AU2003300108A1 - Vegetable tenderizer - Google Patents

Vegetable tenderizer Download PDF

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Publication number
AU2003300108A1
AU2003300108A1 AU2003300108A AU2003300108A AU2003300108A1 AU 2003300108 A1 AU2003300108 A1 AU 2003300108A1 AU 2003300108 A AU2003300108 A AU 2003300108A AU 2003300108 A AU2003300108 A AU 2003300108A AU 2003300108 A1 AU2003300108 A1 AU 2003300108A1
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composition
enzyme
raw vegetable
raw
vegetable composition
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AU2003300108A
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AU2003300108B2 (en
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Uchenna N. Chukwu
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CHI'S RESEARCH Corp
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CHI S RES CORP
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Priority claimed from US10/619,403 external-priority patent/US7407678B2/en
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Priority to AU2006200153A priority Critical patent/AU2006200153B2/en
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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
    • A23B7/00—Preservation or chemical ripening of fruit or vegetables
    • A23B7/14—Preserving or ripening with chemicals not covered by groups A23B7/08 or A23B7/10
    • A23B7/153—Preserving or ripening with chemicals not covered by groups A23B7/08 or A23B7/10 in the form of liquids or solids
    • A23B7/154—Organic compounds; Microorganisms; Enzymes
    • A23B7/155—Microorganisms; Enzymes; Antibiotics
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
    • A23B7/00—Preservation or chemical ripening of fruit or vegetables
    • A23B7/02—Dehydrating; Subsequent reconstitution
    • A23B7/022—Dehydrating; Subsequent reconstitution with addition of chemicals before or during drying, e.g. semi-moist products
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
    • A23B7/00—Preservation or chemical ripening of fruit or vegetables
    • A23B7/04—Freezing; Subsequent thawing; Cooling
    • A23B7/0441—Treatment other than blanching preparatory to freezing
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
    • A23B7/00—Preservation or chemical ripening of fruit or vegetables
    • A23B7/06—Blanching
    • 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
    • A23L11/00—Pulses, i.e. fruits of leguminous plants, for production of food; Products from legumes; Preparation or treatment thereof
    • A23L11/10—Rapid cooking pulses
    • 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
    • A23L11/00—Pulses, i.e. fruits of leguminous plants, for production of food; Products from legumes; Preparation or treatment thereof
    • A23L11/30—Removing undesirable substances, e.g. bitter substances
    • A23L11/33—Removing undesirable substances, e.g. bitter substances using enzymes; Enzymatic transformation of pulses or legumes
    • 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
    • A23L19/00—Products from fruits or vegetables; Preparation or treatment thereof
    • A23L19/03—Products from fruits or vegetables; Preparation or treatment thereof consisting of whole pieces or fragments without mashing the original pieces
    • 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
    • A23L19/00—Products from fruits or vegetables; Preparation or treatment thereof
    • A23L19/20—Products from fruits or vegetables; Preparation or treatment thereof by pickling, e.g. sauerkraut or pickles
    • 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
    • A23L25/00—Food consisting mainly of nutmeat or seeds; Preparation or treatment thereof
    • A23L25/40—Fermented products; Products treated with microorganisms or enzymes
    • 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
    • A23L7/107—Addition or treatment with enzymes not combined with fermentation with 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
    • A23L7/10—Cereal-derived products
    • A23L7/161—Puffed cereals, e.g. popcorn or puffed rice
    • A23L7/174—Preparation of puffed cereals from wholegrain or grain pieces without preparation of meal or dough
    • A23L7/183—Preparation of puffed cereals from wholegrain or grain pieces without preparation of meal or dough by heating without using a pressure release device
    • 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
    • A23L7/196—Products in which the original granular shape is maintained, e.g. parboiled rice

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  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Food Science & Technology (AREA)
  • Polymers & Plastics (AREA)
  • Health & Medical Sciences (AREA)
  • Nutrition Science (AREA)
  • Microbiology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Agronomy & Crop Science (AREA)
  • Botany (AREA)
  • Biotechnology (AREA)
  • Preparation Of Fruits And Vegetables (AREA)

Description

WO 2004/060072 PCT/US2003/041646 VEGETABLE TENDERIZER BACKGROUND OF THE INVENTION The present invention generally relates to using enzymes to degrade and tenderize raw vegetables prior to implementation of conventional processing 5 techniques. More specifically, the present invention relates to forming a enzyme degraded vegetable product with improved processing and nutritional characteristics and to a method of making the vegetable product. During the last several years, consumer interest in eating foods that are nutritionally balanced with an adequate source of protein, fat, carbohydrates, 10 fiber, vitamins and minerals has increased. Growing concern over chronic diseases, such as cancer, diabetes and heart disease have motivated consumers to seek foods for consumption that are effective in treating chronic diseases while promoting a healthier lifestyle. Such foods may include vegetables that containphytochemicals. Unfortunately, consumption of vegetables having phytochemicals 15 may pose several problems. The presence of anti-nutritional components such as indigestible sugars, enzyme inhibitors, nutrient-binding substances or toxic compounds typically render a vegetable containing the beneficial phytochemicals unfit for consumption. Low concentrations of the desired phytochemical in the vegetable is another problem for consumers, especially if the concentration of the 20 phytochemical is considered too low to deliver a health benefit. Heat and/or pressure processing of vegetables to eliminate anti nutritional components in the vegetable prior to consumption is the traditional approach used by food manufacturers. However, heat and/or pressure processing may eliminate most, if not all phytochemical levels during the manufacturing 25 process. In addition, the manufacturing process may require physical and/or chemical pre-treatment strategies, such as cooking, boiling, application of strong acids, and/or hydration of the raw vegetable prior to processing, in order to adequately process the vegetable. Unfortunately, physical and/or chemical pretreatment strategies ofthe vegetable prior to processing may include complicated 30 steps that increase the overall costs associated with vegetable production.
WO 2004/060072 PCT/US2003/041646 2 BRIEF SUMMARY OF THE INVENTION The present invention includes a method of processing vegetables prior to human consumption by applying enzyme(s) to a raw vegetable for a time that is sufficient to form an enzyme-degraded vegetable under normal atmospheric 5 pressures followed by deactivation of the enzyme(s). DETAILED DESCRIPTION The present invention includes a method of processing vegetables. In the method, an aqueous enzyme composition is applied to a raw vegetable composition under normal atmospheric pressures for a time that is effective to form 10 an enzyme-degraded vegetable composition. After degrading, the enzyme-degraded vegetable composition can be processed by one or more additional processing steps that transforms the enzyme-degraded into a vegetable product destined for human consumption. The present invention preferably uses enzymes that degrade, hydrolyze and/or tenderize the raw vegetable composition. 15 Traditional vegetable processing techniques often require the use of high temperatures and/or high pressure during the manufacturing process due in part to the presence of a tough outer layer on vegetables that functions as a barrier. Such high temperatures and/or pressures increase the cost and complexity of processing vegetables. In addition, such high temperatures and/or high pressures 20 may ultimately reduce the nutritional quality of processed vegetables by lowering phytochemical levels in a manner that reduces consumer acceptability and consumption. The present invention includes enzymatic degradation of raw vegetables under normal atmospheric pressures prior to (1) human consumption or 25 (2) the use of more traditional processing techniques that may require high pressures and/or temperatures to complete production. Additionally, the present invention represents a novel approach that helps to reduce the need for high temperatures and/or pressures during vegetable processing. In addition, since WO 2004/060072 PCT/US2003/041646 3 enzymatic processing of vegetables in accordance with the present invention typically occurs under normal atmospheric pressures, specialized equipment is typically not required and subsequent reduction in the cost and complexity of manufacturing vegetables maybe realized. Furthermore, enzymatic degradation of 5 vegetables prior to using more traditional processing techniques may also permit a reduction in time, energy and/or other resources that are required to complete processing of raw vegetables. While not wanting to be bound to theory, it is believed that when one or more enzyme(s) that are capable of degrading one or more target substrates 10 in a first outer layer of a raw vegetable composition, are applied to the first outer layer of the raw vegetable composition in accordance with the present invention, the enzyme(s) degrade the target substrates of the first outer layer of the raw vegetable composition to form an enzyme-degraded vegetable composition having a compromised first outer layer. In addition, the use of the aqueous enzyme 15 composition to degrade the raw vegetable composition tenderizes the raw vegetable composition which can permit a reduction in cook time and may also permit subsequent in situ modification of the raw vegetable composition by addition of ingredients like vitamins, minerals, or other enzymes that catalyze specific reactions within the raw vegetable composition. 20 As used herein, the term "enzyme" means any complex protein produced by a living cell that is capable of at least catalyzing a specific biochemical reaction on one or more target substrates. The term "enzyme" is also meant to encompass any complex protein capable of catalyzing a specific biochemical reaction that is substantially free of any microorganism. All references to enzyme 25 is also understood as encompassing any synthetically- or genetically-produced identical copy of the enzyme that is identical in molecular structure to the enzyme that originated in a living organism.
