Oxidation of Various Kraft Lignins with a Bacterial Laccase Enzyme

Modification of kraft lignin (KL), traditionally uses harsh and energy-demanding physical and chemical processes. In this study, the potential of the bacterial laccase CotA (spore coating protein A) for oxidation of KL under mild conditions was assessed. Thereby, the efficiency of CotA to oxidize both softwood and hardwood KL of varying purity at alkaline conditions was examined. For the respective type of wood, the highest oxidation activity by CotA was determined for the medium ash content softwood KL (MA_S) and the medium ash content hardwood KL (MA_H), respectively. By an up to 95% decrease in fluorescence and up to 65% in phenol content coupling of the structural lignin units was indicated. These results correlated with an increase in viscosity and molecular weight, which increased nearly 2 and 20-fold for MA_H and about 1.3 and 6.0-fold for MA_S, respectively. Thus, this study confirms that the CotA laccase can oxidize a variety of KL at alkaline conditions, while the origin and purity of KL were found to have a major impact on the efficiency of oxidation. Under the herein tested conditions, it was observed that the MA_H KL showed the highest susceptibility to CotA oxidation when compared to the other hardwood KLs and the softwood KLs. Therefore, this could be a viable method to produce sustainable resins and adhesives.

Melanin production and laccase mediated oxidative stress alleviation during fungal-fungal interaction among basidiomycete fungi

Fungal-fungal interaction often leads to the change in metabolite profile of both the interacting fungus which may have potential implication in industry or agriculture. In the present study, we performed two sets of fungal-fungal interaction—Trametes coccinea (F3) with Leiotrametes lactinea (F9) and T. coccinea (F3) with T. versicolor (F1) to understand the changes in the metabolite profile during the interaction process and how this process impacts the hyphal/mycelial morphology of the participating fungi. The metabolites produced during interaction of T. coccinea (F3) with L. lactinea (F9) and T. coccinea (F3) with T. versicolor (F1) was analysed through liquid chromatography coupled to mass spectroscopy (LC-MS). Most of the metabolites secreted or produced during interaction are associated with defensive response. Further, visualization with scanning electron microscopy revealed that interaction between the tested fungi led to the changes in the hyphal morphology. The bipartite fungal interaction resulted in the production of a dark brown colour pigment—melanin as confirmed by the LC-MS, FTIR and NMR analysis. Moreover, the fungal–fungal interaction also led to increase in the production of laccase, a group of multicopper oxidases involved in detoxification of toxic compounds. Further, increased activity of superoxide dismutase, an enzyme that catalyzes the dismutation of the superoxide anion to hydrogen peroxide was also recorded during fungal–fungal interaction. Quantitative real-time PCR revealed upregulation of lcc1 (encoding a laccase enzyme) and few other stress related genes of T. versicolor during its hyphal interaction with T. coccinea, suggesting a direct correlation between laccase production and melanin production.

Biodelignification of sugarcane shoots: agricultural waste management strategy as an alternative feed for ruminants

This research aimed to reduce the lignin content and increase the nutrients content of sugarcane shoots as forages for a ruminant. This research used an experimental method using a completely randomized design (CRD) in factorial patterns, whereas A factor was a type of fungi (Pleurotus ostreatus and Aspergillus oryzae), and B factor was biodelignification time (14. 21 and 28 days). The variables observed in the experiment were laccase enzyme activity, Crude Protein (CP) content, and the percentage of lignin decreased. The research results showed that biodelignification using Pleurotus ostreatus fungi for 28 days resulted: 1,62 U/ml of laccase enzyme activity, 9.23% crude protein content, and 12.83 % of lignin decreased. From this research, the best treatment for bio-delignification of sugarcane shoots was with Pleurotus ostreatus fungi for 28 days, producing the best sugarcane shoots with 9.23 % of crude protein 12.83% of lignin decreased.

