A method for preparing a highly dispersible and highly emulsifying protein emulsifier and its application in cosmetics.

Abstract

This invention discloses a method for preparing a highly dispersible and highly emulsifying protein emulsifier and its application in cosmetics. The method includes: dispersing pumpkin seed protein isolate in water to obtain a protein solution; adjusting the pH of the protein solution to alkaline and heating it; then adjusting the pH of the preheated protein solution to 6-7 with acid; and freeze-drying to obtain improved pumpkin seed protein powder. The obtained pumpkin seed protein powder is then dispersed in deionized water and magnetically stirred to obtain a pumpkin seed protein solution. An oil phase is added to the obtained pumpkin seed protein solution, and homogenization is performed to prepare the highly dispersible and highly emulsifying protein emulsifier. The pumpkin seed protein powder of this invention exhibits good emulsifying properties; 1 wt% of the protein dispersion can emulsify 80% of the oil phase. Furthermore, the prepared ordinary emulsion, after being stored at 45°C for 30 days, 60°C for 15 days, and 90°C for 6 hours, showed no oil release or demulsification. Centrifugation at 10000g for 30 minutes also showed no oil release or demulsification.

Description

Technical Field

[0001] This invention belongs to the fields of pharmaceuticals, cosmetics and food applications, and specifically relates to a method for preparing a highly dispersible and highly emulsifying protein emulsifier and its application in cosmetics. Background Technology

[0002] Pumpkin seeds, as a major by-product of pumpkin, are rich in protein, oil, vitamins, etc. However, the current development of pumpkin seeds mainly focuses on snack foods and pumpkin seed oil, while pumpkin seed cake, which is rich in protein, is sold at a low price as a by-product, resulting in a great waste of resources. Therefore, developing pumpkin seed protein from pumpkin seed cake has important social significance and economic value.

[0003] Pumpkin seed protein contains all eight essential amino acids for the human body, including histidine, which is essential for children. The content exceeds the standards set by the WHO (World Health Organization), giving it extremely high nutritional value. Current research on pumpkin seed protein mainly focuses on the enzymatic preparation of pumpkin seed polypeptides and their antioxidant properties, as well as the isolation and purification of pumpkin seed antifungal proteins.

[0004] Plant proteins have garnered increasing attention as emulsifiers due to their excellent biocompatibility, biodegradability, and emulsion stability. To date, various protein molecules (such as zein, soy protein isolate, and wheat protein) have been used to stabilize emulsions. However, these natural plant proteins often exhibit poor dispersibility, resulting in unsatisfactory emulsifying performance in practical applications.

[0005] Currently, the main methods for protein modification include enzymatic methods, thermal induction methods, and alkali induction methods. However, there are no reports on preparing a protein emulsifier with high dispersibility and high emulsifying properties by combining alkali induction with thermal induction of pumpkin seed protein. Summary of the Invention

[0006] The purpose of this section is to outline some aspects of embodiments of the present invention and to briefly describe some preferred embodiments. Simplifications or omissions may be made in this section, as well as in the abstract and title of this application, to avoid obscuring the purpose of these documents; however, such simplifications or omissions should not be construed as limiting the scope of the invention.

[0007] In view of the problems existing in the above and / or prior art, the present invention is proposed.

[0008] Therefore, the purpose of this invention is to overcome the shortcomings of the prior art and provide a method for preparing a protein emulsifier with high dispersibility and high emulsifying properties.

[0009] To solve the above-mentioned technical problems, the present invention provides the following technical solution: a method for preparing a highly dispersible and highly emulsifying protein emulsifier, comprising,

[0010] Pumpkin seed protein was dispersed in water to obtain a protein solution. The pH of the protein solution was adjusted to alkaline and heated. Then, the pH of the preheated protein solution was adjusted to 6-7 with acid. After freeze-drying, the improved pumpkin seed protein powder was obtained.

