A method and system for preventing and controlling traditional dry-cured ham cheese fly and insect pests in stages

Abstract

The application discloses a kind of stage type air conditioning prevention and control traditional dry-cured ham cheese fly pest method and system, according to the fermentation stage of ham and / or cheese fly life history, carry out multiple independent air conditioning treatment cycles at multiple predetermined time points, break through the air conditioning application limit "fresh-keeping" rather than "fermentation" traditional cognition.Flavor quality, metabolomics and microbiome technology are integrated to establish a stage type air conditioning pest control technology system suitable for Yunnan dry-cured ham, effectively inhibit cheese fly pest while maintaining the normal fermentation process and quality characteristics of ham, provide new ideas and new methods for replacing chemical fumigation and traditional fermented meat products Green control has important significance to promote the sustainable development of Yunnan ham industry.

Description

Technical Field

[0001] This invention belongs to the field of food processing and storage technology, specifically relating to a green control method for cheese fly pests during the long-term fermentation and maturation process of traditional dry-cured ham, and in particular a staged modified atmosphere treatment method that does not interfere with the normal fermentation process of ham and synergistically improves its quality. Background Technology

[0002] Yunnan dry-cured ham, a typical representative of traditional Chinese fermented meat products, enjoys a high reputation both domestically and internationally for its unique regional flavor and long processing history. The traditional processing cycle for Yunnan ham typically lasts 8-12 months, and can even reach 36 months, involving complex processes such as salting, washing and drying, air-drying, and fermentation. However, the infestation of cheese flies (commonly known as ham worms or cheese flies) on the surface and in the crevices of the traditionally open-fermented ham is one of the most serious and difficult-to-eradicate biological hazards.

[0003] The cheese fly belongs to the family Typhonidae in the order Diptera. Its larvae (maggots) possess a strong ability to hydrolyze proteins and are highly adaptable to their environment. Overwintering adults become active in April and May as temperatures rise (daily average temperature > 18℃), with peak egg-laying occurring from June to August. A single female can lay 300-500 eggs, which hatch into first-instar larvae in 24-48 hours at 25-30℃ and 70-80% relative humidity. The larvae feed on ham muscle and adipose tissue, secreting potent proteases and lipases through their mouthparts. During feeding, they cause liquefaction of muscle tissue and saponification of adipose tissue, resulting in noticeable tunnels and putrefactive spots.

[0004] Existing prevention and control technologies mainly include: 1. Chemical control: mainly relies on aluminum phosphide fumigation and pyrethroid spraying. Aluminum phosphide releases highly toxic phosphine (PH3) gas by reacting with moisture in the air. Although it is fast-acting and low-cost, it has the problem of phosphine residue (0.05-0.12 mg / kg of residue accumulates in the fat tissue of ham after traditional fumigation treatment, exceeding the EU limit of 0.01 mg / kg). Moreover, long-term use of this single agent has led to a significant increase in the resistance of strata fly populations (in some areas of Yunnan, strata flies have developed resistance to deltamethrin by 15-20 times).

[0005] 2. Physical control methods: These include low-temperature freezing (below -18℃), high-temperature heat treatment (>50℃), irradiation (Co-60 gamma rays, dose 3-5 kGy), and controlled atmosphere storage. While low-temperature freezing can effectively kill insects at all stages, it can cause ice crystal damage to ham muscle tissue, increase juice loss after thawing (>15%), and interrupt the fermentation process, affecting the continuous accumulation of flavor compounds. Irradiation treatment induces a chain reaction of lipid oxidation, producing the typical "irradiation off-flavor."

[0006] 3. Biological control: Entomopathogenic nematodes and Beauveria bassiana are pathogenic to jack fly larvae, but in the high-salt (NaCl content > 8%) and low-water-activity (Aw < 0.85) environment on the surface of ham, the survival rate and infectivity of biological control agents are significantly reduced.

[0007] The aforementioned prior art has the following main defects and problems: 1. Chemical fumigation has problems with pesticide residues and environmental pollution, which does not meet the requirements of green manufacturing, and pests are becoming increasingly resistant to pesticides.

[0008] 2. Physical prevention methods (low-temperature freezing, irradiation) can interfere with the normal fermentation process of ham, affecting the formation of flavor and quality, and consumers have a low acceptance of irradiated food.

[0009] 3. Existing prevention and control technologies focus on the single indicator of "insecticide efficiency" and neglect the special characteristics of ham as a living fermentation system - any external intervention will affect the quality formation through the microecological-chemical network.

[0010] 4. There is a lack of graded prevention and control strategies targeting the stage characteristics of Yunnan ham fermentation, and fixed treatment methods are difficult to adapt to the needs of different fermentation stages from May to October.

[0011] Therefore, based on the characteristics of the life history of chrysalis flies at different stages of ham fermentation (early, middle, and late stages) and the fermentation requirements of ham itself, it is crucial to study a solution that can "kill insects without harming the fermentation process, preserve quality, and increase efficiency". Summary of the Invention

[0012] To address the problems existing in the background art, the purpose of this invention is to provide a staged modified atmosphere packaging method for controlling cheese fly infestation in traditional dry-cured ham, in order to solve the following technical problems: 1. Provide a green, chemical-residue-free technology for controlling jackfly pests, replacing traditional aluminum phosphide fumigation, and avoiding pesticide residues and environmental pollution.

