Slowly digestible protein gel as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of food nutrition and digestion, and particularly relates to slowly digestible protein gel as well as a preparation method and application thereof. The preparation method of the slowly digestible protein gel mainly comprises the following steps: dissolving protein in water, adding iota-carrageenan or iota-carrageenan oligosaccharide, dissolving, adjusting the pH value, heating in a water bath, and cooling to obtain the slowly digestible protein gel. In the preparation method, the dosage of iota-carrageenan and iota-carrageenan oligosaccharide is relatively low, an acid inducer does not need to be used, the operation is simple and convenient, the production cost is low, and the safety is relatively high. The slowly digestible protein gel prepared by the preparation method has relatively low protein digestibility in gastric digestive juice and intestinal digestive juice, and can be used for preparing products for delaying gastric emptying and promoting satiety.

Description

Technical Field

[0001] The present invention belongs to the technical field of food nutrition and digestion, and particularly relates to a slowly digested protein gel, a preparation method and an application thereof. Background Art

[0002] With the improvement of people's living standards and the formation of unhealthy eating habits, problems such as obesity, type II diabetes, gastrointestinal diseases, etc. caused by overnutrition are increasing. The current nutritional status of the general public has put forward new requirements for food, and characteristics such as prolonging the digestion and absorption time of food, delaying gastric emptying, and enhancing satiety have received more attention. Protein is the most important and essential nutrient in whole-nutrition formula foods. Among them, eggs are an important source of high-quality protein and are also one of the indispensable foods for people. Ovalbumin (OVA) is the main component of egg white protein, accounting for about 54% of the total protein.

[0003] The digestive properties of proteins are closely related to the microstructure of protein gels. The addition of saccharide substances can significantly improve the stability and gelling properties of OVA under various processing conditions, and thus affect the digestive performance of OVA. ι-Carrageenan (ι-CG) is a hydrophilic linear sulfated polysaccharide extracted from marine edible red algae, and has various health functional characteristics, including antioxidant and immunomodulatory activities and prebiotic effects. ι-CG has thermoreversibility, which makes it widely used as a thickener, stabilizer, etc. in the food industry. Some researchers have proposed that the digestibility of the gel formed by ι-carrageenan and whey protein isolate is reduced, but the digestibility of the gel can only be reduced to an ideal level when the addition amount of ι-carrageenan reaches 0.23%, and there are safety problems in the application of this gel because an acid inducer needs to be introduced in the preparation process. In addition, the main digestion sites of proteins are the stomach and small intestine. The existing technologies for studying the reduction of protein digestibility mainly focus on its digestibility in the stomach, and do not consider the digestion of proteins in the small intestine. Summary of the Invention

[0004] Aiming at the above technical problems, the present invention provides a slowly digested protein gel, a preparation method and an application thereof. In the preparation method of the slowly digested protein gel provided by the present invention, the usage amounts of ι-carrageenan and ι-carrageenan oligosaccharide are both low, and no acid inducer is required, the operation is simple, the production cost is low, and it has higher safety. During the simulated gastric digestion process, the slowly digested protein gel prepared by this preparation method has a low protein digestibility in both gastric digestive juice and intestinal digestive juice, and thus can be used to prepare products for delaying gastric emptying and promoting satiety.

[0005] To achieve the above invention purpose, the present invention adopts the following technical solutions: The first aspect of the present invention provides a method for preparing a slowly digestible protein gel, which specifically includes the following steps: S1. Dissolve the protein in water to obtain an aqueous protein solution after sufficient hydration; add ι-carrageenan (ι-CG) or ι-carrageenan oligosaccharide (ι-CO) to the aqueous protein solution, dissolve it, and adjust the pH to be less than the isoelectric point of the protein to obtain a composite solution; the isoelectric point of the protein is greater than 4.4; the mass of ι-carrageenan or ι-carrageenan is at least 0.03% of the mass of the aqueous protein solution; S2. Heat the composite solution in a water bath at 85-95 °C for 20-60 min, and then immediately cool it in an ice-water bath for 5-15 min to obtain the slowly digestible protein gel.

[0006] ι-Carrageenan oligosaccharide is a degradation product of ι-CG, with a molecular weight usually <2500 Da, having various physiological activities and excellent physicochemical properties. In this preparation method, the dosages of ι-carrageenan and ι-carrageenan oligosaccharide are both relatively low, and they can be dissolved in the aqueous protein solution without heating, and no acid inducer is required. Therefore, the preparation method is simple to operate, has low production costs, and has higher safety.

[0007] The slowly digestible protein gel prepared by this preparation method has a low protein digestibility during the simulated in vitro digestion process.

[0008] In this preparation method, the type of carrageenan is crucial for the digestibility of the resulting gel during digestion. Compared with common κ-carrageenan, ι-carrageenan has two sulfate groups, a higher charge, stronger electrostatic interaction with proteins with an isoelectric point greater than 4.4, and a higher ability to inhibit protein thermal aggregation. If ι-carrageenan is replaced with κ-carrageenan, it cannot significantly affect the properties such as gel hardness and gel network structure, thus cannot effectively reduce the accessibility of proteases to peptide bonds, and the protein digestibility of the gel cannot be effectively reduced.

