The present invention also concerns use of a fermented product according to the present invention in the prevention and/or treatment of said diseases and disorders and for lowering serum levels of total cholesterol and/or LDL-cholesterol and/or

triglycerides and/or homocystein in subjects. In addition, the present invention also provides methods for preventing, treating, prophylactically treating and/or alleviating by therapy said diseases and disorders.

BACKGROUND OF THE INVENTION The field of functional foods has increased tremendously in recent years as the health awareness of the population in the industrialized parts of the world has gone up coincident with an increase in lifestyle related syndromes such as obesity, cardiovascular diseases and type 11 diabetes. Several studies show a significant correlation between the diet of a subject and the risk of the subject of contracting one or more of these diseases and syndromes. This has put the spotlight on the healthiness of everyday diet and as research into the field continues many items are added to the group of ingredients that should be present in a healthy diet. Functional foods can be defined as ordinary food items, which include one or more ingredients that can be beneficial to human health in one way or another. Functional foods can be fermented products, for instance dairy products, such as cultured dairy products.

Intake of soy protein, either alone or in combination with other soybean components, has been shown to have several beneficial effects on humans. Naturally, the soy protein could be consumed by eating soybeans, but this has little appeal, as soybeans are conceived by some to have an objectionable flavour and furthermore are not part of a traditional diet in the Western world. An alternative way of consuming soy protein is as part of a functional food product containing soy protein. An example of a desirable functional food product may be a cultured dairy product, such as a yoghurt or a fermented yoghurt-like product. These products do not require any cooking on the part of the consumer and can be eaten as they are.

Traditionally there has been an upper limit as to how much exogenous protein the cultured dairy product could contain without for example the flavour, texture, mouthfeel, taste and/or fermentation time being adversely affected. When manufacturing a well-fermented dairy product, such as for instance a yoghurt, with a low pH value, all the major components (excluding for instance flavourings) are preferably added before the addition of the bacterial culture and attempts to incorporate soy protein into yoghurt in increased amounts by use of traditional techniques will result in precipitation of soy proteins with free calcium ions of milk and lactic acid produced during the fermentation, giving a product with undesirable texture

and mouthfeel. Furthermore, most previous attempts to incorporate soy protein into yoghurt in increased amounts have also resulted in a prolonged fermentation time.

US 4.066. 792 concerns a method of producing soybean milk yoghurt. The soybeans are soaked in water at 20-30°C for about 8-24 hours (an amount of 3 to 5 parts water per part soybean). Then the soaked soybeans are treated with 0.1-1. 0% sodium bicarbonate at-70-100°C for 5-20 minutes using 5-15 parts sodium bicarbonate solution per part soybean, blended and treated with ultrasound to produce a soybean milk. The soybean milk is then mixed with 0-3% milk whey solids, heated to 90-121 °C for 10-20 minutes followed by rapid cooling. A solution of gelatine is added prior to the fermentation, which takes place for 16-24 hours using two strains of Lactobaccilus acidophilus. The soybean milk is described as having a protein content of 3.6%. Whey contains the water-soluble constituents of milk and is essentially a 5 percent solution of lactose in water, with small amounts of minerals, lactalbumin and beta-lactoglobulin.

Gelatine is a nutritionally incomplete animal protein substance derived from collagen.

Accordingly, it follows that the soy protein content of the end product will then be less than 3.6%.

US 4.797. 289 concerns a method of preparing a cultured dairy product by pasteurizing or heat treating a base mixture containing milk as the primary ingredient, cooling the mixture and fermenting the mixture to obtain a product with an increased amount of viable bacteria. It also describes adding fibres to the base mixture to enhance the growth of the bacteria and to obtain a dairy product with significant amounts of dietary fibres. The claimed methods provide a cultured dairy product with a content of dietary fibre of 0.1-3. 0 weight percent. Nothing is mentioned about the addition of exogenous protein.

US 5.147. 668 concerns a process for the production of a solid, dried product, which is reconstitutable by saliva, which process comprises homogenizing, heating, cooling and fermenting an aqueous mixture of milk or a milk product and carbohydrates followed by several steps to obtain the solid, dried product. It is stated that the aqueous mixture can include milk products enriched with soy protein. According to the examples, one of the initial steps of this process is a homogenization step comprising preheating of the aqueous mixture containing milk products and optionally soy protein at 60°C followed by homogenization with 200 bar. There is no description of any treatment of the soy protein prior to the addition thereof to the aqueous mixture containing milk products.

EP 0.486. 738 concerns a method for the production of a milk-fermented food, which method comprises addition of isolated soy protein to a medium comprising milk at a time just prior to the fermentation in order to obtain a lower fermentation time. The growth of bacteria proceeds remarkably fast and generation of acids is also accelerated by the addition of the isolated soy protein. The isolated soy protein is not treated prior to being added to the medium. The isolated protein is added in amounts up to 7.0 weight percent. It is stated that when the isolated soy protein is added at 4.8 weight percent, the concentration thereof in the final food product is 1.5 weight percent. It is stated that adding larger amounts of soy protein will not improve the growth accelerating effect but will result in a characteristic soybean odour in certain kinds of product. The inventors also suggest adding the soy protein after the fermentation and recommends that the isolated soy protein is added in an amount of about 0.1 to 1.5 weight percent as the addition of higher amounts will result in a product with a characteristic soybean odour.

US 5.017. 387 concerns a process for preparing yoghurt compositions using from 30 to 85 percent soy milk. Whole soybeans are soaked in water for 12-18 hours at room temperature with 3 parts water to one part soybean. After this, the soaked soybeans are blanched for 10-25 minutes at 90-95 °C and then milled to produce soy milk with a soy protein content of 3.1 to 3.6 percent. This soy milk is mixed with cow's milk, ewe's milk or other suitable milk and then pasteurized at preferably 85-95°C under high pressure, cooled and finally fermented. Appropriate times for the fermentation is stated to be from 5 to 10 hours with 5 to 6 hours being preferred.

EP 0.741. 975 concerns a process for producing yoghurt suitable for use in baking, which process comprises fermenting a cream mix comprising among others 2-15 weight percent of a protein component selected among milk, skimmed milk powder, milk proteins derived, for example, from whey protein and various vegetable proteins or combinations thereof. The cream mix is subjected to fermentation for 10 hours after for example pasteurization at 90°C for 5 minutes, homogenization at 100 bar and cooling.

US 4.971. 810 concerns a method of preparing fibre enriched yoghurt, which method comprises preparing a base mixture comprising milk and a fibre source, such as soy fibre, pasteurizing and homogenizing the base mixture followed by cooling and fermenting. This method results in the production of yoghurts with a fibre content of up

to 6 grams of fibre per 8 ounces, which corresponds to approximately 0.02 g/ml or 2.5%.

Only recently it has been found that by use of a process involving protein-hydration (EP patent application no. 00610033.3) it is possible to incorporate very high amounts of exogenous protein into cultured dairy products and at the same time avoid any adverse effects on flavour, texture, mouthfeel or taste of the immediate product or fermentation time. EP patent application no. 00610033.3 discloses cultured dairy products, such as yoghurt or yoghurt-like products, having high amounts of exogenously added protein, such as 7-8 weight percent, optionally in conjunction with dietary fibres. The amount of protein, which a cultured dairy product can contain, can thereby be increased with up to 50 percent compared to what was previously thought possible.

With EP patent application no. 00610033.3 a solution to perhaps the principal problem encountered, when trying to incorporate high amounts of exogenous protein, e. g. soy protein, in fermented milk products, has been provided. However, even though the teaching contained in EP patent application no. 00610033.3 was indeed a leap forward within the art, additional research has revealed that in order to be able to produce commercially interesting fermented milk products containing high amounts of exogenous protein, e. g. soy protein, additional problems have to be solved. In fact it has been found that the possibility to incorporate high amounts of exogenous protein into cultured dairy products without adversely affect the flavour, texture, mouthfeel or taste of the immediate product has revealed and to some extent emphasised and worsened other known problems. These problems must be solved in order to provide a commercial product attractive to the average consumer.

In particular the development and testing of cultured dairy products containing high amounts of exogenous protein, e. g. 7-8 weight percent soy protein, has revealed that even though the flavour, texture, mouthfeel or taste of the immediate products are not affected, the shelf-life of such high-protein-products is affected in a way, which severely compromises the consumer-estimated quality of, and hence commercial interest in, most products containing high amounts of exogenous protein, e. g. 7-8 weight percent soy protein. The primary effect on shelf-life observed is a time- dependent development of a bitter taste.

Bitterness as such has been recognised as a problem in traditional dairy products, e. g. cheese, for decades. Moreover, modern consumer preference for mild-flavored Cheddar-type cheese has lent greater significance to the impact of bitterness on dairy economics, since bitterness is a significant problem in Cheddar cheese and other common low-fat cheeses. Extensive research has revealed that within the field of traditional cultured dairy products, this off-flavour partially originates from bitter tasting peptides released by the breakdown of milk-proteins by proteolytic enzymes contained either in the raw-milk or released from the fermentation culture used. In addition, bitterness is caused by the accumulation of hydrophobic peptides produced by certain bacteria, supplied as part of the fermentation culture, and the action of chymosin on milk protein. The proteinase specificity of the fermentation culture used is the primary determinant in whether or not bitter peptides are produced. It is known that bitter peptides produced by the action of chymosin and the fermentation culture used can be degraded by intracellular peptidases from most commonly used fermentation cultures and adjunct bacteria, but the relative contribution of individual peptidases to these reactions remains unknown.

From the above it should be obvious that within the dairy-industry, it is a well known fact that the shelf-life of a cultured dairy product having a high milk-protein content can be affected by a time-dependent increase in the content of bitter tasting peptides.

Hence, the very nature of the problem is intimately linked to the specific kind of protein contained in the product, i. e. milk-protein, and its peptide composition, but also closely depends on other components in the raw-milk and the fermentation culture used.

The peptide composition and specific peptide-bonds present in a specific kind of protein, e. g. soy protein, is part of what inherently differentiates it from other kinds of protein, e. g. milk-protein. Hence, it will be easily understood that a shelf-life problem of the above nature, known to be caused partly by a high content of milk-proteins and partly by the specific fermentation culture used, would not per se be expected by a person skilled in the art to arise in the manufacturing of cultured dairy products containing high amounts of exogenous protein, e. g. soy protein.

In addition to the bitter tasting peptides, it has been contemplated that components that are associated with, but are not part of the exogenous protein, may add to the unexpected shelf-life problem. Two such components are isoflavons and saponins, both of which can be associated with soy protein. Due to the hydrophobicity of these components they are believed to be associated with the interior of the globular soy

protein molecule. According to one presently believed hypothesis the soy protein serves as a nitrogen source for the starter culture during fermentation and the proteolytic activity of the starter culture partly degrades the soy protein, thus facilitating the release of the bitter tasting components into the dairy product. The content of said components and their release into the dairy product will depend on both the source of the exogenously added protein and on the preparation. The release of bitter tasting components associated with the exogenously added protein have not previously been described and hence, would not per se be expected by a person skilled in the art to arise in the manufacturing of fermented products, for instance cultured dairy products, containing high amounts of exogenous protein, e. g. soy protein.

The problems with reduced shelf-life of cultured dairy products containing high amounts of exogenous protein, could furthermore never have been contemplated previously, since it has not previously, i. e. before EP patent application no.

00610033.3, been possible to incorporate so high amounts of exogenous protein in cultured dairy products, without adversely affecting flavour, texture, mouthfeel, taste and/or fermentation time. In conclusion, this indicates that, when exogenous proteins, e. g. soy protein, are concerned, neither the emergence nor the solution of this problem would be straight forward or in any way trivial to a person skilled in the art, even though similar problems have previously been described in the production of traditional cultured dairy products having a high milk-protein content.

According to EP patent application no. 00610033.3 the mixture to be fermented is heated in a vessel suitable for heating prior to the fermentation to achieve pasteurization, coagulation and resultant inactivation of any proteolytic enzymes contained therein. The reasoning behind this heat treatment is questionable and not explicitly stated. However, it might be inferred from EP patent application no.

00610033.3 that the reasoning behind the above-mentioned heating procedure, is similar to the reasoning behind the limited fermentation time preferably used, which allegedly eliminates the risk of the development of any bitter taste in the immediate resultant product stemming from the breakdown of the exogenously added protein during fermentation. However, the heat treatment described in EP patent application no. 00610033.3 does not eliminate the risk of bitter taste emerging in the product during storage and the shelf-life of a product manufactured according to EP patent application no. 00610033.3 is still affected by a time-dependent development of a bitter taste.

It has now been found that it is possible by use of a new process as described below, to avoid any negative effects on the shelf-life of a fermented product for instance a cultured dairy product, such as yoghurt or yoghurt-like products, containing high amounts of exogenous protein, such as 7-8 weight percent, optionally in conjunction with dietary fibres, which are due to a time-dependent increase in the content of bitter taste, in the form of bitter tasting peptides or the release of bitter components originating from the breakdown of the exogenously added protein.

In view hereof, it is an object of the present invention to provide a fermented product, for instance a cultured dairy product, such as a yoghurt or a yoghurt-like product, containing at least 5 weight percent of exogenously added protein, with an improved shelf-life, while retaining a smooth texture, avoiding soy odour.

SUMMARY OF THE INVENTION The present invention provides a method for manufacturing a fermented product containing a high amount of incorporated protein having an improved shelf-life, characterised by the addition of material enabling a reduction of bitter taste. The present invention further provides a method for manufacturing a cultured dairy product containing exogenously added protein having an improved shelf-life, characterised by the addition of material enabling a reduction of bitter taste. Said method is further characterised by the addition of one or several entities exhibiting proteinase activity or excreting or liberating proteinases enabling the degradation of bitter tasting peptides and/or the addition of a hydrolysate resulting in reduced release of bitter tasting components associated with exogenously added protein. According to one hypothesis the addition of a hydrolysate which may serve as an easy accessible source of nitrogen for the starter culture will reduce the breakdown of the exogenous protein and thus reduce the release of bitter components associated with the exogenious protein as well as the release of bitter peptides into the product. Any negative effects on the shelf-life of the cultured dairy product due to a time-dependent increase in the content of bitter tasting peptides and/or the release of bitter tasting components originating from the breakdown of exogenously added protein can thus be avoided by the above means. The present invention also provides a cultured dairy product having a smooth texture and comprising exogenously added protein having an improved shelf-life.

Intake of a protein enriched cultured dairy product prepared by a method according to the present invention will provide any beneficial effects associated with the

exogenously added proteins. Furthermore, a protein enriched cultured dairy product prepared by a method according to the present invention will be able to comply better with consumer-expectations than cultured dairy products prepared by use of previously available methods for protein fortification of cultured dairy products, since, due to the fact that a time-dependent increase in the content of bitter tasting peptides originating from the breakdown of exogenously added protein as well as the release of bitter tasting components associated with the exogenously added protein is avoided, it has an improved shelf-life.

In one aspect, the present invention provides a method for manufacturing a fermented product containing exogenously added protein having an improved shelf-life, characterised by the addition of material enabling the reduction of bitter taste. Said material more specifically comprises one or several entities exhibiting proteinase activity or excreting or liberating proteinases enabling the degradation of bitter tasting peptides and/or a hydrolysate resulting in reduced release of bitter tasting components associated with the exogenously added protein.

In another aspect, the present invention provides a method for manufacturing a cultured dairy product containing exogenously added protein having an improved shel- life which method comprises the steps of i) preparing a mixture of a protein source and a milk composition, ii) adding a fermentation culture to the mixture from step i), and iii) fermenting to obtain a cultured dairy product. wherein said fermentation culture added in step ii) at least partly comprises one or several entities exhibiting proteinase activity or excreting or liberating proteinases enabling the degradation of bitter tasting peptides and/or that said mixture in step i) at least partly comprises a hydrolysate resulting in reduced release of bitter tasting components associated with the exogenous protein.

In another aspect, the present invention provides a fermented product prepared by a method according to the present invention.

In yet another aspect, the present invention provides a cultured dairy product having a smooth texture and comprising exogenously added protein in an amount of from about

5 to 25 weight percent, optionally exogenously added dietary fibres, and containing material to reduce bitter taste e. g. entities exhibiting proteinase activity or excreting or liberating proteinases enabling the degradation of bitter tasting peptides and/or a hydrolysate resulting in reduced release of bitter tasting components associated with the exogenously added protein.

In yet another aspect, the present invention provides the use of a cultured dairy product having a smooth texture and comprising exogenously added soy protein in an amount of from about 5 to 25 weight percent, and exogenously added soy fibres, and which is devoid of any significant content of bitter tasting peptides and other bitter tasting components for lowering serum levels of LDL-cholesterol.

The exogenously added protein may be any type of protein including dairy proteins or any combination of proteins and may consequently be provided by any protein source or by a combination of protein sources. Examples of exogenously added proteins in the context of the present invention are vegetable proteins, such as soy proteins, and animal proteins. The exogenously added protein is preferably non-dairy protein and more preferably soy protein. The soy protein is preferably provided by isolated soy protein, soy protein concentrate, soy flour or the like or any combination thereof. The soy protein is preferably provided by isolated soy protein.

Isolated soy protein is the major proteinacious fraction of soybeans. It is prepared from high quality, dehulled, defatted soybeans by removing a preponderance of the non- protein components resulting in an isolated soy protein fraction which shall contain at least 80 percent protein on a moisture free basis. Soy protein concentrates are made by removing most of the oil and water-soluble non-protein constituents from defatted and dehulled soybeans. In the present context a soy protein concentrate shall preferably contain at least 65 percent protein on a moisture-free basis. The soy protein may also be provided by soy flour, which may be full-fat or defatted soy flour. Full-fat soy flour comes from whole, dehulled soybeans that have been ground into a fine powder and, as the name implies, still contains the fat naturally found in soybeans.

Defatted soy flour comes from whole, dehulled, defatted soybeans that have been ground into a fine powder. Soy flour contains approximately 50 percent protein on a dry weight basis in the present context.

Preferred isolated soy protein products are supplied by Protein Technologies International, Inc. under the brand name of SUPROO. SUPROO isolated soy protein

products are supplied in many different qualities and SUPRA@ XT 12C, SUPROt) PLUS 159, SUPRO@ PLUS 161 and SUPRA PLUS 219 are the presently preferred qualities. SUPRO@ PLUS 161 and SUPRO PLUS 219 has a higher content of soy protein than SUPRA PLUS 159 (87.3% in SUPRO@ PLUS 161 and SUPRO PLUS 219 compared to 80% in SUPRO PLUS 159) and might give a bit more soy flavour in the cultured dairy product, but has less powdery feeling in itself and reduces fermentation times compared with SUPROtl PLUS 159 and are thus more preferred qualities. SUPRA0 PLUS 219 results in a product less prone to develop a bitter taste during storage than a product based on SUPRO@ PLUS 161 and is thus the most preferred quality. The source of the exogenously added protein may also be a mixture of any of the protein sources mentioned above.

A protein source for use in a method according to the present invention comprises the source or sources of exogenously added proteins. The amount of protein in a protein source for use in a method according to the present invention and the amount of the protein source is preferably such that the amount of exogenously added protein in the resulting cultured dairy product is at least about 5 weight percent, such as at least about 6 weight percent, for example at least about 7 weight percent, such as at least about 8 weight percent, for example at least about 9 weight percent, such as at least about 10 weight percent, for example at least about 11 weight percent, such as at least about 12 weight percent.

Optionally, a protein source for use in a method according to the present invention also comprises dietary fibres. Examples of dietary fibres comprised in a protein source for use in step i) of a method according to the present invention include fibres from apples, oats, and soybeans. The dietary fibres are preferably provided by soybean fibres, and more preferably by soy cotyledon fibres. Preferred soy cotyledon fibre products are supplied by Protein Technologies International, Inc. under the brand name of FIBRIM (E). Among the various soybean fibres produced under the FIBRIM (g) brand, FIBRIMO 1020 and FIBRIMO 2000 are particularly preferred in the present invention. These two products also contain soy protein in an amount of 12.4% and 13.3%, respectively. The manufacturers recommend FIBRIM# 2000 if a powdery mouth feel is to be avoided. FIBRIM (g) 1020 is another quality of cotyledon fibres, which in our independent studies contributed less to a powdery mouth feel than FIBRIMO 2000.

The amount of dietary fibres in a protein source for use in step i) of a method according to the present invention may preferably vary from 0 to about 40 weight percent based on the weight of the protein.

Preferably, the amount of protein and the amount of dietary fibres in a protein source for use in a method according to the present invention and the amount of said protein source is such that the resulting cultured dairy product has a content of exogenously added protein of about 9 weight percent and a content of exogenously added dietary fibres of about 2.25 weight percent.

A protein source for use in a method according to the present invention may also optionally comprise carbohydrate sources, fat sources, flavouring agents, vitamins, minerals, electrolytes, trace elements and other conventional additives and the like. If a fat source is present in a protein source for use in step i) of a method according to the present invention, a preferred fat source is lecithin, especially soy lecithin. When lecithin, or soy lecithin, is present in a protein source for use in step i) of a method according to the present invention, it is usually present in an amount of from about 0.5 to 10 weight percent, preferably from about 2.5 to 6 weight percent of the total protein source.

