Chinese yam polysaccharide-whey protein Maillard reactant as well as preparation method and application thereof
By carrying out Maillard reaction between yam polysaccharide and whey protein, a yam polysaccharide-whey protein Maillard reactant with high emulsification and antioxidant activity was prepared, which solved the problem of insufficient emulsification and antioxidant activity of reactants in the prior art, and achieved its efficient application in the emulsion system.
Patent Information
- Application Number
- CN202510015567.4
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-01-06
- Publication Date
- 2025-04-11
AI Technical Summary
The prior art is difficult to improve the emulsification and antioxidant activity of the reactants through the Maillard reaction. It is used as an emulsifier in the O/W emulsion system, and there is insufficient understanding of the relationship between the Maillard reaction process and the product structure and functional properties.
The Maillard reaction was carried out with yam polysaccharide and whey protein, and the yam polysaccharide-whey protein Maillard reactant was prepared by the treatment of α amylase and alkaline protease. The method includes homogenizing fresh yam with water, adding alpha amylase for water bath treatment, then adding alkaline protease and concentrating in water bath, and finally mixing with whey protein at a certain mass ratio, adjusting the pH value and performing Maillard reaction.
The DPPH scavenging rate, ABTS radical scavenging rate, hydroxyl radical scavenging rate and reducing power of the yam polysaccharide-whey protein Maillard reactant was improved, and the emulsification characteristics, protein solubility and foaming properties were enhanced, and its efficient and stable application in the food and medicine fields was achieved.
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Abstract
Description
Technical Field
[0001] The invention belongs to the technical field of protein processing, and particularly relates to a yam polysaccharide-whey protein Maillard reaction product and a preparation method and application thereof. Background Art
[0002] Whey protein isolate (WPI) is a high-quality protein that contains a variety of essential amino acids and bioactive ingredients and has high nutritional value. However, its functional properties are often affected by factors such as pH, temperature, and ionic strength, which greatly limits the application of WPI in food engineering. Therefore, WPI needs to be appropriately modified to improve the processing characteristics of whey protein.
[0003] The Maillard reaction refers to a complex chemical reaction between amino-containing compounds and carbonyl-containing compounds. It consists of three stages: the primary stage, the intermediate stage, and the late stage. In the initial stage, the reducing power of the reaction system is significantly improved, and the functional properties of the protein / peptide in the system, such as emulsification, gel properties, and solubility, will be significantly improved. As a method of chemical modification, the Maillard reaction has been applied to the food field. Therefore, research on improving the functional properties and biological activities of polysaccharides and protein / peptide substances through the Maillard reaction has attracted more and more attention.
[0004] However, there are few research reports on using Maillard modification technology to simultaneously improve the emulsification and antioxidant activity of reactants and its application as an emulsifier in O / W emulsion systems; the relationship between the Maillard reaction process and the structure and functional properties of the products also needs further in-depth discussion. Summary of the invention
[0005] The present invention aims to solve the technical gap in the prior art of using Maillard modification technology to simultaneously improve the emulsification and antioxidant activity of reactants and to use them as emulsifiers in O / W emulsions, and to further explore the relationship between the Maillard reaction process and the product structure and functional properties. A yam polysaccharide-whey protein Maillard reactant and a preparation method and application thereof are provided.
[0006] One of the purposes of the present invention is to provide a method for preparing a yam polysaccharide-whey protein Maillard reaction product, the preparation method comprising the following steps:
[0007] S1: freshly washed and peeled yam and water were homogenized in a volume ratio of 1:10, α-amylase was added thereto, the mixture was in a water bath at 80°C for 4 hours, cooled to room temperature, filtered, centrifuged at 8000 rpm for 20 minutes, the precipitate was discarded, and the supernatant was taken to obtain crude yam polysaccharide supernatant;
[0008] S2: adding alkaline protease to the crude yam polysaccharide supernatant obtained in S1, incubating in a water bath at 55°C for 6 hours, cooling and concentrating, adding 3 times the volume of anhydrous ethanol, washing, re-dissolving and freeze-drying to obtain yam polysaccharide powder;
[0009] S3: The yam polysaccharide powder obtained in S2 is uniformly mixed with whey protein in a certain mass ratio, the pH value of the mixture is adjusted with 0.2 mol hydrochloric acid, and the mixture is hydrated overnight in a 4°C refrigerator, and reacted at 55°C and 130 r / min for 5 hours to obtain a yam polysaccharide-whey protein Maillard reaction product.
