Egg white protein nanogel as well as preparation method and application thereof

Abstract

The invention discloses a preparation method of egg white protein nanogel. The preparation method comprises the following steps: firstly, adding an ethanol solution with the volume fraction of 0.1-11.5% into egg white liquid, and diluting until the protein concentration is 1-2%; the egg white protein ethanol solution is subjected to microwave and ultrasonic combined treatment, the treatment time is set to be 3-7 min, ultrasonic is in an intermittent mode, the ultrasonic power is 200-800 W, and the microwave heating temperature is 65-75 DEG C. And finally, removing ethanol in the solution subjected to microwave ultrasonic treatment to obtain the egg white protein nanogel particles. The method is different from an existing'bottom-to-top 'preparation method and comprises the following steps: firstly, carrying out mild denaturation on protein by adopting low-concentration ethanol, changing a protein structure and an intermolecular acting force to expose a hydrophobic structure and expand the structure, and then moderately gathering and crushing the protein into microgel by utilizing an ultrasonic cavitation effect and a microwave heat effect; the nanogel has smaller particle size, higher system stability and interface stability, and the preparation method is simple and low in cost.

Description

Technical Field

[0001] This invention relates to the field of food processing, and in particular to an egg white protein nanogel, its preparation method, and its application. Background Technology

[0002] Egg white protein is inexpensive and readily available, and is a high-quality protein with excellent functional properties (such as gelling and foaming properties). my country is one of the world's major egg producers, but its processing level is relatively low. Therefore, exploring the preparation and application of egg white nanogels can help improve the processing utilization rate of poultry eggs and promote the development of the poultry egg processing industry.

[0003] Nanogels are important functional food ingredients and drug delivery systems with broad application prospects in the food industry and other fields. Egg white molecules contain abundant functional groups. When heated above their denaturation temperature, the protein polypeptide chains unfold, followed by intermolecular association under the influence of hydrophobic and covalent bonds, further cross-linking to form a nanogel network structure. The particle size of nanogels is one of the key factors determining their function and application. Studies have shown that smaller nanogel particle sizes are more stable and better at improving interfacial properties, thereby enhancing functional properties such as foaming properties. Currently, nanogels with particle sizes around 100 nm are rarely reported. Therefore, how to effectively control the particle size of nanogels has become one of the current research hotspots. Commonly used methods for preparing nanogels include shearing, high-pressure homogenization, and microfluidics, but these are difficult to use for precisely preparing small-particle-size nanogels. Microfluidics can achieve precise control of nanogel particle size, but the cost is too high. Therefore, a new method needs to be explored to control the particle size of nanogels and improve their interfacial stability and functionality. Summary of the Invention

[0004] The purpose of this invention is to overcome the shortcomings of the existing technology and to design a method for preparing egg white protein nanogel, aiming to prepare a nanogel with small particle size, high system stability and interface stability, so as to better apply it to multiphase systems.

[0005] To achieve the above objectives, the present invention adopts the following technical solution:

[0006] A method for preparing egg white protein nanogel specifically includes the following steps:

[0007] 1) Add 0.1-11.5% ethanol solution to the egg white liquid to dilute it to a protein concentration of 1-2%.

[0008] 2) The egg white protein ethanol solution was subjected to a combination of microwave and ultrasonic treatment. The treatment time was set to 3-7 min, the ultrasonic treatment was in intermittent mode, the ultrasonic power was 200-800W, and the microwave heating temperature was 65-75℃.

[0009] 3) Remove the ethanol from the solution after microwave ultrasonic treatment to obtain egg white protein nanogel particles.

[0010] In step 1), the volume fraction of the ethanol solution is preferably 5-10%.

[0011] In step 2), the processing time is 3-7 minutes, preferably 5 minutes.

[0012] In step 2), as the ultrasonic power increases, the particle size of the gel particles first decreases and then increases. The ultrasonic power is preferably 400-700W.

[0013] In step 2), the heating temperature is 65-75℃.