Preferred examples of protein sources for use in a method according to the present invention are the compositions described in WO 97/31546, which are hereby incorporated by reference. Said patent application discloses compositions comprising (a) isolated soy protein, (b) soybean fibres, preferably soy cotyledon fibres, the amount of (a) being such that the protein content provides at least 15% of the total energy content of the composition, and the weight ratio between (a) and (b) being at least 2, preferably at least 3. These compositions are useful for lowering serum levels of cholesterol and triglycerides and for increasing the HDL/LDL-cholesterol ratio in subjects and for treating obesity.

Additionally preferred examples of protein sources are the compositions described in PCT/IB99/01992, PCT/IB99/01997 and PCT/IB99/01998. Said patent applications disclose compositions comprising (a) a soy protein source, selected from isolated soy protein, soy protein concentrate, or soy flour, of which isolated soy protein is most preferred, said soy protein source providing an amount of soy protein, which is at least 45 weight percent of the total protein content of the composition, preferably at least 50 weight percent of the total protein content of the composition, said total protein content

providing at least 15 percent of the total energy content of the composition, (b) at least one phytoestrogen compound in an amount of more than 0.10 weight percent of the soy protein content of the composition, and (c) dietary fibres, preferably soybean fibres, more preferably soy cotyledon fibres, in an amount of more than 4 weight percent of the total weight of the composition on a dry basis. These compositions are particularly useful for lowering serum levels of total cholesterol, LDL-cholesterol, triglycerides, homocystein, reducing the influx of cholesterol and/or triglycerides into the arterial wall, reducing the amount of oxidized LDL-cholesterol present in the arterial wall, increasing the serum HDL/LDL-cholesterol ratio and/or the serum level of HDL-cholesterol in a subject, including a diabetic subject, reducing and/or eliminating mucus hypersecretion and/or dyspnea in a subject suffering from asthma and/or increasing FEV1 of a subject as measured by forced expiratory volume in the first second of expiration. An example of these compositions is shown to be able to lower serum levels of inter alia LDL-cholesterol with as much as 13%. These compositions may therefore be effective in preventing, treating, prophylactically treating and/or alleviating diseases such as cardiovascular diseases, type 11 diabetes, cardiovascular diseases in diabetics, the metabolic syndrome and pulmonary diseases as described in said applications.

One embodiment of the present invention provides methods by use of which the above-mentioned compositions may be incorporated into cultured dairy products in amounts, which methods result in cultured dairy products with increased amounts of exogenously added soy protein having improved shelf-life. Another embodiment of the present invention provides cultured dairy products into which the above-mentioned compositions have been incorporated and having improved shelf-life. Intake of such a cultured dairy product according to the present invention will provide the beneficial effects associated with said compositions. Furthermore, a cultured dairy product prepared by a method according to the present invention will be able to comply better with consumer-expectations than cultured dairy products prepared by use of previously available methods for protein fortification of cultured dairy products, since, due to the fact that a time-dependent increase in the content of bitter tasting peptides originating from the breakdown of exogenously added protein and/or the release of bitter tasting components associated with the exogenously added protein is avoided, it has an improved shelf-life.

One aspect of the present invention provides the use of cultured dairy products as described in the above paragraph for lowering serum levels of glucose and/or total

cholesterol and/or LDL-cholesterol and/or triglycerides and/or homocystein and/or for increasing serum levels of HDL-cholesterol and/or the serum HDULDL-cholesterol ratio in a subject.

The milk ingredients in a milk composition for use in step ia method according to the present invention may for instance be provided by cream, whole milk, partially skimmed milk, skimmed milk, reconstituted non-fat dairy milk, cream powder, whole milk powder, partially skimmed milk powder, skimmed milk powder and/or combinations thereof. A milk composition for use in a method according to the present invention preferably contains a combination of milk and milk powder, such as for example a combination of skimmed milk and skimmed milk powder. The amount of milk powder added to the liquid milk may preferably be such that it can make up for the dilution that takes place when a hydrated protein source is added to the milk composition in a method according to the present invention. The milk ingredients of a milk composition for use in a method according to the present invention seem to contribute to a better taste of the resulting cultured dairy product as it helps in masking any soy taste that might otherwise be present. The milk ingredients of a milk composition for use in a method according to the present invention should be present in an amount which is sufficient to ensure that the resulting cultured dairy product contains at least about 2 g, preferably from about 5 to about 5.5 g of milk ingredients (dry matter) per 100 g of cultured dairy product.

A milk composition for use in a method according to the present invention may also comprise other ingredients, such as water, stabilizers and for instance soy milk.

A milk composition for use in a method according to the present invention may also contain a water soluble calcium ion binder such as a salt selected from the group comprising sodium or potassium citrate, sodium or potassium phosphate, or sodium or potassium hexametaphosphate, or a combination of these, whereof sodium hexametaphosphate is most preferred. A preferred sodium hexametaphosphate is supplied by Ellis & Everand, Bradford, UK, under the brand name Calgon. Sodium citrate, potassium citrate, sodium phosphate, potassium phosphate or a combination thereof is preferably present in a milk composition for use in a method according to the present invention in an amount of from 0 to about 0.2 weight percent, preferably in an amount of about 0.1 weight percent of the cultured dairy product. Sodium hexametaphosphate or potassium hexametaphosphate or a combination thereof is preferably present in a milk composition for use in a method according to the present

invention in an amount of from 0 to about 0.15 weight percent, preferably in an amount of from about 0.08 to about 0.10 weight percent of the cultured dairy product. The presence of a calcium ion binder in a milk composition for use in a method according to the present invention ensures that fewer free calcium ions are available for precipitation with exogenously added proteins. For the purpose of the present invention up to 15 memo)/) of the calcium ions in milk may be bound by a calcium ion binder.

A hydrolysate for use in a method according to the present invention comprises a compound or mixture of compounds comprising a nitrogen-source for instance in the form of hydrolised casein, as contained in the product marketed under the name of Casamino acids and manufactured by Difco. The nitrogen-source for use in a method according to the present invention is preferably present in an amount corresponding to that present when the amount of Casamino acids in the resulting fermented product is for instance between about 0.02 to about 2 weight percent, such as for example about 0.05 weight percent, such as for example about 0.10 weight percent, such as for example about 0.15 weight percent, such as for example about 0.20 weight percent, such as for example about 0.30 weight percent, such as for example about 0.50 weight percent, such as for example about 0.75 weight percent, such as for example about 1.0 weight percent, such as for example about 1.5 weight percent.

The entities exhibiting proteinase activity for use in a method according to the present invention comprises bacterial proteinases for instance in the form of heat inactivated Lactobacillus helveticus, as contained in the product marketed under the name of Enzobact and manufactured-by Medipharm. The bacterial proteinases for use in a method according to the present invention is preferably present in an amount corresponding to that present when the amount of Enzobact in the resulting fermented product is for instance between about 0.02 to about 0.2 weight percent, such as for example about 0.03 weight percent, such as for example about 0.04 weight percent, such as for example about 0.05 weight percent, such as for example about 0.06 weight percent, such as for example about 0.07 weight percent, such as for example about 0.08 weight percent, such as for example about 0.09 weight percent, such as for example about 0.10 weight percent, such as for example about 0.12 weight percent, such as for example about 0.14 weight percent, such as for example about 0.16 weight percent, such as for example about 0.18 weight percent.

The present invention provides a fermented product comprising a protein source, preferably having a high, fixed amount of a phytoestrogen compound such as e. g. naturally occurring isoflavones, and a phospolipid source. More particularly the present invention provides a fermented product on basis of soybean extractable ingredients comprising soy lecithin, preferably having a high fixed level of phosphatidyl choline, and having a high, fixed amount of a phytoestrogen compound such as e. g. naturally occurring isoflavones.

The present invention provides a fermented product comprising a) soy protein, preferably isolated soy protein, b) a high content of a plant hormone in the form of a phytoestrogen compound, preferably naturally occurring isoflavones, (c) a phospholipid source, more preferably lecithin, and even more preferably soy lecithin and preferably having a high fixed level of phosphatidyl choline and optionally (d) dietary fibers, preferably soybean fibers, more preferably soybean fibers manufactured from the cotyledon of soybeans hereinafter referred to as soy cotyledon fibers and the present invention furthermore represents a potential new breakthrough in the treatment of cardiovascular diseases, diabetes and pulmonary diseases.

A fermented product according to the present invention is useful in treating including prophylactically treating cardiovascular diseases such as hypercholesterolemia, hypertriglyceridemia, hyperlipidemia and other cardiovascular diseases such as e. g. arteriosclerosis. It is one objective of the present invention to significantly lower levels of total serum cholesterol and LDL-cholesterol and triglycerides in a mildly hypercholesterolemic subject. It is another objective of the present invention to significantly lower serum levels of total cholesterol and/or LDL-cholesterol and/or triglycerides in a subject suffering from hypercholesterolemia and/or hyperlipidemia. It is another objective of the present invention to render the arterial wall more resistant to the accumulation of lipoproteins. It is a further objective of the present invention to provide a fermented product effective in preventing, treating, prophylactically treating and/or alleviating an arteriosclerotic condition by reducing the influx of cholesterol and/or triglycerides into the endocelium of the arterial wall and/or by causing the dilation of blood vessels. Yet another objective of the present invention is to reduce lipid plaque formation.

The present invention is also useful in the prevention and/or treatment of type 2 diabetes and/or a cardiovascular disease in diabetic subjects. Accordingly, it is an objective of the present invention to effectively lower serum levels of both glucose and

cholesterol and/or triglycerides. No treatment is currently available for concomitantly lowering serum levels of glucose as well as lipid serum levels of total cholesterol and/or LDL-cholesterol and/or triglycerides. It is to be understood that diabetic subjects according to the present invention have a fasting plasma glucose 0 7.0 mmol/l.

A fermented product according to the present invention represents a new approach to treatment of type 2 diabetes and is believed to be capable of i) lowering total serum levels of cholesterol and/or LDL-cholesterol and/or triglycerides and/or increasing serum levels of HLDL-cholesterol, ii) increasing glucose tolerance and/or insulin sensitivity and/or, iii) lowering serum levels of glucose, iv) preventing, treating and/or alleviating impaired glucose tolerance and/or insulin secretory failure in diabetic subjects and/or v) preventing, treating and/or alleviating an arteriosclerotic condition by reducing the influx of cholesterol and/or triglycerides into the endocelium of the arterial wall of a diabetic subject suffering from a cardiovascular disease and/or by causing the dilation of blood vessels. No other known fermented products are effective in lowering serum levels of both lipids and glucose and/or reducing the influx of lipids such as e. g. cholesterol and/or triglycerides into the arterial wall.

The present invention is also useful in the prevention and/or effective treatment of pulmonary diseases such as e. g. airway inflammation, asthma, bronchitis and small airways diseases, in particular asthma including chronic asthma such as e. g. asthma characterized by a chronic inflammatory condition. The present invention is believed to be capable of increasing FEV1 of a subject, measured by forced expiratory volume in the first second of expiration, as well as being capable of treating, alleviating and/or eliminating in particular i) inflammation of the airways, ii) mucus hypersecretion, and iii) bronchoconstriction.

Phytoestrogen compounds are naturally occurring plant hormones showing a structural similarity to 17a-estradiol. Phytoestrogens consist of a number of classes including isoflavones, coumestans, lignans and resorcylic acid lactones. The class of isoflavones consists of among others genistein, daidzein, equol, glycitein, biochanin A, formononetin, and O-desmethylangolesin. The isoflavones genistein and daidzein are found almost uniquely in soybeans. When present in the plant the isoflavones are mainly in a glucoside form, i. e. attached to a sugar molecule. Isoflavones in this glucoside form can be deconjugated to yield isoflavones in a so-called aglycone form, which is the biologically more active form of isoflavones and which is absorbed faster and to a greater extent in the human gut than isoflavones in the glucoside form. In vitro

studies have examined the relative estrogenic effect exerted by various phytoestrogens including isoflavones. The resulting potencies as compared to estradiol (having a relative potency of 100), have been reported by Knight (Maturitas 22,167-175 (1995) ) for among others genistein (0.084) and daidzein (0.013).

However, the results also showed that the estrogen receptor complexes formed by estradiol and isoflavones such as genistein and daidzein are functionally equivalent.

The comparative dissociation constant of genistein for the estrogen receptor, as determined in competitive binding assays, was found to be from 100 to 10.000 times higher than that of estradiol.

Soy proteins are involved in a reduction of cholesterol and triglyceride levels, they are easily digestible, and they represent an efficient sole protein source for maintaining the nitrogen balance. Soy isoflavones in high intakes further enhances this effect.

Phospholipids, such as soy lecithins, especially soy phosphatidyl choline have been shown to effect total serum cholesterol levels and/or to increase serum HLDL- cholesterol levels. Dietary fibers, such as soybean fibers, especially soy cotyledon fibers have been shown to lower total serum cholesterol levels, to improve glucose tolerance, to increase insulin sensitivity, to normalize the gastrointestinal function, and to exert no influence on the absorption of essential minerals.

The term"naturally occurring"substance as used in the present specification refers to a substance originally isolated from a natural source, such as an animal or a plant, for example a soy plant, or modified forms of such a substance. The naturally occurring substance for use in a fermented product according to the present invention may be included in a fermented product according to the present invention as part of the natural source or in any type of extract, isolate or the like thereof, or it may have been isolated from a plant source or synthesized biologically, microbiologically, or chemically or by any other means.

The term"cultured dairy product"as used throughout the present specification and the appended claims shall be taken to mean a product comprising dairy ingredients which product has been treated by way of fermentation. A cultured dairy product may also comprise any number of non-dairy ingredients. Cultured dairy products include, but are not limited to, yoghurts, yoghurt-like products, buttermilk, cream cheese and sour cream and similar products. Flavourings, jams, herbs and such like may be added to a cultured dairy product after fermentation. When amounts in the present specification and the appended claims are stated in relation to a cultured dairy product, this shall be

taken to mean in relation to a cultured dairy product before any such addition unless otherwise stated.

The terms"exogenously added protein"and"exogenously added dietary fibres"as used throughout the present specification and the appended claims shall be taken to mean protein or dietary fibres which ends up in the fermented product by virtue of having been added to or being among or present in the starting materials or the intermediate products in a process for manufacturing said fermented product. When the fermented product is a cultured dairy product said terms shall be taken to mean protein or dietary fibres which are not part of a milk composition for use in a method according to the present invention as such, but which ends up in the cultured dairy product by virtue of having been added to or being among or present in the starting materials or the intermediate products in a process for manufacturing said cultured dairy product, in this case for instance the protein source for use in a method according to the present invention. This should not be taken to mean that for instance exogenously added protein may not be of dairy origin or that exogenously added protein may not be of a kind that is also part of a milk composition for use in a method according to the present invention.

The term"protein source"as used throughout the present specification and the appended claims shall be taken to mean a composition comprising protein. For the purpose of the present invention, a protein source may be the source of any number of proteins of any origin. The term itself shall provide no limitations as to the amount of protein present in the protein source and a protein source for use in a method according to the present invention may additionally comprise any number of non- protein components.

The term"hydration", in this case of proteins, as used throughout the present specification and the appended claims shall be taken to mean a process by which water molecules are forced around and in between different structural moieties of the individual protein molecules and in between polypeptide chains to such a degree, that it is substantially impossible to"squeeze"any additional water molecules into the close proximity of the protein molecule. The term"hydrated", in this case protein, as used throughout the present specification and the appended claims shall be taken to mean a protein that has undergone hydration or is present in a state of hydration.

The term"milk composition"as used throughout the present specification and the appended claims shall be taken to mean any composition containing milk ingredients.

The term"milk ingredients"as used throughout the present specification and the appended claims shall be taken to mean any ingredient which are present in or which may be derived from any kind of milk, such as for example cow's milk, ewe's milk, and goat's milk. The term"milk ingredients"may thus comprise milk carbohydrates, such as lactose, milk proteins, such as cassein, milk fats, such as palmitic acid and lecithin, salts and ions, such as calcium ions, and vitamins, such as vitamin A and D, and any derivatives thereof. The milk ingredients may be provided by cream, whole milk, partially skimmed milk, skimmed milk, reconstituted non-fat dairy milk, cream powder, whole milk powder, partially skimmed milk powder and skimmed milk powder and/or combinations thereof as described above.

The term"shelf-life"as used throughout the present specification and the appended claims shall be taken to mean the time-period from the finalisation of a product and until this product, when stored properly and under the conditions recommended by the manufacturer, becomes unacceptable to the consumer.

The term"proteinase"as used throughout the present specification and the appended claims shall be taken to mean an enzyme that degrades proteins and peptides by splitting internal and terminal peptide bonds to produce peptides and amino acids. The term"proteinase"may thus comprise any proteolytic enzyme, endopeptidase, exopeptidase, peptidase or peptidyl-peptide hydrolase.

The term"proteinase activity"as used throughout the present specification and the appended claims shall be taken to mean an enzymatic activity resulting in the degradation of proteins and peptides by the splitting of internal and terminal peptide bonds to produce peptides and amino acids. The term"proteinase activity"may thus comprise the activity shown by any proteolytic enzyme, endopeptidase, exopeptidase, peptidase or peptidyl-peptide hydrolase.

The term"hydrolysate"as used throughout the present specification and the appended claims shall be taken to mean any compound or mixture of compounds comprising a nitrogen-source which, when used in a method according to the present invention, reduce the release of bitter-tasting components associated with exogenously added protein. The term"hydrolysate"can thus mean, but is not limited to, a preparation of

hydrolised proteins/peptides such as e. g. hydrolyse casien or hydrolyse whey protein.

The term"bitter-tasting component"as used throughout the present specification and the appended claims shall be taken to mean any component with a bitter taste associated with the exogenously added protein. The term"bitter-tasting component" can thus mean, but is not limited to, isoflavons and saponins.

The terms"material to reduce bitter taste"and"material counteracting bitter taste"as used throughout the present specification and the appended claims shall be taken to mean material which is added prior, during or after the fermentation and which reduce the development of bitterness in the product or reduces bitterness already present.

The terms"material to reduce bitter taste"and"material counteracting bitter taste"can thus mean, but is not limited to, a hydrolysate or entities exhibiting proteinase activity.

In a further embodiment, the present invention provides a method for manufacturing a cultured dairy product containing exogenously added protein having an improved shelf-life which method comprises the steps of i) preparing a mixture of a protein source and a milk composition, ii) adding a fermentation culture to the mixture from step i), and iii) fermenting to obtain a cultured dairy product, and iv) cooling the fermented product to a temperature of about 5°C or below for intermediate storage and packing. wherein said fermentation culture added in step ii) at least partly comprises one or several entities exhibiting proteinase activity or excreting or liberating proteinases enabling the degradation of bitter tasting peptides and/or that said mixture in step i) at least partly comprises a hydrolysate resulting in reduced release of bitter tasting components associated with the exogenously added protein.

In a still further embodiment, the present invention provides a method for manufacturing a cultured dairy product containing exogenously added protein having an improved shelf-life which method comprises the steps of i) preparing a mixture of a protein source and a milk composition, ii) adding a fermentation culture to the mixture from step i), and

iii) fermenting to obtain a cultured dairy product, and iv) cooling the fermented product to a temperature of about 10°C and optionally adding flavourings and/or preservatives and finally cooling the fermented product to a temperature of about 5°C or below for intermediate storage and packing. wherein said fermentation culture added in step ii) at least partly comprises one or several entities exhibiting proteinase activity or excreting or liberating proteinases enabling the degradation of bitter tasting peptides and/or that said mixture in step i) at least partly comprises a hydrolysate resulting in reduced release of bitter tasting components associated with the exogenously added protein.

DETAILED DESCRIPTION OF THE INVENTION As described above, a fermentation of a mixture comprising high amounts of an exogenous protein source and a milk composition making use of traditional fermentation cultures results in a fermented product having a decreased shelf-life compared to traditional yoghurt products. Hence, in order to improve the shelf-life of a cultured dairy product according to the present invention, a fermentation culture for use in step ii) of a method according to the present invention at least partly comprises one or several entities exhibiting proteinase activity or excreting proteinases enabling the degradation of bitter tasting peptides and/or the mixture in step i) of a method according to the present invention, at least partly comprises a hydrolysate resulting in reduced release of bitter tasting components associated with the exogenously added protein. Preferably the entities exhibiting proteinase activity enabling the degradation of bitter tasting peptides comprise a bacterial strain capable of producing and secreting proteinases and belonging to the species Lactobacillus helveticus, which has preferably been inactivated by heating or freezing. Preferably the hydrolysate resulting in reduced release of bitter tasting components from the exogenous protein comprise a preparation of hydrolised proteins.

The protein source in step i) of a method according to the present invention is preferably hydrated prior to addition to the milk composition, and preferably by the use of shear forces and essentially as described in EP patent application no. 00610033.3, which is incorporated herein by reference. In nature, proteins are curled together and therefore vulnerable to both acidic and alkaline pH-values and interaction with ions such as calcium ions and will have a lower solubility in water based systems. To avoid this, the proteins in the protein source are hydrated as this causes the proteins to be

stabilized by forcing a lot of water molecules around and in between the protein molecules and side chains thereof. Without wishing to be bound by any specific theory it is believed that this makes them less vulnerable to acidic and alkaline pH-values and interactions with ions such as calcium ions.

