The successful strategy to produce cellulolytic enzymes includes both microorganism selection and improved fermentation process conditions. This work describes the isolation, screening and selection ...of biomass-degrading fungi species from the Amazon forest and analyzes the enzymatic complex produced by a selected strain of Aspergillus fumigatus cultivated using different agro-industrial residues (wheat bran, sugarcane bagasse, soybean bran, and orange peel) as substrate in solid state fermentation (SSF). The profile of endoglucanase (CMCase), FPase, β-glucosidase and xylanase enzymatic activities obtained during 120 h of cultivation is presented. Enzyme activities up 160.1 IU g−1 for CMCase, 5.0 FPU g−1 for FPAse, 105.82 IU g−1 for β-glucosidase and 1055.62 IU g−1 for xylanase were achieved. The enzymatic extract with higher CMCase activity was used to run a zymogram analysis that showed 3 bands of endoglucanase activity. Characterization studies of this extract showed that the CMCase was most active at either 65 °C or pH 3–3.5, indicating that this microorganism produces a thermophilic and acid endoglucanase. These data demonstrate that the fungal isolates from the Amazon forest are a potential source of cellulases and xylanases, providing support to further studies related to the use of these microorganisms to obtain the enzymes needed for biomass conversion.
► Cellulase production is a critical step in the economics of second generation ethanol. ► Screening of microorganism can contribute to finding new cellulolytic enzymes. ► Amazon forest fungi were able to efficiently produce cellulases and xylanases.
Filamentous fungi are attractive hosts for heterologous protein expression due to their capacity to secrete large amounts of enzymes into the extracellular medium. Xyloglucanases, which specifically ...hydrolyze xyloglucan, have been recently applied in lignocellulosic biomass degradation and conversion in many other industrial processes. In this context, this work aimed to clone, express, and determine the functional properties of a recombinant xyloglucanase (AtXEG12) from
Aspergillus terreus
, and also its solid-state (SSF) and submerged (SmF) fermentation in bioreactors. The purified AtXEG12 showed optimum pH and temperature of 5.5 and 65 °C, respectively, demonstrating to be 90 % stable after 24 h of incubation at 50 °C. AtXEG12 activity increased in the presence of 2-mercaptoethanol (65 %) and Zn
+2
(45 %), while Cu
+2
and Ag
+
ions drastically decreased its activity. A substrate assay showed, for the first time for this enzyme’s family, xylanase activity. The enzyme exhibited high specificity for tamarind xyloglucan (
K
M
1.2 mg mL
−1
) and
V
max
of 17.4 μmol min
−1
mg
−1
of protein. The capillary zone electrophoresis analysis revealed that AtXEG12 is an endo-xyloglucanase. The heterologous xyloglucanase secretion was greater than the production by wild-type
A. terreus
cultivated in SmF. On the other hand, AtXEG12 activity reached by SSF was sevenfold higher than values achieved by SmF, showing that the expression of recombinant enzymes can be significantly improved by cultivation under SSF.
► The Amazon rainforest Trichoderma harzianum is a potential candidate for saccharification of plant biomass. ► Important roles for pectinase and α-l-arabinofuranosidase in the enzymatic synergies. ► ...Sugarcane bagasse hydrolysis is more effective and competitive for second generation ethanol.
Supplementation of cellulase cocktails with accessory enzymes can contribute to a higher hydrolytic capacity in releasing fermentable sugars from plant biomass. This study investigated which enzymes were complementary to the enzyme set of Trichoderma harzianum in the degradation of sugarcane bagasse. Specific activities of T. harzianum extract on different substrates were compared with the extracts of Penicillium echinulatum and Trichoderma reesei, and two commercial cellulase preparations. Complementary analysis of the secretome of T. harzianum was also used to identify which enzymes were produced during growth on pretreated sugarcane bagasse. These analyses enabled the selection of the enzymes pectinase and α-l-arabinofuranosidase (AF) to be further investigated as supplements to the T. harzianum extract. The effect of enzyme supplementation on the efficiency of sugarcane bagasse saccharification was evaluated using response surface methodology. The supplementation of T. harzianum enzymatic extract with pectinase and AF increased the efficiency of hydrolysis by up to 116%.
