The Brazilian Amazonian rainforest harbors a high diversity of yeasts associated with rotting wood, including many candidates for new yeast species.

This study investigated the diversity of yeast species associated with rotting wood in Brazilian Amazonian rainforests. A total of 569 yeast strains were isolated from rotting wood samples collected in three Amazonian areas (Universidade Federal do Amazonas-Universidade Federal do Amazonas [UFAM], Piquiá, and Carú) in the municipality of Itacoatiara, Amazon state. The samples were cultured in yeast nitrogen base (YNB)-d-xylose, YNB-xylan, and sugarcane bagasse and corncob hemicellulosic hydrolysates (undiluted and diluted 1:2 and 1:5). Sugiyamaella was the most prevalent genus identified in this work, followed by Kazachstania. The most frequently isolated yeast species were Schwanniomyces polymorphus, Scheffersomyces amazonensis, and Wickerhamomyces sp., respectively. The alpha diversity analyses showed that the dryland forest of UFAM was the most diverse area, while the floodplain forest of Carú was the least. Additionally, the difference in diversity between UFAM and Carú was the highest among the comparisons. Thirty candidates for new yeast species were obtained, representing 36% of the species identified and totaling 101 isolates. Among them were species belonging to the clades Spathaspora, Scheffersomyces, and Sugiyamaella, which are recognized as genera with natural xylose-fermenting yeasts that are often studied for biotechnological and ecological purposes. The results of this work showed that rotting wood collected from the Amazonian rainforest is a tremendous source of diverse yeasts, including candidates for new species.

Strategy to reduce acetic acid in sugarcane bagasse hemicellulose hydrolysate concomitantly with xylitol production by the promising yeast Cyberlindnera xylosilytica in a bioreactor.

The yeast Cyberlindnera xylosilytica UFMG-CM-Y309 has been identified as a promising new xylitol producer from sugarcane bagasse hemicellulosic hydrolysate (SCHH). However, SCHH pretreatment process generates byproducts, which are toxic to cell metabolism, including furans, phenolic compounds, and carboxylic acids, such as acetic acid, typically released at high concentrations. This research aims to reduce acetic acid in sugarcane hemicellulose hydrolysate concomitantly with xylitol production by yeast strain Cy. xylosilytica UFMG-CM-Y309 in a bioreactor by strategically evaluating the influence of volumetric oxygen transfer coefficient (kLa) (21 and 35 h-1). Experiments were conducted on a bench bioreactor (2 L volumetric capacity) at different initial kLa values (21 and 35 h-1). SCHH medium was supplemented with rice bran extract (10 g L-1) and yeast extract (1 g L-1). Cy. xylosilytica showed high xylitol production performance (19.56 g L-1), xylitol yield (0.56 g g-1) and, maximum xylitol-specific production rate (µpmáx 0.20 gxylitol·g-1 h-1) at kLa value of 21 h-1, concomitantly slowing the rate of acetic acid consumption. A faster acetic acid consumption (100%) by Cy. xylosilytica was observed at kLa of 35 h-1, concomitantly with an increase in maximum cellular growth (14.60 g L-1) and reduction in maximum xylitol production (14.56 g L-1 and Yp/s 0.34 g g-1). This study contributes to pioneering research regarding this yeast performance in bioreactors, emphasizing culture medium detoxification and xylitol production.

Practical way to apply fourth-generation assessment tools integrated into creating meaningful learning experiences in biology at high school.

The learning process for a Biology topic regarding organisms and animal kingdom diversity was investigated through an innovative Interactive Didactic Sequence (IDS) which integrated the idea of "concept maps" with the Hermeneutic-Dialectic Circle (HDC). HDC is a tool for data collection and a reference for pluralist-constructivist thinking, considered a form of fourth-generation evaluation. Hofstede's cultural dimensions were also integrated into the investigation in order to facilitate mediation in an evaluative context. Students' performances (N = 25) from a São Paulo-Brazil public school were statistically evaluated. Their cultural profile was determined via the Hofstede Value Survey Model 1994 questionnaire. The elaborative process of arranging concept maps was individual (CM-individual) and integrated with HDC in groups (CM-HDC). Concept map assessment methods were based off existing literature. An improvement in students' performances (p < 0.05) that presented concept maps integrated to HDC in a more complex structure when compared to individually-built maps was observed. Employment of HDC helped form motivational/interactive dialogues between students and teachers, which, in turn, assisted in achieving greater learning through the use of concept maps. The application of the fourth-generation evaluation was improved via knowledge regarding students' cultural profiles.

Comprehensive approach of methods for microstructural analysis and analytical tools in lignocellulosic biomass assessment - A review.

