US20040234649A1 - Method for producing ethanol using raw starch - Google Patents

Method for producing ethanol using raw starch Download PDF

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Publication number
US20040234649A1
US20040234649A1 US10/798,226 US79822604A US2004234649A1 US 20040234649 A1 US20040234649 A1 US 20040234649A1 US 79822604 A US79822604 A US 79822604A US 2004234649 A1 US2004234649 A1 US 2004234649A1
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United States
Prior art keywords
fermenting
saccharifying
fermentation
ethanol
plant material
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Abandoned
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US10/798,226
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English (en)
Inventor
Stephen Lewis
Shon Van Hulzen
John Finck
Debbie Roth
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Broin and Associates Inc
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Broin and Associates Inc
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Priority to US10/798,226 priority Critical patent/US20040234649A1/en
Application filed by Broin and Associates Inc filed Critical Broin and Associates Inc
Assigned to BROIN AND ASSOCIATES, INC. reassignment BROIN AND ASSOCIATES, INC. ASSIGNMENT OF ASSIGNORS INTEREST (SEE DOCUMENT FOR DETAILS). Assignors: FINCK, JOHN MICHALE, ROTH, DEBBIE LYNN, LEWIS, STEPHEN M., VAN HULZEN, SHON ERRON
Publication of US20040234649A1 publication Critical patent/US20040234649A1/en
Priority to US11/077,969 priority patent/US20050233030A1/en
Priority to US11/077,971 priority patent/US20050239181A1/en
Assigned to BANK OF OKLAHOMA, NATIONAL ASSOCIATION, AS ADMINISTRATIVE AGENT reassignment BANK OF OKLAHOMA, NATIONAL ASSOCIATION, AS ADMINISTRATIVE AGENT SECURITY AGREEMENT Assignors: BROIN AND ASSOCIATES, INC.
Priority to US11/676,965 priority patent/US7842484B2/en
Priority to US11/682,195 priority patent/US8409640B2/en
Priority to US12/405,125 priority patent/US7919291B2/en
Priority to US12/886,483 priority patent/US8470550B2/en
Priority to US12/944,618 priority patent/US8409639B2/en
Priority to US12/944,606 priority patent/US8497082B2/en
Priority to US13/850,196 priority patent/US8748141B2/en
Priority to US13/866,002 priority patent/US8679793B2/en
Priority to US14/285,030 priority patent/US20140283226A1/en
Assigned to POET, LLC F/K/A BROIN COMPANIES, LLC, POET RESEARCH, INC. F/K/A BROIN AND ASSOCIATES, INC., POET PLANT MANAGEMENT, LLC, POET INVESTMENTS, INC. F/K/A BROIN ENTERPRISES, INC. reassignment POET, LLC F/K/A BROIN COMPANIES, LLC RELEASE OF PATENT, TRADEMARK AND COPYRIGHT SECURITY INTEREST Assignors: BOKF, N.A., DBA BANK OF OKLAHOMA
Priority to US15/957,818 priority patent/US20180235167A1/en
Abandoned legal-status Critical Current

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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L7/00—Cereal-derived products; Malt products; Preparation or treatment thereof
    • A23L7/10—Cereal-derived products
    • A23L7/104—Fermentation of farinaceous cereal or cereal material; Addition of enzymes or microorganisms
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L7/00—Cereal-derived products; Malt products; Preparation or treatment thereof
    • A23L7/20—Malt products
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
    • C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
    • C12P7/06—Ethanol, i.e. non-beverage
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
    • Y02E50/00—Technologies for the production of fuel of non-fossil origin
    • Y02E50/10—Biofuels, e.g. bio-diesel

Definitions

  • the present invention relates to methods for producing high levels of alcohol during fermentation of plant material, and to the high alcohol beer produced.
  • the present invention also relates to methods for producing high protein distiller's dried grain from fermentation of plant material, and to the high protein distiller's dried grain produced.
  • the present invention further relates to reduced stack emissions from drying distillation products from the production of ethanol.
  • the present invention relates to methods for producing high levels of alcohol during fermentation of plant material, and to the high alcohol beer produced.
  • the present invention also relates to methods for producing high protein distiller's dried grain from fermentation of plant material, and to the high protein distiller's dried grain produced.
  • the present invention relates to a process for producing ethanol from plant material.
  • This method includes grinding the plant material to produce ground plant material including starch; saccharifying the starch, without cooking; fermenting the incubated starch; and recovering the ethanol from the fermentation.
  • the present method can include varying the temperature during fermentation.
  • the present method can include employing a plant material with a particle size such that more than 50% of the material fits though a sieve with a 0.5 mm mesh.
  • the present method can yield a composition including at least 18 vol-% ethanol.
  • the present invention relates to a process for producing high protein distiller's dried grain from plant material.
  • This method includes grinding the plant material to produce ground plant material including starch; producing sugars from the starch without cooking; fermenting the uncooked sugars to yield a composition including ethanol; and recovering distiller's dried grain from the fermentation.
  • the distiller's dried grain can include at least about 30% protein.
  • the distillers dried grain can include increased levels of the protein zein.
  • the present invention relates to a process of producing ethanol from corn.
  • This process includes producing starch from corn and ethanol from the starch; producing dryer stack emissions including a significantly lower level of volatile organic compounds than conventional technologies.
  • FIGS. 1 A-E schematically illustrate a comparison of the yield of the process of the present invention compared to the conventional process.
  • FIGS. 2A-2C schematically illustrate the effect of dosages of glucoamylase and acid fungal amylase in the present process.
  • FIGS. 3A-3D schematically illustrate the effect of grind size and enzyme dosage on fermentation efficiency in the present process.
  • FIGS. 4A-4C schematically illustrate the effect of grind particle size, glucoamylase type, and acid fungal amylase dosage on fermentation efficiency in the present process.
  • FIGS. 5A-5J schematically illustrate the effect of initial dry solids and temperature on fermentation performance in the present process.
  • FIGS. 6A and 6B schematically illustrate high levels of ethanol production from the process of the present invention using simultaneous saccharification and fermentation (SSF) batch or continuous modes of operation.
  • SSF simultaneous saccharification and fermentation
  • FIG. 7 schematically illustrates that the present process maintained low levels of glycerol during SSF batch operations.
  • FIG. 8 schematically illustrates that the present process maintained low levels of fusel oils during SSF batch operations.
  • FIGS. 9A and 9B schematically illustrate that the present process maintained low levels of glucose during SSF batch or continuous fermentation modes of operation.
  • FIGS. 10A and 10B schematically illustrate that the present process maintained low levels of maltose during SSF batch or continuous fermentation modes of operation.
  • FIGS. 11A and 11B schematically illustrate that the present process maintained low levels of maltotriose (DP3) during SSF batch or continuous fermentation modes of operation.
  • DP3 maltotriose
  • FIGS. 12A and 12B schematically illustrate that the present process maintained low levels of dextrins (DP4+) during SSF batch or continuous fermentation modes of operation.
  • FIG. 13 schematically illustrates that the present process impacts DDGS quality favorably based on caking tendency.
  • FIGS. 14A and 14B schematically illustrate mass balance of the present process related to proximate separations during the centrifugation step of ethanol production.
  • FIGS. 15 A-D schematically illustrate that the present process affords advantageous fermentation of non traditional feedstocks.
  • FIGS. 16 A-C schematically illustrate that the process of the present invention is capable of stable operation in a continuous mode of operation without significant loss due to acid producing bacterial contaminants.
  • FIG. 17 schematically illustrates that the present process is capable of achieving low residual starch levels in a continuous mode of operation.
  • without cooking refers to a process for converting starch to ethanol without heat treatment for gelatinization and dextrinization of starch using alpha-amylase.
  • “without cooking” refers to maintaining a temperature below starch gelatinization temperatures, so that saccharification occurs directly from the raw native insoluble starch to soluble glucose while bypassing conventional starch gelatinization conditions.
  • Starch gelatinization temperatures are typically in a range of 57° C. to 93° C. depending on the starch source and polymer type.
  • dextrinization of starch using conventional liquefaction techniques is not necessary for efficient fermentation of the carbohydrate in the grain.
  • plant material refers to all or part of any plant (e.g., cereal grain), typically a material including starch.
  • Suitable plant material includes grains such as maize (corn, e.g., whole ground corn), sorghum (milo), barley, wheat, rye, rice, and millet; and starchy root crops, tubers, or roots such as sweet potato and cassaya.
  • the plant material can be a mixture of such materials and byproducts of such materials, e.g., corn fiber, corn cobs, stover, or other cellulose and hemicellulose containing materials such as wood or plant residues.
  • Suitable plant materials include corn, either standard corn or waxy corn.
  • saccharification and “saccharifying” refer to the process of converting starch to smaller polysaccharides and eventually to monosaccharides, such as glucose.
  • Conventional saccharification uses liquefaction of gelatinized starch to create soluble dextrinized substrate which glucoamylase enzyme hydrolyzes to glucose.
  • saccharification refers to converting raw starch to glucose with enzymes, e.g., glucoamylase and acid fungal amylase (AFAU).
  • AFAU acid fungal amylase
  • the raw starch is not subjected to conventional liquefaction and gelatinization to create a conventional dextrinized substrate.
