Comparative analysis of Chrysoporthe cubensis exoproteomes and their specificity for saccharification of sugarcane bagasse.

The phytopathogenic fungus Chrysoporthe cubensis is a relevant source of lignocellulolytic enzymes. This work aimed to compare the profile of lignocellulose-degrading proteins secreted by C. cubensis grown under semi-solid state fermentation using wheat bran (WB) and sugarcane bagasse (SB). The exoproteomes of the fungus grown in wheat bran (WBE) and sugarcane bagasse (SBE) were qualitative and quantitatively analyzed by liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). Data are available via ProteomeXchange with identifier PXD046075. Label-free proteomic analysis of WBE and SBE showed that the fungus produced a spectrum of carbohydrate-active enzymes (CAZymes) with exclusive characteristics from each extract. While SBE resulted in an enzymatic profile directed towards the depolymerization of cellulose, the enzymes in WBE were more adaptable to the degradation of biomass rich in hemicellulose and other non-lignocellulosic polymers. Saccharification of alkaline pre-treated sugarcane bagasse with SBE promoted glucose release higher than commercial cocktails (8.11 g L-1), while WBE promoted the higher release of xylose (5.71 g L-1). Our results allowed an in-depth knowledge of the complex set of enzymes secreted by C. cubensis responsible for its high lignocellulolytic activity and still provided the identification of promising target proteins for biotechnological applications in the context of biorefinery.

Penicillium Ochrochloron RLS11 Secretome Containing Carbohydrate-Active Enzymes Improves Commercial Enzyme Mixtures During Sugarcane Straw Saccharification.

Filamentous fungi are prolific producers of carbohydrate-active enzymes (CAZymes) and important agents that carry out plant cell wall degradation in natural environments. The number of fungal species is frequently reported in the millions range, with a huge diversity and genetic variability, reflecting on a vast repertoire of CAZymes that these organisms can produce. In this study, we evaluated the ability of previously selected ascomycete and basidiomycete fungi to produce plant cell wall-degrading enzyme (PCWDE) activities and the potential of the culture supernatants to increase the efficiency of the Cellic® CTec2/HTec2 for steam-exploded sugarcane straw saccharification. The culture supernatant of Penicillium ochrochloron RLS11 showed a promising supplementation effect on Cellic® CTec2/HTec2, and we conducted the whole-genome sequencing and proteomic analysis for this fungus. The size of the assembled genome was 38.06 Mbp, and a total of 12,015 protein-coding genes were identified. The repertoire of PCWDE-coding genes was comparatively high among Penicillium spp. and showed an expansion in important cellulases and xylanases families, such as GH3, GH6, GH7, and GH11. The proteomic analysis indicated cellulases that probably enhanced the biomass saccharification performance of the Cellic® CTec2/HTec2, which included enzymes from GH3, GH6, and GH7 families.

Extraction, purification by cation exchange supermacroporous cryogel and physico-chemical characterization of γ-conglutin from lupin seeds (Lupinus albus L.).

This study focused on the extraction, purification, and physicochemical characterization of γ-conglutin, a protein present in lupin seeds with properties of reducing blood glucose levels. Total protein was extracted with an alkaline-saline solvent, followed by isoelectric precipitation. Chromatographic purification of the precipitated fraction was performed using a cation exchange supermacroporous cryogel column. Electrophoresis of the eluted fraction from chromatography presented a single band of ∼48 kDa under non-reducing conditions (two bands of ∼30 and ∼17 kDa, under reducing conditions) confirming the success of the purification protocol. Liquid chromatography-tandem mass spectrometry analysis confirmed the identity of the protein as γ-conglutin. The purified γ-conglutin had an isoelectric point of 7.51, ß-sheets prevailing as a secondary structure, and denaturation temperature close to 68°C. The outcome of this work showed that γ-conglutin was obtained with a high degree of purity. The proposed purification protocol is simple and can be easily scaled up.

