High Bioethanol Production Research Articles

Mild chemical pretreatments are sufficient for bioethanol production in transgenic rice straws overproducing glucosidase

A cost-effective biomass conversion technology for high bioethanol production under mild chemical pretreatment in the transgenic rice straw.

Biomass saccharification is largely enhanced by altering wall polymer features and reducing silicon accumulation in rice cultivars harvested from nitrogen fertilizer supply

In this study, two rice cultivars were collected from experimental fields with seven nitrogen fertilizer treatments. All biomass samples contained significantly increased cellulose contents and reduced silica levels, with variable amounts of hemicellulose and lignin from different nitrogen treatments. Under chemical (NaOH, CaO, H2SO4) and physical (hot water) pretreatments, biomass samples exhibited much enhanced hexoses yields from enzymatic hydrolysis, with high bioethanol production from yeast fermentation. Notably, both degree of polymerization (DP) of cellulose and xylose/arabinose (Xyl/Ara) ratio of hemicellulose were reduced in biomass residues, whereas other wall polymer features (cellulose crystallinity and monolignol proportion) were variable. Integrative analysis indicated that cellulose DP, hemicellulosic Xyl/Ara and silica are the major factors that significantly affect cellulose crystallinity and biomass saccharification. Hence, this study has demonstrated that nitrogen fertilizer supply could largely enhance biomass saccharification in rice cultivars, mainly by reducing cellulose DP, hemicellulosic Xyl/Ara and silica in cell walls.

Development of combined nanofiltration and forward osmosis process for production of ethanol from pretreated rice straw

A membrane process combining nanofiltraion (NF) and forward osmosis (FO) was developed for the sugar concentration with the aim of high bio-ethanol production from the liquid fraction of rice straw. The commercial NF membrane, ESNA3, was more adequate for removal of fermentation inhibitors (such as acetic acid) than the FO membrane, whereas the commercial FO membrane, TFC-ES, was more adequate for concentration of the sugars than the NF membrane. The liquid fraction was subjected to the following process: NF concentration with water addition (NF(+H2O))→enzymatic hydrolysis→FO concentration. This NF(+H2O)-FO hybrid process generated a total sugar content of 107g·L−1. Xylose-assimilating S. cerevisiae produced 24g·L−1 ethanol from the liquid fraction that was diluted 1.5-fold and then concentrated by the NF(+H2O)-FO hybrid process. The NF(+H2O)-FO hybrid process has the potential for optimized ethanol production from pretreated lignocellulosic biomass.

Life cycle assessment of bioethanol production from three feedstocks and two fermentation waste reutilization schemes

More than 95% of energy supplies for Taiwan are imported. Conversely, approximately 150,000 ha half-fallow and 50,000 ha fallow paddy fields exist because of increasing labor costs and competition from rice imports. The potential for energy crop production from such underutilized paddy fields merits significant attention. However, selecting crops for optimal environmental performance when producing bioethanol is equally important. The life cycle assessments (LCAs) of rice straw (agricultural waste), Napier grass (energy crop), and Eucalyptus spp. (short rotation coppice) as bioethanol feedstocks in such fallow paddy fields were investigated in this study. The LCAs of two schemes for on-site fermentation waste utilization were also investigated as pellet fuel and molded pulp feedstocks. Experimental and field survey data were used in this study. Due to higher biomass yields, Napier grass and Eucalyptus spp. resulted in 47% and 28%, respectively, less weight-based negative impacts compared to rice. Pursuing high bioethanol production requires consideration of the overall environmental loading. Conversely, the rankings of the three crops based on acreage and bioethanol yield differed due to extensive farming practices and bioethanol conversion yields. On-site production of pellet fuel using fermentation waste was 7% and 31% higher than that of Eucalyptus biomass pellets and imported coal, respectively, for Taiwan. On-site production of molded pulp products using fermentation waste was 5% and 49% higher than that of recycled newspaper and virgin chemical pulp, respectively, for Taiwan. Thus, fermentation waste utilization schemes could provide a broader evaluation for planning alternative crops for bioethanol production.

