Search Results (19)

In the present investigation, an eco-friendly biocatalyst was developed using Pleurotus eryngii laccase (PeLac) through a copper (Cu)-based protein–inorganic hybrid system for the degradation of bisphenol A, a representative xenobiotic. After partial purification, the specific activity of crude PeLac was 92.6 U/mg of total protein. Immobilization of PeLac as Cu₃(PO₄)₂–Lac (Cu–PeLac) nanoflowers (NFs) at 4 °C resulted in a relative activity 333% higher than that of the free enzyme. The Cu–PeLac NFs exhibited greater pH and temperature stability and enhanced catalytic activity compared to free laccase. This enhanced activity was validated through improved electrochemical properties. After immobilization, Cu–PeLac NFs retained up to 8.7-fold higher residual activity after storage at 4 °C for 30 days. Free and immobilized laccase degraded bisphenol A by 41.6% and 99.8%, respectively, after 2 h of incubation at 30 °C. After ten cycles, Cu–PeLac NFs retained 91.2% degradation efficiency. In the presence of potent laccase inhibitors, Cu–PeLac NFs exhibited a 47.3-fold improvement in bisphenol A degradation compared to free PeLac. Additionally, the synthesized Cu–PeLac NFs demonstrated lower acute toxicity against Vibrio fischeri than Cu nanoparticles. This study presents the first report of PeLac immobilization through an eco-friendly protein–inorganic hybrid system, with promising potential for degrading bisphenol A in the presence of inhibitors to support sustainable development. Full article

Improving the utilization of spent mushroom substrate and enhancing the digestibility of straw-based feed are critical for promoting environmental sustainability. However, the effects of replacing sawdust with straw in the cultivation of Pleurotus ostreatus—including changes in physicochemical properties, enzyme activities, and microbial community structure and function—remain unclear. In this study, corn straw was used as the substrate for P. ostreatus cultivation. Dynamic changes during the fermentation process were investigated through analyses of biological growth characteristics, physicochemical properties, enzyme activities, and amplicon sequencing. The results indicated a significant increase in mushroom yield, with the M80% treatment group achieving a yield of 156.09 ± 7.15 g. The nutritional value of the fermented feed was markedly improved; after 50 days of fermentation, crude protein (CP) and ether extract (EE) contents increased by 5.42% and 0.79%, respectively, while acid detergent fiber (ADF) and neutral detergent fiber (NDF) contents decreased by 18.5% and 22.3%, compared to day 0. Activities of cellulase, xylanase, and laccase were also elevated, contributing to more effective lignocellulose degradation. Furthermore, Illumina sequencing revealed shifts in bacterial and fungal metabolic pathways. The fungal community was dominated by Ascomycota and Basidiomycota, with Pleurotus as the prevailing genus, while the bacterial community was mainly composed of antagonistic genera such as Bacillus and Bacteroides. These findings provide a theoretical basis for understanding the role of microbial interactions during straw substrate fermentation in improving feed quality and increasing P. ostreatus yield. Full article

Laccases are multicopper oxidases that exhibit a broad substrate spectrum, making them excellent biocatalysts for clean technological processes. The study isolated novel laccase-producing bacteria from decomposed wood samples and characterized the enzyme for potential industrial and biotechnological applications. The results showed that three bacteria, SP-2, SP-1, and WP-2, out of the eight isolated bacteria, oxidized both guaiacol and α-naphthol in the plate assay and exhibited extracellular laccase activity of 7.0 ± 0.01, 6.67 ± 0.02, and 7.40 ± 0.04 (U/mL), respectively. WP-2 was selected for further study and was identified as Bacillus sp. GLN (accessioned number: MK290989). Strain GLN maximally secreted laccase 48 h post-fermentation, with an enzyme yield of 36.83 ± 2.47 U/mL in optimized conditions. The crude laccase was optimally active at pH 9.0 and 90 °C and showed excellent pH and thermal stability, retaining approximately 65% residual activity at 100 °C for 270 min. GLN laccase demonstrated remarkable stability after treatment with organic solvents and metal ions, retaining more than 50% of its original activity in the presence of 100 mM inhibitors. The data from this study highlight the relevance of Bacillus sp. GLN and its laccase in promoting clean technology. Full article

