Steam explosion of straw in batch and continuous systems

The effects of the steam-explosion treatment on aqueous fractionation and bioconversion of wheat straw have been investigated. The treatments have been carried out in batch and continuous reactors with capacity of 0.5 Kg/cycle and 150 Kg/h, respectively. The exploded materials have been sequentially extracted with water at 65°C and sodium hydroxide 1.5%. Analytical determinations of liquid fractions and solid residues haveled to the fractionation pattern of the carbohydrates as, monomers, oligomers, and polymers. Evaluations of total acidity, ash content, and lignin recovery have improved understanding of the process. This part of the work has allowed us to derive: the empiric relationship between the batch and the continuous reactors and the yield and availability of pentoses and hexoses in various phases. Selected samples have been tested in enzymatic-hydrolysis experiments, pointing out the effect of treatment severity and reactor used on the saccharification yield.     

Optimization of steam pretreatment of corn stover to enhance enzymatic digestibility

Among the available agricultural byproducts, corn stover, with its yearly production of 10 million t (dry basis), is the most abundant promising raw material for fuel ethanol production in Hungary. In the United States, more than 216 million to fcorn stover is produced annually, of which a portion also could possibly be collected for conversion to ethanol. However, a network of lignin and hemicellulose protects cellulose, which is the major source of fermentable sugars in corn stover (approx 40% of the dry matter [DM]). Steam pretreatment removes the major part of the hemicellulose from the solid material and makes the cellulose more susceptible to enzymatic digestion. We studied 12 different combinations of reaction temperature, time, and pH during steam pretreatment. The best conditions (200°C, 5 min, 2% H₂SO₄) increased the enzymatic conversion (from cellulose to glucose) of corn stover more then four times, compared to untreated material. However, steam pretreatment at 190°C for 5 min with 2% sulfuric acid resulted in the highest overall yield of sugars, 56.1 g from 100 g of untreated material (DM), corresponding to 73% of the theoretical. The liquor following steam explosion was fermented using Saccharomyces cerevisiae to investigate the inhibitory effect of the pretreatment. The achieved ethanol yield was slightly higher than that obtained with a reference sugar solution. This demonstrates that baker's yeast could adapt to the pretreated liquor and ferment the glucose to ethanol efficiently.     

Combined steam pretreatment and enzymatic hydrolysis of starch-free wheat fibers

Steam treatment of an industrial process stream, denoted starch-free wheat fiber, was investigated to improve the formation of monomeric sugars in subsequent enzymatic hydrolysis for further bioconversion into ethanol. The solid fraction in the process stream, derived from a combined starch and ethanol factory, was rich in arabinose (21.1%), xylose (30.1%), and glucose (18.6%), in the form of polysaccharides. Various conditions of steam pretreatment (170–220°C for 5–30 min) were evaluated, and their effect was assessed by enzymatic hydrolysis with 2 g of Celluclast + Ultraflo mixture/ 100 g of starch-free fiber (SFF) slurry at 5% dry matter (DM). The highest overall sugar yield for the combined steam pretreatment and enzymatic hydrolysis, 52g/100 g of DM of SFF, corresponding to 74% of the theoretical, was achieved with pretreatment at 190°C for 10 min followed by enzymatic hydrolysis.     

Effect of chip size on steam explosion pretreatment of softwood

Although considerable progress has been made in technology for converting lignocellulosic biomass into ethanol, substantial opportunities still exist toreduce production costs. In biomass pretreatment, reducing milling power is a technological improvement that will substantially lower production costs for ethanol. Improving sugar yield from hemicellulose hydrolysis would also reduce ethanol production costs. Thus, it would be desirable to test innovative pretreatment conditions to improve the economics by reducing electrical power of the milling stage and by optimizing pretreatment recovery of hemicellulose, as well as to enhance cellulose hydrolysis. The objective of this study was to evaluate the effect of chip size (2–5, 5–8, and 8–12 mm) on steam-explosion pretreatment (190 and 210°C, 4 and 8 min) of softwood (Pinus pinater).     