[0009] The type of oligosaccharide is also important for the digestibility of the resulting gel during digestion. Different oligosaccharides have different biological activities and properties, and not all oligosaccharides can reduce protein digestibility. It is reported that fucoidan can improve the digestibility of scallop myofibrillar protein, and another study shows that 1% chitosan oligosaccharide has no significant effect on the hydrolysis degree of whey protein isolate during gastric digestion. The present invention creatively uses ι-carrageenan oligosaccharide, and its addition amount reaches 0.03% to effectively reduce the protein digestibility of the resulting gel in gastric digestive juice and intestinal digestive juice, thus producing a good effect of delaying gastric emptying and promoting satiety.

[0010] Preferably, the protein is ovalbumin (OVA).

[0011] Both ι-CG and ι-CO used in the present invention are anionic saccharide substances. After binding to OVA, they form a complex, and through heat cross-linking, a protein-sugar composite gel is obtained. This composite gel can reduce the hydrolysis degree of OVA during gastrointestinal digestion, has the potential to delay gastric emptying and promote satiety, and is beneficial for weight control and achieving precise nutrition.

[0012] Further preferably, the preparation method of the ovalbumin is as follows: After beating fresh egg white and mixing it with polyethylene glycol-8000, adjust the pH to 6.5, add NaCl to make its concentration 100 mmol / L, centrifuge after reacting for 25 - 35 min, and collect the supernatant; adjust the pH of the supernatant to 4.5 and centrifuge, and the obtained precipitate is the ovalbumin.

[0013] Preferably, the method for dissolving after adding ι-carrageenan or ι-carrageenan oligosaccharide to the protein aqueous solution in S1 is: Stir at 500 - 700 r / min for 1.5 - 2.5 h.

[0014] Preferably, the mass of ι-carrageenan or ι-carrageenan in S1 is at least 0.03% - 0.10% of the mass of the protein aqueous solution. Compared with the pure ovalbumin gel, adding 0.03% - 0.10% of ι-carrageenan or ι-carrageenan can achieve a better effect of controlling protein digestion and significantly reduce the protein digestibility.

[0015] The second aspect of the present invention provides a slowly digested protein gel prepared by the above preparation method.

[0016] The third aspect of the present invention provides the application of the above slowly digested protein gel in the preparation of products for delaying gastric emptying and / or promoting satiety.

[0017] Preferably, the products include foods, health foods or foods for special medical purposes.

[0018] The beneficial effects of the present invention are as follows: In the preparation method provided by the present invention, the dosages of ι-carrageenan and ι-carrageenan oligosaccharide are both low, they can be dissolved in the protein aqueous solution without heating, and no acid inducer is required. Therefore, the operation is simple, easy to scale up production, and has a low production cost, and has higher safety. Experiments prove that the slowly digested protein gel obtained by the preparation method of the present invention is not easily hydrolyzed by pepsin and pancreatic enzymes under simulated human gastrointestinal digestion conditions, has a lower digestibility, and has a stronger gel structure, and has the ability to delay gastric emptying and promote satiety, which is beneficial for meeting the needs of special populations and achieving precise nutrition.

[0019] All raw materials used in the preparation method of the present invention are food-grade, so the prepared slowly digested protein gel can be applied in food fields such as ordinary foods, health foods, and foods for special medical purposes. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 This is for the digestibility of OVA / ι-CG thermally induced composite gel and OVA / ι-CO thermally induced composite gel in gastrointestinal digestive juices in Test Example 2 of the present invention; Figure 2 This is for the free amino group content in gastrointestinal digestive juices in Test Example 2 of the present invention; Figure 3 This is for the particle size distribution of OVA / ι-CG thermally induced composite gel and OVA / ι-CO thermally induced composite gel in gastrointestinal digestive juices after digestion in Test Example 2 of the present invention; Figure 4 This is for the frequency scanning results of gastrointestinal digestive juices in Test Example 2 of the present invention; Figure 5 This is for the peptide molecular weight distribution of gastrointestinal digestive juices in Test Example 2 of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0021] In order to make the objectives, technical solutions and advantages of the present invention clearer and more understandable, the following further describes the present invention in detail with reference to specific embodiments. It should be understood that the specific embodiments described herein are only for explaining the present invention and are not used to limit the present invention. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts fall within the protection scope of the present invention.

[0022] Excessive food intake and problems of overnutrition can cause various health problems. Ways such as prolonging the digestion and absorption time of food, delaying gastric emptying, and promoting satiety are expected to solve this problem. Protein is the most important and essential nutrient in whole-nutrition formula foods. A reduction in its digestion rate is expected to improve protein utilization, delay gastric emptying, and enhance satiety.

[0023] The digestion characteristics of proteins are closely related to the microstructure of protein gels. In order to slow down the digestion rate of protein gels, developing protein gels with slow digestion characteristics is an urgent need in the fields of food nutrition, healthcare, etc.

