Source: UNIVERSITY OF CALIFORNIA, DAVIS submitted to
THE SCIENCE AND ENGINEERING FOR A BIOBASED INDUSTRY AND ECONOMY
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
0216503
Grant No.
(N/A)
Project No.
CA-D-BAE-2005-RR
Proposal No.
(N/A)
Multistate No.
-OLD S1041
Program Code
(N/A)
Project Start Date
Oct 1, 2008
Project End Date
Sep 30, 2013
Grant Year
(N/A)
Project Director
Zhang, R.
Recipient Organization
UNIVERSITY OF CALIFORNIA, DAVIS
410 MRAK HALL
DAVIS,CA 95616-8671
Performing Department
Biological and Agricultural Engineering
Non Technical Summary
A need for biofuels and other biobased products has been recognized as a national priority. The overall goal of this project address research relating directly to biobased products and processing agricultural coproducts. The specific objectives of the projects include: 1. Reduce costs of harvesting, handling and transporting biomass to increase the competitiveness of biomass as a feedstock for biofuels, biomaterials and biochemicals 2. Improve biofuel production processes 3. Identify, develop and evaluate sustainable processes to convert biomass resources into biochemicals, biocatalysts and biomaterials 4. Identify and develop needed educational resources, develop distance based delivery methods, and develop a trained work force for the biobased economy Measurement of Progress and Results Outputs: A large portion of the efforts outlined in Objectives 1 through 4 are application oriented and will be useful to develop pilot projects, demonstrations and commercialization of biomass conversion to biobased products Other outputs include educational materials that could be used in traditional classroom settings or for distance education and web based distribution Publications in peer reviewed journals, trade journals and popular magazines. Development of intellectual property. Presentations to economic development groups, legislative groups, and to the general public. Development of management recommendations for producers of biobased products. Outcomes or projected Impacts: The committee has served and will continue to serve as a resource for: Bioresearch and Development Initiative (BRDI), Biomass, Research and Development Board working groups, SBIR panel Biofuels 8.8, USDA/DOE Biomass Initiative Project Review Teams, NRI 71.2 panel and reviewers for the Sun Grant Initiative. The multi-state membership will contribute to the implementation of the REE energy science strategic plan. Multi-state membership will contribute to identification of funding priorities and shaping policy of Federal agencies Research as a result of this project will create technology adopted by industry with at least two licensed technology per year. Research will enable reduced dependency on foreign-based fuels and chemicals.
Animal Health Component
(N/A)
Research Effort Categories
Basic
(N/A)
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
51124202020100%
Knowledge Area
511 - New and Improved Non-Food Products and Processes;

Subject Of Investigation
2420 - Noncrop plant research;

Field Of Science
2020 - Engineering;
Goals / Objectives
Reduce costs of harvesting, handling and transporting biomass to increase the competitiveness of biomass as a feedstock for biofuels, biomaterials and biochemicals Improve biofuel production processes Identify, develop and evaluate sustainable processes to convert biomass resources into biochemicals, biocatalysts and biomaterials Identify and develop needed educational resources, develop distance based delivery methods, and develop a trained work force for the biobased economy
Project Methods
Objective A. Reduce costs of harvesting, handling, and transporting biomass to increase competitiveness of biomass as a feedstock for biofuels, biomaterials and biochemicals. Task 1: Quantify and characterize biological feedstocks. (CA, FL, MT, OR, TN) Work in this area will seek to quantify and characterize biological feedstocks to be grown, produced, and converted to biofuels and value-added products. Task 2: Develop and evaluate harvest, process and handling methods. (CA, GA, IL, IN, KY, MS, MT, TN, TX, VA, WI ) Task 3: Model and analyze integrated feedstock supply and process systems. (GA, IL, IN, NE, TN, TX) Objective B. Improve biofuel production processes B.1. Biological conversion processes Task 1: Develop pretreatment methods for biological conversion processes (AR, CA, FL, IN, KS, MI, NC, NE, OK, OR, SD, UT, WI) Task 2: Develop conversion processes (CA, FL, KS, IL, NC, OK, OR, SC, SD, UT, WI) Task 3: Develop value-added products from hemicellulose and lignin (WI, NE) Hemicellulose and/or lignin fractions from cellulosic ethanol production and lignin-based chemicals and materials and/or applications of pentoses from hemicelluloses will be investigated. B.2. Thermochemical conversion processes Task 1: Develop pretreatment methods (GA, MN) Task 2: Develop conversion processes (CA, FL, GA, IN, KS, OK, MN, NE, TN, TX,) Task 3: Improve methods for characterization of intermediate products and process control (IN, MN, MS, SD, TN, TX) B.3. Biodiesel production processes Task 1: Characterize new feedstocks (AZ, GA, IN, MI, MT, ND, NE, SC, UT) Task 2: Develop an understanding of fuel quality and performance issues (IL, KS, MI, MT, ND, NE, SC, UT and VA ) Task 3: Develop and characterize innovative processes for biodiesel production (AZ, CA, FL, SD, MN, UT) Task 4: Utilize coproducts (AZ, CA, FL, KS, MI, MN, MT, ND, TN, SC, VA) Objective C. Identify, develop, and evaluate sustainable processes to convert biomass resources into biochemicals, biocatalysts, and biomaterials (non-fuel uses) Task 1: Discover and characterize biochemicals, biocatalysts, and biomaterials in biomass. (AR, FL, HI, IL, KS, KY, NC, ND, NE, OK, SC, SD, UT, VA, WA, WI) Task 2: Develop separation processes for biochemicals, biocatalysts, and biomaterials. (AR, GA, IN, KS, SD, UT, VA) Task 3: Develop applications for biochemicals and biocatalysts with biological activity. (AR, HI, MI, VA) Task 4: Develop enabling technologies for biochemical production. (HI, IN, KS, LA, MI, ND, SD) Task 5: Develop and evaluate integrated process systems for commercial feasibility. (SD, UT) Objective D. Identify and develop needed educational resources, develop distance based delivery methods, and develop a trained work force for the biobased economy Task 1: Serve as a knowledge resource base for biobased processing and products Task 2: Distribute new knowledge to train the work force and general public in biobased products and processing Task 3: Develop and disseminate educational materials in high-priority topic areas.

