Background:
Simultaneous saccharification and co-fermentation (SSCF) has been recognized as a feasible option for ethanol production from xylose-rich lignocellulosic materials. To reach high ethanol concentration in the broth, a high content of water-insoluble solids (WIS) is needed, which creates mixing problems and, furthermore, may decrease xylose uptake. Feeding of substrate has already been proven to give a higher xylose conversion than a batch SSCF. In the current work, enzyme feeding, in addition to substrate feeding, was investigated as a means of enabling a higher WIS content with a high xylose conversion in SSCF of a xylose-rich material. A recombinant xylose-fermenting strain of Saccharomyces cerevisiae (TMB3400) was used for this purpose in fed-batch SSCF experiments of steam-pretreated wheat straw.
Results:
By using both enzyme and substrate feeding, the xylose conversion in SSCF could be increased from 40% to 50% in comparison to substrate feeding only. In addition, by this design of the feeding strategy, it was possible to process a WIS content corresponding to 11% in SSCF and obtain an ethanol yield on fermentable sugars of 0.35 g g-1.
Conclusion:
A combination of enzyme and substrate feeding was shown to enhance xylose uptake by yeast and increase overall ethanol yield in SSCF. This is conceptually important for the design of novel SSCF processes aiming at high-ethanol titers. Substrate feeding prevents viscosity from becoming too high and thereby allows a higher total amount of WIS to be added in the process. The enzyme feeding, furthermore, enables keeping the glucose concentration low, which kinetically favors xylose uptake and results in a higher xylose conversion.
Mobile phone recycling is a fast increasing notion that is taking the world by storm, and a way of earning yourself some additional money. Even so, people across the world are still very much unacquainted towards the potential behind this new cash earning arrangement.
Should you or any person you know have any old mobile phones lying around their abode then now is the best time to see how much they could be worth by using a mobile phone recycling comparison website like SellMyMobile.com, and the course of action couldn’t be any simpler. When you come to sell your mobile, you can calculate all the UK’s leading mobile phone recycle companies to give you the utmost price possible on your broken, unused or even new mobile phone, saving you time and cash from having to actually visit each web site and take note of the prices.
The course of action to selling your phones for money is clear-cut and will literally only take you a matter of minutes to finalize. To sell mobile handsets simply search for your phone, look out for the best price label and send it in using information that will be emailed across to you. It doesn’t cost you a penny and you will obtain the money for it within the post within several working days. Help declutter your house, get yourself some extra money, and help the environment by recycling your mobile phones with SellMyMobile.com.
Background:
Bioethanol can be produced from sugar-rich, starch-rich (first generation; 1G) or lignocellulosic (second generation; 2G) raw materials. Integration of 2G ethanol with 1G could facilitate the introduction of the 2G technology. The capital cost per ton of fuel produced would be diminished and better utilization of the biomass can be achieved. It would, furthermore, decrease the energy demand of 2G ethanol production and also provide both 1G and 2G plants with heat and electricity. In the current study, steam-pretreated wheat straw (SPWS) was mixed with presaccharified wheat meal (PWM) and converted to ethanol in simultaneous saccharification and fermentation (SSF).
Results:
Both the ethanol concentration and the ethanol yield increased with increasing amounts of PWM in mixtures with SPWS. The maximum ethanol yield (99% of the theoretical yield, based on the available C6 sugars) was obtained with a mixture of SPWS containing 2.5% water-insoluble solids (WIS) and PWM containing 2.5% WIS, resulting in an ethanol concentration of 56.5 g/L. This yield was higher than those obtained with SSF of either SPWS (68%) or PWM alone (91%).
Conclusions:
Mixing wheat straw with wheat meal would be beneficial for both 1G and 2G ethanol production. However, increasing the proportion of WIS as wheat straw and the possibility of consuming the xylose fraction with a pentose-fermenting yeast should be further investigated.
