FIELD OF THE INVENTION

The present invention relates to a method of producing protein con- taining extruded food products, such as extruded snack products or breakfast cereals. In particular, the present invention relates to a method of producing such extruded food products involving an enzyme treatment step with a cross- linking and/or a protein modifying enzyme. The present invention relates also to a feed composition comprising a protein component, a starch component and a crosslinking and/or a protein modifying enzyme. In addition, the present invention relates an extruded food product comprising a protein component, a starch component and a crosslinking and/or a protein modifying enzyme.

BACKGROUND OF THE INVENTION

Currently commercially available extruded food products, such as snack bars contain much sugar, fat and salt and little protein and fiber as well as calcium, other minerals and vitamins.

Supplementation and/or fortification of proteins, including also dairy- based proteins, in extruded snack products is known in the art. The fortification of proteins is known, however, to reduce the expansion of the extruded product and thus make it less crispy compared to a corresponding product which does not contain proteins (Onwulata, C.I., Use of extrusion-texturized whey protein isolates in puffered corn meal, J. of Food Processing and preservation 34, Supplement S2 (2010) 571 -586).

Patent publication WO 2012/036910 discloses a method for incorpo- rating protein into a puffed snack food product which method comprises mixing a protein component with starch, admixing dry components including an expansion controlling agent to the mixture, adding a water-based solution to the mixture to form an extrudate dough, and extruding the dough to form a direct expanded snack food product. It is disclosed in WO 2012/036910 that the pro- cessing conditions can be manipulated to increase expansion through the use of chelating agents to disrupt the matrix of the casein micelle and acids to lower the pH and impact the structure of the proteins. Alternative methods for producing extruded protein containing food products as well as means for fortifying extruded food products with protein supplements are required for fulfilling the increasing commercial need of extruded food products having balanced nutritional value. BRIEF DESCRIPTION OF THE INVENTION

An object of the present invention relates to a method for producing an extruded protein containing food product comprising an enzyme treatment step with a crosslinking and/or protein modifying enzyme.

An object of the present invention relates also to a feed composition comprising a protein component, a starch component and a crosslinking and/or a protein modifying enzyme.

An object of the present invention relates also to an extruded food product comprising a protein component, a starch component and a crosslinking and/or a protein modifying enzyme.

In addition, an object of the present invention relates to use of a crosslinking enzyme and/or a protein modifying enzyme in the manufacture of an extruded food product.

The objects of the invention are achieved by methods, products and uses characterized by what is stated in the independent claims. The preferred embodiments of the invention are disclosed in the dependent claims.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 shows the expansion and structure of extrudates containing 0% of whey protein, 15% of whey protein, and 15% of whey protein and transglutaminase (TG).

Figure 2 shows the effect of transglutaminase (TG) on the expansion and structure of extrudates having 15, 20 or 30% of casein. Extrudates without casein (RF), etxtrudates with casein (15Cas, 20Cas and 30Cas) and extrudastes with casein and treated with TG (15CasLTG, 20CasLTG and 30CasLTG). The addition of TG improved the expansion and made cell size smaller and/or thinner and evenly distributed.

DETAILED DESCRIPTION OF THE INVENTION

In food processing extrusion is a means of manufacturing a product by treating feed composition with pressure, temperature and screws and forcing it through an opening to form a shaped cooked product. Extrusion is accom- plished by single screw and twin-screw extruders. The choice of extruders is determined by the moisture content of the extrusion raw material mixture, for example. Food products produced by extrusion techniques are typically dry and the mouth feel of such products is crisp and chewable. On the other hand, a so-called co-extrusion technique has been developed, which produces simultaneously a crispy shell which contains a filling, such as jam, inside it. The product base produced by extrusion can be coated by desired components, such as chocolate, honey, coloring agents and/or dried cheese, using spray coating, pan coating or other coating techniques known to a person skilled in the art. Alternatively, flavor and/or color components can be impregnated into such extruded product base.

Extruded snacks and breakfast cereals have traditionally been produced using starchy fractions, such as corn meal, wheat flour, potato flour, rice or buckwheat. The addition of protein into the product reduces the expansion of the product and makes it less crisp compared with a corresponding starchy product not containing protein (Onwulata, C.I., J. of Food Processing and Preservation 34, Supplement S2 (2010) 571 -586).