[0024] In view of the above problems, the embodiments of the present invention provide a method for preparing a slow-digesting protein gel, which specifically includes the following steps: S1. Dissolve the protein in water, and obtain a protein aqueous solution after sufficient hydration; add ι-carrageenan (ι-CG) or ι-carrageenan oligosaccharide (ι-CO) to the protein aqueous solution, dissolve it, and adjust the pH to be less than the isoelectric point of the protein to obtain a composite solution; the isoelectric point of the protein is greater than 4.4; the mass of the ι-carrageenan or ι-carrageenan is at least 0.03% of the mass of the protein aqueous solution; S2. Heat the composite solution in a water bath at 85 - 95 °C for 20 - 60 min, and then immediately cool it in an ice - water bath for 5 - 15 min to obtain the slow - digestible protein gel.

[0025] The embodiment of the present invention also provides a slow - digestible protein gel prepared by the above - mentioned preparation method.

[0026] The embodiment of the present invention also provides the application of the above - mentioned slow - digestible protein gel in the preparation of products for delaying gastric emptying and / or promoting satiety.

[0027] The following uses specific examples to illustrate the solution of the present invention.

[0028] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods. Unless otherwise specified, the materials, reagents, etc. used in the following examples can all be obtained from commercial channels.

[0029] Example 1 This example provides a slow - digestible protein gel, and the specific steps of its preparation method are as follows: (1) OVA extraction and OVA solution preparation OVA extraction: Take 1 kg of fresh egg white, stir it with a magnetic stirrer for 15 min (500 r / min), add polyethylene glycol - 8000 to the egg white to prepare an egg white - polyethylene glycol mixed solution with a polyethylene glycol concentration of 15% wt. Adjust the pH of the egg white - polyethylene glycol mixed solution to 6.5, then add NaCl to the egg white - polyethylene glycol mixed solution to a final concentration of 100 mmol / L, then stir for 30 min (300 r / min), and then centrifuge at 15000×g for 15 min to collect the supernatant. Finally, adjust the pH of the supernatant to 4.5 with 10M hydrochloric acid, and centrifuge at 15000×g for 15 min to collect the precipitate. The precipitate is OVA. Lyophilize it to obtain OVA powder.

[0030] OVA solution preparation: Weigh OVA powder and dissolve it in distilled water to make the concentration of OVA 10% (m / v). Stir it on a magnetic stirrer at 300 r / min for 2 h, and then allow OVA to fully hydrate in a 4 °C refrigerator for 13 h to obtain an OVA solution.

[0031] (2) Preparation of OVA / ι - CG composite solution Weigh a certain amount of ι - CG (the mass percentage of ι - CG in the OVA solution is 0.03%) and add it to the OVA solution obtained in step (1). Stir it on a magnetic stirrer at 600 r / min for 2 h, and then adjust the pH to 4.0 to obtain an OVA / ι - CG composite solution.

[0032] (3) Preparation of OVA / ι-CG thermosensitive gel Pour the OVA / ι-CG composite solution obtained in step (2) into a cylindrical mold with a diameter of 1.5 cm, heat it in a water bath at 90 °C for 30 min, and then immediately cool it in an ice-water bath for 10 min to obtain the OVA / ι-CG thermosensitive composite gel. Store the obtained composite gel in a refrigerator at 4 °C.

[0033] Example 2 This example provides a slowly digestible protein gel, and the specific steps of its preparation method are as follows: (1) OVA extraction and OVA solution preparation: The same as Example 1.

[0034] (2) Preparation of OVA / ι-CG composite solution: Weigh a certain amount of ι-CG (the mass percentage in the OVA solution is 0.06%) and add it to the OVA solution obtained in step (1), stir it on a magnetic stirrer at 600 r / min for 2 h, and then adjust the pH to 4.0 to obtain the OVA / ι-CG composite solution.

[0035] (3) Preparation of OVA / ι-CG thermosensitive gel: The same as Example 1.

[0036] Example 3 This example provides a slowly digestible protein gel, and the specific steps of its preparation method are as follows: (1) OVA extraction and OVA solution preparation: The same as Example 1.

[0037] (2) Preparation of OVA / ι-CG composite solution: Weigh a certain amount of ι-CG (the mass percentage in the OVA solution is 0.1%) and add it to the OVA solution obtained in step (1), stir it on a magnetic stirrer at 600 r / min for 2 h, and then adjust the pH to 4.0 to obtain the OVA / ι-CG composite solution.

[0038] (3) Preparation of OVA / ι-CG thermosensitive gel: The same as Example 1.

[0039] Example 4 This example provides a slowly digestible protein gel, and the specific steps of its preparation method are as follows: (1) OVA extraction and OVA solution preparation: OVA extraction: The same as in Example 1. Take 1 kg of fresh egg white and stir it with a magnetic stirrer for 15 min (500 r / min). Add polyethylene glycol-8000 to the egg white to prepare an egg white-polyethylene glycol mixed solution with a polyethylene glycol concentration of 15% wt. Adjust the pH of the egg white-polyethylene glycol mixed solution to 6.5. Subsequently, add NaCl to the egg white-polyethylene glycol mixed solution to a final concentration of 100 mmol / L, then stir for 25 min (300 r / min), and then centrifuge at 15000×g for 15 min to collect the supernatant. Finally, adjust the pH of the supernatant to 4.5 with 10M hydrochloric acid and centrifuge at 15000×g for 15 min to collect the precipitate, which is OVA. Freeze-dry it to obtain OVA powder.

