Formation of Kynurenic and Xanthurenic Acids

Kynurenic acid was discovered in 1853 by Liebig and has been isolated from the urine of many mammalian species. It was shown to be a metabolic product of tryptophan by Ellinger by feeding tryptophan to rats. He also proposed a structural formula for it which, however, was not quite correct. 

The correct structure was established by Homer. Kynurenic acid is formed from L-tryptophan and from indolepyruvic acid, but not from the D-amino acid.  

Xanthurenic acid was isolated from urine by Musajo in 1935 and shown to be 4,8-dihydroxyquinoline-2-carboxylic acid. Because of its 8-hydroxyquinoline structure it forms colored chelates with metal ions and, in particular, gives an intense green color with ferric salts. Xanthurenic acid is only excreted in pyridoxine deficiency, as was discovered by Lepkovsky and co-workers. It is found only in pyridoxine-defi-cient animals fed L-tryptophan or kynurenine. Addition of pyridoxine or the elimination of tryptophan from the diet causes it to disappear from the urine.  

Xanthurenic acid fed to normal animals does not accumulate in the body and is not lost in the urine. This shows that a mechanism for the degradation of this compoimd must exist in the body, which is a further 

Musajo, L., Atti reals accad. nad. Lincei 81, 368 (1935), Gaez. chim. ital. 67, 165, 

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Tryptophan catabolism is also associated with several dead-end pathways, for example the formation of kynurenic and xanthurenic acids. Normal urine contains small amounts of hydroxykynurenine, kynurenine, kynurenic acid, and xanthurenic add. When large amounts of tryptophan are fed to animals, the excretion of these compounds increases. Xanthurenic acid is excreted in massive quantities in vitamin B6 deficiency. 

Kynureninase (Figure 11.16) is a pyridoxal phosphate-dependent enzyme, and its activity falls markedly in vitamin deficiency, at least partly because it undergoes a slow mechanism-dependent inactivation that leaves catalytically inactive pyridoxamine phosphate at the active site of the enzyme. The enzyme can only be reactivated if there is an adequate supply of pyridoxal phosphate. This means that in vitamin deficiency there is a considerable accumulation of both hydroxykynurenine and kynurenine, sufficient to permit greater metabolic flux than usual through kynurenine transaminase, resulting in increased formation of kynurenic and xanthurenic acids. 

The oxidative pathway of tryptophan metabolism is shown in Figure 3. Kynureninase is a pyridoxal phosphate-dependent enzyme, and in deficiency its activity is lower than that of tryptophan dioxygenase, so that there is an accumulation of hydroxy-kynurenine and kynurenine, resulting in greater metabolic flux through kynurenine transaminase and increased formation of kynurenic and xanthurenic acids. Kynureninase is exquisitely sensitive to vitamin Bg deficiency because it undergoes a slow inactivation as a result of catalysing the half-reaction of transamination instead of its normal reaction. The resultant enzyme with pyridoxamine phosphate at the catalytic site is catalytically inactive and can only be reactivated if there is an adequate concentration of pyridoxal phosphate to displace the pyridoxamine phosphate. 

The ability to metabolise a test dose of tryptophan has been widely adopted as a convenient and sensitive index of vitamin Bg nutritional status. However, induction of tryptophan dioxygenase by glucocorticoid hormones will result in a greater rate of formation of kynurenine and hydroxykynurenine than the capacity of kynureninase, and will thus lead to increased formation of kynurenic and xanthurenic acids—an effect similar to that seen in vitamin Bg deficiency. Such results may be erroneously interpreted as indicating vitamin Bg deficiency in a variety of subjects whose problem is increased glucocorticoid secretion as a result of stress or illness, not vitamin Bg deficiency. 

Inhibition of kynureninase (e.g., by estrogen metabolites) also results in accumulation of kynurenine and hydroxykynurenine, and hence increased formation of kynurenic and xanthurenic acids, again giving results which falsely suggest vitamin Bg deficiency. This has been widely, but incorrectly, interpreted as estrogen-induced vitamin Bg deficiency it is in fact simple competitive inhibition of the enzyme that is the basis of the tryptophan load test by estrogen metabolites. 

The increased excretion of kynurenic and xanthurenic acids observed in pyridoxine deficiency is probably due to the preferential combination of the pyridoxal phosphate coenzyme with the transaminase. By preventing the loss of the side chain, as a result of a decreased activity of kynureninase in pyridoxine deficiency, cyclization is favored leading to increased formation of the two acids above. 

