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.

Effect of synthetic and natural media on lipid production from Fusarium oxysporum

Background: Dependence on fossil resources, for the production of fuels and energy, has resulted in environmental and financial problems, which require our immediate action in order to reverse the situation. Use of renewable sources for the production of fuels and energy is an important alternative with biodiesel remains as one of the promising options. Aim of this work is to evaluate the fungus Fusarium oxysporum for its potentials to accumulate microbial lipids when grown on synthetic media and saccharified sweet sorghum stalks. Results: The effect of different carbon sources, nitrogen sources and C/N ratio on the lipid production was initially examined, which resulted in a lipid concentration of 4.4 g/L, with lipid content of 42.6% w/w. Sweet sorghum stalks were able to support growth and lipid production of the fungus, both as carbon source and as nitrogen source. It was also shown that saccharification of the dried stalks is an important step to increase lipid production. Removal of the remaining stalk solids enabled the lipid production during cultivation in increased initial solids of up to 16 w/w. This resulted in a lipid production of 3.81 g/L. Conclusions: It was demonstrated that F. oxysporum can be used as an efficient oleaginous microorganism, with sweet sorghum serving as an excellent raw material for the cultivation of the fungus. The lipids obtained during this work were also found to have a fatty acid profile with good potentials to be used for biodiesel production.

Highly efficient enzymatic preparation of isomalto-oligosaccharides from starch using an enzyme cocktail

Background: Current commercial production of isomalto-oligosaccharides (IMOs) commonly involves a lengthy multistage process with low yields. Results: To improve the process efficiency for production of IMOs, we developed a simple and efficient method by using enzyme cocktails composed of the recombinant Bacillus naganoensis pullulanase produced by Bacillus licheniformis, α-amylase from Bacillus amyloliquefaciens, barley bran ß-amylase, and α-transglucosidase from Aspergillus niger to perform simultaneous saccharification and transglycosylation to process the liquefied starch. After 13 h of reacting time, 49.09% IMOs (calculated from the total amount of isomaltose, isomaltotriose, and panose) were produced. Conclusions: Our method of using an enzyme cocktail for the efficient production of IMOs offers an attractive alternative to the process presently in use.

High levels of ß-xylosidase in Thermomyces lanuginosus: potential use for saccharification

ABSTRACT A new strain of Thermomyces lanuginosus was isolated from the Atlantic Forest biome, and its β-xylosidases optimization in response to agro-industrial residues was performed. Using statistical approach as a strategy for optimization, the induction of β-xylosidases activity was evaluated in residual corn straw, and improved so that the optimum condition achieved high β-xylosidases activities 1003 U/mL. According our known, this study is the first to show so high levels of β-xylosidases activities induction. In addition, the application of an experimental design with this microorganism to induce β-xylosidases has not been reported until the present work. The optimal conditions for the crude enzyme extract were pH 5.5 and 60 °C showing better thermostability at 55 °C. The saccharification ability of β-xylosidase in the presence of hemicellulose obtained from corn straw raw and xylan from beechwood substrates showed a xylo-oligosaccharide to xylose conversion yield of 80 and 50%, respectively, at 50 °C. Our data strongly indicated that the β-xylosidases activities was not subjected to the effects of potential enzyme inhibitors often produced during fermentation process. These data suggest the application of this enzyme studied for saccharification of hemicellulose, an abundant residue in the American continents, thus providing an interesting alternative for future tests for energy production.

Enzymatic saccharification and fermentation of cellulosic date palm wastes to glucose and lactic acid

Abstract The bioconversion of cellulosic wastes into high-value bio-products by saccharification and fermentation processes is an important step that can reduce the environmental pollution caused by agricultural wastes. In this study, enzymatic saccharification of treated and untreated date palm cellulosic wastes by the cellulases from Geobacillus stearothermophilus was optimized. The alkaline pre-treatment of the date palm wastes was found to be effective in increasing the saccharification percentage. The maximum rate of saccharification was found at a substrate concentration of 4% and enzyme concentration of 30 FPU/g of substrate. The optimum pH and temperature for the bioconversions were 5.0 and 50 °C, respectively, after 24 h of incubation, with a yield of 31.56 mg/mL of glucose at a saccharification degree of 71.03%. The saccharification was increased to 94.88% by removal of the hydrolysate after 24 h by using a two-step hydrolysis. Significant lactic acid production (27.8 mg/mL) was obtained by separate saccharification and fermentation after 72 h of incubation. The results indicate that production of fermentable sugar and lactic acid is feasible and may reduce environmental pollution by using date palm wastes as a cheap substrate.