[0040] OVA solution preparation: Weigh OVA powder and dissolve it in distilled water to make the concentration of OVA 10% (m / v). Stir it on a magnetic stirrer at 300 r / min for 2 h, and then allow OVA to fully hydrate in a 4°C refrigerator for 12 h to obtain an OVA solution.

[0041] (2)Preparation of OVA / ι-CG composite solution Weigh a certain amount of ι-CG (the mass percentage in the OVA solution is 0.1%) and add it to the OVA solution obtained in step (1). Stir it on a magnetic stirrer at 600 r / min for 1.5 h, and then adjust the pH to 4.0 to obtain an OVA / ι-CG composite solution.

[0042] (3)Preparation of OVA / ι-CG thermogel Pour the OVA / ι-CG composite solution obtained in step (2) into a cylindrical mold with a diameter of 1.5 cm, heat it in a water bath at 85°C for 60 min, and then immediately place it in an ice-water bath to cool for 5 min to obtain an OVA / ι-CG thermo-composite gel. Store the obtained composite gel in a 4°C refrigerator.

[0043] Example 5 This example provides a slow-digesting protein gel, and the specific steps of its preparation method are as follows: (1)OVA extraction and OVA solution preparation: OVA extraction: The same as in Example 1. Take 1 kg of fresh egg white, stir it with a magnetic stirrer for 15 min (500 r / min), add polyethylene glycol-8000 to the egg white to prepare an egg white-polyethylene glycol mixed solution with a polyethylene glycol concentration of 15% wt. Adjust the pH of the egg white-polyethylene glycol mixed solution to 6.5, then add NaCl to the egg white-polyethylene glycol mixed solution to a final concentration of 100 mmol / L, then stir for 35 min (300 r / min), and then centrifuge at 15000×g for 15 min to collect the supernatant. Finally, adjust the pH of the supernatant to 4.5 with 10M hydrochloric acid, centrifuge at 15000×g for 15 min to collect the precipitate, and the precipitate is OVA. Lyophilize it to obtain OVA powder.

[0044] OVA solution preparation: Weigh OVA powder and dissolve it in distilled water to make the concentration of OVA 10% (m / v). Stir it on a magnetic stirrer at 300 r / min for 2 h, and then allow OVA to hydrate fully in a 4°C refrigerator for 14 h to obtain an OVA solution.

[0045] (2)Preparation of OVA / ι-CG composite solution Weigh a certain amount of ι-CG (0.1% by mass percentage of the OVA solution) and add it to the OVA solution obtained in step (1). Stir it on a magnetic stirrer at 600 r / min for 2.5 h, and then adjust the pH to 4.0 to obtain an OVA / ι-CG composite solution.

[0046] (3)Preparation of OVA / ι-CG thermogel Pour the OVA / ι-CG composite solution obtained in step (2) into a cylindrical mold with a diameter of 1.5 cm, heat it in a water bath at 95°C for 20 min, and then immediately place it in an ice-water bath to cool for 15 min to obtain an OVA / ι-CG thermo-composite gel. Store the obtained composite gel in a 4°C refrigerator.

[0047] Example 6 This example provides a slow-digesting protein gel, and the specific steps of its preparation method are as follows: (1)OVA extraction and OVA solution preparation: The same as in Example 1.

[0048] (2)Preparation of OVA / ι-CO composite solution: Weigh a certain amount of ι-CO (0.03% by mass percentage of the OVA solution) and add it to the OVA solution obtained in step (1). Stir it on a magnetic stirrer at 600 r / min for 2 h, and then adjust the pH to 4.0 to obtain an OVA / ι-CO composite solution.

[0049] (3) Preparation of OVA / ι-CO thermosensitive gel: Pour the OVA / ι-CO composite solution obtained in step (2) into a cylindrical mold with a diameter of 1.5 cm, heat it in a water bath at 90 °C for 30 min, and then immediately place it in an ice-water bath to cool for 10 min to obtain OVA / ι-CG thermosensitive composite gel and OVA / ι-CO thermosensitive composite gel. Store the obtained composite gel in a refrigerator at 4 °C.

[0050] Example 7 This example provides a slow-digesting protein gel, and the specific steps of its preparation method are as follows: (1)OVA extraction and OVA solution preparation: The same as Example 1.

[0051] (2) Preparation of OVA / ι-CO composite solution: Weigh a certain amount of ι-CO (the mass percentage of ι-CO in the OVA solution is 0.06%) and add it to the OVA solution obtained in step (1), stir it on a magnetic stirrer at 600 r / min for 2 h, and then adjust the pH to 4.0 to obtain the OVA / ι-CO composite solution.

