Laccase In Combination Research Articles

Ultrafast synthesis of laccase-copper phosphate hybrid nanoflowers for efficient degradation of tetracycline antibiotics

The presence of antibiotics in the environment causes increasing attention due to their potential risks to ecosystems and public health. Laccases are versatile oxidases capable of degrading various organic contaminants including pharmaceuticals. However, the performance of bacterial laccases on tetracycline antibiotics (TCs) degradation is seldom investigated. In this work, a bacterial laccase from Bacillus amyloliquefaciens was immobilized as laccase-inorganic hybrid nanoflowers (Lac-hNFs) by a facile and rapid method. The immobilized laccase was employed to remove different TCs including tigecycline, which is a third-generation TC that its degradation by laccase has not been reported. Lac-hNFs were synthesized by sonication-mediated self-assembly of laccase and copper ions in 5 min at room temperature. About 95% of laccase could be encapsulated in the nanoflowers, and the obtained Lac-hNFs exhibited great enhancement in stability under harsh conditions. The immobilized laccase showed a half-life of 11.7 h at 60 °C, which was about 1.4-fold higher than that of the free enzyme. Meanwhile, Lac-hNFs retained 81% of the initial activity after incubation at 25 °C for 10 days. The laccase in combination with acetosyringone could efficiently decompose tetracycline, doxycycline, and tigecycline. More than 79% of the three TCs were transformed in 1 h. Compared with the free enzyme, Lac-hNFs demonstrated higher capacity in the removal of TCs. Furthermore, Lac-hNFs remained their high degradation capacity after five cycles of reuse. Bacterial growth inhibition test revealed that most of the toxicity of TCs was eliminated after Lac-hNFs treatment. The main transformation products were identified by LC-MS, and the possible degradation pathways were proposed. The interaction mechanism between laccase and TCs was also analyzed using molecular docking. This work provides an efficient way to remove toxic organic pollutants.

A review of a novel enzyme system for the management of thatch and soil water repellency in turfgrass

AbstractManaging soil organic matter is a crucial component of greenkeeping and turfgrass maintenance. Organic matter accumulation as thatch is a major problem in modern turfgrass management. Additionally, soil water repellency (SWR) caused by hydrophobic organic coatings on sand particles can create serious soil water infiltration and potential runoff problems and contribute to a reduction in turf quality and playability. Lignin is a major component of thatch and acts as protective matrix limiting microbial degradation of cellulose and hemi‐celluloses. This article reviews lignin structure and its unique oxidation mechanism that makes it suitable for use in turfgrass systems and the research progress to develop laccase as a tool to manage thatch layer and SWR. A series of experiments explored the potential for direct application of fungal produced enzymes to degrade lignin, speed thatch degradation, and thereby reduce thatch accumulation. Greenhouse studies provided proof of concept and confirmed laccase impact on physical and chemical properties of thatch. Field studies verified the effectiveness of laccase application on different turfgrass species and optimized application rates and frequency. The reduction in thatch layer thickness by laccase treatments was similar to reductions achieved by cultural management and greater reductions occurred when applied in combination with cultural management. The use of laccase and other enzymes to ameliorate SWR was verified by a series of laboratory and field studies. Single applications of laccase were effective in reducing SWR but not as effective as wetting agents. Enzyme applications combined with a wetting agent to provide maximum relief of SWR. Application of laccase in combination with core‐aeration and sand topdressing could reduce the number of mechanical cultivations necessary to keep thatch layers at desired levels and reduce the potential for SWR. Laccase could provide a nondisruptive organic approach to manage thatch and SWR on highly managed turfgrass.

Stable ABTS Immobilized in the MIL-100(Fe) Metal-Organic Framework as an Efficient Mediator for Laccase-Catalyzed Decolorization.