  • a unit of acid fungal amylase activity refers to the standard Novozymes units for measuring acid fungal amylase activity.
  • the Novozymes units are described in a Novozymes technical bulletin SOP No. EB-SM-0259.02/01. Such units can be measured by detecting products of starch degradation by iodine titration.
  • 1 unit is defined as the amount of enzyme that degrades 5.260 mg starch dry matter per hour under standard conditions.
  • a unit of glucoamylase activity refers to the standard Novozymes units for measuring glucoamylase activity.
  • the Novozymes units and assays for determining glucoamylase activity are described in a publicly available Novozymes technical bulletin.
  • a unit of amyloglucosidase activity refers to the standard Novozymes units for measuring amyloglucosidase activity.
  • the Novozymes units are described in a Novozymes technical bulletin SOP No EB-SM-0131.02/01.
  • Such units can be measured by detecting conversion of maltose to glucose.
  • the glucose can be determined using the glucose dehydrogenase reaction.
  • 1 unit is defined as the amount of enzyme that catalyzes the conversion of 1 mmol maltose per minute under the given conditions.
  • the term “about” modifying any amount refers to the variation in that amount encountered in real world conditions of producing sugars and ethanol, e.g., in the lab, pilot plant, or production facility.
  • an amount of an ingredient employed in a mixture when modified by “about” includes the variation and degree of care typically employed in measuring in an ethanol production plant or lab.
  • the amount of a component of a product when modified by “about” includes the variation between batches in an ethanol production plant or lab and the variation inherent in the analytical method. Whether or not modified by “about,” the amounts include equivalents to those amounts. Any quantity stated herein and modified by “about” can also be employed in the present invention as the amount not modified by “about.”
  • the present invention relates to methods for producing high levels of alcohol during fermentation of plant material, and to the high alcohol beer produced.
  • the present invention also relates to methods for producing high protein distiller's dried grain from fermentation of plant material, to the high protein distiller's dried grain produced, and to the cleaner dryer stack emissions.
  • the present method converts starch from plant material to ethanol.
  • the present method can include preparing the plant material for saccharification, converting the prepared plant material to sugars without cooking, and fermenting the sugars.
  • the plant material can be prepared for saccharification by any a variety of methods, e.g., by grinding, to make the starch available for saccharification and fermentation.
  • the vegetable material can be ground so that a substantial portion, e.g., a majority, of the ground material fits through a sieve with a 0.1-0.5 mm screen.
  • about 70% or more, of the ground vegetable material can fit through a sieve with a 0.1-0.5 mm screen.
  • the reduced plant material can be mixed with liquid at about 20 to about 50 wt-% or about 25 to about 45 wt-% dry reduced plant material.
  • the present process can include converting reduced plant material to sugars that can be fermented by a microorganism such as yeast.
  • This conversion can be effected by saccharifying the reduced plant material with an enzyme preparation, such as a saccharifying enzyme composition.
  • a saccharifying enzyme composition can include any of a variety of known enzymes suitable for converting reduced plant material to fermentable sugars, such as amylases (e.g., ⁇ -amylase and/or glucoamylase).
  • saccharification is conducted at a pH of about 6.0 or less, for example, about 4.5 to about 5.0.
  • the present process includes fermenting sugars from reduced plant material to ethanol. Fermenting can be effected by a microorganism, such as yeast. In an embodiment, fermentation is conducted at a pH of about 6 or less, for example, about 4.5 to about 5. In an embodiment, the present method can include varying the pH. For example, fermentation can include filling the fermenter at pH of about 3 to about 4.5 during the first half of fill and at a pH of about 4.5 to about 6 during the second half of the fermenter fill cycle. In an embodiment, fermentation is conducted at a temperature of about 25 to about 40° C. or about to about 35° C. In an embodiment, during fermentation the temperature is decreased from about 40° C. to about 30° C.
  • fermentation is conducted for about to (e.g., 24) to about to 150 hours, for example, for about 48 (e.g., 47) to about 96 hours.
  • the present process can include simultaneously converting reduced plant material to sugars and fermenting those sugars with a microorganism such as yeast.
  • the product of the fermentation process is referred to herein as “beer”.
  • Ethanol can be recovered from the fermentation mixture, from the beer, by any of a variety of known processes, such as by distilling.
  • the remaining stillage includes both liquid and solid material.
  • the liquid and solid can be separated by, for example, centrifugation.
  • the present method converts starch from plant material to ethanol.
  • the plant material can be reduced by a variety of methods, e.g., by grinding, to make the starch available for saccharification and fermentation.
  • Other methods of plant material reduction are available.
  • vegetable material such as kernels of corn
  • the use of emulsion technology, rotary pulsation, and other means of particle size reduction can be employed to increase surface area of plant material while raising the effectiveness of flowing the liquefied media.
  • the prepared plant material can be referred to as being or including “raw starch”.
  • a fine grind exposes more surface area of the plant material, or vegetable material, and can facilitate saccharification and fermentation.
  • the vegetable material is ground so that a substantial portion, e.g., a majority, of the ground material fits through a sieve with a 0.1-0.5 mm screen.
  • about 35% or more of the ground vegetable material can fit through a sieve with a 0.1-0.5 mm screen.
  • about 35 to about 70% of the ground vegetable material can fit through a sieve with a 0.1-0.5 mm screen.
  • about 50% or more of the ground vegetable material can fit through a sieve with a 0.1-0.5 mm screen.
  • about 90% of the ground vegetable material can fit through a sieve with a 0.1-0.5 mm screen.
  • all of the ground vegetable material can fit through a sieve with a 0.1-0.5 mm screen.
  • the vegetable material can be fractionated into one or more components.
  • a vegetable material such as a cereal grain or corn can be fractionated into components such as fiber (e.g., corn fiber), germ (e.g., corn germ), and a mixture of starch and protein (e.g., a mixture of corn starch and corn protein).
  • fiber e.g., corn fiber
  • germ e.g., corn germ
  • starch and protein e.g., a mixture of corn starch and corn protein
  • One or a mixture of these components can be fermented in a process according to the present invention.
  • Fractionation of corn or another plant material can be accomplished by any of a variety of methods or apparatus.
  • a system manufactured by Satake can be used to fractionate plant material such as corn.
  • the present process can include converting reduced plant material to sugars that can be fermented by a microorganism such as yeast.
  • This conversion can be effected by saccharifying the reduced plant material with any of a variety of known saccharifying enzyme compositions.
  • the saccharifying enzyme composition includes an amylase, such as an alpha amylase (e.g., acid fungal amylase).
  • the enzyme preparation can also include glucoamylase.
  • the enzyme preparation need not, and, in an embodiment, does not include protease.
  • ethanol production methods according to the present invention can conserve water by reusing process waters (backset) which may contain protease.
  • the present method employs acid fungal amylase for hydrolyzing raw starch.
  • Saccharifying can be conducted without cooking.
  • saccharifying can be conducted by mixing source of saccharifying enzyme composition (e.g., commercial enzyme), yeast, and fermentation ingredients with ground grain and process waters without cooking.
  • source of saccharifying enzyme composition e.g., commercial enzyme
  • yeast e.g., yeast, and fermentation ingredients with ground grain and process waters without cooking.
  • saccharifying can include mixing the reduced plant material with a liquid, which can form a slurry or suspension and adding saccharifying enzyme composition (e.g., at least one of acid fungal amylase and glucoamylase) to the liquid.
  • the method includes mixing the reduced plant material and liquid and then adding the saccharifying enzyme composition (e.g., at least one of acid fungal amylase and glucoamylase).
  • adding enzyme composition can precede or occur simultaneously with mixing.
  • the reduced plant material can be mixed with liquid at about 20 to about 50 wt-%, about 25 to about 45 (e.g., 44) wt-%, about 30 to about 40 (e.g., 39) wt-%, or about 35 wt-% dry reduced plant material.
  • wt-% of reduced plant material in a liquid refers to the percentage of dry substance reduced plant material or dry solids.
  • the method of the present invention can convert raw or native starch (e.g., in dry reduced plant material) to ethanol at a faster rate at higher dry solids levels compared to conventional saccharification with cooking. Although not limiting to the present invention, it is believed that the present method can be practiced at higher dry solids levels because, unlike the conventional process, it does not include gelatinization, which increases viscosity.
  • Suitable liquids include water and a mixture of water and process waters, such as stillage (backset), scrubber water, evaporator condensate or distillate, side stripper water from distillation, or other ethanol plant process waters.
  • the liquid includes water.
  • the liquid includes water in a mixture with about 1 to about 70 vol-% stillage, about 15 to about 60 vol-% stillage, about 30 to about 50 vol-% stillage, or about 40 vol-% stillage.
  • stillage provides nutrients for efficient yeast fermentation, especially free amino nitrogen (FAN) required by yeast.
  • FAN free amino nitrogen
  • the present invention can provide effective fermentation with reduced levels of stillage and even without added stillage.
  • the present method employs a preparation of plant material that supplies sufficient quantity and quality of nitrogen for efficient fermentation under high gravity conditions (e.g., in the presence of high levels of reduced plant material).