The now hydrated protein source is mixed with a milk composition in step i) of a method according to the present invention, preferably under thorough stirring. If the hydrated protein source has a temperature above 60°C, it may be cooled to a temperature of about 50 to 60°C as rapidly as possible before being mixed with a milk composition in step i). This cooling is preferably achieved by use of a heat exchanger.

The use of a heat exchanger is practical and fast technology suitable for changing the temperature of a liquid. It normally works as a continuous in-line unit operation in a production process. More preferably, a heat exchanger is used to increase the temperature of the combined water and protein source, as described above, which is then pumped directly through a homogenizer at this temperature. These operations should be done as fast as unwanted reactions might occur during longer processing times and it is definitely also more economical.

A milk composition for use in a method according to the present invention is preferably prepared as follows: Water or milk or a combination thereof is admixed with a water soluble calcium ion binder, such as sodium or potassium citrate, sodium or potassium phosphate, or sodium or potassium hexametaphosphate, or a combination of these, as described previously, which is subsequently dissolved. Any powdery ingredients, such as skimmed milk powder, are added, and the mixture is stirred vigorously until the powder is dispersed and the mixture is homogenous. However, a milk composition for use in step i) of a method according to the present invention may be prepared in any way known in the art.

The hydrolysate resulting in reduced release of bitter tasting components associated with the exogenously added protein preferably comprises hydrolised proteins/peptides.

More preferably the hydrolysate resulting in reduced release of bitter tasting components associated with the exogenously added protein comprise hydrolyse whey protein or hydrolised casein. Even more preferably the hydrolysate resulting in reduced release of bitter tasting components associated with the exogenously added protein comprise hydrolised casein in the form of Casamino acids (Difco). The hydrolysate is preferably added in an amount of about 0,2 weight percent of the

resulting fermented product. The hydrolysate can be added at any point of a method according to the present invention but is preferably added to the mixture of step i).

The mixture obtained from step i) is heated in a vessel suitable for heating prior to step ii) of a method according to the present invention to achieve pasteurization and coagulation of proteolytic enzymes in order to inactivate these. This heating may preferably be obtained in the temperature range from about 75°C to about 95°C, to ensure a complete pasteurization and inactivation of all possible proteolytic enzyme activity. The heating of the resultant mixture from step i) preferably takes place at about 85°C for about 30 minutes, more preferred at from about 90°C to about 92°C for about 10 minutes, and most preferred at about 85 to about 90°C for about 5 or about 7 minutes. The heating of the resultant mixture may begin while the milk composition and the hydrated protein source are being mixed, in which case the actual heating of the mixture at the aforementioned temperature should take place for a shorter time period. This heating is preferably achieved by use of a suitable heat exchanger.

The temperature of the mixture of step i), whether it has been heated as described in the previous paragraph or not, is then brought to a temperature suitable for fermentation either by heating or by cooling whichever is appropriate.

When the temperature, which for each individual case is deemed optimal for the fermentation, is reached, a fermentation culture is added and the fermentation starts.

The appropriately cooled or heated mixture as described in the previous paragraph, may be fermented by use of any method of fermentation, which is known in the art, but the culture used for fermentation in a method according to the present invention partly consists of any type of culture known in the art for use in fermentation to production of cultured dairy products or a combination of two or more such types, for instance the standard cultures used for fermentation to production of milk yoghurts, such as a combination of Lactobacillus bulgaricus and Streptococcus thermophilius, for instance YC180, YC 380, YC 460, YC 470, YT 10 and/or CH1 (all from Chr. Hansen, Horsholm, DK) and/or MYE 98, MYO 87, BY 118TG, B3 (all from Rhodia Texel, Manchester, GB).

This part of the fermentation culture may for instance be added to the resulting mixture from step i) in an amount of from about 0.01 weight percent to about 0.05 weight percent of the cultured dairy product. As described in EP patent application no.

0061033.3 the strains from Rhodia Texel ferments a mixture according to a particular preferred embodiment of the present invention at slower rate than the strains from Chr.

Hansen. Among the strains from Chr. Hansen, CH1 is well suited for the purpose of

the present invention. Since, as described above, a fermentation making use of traditional fermentation cultures results in a fermented product having a decreased shelf-life compared to traditional yoghurt products, a fermentation culture for use in step ii) of a method according to the present invention at least partly comprises one or several entities exhibiting proteinase activity or excreting proteinases enabling the degradation of bitter tasting peptides.

The proteinases provided by the entities exhibiting proteinase activity or excreting proteinases enabling the degradation of bitter tasting peptides are preferably purified or immobilized bacterial proteinases. More preferably the entities exhibiting proteinase activity or excreting proteinases enabling the degradation of bitter tasting peptides comprise a bacterial strain, this being either a natural isolate or genetically modified, capable of producing and secreting proteinases. Furthermore, said bacterial strain has preferably been inactivated or attenuated by freezing, heating, treating with lysozyme, treating with an organic solvent or by neutralization or in any other possible way as described by F. Rallu et al. 1996. Acceleration of cheese ripening. Antonie van Leeuwenhoek 70: 271-297, which is incorporated herein by reference. Even more preferably the entities exhibiting proteinase activity or excreting proteinases enabling the degradation of bitter tasting peptides comprise a starter or non-starter lactic acid bacterial strain capable of producing and secreting proteinases and belonging to genera Lactobacillus, Lactococcus or Streptococcus. Most preferably the entities exhibiting proteinase activity or excreting proteinases enabling the degradation of bitter tasting peptides comprise a bacterial strain capable of producing and secreting proteinases and belonging to the species Lactobacillus helveticus, which has preferably been inactivated by heating or freezing. Particularly preferred entities exhibiting proteinase activity or excreting proteinases enabling the degradation of bitter tasting peptides are heat inactivated Lactobacillus helveticus, as contained in the product marketed under the name of Enzobact and manufactured by Medipharm. Heat inactivated Lactobacillus helveticus in the form of Enzobact may for instance be added as part of a fermentation culture for use in a method according to the present invention in an amount of for instance about 0.02 to about 0.2 weight percent of the cultured dairy product, but is preferably added as part of a fermentation culture for use in a method according to the present invention in an amount of for instance about 0.05 to about 0.1 weight percent of the cultured dairy product. For example, heat inactivated Lactobacillus helveticus in the form of Enzobact may for instance be added as part of a fermentation culture for use in a method according to the present invention in an amount of about 0.0435 weight percent of the cultured dairy product. The addition of one or several entities exhibiting proteinase activity or excreting proteinases enabling

the degradation of bitter tasting peptides will provide for a cultured dairy product with a prolonged shelf-life.

The fermentation culture described above is preferably added under stirring. When the fermentation culture has been added, the fermentation begins. For a normal fermentation of milk the fermentation temperature should be around 42-44°C.

Preliminary trials showed that fermenting according to the present invention at a temperature of about 44°C provided for a faster fermentation and a larger production of lactic acid compared with other temperatures. It seems like fermenting at about 44°C is advantageous when fermenting a fermentation mixture with a high amount of protein according to the present invention, most likely since this temperature is more optimal for Streptococcus thermophilius. Accordingly, the fermentation according to the present invention preferably takes place at a temperature of about 44°C.

The fermentation mixture of step ii) should contain as little oxygen as possible before fermentation since most lactic bacteria are only moderately aerotolerant anaerobes.

This is due to the fact that although most lactic acid bacteria are capable of growth in the presence of oxygen, some species intensively used in the dairy industry are sensitive to high amounts of oxygen due to an accompanying production of hydrogen peroxide. The oxygen content of the fermentation mixture of step ii) is kept as low as possible by restricting the amount of air, which is allowed to be absorbed into the mixture during the first steps of the process. Furthermore heating the resulting mixture of step i) prior to the fermentation as described previously reduces the viscosity, which will make much of the dissolved oxygen disappear from the mixture. If there is still too much oxygen trapped in the mixture, vacuum is applied for a short period of time just before the mixture is brought to the fermentation temperature or just before the addition of the fermentation culture.

The temperature of the fermentation mixture is kept constant preferably by use of a thermostat and fermentation is stopped, preferably after no more than 6 hours although longer fermentation times, such as 10 or 20 hours, can be envisioned. A shorter fermentation time (preferably about 5.5 hours) is nevertheless more preferred as a longer fermentation time, among others, will increase the possibility of invasion of other types of bacteria. Furthermore, a longer fermentation period may increase the proteolytic effect of part of the bacteria present in the fermentation culture which will result in an unfavourable ratio between enzymes responsible for degrading the protein

chains of the exogenously added proteins into bitter tasting peptides and enzymes responsible for degrading these bitter tasting peptides into their component amino- acids. In addition, a longer fermentation period may increase the release of bitter tasting components, such as isoflavons and saponins, associated with the exogenous protein.

In an alternative embodiment of the present invention the fermentation takes place at about 28-30°C and the fermentation time is in the order of 20-24 hours.

A fermented product is in some instances found to contain white spots due to precipitation of casein and to have a grainy texture. To avoid such problems, the product may be led through a homogenizer at a pressure up to 100 bar, preferably from about 30 to about 40 bar, which results in a product with a smoother and less viscous texture and without any particles which could be felt in the mouth, especially when agar-agar, pectin and/or gum arabic is present in the cultured dairy product as described previously. Alternative types of suitable process technology can for instance be filter systems such as a steel mesh with small apertures, which the fermented product can be passed through.

After the fermentation the fermentation vessel is optionally cooled immediately, for instance by the use of cold water, until the fermented product reaches a temperature of about 5°C or below for storage.

Jams, fruit concentrates, flavouring, preservatives and/or other additives may be added to the cultured dairy product at any time after the fermentation or, if need be, before or during the fermentation. If such an addition is to take place, the fermented product in step iii) is preferably cooled to a temperature of about 10 to 12°C at which temperature the addition of the jams and/or fruit concentrates takes place.

A cultured dairy product according to the present invention preferably contains exogenously added protein in an amount of from about 5 weight percent to about 25 weight percent, such as from about 6 weight percent to about 15 weight percent, for example from about 7 weight percent to about 12 weight percent of the cultured dairy product.

Examples of exogenously added proteins in the context of the present invention are vegetable proteins, such as soy proteins, and animal proteins. The exogenously added

protein is preferably non-dairy protein and more preferably soy protein. The soy protein is preferably provided by isolated soy protein, soy protein concentrate, soy flour or the like or any combination thereof. The soy protein is preferably provided by an isolated soy protein product.

A cultured dairy product according to the present invention may additionally contain exogenously added dietary fibres. Examples of dietary fibres comprised in a cultured dairy product according to the present invention include fibres from apples, oats, and soybeans. The dietary fibres are preferably provided by soybean fibres, and more preferably by soy cotyledon fibres. The dietary fibres is preferably present in an amount of from 0 to about 40 weight percent based on the weight of the exogenously added protein.

A preferred cultured dairy product according to the present invention has a content of exogenously added protein of about 9 weight percent and a content of exogenously added dietary fibres of about 2.25 weight percent.

A cultured dairy product according to the present invention may additionally contain a calcium ion binder such as for example a salt selected from the group comprising sodium or potassium citrate, sodium or potassium phosphate, or sodium or potassium hexametaphosphate, or a combination of these, whereof sodium hexametaphosphate is most preferred. A calcium ion binder is preferably present in a cultured dairy product according to the present invention in an amount of from 0 to about 0.2 weight percent.

When the calcium ion binder is sodium or potassium hexametaphosphate, it is preferably present in an amount of from 0 to about 0. 15 weight percent, and more preferably from about 0.08 to about 0.10 weight percent of the cultured dairy product.

A cultured dairy product according to the present invention may furthermore contain a fat source such as lecithin for example soy lecithin. When soy lecithin is present in a cultured dairy product according to the present invention it is preferably present in an amount of from about 0.1 to about 1 weight percent, such as about 0.6 weight percent of the cultured dairy product. A cultured dairy product according to the present invention may furthermore contain a thickening agent such as agar-agar, gum arabic or pectin, or any combination thereof, preferably a combination of agar-agar and pectin. The pectin is preferably a mixture of amidated low ester and high ester pectin, such as GENU pectin type LM-107 AS-YA. When gum arabic and/or pectin is present in a cultured dairy product according to the present invention, each should be

present in an amount of from about 0.08 to about 1.0 weight percent, preferably about 0.25 weight percent of the cultured dairy product, and when agar-agar is present, it should be present in an amount of from about 0.005 to about 0.15 weight percent, preferably about 0.06 weight percent.

A cultured dairy product according to the present invention may furthermore contain soy milk. Preferably, a cultured dairy product according to the present invention may contain up to 35 weight percent soy milk.

A cultured dairy product according to the present invention or prepared by a method according to the present invention also has an increased acidity contributing to a fresh taste. The acidity of a cultured dairy product prepared by a method according to the present invention may be increased with up to 40 to 50% compared to the acidity of cultured dairy products, which are known in the art.

Phytoestrogen compounds according to the present invention are defined as naturally occurring plant substances, which are either structurally or functionally similar to 17El- estradiol or generate estrogenic effects. Phytoestrogens consist of a number of classes including isoflavones, coumestans, lignans and resorcylic acid lactones.

Examples of isoflavones according to the present invention are genistein, daidzein, equol, glycitein, biochanin A, formononetin, and O-desmethylangolesin. The phytoestrogen compounds of a fermented product according to the present invention are preferably isoflavones, more preferably genistein, daidzein, glycitein and/or equol, yet more preferably genistein and/or daidzein and even more preferably genistein.

Genistein and daidzein are found almost uniquely in soybeans. A preferred fermented product according to the present invention may accordingly comprise a single isoflavone, such as genistein, daidzein, glycitein or equol, or it may comprise at least one isoflavone selected from the group comprising at least genistein, daidzein, glycitein and equol. Furthermore, a preferred fermented product according to the present invention may accordingly comprise isoflavones being naturally part of the soy protein source employed.

Phospholipid sources according to the present invention are defined as fat substances comprising at least about 5% phosphatidyl choline. However, phospholipid sources according to the present invention may contain as much as 100% phosphatidyl choline. Furthermore phospholipid sources according to the present invention will preferably comprise polyunsaturated fatty acids and monounsaturated fatty acids and

optionally also saturated fatty acids. The phospholipid sources will preferably comprise polyunsaturated fatty acids and monounsaturated fatty acids and optionally also saturated fatty acids. The amount of polyunsaturated fatty acids and monounsaturated fatty acids, including the essential fatty acids, may range from 35 to 50, preferably 38 to 44, weight percent of the total amount of the phospholipid source source. The essential fatty acids are also called omega-6 and omega-3 fatty acids and include linolic acid and/or linolenic acid (l-linolenic acid). The amount of saturated fatty acids may be from 20 to 30 weight percent, preferably 22 to 26 weight percent, of the total amount of the phospholipid source. Lecithins and soy lecithins having a high content of C-linolenic acid are particularly preferred phospholipid sources according to the present invention.

A fermented product according to the present invention may be capable of preventing, treating, prophylactically treating and/or alleviating an arteriosclerotic condition by reducing the accumulation of cholesterol in the arterial wall and/or causing the dilation of blood vessels. This inhibitory effect may be mediated by the binding of naturally occurring isoflavones and/or soy peptides to an estrogen receptor or estrogen-like receptor present in the endothelium of an artery and/or through the action of the 1- alkyl-2-acetyl analog of phosphatidyl choline. The soy peptides are preferably provided by partial hydrolysis of soy protein.

Plasma cholesterol and triglyceride levels are usually increased in individuals treated for a cardiovascular disease and plasma triglyceride and lipoprotein levels are usually increased in individuals treated for type 2 diabetes and/or the metabolic syndrome.

These increased levels, unless reduced by treatment, are likely to promote atherosclerosis and/or coronary heart disease (CHD). Beta-2-adrenergic receptors are present on many different types of cells including i) cells of the arterial wall ii) cells of the airways and iii) fat cells. Beta-2-adrenergic receptors are involved in the regulation of triglyceride synthesis in fat cells and according to one presently preferred hypothesis, binding of soy peptides and/or a phytoestrogen compound such as e. g. a naturally occurring isoflavone to a beta-2-adrenergic receptor present on a fat cell or in an arterial wall is effective in reducing e. g. the synthesis of triglycerides in fat cells and/or the release of triglycerides into the blood stream and/or reducing the influx of cholesterol and/or triglycerides into the arterial wall. The soy peptides are e. g. obtainable by partial hydrolysis of soy protein.

Several mechanisms for the association between elevated homocysteine and vascular diseases have been proposed, including effect on endothelial function, vascular smooth muscle cells, LDL-C action, coagulation pathways, and oxidative status. In addition to the effect of homocysteine on CVD, some data indicate elevated homocysteine levels are related to poor cognitive function and cancer. Three nutrients- -folate, vitamin B6 and vitamin B12--have been shown to influence homocysteine levels. However, an additional compound, betaine, is required for the conversion of homocysteine to methionine. Betaine, which acts as a methyl donor, is synthesized endogenously from choline through the action of choline dehydrogenase and betaine aldehyde dehydrogenase. In children with cystathione beta-synthase deficiency, which results in the accumulation of homocysteine due to the inability to convert homocysteine to cystathionine, betaine administration has been shown to lower homocysteine levels significantly. In other cases, homocysteine lowering by folate requires the addition of choline or betaine. Thus according to a preferred hypothesis soy phospholipids, e. g. phosphatidyl choline may act as a direct controlling factor in lowering homocysteine levels and thereby reduce and/or eliminate one of the risk factors associated with cardiovascular diseases.

As an integral part of cell membranes, phospholipids are directly involved in cell signaling. As a constituent of cell membranes, phospholipids consist of a variety of molecular species, including ester-, ether-, and vinyl-linked forms. In response to cell stimuli, phospholipids are broken down by specific phospholipases, resulting in a number of hydrolysis products that have the potential to act as second messengers and thus markedly influence cellular processes, including cholesterol metabolism.

According to a preferred hypothesis, a fermented product according to the present invention will reduce and/or eliminate one or more of the risk factors for cardiovascular diseases. Accordingly, a fermented product according to the present invention may be effective in preventing, treating, prophylactically treating and/or alleviating conditions such as e. g. hypercholesterolemia, hypertriglyceridemia, hypertension and hyperglycemia. A fermented product according to the present invention may also be capable of reducing, preventing and/or eliminating fatty streak formation and/or fibrous plaque development and/or effective in mediating a regression of one or both of said arteriosclerotic conditions.

A fermented product according to the present invention may be effective in preventing and/or treating type 2 diabetes and/or the metabolic syndrome and/or reducing and/or

eliminating one or more of the risk factors for cardiovascular diseases associated with diabetes and/or the metabolic syndrome. Accordingly, a fermented product according to the present invention may be effective in preventing, treating, prophylactically treating and/or alleviating conditions such as e. g. increased serum levels of glucose, hypercholesterolemia, hypertriglyceridemia, hypertension, and hyperinsulinemia in diabetic individuals. A fermented product according to the present invention may also be capable of reducing, preventing and/or eliminating fatty streak formation and/or fibrous plaque development and/or effective in mediating a regression of one or both of said arteriosclerotic conditions in diabetic individuals.

According to a preferred hypothesis, a fermented product according to the present invention will be effective in lowering serum levels of total cholesterol and/or LDL- cholesterol and/or triglycerides and/or in increasing the serum HDL/LDL-cholesterol ratio and/or increasing serum levels of high-density lipoproteins (HDL) and/or in generating a decrease in serum levels of low-density lipoproteins (LDL). It is desirable to achieve an elevated serum HDULDL-cholesterol ratio since this may result in an increased reverse cholesterol transport and a subsequent excretion.

Also, it is believed that a fermented product according to the present invention will affect ApoB lipoprotein metabolism including the metabolism of a recently discovered class of ApoB comprising lipoprotein particles called small, dense LDL particles. The LDL class of lipoproteins is in fact composed of several components with distinct properties. The basis for this heterogeneity and the consequences for disease are at present not thoroughly understood. An increased level of small, dense LDL particles is one of the most common dyslipoproteinemias associated with coronary artery disease, and serum levels of ApoB are often disproportionately elevated compared with LDL- cholesterol in dyslipoproteinemic patients.

Heterogeneity within lipoprotein classes may be the result of a differing lipid content, a different apoprotein composition, an altered protein conformation or an as yet unidentified structural variation. Subjects with a preponderance of small, dense LDL have an increased risk of suffering a myocardial infarction independent of the total concentration of serum LDL. Accordingly, a fermented product according to the present invention may be effective in lowering elevated levels of small, dense LDL.

Hypertriglyceridemia in non-diabetic and diabetic subjects alike is associated with an increase in the clotting activities of thrombogenic factors such as e. g. factor Vll and/or

factor X and/or factor XI I and an increase in the level of the inhibitor of tissue plasminogen activator, PAI-1. The increased inhibitor concentration results in a decreased level of plasminogen synthesis and thus a decreased level of plasminogen stimulated clot lysis. These changes in clotting activities no doubt contribute to the procoagulant state, which is also observed in diabetes. Accordingly, the present invention provides a fermented product, which may be effective in normalizing levels of homocystein and/or the clotting activities of at least one thrombogenic factor such as e. g. factor VII and/or factor X and/or factor Xll by e. g. decreasing the increased activity thereof, which is also observed in a subject diagnosed as having type 2 diabetes or diagnosed as having an impaired glucose tolerance or a decreased insulin sensitivity.