[0010] In a preferred embodiment of the present invention, the added amount of the α-amylase in S1 is 20 μ / mL.
[0011] In a preferred embodiment of the present invention, the amount of alkaline protease added in S2 is 100 μ / mL.
[0012] In a preferred embodiment of the present invention, the concentration of the whey protein in S3 is 10 mg / mL.
[0013] In a preferred embodiment of the present invention, the mixing mass ratio of the yam polysaccharide powder to the whey protein in S3 is 4:8.
[0014] In a preferred embodiment of the present invention, the pH value in S3 is 7.0.
[0015] The second object of the present invention is to provide a yam polysaccharide-whey protein Maillard reaction product, which is obtained by the above-mentioned preparation method.
[0016] The third object of the present invention is to provide the application of the above-mentioned yam polysaccharide-whey protein Maillard reaction product in the food field and the medical field.
[0017] The beneficial effects of the present invention are as follows: yam polysaccharide is a heteropolysaccharide. Compared with other monosaccharides or polysaccharides, yam polysaccharide has a larger molecular weight. After being modified and combined with whey protein, the adsorption layer adsorbed on the surface of oil droplets will also be thicker. Therefore, the protein modified with yam polysaccharide has better emulsification than other monosaccharide or polysaccharide modified proteins and has the potential for stronger emulsification performance.
[0018] The invention provides a method for preparing a yam polysaccharide-whey protein Maillard reactant. The yam polysaccharide is used to modify whey protein through the Maillard reaction to prepare the yam polysaccharide-whey protein Maillard reactant. Compared with single-treated whey protein, the Maillard reactant obtained by the preparation method provided by the invention has higher DPPH scavenging rate, ABTS free radical scavenging rate, hydroxyl free radical scavenging rate and reducing power, and improves the emulsifying properties, as well as the protein solubility and foaming properties.
[0019] Chinese yam polysaccharide has functions such as immunomodulatory activity, anti-tumor activity, hypoglycemic and blood lipid activity, antioxidant activity, anti-inflammatory and antibacterial activity, etc. Therefore, the present invention selects Chinese yam polysaccharide to modify whey protein, combines Chinese yam polysaccharide with whey protein to exert the synergistic effect of the two, and achieves the purpose of improving the DPPH scavenging rate, ABTS free radical scavenging rate, hydroxyl free radical scavenging rate and reducing power of whey protein, emulsifying properties, protein solubility and foaming properties; at the same time, by utilizing the various physiological functions of Chinese yam polysaccharide, the Chinese yam polysaccharide-whey protein Maillard reaction product can be applied to the food field and the medical field, and a highly efficient and stable new oral emulsifier is prepared, thereby improving the utilization rate of Chinese yam polysaccharide and whey protein. BRIEF DESCRIPTION OF THE DRAWINGS
[0020] Figure 1 This is the statistical chart of DPPH clearance test results:
[0021] Figure 2 This is the statistical chart of ABTS free radical scavenging rate test results:
[0022] Figure 3 This is a statistical chart of the hydroxyl radical scavenging rate test results:
[0023] Figure 4 This is a statistical chart of the reduction test results:
[0024] Figure 5 This is the statistical chart of the emulsifying activity EAI test results:
[0025] Figure 6 This is the statistical chart of the ESI test results of emulsion stability:
[0026] Figure 7 This is a statistical chart of the foaming activity test results:
[0027] Figure 8 This is a statistical chart of the foaming activity stability test results:
[0028] Fig. 9 This is a statistical chart of protein solubility test results. DETAILED DESCRIPTION
[0029] Those skilled in the art can refer to the content of this article and appropriately improve the process parameters. It should be particularly noted that all similar replacements and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention. The method and application of the present invention have been described through preferred embodiments, and relevant personnel can obviously modify or appropriately change and combine the methods and applications described herein without departing from the content and scope of the present invention to implement and apply the technology of the present invention.
