Raw fresh meat composite antibacterial fresh-keeping agent and low-temperature synergistic fresh-keeping process

Abstract

The invention discloses a raw fresh meat composite antibacterial fresh-keeping agent and a low-temperature synergistic fresh-keeping process, and relates to the technical field of food fresh-keeping. The preservative is formed by mixing a microcapsule component and an emulsion component, the microcapsule component is a microcapsule with epsilon-polylysine as a capsule core and a chitosan-nisin compound as a capsule wall, and the emulsion component comprises D-sodium erythorbate, lysozyme, tea polyphenol, glycerin monostearate and water. The fresh-keeping process comprises the following steps: immersing meat into a fresh-keeping agent, applying variable-frequency ultrasonic-assisted permeation, and storing at ice temperature. The fresh-keeping agent disclosed by the invention realizes full-period long-acting bacteriostasis of the fresh meat. The process disclosed by the invention realizes high-efficiency permeation of the preservative and low-loss storage of meat products, and is suitable for industrial preservation of fresh meat and industrial popularization of food.

Description

Technical Field

[0001] This invention relates to the field of food preservation technology. More specifically, this invention relates to a compound antibacterial preservative for fresh meat and a low-temperature synergistic preservation process. Background Technology

[0002] Fresh meat is rich in nutrients such as protein, free water, and fat. After slaughter, the meat's own endogenous enzymes continue to degrade it, while microorganisms in the environment easily contaminate the surface of the meat, leading to problems such as spoilage, browning, and flavor deterioration. Conventional low-temperature cold chain storage can slow down the rate of spoilage, but it cannot inhibit the growth of psychrophilic microorganisms or block the process of fat oxidation. The preservation time of low-temperature storage alone is relatively short, which is difficult to meet the actual needs of long-distance cold chain transportation and long-term sales. Therefore, safe and efficient fresh meat preservation technology has become a research focus in this field.

[0003] Currently, most fresh meat preservation methods focus on the application of natural food-grade active ingredients, primarily using single-system formulations or simple mixtures of multiple active ingredients to create conventional aqueous solutions. While these methods can exert certain antibacterial or antioxidant effects in a short period, they have significant shortcomings in large-scale applications that have remained unresolved for a long time. First, existing preservatives cannot meet the requirements for long-term preservation. After most existing preservatives are applied to the surface of meat, the core active ingredients will quickly diffuse, be lost, or be degraded, failing to fully inhibit the continuous proliferation of spoilage bacteria and meat oxidation during storage, causing fresh meat to begin to show signs of spoilage in the middle of storage.

[0004] Secondly, existing preservatives cannot simultaneously achieve uniform dispersion of components and long-lasting activity. One type suffers from poor dispersibility, failing to form a uniform film on the surface of meat, resulting in localized preservation deficiencies. Another type exhibits good dispersibility, but the active ingredients are released prematurely and become ineffective. Yet another type involves the use of multiple components in combination, but this can easily lead to problems such as system stratification, component aggregation, and antagonistic activity.

[0005] With the development of international trade in fresh meat, the demand for preservation with a shelf life of 2-4 weeks has emerged. While existing preservation technologies extend shelf life, they often struggle to maintain quality indicators such as meat color and juice loss rate, potentially increasing food safety risks. Therefore, there is an urgent need for an antibacterial preservative suitable for fresh meat to overcome these problems and achieve a dual improvement in both shelf life and meat quality. Summary of the Invention

[0006] This invention provides a compound antibacterial preservative for fresh meat, which achieves long-lasting antibacterial effect throughout the entire life cycle of fresh meat through the spatiotemporal synergistic effect of microcapsules and emulsion.

[0007] This invention provides a low-temperature synergistic preservation process, which achieves efficient penetration of preservatives and low-loss storage of meat products through ultrasonic-assisted impregnation and ice-temperature acclimatization, and is suitable for industrial promotion in the food industry.

[0008] To achieve these objectives and other advantages according to the present invention, a compound antibacterial preservative for fresh meat is provided, which is a mixture of microcapsule components and emulsion components; The microcapsule component is a microcapsule with ε-polylysine as the core and chitosan-nisin complex as the wall. The average particle size of the microcapsule is 10 μm to 50 μm, the mass ratio of the core to the wall is 1:2 to 1:5, and the mass ratio of chitosan to nisin in the chitosan-nisin complex is 10:1 to 20:1. The emulsion component is an emulsion containing the following components at mass-volume concentrations: sodium D-isoascorbate 0.5 g / L to 2.0 g / L, lysozyme 0.1 g / L to 0.8 g / L, tea polyphenols 0.3 g / L to 1.2 g / L, and glyceryl monostearate 0.5 g / L to 2.5 g / L, in sterile water as the solvent; The microcapsule component and the emulsion component are mixed in a mass ratio of 1:3 to 1:8. After mixing, the pH value is adjusted to 4.5 to 5.8 with food-grade citric acid to obtain the preservative.

[0009] Preferably, the microcapsules are prepared using an emulsification-chemical crosslinking method. The preparation process includes: dissolving chitosan and nisin in an acetic acid solution with a mass ratio of 10:1 to 20:1 in a volume fraction of 1% to 3% to obtain an aqueous phase; adding an emulsifier to the aqueous phase to achieve a mass-volume concentration of 1% to 5% to obtain a mixture; adding the mixture to the oil phase under stirring to form a water-in-oil emulsion; then adding an ε-polylysine aqueous solution to the water-in-oil emulsion, wherein the amount of ε-polylysine added is 20% to 50% of the total mass of chitosan and nisin, and the mass-volume concentration of the ε-polylysine aqueous solution is 10 g / L to 50 g / L; finally adding a crosslinking agent, wherein the amount of crosslinking agent added is 5% to 15% of the mass of chitosan; the reaction temperature is 30°C to 50°C; and the reaction time is 2 h to 6 h.

[0010] Preferably, the emulsifier consists of a primary emulsifier and a co-emulsifier, wherein the primary emulsifier is Span 80 and the co-emulsifier is Tween 80, and the mass ratio of the primary emulsifier to the co-emulsifier is 2:1.

[0011] Preferably, the crosslinking agent is at least one of glutaraldehyde, genipin, or sodium tripolyphosphate.

[0012] Preferably, when mixing the microcapsule component with the emulsion component, the microcapsule component is first ground to an average particle size of 5 μm to 15 μm, then pretreated with an ultrasonic disperser at a frequency of 20 kHz to 50 kHz for 5 min to 8 min, and then stirred and mixed at a speed of 200 rpm to 600 rpm under conditions of 4 °C to 6 °C. After mixing, the turbidity value of the system is stabilized at 50 NTU to 80 NTU.

[0013] The low-temperature synergistic preservation process using the aforementioned compound antibacterial preservative for fresh meat includes the following steps: The microcapsule component and the emulsion component were mixed at a mass ratio of 1:3 to 1:8 and dispersed by stirring at 200 rpm to 600 rpm for 5 to 15 minutes at 4°C to 10°C to obtain the composite preservation working solution. Fresh meat that has undergone post-slaughter inspection is immersed in a compound preservation working solution. The temperature of the working solution is controlled at 0℃ to 4℃, and the immersion time is maintained at 120s to 360s. During this period, alternating 20kHz low-frequency ultrasound and 60kHz high-frequency ultrasound are applied to the working solution. The ratio of the action time of low-frequency ultrasound to high-frequency ultrasound is 1:1 to 1:3. Remove the soaked fresh meat and place it in a clean environment at 0℃ to 4℃ to air dry until all free moisture on the surface of the fresh meat is removed. The air-dried fresh meat is placed in a sterile workbench and vacuum-sealed using a high-barrier food packaging film, ensuring that the oxygen volume content of the residual air inside the packaging is less than 0.5%. Vacuum-packed fresh meat is placed in a cold storage room and stored at a temperature within ±0.5℃ of its initial freezing point.

[0014] Preferably, the alternating low-frequency and high-frequency ultrasound is applied intermittently, with each 30-second ultrasound cycle consisting of 10 seconds of low-frequency ultrasound and 20 seconds of high-frequency ultrasound, followed by a 10-second pause. The ultrasound power density is 0.5 W / cm². 2 Up to 0.6W / cm 2 The distance between the ultrasound probe and the raw meat should be maintained between 5cm and 8cm.

[0015] Preferably, in the air-drying step, purified air with a wind speed of 1.0 m / s to 3.0 m / s is used for air drying for 3 to 8 minutes.

[0016] Preferably, before sending the fresh meat into the cold storage, the vacuum-packed fresh meat is first treated in an environment of -10°C to -5°C for 30 to 90 seconds.

[0017] Preferably, in the ice-temperature storage step, the vacuum-packed fresh meat is placed in an ice-temperature storage room for slow cooling. When the temperature drops to 0.5°C above the initial freezing point of the fresh meat, the cooling is paused and the meat is kept warm for 10 to 12 hours. Then, the temperature is lowered to the initial freezing point ±0.2°C at a rate of 0.1°C / h to 0.2°C / h.

[0018] The present invention has at least the following beneficial effects: First, this invention achieves rapid onset and long-lasting spatiotemporal synergistic antibacterial effect through a composite system of microcapsules and emulsions, significantly extending the shelf life of fresh meat.

[0019] Secondly, this invention achieves continuous and stable release of active ingredients by encapsulating ε-polylysine in the chitosan-nisin cross-linked capsule wall. The capsule wall and capsule core work together to destroy the outer membrane of Gram-negative bacteria, thus achieving broad-spectrum and long-lasting antibacterial effect.

[0020] Third, the present invention ensures uniform dispersion of microcapsules by forming a stable emulsion with glyceryl monostearate, and protects emulsifiers and active ingredients with sodium D-isoascorbate, so that a complete protective film is formed on the surface of meat products.