Also, a fermented product according to the present invention may be effective in promoting a decrease in the level of the inhibitor of tissue plasminogen activator, PAI- 1, which in turn leads to an increased plasminogen stimulated clot lysis. A fermented product according to the present invention may also be effective in reducing an increased platelet aggregatability and/or mediating directly or indirectly a reduction in the increased level of lipoprotein (a) associated with a procoagulant state in an arteriosclerotic condition and/or a diabetic condition.

Accordingly, in one embodiment the present invention provides a fermented product effective in reducing and/or eliminating risk factors for coronary heart disease (CHD) in obese subjects and in obese subjects suffering from a diabetic condition and/or the metabolic syndrome. Consequently, a fermented product according to the present invention may be capable of preventing, treating, prophylactically treating, alleviating and/or eliminating hyperinsulinemia and/or hypertriglyceridemia and/or hypercholesterolemia and/or hyperglycemia and/or hypertension and/or effective in mediating an increase in the low serum levels of HDL-cholesterol and/or effective in mediating an increased serum HDULDL-cholesterol ratio.

A fermented product according to the present invention may also be effective in treating dyslipidemia such as e. g. hypertriglyceridemia and/or hypercholesterolemia in connection with increased serum levels of VLDL, decreased and altered serum levels of HDL and increased serum levels of small dense LDL, and hypertension, all of which are risk factors for atherosclerosis. Accordingly, in one embodiment, a fermented product according to the present invention may be capable of effectively lowering and/or eliminating increased serum levels of VLDL, and/or effectively increasing decreased serum levels of HDL, and/or effectively lowering serum LDL levels including serum levels of small dense LDL. A fermented product according to the present

invention may be capable of preventing, treating, prophylactically treating and/or alleviating hypertension.

A fermented product according to the present invention may also be effective in suppressing any effect that would otherwise generate an increased turnover of arterial smooth muscle cells, i. e. an enhanced arterial smooth muscle cell proliferation, and/or lead to an increased cholesterol ester accumulation in the arterial wall.

In hypercholesterolemia characterized by increased levels of intracellular cholesterol resulting from e. g. increased delivery of LDL-cholesterol via the LDL receptor, a fermented product according to the present invention may be effective in reducing the increased activity of the LDL receptor. It is also possible that insulin and other growth factors have the potential to promote the accumulation of cholesterol intracellularly.

This may in fact well occur in a diabetic subject and more generally under conditions when cells are stimulated, but cannot proliferate normally. Accordingly, a fermented product according to the present invention may also be capable of treating, alleviating and/or eliminating any decrease, including any insulin mediated decrease, in the HDL receptor-mediated cholesterol efflux. Accordingly, a fermented product according to the present invention may be capable of reducing and/or eliminating any enhanced retention of intracellular cholesterol caused by a decreasing HDL receptor-mediated cholesterol efflux.

A fermented product according to the present invention may be effective in reducing insulin resistance by stimulating cells or receptors located thereon that are normally stimulated by insulin, but less sensitive to the hormone in a subject diagnosed with type 2 diabetes and/or the metabolic syndrome. A fermented product according to the present invention may also be effective in stimulating cells comprising a beta-2- adrenergic receptor or a receptor belonging to the class of beta-2-adrenergic receptors. The final phase of type 2 diabetes development is characterized by insulin secretory failure (ISF), and in one presently preferred hypothesis, this failure is at least preventable by a fermented product according to the present invention effective in stimulating insulin secretion.

Hypertriglyceridemia in diabetes has been associated with a variety of changes in circulating lipoproteins, and a fermented product according to the present invention may be capable of preventing, treating, alleviating and/or eliminating cardiovascular risk factors such as e. g. chylomicronemia, an increased level of VLDL, an increased

level of remnants (VLDL and chylomicrons), and LDL and HDL containing increased levels of triglycerides.

Lipoprotein fractions obtained from type 2 diabetic subjects tend to lose their typical sharp LDL peak and instead have a broad diffuse LDL band termed polydisperse LDL.

Dissection of polydisperse LDL reveals that diabetics have an increased serum level of intermediate-density-lipoprotein (IDL), an abnormal LDL peak, and an increase in the amount of small dense LDL. While small dense LDL particles have been associated with CHD in the general population, a similar association in diabetes remains to be established. Accordingly, a fermented product according to the present invention may be effective in promoting a decreased serum level of intermediate density lipoprotein (IDL), a normal, sharp LDL peak, and a decreased amount of small dense LDL.

Accordingly, diabetic dyslipidemia of type 2 diabetes is generally associated with abnormalities of apolipoprotein and lipoprotein particle distributions and results in increased plasma VLDL and remnant levels, an increase in the apoE concentration in VLDL and remnants, an increase in the amount of small dense LDL, and an altered HDL particle distribution.

According to one presently preferred hypothesis, a fermented product according to the present invention will alleviate abnormalities associated with apolipoprotein and lipoprotein particle distribution and promote a decreased plasma VLDL and remnant level, a decrease in the apoE concentration in VLDL and remnants, a decrease in the amount of small dense LDL, and a HDL particle distribution similar to that of a comparable non-diabetic, healthy individual.

Hyperinsulinemia is also considered a risk factor for coronary heart disease (CHD) in diabetic subjects due to the association of high insulin levels with increased incidence and mortality rates of CHD. A fermented product according to the present invention may be effective in lowering serum insulin levels in subjects diagnosed with type 2 diabetes. Diabetic patients having increased endogenous insulin levels, i. e. subjects diagnosed with type 2 diabetes, or having increased peripheral circulating insulin levels as a result of intermittent injections of large amounts of exogenous insulin are particularly prone to hyperinsulinemia.

Hyperinsulinemia in both normal persons, persons with the metabolic syndrome and those with type 2 diabetes appears to be related to obesity. Insulin levels are very

often increased in both the fasted state and after intake of a diet rich in carbohydrates in obese individuals, irrespective of whether they suffer from a diabetic condition or not. Furthermore, hyperinsulinemia appears to be directly correlated to the degree of obesity. Accordingly, hyperinsulinemia is one of the many risk factors for CHD associated with obesity, and insulin may modulate many other obesity-related risk factors. Accordingly, a fermented product according to the present invention may be effective in lowering insulin levels in obese subjects with diabetes or the metabolic syndrome.

In obese subjects diagnosed as diabetic, LDL particle size is independently correlated with factors such as e. g. serum triglyceride and serum insulin levels. Consequently, it is possible that the extent of adiposity and concomitant insulin resistance in hyperinsulinemic individuals is associated with the occurrence small dense LDL, independently of hypertriglyceridemia, which is another diabetic condition also putatively associated with small dense LDL formation. Accordingly, both insulin resistance and hyperinsulinemia appear to play a central role in the pathogenesis of atherosclerosis in diabetes. A fermented product according to the present invention may be effective in treating and/or alleviating insulin resistance and/or hyperinsulinemia.

It is very possible that type 2 diabetes is also associated with insulin resistance and hyperinsulinemia independently of an increase in abdominal lipids. Hyperinsulinemia in turn is associated with dyslipidemia, i. e. increased VLDL, decreased and altered HDL and increased small dense LDL, and with hypertension, all of which are risk factors for atherosclerosis. This array of abnormalities and disorders, or a part of thereof, is generally termed the insulin resistance syndrome, or syndrome X, or metabolic syndrome.

In one embodiment, a fermented product according to the present invention may be capable of effectively decreasing and/or eliminating increased serum levels of VLDL and/or LDL, and/or increasing decreased serum levels of HDL, and of decreasing and/or eliminating serum LDL levels including serum levels of small, dense LDL. A fermented product according to the present invention may also be capable of reducing an elevated level of small, dense LDL particles and/or reducing an elevated ratio of LDL-apoB to LDL-cholesterol and/or preventing, treating or alleviating hypertension.

Even though there is no internationally agreed definition for the metabolic syndrome, the term as used herein shall be understood to relate to the occurrence in a subject of at least two of the following: i) impaired glucose tolerance, ii) elevated blood pressure, iii) hypertriglyceridemia and low HDL-cholesterol, iv) insulin resistance, and v) obesity.

The occurrence of a condition characterized by one or more of impaired glucose tolerance, elevated blood pressure, hypertriglyceridemia and low HDL-cholesterol, insulin resistance, and obesity will depend on variables such as sex, age, body weight, physical condition and the like, and general WHO guidelines will generally be adhered to when evaluating the occurrence of any one of the above-mentioned conditions.

Hyperinsulinemia in itself may well be capable of affecting the arterial wall either directly or indirectly by promoting or facilitating the promotion of changes similar to those leading to severe atherogenesis. Insulin may well promote both arterial smooth muscle cell proliferation and cholesterol ester accumulation in the arterial wall. A fermented product according to the present invention may in one embodiment be effective in preventing, treating, alleviating and/or eliminating fatty streak formation, fibrous plaque development, complicated lesion formation, thrombosis, platelet aggregation and/or myocardial infarction.

Since insulin can be expected to be capable, either in combination with other compounds such as additional growth factors, or on its own, of increasing the levels of intracellular cholesterol, by e. g. increasing a delivery of LDL-cholesterol via the LDL receptor, and concomitantly therewith increase an endogenous biosynthesis of cholesterol that makes yet more cholesterol available for new membrane synthesis in the cell proliferation process, it is an object of the present invention to counteract any increased activity including any insulin stimulated increased activity of the LDL receptor.

Modifications of lipoproteins constitute another risk factor for cardiovascular disease, including cardiovascular disease in diabetes. The modification characterized by protein glycosylation is associated with e. g. arteriosclerosis and diabetes, and glycosylated lipoproteins such as e. g. LDL, IDL, VLDL and HDL can be expected to be functionally abnormal. Accordingly, the accumulation of glycosylated LDL in the plasma, including the plasma of a diabetic subject, can be perceived to enhance cholesterol ester accumulation. Also, glycosylation of HDL can be expected to impair the ability of HDL binding to the HDL receptor. This impaired binding is likely to reduce the level of intracellular cholesterol efflux. Accordingly, glycosylated HDL may well be another

factor potentially contributing to the accumulation of cholesterol in the arterial cell wall.

A fermented product according to the present invention may be effective in preventing, treating, alleviating, reducing and/or eliminating lipoprotein glycosylation in the serum of subjects, including diabetic subjects. In addition, a fermented product according to the present invention may also be effective in preventing lipoprotein modification caused e. g. by oxidation, chemical modification such as chemical cross-linking, or modifications caused by an alteration in the lipid composition of the lipoprotein, such as any increase or decrease in the content of triglycerides, cholesterol esters, free cholesterol, and apolipoproteins.

Glycosylated lipoproteins have been suggested to be the subject of further processing leading to the formation of hyperglycosylated compounds. Glycosylation and hyperglycosylation of proteins including lipoproteins in both plasma and the arterial wall can also be expected to be a risk factor for cardiovascular disease including arteriosclerosis, also in diabetic subjects. Accordingly, a fermented product according to the present invention may be capable of preventing, treating, alleviating, reducing and/or eliminating the accumulation of hyperglycosylated proteins in both serum and cells of the arterial wall. By doing so, the fermented product is acting to decrease the amount of LDL becoming"trapped"in the arterial wall due to the high degree of glycosylation of arterial wall proteins. A fermented product according to the present invention may also be effective in preventing and/or alleviating any change to the endothelial cell wall that increase LDL"trapping", and it may be effective in restoring the formation of cells with normal permeability and adhesion parameters.

Lipoprotein glycosylation, hyperglycosylation, oxidation and/or auto-oxidative glycosylation, are risk factors for cardiovascular disease such as arteriosclerosis, including arteriosclerosis in diabetes. Accordingly, a fermented product according to the present invention may be effective in preventing, treating, alleviating, eliminating and/or reducing any incidence of lipoprotein glycosylation, hyperglycosylation, oxidation and/or auto-oxidative glycosylation. According to one presently preferred hypothesis, the phytoestrogen compound of a fermented product according to the present invention is capable of counteracting such incidences. The phytoestrogen compound may also be capable of preventing, reducing and/or eliminating the formation of e. g. free radicals that are likely to be involved in such processes, and a fermented product according to the present invention may be effective in being, promoting, and/or facilitating the formation of an effective antioxidant defense system for counteracting glycosylation, hyperglycosylation, oxidation and/or auto-oxidative

glycosylation of serum proteins and proteins including lipoproteins of the arterial cell wall.

Since oxidative stress is a characteristic of diabetes and possibly a contributory factor to among others lipoprotein oxidation and/or glycosylation, and since no efficient antioxidant protection exists due to e. g. significantly decreased levels in diabetic subjects of antioxidants such as e. g. ascorbic acid, a fermented product according to the present invention may be effectively acting as an antioxidant in preventing lipoprotein oxidation and/or glycosylation.

Generally, a fermented product according to the present invention may be effectively acting as an antioxidant in preventing lipoprotein oxidation and/or glycosylation. By the term auto-oxidative glycosylation, or glycoxidation, is understood a reaction catalyzed e. g. by reducing sugars that leads to an oxidative modification and/or cross-linking of proteins. The rate of such a process can be expected to be increased in the presence of high glucose concentrations since the oxidizing potential is significantly increased under such circumstances. An increased production of free radicals and lipid peroxidation may also contribute to the formation of auto-oxidative glycosylated lipoproteins and this contribution may also be effectively prevented and/or eliminated by a fermented product according to the present invention.

According to another presently preferred hypothesis, the binding of phytoestrogen compounds such as e. g. isoflavones, optionally in combination with soy peptides e. g. obtainable by partial hydrolysis of soy protein, to a receptor in the arterial wall, such as e. g. the estrogen receptor, or an estrogen-like receptor, and optionally influenced by the level of specific phospholipids, such as phosphatidyl choline, is involved in or effective in controlling accumulation of lipoproteins and uptake of cholesterol and/or triglycerides in the arterial wall, possibly by regulating the permeability of said wall and/or the mechanism of lipoprotein and/or cholesterol and/or triglyceride transport across cellular membranes. Consequently, the binding of isoflavones such as e. g. genistein and/or daidzein to a receptor in the arterial wall may reduce the flux of lipoproteins into the arterial wall and/or prevent cholesterol and/or triglycerides from entering the arterial wall, or reduce and/or substantially eliminate the amount of cholesterol and/or triglycerides that enters the arterial wall. Receptor binding of isoflavones in the arterial wall is particularly effective in controlling, preventing and/or eliminating fatty streak formation and/or fibrous plaque development and/or effective in mediating a regression of one or both of said arteriosclerotic conditions.

According to a particularly preferred hypothesis, binding of an isoflavones such as e. g. genistein and/or daidzein to a receptor in the arterial wall, preferably an estrogen receptor or an estrogen-like receptor, results in an increased nitric oxide synthesis in the endothelial cells of the arterial wall. Nitric oxide is known to exert anti- arteriosclerotic effects including inhibition of platelet adhesion and aggregation, and inhibition of smooth muscle cell proliferation. Soy peptides obtainable by hydrolysis of soy protein may participate in the binding of isoflavones to an estrogen receptor or an estrogen-like receptor or the soy peptides may themselves bind to said receptor and exert an action leading to an increased nitric oxide synthesis. Furthermore, specific phospholipids, e. g. phosphatidyl choline, may in response to cell stimuli, be broken down by specific phospholipases, thereby resulting in a number of hydrolysis products having the potential to act as second messengers and thus markedly influence the effect of phytoestrogens.

In another presently preferred hypothesis, the establishment of an oxidative potential that promotes lipoprotein oxidation and/or lipoprotein auto-oxidative glycosylation occurs concomitantly with, and is very likely caused by, a decrease in cellular antioxidative defense systems. This hypothesis is supported by the fact that e. g. ascorbic acid concentrations are decreased in many diabetic individuals. Accordingly, a fermented product according to the present invention may be effective in acting as an antioxidant. This action reduces and/or eliminates LDL, VLDL, IDL and/or HDL susceptibility to oxidation. Concomitantly with a direct anti-oxidative effect, a fermented product according to the present invention may also lower the increased serum glucose levels and by doing so, a fermented product according to the present invention may be effective in reducing the oxidizing potential causing and/or contributing to oxidative stress.

Furthermore, a fermented product according to the present invention may also be effective in reducing an enhanced susceptibility to endothelial injury and/or for alleviating and/or restoring and/or improving an inefficient endothelial cell repair mechanism leading to endothelial dysfunction. One effect of such an action exerted by a fermented product according to the present invention is to direct macrophage development away from foam cell formation and to increase the potential of generating arterial smooth muscle cells.

The unique dyslipidemia associated with type 2 diabetes is a major risk factor for cardiovascular disease, and prevention, alleviation, reduction and/or elimination of dyslipidemia in diabetic subjects is a prime objective of administration of a fermented product according to the present invention to a diabetic individual. Another important objective of such an administration is the development in a diabetic subject of a gradually reduced insulin resistance and/or a gradually improved glucose tolerance.

Since increasing insulin resistance and impaired glucose tolerance are key elements in the progression of type 2 diabetes, the same factors most also be a natural focus of any preventive treatment.

In another presently preferred hypothesis, a fermented product according to the present invention will promote and/or mediate a reduction in arterial wall thickness and lead to a reduction in the amount of LDL entering the wall. It is believed that an increased thickness of the arterial wall is positively associated with an increased uptake of LDL particles that are likely to either aggregate or oxidize within the cells of the arterial wall.

Also, a fermented product according to the present invention may be capable of reducing, eliminating and/or preventing the formation of increased serum levels of lipoprotein (a) in a subject, including a diabetic subject. Lipoprotein (a) levels may primarily be genetically determined, and no current cardiovascular medications are thought effective in lowering serum levels of lipoprotein (a).

Obstructive pulmonary disease (OPD) including chronic obstructive pulmonary disease (COPD) as used herein is defined as a condition comprising subjects with airways limitations or obstructions or subjects with a mucus hypersecretory condition including chronic mucus hypersecretion, i. e. subjects with asthma including chronic asthma and subjects with bronchitis including chronic bronchitis. However, a clear distinction between e. g. bronchial asthma and chronic bronchitis can be difficult and sometimes impossible to make, and a sharp distinction between COPD and OPD is therefore not always possible.

Mucus hypersecretion and a limited or obstructed airflow are two major characteristics of COPD. According to one presently preferred theory, mucus hypersecretion is an initial mechanism that leads to recurrent respiratory infections, that in turn generates a destruction of the airways and promotes a development of pulmonary parenchyma and airflow obstruction. At least two separate conditions, i) mucus hypersecretion and ii)

dyspnea, are identifiable due to an obstructive or limited lung function. Chronic mucus hypersecretion and obstructive airflow are not necessarily related, since an individual may have a hypersecretory disorder only, or an obstructive disorder only, or both a hypersecretory and an obstructive disorder. Chronic mucus hypersecretion is associated with an impaired mucociliary clearance and may therefore predispose to lung cancer by causing a prolonged contact between potential carcinogens with the bronchial epithelium. Accordingly, a fermented product according to the present invention may be effective in treating and/or alleviating mucus hypersecretion and dyspnea in a subject.

Asthma as used herein is defined as a respiratory disease in which spasm and constriction of the bronchial passages and swelling of their mucous lining cause obstruction of breathing, often, but not exclusively, due to allergy. One mechanism for expiratory airflow limitation in asthma is a smooth muscle contraction leading to a narrowing of the airway lumen. Asthma is frequently divided clinically into extrinsic and intrinsic asthma, separating asthma triggered by environmental allergens from that in which atopy does not appear to play a major role. Consequently, a fermented product according to the present invention may be effective in preventing, treating and/or alleviating smooth muscle contraction.

In asthma the airways are occluded by tenacious plugs of exudate and mucus, and there occurs a fragility of airway surface epithelium, thickening of the reticular layer beneath the epithelial basal lamina, bronchial vessel congestion and edema. An increased inflammatory infiltrate comprising"activated"lymphocytes and eosinophils, and an enlargement of bronchial smooth muscle, particularly in medium-sized bronchi, is also observed. Asthma comprises at least extrinsic (atopic or allergic) and intrinsic (non-atopic) divisions, each of which present clinically in a variety of ways. A fermented product according to the present invention may be effective in preventing and/or alleviating the formation of tenacious plugs of exudate and mucus, effective in preventing, treating and/or alleviating a fragility of airway surface epithelium subsequently generated by mucus secretion, effective in preventing, reducing and/or eliminating any thickening of the reticular layer beneath the epithelial basal lamina, and effective in preventing, treating and/or alleviating bronchial vessel congestion and/or edema.

Asthma may in some cases be regarded as a chronic inflammatory disease. Since the term chronic asthmatic bronchitis has no clearly defined pathologic equivalent, patients

having a chronic productive cough normally associated with chronic bronchitis, as well as bronchospasms, at the same time as having an airflow obstruction, will be regarded as suffering from both chronic bronchitis as well as small airways disease (chronic obstructive bronchitis) and asthma, since the pathology presumably would be that of those conditions.

A fermented product according to the present invention may be effective in preventing, alleviating and/or curing inflammation of the airways, whether transient or chronic.

