Search Images Maps Play YouTube News Gmail Drive More »
Sign in
Screen reader users: click this link for accessible mode. Accessible mode has the same essential features but works better with your reader.

Patents

  1. Advanced Patent Search
Publication numberUSRE43341 E1
Publication typeGrant
Application numberUS 11/896,132
Publication dateMay 1, 2012
Filing dateAug 29, 2007
Priority dateJun 7, 1995
Fee statusPaid
Publication number11896132, 896132, US RE43341 E1, US RE43341E1, US-E1-RE43341, USRE43341 E1, USRE43341E1
InventorsTorkil Steenholt Olsen, Inge Lise Povlsen, Jorn Borch Soe, Charlotte Horsmans Poulsen, Pernille Bak Hostrup
Original AssigneeDanisco A/S
Export CitationBiBTeX, EndNote, RefMan
External Links: USPTO, USPTO Assignment, Espacenet
Method of improving the properties of a flour dough, a flour dough improving composition and improved food products
US RE43341 E1
Abstract
A method of improving the rheological and/or machineability properties of a flour dough and/or the quality of the product made from the dough, comprising adding to the dough a combination comprising a Hox and an emulsifying agent.
Images(3)
Previous page
Next page
Claims(56)
1. A method of improving the rheological and/or machineability properties of a flour dough and/or the quality of the product made from the dough, comprising adding to the dough a combination comprising a hexose oxidase and an emulsifying agent, wherein the emulsifying agent is a lipase.
2. A method according to claim 1 wherein the lipase comprises a triacylglycerol lipase, a galactolipase, or a phospholipase.
3. A method according to claim 1 wherein the hexose oxidase is isolated from a red algae.
4. A method according to claim 1 wherein the flour dough comprises flour, water and at least one further dough additive or ingredient.
5. A method according to claim 1 wherein the flour dough comprises flour, water and at least one further dough additive or ingredient and wherein the further dough additive or ingredient is selected from the group consisting of a vegetable oil, a vegetable fat, an animal fat, shortening, butterfat, glycerol, milk fat and a mixture thereof.
6. A method according to claim 1 wherein the flour dough comprises a hard flour.
7. A method according to claim 1 wherein the product is a bread product.
8. A method according to claim 1 wherein at least one further enzyme is added to the dough.
9. A method according to claim 1 wherein at least one further enzyme is added to the dough, and wherein the further enzyme comprises a xylanase, a cellulase, a hemicellulase, a starch degrading enzyme, a protease, a lipoxygenase, an oxidoreductase or a lipase.
10. A dough improving composition comprising a hexose oxidase and an emulsifying agent, wherein the emulsifying agent is a lipase.
11. A dough improving composition according to claim 10 wherein the lipase comprises a triacylglycerol lipase, a galactolipase, or a phospholipase.
12. A dough improving composition according to claim 10 wherein the lipase comprises a triacylglycerol lipase, a galactolipase, or a phospholipase and wherein the hexose oxidase is isolated from red algae.
13. A dough improving composition according to claim 10 wherein the dough improving composition comprises at least one further dough additive or ingredient.
14. A dough improving composition according to claim 10 wherein the dough improving composition comprises at least one further dough additive or ingredient and wherein the further dough additive or ingredient comprises a vegetable oil, a vegetable fat, an animal fat, shortening, butterfat, glycerol or milk fat.
15. A dough improving composition according to claim 10 wherein the dough improving composition comprises at least one further dough additive or ingredient and wherein the further dough additive or ingredient is a hard wheat flour.
16. A method of preparing a bread product comprising adding a dough improving composition according to claim 10 to dough ingredients, dough additives or a dough and baking the dough comprising the dough improving composition to obtain the bread product.
17. A dough improving composition according to claim 10 wherein at least one further enzyme is added to the dough improving composition.
18. A dough improving composition according to claim 9 wherein at least one further enzyme is added to the dough improving composition and wherein the further enzyme comprises a xylanase, a cellulase, a hemicellulase, a starch degrading enzyme, a protease, a lipoxygenase, an oxidoreductase or a lipase.
19. A method of improving the rheological and/or machineability properties of a flour dough comprising adding to the dough a dough improving composition of claim 10.
20. A method of improving the volume or a baked product made from a flour dough comprising adding to the dough a dough improving composition of claim 10.
21. A method of improving the rheological and/or machineability properties of a flour dough and/or the quality of the product made from the dough, comprising adding to the dough a combination comprising a hexose oxidase and a triacylglycerol lipase.
22. A method of improving the rheological and/or machineability properties of a flour dough and/or the quality of the product made from the dough, comprising adding to the dough a combination comprising a hexose oxidase and a galactolipase.
23. A method of improving the rheological and/or machineability properties of a flour dough and/or the quality of the product made from the dough, comprising adding to the dough a combination comprising a hexose oxidase and a phospholipase.
24. A method according to claim 1 wherein at least one further enzyme is added to the dough and wherein the further enzyme comprises a xylanase, an amylase or a mixture of a xylanase and an amylase.
25. A dough improving composition according to claim 1 wherein the hexose oxidase is isolated from red algae and wherein the red algae comprises Iridophycus flaccidum, Chondrus crispus, or Euthora cristata.
26. A dough improving composition of claim 10 wherein at least one further enzyme is added to the dough improving composition and wherein the further enzyme comprises a xylanase, an amylase or a mixture of a xylanase and an amylase.
27. A method according to claim 1, wherein the hexose oxidase is isolated from a red algae and wherein the red algae comprises Iridophycus flaccidum, Chondrus crispus or Euthora cristata.
28. A method of improving the rheological and/or machineability properties of a flour dough and/or the quality of the product made from the dough, comprising adding to the dough a combination comprising a hexose oxidase and an emulsifying agent; wherein said flour dough comprises flour, water and at least one further dough additive or ingredient; wherein said further dough additive or ingredient is selected from the group consisting of a vegetable oil, a vegetable fat, an animal fat, shortening, butterfat, glycerol, milk fat and a mixture thereof and wherein said further dough additive or ingredient is present in an amount from 1 to 5% by the weight of the flour component of the dough.
29. A method according to claim 28 wherein the emulsifying agent is a lipase.
30. A method according to claim 28 wherein the emulsifying agent is a lipase and wherein the lipase comprises a triacylglycerol lipase, a galactolipase, or a phospholipase.
31. A method according to claim 28 wherein the hexose oxidase is isolated from red algae.
32. A method according to claim 28 wherein the flour dough comprises at least one further dough additive or ingredient.
33. A method according to claim 28 wherein the flour dough comprises at least one further dough additive or ingredient and wherein the further dough additive or ingredient is a hard flour.
34. A method according to claim 28 wherein the product is a bread product.
35. A method according to claim 28 wherein at least one further enzyme is added to the dough.
36. A method according to claim 28 wherein at least one further enzyme is added to the dough and wherein the further enzyme is selected from the group consisting of a xylanase, a cellulase, a hemicellulase, a starch degrading enzyme, a protease, a lipoxygenase, an oxidoreductase, a lipase and a mixture thereof.
37. A dough comprising a dough improving composition wherein said dough improving composition comprises a hexose oxidase, an emulsifying agent and a further dough additive or ingredient; wherein said dough comprises flour and water; wherein said further dough additive or ingredient is selected from the group consisting of a vegetable oil, a vegetable fat, an animal fat, shortening, butterfat, glycerol, milk fat and a mixture thereof and wherein said further dough additive or ingredient is present in an amount of from 1 to 5% by weight of the flour component of the dough.
38. A dough according to claim 37 wherein the emulsifying agent is a lipase.
39. A dough according to claim 37 wherein the emulsifying agent is a lipase and wherein the lipase comprises a triacylglycerol lipase, a galactolipase, or a phospholipase.
40. A dough according to claim 37 wherein the hexose oxidase is isolated from red algae.
41. A dough according to claim 37 wherein the dough improving composition comprises at least one further dough additive or ingredient.
42. A dough according to claim 37 wherein the dough improving composition comprises at least one further dough additive or ingredient and wherein the further dough additive or ingredient is a hard wheat flour.
43. A dough according to claim 37 wherein at least one further enzyme is added to the dough ingredients, dough additives or the dough.
44. A dough according to claim 37 wherein at least one further enzyme is added to the dough ingredients, dough additives or the dough and wherein the further enzyme is selected from the group consisting of a xylanase, an amylase, a cellulase, a hemicellulase, a starch degrading enzyme, a protease, a lipoxygenase, an oxidoreductase, a lipase, and a mixture thereof.
45. A dough according to claim 37 wherein at least one further enzyme is added to the dough ingredients, dough additives or the dough and wherein the further enzyme includes a xylanase, an amylase or a mixture thereof.
46. A method for improving a flour dough or the quality of a product made from dough, said method comprising adding to the dough a combination comprising a hexose oxidase isolated from a red algae and an emulsifying agent.
47. The method of claim 46 wherein the red algae is Iridophycus flaccidum.
48. The method of claim 46 wherein the red algae is Chondrus crispus.
49. The method of claim 46 wherein the red algae is Euthora cristata.
50. A dough improving composition comprising a hexose oxidase isolated from a red algae and an emulsifying agent.
51. The dough improving composition of claim 50 wherein the red algae is Iridophycus flaccidum.
52. The dough improving composition of claim 50 wherein the red algae is Chondrus crispus.
53. The dough improving composition of claim 50 wherein the red algae is Euthora cristata.
54. A method for improving a flour dough or the quality of a product made from dough, said method comprising adding to the dough a combination comprising a hexose oxidase and an emulsifying agent comprising an enzyme.
55. The method of claim 54 wherein the enzyme is a lipase.
56. A dough improving composition comprising a hexose oxidase and an emulsifying agent comprising an enzyme.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of priority of the filing date of U.S. Provisional Patent Application No. 60/398,020, filed Jul. 24, 2002. This application is also a continuation-in-part of U.S. patent application Ser. No. 09/932,923, filed Aug. 21, 2001, now U.S. Pat. No. 6,726,942, which is a continuation of application Ser. No. 08/676,186 filed Sep. 12, 1996, now U.S. Pat. No. 6,358,543, which was a continuation-in-part of application Ser. No. 08/483,870, filed Jun. 7, 1995, abandoned. The Ser. No. 08/483,870 application was a U.S. national phase of PCT/DK96/00239, filed Jun. 4, 1996. This application is additionally a continuation-in-part of application Ser. No. 10/040,394, filed Jan. 9, 2002, now U.S. Pat. No. 6,852,346, which was a divisional of application Ser. No. 09/402,664, filed Oct. 22, 1999, now U.S. Pat. No. 6,406,723. U.S. Pat. No. 6,406,723 was the U.S. national phase of PCT/DK98/00136, filed Apr. 3, 1998, which claimed priority from DK0400/97, filed Apr. 9, 1997. This application is also a continuation-in-part of U.S. application Ser. No. 10/150,429, filed May 17, 2002, now U.S. Pat. No. 6,967,035, which claims priority from UK Application 0112226.6, filed May 18, 2001, and U.S. Application No. 60/347,007, filed Jan. 9, 2002. We claim the benefit of priority dates of all the above applications. This application also claims the benefit of priority of the filing date of Great Britain Application 0211975.8, filed May 24, 2002. The contents of all of the above applications are incorporated herein by reference to the extent they are consistent with this application and inventions described herein.

FIELD OF INVENTION

The invention pertains to the provision of flour doughs having improved rheological properties and farinaceous food products having improved quality characteristics and it provides a maltose oxidizing oxidoreductase-containing compo-sition capable of conferring such improved properties on doughs and finished food products made herefrom when it is added as a component to the doughs, and a method of preparing improved doughs and farinaceous food products.

More in particular, the present invention relates to the field of food manufacturing, in particular to the preparation of improved bakery products and other farinaceous food products. Specifically, the invention concerns the use of a new combination for improving the stability and/or machineability of dough and/or improving the quality of baked and dried products made from such doughs.

In a preferred aspect, the present invention relates to:

    • a method of improving the rheological and/or machineability properties of a flour dough and/or the quality of the product made from the dough, comprising adding to the dough a combination comprising a Hox and an emulsifying agent.

Teachings relating to this preferred aspect now follow.

TECHNICAL BACKGROUND AND PRIOR ART

The invention relates in particular to a method of providing flour doughs having improved rheological properties and to finished baked or dried products made from such doughs, which have improved textural, eating quality and dimensional characteristics.

In this connection, the “strength” or “weakness” of doughs is an important aspect of making farinaceous finished products from doughs, including baking. The “strength” or “weakness” of a dough is primarily determined by its content of protein and in particular the content and the quality of the gluten protein is an important factor in that respect. Flours with a low protein content are generally characterized as “weak”. Thus, the cohesive, extensible, rubbery mass which is formed by mixing water and weak flour will usually be highly extensible when subjected to stress, but it will not return to its original dimensions when the stress is removed.

Flours with a high protein content are generally characterized as “strong” flours and the mass formed by mixing such a flour and water will be less extensible than the mass formed from a weak flour, and stress which is applied during mixing will be restored without breakdown to a greater extent than is the case with a dough mass formed from a weak flour.

Strong flour is generally preferred in most baking contexts because of the superior rheological and handling properties of the dough and the superior form and texture qualities of the finished baked or dried products made from the strong flour dough.

Dough quality may be largely dependent on the type or types of flour present in the dough and/or the age of the flour or flours.

Doughs made from strong flours are generally more stable. Stability of a dough is one of the most important characteristics of flour doughs. According to American Association of Cereal Chemists (AACC) Method 36-01A the term “stability” can be defined as “the range of dough time over which a positive response is obtained and that property of a rounded dough by which it resists flattening under its own weight over a course of time”. According to the same method, the term “response” is defined as “the reaction of dough to a known and specific stimulus, substance or set of conditions, usually determined by baking it in comparison with a control”

Within the bakery and milling industries it is known to use dough “conditioners” to strengthen the dough to increase its stability and strength. Such dough conditioners are normally non-specific oxidizing agents such as eg iodates, peroxides, ascorbic acid, K-bromate or azodi-carbonamide and they are added to dough with the aims of improving the baking performance of flour to achieve a dough with improved stretchability and thus having a desirable strength and stability. The mechanism behind this effect of oxidizing agents is that the flour proteins, in particular gluten contains thiol groups which, when they become oxidized, form disulphide bonds whereby the protein forms a more stable matrix resulting in a better dough quality and improvements of the volume and crumb structure of the baked products.

In addition to the above usefulness of ascorbic acid/ascorbate as a dough conditioner due to its oxidizing capacity, these compounds may also act as substrate for an oxidoreductase, ascorbate oxidase which is disclosed in EP 0 682 116 A1. In the presence of its substrate, this enzyme converts ascorbic acid/ascorbate to dehydroascorbic acid and H2O2. This prior art does not suggest that ascorbic acid oxidase in the presence of ascorbic acid/ascorbate might have a dough conditioning effect, but assumingly this is the case.

However, the use of several of the currently available oxidizing agents is either objected to by consumers or is not permitted by regulatory bodies and accordingly, it has been attempted to find alternatives to these conventional flour and dough additives and the prior art has i.a. suggested the use of glucose oxidase for this purpose. In addition, the prior art has inter alia (i.a.) suggested the use of oxidoreductases such as carbohydrate oxidase, glycerol oxidase and hexose oxidase for this purpose.

Thus, U.S. Pat. No. 2,783,150 discloses the addition of glucose oxidase to flour to improve dough strength and texture and appearance of baked bread.

CA 2,012,723 discloses bread improving compositions comprising cellulolytic enzymes such as xylanases and glucose oxidase, the latter enzyme being added to reduce certain disadvantageous effects of the cellulolytic enzymes (reduced dough strength and stickiness) and it is disclosed that addition of glucose to the dough is required to obtain a sufficient glucose oxidase activity.

JP-A-92-084848 suggests the use of a bread improving composition comprising glucose oxidase and lipase.

EP-B1-321 811 discloses the use of an enzyme composition comprising sulfhydryl oxidase and glucose oxidase to improve the rheological characteristics of doughs. It is mentioned in this prior art document that the use of glucose oxidase alone has not been successful.

In EP-B1-338 452 is disclosed an enzyme composition for improving dough stability, comprising a mixture of cellulase/hemicellulase, glucose oxidase and optionally sulfhydryl oxidase.

However, the use of glucose oxidase as a dough improving additive has the limitation that this enzyme requires the presence of sufficient amounts of glucose as substrate in order to be effective in a dough system and generally, the glucose content in cereal flours is low. Therefore, the absence of glucose in doughs or the low content hereof in doughs will be a limiting factor for the effectiveness of glucose oxidase as a dough improving agent.

In contrast hereto, the content of maltose in cereal flours is generally significantly higher than that of glucose and accordingly, a freshly prepared dough will normally contain more maltose than glucose. Thus, in an experiment where the content of sugars in supernatants from suspensions of wheat flour and a dough prepared from the flour and further comprising water, yeast, salt and sucrose (as described in the following example 2.3) were analyzed, the following values (% by weight calculated on flour) were found:

Flour Dough
Sucrose 0.3 <0.01
Galactose 0.001 0.01
Glucose 0.25 0.72
Maltose 2.6 1.4
Fructose 0.06 0.67
Lactose <0.01 <0.01

In addition, the content of maltose remains at a relatively high level in a dough which is leavened by yeast, since the yeast primarily utilizes glucose, or it may even increase in the dough e.g. during proofing due to the activity of starch degrading enzymes such as e.g. β-amylase, which is inherently present in the flour or is added to the dough.

Whereas the prior art has recognized the useful improving effects of glucose oxidase on the rheological characteristics of bread doughs and on the quality of the corresponding baked products, it has also been realized that the use of this enzyme has several drawbacks. Thus, it may be required to add sucrose or glucose as substrate to the dough to obtain a sufficient effect and glucose oxidase does not constantly provide a desired dough or bread improving effect when used alone without the addition of other enzymes.

However, it has now been found that the addition of an oxidoreductase, which is capable of oxidizing maltose, including hexose oxidase as a sole dough conditioning agent, i.e. without concomitant addition of substrate for the added enzyme, or of other enzymes, to a farinaceous dough results in an increased resistance hereof to breaking when the dough is stretched, i.e. this enzyme confers in itself to the dough an increased strength whereby the dough becomes less prone to mechanical deformation. It is contemplated that this effect of addition of hexose oxidase to a dough is the result of the formation of cross-links between thiol groups in sulphur-containing amino acids in wheat gluten which occurs when the H2O2 generated by the enzyme in the dough reacts with the thiol groups which are hereby oxidized.

Hexose oxidase (D-hexose: O2-oxidoreductase, EC 1.1.3.5) is an enzyme which in the presence of oxygen is capable of oxidizing D-glucose and several other reducing sugars including maltose, glucose, lactose, galactose, xylose, arabinose and cellobiose to their corresponding lactones with subsequent hydrolysis to the respective aldobionic acids. Accordingly, hexose oxidases differ from glucose oxidase which can only convert D-glucose, in that hexose oxidases can utilize a broader range of sugar substrates. The oxidation catalyzed by the enzyme can be illustrated as follows:

D-Glucose+O2->δ-D-gluconolactone+H2O2, or

D-Galactose+O2->γ-D-galactogalactone+H2O2

Hexose oxidase (in the following also referred to as HOX) has been isolated from several red algal species such as Iridophycus flaccidum (Bean and Hassid, 1956, J. Biol. Chem., 218:425-436) and Chondrus crispus (Ikawa 1982, Methods Enzymol., 89:145-149). Additionally, the algal species Euthora cristata (Sullivan et al. 1973, Biochemica et Biophysica Acta, 309:11-22) has been shown to produce HOX.

Other potential sources of hexose oxidase according to the invention include microbial species or land-growing plant species. Thus, as an example of such a plant source, Bean et al., Journal of Biological Chemistry (1961) 236: 1235-1240, have disclosed an oxidoreductase from citrus fruits which is capable of oxidizing a broad range of sugars including D-glucose, D-galactose, cellobiose, lactose, maltose, D-2-deoxyglucose, D-mannose, D-glucosamine and D-xylose. Another example of an enzyme having hexose oxidase activity is the enzyme system of Malleomyces mallei disclosed by Dowling et al., Journal of Bacteriology (1956) 72:555-560.

It has been reported that hexose oxidase isolated from the above natural sources may be of potential use in the manufacturing of certain food products. Thus, hexose oxidase isolated from Iridophycus flaccidum has been shown to be capable of converting lactose in milk with the production of the corresponding aldobionic acid and has been shown to be of potential interest as an acidifying agent in milk, e.g. to replace acidifying microbial cultures for that purpose (Rand, 1972, Journal of Food Science, 37:698-701). In that respect, hexose oxidase has been mentioned as a more interesting enzyme than glucose oxidase, since this latter enzyme can only be enzymatically effective in milk or other food products not containing glucose or having a low content of glucose, if glucose or the lactose-degrading enzyme lactase which convert the lactose to glucose and galactose, is also added.

The capability of oxidoreductases including that of hexose oxidase to generate hydrogen peroxide has also been utilized to improve the storage stability of certain food products including cheese, butter and fruit juice as it is disclosed in JP-B-73/016612. It has also been suggested that oxidoreductases may be potentially useful as antioxidants in food products.

However, the present invention has demonstrated that hexose oxidase is highly useful as a dough conditioning agent in the manufacturing of flour dough products including not only bread products but also other products made from flour doughs such as noodles and alimentary paste products.

WO 94/04035 discloses a method of improving properties of a dough (with and without fat) and/or baked product made from dough by adding a lipase of microbial origin to the dough. The use of the microbial lipase resulted in an increased volume and improved softness of the baked product. Furthermore an antistaling effect was found.

EP 1 108 360 A1 discloses a method of preparing a flour dough. The method comprises adding to the dough components an enzyme that under dough conditions is capable of hydrolysing a nonpolar lipid, a glycolipid and a phospholipid, or a composition containing said enzyme and mixing the dough components to obtain the dough.

WO 02/03805 discloses that the addition to dough of a combination of two lipases with different substrate specificities. The combination produces a synergistic effect on the dough or on a baked product made from the dough. Optionally, an additional enzyme may be used together with the lipase.

SUMMARY OF THE INVENTION

Accordingly, the invention relates in a first aspect to a method of improving the rheological properties of a flour dough and the quality of the finished product made from the dough, comprising adding to the dough ingredients, dough additives or the dough an effective amount of an oxidoreductase which at least is capable of oxidizing maltose, such as e.g. a hexose oxidase.

In a further aspect, there is also provided a dough bakery product improving composition comprising an oxidoreductase which at least is capable of oxidizing maltose, and at least one further dough ingredient or dough additive.

In still further aspects, the invention pertains to a method of preparing a bakery product, comprising preparing a flour dough including adding an effective amount of an oxidoreductase which at least is capable of oxidizing maltose and baking the dough, and a method of preparing a dough-based food product comprising adding to the dough an effective amount of a maltose oxidizing oxidoreductase.

In addition, we have surprisingly found that a combination of a Hox and an emulsifying agent results in particularly advantageous properties in dough and dough products and/or in baked products therefrom. In particular the stability (e.g. shock stability) and/or rheological (e.g. decrease in stickiness) and/or machineability properties and/or the resultant volume of either the dough and/or baked products (e.g. baked products with better crumb structure and/or homogeneity) is/are improved. Furthermore, the combination of the Hox and emulsifying agent results in an improvement in bread quality, in particular in respect of specific volume and/or crumb homogeneity, which is not a simple additive effect, but may reflect a synergistic effect of these types of enzymes.

The invention further relates to the use of a Hox and an emulsifying agent to improve the rheological and/or machineability properties of dough.

The invention further relates to the use of a Hox and an emulsifying agent to improve the volume of a baked product made from a dough.

DETAILED DISCLOSURE OF THE INVENTION

In one aspect, the present method contemplates a method of improving the rheological properties of flour doughs. The method comprises, as it is mentioned above, the addition of an effective amount of a maltose oxidizing oxidoreductase either to a component of the dough recipe or to the dough resulting from mixing all of the components for the dough. In the present context, “an effective amount” is used to indicate that the amount is sufficient to confer to the dough and/or the finished product improved characteristics as defined herein.

In another aspect the invention provides a method of improving the rheological and/or machineability properties of a flour dough and/or the quality (e.g. volume) of the product made from the dough, comprising adding to the dough a combination comprising a Hox and an emulsifying agent.

Factors which influence the rheological properties and/or the machineability include stickiness and extensibility.

In another aspect the invention provides a method of improving the theological and/or machineability properties of a flour dough and/or the quality (e.g. volume) of the product made from the dough, comprising adding to the dough a combination comprising a Hox and an emulsifying agent wherein the flour dough comprises at least one further dough additive or ingredient.

In another aspect the invention provides a method of improving the rheological and/or machineability properties of a flour dough and/or the quality (e.g. volume) of the product made from the dough, comprising adding to the dough a combination comprising a Hox and an emulsifying agent wherein the product is selected from the group consisting of a bread product, a noodle product, a cake product, a pasta product and an alimentary paste product.

In another aspect the invention provides a method of improving the rheological and/or machineability properties of a flour dough and/or the quality (e.g. volume) of the product made from the dough, comprising adding to the dough a combination comprising a Hox and an emulsifying agent wherein at least one further enzyme is added to the dough ingredients, dough additives or the dough.

In another aspect the invention provides a dough improving composition comprising a Hox and an emulsifying agent.

In another aspect the invention provides a dough improving composition comprising a Hox and an emulsifying agent wherein the flour dough comprises at least one further dough additive or ingredient.

In another aspect the invention provides use of a dough improving composition comprising a Hox and an emulsifying agent in the manufacture of a product made from dough wherein the product is selected from the group consisting of a bread product, a noodle product, a cake product, a pasta product and an alimentary paste product.

In another aspect the invention provides a dough improving composition comprising a Hox and an emulsifying agent wherein at least one further enzyme is added to the dough ingredients, dough additives or the dough.

In another aspect the invention provides use of a dough improving composition comprising a Hox and an emulsifying agent wherein said composition improves the rheological and/or machineability properties of flour dough.

In another aspect the invention provides use of a dough improving composition comprising a Hox and an emulsifying agent wherein said composition improves the volume of a baked product made from a flour dough.

In another aspect the invention provides a dough for addition to a sponge wherein said dough comprises a Hox and an emulsifying agent.

In another aspect the invention provides a dough for addition to a sponge wherein said dough comprises a Hox and an emulsifying agent and wherein the dough comprises at least one further dough additive or ingredient.

Hexose Oxidase

In one useful embodiment of the method according to the invention, the oxidoreductase is a hexose oxidase.

The term “Hox” as used herein refers to Hexose oxidase (D-hexose:O2-oxidoreductase, EC 1.1.3.5). Below discloses some of the sources of Hox. WO 96/40935 discloses a method of producing Hox by recombinant DNA technology. U.S. Pat. No. 6,251,626 discloses hexose oxidase sequences.

The Hox may be isolated and/or purified from natural sources or it may be prepared by use of recombinant DNA techniques.

The Hox may be a variant or derivative of a natural Hox.

The Hox, or the variant or derivative of a natural Hox, is capable of oxidising maltose in the dough.

Preferably the Hox is added in a substantially pure and/or substantially isolated form.

Hexose oxidase can, as it is described in details herein, be isolated from marine algal species naturally producing that enzyme. Such species are found in the family Gigartinaceae which belong to the order Gigartinales. Examples of hexose oxidase producing algal species belonging to Gigartinaceae are Chondrus crispus and Iridophycus flaccidum. Also algal species of the order Cryptomeniales including the species Euthora cristata are potential sources of hexose oxidase.

When using such natural sources for hexose oxidase, the enzyme is typically isolated from the algal starting material by extraction using an aqueous extraction medium. As starting material may be used algae in their fresh state as harvested from the marine area where they grow, or the algal material can be used for extraction of hexose oxidase after drying the fronds e.g. by air-drying at ambient temperatures or by any appropriate industrial drying method such as drying in circulated heated air or by freeze-drying. In order to facilitate the subsequent extraction step, the fresh or dried starting material may advantageously be comminuted e.g. by grinding or blending.

As the aqueous extraction medium, buffer solutions e.g. having a pH in the range of 5-8, such as 0.1 M sodium phosphate buffer, 20 mM triethanolamine buffer or 20 mM Tris-HCl buffer are suitable. The hexose oxidase is typically extracted from the algal material by suspending the starting material in the buffer and keeping the suspension at a temperature in the range of 0-20° C. such as at about 5° C. for 1 to 5 days, preferably under agitation.

The suspended algal material is then separated from the aqueous medium by an appropriate separation method such as filtration, sieving or centrifugation and the hexose oxidase is subsequently recovered from the filtrate or supernatant. Optionally, the separated algal material is subjected to one or more further extraction steps.

Since several marine algae contain coloured pigments such as phycocyanins, it may be required to subject the filtrate or supernatant to a further purification step whereby these pigments are removed. As an example, the pigments may be removed by treating the filtrate or supernatant with an organic solvent in which the pigments are soluble and subsequently separating the solvent containing the dissolved pigments from the aqueous medium. Alternatively, pigments may be removed by subjecting the filtrate or supernatant to a hydrophobic interaction chromatography step.

The recovery of hexose oxidase from the aqueous extraction medium is carried out by any suitable conventional methods allowing isolation of proteins from aqueous media. Such methods, examples of which will be described in details in the following, include conventional methods for isolation of proteins such as ion exchange chromatography, optionally followed by a concentration step such as ultrafiltration. It is also possible to recover the enzyme by adding substances such as e.g. (NH4)2SO4 or polyethylene glycol (PEG) which causes the protein to precipitate, followed by separating the precipitate and optionally subjecting it to conditions allowing the protein to dissolve.

For certain applications of hexose oxidase it is desirable to provide the enzyme in a substantially pure form e.g. as a preparation essentially without other proteins or non-protein contaminants and accordingly, the relatively crude enzyme preparation resulting from the above extraction and isolation steps may be subjected to further purification steps such as further chromatography steps, gel filtration or chromatofocusing as it will also be described by way of example in the following.

