Improved HPLC Conditions to Determine Eumelanin and Pheomelanin Contents in Biological Samples Using an Ion Pair Reagent.

International Journal of Molecular Sciences
Shosuke ItoKazumasa Wakamatsu

Abstract

Alkaline hydrogen peroxide oxidation (AHPO) of eumelanin and pheomelanin, two major classes of melanin pigments, affords pyrrole-2,3,5-tricarboxylic acid (PTCA), pyrrole-2,3-dicarboxylic acid (PDCA) and pyrrole-2,3,4,5-tetracarboxylic acid (PTeCA) from eumelanin and thiazole-2,4,5-tricarboxylic acid (TTCA) and thiazole-4,5-dicarboxylic acid (TDCA) from pheomelanin. Quantification of these five markers by HPLC provides useful information on the quantity and structural diversity of melanins in various biological samples. HPLC analysis of these markers using the original method of 0.1 M potassium phosphate buffer (pH 2.1):methanol = 99:1 (85:15 for PTeCA) on a reversed-phase column had some problems, including the short lifetime of the column and, except for the major eumelanin marker PTCA, other markers were occasionally overlapped by interfering peaks in samples containing only trace levels of these markers. These problems can be overcome by the addition of an ion pair reagent for anions, such as tetra-n-butylammonium bromide (1 mM), to retard the elution of di-, tri- and tetra-carboxylic acids. The methanol concentration was increased to 17% (30% for PTeCA) and the linearity, reproducibility, and recovery of the markers with th...Continue Reading

References

Aug 6, 1986·Biochimica Et Biophysica Acta·S Ito
Oct 1, 1996·Pigment Cell Research·H OzekiA J Thody
May 25, 2002·Pigment Cell Research·Kazumasa Wakamatsu, Shosuke Ito
May 25, 2002·Pigment Cell Research·Kazumasa WakamatsuJonathan L Rees
Nov 7, 2006·Pigment Cell Research·S Del BinoF Bernerd
Apr 26, 2008·Photochemistry and Photobiology·Shosuke Ito, Kazumasa Wakamatsu
Mar 8, 2011·Pigment Cell & Melanoma Research·Tomohisa Hirobe
May 23, 2012·Proceedings of the National Academy of Sciences of the United States of America·Keely GlassJohn D Simon
May 29, 2013·Pigment Cell & Melanoma Research·Marco d'IschiaShosuke Ito
Jul 16, 2015·Pigment Cell & Melanoma Research·Marco d'IschiaShosuke Ito
Aug 19, 2015·Pigment Cell & Melanoma Research·Sandra Del BinoFrançoise Bernerd
Jul 19, 2016·Skin Research and Technology : Official Journal of International Society for Bioengineering and the Skin (ISBS) [and] International Society for Digital Imaging of Skin (ISDIS) [and] International Society for Skin Imaging (ISSI)·H MatsunakaF Furukawa
Aug 10, 2016·Pigment Cell & Melanoma Research·Grzegorz SzewczykTadeusz Sarna
Nov 25, 2017·Pigment Cell & Melanoma Research·Shosuke ItoKazumasa Wakamatsu
Mar 22, 2019·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·Benjamin RiouxMercedes Quintana
Jun 25, 2019·Pigment Cell & Melanoma Research·Catharina Margrethe LercheHans Christian Wulf
Aug 21, 2019·Proceedings of the National Academy of Sciences of the United States of America·Valentina RossiKazumasa Wakamatsu
Dec 23, 2019·The Journal of Investigative Dermatology·Bao Chau K LySandra Del Bino

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Citations

Dec 31, 2020·International Journal of Molecular Sciences·Martin JarenmarkJohan Lindgren
Mar 23, 2021·Pigment Cell & Melanoma Research·Kazumasa WakamatsuShosuke Ito
Aug 27, 2021·Photochemistry and Photobiology·Kazumasa WakamatsuShosuke Ito

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