[0014] In step 2), the egg white protein ethanol solution is subjected to microwave and ultrasonic combined treatment using a microwave and ultrasonic combined extractor.

[0015] In step 3), conventional methods such as rotary evaporation and heating are used to remove ethanol from the solution.

[0016] The egg white protein nanogel particles prepared by the above method have a particle size range of 105 nm to 200 nm, preferably 105 nm to 167 nm, and more preferably 105 nm to 132 nm.

[0017] The egg white protein nanogel particles can be used as emulsifiers and foaming agents in food.

[0018] This method differs from existing "bottom-up" preparation methods. This invention first uses low-concentration ethanol to mildly denature the protein, changing the protein structure and intermolecular forces, exposing the hydrophobic structure and allowing it to unfold. Then, ultrasonic cavitation and microwave thermal effects are used to moderately aggregate and break the protein into microgels.

[0019] The beneficial technical effects of the present invention are: (1) Egg white protein nanogels are prepared by ethanol-assisted microwave ultrasonic treatment of egg white protein solution, so as to achieve precise control of the particle size and properties of nanogels, and make the nanogels have smaller particle size, higher system stability and interface stability; (2) The preparation method is simple and low cost. Attached Figure Description

[0020] Figure 1 The images shown are scanning electron microscope (SEM) images of the nanogels prepared according to the present invention, wherein A is Comparative Example 1; B is Example 1; and C is Example 2.

[0021] Figure 2 The images shown are atomic force microscopy images and cross-sectional height diagrams of the nanogels prepared in this invention, wherein A is Comparative Example 1; B is Example 1; and C is Example 2.

[0022] Figure 3 The image shows the contact angle of the nanogel prepared according to the present invention, where A is Comparative Example 1; B is Example 1; and C is Example 2. Detailed Implementation

[0023] To facilitate understanding of the present invention, the present invention will be further described below with reference to embodiments, but the implementation of the present invention is not limited thereto.

[0024] A method for preparing egg white protein nanogel specifically includes the following steps:

[0025] 1) Add 0.1-11.5% ethanol solution to the egg white liquid to dilute it to a protein concentration of 1-2%.

[0026] 2) The egg white protein ethanol solution was subjected to a combination of microwave and ultrasonic treatment. The treatment time was set to 3-7 min, the ultrasonic treatment was in intermittent mode, the ultrasonic power was 200-800W, and the microwave heating temperature was 65-75℃.

[0027] 3) Remove the ethanol from the solution after microwave ultrasonic treatment to obtain egg white protein nanogel particles.

[0028] In step 1), the volume fraction of the ethanol solution affects the formation of egg white protein nanogel particles. As the volume fraction increases, the gel particle size gradually decreases. When the volume fraction reaches 12.5%, a blocky gel is obtained, and gel particles cannot be formed. The volume fraction of the ethanol solution is 0.1-11.5%, such as 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, 5.0%, 5.5%, 6.0%, 6.5%, 7.0%, 7.5%, 8.0%, 8.5%, 9.0%, 9.5%, 10.0%, 10.5%, 11.0%, 11.5%, or any two of the above values. The preferred volume fraction of the ethanol solution is 5-10%.

[0029] In step 2), the processing time includes both ultrasonic and microwave heating time. As the processing time increases, the gel particle size first decreases and then increases. The processing time is 3-7 minutes, preferably 5 minutes.

[0030] In step 2), the ultrasound is performed in intermittent mode, alternating between working and stopping. When stopped, it dissipates the large amount of heat generated during operation, such as working for 1-3 seconds and stopping for 1-3 seconds. The ultrasound process generates a significant amount of heat, and higher temperatures can lead to an increase in the size of the nanogel particles. Setting the intermittent mode effectively controls the temperature during the process.

[0031] In step 2), as the ultrasonic power increases, the particle size of the gel particles first decreases and then increases. The ultrasonic power is 200-800W, such as 200W, 300W, 400W, 500W, 600W, 700W, 800W, or any two of the above values, preferably 400-700W.

[0032] In step 2), the heating temperature is 65-75℃, such as 65℃, 70℃, 75℃, or any two of the above values ​​within the range.

