Determination of serotonin and melatonin in rat pineal gland by high-performance liquid chromatography with ultraviolet and fluorometric dual detection.
High-performance liquid-chromatography (HPLC) of serotonin and melatonin was carried out on a reversed-phase column; the compounds were determined by fluorescence measurement with excitation and emission wavelengths of 285 and 345 nm, respectively. The indoles were clearly separated and eluted with a single mobile phase within 20 min. 4-Aminoantipyrine was used as an internal standard (detected by ultraviolet (UV) absorbance measurement). Satisfactory recovery of serotonin and melatonin was obtained by using this dual detection system (UV and fluorescence). The detection limits of the indoles were all in the low picogram range. The proposed method was applied to the determination of serotonin and melatonin in rat pineal gland. A distinct variation between light and dark periods was found in both serotonin and melatonin levels in the rat pineal gland.