Distribution of Structural Carbohydrates in Corn Plants Across the Southeastern USA

Quantifying lignin and carbohydrate composition of corn (Zea mays L.) is important to support the emerging cellulosic biofuels industry. Therefore, field studies with 0 or 100 % stover removal were established in Alabama and South Carolina as part of the Sun Grant Regional Partnership Corn Stover Project. In Alabama, cereal rye (Secale cereale L.) was also included as an additional experimental factor, serving as a winter cover crop. Plots were located on major soil types representative of their respective states: Compass and Decatur soils in Alabama and a Coxville/Rains-Goldsboro-Lynchburg soil association in South Carolina. Lignin and structural carbohydrate concentrations in the whole (above-ground) plant, cobs, vegetation excluding cobs above the primary ear (top), vegetation below the primary ear (bottom), and vegetation from above the primary ear including cobs (above-ear fraction) were determined using near-infrared spectroscopy (NIRS). The distribution of lignin, ash, and structural carbohydrates varied among plant fractions, but neither inclusion of a rye cover crop nor the stover harvest treatments consistently affected carbohydrate concentrations within locations. Total precipitation and average air temperature during the growing season were strongly correlated with stover composition indicating that weather conditions may have multiple effects on potential biofuel production (i.e., not only yield but also stover quality). When compared to the above-ear fractions, bottom plant partitions contained greater lignin concentrations. Holocellulose concentration was consistently greater in the above-ear fractions at all three locations. Data from this study suggests that the above-ear plant portions have the most desirable characteristics for cellulosic ethanol production via fermentation in the southeastern USA.     

Significant Variation for Bio-oil Compounds After Pyrolysis/Gas Chromatography–Mass Spectrometry of Cobs and Stover Among Five Near-Isogenic Brown Midrib Hybrids in Maize

We analyzed five near-isogenic brown midrib hybrids in maize via pyrolysis/gas chromatography–mass spectrometry (Py/GC-MS) in order to determine how differing lignin composition and structure impacts individual bio-oil compounds. Twenty-six compounds were analyzed for differences among the five hybrids and between cob and stover materials. We found statistically significant differences for 9 compounds, when comparing the 5 hybrids, and 17 significant differences when comparing maize cobs with stover. Our data indicate that it may be possible to predict phenolic compounds within bio-oil based on cell wall lignin composition. The genetic variation observed in this study suggests that bio-oil quality can be improved by plant breeding.     

Forage quality and composition measurements as predictors of ethanol yield from maize (Zea mays L.) stover

Background  

Improvement of biofeedstock quality for cellulosic ethanol production will be facilitated by inexpensive and rapid methods of evaluation, such as those already employed in the field of ruminant nutrition. Our objective was to evaluate whether forage quality and compositional measurements could be used to estimate ethanol yield of maize stover as measured by a simplified pretreatment and simultaneous saccharification and fermentation assay. Twelve maize varieties selected to be diverse for stover digestibility and composition were evaluated.     

The Feasibility of Producing Adequate Feedstock for Year-Round Cellulosic Ethanol Production in an Intensive Agricultural Fuelshed

To date, cellulosic ethanol production has not been commercialized in the United States. However, government mandates aimed at increasing second-generation biofuel production could spur exploratory development in the cellulosic ethanol industry. We conducted an in-depth analysis of the fuelshed surrounding a starch-based ethanol plant near York, Nebraska that has the potential for cellulosic ethanol production. To assess the feasibility of supplying adequate biomass for year-round cellulosic ethanol production from residual maize (Zea mays) stover and bioenergy switchgrass (Panicum virgatum) within a 40-km road network service area of the existing ethanol plant, we identified ～14,000 ha of marginally productive cropland within the service area suitable for conversion from annual rowcrops to switchgrass and ～132,000 ha of maize-enrolled cropland from which maize stover could be collected. Annual maize stover and switchgrass biomass supplies within the 40-km service area could range between 429,000 and 752,000 metric tons (mT). Approximately 140–250 million liters (l) of cellulosic ethanol could be produced, rivaling the current 208 million l annual starch-based ethanol production capacity of the plant. We conclude that sufficient quantities of biomass could be produced from maize stover and switchgrass near the plant to support year-round cellulosic ethanol production at current feedstock yields, sustainable removal rates and bioconversion efficiencies. Modifying existing starch-based ethanol plants in intensive agricultural fuelsheds could increase ethanol output, return marginally productive cropland to perennial vegetation, and remove maize stover from productive cropland to meet feedstock demand.     