[0033] In step 2), the egg white protein ethanol solution is subjected to microwave and ultrasonic combined treatment using a microwave and ultrasonic combined extractor.

[0034] In step 3), the obtained egg white protein nanogel particles have a particle size range of 105 nm to 200 nm, preferably 105 nm to 167 nm, and more preferably 105 nm to 132 nm.

[0035] In step 3), conventional methods such as rotary evaporation and heating are used to remove ethanol from the solution.

[0036] The egg white protein nanogel particles can be used as emulsifiers and foaming agents in food.

[0037] Example 1: A method for preparing an egg white protein nanogel

[0038] 1) Clean the eggs, separate the egg whites, and disperse them at 5000 rpm for 2 minutes using a high-speed disperser to improve their fluidity. Then centrifuge at 10000 rpm for 5 minutes to remove insoluble impurities and finally obtain egg white liquid.

[0039] 2) Dilute the collected egg white solution with 5% ethanol solution to a protein concentration of 2%.

[0040] 3) Add 50 mL of the diluted solution to a three-necked flask and treat it with a microwave-ultrasound combined extractor. Set the treatment time to 5 min, the ultrasound to intermittent mode (1 s on, 1 s off), the ultrasound power to 500 W, and the microwave heating temperature to 75 °C.

[0041] 4) After microwave ultrasonic treatment, the solution was rotary evaporated to remove ethanol, yielding egg white protein nanogel particles.

[0042] Examples 2-3: A method for preparing egg white protein nanogels

[0043] Except for step 2), all other steps in Examples 2-3 are the same as in Example 1.

[0044] 2) The collected egg white solution was diluted to a protein concentration of 2% with 10% (Example 2) and 7.5% (Example 3) ethanol solutions respectively.

[0045] Example 4: A method for preparing egg white protein nanogel

[0046] Example 4 is the same as Example 3 except for step 3).

[0047] 3) Add 50 mL of the diluted solution to a three-necked flask and treat it with a microwave-ultrasound combined extractor. Set the treatment time to 5 min, the ultrasound to intermittent mode (working for 1 s, stopping for 1 s), the ultrasound power to 700 W, and the microwave heating temperature to 75 °C.

[0048] Comparative Example 1: A method for preparing egg white protein nanogels

[0049] Comparative Example 1 is the same as Example 1 except for step 2).

[0050] 2) Dilute the collected egg white solution with deionized water to a protein concentration of 2%.

[0051] Comparative Example 2: A method for preparing egg white protein nanogels

[0052] Comparative Example 2 is the same as Example 1 except for step 3).

[0053] 3) Add 50 mL of the diluted solution to a three-necked flask and treat it with a microwave-ultrasonic extraction device. Set the microwave treatment time to 5 min and the microwave heating temperature to 75℃.

[0054] Comparative Example 3: A method for preparing egg white protein nanogels

[0055] Comparative Example 3 is the same as Example 1 except for step 3).

[0056] 3) Add 50 mL of the diluted solution to a three-necked flask and process it with a microwave-ultrasound combined extractor. Set the ultrasound to intermittent mode (working for 1 second and stopping for 1 second), set the ultrasound time to 5 min, and the ultrasound power to 500W.

[0057] Test example:

[0058] The following testing methods were used during the invention process to test the prepared egg white protein nanogel.

[0059] (1) Particle size potential

[0060] The particle size potential of the nanogels prepared in Examples 1-4 and Comparative Examples 1-3 was determined using a particle size and potential analyzer. The nanogels were diluted with deionized water to 0.2% (W / V), and 1 mL was added to the particle size and potential cups, respectively. The temperature was set at 25℃, the scattering angle was 90°, and the experiment was repeated 10 times. The results are shown in Table 1.

