WO2008013455A1

WO2008013455A1 - Means and methods for preventing elevated blood glucose levels in humans with animal antibodies produced in milk

Info

Publication number: WO2008013455A1
Application number: PCT/NL2007/050375
Authority: WO
Inventors: Herman Albert De Boer; Willem Thomas Fokkema; Irma Maria Caecilia Vijn
Original assignee: Lactive B.V.
Priority date: 2006-07-27
Filing date: 2007-07-27
Publication date: 2008-01-31

Abstract

The invention relates to the generation, production and use of binding bodies that are directed to enzymes that are normally present in the human intestine. Such binding bodies are preferably used for the treatment of and/or prevention of diabetes and other (related) diseases in numans, both healty people and persons at risk. The binding bodies are in general useful in the prevention of diseases related to postprandial high glucose peaks and swings in glucose levels, such as diabetes, metabolic syndrome and atherosclerosis. In a preferred embodiment the binding bodies are antibodies raised in mammals and collected from the mammary gland of said mammal. Binding bodies are preferably specific for a digestive enzyme of the human intestinal.

Description

Means and methods for preventing elevated blood glucose levels in humans with animal antibodies produced in milk

The invention relates among others to the fields of overweight and obesity related disorders, such as risks for development of Diabetes Mellitus type II; Metabolic Syndrome and the development of atherosclerosis and cardiovascular disease; endothelial dysfunction and related microvascular diseases

Overweight and obesity associated disorders are a growing epidemic that threatens both industrialized and developing countries. In the United States it has been calculated that 64% of the adult U.S. citizens are overweight or obese. The Surgeon General of the U.S. has designated obesity as the most important public health challenge of our time. Overweight and obesity does not only impose direct threats to the health of an individual, such as strain on the joints. Serious overweight and obesity condition also causes major indirect threats, as it brings various obesity-related disorders with it. Obesity-related disorders are for instance type II Diabetes, metabolic syndrome, hypertension, cardiovascular pathology, microvascular diseases, non-alcoholic fatty liver disease and fetal macrosomia. At present there is no effective cure available for preventing and controlling overweight and obesity and its associated disorders,

Overweight and obesity are major causative factors and predictors for the above mentioned diseases. Glucose excursions beyond the normal range of 4 to 7 mmol/1 is generally accepted to be the mediator of these diseases. Elevated glucose levels and especially repeated glucose peaks, after eating starchy meals, can have major detrimental effects on the arteries. Acute hyperglycemia can exert such deleterious effects on the arterial walls through mechanisms that include oxidative stress, endothelial dysfunction, activation of the coagulation cascade (Bonora et al 2002) and monocyte adhesion to endothelial cells. This also pertains to endothelia of the aorta (Azuma et al 2006). The cardinal features of metabolic syndrome, obesity and insulin resistance, are closely associated with a state of low-grade inflammation (Shoelson et al 2006; Hotamisligil 2006). In adipose tissue chronic overnutrition leads to macrophage infiltration, resulting in local inflammation which in turn predisposes to development of insulin resistance (Weisberg et al. 2003 and Xu et al 2003).

The major, and first in line, overweight and obesity associated disease is type II diabetes (TDM). TDM is characterized by a gradual decline in insulin secretion in response to starchy nutrient uptake. It is therefore primarily a disorder of postprandial glucose regulation. However, at present physicians rely on fasting glucose levels and glycosylated hemoglobin levels to guide management of their patients. Apart from being the historically accepted practice, these parameters are also much easier to obtain from patients then determining postprandial glucose levels because the transient postprandial glucose levels are strongly influenced by timing of blood sampling and the type and amount of food taken by the patients under scrutiny. In order to obtain an accurate picture a series of glucose data points need to be obtained. This makes comparison difficult and requiring standardization of food intake prior to seeing the physician among other practical problems.

Notwithstanding the traditional routine, it is not the fasting glucose levels that determine the damage but the postprandial glucose levels. By now it is becoming clear that fasting glucose levels are poor predictors for glucose levels at other times (Bonora 2002). Therefore, fasting glucose levels are poor predictors for risk of vascular damage. Leiter et al (2005) showed that about 33% of people diagnosed as having type 2 diabetes based on postprandial hyperglycemia have normal fasting glucose levels. Furthermore, more then 70% of the total glycemic load is caused by postprandial glucose peaks even in patients that are considered to be well controlled by anti diabetic medication. Furthermore, Leiter et al (2205) state that there is a linear relationship between the risk of cardiovascular death and the 2 hour oral glucose tolerance test which is a surrogate marker for postprandial glucose levels. Bonora (2002) showed that cardiovascular morbidity and mortality is independent of fasting glucose levels. The present invention provides methods and means for preventing and controlling elevated postprandial glucose levels and swings in blood glucose levels. This pertains especially, but not exclusively, to persons at risk for developing TDM, the overweight and obese humans, and patients that have already developed overt TDM. The present invention results: in a reduced risk of cardiovascular morbidity and mortality; in a lower glycosylated hemoglobin level and/ in a prevention of a further rise in HbAIc and, in case of pregnancy of overweight and obese women. The means and methods of the invention further allows to treat such women and at the same time prevent fetal macrosomia. Preferred means of this invention are antibodies which are specific for digestive enzymes operating in the human gut and which are obtained from mammals preferably from colostrums and milk for example from cattle. These specific enzyme specific antibodies are induced in the preferably the normal or post partum milk of mammals via immunization methods. A preferred but non-limiting example is antibodies against alpha-amylase.

Whereas certain compounds on the market are able to inhibit alpha- amylase, they do so in a nearly irreversible manner and also inhibit alpha- glucosidase. It concerns pharmaceutical compounds such as acarbose and miglitol which are on the market, and that are presently only administered via prescription. Wachters-Hagedoorn (2007) et al has demonstrated that low dose acarbose does not delay digestion of starch but reduces its bioavailability. They demonstrate that the cumulative amount of glucose, released from a starchy meal given to volunteers together with acarbose, and which was measured as a sequential series of glucose concentrations in serum, was reduced by 22%. This means that a fraction of the required alpha -amylase and alpha-glucosidase is irreversibly inactivated and that the remainder is not sufficiently available for the enzymatic conversion of all the starch supplied with the food. This effect was already evident with a low dose of acarbose (12.5 mg per person), an amount several fold less than the usually prescribed dose of 50 to 200 mg per person. Clearly, the corresponding remaining fraction of starch must be passed on to the colon. Because of the nearly irreversible nature of the binding to the enzyme, part of the starchy foods does enter the colon where they are a novel source and an abundant nutrient for starch fermenting bacteria causing fermentative processes leading to flatulence, gas formation, notably butyrate, and other disturbances in the colonic microflora and intestinal lining with unknown effects. This microflora is extremely complex as far as its inhabitants are concerned and the processes that they carry out. Most of these inhabitants are not cultivable and are only known by their DNA fingerprint. By consequence their physiological role in the gut is unknown as yet. Disturbances in microbial content and processes may also have hitherto unknown effects on the host for example on the nutrient and vitamin supply and unknown effects on e.g. the colonic epithelial lining. For example, Dehgan- Kooshkghzazi reported that the butyrate levels in serum treated with acarbose were elevated and that the caecal short chain fatty acid pool size was increased four-fold with an even larger increase for butyrate.

These side effects of acarbose may also pertain for small molecule natural compounds blocking alpha-amylase. For example, Morita et al reported such similar effects on colonic production of short chain fatty acid including butyrate in the distal colon of rats fed with psyllium polysaccharide. Although these alpha-glucosidase inhibitors are very useful products aiding in human health care, notably in regulating glucose release into the bloodstream and as such should be considered as compounds promoting health; they are associated with inconveniences arising from altered colonic processes. Clearly, compounds are needed that have similar or greater benefits to persons at risk for developing TDM and overt TDM patients, without the side effects just mentioned.