Further Dough Additives or Ingredients (Components)

In a preferred embodiment of the method according to the invention, a flour dough is prepared by mixing flour with water, a leavening agent such as yeast or a conventional chemical leavening agent, and an effective amount of hexose oxidase under dough forming conditions. It is, however, within the scope of the invention that further components can be added to the dough mixture.

Typically, such further dough components include conventionally used dough components such as salt (such as sodium chloride, calcium acetate, sodium sulfate or calcium sulfate), sweetening agents such as sugars, syrups or artificial sweetening agents, lipid substances including shortening, margarine, butter or an animal or vegetable oil, glycerol and one or more dough additives such as emulsifying agents, starch degrading enzymes, cellulose or hemicellulose degrading enzymes, proteases, lipases, non-specific oxidizing agents such as those mentioned above, flavouring agents, lactic acid bacterial cultures, vitamins, minerals, hydrocolloids such as alginates, carrageenans, pectins, vegetable gums including e.g. guar gum and locust bean gum, and dietary fiber substances.

The dough may also comprise other conventional dough ingredients, e.g.: proteins, such as milk powder, gluten, and soy; eggs (either whole eggs, egg yolks or egg white); an oxidant such as ascorbic acid, potassium bromate, potassium iodate, azodicarbonamide (ADA) or ammonium persulfate; an amino acid such as L-cysteine; a sugar.

The dough may comprise fat such as granulated fat or shortening.

The further dough additive or ingredient can be added together with any dough ingredient including the flour, water or optional other ingredients or additives, or the dough improving composition. The further dough additive or ingredient can be added before the flour, water, optional other ingredients and additives or the dough improving composition. The further dough additive or ingredient can be added after the flour, water, optional other ingredients and additives or the dough improving composition.

The further dough additive or ingredient may conveniently be a liquid preparation. However, the further dough additive or ingredient may be conveniently in the form of a dry composition.

Preferably the further dough additive or ingredient is selected from the group consisting of a vegetable oil, a vegetable fat, an animal fat, shortening, glycerol, margarine, butter, butterfat and milk fat.

Preferably the further dough additive or ingredient is at least 1% the weight of the flour component of dough. More preferably, the further dough additive or ingredient is at least 2%, preferably at least 3%, preferably at least 4%, preferably at least 5%, preferably at least 6%.

If the additive is a fat, then typically the fat may be present in an amount of from 1 to 5%, typically 1 to 3%, more typically about 2%.

Further Enzymes

In one advantageous embodiment of the above method at least one further enzyme is added to the dough. Suitable examples hereof include a cellulase, a hemicellulase, a xylanase, a starch degrading enzyme, a glucose oxidase, a lipase, a lipoxygenase, an oxidoreductase and a protease.

Among starch degrading enzymes, amylases are particularly useful as dough improving additives. Other useful starch degrading enzymes which may be added to a dough composition include glucoamylases and pullulanases.

The term “xylanase” as used herein refers to xylanases (EC 3.2.1.32) which hydrolyse xylosidic linkages.

The further enzyme can be added together with any dough ingredient including the flour, water or optional other ingredients or additives, or the dough improving composition. The further enzyme can be added before the flour, water, and optionally other ingredients and additives or the dough improving composition. The further enzyme can be added after the flour, water, and optionally other ingredients and additives or the dough improving composition.

The further enzyme may conveniently be a liquid preparation. However, the composition may be conveniently in the form of a dry composition.

In some aspects of the present invention it may be found that some enzymes of the dough improving composition of the invention are capable of interacting with each other under the dough conditions to an extent where the effect on improvement of the rheological and/or machineability properties of a flour dough and/or the quality of the product made from dough by the enzymes is not only additive, but the effect is synergistic.

In relation to improvement of the product made from dough (finished product), it may be found that the combination results in a substantial synergistic effect in respect to crumb homogeneity as defined herein. Also, with respect to the specific volume of baked product a synergistic effect may be found.

Emulsifying Agent

The dough may further comprise a further emulsifier such as mono- or diglycerides, sugar esters of fatty acids, polyglycerol esters of fatty acids, lactic acid esters of monoglycerides, acetic acid esters of monoglycerides, polyoxethylene stearates, or lysolecithin. Among starch degrading enzymes, amylases are particularly useful as dough improving additives. α-amylase breaks down starch into dextrins which are further broken down by β-amylase into maltose. Other useful starch degrading enzymes which may be added to a dough composition include glucoamylases and pullulanases. In the present context, further interesting enzymes are xylanases and other oxidoreductases such as glucose oxidase, pyranose oxidase and sulfhydryl oxidase.

Conventional emulsifiers used in making flour dough products include as examples monoglycerides, diacetyl tartaric acid esters of mono- and diglycerides of fatty acids, and lecithins e.g. obtained from soya.

The emulsifying agent may be an emulsifier per se or an agent that generates an emulsifier in situ.

Examples of emulsifying agents that can generate an emulsifier in situ include enzymes.

Preferably the emulsifying agent is a lipase.

Lipase

The term “lipase” as used herein refers to enzymes which are capable of hydrolysing carboxylic ester bonds to release carboxylate (EC 3.1.1). Examples of lipases include but are not limited to triacylglycerol lipase (EC 3.1.1.3), galactolipase (EC 3.1.1.26), phospholipase (EC 3.1.1.32).

The lipase may be isolated and/or purified from natural sources or it may be prepared by use of recombinant DNA techniques.

Preferably the lipase is selected from the group comprising triacylglycerol lipase, a galactolipase, phospholipase.

More preferably the lipase(s) may be one or more of: triacylglycerol lipase (EC 3.1.1.3), phospholipase A2 (EC 3.1.1.4), galactolipase (EC 3.1.1.26), phospholipase A1 (EC 3.1.1.32), lipoprotein lipase A2 (EC 3.1.1.34).

The lipase may be a variant or derivative of a natural lipase.

For some aspects, preferably the lipase is a phospholipase (including a variant phospholipase).

Preferably the lipase is added in a substantially pure and/or substantially isolated form.

Lipases that are useful in the present invention can be derived from a bacterial species, a fungal species, a yeast species, an animal cell and a plant cell. Whereas the enzyme may be provided by cultivating cultures of such source organisms naturally producing lipase, it may be more convenient and cost-effective to produce it by means of genetically modified cells such as it is described WO 9800136. The term “derived” may imply that a gene coding for the lipase is isolated from a source organism and inserted into a host cell capable of expressing the gene.

WO 02/03805 teaches some of the sources of lipases. The lipases that are taught therein are incorporated herein by reference.

For some aspects of the present invention the lipase may be Lipopan F (supplied by Novozymes) or a variant thereof.

Dough Preparation

The dough is prepared by admixing flour, water, the oxidoreductase according to the invention or the dough improving composition and optionally other possible ingredients and additives. The oxidoreductase or dough improving composition can be added together with any dough ingredient including the flour, the water or dough ingredient mixture or with any additive or additive mixture. The dough improving composition can be added before the flour or water or optional other ingredients and additives. The dough improving composition can be added after the flour or water, or optional other ingredients and additives. The dough can be prepared by any conventional dough preparation method common in the baking industry or in any other industry making flour dough based products.

The dough of the invention generally comprises wheat meal or wheat flour and/or other types of meal, flour or starch such as corn flour, corn starch, maize flour, rice flour, rye meal, rye flour, oat flour, oat meal, soy flour, sorghum meal, sorghum flour, potato meal, potato flour or potato starch.

A preferred flour is wheat flour, but doughs comprising flour derived from other cereal species such as from rice, maize, corn, oat, barley, rye, durra, soy, sorghum and potato are also contemplated.

Preferably the flour dough comprises a hard flour.

The term “hard flour” as used herein refers to flour which has a higher protein content such as gluten than other flours and is suitable for the production of, for example, bread. The term “hard flour” as used herein is synonymous with the term “strong flour”.

Preferably the flour dough comprises a hard wheat flour.

The invention also provides a pre-mix comprising flour together with the combination as described herein. The pre-mix may contain other dough-improving and/or bread-improving additives, e.g. any of the additives, including enzymes, mentioned herein.

The dough of the invention may be fresh, frozen, or part-baked.

The dough of the invention can be a leavened dough or a dough to be subjected to leavening. The dough may be leavened in various ways, such as by adding chemical leavening agents, e.g., sodium bicarbonate or by adding a leaven (fermenting dough), but it is preferred to leaven the dough by adding a suitable yeast culture, such as a culture of Saccharomyces cerevisiae (baker's yeast), e.g. a commercially available strain of S. cerevisiae.

The oxidoreductase or the dough improving composition can be added as a liquid preparation or in the form of a dry powder composition either comprising the enzyme as the sole active component or in admixture with one or more other dough ingredients or additive.

The amount of the enzyme component added normally is an amount which results in the presence in the finished dough of 1 to 10,000 units per kg of flour, preferably 5 to 5000 units such as 10 to 1000 units. In useful embodiments, the amount is in the range of 20 to 500 units per kg of flour. In the present context 1 oxidoreductase unit corresponds to the amount of enzyme which under specified conditions results in the conversion of 1 μmole glucose per minute. The activity is stated as units per g of enzyme preparation.

Rheological Properties

The phrase “rheological properties” as used herein relates to the physical and chemical phenomena described herein which in combination will determine the performance of flour doughs and thereby also the quality of the resulting products.

The phrase “machineability of a flour dough” as used herein refers to the improved manipulation by machinery of the dough. The dough is less sticky compared to the dough without the addition of the combination.

In a further embodiment, the invention relates to improvement of the rheological characteristics of the dough including that the gluten index in the dough is increased by at least 5%, relative to a dough without addition of a combination, the gluten index is determined by means of a Glutomatic 2200 apparatus.

The phrase “rheological properties” as used herein refers to the effects of dough conditioners on dough strength and stability as the most important characteristics of flour doughs. According to American Association of Cereal Chemists (AACC) Method 36-01A the term “stability” can be defined as “the range of dough time over which a positive response is obtained and that property of a rounded dough by which it resists flattening under its own weight over a course of time”. According to the same method, the term “response” is defined as “the reaction of dough to a known and specific stimulus, substance or set of conditions, usually determined by baking it in comparison with a control”.

As it is mentioned herein, it is generally desirable to improve the baking performance of flour to achieve a dough with improved stretchability and thus having a desirable strength and stability by adding oxidising agents which cause the formation of protein disulphide bonds whereby the protein forms a more stable matrix resulting in a better dough quality and improvements of the volume and crumb structure of baked products.

The effect of the oxidoreductase or the dough improving composition on the rheological properties of the dough can be measured by standard methods according to the International Association of Cereal Chemistry (ICC) and the American Association of Cereal Chemistry (AACC) including the amylograph method (ICC 126), the farinograph method (AACC 54-21) and the extensigraph method (AACC 54-10). The extensigraph method measures e.g. the doughs ability to retain gas evolved by yeast and the ability to withstand proofing. In effect, the extensigraph method measures the relative strength of a dough. A strong dough exhibits a higher and, in some cases, a longer extensigraph curve than does a weak dough. AACC method 54-10 defines the extensigraph in the following manner: “the extensigraph records a load-extension curve for a test piece of dough until it breaks. Characteristics of load-extension curves or extensigrams are used to assess general quality of flour and its responses to improving agents”.

In a preferred embodiment of the invention, the resistance to extension of the dough in terms of the ratio between the resistance to extension (height of curve, B) and the extensibility (length of curve, C), i.e. the B/C ratio as measured by the AACC method 54-10 is increased by at least 10% relative to that of an otherwise similar dough not containing oxidoreductase. In more preferred embodiments, the resistance to extension is increased by at least 20%, such as at least 50% and in particular by at least 100%.

The method according to the invention can be used for any type of flour dough with the aims of improving the rheologi-cal properties hereof and the quality of the finished prod-ucts made from the particular type of dough. Thus, the method is highly suitable for the making of conventional types of yeast leavened bread products including wheat flour based bread products such as loaves and rolls. However, it is contemplated that the method also can improve the properties of doughs in which leavening is caused by the addition of chemical leavening agents, including sweet bakery products such as cake products including as examples pound cakes and muffins, or scones.

Noodles

In one interesting aspect, the invention is used to improve the rheological properties of doughs intended for noodle products including “white noodles” and “chinese noodles” and to improve the textural qualities of the finished noodle products. A typical basic recipe for the manufacturing of noodles comprises the following ingredients: wheat flour 100 parts, salt 0.5 parts and water 33 parts. Furthermore, glycerol is often added to the noodle dough. The noodles are typically prepared by mixing the ingredients in an appropriate mixing apparatus followed by rolling out the noodle dough using an appropriate noodle machine to form the noodle strings which are subsequently air dried.

The quality of the finished noodles is assessed inter alia (i.a.) by their colour, cooking quality and texture. The noodles should cook as quickly as possible, remain firm after cooking and should preferably not loose any solids to the cooking water. On serving the noodles should preferably have a smooth and firm surface not showing stickiness and provide a firm “bite” and a good mouthfeel. Furthermore, it is important that the noodles have a light colour.

Since the appropriateness of wheat flour for providing noodles having the desired textural and eating qualities may vary according to the year and the growth area, it is usual to add noodle improvers to the dough in order to compensate for sub-optimal quality of the flour. Typically, such improvers will comprise dietary fiber substances, vegetable proteins, emulsifiers and hydrocolloids such as e.g. alginates, carrageenans, pectins, vegetable gums including guar gum and locust bean gum, and amylases, and glycerol.

It has been attempted to use glucose oxidase as a noodle improving agent. However, as mentioned above, the content of glucose may be so low in wheat flour that this enzyme will not be effective.

It is therefore an important aspect of the invention that the oxidoreductase according to the invention and the composition according to the invention is useful as a noodle improving agent, optionally in combination with glycerol and other components currently used to improve the quality of noodles. Thus, it is contemplated that noodles prepared in accordance with the above method will have improved properties with respect to colour, cooking and eating qualities including a firm, elastic and non-sticky texture and consistency.

Alimentary Paste Product

In a further useful embodiment the dough which is prepared by the method according to the invention is a dough for preparing an alimentary paste product. Such products which include as examples spaghetti and maccaroni are typically prepared from a dough comprising as the main ingredients such as flour, eggs or egg powder and/or water. After mixing of the ingredient, the dough is formed to the desired type of paste product and air dried. It is contemplated that the addition of the combination to a paste dough, optionally in combination with its substrate, will have a significant improving effect on the extensibility and stability hereof resulting in finished paste product having textural and eating qualities.

In a further aspect of the invention there is provided a dough improving composition comprising the oxidoreductase according to the invention and at least one further dough ingredient or dough additive.

Bread

In the invention the improvement of the rheological properties of the dough include that the resistance to extension of the dough in terms of the ratio between resistance to extension (height of curve, B) and the extensibility (length of curve, C), i.e. the B/C ratio, as measured by the AACC method 54-10 is increased by at least 10% relative to that of an otherwise similar dough that does not comprise the combination and wherein the improvement of the quality of the finished product made from the dough is that the average pore diameter of the crumb of the bread made from the dough is reduced by at least 10%, relative to a bread which is made from a bread dough without addition of the combination.

In a further embodiment, the invention, implies that the improvement of the quality of the product made from the dough consists in that the pore homogeneity of the crumb of the bread made from the dough is increased by at least 5%, relative to a bread which is made from a bread dough without addition of the combination. The pore homogeneity of bread is conveniently measured by means of an image analyser composed of a standard CCD-video camera, a video digitiser and a personal computer with WinGrain software. Using such an analyzer, the results of pore diameter in mm and pore homogeneity can be calculated as an average of measurements from 10 slices of bread. The pore homogeneity is expressed in % of pores that are larger than 0.5 times the average of pore diameter and smaller than 2 times the average diameter.

Preferably, the dough is a yeast leavened dough. Although, it is preferred to use the method of the present invention for the manufacture of yeast leavened bread products such as bread loaves, rolls or toast bread, the use of the method for any other type of dough and dough based products such as noodle and pasta products and cakes, the quality of which can be improved by the addition of the combination according to the present invention, is also contemplated.

Preferably the method comprises a further step that the dough is baked to obtain a baked product.

Preferably, when the dough is a bread dough, the method comprises as a further step that the dough is baked to obtain a baked product. One particularly desired property of baked bread products is a high specific volume as defined in the examples. Accordingly, the addition of the combination of the invention preferably results in an increase of the specific volume of the baked product that is at least 10%, relative to a baked product made under identical conditions except that the enzyme is not added. More preferably, the increase of the specific volume is at least 20% such as at least 30%, e.g. at least 40%. Alternatively, the dough is a dough selected from the group consisting of a pasta dough, a noodle dough, and a cake dough or batter.

The phrase “quality of the product” as used herein refers to the final and stable volume and/or crust colour and/or texture and taste.

The term “product made from dough” as used herein refers to a bread product such as in the form of loaves or rolls, french baguette type bread, pita bread, tacos and crisp bread. Preferably the term refers to cakes, pan-cakes, biscuits. More preferably the term refers to pasta. More preferably the term refers to noodles. More preferably the term refers to alimentary paste product.

In a preferred embodiment, the oxidoreductase is hexose oxidase. The further ingredients or additive can be any of the ingredients or additives which are described above. The composition may conveniently be a liquid preparation comprising the oxidoreductase. However, the composition is conveniently in the form of dry composition. It will be understood that the amount of oxidoreductase activity in the composition will depend on the types and amounts of the further ingredients or additives. However, the amount of oxidoreductase activity is preferably in the range of 10 to 100,000 units, preferably in the range of 100 to 50,000 units such as 1,000 to 10,000 units including 2,000 to 5,000 units.

Optionally, the composition may be in the form of a complete dough additive mixture or pre-mixture for a making a particular finished product and containing all of the dry ingredients and additives for such a dough. In specific embodiments, the composition may be one particularly useful for preparing a baking product or in the making of a noodle product or an alimentary paste product.

As mentioned above, the present invention provides a method for preparing a bakery product including the addition to the dough of an oxidoreductase such as e.g. hexose oxidase. In particular, this method results in bakery products such as the above mentioned products in which the specific volume is increased relative to an otherwise similar bakery product, prepared from a dough not containing oxidoreductase. It has been found that the addition of the composition of the present invention to bakery product doughs results in bakery products such as yeast leavened and chemically leavened products in which the specific volume is increased relative to an otherwise similar bakery product. In this context, the expression “specific volume” is used to indicate the ratio between volume and weight of the product. It has been found that, in accordance with the method described herein, the specific volume can be increased significantly such as by at least 10%, preferably by at least 20%, including by at least 30%, preferably by at least 40% and more preferably by at least 50%.

The present invention is highly suitable for improving the rheological and/or machineability properties and/or quality (e.g. volume) of the finished products (products made from the dough) of conventional types of yeast leavened bread products based on wheat Hour, such as loaves and rolls. The present invention is also suitable for improving the rheological properties of doughs containing chemical leavening agents (baking powder) and the quality (e.g. volume) of products made from such doughs. Such product include as examples breads, sponge cakes and muffins.

Enzyme Amount

Preferably the or each enzyme is added in an amount from 1-1000 ppm, preferably 25-500 ppm, more preferably 50-300 ppm.

Nucleotide Sequence

The enzyme need not be a native enzyme. In this regard, the term “native enzyme” means an entire enzyme that is in its native environment and when it has been expressed by its native nucleotide sequence.

The nucleotide sequence of the present invention may be prepared using recombinant DNA techniques (i.e. recombinant DNA). However, in an alternative embodiment of the invention, the nucleotide sequence could be synthesised, in whole or in part, using chemical methods well known in the art (see Caruthers M H et al (1980) Nuc Acids Res Symp Ser 215-23 and Horn T et al (1980) Nuc Acids Res Symp Ser 225-232).

Amino Acid Sequences

The enzyme may be prepared/isolated from a suitable source, or it may be made synthetically or it may be prepared by use of recombinant DNA techniques.

Variants/Homologues/Derivatives

The present invention also encompasses the use of variants, homologues and derivatives of any amino acid sequence of an enzyme of the present invention or of any nucleotide sequence encoding such an enzyme. Here, the term “homologue” means an entity having a certain homology with the subject amino acid sequences and the subject nucleotide sequences. Here, the term “homology” can be equated with “identity”.

In the present context, an homologous sequence is taken to include an amino acid sequence which may be at least 75, 85 or 90% identical, preferably at least 95 or 98% identical to the subject sequence. Typically, the homologues will comprise the same active sites etc. as the subject amino acid sequence. Although homology can also be considered in terms of similarity (i.e. amino acid residues having similar chemical properties/functions), in the context of the present invention it is preferred to express homology in terms of sequence identity.

Homology comparisons can be conducted by eye, or more usually, with the aid of readily available sequence comparison programs. These commercially available computer programs can calculate % homology between two or more sequences.

The invention will now be described by way of illustration in the following non-limiting examples and the drawings, in which:

FIG. 1 which is a photographic image of a bread;

FIG. 2 which is a photographic image of a bread; and

FIG. 3 which is a photographic image of a bread.

EXAMPLE 1

1.1. Purification of Hexose Oxidase from Chondrus crispus

A purified hexose oxidase preparation was obtained using the below extraction and purification procedures. During these procedures and the following characterizations of the purified enzyme, the following assay for determination of hexose oxidase activity was used:

1.1.1. Assay of Hexose Oxidase Activity

The assay was based on the method described by Sullivan and Ikawa (Biochimica et Biophysica Acta, 1973, 309:11-22), but modified to run in microtiter plates. An assay mixture contained 150 μl β-D-glucose (0.1 M in 0.1 M sodium phosphate buffer, pH 6.3), 120 μl 0.1 M sodium phosphate buffer, pH 6.3, 10 μl o-dianisidinedihydrochloride (Sigma D-3252, 3.0 mg/ml in H2O), 10 μl peroxidase (POD) (Sigma P-8125, 0.1 ml in 0.1 M sodium phosphate buffer, pH 6.3) and 10 μl enzyme (HOX) solution. Blanks were made by adding buffer in place of enzyme solution.

The incubation was started by the addition of glucose. After 15 minutes of incubation at 25° C. the absorbance at 405 nm was read in an ELISA reader. A standard curve was constructed using varying concentrations of H2O2 in place of the enzyme solution.

The reaction can be described in the following manner:

HOX

β-D-glucose+H2O+O2->gluconic acid+H2O2

H2O2+β-dianisidinered->2H2O+o-dianisidineox

Oxidized o-dianisidine has a yellow colour absorbing at 405 nm.

1.1.2. Extraction

Fresh Chondrus crispus fronds were harvested along the coast of Brittany, France. This fresh material was homogenized in a pin mill (Alpine). To a 100 g sample of the resulting homogenized frond material was added 300 ml of 0.1 M sodium phosphate buffer, pH 6.8. The mixture was subsequently sonicated in a sonication bath for 5 minutes and then extracted under constant rotation for 4 days at 5° C., followed by centrifugation of the mixture at 47,000×g for 20 minutes.

300 ml of the resulting clear pink supernatant was desalted by ultrafiltration using an Amicon ultrafiltration unit equipped with an Omega (10 kD cut off, Filtron) ultrafiltration membrane.

1.1.3. Anion Exchange Step

The retentate resulting from 1.1.2 was applied to a 5×10 cm column with 200 ml Q-Sepharose FF equilibrated in 20 mM triethanolamine, pH 7.3. The column was washed with the equilibration buffer and hexose oxidase eluted with a 450 ml gradient of 0 to 1 M of NaCl in equilibration buffer. The column was eluted at 6 ml/minute, and fractions of 14 ml collected. Fractions 9-17 (total 125 ml) were pooled and concentrated by ultrafiltration using an Amicon 8400 unit equipped with an Omega (10 kD cut off, Filtron) ultrafiltration membrane to 7.5 ml.

1.1.4. Gel Filtration

The above 7.5 ml retentate was applied to a Superdex 200 2.6×60 cm gel filtration column equilibrated in 50 mM sodium phosphate buffer, pH 6.4 and eluted at a flow rate of 1 ml/-minute. Fractions of 4 ml were collected. Fractions 17-28 (total volume 50 ml) containing the hexose oxidase activity were pooled.

1.1.5. Hydrophobic Interaction Chromatography

To the pool resulting from the gel filtration step 1.1.4 ammonium sulphate was added to a final concentration of 2 M. This mixture was then applied to a 1.6×16 cm column with 32 ml phenyl sepharose equilibrated in 20 mM sodium phosphate buffer, pH 6.3 and 2 M (NH4)2SO4. The column was washed with equilibration buffer followed by elution of hexose oxidase at a flow rate of 2 ml/minute using a 140 linear gradient from 2 M to 0 M (NH4)2SO4 in 20 mM sodium phosphate buffer. Fractions of 4 ml were collected and fractions 24-33 containing the hexose oxidase activity were pooled.

The above mentioned pink colour accompanies the enzyme, but it is separated from hexose oxidase in this purification step.

1.1.6. Mono Q Anion Exchange

The above pool resulting from the above phenyl sepharose chromatography step was desalted by ultrafiltration as described above. 2 ml of this pool was applied to a Mono Q HR 5/5 column equilibrated in 20 mM triethanolamine, pH 7.3. The column was subsequently eluted using a 45 ml linear gradient from 0 to 0.65 M NaCl in equilibration buffer at a flow rate 1.5 ml/minute. Fractions of 1.5 ml were collected and fractions 14-24 were pooled.

1.1.7. Mono P Anion Exchange

The hexose oxidase-containing pool from the above step 1.1.6 was applied to a Mono P HR 5/5 column equilibrated in 20 mM bis-Tris buffer, pH 6.5. The enzyme was eluted using a 45 ml linear gradient from 0 to 0.65 M NaCl in equilibration buffer at a flow rate of 1.5 ml/minute, and fractions of 0.75 ml were collected. The highest hexose oxidase activity was found in fraction 12.

1.2. Characterization of the Purified Hexose Oxidase

The hexose oxidase-containing pools from the above steps 1.1.6 and 1.1.7 were used in the below characterization experiments:

1.2.1. Determination of Molecular Weight

The size of the purified native hexose oxidase was determined by gel permeation chromatography using a Superose 6 HR 10/30 column at a flow rate of 0.5 ml/minute in 50 mM sodium phosphate buffer, pH 6.4. Ferritin (440 kD), catalase (232 kD), aldolase (158 kD), bovine serum albumin (67 kD) and chymotrypsinogen (25 kD) were used as size standards. The molecular weight of the purified hexose oxidase was determined to be 120+10 kD.

1.2.2. Determination of pH Optimum

Assay mixtures for the determination of pH optimum (final volume 300 μl) contained 120 μl of 0.1 M stock solution of sodium phosphate/citrate buffer of varying pH values. All other assay mixture components were dissolved in H2O. The pH was determined in the diluted stock buffer solutions at 25° C. The hexose oxidase showed enzymatic activity from pH 3 to pH 8, but with optimum in the range of 3.5 to 5.5.

1.2.3. Km of the Hexose Oxidase for Glucose and Maltose Respectively

Kinetic data were fitted to V=VmaxS/(Km+S), where Vmax is the maximum velocity, S is the substrate concentration and Km is the concentration giving 50% of the maximum rate (Michaelis constant) using the EZ-FIT curve fitting microcomputer programme (Perrella, F. W., 1988, Analytical Biochemistry, 174:437-447).

A typical hyperbolic saturation curve was obtained for the enzyme activity as a function of glucose and maltose, respectively. Km for glucose was calculated to be 2.7 mM±0.7 mM and fur maltose the Km was found to be 43.7±5.6 mM.

EXAMPLE 2 Dough Improving Effect of Hexose Oxidase Extracted from Chondrus crispus

2.1. Purification of Hexose Oxidase from Chondrus crispus

For this experiment, hexose oxidase was prepared in the following manner:

Fresh Chondrus crispus material was collected at the coast of Brittany, France. The material was freeze-dried and subsequently ground. 40 g of this ground material was suspended in 1000 ml of 20 mM triethanolamine (TEA) buffer, pH 7.3 and left to stand at 5° C. for about 64 hours with gentle agitation and then centrifuged at 2000×g for 10 minutes. The supernatant was filtered through GF/A and GF/C glass filters followed by filtering through a 45 μm pore size filter to obtain a filtrate preparation of 800 ml having hexose oxidase activity corresponding to a glucose oxidase activity of 0.44 units per g of preparation. The activity was determined using the below procedure.

The supernatant was applied onto a 330 ml bed volume chromatographic column with anionic exchange Q Sepharose Big Beads (dead volume 120 ml). The bound proteins were eluted over 180 minutes using a gradient from 0 to 0.5 M NaCl in 20 mM TEA buffer, pH 7.3 followed by 1 M NaCl in 20 mM TEA buffer, and fractions of 9 ml were collected and analyzed for hexose oxidase activity using the below analytical procedure.

Hexose oxidase activity-containing fractions 60-83 were pooled (about 250 ml) and concentrated and desalted by ultrafiltration to about 25 ml. This step was repeated twice on the retentates to which was added 100 ml 0.05 mM TEA. The resulting retentate of 25 ml contained 0.95 glucose oxidase activity units per g.

2.2. Determination of Glucose Oxidase Activity

Definition: 1 glucose oxidase (GOD) unit corresponds to the amount of enzyme which under the specified conditions results in the conversion of 1 μmole glucose per min. The activity is stated as units per g of enzyme preparation.

Reagents: (i) Buffer: 20 g Na2HPO4-2H2O is dissolved in 900 ml distilled water, pH is adjusted to 6.5; (ii) dye reagent (stock solution): 200 mg of 2,6-dichloro-phenol-indophenol, Sigma No. D-1878 is dissolved in 1000 ml distilled water under vigorous agitation for 1 hour; (iii) peroxidase (stock solution): Boehringer Mannheim No. 127 361, 10,000 units is dissolved in 10 ml distilled water and 4.2 g of ammonium sulphate added; (iv) substrate: 10% w/v D-glucose solution in buffer, (v) standard enzyme: hydrase #1423 from Amano.