Investigation of enzymatic hydrolysis of lipid-like substrates in monolayers

The parameters of enzymatic hydrolysis of novel lipid-like substrates assembled in monolayers at water–air interface were estimated by a simple method. The method is based on measurement of the initial velocity of the reaction registered by the decrease of the monolayer area (caused by the enzymatic hydrolysis) at surface pressure of 10 mN/m in a single–compartment trough. Hydrolysis of trilaurin and three 1,3-dilaurylpseudoglycerides acylated by phenylalanine, leucine and valine was characterized by catalytic constants k_cat and apparent Michael's constants K_m(app) (using lipase from Pseudomonas fluorescens as catalyst). It was found that k_cat of the synthetic pseudoglycerides (7–13 per s) are higher than k_cat of trilaurin (4 per s) that can be explained by the presence of positively charged primary aminogroups in the substrates. K_m(app) values were found to be similar for all the substrates studied (ca. 2×10⁻⁶ M). The proposed method allows estimation of the kinetic constants in traditional dimensions.     

Enhancing the enzymatic hydrolysis of cellulosic materials using simultaneous ball milling

One of the limiting factors restricting the effective and efficient bioconversion of softwood-derived lignocellulosic residues is the recalcitrance of the substrate following pretreatment. Consequently, the ensuing enzymatic process requires relatively high enzyme loadings to produce monomeric carbohydrates that are readily fermentable by ethanologenic microorganisms. In an attempt to circumvent the need for larger enzyme loadings, a simultaneous physical and enzymatic hydrolysis treatment was evaluated. A ball-mill reactor was used as the digestion vessel, and the extent and rate of hydrolysis were monitored. Concurrently, enzyme adsorption profiles and the rate of conversion during the course of hydrolysis were monitored. α-Cellulose, employed as a model substrate, and SO₂-impregnated steam-exploded Douglas-fir wood chips were assessed as the cellulosic substrates. The softwood-derived substrate was further posttreated with water and hot alkaline hydrogen peroxide to remove >90% of the original lignin. Experiments at different reaction conditions were evaluated, including substrate concentration, enzyme loading, reaction volumes, and number of ball beads employed during mechanical milling. It was apparent that the best conditions for the enzymatic hydrolysis of α-cellulose were attained using a higher number of beads, while the presence of air-liquid interface did not seem to affect the rate of saccharification. Similarly, when employing the lignocellulosic substrate, up to 100% hydrolysis could be achieved with a minimum enzyme loading (10 filter paper units/g of cellulose), at lower substrate concentrations and with a greater number of reaction beads during milling. It was apparent that the combined strategy of simultaneous ball milling and enzymatic hydrolysis could improve the rate of saccharification and/or reduce the enzyme loading required to attain total hydrolysis of the carbohydrate moieties.     

Strategies to enhance the enzymatic hydrolysis of pretreated softwood with high residual lignin content

Pretreatment of Douglas-fir by steam explosion produces a substrate containing approx 43% lignin. Two strategies were investigated for reducing the effect of this residual lignin on enzymatic hydrolysis of cellulose: mild alkali extraction and protein addition. Extraction with cold 1% NaOH reduced the lignin content by only approx 7%, but cellulose to glucose conversion was enhanced by about 30%. Before alkali extraction, addition of exogenous protein resulted in a significant improvement in cellulose hydrolysis, but this protein effect was substantially diminished after alkali treatment. Lignin appears to reduce cellulose hydrolysis by two distinct mechanisms: by forming a physical barrier that prevents enzyme access and by non-productively binding cellulolytic enzymes. Cold alkali appears to selectively remove a fraction of lignin from steam-exploded Douglas-fir with high affinity for protein. Corresponding data for mixed softwood pretreated by organosolv extraction indicates that the relative importance of the two mechanisms by which residual lignin affects hydrolysis is different according to the pre- and post-treatment method used.     

Steam pretreatment of acid-sprayed and acid-soaked barley straw for production of ethanol

