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Use of a Pyranose Oxidase in Baking Verwendung Von Pyranose-Oxidase Beim Backen Utilisation D’Une Pyranose-Oxydase En Boulangerie

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Use of a Pyranose Oxidase in Baking Verwendung Von Pyranose-Oxidase Beim Backen Utilisation D’Une Pyranose-Oxydase En Boulangerie Europäisches Patentamt *EP000869716B1* (19) European Patent Office Office européen des brevets (11) EP 0 869 716 B1 (12) EUROPEAN PATENT SPECIFICATION (45) Date of publication and mention (51) Int Cl.7: A21D 8/04 of the grant of the patent: 02.05.2001 Bulletin 2001/18 (86) International application number: PCT/DK96/00550 (21) Application number: 96943883.7 (87) International publication number: (22) Date of filing: 20.12.1996 WO 97/22257 (26.06.1997 Gazette 1997/27) (54) USE OF A PYRANOSE OXIDASE IN BAKING VERWENDUNG VON PYRANOSE-OXIDASE BEIM BACKEN UTILISATION D’UNE PYRANOSE-OXYDASE EN BOULANGERIE (84) Designated Contracting States: (72) Inventors: AT BE CH DE DK ES FI FR GB GR IE IT LI NL PT • WAGNER, Peter SE DK-2880 Bagsvaerd (DK) • SI, Joan, Qi (30) Priority: 20.12.1995 DK 144795 DK-2880 Bagsvaerd (DK) (43) Date of publication of application: (56) References cited: 14.10.1998 Bulletin 1998/42 EP-A- 0 321 811 WO-A-94/28728 WO-A-95/23515 (73) Proprietor: Novozymes A/S 2880 Bagsvaerd (DK) Note: Within nine months from the publication of the mention of the grant of the European patent, any person may give notice to the European Patent Office of opposition to the European patent granted. Notice of opposition shall be filed in a written reasoned statement. It shall not be deemed to have been filed until the opposition fee has been paid. (Art. 99(1) European Patent Convention). EP 0 869 716 B1 Printed by Jouve, 75001 PARIS (FR) EP 0 869 716 B1 Description FIELD OF THE INVENTION 5 [0001] The present invention relates to a method of preparing a dough and/or a baked product by. use of the com- position. BACKGROUND OF THE INVENTION 10 [0002] In the bread-making process it is known to add bread-improving and/or dough-improving additives to the bread dough, the action of which, inter alia, results in improved texture, volume, flavour and freshness of the bread as well as improved machinability of the dough. [0003] In recent years a number of enzymes have been used as dough and/or bread improving agents, in particular, enzymes which act on components present in large amounts in the dough. Examples of such enzymes are found within 15 the groups of amylases, proteases, glucose oxidases and (hemi)cellulases, including pentosanases. [0004] EP 321 811 and EP 338 452 disclose the use, in baking, of glucose oxidase in combination with other enzymes (sulfhydryl oxidase, hemicellulase, cellulase). Glucose oxidase catalyzes the oxidation of β-D-glucose to produce D- glucono-1,5-lactone and hydrogen peroxide. [0005] Pyranose oxidases are enzymes which catalyse the oxidation of several monosaccharides at position C2 with 20 the concomitant release of hydrogen peroxide. Glucose, in its pyranose form, tends to be the preferred substrate. However, a number of other substrates, e.g., furanoses such as xylose, can also be oxidised by the enzyme. The reaction product of glucose oxidation is 2-keto-glucose, and of xylose oxidation, 2-keto-xylose. The pyranose oxidase is distinct from glucose oxidase (E.C.1.1.34) which catalyses the oxidation of glucose at position C1 with the concom- itant formation of gluconic acid. 25 [0006] Pyranose oxidases are of widespread occurrence, but in particular, in Basidiomycete fungi. Pyranose oxidases have been characterised or isolated, e.g., from the following sources: Peniophora gigantea (Huwig et al., 1994, Journal of Biotechnology 32, 309-315; Huwig et el., 1992, Med. Fac. Landbouww, Univ. Gent, 57/4a, 1749-1753; Danneel et al., 1993, Eur. J. Biochem. 214, 795-802), genera belonging to the Aphyllophorales (Volc et al., 1985, Folia Microbiol. 30, 141-147), Phanerochaete chrysosporium (Volc et al., 1991, Arch. Mirobiol. 156, 297-301, Volc and Eriksson, 1988, 30 Methods Enzymol 161B, 316-322), Polyporus pinsitus (Ruelius et al., 1968, Biochim. Biophys. Acta, 167, 493-500) and Bierkandera adusta and Phlebiopsis gigantea (Huwig et al., 1992, op. cit.) [0007] The use, in baking, of a pyranose oxidase has never been suggested before. [0008] It is the object of the present invention to improve the properties of dough and/or baked products by the use of a pyranose oxidase enzyme. 35 BRIEF DISCLOSURE OF THE INVENTION [0009] Accordingly, in a first aspect the present invention relates to a bread-improving and/or a dough-improving composition comprising a pyranose oxidase enzyme and a second component which is another enzyme, a milk powder, 40 gluten, an emulsifier, granulated fat, an oxidant, an amino acid or a sugar. [0010] In the present context the terms "bread-improving composition" and "dough-improving composition" are in- tended to indicate compositions which, in addition to the enzyme component, may comprise other substances con- ventionally used in baking to improve the properties of dough and/or baked products. Examples of such components are given below. 