The present invention is based on a finding that when a protein containing raw material was treated with a transglutaminase enzyme before and/or during extrusion, the expansion of the extruded products was increased and the products were crispier than the ones not treated with transglutaminase. Accordingly, using the method of the present invention, it is possible to produce protein containing extruded food products, such as snacks and breakfast cereals, which have better or at least equal textural properties than the current- ly commercially available extruded starchy products which do not contain protein. Further, nutritional quality of the extruded products of the present invention is better, because Maillard reaction, for example, is prevented by the enzyme treatment and thus lysine can be metabolized.

Accordingly, the present invention relates to a method for producing an extruded protein containing food product comprising an enzyme treatment step with a crosslinking and/or protein modifying enzyme. In the present method, the extent of expansion of an extruded protein containing product is increased and also the texture of such product is improved. In one embodiment of the present invention, the protein modifying enzyme refers to a protein de- amidating enzyme. In one embodiment, the method of the present invention comprises the steps of:

- providing a starch component and a protein component;

- mixing the starch component and the protein component;

- treating the protein component with a crosslinking and/or protein deamidating enzyme;

- processing the ingredients to provide an extruded protein containing food product.

In the present invention, the starch component and the protein com- ponent may be mixed together before treating the protein component with a crosslinking and/or protein deamidating enzyme. Alternatively, the protein component may be treated with a crosslinking and/or protein deamidating enzyme before it is mixed with the starch component. In the present invention, the enzyme treatment step with a crosslinking and/or protein deamidating en- zyme may be performed before extrusion step. Alternatively, the enzyme treatment step with a crosslinking and/or protein deamidating enzyme is performed during extrusion.

Accordingly, in one embodiment of the invention, the enzyme is added to a mixture of a protein component and a starch component, and then this mixture is extruded. Here the enzyme may be in powder form, for example. Hence, in this embodiment, the method comprises the steps of:

- providing a starch component and a protein component;

- forming a mixture comprising a starch component and a protein component;

- adding a crosslinking and/or protein deamidating enzyme to the mixture;

- processing the mixture to provide an extruded protein containing food product.

In another embodiment of the invention, the enzyme is added to a mixture of a protein component and a starch component during extrusion. Here the enzyme may be in liquid form, for example. Hence, in this embodiment, the method comprises the steps of:

- providing a starch component and a protein component;

- forming a mixture comprising a starch component and a protein component; - adding a crosslinking and/or protein deamidating enzyme to the mixture during processing the mixture to an extruded protein containing food product.

In a further embodiment of the invention, the protein component is subjected to enzyme treatment, the enzyme-treated protein is mixed with the starch component, and then the mixture is extruded. Hence, in this embodiment, the method comprises the steps of:

- providing a starch component and a protein component;

- subjecting the protein component to a treatment with a crosslink- ing and/or protein deamidating enzyme

- forming a mixture comprising a starch component and the enzyme-treated protein component;

- processing the mixture to provide an extruded protein containing food product.

In this embodiment, the protein component can be in liquid form, which after the enzyme treatment is dried into a powder form, which is then mixed with the starch component before subjecting the mixture to extrusion. Alternatively, the protein component is in powder form and it is dissolved in a liquid and then treated with the enzyme. This enzyme-treated protein is mixed with the starch component, and the mixture is extruded to provide an extruded protein containing food product.

The present invention relates also to a feed composition comprising a protein component, a starch component and a crosslinking and/or a protein modifying enzyme. In one embodiment, the protein modifying enzyme is a pro- tein deamidating enzyme. The feed composition is subjected to extrusion resulting in a protein containing extruded food product.

