Dilute acid pretreatment, enzymatic saccharification and fermentation of wheat straw to ethanol

Wheat straw consists of 48.57 ± 0.30% cellulose and 27.70 ± 0.12% hemicellulose on dry solid (DS) basis and has the potential to serve as a low cost feedstock for production of ethanol. Dilute acid pretreatment at varied temperature and enzymatic saccharification were evaluated for conversion of wheat straw cellulose and hemicellulose to monomeric sugars. The maximum yield of monomeric sugars from wheat straw (7.83%, w/v, DS) by dilute H₂SO₄ (0.75%, v/v) pretreatment and enzymatic saccharification (45 °C, pH 5.0, 72 h) using cellulase, β-glucosidase, xylanase and esterase was 565 ± 10 mg/g. Under this condition, no measurable quantities of furfural and hydroxymethyl furfural were produced. The yield of ethanol (per litre) from acid pretreated enzyme saccharified wheat straw (78.3 g) hydrolyzate by recombinant Escherichia coli strain FBR5 was 19 ± 1 g with a yield of 0.24 g/g DS. Detoxification of the acid and enzyme treated wheat straw hydrolyzate by overliming reduced the fermentation time from 118 to 39 h in the case of separate hydrolysis and fermentation (35 °C, pH 6.5), and increased the ethanol yield from 13 ± 2 to 17 ± 0 g/l and decreased the fermentation time from 136 to 112 h in the case of simultaneous saccharification and fermentation (35 °C, pH 6.0).     

Dilute acid pretreatment of barley straw and its saccharification and fermentation

In this study, the optimization of the major factors for efficient dilute acid pretreatment (DAP) of Korean barley straw was conducted by response surface method (RSM). In addition, saccharification of the optimized pretreated barley straw as well as fermentation of solubilized hemicellulose and enzymatic hydrolysates was performed for bioethanol production. The factors optimized by RSM were concentration of sulfuric acid, reaction time and temperature. Optimization experiments were carried out within the scope of 0.16 ∼ 1.84% sulfuric acid, 10 ∼ 20 min of reaction time, and 116 ∼ 183°C of temperature using a statistical program, and optimal conditions (1.16% of sulfuric acid, 16.9 min of reaction time, and 150°C) were determined based on reliable statistical indicators. The predicted value at stationary point and the experimental value were 81.38 and 80.66%, respectively. Saccharification was performed at 50°C using Celluclast (cellulase) and Novozyme 188 (β-glucosidase) as biocatalysts in an enzyme loading test. Conversion of the saccharification process was approximately 65%. In addition, fermentation of glucose after saccharification and solubilization of xylose solution by DAP were performed using Saccharomyces cerevisiae and Pichia stipitis at 30°C and 200 rpm for 12 h.     

Dilute acid pretreatment and enzymatic saccharification of sugarcane tops for bioethanol production

The aim of this work was to study the feasibility of using sugarcane tops as feedstock for the production of bioethanol. The process involved the pretreatment using acid followed by enzymatic saccharification using cellulases and the process was optimized for various parameters such as biomass loading, enzyme loading, surfactant concentration and incubation time using Box–Behnken design. Under optimum hydrolysis conditions, 0.685 g/g of reducing sugar was produced per gram of pretreated biomass. The fermentation of the hydrolyzate using Saccharomyces cerevisae produced 11.365 g/L of bioethanol with an efficiency of about 50%. This is the first report on utilization of sugarcane tops for bioethanol production.     

Enhanced enzymatic saccharification of rice straw by microwave pretreatment

In this study, Box-Behnken design and response surface methodology were employed to plan experiments and optimize the microwave pretreatment of rice straw. Experimental results show that microwave intensity (MI), irradiation time (IT) and substrate concentration (SC) were main factors governing the enzymatic saccharification of rice straw. The maximal efficiencies of cellulose, hemicellulose and total saccharification were respectively increased by 30.6%, 43.3% and 30.3% under the optimal conditions of MI 680 W, IT 24 min and SC 75 g/L. The chemical composition analysis of straw further confirmed that microwave pretreatment could disrupt the silicified waxy surface, break down the lignin-hemicellulose complex and partially remove silicon and lignin.     

Simultaneous saccharification and fermentation of broken rice: an enzymatic extrusion liquefaction pretreatment for Chinese rice wine production

Broken rice, pretreated by enzymatic extrusion liquefaction, was used to produce Chinese rice wine by simultaneous saccharification and fermentation (SSF) process in this study. The study compared the novel process and traditional process for Chinese rice wine fermentation utilizing broken rice and head rice, respectively. With the optimum extrusion parameters (barrel temperature, 98 °C; moisture content, 42 % and amylase concentration, 1 ‰), 18 % (v/v at 20 °C) alcoholic degree, 37.66 % fermentation recovery and 93.63 % fermentation efficiency were achieved, indicating enzymatic extrusion-processed rice wine from broken rice exhibited much higher fermentation rate and efficiency than traditional-processed rice wine from head rice during SSF. The starch molecule distribution data indicated that the alcoholic degree was related to the oligosaccharides’ formation during enzymatic extrusion. Sum of amino acid (AA) in the extrusion-processed wine was 53.7 % higher than that in the traditional one. These results suggest that the enzymatic extrusion pretreatment for broken rice is a feasible and alternative process in the fermentation of Chinese rice wine.     