  • high gravity conditions e.g., in the presence of high levels of reduced plant material.
  • the present method provides the flexibility to employ high levels of stillage if desired.
  • the present method does not employ conventional liquefaction.
  • Conventional liquefaction increases viscosity of the fermentation mixture and the resulting stillage.
  • the present method produces lower viscosity stillage. Therefore, in an embodiment, increased levels of stillage can be employed in the present method without detrimental increases in viscosity of the fermentation mixture or resulting stillage.
  • Saccharification can employ any of a variety of known enzyme sources (e.g., a microorganism) or compositions to produce fermentable sugars from the reduced plant material.
  • the saccharifying enzyme composition includes an amylase, such as an alpha amylase (e.g., acid fungal amylase) or a glucoamylase.
  • saccharification is conducted at a pH of about 6.0 or less, pH of about 3.0 to about 6.0, about 3.5 to about 6.0, about 4.0 to about 5.0, about 4.0 to about 4.5, or about 4.5 to about 5.0.
  • the initial pH of the saccharification mixture can be adjusted by addition of, for example, ammonia, sulfuric acid, phosphoric acid, process waters (e.g., stillage (backset), evaporator condensate (distillate), side stripper bottoms, and the like), and the like.
  • Activity of certain saccharifying enzyme compositions e.g., at least one of acid fungal amylase and glucoamylase
  • saccharification is conducted at a temperature of about 25 to about 40° C. or about 30 to about 35° C.
  • saccharifying can be carried out employing quantities of saccharifying enzyme composition (e.g., at least one of acid fungal amylase and glucoamylase) selected to maintain low concentrations of dextrin in the fermentation broth.
  • the present process can employ quantities of saccharifying enzyme composition (e.g., at least one of acid fungal amylase and glucoamylase) selected to maintain maltotriose (DP3) at levels at or below about 0.2 wt-% or at or below about 0.1 wt-%.
  • saccharifying enzyme composition e.g., at least one of acid fungal amylase and glucoamylase
  • DP3 maltotriose
  • the present process can employ quantities of saccharifying enzyme composition (e.g., at least one of acid fungal amylase and glucoamylase) selected to maintain dextrin with a degree of polymerization of 4 or more (DP4+) at levels at or below about 1 wt-% or at or below about 0.5 wt-%.
  • saccharifying enzyme composition e.g., at least one of acid fungal amylase and glucoamylase
  • suitable levels of acid fungal amylase and glucoamylase include about 0.3 to about 3 AFAU/gram dry solids reduced plant material (e.g., DSC) of acid fungal amylase and about 1 to about 2.5 (e.g., 2.4) AGU per gram dry solids reduced plant material (e.g., DSC) of glucoamylase.
  • the reaction mixture includes about 1 to about 2 AFAU/gram dry solids reduced plant material (e.g., DSC) of acid fungal amylase and about 1 to about 1.5 AGU per gram dry solids reduced plant material (e.g., DSC) of glucoamylase.
  • saccharifying can be carried out employing quantities of saccharifying enzyme composition (e.g., at least one of acid fungal amylase and glucoamylase) selected to maintain low concentrations of maltose in the fermentation broth.
  • the present process can employ quantities of saccharifying enzyme composition (e.g., at least one of acid fungal amylase and glucoamylase) selected to maintain maltose at levels at or below about 0.3 wt-%.
  • suitable levels of acid fungal amylase and glucoamylase include about 0.3 to about 3 AFAU/gram dry solids reduced plant material (e.g., DSC) of acid fungal amylase and about 1 to about 2.5 (e.g., 2.4) AGU per gram dry solids reduced plant material (e.g., DSC) of glucoamylase.
  • the reaction mixture includes about 1 to about 2 AFAU/gram dry solids reduced plant material (e.g., DSC) of acid fungal amylase and about 1 to about 1.5 AGU per gram dry solids reduced plant material (e.g., DSC) of glucoamylase.
  • the present method employs an ⁇ -amylase.
  • the ⁇ -amylase can be one produced by fungi.
  • the ⁇ -amylase can be one characterized by its ability to hydrolyze carbohydrates under acidic conditions.
  • An amylase produced by fungi and able to hydrolyze carbohydrates under acidic conditions is referred to herein as acid fungal amylase, and is also known as an acid stable fungal ⁇ -amylase.
  • Acid fungal amylase can catalyze the hydrolysis of partially hydrolyzed starch and large oligosaccharides to sugars such as glucose.
  • the acid fungal amylase that can be employed in the present process can be characterized by its ability to aid the hydrolysis of raw or native starch, enhancing the saccharification provided by glucoamylase.
  • the acid fungal amylase produces more maltose than conventional (e.g., bacterial) ⁇ -amylases.
  • Suitable acid fungal amylase can be isolated from any of a variety of fungal species, including Aspergillus, Rhizopus, Mucor, Candida, Coriolus, Endothia, Enthomophtora, Irpex, Penicillium, Sclerotium and Torulopsis species.
  • the acid fungal amylase is thermally stable and is isolated from Aspergillus species, such as A. niger, A. saitoi or A. oryzae , from Mucor species such as M. pusillus or M. miehei , or from Endothia species such as E. parasitica .
  • the acid fungal amylase is isolated from Aspergillus niger .
  • the acid fungal amylase activity can be supplied as an activity in a glucoamylase preparation, or it can be added as a separate enzyme.
  • a suitable acid fungal amylase can be obtained from Novozymes, for example in combination with glucoamylase.
  • the amount of acid fungal amylase employed in the present process can vary according to the enzymatic activity of the amylase preparation. Suitable amounts include about 0.1 to about 10 acid fungal amylase units (AFAU) per gram of dry solids reduced plant material (e.g., dry solids corn (DSC). In an embodiment, the reaction mixture can include about 0.3 to about 3 AFAU/gram dry solids reduced plant material (e.g., DSC). In an embodiment, the reaction mixture can include about 1 to about 2 AFAU/gram dry solids reduced plant material (e.g., DSC).
  • AFAU acid fungal amylase units
  • DSC dry solids corn
  • the reaction mixture can include about 0.3 to about 3 AFAU/gram dry solids reduced plant material (e.g., DSC). In an embodiment, the reaction mixture can include about 1 to about 2 AFAU/gram dry solids reduced plant material (e.g., DSC).
  • the present method can employ a glucoamylase.
  • Glucoamylase is also known as amyloglucosidase and has the systematic name 1,4- alpha -D-glucan glucohydrolase (E.C. 3.2.1.3).
  • Glucoamylase refers to an enzyme that removes successive glucose units from the non-reducing ends of starch.
  • certain glucoamylases can hydrolyze both the linear and branched glucosidic linkages of starch, amylose, and amylopectin.
  • suitable glucoamylases are known and commercially available. For example, suppliers such as Novozymes and Genencor provide glucoamylases.
  • the glucoamylase can be of fungal origin.
  • the amount of glucoamylase employed in the present process can vary according to the enzymatic activity of the amylase preparation. Suitable amounts include about 0.1 to about 6.0 glucoamylase units (AGU) per gram dry solids reduced plant material (e.g., DSC). In an embodiment, the reaction mixture can include about 1 to about 3 AGU per gram dry solids reduced plant material (e.g., DSC). In an embodiment, the reaction mixture can include about 1 to about 2.5 (e.g., 2.4) AGU per gram dry solids reduced plant material (e.g., DSC). In an embodiment, the reaction mixture can include about 1 to about 2 AGU per gram dry solids reduced plant material (e.g., DSC).
  • AGU glucoamylase units
  • the reaction mixture can include about 1 to about 1.5 AGU per gram dry solids reduced plant material (e.g., DSC). In an embodiment, the reaction mixture can include about 1.2 to about 1.5 AGU per gram dry solids reduced plant material (e.g., DSC).
  • the present process includes fermenting sugars from reduced plant material to ethanol.
  • Fermenting can be effected by a microorganism, such as yeast.
  • the fermentation mixture need not, and in an embodiment does not, include protease.
  • the process waters may contain protease.
  • the amount of protease can be less than that used in the conventional process.
  • fermenting is conducted on a starch composition that has not been cooked.
  • the present fermentation process produces potable alcohol. Potable alcohol has only acceptable, nontoxic levels of other alcohols, such as fusel oils.
  • Fermenting can include contacting a mixture including sugars from the reduced plant material with yeast under conditions suitable for growth of the yeast and production of ethanol.
  • fermenting employs the saccharification mixture.
  • yeasts can be employed as the yeast starter in the present process.
  • Suitable yeasts include any of a variety of commercially available yeasts, such as commercial strains of Saccharomyces cerevisiae .
  • Suitable strains include “Fali” (Fleischmann's), Thermosac (Alltech), Ethanol Red (LeSafre), BioFerm AFT (North American Bioproducts), and the like.
  • the yeast is selected to provide rapid growth and fermentation rates in the presence of high temperature and high ethanol levels.
  • Fali yeast has been found to provide good performance as measured by final alcohol content of greater than 17% by volume.
  • the amount of yeast starter employed is selected to effectively produce a commercially significant quantity of ethanol in a suitable time, e.g., less than 75 hours.
  • yeast starter can be added by as a dry batch, or by conditioning/propagating.
