Serotonin monitoring in microdialysate from rat brain by microbore-liquid chromatography with fluorescence detection

A sensitive fluorimetric microbore-liquid chromatographic method for the determination of serotonin in microdialysate from rat brain was developed. The method is based on the fluorescence derivatization of serotonin by reaction with benzylamine in the presence of potassium hexacyanoferrate(III). A microdialysis probe was implanted in the rat brain, and continuously perfused at 2.0 μl min−1 with Ringer solution. The microdialysate, collected every 5 min, was added with a reagent solution composed of 0.3 M CAPS buffer (pH 12.0), 0.2 M benzylamine, 0.1 M potassium hexacyanoferrate(III) and methanol (1:1:5:10, v/v). The derivatization reaction was completed by standing at room temperature for 2 min. The benzylamine derivative of serotonin could be separated within 10 min on a reversed-phase microbore column (100×1.0 mm i.d., 5 μm TSK gel ODS-80TM) with isocratic elution. The detection limit (signal-to-noise ratio=3) of serotonin is 80 amol for a 5 μl injection. The effects of increased potassium ion concentration in the Ringer solution and a single intraperitoneal injection of methamphetamine on the serotonin level in microdialysate were examined.