Identification and characterization of reaction products of 5‐hydroxytryptamine with methylglyoxal and glyoxal by liquid chromatography/tandem mass spectrometry

Rationale Methylglyoxal (MGO) and glyoxal (GO) are known to be at high levels in humans with diabetes. They react with amine‐containing proteins and amino acids to form advanced glycation end products, however, their reactivity with other amine‐containing metabolites, such as neurotransmitters, has not been explored. In this study, we aimed at studying the reactivity of 5‐hydroxytryptamine (5‐HT) with MGO or GO, which may alter the metabolic function of 5‐HT. Methods Stock solutions of 5‐HT, MGO and GO were made in PBS buffer at pH 7.4 and 5‐HT was incubated with MGO or GO at different concentrations. The reactions were also performed at physiological concentrations. The reaction mixtures collected at different incubation times were analyzed by direct ESI‐HRMS, LC/MS and LC/MS/MS to detect/characterize the products. Agilent 6545 Q‐TOF and Agilent 6420 triple quadrupole mass spectrometers were used for the study, and LC separations were performed on a C18 column. Results The direct ESI‐HRMS data of the reaction mixtures showed formation of three and four reaction products when 5‐HT was reacted with MGO and GO, respectively. All the products showed dominant [M + H]+ ions. The products were characterized by HRMS, LC/MS/MS and literature reports on similar compounds. The products can easily be identified by LC/MS based on the accurate mass values together with retention time information. The MS/MS of the reaction products showed structure‐indicative fragment ions. Conclusions 5‐HT reacts with one or two MGO/GO to form a set of reaction products. The reaction between 5‐HT and MGO or GO was faster at higher concentrations of MGO/GO (