[0013] 2. Develop a control strategy that balances "insecticide efficiency" and "fermentation quality" to effectively block the life cycle of jack flies while maintaining the normal fermentation process of ham.

[0014] 3. Enhance the antioxidant activity of ham, giving traditional fermented meat products new health benefits.

[0015] To achieve the above objectives, the present invention provides the following technical solution: A staged modified atmosphere packaging (MAP) method for controlling cheese fly infestation in traditional dry-cured ham involves treating the ham multiple times at predetermined time points, based on the fermentation and maturation process of the dry-cured ham (divided into early, middle, and late stages) and the life cycle of the cheese fly. Each MAP treatment includes placing the ham in a sealed environment, introducing a predetermined concentration of CO2 gas, maintaining specific duration and environmental conditions (temperature, relative humidity), and then restoring it to the natural environment for continued fermentation after the treatment. By using different CO2 concentrations at different stages (e.g., high concentration during the middle stage when the infestation is high, and lower concentration during the early and late stages), complete suppression of the cheese fly infestation and positive synergistic regulation of the ham fermentation quality are achieved.

[0016] Specifically, the following steps are included: S1. Fermentation Stage Identification and Pretreatment: Identify the fermentation stage of the dry-cured ham, which should be at least divided into: early stage (May-June), middle stage (July-August), and late stage (September-October). Place the hams to be processed in a sealed modified atmosphere unit, maintaining sufficient spacing between the hams (e.g., >15cm).

[0017] S2. Staged Controlled Atmosphere Processing: Based on the identified fermentation stage, execute the corresponding controlled atmosphere processing cycle for that stage. Each cycle includes: S2.1 Initial Controlled Atmosphere Treatment: In the initial stage of fermentation, a relatively low CO2 concentration is used for treatment. For example, the temperature is 25±1℃, the relative humidity is 75±5%, the CO2 concentration is 20%-35%, and the treatment time is 7 days.

[0018] S2.2 Mid-term controlled atmosphere treatment: During the mid-fermentation period (peak period of strata fly infestation), a higher CO2 concentration is used for treatment. For example, the temperature is 25±1℃, the relative humidity is 75±5%, the CO2 concentration is 35%-40%, and the treatment time is 7 days.

[0019] S2.3 Later-stage controlled atmosphere treatment: In the later stage of fermentation, a relatively low CO2 concentration is used for treatment again. For example, the temperature is 25±1℃, the relative humidity is 75±5%, the CO2 concentration is 20%-35%, and the treatment time is 7 days.

[0020] S3. Restoration and Natural Fermentation: After the single controlled atmosphere treatment, CO2 is slowly released (e.g., depressurization rate < 0.01 MPa / min) to restore the environment to normal pressure and natural air. The ham is then transferred back to a conventional fermentation workshop or kept well-ventilated to continue natural maturation until the next stage of controlled atmosphere treatment or the final product matures.

[0021] In the above controlled atmosphere treatment, the O2 concentration of the mixed gas is less than 1%. After the controlled atmosphere treatment is completed, there is also a 0-10 day in-store settling period.

[0022] This method primarily relies on a controlled atmosphere processing system, including: Hermetically Controlled Atmosphere Unit (HATU): One or more sealable HAT units / cabins for containing ham and providing an airtight environment. Includes gas inlet / outlet, sealed doors, and internal shelving.

[0023] Gas supply and distribution unit: Includes a CO2 gas source (such as a gas storage tank), gas mixing device, pipelines, and valves, used to fill the gas conditioning unit with CO2 gas at a preset concentration. It may include an N2 gas source for dilution or pressure balancing.

[0024] Environmental control and monitoring unit: Includes temperature sensor, humidity sensor, CO2 concentration sensor, and controller connected to heating / cooling and humidification / dehumidification devices. Used for real-time monitoring and adjustment of temperature, relative humidity, and CO2 concentration within the controlled atmosphere unit.

[0025] Control unit: PLC or industrial computer, connected to the gas supply and distribution unit and the environmental control and monitoring unit, used to automatically execute processes such as gas filling, pressure holding, gas exhaust, and environmental control according to preset staged gas control parameters (time, CO2 concentration, temperature and humidity).

[0026] Data flow: User inputs parameters for each stage to the control unit → Control unit controls environmental control and monitoring unit to start environmental pre-conditioning → Control unit controls gas supply and distribution unit to fill the sealed atmosphere control unit with CO2 of a set concentration → Environmental control and monitoring unit monitors and feeds back data to control unit in real time → Control unit maintains environmental stability for a preset time through environmental control and monitoring unit and gas supply and distribution unit → Control unit controls sealed atmosphere control unit to exhaust gas and restore normal pressure.

