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United States Department of Agriculture

Agricultural Research Service

Research Project: AQUEOUS ENZYMATIC EXTRACTION OF CORN OIL AND VALUE-ADDED PRODUCTS FROM CORN GERM PRODUCED IN NEW GENERATION DRY-GRIND ETHANOL PROCESSES
2005 Annual Report


1.What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? What does it matter?
Fuel ethanol production in the U.S. reached 3.57 billion gallons in 2004 (a 27% increase from the 2.81 billion gallons produced in 2003 and a 233% increase from the 1.53 billion gallons produced in 2000). Most of the expansion in the fuel ethanol industry has come from building moderately sized (30-50 Million Gallon per Year, MGY) dry-grind corn-to-ethanol plants. While dry-grind plants produce fewer and less-valuable coproducts (DDGS and sometimes carbon dioxide) than wet-mill ethanol plants (coproducts include corn germ/corn oil, corn gluten meal, corn gluten feed, and sometimes CO2), the capital expense to build wet mills is prohibitive. Hence, almost all new ethanol plants are of the dry-grind design. Each generation of dry-grind plant has become more efficient, through constant research and innovation. A current major target in planning for the next generation of dry-grind ethanol plants is to create modified dry-grind facilities that, like wet mills, produce higher valued coproducts.

The Quick Germ and Quick Fiber processes developed at the University of IL are two examples of higher value coproducts that could be produced by new generation dry grind ethanol processes. At least nine other new-generation dry grind processes are in development. The 2002 DOE-USDA Biomass R&D Grants program awarded major grants to a dry grind ethanol plant designer/builder (Broin and Associates, Sioux Falls, SD, Second Generation Dry Mill Biorefinery), and an R&D firm (Grain Value, St. Paul, MN, GrainValue Process). Both grants were awarded to assist in the development of processes to remove germ and fiber coproducts from kernels prior to fermentation. A key to the success of these projects will be the actual value that a dry-grind facility can obtain from these coproducts. Crude corn oil is presently at an all time high value, over $.30 per pound. High quality corn germ, such as that produced from wet-mills, brings a premium price due to the strong demand and price for corn oil. Corn germ is processed to corn oil, however, in a small number of hexane extraction facilities. Most of these facilities are operated by large wet mill companies. Because of this, if dry-grind facilities started producing corn germ, they would be dependent on a limited number of oil extractors to purchase their germ. Whereas wet-milled germ has nearly 45-50% oil content, germ from dry-grind processes may only have 20-25% oil plus other soluble protein and carbohydrate components. This new type of germ would sell at an even greater discount, possibly eroding the economic benefit of producing a corn germ coproduct. One solution to this oil extraction problem would be for modified dry-grind plants to build their own hexane extraction facilities as part of their unit operation, however, because of the issues noted below, this is unlikely.

Problems associated with hexane have made hexane extraction plants expensive to build, run and maintain, due to the explosion hazard potential and the human and environmental safety issues and regulations. In fact, because of these issues, some wet mills have even shut down their own extraction facilities and now sell their germ to competitors. To maximize the value of new process streams of corn germ, this project is being proposed. Our goal is to develop a new aqueous/enzymatic process for the extraction of corn oil from the new types of corn germ produced at new generation ethanol plants using technology that could be scaled up and commercialized without the large capital investment and operating cost of a hexane extraction facility. Similar processes have been proposed before but they were not successful due to technology limitations. In this project, we plan to take advantage of new enzymes, not previously available, to develop new engineering techniques for oil separations, and to separate corn germ protein and carbohydrate coproducts that will be integrated into the overall process design. The process will be modeled using SuperPro Designer computer simulation software and an integrated cost model will be developed. Sensitivity studies will be conducted on key process parameters to guide the research toward more cost effective processes. The result of the project, if successful, will be a state-of-the-art enzymatic process for production of corn oil, proteins, and carbohydrates that could be incorporated into moderate sized and larger new generation dry-grind ethanol plants. The process would solve a major problem for value-added utilization of corn germ from these new generation ethanol processes. Additionally, it is likely that some wet-mills and traditional corn dry milling (food) plants that currently do not have in-house oil extraction facilities will adopt this process for their germ, as these industries have recognized a need for new oil extraction technologies in the past.