Turning Maize Cobs into a Valuable Feedstock

With rising energy demand and limited resources, the need for alternative energy production is increasing. Maize cobs have an advantageous composition for the production of biofuels such as cellulosic ethanol but are currently left unused on the fields after harvest. Furthermore, cobs contain a low concentration of nitrogen (<1%). Therefore, cob harvest will not deplete soil fertility. Consequently, maize cobs are a cheap and promising source for sustainable energy production. Yet with primary focus on grain yield, no or little effort has been spent to increase cob biomass yield in addition to grain yield. Both cob and grain yield are complex inherited traits affected by the environment. Breeding of dual-purpose maize varieties with simultaneously increased cob and grain yield requires a deeper understanding of factors influencing both cob and grain development. In this article, the available knowledge on the genetics of cob formation and current and future applications of maize cob utilization are discussed to evaluate the prospects for development of dual-purpose maize varieties.     

Effect of stover fraction and storage method on glucose production during enzymatic hydrolysis

One avenue for overcoming the economic challenges associated with the production of ethanol from renewable resources is to reduce the cost of the biomass feedstock. The balance between storage costs and benefits depend on the storage method and composition changes of individual stover fractions. Corn stover from bales stored inside and outside of a barn was separated into an interior and exterior layer after approximately 10 months of storage. The cobs, stalks, and leaves and husks were separated, dried, and ground through a 2 mm screen. Stover, sodium acetate (buffer), cellulase, and deionized water were added to 125 ml flasks. The mixture was held at 50 degrees C in an incubator and samples taken for glucose determination. The average glucose concentration after 60 h of hydrolysis from cobs, leaves and husks, and stalks was 10.5, 9.6, and 3.1 g/l, respectively. Cobs, leaves, and husks produced over 300% more glucose than stalks. Storage outside of the barn decreased the glucose production from individual stover components between 4% and 8%. The effect of stover fraction type on glucose production was significant, while the storage treatment effect was not significant. Fractionation of corn stover may be a method to increase the value of corn stover as a feedstock for glucose production.     

Forage maize in the rations of dairy cows and fattening cattle in Bulgaria

For maize silage containing about 20% dry matter, the most suitable silage: grain ratio is 2:1 for daily milk production over 20 kg, and 3:1 for the rest of the lactation and dry period. Dehydrated whole maize pellets, given in combination with fresh alfalfa to dairy cows, promoted good milk yields.A complete ration containing 50% maize cobs or stover for fattening calves resulted in an average daily gain of 1200–1300 g in the age range 7–14 months. An improvement in energy value was attributed to steam pelleting and the consequent changes in the content of crude fibre, acid detergent fibre and lignin in the maize cobs or stover.     

Effect of anatomical fractionation on the enzymatic hydrolysis of acid and alkaline pretreated corn stover

Due to concerns with biomass collection systems and soil sustainability there are opportunities to investigate the optimal plant fractions to collect for conversion. An ideal feedstock would require a low severity pretreatment to release a maximum amount of sugar during enzymatic hydrolysis. Corn stover fractions were separated manually and analyzed for glucan, xylan, acid soluble lignin, acid insoluble lignin, and ash composition. The stover fractions were also pretreated with either 0%, 0.4%, or 0.8% NaOH for 2 h at room temperature, washed, autoclaved and saccharified. In addition, dilute sulfuric acid pretreated samples underwent simultaneous saccharification and fermentation (SSF) to ethanol. In general, the two pretreatments produced similar trends with cobs, husks, and leaves responding best to the pretreatments, the tops of stalks responding slightly less, and the bottom of the stalks responding the least. For example, corn husks pretreated with 0.8% NaOH released over 90% (standard error of 3.8%) of the available glucan, while only 45% (standard error of 1.1%) of the glucan was produced from identically treated stalk bottoms. Estimates of the theoretical ethanol yield using acid pretreatment followed by SSF were 65% (standard error of 15.9%) for husks and 29% (standard error of 1.8%) for stalk bottoms. This suggests that integration of biomass collection systems to remove sustainable feedstocks could be integrated with the processes within a biorefinery to minimize overall ethanol production costs.     

