Simultaneous Measurement of Kynurenine and Tryptophan in Human Plasma and Supernatants of Cultured Human Cells by HPLC with Coulometric Detection

l-Tryptophan (Trp) is an essential amino acid and an abundant protein component. Trp is metabolized in mammals via two pathways: biosynthesis of the neurotransmitter serotonin and the kynurenine (Kyn) pathway (1)(2). Trp is the substrate for the first step in both of these pathways. In the Kyn pathway, the indole ring of Trp can be opened by the enzymes tryptophan pyrrolase \[tryptophan 2,3-dioxygenase and indoleamine 2,3-dioxygenase (IDO)\] (3). The reactions catalyzed by both of these enzymes constitute the rate-limiting step in the Kyn metabolic pathway (4). The organ distribution of the two enzymes differs, allowing them to be clearly distinguished (5)(6). Tryptophan 2,3-dioxygenase is located primarily in the liver, and its activity is up-regulated in response to Trp and metabolic steroids. IDO can be found in various cells, but it is not active in healthy humans and its activity is induced only during immune responses mediated by proinflammatory cytokines such interferon-γ (7)(8).The enzymatic reactions of the Kyn pathway produce NAD and other intermediates, including Kyn and quinolinic acid (5)(9). The removal of Trp from the microenvironment via this pathway protects the organism by limiting the growth of intracellular pathogens and malignant cells and may also prevent maternal immune reactions against the fetus during pregnancy (10)(11)(12)(13)(14). The Kyn-to-Trp ratio provides an estimate of IDO activity that is independent of baseline Trp concentrations (15)(16)(17). In plasma, Kyn/Trp may be correlated with the concentrations of inflammatory markers, such as neopterin, to ensure that Trp degradation is induced by interferon-γ and involves IDO (18). Kyn/Trp is higher in individuals with diseases that involve an activated cellular immune response or those undergoing cytokine immunotherapy (16)(17)(19).Here we …