http://www.biotechnologyforbiofuels.com The most accessed research articles published by Biotechnology for Biofuels 2010-02-23T00:00:00Z This is an RSS newsfeed from BioMed Central It is intended to be used with an RSS reader. For more information about RSS newsfeeds from BioMed Central, visit http://www.biomedcentral.com/info/about/rss/ 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. http://www.biotechnologyforbiofuels.com/content/3/1/4 Valdeir Arantes Jack Saddler Biotechnology for Biofuels 2010, 3:4 2010-02-23T00:00:00Z doi:10.1186/1754-6834-3-4 Biotechnology for Biofuels 1754-6834 3 4 2010-02-23T00:00:00Z PDF 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. http://www.biotechnologyforbiofuels.com/content/3/1/3 Marie Chauve Hugues Mathis Delphine Huc Dominique Casanave Frederic Monot Nicolas Lopes Ferreira Biotechnology for Biofuels 2010, 3:3 2010-02-11T00:00:00Z doi:10.1186/1754-6834-3-3 Biotechnology for Biofuels 1754-6834 3 3 2010-02-11T00:00:00Z PDF Background: Two economic factors make watermelon worthy of consideration as a feedstock for ethanol biofuel production. First, about 20% of each annual watermelon crop is left in the field because of surface blemishes or because they are misshapen; currently these are lost to growers as a source of revenue. Second, the neutraceutical value of lycopene and L-citrulline obtained from watermelon is at a threshold whereby watermelon could serve as starting material to extract and manufacture these products. Processing of watermelons to produce lycopene and L-citrulline, yields a waste stream of watermelon juice at the rate of over 500 L/t of watermelons. Since watermelon juice contains 7 to 10% (w/v) directly fermentable sugars and 15 to 35 μmol/ml of free amino acids, its potential as feedstock, diluent, and nitrogen supplement was investigated in fermentations to produce bioethanol. Results: Complete watermelon juice and that which did not contain the chromoplasts (lycopene), but did contain free amino acids, were readily fermentable as the sole feedstock or as diluent, feedstock supplement, and nitrogen supplement to granulated sugar or molasses. A minimum level of ~400 mg N/L (~15 μmol/ml amino nitrogen) in watermelon juice was required to achieve maximal fermentation rates when it was employed as the sole nitrogen source for the fermentation. Fermentation at pH 5 produced the highest rate of fermentation for the yeast system that was employed. Utilizing watermelon juice as diluent, supplemental feedstock, and nitrogen source for fermentation of processed sugar or molasses allowed complete fermentation of up to 25% (w/v) sugar concentration at pH 3 (0.41 to 0.46 g ethanol per g sugar) or up to 35% (w/v) sugar concentration at pH 5 with a conversion to 0.36 to 0.41 g ethanol per g sugar. Conclusion: Although watermelon juice would have to be concentrated 2.5- to 3-fold to serve as the sole feedstock for ethanol biofuel production, the results of this investigation indicate that watermelon juice, either as whole juice fermented on-site or as a waste stream from neutraceutical production, could easily integrate with other more concentrated feedstocks where it could serve as diluent, supplemental feedstock, and nitrogen supplement. http://www.biotechnologyforbiofuels.com/content/2/1/18 Wayne Fish Benny Bruton Vincent Russo Biotechnology for Biofuels 2009, 2:18 2009-08-26T00:00:00Z doi:10.1186/1754-6834-2-18 Biotechnology for Biofuels 1754-6834 2 18 2009-08-26T00:00:00Z XML Ethanol is a biofuel that is used as a replacement for approximately 3% of the fossil-based gasoline consumed in the world today. Most of this biofuel is produced from sugarcane in Brazil and corn in the United States. We present here the rationale for the ethanol program in Brazil, its present 'status' and its perspectives. The environmental benefits of the program, particularly the contribution of ethanol to reducing the emission of greenhouse gases, are discussed, as well as the limitations to its