The enzymatic cocktail of cellulases is one of the most costly inputs affecting the economic viability of the biochemical route for biomass conversion into biofuels and other chemicals. Here, the ...influence of liquid hot water, dilute acid, alkali, and combined acid/alkali pretreatments on sugarcane bagasse (SCB) used for cellulase production was investigated by means of spectroscopic and imaging techniques. Chemical composition and structural characteristics, such as crystallinity (determined by X-ray diffraction), functional groups (Fourier transform infrared spectroscopy), and microstructure (scanning electron microscopy), were used to correlate SCB pretreatments with enzymatic biosynthesis by a strain of the filamentous fungus Aspergillus niger under solid-state fermentation. The combined acid/alkali pretreatment resulted in a SCB with higher cellulose content (86.7 %). However, the high crystallinity (74 %) of the resulting biomass was detrimental to microbial uptake and enzyme production. SCB pretreated with liquid hot water yielded the highest filter paper cellulase (FPase), carboxymethyl cellulase (CMCase), and xylanase activities (0.4, 14.9, and 26.1 U g⁻¹, respectively). The results showed that a suitable pretreatment for SCB to be used as a substrate for cellulase production should avoid severe conditions in order to preserve amorphous cellulose and to enhance the physical properties that assist microbial access.
► Glycosyl hydrolases (GH) play a vital role in the conversion of biomass into biofuels. ► Experimental mixture design is a powerful tool in bioprocess development. ► Growth conditions can ...significantly influence GH biosynthesis. ► The Amazon rainforest T. harzianum strain is a useful candidate for GH production.
This work investigates the glycosyl hydrolase (GH) profile of a new Trichoderma harzianum strain cultivated under controlled bioreactor submerged fermentation. The influence of different medium components (delignified steam-exploded sugarcane bagasse, sucrose, and soybean flour) on GH biosynthesis was assessed using experimental mixture design (EMD). Additionally, the effect of increased component concentrations in culture media selected from the EMD was studied. It was found that that a mixed culture medium could significantly maximize GH biosynthesis rate, especially for xylanase enzymes which achieved a 2-fold increment. Overall, it was demonstrated that T. harzianumP49P11 enzymes have a great potential to be used in the deconstruction of biomass.
Asbestos cement materials used in the past by the building construction sector still poses environmental problems, requiring proper treatment and storage. Here we investigated an alternative ...microbiological-induced treatment of raw chrysotile and asbestos cement materials. Based on the knowledge that some microorganisms, such as Aspergillus niger (fungus) and Acidithiobacillus thiooxidans (bacteria) are potential organic or inorganic acid producers, we evaluated the dissolution of the chrysotile structure, which is the toxic component in these materials, under submerged and solid-state cultivation conditions. The Mg, and Ca release percentage evaluated and SEM/EDX, XRD, and FTIR characterization data have shown a high potential of both microorganisms studied to total chrysotile degradation (up to 100%, based on Mg content), using either the microbial filtrate, submerged and solid-state cultivation. The present study is a potential biological treatment of asbestos cement materials that can also be applied in situ in areas contaminated with asbestos, which is an alternative not applied to energy-intensive methods such as mechanochemical or thermal treatment.
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•Asbestos cement wastes bioremediation;•Biological routes to detoxify asbestos cement wastes;•Mg2+ and Ca2+ bioleaching;•Chrysotile biological mineralization;•Environmentally-friendly strategy to detoxify asbestos cement wastes.