Progress 10/01/08 to 09/30/13

Outputs
Target Audience: The scientific community, policy makers, industry and other stakeholders interested in bioenergy production and biomass conversion. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? This project contributes to the development of knowledge and human resource skills. We are training many graduate students and post doctoral scholars on this project. Their training is unique in that it has a focus in both engineering and cell biology with application to sustainable agriculture and energy production. Graduate students are also trained in the mentoring of undergraduates offering multiple opportunities for human resource development. Partner organizations include government agencies and private companies, such as US Department of Agriculture, California Energy Commission, Chevron Technology Ventures, Mendota Bioenergy, CleanWorld, and others. How have the results been disseminated to communities of interest? The research results have been published in scientific articles, conference papers, presentations, lecture notes and research reports. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? A geospatial bioenergy systems model (GBSM) was developed and implemented to assess potential biofuel supply across the United States and more regionally from agricultural, forest, urban, and energy crop biomass. Spatial information including feedstock resources, national fuel demand, existing and potential refinery locations and a transportation network model is provided to a mixed integer-linear optimization model that determines the optimal locations, technology types and sizes of biorefineries and other facilities to satisfy a maximum profit or other objective function applied across the biofuel supply and demand chain from site of feedstock production to product final demand. The model is also capable of addressing carbon impacts, air quality, water, and other resource sustainability issues. In addition to regional applications in California and the western U.S., the model has also been applied to a regional study of poplar biomass production in the Pacific Northwest. The model was integrated with a poplar growth model to identify locally specific biomass yield and production information, and with a farm level crop adoption model for assessing crop shifting under different economic conditions. The results of the integrated modeling are output to a socioeconomic impact model to evaluate overall impacts on the local and regional economies associated with bioenergy development. Extensive research was carried out to accomplish the goals of evelopment of cost-effective methods for stabilization, storage and supply of seasonally produced biomass feedstock for sustainable biofuel production, and to improve the biofuel production yield from these feedstocks. Objectives included the development of (1) rapid stabilization methods for highly putrescent biomass feedstocks, and (2) delignification methods for improved bioconversion of lignocellulosic feedstocks. Studies were done to identify the minimum pectinase and cellulase levels needed for saccharification of tomato pomace (TP). What was particularly noteworthy was that the ethanol yield from ensiled TP under low-enzyme loading conditions was significantly higher than non-ensiled TP treated using the highest enzyme loading. The results strongly suggest that ensilage serves to pretreat TP during storage. Our work has implications for reducing the enzyme inputs needed for saccharification of TP, which may significantly improve process economics. Ensiling was also investigated for the long-term storage of Sugar Beet Pulp (SBP). Both SBP preservation and prevention of cellulose and hemicellulose loss were better when SBP was treated with Lactobacillus fermentum NRRL B-4524 compared to the non-inoculated control. Additionally, the ensiling process was found to significantly improve the enzymatic digestibility of SBP by as much as 35%. The results suggest that ensiling may be a promising technology for SBP stabilization and pretreatment for bioconversion to products. An integrated system for production of ethanol and biogas from a variety of biomass materials, including alkaline pretreated rice and wheat straw, sugar beet, grape pomace, and other biomass feedstock was developed. New pretreatment processes using sodium hydroxide (NaOH) and potassium hydroxide (KOH) with effective chemical recycling and recovery were developed. New methods for chemical and lignin separation and recovery were also developed. The new biomass pretreatment technologies can be applied to bioconversion systems, including anaerobic digestion and ethanol fermentation. Research was also conducted to produce cellulase enzymes using T. reesei on various lignocellulosic substrates, including rice straw, grape pomace and pure celloluse. New technologies were developed to convert sugar beets into biofuels (ethanol and biogas) and fertilizer products. New processes were developed and optimized to reduce the enzyme and energy requirement for liquefaction and hydrolysis of sugar beets. Simultaneous and separate hydrolysis and fermentation processes were developed for ethanol production. A continuous mesophilic anaerobic digester was tested with the stillage from ethanol processes. The pilot test of an integrated ethanol and biogas production system was highly successful to provide the design and operation data for scale up. Process and system engineering design and system model were developed for a demonstration plant that has a capacity of processing 100 tons per day. A high-rate anaerobic digestion of food and agricultural waste was developed and have been applied to the high solids organic waste in both pilot and commercial scale. Integrated biological treatment and advanced separation technologies were developed to fractionate the digester effluent and concentrate the solids and nutrients into marketable products. A novel biochemical route for producing fuels and chemicals from cellulosic biomass was developed. Instead of making sugars as the reactive intermediates for subsequent fermentation, the new route generates sugar aldonates and their salts as the reaction intermediates directly from cellulose using an genetically engineered fungus. The sugar aldonates are then fermented to fuels and chemicals. We demonstrated that sugar aldonates can be produced from cellulose after six beta-glucosidase genes were deleted and exogenous cellobiose dehydrogenase was added; and sugar aldonates were able to be converted to ethanol at a rate which is even faster than that of conventional substrate glucose. We also conducted some fundamental research on the beta-glucosidase (BGL) production, and cellulase regulation in the model microorganism N. crassa. We constructed a library of sextuple bgl knockout strains (with only one bgl gene remaining active) and one septuple bgl knockout strain (with no active bgl gene remaining). With these mutants, we studied the direct cellobiose production from cellulose by the sextuple bgl deletion strains without exogenous cellulase addition, and the cellular location of the each BGL and the contribution of each BGL to the overall BGL activity. We also tested a new a bacterial recombination system employing β-recombinase acting on six recognition sequences (β-rec/six) which allowed repetitive site-specific gene deletion and marker recycling in N. crassa. We demonstrated the functionality of the beta-recombinase based gene knockout system in N. crassa for the first time. We further improve the system by adding a negative selection marker in the knock-in cassette to enable one-step homokaryon purification and using a more regulative native promoter in N. crassa to improve the cassette eviction efficiency.