[0052] (3) Preparation of OVA / ι-CO thermosensitive gel: The same as Example 6.

[0053] Example 8 This example provides a slow-digesting protein gel, and the specific steps of its preparation method are as follows: (1)OVA extraction and OVA solution preparation: The same as Example 1.

[0054] (2) Preparation of OVA / ι-CO composite solution: Weigh a certain amount of ι-CO (the mass percentage of ι-CO in the OVA solution is 0.1%) and add it to the OVA solution obtained in step (1), stir it on a magnetic stirrer at 600 r / min for 2 h, and then adjust the pH to 4.0 to obtain the OVA / ι-CO composite solution.

[0055] (3) Preparation of OVA / ι-CO thermosensitive gel: The same as Example 6.

[0056] Example 9 This example provides a slow-digesting protein gel, and the specific steps of its preparation method are as follows: (1)OVA extraction and OVA solution preparation: The same as Example 4.

[0057] (2) Preparation of OVA / ι-CO composite solution: Weigh a certain amount of ι-CO (0.1% of the mass percentage of the OVA solution) and add it to the OVA solution obtained in step (1). Stir it on a magnetic stirrer at 600 r / min for 1.5 h, and then adjust the pH to 4.0 to obtain the OVA / ι-CO composite solution.

[0058] (3)Preparation of OVA / ι-CO thermogel Pour the OVA / ι-CO composite solution obtained in step (2) into a cylindrical mold with a diameter of 1.5 cm, heat it in a water bath at 85 °C for 60 min, and then immediately place it in an ice-water bath to cool for 5 min to obtain the OVA / ι-CO thermogel. Store the obtained composite gel in a refrigerator at 4 °C.

[0059] Example 10 This example provides a slow-digesting protein gel, and the specific steps of its preparation method are as follows: (1)OVA extraction and OVA solution preparation: The same as in Example 4.

[0060] (2)Preparation of OVA / ι-CO composite solution: Weigh a certain amount of ι-CO (0.1% of the mass percentage of the OVA solution) and add it to the OVA solution obtained in step (1). Stir it on a magnetic stirrer at 600 r / min for 2.5 h, and then adjust the pH to 4.0 to obtain the OVA / ι-CO composite solution.

[0061] (3)Preparation of OVA / ι-CO thermogel Pour the OVA / ι-CO composite solution obtained in step (2) into a cylindrical mold with a diameter of 1.5 cm, heat it in a water bath at 95 °C for 20 min, and then immediately place it in an ice-water bath to cool for 15 min to obtain the OVA / ι-CO thermogel. Store the obtained composite gel in a refrigerator at 4 °C.

[0062] Comparative Example 1 This example provides a protein gel, and the specific steps of its preparation method are as follows: (1)OVA extraction and OVA solution preparation: The same as in Example 1.

[0063] (2)Stir the OVA solution obtained in step (1) on a magnetic stirrer at 600 r / min for 2 h, and then adjust the pH to 4.0 and pour it into a cylindrical mold with a diameter of 1.5 cm. Heat it in a water bath at 90 °C for 30 min, and then immediately place it in an ice-water bath to cool for 10 min to obtain the protein gel. Store the obtained protein gel in a refrigerator at 4 °C.

[0064] Comparative Example 2 This embodiment provides a protein gel, and the specific steps of its preparation method are as follows: (1) OVA extraction and OVA solution preparation: The same as in Example 1.

[0065] (2) Preparation of OVA / ι-CG composite solution: Weigh a certain amount of ι-CG (the mass percentage in the OVA solution is 0.01%) and add it to the OVA solution obtained in step (1). Stir on a magnetic stirrer at 600 r / min for 2 h, and then adjust the pH to 4.0 to obtain the OVA / ι-CG composite solution.

[0066] (3) Preparation of OVA / ι-CG thermogel: The same as in Example 1.

[0067] Comparative Example 3 This embodiment provides a protein gel, and the specific steps of its preparation method are as follows: (1) OVA extraction and OVA solution preparation: The same as in Example 1.

[0068] (2) Preparation of OVA / ι-CO composite solution: Weigh a certain amount of ι-CO (the mass percentage in the OVA solution is 0.01%) and add it to the OVA solution obtained in step (1). Stir on a magnetic stirrer at 600 r / min for 2 h, and then adjust the pH to 4.0 to obtain the OVA / ι-CO composite solution.

[0069] (3) Preparation of OVA / ι-CO thermogel: The same as in Example 6.

[0070] Test Example 1 Prepare an OVA solution according to the method of Example 1. Using the same batch of OVA solution, prepare slow-digesting protein gels by the methods of Examples 1 to 3 and Examples 6 to 8 respectively, and prepare protein gels by the methods of Comparative Examples 1 to 3. Then measure the texture data of each gel. The results are shown in Table 1.

[0071] Table 1

[0072] Note: The same letters indicate that there is no significant difference in the test results under the same item, and different letters indicate that there is a significant difference in the test results under the same item ( P <0.05).