WO 2004/060072 PCT/US2003/041646 4 The enzyme(s) that may be included as part of the aqueous enzyme composition may be generally characterized as lipase(s). As used herein, the term "lipase" means any enzyme that is capable of at least catalyzing hydrolysis of a fat containing or lipid-containing target substrate. By"hydrolysis" is meant enzymatic 5 degradation of the fat-containing or lipid-containing target substrate that includes triglycerides, diglycerides, monoglycerides, cutin, wax-containing substrates, chemically combined fatty acids and long chain alcohols, phosphatides, cerebrosides, sterols, terpenes, fatty alcohols, solid fat, liquid fat (oils), fatty acids, fat-soluble vitamins, waxes or any combination of any of these. In addition, the 10 terms "fat-containing" and "lipid-containing" are used interchangeably throughout the specification. Furthermore, the term "hydrolysis" is not meant to include the use of microorganisms that produce carbohydrases to hydrolyze and/or degrade raw vegetable compositions in accordance with the present invention. The application 15 of microorganisms that produces carbohydrases and other enzymes to process raw vegetable compositions is commonly referred to as a microbial fermentation. Additionally, although microbial fermentation may involve some degree of hydrolysis, microbial fermentation is known to further transform sugar components like pentoses or hexoses into organic acids that increases the acidity, 20 reduces the pH, and alters the texture and taste of the fermented vegetable composition. In contrast, the present invention uses enzymes that are substantially free of microorganisms to hydrolyze, tenderize, and/or degrade the raw vegetable composition. Furthermore, use of the aqueous enzyme composition in accordance with the present invention typically results in a decrease in the acidity of, and/or 25 increase in the pH of the aqueous enzyme composition after degradation. Lipases generally hydrolyze or degrade fat-containing or lipid containing molecules into free fatty acids, glycerol, mono- and di-glyecerides. As an example, suitable lipases for the present invention include lipases that can WO 2004/060072 PCT/US2003/041646 5 hydrolyze short, medium or long-chain fatty acids from the 1, 2 or 3 position of triglycerides. In addition, any enzyme that is effective in hydrolyzing wax containing substrates or combinations of long chain alcohols and fatty acids to form long chain alcohols and free fatty acids is also suitable for use in the present 5 invention. Preferably, a lipase that is effective in hydrolyzing at least one long chain fatty acid from a fat-containing or lipid-containing target substrate like a triglyceride is used to practice the present invention. More preferably, a lipase that hydrolyzes at least one long chain fatty acid and/or at least one medium chain fatty 10 acid from a fat-containing or lipid containing target substrate, such as a triglyceride is used in accordance with the present invention. Still more preferably, a lipase that is effective in hydrolyzing a waxy coat or lipid-containing coat of a raw vegetable compositions and is substantially free of any microorganism is included as part of the aqueous enzyme composition. 15 Lipase may be derived from a number of different sources, such as fungal sources, plant sources, microbial sources, animal sources, or any combination of any of these. As examples, Lipase A "Amano" 12, Lipase AY "Amano" 30, Lipase G "Amano" 50, Lipase M "Amano" 10 that are available from Amano Enzyme Co. Ltd. of Lombard, Illinois may be used to degrade the waxy coat of raw vegetable 20 compositions when practicing the present invention. Another enzyme that maybe included as part ofthe aqueous enzyme composition maybe generally characterized as carbohydrase(s). As used herein, the term "carbohydrase" means any enzyme that is capable of at least catalyzing hydrolysis of a carbohydrate-containing target substrate. By "hydrolysis" is meant 25 enzymatic degradation ofthe carbohydrate-containing target substrate that includes complex carbohydrates like cellulose, hemicellulose, pectin, xylan chains of hemicellulose, and/or polymers of other 5-carbon sugars into their sugar components like pentoses or hexoses.
WO 2004/060072 PCT/US2003/041646 6 Preferably, cellulase is one carbohydrase that is also used as part of the aqueous enzyme composition. Still more preferably, cellulase that is substantially free of any microorganism is included as part of the aqueous enzyme composition. Most preferably, cellulase that is substantially free of any 5 microorganism is used to degrade, hydrolyze and/or tenderize the raw vegetable composition when practicing the present invention. Cellulase may be derived from a number of different sources, such as fungal sources, plant sources, microbial sources, animal sources, or any combination of any of these. Besides cellulase, it is believed that other carbohydrases, such as 10 hemicellulase, alpha-galactosidase, invertase, mannanase, beta-gluconase, beta glucanase, arabanase, polygalacturonase, ferulic acid esterase, xylanase, beta galactosidase, beta-fructofuranosidase, alpha-amylase, beta-amylase, pectinase, pectin depolymerase, pectin methyl esterase, pectin lyase, glucoamylase, oligo- 1,6 glucosidase, lactase, beta-d-glucosidase, or any combination of any of these are 15 suitable additional non-exhaustive examples of carbohydrases that may be used separately or in combination with cellulase in accordance with the present invention. Preferably, the aqueous enzyme composition includes lipase, cellulase and any combination of hemicellulase, alpha-galactosidase, mannanase, 20 beta-gluconase, beta-glucanase, arabanase, polygalacturonase, xylanase, beta galactosidase, beta-fructofuranosidase, alpha-amylase, beta-amylase, pectinase, invertase, pectin depolymerase, pectin methyl esterase, pectin lyase, glucoamylase, oligo-1,6 glucosidase, lactase, or beta-d-glucosidase to degrade the raw vegetable composition under normal atmospheric pressures, prior to (1) human consumption 25 or (2) application of traditional processing techniques like cooking, pressure cooking or the like. More preferably, a blend of lipase, cellulase and hemicellulase is used in the present invention to degrade, tenderize and/or render the raw vegetable WO 2004/060072 PCT/US2003/041646 7 composition more absorbent to water, enzymes, additives or the like. Still more preferably, a blend of lipase, cellulase, hemicellulase and pectinase is used in the present invention to degrade the raw vegetable composition so that subsequent processing can be practiced with reduced temperature and/or pressure requirements. 5 A preferred example of a carbohydrase that may be used as part of the aqueous enzyme composition is Viscozyme*L, available from Novozymes of Franklinton, North Carolina. Alternate examples of carbohydrases that are suitable for use as part of the aqueous enzyme composition is Econase* CE, available from Enzyme Development Corporation of New York, New York, Cellulase 4000 or 10 Crystalzyme Cran that is available from Valley Research Inc., of South Bend, Indiana. When enzymes are used during vegetable processing, enzymes may be applied in any form, such as a granular form, or a vapor form, or as part of the aqueous enzyme composition as noted above. The application form that is selected 15 preferably permits the enzyme to (1) contact the vegetable composition being treated, and (2) remain in contact with the vegetable composition being treated for a time that is sufficient to degrade the target substrate. Preferably, the enzyme(s) is applied to the raw vegetable composition as part of the aqueous enzyme composition. 20 The aqueous enzyme composition may include one or more enzyme componentss, one or more optional catalyst component(s), one or more optional pH-modifying component(s), one or more optional additive(s) or one or more optional solvent component(s). The components of the aqueous enzyme composition may be supplied as individual components, or supplied in various 25 prepared mixtures of two or more components, that are subsequently combined to form the aqueous enzyme composition. The enzyme component(s) may include only the enzyme(s), the enzyme(s) and water, or may optionally include additional components. The WO 2004/060072 PCT/US2003/041646 8 enzyme component(s) may be supplied as individual components, or supplied in various prepared mixtures of two or more components, that are subsequently combined to form the enzyme component(s). Additionally, the enzyme component may be supplied in granular form, vapor form, or as part of an aqueous enzyme 5 component. The concentration of the enzyme(s) in the enzyme component may generally range from about 0.0001 weight percent to about 100 weight percent, based on the total weight of the enzyme component. The enzyme component may optionally include sucrose, fructose, ash, alcohol, and any other components that 10 are compatible with, and do not interfere with the biochemical rate of catalysis of the enzyme. Preferably, the concentration ofthe enzyme component is an amount that is effective to tenderize, hydrolyze, modify and/or degrade the raw vegetable composition. Still more preferably, the concentration of the enzyme component is 15 an amount that is effective to degrade the first outer layer of a raw vegetable composition. Most preferably, the concentration of the enzyme component that is used in accordance with the present invention is an amount that is effective to degrade the first outer layer of a raw vegetable composition, tenderize, hydrolyze, modify and/or degrade the raw vegetable composition, and pennit further 20 modification of an inner portion of the raw vegetable composition. Furthermore, it is to be understood that the concentration of the enzyme component(s) may vary depending on the amount of time that the enzymes remain in contact with the raw vegetable composition. Furthermore, if a short exposure time is employed, then higher concentrations ofthe enzyme component(s) 25 would be required to achieve the desired degree of degradation, tenderization, hydrolysis and/or modification of the raw vegetable compositions. Similarly, if longer exposure times are employed, then the concentration of the enzyme component(s) would be reduced to arrive at the desired result.