In addition, the present invention relates to an extruded food product comprising a protein component, a starch component and a crosslinking and/or a protein modifying enzyme. In one embodiment, the protein modifying enzyme is a protein deamidating enzyme. The extruded food product of the invention can be in a puffed or in a flake form, for example. In one embodiment, the protein content of the product of the present invention is high, i.e. at least 20% of the energy value of the food product is provided by protein. In another embodiment, about 9 - about 50% of the energy of the extruded prod- uct of the present invention comes from the protein. In one embodiment, the fibre content of the product is high, i.e. the product contains fibre in an amount of at least 6 g/100 g of the product or at least 3 g/100 kcal. The crosslinking and/or the protein deamidating enzyme treatment was found to improve the expansion and make the air cell size smaller/thinner and evenly distributed in the extruded product. The even distribution of air cells and smooth surface may have a positive impact on the flavor retention/release as well as on the coating of the extrudates. Thus, the products of the present invention have an increased extent of expansion and improved texture compared to an extruded protein containing product produced without a crosslinking and/or protein deamidating enzyme. Further, the protein containing extruded food products, such as snacks and breakfast cereals, of the present invention have better or at least equal textural properties than the currently commercially available extruded starchy products which do not contain a protein component.

In one embodiment of the present invention, the protein-modifying enzyme is a protein deamidating enzyme. Accordingly, in the present inven- tion, the crosslinking and/or protein-deamidating enzyme is a transglutaminase, and/or a protein glutaminase, for example. In one embodiment, the enzyme is transglutaminase. In another embodiment, the enzyme is protein glutaminase. In an even further embodiment, the enzyme is a mixture of a transglutaminase and a protein glutaminase.

Transglutaminases are a family of enzymes (EC 2.3.2.13) that catalyze the generation of covalent linkages between the glutamine and lysine amino acid residues present in the protein molecules. In the present invention, transglutaminase may originate from a microbe, yeast, mould, fish or mammal. There are several different commercially available transglutaminase enzyme preparations that are suitable for use in the process of the invention. These include Activa®YG (Ajinomoto, Japan) and Activa®MP (Ajinomoto, Japan).

Protein glutaminase catalyzes the deamination of protein bound glutamine, and glutamine is converted to glutamic acid. There are several commercially available protein glutaminase containing preparations, i.e. commer- cialized by Ajinomoto (Japan) that are suitable for use in the process of the invention.

Said enzymes may be used either alone or in combination to achieve the desired result. Optimum conditions for the enzyme treatment depend on the used enzyme, and they can be obtained from the manufacturers of the commercial enzymes. It was found in the present invention, that transglutaminase (TG) can be used to improve the structure of extrudates for better coating and flavor encapsulation. TG has effect on expansion and textural attributes of an extruded protein containing product. The addition of TG benefits also by providing uniform air cell (bubble) distribution and smooth outer surface that eventually improves the flavor perception. The enzyme is used in the present invention in an amount from about 0.1 U/g protein to about 100 U/g protein depending on the time point the enzyme is added to the process and/or the physical form of the enzyme. In one embodiment the amount of enzyme used is in the range of about 10 U/g protein - about 40 U/g protein. In another embodiment, the amount of enzyme used is about 10 U/g protein. In further embodiment, the amount of enzyme used is about 40 U/g protein. There are several stages of the process when the enzyme may be added into it. In one embodiment, the enzyme is added to a mixture of a starch and protein components before the extrusion. In another embodiment, the enzyme is added to the mixture a starch and protein components during extrusion process. In a further embodiment, the protein component is subjected to the enzyme treatment before it is mixed with the starch component. The enzyme may be used in powder form or in liquid form. In one embodiment, the liquid enzyme formulation is based on a pol- yol-water suspension. Polyols, such as glycerol, sorbitol, xylitol, mannitol or mixtures thereof can be used in such liquid enzyme formulation. The formulation may contain polyol(s) from 25% up to 100%, preferably from 50% up to 100% (w/w).

In the present invention, the protein component can be derived from any food grade protein source such as milk protein, pea proteins, plant proteins and/or cereal proteins or mixtures thereof. In one embodiment, the protein source is milk. The proteins of milk belong to casein and whey proteins. The major whey proteins in milk are beta-lactoglobulin and alpha-lactalbumin. Caseins which make up about 80% of the proteins in cow milk are divided into alpha-, beta- and kappa-caseins. In milk, casein exists in groups of molecules that are called micelles which consist of casein, calcium, inorganic phosphate and citrate ions. In the present invention, the milk-derived protein may be in liquid or in powder form. In an embodiment the protein source is milk, plant and/or cereal. In another embodiment, the protein source is milk and cereal, or milk and plant. In one embodiment, the protein component is derived from milk protein. In a certain embodiment of the present invention, the milk-derived protein is whey protein. The whey protein may be in the form of a whey protein concentrate (WPC), a whey protein isolate (WPI), a whey protein hydrolysate, or an ideal whey protein solution, for example.