Organic acid pretreatment of oil palm trunk: effect on enzymatic saccharification and ethanol production

Bioprocess and Biosystems Engineering - Effective lignocellulosic biomass saccharification is one of the crucial requirements of biofuel production via fermentation process. Organic acid...     

Wet oxidation pretreatment, enzymatic hydrolysis and simultaneous saccharification and fermentation of clover-ryegrass mixtures

The potential of clover (Trifolium repens) and ryegrass (Lolium perenne) mixtures as raw materials for ethanol production was investigated. Wet oxidation, at 175, 185 or 195 degrees C during 10min at two different oxygen pressures and with either addition or no addition of sodium carbonate, was evaluated as pretreatment method for clover-ryegrass mixtures. The enzymatic hydrolysis of cellulose was significantly improved after pretreatment. The highest conversion efficiency, 93.6%, was achieved for the sample pretreated at 195 degrees C, 10min, 1.2MPa and no addition of Na(2)CO(3). For that sample, the overall glucose yield after pretreatment and hydrolysis was 75.5%. No inhibition of cellulose enzymatic conversion by the filtrates was observed. The simultaneous saccharification and fermentation of the pretreated material yielded cellulose conversions of 87.5 and 86.6%, respectively, with Saccharomyces cerevisiae and the filamentous fungus Mucor indicus, and revealed that no addition of nutrients is needed for the fermentation of clover-ryegrass hydrolysates.     

Simultaneous saccharification and fermentation of cellulose: effect of ethanol on enzymatic saccharification of cellulose

It was confirmed that simultaneous saccharification and fermentation are effective for accelerating enzymatic saccharification of cellulose. In this work, the effects of ethanol on the saccharification of tissue paper by Trichoderma cellulase (Meicelase CEPB) have been investigated. The following results were obtained. (1) Saccharification was inhibited by at least 0.2M ethanol. (2) Less than 4M ethanol did not affect the enzymatic activities of beta-glucosidase and endoglucanase (C(x)) at all. The thermal stability of endoglucanase was not also varied by ethanol. (3) It is suggested that ethanol depresses the adsorption of exoglucanase on cellulose. (4) The rate expression of saccharification of cellulose in the presense of ethanol is proposed. (5) The inhibititory effect of ethanol was found to become more significant in the later stages of the reaction than just the initial stage.     

Aqueous ammonia pretreatment, saccharification, and fermentation evaluation of oil palm fronds for ethanol production

Oil palm fronds are the most abundant lignocellulosic biomass in Malaysia. In this study, fronds were tested as the potential renewable biomass for ethanol production. The soaking in aqueous ammonia pretreatment was applied, and the fermentability of pretreated fronds was evaluated using simultaneous saccharification and fermentation. The optimal pretreatment conditions were 7?% (w/w) ammonia, 80?°C, 20?h of pretreatment, and 1:12 S/L ratio, where the enzymatic digestibility was 41.4?% with cellulase of 60?FPU/g-glucan. When increasing the cellulase loading in the hydrolysis of pretreated fronds, the enzymatic digestibility increased until the enzyme loading reached 60?FPU/g-glucan. With 3?% glucan loading in the SSF of pretreated fronds, the ethanol concentration and yield based on the theoretical maximum after 12 and 48?h of the SSF were 7.5 and 9.7?g/L and 43.8 and 56.8?%, respectively. The ethanol productivities found at 12 and 24?h from pretreated fronds were 0.62 and 0.36?g/L/h, respectively.     

Reducing acid in dilute acid pretreatment and the impact on enzymatic saccharification