WO 2004/060072 PCT/US2003/041646 9 As an example, when the enzyme component is supplied in the form of a liquid, the enzyme component can be applied at a concentration of less than about 10 weight percent, based on the total weight ofthe raw vegetable composition to tenderize the raw vegetable compositions like dry edible beans. More preferably, 5 when the enzyme component is supplied in the form of a liquid, the enzyme component is applied at a concentration of less than about 5 weight percent, based on the total weight of the raw vegetable composition to tenderize raw vegetable compositions. Similarly, when the enzyme component is supplied in the fonn of 10 a granular powder, the enzymes can be applied at a concentration of less than about 5 weight percent, based on the total weight of the raw vegetable composition to tenderize, hydrolyze and/or enzymatically modify the raw vegetable composition. More preferably, when the enzyme component is supplied in the form of a granular powder, the enzyme component is applied at a concentration of less than about 1 15 weight percent, based on the total weight of the raw vegetable composition to tenderize, hydrolyze and/or enzymatically modify the raw vegetable compositions. The aqueous enzyme composition may optionally include one or more catalyst component(s) in a form that is readily applied to the raw vegetable composition. A catalyst, when included as part of the aqueous enzyme 20 composition, generally enhances the biochemical rate of catalysis of the enzyme component(s). Increasing the biochemical rate of catalysis of the enzyme component(s) may decrease the application time of the aqueous enzyme composition to the rawvegetable composition or the amount ofthe aqueous enzyme composition applied to the raw vegetable composition. 25 Alternatively, the catalyst component may be applied separately from the aqueous enzyme composition, either before, during, or after application of the aqueous enzyme composition to the raw vegetable composition. Additionally, the source(s) of the catalyst may be applied in particulate form, as part WO 2004/060072 PCT/US2003/041646 10 of an aqueous composition, or in a vapor form so long as the particular form selected results in application to and uptake by the vegetable composition. Some non-exhaustive examples of catalysts that may be included as part of the aqueous enzyme composition are salts that include calcium ions, copper ions, magnesium 5 ions, iron ions, sodium ions, zinc ions, manganese ions, potassium ions, or any combination thereof. The catalyst component(s) may be supplied as individual components, or supplied in various prepared mixtures of two or more components, that are subsequently combined to form the catalyst component(s). The aqueous enzyme composition may include one or more pH 10 modifying component(s) that are capable of adjusting the acidity, hereinafter referred to as the pH, of the aqueous enzyme composition. Furthermore, the pH of the aqueous enzyme composition will vary depending on the enzyme(s) present in the aqueous enzyme composition. Preferably, the pH of the aqueous enzyme composition is about 2.0 15 to about 7.0. Still more preferably, the pH of the aqueous enzyme composition is about 3.0 to about 7.0 when degrading, tenderizing, hydrolyzing and/or enzymatically modifying the raw vegetable composition. In addition, extremely low pH values of less than about 1.0 are typically effective in deactivating the enzyme component(s) when practicing the present invention. Consequently, human 20 ingestion or addition of strong acids that reduce the pH of the aqueous enzyme composition to below a pH of about 2.0 are believed effective in deactivating the enzyme component(s) of the aqueous enzyme composition. Some non-exhaustive examples ofpH modifying substances include organic acids, such as acetic acid, tartaric acid, malic acid, succinic acid, citric acid, 25 or the like; phosphoric acid; or buffering agents of such organic acids, such as sodium acetate, sodium malate, sodium succinate, sodium citrate, or the like. Basic compounds like sodium hydroxide or the like may be included as part of the pH modifying substances that are suitable for use in the present invention.
WO 2004/060072 PCT/US2003/041646 11 As lipases and/or cutinases are used to hydrolyze wax- or fat containing substrates of the raw vegetable composition, an aqueous composition that contains an emulsifer or surfactant may be required in order to attain sufficient contact of the enzyme with the waxy coat of the raw vegetable composition. Fats 5 or waxes are typically hydrophobic while water is required for enzymatic hydrolysis. Therefore, in order to realize an efficient hydrolytic lipase reaction, at least 10 percent water must be available and the lipase and/or cutinase must be in intimate contact ofthe raw vegetable composition. Some non-exhaustive examples of emulsifiers or surfactants include such as mono-glycerides, distilled mono 10 glycerides, di-glycerides, distilled di-glycerides, or lecithin (natural or artificial), vegetable glycerin, glycerol, glycerin, or any combination of any of these. Preferably, the emulsifier and/or surfactant that is used in the present invention is any material that is compatible the process requirements and final vegetable product attributes. As an example, vegetable glycerin is used in accordance with the present 15 invention. Other examples of optional additives that may be included as part of the aqueous enzyme composition include natural and/or artificial flavors; artificial colors; naturally-occurring pigments, such as, for example, chlorophyll, anthocyanin, betalain, betaine, carotenoid, anthoxanthins; herbs; spices; vitamins; 20 minerals; plant extracts; essential oils; sugars such as sucrose, fructose, glucose, or maltose; preservatives; any additive that improves the aqueous enzyme composition application to, uptake by, or subsequent processing of the vegetable composition; or any combination of any of these. The aqueous enzyme composition may also include one or more 25 solvent component(s). The solvent component(s) preferably facilitate homogenous blending of the enzyme component(s), the optional catalyst component(s), the optional pH-modifying component(s), the optional additives, or any combination thereof. The solvent component(s) preferably facilitate aqueous enzyme WO 2004/060072 PCT/US2003/041646 12 composition application to, and uptake by the vegetable composition. Some non exhaustive examples of solvents that may be included in the aqueous enzyme composition include water; oils; alcohol, such as ethanol, methanol, propanol, butanol, or the like; hexane; or any combination thereof. The solvent component(s) 5 may be supplied as individual components, or supplied in various prepared mixtures of two or more components, that are subsequently combined to form the solvent component(s). Liquid water is the preferred solvent for the aqueous enzyme composition as water is typically required for enzymatic degradation, tenderization 10 and/or hydrolysis. The amount of liquid water included as part of the aqueous enzyme composition depends on an initial concentration of water in the raw vegetable composition, the biochemical rate of catalysis, and/or the desired final product characteristics of the enzyme-degraded raw vegetable composition. Generally, the amount of the aqueous enzyme composition is such that the raw 15 vegetable composition is completely contacted bythe aqueous enzyme composition. As an example, when degrading raw edible beans, water is included as part of the aqueous enzyme composition at a range of about 1.25 to about 5 times the weight of raw edible beans. Similarly, when tenderizing raw greens like collards, kale, turnip, or mustard greens, water is included as part of the aqueous enzyme 20 composition at a range of about 1 to about 2 times the weight of the raw greens. In general, any conventional blending apparatus and technique that is suitable for homogeneously blending the enzyme component(s), the optional catalyst component(s), the optional pH-modifying component(s), the optional additives, the optional solvent component(s), or any combination thereof, such as 25 a mixer, may be used to form the aqueous enzyme composition. As used herein, the tenn "application" means to apply the aqueous enzyme composition to the raw vegetable composition by spraying, knife-coating, spreading, soaking, exposing, immersing, slop-coating, dip-coating, roller- coating, WO 2004/060072 PCT/US2003/041646 13 dipping, contacting, brush-coating, squirting, submerging, foam padding, leaf sprinkling, sprinkling, pouring, slop-padding, or any combination thereof. The temperature ofthe aqueous enzyme composition depends on the initial temperature of the vegetable composition, the temperature for the optimum 5 biochemical rate of catalysis of the enzyme component(s), and/or the desired characteristics of the enzyme-degraded vegetable composition. The temperature of the aqueous enzyme composition is at the optimum temperature for a maximum biochemical rate of catalysis of the enzyme component(s) of the aqueous enzyme composition. 