[0021] Fourth, this invention breaks down the secondary aggregation of microcapsules by grinding combined with ultrasonic pretreatment, and promotes the deep penetration of preservatives into meat products by intermittent variable frequency ultrasonic impregnation using cavitation effect, while avoiding muscle fiber damage caused by continuous ultrasound.

[0022] Fifth, this invention reduces water sublimation in the early stages of ice-temperature storage through rapid surface crusting treatment, and reduces intracellular ice crystal formation through pre-freezing insulation, thereby synergistically reducing juice loss and enhancing antibacterial effects.

[0023] Other advantages, objectives and features of the present invention will become apparent in part from the following description, and in part from those skilled in the art through study and practice of the invention. Detailed Implementation

[0024] The present invention will be further described in detail below with reference to examples, so that those skilled in the art can implement it based on the description.

[0025] It should be understood that terms such as "having," "comprising," and "including" as used herein do not exclude the presence or addition of one or more other elements or combinations thereof. It should be noted that the experimental methods described in the following embodiments, unless otherwise specified, are conventional methods, and the reagents and materials, unless otherwise specified, are commercially available and therefore should not be construed as limiting the invention.

[0026] <Example 1> Preservation techniques for fresh meat include; (1) Microcapsule preparation: Weigh 15.0 g of chitosan (degree of deacetylation 90%, viscosity-average molecular weight 10000 Da) and 1.0 g of nisin (potency 1000 IU / mg), with a mass ratio of 15:1. Dissolve them in 500 mL of 2% acetic acid solution and stir until completely dissolved to obtain the aqueous phase.

[0027] Weigh out 10.0g of Span 80 and 5.0g of Tween 80 (mass ratio 2:1), add them to the above aqueous phase, stir to dissolve, so that the mass volume concentration of the emulsifier in the aqueous phase is (15g / 0.5L) = 30g / L, that is, 3.0%.

[0028] While stirring at 500 rpm, the above aqueous phase was slowly added to a reaction vessel pre-filled with 1000 mL of liquid paraffin (oil phase), and stirring was continued for 30 min to form a stable water-in-oil emulsion.

[0029] Based on a core-to-wall mass ratio of 1:3, and a total wall mass of 16.0 g (chitosan + nisin), the core ε-polylysine should be 5.33 g. Weigh 5.33 g of ε-polylysine, dissolve it in 178 mL of sterile water to prepare a solution with a volume concentration of 30 g / L, and slowly add it to the above emulsion, continuing to stir for 30 min.

[0030] Finally, add 6.0 mL of glutaraldehyde (25% aqueous solution) (containing 1.5 g of pure glutaraldehyde, which is 10% of the mass of chitosan), and react in a water bath at 40°C for 4 h.

[0031] After the reaction was complete, the emulsion was centrifuged (4000 rpm, 10 min), the supernatant was discarded, and the microcapsules were collected. The microcapsules were washed three times with anhydrous ethanol and three times with distilled water, centrifuged after each wash. Finally, the microcapsules were dried in a vacuum drying oven at 40℃ for 12 h to obtain microcapsule powder. The average particle size was determined to be 30 μm using a laser particle size analyzer; the mass ratio of the capsule core to the capsule wall was approximately 1:3 by gravimetric analysis.

[0032] (2) Emulsion preparation: Take 1.5g of glyceryl monostearate, add it to 200mL of sterile water at 45℃, and stir at 300rpm until completely melted and dispersed to obtain an emulsifier dispersion. Let it cool to room temperature for later use.

[0033] Take another 800mL of sterile water, add 1.0g of sodium D-isoascorbate and 0.5g of lysozyme (activity 20000U / mg) in sequence, stir at 500rpm until completely dissolved; then add 0.8g of tea polyphenols (content 98%), and continue stirring for 10min until completely dispersed.

[0034] The above glyceryl monostearate dispersion was slowly poured into the aqueous phase and stirred at 600 rpm for 15 min to obtain a preliminary emulsion.

[0035] The initial emulsion was homogenized twice using a high-pressure homogenizer at 25 MPa for 3 minutes each time, resulting in a milky white, homogeneous emulsion. The final volume was approximately 1 L, and the concentrations of each component met the requirements.

[0036] (3) Compound preservatives: Take 20g of the above microcapsule powder and add it to 100mL of emulsion (mass ratio 1:5). Adjust the pH to 5.2 with food-grade citric acid and stir evenly to obtain a compound preservative.

[0037] (4) Preservation process: The longissimus dorsi muscle of pigs within 24 hours post-slaughter (initial total bacterial count 3.2 log CFU / g) was cut into pieces of approximately 200g each. The compound preservative (microcapsules and emulsion were mixed) was dispersed at 400 rpm for 10 minutes at 6°C to obtain the working solution.

[0038] The meat was completely immersed in the working solution (4°C) for 240 seconds. During this time, alternating 20kHz and 60kHz ultrasound was applied to the working solution, with a low-frequency to high-frequency duration ratio of 1:2 and an ultrasound power density of 0.5W / cm³. 2 The probe is about 6cm away from the surface of the meat.

[0039] Remove the meat, drain off excess liquid from the surface, and place it in a clean environment at 4°C with ventilation to help it air dry until there is no free moisture on the surface.

[0040] Use high-barrier food packaging film (oxygen permeability <10cm) 3 / (m 2 Vacuum skin packaging was performed at 0.1 MPa for 24 hours to control the residual oxygen volume content inside the packaging to be less than 0.5%.

[0041] The packaged meat was placed in a cold storage room and slowly cooled to -1.5℃ at a rate of 0.2℃ / h (the initial freezing point of this batch of pork was measured to be -1.8℃), and stored at this temperature for 28 days.

[0042] <Example 2> Preservation techniques for fresh meat include; (1) Microcapsule preparation: Weigh 10.0g of chitosan and 1.0g of nisin, with a mass ratio of 10:1, and dissolve them in 500mL of 2% acetic acid solution. Stir until completely dissolved to obtain an aqueous phase.

[0043] Weigh 10.0g of Span 80 and add it to the above aqueous phase. Stir to dissolve, so that the mass-volume concentration of the emulsifier in the aqueous phase is (10g / 0.5L) = 20g / L, or 2.0%.

[0044] While stirring at 500 rpm, the above aqueous phase was slowly added to a reaction vessel pre-filled with 1000 mL of liquid paraffin (oil phase), and stirring was continued for 30 min to form a stable water-in-oil emulsion.

[0045] Based on a core-to-wall mass ratio of 1:2, and a total wall mass of 11.0 g (chitosan + nisin), the core ε-polylysine should be 5.5 g. Weigh 5.5 g of ε-polylysine, dissolve it in 200 mL of sterile water to prepare a solution with a volume concentration of 27.5 g / L, and slowly add it to the above emulsion, continuing to stir for 30 min.

[0046] Finally, 0.8 g of genipin (8% of the chitosan mass) was added, and the mixture was reacted in a 40°C water bath for 4 hours. After the reaction was complete, the emulsion was centrifuged (4000 rpm, 10 min), the supernatant was discarded, and the microcapsules were collected. The microcapsules were washed three times with anhydrous ethanol and three times with distilled water, centrifuged after each wash.

[0047] Finally, the microcapsules were dried in a vacuum drying oven at 40℃ for 12 hours to obtain microcapsule powder. The average particle size was measured to be 20 μm by a laser particle size analyzer; the mass ratio of the capsule core to the capsule wall was determined to be approximately 1:2 by weighing.

[0048] (2) Emulsion preparation: Take 2.0 g of glyceryl monostearate and add it to 200 mL of sterile water at 45 °C. Stir at 300 rpm until completely melted and dispersed to obtain an emulsifier dispersion. Let it cool to room temperature for later use. Take another 800 mL of sterile water and add 0.5 g of sodium D-isoascorbate and 0.8 g of lysozyme in sequence. Stir at 500 rpm until completely dissolved. Then add 1.2 g of tea polyphenols and continue stirring for 10 min until completely dispersed. Slowly pour the above glyceryl monostearate dispersion into the aqueous phase and stir at 600 rpm for 15 min to obtain a preliminary emulsion. Homogenize the preliminary emulsion twice at 25 MPa pressure for 3 min each time to obtain a milky white uniform emulsion. The final volume is about 1 L, and the concentration of each component meets the requirements.

[0049] (3) Preservative compounding: Take 20g of the above microcapsule powder, add it to 100mL of emulsion, adjust the pH to 4.8 with food-grade citric acid, stir evenly, and obtain a compound preservative.

[0050] (4) Preservation process: Take the longissimus dorsi muscle of cattle within 24 hours after slaughter (initial freezing point -1.5℃) and cut it into pieces of about 200g each. Disperse the compound preservative at 400rpm for 10min at 6℃ to obtain the working solution. Immerse the meat completely in the working solution (temperature 4℃) for 300s. During this period, apply alternating 20kHz and 60kHz ultrasound to the working solution, with a low-frequency to high-frequency action time ratio of 1:1 and an ultrasound power density of 0.5W / cm³. 2 The probe should be approximately 6cm from the surface of the meat. Remove the meat, drain excess liquid, and place it in a clean, ventilated environment at 4℃ to air dry until no free moisture remains on the surface. Use a high-barrier food packaging film (oxygen permeability <10cm). 3 / (m 2 Vacuum-sealed packaging was performed at 0.1 MPa for 24 hours, controlling the residual oxygen volume content inside the packaging to be below 0.5%. The packaged meat was then placed in an ice-temperature storage room and slowly cooled to -1.2℃ at a rate of 0.25℃ / h, where it was stored for 28 days.