expansion. http://www.biotechnologyforbiofuels.com/content/1/1/6 Jose Goldemberg Biotechnology for Biofuels 2008, 1:6 2008-05-01T00:00:00Z doi:10.1186/1754-6834-1-6 Biotechnology for Biofuels 1754-6834 1 6 2008-05-01T00:00:00Z XML Transgenic modification of plants is a key enabling technology for developing sustainable biofeedstocks for biofuels production. Regulatory decisions and the wider acceptance and development of transgenic biofeedstock crops are considered from the context of science-based risk assessment. The risk assessment paradigm for transgenic biofeedstock crops is fundamentally no different from that of current generation transgenic crops, except that the focus of the assessment must consider the unique attributes of a given biofeedstock crop and its environmental release. For currently envisioned biofeedstock crops, particular emphasis in risk assessment will be given to characterization of altered metabolic profiles and their implications relative to non-target environmental effects and food safety; weediness and invasiveness when plants are modified for abiotic stress tolerance or are domesticated; and aggregate risk when plants are platforms for multi-product production. Robust risk assessments for transgenic biofeedstock crops are case-specific, initiated through problem formulation, and use tiered approaches for risk characterization. http://www.biotechnologyforbiofuels.com/content/2/1/27 Jeffrey Wolt Biotechnology for Biofuels 2009, 2:27 2009-11-02T00:00:00Z doi:10.1186/1754-6834-2-27 Biotechnology for Biofuels 1754-6834 2 27 2009-11-02T00:00:00Z XML 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 €/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°C, with 46 mM maleic acid, resulting in a yield of 65 €/Mg (megagram = metric ton) dry straw, consisting of 68 €/Mg glucose benefits (from solids: 85% of all glucan), 17 €/Mg xylose benefits (from liquid: 80% of all xylan), 17 €/Mg maleic acid costs, 2.0 €/Mg heating costs and 0.68 €/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 €/Mg dry feedstock. This is reached when process conditions are: 50 min at 170°C, with a maleic acid concentration of 46 mM. Maleic acid replenishment is the most important of the studied cost factors. http://www.biotechnologyforbiofuels.com/content/2/1/31 A. Maarten Kootstra Hendrik Beeftink Elinor Scott Johan Sanders Biotechnology for Biofuels 2009, 2:31 2009-12-21T00:00:00Z doi:10.1186/1754-6834-2-31 Biotechnology for Biofuels 1754-6834 2 31 2009-12-21T00:00:00Z XML Background: Transesterification of Jatropha oil was carried out in t-butanol solvent using immobilized lipase from Enterobacter aerogenes. The presence of t-butanol significantly reduced the negative effects caused by both methanol and glycerol. The effects of various reaction parameters on transesterification of Jatropha oil were studied. Results: The maximum yield of biodiesel was 94% (of which 68% conversion was achieved with respect to methyl oleate) with an oil:methanol molar ratio of 1:4, 50 U of immobilized lipase/g of oil, and a t-butanol:oil volume ratio of 0.8:1 at 55°C after 48 h of reaction time. There was negligible loss in lipase activity even after repeated use for seven cycles. Conclusion: To the best of our knowledge this is the first report on biodiesel synthesis using immobilized E. aerogenes lipase. http://www.biotechnologyforbiofuels.com/content/2/1/1 Annapurna Kumari Paramita Mahapatra Vijay Kumar Garlapati Rintu Banerjee Biotechnology for Biofuels 2009, 2:1 2009-01-14T00:00:00Z doi:10.1186/1754-6834-2-1 Biotechnology for Biofuels 1754-6834 2 1 2009-01-14T00:00:00Z XML Background: Pretreatment chemistry is of central importance due to its impacts on cellulosic biomass processing and biofuels conversion. Ammonia fiber expansion (AFEX) and dilute acid are two promising pretreatments using alkaline and acidic pH that have distinctive differences in pretreatment chemistries. Results: Comparative evaluation on these two pretreatments reveal