• Feedstocks Analysed at Celignis
    Corn Stover

Background on Corn Stover

Corn stover (also referred to as maize stover) is an agricultural residue similar to straw that is considered to be a substantial potential biorefining resource in the USA - an availability of 200 million tonnes has been estimated, equivalent to more than half the total amount of agricultural residues produced.

This resource is so large partially because of the amount of corn grown in the US, but also because of the amount of residues produced by the crop; it has been estimated that corn in the US produces 1.7 times more residue than other cereals.

This field harvest residue consists of the stalk (around 48% of the total dry mass), leaves (28%), and husks (8%) of the plant, as well as the cob (15%).

For stover and other straws, the amount of residue that needs to be left on the soil to preserve soil fertility and prevent against soil erosion is an important consideration. This is a complex issue and the field stover requirements will vary according to the location and factors such as climate, soil characteristics, tillage type and crop rotations.

Corn stover can be harvested wet or after drying on the field for a period of time. A dry harvest involves the residue being shredded and raked before being left to field dry and at a later point the stover is windrowed and baled. Since this is a multi-pass operation it can be costly and means that grain harvesting and residue collecting operations cannot be integrated. An alternative is to harvest the product wet in which case the shredding, windrowing and baling can take place on the same day as grain harvest with the first two operations possible from the grain harvester itself. The stover can then be ensiled to prevent storage losses.

The target of many researchers developing integrated harvesting practices is a one-pass system that allows for the simultaneous harvest of grain and stover while also leaving enough residue on the field to preserve soil fertility. There is also work on modifying the header of such an integrated combine so that the relative proportions of the stover fractions (cobs, stalks etc.) can be varied according to the end needs of the biorefinery.

Analysis of Corn Stover at Celignis



Celignis Analytical can determine the following properties of Corn Stover samples:



Lignocellulosic Properties of Corn Stover

Cellulose Content of Corn Stover

Each of the different corn stover fractions (stalk, leaves, husks, and cob) have different lignocellulosic compositions.

For example, the cellulose content is typically highest in the stalk, and less in the husks, lower still in the cob, and lowest in the leaves.

Furthermore the composition of these residues will also vary due to the variety of the plant and its productivity.

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Hemicellulose Content of Corn Stover

The total hemicellulose content, and its constituent sugars, varies greatly according to the stover fraction. For instance, xylan content is usually greatest in the cob whilst the husks tend to have more arabinose than in present in the other fractions.

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Lignin Content of Corn Stover

Each of the different corn stover fractions (stalk, leaves, husks, and cob) have different lignin contents.

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Starch Content of Corn Stover

The starch content of corn stover will vary according to the anatomical fraction of the plant. For instance, it will be higher in the cobs and lower in the stem.

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Uronic Acid Content of Corn Stover

Uronic acids can be present as side chains attached to the main backbone of hemicelluloses in corn stover. They concentrations of uronic acids tends to be greatest in the nodes, lower in the internodes, and at intermediate levels in the leaves. Corn cobs can also contain uronic acids.

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Enzymatic Hydrolysis of Corn Stover

We can undertake tests involving the enzymatic hydrolysis of Corn Stover. In these experiments we can either use a commercial enzyme mix or you can supply your own enzymes. We also offer analysis packages that compare the enzymatic hydrolysis of a pre-treated sample with that of the native original material.

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Bioenergy Properties of Corn Stover

Ash Content of Corn Stover

As with many other plants, the ash content is typically greatest in the leaf fraction of corn stover.

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Heating (Calorific) Value of Corn Stover

The heating value of corn stover will depend on the relative mass proportions of the different plant fractions and also on the moisture content of the feedstock.

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Ash Melting Behaviour of Corn Stover

Ash melting, also known as ash fusion and ash softening, can lead to slagging, fouling and corrosion in boilers which may reduce conversion efficiency. We can determine the ash melting behaviour of Corn Stover using our Carbolite CAF G5 BIO ash melting furnace. It can record the following temperatures:

Ash Shrinkage Starting Temperature (SST) - This occurs when the area of the test piece of Corn Stover ash falls below 95% of the original test piece area.

Ash Deformation Temperature (DT) - The temperature at which the first signs of rounding of the edges of the test piece occurs due to melting.

Ash Hemisphere Temperature (HT) - When the test piece of Corn Stover ash forms a hemisphere (i.e. the height becomes equal to half the base diameter).

Ash Flow Temperature (FT) - The temperature at which the Corn Stover ash is spread out over the supporting tile in a layer, the height of which is half of the test piece at the hemisphere temperature.



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Major and Minor Elements in Corn Stover

Examples of major elements that may be present in Corn Stover include potassium and sodium which are present in biomass ash in the forms of oxides. These can lead to fouling, ash deposition in the convective section of the boiler. Alkali chlorides can also lead to slagging, the fusion and sintering of ash particles which can lead to deposits on boiler tubes and walls.

We can also determine the levels of 13 different minor elements (such as arsenic, copper, and zinc) that may be present in Corn Stover.