[0051] <Example 3> Preservation techniques for fresh meat include; (1) Microcapsule preparation: Weigh 20.0g of chitosan and 1.0g of nisin, with a mass ratio of 20:1, and dissolve them in 500mL of 2% acetic acid solution. Stir until completely dissolved to obtain an aqueous phase.

[0052] Weigh 20.0g of Tween 80 and add it to the above aqueous phase. Stir to dissolve, so that the mass-volume concentration of the emulsifier in the aqueous phase is (20g / 0.5L) = 40g / L, or 4.0%.

[0053] While stirring at 500 rpm, the above aqueous phase was slowly added to a reaction vessel pre-filled with 1000 mL of liquid paraffin (oil phase), and stirring was continued for 30 min to form a stable water-in-oil emulsion.

[0054] Based on a core-to-wall mass ratio of 1:5, and a total wall mass of 21.0g (chitosan + nisin), the core ε-polylysine should be 4.2g. Weigh 4.2g of ε-polylysine, dissolve it in 200mL of sterile water to prepare a solution with a mass-to-volume concentration of 21g / L, and slowly add it to the above emulsion, continuing to stir for 30min.

[0055] Finally, add 2.4g of sodium tripolyphosphate (12% of the chitosan mass) and react in a 40℃ water bath for 4 hours.

[0056] After the reaction was complete, the emulsion was centrifuged (4000 rpm, 10 min), the supernatant was discarded, and the microcapsules were collected. The microcapsules were washed three times with anhydrous ethanol and three times with distilled water, and centrifuged after each wash.

[0057] Finally, the microcapsules were dried in a vacuum drying oven at 40℃ for 12 hours to obtain microcapsule powder. The average particle size was measured to be 45 μm by a laser particle size analyzer; the mass ratio of the capsule core to the capsule wall was determined to be approximately 1:5 by weighing.

[0058] (2) Emulsion preparation: Take 0.8g of glyceryl monostearate and add it to 200mL of sterile water at 45℃. Stir at 300rpm until completely melted and dispersed to obtain an emulsifier dispersion. Let it cool to room temperature for later use. Take another 800mL of sterile water and add 2.0g of sodium D-isoascorbate and 0.2g of lysozyme in sequence. Stir at 500rpm until completely dissolved. Then add 0.4g of tea polyphenols and continue stirring for 10min until completely dispersed. Slowly pour the above glyceryl monostearate dispersion into the aqueous phase and stir at 600rpm for 15min to obtain a preliminary emulsion. Homogenize the preliminary emulsion twice at 25MPa pressure for 3min each time to obtain a milky white uniform emulsion. The final volume is about 1L, and the concentration of each component meets the requirements.

[0059] (3) Preservative compounding: Take 20g of the above microcapsule powder, add it to 100mL of emulsion, adjust the pH to 5.8 with food-grade citric acid, stir evenly, and obtain a compound preservative.

[0060] (4) Preservation process: Take the longissimus dorsi muscle of sheep within 24 hours after slaughter (initial freezing point -2.0℃) and cut it into pieces of about 200g. Disperse the compound preservative at 400rpm for 10min at 6℃ to obtain the working solution. Immerse the meat completely in the working solution (temperature 4℃) for 150s. During this period, apply alternating 20kHz and 60kHz ultrasound to the working solution, with a low-frequency to high-frequency action time ratio of 1:3 and an ultrasound power density of 0.5W / cm³. 2 The probe should be approximately 6cm from the surface of the meat. Remove the meat, drain excess liquid, and place it in a clean, ventilated environment at 4℃ to air dry until no free moisture remains on the surface. Use a high-barrier food packaging film (oxygen permeability <10cm). 3 / (m 2 Vacuum-sealed packaging was performed at 0.1 MPa for 24 hours, controlling the residual oxygen volume content inside the packaging to be below 0.5%. The packaged meat was then placed in an ice-temperature storage room and slowly cooled to -1.8℃ at a rate of 0.15℃ / h, where it was stored for 28 days.

[0061] <Example 4> Preservation techniques for fresh meat include; (1) Microcapsule preparation: Weigh 15.0g of chitosan (90% degree of deacetylation, viscosity-average molecular weight 10000Da) and 1.0g of nisin (potency 1000IU / mg), with a mass ratio of 15:1. Dissolve them in 500mL of 2% acetic acid solution and stir until completely dissolved to obtain an aqueous phase.

[0062] Weigh out 10.0g of Span 80 and 5.0g of Tween 80 (mass ratio 2:1), add them to the above aqueous phase, stir to dissolve, so that the mass volume concentration of the emulsifier in the aqueous phase is (15g / 0.5L) = 30g / L, that is, 3.0%.

[0063] While stirring at 500 rpm, the above aqueous phase was slowly added to a reaction vessel pre-filled with 1000 mL of liquid paraffin (oil phase), and stirring was continued for 30 min to form a stable water-in-oil emulsion.

[0064] Based on the core-to-wall mass ratio of 1:3, the total mass of the wall (chitosan + nisin) is 16.0g, therefore the core ε-polylysine should be 5.33g.

[0065] Weigh 5.33 g of ε-polylysine, dissolve it in 178 mL of sterile water to prepare a solution with a mass-volume concentration of 30 g / L, slowly add it to the above emulsion, and continue stirring for 30 min.

[0066] Finally, add 6.0 mL of glutaraldehyde (25% aqueous solution) (containing 1.5 g of pure glutaraldehyde, which is 10% of the mass of chitosan), and react in a water bath at 40°C for 4 h.

[0067] After the reaction was complete, the emulsion was centrifuged (4000 rpm, 10 min), the supernatant was discarded, and the microcapsules were collected. The microcapsules were washed three times with anhydrous ethanol and three times with distilled water, and centrifuged after each wash.

[0068] Finally, the microcapsules were dried in a vacuum drying oven at 40℃ for 12 hours to obtain microcapsule powder. The average particle size was measured to be 30 μm by a laser particle size analyzer; the mass ratio of the capsule core to the capsule wall was determined to be approximately 1:3 by weighing.

[0069] (2) Emulsion preparation: Take 1.5g of glyceryl monostearate and add it to 200mL of sterile water at 45℃. Stir at 300rpm until completely melted and dispersed to obtain an emulsifier dispersion. Let it cool to room temperature for later use. Take another 800mL of sterile water and add 1.0g of sodium D-isoascorbate and 0.5g of lysozyme (activity 20000U / mg) sequentially. Stir at 500rpm until completely dissolved. Then add 0.8g of tea polyphenols (content 98%) and continue stirring for 10min until completely dispersed. Slowly pour the above glyceryl monostearate dispersion into the aqueous phase and stir at 600rpm for 15min to obtain a preliminary emulsion. Homogenize the preliminary emulsion twice at 25MPa pressure for 3min each time to obtain a milky white uniform emulsion. The final volume is about 1L, and the concentration of each component meets the requirements.

[0070] (3) Preservative compounding: Take the microcapsules obtained in step (1) above and grind them for 30 minutes using a planetary ball mill to reduce the average particle size of the microcapsules to 10 μm. Weigh 20 g of the ground microcapsules and add them to 100 mL of sterile water. Use an ultrasonic disperser at a frequency of 30 kHz and a power density of 0.3 W / cm³. 2 Pre-treat for 6 min to obtain microcapsule pre-treatment solution; mix microcapsule pre-treatment solution with 100 mL of emulsion prepared in step (2) above, and stir at 400 rpm for 10 min at 6℃ to obtain composite preservative working solution.

[0071] (4) Preservation process: Take the longissimus dorsi muscle of pigs within 24 hours after slaughter (initial total bacterial count 3.2 log CFU / g, initial freezing point -1.8℃), and cut it into pieces of about 200g. Maintain the temperature of the compound preservative working solution at 4℃ to obtain the immersion working solution. Immerse the meat completely in the immersion working solution for 240s. During this period, apply intermittent ultrasound to the working solution: every 30s of ultrasound consists of 10s of low frequency (20kHz) + 20s of high frequency (60kHz), followed by a 10s pause, with an ultrasound power density of 0.55W / cm³. 2 The probe was held approximately 6cm from the meat surface (actual ultrasonic treatment time 160s, pause 80s). The meat was removed, excess liquid drained, and placed in a clean, ventilated environment at 4℃ for assisted air drying until no free moisture remained on the surface. A high-barrier food packaging film (oxygen permeability <10cm) was used. 3 / (m 2Vacuum-sealed packaging was performed at 0.1 MPa for 24 hours, controlling the residual oxygen volume content inside the packaging to be below 0.5%. The packaged meat was then placed in an environment of -8℃ for 60 seconds to rapidly form a continuous ice layer with a thickness of approximately 0.5-1 mm on its surface. The meat with the continuous ice layer was then transferred to an ice-temperature storage room and slowly cooled to -1.3℃ at a rate of 0.15℃ / h. Cooling was then paused and the temperature was maintained for 11 hours. The temperature was then further slowly cooled to the range of -1.8℃ to -1.6℃ at a rate of 0.15℃ / h, and stored at this temperature for 28 days.

[0072] <Experiment on the Preservation Effect of Fresh Meat> This experiment systematically verified the synergistic effect and safety of the microcapsule-emulsion composite preservative of the present invention through four indicators: total bacterial count, TVB-N value, sensory score, and residual amount of active ingredients.

[0073] 1.1 Experimental Grouping: Experimental Group - Example 1 (1) Preservative formulation: It is composed of microcapsules and emulsion in a mass ratio of 1:5 and the pH after compounding is 5.2. The raw materials of microcapsules are chitosan 15.0g, nisin 1.0g, and ε-polylysine 5.33g. The emulsion contains 1.0g of sodium D-isoascorbate, 0.5g of lysozyme, 0.8g of tea polyphenols, and 1.5g of glyceryl monostearate per liter.