In this study, Candida utilis, Lactobacillus plantarum, and non-starch polysaccharide enzymes (cellulase, laccase, β-glucanase, xylanase, and mannanase) were employed to examine the effects of various microorganism–enzyme combinations on the nutritional composition, fiber structure, and fermentation quality of corn stover and wheat straw. Furthermore, the synergistic effects of these treatments were assessed through the use of in vitro rumen fermentation. The results showed that the microorganism–enzyme combinations significantly increased the crude protein content (p < 0.05), while reducing the acid detergent fiber and neutral detergent fiber levels (p < 0.05) in both substrates. The fermentation broth pH decreased (p = 0.06 for corn stover; p < 0.05 for wheat straw) as a result of the treatments, with a significant increase in the lactate concentration (p < 0.05). The reducing sugar levels varied across the treatments (p < 0.05). Mycotoxin analysis revealed trace amounts of zearalenone, well below the Chinese feed hygiene standard. Scanning electron microscopy showed structural modifications, including fiber breakage and surface wrinkling, in the treated substrates. In vitro rumen fermentation demonstrated significant changes in the NH₃-N production and volatile fatty acid profiles (p < 0.05). In conclusion, the addition of different microorganism–enzyme combinations can effectively improve the nutritional composition, fiber structure, and fermentation quality of corn stover and wheat straw. Among the treatments, the T3 group (25% each of C. utilis, L. plantarum, cellulase, and laccase, with a total addition ratio of 0.3% w/w) exhibited the most pronounced improvement in nutritional value for both corn stover and wheat straw. These findings suggest that microorganism–enzyme combinations effectively enhance the nutritional and fermentative quality of agricultural residues. Full article

Fungal endophytes of medicinal plants produce diverse secondary metabolites and extracellular enzymes with therapeutic and biotechnological potential. However, the biological and biotechnological potential of fungal endophytes from South African medicinal plants remain relatively underexplored. In this study, the antimicrobial, antioxidant, anti-inflammatory and extracellular enzymatic capabilities of five fungal endophytes previously isolated from the leaves of Kirkia acuminata Oliv. were investigated. Sequencing of the internal transcribed spacer (ITS) regions revealed that the isolates belonged to the genera Setosphaeria, Diaporthe and Corynespora. The broth micro-dilution assay and the Folin–Ciocalteau reagent method were used to assess the antibacterial activity and the total phenolic content (TPC) of the fungal endophytes’ ethyl acetate crude extracts (CEs), respectively. The antioxidant activity was assessed using the ferric reducing antioxidant power (FRAP) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging assays. The influence of the CE of the Setosphaeria rostrata KaL-4 on the viability and LPS-induced interleukin-6 (IL-6) production in Raw 264.7 macrophages was assessed using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and an ELISA, respectively. The ability of the isolates to produce extracellular proteases, laccases and peroxidases was also determined. The CEs displayed antimicrobial activity with MICs ranging from 0.63 to 1.25 mg/mL and reducing power and scavenging activity ranging from 40% to 18% and from 60% to 48%, respectively. The S. rostrata KaL-4 CE possessed the highest TPC and demonstrated dose-dependent cytotoxicity. The CE further demonstrated a significant reduction in IL-6 production at a concentration of 0.75 µg/mL. Only one isolate demonstrated the ability to produce proteases with an enzymatic index (EI) of 0.66, while laccases (EI range of 0.14 to 1.15) and peroxidases were produced by all of the isolates. These findings suggest that fungal endophytes from South African medicinal plants are promising sources of bioactive compounds and industry-significant extracellular enzymes. Full article

The aim of the present research was the efficient degradation of industrial textile wastewater dyes using a very active cloned laccase enzyme. For this purpose, potent laccase-producing bacteria were isolated from soil samples collected from wastewater-replenished textile sites in Punjab, Pakistan. The laccase gene from locally isolated strain LI-81, identified as Bacillus megaterium, was cloned into vector pET21a, which was further transformed into E. coli BL21 codon plus. The optimized conditions for the increased production of laccase include fermentation in a 2% glucose, 5% yeast extract and 250 mg/L CuSO₄ medium with pH 7.5; inoculation with 5% inoculum; induction with 0.1 mM IPTG at 0.5 O.D.; and incubation for 36 h at 37 °C. The crude enzyme produced was employed for the removal of commercially used textile dyes. The dyes were quickly precipitated under optimized reaction conditions. Rose bengal, brilliant green, brilliant blue G, Coomassie brilliant blue R and methylene blue were precipitated at rates of 10.69, 54.47, 84.04, 78.99 and 7.40%, respectively. The FTIR and UV–Vis spectroscopic analyses of dyes before and after confirmed the chemical changes brought about by the cloned laccase that led to the dye removal. Full article

