histochemie 32, 113–131 (1972). https://doi.org/10.1007/BF00303727
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DOI: https://doi.org/10.1007/BF00303727