The ideal whey protein solution of the present invention is prepared by microfiltering skim milk and concentrating the obtained microfiltration permeate by ultrafiltration. Accordingly, the ideal whey protein solution is an ultrafiltration retentate of the microfiltration permeate of skim milk. Microfiltration of the skim milk is typically carried out at a temperature of about 2°C to about 55°C. In an embodiment, the microfiltration is carried out at about 10°C. Ultrafiltration is typically performed at about 5°C to about 55°C. In an embodiment, the ultrafiltration is carried at about 10°C. The ideal whey protein solution contains β-casein. It may contain traces of fat, but it does not contain other micel- lar casein monomers or any other by-products from the cheese manufacture. Further, it is free of caseinmacropeptides (glycomacropeptide, GMP) and thermally formed k-casein β-lactoglobulin complexes.

The protein content of the ideal whey protein solution can range from about 4% to about 25%. In an embodiment, the protein content of the solution is about 9% and β-casein is about 20% based on total protein. The lac- tose content of the ideal whey protein solution can reduced, if desired. The lactose removal can be accomplished with methods known in the art, for example. The ideal whey protein solution can be used in the method of the present invention as a liquid. Alternatively, the ideal whey protein solution may be dried, by spray-drying for example, and used in extrusion process in powder form. In one embodiment, the milk protein is ideal whey. In another embodiment, the milk protein is ideal whey comprising about 20% β-casein is based on total protein.

In a certain embodiment of the present invention, the milk-derived protein is casein protein. The casein protein may be in the form of a casein concentrate, a caseinate or an ideal casein solution, for example.

In one embodiment, the casein protein is casein concentrate produced by membrane filtration of a milk raw material, such as skim milk.

For fractionation of casein from milk raw material, membrane filtration techniques, such as microfiltration, ultrafiltration, nanofiltration, reverse osmosis or their combinations, can be used. Microfiltration of the milk raw material is performed in such a manner that the milk raw material is concentrated by a factor of 1 to 4.5 times by volume, preferably 3.5 to 4.5 times by volume. The concentration factor (cf=K) refers to the ratio of the volume of the liquid fed to the filtration to the retentate, and it is defined with the following formula: K = feed (L) / retentate (L) (L = volume). The concentration factor of ultrafiltration is typically in the range of 1 to 10. In an embodiment, the concentration coefficient is 2 to 5. In one embodiment, the microfiltration is performed in a temperature below 20°C. In another embodiment the microfiltration is performed in a temperature range of 2°C to 20°C. In a further embodiment the temperature during microfiltration is in the range of 10°C to 14°C. The microfiltration of the milk raw material retains major portion of the casein in the retentate whereas a major portion of the whey proteins passes into the permeate.

The microfiltration may comprise a plurality of microfiltration steps. Different steps may comprise, for instance, changing of process conditions, such as filtration temperature, filtration pressure filtration and/or concentration factor of filtration, and/or filtration membranes. In one embodiment, the casein concentrate is obtained using a combination of microfiltration, ultrafiltration, nanofiltration and reverse osmosis.

In the membrane filtration, substances inhibiting the activity of a protein crosslinking enzyme are passed into the permeate while casein protein is concentrated in the retentate. The membrane filtration is preferably carried out, since the substances inhibiting a protein crosslinking enzyme are removed and the enzyme can act actively. The membrane filtration of the milk raw material provides a casein concentrate containing casein, lactose, and calcium- and phosphate-ions. The casein concentrate can be used in the method of the pre- sent invention as a liquid. Alternatively, the casein concentrate may be dried, by spray-drying for example, and used in an extrusion process in powder form.

