CN1620255A - Protein-containing foodstuff comprising a coss-linking enzyme and a hydrocolloid - Google Patents

Abstract

The present invention provides a composition comprising a hydrocolloid, and an enzyme, wherein the enzyme is a cross-linking enzyme and the hydrocolloid and enzyme are present in an amount to provide a dosage of the enzyme in a protein containing foodstuff of no greater than 20 U/g and a concentration of the hydrocolloid in the foodstuff of less than 1%.

Description

Protein-containing foods including cross-linking enzymes and hydrocolloids

The present invention relates to a composition comprising a hydrocolloid and an enzyme.

A number of hydrocolloids have been used for many years as food additives for gelling and stabilization. For example, carrageenan has been used as a gelling agent in puddings and other confections and to stabilize cocoa particles from settling in cocoa milk. Pectin has been used to impart texture to many products, one major use of pectin is in jams and jellies where pectin imparts strength to the gel. Pectin is also used in other foods, for example in fermented dairy products to increase gel strength and to increase viscosity and stability against whey separation. Research on the use of hydrocolloids in food is reviewed, for example, by Lapasin and Pricl (1995) in Rheology of industrial polysaccharides: Theory and Applications, Chapter 2: Industrial Applications of polysaccharides, Chapman & Hall, London, UK.

Enzymatic cross-linking of proteins is a somewhat new stabilization mechanism for protein-containing food products. Research in the last 10-15 years has shown many interesting applications for enzymatic crosslinking. However, studies to date related to enzymatic crosslinking have addressed the beneficial effects of protein crosslinking either by itself or in combination with other types of enzymes such as proteases.

Enzymatic protein crosslinking is an enzyme-catalyzed process that directly or indirectly joins proteins or peptides together through chemical bonds. Transglutaminases (TGases) are a class of enzymes with the systematic name R-glutamylamino-peptides: amine γ-glutamyl-transferases. These enzymes catalyze acyl transfer reactions between the γ-carboxamide group of the glutamine residue of the bound peptide as an acyl donor and various primary amines as acceptors. ε-(γ-glutamyl)lysine crosslinks are formed when the ε-amino group of a peptide-bound lysine acts as an acyl acceptor (Folk and Finlayson, 1977, ε-(γ-glutamyl) Lysine crosslinking and catalysis by transglutaminase, Advances in Protein Chemistry 31, 2-120). Furthermore, several oxidases such as amine oxidase, diamine oxidase and lysyl oxidase have been shown to induce protein cross-linking (Matheis and Whitaker, 1987, a review: enzymatic cross-linking of proteins applicable to foods. Journal of Food Biochemistry 11, 309-327). Typically this _occurs through the formation of _H2O2 which induces the formation of free _radicals (eg reactive quinines or aldehydes) within _the protein. As for these types of enzymes, some proteases are known to hydrolyze proteins in a specific manner, during which peptides are cross-linked by hydrophobic interactions (Otte, J., Ju, ZY, Faergemand, M., Lomholt, SB and Qvist, KB, 1996, Protease-induced aggregation and gelation of whey proteins. Journal of FoodScience 65, 911-915).

Enzymatic crosslinking has been shown to induce gelation and affect many functional properties of various food proteins including milk proteins. At acidic pH (Budolfsen, G., Nielsen, P.M., 1999, Method for production of an acidified edible gel on milkbasis, US 5,866,180) and by unacidified milk (Budolfsen, G., Nielsen, P.M., 1994, Method for production of a not acidified edible gel on milk basis, and use of such a gel, WO 94/21130) produces a milk gel based on cross-linked proteins. Moreover, several patents have used transglutaminase for ice cream (Motoki, M., Atsushi, O., Nonaka, M., Tanaka, H., Uchio, R., Matsuura, A., Ando, H. , Umeda, K., 1992, transglutaminase, US5,156,956; Yuzo, O., Kazuyoshi, M., Takahiko, S., 1993, the preparation method of low-calorie ice cream, JP 5091840A2; Hirobumi, M., Isao, K., 1994, Method for Improving Ice Cream Quality, JP 6303912A2.; Takahiko, S., Katsutoshi, Y., 1995, Preparation of Ice Cream, JP 7184554A2).

WO 99/29186 relates to a method of increasing the rate of digestion of a proteinaceous material which comprises treating the proteinaceous material with transglutaminase and mixing it with anionic polysaccharides.

US 5,156,956 relates to a method for the preparation of a protein gelled product comprising contacting a protein-containing solution or slurry with transglutaminase, which catalyzes the Ca2 ⁺ -independent gamma conversion of glutamine residues in peptides or protein chains - Acyl transfer reaction of the carboxamide group. The composition additionally includes a polysaccharide.

The present invention alleviates the problems of the prior art.

In one aspect, the present invention provides a composition comprising a hydrocolloid and an enzyme, wherein the enzyme is a crosslinking enzyme, and the hydrocolloid and enzyme are present in an amount such that the enzyme is present in a protein-containing The dosage in the food is not more than 20 U/g and the concentration of the hydrocolloid in the food is less than 1%.

In one aspect, the present invention provides a composition comprising hydrocolloid, protein and enzyme, wherein the enzyme is a cross-linking enzyme, and the dosage of the enzyme is not more than 20 U/g protein, and the concentration of hydrocolloid is less than 1%.

In one aspect, the invention provides a protein-containing beverage comprising a composition as defined herein and milk protein.

In one aspect, the present invention provides a method for the preparation of a composition comprising a cross-linked protein, the method comprising the steps of contacting the protein with a hydrocolloid and an enzyme; wherein the enzyme is a cross-linking enzyme and the enzyme is present in an amount of More than 20U/g, the concentration of hydrocolloid is less than 1%.

In one aspect, the present invention provides a use of a hydrocolloid and a crosslinking enzyme to synergistically form a gel in a protein-containing food product.

The term "cooperatively formed" refers to the formation of a gel with a higher complex modulus (gel stiffness) and/or a lower phase angle (as described herein) than would be predicted from the addition of hydrocolloid and crosslinking enzyme alone.

In one aspect, the present invention provides a use of a composition comprising a hydrocolloid, a protein and a crosslinking enzyme in the preparation of a food product selected from the group consisting of confectionary, acidified gel, drinkable protein-containing beverage and dough.

In one aspect, the present invention provides a use of a composition comprising a hydrocolloid and a cross-linking enzyme in the preparation of protein-containing ice cream, wherein the dosage of the enzyme is not more than 20 U/g.

It has surprisingly been found that there is a synergy between added hydrocolloids and enzymatic crosslinking in protein-containing foods.

It has been found that enzymatic cross-linking using hydrocolloids in combination with proteins unexpectedly imparts strong gelation to compositions (food or food fractions) containing proteins and hydrocolloids, and where either used alone has a much weaker gelling effect. gelling effect.

In one aspect, the present invention provides a use of a hydrocolloid and a crosslinking enzyme for the synergistic formation of a gel in a protein-containing cosmetic.

It has surprisingly been found that there is a synergy between added hydrocolloids and enzymatic crosslinking in protein-containing cosmetics.

It has been found that the use of hydrocolloids in combination with enzymatic cross-linking of proteins unexpectedly imparts strong gelation to compositions (cosmetics or cosmetic parts) containing proteins and hydrocolloids, and the use of either of them has a much weaker gelling effect. Gelation.

The term "protein" as used herein is equivalent to "polypeptide" or "proteinaceous".

The term "crosslinking enzyme" means that the enzyme catalyzes the direct or indirect crosslinking of proteins. Examples of protein crosslinking enzymes include transferases such as transglutaminase, oxidoreductases, and some proteases.

