A new one-pot technique for obtaining potential indapamide metabolites by oxidation and conjugation on MALDI target

Introduction. Metabolic activation of xenobiotics, including pharma drugs, is considered to be one of the main mechanisms for the development of idiosyncratic reactions. Accordingly, the potential bioactivation of a xenobiotic should be carefully evaluated in the early stages of drug development. In this regard, the search for new rapid and effective screening techniques for reactive metabolites of xenobiotics presents particular interest. Objective . Development of a new technique for modeling the processes of xenobiotic biotransformation in vitro to identify potential metabolites of indapamide. Materials and methods . Non-enzymatic instrumental methods, such as electrochemical oxidation (ECO) and photocatalytic oxidation (PCO) in volume, were used as comparison methods. The second phase of metabolism was modeled by incubating the oxidation products of indapamide with a trapping agent (glutathione, GSH). The oxidation products, as well as their conjugates with GSH, were then analyzed by high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS). The developed one-pot technique for metabolism modeling is based on a UV-induced PCO of a xenobiotic in the presence of GSH on the surface of a target functionalized with titanium dioxide followed by detection of the products by matrix-assisted laser desorption/ionization mass spectrometry (MALDI). Results. In use of ECO resulted in the detection of 5 metabolites and 3 adducts with GSH, while the use of PCO in the volume allowed detection of 7 metabolites and 1 adduct with GSH. The new one-pot technique detected 8 adducts with GSH. In addition to the detection of a number of known indapamide metabolites and their conjugates with GSH, a total of 4 previously unstudied metabolites and adducts with GSH were each detected for indapamide by the three methods. Conclusions. In comparison with ECO and PCO in volume, the proposed analytical technique for modeling indapamide metabolism showed its higher informativity combined with simplicity and rapidity, which makes it a promising candidate for use in preclinical studies of drugs in predicting the metabolism and toxicity of pharmaceutical objects, as well as in studying the biotransformation processes of various xenobiotics.