title: Determination of endogenous melatonin in the individual pineal glands of inbred mice using precolumn oxidation reversed-phase micro-high-performance liquid chromatography.
abstract: The amount of endogenous melatonin in the individual pineal glands of inbred mice has been determined using reversed-phase micro-high-performance liquid chromatography after precolumn oxidation of melatonin to a compound having strong fluorescence. The fluorescent compound was identified as N-[(6-methoxy-4-oxo-1,4-dihydroquinolin-3-yl)methyl]acetamide. The excitation and emission wavelengths of this compound are 245 and 380 nm, respectively, and the fluorescence intensity is 6.8 times greater than that of melatonin. Molar absorptivity and fluorescence quantum yield of this compound are 46,300[L mol(-1)cm(-1)] and 0.31 (245 nm), respectively. The lower quantification limit of melatonin in biological samples using this precolumn oxidation method is 200 amol, and the calibration curve of spiked melatonin is linear from 200 amol to 50 fmol (r>0.999). The sensitivity of the present method is almost 10 times higher than that of the previous method. The values of endogenous melatonin obtained for ICR, C57BL, BALB/c, and AKR mice are 4.7, 6.1, 7.4, and 18.8 fmol/pineal gland, respectively. The amounts of endogenous pineal melatonin of these strains had not been clearly reported due to the poor enzymatic activities for melatonin biosynthesis; this is the first report that clearly demonstrates the existence of endogenous melatonin in these inbred mice.
authors.affiliation: Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Tatsunosuke
authors.initials: T
authors.lastname: Tomita
authors.forename: Kenji
authors.initials: K
authors.lastname: Hamase
authors.forename: Hiromi
authors.initials: H
authors.lastname: Hayashi
authors.forename: Hiroko
authors.initials: H
authors.lastname: Fukuda
authors.forename: Junzo
authors.initials: J
authors.lastname: Hirano
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Zaitsu
mesh_terms: D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005453:Fluorescence; D007460:Iodoacetamide / Q000031:analogs & derivatives* / Q000592:standards; D009682:Magnetic Resonance Spectroscopy; D008550:Melatonin / Q000031:analogs & derivatives / Q000032:analysis*; D051379:Mice; D008815:Mice, Inbred Strains; D008832:Microchemistry / Q000379:methods; D015394:Molecular Structure; D010084:Oxidation-Reduction; D010870:Pineal Gland / Q000737:chemistry / Q000378:metabolism*; D012015:Reference Standards
chemical_list: C091256:N-methyliodoacetamide; D008550:Melatonin; D007460:Iodoacetamide