Ethanolic fermentation of hydrolysates from ammonia fiber expansion (AFEX) treated corn stover and distillers grain without detoxification and external nutrient supplementation

External nutrient supplementation and detoxification of hydrolysate significantly increase the production cost of cellulosic ethanol. In this study, we investigated the feasibility of fermenting cellulosic hydrolysates without washing, detoxification or external nutrient supplementation using ethanologens Escherichia coli KO11 and the adapted strain ML01 at low initial cell density (16 mg dry weight/L). The cellulosic hydrolysates were derived from enzymatically digested ammonia fiber expansion (AFEX)-treated corn stover and dry distiller's grain and solubles (DDGS) at high solids loading (18% by weight). The adaptation was achieved through selective evolution of KO11 on hydrolysate from AFEX-treated corn stover. All cellulosic hydrolysates tested (36-52 g/L glucose) were fermentable. Regardless of strains, metabolic ethanol yields were near the theoretical limit (0.51 g ethanol/g consumed sugar). Volumetric ethanol productivity of 1.2 g/h/L was achieved in fermentation on DDGS hydrolysate and DDGS improved the fermentability of hydrolysate from corn stover. However, enzymatic hydrolysis and xylose utilization during fermentation were the bottlenecks for ethanol production from corn stover at these experimental conditions. In conclusion, fermentation under the baseline conditions was feasible. Utilization of nutrient-rich feedstocks such as DDGS in fermentation can replace expensive media supplementation.     

Effects of Forage Quality and Cell Wall Constituents of Bermuda Grass on Biochemical Conversion to Ethanol

Bermuda grass is an attractive candidate as a feedstock for biofuel production because over four million hectares of Bermuda grass are already grown for forage in the Southern USA. Because both rumen digestion and biochemical conversion to ethanol depend upon enzymatic conversion of the cell wall polysaccharides into fermentable sugars, it is probable that grasses bred for increased forage quality would be more amenable for ethanol production. However, it is not known how variation in rumen digestibility and cell wall/fiber components correlates with efficiency of conversion to ethanol via fermentation. The objective of this research was to determine relationships between ethanol production evaluated by simultaneous saccharification and fermentation (SSF), 72-h in vitro ruminal dry matter digestibility (IVDMD), in vitro ruminal gas production after 24 and 96 h, and biomass composition for 50 genetically diverse Bermuda grass accessions. The Bermuda grass samples were subjected to standard 72-h IVDMD and forage fiber analyses. Also, in separate labs, gas production was measured in sealed volume-calibrated vials after 24 (NNG24) and 96 h (NNG96) of in vitro fermentation by ruminal fluid; ethanol and pentose sugar productions were measured from a bench-top SSF procedure; cell wall constituents were determined by the Uppsala Dietary Fiber Method; and total nitrogen, carbon, and ash concentrations were determined by using the LECO combustion method. Ethanol production was moderately correlated with IVDMD (r?=?0.55) and NNG96 (r?=?0.63) but highly correlated with NNG24 (r?=?0.93). Ethanol was negatively correlated with neutral detergent fiber (NDF; r?=??0.53) and pentose sugars (r?=??0.60), but not correlated with glucose content. Regression models indicated that NDF and cell wall pentose sugar concentrations had significant negative effects on ethanol production. Variation among entries for IVDMD was affected by variability of NDF, pentose sugar concentrations, and biomass nitrogen content. Variation in Klason lignin content had only minor negative impacts on ethanol production and IVDMD. Biochemical conversion efficiency of Bermuda grass by SSF can be best estimated by NNG24 but not by IVDMD.     