Barley is an abundant crop in Europe, which makes its straw residues an interesting cellulose source for ethanol production. Steam pretreatment of the straw followed by enzymatic hydrolysis converts the cellulose to fermentable sugars. Prior to pretreatment the material is impregnated with a catalyst, for example, H₂SO₄, to enhance enzymatic digestibility of the pretreated straw. Different impregnation techniques can be applied. In this study, soaking and spraying were investigated and compared at the same pretreatment condition in terms of overall yield of glucose and xylose. The overall yield includes the soluble sugars in the liquid from pretreatment, including soluble oligomers, and monomer sugars obtained in the enzymatic hydrolysis. The yields obtained differed for the impregnation techniques. Acid-soaked barley straw gave the highest overall yield of glucose, regardless of impregnation time (10 or 30 min) or acid concentration (0.2 or 1.0 wt%). For xylose, soaking gave the highest overall yield at 0.2 wt% H₂SO₄. An increase in acid concentration resulted in a decrease in xylose yield for both acid-soaked and acid-sprayed barley straw. Optimization of the pretreatment conditions for acid-sprayed barley straw was performed to obtain yields using spraying that were as high as those with soaking. For acid-sprayed barley straw the optimum pretreatment condition for glucose, 1.0 wt% H₂SO₄ and 220°C for 5 min, gave an overall glucose yield of 92% of theoretical based on the composition of the raw material. Pretreatment with 0.2wt% H₂SO₄ at 190°C for 5 min resulted in the highest overall xylose yield, 67% of theoretical based on the composition of the raw material.     

Effect of acid steam explosion on enzymatic hydrolysis ofO. nervosum andC. cardunculus

Applied Biochemistry and Biotechnology -     

Effect of pretreatment reagent and hydrogen peroxide on enzymatic hydrolysis of oak in percolation process

The effect of pretreatment reagent and hydrogen peroxide on enzymatic digestibility of oak was investigated to compare pretreatment performance. Pretreatment reagents used were ammonia, sulfuric acid, and water. These solutions were used without or in combination with hydrogen peroxide in the percolation reactor. The reaction was carried out at 170°C for the predetermined reaction time. Ammonia treatment showed the highest delignification but the lowest digestibility and hemicellulose removal among the three treatments. Acid treatment proved to be a very effective method in terms of hemicellulose recovery and cellulose digestibility. Hemicellulose recovery was 65–90% and digestibilities were >90% in the range of 0.01–0.2% acid concentration. In both treatments, hydrogen peroxide had some effect on digestibility but decomposed soluble sugars produced during pretreatment. Unlike ammonia and acid treatments, hydrogen peroxide in water treatment has a certain effect on hemicellulose recovery as well as delignification. At 1.6% hydrogen peroxide concentration, both hemicellulose recovery and digestibility were about 90%, which were almost the same as those of 0.2% sulfuric acid treatment. Also, digestibility was investigated as a function of hemicellulose removal or delignification. It was found that digestibility was more directly related to hemicellulose removal rather than delignification.     

Evaluation of wet oxidation pretreatment for enzymatic hydrolysis of softwood

The wet oxidation pretreatment (water, oxygen, elevated temperature, and pressure) of softwood (Picea abies) was investigated for enhancing enzymatic hydrolysis. The pretreatment was preliminarily optimized. Six different combinations of reaction time, temperature, and pH were applied, and the compositions of solid and liquid fractions were analyzed. The solid fraction after wet oxidation contained 58–64% cellulose, 2–16% hemicellulose, and 24–30% lignin. The pretreatment series gave information about the roles of lignin and hemicellulose in the enzymatic hydrolysis. The temperature of the pretreatment, the residual hemicellulose content of the substrate, and the type of the commercial cellulase preparation used were the most important factors affecting the enzymatic hydrolysis. The highest sugar yield in a 72-h hydrolysis, 79% of theoretical, was obtained using a pretreatment of 200°C for 10 min at neutral pH.     

Effect of alkaline pretreatment on delignification of wheat straw

This study was conducted to analyse structural changes through scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) after alkaline pretreatment of wheat straw for optimum steaming period. During the study, 2 mm size of substrate was soaked in 2.5% NaOH for 1 h at room temperature and then autoclaved at 121°C for various steaming time (30, 60, 90 and 120 min). Results revealed that residence time of 90 min at 121°C has strong effect on substrate, achieving a maximum cellulose content of 83%, delignification of 81% and hemicellulose content of 10.5%. Further SEM and FTIR spectroscopy confirmed structural modification caused by alkaline pretreatment in substrate. Maximum saccharification yield of 52.93% was achieved with 0.5% enzyme concentration using 2.5% substrate concentration for 8 h of incubation at 50°C. This result indicates that the above-mentioned pretreatment conditions create accessible areas for enzymatic hydrolysis.     