[0073] Test Example 2 Prepare the OVA solution according to the method of Example 1. Using the same batch of OVA solution, prepare slow-digesting protein gels by the methods of Examples 1-3 and Examples 6-8 respectively, and prepare protein gels by the methods of Comparative Examples 1-3. Then, evaluate the in vitro simulated digestion performance of the obtained gels respectively.

[0074] 1. Simulated digestive fluids used in the in vitro simulated digestion experiment 1.1 In vitro simulated oral digestive fluid 15.1 mmol / L KCl, 3.7 mmol / L KH 2 PO 4 ₄, 13.6 mmol / L NaHCO 3 ₃, 0.15 mmol / L MgCl 2 ₂, 0.06 mmol / L (NH 4 ) 2 ₂CO 3 ₃.

[0075] 1.2 In vitro simulated gastric digestive fluid 6.9 mmol / L KCl, 0.9 mmol / L KH 2 PO 4 ₄, 25.0 mmol / L NaHCO 3 ₃, 47.2 mmol / L NaCl, 0.12 mmol / L MgCl 2 (H 2 ) 6 ₂O, 0.5 mmol / L (NH 4 ) 2 ₂CO 3 ₃ and 0.15 mmol / L CaCl 2 ₂.

[0076] 1.3 In vitro simulated intestinal digestive fluid 6.8 mmol / L KCl, 0.8 mmol / L KH 2 PO 4 ₄, 85.0 mmol / L NaHCO 3 ₃, 38.4 mmol / L NaCl, 0.33 mmol / L MgCl 2 (H 2 ) 6 ₂O and 0.15 mmol / L CaCl 2 ₂.

[0077] 2. Experimental method 2.1 Simulated oral digestion Before simulated digestion, an appropriate amount of gel blocks prepared in Examples 1-3, Examples 6-8, and Comparative Examples 1-3 were broken up (particle size 0.5-3 mm) using a homogenizer (8000 rpm, 1 min). 2.0 g of gel particles were taken and mixed with 1.6 mL of in vitro simulated oral digestive fluid, and 75 U / mL of α-amylase and 1.4 mmol / L of CaCl 2 , and the pH was adjusted to 7.0. The final mixture was shaken (130 rpm) in a 37 °C water bath shaker for 5 min to simulate oral digestion, obtaining the protein oral digestive fluid.

[0078] 2.2 Simulated gastric digestion 3.2 mL of in vitro simulated gastric digestive fluid, 0.538 mL of distilled water, and 0.15 mml / L of CaCl 2 were added to the protein oral digestive fluid obtained in "2.1", along with 2000 U / mL of pepsin, and the pH was adjusted to 2.0. The final mixture was shaken (130 rpm) in a 37 °C water bath shaker for 2 h to simulate gastric digestion. After the gastric digestion was completed, it was heated at 90 °C for 10 min to inactivate pepsin, and then cooled in an ice water bath for 10 min to obtain the protein gastric digestive fluid. A small amount of the protein gastric digestive fluid was centrifuged at 8000 rpm for 20 min, and the supernatant was taken for subsequent analysis.

[0079] 2.3 Simulated intestinal digestion 6.4 mL of in vitro simulated intestinal digestive fluid, 1.392 mL of distilled water, 0.15 mml / L of CaCl 2 , 0.07 g of bile salts, and 800 U / mL of trypsin were added to the protein gastric digestive fluid obtained in "2.2", and the pH was adjusted to 7.0, and digestion continued for 2 h. After the intestinal digestion was completed, it was heated at 90 °C for 10 min to inactivate trypsin, and then cooled in an ice water bath for 10 min to obtain the protein intestinal digestive fluid. A small amount of the protein intestinal digestive fluid was centrifuged at 8000 rpm for 20 min, and the supernatant was taken for subsequent analysis.

[0080] 2.4 Calculate the digestion rate of the gel in gastrointestinal digestive fluids The supernatants of the protein gastric digestive fluid and the protein intestinal digestive fluid obtained in "2.2" and "2.3" were respectively mixed with an equal volume of 10%wt trichloroacetic acid aqueous solution and allowed to stand for 30 min to remove undissolved macromolecular proteins, and then centrifuged at 8000×g for 15 min. The protein concentration of the supernatant was measured using a BCA kit.

[0081] Digestion rate (%) = protein content in the supernatant after digestion / total protein content × 100 2.5 Calculate the free amino group content in gastrointestinal digestive fluids Preparation of o-phthalaldehyde (OPA) working solution: Dissolve 40 mg of OPA in 1 mL of methanol, and mix it with 25 mL of 10 mM sodium tetraborate solution, 2.5 mL of 20% (w / w) SDS solution, and 100 μL of β-mercaptoethanol. Make the solution up to 50 mL with distilled water. Subsequently, mix 4 mL of the OPA working solution with 200 μL of the supernatant of the protein gastric digestive juice and the supernatant of the protein intestinal digestive juice obtained in "2.2" and "2.3" respectively, let it stand at 37 °C for 2 min, and measure the absorbance at a wavelength of 340 nm using a microplate reader. Draw a standard curve using L-leucine (0.2 - 1.0 mM). Calculate the free amino acid content in the supernatant using this standard curve.