Airway inflammation is thought to be an important contributor to asthma, and airway inflammation may well be present even in the absence of severe symptoms of asthma.

In one particularly preferred aspect the present invention provides a treatment and/or alleviation of an inflammation of the airways by means of an anti-oxidative effect exerted by a fermented product according to the present invention. The anti-oxidative effect may in particular be exerted by naturally occurring isoflavones forming part of a fermented product according to the present invention.

A fermented product according to the present invention may be effective in increasing FEV1, as measured by forced expiratory volume in the first second of expiration, said effect being exerted by the binding of a component of the fermented product, particularly naturally occurring isoflavones, to beta-2 receptors or receptors belonging to the class of beta-2 receptors. Beta-2 receptors are present on many different types of cells including cells in airways and vessels. A fermented product according to the present invention may also be effective in generating a dilatation of the airways in a subject, preferably a subject suffering from a pulmonary disease.

The occurrence of bronchial inflammation in asthma is, according to one presently preferred hypothesis, thought to arise at least in part from an airway response to an antigen in an allergic subject. The response includes immediate pulmonary mast-cell activation and initiation of an inflammatory response that develops over hours and is important in the later and more persistent development of bronchial obstruction. A fermented product according to the present invention may be effective in treating, alleviating and/or eliminating several of the causes of airway obstruction that-alone or in combination-contributes to bronchial hyperresponsiveness, i. e. the fundamental defect in asthma. Importantly, airway inflammation is believed to be a crucial component for i) the chronicity of asthma, ii) the intensity of airways hyperresponsiveness, and iii) the absence of a complete therapeutic control, when bronchodilator therapy is used alone. Consequently, a fermented product according to

the present invention may be effective in controlling, reducing and/or eliminating edema, mucus secretion, and inflammation of the airways resulting at least in part from a response to an allergen.

Although the precise pathogenesis of asthma has yet to be discovered, allergic reactions and respiratory infections are particularly important. Both are frequent factors in asthma and exacerbations of asthma, and both not only trigger acute asthmatic symptoms but may also enhance the degree of airway hyperresponsiveness long after the initial stimulus has been removed. Of particular interest has been the airway's response to an inhaled antigen. Almost all subjects with allergic asthma experience immediate bronchospasm following inhalation of an antigen, i. e. acute airway obstruction, within 15 min of antigen exposure. In these subjects, antigen inhalation initiates not only immediate bronchocontraction, but also the reappearance of airway obstruction 4 to 6 hours later, a condition known as late asthmatic reaction or LAR.

The late asthmatic response has a number of features that are characteristic of chronic asthma such as e. g. less responsiveness to bronchodilator therapy than the isolated acute event, an increased airway responsiveness, and the development of bronchial inflammation. Two features of the LAR to antigen inhalation suggest a linkage to the pathogenesis of asthma: The presence of bronchial inflammation and the enhancement of bronchial responsiveness. Consequently, a fermented product according to the present invention may be capable of preventing both immediate bronchocontraction as well as a late asthmatic reaction.

Asthmatic reactions following inhalation of an antigen include an immediate release from pulmonary mast cells of preformed mediators and a generation of a variety of factors needed to initiate an acute allergic airway reaction. Because the airways of patients with asthma are hyperresponsive, the immediate bronchial reaction to mast cell bronchospastic mediators is accentuated beyond the pharmacological properties of these substances. With cellular activation by antigen and membrane-bound IgE interaction, the mast cell initiates a generation of leukotrienes and prostaglandins. The leukotrienes, C4, D4, E4, along with histamine, are undoubtedly involved in the acute bronchospastic response because of their airway smooth muscle contractile properties. The generation and release by mast cells of chemotaxic factors is important for the recruitment of inflammatory cells to the airway and for the subsequent development of the late asthmatic response. Accordingly, a fermented product according to the present invention may be capable of effectively reducing or

eliminating mast cell mediated secretion of mediators such as e. g. heparin, histamine and sulphidopeptide leukotrienes C4, D4, and E4.

Associated with the development of the LAR is a recruitment of inflammatory cells to the airway, including neutrophils, macrophages, lymphocytes, eosineophils, monocytes, and basophils. With their entry into the airways, and presumable cellular activation, airway obstruction reappears. It is thought that components of airway obstruction in LAR include bronchospasm, edema, and inflammation. An additional consequence of the LAR is an increase in airway hyperresponsiveness; thus, the asthmatic process is further perpetuated and positively reinforced. Consequently, a fermented product according to the present invention may be capable of effectively controlling in a late asthmatic response the symptoms of bronchospasm, edema, and inflammation, and in addition also effectively controlling such as reducing and/or eliminating any increase in airway hyperresponsiveness.

Furthermore, mast cells may according to another presently preferred hypothesis produce various cytokines, interleukin 3 (IL-3), interleukin 5 (IL-5), and granulocyte/macrophage colony-stimulating factor (GM-CSF), which can perpetuate the allergic reaction by further priming inflammatory cells. Consequently, a fermented product according to the present invention may be capable of effectively controlling i. e. reducing and/or eliminating the production of various cytokines, interleukins such as e. g. interleukin 3 (IL-3) and interleukin 5 (IL-5), and granulocyte/macrophage colony- stimulating factor (GM-CSF), and reduce any further priming of inflammatory cells during an early and/or late asthmatic response.

A class of cells termed neutrophils can be found in lavage fluid from an asthmatic subject, but the precise role of neotrophils in the generation of a late allergic reaction has not yet been established. The neutrophil cell has a potential for generating inflammation by releasing e. g. lysosomal enzymes, oxygen metabolites, leukotriene B4 and by synthesizing histamine-releasing factor (HRF). HRF can amplify the allergic reaction by causing mediator release from a class of cells termed basophils that also appear during a late allergic reaction. Consequently, a fermented product according to the present invention may be capable of effectively controlling i. e. reducing and/or eliminating a neutrophil production of e. g. lysosomal enzymes, oxygen metabolites, leukotriene B4 and histamine-releasing factor (HRF).

Evidence also exists for an implication of the group of cells termed eosinophils in an asthmatic response. Circulation of eosinophils leads to an increased severity of airway obstruction. Eosinophil granular associated proteins, including major basic protein (MBP), eosinophil cationic protein, eosinophil-derived neurotoxin, and eosinophil peroxidase are known to have profound effects on airway and cell function. MBP in particular has a number of unique properties accentuating the asthmatic response.

MBP can directly injure airway epithelium, promote bronchial responsiveness, and mediate smooth muscle contraction. MBP further activates the release of mediators from mast cells and basophils. Eosinophils may also be involved in initiating tissue damage associated with various allergic diseases, such as e. g. the epithelial desquamation observed in asthmatics. This tissue damage has been suggested to be mediated in part via the release of cytotoxic mediators such as major basic protein (MBP), eosinophil cationic protein (ECP), and eosinophil peroxidase (EPO).

Eosinophil activation results in the release of a number of important mediators, including leukotriene C4, which can contract airway smooth muscle, and platelet- activating factor (PAF). The release process of PAF has not been fully defined, but if secreted, this lipid mediator could contract airway smooth muscle as well as increase bronchial responsiveness. Furthermore, PAF is a potent eosinophil chemoattractant and a functional primer. Accordingly, eosinophils possess properties directly and indirectly causing airway obstruction and promoting bronchial hyperresponsiveness. Consequently, a fermented product according to the present invention may be capable of effectively controlling i. e. reducing and/or eliminating any increase in the formation of eosinophils during an asthmatic response. A fermented product according to the present invention may further be effective in controlling the production of eosinophil granular associated proteins including major basic protein (MBP), eosinophil cationic protein, eosinophil-derived neurotoxin, and eosinophil peroxidase. In an even further embodiment, a fermented product according to the present invention may be effective in controlling i. e. reducing and/or eliminating the release of mediators from mast cells, neutrophils, basophils and eosinophils, in particular the release of mediators such as e. g. leukotriene C4 and platelet-activating factor (PAF), IL-3, GM-CSF and IL-5.

To generate airway inflammation after eosinophil recruitment, a number of events need to occur such as e. g. eosinophil migration to the lung and eosinophil activation.

The last event is likely to involve eosinophil adhesion to endothelium and, eventually, airway epithelium. Accordingly, a fermented product according to the present invention

may be effective in preventing eosinophil participation in the bronchial responsiveness process by inhibiting eosinophil adhesion to endothelium and epithelium.

Mast cells may also release compounds such as heparin and related proteoglycans, but the release of such mediators have so far not received much attention from allergy researchers. These highly anionic molecules are normally only associated with the binding histamine within mast cell granules. These molecules may act as natural antiinflammatory molecules and, thus, have a far greater role in the pathogenesis of allergic diseases. Accordingly, a fermented product according to the present invention may be effective in promoting the release of potentially antiinflammatory molecules such as e. g. heparin and related proteoglycans. Also, it has been reported that another cationic protein, platelet factor 4 (PF4), is a chemotaxic agent for human eosinophils and is a molecule well recognized for its ability to bind heparin. It is therefore plausible that endogenous heparin could be released to limit both the extent of eosinophil recruitment into sites of allergic inflammation as well as the extent of tissue damage induced by cationic proteins. Lymphocytes are also likely to be involved in the pathogenesis of allergic asthma. Recent studies have suggested that heparin acts as an immunomodulator inhibiting lymphocyte activation and trafficking and, like glucocorticosteroids, can also inhibit delayed hypersensitivity responses.

Further evidence of asthma being a chronic inflammatory disease is provided by the observation that an exposure to an allergen that results in tissue damage is likely to lead to a repair of the damage. Evidence of this repair process can most likely be seen in the asthmatic lung, where a thickened basement membrane is believed to be related to subepithelial fibrosis, the presence of myofibroblasts, and collagen deposition. Also, asthma is further characterized by a thickened smooth muscle layer.

The above-mentioned changes appear very early in the disease and are not constricted to patients with chronic asthma. Furthermore, even following chronic treatment with inhaled glucocorticosteroids for periods of up 10 years, the thickness of the basement membrane is not reduced, although such therapy reduces the number of inflammatory cells present in the biopsies and the extent of the epithelial damage.

Such clinical observations suggest that once these anatomic changes have appeared they may not be readily reversible, even with the most aggressive therapy currently available. Thus, it is plausible that once established, such anatomic changes may underlie the irreversible component of the disease, and by altering the geometry of the airway wall, these changes may contribute to the persistent airways hyperresponsiveness that does not respond to treatment. Accordingly, a fermented

product according to the present invention may, in one particularly preferred embodiment, be capable of effectively preventing and/or alleviating the formation of a thickened basement membrane or a smooth muscle layer, subepithelial fibrosis, the presence of myofibroblasts, and a deposition of collagen.

Bronchitis as used herein is defined as an acute or chronic inflammation of any part of the bronchi and bronchial tubes. The bronchi are large delicate tubes in the lungs that are attached to the trachea and carry air to smaller tubes in the lungs. In bronchitis, including chronic bronchitis, there is mucous hypersecretion, an enlargement of tracheobronchial submucosal glands, and a disproportionate increase of mucous acini.

Acute bronchitis is often characterized by fever, chest pain, severe coughing, and secretion of mucous material coughed up from the respiratory tract. Acute bronchitis affects the branches of the bronchi and may develop into bronchial or lobular pneumonia. Chronic bronchitis may result from repeated attacks of acute bronchitis.

Consequently, a fermented product according to the present invention may be effective in controlling mucous hypersecretion, preventing, treating and/or alleviating an enlargement of tracheobronchial submucosal glands, and reduce and/or eliminate a disproportionate increase of mucous acini. The pathologic equivalent to chronic bronchitis is a non-specific series of changes in the bronchial wall generally characterized by an increase in the size and number of mucous glands and an increased number of goblet cells in the epithelium. When progressing into a chronic condition, bronchitis is a serious and incurable disorder. Consequently, a fermented product according to the present invention may be effective in controlling a series of changes in the bronchial wall generally characterized by an increase in the size and number of mucous glands and an increase in the number of goblet cells. A fermented product according to the present invention may also be capable of reducing and/or eliminating any mucos production including an increased mucos production.

Bronchial infections usually remain confined to the mucosa, and some resolve spontaneously without the need for treatment. Chronic bronchitis affects both the large and small airways. In the large airways, hypertrophy and hyperplasia of glandular structures and goblet cell metaplasia are prominent features of the condition. In the small airways, peribronchiolar fibrosis and airway narrowing may be prominent features. In chronic bronchitis hypertrophy of glandular structures and goblet cell metaplasia in the proximal airways likely contribute to an increased mucus production, the expectoration of which is one defining characteristic of chronic bronchitis.

Consequently, a fermented product according to the present invention may be

effective in preventing an airflow limitation in a subject prone to contracting bronchitis and/or to alleviate any airflow limitation or obstruction already present in said subject. Particularly, a fermented product according to the present invention may be effective in controlling hypertrophy and hyperplasia of glandular structures and goblet cell metaplasia, as well as peribronchiolar fibrosis and a narrowing of the small airways.

Bronchitis may be caused by a number of factors including viral and/or bacterial infection, environmental pollutants including cigarette smoke, and allergy. These factors may occur together or separately. A viral infection may e. g. predispose an individual to a subsequent bacterial infection. Bronchial infections occur in patients with abnormal airways who have reduced host defenses. The three major bacterial pathogens isolated during bronchial infections are non-typable Haemophlus influenzae, moraxella catarrhalis, and Streptococcus pneumoniae. A fermented product according to the present invention may especially be effective in preventing viral and/or bacterial infection in a subject by e. g. increasing the host defenses of said subject.

The term small airways as used herein is defined as small bronchi and bronchioles that contain no cartilage, glands or alveoli in their walls and measure 2 mm or less in internal diameter. The term small airways disease is used for a group of non-specific histological changes of peripheral airways found in individuals with a limited or obstructed airflow, including individuals having features such as mucus plugging, chronic inflammation, and muscular enlargement of small airway walls. Small airways disease is present in some patients with the clinical picture of chronic bronchitis.

Consequently, a fermented product according to the present invention may be effective in preventing, treating, prophylactically treating and/or alleviating a limited or obstructed flow of air through the small airways.

In small or peripheral airways disease, there is inflammation of bronchioli and mucous metaplasia and hyperplasia, increased intraluminal mucus and increased wall muscle.

Consequently, a fermented product according to the present invention may be effective in controlling inflammation of the bronchioli and mucous metaplasia and hyperplasia, and effective in reducing and/or eliminating any increased intraluminal mucus formation and/or any increased wall muscle development.

Detailed The soy protein for use in a fermented product according to the present invention can be provided by isolated soy protein, soy protein concentrate, soy flour or the like or any combination thereof. Isolated soy protein is preferred. Processed Isolated soy protein is particularly preferred.

Isolated soy protein is the major proteinacious fraction of soybeans. It is prepared from high quality, dehulled, defatted soybeans by removing a preponderance of the non- protein components resulting in an isolated soy protein fraction which in the present context shall contain at least 90 percent protein (N x 6.25) on a moisture free basis.

The preparation takes place through a series of steps in which the soybean protein portion is separated from the rest of the soybean. The removal of carbohydrate results in a product, which is essentially bland in flavor and therefore particularly useful in a fermented product for humans.

Soy protein concentrates are made by removing most of the oil and water-soluble non- protein constituents from defatted and dehulled soybeans. In the present context a soy protein concentrate shall preferably contain at least 65 percent protein on a moisture- free basis.

The soy protein can also be provided by soy flour, which can be full-fat or defatted soy flour. Full-fat soy flour comes from whole, dehulled soybeans that have been ground into a fine powder and, as the name implies, still contains the fat naturally found in soybeans. Defatted soy flour comes from whole, dehulled, defatted soybeans that have been ground into a fine powder. Soy flour is approximately 50 percent soy protein on a dry weight basis in the present context.

The soy protein used in a fermented product according to the present invention should preferably supply all the essential amino acids in the amounts required for humans.

Preferably, the soy protein should also meet or exceed the essential amino acid requirement pattern for children and adults as established by the Food and Agricultural Organization, World Health Organization and United Nations University (FAO/WHO, UNU). Furthermore, the preferred soy protein should be comparable in digestibility to milk, meat, fish, and egg protein. Finally, the preferred soy protein shall be effective in maintaining nitrogen balance when consumed at the recommended protein intake level.

Researcher have shown that specific amino acids may to some extent effect serum lipid levels and potentially alleviate cardiovascular diseases. Animal studies have indicated that the amino acid lysine increases serum cholesterol levels, while arginine counteracts this effect (Kurowska et al., J. Nutr. 124,364-370 (1994) and Sanchez et al., Med. Hypotheses 35,324-329 (1991). This observation appears to be in correspondence with the well established influence of NO on vasodilation, since arginine may potentially be converted to citrullin and NO by NO-synthetase. Thus according to a presently preferred hyphothesis soy protein having a high arginine to lysine ratio effects serum lipid levels and alleviates symptoms of cardiovascular diseases to a greater extent than soy protein having a lower or normal arginine to lysine ratio. Consequently, isolated, potentially processed, soy protein having a high arginine to lysine ratio is a particularly preferred soy protein source in a fermented product according to the present invention. Preferably the soy protein of the soy protein source in a fermented product according to the present invention should have an arginine to lysine ratio of at least about 1.0, such as at least about 1.1, for example at least about 1.2, such as at least about 1.3, for example at least about 1.4, such as at least about 1.5, for example at least about 1.6, such as at least about 1.7, for example at least about 1.8, such as at least about 1.9, for example more than about 2, such as at least about 2.1, for example at least about 2.2, such as at least about 2.5, for example at least about 2.75, such as at least about 3, for example more than about 3.3, such as at least about 3.6, for example at least about 4, such as at least about 4.5, for example at least about 5, such as at least about 6, for example at least about 7, such as at least about 8, for example at least about 9, such as at least about 10, for example at least about 11, such as at least about 12, for example at least about 13, such as at least about 14.

Preferred isolated soy protein products meeting some or all of the foregoing requirements are supplied by Protein Technologies International, Inc. under the brand name SUPROO. SUPROE isolated soy proteins are supplied in many different qualities and SUPROO XT 12C is one particularly preferred quality. The currently most preferred quality is termed SUPROO FXP-HO159.

The soy protein is preferably the main protein source in a fermented product according to the present invention. However, parts of the protein source may be provided by other proteins such as e. g. skimmed milk, preferably as a powder, and other vegetable or animal proteins including diary proteins.

In a preferred embodiment of the invention the soy protein is provided by isolated soy protein Phytoestrogen compounds according to the present invention are defined as naturally occurring plant substances, said substances being either structurally or functionally similar to 170-estradiol or generating estrogenic effects. Phytoestrogens consist of a number of classes including isoflavones, coumestans, lignans and resorcylic acid lactones. Examples of isoflavones according to the present invention are genistein, daidzein, equol, glycitein, biochanin A, formononetin, and O-desmethylangolesin. The phytoestrogen compounds of a fermented product according to the present invention are preferably isoflavones, more preferably genistein, daidzein, glycitein and/or equol, yet more preferably genistein and/or daidzein, and even more preferably genistein. A preferred fermented product according to the present invention may accordingly comprise a single isoflavone, such as genistein, daidzein, glycitein or equol, or it may comprise at least one isoflavone selected from the group comprising at least genistein, daidzein, glycitein and equol. When present in the plant the isoflavones are mainly in a glucoside form, i. e. attached to a sugar molecule. This glucoside form can be deconjugated to yield a so-called aglycone form, which is the biologically active species. A fermented product according to the present invention may comprise isoflavones in glucoside and/or aglycone forms regardless of whether the deconjugation to the aglycone form has taken place biologically, in vitro or by any other means whereby the isoflavones are included in a fermented product according to the present invention or if the aglycone forms are the native form of the isoflavones.

The phytoestrogen compound is preferably present in an amount of at least about 0.10 weight percent of the soy protein content. More preferably the phytoestrogen compound is present in an amount of at least 0.10 weight percent of the soy protein content, such as at least about 0.11 weight percent, for example at least about 0.12 weight percent, such as at least about 0.14 weight percent, for example at least about 0.16 weight percent, such as at least about 0.18 weight percent, for example at least about 0.20 weight percent, such as at least about 0.22 weight percent, for example at least about 0.24 weight percent, such as at least about 0.25 weight percent, for example more than about 0.25 weight percent, such as at least about 0.26 weight percent, for example at least about 0.28 weight percent, such as at least about 0.30 weight percent, for example at least about 0.32 weight percent, such as at least about 0.33 weight percent, for example more than about 0.33 weight percent, such as at least about 0.35 weight percent, for example at least about 0.40 weight percent, such

as at least about 0.45 weight percent, for example at least about 0.50 weight percent, such as at least about 0.55 weight percent, for example at least about 0.60 weight percent, such as at least about 0.65 weight percent, for example at least about 0.70 weight percent, such as at least about 0.75 weight percent, for example at least about 0.80 weight percent, such as at least about 0.85 weight percent, for example at least about 0.90 weight percent, such as at least about 1.0 weight percent of the soy protein content, and preferably less than 2.50 weight percent of the soy protein content.