Background:
The two-step dilute acid hydrolysis (DAH) of softwood is costly in energy demands and capital costs. However, it has the advantage that hydrolysis and subsequent removal of hemicellulose-derived sugars can be carried out under conditions of low severity, resulting in a reduction in the level of sugar degradation products during the more severe subsequent steps of cellulose hydrolysis. In this paper, we discuss a single-step DAH method that incorporates a temperature profile at two levels. This profile should simulate the two-step process while removing its major disadvantage, that is, the washing step between the runs, which leads to increased energy demand.
Results:
The experiments were conducted in a reactor with a controlled temperature profile. The total dry matter content of the hydrolysate was up to 21.1% w/w, corresponding to a content of 15.5% w/w of water insoluble solids. The highest measured glucose yield, (18.3 g glucose per 100 g dry raw material), was obtained after DAH cycles of 3 min at 209degreesC and 6 min at 211degreesC with 1% H2SO4, which resulted in a total of 26.3 g solubilized C6 sugars per 100g dry raw material. To estimate the remaining sugar potential, enzymatic hydrolysis (EH) of the solid fraction was also performed. EH of the solid residue increased the total level of solubilized C6 sugars to a maximum of 35.5 g per 100 g dry raw material when DAH was performed as described above (3 min at 210degreesC and 2 min at 211degreesC with 1% H2SO4).
Conclusion:
The dual-temperature DAH method did not yield decisively better results than the single-temperature, one-step DAH. When we compared the results with those of earlier studies, the hydrolysis performance was better than with the one-step DAH but not as well as that of the two-step, single-temperature DAH. Additional enzymatic hydrolysis resulted in lower levels of solubilized sugars compared with other studies on one-step DAH and two-step DAH followed by enzymatic hydrolysis. A two-step steam pretreatment with EH gave rise to a considerably higher sugar yield in this study.
Background:
To make lignocellulosic fuel ethanol economically competitive with fossil fuels, it is necessary to reduce the production cost. One way to achieve this is by increasing the substrate concentration in the production process, and thus reduce the energy demand in the final distillation of the fermentation broth. However, increased substrate concentration in simultaneous saccharification and fermentation (SSF) processes has been shown to result in reduced ethanol yields and severe stirring problems. Because the SSF medium is being continuously hydrolyzed, running the process in fed-batch mode could potentially reduce the stirring problems and lead to increased ethanol yields in high-solids SSF. Different enzyme feeding strategies, with the enzymes either present in the reactor from start-up or fed into the reactor together with the substrate, have been studied, along with the influence of the enzyme feeding strategy on the final ethanol yield and productivity.
Results:
In the present study, SSF was run successfully with 10% and 14% water-insoluble solids (WIS) in batch and fed-batch mode. The mixing of the material in the reactor was significantly better in fed-batch than batch mode, and similarly high or higher ethanol yields were achieved in fed-batch mode compared with batch SSF in some cases. No general trend in the dependence of ethanol yield on enzyme feeding strategy was found.
Conclusions:
The optimum enzyme feeding strategy appears to depend on the conditions during SSF, such as the WIS concentration and the concentration of inhibitory compounds in the SSF medium.
Background:
Cost-effective fermentation of lignocellulosic hydrolysate to ethanol by Saccharomyces cerevisiae requires efficient mixed sugar utilization. Notably, the rate and yield of xylose and arabinose co-fermentation to ethanol must be enhanced.
Results:
Evolutionary engineering was used to improve the simultaneous conversion of xylose and arabinose to ethanol in a recombinant industrial Saccharomyces cerevisiae strain carrying the heterologous genes for xylose and arabinose utilization pathways integrated in the genome. The evolved strain TMB3130 displayed an increased consumption rate of xylose and arabinose under aerobic and anaerobic conditions. Improved anaerobic ethanol production was achieved at the expense of xylitol and glycerol but arabinose was almost stoichiometrically converted to arabitol. Further characterization of the strain indicated that the selection pressure during prolonged continuous culture in xylose and arabinose medium resulted in the improved transport of xylose and arabinose as well as increased levels of the enzymes from the introduced fungal xylose pathway. No mutation was found in any of the genes from the pentose converting pathways.