Batch Fermentation Options for High Titer Bioethanol Production from a SPORL Pretreated Douglas-Fir Forest Residue without Detoxification

This study evaluated batch fermentation modes, namely, separate hydrolysis and fermentation (SHF), quasi-simultaneous saccharification and fermentation (Q-SSF), and simultaneous saccharification and fermentation (SSF), and fermentation conditions, i.e., enzyme and yeast loadings, nutrient supplementation and sterilization, on high titer bioethanol production from SPORL-pretreated Douglas-fir forest residue without detoxification. The results indicated that Q-SSF and SSF were obviously superior to SHF operation in terms of ethanol yield. Enzyme loading had a strong positive correlation with ethanol yield in the range studied. Nutrient supplementation and sterility were not necessary for ethanol production from SPORL-pretreated Douglas-fir. Yeast loading had no substantial influence on ethanol yield for typical SSF conditions. After 96 h fermentation at 38 °C on shake flask at 150 rpm, terminal ethanol titer of 43.2 g/L, or 75.1% theoretical based on untreated feedstock glucan, mannan, and xylan content was achieved, when SSF was conducted at whole slurry solids loading of 15% with enzyme and yeast loading of 20 FPU/g glucan and 1.8 g/kg (wet), respectively, without nutrition supplementation and sterilization. It is believed that with mechanical mixing, enzyme loading can be reduced without reducing ethanol yield with extended fermentation duration.

Production of bioethanol from a mixture of agricultural feedstocks: Biofuels characterization

Bioethanol production from a mixture of agricultural feedstock was carried out in a batch fermenter. To study the effect of optimal mixture ratio of fruit juices (100% dates; 70% dates+30% grapes, 50% dates+50% grapes, 30% dates+70% grapes, 100% grapes) on ethanol yield (g ethanol/g sugar), different parameters have been monitored such as ethanol yield, total sugar, ammoniacal nitrogen, pH. It was found that the mixture of Mech Degla dates and grapes juices has the higher bioethanol concentration than that obtained from a single juice from one kind of fruit. Higher bioethanol production 155.5g/L at 72h was obtained when 30% dates juice was blended with 70% grapes juice. In such case, the higher initial sugar concentration is close to 228.34g/L, and the pH droped from 4.5 to 3.87. The Luedeking-Piret model was used to describe the bioethanol production. A good agreement was found between simulated and experimental data. The influence of some physical properties on the addition of ethanol to lead-free gasoline has been studied. It was found that the addition of 5% of ethanol, to lead-free gasoline raises the RON to around 96.4, that can improve its octane number by 2 points.

Immobilization of Zymomonas mobilis with Fe2O3-modified polyvinyl alcohol for continuous ethanol fermentation

Zymomonas mobilis cells were immobilized with a novel Fe2O3-modified polyvinyl alcohol (PVA) matrix to perform continuous ethanol fermentation with very high efficiency. The key operating parameters, such as glucose feeding concentration, dilution rate, and cells loading were optimized to achieve high bioethanol production performance at a high dilution rate and high glucose utilization efficiency. The optimal conditions for ethanol fermentation with the modified PVA immobilized cells were determined as: cell loading, 40% (w/v); glucose feeding concentration, 125g/L; hydraulic retention time (HRT), 2h (or 0.5h⿿1 dilution rate). The bioethanol productivity was nearly doubled (31.09g/L/h) when the PVA-immobilized cells were modified with 1% (w/v) Fe2O3. The modified PVA-immobilized cells could be utilized for more than 20days without losing their activity.

Experimental investigations on bioethanol production from halophilic microalgal biomass

Although the microalgae biomass is a significant feedstock for bioethanol production, studies on “third generation” of algal bioethanol is at its early stages of investigation. In this study halophilic microalgae Dunaliella sp. was used for producing bioethanol. Physical and physicochemical methods of pretreatment procedures were used. Some important parameters such as KNO3 (0–1000mg/L), KH2PO4 (0–35mg/L), MgSO4 (0–500mg/L) nutrient amendments and NaCl (15%–30%) concentrations for Dunaliella sp. cultivation were optimized. Changing fermentation pH within 4–7 range, and yeast inoculum amount by inoculation of 0.5–3ml volumes were used for optimization of the bioethanol production processes. Dilute acidic hydrolysis with 1% sulfuric acid in autoclave was found to be very effective in saccharifying algal biomass and higher bioethanol production values were obtained under nitrogen-depletion conditions. The highest bioethanol production by Saccharomyces cerevisiae was obtained at 72h of incubation at substrate concentration of 30g/L microalgal biomass and 3ml inoculums at pH 6. The results demonstrated that at the optimized saccharification and fermentation conditions the bioethanol production could be increased up to 7.26g/L, which was 10.7-fold higher than the value obtained by using the microalgal biomass was not exposed to any pretreatment. In the present study we showed that bioethanol production from biomass of a halophilic microalgae, which is known as a promising feedstock for bioethanol production, can be increased by appropriate pretreatments.