In one embodiment, the casein protein is an ideal casein solution. The ideal casein solution of the present invention is prepared by microfiltrating skim milk and recovering the retentate obtained therein. Microfiltration of the skim milk is typically carried out at a temperature of about 2°C to about 55°C. In an embodiment, the microfiltration is carried out at about 10°C. The ideal casein solution can be used in the method of the present invention as a liquid. Alternatively, the ideal casein solution may be dried, by spray-drying for example, and used in an extrusion process in powder form. The ideal casein solu- tion as well as the corresponding powder contains immunoglobulins and lac- toferrin. The ideal casein solution as well as the corresponding powder, how- ever, contains only very small amounts of a-lactalbumin and β-lactoglobuline and in addition it contains less β-casein than normal milk. Without wishing to be bound by theory, during microfiltration, the structure of a casein micelle is disrupted due to the mechanical treatment, when β-casein loosens from the structure and it transfers with the whey protein. The disrupted micelles like this are more open and their surface structure is more porous, which enable an enzyme, especially transglutaminase, to enter the micelle. Additionally, trans- lutaminase inhibitor of milk goes to the permeate during microfiltration, and accordingly there is no need for heat treatment of the casein concentrate.

In one embodiment, the milk protein is ideal casein. In another embodiment, the milk protein is ideal casein which is essentially free of gly- comacropeptides (GMP),

In a certain embodiment, the casein protein is in the form of a ca- seinate. Caseinates, such as sodium caseinate, calcium caseinate and potas- sium caseinate, are soluble salts of casein. Caseinates provide outstanding nutritional properties, contain all of the essential amino acids, have a protein efficiency ratio (P.E.R.) of 2.5, and have a minimum protein content of 90% (dry solids basis). In one embodiment, the casein concentrate is calcium caseinate.

In a certain embodiment, the casein protein is in the form of micellar casein, acid casein, hydrolyzed casein or rennet casein, for example. Micellar casein is ultrafiltered casein extracted from milk without acidification. Acid casein is a dry free flowing high quality protein food ingredient that has been isolated from skim milk. Hydrolyzed casein is a soluble, enzymatic digest of casein. Rennet casein is produced by the controlled precipitation of casein from pure, pasteurized skim milk through the action of rennet.

In one embodiment, the milk-derived protein is a mixture of whey protein and casein protein.

The amount of protein included in the mixture of the protein and the starch components is up to about 60 weight-%. In one embodiment, the amount of protein included in the mixture of the protein and the starch components is up to about 15 to 60 weight-%. In one embodiment, the mixture contains protein up to about 30 weight-%. In another embodiment, the mixture contains protein from about 15 weight-% to about 20 weight-%. In one embodiment, the mixture contains milk protein up to about 30 weight-%. In another embodiment, the mixture contains milk protein from about 15 weight-% to about 20 weight-% In one embodiment, the mixture contains casein in an amount of about 20 weight-%. In another embodiment, the mixture contains whey protein in an amount of about 15 weight-%.

In the present invention, the starch component can be or can be derived from any kind of a cereal known to be suitable by a person skilled in the art or any mixture thereof. Accordingly, the starch component can be derived from any of oat, rye, wheat, barley, corn, buckwheat, rice, potato or any mixture thereof. Further, various treated starches, such as modified starches, are also useful in the present invention. In one embodiment, the starch component is derived from rye. In another embodiment, the starch component is a mixture of rye and wheat flours. In a further embodiment, the starch component comprises a mixture of oat and wheat flours and rye bran. In a further embodiment, the starch component comprises a combination of rice flour, modified starch and oat bran. The mixture of the protein and the starch components comprises the starch component from 40 weight-% to 98 weight-%, typically about 70 - 90 weight-%. In one embodiment, the mixture contains the starch component from about 80 weight-% to about 85 weight-%. In another embodiment, the mixture contains the starch component in an amount of about 80 weight-%. In a further embodiment, the mixture contains starch component in an amount of about 85 weight-%.

In one embodiment, the method of the present invention comprises a step of adding a liquid component to the feed mixture before or during the extrusion. The liquid component may be water, a carbohydrate containing syrup having brix 40-60, a polyol, and/or a salt solution, for example. The addition of the liquid component gives elasticity in the flakes produced by extrusion.

In the present invention, additional ingredients such as fibers (in the form of berry press cake, rye bran, oat bran, for example), can be mixed with the starch component and the protein component. In addition, berries and/or fruits as fresh, dried, frozen and or/in the form of a press cake, can be mixed with the starch and protein components to give taste and/or colour. Alternative- ly, such additional ingredients can be admixed with the starch-protein mixture in a separate step.

