laccase and syringaldehyde

As a new type of environmental pollutant, environmental antibiotic residues have attracted widespread attention, and the degradation and removal of antibiotics has become an engaging topic for scholars. In this paper, Novozym 51003 industrialized laccase and syringaldehyde were combined to degrade sulfonamides in aquaculture wastewater. Design Expert10 software was used for multiple regression analysis, and a response surface regression model was established to obtain the optimal degradation parameters. In the actual application, the degradation system could maintain a stable performance within 9 h, and timely supplement of the mediator could achieve a better continuous degradation effect. Low concentrations of heavy metals and organic matter would not significantly affect the degradation performance of the laccase-mediator system, making the degradation system suitable for a wide range of water quality. Enzymatic reaction kinetics demonstrated a strong affinity of sulfadiazine to the substrate. Ten degradation products were speculated using high-resolution mass spectrum based on the mass/charge ratios and the publication results. Four types of possible degradation pathways of sulfadiazine were deduced. This work provides a practical method for the degradation and removal of sulfonamide antibiotics in actual sewage.

Topics: Anti-Bacterial Agents; Aquaculture; Benzaldehydes; Kinetics; Laccase; Sulfadiazine; Sulfanilamide; Sulfonamides; Wastewater

Metabolite formation from radical-based oxidation of the environmental pollutant triclosan (TCS) was compared using an ascomycete (Phoma sp. UHH 5-1-03) and a basidiomycete (Trametes versicolor) laccase, laccase-redox mediator systems, and electrochemical oxidation (EC). Laccase oxidation predominantly yielded TCS di- and trimers, but notably also caused TCS ether bond cleavage. The latter was more prominent during EC-catalysed TCS oxidation, which generally resulted in a broader and more divergent product spectrum. By contrast, only quantitative but not qualitative differences in TCS metabolite formation were observed for the two laccases. Application of the presumable natural laccase redox mediator syringaldehyde (SYD) shifted the TCS-transforming reactions of laccase systems from oligomerization more towards ether bond cleavage. However, the observed rapid removal of SYD from reaction systems caused by predominant adduct formation from SYD and TCS, and concomitant conversion of SYD into 2,6-dimethoxy-1,4-benzoquinone (DMBQ) clearly demonstrates that SYD does not function as a "true" laccase redox mediator in the sense of being recycled during TCS oxidation. Laccase treatment of TCS without SYD decreased the anti-bacterial TCS activity more than treatment employing SYD in addition, indicating that SYD and/or its transformation products contribute to bacterial toxicity. DMBQ was found to be about 80% more active in a bacterial growth inhibition test than its parent compound SYD in terms of IC

Topics: Ascomycota; Benzaldehydes; Benzoquinones; Biotransformation; Disinfectants; Electrochemical Techniques; Environmental Pollutants; Escherichia coli K12; Laccase; Oxidation-Reduction; Trametes; Triclosan; Water Purification

Redox-mediators such as syringaldehyde (SA) can improve laccase-catalyzed degradation of trace organic contaminants (TrOCs) but may increase effluent toxicity. The degradation performance of 14 phenolic and 17 non-phenolic TrOCs by a continuous flow enzymatic membrane reactor (EMR) at different TrOC and SA loadings was assessed. A specific emphasis was placed on the investigation of the toxicity of the enzyme (laccase), SA, TrOCs and the treated effluent. Batch tests demonstrated significant individual and interactive toxicity of the laccase and SA preparations. Reduced removal of resistant TrOCs by the EMR was observed for dosages over 50μg/L. SA addition at a concentration of 10μM significantly improved TrOC removal, but no removal improvement was observed at the elevated SA concentrations of 50 and 100μM. The treated effluent showed significant toxicity at SA concentrations beyond 10μM, providing further evidence that higher dosage of SA must be avoided.