Yeasts Associated with Roots of the Endemic Plant Mankyua chejuense

BACKGROUND: Identification of endophytic yeasts inhabiting the internal roots of the Mankyua chejuense tree requires techniques involving biotechnology. There is a need for a culture-based method to isolate and identify yeast strains associated with M. chejuense.METHODS AND RESULTS: We spread homogenized M. chejuense root samples onto glucose-peptone- yeast agar containing antibiotics, Triton X-100, and L-sorbose. A total of 152 yeast isolates were obtained and identified via phylogenetic analysis based on ITS gene sequencing. The results revealed that the root-associated yeast species included the genera Cyberlindnera (140 isolates), Candida (11 isolates), and Kluyveromyces (one isolate). Additionally, three yeast isolates showed high bioethanol production.CONCLUSION: We identified the specific yeast community associated with M. chejuense roots. These yeast isolates may have industrial applications as bioethanol producers. Our findings revealed that Cyberlindnera isolates included C. suaverolens and C. satumus, while Kluyveromyces isolates showed high bioethanol production.

Bioethanol production by Saccharomyces cerevisiae, Pichia stipitis and Zymomonas mobilis from delignified coconut fibre mature and lignin extraction according to biorefinery concept

In search to increase the offer of liquid, clean, renewable and sustainable energy in the world energy matrix, the use of lignocellulosic materials (LCMs) for bioethanol production arises as a valuable alternative. The objective of this work was to analyze and compare the performance of Saccharomyces cerevisiae, Pichia stipitis and Zymomonas mobilis in the production of bioethanol from coconut fibre mature (CFM) using different strategies: simultaneous saccharification and fermentation (SSF) and semi-simultaneous saccharification and fermentation (SSSF). The CFM was pretreated by hydrothermal pretreatment catalyzed with sodium hydroxide (HPCSH). The pretreated CFM was characterized by X-ray diffractometry and SEM, and the lignin recovered in the liquid phase by FTIR and TGA. After the HPCSH pretreatment (2.5% (v/v) sodium hydroxide at 180 °C for 30 min), the cellulose content was 56.44%, while the hemicellulose and lignin were reduced 69.04% and 89.13%, respectively. Following pretreatment, the obtained cellulosic fraction was submitted to SSF and SSSF. Pichia stipitis allowed for the highest ethanol yield – 90.18% – in SSSF, 91.17% and 91.03% were obtained with Saccharomyces cerevisiae and Zymomonas mobilis, respectively. It may be concluded that the selection of the most efficient microorganism for the obtention of high bioethanol production yields from cellulose pretreated by HPCSH depends on the operational strategy used and this pretreatment is an interesting alternative for add value of coconut fibre mature compounds (lignin, phenolics) being in accordance with the biorefinery concept.

Optimization of Bioethanol Production Using Whole Plant of Water Hyacinth as Substrate in Simultaneous Saccharification and Fermentation Process.

Water hyacinth was used as substrate for bioethanol production in the present study. Combination of acid pretreatment and enzymatic hydrolysis was the most effective process for sugar production that resulted in the production of 402.93 mg reducing sugar at optimal condition. A regression model was built to optimize the fermentation factors according to response surface method in saccharification and fermentation (SSF) process. The optimized condition for ethanol production by SSF process was fermented at 38.87°C in 81.87 h when inoculated with 6.11 ml yeast, where 1.291 g/L bioethanol was produced. Meanwhile, 1.289 g/L ethanol was produced during experimentation, which showed reliability of presented regression model in this research. The optimization method discussed in the present study leading to relatively high bioethanol production could provide a promising way for Alien Invasive Species with high cellulose content.