Grape pomace (GP) is considered a natural source of bioactive compounds. To improve the extractability of bioactive compounds, in this work, GP was biologically treated for 15 days with the white-rot fungus Trametes versicolor in laboratory jars and a tray bioreactor under solid-state fermentation (SSF) conditions. During SSF, the activity of lignolytic (laccase and manganese peroxidase) and hydrolytic (xylanase, cellulase, β-glucosidase, and invertase) enzymes was measured, with the activities of laccase (2.66 U/g_db in jars and 0.96 U/g_db in the bioreactor) and xylanase (346.04 U/g_db in jars and 200.65 U/g_db in the bioreactor) being the highest. The effect of the complex enzyme system was reflected in the changes in the chemical composition of GP with increasing ash, crude protein, and free fat content: 28%, 10%, and 17% in the laboratory jars, and 29%, 11%, and 7% in the bioreactor, respectively. In addition, the biological treatment improved the extractability of 13 individual phenolic compounds. Therefore, the applied SSF technique represents an effective strategy to improve the profile of phenolic compounds and the nutritional composition of GP, promoting their valorization and opening the door for potential applications in the food industry and other sectors. Full article

Edible fungi have certain photo-sensitivity during the mushroom emergence stage, but there have been few relevant studies on the responses of Ganoderma lucidum to different light irradiation conditions. Ganoderma lucidum were planted in an environmentally controllable mushroom room with different light supply modes that were, respectively, continuous white light (CK), red light (R), green light (G), blue light (B), and intermittent red light (R-), green light (G-), and blue light (B-), with a total light intensity of 15 μmol·m⁻²·s⁻¹ and a light/dark (L/D) period of 12 h/12 h for each treatment. The interval in intermittent light treatments was 30 min. The optimal light supply mode suitable for the growth of Ganoderma lucidum was explored by analyzing the characteristics, nutritional quality, and extracellular enzyme activity in mushrooms exposed to different light treatments. The results showed that red light (whether in continuous or intermittent supply modes) inhibited the fruiting body differentiation of Ganoderma lucidum, showing delayed differentiation or complete undifferentiation. The highest stipe length and pileus diameter of fruiting bodies were detected under G- treatment, which were, respectively, increased by 71.3% and 3.2% relative to the control. The highest weight of fruiting bodies was detected under G treatment, which was significantly increased by 21.4% compared to the control (p < 0.05). Intermittent light mode seemed to be more conducive to the size development of the fruiting body, while continuous light mode was beneficial for increasing the weight. The highest contents of crude protein and total triterpenes in pileus were detected under G treatment (significantly 14.9% and 28.1% higher than the control, respectively), while that of the crude polysaccharide was detected under G- treatment (significantly 35.7% higher than the control) (p < 0.05). The highest activities of extracellular enzymes such as cellulase, hemicellulase, laccase, lignin peroxidase, and amylase were detected in fruiting bodies subjected to G treatment, which were significantly increased by 11.9%~30.7% in the pileus and 9.5%~44.5% in the stipe. Green light might increase the weight and nutrient accumulation in the pileus of Ganoderma lucidum via up-regulating the extracellular enzyme activities. This study provides an effective light supply strategy for regulating the light environment in the industrial production of Ganoderma lucidum. Full article

In the present investigation, an ecofriendly magnetic inorganic-protein hybrid system-based enzyme immobilization was developed using partially purified laccase from Trametes versicolor (TvLac), Fe₃O₄ nanoparticles, and manganese (Mn), and was successfully applied for synthetic dye decolorization in the presence of enzyme inhibitors. After the partial purification of crude TvLac, the specific enzyme activity reached 212 U∙mg total protein⁻¹. The synthesized Fe₃O₄/Mn₃(PO₄)₂-laccase (Fe₃O₄/Mn-TvLac) and Mn₃(PO₄)₂-laccase (Mn-TvLac) nanoflowers (NFs) exhibited encapsulation yields of 85.5% and 90.3%, respectively, with relative activities of 245% and 260%, respectively, compared with those of free TvLac. One-pot synthesized Fe₃O₄/Mn-TvLac exhibited significant improvements in catalytic properties and stability compared to those of the free enzyme. Fe₃O₄/Mn-TvLac retained a significantly higher residual activity of 96.8% over that of Mn-TvLac (47.1%) after 10 reuse cycles. The NFs showed potential for the efficient decolorization of synthetic dyes in the presence of enzyme inhibitors. For up to five reuse cycles, Fe₃O₄/Mn-TvLac retained a decolorization potential of 81.1% and 86.3% for Coomassie Brilliant Blue R-250 and xylene cyanol, respectively. The synthesized Fe₃O₄/Mn-TvLac showed a lower acute toxicity towards Vibrio fischeri than pure Fe₃O₄ nanoparticles did. This is the first report of the one-pot synthesis of biofriendly magnetic protein-inorganic hybrids using partially purified TvLac and Mn. Full article

