Method for dough relaxation involving endopeptidases

Abstract

A method for increasing the softness and relaxation of dough, the method comprising a) adding at least one endopeptidase having at least 60% identity to SEQ ID NO: 1 or SEQ ID NO: 2 to flour or to a dough comprising flour, wherein L-cysteine ​​and / or sodium metabisulfite is not added to the dough; b) kneading the dough; and c) forming the dough into an edible product.

Description

[0001] References to sequence listings

[0002] This application contains a Sequence Listing in computer readable form, which is incorporated herein by reference. Technical Field

[0003] The present invention relates to a method for improving dough rheology, for example improving the relaxation and softness of the dough when producing, for example, bread, flatbreads, crackers, pizza, pasta, noodles, laminated bakery products, biscuits, baguettes and hamburgers. Background Art

[0004] Currently, in industrial dough making processes, it is known to add dough improving additives to the dough in order to improve parameters of the dough, such as texture, volume, extensibility, pan flow and mechanical capabilities.

[0005] L-cysteine ​​and sodium metabisulfite (SMS) are known dough improving additives used to improve the rheological properties of dough, such as softness, stretchability, relaxation, etc.

[0006] Biological solutions for improving dough rheology in dough production, such as relaxation, stretchability and softness, are sought when making products such as bread, flatbreads, crackers, pizza, pasta, noodles, laminated bakery products, cookies, baguettes and hamburgers. Summary of the invention

[0007] Surprisingly, the inventors have found that a specific type of endopeptidase improves dough relaxation, thereby avoiding the addition of compounds such as L-cysteine ​​and / or sodium metabisulfite (SMS), and we therefore claim:

[0008] A method for increasing the softness and relaxation of dough, the method comprising

[0009] a) adding at least one endopeptidase having at least 60% identity to SEQ ID NO: 1 or SEQ ID NO: 2 to flour or to a dough comprising flour, wherein no L-cysteine ​​and / or sodium metabisulfite is added to the dough;

[0010] b) kneading the dough; and

[0011] c) forming the dough into an edible product.

[0012] In one embodiment, the edible product is selected from the group consisting of bread, flatbread, crackers, pasta, noodles, laminated bakery products, cookies, baguettes, hamburgers and pizza.

[0013] In one embodiment, the dough is formed into cookies.

[0014] In one embodiment, the flour is selected from the group consisting of wheat flour, corn flour, rye flour, barley flour, oat flour, rice flour, sorghum flour, and combinations thereof.

[0015] In one embodiment, the amount of energy used to knead the dough is reduced compared to a method wherein an endopeptidase having at least 60% identity to SEQ ID NO: 1 or SEQ ID NO: 2 is not added to the dough.

[0016] In one embodiment the endopeptidase is added in an amount ranging from 0.1-1000 mg enzyme protein / kg flour.

[0017] In one embodiment, the shape of the edible product is better maintained compared to a method wherein an endopeptidase having at least 60% identity to SEQ ID NO: 1 or SEQ ID NO: 2 is not added to the dough.

[0018] In one embodiment, the extensibility of the dough is better than the extensibility of a dough prepared under the same conditions but not treated with endopeptidase.

[0019] In one embodiment, the eating quality of the edible product is better than the eating quality of an edible product prepared under the same conditions but not treated with the endopeptidase.

[0020] In one embodiment, the dough further comprises one or more enzymes selected from the group consisting of amylases, maltogenic amylases, β-amylases, aminopeptidases, carboxypeptidases, catalases, cellulolytic enzymes, chitinases, cutinases, cyclodextrin glycosyltransferases, deoxyribonucleases, esterases, glucan 1,4-α-maltotetraosyl hydrolases, glucanases, galactanases, α-galactosidases, β-galactosidases, glucoamylases, glucose oxidases, α-glucosidases, β-glucosidases, haloperoxidases, hemicellulolytic enzymes, invertases, laccases, lipases, mannanases, mannosidases, oxidases, pectinolytic enzymes, peroxidases, phospholipases, phytases, polyphenol oxidases, proteolytic enzymes, ribonucleases, transglutaminases, and xylanases.

[0021] In one embodiment we claim a premix comprising an endopeptidase having at least 60% identity to SEQ ID NO: 1 or SEQ ID NO: 2 and flour.

