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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
2006 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? Why 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.

This project is part of National Program 307 and it specifically addresses the Ethanol component "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 by year the currently approved milestones (indicators of research progress)
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.

Years 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.

Years 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.

Years 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.List the single most significant research accomplishment during FY 2006.
Processes to Produce Corn Germ Peptides with Valuable Biological Properties. Recently there has been considerable interest in nutraceuticals or engineered foods containing bioactive compounds with health promoting or disease preventing properties. It is important to identify bioactive compounds from food proteins whenever possible. Numerous bioactive peptides have been identified, and most are derived from milk and dairy products. Another excellent food source with the potential to be developed into high valued nutritional products is corn germ proteins which have been basically ignored. One way to increase the value of corn germ proteins would be to treat them with enzymes to create peptides with valuable biological properties. Biologically active peptides are usually short chain peptides that can act as physiological modulators in the body. Previous researchers have reported that some peptides from dairy proteins could inhibit Angiotensin I Converting Enzyme and this inhibition resulted in a reduction of blood pressure and several other physiological effects. Although no inhibitory peptides were detected in the unhydrolyzed wet- or dry-milled corn germ, treatment of wet milled corn germ with protease (thermolysin) produced peptides that exhibited inhibitory properties. Similar experiments conducted with dry milled corn germ revealed that thermolysin and two other proteases (trypsin and flavourzyme) produced peptides that showed inhibition. This is the first demonstration that specific protein hydrolysates from corn germ might have biologically useful properties. These results indicate that it may be possible to treat corn germ proteins with enzymes to increase their inhibitory properties and increase their value. This research supports the ARS National Program 307, Bioenergy and Energy Alternatives, Component 1, Ethanol Coproduct Development.


4b.List other significant research accomplishment(s), if any.
Development of new processes to obtain corn oil from corn germ by Aqueous Enzymatic Oil Extraction (AEOE) There is a need to develop new environmentally safe processes for the extraction of corn oil from wet milled corn germ. We are developing aqueous enzymatic corn oil extraction methods as a solution to this problem. During the last year we scaled this process from 6 grams of germ to 200 grams of germ. Corn germ dispersions were created by preheating corn germ in a pressure reactor, blending, mixing with a buffer solution to produce germ concentrations of 5 to 20% and churning in an incubator/shaker. Preheating improved the oil yield by swelling the germ and making it more susceptible to blending. The incubator churning produced a maximum corn oil yield from dispersions with between 0.8 and 1.1 g/cm2 of churning vessel cross section. Based on this finding, and churning trials with a variety of internal rollers, it now appears that milling the germ controls the oil yield and agitation of the mixture during churning is important to collecting the released oil droplets. For aqueous enzymatic oil extraction (AEOE) runs, a solution of enzymes was added to the germ dispersion prior to churning. The AEOE runs exhibited 90% higher oil yields than comparable non-enzymatic aqueous extractions. Both types of extraction gave the highest oil yields after cooking at 120°C and then churning at 70°C. These results indicate that AEOE methods may be a safer alternative to hexane extraction of corn oil. This research supports the ARS National Program 307, Bioenergy and Energy Alternatives, Component 1, Ethanol Coproduct Development.

Processes to Produce Corn Germ Peptides with Antimicrobial Properties. There is a need to increase the value and commercial application for corn germ proteins. In addition to the corn germ peptide project described in 4a, research efforts have also been focused on developing processes to produce antimicrobial peptides from corn germ. Numerous peptides with antimicrobial activity have been isolated from dairy products, amphibian sources, marine fish sources and cereals. Although the mechanism of action is not completely understood, antimicrobial peptide activity is usually expressed by the disintegration of cell membranes. This project has practical importance because there is evidence that increasing levels of bacterial resistance to antibiotics have been caused by the overuse of antibiotics in humans and as an additive in livestock feeds. Antimicrobial peptides might be useful replacements for antibiotics in animal feeds, which should help to prevent the development of future antibiotic-resistant strains. As part of this project, on the aqueous enzymatic extraction of oil from corn germ, the objective of our research was to develop bioprocesses to enhance the value of proteins from the de-oiled germ. Albumin and globulin proteins, in de-oiled commercial wet- and dry-milled corn germ, were extracted with saline solution followed by aqueous alcohol extraction of zein proteins from the residue after centrifugation. Inhibitory activity of these proteins and their peptides, after hydrolysis with trypsin or thermolysin was determined against Escherichia coli O157:H7, Listeria monocytogenes or Salmonella Anatum. Zein hydrolyzed with trypsin exhibited strongest inhibitory activity against L. monocytogenes in initial bioassays. These results indicate that it may be possible to treat corn germ proteins with enzymes to increase their antibiotic properties and increase their value as feed ingredients. This research supports the ARS National Program 307, Bioenergy and Energy Alternatives, Component 1, Ethanol Coproduct Development.

Cooking Dry Milled Corn Germ Increases Corn Oil Yields using Aqueous Enzymatic Oil Extraction (AEOE) Previously we developed an AEOE process to obtain corn oil from corn germ from a commercial corn wet mill. We are now adapting this process to obtain corn oil from corn germ obtained from a commercial corn dry mill. We found that when "raw" (uncooked or undried) dry milled corn germ is used as feedstock for our aqueous enzymatic extraction method, no oil is obtained unless the corn germ is cooked or heat-dried. We compared various methods of cooking and drying the germ in microwave and conventional ovens before AEOE. We believe that cooking probably weakens or disrupts the oil body membrane and this is probably required to release the oil. In our previous studies with wet milled corn germ excellent oil yields were achieved without cooking, but wet milled corn germ is subjected to harsher conditions than dry milled corn germ (it is subjected to steeping with SO2 and lactic acid and it is dried in driers where it is subjected to high temperatures). Although cooking may be required before AEOE with dry milled corn germ, cooking is energy-intensive and we have some preliminary data that suggests that it may be possible to replace cooking with various types of enzyme pretreatments. This research should result in the development of a process to obtain corn oil from dry milled corn germ without the use of hexane, which is becoming increasingly expensive to use industrially because of tougher environmental and safety regulations. This research supports the ARS National Program 307, Bioenergy and Energy Alternatives, Component 1, Ethanol Coproduct Development.


