SBIR-STTR Award

The USDA has established research on renewable energy as a high priority. Cellulosic ethanol from perennial grasses has the potential to become an important component of America's effort to reduce its dependence on foreign oil and alleviate the buildup of greenhouse gases. For energy crops to become viable biofuels, they must become cost-competitive with foreign oil and provide environmental benefits. This will require improvements in refining technologies to efficiently convert cellulose to ethanol, improvements in agronomic productivity, and maximal greenhouse gas reduction. GrassRoots Biotechnology is working to create enhanced energy crops by focusing on improved root architecture. In this project, we are focusing on two main components of root architecture: 1) greater deep root extension to increase drought resistance and carbon sequestration; 2) maintaining sufficient shallow root spread for nutrient acquisition from topsoil. OBJECTIVES: In Phase I, we propose to determine the ideal growing conditions for imaging root growth of switchgrass and Brachypodium distachyon, an emerging grass model whose genome sequence is now available. We will also image and analyze cultivars of switchgrass and Brachypodium distachyon to identify variation in root traits. In Phase II, we propose to identify root traits associated with deep root extension and shallow root spread, and determine the heritability of these traits. We will also determine the effect of different nutritional and environmental stimuli on these traits. The ultimate goal of this work is to modify root architecture in energy crops for enhanced agronomic productivity and carbon sequestration. APPROACH: To identify the genetic determinants that allow a root system to integrate and respond to its environment we need to be able to "watch" roots grow over time and have a means of quantitatively comparing the system architectures that are formed. Two non-invasive imaging technologies will be used to acquire images of growing switchgrass and Brachypodium roots. We have developed a pipeline for analysis of 2D root images that is both flexible and efficient. Images are pre-processed with a variety of conventional image analysis methods and then analyzed using a set of automated computational tools that extract (i) geometric; (ii) network; and (iii) topological information and build upon prior methods to characterize root system architecture traits. We have successfully developed a prototype automatic classification pipeline for analysis of root system architecture using support vector machine (SVM) methods. The pipeline takes individual root images, calculates ten distinct statistical features, and then uses an SVM classifier to find a hyperplane (or hypersurface) that best separates the data into two categories. The SVM approach has the additional benefit of indicating which root architecture features differentiate between varieties. We will measure many root parameters and determine which are likely to be associated with deep root extension to increase drought resistance and carbon sequestration and which are associated with shallow root spread for nutrient acquisition from topsoil. We will then determine how these traits are affected by altering the availability of two essential nutrients, nitrogen and phosphorous. We will perform a heritability analyses for one or more of the traits that we identify. This could be a QTL analysis for traits under normal growth conditions or under nutrient deprivation conditions. A draft assembly of the Brachypodium genome sequence is already available and an annotated sequence should be available by the time we enter Phase II of the project. Thus, performing the QTL analysis in B. dystachion will facilitate going from QTL to gene. Once we have identified genes that will optimize root development in switchgrass and other energy crops, we will seek to form strategic partnerships with plant breeders and/or agricultural biotechnology companies to exploit these discoveries

Despite its clean and green nature when utilized in fuel cells and other devices, most hydrogen is currently produced primarily from non-renewable sources, such as natural gas, oil, and coal. Anaerobic digestion provides a potentially improved alternative to manufacturing hydrogen from petroleum and natural gas. Anaerobic digesters can produce hydrogen from inexpensive and renewable energy sources such as organic wastes (e.g. food processing waste and animal waste). Recent studies have shown that certain strains of bacteria (e.g. bacteria from the genus Clostridium) are effective at producing hydrogen as a by-product during anaerobic digestion With help from two USDA grants, one of them an SBIR grant in 2005, a new type of high rate anaerobic digester was develop and patented by Hansen Energy and Environmental (HEE) and called the Andigen Induced Blanket Reactor (IBR). Andigen was licensed to sells the patented IBR anaerobic digester. Then again in 2006 with the help from a USDA Rural Development Grant, HEE operated a 1996 Chevy truck on biogas (methane) produced by the IBR digester. A review of literature revealed that if a small percentage (10% to 20%) of hydrogen is added to methane (biogas) it crates a much better fuel and reduces emissions by as much as 50%. Therefore, research into the production of hydrogen was started that resulted in the SBIR phase I award. The technology of the Andigen IBR high rate digester has the ability to control the parameters needed to produce hydrogen on a continuous flow through basis using agricultural