[0027] This invention achieves the following technical effects by using a staged controlled atmosphere treatment method and dynamically adjusting the CO2 concentration at different fermentation stages: 1. Highly effective insecticidal effect: The high-concentration modified atmosphere group (35%-40% CO2) achieved a 99.6% inhibition rate against the emergence of botfly pupae, while the low-concentration group (20%-35% CO2) also achieved a 97.8% inhibition effect. Moreover, there was no "delayed emergence" phenomenon within 10 days after treatment, and the control effect was certain and thorough.

[0028] 2. Maintaining normal fermentation process: Modified atmosphere treatment has little impact on key physicochemical indicators of ham such as water activity, pH value, crude protein, crude fat, and ash content. There are no significant differences between the treatment groups and the control group, indicating that the staged modified atmosphere technology can effectively control pests while maintaining the normal physicochemical properties of ham.

[0029] 3. Inhibition of lipid oxidation: The MDA content in the high-concentration modified atmosphere group was 40.5%-57.1% lower than that in the control group, which effectively inhibited lipid oxidation and extended the product shelf life.

[0030] 4. Enhanced antioxidant activity: High-concentration modified atmosphere treatment significantly enhances the antioxidant activity of ham. The DPPH, ABTS free radical scavenging capacity and hydroxyl free radical scavenging capacity are increased by 15%-25% compared with the control group, giving the product higher nutritional and health value.

[0031] 5. Maintaining flavor quality: GC-MS identified 128 volatile flavor compounds. The OAV values ​​of key aroma active substances remained at a high level. The modified atmosphere treatment group still maintained the typical flavor characteristics of Yunnan dry-cured ham, and the overall acceptability score reached 8.5 points.

[0032] 6. Green and environmentally friendly: This invention uses a purely physical method to control pests, leaving no chemical residues. Attached Figure Description

[0033] Figure 1 This is a flowchart of the method for controlling cheese fly infestation in traditional dry-cured ham using a staged modified atmosphere packaging according to the present invention.

[0034] Figure 2 This is a comparison chart showing the effect of different treatment groups on the inhibition rate of tyrosinus emergence in embodiments of the present invention.

[0035] Figure 3 This represents the change in cumulative feathering inhibition rate during the static period in the storage chamber after controlled atmosphere treatment in this embodiment of the invention.

[0036] Figure 4 This describes the dynamic changes in the water activity of ham during the staged modified atmosphere treatment in this embodiment of the invention.

[0037] Figure 5 This describes the dynamic changes in the moisture content of ham during the staged modified atmosphere processing in this embodiment of the invention.

[0038] Figure 6 This describes the dynamic change of ham pH value during the staged modified atmosphere treatment in this embodiment of the invention.

[0039] Figure 7 This illustrates the dynamic changes in crude fat content of ham during the staged modified atmosphere processing in this embodiment of the invention.

[0040] Figure 8 This describes the dynamic changes in crude protein content of ham during the staged modified atmosphere treatment in this embodiment of the invention.

[0041] Figure 9 This is a dynamic change graph showing the effect of different treatment groups on the malondialdehyde (MDA) content of ham in the embodiments of the present invention.

[0042] Figure 10 This is a radar chart showing the sensory evaluation of ham processed in a staged modified atmosphere manner according to an embodiment of the present invention.

[0043] Figure 11This is a radar diagram showing the response of the electronic nose sensor for a staged modified atmosphere processing ham according to an embodiment of the present invention.

[0044] Figure 12 This is a principal component analysis (PCA) score graph of the electronic nose data of the staged modified atmosphere treatment ham in an embodiment of the present invention.

[0045] Figure 13 This is a bacterial community structure at the phylum level, as described in an embodiment of the present invention.

[0046] Figure 14 The variation of ham peptide content during staged modified atmosphere treatment is shown in an embodiment of the present invention.

[0047] Figure 15 This invention provides an analysis of the ability of staged modified atmosphere treatment to scavenge free radicals from crude peptides in ham.

[0048] Figure 16 This invention provides a staged modified atmosphere treatment process for ham with antioxidant activity IC. 50 Value analysis. Detailed Implementation

[0049] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0050] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods have been described herein, any methods similar or equivalent to those described herein may be used in the implementation or testing of this invention.

[0051] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be obvious to those skilled in the art. This specification and embodiments are merely exemplary.

[0052] The materials, reagents, and instruments used in this invention are as follows: 1. Experimental Materials 1.1 Raw Ham: The hind leg of Dugao Sa pig, processed using traditional Yunnan techniques, is dry-cured ham with a processing cycle of 12 months. Raw materials were collected from the same batch and fermentation workshop of the ham processing plant of Qujing Chuanfeng Food Co., Ltd., Yunnan Province, between June and October 2025. Ham samples with uniform size (weight 10.5±0.5kg, length 70±3cm), no mold, and no visible insect infestation were selected.

[0053] Insect source material: Stratiosperm larvae were collected from historically infected hams from the same factory. Healthy and active larvae (8-10mm in length) were selected for modified atmosphere insecticide verification experiments, with 20-50 test larvae per replicate.