Conclusion:
To the best of our knowledge, this is the first report that characterizes the molecular mechanisms for improved mixed-pentose utilization obtained by evolutionary engineering of a recombinant S. cerevisiae strain. Increased transport of pentoses and increased activities of xylose converting enzymes contributed to the improved phenotype.
Background:
Corn grain is an important renewable source for bioethanol production in the USA. Corn ethanol is currently produced by steam liquefaction of starch-rich grains followed by enzymatic saccharification and fermentation. Corn stover (the non-grain parts of the plant) is a potential feedstock to produce cellulosic ethanol in second-generation biorefineries. At present, corn grain is harvested by removing the grain from the living plant while leaving the stover behind on the field. Alternatively, whole corn plants can be harvested to cohydrolyze both starch and cellulose after a suitable thermochemical pretreatment to produce fermentable monomeric sugars. In this study, we used physiologically immature corn silage (CS) and matured whole corn plants (WCP) as feedstocks to produce ethanol using ammonia fiber expansion (AFEX) pretreatment followed by enzymatic hydrolysis (at low enzyme loadings) and cofermentation (for both glucose and xylose) using a cellulase-amylase-based cocktail and a recombinant Saccharomyces cerevisiae 424A (LNH-ST) strain, respectively. The effect on hydrolysis yields of AFEX pretreatment conditions and a starch/cellulose-degrading enzyme addition sequence for both substrates was also studied.
Results:
AFEX-pretreated starch-rich substrates (for example, corn grain, soluble starch) had a 1.5-3-fold higher enzymatic hydrolysis yield compared with the untreated substrates. Sequential addition of cellulases after hydrolysis of starch within WCP resulted in 15-20% higher hydrolysis yield compared with simultaneous addition of hydrolytic enzymes. AFEX-pretreated CS gave 70% glucan conversion after 72 h of hydrolysis for 6% glucan loading (at 8 mg total enzyme loading per gram glucan). Microbial inoculation of CS before ensilation yielded a 10-15% lower glucose hydrolysis yield for the pretreated substrate, due to loss in starch content. Ethanol fermentation of AFEX-treated (at 6% w/w glucan loading) CS hydrolyzate (resulting in 28 g/L ethanol at 93% metabolic yield) and WCP (resulting in 30 g/L ethanol at 89% metabolic yield) is reported in this work.
Conclusions:
The current results indicate the feasibility of co-utilization of whole plants (that is, starchy grains plus cellulosic residues) using an ammonia-based (AFEX) pretreatment to increase bioethanol yield and reduce overall production cost.
Background:
Fermentations using Escherichia coli KO11, Saccharomyces cerevisiae 424A(LNH-ST), and Zymomonas mobilis AX101 are compared side-by-side on corn steep liquor (CSL) media and the water extract and enzymatic hydrolysate from ammonia fiber expansion (AFEX)-pretreated corn stover.
Results:
The three ethanologens are able produce ethanol from a CSL-supplemented co-fermentation at a metabolic yield, final concentration and rate greater than 0.42 g/g consumed sugars, 40 g/L and 0.7 g/L/h (0-48h), respectively. Xylose-only fermentation of the tested ethanologenic bacteria are five to eight times faster than 424A(LNH-ST) in the CSL fermentation.All tested strains grow and co-ferment sugars at 15% w/v solids loading equivalent of ammonia fiber explosion (AFEX)-pretreated corn stover water extract. However, both KO11 and 424A(LNH-ST) exhibit higher growth robustness than AX101. In 18% w/w solids loading lignocellulosic hydrolysate from AFEX pretreatment, complete glucose fermentations can be achieved at a rate greater than 0.77 g/L/h. In contrast to results from fermentation in CSL, S. cerevisiae 424A(LNH-ST) consumed xylose at the greatest extent and rate in the hydrolysate compared to the bacteria tested.