First report on the enzymatic profile of Kretzschmaria zonata.

Enzymes from phytopathogenic fungi are desirable for biotechnological applications and a highly virulent phytopathogen shows great appeal for enzymes production. To assess the biotechnological potential of Kretzschmaria zonata, a plant pathogenic fungus, we analyzed its enzymatic profile after growth on six different types of lignocellulosic biomasses. The fungus was able to produce a wide variety of enzymes with superior xylanase activity. The corn cob induced the highest specific activity of xylanase, 56.30 U/mg of protein, as well as other important enzymatic activities such as endoglucanase, 11.20 U/mg of protein; pectinase, 4.52 U/mg of protein; and ß-glucosidase, 2.77 U/mg of protein. The highest release of xylose, 0.88 g/L, was observed after saccharification of 10% of alkaline pretreated sugarcane bagasse by a commercial cocktail supplemented with the crude extract from K. zonata after growth on corn cob. The fungus extract is rich in hemicellulases and accessory enzymes and the result showed synergism between the enzymes present in the commercial mixture and in the K. zonata extract. This is the first report concerning the biotechnological potential of the fungus K. zonata, especially regarding to its ability to produce plant biomass degrading enzymes related to second generation ethanol production.

Secretomic insight into the biomass hydrolysis potential of the phytopathogenic fungus Chrysoporthe cubensis.

The phytopathogenic fungus Chrysoporthe cubensis has a great capacity to produce highly efficient enzymes for the hydrolysis of lignocellulosic biomass. The bioinfosecretome of C. cubensis was identified by computational predictions of secreted proteins combined with protein analysis using 1D-LC-MS/MS. The in silico secretome predicted 562 putative genes capable of encoding secreted proteins, including 273 CAZymes. Proteomics analysis confirmed the existence of 313 proteins, including 137 CAZymes classified as Glycosyl Hydrolases (GH), Polysaccharide Lyases (PL), Carbohydrate Esterases (CE) and Auxiliary Activities enzymes (AA), which indicates the presence of classical and oxidative cellulolytic mechanisms. The enzymes diversity in the extract shows fungal versatility to act in complex biomasses. This study provides an insight into the lignocellulose-degradation mechanisms by C. cubensis and allows the identification of the enzymes that are potentially useful in improving industrial process of bioconversion of lignocellulose. SIGNIFICANCE: Chrysoporthe cubensis is an important deadly canker pathogen of commercially cultivated Eucalyptus species. The effective depolymerisation of the recalcitrant plant cell wall performed by this fungus is closely related to its high potential of lignocellulolytic enzymes secretion. Since the degradation of biomass occurs in nature almost exclusively by enzyme secretion systems, it is reasonable to suggest that the identification of C. cubensis lignocellulolytic enzymes is relevant in contributing to new sustainable alternatives for industrial solutions. As far as we know, this work is the first accurate proteomic evaluation of the enzymes secreted by this species of fungus. The integration of the gel-based proteomic approach, the bioinformatic prediction of the secretome and the analyses of enzymatic activity are powerful tools in the evaluation of biotechnological potential of C. cubensis in producing carbohydrate-active enzymes. In addition, analysis of the C. cubensis secretome grown in wheat bran draws attention to this plant pathogen and its extracellular enzymatic machinery, especially regarding the identification of promising new enzymes for industrial applications. The results from this work allowed for explanation and reinforce previous research that revealed C. cubensis as a strong candidate to produce enzymes to hydrolyse sugarcane bagasse and similar substrates.