It is a further matter of this invention that the antibodies obtained from mammals and used orally in humans to bind to digestive enzymes such as alpha amylase, are proteins themselves and therefore subject to digestion by the in the gut residing proteases. By consequence these antibodies are, like other food proteins, gradually cleaved during passage of the ileum thereby slowly releasing the digestive enzymes they hold in their binding pocket. Upon gradual cleavage of the binding antibodies, the digestive enzymes will be released and can resume its enzymatic activity. It is a matter of this invention that, in contrast to the pharmaceutical compounds referred to above all starch will eventually be converted to glucose but in a slower and delayed process.

It is an object of the invention to spread out glucose release from the gut over a longer period of time after eating starchy foods. Reducing at least the rate of calorie absorption preferably results in a lengthening of the time span wherein calories, present in the food, are absorbed. In this aspect of the invention binding bodies against alpha amylase and related polysaccharide converting enzymes and enzymes converting oligosaccharides are preferably produced and used. It is preferred that an antibody of the invention is specific for alpha amylase.

The intestines take up calories by absorption of absorbable subunits of proteins, lipids and carbohydrates. Absorbable subunits are amino acids, short chain fatty acids and oligo- and mono-saccharides respectively. In a preferred embodiment said oligo- and monosaccharides comprise glucose molecules. Together they form the bulk of the calories that are taken up from foodstuffs. Absorbable subunits mainly become available to the body in the digestive tract. A digestive tract is composed of any organ that food optionally passes between entry and exit of the body. Additionally, all glands that open into the digestive tract and that are involved in the digestive process, such as the pancreas, are considered part of the digestive tract. Parts of the digestive tract that are of specific interest for the invention are the intestines. A method of the invention may be used for any animal. Preferably the animal is a mammal, more preferably a domestic animal, most preferably a human. The terms animal and individual as used herein, are interchangeable terms.

The digestion process in an animal body is aided by various digestive enzymes. A digestive enzyme as used in the invention is a proteinaceous molecule that promotes conversion of a polymeric carbohydrate or lipid from food into subunits. A digestive enzyme is preferably excreted into the digestive of said animal, preferably human. A digestive enzyme according to the invention preferably promotes conversion of a polymeric carbohydrate or lipid from food into absorbable subunits. In one embodiment the invention provides a method according to the invention, wherein said digestive enzyme is an endogenous enzyme secreted by said animal. Preferably, said digestive enzyme is an alpha-amylase; a poly-saccharidase and/or a glucosidase. In a particularly preferred embodiment said digestive enzyme produced by the animal itself, comprises an alpha-amylase. In an alternative embodiment of the invention said digestive enzyme is a lipase.

Inhibiting conversion of a polymeric carbohydrate or lipid from food can result in the maintenance of the undigested component as it was, or alternatively that the food component is converted into smaller units but not into the most basic units that can be taken up by the intestines. For instance, from polysaccharides into oligosaccharides instead of conversion of polysaccharides into di- or mono-saccharides as would have been the case in a situation without intervention. A subunit is any part that was present in the original food polymeric carbohydrate. Inhibited conversion can be, but does not need to be, a result of an altered location where the conversion predominantly or totally occurs.

A binding body specific for a digestive enzyme as used in the invention, is any component that can specifically bind tb the digestive enzyme. Binding of said binding body preferably at least in part prevents activity and/or reduces activity of said digestive enzyme. Prevention of activity means that the digestive enzyme will not catalyze conversion of a polymeric carbohydrate or lipid from food. When activity of the digestive enzyme is reduced, the enzyme catalyzes digestion of a polymeric carbohydrate or lipid from food to a lesser extent and/or in a less effective manner than would have been the case in a situation without intervention. In order to prevent and/or reduce activity of a digestive enzyme, the binding body for instance binds to an enzymatic active region of said digestive enzyme and/or to an activity regulatory region of said digestive enzyme. Additionally or alternatively the binding body causes digestive enzymes to aggregate with one another or with other particles thereby at least preventing exposure of substrate to the digestive enzyme. Another mechanism that causes inhibition of digestive enzyme activity by a binding body of the invention is that the binding body binds to said digestive enzyme such that the enzymatic active region becomes inaccessible for the environment and thus can't perform its enzymatic activity. Inaccessibility is for instance caused by conformational changes of the digestive enzyme.

The invention preferably uses proteinaceous binding bodies. In a preferred embodiment the invention provides a binding body specific for a polysaccharidase/ glycosidase. In another embodiment the invention provides a binding body specific for a lipase. A binding body according to the invention preferably comprises a protein molecule. A protein molecule is generally degradable and as such no burden for a natural environment. In the art many different specific binding bodies are available. In a particularly preferred embodiment said binding body comprises an antibody or a functional part, derivative and/or analogue thereof. Currently many different parts, derivatives and/or analogues of antibodies are in use. Non-limiting examples of such parts, derivatives and/or analogues are, single chain Fv-fragments, monobodies, VHH, Fab-fragments, or artificial binding proteins such as for example avimers, and the like. A common denominator of such specific binding bodies is the presence of an affinity region (a binding peptide) that is present on a structural body that provides the correct structure for presenting the binding peptide. Binding peptides are typically derived from or similar to CDR sequences (typically at least CDR3 sequences) of antibodies, whereas the structure providing body is typically, though not necessarily derived from or similar to framework regions of antibodies. In a preferred embodiment the invention thus provides a method according to the invention, wherein said binding body is an antibody or a functional part, derivative and/or analogue thereof.

There are many ways to produce an antibody. An antibody is in one embodiment of the invention artificially synthesized. Alternatively, an antibody is produced by an organism or cells of an organism in vivo or in vitro. One of the advantages of the use of antibodies in comparison with the use of other compounds is that antibodies are specific for the selected digestive enzyme wherefore these are produced. The antibodies will therefore not inhibit other enzymes when that is not desired. Furthermore, intact antibodies administered to an animal via the digestive tract do not enter the bloodstream of the animal. Side effects caused by entrance of the bloodstream are thus avoided. The antibody can be a mono-clonal antibody or a polyclonal antibody. An antibody of the invention is preferably produced in vivo. In a preferred method of the invention, an antibody is generated in an animal thereby producing a polyclonal antibody preparation. This can be done by raising an immune response to said digestive enzyme in said animal. Said animal is preferably a mammal. A mammal is preferably immunized by injection of an antigenic agent, thereby inducing an antibody response specific for the injected antigenic agent. Systemic immunization can be induced and/or complemented by other forms of immunization having an inducing and/or boosting effect and carried by according to methods familiar by those skilled in the art. Preferably, the antigens can be delivered via intramammary injection, or injection into the supramammary lymphnodes or intranasal delivery of antigens. Alternatively, or in addition to the above methods, novel methods are developed and are being developed by authors and owners of this patent application that lead to animal friendly immunization protocols. Antibodies are for instance produced in small mammals, such as a mouse, a rat or a hamster. In a further preferred embodiment of the invention, said antibody is secreted in milk of said mammal. As the antibody is secreted in the milk, the antibody is easily obtained and readily available for administration to other animals of the same or other species. Any milk producing mammal is included in this embodiment. A mammal that produces substantial amounts of milk is however preferred. Another preferred mammal is a mammal with suitable characteristics for housing in an industrial farming environment. Examples of preferred mammals are a goat, a sheep, a horse, a camel or related mammals and members of the bovine species. The invention in a preferred embodiment thus provides a goat, sheep or cow antibody according to the invention. Further, the invention provides a method according to the invention, wherein said animal is of a bovine species. A preferred member of a bovine species is a cow as these animals are domesticated and there is a lot of experience in keeping, and breeding, these animals for milk production. Therefore, the invention preferably provides a bovine antibody according to the invention.