Effect of ensilage on bioconversion of switchgrass to ethanol based on liquid hot water pretreatment / 生物工程学报

Ensilage is a traditional way of preserving fresh biomass. However, in order to apply ensilage to the ethanol biorefinery, two parameters need to be evaluated: quantity and quality changes of the biomass; and its effects on bioconversion process. To study these two aspects, switchgrass harvested on three different time points (Early, mid and late fall) were used as feedstock. The early fall harvested biomass was ensiled at 5 moisture levels ranging from 30% to 70%. Silage of 40% moisture and 3 other raw switchgrass were pretreated with liquid hot water, followed by enzymatic hydrolysis as well as simultaneous saccharification and fermentation. After 21 days storage pH values of all silages decreased below 4.0 and the dry matter losses were less than 2.0%, and structural sugars contents did not change dramatically. Liquid hot water caused more hemicellulose dissolution in the silage than in unensiled switchgrass. However, ensilage also increased the risk of releasing more sugar degradation products; After enzymatic hydrolysis, silage obtained higher total glucose, xylose and galactose yields than raw materials; After simultaneous saccharification and fermentation, ethanol concentration in silage was 12.1 g/L, higher than the unensiled switchgrass (10.3 g/L, 9.7 g/L and 10.6 g/L for early, mid and late fall respectively). Our results suggest that ensilage helps increase pretreatment efficiency and sugar yield, which increases final ethanol production.

Adsorption Kinetics Studies of Endoglucanase from Rhizopus Oryzae on Activated Charcoal.

The adsorption kinetics and activities of endoglucanase enzyme from Rhizopus oryzae were evaluated using activated commercial charcoal as adsorbent. The effect of various experimental parameters on adsorption of endoglucanase such as initial enzyme concentration, amount of adsorbent, contact time and temperature were investigated. The pseudo-first-order and pseudo second-order kinetic models were used to describe the kinetic data which showed that the adsorption of the enzyme followed the pseudo-first order rate expression. The adsorbed enzyme was subjected to saccharification in presence of commercially available carboxy methyl cellulose and the reusability of the adsorbed enzyme was also tested.

Isolation and Optimization of Cellulolytic and Xylanolytic Microbes from Nepal and their Utilization for Lignocellulosic Biomass Degradation.

Microorganisms harbored by nature and guts of herbivorous animals can degrade different plant related biomass. One of the in-between steps for conversion of lignocellulosic biomass to ethanol entails isolation and identification of microorganisms that could convert pretreated biomass into a suitable form, which could then be fermented into bioethanol. We isolated 36 different microorganisms from hot spring, 6 from ruminant’s (goat) gut, 2 sample from hay spray on the basis of their ability to secrete enzymes that hydrolyzed different plant constituents. Similarly 3 microorganisms were isolated from the rotten wood available around Kathmandu University on the basis of utilization of xylose and glucose. 3 varieties of thermophiles, gut microorganism and microorganism from hay spray that showed the highest cellulolytic and xylanolytic activities by saccharification of cellulose and xylan into their monosaccharide glucose and xylose units respectively were then applied on different biomass (rice straw, corn stover and sugarcane bagasse). Before Saccharification biomass was made accessible for the digestion by enzymes through 3 different pretreatment strategies (3.35% H2SO4, NaOH and H2O2 with 1:10 substrate: chemical ratio) following thermal strategy of steam explosion. Also, different conditions like incubation time, pH and temperature for saccharification were assessed with the highest liberation of reducing sugar at pH 5, temperature of 5̊ C and incubation time of 4 days. Microorganism from rotten wood was able to utilize both xylose and glucose and yielded highest amount (5.567 mg/ml) of bioethanol.

Optimization of enzymatic saccharification of microwave pretreated sugarcane tops through response surface methodology for biofuel.

The optimization of biomass loading enzyme loading, surfactant concentration and incubation time, using response surface methodology (RSM) and Box Behnken design for enzymatic saccharification of sugarcane tops (SCT) for maximum recovery of fermentable sugars using crude cellulases, resulted in 90.24% saccharification efficiency. Maximum saccharification yield of 0.376 g/g glucose as substrate for ethanol production was observed at optimal conditions of 10% biomass loading (pretreated), 100FPU/g of cellulase loading, 0.04% (w/w) surfactant concentration and 72 h of incubation time.