10 Generally, the temperature ofthe aqueous enzyme composition may range from about 30 F to about 250F. Preferably, the temperature of the aqueous enzyme composition ranges from about 30'F to about 250'F. Still more preferably, the temperature of the enzyme composition ranges from about 40'F to about 200 F. Most preferably, the temperature ofthe aqueous enzyme composition 15 ranges from about 40'F to about 195 F. Although the aqueous enzyme composition may be applied to the raw vegetable composition at a constant temperature, the temperature of the aqueous enzyme composition may be increased at any time during application of the aqueous enzyme composition to the raw vegetable composition. Generally, 20 increasing the temperature increases the biochemical rate of catalysis, and/or water absorption. However, a negative impact on the texture of the raw vegetable composition may occur if the temperature of the aqueous enzyme composition is too high, such as more than about 250 0 F, or the temperature of the aqueous enzyme 25 composition is changed too rapidly during application. Furthermore, too high temperatures may inactivate the enzyme component of the aqueous enzyme composition, therefore care is required to avoid premature inactivation of the enzyme component(s) before attaining the desired degree of hydrolysis, WO 2004/060072 PCT/US2003/041646 14 tenderization, degradation and/or enzymatic modification when practicing the present invention. Steam can also be injected into the aqueous enzyme composition to during or after application of the aqueous enzyme composition to the raw vegetable 5 composition to (1) optionally increase the temperature of the aqueous enzyme composition applied to the raw vegetable composition, (2) optionally increase the moisture content of the vegetable composition, (3) optionally gelatinize any starch granules of the vegetable composition, (3) optionally increase the efficacy of the biochemical rate of catalysis of the aqueous enzyme composition, or (3) optionally 10 deactivate the enzyme component in the aqueous enzyme composition. Preferably, steam is injected after the aqueous enzyme composition is applied to the raw vegetable composition if steam is included as part of the vegetable manufacturing process. As noted, inactivation of the enzyme component(s) readily occurs at high temperatures, such as temperatures that occurs with steam application, therefore, 15 care is required to avoid premature inactivation of the enzyme component(s) prior to attaining the desired degree of degradation, tenderization and/or hydrolysis ofthe raw vegetable composition. The aqueous enzyme composition is typically applied to the raw vegetable composition at normal atmospheric pressures. By "normal atmospheric 20 pressures" is meant atmospheric pressures of about 14.7 psi. Furthermore, it is to be understood that "normal atmospheric pressures" also includes atmospheric pressures that occurs even under varying altitudes, temperatures, humidities, or the like. Additionally, the term "normal atmospheric pressures" is not meant 25 to include application of positive pressure (more than about 14.7 psi) or negative pressures (less than about 14.7 psi or vacuum pressure conditions) to the raw vegetable composition prior to or during application of the aqueous enzyme WO 2004/060072 PCT/US2003/041646 15 composition in a manner that facilitates degradation, tenderization, hydration and/or hydrolysis. As used herein, the term "vegetable" means a plant-based food that originated as a living organism of the Plantae kingdom. All references to 5 "vegetable" are to be understood as encompassing any genetically-altered copy of the plant that originated as a living organism of the Plantae kingdom. Furthermore, the term "vegetable" encompasses leaves, seeds, roots, tubers, bulbs, flowers, fruits, stems, shoots, nuts, or any combination of any of these that originated as a living organism of the Plantae kingdom. 10 The raw vegetable composition of the present invention typically contains a first outer layer that substantially covers, overlays, and/or is in adhesive contact with a second inner layer of the raw vegetable composition when practicing the present invention. When the first outer layer is in adhesive contact with the second inner layer, adhesive contact may be accomplished through bonding via 15 cementing substances like pectic substances. The first outer layer of the raw vegetable composition may be characterized as a water-impermeable layer that typically includes a fibrous network of cellulose; xylan chains ofhemicellulose; hemicellulose; polysacecharides of five carbon sugars; lignin; pectic substances, such as protopectin, pectic acid, pectin, or 20 any combination thereof; vitamins; minerals; fats; anti-nutritional components; or any combination of any of these. Some non-exhaustive examples of the first outer layer may include a seed coat of a legume or lentil; a bran layer of a grain; a stem wall of a vegetable; a skin of a root, tuber, and/or bulb vegetable; a peel of a fruit; a testa or a seed wall of a nut. 25 The second inner layer of the raw vegetable composition generally includes a network of starch granules, fat globules, fiber, proteins, vitamins, minerals, water, phytochemicals, anti-nutritional components, or any combination of any of these. In addition, all references to the second inner layer is also WO 2004/060072 PCT/US2003/041646 16 understood to encompass the inner portion of the raw vegetable composition and thus, the second inner layer may also include seeds embedded in the vegetable composition. Some non-exhaustive examples of anti-nutritional components of a vegetable composition include flatulence-causing sugars, such as, for example, 5 raffinose, verbascose and stachyose; lectins; nutrient-binding substances, such as phytic acid; other indigestible polysaccharides; enzyme inhibitors, such as trypsin inhibitor; or toxic compounds, such as goitrogens, solanine, or oxalic acid. Preferred raw vegetable compositions of the present invention possess a tough hard second inner layer. As an example, legumes contain at least 10 one cotyledon that may be characterized as a tough fibrous network of starch, protein, anti-nutritional factors, fat, vitamins, and minerals. Similarly, grains, such as whole wheat or hominy contain an endosperm that may also be characterized as a tough fibrous network of starch, protein, anti-nutritional factors, fat, vitamins and minerals. Furthermore, raw vegetable compositions that have a hard second inner 15 layer typically have less than about 40 weight percent moisture content, and preferably less than about 30 weight percent moisture content. Raw vegetable compositions having a moisture content of less than about 40 weight percent and preferably less than about 30 weight percent can be effectively tenderized when practicing the present invention. 20 Preferably, the first outer layer is connected or in adhesive contact to the second inner layer or inner portion of raw vegetable compositions when practicing the present invention. By "connected or in adhesive contact" is meant that the raw vegetable composition has a substantial portion of the first outer layer connected to the inner portion or second inner layer of the raw vegetable 25 composition. Additionally, removal of the first outer layer of the raw vegetable composition by peeling, chemicals, grating for example is preferably avoided when practicing the present invention.