In a preferred embodiment, a binding body of the invention is a polyclonal antibody. A polyclonal antibody preparation recognizes different epitopes on the digestive enzyme. This renders the methods of the invention at least more robust. The invention therefore provides a binding body according to the invention, wherein said antibody is a polyclonal antibody. The polyclonal antibodies of the invention have specificity for different epitopes of a digestive enzyme and/or have specificity for epitopes of different digestive enzymes. Polyclonal antibodies are preferably derived from different cell lines. In a preferred embodiment of the invention, the polyclonal antibodies are produced by immunization of an animal, preferably a mammal. Immunizing an animal with different antigenic agents reflecting the different digestive enzymes, induces production of polyclonal antibodies specific for a variety of digestive enzymes. A selected composition of antibodies is thus produced. Such a selected composition for instance comprises antibodies specific for all or almost all digestive enzymes that brake down carbohydrates to glucose. In one embodiment of the invention, IgG and IgA type polyclonal antibodies are produced. An advantage of use of polyclonal antibodies is that different antibody-digestive enzyme complexes degrade with a different rate. For instance, a functional half life, in the gastro intestinal tract of a human of a IgA-digestive enzyme complex will generally be longer than that of a IgG- digestive enzyme complex. Because of the difference in functional half life between the complexes, a biphasic release occurs. Within the same type of antibody, the degradation time of antibody-digestive enzyme complexes formed with this type of antibody varies as well. One of the factors influencing this degradation time is the specificity of an antibody for an enzyme. Independent of the type of antibody involved, antibody-digestive enzyme complexes take time to dissociate and to degrade, and thus part of the enzyme antibody complex will disintegrate and part will dissociate leading to a slow release of the enzymes. As a result of the aforementioned characteristics, a composition of polyclonal antibodies of mammals and particularly bovines comprise different types of antibodies that display a multiphasic and preferably biphasic slow release of enzymes. Within the gut, digestion of the antibody will take place which in turn affects the binding and dissociation properties of the antibody thereby affecting the rate of enzyme release as well. The invention in a preferred embodiment provides a composition, comprising polyclonal antibodies with an affinity specific for a digestive enzyme. It is preferred that the specific polyclonal antibody comprises at least two different Ig-classes. Preferably IgG and IgA.

In a preferred embodiment of the invention, antibodies of the secretory IgA type are induced in the mammal and induction of their transport to the milk compartment. In this embodiment the IgA is produced as a complex with its receptor; the secretory component (SC) which confers a greater degree of protection against proteolysis in the GI tract. This SC is a remnant of the IgA receptor which facilitates the transport of the dimeric IgA across the epithelial cells. The IgA receptor transports the IgA from the basolateral side to the apical side of the mammary gland and of other mucosal tissues. Due to the SC bound to IgA, the complex is much more stable in the environment of the human GI tract. Likewise, the complex when it is bound to polysaccharide degrading enzymes, be it from endogenous origin or from a bacterial source, will be more stable under these conditions. This increased stability of the IgA .SC type of antibodies over IgG type of antibodies bound to the enzyme has the advantage that it is released more slowly then its IgG counterpart. The enzyme will therefore be released further down the GI tract. The slower release is due to the binding of SC to IgA, conferring better protection against proteases, and due to the fact that IgA forms multivalent complexes with the enzyme also conferring an additional level of protection. Both factors cause a delayed and slow release of the antigen further down the GI tract at a moment where the IgG enzyme complexes have been dissociated already. Once bound to the antigen, it will degrade more slowly leading to a prolonged decay of polysaccharides and it will persist longer in the GI tract.

A preferred binding body of the invention is a binding body with an affinity specific for an alpha-glucosidase. In the art a few alpha-glucosidase inhibitors are available. Those inhibitors have disadvantages that are at least partially overcome by a method of the invention. One of the known inhibitors is acarbose, an oligosaccharide which is obtained from fermentation processes of a microorganism. Acarbose binds strongly to alpha-glucosidase and thereby causes undigested carbohydrates to end up in the colon. The undigested carbohydrates result in a change of local microflora and because of this in excess fermentation processes. An antibody binds reversibly to digestive enzymes. Furthermore, antibodies are proteins which are subject to digestion in the gastro intestinal tract. As a result, and for both reasons, the digestive enzymes are reversibly inhibited. Use of a binding body of the invention as indicated prevents a radical change of local microflora. Furthermore, side effects occur by entrance of the blood stream by acarbose where it is reported to block also tissue alpha-glucosidases. The production of acarbose in a fungus is expensive. Subsequent purification is needed and implies a difficult procedure. Acarbose inhibits only one enzyme that is involved in glucose formation, whereas binding bodies of the invention inhibit each selected enzyme, depending on the chosen specificity. Moreover, binding bodies of the invention are in one embodiment of the invention combined in a composition for inhibiting at least 2, preferably 3, 4, 5 or more enzymes involved in glucose formation. In a preferred embodiment said composition comprises at least two antibodies for different digestive enzymes, wherein at least one of said binding bodies is specific for a digestive enzyme that promotes conversion of a polymeric carbohydrate into subunits and wherein at least one other of said at least two antibodies is specific for a digestive enzyme that promotes conversion of a lipid into subunits. This embodiment allows further delay and/or spread of calorie uptake via the intestine. It is preferred that said binding body that is specific for a digestive enzyme that promotes conversion of a polymeric carbohydrate into subunits is specific for pancreatic amylase and that said binding body that is specific for a digestive enzyme that promotes conversion of a lipid into subunits is a pancreatic lipase. In a preferred embodiment the invention provides a composition comprising binding bodies specific for all or almost all, i.e. the main, digestive enzymes involved in glucose formation in a digestive tract of an animal.

Besides acarbose, some compounds from a plant have been found to be exerting alpha-glucosidase inhibition. Those compounds have the same disadvantages as the disadvantages above described for acarbose. This also concerns Miglitol. Another disadvantage is that these compounds concern prescribed drugs which are not likely to become a commonly used food additive for use in persons that are at risk for developing TDM but who are as yet healthy. The use of antibodies produced in milk of normal cows can, on the other hand, be easily used as a food additive because milk antibodies are already a common ingredient in all milk based foods and drinks and hence be made available for everybody.

In a preferred embodiment the invention provides means and methods for at least inhibiting enzymes involved in the formation of glucose in the digestive tract of an animal. In another preferred embodiment the invention provides means and methods for inhibiting glucose formation in the digestive tract, preferably the intestines, of an animal. The inventors have found that a method according to the invention prevents a plasma glucose peak. Glucose peaks are suspected to have a causative effect on the development of diabetes type II and glycation damage and swings in glucose levels is a suspected causative factor for the development of atherosclerosis. Therefore, the invention provides a method according to the invention, wherein conversion of carbohydrate is inhibited, for at least in part reducing the magnitude of a plasma glucose peak in the blood of said animal. At least in part preventing a plasma glucose peak means that the glucose level in plasma of the blood will not rise to maximal peak levels due to digestion of carbohydrates from the food. A peak in plasma glucose level is preferably prevented. Alternatively, peak plasma glucose levels will be reduced. Prevention and reduction are related and compared to values that are reached in case of digestion without intervention. The instance wherein the risk on high peak plasma glucose levels is highest is postprandial. Therefore, the invention provides a method according to the invention, wherein said plasma glucose peak is postprandial. Postprandial is defined as after intake of food, preferably a meal.

In a further preferred embodiment, the invention provides a composition, comprising polyclonal antibodies with an affinity specific for a polysaccharidase and/or glycosidase. Such a composition forms antibody- digestive enzyme complexes in the intestines wherefrom the enzymes are slowly released. One of the advantages of this slow release system is that glucose is slowly released from ingested food; a plasma glucose peak is thus at least in part prevented. Another advantage of the slow release system is that carbohydrates in food are slowly digested instead of not digested as is the case when digestive enzymes are blocked with compounds such as acarbose. If carbohydrates are not digested at all, fermentative processes in the colon are disturbed. In the case of acarbose treatment this disturbance gives all sorts of unwanted side effects such as cramps and flatulence. Such side effects are at least in part prevented by the use of binding bodies according to the invention.