[0061] Table 1. Nanogel particle size and potential

[0062]

[0063] Compared to Comparative Example 1, the egg white protein nanogels in Examples 1-4 showed a significant reduction in particle size, indicating that the method of treating with ethanol combined with microwave ultrasound is beneficial in reducing the formation of large aggregates. Furthermore, the egg white protein nanogel in Example 2 had the smallest particle size and the lowest PDI, indicating the most uniform particle size distribution. The absolute value of the Zeta potential in the egg white protein nanogels in Examples 1-4 was increased. A high absolute value of the Zeta potential can inhibit aggregation, making the nanogel system more stable.

[0064] (2) Scanning electron microscope

[0065] 20 μL of the nanogel samples prepared in Examples 1-2 and Comparative Example 1 were respectively added to mica sheets and dried at room temperature. After platinum coating on the sample surface, the samples were observed under an accelerating voltage of 5.0 kV. Figure 1 As shown, the nanogels in Comparative Example 1 exhibited a small amount of aggregation and stacking distribution, with slightly larger particle sizes. The nanogels in Examples 1 and 2 showed more uniform distribution and smaller particle sizes, with the nanogels in Example 2 having the smallest particle size.

[0066] (3) Atomic force microscope

[0067] Take 20 μL of the nanogel samples prepared in Examples 1-2 and Comparative Example 1, respectively, and drop them onto mica sheets. Let them air dry at room temperature. Observe the morphology of the samples with different treatments in tapping mode.

[0068] like Figure 2 As shown, the atomic force scanning electron microscopy (AFM) images are consistent with the particle size and SFM results, with the nanogel in Example 2 having the smallest particle size. The cross-sectional height diagram shows that the maximum heights of Comparative Example 1, Example 1, and Example 2 are 18.5 nm, 13.4 nm, and 6.07 nm, respectively. The nanogel in Example 2 has the lowest height, indicating that ethanol combined with microwave ultrasound can regulate the size of the nanogel and reduce protein aggregation. The height of all nanogels is smaller than their diameter, indicating that they are soft particles capable of deformation on the mica sheet surface.

[0069] (4) Contact angle measurement

[0070] 100 μL of the nanogel samples prepared in Examples 1-2 and Comparative Example 1 were dropped onto a glass slide, spread evenly, and dried at 40°C to form a nanogel film. The contact angle of the samples was measured using a video optical angle analyzer. The contact angle was calculated using OneAttension software, and the results are shown in Table 3. Compared with Comparative Example 1, the contact angle of the egg white protein nanogels in Examples 1 and 2 was increased. The closer the contact angle is to 90°, the higher the interfacial stability. The contact angle of Example 2 was closest to 90°, which is due to the increased protein unfolding under the synergistic effect of ethanol and ultrasonic microwaves, exposing more hydrophobic groups and enhancing the stability of the water-oil interface.

[0071] (5) Foaming and emulsifying properties

[0072] The nanogels obtained in Examples 1, 2, and Comparative Example 1 were diluted to 10% (v / v) with deionized water. 25 mL of the diluted solution was stirred at 8000 rpm for 2 min in a high-speed homogenizer and then allowed to stand. The foam volume was recorded at 2 min and 30 min. The foaming performance (FA) and foam stability (FS) were calculated as follows:

[0073]

[0074]

[0075] Where V1 is the foam volume at 2 min; V2 is the foam volume at 30 min.

[0076] The nanogels obtained in Examples 1, 2, and Comparative Example 1 were diluted to 0.1% (V / V) with deionized water. The diluted solution was mixed with soybean oil at a ratio of 3:1 (V / V), homogenized at 10,000 rpm for 1 min with a high-speed dispersant, and then 50 μL of the emulsion was added to 5 mL of 1% w / v sodium dodecyl sulfate (SDS) aqueous solution. The absorbance was measured at 500 nm using a spectrophotometer. The SDS solution was used as a blank solution. The emulsifying properties (EC) and emulsion stability (ES) were calculated as follows:

[0077]

[0078]

[0079] Where A0 is the absorbance measured for the first time, A 10 The absorbance was measured after 10 minutes.