Dilute acid pretreatment is a leading pretreatment technology for biomass to ethanol conversion due to the comparatively low chemical cost and effective hemicellulose solubilization. The conventional dilute acid pretreatment processes use relatively large quantities of sulfuric acid and require alkali for pH adjustment afterwards. Significant amounts of sulfate salts are generated as by-products, which have to be properly treated before disposal. Wastewater treatment is an expensive, yet indispensable part of commercial level biomass-to-ethanol plants. Therefore, reducing acid use to the lowest level possible would be of great interest to the emerging biomass-to-ethanol industry. In this study, a dilute acid pretreatment process was developed for the pretreatment of corn stover. The pretreatment was conducted at lower acid levels than the conventional process reported in the literature while using longer residence times. The study indicates that a 50% reduction in acid consumption can be achieved without compromising pretreatment efficiency when the pretreatment time was extended from 1–5 min to 15–20 min. To avoid undesirable sugar degradation and inhibitor generation, temperatures should be controlled below 170°C. When the sulfuric acid-to-lignocellulosic biomass ratio was kept at 0.025 g acid/g dry biomass, a cellulose-to-glucose conversion of 72.7% can be achieved at an enzyme loading of 0.016 g/g corn stover. It was also found that acid loading based on total solids (g acid/g dry biomass) governs the pretreatment efficiency rather than the acid concentration (g acid/g pretreatment liquid). While the acid loading on lignocellulosic biomass may be achieved through various combinations of solids loading and acid concentration in the pretreatment step, this work shows that it is unlikely to reduce acid use without undermining pretreatment efficiency simply by increasing the solid content in pretreatment reactors, therefore acid loading on biomass is indicated to be the key factor in effective dilute acid pretreatment.     

Simultaneous enzymatic wheat starch saccharification and fermentation to lactic acid by Lactococcus lactis

Simultaneous saccharification of starch from whole-wheat flour and fermentation to lactic acid (SSF) was investigated. For saccharification the commercial enzyme mixture SAN Super 240 L, having α-amylase, amyloglucosidase and protease activity, was used, and Lactococcus lactis ssp. lactis ATCC 19435 was used for the fermentation. SSF was studied at flour concentrations corresponding to starch concentrations of 90 g/l and 180 g/l and SAN Super concentrations between 3 μl/g and 8 μl/g starch. Kinetic models, developed for the saccharification and fermentation, respectively, were used for simulation and data from SSF experiments were used for model verification. The model simulated SSF when sufficient amounts of nutrients were available during fermentation. This was achieved with high wheat flour concentrations or with addition of yeast extract or amino acids. Nutrient release was dependent on the level of enzyme activity. Received: 26 January 1999 / Accepted: 20 February 1999     

Dilute acid pretreatment of rye straw and bermudagrass for ethanol production

Ethanol production from lignocellulosic materials provides an alternative energy production system. Rye and bermudagrass that are used in hog farms for nutrient uptake from swine wastewater have the potential for fuel ethanol production because they have a relative high cellulose and hemicellulose content. Dilute sulfuric acid pretreatment of rye straw and bermudagrass before enzymatic hydrolysis of cellulose was investigated in this study. The biomass at a solid loading rate of 10% was pretreated at 121 degrees C with different sulfuric acid concentrations (0.6, 0.9, 1.2 and 1.5%, w/w) and residence times (30, 60, and 90 min). Total reducing sugars, arabinose, galactose, glucose, and xylose in the prehydrolyzate were analyzed. In addition, the solid residues were hydrolyzed by cellulases to investigate the enzymatic digestibility. With the increasing acid concentration and residence time, the amount of arabinose and galactose in the filtrates increased. The glucose concentration in the prehydrolyzate of rye straw was not significantly influenced by the sulfuric acid concentration and residence time, but it increased in the prehydrolyzate of bermudagrass with the increase of pretreatment severity. The xylose concentration in the filtrates increased with the increase of sulfuric acid concentration and residence time. Most of the arabinan, galactan and xylan in the biomass were hydrolyzed during the acid pretreatment. Cellulose remaining in the pretreated feedstock was highly digestible by cellulases from Trichoderma reesei.     

Acid-catalyzed steam pretreatment of lodgepole pine and subsequent enzymatic hydrolysis and fermentation to ethanol

Utilization of ethanol produced from biomass has the potential to offset the use of gasoline and reduce CO(2) emissions. This could reduce the effects of global warming, one of which is the current outbreak of epidemic proportions of the mountain pine beetle (MPB) in British Columbia (BC), Canada. The result of this is increasing volumes of dead lodgepole pine with increasingly limited commercial uses. Bioconversion of lodgepole pine to ethanol using SO(2)-catalyzed steam explosion was investigated. The optimum pretreatment condition for this feedstock was determined to be 200 degrees C, 5 min, and 4% SO(2) (w/w). Simultaneous saccharification and fermentation (SSF) of this material provided an overall ethanol yield of 77% of the theoretical yield from raw material based on starting glucan, mannan, and galactan, which corresponds to 244 g ethanol/kg raw material within 30 h. Three conditions representing low (L), medium (M), and high (H) severity were also applied to healthy lodgepole pine. Although the M severity conditions of 200 degrees C, 5 min, and 4% SO(2) were sufficiently robust to pretreat healthy wood, the substrate produced from beetle-killed (BK) wood provided consistently higher ethanol yields after SSF than the other substrates tested. BK lodgepole pine appears to be an excellent candidate for efficient and productive bioconversion to ethanol.     