In one embodiment the invention provides a food product, comprising a binding body according to the invention. A food product as used herein is defined as any food product suitable for consumption by an animal. Such an animal is preferably a human or a domestic animal. A food product is a product that is part of the usual daily diet of an animal, or alternatively is a product that is specifically designed to comprise the binding bodies according to the invention. In a preferred embodiment, the invention provides a food product according to the invention, wherein said food product is a dairy product. A dairy product is at least partially produced in a natural way. A dairy product is for instance part of a healthy diet for a human being. In a further preferred embodiment the invention provides a food product according to the invention, wherein said dairy product is milk. Milk is a preferred food product of the invention for several reasons. An antibody of the invention is for instance produced by immunization of a milk producing animal. The milk thus produced comprises binding bodies according to the invention and the milk is thus readily available to provide to man. Milk is furthermore a product with a GRAS-status, wherein GRAS stands for 'Generally Recognized as Safe'. Also, milk is a product that is tolerated by most human individuals. For instance, most young people, elderly persons and even hospitalized persons tolerate milk. Moreover, milk is considered to be a healthy constituent of the diet of a human individual. Milk comprising binding bodies of the invention is a source of protein for an animal. Milk is in a preferred embodiment processed into whey. In a further preferred embodiment antibodies, preferably immunoglobulin (Ig) A and G, are purified from the whey. Purification means that the resulting composition has a 50-100% immunoglobulin content, preferably an immunoglobulin content of at least 70%, more preferably at least 80%, most preferably at least 90%. The purified antibodies are in one embodiment comprised in a pharmaceutical composition.

A binding body of the invention is in one preferred embodiment comprised in a pharmaceutical composition. The invention therefore provides a pharmaceutical composition comprising a binding body according to the invention. A pharmaceutical composition comprises any composition for pharmaceutical purposes. A pharmaceutical composition optionally comprises additives such as stabilizers and/or colorants. Pharmaceutical compositions are for instance formulated as drinks, tablets or capsules. In a preferred embodiment the invention provides a pharmaceutical composition formulated as a capsule. A capsule is in one embodiment for oral administration. In this embodiment the capsule preferably comprises a coating that is resistant against stomach conditions, of which an important aspect is the low pH that is present in the stomach. A preferred capsule of the invention is thus acid- resistant. The invention also provides the use of a binding body of the invention for the manufacture of a pharmaceutical composition for reducing at least the rate of calorie absorption from food in the digestive tract of an animal. The invention further provides the use of a binding body of the invention for the manufacture of a pharmaceutical composition for at least in part preventing a plasma glucose peak.

A binding body of the invention is in one preferred embodiment used for reducing at least the rate of calorie absorption from food in the digestive tract of an animal. In a preferred embodiment the invention thus provides the use of a binding body according to the invention, a food product according to the invention, and/or a pharmaceutical composition according to the invention, for reducing at least the rate of calorie absorption from food in the digestive tract of an animal. A binding body of the invention is in another embodiment used for at least in part preventing a plasma glucose peak. In a preferred embodiment the invention therefore provides the use of a binding body according to the invention, a food product according to the invention, and/or a pharmaceutical composition according to the invention, wherein conversion of carbohydrate is inhibited, for at least in part preventing a plasma glucose peak in the blood of said animal. A binding body of the invention is in a further preferred embodiment used for reducing and preferably blocking the activity of said digestive enzyme. In a further preferred embodiment the invention provides the use of a binding body according to the invention, a food product (containing an ingredient prepared according to the invention) according to the invention, and/or a pharmaceutical composition according to the invention, for reducing and preferably blocking the activity of said digestive enzyme.

A method according to the invention at least in part prevents and/or reduces obesity and/or obesity related disorders. The invention therefore provides a method according to the invention, for at least in part preventing and/or reducing obesity and/or obesity related disorders. The invention specifically provides a method for at least in part preventing and/or reducing obesity and/or obesity related disorders, comprising administering a binding body according to the invention, a food product according to the invention, and/or a pharmaceutical composition according to the invention. Administration is preferably orally. In a preferred embodiment the invention therefore provides a method according to the invention, wherein said binding body, food product and/or pharmaceutical composition is administered orally. In an alternative embodiment of the invention, a binding body and/or a food product and/or a pharmaceutical composition is administered to the digestive tract by means of an alternative route.

A method of the invention reduces at least the rate of calorie absorption from food in an animal. A method of the invention causes some absolute calorie loss of the ingested food. A more important effect of a method of the invention is however, especially when a binding body specific for a digestive enzyme that is involved in glucose formation is administered, that the food is digested slower and thus the food becomes available to the body in a more gradual manner. As a result of this, the animal that has received binding bodies of the invention is inclined to eat less as the animal is triggered less often by a feeling of hunger. A preferred effect obtained with the provision of a binding body specific for a digestive enzyme that is involved in glucose formation such as alpha amylase; polysaccharidase/ glucosidase and/or glycosylhydrolase, is the inhibition of a plasma glucose peak. A least in part preventing a plasma glucose peak and specifically preventing hyperglycemia prevents development of micro- and - macrovascular complications. Further, a method of the invention aids to maintain glycemic control. Glycemic control is an important factor in the prevention of impaired glucose tolerance (IGT), also called prediabetes. Impaired glucose tolerance as well as obesity is main characteristics of the Metabolic Syndrome. IGT and the Metabolic Syndrome are often pre-stages of Diabetes Mellitus type II. A method of the invention is in one embodiment applied in an animal, preferably a human being, in order to prevent type II Diabetes. In case an animal has already reached a prediabetes stage, a method according to the invention at least in part prevents development of the disease from IGT to type II Diabetes. In a preferred embodiment the invention therefore provides a method according to the invention, wherein the obesity related disorders are Diabetes Mellitus type II and/or Metabolic Syndrome.

A method of the invention is particularly beneficial for animals with obesity or signs of obesity, because these animals are at risk for developing type II Diabetes. Humans preferably are provided with both an antibody specific for alpha-glucosidase and an antibody specific for a digestive enzymes excreted by a microorganism. Groups of human beings wherefore a method of the invention is particularly beneficial are for example youngsters with signs of obesity, adults at risk for developing type II Diabetes, Metabolic Syndrome patients, or type II Diabetes patients. Establishing whether an animal is at risk for developing prediabetes and/or diabetes requires the consideration and measurement of several factors. Those factors are for instance the weight and/or fat percentage of an animal in relation to the size of the animal. The manner in which the size of an animal is determined is dependent on the species. For a human being, the risk of developing prebiabetes and other overweigth related disorders is often measured by means of the Body Mass Index (BMI), although other methods to measure the relative fat percentage are also used. As a measure, BMI became popular during the early 1980s as obesity started to become a discernible issue in prosperous Western society. BMI provided a simple numeric measure of a person's "fatness" or "thinness", allowing health professionals to discuss over- and under-weight problems more objectively with their patients. It is meant to be used as a simple means of classifying sedentary (physically inactive) individuals with an average body composition. For these individuals, the current value settings are as follows: a BMI of 18.5 to 25 may indicate optimal weight; a BMI lower than 18.5 suggests the person is underweight while a number above 25 indicates that the person is overweight; a number above 30 suggests the person is obese (over 40, morbidly obese). For the present invention it is said that humans are at risk of developing prediabetes when their BMI index is above 25. Other factors in determining whether an animal is at risk for developing prediabetes and/or diabetes are for instance its genetic predisposition for obesity and/ or its genetic predisposition for Diabetes Mellitus type II. In a preferred embodiment of the invention, a method according to the invention is combined with other weight and blood glucose controlling measures. For the overall health of an animal a healthy lifestyle, including a diet with moderate amounts of food and sufficient physical exercise, is very beneficial. Therefore, a method according to the invention is preferably combined with a healthy lifestyle. Combining a method of the invention with other weight and blood glucose controlling measures, further generally leads to faster and longer lasting results.

