A processing method for improving the emulsification properties of chicken protein
Through pH field drive technology and gluconolactone treatment, combined with low temperature and high-speed centrifugation, the emulsification characteristics of chicken protein are improved, and the problem of low water and oil holding capacity of emulsified meat products in the production process is solved, achieving better emulsification Characteristics and stability provide a theoretical basis for the development of new healthy emulsified meat products.
Patent Information
- Application Number
- CN202211605226.5
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2022-12-14
- Publication Date
- 2024-12-10
- Estimated Expiration
- 2042-12-14
AI Technical Summary
In the production, processing, storage and transportation of existing emulsified meat products, there are problems such as low water and oil holding capacity and loose quality structure. The use of high saturated fatty acids in traditional methods leads to health risks, making it difficult to develop new emulsified meat products.
The pH field-driven technology was used to dilute myofibrillar protein through phosphate buffer, and the pH was adjusted using gluconolactone, combined with low temperature and high-speed centrifugation, to improve the emulsification characteristics of chicken protein.
It improves the emulsification characteristics of chicken protein, has good repeatability of experimental results, is convenient to operate, and is cost-effective, provides a theoretical basis for the development of new emulsified meat products, and enriches the theory of muscle protein emulsification.
Smart Images
Figure CN115918769B_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of protein processing, and more particularly to a novel processing method for improving the emulsification properties of chicken protein. Background Art
[0002] As an important type of deep-processed meat products on the market, emulsified meat products still have problems such as low water and oil holding capacity and loose texture during actual production, processing, storage and transportation. At the same time, the high saturated fatty acid content of traditional emulsified meat products can easily lead to chronic diseases such as obesity, cardiovascular metabolic abnormalities and diabetes, posing a huge hidden danger to consumers' health. In the post-epidemic era, the public is no longer only concerned about the sensory flavor of the food itself, but also pays more attention to whether it is healthy and nutritious. In order to reduce consumers' concerns about emulsified meat products, the use of vegetable oils containing a high proportion of unsaturated fatty acids to replace animal fats is an effective method and research hotspot to improve the quality of emulsified meat products to a certain extent. However, due to the characteristics of vegetable oils that are easier to flow and oxidize than animal oils, the development of new emulsified meat products is restricted.
[0003] Myofibrillar protein (MP) is the most critical functional protein in muscle, accounting for 55-60% of the total protein content. As the most important "endogenous" emulsifier, the emulsifying properties of MP determine the quality and value of emulsified meat products. pH field driven technology is to first unfold the protein at extreme pH, and then refold it at neutral pH, so that the protein is in a molten state. Proteins in this state often show good gel emulsification processing characteristics. Gluconodeltalactone (GDL) is a good food-grade slow-release acid. By controlling the active H + The release rate can achieve the effect of slow acid release, which has great potential in the research of pH field driven technology for MP emulsion optimization. Summary of the Invention
[0004] Based on the above, the present invention overcomes the deficiencies of the prior art and provides a novel emulsion preparation technology that is economical, convenient, and effectively improves emulsification properties.
[0005] To achieve the above object, the present invention adopts the following technical solutions:
[0006] A novel processing method for improving the emulsification properties of chicken protein comprises the following steps:
[0007] (1) Remove the fat and fascia on the surface of animal muscles and then extract myofibrillar protein (MP);
[0008] (2) diluting the myofibrillar protein extracted in step (1) with phosphate buffered saline (PBS), adjusting the pH to 11.0 with 2M NaOH, and allowing the pH to balance to obtain an alkaline myofibrillar protein solution;
[0009] (3) adjusting the pH of the alkaline myofibrillar protein solution obtained in step (2) to 7.0 using gluconolactone (GDL), and allowing the solution to stand to achieve pH equilibrium, thereby obtaining a myofibrillar protein aggregate solution;
[0010] (4) The myofibrillar protein aggregate solution obtained in step (3) is centrifuged at high speed under low temperature, and the obtained precipitate is the improved chicken protein.
[0011] Preferably, the steps of extracting myofibrillar protein in step (1) are as follows:
[0012] The processed animal muscle is chopped into minced meat, added with standard salt solution, mixed evenly and homogenized. The filtered homogenate is centrifuged at low temperature and the precipitate is retained. The homogenization, filtration and low temperature centrifugation steps are repeated twice. The final precipitate is washed twice with low salt solution to obtain myofibrillar protein.