Publications

  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Li, Y., R. Zhang, G. Liu, C. Chen, Y. He and X. Liu. 2013. Comparison of methane production potential, biodegradability, and kinetics of different organic substrates . Bioresource Technology, 2013, 149: 565-569.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Li, Y., R. Zhang, C. Chen, G. Liu, Y. He and X. Liu. 2013. Biogas production from co- digestion of corn stover and chicken manure under anaerobic wet, hemi-solid, and solid state conditions [J]. Bioresource Technology. 149: 406-412.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Li, Y., L. Feng, R. Zhang, Y. He, X. Liu, X. Xiao, X. Ma, C. Chen and G. Liu. 2013. Influence of inoculum source and pre-incubation on bio-methane potential of chicken manure and corn stover. Applied Biochemistry and Biotechnology. 171(1): 117-127.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Lu, Feng, Y. Li, C. Chen, X. Liu, X. Xiao, X. Ma, R. Zhang, Y. He and G. Liu. 2013. Biochemical methane potential (BMP) of vinegar residue and the influence of feed to inoculum ratios on biogas production. BioResources, 2013, 8(2): 2487-2498.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Li, Y., R. Zhang, X. Liu, C. Chen, X. Xiao, F. Lu, Y. He and G. Liu. 2013. Evaluating methane production from anaerobic mono- and co-digestion of kitchen waste, corn stover, and chicken manure [J]. Energy & Fuels, 27(4): 2085-2091.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Simmons CW, Claypool JT, Marshall MN, Jabusch LK, Reddy AP, Simmons BA, Singer SW, Stapleton JJ, VanderGheynst JS. 2014. Characterization of bacterial communities in solarized soil amended with lignocellulosic organic matter. Applied Soil Ecology 73:97-104.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Reddy AP, Simmons CW, D'haeseleer P, Khudyakov J, Burd H, Hadi MZ, Simmons BA, Singer SW, Thelen MP, VanderGheynst JS. 2013. Discovery of microorganisms and enzymes involved in high-solids decomposition of rice straw using metagenomic analyses. PLoS One In press.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Marshall MN, Rumsey TR, Stapleton JJ, VanderGheynst JS. 2013. A Predictive Model for Soil Temperature During Solarization and Its Validation at Two California Field Sites. Transactions of the ASABE 56(1):117-133.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Williams, K., Y. Zheng, J. McGarvey, Z. Fan and R. Zhang. Ethanol and volatile fatty acid production from lignocellulose by clostridium cellulolyticum. ISRN Biotechnology. vol. 2013, Article ID 137835, 7 pages, 2013. doi:10.5402/2013/137835.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Zheng Y, Lee C, Yu C, Cheng Y-S, Zhang R, Jenkins BM, VanderGheynst JS. 2013. Dilute acid pretreatment and fermentation of sugar beet pulp to ethanol. Applied Energy 105(0):1-7.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Wu, W., Kasuga, T., Xiong, X., Ma, D., and Fan, Z. (2013) Location and contribution of individual beta-glucosidases from Neurospora crassa to total beta-glucosidase activity Archives of Microbiology, 195:823-9
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Hildebrand, A., Schlacta, T., Warmack, R., Kasuga, T., and Fan, Z. (2013) Engineering Escherichia coli for improved ethanol yield from gluconate, Journal of Biotechnology, 168: 101-106.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Hildebrand, A., T. Schlacta, T., R. Warmack, T. Kasuga, and Z. Fan: Engineering Escherichia coli for improved ethanol yield from gluconate, Presented at the 35th Symposium on Biotechnology for Fuels and Chemicals, Portland, OR.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Szewczyk, E., T. Kasuga, and Z. Fan. A new variant of self-excising beta� recombinase/six cassette for repetitive gene deletion and homokaryon purification in Neurospora crassa, presented at the 2013 AICHE annual meeting
  • Type: Journal Articles Status: Accepted Year Published: 2014 Citation: Murphy, C. and N. Parker. 2014. Effects of air pollution control costs on U.S. biofuel system development. Environmental Science & Technology (In press).
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Xiao, X., R. Zhang, Y. He, Y. Li, L. Feng, C. Chen and G. Liu. 2013. Influence of particle size and alkaline pretreatment on the anareobic digestion of corn stover. BioResources.8(4): 5850-5860.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Simmons CW, Guo H, Claypool JT, Marshall MN, Perano KM, Stapleton JJ, VanderGheynst JS. 2013. Managing compost stability and amendment to soil to enhance soil heating during soil solarization. Waste Management 33(5):1090-1096.


Progress 01/01/12 to 12/31/12

Outputs
OUTPUTS: Development of a large scale geospatial bioenergy system model was continued for evaluating technical and policy outcomes in relation to the national renewable fuel standard and the potential application of a state and national low carbon fuel standard. The model is also being applied in a regional study of advanced biofuels production from hardwood poplars in the Pacific Northwest. In addition to quantifying sustainability metrics and identifying optimal biorefinery site locations, the model has been used to develop information on the anticipated costs to take the emerging biofuel industry from first to nth plant along different learning scenarios, providing estimates for potential incentives or other alternatives necessary to finance the industry to the point of independent commercial profitability. An innovative conversion technology for cellulose biomass was investigated with cellulose first converted to cellobionate by a genetically modified fungus and cellobionate is then converted to fuels and chemicals. Neurospora crassa was used as the model microorganism to construct the strain for conversion of the cellulose to cellobionate. A library of sextuple bgl deletion N. crassa strains and the septuple bgl deletion N. crassa strain were constructed by genetic crossing, using a new targeted repetitive gene deletion method with marker recycle was used. A new self-excising resistance marker system featuring beta-recombinase/six cassette for serial gene replacement in N. crassa was developed and the functionality of the system in N. crassa was validated for the first time. A new integrated system was developed and proven in the laboratory and then scaled up to the pilot scale for converting whole sugar beets into ethanol and biogas fuels. A pilot scale ethanol fermentation system capable of fermenting up to 10 tons per batch was developed and successfully tested. The ethanol yield from sugar beets was determined to be about 0.4 gram per gram of total solids in the beets, which was 90% of the ethanol yield obtained in the laboratory research. The stillage from the ethanol fermentation was processed through anaerobic digester with a biogas yield over 0.6 L/gVS. Sugar beet pulp (SBP) is also investigated as biomass feedstock for fermentable sugar and ethanol production. The enzymatic hydrolysis of SBP was studied to examine the effects of solid loading (2-10%, dry basis), enzyme preparation and enzyme recycle on the production of fermentable sugars. The enzyme partitioning to the solid and liquid phases during SBP enzymatic hydrolysis and loss during recycling were investigated using SDS-PAGE and Zymogram analysis. Without considering product inhibition, the cellulase added initially to the SBP hydrolysis lost only 6% filter paper activity and negligible carboxymethyl cellulose activity upon multiple cycles of SBP hydrolysis. The results have been disseminated to government agencies and industry sponsors, and have also been presented to the scientific communities with conference papers and scientific publications. PARTICIPANTS: This project contributes to the development of knowledge and human resource skills. We are training many graduate students and post doctoral scholars on this project. Their training is unique in that it has a focus in both engineering and cell biology with application to sustainable agriculture and energy production. Graduate students are also trained in the mentoring of undergraduates offering multiple opportunities for human resource development. Partner organizations include government agencies and private companies, such as US Department of Agriculture, California Energy Commission, Chevron Technology Ventures, Mendota Bioenergy, Clean World Partners, and others. TARGET AUDIENCES: The research results are useful to researchers, technology developers and users, government agencies, companies working in the areas related to bioenergy, biofuel, and waste treatment. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Results from the modeling have been used in evaluating policy alternatives around low carbon fuels in California and nationally and in quantifying uncertainties in national biomass resources and potential biofuel supplies. New methods and improved fungal and bacterial strains are highly significant. A new self-excising resistance marker system for serial gene replacement employing a beta-recombinase acting on six sequences was validated in N. crassa for the first time. The new method is more superior than the widely used Cre-loxP systems in the following two aspects: 1) only one transformation per gene deletion is needed using the beta-Rec/six sequence system, while the Cre-loxP system needs three. 2) Using the beta-Rec/six sequence system eliminates the risk of unwanted chromosome rearrangement, as occurs in the Cre/loxP system. This method is very useful for other researchers who are interested in constructing multiple gene deletions in N. crassa. An integrated ethanol and biogas production system using whole sugar beets will enable the establishment of biorefinery operations with higher biofuel production efficiency and lower costs. Improvement of ethanol and biogas yields by applying combined approaches of enzymatic and microbial conversion will improve the overall efficiencies of the biofuel production systems. High rate anaerobic digestion technologies will allow more applications of biogas energy technologies for solid organic waste streams. In the hydrolysis of sugar beet pulp (SBP), it was found that enzyme dosage can be reduced by 50% while maintaining similar, and in some cases higher fermentable sugar yield. The removal of hydrolysis products will further improve enzymatic hydrolysis of SBP for biofuel production.