Conclusions:
Our results confirm that glucose fermentations among the tested strains are effective even at high solids loading (18% by weight). However, xylose consumption in the lignocellulosic hydrolysate is the major bottleneck affecting overall yield, titer or rate of the process. In comparison, Saccharomyces cerevisiae 424A(LNH-ST) is the most relevant strains for industrial production for its ability to ferment both glucose and xylose from undetoxified and unsupplemented hydrolysate from AFEX-pretreated corn stover at high yield.
Although measurements of crystallinity index (CI) have a long history, it has been found that CI varies significantly depending on the choice of measurement method. In this study, four different techniques incorporating X-ray diffraction and solid-state 13C nuclear magnetic resonance (NMR) were compared using eight different cellulose preparations. We found that the simplest method, which is also the most widely used, and which involves measurement of just two heights in the X-ray diffractogram, produced significantly higher crystallinity values than did the other methods. Data in the literature for the cellulose preparation used (Avicel PH-101) support this observation. We believe that the alternative X-ray diffraction (XRD) and NMR methods presented here, which consider the contributions from amorphous and crystalline cellulose to the entire XRD and NMR spectra, provide a more accurate measure of the crystallinity of cellulose. Although celluloses having a high amorphous content are usually more easily digested by enzymes, it is unclear, based on studies published in the literature, whether CI actually provides a clear indication of the digestibility of a cellulose sample. Cellulose accessibility should be affected by crystallinity, but is also likely to be affected by several other parameters, such as lignin/hemicellulose contents and distribution, porosity, and particle size. Given the methodological dependency of cellulose CI values and the complex nature of cellulase interactions with amorphous and crystalline celluloses, we caution against trying to correlate relatively small changes in CI with changes in cellulose digestibility. In addition, the prediction of cellulase performance based on low levels of cellulose conversion may not include sufficient digestion of the crystalline component to be meaningful.
Background:
Grasses are relatively recalcitrant to genetic transformation in comparison to certain dicotyledons, yet they constitute some of the most important biofuel crops. Genetic transformation of switchgrass (Panicum virgatum L.) has previously been reported after cocultivation of explants with Agrobacterium and biolistics of embryogenic calli. Experiments to increase transient gene expression in planta may lead to stable transformation methods with increased efficiency.
Results:
A high-throughput Agrobacterium-mediated transient gene expression system has been developed for in planta inoculation of germinating switchgrass seedlings. Four different Agrobacterium strains were compared for their ability to infect switchgrass seedlings, and strain AGL1 was found to be the most infective. Wounding pretreatments such as sonication, mixing by vortex with carborundum, separation by centrifugation, vacuum infiltration, and high temperature shock significantly increased transient expression of a reporter gene (GUSPlus, a variation of the beta-glucuronidase (GUS) gene). The addition of L-cysteine and dithiothreitol in the presence of acetosyringone significantly increased GUS expression compared with control treatments, whereas the addition of 0.1% surfactants such as Silwet L77 or Li700 decreased GUS expression. 4-Methylumbelliferyl beta-D-galactopyranoside (MUG) assays showed a peak of beta-glucuronidase (GUS) enzyme activity 3 days after cocultivation with Agrobacterium harboring pCambia1305.2, whereas MUG assays showed a peak of enzyme activity 5 days after cocultivation with Agrobacterium harboring pCambia1305.1.
Conclusion:
Agrobacterium strains C58, GV3101 and EHA105 are less able to deliver transfer DNA to switchgrass seedlings (cultivar Alamo) compared with strain AGL1. Transient expression was increased by double or triple wounding treatments such as mixing by vortex with carborundum, sonication, separation by centrifugation, and heat shock. The addition of thiol compounds such as L-cysteine and dithiothreitol in combination with acetosyringone during cocultivation also increased transient expression. The combination of multiple wounding treatments along with the addition of thiol compounds during cocultivation increased transient expression levels from 6% to 54%. There were differences in temporal GUS expression induced by pCambia1305.1 and pCambia1305.2.