[0030] In order to make the purpose, technical scheme and advantages of the present invention more clear, the present invention is further described in detail below in conjunction with specific embodiments and the accompanying drawings of the specification. The test methods used in the following examples are conventional methods unless otherwise specified, and the materials, reagents, methods and instruments used are conventional materials, reagents, methods and instruments in the art unless otherwise specified, and can be obtained by those skilled in the art through commercial channels.
[0031] Embodiment 1:
[0032] S1: freshly washed and peeled yam and water were homogenized in a volume ratio of 1:10, 20 μ / mL α-amylase was added thereto, the mixture was incubated in a water bath at 80°C for 4 h, cooled to room temperature, filtered, centrifuged at 8000 rpm for 20 min, the precipitate was discarded, and the supernatant was taken to obtain crude yam polysaccharide supernatant;
[0033] S2: add 100 μ / mL alkaline protease to the crude yam polysaccharide supernatant obtained in S1, incubate in a water bath at 55°C for 6 h, cool and concentrate, add 3 times the volume of anhydrous ethanol, wash 3 times, re-dissolve and freeze-dry to obtain yam polysaccharide powder;
[0034] S3: The yam polysaccharide powder obtained in S2 was uniformly mixed with 10 mg / mL whey protein in a mass ratio of 4:8, the pH value of the mixture was adjusted to 7.0 using 0.2 mol hydrochloric acid, hydrated in a 4°C refrigerator overnight, and reacted at 55°C and 130 r / min for 5 hours to obtain a yam polysaccharide-whey protein Maillard reaction product.
[0035] Comparative Example 1:
[0036] The pH value of 10 mg / mL whey protein was adjusted to 7.0 using 0.2 mol hydrochloric acid, hydrated overnight in a 4° C. refrigerator, and reacted at 55° C. and 130 r / min for 5 h to obtain a whey reactant.
[0037] Comparative Example 2:
[0038] The difference between this comparative example and Example 1 is that the mixing mass ratio of the yam polysaccharide powder and the whey protein in S3 is 4:1; the remaining steps are the same as in Example 1.
[0039] Comparative Example 3:
[0040] The difference between this comparative example and Example 1 is that the mixing mass ratio of the yam polysaccharide powder and the whey protein in S3 is 4:3; the remaining steps are the same as in Example 1.
[0041] Comparative Example 4:
[0042] The difference between this comparative example and Example 1 is that the mixing mass ratio of the yam polysaccharide powder and the whey protein in S3 is 4:5; the remaining steps are the same as in Example 1.
[0043] Effect experiment:
[0044] (1) DPPH clearance rate
[0045] The five reactants prepared in Example 1 and Comparative Examples 1-4 were used as samples and diluted with deionized water to a sample concentration of 1 mg / mL; DPPH was weighed and dissolved in anhydrous ethanol to prepare a DPPH solution with a concentration of 0.2 mmol / L, which was stored in the dark for later use; 50 μL of DPPH solution was added to each of the five reactant samples of 100 μL, mixed evenly, reacted at room temperature in the dark for 30 minutes, and the absorbance of the mixed solution was measured at 517 nm, recorded as A1; 50 μL of anhydrous ethanol was used instead of DPPH solution as a blank group, and the absorbance after mixing was measured, recorded as A2; 100 μL of deionized water was mixed with DPPH solution as a control group, and the absorbance after mixing was measured, recorded as A0.
