Cold sterilization and anti-browning method for uncooked aquatic products based on polypeptide-polyphenol synergistic medium and high pressure

Abstract

The invention relates to the field of non-thermal processing of aquatic products, in particular to a method for cold sterilization and browning resistance of uncooked aquatic products based on polypeptide-polyphenol synergistic medium-high pressure, which comprises the following steps: preparing natural composite active liquid containing natural antibacterial polypeptide and natural polyphenol; infiltrating the active liquid into the aquatic product by using a pulse vacuum technology; and then carrying out middle-high pressure synergistic treatment under the medium pressure of 300-450 MPa and at normal temperature. According to the method disclosed by the invention, bacterial cell membranes are subjected to sublethal injury and polyphenol oxidase (PPO) conformation is induced to expand by utilizing physical pressure, so that polypeptide components are promoted to efficiently permeate into thalli for sterilization, and polyphenol components are accurately embedded into enzyme active centers for inhibiting browning. According to the method disclosed by the invention, on the premise of avoiding heat damage and high-pressure discoloration, the shelf life of the uncooked aquatic product is remarkably prolonged, and the fresh, tender and compact uncooked texture of the uncooked aquatic product is maintained.

Description

Technical Field

[0001] This invention relates to the field of non-thermal processing of aquatic products. Specifically, it is a method for cold sterilization and anti-browning of raw aquatic products based on the synergistic medium-high pressure treatment of natural antibacterial peptides and natural polyphenols. It is applicable to the safe processing and quality maintenance of raw-grade and high-end ready-to-eat aquatic products. Background Technology

[0002] With the upgrading of consumption, high-end aquatic products such as raw salmon, ready-to-eat shrimp, and ready-to-eat shellfish are favored by the market because they retain their nutritional value and fresh flavor. However, these products face multiple technical challenges in processing and storage.

[0003] First, there are significant microbial safety risks. Raw aquatic products often carry microorganisms such as Pseudomonas, Vibrio, and Enterobacteriaceae, and some strains exhibit a certain degree of tolerance to low-temperature and pressure treatments. Simple refrigeration or a single medium-to-high-pressure treatment can easily put microorganisms in a sub-lethal state, posing a risk of regeneration and revival.

[0004] Secondly, enzymatic browning (blackening) is a serious problem. Polyphenol oxidase (PPO) is widely present in aquatic product tissues and is easily activated during cutting, processing, or storage, triggering a browning reaction that significantly affects the color and commercial value of raw aquatic products.

[0005] Secondly, the quality requirements are extremely high. High-end raw aquatic products are highly sensitive to texture, color, and flavor. Existing high-temperature sterilization or high-dose chemical preservative treatments easily cause protein denaturation and deterioration of taste, making them unacceptable to the high-end market.

[0006] High pressure processing (HPP), as a non-thermal processing technology, can disrupt microbial structure and inhibit enzyme activity to some extent. However, it is often difficult to completely inactivate microorganisms within the 300-450 MPa range, and its long-term inhibitory effect on enzymatic browning is limited. Existing technologies mostly use broad-spectrum antibacterial agents or single polyphenols in synergy with high pressure, which suffers from problems such as insufficient antibacterial specificity, unstable anti-browning effects, or safety controversies.

[0007] Therefore, there is an urgent need for a new non-thermal processing method that can simultaneously achieve cold sterilization and anti-browning under moderate pressure conditions while maintaining the quality of raw aquatic products. Summary of the Invention

[0008] The purpose of this invention is to provide a method for cold sterilization and anti-browning of raw aquatic products based on the synergistic medium and high pressure of polypeptide-polyphenols.

[0009] To achieve the above objectives, a first aspect of the present invention provides a method for cold sterilization and anti-browning of raw aquatic products based on polypeptide-polyphenol synergistic high-pressure treatment, comprising the following steps:

[0010] (a) Raw material pretreatment

[0011] Select aquatic product raw materials that meet the standards for raw or ready-to-eat consumption. The aquatic products are one or a combination of fish, shellfish or crustaceans. Remove inedible parts, wash three times with sterile water at 0-4 ℃, and drain the surface water for later use.