Background:
US legislation requires the use of advanced biofuels to be made from non-food feedstocks. However, commercialization of lignocellulosic ethanol technology is more complex than expected and is therefore running behind schedule. This is creating a demand for non-food, but more easily converted, starch-based feedstocks other than corn that can fill the gap until the second generation technologies are commercially viable. Winter barley is such a feedstock but its mash has very high viscosity due to its high content of -glucans. This fact, along with a lower starch content than corn, makes ethanol production at the commercial scale a real challenge.
Results:
A new fermentation process for ethanol production from Thoroughbred, a winter barley variety with a high starch content, was developed. The new process was designated the EDGE (enhanced dry grind enzymatic) process. In this process, in addition to the normal starch-converting enzymes, two accessory enzymes were used to solve the beta-glucan problem. First, beta-glucanases were used to hydrolyze the beta-glucans to oligomeric fractions, thus significantly reducing the viscosity to allow good mixing for the distribution of the yeast and nutrients. Next, beta-glucosidase was used to complete the beta-glucan hydrolysis and to generate glucose, which was subsequently fermented in order to produce additional ethanol. While beta-glucanases have been previously used to improve barley ethanol production by lowering viscosity, this is the first full report on the benefits of adding beta-glucosidases to increase the ethanol yield.
Conclusions:
In the EDGE process, 30% of total dry solids could be used to produce 15% v/v ethanol. Under optimum conditions an ethanol yield of 402 L/MT (dry basis) or 2.17 gallons/53 lb bushel of barley with 15% moisture was achieved. The distillers dried grains with solubles (DDGS) co-product had extremely low beta-glucan (below 0.2%) making it suitable for use in both ruminant and mono-gastric animal feeds.
Mobile phone recycling is {starting to increase in popularity with numerous individuals from around the world, but what mobile phone is selling for the most? And which cell phone is the best selling?
Well predictably the iPhone 3G is the most valuable at the moment clocking in at a staggering £350 at MobilePhoneXchange, so if you are in possession of a new Apple iPhone that you no longer use, or just can’t get used to then you could make yourself a quite a bit of money.
The most popular phone that can be used for mobile phone recycling at the present time is the Nokia N95 and if you have one that you wish to get rid of you can get up to £118 if you trade it in now at Love2Recycle. So it’s time to dust down your old mobile phones and put some money in your pocket!
Background:
The conditions for steam pretreatment of sugar cane bagasse and leaves were studied using CO2 as an impregnating agent. The following conditions were investigated: time (5 to 15 min) and temperature (190 to 220degreesC). The pretreatment was assessed in terms of glucose and xylose yields after enzymatic hydrolysis and inhibitor formation (furfural and hydroxymethylfurfural) in the pretreatment. Results from pretreatment using SO2 as impregnating agent was used as reference.
Results:
For sugar cane bagasse, the highest glucose yield (86.6% of theoretical) was obtained after pretreatment at 205degreesC for 15 min. For sugar cane leaves the highest glucose yield (97.2% of theoretical) was obtained after pretreatment at 220degreesC for 5 min. The reference pretreatment, using impregnation with SO2 and performed at 190degreesC for 5 min, resulted in an overall glucose yield of 79.7% and 91.9% for bagasse and leaves, respectively.
Conclusions:
Comparable pretreatment performance was obtained with CO2 as compared to when SO2 is used, although higher temperature and pressure were needed. The results are encouraging as some characteristics of CO2 are very attractive, such as high availability, low cost, low toxicity, low corrosivity and low occupational risk.
