␣-1-Microglobulin carries a set of covalently linked chromophores that give it a peculiar yellow-brown color, fluorescence properties, and both charge and size heterogeneity. In this report it is shown that these features are due to the adducts with the tryptophan metabolite, 3-hydroxykynurenine, and its autoxidation products and that the modification is more pronounced in the protein isolated from urine of hemodialyzed patients. The light yellow amniotic fluid ␣-1-microglobulin acquires the optical properties and charge heterogeneity of the urinary counterpart following incubation with kynurenines. The colored amino acid adducts of urinary and amniotic fluid ␣-1-microglobulins were separated by chromatography after acid hydrolysis and analyzed by mass spectrometry. Human serum albumin samples, native and treated with 3-hydroxykynurenine in the presence of oxygen, were used as a control. The retention times and mass fragmentation products were compared, and a lysyl adduct with hydroxantommathin was identified in the urinary ␣-1-microglobulin and in the modified albumin samples. The more extensive modification of the urinary protein appears to be correlated with uremia, a condition in which the catabolism of tryptophan via the kynurenine pathway is increased, and the consequent rise in the concentration of its derivatives is accompanied by the oxidative processes due to the hemodialysis treatment. The oxidative derivatives of 3-hydroxykynurenine, which are known to act as protein crosslinking agents, are the likely cause of the propensity of urinary ␣-1-microglobulin to form dimers and oligomers. This process, as well as the redox properties of these metabolites, may contribute to the toxic effects of the kynurenine species.1 also known as protein HC, is a widely distributed glycosylated protein that on the basis of sequence homology has been included in the lipocalin family. This large group of predominantly extracellular molecules shares a common -barrel fold but has been assigned different functional roles (1). ␣-1-m is present in many tissues and, although its function is not known, several reports hint at a role as an immunomodulator (2, 3). Its single polypeptide chain consists of 183 residues (4) and contains one O-linked and two N-linked oligosaccharide moieties (5). The synthesis takes place mainly in the liver and, in humans, the ␣-1-m gene has been mapped to chromosome 9 in a region where other lipocalin genes are clustered (4, 6). As for all other species, it encodes also bikunin, the light chain of a plasma proteinase inhibitor family (7), and the precursor polypeptide chain contains an internal basic tripeptide recognized by a specific endoprotease. The cleavage occurs in the Golgi apparatus before secretion and, so far, no functional correlation between the lipocalin and bikunin has been found. ␣-1-m is then catabolized in the proximal tubule cells of the kidneys, and its concentration increases in urines of patients with renal failure (8). The protein has thus been recognized as a very sensitive ...