In the past, the downstream processing techniques used in the preparation of soy proteins have included steps that removed and/or destroyed isoflavones. Methods are available today, which provide soy protein products with high, fixed levels of naturally occurring isoflavones. The isoflavones according to the present invention in glucoside and/or aglycone forms can be included in a fermented product according to the present invention as part of such soy protein products and/or by themselves and/or as part of any other fermented product comprising isoflavones.

According to a preferred embodiment of the present invention the soy protein used in a process for the preparation of a fermented product according to the present invention retain the ability to lower blood cholesterol levels upon ingestion, characterised by having more than a further specified minimum content of intact 7S (a + a'+ ß) and/or 11 S subunits (A + B).

Without wishing to be bound by theory it is believed that ingestion of soy proteins with minimum content of intact 7S (a + a'+ (3) and/or US subunits (A + B) facilitate the formation of breakdown products in the digestive tract, in the form of peptides which elicit an effect in the form of cholesterol lowering.

The present invention also provides the use of soy protein in a fermented product according to the present invention, which retains the ability to lower blood cholesterol levels upon ingestion, characterised by minimum content of intact 7S (a + a'+ (3) and/or 11 S subunits (A + B), and also provides for the use of such a fermented product in which the soy protein retains the ability to lower blood cholesterol levels upon ingestion to obtain a health benefit.

In one aspect the present invention provides fermented product comprising soy protein, which retains the ability to lower blood cholesterol levels upon ingestion, characterised by a minimum content of intact 7S (a + a'+ (3) and/or 11 S subunits (A +

B). The soy protein of such a fermented product more specifically are believed facilitate the formation of peptides which has a cholesterol lowering effect.

In another aspect, the present invention provides a method for manufacturing a fermented product product, wherein the soy protein retains the ability to lower blood cholesterol levels upon ingestion and characterised by a minimum content of intact 7S (a + a'+ B) and/or 11S subunits (A + B) The amount of intact 7S subunits (a + a'+ ß) and 11 S subunits (A + B) in the soy protein used in the preparation of a fermented product according to the invention preferably constitute more than 5 % of the total soy protein content, such as more than 10 %, for example more than 15 %, , such as more than 20 %, for example more than 25 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 51 %, such as more than 52 %, for example more than 53 %, such as more than 54 %, for example more than 55 %, such as more than 56 %, for example more than 57 %, such as more than 58 %, for example more than 59 %, such as more than 60 %, for example more than 61 %, such as more than 62 %, for example more than 63 %, such as more than 64 %, for example more than 65 %, such as more than 66 %, for example more than 67 %, such as more than 68 %, for example more than 69 %, such as more than 70 %, for example more than 71 %, such as more than 72 %, for example more than 73 %, such as more than 74 %, for example more than 75 %, such as more than 77 %, for example more than 79 %, such as more than 81 %, for example more than 83 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

The amount of intact 7S subunits (a + a'+ ß) in the soy protein used in the preparation of a fermented product according to the invention preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example

more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

The amount of intact 7S subunit a in the soy protein used in the preparation of a fermented product according to the invention preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such

as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

The amount of intact 7S subunit a'in the soy protein used in the preparation of a fermented product according to the invention preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11%, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

According to a particularly preferred embodiment of the present invention the soy protein used in the preparation of a fermented product according to the invention also contain phytoestrogens, such as isoflavons. The phytoestrogen compound is preferably present in an amount of at least about 0.12 weight percent of the soy protein content, such as at least about 0.14 weight percent, for example at least about 0.16 weight percent, such as at least about 0.18 weight percent, for example at least about 0.20 weight percent, such as at least about 0.22 weight percent, for example at least about 0.24 weight percent, such as at least about 0.25 weight percent, for example more than about 0.25 weight percent, such as at least about 0.26 weight percent, for example at least about 0.28 weight percent, such as at least about 0.30 weight percent, for example at least about 0.32 weight percent, such as at least about

0.33 weight percent, for example more than about 0.33 weight percent, such as at least about 0.35 weight percent, for example at least about 0.40 weight percent, such as at least about 0.45 weight percent, for example at least about 0.50 weight percent, such as at least about 0.55 weight percent, for example at least about 0.60 weight percent, such as at least about 0.65 weight percent, for example at least about 0.70 weight percent, such as at least about 0.75 weight percent, for example at least about 0.80 weight percent, such as at least about 0.85 weight percent, for example at least about 0.90 weight percent, such as at least about 1.0 weight percent of the soy protein content, and preferably less than 2.50 weight percent of the soy protein content.

In soy protein used in the preparation of a fermented product according to the invention with an amount of isoflavones of at least 0.16 weight % of the soy protein, the amount of intact 7S subunits (a + a'+ ß) and 11S subunits (A + B) preferably constitute more 5 % of the total soy protein content, such as more than 10 %, for example more than 15 %, , such as more than 20 %, for example more than 25 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 51 %, such as more than 52 %, for example more than 53 %, such as more than 54 %, for example more than 55 %, such as more than 56 %, for example more than 57 %, such as more than 58 %, for example more than 59 %, such as more than 60 %, for example more than 61 %, such as more than 62 %, for example more than 63 %, such as more than 64 %, for example more than 65 %, such as more than 66 %, for example more than 67 %, such as more than 68 %, for example more than 69 %, such as more than 70 %, for example more than 71 %, such as more than 72 %, for example more than 73 %, such as more than 74 %, for example more than 75 %, such as more than 77 %, for example more than 79 %, such as more than 81 %, for example more than 83 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

In soy protein used in the preparation of a fermented product according to the invention with an amount of isoflavones of at least 0.16 %, the amount of intact 7S subunits (a + a'+ ) preferably constitute more than than 3 %, such as more than 4 %,

for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

In soy protein used in the preparation of a fermented product according to the invention with an amount of isoflavones of at least 0. 16 % of the soy protein, the amount of intact 7S subunit a preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for

example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

In soy protein used in the preparation of a fermented product according to the invention with an amount of isoflavones of at least 0.16 % of the soy protein, the amount of intact 7S subunit a'preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

Optionally a soy protein product used in the preparation of a fermented product according to the invention may have the trypsin inhibitors partly or fully destroyed or removed. The amount of ACTIVE trypsin inhibitor in a soy protein product used in the preparation of a fermented product according to the invention may preferably be less than 50% of the amount in the original soy bean, such as less than 40%, for example less than 30%, such as less than 25%, for example less than 20%, such as less than 15%, for example less than 10%, such as less than 5 %, for example less than 1 %.

The amount of intact 7S subunits (a + a'+ ß) and 11 S subunits (A + B) in the soy protein in a fermented product according to the invention preferably constitute more than 5 % of the total soy protein content, such as more than 10 %, for example more than 15 %, , such as more than 20 %, for example more than 25 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 51 %, such as more than 52 %, for example more than 53 %, such as more than 54 %, for example more than 55 %, such as more than 56 %, for example more than 57 %, such as more than 58 %, for example more than 59 %, such as more than 60 %, for example more than 61 %, such as more than 62 %, for example more than 63 %, such as more than 64 %, for example more than 65 %, such as more than 66 %, for example more than 67 %, such as more than 68 %, for example more than 69 %, such as more than 70 %, for example more than 71 %, such as more than 72 %, for example more than 73 %, such as more than 74 %, for example more than 75 %, such as more than 77 %, for example more than 79 %, such as more than 81 %, for example more than 83 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

The amount of intact 7S subunits (a + a'+ (3) in the soy protein in a fermented product according to the invention preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such

as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

The amount of intact 7S subunit a in the soy protein in a fermented product according to the invention preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

The amount of intact 7S subunit a'in the soy protein in a fermented product according to the invention preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more

than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

According to a particularly preferred embodiment of the present invention the soy protein in a fermented product according to the invention also contain phytoestrogens, such as isoflavons. The phytoestrogen compound is preferably present in an amount of at least about 0.12 weight percent of the soy protein content, such as at least about 0.14 weight percent, for example at least about 0.16 weight percent, such as at least about 0.18 weight percent, for example at least about 0.20 weight percent, such as at least about 0.22 weight percent, for example at least about 0.24 weight percent, such as at least about 0.25 weight percent, for example more than about 0.25 weight percent, such as at least about 0.26 weight percent, for example at least about 0.28 weight percent, such as at least about 0.30 weight percent, for example at least about 0.32 weight percent, such as at least about 0.33 weight percent, for example more than about 0.33 weight percent, such as at least about 0.35 weight percent, for example at least about 0.40 weight percent, such as at least about 0.45 weight percent, for example at least about 0.50 weight percent, such as at least about 0.55 weight percent, for example at least about 0.60 weight percent, such as at least about 0.65 weight percent, for example at least about 0.70 weight percent, such as at least about 0.75 weight percent, for example at least about 0.80 weight percent, such as at least about 0.85 weight percent, for example at least about 0.90 weight percent, such as at least about 1.0 weight percent of the soy protein content, and preferably less than 2.50 weight percent of the soy protein content.

In soy protein in a fermented product according to the invention with an amount of isoflavones of at least 0.16 weight % of the soy protein, the amount of intact 7S subunits (a + a'+ (3) and US subunits (A + B) preferably constitute more 5 % of the total soy protein content, such as more than 10 %, for example more than 15 %, , such as more than 20 %, for example more than 25 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 51 %, such as more than 52 %, for example more than 53 %, such as more than 54 %, for example more than 55 %, such as more than 56 %, for example more than 57 %, such as more than 58 %, for example more than 59 %, such as more than 60 %, for example more than 61 %, such as more than 62 %, for example more than 63 %, such as more than 64 %, for example more than 65 %, such as more than 66 %, for example more than 67 %, such as more than 68 %, for example more than 69 %, such as more than 70 %, for example more than 71 %, such as more than 72 %, for example more than 73 %, such as more than 74 %, for example more than 75 %, such as more than 77 %, for example more than 79 %, such as more than 81 %, for example more than 83 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

In soy protein in a fermented product according to the invention with an amount of isoflavones of at least 0.16 %, the amount of intact 7S subunits (a + a'+ (3) preferably constitute more than than 3 %, such as more than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such

as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

In soy protein in a fermented product according to the invention with an amount of isoflavones of at least 0.16 % of the soy protein, the amount of intact 7S subunit a preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 %, for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

In soy protein in a fermented product according to the invention with an amount of isoflavones of at least 0.16 % of the soy protein, the amount of intact 7S subunit a' preferably constitute more than 1 % of the total soy protein content, such as more than 2 %, for example more than 3 %, such asmore than 4 %, for example more than 5 %, such as more than 6 %, for example more than 7 %, such as more than 8 %, for

example more than 9 %, such as more than 10 %, for example more than 11 %, such as more than 12 %, for example more than 13 %, such as more than 14 %, for example more than 15 %, such as more than 16 %, for example more than 17 %, such as more than 18 %, for example more than 19 %, such as more than 20 %, for example more than 21 %, such as more than 22 %, for example more than 23 %, such as more than 24 %, for example more than 25 %, such as more than 26 %, for example more than 27 %, such as more than 28 %, for example more than 29 %, such as more than 30 for example more than 31 %, such as more than 32 %, for example more than 33 %, such as more than 34 %, for example more than 35 %, such as more than 36 %, for example more than 37 %, such as more than 38 %, for example more than 39 %, such as more than 40 %, for example more than 41 %, such as more than 42 %, for example more than 43 %, such as more than 44 %, for example more than 45 %, such as more than 46 %, for example more than 47 %, such as more than 48 %, for example more than 49 %, such as more than 50 %, for example more than 55 %, such as more than 60 %, for example more than 65 %, such as more than 70 %, for example more than 75 %, such as more than 80 %, such as more than 85 %, for example more than 90 %, such as more than 95 %.

The phospholipid source according to the present invention will preferably comprise polyunsaturated fatty acids and monounsaturated fatty acids and optionally also saturated fatty acids. Soy lecithins and [l-linolenic acid are particularly preferred. The phospholipid source will preferably comprise at least about 5% phosphatidyl choline, such as at least 10% phosphatidyl choline. The phospholipid source will more preferably comprise at least about 20% phosphatidyl choline, such as at least about 30% phosphatidyl choline, for example at least about 35% phosphatidyl choline, such as at least about 40% phosphatidyl choline, for example at least about 45% phosphatidyl choline, such as at least about 50% phosphatidyl choline, for example more than about 55% phosphatidyl choline phosphatidyl choline by weight, such as at least 60% phosphatidyl choline, for example at least about 65% phosphatidyl choline, such as at least about 70% phosphatidyl choline, for example at least about 71 % phosphatidyl choline, such as at least about 72% phosphatidyl choline, for example at least about 73% phosphatidyl choline, such as at least about 74% phosphatidyl choline, for example more than about 75% phosphatidyl choline, such as at least about 76% phosphatidyl choline, for example at least about 77% phosphatidyl choline, such as at least about 78% phosphatidyl choline, for example at least about 79% phosphatidyl choline, for example more than about 80% phosphatidyl choline, such as at least about 85% phosphatidyl choline, for example at least about 90% phosphatidyl

choline, such as at least about 98% phosphatidyl choline, for example 100% phosphatidyl choline by weight.

The phospholipid source will preferably comprise polyunsaturated fatty acids and monounsaturated fatty acids and optionally also saturated fatty acids. The amount of polyunsaturated fatty acids and monounsaturated fatty acids, including the essential fatty acids, may range from 35 to 50, preferably 38 to 44, weight percent of the total amount of the fat source. The essential fatty acids are also called omega-6 and omega-3 fatty acids and include linolic acid and/or linolenic acid (D-linolenic acid). The amount of saturated fatty acids may be from 20 to 30 weight percent, preferably 22 to 26 weight percent, of the total amount of the phospholipid source. In a fermented product according to the present invention, the phospholipid source usually provides from 5 to 70 percent, preferably 10 to 60 percent, such as from 15 to 50 percent, for example from 20 to 40 percent, such as from 25 to 35 percent of the total energy content of the fermented product.

The phospholipid source preferably provides at least about 5 percent of the total energy content of the fermented product, such as at least about 10 percent, for example at least about 15 percent, such as at least about 20 percent, for example at least about 21 percent, such as at least about 22 percent, for example at least about 23 percent, such as at least about 24 percent, for example more than about 25 percent, such as at least about 26 percent, for example at least about 27 percent, such as at least about 28 percent, for example at least about 29 percent, such as at least about 30 percent, for example more than about 31 percent, such as at least about 32 percent, for example at least about 33 percent, such as at least about 34 percent, for example at least about 35 percent, such as at least about 36 percent, for example at least about 37 percent, such as at least about 38 percent, for example at least about 39 percent, such as at least about 40 percent, for example at least about 45 percent, such as at least about 50 percent, for example at least about 55 percent, such as at least about 60 percent, for example at least about 65 percent of the total energy content of the fermented product, and preferably less than 70 percent of the total energy content of the fermented product.

Preferred phospholipid sources are lecithins and even more preferably soy lecithin.

Currently preferred lecithin products are manufactured by SKW Nature Products, BioActives, Freising, Germany are marketed under the brand name of Epikuron 1000, Epikuron 1300.

The dietary fibers used in a presently preferred embodiment of the present invention should preferably comprise a mixture of insoluble fibers and water-soluble fibers also referred to as soluble fibers. Soluble fibers have a lowering effect on blood cholesterol levels. Examples of dietary fibers comprising soluble fibers are fibers from apples, bananas, oranges, carrots, oats, and soybeans. The dietary fibers preferably comprise soluble fibers in an amount of about 5 weight percent, such as about 10 weight percent, for example about 15 weight percent, such as about 20 weight percent, for example about 25 weight percent, such as about 30 weight percent, for example about 35 weight percent, such as about 40 weight percent, for example about 45 weight percent, such as about 50 weight percent, for example about 55 weight percent, such as about 60 weight percent, for example about 65 weight percent, such as about 70 weight percent, for example about 75 weight percent, such as about 80 weight percent, for example about 85 weight percent, such as about 90 weight percent, for example about 95 weight percent. The dietary fibers used in the present invention are preferably soybean fibers, more preferably soy cotyledon fibers. Such fibers are derived from dehulled and defatted soybean cotyledon and are comprised of a mixture of soluble and insoluble fibers. Soy cotyledon fibers are distinctly different from soybean fibers derived from soy hulls as well as other fiber sources. Soy cotyledon fibers are bland tasting, contain no cholesterol, are low in fat and sodium, and they have good water-binding properties and low caloric content.

Soy cotyledon fibers supplied in a fat-modified and low-cholesterol diet are known to further reduce serum cholesterol levels in a subject suffering from mild to severe hypercholesterolemia. The effect is a lowering of the serum levels of total cholesterol including a lowering of the serum levels of LDL-cholesterol. However, HDL-cholesterol and total triglycerides are not significantly affected by soy cotyledon fibers. Soybean fibers, in particular soy cotyledon fibers, are believed to provide a synergistic effect in combination with soy protein and/or with a phytoestrogen compound, such as naturally occurring isoflavones, or to exert a potentiating effect on the soy protein and/or the phytoestrogen compound, said synergistic or potentiating effect being effective in lowering serum levels of lipid and cholesterol in subjects having normal as well as elevated serum levels of total cholesterol and total triglycerides.

Without wishing to be bound by any specific theory it is presently believed that both soluble dietary fibers (working as nutrients) and insoluble dietary fibers (working as bulking agents), in particular from soybean fibers, more particularly soy cotyledon

fibers, provide favorable growth conditions for the microflora in the human gut, which makes the microflora more effective in deconjugating isoflavones in the glucoside form to the aglycone form. Isoflavones in the aglycone form are absorbed faster and to a greater extent in the human gut than isoflavones in the glucoside form, and isoflavones in the aglycone form are the biologically more active species in the present context. In view hereof it can be understood that administration of a combination of soy proteins, a high, fixed level of isoflavones and a combination of soluble and insoluble fibers may be effective in providing an increased uptake of isoflavones.

Furthermore, again without wishing to be bound by any specific theory, it is presently believed that both soluble dietary fibers (working as nutrients) and insoluble dietary fibers (working as bulking agents), in particular from soybean fibers, more particularly soy cotyledon fibers, provide favorable growth conditions for the microflora in the human gut, which makes the microflora more effective in converting phosphatidyl serine and phosphatidyl ethanolamine into phosphatidyl choline. This capability to decarboxylate phosphatidyl serine into phosphatidyl ethanolamine by the action of pyridoxal phosphate enzymes and further methylat phosphatidyl ethanolamine into phosphatidyl choline has presently only been proven for bacteria. Phosphatidyl choline are absorbed faster and to a greater extent in the human gut than phosphatidyl serine and phosphatidyl ethanolamine, and phosphatidyl choline is the biologically more active species in the present context. In view hereof it can be understood that administration of a combination of soy proteins, a phospholipid source having a high fixed level of phosphoglycerides and a combination of soluble and insoluble fibers may be effective in providing increased levels of phosphatidyl choline from a given phospholipid source and hence provide an increased uptake of phosphatidyl choline from a given phospholipid source.

The amount of dietary fibers of the total weight of a fermented product according to the present invention on a dry basis is preferably more than 2 weight percent, for example at least 4 weight percent, such as at least 6 weight percent, for example at least 7 weight percent, such as at least 8 weight percent, for example at least 9 weight percent, such as at least 10 weight percent, for example at least 11 weight percent, such as at least 12 weight percent, for example at least 13 weight percent, such as at least 14 weight percent, for example at least 15 weight percent, such as at least 16 weight percent, for example at least 17 weight percent, such as at least 18 weight percent, for example at least 19 weight percent, such as at least 20 weight percent, and preferably less than 50 weight percent.

The preferred daily dosage of soybean fibers is from at least 1 g to about 100 g soybean fibers, for example from at least 2 to about 75 g soybean fibers, such as from at least 3 g to about 50 g, for example from at least 4 g to about 40 g, such as from at least 5 to about 30 g, such as from at least 10 g to about 20 g soybean fibers.

Preferred soy cotyledon fiber products manufactured by Protein Technologies International, Inc. are marketed under the brand name of FIBRIMO. Among the various soybean fibers produced under the FIBRIM (D brand, FIBRIMO 1020 is particularly preferred because of a particularly pleasant mouth feel and dispersability for dry blended beverage applications. FIBRIMO 2000 is presently preferred for use in ready- made liquids.

Some compositions of isolated soy protein and soy cotyledon fiber are preferred in order to maximize the content of soy protein and isoflavones contained therein namely SUPRO FXP-H0159, SUPRO FXP-H0161, FIBRIMO 1450, FIBRIM 2000 and FIBRIMO 1020 for dry blended beverage applications and SUPRO FXP-H0159, SUPROO FXP-H0161, FIBRIMO 1450, FIBRIMO 2000 and FIBRIM (g) 1020 for use in ready made liquids.

When a fermented product according to the present invention is for use in the prevention and/or treatment of type 2 diabetes, the metabolic syndrome and associated cardiovascular diseases, lecithinated fat reduced cacao is particularly preferred. Other preferred carbohydrates for use in a fermented product according to the present invention for use in the prevention and/or treatment of type 2 diabetes, the metabolic syndrome and associated cardiovascular diseases are polydextrose or saccharose, but these should be limited using other sweeteners like e. g. aspartame.