Background:
Different mechanistic models have been used in the literature to describe the enzymatic hydrolysis of pretreated biomass. Although these different models have been applied to different substrates, most of these mechanistic models fit into two- and three-parameter mechanistic models. The purpose of this study is to compare the models and determine the activation energy and the enthalpy of adsorption of Trichoderma reesei enzymes on ammonia fibre explosion (AFEX)-treated wheat straw. Experimental enzymatic hydrolysis data from AFEX-treated wheat straw were modelled with two- and three-parameter mechanistic models from the literature. In order to discriminate between the models, initial rate data at 49oC were subjected to statistical analysis (analysis of variance and scatter plots).
Results:
For three-parameter models, the HCH-1 model best fitted the experimental data; for two-parameter models Michaelis-Menten (M-M) best fitted the experimental data. All the three-parameter models fitted the data better than the two-parameter models. The best three models at 49oC (HCH-1, Huang and M-M) were compared using initial rate data at three temperatures (35o, 42o and 49oC). The HCH-1 model provided the best fit based on the F values, the scatter plot and the residual sum of squares. Also, its kinetic parameters were linear in Arrhenius/van’t Hoff’s plots, unlike the other models. The activation energy (Ea) is 47.6 kJ/mol and the enthalpy change of adsorption (H) is -118 kJ/mol for T. reesei enzymes on AFEX-treated wheat straw.
Conclusion:
Among the two-parameter models, Michaelis-Menten model provided the best fit compared to models proposed by Humphrey and Wald. For the three-parameter models, HCH-1 provided the best fit because the model includes a fractional coverage parameter which accounts for the number of reactive sites covered by the enzymes.
Background:
Baker’s yeast (Saccharomyces cerevisiae) has been engineered for xylose utilization to enable production of fuel ethanol from lignocellulose raw material. One unresolved challenge is that S. cerevisiae lacks a dedicated transport system for pentose sugars, which means that xylose is transported by non-specific Hxt transporters with comparatively low transport rate and affinity for xylose.
Results:
In this study, we compared three heterologous xylose transporters that have recently been shown to improve xylose uptake under different experimental conditions. The transporters Gxf1, Sut1 and At5g59250 from Candida intermedia, Pichia stipitis and Arabidopsis thaliana, respectively, were expressed in isogenic strains of S. cerevisiae and the transport kinetics and utilization of xylose was evaluated. Expression of the Gxf1 and Sut1 transporters led to significantly increased affinity and transport rates of xylose. In batch cultivation at 4 g/L xylose concentration, improved transport kinetics led to a corresponding increase in xylose utilization, whereas no correlation could be demonstrated at xylose concentrations greater than 15 g/L. The relative contribution of native sugar transporters to the overall xylose transport capacity was also estimated during growth on glucose and xylose.
Conclusions:
Kinetic characterization and aerobic batch cultivation of strains expressing the Gxf1, Sut1 and At5g59250 transporters showed a direct relationship between transport kinetics and xylose growth. The Gxf1 transporter had the highest transport capacity and the highest xylose growth rate, followed by the Sut1 transporter. The range in which transport controlled the growth rate was determined to between 0 and 15 g/L xylose. The role of catabolite repression in regulation of native transporters was also confirmed by the observation that xylose transport by native S. cerevisiae transporters increased significantly during cultivation in xylose and at low glucose concentration.
The efficient enzymatic saccharification of cellulose at low cellulase (protein) loadings continues to be a challenge for commercialization of a process for bioconversion of lignocellulose to ethanol. Currently, effective pretreatment followed by high enzyme loading is needed to overcome several substrate and enzyme factors that limit rapid and complete hydrolysis of the cellulosic fraction of biomass substrates. One of the major barriers faced by cellulase enzymes is their limited access to much of the cellulose that is buried within the highly ordered and tightly packed fibrillar architecture of the cellulose microfibrils. Rather than a sequential ’shaving’ or ‘planing’ of the cellulose fibrils from the outside, it has been suggested that these inaccessible regions are disrupted or loosened by non-hydrolytic proteins, thereby increasing the cellulose surface area and making it more accessible to the cellulase enzyme complex. This initial stage in enzymatic saccharification of cellulose has been termed amorphogenesis. In this review, we describe the various amorphogenesis-inducing agents that have been suggested, and their possible role in enhancing the enzymatic hydrolysis of cellulose.