[0046]
[0047] The results are as follows Figure 1 As shown, compared with whey protein that has only undergone a single treatment (Comparative Example 1), the DPPH clearance rate of the Maillard reactant prepared by whey protein and adding yam polysaccharide (Example 1 and Comparative Examples 2-4) is significantly increased, among which, when the mixing mass ratio of yam polysaccharide to whey protein is 4:8, the DPPH clearance rate is the highest; indicating that the whey protein is modified by the Maillard reaction using yam polysaccharide, so that the antioxidant property of the whey protein is changed, and when the mixing mass ratio of yam polysaccharide powder to whey protein is 4:8, the prepared yam polysaccharide-whey protein Maillard reactant has the strongest antioxidant capacity.
[0048] Analysis shows that this is because the addition of yam polysaccharides will introduce sugar molecules into whey protein, and this process changes the conformation of whey protein, exposing some amino acid residues with antioxidant activity on the molecular surface, thereby enhancing its contact and reaction ability with DPPH free radicals, thereby increasing the antioxidant properties of the treated whey protein.
[0049] (2) ABTS free radical scavenging rate
[0050] Take the five reactants prepared in Example 1 and Comparative Examples 1-4 as samples, dilute them with deionized water to a sample concentration of 1 mg / mL; prepare 7.4 mmol / L ABTS stock solution and 2.6 mmol / L K2S2O8 stock solution, mix the two stock solutions in a mass ratio of 1:1, and keep them in a dark place for 12-16 hours to obtain ABTS working solution; dilute the working solution with anhydrous ethanol so that its absorbance value at room temperature 734 nm is 0.7±0.02; add 0.16 mL ABTS to each of the five reactant samples of 20 μL. + The working solution was placed at room temperature and away from light for 6 min, and the absorbance was measured at a wavelength of 734 nm. This was repeated three times and recorded as A1. The blank group was the one without adding reactant samples and recorded as A0.
[0051] ABTS free radical scavenging rate (%) = [A0-A1 / A0] × 100%
[0052] The results are as follows Figure 2 As shown, compared with whey protein (Comparative Example 1) that has only been subjected to a single treatment, the Maillard reaction products prepared by whey protein and adding yam polysaccharide (Example 1 and Comparative Examples 2-4) have a stronger ABTS free radical scavenging rate. Among them, when the mixing mass ratio of yam polysaccharide to whey protein is 4:8, the addition of yam polysaccharide will change the surface properties and structure of whey protein, so it is easier to combine with free radicals.
[0053] (3) Hydroxyl radical (OH - ) Clearance
[0054] The five reactants prepared in Example 1 and Comparative Examples 1-4 were used as samples and diluted with deionized water to a sample protein concentration of 1 mg / mL; 1 mL of each of the five sample solutions prepared in Example 1 and Comparative Examples 1-4 was taken, and mixed with 1 mL of salicylic acid solution (4.5 mmol / L) and 1 mL of ferrous sulfate solution (4.5 mmol / L), respectively, and then 2 mL of hydrogen peroxide solution (6 mmol / L) was added, respectively, and after being fully mixed, placed in a 37° C. light-proof water bath for 30 min, and the absorbance was measured at a wavelength of 510 nm, recorded as A1; then centrifuged at 4000 r / min for 10 min, and the supernatant was taken to measure the absorbance at 510 nm, deionized water was used instead of hydrogen peroxide as a control, recorded as A2, and deionized water was used instead of the sample to measure the absorbance as a blank, recorded as A0.
[0055]
[0056] The results are as follows Figure 3As shown in the figure, compared with the whey protein (Comparative Example 1) that has only been subjected to a single treatment, the Maillard reaction products prepared by adding whey protein and yam polysaccharide (Example 1 and Comparative Examples 2-4) have stronger hydroxyl radical (OH - ) clearance rate. Among them, when the mixing mass ratio of yam polysaccharide to whey protein is 4:8, the hydroxyl radical (OH - ) clearance rate was significantly higher than that of Maillard reactants prepared with other mixing mass ratios.
