Acetosolv delignification of Dichrostachys cinerea biomass for ethanol production

The interest in production of fuel ethanol from lignocellulosic materials is continuouslyincreasing due to the urgency of finding non-food substrates for production of bio-fuels.Marabou (Dichrostachys cinerea) is one of the abundant lignocellulosic bio-resources in Cuba,and it could be useful to produce bio-ethanol. Pre-treatment is an important step to produceethanol from lignocellulosic materials since it allows the separation of cellulose, hemicelluloseand lignin, and activates cellulose towards enzymatic hydrolysis. During the past few years,organosolv methods have been reported for effective separation of the main components oflignocellulosic materials and improvement of the enzymatic hydrolysis of cellulose. By usingacetosolv method lignin is separated under mild conditions and many of the lignin properties arewell preserved.The present work was aimed to perform a chemical characterisation of marabou biomass and toevaluate acetosolv delignification of the material. In this work the content of moisture, ash,extractives, easy-to-hydrolyze polysaccharides, difficult-to-hydrolyze polysaccharides, andKlason lignin of marabou biomass were analyzed. Klason lignin of the marabou biomass was23.4% of the mass. Acetosolv delignification was performed at normal boiling temperature(NBT) and 121oC, using 50-50, 70-30 and 90-10 acetic acid – water mixtures with 10% of solidsload during 1h. Hydrochloric acid (0.2g / 100g of mixture) was used as catalyst. Thedelignification of marabou biomass was also evaluated for the combination of dilute acid prehydrolysis(DAPH) and acetosolv with the same reaction conditions. This investigation provedthat acetosolv pretreatment was effective for solubilizing lignin contained in marabou biomass.The degree of lignin solubilisation increased with increasing acetic acid concentration in thereaction mixture. Lignin removals above 80% were achieved consistently both at NBT and121oC with 90% acetic acid, while only around 44.6 and 6.8% of the initial lignin was removedusing, respectively, 70 and 50% acetic acid at 121oC. The effect of temperature ondelignification was only marginal when acetosolv was conducted with 90% acetic acid, but itwas remarkable for lower acetic acid concentrations. A two-fold decrease of lignin removal wasobserved for the NBT acetosolv compared with the process performed at 121oC using both 70and 50% acetic acid. The insertion of a DAPH step prior to acetosolv considerably improvedlignin removal using 70 and 50% acetic acid at both temperatures, but its effect on the processesusing 90% acetic acid was minimal. High lignin yields were achieved upon its precipitation fromacetosolv liquors

Acetosolv delignification of Dichrostachys cinerea biomass for ethanol production

The interest in production of fuel ethanol from lignocellulosic materials is continuouslyincreasing due to the urgency of finding non-food substrates for production of bio-fuels.Marabou (Dichrostachys cinerea) is one of the abundant lignocellulosic bio-resources in Cuba,and it could be useful to produce bio-ethanol. Pre-treatment is an important step to produceethanol from lignocellulosic materials since it allows the separation of cellulose, hemicelluloseand lignin, and activates cellulose towards enzymatic hydrolysis. During the past few years,organosolv methods have been reported for effective separation of the main components oflignocellulosic materials and improvement of the enzymatic hydrolysis of cellulose. By usingacetosolv method lignin is separated under mild conditions and many of the lignin properties arewell preserved.The present work was aimed to perform a chemical characterisation of marabou biomass and toevaluate acetosolv delignification of the material. In this work the content of moisture, ash,extractives, easy-to-hydrolyze polysaccharides, difficult-to-hydrolyze polysaccharides, andKlason lignin of marabou biomass were analyzed. Klason lignin of the marabou biomass was23.4% of the mass. Acetosolv delignification was performed at normal boiling temperature(NBT) and 121oC, using 50-50, 70-30 and 90-10 acetic acid – water mixtures with 10% of solidsload during 1h. Hydrochloric acid (0.2g / 100g of mixture) was used as catalyst. Thedelignification of marabou biomass was also evaluated for the combination of dilute acid prehydrolysis(DAPH) and acetosolv with the same reaction conditions. This investigation provedthat acetosolv pretreatment was effective for solubilizing lignin contained in marabou biomass.The degree of lignin solubilisation increased with increasing acetic acid concentration in thereaction mixture. Lignin removals above 80% were achieved consistently both at NBT and121oC with 90% acetic acid, while only around 44.6 and 6.8% of the initial lignin was removedusing, respectively, 70 and 50% acetic acid at 121oC. The effect of temperature ondelignification was only marginal when acetosolv was conducted with 90% acetic acid, but itwas remarkable for lower acetic acid concentrations. A two-fold decrease of lignin removal wasobserved for the NBT acetosolv compared with the process performed at 121oC using both 70and 50% acetic acid. The insertion of a DAPH step prior to acetosolv considerably improvedlignin removal using 70 and 50% acetic acid at both temperatures, but its effect on the processesusing 90% acetic acid was minimal. High lignin yields were achieved upon its precipitation fromacetosolv liquors