[0074] (2) Preparation of preservative: Microcapsules were prepared by emulsification-chemical cross-linking method, emulsion was prepared according to the ratio, the two were mixed and the pH was adjusted to 5.2, and stirred evenly to obtain the product.

[0075] (3) Preservation target: longissimus dorsi muscle of pig.

[0076] (4) Preservation process: Immerse the meat in the above-mentioned compound preservative (4°C) for 240s, during which 20kHz and 60kHz ultrasound are applied alternately (action time ratio 1:2). The remaining air drying, packaging and ice-temperature storage conditions are the same as the basic control process.

[0077] Experimental Group - Example 2 (1) Preservative formulation: It is composed of microcapsules and emulsion in a mass ratio of 1:5 and the pH after compounding is 4.8. The raw materials of microcapsules are chitosan 10.0g, nisin 1.0g, and ε-polylysine 5.5g. The emulsion contains sodium D-isoascorbate 0.5g, lysozyme 0.8g, tea polyphenols 1.2g, and glyceryl monostearate 2.0g per liter.

[0078] (2) Preparation of preservative: Microcapsules were prepared by emulsification-chemical cross-linking method, emulsion was prepared according to the ratio, the two were mixed and the pH was adjusted to 4.8, and stirred evenly to obtain the product.

[0079] (3) Preservation target: longissimus dorsi muscle of cow.

[0080] (4) Preservation process: Immerse the meat in the above-mentioned compound preservative (4°C) for 300s, during which time alternating 20kHz and 60kHz ultrasound (action time ratio 1:1) are applied. The remaining air drying, packaging and ice-temperature storage conditions are the same as in Example 1.

[0081] Experimental Group - Example 3 (1) Preservative formulation: It is composed of microcapsules and emulsion in a mass ratio of 1:5 and the pH after compounding is 5.8. The raw materials of microcapsules are chitosan 20.0g, nisin 1.0g and ε-polylysine 4.2g. The emulsion contains sodium D-isoascorbate 2.0g, lysozyme 0.2g, tea polyphenols 0.4g and glyceryl monostearate 0.8g per liter.

[0082] (2) Preparation of preservative: Microcapsules were prepared by emulsification-chemical cross-linking method, emulsion was prepared according to the ratio, the two were mixed and the pH was adjusted to 5.8, and stirred evenly to obtain the product.

[0083] (3) Preservation target: longissimus dorsi muscle of sheep.

[0084] (4) Preservation process: Immerse the meat in the above-mentioned compound preservative (4°C) for 150s, during which 20kHz and 60kHz ultrasound are applied alternately (action time ratio 1:3). The remaining air drying, packaging and ice-temperature storage conditions are the same as in Example 1.

[0085] Experimental Group - Comparative Example 1 (1) Preservative formulation: Only the emulsion of Example 1 is used, without adding microcapsules. Emulsion composition: 1.0g of sodium D-isoascorbate, 0.5g of lysozyme, 0.8g of tea polyphenols, and 1.5g of glyceryl monostearate per liter.

[0086] (2) Preparation of preservative: 1L of emulsion was prepared according to the emulsion preparation method in Example 1.

[0087] (3) Preservation target: longissimus dorsi muscle of pig.

[0088] (4) Preservation process: Immerse the meat in the above emulsion (4°C) for 240s, during which 20kHz and 60kHz ultrasound are applied alternately (time ratio 1:2). The remaining air drying, packaging and ice-temperature storage conditions are the same as in Example 1.

[0089] Experimental Group - Comparative Example 2 (1) Preservative formulation: Only the microcapsules of Example 1 are used, without adding emulsion. The microcapsules are prepared in the same way as in Example 1.

[0090] (2) Preparation of preservative: Take 20g of microcapsules and disperse them in 100mL of sterile water to make a suspension.

[0091] (3) Preservation target: longissimus dorsi muscle of pig.

[0092] (4) Preservation process: Immerse the meat in the above microcapsule suspension (4°C) for 240s, during which alternating 20kHz and 60kHz ultrasound (action time ratio 1:2) are applied, and the rest is the same as in Example 1.

[0093] Experimental Group - Comparative Example 3 (1) Preservative formula: sterile water.

[0094] (2) Preservation target: longissimus dorsi muscle of pig.

[0095] (3) Preservation process: Immerse the meat in sterile water (4°C) for 240s, during which time alternating 20kHz and 60kHz ultrasound (action time ratio 1:2) are applied, and the rest is the same as in Example 1.

[0096] Experimental Group - Comparative Example 4 (1) Preservative formula: Each liter of aqueous solution contains 2.0g of chitosan, 1.0g of tea polyphenols and 0.5g of lysozyme.

[0097] (2) Preparation of preservative: Weigh 2.0g of chitosan, 1.0g of tea polyphenols and 0.5g of lysozyme, and dissolve them in 1L of sterile water and stir to dissolve.

[0098] (3) Preservation target: longissimus dorsi muscle of pig.

[0099] (4) Preservation process: Immerse the meat in the solution (4°C) for 240s, during which alternating 20kHz and 60kHz ultrasound (action time ratio 1:2) are applied, and the rest is the same as in Example 1.

[0100] 1.2 Test Content: (1) Total bacterial count determination: Performed according to GB4789.2-2022. Take 25g of meat sample, cut it into small pieces, add 225mL of sterile physiological saline, homogenize for 2min, and prepare a 1:10 dilution. Dilute 10-fold serially, select 2-3 suitable dilutions, take 1mL of each and pour it onto a plate counting agar, incubate at 36℃±1℃ for 48h±2h, count and calculate the total bacterial count, and the result is expressed as log CFU / g.

[0101] (2) Determination of volatile basic nitrogen (TVB-N): The semi-micro nitrogen determination method was performed according to GB5009.228-2016. 10 g of minced meat sample was weighed, 75 mL of water was added, and the mixture was shaken for 30 min and then filtered. 5.0 mL of the filtrate was distilled, absorbed with boric acid, and titrated with hydrochloric acid standard solution. The TVB-N content was calculated, and the result was expressed as mg / 100 g.

[0102] (3) Sensory evaluation: A panel of 10 trained sensory evaluators will comprehensively evaluate the color, odor, and texture of the meat samples using a 10-point scale. Evaluation criteria: 9-10 points: bright red (or natural meat color), with the inherent odor of fresh meat, and a firm, elastic texture; 7-8 points: slightly darker color, normal odor, and a relatively firm texture; 5-6 points: dark red color, slight off-odor, and a soft texture; 3-4 points: dark brown color, a noticeable off-odor, and a soft, inelastic texture; 1-2 points: browned color, a putrid odor, and increased mucus in the tissue. The average score for each item will be used as the sensory evaluation score.

[0103] (4) Determination of residual active ingredients: ε-polylysine was determined by high performance liquid chromatography according to GB5009.243-2016; nisin was determined by tube-disc method according to GB1886.231-2016.

[0104] 1.3 Experimental Results: The effects of different treatments on the total bacterial count (log CFU / g) are shown in Table 1. The effects of different treatments on the TVB-N value (mg / 100g) are shown in Table 2. The effects of different treatments on the sensory score (out of 10) are shown in Table 3. The results of the residual amount of active ingredients after 28 days of storage in Example 1 are shown in Table 4.

[0105] Table 1 Note: Comparative Example 3 had a total bacterial count exceeding 8.5 log CFU / g at 21 days, indicating severe meat spoilage, so the assay was terminated, and data from 28 days were no longer tested.

[0106] As shown in Table 1, the total bacterial counts of Examples 1-3 at 28 days were 5.9, 5.8, and 6.1 log CFU / g, respectively, all near the critical spoilage value (6 log CFU / g), and the differences among the three were within the experimental error range. Comparative Example 1 had a count of 7.8, Comparative Example 2 7.3, and Comparative Example 4 8.3, while Comparative Example 3 reached 8.9 at 21 days. Comparative Example 1 contained only an emulsion; lysozyme and tea polyphenols effectively inhibited microorganisms from 0 to 7 days, but after 7 days, the small molecule active ingredients rapidly degraded and became inactive, and without protection in the later stages, microorganisms entered the logarithmic phase and proliferated rapidly. Comparative Example 2 contained only microcapsules; the initial release of ε-polylysine was insufficient (4.2 colonies at 7 days), and some microorganisms adapted early and entered the logarithmic phase prematurely. Comparative Example 4 was a chitosan-tea polyphenol-lysozyme aqueous solution, with a single component and no sustained release; the active ingredients were depleted within 7-14 days. Comparative Example 3 (blank group) showed no antibacterial activity. Examples 1-3 demonstrate that by providing high concentrations of antibacterial agents through emulsions from 0 to 7 days and continuously releasing ε-polylysine through microcapsules from 7 to 28 days, microorganisms are unable to obtain the logarithmic growth window.

[0107] Table 2 Note: Comparative Example 3 was severely decomposed at 21 days, so subsequent measurements were terminated, and data from 28 days were no longer tested.

[0108] As shown in Table 2, the 28-day TVB-N value of Example 1 was 16.7 mg / 100g, slightly exceeding the first-grade freshness limit (15 mg / 100g), while Comparative Example 1 was 23.4, Comparative Example 2 was 21.1, Comparative Example 4 was 25.0, and Comparative Example 3 reached 28.3 by day 21. In Comparative Example 1, microorganisms proliferated significantly in the later stages. When the bacterial count of Pseudomonas and other putrefactive bacteria exceeded 6 log CFU / g, they secreted large amounts of extracellular proteases, accelerating protein decomposition. Simultaneously, the lack of ε-polylysine to inhibit proteases led to the accumulation of decomposition products. In Comparative Example 2, microbial control was insufficient in the early stages (4.1 colonies at 7 days), and some proteins were already enzymatically degraded initially. Although the microcapsules released ε-polylysine later, the losses from the early stages could not be compensated. Comparative Example 4 lacked the enzyme-inhibiting function of ε-polylysine and also lacked sodium D-isoascorbate, making proteins more easily hydrolyzed after oxidation. Comparative Example 3 had the highest total number of microorganisms and the most active metabolism, resulting in the extensive decomposition of proteins. The TVB-N growth in Example 1 was the slowest, partly because the total number of microorganisms was kept at a low level, reducing protease secretion; and partly because the antioxidant effect of sodium D-isoascorbate reduced protein oxidative modification. The cationic polypeptide structure of ε-polylysine has the property of adsorbing onto the protein surface, and it is known in the art that it may interfere with the binding of protease to substrate through steric hindrance.