Hydrothermal Pretreatment as a Strategy for the Improvement of Sugarcane Bagasse Saccharification by Fungal Enzyme Blend

Abstract Obtaining low cost lignocellulolytic enzymes and efficient biomass pretreatment are key to increase the competitiveness of second-generation ethanol in comparison with fossil fuels. The enzymatic cocktail produced by the Chrysoporthe cubensis fungus as well as the mixture prepared with the cocktails of the Chrysoporthe cubensis and Penicillium pinophilum fungi have already proven to be efficient for hydrolyzing biomass pretreated with alkali. In this study, they were evaluated in saccharification of sugarcane bagasse pretreated with dilute acid or hot water at 121°C using an enzyme loading equal to 8 filter paper units per gram of biomass. The most promising results were obtained from the hydrolysis of biomass pretreated with hot water by the C. cubensis-P. pinophilum enzymes blend. In this condition, the glucose and xylose production were 25.2 g.L-1 and 4.6 g.L-1, respectively, that resulted in the conversion of 68% of glucan and 23% of xylan in only 48 hours. This study shows that the hydrothermal pretreatment is a promising alternative to improve the enzymes performance, produced by the fungi C. cubensis and P. pinophilum, in the sugarcane bagasse hydrolysis without the need of chemical compounds, generally used in the acid and alkali pretreatments. Furthermore, the hydrothermal pretreatment for 60 min allowed all cocktails applied to convert the cellulose efficiently with only 24 h of saccharification, which contributes to the energy savings employed in the process.

Brachiaria brizantha Grass as a Feedstock for Ethanol Production

HIGHLIGHTS Brachiaria brizantha proved to be a promising biomass for ethanol production. Fermentation was not impaired by the inhibitors furfural and hydroxymethylfurfural.

Abstract Different lignocellulosic biomasses are found worldwide and each country has its own important industrial crop that can be converted into high-value products, such as ethanol. Therefore, evaluation of new biomasses to be used in biorefineries is important to decrease the dependence on non-renewable resources and to guarantee sustainable development. This work evaluated Brachiaria brizantha, a grass commonly used as animal forage, and the standard biomass for 2G-ethanol, sugarcane bagasse. The chemical compositions of both biomasses were determined and different times and temperature of acid pretreatment were tested. Morphological analysis via scanning electron microscopy showed more deconstructed fibers after harsher biomass pretreatments. Simultaneous saccharification and fermentation of pretreated Brachiaria brizantha presented higher efficiency than when using sugarcane bagasse as the carbon source. A biomass conversion of 46 % was achieved when Brachiaria brizantha grass was pretreated with 2% sulfuric acid for 60 minutes. Moreover, fermentation was not impaired by the inhibitors furfural and hydroxymethylfurfural. It was concluded that Brachiaria brizantha is a promising biomass for ethanol production.

Synbiotic VSL#3 and yacon-based product modulate the intestinal microbiota and prevent the development of pre-neoplastic lesions in a colorectal carcinogenesis model.

Colorectal cancer is a public health problem, with dysbiosis being one of the risk factors due to its role in intestinal inflammation. Probiotics and synbiotics have been used in order to restore the microbiota balance and to prevent colorectal carcinogenesis. We aimed to investigate the effects of the probiotic VSL#3® alone or in combination with a yacon-based prebiotic concentrate on the microbiota modulation and its influence on colorectal carcinogenesis in an animal model. C57BL/6J mice were divided into three groups: control (control diet), probiotic (control diet + VSL#3®), and synbiotic (yacon diet + VSL#3®). The diets were provided for 13 weeks and, from the third one, all animals were subjected to induction of colorectal cancer precursor lesions. Stool samples were collected to evaluate organic acids, feces pH, ß-glucuronidase activity, and microbiota composition. The colon was used to count pre-neoplastic lesions and to determine the cytokines. The microbiota composition was influenced by the use of probiotic and synbiotic. Modifications were also observed in the abundance of bacterial genera with respect to the control group, which confirms the interference of carcinogenesis in the microbiota. Pre-neoplastic lesions were reduced by the use of the synbiotic, but not with the probiotic. The protection provided by the synbiotic can be attributed to the modulation of the intestinal inflammatory response, to the inhibition of a pro-carcinogenic enzyme, and to the production of organic acids. The modulation of the composition and activity of the microbiota contributed to beneficial changes in the intestinal microenvironment, which led to a reduction in carcinogenesis. KEY POINTS: • Synbiotic reduces the incidence of colorectal cancer precursor lesions. • Synbiotic modulates the composition and activity of intestinal microbiota. • Synbiotic increases the abundance of butyrate-producing bacteria.