Biochemical conversion of biomass to biofuels : pretreatment–detoxification–hydrolysis–fermentation

The use of lignocellulosic materials to replace fossil resources for the industrial production of fuels, chemicals, and materials is increasing. The carbohydrate composition of lignocellulose (i.e. cellulose and hemicellulose) is an abundant source of sugars. However, due to the feedstock recalcitrance, rigid and compact structure of plant cell walls, access to polysaccharides is hindered and release of fermentable sugars has become a bottle-neck. Thus, to overcome the recalcitrant barriers, thermochemical pretreatment with an acid catalyst is usually employed for the physical or chemical disruption of plant cell wall. After pretreatment, enzymatic hydrolysis is the preferred option to produce sugars that can be further converted into liquid fuels (e.g. ethanol) via fermentation by microbial biocatalysts. However, during acid pretreatment, several inhibitory compounds namely furfural, 5-hydroxymethyl furfural, phenols, and aliphatic acids are released from the lignocellulose components. The presence of these compounds can greatly effect both enzymatic hydrolysis and microbial fermentation. For instance, when Avicel cellulose and acid treated spruce wood hydrolysate were mixed, 63% decrease in the enzymatic hydrolysis efficiency was observed compared to when Avicel was hydrolyzed in aqueous citrate buffer. In addition, the acid hydrolysates were essentially non-fermentable. Therefore, the associated problems of lignocellulose conversion can be addressed either by using feedstocks that are less recalcitrant or by developing efficient pretreatment techniques that do not cause formation of inhibitory byproducts and simultaneously give high sugar yields. A variety of lignocellulose materials including woody substrates (spruce, pine, and birch), agricultural residues (sugarcane bagasse and reed canary grass), bark (pine bark), and transgenic aspens were evaluated for their saccharification potential. Apparently, woody substrates were more recalcitrant than the rest of the species and bark was essentially amorphous. However, the saccharification efficiency of these substrates varied based on the pretreatment method used. For instance, untreated reed canary grass was more recalcitrant than woody materials whereas the acid treated reed canary grass gave a higher sugar yield (64%) than the woody substrates (max 34%). Genetic modification of plants was beneficial, since under similar pretreatment and enzymatic hydrolysis conditions, up to 28% higher sugar production was achieved from the transgenic plants compare to the wild type. As an alternative to the commonly used acid catalysed pretreatments (prior to enzymatic hydrolysis) lignocellulose materials were treated with four ionic liquid solvents (ILs): two switchable ILs (SILs) -SO2DBUMEASIL and CO2DBUMEASIL, and two other ILs [Amim][HCO2] and [AMMorp][OAc]. viii After enzymatic hydrolysis of IL treated substrates, a maximum amount of glucan to glucose conversion of between 75% and 97% and a maximum total sugar yields of between 71% and 94% were obtained. When using acid pretreatment these values varied between 13-77% for glucan to glucose conversion and 26-83% for total sugar yield. For woody substrates, the hemicellulose recovery (max 92%) was higher for the IL treated substrates than compared to acid treated samples. However, in case of reed canary grass and pine bark the hemicellulose recovery (90% and 88%, respectively) was significantly higher for the acid treated substrates than the IL treated samples. To overcome the inhibitory problems associated with the lignocellulose hydrolysates, three chemical conditioning methods were used 1. detoxification with ferrous sulfate (FeSO4) and hydrogen peroxide (H2O2) 2. application of reducing agents (sulfite, dithionite, or dithiothreitol) and 3. treatment with alkali: Ca(OH)2, NaOH, and NH4OH. The concentrations of inhibitory compounds were significantly lower after treatments with FeSO4 and H2O2 or alkali. Using reducing agents did not cause any decrease in the concentration of inhibitors, but detoxification of spruce acid hydrolysates resulted in up to 54% improvement of the hydrolysis efficiency (in terms of sugar release) compared to untreated samples. On the other hand, application of detoxification procedures to the aqueous buffer resulted in up to 39% decrease in hydrolysis efficiency, thus confirming that the positive effect of detoxification was due to the chemical alteration of inhibitory compounds. In addition, the fermentability of detoxified hydrolysates were investigated using the yeast Saccharomyces cerevisiae. The detoxified hydrolysates were readily fermented to ethanol yielding a maximum ethanol concentration of 8.3 g/l while the undetoxified hydrolysates were basically non-fermentable