[0012] (b) Preparation of natural compound active liquid

[0013] A natural complex active liquid is prepared, wherein the natural complex active liquid comprises natural antibacterial peptides and natural polyphenols, wherein:

[0014] The concentration of the natural antimicrobial polypeptide is 10–500 mg / L;

[0015] The concentration of the natural polyphenols is 50–1500 mg / L;

[0016] The solvent is food-grade sterile water or citric acid-sodium citrate buffer solution with a pH of 5.5–6.8;

[0017] The natural antimicrobial polypeptide is an aquatic antimicrobial polypeptide, selected from one or more of the grouper antimicrobial peptide Epinecidin-1, fish-derived antimicrobial peptides, or chitosan-derived peptides; preferably, grouper antimicrobial peptide Epinecidin-1.

[0018] The natural polyphenols are selected from one or more of epigallocatechin gallate, gallic acid or its derivatives; preferably epigallocatechin gallate (EGCG).

[0019] (c) Pulse vacuum permeation treatment

[0020] The aquatic products treated in step (a) are immersed in the natural composite active liquid prepared in step (b) (the material-to-liquid ratio is 1:2 (g / mL)) and treated by pulsed vacuum permeation. The vacuum degree of the pulsed vacuum treatment is -0.06 to -0.09 MPa, the vacuum holding time is 1 to 5 min, and the number of pulses is 2 to 5 times, so as to promote the uniform distribution of the natural antibacterial peptides and natural polyphenols in the tissue of the aquatic products.

[0021] (d) Co-processing of medium and high voltage

[0022] The aquatic products processed in step c) are sealed and packaged, and then subjected to medium- and high-pressure treatment at 10–25 °C. The pressure of the medium- and high-pressure treatment is 300–450 MPa, and the treatment time is 2–8 min.

[0023] (e) Cooling and storage

[0024] After the medium- and high-pressure treatment is completed, the aquatic products are quickly cooled to 0-4℃ (preferably 4℃) and then stored in a cold storage.

[0025] Furthermore, the concentration of natural antibacterial peptides in the natural composite active liquid is 300 mg / L; the concentration of natural polyphenols is 1000 mg / L.

[0026] Furthermore, the natural composite active liquid is prepared by dissolving grouper antimicrobial peptide (Epinecidin-1) and epigallocatechin gallate (EGCG) in a citrate-sodium citrate buffer solution at pH 5.5. The concentration of Epinecidin-1 is 300 mg / L, and the concentration of EGCG is 1000 mg / L. The mixture is then filtered through a 0.22 μm microporous membrane for sterilization, yielding a clear and transparent natural composite active liquid.

[0027] Furthermore, the natural composite active liquid may selectively include 200-1500 mg / L (preferably 1000 mg / L) of ε-polylysine as an auxiliary antibacterial agent.

[0028] Further, the pulsed vacuum treatment in step (c) involves evacuating to -0.085 MPa and holding for 4 minutes; then rapidly restoring to atmospheric pressure and holding for 4 minutes; repeating this "vacuum-atmospheric pressure" cycle 3 times. This step utilizes the pressure difference to "pump" the active liquid into the deep layers of muscle tissue, constructing an internal defense network.

[0029] Further, in step (d), the pressure of the medium-high pressure treatment is 350 MPa (crustaceans) or 400 MPa (fish); the initial temperature inside the chamber is set to 10-15℃, and the temperature of the pressure transmission medium is adjusted in real time through the temperature control and cold circulation system of the ultra-high pressure equipment to ensure that the core temperature of the material does not exceed 25℃ during the pressure holding process, so as to maintain the natural texture of the raw food product; the pressure holding time is 5-8 minutes; the pressure increase rate is 200 MPa / min, and the pressure release time is <3 seconds.

[0030] The advantages of this invention are:

[0031] 1. The present invention provides a method for cold sterilization and anti-browning of raw aquatic products based on peptide-polyphenol synergistic medium-high pressure. By constructing a dual synergistic mechanism of "physical pressure damage + biological targeted supplementary killing," the method significantly improves the stability and thoroughness of sterilization. Under moderate pressure of 300-450 MPa, the permeability of microbial cell membranes increases significantly, placing them in a sub-lethal state. At this time, natural antimicrobial peptides can penetrate the damaged cell membrane, destroy the microbial membrane structure, or interfere with its intracellular metabolism, thereby effectively killing sub-lethal microorganisms. This greatly reduces the risk of microbial reactivation after single high-pressure treatment, ensuring the biosafety of raw aquatic products.