The greenhouse gas intensity and potential biofuel production capacity of maize stover harvest in the US Midwest

Agricultural residues are important sources of feedstock for a cellulosic biofuels industry that is being developed to reduce greenhouse gas emissions and improve energy independence. While the US Midwest has been recognized as key to providing maize stover for meeting near‐term cellulosic biofuel production goals, there is uncertainty that such feedstocks can produce biofuels that meet federal cellulosic standards. Here, we conducted extensive site‐level calibration of the Environmental Policy Integrated Climate (EPIC) terrestrial ecosystems model and applied the model at high spatial resolution across the US Midwest to improve estimates of the maximum production potential and greenhouse gas emissions expected from continuous maize residue‐derived biofuels. A comparison of methodologies for calculating the soil carbon impacts of residue harvesting demonstrates the large impact of study duration, depth of soil considered, and inclusion of litter carbon in soil carbon change calculations on the estimated greenhouse gas intensity of maize stover‐derived biofuels. Using the most representative methodology for assessing long‐term residue harvesting impacts, we estimate that only 5.3 billion liters per year (bly) of ethanol, or 8.7% of the near‐term US cellulosic biofuel demand, could be met under common no‐till farming practices. However, appreciably more feedstock becomes available at modestly higher emissions levels, with potential for 89.0 bly of ethanol production meeting US advanced biofuel standards. Adjustments to management practices, such as adding cover crops to no‐till management, will be required to produce sufficient quantities of residue meeting the greenhouse gas emission reduction standard for cellulosic biofuels. Considering the rapid increase in residue availability with modest relaxations in GHG reduction level, it is expected that management practices with modest benefits to soil carbon would allow considerable expansion of potential cellulosic biofuel production.     

Development of Sustainable Corn Stover Harvest Strategies for Cellulosic Ethanol Production

To prepare for a 2014 launch of commercial scale cellulosic ethanol production from corn/maize (Zea mays L.) stover, POET-DSM near Emmetsburg, IA has been working with farmers, researchers, and equipment dealers through “Project Liberty” on harvest, transportation, and storage logistics of corn stover for the past several years. Our objective was to evaluate seven stover harvest strategies within a 50-ha (125 acres) site on very deep, moderately well to poorly drained Mollisols, developed in calcareous glacial till. The treatments included the following: conventional grain harvest (no stover harvest), grain plus a second-pass rake and bale stover harvest, and single-pass grain plus cob-only biomass, grain plus vegetative material other than grain [(MOG) consisting of cobs, husks, and upper plant parts], grain plus all vegetative material from the ear shank upward (high cut), and all vegetative material above a 10 cm stubble height (low cut), with a John Deere 9750 STS combine, and grain plus direct baling of MOG with an AgCo harvesting system. Average grain yields were 11.4, 10.1, 9.7, and 9.5 Mg ha^?1 for 2008, 2009, 2010, and 2011, respectively. Average stover harvest ranged from 0 to 5.6 Mg ha^?1 and increased N, P, and K removal by an average of 11, 1.6, and 15 kg Mg^?1, respectively. Grain yield in 2009 showed a significant positive response to higher 2008 stover removal rates, but grain yield was not increased in 2010 or 2011 due to prior-year stover harvest. High field losses caused the direct-bale treatment to have significantly lower grain yield in 2011 because the AgCo system could not pick up the severely lodged crop. We conclude that decreases in grain yield across the 4 years were due more to seasonal weather patterns, spatial variability, and not rotating crops than to stover harvest.     

Genetic and Morphometric Analysis of Cob Architecture and Biomass-Related Traits in the Intermated B73 × Mo17 Recombinant Inbred Lines of Maize

Expected future cellulosic ethanol production increases the demand for biomass in the US Corn Belt. With low nutritious value, low nitrogen content, and compact biomass, maize cobs can provide a significant amount of cellulosic materials. The value of maize cobs depends on cob architecture, chemical composition, and their relation to grain yield as primary trait. Eight traits including cob volume, fractional diameters, length, weight, tissue density, and grain yield have been analyzed in this quantitative trait locus (QTL) mapping experiment to evaluate their inheritance and inter-relations. One hundred eighty-four recombinant inbred lines of the intermated B73?×?Mo17 (IBM) Syn 4 population were evaluated from an experiment carried out at three locations and analyzed using genotypic information of 1,339 public SNP markers. QTL detection was performed using (1) comparison-wise thresholds with reselection of cofactors (α?=?0.001) and (2) empirical logarithm of odds score thresholds (P?=?0.05). Several QTL with small genetic effects (R ²?=?2.9–13.4 %) were found, suggesting a complex quantitative inheritance of all traits. Increased cob tissue density was found to add value to the residual without a commensurate negative impact on grain yield and therefore enables for simultaneous selection for cob biomass and grain yield.     