Topics: Aspergillus oryzae; Benzaldehydes; Biodegradation, Environmental; Bioreactors; Catalysis; Clostridium; Dose-Response Relationship, Drug; Inhibitory Concentration 50; Kinetics; Laccase; Luminescence; Membranes, Artificial; Organic Chemicals; Oxidation-Reduction; Photobacterium

Laccases (LCs) are multicopper oxidases that find application as versatile biocatalysts for the green bioremediation of environmental pollutants and xenobiotics. In this study we elucidate the degrading activity of Lac2 pure enzyme form Pleurotus pulmonarius towards aflatoxin B₁ (AFB₁) and M₁ (AFM₁). LC enzyme was purified using three chromatographic steps and identified as Lac2 through zymogram and LC-MS/MS. The degradation assays were performed in vitro at 25 °C for 72 h in buffer solution. AFB₁ degradation by Lac2 direct oxidation was 23%. Toxin degradation was also investigated in the presence of three redox mediators, (2,2'-azino-bis-[3-ethylbenzothiazoline-6-sulfonic acid]) (ABTS) and two naturally-occurring phenols, acetosyringone (AS) and syringaldehyde (SA). The direct effect of the enzyme and the mediated action of Lac2 with redox mediators univocally proved the correlation between Lac2 activity and aflatoxins degradation. The degradation of AFB₁ was enhanced by the addition of all mediators at 10 mM, with AS being the most effective (90% of degradation). AFM₁ was completely degraded by Lac2 with all mediators at 10 mM. The novelty of this study relies on the identification of a pure enzyme as capable of degrading AFB₁ and, for the first time, AFM₁, and on the evidence that the mechanism of an effective degradation occurs via the mediation of natural phenolic compounds. These results opened new perspective for Lac2 application in the food and feed supply chains as a biotransforming agent of AFB₁ and AFM₁.

Topics: Acetophenones; Aflatoxin B1; Aflatoxin M1; Benzaldehydes; Benzothiazoles; Biodegradation, Environmental; Food Microbiology; Fungal Proteins; Laccase; Oxidation-Reduction; Pleurotus; Proteolysis; Substrate Specificity; Sulfonic Acids; Time Factors

A Structure Activity Relationship (SAR) study for laccase mediator systems was performed in order to correctly classify different natural phenolic mediators. Decision tree (DT) classification models with a set of five quantum-chemical calculated molecular descriptors were used. These descriptors included redox potential (ɛ°), ionization energy (E(i)), pK(a), enthalpy of formation of radical (Δ(f)H), and OH bond dissociation energy (D(O-H)). The rationale for selecting these descriptors is derived from the laccase-mediator mechanism. To validate the DT predictions, the kinetic constants of different compounds as laccase substrates, their ability for pesticide transformation as laccase-mediators, and radical stability were experimentally determined using Coriolopsis gallica laccase and the pesticide dichlorophen. The prediction capability of the DT model based on three proposed descriptors showed a complete agreement with the obtained experimental results.

Topics: Acetophenones; Benzaldehydes; Biocatalysis; Catechols; Coumaric Acids; Decision Trees; Dichlorophen; Fungal Proteins; Hydrazones; Laccase; Models, Chemical; Models, Molecular; Molecular Structure; Nitrophenols; Oxidation-Reduction; Phenols; Polyporales; Protein Conformation; Quantitative Structure-Activity Relationship; Vanillic Acid

The ability of two natural phenols to act as mediators of the recombinant Myceliophthora thermophila laccase (MtL) in eucalypt-pulp delignification was investigated. After alkaline peroxide extraction, the properties of the enzymatically-treated pulps improved with respect to the control. The pulp brightness increased (3.1 points) after the enzymatic treatment with MtL alone, but the highest improvements were obtained after the MtL treatment using syringaldehyde (4.7 points) and especially methyl syringate (8.3 points) as mediators. Likewise, a decrease in kappa number up to 2.7 points was obtained after the MtL-methyl syringate treatment, followed by decreases of 1.4 and 0.9 points after the treatments with MtL-syringaldehyde and MtL alone, respectively. On the other hand, removal of the main lipophilic extractives present in eucalypt pulp was observed after the above laccase-mediator treatments. Finally, the doses of both MtL and methyl syringate were reduced, and results compatible with industrial implementation were obtained.

Topics: Benzaldehydes; Bioreactors; Eucalyptus; Gallic Acid; Gas Chromatography-Mass Spectrometry; Laccase; Lignin; Paper; Polysorbates; Sordariales; Waste Disposal, Fluid

This paper presents the use of a membrane-integrated reactor system with recycling of laccase and mediator for azo dye decolorization. From initial screening of different laccases and mediators, Trametes versicolor laccase and syringaldehyde provided the best system for decolorization. Decolorization yields of 98, 88, 80 and 78% were obtained for Red FN-2BL, Red BWS, Remazol Blue RR and Blue 4BL, respectively. The reaction parameters were optimized and a membrane reactor was set up for dye decolorization in batch mode with reuse of the enzyme. Between 10 and 20 batches could be run with decolorization yields from 95 to 52% depending on the dye type. To study the possibility of reusing both enzyme and mediator, the reactor was run using 2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO) coupled to polyethylene glycol (PEG). Nine batches were run for the treatment of Remazol Blue RR, providing decolorization yields of 96-78%. Cost analysis of the processes showed that the costs of laccase/syringaldehyde or laccase/TEMPO were almost equal when running 20 batches, but the cost for the PEG-TEMPO was higher. However, the advantages associated with reuse of the mediator should motivate further development of the concept.

