Project Research Publications: Conversion

  • Agyin-Birikorang, S., G.A. O’Connor and J.E. Erickson (2013). Sustainable nutrient management package for cost-effective bioenergy biomass production. Journal of Plant Nutrition 36(12):1881-1900. DOI:10.1080/01904167.2013.818154

      • Abstract:

      • Commercial fertilizer (particularly nitrogen) costs account for a substantial portion of the total production costs of cellulosic biomass and can be a major obstacle to biofuel production. In a series of greenhouse studies, we evaluated the feasibility of co-applying Gibberellins (GA) and reduced nitrogen (N) rates to produce a bioenergy crop less expensively. In a preliminary study, we determined the minimum combined application rates of GA and N required for efficient biomass (sweet sorghum, Sorghum bicolor) production. Co-application of 75 kg ha−1 (one-half of the recommended N rate for sorghum) and a modest GA rate of 3 g ha−1 optimized dry matter yield (DMY) and N and phosphorus (P) uptake efficiencies, resulting in a reduction of N and P leaching. Organic nutrient sources such as manures and biosolids can be substituted for commercial N fertilizers (and incidentally supply P) to further reduce the cost of nutrient supply for biomass production. Based on the results of the preliminary study, we conducted a second greenhouse study using sweet sorghum as a test bioenergy crop. We co-applied organic sources of N (manure and biosolids) at 75 and 150 kg PAN ha−1 (representing 50 and 100% N rate respectively) with 3 g GA ha−1. In each batch of experiment, the crop was grown for 8 wk on Immokalee fine sand of minimal native fertility. After harvest, sufficient water was applied to soil in each pot to yield ∼1.5 L (∼0.75 pore volume) of leachate, and analyzed for total N and soluble reactive P (SRP). The reduced (50%) N application rate, together with GA, optimized biomass production. Application of GA at 3 g ha−1, and the organic sources of N at 50% of the recommended N rate, decreased nutrient cost of producing the bioenergy biomass by ∼$ 375 ha−1 (>90% of total nutrient cost), and could reduce offsite N and P losses from vulnerable soils.

      • http://dx.doi.org/10.1080/01904167.2013.8181547

  • Agyin-Birikorang, S., G.A. O’Connor, P.C. Pullammanappallil and G.R. Mohan (2013). Recovery of essential plant nutrients from biofuel residual. Journal of Sustainable Bioenergy Systems 3(2):149-159.

      • Abstract:

      • Essential plant nutrients contained in residues and wastes generated during biofuel processing can be recovered for further production of bioenergy biomass. The objective of this study was to determine the relative agronomic efficiency of “processed” biofuel residual (PBR). Liquid biofuel residual was “processed” by precipitating phosphate and ammonium in the residual with magnesium into a struvite-like material. Then, in a series of greenhouse experiments, we evaluated the fertility potential of PBR, using sweet sorghum (Sorghum bicolor (L.) Moench), as a test bioenergy crop. We compared the agronomic effectiveness of PBR to inorganic commercial fertilizers, biosolids, and poultry manure as nutrient sources. The sources were either applied alone or in combination with supplemental essential plant nutrients (S, K, Mg, and micronutrients). In each of the greenhouse experiments, the crop was grown for 12 wk on soil of minimal native fertility. After each harvest, sufficient water was applied to the soil in each pot over a 6-wk period to yield ∼2 L (∼one pore volume) of leachate to assess potential total N and soluble reactive phosphorus (SRP) losses. Dry matter yields from the PBR treatment applied alone were significantly greater than yields from inorganic fertilizers, biosolids, and poultry manure treatments applied alone, and similar to yields obtained when the supplemental essential plant nutrients were added to the inorganic fertilizer, biosolids, and manure treatments. Leachate N and SRP concentrations from the PBR treatment were significantly lower than in the treatments with inorganic fertilizers, poultry manure, and biosolids. We conclude that PBR can substitute for inorganic fertilizers and other organic sources of plant nutrients to produce bioenergy biomass cheaply, without causing offsite N and P losses in vulnerable soils.

