Determination of octopamine and tyramine traces in dietary supplements and phytoextracts by high performance liquid chromatography after derivatization with 2,5-dimethyl-1H-pyrrole-3,4-dicarbaldehyde

The use of 2,5-dimethyl-1H-pyrrole-3,4-dicarbaldehyde (DPD) as a pre-column derivatization reagent for HPLC (high performance liquid chromatography) analysis of octopamine (oct) and tyramine (tyr) is proposed. The compound reacts under mild conditions (2 min at ambient temperature) with primary amino groups. The derivatization conditions were optimized by considering different parameters (temperature, time and reagent concentration). The synthesized oct and tyr adducts were characterized by H-1 NMR (nuclear magnetic resonance). ESI-MS (electrospray ionization mass spectrometry), IR (infrared) and UV (ultraviolet). Derivative chromatographic separations were performed on a Sinergy Hydro-RP column (150 mm x 4.6 mm i.d.) using a mobile phase consisting of methanol and triethylammonium phosphate buffer (pH 3; 10 mM) at varying composition gradient elution and at a flow rate of 0.8 mL/min. Detection was set at lambda = 320 nm. The obtained results were compared with those achieved by a validated direct HPLC method with detection at lambda = 275 nm using a Sinergy Polar-RP column (250 mm x 3 mm i.d.) by isocratic elution conditions with a mobile phase consisting of methanol/acetonitrile/sodium pentane-sulphonate (SPS; pH 3; 10 mM), 7.5:7.5:85 (v/v/v) at a flow rate of 0.3 mL/min. Derivatization method sensitivity proved to be ten times higher than direct method. Limit of detection of oct and tyr was 0.010 and 0.008 mu g/mL, respectively. The reliability of the pre-column method was satisfactory also in terms of linearity (from 0.028 to 1.255 and 0.024 to 1.244 mu g/mL for oct and tyr, respectively), precision (relative standard deviation <= 2, without significant differences between intra-day and inter-day data) and recovery (from 98.9 to 101.2%). The proposed method showed to be suitable for a reliable determination of oct and tyr traces in commercially available phytoproducts using the instrumentation usually present in any analytical laboratory. (C) 2011 Elsevier B.V. All rights reserved.