Methods for improvement of enzymatic hydrolysis of lignocellullosic material

The present invention relates to a method of enzymatic hydrolysis of a lignocellulosic material, comprising the steps of: a) pretreating the lignocellulosic material to obtain a slurry having a pH of less than 6; b) adding NaOH, Ca(OH)2 and/or CaO to the slurry to increase its pH to at least 8, said addition being carried out at a slurry temperature of at least 60 °C; c) reducing the pH of the slurry to below 7; and optionally cooling the slurry from step b) to a temperature below 60 °C; and d) adding hydrolytic enzymes to the slurry from c) and allowing the slurry to hydrolyze wherein no washing of the slurry is performed prior to step d)WO 2013/000696 A1, PCT/EP2012/061021</p

Coupled Enzymatic Hydrolysis and Ethanol Fermentation : Ionic Liquid Pretreatment for Enhanced Yields

Background Pretreatment is a vital step upon biochemical conversion of lignocellulose materials into biofuels. An acid catalyzed thermochemical treatment is the most commonly employed method for this purpose. Alternatively, ionic liquids (ILs), a class of neoteric solvents, provide unique opportunities as solvents for the pretreatment of a wide range of lignocellulose materials. In the present study, four ionic liquid solvents (ILs), two switchable ILs (SILs) DBU–MEA–SO 2 and DBU–MEA–CO 2 , as well as two ‘classical’ ILs [Amim][HCO 2 ] and [AMMorp][OAc], were applied in the pretreatment of five different lignocellulosic materials: Spruce (Picea abies) wood, Pine (Pinus sylvestris) stem wood, Birch (Betula pendula) wood, Reed canary grass (RCG, Phalaris arundinacea), and Pine bark. Pure cellulosic substrate, Avicel, was also included in the study. The investigations were carried out in comparison to acid pretreatments. The efficiency of different pretreatments was then evaluated in terms of sugar release and ethanol fermentation. Results Excellent glucan-to-glucose conversion levels (between 75 and 97 %, depending on the biomass and pretreatment process applied) were obtained after the enzymatic hydrolysis of IL-treated substrates. This corresponded between 13 and 77 % for the combined acid treatment and enzymatic hydrolysis. With the exception of 77 % for pine bark, the glucan conversions for the non-treated lignocelluloses were much lower. Upon enzymatic hydrolysis of IL-treated lignocelluloses, a maximum of 92 % hemicelluloses were also released. As expected, the ethanol production upon fermentation of hydrolysates reflected their sugar concentrations, respectively. Conclusions Utilization of various ILs as pretreatment solvents for different lignocelluloses was explored. SIL DBU–MEA–SO 2 was found to be superior solvent for the pretreatment of lignocelluloses, especially in case of softwood substrates (i.e., spruce and pine). In case of birch and RCG, the hydrolysis efficiency of the SIL DBU–MEA–CO 2 was similar or even better than that of DBU–MEA–SO 2 . Further, the IL [AMMorp][OAc] was found as comparably efficient as DBU–MEA–CO 2. Pine bark was highly amorphous and none of the pretreatments applied resulted in clear benefits to improve the product yields.Originally included in thesis in manuscript form.</p