WO 2004/060072 PCT/US2003/041646 17 As used herein, the term "raw" refers to vegetable composition(s) that are uncooked, un-boiled, dry, edible, as being in a natural condition, or any combination of any of these. Furthermore, it is to be understood that the term "raw" refers to the condition of the first outer layer, the second inner layer or both the first 5 and second layers of the vegetable composition when practicing the present invention. In addition, the raw vegetable composition is preferably a whole raw vegetable composition. By "whole" is meant that the raw vegetable composition has not been subjected to techniques like maceration, pulverization, grating, 10 grinding or the like. For example, dry edible beans that have not been ground to a powder (flour), grated to form flakes, macerated or pulverized are examples of whole raw vegetable compositions. Similarly, green leafy vegetables such as collards, kale or the like that have not been ground, grated, macerated or pulverized are preferred examples of whole raw vegetables that may be used in accordance 15 with the present invention. In addition, preferred raw vegetable compositions for the present invention include raw vegetable compositions that contain an additional outer layer on top of the first outer layer. Examples of an additional outer layer on top of the first outer layer includes waxy layers (coats) of seeds, grains, legumes or the like. 20 Typically, the waxy coats include cutin or other wax- and/or lipid-containing molecules. Example of raw vegetable compositions that may contain a waxy coat/outer layer include beans like pinto beans, navy beans, light red kidney beans, black-eye peas, lentils, mung beans, pinkie beans, great northern beans, green lima 25 beans, yellow lima beans, garbanzo beans, carob beans, cacao beans, coffee beans, split and/or whole peas, peanuts, yellow peas, green peas, soybeans, black beans, vanillabean, or any other edible seed from plants ofthe Leguninosae family; seeds, such as amaranth seeds, watermelon seeds, pomegranate seeds, sunflower seeds, WO 2004/060072 PCT/US2003/041646 18 safflower seeds, poppy seeds, sesame seeds, alfalfa seeds, caraway seeds, cardamom seeds, celery seeds, chia seeds, coriander seeds, dill seeds, fennel seeds, fenugreek seeds, flax seeds, milk thistle seeds, nutmeg seeds, mustard seeds, psyllium seed; grains such as barley, buckwheat, hominy, pearl barley, bulghur, 5 amaranth, corn, millet, oats, rice, rye, triticale, wheat, wild rice, brown rice; or any seed from recognized edible vegetable source. In addition, raw vegetable compositions having a moisture content of more than about 40 weight percent, as disclosed in pending Application Serial Application No. 10/619,403 may also be effectively tenderized when practicing the 10 present invention. For example, green leafy vegetables that contain an outer layer of a waxy coat were tenderized using a blend of cellulase, hemicellulase, and pectinases. Thus, the inclusion of a lipase was not required. However, when practicing the present invention on grains such as wheat, rice or seeds/vegetable compositions that include the waxy coat, the inclusion of a lipase and/or cutinase 15 is preferred. Raw vegetable compositions that are generally in the form of a nut may also have less than about 40 weight percent moisture content and may also be included as part of the raw vegetable composition when practicing the present invention. As used herein, a "nut" means a hard shelled dry fruit or seed with a 20 separable first outer layer that substantially encloses an interior kernel. Some non-exhaustive examples of vegetable compositions in the form of a nut that may be used in accordance with the present invention include an acorn nut, an almond nut, a brazil nut, a butternut, a cashew nut, a chestnut, a coconut, a filbert nut, a hazelnut, a hickory nut, a macadamia nut, a pecan nut, a 25 pine nut, a pistachio nut, a walnut, or any recognized edible nut from a recognized edible vegetable source. It is also to be understood that the term "raw vegetable composition" is meant to encompass raw vegetable compositions that mayhave been washed with WO 2004/060072 PCT/US2003/041646 19 steam, hot, warm and/or cold water in an attempt to remove dirt and the like from the raw vegetable composition. Cleaning, washing or dirt removal from the vegetable composition may also include the application of food-grade detergents or chemicals using sprinkler-type equipment or soaking equipment. Such cleaning, 5 washing or dirt removal techniques are believed to not (1) significantly remove the first outer layer of the raw vegetable composition from the second inner layer or inner portions of the raw vegetable compositions, and/or (2) substantially reduce the fibrous components of the raw vegetable composition prior to enzyme application, such as by reducing the fibrous component by more than about 1 10 weight percent, based on the total weight of the raw vegetable composition. Therefore, use of such cleaning, washing or dirt removal techniques prior to application ofthe aqueous enzyme composition are permissible when practicing the present invention. In addition, physical and/or chemical pretreatment strategies 15 designed to initiate breakdown, improve the porosity of the first outer layer of raw vegetable compositions, remove certain cellulose and hemicellulose fractions or expose degradation sites have been practiced in the widespread belief that enzymatic degradation cannot proceed without such pre-treatment strategies. Physical pre-treatment strategies includes application of positive or negative 20 pressure prior to application of the aqueous enzyme composition to vegetable compositions. Furthermore, chemical pre-treatment strategies include application of strong acid solutions, pre-soaking, boiling or cooking of vegetable compositions that typically increase the moisture content of vegetables prior to application of the aqueous enzyme composition. 25 The present invention avoids these complicated processing strategies by applying the aqueous enzyme composition to the raw vegetable composition under normal atmospheric pressures without having first subjected the raw vegetable composition to strong acidic solutions, pre-soaking, boiling or cooking WO 2004/060072 PCT/US2003/041646 20 prior to application of the aqueous enzyme composition. Such physical and/or chemical treatments are typically reserved, and preferably conducted after the aqueous enzyme composition has degraded the raw vegetable composition to deactivate the enzyme component(s). 5 It is also to be understood that the term "whole raw vegetable composition" is meant to encompass broken a raw vegetable composition that (1) has a first outer layer that is in adhesive contact with the second layer, or (2) an exposed second inner layer or inner portion of the raw vegetable composition. For example, in the manufacture of refried beans, broken portions of whole beans still 10 contain a seed coat and exposed cotyledons. Such broken portions of whole raw beans with a moisture content of less than about 30 weight percent can be soaked or exposed to the aqueous enzyme composition of the present invention to degrade the seed coat and tenderize the cotyledons prior to human consumption or any subjecting the beans to any other remaining processing steps required for 15 manufacturing refried beans. Similarly, raw green leafy vegetables may be chopped prior to application of the aqueous enzyme composition to permit enzymatic degradation and subsequent tenderization of the raw green leafy vegetables as chopping is not believed to remove or diminish the fibrous network present in raw green leafy 20 vegetables. Similarly, the common industrial practice of application of heat (wilting) of greens prior to canning is permissible when practicing the present invention as the wilting step is not believed to substantially reduce the fibrous components present in leafy greens, such as by more than about 1 weight percent fiber, based on the total weight of the raw vegetable composition. Rather, the 25 wilting step is believed to improve compaction of the leafy greens for subsequent inclusion into cans. As noted above, the length oftime the aqueous enzyme composition is applied to the raw vegetable composition typically depends on the raw vegetable WO 2004/060072 PCT/US2003/041646 21 composition, the desired degree of degradation, the concentration of the enzyme components) and/or the desired characteristics of the enzyme-degraded vegetable composition. The length of time used in practicing the present invention may range from about 1 second to more than about 24 hours. As examples, the length of time 5 to degrade raw vegetable compositions having a moisture content of less than about 30 weight percent is about 1 second to about 2 hours while the length of time to tenderize raw vegetable compositions is about 1 second to about 2 hours as well. While not wanting to be bound to theory, it is believed that the lipase degrades the additional outer waxy layer or lipid-containing layer of the raw 10 vegetable compositions to form a network of degraded sites. Next, the preferred cellulase, hemicellulase and pectinase are able to penetrate the network of degraded sites and initiate hydrolysis of carbohydrate containing target substrates of the first outer layer. As hydrolysis of carbohydrate-containing target substrates is also believed to form a network of degraded sites or holes, the enzymes that are included 15 as part of the aqueous enzyme composition are able to facilitate degradation so that effective tenderization of the raw vegetable compositions can occur. Therefore, the aqueous enzyme composition may generate holes throughout the waxy top layer, the first outer layer and/or the second inner layer of the vegetable composition so that absorption of additives or enzymes and effective 20 hydrolysis of target substrates are observed. The aqueous enzyme composition may also target a wide range of substrates within the raw vegetable composition, therefore, the breakdown of these substrates may occur and aid in the reduction of cook time of the enzyme-degraded raw vegetable composition. The benefits of the enzyme-degraded raw vegetable composition 25 include formation of a tenderized raw vegetable composition or a reduction in the fibrous components of the raw vegetable composition. For example, when practicing the present invention, the fibrous components of the raw vegetable composition may be reduced by more than about 0.5 weight percent, preferably WO 2004/060072 PCT/US2003/041646 22 more than about 1 weight percent, still more preferably more than about 5 weight percent, based on the total weight of the initial raw vegetable composition. In addition, processing the enzyme-degraded raw vegetable composition by conventional means, after enzymatic degradation, such as by freezing, hydrating, 5 steaming, freeze-drying, canning, frying, boiling, drying, extrusion, cooking, baking, roasting, pulverizing, fermenting, enzyme, pasteurizing, extracting, milling, puffing, steam-pressure cooking, or any combination thereof, is improved since the first outer layer of the raw vegetable composition that typically functions as a barrier during conventional processing has been degraded. For example, cooking 10 times of raw vegetables, such as grains or legumes have been reduced by about 10 to about 75 percent when compared to untreated controls. Once sufficient degradation of the raw vegetable composition has occurred to form the enzyme-degraded raw vegetable composition, the enzyme degraded raw vegetable composition may be separated from the aqueous enzyme 15 composition and further subjected to processing steps, such as, for example, blanching, that inactivates any enzyme component(s) remaining in the enzyme degraded raw vegetable composition. Alternatively, transferring both the raw enzyme-degraded vegetable composition and the aqueous enzyme composition to equipment that permits further processing by freezing, hydrating, steaming, freeze 20 drying, canning, frying, boiling, drying, extrusion, cooking, baking, roasting, pulverizing, fermenting, enzyme, pasteurizing, extracting, milling, puffing, steam pressure cooking, or any combination thereof, is also effective in deactivating any enzyme component(s) remaining in the enzyme-degraded raw vegetable composition and the aqueous enzyme composition. 25 The present invention is more particularly described in the following examples that are intended as illustrations only since numerous modifications and variations within the scope of the present invention will be apparent to those skilled in the art.