[0011] The obtained pumpkin seed protein powder was dispersed in deionized water and magnetically stirred to obtain a pumpkin seed protein solution.

[0012] An oil phase was added to the obtained pumpkin seed protein solution, and homogenization was carried out to prepare a protein emulsifier with high dispersibility and high emulsification.

[0013] As a preferred embodiment of the preparation method of the present invention, the step of dispersing pumpkin seed protein isolate in water to obtain a protein solution, wherein the mass fraction of the protein solution is 0.5-5%.

[0014] In a preferred embodiment of the preparation method described in this invention, the protein solution is adjusted to alkaline and then heated, wherein the alkaline pH is 10.0 to 12.0.

[0015] In a preferred embodiment of the preparation method described in this invention, the heating temperature is 70–100°C and the heating time is 60–240 min.

[0016] In a preferred embodiment of the preparation method described in this invention, the mass fraction of protein in the pumpkin seed protein solution is 0.5-5%.

[0017] As a preferred embodiment of the preparation method described in this invention, the oil phase includes sunflower seed oil, olive oil, caprylic / capric triglyceride (GTCC), isohexadecane, isododecane, D5 silicone oil, liquid paraffin, and squalane.

[0018] As a preferred embodiment of the preparation method of the present invention, an oil phase is added to the obtained pumpkin seed protein solution, wherein the volume fraction of the oil phase is 10% to 80%.

[0019] In a preferred embodiment of the preparation method described in this invention, the homogenization speed is 10000 rpm to 18000 rpm, and the homogenization time is 1 min to 5 min.

[0020] Another objective of this invention is to overcome the shortcomings of the prior art and provide a method for preparing a highly dispersible and highly emulsifying protein emulsifier.

[0021] Another objective of this invention is to overcome the shortcomings of the prior art and provide a highly dispersible and highly emulsifying protein emulsifier for use in the food, pharmaceutical, and cosmetic fields.

[0022] Beneficial effects of this invention:

[0023] (1) The improved pumpkin seed protein powder preparation process of this invention is simple, the equipment used is common, and it is suitable for industrial production. At the same time, the pumpkin seed protein powder obtained has excellent dispersibility and solubility, its dispersion index is increased by 1.95 times, and it can be stably dispersed at room temperature for more than 30 days. The modified protein powder is completely wetted within 5 seconds, while the unmodified protein cannot be completely wetted after 24 hours.

[0024] (2) The pumpkin seed protein powder of the present invention has good emulsification performance. 1 wt% of protein dispersion can emulsify 80% of the oil phase. In addition, the prepared ordinary emulsion did not show oil release or demulsification when stored at 45°C for 30 days, 60°C for 15 days, and 90°C for 6 hours. No oil release or demulsification was also observed when centrifuged at 10000g for 30 minutes.

[0025] (3) The stable emulsion system of pumpkin seed protein powder of the present invention has high biosafety, which avoids the use of surfactants and is in line with the concept of green and sustainable development. Attached Figure Description

[0026] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort. Wherein:

[0027] Figure 1 The images show the appearance of the protein dispersions obtained in Examples 1-3 and Comparative Examples 1-4.

[0028] Figure 2 The images show the wettability of the lyophilized protein powders obtained in Example 1 and Comparative Example 1.

[0029] Figure 3 The protein three-phase contact angle diagrams obtained in Examples 1-3 and Comparative Examples 1-4 are shown.

[0030] Figure 4 The images show the appearance of the emulsions obtained from Examples 1-3, Comparative Examples 1-5, and unmodified soybean protein.

[0031] Figure 5 Microscopic images of the emulsions obtained in Examples 1-3;

[0032] Figure 6 The images show the appearance of the modified protein dispersions obtained in Examples 4-5 and Comparative Example 6;

[0033] Figure 7 The images show the appearance of the emulsions obtained in Examples 9-11 and Comparative Example 9;

[0034] Figure 8 The images show the appearance of Examples 9 and Comparative Example 10 after being stored at 45°C for 30 days.