The trend in the modern world is to replace fossil fuels with green energy sources in order to reduce their environmental impact. The biorefinery industry, within this premise, needs to establish quantitative and qualitative analytical methods to better understand lignocellulosic biomass composition and structure. This paper presents chemical techniques (chromatography, thermal analysis, HRMS, FTIR, NIR, and NMR) and physicochemical techniques (XRD, optical and electron microscopy techniques - Confocal fluorescence, Raman, SPM, AFM, SEM, and TEM) for the microstructural characterization of lignocellulosic biomass and its derivatives. Each of these tools provides different and complementary information regarding molecular and microstructural composition of lignocellulosic biomass. Understanding these properties is essential for the design and operation of associated biomass conversion processing facilities. PAT, monitored in real-time, ensures an economical and balanced mass-energy process. This review aimed to help researchers select the most suitable analytical technique with which to investigate biomass feedstocks with recalcitrant natures.

Diversity and physiological characterization of D-xylose-fermenting yeasts isolated from the Brazilian Amazonian Forest.

BACKGROUND: This 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 FINDINGS: A 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/SIGNIFICANCE: This 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.

Response surface methodology (RSM) to evaluate moisture effects on corn stover in recovering xylose by DEO hydrolysis.

Response surface methodology (RSM), based on a 2(2) full factorial design, evaluated the moisture effects in recovering xylose by diethyloxalate (DEO) hydrolysis. Experiments were carried out in laboratory reactors (10 mL glass ampoules) containing corn stover (0.5 g) properly ground. The ampoules were kept at 160 °C for 90 min. Both DEO concentration and corn stover moisture content were statistically significant at 99% confidence level. The maximum xylose recovery by the response surface methodology was achieved employing both DEO concentration and corn stover moisture at near their highest levels area. We amplified this area by using an overlay plot as a graphical optimization using a response of xylose recovery more than 80%. The mathematical statistical model was validated by testing a specific condition in the satisfied overlay plot area. Experimentally, a maximum xylose recovery (81.2%) was achieved by using initial corn stover moisture of 60% and a DEO concentration of 4% w/w. The mathematical statistical model showed that xylose recovery increases during DEO corn stover acid hydrolysis as the corn stover moisture level increases. This observation could be important during the harvesting of corn before it is fully dried in the field. The corn stover moisture was an important variable to improve xylose recovery by DEO acid hydrolysis.

Xylitol production from DEO hydrolysate of corn stover by Pichia stipitis YS-30.

Corn stover that had been treated with vapor-phase diethyl oxalate released a mixture of mono- and oligosaccharides consisting mainly of xylose and glucose. Following overliming and neutralization, a D-xylulokinase mutant of Pichia stipitis, FPL-YS30 (xyl3-∆1), converted the stover hydrolysate into xylitol. This research examined the effects of phosphoric or gluconic acids used for neutralization and urea or ammonium sulfate used as nitrogen sources. Phosphoric acid improved color and removal of phenolic compounds. D-Gluconic acid enhanced cell growth. Ammonium sulfate increased cell yield and maximum specific cell growth rate independently of the acid used for neutralization. The highest xylitol yield (0.61 g(xylitol)/g(xylose)) and volumetric productivity (0.18 g(xylitol)/g(xylose )l) were obtained in hydrolysate neutralized with phosphoric acid. However, when urea was the nitrogen source the cell yield was less than half of that obtained with ammonium sulfate.

The roles of xylan and lignin in oxalic acid pretreated corncob during separate enzymatic hydrolysis and ethanol fermentation.

High yields of hemicellulosic and cellulosic sugars are critical in obtaining economical conversion of agricultural residues to ethanol. To optimize pretreatment conditions, we evaluated oxalic acid loading rates, treatment temperatures and times in a 2(3) full factorial design. Response-surface analysis revealed an optimal oxalic acid pretreatment condition to release sugar from the cob of Zea mays L. ssp. and for Pichia stipitis CBS 6054. To ferment the residual cellulosic sugars to ethanol following enzymatic hydrolysis, highest saccharification and fermentation yields were obtained following pretreatment at 180 degrees C for 50 min with 0.024 g oxalic acid/g substrate. Under these conditions, only 7.5% hemicellulose remained in the pretreated substrate. The rate of cellulose degradation was significantly less than that of hemicellulose and its hydrolysis was not as extensive. Subsequent enzymatic saccharification of the residual cellulose was strongly affected by the pretreatment condition with cellulose hydrolysis ranging between 26.0% and 76.2%. The residual xylan/lignin ratio ranged from 0.31 to 1.85 depending on the pretreatment condition. Fermentable sugar and ethanol were maximal at the lowest ratio of xylan/lignin and at high glucan contents. The model predicts optimal condition of oxalic acid pretreatment at 168 degrees C, 74 min and 0.027 g/g of oxalic acid. From these findings, we surmised that low residual xylan was critical in obtaining maximal glucose yields from saccharification.

