Location: Renewable Product Technology Research
2019 Annual Report
Objectives
This project develops commercially targeted technologies for producing value added bioproducts, such as specialty/commodity chemicals and biopolymers made from renewable agriculture feedstocks or biomass. Materials being investigated in this project have potential for significant market expansion and address the growing demand for improved manufacturing of products made with renewable technology. We work closely with industrial collaborators, stakeholders, and customers to ensure that goals are compatible with market needs and will ultimately strengthen our energy independence, improve sustainable agriculture, and provide economic support to rural communities.
Goals for this project include the following specific objectives:
Objective 1: Enable, from a technological standpoint, fungal-based processes for the commercial production of carboxylic acids and microbial oils. Sub-Objective 1.1: Enhance productivity and yield of microbial oils synthesized by Aureobasidium pullulans. Sub-Objective 1.2: Improve current methods for the fermentative production of carboxylic acids by Rhizopus.
Objective 2: Enable chemical and enzymatic processes for the commercial production of (1) sugar-based biopolymers/oligosaccharides and (2) ethers derived from sugars or polyols. Sub-Objective 2.1: Develop biocatalytic processes for the production of novel biopolymers and oligomers from agricultural feedstocks. Sub-Objective 2.2: Develop renewable chemical processes for the synthesis of valuable sugar/polyol-based ethers.
Approach
The objectives of this research are achieved using strategies that include microbial strain development, fermentation technology, bacterial/fungal/yeast biotechnology, microbial bioengineering, enzyme technology, chemical/biochemical syntheses, and analytical analyses using state of the art equipment. Approaches for this project currently include the following areas of research:
Specialty Oils. In this project, we develop advanced technologies for the production of specialty microbial oils, called liamocins, which are produced by certain strains of the fungus Aureobasidium. Liamocins are a family of novel oils that have significant potential for numerous veterinary, medical, industrial and food applications. However, the technology for large-scale production of liamocin is currently underdeveloped and is only economical for high-value applications. This work provides further development towards the commercialization of liamocins by increasing the yield and desired type of product through a combination of specialized techniques.
Carboxylic Acids. We utilize metabolic engineering technology to enhance the production of carboxylic acids by the fungus Rhizopus, which is used in industry to convert sugars obtained from agricultural crops to this important commodity chemical. Carboxylic acids, such as fumaric and lactic acid, are natural fermentation biochemicals that are utilized for the manufacture of several environmentally friendly products, such as biodegradable plastics and cleaning solvents. In order to allow the market potential to continue expanding, it is important that the production costs are minimized by the development of new and improved technologies.
Novel Biopolymers and Oligomers. We work on technologies to synthesize unique water-insoluble biopolymers using enzymatic conversion of agriculturally-derived sugars. These polymers are similar to dextrans, which are utilized in a large number of industrial, medical, and food applications. We identify, characterize, and modify novel microorganisms/enzymes that have potential for production of biodegradable products (e.g., fibers, films, encapsulation materials) for a broad number of consumer applications. In addition, we develop novel oligomers (i.e., short sugar chains) that have potential to promote the growth of healthy intestinal bacteria and potentially inhibit pathogens. In order to bring this technology to maturity, we continue improving these processes and develop further novel products made with these methods.
Chemical Conversion of Sugars. We develop environmentally-friendly technologies that are capable of converting sugars to a class of compounds, called ethers, which are used extensively in many industrial applications. Ethers made from sugars have valuable potential applications as drop-in renewable alternatives for solvents, lubricants, and waxes. Chemical based conversion of sugars has immense potential to synthesize these important compounds, but progress is hampered by difficulties with reactions that typically involve toxic compounds. Therefore, we continue to explore and develop safer technologies and examine additional applications and products.
Progress Report
Progress was made on both objectives of the research project which addresses research needs to discover and develop commercially viable biobased materials and conversion processes; and to improve biobased material performance and processing through enhanced knowledge of their structure/property relationships. This project addresses the National Program (NP) 306 (Quality and Utilization of Agricultural Products) Action Plan, Statement 2B-Enable technologies for (1) expanding market applications for existing biobased products, or (2) producing new marketable non-food biobased products derived from agricultural products and byproducts, and ensure that these technologies will generate economic impact by estimating their potential economic value. In addition, ARS scientists in Peoria, Illinois, continue to develop new technologies that support these efforts and lead to new areas of research.
Specific examples of significant developments in FY 2019 include the following:
Under Objective 1, progress was made on improving genetic modification methods for the fungus Aureobasidium. This allowed us to more easily delete genes involved in the production of a dark pigment, called melanin, which often contaminants bioproducts made by this organism. Eliminating the requirement to perform post-production cleanup to remove this impurity facilitates the commercial development of liamocin and other important Aureobasidium bioproducts such as pullulan. ARS scientists are now utilizing this technology in collaboration with industrial partners to improve existing production methods of commercially available bioproducts.