[0054] 1.2 Main Reagents Methanol, acetonitrile (chromatographic grade, Thermo Fisher Scientific, USA); n-hexane, petroleum ether, anhydrous ethanol (analytical grade, Xilong Scientific Co., Ltd.); 2-methyl-3-heptanone (internal standard, purity ≥98%, Shanghai Aladdin Biochemical Technology Co., Ltd.); thiobarbituric acid (TBA), trichloroacetic acid (TCA), ethylenediaminetetraacetic acid (EDTA) (analytical grade, Sinopharm Chemical Reagent Co., Ltd.); DPPH (1,1-diphenyl-2-trinitrophenylhydrazine), ABTS (2,2'-azido-bis-3-ethylbenzothiazoline-6-sulfonic acid) (purity ≥98%, Sigma-Aldrich, USA); DNA extraction kit (Qiagen PowerSoil® DNA Isolation Kit, Qiagen GmbH, Germany); PCR amplification primers (synthesized by Shanghai Sangon Biotech Co., Ltd.).

[0055] 1.3 Main Instruments and Equipment Controlled atmosphere chamber (Guangyu Acrylic Glass Co., Ltd.); Gas chromatography-mass spectrometry (GC-MS, TQ8040NX, Shimadzu Corporation, Japan); Liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QTOF-MS, 1290Infinity II-6545, Agilent Technologies, USA); Electronic nose (PEN3, Airsense, Germany); High-throughput sequencing platform (NovaSeq 6000, Illumina, USA); Texture analyzer (TA.XT Plus, Stable Micro Systems, UK); Colorimeter (CR-400, Konica Minolta, Japan); Water activity meter (LabSwift-Aw, Novasina, Switzerland).

[0056] 2. Experimental Methods 2.1 Stage-based modified atmosphere treatment experimental method Experimental group design: A randomized block design was adopted, dividing the ham into two treatment groups (high concentration group H and low concentration group L), with three replicates in each treatment group. A natural fermentation control group B was set up, which matured naturally in the same fermentation workshop without controlled atmosphere treatment, and also had three replicates. Based on the investigation of the natural infection pattern of chrysalis flies, three key control points were identified: May-June (early stage), July-August (mid-stage), and September-October (late stage). Based on the preliminary factory survey and the design of staged controlled atmosphere parameters for the preliminary experiment, control group B, low concentration group L, and high concentration group H were set up.

[0057] Sampling method: Sampling sites were the surface of the ham (0-2 cm from the skin) and deep muscle (5-8 cm from the skin, avoiding the bone). Samples were immediately flash-frozen in liquid nitrogen after collection and stored in an ultra-low temperature freezer at -80°C for later use.

[0058] The specific parameters for the three stages of controlled atmosphere treatment are as follows: Stage 1 (Initial Stage): May 11-17, 2025 (7 days of treatment) and June 15-21, 2025 (7 days of treatment), temperature 25±1℃, relative humidity 75±5%, CO2 concentration 20% (low concentration group L), 35% (high concentration group H); Stage 2 (Mid-Stage Stage): July 13-19, 2025 (7 days of treatment) and August 10-16, 2025 (7 days of treatment), temperature 25±1℃, relative humidity 75±5%, CO2 concentration 35% (low concentration group L), 40% (high concentration group H). This stage coincides with the peak period of jackfly infestation, and the focus is on monitoring the insecticidal effect. Phase 3 (Later Stage): September 14-20, 2025 (7 days of treatment) and October 12-18, 2025 (7 days of treatment), with a temperature of 25±1℃, relative humidity of 75±5%, and CO2 concentration of 20% (low concentration group L) and 35% (high concentration group H).

[0059] Controlled Atmosphere (CAT) Operation Procedure: 1. Pretreatment: Place the hams on a stainless steel mesh rack inside the CAT chamber, spacing each ham 15cm apart to ensure gas flow. 2. Sealing and Inflation: Close the CAT chamber door and check the seal (pressure drop < 5% per 24h). Inflate CO2 according to the settings, monitoring in real time using a CO2 concentration detector. 3. Parameter Monitoring: Record temperature, humidity, and CO2 concentration every 2 hours, ensuring fluctuations are within ±5% of the set values. 4. Recovery Phase: After CAT treatment, slowly release CO2 (pressure drop rate < 0.01MPa / min) to avoid decompression damage. Immediately after treatment, switch to natural ventilation in the mesh shed and continue natural maturation in a conventional fermentation workshop.

[0060] Insecticidal validation experiment for botflies: During the modified atmosphere treatment, the survival status of larvae was observed and recorded every 24 hours. After the treatment, observation continued for 10 days, and the pupation rate and emergence rate were recorded. The emergence inhibition rate (IRE) was calculated.

[0061] ; In the formula, N control N represents the number of feathered cells in the control group. treatment To handle the number of feathered groups.

[0062] 3. Results and Analysis 3.1 Feathering Inhibition Rate Based on the preliminary experimental results, the inhibitory effects of different CO2 concentrations on the emergence of tyrosinus flies at various stages were as follows: Figure 2 As shown, the significance markers are: ( The results showed that controlled atmosphere treatment significantly inhibited zygote emergence. The control group (natural air) showed an emergence inhibition rate of less than 5.0% in each month, indicating that zygotes can complete development and emerge normally under natural conditions, with an emergence rate maintained between 89.5% and 94.2%, consistent with normal developmental levels. This result confirms the vitality and developmental capacity of the zygote population under the experimental conditions, providing a reliable benchmark for subsequent evaluation of the effects of controlled atmosphere treatment.