[0013] Furthermore, the standard salt solution formula components are: 0.1M KCl, 20mM K2HPO4 / KH2PO4, 1mM EGTA, 2mM MgCl2, pH=7.0;
[0014] The low salt solution is a 0.1M KCl solution.
[0015] Furthermore, the volume ratio of the standard salt solution to the minced meat is 2 to 8:1, more preferably 3 to 5:1; the volume ratio of the added amount of the low-salt solution to the precipitate is 2 to 8:1, more preferably 3 to 5:1.
[0016] Further, the low salt solution washing steps are as follows:
[0017] The precipitate and low-salt solution were mixed evenly and homogenized, and then centrifuged at low temperature. The homogenization and centrifugation steps were repeated once, and the final precipitate obtained was myofibrillar protein.
[0018] Furthermore, the homogenization speed is 6700 rpm / min, the time is 30 s, and the operation is performed 1 to 2 times. The low-temperature centrifugation speed is 2000 to 2500×g, the time is 10 min, and the temperature is 4°C.
[0019] Preferably, the phosphate buffer in step (2) comprises: 0.6M KCl, 50mM K2HPO4 / KH2PO4, pH=7.0; the concentration of myofibrillar protein after dilution with the phosphate buffer is 12-18 mg / ml, more preferably 12-16 mg / ml.
[0020] Preferably, in step (3), based on the alkaline myofibrillar protein solution, the added amount of gluconolactone is 0.30-0.60 wt%, more preferably 0.40-0.45 wt%, and most preferably 0.42 wt%.
[0021] Preferably, the myofibrillar protein aggregate solution in step (4) is centrifuged at a low temperature of 8000 g for 15 min at a temperature of 4°C.
[0022] It can be seen from the above technical solution that, compared with the prior art, the present invention discloses a new processing method for improving the emulsification properties of chicken protein, which has the following beneficial effects:
[0023] 1) The experimental results of the present invention are highly reproducible. The operation is convenient and the cost is low.
[0024] 2) The present invention is based on pH field driven NaOH-GDL treatment, which can effectively improve the emulsification properties of chicken protein;
[0025] 3) The present invention can provide a theoretical basis for the development of healthy new emulsified meat products and enrich the theory of muscle protein emulsification. BRIEF DESCRIPTION OF THE DRAWINGS
[0026] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are merely embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on the provided drawings without paying any creative work.
[0027] Figure 1 is the effect of NaOH-GDL treatment on the hydrophobicity of MP surface;
[0028] Figure 2 is the effect of NaOH-GDL treatment on the intrinsic fluorescence intensity of MP;
[0029] Figure 3 is the effect of NaOH-GDL treatment on the flexibility of MP molecules;
[0030] Figure 4 is the effect of NaOH-GDL treatment on MP emulsification activity;
[0031] Figure 5 is the effect of NaOH-GDL treatment on MP emulsification stability;
[0032] Figure 6 The effect of NaOH-GDL treatment on the macroscopic stability of MP emulsion. DETAILED DESCRIPTION
[0033] The following will clearly and completely describe the technical solutions of the present invention in conjunction with the embodiments of the present invention. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts shall fall within the scope of protection of the present invention.
[0034] Example
[0035] Step 1: Pre-processing the chicken breast
[0036] Use absorbent paper to wipe the surface of the chicken breast dry and remove the surface fat and fascia; then cut the pre-treated chicken breast into about 1cm 3 Use a meat grinder to chop the chicken into cubes of different sizes at 3000 rpm for 30 seconds to obtain chicken mince;
[0037] Step 2: MP extraction
[0038] The minced meat and 4 volumes of standard salt solution (0.1M KCl, 20mM K2HPO4 / KH2PO4, 1mM EGTA, 2mM MgCl2, pH = 7.0) were uniformly mixed and homogenized (6700 rpm / min, 30s×2 times), the homogenate was filtered through a double layer of medical sterile gauze and centrifuged at low temperature (2000×g, 10min, 4°C); the lower precipitate obtained by centrifugation was repeated twice; then the final precipitate was uniformly mixed with 4 volumes of low salt solution (0.1M KCl) and homogenized (6700 rpm / min, 30s×2 times), the homogenate was centrifuged at low temperature (2500×g, 10min, 4°C), and the lower precipitate was repeated once to obtain the final precipitate, which is MP;
[0039] Step 3: NaOH-GDL treatment:
[0040] Dilute the MP concentration to 12-16 mg / ml with PBS (0.6 mM KCl, 50 mM KHPO / KHPO, pH = 7.0, the same below). Adjust the pH of the MP solution to 11.0 with NaOH solution and let it stand for 10 minutes to allow the pH to balance. Add 0.42 wt% GDL to adjust the pH of the solution to a neutral pH of 7.0 and let it stand for 10 minutes to allow the pH to balance.