Molecular characterization of glycosyl hydrolases of Trichoderma harzianum WF5 – A potential strain isolated from decaying wood and their application in bioconversion of poplar wood to ethanol under separate hydrolysis and fermentation

Populous deltoides wood is a suitable raw material for ethanol production, since it has high cellulose content. Efficient degradation and subsequent conversion of lignocellulosic biomass to bio-ethanol, requires physico-chemical pretreatments followed by enzymatic hydrolysis to obtain high overall yields of sugars. Microbial hydrolytic enzymes are actively utilized for biomass degradation and bio-ethanol production. Four glycosyl hydrolases (GHs) namely; three subunits of cellulase, endoglucanase (ThWF5-endoglucanase), exoglucanase (ThWF5-exoglucanase) and β-glucosidase (ThWF5-glucosidase), and xylanase (ThWF5-xylanase), from fungal strain Trichoderma harzianum WF5 - isolated from decayed wood, were pulled out and subsequently characterized. Analysis of full length sequences and prediction of secondary and tertiary structure showed that these enzymes shared homology with the respective enzymes of other Trichoderma species and have comparatively evolved structure to express catalytic activity under broad range of environmental conditions. Respective GHs were partially purified and used for the enzymatic hydrolysis of P. deltoids wood after pretreatments. The pretreatment of P. deltoides wood with sulfuric acid and hydrogen peroxide (H2SO4 + H2O2) at 121 °C for 60 min was determined as the best condition to high recovery of reducing sugars after enzymatic hydrolysis. The overall high ethanol yield of 15.6 g/l and fermentation efficiency of 61.1% was obtained when fermentation was performed on pretreated poplar wood slurry followed by detoxification with calcium hydroxide. High bio-ethanol production from P. deltoides wood and molecular characterization of glycosyl hydrolases used, provide insights that this wood offers a potential source for biofuel production while T. harzianum WF5 is a treasure house of robust degrading enzymes.

An isolated Amycolatopsis sp. GDS for cellulase and xylanase production using agricultural waste biomass.

The aim of this study was to evaluate an isolate of Amycolatopsis sp. GDS for cellulase and xylanase production, their characterization, and its application to the preparation of biomass feedstock for ethanol production. A novel potent cellulolytic bacterial strain was isolated and identified as Amycolatopsis sp. GDS. The strain secreted high levels of cellulase and xylanase in the presence of agricultural waste biomass. The enzymes were thermostable and active up to 70°C. Interestingly, the enzymes were expressed well at higher NaCl (up to 2·5moll(-1) ) and ionic liquid (10%) concentrations, so that they could be used during the pretreatment of biomass. Enzyme stability in the presence of organic solvents, surfactants and oxidizing agents was also noted. Crude enzymes from Amycolatopsis sp. GDS resulted in comparable saccharification (60%) of wheat straw to commercial enzymes (64%). The cellulolytic enzymes from Amycolatopsis sp. GDS were stable, expressed well under conditions with various chemicals, and yielded significant amounts of hydrolysates from the biomass. The high bioethanol production using yeast co-cultures with enzymatic hydrolysates highlights the significance of selecting the strain and substrate for biofuel production. This study demonstrates the importance of the isolate Amycolatopsis sp. GDS that secretes high levels of cellulase and hemicellulase by utilizing agricultural waste biomass and its application in the preparation of biomass feedstock and sequential ethanol fermentation.

Evaluation of bioethanol production from juice and bagasse of some sweet sorghum varieties