45 [0011] Without being limited to any theory, it is presently contemplated that when a pyranose oxidase enzyme is added to dough intended for use in the preparation of baked products, it may exert an oxidizing effect on dough con- stituents and thereby serve, e.g., to improve the strength of gluten structures in the dough and/or baked products and thereby improve the strength of the dough in addition to the rheological and the handling properties of the dough. [0012] More specifically, the oxidizing effect is believed to be achieved when the enzyme catalyses oxidation of 50 monosaccharides present in the flour or dough with the concomitant formation of hydrogen peroxide. [0013] In the present context the term "improved properties" is intended to indicate any property which may be im- proved by the action of a pyranose oxidase enzyme. In addition to, or as a consequence of, the gluten strengthening effect mentioned above, the use of a pyranose oxidase may result in an increased volume and an improved crumb structure and softness of the baked product, as well as an increased strength, stability and reduced stickiness of the 55 dough, thus resulting in improved machinability. The effect on the dough may be particularly advantageous when a poor quality flour is used. The improved machinability is of particular importance in connection with dough which is to be processed industrially. [0014] The improved properties are evaluated by comparison with dough and/or baked products prepared without 2 EP 0 869 716 B1 addition of a pyranose oxidase in accordance with the present invention. [0015] In a second aspect, the present invention relates to a method of preparing a dough and/or a baked product prepared from the dough comprising adding a pyranose oxidase enzyme, optionally present in a bread-improving or dough-improving composition, to the dough and/or to any ingredient of the dough and/or to any mixture of the dough 5 ingredients. [0016] In further aspects, the present invention relates to a dough and a pre-mix comprising a pyranose oxidase enzyme. In the present context, the term "pre-mix" is intended to be understood in its conventional meaning, i.e. as a mix of baking agents including flour, which may be used not only in industrial bread-baking plants/facilities, but also in retail bakeries. 10 [0017] In final aspects, the present invention relates to the use of a pyranose oxidase enzyme as a dough-improving or bread-improving agent and use of a pyranose oxidase enzyme in the preparation of pasta. DETAILED DISCLOSURE OF THE INVENTION 15 [0018] As mentioned in the Background of the Invention section above pyranose oxidases have been found to be of widespread occurrence, in particular among Basidomycete fungi. It is contemplated that the origin of the pyranose oxidase to be used in the present invention is not critical with a view to exerting its dough and/or baked product improving capability. Accordingly, the pyranose oxidase enzyme of the dough- and/or bread-improving composition of the inven- tion may be of any origin. 20 [0019] The pyranose oxidase may be derived, e.g., from a microbial source, such as a fungus, e.g., a filamentous fungus or a yeast, in particular a Basidomycete fungus. The pyranose oxidase may be derived from genera belonging to Agaricales, such as Oudemansiella or Mycena, to Aphyllophorales, such as Trametes, e.g. T. hirsuta, T. versicolor T. gibbosa, T. suaveolens, T. ochracea, T. pubescens,ortoPhanerochaete, Lenzites or Peniophora, and may be any of the specific pyranose oxidases described in the references cited in the Background of the Invention section above. 25 [0020] The pyranose oxidase may be obtained from the organism in question by use of any suitable technique. For instance, a pyranose oxidase preparation may be obtained by use of recombinant DNA techniques as known in the art. Such method normally comprises cultivation of a host cell transformed with a recombinant DNA vector capable of expressing and carrying a DNA sequence encoding the pyranose oxidase in question, in a culture medium under conditions permitting the expression of the enzyme and recovering the enzyme from the culture. The DNA sequence 30 may be of genomic, cDNA or synthetic origin, or any mixture of these, and may be isolated or synthesized in accordance with methods known in the art. The pyranose enzyme may also be extracted from the organism, or relevant part thereof, by which it is produced in nature. [0021] The pyranose oxidase enzyme to be included in the bread-improving and/or dough-improving composition of the invention may be in any form suited for the use in question, e.g. in the form of a dry powder or granulate, in particular 35 a non-dusting granulate, a liquid, in particular a stabilized liquid, or a protected enzyme.
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