[0032] 2. This invention creatively solves the problem of enzymatic browning in raw aquatic products by utilizing the principle of medium- and high-pressure-induced enzyme conformational changes. Through medium-pressure treatment, the spatial conformation of polyphenol oxidase (PPO) is relaxed or unfolded, fully exposing its active site. Natural polyphenol molecules (such as EGCG) can then precisely embed into the enzyme's active site, forming stable compounds with key amino acid residues, thereby achieving long-term stable inhibition of the enzymatic browning reaction, with no significant recovery observed during storage. This mechanism significantly improves the sensory quality stability of high-end aquatic products (such as shrimp and shellfish) without damaging the product's color.

[0033] 3. The method of this invention can achieve efficient preservation while significantly reducing processing pressure (<450 MPa) and shortening processing time. Compared with traditional ultra-high pressure processing of over 600 MPa, this invention avoids problems such as excessive muscle fiber contraction, whitening of meat color (pressure whitening effect), and hardening of texture caused by high-intensity pressure, perfectly preserving the original tender texture, water-holding capacity, and flavor substances of raw aquatic products, providing a gentle and non-destructive processing solution for high-end raw aquatic products.

[0034] 4. The natural antibacterial peptides and natural polyphenols used in this invention are both biologically derived active substances with extremely high safety and biocompatibility, aligning with the modern food industry's "clean label" development trend. This method is not only applicable to various high-end raw aquatic products, but also ensures the uniform distribution of active ingredients within the tissue through pulsed vacuum permeation technology, overcoming the shortcomings of traditional soaking methods such as shallow penetration and poor effectiveness. This technical route is clear, easy to operate, and has low energy consumption, demonstrating good economic benefits and promising prospects for industrialization, providing strong technical support for the green and high-quality development of aquaculture and processing industries. Attached Figure Description

[0035] Figure 1 This is a schematic diagram of the process for the present invention of cold sterilization and anti-browning method for raw aquatic products based on polypeptide-polyphenol synergy under medium and high pressure.

[0036] Figure 2This is a schematic diagram showing the changes in total bacterial count (TVC) of raw shrimp in different treatment groups during storage, as per the method of this invention.

[0037] Figure 3 This is a schematic diagram showing the changes in TVB-N content of raw shrimp in different treatment groups during storage, as described in the method of this invention.

[0038] Figure 4 This is a schematic diagram showing the changes in total bacterial count (TVC) of raw salmon slices in different treatment groups during storage, as described in the method of this invention.

[0039] Figure 5 This is a schematic diagram showing the changes in TVB-N content of raw salmon fillets in different treatment groups during storage, as per the method of this invention. Detailed Implementation

[0040] The specific implementation methods provided by the present invention will be described in detail below with reference to the embodiments.

[0041] Example 1: A method for cold sterilization and anti-browning of raw aquatic products based on peptide-polyphenol synergistic medium and high pressure

[0042] Figure 1 This is a schematic flowchart of the present invention, which describes a method for cold sterilization and anti-browning of raw aquatic products based on the synergistic medium-high pressure of polypeptide-polyphenols, including the following steps:

[0043] The first step is the pretreatment of raw materials and the preparation of the active liquid.

[0044] Select fresh, raw-eating grade aquatic products (such as whiteleg shrimp or salmon), remove inedible parts, wash three times with sterile water at 0~4℃, and drain the surface moisture.

[0045] Preparation of the natural complex active solution: Epinecidin-1 (purity ≥95%, purchased from Shanghai Qiangyao Biotechnology Co., Ltd.) and epigallocatechin gallate (EGCG) (purity ≥98%, purchased from Chengdu Institute of Biological Products Co., Ltd.) were dissolved in a citrate-sodium citrate buffer solution at pH 5.5. The concentration of Epinecidin-1 was 300 mg / L, and the concentration of EGCG was 1000 mg / L. The mixture was filtered through a 0.22 μm microporous membrane for sterilization to obtain a clear and transparent natural complex active solution.

[0046] The second step is pulsed vacuum permeation treatment.

[0047] The pretreated aquatic products were completely immersed in the above-mentioned natural compound active liquid at a material-to-liquid ratio of 1:2 (g / mL). The container was then placed in a vacuum tumbler for pulsed vacuum treatment.