From the aqueous enzymatic corn oil extraction research, we anticipate: A) increased amounts of corn oil that is comparable in quality and safer to produce than from current technology, B) new types of animal feed with higher protein and perhaps lower fiber, and C) increased yields of fuel ethanol (per bushel of corn) produced via fermentation of corn germ carbohydrates. The identification and development of new corn peptide products with antimicrobial and prebiotic properties may have valuable applications in the animal feed industry and other areas of agriculture.

Dry-grind ethanol companies will benefit from the research directly and opportunities for research partnerships and CRADAS with them will be explored. In addition, development of a more economical and safer extraction process, such as an aqueous enzymatic extraction process, may also benefit both corn wet mills and corn dry mills by providing alternatives to hexane extraction. Our research focuses on NP 307 Action Plan Component I. Ethanol, and the goal is to perform research "To improve process economics, value-added coproducts will be developed from current byproducts and separation technologies will be improved for ethanol as well as coproduct recovery."


2.List the milestones (indicators of progress) from your Project Plan.
Year 1 (FY 2005) A1. Begin development of a bench scale AEE method to maximize oil yields from commercial wet milled corn germ.

A2. Begin development of a bench scale AEE method to maximize oil yields from commercial dry milled corn germ.

A3. Begin development of a bench scale AEE method for corn germ from "new generation" processes.

B1. Begin characterization of the protein types and carbohydrate types in corn germ from various sources to determine potential uses and value.

B2. Begin development of processes to separate the water soluble and insoluble components in corn germ* from various sources.

C1. Begin identification and analyses of the proteins in protein-rich germ fractions and begin evaluation of feed applications.

C3. Begin treatments of proteins in protein-rich germ fractions with proteases and characterize the structure and potential uses/value of the peptides.

Year 2 (FY 2006) A1. Complete development of a bench scale AEE method to maximize oil yields from commercial wet milled corn germ.

A2. Complete development of a bench scale AEE method to maximize oil yields from commercial dry milled corn germ.

A3. Continue development of a bench scale AEE method for corn germ from "new generation" processes.

A4. Begin scale up and economic analysis of an AEE method for corn germ from wet mill, dry mill, and “new generation" processes.

B1. Complete characterization of the protein types and carbohydrate types in corn germ from various sources to determine potential uses and value.

B2. Complete development of processes to separate the water soluble and insoluble components in corn germ from various sources.

B3. Begin the development of bench scale processes to separate a protein-rich fraction and a carbohydrate-rich fraction from corn germ.

B4. Begin the development and economic analysis of pilot scale processes to separate a protein-rich fraction and a carbohydrate-rich fraction from corn germ.

C1. Continue identification and analyses of the proteins in protein-rich germ fractions and continue evaluation of feed applications.

C2. Begin evaluation of feed, gum and fermentation applications in carbohydrate-rich germ fractions.

C3. Continue treatments of proteins in protein-rich germ fractions with proteases and continue to characterize the structure and potential uses/value of the peptides.

C4. Begin providing corn germ peptide fractions to various collaborators to evaluate anti-microbial and prebiotic activities.

Year 3 (FY 2007) A3. Continue development of a bench scale AEE method for corn germ from "new generation" processes.

A4. Continue scale up and economic analysis of an AEE method for corn germ from wet mill, dry mill, and “new generation" processes.

B3. Complete development of bench scale processes to separate a protein-rich fraction and a carbohydrate-rich fraction from corn germ.

B4. Continue development and economic analysis of pilot scale processes to separate a protein-rich fraction and a carbohydrate-rich fraction from corn germ.

C1. Complete identification and analyses of the proteins in protein-rich germ fractions and complete evaluation of feed applications.

C2. Continue evaluation of feed, gum and fermentation applications in carbohydrate-rich germ fractions.

C3. Complete treatments of proteins in protein-rich germ fractions with proteases and complete the characterization of the structure and potential uses/value of the peptides

C4. Continue providing corn germ peptide fractions to various collaborators to evaluate anti-microbial and prebiotic activities.

Year 4 (FY 2008) A3. Complete development of a bench scale AEE method for corn germ from "new generation" processes.

A4. Continue scale up and economic analysis of an AEE method for corn germ from wet mill, dry mill, and "new generation" processes.

B4. Continue development and economic analysis of pilot scale processes to separate a protein-rich fraction and a carbohydrate-rich fraction from corn germ.

C2. Continue evaluation of feed, gum and fermentation applications in carbohydrate-rich germ fractions.

C4. Continue providing corn germ peptide fractions to various collaborators to evaluate anti-microbial and prebiotic activities.