WO 2004/060072 PCT/US2003/041646 23 A method of degrading and/or tenderizing raw vegetable compositions A series of experiments were conducted to degrade and/or tenderize raw vegetable compositions. Tenderization of raw vegetable compositions can be measured by observing a reduction in the cook and/orprocess time ofraw vegetable 5 compositions when compared to raw vegetable compositions that have not been subjected to enzymatic degradation. An amount of raw vegetable compositions were contacted with an aqueous enzyme composition containing an amount of water, vinegar, enzyme and surfactant (see Table 1 below). Viscozyme L 120 that is available from Novozymes 10 of Franklinton, North Carolina was used as the carbohydrase. The density of Viscozyme L 120 is about 1.2 grams per milliliter. Therefore, one teaspoon of Viscozyme L 120 contains about 6 grams of enzyme. Lipase "A" Amano 12 that is available from Amano Company of Japan was used in these experiments. Blanching, if implemented was conducted at about 200'F for about 5 minutes. 15 Next, the blanched vegetables were cooked until done or until there was (1) no observable ungelatnized starch portions (or uncooked portions) in the raw vegetable composition and/or (2) no detection of any fibrous components when chewing the cooked vegetable composition. The control experiments did not include any enzyme. Rather, the 20 control experiments contained (same) equal amounts of the raw vegetable that was used during the enzyme-treated experiments, vegetable glycerin (Whole Foods Market, Minneapolis, MN), water and vinegar. The aqueous composition (water, vegetable glycerin, water and vinegar) was heated to approximately the same initial temperature range that was used during the enzyme-treated experiments. The 25 amount of the aqueous composition that was absorbed by the raw vegetables was substantially the same as the amount of the aqueous enzyme composition that was absorbed during enzymatic treatment of the raw vegetables. However, the cooking time was significantly shorter for the enzyme-treated vegetables when compared to WO 2004/060072 PCT/US2003/041646 24 the controls that were not subjected to any enzyme treatment. The details of the experiments are presented in Table 1 below: TABLE 1 5 Conditions uncooked white uncooked uncooked rice brown rice hominy Initial Wt (gr) about 250 about 250 about 250 Lipase (teaspoon) about one-eighth about one- about one eighth eighth Carbohydrase (gr) abut 3 about 3 about 3 Water (gr) about 750 about 750 about 750 10 Vegetable glycerin (teaspoon) about 1 about 1 about 1 Vinegar (teaspoon) about 1 about 1 about 1 Initial Temp (F) about 109.1 about 110 about 110.1 Initial pH about 4.71 about 5.12 about 5.08 Final Temp (F) about 92.4 about 93.3 15 Final pH - about 5.34 Application technique soaking soaking soaking Application Time (minutes) about 60 about 60 about 60 Final weight (gr)* about 420 about 318 about 319 Blanch weight (gr)# - 20 Cook time (minutes) - about 21 about 60 Control experiment cook time - - about 40 to 45 about 90 (minutes) - - indicates this step was not performed or the variable was not measured.
WO 2004/060072 PCT/US2003/041646 25 TABLE 2 Conditions uncooked whole uncooked uncooked grain wheat pearl barley navy beans Initial Wt (gr) about 250 about 250 about 250 5 Lipase (teaspoon) about one-eighth about one- about one eighth eighth Carbohydrase (gr) abut 3 about 3 about 3 Water (gr) about 750 about 750 about 750 Vegetable glycerin (teaspoon) about 1 about 1 about 1 Vinegar (teaspoon) about 1 about 1 about 1 10 Initial Temp (F) about 106.1 about 105.4 about 105.7 Initial pH about 4.98 about 5.31 about 5.37 Final Temp (F) 91.3 about 92.8 about 90.1 Final pH 5.21 5.27 about 5.65 Application technique soaking soaking soaking 15 Application Time (minutes) about 60 about 60 about 60 Final weight (gr)* about 313 about 388 about 447 Blanch weight (gr)" - Cook time (minutes) about 30 about 17 about 30 Control experiment cook time about 60 about 40 about 45 20 (minutes) - - indicates this step was not performed or the variable was not measured. TABLE 3 Conditions uncooked uncooked uncooked amaranth white corn navy beans 25 Initial Wt (gr) about 250 about 264 about 250 Lipase (teaspoon) about 1/8 about 1/8 'about 1/8 WO 2004/060072 PCT/US2003/041646 26 Carbohydrase (gr) abut 3 about 3 about 3 Water (gr) about 750 about 750 about 750 Vegetable glycerin (teaspoon) about 1 about 1 about 1 Vinegar (teaspoon) about 1 about 1 about 1 5 Initial Temp (F) about 105.7 about 113.9 about 105.7 Initial pH about 4.91 about 5.11 about 5.37 Final Temp (F) 92.6 about 92.5 about 90.1 Final pH 5.38 4.57 about 5.65 Application technique soaking soaking soaking 10 Application Time (minutes) about 60 about 60 about 60 Final weight (gr)* about 475 about 331 about 447 The enzyme-degraded white corn kernels were white popcorn kernels. After soaking, the popcorn kernels were dried to a weight of about 245 15 grams. After drying, the popcorn kernels were popped using about 1 tablespoon of butter. About one-quarter cup of popcorn kernels yielded about 1.5 quarts of popcorn. Similarly, about 250 grams of raw collards were sprayed with an aqueous enzyme composition that contained about 12-13 grams ofViscozyme, 740 20 grams of water and enough vinegar to reach an initial pH of about 4.0. In addition, the initial temperature of the aqueous enzyme composition was about 110 F. The raw collards were allowed to soak for about 60 minutes and then cooked in the aqueous enzyme composition. The raw greens cooked in about 45 minutes compared to the time of more than 2 hours that were required to cook raw greens 25 not subjected to enzymatic treatment. Subsequent experiments using the same amount of greens, enzyme and vinegar conditions that were permitted to soak for about 15 minutes or about 30 minutes also cooked in about 45 minutes compared to the time of more than 2 hours that were required to cook raw greens not subjected to enzymatic treatment.
WO 2004/060072 PCT/US2003/041646 27 A method of enzymatically processing a raw vegetable composition Typically application of the aqueous enzyme composition that includes first and second enzyme components occur under normal atmospheric pressures and temperatures that can range from about 40 F to about 250F and 5 preferably from about 40 F to about 195 F. In addition, the pH values range from about 2.0 to about 7.0. Alternatively, two or more separate aqueous enzyme compositions may be applied to the raw vegetable composition when practicing the present invention. For example, a first aqueous enzyme composition that includes cellulase 10 and hemicellulase maybe applied to a raw vegetable composition having a moisture content of less than about 30 weight percent under normal atmospheric pressures and temperatures to form an enzyme-degraded raw vegetable composition. Next, a second aqueous enzyme composition that contains an enzyme component that is effective to degrade and/or hydrolyze target substrates either in the first outer layer 15 or the second inner layer can be applied to the enzyme-degraded vegetable composition. The second aqueous enzyme composition is able to penetrate the enzyme-degraded vegetable composition and therefore, is capable of degrading and/or hydrolyzing desired target substrates in the enzyme-degraded vegetable composition. 20 It is believed that the compromised first outer layer of the enzyme degraded raw vegetable composition allows the second enzyme component or the second aqueous enzyme composition to enter and degrade any anti-nutritional components in the enzyme-degraded raw vegetable composition. Additionally, water included as part of the aqueous enzyme composition(s) may also enter 25 through the degraded first outer layer to hydrate the enzyme-degraded raw vegetable composition. If water is absorbed by the enzyme-degraded raw vegetable composition, the water in the enzyme-degraded raw vegetable composition may facilitate degradation of anti-nutritional components of the enzyme-degraded raw WO 2004/060072 PCT/US2003/041646 28 vegetable composition by the second enzyme component or the second aqueous enzyme composition. After sufficient enzymatic degradation by the second enzyme component, or the second aqueous enzyme composition, an enzyme-processed raw 5 vegetable composition is formed that can be further subjected to other processing steps, such as, for example, blanching, that inactivates any enzyme component(s) remaining in the enzyme-processed vegetable composition. Preferably, the enzymes that are included as part of the first enzyme component include the above noted enzymes that are effective in degrading the first 10 outer layer of raw vegetable compositions. The enzyme(s) that may be included as part of the second enzyme component or second aqueous enzyme composition are carbohydrases, proteases or any combination thereof. Any of the examples of carbohydrases as suitable for use during application of the first enzyme component may be used as part of the second enzyme component in any combination with the 15 first enzyme component, for degradation of any anti-nutritional component of the raw vegetable composition in accordance with the present invention. As used herein, the term "protease" means any enzyme that is capable of at least catalyzing degradation of a protein-containing target substrate. One particular form of a protease that may be used as part of the second enzyme 20 component in accordance with the present invention is an endoprotease. As used herein, an "endoprotease" means any enzyme that is capable of degrading an internal peptide bond on a target substrate having one or more peptide bonds. Another particular form of a protease that may be used as part of the second enzyme component in accordance with the present invention is an "exoprotease". As used 25 