Cultivation of Trichosporon mycotoxinivorans in sugarcane bagasse hemicellulosic hydrolyzate

Background: The yeast strain IB09 was isolated from the gut of Calosoma sp. (Carabidae, Coleoptera, Insecta) that were collected in the central Amazon rainforest. First, tolerance of the strain to ethanol and heat was tested. Then, IB09 was cultivated in a medium using sugarcane bagasse hemicellulosic hydrolyzate as a carbon source, and cell growth (OD600), specific growth rate (uMAX, h-1), biomass yield (Y B, g.g-1) and relative sugar consumption (RSC, percent) were evaluated. Taxonomic identification was determined by sequencing the ITS1 region of IB09 and comparing it to sequences obtained from the GenBank database (NCBI). Results: IB09 showed both ethanol tolerance and thermotolerance. Relative sugar consumption indicated that IB09 was able to perform saccharification of sugarcane bagasse hemicellulosic hydrolyzate, increasing the total reducing sugar concentration by approximately 50 percent. The μMAX value obtained was 0,20, indicating that cell growth was slow under the assessed conditions. Biomass yield was 0,701 g per g of consumed sugar, which is relatively high when compared with other findings in the literature. After 120 hrs of cultivation, 80,1 percent of total reducing sugar had been consumed. Sequencing of the ITS1 region identified IB09 as Trichosporon mycotoxinivorans. Conclusion: This is the first report to document this species in the central Amazon rainforest at this host. Trichosporon mycotoxinivorans has great biotechnological potential for use in the saccharification of sugarcane bagasse hemicellulosic hydrolyzate and for biomass production with this substrate as carbon source.

Bioconversion and saccharification of some lignocellulosic wastes by aspergillus oryzae ITCC-4857.01 for fermentable sugar production

The recent interest in bioconversion of agricultural and industrial wastes to chemical feedstock has led to extensive studies on cellulolytic enzymes produced by microorganisms. In the present study three lignocellulosic substrates viz. sugarcane bagasse, sawdust and water hyacinth were pre-treated with alkali and enzyme and their effect on bioconversion has been investigated. The ability of selected substrates for induction of cellulase enzyme by A. oryzae ITCC 4857.01 and for the potentiality of the induced enzyme to saccharify the substrates were also assessed. The maximum degree of conversion of substrate (0.415 percent) and improved specific substrate consumption (0.99 g substrate/g dry biomass) was exhibited in sugarcane bagasse after alkali treatment at 96 hrs. Both alkali-treatment and enzyme-treatment, water hyacinth was the best for cellulase induction and showed maximum endoglucanase activity of 11.42 U/ml. Reducing sugar yield ranged from 1.12 mg/ml for enzyme treated sawdust at 48 hrs to 7.53 mg/ml for alkali treated sugarcane bagasse at 96 hrs. Alkali-treated sugarcane bagasse gave the highest saccharification rate of 9.03 percent after 96 hrs. The most resistant substrate was sawdust which produced 5.92 percent saccharification by alkaline treatment. The saccharification of lignocellulosic substrates by enzyme produced by A. oryzae ITCC 4857.01 indicates the enzymes specificity towards the substrates. The use of such enzyme in lingo-cellulose hydrolysis will lead to efficient conversion of cellulose materials to other important products.

Evaluation of Potential Fungal Species for the in situ Simultaneous Saccharification and Fermentation (SSF) of Cellulosic Material

Three fungal species were evaluated for their abilities to saccharify pure cellulose. The three species chosen represented three major wood-rot molds; brown rot (Gloeophyllum trabeum), white rot (Phanerochaete chrysosporium) and soft rot (Trichoderma reesei). After solid state fermentation of the fungi on the filter paper for four days, the saccharified cellulose was then fermented to ethanol by using Saccharomyces cerevisiae. The efficiency of the fungal species in saccharifying the filter paper was compared against a low dose (25 FPU/g cellulose) of a commercial cellulase. Total sugar, cellobiose and glucose were monitored during the fermentation period, along with ethanol, acetic acid and lactic acid. Results indicated that the most efficient fungal species in saccharifying the filter paper was T. reesei with 5.13 g/100 g filter paper of ethanol being produced at days 5, followed by P. chrysosporium at 1.79 g/100 g filter paper. No ethanol was detected for the filter paper treated with G. trabeum throughout the five day fermentation stage. Acetic acid was only produced in the sample treated with T. reesei and the commercial enzyme, with concentration 0.95 and 2.57 g/100 g filter paper, respectively at day 5. Lactic acid production was not detected for all the fungal treated filter paper after day 5. Our study indicated that there is potential in utilizing in situ enzymatic saccharification of biomass by using T. reesei and P. chrysosporium that may lead to an economical simultaneous saccharification and fermentation process for the production of fuel ethanol.