Topics: Azo Compounds; Benzaldehydes; Biodegradation, Environmental; Bioreactors; Color; Coloring Agents; Costs and Cost Analysis; Cyclic N-Oxides; Enzyme Stability; Laccase; Spectrophotometry

The great amount of pollutants released from kraft pulp processes, mainly from cooking and bleaching stages, is one of the most relevant environmental problems in this type of industry. New bleaching sequences are being studied based on the use of oxidative enzymes from fungal cultures. In this study, the bleaching systems consisting of Laccase and different mediators such as 1-hydroxybenzotriazole, violuric acid, syringaldehyde and methyl syringate in the bleaching sequence of Eucalyptus globulus kraft pulp were applied. The main objective of this study is to evaluate the aerobic and anaerobic biodegradability and toxicity to Vibrium fischeri of generated L-stage and total bleaching sequence effluents. The highest levels of aerobic and anaerobic degradation of the generated effluents were achieved for treatments with laccase plus violuric acid, with 80% of aerobic degradation and 68% of anaerobic biodegradation. V. fischeri toxicity was remarkably reduced for all the effluents after aerobic degradation.

Topics: Aerobiosis; Aliivibrio fischeri; Anaerobiosis; Barbiturates; Benzaldehydes; Biodegradation, Environmental; Biomass; Eucalyptus; Gallic Acid; Industrial Waste; Laccase; Microbial Viability; Paper; Sewage; Triazoles; Waste Disposal, Fluid

The phenols syringaldehyde (SA), acetosyringone (AS) and p-coumaric acid (PCA) were used as natural laccase mediators in combination with a laccase from Pycnoporus cinnabarinus to bleach flax fibres. Their performance was compared with 1-hydroxybenzotriazole (HBT) in terms of enzyme stability, and pulp and effluent properties. HBT and PCA were found to inactivate laccase in the absence of pulp. However, in the presence of unbleached flax pulp stability was increased; for example with PCA, laccase retained 77% of its initial activity, in contrast with complete inactivation in the absence of pulp. This suggests a protective effect of the pulp against denaturalization of the enzyme. All natural mediators resulted in a reduced kappa number after the subsequent alkaline treatment with hydrogen peroxide; the reduction being especially marked with SA (about 2 units - with respect to the control sample) and comparable to that obtained by HBT. Brightness was significantly increased by all natural mediators, but especially by AS and SA (23% with both), which performed very similarly to HBT in this respect. Natural mediators therefore might constitute an effective alternative to synthetic mediators for flax pulp biobleaching. This paper demonstrates for the first time the use of natural mediators in the laccase-assisted delignification of flax pulp and their effect on the properties of the resulting effluents.

Topics: Acetophenones; Basidiomycota; Benzaldehydes; Color; Coumaric Acids; Enzyme Stability; Flax; Laccase; Propionates; Temperature

The transformation of organic halogenated pesticides by laccase-mediator system has been investigated. Twelve pesticides were assayed in the presence of nine different mediators. Acetosyringone and syringaldehyde showed to be the best mediators. The halogenated pesticides bromoxynil, niclosamide, bromofenoxim and dichlorophen were transformed by the laccase-syringaldehyde system showing catalytic activities of 48.8, 142.0, 166.2 and 1257.6nmolmin(-1)U(-1), respectively. The highest pesticide transformation rates were obtained with a mediator-substrate proportion of 5:1, one of the lowest reported so far for the laccase-mediator systems. The analysis of the main product from the dichlorophen transformation showed that an oxidative dehalogenation is involved in the catalytic mechanism. Adduct formation between the mediator syringaldehyde and the pesticides dichlorophen or bromoxynil was also found after enzymatic oxidation. The main goal of this work is to evaluate environmental-friendly mediators for the pesticide transformation, and the potential of laccase-mediator system to efficiently reduce the environmental impact of organic halogenated pesticides is discussed.