Analytical principle and procedure: Glucose is converted to gluconic acid and H2O2 which is subsequently converted by peroxidase to H2O and O2. The generated oxygen oxidizes the blue dye reagent 2,6-dichloro-phenol-indophenol which thereby changes its colour to purple. The oxidized colour is measured spectrophotometrically at 590 nm and the enzymatic activity values calculated relative to a standard.

2.3. The Effect of the Hexose Oxidase Preparation on Crosslinking between Thiol Groups in a Wheat Flour Based Dough

The effect of hexose oxidase on the formation of thiol group cross-linking was studied by measuring the content of free thiol groups in a dough prepared from 1500 g of wheat flour, 400 Brabender Units (BU) of water, 90 g of yeast, 20 g of sucrose and 20 g of salt to which was added 0, 100, 250, 875 and 1250 units per kg of flour, respectively of the above hexose oxidase preparation. The measurement was carried out essentially in accordance with the colorimetric method of Ellman (1958) as also described in Cereal Chemistry, 1983, 70, 22-26. This method is based on the principle that 5.5′-dithio-bis(2-nitrobenzoic acid) (DTNB) reacts with thiol groups in the dough to form a highly coloured anion of 2-nitro-5-mercapto-benzoic acid, which is measured spectrophotometrically at 412 nm.

Assuming that the relative change of the amount of thiol groups in a dough is reflected as the change in the optical density (OD) resulting from the reaction between thiol groups and DTNB in the dough, the following results were obtained:

Hexose oxidase
GOD units/kg flour OD412
0 0.297
100 0.285
250 0.265
875 0.187
1250 0.138

Thus, this experiment showed a significant decrease in OD indicating a reduction of the content of free thiol groups which was proportionate to the amount of hexose oxidase activity added.

2.4. Improvement of the Rheological Characteristics of Dough by the Addition of Hexose Oxidase

The above dough was subjected to extensigraph measurements according to AACC Method 54-10 with and without the addition of an amount of the hexose oxidase preparation corresponding to 100 units/kg flour of hexose oxidase activity. The dough without addition of enzyme served as a control.

The principle of the above method is that the dough after forming is subjected to a load-extension test after resting at 30° C. for 45, 90, 135 and 180 minutes, respectively, using an extensigraph capable of recording a load-extension curve (extensigram) which is an indication of the doughs resistance to physical deformation when stretched. From this curve, the resistance to extension, B (height of curve) and the extensibility, C (total length of curve) can be calculated. The B/C ratio (D) is an indication of the baking strength of the flour dough.

The results of the experiment is summarized in Table 2.1 below.

TABLE 2.1
Extensigraph measurements of dough supplemented
with 100 GOD units/kq flour of hexose oxidase (HOX).
Sample Time, min B C D = B/C
Control 45 230 180 1.3
HOX 45 320 180 1.8
Control 90 290 161 1.8
HOX 90 450 148 3.0
Control 135 290 167 1.7
HOX 135 490 146 3.4
Control 180 300 168 1.8
HOX 180 500 154 3.2

It is apparent from this table that the addition of hexose oxidase (HOX) has an improving effect on the doughs resistance to extension as indicated by the increase in B-values. This is reflected in almost a doubling of the B/C ratio as a clear indication that the baking strength of the flour is significantly enhanced by the hexose oxidase addition.

In a similar experiment, 100 units/kg flour of a commercial glucose oxidase product was added and the above parameters measured in the same manner using a dough without enzyme addition as a control. The results of this experiment is shown in Table 2.2 below:

TABLE 2.2
Extensigraph measurements of dough supplemented
with 100 GOD units/kg flour of glucose oxidase (GOX).
Sample Time, min B C D = B/C
Control 45 240 180 1.3
GOX 45 290 170 1.7
Control 90 260 175 1.5
GOX 90 360 156 2.3
Control 135 270 171 1.6
GOX 135 420 141 3.0

When the results for the above two experiments are compared with regard to differences between control dough and the hexose oxidase or glucose oxidase supplemented doughs it appeared that hexose oxidase has a stronger strengthening effect than glucose oxidase. Furthermore, the B/C ratio increased more rapidly with hexose oxidase relative to glucose oxidase which is a clear indication that enhancement of the baking strength is being conferred more efficiently by hexose oxidase than by glucose oxidase.

EXAMPLE 3 Dough Improving Effect of Hexose Oxidase Extracted from Chondrus crispus

For this experiment fresh Chondrus crispus seaweed fronds were harvested along the coast of Hirsholmene, Denmark. Hexose oxidase was isolated using two different extraction procedures, and the materials from both were pooled for the below dough improving experiment.

3.1. Purification of Hexose Oxidase from Chondrus crispus I

954 g of the fresh fronds was rinsed in distilled water, dried with a towel and stored in liquid nitrogen. The seaweed was blended using a Waring blender and 1908 ml of 0.1 M sodium phosphate buffer, 1 M Na Cl, pH 6.8 was added to the blended seaweed. The mixture was extracted under constant stirring for 4 days at 5° C., followed by centrifugation of the mixture at 20,000×g for 30 minutes.

The resulting 1910 ml supernatant (351.1 U/ml) was concentrated to 440 ml at 40° C. in a Buchi Rotavapor R110. The concentrate was ammonium sulphate fractionated to 25% The mixture was stirred for 30 minutes and centrifuged for 20 minutes at 47,000×g. The supernatant (395 ml) was dialysed overnight against 20 l of 10 mM triethanolamine (TEA) buffer, pH 7.3 to a final volume of 610 ml (367.1 U/ml).

The above 610 ml was applied in two runs to a 2.6×25 cm column with 130 ml Q-Sepharose FF equilibrated in 20 mM TEA buffer, pH 7.3. The column was washed with the equilibration buffer and the bound proteins were eluted using 800 ml gra dient from 0 to 0.8 M NaCl in equilibration buffer. The column was eluted at 4 ml/minute and fractions of 12 ml collected. Fractions containing the hexose oxidase activity were collected and pooled to a final volume of 545 ml (241.4 U/ml).

3.2. Purification of Hexose Oxidase from Chondrus crispus II

1250 g of the fresh fronds was rinsed in distilled water, dried with a towel and stored in liquid nitrogen. The seaweed was blended in a Waring blender followed by the addition of 2500 ml 0.1 M sodium phosphate buffer, 1 M NaCl. pH 6.8. The mixture was extracted under continuous stirring for 4 days at 5° C. followed by centrifugation at 20,000×g for 30 minutes.

The resulting 2200 ml supernatant (332.8 U/ml) was concentrated to 445 ml at 40° C. using a Buchi Rotavapor R110. The resulting concentrate was ammonium sulphate fractionated to 25%. The mixture was stirred for 30 minutes and centrifuged for 20 minutes at 47,000×g. The precipitate was discarded. The 380 ml supernatant was dialysed overnight against 20 l 10 mM TEA buffer, pH 7.3, to a final volume of 850 ml (319.2 U/ml).

The above 850 ml was applied to a 2.6×25 cm column with 130 ml Q-Sepharose FF equilibrated in 20 mM TEA buffer, pH 7.3.

The column was washed with the equilibration buffer and the bound proteins were eluted using 800 ml gradient from 0 to 0.8 M NaCl in equilibration buffer. The column was eluted at 4 ml/minute and fractions of 12 ml collected. Fractions containing the hexose oxidase activity were collected and pooled to a final volume of 288 ml.

The retentate from the above step was applied to a 2.6×31 cm column with 185 ml metal chelating sepharose FF loaded with Ni2+ and equilibrated in 50 mM sodium phosphate, 1 M NaCl, pH 7.4. The bound proteins were eluted with a 740 ml gradient of 0 to 35 mM imidazole, pH 4.7 in equilibration buffer. The column was eluted at 2 ml/minute and fractions of 11 ml was collected. Fractions 41-54 (140 ml, 352.3 U/ml) were pooled. Some hexose oxidase did run through the column.

3.3. Pooling and Concentrating of Extracts

The run through and the 140 ml from purification II and the 545 ml from purification I were pooled to a final volume of 1120 ml (303.6 U/ml). The 1120 ml was rotation evaporated into a volume of 210 ml followed by dialysis overnight 30 against 20 l of 10 mM TEA buffer, pH 7.3, to a final volume of 207 ml (1200.4 U/ml)

3.3.1. Anion Exchange Step

The retentate resulting from the above step was applied to a 2.6×25 cm column with 130 ml Q-sepharose FF equilibrated in 20 mM triethanolamine, pH 7.3. The column was washed with the equilibration buffer and the bound proteins eluted using 800 ml gradient from 0 to 0.8 M NaCl in equilibration buffer. The column was eluted at 4 ml/minute and fractions of 12 ml collected. Fractions 30-50 containing the hexose oxidase activity (260 ml, 764.1 U/ml) were collected and pooled.

3.3.2. Other Enzyme Activity

The above pooled solution was tested for the following enzymatic side activities catalase, protease, xylanase, α- and β-amylase and lipase. None of these activities were found in the solution.

3.4. Improvement of the Rheological Characteristics of Dough by the Addition of Hexose Oxidase

A dough was prepared from wheat flour, water and salt and 0, 72, 216 and 360 units per kg of flour, respectively of the above hexose oxidase preparation was added hereto. The dough without addition of enzyme served as a control. In addition two doughs were prepared to which was added 216 and 360 units per kg of flour respectively, of Gluzyme, a glucose oxidase available from Novo Nordisk A/S. Denmark.

The doughs were subjected to extensigraph measurements according to a modification of the above AACC Method 54-10. The results of the experiment are summarized in Table 3.1 below.

TABLE 3.1
Extensigraph measurements of dough supplemented
with hexose oxidase (HOX) or glucose oxidase (units per kg flour)
Sample Time, Min B C D = B/C
Control 45 250 158 1.6
HOX 72 U/kg 45 330 156 2.1
HOX 216 U/kg 45 460 153 3.0
HOX 360 U/kg 45 580 130 4.5
Gluzyme 72 U/kg 45 350 159 2.2
Gluzyme 216 U/kg 45 340 148 2.3
Gluzyme 360 U/kg 45 480 157 3.1
Control 90 290 164 1.8
HOX 72 U/kg 90 470 145 3.2
HOX 216 U/kg 90 650 142 4.6
HOX 360 U/kg 90 870 116 7.5
Gluzyme 72 U/kg 90 450 147 3.1
Gluzyme 216 U/kg 90 480 138 3.5
Gluzyme 360 U/kg 90 500 152 3.2
Control 135 330 156 2.1
HOX 72 U/kg 135 540 129 4.2
HOX 216 U/kg 135 750 125 6.0
HOX 360 U/kg 135 880 117 7.5
Gluzyme 72 U/kg 135 510 136 3.8
Gluzyme 216 U/kg 135 550 122 4.5
Gluzyme 360 U/kg 135 560 121 4.6

It is evident from the above table that the addition of hexose oxidase (HOX) or glucose oxidase had an improving effect on the resistance of doughs to extension as indicated by the increase in B-values. This is reflected in an increase of the B/C ratio as a clear indication that the baking strength of the flour was enhanced significantly by the addition of enzymes.

It is also evident that the hexose oxidase had a higher strengthening effect than glucose oxidase. Furthermore, the B/C ratio increased more rapidly with hexose oxidase relative to glucose oxidase which is a clear indication that enhancement of the baking strength is being conferred more efficiently by hexose oxidase than by glucose oxidase.

EXAMPLE 4 Dough Improving Effect of Hexose Oxidase Extracted from Chondrus crispus

4.1. Purification of Hexose Oxidase from Chondrus crispus

Fresh Chondrus crispus fronds were harvested along the coast of Brittany, France. 2285 g of this fresh material was rinsed in distilled water, dried with a towel and stored in liquid nitrogen. The seaweed was blended in a Waring blender followed by addition of 4570 ml 0.1 M sodium phosphate buffer, 1 M NaCl pH 6.8. The mixture was extracted under continuous magnetic stirring for 4 days at 5° C. followed by centrifugation at 20,000×g for 30 minutes.

The resulting 4930 ml supernatant (624.4 U/ml) was concentrated to 1508 ml at 40° C. using a Buchi Rotavapor R110. The obtained concentrate was polyethylenglycol fractionated to 3% (w/v). The mixture was stirred for 30 minutes and centrifuged for 30 minutes at 47,000×g. The pellet was discarded. The 1470 ml supernatant (2118.7 U/ml) was PEG fractionated to 24%. The mixture was stirred for 30 minutes and centrifuged for 30 minutes at 47,000×g. The supernatant was discarded and the 414.15 g of precipitate was resuspended in 200 ml 20 mM TEA buffer, pH 7.3, followed by dialysis over night at 5° C. against 20 l 10 mM TEA buffer, pH 7.3.

After dialysis the volume was 650 ml (2968.6 U/ml). The suspension was centrifuged for 30 minutes at 20,000×g. The precipitate was discarded and the supernatant was diluted to 3200 ml with distilled water.

The above 3200 ml (829.9 U/ml) was applied to a 10×14 cm column with 1100 ml Q-Sepharose FF equilibrated in 20 mM TEA buffer, pH 7.3. The column was washed with the equilibration buffer and the bound proteins were eluted using 15,000 ml gradient from 0 to 0.8 M NaCl in equilibration buffer. The column was eluted at 50 ml/minute. Hexose oxidase did run through the column and 840 ml of this was collected.

The 840 ml suspension was treated with kieselguhr and concentrated to 335 ml (2693.3 U/ml).

The above 335 ml was applied to a 3 l Sephadex G25C desalting column 10×40 cm. The column was equilibrated in 20 mM TEA buffer, pH 7.3, eluted at a flow rate of 100 ml/minute and 970 ml eluate was collected. This eluate was applied to a 10×14 cm column with 1100 ml Q-Sepharose FF equilibrated in 20 mM TEA, pH 7.3. The column was washed with the equilibration buffer and bound proteins eluted using a 15,000 ml gradient of 0 to 0.8 M NaCl in equilibration buffer. The column was eluted at 50 ml/min. Hexose oxidase did run through the column and 1035 ml of this was collected.

To the above eluate (1035 ml) ammonium sulphate was added to a final concentration of 2 M. The mixture was then applied in two runs to a 5×10 cm column with 200 ml phenyl sepharose HP equilibrated in 25 mM sodium phosphate buffer, pH 6.3 and 2 M (NH4)2SO4. The column was washed with equilibration buffer followed by eluting the bound proteins at a flow rate of 50 ml/minute using 5,000 ml gradient from 2 M to 0 M (NH4)2SO4 in 25 mM sodium phosphate buffer. Fractions of 500 and 29 ml, respectively were collected from run 1 and 2. Fraction 5 in run 1 and fractions 27-42 in run 2 containing the hexose oxidase activity were pooled to a total of 1050 ml (563.9 U/ml).

The above pool was desalted by a 3 l Sephadex G25C gel filtration column. The column was equilibrated in 20 mM TEA buffer, pH 7.3, eluted at a flow rate of 100 ml/minute and 1,000 ml eluate was collected.

The 1,000 ml eluate was concentrated to 202 ml (2310.2 U/ml) and this preparation was used for following rheology testing.

4.2. Improvement of the Rheological Characteristics of Dough by the Addition of Hexose Oxidase

A dough was prepared from wheat flour, water and salt and 0, 288, 504 and 720 oxidoreductase units per kg of flour, respectively of the above hexose oxidase preparation was added hereto. The dough without addition of enzyme served as a control. In addition two doughs were prepared to which was added 288 and 504 oxidoreductase units per kg of flour respectively, of Gluzyme, a glucose oxidase available from Novo Nordisk A/S. Denmark.

The doughs were subjected to extensigraph measurements according to a modification of AACC Method 54-10.

The results of the experiment are summarized in Table 4.1 below.

TABLE 4.1
Extensigraph measurements of dough supplemented
with hexose oxidase (HOX) or glucose oxidase (Units per kg flour).
Sample Time, Min B C D = B/C
Control 45 210 171 1.2
HOX 288 U/kg 45 490 139 3.5
HOX 504 U/kg 45 640 122 5.2
HOX 720 U/kg 45 730 109 6.7
Gluzyme 288 U/kg 45 350 165 2.1
Gluzyme 504 U/kg 45 385 153 2.5
Gluzyme 720 U/kg 45 435 148 2.9
Control 90 275 182 1.5
HOX 288 U/kg 90 710 130 5.5
HOX 504 U/kg 90 825 106 7.8
HOX 720 U/kg 90 905 107 8.5
Gluzyme 288 U/kg 90 465 153 3.0
Gluzyme 504 U/kg 90 515 135 3.8
Gluzyme 720 U/kg 90 540 140 3.9
Control 135 280 175 1.6
HOX 288 U/kg 135 745 102 7.3
HOX 504 U/kg 135 920 94 9.8
HOX 720 U/kg 135 80
Gluzyme 288 U/kg 135 525 129 4.1
Gluzyme 504 U/kg 135 595 129 4.6
Gluzyme 720 U/kg 135 630 121 5.2

It is apparent from the above results that the addition of hexose oxidase (HOX) or glucose oxidase has an improving effect on the resistance of doughs to extension as indicated by the increase in B-values. This is reflected in an increase of the B/C ratio.

It is also apparent that hexose oxidase has a stronger s strengthening effect than that of glucose oxidase, the strengthening effect of both enzymes being proportional to the amount of enzyme added. Furthermore, the B/C ratio increased more rapidly with hexose oxidase relative to glucose oxidase which is a clear indication that enhancement of the baking strength is being conferred more efficiently by hexose oxidase than by glucose oxidase.

EXAMPLE 5 Improving Effect of Hexose Oxidase Extracted from Chondrus crispus on the Specific Volume of Bread

5.1. Purification of Hexose Oxidase from Chondrus crispus

Fresh Chondrus crispus fronds were harvested along the coast of Brittany, France. 2191 g of this fresh material was rinsed in distilled water, dried with a towel and stored in liquid nitrogen. The seaweed was blended in a Waring blender fol lowed by addition of 4382 ml 0.1 M sodium phosphate buffer, 1 M NaCl and pH 6.8. The mixture was extracted under continuously magnetic stirring for 4 days at 5° C. followed by centrifugation at 20,000×g for 20 minutes.

The resulting 4600 ml supernatant (746.1 U/ml) was concentrated to 850 ml at 40° C. in a Buchi Rotavapor R110. This concentrate (3626.9 U/ml) was polyethylene glycol fractionated to 3% (w/v). The mixture was stirred for 30 minutes and centrifuged for 30 minutes at 20,000×g. The precipitate was discarded. The 705 ml supernatant (2489.8 U/ml) was PEG fractionated to 25%. The mixture was stirred for 30 minutes and centrifuged for 30 minutes at 20,000×g. The supernatant was discarded and the 341 g of precipitate was resuspended in 225 ml 20 mM TEA buffer, pH 7.3. The suspension (500 ml) was desalted on a 3 l Sephadex G25C desalting column 10×40 cm. The column was equilibrated in 20 mM TEA buffer, pH 7.3, and eluted at a flow rate of 100 ml/minute. 1605 ml eluate was collected.

To the above eluate (687.5 U/ml) ammonium sulphate was added to a final concentration of 2M. The mixture was then applied in two runs to a 5×10 cm column with 200 ml phenyl sepharose HP equilibrated in 25 mM sodium phosphate buffer, pH 6.3 and 2 M (NH4)2SO4. The column was washed with equilibration buffer followed by elution of the bound proteins at a flow rate of 50 ml/minute using 5,000 ml gradient from 2 M to 0 M (NH4)2SO4 in 25 mM sodium phosphate buffer. Fractions of 29 ml was collected. Fractions 85-105 in run 1 and fractions 36-69 in run 2 containing the hexose activity were pooled to a total of 1485 ml (194.7 U/ml).

The above pool was desalted by a 3 l Sephadex G25C gelfiltration column, the same as used in 4.1. The column was equilibrated in 20 mM TEA buffer, pH 7.3, and eluted at a flow rate of 100 ml/minute. 1,200 ml eluate was collected.

The 1,200 ml eluate was concentrated to 685 ml (726.2 U/ml) and used for baking experiments.

5.2. Improvement of the Specific Volume of Bread by Adding Hexose Oxidase to the Dough

A dough was prepared from 1500 g of flour, 90 g of yeast, 24 g of salt, 24 g of sugar and 400 BU of water and 0 or 108 units of the above purified hexose oxidase and 108 units of Gluzyme (glucose oxidase available from Novo Nordisk, Denmark) per kg flour, respectively was added hereto. The dough was mixed on a Hobart mixer for 2+9 minutes at 26° C. and divided into two parts followed by resting for 10 minutes at 30° C. in a heating cabinet, moulding with a Fortuna 3/17/7 and proofing for 45 minutes at 34° C. and 85% RH. The thus proofed dough was baked at 220° C. for 17 minutes with 12 sec. steam in a Bago oven.

The results of the experiment are summarized in table 5.1 below.

TABLE 5.1
Improvement of specific volumes of bread prepared from
dough supplemented with hexose oxidase or glucose
oxidase (Units per kg flour)
Total volume Total weight Specific volume
control 5325 1027 5.18
Hexose oxidase 108 U/kg 6650 1036 6.41
Gluzyme 108 U/kg 6075 1030 5.89

It is evident from the above table that the addition of hexose oxidase or glucose oxidase had an increasing effect on the total volume, the weight being essentially the same. This is reflected in an increase of the specific volume as compared to the bread baked without addition of enzymes.

It is also evident that hexose oxidase has a significantly larger effect on the increase of the specific volume than had glucose oxidase at the same dosage.

EXAMPLE 6 Characterization of the Purified Hexose Oxidase

Preparations from the above purifications were used for characterization of hexose oxidase.

6.1. Staining for Hexose Activity After Non-Denaturing PAGE

Hexose oxidase activity was analyzed by native PAGE using precast 8-16% Tris-glycine Novex gels according to the manufactures instructions (Novex, San Diego, USA). After electrophoresis the gels were stained for hexose oxidase activity by incubation of the gel in a solution containing 50 mM sodium phosphate buffer, pH 6.0, 100 mM glucose, 50 mg/l phenazine methosulphate (Sigma P9625) and 250 mg/l nitroblue tetrazolium (Sigma N6876) as described in the PhD thesis by Witteveen, C. F. B. (1993) “Gluconate formation and polyol metabolism in Aspergillus niger”. After about 30 minutes the hexose oxidase activity was visible as a double hand very close to each other. The same double band was also seen when a native PAGE of hexose oxidase was silver stained. The molecular weight of purified hexose oxidase was determined to 144 kD by native PAGE. Half the gel was silver stained, the other half was activity stained. As standards were used bovine serum albumin (67 kD), lactate dehydrogenase (140 kD), catalase (232 kD), ferritin (440 kD) and thyroglobulin (669 kD).

6.2 Determination of Molecular Weight by SDS-Page

The molecular weight was also determined on material which was first applied to a native PAGE as described above, after activity staining the hexose oxidase band was excised from the gel and then electroeluted using an Electro-Eluter (model 422, Bio-Rad, CA, USA) according to the manufacturer's recommendations. The electroeluted protein was subjected to SDSPAGE and silver stained. This material gave “one” double band at about 70 kDa in SDS-PAGE gels. The electroeluted hexose oxidase is therefore a dimer of two subunits.

6.3 Determination of pI of Hexose Oxidase

Samples containing hexose oxidase activity were analyzed by isoelectric focusing (IEF) using a precast 3-10 IEF gel according to the manufacturer's recommendations (Novex, San Diego, US). After electrophoresis half of the gel was silver stained and the other half nitroblue tetrazolium stained as described in 6.1.

Hexose oxidase stained as a double band. The pI of the first band was 4.79, pI of the second band was 4.64. As standards were used trypsinogen (9.30), lentil lectin basic band (8.65), lentil lectin middle band (8.45), lentil lectin acid band (8.15), horse myoglobin acidic band (6.85), human carbonic anhydrase B (5.85), β-lactoglobulin A (5.20), soy bean trypsin inhibitor (4.55) and amyloglucosidase (3.50).

6.4 Determination of Km Hexose Oxidase for Different Sugars

Km of hexose oxidase was determined for 7 different sugars as described in 1.2.3. Results are summarized in table 6.1 below.

TABLE 6.1
Determination of Km of hexose oxidase for different sugars
Substrate Km (mM) cv (mM)
D-glucose 2.7 0.7
D-galactose 3.6 1
cellobiose 20.2 7.8
maltose 43.7 5.6
lactose 90.3 20.6
xylose 102 26
arabinose 531 158
(cv = coefficient of variation)

6.5 Determination of a Peptide Sequence of Hexose Oxidase

50 μl from the electroeluted mixture in 6.2 was suspended in 450 μl 0.1% triflouracetic acid (TFA).

To remove the Tris, glycine and SDS, the above mixture was subjected to chromatography on reverse-phase HPLC. The resulting solution was applied in 9 runs to a 4.6×30 cm Brownlee C2 column equilibrated in 0.1% TFA. The column was washed in equilibration buffer and bound peptides eluted with a 14 ml gradient from 10 to 80% acetonitrile in 0.1% TFA, at a flow rate of 0.7 ml/min. Fractions from the largest peak containing the enzyme were collected and freeze dried.

6.5.1 Endoproteinase Lys-C Digestion

The resulting freeze dried enzyme was dissolved in 50 μl 8 M urea, 0.4 M NH4HCO3, pH 8.4. Denaturation and reduction of the protein was carried out by the addition of 5 μl 45 mM di-thiothreitol and under an overlay of N2 at 50° C. for 15 min. The solution was cooled to room temperature and 5 μl 100 mM iodoacetamide was added, the cysteines being derivatized for 15 min. at room temperature in the dark under N2. Subsequently, the solution was suspended in 135 μl water and digestion was carried out at 37° C. under N2 for 24 hours by addition of 5 μg endoproteinase Lys-C dissolved in 5 l water. The reaction was terminated by freezing the reaction mixture at −20° C.

6.5.2 Reverse-Phase HPLC Separation of Peptides

The resulting peptides were separated by reverse-phase HPLC on a VYDAC c18 column 0.46×15 cm (The Separation Group, CA, USA) using as solvent A 0.1% TFA in water and as solvent B 0.1% TFA in acetonitrile.

6.5.3 Peptide Sequencing

Sequencing was performed on an Applied Biosystems 476A sequencer (Applied Biosystems, CA, USA) using pulsed-liquid fast cycles according to the manufacturer's instructions. A peptide having the below amino acid sequence was identified:

D P C Y I V I D V N A G T P O K P D P.

EXAMPLES 7 TO 10

Definitions

All PANODAN™ products contain DATEM (Di-acetyl tartaric acid ester of monoglycerides) and are obtained from Danisco A/S.

PANODAN™ 521: DATEM containing bacterial xylanase and fungal amylase

TS-E 662™ (obtained from Danisco A/S) is a product containing hexose oxidase (Hox) (EC 1.1.1.5) from Chondrus chrispus expressed in Hansenula polymorpha.

TS-E 680™ (obtained from Danisco A/S) is a product containing fungal xylanase (EC 3.2.1.8) from Aspergillus niger.

TS-E 861™ (obtained from Danisco A/S) is a product containing fungal xylanase (EC 3.2.1.8) from Aspergillus niger, lipase (EC 3.1.1.3) from Thermomyces lanuginosa expressed in Aspergillus oryzae, and hexose oxidase (EC 1.1.1.5) from Chondrus crispus expressed in Hansenula polymorepha.

GRINDAMYL™ H 640 (obtained from Danisco A/S): contains bacterial xylanase

Grindamyl™ H 121 (obtained from Danisco A/S) is a fungal xylanase (EC 3.2.1.8) from Aspergillus niger.

Grindamyl™ EXEL 16 (obtained from Danisco A/S) is lipase (EC 3.1.1.3) from Thermomyces lanuginosa expressed in Aspergillus oryzae.

Grindamyl™ EXEL 66 (obtained from Danisco A/S) is a mixture of lipase (EC 3.1.1.3) from Thermomyces lanuginosa expressed in Aspergillus oryzae and a fungal xylanase (EC 3.2.1.8) from Aspergillus niger.

Lipopan F™ (Lipopan F BG) (obtained from Novozymes) is according to its producer (Novozymes) a purified lipolytic enzyme from Fusarium oxysporum produced by submerged fermentation of a genetically modified Aspergillus oryzae microorganism. According to its producer, Lipopan F has inherent activity toward phospholipids, glycolipids and triglycerides.

