| 15560919 |
Eggenreich K, Golouch S, Toscher B, Beck H, Kuehnelt D, Wintersteiger R: Determination of 4-amino-m-cresol and 5-amino-o-cresol and metabolites in human keratinocytes (HaCaT) by high-performance liquid chromatography with DAD and MS detection. J Biochem Biophys Methods. 2004 Oct 29;61(1-2):23-34.
A multi-step gradient HPLC system combined with DAD and MS detection has been developed for the determination of the oxidation hair dyes 4-amino-m-cresol (4-AC) and 5-amino-o-cresol (5-AC) and their metabolites in the alternative testing system human keratinocytes (HaCaT) cell culture. The culture medium induced by 3-methylcholanthrene (3-MC) was fortified with 4-AC or 5-AC and incubated for 24 h at 37 degrees C in order to produce metabolites. After several pre-cleaning steps, further cleaning was done by solid-phase extraction using C18 phenyl cartridges. Optimizing chromatographic conditions, a hybrid-based RP8 column was most suitable for the separation of the metabolites formed in HaCaT. Only one conjugation product, the N-acetylated derivative, could be identified for both 4-AC and 5-AC by LC/DAD/MS. The ionisation technique used for MS analysis was Atmospheric Pressure Ionization (API). |
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