Determination of eumelanin and pheomelanin in melanomas using solid-phase extraction and high performance liquid chromatography-diode array detection (HPLC-DAD) analysis.
J Chromatogr B Analyt Technol Biomed Life Sci
; 1113: 60-68, 2019 Apr 15.
Article
em En
| MEDLINE
| ID: mdl-30897406
ABSTRACT
Determination of eumelanin and pheomelanin in melanomas that exhibit different pigmentation was carried using a solid-phase extraction (SPE) preparation method based on weak anion exchange chemistry. This extraction significantly enhanced the chromatographic profile obtained by reverse phase high performance liquid chromatography-diode array detection (RP-HPLC-DAD). The SPE method was developed using aqueous standards of melanin markers thiazole-2,4,5-tricarboxylic acid (TTCA), thiazole-4,5-dicarboxylic acid (TDCA), pyrrole-2,3-dicarboxylic acid (PDCA) and pyrrole-2,3,5-tricarboxylic acid (PTCA) and non-pigmented cell lines spiked with those markers. An excellent average recovery, above 90%, was obtained for the four markers with a relative standard deviation below 7%. We have also optimized the stationary phase and the mobile phase (phosphate concentration and pH) to improve sensitivity and to reduce the analysis time. Elution of the four markers is achieved in 5â¯min and total analysis of biological samples is completed in 15â¯min. The quantification limits for TDCA, TTCA, PDCA and PTCA are 60, 50, 47 and 48â¯ng/mL respectively. Furthermore, DAD detection improves the marker identification in complex matrices through the analysis of UV spectra. We have successfully applied this method to melanoma tumors and cells. Murine B16BL6 tumor are highly pigmented with mostly eumelanin (98.1% of eumelanin) while human SK-MEL-3 tumor contain about 30% pheomelanin. B16BL6 and B16F10 are eumelanic cells lines and NHEM melanocytes contain about 24% of pheomelanin.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Temas:
Geral
/
Tipos_de_cancer
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Pele
Base de dados:
MEDLINE
Assunto principal:
Cromatografia Líquida de Alta Pressão
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Extração em Fase Sólida
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Melaninas
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Melanoma
Tipo de estudo:
Diagnostic_studies
/
Prognostic_studies
Limite:
Animals
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Humans
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Male
Idioma:
En
Revista:
J Chromatogr B Analyt Technol Biomed Life Sci
Assunto da revista:
ENGENHARIA BIOMEDICA
Ano de publicação:
2019
Tipo de documento:
Article
País de afiliação:
França