[0022] In one embodiment, the premix further comprises one or more enzymes selected from the group consisting of amylases, maltogenic amylases, β-amylases, aminopeptidases, carboxypeptidases, catalases, cellulolytic enzymes, chitinases, cutinases, cyclodextrin glycosyltransferases, deoxyribonucleases, esterases, glucan 1,4-α-maltotetraosyl hydrolases, glucanases, galactanases, α-galactosidases, β-galactosidases, glucoamylases, glucose oxidases, α-glucosidases, β-glucosidases, haloperoxidases, hemicellulolytic enzymes, invertases, laccases, lipases, mannanases, mannosidases, oxidases, pectinolytic enzymes, peroxidases, phospholipases, phytases, polyphenol oxidases, proteolytic enzymes, ribonucleases, transglutaminases, and xylanases.

[0023] In one embodiment we claim the use of an endopeptidase having at least 60% identity to SEQ ID NO: 1 or SEQ ID NO: 2 for increasing the extensibility of a dough, wherein no L-cysteine ​​and / or sodium metabisulfite is added to the dough.

[0024] In one embodiment we claim the use of an endopeptidase having at least 60% identity to SEQ ID NO: 1 or SEQ ID NO: 2 for improving the eating quality of an edible product, wherein L-cysteine ​​and / or sodium metabisulfite is not added to the edible product. DETAILED DESCRIPTION

[0025] definition

[0026] Sequence identity: The relatedness between two amino acid sequences or between two nucleotide sequences is described by the parameter "sequence identity".

[0027] For purposes of the present invention, the sequence identity between two amino acid sequences is determined using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, J.Mol.Biol. [Journal of Molecular Biology] 48:443-453) implemented in the Needle program of the EMBOSS package (EMBOSS:The European Molecular Biology Open Software Suite [EMBOSS:European Molecular Biology Open Software Suite], Rice et al., 2000, Trends Genet. [Genetics Trend] 16:276-277) (preferred 5.0.0 version or updated version). The parameters used are gap opening penalty 10, gap extension penalty 0.5 and EBLOSUM62 (EMBOSS version of BLOSUM62) substitution matrix. The output result (obtained using - non-simplified (no brief) option) marked as "highest identity (longest identity)" using Needle is used as the identity percentage, and is calculated as follows:

[0028] (number of identical residues x 100) / (length of alignment - total number of gaps in the alignment)

[0029] Variant: The term "variant" means a polypeptide having endopeptidase activity comprising an alteration (i.e., a substitution, insertion, and / or deletion) at one or more (e.g., several) positions. Substitution means replacing the amino acid occupying a position with a different amino acid; deletion means removing the amino acid occupying a position; and insertion means adding one or more amino acids adjacent to and immediately following the amino acid occupying a position.

[0030] Improved properties: When an endopeptidase according to the invention is incorporated into flour and / or dough in an effective amount, one or more properties are improved compared to flour and / or dough without added enzyme.

[0031] Improved properties can be determined by comparing dough and / or baked products prepared according to the methods described below, with and without the addition of an enzyme of the invention.

[0032] Organoleptic qualities may be assessed using procedures that are well established in the baking industry, and may include, for example, the use of a panel of trained taste testers.

[0033] Improved stretchability: The term "improved stretchability of the dough" is defined herein as the property of the dough that it can withstand increased stretching without breaking.

[0034] Increased stretching is a very important parameter since it means that it is possible, for example, to obtain very thin doughs.

[0035] Increased relaxation: The term "increased relaxation" is defined herein as the property that the endopeptidase according to the invention may be able to replace chemical relaxants for optimizing dough rheology and ensuring that the final product has improved quality, smooth surface and rounded edges.

[0036] Increased softness: The term "increased softness of the dough" is defined herein as the property of a dough that is softer than a control dough (to which no endopeptidase according to the invention has been added), as assessed by a skilled baker.

[0037] Increased elasticity: The term "increased elasticity of the dough" is defined herein as the property of a dough that has a higher tendency to recover its original shape after being subjected to a certain physical stress.

[0038] Increased stability of the dough: The term "increased stability of the dough" is defined herein as the property of a dough that is less susceptible to mechanical damage, thus better retaining its shape and volume, and is assessed by the height:width ratio of a cross-section of a loaf after normal and / or extended proofing.

[0039] Improved mechanical capabilities: The term "improved mechanical capabilities of the dough" is defined herein as properties of a dough that is generally less sticky and / or firmer and / or more elastic.