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


4d.Progress report.
The following section serves to document research conducted under a CRADA between ARS and the Illinois Corn Marketing Board. During the last year researchers at ERRC continued to compare the composition of conventional corn oil (corn oil obtained by pressing and/or hexane extraction of corn germ) versus corn oil obtained by extracting ground corn with ethanol. The analyses focused on comparing the levels of "functional lipids" such as phytosterols, tocopherols, tocotrienols, and carotenoids in the two oils. The results indicated that the levels of each of these functional lipids were higher in the corn oil from ethanol-extracted ground corn than in conventional corn oil.


5.Describe the major accomplishments to date and their predicted or actual impact.
A comparison of the properties of the proteins in corn germ from wet and dry mills. Corn germ is a coproduct of corn wet milling (an industrial process to obtain cornstarch), corn dry milling (an industrial process to produce stable corn ingredients for human food use), 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%). 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. This research supports the ARS National Program 307, Bioenergy and Energy Alternatives, Component 1, Ethanol Coproduct Development.


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 was 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. Scientists from the two largest enzyme companies have visited ERRC to learn about our Aqueous Enzymatic Corn Oil Extraction process. Both companies have expressed interest in developing a CRADA with ARS on this technology. Scientists from a major US food company have expressed interest in our process for encapsulating bioactive materials in zein for delivery into the colon.

Our US Patent Application, "Methods of Preparing Corn Fiber Oil and of Recovering Corn Aleurone Cells from Corn Fiber" (0012.01) was recently "Allowed" by the USPTO.


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, R.A., D.B. Johnston, and K.B. Hicks, The Influence of Moisture Content and Cooking on the Screw Pressing and Pre-Pressing of Corn Oil from Corn Germ, Oil Mill Gazetteer 111:8-10, March 2006.

Moreau, R.A., D.B. Johnston, and K.B. Hicks, The Influence of Moisture Content and Cooking on the Screw Pressing and Pre-Pressing of Corn Oil from Corn Germ, INFORM 17: 348-350, June 2006.


Review Publications
Dickey, L.C., Kurantz, M.J., Goldberg, N.M., Parris, N. 2005. Separation of particles from ethanol/maize extracts: an inexpensive alternative to centrifugation. Industrial Crops and Products. 23/3, p.264-272.

Parris, N., Moreau, R., Johnston, D. B., Singh, V. and Dickey, L.C., 2006. Protein Distribution in Commercial Wet- and Dry-Milled Corn Germ. Journal of Agricultural and Food Chemistry. 54:4868-4872.

Moreau, R.A., Johnston, D., Hicks, K.B. 2005. The influence of moisture content and cooking on the screw pressing and pre-pressing of corn oil from corn germ. Journal of the American Oil Chemists' Society. Vol. 82, No. 11,p.851-854.

Moreau, R.A., Hicks, K.B. 2005. The composition of corn oil obtained by the alcohol extraction of ground corn. Journal of the American Oil Chemists' Society. V. 82, No. 11. p. 809-815.

Moreau, R.A. 2006. An overview of modern mass spectrometry methods in the toolbox of lipid chemists and biochemists. In "New Techniques and Applications in Lipid Analysis and Lipidomics" ed. by M. M. Mossoba, J.K.G.Kramer, J.T. Brenna, and R. E. McDonald, AOCS Press, Champaign, 2006. 448pp.

Dickey, L.C., Moreau, R.A., Parris, N. Churning corn germ dispersions to separate oil. Presentation at the Corn Utilization and Technology Conference, Dallas, TX, June 5-7, 2006. Poster Session.

Parris, N., Moreau, R.A., Johnston, D., Singh, V., Dickey, L.C. 2006.Protein distribution in commercial wet- and dry-milled corn germ. Presentation at the AOCS Annual meeting, April 30-May 4, 2006, St. Louis, MO., Protein and Co-Products Poster.

Moreau, R.A., Hicks, K.B., Parris, N., Dickey, L.C. 2006. Processes to produce functional foods during the conversion of grains to fuel ethanol. Presentation at the ACS Annual Meeting, Atlanta, GA, March 26-30, 2006. Symposium on Healthy Products from Agricultural By-Products, Oral Paper 6.

Wanjie, S.W., Welti, R., Moreau, R.A., Chapman, K.D. 2005. Identification and quantification of lipid metabolites in cotton fibers: reconciliation with metabolic pathway predictions from dna databases. Lipids Journal. V. 40, No. 8, p.773-785.

Srinivasan, R., Singh, V., Belyea, R.L., Rausch, K.D., Moreau, R.A., Tumbleson, M.E. 2006. Economics of fiber separation from distillers dried grains with solubles (ddgs) using sieving and elutriation. Cereal Chemistry.83(4):p.324-330.

Srinivasan, R., Moreau, R.A., Rausch, K.D., Belyea, R.L., Tumbleson, M.E., Singh, V. 2005. Removal of fiber from distillers dried grains with solubles (ddgs) using sieving and elutriation. Cereal Chemistry 82, p.528-533.

Last Modified: 4/20/2014
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