[0041] Step 4: Protein recovery
[0042] The protein solution treated in step 3 was centrifuged at low temperature and high speed (8000 g, 15 min, 4° C.), and the precipitate was collected to obtain the recovered protein.
[0043] Comparative Example 1
[0044] The difference from the embodiment is that:
[0045] In step 3, the MP concentration was directly diluted to 12-16 mg / ml with PBS (0.6 mM KCl, 50 mM K2HPO4 / KH2PO4, pH = 7.0, the same below) without adjusting the pH.
[0046] Comparative Example 2
[0047] The difference from the embodiment is that:
[0048] In step 3, no GDL was added and the pH was adjusted to neutral with 2 M HCl.
[0049] Experimental example
[0050] Experiment 1: Effect of NaOH-GDL treatment on MP surface hydrophobicity
[0051] The protein solution was diluted to 5 mg / ml with PBS. Then, 200 μl of bromophenol blue (BPB) solution (1 mg / ml in deionized water) was added to 1 ml of the protein solution and 1 ml of deionized water and thoroughly vortexed for 10 minutes at room temperature to serve as the treated and control groups. After centrifugation at 2000 × g for 15 minutes, the absorbance of the supernatant was measured at 595 nm.
[0052] The BPB binding protein content was calculated according to the following formula:
[0053] BPB binding amount (μg) = 20 μg (A c -A s ) / A c
[0054] Among them, A c is the absorbance of the control group, A s is the absorbance of the sample.
[0055] Experiment 2: Effect of NaOH-GDL treatment on the endogenous fluorescence intensity of MP tryptophan
[0056] Dilute the protein sample to 0.8 mg / ml in PBS. Measure the fluorescence emission spectrum of each sample. Excitation and emission wavelengths were 280 nm and 315–400 nm, respectively, with a scan speed of 100 nm / min and a slit width of 10 nm. Protect from light throughout the entire process.
[0057] Experiment 3: Effect of NaOH-GDL treatment on MP molecular flexibility
[0058] Adjust the protein concentration to 1 mg / ml with PBS. Add 250 μl of prepared trypsin (1 mg / ml in 0.05 M Tris-HCl buffer, pH 8.0) to 4 ml of diluted sample. Incubate at 38°C for 5 minutes. Terminate the reaction by adding 4 ml of 50 g / L trichloroacetic acid. Centrifuge at 4°C for 10 minutes, and measure the absorbance of each supernatant at 280 nm.
[0059] Experiment 4: Effects of NaOH-GDL treatment on MP emulsifying activity (EAI, emulsifying activity index) and emulsifying stability (ESI, emulsifying stability index)
[0060] Mix the protein solution and soybean oil in a 4:1 (v / v) ratio and homogenize at high speed (15,000 rpm / min, 40 s x 3 times) to prepare an oil-in-water emulsion. Disperse 50 μl of the fresh emulsion into 5 ml of 0.1% SDS solution and vortex thoroughly. Measure the absorbance at 500 nm. Let the remaining diluted solution stand at room temperature for 30 minutes and repeat the absorbance measurement described above.
[0061] EAI and ESI are calculated according to the following formulas:
[0062]
[0063]
[0064] Among them, A0, A 30 are the absorbance values at 500 nm corresponding to time 0 and 30 min, respectively; C is the protein concentration (g / ml); n is the dilution factor. is the ratio of oil volume fraction (v / v) to total volume.
[0065] Experiment 5: Effect of NaOH-GDL treatment on the macroscopic stability of MP emulsion
[0066] 20 ml of freshly prepared emulsion (preparation steps are the same as in Experiment 4) was pipetted into glass tubes, with each sample kept at the same height, and immediately placed in the chamber of the instrument. Scanning was then performed at 25°C at 880 nm near-infrared for 3 h.
[0067] Data analysis and graphics:
[0068] The above experimental procedures were repeated three times, and statistical analysis was performed using SAS system for Windows V8, including one-way analysis of variance (ANOVA, P < 0.05) and multivariate comparative analysis (P < 0.05). Images were drawn using Origin 2022b and Graphpad Prism 9.4.0.