New Laccase-Mediated System Utilized for Bio-Discoloration of Indigo-Dyed Denim Fabrics

In this study, indigo-dyed denim fabric was decolorized by washing and printing with separate and simultaneous applications of laccase enzyme, sodium hydrosulfite, and cellulase enzyme. In this regard, the surface reflectance and color coordinates of the discolored fabrics were analyzed, and SEM photographs of the treated fabrics were prepared to analyze their surfaces. Finally, the effects of the discoloration process and materials on various parameters of the treated samples were investigated, including moisture content, creaserecovery angle, air permeability, and abrasion resistance. The color experiments showed that the discoloration mechanism with the combined use of laccase enzyme, sodium hydrosulfite, and cellulose enzyme had a significant effect on the improvement of the lightness (L*) of the samples, as the lightness of the treated samples was improved by 101.18 percent and 55.79 percent in both printing and washing, respectively. Furthermore, examination of specimen color coordinates revealed that the hue of the treated samples was changed to blue and green, and the purity of color (C*) was improved. The increased moisture content and air permeability of the treated specimens suggested that the comfort of the jeans clothing provided by these treatment methods had improved. As a result, it should be noted that the mediating action of sodium hydrosulfite was significantly influential for discoloration of denim with the laccase enzyme.

Deciphering the Amendment-Induced Secretion of Ligninolytic Enzymes by Pleurotus pulmonarius

Pleurotus pulmonarius belonging to the white-rot fungal basidiomycetes group secretes extracellular ligninolytic enzymes for the degradation of agroresidues. The present study was carried out to evaluate the efficacy of different agro-residues for the enhanced production of ligninolytic enzymes and to authenticate their ability by protein analysis. The morphological and molecular sequences of white-rot fungi were characterised. Besides, the efficacy of organic and inorganic amendments in the secretion of ligninolytic enzymes by P. pulmonarius was characterised using SDS-PAGE and native PAGE analysis. The characterised strain of P. pulmonarius secreted enhanced laccase enzyme levels in the liquid medium through supplementation with organic and inorganic amendments. Wheat bran and groundnut cake each @5% enhanced secretions of Laccase, LiP and MnP. Copper sulphate at 150 μM enhanced the laccase enzyme and at 100 μM enhanced the LiP enzyme level by P. pulmonarius. Similarly, supplementation with manganese sulphate at 150 μM enhanced laccase, LiP and MnP enzyme levels compared to control. SDS-PAGE results showed protein banding patterns in the range of 50–85 kDa for the Lac enzyme in samples drawn from wheat bran and groundnut cake-supplemented substrates. Native PAGE results of laccase enzymes also showed that wheat bran (5%) + groundnut cake (5%) + CuSO4 (150 M) + MnSO4 (150 M) induced four laccase isozymes. Supplementing organic and inorganic amendments to the substrates would enhance the secretion of laccase enzyme that would aid in better breakdown of lignin.

Recovery of Lignocellulolytic Enzymes and Valorization of Spent Mushroom Substrate

Spent Mushroom Substrate (SMS) comprises sugarcane bagasse, coconut coir, chicken manure, and paddy straw; inoculated with and farmed for Agaricus bisporus. At present, the waste generation at a mushroom cultivation plant in Goa is 40 tons/day (15,000 tons annually). Valorization of this waste has been explored in terms of extracting lignocellulolytic enzymes and briquette production. SMS was screened for the presence of lignocellulolytic enzymes and then was used to make briquettes. The enzymes found in SMS were cellulase and laccase, which were further concentrated via tangential flow filtration (TFF). Enzyme activity for Cellulase increased by four-fold (from 255.34±1.30 U/mL increased to 1022.21±4.84 U/mL) and Laccase increased by three-fold (from 4.83±0.02 U/mL to 13.21±0.05 U/mL). The concentrated enzyme cocktail was used to decolorize congo red dye. After only eight hours of enzymatic treatment at pH 4.8 on congo red, approx. 40-49% decolorization was accomplished. The color removal was due to the presence of the laccase enzyme. After enzyme extraction, all the residual SMS was utilized to generate briquettes with an initial reduction in its moisture content from 50% to 10%. The resulting briquette gave a Gross Calorific Value of 4,143 Kcal/kg with 12.60% ash content. Thus, SMS proves to be a valuable source for recovering enzymes and a cost-effective material for briquette production rather than going into landfills.