The invention further provides a method for delaying calorie uptake from food in an animal, comprising providing the digestive tract of said animal with a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme in the digestive tract of said animal, thereby reducing the rate of conversion of polymeric carbohydrate from said food into subunits in said digestive tract. Delaying or slowing down the calorie uptake from food reduces the postprandial calorie peak and particularly the glucose peak in the bloodstream. The calorie uptake is smoothened out over a longer period. This reduces the chances of diseases associated with, in particularly, these phenomena. It also fits well with a healthy life stile, and can be included by healthy individuals in their life design. The total calorie uptake does not necessarily have to be reduced. It is preferred that the content of the digested food is not essentially altered upon entering the colon. Many different bacteria are present in the colon that are preferentially provided with a normal input. Essential changes in the input are associated with undesired side effects. Such side effects occur typically using irreversible digestive enzyme inhibitors such as acarbose. The invention further provides a method for reducing a post prandial glucose peak in the blood stream of an individual upon ingestion of a food comprising orally administering a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme essentially together with said food. The (proteinaceous) binding body preferably comprises an antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme, preferably an antibody obtained from normal or postpartum milk of a mammal. Further provided is a method for inhibiting the activity of a digestive enzyme in the digestive tract of an individual upon ingestion of food said method comprising orally providing said individual with an inhibitor of the activity of said digestive enzyme essentially together with said food said method characterised in that said inhibitor comprises an antibody obtained from normal or post-partum milk of a mammal. Inhibition of the enzyme with an antibody of a mammal results in a delay and/or slowing down of the calorie and preferably the glucose uptake. The mammal is a preferably bovine. The antibody is a polyclonal antibody. The polyclonal antibody specific for said digestive enzyme preferably comprises at least two Ig classes. Different Ig classes have different stabilities in the gasto-intestinal tract. This difference in stability results in differential breakdown of the antibody and thereby differential release of functional digestive enzyme in the intestine. This results in a more gradual calorie uptake. This reduces the postprandial glucose peak while leaving the total calorie uptake preferably unchanged. Thus preferably said at least two Ig classes have different stability in the intestinal tract, preferably in the small intestine. The polyclonal antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme preferably comprises an antibody of the Ig class IgA and an antibody of Ig class IgG. The IgG part in the antibody preparation provided acts predominantly in the first part of the calorie uptake process whereas the IgA part, being the more stable of the two, is also effective later in the digestive process. This allows a more gradual digestion of the polymeric substrates and thereby a more gradual uptake of the absorbable subunits. Said proteinaceous binding body is preferably provided at a dose of between about 0,1 to 5 g antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme per meal of about 700 kcal. Preferably said proteinaceous binding body is provided at a dose of between 0,2 to 2,5 g antibody per meal of about 700 kcal. These amounts are given for the total amount of antibody in the preparation, i.e. obtained from the milk. The antibody is thus preferably provided in the context of the other antibodies of the milk. In one embodiment, the antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme is purified from unspecific antibody (present in the milk). Purification, although typically not necessary, results in a more defined antibody preparation. When purified antibodies are used it is preferred that a dose is provided of between 0.05 to 0.25 g specific antibody per 700 kcal meal, preferably between 0,1 to 0,13 g specific antibody per 700 kcal meal. When antibody is formulated such that degradation in the stomach is prevented such that at least 80% of the antibody is active when entering the small intestine, it is preferred that the specific antibody is given at a dose that is between about 5 to 10 x lower than otherwise formulated. The dose for unpurified antibody can also be adjusted accordingly. In one embodiment, antibody is formulated such that degradation in the stomach is prevented such that at least 80% of the antibody is active when entering the small intestine. This can be achieved by providing the antibody with a buffer that adjust the stomach pH above a pH of 4. In a preferred embodiment said antibody is coated or enveloped into a delivery vehicle for protecting said antibody from degradation in the stomach and delivering and releasing said antibody in the small intestine. Providing said proteinaceous binding body preferably at least delays the calorie absorption from said food by said animal. Binding of the said proteinaceous binding body preferably at least in part inhibits reduces and or delays activity of said digestive enzyme. The digestive enzyme is preferably a polysaccharidase, a glucosidase and/or a glycosylhydrolase. In a particularly preferred embodiment said digestive enzyme is pancreatic amylase or pancreatic lipase. In a preferred embodiment said antibody preparation comprises an antibody specific for pancreatic amylase and an antibody specific for pancreatic lipase. The digestive enzyme is preferably a human digestive enzyme. Conversion of carbohydrate is preferably inhibited and or delayed, for at least in part preventing a plasma glucose peak in the blood of said animal.

The invention further provides an isolated and/or recombinant antibody of a goat, camel or bovine specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme. Also provided is a composition comprising normal or post-partum milk from a goat, camel or bovine comprising an antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme. The invention also provides a composition comprising an antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme obtained from normal or post-partum milk of a goat, camel or bovine.

The antibody is preferably administered orally. To allow good admixture with the food it is preferred that the antibody preparation or composition comprising the antibody is taken around the time of food ingestion. In a particularly preferred embodiment the antibody is taken or administered together with the food. To this end the invention further provides a food or feed product comprising a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme. Preferably said food comprises an antibody of a goat, camel or bovine specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme. The antibody can be provided to the food as such. However, it is preferably administered in the context of a food additive. This allows phasing of the food preparation process. The invention thus further provides a food additive comprising an antibody of a goat, camel or bovine specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme.

In a clinical setting, when used in the context of individuals suffering from or at risk of suffering from a disease the proteinaceous binding body can also be administered as a pharmaceutical. The pharmaceutical at least reduces a risk associated with obesity as mentioned herein above. Thus the invention further provides a pharmaceutical composition comprising a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme. The pharmaceutical composition is preferably formulated for oral administration. An antibody of the invention is preferably used as such. However, the antibody may be modified to accommodate desirable characteristics such as acid tolerance, or resistance, improved admixture of the antibody, solubility ect. In one embodiment the antibody is comprised in a delivery vehicle separating the antibody from the immediate environment. This is typically done to provide additional stability before use and/or during passage of the stomach. However, in one embodiment of the invention it is preferred that the antibody is readily available for interaction with the digestive enzyme upon exposure of the food to the digestive enzyme. In this embodiment it is preferred that the antibody not comprised in a delivery vehicle for delivering the antibody to the (small) intestine, as this typically delays the availability of the antibody. In one embodiment it is preferred that said delivery vehicle allows delayed delivery in the stomach. This allows for delivery of more intact antibody to the small intestine while simultaneously not preventing availability of the antibody upon contact with the digestive enzyme.

The invention further provides use of a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme, an isolated and/or recombinant antibody according to the invention, a composition according to the invention, a food or feed product or a food additive according to the invention and/or a pharmaceutical composition according to the invention, for reducing at least the rate of glucose release from food in the digestive tract of an animal.

The invention further provides use of a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme, an isolated and/or recombinant antibody according to the invention, a composition according to the invention, a food or feed product or a food additive according to the invention and/or a pharmaceutical composition according to the invention, for inhibiting conversion of carbohydrate or lipid into subunits. Conversion of carbohydrate is preferably inhibited for at least in part preventing a plasma glucose peak in the blood of said animal. The invention further provides a method of the invention for at least in part preventing and/or reducing disorders that are related to glucose peaks and swings in glucose levels in the blood of a mammal. The invention thus further provides a method for at least in part preventing and/or delaying disorders that are related to glucose peaks and swings in glucose levels in an individual, comprising administering to said individual a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme, an isolated and/or recombinant antibody according to the invention, a composition according to the invention, a food or feed product or a food additive according to the invention, and/or a pharmaceutical composition according to the invention. Said binding body, food product and/or pharmaceutical composition is preferably administered orally. For pharmaceutical purposes said individual is preferably suffering from or at risk of suffering from Diabetes Mellitus type II, Metabolic Syndrome, Impaired Glucose tolerance (IGT), Post-prandrial glycemia (PPG) and/or atherosclerosis.

The invention is further illustrated by the following examples. The examples are not to be interpreted as limiting the scope of the invention in any way.