As used herein, the term "hydrocolloid" refers to molecules or multimolecular particles that are dispersed/dispersible in water or an aqueous solution. Hydrocolloids may include polysaccharides. Hydrocolloids cannot or only slowly pass through semipermeable membranes. Examples of hydrocolloids include carrageenan, starch, pectin, guar gum, alginates, locust bean gum (LBG), gellan gum, xanthan gum, carboxymethylcellulose (CMC), guar Gum, Acacia Gum.

In one aspect, the hydrocolloid is not a protein to be cross-linked or has been cross-linked.

preferred aspect

enzyme

In a preferred aspect, the enzyme is transglutaminase (TGase).

In one aspect, it is preferred that the dosage of the enzyme is no more than 20 U/g, preferably no more than 18 U/g, preferably no more than 16 U/g, preferably no more than 14 U/g, preferably no more than 12 U/g, preferably no more than 10 U/g, preferably Not more than 6.25U/g, preferably not more than 4U/g, preferably not more than 3.5U/g, preferably not more than 2U/g, preferably not more than 1.6U/g, preferably not more than 1.3U/g, preferably not more than 0.5U /g, preferably not more than 0.3U/g, preferably not more than 0.15U/g.

Enzyme activity is to measure (Folk and Cole, 1966, Mechanism of action of guinea pigliver transglutaminase, J.Biol.Chem.241,5518 with CBZ-L-glutamyl aminoglycine as the hydroxamate method of substrate -5525). As used herein, the "unit (U)" of enzyme activity is defined as 1 unit resulting in the formation of 1 M (mol) hydroxamic acid/minute at pH 6.0 and 37°C. U/g refers to enzyme activity/g substrate protein.

The specific activity of the enzyme preparation may be 100 U/g (enzyme activity/g product).

The specific activity of the enzyme preparation may be 100 U/g (enzyme activity/g product).

Hydrolyzed amine

In a preferred aspect, the hydrocolloid is selected from carrageenan, starch, pectin, alginate, locust bean gum (LBG), gellan gum, xanthan gum, CMC, guar gum, acacia gum and its combination.

In one aspect, preferably the concentration of hydrocolloid is less than 1%, preferably not more than 0.95%, preferably not more than 0.8%, preferably not more than 0.65%, preferably not more than 0.6%, preferably not more than 0.55%, preferably not more than 0.5%, preferably Not more than 0.45%, preferably not more than 0.4%.

All percentages given herein are by weight of all food items unless otherwise stated.

In one aspect, it is preferred that the concentration of hydrocolloid is not greater than 0.35%, preferably not greater than 0.3%, preferably not greater than 0.25%, preferably not greater than 0.2%.

In one aspect, it is preferred that the concentration of hydrocolloid is not greater than 0.15%, preferably not greater than 0.1%, preferably not greater than 0.02%.

In some respects, it is preferable

The concentration of hydrocolloid is not more than 0.3%, the concentration of enzyme is not more than 10U/g

The concentration of hydrocolloid is not more than 0.25%, the concentration of enzyme is not more than 6.25U/g

The concentration of hydrocolloid is not more than 0.2%, the concentration of enzyme is not more than 2U/g

protein

In a preferred aspect, the protein is selected from soy protein, milk protein, whey protein, flour protein, meat protein, and combinations thereof, or is present in, obtained from, or can be obtained from meat, meat fillings, and protein-containing beverages. its earned.

In one aspect, it is preferred that the dosage of protein is not greater than 90%, preferably not greater than 75%, preferably not greater than 50%, preferably not greater than 25%.

In one aspect, it is preferred that the dosage of protein is not more than 12%, preferably not more than 10%, preferably not more than 9%, preferably not more than 8%, preferably not more than 7.5%, preferably not more than 5%, preferably not more than 2.5%, preferably Not more than 2%.

A preferred protein is soy protein. In this regard, the preferred

· Hydrocolloid is carrageenan

The preferred concentration of carrageenan is not more than 0.5%

· Preferably the concentration of carrageenan is not more than 0.45%

·The concentration of carrageenan is preferably not more than 0.4%

The preferred concentration of carrageenan is not more than 0.3%

The preferred concentration of carrageenan is not more than 0.2%

• Preferably, the carrageenan is first contacted with the soy protein before the enzyme is contacted with the soy protein.

· Hydrocolloid is starch

·Preferably the concentration of starch is not more than 0.5%

· Preferably the concentration of starch is not more than 0.45%

·Preferably the concentration of starch is not more than 0.4%

· Preferably the concentration of starch is not more than 0.2%

Preferably starch and enzymes are contacted with milk protein at the same time

• Preferably, the enzyme is first contacted with the soy protein, followed by contacting the starch with the soy protein.

· Hydrocolloid is pectin

Preferable concentration of pectin is less than 1%

·Preferably the concentration of pectin is not more than 0.5%

- Preferably the concentration of pectin is not greater than 0.2%.

Preferably, the enzyme is first contacted with the soy protein, and the pectin is then contacted with the soy protein

· Preferably pectin is contacted with soy protein followed by heat treatment of the mixture (e.g. 80°C for 15 minutes) before enzyme is contacted with soy protein

Preferably the protein is milk protein. In this regard, the preferred

· Hydrocolloid is carrageenan

Preferable simultaneous contact of carrageenan and enzymes with milk protein

• Preferably, the carrageenan is first contacted with the milk protein before the enzyme is contacted with the milk protein.

· Hydrocolloid is starch

Preferably the starch is contacted with the milk protein, followed by heat treatment (eg 80°C for 15 minutes) of the mixture, after which the enzyme is contacted with the milk protein

Preferably starch and enzymes are contacted with milk protein at the same time

· Hydrocolloid is pectin

·The concentration of pectin is preferably less than 1%, and the concentration of enzyme is not more than 10U/g

·Preferably the concentration of pectin is not more than 0.5%

·Preferably the concentration of pectin is not more than 0.2%

The preferred enzyme concentration is not more than 5U/g

The concentration of the preferred enzyme is not more than 2U/g

Preferably the protein is whey protein. In this regard, the preferred

・The hydrocolloid is carrageenan.

- Preferably the enzymes are contacted with the whey milk protein, after which the starch is contacted with the whey protein.

· Hydrocolloid is starch

• The enzymes are preferably contacted with the whey protein first, followed by the starch with the whey protein.

Preferably the protein is present in, obtained or obtainable from the protein-containing beverage. In this regard, the preferred

· Hydrocolloid is carrageenan

·The concentration of carrageenan is preferably not more than 0.04%, and the dosage of enzyme is not more than 10U/g

· Preferably the concentration of carrageenan is not more than 0.02%

The preferred dosage of enzyme is not more than 2U/g

The preferred dosage of enzyme is not more than 1.6U/g

The preferred dosage of enzyme is not more than 1.3U/g

· Preferably the composition also includes a flavoring agent

The preferred flavoring agent is cocoa solids

Preferred flavoring agents are selected from chocolate, strawberry, cranberry, banana, citrus, mango, lemon, lime, cherry, peach, pear, apple, pineapple or combinations thereof

As used herein, the phrase "protein-containing beverage" refers to a protein solution. For example, cow's milk, soy milk, and reconstituted milk; reconstituted milk is usually made from dry milk powder, (anhydrous) milkfat, and water

A preferred protein is gluten. In this regard, the preferred

The hydrocolloid is guar gum

The dosage of enzyme is not more than 0.3U/g

The dosage of enzyme is not more than 0.15U/g

method

In the methods of the present invention, enzymes, proteins and hydrocolloids may be provided individually or in combination. Where they are provided together, preferably the hydrocolloid and the enzyme are provided in a composition as defined herein.