Genome-Scale Characterization of Fungal Phytases and a Comparative Study Between Beta-Propeller Phytases and Histidine Acid Phosphatases.

This work intended to prospect new phytase-producing organisms. In silico genomic analyses allowed the selection of twelve potential phytase-producing fungi. Based on gene sequence, it was possible to identify four well-defined groups of phytate-degrading enzymes: esterase-like, ß-propeller phytases (ßPP), phosphoglycerate mutase-like, and phytases of the histidine acid phosphatases (HAP) family. Analysis of the predicted genes encoding phytases belonging to the HAP family and ßPP phytases and in silico characterization of these enzymes indicated divergence among the catalytic activities. Predicted fungal ßPP phytases exhibited higher molecular mass (around 77 kDa) probably due to the epidermal growth factor-like domain. Twelve sequences of phytases contained signal peptides, of which seven were classified as HAP and five as ßPP phytases, while ten sequences were predicted as phytases secreted by non-classical pathways. These fungi were grown in liquid or semi-solid medium, and the fungal enzymatic extracts were evaluated for their ability to hydrolyze sodium phytate at 50 °C and pH ranging from 2.0 to 9.0. Seven fungi were identified as phytase producers based on phosphate release under enzyme assay conditions. Results obtained from in silico analyses combining experimental enzymatic activities suggest that some selected fungi could secrete ßPP phytases and HAP phytases.

Production of Fungal Phytases from Agroindustrial Byproducts for Pig Diets.

The application of phytases for animal feed in developing countries is limited due to the high cost of these enzymes, determined by the importation fees and the expensive substrates used for their production. In this work, we have used agroindustrial byproducts for the production of extracts containing phytases, which were accessed for their stability focusing on the conditions found in the gastrointestinal tract of pigs. The fungus Acremonim zeae presented higher phytase production in medium containing cornmeal, while the yeast Kluyveromyces marxianus produced 10-fold more phytase when cultivated on rice bran. Process optimization increased the difference in productivity to more than 300 fold. The phytase from A. zeae was thermostable, with higher activity at neutral pH and 50 °C, but was inhibited at pH 2.5 and by various ions. The phytase activity in the K. marxianus extract was stable at a wide range of conditions, which indicates the presence of at least two enzymes. As far as we know, this manuscript describes for the first time the phytase production and the characteristics of the extracts produced by both these microbial species. These enzymes could be produced at low cost and have potential to replace enzymes currently imported for this purpose.

Engineered GH11 xylanases from Orpinomyces sp. PC-2 improve techno-functional properties of bread dough.

BACKGROUND: Endo-1,4-ß-xylanases have marked hydrolytic activity towards arabinoxylans. Xylanases (xynA) produced by the anaerobic fungus Orpinomyces sp. strain PC-2 have been shown to be superior in specific activity, which strongly suggests their applicability in the bakery industry for the processing of whole-wheat flour containing xylans. In the present study, two xylanases from this source, the small wild-type xylanase SWT and the small mutant xylanase SM2 (V108A, A199T), were expressed in Escherichia coli, purified, characterized, tested for their ability to hydrolyze whole-wheat flour and applied in dough processing. RESULTS: Both purified SM2 and SWT showed high specific activity against oat spelt xylan and wheat arabinoxylan, exhibiting maximum activity at pH 3-7 and 60 °C. SM2 was more thermostable than SWT, which suggests that the mutations enhanced its stability. Both SWT and SM2 were able to hydrolyze whole-wheat flour, and evaluation of their applicability in dough processing by the sponge method indicated that use of these enzymes increased dough volume by 60% and reduced texture hardness by more than 50%, while gumminess and chewiness were reduced by 40%. CONCLUSION: The recombinant xylanases showed potential for application in bakery processing and can improve techno-functional properties in sponges. © 2018 Society of Chemical Industry.