Examples Example 1: production of anti-amylase specific immunoglobulins

Materials and Methods

For the production of anti-amylase specific immunoglobulin porcine pancreatic amylase (Sigma-Aldrich Inc., BioChemika) was used as antigen. Cows

(Holstein-Frisian) were intra-muscularly immunized with 0,5 mg amylase 4 ml PBS. Immunization was performed twice, 5 and 2 weeks before parturition. For continuous production of amylase specific immunoglobulin in milk cows can be immunized according to protocols as described by Lee, SH (WO 01/32713 Al). Cows were housed and maintained according to the generally accepted dairy management practices in the Netherlands. For the production of anti-amylase specific immunoglobulin used in this example colostrum was harvested and sweet whey was prepared by removal of the fat by centrifugation for 45 min at 4 °C, 11.000 rpm ((Beckman J2-HS, Beckman Coulter, Inc., Fullerton, USA). Casein was precipitated by addition of 10 ml rennet per liter fat-free colostrum followed by incubation for 2 hours at 37 °C (occasionally stirring is required). Precipitated casein is removed by centrifugation for 45 min 20⁰C, 11.000 rpm (Beckman J2-HS, Beckman Coulter, Inc., Fullerton, USA). After freeze drying the resulting powder was analysed for the presence of specific anti-amylase antibodies by a direct ELISA. Microtiter plates (Greiner nr 655902; Greiner Bio-One, Alphen a/d Rijn, The Netherlands) were coated o/n at 4 °C with 1 μg/ml porcine pancreatic amylase in carbonic acid buffer (pH 9, 5). After each incubation step the plates were washed six times with PBS containing 0,05% Tween. The plates were blocked with 2% skimmilk (Difco™) for 1 hour at room temperature (RT).

Samples were dissolved (50 mg/ml) and diluted in PBS and added in a two fold dilution series to the plate and allowed to incubate for 1 hour at RT. After washing anti-amylase specific antibodies were detected by using a sheep anti- cow IgG specific Horseradish Peroxidase labeled secondary antibody (AbD Serotec, Dusseldorf, Germany). Tetramethylbenzidine was used as the substrate for horseradish peroxidase. After incubation for 10 min at RT the coloring reaction was stopped with sulphuric acid (0,5M) and the optical density was measured at 450 nm (SpectraMAX 340, SLT labinstruments, Austria). Titers were expressed as fold dilution to obtain an OD450 = 1 compared to a standard that was set to 1. The standard was a commercially available sheep anti-amylase antibody (AbD Serotec, Dusseldorf, Germany) that was detected with a rabbit anti-sheep IgG Horeradish Peroxidase labeled secondary antibody (ITK Diagnostics, Uithoorn, The Netherlands)

Results

In table 1 the results are shown of five whey powders from five different cows immunized with porcine pancreatic amylase. As control, two different whey powders from non-immunized cows were analysed as well. The results show clearly the production of anti-amylase specific antibodies in the immunized cows. The powders with the highest titer (LA2902 and LA2905) were used in the inhibition assays as described in example 3 and 4. As negative control in these assays the batches LA2904 and LA2907 were used.

Table 1: anti-amylase titer of whey powder from immunized and non- immunized cows.

Example 2: production of anti-lipase specific immunoglobulins

Materials and Methods For the production of anti-lipase specific immunoglobulin porcine pancreatic lipase (Sigma-Aldrich Inc., BioChemika) was used as antigen. Cows (Holstein- Frisian) were intra -muscular Iy immunized with. 0,5 mg lipase in 4 ml PBS as described in example 1. Colostrum and milk was harvested and whey was produced as described in example 1. the freeze-dried powder was analysed for the presence of specific anti-lipase antibodies by a direct ELISA. Microtiter plates (Greiner nr 655902) were coated o/n at 4 °C with 1 μg/ml porcine pancreatic lipase in carbonic acid buffer (pH 9,5). After each incubation step the plates were washed six times with PBS containing 0,05% Tween. The plates were blocked with 2% skimmilk (Difco™) for 1 hour at room temperature (RT). Samples were dissolved (50 mg/ml) and diluted in PBS and added in a two fold dilution series to the plate and allowed to incubate for 1 hour at RT. After washing anti-lipase specific antibodies were detected by using a sheep anti-cow IgG specific Horseradish Peroxidase labeled secondary antibody (AbD Serotec, Dusseldorf, Germany). Tetramethylbenzidine was used as the substrate for horseradish peroxidase. After incubation for 10 min at RT the coloring reaction was stopped with sulphuric acid (0,5M) and the optical density was measured at 450 nm (SpectraMAX 340, SLT labinstruments, Austria). Titers were expressed as fold dilution to obtain an OD450 = 1. On each ELISA plate a standard sample was taken along for comparison and correction.

Results

In table 2 the results are shown of whey powders from cows either immunized with porcine lipase (LA2908) or non-immunized (LA2904). Although the whey powder of the non-immunized cow does react in the direct ELISA with porcine lipase, resulting in a titer of 44, the titer if the whey powder of the immunized cow is more than three times higher (156).

Table 2: Anti-lipase titer of whey powder from immunized and non-immunized cows.

Example 3: Inhibition of amylase activity by anti-amylase specific immunoglobulins - in vitro

Materials and Methods

The inhibition rate of amylase was tested with porcine pancreatic amylase (Sigma- Aldrich Inc., BioChemika) and the Phadebas® Amylase test (Phadebas, Lund, Sweden). A solution (50 mg/ml) of the whey powder containing the anti- amylase specific immunoglobulins was mixed in a dilution series with 62,5 ng amylase (300 U/L) in PBS. As a control whey powder from non-immunized cows were mixed in an identical dilution series with the same amount of amylase. In addition a positive control, PBS with only the amylase, and a negative control, only PBS, were treated identical. The amylase activity in these samples was measured according to the manufacturer's protocol. Briefly, the samples were incubated for 5 minutes in a waterbath at 37 ⁰C followed by addition of a Phadebas® tablet, immediate vortexing and placed back in the waterbath. All samples were incubated for exactly 15 minutes. The reaction was stopped by the addition of 1 ml 0,5 M NaOH. After centrifigation for 5 minutes at 1500 rpm (Heraeus Multifuge IS-R; Heraeus Instruments GMBH, Dusseldorf, Germany) the supernatants were measured in a spectrofotometer (UVmini 1240, Shimadzu, Kyoto, Japan) at 620 nm. The enzymatic activity was calculated using the standard curve as delivered by Phadebas. The activity of the positive control was set to 100%. The percentage activity was calculated by the formula:

% activity = AS (Units/L)/AC (units/L) xlOO

AS: activity sample (units/L) AC: activity positive control

Results

The influence of several whey powders on the amylase activity is shown in figure 1.

The whey powder from a non-immunized cow (LA2904) does not influence the enzymatic activity of amylase. The proteins present in the whey powder most likely stabilize the amylase, resulting in an activity higher than 100%, compared to the control in which no whey powder was added and of which the activity was set to 100%. When only whey powder was added and no amylase, no amylase activity could be detected (results not shown). When whey powders from cows immunized with amylase (LA 2902, LA2903, LA2905 and LA2906) were used a clear reduction in amylase activity was observed (see figure 1). This shows that these immunoglobulins can specifically reduce the activity of amylase; a crucial digestive enzyme involved in starch breakdown in the duodenum, and as such slows down the degradation of starch and thereby preventing or reducing the post-prandial glucose peak. As such these immunoglobulins can prevent or reduce the symptoms of diabetes.

To show that these immunoglobulins indeed can play a role in preventing or reducing the symptoms of diabetes we compared the inhibition of amylase by the immunoglobulins with the inhibition of amylase by acarbose (Glucobay™, Bayer Healtcare AG, Germany). Acarbose is a well known pharmaceutical belonging to the class of alpha-glucosidase inhibitors (including pancreatic alpha-amylase), and is being prescribed to diabetes patients to control their blood glucose level. In a similar test as described above acarbose was added in a serial dilution to the amylase solution (300 U/L) and amylase activity was measured. Figure 2 shows that 50 μmol of acarbose results in 90% inhibition of amylase activity (10% rest activity), while only 2,5 to 3 μmol of total IgGl is necessary to obtain a 90% inhibition. This shows that the immunoglobulins are 20 times more effective in inhibition of enzyme activity. When taking into account that of these added IgGl antibodies only 3-5% are anti-amylase specific (no IgGl specific purification step has been performed) the effectiveness of the immunoglobulins compared to acarbose grows even further up to 400 times. This shows one of the advantages of using cow antibodies for inhibition of digestive enzymes.