Enzymes and hydrocolloids can be contacted with proteins in any order. They can be contacted with the protein simultaneously, the enzyme can be contacted with the protein first and then the hydrocolloid, or the hydrocolloid can be contacted with the protein first and then the enzyme. In some cases, the amount of hydrocolloid and/or enzyme can be split and the contacting can be a combination of the above.

other aspects

In other aspects, the present invention provides

• Use of hydrocolloids and crosslinking enzymes for synergistic gel formation in protein-containing food products.

• Use of hydrocolloids and crosslinking enzymes in the preparation of confectionery.

• Use of hydrocolloids and crosslinking enzymes for the preparation of yogurt or acidified confectionary products (acidified gel).

• Use of hydrocolloids and crosslinking enzymes in the preparation of protein-containing beverages, eg cocoa milk beverages, drinkable yoghurts, whey-based beverages.

• Use of a composition as defined herein for the preparation of ice cream.

In this regard, it is preferred that the hydrocolloid is carrageenan and the enzyme is TGase

• Use of hydrocolloids and crosslinking enzymes in the preparation of bakery products.

In this regard, it is preferred that the hydrocolloid is guar gum and the enzyme is TGase

• Use of hydrocolloids and crosslinking enzymes in the preparation of dough.

In this regard, it is preferred that the hydrocolloid is guar gum and the enzyme is TGase

• Use of hydrocolloids and crosslinking enzymes in the preparation of meat products.

• Use of hydrocolloids and crosslinking enzymes for synergistic gel formation in protein-containing cosmetics.

The phrase "acidified confectionery product" as used herein is comparable to the term "acidified gel" and/or the term "yoghurt".

The invention will now be described in more detail by way of example with reference only to the accompanying drawings, in which:

Figure 1 shows the gel stiffness of a sweet cream containing soy protein and carrageenan

Figure 2 shows the effect of increasing the dosage of carrageenan on gel stiffness (G*) and phase angle

Figure 3 shows the gel stiffness of sweet cream containing soybean protein, starch and TGase

Figure 4 shows the phase angle of sweet cream containing soybean protein, waxy corn starch and TGase

Figure 5 shows the gel stiffness (complex modulus) and phase angle of skim milk-based sweet cream containing carrageenan and TGase

Figure 6 shows the gel stiffness and phase angle of sweet cream containing whey protein, carrageenan and TGase

Figure 7 shows the gel stiffness and phase angle of sweet cream containing whey protein, waxy corn starch and TGase

Figure 8 shows the complex modulus (gel stiffness) of milk after acidification with GDL (glucono-delta-lactone) in a rheometer at pH 4.5

Figure 9 shows the gel firmness of acidified skim milk gels containing pectin and TGase

Figure 10 shows the effect of increasing pectin dosage on the gel firmness of acidified skim milk gels

Figure 11 shows the effect of different combinations of pectin and TGase doses on the gel firmness of acidified skim milk gels

Figure 12 shows the gel firmness of acidified skim milk gels comprising waxy corn starch and TGase

Figure 13 shows the gel hardness of acidified soybean protein gels containing pectin and TGase

Figure 14 shows the effect of pectin and TGase on the gel firmness of acidified soybean protein gel

Figure 15 shows the sedimentation measured as an increase in light scattering at the bottom of the sample

Figure 16 shows the sedimentation measured as an increase in light scattering at the bottom of the sample

Figure 17 shows the melting of the ice cream

Figure 18 shows the effect of guar gum and TGase on dough stability. The percentage of guar gum added to the composition is shown on the graph.

Figure 19 shows the extensibility curves from Kieffer Rig

Figure 20 shows the effect of guar gum and TGase on Kieffer Rig force. The percentage of guar gum added to the dough is shown on the graph.

Figure 21 shows the effect of guar gum and TGase on the Keiffer rig distance. The percentage of guar gum added to the dough is shown on the graph.

Figure 22 shows the effect of guar gum and TGase on Keiffer rig area. The percentage of guar gum added to the dough is shown on the graph.

Figure 23 shows processed cheese samples comprising combinations of alginate and/or Tgase

In all figures, when error bars are shown, the values shown are the mean of duplicate experiments - error bars refer to standard deviation unless otherwise stated.

The invention will now be described in more detail in the following examples.

Example

Materials and methods

The enzyme preparation used in the following examples is Ajinomoto Active VM (Ajinomoto, Japan), and the stated activity is 100 U/g.

The concept of synergy between hydrocolloids and enzymatic crosslinking was tested in the following 5 systems: (1) dessert model system, (2) acidified milk system, (3) cocoa milk model, (4) ice cream system and (5) Dough system.

In each system, the order of addition and the simultaneous addition of enzyme and hydrocolloid were tested without changing other parameters. Also, in some systems, experiments were performed to remove one or both of the enzyme and/or the hydrocolloid from the product.

Sweets Model System

The basic recipe for this model is as follows:

- Dissolve 37.5 g of soybean isolate (Supro XT12) in 453.5 g of demineralized water at 60°C within 30 minutes with stirring. It was then cooled to 40°C.

- To 50 ml of the above mixture was added 10 U/g TGase (Active WM, 100 U/g, Ajinomoto Co.), and the mixture was incubated at 40°C for 60 minutes.

• A mixture of 0.1 g of carrageenan (Grindsted ^™ Carrageenan CL 360) and 0.5 g of sugar was added to the above mixture under stirring.

• Heat the final mixture to 80°C and hold at 80°C for 10 minutes.

• After these 10 minutes, the final mixture was transferred to a StressTech controlled stress rheometer where gelling occurred during cooling from 80°C to 5°C at 1°C/min.

This was followed by incubation at 5°C for 60 minutes to measure gel aggregation.

Measurement setup on a StressTech rheometer

Measurement Type: Oscillation of Controlled Strain

Measuring system: C25, concentric cylinder

Strain: 0.005

Frequency: 1Hz

Temperature: 80°C-5°C, 1°C/min-60 minutes 5°C constant

Initial balance time: 300 seconds

The following parameters are derived from these rheological metrics:

G*: complex modulus or gel stiffness (Pa); it is a measure of the total resistance of the sample to small deformations.

Phase angle, δ: The phase angle describes whether the sample is predominantly solid (elastic) or liquid (viscous). A sample that is more elastic has a phase angle of 0°, while a liquid that is more viscous (such as water) has a phase angle of 90°. Preferably the phase angle is 45° or less.

Other dessert models tested soy (7.5% protein in these trials) for whey protein isolate (5% protein) or skim milk powder (3% protein). The enzyme level was kept constant in all experiments, the same as described in the examples above, while the carrageenan concentration (of the total product) was kept constant in all experiments.

Other dessert models (soy, whey or skim milk based) tested carrageenan exchanged for starch (waxy corn, 1%).

Acidified gelled products (e.g. yogurt molds)

The basic recipe for this model is as follows:

• Heat the skim milk to 80°C for 15 minutes. Then cool to 40°C.

Add 10U/g TGase (Ajinomoto), and incubate the sample at 40°C for 60 minutes

Dry blend 0.1% GRINDSTED® Pectin LC 710 with 0.5 g sugar and add to the milk mixture within 15 minutes under stirring

• Add 2% glucono-delta-lactone (GDL) and incubate the sample at 40°C for acidification. When the pH had dropped to 4.5, the samples were cooled and stored at 5°C overnight prior to assay.

In some experiments, gelation of acidified milk products was performed on a StressTech rheometer, where the samples were used after addition of GDL. The pH was lowered in parallel samples and the rheological determination was stopped at pH 4.5.