Anti-Hypertensive Peptides Derived from Caseins: Mechanism of Physiological Action, Production Bioprocesses, and Challenges for Food Applications.

This review is focused on the state-of-art of peptides with inhibitory activity towards angiotensin I-converting enzyme (ACE) - thus, with anti-hypertensive potential - derived from enzymatic hydrolysis of caseins. Firstly, molecular characteristics of caseins relevant to a better understanding of this subject were concisely commented. Next, a brief description of the pathophysiology of hypertension was explained, focusing on the ACE role in regulation of blood pressure in human body. Then, casein-derived peptides with ACE inhibitory capacity were specifically addressed. The main in vitro and in vivo bioassays often reported in literature to assess the anti-hypertensive potential of peptides were presented, illustrated with recently published studies, and discussed in terms of advantages and limitations of both approaches. Characteristics related to amino acid composition and sequence of peptides with high ACE-inhibitory potential were also commented. Process parameters of enzymatic hydrolysis (types and origins of casein substrates, types of enzymes, pH, temperature, and times of reactions) were discussed. Patents dealing with casein-derived anti-hypertensive peptides were examined not only in terms of amino acid sequences, but also regarding their novelty claims in hydrolysis process parameters. Finally, some trends, challenges, and opportunities inferred from this literature analysis were commented, emphasizing the importance of this research topic in food products development.

Impact of the removal of N-terminal non-structured amino acids on activity and stability of xylanases from Orpinomyces sp. PC-2.

Xylanases catalyze the random hydrolysis of xylan backbone from plant biomass and thus, they have application in the production of biofuels, Kraft pulps biobleaching and feed industry. Here, xylanases derived from Orpinomyces sp. PC-2 were engineered guided by molecular dynamics methods to obtain more thermostable enzymes. Based on these models, 27 amino acid residues from the N-terminal were predicted to reduce protein stability and the impact of this removal was validated to two enzyme constructs: small xylanase Wild-Type (SWT) obtained from Wild-Type xylanase (WT) and small xylanase Mutant (SM2) generated from M2 mutant xylanase (V135A, A226T). The tail removal promoted increase in specific activity of purified SWT and SM2, which achieved 5,801.7 and 5,106.8Umg-1 of protein, respectively, while the WT activity was 444.1Umg-1 of protein. WT, SWT and SM2 showed half-life values at 50°C of 0.8, 2.3 and 29.5h, respectively. Overall, in view of the results, we propose that the presence of non-structured amino acid in the N-terminal leads to destabilization of the xylanases and may promote less access of the substrate to the active site. Therefore, its removal may promote increased stability and enzymatic activity, interesting properties that make them suitable for biotechnological applications.

A Chrysoporthe cubensis enzyme cocktail produced from a low-cost carbon source with high biomass hydrolysis efficiency.

Low cost and high efficiency cellulolytic cocktails can consolidate lignocellulosic ethanol technologies. Sugarcane bagasse (SCB) is a low cost agro-industrial residue, and its use as a carbon source can reduce the costs of fungi cultivation for enzyme production. Chrysoporthe cubensis grown under solid state fermentation (SSF) with wheat bran has potential to produce efficient enzymatic extracts for SCB saccharification. This fungus was grown under submersed fermentation (SmF) and SSF with in natura SCB, pretreated with acid or alkali and with others carbon sources. In natura SCB induced the highest carboxymethylcellulase (CMCase), xylanase, ß-xylosidase, α-galactosidase and mannanase activities by C. cubensis under SSF. In natura and washed SCB, inducers of enzyme production under SSF, did not induce high cellulases and hemicellulases production by C. cubensis in SmF. The C. cubensis enzymatic extract produced under SSF with in natura SCB as a carbon source was more efficient for lignocelulolic biomass hydrolysis than extracts produced under SSF with wheat bran and commercial cellulolytic extract. Chrysoporthe cubensis showed high potential for cellulases and hemicellulases production, especially when grown under SSF with in natura SCB as carbon source.