  • yeast starter is added as a single inoculation.
  • yeast is added to the fermentation during the fermenter fill at a rate of 5 to 100 pounds of active dry yeast (ADY) per 100,000 gallons of fermentation mash.
  • the yeast can be acclimated or conditioned by incubating about 5 to 50 pounds of ADY per 10,000 gallon volume of fermenter volume in a prefermenter or propagation tank. Incubation can be from 8 to 16 hours during the propagation stage, which is also aerated to encourage yeast growth.
  • the prefermenter used to inoculate the main fermenter is can be from 1 to 10% by volume capacity of the main fermenter, for example, from 2.5 to 5% by volume capacity relative to the main fermenter.
  • the fermentation is conducted at a pH of about 6 or less, pH of about 3 to about 6, about 3.5 to about 6, about 4 to about 5, about 4 to about 4.5, or about 4.5 to about 5.
  • the initial pH of the fermentation mixture can be adjusted by addition of, for example, ammonia, sulfuric acid, phosphoric acid, process waters (e.g., stillage (backset), evaporator condensate (distillate), side stripper bottoms, and the like), and the like.
  • distillery yeast grow well over the pH range of 3 to 6, but are more tolerant of lower pH's down to 3.0 than most contaminant bacterial strains. Contaminating lactic and acetic acid bacteria grow best at pH of 5.0 and above. Thus, in the pH range of 3.0 to 3.5, it is believed that ethanol fermentation will predominate because yeast will grow better than contaminating bacteria.
  • the present method can include varying the pH. It is believed that varying the pH can be conducted to reduce the likelihood of contamination early in fermentation and/or to increase yeast growth and fermentation during the latter stages of fermentation.
  • fermentation can include filling the fermenter at pH of about 3 to about 4.5 during the first half of fill.
  • Fermentation can include increasing the slurry pH to pH of about 4.5 to about 6 during the second half of the fermenter fill cycle.
  • Fermentation can include maintaining pH by adding fresh substrate slurry at the desired pH as described above.
  • pH is not adjusted. Rather, in this embodiment, the pH is determined by the pH of the components during filling.
  • the pH is decreased to about five (5) or below in the corn process waters.
  • the pH is about pH 4 (e.g. 4.1) at the start of fermentation fill and is increased to about pH 5 (e.g. 5.2) toward the end of fermentation fill.
  • the method includes stopping pH control of the mash slurry after the yeast culture becomes established during the initial process of filling the fermenter, and then allowing the pH to drift up in the corn process waters during the end stages of filling the fermenter.
  • fermentation is conducted for about to 25 (e.g., 24) to about to 150 hours, about 25 (e.g., 24) to about 96 hours, about 40 to about 96 hours, about 45 (e.g., 44) to about 96 hours, about 48 (e.g., 47) to about 96 hours.
  • fermentation can be conducted for about 30, about 40, about 50, about 60, or about 70 hours.
  • fermentation can be conducted for about 35, about 45, about 55, about 65, or about 75 hours.
  • fermentation is conducted at a temperature of about 25 to about 40° C. or about 30 to about 35° C.
  • the temperature is decreased from about 40° C. to about 30° C. or about 25° C., or from about 35° C. to about 30° C., during the first half of the fermentation, and the temperature is held at the lower temperature for the second half of the fermentation.
  • the temperature can be decreased as ethanol is produced.
  • the temperature can be as high as about 99° F. and then reduced to about 79° F. This temperature reduction can be coordinated with increased ethanol titers (%) in the fermenter.
  • the present method includes solids staging.
  • Solids staging includes filling at a disproportionately higher level of solids during the initial phase of the fermenter fill cycle to increase initial fermentation rates.
  • the solids concentration of the mash entering the fermenter can then be decreased as ethanol titers increase and/or as the fermenter fill cycle nears completion.
  • the solids concentration can be about 40% (e.g. 41%) during the first half of the fermentation fill. This can be decreased to about 25% after the fermenter is 50% full and continuing until the fermenter fill cycle is concluded. In the above example, such a strategy results in a full fermenter with solids at 33%.
  • solids staging can accelerate enzyme hydrolysis rates and encourage a rapid onset to fermentation by using higher initial fill solids. It is believed that lowering solids in the last half of fill can reduce osmotic pressure related stress effects on the yeast. By maintaining overall fermenter fill solids within a specified range of fermentability, solids staging improves the capacity of the yeast to ferment high gravity mashes toward the end of fermentation.
  • the present process can include simultaneously converting reduced plant material to sugars and fermenting those sugars with a microorganism such as yeast. Simultaneous saccharifying and fermenting can be conducted using the reagents and conditions described above for saccharifying and fermenting.
  • saccharification and fermentation is conducted at a temperature of about 25 to about 40° C. or about 30 to about 35° C.
  • the temperature is decreased from about 40 to about 25° C. or from about 35 to about 30° C. during the first half of the saccharification, and the temperature is held at the lower temperature for the second half of the saccharification.
  • saccharification and fermentation is conducted at a pH of about 6 or less, pH of about 3 to about 6, about 3.5 to about 6, about 4 to about 5, about 4 to about 4.5, or about 4.5 to about 5.
  • the initial pH of the saccharification and fermentation mixture can be adjusted by addition of, for example, ammonia, sulfuric acid, phosphoric acid, process waters (e.g., stillage (backset), evaporator condensate (distillate), side stripper bottoms, and the like), and the like.
  • saccharification and fermentation is conducted for about to 25 (e.g., 24) to about to 150 hours, about 25 (e.g., 24) to about 72 hours, about 45 to about 55 hours, about 50 (e.g., 48) to about 96 hours, about 50 to about 75 hours, or about 60 to about 70 hours.
  • saccharification and fermentation can be conducted for about 30, about 40, about 50, about 60, or about 70 hours.
  • saccharification and fermentation can be conducted for about 35, about 45, about 55, about 65, or about 75 hours.
  • simultaneous saccharifying and fermenting can be carried out employing quantities of enzyme and yeast selected to maintain high concentrations of yeast and high levels of budding of the yeast in the fermentation broth.
  • the present process can employ quantities of enzyme and yeast selected to maintain yeast at or above about 300 cells/mL or at about 300 to about 600 cells/mL.
  • simultaneous saccharifying and fermenting can be carried out employing quantities of enzyme and yeast selected for effective fermentation without added exogenous nitrogen; without added protease; and/or without added backset.
  • Backset can be added, if desired, to consume process water and reduce the amount of wastewater produced by the process.
  • the present process maintains low viscosity during saccharifying and fermenting.
  • simultaneous saccharifying and fermenting can employ acid fungal amylase at about 0.1 to about 10 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 0.5 to about 6 AGU per gram dry solids reduced plant material (e.g., DSC).
  • simultaneous saccharifying and fermenting can employ acid fungal amylase at about 0.3 to about 3 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 3 AGU per gram dry solids reduced plant material (e.g., DSC).
  • simultaneous saccharifying and fermenting can employ acid fungal amylase at about 1 to about 2 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 1.5 AGU per gram dry solids reduced plant material (e.g., DSC).
  • dry solids reduced plant material e.g., DSC
  • glucoamylase at about 1 to about 1.5 AGU per gram dry solids reduced plant material
  • simultaneous saccharifying and fermenting can be carried out employing quantities of enzyme and yeast selected to maintain low concentrations of glucose in the fermentation broth.
  • the present process can employ quantities of enzyme and yeast selected to maintain glucose at levels at or below about 2 wt-%, at or below about 1 wt-%, at or below about 0.5 wt-%, or at or below about 0.1 wt-%.
  • the present process can employ quantities of enzyme and yeast selected to maintain glucose at levels at or below about 2 wt-% during saccharifying and fermenting.
  • the present process can employ quantities of enzyme and yeast selected to maintain glucose at levels at or below about 2 wt-% from hours 0-10 (or from 0 to about 15% of the time) of saccharifying and fermenting.
  • the present process can employ quantities of enzyme and yeast selected to maintain glucose at levels at or below about 1 wt-%, at or below about 0.5 wt-%, or at or below about 0.1 wt-% from hours 12-54 (or from about 15% to about 80% of the time) of saccharifying and fermenting.
  • the present process can employ quantities of enzyme and yeast selected to maintain glucose at levels at or below about 1 wt-% from hours 54-66 (or about from 80% to about 100% of the time) of saccharifying and fermenting.
  • Suitable levels of enzyme include acid fungal amylase at about 0.3 to about 3 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 3 AGU per gram dry solids reduced plant material (e.g., DSC).
  • simultaneous saccharifying and fermenting can employ acid fungal amylase at about 1 to about 2 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 1.5 AGU per gram dry solids reduced plant material (e.g., DSC).
  • simultaneous saccharifying and fermenting can be carried out employing quantities of enzyme and yeast selected to maintain low concentrations of maltose (DP2) in the fermentation broth.
  • the present process can employ quantities of enzyme and yeast selected to maintain maltose at levels at or below about 0.5 wt-% or at or below about 0.2 wt-%.
  • Suitable levels of enzyme include acid fungal amylase at about 0.3 to about 3 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 3 AGU per gram dry solids reduced plant material (e.g., DSC).