[0109] Table 3 Note: Comparative Example 3 was severely decomposed at 21 days, so subsequent measurements were terminated, and data from 28 days were no longer tested.

[0110] As shown in Table 3, the sensory score of Example 1 at 28 days was 7.0, still within an acceptable level (>6 points), while Comparative Example 1 scored 3.9, Comparative Example 2 scored 4.4, Comparative Example 4 scored 3.4, and Comparative Example 3 had already dropped to 4.1 by day 14. In Comparative Example 1, microorganisms proliferated rapidly in the later stages, producing odorous substances such as hydrogen sulfide and ammonia; myoglobin was oxidized to methemoglobin, resulting in a dark brown color; and protein decomposition made the tissue soft. In Comparative Example 2, microbial control was insufficient in the early stages, with some areas showing slight spoilage, resulting in impaired color and odor. Comparative Example 4 lacked sodium D-isoascorbate, resulting in a lack of antioxidants; its color began to darken after 7 days; it also lacked long-lasting antibacterial effects, and a noticeable odor appeared by day 21. Comparative Example 3 was completely unprotected, and all its qualities deteriorated rapidly. In Example 1, sodium D-isoascorbate continuously reduces metmyoglobin, ε-polylysine inhibits hydrogen sulfide-producing bacteria, and the protective film formed by the microcapsules on the surface of the meat can physically block water migration, which may be one of the reasons for the reduced juice loss rate. The three work together to maintain color, aroma and texture.

[0111] Table 4 As shown in Table 4, after 28 days of storage, the residual amount of ε-polylysine in Example 1 was 12.5 mg / kg, and the residual amount of nisin was 0.18 mg / kg. According to GB 2760-2024, the maximum usage of nisin in meat products is 0.25 g / kg, and ε-polylysine can be used in appropriate amounts as needed for production. The residual amount of the active ingredients in this invention is far below the national standard limit, meeting food safety requirements. It should be noted that the residual amount of ε-polylysine was determined using GB5009.243-2016, and the result represents the total ε-polylysine content in the sample; some ε-polylysine remains encapsulated in microcapsules and does not directly contact the meat product. The actual free ε-polylysine that can migrate into the meat product is lower than this measured value. Therefore, the preservative of this invention has higher safety.

[0112] <Experiment on the Influence of Microcapsule Structure on Release Performance and Antibacterial Activity> This experiment verified the controlled-release advantage and broad-spectrum antibacterial activity of the microcapsule with a specific structure (ε-polylysine core + chitosan-nisin capsule wall) using two indicators: cumulative release rate of ε-polylysine and diameter of inhibition zone.

[0113] 2.1 Experimental Grouping: Experimental Group - Example 1 (1) Preservative formulation: It is composed of microcapsules and emulsion in a mass ratio of 1:5 and the pH after compounding is 5.2. The raw materials of microcapsules are chitosan 15.0g, nisin 1.0g, and ε-polylysine 5.33g. The emulsion contains 1.0g of sodium D-isoascorbate, 0.5g of lysozyme, 0.8g of tea polyphenols, and 1.5g of glyceryl monostearate per liter.

[0114] (2) Preparation of preservative: Microcapsules were prepared by emulsification-chemical cross-linking method, emulsion was prepared according to the ratio, the two were mixed and the pH was adjusted to 5.2, and stirred evenly to obtain the product.

[0115] (3) Preservation target: longissimus dorsi muscle of pig.

[0116] (4) Preservation process: Immerse the meat in the above-mentioned compound preservative (4°C) for 240s, during which 20kHz and 60kHz ultrasound are applied alternately (action time ratio 1:2). The remaining air drying, packaging and ice-temperature storage conditions are the same as the basic control process.

[0117] Experimental Group - Comparative Example 5 (1) Microcapsule preparation: 16.0 g of chitosan (replacing 15 g of original chitosan + 1 g of nisin) was dissolved in 500 mL of 2% acetic acid solution, 10.0 g of Span 80 and 5.0 g of Tween 80 were added, and after emulsification, ε-polylysine aqueous solution (same as in Example 1) was added, and 6.0 mL of glutaraldehyde was added. The reaction was carried out, washed, and dried to obtain microcapsules.

[0118] (2) Emulsion: Same as in Example 1.

[0119] (3) Preservative compounding: Take 20 g of the above microcapsules and mix with 100 mL of the emulsion in Example 1, and adjust the pH to 5.2. (4) Preservation target: longissimus dorsi muscle of pig.

[0120] (5) Preservation process: Same as in Example 1.

[0121] Experimental Group - Comparative Example 6 (1) Microcapsule preparation: 15.0 g of chitosan, 1.0 g of nisin and 5.33 g of ε-polylysine were mixed and dissolved in 500 mL of 2% acetic acid solution. The mixture was then spray-dried (inlet temperature 160℃, outlet temperature 80℃) to obtain microspheres.

[0122] (2) Emulsion: Same as in Example 1.

[0123] (3) Preservative compounding: Take 20 g of the above microspheres and mix them with 100 mL of the emulsion from Example 1, and adjust the pH to 5.2.

[0124] (4) Preservation target: longissimus dorsi muscle of pig.

[0125] (5) Preservation process: Same as in Example 1.

[0126] Experimental Group - Comparative Example 7 (1) Preservative powder: 15.0 g of chitosan, 1.0 g of nisin and 5.33 g of ε-polylysine are directly mixed and ground into fine powder.

[0127] (2) Emulsion: Same as in Example 1.

[0128] (3) Preservative compounding: Take 20 g of the above mixed powder, add it to 100 mL of sterile water, and stir evenly to obtain a suspension; at the same time, prepare 100 mL of the emulsion of Example 1. Mix the suspension and the emulsion and adjust the pH to 5.2.

[0129] (4) Preservation target: longissimus dorsi muscle of pig.

[0130] (5) Preservation process: Same as in Example 1.

[0131] 2.2 Test Content: Cumulative release rate determination of ε-polylysine: Weigh 50 mg of microcapsule sample containing ε-polylysine and place it in a dialysis bag (molecular weight cutoff 8000-14000 Da). Add 10 mL of pH 5.5 phosphate buffer, seal, and place in 100 mL of release medium (same buffer). Incubate at 4°C. At predetermined time points, take 1.0 mL of release medium (simultaneously replenish with an equal volume of fresh medium), determine the ε-polylysine concentration using the phthalaldehyde method, and calculate the cumulative release rate.

[0132] Inhibition zone diameter determination: Oxford cup method was used. Indicator bacteria (Pseudomonas aeruginosa CGMCC 1.3342, Listeria monocytogenes CGMCC 1.10790, Escherichia coli CGMCC 1.3373) were activated and then a bacterial suspension (concentration approximately 10) was prepared. 7 Take 0.1 mL of the sample solution (CFU / mL) and spread it on a nutrient agar plate. Place an Oxford cup evenly on the plate and add 0.2 mL of the sample solution to be tested (the microcapsule sample needs to be ground and dispersed in sterile water at a concentration of 10 mg / mL). After diffusion at 4°C for 2 h, incubate at 36°C for 24 h and measure the diameter of the inhibition zone (mm). Repeat the test 3 times for each sample and take the average value.

[0133] 2.3 Experimental Results: The effects of different microcapsule structures on the cumulative release rate of ε-polylysine (%, 4℃, pH 5.5) are shown in Table 5, and the effects of different microcapsule structures on the diameter of the inhibition zone (mm) are shown in Table 6.

[0134] Table 5 As shown in Table 5, Example 1 exhibited a release rate of 74% at 28 days, showing a gradual and gradual release; Comparative Example 5 reached 97%, Comparative Example 6 reached 98%, and Comparative Example 7 reached 83% at 3 days. Comparative Example 5, lacking nisin in its capsule wall, showed a significantly faster release rate, indicating that the incorporation of nisin may have increased the degree of cross-linking in the capsule wall and delayed the diffusion of ε-polylysine. This is consistent with the property that the amino and thiol groups in the nisin molecule can participate in cross-linking reactions. Comparative Example 6, using spray drying, resulted in disordered solidification of chitosan molecules at high temperatures, leading to a loose network and partial thermal denaturation and inactivation of nisin, resulting in a faster initial release (29% at 1 day). Comparative Example 7, being a physical mixture without a capsule wall barrier, directly exposed ε-polylysine to the medium, exhibiting explosive release characteristics, releasing 46% at 1 day and exhausting it at 3 days. In Example 1, because nisin is embedded in the capsule wall, the cross-linking density is increased, and the emulsification-chemical cross-linking forms a gradient structure. The outer layer is dense to control the initial release, while the inner layer is loose to maintain the later release, thus achieving continuous and stable sustained release.