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Analysis of Corn Stover for Anaerobic Digestion



Biomethane potential (BMP) of Corn Stover

At Celignis we can provide you with crucial data on feedstock suitability for AD as well as on the composition of process residues. For example, we can determine the biomethane potential (BMP) of Corn Stover. The BMP can be considered to be the experimental theoretical maximum amount of methane produced from a feedstock. We moniotor the volume of biogas produced allowing for a cumulative plot over time, accessed via the Celignis Database. Our BMP packages also involve routine analysis of biogas composition (biomethane, carbon dioxide, hydrogen sulphide, ammonia, oxygen). We also provide detailed analysis of the digestate, the residue that remains after a sample has been digested. Our expertise in lignocellulosic analysis can allow for detailed insight regarding the fate of the different biogenic polymers during digestion.



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Physical Properties of Corn Stover



Bulk Density of Corn Stover

At Celignis we can determine the bulk density of biomass samples, including Corn Stover, according to ISO standard 17828 (2015). This method requires the biomass to be in an appropriate form (chips or powder) for density determination.



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Particle Size of Corn Stover

Our lab is equipped with a Retsch AS 400 sieve shaker. It can accommodate sieves of up to 40 cm diameter, corresponding to a surface area of 1256 square centimetres. This allows us to determine the particle size distribution of a range of samples, including Corn Stover, by following European Standard methods EN 15149- 1:2010 and EN 15149-2:2010.



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Publications on Corn Stover By The Celignis Team

Gottumukkala L.D. Gorgens J.F (2016) Biobutanol production from lignocellulosics, Biofuels Production and future perspectives, Taylor & Francis group

Next-generation biofuels from renewable sources have gained interest among research investigators, industrialists, and governments due to major concerns on the volatility of oil prices, climate change, and depletion of oil reserves. Biobutanol has drawn signicant attention as an alternative transportation fuel due to its superior fuel properties over ethanol. e advantages of butanol are its high energy content, better blending with gasoline, less hydroscopic nature, lower volatility, direct use in convention engines, low corrosiveness, etc. Butanol production through (acetone, butanol, and ethanol) ABE fermentation is a well-established process, but it has several drawbacks like feedstock cost, strain degeneration, product toxicity, and low product concentrations. Lignocellulosic biomass is considered as the most abundant, renewable, low-cost feedstock for biofuels. Production of butanol from lignocellulosic biomass is more complicated due to the recalcitrance of feedstock and inhibitors generated during the pretreatment and hydrolysis process. Advanced fermentation and product recovery techniques are being researched to make biobutanol industrially viable.

V.P. Zambare, Lew P. Christopher (2012) Optimization of enzymatic hydrolysis of corn stover for improved ethanol production, Energy Exploration & Exploitation 30(2): 193-205

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Response surface methodology (RSM) was used to optimize the enzymatic hydrolysis of corn stover (CS), an abundant agricultural residue in the USA. A five-level, three-variable central composite design (CCD) was employed in a total of 20 experiments to model and evaluate the impact of pH (4.1–6.0), solids loadings (6.6–23.4%), and enzyme loadings (6.6?23.4 FPU g?1 DM) on glucose yield from thermo-mechanically extruded CS. The extruded CS was first hydrolyzed with the crude cellulase of Penicillium pinophilum ATCC 200401 and then fermented to ethanol with Saccharomyces cerevisiae ATCC 24860. Although all three variables had a significant impact, the enzyme loadings proved the most significant parameter for maximizing the glucose yield. A partial cubic equation could accurately model the response surface of enzymatic hydrolysis as the analysis of variance (ANOVA) showed a coefficient of determination (R2 ) of 0.82. At the optimal conditions of pH of 4.5, solids loadings of 10% and enzyme loadings of 20 FPU g?1 DM, the enzymatic hydrolysis of pretreated CS produced a glucose yield of 57.6% of the glucose maximum yield which was an increase of 10.4% over the non-optimized controls at zero-level central points. The predicted results based on the RSM regression model were in good agreement with the actual experimental values. The model can present a rapid means for estimating lignocellulose conversion yields within the selected ranges.

V.P. Zambare, A. Bhalla, K. Muthukumarappan, R. Sani, L. Christopher (2011) Bioprocessing of agricultural residues to ethanol utilizing a cellulolytic extremophile, Extremophiles 15: 611-618

Link

A recently discovered thermophilic isolate, Geobacillus sp. R7, was shown to produce a thermostable cellulase with a high hydrolytic potential when grown on extrusion-pretreated agricultural residues such corn stover and prairie cord grass. At 70°C and 15–20% solids, the thermostable cellulase was able to partially liquefy solid biomass only after 36 h of hydrolysis time. The hydrolytic capabilities of Geobacillus sp. R7 cellulase were comparable to those of a commercial cellulase. Fermentation of the enzymatic hydrolyzates with Saccharomyces cerevisiae ATCC 24860 produced ethanol yields of 0.45–0.50 g ethanol/g glucose with more than 99% glucose utilization. It was further demonstrated that Geobacillus sp. R7 can ferment the lignocellulosic substrates to ethanol in a single step that could facilitate the development of a consolidated bioprocessing as an alternative approach for bioethanol production with outstanding potential for cost reductions.