Year 5 (FY 2009) A4. Complete scale up and economic analysis of an AEE method for corn germ from wet mill, dry mill, and "new generation" processes and in the latter, determine the effects on the economics of an ethanol plant using this process.

B4. Complete development and economic analysis of pilot scale processes to separate a protein-rich fraction and a carbohydrate-rich fraction from corn germ. Determine effect on economics of an ethanol plant using this process.

C2. Complete evaluation of feed, gum and fermentation applications in carbohydrate-rich germ fractions.

C4. Complete the process of providing corn germ peptide fractions to various collaborators and determine their anti-microbial and prebiotic potential and uses. Summarize evaluation results.


4a.What was the single most significant accomplishment this past year?
Bioactive peptides from corn germ. There is a need to create more valuable co-products from corn germ proteins that are currently sold as low-valued cattle feed. Demonstrated for the first time that bioactive peptides could be generated by treating proteins from dry milled corn germ with commercial proteases (trypsin and thermolysin). These corn germ peptides were found to have two different biological activities, a) they caused inhibition of growth of three pathogenic bacterial species, Escherichia coli 0157:H7, Listeria monocytogenes, and Salmonella anatum and b) they caused inhibition of angiotensin converting enzyme (ACE). The identification of new antimicrobial peptides is of practical importance because they could potentially be valuable for use in animal feeds. Antimicrobial peptides may provide a natural alternative to the use of antibiotics in animal feeds, as a means to inhibit the growth of pathogenic microbes and encourage the growth of beneficial colonic microbes (e.g. Lactobaccillus and Biffidobacteria species). The practice of adding antibiotics to animal feeds is being curtailed in many countries, partly due to the growing problem caused by bacterial resistance to the current antibiotics because of their overuse in humans and livestock. In addition, bioactive peptides from dry milled corn germ that demonstrate ACE inhibition could potentially be used for the treatment of hypertension (high blood pressure).


4b.List other significant accomplishments, if any.
Development of an aqueous enzymatic extraction method for dry milled corn germ. There is a need to develop cost-effective alternatives to hexane extraction for corn oil extraction. A method was developed to use dry milled corn germ as a feedstock to obtain corn oil using aqueous enzymatic extraction. Dry milled corn germ is a coproduct of a corn dry mill which produces food grade corn products such as grits for use in breakfast cereals. Currently, commercial corn oil is obtained from dry milled corn germ (which contains about 20 gram of oil per 100 grams of germ) via screw-pressing or hexane extraction. Using aqueous methods, combined with various commercial enzymes, we have been able to remove about half of the oil from dry milled corn germ. Additional work is required to identify the correct combination of enzymes and the correct process steps to achieve higher yields of corn oil from dry milled corn germ. Higher yields will be necessary for our aqueous enzymatic oil extraction process to compete economically with current commercial hexane extraction and pressing processes.

Characterization of the proteins in corn germ. Corn germ is a coproduct of corn wet milling, corn dry milling, and very recently corn dry grind ethanol production. All commercial corn oil is obtained by extracting corn germ. Although corn germ typically contains more oil when it is produced by wet (40-50% oil) versus dry (20-25% oil) processes, the levels of protein in corn germ produced by all processes have been reported to be similar, approximately 14-16% protein (based on nitrogen). Our recent research indicates that the levels of digestible protein in corn germ may actually be much lower than previously reported – we measured 11.7% protein in dry milled germ and 4.9% protein in wet milled germ. These large differences in protein values in the two types of germ are probably attributable to the much higher levels of nonprotein nitrogen that we measured in wet milled corn germ (6.6%) versus dry milled corn germ (1.5%). During wet milling, it is thought that some of the water- and salt-soluble proteins are probably removed during the SO2 steeping step. Gel electrophoresis (used to separate and visualize the individual proteins) revealed that many of the proteins in wet milled germ had very low molecular weights, indicating that they had undergone degradation/hydrolysis, perhaps as a result of steeping. To better understand the composition of corn germ proteins during commercial processing, we carefully excised the germ from yellow dent corn kernels and found that the protein pattern, as visualized by gel electrophoresis, more closely resembles that of dry- rather than wet-milled corn. This information should aid in developing bioprocesses to enhance the value of proteins from corn germ. As new generation processes are developed to remove germ before dry grind ethanol production, our research indicates that it will be important to evaluate and compare the quality of the protein in germ produced by various processes.