Large quantities of wine lees are produced annually by the wine industry. The high phenolic content makes them unsuitable for disposal in the environment or animal feed without a suitable treatment. In this study, wine lees were treated by Pleurotus ostreatus in submerged cultivation, producing a high-value biomass and elevated levels of laccase, an important industrial enzyme. Biomass and laccase production reached 21 g/L and 74,000 Units/L, respectively, at the optimal conditions of initial pH 6.0, 20% v/v wine lees, 30 g/L glucose, and 20 g/L yeast extract, while decolorization and dephenolization rates of the waste were over 90%. The mycelial biomass was rich in proteins and essential amino acids reaching up to 43% and 16% per dry weight, respectively. Carbohydrates and lipids were the second richest bioactive compound in biomass, with values of 29.4 ± 2.7% and 29.5 ± 2.7%, respectively. The crude laccase in the culture supernatant was purified via a simple two-step purification procedure by 4.4-fold with a recovery of 44%. The molecular weight of the enzyme was determined to be 62 kDa via SDS electrophoresis. Enzyme activity was optimal at pH 5.0 and 70 °C. The activation energy of the enzyme was calculated at a value of 20.0 ± 0.2 kJ/mol. The pH stability and thermostability of the purified laccase were studied. The enzyme was remarkably stable at pH 8.0 and at temperatures up to 40 °C. The thermal inactivation energy of the enzyme was determined to be 76.0 ± 1.2 kJ/mol. The thermodynamic parameters (ΔH*, ΔG*, and ΔS*) for the thermal deactivation of the purified laccase at a temperature range of 20–60 °C were: 73.8 ≤ ΔH* ≤ 74.3 kJ·mol⁻¹, 98.7 ≤ ΔG* ≤ 101.9 kJ·mol⁻¹, and −90.5 ≤ ΔS* ≤ −84.3 J·mol⁻¹·K⁻¹. Wine lees could be ideal substrates of fungal cultivation for laccase production and biomass with a high protein content in an eco-friendlier way. Full article

This work focused on obtaining fermented oil cake (cotton or Jatropha) via macrofungi growth with potential characteristics for animal feed formulations, such as the presence of extracellular enzymes, bioactive (ergosterol and antioxidants), and detoxification of antinutritional compounds. The concentration of phorbol esters was reduced by four macrofungi in Jatropha seed cake (JSC) to non-toxic levels. At least two macrofungi efficiently degraded free gossypol in cottonseed cake (CSC). Fermentation with Coriolopsis sp. INPA1646 and Tyromyces sp. INPA1696 resulted in increased ergosterol concentrations, antioxidant activity reduction, and high activity of laccases and proteases. Bromatological analysis indicated high crude protein concentrations, with partial solubilization by fungal proteases. Fermented products from Coriolopsis sp. and Tyromyces sp. in JSC or CSC can be considered important biological inputs for monogastric and polygastric animal feed. Full article

One of the main challenges in both the design of new wastewater treatment plants and the expansion and improvement of existing ones is the removal of emerging pollutants. Therefore, the search for economic and sustainable treatments is needed to enhance the removal of pharmaceuticals. The potential of a lignocellulosic substrate colonized by Pleurotus ostreatus, a waste from mushroom production, to remove fluoxetine from aqueous solutions was studied. Batch assays were performed to remove 600 µg∙L⁻¹ fluoxetine from aqueous solutions using the colonized mushroom substrate (CMS) and crude enzyme extracts. The removal efficiencies achieved were, respectively, ≥83.1% and 19.6% in 10 min. Batch assays with sterilized CMS and 1-aminobenzotriazole (to inhibit cytochrome P450 enzymes) showed that the higher removal efficiencies achieved in the CMS assays may be attributed to the synergistic contribution of biosorption onto the CMS and lignin modifying enzymes activity, namely laccase activity. A column assay was performed with the CMS, fed with 750 µg∙L⁻¹ fluoxetine aqueous solution. The removal efficiency was 100% during 30 min, decreasing to a final value of 70% after 8 h of operation. The results suggested that CMS can be a promising eco-friendly alternative to remove fluoxetine from aqueous solutions. Full article