Sweet sorghum, with sugar-rich stalks and water-use efficiency, has a very good potential as an alternative feedstock for ethanol and also non-competing with human feed on land. The present study evaluates the exploitation of juice and bagasse of five varieties of sweet sorghum for bioethanol production which can further improve the energy yield of the crop. The sweet sorghum varieties, GK-coba, Mn-1054, Ramada, Mn-4508 and SS-301, were analyzed for their productivity, and sugar and fiber contents. All varieties significantly differed in yield of stripped stalk, juice and bagasse. The sugar-rich juice and the fiber-rich bagasse, resulting from squeezing the striped stalks, were used for bioethanol production by two microorganisms; Saccharomyces cerevisiae ATCC 7754 and Zymomonas mobilis ATCC 29191. Stalks of varieties GK-coba, Mn-4508 and SS-301 contained high sugar contents and thus were utilized for bioethanol production directly from juice. Stalks of varieties Mn-1054, Ramada and SS-301 had higher content of fibers, so their bagasses were used for bioethanol production. Bagasse was pretreated and hydrolyzed thermo-chemically with 2% (v/v) sulfuric acid (98%) at 120°C for 60min and filtered and the sugar-rich filtrate was neutralized and supplemented with nutrients for bioethanol production. Fermentation of sweet sorghum sugars or acid-hydrolyzed neutralized bagasse into bioethanol was conducted by Sacch. cerevisiae, Z. mobilis or mixed-culture of both organisms at 1:1 ratio. The highest bioethanol production was obtained from juice and bagasse of variety SS-301, by the mixed-culture treatment. From the juice, bioethanol concentration was 50.26mLL−1, whereas from bagasse, bioethanol concentration was 10.5mLL−1. Finally, it could be estimated that 160mL of bioethanol can be produced out of each 1kg of variety SS-301, when using both juice and bagasse.

Potential of Jerusalem artichoke (Helianthus tuberosus L.) as a biorefinery crop

The utilization of Jerusalem artichoke in a biorefinery context was not investigated so far. Therefore the aim of this study was to evaluate the potential of this plant as feedstock for production of bioethanol, protein and inulin. We investigated the biomass productivity and chemical composition of 11 different clones, harvested on three occasions during the fall and early winter of 2011. The results obtained showed that cellulose production per hectare was at least double compared to corn stover, rice straw, sugarcane bagasse and wheat straw, showing high bioethanol production potential of Jerusalem artichoke. Although not high (in total <6% of dry matter), protein and lipid levels could be of importance when biomass is utilized in a biorefinery concept. Harvest occasion determined the biomass productivity and chemical composition to a higher degree than the choice of clone. The average biomass productivity was 88% higher in September than in December. Fresh tuber biomass productivity showed large variations between harvests, where the maximum average productivity in December was 3.4 times higher than in September. Inulin content in dry tubers was between 76 and 85% making the plant an excellent crop, for e.g. inulin extraction, production of high fructose syrup or fermentations. Less mature plants were shown to have degree of polymerization (DP) up to 14, which makes biomass useful as dietary fibre, while the inulin DP in tubers harvested later became as low as 6, showing lower potential with respect to plant utilization for dietary purposes. Results obtained clearly demonstrated that crop harvesting time was an important factor affecting biomass productivity and composition.

【ミニレビュー-第1回フレッシュシンポジウム-】 高効率エタノール生産を目指した近赤外スペクトルによる前処理バイオマスのハイスループット解析

【ミニレビュー-第1回フレッシュシンポジウム-】 高効率エタノール生産を目指した近赤外スペクトルによる前処理バイオマスのハイスループット解析

Stochastic optimization of a multi-feedstock lignocellulosic-based bioethanol supply chain under multiple uncertainties

An integrated multi-feedstock (i.e. switchgrass and crop residue) lignocellulosic-based bioethanol supply chain is studied under jointly occurring uncertainties in switchgrass yield, crop residue purchase price, bioethanol demand and sales price. A two-stage stochastic mathematical model is proposed to maximize expected profit by optimizing the strategic and tactical decisions. A case study based on ND (North Dakota) state in the U.S. demonstrates that in a stochastic environment it is cost effective to meet 100% of ND's annual gasoline demand from bioethanol by using switchgrass as a primary and crop residue as a secondary biomass feedstock. Although results show that the financial performance is degraded as variability of the uncertain parameters increases, the proposed stochastic model increasingly outperforms the deterministic model under uncertainties. The locations of biorefineries (i.e. first-stage integer variables) are insensitive to the uncertainties. Sensitivity analysis shows that “mean” value of stochastic parameters has a significant impact on the expected profit and optimal values of first-stage continuous variables. Increase in level of mean ethanol demand and mean sale price results in higher bioethanol production. When mean switchgrass yield is at low level and mean crop residue price is at high level, all the available marginal land is used for switchgrass cultivation.