The successful encapsulation of 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), a well-known laccase mediator, within a mesoporous metal-organic framework sample (i.e., MIL-100(Fe)) was achieved using a one-pot hydrothermal synthetic method. The as-prepared ABTS@MIL-100(Fe) was characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD), Fourier transform infrared (FT-IR) spectroscopy, nitrogen sorption, and cyclic voltammetry (CV). Our ABTS@MIL-100(Fe)-based electrode exhibited an excellent electrochemical response, indicating that MIL-100(Fe) provides an appropriate microenvironment for the immobilization and electroactivity of ABTS molecules. ABTS@MIL-100(Fe) was then evaluated as an immobilized laccase mediator for dye removal using indigo carmine (IC) as a model dye. Through the application of laccase in combination with a free (ABTS) or immobilized (ABTS@MIL-100(Fe)) mediator, decolorization yields of 95% and 94%, respectively, were obtained for IC after 50 min. In addition, following seven reuse cycles of ABTS@MIL-100(Fe) for dye treatment, a decolorization yield of 74% was obtained. Dye decolorization occurred through the breakdown of the chromophoric group by the Laccase/ABTS@MIL-100(Fe) system, and a catalytic mechanism was proposed. We therefore expect that the stability, reusability, and validity of ABTS@MIL-100(Fe) as a laccase mediator potentially render it a promising tool for dye removal, in addition to reducing the high running costs and potential toxicity associated with synthetic mediators.

Design of Laccase-Metal Organic Framework-Based Bioelectrodes for Biocatalytic Oxygen Reduction Reaction.

Laccase in combination with 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) as a mediator is a well-known bioelectrocatalyst for the 4-electron oxygen reduction reactions (ORR). The present work deals with the first exploitation of mesoporous iron(III) trimesate-based metal organic frameworks (MOF) MIL-100(Fe) (MIL stands for materials from Institut Lavoisier) as a new and efficient immobilization matrix of laccase for the building up of biocathodes for ORR. First, the immobilization of ABTS in the pores of the MOF was studied by combining micro-Raman spectroscopy, X-ray powder diffraction (XRPD), and N2 porosimetry. The ABTS-MIL-100(Fe)-based modified electrode presents excellent properties in terms of charge transfer kinetics and ionic conductivity as well as a very stable and reproducible electrochemical response, showing that MIL-100(Fe) provides a suitable and stabilizing microenvironment for electroactive ABTS molecules. In a second step, laccase was further immobilized on the MIL-100(Fe)-ABTS matrix. The Lac-ABTS-MIL-100(Fe)-CIE bioelectrode presents a high electrocatalytic current density of oxygen reduction and a reproducible electrochemical response characterized by a high stability over a long period of time (3 weeks). These results constitute a significant advance in the field of laccase-based bioelectrocatalysts for ORR. According to our work, it appears that the high catalytic efficiency of Lac-ABTS-MIL-100(Fe) for ORR may result from a synergy of chemical and catalytic properties of MIL-100(Fe) and laccase.

ChemInform Abstract: Ultrasound Enhanced Laccase Applications

AbstractReview: 142 refs.

A Sequential Combination of Laccase Pretreatment and Enzymatic Hydrolysis for Glucose Production from Furfural Residues

Furfural residues (FRs) were pretreated with laccase or a laccase-mediator (1-hydroxybenzotriazole, HBT) system to produce fermentable sugar for bioethanol production. Compared to laccase-only pretreatment, laccase-mediator pretreatment dissolved more lignin. Approximately 10.5% of the initially present lignin was removed when FRs were treated with a laccase loading of 100 U/g of dry substrate in 1% (w/w) HBT at 48 °C for 24 h in an acetate buffer (pH 4.8). The enzymatic saccharification process was carried out by a combined laccase or laccase-mediator pretreatment without washing of the treated solids. The results showed that active laccase had a negative effect on the rate and yield of enzymatic hydrolysis. Laccase-oxidized HBT seriously reduced glucose yield. However, non-oxidized HBT increased glucose yield when laccase was deactivated at 121 °C for 20 min prior to enzymatic hydrolysis. The highest glucose yield, 80.9%, was obtained from the substrate pretreated with 100 U/g of dry substrate laccase and 1% (w/w) HBT at 48 °C for 24 h in an acetate buffer (pH 4.8). Furthermore, the structures of FRs before and after laccase-mediator pretreatment were characterized by scanning electron microscopy (SEM) and Fourier Transform Infrared spectroscopy (FT-IR).