Publications

  • Aramrueang, N. S. Zicari, J. Li, S. Kaffka, J. Tischer and R. Zhang: 2012. Characterization and compositional analysis of herbaceous crops for biofuel production, Presentation at ASABE Annual International Meeting. July 29-August 1. Dallas, Texas
  • Aramrueang, N. S. Zicari, K. Williams and R. Zhang: 2012. Enzymatic hydrolysis of sugar beet leaves for biofuel production. Presentation at ASABE Annual International Meeting. July 29-August 1. Dallas, Texas.
  • Kim. H. S. Moon, A. Abug, S. Choi, R. Zhang, Y. Oh. 2012. Effect of fermentation conditions on biohydrogen production from lipid-rich food material. International Journal of Hydrogen Energy. 37:15062-15069.
  • Parker, N. 2012. Spatially explicit projection of biofuel supply for meeting renewable fuel standard. Transportation Research Record: Journal of the Transportation Research Board No. 2287:72-89.
  • Wu, W., A. Hildebrand, T. Kasuga, X. Xiong, Z. Fan. 2013 Direct cellobiose production from cellulose using sextuple beta-glucosidase gene deletion Neurospora crassa mutants. in press Enzyme and Microbial Technology
  • Szewczyk, E., T. Kasuga, Z. Fan. 2013 Efficient sequential repetitive gene deletions in Neurospora crassa, employing self-excising beta-recombinase/six cassette. In press Journal of Microbiological Methods
  • Szewczyk, E. W. Wu, T. Kasuga, Z. Fan. 2012. An integrated bioprocessing and separation process for biofuels production from cellulosic biomass presented at the poster session at the Spring 2012 ACS National Meeting , San Diego, CA
  • Szewczyk, E. W. Wu, T. Kasuga, Z. Fan. 2012. An integrated bioprocessing and separation process for biofuels production from cellulosic biomass presented at the poster session at the 34st Symposium on Biotechnology for Fuels and Chemicals, New Orleans, LA
  • Szewczyk, E. W. Wu, X. Xiong, T. Kasuga, Z. Fan 2012. Genetic engineering of Neurospora crassa for direct cellobiose production from cellulose presented at the poster session at the 34st Symposium on Biotechnology for Fuels and Chemicals, New Orleans, LA
  • Zhang, R.H., S. Zicari, C. Chang, N. Aramrueang: 2012. Ethanol Production from Sugar Beets, Research report submitted to Califronia Energy Commission. June 4, 2012.
  • Zheng Y, Cheng Y-S, Yu C, Zhang R, Jenkins B, VanderGheynst J. 2012. Improving the efficiency of enzyme utilization for sugar beet pulp hydrolysis. Bioprocess and Biosystems Engineering 35(9):1531-1539.
  • Zheng Y, Lee C, Yu C, Cheng Y-S, Simmons CW, Zhang R, Jenkins BM, VanderGheynst JS. 2012. Ensilage and Bioconversion of Grape Pomace into Fuel Ethanol. Journal of Agricultural and Food Chemistry 60(44):11128-11134.
  • Zheng Y, Lee C, Yu C, Cheng Y-S, Zhang R, Jenkins BM, VanderGheynst JS. 2013. Dilute acid pretreatment and fermentation of sugar beet pulp to ethanol. Applied Energy 105(0):1-7.
  • Zicari, S., R. Zhang, N. Aramurueang, A. Kendall, S. Kafflka, J. Tischer and J. Manternach: 2012. Sugar beets for advanced bioethanol and biogas production in California, Poster.


