Evaluation of Chlorella (Chlorophyta) as Source of Fermentable Sugars via Cell Wall Enzymatic Hydrolysis

The cell wall of Chlorella is composed of up to 80% carbohydrates including cellulose. In this study, Chlorella homosphaera and Chlorella zofingiensis were evaluated as source of fermentable sugars via their cell wall enzymatic degradation. The algae were cultivated in inorganic medium, collected at the stationary growth phase and centrifuged. The cell pellet was suspended in citrate buffer, pH 4.8 and subjected to 24 hours hydrolysis at 50°C using a cellulases, xylanases, and amylases blend. The measurement of glucose and reducing sugars concentration in the reaction mixture supernatant, on a dry biomass base, showed hydrolysis yields of 2.9% and 5.03% glucose and 4.8% and 8.6% reducing sugars, for C. homosphaera and C. zofingiensis, respectively. However if cells were washed with chilled ethanol, cold dried, and grounded the biomass hydrolysis yields increased to 23.3% and 18.4% glucose and 24.5% and 19.3% reducing sugars for C. homosphaera and C. zofingiensis, respectively

Processo de produção de moléculas orgânicas a partir da biomassa

DepositadaProcesso de produção de moléculas orgânicas a partir da biomassa que compreende as etapas de: pré-tratamento contínuo de uma biomassa com líquidos iônicos em um misturador; hidrólise de biomassa pré-tratada; e separação do xarope de açúcares do sólido residual não hidrolisado rico em lignina; e, fermentação do xarope de açúcares, onde uma etapa ocorre de forma contínua em um misturador, no qual uma razão de elevada de biomassa; líquido iônico é misturado possibilitando a atuação efetiva do líquido iônico na biomassa

A thermotolerant xylan-degrading enzyme is produced by Streptomyces malaysiensis AMT-3 using by-products from the food industry

ABSTRACT: This study evaluated the production of endoxylanases by Streptomyces malaysiensis AMT-3 in submerged fermentation using by-products of the food industry at 28ºC. In shake-flasks experiments, the highest endoxylanase activity of 45.8 U.mL-1 was observed within 6 days in a medium containing (w/v) 2.5% wheat bran and 1.2% corn steep liquor. The same culture conditions were used to evaluate the enzyme production in a 2 L stirred tank reactor under different agitation (300, 450 and 600 rev.min-1) and aeration (30 and 60 L.h-1) conditions. The use of 450 rev.min-1 coupled to an aeration of 90 L.h-1 resulted on 81.3 U.mL-1 endoxylanase activity within 5 days. The effect of temperature and pH on endoxylanase activity and stability showed the highest activity at 60 ºC and pH 6.0. Zymography showed the presence of three xylanolytic bands with molecular masses of 690, 180 and 142 kDa. The results showed that the thermotolerant actinobacterial endoxylanase can be produced in high titers using by-product of the food industry.info:eu-repo/semantics/publishedVersio

Production of thermophilic and acidophilic endoglucanases by mutant Trichoderma atroviride 102C1 using agro-industrial by-products

Many traditional mutagenic strategies have been used to improve cellulase production by microorganisms, especially fungi species. Trichoderma species are among cellulolytic fungi, those that have been most extensively studied, due to their efficient production of these enzymes. In the present study, N-methyl-N´-nitro-N nitrosoguanidine (NTG) was used as mutagenic agent to obtain cellulolytic mutant from wild strain T. atroviride 676. After mutagenic procedures, two strains (102C1 and 104C2) were selected as promising cellulase-producing mutant. The effect of the carbon (sugarcane bagasse: SCB) and nitrogen (corn steep liquor: CSL) sources on endoglucanase production by the mutants 102C1 and 104C2 was studied using submerged cultivations at 28°C. Different concentrations of SCB and CSL were used and nine different media were generated. Mutant 102C1 showed the best results when using 2.5% SCB and 0.7% CSL. A central composite rotational design (CCRD) was performed to estimate optimal conditions of pH and temperature for endoglucanase activity of strain 102C1, which were pH 3.6 and temperature 66°C. The characterization of this acidophilic and thermophilic endoglucanase activity produced by the mutant strain 102C1 allows its use in biotechnological applications, particularly in the hydrolysis of agro industrial residues, such as SCB, for bioethanol production.Keywords: Trichoderma atroviride, CMCase, nitrosoguanidine, sugarcane bagasse, corn steep liquo