Metabolic Adaption of Ethanol-Tolerant Clostridium thermocellum

Clostridium thermocellum is a major candidate for bioethanol production via consolidated bioprocessing. However, the low ethanol tolerance of the organism dramatically impedes its usage in industry. To explore the mechanism of ethanol tolerance in this microorganism, systematic metabolomics was adopted to analyse the metabolic phenotypes of a C. thermocellum wild-type (WT) strain and an ethanol-tolerant strain cultivated without (ET0) or with (ET3) 3% (v/v) exogenous ethanol. Metabolomics analysis elucidated that the levels of numerous metabolites in different pathways were changed for the metabolic adaption of ethanol-tolerant C. thermocellum. The most interesting phenomenon was that cellodextrin was significantly more accumulated in the ethanol-tolerant strain compared with the WT strain, although cellobiose was completely consumed in both the ethanol-tolerant and wild-type strains. These results suggest that the cellodextrin synthesis was active, which might be a potential mechanism for stress resistance. Moreover, the overflow of many intermediate metabolites, which indicates the metabolic imbalance, in the ET0 cultivation was more significant than in the WT and ET3 cultivations. This indicates that the metabolic balance of the ethanol-tolerant strain was adapted better to the condition of ethanol stress. This study provides additional insight into the mechanism of ethanol tolerance and is valuable for further metabolic engineering aimed at higher bioethanol production.

Dilute-Acid Hydrolysis of Apple, Orange, Apricot and Peach Pomaces as Potential Candidates for Bioethanol Production

Chemical composition of four selected fruit pomaces (agro industrial wastes) was evaluated. The effect of temperature, time, acid concentration and solid:liquid (S:L) ratio on dilute-acid hydrolysis of selected pomaces were investigated using 2(4) factorial and central composite design and optimum hydrolysis conditions were determined. A preliminary study was initiated using apple hydrolysate and the fungus Tricoderma harzianum in order to explore and demonstrate their potential uses in bioethanol production. Chemical composition of pomaces was promising as fermentation media. The highest reducing sugar yield from the optimization step was 31%, 49 /0, 56% and 52% for apple, apricot, orange and peach pomaces, respectively under optimum hydrolysis conditions. Neither furfural nor hydroxmethylfurfural (HMF) were detected in hydrolysates. The highest bioethanol production (1.67 g/L) was obtained in 6 days in a non-static incubator using Tricoderma harzianum pre-grown in minimal medium. It was demonstrated that selected pomaces holds potential for bioethanol production and can be eliminated without being potential waste problem to the environment, with economical return. Although, this work is an initial study in showing the potential of these pomaces and T harzianum in bioethanol production, an extensive study on the optimization of fermentation parameters is recommended for further increase of bioethanol production.

Bioethanol fuel production from rotten banana as an environmental waste management and sustainable energy

Banana waste is discarded due to the imperfection during grading process. Banana biomass can be used as raw material to produce bioethanol. In this study, fermentation of banana waste was conducted using Saccharomyces cerevisiae, Type II under anaerobic condition. Production of bioethanol was determined and the effects of various operating conditions which included different temperatures, shaking period, rotten and fresh banana fruit and saccharification method were observed. Overall, the fermented banana fruit waste produced 4.1 to 7.1% bioethanol. The bioethanol yield from mixture of rotten banana fruit increased with increase fermentation period. It is also increased with yeast concentration, using 35% of water at 35°C. The optimum shaking hours for fermentation was 6 h at pH 5.8. Combination of enzyme (pectinase and cellulase) produced higher bioethanol than enzyme alone. Viscosity and acid value of the produced bioethanol followed the ASTM (American Standard for Testing Materials) and EN (European Norms) standards. Fermented banana treated with mixture of enzymes was the best method used for higher bioethanol production. The results showed that, utilization of mixture (skin and pulp) of rotten fruit was more suitable for bioethanol production as renewable energy which could reduce the cost of the initial process. In addition, it did not compete with the consumer food supply and could avoid the overloaded waste for compose, as well as could be used as fuel in the normal petrol engine. In addition to that, energy could be produced from waste banana fruit as environmental recycling process. Key words: Bioethanol, rotten banana, yeast, enzyme, temperature.