Purification and Characterization of Xylanases from the Fungus Chrysoporthe cubensis for Production of Xylooligosaccharides and Fermentable Sugars.

Xylanases from the pathogen fungus Chrysoporthe cubensis were produced under solid state fermentation (SSF) using wheat bran as carbon source. The enzymatic extracts were submitted to ion exchange (Q Sepharose) and gel filtration chromatography methods (Sephadex S-200) for purification. The xylanases were divided into three groups: P1 showed better performance at 60 °C and pH 4.0, P2 at 55 °C and pH 3.0, and P3 at 80 °C and pH 3.0. Oat spelt xylan was the best substrate hydrolyzed by P1 and P3, while beechwood xylan was better degraded by P2. Carboxymethyl cellulose (CMC) and p-nitrophenyl-ß-D-xylopyranoside (p-NPßXyl) were not hydrolyzed by any of the xylanases. The K M' or K M values, using oat spelt xylan as substrate, were 2.65 mg/mL for P1, 1.81 mg/mL for P2, and 1.18 mg/mL for P3. Xylobiose and xylotriose were the main xylooligosaccharides of oat spelt xylan degradation, indicating that the xylanases act as endo-ß-1,4-xylanases. Xylanases also proved to be efficient for hydrolysis of sugarcane bagasse when used as supplement of a commercial cocktail due to the increase of the reducing sugar release.

The influence of pretreatment methods on saccharification of sugarcane bagasse by an enzyme extract from Chrysoporthe cubensis and commercial cocktails: A comparative study.

Biomass enzymatic hydrolysis depends on the pretreatment methods employed, the composition of initial feedstock and the enzyme cocktail used to release sugars for subsequent fermentation into ethanol. In this study, sugarcane bagasse was pretreated with 1% H2SO4 and 1% NaOH and the biomass saccharification was performed with 8% solids loading using 10 FPase units/g of bagasse of the enzymatic extract from Chrysoporthe cubensis and three commercial cocktails for a comparative study. Overall, the best glucose and xylose release was obtained from alkaline pretreated sugarcane bagasse. The C. cubensis extract promoted higher release of glucose (5.32 g/L) and xylose (9.00 g/L) than the commercial mixtures. Moreover, the C. cubensis extract presented high specific enzyme activities when compared to commercial cocktails mainly concerning to endoglucanase (331.84 U/mg of protein), ß-glucosidase (29.48 U/mg of protein), ß-xylosidase (2.95 U/mg of protein), pectinase (127.46 U/mg of protein) and laccase (2.49 U/mg of protein).

Increased enzymatic hydrolysis of sugarcane bagasse from enzyme recycling.

BACKGROUND: Development of efficient methods for production of renewable fuels from lignocellulosic biomass is necessary to maximize yields and reduce operating costs. One of the main challenges to industrial application of the lignocellulosic conversion process is the high costs of cellulolytic enzymes. Recycling of enzymes may present a potential solution to alleviate this problem. In the present study enzymes associated with the insoluble fraction were recycled after enzymatic hydrolysis of pretreated sugarcane bagasse, utilizing different processing conditions, enzyme loadings, and solid loadings. RESULTS: It was found that the enzyme blend from Chrysoporthe cubensis and Penicillium pinophilum was efficient for enzymatic hydrolysis and that a significant portion of enzyme activity could be recovered upon recycling of the insoluble fraction. Enzyme productivity values (g glucose/mg enzyme protein) over all recycle periods were 2.4 and 3.7 for application of 15 and 30 FPU/g of glucan, representing an increase in excess of ten times that obtained in a batch process with the same enzyme blend and an even greater increase compared to commercial cellulase enzymes. CONCLUSIONS: Contrary to what may be expected, increasing lignin concentrations throughout the recycle period did not negatively influence hydrolysis efficiency, but conversion efficiencies continuously improved. Recycling of the entire insoluble solids fraction was sufficient for recycling of adhered enzymes together with biomass, indicative of an effective method to increase enzyme productivity.

