Abstract: Triacylglycerol (TAG) biosynthesis is a principal metabolic pathway in most organisms, and TAG is the major form of carbon storage in many plant seeds. Acyl-CoA:diacylglycerol acyltransferase 1 (DGAT1) is the only acyltransferase enzyme that has been confirmed to contribute to TAG biosynthesis in Arabidopsis thaliana seeds. However, dgat1 null mutants display only a 20 to 40% decrease in seed oil content. To determine whether other enzymes contribute to TAG synthesis, candidate genes were expressed in TAG-deficient yeast, candidate mutants were crossed with the dgat1-1 mutant, and target genes were suppressed by RNA interference (RNAi). An in vivo role for phospholipid:diacylglycerol acyltransferase 1 (PDAT1; At5g13640) in TAG synthesis was revealed in this study. After failing to obtain double homozygous plants from crossing dgat1-1 and pdat1-2, further investigation showed that the dgat1-1 pdat1-2 double mutation resulted in sterile pollen that lacked visible oil bodies. RNAi silencing of PDAT1 in a dgat1-1 background or DGAT1 in pdat1-1 background resulted in 70 to 80% decreases in oil content per seed and in disruptions of embryo development. These results establish in vivo involvement of PDAT1 in TAG biosynthesis, rule out major contributions by other candidate enzymes, and indicate that PDAT1 and DGAT1 have overlapping functions that are essential for normal pollen and seed development of Arabidopsis.

Abstract: Cellulosic biofuel systems have the potential to significantly reduce the environmental impact of the world’s transportation energy requirements. However, realizing this potential will require systems level thinking and scale integration. Until now, we have lacked modeling tools for studying the behavior of integrated cellulosic biofuel systems. In this paper, we describe a new research tool, the Biorefinery and Farm Integration Tool (BFIT) in which the production of fuel ethanol from cellulosic biomass is integrated with crop and animal (agricultural) production models. Uniting these three subsystems in a single combined model has allowed, for the first time, basic environmental and economic analysis of biomass production, possible secondary products, fertilizer production, and bioenergy production across various regions of the United States. Using BFIT, we simulate cellulosic ethanol production embedded in realistic agricultural landscapes in nine locations under a collection of farm management scenarios. This combined modeling approach permits analysis of economic profitability and highlights key areas for environmental improvement. These results show the advantages of introducing integrated biorefinery systems within agricultural landscapes. This is particularly true in the Midwest, which our results suggest is a good setting for the cellulosic ethanol industry. Specifically, results show that inclusion of cellulosic biofuel systems into existing agriculture enhances farm economics and reduces total landscape emissions. Model results also indicate a limited ethanol price effect from increased biomass transportation distance. Sensitivity analysis using BFIT revealed those variables having the strongest effects on the overall system performance, namely: biorefinery size, switchgrass yield, and biomass farm gate price.

The accounting now used for assessing compliance with carbon limits in the Kyoto Protocol and in climate legislation contains a far-reaching but fixable flaw that will severely undermine greenhouse gas reduction goals (1). It does not count CO₂ emitted from tailpipes and smokestacks when bioenergy is being used, but it also does not count changes in emissions from land use when biomass for energy is harvested or grown. This accounting erroneously treats all bioenergy as carbon neutral regardless of the source of the biomass, which may cause large differences in net emissions. For example, the clearing of long-established forests to burn wood or to grow energy crops is counted as a 100% reduction in energy emissions despite causing large releases of carbon.

Abstract: The development of a viable lignocellulosic ethanol industry requires multiple improvements in the process of converting biomass to ethanol. A key step is the improvement of the plants that are to be used as biomass feedstocks. To facilitate the identification and evaluation of feedstock plants, it would be useful to have a method to screen large numbers of individual plants for enhanced digestibility in response to combinations of specific pretreatments and enzymes. This paper describes a high-throughput digestibility platform (HTDP) for screening collections of germplasm for improved digestibility, which was developed under the auspices of the Department of Energy-Great Lakes Bioenergy Research Center (DOE-GLBRC). A key component of this platform is a custom-designed workstation that can grind and dispense 1–5 mg quantities of more than 250 different plant tissue samples in 16 h. The other steps in the processing (pretreatment, enzyme digestion, and sugar analysis) have also been largely automated and require 36 h. The process is adaptable to diverse acidic and basic, low-temperature pretreatments. Total throughput of the HTDP is 972 independent biomass samples per week. Validation of the platform was performed on brown midrib mutants of maize, which are known to have enhanced digestibility. Additional validation was performed by screening approximately 1,200 Arabidopsis mutant lines with T-DNA insertions in genes known or suspected to be involved in cell wall biosynthesis. Several lines showed highly significant (p < 0.01) increases in glucose and xylose release (20–40% above the mean). The platform should be useful for screening populations of plants to identify superior germplasm for lignocellulosic ethanol applications and also for screening populations of mutant model plants to identify specific genes affecting digestibility.

Nitrogen (N) is central to living systems, and its addition to agricultural cropping systems is an essential facet of modern crop management and one of the major reasons that crop production has kept pace with human population growth. The benefits of N added to cropping systems come, however, at well-documented environmental costs: Increased coastal hypoxia, atmospheric nitrous oxide (N₂O), reactive N gases in the troposphere, and N deposition onto forests and other natural areas are some of the consequences of our inability to keep fertilizer N from leaving cropped ecosystems via unmanaged pathways. The N cycle is complex, and solutions require a thorough understanding of both the biogeochemical pathways of N in agricultural systems and the consequences of different management practices. Despite the complexity of this challenge, however, a number of technologies are available today to reduce N loss. These include adding rotational complexity to cropping systems to improve N capture by crops, providing farmers with decision support tools for better predicting crop fertilizer N requirements, improving methods for optimizing fertilizer timing and placement, and developing watershed-level strategies to recapture N lost from fields. Solutions to the problem of agricultural N loss will require a portfolio approach in which different technologies are used in different combinations to address site-specific challenges. Solutions will also require incentives that promote their adoption.