Example 4: Effect of anti-amylase immunoglobulins on the postprandial blood glucose levels in piglets.

Materials and Methods

Seven crossbred sows (Yorkshire x Landrace) with an initial body weight of 15 kg were used in this study. The animals were housed individually in metabolism cages and adapted for 5 days to the light-dark cycle (lights on from 05:00 to 22:00 h) and the feeding regime (from a pelleted commercial ad lib feeding to two times a limited soaked meal feeding). The average ambient room temperature was 2O⁰C. On day 6 the piglets were equipped with catheters in the left external jugular vein and one in the left carotid artery. Surgery was performed as described by Koopmans et al.2005 (Physiology & Behaviour, vol 85, pp 497-503, 2005). In the 5 days recovery period after surgery, the piglets were habituated to be fed first a small portion of 30 g with 30 ml water (feed:water ratio of 1:1), followed by the remainder of the feed. The piglets were fed a diet with 60% maize starch. The diet was mixed with water at a ratio of 1:4 (wt/vol). The diet was offered twice daily (07:00 and 15:00 h) at 2.6-fold maintenance requirements for metabolizable energy (ME). They were also habituated to the blood sampling procedure, during which their living area in the metabolism cage was reduced to about 0.35 x 0.90 m.

After the recovery period experimental days were performed, on which blood samples were taken at time points -1/2, -1/4, 0, 1/4, 1/2, 1, 11/2, 2, 3, 4, 5, 6 h of the meal. Pigs were fed either 300 gram of the feed with addition of 20 g whey powder containing specific anti-amylase immunoglobulins, with addition of 20 g whey powder containing only non-specific immunoglobulins or with addition of 25 mg acarbose (a crushed half tablet Glucobay^® 50, Bayer Healthcare, Germany). The acarbose was completely added to the first small portion of 30 g of the feed, while the whey powder was mixed to the total feed before the small portion of 30 g was extracted. Blood samples were taken from the arterial catheter unless this catheter failed, then the catheter in the jugular vein was used. Blood glucose concentration was analysed on a Precision Xtra™ blood glucose analyzer with Precision Xtra™ Plus blood glucose test strips (Medisense UK LTd, Abingdon, UK).

Results

The results of two representative piglets are shown in figure 3 and 4. Figure 3 shows an animal having a normal glucose response after being fed a meal containing non-specific immunoglobulins. When the same animal is fed a meal containing specific anti-amylase antibodies the blood glucose level does not rise after the meal, but follows a pattern as when the animal is fed a meal containing acarbose. Figure 4 shows an animal with a pre-diabetic postprandial glucose response after being fed a meal containing non-specific immunoglobulins. This animal received a 3x higher dose of specific immunoglobulins in the meal en showed after this meal a much lower (at least a 25% reduced) and a much shorter blood glucose peak. After 2 hours the blood glucose level was reduced to a normal 5 mmol/1, while after the meal with nonspecific immunoglobulins it lasted at least 6 hours to come back to this level. After a meal with the normal dose of specific immunoglobulins also a clear reduction in blood glucose level was observed (data not shown). These results show clearly that immunoglobulins containing specific anti-amylase antibodies can contribute in controlling post-prandial blood-glucose levels. In addition the results do not show the severe effect of acarbose. When animals were fed a meal containing acarbose their blood glucose level did not rise at all after the meal.

Example 5

In order to contribute to the solution of the problem of obesity and obesity-related disorders, a study will be performed on the inhibition of carbohydrate digestion. This study will concentrate on the generation of natural antibodies in serum and milk of cows. Subsequently the antibodies will be tested in several test-models for the ability to inhibit carbohydrate digestion.

Materials and methods Purified digestive enzymes that are involved in carbohydrate digestion will be obtained. The enzymes will at least comprise Alpha glucosidase, alpha amylase, and sucrose converting enzymes such as sucrase. Cows will be immunized with these enzymes using methods known to those skilled in the art and which can be further improved by the inventor. To further enhance the immunogenicity the enzymes or functional parts thereof, cell targeting devices and/or antigenic compounds will be attached. Preferred antigenic compounds are immune enhancing peptides. The resulting antigens will be formulated in microp article s. Part of the microparticles will comprise a selection of one or a few enzymes. Another part of the microparticles will comprise a collection of enzymes that covers the main carbohydrate digestive enzymes in a digestive tract.

Normal production cows, such as Holstein-Friesian*Zwartbont will be immunized with the antigen comprising microparticles. Additionally, Brahman cows and/or Shorthorn cows will be immunized. Alternatively Jersey cows are used. Part of the immunized cows will be immunized in the dry period, the other part will be immunized in the lactating period. The milk produced by the cows will be collected. Part of the milk will be processed into whey. Antibodies will be purified from the whey. Part of the antibodies will be purified to be comprised in a 90% Immunoglobulin (Ig) G and IgA composition. The composition will be formulated in at least two pharmaceuticals. Capsules will be formed, comprising a coating that is resistant to stomach conditions. Release will occur depending on the strength of the coating. At least part of the capsules will be formulated to release enzymes in the ileum. Another formulation will be a drink.

The antibody comprising products will be tested in several test-models. One of the models will be a rat model, for instance the rat model as described by Key FB and Mathers JC (1995). Furthermore a pig model will used. A suitable pig model for metabolic research is provided by ASG Lelystad, The Netherlands (Koopmans et al 2005). From this source, pigs are derived that are obese models, as well as models for IGT, Metabolic Syndrome and for Diabetes. Pigs are a very appropriate model for human beings as there are numerous anatomical, physiological and metabolic similarities. Another model will be provided by human volunteers. To all the test individuals, antibody comprising products will be provided. Digestive enzyme levels will be determined for species. The amount of antibody provided will be a 1:1 ratio of antibody over enzyme to start with. Later on the amount will be fine tuned. Human volunteers that will be included in the study in an earlier or later stage will be young people and adults with signs of obesity and patients with prediabetes or type II Diabetes. Antibodies will be administered approximately 2-6 times a day. In prediabetes or diabetes patients antibodies are preferably administered before, during and after each meal.

In order to facilitate regular sampling and reduce stress, part of the test-animals will be provided with permanent catheters and, if needed, stomas. Pigs will be sampled at the portal vein, the carotid artery or the jugular vein.

Results

Several parameters will be measured in the animal models. Inhibition of enzyme activity will be measured in samples derived from the intestinal tract. Such samples comprise faeces, samples obtained via surgery of animals. Glucose levels and levels of glucose derivatives in the blood will be measured at diverse points in time, as well as insulin levels in the blood. Further cholesterol and triglycerid levels in the blood will be assessed. Urine samples will be checked, for instance for glucose and protein content. Hepatic activity such as glucose en fat storage will be measured. The composition of the microflora in the test animals will be determined in several stages. Effects of treatment on the body weight and body composition will be assessed. The effects of treatment on the development of diabetes and other obesity-related disorders and, in patients, amelioration of existing diabetes and other obesity- related disorders will be measured in detail. Known obesity and diabetes related diseases will be monitored, such as cataract and vessel condition.

BRIEF DESCRIPTION OF THE FIGURES

Figure 1 is a graphic representation of the influence of whey powder from immunized cows (LA2902, LA2903, LA2905 and LA2906) containing anti- amylase specific antibodies (IgGl) and whey powder of a non-immunized cow (LA2904) on amylase activity.

Figure 2 is a graphic representation of the influence of whey powder from immunized cows (LA2902 and LA2905) containing anti-amylase specific antibodies on amylase activity compared to the influence of acarbose on amylase activity.

Figure 3 is a graphic representation of the blood glucose levels of a piglet being fed a starch rich meals supplemented with either whey powder from non- immunized cows (non-specific immunoglobulins), whey powder from immunized cows (specific immunoglobulins) or acarbose.