[0035] Figure 9 The high-temperature stability graph for Example 9 is shown.

[0036] Figure 10 The following are the stability graphs of 10000g centrifugation for Example 9 and Comparative Example 10;

[0037] Figure 11 The images show the appearance of the emulsions obtained in Example 12 and Comparative Example 11;

[0038] Figure 12 These are microscopic images of the emulsions obtained in Examples 16-20. Detailed Implementation

[0039] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the specific embodiments of the present invention will be described in detail below with reference to the examples in the specification.

[0040] Many specific details are set forth in the following description in order to provide a full understanding of the invention. However, the invention may also be practiced in other ways different from those described herein, and those skilled in the art can make similar extensions without departing from the spirit of the invention. Therefore, the invention is not limited to the specific embodiments disclosed below.

[0041] Secondly, the term "one embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in different places in this specification does not necessarily refer to the same embodiment, nor is it a single or selective embodiment that is mutually exclusive with other embodiments.

[0042] Unless otherwise specified, the materials and reagents used in this embodiment are commercially available.

[0043] Preparation of pumpkin seed protein isolate: Specifically, pumpkin seed powder was added to petroleum ether at a material-to-liquid ratio of 1:4 g / mL, and magnetically stirred in a water bath at 40℃ for 4 hours. The mixture was then filtered, and the supernatant was removed. This process was repeated three times to obtain defatted pumpkin seed powder. The defatted pumpkin seed powder was dried and added to deionized water at a material-to-liquid ratio of 1:30 g / mL. The pH was adjusted to 10, and extraction was performed using ultrasound-assisted extraction for 3 hours, followed by mechanical stirring-assisted extraction for another 3 hours. The mixture was centrifuged at 8000 rpm for 10 minutes to obtain the supernatant. The supernatant was adjusted to approximately pH 4.5, centrifuged at 8000 rpm for 10 minutes, and the precipitate was collected. The precipitate was washed with water until neutral and then freeze-dried to obtain pumpkin seed protein isolate.

[0044] Protein dispersibility test: Dispersions of proteins modified at different temperatures (10 mg / mL) were packaged in glass sample vials and photographed at 0 hours, 1 day, 7 days, 14 days and 30 days. The supernatant was aspirated at each time point and the supernatant protein concentration of each sample was determined using a BCA kit.

[0045] Dispersibility (%) = supernatant protein concentration / initial dispersion protein concentration × 100%, each sample was measured 3 times.

[0046] Stability experiment of modified pumpkin seed protein powder stabilized emulsion: The prepared modified pumpkin seed protein dispersion was mixed with the oil phase and homogenized to prepare a protein emulsion.

[0047] The prepared protein emulsions were placed in room temperature, high temperature and centrifuge respectively, and their appearance was observed and optical microscope images were taken regularly.

[0048] Example 1

[0049] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0050] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0051] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0052] Example 2

[0053] The difference between this embodiment and Embodiment 1 is that the heating temperature in step 1 is adjusted, specifically:

[0054] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 85°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0055] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0056] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0057] Example 3

[0058] The difference between this embodiment and Embodiment 1 is that the heating temperature in step 1 is adjusted, specifically:

[0059] 1) Dissolve pumpkin seed protein isolate in deionized water to prepare a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 100℃ for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0060] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0061] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0062] Comparative Example 1

[0063] The difference between this comparative example and Example 1 is that the emulsion was prepared using untreated pumpkin seed protein, specifically:

[0064] 1) Dissolve pumpkin seed protein isolate in deionized water to prepare a pumpkin seed protein isolate dispersion, wherein the mass fraction of protein is 1 wt%.

[0065] 3) Olive oil was added to the pumpkin seed protein isolate dispersion to make the oil phase volume fraction 80% (v / v), and then the mixture was sheared at 16000 rpm for 2 min using a homogenizer. No emulsion could be obtained.