The determination of phenol in urine by enzymatic hydrolysis/headspace gas chromatography

Summary A new method has been developed for the determination of phenol in urine in which the phenol conjugates are hydrolysed enzymatically and the liberated phenol is analysed by headspace chromatography. The results compare favourably with those obtained by the method of van Haaften and Sie in which acid hydrolysis is carried out in a heated GC precolumn. The enzymatic hydrolysis headspace technique appears to be very reliable and does not suffer from the disadvantages of precolumn acid hydrolysis.Since this paper was submitted, we have found a report on the enzymatic hydrolysis of phenyl glucuronide in urine followed by gas chromatographic analysis which seems to have been overlooked by later workers. The paper is by H. Desoille, T. F. Gandenzi-Britto and M. Philbert in Archives des Maladies Professionelles, de Medecine du Travail et de Securite Sociale (Paris),30 (3), 129 (1969).     

On the enzymatic hydrolysis of methyl 2-fluoro-2-arylpropionates by lipases

The enzymatic hydrolysis of methyl 2-fluoro-2-arylpropionates was performed using lipases from Candida rugosa and Candida cylindracea (OF-360). A careful analysis of the reaction products revealed that racemic 2-hydroxy-2-arylpropionic acid and traces of 2-arylacrylic acid are formed, in addition to the expected 2-aryl-2-fluoropropionic acid. The presence of powerful electron-releasing groups in the aromatic ring of the substrate increase the amount of 2-hydroxypropionic acid. A mechanistic hypothesis has been formulated according to which the enzyme facilitates the elimination of fluoride ion from the hydrolysed acid with the formation of an α-carboxy-stabilized carbocation which provides 2-hydroxypropionic acids by nucleophilic attack of H₂O and 2-arylacrylic acids by a β-elimination process.     

Optimization of steam explosion to enhance hemicellulose recovery and enzymatic hydrolysis of cellulose in softwoods

A combination of Douglas fir heartwood and sapwood chips were steam pretreated under three conditions as measured by the Severity Factor (log R_o), which incorporated the time, temperature/pressure of pretreatment. By adjusting the steam pretreatment conditions, it was hoped to recover the majority of the hemicellulose component as monomers in the water-soluble stream, while providing a cellulosic-rich, water-insoluble fraction that could be readily hydrolyzed by cellulases. These three conditions were chosen to represent either high hemicellulose sugar recovery (low severity [L], log R_o=3.08), high-enzyme hydrolyzability of the cellulosic component (high severity [H], log R_o=4.21), and a compromise between the two conditions (medium severity [M], log R_o=3.45). The medium-severity pretreatment conditions (195°C, 4.5 min, 4.5% SO₂ logR_o=3.45) gave the best compromise in terms of relatively high hemicellulose recovery after stream pretreatment and the subsequent efficiency of enzymatic hydrolysis of the water-insoluble cellulosic fraction. The percent recovery of the original hemicellulose in the water-soluble fraction dropped significantly when the severity was increased (L-76.8%, M-64.7%, and H-37.5%). However, the ease of enzymatic hydrolysis of the cellulose-rich, water-insoluble fraction increased with increasing severity (L-24%, M-86.6%, and H-97.9%). Although more severe pretreatment conditions provided optimum hydrolysis of the cellulosic component, less severe conditions resulted in better recovery of the combined hemicellulose and cellulosic components.     

Nanocomposites of aliphatic polyesters: An overview of the effect of different nanofillers on enzymatic hydrolysis and biodegradation of polyesters

In the present review the findings concerning the effect of nanofillers to biodegradation and enzymatic hydrolysis of aliphatic polyesters were summarized and discussed. Most of the published works are dealing with the effect of layered silicates such as montmorillonite (unmodified and modified with organic compounds), carbon nanotubes and spherical shape additives like SiO₂ and TiO₂. The degradation of polyester due to the enzymatic hydrolysis is a complex process involving different phenomena, namely, water absorption from the polyesters, enzymatic attack to the polyester surface, ester cleavage, formation of oligomer fragments due to endo- or exo-type hydrolysis, solubilization of oligomer fragments in the surrounding environment, diffusion of soluble oligomers by bacteria and finally consumption of the oligomers and formation of CO₂ and H₂O. By studying the published works in nanocomposites, different and sometimes contradictory results have been reported concerning the effect of the nanofillers on aliphatic polyesters biodegradation. Most of the papers suggested that the addition of nanofillers provokes a substantial enhancement of polyester hydrolysis due to the catalyzing effect of the existed reactive groups (–OH and –COOH), to the crystallinity decrease, to the higher hydrophilicity of nanofillers and thus higher water uptake, to the higher interactions, etc. However, there are also some papers that suggested a delay effect of nanofillers to the polyesters degradation mainly due to the barrier effect of nanofillers and the lower available surface for enzymatic hydrolysis.     