Measurement setup on a StressTech rheometer

Measurement Type: Oscillation of Controlled Strain

Measuring system: C25, concentric cylinder

Strain: 0.005

Frequency: 1Hz

Temperature: constant at 40°C

Initial balance time: 300 seconds

In other tests, samples were tested for resistance to stripping after overnight incubation at 5[deg.] C. using a texture analyzer to measure the macrodeformability of acidified gels.

Setup of the measurements on the texture analyzer

Measuring Mode: Measuring Compression Force

Probe: stripping device 35mm

Test speed: 1mm/s

Temperature: room temperature/25°C

Load unit: 5kg

Distance: 20mm

Trigger: Auto, 10g

Samples were also prepared in which pectin (0.1%) was replaced with starch (0.5%).

Beverage Model Containing Protein - Cocoa Milk

The cocoa milk model is prepared from the following basic recipe:

- Dissolve 37.5g skimmed milk powder, 30g sugar and 10g skimmed cocoa powder in 420g demineralized water under stirring at 60°C within 30 minutes. Then cool to 40°C.

・Add 10U/g TGase (Active WM, 100U/g, Ajinomoto Co.) to 50ml of the above mixture, and incubate the mixture at 40°C for 60 minutes

• A mixture of 0.02 g of carrageenan (GRINDSTED® Carrageenan CL 220) and 0.10 g of sugar was added to the above mixture with stirring.

·Heat the mixture to 80°C for 15 minutes

• Pipette the cocoa milk into Turbiscan test tubes, then cool to room temperature and determine the stability (settling or clarification) of the cocoa milk during storage at 5°C.

In some trials, soy isolate was used instead of skim milk protein.

Sedimentation during storage was determined by measuring backscatter at the bottom using a Turbiscan instrument (Formulation, France). This backscatter is a measure of the particle density (or size) of a particular layer in the sample (ie in the case of the bottom). At the bottom of the sample, there is an increase in backscattering due to the increased density of the particles as they settle.

ice cream model

Ice cream models are prepared with the following basic recipe:

Water 61.778%

Hardened Coconut Oil (Cocowar 31) 7.888%

Skimmed milk powder 11.173%

Sucrose 14.000%

Glucose syrup 4.211%

CREMODAN® SE 30* 0.6%

Vanilla Flavor NA U35035 0.300%

Colorant, a-160-ws 0.05%

In some trials:

CREMODAN® SUPER* 0.3%

GRINDSTED® Carrageenan ICF 0.05%

TGase (Activa MP) 0.25% (6.25U/g protein)

In some trials, CREMODAN® SUPER (0.3%) and GRINDSTED® Carrageenan ICF (0.05%) and/or TGase (0.25%) were used instead of the standard emulsifier/stabilizer blend (CREMODAN® SE 30). In some cases TGase was mixed with the standard emulsifier/stabilizer blend (CREMODAN® SE 30).

*CREMODAN® SE 30 is an integrated blend of food grade emulsifiers (mono- and diglycerides) and stabilizers (carrageenan, LBG, sodium alginate, guar gum).

*CREMODAN® SUPER is a mono-diglyceride derived from edible fully hydrogenated vegetable fats.

method:

1. Melt the fat at about 50°C

2. Mix the liquid components at 20-22°C

3. Combine the dry ingredients

4. Add vanilla flavor

5. Add colorant

6. Add the fat and heat up to 30°C

7. Sterilize at 78°C for 2-3 minutes

8. Homogenize at 78°C at optimum pressure based on fat percentage

9. Cool to 40°C in a homogenizer cooler

10. Add TGase and keep at 40°C for 45 minutes

11. Cool to 5°C in an ice-water bath

12. Aging overnight in ice water (1-2°C)

13. Stirring

14. Freeze with 60% excess at -2.8°C in a continuous freezing tunnel

15. Fill into 1 liter cup

16. Fill into molds and insert sticks to freeze

17. Freeze overnight at -30°C in a hardening tunnel

18. Store at -18°C

The ice cream was evaluated for melting by placing the ice cream on a net at a controlled temperature (20° C.) and determining how many melted ice droplets passed through the net.

Dough Modeling System

The dough was prepared with the following basic recipe.

Flour 10.0g

Salt 0.2g

Water 500 Brabender Unit(BU)

(BU is determined in accordance with AACC Law 54-21)

+ Enzymes and Guar Gum

The rheological influence of the dough was investigated by Doughometer testing followed by extensometer measurements using a Texture Analyzer with a Kieffer Rig.

The dough was mixed for 6 minutes at 26°C on a Dough Meter. (According to the AACC method, analyze the dough physical property tester curve and record stability and resting time.)

Place the plastic strips on the grooved substrate of the lattice. 15 g of dough samples (prepared) were placed on the grooved base of the grid. Place the top half of the grid over the sample and press down firmly until the two pieces are together. Remove excess dough from sides. The lattice containing the dough is clamped in the lattice press in a plastic bag at 34°C for 40 minutes; this cuts the sample into strips, relaxes the dough and prevents moisture loss. The dough grid is then removed from the press, the dough strips uncovered one by one, and the upper grid pieces are carefully slid over the grooved base plate as needed.

Test Setup: Carefully remove each plastic strip of dough with a spatula, being careful not to puncture, stretch, and deform the dough. Place the strip in the recessed area of the sample plate and insert the plate into the apparatus with the spring clamping down on the loaded clamp bar. Tension measurements on the Texture Analyzer were then started.

Sample results: Determination results obtained from about 8 dough samples (same preparation) yielded average peak force (g) and distance values (mm) (at the point of limit of elongation), and their respective coefficients of variation (C.V.): also evaluated Integral area of force x distance (g x mm).

Example 1-3 - Dessert Model

Embodiment 1-soybean protein system

Embodiment 1.1 soybean, carrageenan and TGase

A test was carried out to determine the gel stiffness of sweet cream containing soybean protein, 0.3% carrageenan, 10 U/g TGase/substrate protein.

The tests performed were

A: Add TGase first, add carrageenan after 1 hour

B: Add TGase and carrageenan together

C: Add carrageenan first, then add enzyme

D: Add only carrageenan

E: only join TGase

F: control.

A clear synergy between carrageenan and enzymatic crosslinking was observed in a soy-based confectionary model product. Figure 1 shows the results of experiments varying the order of addition. Clearly, the highest degree of synergy was obtained when the carrageenan was first reacted with the soy protein and then the crosslinking enzyme was added (test C). When the cross-linking enzyme was added before the carrageenan, the gel stiffness obtained was about 5 times lower than when these components were added in the reverse order. This suggests that the mechanism for the very high gel stiffness when using crosslinking enzymes and carrageenan is the possibility of protein-hydrocolloid network-crosslinking due to the reaction of carrageenan with soy protein to make the particles much larger so that when A network can be easily formed when a cross-linking enzyme is added.

For comparison, Figure 2 shows the effect of increasing carrageenan dosage

Embodiment 1.2 soybean, waxy cornstarch and TGase

Example 1.1 was repeated. Use starch (another ingredient used in many foods) instead of carrageenan.

The tests performed were:

A: Add TGase first, then add starch after 1 hour

B: Add TGase and starch together

C: Add starch first, then enzymes

D: Add only starch

E: only join TGase

A beneficial effect of adding starch together with a crosslinking enzyme was found - see Figure 3. Apparently, there is also a synergy between this enzyme and starch. In contrast to the experiments with carrageenan and cross-linking enzyme, the order of addition of starch had no appreciable effect. The reason this may be so is that there is less specific reaction between starch and protein than between carrageenan and protein. However, a synergy between the addition of starch and the crosslinking enzyme is evident, especially from a phase perspective - see Figure 4. Neither component formed a gel when added alone (at this dose). However, when added together, the phase angle was below 45°, indicating gel formation.

