Coupling chemical oxidation processes and Leptosphaerulina sp. myco-remediation to enhance the removal of recalcitrant organic pollutants in aqueous systems.

This research evaluated for the first time, the coupling of chemical oxidation processes with Leptosphaerulina sp. (a Colombian fungus), to degrade a refractory pollutant. For such purpose, a model contaminant (crystal violet, CV) was considered. Initially, the pollutant, at high concentrations (i.e., 200 and 50 mg L-1), was submitted to the fungus action. However, the CV inhibited the growth and enzymatic production of the fungus. Then, three chemical oxidation processes: TiO2-photocatalysis, sonochemistry, or electrochemistry (with a Ti/IrO2 anode in sodium chloride) were used as treatments previous to the myco-remediation. These oxidative treatments led to the pollutant degradation (~100%) by the action of radicals or active chlorine species, but they showed low mineralization. Indeed, the total organic carbon removal (TOC) was 54, ~15, and 31% to TiO2-photocatalysis (after 12 h), sonochemistry (after 12 h), and electrochemistry (after 1.33 h), respectively. Thus, the resultant solutions from the chemical oxidations were submitted to the action of Leptosphaerulina sp. (this time effective fungus growth and enzymes production were observed). It was found that the TOC removals by the fungus were 87, 84, and 83% for solutions pre-treated by TiO2-photocatalysis (12 h), sonochemical (12 h), and electrochemical (1.33 h) treatments, respectively. Regarding the enzymatic production, TiO2-photocatalysis/Leptosphaerulina sp., ultrasonication/Leptosphaerulina sp., and electrochemical oxidation/Leptosphaerulina sp. combinations reached the highest activities of laccase (0.6 U mg-1, at day 15), manganese peroxidase (1.35 U mg-1, at day 7) and versatile peroxidase (1.72 U mg-1, at day 15), respectively. The results from this work evidence feasibility of the pre-treatment with chemical oxidation processes as a strategy to enhance Leptosphaerulina sp. action toward recalcitrant organic pollutants (as CV) in water.

Enhancement of ligninolytic enzymes production and decolourising activity in Leptosphaerulina sp. by co-cultivation with Trichoderma viride and Aspergillus terreus.

This work investigated fungal co-culture as inducer of ligninolytic enzymes and decolourising activity in the Colombian strain Leptosphaerulina sp., an ascomycete white-rot fungus isolated from lignocellulosic material. Aspergillus niger, Aspergillus fumigatus, Aspergillus terreus, Trichoderma viride, Fusarium sp. and Penicillium chrysogenum were tested as Leptosphaerulina sp. inducers. The best fungal combinations in terms of enzyme production, fungal growth and decolourising activity were selected from solid media experiments. Response surface methodology (RSM) was utilised to optimise enzyme production and decolourising activity in liquid media. Solid media assays evidenced T. viride and A. terreus as the best Leptosphaerulina sp. inducers. The RSM identified a triple co-culture inoculated with T. viride (1000 µL) and A. terreus (1000 µL) into a 7-day culture of Leptosphaerulina sp. as the best treatment. This triple combination significantly improved ligninolytic enzymes production and Reactive Black 5 dye removal when compared to the Leptosphaerulina sp. monoculture and previously used chemical inducers. These results demonstrated the potential of fungal co-culture as an environmentally-friendly method to enhance Leptosphaerulina sp. enzymes production and decolourising activity.

Decolorization of Reactive Black 5 Dye by Heterogeneous Photocatalysis with TiO 2 /UV / Decoloración del tinte Negro Remazol B mediante fotocatálisis heterogénea con TiO 2 /UV / Descoloração do corante sintético Negro Remazol B por fotocatálise heterogênea com TiO 2 /UV

Abstract Reactive Black 5 (RB5) is an azo dye widely used in the textile industry because of its high chemical stability. Since it does not entirely fix on the fabrics, it pollutes water sources. In this work, the decolorization of aqueous solutions with RB5 was performed by heterogeneous photocatalysis with TiO2/UV. The reaction was carried out in an aluminum photoreactor equipped with five lamps. The effect of TiO2 (0.1, 0.175, and 0.25 g L-1), RB5 concentration (50, 75, and 100 mg L-1), and pH (3, 7, and 11) was evaluated for 14 h, using a Box-Behnken experimental statistical design. Complete decolorization of RB5 was obtained at 14 h, employing 0.175 g L-1 TiO2, 50 mg L-1 RB5, and pH 3. A 98.44% of decolorization was achieved in 10 h (0.25 g L-1 of TiO2, 50 mg L-1 of RB5, and pH 7). The highest decolorization percentage of RB5 (99.51%) was obtained at 10 h of exposure to UV light, using 0.5 g L-1 of TiO2, 50 mg L-1 of the dye, and a pH of 3. Cytotoxicity tests on the HepG2 cell line indicated that photocatalytic degradation of RB5 did not generate cytotoxic byproducts.