Vitamins and minerals may optionally be added to a fermented product according to the present invention in accordance with the limits laid down by health authorities. A fermented product according to the present invention may comprise all recommended vitamins and minerals. The vitamins will typically include A, B1, B2, B12, folic acid, niacin, panthotenic acid, biotin, C, D, E and K. The minerals will typically include iron, zinc, iodine, copper, manganese, chromium and selenium. Electrolytes, such as sodium, potassium and chlorides, trace elements and other conventional additives may also be added in recommended amounts.

A fermented product according to the present invention may be used for special dietary use, preferably for lowering serum levels of total cholesterol and/or LDL- cholesterol and/or triglycerides in subjects such as hyperlipidemic patients or normocholesterolemic patients suffering from a cardiovascular disease, and/or for lowering serum levels of glucose and/or insulin and/or total cholesterol and/or LDL- cholesterol and/or triglycerides and/or for increasing glucose tolerance and/or insulin sensitivity and/or for preventing, treating and/or alleviating impaired glucose tolerance and/or insulin secretory failure in diabetic subjects and/or for preventing, treating and/or alleviating an arteriosclerotic condition by reducing the influx of lipoproteins and/or cholesterol and/or triglycerides into the endocelium of the arterial wall of a diabetic subject suffering from a cardiovascular disease. For example, from one to three daily meals of ordinary food can be supplemented or replaced by a fermented product according to the present invention. Hereby, significant reductions in serum levels of cholesterol and/or LDL-cholesterol and/or triglycerides can be obtained, as well as an improvement of serum HDL/LDL-cholesterol ratio and/or an increase in serum HDL-cholesterol levels. The fermented product may provide from about 50 to about 250 kcal per serving.

The fermented product according to the present invention is effective in lowering levels of cholesterol in normocholesterolemic patients by at least 2%, for example at least 5%, such as at least 8%, for example at least 10%, such as at least 12%, for example at least 14%, such as at least 16%, for example at least 18%, such as at least 20%, for example at least 25%, such as at least 30%. The fermented product according to the present invention is effective in lowering levels of triglycerides in normocholesterolemic patients by at least 10%, such as at least 12%, for example at least 14%, such as at least 16%, for example at least 18%, such as at least 20%, for example at least 25%, such as at least 30%.

The fermented product according to the present invention is effective in lowering levels of cholesterol in mildly hypercholesterolemic patients by at least 3%, for example at least 5%, such as at least 8%, for example at least 10%, such as at least 12%, for example at least 15%, such as at least 20%, for example at least 25%, such as at least 30%, for example at least 35%, such as at least 40%, for example at least 45%.

The fermented product according to the present invention is effective in lowering levels of triglycerides in mildly hypercholesterolemic patients by at least 15%, such as at

least 20%, for example at least 25%, such as at least 30%, for example at least 35%, such as at least 40%, for example at least 45%.

The fermented product according to the present invention is effective in lowering levels of cholesterol in severely hypercholesterolemic patients by at least 3%, for example at least 5%, such as at least 8%, for example at least 10%, such as at least 12%, for example at least 15%, such as at least 20%, for example at least 25%, such as at least 30%, for example at least 35%, such as at least 40%, for example at least 45%, such as at least 50%, for example at least 55%, such as at least 60%. The fermented product according to the present invention is effective in lowering levels of triglycerides in severely hypercholesterolemic patients by at least 20%, for example at least 25%, such as at least 30%, for example at least 35%, such as at least 40%, for example at least 45%, such as at least 50%, for example at least 55%, such as at least 60%.

In another embodiment the present invention provides the use of a fermented product according to the present invention in the treatment of cardiovascular diseases in the human or animal body in an amount effective in lowering serum levels of total cholesterol and/or LDL-cholesterol and/or triglycerides and/or homocystein and/or increasing the serum HDULDL-cholesterol ratio and/or serum HDL-cholesterol levels and/or reducing the influx of cholesterol and/or triglycerides into the arterial wall and/or reducing the amount of oxidized LDL-cholesterol present in the arterial wall and/or preventing, reducing or eliminating fatty streak formation and/or preventing, reducing or eliminating fibrous plaque formation and/or preventing, reducing or eliminating complicated lesion formation and/or reducing or eliminating the risk of a subject contracting angina pectoris and/or reducing or eliminating the risk of a subject contracting a myocardial infarction, and/or alleviating the clinical condition of patients contracting a myocardial infection. The cardiovascular disease is preferably a cardiovascular disease selected from the group comprising hypercholesterolemia, hypertriglyceridemia, other hyperlipidemias, arteriosclerosis, atherosclerosis, arteriolosclerosis, coronary heart disease, angina pectoris, thrombosis, myocardial infarction, and hypertension and more preferred selected from arteriosclerosis and atherosclerosis.

In another embodiment the present invention provides the use of a fermented product according to the present invention in the treatment of type 2 diabetes and/or the metabolic syndrome in an amount effective in lowering serum levels of total cholesterol and/or LDL-cholesterol and/or triglycerides and/or glucose and/or increasing serum

levels of HLDL-cholesterol and/or homocystein and/or reducing the influx of cholesterol and/or triglycerides into the arterial wall and/or reducing the amount of oxidized LDL-cholesterol present in the arterial wall and/or improving glucose tolerance and/or increasing insulin sensitivity and/or alleviating impaired glucose tolerance and/or improving insulin secretion and/or reducing or eliminating fatty streak formation and/or preventing, reducing or eliminating fibrous plaque formation and/or preventing, reducing or eliminating complicated lesion formation and/or preventing, reducing or eliminating the risk of a subject contracting angina pectoris and/or preventing, reducing or eliminating the risk of a subject contracting a myocardial infarction and/or preventing, treating, prophylactically treating, alleviating and/or eliminating hypertension and/or hyperglycemia and/or hyperinsulinemia and/or hypercholesterolemia and/or hypertriglyceridemia and/or arteriosclerosis and/or atherosclerosis and/or arteriolosclerosis in a diabetic subject.

In another embodiment the present invention provides the use of a fermented product according to the present invention in the treatment of a pulmonary disease in a human or animal body, preferably a disease selected from the group comprising inflammation of the airways, bronchoconstriction, bronchitis, asthma, and small airways diseases, in an amount effective in preventing, treating, prophylactically treating and/or alleviating inflammation of the airways and/or bronchoconstriction and/or bronchitis and/or small airways diseases and/or asthma and/or reducing and/or eliminating mucus hypersecretion and/or dyspnea in a subject suffering from asthma and/or increasing FEV1 of a subject as measured by forced expiratory volume in the first second of expiration.

The present invention also provides a method of preventing, treating, prophylactically treating and/or alleviating by therapy a cardiovascular disease in the human or animal body such as an arteriosclerotic condition of a human or animal body, said method comprising administration of a fermented product according to the present invention in an amount effective in lowering serum levels of total cholesterol and/or LDL- cholesterol and/or triglycerides and/or homocystein and/or increasing the serum HDL/LDL-cholesterol ratio and/or serum HDL-cholesterol levels and/or reducing the influx of cholesterol and/or triglycerides into the arterial wall and/or reducing the amount of oxidized LDL-cholesterol present in the arterial wall and/or preventing, reducing or eliminating fatty streak formation and/or preventing, reducing or eliminating fibrous plaque formation and/or preventing, reducing or eliminating complicated lesion formation and/or reducing or eliminating the risk of a subject contracting angina

pectoris and/or reducing or eliminating the risk of a subject contracting a myocardial infarction, and/or alleviating the clinical condition of patients contracting a myocardial infection. The cardiovascular disease is preferably a cardiovascular disease selected from the group comprising hypercholesterolemia, hypertriglyceridemia, other hyperlipidemias, arteriosclerosis, atherosclerosis, arteriolosclerosis, coronary heart disease, angina pectoris, thrombosis, myocardial infarction, and hypertension and more preferred selected from arteriosclerosis and atherosclerosis.

The present invention also provides a method of preventing and/or treating by therapy type 2 diabetes and/or the metabolic syndrome in a human or animal body, said method comprising administration to said human or animal body of a fermented product according to the present invention in an amount effective in lowering serum levels of total cholesterol and/or LDL-cholesterol and/or triglycerides and/or glucose and/or increasing serum levels of HLDL-cholesterol and/or homocystein and/or reducing the influx of cholesterol and/or triglycerides into the arterial wall and/or reducing the amount of oxidized LDL-cholesterol present in the arterial wall and/or improving glucose tolerance and/or increasing insulin sensitivity and/or alleviating impaired glucose tolerance and/or improving insulin secretion and/or reducing or eliminating fatty streak formation and/or preventing, reducing or eliminating fibrous plaque formation and/or preventing, reducing or eliminating complicated lesion formation and/or preventing, reducing or eliminating the risk of a subject contracting angina pectoris and/or preventing, reducing or eliminating the risk of a subject contracting a myocardial infarction and/or preventing, treating, prophylactically treating, alleviating and/or eliminating hypertension and/or hyperglycemia and/or hyperinsulinemia and/or hypercholesterolemia and/or hypertriglyceridemia and/or arteriosclerosis and/or atherosclerosis and/or arteriolosclerosis in a diabetic subject.

The present invention also provides a method of preventing, treating, prophylactically treating and/or alleviating by therapy a pulmonary disease in a human or animal body, preferably a disease selected from the group comprising inflammation of the airways, bronchoconstriction, bronchitis, asthma, and small airways diseases, said method comprising administration to said human or animal body of a fermented product according to the present invention in an amount effective in preventing, treating, prophylactically treating and/or alleviating inflammation of the airways and/or bronchoconstriction and/or bronchitis and/or asthma and/or small airways diseases and/or reducing and/or eliminating mucus hypersecretion and/or dyspnea in a subject

suffering from asthma and/or increasing FEV1 of a subject as measured by forced expiratory volume in the first second of expiration.

The period of treatment is preferably in the range of from 1 to 12 months or more, such as from 2 weeks to 9 months, for example from 3 weeks to 6 months, such as from 4 weeks to 4 months, such as from 6 weeks to 3 months. However, the period of treatment shall not be limited to these periods and may e. g. be longer than 12 months, such as e. g. a lifelong treatment in order to prevent cardiovascular diseases or in order to prevent and/or alleviate type 2 diabetes and/or a cardiovascular disease in connection therewith or in order to prevent pulmonary diseases.

In another embodiment, the present invention relates to the use of a fermented product according to the present invention as a partial or total diet for an overweight subject, an overweight subject suffering from an arteriosclerotic condition or an overweight subject suffering from a diabetic condition. Obesity is believed to be one of the major causes of diabetes including type 2 diabetes. Overweight subjects, including overweight diabetic subjects, often have increased serum cholesterol levels and increased triglyceride levels and are therefore more likely to develop cardiovascular diseases. However, the present invention is not limited to treating subjects with an increased risk of contracting a cardiovascular disease, i. e. subjects likely to have increased serum levels of cholesterol and/or triglycerides, or to treating obese diabetic subjects with an increased risk of contracting a cardiovascular disease, i. e. obese diabetic subjects likely to have increased serum levels of cholesterol and/or triglycerides. A fermented product according to the present invention also has substantial serum cholesterol, serum LDL-cholesterol and serum triglyceride lowering effects in subjects having a more normal lipid profile and in diabetic subjects that do not also suffer from overweight. The medical use of a fermented product to the present invention is not limited to overweight or obese subjects, including diabetic subjects, but may be used for normal weight subjects having increased serum levels of cholesterol and/or LDL-cholesterol and/or triglycerides or for subjects with a cardiovascular condition such as e. g. arteriosclerosis or a related condition who have normal serum levels of cholesterol and/or LDL-cholesterol and/or triglycerides. Such increased serum levels of cholesterol and/or LDL-cholesterol and/or triglycerides may be caused by intake of a diet rich in fats or it may be genetically related.

EXAMPLE 1 Preparation of a cultured dairy product containing exogenously added soy protein 1 A cultured dairy product is prepared using the following ingredients: Weight percent of Protein source cultured dairy product Isolated soy protein product, SUPROO PLUS 10.66 161 Soy cotyledon fibres, F) BR) M@ 1020 2.67 Soy lecithin Epikuron 100SP* 0.76 Water Distilled water 64.17 Milk composition Skimmed milk 16.61 Skimmed milk powder 3.99 Distilled water 0.72 Calgon 0. 087 Pectin LM-107 AS-YA 0.250 Agar-agar 0.063 Fermentation culture CH1 0.03 Sum 100 * = supplied by Lucas Meyer, Hamburg, Germany The protein source is dispersed in distilled water having a temperature of 50°C and the dispersion is heated to 82°C and homogenised at 300 bar. In the meantime, the milk composition is prepared by dissolving Calgon@ in milk/water and then adding skimmed milk powder along with pectin and agar-agar. The homogenized dispersion and the milk composition is mixed and heated to a temperature of 90°C for 5 minutes, cooled to 43°C and inoculated with the fermentation culture, CH1 (Chr. Hansen). The fermentation proceeds for 5.5 hours and then the fermented mixture is homogenized at 30 bar, filled into small aseptic jars (0.5-1 I) and cooled to 4-5°C.

A cultured dairy product prepared according to this example contains isolated soy protein product in an amount of about 10.7 weight percent corresponding to an amount

of exogenously added soy protein of about 9.3 weight percent. When the amount of soy protein provided by the soy fibre product is included, the cultured dairy product contains exogenously added soy protein in an amount of about 9.6 weight percent.

The content of the final fruited product is as follows : Content per 100 g Cultured dairy product 83 g Fruit log Sugar 7 g Sum 100 g This fruited cultured dairy product contains isolated soy protein product in an amount of 8.9 weight percent corresponding to an amount of exogenously added soy protein of about 7.7 weight percent. When the amount of soy protein provided by the soy fibre product is included, the fruited cultured dairy product contains exogenously added soy protein in an amount of about 8.0 weight percent.

The cultured dairy product appears like yoghurt and has no soy odour and a smooth texture without powdery mouthfeel.

EXAMPLE 2 Preparation of a cultured dairv product containing exogenously added soy protein and heat inactivated Lactobacillus helveticus 1 A cultured dairy product prepared according to EXAMPLE 1 was found to develop a bitter taste relatively rapidly during storage, which would make the product appalling to people sensitive to bitterness and hence reduce the shelf-life of the product. The flavour was found to be impossible to overshadow by adding increasing amounts of sugar and fruit compounds. In order to counteract the development of bitterness in the product the fermentation culture was supplemented with heat inactivated Lactobacillus helveticus, in the form of Enzobact@. Accordingly, a cultured dairy product was prepared using the following ingredients: Weight percent of Protein source cultured dairy product Isolated soy protein product, SUPRO PLUS 10.66 161 Soy cotyledon fibres, FIBRIM (D 1020 2.67 Soy lecithin Epikuron 100SP* 0.76 Water Distilled water 64.17 Milk composition Skimmed milk 16.54 Skimmed milk powder 3.97 Distilled water 0.72 Ca) gon@ 0.086 Pectin LM-107 AS-YA 0.249 Agar-agar 0.062 Fermentation culture CH1 0. 03 Enzobact 0.1 Sum 100 The protein source is dispersed in distilled water having a temperature of 50°C and the dispersion is heated to 82°C and homogenised at 300 bar. In the meantime, the milk composition is prepared by dissolving Calgon in milk/water and then adding

skimmed milk powder along with pectin and agar-agar. The homogenized dispersion and the milk composition is mixed and heated to a temperature of 90°C for 5 minutes, cooled to 46°C and inoculated with the fermentation culture, CH1 (Chr. Hansen) and Enzobact (MediPharm). The fermentation proceeds for 6 hours and then the fermented mixture is homogenized at 30 bar, filled into small aseptic jars (0. 5-1 1) and cooled to 4-5°C.

A cultured dairy product prepared according to this example contains isolated soy protein product in an amount of about 10.7 weight percent corresponding to an amount of exogenously added soy protein of about 9.3 weight percent. When the amount of soy protein provided by the soy fibre product is included, the cultured dairy product contains exogenously added soy protein in an amount of about 9.6 weight percent.

The content of the final fruited product is as follows : Content per 100 g Cultured dairy product 83 g Fruit log Sugar 7 g Sum 100 g This fruited cultured dairy product contains isolated soy protein product in an amount of 8.8 weight percent corresponding to an amount of exogenously added soy protein of about 7.6 weight percent. When the amount of soy protein provided by the soy fibre product is included, the fruited cultured dairy product contains exogenously added soy protein in an amount of about 8.0 weight percent.

The cultured dairy product appears like yoghurt has a pH of 4.83, a lactic acid content of 1.15%, and displays no soy odour, a smooth texture without powdery mouthfeel and a reduced tendency to become bitter upon storing compared to the product prepared according to EXAMPLE 1.

EXAMPLE 3 Preparation of a cultured dairy product containing exogenousiv added sov protein and heat inactivated Lactobacillus helveticus 2 A cultured dairy product prepared according to EXAMPLE 1 was found to develop a bitter taste relatively rapidly during storage, which would make the product appalling to people sensitive to bitterness and hence reduce the shelf-life, while preparing a product according to EXAMPLE 2, was found to partially counteract the development of bitter taste. In order to estimate whether the development of bitterness in the product could be counteracted by supplementing the fermentation culture with reduced amounts of heat inactivated Lactobacillus helveticus, in the form of Enzobact@, a cultured dairy product was prepared using the following ingredients: Weight percent of Protein source cultured dairy product Isolated soy protein product, SUPRO PLUS 10.66 161 Soy cotyledon fibres, FIBRIM# 1020 2.67 Soy lecithin Epikuron 100SP* 0.76 Water Distilled water 64.17 Milk composition Skimmed milk 16.58 Skimmed milk powder 3.98 Distilled water 0.72 Ca ! gon@ 0.087 Pectin LM-107 AS-YA 0.249 Agar-agar 0.062 Fermentation culture CH1 0.03 Enzobact 0. 0435 Sum 100 The protein source is dispersed in distilled water having a temperature of 50°C and the dispersion is heated to 82°C and homogenised at 300 bar. In the meantime, the milk composition is prepared by dissolving Calgon@ in milk/water and then adding

skimmed milk powder along with pectin and agar-agar. The homogenized dispersion and the milk composition is mixed and heated to a temperature of 90°C for 5 minutes, cooled to 46°C and inoculated with the fermentation culture, CH1 (Chr. Hansen) and Enzobact (MediPharm). The fermentation proceeds for 6 hours and then the fermented mixture is homogenized at 30 bar, filled into small aseptic jars (0.5-1 I) and cooled to 4-5°C.

A cultured dairy product prepared according to this example contains isolated soy protein product in an amount of about 10.6 weight percent corresponding to an amount of exogenously added soy protein of about 9.2 weight percent. When the amount of soy protein provided by the soy fibre product is included, the cultured dairy product contains exogenously added soy protein in an amount of about 9.6 weight percent.

The content of the final fruited product is as follows : Content per 100 g Cultured dairy product 83 g Fruit log Sugar 7 g Sum 100 g This fruited cultured dairy product contains isolated soy protein product in an amount of 8.7 weight percent corresponding to an amount of exogenously added soy protein of about 7.6 weight percent. When the amount of soy protein provided by the soy fibre product is included, the fruited cultured dairy product contains exogenously added soy protein in an amount of about 8.0 weight percent.

The cultured dairy product appears like yoghurt has a pH of 4.50, a lactic acid content of 1.54%, and displays no soy odour, a smooth texture without powdery mouthfeel and a reduced tendency to become bitter upon storing compared to the product prepared according to EXAMPLE 1. A comparison of the tendency to develop bitterness between the products prepared according to EXAMPLE 2 and the present EXAMPLE revealed that the product prepared according to the present EXAMPLE had a slightly higher tendency to develop bitterness upon storage.

EXAMPLE 4 Preparation of a cultured dairy product containing improved exogenousiv added soy protein and heat inactivated Lactobacillus helveticus 1 A cultured dairy product prepared according to EXAMPLE 2was found to partially counteract the development of bitter taste during storage, and i. e. addition of Enzobact prolonged the shelf-life of the product. In order to estimate the effect of the isolated soy protein product source used on the development of bitterness in the product the isolated soy protein product, SUPRO PLUS 161 used in EXAMPLE 2 was replaced with another isolated soy protein product, SUPROX PLUS 219.

Accordingly, a cultured dairy product was prepared using the following ingredients: Weight percent of Protein source cultured dairy product Isolated soy protein product, SUPRA0 PLUS 10.66 219 Soy cotyledon fibres, FIBRIM# 1020 2.67 Soy lecithin Epikuron 100SP* 0.76 Water Distilled water 64.17 Milk composition Skimmed milk 16.54 Skimmed milk powder 3.97 Distilled water 0.72 Calgon 0. 086 Pectin LM-107 AS-YA 0.249 Agar-agar 0.062 Fermentation culture CH1 0.03 Enzobact 0.1 Sum 100 The protein source is dispersed in distilled water having a temperature of 50°C and the dispersion is heated to 82°C and homogenised at 300 bar. In the meantime, the milk composition is prepared by dissolving Ca ! gon@ in milk/water and then adding skimmed milk powder along with pectin and agar-agar. The homogenized dispersion

and the milk composition is mixed and heated to a temperature of 90°C for 5 minutes, cooled to 46°C and inoculated with the fermentation culture, CH1 (Chr. Hansen) and Enzobact (MediPharm). The fermentation proceeds for 6 hours and then the fermented mixture is homogenized at 30 bar, filled into small aseptic jars (0.5-1 I) and cooled to 4-5°C.