Under Objective 2, progress was made on increasing production of a novel sugar called, isomelezitose. This rare sugar is often produced as a byproduct in small amounts by a group of called glucansucrases. ARS scientists have used genetic engineering to modify one of these enzymes to significantly improve synthesis of isomelezitose. Optimization of production and purification methods have further enhanced yields of this product to levels far in excess of any previously reported studies. ARS scientists have also demonstrated that this unique sugar may be useful in numerous food and biotechnology applications. We are currently working with industrial partners to accelerate the commercialization of this product.
ARS scientists have also identified several unique glucansucrase enzymes through our previous genome sequencing efforts. These enzymes were shown to be very efficient for synthesis of novel oligosaccharides, which are comprised of a small number of sugars linked together. These types of oligosaccharides are important because they often encourage growth of probiotics (i.e. healthy bacteria) and have been shown to have anti-inflammatory properties. Efforts are now focused on increasing production of these oligosaccharides for further testing.
Other significant progress:
Tunicamycin is a unique antibiotic that that can be combined with other antibiotics as a method to overcome antibiotic resistance in certain types of bacteria, but the toxicity of this antibiotic prevents it from being used for clinical applications. ARS scientists recently developed technology to chemically modify tunicamycin so that it has drastically reduced toxicity, while still increasing the efficacy of penicillin- based drugs up to 128-fold. They are now working with other ARS researchers to test these modified tunicamycins in combination with penicillins for the treatment of Johne’s Disease, a highly contagious and usually fatal infection in cattle, has been conducted.
Accomplishments
1. Improving antimicrobial activity of modified antibiotics. The alarming growth of antibiotic resistance threatens agriculture and human health, so development of new antimicrobial strategies is critical to combating this problem. Tunicamycin is a powerful antibiotic that can be combined with other antibiotics in order to improve their efficacy and often overcome antimicrobial resistance, but toxicity in human and animal cells prevents it from being used for therapeutic applications. ARS scientists in Peoria, Illinois, have developed technology to chemically modify tunicamycin to have significantly less toxicity while still retaining the antimicrobial properties. These same researchers also recently determined that natural structural variants of tunicamycin, which differ in length and branching of an attached fatty acid chain, have significantly altered binding to bacterial cell wall components and distinct antimicrobial activities. This significant discovery will now allow ARS scientists to specifically focus research efforts on synthesis of tunicamycin structures that have the desired characteristics. This will further improve the use of these modified antibiotics and is an important step towards combating drug resistance.
Review Publications
Ispirli, H., Simsek, Ö., Skory, C.D., Sagdic, O., Dertli, E. 2018. Characterization of a 4,6-a-glucanotransferase from Lactobacillus reuteri E81 and production of malto-oligosaccharides with immune-modulatory roles. International Journal of Biological Macromolecules. 124:1213-1219. https://doi.org/10.1016/j.ijbiomac.2018.12.050.
Leathers, T.D., Rich, J.O., Bischoff, K.M., Skory, C.D., Nunnally, M.S. 2019. Inhibition of Streptococcus mutans and S. sobrinus biofilms by liamocins from Aureobasidium pullulans. Biotechnology Reports. 21:e00300. https://doi.org/10.1016/j.btre.2018.e00300.
Hay, W.T., Fanta, G.F., Felker, F.C., Peterson, S.C., Skory, C.D., Hojilla-Evangelista, M.P., Biresaw, G., Selling, G.W. 2019. Emulsification properties of amylose-fatty sodium salt inclusion complexes. Food Hydrocolloids. 90:490-499. https://doi.org/10.1016/j.foodhyd.2018.12.038.
Saunders, L.P., Bischoff, K., Bowman, M.J., Leathers, T.D. 2018. Inhibition of Lactobacillus biofilm growth in fuel ethanol fermentations by Bacillus. Bioresource Technology. 272:156-161. https://doi.org/10.1016/j.biortech.2018.10.016.
Price, N.P.J., Hartman, T.M., Vermillion, K.E. 2018. Thiazolidine peracetates: carbohydrate derivatives that readily assign cis-, trans-2,3- monosaccharides by gas chromatography - mass spectrometry analysis. Analytical Chemistry. 90(13):8044-8050. doi:10.1021/acs.analchem.8b00976.
Naumann, T.A., Price, N.P.J. 2018. Purification and in vitro activities of a chitinase-modifying protein from the corn ear rot pathogen Stenocarpella maydis. Physiological and Molecular Plant Pathology 106:74-80. https://doi.org/10.1016/j.pmpp.2018.12.004.
Jackson, M.A., Price, N.P., Blackburn, J.A., Peterson, S.C., Kenar, J.A., Haasch, R., Chen, C. 2019. Partial hydrodeoxygenation of corn cob hydrolysate over palladium catalysts to produce 1-hydroxy-2-pentanone. Applied Catalysis A: General. 577:52-61. https://doi.org/10.1016/j.apcata.2019.03.019.
Bantchev, G.B., Cermak, S.C., Durham, A.L., Price, N.P. 2019. Estolide molecular weight distribution via gel permeation chromatography. Journal of the American Oil Chemists' Society. 96(4):365-380. https://doi.org/10.1002/aocs.12165.