Progress 01/01/11 to 12/31/11

Outputs
OUTPUTS: Significant research was carried out to develop new methods for biomass storage and conversion and novel microorganisms and pathways to produce ethanol and other bilfuels. Studies were done to identify the minimum pectinase and cellulase levels needed for saccharification of tomato pomace (TP). Outputs from this study included enzyme levels required for simultaneous saccharification and co-fermentation of TP to ethanol. Additional studies were completed to examine SSCF of ensiled and non-ensiled TP. The conversion rate of cellulose for ensiled TP at low enzyme loadings was equivalent to or higher than conversion using high enzyme loadings. Furthermore, higher ethanol yields were obtained for ensiled TP compared to non-ensiled TP. An output from this work is storage conditions for TP that improve conversion to biofuel. An alternative two step method for conversion of cellulose biomass was investigated. Cellulose will be converted to cellobionate by a genetically modified fungus without exiguous cellulase addition in an aerobic fermentation step. Cellobionate is then converted to fuels and chemicals in a second anaerobic step. We use the Neurospora crassa as the model microorganism to construct the strain for conversion of the cellulose to cellobionate. During the process of construct the mutant strains with multiple copies of beta-glucosidase knocked out, we also studied the relationship between beta-glucosidase and cellulase production, the locality of individual beta-glucosidase gene products, and characterized multiple mutants with multiple copies of the be-ta-glucosidase knockout. New research was initiated to develop an integrated system to convert whole sugar beets into ethanol and biogas fuels. Sugar beets of different varieties were characterized for sugar and structural component. Laboratory experiments were partially performed for converting sugar and cellulosic compounds into ethanol using yeast and enzymes. Anaerobic digestion of solid organic waste continued for bioenergy production. Community based anaerobic digestion system was developed to conversion multiple streams of organic wastes into biogas energy. A new high rate anaerobic digestion system was developed for fast conversion of food waste into biogas fuel. Anaerobic digestion research was carried out in both laboratory and pilot scale. The results have been disseminated to industry sponsors, including Novozymes (Davis, CA), Chevron Technologies Ventures, Mendota Bioenergy, Clean World Partners and have also presented to the scientific communities with conference papers and scientific publications. PARTICIPANTS: This project contributes to the development of knowledge and human resource skills. We are training many graduate students and post doctoral scholars on this project. Their training is unique in that it has a focus in both engineering and cell biology with application to sustainable agriculture and energy production. Graduate students are also trained in the mentoring of undergraduates offering multiple opportunities for human resource development. Partner organizations include government agencies and private companies, such as US Department of Agriculture, California Energy Commission, Chevron Technology Ventures, Mendota Bioenergy, Clean World Partners, and others. TARGET AUDIENCES: The research results are useful to researchers, technology developers and users, government agencies, companies working in the areas related to bioenergy, biofuel, and waste treatment. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
The results from ensilage and bioconversion of tomato pomace to ethanol strongly suggest that ensilage serves to pretreat TP during storage. Our work has implications for reducing the enzyme inputs needed for saccharification of TP, which may significantly improve process economics. Disruption of beta-glucosidase gene on cellulase production was studied with the results showing that the disruption of individual beta-glucosidase genes did not have a negative effect on cellulase production. On the contrary, some single beta-glucosidase gene knockout strains produced more cellulase than the wild type strain. The expression patterns of the individual beta-glucosidase genes were determined. Such new knowledge is of interest to the cellulase community. An integrated ethanol and biogas production system using whole sugar beets will enable the establishment of biorefinery operations with higher biofuel production efficiency and lower costs. Improvement of ethanol and biogas yields by applying combined approaches of enzymatic and microbial conversion will improve the overall efficiencies of the biofuel production systems. High rate anaerobic digestion technologies will allow more applications of biogas energy technologies for solid organic waste streams.

Publications

  • Clark, I. C., R. Zhang, Z. Pan, B. Brown, J. Ambuel and M. Delwiche. 2011. Development of a low flow meter for measuring gas production from bioreactors. Transactions of the ASABE; vol. 54(5): 1959-1964.
  • Fan, Z., Wu, W., Zhang R., Kasuga, T., Szewczyk E., and Xiong X. 2012 A novel biochemical route for fuels and chemicals production from cellulosic biomass In press, PLOS One
  • El-Mashad, H.M., R. Zhang, and J.P. Green, 2011. Anaerobic Biodegradability of Selected Biodegradable Plastics and Biobased Products . Journal of Environmental Science and Engineering.
  • Liu, G., R. Zhang, H. El-Mashad, R. Dong and X. Liu. 2011. Biogas production from green and food wastes using anaerobic phased solids digester system. Applied Biochemistry and Biotechnology. DOI 10.1007/s12010-011-9322-z
  • Romano, R.R. and R.H. Zhang. 2011. Anaerobic Digestion of Onion Waste Using a Mesophilic Anaerobic Phased Solids Digester. Biomass and Bioenergy.
  • Zheng, Y., Z. Pan and R. Zhang. 2010. Fermentable Sugar Production from Lignocellulosic Biomass. In: Advances in Energy Research. Volume 2 (.Editor: Morena J. Acosta). Nova Science Publishers, Inc.
  • Wu, W., kasuga, T., and Fan, Z. 2011. Analysis of cellulase expression in beta-glucosidase knockout strains of Neurospora crassa 33rd Symposium on Biotechnology for Fuels and Chemicals. Seattle, Washington
  • Zheng Y, Yu C, Cheng Y-S, Lee C, Simmons C, Dooley T, Zhang R, Jenkins B, VanderGheynst JS. 2012. Integrating Sugar Beet Pulp Storage, Hydrolysis and Fermentation for Fuel Ethanol Production. Applied Energy. In press.


