Determination of serotonin and its precursors in chocolate samples by capillary liquid chromatography with mass spectrometry detection

A method for the analysis of serotonin (5-HT) and its precursors, 5-hydroxytryptophan (5-HTP) and L-tryptophan (TP) in chocolate samples by capillary liquid chromatography-mass spectrometry (cLC-MS) has been developed. Optimum chromatographic conditions were established by using a personalized multifactorial experimental design. Finally the cLC separation was achieved through a mixture of acetonitrile and 5 mM ammonium formate at pH 4 (3:97, v/v) as mobile phase in gradient elution, setting the injection volume at 10 mu L and using pure water as injection solvent for focusing purposes on the head of the capillary column. For extraction of targets in chocolate samples a new, fast and simple procedure based on the use of acidic extraction medium and sonication was developed. Working in selected ion mode (m/z 177 for 5-HT, m/z 205 for L-tryptophan and m/z 221 for 5-HTP) detection limits were between 0.01 and 0.11 mu g g(-1) and linearity was in the concentration range of 0.5-25 mu g g(-1). Recoveries higher than 76% with RSDs lower than 8% were obtained from spiked samples for all analytes, showing the effectiveness of the proposed method. Serotonin and its precursors were determined in 5 kinds of commonly consumed chocolates with different cocoa contents (70-100%). The highest serotonin content was found in chocolate with a cocoa content of 85% (2.93 mu g g(-1)). Regarding L-tryptophan, the highest content of this amino acid (13.27-13.34 mu g g(-1)) was found in chocolate samples with the lowest cocoa content (70-85%). 5-Hydroxytryptophan was not detected in any chocolate samples. (C) 2011 Elsevier B.V. All rights reserved.