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•Sugarcane bagasse and soybean bran favored oxalic acid production by A. niger.•Addition of methanol significantly increased oxalic acid production by A. niger.•PRs can be efficiently ...solubilized under optimized oxalic acid production via SSF.•Solubilization of PRs by SSF can promote additional P release from substrates.
This study explores the enhancement of phosphate rock (PR) solubilization through solid-state fermentation (SSF) by optimizing oxalic acid production using Aspergillus niger. Key process parameters, including the use of agro-industrial by-products (sugarcane bagasse (SCB), wheat bran (WB), soybean bran (SB)), pH levels, sucrose supplementation, and methanol addition, were systematically evaluated through sequential experimental designs. The results identified SCB and SB in a 1:1 ratio as the most effective substrate. Remarkably, the inclusion of methanol (7 %) and sucrose (0.5 %) resulted in a 3-fold increase in oxalic acid production. Under these optimized conditions, significant phosphorus solubilization of Bayóvar, Itafós, and Registro PRs was achieved, with Bayóvar rock releasing up to 12.1 g/kgds of soluble P (63.8 % efficiency). Additionally, the SSF process effectively released organic phosphorus from the agro-industrial substrates. These findings hold promise for advancing the bio-based economy and developing future industrial biofertilizers.
A conversão da biomassa vegetal proveniente de resíduos agroindustriais e florestais em biocombustíveis e bioprodutos, dentro do conceito de biorrefinarias, é de grande interesse, principalmente para ...o Brasil, onde a agroenergia possui um enorme potencial de desenvolvimento. Entretanto, para garantir a viabilidade do processo de conversão, é fundamental reduzir o custo das enzimas utilizadas na etapa de hidrólise. Para isso, deve-se dispor da peça chave deste processo, que é o microrganismo. Nesse contexto, o objetivo deste trabalho foi avaliar fungos isolados da região Amazônica em relação ao potencial de produção das enzimas celulases e xilanases. De um total de 40 isolados cultivados por fermentação em estado sólido (FES), durante 10 dias, os fungos que se destacaram quanto à produção de endoglucanase (351,79Ug -1 em 120h) e β-glicosidase (62,31Ug -1 em 72h) foi o P47C3 (A. niger), e na produção de xilanase (1076,94Ug -1 em 72h) e FPase (2,46Ug -1 em 120h) foram o P6B2 (A. oryzae) e o P40B3, respectivamente. Os resultados obtidos demonstram o enorme potencial de aplicação das enzimas produzidas pelos fungos isolados da Amazônia, contribuindo, assim, para gerar os avanços tecnológicos necessários para o aumento da eficiência do uso da biomassa vegetal como fonte de energia renovável
O objetivo deste trabalho foi avaliar a produção enzimática de celulases pelo fungo filamentoso Aspergillus niger por fermentação em estado sólido de diferentes substratos. Foram avaliados os ...substratos sólidos bagaço de cana-de-açúcar, farelo de soja, farelo de trigo e misturas entre os substratos. Em substrato com 90% de bagaço e 10% de farelo de soja, avaliaram-se os efeitos do conteúdo de umidade (60, 70 e 80%, base úmida) e da suplementação com os meios indutores da atividade enzimática: sacarose, Mandels & Weber básico, Mandels & Weber modificado, com acréscimo de carboximetilcelulose, e Czapeck Dox. As maiores atividades de celulase total e endoglucanase, em farelo de trigo, foram obtidas após 72 horas de cultivo: 0,4 e 21,0 UI g-1, respectivamente. Observou-se expressivo aumento nas atividades enzimáticas na medida em que se aumentou a proporção de farelos no substrato, em comparação à fermentação com bagaço de cana apenas. O conteúdo de umidade de 50% foi insuficiente para conseguir completa hidratação do bagaço de cana, e a umidade ideal varia de acordo com o meio utilizado para suplementação e encontra-se entre 70 e 80%. O meio de Mandels & Weber modificado apresenta o melhor resultado como indutor da atividade enzimática.
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