Recipes/Procedures

High Volume Tweedy

Recipe

Product Name % Gram ppm
Ijsvogel flour 3000
Water 58
Salt 60
Compressed yeast 180
Ascorbic acid 30

Procedure:

    • Dough temperature: 29° C. (dough temp.-flour temp.+4° C. water temp.)
    • Mixing: 55 WH no vacuum
    • Resting: 5 min. at room temperature
    • Scaling: 500 g (bread), 1350 g (rolls)
    • Resting: 5 min. at room temperature
    • Moulding: Puma I 13 II 18 (bread), Fortuna 3/17/7 (rolls), Glimek (moulding machine) 1:4, 2:3, 3:12, 4:14
    • Proofing: 70 min. at 43° C., 70% RH. (bread), 50 min. at 34° C., 85% RH. (rolls)
    • Baking: BAGO, 35 min.+5 min. with the steamer open at 220° C., 12 sec. steam (bread), 17 min. at 220° C., 17 sec. steam (rolls)
      Turkish Batard
      Recipe

Product name % Gram ppm
Ijsvogel flour 2000
Water 57,00
Compressed yeast 80
Salt 30
Ascorbic acid 70

Procedure:

    • Flour temperature: 15-17° C. (for trials—storage day before use at 15° C.)
    • Mixing: 35 min. After 25 min. acid salt
    • After 30 min. add yeast
    • Dough temp.: 23-25° C.
    • Resting: 30 min. Bulk rest on table (table=22° C. & 80% RH)
    • Scaling 300 g. pieces
    • Rounding: By hand
    • Resting: 25 min. on table (table=22° C. & 80% RH) . . . start clock when scaling starts
    • Molding=Glimek: 1:5, 2:4, 3:15, 4:10 . . . 10 in innerpos.
    • Proofing: 60 min & 90 min. for this trial at 30° C. & 85% RH
    • Shock test
    • Baking: 20 min. in Bago1 & 25 min. in Bago2 . . . the last 5 min. is with the damper open for both ovens.
    • Bago1: 250° C. start temp. 5 sec. steam with damper open. Oven temp. clown to 230° C. at once. Close damper after 11% min.
    • Bago2: 275° C. start temp. 8 sec. steam with damper open. Oven temp. down to 260° C. at once. Close damper after 11% min.
      Crispy Rolls
      Recipe:

Product Name % Gram ppm
Danish silver flour 2000
Water 58/60
Compressed yeast 120
Salt 32
Sugar 32
Ascorbic acid 40

Procedure:

    • Mixing: Diosna 2+5 min. (depending on flour)
    • Dough temperature: 26° C.
    • Scaling: 1350 g
    • Resting: 10 min. at 30° C. in heating cabinet
    • Moulding: Fortuna 3/17/7
    • Proofing: 45 min alternatively 90 min at 34° C., 85% RH.
    • Baking: 18 min. at 220° C., 8 sec. steam (Bago-oven), 7 sec. steam (Wachtel-oven)
    • (MIWE program 28) (0.35 liter steam, 15 min. at 2000° C., ½ min. at 2200° C.)
      US Toast

Here a sponge as a pre-mix is prepared, to all of which is then added the dough.

Recipe
Gr %
US Flour 900.000 g 50.000%
Sponge: Water 900.000 g 50.000%
Dry 23.400 g  1.300%
Yeast
Yeast Food 5.400 g  0.300%
Enzyme 0.054 g  0.003%
Complex
ADA 0.036 g  0.002%
US Flour 900.000 g 50.000%
Dough: Water 234.000 g 13.000%
Dry 25.200 g  1.400%
Yeast
Sugar 153.000 g  8.500%
Salt 43.200 g  2.400%
Shortening (fat) 36.000 g  2.000%
Sod.Prop. 8.100 g  0.450%
Dimodan SDM-T (P100/B) 9.000 g  0.500%
Asc. Acid. 0.072 g  0.004%
′→ (=7,200 g to
1000 ml.
Take 10 ml.
from the
solution)
Total flour amount: 1.800,000 g.
Datem 22-CA-60 4.500 g 0.2500%
S685 300 PPM
H640 20 PPM
TS-E 662 100 PPM

    • Care has to be taken with the water amount added from asc. acid solution and other water based solutions ex. enzymes.
    • The extra added water amount should be be withdrawn from the water amount on the Dough-side of the recipe.

The enzyme complex is a mix of alpha amylase and amyloclucosidase.

DIMODAN SDM-T (P100/B) (obtained from Danisco A/S) is a distilled monoglyceride.

Procedure:

For the Sponge:
Water Temp.: 25° C.
Hobart mixer
Step 1, 1 min.
Step 2, 1 min.
Step 3, 1 min.

Fermentation: 2 h & 15 min. 40° C. & 80% RH (relative humidity) 45 min. in freezer.

    • For the Dough:
    • Mix all ingredients together
    • Diosna-Mixer: Speed 1, 120 secs & Speed 2, 450 secs (or 28 degrees dough temp.)
    • On table—rest 5 min.
    • Weigh out the breads at 450 g pr. bread—rest 5 min.

Glimek (moulding machine) adjustments: 1, 2, 14, 11—& 9 cm—read on outer position.

Fermentation:

    • 1 h & 10 min. 45 degrees Celsius & 90% RH

Bake-off:

    • Start temp.=250 degrees Celsius in 25 min.

Insert the breads and adjust bake-off temperature to 200 degrees Celsius at once.

Baking Trials

In each trial the dough characteristic, stickiness and all over bread score have been evaluated. The dough characteristic is a total of three different parameters: dough extensibility evaluated just after mixing and again after resting and stickiness after resting. Each parameter has been evaluated by bakers on a scale from 1-10, where 10 are the best. The score in the examples are a total of these different scores.

Stickiness evaluation has been subjectively evaluated by bakers just after mixing on a scale from 1 to 10, where 10 is the best, meaning non sticky.

All over bread score is a total of an evaluation made on bread crust, -crumb, possible capping and all over energy of the bread. Again each parameter is evaluated on a scale from 1-10, where 10 is the best.

EXAMPLE 7 Testing Alternatives in Tweedy Bread (UK Procedure)

The breads were rested for 70 min each and after a full proofing, each bread was shock treated in order to evaluate the shock resistance and thereby the dough stability.

In the baking trials, both pure enzyme solutions and combinations of DATUM and enzymes were tested as alternative to Lipopan F.

Baking Trials 4969-29

All
Specific Shocked Dough Dough over
volume, volume, charac- sticki- bread
Test ccm/g ccm/g teristic ness score
0.4% PANODAN GB 5.6 4.64 15 4 29
0.2% PANODAN GB, 5.75 4.92 14 4 30
100 ppm
GRINDAMYL H121,
100 ppm TS-E 662
100 ppm TS-E 662, 5.57 4.47 14 4 20
100 ppm
GRINDAMYL H121,
100 ppm
GRINDAMYL
EXEL 16
40 ppm Lipopan F 5.7 4.6 13 4 29
0.2% PANODAN GB, 5.88 4.6 14 4 27
20 ppm Lipopan F
20 ppm Lipopan F, 5.65 4.78 14 4 29
100 ppm TS-E 662,
100 ppm
GRINDAMYL H121
40 ppm Lipopan F, 5.79 4.82 13 4 29
100 ppm TS-E 662,
100 ppm
GRINDAMYL H121

From the results it can be concluded that PANODAN GB results in a better crust of the product and a product.

The combination of PANODAN GB in combination with xylanase and hexose oxidase yields a beneficial effect.

When using DATEM and/or HOX in combination with GRINDAMYL EXEL 66 the volume is increased significantly and the crust is considerably improved. The test with 0.1% PANODAN GB 100 ppm GRINDAMYL EXEL 66 and 100 ppm TS-E 662 (HOX), gave a significantly good result at the same level as 0.4% PANODAN GB. Use of DATEM clearly gives a significantly positive effect on the crust as compared to pure enzyme solutions.

EXAMPLE 8 Testing Alternatives in Turkish Batard

Baking Trials 7258-2

Specific Dough Dough All over
volume, characteristic stickiness bread
Test ccm/g * ** score***
15 ppm Lipopan F, 60 5.01 14 4 33
ppm TS-E 680
40 ppm Lipopan F 3.78 15 5 32
100 ppm TS-861* 5.03 16 5 44
*A combination of fungal xylanase. 1,3 triglyceride degrading lipase and hexose oxidase.

Both from the specific volume in the table as well as the pictures shown in FIGS. 1-3 it can be concluded that TS-E 861 performs better.

EXAMPLE 9 Testing Alternatives in Crispy Rolls

The rolls were fermented at two different fermentation times—45 and 90 min in order to stress the system and thereby give a better picture of the dough strengthening effect of the products. In general it can be said that 90 min of fermentation for a small crispy roll is quite long.

Baking Test: 4969-28

Specific Specific Dough All over
volume volume Dough sticki- bread
45 min, 90 min, characteristic ness score
Test ccm/g ccm/g * ** ***
0.3% PANODAN 7.15 8.48 14 5 25
A2020
30 ppm Lipopan F 6.83 8.1 14 4 26
100 ppm TS-E 662, 6.98 8.98 14 5 27
100 ppm
GRINDAMYL
H121, 100 ppm
GRINDAMYL
EXEL 16

From the results it can be seen that use of the combination of xylanase, 1,3 triglyceride degrading lipase and hexose oxidase produces beneficial results.

In short fermentation times (45 min.) at certain concentrations PANODAN A2020 and Lipopan F gave comparable volume results. However, 0.3% PANODAN A2020 showed better results with regard to crispiness of the crust and a better dough stability in general. We found that Lipopan F often gave a slightly more “wet” crust.

Using HOX in combination with GRINDAMYL EXEL 66 and PANODAN 660 results in an increase in dough stability.

With prolonged fermentation times (90 min.) all buns become relatively unstable. At some concentrations PANODAN A2020 does, however, give the best result.

EXAMPLE 10 Testing Alternatives in US Toast

Test of Lipopan F in a US sponge and dough using flour from Mexico—hard wheat type. The breads have all been fully proofed and after that each bread have been shock treated in order to evaluate the shock resistance and thereby the dough stability.

Baking Trials 7230-1:

Specific Shocked
Test volume, ccm/g volume, ccm/g
0.5% PANODAN 521 6.88 5.47
10 ppm Lipopan F 6.16 5.36
20 ppm Lipopan F 6.44 5.30
40 ppm Lipopan F 6.28 5.52
0.25% PANODAN 521, 7.15 5.74
20 ppm GRINDAMYL H
640, 100 ppm TS-E 662

From these tests it is clear that the use of Hox results in a far better dough stability and consequently an increase of volume.

All publications mentioned in the above specification are herein incorporated by reference. Various modifications and variations of the described methods and system of the present invention may be apparent to those skilled in the art without departing from the scope and spirit of the present invention. Although the present invention has been described in connection with specific preferred embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the described modes for carrying out the invention which are obvious to those skilled in biochemistry and biotechnology or related fields are intended to be within the scope of the claims.

Summary Paragraphs

Some aspects of the present invention are now described by way of Summary Paragraphs.

1. A method of improving the rheological properties of a flour dough and the quality of the finished product made from the dough, comprising adding to the dough ingredients, dough additives or the dough an effective amount of an oxidoreductase which is at least capable of oxidizing maltose.

2. A method according to paragraph 1 wherein the oxidoreductase is hexose oxidase.

3. A method according to paragraph 2 wherein the hexose oxidase is derived from a source selected from an algal species, a plant species and a microbial species.

4. A method according to paragraph 3 wherein the hexose oxidase is derived from Chondrus crispus.

5. A method according to paragraph 2 wherein hexose oxidase is added in an amount which is in the range of 1 to 10,000 units per kg of flour.

6. A method according to paragraph 5 wherein the hexose oxidase is added in an amount which is in the range of 10 to 1000 units per kg of flour.

7. A method according to paragraph 1 or 2 wherein the resistance to extension of the dough in terms of the ratio between the resistance to extension (height of curve, B) and the extensibility (length of curve, C), i.e. the B/C ratio, as measured by the AACC method 54-10 is increased by at least 10% relative to that of an otherwise similar dough not containing oxidoreductase.

8. A method according to paragraph 1 wherein the finished product is bread.

9. A method according to paragraph 1 wherein the finished product is a noodle product

10. A method according to paragraph 1 wherein the finished product is an alimentary paste product.

11. A method according to paragraph 1 wherein at least one further enzyme is added to the dough ingredients, dough additives or the dough.

12. A method according to paragraph 11 wherein the further enzyme is selected from the group consisting of a cellulase, a hemicellulase, a xylanase, a starch degrading enzyme, a glucose oxidase, a lipase and a protease.

13. A dough improving composition comprising an oxidoreductase which is at least capable of oxidising maltose and at least one further dough ingredient or dough additive.

14. A composition according to paragraph 13 wherein the oxidoreductase is derived from a source selected from an algal species, a plant species and a microbial species.

15. A composition according to paragraph 14 wherein the oxidoreductase is hexose oxidase.

16. A composition according to paragraph 15 wherein the hexose oxidase is derived from Chondrus crispus.

17. A composition according to paragraph 13 which is a pre-mixture useful for preparing a baked product or in making a noodle product or an alimentary paste product.

18. A composition according to paragraph 13 which comprises an additive from the group consisting of an emulsifying agent and a hydrocolloid.

19. A composition according to paragraph 18 wherein the hydrocolloid is selected from the group consisting of an alginate, a carrageenan, a pectin and a vegetable gum.

20. A method of preparing a bakery product the method comprising preparing a flour dough to which is added an effective amount of an oxidoreductase which is at least capable of oxidising maltose, and baking the dough.

21. A method according to paragraph 20 wherein the specific volume of the bakery product is increased relative to an otherwise similar bakery product prepared from a dough not containing oxidoreductase.

22. A method according to paragraph 21 wherein the specific volume is increased by at least 20%.

23. A method according to paragraph 20 wherein at least one further enzyme is added to the dough.

24. A method according to paragraph 20 wherein the further enzyme is selected from the group consisting of a cellulase, hemicellulase, a xylanase, a starch degrading enzyme, a glucose oxidase, a lipase and a protease.

25. A method according to paragraph 20 wherein the oxidoreductase is hexose oxidase.

26. A method of preparing a flour dough-based food product, comprising adding to the dough an effective amount of a maltose oxidising oxidoreductase.