Simultaneous saccharification and ethanol fermentation of oxalic acid pretreated corncob assessed with response surface methodology.

Response surface methodology was used to evaluate optimal time, temperature and oxalic acid concentration for simultaneous saccharification and fermentation (SSF) of corncob particles by Pichia stipitis CBS 6054. Fifteen different conditions for pretreatment were examined in a 2(3) full factorial design with six axial points. Temperatures ranged from 132 to 180 degrees C, time from 10 to 90 min and oxalic acid loadings from 0.01 to 0.038 g/g solids. Separate maxima were found for enzymatic saccharification and hemicellulose fermentation, respectively, with the condition for maximum saccharification being significantly more severe. Ethanol production was affected by reaction temperature more than by oxalic acid and reaction time over the ranges examined. The effect of reaction temperature was significant at a 95% confidence level in its effect on ethanol production. Oxalic acid and reaction time were statistically significant at the 90% level. The highest ethanol concentration (20 g/l) was obtained after 48 h with an ethanol volumetric production rate of 0.42 g ethanol l(-1) h(-1). The ethanol yield after SSF with P. stipitis was significantly higher than predicted by sequential saccharification and fermentation of substrate pretreated under the same condition. This was attributed to the secretion of beta-glucosidase by P. stipitis. During SSF, free extracellular beta-glucosidase activity was 1.30 pNPG U/g with P. stipitis, while saccharification without the yeast was 0.66 pNPG U/g.

Fermentation kinetics for xylitol production by a Pichia stipitis D: -xylulokinase mutant previously grown in spent sulfite liquor.

Spent sulfite pulping liquor (SSL) contains lignin, which is present as lignosulfonate, and hemicelluloses that are present as hydrolyzed carbohydrates. To reduce the biological oxygen demand of SSL associated with dissolved sugars, we studied the capacity of Pichia stipitis FPL-YS30 (xyl3Delta) to convert these sugars into useful products. FPL-YS30 produces a negligible amount of ethanol while converting xylose into xylitol. This work describes the xylose fermentation kinetics of yeast strain P.stipitis FPL-YS30. Yeast was grown in rich medium supplemented with different carbon sources: glucose, xylose, or ammonia-base SSL. The SSL and glucose-acclimatized cells showed similar maximum specific growth rates (0.146 h(-1)). The highest xylose consumption at the beginning of the fermentation process occurred using cells precultivated in xylose, which showed relatively high specific activity of glucose-6-phosphate dehydrogenase (EC 1.1.1.49). However, the maximum specific rates of xylose consumption (0.19 g(xylose)/g(cel) h) and xylitol production (0.059 g(xylitol)/g(cel) h) were obtained with cells acclimatized in glucose, in which the ratio between xylose reductase (EC 1.1.1.21) and xylitol dehydrogenase (EC 1.1.1.9) was kept at higher level (0.82). In this case, xylitol production (31.6 g/l) was 19 and 8% higher than in SSL and xylose-acclimatized cells, respectively. Maximum glycerol (6.26 g/l) and arabitol (0.206 g/l) production were obtained using SSL and xylose-acclimatized cells, respectively. The medium composition used for the yeast precultivation directly reflected their xylose fermentation performance. The SSL could be used as a carbon source for cell production. However, the inoculum condition to obtain a high cell concentration in SSL needs to be optimized.

Enhanced xylitol production by precultivation of Candida guilliermondii cells in sugarcane bagasse hemicellulosic hydrolysate.

The present work evaluated the key enzymes involved in xylitol production (xylose reductase [XR] and xylitol dehydrogenase [XDH]) and their correlation with xylose, arabinose, and acetic acid assimilation during cultivation of Candida guilliermondii FTI 20037 cells in sugarcane bagasse hemicellulosic hydrolysate. For this purpose, inocula previously grown either in sugarcane bagasse hemicellulosic hydrolysate (SBHH) or in semidefined medium (xylose as a substrate) were used. The highest xylose/acetic acid consumption ratio (1.78) and the lowest arabinose consumption (13%) were attained in the fermentation using inoculum previously grown in semidefined medium (without acetic acid and arabinose). In this case, the highest values of XR (1.37 U mg prot(-1)) and XDH (0.91 U mg prot(-1)) activities were observed. The highest xylitol yield (approximately 0.55 g g(-1)) and byproducts (ethanol and glycerol) formation were not influenced by inoculum procedure. However, the cell previously grown in the hydrolysate was effective in enhancing xylitol production by keeping the XR enzyme activity at high levels (around 0.99 U.mg(prot) (-1)), reducing the XDH activity (34.0%) and increasing xylitol volumetric productivity (26.5%) with respect to the inoculum cultivated in semidefined medium. Therefore, inoculum adaptation to SBHH was shown to be an important strategy to improve xylitol productivity.