[0057] (4) Reduction power
[0058] The five reactants prepared in Example 1 and Comparative Examples 1-4 were used as samples and diluted with deionized water to a sample protein concentration of 5 mg / mL; 2 mL of potassium ferricyanide solution (1.0%, w / v) and 2 mL of sodium phosphate buffer solution (0.2 mol / L, pH 6.8) were mixed, 0.8 mL of sample solution (5 mg / mL) was added, reacted at 50° C. for 20 min, cooled to room temperature, and then 2 mL of trichloroacetic acid solution (10%, w / v) was added, centrifuged at 5000 r / min for 10 min, 2 mL of supernatant was taken, 400 μL of ferric chloride solution (0.1%, w / v) and 2 mL of deionized water were added and mixed evenly, and the absorbance at 700 nm was measured after being placed away from light for 10 min.
[0059] The results are as follows Figure 4 As shown, compared with whey protein that has only undergone a single treatment (Comparative Example 1), the reducing power of the Maillard reactant prepared by whey protein and adding yam polysaccharide (Example 1 and Comparative Examples 2-4) is significantly increased; when the mixing mass ratio of yam polysaccharide to whey protein is 4:8, the reducing power of the prepared Maillard reactant is the highest, which is about 5 times that of whey protein that has only undergone a single treatment; and it is significantly higher than the reducing power of the Maillard reactants prepared by other mixing mass ratios.
[0060] (5) Determination of emulsification properties
[0061] The five reactants prepared in Example 1 and Comparative Examples 1-4 were used as samples and diluted with deionized water to a sample protein concentration of 1 mg / mL; the five reactant samples were mixed with soybean oil in a mass ratio of 3:1 in a 50 mL centrifuge tube, homogenized at 10000 rpm / min for 3 min, 50 μL was quickly drawn from the bottom of the emulsion and placed in 5 mL of a 0.1% SDS solution, the SDS solution was used as a blank group, and the absorbance was measured at 500 nm, recorded as A0; after standing for 10 min, the above method was repeated, and the absorbance value after 10 min was measured at 500 nm, recorded as A 10 .
[0062]
[0063] Among them, DF is the dilution factor: 100, θ is the oil phase ratio: 0.25, Dish diameter: 1 cm, C: sample concentration: 0.001 g / ml.
[0064] The results are as follows Figure 5 As shown, compared with whey protein that has only undergone a single treatment (Comparative Example 1), the emulsifying activity of the Maillard reactant prepared by adding whey protein and yam polysaccharide (Example 1 and Comparative Examples 2-4) first increases and then decreases, indicating that yam polysaccharide can effectively improve the emulsifying activity of whey protein, and when the mixing mass ratio of yam polysaccharide to whey protein is 4:3, the prepared Maillard reactant has the strongest emulsifying activity.
[0065] The results are as follows Figure 6 As shown, when the mixing mass ratio of yam polysaccharide to whey protein is 4:3 and 4:5, the emulsification stability of the prepared Maillard reaction product is significantly lower than that of the single-treated whey protein, and when the mixing mass ratio of yam polysaccharide to whey protein is 4:1 and 4:8, the emulsification stability of the prepared Maillard reaction product is
[0066] (6) Determination of foaming properties
[0067] The five reactants prepared in Example 1 and Comparative Examples 1-4 were used as samples and diluted with deionized water to a sample protein concentration of 1 mg / mL; 20 mL of each of the five reactant samples was placed in a 50 mL centrifuge tube and homogenized at 10000 rpm / min for 2 min, and the total volume of the liquid and foam in the centrifuge tube was immediately measured, recorded as V0 (mL); after standing for 30 min, the total volume of the liquid and foam in the centrifuge tube was measured again, recorded as V 30 .
[0068]
[0069]
[0070] Where V represents the sample volume.
[0071] The results are as follows Figure 7-8 As shown, compared with whey protein that has only undergone a single treatment (Comparative Example 1), the foaming activity and foaming stability of the Maillard reactant prepared by whey protein and adding yam polysaccharide (Example 1 and Comparative Examples 2-4) are continuously improved with the increase of the mass of yam polysaccharide; it shows that yam polysaccharide and whey protein undergo a cross-linking reaction, and the presence of sugar molecules can increase the viscosity of the complex system, thereby enhancing the foaming ability of the yam polysaccharide-whey protein Maillard reactant; among them, when the mixing mass ratio of yam polysaccharide to whey protein is 4:8, the emulsification activity and emulsification stability of the prepared Maillard reactant are optimal.