[0066] Comparative Example 2

[0067] The difference between this comparative example and Example 1 is that the heating temperature in step 1 is adjusted, specifically:

[0068] 1) Dissolve pumpkin seed protein isolate in deionized water to prepare a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, stir at room temperature for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, obtain alkali-modified pumpkin seed protein powder.

[0069] 2) Dissolve the alkaline pumpkin seed protein powder separately in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of the protein is 1 wt%.

[0070] 3) Olive oil was added to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then the mixture was homogenized and sheared at 16000 rpm for 2 min, but no emulsion could be obtained.

[0071] Comparative Example 3

[0072] The difference between this comparative example and Example 1 is that the heating temperature in step 1 is adjusted, specifically:

[0073] 1) Dissolve pumpkin seed protein in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 55°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0074] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0075] 3) Olive oil was added to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then the mixture was homogenized and sheared at 16000 rpm for 2 min, but no emulsion could be obtained.

[0076] Comparative Example 4

[0077] The difference between this comparative example and Example 1 is that the pH of the solution in step 1 is adjusted, specifically:

[0078] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 6.5 and heat at 70°C for 2 hours. After cooling, freeze-dry the protein solution to obtain individually thermally modified pumpkin seed protein powder.

[0079] 2) Dissolve the separately thermally modified pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of the protein is 1 wt%.

[0080] 3) Olive oil was added to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then the mixture was homogenized and sheared at 16000 rpm for 2 min, but no emulsion could be obtained.

[0081] Comparative Example 5

[0082] The difference between this comparative example and Example 1 is the type of regulatory protein, specifically:

[0083] 1) Soy protein was dissolved in deionized water to prepare a soy protein solution with a protein mass fraction of 1 wt%. The pH of the solution was adjusted to 12, and the solution was heated at 70°C for 2 hours. Then, the pH of the cooled protein solution was adjusted to 6.5 with HCl solution, and the modified soy protein powder was obtained by freeze drying.

[0084] 2) Dissolve soybean protein powder in deionized water to prepare a soybean protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0085] 3) Olive oil was added to the soybean protein dispersion to make the oil phase volume fraction 80% (v / v), and then the mixture was sheared at 16000 rpm for 2 min using a homogenizer, but no emulsion could be obtained.

[0086] The dispersibility of the proteins obtained in Examples 1 to 3 and Comparative Examples 1 to 5 and the particle size of the prepared emulsions were tested, and the results are shown in Table 1.

[0087] Table 1

[0088]

[0089] As shown in Table 1 and Figure 1 and Figure 2 As shown, after alkaline heat treatment, the dispersibility of the protein was significantly improved, increasing from 40.4% for the unmodified protein to a maximum of 78.7%, an improvement of nearly 2 times. The wettability of the protein was also significantly improved; the modified protein was completely wetted within 5 seconds, while the unmodified protein could not be completely wetted even after 24 hours.

[0090] At the same time Figure 3 It can be seen that the surface hydrophilicity and hydrophobicity of proteins are improved by alkaline heat treatment, making them more hydrophilic.

[0091] from Figure 4 and Figure 5It was found that untreated pumpkin seed protein isolate at a concentration of 1 wt% could not emulsify 80% of olive oil, nor could heat-modified pumpkin seed protein isolate alone, nor could alkali-modified pumpkin seed protein isolate alone emulsify 80% of olive oil. However, after alkali heat treatment, at temperatures above 55℃, 80% of olive oil could be well stabilized. But at temperatures below or equal to 55℃, 80% of olive oil could not be stabilized. This indicates that not all high-temperature treatments yield good emulsification; only sufficiently high temperatures can achieve a good emulsification effect.

[0092] At the same time, from Figure 4 It can be seen that untreated soy protein can emulsify 80% of olive oil, but the emulsion stability is poor, and oil leakage occurs quickly after emulsification, making it impossible to obtain a stable emulsion. In contrast, the emulsifying properties of soy protein treated with alkali heat were not improved; using 80% olive oil as the inner oil phase, it was impossible to form an emulsion at all. This indicates that using alkali heat treatment to improve the emulsifying properties of protein is not suitable for all proteins.