Topics: Benzaldehydes; Biotransformation; Dichlorophen; Fungi; Halogenation; Laccase; Niclosamide; Nitriles; Oxidation-Reduction; Oximes; Pesticides; Trametes

The natural phenolic compounds syringaldehyde and vanillin were compared to the synthetic mediators 1-hydroxybenzotriazole, violuric acid and promazine in terms of boosting efficiency in a laccase-assisted biobleaching of eucalyptus kraft pulp. Violuric acid and 1-hydroxybenzotriazole revealed to be the most effective mediators of the bioprocess. Nevertheless, laccase-syringaldehyde system also improved the final pulp properties (28% delignification and 63.5% ISO brightness) compared to the process without mediator (23% and 61.5% respectively), in addition to insignificant denaturation effect over laccase. The efficiency of the biobleaching process was further related to changes in non-conventionally used optical and chromatic parameters of pulp, such as (L*), chroma (C*) and dye removal index (DRI) showing good correlation. Adverse coupling reactions of the natural phenolic mediators on pulp lignin were predicted by electrochemical studies, demonstrating the complexity of the laccase-mediator reaction on pulp.

Topics: Barbiturates; Benzaldehydes; Color; Eucalyptus; Industrial Waste; Laccase; Paper; Peroxides; Phenols; Promazine; Spectrum Analysis; Triazoles

In this paper, we show for the first time that lignin-derived phenols can act as laccase mediators for the removal of lipophilic compounds from paper pulp. These natural mediators represent an alternative to synthetic mediators, such as 1-hydroxybenzotriazole (HBT), that cause some economic and environmental concerns. Unbleached kraft pulp from eucalypt wood, which contained free and conjugated sterols responsible for pitch deposition in the manufacture of totally chlorine free paper, was treated with a fungal laccase in the presence of syringaldehyde, acetosyringone, and p-coumaric acid as mediators. The composition of lipophilic extractives in the pulps after the enzymatic treatment followed by a hydrogen peroxide stage was analyzed by gas chromatography and gas chromatography/mass spectrometry. The enzymatic treatment using syringaldehyde as laccase mediator caused the highest removal (over 90%) of free and conjugated sitosterol, similar to that attained with HBT, followed by acetosyringone (over 60% removal), whereas p-coumaric acid was barely effective. Moreover, recalcitrant oxidized steroids surviving laccase-HBT treatment could be removed when using these natural mediators. Pulp brightness was also improved (from 57% to 66% ISO brightness) by the laccase treatment in the presence of the above phenols followed by the peroxide stage due to the simultaneous removal of lignin.

Topics: Acetophenones; Benzaldehydes; Coumaric Acids; Eucalyptus; Industrial Waste; Laccase; Lignin; Paper; Phenols; Plant Extracts; Propionates; Waste Management; Wood

Chlorinated phenols and anilines were transformed by oxidoreductive catalysts with release of chloride ions in both the absence and the presence of humic substances (syringaldehyde, catechol, and humic acid). Dehalogenation of these xenobiotics resulted from oxidative coupling reactions occurring at the chlorinated sites of the substrates. The effect of humic substances on dehalogenation depended on the mechanism of oxidative coupling. In a free-radical reaction mediated by peroxidase, laccase, or birnessite (delta-MnO2), syringaldehyde enhanced the dehalogenation of most of the chlorinated phenols, but it did not enhance the dehalogenation of the chloroanilines. With catechol, which does not form free radicals, dehalogenation was reduced or remained the same for both the chlorophenols and the chloroanilines. However, in tyrosinase-mediated reactions controlled by nucleophilic addition, catechol enhanced the dehalogenation of most of the chlorophenols, whereas syringaldehyde had little effect. Humic acid in most cases enhanced the dehalogenation of the chlorophenols, but it had little effect on the dehalogenation of the chloroanilines. On a molar basis, changes in dehalogenation caused by humic substances were proportional to the respective changes in substrate transformation. Only syringaldehyde was capable of releasing disproportionately high amounts of chloride ions from chlorophenols, apparently as a result of multiple crosscouplings to one molecule of the substrate.

Topics: Benzaldehydes; Biodegradation, Environmental; Catechols; Chlorophenols; Humic Substances; Laccase; Manganese Compounds; Oxidation-Reduction; Oxides; Oxidoreductases; Peroxidase