      • http://dx.doi.org/10.4236/jsbs.2013.32021

  • Castro, E., Nieves, I.U., Mullinnix, M.T., Sagues, W.J., Hoffman, R.W., Fernández-Sandoval, M.T., Tian, Z., Rockwood, D.L., Tamang, B. and Ingram, L.O. (2014). Optimization of dilute-phosphoric-acid steam pretreatment of Eucalyptus benthamii for biofuel production. Applied Energy 125:76-83. DOI: 10.1016/j.apenergy.2014.03.047

      • Abstract:

      • This work deals with the production of ethanol from phosphoric acid-impregnated, steam-exploded Eucalyptus benthamii. The whole conversion process, addressing pretreatment, enzymatic hydrolysis of the whole slurry, and fermentation of both C5 and C6-sugars including a presaccharification step, is covered in this study.

        Two separate models were developed to maximize sugar content and minimize inhibitor concentrations, resulting in xylose yields of ∼50% and ∼60% after pretreatment. In addition, a Liquefaction plus Simultaneous Saccharification and co-Fermentation (L+SScF) was performed to compare the fermentability of the resulting pretreated biomass. After the 6-h liquefaction step using the Cellic CTec2 enzyme from Novozyme and 10% DW pretreated biomass, the total sugar concentration in the slurry was 47 g/L and 51 g/L for the two conditions respectively. Enzymatic hydrolysis continued during fermentation using an ethanologenic derivative of Escherichia coli KO11. The sugars were completely consumed in 96 h with product yields of 0.217 and 0.243 g ethanol/g DW biomass for each condition, respectively. These yields are equivalent to 275 and 304 L/tonne DW, confirming the effectiveness of the L+SScF process using phosphoric-acid-pretreated Eucalyptus.

      • https://doi.org/10.1016/j.apenergy.2014.03.047

  • Chauliac, D., Pullammanappallil, P.C., Ingram, L.O. and Shanmugam, K.T. (2015). Removing chiral contamination of lactate solutions by selective metabolism of the D-enantiomer. Biotechnology Letters 37(12):2411-2418. DOI: 10.1007/s10529-015-1924-z

      • Abstract:

      • Objective

        A bio-based process is appealing for purification of l-lactic acid, the major enantiomer of polylactic acid syrup, generated by thermochemical processes at the end of life of PLA-based plastics, from its chiral impurity, d-lactic acid, before reuse.

        Results

        Polylactic acid (PLA), a renewable alternative to petroleum-derived plastics, contains a mixture of l- and d-lactic acid (LA) isomers with the l-isomer dominating (up to 95 %). A novel bio-based process was developed to produce chirally pure l-LA from syrup produced during recycling of PLA-plastics. This process utilizes an engineered Escherichia coli (strain DC1001) containing novel gene deletions (lld, ykg) that eliminated the oxidative metabolism of l-lactate, leaving the membrane-bound d-lactate dehydrogenases to selectively metabolize the d-isomer. Strain DC1001 removed 8.7 g d-lactate l−1 from a PLA-syrup containing 135 g total lactic acid l−1 in 24 h. Average rates of removal of d-lactic acid were 0.25 g d-lactate h−1 (g cell dry weight)−1 and 0.36 g d-lactate l−1 h−1.

        Conclusion

        Bio-based purification of PLA-syrup utilizing E. coli strain DC1001 is an attractive process step during recycling of PLA-plastics. This selective oxidation process can also be used to remove chiral contamination of l-lactate in medical applications.

      • https://doi.org/10.1007/s10529-015-1924-z

  • Geddes, R.D., Wang, X, Yomano, L.P., Miller, E.N., Zheng, H., Shanmugam, K.T. and Ingram, L.O. (2014). Polyamine transporters and polyamines increase furfural tolerance during xylose fermentation with ethanologenic Escherichia coli strain LY180. Applied and Environmental Microbiology 80(19):5955-5964. DOI: 10.1128/AEM.01913-14

      • Abstract:

