TY - JOUR
T1 - Determination of endogenous melatonin in the individual pineal glands of inbred mice using precolumn oxidation reversed-phase micro-high-performance liquid chromatography
AU - Tomita, Tatsunosuke
AU - Hamase, Kenji
AU - Hayashi, Hiromi
AU - Fukuda, Hiroko
AU - Hirano, Junzo
AU - Zaitsu, Kiyoshi
N1 - Funding Information:
The authors gratefully acknowledge Dr. Tomofumi Miyamoto and Dr. Takeyuki Akita of Kyushu University for their valuable help in the NMR and X-ray diffraction analysis. This work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan.
PY - 2003/5/15
Y1 - 2003/5/15
N2 - 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 380nm, 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[Lmol-1cm-1] and 0.31 (245nm), respectively. The lower quantification limit of melatonin in biological samples using this precolumn oxidation method is 200amol, and the calibration curve of spiked melatonin is linear from 200amol to 50fmol (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 18.8fmol/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.
AB - 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 380nm, 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[Lmol-1cm-1] and 0.31 (245nm), respectively. The lower quantification limit of melatonin in biological samples using this precolumn oxidation method is 200amol, and the calibration curve of spiked melatonin is linear from 200amol to 50fmol (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 18.8fmol/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.
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U2 - 10.1016/S0003-2697(03)00079-4
DO - 10.1016/S0003-2697(03)00079-4
M3 - Article
C2 - 12711335
AN - SCOPUS:0037447866
VL - 316
SP - 154
EP - 161
JO - Analytical Biochemistry
JF - Analytical Biochemistry
SN - 0003-2697
IS - 2
ER -