Effect of dilute acid pretreatment on the saccharification and fermentation of rye straw

This research shows the effect of dilute acid pretreatment with various sulfuric acid concentrations (0.5–2.0% [wt/vol]) on enzymatic saccharification and fermentation yield of rye straw. After pretreatment, solids of rye straw were suspended in Na citrate buffer or post-pretreatment liquids (prehydrolysates) containing sugars liberated after hemicellulose hydrolysis. Saccharification was conducted using enzymes dosage of 15 or 25 FPU/g cellulose. Cellulose saccharification rate after rye straw pretreatment was enhanced by performing enzymatic hydrolysis in sodium citrate buffer in comparison with hemicellulose prehydrolysate. The maximum cellulose saccharification rate (69%) was reached in sodium citrate buffer (biomass pretreated with 2.0% [wt/vol] H₂SO₄). Lignocellulosic complex of rye straw after pretreatment was subjected to separate hydrolysis and fermentation (SHF) or separate hydrolysis and co-fermentation (SHCF). The SHF processes conducted in the sodium citrate buffer using monoculture of Saccharomyces cerevisiae (Ethanol Red) were more efficient compared to hemicellulose prehydrolysate in respect with ethanol yields. Maximum fermentation efficiency of SHF processes obtained after rye straw pretreatment at 1.5% [wt/vol] H₂SO₄ and saccharification using enzymes dosage of 25 FPU/g in sodium citrate buffer, achieving 40.6% of theoretical yield. However, SHCF process using cocultures of pentose-fermenting yeast, after pretreatment of raw material at 1.5% [wt/vol] H₂SO₄ and hydrolysis using enzymes dosage of 25 FPU/g, resulted in the highest ethanol yield among studied methods, achieving 9.4 g/L of ethanol, corresponding to 55% of theoretical yield.     

Optimization of enzymatic hydrolysis and ethanol fermentation from AFEX-treated rice straw

An abundant agricultural residue, rice straw (RS) was pretreated using ammonia fiber expansion (AFEX) process with less than 3% sugar loss. Along with commercial cellulase (Spezyme® CP) at 15 filter paper unit/g of glucan, the addition of Multifect® Xylanase at 2.67 mg protein/g glucan and Multifect® Pectinase at 3.65 mg protein/g glucan was optimized to greatly increase sugar conversion of AFEX-treated RS. During enzymatic hydrolysis even at 6% glucan loading (equivalent to 17.8% solid loading), about 80.6% of glucan and 89.6% of xylan conversions (including monomeric and oligomeric sugars) were achieved. However, oligomeric glucose and xylose accounted for 12.3% of the total glucose and 37.0% of the total xylose, respectively. Comparison among the three ethanologenic strains revealed Saccharomyces cerevisiae 424A(LNH-ST) to be a promising candidate for RS hydrolysate with maximum ethanol metabolic yield of 95.3% and ethanol volumetric productivity of 0.26 g/L/h. The final concentration of ethanol at 37.0 g/L was obtained by S. cerevisiae 424A(LNH-ST) even with low cell density inoculum. A biorefinery combining AFEX pretreatment with S. cerevisiae 424A(LNH-ST) in separate hydrolysis and fermentation could achieve 175.6 g EtOH/kg untreated rice straw at low initial cell density (0.28 g dw/L) without washing pretreated biomass, detoxification, or nutrient supplementation.     

Cost reduction and feedstock diversity for sulfuric acid-free ethanol cooking of lignocellulosic biomass as a pretreatment to enzymatic saccharification

We have previously demonstrated that a sulfuric acid-free ethanol (EtOH) cooking treatment enhances the enzymatic digestibility of eucalyptus wood and bagasse flour. In the present study, a reconfigured process that achieves similar performance was developed by identifying possible cost-competitive pretreatments that provide high cellulose-to-glucose conversion during subsequent enzymatic hydrolysis. The series of reconfigurations reduced EtOH usage in the pretreatment by more than 80% in comparison with our previous research. Higher initial pressures and intensive size reduction of the starting material are not required. The reconfigured process was applied to rice straw and Douglas fir, in order to confirm the feasibility of feedstock diversity.     

Simultaneous saccharification and fermentation of pretreated sugar cane leaves to ethanol

The simultaneous saccharification and fermentation (SSF) of pretreated sugar cane leaves to produce ethanol using a cellulolytic enzyme complex from Trichoderma reesei QM 9414 and Saccharomyces cerevisiae NRRL-Y-132 was optimized. Enzymic saccharification parameters were evaluated prior to SSF studies. A 92% conversion of 2·5% substrate (alkaline hydrogen peroxide pretreated) to sugars was achieved at 50°C and pH 4·5, using T. reesei cellulase (40 FPU/g substrate), in 48 h. The pretreated substrate was then subjected to an SSF process using the cellulase complex and S. cerevisiae cells. Optimization of the SSF system is described.