      • Expression of genes encoding polyamine transporters from plasmids and polyamine supplements increased furfural tolerance (growth and ethanol production) in ethanologenic Escherichia coli LY180 (in AM1 mineral salts medium containing xylose). This represents a new approach to increase furfural tolerance and may be useful for other organisms. Microarray comparisons of two furfural-resistant mutants (EMFR9 and EMFR35) provided initial evidence for the importance of polyamine transporters. Each mutant contained a single polyamine transporter gene that was upregulated over 100-fold (microarrays) compared to that in the parent LY180, as well as a mutation that silenced the expression of yqhD. Based on these genetic changes, furfural tolerance was substantially reconstructed in the parent, LY180. Deletion of potE in EMFR9 lowered furfural tolerance to that of the parent. Deletion of potE and puuP in LY180 also decreased furfural tolerance, indicating functional importance of the native genes. Of the 8 polyamine transporters (18 genes) cloned and tested, half were beneficial for furfural tolerance (PotE, PuuP, PlaP, and PotABCD). Supplementing AM1 mineral salts medium with individual polyamines (agmatine, putrescine, and cadaverine) also increased furfural tolerance but to a smaller extent. In pH-controlled fermentations, polyamine transporter plasmids were shown to promote the metabolism of furfural and substantially reduce the time required to complete xylose fermentation. This increase in furfural tolerance is proposed to result from polyamine binding to negatively charged cellular constituents such as nucleic acids and phospholipids, providing protection from damage by furfural.

      • https://doi.org/10.1128/AEM.01913-14

  • Geddes, R., Shanmugam, K.T. and Ingram, L.O. (2015). Combining treatments to improve the fermentation of sugarcane bagasse hydrolysates by ethanologenic Escherichia coli LY180. Bioresource Technology 189:15-22. DOI: 10.1016/j.biortech.2015.03.141

      • Abstract:

      • Inhibitory side products from dilute acid pretreatment is a major challenge for conversion of lignocellulose into ethanol. Six strategies to detoxify sugarcane hydrolysates were investigated alone, and in combinations (vacuum evaporation of volatiles, high pH treatment with ammonia, laccase, bisulfite, microaeration, and inoculum size). High pH was the most beneficial single treatment, increasing the minimum inhibitory concentration (measured by ethanol production) from 15% (control) to 70% hydrolysate. Combining treatments provided incremental improvements, consistent with different modes of action and multiple inhibitory compounds. Screening toxicity using tube cultures proved to be an excellent predictor of relative performance in pH-controlled fermenters. A combination of treatments (vacuum evaporation, laccase, high pH, bisulfite, microaeration) completely eliminated all inhibitory activity present in hydrolysate. With this combination, fermentation of hemicellulose sugars (90% hydrolysate) to ethanol was complete within 48 h, identical to the fermentation of laboratory xylose (50 g/L) in AM1 mineral salts medium (without hydrolysate).

      • https://doi.org/10.1016/j.biortech.2015.03.141

  • Gubicza, K., Nieves, I.U., Sagues, W.J., Barta, Z., Shanmugam, K.T. and Ingram, L.O. (2016). Techno-economic analysis of ethanol production from sugarcane bagasse using a Liquefaction plus Simultaneous Saccharification and co-Fermentation process. Bioresource Technology 208:42-48. DOI: 10.1016/j.biortech.2016.01.093

      • Abstract:

      • A techno-economic analysis was conducted for a simplified lignocellulosic ethanol production process developed and proven by the University of Florida at laboratory, pilot, and demonstration scales. Data obtained from all three scales of development were used with Aspen Plus to create models for an experimentally-proven base-case and 5 hypothetical scenarios. The model input parameters that differed among the hypothetical scenarios were fermentation time, enzyme loading, enzymatic conversion, solids loading, and overall process yield. The minimum ethanol selling price (MESP) varied between 50.38 and 62.72 US cents/L. The feedstock and the capital cost were the main contributors to the production cost, comprising between 23–28% and 40–49% of the MESP, respectively. A sensitivity analysis showed that overall ethanol yield had the greatest effect on the MESP. These findings suggest that future efforts to increase the economic feasibility of a cellulosic ethanol process should focus on optimization for highest ethanol yield.

      • https://doi.org/10.1016/j.biortech.2016.01.093

  • Hafner, K.D. and Wilkie, A.C. (2015). Cellulosic Ethanol Stillage as a Cultivation Medium for Spirulina. Abstract. 16th Annual Soil and Water Science Research Forum, Gainesville, Florida, September 17, 2015.