Functional group and fertilization affect the composition and bioenergy yields of prairie plants

Prairies used for bioenergy production have potential to generate marketable products while enhancing environmental quality, but little is known about how prairie species composition and nutrient management affect the suitability of prairie biomass for bioenergy production. We determined how functional‐group identity and nitrogen fertilization affected feedstock characteristics and estimated bioenergy yields of prairie plants, and compared those prairie characteristics to that of corn stover. We tested our objectives with a field experiment that was set up as a 5 × 2 incomplete factorial design with C₃ grasses, C₄ grasses, legumes, and multi‐functional‐group mixtures grown with and without nitrogen fertilizer; a fertilized corn treatment was also included. We determined cell wall, hemicellulose, cellulose, and ash concentrations; ethanol conversion ratios; gross caloric ratios; aboveground biomass production; ethanol yields; and energy yields for all treatments. Prairie functional‐group identity affected the biomass feedstock characteristics, whereas nitrogen fertilization did not. Functional group and fertilization had a strong effect on aboveground biomass production, which was the major predictor of ethanol and energy yields. C₄ grasses, especially when fertilized, had among the most favorable bioenergy characteristics with high estimated ethanol conversion ratios and nongrain biomass production, relatively high gross caloric ratios, and low ash concentrations. The bioenergy characteristics of corn stover, from an annual C₄ grass, were similar to those of the biomass of perennial C₄ grasses. Both functional‐group composition and nitrogen fertility management were found to be important in optimizing bioenergy production from prairies.     

Crop Residue Considerations for Sustainable Bioenergy Feedstock Supplies

The anticipated 2014 launch of three full-scale corn stover bioenergy conversion facilities is a strong US market signal that cellulosic feedstock supplies must increase dramatically to supply the required biomass in a sustainable manner. This overview highlights research conducted by the USDA-Agricultural Research Service Renewable Energy Assessment Project (now known as the Resilient Economic Agricultural Practices) team as part of the National Institute for Food and Agriculture Sun Grant Regional Feedstock Partnership Corn Stover team. Stover and grain yield, soil organic carbon, soil aggregation, greenhouse gas, energy content of the stover, and several other factors affecting the fledgling bioenergy industry are addressed in this special issue of the journal.     

A whole cell biocatalyst for cellulosic ethanol production from dilute acid-pretreated corn stover hydrolyzates

In this research, a recombinant whole cell biocatalyst was developed by expressing three cellulases from Clostridium cellulolyticum--endoglucanase (Cel5A), exoglucanase (Cel9E), and β-glucosidase--on the surface of the Escherichia coli LY01. The modified strain is identified as LY01/pRE1H-AEB. The cellulases were displayed on the surface of the cell by fusing with an anchor protein, PgsA. The developed whole cell biocatalyst was used for single-step ethanol fermentation using the phosphoric acid-swollen cellulose (PASC) and the dilute acid-pretreated corn stover. Ethanol production was 3.59 ± 0.15 g/L using 10 g/L of PASC, which corresponds to a theoretical yield of 95.4 ± 0.15%. Ethanol production was 0.30 ± 0.02 g/L when 1 g/L equivalent of glucose in the cellulosic fraction of the dilute sulfuric acid-pretreated corn stover (PCS) was fermented for 84 h. A total of 0.71 ± 0.12 g/L ethanol was produced in 48 h when the PCS was fermented in the simultaneous saccharification and co-fermentation mode using the hemicellulosic (1 g/L of total soluble sugar) and as well as the cellulosic (1 g/L of glucose equivalent) parts of PCS. In a control experiment, 0.48 g/L ethanol was obtained from 1 g/L of hemicellulosic PCS. It was concluded that the whole cell biocatalyst could convert both cellulosic and hemicellulosic substrates into ethanol in a single reactor. The developed C. cellulolyticum-E. coli whole cell biocatalyst also overcame the incompatible temperature problem of the frequently reported fungal-yeast systems.     

