ABSTRACT
The multistep synthesis and negative ion-ESI fragmentation pattern of [methyl-D(3)](2)hypericin (1-D(6)) is described. The application of 1-d(6) as internal standard for the quantification of hypericin (1) in the ng mL(-1) range in human plasma by isotope-dilution LC-MS is demonstrated. The hypericin-containing plasma samples are spiked with 1-D(6), deproteinized and extracted with ethyl acetate. The extracts are injected into a HPLC-ESI-ion-trap system and the mass-separated negative ions from 1 and 1-D(6) are analysed. From their intensities linear standard curves over the concentration range from 1 to 10 ng mL(-1) are obtained. Accuracy, precision and recovery are discussed.
Subject(s)
Perylene/analogs & derivatives , Perylene/blood , Spectrometry, Mass, Electrospray Ionization/methods , Anthracenes , Deuterium , Humans , Molecular Structure , Perylene/chemistry , Reference Standards , Sensitivity and Specificity , Spectrometry, Mass, Electrospray Ionization/standardsABSTRACT
The oxidative biotransformation of the anticancer drug 7-hydroxy-2-[2-[(2-hydroxyethyl)amino]ethyl]-5-[[2-[(2-hydroxyethyl)amino]ethyl]amino]anthra[1,9-cd]pyrazol-6(2H)-one dihydrochloride (losoxantrone, CI-941) after incubation of primary cultures of rat hepatocytes has been investigated. The structures of twelve losoxantrone metabolites have been elucidated by means of high-performance liquid chromatography-mass spectometry, tandem mass spectrometry, and two-dimensional NMR. In these mammalian hepatocytes, the CI-941 biotransformation includes a monohydroxylation of the phenolic substructure of the CI-941-chromophore via cytochrome P450 catalysis, resulting in metabolites having an ortho- and para-hydroquinonoid substructure, respectively. The identification of a glutathione conjugate as a follow-up metabolite confirms the oxidative activation of the ortho-hydroxylated losoxantrone metabolite. The oxidative activation establishes the ability of CI-941 to form covalent bonds to intracellular nucleophilic targets. Furthermore, the CI-941 metabolism was shown to be extremely suppressed in rat hepatocytes incubated with metyrapone. In contrast to these results, human tumor HepG2 cells did not show any CI-941 biotransformation after incubation.
