Quantitation of eumelanin and pheomelanin markers in diverse biological samples by HPLC-UV-MS following solid-phase extraction

Autoři: Susanne Affenzeller aff001;  Holm Frauendorf aff002;  Tobias Licha aff003;  Daniel J. Jackson aff001;  Klaus Wolkenstein aff001
Působiště autorů: Department of Geobiology, Georg-August University, Göttingen, Germany aff001;  Institute of Organic & Biomolecular Chemistry, Georg-August University, Göttingen, Germany aff002;  Department of Applied Geology, Georg-August University, Göttingen, Germany aff003
Vyšlo v časopise: PLoS ONE 14(10)
Kategorie: Research Article
doi: https://doi.org/10.1371/journal.pone.0223552


Eumelanin and pheomelanin are well known and common pigments found in nature. However, their complex polymer structure and high thermostability complicate their direct chemical identification. A widely used analytical method is indirect determination using HPLC with UV detection of both types of melanin by their most abundant oxidation products: pyrrole-2,3-dicarboxylic acid (PDCA), pyrrole-2,3,5-tricarboxylic acid (PTCA), thiazole-4,5-dicarboxylic acid (TDCA), and thiazole-2,4,5-tricarboxylic acid (TTCA). An increasing interest in pigmentation in biological research led us to develop a highly sensitive and selective method to identify and quantify these melanin markers in diverse biological samples with complex matrices. By introducing solid-phase extraction (SPE, reversed-phase) following alkaline oxidation we could significantly decrease background signals while maintaining recoveries greater than 70%. Our HPLC-UV-MS method allows for confident peak identification via exact mass information in corresponding UV signals used for quantitation. In addition to synthetic melanin and Sepia officinalis ink as reference compounds eumelanin markers were detected in brown human hair and a brown bivalve shell (Mytilus edulis). Brown feathers from the common chicken (Gallus g. domesticus) yielded all four eumelanin and pheomelanin markers. The present method can be easily adapted for a wide range of future studies on biological samples with unknown melanin content.

Klíčová slova:

High performance liquid chromatography – Chickens – Melanin – Oxidation – Pigments – Percutaneous transluminal coronary angioplasty – Feathers – Ultraviolet spectroscopy


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