title: Radioimmunoassay of N-acetyl-N-formyl-5-methoxykynuramine (AFMK): a melatonin oxidative metabolite.
abstract: N-acetyl-N-formyl-5-methoxykynuramine (AFMK) is a melatonin metabolite identified in rat brain by Hirata et al. (The Journal of Biological Chemistry 249 (1974) 1311). Since no assay has been described for its routine measurement, we have developed and validated such a radioimmunoassay. We synthesized AFMK and N-acetyl-5-methoxykynuramine (AMK), in order to produce anti-AFMK antibodies and to standardize the assay. The tracer [3H]-AFMK was obtained from [3H]-melatonin. The assay was preceded by a chromatographic step on Celite microcolumn in order to increase its specificity. The assay was suitable for the measurement of AFMK levels ranging from 59 to 1894 pmol/L. The detection limit of the assay was routinely set at 65 pmol/L. The intra- and inter-assay coefficients of variation were 3.5% and 11% respectively. Investigation of the 24 h plasma pattern in healthy volunteers did not reveal any AFMK levels in plasma samples. In rats, plasma AFMK showed a peak after melatonin injection, which confirmed the in vivo AFMK production as a melatonin metabolite. This AFMK assay is suitable for studies on melatonin metabolism.
authors.affiliation: Service de Radioanalyse, Hôpital Neurocardiologique, Centre de Médecine Nucléaire, 59 boulevard Pinel, 69394 Lyon, France.
authors.forename: Catherine
authors.initials: C
authors.lastname: Harthé
authors.forename: Danièle
authors.initials: D
authors.lastname: Claudy
authors.forename: Henri
authors.initials: H
authors.lastname: Déchaud
authors.forename: Berthe
authors.initials: B
authors.lastname: Vivien-Roels
authors.forename: Paul
authors.initials: P
authors.lastname: Pévet
authors.forename: Bruno
authors.initials: B
authors.lastname: Claustrat
mesh_terms: D000818:Animals; D007692:Diatomaceous Earth / Q000737:chemistry; D006801:Humans; D007735:Kynuramine / Q000031:analogs & derivatives* / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D008297:Male; D008550:Melatonin / Q000494:pharmacology; D000442:Octanols / Q000737:chemistry; D010084:Oxidation-Reduction; D011817:Rabbits; D011863:Radioimmunoassay / Q000379:methods*; D051381:Rats; D017208:Rats, Wistar
chemical_list: D000442:Octanols; D007735:Kynuramine; C041550:N-acetyl-N-formyl-5-methoxykynurenamine; C032131:N-acetyl-5-methoxy kynurenamine; D007692:Diatomaceous Earth; D008550:Melatonin