[0135] Table 6 As shown in Table 6, after 14 days, Example 1 exhibited an inhibition zone of 10.1 mm against Pseudomonas and 9.4 mm against Escherichia coli, while Comparative Example 5 showed inhibition zones of only 8.4 and 6.1 mm, Comparative Example 6 only 7.7 and 5.7 mm, and Comparative Example 7 only 5.1 and 4.4 mm. Comparative Example 5 lacked nisin in its capsule wall and relied solely on ε-polylysine for inhibition against G. - The ability to destroy the outer membrane is limited, and there is a lack of synergistic effect from nisin; the antibacterial activity significantly decreases after 14 days. Comparative Example 6: Spray drying caused partial thermal denaturation of nisin; although the release rate was similar to Comparative Example 5, the active ingredient itself was damaged, resulting in a worse antibacterial effect. Comparative Example 7: The active ingredient was directly exposed, rapidly degraded and inactivated within 14 days; although the inhibition zone was large at 0 days, it was almost completely lost later. Example 1: The capsule wall contains nisin (mainly anti-G...). + (bacteria), the core of which is ε-polylysine (destroying G) - (outer membrane of bacteria), the two work synergistically to enhance the effect on G. - It inhibits bacterial growth, and the microcapsule protection allows the active ingredients to retain more than 80% of their activity after 14 days.

[0136] <Experimental Study on the Compatibility of Emulsion Components with Microcapsules> This experiment verified the effects of emulsion components (glyceryl monostearate and sodium D-isoascorbate) on the dispersion stability and antibacterial effect of microcapsules using two indicators: turbidity change and total bacterial count after 14 days.

[0137] 3.1 Experimental Grouping: Experimental Group - Example 1 (1) Preservative formulation: It is composed of microcapsules and emulsion in a mass ratio of 1:5 and the pH after compounding is 5.2. The raw materials of microcapsules are chitosan 15.0g, nisin 1.0g, and ε-polylysine 5.33g. The emulsion contains 1.0g of sodium D-isoascorbate, 0.5g of lysozyme, 0.8g of tea polyphenols, and 1.5g of glyceryl monostearate per liter.

[0138] (2) Preparation of preservative: Microcapsules were prepared by emulsification-chemical cross-linking method, emulsion was prepared according to the ratio, the two were mixed and the pH was adjusted to 5.2, and stirred evenly to obtain the product.

[0139] (3) Preservation target: longissimus dorsi muscle of pig.

[0140] (4) Preservation process: Immerse the meat in the above-mentioned compound preservative (4°C) for 240s, during which 20kHz and 60kHz ultrasound are applied alternately (action time ratio 1:2). The remaining air drying, packaging and ice-temperature storage conditions are the same as the basic control process.

[0141] Experimental Group - Comparative Example 8 (1) Emulsion preparation: Each liter contains 0.5g of lysozyme, 0.8g of tea polyphenols, and 1.5g of glyceryl monostearate, without adding sodium D-isoascorbate, and the rest is the same as in Example 1.

[0142] (2) Microcapsules: Same as in Example 1.

[0143] (3) Preservative compounding: Mix 20g of microcapsules with 100mL of the above emulsion and adjust the pH to 5.2.

[0144] (4) Preservation target: longissimus dorsi muscle of pig.

[0145] (5) Preservation process: Same as in Example 1.

[0146] Experimental Group - Comparative Example 9 (1) Emulsion preparation: Each liter contains 1.0g of sodium D-isoascorbate, 0.5g of lysozyme, and 0.8g of tea polyphenols. Glyceryl monostearate is not added. It is directly dissolved in water to obtain a transparent solution.

[0147] (2) Microcapsules: Same as in Example 1.

[0148] (3) Preservative compounding: Add 20g of microcapsules to 100mL of the above aqueous solution and stir to disperse. During impregnation, the working solution needs to be continuously stirred to prevent the microcapsules from settling.

[0149] (4) Preservation target: longissimus dorsi muscle of pig.

[0150] (5) Preservation process: Same as in Example 1 (working solution is continuously stirred during the immersion process).

[0151] Experimental Group - Comparative Example 10 (1) Emulsion preparation: Each liter contains 0.5g of lysozyme and 0.8g of tea polyphenols, without adding sodium D-isoascorbate and glyceryl monostearate, and is directly dissolved in water.

[0152] (2) Microcapsules: Same as in Example 1.

[0153] (3) Preservative compounding: Take 20g of microcapsules and add them to 100mL of the above solution. Stir to disperse and continue stirring during impregnation.

[0154] (4) Preservation target: longissimus dorsi muscle of pig.

[0155] (5) Preservation process: Same as in Example 1 (continuous stirring during the soaking process).

[0156] 3.2 Test Content: (1) Turbidity determination: The liquid sample was directly measured using a Hach 2100Q turbidity meter. The result is expressed in NTU. Shake well before measurement.

[0157] (2) The total number of colonies was determined as described in section 1.2 above.

[0158] 3.3 Test Results: The effects of different emulsion components on physical stability (after standing at 4℃) are shown in Table 7. The dispersion stability of different emulsions mixed with microcapsules is shown in Table 8. The effects of different emulsions mixed with microcapsules on antibacterial effect (total bacterial count at 14 days, log CFU / g) are shown in Table 9.

[0159] Table 7 As shown in Table 7, the turbidity of Example 1 decreased only from 86 NTU to 84 NTU within 72 hours, a decrease of only 2.3%, and remained a milky white and homogeneous solution throughout. The turbidity of Comparative Example 8 decreased from 83 NTU to 77 NTU, a decrease of 7.2%, and slight stratification and bottom precipitation occurred after 72 hours. The turbidity of Comparative Example 9 increased from 13 NTU to 19 NTU, and that of Comparative Example 10 increased from 11 NTU to 17 NTU; both were light yellow and transparent solutions, and showed no stratification.

[0160] The reason for the decrease in turbidity in Comparative Example 8, which lacks D-isoascorbic acid sodium, is that the glyceryl monostearate molecule contains unsaturated bonds, which, in trace amounts of Fe... 2+ Cu 2+ Under the catalysis of metal ions, dissolved oxygen initiates a free radical chain reaction, leading to the oxidative breakage of emulsifier molecules. As the emulsifier gradually degrades, defects appear in the oil-water interface film, oil droplets begin to coalesce, their size increases, and the system's ability to scatter light decreases, manifesting as a reduction in turbidity. The white precipitate appearing at the bottom after 72 hours is either the coalesced large oil droplets or emulsifier degradation products.

[0161] Comparative Examples 9 and 10 lacked glyceryl monostearate, thus failing to form water-in-oil or oil-in-water emulsions. Water-soluble components such as sodium D-isoascorbate, lysozyme, and tea polyphenols completely dissolved in water, forming true solutions. The turbidity of these true solutions is typically below 20 NTU, mainly due to microbubbles or trace amounts of insoluble impurities. A slight increase in turbidity within 72 hours may be due to microbial growth producing tiny particles or the slow oxidation and polymerization of tea polyphenols to form soluble aggregates.

[0162] Example 1 contains both glyceryl monostearate and sodium D-isoascorbate, exhibiting stable turbidity: Glyceryl monostearate forms a hydrated dispersion in water at 45°C. Upon cooling, it oriented at the oil-water interface, with the lipophilic end extending into the oil phase and the hydrophilic end into the water phase, forming a dense interfacial film that significantly reduces interfacial tension and stabilizes oil droplet dispersion. Sodium D-isoascorbate, through the strong reducing properties of its enediol structure, scavenge free radicals catalyzed by metal ions in the system, blocking the oxidation chain reaction of the emulsifier, protecting the integrity of the interfacial film, and maintaining the long-term stability of the emulsion.

[0163] Table 8 As can be seen from Table 8, the turbidity of Example 1 decreased by only 2.4% after 2 hours of mixing and there was no precipitation, while the turbidity of Comparative Example 8 decreased by 17.9% and precipitation was visible at the bottom. This is because the emulsion of Comparative Example 8 had already begun to destabilize. When the microcapsules were added, the unstable oil droplets accelerated their aggregation, which led to a decrease in the viscosity of the system. This resulted in a significant increase in the sedimentation rate of the microcapsules, and thus the turbidity decreased significantly within 2 hours and precipitation occurred.

[0164] As shown in Table 8, the turbidity of Example 1 decreased by only 2.4% after 2 hours of mixing, with no precipitation. In contrast, the turbidity of Comparative Example 9 was only 45 NTU after mixing, and decreased by 37.0% to 28 NTU after 2 hours, with a large amount of precipitate at the bottom. This is because the continuous phase of Comparative Example 9 was a pure aqueous solution with a viscosity close to 1 cP, resulting in extremely weak suspending ability for the microcapsules. Based on Stokes' law, the particle size was estimated to be 30 μm, and the density difference was 0.15 g / cm³. 3 The microcapsules settle at a rate of about 0.15 mm / s in pure water and can settle to about 1 m within 2 hours, so the vast majority of the microcapsules settle to the bottom.

[0165] As shown in Table 8, the turbidity of Example 1 decreased by only 2.4% after 2 hours of mixing and there was no precipitation. In contrast, the turbidity of Comparative Example 10 was only 42 NTU after mixing and decreased by 39.5% to 25 NTU after 2 hours, with a large amount of precipitation at the bottom. This is because the continuous phase of Comparative Example 10 was a pure aqueous solution, and the lack of emulsifier resulted in the system having no viscosity support. At the same time, the absence of sodium D-isoascorbate was not the dominant factor in this system. The sedimentation of the microcapsules was entirely determined by the extremely low viscosity of the continuous phase, and its sedimentation kinetics were basically the same as those of Comparative Example 9.

[0166] Table 9 As shown in Table 9, the total bacterial count of Example 1 after 14 days was 4.4 log CFU / g, while that of Comparative Example 8 was 5.0 log CFU / g. This is because in Comparative Example 8, some microcapsules settled, resulting in a reduced amount of microcapsules adhering to the surface of the meat and uneven distribution, with insufficient protection in some areas; at the same time, although the emulsion contained lysozyme and tea polyphenols, it lacked the antioxidant protection of sodium D-isoascorbate, leading to a decrease in the stability of the active ingredients. The combination of these two factors weakened the antibacterial effect.