      • Abstract:

      • Algae are high-yielding plants and a potential alternative to conventional fossil fuels that can alleviate the greenhouse effect while simultaneously treating wastewater. Cultivating algae requires high nitrogen inputs to sustain growth and produce feedstock biomass, providing a possible bioremediation option for high-ammonia wastewaters such as stillage from cellulosic ethanol production. Cultivating algae on stillage can help to offset the energy consumed in the pretreatment and distillation operations of bioethanol production as it combines nutrient removal and algal production for potential use as a biofuel feedstock. Thus, this creates a sustainable, closed-loop process. The objective of this study was to determine the growth and remediation potential of Spirulina, a filamentous, blue-green algae with high biomass productivity, on sugarcane bagasse stillage. Cultures were prepared with 10% inoculum in 125 mL Erlenmeyer flasks (50 mL active volume). The treatment mediums consisted of 100% thin stillage (negative control), Modified Spirulina Medium (positive control), and 2%, 5% and 10% concentrations of thin stillage with 1% (v/v) sodium bicarbonate. The cultures were cultivated for 72 hours under 120 µmol photons/m 2/s fluorescent lighting on a 24:0 (light:dark) photoperiod. Cells were subjected to moderate mixing (120 rpm) provided by a mechanical shaker. Algal growth was monitored spectrophotometrically using absorbance at 680 nm. The data revealed that thin stillage in dilutions is promising for cultivating Spirulina. Compared to algal growth in Modified Spirulina Medium (2.31 g/L), the 2%, 5% and 10% thin stillage mediums exhibited biomass yields of 1.60, 1.96, and 1.93 g/L, respectively.

      • http://soils.ifas.ufl.edu/media/soilsifasufledu/sws-main-site/pdf/research/forum/Judged_Poster_Presentation_Abstracts_9_10_15.pdf

  • Jairam S., Bucklin, R., Correll, M., Sakthivel, T.S., Seal, S., Truett, J. and Tong, Z. (2016). UV resistance of polystyrene co-butyl acrylate (PSBA) encapsulated lignin-saponite nanohybrid composite film. Materials and Design 90:151-156. DOI: 10.1016/j.matdes.2015.10.118

      • Abstract:

      • Protecting biological systems and organic materials from high ultraviolet (UV) radiation is a significant issue for engineering materials in both terrestrial and space applications. In this study, the synthesized composites of polystyrene butyl acrylate (PSBA) encapsulated lignin based nanohybrid were evaluated as UV resistant cladding materials for both applications. The UV transmittance of the composite films and their physical, thermal, and chemical degradation after prolonged exposure to high UV radiation (10 Earth days and 30 Mars equivalent days) at low temperatures of 3.4 °C were examined. The results showed that the composite films blocked the UV radiation and there was no significant effect of UV radiation on film integrity, mechanical and thermal properties of the composite films when they were exposed to high intensity UV radiation (132 W/m2) within 24 h. Further, FTIR spectra of the composite films before and after UV radiation demonstrated that the films did not have notable oxidation degradation, although the glass transition temperature was slightly decreased with increased exposure time.

      • https://doi.org/10.1016/j.matdes.2015.10.118

  • Lovato, T.B. and Wilkie, A.C. (2014). Characterization of cellulosic ethanol stillage and use as an algal growth medium. Abstracts of the 2014 Florida Energy Systems Consortium Workshop, Gainesville, Florida, May 12-13, 2014.

      • Abstract:

      • Cellulosic ethanol is a potential alternative to petroleum-based fuels and, because it is created from lignocellulose found in woody plant materials, it does not compete directly with food production, unlike corn-based ethanol. One obstacle in cellulosic ethanol production is dealing with the stillage by-product that is typically high in nutrients and chemical oxygen demand. Growing algae for biofuels and high-value pigments like β-carotene or astaxanthin requires high nitrogen inputs to sustain growth and produce more biomass, providing a possible bioremediation option for stillage. Various treatment options exist for stillage including anaerobic digestion that has been shown to substantially reduce chemical oxygen demand. In addition, algae can serve as a potential feedstock for anaerobic digestion and the subsequent creation of biogas introducing a conceivable link between multiple bioenergy prospects. The objective of this study was to cultivate algae using stillage as a nutrient source. Sugarcane bagasse stillage from the UF-IFAS Stan Mayfield Biorefinery Pilot Plant was characterized, measuring pH, electrical conductivity, light transmission, total nitrogen, total ammoniacal nitrogen, total and soluble phosphorus, and total and soluble chemical oxygen demand. A strain of the microfilamentous cyanobacterial algae Spirulina sp. was isolated using a modified Spirulina standard culture medium replacing nitrate ion with ammonium as the nitrogen source. This culture was then inoculated into flasks with 2% dilutions of stillage supplemented with Spirulina nutrients using Spirulina’s preferred sodium bicarbonate as a carbon source. The experimental group using the 2% stillage dilutions produced more biomass than the control under the same conditions, as measured by optical density absorbance readings. Lipid analysis using nuclear magnetic resonance based against a triolein standard showed that the algae biomass had low neutral oil content and was not ideal for algal biodiesel production. However, the biomass growth under experimental conditions points towards a potential use of stillage as a nutrient source in algae production. The algal biomass can be utilized as feedstock for biogas production via anaerobic digestion.