[0063] The low-concentration group (20%-35% CO2) showed a significant feathering inhibition effect in all treatment months. In May, the feathering inhibition rate was 88.2±3.5%, and the inhibition effect gradually increased with the progress of the treatment months.

[0064] The initial treatment in June (stage 1 of ham processing) achieved a feather emergence inhibition rate of 85.4±4.2%, which was significantly different from the control group (p<0.05). The feather emergence inhibition rates for the July and August treatments were 90.8±3.8% and 92.5±4.5%, respectively, indicating that controlled atmosphere packaging (CAP) maintained good control effects during the high-temperature summer period. The feather emergence inhibition rates for the September and October treatments further increased to 93.3±3.9% and 93.6±3.2%, respectively, which may be related to the suitable ambient temperature in autumn and the effectiveness of CAP. The annual average feather emergence inhibition rate for the low-concentration group was 92.5±3.9%.

[0065] The high-concentration group (35%-40% CO2) exhibited superior emergence inhibition. In May, the emergence inhibition rate reached 97.5±2.8%, significantly higher than the low-concentration group at the same time. In early June, the emergence inhibition rate was 98.1±2.2%, at which point the ham was in the early fermentation stage, and chrysalis fly activity began to increase; high-concentration controlled atmosphere treatment effectively blocked pest establishment. In July, the emergence inhibition rate reached a peak of 99.6±2.5%. This period coincided with the rainy season in Yunnan, and the ambient temperature (25±1℃) and relative humidity (75±5%) were conducive to the full effect of controlled atmosphere treatment. In mid-August, the emergence inhibition rate was 99.4±3.3%. At this time, the ham entered the mid-fermentation stage, protein and fat decomposition was active, odor emission was enhanced, and the risk of chrysalis fly attraction increased; high-concentration controlled atmosphere treatment provided an effective protective barrier. The feathering inhibition rates in September and October were 99.2±2.7% and 99.8±2.1%, respectively, indicating that a CO2 concentration of 35%-40% can achieve near-complete feathering inhibition in the later stages of ham processing. The high-concentration group had an average feathering inhibition rate of 99.6±2.8% throughout the year, which was significantly better than the low-concentration group.

[0066] 3.2 Cumulative feathering inhibition rate To verify the sustainability of pest control effects after controlled atmosphere treatment, this study monitored the cumulative emergence inhibition rate during the 0-10 day period of static storage after treatment. The results are as follows: Figure 3 As shown in the figure, the cumulative emergence inhibition rate of the control group remained at a low level (0-3.5%) during the 10-day static period, and the change over time was not significant, indicating that botflies can emerge normally under natural conditions, and the storage environment has no significant inhibitory effect on botfly development. After 7 days of treatment, both the low-concentration and high-concentration groups achieved stable control effects after only 1 day of static storage, with the cumulative emergence inhibition rate stabilizing at 100%. The 10-day no-emergence verification results showed that the high-concentration group had a cumulative emergence inhibition rate of 100% on the 10th day, indicating that the modified atmosphere treatment did not exhibit a "delayed emergence" phenomenon, and the treatment effect was definite and thorough. This result is of great significance for practical production applications, allowing ham processing plants to maintain a pest-free state for 7-10 days after the modified atmosphere treatment, providing a safety guarantee for subsequent processing steps. By combining the two indicators of emergence inhibition rate and cumulative emergence inhibition rate, a staged controlled atmosphere strategy of 35%-40% CO2 concentration, 7 days of treatment, and 10 days of static storage in the storage room can achieve an inhibition effect of more than 100% on tyrosinus emergence, and the effect is stable and long-lasting, making it feasible for promotion and application in actual production.

[0067] 3.3 Effects of modified atmosphere treatment on the physicochemical quality of ham Measurement indicators and methods: (1) Moisture content: Refer to GB 5009.3-2016 "National Food Safety Standard - Determination of Moisture in Food" - Method 1 (Direct Drying Method); (2) Water activity (Aw): Measured using a water activity meter, and read after equilibration at 25℃ for 30 min; (3) pH value: According to GB 5009.237-2016, weigh 10g of sample, add 100mL of deionized water, homogenize for 2min, let stand for 10min, and then filter and measure. (4) Crude protein: Calculated according to GB 5009.5-2025, Method III (combustion method), with a nitrogen-protein conversion factor of 6.25; (5) Crude fat: Refer to GB 5009.6-2016 Method II (Soxhlet extraction); (6) MDA content: Refer to GB 5009.181-2016 (spectrophotometric method), and the results are expressed as mg MDA / kg sample.