[0040] Increased volume of dough / baked product: The term "increased volume of dough / baked product" is measured as the volume of the dough or the volume of a given loaf of bread. The volume can be determined, for example, by the rapeseed displacement method, or by a skilled baker, or by using a food volumeter (Volscan profiler 600).

[0041] Improved crumb structure of a baked product: The term "improved crumb structure of a baked product" is defined herein as the properties of a baked product with respect to crumb uniformity, cell wall thickness, and size of individual cell cavities on a slice of bread.

[0042] The crumb structure of baked products is typically assessed visually by the baker or by digital image analysis as known in the art (e.g., C-cell, Calibre Control International Ltd, Appleton, Warrington, UK).

[0043] Improved softness of baked products: The term "improved softness of baked products" is the opposite of "hardness" and is defined herein as that property of a baked product which is more easily compressed and is assessed empirically by a skilled test baker or measured by using a texture analyzer as known in the art (e.g., TAXT2 or TA-XT Plus from Stable Micro Systems Ltd, Surrey, UK).

[0044] Dough composition

[0045] The present invention can solve the problem caused by uneven flour quality, because overly strong flour will cause the dough to not flow properly in the baking pan, resulting in deformation of bread, buns, pizza, etc.

[0046] The present invention can help ensure the relaxation of yeast dough for baked goods such as hamburger buns, yeast baked goods, pizza, pan bread, etc., allowing it to be made with various flour qualities, improving dough rheological properties, dough extensibility, pan flowability of dough, and producing baked goods with attractive appearance, better volume and higher quality pan bread.

[0047] The present invention can solve the problem of flour quality used in biscuit production, help weaken gluten, increase dough extensibility, reduce the energy used for kneading, maintain the shape of laminated dough biscuits, ensure the size and shape of biscuits, improve taste, appearance, etc.

[0048] As used herein, "dough" means any dough used to prepare baked or culinary products.

[0049] According to the present invention, the dough for preparing the baked product or the culinary product can be made from any suitable dough component containing flour. The flour for preparing the dough according to the present invention can be any suitable flour source, for example flour derived from cereals, such as wheat flour, corn flour, rye flour, barley flour, oat flour, rice flour, sorghum flour, potato flour, soy flour, flour from beans and combinations thereof.

[0050] As used herein, "flat dough" means a dough that typically has a thickness of one millimeter to several centimeters.

[0051] According to the invention, the dough can be used to make any baked or cooked product, such as cookies, crackers, pizza, flatbreads, bread, especially pan bread, buns, especially hamburger buns, baguettes, pasta and noodles.

[0052] The dough according to the invention can be used in particular for making biscuits.

[0053] According to the invention, biscuits can be defined as hard dough or short dough:

[0054] Hard dough: This dough is similar to bread dough and has a stiff consistency. It is a lean dough because the fat and sugar content is low relative to the flour content. Semisweet, unsweetened, and salty biscuits are made from lean dough.

[0055] For the brioche dough: This dough is more closely related to cake dough, but has much less water. The name refers to its high level of shortening, or fat, relative to the flour content. This fat reduces the stretchability of the dough, meaning these cookies are more likely to crack. This dough is also high in sugar.

[0056] The biscuits according to the present invention may be any biscuits known in the art.

[0057] An example of a hard dough biscuit is a cracker. Crackers are baked food products made from flaky dough and are typically not sweet. As is known in the art, condiments or seasonings (such as salt, herbs, seeds, and / or cheese) may be added to the dough or sprinkled on the dough before baking. Crackers come in many shapes and sizes - round, square, triangular, etc. If a leavening agent is used, sodium acid pyrophosphate (SAPP) and / or sodium bicarbonate and / or ammonium bicarbonate may be used.

[0058] Flatbreads can be made from a simple mixture of flour, water, and salt, which is then well rolled into a flat dough. Flatbreads have a very fast baking time (usually < 2 minutes).

[0059] Flatbreads can be unleavened (i.e., made without yeast) or leavened (e.g., made with yeast). Flatbreads can further include optional ingredients such as olive oil, sesame oil, shortening, and spices. Examples of flatbreads include tortillas, pita bread, Arabic bread, and Indian flatbreads, including wheat flatbreads and gluten-free flatbreads.