[0048] The specific parameters of the pulsed vacuum are as follows: evacuate to -0.085 MPa and maintain for 4 minutes; then quickly restore to normal pressure and maintain for 4 minutes; repeat the above "vacuum-normal pressure" cycle 3 times. This step utilizes the pressure difference to "pump" the active liquid into the deep layers of muscle tissue, constructing an internal defense network.

[0049] The third step involves vacuum packaging and combined medium- and high-pressure processing.

[0050] After the aquatic products have undergone osmosis treatment, remove them, drain off the surface moisture, and pack them into high-barrier EVOH / PE composite packaging bags (oxygen permeability <0.5 cm). 3 / m 2 (24h·0.1MPa), sealed using a vacuum packaging machine, with a vacuum degree ≥-0.095MPa.

[0051] The packaged samples were placed in the processing chamber of the ultra-high pressure equipment (HPP-600 type, Baotou Kefa High Pressure Technology Development Co., Ltd.), with pure water as the pressure transmission medium.

[0052] The processing parameters are set as follows: pressure 350 MPa (shrimp) or 400 MPa (salmon); initial temperature inside the chamber is set to 10-15℃; the temperature of the pressure transmission medium is adjusted in real time through the temperature control and cold circulation system of the ultra-high pressure equipment to ensure that the core temperature of the material does not exceed 25℃ during the pressure holding process, so as to maintain the natural texture of raw food products; pressure holding time is 5-8 minutes; pressure increase rate is 200 MPa / min; pressure release time is <3 seconds.

[0053] The fourth step is depressurization, refrigeration, and indicator monitoring.

[0054] After processing, remove the product, wipe the surface of the packaging bag dry, and store it in a 4°C constant temperature refrigerator away from light.

[0055] Starting from day 0 of the experiment, samples were collected and tested at different time points (such as days 3, 6, 9, 12, and 15). The monitoring indicators included color score, total bacterial count, TVB-N value, etc., to evaluate the preservation effect.

[0056] Example 2:

[0057] This embodiment tests the anti-metastasis and antibacterial effects of the technical solution of the present invention on raw Litopenaeus vannamei shrimp. Three treatment groups were set up in the experiment: a synergistic treatment group (T1), a single high-pressure group (T2), and a blank control group (T3). These treatment groups simulated different processing conditions to obtain the preservation effect of the present invention under different conditions.

[0058] Synergistic treatment group (T1, i.e., the present invention): treated with "pulse vacuum permeation of active liquid + 350 MPa medium and high pressure".

[0059] Single high-pressure group (T2): Only "350 MPa medium and high pressure" treatment is performed, without adding active liquid.

[0060] Blank control group (T3): No treatment was performed, and the shrimp were directly vacuum-packed (simulating commercially available raw shrimp).

[0061] Experimental environment and sample collection: 500g of fresh whiteleg shrimp meat of the same size (about 15g each) were selected for each treatment group.

[0062] Active solution parameters: Epinecidin-1 concentration 300 mg / L, EGCG concentration 1000 mg / L.

[0063] HPP parameters: pressure 350 MPa, temperature 20℃, pressure holding time 5 min.

[0064] Testing frequency: days 0, 3, 6, 9, 12, and 15. Three independently packaged samples were randomly selected at each time point for parallel testing (n=3), and the results are expressed as mean ± standard deviation.

[0065] To ensure sample integrity, collection was performed in a sterile operating table to avoid secondary contamination. Immediately after collection, blackening score (sensory indicator), total bacterial count (microbiological indicator), and TVB-N value (physicochemical indicator) were analyzed.

[0066] Data Analysis: Experimental data analysis is based on tables and charts, including the following:

[0067] 1. Sensory index analysis: Blackening score.

[0068] Melting is the primary factor affecting the commercial value of crustacean aquatic products. A 10-point scoring method was used (0 points = no melaninization, 10 points = severe melaninization), and the results are shown in Table 1.

[0069] Table 1. Changes in melanosis score of raw shrimp in different treatment groups (n=3)

[0070] (On a 10-point scale, 0 = no blacklisting, 10 = severe blacklisting)

[0071]

[0072] Note: Different lowercase letters (a, b, c) after the same row of data indicate significant differences between groups (P < 0.05).

