Pheomelanin-related benzothiazole isomers in the urine of patients with diffuse melanosis of melanoma

Abstract

Background: Currently used as structural markers for pheomelanin identification and quantitation, benzothiazole compounds derived from isomers of cysteinyldopa have been indicated by recent in vitro studies as new potential pheomelanogenesis intermediates. The presence of benzothiazole compounds in the urine of patients with melanoma with or without diffuse melanosis was investigated.

Methods: Hydrophilic interaction liquid chromatography with zwitterionic stationary phase (ZIC-HILIC) and photo-diode array (PDA) detection was used for analysis of 6-(2-amino-2-carboxyethyl)-4-hydroxybenzothiazole-2-carboxylic acid (BTCA-5), and 7-(2-amino-2-carboxyethyl)-4-hydroxybenzothiazole-2-carboxylic acid (BTCA-2), derived from 5-S-cysteinyldopa (5-S-CD) and 2-S-cysteinyldopa (2-S-CD) isomers, respectively. After minimal sample preparation, isocratic chromatography allowed efficient separation of the compounds, which were safely identified by their typical absorption features.

Results: Three patients with diffuse melanosis, 16 patients with melanoma (stages III and IV) and three healthy subjects were investigated. The urinary BTCAs were found to be highly associated with melanosis but more loosely to excreted 5-S-CD. Analysis of the pigmented fraction of urine following alkaline hydrogen peroxide degradation and quantitation of BTCAs provided evidence for the presence of pheomelanins at high levels in patients with melanosis.

Conclusion: Identification of free BTCA isomers in urine provides a significant contribution in the field of urinary melanogens, and has important implications for biosynthetic activity of normal and pathologic melanocytes.