A cultured dairy product prepared according to this example contains isolated soy protein product in an amount of about 10.7 weight percent corresponding to an amount of exogenously added soy protein of about 9.3 weight percent. When the amount of soy protein provided by the soy fibre product is included, the cultured dairy product contains exogenously added soy protein in an amount of about 9.6 weight percent.

The content of the final fruited product is as follows : Content per 100 g Cultured dairy product 83 g Fruit log Sugar 7 g Sum 100 g This fruited cultured dairy product contains isolated soy protein product in an amount of 8.8 weight percent corresponding to an amount of exogenously added soy protein of about 7.6 weight percent. When the amount of soy protein provided by the soy fibre product is included, the fruited cultured dairy product contains exogenously added soy protein in an amount of about 8.0 weight percent.

The cultured dairy product appears like yoghurt has a pH of 4.76, a lactic acid content of 1.23%, and displays no soy odour, a smooth texture without powdery mouthfeel and does not become bitter upon storing.

EXAMPLE 5 Preparation of a cultured dairy product containing improved exogenous) v added soy protein and reduced amounts of heat inactivated Lactobacillus helveticus 2 A cultured dairy product prepared according to EXAMPLE 3 was found to partially counteract the development of bitter taste during storage, and i. e. addition of Enzobact# prolonged the shelf-life of the product. In order to estimate the effect of the isolated soy protein product source used on the development of bitterness in the product the isolated soy protein product, SUPRO PLUS 161 used in EXAMPLE 3, was replaced with another isolated soy protein product, SUPRO PLUS 219.

Accordingly, a cultured dairy product was prepared using the following ingredients: Weight percent of Protein source cultured dairy product Isolated soy protein product, SUPRO PLUS 10.66 219 Soy cotyledon fibres, FIBRIM (g) 1020 2.67 Soy lecithin Epikuron 100SP* 0.76 Water Distilled water 64.17 Milk composition Skimmed milk 16.58 Skimmed milk powder 3.98 Distilled water 0.72 Ca) gon@ 0. 087 Pectin LM-107 AS-YA 0.249 Agar-agar 0.062 Fermentation culture CH1 0.03 Enzobact 0.0435 Sum 100 The protein source is dispersed in distilled water having a temperature of 50°C and the dispersion is heated to 82°C and homogenised at 300 bar. In the meantime, the milk composition is prepared by dissolving Calgon@ in milk/water and then adding skimmed milk powder along with pectin and agar-agar. The homogenized dispersion

and the milk composition is mixed and heated to a temperature of 90°C for 5 minutes, cooled to 46°C and inoculated with the fermentation culture, CH1 (Chr. Hansen) and Enzobact (MediPharm). The fermentation proceeds for 6 hours and then the fermented mixture is homogenized at 30 bar, filled into small aseptic jars (0. 5-1 1) and cooled to 4-5°C.

A cultured dairy product prepared according to this example contains isolated soy protein product in an amount of about 10.7 weight percent corresponding to an amount of exogenously added soy protein of about 9.3 weight percent. When the amount of soy protein provided by the soy fibre product is included, the cultured dairy product contains exogenously added soy protein in an amount of about 9.6 weight percent.

The content of the final fruited product is as follows : Content per 100 g Cultured dairy product 83 g Fruit log Sugar 7 g Sum 100 g This fruited cultured dairy product contains isolated soy protein product in an amount of 8.8 weight percent corresponding to an amount of exogenously added soy protein of about 7.6 weight percent. When the amount of soy protein provided by the soy fibre product is included, the fruited cultured dairy product contains exogenously added soy protein in an amount of about 8.0 weight percent.

The cultured dairy product appears like yoghurt has a pH of 4.62, a lactic acid content of 1.34%, and displays no soy odour, a smooth texture without powdery mouthfeel and does not become bitter upon storing. A comparison of the tendency of the products prepared according to EXAMPLE 4 and the present EXAMPLE did not reveal any significant difference.

EXAMPLE 6 Preparation of a cultured dairy product containing exoaenoustv added soy protein heat inactivated Lactobacillus helveticus and hedrolysate.

A cultured dairy product prepared according to EXAMPLE 2 and 3, i. e. comprising addition of Enzobact@, was found to partially counteract the development of bitter taste during storage, and thus prolonged the shelf-life of the product. By further replacing the isolated soy protein product source SUPRA PLUS 161 with SUPRA@ PLUS 219 according to example 4 and 5 the products did no longer become bitter upon storing.

In order to estimate the effect of adding a hydrolysate on the development of bitterness in the product when using the isolated soy protein product SUPRA@ PLUS 161, a preparation of hydrolyse casein, Casamino acids (Difco) was added to the milk composition. Accordingly, a cultured dairy product was prepared using the following ingredients: Weight percent of Protein source cultured dairy product Isolated soy protein product, SUPRA@ PLUS 10.66 161 Soy cotyledon fibres, FIBRIMO 1020 2.67 Soy lecithin Epikuron 100SP* 0.76 Water Distilled water 64.17 Milk composition Skimmed milk 16.58 Skimmed milk powder 3.98 Distilled water 0.41 Di-sodium tartrate x 2 H20 0.15 Potassium sorbate 0.043 Pectin LM-107 AS-YA 0.249 Agar-agar 0.062 Hydrolised casein, Difco 0.20 Fermentation culture CH1 0.03 Enzobact 0.0435 Sum 100

The protein source is dispersed in distilled water having a temperature of 50°C and the dispersion is heated to 82°C and homogenised at 300 bar. In the meantime, the milk composition is prepared by dissolving diNa-tartrate in milk/water and then adding skimmed milk powder along with pectin, agar-agar and potassium sorbate. The homogenized dispersion and the milk composition is mixed and heated to a temperature of 85°C for 6 minutes, cooled to 46°C and inoculated with the fermentation culture, CH1 (Chr. Hansen) and Enzobact (MediPharm). The fermentation proceeds for 6 hours at 44°C and then the fermented mixture is homogenized at 30 bar, filled into small aseptic jars (0. 5-1 I) and cooled to 4-5°C.

A cultured dairy product prepared according to this example contains isolated soy protein product in an amount of about 10.6 weight percent corresponding to an amount of exogenously added soy protein of about 9.2 weight percent. When the amount of soy protein provided by the soy fibre product is included, the cultured dairy product contains exogenously added soy protein in an amount of about 9.6 weight percent.

The content of the final fruited product is as follows : Content per 100 g Cultured dairy product 83 g Fruit log Sugar 7 g Sum 100 g This fruited cultured dairy product contains isolated soy protein product in an amount of 8.7 weight percent corresponding to an amount of exogenously added soy protein of about 7.6 weight percent. When the amount of soy protein provided by the soy fibre product is included, the fruited cultured dairy product contains exogenously added soy protein in an amount of about 8.0 weight percent.

The cultured dairy product appears like yoghurt has a pH of 4.52, displays no soy odour, has a smooth texture without powdery mouthfeel and does not become bitter upon storing. A comparison of the tendency to become bitter, of the products prepared according to EXAMPLEs 4 and 5 and the present EXAMPLE, did not reveal any significant difference. EXAMPLE 7-Pilot scale preparation of new ISP's Table 1: Separation Scheme for Isolate Step Action Fractions Time (min) 1 Soy flour (40 kg) was dispersed in water (3001) at 20-10 to 21°C in Tank 1 with a portion of anti-foam FDP (75 ml). 2 The mixture was adjusted to pH 8.0 with 2 M KOH-5. 0 (-3. 61) 3 Extraction with continuous stirring (30 min. ) was 60 followed by holding without stirring (30 min.) 4 The slurry was centrifuge to remove solids with feed Fibre + # 90 rate of 726 ich and a pressure of 4.25 kg/cm2 into high Mwt. Tank 2. proteins The partially cleared slurry was re-centrifuged and returned to Tank1. The fibre-rich solids collected by the centrifuge were discarded. 5 Small ice pieces (48kg) were added to bring-5. 0 temperature # 7.0° C 6 The pH of the solution was reduced to 5.4 with 5.5 M-5. 0 HCI (-1. 7 I) 7 Precipitation was allowed to occur without stirring 90 8 Isolate was separated by centrifuging at 726 ich and ISP, # 90 a pressure of 4.25 kg/cm2 from Tank1 to Tank 2 and fraction 1 then back to Tank 1. The precipitate recovered from the centrifuge. Antifoam FDP (75 mi) was added during the separation. 9 Small ice pieces (12 kg) were added to the solution ~ 3. 0 to bring temperature- 10° C 10 The pH was reduced to 3.5 with 5.5 M HCl # 5. 0 (approximately 3. 71) 11 Precipitation was allowed to occur without stirring 60 12 The isolate was separated by centrifugation at a feed SP,-40 a rate of 726 I/h and a pressure of 4.25 kg/cm2 in a fraction 2 single pass from Tank 1 to Tank 2. The insoluble isolate was collected and the liquor of solubles discarded. 13 Both isolates were combined in a 251-vessel and-10 mixed with a Silverson. Samples were collected for microbiology, protein and moisture. 14 The pH was raised to between 6.0 to 6.5 with a-10 concentrated solution of KOH (415g in approx. 500 mi water). 15 Portions (6.0 kg) of the slurry were deposited in-60 freezing trays (1000mm x 495mm) lined with plastic sheets and frozen at-21 °C in walk-in freezer.

RESULTS The results from the analysis if the FM for moisture, fibre, ash, and protein content.

Table 2. Analysis of freeze-dried isolate Moisture Fibre Fibre dry Ash Ash Protein Protein Fibre (%) basis (dry (as is) (dry basis) Ash+Prot basis) Final 5.3 4. 30 4. 5 6. 64 7.0 77. 0 81. 3 92.8 mix FM

Physical analysis of free-dried isolate Differential thermal analysis Table 3: DSC data for flour, isolate and some commercial isolates Weight used in Enthalpy Peak 1 Enthalpy Peak 2 DSC pan (mg) (J/g dry protein) (J/g dry protein) Commercial) soy 46. 3 1. 48 5. 64 flour Soya isolate final 32. 5 1. 10 5. 41 mix A-FXP H0161 52. 8 0 D-XT 10 34. 6 0 The two graphs for soy flour and soy isolate FM are plotted on the same graph (Figure 1) with the same scale for the peaks. The difference in baseline due to sample mass has been reduced by shifting the flour graph downwards by 7.0 units.

As shown in the graphs depicted in Figure 2: two commercial available qualities of ISP from Du Pont Protein Technologies, the curves are without peaks, indicating that there is not any un-denatured globular protein structure left in the products.

The PAGE data showed strong bands for the 7 and 11 S proteins that were more pronounced than for the flour, supporting the DSC finding.

Table 4: Values of the peak areas shown in graph of densitometry in the soy flour and the FM isolate at equivalent protein levels for loading the PAGE.

Mol. weight 79. 5 71. 7 64. 6 43. 1 29. 3 14 9. 6 in Da Protein 7S 7S 7S 7S 11 S 11 S PEPTI DES type Soya flour 49 81 125 118 160 176 23 OD value Isolate FM 50 105 135 180 210 210 20 OD value OD is the optical density reading of the scanning device used to"read"the gels.

Solubility of isolated precipitated at 5.4 and 3.5

Table 5: Results of analysis of commercial isolates for % protein solubility Samples Total Soluble % protein, % protein, % soluble A FX H0 161 84.5 46.5 55 B Supro 760 84. 5 23 27. 2 C 661 87. 5 19 21. 7 D XT10 82 34 41. 5 E FX H0 159 77.5 40 51. 6 F 219 83 43.5 52. 4 G 219D 82. 56 44 53. 3 H LH (Bunge) 85 35. 5 41. 8 I NB (Bunge) 86. 5 24 27. 8 J Profam 940 (ADM) 84 28 33. 3 K Fibrim 1020 8.5 3 35. 3 L Supro ST 79 16 20. 3 M Supro 770LN 84.5 25 29. 6 N Supro XT34 80 22 27. 5 Isolates 82. 1 78. 8 96. 0 from Example 80.1 77.7 97.1 7 batches 18/20 The solubility of the isolates prepared according to this example were approximately 96-97% after freeze-drying showing no denaturation had occurred.

Table 6 : Selected values for comparison on total isoflavon level Identification Genistein Daidzein Glycitein Total as Total as Aglycon Glycoside mcg/g mg/g Commercial 770 654 13 1437 2. 44 soy flour used for preparing ISP ISP 667 641 94 1402 2, 38

From the above data it is clear that the process according to the present invention allows for the preparation of soy protein products retaining almost all the isoflavones present in the starting material. This is in line with the undenatured state of the soy protein as the isoflavones are associated with the interior of the proteins and would thus remain associated with the proteins when the globular structure is intact.

PAGE densitograms: The isolate was run at 3 concentrations of material added to the gel. In general terms the O. D. values are linear from 0.01 to 0.8 and it is unwise to use higher values. The most diluted sample gave the best separation of the peaks and the densitogram from this was consequently used for further analysis. This was for the isolate obtained by mixing 0. 025moi of the extract with 0. 375ml of Laemmli buffer.

The same was true for the densitograms for the soy flour. The lowest concentration of material added to the gel gave a better separation.

Table 7: 7 and 11 S protein content in Flour and ISP Product 7S US Other, soy proteins Flour, % 28 29 43 ISP, % 34 39 27 Table 8 : Composition of the basic components in 7S and 11 S of the ISP

7S US Components a'a p A B Amounts % of total prot. 9 12 13 19 20 The amounts of 7 and 11 S proteins are considerably increased in concentration during the processing. This is because the highest molecular weight proteins, the 15 S fraction will be separated out together with the insoluble fibres and much of the 2S fraction is more water-soluble and will be a part of the water solution and removed.

The SDS-PAGE data indicated that the second washing and precipitation of ISP at pH 5.4 would have contributed to higher yield of ISP, but also to an increased amount of 7S as this ISP fraction showed a different relation between 7 and 11 S than from the first precipitation. The result would most probably have been a result of more equal quantities of 7 and 11 S than shown in the Table 7 above.

Material and Equipment.

1. Two deliveries (1000 kg) of soy flour were purchased from Cargill as lots 1 and 2.

Lot 1 was processed as batches Nos. 14 to 31 and lot 2 as batches Nos. 32 to 40.

2. Reagent grade Hydrochloric acid (specific gravity 1.18) and 3. Potassium hydroxide (56. 1 g/mol.) was purchased from BDH and tap water was used to make the slurry.

4. A foam-depressing reagent, antifoam FDP, was obtained from Basildon Chemical Company Ltd. , Kimber Road Abingdon Oxon., OX14 1 RZ.

The table below shows the main data for the two batches of Soy Flour used as raw material for the process and the pilot scale production: Table 9. Soy flour-starting material Detail Name De-fatted soy De-fatted soy Provabis 200/80 Provabis 200/80 Batch number 820423 8198859 Production 18/04/02 18/04/02 Protein, % > 52 > 52 Fibre, % 3.0-3. 5 3. 0-3.5 Oil, % 0.7-1. 2 0. 7-1.2 Moisture, % < 10 < 10 Total plate count, In max. 5.2 max. 5.2 Enterobacteriacae, In max. 4.0 max. 4.0 Salmonella nil nil Granulation % < 6 < 6 Used in batches 1-13 14-40

Equipment 1. Centrifugal separators from Westphalia Separators, Model SA 7-06-476 with self- cleaning bowl and a set of 69 separating cones, and 2. A paddle mixer was used to make the flour/water slurry and for mixing during pH adjustment.

3. A Silverson mixer (without shearing element) model D, Silverson Machine Ltd, Waterside, Chesham, Bucks, HP5 1 PQ was used to mix the isolates during pH adjustment.

4. APV 454 litres capacity steam jacketed kettles were used as holding tanks.

5. A NORD, model SK 20R150U9OU433, CIP lobe pump was used to feed slurry to the separator.

6. An Edwards freeze Dryer Modulyo was used for small-scale drying of isolates.

7. Commercial Freeze Drying was performed with N° Three and N° Seven freeze dryers.

Freeze-drying Comment: Freeze-drying was chosen as a suitable method in this case. To avoid microbiological growth in all produced ISP in the established pilot process was frozen on trays immediately after production. Instead of having to defrost all material again with increased microbiological risk, we decided to use freeze drying in this particular case.

Freeze-drying is also a rather gentle drying method.

Generally all kind of drying methods can be used, but the drying conditions must in all cases be chosen carefully to avoid further denaturation. The standard spray drying method is preferred for full production scale operations. But in such cases it is an advantage not to dry to low water level as this might influence the dispersion and flavour properties of the final product.

Laboratory scale to obtain analytical values Samples were frozen at-21°C. Edwards freeze Dryer Modulyo was used for drying the isolate at lab scale. Samples were kept at-55°C and a vacuum pressure of 10-'bar (0. 1 mm Hg) was used. It took 4 to 5 days to dry even very small samples.

Commercial Freeze Drying: The process was undertaken in two dryers, N° Three and N° Seven designed and built by Commercial Freeze Drying Ltd. Both dryers were CFD. N° Three has 28 trays of approximately 1000mm x 495mm and N° Seven holds 102 trays of the same size.

The frozen product was received by Commercial Freeze Dry Ltd. was held in cold storage at-20°C. Clean trays were then lined and the product weighed onto them at 5 kg/tray. The trayed product was then held overnight at-20°C to stabilise and the freeze dryer was cleaned with Commercial Freeze Dry Ltd. normal cleaning procedures and cooled to-20°C ready for loading. After loading a vacuum was created in the freeze dryer and it was run at a vacuum pressure < 2 mbar. Since the product was heat sensitive, a heat profile was run which prevented the product from reaching a temperature of greater than +30°C. This considerably extended the drying time.

First batch of 140 kg of wet product was dried In ? Three dryer on 5th September through 7th September. Dried product was then ground with an Apex comminuting mill model 114 type S2 at a fast hammer rotation speed and a screen aperture of 0.125".

After examination of the powder sample, it was too coarse and to reduce the particle size further, the finished product screen aperture was reduced to 0. 107"for subsequent milling of the product. The product was then dispatched to CCFRA.

The remaining wet product was dried in five lots of 140 kg each in ? Three dryer and two lots of 510 kg each in N°Seven dryer. The dried product was then ground using 0. 107"screen and dispatched to CCFRA in sealed in plastic sacks for further processing. Grinding screens were restricted by concerns for the temperature sensitivity of the product. A fine screen necessary to obtain particles < 150 um caused a build up of heat in the product, so a larger screen was used.

EXAMPLE 8 Comparison of the clinical effect of ingesting undenatured soy protein according to the present invention or a commercial ISP with a high isoflavone content.

Two Isolated soy protein products A (SuproSoy) and B (Undenatured ISP) were studied with a placebo C (casein) in a randomized placebo-controlled trial according to GCP with respect to their lipid lowering effects.

Study details: The patients took for 8 weeks daily 25g of the 3 products in 2 dosages in the morning and in the evening. The products were dissolved in water. Intermediate visits occurred after 2 and 4 weeks.

Patients: 120 patients of both sexes (73 women and 47 men in the age of 32 to 70) were included if they fulfilled the inclusion criteria of 200-300mg/dl or 5.2-7. 8 mmol/i total cholesterol.

Statistics: For the primary (total cholesterol) and secondary parameters (LDL-and HDLcholesterol) the mean differences between the first and fourth visit were tested using the group comparison analysis of variance.

Dropouts: During the study the following dropouts occurred: Group A: 8 cases, group B: 10 cases, and group C: 11 cases; from those only partial results are available.

Therefore the analysis was done for the per protocol group; in addition a intention-to- treat analysis was performed.

Side effects : No severe side effects occurred.

Description of patients: 120 patients included; 91 finished per protocol.

Results : Of the protocol analysis : The mean age was 55.1 years Weight: the weight increase during the study was 0.2-0. 6 kg 1) total cholesterol : Difference visit 4 vs visit 1: A:-12. 8 mg/dl-5. 0% B:-24. 3 mg/dl-9. 4% C: + 1.1 mg/dl +0.4% Percentage changes Active vs Placebo : A-C:-5. 4% B-C:-9. 8% Significances: A: B 0.017 A: C 0.013 B: C 0.001 2) LDL cholesterol Difference visit 4 vs visit 1: A:-12. 3 mg/dl - 7. 5% B:-19. 4 mg/dl-11. 8% C:-5. 8 mg/dl-3. 6% Percentages changes Active vs Placebo :

A-C: 3.9 % B-C: 8.2 % Significances: A: B 0.081 A: C 0. 159 B: C 0.006 3) HDL cholesterol No significant changes occurred Summary: The new undenaturated ISP formulation has proven to be significantly more effective to reduce total cholesterol than SuproSoy ISP. LDL cholesterol was significantly lowered with the new undenaturated ISP, but not for SuproSoy, in 91 patients with hypercholesterolemia in 8 weeks in a randomized placebo-controlled trial.

The percentage improvement of total cholesterol of the new undenaturated ISP was 9. 8% compared to SuproSoy ISP with 5.4% ; an improvement of over 80%.

The improvement of LDL cholesterol was 8.2% for the new undenaturated ISP compared to 3.9% for SuproSoy, an improvement of over 100%.