[0073] Data analysis: As can be seen from Table 1, there are significant differences in the blackening scores of the three groups of samples (P<0.05).

[0074] Synergistic treatment group (T1, i.e., the present invention): During the 15-day storage period, the melanosis score increased extremely slowly. Until day 15, the score was only 1.20 ± 0.18, and the shrimp body still maintained a translucent jade color, with excellent sensory acceptance. On day 15, the score of the synergistic treatment group (T1) was significantly lower than that of the single high-pressure group (T2) and the blank control group (T3), indicating that the process of the present invention has a statistically significant advantage in inhibiting melanosis, and the process has a highly stable and uniform inhibitory effect on PPO enzyme.

[0075] High-pressure group (T2): The score rose rapidly over time, reaching 2.51 on day 3 (visible black spots appeared) and as high as 6.51 on day 15 (severe black spots). This indicates that the moderate pressure of 350 MPa can only temporarily reduce PPO enzyme activity, but cannot cause irreversible inactivation. Enzyme activity recovery during storage leads to browning.

[0076] Blank control group (T3): showed a linear deterioration trend, with the score exceeding 6.0 on day 6, thus losing its commercial value.

[0077] This result demonstrates that the present invention utilizes the mechanism of medium-high pressure inducing conformational changes in PPO enzymes and synergistically targeting and embedding EGCG into the active site to successfully achieve permanent inhibition of enzymatic browning, with significantly better results than single physical treatment.

[0078] 2. Microbiological indicator analysis: Total bacterial count (TVC)

[0079] Total bacterial count is a core indicator for evaluating food hygiene and safety. Referring to the relevant microbial limits for raw aquatic products in GB 10136-2015 "National Food Safety Standard for Animal-Derived Aquatic Products" (usually with an upper limit of 5.0 lg CFU / g), the results are as follows: Figure 2 As shown.

[0080] Chart analysis (based on) Figure 2 (Cumulative count change curve)

[0081] T1 (This invention): The curve is nearly horizontal. The initial bacterial count was 2.10 ± 0.10 lg CFU / g, which only slightly increased to 2.30 ± 0.15 lg CFU / g on day 15 of storage. Throughout the shelf life, the total bacterial count remained at an extremely low level of 2.0~2.5, far below the national standard limit (5.0). This indicates that the dual mechanism of "pressure sensitization + peptide penetration" not only provides thorough instantaneous sterilization, but also that the long-lasting antibacterial properties of Epinecidin-1 effectively prevent the repair and proliferation of residual microorganisms.

[0082] T2 (single high pressure): The curve shows a "J"-shaped increase. Although the initial sterilization effect was acceptable (3.51 lg CFU / g), the slope increased sharply after day 9, exceeding the 5.0 limit on day 12, and reaching 5.80 ± 0.88 lg CFU / g on day 15. This confirms that the bacterial damage caused by single medium pressure treatment is repairable (resurrection from a sublethal state), posing a potential food safety hazard.

[0083] T3 (blank control): showed exponential rapid growth, exceeding the limit standard on day 3, and reaching 9.20 lgCFU / g on day 15, indicating severe spoilage.

[0084] This result demonstrates that the present invention overcomes the technical challenges of incomplete sterilization under medium pressure and bacterial reactivation by constructing a physical-biological barrier, thus ensuring the biosafety of the product within 15 days.

[0085] 3. Physicochemical index analysis: Volatile basic nitrogen (TVB-N)

[0086] TVB-N reflects the degree of protein breakdown and is the gold standard for freshness evaluation. The limit for raw consumption is ≤ 25 mg / 100g, and for Grade 1 freshness, ≤ 15 mg / 100g. Results are as follows... Figure 3 As shown.

[0087] Chart analysis (based on) Figure 3 TVB-N content change curve):

[0088] T1 (Invention): The curve is flat, with a lagging growth rate. On day 15, the TVB-N content was 12.51 ± 2.05 mg / 100g, still within the first-grade freshness range. This indicates that the synergistic treatment effectively inhibited the metabolic activity of amine-producing bacteria (such as Pseudomonas and Shewanella), significantly delaying the spoilage and degradation of shrimp meat protein.

