Application of Deconica castanella ligninolytic enzymatic system in the degradation of hexachlorobenzene in soil.

Hexachlorobenzene (HCB) is a pollutant still found in the environment despite being widely banned. Considering that basidiomycetes are useful to degrade a variety of organochlorinated pollutants, we therefore report the influence of HCB on the ligninolytic enzymatic system of Deconica castanella. The inoculum was prepared with sugarcane bagasse and soybean flour and was added in soil with and without HCB (2000 mg kg soil-1 ), 5% emulsion containing soybean oil and Tween 20 at proportion 9:1, v:v; with 70% moisture at 25°C. Fungal biomass was quantified by widely acknowledged growth biomarker ergosterol. The extraction of the enzymatic complex was performed and laccase, Mn-dependent peroxidase (MnP), and lignin peroxidase (LiP) activities were determined. Furthermore, HCB and its metabolites were quantified by gas chromatography and chlorides by potentiometric titration. Results evidenced that HCB did not interfere in fungal growth, though the only detected enzymatic activity was laccase. MnP and Lip were not detected during D. castanella growth in soil. The peak of laccase enzymatic activity occurred in the presence of HCB. In addition, the laccase exhibited thermostability. Therefore, we hereby shed light on the role of laccase in the degradation of HCB by an efficient low-cost and environmentally safe detoxification mechanism.

Characterization of a thermostable Deconica castanella Laccase and application toward pentachlorophenol degradation.

Pentachlorophenol (PCP) is an organochlorine pesticide whose toxicity led it to be banned in several countries. In Brazil however, this compound is widely accessible, and its indiscriminate use leads to extensive soil contamination, which requires henceforth, the development of new approaches to manage PCP presence in environment. Considering that PCP is susceptible to undergo enzymatic degradation, this investigation is thence, aimed at the purification, and characterization, of a thermostable fungal enzyme (i.e. Laccase of Deconica castanella (Dc-Lac), and assess its role in PCP in vitro biodegradation. Results evidenced that, molecular mass of the partially purified Dc-Lac was estimated to be of 64 kDa, presenting apparent Km of 0.47 µmol, and Vmax of 11.56 U mg-1. The optimum temperature and pH were 55 °C and 2.5, respectively. The T½ verified at 55, 60, and 80 °C were 19 and 17 hr, and 47 min, respectively. The highest PCP biodegradation was of 23% at pollutant concentration of 100 µg L-1, which evidenced that Dc-Lac is a thermostable enzyme that acted directly in PCP degradation and may be a useful asset to remediate this pollutant.

Voltammetric Evaluation of Diclofenac Tablets Samples through Carbon Black-Based Electrodes.

Diclofenac (DIC) is a non-steroidal anti-inflammatory drug of wide use around the world. Electroanalytical methods display a high analytical potential for application in pharmaceutical samples but the drawbacks concerning electrode fouling and reproducibility are of major concern. Henceforth, the aim of this work was to propose the use of alternative low-cost carbon black (CB) and ionic liquid (IL) matrix to modify the surface of pencil graphite electrodes (PGE) in order to quantify DIC in raw materials, intermediates, and final products, as well as in stability assays of tablets. The proposed method using CB+IL/PGE displayed good recovery (99.4%) as well as limits of detection (LOD) of 0.08 µmol L-1 and limits of quantification (LOQ) of 0.28 µmol L-1. CB+IL/PGE response was five times greater than the unmodified PGE. CB+IL-PGE stands as an interesting alternative for DIC assessment in different pharmaceutical samples.

Electrochemical Characterization of Central Action Tricyclic Drugs by Voltammetric Techniques and Density Functional Theory Calculations.

This work details the study of the redox behavior of the drugs cyclobenzaprine (CBP), amitriptyline (AMP) and nortriptyline (NOR) through voltammetric methods and computational chemistry. Results obtained in this study show that the amine moiety of each compound is more likely to undergo oxidation at 1a at Ep1a ≈ 0.69, 0.79, 0.93 V (vs. Ag/AgCl/KClsat) for CBP, AMP and NOR, respectively. Moreover, CBP presented a second peak, 2a at Ep2a ≈ 0.98 V (vs. Ag/AgCl/KClsat) at pH 7.0. Furthermore, the electronic structure calculation results corroborate the electrochemical assays regarding the HOMO energies of the lowest energy conformers of each molecule. The mechanism for each anodic process is proposed according to electroanalytical and computational chemistry findings, which show evidence that the methods herein employed may be a valuable alternative to study the redox behavior of structurally similar drugs.