[0082] 2.6 Measurement of the particle size distribution of the gel after gastrointestinal digestion Take appropriate amounts of the supernatant of the protein gastric digestive juice and the supernatant of the protein intestinal digestive juice obtained in "2.2" and "2.3" respectively, shake well to make the digested chyme evenly distributed in the digestive juice, dilute it 100 times with distilled water, pass it through a 0.45 μm pore membrane, and measure the particle structure (average particle size and polydispersity coefficient) of the digestion product using a Malvern laser nanosizer. Set the measurement temperature to 25 °C, the scattering angle to 173°, the protein absorbance to 0.001, and the relative refractive index to 1.450.

[0083] 2.7 Frequency sweep in gastrointestinal digestive juice Take appropriate amounts of the supernatant of the protein gastric digestive juice and the supernatant of the protein intestinal digestive juice obtained in "2.2" and "2.3", and perform a frequency sweep on them using a rotational rheometer. Measurement conditions: The fixture is 40 mm, the temperature is 37 °C, and the gap is set to 1 mm. Perform a frequency sweep in the frequency range of 0.1 - 10 Hz at a strain of 1% (within the linear viscoelastic region), and collect data on the storage modulus ( G' G') and the loss modulus ( G G'').

[0084] 2.8 Measurement of the peptide molecular weight distribution in gastrointestinal digestive juice The supernatants of the gastric digestive fluid and intestinal digestive fluid of the proteins obtained from "2.2" and "2.3" were filtered through a 0.45 μm microporous filter membrane, degassed by ultrasound for 30 min, and the peptide molecular weight distribution was detected by HPLC. The chromatographic conditions of HPLC were as follows: mobile phase: acetonitrile / water / trifluoroacetic acid = 45 / 55 / 0.1 (V / V / V); chromatographic column: TSKgel G2000 SWXL (300mm * 7.8mm, 0.5um); detection wavelength: 220 nm; elution speed: 0.5 mL / min; injection volume: 10.0 uL. Protein standards: Cytochrome C (12327.0 Da), Aprotinin (6511.0 Da), Bacitracin (1422.0 Da), Ala-Ala-Tyr-Arg (451.0 Da), Gly-Gly-Gly (189.0 Da).

[0085] 3. Results 3.1 Digestibility As Figure 1 shown. From Figure 1 (A), it can be seen that adding ι-CG and ι-CO at a concentration of 0.01% had no significant effect on the gastric digestibility of OVA gel; when the concentration of ι-CG and ι-CO was higher than 0.03%, the gastric digestibility of OVA gel gradually decreased; when the concentration of ι-CG and ι-CO was 0.1%, the gastric digestibility of OVA gel decreased from 36.43 ± 0.61% to 21.32 ± 0.59% ( P <0.001) and 18.96 ± 0.95% ( P <0.001), respectively. In addition, the digestibility of OVA / ι-CO composite gel was lower than that of OVA / ι-CG composite gel ( P <0.05). From Figure 1 (B), it can be seen that when the added concentration of ι-CG and ι-CO was 0.1%, the intestinal digestibility of OVA gel decreased from 88.25 ± 1.64% to 42.65 ± 1.99% ( P <0.001) and 34.09 ± 1.59% ( P <0.001), respectively. The digestibility of OVA / ι-CO composite gel was lower than that of OVA / ι-CG composite gel ( P <0.05). The reason for this result may be that the electrostatic interaction between ι-CG, ι-CO and OVA reduces the accessibility of protease to peptide bonds, thereby reducing the digestibility of proteins, making the gel have the potential to delay gastric emptying and enhance satiety.

[0086] 3.2 Free amino group content As Figure 2As shown in the figure. With the addition of ι-CG and ι-CO, the free amino acid content in gastrointestinal digestive juices decreased significantly. In the gastric digestion products, the free amino acid content decreased from 13.25 ± 0.19 mmol / L to 2.69 ± 0.35 mmol / L ( P <0.001), 2.48 ± 0.26 mmol / L ( P <0.001). In the intestinal digestion products, the free amino acid content decreased from 144.43 ± 3.07 mmol / L to 65.64 ± 5.06 mmol / L ( P <0.001), 59.42 ± 2.06 mmol / L ( P <0.001). Among them, the free amino acids in the intestinal digestive juices of OVA / ι-CO composite gels were significantly lower than those of OVA / ι-CG composite gels. The above results indicate that ι-CG and ι-CO can effectively delay the digestion of OVA gels.