Progress 01/01/10 to 12/31/10

Outputs
OUTPUTS: Research on large-scale storage of four food processing wastes -- fresh grape pomace (FrGP), fermented grape pomace (FeGP), sugar beet pulp (SBP), and tomato pomace (TP) - was completed. The storage volume of 1200-L was used. Outputs included data on best management practices for storage of wet feedstocks. Raw and ensiled grape pomace underwent simultaneous saccharification and fermentation (SSF) using Escherichia coli KO11. Outputs from the research included hydrolysis and fermentation approaches for conversion of grape pomace to ethanol. Finally, studies were completed to examine the pretreatment of SBP by acid pretreatment. Results demonstrated that mild acid pretreatment conditions improved the conversion of SBP to ethanol. The results have been disseminated to communities of interest including Novozymes (Davis, CA) and Chevron Technologies Ventures. Alkaline pretreatment of straw, grape pomace and woody biomass for increasing the biodegradability was investigated. New pretreatment processes using sodium hydroxide (NaOH) was developed. New methods for chemical and lignin separation and recovery were also developed. The research outputs include new process design and optimized conditions for achieving effective biomass pretreatment. The new biomass pretreatment technologies can be applied to bioconversion systems, including anaerobic digestion and ethanol fermentation. Research was also conducted to produce cellulase enzymes using T. reesei on various lignocellulosic substrates, including rice straw, grape pomace and pure celloluse. Anaerobic digestion of various organic residues, including poultry manure, vegetable waste, sugar beet pulp and leaves were investigated for the purpose of developing effective biogas production system from these biomass materials. Both batch and continuous digesters were tested and recommendations were developed for the optimum conditions for designing and operating anaerobic digesters. Research results have been presented to industry sponsor and collaborators, at scientific meetings, in research reports and articles. PARTICIPANTS: This project contributes to the development of knowledge and human resource skills. We are training many graduate students and post doctoral scholars. Their training is unique in that it has a focus in several disciplines, including engineering, microbiology, and cell biology with application to sustainable agriculture and energy production. Graduate students are also trained in the mentoring of undergraduates offering multiple opportunities for human resource development. Partner organizations include government agencies and private companies, such as California Energy Commission, Chevron Technology Ventures, Biostar Systems, Clean World Partners, and others. TARGET AUDIENCES: The research results are useful to researchers, technology developers and users, government agencies, companies working in the areas related to bioenergy, biofuel, and waste treatment. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Ensilage at the 1200-L scale varied by material, suggesting each feedstock requires unique storage strategies. Ethanol, whether present from yeast fermentation or as process residue, was effective in protecting FrGP and FeGP from spoiling. Ethanol could be recovered from FrGP and FeGP either before or after storage. Airtight storage of SBP was required to preserve anaerobic conditions and prevent aerobic mold contamination. Dilute acid pretreatment was selected for pretreatment of SBP. Three critical parameters including temperature (120-160˚C), acid concentration (0 -1 wt%) and solid loading (2 -6 wt%) were tested. The pretreatment time was set at 5 min. Dilute acid pretreatment significantly improved the digestibility of SBP to achieve high sugar and ethanol yield upon hydrolysis and fermentation. Alkaline pretreatment using NaOH at ambient temperature (20˚C) was shown to be very effective in increasing the biogas yield via anaerobic digestion and sugar yield via enzyme hydrolysis from rice straw and grape pomace. The chemical requirement varies with different materials due to different lignin contents. For grape pomace and rice straw, recommended chemical loading was 0.02 and 0.10 gNaOH per g of dry biomass (10%) for 24 hour treatment with corresponding methane yields of 362 and 317 ml/gVS. For woody biomass from orchards, pretreatment at a higher temperature was needed. After 1 hour pretreatment with 10% NaOH loading and at 120˚C, methane yield of woody biomass was 213 ml/gVS. Separation of lignin in black liquor after the biomass pretreatment would enable recovery of lignin for industrial or energy uses and also improve the quality of water for later reuse in the pretreatment process. Two step alkaline pretreatment was found to be better than single step process. More research is underway to optimize the conditions for the two step process. Cellulase enzyme production research results indicated that alkaline pretreated rice straw was a better substrate for cellulase production as compared to raw rice straw and pure cellulose. The activity of cellulase produced from rice straw was higher than the commercial cellulase tested. More research is underway to optimize the conditions for enzyme production. The results of anaerobic digestion research led to improved designs of anaerobic digester system for treatment of vegetable waste and an invention of three-stage high rate anaerobic digestion system (HRD).

Publications

  • Yu, C. B. Jenkins, P. Thy, L. Wang, A. Bhat, S. Anderson, Y. Zheng, J. VanderGheynst, R. Zhang, T. Jeoh. 2010. Leaching pretreatments to improve biomass feedstock quality and resource recovery. Abstract and Poster Presented at 3ndst Symposium on Biotechnology for Fuels and Chemicals. April 19-22, 2010. Clearwater Beach, FL.
  • Zheng Y, Yu C, Cheng Y-S, Zhang R, Jenkins B, VanderGheynst JS. 2011. Effects of ensilage on the storage and the improvement of enzymatic degradability of sugar beet pulp. Bioresource Technology. 102:1489-1495.
  • Zhu, B., R. H. Zhang, P. Gikas, J. Rapport, B.M. Jenkins and X. Li. 2010. Biogas production from municipal solid waste using integrated rotary drum and anaerobic- phased solid digester system. Bioresource Technology, 101 (2010):6374-6380.
  • Zheng, Y. and Zhang, R.H., 2010. Lignocellulosic biomass pretreatment for bioethanol production. I In Erbaum, J. B. Bioethanol: Production, Benefits and Economics. pp. 1-48; Nova Science Publishers, Inc. Hauppauge, New York. Zheng, Y. C. Lee, C. Yu, Y.S. Cheng, R. Zhang, J. VanderGheynst, and B. Jenkins. 2010. Pretreatment and fermentation of sugar beet pulp into fuel ethanol. Abstract and Poster Presented at 3ndst Symposium on Biotechnology for Fuels and Chemicals. April 19-22, 2010. Clearwater Beach, FL.
  • Zhang, R., P. Gikas, B. Zhu, J. Lord, C. Choate, J. Rapport, H. El-Mashed, B. Jenkins. 2010 Integration of Rotary Drum Reactor and Anaerobic Digestion Technologies for Treatment of Municipal Solid Waste. California Department of Resources Recycling and Recovery. Publication # DRRR-2010-004. http://www.calrecycle.ca.gov/Publications/Organics/2010004.pdf
  • Chen, X., R. Zhang, S. Shoemaker, and E. Hughes. 2010. Alkaline pretreatment of grape pomace for enhancing its biogas energy yield. Abstract and Poster Presented at 3ndst Symposium on Biotechnology for Fuels and Chemicals. April 19-22, 2010. Clearwater Beach, FL.
  • Qu,W. Z. Pan, R. Zhang, H. Ma, X. Chen, B. Zhu, Z. Wang and G. G. Atungulu.2010. Integrated extraction and anaerobic digestion process for recovery of nutraceuticals and biogas from pomegranate marc. Transactions of the ASABE, 52(6):1997-2006. El-Mashad, H.M. and R.H. Zhang. 2010. Biogas production from co-digestion of dairy manrue and food Waste. Bioresource Technology, 101:4021-4028. Everhart, D. and R. Zhang. 2010. Alchemy in the desert: The UC-Davis/Auc Biodigester project. Energetica India. July/August Issue. 74-75.
  • Gikas, P., H. El-Mashad, R. Zhang. 2010. Biogas production from autoclaved and rotary drum reactor-treated municipal solid waste. Paper Presented at 3rd International Conference on Engineering for Waste and Biomass Valorisation, May 17-19, 2010 Beijing, China.
  • Rezaei F, VanderGheynst JS. 2010. Critical moisture content for microbial growth in dried food-processing residues. Journal of the Science of Food and Agriculture 90(12):2000-2005.
  • Williams, K., R. Zhang, and T. Jeoh. 2010. Ehtanol production from agricultural residues using integrated enzyme production and microbial fermentation system. Abstract and Poster Presented at 3ndst Symposium on Biotechnology for Fuels and Chemicals. April 19-22, 2010. Clearwater Beach, FL.
  • Yu C, Zheng Y, Cheng Y-S, Jenkins BM, Zhang R, VanderGheynst JS. 2010. Solid liquid extraction of alkali metals and organic compounds by leaching of food industry residues Bioresource Technology 101:4331 4336.