An enzyme-catalysed bleaching treatment to meet dissolving pulp characteristics for cellulose derivatives applications

Bleached cellulose with good end-properties (≈ 90% ISO brightness and 62% cellulose preservation) was obtained by using a totally chlorine-free biobleaching process (TCF). Unbleached sulphite cellulose was treated with Trametes villosa laccase in combination with violuric acid. This enzymatic stage (L) was followed by a chelating stage (Q) and then by a hydrogen peroxide stage reinforced with pressurized oxygen (Po), resulting to an overall LQPo sequence. The use of violuric acid was dictated by the results of a preliminary study, where the bleaching efficiency of various natural (syringaldehyde and p-coumaric acid) and synthetic mediators (violuric acid and 1-hydroxybenzotriazole) were assessed. The outstanding results obtained with laccase-violuric acid system fulfil most of the characteristics of commercial dissolving pulp, totally acceptable for viscose manufacturing or CMC derivatives, with the added advantage that the enzymatic treatment saved 2h of reaction time and about 70% of hydrogen peroxide consumption, relative to a conventional sequence (Po).

Laccase from Pycnoporus cinnabarinus and phenolic compounds: Can the efficiency of an enzyme mediator for delignifying kenaf pulp be predicted?

In this work, kenaf pulp was delignified by using laccase in combination with various redox mediators and the efficiency of the different laccase–mediator systems assessed in terms of the changes in pulp properties after bleaching. The oxidative ability of the individual mediators used (acetosyringone, syringaldehyde, p-coumaric acid, vanillin and actovanillone) and the laccase–mediator systems was determined by monitoring the oxidation–reduction potential (ORP) during process. The results confirmed the production of phenoxy radicals of variable reactivity and stressed the significant role of lignin structure in the enzymatic process. Although changes in ORP were correlated with the oxidative ability of the mediators, pulp properties as determined after the bleaching stage were also influenced by condensation and grafting reactions. As shown here, ORP measurements provide a first estimation of the delignification efficiency of a laccase–mediator system.

A new procedure for the hydrophobization of cellulose fibre using laccase and a hydrophobic phenolic compound

A new biotechnological procedure using laccase in combination with a hydrophobic phenolic compound (lauryl gallate) for the hydrophobization of cellulose fibres and internal sizing of paper was developed. Cellulose fibres from hardwood kraft pulp were incubated with laccase (Lac), in combination with lauryl gallate (LG). The Lac-LG treatment resulted in the internal sizing of paper, and also in significantly reduced water penetration in the handsheets and wettability of the paper surface. Paper was found not to be effectively rendered hydrophobic by LG alone. SEM images of the fibre network revealed the presence of the sizing agent: a product of the reaction between laccase and lauryl gallate. Binding of lauryl gallate to cellulose fibres was suggested by the increase in kappa number of the pulp and further confirmed by IR spectroscopy.

Enhanced transformation of triclosan by laccase in the presence of redox mediators

Triclosan (TCS), an antimicrobial agent, is an emerging and persistent environmental pollutant that is often found as a contaminant in surface waters and sediments; hence, knowledge of its degradability is important. In this study we investigated laccase-mediated TCS transformation and detoxification, using laccase (from the fungus Ganoderma lucidum) in the presence and absence of redox mediators. Transformation products were identified using HPLC, ESI-MS and GC–MS, and transformation mechanisms were proposed. In the absence of redox mediator, 56.5% TCS removal was observed within 24h, concomitant with formation of new products with molecular weights greater than that of TCS. These products were dimers and trimers of TCS, as confirmed by ESI-MS analysis. Among the various mediators tested, 1-hydroxybenzotriazole (HBT) and syringaldehyde (SYD) significantly enhanced TCS transformation (∼90%). The presence of these mediators resulted in products with lower molecular weights than TCS, including 2,4-dichlorophenol (2,4-DCP; confirmed by GC–MS) and dechlorinated forms of 2,4-DCP. When SYD was used as the mediator, dechlorination resulted in 2-chlorohydroquinone (2-CHQ). Bacterial growth inhibition studies revealed that laccase-mediated transformation of TCS effectively decreased its toxicity, with ultimate conversion to less toxic or nontoxic products. Our results confirmed the involvement of two mechanisms of laccase-catalyzed TCS removal: (i) oligomerization in the absence of redox mediators, and (ii) ether bond cleavage followed by dechlorination in the presence of redox mediators. These results suggest that laccase in combination with natural redox mediator systems may be a useful strategy for the detoxification and elimination of TCS from aqueous systems.