27. A method according to paragraph 26 wherein the oxidoreductase is hexose oxidase.

Patent Citations
Cited PatentFiling datePublication dateApplicantTitle
US2783150Sep 25, 1952Feb 26, 1957Pfizer & Co CTreatment of flour with glucose oxidase
US2888385Nov 27, 1953May 26, 1959Grandel FelixProcess of making a preparation containing lipases and oxidases
US3260606Apr 29, 1964Jul 12, 1966Taiyo Food Co LtdEnzymatic treatment of egg
US3368903Feb 18, 1966Feb 13, 1968Vanderbilt Co R TBaked goods dough and method
US3520702May 1, 1967Jul 14, 1970Menzi RobertMethod of making dried pasta having a protein network that withstands cooking
US3634195Sep 8, 1969Jan 11, 1972Miles LabProduction of lipase
US3652397Jan 8, 1970Mar 28, 1972Lever Brothers LtdPreparation of phosphatides
US3677902Jun 21, 1966Jul 18, 1972Novo Terapeutisk Labor AsPreparation of amyloglucosidase
US3852260Feb 2, 1973Dec 3, 1974Astra Nutrition AbProcess for preparing fish protein using propanol or butanol
US3973042May 10, 1974Aug 3, 1976Cornell Research Foundation, Inc.Flavor development by microbial lipases in pasteurized milk blue cheese
US4034124Nov 19, 1975Jul 5, 1977Lever Brothers CompanyPhospholipase a modified phospholipoprotein, oils, water
US4065580Nov 17, 1975Dec 27, 1977Gb Fermentation Industries, Inc.Lipolytic enzyme flavoring system
US4160848Apr 18, 1977Jul 10, 1979Pennwalt CorporationAntistaling agent for bakery products
US4202941Apr 19, 1978May 13, 1980Kyowa Hakko Kogyo, Co. Ltd.Aspergillus or neurospora cultrues
US4399218Jan 23, 1981Aug 16, 1983Boehringer Mannheim GmbhMethod and reagent for the determination of glycerin
US4567046Nov 3, 1983Jan 28, 1986Kyowa Hakko Kogyo Co., Ltd.Bread or other cereal-based food improver composition involving the addition of phospholipase A to the flour
US4575487May 25, 1983Mar 11, 1986Miles Laboratories, Inc.In glucoamylase preparation
US4683202Oct 25, 1985Nov 27, 1990Cetus CorpTitle not available
US4689297Mar 5, 1985Aug 25, 1987Miles Laboratories, Inc.Dust free particulate enzyme formulation
US4707291Jun 3, 1986Nov 17, 1987Lever Brothers CompanyAnionic or nonionic surfactants
US4707364Nov 1, 1985Nov 17, 1987Miles Laboratories, Inc.Freeze drying, lipase
US4708876Apr 22, 1986Nov 24, 1987Fuji Oil Company, LimitedMethod for preparing cheese flavor concentrate
US4798793Jul 31, 1987Jan 17, 1989Novo Industri A/SImmobilized Mucor miehei lipase for transesterification
US4808417Dec 8, 1987Feb 28, 1989Toa Pharmaceutical Co., Ltd.Feed additive for fish cultivation
US4810414Aug 28, 1987Mar 7, 1989Novo Industri A/SEnzymatic detergent additive
US4814331Sep 25, 1985Mar 21, 1989Nederlandse Organisatie Voor Et Al.Method for controlling vascular withering diseases in plants, especially dutch elm disease
US4818695Jul 31, 1987Apr 4, 1989Novo Industri A/SImmobilized Mucor miehe lipase for transesterification
US4826767Oct 31, 1986May 2, 1989Novo Industri A/SEnzymatic synthesis of waxes
US4865866Apr 26, 1988Sep 12, 1989Lever Brothers CompanyPlastic emulsion food product with a hardstock fat
US4904483Feb 15, 1989Feb 27, 1990Novo Industri A/SMethod for production of an upgraded coconut product
US4916064Oct 6, 1986Apr 10, 1990Cpc International Inc.Carbohydrate refining process and novel enzyme compositions suitable for use therein
US5059430Sep 12, 1990Oct 22, 1991Enzyme Bio-Systems Ltd.Acid stable microbial alpha-amylase and a bacterial alpha-amylase; improves softness; added to sponge and/or dough
US5094951Jun 19, 1989Mar 10, 1992Chiron CorporationGenetic engineering
US5108765Mar 23, 1990Apr 28, 1992Van Den Bergh Foods Co.Containing cellulase, peroxidase and xylanase
US5112624Jun 19, 1990May 12, 1992Knoll AgPrevention of digestive disturbances in herbivores
US5213968Dec 31, 1991May 25, 1993Nestec S.A.Process for preparing emulsifying agents
US5219733Aug 7, 1990Jun 15, 1993Yoshikawa Oil & Fat Co., Ltd.Reacting sterol or fatty alcohol with fatty acid or ester in contact with microorganism-derived lipase in water or mixture of water and organic solvent
US5219744Jun 26, 1991Jun 15, 1993Ajinomoto Co., Inc.Process for modifying fats and oils
US5232846Sep 3, 1992Aug 3, 1993Unitika Ltd.Method for producing a thermostable lipoprotein lipase from streptomyces
US5264367May 14, 1992Nov 23, 1993Rohm GmbhEnzymatic treatment of edible oils
US5273898Oct 16, 1992Dec 28, 1993Noro Nordisk A/SThermally stable and positionally non-specific lipase isolated from Candida
US5288619Jun 11, 1992Feb 22, 1994Kraft General Foods, Inc.Enzymatic method for preparing transesterified oils
US5290694Feb 28, 1989Mar 1, 1994Amano Pharmaceutical Co., Ltd.Recombinant DNA, bacterium of the genus Pseudomonas containing it, and process for preparing lipase by using it
US5318785Feb 8, 1993Jun 7, 1994Elf Atochem North America, Inc.Benzoyl peroxide to improve the performance of oxidants in breadmaking
US5378623Jun 15, 1993Jan 3, 1995Sankyo Company, LimitedPhospholipase A1, process for its preparation and the use thereof
US5451413Sep 16, 1993Sep 19, 1995Gist-Brocades, B.V.Yeast derivative and method to improve bread quality
US5516689Apr 30, 1993May 14, 1996Solvay Enzymes, Inc.Method for the treatment of sticky cotton fiber with transglucosidase from Aspergillus niger
US5523237Feb 6, 1992Jun 4, 1996Novo Nordisk A/SProtein preparations
US5536661Mar 7, 1994Jul 16, 1996Novo Nordisk A/SProcess for the production of protein products in aspergillus
US5558781Nov 16, 1994Sep 24, 1996Metallgesellschaft AktiengesellschaftProcess for enzymatically degumming vegetable oil
US5650188Oct 28, 1994Jul 22, 1997Gist-Brocades, B.V.Baking improver compositions
US5677160Dec 14, 1994Oct 14, 1997Henkel CorporationPrehydrolysis in presence of lipase; pressure splitting
US5695802Jun 13, 1995Dec 9, 1997Firmenich SaFlavoring composition and process
US5716654Dec 23, 1994Feb 10, 1998Gist-Brocades, N.V.Homogeneous mixture comprising a baking promoter coated on as a film or adhered particles; shelf life, storage stability
US5763383Dec 22, 1993Jun 9, 1998Novo Nordisk A/SAlkaline lipases
US5766912Apr 20, 1994Jun 16, 1998Novo Nordisk A/SHumicola lipase produced in aspergillus
US5776741Feb 21, 1995Jul 7, 1998Novo Nordisk A/SMethod of enzyme immobilization on a particulate silica carrier for synthesis inorganic media
US5814501Feb 15, 1995Sep 29, 1998Genencor International, Inc.Process for making dust-free enzyme-containing particles from an enzyme-containing fermentation broth
US5821102Jan 21, 1997Oct 13, 1998Novo Nordisk Biotech Inc.Recombinant production of lipase for use in alkaline detergents
US5827719Oct 26, 1995Oct 27, 1998Novo Nordisk A/SDna sequence isolated for saccharomyces cerevisiae dsm
US5830736Oct 26, 1995Nov 3, 1998Novo Nordisk A/SLipolytic enzyme
US5834280Nov 7, 1996Nov 10, 1998Novo Nordisk A/SGlucose oxidases obtained from a cladosporium
US5856163Jun 3, 1993Jan 5, 1999Novo Nordisk A/SLipases from hyphozyma
US5863759Jun 5, 1995Jan 26, 1999Novo Nordisk A/SProcess for the production of protein products in aspergillus
US5869438Jun 7, 1995Feb 9, 1999Novo Nordisk A/SLipase variants
US5874558May 17, 1996Feb 23, 1999Novo NordiskVector system; greater glycosylation, improved thermostability
US5879920Dec 22, 1995Mar 9, 1999Genencor International, Inc.Coated enzyme-containing granule
US5892013Jun 7, 1995Apr 6, 1999Novo Nordisk A/SLipolytic enzymes have been used in detergents to remove lipid or fatty stains from clothes and other textiles.
US5914306May 1, 1992Jun 22, 1999Novo Nordisk A/SContaining nucleotide sequence having proline substituted at specified position(s)
US5916607May 27, 1997Jun 29, 1999Gist-Brocades B.V.Process for increasing the volume of a baked product
US5916619Feb 28, 1996Jun 29, 1999Nisshin Flour Milling Co., Ltd.Chemical leavening agent and atleast one enzyme selected from amylase or protease is added in the cereal flour, steaming and frying the string noodles formed by above mixture, low oil absorption, lower calories
US5919746Aug 28, 1997Jul 6, 1999Novo Nordisk A/SDerived from a strain of botryosphaeria or guignardia; high activity at high ph for use in detergent systems
US5929017Oct 26, 1995Jul 27, 1999Novonordisk A/SEnzymatic detergent composition
US5965384Jun 5, 1995Oct 12, 1999Novo Nordisk A/SCulturing aspergillus oryzae transformed with expression vector of regulatory sequences and dna sequences coding for humicola lanuginosa lipase, recovering lipase with different glycosylation and heat and/or proteolytic resistance
US5965422May 20, 1997Oct 12, 1999Rohm GmbhLysophospholipase produced from aspergillus by recombinant methods
US5976855Aug 22, 1996Nov 2, 1999Novo Nordisk A/SMethod of preparing a variant of a lipolytic enzyme
US5989599Apr 24, 1995Nov 23, 1999Nestec S.A.Process for the interesterification of phospholipids
US5990069Dec 2, 1995Nov 23, 1999Genencor International, Inc.Isolated lipase produced by fungus of species fusarium solanii characterized by specific isoelectric points and apparent moleuclar weight; use in detergent compositions to remove fat stains
US6001586Mar 29, 1996Dec 14, 1999Genencor International, Inc.Compartmentalization method for screening microorganisms
US6001640Jul 4, 1996Dec 14, 1999Roehm GmbhVegetable oil enzymatic degumming process by means of aspergillus phospholipase
US6020180Jul 8, 1998Feb 1, 2000Novo Nordisk A/SC. antarctica lipase and lipase variants
US6039983Apr 28, 1998Mar 21, 2000Novo Nordisk A/SUse of a pyranose oxidase in baking
US6066482Mar 13, 1998May 23, 2000Cornell Research Foundation, Inc.Acyltransferase and gene encoding acyltransferase
US6074863Jul 5, 1993Jun 13, 2000Novo Nordisk A/SHydrolytic enzyme for the treatment of paper products or use as a detergent enzyme
US6103505Dec 9, 1997Aug 15, 2000Novo Nordisk A/SPolypeptide exhibiting phospholipase activity; for the hydrolysis of fatty acyl groups in food production
US6110508Jan 4, 1999Aug 29, 2000Novo Nordisk A/SUse of lipase in baking
US6140094Jan 8, 1998Oct 31, 2000Rohm GmbhAspergillus hydrolase; for degumming vegetable oils; for preparation of foods
US6143543Nov 21, 1997Nov 7, 2000Danisco A/SNucleotide sequence coding a hydrolase; for use in food processing
US6143545Sep 1, 1999Nov 7, 2000Novo Nordisk A/SNucleotide sequence coding phospholipase; for removal of non-hydratable phosphatides from crude oils
US6146869Oct 21, 1999Nov 14, 2000Novo Nordisk Biotech, Inc.Nucleotide and amino acid sequence of aspergillus enzyme
US6156548Dec 11, 1998Dec 5, 2000Novo Nordisk A/SFluidizing particulate porous carrier in bed, atomizing liquid containing enzyme into bed, and removing volatile components; for lipases used in transesterification of triglycerides
US6180406Jun 17, 1998Jan 30, 2001Maxygen, Inc.Methods for generating polynucleotides having desired characteristics by iterative selection and recombination
US6251626Jun 4, 1996Jun 26, 2001Bioteknologisk InstitutGenerating enzymatic polypeptide; insert expression vector into cell, culture cells, recover enzymatic polypeptide
US6254645Nov 5, 1999Jul 3, 2001Genencor International, Inc.Enzymatic modification of the surface of a polyester fiber or article
US6254903Nov 13, 1997Jul 3, 2001Roehm GmbhProcess for making baked articles that retain freshness
US6344328Apr 24, 2000Feb 5, 2002Diversa CorporationMethod for screening for enzyme activity
US6350604Oct 22, 1998Feb 26, 2002Novozymes A/SAlkaline lipolytic enzyme
US6358543Jun 4, 1996Mar 19, 2002Danisco A/SMethod of improving the properties of a flour dough, a flour dough improving composition and improved food products
US6361974Mar 27, 2000Mar 26, 2002Diversa CorporationExonuclease-mediated nucleic acid reassembly in directed evolution
Non-Patent Citations
Reference
1"Baking Science & Technology", E.J. Pyler (1982), vol. 1, pp. 314-316.
2"DEEO®, "A glucose oxidase and catalase enzyme system product sheet from Miles laboratories-Enzymes from Miles (technical Information) (1976), 5 pages.
3"Effect of Different Hexose Oxidase and Other Oxide Reductases in Dough", Experimental Data Submitted by Applicants in European Counterpart Application 96917368.
4"Enzyme Function", Experimental Report from Novo Nordisk, Mar. 13, 1997, 2 pages.
5"Enzyme Nomenclature (Recommendations of the Nomenclature Committee of the International Union of Biochemistry and Molecular Biology on the Nomenclature and Classification of Enzymes)" (1992), p. 56.
6"Enzyme Nomenclature 1984 (Recommendations of the Nomenclature Committee of the International Union of Biochemistry on the Nomenclature and Classification of Enzyme-Catalysed Reactions)" (1984), pp. v, ix, and 50-51.
7"Enzyme Technology in Flour Milling and Baking", Baking Industry Europe (Alan Gordon, editor), S. Haarasilta and T. Pullinen (1993), pp. 49-52.
8"Enzymes in Food Processing", 2nd Ed. By G. Reed, Universal Foods Corporation, Academic Press (1975), p. 222-229.
9"Glucose Oxidase: Production, Properties, Present and Potential Applications", Soc. Chem. Ind. (Londen), (1961), L.A. Undetkofler, p. 72-86.
10"Gluzyme.(TM)" product sheet from Novo Nordisk Enzyme Process Division, Jan. 1994, 2 pages. . Derwent Publications Ltd., London, GB; AN 73-30288u XP002012361 & JP, A48016612 (EISAI Co. Ltd).
11"Methods in Enzymology", Biomass Part B Glucose Oxidase of Phanerochaete chrysosporium, R.L. Kelley and C.A. Reddy (1988), 161, pp. 306-317.
12"Novel Enzyme Combinations A New Tool to Improve Baking Results", Agro-Industry Hi-Tech. S. Haarasilta and T. Pullinen, (May/Jun. 1992), p. 12-13.
13"Properties and Applications of the Fungal Enzyme Glucose Oxidase", reprinted from "Proceedings of the International Symposium on Enzyme Chemistry", Tokyo and Kyoto, (1957) L.A.
14"Technology of Cereals (with special reference to wheat)", 2nd Ed., Pergamom Press Ltd. N. L. Kent, (1975), pp. iv-v, 48-49, and 72-73.
15"The Oxidation of Glucose and Related Compounds by Glucose Oxidase from Aspergillus niger", Biochemistry, Pazur et al., vol. 3(4), 1964, 578-583.
16"DEEO®, "A glucose oxidase and catalase enzyme system product sheet from Miles laboratories—Enzymes from Miles (technical Information) (1976), 5 pages.
17"DEEOθ" A glucose oxidase and catalase enzyme system product sheet from Miles laboratories—Enzymes from Miles (technical Information) (1976), 5 pages.
18"Enzymes in Food Processing", 2nd Ed. by G. Reed, Universal Foods Corporation, Academic Press (1975), p. 222-229.
19"Glucose Oxidase: Production, Properties, Present and Potential Applications", Soc. Chem. Ind. (Londen), (1961), L.A. Underkofler, p. 72-86.
20"Gluzyme.™" product sheet from Novo Nordisk Enzyme Process Division, Jan. 1994, 2 pages. . Derwent Publications Ltd., London, GB; AN 73-30288u XP002012361 & JP, A48016612 (EISAI Co. Ltd).
21"Gluzyme™" product sheet from Novo Nordisk Enzyme Process Division, Jan. 1994, 2 pages.
22"Properties and Applications of the Fungal Enzyme Glucose Oxidase", reprinted from "Proceedings of the International Symposium on Enzyme Chemistry", Tokyo and Kyoto, (1957) I.A. Underkofler, (1958), pp. 486-490.
23"Unique Chance for Better Bread," Direct, A Newsletter from Danisco Ingredients, (1996).
24AACC Method 36-01 A.
25AACC Method 54-10.
26Acker, L. "Die Lipide des Getreides, ihre Zusammense und inre Bedeutung", Getreide Mehl Brot (1974) 28:181-187.
27Adamzcak, Marek, et al., "Application of Enzymatic Glycerolysis for Production of Monoglycerides from Waste Fats", Polish Journal of Food and Nutrition Science, Mar. 1994.
28Adhikari, B., et al., "Stickiness in Foods: A Review of Mechanisms and Test Methods", International Journal of Food Properties, vol. 4, No. 1, 2001.
29Agarwal et al., "Lipase Activity of Some Fungi Isolated from Groundnut", Current Science, Dec. 5, 1984, vol. 53, No. 23.
30Aires-Barros et at (1994) Isolation and purification of lipases, Cambridge Unversity Press.
31Aisaka, Kazuo et al., "Production of Lipoprotein Lipase and Lipase by Rhizopus japonicu", Agri. Biol. Chem., vol. 43, No. 10, pp. 2125-2129, 1979.
32Akoh, Casimir C., et al., "GDSL family of serine esterases/lipases" Progress in Lipid Research, vol. 43, 2004, pp. 534-552.
33Allan Svendsen et al., "Biochemical properties of cloned lipases from the Pseudomonas family", Biochimica et Biophysica Acta, vol. 1259, 1995, pp. 9-17.
34Allen, R.M. et al., "Low-Level Electrochemical Detection of Glucose Oxidase and a Glucose Oxidase Conjugate", Biosensors and Bioelectronics, 10:621-631 (1995).
35Al-Obaidy, K A, Dissertation Abstracts International B (1987) vol. 47(9) 3597, order No. DA8624641, pp. 266.
36Amano Enzyme Inc. (2004). Http://www.amano-enzyme.co.jp/english/productuse/oil—fat.html. Dato Jun. 21, 2004.
37Amano Enzymes "Enzymes for Gastrointestinal Digestion" Oct. 1997.
38Amano Enzymes, Amano Enzyme Europe Ltd, Sep. 1994.
39Amendment in response to Office Action dated Jan. 3, 1999.
40Amin, Neelam S., et al., "Direct transformation of site-saturation libraries in Bacillus subtilis", BioTechniques, Dec. 2003, 35:1134-1140.
41Amino acid composition of lipases.
42Andersson, L., et al., "Hydrolysis of galactolipids by human pancreatic lipolytic enzymes and duidenal contents", Journal of Lipid Research, 1995, vol. 36, pp. 1392-1400.
43Andreas Sander, Eberhand Eilers, Andrea Heilemann, Edith von Kreis.Fett/lipid 99 (1997) Nr. 4, 115-120.
44Angelino, S.A.G.F., et al., "The first European Symposium on Enzymes and Grain Processing".
45An-I Yeh et al., "Effects of Oxido-reductants on rheological properties of wheat flour dough and comparison with some characteristics of extruded noodles", Cereal Chemistry, 1999, vol. 76, No. 5, pp. 614-620.
46AOCS Introduction to the Processing of Fats and Oils, American Oil Chemists, p. III-16-III-19 (2003).
47Application of F. oxysporum phospholipase (FoL) in baking.
48AR Patent Application-Variantes De Lipasa Y Metodos Para Su Preparacion, May 21, 1996, Argentina.
49AR Patent Application—Variantes De Lipasa Y Metodos Para Su Preparacion, May 21, 1996, Argentina.
50AR Patent—Lipasa Alcalina, Argentina.
51AR Patent—Variantes De Lipasa Y Metodos Para Su Preparacion, Argentina.
52Arbige, Michael A et al, Novel lipase for cheddar cheese flavor development.
53Archer, David B., et al., "Proteolytic degradation of heterologous proteins expressed in Aspergillus Niger", Biotechnology Letter, vol. 14, No. 5, May 1992, pp. 357-362.
54Arcos J.A. et al, "Quantative Enzymatic Production of 6.0-Acylglucose Esters", Biotechnology and Bioengineering 1998 57(5).
55Arpigny Jean Louis et al, "Bacterial lipolytic enzymes: Classification and properties", Biochemical Journal, vol. 343, No. 1, Oct. 1, 1999, pp. 177-183, XP002375631.
56Arskog and Joergensen, "Baking of prior art lipases from Candida cylindraceaa and Aspergillus foeditus and their activity on galactolipids on dough," Novozymes Report 2005.
57Arskog and Joergensen, "Baking of prior art lipases from Humicola lanuginosea, Aspergillus tubigensis, Rhizopus delemar and Thizomucor miehei, and their activity on galactolipids on dough," Novozymes Report 2005.
58Assignment Document for Enzymatisk detergent additiv, detergent og vaskemetode.
59Atomi, et al.; "Microbial Lipases—from Screening to Design"; pp. 49-51.
60August C.A.P.A. et al. "The use of genetic engineering to obtain efficient production of porcine pancreatic phospholipase A2", Biochimica et Biophysica Acta, vol. 1089, 1991, pp. 345-351.
61Aunstrup, Knud et al., "Production of Microbiol Enzymes", Microbiol Technology, vol. 1.
62Aust K., Applications of lecithin in bakery foods, AIB Research Technical Bulletin, vol. XV, issue 12, Dec. 1993, p. 1-6.
63Ausubel, Frederick M., et al., "Short Protocols in Molecular Biology—A Compendium of Methods from Current Protocols in Molecular Biology", 1995, John Wiley & Sons, Inc.
64Bachmatova, I., et al., "Lipase of Pseudomonas mendocina 3121-1 and its Substrate Specificty", Biologija, 1995.
65Bailey's Industrial Oils and Fat Products, vol. 2, 4th Edition, John Wiley and Sons, New York pp. 97-173.
66Bak et al., "A Method for Testing the Strengthening Effect of Oxidative Enzymes in Dough", presented at a symposium entitled "Wheat Structure, Biochemistry and Functionality", Reading UK, Apr. 10-12, 1995.
67Bakezyme PH 800.
68Balashev, Konstantin, Surface studies of enzymes using Atomic force microscopy (AFM).
69Balcao V.M., Pavia A.L. Malcata F.X., Enzyme Microb Technhol, May 1, 1996; 18(6):392-416.
70Balcao, Victor M and Malcata F. Xavier (1998), Biotechnology Advances, vol. 16, No. 2, pp. 309-341.
71Ballance, D.J., et al., "Transformation of Aspergillus Nidulans by the orotidine-5′-phosphate decarboxylase gene of neurospora crassa", Biochemical and biophysical Research Communications, vol. 112, No. 1, 1983, pp. 284-289.
72Ballance, Molecular Industrial Mycology, Systems and Applications for Filamentous Fungi, Leong and Berka (eds.), Marcel Dekker Inc, New York 1991, pp. 1-29.
73Barbesgaard, Peder et al Applied Microbiology and Biotechnology (1992) 36: 569-572.
74Barkholt, V. and A.L. Jensen, 1989, Amino Acid Analysis: Determination of Cysteine plus Half-Cysteine in Proteins after Hydrochloric Acid Hydrolysis with a Disulfide Compound as Additive, Analytical Biochemistry, 177:318-322.
75Barnes, P.J., "Lipids in Cereal Technology", Food and Science Technology, Academic Press, 1983, NZNA-0006131.
76Basrl, M., et al., "Amidination of Lipase with Hyrdophobic Imidoesters", JAOCS, vol. 69, No. 6, Jun. 1992.
77Bateman A and Haft DH (2002) Brief Bioinform 3, 236-245.
78Bateman A et al, (2002) Nucleic Acids Res. 30, 276-280.
79Bean and Hassid, 1956, J. Biol. Chem. , 218: 425-436.
80Bean and Hassid, 1956, J. Biol. Chem., 218: 425-436.
81Bean et al., "Carbohydrate Metabolism of Citrus Fruits", Journal of Biological Chemistry (1961) 236: 1235-1240.
82Becker T. "Separation and Purification Processes for Recovery of Industrial Enzymes" in R.K., Singh, S.S.H. Rizvi (eds): Bioseparation processes in Foods, Marcel Dekker, New York, pp. 427-445.
83Bedre Brod med nyt enzym.
84Bekkers et al, The use of genetic engineering to obtain efficient production of porcine pancreatic phospholipase A2 by Saccharomyces cerevisiae, (1991) Biochim Biophys Acta 1089(3), 345-51.
85Bengtsson Olivecrona Gunilla et al. Phospholipase activity of milk lipoprotein lipase, Methods in Enzymology, vol. 197, 1991.
86Bentley S D et al, Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2), Nature vol. 417, 2002, pp. 141-147.
87Berger K.G. (1990) Recent developments in palm oil. In Oleagineux 45:437-443.
88Berks, Ben C., "A common export pathway for proteins binding complex redox cofactors?" Molecular Microbiology, 1996, vol. 22, pp. 393-404.
89Beucage S.L. et al, (1981) Tetrahedron Letters 22, p. 1859-1869.
90Bieleski R.L., Chapter 5, Sugar Alcohols.
91Bilyk, Alexander, et al., "Lipase-catalyzed triglyceride Hydrolysis in Organic Solvent", pp. 320-323, JAOCS, vol. 68, No. 5, May 1991.
92Biocatalysts, Limited, Product Sheet for Lipomod(TM) 627P-L627P.
93Biotekkomet falder hardt til jorden.
94Birch et al., "Evidence of Multiple Extracellular Phospholipase Activities of Aspergillus fumigatus", Infection and Immunity, Mar. 1996, vol. 64, No. 3, 1996.
95Birgitte Hugh-Jensen et al., "Rhizomucor miehei Triglyceride Lipase is Processed and Secreted from Transformed Aspergillus oryzae", Lipids, vol. 24, No. 9, 1989, NZAS-0214608.
96Biswas, et al., "Interfacial Behavior of Wheat Puroindolines: Study of Adsorption at the Air-Water Interface from Surface Tension Measurement Using Wilhelmy Plate Method", Journal of Colloid and Interface Science, vol. 244, pp. 245-253, 2001, NZAS - 0204251.
97Bjorkling, F., et al., "Lipase Catalyzed Organic Synthesis", S. Servie (ed.), Microbial Reagents in Organic Synthesis, pp. 249-260, 1992, NZAS-0214653.
98Bjorkling, Frederik, et al., "Lipase Catalyzed Synthesis of Perozycarboxylic Acids and Lipase Mediated Oxidations", Tetrahedron, vol. 48, No. 22, pp. 4587-4592, 1992.
99Bjorkling, Frederik, et al., "Lipase -mediated Formation of Peroxycarboxylic acids used in Catalytic Epoxidation of Alkenes", J. Chem. Soc., Chemical Communications, Issue 19, 1990.
100Bjurlin et al. Identification of carboxylesterase activities of commercial triacylglycerol hydrolase (lipase) preparations, Eur. J. Lipid Sci. Technol. 104 (2002) 143-155.
101Blain JA et al, The Nature of Mycelial Lipolytic enzymes in filamentous fungi, Fems Microbiol. Lett., 1978, vol. 3, 85-87.
102Blecker et al, Improved emulsifying and foaming of whey proteins after enzymic fat hydrolysis, (1997) J Food Science, vol. 62, No. 1.
103Blumenthal, Cynthia Z., "Production of toxic metabolites in Aspergillus niger, Aspergillus oryzae, and Trichoderma reesei: justification of mycotoxin testing in food grade enzyme preparations derived from the three fungi", Regulatory Toxicology and Pharmacology, vol. 39, 2004, pg. 214-228.
104Boel, Esper, et al.; "Rhizomucor miehei Triglyceride Lipase is Synthesized as a Precursor"; Novo Research Institute; vol. 23; No. 7; Jul. 1988.
105Bornscheuer U T et al, Trends in Biotechnology, Elsevier Publications, Cambridge GB, vol. 20, No. 10, Oct 1, 2002, pp. 433-437.
106Bornscheuer, U.T., "Lipase-Catalyzed Syntheses of Monoacylglycerols", Enzyme and Microbial Technology, 17: 578-586 (1995).
107Bornscheuer, Uwe T., Lipase-catalyzed syntheses of monoacylglycerols, Enzyme and Microbiol Technology, vol. 17, pp. 578-586, 1995.
108Brady, Leo, et al., "A serine protease triad forms the catalytic centre of a triacylglycerol lipase", Nature, vol. 343, 1990.
109Briand et al., "Lipids," Aug. 1995, vol. 30, No. 8, pp. 747-754.
110Brockerhoff, Hans, et al., "Lipolytic Enzymes", Academic Press, 1974.
111Brumlik, Michael J., et al., "Identification of the Catalytic Triad of the Lipase/Acyltransferase from Aeromonas hydrophila", Journal of Bacteriology, Apr. 1996, vol. 178, No. 7, pp. 2060-2064.
112Brzozowski, A.M., et al., "A model for interfacial activation in lipases from the structure of a fungal lipase-inhibitor comples", Nature, vol. 351, 1991.
113Buckley J. Thomas et al, Journal of Biological Chemistry, vol. 257, No. 6, pp. 3320-3325, 1982.
114Buckley, Biochemistry 1983, 22, 5490-5493.
115Bulkacz J et al, Biochim. Biophys. Acta (1981) vol. 664, pp. 148-155.
116Bulletin of the IDF 294: 1994.
117Burdge, Graham C., et al., "A method for separation of phosphatidylcholine, triacylglycerol, non-esterified fatty acids and cholesterol esters from plasma by solid-phase extraction", British Journal of Nutrition, 2000, vol. 84, pp. 281-787.
118Butcher, Bronwyn G., et al., Microbiology, 2002, vol. 148, pp. 3983-3992.
119Buxton et al, Gene, 1985, 37:207-214.
120C.H. Poulsen, et al., "Effect and Functionality of Lipases in Dough and Bread," The First European Symposium on Enzymes and Grain Processing, pp. 204-214 (1997).
121Cammann, K., et al., "Chemical Sensors and Biosensors-Principles and Applications", Angew. Chem. Int. Ed. Engl., 30: 516-539 (1991).
122Cao, Shu-Gui, et al., "Enzymatic Preparation of Monoglycerides via Glycerolysis of Fats and Oils Catalyzed by Lipase from Pseudomonas Species" National Laboratory of Enzyme Engineering.
123Carriere et al, "Pancreatic Lipase Structure—Function Relationships by Domain Exchange", American Chemical Society-Biochemistry (1997), 36, pp. 239-248.
124Carriere, F., et al., "Pancreatic Lipase Structure-Function Relationships by Domain Exchange," Biochemistry, 36: 239-248 (1997).
125Carriére, Frédéric , et al., "Structural basis for the substrate selectivity of pancreatic lipases and some related proteins", Biochemica et Biophysica Acta, vol. 1376, pp. 417-432, 1998.
126Caruthers MH et al (1980) Nuc Acids Res Symp Ser 215-23.
127Casimir C A et al Progress in Lipid Research, 2004, pp. 534-552.
128Castello, P., et al., "Technological and Biochemical effects of exogenous lipases in breadmaking", 2nd European Symposium on enzymes in Grain Processing.
129Castello, Phillippe, et al., "Effect of exogenous lipase on dough lipids during mixing of wheat flours", Cereal Chemistry, 1998, vol. 75, No. 5, pp. 595-601.
130Castello, Phillippe, et al., "Effects of mixing conditions and wheat flour dough composition on lipid hydrolysis and oxidation levels in the presence of exogenous lipase", Cereal Chemistry, 1999, vol. 76, No. 4. pp. 476-482.
131Certificate of Analysis for Maltose Monohydrate, SIGMA.
132Chakravarti DN et al, Biol. Abstracts, 1981, vol. 72, abstract No. 012592.
133Chan and Wasserman, Cereal Chem. vol. 70(1), p. 22-26.
134Chemistry Comes Alive-2001.
135Cheng Cheng et al., "Transformation of Trichoderma viride using the Neurospora crassa pyr4 gene and its use in the expression of a Taka-amylase a gene from Aspergillus oryzae", Curr. Genet., 18: 453-456, 1990.
136Chica et al., Current Opinion in Biotechnology, vol. 16, p. 378-384 (2005).
137Christensen et al, "A new and simple method to immobilise lipases by means of granulation", 1998 Nachwachsende Rohstoff 10, 98-105.
138Christiansen, 1993, "Application of Oxidoreductases for Food Preservation" in Progress Report of R&D Projects and Concerted Actions published by the European Communities, Luxembourg, 1993, p. 32-36.