[0089] T2 (single high pressure): The growth was relatively rapid, reaching 22.84 ± 3.48 mg / 100g on day 15. Although it was barely qualified, it was close to the deterioration threshold and the standard deviation was large, indicating that the product quality was unstable.

[0090] T3 (blank control): The dose was close to the limit of 25 mg / 100g on day 6 and reached as high as 45.00 mg / 100g on day 15, producing a strong putrid odor.

[0091] This result shows that the technical solution of the present invention extends the shelf life of raw shrimp with first-grade freshness to more than 15 days, which is significantly better than the existing technology.

[0092] Based on the above sensory, microbiological, and physicochemical analyses, the "pulsed vacuum permeation + polypeptide-polyphenol synergy + medium-high pressure treatment" technology employed in this invention simultaneously achieves: thorough anti-browning effect; excellent cold sterilization effect; and ultimate freshness preservation under a gentle pressure of 350 MPa. This result demonstrates that this invention solves the industry pain point of traditional single technologies being unable to simultaneously address both "sterilization" and "color protection," and possesses extremely high application value.

[0093] Example 3:

[0094] This embodiment tests the preservation effect of the technical solution of the present invention on raw Atlantic salmon fillets. Given the high lipid content and different composition of putrefactive bacteria in fish muscle tissue, to further enhance the antibacterial stability and synergistic antioxidant effect, this embodiment adaptively adjusts the proportion of active components based on the aforementioned optimized system and introduces ε-polylysine as an auxiliary antibacterial component. Three treatment groups were set up in the experiment: synergistic treatment group (T1), single high-pressure group (T2), and blank control group (T3).

[0095] Synergistic treatment group (T1, i.e., the present invention): treated with "pulse vacuum permeation of active liquid + 400 MPa medium and high pressure".

[0096] Single high-voltage group (T2): Only "400 MPa medium and high voltage" treatment is performed.

[0097] Blank control group (T3): No treatment was given.

[0098] Experimental environment and sample collection: Fresh salmon midsection was used for each treatment group and cut into 1.0 cm thick slices.

[0099] Active solution parameters: Epinecidin-1 200 mg / L + EGCG 500 mg / L + ε-polylysine 1000 mg / L.

[0100] Testing frequency: days 0, 3, 6, 9, 12, 15, and 18. Three independently packaged samples were randomly selected at each time point for parallel testing (n=3), and the results are expressed as mean ± standard deviation.

[0101] To ensure sample integrity, collection was performed in a sterile operating table to avoid secondary contamination. Immediately after collection, color deterioration scoring (sensory indicator), total bacterial count (microbiological indicator), and TVB-N value (physicochemical indicator) were analyzed.

[0102] Data Analysis: Experimental data analysis is based on tables and charts, including the following:

[0103] 1. Sensory index analysis: Color deterioration score.

[0104] The color of salmon is the primary factor in judging its freshness. Considering the tendency of salmon to fade and brown, a color deterioration scoring method was used (0 points = bright orange-red / excellent, 10 points = severe browning / discoloration / unacceptable). The results are shown in Table 2.

[0105] Table 2. Color deterioration score of raw salmon in different treatment groups during storage (n=3) (10-point scale, 0=bright, 10=browning / discoloration)

[0106]

[0107] Note: Different lowercase letters (a, b, c) after the data in the same row indicate significant differences between groups (P < 0.05).

[0108] Data analysis: As can be seen from Table 2, there are significant differences in the color scores of the three groups of samples (P<0.05).

[0109] Co-processing group (T1, i.e., the present invention): The score increased extremely slowly. Throughout the 18-day storage period, the fish meat maintained an appealing orange-red color with clear fat texture. Until day 18, the score was only 1.49 ± 0.22, still within the excellent sensory acceptance range. On day 18, the score of group T1 was significantly lower than that of groups T2 and T3, indicating that the EGCG permeated into the system, as a powerful antioxidant, effectively protected myoglobin and astaxanthin from oxidation, achieving excellent color preservation.

[0110] The single high-pressure group (T2) showed a significant increase in the mid-to-late stages, reaching 5.51 on day 18, with noticeable discoloration and a dull appearance of the fish meat. This indicates that single physical pressure (400 MPa) accelerated the oxidation of ferromyoglobin to some extent and failed to inhibit the yellowing caused by fat auto-oxidation.