[0060] Additional non-limiting examples of flatbreads include lavash, baladi, barbari, Sangak, tanoor, taftoon, shami, halabi, mafrood, burr, bairuti, pocket bread, naan, phulka, chapatti, paratha, Arabic pita, Lebanese, mafrood, hapati, sangak, roti, taboon, shrak, mashrouh, nasir, tannoor, taftan, and taftan.

[0061] The dough according to the invention can also be used to make pizza. Pizza is a typical yeast flat bread, covered with e.g. tomato sauce and cheese, and baked in an oven.

[0062] The flat dough according to the invention can also be used to make noodles and pasta. Noodles are made from unfermented dough that is stretched, squeezed or flattened and cut into one of various shapes. Noodles are usually cooked in boiling water, sometimes with the addition of edible oil and / or salt. They can be pan-fried or deep-fried. Pasta is typically a noodle made from unfermented dough of durum wheat flour mixed with water and / or eggs and formed into sheets or various shapes and then cooked by boiling. Pasta can also be made from flour from other cereals or grains.

[0063] The dough according to the invention can also be used to make laminated baked products.

[0064] Laminated dough is a culinary preparation consisting of many thin layers of butter-separated dough produced by repeated folding and rolling. Such doughs can contain multiple layers, i.e., more than 10 layers. During baking, the water in the butter evaporates and expands, causing the dough to expand and separate, while the lipids in the butter essentially fry the dough, resulting in a light, flaky product. Examples of laminated doughs include breakfast croissants and other pastries, such as Danish pastry, flaky pastry, and puff pastry.

[0065] It is not usually necessary, but the dough according to the invention may also contain other conventional dough relaxing ingredients, such as deactivated yeast, glutathione, malt, sorbic acid and / or yeast extract.

[0066] Dough according to the present invention can also include one or more emulsifiers. Emulsifiers can be used to improve dough extensibility. Examples of suitable emulsifiers are monoglycerides or diglycerides, polyoxyethylene stearates, diacetyl tartaric acid esters of monoglycerides, sugar esters of fatty acids, propylene glycol esters of fatty acids, polyglycerol esters of fatty acids, lactic acid esters of monoglycerides, acetate esters of monoglycerides, lecithin or phospholipids, or ethoxylated monoglycerides. Special emulsifiers include monoglycerides, diacetyl tartaric acid esters of monoglycerides (DATEM) and sodium stearoyl lactylate (SSL).

[0067] Other conventional ingredients that can be added to the dough include proteins, such as milk powder, gluten and soy; eggs (whole eggs, egg yolks or egg whites); oxidants, such as ascorbic acid, potassium bromide, potassium iodide, azodicarbonamide (ADA), ammonium persulfate or potassium persulfate; sugars, such as sucrose, dextrose, glucose, etc.; salts, such as sodium chloride, calcium acetate, sodium sulfate or calcium sulfate, diluents such as silicon dioxide, starch from different sources. Other conventional ingredients also include hydrocolloids, such as CMC, guar gum, xanthan gum, locust bean gum, etc. Modified starch can also be used.

[0068] The dough according to the invention may be a fibrous dough, for example the dough may contain cereals (e.g. whole wheat) and / or be enriched with additional fiber in the form of, for example, cereal brans (e.g. wheat bran). Wheat bran is produced as a by-product of grinding wheat into white flour.

[0069] Typically, fibers are classified into fine fibers, medium fibers, and coarse fibers as is known in the art. Fine fibers are particularly useful in the present invention.

[0070] In addition to the preparation of fresh dough or fresh dough products, the present invention also relates to a method for preparing frozen dough or frozen dough products.

[0071] The invention is particularly useful for preparing flat doughs and products obtained therefrom in an industrial process, wherein these products are prepared mechanically using automated or semi-automated equipment.

[0072] Enzymes

[0073] Endopeptidase according to the invention

[0074] As used herein, the term endopeptidase is an enzyme that hydrolyzes internal peptide bonds (has endopeptidase activity).

[0075] There is no restriction on the source of the endopeptidase used according to the present invention. Therefore, the term endopeptidase includes not only natural or wild-type endopeptidase, but also any mutant, variant, fragment, etc. thereof exhibiting endopeptidase activity, as well as synthetic endopeptidase, such as shuffled endopeptidase.

[0076] Genetically engineered endopeptidase variants can be prepared as known in the art.Examples of endopeptidase variants are endopeptidases in which one or more amino acids have been deleted, inserted or substituted with other amino acids.