[0093] Example 4

[0094] The difference between this embodiment and Embodiment 1 is that the heating time is adjusted, specifically:

[0095] 1) Dissolve pumpkin seed protein isolate in deionized water to prepare a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 1 hour, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0096] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0097] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0098] Example 5

[0099] The difference between this embodiment and Embodiment 1 is that the heating time is adjusted, specifically:

[0100] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 4 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0101] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0102] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0103] Comparative Example 6

[0104] The difference between this embodiment and Embodiment 1 is that the heating time is adjusted, specifically:

[0105] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 0.5 hours, then adjust the pH of the cooled protein solution to 6.5 with HCl solution, and freeze-dry to obtain modified pumpkin seed protein powder.

[0106] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0107] 3) Olive oil was added to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then the mixture was homogenized and sheared at 16000 rpm for 2 min, but no emulsion could be obtained.

[0108] like Figure 6 As shown, when the alkaline heat treatment time is too short, the dispersibility and emulsifying properties of the modified pumpkin seed protein are not significantly improved, and the difference from the untreated protein is not significant. This indicates that alkaline heat treatment requires sufficient time to open the tertiary structure of the protein, thereby improving its emulsifying and dispersible properties.

[0109] Example 6

[0110] The difference between this embodiment and Example 1 is that the protein concentration is adjusted, specifically:

[0111] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 5 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0112] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0113] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0114] Example 7

[0115] The difference between this embodiment and Example 1 is that the protein concentration is adjusted, specifically:

[0116] 1) Dissolve pumpkin seed protein in deionized water to obtain a protein solution with a protein mass fraction of 0.5 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0117] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0118] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0119] Comparative Example 7

[0120] The difference between this embodiment and Example 1 is that the protein concentration is adjusted, specifically:

[0121] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 10 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0122] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0123] 3) Olive oil was added to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then the mixture was homogenized and sheared at 16000 rpm for 2 min, but no emulsion could be obtained.

[0124] Examples 6-7 and Comparative Example 7 show that when the protein concentration is too high during alkaline heat treatment, the dispersibility and emulsifying properties of the modified pumpkin seed protein are not significantly improved, and the difference from the untreated protein is not significant. This indicates that the protein concentration during alkaline heat treatment should not be too high, otherwise it will weaken the effect of alkaline heat treatment.

[0125] Example 8

[0126] The difference between this embodiment and Embodiment 1 is that the pH of the solution is adjusted, specifically:

[0127] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 10, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0128] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0129] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0130] Comparative Example 8

[0131] The difference between this embodiment and Embodiment 1 is that the pH of the solution is adjusted, specifically:

[0132] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 9, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0133] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0134] 3) Olive oil was added to the pumpkin seed protein dispersion to make the oil phase volume fraction 80% (v / v), and then the mixture was homogenized and sheared at 16000 rpm for 2 min, but no emulsion could be obtained.

[0135] From Example 8 and Comparative Example 8, it can be concluded that when alkaline heat treatment is performed, the pH value of the solution should not be too low. If the pH is too low, the tertiary structure of the protein cannot be opened, and the dispersibility and emulsification properties of the modified pumpkin seed protein cannot be significantly improved, and there is little difference from the untreated protein.