Asparaginase II-GFP fusion as a tool for studying the secretion of the enzyme under nitrogen starvation Fusão asparaginase II-GFP como ferramenta para estudo da via secretora de enzima sobre depleção por nitrogênio

Production of asparaginase II of Saccharomyces cerevisiae is regulated by nitrogen and can be used as a model system for studying other secreted proteins in yeast. Green fluorescent protein (GFP) from Aequorea victoria was fused to the carboxy-terminus of the enzyme by genomic integration to the locus ASP3 of S. cerevisiae. We determined asparaginase II activity, mRNA ASP3, mRNA ASP3-GFP and GFP fluorescence. Nitrogen starvation in cells carrying the chimera ASP3-GFP caused an increase in fluorescence and in the expression of ASP3. We have shown that cells producing the chimera Asp3-GFPp displayed the same response to nitrogen starvation as control cells. We demonstrated that Asp3-GFPp can be used for studying asparaginase II secretion under nitrogen starvation in vivo.<br>A produção de asparaginase II de Saccharomyces cerevisiae é regulada por nitrogênio e pode ser utilizada como um sistema modelo para estudar outras proteínas secretadas, em leveduras. A proteína "green fluorescent protein" (GFP) de Aequorea victoria foi fusionada à porção carboxi-terminal de Asp3p por integração genômica da sequência de GFP ao locus ASP3. Determinaram-se os níveis de atividade de asparaginase II, mRNA ASP3, mRNA ASP3-GFP e de fluorescência para GFP. A depleção para nitrogênio, em células portadoras do gene quimérico ASP3-GFP, fez aumentar a fluorescência, assim como a expressão de ASP3. Demonstramos que Asp3-GFPp pode ser utilizada para estudar a secreção de asparaginase II em células submetidas à privação de nitrogênio in vivo

Production of cellulases and β-glucosidases by Trichoderma reesei Rut C30 using steam-pretreated sugarcane bagasse : an integrated approach for onsite enzyme production

The high cost of commercial cellulases still hampers the economic competitiveness of the production of fuels and chemicals from lignocellulosic biomasses. This cost may be decreased by the on-site production of cellulases with the integrated use of the lignocellulosic biomass as carbon source. This integrated approach was evaluated in the present study whereby steam-pretreated sugarcane bagasse (SPSB) was used as carbon source for the production of cellulases by Trichoderma reesei Rut C30. The use of SPSB resulted in an enzyme preparation with a cellulase activity of 1.93 FPU/mL. Additionally, a significant beta-glucosidase activity, of 0.37 BGU/mL, was achieved in buffered media, which is not a common feature of T. reesei culture supernatants, indicating the importance of pH control during enzyme production. The hydrolysis of SPSB with the laboratory-made mixture resulted in a glucose yield of 80%, which was equivalent to those observed for control experiments using commercial enzymes. It was shown that SPSB is a promising carbon source for the production of cellulases and beta-glucosidases by T. reesei Rut C30 and that the enzyme preparation obtained is effective for the hydrolysis of SPSB without the need for beta-glucosidases supplementation, supporting the on-site integrated approach to decrease the cost of the enzymatic hydrolysis of lignocellulosic biomass

On-site integrated production of cellulases and β-glucosidases by Trichoderma reesei Rut C30 using steam-pretreated sugarcane bagasse

The high cost of commercial cellulases still hampers the economic competitiveness of the production of fuels and chemicals from lignocellulosic biomasses. This cost may be decreased by the on-site production of cellulases with the integrated use of the lignocellulosic biomass as carbon source. This integrated approach was evaluated in the present study whereby steam-pretreated sugarcane bagasse (SPSB) was used as carbon source for the production of cellulases by Trichoderma reesei Rut C30 and the produced enzymes were subsequently used for SPSB hydrolysis. An enzyme preparation with a high cellulase activity, of 1.93 FPU/mL, was obtained, and a significant β-glucosidase activity was achieved in buffered media, indicating the importance of pH control during enzyme production. The hydrolysis of SPSB with the laboratory-made mixture resulted in a glucose yield of 80%, which was equivalent to those observed for control experiments using commercial enzymes. Even though the supplementation of this mixture with external β-glucosidase from Aspergillus awamori was found to increase the initial hydrolysis rates, it had no impact on the final hydrolysis yield. It was shown that SPSB is a promising carbon source for the production of cellulases and β-glucosidases by T. reesei Rut C30 and that the enzyme preparation obtained is effective for the hydrolysis of SPSB, supporting the on-site integrated approach to decrease the cost of the enzymatic hydrolysis of lignocellulosic biomass