[0077] According to the present invention, the endopeptidase has at least 60% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 61% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 62% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 63% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 64% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 65% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 66% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 67% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 68% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 69% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 70% identity with SEQ ID NO: 1 or SEQ ID NO: 2, such as at least 71% identity with SEQ ID NO: 1 or SEQ ID NO: 2, NO:2 is at least 71% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 72% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 73% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 74% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 75% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 76% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 77% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 78% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 79% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 80% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 81% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 82% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 83% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 84% identical to SEQ ID NO: 1 or SEQ ID NO: 2 NO:1 or SEQ ID NO:2 has at least 83% identity, such as at least 84% identity, such as at least 85% identity, such as at least 86% identity, such as at least 87% identity, such as at least 88% identity, such as at least 89% identity, such as at least 90% identity, such as at least 91% identity, such as at least 92% identity, such as at least 93% identity, such as at least 94% identity, such as at least 95% identity,NO:2 is at least 86% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 87% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 88% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 89% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 90% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 91% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 92% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 93% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 94% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 95% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 96% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 97% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 98% identical to SEQ ID NO: 1 or SEQ ID NO: 2, for example, at least 99% identical to SEQ ID NO: 1 or SEQ ID NO: 2 NO:1 or SEQ ID NO:2 has at least 98% identity, such as at least 99% identity to SEQ ID NO:1 or SEQ ID NO:2.

[0078] In one embodiment, the endopeptidase is SEQ ID NO:1 or SEQ ID NO:2.

[0079] For purposes of the present invention, the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, J.Mol.Biol. [J.Molecular Biology] 48:443-453) implemented in the Needle program of the EMBOSS package (EMBOSS:The European Molecular Biology Open Software Suite [EMBOSS:European Molecular Biology Open Software Suite], Rice et al., 2000, Trends Genet. [Genetics Trend] 16:276-277) (preferred 5.0.0 version or updated version) is used to determine the sequence identity between two amino acid sequences. The parameters used are gap opening penalty 10, gap extension penalty 0.5 and EBLOSUM62 (EMBOSS version of BLOSUM62) substitution matrix. The output result (using-non-simplified option to obtain) marked as "highest identity" using Needle is used as the identity percentage, and is calculated as follows:

[0080] (number of identical residues x 100) / (length of alignment - total number of gaps in the alignment)

[0081] The endopeptidase used according to the present invention may be a microbial endopeptidase, preferably a bacterial endopeptidase, the term bacterial indicating that the endopeptidase is derived from or originates from bacteria, or is an analog, fragment, variant, mutant or synthetic endopeptidase derived from bacteria. The endopeptidase may be produced or expressed in an original wild-type bacterial strain, another microbial strain or a plant; i.e., the term encompasses the expression of a wild-type, naturally occurring endopeptidase, as well as the expression of a recombinant, genetically engineered or synthetic endopeptidase in any host.

[0082] An example of a bacterial endopeptidase suitable for use according to the present invention is the endopeptidase from Nocardiopsis, in particular Nocardiopsis alba (formerly Nocardiopsis dassonvillei) NRRL 18133 disclosed in WO 88 / 03947.

[0083] In the method for the present invention, the endopeptidase can be purified. As used herein, the term "purified" encompasses enzyme protein preparations, wherein the preparation has been enriched with the enzyme protein in question. This enrichment can be, for example, removal of cells from the organism producing the extracellular enzyme protein, removal of non-proteinaceous materials by protein-specific precipitation or using a chromatographic procedure, wherein the enzyme protein in question is selectively adsorbed and eluted from a chromatographic matrix. The endopeptidase may have been purified to a certain extent so that only a small amount of other proteins are present. The statement "other proteins" particularly relates to other enzymes. The endopeptidase used in the method for the present invention can be "substantially pure", that is, substantially free of other components from the organism producing the endopeptidase, which can be a naturally occurring microorganism, or a genetically modified host microorganism for the recombinant production of the endopeptidase.

[0084] The endopeptidase according to the invention may typically be added in an effective amount, such as in the range of 0.1-1000 mg enzyme protein / kg flour, such as 1-500 mg enzyme protein / kg flour, such as 1-200 mg enzyme protein / kg flour.