      • http://floridaenergy.ufl.edu/fesc-conferences/florida-energy-systems-consortium-fesc-workshop/2014-poster-session/additional-posters/

  • Mondala, A., Hernandez, R., French, T., Green, M., McFarland, L. and Ingram, L. (2015). Enhanced microbial oil production by activated sludge microorganisms from sugarcane bagasse hydrolysate. Renewable Energy 78:114-118. DOI: 10.1016/j.renene.2014.12.073

      • Abstract:

      • The use of sugarcane bagasse hydrolyzate as a carbon source for enhanced oil production by activated sludge microbial cultures was investigated. Cultivation of raw activated sludge inoculum using pure xylose as carbon source was necessary prior to bagasse hydrolyzate feeding for microbial acclimation to this pentose sugar, the major component of the hydrolyzate. Lipid contents from 40 to 47% (dry cell weight) were achieved under high C:N ratio following bagasse hydrolyzate feeding; however nutrient supplementation was found to be necessary in order to maintain viable cell biomass levels (>10 g/L) to achieve a high lipid titer (7.62 g/L). Hence, a process involving sequential batch feeding of hydrolyzate with and without nutrients was proposed and simulated using the Logistic and Luedeking–Piret models. Analysis of the product lipids showed up to 50% saponifiable fractions and dominance of C16 and C18 fatty acids, demonstrating their suitability as biofuel feedstock.

      • https://doi.org/10.1016/j.renene.2014.12.073

  • Ou, M.S., Awasthi, D., Nieves, I., Wang, L., Erickson, J., Vermerris, W., Ingram, L.O. and Shanmugam, K.T. (2016). Sweet sorghum juice and bagasse as feedstocks for the production of optically pure lactic acid by native and engineered Bacillus coagulans strains. BioEnergy Research 9(1):123-131. DOI: 10.1007/s12155-015-9670-6

      • Abstract:

      • Sweet sorghum is a bioenergy crop that produces large amounts of soluble sugars in its stems (3–7 Mg ha−1) and generates significant amounts of bagasse (15–20 Mg ha−1) as a lignocellulosic feedstock. These sugars can be fermented not only to biofuels but also to bio-based chemicals. The market potential of the latter may be higher given the current prices of petroleum and natural gas. The yield and rate of production of optically pure d-(−)- and l-(+)-lactic acid as precursors for the biodegradable plastic polylactide was optimized for two thermotolerant Bacillus coagulans strains. Strain 36D1 fermented the sugars in unsterilized sweet sorghum juice at 50 °C to l-(+)-lactic acid (∼150 g L−1; productivity, 7.2 g L−1 h−1). B. coagulans strain QZ19-2 was used to ferment sorghum juice to d-(−)-lactic acid (∼125 g L−1; productivity, 5 g L−1 h−1). Carbohydrates in the sorghum bagasse were also fermented after pretreatment with 0.5 % phosphoric acid at 190 °C for 5 min. Simultaneous saccharification and co-fermentation of all the sugars (SScF) by B. coagulans resulted in a conversion of 80 % of available carbohydrates to optically pure lactic acid depending on the B. coagulans strain used as the microbial biocatalyst. Liquefaction of pretreated bagasse with cellulases before SScF (L + SScF) increased the productivity of lactic acid. These results show that B. coagulans is an effective biocatalyst for fermentation of all the sugars present in sweet sorghum juice and bagasse to optically pure lactic acid at high titer and productivity as feedstock for bio-based plastics.