Background:
The enzymatic hydrolysis of cellulose is still considered as one of the main limiting steps of the biological production of biofuels from lignocellulosic biomass. It is a complex multistep process, and various kinetic models have been proposed. The cellulase enzymatic cocktail secreted by Trichoderma reesei has been intensively investigated. beta-glucosidases are one of a number of cellulolytic enzymes, and catalyze the last step releasing glucose from the inhibitory cellobiose. beta-glucosidase (BGL1) is very poorly secreted by Trichoderma reesei strains, and complete hydrolysis of cellulose often requires supplementation with a commercial beta-glucosidase preparation such as that from Aspergillus niger (Novozymes SP188). Surprisingly, kinetic modeling of beta-glucosidases lacks reliable data, and the possible differences between native T. reesei and supplemented beta-glucosidases are not taken into consideration, possibly because of the difficulty of purifying BGL1.
Results:
A comparative kinetic analysis of beta-glucosidase from Aspergillus niger and BGL1 from Trichoderma reesei, purified using a new and efficient fast protein liquid chromatography protocol, was performed. This purification is characterized by two major steps, including the adsorption of the major cellulases onto crystalline cellulose, and a final purification factor of 53. Quantitative analysis of the resulting beta-glucosidase fraction from T. reesei showed it to be 95% pure. Kinetic parameters were determined using cellobiose and a chromogenic artificial substrate. A new method allowing easy and rapid determination of the kinetic parameters was also developed. beta-Glucosidase SP188 (Km = 0.57 mM; Kp = 2.70 mM) has a lower specific activity than BGL1 (Km = 0.38 mM; Kp = 3.25 mM) and is also more sensitive to glucose inhibition. A Michaelis-Menten model integrating competitive inhibition by the product (glucose) has been validated and is able to predict the beta-glucosidase activity of both enzymes.
Conclusions:
This article provides a useful comparison between the activity of beta-glucosidases from two different fungi, and shows the importance of fully characterizing both enzymes. A Michaelis-Menten model was developed, including glucose inhibition and kinetic parameters, which were accurately determined and compared. This model can be further integrated into a cellulose hydrolysis model dissociating beta-glucosidase activity from that of other cellulases. It can also help to define the optimal enzymatic cocktails for new beta-glucosidase activities.
Background:
Biofuels offer a viable alternative to petroleum-based fuel. However, current methods are not sufficient and the technology required in order to use lignocellulosic biomass as a fermentation substrate faces several challenges. One challenge is the need for a robust fermentative microorganism that can tolerate the inhibitors present during lignocellulosic fermentation. These inhibitors include the furan aldehyde, furfural, which is released as a byproduct of pentose dehydration during the weak acid pretreatment of lignocellulose. In order to survive in the presence of furfural, yeast cells need not only to reduce furfural to the less toxic furan methanol, but also to protect themselves and repair any damage caused by the furfural. Since furfural tolerance in yeast requires a functional pentose phosphate pathway (PPP), and the PPP is associated with reactive oxygen species (ROS) tolerance, we decided to investigate whether or not furfural induces ROS and its related cellular damage in yeast.
Results:
We demonstrated that furfural induces the accumulation of ROS in Saccharomyces cerevisiae. In addition, furfural was shown to cause cellular damage that is consistent with ROS accumulation in cells which includes damage to mitochondria and vacuole membranes, the actin cytoskeleton and nuclear chromatin. The furfural-induced damage is less severe when yeast are grown in a furfural concentration (25 mM) that allows for eventual growth after an extended lag compared to a concentration of furfural (50 mM) that prevents growth.