herein, an "exoprotease" means any enzyme that is capable degrading a peptide bond located at a terminal portion of a target substrate having one or more peptide bonds. Either the endoprotease or the exoprotease may be derived from a number WO 2004/060072 PCT/US2003/041646 29 of different sources, such as fungal sources, plant sources, microbial sources, animal sources, or any combination of any of these. Some non-exhaustive examples of endoproteases and exoproteases include Alcalase, Neutrase* Esperase*, Protamex, Novozym* FM, Flavourzyne*, 5 and Kojizyrne@, all available from Novo Nordisk Biochem North America of Franklinton, North Carolina, and Enzeco@ exoprotease that is available from Enzyme Development Corporation of New York, New York. The enzyme-processed vegetable composition may also be further processed by freezing, hydrating, steaming, freeze-drying, canning, frying, boiling, 10 drying, extrusion, cooking, baking, roasting, pulverizing, fermenting, enzyme, pasteurizing, extracting, milling, puffing, steam-pressure cooking, or any combination thereof after enzymatic degradation has occurred. These additional processing steps are also generally effective in deactivating any enzyme component(s) in the enzyme-processed vegetable composition. Partial degradation 15 of anti-nutritional components in the first outer layer or the second inner layer of the vegetable composition may also occur during application of aqueous enzyme compositions that includes first and second enzyme components or the second aqueous enzyme composition. As a first example, an enzyme-degraded raw vegetable composition 20 that includes flatulence-causing substrates located in an inner portion of the raw vegetable composition is formed by applying a first aqueous enzyme composition that degrades the raw vegetable composition in accordance with the present invention. If a second aqueous enzyme composition that includes any enzyme component capable of degrading any flatulence-causing substrates that cause 25 flatulence in human, such as alpha-galactosidase, beta-fructofuranosidase, beta galactosidase, invertase, or any combination thereof, is applied to the enzyme degraded raw vegetable composition either during or after the first aqueous enzyme composition is applied to the raw vegetable composition, degradation of flatulence- WO 2004/060072 PCT/US2003/041646 30 causing substrates, such as raffinose, verbascose and stachyose of the enzyme degraded raw vegetable composition typically occurs. Generally, the second aqueous enzyme composition is applied to the enzyme-degraded vegetable composition for a time that is sufficient for the second 5 enzyme composition to degrade the flatulence-causing substrates, such as, for example, about 1 minute to about 12 hours. Preferably, the second aqueous enzyme composition remains in contact with the enzyme-degraded raw vegetable composition for about 5 minutes to about 120 minutes so that more than about 5 weight percent of flatulence-causing substrates are degraded in the enzyme 10 degraded raw vegetable composition when practicing the present invention. As a second example, an enzyme-degraded raw vegetable composition that includes methylxanthine is formed in accordance with the present invention. As used herein, the term "methylxanthine" refers to the group of compounds used as a stimulant and diuretic typically found in vegetable compositions, such as tea, 15 coffee, kola nuts, mat6 leaves, cacao bean, guarana and the like. It is understood that "methylxanthine" includes substituted forms of methylxanthine, such as, for example, caffeine. If a second aqueous enzyme composition that is capable of degrading methylxanthine, is applied to the enzyme-degraded raw vegetable composition either during or after application of a first aqueous enzyme 20 composition is applied to the raw vegetable composition, degradation of methylxanthine in the enzyme-degraded raw vegetable composition thereby occurs to reduce the level of methylxanthine in the enzyme-degraded raw vegetable composition. Preferably, the second aqueous enzyme composition is applied to the 25 enzyme-degraded vegetable composition for a time that is sufficient to degrade methylxanthine, such as, for example, about 1 minute to about 8 hours. Still more preferably, the second enzyme composition that is capable of degrading methylxanthine remains in contact with the enzyme-degraded raw vegetable WO 2004/060072 PCT/US2003/041646 31 composition for a time that is sufficient to degrade more than about 5 weight percent methylxanthine of the raw vegetable composition when practicing the present invention. After enzymatic degradation, the enzyme-processed vegetable composition having reduced levels of methylxanthine can then be blanched to 5 inactivate anyremaining enzymes and/or further processed bypulverizing, grinding, milling, roasting, freezing, drying, freeze-drying, or any combination thereof. Subsequent additional processing bypulverizing, blending, grinding, paste-forming, roasting, freezing, drying, freeze-drying, extraction, or any combination thereof, may also deactivate the enzyme(s). 10 The benefits of processing vegetable compositions that include methylxanthine in accordance with the present invention include reducing the need for expensive solvent extraction equipment and chemicals that are traditionally required to decaffeinate raw vegetable compositions, such as, for example coffee bean. Additionally, the present embodiment may improve the flavor of 15 decaffeinated coffee beans which may result in an increase in market share for a decaffeinated coffee manufacturer selling the enzyme-processed coffee beans. As a third example, bitter flavor notes that characterize green unfermented cocoa beans may be reduced in accordance with the present invention. Cocoa beans can be divided into four categories, according to their color: fully 20 fermented, i.e., predominantly a brown hue; purple/brown; purple; and slaty, in which slatybeans represent unfermented or green cocoa beans. Purple/brown cocoa beans include all beans showing any blue, purple or violet color on an exposed surface, whether suffused or as a patch of the cocoa beans. Purple cocoa beans should include all cocoa beans showing a completely blue, purple or violet color 25 over the whole exposed surface. An enzyme-degraded green unfermented cacao bean is formed in accordance with the present invention after application of an aqueous enzyme composition that includes cellulase, hemicelluase and pectinase degrades the first WO 2004/060072 PCT/US2003/041646 32 outer layer of green, unfermented or slaty cocoa beans. As used herein, a "green or unfermented cacao bean" includes cacao beans that do not have a sufficient quantity of amino acids and peptides required to form an acceptable cocoa flavor during subsequent roasting. Furthermore, the "green or unfermented cacao bean" include 5 beans having less than about 40 weight percent moisture content and that have not been subjected to a fermentation step. Some non-exhaustive examples of green unfermented cacao bean that may be used in accordance with the present invention include beans derived from Theobrona sp., such as Ghanian cacao beans, Amelonado sp., Criollo, Forastero, or Trinitario. 10 If a second aqueous enzyme composition that includes an endoprotease, an exoprotease, or any combination thereof, is absorbed by the enzyme-degraded green, slaty or unfermented cacao bean, degradation of protein in the enzyme-degraded green unfermented cacao bean occurs. Preferably, the endoprotease, the exoprotease, or any combination thereof, is capable of degrading 15 protein that include hydrophobic amino acids and peptides that typically contribute to the bitter flavor notes that characterize green or unfermented cacao beans. Still more preferably, the endoprotease, the exoprotease, or any combination thereof, is applied to the green or unfermented enzyme-degraded cacao bean for a time, temperature, pH and moisture content of the green unfermented enzyme-degraded 20 cacao bean that is sufficient to degrade the bitter flavor notes in the green unfermented cacao bean. While not wanting to be bound to theory, it is believed that the enzyme-degraded cacao bean includes sites through which the endoprotease, exoprotease, or any combination thereof, may be absorbed. Furthennore, the sites 25 in the enzyme-degraded green unfermented cacao bean may facilitate subsequent natural fermentation of the green unfermented cacao bean by enhancing the capability of indigenous microflora of the natural fermentation process to colonize WO 2004/060072 PCT/US2003/041646 33 and digest the green unfermented cacao bean at the sites during the fermentation process. The benefit of processing green unfermented cacao beans in accordance with the present invention include obtaining a less bitter cacao bean 5 after fermentation and roasting steps. A less bitter cacao bean requires little flavor modification required during manufacture of cocoa containing products. In an example of practicing the method of reducing the flatulence causing substrates in a raw vegetable composition, about 740 grams of water was added to about 7.5 grams of vinegar and brought up to a temperature of about 10 150'F. About 2.5 milliliters of Viscozyme*L, and 2.5 milliliters of Alpha-GalTM 600L, supplied by Novo Nordisk Biochem North America Inc., of Franklinton, North Carolina, were added to the vinegar and water mixture to form an aqueous enzyme composition with an initial pH of about 5.0. About 250 grams of raw collard greens were added to the aqueous enzyme composition and allowed to soak 15 for about 30 minutes. The raw collard greens were then cooked for about 30 minutes at about 200 F in the soak water. After cooking, the collard greens were drained and evaluated. Little, if any, flatulence was experienced after consumption of about 100 grams of collards even after 4 hours from the time of consumption of the collard greens. 