A specific, sensitive and reproducible high-performance liquid chromatographic (HPLC) method is described for the simultaneous measurement of 6-hydroxymelatonin and melatonin in rat plasma. Using this technique, a dose-dependent increase in 6-hydroxymelatonin concentration was observed 10 min after administration of different doses of melatonin. Plasma 6-hydroxymelatonin represented 1% of plasma melatonin, irrespective of the melatonin dose administered. After administration of 100 µg melatonin, 6-hydroxymelatonin could be detected in plasma from 7.5 to 60 min. In the same experiment, plasma 6-sulfatoxymelatonin concentrations determined by radioimmunoassay could be detected from 7.5 to 120 min. This HPLC technique allows the measurement of 6-hydroxymelatonin together with melatonin. Combined with an RIA for 6-sulfatoxymelatonin, it could be useful in determining the site of action of drugs affecting melatonin metabolism.
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