Physiological Characterization of Fungal Inoculum for Biotechnological Remediation of Soils

The aim of this work was to study the bioremediating potential of Lentinus crinitus CCIBt2611 according to the physiological condition of the inoculum. Inoculum was prepared using sugarcane ground husk (C:N 90), at several physiological ages and applied in soil contaminated with pentachlorophenol. The inoculum's potential was assessed by evaluating the mycelium's vigor at soil's colonization, determination of peroxidase and phenoloxidase activities, in vitro degradation of Remazol Brilliant Blue R and in vivo degradation of pentachlorophenol. The results showed that the assessed parameters were relevant to identify the quality of the inoculum. For L. crinitus, 10 day old inoculum showed good soil-colonization speed with significant enzymatic activities, indicating the role of Manganese-dependent peroxidase and laccase in degradation, and efficient degradation of pentachlorophenol.

Interference of a commercial catalase preparation in laccase and peroxidase activities

The influence of commercial catalase preparations (fungal and bovine origin) on laccase and peroxidase activity assays was evaluated using enzymatic extracts obtained from several basidiomycetes grown under different culture conditions. No hydrogen peroxide was detected in the extracts. Inhibition of laccase activity by 40 to 80 percent was related to the catalase source. In addition, oxidation of the substrate (ABTS) by fungal catalase in the absence of the enzymatic extract from basidiomycetes was observed. The results demonstrated the need for the evaluation of interference of the commercial catalase preparation when its use was required in the reaction mixture.

A influência da preparação comercial de catalase (origem fúngica e bovina) nos ensaios de atividade absence and presence of a fungal or bovine de lacase e de peroxidases foi avaliada empregando-se extratos enzimáticos obtidos do crescimento de diversos basidiomicetos em diferentes condições de cultivo. Não foi detectado H2O2 nos extratos. Inibição de 40 a 80 por cento da atividade de lacase foi relacionada à fonte de catalase. Além disso, foi observada oxidação do substrato (ABTS) pela catalase fúngica, na ausência de extrato enzimático do basidiomiceto. Os resultados evidenciaram a necessidade de se proceder a uma avaliação da interferência da preparação comercial de catalase, quando o seu uso se fizer necessário na mistura reacional.

Characterization of fungal inoculum used in soil bioremediation

Studies have indicated the capacity of basidiomycetes to degrade recalcitrant organopollutants. However, the age of the fungal inoculum to obtain a more effective degradation has not been defined. The criterion used is total colonization of the substrate. Psilocybe castanella CCB444 and Lentinus crinitus CCB274 have been evaluated in soils containing hexachlorobenzene. In the present study, the physiological conditions of the fungal inocula were characterized on solid substrate (sugarcane bagasse, starch and soy flour). Colonization of the substrate, loss of organic matter, pH variation, organic carbon, total nitrogen, fungal biomass and enzymatic activity were evaluated over 30 days of incubation. Colonization of the substrate was almost complete after 20 days for both species, with about 90 percent of organic matter remaining on the substrates. The pH continued to be acid during incubation. The highest enzymatic production was observed at 10 days for L. crinitus and at 5 days for P. castanella. The fungi presented growth up to 30 days. The C/N ratio of the inocula showed little variation. The use among 10 and 15-day-old inoculum is adequate since sufficient nutrients are left to guarantee survival of the fungus, vigorous colonization of the substrate, a growing biomass and an active enzymatic system, thus permitting fungal growth in soil.

Estudos indicam capacidade de basidiomicetos em degradar organopoluentes recalcitrantes. Porém, ainda não foi padronizada a idade do inóculo fúngico a ser aplicado para que ocorra uma degradação mais efetiva. O critério utilizado é colonização total do substrato. Psilocybe castanella CCB444 e Lentinus crinitus CCB274 têm sido avaliados em solos com hexaclorobenzeno. No presente trabalho, foram caracterizadas as condições fisiológicas dos inóculos fúngicos em substrato sólido (bagaço de cana-de-açúcar , amido e farinha de soja). Determinou-se a colonização do substrato, perda de matéria orgânica, variação de pH, carbono orgânico, nitrogênio total, biomassa fúngica e atividade enzimática durante 30 dias de incubação. A colonização do substrato foi quase completa após 20 dias, para ambos os fungos, restando cerca de 90 por cento de matéria orgânica nos substratos. O pH permaneceu ácido durante incubação. A maior produção enzimática foi aos 10 dias para L. crinitus e aos 5 dias para P. castanella. Os fungos apresentaram crescimento até os 30 dias. A relação C/N dos inóculos variou pouco. Conclui-se que, inóculos entre 10 e 15 dias de idade podem ser utilizados, pois ainda restam nutrientes suficientes para manter a sobrevivência do fungo, colonização vigorosa do substrato, biomassa crescente e sistema enzimático ativo, permitindo seu crescimento no solo.