[0072] (7) Determination of protein solubility
[0073] The five reactants prepared in Example 1 and Comparative Examples 1-4 were used as samples and diluted with deionized water to a sample protein concentration of 1 mg / mL; the solutions of the five reactant samples were respectively placed under a constant temperature condition of 25° C. and oscillated for 1 h. After oscillation, they were centrifuged at a speed of 1000 rpm / min for 25 min to obtain separated undissolved proteins and supernatants; the protein content in the supernatant was determined by the Folin phenol method, and the experiment was repeated 3 times for each sample to ensure the accuracy of the results. The solubility was expressed as NSI (nitrogen solubility index);
[0074] NSI = (protein content in supernatant / protein content in sample) × 100%
[0075] The results are as follows Fig. 9 As shown, compared with whey protein (Comparative Example 1) that has only undergone a single treatment, the protein solubility of the Maillard reactant prepared by whey protein and adding yam polysaccharide (Example 1 and Comparative Examples 2-4) is significantly improved, indicating that yam polysaccharide reacts with whey protein, introduces hydrophilic hydroxyl groups, and effectively improves the technical problem of low solubility of whey protein. As the mass ratio of yam polysaccharide added increases, the solubility of the prepared Maillard reactant shows an upward trend. When the mixing mass ratio of yam polysaccharide to whey protein is 4:8, the protein solubility of the prepared Maillard reactant reaches a maximum of 71.92%.
[0076] The contents not described in detail in the specification of the present invention are well-known technologies for those skilled in the art. Although the present invention has been disclosed as above with preferred embodiments, it is not intended to limit the present invention. Anyone familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention should be based on the definition of the claims.
Claims
1. A method for preparing a yam polysaccharide-whey protein Maillard reaction product, characterized in that: The preparation method comprises the following steps: S1: freshly washed and peeled yam and water were homogenized in a volume ratio of 1:10, α-amylase was added thereto, the mixture was in a water bath at 80°C for 4 hours, cooled to room temperature, filtered, centrifuged at 8000 rpm for 20 minutes, the precipitate was discarded, and the supernatant was taken to obtain crude yam polysaccharide supernatant; S2: adding alkaline protease to the crude yam polysaccharide supernatant obtained in S1, incubating in a water bath at 55°C for 6 hours, cooling and concentrating, adding 3 times the volume of anhydrous ethanol, washing, re-dissolving and freeze-drying to obtain yam polysaccharide powder; S3: The yam polysaccharide powder obtained in S2 is uniformly mixed with whey protein in a certain mass ratio, the pH value of the mixture is adjusted with 0.2 mol hydrochloric acid, and the mixture is hydrated overnight in a 4°C refrigerator, and reacted at 55°C and 130 r / min for 5 hours to obtain a yam polysaccharide-whey protein Maillard reaction product.
2. The preparation method according to claim 1, characterized in that: The amount of α-amylase added in S1 was 20 μ / mL.
3. The preparation method according to claim 1, characterized in that: The amount of alkaline protease added in S2 is 100 μ / mL.
4. The preparation method according to claim 1, characterized in that: The concentration of whey protein in S3 is 10 mg / mL.
5. The preparation method according to claim 1, characterized in that: The mixing mass ratio of the yam polysaccharide powder and whey protein in S3 is 4:
8.
6. The preparation method according to claim 1, characterized in that: The pH value described in S3 is 7.
0.
7. A yam polysaccharide-whey protein Maillard reaction product, characterized in that: The medicinal polysaccharide-whey protein Maillard reaction product is obtained by the preparation method described in any one of claims 1 to 6.
8. Application of the yam polysaccharide-whey protein Maillard reaction product according to claim 7 in the food field and the medical field.