[0068] 3.3.1 Changes in water activity of ham Water activity (Aw) is a key parameter affecting the microbial stability and quality formation of dry-cured ham. During the dry-curing process, the gradual decrease in water activity is closely related to muscle tissue dehydration and shrinkage, salt penetration, and the drying process. For example... Figure 4As shown, the water activity of all three ham groups decreased with increasing fermentation time, a typical characteristic of the ham dehydration and maturation process. The water activity of all three groups exhibited an inverted V-shaped trend, first increasing and then decreasing. During the early fermentation period from May to June, the water activity rose from approximately 0.875 to its peak (0.885 for group H, and approximately 0.883 for groups B and L). During this stage, the free water content in the substrate increased, mainly due to the release of moisture from the raw materials themselves and the initial metabolic water production by microorganisms. After June, the fermentation entered the middle and late stages, and the water activity continued to decrease. By October, group H had dropped to 0.838, while groups B and L maintained at 0.847 and 0.850, respectively, indicating that the fermentation system gradually became drier and more stable. The high-concentration controlled atmosphere group (H) showed a more significant decreasing trend in water activity in the later stages (August-October). In September and October, the water activity of group H was significantly lower than the control group, decreasing by 1.19% and 1.06% compared to group B, respectively. This phenomenon may be attributed to the high concentration of CO2 inhibiting the metabolic activity of microorganisms and reducing metabolic water production. Simultaneously, the modified atmosphere packaging (MAP) environment may have accelerated the migration and loss of moisture from the substrate surface. CO2 MAP inhibits microbial metabolism by creating an acidic environment, affecting the moisture distribution of meat products; the CO2 environment has a reducing effect on water activity. Notably, there was no significant difference between the low concentration group (L) and the control group (B) throughout the fermentation process, suggesting that the MAP concentration threshold has a dose-dependent effect on water activity regulation. The optimal water activity range for tyrosinfly egg hatching and larval growth is 0.90-0.95. In this invention, although the water activities of all three groups were below this threshold, group H further reduced Aw to below 0.85 in the later stages, closer to the critical lower limit for tyrosinfly survival (Aw≈0.80), thus effectively suppressing the risk of pest outbreaks.

[0069] 3.3.2 Changes in the moisture content of ham like Figure 5As shown, the moisture content of the three groups of samples exhibited a three-stage change characteristic of "stable-sudden decrease-slow decrease". From May to July, during the initial fermentation period, the moisture content remained at a high level (46.0%-46.8%), with no significant difference between groups (p>0.05), indicating that the controlled atmosphere treatment had not yet significantly affected the total moisture content of the substrate during this stage. From July to August, a rapid dehydration period began, with the moisture content of the three groups decreasing sharply by approximately 4.5-5.0 percentage points, mainly attributed to increased water evaporation due to higher summer temperatures and water consumption by microbial metabolism. From August to October, a slow drying period occurred, with the moisture content continuing to decrease but at a slower rate. By October, group H had decreased to 37.5%, while groups B and L had 38.0% and 38.2%, respectively. The high-concentration controlled atmosphere group (H) showed a more significant dehydration-promoting effect in the middle and later stages of fermentation. In August, the moisture content of group H (42.8%) was already lower than that of the control group (41.8% vs 42.0%), and the difference widened further by October, with group H showing a 1.32% decrease compared to group B (relative decrease rate). This result corroborates the water activity data, indicating that a high-concentration CO2 environment may accelerate water loss in the fermentation system through mechanisms such as inhibiting microbial metabolic water production and altering the water vapor pressure gradient on the substrate surface. Notably, the moisture content of the low-concentration group (L) and the control group (B) highly overlapped throughout the monitoring period, with significant overlap in the error bars, indicating that the regulatory effect of low-concentration controlled atmosphere on total moisture content is limited. In the later stages of fermentation (September-October), the moisture content dropped below 40%, which is lower than the suitable moisture threshold (40%-50%) for jackfly larval development. Combined with a low water activity environment, this effectively interrupted the pest life cycle. Simultaneously, moderate dehydration helps inhibit the proliferation of spoilage microorganisms and extend the product shelf life. This invention demonstrates the significant advantages of high-concentration staged controlled atmosphere in promoting fermentation substrate dehydration and synergistic pest control.

