Fungal laccase-catalyzed biotransformation of naturally occurring phenols for biotechnological applications

Abstract

Laccases are multi-copper oxidases that catalyze single-electron oxidations of phenolic compounds with the concomitant reduction of oxygen to water. Applicability of laccases has been highlighted due to green nature and versatile biochemical features of the enzymes. In particular, use of fungal laccases that are commercially available has proven to be effective for several biotechnological areas including pulp or textile bleaching, bioremediation, organic synthesis, and polymer surface modification. The green nature of the laccase applications can be markedly enhanced by the use of low-molecular-weight naturally occurring phenols, whose involvements in laccase-catalyzed in vivo metabolisms contributing carbon recycling in land ecosystem and morphogenesis of bio-matrices are also evident. Although several review papers focusing on laccase biotechnological applicability have been reported, connection between the in vivo metabolisms and the application concepts has not been attracted. Thus, we here reviewed current advances in laccase-catalyzed oxidations of naturally occurring phenols particularly focusing on laccase-mediator system (LMS) and organic synthesis, and discussed how both the application strategies can mimic in vivo biometabolism to make the application processes much greener and to extend applicability of the green reactions. Laccases have low redox potentials limiting their environmental and industrial applications. The use of laccase mediators has proven to be an effective approach for overcoming the low redox potentials. However, knowledge about the role played by the mediator cocktails in such a laccase-mediator system (LMS) is scarce. Here we assembled different dual-agent mediator cocktails containing 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulfonate (ABTS), vanillin, and/or acetovanillone, and compared their mediating capabilities with those of each individual mediator alone in oxidation of pentachlorophenol (PCP) by Ganoderma lucidum laccase. Cocktails containing ABTS and either vanillin or acetovanillone strongly promoted PCP removal compared to the use of each mediator alone. The removal enhancement was correlated with mediator molar ratios of the cocktails and incubation times. Analysis of the kinetic constants for each mediator compound showed that G. lucidum laccase was very prone to react with ABTS rather than vanillin and acetovanillone in the cocktails. Moreover, the presence of the ABTS radical (ABTS+&#8226

) and vanillin or acetovanillone significantly enhanced PCP removal concomitant with electron transfer from vanillin or acetovanillone to ABTS+&#8226

. These results strongly suggest that vanillin and acetovanillone mediate the reaction between ABTS and PCP via multiple sequential electron transfers among laccase and its mediators.Laccase efficiently catalyzes polymerization of phenolic compounds. However, knowledge on applications of polymers synthesized in this manner remains scarce. Here, the potential of laccase-catalyzed polymerization of natural phenols to form products useful in hair dyeing was investigated. All 15 tested phenols yielded colored products after laccase treatment and color diversity was attained by using mixtures of two phenolic monomers. After exploring color differentiation pattern of 120 different reactions with statistical regression analysis, three monomer combinations, namely gallic acid and syringic acid, catechin and catechol, and ferulic acid and syringic acid, giving rise to brown, black, and red materials, respectively, were further characterized because such colors are commercially important for gray hair dyeing. Selected polymers could strongly absorb visible light and their hydrodynamic sizes ranged from 100 to 400 nm. Analyses of enzyme kinetic constants, liquid chromatography, and electrospray ionization-mass spectrometry (ESI-MS) coupled with collision-induced dissociation MS/MS indicate that both monomers in reactions involving catechin and catechol, and ferulic acid and syringic acid, are colored by heteropolymer synthesis, but the gallic acid/syringic acid combination is based on homopolymer mixture formation. Comparison of color parameters from these three reactions with those of corresponding artificial homopolymer mixtures also supported the idea that laccase may catalyze either hetero- or homo-polymer synthesis. We finally used selected materials to dye gray hair. Each material colored hair appropriately and the dyeing showed excellent resistance to conventional shampooing. Our study indicates that laccase-catalyzed polymerization of natural phenols is applicable to the development of new cosmetic pigments.Fungal laccase-based eco-coloration with plant-derived phenols is highly applicable to cosmetic or food processing industries due to eco-friendly features of the dye synthesis process. However, the synthesis procedure is not based on any theoretical rationale so that considerable experimental work is required for finding desirable colors. Here, the coloration reaction using natural phenol blends was numerically predicted using a well-defined learning algorithm, the support vector machine. Both forward and backward predictions were successfully developed using 1,439 different colorations for training and testing. The forward calculations commenced with known monomer compositions and yielded color parameters

the mean absolute errors for each parameter were 1.4222 for L*, 0.7583 for a*, and 2.1442 for b*. Reversely, the backward calculations with known color parameters yielded monomer compositions

average accuracy was 71.9%. Two variables regarding color differentiation of each natural phenol were identified to critically affect the backward prediction accuracy, suggesting dependence of the accuracy on monomer compositions and color differentiation capacity of each monomer. Based on true or false (either -positive or -negative) ratio analysis, relation between the accuracy and the monomer composition was further discussed. Our coloration data pool and predictive models will aid to develop synthetic routes to give rise to desirable colors via fungal laccase-catalyzed polymerization of plant-derived phenols. The color SVM software is available at http://home.postech.ac.kr/~fenrir/colorsvm/colorsvm.tar.gz.Natural organic coagulants (NOCs) such as chitosan and Moringa oleifera seeds have been extensively characterized for potential application in water treatment as an alternative to metal-based coagulants. However, the action of both chitosan and Moringa oleifera seeds is mainly restricted to anionic organic pollutants because of their cationic functional groups affording poor cationic pollutant coagulation by electrostatic repulsion. In this study, we employed ethanolic grape seed extract (GSE) and grape seed-derived polyphenols such as tannic acid and catechin in an effort to find novel NOCs showing stable anionic forms for removal of cationic organic pollutants. The target substances tested were malachite green (MG) and crystal violet (CV), both mutagenic cationic dyes. Polyphenol treatment induced fast decolorization followed by gradual floc formation concomitant with red or blue shifts in maximum absorbance wavelengths of the cationic dyes. Liquid chromatography analysis of flocs formed by polyphenols directly showed that initial supramolecular complexes attributed mainly to electrostatic attraction between polyphenol hydroxyphenyl groups and cationic dyes further progressed into stronger aggregates, leading to precipitation of dye-polyphenol complexes. Consistent with the results obtained using catechin and tannic acid, use of GSE also resulted in effective decolorization and coagulation of soluble MG and CV in aqueous solutions. Screening of several organic GSE components for NOC activity strongly suggested that natural polyphenols are the main organic ingredients causing MG and CV removal via gradual floc formation. The treatment by natural polyphenols and GSE decreased toxicity of MG- or CV-contaminated water.