In contrast to the case with carrageenan, the interaction between soybean protein, starch and crosslinking enzyme seems to be slightly better (lower phase angle, see Figure 4) by adding TGase first and then starch. It is possible that swelling of the inert starch granules (enzymatically and forming a protein network) sterically prevents some crosslinking when the starch is added before the crosslinking reaction takes place.

Embodiment 2-milk protein system

Embodiment 2.1 milk, carrageenan and TGase

A test was carried out to measure the gel stiffness (complex modulus) and phase angle of a sweet cream mainly composed of skim milk and containing carrageenan and TGase. These tests are:

A: Add TGase first, add carrageenan after 1 hour

B: Add TGase and carrageenan together

C: Add carrageenan first, then add enzyme

D: Add only carrageenan

E: only join TGase

F: control.

In the milk-only micellar protein system, the effect of adding the cross-linking enzyme together with carrageenan was less pronounced - see Figure 5.

No positive effect on gel stiffness was observed when the two were added together; the highest gel stiffness was observed when carrageenan was added alone.

A synergistic effect on the phase angle (elasticity) was observed, where the addition of the cross-linking enzyme slightly lowered the phase angle (especially when added after the carrageenan; test C), thus forming a more elastic gel, but the total gel Reduced stiffness. Possibly, the negative effect on the stiffness of the crosslinked gel in this system is due to the formation of a coarser network. Even little cross-linking between casein micelles may produce very large particles that may disturb the particle network. Thus, a fractured, but strong network of lines can be formed.

Example 3 - Whey Protein System

Embodiment 3.1 whey, carrageenan and TGase

A test was performed to determine the gel stiffness of a whey protein-based confectionary model (5% protein) containing carrageenan and TGase.

The tests performed were

A: Add TGase first, add carrageenan after 1 hour

B: Add TGase and carrageenan together

C: Add carrageenan first, then add enzyme

D: Add only carrageenan

E: only join TGase

F: control.

Addition of carrageenan alone does not induce gelation (i.e. phase angle greater than 45°), but the viscosity of the solution (viscosity modulus, not shown) increases as shown by the complex modulus (which is the sum of elastic and viscous moduli) - See Figure 6. Addition of cross-linking enzyme alone had a similar effect (increased viscous modulus (not shown), but higher phase angle).

When both carrageenan and cross-linking enzyme were added, gelation was found and the overall gel stiffness increased considerably (approximately twofold) compared to carrageenan alone. Therefore, when only any one of the components was added, the overall stiffness (complex modulus) was found to increase due to the increase in the viscous modulus. However, when both components were added, a gel was formed and the gel stiffness increased compared to the control. Therefore, there is a synergy between the addition of these two components.

Embodiment 3.2 whey, starch and TGase

A test was performed to determine the gel stiffness of a whey protein based confectionary model (5% protein) containing waxy corn starch and TGase.

The tests performed were:

A: Add TGase first, then add starch after 1 hour

B: Add TGase and starch together

C: Add starch first, then enzymes

D: Add only starch

E: only join TGase

A curious synergy was found when starch was used in a whey based dessert model product - see Figure 7. When the crosslinking enzyme was reacted with whey protein (at 80°C) prior to the addition of starch, a 20-fold increase in gel stiffness was found to form significantly stronger gels compared to when either starch or crosslinking enzyme alone (Test A) was used , which is shown by a decrease in phase angle from about 22° (starch) or 62° (enzyme only) to 9° (using both). This synergy was especially observed when starch was added at 80°C after whey protein crosslinking (the benefit of adding TGase before starch was the same, although to a lesser extent observed in soy based systems - see Figure 4) .

Example 4-5-acidified gel

A test was carried out to measure the gel hardness of a dessert model containing mainly milk protein/soybean protein and containing hydrocolloid and TGase.

Embodiment 4-milk protein system

Example 4.1 - Milk, Pectin and TGase

In acidified milk products, such as yoghurt and acidified milk desserts, the use of stabilizers such as pectin imparts increased body, eg higher viscosity in stirred products and higher gel strength in set products. We used chemically acidified milk gel as a model for this product type.

In acidified skim milk, there was a clear synergy between the use of pectin and enzymatic crosslinking, as shown in Figure 8 (values shown are means of duplicate experiments - error bars represent standard deviation). While 0.1% content of pectin itself can moderately increase the gel strength by 10%, TGase can increase the gel stiffness by about 50% at 10U/g enzyme preparation. However, using the two in combination resulted in an increase in gel stiffness of more than 100%, suggesting that there is a synergy between pectin and enzymatic crosslinking in this system.

These results were corroborated by large deformation assays - assays often just about perception. The results of back extrusion on a texture analyzer for acidified milk gels are shown in Figure 9 below.

Figure 9 shows the effect of pectin and TGase on the gel firmness of acidified skim milk gels. Numbers represent the following trials:

1: No substance added

2: Add 0.1% pectin, then heat the milk at 80°C for 15 minutes

3: Add 0.1% pectin after heating the milk at 80°C for 15 minutes

4: Add 10U/g TGase after heat treatment of the milk (80°C, 15 minutes), then incubate at 40°C for 1 hour, then add 0.1% pectin

5: Add pectin and TGase together, then incubate at 40°C for 1 hour, then heat treatment (80°C, 15 minutes)

6: Add pectin, then heat treatment (80°C, 15 minutes), then add TGase, and incubate at 40°C for 1 hour

7: Heat treatment (80°C, 15 minutes), then cooling to 40°C, after which TGase was added and incubated at 40°C for 1 hour.

GDL was added as an acidifier to all samples after each treatment and these samples were incubated at 40°C until the pH dropped to 4.5. These samples were then cooled and stored overnight at 5°C prior to assay.

For comparison with the results obtained with the combination of pectin and cross-linking enzyme, the effect of increasing pectin dosage is shown in Figure 10. Figure 10 shows the effect of increasing pectin dosage on the gel firmness of acidified skim milk gels. No benefit on firmness was found, and at high pectin doses the gel became gritty in texture (as seen by the naked eye). Apparently, gel firmness cannot be increased by increasing pectin dosage.

The synergy between pectin and TGase in the yogurt model system was much more pronounced when the pectin dosage was increased to 0.2% - see Figure 11. Figure 11 shows the effect of different combinations of pectin and TGase on the gel firmness of acidified skim milk gels. Concentrations are shown on the graph. The samples were all prepared as sample 4 in FIG. 9 . The addition of only 0.2% pectin resulted in a decrease in gel firmness (see Figure 10), however when added together with Tgase, a strong synergistic effect was found when combined with 0.2% pectin and a small amount of 2.5 U/g TGase.

Example 4.2 - Milk, Starch and TGase

Trials were carried out with starch instead of pectin - weak synergy was found in this system, rather than strong synergy - see Figure 12.

The numbers in Figure 12 represent the following trials:

1: No substance added

2: Add 0.5% starch, then heat the milk at 80°C for 15 minutes

3: Add 0.5% starch after heating the milk at 80°C for 15 minutes

4: Add 10 U/g TGase after heat treatment of the milk (80°C, 15 minutes), then incubate at 40°C for 1 hour, then add 0.5% starch

5: Add starch and TGase together, then incubate at 40°C for 1 hour, then heat treatment (80°C, 15 minutes)

6: Add starch, then heat treatment (80°C, 15 minutes), then add TGase, and incubate at 40°C for 1 hour

7: Heat treatment (80°C, 15 minutes), then cooling to 40°C, after which TGase was added and incubated at 40°C for 1 hour.

GDL as an acidifier was added to all samples after each treatment and these samples were incubated at 40°C until the pH dropped to 4.5. These samples were then cooled and stored overnight at 5°C prior to assay.

