The kynurenine pathway generates multiple tryptophan metabolites called collectively kynurenines and leads to formation of the enzyme cofactor nicotinamide adenine dinucleotide. The first step in this pathway is tryptophan degradation, initiated by the rate‐limiting enzymes indoleamine 2,3‐dioxygenase, or tryptophan 2,3‐dioxygenase, depending on the tissue. The balanced kynurenine metabolism, which has been a subject of multiple studies in last decades, plays an important role in several physiological and pathological conditions such as infections, autoimmunity, neurological disorders, cancer, cataracts, as well as pregnancy. Understanding the regulation of tryptophan depletion provide novel diagnostic and treatment opportunities, however it requires reliable methods for quantification of kynurenines in biological samples with complex composition (body fluids, tissues, or cells). Trace concentrations, interference of sample components, and instability of some tryptophan metabolites need to be addressed using analytical methods. The novel separation approaches and optimized extraction protocols help to overcome difficulties in analyzing kynurenines within the complex tissue material. Recent developments in chromatography coupled with mass spectrometry provide new opportunity for quantification of tryptophan and its degradation products in various biological samples. In this review, we present current accomplishments in the chromatographic methodologies proposed for detection of tryptophan metabolites and provide a guide for choosing the optimal approach.

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Metabolites and enzymes involved in the kynurenine pathway (KP) are highly promising targets for cancer treatment, including gastrointestinal tract diseases. Thus, accurate quantification of these compounds in body fluids becomes increasingly important. The aim of this study was the development and validation of the UHPLC-ESI-MS/MS methods for targeted quantification of biologically important KP substrates (tryptophan and nicotinamide) and metabolites(kynurenines) in samples of serum and peritoneal fluid from gastric cancer patients. The serum samples were simply pretreated with trichloroacetic acid to precipitate proteins. The peritoneal fluid was purified by solid-phase extraction before analysis. Validation was carried out for both matrices independently. Analysis of the samples from gastric cancer patients showed different accumulations of tryptophan and its metabolites in different biofluids of the same patient. The protocols will be used for the evaluation of tryptophan and kynurenines in blood and peritoneal fluid to determine correlation with the clinicopathological status of gastric cancer or the disease’s prognosis.

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