New insights and advances on pyomelanin production: from microbial synthesis to applications

Abstract

Pyomelanin is a brown-black phenolic polymer and results from the oxidation of homogentisic acid (HGA) in the L-tyrosine pathway. As part of the research for natural and active ingredients issued from realistic bioprocesses, this work re-evaluates the HGA pigment and makes an updated inventory of its syntheses, microbial pathways, and properties, with tracks and recent advances for its large-scale production. The mechanism of the HGA polymerization is also well documented. In alkaptonuria, pyomelanin formation leads to connective tissue damage and arthritis, most probably due to the ROS issued from HGA oxidation. While UV radiation on human melanin may generate degradation products, pyomelanin is not photodegradable, is hyperthermostable, and has other properties better than L-Dopa melanin. This review aims to raise awareness about the potential of this pigment for various applications, not only for skin coloring and protection but also for other cells, materials, and as a promising (semi)conductor for bioelectronics and energy.

Keywords: Applications; Hydroxylase; Laccases; Polymerization; Pyomelanin.

Graphical Abstract

Fig. 1

Biosynthesis of pyomelanin through the homogentisate pathways is redefined as the HGA catabolon. This catabolon is involved in the degradation of L-Phe, L-Tyr, PA, and two hydroxy-PA derivatives into the common HGA and further toward the same route of catabolic convergence until fumarate and acetoacetate. Compounds: CHMS, 5-carboxymethyl-2-hydroxymuconic semialdehyde acid; 3,4-DHPA, 3,4-dihydroxyphenylacetic acid; FAA, fumarylacetoacetate; FAH, FAA hydrolase; GSH, glutathione; HGA, homogentisic acid; HGO, homogentisic acid 1,2-dioxygenase; x-HPA, x-hydroxyphenylacetic acid (x = 2, 3, or 4); 3,4-HPADO, 3,4-dihydroxyphenylacetate dioxygenase; 4-HPAH-y, 4-hydroxyphenylacetate hydroxylase-(y = 1, 2, 3, 5, or 6); 4-HPP, 4-hydroxyphenylpyruvic acid; 4-HPPD, 4-hydroxyphenylpyruvate dioxygenase; 4-HPPO, 4-hydroxyphenylpyruvate oxidase; MAA, 4-maleylacetoacetate; MAAI, MAA isomerase; PA, phenylacetic acid; L-PAH, L-phenylalanine hydroxylase; PAH-z, phenylacetic acid hydroxylase-(z = 2 or 5); L-Phe, L-phenylalanine; TAT, tyrosine transaminase; and L-Tyr, L-tyrosine.

Fig. 2

Inhibitors of pyomelanin synthesis and other inhibitors to differentiate pyomelanin from other melanin. (A) Inhibitors of pyomelanin synthesis are represented in red, compared to those of L-Dopa (blue) and DHN-melanin (green). Reactions are detailed in Fig. 1. (B) Chemical structure of the main inhibitors. Abbreviations: L-Cys, L-cysteine; DHBTPI, 1-(4,9-dihydrobenzo[e]thieno[2,3-b]thiepin-4-yl)-1H-imidazole; 4-HR, 4-hexylresorcinol; kojic acid, 5-hydroxy-2-(hydroxymethyl)-4H-pyran-4-one; MA, mandelic acid; medicarpin, 3-hydroxy-9-methoxypterocarpan; mesotrione, 2-[4-(methylsulfonyl)-2-nitrobenzoyl]cyclohexane-1,3-dione; NaN₃, sodium azide; NTBC (nitisinone), 2-[2-nitro-4-(trifluoromethyl)benzoyl]cyclohexane-1,3-dione; PA, phenylacetic acid; PP, phenylpropionic acid; sulcotrione, 2-[4-(methylsulfonyl)-2-chlorobenzoyl]cyclohexane-1,3-dione; and tricyclazole, 5-methyl-1,2,4-triazolo(3,4-b)benzothiazole.

Fig. 3

Schematic diagram of the production of pyomelanin by the laccase process (in blue) (reproduced with permission of Lorquin et al., 2021). Comparison to the abiotic autoxidation process (in red) via steps 2’ and 3’. The laccase oxidation generates a decarboxylation mechanism giving gentisyl alcohol and gentisaldehyde identified in the polymer. In step 1, 2,5-DMPA is demethylated by HBr at reflux and gives HGA-lactone at 99% yield. In step 2, the lactone is opened and the laccase is added at its optimal activity pH. In the case of the rMt laccase, the reaction medium is previously buffered at pH 6.8 (optimal activity). Step 4 is the final HCl precipitation, followed by washing and drying. Compounds: BQA, 1,4-benzoquinone acetic acid; gentisaldehyde, 2,5-dihydroxybenzaldehyde; gentisyl alcohol, 2,5-dihydroxybenzyl alcohol; 2,5-DMPA, 2,5-dimethoxyphenylacetic acid; and HGA, homogentisic acid.

Fig. 4

The proposed mechanism for the radical polymerization of HGA, showing the most probable structures (A, in red) and the detailed representations of the corresponding dimers in (B). R = -CH₂-COOH. Gentisyl alcohol (major) and gentisaldehyde (minor) issued from the decarboxylation mechanism (laccase process, bacteria cultures) are not represented here but are incorporated into the polymer in the same manner as HGA radicals at locations of the chain that could not be determined at this time.

Fig. 4

The proposed mechanism for the radical polymerization of HGA, showing the most probable structures (A, in red) and the detailed representations of the corresponding dimers in (B). R = -CH₂-COOH. Gentisyl alcohol (major) and gentisaldehyde (minor) issued from the decarboxylation mechanism (laccase process, bacteria cultures) are not represented here but are incorporated into the polymer in the same manner as HGA radicals at locations of the chain that could not be determined at this time.