BackgroundThis study is the first to investigate the Brazilian Amazonian Forest to identify new D-xylose-fermenting yeasts that might potentially be used in the production of ethanol from sugarcane bagasse hemicellulosic hydrolysates.Methodology/Principal FindingsA total of 224 yeast strains were isolated from rotting wood samples collected in two Amazonian forest reserve sites. These samples were cultured in yeast nitrogen base (YNB)-D-xylose or YNB-xylan media. Candida tropicalis, Asterotremella humicola, Candida boidinii and Debaryomyces hansenii were the most frequently isolated yeasts. Among D-xylose-fermenting yeasts, six strains of Spathaspora passalidarum, two of Scheffersomyces stipitis, and representatives of five new species were identified. The new species included Candida amazonensis of the Scheffersomyces clade and Spathaspora sp. 1, Spathaspora sp. 2, Spathaspora sp. 3, and Candida sp. 1 of the Spathaspora clade. In fermentation assays using D-xylose (50 g/L) culture medium, S. passalidarum strains showed the highest ethanol yields (0.31 g/g to 0.37 g/g) and productivities (0.62 g/L·h to 0.75 g/L·h). Candida amazonensis exhibited a virtually complete D-xylose consumption and the highest xylitol yields (0.55 g/g to 0.59 g/g), with concentrations up to 25.2 g/L. The new Spathaspora species produced ethanol and/or xylitol in different concentrations as the main fermentation products. In sugarcane bagasse hemicellulosic fermentation assays, S. stipitis UFMG-XMD-15.2 generated the highest ethanol yield (0.34 g/g) and productivity (0.2 g/L·h), while the new species Spathaspora sp. 1 UFMG-XMD-16.2 and Spathaspora sp. 2 UFMG-XMD-23.2 were very good xylitol producers.Conclusions/SignificanceThis study demonstrates the promise of using new D-xylose-fermenting yeast strains from the Brazilian Amazonian Forest for ethanol or xylitol production from sugarcane bagasse hemicellulosic hydrolysates.

A B S T R A C TEndophytic fungi are widely studied as producers of secondary metabolites of biotechnological interest. In recent years, the interest in these fungi as new sources of enzymes, especially hydrolytic, has increased. In the present study, 14 strains of endophytic fungi not yet explored as enzymes sources were randomly chosen and prospected for cellulases and xylanases production by solid-state fermentation. Initially, fungi were cultivated in a mixture (1:1 w/w) of sugarcane bagasse and wheat bran for 7 days, at 28°C. In this initial screening, 4 fungi excelled in endoglucanase activity (U/g): Cladosporium cladosporioides PAJ 03 (88.51 ± 1.0), Phomopsis stipata SC 04 (83.44 ± 7.7), Trichoderma viridae PAJ 01 (64.56 ± 4.0) and Botryosphaeria sp. AM 01 (42.79 ± 1.6). On the other hand, the following 4 fungi stood out in relation to β-glucosidase activity (U/g): Saccharicola sp. EJC 04 (51.56 ± 2.7), Paecilomyces sp. SF 021 (33.19 ± 9.2), Ustilaginoidea sp. CV 04 (29.75 ± 0.8) and Ustilaginoidea sp. XYA 04 (21.72 ± 3.05). Among these fungi, P. stipata SC 04 and Botryosphaeria sp. AM 01 were the best producers of xylanase and β-xilosidase (694,33 and 4,87 U/g, respectively). These 8 fungi were then cultured in new mixtures (1:1 w/w) of lignocellulosic substrates. Botryosphaeria sp. AM01and Saccharicola sp. EJC04 stood out regarding endoglucanase and β-glucosidase activities (184.74 ± 6.0 and 92.28 ± 9.57 U/g, respectively) when cultivated on cotton seed meal and wheat bran and were selected to continue the study. The influence of time cultivation, inoculum amount and substrate initial moisture content was evaluated and the best condition for cellulases production was 192 h, six mycelial plugs and 65%, respectively, for both fungi. Cellulases and xylanases produced under these conditions were characterized and optimum pH and temperature values were between 4.5-6 and 60-75°C, respectively. The enzymes were stable over a wide pH range and under 30-70°C. β-glucosidase from both isolates retained about 75-80% of their activity in the presence of glucose at 6 mM. The presence of ethanol stimulated β-glucosidase activity from Botryosphaeria sp. AM01 (about 60% higher in the presence of ethanol at 15%). On the other hand, the activity of β-glucosidase produced by Saccharicola sp. EJC 04 was reduced at ethanol concentrations above 15%. A blend of the enzymatic extracts was used to saccharify pretreated sugarcane bagasse and a face-centered central composite design was used to find the best conditions. Under the predicted optimum condition (50°C, 5% of sugarcane bagasse, 150 U g −1 of endoglucanase and 20 h), glucose and xylose concentrations obtained were 3.56 and 1.66 mg mL −1 , respectively. These results show that the 14 endophytic fungi studied have potential to be explored as producers of plant material degrading enzymes. Botryosphaeria sp. AM01 and Saccharicola sp. EJC 04 are promising in relation