[0080] The results are shown in Table 2. Compared with Comparative Example 1, the egg white protein nanogels in Examples 1 and 2 showed significantly improved foaming and emulsifying properties. The smallest particle size and high interfacial stability of the nanogel in Example 1 facilitated its faster and more stable adsorption at the water-oil and gas-liquid interfaces, resulting in the highest foaming and emulsifying properties.

[0081] Table 2 Foaming and Emulsifying Properties of Nanogels

[0082] Foaming properties Foam stability emulsifying Emulsion stability Comparative Example 1 <![CDATA[43.33±0.72 c ]]> <![CDATA[64.23±0.90 c ]]> <![CDATA[63.24±0.61 c ]]> <![CDATA[50.71±0.34 c ]]> Example 1 <![CDATA[47.16±0.33 b ]]> <![CDATA[66.47±0.72 b ]]> <![CDATA[67.52±0.53 b ]]> <![CDATA[53.45±0.59 b ]]> Example 2 <![CDATA[49.09±0.41 a ]]> <![CDATA[68.18±0.83 a ]]> <![CDATA[70.11±0.68 a ]]> <![CDATA[55.10±0.42 a ]]>

[0083] Application examples

[0084] A method for preparing melt-resistant ice cream includes the following steps:

[0085] (1) By mass percentage: 11% milk powder, 10% animal cream, 0-5% egg white protein microgel (see Table 3 for details), 15% sucrose, 0.09% stabilizer, 0.11% emulsifier and 63.8% water, stir and mix to obtain a mixed liquid;

[0086] (2) The mixture is homogenized and sterilized sequentially to obtain the liquid before stirring;

[0087] (3) The pre-mixing liquid is aged at 4°C for 8-12 hours to obtain aged liquid. The aged liquid is then mixed and frozen in an ice cream machine. After freezing, it is quickly placed at -20°C to harden, thus obtaining melt-resistant ice cream.

[0088] The ice cream was placed on a perforated plate (at room temperature) and allowed to melt naturally. The mass of the melted ice cream was recorded at regular intervals. The melting rate was calculated based on the following formula, and the results are shown in Table 3.

[0089]

[0090] Where m1 is the mass of the ice cream before melting, and m2 is the mass of the ice cream after melting.

[0091] As the amount of egg white protein nanogel added increases, the melting rate of ice cream decreases. The high foaming and emulsifying properties of the microgel make the fat globule structure more stable, the foam distribution more uniform, and the resulting ice cream has a lower melting rate. In addition, the cross-linking of the microgel forms a spatial network structure, which increases the viscosity of the system, thereby restricting the movement of water molecules and reducing the melting rate of ice cream.

[0092] Table 3 Ice Cream Melting Rate

[0093]

Claims

1. A method for preparing egg white protein nanogel, characterized in that, Specifically, the following steps are included: 1) Add 0.1-11.5% ethanol solution to the egg white liquid to dilute it to a protein concentration of 1-2%. 2) The egg white protein ethanol solution was subjected to a combination of microwave and ultrasonic treatment. The treatment time was set to 3-7 min, the ultrasonic treatment was in intermittent mode, the ultrasonic power was 200-800W, and the microwave heating temperature was 65-75℃. 3) Remove the ethanol from the solution after microwave ultrasonic treatment to obtain egg white protein nanogel particles.

2. The method for preparing egg white protein nanogel according to claim 1, characterized in that, In step 1), the volume fraction of the ethanol solution is 5-10%.

3. The method for preparing egg white protein nanogel according to claim 1, characterized in that, In step 2), the ultrasonic power is 400-700W.

4. The method for preparing egg white protein nanogel according to claim 1, characterized in that, In step 2), the egg white protein ethanol solution is subjected to microwave and ultrasonic combined treatment using a microwave and ultrasonic combined extractor.

5. The method for preparing egg white protein nanogel according to claim 1, characterized in that, The egg white protein nanogel particles prepared by the above method have a particle size range of 105 nm to 200 nm.

6. Egg white protein nanogel prepared by the method according to any one of claims 1-5.

7. The use of the egg white protein nanogel particles according to claim 6 as an emulsifier or foaming agent in food.

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