Enhanced enzymatic hydrolysis of steam-exploded douglas fir wood by alkali-oxygen post-treatment

Good enzymatic hydrolysis of steam-exploded Douglas fir wood (SEDW) cannot be achieved owing to the very high lignin content (>40%) that remains associated with this substrate. Thus, in this study, we investigated the use of alkali-oxygen treatment as a posttreatment to delignify SEDW and also considered the enzymatic hydrolyzability of the delignified SEDW. The results showed that under optimized conditions of 15% NaOH, 5% consistency, 110°C, and 3h, approx84% of the lignin in SEDW could be removed. The resulting delignified SEDW had good hydrolyzability, and cellulose-to-glucose conversion yields of over 90 and 100% could be achieved within 48 h with 20 and 40 filter paper units/g of cellulose enzyme loadings, respectively. It was also indicated that severe conditions, such as high NaOH concentration and high temperature, should not be utilized in oxygen delignification of SEDW in order to avoid extensive condensation of lignin and significant degradation of cellulose.     

超临界二氧化碳偶合超声预处理强化玉米秸秆酶水解(英)

提出了一个木质纤维素生物质预处理的全绿色加工过程.以玉米秸秆和玉米芯为原料,以超临界CO₂和超声偶合法对木质纤维素进行预处理.超临界CO₂预处理条件为:压力15-25 MPa,温度120₁70℃,含水量50%,反应时间0.5₄ h.超声场功率600W,温度80℃,作用时间2-8 h.用纤维素酶水解反应获得的还原糖总量来评价预处理效果.结果表明,单纯超临界CO₂和超临界CO₂偶合超声预处理都能够提高生物质水解反应还原糖产量.对于玉米芯,超临界CO₂预处理（170℃,20 MPa,3 0min）后,还原糖产率为62%（未预处理的为12%）.对于玉米秸秆（170℃,20 MPa,2.5 h）,还原糖产率为46.4%.对于玉米芯,超临界CO₂偶合超声预处理（600 W,80℃下超声处理6 h,然后用170℃,20 MPa超临界CO₂预处理30 min）后,还原糖产率为87%.对于玉米秸秆,超临界CO₂偶合超声预处理（600 W,80℃下超声处理8 h,然后用170℃,20 MPa超临界CO₂预处理1 h）后,还原糖产率为25.5%.与未处理生物质相比,X射线衍射结果表明玉米秸秆和玉米芯在超临界CO₂和超声预处理后其结晶度没有明显变化.扫描电镜分析则发现木质纤维素的表面积显著增加.     

复合酶耦合酶解酸处理后玉米秸秆的研究

以玉米秸秆为研究对象,经过2%硫酸预处理后,利用果胶酶、β-葡萄糖苷酶、纤维素酶三种酶协同酶解,以提高玉米秸秆的酶解产糖量。结果表明:当酶解时间为48h,果胶酶、β-葡萄糖苷酶、纤维素酶分别为45U/mL、30U/mL、60U/mL时,葡萄糖、木糖和酶水解得率分别为67.83%、3.25%、73.65%,相比纤维素酶单一酶解的葡萄糖、木糖和酶水解得率分别提高了65.04%、20.82%、65.06%。分步糖化发酵5天后,相比单一酶解发酵乙醇含量提高了72.5%。说明利用三种酶复合处理,能明显提高酶解产糖量。研究结果为玉米秸秆转化为可发酵糖技术的研究提供重要参考。     

底物水解预处理对玉米秸秆发酵生物产氢的影响

水解预处理是影响纤维索类生物质发酵产氢效率的关键因素之一.在批式试验条件下,分别采用乳酸处理(方法A),生物处理(方法B)和生物/乳酸两步处理(方法C)方法对玉米秸秆进行糖化水解预处理;考察了水解预处理对产氢效率的影响.结果表明:经乳酸预处理、生物预处理和乳酸/生物两步处理的玉米秸秆的累积氢产量分别为132 mL/g,...