[0167] As shown in Table 9, the total bacterial count of Example 1 after 14 days was 4.4 log CFU / g, while that of Comparative Example 9 was 6.1 log CFU / g, which is close to the spoilage threshold. This is because in Comparative Example 9, a large number of microcapsules settled, and the actual amount of microcapsules in contact with the meat surface was extremely small, resulting in the near loss of the long-lasting antibacterial effect of ε-polylysine. The limited antibacterial effect was provided only by the aqueous solution of lysozyme and tea polyphenols in the emulsion, and because there was no emulsifier system, these components were unevenly distributed on the meat surface, leading to a severe decrease in the antibacterial effect.

[0168] As shown in Table 9, the total bacterial count of Example 1 after 14 days was 4.4 log CFU / g, while that of Comparative Example 10 was 6.2 log CFU / g, which is close to the spoilage threshold. This is because the microcapsules in Comparative Example 10 also precipitated in large quantities, and the emulsion lacked all the key components (sodium D-isoascorbate and glyceryl monostearate), leaving only an aqueous solution of lysozyme and tea polyphenols. Without emulsifiers to maintain dispersion or antioxidants to protect the active ingredients, its antibacterial ability was weaker than that of Comparative Example 9, hence the slightly higher total bacterial count.

[0169] In actual production, it is recommended to prepare and use the preservative immediately to ensure optimal dispersion. For short-term storage, it is advised to store it in a sealed container at 4°C and use it within 24 hours.

[0170] <Microencapsulation-Emulsion Mixing Process Optimization Experiment> 4.1 Experimental Grouping: Experimental Group - Example 4 (1) Preservative formulation: It is composed of microcapsules and emulsion. The raw material ratio, components and compound pH are the same as in Example 1.

[0171] (2) Preparation of preservative: Microcapsules and emulsion were prepared according to the method in Example 1; the microcapsules were ground and pretreated by ultrasonication, and then mixed with the emulsion to obtain the preservative working solution.

[0172] (3) Preservation target: longissimus dorsi muscle of pig.

[0173] (4) Preservation process: Immerse the meat in the above-mentioned preservation working solution (4°C) for 240s, during which intermittent alternating ultrasound is applied. The remaining air drying, packaging, ice-temperature pretreatment and storage conditions are the same as in Example 1.

[0174] Experimental Group - Comparative Example 11 (1) Microcapsules: Same as in Example 1.

[0175] (2) Emulsion: Same as in Example 1.

[0176] (3) Pretreatment: Take 20g of microcapsules, mix them directly with 100mL of emulsion without grinding or ultrasonic pretreatment, and stir at 400rpm for 10min at 6℃.

[0177] (4) Preservation target: longissimus dorsi muscle of pig.

[0178] (5) Preservation process: Same as in Example 4 (including intermittent ultrasound, surface crusting, and pre-freezing insulation).

[0179] Experimental Group - Comparative Example 12 (1) Microcapsules: Same as in Example 1, grinding + ultrasonic pretreatment is the same as in Example 4.

[0180] (2) Emulsion: Same as in Example 1.

[0181] (3) Preservative compounding: The pretreated microcapsules are mixed with the emulsion, as in Example 4.

[0182] (4) Preservation target: longissimus dorsi muscle of pig.

[0183] (5) Preservation process: No ultrasound is applied during soaking. The meat is simply soaked in a 4°C working solution for 240 seconds. The remaining steps (air drying, packaging, and ice-temperature storage) are the same as in Example 4.

[0184] Experimental Group - Comparative Example 13 (1) Microcapsules: Same as in Example 1, grinding + ultrasonic pretreatment is the same as in Example 4.

[0185] (2) Emulsion: Same as in Example 1.

[0186] (3) Preservative compound: Same as Example 4.

[0187] (4) Preservation target: longissimus dorsi muscle of pig.

[0188] (5) Preservation process: During impregnation, continuous 20kHz ultrasound (without interruption) is used with a power density of 0.55W / cm³. 2 The probe distance was 6cm, the immersion time was 240s, and the rest was the same as in Example 4.

[0189] Experimental Group - Comparative Example 14 (1) Microcapsules: Same as in Example 1, grinding + ultrasonic pretreatment is the same as in Example 4.

[0190] (2) Emulsion: Same as in Example 1.

[0191] (3) Preservative compound: Same as Example 4.

[0192] (4) Preservation target: longissimus dorsi muscle of pig.

[0193] (5) Preservation process: Same as in Example 4, but the steps of “treating the vacuum-packed meat in an environment of -8℃ for 60s” and “keeping it at -1.3℃ for 11h” are omitted. Instead, the temperature is directly reduced to -1.8℃ at a uniform rate of 0.15℃ / h.

[0194] 4.2 Test Content: (1) Turbidity measurement is the same as described in 3.2 above.

[0195] (2) Determination of preservative penetration depth: The meat samples after immersion treatment were immediately sliced ​​using a cryostat (20 μm thick), placed on a glass slide, and stained with 0.1% methylene blue solution for 2 min. After rinsing with water, the samples were examined under a microscope. The preservative penetration area was blue. The distance (μm) from the surface of the meat to the deepest point of the stained area was measured. Five fields of view were measured for each sample and the average value was taken.

[0196] (3) Juice loss rate determination: The vacuum-packed meat sample was taken out at the predetermined storage time point, the package was opened, the surface free water was absorbed with filter paper and weighed (m1), and compared with the weight of the meat sample before packaging (m0) to calculate the juice loss rate: Juice loss rate (%) = (m0-m1) / m0×100%.

[0197] (4) The total number of colonies was determined as described in section 1.2 above.

[0198] 4.3 Test Results: The effects of different pretreatments on the turbidity stability of the mixed liquor are shown in Table 10. The effects of different impregnation processes on the penetration depth of the preservative are shown in Table 11. The effects of different treatments on the juice loss rate (%) are shown in Table 12. The effects of different treatments on the total bacterial count (log CFU / g) are shown in Table 13.

[0199] Table 10 As shown in Table 10, the turbidity of Example 4 decreased from 218 NTU to 202 NTU within 4 hours, indicating a relatively stable system. The turbidity of Comparative Example 11 decreased from 176 NTU to 92 NTU. This is because the lack of grinding and ultrasonic pretreatment meant that the secondary aggregates formed during microcapsule preparation were not destroyed. These large-particle aggregates settled much faster than monodisperse microcapsules, leading to a rapid decrease in turbidity. Furthermore, the aggregation of microcapsules in Comparative Example 11 not only resulted in a rapid settling rate but also reduced the total surface area per unit mass of microcapsules, leading to a decrease in the effective contact area with the meat and further reducing the amount of adhesion.

[0200] The turbidity change trends of Comparative Examples 12, 13, and 14 were basically consistent with those of Example 4, with a 4-hour decrease of 16 NTU, 18 NTU, and 16 NTU, respectively. The values ​​at each time point fluctuated within the range of ±6 NTU, indicating that the immersion ultrasonic mode and ice-temperature optimization steps did not affect the initial dispersion state of the working fluid, and the turbidity difference only came from random fluctuations during the sampling and measurement process.

[0201] Table 11 As shown in Table 11, the penetration depth of Example 4 was 624 μm, while that of Comparative Example 11 was 538 μm. This is because the microcapsule aggregates in Comparative Example 11 had large particle sizes, resulting in large particle sizes and small specific surface areas on the meat surface. Consequently, the contact efficiency with the meat interface was low, making it difficult for ultrasonic cavitation to push the large particles to deeper layers. At the same time, the aggregates may block the pores on the meat surface, hindering overall penetration.

[0202] As can be seen from Table 11, the penetration depth of Comparative Example 12 is only 208 μm, which is lower than that of Example 4. This is because Comparative Example 12 did not involve immersion ultrasound, and the preservative entered the meat product solely through molecular diffusion; the ultrasonic cavitation effect can disrupt the boundary layer on the surface of the meat product, forming microjets and shock waves, significantly increasing the mass transfer driving force and increasing the penetration depth by about 3 times.

[0203] As can be seen from Table 11, the penetration depth of Comparative Example 13 is 657 μm, which is slightly higher than that of Example 4. This is because Comparative Example 13 uses continuous ultrasound, and the cavitation effect is continuous, with no intermittent mass transfer driving force; however, this slight gain comes at the cost of damage to the muscle fiber structure (see Table 12), and 624 μm is already close to the upper limit of meat thickness, so there is limited practical significance in further increasing the penetration depth.

[0204] As can be seen from Table 11, the penetration depth of Comparative Example 14 is not significantly different from that of Example 4. This is because the penetration depth depends on the impregnation process stage (ultrasonic parameters, time, and microcapsule dispersion state), while the ice temperature optimization step (surface crusting and pre-freezing temperature preservation) is temperature control performed after impregnation and packaging, which affects ice crystal morphology and moisture migration, and does not affect the initial penetration process of the preservative in the meat. Therefore, the penetration depth does not change much compared to Example 4.

[0205] Table 12 As can be seen from Table 12, the juice loss rate of Example 4 was 5.2% after 28 days. In the ultrasonic process of the present invention, 20kHz low-frequency ultrasound is usually used to promote liquid penetration, and 60kHz high-frequency ultrasound is often used to disperse particulate matter; the intermittent application method can reduce the thermal effect caused by continuous ultrasound.

[0206] As shown in Table 12, the juice loss rate of Comparative Example 11 was 7.5% after 28 days, which was higher than that of Example 4. This is because the microcapsules of Comparative Example 11 were unevenly dispersed, the protective film on the surface of the meat was incomplete, and there were defective areas. Moisture preferentially migrated to the surface through these areas and sublimated, resulting in a higher juice loss rate.