139Christie, William et al., "New Procedures for Rapid Screening of Leaf Lipid Components from Arabidopsis", Phytochemical Analysis, vol. 9, pp. 53-57, 1998.
140Christophersen, Claus, et al., "Enzymatic Characterisation of Novamyl a Thermostable α-Amylase", Starch/Sturke, vol. 50, 1998.
141Chung O K et al, "Defatted and Reconstituted wheat flours. VI. Response to shortening addition and Lipid Removal in Flours that vary in Bread-making Quality" Cereal Chemistry (1980), vol. 57(2), p. 111-117.
142Chung OK et al, "Recent Research on Wheat Lipids" Bakers Digest Oct. 1981.
143Ciuffreda, Pierangela, et al., "Spectrophotometric Assay of Lipase Activity: A New 40nitrophenyl Ester of a Dialkylglycerol Suitable as a Chromogenic Substrate of Pseudomonas cepacia Lipase", Biocatalysis and Biotransformation, vol. 21, No. 3, pp. 123-127, 2003.
144Claesson et al., "Techniques for measuring surface forces", Advances in Colloid and Interface Science, vol. 67, 1996, pp. 119-183.
145Clare et al., 1991, Bio/Technology 9:455-460 [3].
146Clausen, Kim, "Enzymatic oil-degumming by a novel microbial phospholipase", European Journal of Lipid Science And Technology, vol. 103, 2001, pp. 333-340.
147Clausen, Kim, "New enzyme for degumming", Oils and Fats International, vol. 17, No. 4, Jun. 2001, pp. 24-25.
148Cloning of rad51 and rad52 homologues from Aspergillus oryzae and the effect of their overexpression on homologous recombination, no date.
149Collar C, et al, "Lipid binding fresh and stored formulated wheat breads. Relationships with dough and bread technological performance", Lab de Cereales Inst de Agroquimica y Tec de Alimentos, CSIC, Food Science and Technology International 2001, vol. 7(6), p. 501-510.
150Colombo, Diego, et al., "Optically Pure 1-0- and 3-0-β-D-Glucosylk—and Galactosyl-sn-glycerols through Lipase-catalyzed Transformations", Tetrahedron Letters, vol. 36, No. 27, pp. 2865-4868, 1995.
151Conference May 6-8, 1999 in Santorini, Greece, "Lipases of Lipids Structure, Function and Biotechnological Applications," Slides presented by Charlotte Poulsen.
152Conference May 6-8, 1999 in Santorini, Greece—Lipases & Lipids Structure, Function and Biotechnological Applications—Slides presented by Charlotte Poulsen.
153Cordle et al, "The hydrophobic surface of colipase influences lipase activity at an oil-water interface", Journal of Lipid Research, vol. 39 (1998), 1759-1767.
154Cordle, R.A., "The Hydrophobic Surface of Colipase Influences Lipase Activity at an Oil-Water Interface", Journal of Lipid Research, 39: 1759-1767 (1998).
155Coteron, A., et al., "Reactions of Olive Oil and Glycerol over Immobilized Lipases", JAOCS, vol. 75, No. 5, 1998.
156Council Directive of Dec. 21, 1988 (89/107/EEC).
157Council Regulation (EC) No. 2991/94 May 12, 1994 Official Journal of the European Communities, Sep. 12, 1994, No. L316/2-7.
158Courtin, Christophe M., et al., "Recent Advances in Enzymes in Grain Processing".
159Cregg et al., 1987, In: Biological Research on Industrial Yeast, vol. II, Stewart, G.G. et al. (Eds.), pp. 1-18 [4].
160Creveld, Lucia D, et al., "Identification of Functional and Unfolding Motions of Cutinase as Obtained from Molecular Dynamics Computer Simulations", Proteins: Structure, Function, and Genetics, 33:253-264, 1998.
161Cromie, Susan. Psychrotrophs and their Enzyme residues in cheese milk, The Australian Journal of Dairy Technology, vol. 47, Nov. 1992.
162Cui et al., "Purification and characterization of an intracellular carboxylesterase from Arthrobacter viscosus NRRL B-1973", Enzyme and Microbial Technology, vol. 24, pp. 200-208, 1999, NZAS-0683611.
163Curriculum vitae—Jorn Born Soe.
164D. Marion, et al., "Wheat Lipids and Lipid-Binding Proteins: Structure and Function," Wheat Structure Biochemistry and Functionality, ed. Scholfield JP), pp. 245-260 (1995).
165Daboussi et al, Heterologous expression of the Aspergillus nidulans regulatory gene nirA in Fusarium oxysporum, (1991) Gene 109(1), 155-60.
166Daboussi et al., "Transformation of seven species of filamentous fungi using the nitrate reductase gene of Aspergillus nidulans", Curr. Genet., 15:453-456, 1989.
167Daftary, R.D., et al., "Functional Bread-Making Properties of Wheat Flour Lipids", Food Technology, vol. 22, No. 237, Mar. 1968-1979, NZAS - 0230175.
168Dahlquist, Anders, et al., "Phospholipid: diacylglycerol acyltransferase: An enzyme that catalyzes the acyl-CoA-independent formation of triacylglycerol in yeast and plants", PNAS, vol. 97, No. 12, pp, 6487-6492, 2000.
169Dalrymple, Brian D., et al., "Three Neocallimastic patriciarum esterases associated with the degradation of complex polysaccharides are members of a new family of hydrolases", Microbiology, vol. 142, pp. 2605-2614, 1997.
170Danisco Experimentation "Effect of Alpha-Glucosidase on Gluconic Acid and Maltose".
171Danisco further experimentation "Effect on dough of HOX and GOX on maltose in dough".
172Danisco further experimentation "Experiments on glucoligosaccharide oxidase (GO) from Acremonium".
173Danisco further experimentation with color photographs.
174Danisco, "Unique Chance for Better Bread" Direct, A Newsletter from Danisco Ingredients (1996).
175Danisco, Hexose oxidase—nyt enzym med mange mulingheder (advert).
176Darnell et al., Eds., "Synthetic Peptide and Nucleotide Sequences: Their Use in Isolating and Identifying Genes", in Molecular Cell Biology, Chapter 6, Manipulating Macromolecules, 1990, Scientific American Books, Baltimore.
177Database accession No. P10480 -& Database UniProt 'Online!, Jul. 1, 1989.
178Database accession No. Q44268 -& Database UniProt 'Online! Nov. 1, 1996.
179Database accession No. Q9F7Y6 Database UniProt 'Online!, Mar. 1, 2001.
180Database FSTA International Food Information Service (IFIS), Frankfurt/Main, De Mine Y:"Application of the enzymatic methods to the determination of contaminated yolk in egg white." XP002077295 see abstract & Food Research International, vol. 29, No. 1, pp. 81-84, 1976.
181Database FSTA International Food Information Service (IFIS), Frankfurt/Main, De Nicolas J:"Action of oxidoreductases in breadmaking. Maturation of soft wheat flours and kneading of doughs." XP002077286 see abstract & Annales De Technologie Agricole, vol. 28, No. 4, 1979, pp. 445-468.
182Database FSTA International Food Information Service (IFIS), Frankfurt/Main, De Qi Si J: "New enzymes for the baking industry" XP002077284 see abstract & Food Tech Europe vol. 3, No. 1, 1996, pp. 60-64, Novo Nordisk Ferment Ltd.
183Database FSTA International Food Information Service (IFIS), Frankfurt/Main, De Weipert D:"Rheologie von Roggenteigen. II. Der einfluss der enzyme unterschiedlicher spezifitat auf das rheologische verhalten des teiges." XP002077285 see abstract & Getreide, Mehl Und Brot, vol. 26, No. 10, 1972, pp. 275-280.
184Database UNIPROTKH Jun. 1, 2003, S. Omura et al: "putative secreted hydrolase from streptomyces avermitilis" XP002376340 retrieved from EBI, Hinxton, UK Database accession No. Q828T4 abstract.
185Database UNIPROTKH May 1, 2000, S.D. Bentley et al: "Putative Secreted Hydrolase from Streptomyces coelicolor" XP002376339 retrieved from EBI, Hinxton, UK Database accession No. Q9S2A5 abstract.
186Davies, Progress in Industrial Microbiology, Martinelli and Kinghorn (eds.), Elsevier, Amsterdam 1994, 29:525-560.
187De Haas GH et al, "Purification and Properties of Phospholipase A from Porcine Pancreas" Biochim. Biophys. ACTA, 1968, vol. 139, pp. 103-117.
188Declaration by Clive Graham Phipps Walter (Dec C).
189Declaration by Dr Jorn Borch Soe (Dec F).
190Declaration by Dr M Turner.
191Declaration by Dr Mark Turner (Dec G).
192Declaration by Henrik Pedersen (Dec A).
193Declaration by Henrik Pedersen, Masoud Rajabi Zargahi and Clive Graham Phipps Walter, (Dec 2).
194Declaration by Janne Brunstedt (Dec D).
195Declaration by Kazuko Kato, Henrik Pedersen, Masoud Rajabi Zaghari, Clive Phipps Walter, and Janne Brunstedt (Dec I).
196Declaration by Kim Borch.
197Declaration by Luise Erlandsen.
198Declaration by Masoud Rajabi Zargahi (Dec B).
199Declaration by Masoud Rajabi Zargahi (Dec E).
200Declaration by Tina Spendler.
201Definition of "hexose", Webster Dictionary, p. 1065., no date.
202Definition of "hexose", Webster Dictionary, p. 1065.
203Delcros, Jean-Francois, et al., "Effect of mixing conditions on the behavior of lipoxygenase, peroxidase, and catalase in wheat flour doughs", Cereal Chemistry, 1998, vol. 75, No. 1, pp. 85-93.
204Dellaporta, et al.; "A Plant DNA Minipreparation Version II"; Plant Molecular Biology Reporter(1983); vol. 1(4); pp. 19-21.
205Derewenda et al, "The crystal and molecular structure of the Rhizomuxor miehei Triacylglyceride Lipase at 1.9 ÅResolution", J. Mol. Biol. 1992, 227:818-839.
206Derewenda, Urszula, et al., "Catalysis at the Interface: The Anatomy of a Conformational Change in a Triglyceride Lipase", Biochemistry, vol. 31, pp. 1532-1541, 1992.
207Derwent Publications Ltd., London, GB; AN 73-30288u XP002012361 & JP, A48016612 (EISAI Co. Ltd).
208Dictionary of Biochemistry and Molecular Biology, Second Edition, p. 16.
209Dinkci. N, Mucor miehei den elde edilen lipaz.
210Direct, A Newsletter from Danisco Ingredients, Sep. 1996.
211Directive 2000/36/EC. Http://europa.eu.int/scadplus/leg/en/lvb/121122b.htm. Dato: Jun. 16, 2004.
212Dowling et al., "Hexose Oxidation by an enzyme system of Malleomyces Pseudomallei", Journal of Bacteriology (1956) 72:555-560.
213Drost-Lustenberger, C and Spendler T Lipopan F BG—Application and Mechanism of a new lipase for baking, Novozymes.
214Drost-Lustenberger, Cornelia, et al., "Lipopan F BG—application and mechanism of a new lipase for bread baking", Cereal Food, 2003.
215Drost-Lustenberger, Cornelia, et al., "Lipopan F BG—unlocking the natural strengthening potential in dough", Cereal Food, 2004.
216Duan, Rui Dong, Fat Digestion and Absorption (2000), p. 25-46, publisher AOCS Press, Champaign III CODEN 69ACBA Conference; general review written in English.
217Dubreil, Laurence, et al., "Localization of Puroinoline-a and Lipids in Bread Dough Using Confocal Scanning Laser Microscopy", J. Agric. Food Chem., 2002, vol. 50, pp. 6078-6085, NZAS-0204271.
218Ducancel, Frederic, et al., "Complete amino acid sequence of a PLA2 from the tiger snake Notechis sculatus scutatus as deduced from a complementary DNA", Nucleic Acids Research, vol. 16, No. 18, 1988, NZAS-0418884.
219Dugi KA et al, "Human hepatic and lipoprotein lipase: the loop covering the catalytic site mediates lipase substrate specificity", Journal of Biological Chemistry (1995), vol. 270, pp. 25, 396-pp. 25, 401.
220Dugruix (Edited by) Crystallization of Nucleic Acids and Proteins A Practical Approach.
221Dutilh & Groger, "Improvement of Product Attributes of Mayonnaise by Enzymic Hydrolysis of Egg Yolk with Phospholipase A2", 1981 J. Sci. Food Agric. 32, 451-458.
222Dybdal, L., et al., "Enzymes in Cereals Processing", NZAS - 0254380.
223Eddine et al, "Cloning and expression analysis of NhL1, a gene encoding an extracellular lipase from the fungal pea pathogen Nextria haematococca MP VI (Fusarium solani f. sp. pisi) that is expressed in planta", Mol. Genet. Genomics (2001) 265: 215-224.
224EFEMA Index of Food Emulsifiers Jan. 2004, 4th Edition.
225Effect of different Hexose Oxidase.
226Efthymiou CC et al. Development of domestic feta cheese.
227Eliasson et al., "Cereals in Breadmaking—A molecular colloidal approach", NZNA- 0006056.
228Ellaiah et al., "Production of lipase by immobilized cells of Aspergillus niger", Process Biochemistry, vol. 39, 2004, pp. 525-528.
229Ellman, George L.: "A Colorimetric Method for Determining Low Concentrations of Mercaptans", Archives of Biochemistry and Biophysics (1958) 74: 443-450.
230Elyk, Alexander, et al., "Lipase-Catalyzed----------------", JAOCS, vol. 08, No. 5, May 1991, pp. 320-323 NZAS-0033036.
231Engelhorn and Raab, "Rapid Electroblotting of Small DNA Fragments from Polyacrylamide Gels", Biotechniques (1991) 11(5):594-6.
232Engelhorn et al., "Rapid Electroblotting of Small DNA Fragments from Polyacrylamide Gels"; Biotechniques(1991); vol. 11(5); pp. 594-596, NZAS-0665838.
233English translation of EIDA, JP 73 01661 (Japanese Unexamined Patent Publication No. 48-16612.
234Enzyme Schemes.
235Enzymes in food processing (3rd Ed.), Academic press 1993.
236EPO, Mobay Chemical Corporation—Decision of the Technical Board of Appeal 3.3.1 dated Jul. 1, 1982, Official Journal EPO, Oct. 1982, pp. 394-402.
237Ettinger, William F. et al., "Structure of Cutinase Gene, cDNA, and the Derived Amino Acid Sequence from Phytopathogenic Fungi", Biochemistry, vol. 26, pp. 7883-7892, 1987.
238Euromonitor International, "The World Market for Dairy Products—Introduction, Executive Summary, Operating Environment, World Market Overview, Key Trends and Developments" in Euromonitor, Strategy 2000, Feb. 2001.
239European Parliament and Council Directive No. 95/2/EC of Feb. 20, 1995 on food additives other than colours and sweeteners.
240European Parliament and Council Directive No. 98/72/EC of Oct. 15, 1998 amending Directive 95/2/EC on food additives other than colours and sweeteners.
241Eurpean Journal of Biochemistry, vol. 166, 1987, Published by Springer International on behalf of the Federation of European Biochemical Societies.
242Examiner's Report on Application for Patent of Invention (CL 939-96) and English translation.
243Exhibit a filed by DSM.
244Exhibit B filed by DSM.
245Experiments by Danisco "A method of improving the properties of flour dough, a flour dough improving composition and improved food products" (Declaration of Jorn Born Soe).
246Experiments by Danisco "Evaluation of enzyme of present invention in the production of German mischbrot" (Declaration of Jorn Born Soe).
247Experiments by Novo in Appeal Statement.
248Extract from Industrial Enzymology.
249Ezra, David, et al., "Coronamycins, peptide antibiotics produced by a verticillate Streptomyces sp. (MSU-2110) endophytic on Monstera sp.", Microbiology, 2004, vol. 150, p. 785-793, NZAS-0216252.
250Fauvel, et al.; "Purification of Two Lipases With High Phospholipase A, Activity from Guinea-Pig Pancreas"; Biochimica et Biophysica Acta(1981); vol. 663; pp. 446-456.
251Fennema, Owen F., "Food Chemistry Second Edition, Revised and Expanded", NZAS-0213736.
252Fernandez et al., 1992, Analytical Biochemistry, 201:255-264 [5].
253Fernandez, J. et al., 1994, An Improved Procedure for Enzymatic Digestion of Polyvinylidene Difluoride-Bound Proteins for Internal Sequence Analysis, Analytical Biochemistry, 218:112-117.
254Fernandez-Garcia et al., "The use of lipolytic and proteolytic enzymees in the manufacture of manchego type cheese from ovine and bovine milk", 1994 J. Dairy Sci. 77: 2139-2149.
255Fernandez-Lafuente, Roberto, et al., The coimmobilization of D-amino acid oxidase and catalase enables the quantitative transformation of D-amino acids (D-phenylalanine) into α-keto acids (phenylpyruvic acid), Enzyme and Microbial Technology, vol. 23, pp. 28-33, 1998, NZAS-0442385.
256Ferrer et al, 2000, J. Chem. Technol. Biotechnol. 75, 569-576.
257Finizym Technical Information, Novo Enzymes, 1981.
258Fødevarenubusteriet (2003). Bekendtgørelse om indhold Of transfedtsyrer I olier og fedtstoffer. Bekendtgørelse nr. 160 af Nov. 3, 2003.
259Food Enzymes: Stalingase L, Gist-brocades Food Ingredients.
260Food R&D. Dairy fields ingredient technology section.
261Forman, Todd, "Enzymes Used in Bread Baking: An Application Update", Technical Bulletin, vol. XXVI, Issue 10, Oct. 2004.
262Fox, et al.; "Isolation and some Properties of Extracellular Heat-Stable Lipases: from Pseudomonas Fluorescens Strain AFT 36"; Journal of Dairy Research (1988); vol. 50; pp. 77-89.
263Frenken N. et al (1992) Appl. Envir. Microbiol. 58 3787-3791.
264Freshzyme, Product Sheet, NZAS-0265916.
265Frohman, et al.;"Rapid Production of Full-Length cDNAs from Rare transcripts: Amplification using a single gene-specific oligonucleotide primer"; Proc. Natl. Acad. Sci. USA (1988); vol. 85; pp. 8998-9002.
266Frost & Sullivan, U.S. Market for Enzymes for food Applications, NZAS-0413133.
267Fugman, Douglas A et al Biochemica et Biophysica acia 795 (1984) 191-195.
268Functional Bread-Making Properties of Lipids, NZAS-0487568.
269Galliard T and Dennis S (1974) Phytochemistry vol. 13, pp. 1731-1735.
270Galliard, "The Enzymic Breakdown of Lipids in Potato Tuber by Phospholipid—And Galactolipid-Acyl Hydrolase Activities and by Lipoxygenase", Phytochemistry, 1970, vol. 9, pp. 1725-1734, NZAS-0487554.
271Gan, Z. et al., "Rapid Communication—Antisera agains: Wheat Diacylgalactosylglycerol (MGDG) and Diacyldigalactosylglycerol (DGDG)", Journal of Cereal Science, vol. 18, pp. 207-210, 1993.
272Ganghro AB & Dahot MU, Sci Int. (Lahore), 1992, vol. 4, pp. 169-172.
273Garcia et al. Journal of Agricultural and Food Chemistry (2004), vol. 52, p. 3946-3953.
274Garcia et al., Methods Enzymol., vol. 71, p. 782-772 (1981).
275Garzillo et al., Biotechnol Appl Biochem (1995) vol. 22, p. 169-178 and Proof of date of publication of Garzillo et al.
276Gemel, Joanna et al., "Comparison of galactolipase activity and free fatty acid levels in chloroplasts of chill-sensitive and chill resistant plants", European Journal of Biochemistry, vol. 166, p. 229-33, 1987.
277Geus et al (1987) Nucleic Acids Research 15(9) p. 3743-3759.
278Giffhorn, Appl. Microbiol. Biotechnol., vol, 54, p. 727-740 (2000).
279Giffhorn, F., "Fungal Pyranose Oxidases: Occurrence, Properties and Biotechnical Applications in Carbohydrate Chemistry", Appl. Microbiol. Biotechnol., 54:727-740 (2000).
280Gilbert, E. Jane, et al., "Purification and properties of extracellular lipase from Pseudomonal aeruginosa EF2", Journal of General Microbiology, 1991, vol. 137, pp. 2223-2229, NZAS-0225579.
281Gillian, B., Turgeon et al., "Cochliobolus heterostrophus using the Aspergillus nidulans amdS gene", Mol Gen Genet, 201: 450-453, 1985.
282Gist-brocades, Amylase P Information Sheets.
283Glucose Oxidase (Gox) E.C. 1.1.3.4.
284Glucose Oxidase, An Extract from the Enzyme Handbook, p. 1-7 (1995).
285Glucose Oxidase: A much used and much loved enzyme in biosensors.
286Godfrey, Tony, et al., "Industrial Enzymology Second Edition".
287Goodey et al, Yeast Biotechnology, Berry et al (eds.), Allen and Unwin, London 1987, pp. 401-429.
288Graille J, Lipid Technology, vol. 5, No. 1, 1993, pp. 11-16.
289GRAS Notification dated Apr. 11, 2001 by Novozymes for LecitaseR and Lipopan™ F.
290Greenough et al (1996) Food Chem Toxicology 34:161-166 and PubMed abstract in respect thereof.
291Greenough R J et al, Food and Chemical Toxicology, vol. 34(2), 1996, pp. 161-166.
292Grindsted Products, Grindsted Bakery News.
293Grindsted, "Emulsifiers for the baking industry".
294Grindsted, "Grindamyl Fungal Alpha-Amylase".
295Groen, B. W., s De Vries, J. A. Duine (1997), Eu. J. Biochem-, vol. 244, pp. 858-861, "Characterization of hexose from the red seaweed Chondrus crispus".
296Groen, B. W., s De Vries, J. A. Duine (1997), Eu. J. Biochem., vol. 244, pp. 858-861, "Characterization of hexose from the red seaweed Chondrus crispus".
297Groppe, J.C. and Morse, D.E., 1993, Isolation of full-length RNA templates for reverse transcription from tissues rich in RNase and proteoglycans, Anal. Biochem., 210:337-343.
298Haarasilta, S., T. Pullinen (1993) in Baking Industry Europe (Alan Gordon, editor), pp. 49-52.
299Haas and Berka, 1991, Gene, 109:107-113.
300Haas, et al., "Enzymatic Phosphatidylcholine Hydrolysis in Organic Solvents: An Examination of Selected Commercially Available Lipases", JAOCS, vol. 71, No. 5, May 1994, pp. 483-490.
301Haas, et al.; "Lipases of the Genera Rhizopus and Rhizomucor: Versatile Catalysts in Nature and the Laboratory"; Food Biotechnology Micro-organisims (1995); pp. 549-588.
302Haggag H F et al. Egypt J Food Sci vol. 22, No. 1 pp. 99-107 (1994).
303Hamer, Rob J., et al., "Interaction: The Keys to Cereal Quality", American Association of Cereal.
304Hanlin, Richard T., "Illustrated Genera of Ascomycetes"; The American Phytopathological Society.
305Hansen, Chr., Danisco and Novozymes, Apr. 3, 2002, Food Ingredients day, R&D—the main ingredients for growth.
306Hara, et al.; "Comparative Study of Comercially Available Lipases in Hydrolysis Reaction of Phosphatidylcholine"; JAOCS (1997); vol. 74; No. 9, pp. 1129-1132.
307Hawker, Kim L., et al., "Heterologous expression and regulation of the Neurospora crassa nit-4 pathway-specific regulartory gene for nitrate assimilation in Aspergillus nidulans", Gene., vol. 100, pp. 237-240, 1991.
308Hedin, Eva M.K., et al., "Selective reduction and chemical modification of oxidized lipase cysteine mutants".
309Helmsing, "Purification and Properties of Galactolipase", Biochim., Biophys., Acta, vol. 178, pp. 519-533, 1969.
310Henderson, H.E., et al., "Structure-function relationships of lipoprotein lipase: mutation analysis and mutagenesis of the loop region", Journal of Lipid Research, vol. 34, 1993, pp. 1593-1602.
311Henke, Erik, et al., "Activity of Lipases and Esterases towards Tertiary Alcohols: Insights into Structure-Function Relationships", Angew. Chem. Int. Ed., 2002, vol. 41, No. 17.
312Hernquist L & Anjou K (1993) Diglycerides as a stabilizer of the β′-crystal form in margarines and fats, in Fette Seifen Anstrichmittel 2:64-66.
313Hernquist L. Herslof B. Larsson K & Podlaha O. (1981) Polymorphism of rapeseed oil with low content of erucic acid and possibilities to stabilize the β′-crystal form in fats, in Journal of Science and Food Agriculture 32:1197-1202.
314Hilton S et al, Biochemistry vol. 29, No. 38, 1990, pp. 9072-9078.
315Hilton S, Buckley JT, J Biol Chem. Jan. 15, 1991; 266(2): 997-1000.
316Hirayama O et al, Biochim Biophys Acta. 1975, vol. 384(1), p. 127-37.
317Hirose, Yoshihiko et al., "Characteristics of Immobilized Lipase PS on Chemically Modified Ceramics", Amano Pharmaceutical, NZAS-0239105.
318Hjorth, Annegrethe, et al., "A Structural Domain (the lid) Found in Pancreatic Lipases is Absent in the Guinea Pic (Phospho) lipase", Biochemistry, vol. 32, pp. 4702-4704, 1993.
319Höfelmann et al, J. Food Sci., 1985, 50:1721-1731.
320Holmquist et al., "Lipases from Rhizomucor miehei and Humicola lanuginosa: Modification of the Lid covering the active site alters enantioselectivity", Journal of Protein Chemistry, vol. 12, No. 6, 1993, NZAS-0668771.
321Holmquist et al., "Probing a Functional Role of Glu87 and Trp89 in the Lid of Humicola lanuginosa Lipase through Transesterification Reactions in Organic Solvent", Journal of Protein Chemistry, 1995, vol. 14, No. 4, pp. 217-224, NZAS-0668747.
322Holmquist et al., "Trp89 in the Lid of Humicola lanuginosa Lipase is Important for Efficient Hydrolysis of Tributyrin", Lipids, vol. 29, No. 9, 1994, NZAS-0668751.
323Horn T et al, (1980) Nuc Acids Res Symp Ser 225-232.
324Hoshino, et al.; "Calcium Ion Regulates the Release of Lipase of Fusarium oxysporum"; J. Biochem (1991); vol. 110; pp. 457-461.
325Hoshino, et al.; "Purification and Some Characteristics of Extracellular Lipase from Fusarium oxysporum f sp. lini"; Biosci. Biotech. Biochem (1992); pp. 660-664.
326Hoshino, Tamotsu, et al., "Purfication and Some Characteristics of Extracellular Lipase from Fusarium oxysporum", Biosci. Biotech. Biochem., vol. 56, No. 4, pp. 660-664, 1992.
327Hossen, Monjur and Hernandez, Ernesto, Lipids, vol. 39, Aug. 2004, pp. 777-782.
328Hossen, Monjur, "Enzyme catalyzed synthesis of structured phospholipids with conjugated linoleic acid and plant sterols," A Dissertation by MD Monjur Hossen, May 2005.
329Hou Ching T, Journal of Industrial Microbiology, vol. 13, No. 4, 1994, pp. 242-248.
330Hou, C.T., "pH Dependence and Thermostability of Lipases from Cultures from the ARS Culture Collection", Journal of Industrial Microbiology, 13:242-248 (1994).
331Hübner et al., "Interactions at the lipid-water interface", Chemistry and physics of Lipids, vol. 96, 1998, pp. 99-123, NZAS-0684011.
332Hugh-Jensen, Birgitte, et al., "Rhizomucor miehei Triglyceride Lipase is Processed and Secreted from Transformed Aspergillus oryzae", Lipids, vol. 24, No. 9, pp. 1989, NZAS-0223016.
333Humum et al., "Enzyme Catalysed Synthesis in Ambient Temperature Ionic Liquids", Biocatalysis and Biotransformation, vol. 19, pp. 331-338, NZAS-0215170.
334Icard-Verniere, Christele, et al., "Effects of mixing conditions on pasta dough development on biochemical changes", Cereal Chemistry, 1999, vol. 76, No. 4, pp. 558-565.
335Identification of alpha-glucosidase (otherwise known as transglucosidase) activity in Bakezyme GO 10000 from DSM.
336Igrejas, Gilberto, et al., "Genetic and Environmental Effects on Puroindoline-a and Puroindoline -b Content and their Relationship to Technological Properties in French Bread Wheats", Journal of Cereal Science, vol. 34, 2001, pp. 37-47.
337Ikawa, 1982, Methods Enzymol., 89: 145-149.
338Ikeda H et al, Nature Biotech, vol. 21, 2003, p. 526-531.
339Industrial enzymology (2nd Ed.), The Macmillan press 1996.
340International Dairy Federation Bulletin Document, Document 116, p. 5. (1979).
341International Search Report for the International Searching Authority in PCT/DK96/00239 issued Sep. 11, 1996.
342Investigation of Glucose Oxidase from Cladosporium Oxysporum.
343Ishihara et al Biochimica et Biophysica Acta 388 (1975) 413-422.
344Isobe and Nokihara, Febs. Lett., 1993, 320:101-106.
345Isobe K et al, Journal of Molecular Catalysis B: Enzymatic 1 (1995), pp. 37-43.
346Iwai and Tsujisaka (in Lipases, Borgström and Brockman (eds.), Elsevier, Amsterdam, 1984, pp. 443-468.
347Iwai, Mieko, et al., "Hydrolytic and Esterifying Actions of Crystalline Lipase of Aspergillus Niger", Osaka Municipal Technical Research Institute, Osaka, Japan.
348Izco et al. Adv Food Sci vol. 21 N 3/4 (10-116) 1999.
349J. Chromatog., Knoll et al., 55 (1971), 425-428.
350J. Chromotog., Knoll et al., 55 (1971), 425-428.
351Jacob, Jules S., et al., "The Effects of Galactolipid Depletion on the Structure of a Photosynthetic Membrane", The Journal of Cell Biology, vol. 103, Oct. 1986, pp. 1337-1347.
352Jacobsberg B. & Oh C.H. (1976) Studies in Palm Oil Crystallisation, in Journal of the American Oil Chemist Society 53:609-616.
353Jan-Willem F. A. Simons et al., "Cloning, purification and characterisation of the lipase from Staphylococcus epidermidis", Eur. J. Biochem., vol. 253, pp. 675-683, 1998, NZAS-0216239.
354Jeng-yen Lin, Matthew, "Wheat Polar Lipids—A Theseis Submitted to the Graduate Faculty of the North Dakota State University of Agriculture and Applied Science", May 1972.
355Jensen B et al "Effect and Activity of Lipases in Dough and Bread" Translation.
356Jensen, B., et al., "Effekt and Wirksamkeit von Lipasen in Teig and Brot".
357JJ Owens. Lecithinase Positive Bacteria in milk.
358Joerger et al., "Alteration of Chain Length Selectivity of a Rhizopus delemar Lipase through Site-Directed Mutagenesis", Lipids, vol. 29, No. 6, 1994, pp. 377-384, NZAS-0668759.
359Jong et al.; "American Type Culture Collection Catalogue of Filamentous Fungi"; Eighteenth edition (1991); NZAS-0025887.
360Joshi, et al.; "Specificity of Fungal Lipase in Hydrolytic Cleavage of Oil"; Acta Microbiologica Hungarica (1987); vol. 34(2); pp. 111-114.
361Joshi, Sunita, et al., "Specificity of Lipase isolated from Fusarium oxysporum", Department of Chemistry, Indian Institute of Technology, vol. 25, No. 1 & 2, pp. 76-78.
362Juffer, A.H., et al., "Adsorption of Proteins onto Charged Surfaces: A Monte Carlo Approach with Explicit Ions", Journal of Computational Chemistry, vol. 17, No. 16, pp. 1783-1803, 1996.
363Jurgens, Catharina, et al., "Directed evolution of a (βα)8-barrel enzyme to catalyze related reactions in two different metabolic pathways", PNAS, Aug. 29, 2000, vol. 97, No. 18, pp. 9925-9930, NZAS-0458259.
364Kaniuga Z, Acta Biochim Pol. (1997), vol. 44(1), p. 21-35.
365Kaplan, Methods in Enzymology, vol. 3, p. 107-110 (1957).
366Kapur J & Sood ML, J. Parasit., 1986, vol. 72, pp. 346-347.
367Kasai, Naoya, et al., "Chiral C3 epoxides and halophydrins: Their preparation and synthetic application", Journal of Molecular Catalysis B: Enzymatic, vol. 4, 1998, pp. 237-252, NZAS-0562172.
368Kasai, Naoya, et al., "Optically Active Chlorohydrins as Chiral C3 and C4 Building Units: Microbial Resolution and Synthetic Applications", Chirality, vol. 10, pp. 682-692, NZAS-0562161.
369Kawamura and Doi, J. of Bacteriology Oct. 1984, p. 442-444.
370Keilin et al., Biochem J (1948) vol. 24, p. 206-207.
371Keller, R.C.A., et al., "Competitive Adsorption Behaviour of Wheat Flour Components and Emulsifiers at an Air-Water Interface", Journal of Cereal Science, vol. 25, 1997, pp. 175-183.
372Kelley and Reddy, J. Bacteriology, vol. 166, p. 269-274 (1986).
373Kelly and Reddy, Methods in Enzymology, vol. 161, p. 307-316 (1988).
374Kerschensteiner, D. D. Diss. Abstr. Int. B 1978, 39(7), 3299, CAN 90: 117113 AN 1978:117113 Caplus, "The mechanism of action and the state of copper in hexose oxidase".
375Kerschensteiner, D. D. Diss. Abstr. Int. B 1978, 39(7), 3299, Can 90:117113 An 1978:117113 Caplus, "The mechanism of action and the state of cooper in hexose oxidase ".