[0085] Other enzymes

[0086] Optionally, one or more additional enzymes (such as amylases, maltogenic amylases, β-amylases, catalases, cellulolytic enzymes, chitinases, cutinases, cyclodextrin glycosyltransferases, deoxyribonucleases, esterases, glucan 1,4-α-maltotetraosylases, glucanases, galactanases, α-galactosidases, β-galactosidases, glucoamylases, glucose oxidases, α-glucosidases, β-glucosidases, haloperoxidases, hemicellulolytic enzymes, invertases, laccases, lipases, mannanases, mannosidases, oxidases, pectinolytic enzymes, peroxidases, phospholipases, phytases, polyphenol oxidases, proteolytic enzymes, ribonucleases, transglutaminases and xylanases) can be used together with the enzyme composition according to the invention.

[0087] The one or more additional enzymes may be of any origin, including mammalian origin, plant origin, and microbial (bacterial, yeast or fungal) origin.

[0088] Suitable commercial alpha-amylase compositions include, for example, BAKEZYME P 300 (available from DSM) and FUNGAMYL 4000 BG, FUNGAMYL 800 L, FUNGAMYL ULTRA BG and FUNGAMYL ULTRA SG (available from Novozymes A / S).

[0089] The maltogenic alpha-amylase (EC 3.2.1.133) may be from Bacillus. A maltogenic alpha-amylase from B. stearothermophilus strain NCIB 11837 is available from Novozymes under the trade name Commercially available.

[0090] The maltogenic alpha-amylase may also be a variant of a maltogenic alpha-amylase from Bacillus stearothermophilus as disclosed in, e.g., WO 1999 / 043794; WO 2006 / 032281; or WO 2008 / 148845, e.g. 3D.

[0091] The anti-staling amylase used in the present invention may also be an amylase from Pseudomonas saccharophilia (glucan 1,4-α-maltotetraose hydrolase (EC 3.2.1.60)) or a variant thereof, such as any amylase disclosed in WO 1999 / 050399, WO 2004 / 111217 or WO 2005 / 003339.

[0092] Glucoamylases useful in the present invention include A. niger G1 or G2 glucoamylase (Boel et al., (1984), EMBO J. [Journal of the European Molecular Biology Association] 3(5), pp. 1097-1102), A. awamori glucoamylase disclosed in WO 84 / 02921, or A. oryzae glucoamylase (Agric. Biol. Chem. [Agriculture, Biology and Chemistry] (1991), 55(4), pp. 941-949). Suitable commercial glucoamylases are available from Novozymes.

[0093] The glucose oxidase may be a fungal glucose oxidase, in particular Aspergillus niger glucose oxidase (eg Available from Novozymes AB).

[0094] The xylanase may be of microbial origin, e.g. derived from a bacteria or fungus, such as a strain of Aspergillus (particularly A. aculeatus, A. niger, A. awamori or A. tubigensis), from a strain of Trichoderma (e.g. T. reesei), or from a strain of Humicola (e.g. H. insolens).

[0095] Suitable commercially available xylanase preparations for use in the present invention include PANZEA BG, PENTOPAN MONO BG, and PENTOPAN 500 BG (available from Novozymes), GRINDAMYL POWERBAKE (available from Danisco), and BAKEZYME BXP 5000 and BAKEZYME BXP 5001 (available from DSM).

[0096] The phospholipase may have phospholipase A1, A2, B, C, D or lysophospholipase activity; it may or may not have lipase activity. It may be of animal origin, for example from pancreas, snake venom or bee venom, or it may be of microbial origin, for example from filamentous fungi, yeast or bacteria, such as Aspergillus or Fusarium, for example Aspergillus niger, Aspergillus oryzae or F. oxysporum. Preferred lipases / phospholipases from F. oxysporum are disclosed in WO 98 / 26057. Likewise, variants described in WO 00 / 32758 may be used.

[0097] Suitable phospholipase compositions are LIPOPAN F and LIPOPAN XTRA (available from Novozymes) or PANAMORE GOLDEN and PANAMORE SPRING (available from DSM).

[0098] Enzyme treatment

[0099] The endopeptidase according to the invention is added to the dough ingredients, for example indirectly to the dough by adding it to the flour used to prepare the dough, or directly to the dough itself.

[0100] The endopeptidase may be added to the flour or dough in any suitable form, such as, for example, in liquid form, in particular a stabilized liquid, or it may be added to the flour or dough as a substantially dry powder or granules.

[0101] For example, granules may be produced as disclosed in U.S. Pat. No. 4,106,991 and U.S. Pat. No. 4,661,452. Liquid enzyme preparations may be stabilized, for example, by adding sugars or sugar alcohols or lactic acid according to established procedures. Other enzyme stabilizers are well known in the art.