[0207] As shown in Table 12, the juice loss rate of Comparative Example 12 after 28 days was 9.4%, which was higher than that of Example 4. This is because Comparative Example 12 did not undergo immersion ultrasound, and the preservative only penetrated to the surface layer of 208 μm, leaving the deeper tissues unprotected. During ice-temperature storage, ice crystals formed and grew in the unprotected deep areas, piercing cell membranes and muscle fibers, resulting in a large release of water after thawing.

[0208] As shown in Table 12, the juice loss rate of Comparative Example 13 after 28 days was 8.7%, which was higher than that of Example 4. This is because although the continuous ultrasound in Comparative Example 13 penetrated deeper, the ultrasound energy continued to act on the meat, causing mechanical damage to the muscle fibers and an increase in local temperature, which reduced the integrity of the cell membrane and made it easier for moisture to seep out during storage.

[0209] As shown in Table 12, the juice loss rate of Comparative Example 14 after 28 days was 6.8%, which was higher than that of Example 4. This is because Comparative Example 14 lacked surface crusting and pre-freezing insulation. According to the conventional process of ice-temperature storage, proper insulation before the temperature drops to the freezing point can make the internal temperature of the meat uniform, which helps to reduce the damage to the cell structure by ice crystals during the subsequent cooling process. In this example, the insulation time (10-12h) is an optimized value determined based on the thickness of 200g meat pieces and thermal conductivity characteristics. Without this step, this effect cannot be obtained.

[0210] Table 13 As shown in Table 13, the total bacterial count in Example 4 after 28 days was 5.7 log CFU / g, while that in Comparative Example 11 was 7.0 log CFU / g. This is because the microcapsules in Comparative Example 11 were unevenly dispersed, and some areas on the surface of the meat were unprotected or poorly protected. Microorganisms preferentially proliferated in these areas, forming localized putrefaction foci, and then spread to the surrounding areas, leading to a rapid increase in the overall bacterial count.

[0211] As shown in Table 13, the total bacterial count of Comparative Example 12 at 28 days was 7.9 log CFU / g, which was higher than that of Example 4. This is because Comparative Example 12 did not use ultrasound and had no deep protection. The putrefactive bacteria migrated to the deeper layers along the pores between muscle bundles and established colonies in the unprotected deep areas, thus damaging the meat from the inside. The surface antibacterial components were unable to control the deep putrefaction.

[0212] As shown in Table 13, the total bacterial count of Comparative Example 13 at 28 days was 6.3 log CFU / g, which was higher than that of Example 4. This is because the continuous ultrasound in Comparative Example 13 resulted in a high juice loss rate. The lost juice is rich in protein and water, which accumulates on the surface of the meat, providing nutrients for residual microorganisms on the surface, allowing them to re-proliferate in the later stages of storage.

[0213] As shown in Table 13, the total bacterial count of Comparative Example 14 at 28 days was 6.6 log CFU / g, which was higher than that of Example 4. This is because Comparative Example 14 lacked ice-temperature optimization, and the intracellular contents released by the loss of juice provided nutrients for the microorganisms. At the same time, the amino acids and nucleotides released by ice crystal damage were more easily utilized by the microorganisms, promoting their growth.

[0214] The preservative and preservation process of this invention can extend the shelf life of fresh meat to 28 days, meeting the preservation cycle requirements for sea freight exports (approximately 21-28 days). Compared with conventional 7-14 day shelf-life preservation technologies, this invention can reduce the cost of discounted sales or air freight substitution due to insufficient shelf life, and has the potential to be applied to the international trade of high-end fresh meat.

[0215] The number of devices and processing scale described herein are for the purpose of simplifying the description of the invention. Applications, modifications, and variations of the invention will be readily apparent to those skilled in the art.

[0216] Although embodiments of the present invention have been disclosed above, they are not limited to the applications listed in the specification and embodiments. They can be applied to various fields suitable for the present invention. For those skilled in the art, other modifications can be easily made. Therefore, without departing from the general concept defined by the claims and their equivalents, the present invention is not limited to the specific details and examples shown and described herein.

Claims

1. A compound antibacterial preservative for fresh meat, characterized in that, It is composed of a mixture of microcapsule components and emulsion components; The microcapsule component is a microcapsule with ε-polylysine as the core and chitosan-nisin complex as the wall. The average particle size of the microcapsule is 10 μm to 50 μm, the mass ratio of the core to the wall is 1:2 to 1:5, and the mass ratio of chitosan to nisin in the chitosan-nisin complex is 10:1 to 20:

1. The emulsion component is an emulsion containing the following components at mass-volume concentrations: sodium D-isoascorbate 0.5 g / L to 2.0 g / L, lysozyme 0.1 g / L to 0.8 g / L, tea polyphenols 0.3 g / L to 1.2 g / L, and glyceryl monostearate 0.5 g / L to 2.5 g / L, in sterile water as the solvent; The microcapsule component and the emulsion component are mixed in a mass ratio of 1:3 to 1:

8. After mixing, the pH value is adjusted to 4.5 to 5.8 with food-grade citric acid to obtain the preservative.

2. The compound antibacterial preservative for fresh meat according to claim 1, characterized in that, The microcapsules were prepared using an emulsification-chemical crosslinking method. The preparation process included: dissolving chitosan and nisin in an acetic acid solution with a mass ratio of 10:1 to 20:1 in a 1% to 3% (v / v) solution to obtain an aqueous phase; adding an emulsifier to the aqueous phase to achieve a mass-volume concentration of 1% to 5% to obtain a mixture; adding the mixture to the oil phase under stirring to form a water-in-oil emulsion; then adding an ε-polylysine aqueous solution to the water-in-oil emulsion, wherein the amount of ε-polylysine added was 20% to 50% of the total mass of chitosan and nisin, and the mass-volume concentration of the ε-polylysine aqueous solution was 10 g / L to 50 g / L; finally, adding a crosslinking agent, wherein the amount of crosslinking agent added was 5% to 15% of the mass of chitosan; the reaction temperature was 30℃ to 50℃; and the reaction time was 2 h to 6 h.

3. The compound antibacterial preservative for fresh meat according to claim 2, characterized in that, The emulsifier consists of a primary emulsifier and a co-emulsifier. The primary emulsifier is Span 80, and the co-emulsifier is Tween 80. The mass ratio of the primary emulsifier to the co-emulsifier is 2:

1.

4. The compound antibacterial preservative for fresh meat according to claim 2, characterized in that, The crosslinking agent is at least one of glutaraldehyde, genipin, or sodium tripolyphosphate.

5. The compound antibacterial preservative for fresh meat according to claim 1, characterized in that, When mixing the microcapsule component with the emulsion component, the microcapsule component is first ground to an average particle size of 5μm to 15μm, then pretreated with an ultrasonic disperser at a frequency of 20kHz to 50kHz for 5min to 8min, and then stirred and mixed at a speed of 200rpm to 600rpm at a temperature of 4℃ to 6℃. After mixing, the turbidity value of the system is stabilized at 50NTU to 80NTU.

6. A low-temperature synergistic preservation process using the composite antibacterial preservative for fresh meat according to any one of claims 1 to 5, characterized in that, Includes the following steps: The microcapsule component and the emulsion component were mixed at a mass ratio of 1:3 to 1:8 and dispersed by stirring at 200 rpm to 600 rpm for 5 to 15 minutes at 4°C to 10°C to obtain the composite preservation working solution. Fresh meat that has undergone post-slaughter inspection is immersed in a compound preservation working solution. The temperature of the working solution is controlled at 0℃ to 4℃, and the immersion time is maintained at 120s to 360s. During this period, alternating 20kHz low-frequency ultrasound and 60kHz high-frequency ultrasound are applied to the working solution. The ratio of the action time of low-frequency ultrasound to high-frequency ultrasound is 1:1 to 1:

3. Remove the soaked fresh meat and place it in a clean environment at 0℃ to 4℃ to air dry until all free moisture on the surface of the fresh meat is removed. The air-dried fresh meat is placed in a sterile workbench and vacuum-sealed using a high-barrier food packaging film, ensuring that the oxygen volume content of the residual air inside the packaging is less than 0.5%. Vacuum-packed fresh meat is placed in a cold storage room and stored at a temperature within ±0.5℃ of its initial freezing point.

7. The low-temperature synergistic preservation process according to claim 6, characterized in that, Alternating low-frequency and high-frequency ultrasound were applied intermittently, with each 30-second ultrasound cycle consisting of 10 seconds of low-frequency ultrasound and 20 seconds of high-frequency ultrasound, followed by a 10-second pause. The ultrasound power density was 0.5 W / cm². 2 Up to 0.6W / cm 2 The distance between the ultrasound probe and the raw meat should be maintained between 5cm and 8cm.

8. The low-temperature synergistic preservation process according to claim 6, characterized in that, During the air-drying step, purified air with a wind speed of 1.0 m / s to 3.0 m / s is used for air drying for 3 to 8 minutes.

9. The low-temperature synergistic preservation process according to claim 6, characterized in that, Before sending fresh meat into the cold storage, the vacuum-packed fresh meat is first treated in an environment of -10℃ to -5℃ for 30 to 90 seconds.

10. The low-temperature synergistic preservation process according to claim 6, characterized in that, In the ice-temperature storage step, the vacuum-packed fresh meat is sent to the ice-temperature storage room for slow cooling. When the temperature drops to 0.5℃ above the initial freezing point of the fresh meat, the cooling is paused and the meat is kept warm for 10 to 12 hours. Then, the temperature continues to drop to the initial freezing point ±0.2℃ at a rate of 0.1℃ / h to 0.2℃ / h.

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