Producción de eaanol a partir de almidón de yuc utilizando la estrategia de prroceso sacarificación- fermentación simultáneas (ssf) / Ethanol prodctioon from cassva starch using the process strategy simultaneoous saccharification-fermentation

La tendencia mundial en el manejo de los combustibles, en especial los biocombustibles como el etanol, ha llevado a explorar nuevas metodologías de proceso para optimizar su producción; por tal razón se aborda en esta investigación el proceso sacarificación fermentación simultáneas, se evalúa la influencia de la concentración de azúcares reductores y la dosificación de la enzima Spirizyme fuel® sobre la productividad y concentración final de etanol, bajo el proceso SSF (sacarificación -fermentación simultáneas), partiendo del licuado de almidón de yuca como sustrato. El proceso SSF se compara con un control con características de sacarificación-fermentación independientes (SHF), proceso convencional. Sólo el factor concentración inicial de sustrato presenta efecto sobre la productividad de etanol. Las cinéticas de proceso, frente a las del control, presentan reducciones de tiempo de 47 y 33% para los niveles de sustrato evaluados. Los niveles de productividad son mayores en un 33% para el nivel de 150 g/l de AR (azúcares reductores) y se mantiene constante para 200 g/l. La glucosa en la estrategia SSF, conforme se produce se transforma en etanol, no permitiendo alcanzar concentraciones superiores a 100 g/l, lo que se traduce en que no se presentan inhibiciones por sustrato. La concentración de etanol no afecta la reacción de la enzima en el proceso de sacarificación. El proceso SSF demuestra su viabilidad técnica en la producción de alcohol, al reducir los tiempos y necesidades de energía en la producción de alcohol carburante a partir de almidón de yuca.

The world trend on fuel management, in special biofuels like ethanol, have gone to explorer new methodologies of process to optimize its production by this reason in this research is about simultaneous sacarification fermentation process and evaluate initial concentration of reducing sugar, and enzyme dosing of Spirizyme fuel® are evaluated on productivity and final concentration of ethanol, under SSF (Simultaneous Saccharification and Fermentation) process, from the product of the licuefaction process of cassava starch as substrate. The SSF process is evaluated against SHF (Independent Saccharification and Fermentation) process as control. Only the factor, initial concentration of substrate presents effect over ethanol productivity. The kinetic of SSF process, in opposite to the SHF process, presents time diminution of the global process around 47 y 33% to substrate levels of 150 and 200 g/l respectively. The productivity values are most at a 33% to 150 g/l of reducing sugar, and they keep constant to 200 g/l reducing sugar. The glucose in SSF strategy, at the time it is producing, it is transformed to ethanol, does not allowing to reach superior concentration to 100 g/l of reducing sugar, this implicates there is not substrate inhibition. The ethanol concentration doesn't affect the enzymatic process of sacharification. The SSF process demonstrates his technical viability on the ethanol production, to reduce time an energy requirements on the ethanol production from cassava flour.

Study on the Saccharification Processes of Lignocellulose Brought about by Ultrasonic Wave / 中国生物工程杂志

Both pretreatment of lignocellulose and its saccharification process are treated with ultrasonic wave.The morphology,structure and crystal performance of the original and treated lignocellulose sample were characterized by SEM and FTIR.Moreover,the changes of raw materials caused by different pretreatment ways and the affect of saccharification rate brought about by ultrasonic wave were also studied.The result shows that the ultrasonic wave decreases the crystallinity of lignocellulose destroying the intermolecular hydrogen bonding effectively and improves the degradation rate of lignin and the saccharification rate of zymohydrolysis availably.The mechanism of activation of ultrasonic wave in zymohydrolysis process was discussed primarily.