The method of the present invention may further comprise additional process steps such as cutting, milling, seasoning, coating and/or packing the extruded product. Depending on the type of the extruded food product, a per- son skilled in the art is able to choose the additional steps required for the manufacture of that specific product type. In one embodiment, the method comprises a step of drying the extruded protein containing food product. In one embodiment, the method comprises a step of coating the extruded protein containing food product. Suitable coating techniques are spraying and pan coating, for example. The extruded protein containing food product can be coated and/or impregnated with a flavour component, such as chocolate, honey, dried cheese and/or with a colouring, for example. Alternatively, such ingredients/components can be impregnated into the product. Accordingly, in one embodiment, the method of the present invention comprises a step of impregnating additional ingredients, such as flavour and/or a colouring component, for example, into the extruded food product.

The extruded food product of the present invention is a ready-to-eat food product, such as a snack product or a breakfast cereal. In one embodiment, the extruded food product of the present invention is puffed product. In another embodiment, the product is in the form of flakes. In one embodiment, the extruded food product of the present invention is a shelf-stable product. In one embodiment, the extruded food product of the present invention has high protein content. In another embodiment, the extruded food product of the present invention has high fiber content. In a further embodiment, the extruded food product of the present invention has high protein content and high fiber content. In one embodiment, the extruded food product of the present invention is gluten-free. In one embodiment, the extruded food product of the present invention contains a decreased amount of salt, sugar and fat. In addition, the extruded food product may be produced using the co-extrusion technique producing a crispy shell which contains a filling inside. The product of the present invention has an increased extent of expansion and an improved texture compared to an extruded protein containing product produced without a cross- linking and/or protein deamidating enzyme. Further, the protein containing extruded food products, such as snacks and breakfast cereals, of the present invention have better or at least equal textural properties than the currently commercially available extruded starchy products which do not contain a protein component.

In one embodiment, fibers from berries, fruits, cereals and/or potato, for example, may be added to the feed mixture. In one embodiment the feed composition and/or the extruded food product comprises fiber, minerals, vitamins or a combination thereof. In the present invention, processing the mixture to provide an extruded protein containing food product is performed using extrusion processes and equipment known to the person skilled in the art, such as twin screw or high temperature extruders. A person skilled in the art is able to choose and optimise the process parameters, such as temperature, pressure and screw speed, depending on the equipment used and/or the type of product produced. In the extrusion processes the temperature, pressure and screw speed may vary in the ranges of 20 - 140°C, 0.1 - 200 bar, 10 - 700 rpm, respectively.

The following examples are given to further illustrate the invention without, however, restricting the invention thereto.

Example 1 - Preparation of ideal casein and whey protein

Skim milk was microfiltered with polymeric microfiltration membranes of 800 kDa (Synder FR-3A-6338) at 10°C. The microfiltration was performed with a concentration factor of 16.5 to provide an ideal casein solution as a microfiltrate retentate. The ideal casein solution contains immunoglobulins and lactoferrin. It contains only very small amounts of a-lactalbumin (less than 0.05%) and β-lactoglobulin (less than 0.2%). The protein content of the ideal casein solution was 9.5%.

The microfiltration permeate obtained from the microfiltration was concentrated by ultrafiltration with an ultrafiltration membrane of 10 kDa (Koch HFK-131 6438-VYT) and with a concentration factor of 36 at 10°C to provide an ideal whey solution as an ultrafiltration retentate. The protein content of the ideal whey solution was 9%. β-casein content is about 20% based on total protein.

Example 2 - Preparation of extruded snack products

Test product was produced by mixing together endosperm rye (85%) and native whey protein in powder form (15%). Transglutaminase TG- WM (Ajinomoto) having activity of 155.7 U/g was added to this mixture in an amount of 15 U/ g whey protein. This mixture was extruded while adding water so that the total moisture of the feed was about 19%. The temperature profile of the extrusion was 120°C (out) - 90°C - 60°C - 50°C (feed), and the screw speed was 430 rpm.

A reference product 1 was produced in the same manner without the addition of transglutaminase powder. A reference product 2 was produced in the same manner from 100% endosperm rye without the addition of transglutaminase powder.

The diameter of the test product was 30% larger than that of the reference product 1 and 8.3% larger than that of the reference product 2. In addition, the test product was crispier than the reference product 1 and as crispy as the reference product 2.