Progress 01/01/09 to 12/31/09

Outputs
OUTPUTS: A resource assessment and biorefinery siting optimization model was developed and implemented to assess potential biofuel supply across the United States from agricultural, forest, urban, and energy crop biomass for the next decade. Spatial information including feedstock resources, national fuel demand, existing and potential refinery locations and a transportation network model is provided to a mixed integer-linear optimization model that determines the optimal locations, technology types and sizes of biorefineries to satisfy a maximum profit objective function applied across the biofuel supply and demand chain from site of feedstock production to the product fuel terminal. The resource basis includes preliminary considerations of crop and residue sustainability. Research was also carried out to develop cost-effective methods for stabilization, storage and supply of seasonally produced biomass feedstock for sustainable biofuel production, and to improve the biofuel production yield from these feedstocks. Significant developments were made in these areas, (1) rapid stabilization methods for highly putrescent biomass feedstocks, and (2) delignification methods for improved bioconversion of lignocellulosic feedstocks. We adapted a 150-L rotating drum reactor for alkaline pretreatment. Several studies have been completed to examine the dynamics in rice straw pretreatment at this scale. In addition engineering design and operation parameters were determined for a high-pressure reactor for dilute acid pretreatment of rice straw and sugar beet pulp; studies were initiated to determine optimal conditions for dilute sulfuric acid pretreatment of sugar beet pulp. Alkaline pretreatment of rice straw and grape pomace at low temperature range (25-50degree C) was found to be a co-effective method for increasing the biodegradability of these biomass for biofuel production. During the pretreatment, part of lignin present in the biomass was solublized in black liquor and removed from the solids, making the solids more digestable with higher biogas yield via anaerobic digestion. An integrated system for production of ethanol and biogas from alkaline pretreated rice straw, sugar beet and other biomass feedstock was developed. Fungal enzyme production using rice straw and rice bran as substrates were also investigated for the purpose of developing an integrated enzyme production and fermentation system for production of ethanol and other valuable products. Production of lactic acid from paper sludge was studied using thermophilic Bacillus coagulan strains 36D1 and P4-102B. Effective separation of organic residuals from municipal solid waste and anaerobic digestion of organic residuals, including various food wastes, were researched for biogas energy production. The results have been disseminated to communities of interest including such companies as Novozymes (Davis, CA), Chevron Technologies Ventures, Mendota Advanced Beet Bioenergy Cooperative, Clean World Partners, LLC. In addition the results have been disseminated in presentations to industry, presentations at scientific meetings and publications. PARTICIPANTS: This project contributes to the development of knowledge and human resource skills. We are training many graduate students, post doctoral scholars and undergraduate students on this project. Their training is unique in that it has a focus in both engineering and biology with application to sustainable agriculture and energy production. Graduate students are also trained in the mentoring of undergraduates offering multiple opportunities for human resource development. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Biomass resources survey and analysis shows that the RFS2 goal of 24 billion ethanol-equivalent gallons for the year 2017 can be met at a cost of $2.50 per gallon gasoline equivalent (gge) at the fuel terminal. Taxes and local distribution bring the total to approximately $3.10/gge. The 2022 RFS2 goal of 36 billion ethanol-equivalent gallons can be achieved at approximately $3.00/gge at the fuel terminal. At $2.50/gge, model predictions include the construction of 228 cellulosic ethanol facilities and 18 Fischer-Tropsch biorefineries. Sensitivity analysis gives a range of $2.25 to $3.75/gge for meeting the 2017 RFS2 goal and $2.90 to $5.50/gge for meeting the 2022 goal. An exploratory survey of the mercury content of some common biomass feedstocks shows that the concentrations are well below EPA toxicity levels with representative feedstock concentrations of 20 ppb for rice straw, 28 ppb for wheat straw, and 32 ppb for whole-tree wood chips. Water leaching has no detectable effects on mercury mobility. Pretreatment of rice straw for enhanced biofuel production shows the following results. The anaerobic digestion and enzymatic hydrolysis results of rice straw treated with three alkali chemicals, including lime, sodium hydroxide, and potassium hydroxide that NaOH was the most effective chemical and temperature ranging from 20 degree C to 50 degree C had no significant effect on lignin removal. The results of using a rotating drum at 70 degree C to pretreat rice straw for 6 hours, with 6% sodium hydroxide (per g dry biomass), target moisture content of 80% (wet basis) and heating for 6 hours showed that both glucose conversion ratio and glucose yield (47.6% and 194.7 mg/g raw biomass) from enzymatic saccharification were greater, compared to preliminary studies done at 55oC in a static system. We found that ensiling significantly increased the ash content of sugar beet pulp and grape pomace. The results of combustion experiments indicated that tomato pomace, grape pomace, and sugar beet pulp may require pretreatment prior to any direct use in high temperature conversion systems. Production of lactic acid from paper sludge was studied using thermophilic Bacillus coagulan strains 36D1 and P4-102B. More than 80% of lactic acid yield and more than 87% of cellulose conversion were achieved using both strains without any pH control due to the buffering effect of CaCO3 in paper sludge. Results of semi-continuous simultaneous saccharification and co-fermentation (SSCF) using medium containing 100 g/L cellulose equivalent paper sludge without pH control showed that when rich medium was utilized, the average lactic acid concentrations in steady state for strains 36D1 and P4-102B were 92 g/L and 91.7 g/L, respectively, and lactic acid yields were 77% and 78%. Various food waste and food processing wastes were characterized for its potential for biogas energy production. A pilot scale, theromophilic anaerobic phased digester system was operated on a food processing waste for over 10 months and achieved 90% solids conversion and 24 therms of biogas energy yield per ton of food waste with moisture content of 80%.

