5.6- Dihydroxyindole 2-carboxylic acid

Eumelanins — These melanins are considered polymers derived from tyrosine derivatives, mainly 5,6-dihydroxyindole-2-carboxylic acid (DHCIA) and dihidrox-yindole (DHl), with high degrees of cross-linking. In vivo eumelanins are associated with proteins and with metals, most frequently copper, zinc, or iron. 

Melanin biosynthesis in animals is a complex process starting with the L-tyrosine amino acid. In the first step, L-tyrosine is converted first into DOPA and then into dopaquinone, a process catalyzed by tyrosinase. In the biosynthesis of eumelanins, dopaquinone undergoes a cyclization to form dopachrome and subsequently a tau-tomerization into 5,6-dihydroxyindole-2-carboxylic acid (DHICA). DHICA is further oxidized to indole-5,6-quinone2-carboxylic acid, the precnrsor of DHICA eumelanins. Tyrosinase-related proteins TRP-2 and TRP-1, respectively, are responsible for the last two steps, and they are under the control of the tyrosinase promoter. 

Pezzella, A. et al., An integrated approach to the structure of sepia melanin evidence for a high proportion of degraded 5,6-dihydroxyindole-2-carboxylic acid units in the pigment backbone, Tetrahedron, 53, 8281, 1997. 

LOX-hydrogen peroxide system catalyzed the conversion of 5,6-dihydroxyindole and 5,6-dihydroxyindole-2-carboxylic acid, which are important intermediates of melanogenesis, into melanin pigments [47]. 

In 1927 Raper showed that the red pigment obtained on oxidation of DOPA [i.e. 2,3-dihydroindole-5,6-quinone-2-carboxylic acid, dopachrome (4)] rearranged spontaneously by an autoreduction process in vacuo to 5,6-dihydroxyindole (29).72 The rearrangement process could be accelerated by the action of alkali or sulfur dioxide.72 In the latter case, decarboxylation did not accompany the rearrangement and the colorless derivative was 5,6-dihydroxyindole-2-carboxylic acid (17).72 Compounds 17 and 29 were isolated as their dimethyl ethers, (30A) and (30B).72 Immediate decolorization of epinochrome (27) solutions on addition of alkali was reported a few years later.134... 

Dihydroxyindole and 5,6-dihydroxyindole-2-carboxylic acid were shown to form after the red pigment stage that occurred during the conversion of DOPA into a melanin, by Raper, who isolated these compounds as their dimethyl ethers.72 The presence of 5,6-dihydroxyindoles in solutions of DOPA, dopamine, and noradrenaline which are undergoing oxidation has subsequently been confirmed by paper and thin-layer chromatography.118,120,222 5,6-Dihydroxyin-dole and 5,6-dihydroxyindole-2-carboxylic acid have recently been isolated from the alkali fusion products of sepiomelanin, indicating... 

In the assay described by Smit et al. (1990), 5,6-dihydroxyindole-2-carboxylic acid was the substrate. Two products are formed, 5-hydroxy-6-methoxyindole-2-carboxylic add and 6-hydroxy-5-methoxyindole-2-carbox-ylic add. 

The enzyme assay contained 0.1 mM 5,6-dihydroxyindole-2-carboxylic acid in 30 mM Tris-HCl buffer (pH 7.8), 2.5 mM MgCl2,1 mM 5-adenosylmethio-nine (as methyl donor), 0.5% Triton X-100,0.5 mM EGTA, and 5 mAf dithio-threitol in a total volume of 250 / L. The reaction was stopped with 25 fiL of 4 M perchloric add. Following centrifugation, the supemate was analyzed by HPLC. Product formation was linear with time for 60 minutes. 

Ty initiates melanin synthesis by the hydroxylation of L-tyrosine to 3,4-dihydroxyphenylalanine (Dopa) and the oxidation of dopa to dopaquinone. In the presence of L-cysteine, dopaquinone rapidly combines with the thiol group to form cysteinyldopas, which undergo nonen-zymatic conversion and polymerization to pheomelanin via benzothiazine intermediates. In the absence of thiol groups, dopaquinone very rapidly undergoes conversion to dopachrome, which is transformed to 5,6-dihydroxyindole-2-carboxylic acid (DHICA) by dopachrome tautomerase. Alternatively, dopachrome is converted nonenzymatically to 5,6-dihydroxyindole (DHI). Oxidation of DHICA and DHI to the corresponding quinones and subsequent polymerization leads to eumelanins. It is still questionable if Ty is involved in this step. 

NaBH4/NaOH treatment yields 5,6-dihydroxyindole-2-carboxylic acid... 

Olivares C et al. (2001) The 5,6-dihydroxyindole-2-carboxylic acid (DHICA) oxidase activity of human tyrosinase. Biochem J 354(Ptl) 131-139... 

Multistep Syntheses Based on Conventional Procedures 5,6-Dihydroxyindole-2-Carboxylic Acid 2 and Related Compounds... 

The first approach to 5,6-dihydroxyindole-2-carboxylic acid 2 and related derivatives was due to Beer and co-workers (49JCS2061) and was based on the reductive cyclization of 2-nitro-4,5-dihydroxyphenylpyruvic acid or a derivative. The critical reduction step is achieved with Fe powder in ethanol/acetic acid. Notably, indole 2 was reported to oxidize poorly and only slowly, giving in the course of 24 h a dark-brown solution but not an insoluble product of the melanin type. This observation led to the conclusion that ... it is unlikely that 5,6-dihydroxyindole-2-carboxylic acid is a precursor of melanin . This conclusion was disproved by subsequent studies (97T8281). In the same paper, the preparations and properties of... 

Napolitano A, Crescenzi O, Prota G (1993) Copolymerization of 5,6-Dihydroxy-indole and 5,6-Dihydroxyindole-2-carboxylic Acid in Melanogenesis. Isolation of a Cross-Coupling Product. Tetrahedron Lett 34 885... 

Eumelanins (black or brown) that are produced in the course of oxidation of tyrosine (and/or phenylalanine) to 3,4-dihydroxyphenylalanine (DOPA) and dopaquinone, which further undergoes cyclization to 5,6-dihydroxyindole or 5,6-dihydroxyindole- 2-carboxylic acid... 

The reaction of peroxynitrite with the melanin precursor 5,6-dihydroxyindole-2-carboxylic acid (DHICA) produces polymeric products. The reaction proceeds through unstable intermediates including dimers and trimers of DHICA in addition to indole-5,6-quinone-2-carboxylic acid thus peroxynitrite, which nitrates phenolic compounds. 

Scheme 1. Biosynthetic pathway of melanin (Chang, 2009 Kim, 2005 Seo, 2003). DOPA, 3,4-dihydroxyphenylalanine DHI, 5,6-dihydroxyindole DHICA, 5,6-dihydroxyindole-2-carboxylic acid.

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