  • simultaneous saccharifying and fermenting can employ acid fungal amylase at about 1 to about 2 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 1.5 AGU per gram dry solids reduced plant material (e.g., DSC).
  • dry solids reduced plant material e.g., DSC
  • glucoamylase at about 1 to about 1.5 AGU per gram dry solids reduced plant material
  • simultaneous saccharifying and fermenting can be carried out employing quantities of enzyme and yeast selected to maintain low concentrations of dextrin in the fermentation broth.
  • the present process can employ quantities of enzyme and yeast selected to maintain maltotriose (DP3) at levels at or below about 0.5 wt-%, at or below about 0.2 wt-%, or at or below about 0.1 wt-%.
  • DP3 maltotriose
  • the present process can employ quantities of enzyme and yeast selected to maintain dextrin with a degree of polymerization of 4 or more (DP4+) at levels at or below about 1 wt-% or at or below about 0.5 wt-%.
  • Suitable levels of enzyme include acid fungal amylase at about 0.3 to about 3 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 3 AGU per gram dry solids reduced plant material (e.g., DSC).
  • simultaneous saccharifying and fermenting can employ acid fungal amylase at about 1 to about 2 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 1.5 AGU per gram dry solids reduced plant material (e.g., DSC).
  • simultaneous saccharifying and fermenting can be carried out employing quantities of enzyme and yeast selected to maintain low concentrations of fusel oils in the fermentation broth.
  • the present process can employ quantities of enzyme and yeast selected to maintain fusel oils at levels at or below about 0.4 to about 0.5 wt-%.
  • Suitable levels of enzyme include acid fungal amylase at about 0.3 to about 3 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 3 AGU per gram dry solids reduced plant material (e.g., DSC).
  • simultaneous saccharifying and fermenting can employ acid fungal amylase at about 1 to about 2 AFAU per gram of dry solids reduced plant material (e.g., DSC) and glucoamylase at about 1 to about 1.5 AGU per gram dry solids reduced plant material (e.g., DSC).
  • dry solids reduced plant material e.g., DSC
  • glucoamylase at about 1 to about 1.5 AGU per gram dry solids reduced plant material
  • the saccharification and/or fermentation mixture can include additional ingredients to increase the effectiveness of the process.
  • the mixture can include added nutrients (e.g., yeast micronutrients), antibiotics, salts, added enzymes, and the like.
  • Nutrients can be derived from stillage or backset added to the liquid.
  • Suitable salts can include zinc or magnesium salts, such as zinc sulfate, magnesium sulfate, and the like.
  • Suitable added enzymes include those added to conventional processes, such as protease, phytase, cellulase, hemicellulase, exo- and endo-glucanase, xylanase, and the like.
  • the product of the fermentation process is referred to herein as “beer”.
  • fermenting corn produces “corn beer”.
  • Ethanol can be recovered from the fermentation mixture, from the beer, by any of a variety of known processes. For example, ethanol can be recovered by distillation.
  • the remaining stillage includes both liquid and solid material.
  • the liquid and solid can be separated by, for example, centrifugation.
  • the recovered liquid, thin stillage can be employed as at least part of the liquid for forming the saccharification and fermentation mixture for subsequent batches or runs.
  • the recovered solids, distiller's dried grain include unfermented grain solids and spent yeast solids. Thin stillage can be concentrated to a syrup, which can be added to the distiller's dried grain and the mixture then dried to form distiller's dried grain plus solubles. Distiller's dried grain and/or distiller's dried grain plus solubles can be sold as animal feed.
  • the present method can include heat treatment of the beer or stillage, e.g., between the beer well and distillation.
  • This heat treatment can convert starches to dextrins and sugars for subsequent fermentation in a process known as burn-out.
  • Such a treatment step can also reduce fouling of distillation trays and evaporator heat exchange surfaces.
  • heat treatment staging can be performed on whole stillage. Following enzymatic treatment of the residual starches, in an embodiment, the resulting dextrins and sugars can be fermented within the main fermentation process as recycled backset or processed in a separate fermentation train to produce ethanol.
  • Large pieces of germ and fiber can ferment the residual starch in the fermenter. After fermentation, the fractions could be removed prior to or after distillation. Removal can be effected with a surface skimmer before to distillation. In an embodiment, screening can be performed on the beer. The screened material can then be separated from the ethanol/water mix by, for example, centrifugation and rotary steam drum drying, which can remove the residual ethanol from the cake. In embodiments in which the larger fiber and germ pieces are removed prior to bulk beer distillation, a separate stripper column for the fiber/germ stream can be utilized. Alternatively, fiber and germ could be removed by screening the whole stillage after distillation.
  • all the components are blended and dried together.
  • the fiber and germ can be removed from the finished product by aspiration and/or size classification.
  • the fiber from the DDGS can be aspirated. Removal of fiber by aspiration after drying increased the amount of oil and protein in the residual DDGS by 0.2 to 1.9% and 0.4 to 1.4%, respectively.
  • the amount of NDF in the residual DDGS decreased by 0.1 to 2.8%.
  • fractionation can employ the larger fiber and germ pieces to increase the particle size of that part of the DDGS derived from the endosperm, as well as to improve syrup carrying capacity.
  • a ring dryer disintegrator can provide some particle size reduction and homogenization.
  • the present process can be run via a batch or continuous process.
  • a continuous process includes moving (pumping) the saccharifying and/or fermenting mixtures through a series of vessels (e.g., tanks) to provide a sufficient duration for the process.
  • vessels e.g., tanks
  • a multiple stage fermentation system can be employed for a continuous process with 48-96 hours residence time.
  • reduced plant material can be fed into the top of a first vessel for saccharifying and fermenting.
  • Partially incubated and fermented mixture can then be drawn out of the bottom of the first vessel and fed in to the top of a second vessel, and so on.
  • the present method is more suitable than conventional methods for running as a continuous process. It is believed that the present process provides reduced opportunity for growth of contaminating organisms in a continuous process.
  • the majority of dry grind ethanol facilities employ batch fermentation technology. This is in part due to the difficulty of preventing losses due to contamination in these conventional processes.
  • the conventional belief is that a separate saccharification stage prior to fermentation is necessary to pre-saccharify the mash for fermentation. Such pre-saccharification insures that there is adequate fermentable glucose for the continuous fermentation process.
  • the present method achieves efficient production of high concentrations of ethanol without a liquefaction or saccharification stage prior to fermentation. This is surprising since this conventional wisdom teaches that it is necessary to have adequate levels of fermentable sugar available during the fermentation process when practiced in a continuous mode. In contrast the present method can provide low concentrations of glucose and efficient fermentation. In the present method, it appears that the glucose is consumed rapidly by the fermenting yeast cell. It is believed that such low glucose levels reduce stress on the yeast, such as stress caused by osmotic inhibition and bacterial contamination pressures. According to the present invention, ethanol levels greater than 18% by volume can be achieved in about to about 96 hours.
  • the present invention also relates to a high alcohol beer.
  • the process of the present invention produces beer containing greater than 18 vol-% ethanol.
  • the present process can produce such a high alcohol beer in about 40 to about 96 hours or about to about 96 hours.
  • the beer includes 18 vol-% to about 23 vol-% ethanol.
  • the present method can produce alcohol contents in the fermenter of 18 to 23% by volume in about 45 to 96 hours.
  • the present method can produce alcohol content in the fermenter of 18 to 23% by volume in about 45 to 96 hours.
  • the majority of the alcohol 80% or more of the final concentration
  • an additional 2 to 5 vol-% alcohol can be produced in the final 12-48 hours.
  • Concentrations of ethanol up to 23 vol-% can be achieved with fermentation time up to 96 hours. It can be economically advantageous to harvest after 48 to 72 hours of fermentation to increase fermenter productivity.
  • the present beer can include this high level of ethanol even when it includes high levels of residual starch.
  • the present beer can include ethanol at 18 to 23 vol-% when it contains 0 to 30% residual starch.
  • the present beer can contain residual starches as low as 0% to as high as 20% residual starch.
  • the present beer includes fewer residual byproducts than conventional beers, even though residual starch can be higher.
  • residual glucose, maltose, and higher dextrins can be as much as 0.8 wt-% lower than in conventional beers produced under similar fermentation conditions.
  • residual glycerol can be as much as 0.45 wt-% less. Lactic acid and fusel oils can also be significantly reduced.
  • the present beer can include less than or equal to about 0.2 wt-% glucose, about 0.4 wt-%, about 0.1 wt-% DP3, undetectable DP4+, 0.45 wt-% glycerol, about 0.01 wt-% lactic acid, and/or about 0.4 wt-% fusel oils.
  • the present invention also relates to a distiller's dried grain product.
  • the distiller's dried grain can also include elevated levels of one or more of protein, fat, fiber (e.g., neutral detergent fiber (NDF)), and starch.
  • NDF neutral detergent fiber
  • the distiller's dried grain can include 34 or more wt-% protein or about 30 to about 45 wt-% protein, or about 1 to about 2 wt-% more protein than produced by the conventional process.
  • the distiller's dried grain can include 15 or more wt-% fat, about 13 to about 17 wt-% fat, or about 1 to about 6 wt-% more fat than produced by the conventional process.