There is abundant evidence that kynurenic and xanthurenic acids are not intermediates in the formation of nicotinic acid. In experiments with tryptophan labeled with C in the /3-position of the side chain it was found that the alanyl side chain of tryptophan is lost during its conversion to nicotinic acid. The same result was obtained with tryptophan labeled with C in the carboxyl group. 

Hydroxyanthranilic Acid. Since kynurenic and xanthurenic acids were eliminated as being precursors of nicotinic acid because tracer experiments showed that the alanyl side chain was lost prior to the formation of the vitamin, it appeared plausible to assume that 3-hydroxyanthranilic acid was an intermediate immediately following 3-hydroxykynurenine. In support of this, it has been found that it can replace niacin in the diet of the rat, but its activity is of the order of tryptophan rather than that of niacin. Furthermore, feeding 3-hydroxyanthranilic acid yields an increased urinary excretion of N -methyl nicotinamide. ... 

Kynurminase, Kynurenine Transaminase, and the Formation of Anthranilic, Kynurenic, Hydroxyanthranilic, and Xanthurenic Acids... 

Kynurenic acid and xanthurenic acid, side products of the reaction, are the products of the transamination of the a-amino group of kynurenine and 3-hydroxy-kynurenine to a-ketoglutaric acid in the presence of pyridoxal phosphate and an enzyme found in mammalian liver and kidney, kynurenine transaminase. The keto acid resulting from the transamination reaction condenses spontaneously. Liver homogenate also decarboxylates 3-hydroxykynurenine to yield 4,8-de-hydroxyquinoline. Kynurenase may catalyze the cleavage of the side chain of kynurenine or 8-hydroxy-kynurenine and lead to the formation of alanine and... 

As discussed in Section 8.3.3, estrogen metabolites inhibit kynureninase and reduce the activity of kynurenine hydroxylase to such an extent that, even without induction of tryptophan dioxygenase (Section 9.5.4.1), the activity of these enzymes is lower than is needed for the rate of flux through the pathway, thus leading to increased formation of xanthurenic and kynurenic acids. 

The increased plasma kynuremne pool and the induced xanthurenic acid urinary excretion have several implications in the assessment of diazinon noncholinergic toxicity. An increase in xanthurenic acid formation may alter glucose metabolism. Xanthurenic acid has been reported to form a complex with insulin and damage pancreatic P cells. Elevated plasma kynurenin may alter kynurenin transport into the brain. Since more than 40% of brain kynurenin originates from the systemic circulation, cerebral biosynthesis of neuroactive kynurenin metabolites such as quinolinic acid and kynurenic acid may change. Finally, the availability of L-iryptophan for other L-lryptophan-dependent processes may be reduced. Tryptophan is the metabolic precursor for. serotonin and nicotinic adenine dinucleotidc. Diabetes, bladder cancer, and neurological disorders may be the toxic consequences of diazinon-altered L-tryptophan metaboli.sm (Seifert and Pewnim, 1992 Pewnim and Seifert, 1993). 

Fic. 1. Metabolism of tryptophan to serotonin (5-hydroxytryptamine) and niacin. Fyiidoxal phosphate (PLP) dependent reactions are indicated. Reactions not shown which may result in formation of products excreted in urine include the acetylation of liymuenine and 3-hydroxykynurenine, conjugation of anthranilic acid with glycine (to form o-aminohippuric acid) and with glucuronic acid, and the dehydroxylation of kynurenic acid and xanthurenic add to quinaldic add and 8-hydroxyquinaldic add, respectively. 

Xanthurenic and kynurenic acids, and kynurenine and hydroxykynurenine, are easy to measure in urine, so the tryptophan load test (the ability to metabolize a test dose of 2—5 g of tryptophan) has been widely adopted as a convenient and very sensitive index of vitamin nutritional status. However, because glucocorticoid hormones increase tryptophan dioxygenase activity, abnormal results of the tryptophan load test must be regarded with caution, and cannot necessarily be interpreted as indicating vitamin B deficiency. Increased entry of tryptophan into the pathway will overwhelm the capacity of kynureninase, leading to increased formation of xanthurenic and kynurenic acids. Similarly, oestrogen metabolites inhibit kynureninase, leading to results that have been misinterpreted as vitamin B deficiency. 

Scheme 5 Biogenetic pathway for the formation of anthranilic (1), kynurenic (D, and xanthurenic [10(8)] acids, and quinolabactin [10(8) ] (176).

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