waste products, including animal manure and food waste Phase I replicated earlier lab trials for anaerobic production of hydrogen and was able to determine the best method to inhibit the growth of methane producing bacteria in order to create an environment where the hydrogen producing bacteria would thrive. The method used needed to have the potential to work in the Andigen IBR digester. The lab trials have resulted in a patented process that has produced 40% hydrogen. With phase II funding a continuous flow through process for the production of hydrogen will be developed using agricultural waste products that have little or no value. This Phase II project will also show that there is a net energy gain from the same amount of waste by producing both biohydrogen and biomethane. By using biogas as a fuel for his trucks and tractors a farmer can realize a much higher return on his digester investment than the generation of electricity. The production of hydrogen has many commercial applications in a "hydrogen society" but the commercial application waiting for the development of this technology is for use as a fuel mixture for the biomethane and biohydrogen produced by the Andigen IBR digesters. This Phase II grant will also include a collaborative effort with Ceramatec of Salt Lake City, Utah to produce a liquid fuel (synthetic Diesel) from hydrogen. An economic analysis will be made to examine the economic feasibility in producing a liquid fuel from the biohydrogen. Ceramatec has made great strides in cutting the cost of producing synthetic diesel from biogas. OBJECTIVES: 1. Produce fermentative hydrogen and volatile fatty acids using manure and cheese processing waste (whey) in a pilot scale Andigen IBR digester system until an optimum mixture for hydrogen production can be determined. 2. Determine flow ratios of cheese whey and manure so as to be able to regulate pH in the digester with low pH whey, stressing the mixed bacterial culture enough to inhibit the production of methane producing bacteria. Procedures for preparing the different cultures will follow the research that has been patented (U.S. patent #7,540,961). 3. Confirm that the control system on the Andigen IBR digester will monitor and control flow rates, temperature and pH in order to produce hydrogen on a continuous basis in a flow through system according to parameters discussed in objective two. 4. Feed the effluent from the hydrogen producing IBR to the methane producing IBR Show that there is a net gain in energy out put. 5. Build a large scale hydrogen producing tank at a full scale IBR facility located at the Blaine Wade dairy. The Hydrogen producing tank will have a capacity of 5000 to 7000 gallons. Show that the biohydrogen producing IBR tank can be operated as a continuous flow through system. 6. Commingle effluent from the hydrogen digestion with manure influent to the full scale IBR digester to replicate the results in Phase I and show an increase in overall energy output. 7. Produce both biohydrogen and biomethane on a continuous basis. 8. Clean the biogas with the onsite biogas cleaning system. 9. Deliver biohydrogen and biomethane to Ceramatec to be made into liquid fuels 10. Calculate the economics of taking both biohydrogen and biomethane to a liquid fuel. APPROACH: Phase II research will rebuild and commence using the pilot scale as a flow through system for hydrogen production. Under the first objective, hydrogen and valitial fatty acids VFA will be produced in the pilot scale by adding the right prepared bacteria (patent #7,540,961) and controlling pH. The pilot scale IBR's will be operated in the USU Bioengineering Lab and/or the Andigen IBR facility on the Blaine Wade dairy. VFA is unavoidably produced when methanogens are inhibited, but in the proposed continuous system, these VFA's will be fed to a methanogenic IBR and methane will be produced Objectives two and three are to demonstrate that hydrogen production can be controlled in a continuous process by controlling the feed ratios of cheese whey (low pH) and manure. In objective four, the process of commingling the effluent from the hydrogen producing IBR with manure to enhance methane production will be demonstrated. Methane is enhanced because the hydrogen producing IBR effluent is rich in VFA that is easily and quickly converted to methane with the methanogenic bacteria in a normally operated IBR. The purpose of objectives five and six are to scale up the process demonstrated in pilot scale. A larger scale (5000 to 7000 gallon) hydrogen producing IBR will be built and operated next to an operating full sized methane producing digester system (the Andigen IBR digester on the Blaine Wade dairy). The larger scale hydrogen producing IBR digester will be operated to produce hydrogen by again controlling bacteria starter and pH. An onsite inexpensive biogas cleaning system (patent pending) will be used to clean the biohydrogen and biomethane for the combined biofuel research. Mixing will be done on a volumetric basis. Objective 9 will be completed by delivering biogas to Ceramatec, 2425 South, 900 West, Salt Lake City, Utah 84119. Ceramatec will make a synthetic diesel from the biohydrogen. The purpose of this objective is to compare the economics of using the combined hydrogen and methane as a fuel as compared to the extra step of turning it into a liquid fuel(synthetic diesel). Through collaborative efforts in the past Ceramatec has processed methane biogas from an Andigen digester into liquid fuels and determined that with additional research it could be economical