376Kerschensteiner, D. D. Diss. Abstr. Int. B 1978, 39(7), 3299, Can 90:117113 An 1978:117113 Caplus, "The mechanism of action and the state of cooper in hexose oxidase".
377Kerschensteiner, D.A. and Klippenstein, D.A., 1978, Purification Mechanism and State of Copper in Hexose Oxidase, Federation Proceedings 37:1816 abstract.
378Kerschensteiner, The Mechanism of Action and the State of Copper in Hexose oxidase, Thesis, 1978, p. iii-xiii.
379Keum J S et al. Korean J Dairy Sci 15 (2): 103-117 1993.
380Kim, Hyung Kwoun, et al., Expression and characterization of Ca2+-independent lipase from Bacillus pumilus B26, Biochimica et Biophysica Acta, vol. 1583, 2002, pp. 205-212, NZAS-0204610.
381Kim, Myo-Jeong, et al., "Thermal Inactivation Kinetics and Application of Phospho and Galactolipid-Degrading Enzymes for Evaluation of Quality Changes in Frozen Vegetables", J. Agric. Food Chem., 2001, vol. 49, pp. 2241-2248.
382Kimura, Yoshiharu, et al., "Application of Immobilized Lipase to Hydrolysis of Triacylglyceride", Eur J. Appl Microbiol Biotechnol, 1983, vol. 17, pp. 107-112 NZAS-0033058.
383Kindstedt et al, Rapid Quantative test for free oil (Oiling off) in melted Mozzarella cheese.
384King et al, Molecular and Cell Biology of Yeasts, Walton and Yarronton (eds.), Blackie, Glasgow, 1989, pp. 107-133.
385Kirk, Ole, et al., "Fatty Acid Specificity in Lipase-Catalyzed Synthesis of Glucoside Esters" Biocatalysis, 1992, vol. 6, pp. 127-134, NZAS-0214549.
386Klein, Robert R., et al., "Additive Effects of Acyl-Binding Site Mutations on the Fatty Acid Selectivity of Rhizopus delemar Lipase", JAOCS, vol. 74, No. 11, 1997.
387Klein, Robert R., et al., "Altered Acyl Chain Length Specificity of Rhizopus delemar Lipase Through Mutagenesis and Molecular Modeling", Lipids, 1997, vol. 32, No. 2, pp. 123-130.
388Kocak et al, Effect of lipase enzyme (palatase A 750 L) on the ripening of tulum cheese.
389Kocak et al, Milchwissenschaft 51(1), 1996.
390Kochubei et al Role of lipids in the organization of the closest surroundings of the reaction centers(1976) Institute of Plant Physiology.
391Kochubei S M et al, Biophysics (1981), vol. 26(2), p. 299-304.
392Kochubei S M et al, Mol Biol (Mosk) (1975), vol. 9(2), (p. 190-3) p. 150-153.
393Kochubei SM et al, Mol Biol (Mosk) (1978), vol. 1, p. 47-54, p. 32-37.
394Kolkovski et al (1991) Fish Nutrition in Practice, Biarritz (France), Jun. 24-27.
395Kostal, Jan, et al., "Enhanced Arsenic Accumulation in Engineered Bacterial Cells Expressing ArsR", Applied and Environmental Microbiology, Aug. 2004, pp. 4582-4587, NZAS-0572557.
396Kouker, et al.; "Specific and Sensitive Plate Assay for Bacterial Lipases"; Applied and Environmental Microbiology (1987); vol. 53(1); pp. 211-213.
397Krishna, Sajja Hari, et al., "Enantioselective transesterification of a tertiary alcohol by lipase A from Candida antarctica", Tetrahedron: Asymmetry, vol. 13, 2002, pp. 2693-2696, NZAS-0457490.
398Kristensen A.C.J. (2004) Preparation of margarine and spreads by enzyme-generated emulsifiers. Master thesis, The Royal Veterinary and Agricultural University, Frederiksberg, Copenhagen.
399Krog, Cereal Foods World, The American Association of Cereal Chemists, p. 10, Jan. 1979, vol. 24, No. 1, pp. 10-11.
400Krog, N.J., "Dynamic and Unique Monoglycerides", Cereal Foods World, 24(1): 10-11 (1979).
401Krupa, Zbigniew et al., "Requirement of Galactolipids for Photosystem J Activity In Lyophilized Spinach Chloroplasts", Biochimica et Biophysica Acta, 408, pp. 26-34, 1975.
402KSV-5000.
403Kuipers, Oscar P., et al., "Enhanced Activity and Altered Specificity of Phospholipase A2 by Deletion of a Surface Loop", Science, vol. 244, 1989, NZAS-0668767.
404Kulp K, Advances in Baking Technology, p. 152-178 (1993).
405Kunze, Hans, et al., "On the mechanism of lysophospholipase activity of secretory phospholipase A2 (EC 3.1.1.4): deacylation of monoacylphosphoglycerides by intrinsic sn-1 specificity and Ph-dependent acyl migration in combination with sn-2 specificity", Biochimica et Biophysica Acta, vol. 1346, 1997, pp. 86-92, NZAS-0399970.
406Kuwabara, et al., "Purification and Some Properties of Water-soluble Phospholipase B from Torulaspora delbrueckii", J. Biochem., vol. 104, pp. 236-241, 1988, NZAS-0418757.
407Kuwabara, et al., "Purification and Some Properties of Water-soluble Phospholipase", Agric. Biol. Chem., vol. 52, No. 10, pp. 2451-2458, 1988, NZAS-0418752.
408Kweon et al., "Phospholipid Hydolysate and Antistaling Amylase Effects on Retrogradation of Starch in Bread", Journal of Food Science, vol. 59, No. 5, 1994, NZAS-0666141.
409Laemmli, U.K., 1970, Cleavage of structural Proteins during the Assembly of the Head of Bacteriophage T4, Nature (London), 227:680-685.
410Larchenkova LP et al. Effect of starter and souring temperature on reproduction of E coli and lactobacili in milk.
411Larsen N G et al, Journal of Cereal Science (1990), vol. 12(2), p. 155-164.
412Lecointe et al Biotechnology Letters, vol. 18, No. 8 (August) pp. 869-874.
413Lee, Keun Hyeung, et al., "Identification and characterization of the antimicrobial peptide corresponding to C-terminal B-sheet domain of tenecin 1, an antibacterial protein of larvae of Tenebrio molitor", Biochem. J., 1996, vol. 334, pp. 99-105, NZAS-0574043.
414Lee, Kyung S., et al., The Saccharomyces cerevisiae PLB1 Gene Encodes a Protein Required for Lysophospholipase and Phospholipase B Activity, The Journal of Biological Chemistry, vol. 269, No. 31, Issue of Aug. 5, pp. 19725-19730, NZAS-0418807.
415Leggio, Leila Lo, et al., "The 1.62 A structure of Thermoascus aurantiacus endoglucanase: completing the structural picture of subfamilies in glycoside hydrolase family 5", FEBS Letters, vol. 523, 2002, pp. 103-108, NZAS-0457668.
416Leidich et al., "Cloning and Disruption of caPLB1, a Phospholipase B Gene Involved in the Pathogenicity of Candida albicans", The Journal of Biological Chemistry, vol. 273, No. 40, oo. 26078-26086, 1998.
417Leon et al., "A new approach to study starchy changes occurring the double-baking process and during bread storage," Z. Lebensn. Unters Forsch A, 1997, vol. 204 p. 316-320.
418Li, W., et al., "Surface properties and locations of gluten proteins and lipids revealed using confocal scanning laser microscopy in bread dough", Journal of Cereal Science, vol. 39, 2004, pp. 403-411, NZAS-0683630.
419Lih-ling Wang et al, J Agric. Food. Chem. (1993), 41, 1000-1005.
420Lima, Vera L.M., et al., "Lecithin-cholesterol acyltransferase (LCAT) as a plasma glycoprotein: an overview", Carbohydrate Polymers, vol. 55, 2004, pp. 179-191, NZAS-0683970.
421Lin M J Y et al, Cereal Chemistry (1974), vol. 51(1), p. 34-45.
422Lin S et al, Enzyme and Microbial Technology 18 (1996), pp. 383-387.
423Lin, Shuen-Fuh et al., "Purification and Characterization of a Novel Glucooligosaccharide Oxidase from Acremonium strictum T1", Biochimica et Biophysica Acta, 1118:41-47 (1991).
424Lipase A "Amano" 6 Assay Note and Product Specification from Armano Pharmaceutical Co Ltd Nagoya Japan, Aug. 27, 1985.
425Lipase A "Amano" 6 Assay Note and Product Specification from Armano Pharmaceutical Co Ltd Nagoya Japan, Dec. 16, 1985.
426Lipase A "Amano" 6 product sheet, Apr. 1, 1999.
427Lipase SP677 as a Baking Enzyme, from Novo Nordisk, Denmark, Mar. 17, 1994.
428Lipomod L338P.
429Lipopan F: Keep the quality—cut your costs 2000 Novozymes A/S. www.enzymes.novo.dk/cgl-bin/bvisapi.dll/biotimes/one—article.jsp?id=16947&lang=en&t=b1.
430Litthauer, Derek, et al., "Pseudomonas luteola lipase: A new member of the 320- residue Pseudomonas lipase family", Enzyme and Microbial Technology, vol. 30, pp. 209-215, 2002.
431Llustenberger, Cornelia, et al., "Application of Noopazyme in Asian Noodles and Non-Durum Pasta", Cereal Food, 2002-18584-01, p. 1, vol. 11, NZAS-0255265.
432Llustenberger, Cornelia, et al., "Enzymes in Frozen Dough and Parbaked Bread", Cereal Food, 2001-17056-01, p. 1, vol. 19, NZAS - 0255182.
433Lo Y-C et al. Crystal structure of Escherichia coli Thioesterase I/Proteasel/Lysophospholipase L1: Consensus sequence blocks constitute the catalytic center of SGNH-hydrolases through a conserved hydrogen bond network. Journal of Molecular Biology, London, GB, vol. 330, No. 3, 539-551.
434Longhi, Sonia, et al., "Atomic Resolution (1.0) Crystal Structure of Fusarium solani Cutinase: Stereochemical Analysis" J. Mol. Biol. vol. 268, pp. 779-799, 1997.
435Lozano et al., "Over-stabilization of Candida antarctica lipase B by ionic liquids in ester synthesis", Biotechnology Letters, vol. 23, pp. 1529-1533, 2001, NZAS-0214942.
436Lustenberger Abstract.
437Luzi, Paola et al, Genomics (1995), vol. 26(2), p. 407-9.
438Madsen J.S. & Qvist K.B. (1997) J. Food Sci. 62, 579-582.
439Maes et al., Analytica Chimica Acta, 284 (1993) 281-290.
440Mancini (1995) Food Formulating.
441Mao, Cungui, et al., "Cloning and Characterization of a Saccharomyces cerevisiae Alkaline Ceramidase with Specificity for Dihydroceramide", The Journal of Biological Chemistry, vol. 275, No. 40, 2000, pp. 31369-31378, NZAS-0683620.
442Maria Teres Neves Petersen, PhD, "Total Internal Reflection Fluorescence Flow System with Electrochemical Control", TIRF-EC Flow System, Sep. 2002, NZAS-0458374.
443Marion D et al pp. 245-260 of Wheat Structure Biochemistry & Functionality (ed Schofield JP) ISBN 085404777-8 published in 2000—(It states that it is the Proceedings of Conference organised by Royal Soc of Chemistry Food Chemistry Group held on Apr. 10-12, 1995, in Reading, UK. However, it is unclear why there was such a delay).
444Marion D et al—Chapter 6, pp. 131-p. 167 of "Interactions The Keys to Cereal Quality" 1998 ISBN 0 913250-99-6 (ed. Hamer & Hoseney).
445Marion Didier, et al., "Lipids, Lipid-Protein Interactions and the Quality of Baked Cereal Products," Interactions: The Keys to Cereal Quality, (ed. Hamer & Hoseney), Chapter 6, pp. 131-167 (1998).
446Marsh, Derek, et al., "Derivatised lipids in membranes. Physico-chemical aspexts of N-biotinyl phosphatidylethanolamines and N-acyl ethanolamines", Chemistry and Physics of Lipids, vol. 105, 2000, pp. 43-69, NZAS-0684062.
447Martinelle et al., "The Role of Glu87 and Trp89 in the lid of Humicola lanuginosa lipase", Protein Engineering, vol. 9, No. 6, 1996, pp. 519-524.
448Martinez, Chrislaine, et al., "Engineering cysteine mutants to obtain crystallographic phases with a cutinase from Fusarium solani pisi", Protein Engineering, vol. 6, No. 2, pp. 157-165, 1993.
449Martinez, Diego, et al., "Genome sequence of the lignocellulose degrading fungus Phanerochaete chrysosporium strain RP78", Nature Biology, May 2, 2004.
450Mase et al., "Purification and Characterization of a new Lipase from Fusarium sp. TM-30", Biosci. Biotech. Biochem., vol. 59, No. 9, pp. 1771-1772, 1995.
451Mason, Research Disclosure, Kenneth Mason Publications, Westbourne GB No. 390, Oct. 1996, pp. 661-662.
452Masuda, Naoko, et al., "Primary structure of protein moiety of Penicillium notatum phospholipase B deduced from the Cdna", Eur. J. Biochem., vol. 202, pp. 783-787, 1991.
453Matos AR, Lipid Catabolism: Lipid Degradation, 2000, p. 779-781.
454Matos, A.R., et al., "A Novel Patatin-like Gene Stimulated by Drought Stress Encodes a Galactolipid Acyl Hydrolase", FEBS Letters, 491: 188-192 (2001).
455Matos, A.R., et al., "A patatin-like protein with galactolipase activity is induced by drought stress in Vigna unguiculata leaves", Biochemical Society Transactions, vol. 28, part 6, 2000.
456Matos, AR et al, Febs Letters, 491 (2001) P188-192.
457Matsuda H et al, Biochim Biophys Acta, (1979), vol. 573(1), p. 155-65.
458Matsuoka, et al.; "Purification and properties of a Phospholipase C That has High Activity toward Sphingomyelin from Aspergillus Saitoi"; Biotiechonology and Applied Biochemistry (1987); vol. 9, pp. 401-409.
459Matthes et al, (1984) EMBO J. 3, p. 801-805.
460Max-Planck-Institut fur Kohlenforschung et al., "Controlling the enantioselectivity of enzymes by directed evolution: Practical and theoretical ramifications", NZAS-0441867.
461McAuley, Katherine E., et al., "Structure of a feruloyl esterase from Aspergillus niger", Acta Crystallographica, Section D, pp. 878-887, 2004.
462McCoy M G et al, Journal of Lipid Research (2002), vol. 43, pp. 921-929.
463McNeill G.P. & Berger R.G. (1993) Enzymatic glycerolysis of palm oil fractions and palm oil based model mixture: Relationship between fatty acid composition and monoglyceride yield, in Food Biotechnology 7: 75-87.
464McNeill, Gerald P., et al., "Further Improvements in the Yield of Monoglycerides During Enzymatic Glycerolysis of Fats and Oils", NZAS-0213370.
465McNeill, Gerald P., et al., "High-Yield Enzymatic Glycerolysis of Fats and Oils", JAOCS, vol. 68, No. 1, Jan. 1991.
466McNeill, Gerald P., et al., "Selective Distribution of Saturated Fatty Acids into the Monoglyceride Fraction During Enzymatic Glycerolysis", JAOCS, vol. 69, No. 11, Nov. 1992, NZAS-0213375.
467McNeill, Gerald P., et al., "Solid Phase Enzymatic Glycerolysis of Beef Tallow Resulting in a High Yield of Monoglyceride", NZAS-0213366, no date.
468Mechanism studies of the new lipase, Article, p. 1, No. 14.
469Memo: From Charlotte Johanson?, "Short introduction/ status on Ferulic Acid Esterases and Acetyl Xylan Esterases", Jan. 9, 2004.
470Meyer, V., et al., "Transcriptional regulation of the Antifungal Protein in Aspergillus giganteus", Mol Genet Genomics, 2002, vol. 266, pp. 747-757, NZAS-0567995.
471Meyers, Robert A., "Molecular Biology and Biotechnology—A Comprehensive Desk Reference" NZAS-015769.
472Michalski et al., "Photosynthetic apparatus in chilling-sensitive plants. VII. Comparison of the effect of galactolipase treatment of chloroplasts and cold-dark storage of leaves on photosynthetic electron flow", Biochimica et Biophysica Acta, vol. 589, pp. 84-99, 1980.
473Mielgo, I., et al., "Covalent immobilisation of manganese peroxidases (MnP) from Phanerochaete chrysosporium and Bjerkandera sp. BOS55", Enzyme and Microbial Technology, vol. 32, 2003, pp. 769-775, NZAS-0442378.
474Miller, Byron S., et al., "A Comparison of Cereal, Fungal, and Bacterial Alpha-Amylases as Supplements for Breadmaking", Food Technology, Jan. 1953, NZAS-0225991.
475Mine Y, Food Research International, 29(1), 1996, pp. 81-84.
476Mine, Y., "Application of the Enzymatic Methods to the Determination of Contaminated Yolk in Egg White", Food Research International, 29(1):81084 (1996).
477Ministerio da Ciencia e Tecnologia, Diario Oficial da Uniao, Jul. 15, 2003.
478Mogensen, Jesper E., et al., "Activation, Inhibition, and Destabilization of Thermomyces lanuginosus Lipase by Detergents", Biochemistry, vol. 44, pp. 1719-1730, 2005.
479Mohsen et al., "Specificity of Lipase Produced by Rhyopus Delemar and Its Utilization in Bread Making", Egypt. J Food. Sci. vol. 14, No. 1, pp. 175-182.
480Molecular Biological Methods for Bacillus—Chapter 3 (Ed. C.R. Harwood and S.M. Cutting) 1990, John Wiley and Sons Ltd, Chichester, UK.
481Mølgaard, Anne, et al., "Rhamnogalacturonan acetylesterase elucidates the structure and function of a new family of hydrolases", Structure, vol. 9, No. 4, 2000.
482Molochnaya Promyshlennost 1980 No. 11 21-25, 47—abstract from Food Sci & Tech Abs.
483Monick John A., Alcohols, Their Chemistry, Properties and Manufacture.
484Monographs for Emulsifiers for Foods, EFEMA Nov. 1985 2nd Edition.
485Moore, Charles M., et al., "Metal ion homeostasis in Bacillus subtilis", Current Opinion in Microbiology, 2005, vol. 8, pp. 188-195, NZAS-0572549.
486Morgan, Keith R., et al., "Stalling in Starch Breads: The Effect of Antistaling α-Amylase", Starch/Stärke, vol. 49, 1997, pp. 59-66.
487Morgan-Jones, Gareth; "Notes on Coelomycetes.II. Concerning the Fusicoccum Anamorph of Botryosphaneria Ribis"; vol. Xxx, pp. 117-125; Oct.-Dec. 1987.
488Morinaga et al Biotechnology (1984) 2, p. 636-639.
489Morten, T. & A., Letter, Rodovre, Jul. 2004.
490Mukherjee, Kumar D. et al., "Enrichment of y-linolenic acid from fungal oil by lipase-catalysed reactions", Appl. Microbiol Biotechnol (1991), vol. 35, pp. 579-584.
491Murakami, Mototake, et al., "Transesterification of Oil by Fatty Acid-Modified Lipase", Technical Research Institute, NZAS-0457255.
492Murakami, Nobutoshi, et al., "Enzymatic Transformation of Glyceroglycolipids into sn-1 and sn-2 Lysoglyceroglycolipids by use of Rhizopus arrhizus Lipase", Tetrahedron, vol. 50, No. 7, pp. 1993-2002, 1994.
493Mustranta, Annikka, et al., "Comparison of Lipases and Phosphlipases in the Hydrolysis of Phospholipids", Process Biochemistry, vol. 30, No. 5, pp. 393-401, 1995.
494N. V. Nederlandsch Octrooibureau Terms and Conditions.
495Nagano, et al.; "Cloning and Nucleotide Sequence of cDNA Encoding a Lipase from Fusarium keteroporum"; J. Biochem (1994); vol. 116; pp. 535-540.
496Nagao et al, J. Biochem 124, 1124-1129, 1998.
497Nagao et al, J. of Bioscience and Bioengineering vol. 89, No. 5, 446-450, 2000.
498Nagao et al, J. of Molecular Catalysis B: Enzymatic 17 (2002) 125-132.
499Nagao et al, JAOCS vol. 78, No. 2, 2001.
500Nagao, Toshihiro et al., "Cloning and Nucleotide Sequence of CDNA Encoding a Lipase from Fusarium heterosporum", J. Biochem., vol. 116, pp. 535-540, 1994, NZAS-0225015.
501Nagao, Toshihiro et al., "Expression of Lipase cDNA from Fusarium heterosporum by Saccharomyces cereviisiae: High-Level Production and Purification", Journal of Fermentation and Bioengineering, 1996, vol. 81, No. 6, pp. 488-492.
502Nagodawlthana et al., "Enzymes in Food Processing", Third Edition, 1993, Academic Press, Inc., NZAS-0665885.
503National Research Council (U.S.) Committee on Specifications of the Food Chemicals Codex, "Lipase Activity" in Food Chemicals Codex (1981) National Academy Press, Washington, D.C. pp. 492-493.
504Needleman & Wunsch (1970), J. of Molecular Biology 48, 443-453.
505Nelson and Long, Analytical Biochemistry (1989), 180, p. 147-151.
506Nerland A H, Journal of Fish Diseases, vol. 19, No. 2, 1996, pp. 145-150.
507Néron, et al., "Effects of lipase and the phosphlipase on the lipids hydrolysis during mixing in correlation with the oxygen consumption by wheat flour dough during kneading" available at http://www.cnam.fr/biochimie.
508Ness, Jon. E., et al., "DNA shuffling of subgenomic sequences of subtilisin" Nature Biotechnology, vol. 17, Sep. 1999, NZAS-0214844.
509Nestle Research Center, Brochure for "Food Colloids 2006" in Montreux, Switzerland, Apr. 23-26, 2006.
510Neugnot Virginie et al, European Journal of Biochemistry, 2002, vol. 269, pp. 1734-1745.
511Newport, G., et al., "KEX2 Influences Candida albicans Proteinase Secretion and Hyphal Formation", The Journal of Biological Chemistry, 1997, vol. 272, No. 46, pp. 28954-28961, NZAS-0685388.
512Nicolas, Anne, et al., "Contribution of Cutinase Serine 42 Side Chain to the Stabilization of the Oxyanion Transition State", Biochemistry, vol. 35, pp. 398-410, 1996.
513Nicolas, J., "Mise au Point sur I'action d'enzymers d'oxydoréduction en technologie boulangère. La maturation des farines de blétendre et le pétrissage des pâtes", Ann. Technol. Agric., 28(4):445-468 (1979); and English language translation of Abstract.
514Nicolas, J., "Mise au Point sur l'action d'enzymers d'oxydoreduction en technologie boulangere. La maturation des farines de bletendre et le petrissage des pates", Ann. Technol. Agric., 28(4):445-468 (1979); and English language translation of Abstract.
515Nielsen et al., "Lipases A and B from the yeast Candida antarctica", NZAS-0214451.
516Nierle W et al, Fette Seifen Anstrichmittel (1981), vol. 83(10), p. 391-395.
517Nierle, Von W. et al. "Weizenlipide: Funktion and Einflub bei der Verarbeitung des Mehles", NZAS-0208047.
518Nierle, W., et al., "Versuche zur Verlangerung der Haltbarkeit von Dartoffelprodukten", Chem. Mikrobiol. Technol. Lebensm., 1975, vol. 3, pp. 172-175, NZAS-0202446.
519Nobutoshi M et al, Tetrahedron Letters (1991), vol. 31(1), p. 1331-4.
520Novozymes Annual Report 2002, "Success for new baking enzyme."
521Novozymes D3.
522Novozymes data dated Jul. 17, 2005 entitled "Baking performance of prior art lipases from Humicola lanuginosa, Aspergillus tubigensis, Rhizopus delemar and Rhizomucor miehei, and their activity on galactolipids in dough".
523Novozymes Data E1.
524Novozymes Memo—Test of lipases for EP1193314B1, Jul. 6, 2005.
525Novozymes Report 2002 Annual Report.
526Novozymes, "Biowhitening—a new concept for steamed bread", BioTimes, Jan. 2005.
527Novozymes, "Breakthrough: Less Fattening Fried Food" BioTimes, Jun. 2001, No. 2.
528Novozymes, "Enzymes for dough strengthening", 2001.
529Novozymes, "Lipopan F BG", Cereal Foods.
530Novozymes, "Lipopan F BG—application and mechanism of a new lipase for bread baking" (Draft) Cereal Food (2003) (Author: Drost-Lustenberger, C. et al.).
531Novozymes, "Mechanism studies of the new lipase".
532Novozymes, "Product Sheet for Lipopan F BG", Cereal Food, (2001).
533Novozymes, "Product Sheet for Lipopan FS BG", Cereal Food (2002).
534Novozymes, "Product Sheet for Lipopan S BG", Cereal Food (2002).
535Novozymes, "Product Sheet for Noopazyme".
536Novozymes, "Product Sheet for Novozym 27016" (draft); Novozymes, "Product Sheet for Novozym 27041" (draft).
537Novozymes, "Product Sheet for Novozym 27019" (draft).
538Novozymes, "Product Sheet for Novozym 27080".
539Novozymes, "Product Sheet for Novozym 27106".
540Novozymes, "Product Sheet: Enzyme Business, Noopazyme" (draft).
541Novozymes, "Product Sheet: Enzyme Business, Novozym 27019" (draft).
542Novozymes, "Product Sheet: Enzyme Business, Novozym 677 BG".
543Novozymes, "Revolutionizing baking", BioTimes (2002) pp. 6-7.
544Novozymes, "Strong sales for lipase that makes dough stronger" BioTimes, Dec. 2003.
545Novozymes, "The Novozyme Touch: Make your mark on the future".
546Novozymes, "The perfect roll every time for steers", BioTimes, Sep. 2003.
547Novozymes, "The value of innovation", BioTimes, Mar. 2004.
548Novozymes, "The vital role of technical service in baking", BioTimes, Jun. 2004.
549Novozymes, Lipopan 50 BG, Product Sheet.
550Novozymes, Lipopan 50 BG, Product Specification.
551Novozymes, Lipopan F BG, Product Data Sheet.
552Novozymes, Lipopan FS BG, Product Sheet.
553Novozymes. Enzymes at work.
554NY metode til aktivitetsbestemme fedtnedbrydende vaskemiddelenzy.
555Nylander et al., "Interaction between lipids and lipases A collection of papers presented at the European Meeting on lipid and lipase interaction at Lund University", NZAS-0458953.
556Ognjenovic Radomir et al, Acceleration of ripening of semi-hard cheese by proteolytic and lipolytic enzymes.
557Ohm, J.B., et al., "Relationships of Free Lipids with Quality Factors in Hard Winter Wheat Flours", Cereal Chem., vol. 79, No. 2, pp. 274-278, 2002.
558Ohta, S. et al., "Application of Enzymatic Modification of Phospholipids on Breadmaking", Abstract from AACC 68th Annual Meeting in Kansas City, MO, Oct. 30th-Nov. 3, 1983, published in Cerial Foods World, p. 561.
559Ohta, Yoshifumi, et al., "Inhibition and Inactivation of Lipase by Fat Peroxide in the Course of Batch and Continuous Glycerolyses of Fat by Lipase", Agric. Biol. Chem., vol. 53, No. 7, pp. 1885-1890, 1989 NZAS-0211980.
560Okiy D.A. (1977) Partial glycerides and palm oil Crystallisation, in Journal of Science and Food Agriculture 28:955.
561Okiy D.A. (1978) Interaction of triglycerides and diglycerides of palm oil, in Oleagineux 33:625-628.
562Okiy D.A., Wright, W.B., Berger, K.G. & Morton I.D. (1978), The physical properties of modified palm oil, in Journal of Science of Food and Agriculture 29:1061-1068.
563Oluwatosin, Yemisi E., et al., "Mutations in the Yeast KEX2 Gene Cause a Vma-Like Phenotype: a Possible Role for the Kex2 Endoprotease in Vacuolar Acidification", Molecular and Cellular Biology, vol. 18, No. 3, pp. 1534-1543, Mar. 1998.
564Oluwatosin, Yemisi E., et al., "Phenotype: a Possible Role for the Kex2 Endoprotease in Vacuolar Acidification", Molecular and Cellular Biology, 1998, pp. 1534-1543, NZAS-0685378.
565O'Mahony et al. Hydrolysis of the lipoprotein fractions of milk by Phospholipase C.
566Orberg, Marie-Louise, "Self-assembly Structures Formed by Wheat Polar Lipids and their Interaction with Lipases", Master of Scient Thesis, Apr. 2005, NZAS-0436213.
567Orskov, Janne, et al., "Solubilisation of poorly water-soluble drugs during in vitro lipolysis of medium- and long-chain triacylglycerols", European Journal of Pharmaceutical Sciences, vol. 23, 2004. pp. 287-296, NZAS-0461574.
568Osman, Mohamed, et al., "Lipolytic activity of Alternaria alternata and Fusarium oxysporum and certain properties of their lipids", Microbios Letters, vol. 39, pp. 131-135, 1988, NZAS-0225041.
569Ostrovskaya L K et al, Dokl Akad Nauk SSSR, (vol. 186(4), p. 961-3) p. 59-61.
570O'Sullivan et al, J Plant Physiol, vol. 313, (1987) p. 393-404.
571O'Sullivan, J., et al., "A Galactolipase Activity Associated with the Thylakoids of Wheat Leaves (Triticum aestivum L.)", J. Plant Physiol., 131:393-404 (1987).
572Outtrup, Günther H., et al., "Properties and Application of a Thermostable Maltogenic Amylase Produced by a Strain of Bacillus Modified by Recombinant-DNA Techniques", Starch/Starke, vol. 36, No. 12, pp. 405-411.
573Palomo, Jose M., et al., "Enzymatic production of (3S, 4R)-(-)-4-(4′-fluorophenyl)-6-oxo-piperidin-3-carboxylic acid using a commerical preparation of lipase A from Candida antarctica: the role of a contaminant esterase" Tetrahedron: Asymmetry, vol. 13, 2002, pp. 2653-2659.
574Palomo, Jose M., et al., "Enzymatic resolution of (±)-glycidyl butyrate in aquenous media. Strong modulation of the properties of the lipase from Rhizopus oryzae via immobilization techniques", Tetrahedron: Asymmetry, vol. 15, 2004, pp. 1157-1161.
575Palomo, Jose M., et al., "Modulation of the enantioselectivity of Candida antarctica B lipase via conformational engineering: kinetic resolution of (±)-α-hydroxy-phenylacetic acid derivatives", Tetrahedron: Asymmetry, vol. 13, 2002, pp. 1337-1345.
576Pariza, Michael, et al., "Evaluating the safety of Microbiol Enzyme Preparations Used in Food Processing: Update for a New Century", Regulatory Toxicology and Pharmacology, vol. 33, pp. 173-186.
577Patent Abstracts of Japan Vo. 016, No. 528 (C-1001).
578Patent Abstracts of Japan vol. 016, No. 528 (C-1001), Oct. 29, 1992 & JP 04 200339 A see abstract.
579Patent Abstracts of Japan vol. 095, No. 001, Feb. 28, 1995 & JP 06 296467 A see abstract.
580Pazur, J.H., et al., "Comparison of the action of Glucoamylase and Glucosyltransferase on D-Glucose, Maltose, and Malto-Oligosaccharaides," Carbohydrate Research, 58:193-202 (1977).
581PCT International Search Report for PCT/DK96/00238, issued Apr. 11, 1996.
582Pedersen et al., 1996, J. Biol. Chem. 271:2514-2522 [10].
583Peelman F, et al, Protein Science Mar. 1998; 7(3): 587-99.
584Penninga et al, Biochemistry (1995), 3368-3376.
585Perella, F.W., Analytical Biochemistry, 174:437-447 (1988).
586Persson, Mattias, et al., "Enzymatic fatty acid exchange in digalactosyldiacylglycerol", Chemistry and Physics of Lipids, vol. 104, 2000, pp. 13-21, NZAS-0462330.
587Peters, G.H., et al., "Active Serine Involved in the Stabilization of the Active Site Loop in the Humicola lanuginosa Lipase", Biochemistry, 1998, vol. 37, pp. 12375-12383.
588Peters, G.H., et al.; "Dynamics of Rhizomucor miehei lipase in a lipid or aqueous environment: Functional role of glycines"; Dept. of Biochemistry and Molecular Biology, University of Leeds; NZAS-0031441.
589Peters, G.H., et al.; "Essential motions in lipases and their relationship to the biological function" NZAS-0031438.
590Peters, Günther H., et al., "Theoretical Investigation of the Dynamics of the Active Site Lid in Rhizomucor miehei Lipase", Biophysical Journal, vol. 71, 1996, pp. 119-129, NZAS-0668792.
591Philippine Patent Application Serial No. 31068 NZAS-0033694.
592Phytochemical Dictionary "Chapter 4, Sugar Alcohols and Cyclitols".
593Picon et al. Biotechnology letters vol. 17 nr 10 pp. 1051-1056.
594Plijter J and JHGM Mutsaers, The surface rheological properties of dough and the influence of lipase on it, Gist-brocades, Bakery Ingredients Division, Oct. 1994.
595Plou et al, J. Biotechnology 92 (2002) 55-66.
596Poldermans B and Schoppink P, "Controlling the baking process and product quality with enzymes", Cereal Foods World, Mar. 1999, 44 (3), p. 132-135.
597Ponte J G, Cereal Chemistry (1969), vol. 46(3), p. 325-29.
598Poulsen, C., et al., "Purification and Characterization of a Hexose Oxidase with Excellent Strengthening Effects in Bread", Cereal Chem., 75(1):51-57 (1998).
599Poulsen, C.H., et al., "Effect and Functionality of Lipases in Dough and Bread", The British Library, NZAS-0158559.
600Poulsen, Charlotte, et al. Purification and Characterization of a Hexose Oxidase with Excellent Strenghening Effects in Bread, NZAS-0409455.
601Product Data Sheet, Bakezyme P 500 BG, DSM Food Specialties, NZAS-0659329.
602Product Description PD 40084-7a Grindamyl Exel 16 Bakery Enzyme.