Use of a screw-press to pre-press oil from corn germ. There is a need to develop cost effective processes to produce corn oil. Experiments were conducted with a screw-press to investigate the effect of pre-pressing on the yields of oil from wet milled corn germ and dry milled corn germ using the aqueous enzymatic extraction. Pressing is a common industrial process to remove oil from oilseeds and corn germ. Our studies indicated that pre-pressing corn germ removed about 2/3 of the oil, but the remaining oil in the germ was not more accessible to removal via aqueous enzymatic extraction. We concluded that pre-pressing is probably not a cost-effective process to include as a pretreatment before aqueous enzymatic extraction.

Evaluation of a published peanut oil aqueous extraction method for corn germ extraction. There is a need to develop cost effective methods to obtain corn oil. Experiments were conducted using a published aqueous oil extraction process (which did not include enzymes) to extract corn oil from corn germ. This method was developed to extract peanut oil from whole peanuts. Some adaptations were made to make it more effective at extracting oil from corn germ. Although the process does not yet provide acceptable oil yields, it provides a control to compare with our aqueous enzymatic extraction process, and it should be helpful in the development of the necessary slurry fractionation steps to develop protein and carbohydrate separation methods.

Bioactive components in ethanol extracted corn kernel oil. Although all commercial corn oil is produced by the hexane extraction of corn germ, there is a need to study the composition of corn oil obtained by other processes, such as extracting oil from the whole corn kernel with ethanol. In 2005 we negotiated a one year CRADA (591-1935-197) An Examination of the Bioactive Components in Corn Germ and Corn Oil Obtained by Alternative Processes, with the Illinois Corn Marketing Board. Funds obtained via the CRADA ($25,000) were partially used to fund the maintenance allowance for a visiting scientist from Korea, who will conduct some of the research on this project. The research results obtained in this CRADA will determine the chemical composition of ethanol-extracted corn oil and determine whether the unique compounds in this oil are beneficial (thus making the oil more valuable) or harmful (creating a need to develop processes to remove them).


4c.List any significant activities that support special target populations.
None


4d.Progress report.
Question 4d: None


5.Describe the major accomplishments over the life of the project, including their predicted or actual impact.
Question 5: Because this project has only been in effect for one year, the major accomplishment over the life of the project is the same as 4A above, the most significant accomplishment during FY 2005.


6.What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end-user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products?
Our patent for Amaizing Oil (US 5,843,499) was licensed to a US company from 2001 until early 2004. It is now available for licensing. In 2005, three companies expressed interest in licensing the patent. Before Amaizing Oil can be commercialized, there is a need to conduct toxicological studies and a clinical feeding study to establish the safety and efficacy of corn fiber oil.

Our corn fiber gum patent (US 6,147,206, co-owned by a major US starch and specialty chemical manufacturer) issued on 11/14/00. Through a CRADA with the patent co-owner, we transferred technology for decreasing capital costs for commercial corn fiber gum production, which is the last barrier to commercialization. During 2005 we learned that that patent co-owner is moving forward with commercialization and they expect to begin production in late 2006.

Samples of our zein isolate were provided to: A) a company for evaluation as formulation in an aqueous dispersion to coat fresh cut produce such as iceberg lettuce and B) a second company for evaluation as a of bio degradable water repellant coating material for seeds. Our cost estimates for the use of our zein isolates for these applications are favorable, but more research is needed to determine if zein is superior to other current coatings.

Samples of essential oils were encapsulated with zein and provided to a company to evaluate their effects on colonic microorganisms as a potential replacement for antibiotics in animal feeds. If these studies demonstrate that encapsulation with zein protects the essential oils from degradation in the stomach and small intestine, zein coating of other materials for delivery in the colon may be a promising new application.


7.List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below).
Moreau, Robert. A., The Aqueous Enzymatic Extraction of Corn Oil from Corn Germ, Presentation at Genencor International, Palo Alto, CA, January 27, 2005.


Review Publications
Dickey, L.C., Parris, N. 2005. Improving particle separation from an ethanol extract to water: settling dependence on fine particle content. Industrial Crops and Products. V.21/3. p.379-385.

Dien, B.S., Nagle, N., Singh, V., Moreau, R.A., Nichols, N.N., Johnston, D., Cotta, M.A., Hicks, K.B., Nguyen, Q., Bothast, R.J. 2004. Fermentation of "quick fiber" produced from a modified corn milling process into ethanol and recovery of corn fiber oil. Journal of Applied Biochemistry and Biotechnology. 115:937-950.

Lampi, A., Moreau, R.A., Piironen, V., Hicks, K.B. 2004. Pearling barley and rye to produce phytosterol-rich fractions. Lipids, V.39, NO. 8.P.783-787.