[0069] Results and Discussion
[0070] 1) Effect of NaOH-GDL treatment on MP surface hydrophobicity
[0071] This test uses the bromophenol blue method to determine the number of surface hydrophobic groups. Figure 1 As shown, the surface hydrophobicity of the embodiment and comparative example 2 treated with NaOH-GDL / HCl was significantly higher than that of the untreated comparative example 1 (P<0.001). This is consistent with the view generally held by scholars at home and abroad that the unfolding and refolding stages caused by pH field-driven changes lead to molecular mismatch, exposing hidden hydrophobic residues on the polar surface. In addition, the enhanced hydrophobicity in MP can promote the formation of interfacial films, thereby improving the emulsification properties of the emulsion. In theory, there is a great correlation between the number of surface hydrophobic groups and the higher-order molecular conformation of the protein. There was no significant difference in surface hydrophobicity between the embodiment and comparative example 2 (P>0.05), indicating that under the same pH field-driven treatment (pH: 11.0→7.0), the use of different types of acids (HCl and GDL) to refold to neutral pH 7.0 had no significant effect on the higher-order conformation of MP.
[0072] 2) Effect of NaOH-GDL treatment on the intrinsic fluorescence of MP tryptophan
[0073] like Figure 2 As shown, MP treated with NaOH-GDL / HCl exhibited a greater fluorescence loss compared to Comparative Example 1. This quenching of the maximum fluorescence is attributed to MP unfolding, exposing tryptophan residues to a hydrophilic environment. Furthermore, the maximum emission wavelength of MP treated with pH field-driven treatment red-shifted by 3 nm compared to the untreated protein, indicating that MP aggregate formation leads to unfolding of the tertiary structure, consistent with the surface hydrophobicity.
[0074] 3) Effect of NaOH-GDL treatment on the flexibility of MP molecules
[0075] When studying the molecular flexibility of MP treated with NaOH-GDL, protease digestion is an effective tool to reflect the differences in conformational flexibility between solution and interface environment. Figure 3 As shown, there was no significant difference between Comparative Example 1 and Comparative Example 2 (P>0.05). Compared with the other two groups, Example 1 exhibited higher molecular flexibility (P<0.05). This suggests that by changing the acid type that refolds to neutral pH (HCl→GDL), the molecular flexibility of MP can be significantly improved, which is beneficial for obtaining better performance at the oil-water interface to promote emulsification properties. The potential mechanism may be that the NaOH-GDL treatment appropriately changes the MP conformation, thereby effectively enhancing molecular flexibility.
[0076] IV) Effect of NaOH-GDL treatment on EAI and ESI of MP emulsion
[0077] EAI is a method to characterize how much oil can be emulsified per unit protein. Figure 4 As shown in Figure 2, the emulsions prepared with NaOH-HCl and NaOH-GDL as matrices have higher EAI values than the untreated proteins. 2 / g) was significantly higher than that of the NaOH-HCl treatment (2.17m 2 / g)(P<0.05), showing better emulsification activity. The potential reason may be that the emulsion formed by MP treated with NaOH-GDL can better form an interfacial film, thereby promoting the dispersion of oil droplets.
[0078] ESI is a characterization of the storage stability of an emulsion over a certain time span. Figure 5 As shown, similar to the trend of the EAI results, the emulsions corresponding to Example 1 and Comparative Example 2 all have higher ESI values compared to Comparative Example 1. Compared with LA (66.60%), SA (93.67%) has a higher ESI value (P < 0.01), which is beneficial to the stability of the emulsion during storage. This is because the increased flexibility of protein molecules after NaOH-GDL treatment will increase the adsorption ratio of proteins on the oil droplet surface, thereby improving the emulsification activity and stability of the emulsion. At the same time, smaller protein aggregates are more easily adsorbed to the water-in-oil interface due to their higher migration and diffusion rates, thereby enhancing the performance of the emulsion.
[0079] 5) Effect of NaOH-GDL treatment on
[0080] TSI is one of the important indicators for quantifying the instability process of emulsion such as coalescence and flocculation. Figure 6 As shown in the figure, the dynamic instability of the three emulsions gradually increases with the extension of the test time. In theory, the TSI value represents the dynamic instability. The lower the TSI value, the higher the macroscopic stability of the emulsion. The TSI values of Example and Comparative Example 2 tend to stabilize after about 1800s and 3800s, respectively, while the TSI value of Comparative Example 1 is still slowly rising until 10800s. This phenomenon shows that the emulsion formed by MP after pH field driven treatment has higher overall stability than the untreated protein. In addition, the TSI value of Example after equilibrium is significantly lower than that of Comparative Example 2 (P<0.05), indicating that a more stable emulsion can be formed under NaOH-GDL treatment, and this result is consistent with the result of ESI.