Many free-living saprobic fungi are nature recruited organisms for the degradation of wastes, ranging from lignocellulose biomass to organic/inorganic chemicals, aided by their production of enzymes. In this study, fungal strains were isolated from contaminated crude-oil fields in Nigeria. The dominant fungi were selected from each site and identified as Aspergillus oryzae and Mucor irregularis based on morphological and molecular characterization, with site percentage incidences of 56.67% and 66.70%, respectively. Selected strains response/tolerance to complex hydrocarbon (used engine oil) was studied by growing them on Bushnell Haas (BH) mineral agar supplemented with the hydrocarbon at different concentrations, i.e., 5%, 10%, 15%, and 20%, with a control having dextrose. Hydrocarbon degradation potentials of these fungi were confirmed in BH broth culture filtrates pre-supplemented with 1% engine oil after 15 days of incubation using GC/MS. In addition, the presence of putative enzymes, laccase (Lac), manganese peroxidase (MnP), and lignin peroxidase (LiP) was confirmed in culture filtrates using appropriate substrates. The analyzed fungi grew in hydrocarbon supplemented medium with no other carbon source and exhibited 39.40% and 45.85% dose inhibition response (DIR) respectively at 20% hydrocarbon concentration. An enzyme activity test revealed that these two fungi produced more Lac than MnP and LiP. It was also observed through the GC/MS analyses that while A. oryzae acted on all hydrocarbon components in the used engine oil, M. irregularis only degraded the long-chain hydrocarbons and BTEX. This study confirms that A. oryzae and M. irregularis have the potential to be exploited in the bio-treatment and removal of hydrocarbons from polluted soils. Full article

The oil and gas industry generates large amounts of oil-derived effluents such as Heavy Crude Oil (HCO) in water (W) emulsions, which pose a significant remediation and recovery challenge due to their high stability and the presence of environmentally concerning compounds. Nanomaterials emerge as a suitable alternative for the recovery of such effluents, as they can separate them under mild conditions. Additionally, different biomolecules with bioremediation and interfacial capabilities have been explored to functionalize such nanomaterials to improve their performance even further. Here, we put forward the notion of combining these technologies for the simultaneous separation and treatment of O/W effluent emulsions by a novel co-immobilization approach where both OmpA (a biosurfactant) and Laccase (a remediation enzyme) were effectively immobilized on polyether amine (PEA)-modified magnetite nanoparticles (MNPs). The obtained bionanocompounds (i.e., MNP-PEA-OmpA, MNP-PEA-Laccase, and MNP-PEA-OmpA-Laccase) were successfully characterized via DLS, XRD, TEM, TGA, and FTIR. The demulsification of O/W emulsions was achieved by MNP-PEA-OmpA and MNP-PEA-OmpA-Laccase at 5000 ppm. This effect was further improved by applying an external magnetic field to approach HCO removal efficiencies of 81% and 88%, respectively. The degradation efficiencies with these two bionanocompounds reached levels of between 5% and 50% for the present compounds. Taken together, our results indicate that the developed nanoplatform holds significant promise for the efficient treatment of emulsified effluents from the oil and gas industry. Full article

Laccase enzyme from white-rot fungi is a potential biocatalyst for the oxidation of emerging contaminants (ECs), such as pesticides, pharmaceuticals and steroid hormones. This study aims to develop a three-step platform to treat ECs: (i) enzyme production, (ii) enzyme concentration and (iii) enzyme application. In the first step, solid culture and liquid culture were compared. The solid culture produced significantly more laccase than the liquid culture (447 vs. 74 µM/min after eight days), demonstrating that white rot fungi thrived on a solid medium. In the second step, the enzyme was concentrated 6.6 times using an ultrafiltration (UF) process, resulting in laccase activity of 2980 µM/min. No enzymatic loss due to filtration and membrane adsorption was observed, suggesting the feasibility of the UF membrane for enzyme concentration. In the third step, concentrated crude enzyme was applied in an enzymatic membrane reactor (EMR) to remove a diverse set of ECs (31 compounds in six groups). The EMR effectively removed of steroid hormones, phytoestrogen, ultraviolet (UV) filters and industrial chemical (above 90%). However, it had low removal of pesticides and pharmaceuticals. Full article