Mcgovern, P.E., Zhang, J., Tang, J., Zhang, Z., Hall, G.R., Moreau, R.A., Nunez, A. 2004. Fermented beverages of pre- and protohistoric china. Nature. PNAS. V. 101, NO. 5. P.17593-17598.

Mellon, J.E., Moreau, R.A. 2004. Inhibition of aflatoxin biosynthesis in Aspergillus flavus by diferuloylputrescine and p-coumaroylferuloylputrescine. Journal of Agricultural and Food Chemistry. 52(21):6660-6663.

Moreau, R.A., Hicks, K.B. 2004. The in vitro rates of hydrolysis of phytosterol conjugates and other plant lipids by mammalian digestive enzymes, compared to saponification. Lipids, v.39, No. 8. p.769-776.

Moreau, R.A., Johnston, D., Powell, M.J., Hicks, K.B. 2004. A comparison of commercial enzymes for the aqueous enzymatic extraction of corn oil from corn germ. Journal of the American Oil Chemists' Society, V.81, No. 11, p.1071-1075.

Moreau, R.A. Phytosterols and phytosterol esters. Book Chapter. 2005. C. Akoh and O-M. Lai (eds.), in Healthful Lipids, AOCS Press, Champaign, pp.335-360.

Moreau, R.A. Extraction and analysis of food lipids. Book Chapter. S. Otles (ed), in Methods of Analysis of Food Components and Additives, CRC Press/Taylor and Francis, Boca Raton, pp.97-110 2005.

Moreau, R.A. The evaporative light-scattering detector as a tool for the analysis of lipids by hplc. Book Chapter. J-T Lin and T.A. McKeon (eds), in HPLC of Acyl Lipids, H.N.B. Publishing, New York, pp.93-116, 2005.

Moreau, R.A. Corn oil. Book Chapter. F. Shahidi (ed), in Bailey's Industrial Oil & Fat Products, Sixth Edition, Volume 2, Edible Oil & Fat Products: Edible Oils, Wiley-Interscience, Hoboken, pp.149-172, 2005.

Moreau, R.A., Nunez, A. 2005. Fermented beverages from ancient china. Inform 16:326-327.

Moreau, R.A., Johnston, D., Dickey, L.C., Hicks, K.B. The influence of moisture content and cooking on the screw pressing of corn oil from corn germ. Meeting Abstract. 96th AOCS Annual Meeting & Expo. Salt Lake City, UT, May 1-5, 2005, Processing Poster 1.

Moreau, R.A., Powell, M.J., Johnston, D., Hicks, K.B. Aqueous enzymatic extraction of corn oil from corn germ. Meeting Abstract. 96th AOCS Anual Meeting & Expo., Salt Lake City, UT, May 1-5, 2005. Biotechnology Poster 1.

Nunez, A., Moreau, R.A., Foglia, T.A. Liquid chromatography/mass spectrometry for the analysis of biosurfant glycolipids secreted by microorganisms. Book Chapter. W. C. Byrdwell (ed) in Modern Methods for Lipids Analysis by Liquid Chromatography/Mass Spectrometry and Related Techniques, pp.447-471, 2005.

Parris, N., Fett, W.F., Dickey, L.C., Moreau, R.A. Bioactive peptides from corn germ proteins. Meeting Abstract. 96th AOCS Annual Meeting & Expo. Salt Lake City, UT, May 1-5, 2005. Session PCP 1.1 Protein and Coproducts: Bioactive Peptides, Oral Paper 9.

Parris, N., Cooke, P.H., Hicks, K.B. 2005. Encapsulation of essential oils in zein nanospherical particles as a delivery system for antimicrobials. Journal of Agricultural and Food Chemistry. V.53,No.12,p.4788-4792

Parris, N., Douds, D.D., Dickey, L.C., Moreau, R.A., Phillips, J.G. 2004. Effect of hydrophilic zein films on the growth of tomato plants and evaporative water loss. Hort Science.39(6).p.1324-1326.

Dien, B.S., Nagle, N., Singh, V., Moreau, R.A., Tucker, M.P., Nichols, N.N., Johnston, D., Cotta, M.A., Hicks, K.B., Nguyen, Q., Bothast, R.J. 2005. Review of process for producing corn fiber oil and ethanol from "Quick Fiber." International Sugar Journal. 107(1275):187-191.

Last Modified: 4/24/2014
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