that (i) AFEX-pretreated corn stover is significantly more fermentable with respect to cell growth and sugar consumption, (ii) both pretreatments can achieve more than 80% of total sugar yield in the enzymatic hydrolysis of washed pretreated solids, and (iii) while AFEX completely preserves plant carbohydrates, dilute acid pretreatment at 5% solids loading degrades 13% of xylose to byproducts. Conclusion: The selection of pretreatment will determine the biomass-processing configuration, requirements for hydrolysate conditioning (if any) and fermentation strategy. Through dilute acid pretreatment, the need for hemicellulase in biomass processing is negligible. AFEX-centered cellulosic technology can alleviate fermentation costs through reducing inoculum size and practically eliminating nutrient costs during bioconversion. However, AFEX requires supplemental xylanases as well as cellulase activity. As for long-term sustainability, AFEX has greater potential to diversify products from a cellulosic biorefinery due to lower levels of inhibitor generation and lignin loss. http://www.biotechnologyforbiofuels.com/content/2/1/30 Ming Lau Christa Gunawan Bruce Dale Biotechnology for Biofuels 2009, 2:30 2009-12-04T00:00:00Z doi:10.1186/1754-6834-2-30 Biotechnology for Biofuels 1754-6834 2 30 2009-12-04T00:00:00Z XML 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. http://www.biotechnologyforbiofuels.com/content/3/1/2 Sandra Allen William Clark J McCaffery Zhen Cai Alisson Lanctot Patricia Slininger Z Liu Steven Gorsich Biotechnology for Biofuels 2010, 3:2 2010-01-15T00:00:00Z doi:10.1186/1754-6834-3-2 Biotechnology for Biofuels 1754-6834 3 2 2010-01-15T00:00:00Z XML Background: Corn stover composition changes considerably throughout the growing season and also varies between the various fractions of the plant. These differences can impact optimal pretreatment conditions, enzymatic digestibility and maximum achievable sugar yields in the process of converting lignocellulosics to ethanol. The goal of this project was to determine which combination of corn stover fractions provides the most benefit to the biorefinery in terms of sugar yields and to determine the preferential order in which fractions should be harvested. Ammonia fiber expansion (AFEX) pretreatment, followed by enzymatic hydrolysis, was performed on early and late harvest corn stover fractions (stem, leaf, husk and cob). Sugar yields were used to optimize scenarios for the selective harvest of corn stover assuming 70% or 30% collection of the total available stover. Results: The optimal AFEX conditions for all stover fractions, regardless of harvest period, were: 1.5 (g NH3 g-1 biomass); 60% moisture content (dry-weight basis; dwb), 90°C and 5 min residence time. Enzymatic hydrolysis was conducted using cellulase, β-glucosidase, and xylanase at 31.3, 41.3, and 3.1 mg g-1 glucan, respectively. The optimal harvest order for selectively harvested corn stover (SHCS) was husk > leaf > stem > cob. This harvest scenario, combined with optimal AFEX pretreatment conditions, gave a theoretical ethanol yield of 2051 L ha-1 and 912 L ha-1 for 70% and 30% corn stover collection, respectively. Conclusion: Changing the proportion of stover fractions collected had a smaller impact on theoretical ethanol yields (29 - 141 L ha-1) compared to the effect of altering pretreatment and enzymatic hydrolysis conditions (150 - 462 L ha-1) or harvesting less stover (852 - 1139 L ha-1). Resources may be more effectively spent on improving sustainable harvesting, thereby increasing potential ethanol yields per hectare harvested, and optimizing biomass processing rather than focusing on the selective harvest of specific corn stover fractions. http://www.biotechnologyforbiofuels.com/content/2/1/29 Rebecca Garlock Shishir Chundawat Venkatesh Balan Bruce Dale Biotechnology for Biofuels 2009, 2:29 2009-11-24T00:00:00Z doi:10.1186/1754-6834-2-29 Biotechnology for Biofuels 1754-6834 2 29 2009-11-24T00:00:00Z XML