  • the distiller's dried grain can include 31 or more wt-% fiber, about 23 to about 37 wt-% fiber, or about 3 to about 13 wt-% more fiber than produced by the conventional process.
  • the distiller's dried grain can include 12 or more wt-% starch, about 1 to about 23 wt-% starch, or about 1 to about 18 wt-% more starch than produced by the conventional process.
  • the present distiller's dried grain includes elevated levels of B vitamins, vitamin C, vitamin E, folic acid, and/or vitamin A, compared to conventional distiller's dried grain products.
  • the present distiller's dried grain has a richer gold color compared to conventional distiller's dried grain products.
  • the present invention also relates to a distiller's dried grain with one or more improved physical characteristics, such as decreased caking or compaction or increase ability to flow.
  • the present process can produce such an improved distiller's dried grain.
  • the present process can produce fermentation solids including higher molecular weight forms of carbohydrates. Such fermentation solids can, it is believed, exhibit a higher glass transition temperature (i.e. higher T g values). For example, residual starches have a high T g value. Thus, through control of starch content in the DDG and DDGS, the present process can manufacture DDG or DDGS with target T g values.
  • adding an alkaline syrup blend e.g., syrup plus added lime or other alkaline material
  • the fermentation solids e.g., distiller's dried grains
  • DDGS distiller's dried grain with solubles
  • organic acids such as lactic, acetic, and succinic acids which are produced in fermentation have a lower T g value than their corresponding calcium salts.
  • process of the present invention can need not destroy protein in the fermented plant material.
  • Corn contains prolamins, such as zein.
  • Grain sorghum for example, contains a class of zein-like proteins known as kafirins, which resemble zein in amino acid composition.
  • the thermal degradation that occurs during liquefaction, distillation, and high temperature drying produces DDG and DDGS including significant amounts of degraded protein. It is believed that the process of the present invention can provides improved levels of the prolamin fraction of cereal grains.
  • efficient fermentation of the present process removes from the DDG or DDGS non zein components such as starch.
  • Fractionating the plant material, e.g., corn can also increase levels of proteins, such as zein, in the DDG or DDGS.
  • proteins such as zein
  • removing the bran and germ fractions prior to fermentation can concentrate zein in the substrate.
  • Zein in corn is isolated in the endosperm. Fermentation of zein enriched endosperm results in concentration of the zein in the residuals from fermentation.
  • the process of the present invention can provide DDG and DDGS with different, predetermined T g values.
  • the process of the present invention can ferment fractions containing high, medium, or low levels of zein, thus varying the glass transition temperature of the resulting DDG or DDGS.
  • the resulting co-product T g can be directly proportional to the prolamin protein (such as zein) content.
  • the process of the current invention is desirable for the fermentation of high protein corn. This also allows production of DDG and DDGS with a higher prolamin (zein) content.
  • the wet cake fraction produced by the present method which can be dried separately to produce DDG, can be higher in prolamin protein (such as zein) than conventional DDG.
  • the present process allows syrup and wet cake blend ratios to be varied. This results in DDG/DDGS with varying ratios of prolamin protein (such as zein) and residual starch. As the residual starch in the wet cake reduces the protein in the wet cake increases. This indicates an inverse relationship. A similar response occurs in the syrup fraction.
  • starch can segregate into the liquid fraction.
  • the amount of starch in the DDGS can be varied by blending syrup at rates ranging from 0 lbs. dry weight of syrup solids to 1.2 lbs. of syrup solids per lb. of wet cake solids before, and various times during drying to create the final DDGS product.
  • the disproportionate segregation of residual starches into the backset or thin stillage fraction can provide both the aforementioned burn-out and secondary fermentation to be performed on these fractions. Since the thin stillage is evaporated to produce syrup, the centrifuge mass balance also enables DDGS production at various T g values depending on the desired properties and their dependence on T g .
  • the present invention has emissions benefits. Emissions benefits result in the reduction in byproducts created in the ethanol manufacturing process. There is a marked reduction in extraction of fats and oils in the mash from the germ fraction of cereal grains. There is a reduction of byproducts from Maillard reactions typically formed during cooking and liquefaction. And there is a reduction in fermentation byproducts. These observations result in reduced emissions during the recovery of co-products. The concentration and emission rates of volatile organic compounds (VOC), carbon monoxide (CO), nitric oxide compounds (NOx), sulfur oxides (SO2), and other emissions are considerably lower. See Table 1. Note that other manufacturers have attempted to lower emissions by manufacturing wet cake instead of drying to DDG or DDGS.
  • VOC volatile organic compounds
  • CO carbon monoxide
  • NOx nitric oxide compounds
  • SO2 sulfur oxides
  • the present invention also relates to volatile organic compounds (VOC), such as those produced by drying products of a fermentation process.
  • VOC volatile organic compounds
  • the present method includes producing ethanol, distiller's dried grain, and additional useful fermentation products with production of lower levels of VOC compared to conventional processes.
  • drying distillation products e.g., spent grain
  • VOC volatile organic compounds
  • VOC produced by fermentation processes include ethanol, acetic acid, formaldehyde, methanol, acetaldehyde, acrolein, furfural, lactic acid, formic acid, and glycerol.
  • the present invention also relates to carbon monoxide (CO), such as those produced by drying products of a fermentation process.
  • CO carbon monoxide
  • the present method includes producing ethanol, distiller's dried grain, and additional useful fermentation products with production of lower levels of CO compared to conventional processes.
  • drying distillation products e.g., spent grain
  • a method according to the present invention was employed to produce distiller's dried grain from corn. This method produced high protein, high fat, and high fiber distiller's dried grain. Comparison with a conventional saccharification and liquefaction process indicates superior performance of the present method.
  • Yeast inoculum was prepared by adding glucoamylase (0.088 ml of Novozyme's Spirizyme Plus gluco-amylase at 400 AGU/g) and protease (0.018 ml of Genencor International's GC 106 protease 1000 SAPU/g) to 400 ml of stillage containing 70 grams of maltodextrin.
  • Stillage (backset) used was prepared from prior conventional or raw starch fermentations by distilling off the alcohol and subjecting the resulting whole stillage to centrifugal separation to produce backset.
  • Corn was obtained from commercial suppliers of seed corn and was ground through a 0.5 mm screen using a hammermill prior to fermentation. Several varieties of conventional number 2 yellow dent corn were compared, and in several experiments their isogenically equivalent of waxy corn was also tested. Different corn varieties were tested to demonstrate that the present methods produce improved DDG using any of a variety of corn hybrids.
  • Fermentations were conducted for 72 hours with sampling conducted at approximately 24 (e.g. 25) hour intervals. All samples were analyzed by HPLC. At the end of fermentation beer samples were placed in metal pans, pH was decreased to ⁇ 3.5 to inactivate residual enzyme activity, and dried.
  • Waxy corn is almost entirely comprised of amylopectin starch, whereas normal #2 yellow corn is about 25 to 28% amylose starch with the remainder being amylopectin.
  • Waxy corn is generally not used in the conventional process because of the high peak viscosity and more rapid rate of viscosity development compared to regular corn. The high initial viscosity makes the corn slurry more difficult to pump during the initial primary high temperature liquefaction. Waxy corn varieties can, however, be readily employed in the present process. Because no cook stage is employed, the high peak viscosity is not a processing issue.
  • the Present Process Provides Improved Yield Potential
  • the yield potential of the method of the present invention was compared to a conventional process.
  • the present method exhibited improved yield using temperature staging.
  • the present method exhibited an increased potential maximum yield for ethanol production. Comparison with conventional saccharification and liquefaction process indicates superior performance of the present method.
  • Fermentations were prepared in a similar manner as in Example 1 except for intentional differences in particle size, alpha amylase enzyme dose, gluco-amylase enzyme dose, or acid fungal amylase enzyme dose. Conditions for this experiment are described in Table 3. Corn for all tests was obtained from Broin Enterprises (BEI), Scotland, S. Dak., USA. Corn representing a coarse particle size by raw starch standards was ground at BEI. Finely ground corn was produced using a lab hammermill through a 0.5 mm screen.
  • the conventional process utilized indicated levels of Liquozyme SC and GC 480.
  • the raw starch process used indicated levels of Spirizyme Plus and SP 288 acid fungal amylase at 1700 AFAU's per gm. Dosages of urea liquor, zinc sulfate, and antibiotic were adjusted accordingly for the conventional process.
  • Stillage (backset) used was prepared from prior conventional or raw starch fermentations by distilling off the alcohol and subjecting the resulting whole stillage to centrifugal separation to produce backset. Fermentation temperatures were staged according to the following set points: 0-18 hours at 90° F., 18-42 hours at 86° F., and 42-72 hours at 82° F. Samples were taken at 65 hours to represent the end of fermentation.
  • FIG. 1C illustrates the potential for the new process to produce more alcohol than the conventional process, even when residual starch levels are higher.
  • Conventional wisdom would suggest the raw starch process is less efficient due to the higher levels of residual starch, however, this is not the case.
