Summary
A new technique based on liquid phase ion exchange chromatography on short column is proposed for quantitative determination of tyrosine and Dopa.
α-Amino, β-guanidinopropionic acid is used as an internal standard of coloration. The role of H2O2 and ascorbic acid on tyrosine and Dopa was checked. Ascorbic acid prevents the auto-oxidation of Dopa, H2O2 has no effect on tyrosine but oxidizes Dopa even in the presence of excess ascorbic acid. This method was tested in mushroom tyrosinase, with and without ascorbic acid. Assays performed with tyrosinase from rabbit occular extracts clearly showed that they do oxidize tyrosine.
Reliability of the method is comparable to radioassay.
Zusammenfassung
Es wird eine neue Technik für die quantitative Bestimmung von Tyrosin und Dopa beschrieben. Methodisch wurde dabei die Ionen-austauschchromatographie ohne kurze Säulen verwandt. α-Amino, β-guanidinopropionic-Säure wurde als Standard der Färbungsskala verwandt. Der Effekt von H2O2 und Ascorbinsäure für die Substrate Tyrosin und Dopa wurde untersucht. Ascorbinsäure verhindert die Autooxydation von Dopa. H2O2 hat keine Wirkung auf Tyrosin, oxydiert aber Dopa, auch wenn Ascorbinsäure in hohem Überschuß vorliegt. Diese Methode wurde an Pilztyrosinase getestet. Versuche mit Tyrosinase vom Kaninchenaugenextrakt zeigten deutlich, daß sie Tyrosin oxydieren. Diese methode ist in ihrer Aussagekraft mit Radioassay vergleichbar.
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This work was supported in part by INSERM grant ATP 78-99
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Menezo, Y., Voulot, C., Ortonne, J.P. et al. A rapid quantitative analysis of tyrosine and its oxydation products by tyrosinase. Arch Dermatol Res 263, 267–272 (1978). https://doi.org/10.1007/BF00446943
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DOI: https://doi.org/10.1007/BF00446943