[0111] Blank control group (T3): The color deteriorated rapidly, and the score exceeded 6.0 on day 9, indicating severe browning and loss of commercial value.

[0112] This result demonstrates that the "peptide-polyphenol synergistic medium-high pressure" technology employed in this invention can effectively overcome the common problem of "color deterioration" in traditional ultra-high pressure processing, and significantly extend the sensory shelf life of high-end raw salmon.

[0113] 2. Microbiological indicator analysis: Total bacterial count (TVC)

[0114] Total bacterial count is a core indicator for evaluating food hygiene and safety. Referring to the relevant microbial limits for raw aquatic products in GB 10136-2015 "National Food Safety Standard for Animal-Derived Aquatic Products" (usually with an upper limit of 5.0 lg CFU / g), the results are as follows: Figure 4 As shown (Note: This is a textual description of the chart content).

[0115] Chart analysis (based on) Figure 4 (Cumulative count change curve)

[0116] T1 (This invention): The curve is flat. The initial bacterial count was 2.50 ± 0.13 lg CFU / g, and the growth was slow throughout the 18-day storage period, reaching only 3.82 ± 0.52 lg CFU / g on day 18, never exceeding the national standard limit (5.0). This indicates that the combined use of Epinecidin-1 and ε-polylysine, under pressure assistance, penetrates deep into the fish meat, demonstrating a strong and long-lasting inhibitory effect on fish spoilage bacteria.

[0117] T2 (single high pressure): The inhibitory effect was acceptable for the first 6 days, but it began to rise rapidly from day 9 (4.64 lg CFU / g), exceeded the 5.0 limit on day 12, and reached 6.18 ± 0.94 lg CFU / g on day 18. This confirms that single medium pressure treatment only causes sublethal damage to some pressure-resistant bacteria, and there is a risk of reactivation in the later stage.

[0118] T3 (blank control): showed exponential growth, exceeding the limit of 5.0 on day 6 and reaching as high as 8.53 lg CFU / g on day 18, indicating severe spoilage.

[0119] This result demonstrates that the technical solution of this invention successfully solved the problem of microbial reactivation under medium pressure conditions, ensuring the biosafety of raw salmon during its 18-day shelf life.

[0120] 3. Physicochemical index analysis: Volatile basic nitrogen (TVB-N)

[0121] TVB-N is the gold standard for evaluating the freshness of aquatic products. Referring to GB 10136-2015, the limit for raw consumption is ≤25 mg / 100g, and for Grade 1 freshness it is ≤15 mg / 100g. The results are as follows... Figure 5 As shown.

[0122] Chart analysis (based on) Figure 5 TVB-N content change curve):

[0123] T1 (This invention): The growth curve is extremely slow. From 6.01 mg / 100g on day 0 to 13.52 ± 2.10 mg / 100g on day 18, it remained below 15 mg / 100g throughout. Remarkably, even on day 18, the product remained at Grade 1 freshness. This indicates that the synergistic treatment effectively disrupted the metabolic pathway of amine-producing bacteria, significantly delaying protein spoilage and decomposition.

[0124] T2 (single high pressure): The increase was relatively rapid, reaching 24.2 mg / 100g on day 15 (close to the limit), and 28.87 ± 4.50 mg / 100g on day 18, which exceeded the national standard limit and was judged to be deteriorated.

[0125] T3 (blank control): On day 6, the dose was close to the limit (22.69 mg / 100g), on day 9 it was seriously exceeded (36.19 mg / 100g), and on day 18 it reached as high as 55.00 mg / 100g, indicating extremely rapid putrefaction.

[0126] This result shows that the technical solution of the present invention extends the shelf life of raw salmon to more than 18 days, which is more than three times longer than the blank control group (about 5 days).

[0127] Based on the above sensory, microbiological, and physicochemical analyses, the "pulsed vacuum permeation + polypeptide-polyphenol synergy + 400 MPa treatment" technology employed in this invention simultaneously achieves: perfect color retention; long-lasting antibacterial ability; and ultimate freshness locking in the preservation of raw salmon. This result demonstrates that this invention is not only applicable to crustaceans but also possesses significant preservation advantages for high-fat fish, achieving a comprehensive reconstruction of the quality of high-end raw aquatic products.

