Vol. 83, No. 4, 2006 317 Dry-Grind Processing of Corn with Endogenous Liquefaction Enzymes Vijay Singh, 1,2 Christopher J. Batie, 3 George W. Aux, 3 Kent D. Rausch, 1 and Carl Miller 3 ABSTRACT Cereal Chem. 83(4):317–320 An amylase corn has been developed that produces an α-amylase enzyme that is activated in the presence of water at elevated temperatures (>70°C). Amylase corn in the dry-grind process was evaluated and com- pared with the performance of exogenous amylases used in dry-grind processing. Amylase corn (1–10% by weight) was added to dent corn (of the same genetic background as the amylase corn) as treatments and resulting samples were evaluated for dry-grind ethanol fermentation using 150-g and 3-kg laboratory procedures. Ethanol concentrations during fermentation were compared with the control treatment (0% amylase corn addition or 100% dent corn) which was processed with a conventional amount of exogenous α-amylase enzymes used in the dry-grind corn process. The 1% amylase corn treatment (adding 1% amylase corn to dent corn) was sufficient to liquefy starch into dextrins. Following fermen- tation, ethanol concentrations from the 1% amylase corn treatment were similar to that of the control. Peak and breakdown viscosities of liquefied slurries for all amylase corn treatments were significantly higher than the control treatment. In contrast, final viscosities of liquefied slurries for all amylase corn treatments were lower than those of the control. Protein, fat, ash, and crude fiber contents of DDGS samples from the 3% amylase corn treatment and control were similar. Production of ethanol in the United States has increased in the last several years. Most of the recent increase in ethanol capacity in the United States has come from construction or expansion of dry-grind corn processing plants. The increase in ethanol capacity from dry-grind corn plants is expected to continue for the next several years due to replacement of MTBE in motor fuels and the passage of the Energy Bill of 2005. In a conventional dry-grind process, corn is ground and mixed with water to produce slurry, which is cooked, liquefied, saccharified, and fermented to produce ethanol. Conventional liquefaction and saccharification of slurry uses exogenous α-amylase and glucoamylase enzymes, respective- ly. Liquefaction enzymes break down starch molecules into lower molecular weight dextrins; glucoamylase enzymes break down the dextrins into glucose. Remaining nonfermentables in corn (germ, fiber, and protein) are recovered at the end of the dry-grind process as a coproduct called distillers dried grains with solubles (DDGS). New advances are being made to improve the dry-grind corn process and increase profitability of ethanol production. These include development of new and modified dry-grind processes (Hojilla-Evangelista et al 1992; Bryan 2005; Singh et al 2005), recovery of valuable coproducts (Singh and Eckhoff 1996; Singh et al 1999; Hojilla-Evangelista and Johnson 2003), better and lower cost enzymes (Anonymous 2005), and high-ethanol-yielding corn hybrids (Haefele et al 2004; Singh and Graeber 2005). Recently, an amylase corn has been developed by transgenic tech- nology that produces and stores α-amylase within the kernel (Lanahan 2003). These enzymes are activated in the presence of water at elevated temperatures (>70°C). In conventional dry-grind corn processing, exogenous α-amylase enzymes are added during liquefaction and are one of the operating costs in dry-grind processing. Expression levels of the α-amylase in the amylase corn are high and, therefore, it is expected that only small amounts of amylase corn will need to be added to regular dent corn to achieve adequate liquefaction. Performance of amylase corn in the dry-grind process and the requirements of amylase corn addition are not known. Use of amy- lase corn in dry-grind ethanol processing was evaluated and the performance was compared with exogenous amylases currently used in the dry-grind process. MATERIALS AND METHODS Experimental Material and Design Transgenic amylase corn and dent corn with the same genetic background were obtained from a commercial seed company (Syn- genta Biotechnology, Inc., Research Triangle Park, NC). Corn samples were hand-cleaned to remove broken corn and foreign material, packaged in plastic bags, and stored at 4°C until pro- cessing. Whole kernel moisture content was measured using the 103°C convection oven method (Approved Method 44-15A, AACC International 2000). Starch content of amylase (71%) and dent (72%) corn was determined by near-infrared transmittance (GrainSpec, Foss North America, Minneapolis, MN) using Corn Refiners Association standard methods (CRA 1980). Three experiments evaluated the performance of amylase corn in the dry-grind process. In the first experiment, four levels (0, 3, 5, and 10%) of amylase corn were added to the dent corn as treat- ments, and samples were evaluated in a 150-g laboratory dry-grind procedure. Zero percent amylase corn treatment (0% amylase corn addition or 100% dent corn) was the control treatment. No exogen- ous α-amylase enzyme was added, except for the control treat- ment in which normal amounts of exogenous α-amylase enzyme was added. A second experiment evaluated four amylase corn treatments (0, 1, 2, and 3%) using a 150-g dry-grind procedure. A third experiment was conducted in which a 3% amylase corn treat- ment was compared with a control treatment in a 3-kg dry-grind procedure. 150-g Dry-Grind Laboratory Process Corn samples were ground in a laboratory hammer mill (model MHM4, Glen Mills, Clifton, NJ) equipped with a 2.0-mm sieve and operated at 500 rpm. Ground corn weighing 150 g was mixed with tap water (35°C) to obtain slurry containing 27% solids (db). All runs were done in 1,000-mL flasks in a shaking water bath (model DHOD-182, Bellco Glass, Vineland, NJ). The first experiment used sequential saccharification and fer- mentation after liquefaction for the control treatment. Samples were liquefied by increasing the temperature of the slurry to 90 ± 0.5°C. No α-amylase enzyme was added to 3, 5, and 10% amylase corn treatments. α-Amylase (0.2 mL, α-amylase solution Bacillus lichenformis, type XII-A saline solution 500–1,000 units/mg of protein, 1,4-α-D-glucan-glucanohydrolase, 9000-85-5, Sigma-Ald- rich, St. Louis, MO) was added to the control treatment (0% amylase corn or 100% dent corn). The slurry was held at 90°C for 90 min with continuous agitation at 150 rpm. After 90 min, slurry 1 Agricultural and Biological Engineering Department, University of Illinois at Urbana- Champaign, Urbana, IL. Mention of brand or firm names does not constitute an endorsement by the University of Illinois above others of a similar nature not mentioned. 2 Corresponding author. Phone: 217-333-9510. Fax: 217-244-0323. E-mail: vsingh @uiuc.edu 3 Syngenta Biotechnology Inc., Research Triangle Park, NC 27709. DOI: 10.1094/CC-83-0317 © 2006 AACC International, Inc.