Cultivar variation in the effect of chlorsulfuron in depressing the uptake of copper in wheat

The application of herbicides has induced symptoms of nutrient deficiencies under some circumstances. This glasshouse study examined the effect of chlorsulfuron on the uptake and utilization of copper (Cu) in four cultivars of wheat plants (Triticum aestivum L. cvs. Kulin, Cranbrook, Gamenya and Bodallin) on a Cu-responsive soil. Application of chlorsulfuron depressed the concentration of Cu in wheat plants receiving either inadequate or adequate Cu. In plants with inadequate Cu supply, chlorsulfuron increased the severity of Cu deficiency. Shoot weight was markedly decreased by chlorsulfuron at all levels of Cu, through decreasing the number of tillers and the elongation of leaves. This decreased growth of shoots occurred prior to the effect on Cu concentration in tissues. The retranslocation of Cu in old tissues over time was unaffected by chlorsulfuron. In all wheat cultivars, the decreased growth of shoots were correlated with the concentration of Cu in the youngest fully emerged leaf blade with critical levels of 1.6−1.7 at day 25 and 0.9−1.0 μg g⁻¹ d. wt. at day 60. The application of chlorsulfuron tended to increase the critical level at day 25 but not at day 60. In addition, Kulin seems to be most, and Cranbrook least, sensitive to chlorsulfuron. This sensitivity was associated with the sensitivity of the cultivars to Cu deficiency. It is suggested that chlorsulfuron application induces Cu deficiency in wheat plants mainly due to effects on the uptake of Cu. This revised version was published online in June 2006 with corrections to the Cover Date.     

Variation due to Growth Environment in Alfalfa Yield, Cellulosic Ethanol Traits, and Paper Pulp Characteristics

Alfalfa (Medicago sativa L.) is a promising bioenergy and bioproduct feedstock because of its high yield, N-fixation capacity, potential for planting in rotation with corn (Zea mays L.), and valuable protein co-product (leaf meal). Our objective was to examine the effect of growth environment on biomass yield, cellulosic ethanol traits, and paper pulp fiber characteristics of alfalfa stems. Landscape position (summit and mild slope), season of harvest (four harvests per season), and multiple years (2005 and 2006) provided environmental variation. Alfalfa stem samples were analyzed for cell wall carbohydrate and lignin concentration. Stems were subjected to dilute acid pre-treatment, enzymatic saccharification, and pulping processes to measure relevant cellulosic ethanol and paper production traits. Landscape position was not a significant source of variation for yield or any biomass quality trait. Yields varied among harvests in 2005 (1,410–3,265 kg ha^?1) and 2006 (1,610–3,795 kg ha^?1). All cell wall, conversion test, and paper production traits exhibited year by harvest interactions with no clear pattern. Total carbohydrates and lignin ranged from 440 to 531 g?kg^?1 DM and from 113 to 161 g?kg^-1 DM, respectively. Release of cell wall sugars by the conversion test ranged widely (419 to 962 g?kg^?1 DM). Fiber traits were similarly variable with length and fine content ranging from 1.24 to 1.59 mm and from 15.2% to 21.9%, respectively. Utilizing alfalfa biomass for cellulosic ethanol and paper pulp production will involve dealing with significant feedstock quality variation due to growth environment.     

Sugarcane Internode Composition During Crop Development

Sugarcane sugar and bagasse can be utilized for the production of ethanol or other biofuels. A better understanding of the changes in composition with development along the stalk and with crop development will maximize the usage of sugarcane for this purpose. Two experiments were designed to elucidate internode composition changes during the growing season. In experiment 1, an internode of stalks of 5 modern cultivars were marked at the start of elongation, and then sampled every 1 to 2?weeks from July until October. Sugars were extracted and assayed, and a sequential detergent method was used to estimate hemicellulose, cellulose, and lignin contents. In experiment 2, internodes 1, 3, 5, 7, 9, and 11 down the stalk were sampled in late July (grand growth) and late September (ripening). Internode length, fresh weight, dry weight, water content, and sugar contents were determined as well as cell wall composition. Both experiments were repeated in 2?years. As internodes elongated, total sugar increased, and hemicellulose decreased as a proportion of neutral detergent fiber, while cellulose and lignin increased. After elongation, sucrose and lignin increased, and cellulose content decreased with internode age. The variability in cell wall composition among the five cultivars suggests that selection for desirable composition may be possible. In Experiment 2, hemicellulose contents were lower, and lignin and ash contents were higher at ripening than during grand growth. Delaying sugarcane harvest to maximize sucrose content may decrease bagasse suitability for cellulosic ethanol production because of the increased lignin content.