Conclusion:
These data suggest that when yeast cells encounter the inhibitor furfural, they not only need to reduce furfural into furan methanol but also to protect themselves from the cellular effects of furfural and repair any damage caused. The reduced cellular damage seen at 25 mM furfural compared to 50 mM furfural may be linked to the observation that at 25 mM furfural yeast were able to exit the furfural-induced lag phase and resume growth. Understanding the cellular effects of furfural will help direct future strain development to engineer strains capable of tolerating or remediating ROS and the effects of ROS.
Background:
When producing biofuels from dedicated feedstock, agronomic factors such as harvest time and location can impact the downstream production. Thus, this paper studies the effectiveness of ammonia fibre expansion (AFEX) pretreatment on two harvest times (July and October) and ecotypes/locations (Cave-in-Rock (CIR) harvested in Michigan and Alamo harvested in Alabama) for switchgrass (Panicum virgatum).
Results:
Both harvest date and ecotype/location determine the pretreatment conditions that produce maximum sugar yields. There was a high degree of correlation between glucose and xylose released regardless of the harvest, pretreatment conditions, or enzyme formulation. Enzyme formulation that produced maximum sugar yields was the same across all harvests except for the CIR October harvest. The least mature sample, the July harvest of CIR switchgrass, released the most sugars (520 g/kg biomass) during enzymatic hydrolysis while requiring the least severe pretreatment conditions. In contrast, the most mature harvest released the least amount of sugars (410 g/kg biomass). All hydrolysates were highly fermentable, although xylose utilisation in the July CIR hydrolysate was poor.
Conclusions:
Each harvest type and location responded differently to AFEX pretreatment, although all harvests successfully produced fermentable sugars. Thus, it is necessary to consider an integrated approach between agricultural production and biochemical processing in order to insure optimal productivity.
Background:
In this study, the dilute maleic acid pretreatment of wheat straw is optimized, using pretreatment time, temperature and maleic acid concentration as design variables. A central composite design was applied to the experimental set up. The response factors used in this study are: (1) glucose benefits from improved enzymatic digestibility of wheat straw solids; (2) xylose benefits from the solubilization of xylan to the liquid phase during the pretreatment; (3) maleic acid replenishment costs; (4) neutralization costs of pretreated material; (5) costs due to furfural production; and (6) heating costs of the input materials. For each response factor, experimental data were fitted mathematically. After data translation to E/Mg dry straw, determining the relative contribution of each response factor, an economic optimization was calculated within the limits of the design variables.
Results:
When costs are disregarded, an almost complete glucan conversion to glucose can be reached (90% from solids, 7%-10% in liquid), after enzymatic hydrolysis. During the pretreatment, up to 90% of all xylan is converted to monomeric xylose. Taking cost factors into account, the optimal process conditions are: 50 min at 170 degreesC, with 46 mM maleic acid, resulting in a yield of 65 E/Mg (megagram = metric ton) dry straw, consisting of 68 E/Mg glucose benefits (from solids: 85% of all glucan), 17 E/Mg xylose benefits (from liquid: 80% of all xylan), 17 E/Mg maleic acid costs, 2.0 E/Mg heating costs and 0.68 E/Mg NaOH costs. In all but the most severe of the studied conditions, furfural formation was so limited that associated costs are considered negligible.
Conclusions:
After the dilute maleic acid pretreatment and subsequent enzymatic hydrolysis, almost complete conversion of wheat straw glucan and xylan is possible. Taking maleic acid replenishment, heating, neutralization and furfural formation into account, the optimum in the dilute maleic acid pretreatment of wheat straw in this study is 65 E/Mg dry feedstock. This is reached when process conditions are: 50 min at 170 degreesC, with a maleic acid concentration of 46 mM. Maleic acid replenishment is the most important of the studied cost factors.