Accordingly, the nutritional quality as well as the physical quality of the test product was better than that of the reference product 2, which did not contain whey protein.

Example 3 - Preparation of extruded snack products and breakfast cereals utilizing dairy proteins and berry side stream products

The materials used in the present example were:

Protein ingredients: casein and whey protein powders (Valio Oy).

Berry materials: bilberry berry press cake and frozen strawberries (Senga Sengana).

For extrusion purposes frozen berry press cake was dried in a malt drying equipment at 40°C and strawberries in a freeze dryer (Christ Epsilon 2- 25, Germany) and milled by a cutting mill (Retch SM300, 1000 rpm with 8mm sieve).

Enzymes: liquid TG (Valio) and powder TG (Ajinomoto YG and Wiberg GmbH). Carrier material: endosperm rye, corn, rice flour, modified starches.

Samples were extruded with a co-rotating twin-screw extruder APV MPF 19/25 (Baker Perkins Group Ltd, Peterborough, U.K.) to obtain directly expanded (125-1 15-90-80°C, 400 rpm) extrudates and flakes (90-90-90-80 °C, 250 rpm). However, temperature scales were lowered (to 120-100-60-50°C for balls or to 90-90-60-50°C for flakes) for the samples which were produced with transglutaminase to have optimal enzyme activity during processing.

Directly puffed extrudates were dried in an oven (Modell 800, Memmert GmbH, Schwabach, Germany) with recirculation air at 100°C for 15 min and the flakes at 100°C for 30 min to obtain crispy texture.

Determination of extrusion processing parameters and carrier matrix screening

Endosperm rye, corn, rice flour, modified starches as carrier matri- als were tested with various screw configurations (low and high shearing profiles) to obtain the best structure and texture. The screw design with less inten- sive shearing profile and shorter paddles gave the best structure. Initial samples were produced with 0-5-10-15-20-30% whey protein or casein in the feed mix. Increasing the protein content gradually decreased the expansion rate with thicker air cell walls and smaller air cells. Dark brown colour was also observed due to increased Maillard reactions. Textural meas- urements revealed that samples containing whey protein were crispier than the samples containing casein. After 15% protein addition there was a dramatic reduction in the crispiness values due to reduced expansion and closed structure. According to the results of a small sensory panel, casein could be added up to 20%, whereas whey protein could be added up to 15% of dry ingredients. However, whey protein was considered tastier than casein.

Use of transglutaminase (TG) to improve structure during extrusion

Increasing protein concentration decreased expansion, because protein started to form a network of its own and interfered with the melt rheolo- gy of starch. According to our hypothesis, an optimal size of inert protein parti- cles might interfere less with starch film. TG was fed at different levels (10 U/g protein and 40 U/g protein) from the water inlet of extruder. A challenge was the very short processing time which was giving less than 60 sec for the TG to react; this was adjusted by using only TG solution as a liquid source instead of water. Alternatively, powder TG can be used by evenly mixing it with the rest of the dry ingredients.

Both powder and liquid TG was used in levels of 10 U/g protein and 40 U/g protein. Compared to the control samples without TG, the TG treated samples (40 U/g of protein) had better expansion, as can be seen from Figures 1 and 2. The degree of expansion was highest at low T profile particularly for samples which had 15 - 20% protein and extruded with TG in an amount of 40 U/g protein. TG had effect also on the air cell size distribution and the structural appearance, as can be seen from Figure 2. Overall, the addition of TG improved the expansion and made the air cell size smaller/thinner and evenly distributed.

Liquid TG treatment during extrusion processing gave an even distribution of air cells and smooth surface which would have a positive impact on the flavor retention/release as well as on the coating of extrudates. The TG treated samples were less hard and crispier compared to the control samples without TG, based on the opinions of a sensory panel. Example 4 - Recipe and nutritional value of an extruded product according to the present invention

Recipe:

In addition, transglutaminase in an amout of 0.1 - 100 U/g protein is used.

Example of the nutritional value of the roduct er 100 :

About 9 - 50% of the energy of the product comes from protein.

It will be obvious to a person skilled in the art that, as the technology advances, the inventive concept can be implemented in various ways. The invention and its embodiments are not limited to the examples described above but may vary within the scope of the claims.