Publications

  • Zheng,Yi., M. Yates, Y. Cheng, C. W. Yu, T. Dooley, R. Zhang, J. VanderGheynst, and B. Jenkins. 2010. Effect of Moisture Content on the Ensilage of Sugar Beet Pulp and Tomato Pomace. Submitted to Biomass and Bioenergy.
  • Cheng, Y., Y. Zheng, C. W. Yu, T. M. Dooley, B. M. Jenkins and J. S. VanderGheynst. 2010. Evaluation of high solids alkaline pretreatment of rice straw. Submitted to Chemical Technology and Biotechnology.
  • Wei, C. W Yu, B. J., J. VanderGheynst, R. Zhang, Y. Zheng, and Y. Cheng. 2010. Leaching of Food Industry Residues to Improve Feedstock Quality and Resource Recovery Bioresource Technology. (In press.)
  • Parker, N., Q. Hart, P. Tittmann, C. Murphy, M. Lay, R. Nelson, K. Skog, E. Gray, A. Schmidt and B. Jenkins. 2010. National Biorefinery Siting Model: Spatial analysis and supply curve development. Final report, Western Governors Association, Contract 20113-03, Denver, CO.
  • Hart, Q., N. Parker, P. Tittmann, M. Lay, R. Nelson, K. Skog, E. Gray, A. Schroeder and B. M. Jenkins. 2010. Strategic development of bioenergy in the United States: spatial analysis and supply curve development. Final report, USDA Project Number 58-3000-7-0158 (fund 30364), USDA, Washington, D.C.
  • Parker, N., P. Tittmann, Q. Hart, R. Nelson, K. Skog, A. Schmidt, E. Gray and B.M. Jenkins. 2010. Development of a biorefinery optimized biofuel supply curve for the Western United States. Biomass & Bioenergy (in press).
  • Jenkins, B.M., R.B. Williams, N. Parker, P. Tittmann, Q. Hart, M.C. Gildart, S. Kaffka, B.R. Hartsough, P. Dempster. 2009. Sustainable use of California biomass resources can help meet state and national bioenergy targets. California Agriculture 63(4):168-177.
  • Jenkins, B.M., C. Somerville, J.J. Stapleton. 2009. Biofuels: growing toward sustainability (editorial). California Agriculture 63(4):168-177.
  • Thy, P., K.H. Esbensen, B.M. Jenkins. 2009. On representative sampling and reliable chemical characterization in thermal biomass conversion studies. Biomass and Bioenergy 33(11):1513-1519.
  • Thy, P., B.M. Jenkins. 2009. Mercury in biomass feedstock and combustion residuals. Water, Air, and Soil Pollution DOI 10.1007/s11270-009-0211-9.
  • Budhavaram, N. K., Fan, Z. 2009. Production of Lactic acid from Paper Sludge using Acid-tolerant, Thermophilic Bacillus coagulan Strains Bioresource. Tech . 100: 5966-5972 El-Mashad, H.M., R.H. Zhang and R.J. Avena-Bustillos. 2008. A two-step process for biodiesel production from fish oil. Biosystems Engineering, 99(2008):220-227.
  • Zheng, Y., Z. Pan, R.H. Zhang, D. Wang, J. Labavitch, B.M. Jenkins. 2008. Non-ionic Surfactants and Noncatalytic Protein on Enzymatic Hydrolysis of Saline Creeping Wild Rye Grass. Applied Biochemistry and Biotechnology. 146:231-248.
  • Jia, Y.H., H.T. Tran, D.H. Kim, S.J. Oh. D.H. Park, R.H. Zhang and D.H. Ahn. 2008. Simultaneous organics removal and bio-electrochemical denitrification in microbial fuel cells. Bioprocess and Biosystems Engineering. 31:315-321.
  • Chiou, B.S., H. M. El-Mashad, R.J. Avena-Bustillos, R.O. Dunn, P. J. Bechtel, T.H. McHugh, S.H. Imam, G.M. Glenn, W.J. Orts, and R. Zhang. 2008. Biodiesel from Waste Salmon Oil. Transactions of the ASABE. 51(2):1-6.
  • Pan, J. R. Zhang, H.M. El-Mashad, H. Sun, Y. Ying. 2008. Effect of food to microorganism ratio on biohydrogen production from food waste via anaerobic fermentation, International Journal of Hydrogen Energy. 33(2008):6968-6975.
  • Rapport, J. L., R.H. Zhang, R.Williams and B.M. Jenkins. 2008. Anaerobic digestion technologies for the treatment of municipal solid waste. International Journal of Environment and Waste Management.
  • Zheng, Y., Z. Pan, R.H. Zhang. 2008. Anaerobic digestion of saline creeping wild ryegrass for biogas production and pretreatment of particleboard material. Bioresource Technology. 100(2009):1582-1588.
  • Zhu, B., P. Gikas, R.H. Zhang, J. Lord, B.M. Jenskins, X. Li. Characteristics and Biogas Production Potential of municipal solid waste pretreated with rotating drum reactor. Bioresource Technology. 100(2009):1122-1129.
  • Romano, R.R., R.H. Zhang, S. Teter, and J.A. McGarvey. The effect of enzyme addition on anaerobic bigestion of wheat grass. Bioresource Technology. 100(2009):4561-4571.
  • Zhang, R.H., Y. Ma, J. A. McGarvey and F.Mitlouhner.. 2008 Effects of anaerobic digestion and aerobic treatment on reduction of gaseous emissions from dairy manure storages. International Journal of Agricultural and Biological Engineering. 1(2):15-20.
  • Zheng, Y., Z. Pan and R.H. Zhang. 2009. Overview of biomass pretreatment for cellulosic ethanol production. International Journal of Agricultural and Biological Engineering. 2(3)1-18.
  • Zheng, Y, Z. Pan, and R.H. Zhang, D. Wang. 2009. Optimization of enzymatic hydrolysis of dilute acid pretreated Creeping Wild Ryegrass for fermentable sugar production. Applied Energy, 86, 2459-2465.
  • El-Mashed, H. M., J.A. McGarvey and R.H. Zhang. Performance and microbial analysis of anaerobic digesters treating food waste and dairy manure. 2009. Biological Engineering. 1(3):
  • Liu, G. R.H. Zhang, H.M. El-Mashed and R. Dong. 2009. Effect of feed to inoculum ratios on biogas yields of food and green wastes. Bioresource Technology. 100(2009)5103-5108.
  • Zhang, R.H., J. Rapport, P. Gikas, B. Zhu, B. Jenkins,J. Lord, C. Choate. 2009. Rotary drum reactors for pretreatment of municipal solid waste prior to anaerobic digestion. Biocycle. April (2009): 20-25.