Modification of High-Lignin Kraft Pulps with Laccase. Part 2. Xylanase-Enhanced Strength Benefits

The effects of xylanase pretreatment of high lignin content softwood (SW) kraft pulp on subsequent pulp treatment with laccase in combination with gallic acid were investigated. Although xylanase pretreatment was ineffective in enhancing the laccase-facilitated biografting of gallic acid to kraft fibers, it was beneficial for subsequent treatment with laccase exclusively. Treating pulp fibers with xylanase followed by laccase provided a collective 25% and 46% increase in dry and wet tensile strength properties, respectively.

Decolourization of recalcitrant dyes with a laccase from Streptomyces coelicolor under alkaline conditions

Colored wastewater from textile industries is a consequence of dye manufacturing processes. Two percent of dyes that are produced are discharged directly in aqueous effluent and more than 10% are subsequently lost during the textile coloration process. It is not surprising that these compounds have become a major environmental concern. In that context, we have evaluated the potential use of Streptomyces coelicolor laccase for decolourization of various dyes with and without a mediator. Results showed that in all cases the combination of laccase and the mediator acetosyringone was able to rapidly decolourize, to various degrees, all the dyes tested. In 10 min, decolourization was achieved at 94% for acid blue 74, 91% for direct sky blue 6b and 65% for reactive black 5. Furthermore, decolourization was achieved at 21% for reactive blue 19 and at 39% for the direct dye Congo red in 60 min. These results demonstrate the potential use of this laccase in combination with acetosyringone, a natural mediator, for dye decolourization.

Paper pulp delignification using laccase and natural mediators

Three plant phenols, namely acetosyringone, syringaldehyde and p-coumaric acid, were selected as laccase redox mediators to investigate the enzymatic delignification of paper pulp (obtained from kraft cooking of eucalypt wood) in combination with peroxide bleaching. The effects of these natural mediators were compared with those obtained using the synthetic mediator 1-hydroxybenzotriazole. p-Coumaric acid only caused minor increase of pulp brightness and did not lower its kappa number (a rough estimation of the lignin content), whereas, the use of acetosyringone or syringaldehyde as laccase mediators enabled over 15% increase of final brightness and a similar decrease of final kappa number. Pulp delignification by laccase in the presence of the two latter natural mediators was demonstrated by analytical pyrolysis, which does not suffer from interferences by other pulp constituents as kappa number does, showing a preferential removal of lignin marker compounds compared with carbohydrate markers (up to 25% decrease of the corresponding ratio). This technique also revealed a modification of the residual lignin composition in terms of phenylpropane units after the laccase-mediator treatment. The use of laccase in combination with natural mediators, widely available from plant materials and pulping liquors, represents a promising alternative for environmentally friendly delignification of paper pulp.

Effects of laccase, xylanase and their combination on the rheological properties of wheat doughs

The effects of Trametes hirsuta laccase alone and in combination with Aspergillus oryzae and Bacillus subtilis xylanases on dough extensibility were studied using the Kieffer test to determine the dough extensibility ( E x) and the resistance to stretching ( R max). Laccase treatment resulted in dough hardening: the R max of dough increased and the E x at R max decreased as a function of dosage (5–50 nkat/g flour). Xylanases softened flour and gluten doughs. Hardening by laccases and softening by xylanases was weaker in gluten doughs. Dough hardening, observed in the laccase treatments, decreased as a function of dough resting time. The softening effect occurred especially at higher laccase dosages (≈50 nkat/g flour). The softening phenomenon was related to the laccase-mediated depolymerization of the cross-linked AX network. In combined laccase and xylanase treatments, the effect of laccase was predominant, especially at low xylanase dosage, but when xylanase was added to flour dough at high concentrations, the hardening effect of laccase on dough was decreased. In combined laccase and xylanase treatments in gluten doughs, similar decreases in laccase-mediated hardening were not seen.

Enzymatic Biobleaching of Two Recalcitrant Paper Dyes with Horseradish and Soybean Peroxidase

A stilbene dye (Direct Yellow 11) and a methine dye, Basazol 46L, recalcitrant to common chemical bleaches, were treated with horseradish and soybean peroxidases. Both enzymes were effective at chromophore removal. When compared to laccase in combination with a mediator (ABTS), soybean peroxidase was more effective at oxidative dye removal, especially for the methine dye.