[0136] Example 9

[0137] The difference between this embodiment and Embodiment 1 is that the oil-to-water ratio of the emulsion is adjusted, specifically:

[0138] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0139] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0140] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0141] Example 10

[0142] The difference between this embodiment and Embodiment 1 is that the oil-to-water ratio of the emulsion is adjusted, specifically:

[0143] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0144] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0145] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 60% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0146] Example 11

[0147] The difference between this embodiment and Embodiment 1 is that the oil-to-water ratio of the emulsion is adjusted, specifically:

[0148] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0149] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0150] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 75% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0151] Comparative Example 9

[0152] The difference between this embodiment and Embodiment 1 is that the oil-to-water ratio of the emulsion is adjusted, specifically:

[0153] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0154] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0155] 3) Olive oil was added to the pumpkin seed protein dispersion to make the oil phase volume fraction 85% (v / v), and then the mixture was sheared at 16000 rpm for 2 min using a homogenizer, but no emulsion could be obtained.

[0156] Comparative Example 10

[0157] The difference between this embodiment and Embodiment 9 is that the emulsion is prepared using untreated pumpkin seed protein, specifically:

[0158] 1) Dissolve pumpkin seed protein isolate in deionized water to prepare pumpkin seed protein dispersion, wherein the mass fraction of protein is 1 wt%.

[0159] 2) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0160] From Examples 9-11 and Comparative Example 9 and Figure 7 As shown, after alkaline heat treatment, the protein has a good emulsifying effect. When the protein concentration is 1 wt%, it can emulsify up to 80% of olive oil. When the oil phase volume fraction is 85%, the emulsion cannot be stabilized.

[0161] from Figure 8 It can be seen that the emulsion prepared in Example 9 can be stable at 45°C for 30 days, while the emulsion obtained in Comparative Example 10 begins to leak oil after 7 days at 45°C, and by 30 days the oil leakage has become very serious.

[0162] from Figure 9It can be seen that the emulsion prepared in Example 9 can be stable at 60°C for 15 days without oil leakage or stratification, and at 90°C for 6 hours without oil leakage or stratification, indicating that the emulsion prepared by modified pumpkin seed protein has good high-temperature stability.

[0163] from Figure 10 It can be seen that the emulsion prepared in Example 9 can be centrifuged at 10,000g for 30 minutes without oil leakage or demulsification, while the emulsion obtained in Comparative Example 10 showed oil leakage and demulsification after centrifugation at 10,000g for 10 minutes, indicating that the emulsion prepared by modified pumpkin seed protein has good centrifugal stability.

[0164] Example 12

[0165] The difference between this embodiment and Embodiment 9 is that the protein mass fraction in step 2 is adjusted, specifically:

[0166] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0167] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 0.125 wt%.

[0168] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0169] Example 13

[0170] The difference between this embodiment and Embodiment 9 is that the protein mass fraction in step 2 is adjusted, specifically:

[0171] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0172] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 0.25 wt%.

[0173] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0174] Example 14

[0175] The difference between this embodiment and Embodiment 9 is that the protein mass fraction in step 2 is adjusted, specifically:

[0176] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0177] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 4 wt%.

[0178] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0179] Example 15

[0180] The difference between this embodiment and Embodiment 9 is that the protein mass fraction in step 2 is adjusted, specifically:

[0181] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0182] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 10 wt%.

[0183] 3) Add olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0184] Comparative Example 11

[0185] The difference between this embodiment and Embodiment 9 is that the protein mass fraction in step 2 is adjusted, specifically:

[0186] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 70°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0187] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 0.0625 wt%.

[0188] 3) Adding olive oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then using a homogenizer to shear at 16000 rpm for 2 min, a stable protein emulsion could not be obtained.

[0189] From Examples 12-13 and Comparative Example 11 and Figure 11 As shown, after alkaline heat treatment, protein has a good emulsifying effect. When the protein concentration is higher than 0.125%, it can stabilize 50% of olive oil. However, when the protein concentration is too low, below 0.125%, it cannot emulsify the emulsion, indicating that a certain amount of protein is required to stabilize the oil phase.

[0190] Example 16

[0191] The difference between this embodiment and Embodiment 9 is that the modification temperature and the type of oil phase are adjusted, specifically:

[0192] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 85°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0193] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 10 wt%.