[0070] 3.3.3 Changes in pH value of ham pH value is an important indicator reflecting the fermentation process and the degree of protein hydrolysis in dry-cured ham. For example... Figure 6As shown, the pH values ​​of the three ham groups exhibited a trend of first decreasing and then increasing, which is a typical characteristic of dry-cured ham fermentation. During protein hydrolysis, the synergistic action of endogenous enzymes and microbial proteases leads to the decomposition of polypeptides into small peptides, free amino acids, and basic amines, causing changes in physicochemical indicators. From May to July, the fermentation acidification reversal period occurred, with the pH value significantly increasing from the initial 6.25 to 6.48, mainly attributed to the deamination of proteins under the action of microbial proteases and the production of basic nitrogen-containing compounds from amino acid degradation. Protein degradation releases low-molecular-weight nitrogen molecules and ammonia, resulting in a significant increase in pH. From July to October, the pH stabilized, with the pH values ​​of all three groups remaining in the weakly acidic to near-neutral range of 6.48-6.50, and the differences between groups gradually narrowed and tended to be consistent. The high-concentration controlled atmosphere group (H) showed a significant lag in pH increase during the early stage of fermentation. In May, the pH value of group H (6.28) was significantly higher than that of the control group (6.19), but the rate of increase from June to July was significantly lower than that of groups B and L. In July, the pH value of group H (6.38) was significantly lower than that of groups B (6.47) and L (6.46) (p < 0.05), with a difference of 0.09 pH units. This result indicates that a high concentration of CO2 effectively inhibits the deammoniation metabolism of microorganisms and the accumulation of alkaline substances, thus delaying the alkalization process of the fermentation system. In modified atmosphere packaging, CO2 dissolves in the aqueous phase of the meat product and promotes the metabolism of lactic acid bacteria to produce lactic acid, leading to a decrease in pH. After August, the pH values ​​of the three groups tended to be consistent, suggesting that the pH regulation effect of modified atmosphere treatment is phased. As fermentation enters the later stage, the microbial community structure tends to stabilize, and the modified atmosphere effect gradually weakens. The optimal pH range for the digestive system of tyrosinfly larvae is 6.5-7.5. The high-concentration controlled atmosphere group maintained a relatively low pH (6.30-6.38) during the critical fermentation period (June-July), reducing the larvae's survival adaptability by inhibiting intestinal protease activity and interfering with nutrient absorption. These results provide a biochemical and environmental theoretical basis for the application of staged controlled atmosphere treatment during the pest-sensitive period (mid-fermentation).

[0071] 3.3.4 Changes in the crude fat content of ham Crude fat content is an important indicator for evaluating the nutritional value and flavor characteristics of dry-cured ham. For example... Figure 7As shown, the crude fat content remained within a narrow range of 5.44%-5.64% with minimal fluctuation (coefficient of variation <2%), indicating that the modified atmosphere packaging and fermentation process had a good retention effect on the total lipids in the matrix. The fatty acid content of the fermented ham exhibited a wave-like change during processing, significantly increasing after maturation. The control group (B) showed a fluctuating trend of "decline-increase-decline-increase," slightly decreasing from May to June (5.46%→5.44%), rising to 5.49% in July, decreasing again to 5.44% in August, and slightly increasing to 5.47% in October, generally stabilizing. The low-concentration group (L) showed a similar trend to the control group, but with a more gradual fluctuation, remaining within the 5.46%-5.53% range from May to October. The high-concentration modified atmosphere packaging group (H) exhibited a unique "first decrease, then increase" pattern. From May to August, the crude fat content of group H was not significantly different from that of groups B and L (p>0.05), and was even slightly lower than that of the control group in June and August, indicating that the high concentration of CO2 did not significantly inhibit lipid metabolism during the mid-fermentation period. Notably, the crude fat content of group H showed a significant upward trend from September to October, reaching 5.64% in October, an increase of 3.11% compared to the control group (5.47%) and 1.99% compared to group L (5.53%). Error bars showed that the difference between groups was statistically significant (p<0.05). This late-stage enrichment phenomenon may be due to the high concentration of modified atmosphere packaging inhibiting lipoxygenase activity and free radical chain reactions, reducing lipid oxidation losses. Simultaneously, the modified atmosphere environment may have inhibited the proliferation of lipophilic microorganisms, reducing lipid microbial degradation. The presence of CO2 in modified atmosphere packaging can slow down the lipid oxidation rate of meat products. Lipids are a key energy source required for the development of tyrosinfly larvae, and their content is positively correlated with larval survival rate. Halving the lipid content in feed leads to a decrease in adult emergence rate, confirming the significant impact of lipids on insect development and reproductive performance. This result indicates that staged controlled atmosphere packaging can achieve a balance between lipid protection and pest control.

[0072] 3.3.5 Changes in crude protein content of ham Crude protein content is a core indicator for evaluating the nutritional value and maturity of cured ham. For example... Figure 8As shown, the protein content exhibited a non-linear change characteristic of "continuous increase - sharp drop at peak". From May to September, the protein accumulation period occurred, with the content continuously climbing from 31.5%-32.5% to 35.5%-36.2%, an increase of 12.7%-13.2%, mainly due to microbial metabolism and the relative concentration effect of substrate dry matter. Protein degradation is the core biochemical process during the dry-cured ham processing; the synergistic action of microbial proteases and endogenous enzymes leads to the degradation of structural proteins, forming small peptides and free amino acids. After reaching its peak in September, the protein content of all three groups showed a significant decline in October, with group H dropping to 32.2%, and groups B and L at 35.0% and 32.5% respectively, a decrease of 10.9%-11.0%, possibly related to the enhanced activity of microbial proteases and the deep degradation of proteins in the later stages. During the maturation process of dry-cured ham, non-protein nitrogen (NPN) continuously increased, and protein degradation was significantly affected by temperature and salt concentration, with high-temperature and low-salt conditions exacerbating protein degradation. The low-concentration controlled atmosphere group (L) showed a significant advantage in protein content during the mid-fermentation stage. From May to August, the protein content of group L was consistently higher than that of groups B and H, reaching 35.5% in August, which was 6.3% higher than the control group (33.4%) and 2.9% higher than group H (34.5%). This indicates that moderate controlled atmosphere concentration may promote protein synthesis or inhibit excessive degradation during the mid-fermentation stage. The protein content of the high-concentration controlled atmosphere group (H) was slightly lower than that of group L from May to August, but quickly caught up to 36.2% in September, matching that of group B (35.9%) and slightly higher than group L (35.2%), showing that the regulation of protein accumulation by high-concentration controlled atmosphere has a lag effect. Notably, in October, the protein content of groups H and L decreased to around 32%, while the control group remained at 35.0%, indicating that controlled atmosphere treatment may have accelerated protein degradation and transformation in the later stages. Protein is an essential nutrient for the development of tyrosinfly larvae, and its content is positively correlated with the risk of pest outbreaks. The high protein accumulation in the low-concentration controlled atmosphere (CPA) group during the mid-term raises concerns about insect pest risks. However, considering the water activity and pH data of this group, the CPA environment can establish a balance between nutrient supply and physiological stress by reducing Aw (air activity) and delaying alkalization. Protein is primarily used for tissue growth, while carbohydrates and lipids are used for energy supply, suggesting an optimal threshold for insect protein requirements. The rapid protein degradation in the high-concentration CPA group during the later stages may help reduce attractability to tyrosinus flies, but its impact on the final nutritional quality of the product needs to be monitored. These results provide a basis for optimizing staged CPA parameters (moderate CPA in the mid-term and enhanced CPA in the late stage) at the protein metabolism level.