Abstract: 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.

Abstract: Studies of aldol condensation/hydrogenation reactions of 2-hexanone were carried out over Pd/CeZrO_x and CeZrO_x catalysts at temperatures between 573 and 673 K, and pressures of 5-26 bar. These studies were formulated to address the catalytic upgrading to transportation fuels of the mono-functional oxygenated compounds (consisting primarily of C₄-C₆ ketones, alcohols, carboxylic acids and heterocyclics) formed by the catalytic conversion of polyols over a Pt-Re/C catalyst. Characterization by XRD, TPR and NH₃/CO₂-TPD showed that Pd/CeZrO_x catalyst consists of a partially reducible solid solution of cerium and zirconium oxides, and possesses both acidic and basic functionalities. Reaction kinetics studies show that in addition to the expected C₁₂ condensation product (7-methyl-5-undecanone), the CeZrO_x-based catalysts produce C₁₈ and C₉ secondary species, along with light alkanes (≤C7). Low loadings of Pd (e.g., 0.25 wt%) lead to optimal activity and selectivity for the production of C₁₂ species. The high apparent activation energy of the formation of C₉ (140 kJ/mol) compared to the formation of C₁₂ and C₁₈ species (15 and 28 kJ/mol, respectively) indicates that these species may be formed as a result of the decomposition of heavier condensation products. The self-coupling of 2-hexanone was found to be positive order in both 2-hexanone and hydrogen. The addition of primary alcohols and carboxylic acids as well as water and CO₂ to the feed was found to reversibly inhibit the self-coupling activity of 2-hexanone. This inhibition is strongest in the presence of CO₂, and TPSR studies indicate that CO₂ is removed from the surface by conversion to CO in the presence of reduced ceria species.

Abstract: Maize stover, including stalks, leaves, and cobs, has potential utility as a cellulosic biofeedstock. Understanding how total stover ethanol potential is affected by the proportion and quality of major plant components would facilitate the genetic improvement of stover quality and inform decisions regarding which plant parts should be targeted for harvesting. Our objectives were to determine how the proportion and composition of plant components affected ethanol potential and whether there are early season predictors of stover quality at maturity. Twenty-three hybrids were evaluated including 20 from a factorial mating design between five silage inbred lines and four commercial inbreds and a brown-midrib3, a Leafy1, and a commercial grain hybrid checks. Plants were harvested and dissected into component parts at developmental stages vegetative 3, vegetative 12, reproductive 3, and reproductive 6 (R6). Tissues were evaluated for acid detergent fiber (ADF), neutral detergent fiber (NDF), and NDF digestibility (NDFD). Stalk was the largest fraction of whole plant dry matter (46.2%) and had the lowest NDFD (3,750 g/kg) at R6. No relationship was found between stalk ADF at early developmental stages and whole plant ADF at R6, suggesting that quality at early developmental stages is not indicative of quality at physiological maturity. Differences were observed among hybrids for ADF and NDF for most plant parts evaluated. Hybrid-by-developmental stage and hybrid-by-plant part interactions were statistically significant. This indicates that there is minimal opportunity to identify superior hybrids for biofuel production based on the proportion of total biomass represented by a plant part and its quality at early developmental stages. Maximum conversion efficiency is attained when leaves are harvested compared to other tissue types at physiological maturity.

Abstract: Mono-functional intermediates produced by catalytic conversion of sugars and polyols over Pt–Re/C catalysts (consisting of alcohols, ketones, carboxylic acids, and heterocyclic compounds) can be upgraded to fuel-grade compounds using two catalytic reactors operated in a cascade mode. The first reactor achieves C–C coupling of mono-functional intermediates using a dual-bed catalyst system, where the upstream catalyst bed (Ce₁Zr₁O_x) is employed to carry out ketonization of carboxylic acids, and the downstream catalyst bed (Pd/ZrO₂) is used to achieve aldol condensation/hydrogenation of alcohols and ketones. This second bed is not significantly inhibited by CO₂ and H₂O produced during ketonization. The high molecular weight ketones produced by C–C coupling reactions in the dual-bed catalyst system are subsequently converted to alkanes by hydrodeoxygenation (i.e., dehydration/hydrogenation) over a Pt/SiO₂–Al₂O₃ catalyst. Using the aforementioned approach, an aqueous feed containing 60 wt% sorbitol was converted to a liquid stream of alkanes, 53% of which consisted of C₇₊ alkanes with minimal branching, desirable for Diesel fuel.

Background: Agricultural residues could potentially be converted to bioenergy, but the sustainable harvest rate is unclear. Results: Residue removal increases soil loss at rates that vary with topography, crop rotation and management; decreases yields (100-year mean yields changed -0.07 to -0.08% for every percent of residue mass removed); decreases soil carbon (approximately 40–90 kg C ha^-1 year^-1 per Mg of residue harvested); and decreases soil nitrogen (3 kg N ha^-1 year^-1 per Mg residue harvested). Conclusion: Even where soil loss is within tolerable limits, harvesting residue is a question of trade-offs in terms of reduction of yield and loss of soil nutrients. The effects of increased residue harvest are highly variable, depending on local climate and soil erodibility and it is thus problematic to apply a single harvest rate globally. However, on flat land under conservation management, the majority of residue could be sustainably harvested for bioenergy.