      • https://doi.org/10.1007/s12155-015-9670-6

  • Riley, M.K. and Vermerris, W. (2017) Recent advances in nanomaterials for gene delivery—A review. Nanomaterials 7(5):94. DOI: 10.3390/nano7050094

      • Abstract:

      • With the rapid development of nanotechnology in the recent decade, novel DNA and RNA delivery systems for gene therapy have become available that can be used instead of viral vectors. These non-viral vectors can be made of a variety of materials, including inorganic nanoparticles, carbon nanotubes, liposomes, protein and peptide-based nanoparticles, as well as nanoscale polymeric materials. They have as advantages over viral vectors a decreased immune response, and additionally offer flexibility in design, allowing them to be functionalized and targeted to specific sites in a biological system with low cytotoxicity. The focus of this review is to provide an overview of novel nanotechnology-based methods to deliver DNA and small interfering RNAs into biological systems.

      • https://doi.org/10.3390/nano7050094

  • Sawisit, A., Jantama, K., Zheng, H., Yomano, L.P., York, S.W., Shanmugam, K.T. and Ingram, L.O. (2015). Mutation in galP improved fermentation of mixed sugars to succinate using engineered Escherichia coli AS 1600a and AM1 mineral salts medium. Bioresource Technology 193:433-441. DOI: 10.1016/j.biortech.2015.06.108

      • Abstract:

      • Escherichia coli KJ122 was engineered to produce succinate from glucose using the wild type GalP for glucose uptake instead of the native phosphotransferase system (ptsI mutation). This strain now ferments 10% xylose poorly. Mutants were selected by serial transfers in AM1 mineral salts medium with 10% xylose. Clones from this population all exhibited a similar improvement, co-fermentation of an equal mixture of xylose and glucose. One of these, AS1600a, produced 84.26 ± 1.37 g/L succinate, equivalent to that produced by the parent (KJ122) from 10% glucose (85.46 ± 1.78 g/L). AS1600a was sequenced and found to contain a mutation in galactose permease (GalP, G236D). This mutation was shown to be responsible for the improvement in fermentation using KJΔgalP as the host and expression vectors with native galP and with mutant galP*. Strain AS1600a and KJΔgalP(pLOI5746; galP*) also co-fermented a mixture of glucose, xylose, arabinose, and galactose in sugarcane bagasse hydrolysate using mineral salts medium.

      • https://doi.org/10.1016/j.biortech.2015.06.108

  • Scully, E.D., Gries, T., Funnell-Harris, D.L., Xin, Z., Kovacs, F.A., Vermerris, W. and Sattler, S.E. (2016). Characterization of novel Brown midrib 6 mutations affecting lignin biosynthesis in sorghum. Journal of Integrative Plant Biology 58(2):136-149. DOI: 10.1111/jipb.12375

      • Abstract:

      • The presence of lignin reduces the quality of lignocellulosic biomass for forage materials and feedstock for biofuels. In C4 grasses, the brown midrib phenotype has been linked to mutations to genes in the monolignol biosynthesis pathway. For example, the Bmr6 gene in sorghum (Sorghum bicolor) has been previously shown to encode cinnamyl alcohol dehydrogenase (CAD), which catalyzes the final step of the monolignol biosynthesis pathway. Mutations in this gene have been shown to reduce the abundance of lignin, enhance digestibility, and improve saccharification efficiencies and ethanol yields. Nine sorghum lines harboring five different bmr6 alleles were identified in an EMS-mutagenized TILLING population. DNA sequencing of Bmr6 revealed that the majority of the mutations impacted evolutionarily conserved amino acids while three-dimensional structural modeling predicted that all of these alleles interfered with the enzyme's ability to bind with its NADPH cofactor. All of the new alleles reduced in vitro CAD activity levels and enhanced glucose yields following saccharification. Further, many of these lines were associated with higher reductions in acid detergent lignin compared to lines harboring the previously characterized bmr6-ref allele. These bmr6 lines represent new breeding tools for manipulating biomass composition to enhance forage and feedstock quality.

      • https://doi.org/10.1111/jipb.12375

  • Shi, J., O’Connor, G.A. and Wilkie, A.C. (2014). Reuse of cellulosic bioethanol residuals. Abstracts of the 2014 Florida Energy Systems Consortium Workshop, Gainesville, Florida, May 12-13, 2014.