[0128] The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited to the embodiments described. Those skilled in the art can make various equivalent modifications or substitutions without departing from the spirit of the present invention, and these equivalent modifications or substitutions are all included within the scope defined by the claims of this application.

Claims

1. A method for cold sterilization and anti-browning of raw aquatic products based on polypeptide-polyphenol synergistic medium-high pressure, characterized in that, Includes the following steps: (a) Raw material pretreatment Select aquatic product raw materials that meet the standards for raw or ready-to-eat consumption. The aquatic products are one or a combination of fish, shellfish or crustaceans. Remove inedible parts, wash with sterile water at 0-4 ℃, and drain the surface moisture for later use. (b) Preparation of natural compound active liquid The natural composite active liquid includes natural antibacterial peptides and natural polyphenols: The concentration of the natural antimicrobial polypeptide is 10–500 mg / L; The concentration of the natural polyphenols is 50–1500 mg / L; The solvent is food-grade sterile water or citric acid-sodium citrate buffer solution with a pH of 5.5–6.8; The natural antimicrobial polypeptide is an aquatic antimicrobial polypeptide selected from one or more of the grouper antimicrobial peptide Epinecidin-1, fish-derived antimicrobial peptides, or chitosan-derived peptides. The natural polyphenols are selected from one or more of epigallocatechin gallate, gallic acid or its derivatives; (c) Pulse vacuum permeation treatment The aquatic products treated in step (a) are immersed in the natural compound active liquid prepared in step (b) and treated by pulsed vacuum permeation. The vacuum degree of the pulsed vacuum treatment is -0.06 to -0.09 MPa, the vacuum holding time is 1 to 5 minutes, and the number of pulses is 2 to 5. (d) Co-processing of medium and high voltage The aquatic products processed in step c) are sealed and packaged, and then subjected to medium- and high-pressure treatment at 10–25 °C. The pressure of the medium- and high-pressure treatment is 300–450 MPa, and the treatment time is 2–8 min. (e) Cooling and storage After the medium- and high-pressure treatment is completed, the aquatic products are quickly cooled to 0-4 ℃ and then stored in cold storage.

2. The method for cold sterilization and anti-browning of raw aquatic products based on polypeptide-polyphenol synergistic high pressure as described in claim 1, characterized in that, The concentration of natural antibacterial peptides in the natural composite active liquid is 30 mg / L.

3. The method for cold sterilization and anti-browning of raw aquatic products based on polypeptide-polyphenol synergistic high pressure as described in claim 1, characterized in that, The concentration of natural polyphenols in the natural composite active liquid is 100 mg / L.

4. The method for cold sterilization and anti-browning of raw aquatic products based on polypeptide-polyphenol synergistic high pressure as described in claim 1, characterized in that, The natural composite active solution is prepared by dissolving the grouper antimicrobial peptide Epinecidin-1 and epigallocatechin gallate (EGCG) in a citrate-sodium citrate buffer solution at pH 5.5; wherein the concentration of Epinecidin-1 is 300 mg / L and the concentration of EGCG is 1000 mg / L.

5. The method for cold sterilization and anti-browning of raw aquatic products based on polypeptide-polyphenol synergistic high pressure according to claim 1, characterized in that, The natural composite active liquid also includes 20~1500 mg / L of ε-polylysine.

6. The method for cold sterilization and anti-browning of raw aquatic products based on polypeptide-polyphenol synergistic high pressure according to claim 1, characterized in that, The pulsed vacuum treatment in step (c) involves evacuating to -0.085 MPa and holding for 4 minutes; then rapidly restoring to atmospheric pressure and holding for 4 minutes; repeating the above "vacuum-atmospheric pressure" cycle 3 times.

7. The method for cold sterilization and anti-browning of raw aquatic products based on polypeptide-polyphenol synergistic high pressure as described in claim 1, characterized in that, The pressure for the medium-high pressure treatment in step (d) is 350 MPa for crustaceans or 400 MPa for fish; the initial temperature inside the chamber is set to 10-15℃, and the temperature of the pressure transmission medium is adjusted in real time through the temperature control and cold circulation system of the ultra-high pressure equipment to ensure that the core temperature of the material does not exceed 25℃ during the pressure holding process, so as to maintain the natural texture of the raw food products; the pressure holding time is 5-8 minutes; the pressure increase rate is 200 MPa / min, and the pressure release time is <3 seconds.

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