  • the present process is superior to the conventional method. Note that fermentation efficiency can also be assessed by examining the fermentation drop solids. This is shown in the composite data comparing both processes in FIG. 1D. Since all fermentations in the above example were started at the same initial set solids, a lower drop solids suggests a more efficient conversion of starch to ethanol. The potential of this process is also indicated by the achievement of an equal to or reduced level of drop solids, despite the higher residual starches observed.
  • FIG. 1E shows the temperature staging done during the present process. Fermentation temperatures were staged according to the following set points: 0-18 hours at approximately 90° F. (ranging from about 95° F. to about 90° F.), 18-42 hours at approximately 86° F. (ranging from 90° F. to 86° F.), and 42-72 hours at about 82° F. (ranging from 86° F. to 84° F.). Staging of temperature helps to increase ethanol production process by reducing stress on yeast. The temperature is decreased as ethanol is produced to reduce the stress on yeast caused by ethanol production. TABLE 3 Comparison of the Yield Potential of Conventional vs.
  • the Present Process Exhibits Improved Results with Increased Levels of Acid Fungal Amylase and Increased Levels of Glucoamylase
  • Results of an embodiment of the method of the present invention were evaluated with increased levels of acid fungal amylase and increased levels of glucoamylase. Increased levels of acid fungal amylase improved results with the present process. The increased levels of glucoamylase improved results with the present process.
  • Glucoamylase Novozymes Spirizyme Plus
  • acid fungal amylase Novozymes SP 288
  • the objective of this test was to examine the effect of a range of dosages of glucoamylase and acid fungal amylase on production of ethanol and other products from raw starch hydrolysis fermentations.
  • dosages above 0.3 AFAU's per gm dry solids corn for acid fungal amylase and dosages above 0.3 AGU's per gram of dry solids corn produce higher alcohol and consistently higher residual glucose. The consistently higher glucose indicates that these fermentations have the potential for even higher ethanol yield.
  • Results of an embodiment of the method of the present invention were evaluated with varying particle size of the ground plant material. Smaller particle sizes improved results with the present process.
  • Results are shown in Table 4, and FIGS. 3A, 3B, 3 C, 3 D.
  • the data illustrates that smaller grind size provided higher ethanol yield and lower residual starch.
  • grind size was a more influential factor.
  • a higher enzyme dosage was required to achieve the same relative results.
  • Results of an embodiment of the method of the present invention were evaluated with varying particle size of the ground plant material, varying glucoamylase type, and dosage of acid fungal amylase.
  • FIGS. 4A, 4B, and 4 C Final fermenter results are shown in FIGS. 4A, 4B, and 4 C.
  • Conventional glucoamylase enzymes such as Distillase from Genencor International contained a very low level of acid fungal amylase activity.
  • Spirizyme Plus contained about 2.5 times as much AFAU activity per ml of enzyme and exhibited improved performance for hydrolyzing raw starch.
  • SP 288 acid fungal amylase contained a relatively low level of glucoamylase.
  • An embodiment of the present invention was employed to produce ethanol from corn. This process produced high alcohol corn beer, high protein, high fat, and high fiber distiller's dried grain. Comparison with conventional saccharification and liquefaction process indicate superior performance of the present method.
  • Example 6 was set up in a manner similar to prior examples except the initial fermentation solids and temperature were varied as described in the presentation of the results.
  • An interesting feature of the present raw starch fermentation process is the ability to enhance the rate of fermentation through increasing the solids content or initial temperature of fermentation. Solids loading, temperature, grind size, glucoamylase dosage, acid fungal amylase dosage, and yeast dosage can be combined to increase the performance of raw starch fermentation.
  • FIGS. 5A, 5B, 5 C, 5 D, 5 E, 5 F, 5 G, 5 H, 5 I, and 5 J illustrate the influence of temperature at different solids loadings.
  • the residual starch values reported for this Example suggest that temperature can be used to improve the efficiency of raw starch fermentations at intermediate fermentation gravities, which are defined as fermentation solids levels which would yield between 15% to 18% ethanol.
  • the fermentation temperature could be used to accelerate raw starch fermentations so that they finish in less than 48 hours, yet still achieve alcohol levels of 15% to 18%, with acceptable residual starch levels.
  • the increased fermentation set point will help to accelerate enzymatic conversion of native starch to glucose, which appears to be the rate limiting step in the raw starch process.
  • Fermentation performance using higher temperature set points is an aspect of the process for intermediate ethanol ranges, especially when viewed from the perspective of prior examples establishing that raw starch fermentations can tolerate a higher level of residual starch in the residual distillers dried grains and with distillers dried grains solubles, and still produce excellent quality DDG or DDGS according to the proximate analysis.
  • the dry substance of raw starch fermentations can be increased by approximately 20% to increase the rate of fermentation, while producing higher alcohol content in the fermenter and more DDGS with excellent quality even if the residual starch levels are high.
  • the present method produced ethanol containing corn beer with greater than 18 vol-% ethanol. Runs produced at least 18 vol-% ethanol and up to 23 vol-% ethanol within 48 to 96 hours of incubation and fermentation. The beer contained these high levels of ethanol even when it also included higher levels of residual starch. After 24 hours of incubating and fermenting the corn beer contained 9-16.5 or 12-15 vol-% ethanol. After 48 hours of incubating and fermenting the corn beer contained 13-20 vol-% ethanol. Ethanol production was linear up to a level of 14-16 vol-%. A compilation of ethanol production results from various runs is illustrated at least in FIGS. 6A and 6B.
  • the beer contained approximately 0.4 to 0.5 wt-% less glycerol than conventional fermentation at otherwise identical fermentation conditions (FIG. 7).
  • the beer contained less extracted oil from the germ fraction, resulting in reduced fouling and lower VOC emissions in the water vapor during drying of the residual animal feed product.
  • Table 1 The beer contained less extracted oil from the germ fraction, resulting in reduced fouling and lower CO emissions in the water vapor during drying of the residual animal feed product (Table 1).
  • the beer contained less fusel oil (FIG. 8), which inhibits yeast cell growth and fermentation if these alcohol compounds are unintentionally recycled in distillation side stripper bottoms streams. Fusel oils are also an undesirable component of potable alcohol manufacturing operations, so the present process offers an improved method of production of potable alcohol.
  • the beer also contained less lactic and acetic acid relative to the conventional process.
  • the beer also contained higher yeast cell counts, which contributes to improved feed products.
  • the present process maintained yeast at or above 300 cells/mL in these numerous runs. Yeast budding was observed in at least 40% of the yeast from hours 0-20 of incubating and fermenting and/or at least 15-20% of the yeast after hours 60 of incubating and fermenting. These yeast counts and budding are higher than observed in the conventional process.
  • the Present Process Maintains Low Levels of Glucose, Maltose (DP2), Maltotriose (DP3), and Dextrins (DP4+)
  • the levels of glucose, maltose (DP2), maltotriose (DP3), and dextrins (DP4+) produced by an embodiment of the present invention was compared to a conventional process.
  • the present method exhibited decreased levels of glucose, maltose (DP2), maltotriose (DP3), and dextrins (DP4+) respectively. Comparison of the level of glucose to the conventional process indicates superior performance of the present method.
  • Batch fermentations were set up in a similar manner as Example 1.
  • Continuous ethanol fermentation was evaluated in a bench top system consisting of a refrigerated cold slurry tank followed by five (5) fermenters operating in continuous mode and finishing with a beerwell collecting the fermented beer.
  • the volume of each fermentation stage was approximately two (2) liters.
  • Average fermenter fill solids was approximately 30-35% dry solids corn, depending on the starch content of the substrate.
  • the mash slurry for feeding fermentation was prepared every 3 to 4 days and maintained between 6 to 12 degrees Centigrade to discourage bacterial growth in the feed tank.
  • the mash preparation procedures did not sterilize the mash prior to fermentation, and the fermentation train was operated with no antibiotic addition to inhibit bacterial contaminants.
  • the mash was stored at a cold temperature to reduce the amount of work required for substrate preparation. 15 to 20 ml of 50% urea liquor was added to the cold slurry tank, which had a final mash volume of approximately 9000 liters.
  • Each fermenter in the continuous series was fed from the prior fermenter, while the first fermenter was fed directly from the cold slurry tank. Fermentation temperature was held at a constant 82° F. through the five (5) stage fermentation. Glucoamylase was dosed into the first fermenter to provide a dosage of approximately 2.0 to 2.4 AGU's per gram dry substance corn. Fali yeast, obtained from Fleischmann's Yeast, was added at a rate of approximately 0.65 grams per liter of slurry makeup, and was batched into the cold slurry each time fresh mash was prepared.
  • a continuous fermentation run was set up employing the procedure described above for experiment 1. Lactic acid and acetic acid measurements were taken at various times and stages during the continuous multistage fermentation process. Toward the end of the run, the initial slurry pH was purposely increased, as shown, to challenge the system microbiologically. In certain circumstances, slurry pH was intermittently lowered to keep contamination in check (see, e.g., FIGS. 16A, 16B, and 16 C).
  • FIGS. 9A and 9B illustrate that the present process maintained low levels of glucose during simultaneous saccharification and fermentation (SSF) and continuous raw starch fermentations.
  • SSF simultaneous saccharification and fermentation
  • this low level of glucose reduces potential reactions such as reversion, condensation, or Maillard Browning Reactions. Such reactions in turn can reduce ethanol yield.