Vasudeo Zambare, Archana Zambare, Kasiviswanathan Muthukumarappan, Lew Christopher (2011) Biochemical characterization of thermophilic lignocellulose degrading enzymes and their potential for biomass bioprocessing, International Journal of Energy and Environment 2(1): 99-112

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A thermophilic microbial consortium (TMC) producing hydrolytic (cellulolytic and xylanolytic) enzymes was isolated from yard waste compost following enrichment with carboxymethyl cellulose and birchwood xylan. When grown on 5% lignocellulosic substrates (corn stover and prairie cord grass) at 600 C, the thermophilic consortium produced more xylanase (up to 489 U/l on corn stover) than cellulase activity (up to 367 U/l on prairie cord grass). Except for the carboxymethyl cellulose-enriched consortium, thermo-mechanical extrusion pretreatment of these substrates had a positive effect on both activities with up to 13% and 21% increase in the xylanase and cellulase production, respectively. The optimum temperatures of the crude cellulase and xylanase were 600 C and 700 C with half-lives of 15 h and 18 h, respectively, suggesting higher thermostability for the TMC xylanase. Sodium dodecyl sulfatepolyacrylamide gel electrophoresis of the crude enzyme exhibited protein bands of 25-77 kDa with multiple enzyme activities containing 3 cellulases and 3 xylanases. The substrate specificity declined in the following descending order: avicel>birchwood xylan>microcrystalline cellulose>filter paper>pine wood saw dust>carboxymethyl cellulose. The crude enzyme was 77% more active on insoluble than soluble cellulose. The Km and Vmax values were 36.49 mg/ml and 2.98 U/mg protein on avicel (cellulase), and 22.25 mg/ml and 2.09 U/mg protein, on birchwood xylan (xylanase). A total of 50 TMC isolates were screened for cellulase and xylanase secretion on agar plates. All single isolates showed significantly lower enzyme activities when compared to the thermophilic consortia. This is indicative of the strong synergistic interactions that exist within the thermophilic microbial consortium and enhance its hydrolytic capabilities. It was further demonstrated that the thermostable enzyme-generated lignocellulosic hydrolyzates can be fermented to bioethanol by a recombinant strain of Escherichia coli. This could have important implications in the enzymatic breakdown of lignocellulosic biomass for the establishment of a robust and cost-efficient process for production of cellulosic ethanol. To the best of our knowledge, this work represents the first report in literature on biochemical characterization of lignocellulose-degrading enzymes from a thermophilic microbial consortium.

V. P. Zambare (2010) Solid state fermentation production of cellulase from Bacillus sp, International Journal of BioScience, Agriculture and Technology 2(1): 1-6

Link

Bacillus sp. was cultured in solid-state fermentation (SSF) of wheat straw to produce cellulase. The fermented biomass was harvested after 36 h of SSF at pH 8 and temperature 400C. It was filtered and centrifuged at 10,000 rpm at 4 0C and supernatant was collected as crude enzyme extract. Maximum activity of cellulase (3.775±0.13U/ml) was obtained after fermentation of wheat straw (10g) medium containing 0.2g soybean meal, 0.04g corn steep liquor (CSL), 80% moisture content (mineral salt medium, pH 8), 2-mL inoculum, and temperature 40 0C. SSF was found to be more productive than submerged fermentation (SmF) in terms of cellulase yields. The partial purification of cellulase was carried out through (NH4)2SO4 precipitation. The partially purified enzyme produced under SSF had molecular weight of 35 and 45kDa. It was active in a broad pH (4-10) and temperature range (25-550C). The optimum, pH and temperature of Bacillus cellulase were pH 5 and 450C, respectively. At 500C and 600C, the half lives of the partially purified cellulase were 194 and 163 min, respectively. All the results indicated that the Bacillus sp. had a promising application of treatment of agro-wastes and cellulase from Bacillus sp. could be potentially used in biofuel industries.

Vasudeo Zambare (2010) Solid state fermentation of Aspergillus oryzae for glucoamylase production on agro residues, International Journal of Life Sciences 4: 16-25

Link

Glucoamylase is a well recognized amylolytic enzyme used in food industry, which is generally produced by Aspergillus genus under solid-state fermentation (SSF). This study presents production of glucoamylase by Aspergillus oryzae on the solid surface of rice husk, wheat bran, rice bran, cotton seed powder, corn steep solids, bagasse powder, coconut oil cake, and groundnut oil cake as substrates. Optimization of the SSF media and parameters resulted in a 24% increase in the glucoamylase activity. Optimum glucoamylase production (1986 µmoles of glucose produced per minute per gram of dry fermented substrate) was observed on wheat bran supplemented with 1%, (w/w) starch, 0.25%, (w/w) urea at pH 6, 100%, (v/w) initial moisture and 300C after incubation 120 hrs. Therefore, A. oryzae can be useful in bioprocessing application for saccharification of agro-residues.





Examples of Other Feedstocks Analysed at Celignis



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