The results ( FIG. 12 ) showed that starch itself resulted in a small increase in hardness, while the combination of starch and TGase resulted in higher hardness. TGase alone also increased firmness significantly, and the combined effect of starch and TGase appeared to be only slightly greater than their sum, and there was a beneficial effect of both only when TGase was added after the addition of starch (ie, gelatinization).

Embodiment 5-soybean protein system

In soy-based acidified gels, the synergy between pectin and TGase was less pronounced than in the milk-based system. However, due to the high protein content, these gels were very hard, and this may have affected the ability to distinguish between samples containing only TGase and those also containing pectin. A weak synergy can be seen in Figure 13, however.

This synergistic effect became more and more pronounced as the pectin level was increased to 0.2%, as shown in Figure 14. Figure 14 shows the effect of pectin and TGase on the gel firmness of acidified soy protein gels. These samples were prepared as described in FIG. 12 . The preparation of the gel with TGase and 0.2% pectin is as in sample 4 in Fig. 12 . Pectin at a dose of only 0.2% did not increase the firmness of the gels, however the gels formed with TGase were much stiffer than those formed with TGase alone.

Figure 13 shows the effect of pectin and TGase on the gel firmness of acidified soy protein gels. Numbers represent the following trials:

1: No substance added

2: Add 0.1% pectin, then heat the soybean solution at 80°C for 15 minutes

3: Add 0.1% pectin after heating the soybean solution at 80°C for 15 minutes

4: After heat-treating the soybean solution (80°C, 15 minutes), add 10U/g TGase, then incubate at 40°C for 1 hour, then add 0.1% pectin

5: Add pectin and TGase together, then incubate at 40°C for 1 hour, then heat treatment (80°C, 15 minutes)

6: Add pectin, then heat treatment (80°C, 15 minutes), then add TGase, and incubate at 40°C for 1 hour

7: Heat treatment (80°C, 15 minutes), then cooling to 40°C, after which TGase was added and incubated at 40°C for 1 hour.

GDL as an acidifier was added to all samples after each treatment and these samples were incubated at 40°C until the pH dropped to 4.5. These samples were then cooled and stored overnight at 5°C prior to assay.

Example 6 - Beverage Model Containing Protein - Cocoa Milk System

Cocoa milk is often stabilized with carrageenan to avoid sedimentation of the cocoa particles during storage. It was observed whether there might be a synergistic stabilizing effect between carrageenan and crosslinking enzyme in this beverage.

Experiments were carried out to determine sedimentation in beverages containing protein and containing cocoa solids, carrageenan and TGase.

The tests performed were:

1: control

2: Only 10U/g TGase

3: only 0.04% carrageenan

4: 0.04% carrageenan and 10U/g TGase.

5: only 0.03% carrageenan

6: Only 0.01% carrageenan

7: 0.01% carrageenan and 1.3U/g TGase

8: 1.3U/g TGase

9: Control

Under the typically used carrageenan concentration of 0.04%, the stability was reduced by adding 10 U/g TGase compared to the sample with only carrageenan added (as shown in Figure 15). Figure 15 shows the sedimentation measured as an increase in light scattering at the bottom of the sample. Doses of carrageenan and TGase are shown. Carrageenan was added to the mixed samples before TGase. Each curve represents 3 determinations. Tgase did not increase the stability of the beverage when added at only 10 U/g, while 0.04% carrageenan completely stabilized the cocoa milk compared to the control. The destabilizing effect of these two stabilizers when used in combination at this dosage illustrates a possible phase separation (minor syneresis) due to the formation of too strong a network in the cocoa milk.

However, when the carrageenan concentration was reduced to 0.01%, the addition of cross-linking enzyme at a low dose significantly improved the stability of the product. Figure 16 shows the sedimentation measured as light scattering increase at the bottom of the sample. Doses of carrageenan and TGase are indicated on the graph. Carrageenan was added to the mixed sample prior to TGase. Each curve represents 3 assays. As shown in Figure 16, the stability of the beverage was significantly improved compared to when either component alone was used. This illustrates the synergy between these two components in this product.

Example 7 - Ice Cream Model

A test was performed to measure the melting of ice cream containing carrageenan and TGase.

The tests performed were:

1: 0.6% CREMODAN® SE 30 (standard)

2: 0.6% CREMODAN® SE 30+6.25U/g TGase

3: 0.3% CREMODAN® Super + 0.05% GRINDSTED® Carrageenan

4: 0.3% CREMODAN® Super + 0.05% GRINDSTED® Carrageenan + 6.25U/g TGase

5: 0.3% CREMODAN® Super

6: 0.3% CREMODAN® Super+6.25U/g TGase

The graph (Fig. 17) shows that the 6 ice creams described above melted at 20°C. Ice creams 1 and 2 were standard ice creams made with a complete emulsifier-stabilizer complex (monoglycerides, carrageenan, guar gum, LBG, alginate) and adding TGase to ice cream 2. Clearly, ice cream 2 melts slower and less melts than ice cream 1.

Ice creams 3 and 4 were made using emulsifiers (CREMODAN® Super) and GRINDSTED® carrageenan (i.e. no other stabilizers); ice cream 4 had TGase added; again clearly this ice cream (ice cream 4) containing TGase melted Slow and melts less.

Ice creams 5 and 6 were made with emulsifiers but no stabilizers; ice cream 6 had TGase added. The use of TGase apparently did not reduce this melting.

There is thus a clear synergy between the added hydrocolloid (or complete emulsifier complex or carrageenan (without other stabilizers)) and TGase.

Example 8 - Dough System

Experiments were performed to determine the stability and extensibility of doughs containing guar gum and TGase.

The tests performed were:

1. No guar gum or TGase added

2.0.95% guar gum

3.150U/kg flour TGase

4.150U/kg flour of TGase and 0.95% guar gum

TGase of 5.300U/kg flour

6.300U/kg flour of TGase and 0.95% guar gum

Doughs with each of the different treatments were made in triplicate.

At the beginning of the experiment, dough was prepared with different amounts of TGase and guar gum added in order to calculate the water absorption. These doughs were then prepared on a Doughometer and analyzed on a KiefferRig as described above. The results of these dough testers are shown in Table 1.

Table 1 - Dough physical property tester test

        Tgase   Guar Gum   Physical Properties of Dough   Resting Time   Stability

Days ppm % % Water Absorption of Meter Minutes Minutes

1 0 0 0 54.2 1 2.7

1 0 0.95 55.3 1.2 1.4

1 1500 0 54.7 1.5 3.6

1 1500 0.95 55.5 1 1.8

1 3000 0 55.5 1.4 2.9

1 3000 0.95 56 1.4 1.5

2 0 0 0.95 55.3 1.3 1.5

2 1500 0.95 55.5 0.9 0.8

2 3000 0 55.5 1.2 2.2

2 3000 0.95 56 2.2 2.8

3 0 0 0 54 1.2 2.8

3 0 0.95 55.3 1 1.5

3 1500 0 54.5 1 5.1

3 1500 0.95 55.5 1.4 2.1

3 3000 0 55.5 1.3 2.8

Multi-factor ANOVA test showed that guar gum and TGase had no significant effect on the resting time. An ANOVA analysis of this dough stability is shown in FIG. 18 . The results indicate that there is an interactive effect on dough stability between guar gum and TGase.

The results described in Figure 18 illustrate that low doses of TGase are beneficial to improve stability, but high doses of TGase reduce stability. The addition of 0.95% guar gum made the dough less stable, but the combined addition of TGase and guar gum restored some stability.