      • Abstract:

      • Increasing demand for fossil fuels, coupled with diminishing fossil fuel reserves, create an urgent need to develop renewable energy sources as a replacement for fossil fuels. Biofuel systems have emerged as one such alternative to fossil fuels. First-generation biofuels are made from sugary or starchy plants and have generated food security concerns. The development of second-generation biofuels emphasizes the use of non-edible (cellulosic) feedstocks and green chemical technologies. However, the commercialization of second-generation biofuel is held up by technical as well as economic issues. Many studies focus on enhancing technology to improve ethanol productivity, while very few studies recognize the importance of the byproducts associated with cellulosic ethanol production. Beneficial reuse of these residues could provide cost savings as well as lessen the negative environmental impacts imposed by chemical fertilizers. An estimated 20 liters of stillage byproduct is generated for every liter of ethanol produced, so an effective solution for byproduct treatment is necessary when considering large-scale production of cellulosic ethanol in the future. This study critically evaluates the possible treatments for residues from cellulosic ethanol production and discusses the potential environmental and economic impacts of byproduct land application. The first step is to characterize and compare the stillage from different types of feedstocks. Treatments applied to first-generation bioethanol residues may or may not work on cellulosic ethanol residuals. Different initial chemical components of cellulosic feedstock, combined with the various pretreatment processes, may also result in byproducts that require different types of treatment prior to reuse management. Land application has been considered as one of the possible approaches for final disposal of cellulosic ethanol byproduct. Several studies have reported positive effects of land application such as improving soil structure, reducing soil erosion, increasing soil organic matter content, and reducing nutrient leaching. These potential benefits as well as the effects of these residuals on crop yields, especially biofuel feedstocks, are being evaluated as part of this research.

      • http://floridaenergy.ufl.edu/fesc-conferences/florida-energy-systems-consortium-fesc-workshop/2014-poster-session/additional-posters/

  • Shi, A., Zheng, H., Yomano, L.P., York, S.W., Shanmugam, K.T. and Ingram, L.O. (2016). Plasmidic expression of nemA and yafC* increased resistance of ethanologenic Escherichia coli LY180 to nonvolatile side products from dilute acid treatment of sugarcane bagasse and artificial hydrolysate. Applied and Environmental Microbiology 82(7):2137-2145. DOI: 10.1128/AEM.03488-15n

      • Abstract:

      • Hydrolysate-resistant Escherichia coli SL100 was previously isolated from ethanologenic LY180 after sequential transfers in AM1 medium containing a dilute acid hydrolysate of sugarcane bagasse and was used as a source of resistance genes. Many genes that affect tolerance to furfural, the most abundant inhibitor, have been described previously. To identify genes associated with inhibitors other than furfural, plasmid clones were selected in an artificial hydrolysate that had been treated with a vacuum to remove furfural. Two new resistance genes were discovered from Sau3A1 libraries of SL100 genomic DNA: nemA (N-ethylmaleimide reductase) and a putative regulatory gene containing a mutation in the coding region, yafC*. The presence of these mutations in SL100 was confirmed by sequencing. A single mutation was found in the upstream regulatory region of nemR (nemRA operon) in SL100. This mutation increased nemA activity 20-fold over that of the parent organism (LY180) in AM1 medium without hydrolysate and increased nemA mRNA levels >200-fold. Addition of hydrolysates induced nemA expression (mRNA and activity), in agreement with transcriptional control. NemA activity was stable in cell extracts (9 h, 37°C), eliminating a role for proteinase in regulation. LY180 with a plasmid expressing nemA or yafC* was more resistant to a vacuum-treated sugarcane bagasse hydrolysate and to a vacuum-treated artificial hydrolysate than LY180 with an empty-vector control. Neither gene affected furfural tolerance. The vacuum-treated hydrolysates inhibited the reduction of N-ethylmaleimide by NemA while also serving as substrates. Expression of the nemA or yafC* plasmid in LY180 doubled the rate of ethanol production from the vacuum-treated sugarcane bagasse hydrolysate.

      • https://doi.org/10.1128/AEM.03488-15

  • Wang, J. and Vermerris, W. (2016). Antimicrobial nanomaterials derived from natural products - A review. Materials 9(4):255. DOI: 10.3390/ma9040255

      • Abstract:

      • Modern medicine has relied heavily on the availability of effective antibiotics to manage infections and enable invasive surgery. With the emergence of antibiotic-resistant bacteria, novel approaches are necessary to prevent the formation of biofilms on sensitive surfaces such as medical implants. Advances in nanotechnology have resulted in novel materials and the ability to create novel surface topographies. This review article provides an overview of advances in the fabrication of antimicrobial nanomaterials that are derived from biological polymers or that rely on the incorporation of natural compounds with antimicrobial activity in nanofibers made from synthetic materials. The availability of these novel materials will contribute to ensuring that the current level of medical care can be maintained as more bacteria are expected to develop resistance against existing antibiotics.