[0087] 3.3 Particle size distribution As Figure 3 shown. During the gastric digestion stage, with the increase in the concentrations of ι-CG and ι-CO, the particle size of OVA gels in gastric digestive juices gradually increased. When the addition concentrations of ι-CG and ι-CO were 0.1%, the particle sizes of OVA gels in the gastric digestion products increased from 511.60 ± 40.16 nm to 1052.33 ± 46.54 nm ( P <0.001), 1105.66 ± 58.70 nm ( P <0.001), indicating that the addition of ι-CG and ι-CO reduced the degree of digestion of OVA gels in the stomach. Among them, the degree of digestion of OVA / ι-CO gels was lower than that of OVA / ι-CG gels. After further experiencing the intestinal digestion stage, the particle sizes of the digestion products decreased significantly. The particle size of the OVA gel digestion products was 150.03 ± 30.00 nm. When the addition concentrations of ι-CG and ι-CO were 0.1%, the particle sizes of OVA / ι-CG and OVA / ι-CO gels in the intestinal digestion products were 642.96 ± 38.6 nm ( P <0.001) and 700.35 ± 10.00 nm ( P <0.001), respectively, indicating that the addition of ι-CG and ι-CO reduced the degree of digestion of OVA gels in the intestine. This result is consistent with the digestion rate results.

[0088] 3.4 Frequency sweep As Figure 4 shown. The storage modulus ( G' ) of all samples after gastrointestinal digestion was greater than the loss modulus ( G′′), indicating that all samples exhibited elastic behavior. During gastrointestinal digestion, the addition of ι-CG and ι-CO increased the G' and G ′′ values ( P <0.05), indicating that ι-CG and ι-CO enhanced the viscoelastic behavior of the OVA gel digestion products and had the potential to delay gastric emptying and enhance satiety.

[0089] 3.5 Peptide molecular weight distribution As Figure 5 shown. During the gastric digestion stage, the OVA gel was mainly hydrolyzed into large molecular peptides. With the increase in the concentration of ι-CG and ι-CO, the percentage of small molecular peptides (<1000 Da) generally showed a downward trend, while the percentage of large molecular peptides (>3000 Da) showed an upward trend. When the concentration of ι-CG and ι-CO was 0.1%, the small molecular peptides (<1000 Da) in the digestion products decreased from 5.02 ± 0.23% to 2.93 ± 0.13% ( P <0.001), 0.80 ± 0.09% ( P <0.001), while the large molecular peptides (>3000 Da) increased from 53.00 ± 0.45% to 59.02 ± 0.65% ( P <0.05), 60.94 ± 0.90% ( P <0.05).

[0090] During the intestinal digestion stage, the large molecular peptides were gradually hydrolyzed into small molecular peptides. When the concentration of ι-CG and ι-CO was 0.1%, the small molecular peptides (<1000 Da) in the digestion products decreased from 73.13 ± 1.01% to 63.76 ± 1.08% ( P <0.01), 60.13 ± 1.11% ( P <0.01), while the large molecular peptides (>3000 Da) increased from 2.2 ± 0.34% to 9.33 ± 0.34% ( P <0.001), 12.77 ± 0.44% ( P <0.001).

[0091] From the above results, it can be seen that the addition of ι-CG and ι-CO had a significant effect on the OVA gel digestion products, reducing the percentage of small molecular peptides, that is, reducing the protein digestibility, and the effect of ι-CO on reducing the digestibility of OVA protein gel was more significant.

[0092] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention shall be included within the protection scope of the present invention.

Claims

1. A method for preparing a slow digestible protein gel, characterized in that: The specific steps include: S1. dissolving a protein in water and obtaining a protein aqueous solution after sufficient hydration; adding iota carrageenan or iota carrageenan oligosaccharide to the protein aqueous solution, dissolving, and adjusting the pH to be less than the isoelectric point of the protein to obtain a composite solution; the isoelectric point of the protein is greater than 4.4; the mass of the iota carrageenan or iota carrageenan is at least 0.03% of the mass of the protein aqueous solution; S2. The composite solution is heated in a water bath at 85-95° C. for 20-60 min, and then immediately cooled in an ice water bath for 5-15 min to obtain the slowly digestible protein gel.

2. The preparation method according to claim 1, characterized in that: The protein is ovalbumin.

3. The preparation method according to claim 2, characterized in that: The preparation method of the ovalbumin is as follows: fresh egg white is beaten and then mixed with polyethylene glycol-8000, the pH is adjusted to 6.5, NaCl is added to make its concentration 100 mmol / L, the mixture is reacted for 25 to 35 minutes, the mixture is centrifuged, and the supernatant is collected; the pH of the supernatant is adjusted to 4.5, the mixture is centrifuged, and the resulting precipitate is the ovalbumin.

4. The preparation method according to any one of claims 1 to 3, characterized in that In S1, after adding iota-carrageenan or iota-carrageenan oligosaccharide to the protein aqueous solution, the dissolution method is: stirring at 500-700 r / min for 1.5-2.5 h.

5. The preparation method according to any one of claims 1 to 3, characterized in that The mass of the iota-carrageenan or iota-carrageenan in S1 is at least 0.03% to 0.10% of the mass of the protein aqueous solution.

6. The slowly digestible protein gel prepared by the preparation method according to any one of claims 1 to 5.

7. Use of the slowly digestible protein gel according to claim 6 in preparing products for delaying gastric emptying and / or promoting satiety.

8. The use according to claim 7, characterized in that: The products include food, health food or special medical purpose formula food.

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