The results of the Kieffer Rig analysis are shown in Table 2 and the average extensibility curves are shown in Figure 19

Table 2 TGase guar gum ppm % Force Distance Area g mm g×mm 0 00 0.951500 01500 0.953000 03000 0.950 0.951500 0.953000 03000 0.950 00 0.951500 01500 0.953000 0 25.423 111.962 1727.5928.324 95.629 1631.0129.188 99.394 1598.2132.516 83.017 1760.3430.735 90.216 1650.8838.962 75.933 1837.0427.448 86.12 1623.4927.835 87.485 1600.0628.841 87.325 1502.4740.894 74.826 1852.125.677 98.77 1628.2528.635 91.831 1708.7228.506 83.892 1523.631.468 91.024 1823.5325.468 84.996 1327.05

The effect of TGase and guar gum on force was evaluated by ANOVA test shown in FIG. 20 . The results indicated a strong interaction between guar gum and Tgase for the maximum force required to stretch the dough in the Kieffer Rig.

The effect of guar gum and TGase on the distance (in mm) before the dough breaks in the Kieffer test is shown in FIG. 21 . Both guar gum and TGase decreased the distance before dough breakage, but ANOVA showed no interaction effect.

The effect of guar gum and TGas on the area under the extensibility curve is a measure of the total work input required to stretch the dough strip. This ANOVA evaluation of the effect on area is shown in FIG. 22 . The ANOVA results show an interaction between guar gum and TGase, and it is interesting to note (Figure 22) that only TGase reduces the area value, only guar gum has no effect on the area value, However, when combined, the area value increases significantly. This effect should predict improved dough stability when used in baking.

TGase and guar gum were tested in a model system in which dough was prepared on a 10 g flour basis in a microdough physical tester. The extensibility of these doughs was tested in a texture analyzer using a Kieffer Rig. The results confirmed the synergistic effect of the combined addition of guar gum and TGase to the dough. This is clearly illustrated by the effect of an increase in the maximum force required to stretch the dough, and a synergistic effect is also observed on the energy required to stretch the dough until it breaks.

Example 9 - Low Fat Sauce

 Materials and methods (low fat sauce)

Low-fat sauce model-low-fat sauce is prepared with the following basic formula:

Water Phase Lipid Phase

Water 55.6 Hydrogenated soybean oil 9.9

(mp 41℃)

Salt (NaCl) 1.2 Rapeseed oil 29.6

Skimmed Milk Powder 1 Dimodan (Monoglyceride) 0.5

Alginate 1.5 β-carotene 4ppm

TGase 0.57

Potassium sorbate 0.1

EDTA 0.015

Total water phase 60% Total lipid phase 40%

In some trials, some or all of the alginate was replaced with water.

In some experiments, TGase was replaced with water.

All aqueous phase components except TGase were mixed and dissolved at 60°C, then cooled to 37°C. TGase was added and the entire aqueous phase was incubated for 1.5 hours.

The lipid phase was mixed at 65°C and cooled to 37°C.

The two phases were combined and homogenized by vigorous stirring. The paste was crystallized and kneaded in a tube freezer. After processing, the samples were filled into 100 ml containers and left to stand at 4° C. for 14 days, after which visual and sensory evaluations were carried out.

The tests performed were:

A: 1.5% Alginate 0% TGase

B: 0.5% Alginate 0% TGase

C: 0% Alginate 0% TGase

D: 1.5% alginate 0.57% TGase

E: 0.5% Alginate 0.57% TGase

F: 0% alginate 0.57% TGase

Any components deleted compared to the base formulation in the Materials and Methods section were replaced with water.

evaluate:

Samples were evaluated by a panel of experts and scored on a scale of 0-8 for each parameter evaluated. Syneresis was evaluated by visual sensation. Stability was evaluated visually after spreading the low-fat sauce on cardboard. A stable sauce is one that maintains a smooth texture after application, ie does not become particulate. Stickiness was evaluated sensory.

table 3 Syneresis (macroscopic observation) 0 = low; 8 = high Stability (by visual observation) 0=smooth; 8=particles Viscosity (mouthfeel) 0 = low; 8 = high A 0 3 8 B 2 4 6 C 8 8 - D. 0 0 3 E. 0 1 0 f 8 8 -

It can be concluded that by combining TGase and alginate, the amount of alginate used can be reduced by at least 2/3 without greater syneresis. By combining alginate and TGase, low viscosity, high stability and low syneresis can be obtained, which indicates the synergistic effect of these two components. Only the samples containing both alginate and TGase were the products acceptable to the naked eye.

Example 10 - Processed Cheese

Materials and methods

processed cheese model

Processed cheese was prepared with the following basic recipe:

Aqueous phase %(w/w)

Water 36.1

Processed cheese 45.26

Joha S9 2.50

(Alginate) FD150 1.00

Calcium lactate 0.31

Lactic acid 0.20

Flavor 4723 2.00

Salt NaCl 0.40

Dimodan OT

Rapeseed oil 8.76

Starch 570

Skimmed Milk Powder 3.00

TGase 0.5

Raw cheese, salt, calcium lactate, oil, flavors and enzymes were mixed with 2/3 of water in a limititech mixer at 40°C and incubated at this temperature for 1 hour. The alginate was dissolved in the remaining water at 75°C and mixed with the cheese cubes. The whole mixture was heated to 95°C for 7 minutes. The pH of the mixture was adjusted to 5.6 with lactic acid, after which the mixture was tapped into 100ml plastic containers.

The tests performed were

A: 0% alginate, 0% TGase

B: 0.5% alginate, 0% TGase

C: 1% alginate, 0% TGase

D: 0% alginate, 0.5% TGase

E: 0.5% alginate, 0.5% TGase

F: 1% alginate, 0.5% TGase

evaluate

Samples were evaluated visually and by texture analysis.

Figure 23 shows graphical samples of cheeses C, D and F.

Since only alginate and Tgase combined we get a firm and firm texture, the synergistic effect of the treatments of these two compounds is immediately seen.

Also the cheese samples were evaluated by texture analysis. The results of the breaking strength of these samples are shown in Table 4.

Table 4 - Breaking Strength of Processed Cheese Samples sample Breaking strength (g/cm ² ) A 16 B 15 C 18 D. 42 E. 528 f 652

This synergy is clearly seen from the results in the table, since only the samples containing both alginate and TGase had a high breaking strength. The effect on cheese containing both alginate and TGase could not be explained by the sum of the individual TGase and alginate effects.

Example 11 - Cream Cheese

Materials and methods

cream cheese model

Cream cheese is prepared using the following basic recipe: %(w/w) water 11 Cream Cheese Base 70+ 40 Quark 47 alginate 0.2 TGase 1.0 ^NisaplinTM 0.017 NaCl 0.5 total 100

In some experiments, some or all of the alginate was replaced with water.

In some experiments, TGase was replaced with water.

Mix the cream cheese base and quark with water at 40°C. TGase was added, and the cheese block was incubated at 40°C for 30 minutes. Alginate, salt and Nisaplin ^™ were mixed and added to the cheese block. The cheese block was heated at 80°C for 3 minutes, then cooled to 70°C and filled into 100ml plastic containers. The cheese was stored at 5°C for 5 days and then evaluated.