Resumen El Negro Remazol B (NRB) es un colorante azoico, usado en la industria textil por su estabilidad química. Este tinte no se fija al 100%, ocasionando contaminación en el agua. En este trabajo se evaluó la decoloración de soluciones acuosas con NRB mediante fotocatálisis heterogénea con TiO2/UV. La reacción se realizó en un fotorreactor de aluminio equipado con cinco lámparas. El efecto de la concentración de TiO2 (0,1; 0,175 y 0,25 g L-1), y NRB (50, 75 y 100 mg L-1) y el pH (3, 7 y 11) fue evaluado durante 14 h, tomando un diseño estadístico experimental Box-Behnken. La decoloración completa del NRB se obtuvo después de 14 h (0,175 g L-1 de TiO2, 50 mg L-1 de NRB, y pH 3). Se alcanzó un 98,44% de decoloración después de 10 h (0,25 g L-1 de TiO2, 50 mg L-1 de NRB, y pH 7). Se encontró que el mejor porcentaje de decoloración del NRB (99,51%) se obtuvo a las 10 h de exposición a luz UV, utilizando 0,5 g L-1 de TiO2, 50 mg L-1 del colorante y un pH de 3. Los ensayos de citotoxicidad sobre la línea celular HepG2 indicaron que la degradación fotocatalítica del NRB no generó subproductos citotóxicos.

Resumo O Negro Remazol B (RNB) é um corante azoico usado na indústria têxtil pela sua estabilidade química. O corante não é fixo no tecido em 100%, então contamina as fontes de água. Neste trabalho, avaliou-se a descoloração de soluções aquosas com RNB por meio de fotocatálise heterogênea com TiO2/UV. A reação foi feita no fotoreactor de alumínio com cinco lâmpadas. O efeito da concentração de TiO2 (0,1; 0,175 e 0,25 g L-1), RNB (50, 75 e 100 mg L-1) e pH (3, 7 e 11) foi avaliado durante 14 h utilizando o modelo estatístico Box-Behnken. Ás 14 h, foi atingida a descoloração completa do RNB (pH 3; 0,175 g L-1 de TiO2 e 50 mg L-1 de RNB). Utilizando 0,25 g L-1 de TiO2, 50 mg L-1 de RNB e pH 7, obteve-se descoloração de 98,44% em 10 h. Encontrou-se que a melhor porcentagem de descoloração do RNB (91,51%) foi obtida com 10 h de exposição à luz UV usando 0,5 g L-1 de TiO2, 50 mg L-1 do corante e pH de 3. Adicionalmente, os testes de citotoxicidade na linha celular HepG2 mostraram que a degradação fotocatalítica do RNB não gerou subprodutos citotóxicos.

Elimination of Isoxazolyl-Penicillins antibiotics in waters by the ligninolytic native Colombian strain Leptosphaerulina sp. considerations on biodegradation process and antimicrobial activity removal.

In this work, Leptosphaerulina sp. (a Colombian native fungus) significantly removed three Isoxazolyl-Penicillin antibiotics (IP): oxacillin (OXA, 16000 µg L-1), cloxacillin (CLX, 17500 µg L-1) and dicloxacillin (DCX, 19000 µg L-1) from water. The biological treatment was performed at pH 5.6, 28 °C, and 160 rpm for 15 days. The biotransformation process and lack of toxicity of the final solutions (antibacterial activity (AA) and cytotoxicity) were tested. The role of enzymes in IP removal was analysed through in vitro studies with enzymatic extracts (crude and pre-purified) from Leptosphaerulina sp., commercial enzymes and enzymatic inhibitors. Furthermore, the applicability of mycoremediation process to a complex matrix (simulated hospital wastewater) was evaluated. IP were considerably abated by the fungus, OXA was the fastest degraded (day 6), followed by CLX (day 7) and DCX (day 8). Antibiotics biodegradation was associated to laccase and versatile peroxidase action. Assays using commercial enzymes (i.e. laccase from Trametes versicolor and horseradish peroxidase) and inhibitors (EDTA, NaCl, sodium acetate, manganese (II) ions) confirmed the significant role of enzymatic transformation. Whereas, biomass sorption was not an important process in the antibiotics elimination. Evaluation of AA against Staphylococcus aureus ATCC 6538 revealed that Leptosphaerulina sp. also eliminated the AA. In addition, the cytotoxicity assay (MTT) on the HepG2 cell line demonstrated that the IP final solutions were non-toxic. Finally, Leptosphaerulina sp. eliminated OXA and its AA from synthetic hospital wastewater at 6 days. All these results evidenced the potential of Leptosphaerulina sp. mycoremediation as a novel environmentally friendly process for the removal of IP from aqueous systems.