Figure 4 is a graphic representation of the blood glucose levels of a piglet being fed a starch rich meals supplemented with either whey powder from non- immunized cows (non-specific immunoglobulins), a high dose whey powder from immunized cows (high dose specific immunoglobulins) or acarbose.

1. References

1. Dehghan-Kooshkghazi M. and Mathers J. C, 2004. Starch digestion, large- bowel fermentation and intestinal mucosal cell proliferation in rats treated with the alpha-glucosidase inhibitor acarbose. Br. J. of Nutr 91:357.

2. Animal Sciences Group (ASG), 8200 AB LeIy stad, the Netherlands.

3. Leiter L.A. et al 2005. Postprandial glucose regulation: new data and new implications. Clin. Ther. 27 : S42-56

4. Bonora E. 2002. Postprandial peaks as a risk factor for cardiovascular disease: epidemiological perspectives. Int. J. Clin. Pract. Suppl. 129 : 5-11

5. Shoelson, S. E. et al 2006. Inflammation and insulin resistance. J. Clin. Invest. 116, 1793-1801

6. Hotamisligil, G.S. 2006. Inflammation and metabolic disorders. Nature 444, 860-867 7. Weisberg, S.P. 2003. Obesity is associated with macrophage accumulation in adipose tissue. J. Clin. Invest. 112, 1796-1808 8. Xu, H. et al Chronic inflammation in fat plays a crucial role in the development of obesity related insulin resistance. J. Clin. Invest. 112, 1821- 1830 9. Gonzales-Quintero 2007. The impact of glycemic control on neonatal outcome in singleton pregnancies complicated by gestational diabetes. Diabetes Care 30 : 467-470 lO.Azuma K. et al 2006. Repetitive fluctuations in blood glucose enhance monocyte adhesion to the endothelium of rat thoracic aorta. Arterioscler. Thromb. Vase. Biol. 10: 2275-2280 ll.Mita T. et al 2007. Swings in blood glucose levels accelerate atherogenesis in apolipoprotein E-deficient mice. Biochem. Biophys. Res. Commun. 358 (3): 679-685 12. Wachters-Hagedoorn, R.E. et al 2007. Diabetic medicine 24 (6):600-606.

Low dose acarbose does not delay digestion of starch but reduces its bioavailability.

13. Weaver G.A. 1997. Acarbose enhances human colonic butyrate production J. Nutr. 127(5):717-23.

14.Morita T. 1999. Psyllium shifts the fermentation site of high-amylose cornstarch toward the distal colon and increases fecal butyrate concentration in rats. J Nutr. 129(ll):2081-7 lδ.Wolin MJ. Et. al. 1999. Appl Environ Microbiol. 65(7):2807-12. Changes of fermentation pathways of fecal microbial communities associated with a drug treatment that increases dietary starch in the human colon. lβ.Koopmans S.J. et. al.2005 (Physiology & Behaviour, vol 85, pp 497-503).

Diurnal rhythms in plasma Cortisol, insulin, glucose, lactate and urea in pigs fed identical meals at 12-hourly intervals. 17. Key FB and Mathers JC 1995. Digestive adaptations of rats given white bread and cooked haricot beans (Phaseolus vulgaris): large-bowel fermentation and digestion of complex carbohydrates. Br J Nutr.

Sep;74(3):393-406

Claims

1. A method for delaying calorie uptake from food in an animal, comprising providing the digestive tract of said animal with a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme in the digestive tract of said animal, thereby reducing the rate of conversion of polymeric carbohydrate from said food into subunits in said digestive tract.

2. A method according to claim 1, wherein said calorie uptake comprises glucose uptake.

3. A method for reducing a post prandial glucose peak in the blood stream of an individual upon ingestion of a food comprising orally administering a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme essentially together with said food.

4. A method according to any one of claims 1-3, wherein said binding body comprises an antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme, preferably an antibody obtained from normal or postpartum milk of a mammal.

5. A method for inhibiting the activity of a digestive enzyme in the digestive tract of an individual upon ingestion of food said method comprising orally providing said individual with an inhibitor of the activity of said digestive enzyme essentially together with said food said method characterised in that said inhibitor comprises an antibody obtained from normal or postpartum milk of a mammal.

6. A method according to claim 4 or claim 5, wherein said mammal is a bovine.

7. A method according to any one of claims 4-6, wherein said antibody is a polyclonal antibody.

8. A method according to any one of claims 4-7, wherein said antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme comprises a polyclonal antibody consisting of at least two Ig classes.

9. A method according to claim 8, wherein said at least two Ig classes have different stability in the intestinal tract, preferably in the small intestine.

10. A method according to any of claims 7-9, wherein said polyclonal antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme comprises an antibody of the Ig class IgA and an antibody of Ig class IgG.

11. A method according to any one of claims 1-10, wherein said proteinaceous binding body is provided at a dose of between about 0, 1 to 5 g antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme per meal of about 700 kcal.

12. A method according to claim 11, wherein said antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme is purified from unspecific antibody.

13. A method according to any one of claims 1-12, wherein providing said proteinaceous binding body at least delays the calorie absorption from said food by said animal.

14. A method according to any one of claims 1-13, wherein binding of said proteinaceous binding body at least in part inhibits reduces and or delays activity of said digestive enzyme.

15. A method according to any one of claims 1-14, wherein said digestive enzyme is a polysaccharidase, a glucosidase and/or a glycosylhydrolase, preferably said digestive enzyme is pancreatic amylase or pancreatic lipase.

16. A method according to any one of claims 1-15, wherein conversion of carbohydrate is inhibited and or delayed, for at least in part preventing a plasma glucose peak in the blood of said animal.

17. An isolated and/or recombinant antibody of a goat, camel or bovine specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme.

18. A composition comprising normal or post-partum milk from a goat, camel or bovine comprising an antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme.

19. A composition comprising an antibody specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme obtained from normal or post-partum milk of a goat, camel or bovine.

20 A food or feed product comprising a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme.

21. A food or feed product according to claim 20, comprising an antibody of a goat, camel or bovine specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme

22. A food additive comprising an antibody of a goat, camel or bovine specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme.

23. Pharmaceutical composition comprising a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme.

24. A pharmaceutical composition according to claim 23, formulated for oral administration.

25. A pharmaceutical composition according to claim 23 or claim 24, wherein said proteinaceous binding body is comprised in a delivery vehicle for delivery of said binding body to the small intestine.

26. Use of a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme, an isolated and/or recombinant antibody according to claim 17, a composition according to claim 18 or 19, a food or feed product or a food additive according to any one of claims 20-22, and/or a pharmaceutical composition according to any one of claims 23-25, for reducing at least the rate of glucose release from food in the digestive tract of an animal.

27. Use of a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme, an isolated and/or recombinant antibody according to claim 17, a composition according to claim 18 or 19, a food or feed product or a food additive according to any one of claims 20-22, and/or a pharmaceutical composition according to any one of claims 23-25, for inhibiting conversion of carbohydrate or lipid into subunits.

28 Use according to claim 27, wherein conversion of carbohydrate is inhibited for at least in part preventing a plasma glucose peak in the blood of said animal.

29. A method according to any one of claims 1-16, for at least in part preventing and/or reducing disorders that are related to glucose peaks and swings in glucose levels in the blood of a mammal.

30. A method for at least in part preventing and or delaying disorders that are related to glucose peaks and swings in glucose levels in an individual, comprising administering to said individual a proteinaceous binding body specific for a carbohydrate digesting enzyme and/or a lipid digesting enzyme, an isolated and/or recombinant antibody according to claim 17, a composition according to claim 18 or 19, a food or feed product or a food additive according to any one of claims 20-22, and/or a pharmaceutical composition according to any one of claims 23-25,

31. A method according to claim 29, wherein said binding body, food product and/or pharmaceutical composition is administered orally.

32. A method according to any one of claims 1-16, 28-30, wherein said individual is suffering from or at risk of suffering from Diabetes Mellitus type II, Metabolic Syndrome, Impaired Glucose tolerance (IGT), Post-prandrial glycemia (PPG) and/or atherosclerosis.