[0102] Premix

[0103] It is often advantageous to provide a mixture of one or more enzymes used in the process of the invention with other ingredients for improving the properties of the dough product. These are often referred to in the art as "premixes" which typically contain flour.

[0104] Therefore, in another aspect, the present invention relates to a premix for improving the quality of dough (for the preparation of one or more flat bread products), the premix comprising an endopeptidase and one or more dough ingredients, in particular flour, for example flour from cereals, such as wheat flour, corn flour, rye flour, barley flour, oat flour, rice flour, or sorghum flour and any combination thereof.

[0105] The premix may further comprise one or more enzymes selected from the group consisting of amylases, maltogenic amylases, β-amylases, catalases, cellulolytic enzymes, chitinases, cutinases, cyclodextrin glycosyltransferases, deoxyribonucleases, esterases, glucan 1,4-α-maltotetraose hydrolases, glucanases, galactanases, α-galactosidases, β-galactosidases, glucoamylases, glucose oxidases, α-glucosidases, β-glucosidases, haloperoxidases, hemicellulolytic enzymes, invertases, laccases, lipases, mannanases, mannosidases, oxidases, pectinolytic enzymes, peptidylglutaminases, peroxidases, phospholipases, phytases, polyphenol oxidases, proteolytic enzymes, ribonucleases, transglutaminases, and xylanases.

[0106] In one embodiment the premix comprises an endopeptidase, a xylanase and / or a phospholipase according to the invention.

[0107] In another embodiment, the present invention relates to a premix comprising an endopeptidase of the present invention and flour, e.g., flour from a cereal such as wheat flour, corn flour, rye flour, barley flour, oat flour, rice flour, sorghum flour, and any combination thereof, and one or more additional enzymes as previously described.

[0108] The premix composition may be in liquid form or in dry or substantially dry form.

[0109] The invention described and claimed herein is not to be limited in scope by the specific embodiments disclosed herein, since these embodiments are intended as illustrative illustrations of several aspects of the invention. Any equivalent embodiments as well as combinations of one or more of these embodiments are intended to be within the scope of the invention.

[0110] Various references are cited herein, the disclosures of which are incorporated herein by reference in their entireties.The present invention is further described by the following examples, which should not be construed as limiting the scope of the invention.

[0111] Examples

[0112] Example 1

[0113] Petit butter cookies beurre Production of biscuits

[0114] Use the following recipe:

[0115] Table 1: Ingredients

[0116]

[0117]

[0118] method:

[0119] Icing sugar, glucose syrup and biscuit margarine were pureed in a Kenwood mixer with a flat beater for 2.30 min.

[0120] Add SAPP 10 and sodium bicarbonate to the above mixture and continue mixing at speed 2 for up to 3 minutes.

[0121] The salt and ammonium bicarbonate (and SMS in the control / reference) were dissolved in 20°C water and then added to the above mixture, after which mixing was continued at minimum speed for up to 4 minutes.

[0122] Add the flour plus enzyme according to the invention and blend at minimum speed for 5 minutes, scrape out, and continue mixing at speed 1 for 6 minutes.

[0123] Finally, use high speed for 2.30 minutes.

[0124] The dough was allowed to rest in a covered condition in a 40°C / 85%RH chamber for 15 min.

[0125] After resting, the dough was sheeted by forming 8 layers to a final sheet thickness of 1.75 mm.

[0126] Mark the cookie sheet with a Dough Roller Docker and cut at the specified weight and bake at 210°C-220°C for 8±1 min.

[0127] The enzyme according to the invention:

[0128] A: Control / reference: 0 mg enzyme. 100 ppm sodium metabisulfite (SMS)

[0129] B: 20 ​​mg SEQ ID NO: 1 enzyme protein / kg flour. 0 ppm sodium metabisulfite (SMS)

[0130] result:

[0131] The sensory test was conducted by 15 people and took into account appearance, crunchiness, taste and taste preference.

[0132] Table 2: Sensory test results

[0133]

[0134] The results of the sensory tests show that the eating properties are particularly improved by using the enzyme solution according to the invention compared to the use of sodium metabisulfite.

Claims

1. A method for increasing the softness and relaxation of dough, the method comprising: a) adding at least one endopeptidase having at least 99% identity to SEQ ID NO: 1 to flour or to a dough comprising flour, wherein no sodium metabisulfite is added to the dough; b) kneading the dough; and c) forming the dough into an edible product.