  • the data compiled in this example demonstrates that the process maintained glucose at levels at or below 3 wt-% for the entire run and at or below 1 wt-% for about 90% of the run. In particular, the process maintained glucose at levels at or below 1 wt-% from hours 12-54 of incubating and fermenting.
  • FIGS. 10-12 illustrate that the present process maintained low levels of dextrin during SSF and continuous raw starch fermentation.
  • FIGS. 10A and 10B illustrate that the present process maintained maltose (DP2) at levels at or below about 0.2 wt-% during simultaneous saccharifying and fermenting and below about 0.34 wt-% during continuous raw starch fermentation.
  • the data shown in FIG. 11A demonstrate that the process maintained low levels of maltotriose (DP3) during simultaneous saccharifying and fermenting at levels at or below 0.2 wt-% and at or below 0.1 wt-%.
  • the data shown in FIG. 11B demonstrate that the present process maintained low levels of maltotriose (DP3) during a continuous raw starch fermentation at levels at or below 0.25 wt-%.
  • the data shown in FIG. 12A demonstrate that the process maintained low levels of dextrins (DP4+) during simultaneous saccharifying and fermenting at levels at or below 1 wt-% and at or below 0.5 wt-%.
  • the data shown in FIG. 12B demonstrate that the process maintained low levels of dextrins (DP4+) during continuous raw starch system at levels at or below 0.3 wt-%.
  • the results of experiment 3 demonstrate that a continuous embodiment of the present method produced residual starch levels lower than that produced by the conventional process (FIG. 17).
  • the residual starch levels produced using this embodiment of the present method remained lower than the residual starch levels of the conventional process (FIG. 17).
  • the percentage of starch produced using this embodiment of the present method remained at about twenty (e.g. 21) or less (FIG. 17) whereas the percentage starch produced using the conventional process was as high as 27 (FIG. 17).
  • temperature staging can be employed to minimize residual glucose production. That is, the temperature of the fermentation can be reduced as the fermentation progresses. It is believed that, in general, for every 10° C. (18° F.) increase in temperature, the rate of an enzymatic reaction approximately doubles.
  • enzyme action can be slowed by decreasing the temperature of the fermentation mixture after a time period, such as after 30 hours. It is believed that cooling also maintains yeast viability, so that fermentation can continue to utilize the glucose that has been formed.
  • the present process does not require a saccharification stage before fermentation and produces improved results.
  • Another conventional continuous process includes aerating the 1 st fermentor, and possibly the second fermentor, to encourage yeast growth.
  • the present process provides improved results and does not require aeration of the fermentor.
  • Some conventional continuous systems employ a yeast recycle method.
  • the present method does not require yeast recycling and provides improved results.
  • This embodiment of the present invention is superior to such conventional continuous fermentation systems.
  • the present invention can employ simultaneous saccharification and fermentation of raw starch and can operate at high gravity. In an embodiment, the process of the present invention can produce ethanol at fast rates despite the apparent lack of adequate fermentable substrate.
  • a continuous ethanol production embodiment of the present method maintained low acidity levels throughout the fermentation cycle.
  • a continuous ethanol production embodiment of the present method maintained lower starch levels throughout the fermentation cycle. Comparison of the present residual starch level to the conventional process provides an indication of advantageous performance from the present method. The mass balance of the present raw starch process suggests that residual starches can actually be higher in this process relative to the conventional, while still achieving a higher ethanol yield and improved proximate mass balance.
  • the DDGS according to an embodiment of the present invention was compared to that produced by a conventional process.
  • the present method produced an inventive DDGS that exhibited less caking compared to DDGS produced by the conventional process.
  • the present DDGS with less caking is superior to conventional DDGS.
  • the DDGS was collected as a co-product of ethanol production from the conventional high temperature liquefaction process and from the process of the present invention.
  • the caking/collapse assay was performed by filling a 500 ml cylinder with approximately 400 ml of DDGS. Attention was given to avoiding physical packing of the DDGS when filling the cylinder. After filling, a 4.4 cm diameter disc weighing 78 grams was placed on top of the DDGS, followed by placement of 1.5 kg of lead shot (in an appropriately sized plastic bag) on top of the disc. Assay preparation was completed by covering each cylinder with a plastic bag and sealing the apparatus with a rubber band to prevent moisture loss.
  • the weight applied to the DDGS is used to exaggerate the effect and approximate the conditions which DDGS is exposed to during transport, for example, in a railcar.
  • the level of the DDGS is noted at the beginning of storage and at various times during storage at a temperature of 50° C.
  • the measured height of the collapsed (caked) DDGS was compared to the initial height of the DDGS.
  • the measured height was compared to the initial height as an estimate of the tendency of the product to collapse or cake.
  • the DDGS from the present invention shows less caking collapse over time (FIG. 13) when compared to the DDGS of the conventional process. Over a twenty-five hour compaction time the DDGS according to the present invention collapsed only 4-5% of the initial volume as compared to 10-14% of the volume collapse for DDGS of the conventional process.
  • DDGS compaction of DDGS at controlled conditions models the DDGS caking observed in the containers of transportation vehicles, for example railcars and trucks.
  • DDGS produced using the process of this invention exhibited less caking related collapse than that of the conventional process, indicating superior performance of the present method.
  • glass transition temperature increases with molecular weight for polymers such as those found in DDGS.
  • the present DDG includes higher levels of such polymers and should exhibit a higher glass transition temperature. It is believed that product moisture, storage temperature, and chemical composition can impact the transition of DDGS from an amorphous glass to an amorphous rubber phase. DDGS in the rubber phase compacts more readily that DDGS in the glass phase.
  • the Present Process can Employ High Protein Corn to Produce High Protein DDGS and High Levels of Ethanol
  • the present invention can include fermenting high protein corn to produce high protein DDGS and high levels of ethanol. This provides for advantageous flexibility for processing high protein corn.
  • DDGS was collected as a co-product of ethanol production from fermentation of various corn hybrids with fermentations set up in a similar manner as Example 1. All fermentations were set up using identical conditions. Different corn hybrids were tested using various grind sizes using a lab scale hammermill. The hammermill screen size was varied from 0.5 mm to 4.0 mm to create flour particle sizes ranging from fine (0.5 mm screen) to coarse (4.0 mm screen).
  • FIG. 15A illustrates the dependence of protein level in DDGS on grind size. This figure illustrates the inverse correlation between grind size and protein: as particle size increases the protein content of DDGS decreases for each tested corn hybrid (FIG. 15A).
  • FIG. 15B illustrates the dependence of starch level in DDGS on grind size. This figure illustrates a positive correlation between grind size and starch content in: as particle size increases the starch content of the DDGS increases for each tested corn hybrid (FIG. 15B).
  • FIG. 15C illustrates the dependence of ethanol production on grind size. This figure illustrates that as particle size decreases there is an increase in ethanol production (FIG. 15C).
  • “highly fermentable corn” hybrids can have a higher starch concentration and not a high protein concentration.
  • This example demonstrates that higher protein corn hybrid varieties of standard #2 yellow corn can be used to obtain high levels of ethanol production.
  • fermenter dry solids can be increased to maintain ethanol % levels in the fermenter while producing a higher protein DDGS.
  • the present invention provides improved access to the prolamin protein (such as zein) fraction of cereal grains.
  • the high protein content of DDG and DDGS is useful in compounding.
  • FIGS. 14A and 14B show the relationship of: wet cake, syrup starch, and protein levels. As the residual starch in the wet cake reduces the protein in the wet cake increases. This indicates an inverse relationship. A similar response occurs in the syrup fraction.

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US11/077,969 US20050233030A1 (en) 2004-03-10 2005-03-10 Methods and systems for producing ethanol using raw starch and fractionation
US11/077,971 US20050239181A1 (en) 2004-03-10 2005-03-10 Continuous process for producing ethanol using raw starch
US11/676,965 US7842484B2 (en) 2003-03-10 2007-02-20 Method for producing ethanol using raw starch
US11/682,195 US8409640B2 (en) 2003-03-10 2007-03-05 Methods and systems for producing ethanol using raw starch and fractionation
US12/405,125 US7919291B2 (en) 2003-03-10 2009-03-16 Method for producing ethanol using raw starch
US12/886,483 US8470550B2 (en) 2003-03-10 2010-09-20 Composition comprising raw starch for the production of ethanol
US12/944,618 US8409639B2 (en) 2003-03-10 2010-11-11 Methods and systems for producing ethanol using raw starch and fractionation
US12/944,606 US8497082B2 (en) 2003-03-10 2010-11-11 Composition comprising corn flour and saccharification enzymes
US13/850,196 US8748141B2 (en) 2003-03-10 2013-03-25 Methods and systems for producing ethanol using raw starch and fractionation
US13/866,002 US8679793B2 (en) 2003-03-10 2013-04-18 Method for producing ethanol using raw starch
US14/285,030 US20140283226A1 (en) 2003-03-10 2014-05-22 Methods and systems for producing ethanol using raw starch and fractionation
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US8679793B2 (en) 2014-03-25
US8497082B2 (en) 2013-07-30
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US7842484B2 (en) 2010-11-30
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