      • https://doi.org/10.3390/ma9040255

  • Wang, L., Ou, M.S., Nieves, I., Erickson, J.E., Vermerris, W., Ingram, L.O. and Shanmugam, K.T. (2015). Fermentation of sweet sorghum derived sugars to butyric acid at high titer and productivity by a moderate thermophile Clostridium thermobutyricum at 50oC. Bioresource Technology 198:533-539. DOI: 10.1016/j.biortech.2015.09.062

      • Abstract:

      • In this study, a moderate thermophile Clostridium thermobutyricum is shown to ferment the sugars in sweet sorghum juice treated with invertase and supplemented with tryptone (10 g L−1) and yeast extract (10 g L−1) at 50 ° C to 44 g L−1 butyrate at a calculated highest volumetric productivity of 1.45 g L−1 h−1 (molar butyrate yield of 0.85 based on sugars fermented). This volumetric productivity is among the highest reported for batch fermentations. Sugars from acid and enzyme-treated sweet sorghum bagasse were also fermented to butyrate by this organism with a molar yield of 0.81 (based on the amount of cellulose and hemicellulose). By combining the results from juice and bagasse, the calculated yield of butyric acid is approximately 90 kg per tonne of fresh sweet sorghum stalk. This study demonstrates that C. thermobutyricum can be an effective microbial biocatalyst for production of bio-based butyrate from renewable feedstocks at 50 ° C.

      • https://doi.org/10.1016/j.biortech.2015.09.062

  • Zeng, J., Tong, Z., Wang, L., Zhu, J.Y. and Ingram, L. (2014). Isolation and structural characterization of sugarcane bagasse lignin after dilute phosphoric acid plus steam explosion pretreatment and its effect on cellulose hydrolysis. Bioresource Technology 154:274-281. DOI: 10.1016/j.biortech.2013.12.072

      • Abstract:

      • The structure of lignin after dilute phosphoric acid plus steam explosion pretreatment process of sugarcane bagasse in a pilot scale and the effect of the lignin extracted by ethanol on subsequent cellulose hydrolysis were investigated. The lignin structural changes caused by pretreatment were identified using advanced nondestructive techniques such as gel permeation chromatography (GPC), quantitative 13C, and 2-D nuclear magnetic resonance (NMR). The structural analysis revealed that ethanol extractable lignin preserved basic lignin structure, but had relatively lower amount of Β-O-4 linkages, syringyl/guaiacyl units ratio (S/G), p-coumarate/ferulate ratio, and other ending structures. The results also indicated that approximately 8% of mass weight was extracted by pure ethanol. The bagasse after ethanol extraction had an approximate 22% higher glucose yield after enzyme hydrolysis compared to pretreated bagasse without extraction.

      • https://doi.org/10.1016/j.biortech.2013.12.0725

  • Zeng, J., Yoo, C.G., Wang, F., Pan, X., Vermerris, W. and Tong, Z. (2015). Biomimetic Fenton-catalyzed lignin depolymerization to high-value aromatics and dicarboxylic acids. ChemSusChem 8(5):861–871. DOI:10.1002/cssc.201403128

      • Abstract:

      • By mimicking natural lignin degradation systems, the Fenton catalyst (Fe3+, H2O2) can effectively facilitate lignin depolymerization in supercritical ethanol (7 MPa, 250 °C) to give organic oils that consist of mono- and oligomeric aromatics, phenols, dicarboxylic acids, and their derivatives in yields up to (66.0±8.5)% . The thermal properties, functional groups, and surface chemistry of lignin before and after Fenton treatment were examined by thermogravimetric analysis, pyrolysis–gas chromatography–mass spectrometry, 31P NMR spectroscopy, and X-ray photoelectron spectroscopy. The results suggest that the Fenton catalyst facilitates lignin depolymerization through cleavage of β-ether bonds between lignin residues. The formation of a lignin–iron chelating complex effectively depresses lignin recondensation; thus minimizing charcoal formation and enhancing the yield of liquid products.

      • https://doi.org/10.1002/cssc.201403128