603Product Sheet B1324a-GB—LecitaseR Novo, Novo Nordisk.
604Product Sheet, Lipozyme® 10.000 L, Novo Nordisk, NZAS-0663762.
605Pub. No. 06-296467 (JP 6296467), Oct. 25, 1994, Section No. FFFFFF, vol. 94, No. 10, p. FFFFFF, FF, FFFF (FFFFFFFF) believed to be Patent Abstracts of Japan vol. 095, No. 001.
606Publication by Danisco (extracts from WO 99/31990).
607Publication by Danisco (Grindamyl SUREBake Bakery Enzymes).
608Publication by Danisco II.
609Publication by Danisco.
610Punt and van den Hondel, Meth. Enzym., 1992, 216:447-457.
611Pyler, E.J., "Baking Science and Technology Third Edition", vol. 1, 1988.
612Pyler, E.J., "Baking Science and Technology Third Edition", vol. II, 1988.
613Qi Si, J., "New Enzymes for the Baking Industry", Food Tech Europe, 3(1):60-64 (1996), Novo Nordisk Ferment Ltd.
614Qi Si, J., "New Enzymes for the Baking Industry", Food Tech Europe, 3(l):60-64 (1996), Novo Nordisk Ferment Ltd.
615Queener et al. (1994) Ann N Y Acad Sci. 721, 178-93.
616Raba, J., et al., "Glucose Oxidase as an Analytical Reagent", Critical Reviews in Analytical Chemistry, 25(1):1-42 (1995).
617Rambosek and Leach, CRC Crit. Rev. Biotechnol., 1987, 6:357-393.
618Rand, 1972, Journal of Food Science, 37:698-701.
619Rapp, Peter, et al., "Formation of extracellular lipases by filamentous fungi, yeasts, and bacteria", Enzyme Microb. Technol., 1992, vol. 14, November, NZAS-0225046.
620Rapp, Peter; "Production, regulation, and some properties of lipase activity from Fusarium Oxysporum f. sp. vasinfectum"; Enzyme and Microbial Technology(1995); vol. 17; pp. 832-838.
621Reetz M.T., Jaeger K.E. Chem Phys Lipids. Jun. 1998; 93(1-2): 3-14.
622Reetz Manfred T, Current Opinion in Chemical Biology, Apr. 2002, vol. 6, No. 2, pp. 145-150.
623Reiser J et al. (1990) Adv Biochem Eng Biotechnol. 43, 75-102.
624Richardson & Hyslop, pp. 371-476 in Food Chemistry, 1985, second edition, Owen R. Fennema (ed), Manel Dekker, Inc, New York and Basel.
625Richardson and Hyslop, "Enzymes: XI—Enzymes Added To Foods During Processing" in Food Chemistry, Marcel Dekker, Inc., New York, NY 1985.
626Richardson, Toby H., et al., "A Novel, High Performance Enzyme for Starch Liquefaction", The Journal of Biological Chemistry, vol. 277, No. 29, Issue of Jul. 19, pp. 25501-26507, 2002, NZAS-0564479.
627Roberts et al. (1992) Gene 122(1), 155-61.
628Roberts, et al.; "Extracellular Lipase Production by Fungi from Sunflower Seed"; Mycologia(1987); vol. 79(2); pp. 265-273.
629Roberts, Ian N., et al., Heterologous gene expression in Aspergillus niger: a glucoamylase-porcine pancreatic prophospholipase A2 fusion protein is secreted and processed to yield mature enzyme.
630Robertson et al, Journal of Biological Chemistry, 1994, 2146-2150.
631Rodrigues, et al.;"Short Communication: Bioseparations with Permeable Particles"; Journal of Chromatography & Biomedical Applications(1995); vol. 655; pp. 233-240.
632Rogalska, Ewa, et al., "Stereoselective Hydrolysis of Triglycerides by Animal and Microbial Lipases", Chirality, vol. 5, pp. 24-30, 1993.
633Rose, et al.;"Codehop (Consensus-Degenerate Hybrid Oligonucleotide Primer) PCR primer design"; Nucleic Acids Research(2003); vol. 31(13); pp. 3763-3766.
634Rousseau, Derick, et al., "Tailoring the Textural Attributes of Butter Fat/Canola Oil Blends via Rhizopus arrhizus Lipase-Catalyzed Interesterification. 2. Modifications of Physical Properties", J. Agric. Food Chem., vol. 1998, vol. 46, pp. 2375-2381, NZAS-0229125.
635Rydel, Timothy J. et al., "The Crystal Structure, Mutagenesis and Activity Studies Reveal that Patatin Is A Lipid Acyl Hydrolase with a Ser-Asp Catalytic Dyad", Biochemistry, 2003, vol. 42, pp. 6696-6708, NZAS-0301072.
636Sahm et al., 1973, Eur. J. Biochem. 37:250-256 [12].
637Sahsah, Y., et al., "Purification and Characterization of a Soluble Lipolytic Acylhydrolase from Cowpea (Vigna unguiculata L.) Leaves", Biochemica et Biophysica Acta, 1215: 66-73 (1994).
638Sahsah, Y., et al., "Enzymatic degradation of polar lipids in Vigna unguiculata leaves and influence of drought stress", Physiologia Plantarum, vol. 104, pp. 577-586, 1998.
639Sahsah, Y., et al., "Purification and characterization of a soluble lipolytic acylhydrolase from Cowpea (vigna unguiculata L.) leaves", Biochimica et Biophysica Acta, vol. 1215, pp. 66-73, 1994.
640Saiki R.K. et al Science (1988) 239, pp. 487-491.
641Saito, Kunihiko, et al., "Phospholipase B from Penicillium notatum", Methods in Enzymology, vol. 197, NZAS-0418833.
642Sakai, Norio, et al., "Human glactocerebrosidase gene: promoter analysis of the 5'-flanking region and structural organization", Biochimica et Biophysica Acta, vol. 1395, pp. 62-67, 1998.
643Sakaki T et al, Advanced Research on Plant Lipids, Proceedings of the International Symposium on Plant Lipids, 15th, Okazaki, Japan, May 12-17, 2002 (2003) p. 291-294, Publisher Kluwer Academic Publishers.
644Sales Range for Baking Improver and Premix Manufacturers from DSM Bakery Ingredients.
645Sambrook et al, Chapters 1, 7, 9, 11, 12 and 13—Molecular Cloning a laboratory manual, Cold Spring Harbor Laboratory Press (1989).
646Sambrook, J., et al. "A Laboratory Manual, Second Edition", Plasmid Vectors, 1989, NZAS-0665591.
647Sambrook, J., Fritsch, E.F. and Maniatis, T., 1989, Molecular Cloning, A Laboratory Manual 2nd Ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.
648Sanchez et al., "Solution and Interface Aggregation States of Crotalus atrox Venom Phospholipase A2 by Two-Photon Excitation Fluorescence Correlation Spectroscopy", Biochemistry, 2001, vol. 40, pp. 6903-6911.
649Sarney Douglas B. et al, "Enzymatic Synthesis of Sorbitan Esters Using a Low-Boiling-Point Azeotrope as Reaction Solvent", Biotechnology and Bioengineering, 1997, vol. 54(4).
650Saxena, et al.; "Purification Strategies for Microbial Lipases"; Journal of Microbilogical Methods (2003); pp. 1-18.
651Schagger, H. and von Jagow, G., 1987, Tricine-Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis for the Separation of Proteins in the Range from 1 to 100 kDa, Analytical Biochemistry 166:368-379.
652Schägger, H. and von Jagow, G., 1987, Tricine-Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis for the Separation of Proteins in the Range from 1 to 100 kDa, Analytical Biochemistry 166:368-379.
653Scheib et al.; "Stereoselectivity of Mucorales lipases toward triradylglycerols—A simple solution to a complex problem"; Protein Science (1999); vol. 8; pp. 215-221 (NZAS-0016084).
654Schiller, Jurgen, et al., "Lipid analysis of human spermatozoa and seminal plasma by MALDI-TOF mass spectrometry and NMR spectroscopy—effects of freezing and thawing" Chemistry and Physics of Lipids, vol. 106, 2000, pp. 145-156, NZAS-0564053.
655Schofield, J. David, "Wheat Structure, Biochemistry and Functionality", Department of Food Science and Technology, NZAS-0206986.
656Scopes, Robert K., "Section 8.4: Ultrafiltration" in Protein Purification Principles and Practice, Third Edition (1994) Springer-Verlag, New York, p. 267-9.
657Seffernick et al., Journal of Bacteriology, vol. 183, No. 8, p. 2405-2410 (2001).
658Seino, H. et al., JAOCS, Nov. 1984, vol. 61, No. 11, pp. 1761-1765.
659Semashko et al., Applied Biochem and Microbiol. (2003), vol. 39, p. 368-374.
660Sen S. et al., Developments in Directed Evolution for Improving Enzyme Functions, Appl. Biochem. Biotechnol., 2007, vol. 143, No. 3, p. 212-223.
661Sequence Alignment of database accession No. Q44268 (database: UNIProtKB/TrEMBL) with SEQ ID No. 16.
662Sequence Alignment of database accession No. Q44268 (database: UNIProtKB/TrEMBL) with SEQ ID No. 70.
663Sequence alignment of the nucleotide sequences of SEQ ID No. 2 of EP'167 and SEQ ID No. 7 of D20 and the amino acid sequences of SEQ ID No. 2 of EP'167 and SEQ ID No. 8 of D20.
664Sequence of enzyme GCAT (glycerophospholipidcholesterolacyltranspherase) (P10480, http://www.ncbi.nim.nih.gov/entrez/viewer.fcgi?db-protein&id=118572649) (1989).
665Shehata PhD Thesis.
666Shillcock, Julian C., et al., "Equilibrium structure and lateral stress distribution of amphiphilic bilayers from dissipative particle dynamics simulations", Journal of Chemical Physics, vol. 117, No. 10, Sep. 8, 2002, NZAS-0231167.
667Shillcock, Julian C., et al., "Tension-induced fusion of bilayer membranes and vesicles", Advance Online Publication, NZAS-0231181.
668Shimada et al, J. of Bioscience and Bioengineering vol. 91, No. 6, 529-538 (2001).
669Shimada et al, J. of Fermentation and Bioengineering vol. 75, No. 5, 349-352 (1993).
670Shimada et al, JAOCS vol. 71, No. 9, (Sep. 1994).
671Shin, et al.; "Butyl-Toyopearl 650 as a New Hydrophobic Adsorbent for Water-Soluable Enzyme Proteins"; Analytical Biochemistry(1984); vol. 138; pp. 259-261.
672Shogren, M.D., et al., "Functional (Breadmaking) and Biochemical Properties of Wheat Flour Components. I. Solubilizing Gluten and Flour Protein", Cereal Chemistry, vol. 46, No. 2, Mar. 1969, NZAS-0213171.
673Si, Joan Qi, "Enzymes, Baking, Bread-Making", NZAS-0255053.
674Si, Joan Qi, "Synergistic Effect of Enzymes for Breadbaking".
675Si, Joan Qi, et al. "Enzymes for bread, noodles and non-durum pasta".
676Si, Joan Qi, et al., "Novamyl—A true Anti-Staling Enzyme", Cereal Food, p. 1, No. 20.
677Si, Joan Qi, et al., "Synergistic Effect of Enzymes for Breadbaking".
678Si, Joan Qi; "New Enzymes for the Baking Industry"; Food Tech Europe (1996) pp. 60-64.
679Sias B et al, Biochemistry, (2004), vol. 43(31), p. 10138-48.
680Siew W.L. & Ng W.L. (1999) Influence of diglycerides on crystalisation of palm oil, in Journal of Science of Food and Agriculture 79:722-726.
681Siew W.L. & Ng W.L. (2000) Differential scanning thermograms of palm oil triglycerides in the presence of diglycerides, in Journal of Oil Palm Research 12:107.
682Siew W.L. (2001) Understanding the Interactions of Diacylglycerols with oil for better product performance, paper presented at the 2001 PIPOC International Palm Oil Congress—Chemistry and Technology Conference Aug. 20-23, 2001, Kuala Lumpur, Malaysia.
683Skovgaard, et al.;"Comparison of Intra—and extracellualr isozyme banding patterns of Fusarium Oxysporum"; Mycol. Res. (1998); vol. 102(9); pp. 1077-1084.
684Slotboom et al Chem. Phys. Lipids 4 (1970) 15-29.
685Smith and Whelan, Biochemical Preparations, vol. 10, p. 126-130 (1963).
686Smith, George P.; "The Progeny of sexual PCR"; Nature; vol. 370; No. 18; Aug. 4, 1994 (NZAS-0031326).
687Smith, Timothy L., et al., "The promoter of the glucoamylase-encoding gene of Aspergillus niger functions in Ustilago maydis", Gene. 88, 259-262, 1990.
688Sock, Jr. and Rohringer, R., 1988, Activity Staining of Blotted Enzymes by Reaction Coupling with Transfer Membrane-Immobilized Auxiliary Enzymes, Analytical Biochemistry 171:310-319.
689Soe, J.B., "Analyses of Monoglycerides and Other Emulsifiers by Gaschromatography", NZNA-0005896.
690Solares, Laura F., et al., "Enzymatic resolution of new carbonate intermediates for the synthesis of (S)-(+)-zopiclone", Tetrahedron: Asymmetry, vol. 13, 2002, pp. 2577-2582, NZAS-0215266.
691Sols and De Le Fuente, "On the substrate specificity of glucose oxidase", Biochem et Biophysica Acta (1957) 24:206-7.
692Sommer et al., "Genetic and Biochemical Characterization of a new Extracellular Lipase from Streptomyces cinnamomeus," Applied Environmental Microbiology, 1997, vol. 63, No. 9, p. 3553-3560.
693Sonntag N.O.V. (1982a) Glycerolysis of Fats and methyl esters—status, review and critique, in Journal of American Oil Chemist Society 59:795-802A.
694Soragni, Elisabetta, et al., "A nutrient-regulated, dual localization phospholipase A2 in the symbiotic fungus" The EMBO Journal, vol. 20, No. 18, pp. 5079-5090, 2001, NZAS-0230692.
695Sorensen, H.R., et al., "Effects of added enzymes on the physico-chemical characteristics on fresh durum-pasta".
696Sosland, Josh, "Alive and kicking", Milling & Baking News, Feb. 24, 2004.
697Soumanou, Mohamed M., et al., "Two-Step Enzymatic Reaction for the Synthesis of Pure Structured Triacylglycerides", JAOCS, vol. 75, No. 6, 1998.
698Spargeon, Brad, "In China, a twist: Forgers file patents" NZAS-0201747.
699Spendler, et al., "Functionality and mechanism of a new 2nd generation lipase for baking industry"—Abstract. 2001 AACC Annual Meeting; Symposia at Charlotte, NC. Oct. 14-18, 2001.
700Spradlin J E, Biocatalysis in Agric. Technol., ACS Symposium, 389(3), 24-43 (1989).
701Sreekrishna K et al (1988) J Basic Microbiol. 28(4), 265-78.
702Stadler et al., "Understanding Lipase Action and Selectivity", CCACAA, vol. 68, No. 3, pp. 649-674, 1995.
703Steinstraesser, et al., "Activity of Novispirin G10 against Pseudomonas aeruginosa in Vitro and in Infected Burns", Antimicrobial Agents and Chemotherapy, Jun. 2002, vol. 46, No. 6, pp. 1837-1844.
704Stemmer, Willem P.C.; "DNA shuffling by random fragmentation and reassembly: In vitro recombination for molecular evolution"; Proc. Natl. Acad. Sci. USA, vol. 91, pp. 10747-10751; Oct. 1994.
705Stemmer, Willem P.C.; "Rapid evolution of a protein in vitro by DNA shuffling"; Affymax Research Institute, Nature, vol. 370, Aug. 4, 1994, NZAS-0031401.
706Sternberg, M., "Purification of Industrial Enzymes with Polyacrylic Acids", Process Biochemistry, Sep. 1976, NZAS-0214848.
707Strickland, James A., et al., "Inhibition of Diabrotica Larval Growth by Patatin, the Lipid Acyl Hydrolase from Potato Tubers", Plant Physiol, vol. 109, pp. 667-674, 1995.
708Stryer, L., 1981, Biochemistry, 2nd ed, W.H. Freeman and Co., San Francisco.
709Sudbery et al (1988) Biochem Soc Trans. 16(6), 1081-3.
710Sugatani, Junko, et al., "Studies of a Phospholipase B from Penicillium Notatum Substrate Specificity and Properties of Active Site", Biochimica et Biophysica Acta, vol. 620, 1980, pp. 372-386, NZAS-0418825.
711Sugimoto et al., Agric. Biol. Chem. 47(6), 1201-1206 (1983).
712Sugiyama et al., "Molecular cloning of a second phospholipase B gene, caPLB2 from Candida albicans", Medical Mycology, vol. 37, 1999.
713Sullivan et al., 1973, Biochemica et Biophysica Acta, 309:11-22.
714Sullivan, James Denis Jr., Diss. Abstr. Int. B, 1973, 34(5), 1875, CAN 80: 105204 AN 1974: 105204 Caplus, "Purification and characterization of hexose oxidase from the red alga Chondrus crispus".
715Sullivan, James Denis Jr., Diss. Abstr. Int. B, 1973, 34(5), 1875, CAN 80: 105204 AN 1974: 105204 CAPLUS, "Purification and characterization of hexose oxidase from the end alga Chondrus crispus".
716Sullivan, James Denis Jr., Diss. Abstr. Int. B, 1973, 34(5), 1875, Can 80: 105204 AN 1974: 105204 Caplus, "Purification and characterization of hexose oxidase from the red alga Chondrus crispus."
717Svendsen, A. "Engineered lipases for practical use", Inform (1994) 5(5):619-623.
718Svendsen, Allan, "Lipase protein engineering" Biochimica et Biophysica Acta, vol. 1543, 2000, pp. 223-238, NZAS - 0157616.
719Svendsen, Allan, et al., "Biochemical properties of cloned lipases from the Pseudomonas family", Biochimica et Biophysica Acta, vol. 1259, 1995, pp. 9-17, NZAS-0214531.
720Sweigard, James A., et al., "Cloning and analysis of CUT1, a cutinase gene from Magnaporthe grisea", Mol. Gen. Genet., 232:174-182, 1992.
721Swinkels et al (1993) Antonie van Leeuwenhoek 64, 187-201.
722Sztajer H et al Acta Biotechnol, vol. 8, 1988, pp. 169-175.
723Talker-Huiber, Cynthia Z., et al., "Esterase EstE from Xanthomonas vesicatoria (Xv—EstE) is an outer membrane protein capable of hydrolyzing long-chain polar esters", Appl. Microbiol Biotechnol, 61:479-487, 2003.
724Terasaki, Masaru, et al., "Glycerolipid Acyl Hydrolase Activity in the Brown Alga Cladosiphon okamuranus Tokida", Biosci. Biotechnol. Biochem., vol. 67, No. 9, pp. 1986-1989, 2003.
725Testing of Hexose Oxidase in Baking.
726The First European Symposium of Enzymes on Grain Processing—Proceedings.
727The New Enzyme Operatives, Ingredient Technology, 50, Aug. 1997.
728Thommy L-G; Carlson, "Law and Order in Wheat Flour Dough; Colloidal Aspects of the Wheat Flour Dough and its Lipid and Protein Constitutents in Aqueous Media", Fortroligt, Lund 1981, NZNA-0006047.
729Thornton et al 1988 Biochem. Et Biophys. Acta. 959, 153-159.
730Tiss, Aly, et al., "Effects of Gum Arabic on Lipase Interfacial Binding and Activity", Analytical Biochemistry, vol. 294, pp. 36-43, 2001, NZAS-0572432.
731Toida J et al, Bioscience, Biotechnology, and Biochemistry, Jul. 1995, vol. 59, No. 7, pp. 1199-1203.
732Tombs and Blake, Biochim. Biophys (1982) 700:81-89.
733Topakas, E., et al. "Purification and characterization of a feruloyl esterase from Fusarium oxysporum catalyzing esterification of phenolic acids in ternary water—organic solvent mixtures", Journal of Biotechnology, vol. 102, 2003, pp. 33-44.
734Torossian and Bell (Biotechnol. Appl. Biochem., 1991, 13:205-211.
735Tsao et al. (1973) J Supramol Struct. 1(6), 490-7.
736Tschopp et al., 1987, Bio/Technology 5:1305-1308 [17].
737Tsuchiya, Atsushi et al, Fems Microbiology Letters, vol. 143, pp. 63-67.
738Tsuneo Yamane et al., "Glycerolysis of Fat by Lipase", Laboratory of Bioreaction Engineering, vol. 35, No. 8, 1986, NZAS-0213396.
739Tsychiya, Atsushi, et al., "Cloning and nucleotide sequence of the mono- and diacylglycerol lipase gene (md1B) of Aspergillus oryzae", FEMS Microbiology Letters, vol. 143, pp. 63-67, 1996.
740Turnbull, K.M., et al., "Early expression of grain hardness in the developing wheat endosperm", Planta, 2003, vol. 216, pp. 699-706.
741Turner, Nigel A., et al., "At what temperature can enzymes maintain their catalytic activity?", Enzyme and Microbial Technology, vol. 27, 2000, pp. 108-113, NZAS-0224067.
742Turner, Progress in Industrial Microbiology, Martinelli and Kinghorn (eds.), Elsevier, Amsterdam, 1994, 29:641-666.
743U.S. Appl. No. 09/824,053, filed Apr. 3, 2001.
744U.S. Appl. No. 09/932,923, filed Aug. 21, 2001.
745U.S. Appl. No. 10/040,394, filed Jan. 9, 2002.
746U.S. Appl. No. 10/150,429, filed May 17, 2002.
747U.S. Appl. No. 60/039,791, filed Mar. 4, 1997, Clausen.
748U.S. Appl. No. 60/039,791, filed Mar. 4, 1997, Kreij.
749U.S. Appl. No. 60/189,780, filed Mar. 16, 2000, Soe.
750U.S. Appl. No. 60/489,441, filed Jul. 23, 2003, Kreij.
751Underkofler, (1958), pp. 486-490.
752Unknown, "Appendix: Classification and Index of Fungi mentioned in the Text" in Unknown, p. 599-616.
753Unknown, "Section I: Structure and Growth—Chapter 1: An Introduction to the Fungi" in Unknown pp. 1-16.
754Unknown, Studies on Lipase (1964) p. 21.
755Uppenberg, Jonas, et al., "Crystallographic and Molecular-Modeling Studies of Lipase B from Candida antarctia Reveal a Stereospecificity Pocket for Secondary alcohols", Biochemistry, 1995, vol. 34, pp. 16838-16851 NZAS-0214433.
756Uppenberg, Jonas, et al., "The Sequence, crystal structure determination and refinement of two crystal forms of lipase B from Candida antarctica", Structure 1994, vol. 2, No. 4.
757Upton C et al TIBS Trends in Biochemical Sciences, Elsevier Publication (1995), vol. 20, pp. 178-179.
758USDA, "Production of an Industrially Useful Fungal Lipase by a Genetically Altered Strain of E. coli", New Technology.
759Uusitalo et al. (1991) J Biotechnol. 17(1), 35-49.
760Uwajima T et al, Agricultural and Biological Chemistry, 43(12), pp. 2633-2634, 1979.
761Uwajima T et al, Agricultural and Biological Chemistry, 44(9), pp. 2039-2045, 1980.
762Uwajima T et al, Methods in Enzymology, 89(41), pp. 243-248.
763Vaidehi, et al.; "Lipase Activity of Some Fungi Isolated from Groundnut"; Current Science (1984); vol. 53(23); p. 1253.
764van Binsbergen, Jan, et al., "Substitution of PHE-5 and ILE-9, Amino Acids Involved in the Active Site of Phospholipase A2 (PLA), and Chemical Modification of Enzymatically Generated (LYS-6)-PLA.", Proceedings of the 20th European Peptide Symposium, Sep. 4-9, 1988, University of Tubingen, Nzas-0457251.
765Van Den Berg. G, Regulatory status and use of lipase in various countries.
766van Gemeren, I.A., et al., "Expression and Secretion of Defined Cutinase Variants by Aspergillus awamori" Applied and Environmental Microbiology, vol. 64, No. 8, pp. 2794-2799, Aug. 1998.
767van Kampen, M.D., et al., "The phospholipase activity of Staphylococcus hyicus lipase strongly depends on a single Ser to Val mutation", Chemistry and Physics of Lipids, vol. 93, 1998, pp. 39-45, NZAS-0668780.
768van Nieuqenhuyzen, "Open Doors to baked goods".
769van Oort, Maarten G et al, Biochemistry 1989 9278-9285.
770van Solingen, Pieter, et al., "The cloning and characterization of the acyltransferase gene of penicillium chrysogenum", Agricultural University, Wageningen, The Netherlands.
771Vaysse et al J. of Biotechnology 53 (1997) 41-46.
772Verenium Corporation leaflet Purifine® Enzyme (Jan. 2008).
773Villeneuve, P., et al., "Lipase Specificities: Potential Application in Lipid Bioconversions", Inform, 8(6): 640-650 (1997).
774Villenueva, Inform, vol. 8, No. 6, Jun. 1997.
775Volc, J., et al., "Glucose-2 Oxidase Activity in Mycelial Cultures of Basidiomycetes", Folia Microbiol., 30:141-147 (1985).
776Vujaklija, Du{hacek over (s)}ica, et al., "A novel streptomycete lipase: cloning, sequencing and high-level expression of the Streptomyces rimosus GDS (L)-lipase gene", Arch. Microbiol, vol. 178, pp. 124-130, 2002.
777Wahnelt S.V., Meusel D, & Tülsner M, (1991) Zur kenntnis des diglyceride influsses auf das kristallisationsverhalten von Fetten, in Fat Science Technology 4:117-121.
778Waninge, Rianne, et al., "Milk membrane lipid vesicle structures studied with Cryo-TEM", Colloids and Surfaces B: Biointerfaces 31 (2003), pp. 257-264.
779Warmuth et al, 1992, Bio Forum 9, 282-283.
780Watanabe et al. Bio sci Biochem 63(5) 820-826, 1999.
781Watanabe, Yasuo et al., "Cloning and sequencing of phospholipase B gene from the yeast Torulaspora delbrueckii", FEMS Microbiology Letters, vol. 124, 1994, pp. 29-34, NZAS-0418803.
782Webb EC, Enzyme Nomenclature, 1992, p. 310.
783Weber et al. J Agric Food Chem 1985, 33, 1093-1096.
784Weipert, D., "Rheologie von Roggenteigen., II. Der Einfluss der Enzyme unterschiedlicher Spezifitat auf dzas rheologische Verhalten des Teiges", Getreide, Mehl Und Brot, 26(10):275-280 (1972); and English language translation of Abstract.
785Weipert, D., "Rheologie von Roggenteigen., II. Der Einfluss der Enzyme unterschiedlicher Spezifität auf dzas rheologische Verhalten des Teiges", Getreide, Mehl Und Brot, 26(10):275-280 (1972); and English language translation of Abstract.
786Welter, et al; "Identification of Recombinant DNA"; pp. 424-431.
787Wen-Chen Suen et al., "Improved activity and thermostability of Candida antarctica lipase B by DNA family shuffling", Protein Engineering, Design & Selection, vol. 17, No. 2, pp. 133-140, 2004, NZAS-0224073.
788Wen-Chen Suen et al., "Improved activity and thermostability of Candida antarctica lipase B by DNA family shuffling", Protein Engineering, Design & Selection, vol. 17, No. 2, pp. 133-140, 2004, NZAS-0717275.
789West S.; "Olive and Other Edible Oils"; Industrial Enzymology (1996); pp. 295-299.
790Whitaker, John R., et al., "Biocatalysis in Agricultural Biotechnology", ACS Symposium Series, NZAS-0213683.
791Whitehead, Michael, et al., "Transformation of a nitrate reductase deficient mutant of Penicillium chrysogenum with the corresponding Aspergillus niger and A. nidulans niaD genes", Mol Gen Genet, 216: 408-411, 1989.
792Wilhelm et al., "A Novel Lipolytic Enzyme Located in the Outer Membrane of Pseudomonas aeruginosa", Journal of Bacteriology, vol. 181, No. 22, Nov. 1999, pp. 6977-6986.
793Williams et al Protein Analysis by Integrated Sample Preparation, Chemistry, and Mass Spectrometry, Edited by Meyers.
794Wilson, R., et al., "Glucose Oxidase: An Ideal Enzyme", Biosensors and Bioelectronics, 7:165-185 (1992).
795Winnacker, Chapter 11, pp. 424-431 In From genes to clones: introduction to gene technology, VCH (1987).
796Winnacker, E. "Chapter 11: Identification of Recombinant DNA" in From Genes to Clones: Introduction to Gene Technology, 1987 John Wiley & Sons.
797Winther, Ole, et al., "Teaching computers to fold proteins", Physical Review, vol. 70, No. 030903, 2004, NZAS-0461499.
798Wirkowski et al., Biochemistry, vol. 38, No. 36, p. 11643-11650 (1999).
799Wirkung von Phospholipiden, "Struktur-Wirkungsbezehungen von Phospholipiden in Backwaren", NZAS-0301096.
800Wiseman, A., "Immobilization of Glucose Oxidase into Membranes as Sensors for Food Analysis", Elsevier Science Publishers, (1987).
801Withers-Martinez, C., et al., "A Pancreatic Lipase with a Phospholipase A1 activity: Crystal Structure of a Chimerica Pancreatic Lipase-Related Protein 2 from Guinea Pig", Structure 4(11): 1363-1374 (1996).
802Withers-Martinez, Chrislaine, et al., "A pancreatic lipase with a phospholipase A1 activity: crystal structure of a chimeric pancreatic lipase-related protein 2 from guinea pig", Structure, 1996, vol. 4, No. 11.
803Witt, Wolfgang et al., "Secretion of Phospholipase B From Saccharomyces Cerevisiae", Biochimica et Biophysica Acta, vol. 795, 1984, pp. 117-124, NZAS-0418813.
804Witteveen, C.F.B.: Thesis "Gluconate formation and polyol metabolism in Aspergillus niger", selected pages (1993).
805Wolff, A. M., O. C. Hansen U. Poulsen, S. Madrid, P. Stougaard (2001), Protein Expression and Purification., vol. 22, pp. 189-199. "Optimization of the Production of Chondrus crispus Hexose Oxidase in Pichia pastoris".
806Wolff, A. M., O. C. Hansen, U. Poulsen, S. Madrid, P. Stougaard (2001), Protein Expression and Purification., vol. 22, pp. 189-199. "Optimization of the Production of Chondrus crispus Hexose Oxidase in Pichia pastoris".
807Wolff, A. M., O. C. Hansen, U. Poulsen, S. Madrid, P. Stougard (2001), Protein Expression and Purification., vol. 22, pp. 189-199. "Optimization of the Production of Chondrus crispus Hexose Oxidase in Pichia pastoris".
808Wood et al., Eds., "Biomass, Part B, Lignin, Pectin, and Chitin", Methods in Enzymology (1988) vol. 161, Academic Press, San Diego.
809Woolley et al., "Lipases their structure, biochemistry and application", Cambridge University Press, NZAS-00354436.
810WPI Acc No. 93-298906(38) and JP05211852 Preparation of low fat content cream-by adding lipase to mixture of fat and water.
811Xu, Jun, et al., "Intron requirement for AFP gene expression in Trichoderma viride", Microbiology, 2003, vol. 149, pp. 3093-3097, NZAS-0572649.
812Yamaguchi et al, 1991, Gene 103:61-67.
813Yamane et al., "High-Yield Diacylglycerol Formation by Solid-Phase Enzymatic Glycerolysis of Hydrogenated Beef Tallow", JAOCS, vol. 71, No. 3, Mar. 1994, NZAS-0213713.
814Yamano Y, Surface activity of lysophosphatidyl choline from soybean.
815Yamauchi, Asao et al., "Evolvability of random polypetides through functional selection within a small library", Protein Engineering, vol. 15, No. 7, pp. 619-626, 2002, NZAS-0564005.
816Yang, Baokang, et al., "Control of Lipase-Mediated Glycerolysis Reactions with Butteroil in Dual Liquid Phase Media Devoid of Organic Solvent", J. Agric. Food Chem., 1993, vol. 41, pp. 1905-1909.
817Yeh, K-W, Juang, R.H and Su, J-C, A Rapid and efficient method for RNA isolation from plants with high-carbohydrate content, Focus 13 (3):102-103, 1991.
818Yount, Nannette Y., et al., "Multidimensional signatures in antimicrobial peptides", NZAS-0572406.
819Zaks, Aleksey, et al., "Enzyme-catalyzed processes in organic solvents", Proc. Natl. Acad. Sci. USA, vol. 82, pp. 3192-3196, May 1985.
820Zaks, Aleksey, et al., "The Effect of Water on Enzyme Action in Organic Media", The Journal of Biological Chemistry, vol. 263, No. 17, Issue of Jun. 15, pp. 8017-8021, 1988.
821Zangenbert, Niels Honberg, et al., "A dynamic in vitro lipolysis model 1. Controlling the rate of lipolysis by continuous addition of calcium", European Journal of Pharmaceutical Sciences, vol. 14, 2001, pp. 115-122.
822Zangenbert, Niels Honberg, et al., "A dynamic in vitro lipolysis model II. Evaluation of the model", European Journal of Pharmaceutical Sciences, vol. 14, 2001, pp. 237-244.
823Zhang, Hong, et al., "Modification of Margarine Fats by Enzymatic Interesterification: Evaluation of a Solid-Fat-Content-Based Exponential Model with Two Groups of Oil Blends", JAOCS, vol. 81, No. 1, 2004.
Referenced by
Citing PatentFiling datePublication dateApplicantTitle
US8460723 *Mar 21, 2011Jun 11, 2013Dupont Nutrition Biosciences ApsMethod of improving the properties of a flour dough, a flour dough improving composition and improved food products
US20120201928 *Mar 21, 2011Aug 9, 2012Danisco A/SMethod of improving the properties of a flour dough, a flour dough improving composition and improved food products
Classifications
U.S. Classification426/20, 426/549
International ClassificationA21D2/00
Cooperative ClassificationA23L1/16, C12Y301/01004, A21D8/042, C12Y301/01003, C12Y301/01026, A23L1/1055, A21D2/26, A21D2/16, C12Y101/03005
European ClassificationA23L1/16, A21D2/26, A23L1/105B, A21D8/04B, A21D2/16, C12Y101/03005, C12Y301/01004, C12Y301/01003, C12Y301/01026
Legal Events
DateCodeEventDescription
Oct 2, 2012FPAYFee payment
Year of fee payment: 8