2. The method according to claim 1, wherein no L-cysteine ​​is added to the dough.

3. The method of claim 1, wherein the edible product is selected from the group consisting of bread, noodles, laminated bakery products, cookies and pizza.

4. The method according to any one of claims 1 to 3, wherein the dough is formed into biscuits.

5. The method according to any one of claims 1-3, wherein the flour is selected from the group consisting of wheat flour, corn flour, rye flour, barley flour, oat flour, rice flour, sorghum flour and combinations thereof.

6. The method according to any one of claims 1-3, wherein the amount of energy used to knead the dough is reduced compared to a method wherein no endopeptidase having at least 99% identity to SEQ ID NO: 1 is added to the dough.

7. The method according to any one of claims 1-3, wherein the shape of the edible product is better maintained compared to a method wherein no endopeptidase having at least 99% identity to SEQ ID NO: 1 is added to the dough.

8. The method according to any one of claims 1 to 3, wherein the endopeptidase is added in an amount ranging from 0.1 to 1000 mg enzyme protein / kg flour.

9. The method according to any one of claims 1 to 3, wherein the extensibility of the dough is better than the extensibility of a dough prepared under the same conditions but not treated with the endopeptidase.

10. The method according to any one of claims 1-3, wherein the eating quality of the edible product is better than the eating quality of an edible product prepared under the same conditions but not treated with the endopeptidase.

11. The method according to any one of claims 1-3, wherein the dough further comprises one or more enzymes selected from the group consisting of: Amylase, aminopeptidase, carboxypeptidase, cellulolytic enzyme, chitinase, cutinase, cyclodextrin glycosyltransferase, deoxyribonuclease, esterase, glucan 1,4-α-maltotetraose hydrolase, glucanase, galactanase, α-galactosidase, β-galactosidase, α-glucosidase, β-glucosidase, haloperoxidase, hemicellulolytic enzyme, invertase, laccase, lipase, mannanase, mannosidase, oxidase, pectinolytic enzyme, peptidoglutaminase, peroxidase, phospholipase, phytase, proteolytic enzyme, ribonuclease, transglutaminase and xylanase.

12. The method of claim 3, wherein the edible product is selected from the group consisting of: flatbreads, buns, crackers, pasta, and baguettes.

13. The method according to claim 11, wherein the dough further comprises one or more enzymes selected from the group consisting of: Produces maltogenic amylase, β-amylase, catalase, glucoamylase, glucose oxidase and polyphenol oxidase.

14. A premix comprising at least one endopeptidase having at least 99% identity to SEQ ID NO: 1 and flour, wherein the premix does not comprise sodium metabisulfite.

15. The premix according to claim 14, wherein the premix does not contain L-cysteine.

16. The premix according to claim 14, wherein the premix further comprises one or more enzymes selected from the group consisting of: Amylase, aminopeptidase, carboxypeptidase, cellulolytic enzyme, chitinase, cutinase, cyclodextrin glycosyltransferase, deoxyribonuclease, esterase, glucan 1,4-α-maltotetraose hydrolase, glucanase, galactanase, α-galactosidase, β-galactosidase, α-glucosidase, β-glucosidase, haloperoxidase, hemicellulolytic enzyme, invertase, laccase, lipase, mannanase, mannosidase, oxidase, pectinolytic enzyme, peroxidase, phospholipase, phytase, proteolytic enzyme, ribonuclease, transglutaminase and xylanase.

17. The premix according to claim 16, wherein the premix further comprises one or more enzymes selected from the group consisting of: Produces maltogenic amylase, β-amylase, catalase, glucoamylase, glucose oxidase and polyphenol oxidase.

18. Use of an endopeptidase having at least 99% identity to SEQ ID NO: 1 for increasing the extensibility of dough, wherein no sodium metabisulfite is added to the dough.

19. Use according to claim 18, wherein no L-cysteine ​​is added to the dough.

20. Use of an endopeptidase having at least 99% identity to SEQ ID NO: 1 for improving the eating quality of an edible product, wherein no sodium metabisulfite is added to the edible product, wherein the edible product is a biscuit and the eating quality is one or more qualities selected from hardness, crispness and crunchiness.

21. Use according to claim 20, wherein no L-cysteine ​​is added to the edible product.

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