The tests performed were

A: 0.2% Alginate 0% TGase

B: 0.1% Alginate 0% TGase

C: 0% Alginate 0% TGase

D: 0.2% alginate 1% TGase

E: 0.1% alginate 1% TGase

F: 0% alginate 1% TGase

Compared to the basic formulation in this Materials and methods section, any components deleted were replaced with water

evaluate

Samples were evaluated by a panel of experts and scored on a scale of 0-8 for each parameter evaluated. Syneresis was evaluated by visual sensation. Stability was assessed by visual perception of the cheese after it had been spread on cardboard. A stable cheese is one that maintains a smooth texture after coating, ie does not become particulate.

table 5

  Syneresis (observation with naked eyes) Stability (observation with naked eyes)

0=Low; 8=High 0=Smooth; 8=Particles

A 2 3

B 4 5

C 8 8

D 0 0

E 1 2

F 8 8

It can be seen from Table 5 that by using alginate and TGase in combination, low syneresis and high stability can be obtained. Only the samples containing both alginate and TGase were acceptable products in terms of texture and naked eyes.

in conclusion

Synergistic gel formation has been demonstrated in many systems. Particularly meaningful synergies are:

• Synergistic effect of carrageenan and crosslinking enzymes on the gel strength of soy-based sweet cream.

· Synergistic effect of carrageenan and cross-linking enzymes on the gel strength of whey-based sweet cream.

• Synergistic effect of pectin and crosslinking enzymes on the gel strength of acidified milk/soy protein gels.

Synergistic effect of carrageenan and cross-linking enzymes on the stability of protein-containing beverages containing cocoa solids

Synergistic effect of carrageenan and cross-linking enzyme on the melting of ice cream

Synergistic effects of guar gum and cross-linking enzymes on dough stability and extensibility

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

Claims (46)

1, a kind of composition, comprise hydrocolloid and enzyme, wherein said enzyme is a kind of cross-linking enzyme, and described hydrocolloid and enzyme exist with following amount: the dosage of described enzyme in proteinaceous food is not more than 20U/g and the concentration of described hydrocolloid in this food less than 1%.

2, a kind of composition comprises hydrocolloid, protein and enzyme, and wherein said enzyme is a cross-linking enzyme, and the dosage of enzyme is not more than 20U/g protein, and the concentration of hydrocolloid is less than 1%.

3, as the composition of claim 1 or 2, wherein said hydrocolloid is selected from carragheen, starch, pectin, alginates, tracasol, gum gellan, xanthans, carboxymethyl cellulose, guar gum, acacin and combination thereof.

4, as claim 1,2 or 3 composition, wherein enzyme is TGase.

5, as each composition among the claim 1-4, wherein said composition also comprises soybean protein.

6, composition as claimed in claim 5, wherein said hydrocolloid is a carragheen, preferred concentration is not more than 0.5%.

7, composition as claimed in claim 5, wherein said hydrocolloid is a starch.

8, composition as claimed in claim 5, wherein said hydrocolloid is a pectin.

9, composition as claimed in claim 8, the content of wherein said pectin is not more than 0.3%.

10, as each composition among the claim 1-4, wherein said composition also comprises lactoprotein.

11, as the composition of claim 10, wherein said hydrocolloid is a carragheen.

12, as the composition of claim 11, wherein said composition comprises a kind of emulsifying agent-stabilizing agent compound in addition.

13, as the composition of claim 10, wherein said hydrocolloid is a starch.

14, as the composition of claim 10, wherein said hydrocolloid is a pectin.

15, as the composition of claim 14, the concentration of wherein said pectin is not more than 0.3%, and the concentration of enzyme is not more than 20U/g.

16, as each composition among the claim 1-4, wherein said composition also comprises whey protein.

17, as the composition of claim 16, wherein said hydrocolloid is a carragheen.

18, as the composition of claim 17, wherein said hydrocolloid is a starch.

19, a kind of proteinaceous beverage, it comprises as each composition and lactoprotein among the claim 1-4.

20, as the proteinaceous beverage of claim 19, wherein said hydrocolloid is a carragheen.

21, as the proteinaceous beverage of claim 20, the concentration of wherein said carragheen is not more than 0.02%, and the dosage of enzyme is not more than 2U/g.

22, as the proteinaceous beverage of claim 19-21, it also comprises a kind of flavouring agent, and wherein said flavouring agent is a cocoa solids.

23, as the proteinaceous beverage of claim 19-21, it also comprises a kind of flavouring agent, and wherein said flavouring agent is selected from chocolate, strawberry, the red certain kind of berries, banana, oranges and tangerines, mango, lemon, bitter orange, cherry, peach, pears, apple, pineapple or its combination.

24, as each composition among the claim 1-4, wherein said composition also comprises gluten.

25, as the composition of claim 24, wherein said hydrocolloid is a guar gum.

26, as the composition of claim 25, wherein said guar gum is less than 1%, and the dosage of described enzyme is not more than 0.3U/g.

27, a kind of preparation of compositions method that comprises cross-linked proteins, this method comprises the step that protein and hydrocolloid are contacted with enzyme; Wherein said enzyme is a cross-linking enzyme, and the dosage of described enzyme is not more than 20U/g, and the concentration of hydrocolloid is less than 1%.

28, as the method for claim 27, wherein said hydrocolloid is selected from carragheen, starch, pectin and combination thereof.

29, as the method for claim 27 or 28, wherein said enzyme is TGase.

30, as claim 27,28 or 29 method, wherein said protein is soybean protein, and described hydrocolloid is a carragheen, and earlier carragheen is contacted with soybean protein, afterwards enzyme is contacted with soybean protein.

31, as claim 27,28 or 29 method, wherein said protein is soybean protein, and described hydrocolloid is a starch, and earlier enzyme is contacted with soybean protein, afterwards starch is contacted with soybean protein.

32, as claim 27,28 or 29 method, wherein said protein is lactoprotein, and described hydrocolloid is a carragheen, and carragheen and enzyme are contacted with lactoprotein simultaneously.

33, as claim 27,28 or 29 method, wherein said protein is lactoprotein, and described hydrocolloid is a starch, and earlier starch is contacted with lactoprotein, afterwards enzyme is contacted with lactoprotein.

34, as claim 27,28 or 29 method, wherein said protein is lactoprotein, and described hydrocolloid is a pectin, and earlier enzyme is contacted with lactoprotein, afterwards pectin is contacted with lactoprotein.

35, as claim 27,28 or 29 method, wherein said protein is whey protein, and described hydrocolloid is a starch, and earlier enzyme is contacted with whey protein, afterwards starch is contacted with whey protein.

36, as claim 27,28 or 29 method, wherein said protein is lactoprotein, and described hydrocolloid is a carragheen, and earlier carragheen is contacted with lactoprotein, afterwards enzyme is contacted with lactoprotein.

37, as claim 27,28 or 29 method, wherein said protein is gluten, and described hydrocolloid is a guar gum, and guar gum and enzyme are contacted with gluten simultaneously.

38, as the method for claim 27, each composition of wherein said hydrocolloid and enzyme such as claim 1-26 provides.

39, a kind of hydrocolloid and cross-linking enzyme are used for the purposes in the collaborative formation of proteinaceous food gel.

40, as the purposes of claim 39, wherein said hydrocolloid and crosslinked be to provide by composition as claim 1-26.

41, a kind of composition that comprises hydrocolloid, protein and cross-linking enzyme is selected from purposes in sweet food, acidifying gel, the drinkable food that contains protein beverage and dough in preparation.

42, a kind of purposes of composition in the proteinaceous ice cream of preparation that comprises hydrocolloid and cross-linking enzyme, the dosage of wherein said enzyme is not more than 20U/g.

43, as the purposes of claim 42, wherein said hydrocolloid is a carragheen, and described enzyme is TGase.

44, a kind of composition is basically as described in this paper front embodiment 1,2,3,4,5,6,7 or 8.

45, a kind of method is basically as described in this paper front embodiment 1,2,3,4,5,6,7 or 8.

46, a kind of purposes is basically as described in this paper front embodiment 1,2,3,4,5,6,7 or 8.

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