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This study evaluated D-xylose-assimilating yeasts that are associated with rotting wood from the Galápagos Archipelago, Ecuador, for xylitol production from hemicellulose hydrolysates. A total of 140 yeast strains were isolated. Yeasts related to the clades Yamadazyma, Kazachstania, Kurtzmaniella, Lodderomyces, Metschnikowia and Saturnispora were predominant. In culture assays using sugarcane bagasse hemicellulose hydrolysate, Candida tropicalis CLQCA-24SC-125 showed the highest xylitol production, yield and productivity (27.1 g L(-1) xylitol, Y p/s (xyl) = 0.67 g g(-1), Qp = 0.38 g L(-1). A new species of Cyberlindnera, strain CLQCA-24SC-025, was responsible for the second highest xylitol production (24 g L(-1), Y p/s (xyl) = 0.64 g g(-1), Qp = 0.33 g L(-1) h(-1)) on sugarcane hydrolysate. The new xylitol-producing species Cyberlindnera galapagoensis f.a., sp. nov., is proposed to accommodate the strain CLQCA-24SC-025(T) (=UFMG-CM-Y517(T); CBS 13997(T)). The MycoBank number is MB 812171.

BACKGROUND: The bioconversion of lignocellulosic biomass to second generation ethanol has the potential to replace some fossil fuels. Sugarcane bagasse (SB) is a readily available feedstock for ethanol production. The aim of this work was to study the enzymatic saccharification of consecutively acid-alkali pretreated SB using different commercial enzyme preparations. RESULTS:Under the most favourable enzymatic hydrolysis conditions, a maximum sugar recovery of 0.89 g of reducing sugars g -1 acid-alkali pretreated SB was obtained. Scanning electron microscopy (SEM) revealed the structural disruption in the SB morphology after acid-alkali pretreatment followed by enzymatic hydrolysis. CONCLUSION: These results improve research strategies and show the susceptibility of SB under different enzymatic actions.This is thought to be the first report on utilization of cellulase AF28918 for SB saccharification into fermentable sugars.

Biotechnological production of xylitol is an attractive route to add value to a sugarcane biorefinery, through utilization of the hemicellulosic fraction of sugarcane straw, whose availability is increasing in Brazil. Herein, supplementation of the sugarcane straw hemicellulosic hydrolyzate (xylose 57gL(-1)) with maltose, sucrose, cellobiose or glycerol was proposed, and their effect as co-substrates on xylitol production by Candida guilliermondii FTI 20037 was studied. Sucrose (10gL(-1)) and glycerol (0.7gL(-1)) supplementation led to significant increase of 8.88% and 6.86% on xylose uptake rate (1.11gL(-1)h(-1) and 1.09gL(-1)), respectively, but only with sucrose, significant increments of 12.88% and 8.69% on final xylitol concentration (36.11gL(-1)) and volumetric productivity (0.75gL(-1)h(-1)), respectively, were achieved. Based on these results, utilization of complex sources of sucrose, derived from agro-industries, as nutritional supplementation for xylitol production can be proposed as a strategy for improving the yeast performance and reducing the cost of this bioprocess by replacing more expensive nutrients.