title: Synthesis and characterization of N-demethylated metabolites of malachite green and leucomalachite green.
abstract: Malachite green (MG), a triphenylmethane dye used to treat fungal and protozoan infections in fish, undergoes sequential oxidation to produce various N-demethylated derivatives (monodes-, dides(sym)-, dides(unsym)-, trides-, and tetrades-) both before and after reduction to leucomalachite green (LMG). The close structure resemblance of the metabolites with aromatic amine carcinogens implicates a potential genotoxicity from exposure to MG. The availability of the synthetic standards is important for metabolic and DNA adduct studies of MG. This paper describes a simple and versatile method for the synthesis of MG, LMG, and their N-demethylated metabolites. The synthesis involves a coupling of 4-(dimethylamino)benzophenone or 4-nitrobenzophenone with the aryllithium reagents derived from appropriately substituted 4-bromoaniline derivatives, followed by treatment with HCl in methanol. The resulting cationic MG and their leuco analogues showed systematic UV/vis spectral and tandem mass fragmentation patterns consistent with sequential N-demethylation. The extensive (1)H and (13)C spectral assignments of the metabolites were aided by the availability of (13)C(7)-labeled MG and LMG. The results indicate the existence of a resonance structure with the cationic charge located in the central methane carbon (C(7)). The synthetic procedure is general in scope so that it can be extended to the preparation of N-demethylated metabolites of other structurally related N-methylated triphenylmethane dyes.
authors.affiliation: Department of Biomedical Sciences, College of Pharmacy, University of Rhode Island, Kingston, Rhode Island 02881, USA. bcho@uri.edu
authors.forename: Bongsup P
authors.initials: BP
authors.lastname: Cho
authors.forename: Tianle
authors.initials: T
authors.lastname: Yang
authors.forename: Lonnie R
authors.initials: LR
authors.lastname: Blankenship
authors.forename: Joanna D
authors.initials: JD
authors.lastname: Moody
authors.forename: Mona
authors.initials: M
authors.lastname: Churchwell
authors.forename: Frederick A
authors.initials: FA
authors.lastname: Beland
authors.forename: Sandra J
authors.initials: SJ
authors.lastname: Culp
mesh_terms: D000814:Aniline Compounds / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D001577:Benzophenones / Q000737:chemistry; D005659:Fungicides, Industrial / Q000737:chemistry / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry / Q000379:methods; D008745:Methylation; D012394:Rosaniline Dyes / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D013056:Spectrophotometry, Ultraviolet
chemical_list: D000814:Aniline Compounds; D001577:Benzophenones; D005659:Fungicides, Industrial; D012394:Rosaniline Dyes; C005095:malachite green; C015320:leucomalachite green