20 In another example of practicing the method of reducing the flatulence-causing sugars in araw vegetable composition, about 740 grams ofwater was added to about 7.5 grams of vinegar and brought up to a temperature of about 119'F to about 123 F. About 2.5 milliliters of Viscozyme*L, and 1.25 milliliters of Alpha-Gal T M 600L, supplied by Novo Nordisk Biochem North America Inc., of 25 Franklinton, North Carolina, were added to the vinegar and water mixture to form an aqueous enzyme composition with an initial pH of about 5.0. About 250 grams of raw great northern beans were added to the aqueous enzyme composition and allowed to soak for about 60 minutes. The great northern beans were then blanched WO 2004/060072 PCT/US2003/041646 34 for about 5 minutes at about 200'F to deactivate the enzymes. The great northern beans did not result in any observable flatulence after human consumption. A method of modifying a vegetable composition As noted, the enzyme-degraded raw vegetable composition of the 5 present invention is capable of absorbing additives to form a modified vegetable composition. The method of modifying a vegetable composition in accordance with the present invention is a significant improvement in the art of vegetable processing. Typically, a vegetable composition is mechanically modified, such as by peeling, grinding, pulverizing, prior to inclusion of an additive, followed by 10 subsequent processing by conventional means. The added step of mechanical modification, along with any safety and health hazards that accompany using equipment involved in the mechanical modification of vegetables, may be eliminated when practicing the present invention. Additionally, the present invention accomplishes in situ modification of a raw vegetable composition. 15 As an example of the present embodiment that modifies raw vegetable composition, about 740 grams of water was added to about 7.5 milliliters of vinegar and brought up to a temperature of about 119 F to about 123 F. About 2.5 milliliters ofViscozyme*L, supplied by Novo Nordisk Biochem North America Inc., of Franklinton, North Carolina, was added to the vinegar and water mixture 20 to form an aqueous enzyme composition with an initial pH of about 4.8. About 250 grams of raw pinto beans were added to the aqueous enzyme composition and allowed to soak for about 60 minutes. The change in pH, indicating the absorption of vinegar is presented in Table 3 below: WO 2004/060072 PCT/US2003/041646 35 TABLE 4 TIME (minute) TEMPERATURE ("F) pH 0 123.1 4.78 15 118.5 5.11 5 30 117.7 5.38 45 118.9 5.56 60 119.2 5.70 Conclusion 10 In view of the foregoing disclosure and embodiments, it is believed that processing a raw vegetable composition in accordance with the present invention represents a significant improvement in the art of vegetable processing. The development of an effective process that reduces the complexity and costs associated with vegetable production, byreducing the first outer layer of a vegetable 15 composition that typically hinders processing, creates a vegetable product with enhanced processing characteristics. Furthermore, the development of an in situ method of processing and modifying a raw vegetable composition greatly enhances the ability of a food manufacturer to produce vegetable products that offer a wide variety of nutritional characteristics to consumers. 20 Although the present invention has been described with reference to preferred embodiments, workers skilled in the art will recognize that changes may be made in form and detail without departing from the spirit and scope of the invention. 25

Claims (13)

1. A method of enzymatically degrading a raw vegetable composition for subsequent human consumption, the method comprising: providing a raw vegetable composition having a moisture content of 5 less than about 30 weight percent; applying an aqueous enzyme composition comprising water, lipase and a cellulase to the raw vegetable composition under normal atmospheric pressures for a time that is sufficient to degrade raw vegetable composition, wherein the aqueous enzyme composition 10 is at a pH of between about 2.0 and 7.0; and deactivating the, first enzyme composition.
2. The method of claim 1 wherein the aqueous enzyme composition is effective to degrade a first outer layer of the raw vegetable composition. 15
3. The method of claim 1 wherein the aqueous enzyme composition is effective to tenderize the raw vegetable composition.
4. The method of claim 1 and further including applying a second 20 aqueous enzyme composition to the raw vegetable composition, wherein the second aqueous enzyme composition is selected from the group consisting of alpha-galactosidase, mannanase, beta-gluconase, beta-gluconase, arabinase, xylanase, beta-galactosidase, invertase, beta-fructofuranosidase, alpha-amylase, beta-amylase, pectinase, pectin depolymerase, pectin methyl esterase, pectin 25 lyase, glucoamylase, oligo-1,6 glucosidase, lactase, beta-d-glucosidase, and any combination thereof. WO 2004/060072 PCT/US2003/041646 37
5. A method of enzymatically processing a vegetable composition for subsequent human consumption, the method comprising: providing a raw vegetable composition having a moisture content of less than about 30 weight percent; 5 applying a first enzyme composition comprising water, lipase and a cellulase to the raw vegetable composition under normal atmospheric pressures for a time that is sufficient to form an enzyme-degraded raw vegetable composition, wherein the first enzyme composition is at a pH of between about 2.0 and 7.0; 10 applying a second enzyme composition comprising water and a carbohydrase to the enzyme-degraded raw vegetable composition; and deactivating the first enzyme composition and the second enzyme composition. 15
6. The method of claim 5 wherein the second enzyme composition is selected from the group consisting of hemicellulase, alpha-galactosidase, mannanase, beta-gluconase, beta-gluconase, arabinase, xylanase, beta galactosidase, invertase, beta-fructofuranosidase, alpha-amylase, beta-amylase, 20 pectinase, pectin depolymerase, pectin methyl esterase, pectin lyase, glucoamylase, oligo-1,6 glucosidase, lactase, beta-d-glucosidase, and any combination thereof.
7. A method of processing a vegetable composition for subsequent 25 human consumption, the method comprising: providing a raw vegetable composition having a moisture content of less than about 40 weight percent; WO 2004/060072 PCT/US2003/041646 38 applying an enzyme composition having a pH of between about 2.0 and 7.0 to the raw vegetable composition under normal atmospheric pressures for a time that is sufficient to degrade raw vegetable composition, wherein the enzyme composition includes 5 water, a first enzyme component, and a second enzyme component, wherein the first enzyme component includes a cellulase that degrades the raw vegetable composition, and wherein the second enzyme component includes a protease that degrades a protein or a peptide; and 10 deactivating the enzyme composition.
8. The method of claim 7 wherein the raw vegetable composition bean is a green or unfermented cocoa bean. 15
9. The method of claim 7 wherein the protease degrades a hydrophobic amino acid containing protein, a hydrophobic amino acid-containing peptide, or any combination of any of these.
10. A method of processing a vegetable composition for subsequent 20 human consumption, the method comprising: providing a raw vegetable composition having a moisture content of less than about 30 weight percent; applying an enzyme composition having a pH of between about 2.0 and 7.0 to the raw vegetable composition for a time that is 25 sufficient to degrade raw vegetable composition, wherein the enzyme composition includes water, a first enzyme component, and a second enzyme component, wherein the first enzyme component includes a cellulase that is effective to degrade the WO 2004/060072 PCT/US2003/041646 39 raw vegetable composition, wherein the second enzyme component is effective to degrade methylxanthine; and deactivating the enzyme composition. 5
11. The method of claim 10 wherein the raw vegetable composition is a green coffee bean, a cacao bean, a guarana, a cola nut, or any combination thereof.
12. A method of tenderizing a vegetable composition for subsequent 10 human consumption, the method comprising: applying an enzyme composition that includes water and a cellulase to a raw vegetable composition under normal atmospheric pressures, wherein the enzyme composition has a pH of between about 2.0 and 7.0, and wherein the enzyme composition is 15 effective to tenderize the raw vegetable composition; and deactivating the enzyme composition.
13. The method of claim 12 wherein the enzyme composition is effective to reduce a cook time of the bean. 20
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EP1906769A2 (en) * 2005-07-15 2008-04-09 Amano Enzyme USA., Ltd. Enzyme compositions that enhance the flavor of food and beverages
ES2343847T3 (en) 2005-09-06 2010-08-11 Biogreen A/S DEMUCILAGINATED LINEN BUDS AND THEIR SUBPRODUCTS AS WELL AS THEIR PRODUCTION AND APPLICATION.
JP5031508B2 (en) * 2007-04-19 2012-09-19 ハウス食品株式会社 Fenugreek seed with reduced bitterness and method for producing the same
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MA37165B1 (en) * 2014-06-26 2016-08-31 Mascir Morrocan Foundation For Advanced Science Innovation & Res Enzymatic method for extracting almonds from dried fruits from a lignocellulosic shell

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US4214007A (en) * 1978-01-16 1980-07-22 Kanebo Foods, Ltd. Processed dried beans as instant food and their manufacturing method
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