[0194] 3) Add GTCC to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0195] Example 17

[0196] The difference between this embodiment and Embodiment 9 is that the modification temperature and the type of oil phase are adjusted, specifically:

[0197] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 85°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0198] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 10 wt%.

[0199] 3) Add isohexadecane to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0200] Example 18

[0201] The difference between this embodiment and Embodiment 9 is that the modification temperature and the type of oil phase are adjusted, specifically:

[0202] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 85°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0203] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 10 wt%.

[0204] 3) Add liquid paraffin to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0205] Example 19

[0206] The difference between this embodiment and Embodiment 9 is that the modification temperature and the type of oil phase are adjusted, specifically:

[0207] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 85°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0208] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 10 wt%.

[0209] 3) Add sunflower seed oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain an emulsion.

[0210] Example 20

[0211] The difference between this embodiment and Embodiment 9 is that the modification temperature and the type of oil phase are adjusted, specifically:

[0212] 1) Dissolve pumpkin seed protein isolate in deionized water to obtain a protein solution with a protein mass fraction of 1 wt%. Adjust the pH of the solution to 12, heat at 85°C for 2 hours, and then adjust the pH of the cooled protein solution to 6.5 with HCl solution. After freeze drying, the modified pumpkin seed protein powder is obtained.

[0213] 2) Dissolve pumpkin seed protein powder in deionized water to prepare a pumpkin seed protein dispersion, wherein the mass fraction of protein is 10 wt%.

[0214] 3) Add D5 silicone oil to the pumpkin seed protein dispersion to make the oil phase volume fraction 50% (v / v), and then use a homogenizer to shear at 16000 rpm for 2 min to obtain the emulsion.

[0215] From Examples 16-20, combined with Figure 12 As shown, the modified protein has excellent emulsifying properties and can emulsify different oil phases.

[0216] This invention is the first to use alkaline heat treatment of pumpkin seed protein isolate as an emulsifier in oil-in-water emulsions to prepare emulsions with different oil-to-water ratios. The dispersibility, wettability, and emulsifying properties of the protein were investigated by adjusting the modification temperature, pH, modification time, and protein concentration during modification. The results show that, compared to unmodified pumpkin seed protein isolate, the modified pumpkin seed protein powder exhibits superior performance in terms of dispersibility, wettability, and emulsifying properties. Furthermore, the stable emulsion prepared by this method demonstrates excellent high-temperature and centrifugal stability, making it a promising green oil-in-water emulsifier.

[0217] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all such modifications or substitutions should be covered within the scope of the present invention.

Claims

1. A method for preparing a highly dispersible and highly emulsifying protein emulsifier, characterized in that: include, Pumpkin seed protein was dispersed in water to obtain a protein solution. The pH of the protein solution was adjusted to alkaline and heated. Then, the pH of the preheated protein solution was adjusted to 6-7 with acid. The improved pumpkin seed protein powder was obtained by freeze drying. The alkaline pH was 10.0-12.

0. The heating temperature was 70-100℃ and the heating time was 60-240 min. The obtained pumpkin seed protein powder was dispersed in deionized water and magnetically stirred to obtain a pumpkin seed protein solution with a protein mass fraction of 0.5-5%. An oil phase was added to the obtained pumpkin seed protein solution, and homogenization was carried out to prepare a protein emulsifier with high dispersibility and high emulsification. The oil phase had a volume fraction of 10% to 80%, and the oil phase included sunflower seed oil, olive oil, caprylic / capric triglyceride (GTCC), isohexadecane, isododecane, D5 silicone oil, liquid paraffin, and squalane.

2. The preparation method according to claim 1, characterized in that: The homogenization speed is 10,000 rpm to 18,000 rpm, and the homogenization time is 1 min to 5 min.

3. A highly dispersible and highly emulsifying protein emulsifier prepared by the preparation method described in claim 1 or 2.

4. The use of the highly dispersible and highly emulsifying protein emulsifier of claim 3 in the preparation of food, pharmaceuticals and cosmetics.

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