Culture supernatant containing laccase produced by Cerrena unicolor strain was used to examine laccase partitioning between phases in an aqueous two-phase system. The investigated system consisted of polyethylene glycol 3000 and sodium phosphate buffer adjusted to pH = 7. Influence of several parameters on partitioning was measured, including phase forming components’ concentrations, tie line lengths, phase volume ratio, supernatant dilution, process temperature and halogen salt supplementation. Partitioning coefficients up to 78 in the bottom phase were achieved with yields of over 90%. Tie line length and phase volume ratio had significant effect on enzyme partitioning.
Foam fractionation process for concentration of laccases from two Basidiomycete strains under different process conditions was investigated. Culture supernatants of Cerrena unicolor and Pleurotus sapidus containing active laccase were used with and without surfactant additives. Two surfactants: cationic cetrimonium bromide (CTAB) and non-ionic Polysorbate 80 were applied in the range from 0.2 mM to 1.5 mM. The pH levels ranging from 3 to 10 were examined with particular attention to pH=4, which is close to the pI of the enzymes. Results show that the source of the enzyme is significant in terms of partitioning efficiency in a foam fractionation process. Laccase from Cerrena unicolor showed the best activity partitioning coefficients between foamate and retentate of almost 200 with yields reaching 50% for pH 7.5 and concentration of CTAB cCTAB = 0.5 mM, whereas laccase from Pleurotus sapidus showed partitioning coefficients of up to 8 with 25% yield for pH 4 and cCTAB = 0.5 mM.
Extracellular laccase produced by the wood-rotting fungus Cerrena unicolor was immobilised covalently on the mesostructured siliceous foam (MCF) and three hexagonally ordered mesoporous silicas (SBA-15) with different pore sizes. The enzyme was attached covalently via glutaraldehyde (GLA) or by simple adsorption and additionally crosslinked with GLA. The experiments indicated that laccase bound by covalent attachment remains very active and stable. The best biocatalysts were MCF and SBA-15 with Si-F moieties on their surface. Thermal inactivation of immobilised and native laccase at 80°C showed a biphasic-type activity decay, that could be modelled with 3- parameter isoenzyme model. It appeared that immobilisation did not significantly change the mechanism of activity loss but stabilised a fraction of a stable isoform. Examination of time needed for 90% initial activity loss revealed that immobilisation prolonged that time from 8 min (native enzyme) up to 155 min (SBA-15SF).
Diclofenac (2-[(2,6-Dichlorophenyl)amino]benzeneacetic acid) is a non-steroidal anti-infl ammatory drug. Due to excessive use of diclofenac, this drug has been detected in surface water, ground water and drinking water. In our study, four fungal strain Trametes trogii, Aspergillus niger, Yarrowia lipolytica and Phanerochaete chrysosporium were investigated in terms of diclofenac degradation potential. Trametes trogii was found to be the most effi cient strain with 100% diclofenac degradation rate. Two hydroxylated diclofenac metabolites have been identifi ed in culture medium. Crude laccase from T. trogii almost completely removed diclofenac with 97% removal in 48 h. We suggest that the degradation of diclofenac depends on the cytochrome P450 enzyme system and laccase activity. After 24 h incubation decrease in toxicity of diclofenac was confi rmed by Microtox test.
The current study was aimed to evaluate the industrial efﬂ uents biodegradation potential of an indigenous microorganism which reduced water pollution caused by these efﬂ uents. In the present study biodegradation of three textile industrial efﬂ uents was performed with locally isolated brown rot fungi named Coniophora puteana IEBL-1. Response Surface Methodology (RSM) was employed under Box Bhenken Design (BBD) for the optimization of physical and nutritional parameters for maximum biodegradation. Quality of treated efﬂ uents was checked by study of BOD, COD and analysis through HPLC. Three ligninolytic enzymes named lignin peroxidase, manganese peroxidase and laccase were also studied during the biodegradation process. The results showed that there was more than 85% biodegradation achieved for all three efﬂ uents with decrease in Biological Oxygen Demand (BOD) and Chemical Oxygen Demand (COD) below the recommended values for industrial efﬂ uent i.e. 80 mg/L for BOD and 220 mg/L for COD after optimization of nutritional parameters in the second stage. Analysis of samples through HPLC revealed the formation of less toxic diphenylamine, 3-methyldiphenylamine and N-methylaniline after treatment. The ligninolytic enzymes assays conﬁ rmed the role of lignin peroxidase (LiP), manganese peroxidase (MnP) and laccase in biodegradation process. Lignin peroxidase with higher activity has more contribution in biodegradation of efﬂ uents under study. It can be concluded through the results that Coniophora buteana IEBL-1 is a potential fungus for the treatment of industrial efﬂuents.