[0081] Conclusion: This invention provides a novel processing method for improving the emulsification properties of chicken protein. This method, based on an optimized pH-field-driven technique, employs GDL as the acid for refolding to neutral pH. NaOH-GDL treatment alters the molecular conformation of MP, significantly increasing its flexibility and significantly enhancing the emulsion activity and stability. These results demonstrate that NaOH-GDL treatment is an effective method for improving emulsion properties and may provide insights for the development of novel, healthy emulsified meat products.
[0082] The various embodiments in this specification are described in a progressive manner, with each embodiment focusing on the differences from other embodiments. Reference can be made to the common and similar parts between the various embodiments. For the devices disclosed in the embodiments, since they correspond to the methods disclosed in the embodiments, the description is relatively simple, and the relevant parts can be referred to the method description.
[0083] The above description of the disclosed embodiments is intended to enable one skilled in the art to implement or use the present invention. Various modifications to these embodiments will be readily apparent to one skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention is not limited to the embodiments shown herein but is intended to conform to the widest scope consistent with the principles and novel features disclosed herein.
Claims
1. A processing method for improving the emulsification properties of chicken protein, characterized in that: The following steps are involved: (1) Remove the fat and fascia on the surface of chicken breast, and then extract myofibrillar protein; (2) diluting the myofibrillar protein extracted in step (1) with a phosphate buffer, adjusting the pH to 11.0 with 2 M NaOH, and allowing the solution to stand to achieve pH equilibrium, thereby obtaining an alkaline myofibrillar protein solution; (3) adjusting the pH value of the alkaline myofibrillar protein solution obtained in step (2) to 7.0 with gluconolactone, and allowing the solution to stand to achieve pH equilibrium, thereby obtaining a myofibrillar protein aggregate solution, wherein the amount of gluconolactone added is 0.40-0.45 wt % based on the alkaline myofibrillar protein solution; (4) The myofibrillar protein aggregate solution obtained in step (3) is centrifuged at high speed under low temperature to obtain the improved chicken protein.
2. A processing method for improving the emulsification properties of chicken protein according to claim 1, characterized in that: The steps for extracting myofibrillar protein in step (1) are as follows: The processed animal muscle is chopped into minced meat, added with standard salt solution, mixed evenly and homogenized. The filtered homogenate is centrifuged at low temperature and the precipitate is retained. The homogenization, filtration and low temperature centrifugation steps are repeated twice. The final precipitate is washed twice with a low salt solution to obtain myofibrillar protein.
3. A processing method for improving the emulsification properties of chicken protein according to claim 2, characterized in that: The standard salt solution formula components are: 0.1M KCl, 20mM K2HPO4 / KH2PO4, 1mM EGTA, 2mM MgCl2, pH=7.0; The low salt solution is 0.1 M KCl solution.
4. A processing method for improving the emulsification properties of chicken protein according to claim 2, characterized in that: The volume ratio of the standard salt solution to the minced meat is 2-8:1; the volume ratio of the added amount of the low-salt solution to the precipitate is 2-8:
1.
5. A processing method for improving the emulsification properties of chicken protein according to claim 2, characterized in that: The low salt solution washing steps are as follows: The precipitate and low-salt solution were mixed evenly and homogenized, and then centrifuged at low temperature. The homogenization and centrifugation steps were repeated once, and the final precipitate obtained was myofibrillar protein.
6. A processing method for improving the emulsification properties of chicken protein according to claim 2 or 5, characterized in that: The homogenization speed is 6700 rpm, the time is 30 s, and the operation is performed 1 to 2 times. The low-temperature centrifugation speed is 2000 to 2500×g, the time is 10 min, and the temperature is 4°C.
7. A processing method for improving the emulsification properties of chicken protein according to claim 1, characterized in that: The formula composition of the phosphate buffer in step (2) is: 0.6M KCl, 50mM K2HPO4 / KH2PO4, pH=7.0; the concentration of myofibrillar protein after dilution with the phosphate buffer is 12-18mg / ml.
8. A processing method for improving the emulsification properties of chicken protein according to claim 1, characterized in that: The low-temperature high-speed centrifugation of the myofibrillar protein aggregate solution in step (4) is performed at a speed of 8000 g, a time of 15 min, and a temperature of 4°C.