Synergistic effect of Aspergillus niger and Trichoderma reesei enzyme sets on the saccharification of wheat straw and sugarcane bagasse.

Plant-degrading enzymes can be produced by fungi on abundantly available low-cost plant biomass. However, enzymes sets after growth on complex substrates need to be better understood, especially with emphasis on differences between fungal species and the influence of inhibitory compounds in plant substrates, such as monosaccharides. In this study, Aspergillus niger and Trichoderma reesei were evaluated for the production of enzyme sets after growth on two "second generation" substrates: wheat straw (WS) and sugarcane bagasse (SCB). A. niger and T. reesei produced different sets of (hemi-)cellulolytic enzymes after growth on WS and SCB. This was reflected in an overall strong synergistic effect in releasing sugars during saccharification using A. niger and T. reesei enzyme sets. T. reesei produced less hydrolytic enzymes after growth on non-washed SCB. The sensitivity to non-washed plant substrates was not reduced by using CreA/Cre1 mutants of T. reesei and A. niger with a defective carbon catabolite repression. The importance of removing monosaccharides for producing enzymes was further underlined by the decrease in hydrolytic activities with increased glucose concentrations in WS media. This study showed the importance of removing monosaccharides from the enzyme production media and combining T. reesei and A. niger enzyme sets to improve plant biomass saccharification.

Production and application of an enzyme blend from Chrysoporthe cubensis and Penicillium pinophilum with potential for hydrolysis of sugarcane bagasse.

Blending of the enzyme extracts produced by different fungi can result in favorable synergetic enhancement of the enzyme blend with regards to the main cellulase activities, as well as the inclusion of accessory enzymes that may not be as abundant in enzyme extracts produced by predominantly cellulase producing fungi. The Chrysoporthe cubensis:Penicillium pinophilum 50:50 (v/v) blend produced herein presented good synergy, especially for FPase and endoglucanase activities which were 76% and 48% greater than theoretical, respectively. This enzyme blend was applied to sugarcane bagasse previously submitted to a simple alkali pretreatment. Glucan hydrolysis efficiency reached an excess of 60% and xylan conversion exceeded 90%. Increasing the hydrolysis temperature from 45 to 50°C also resulted in a 16-20% increase in conversion of both glucan and xylan fractions. The blended enzyme extract obtained therefore showed great potential for application in the lignocellulose hydrolysis process.

Chrysoporthe cubensis: a new source of cellulases and hemicellulases to application in biomass saccharification processes.

The plant pathogenic fungus Chrysoporthe cubensis was cultivated under solid state employing different substrates and the highest endoglucanase (33.84Ug(-1)), FPase (2.52Ug(-1)), ß-glucosidase (21.55Ug(-1)) and xylanase (362.38Ug(-1)) activities were obtained using wheat bran as carbon source. Cellulases and xylanase produced by C. cubensis showed maximal hydrolysis rate at pH 4.0 and in a temperature range of 50-60°C. All enzymatic activities were highly stable at 40 and 50°C through 48h of pre-incubation. Saccharification of alkaline pretreated sugarcane bagasse by crude enzyme extract from C. cubensis resulted in release of 320.8mg/g and 288.7mg/g of glucose and xylose, respectively. On another hand, a similar assay employing commercial cellulase preparation resulted in release of 250.6mg/g and 62.1mg/g of glucose and xylose, respectively. Cellulolytic extract from C. cubensis showed a great potential to be used in biomass saccharification processes.