[0073] 3.3.6 Changes in malondialdehyde (MDA) content in ham Malondialdehyde (MDA) is a major product of lipid oxidation, and its content reflects the degree of oxidation and quality stability of cured ham. For example... Figure 9As shown, MDA content exhibited significant inter-group differentiation. The control group (B) showed a continuous and sharp upward trend, increasing from 0.85 mg / kg in May to 1.82 mg / kg in October, an increase of 114.1%, with a monthly average growth rate of 0.19 mg / kg, indicating that lipid oxidation under conventional fermentation conditions intensifies over time. The low-concentration controlled atmosphere group (L) and the high-concentration controlled atmosphere group (H) showed excellent oxidation inhibition effects, with MDA content in both groups remaining at extremely low levels of 0.70-0.80 mg / kg throughout the monitoring period, with nearly flat curves and no significant time-dependent increase.

[0074] MDA, as a major product of lipid oxidation, has cytotoxicity and mutagenicity, and can significantly reduce the digestibility of myofibrillar proteins, which is consistent with the conclusion in this invention that the accumulation of MDA in the control group affects food safety. There was no significant difference in MDA content between the high-concentration modified atmosphere group (H) and the low-concentration modified atmosphere group (L) (p>0.05), but both were significantly lower than the control group. In July, the MDA inhibition rate of the H group and the L group reached 42.4% and 41.2%, respectively, and by April the inhibition rate had further increased to 57.1% and 56.0%. This strong antioxidant effect may be due to the following mechanisms: (1) The high concentration of CO2 environment significantly reduced the partial pressure of O2 in the system, directly inhibiting the initiation and propagation of free radical chain reactions; (2) Modified atmosphere treatment inhibited the activity of lipoxygenase (LOX) and lipoxygenase, blocking the enzymatic oxidation pathway; (3) The low oxygen environment reduced the amount of transition metal ions (Fe2+). 2＋ Cu ＋The catalytic oxidation effect of CO2 modified atmosphere packaging. CO2 modified atmosphere packaging exhibits strong inhibitory ability against lipid oxidation and rancidity under storage conditions of 25℃, effectively delaying initial and secondary lipid oxidation. In this invention, both low and high concentrations of modified atmosphere packaging achieve similar antioxidant effects at a "plateau." Reducing O2 concentration significantly inhibits lipid oxidation, but high concentrations of CO2 promote oxidation by reducing the activity of superoxide dismutase and glutathione peroxidase, indicating the existence of an optimal CO2 concentration range. Notably, even low concentrations of modified atmosphere packaging achieve antioxidant effects similar to high concentrations, suggesting the existence of an "plateau" in modified atmosphere concentration for lipid oxidation control. MDA, as a secondary oxidation product, is cytotoxic and mutagenic. Its accumulation not only affects product flavor (producing rancidity) but also threatens food safety. As a byproduct of lipid oxidation, MDA can significantly reduce the oxidative stability of meat myofibrillar proteins by promoting protein carbonylation and cross-linking reactions, further demonstrating the negative impact of MDA accumulation on meat quality. In this invention, the MDA content in the control group (1.82 mg / kg) after 10 months was close to the critical threshold (2.0 mg / kg) for oxidative deterioration of fermented foods, while the modified atmosphere treatment group consistently maintained below the safe level. This result strongly demonstrates the significant advantages of "staged modified atmosphere" technology in delaying lipid oxidation and ensuring the oxidative stability of fermented foods, providing an important technical means for extending product shelf life.

[0075] In summary, the water activity of all treatment groups decreased with increasing fermentation time, eventually reaching 0.838 in group H, 0.847 in group B, and 0.850 in group L. The MDA content in the high-concentration modified atmosphere groups was 40.5%-57.1% lower than that in the control group, effectively inhibiting lipid oxidation. The pH value of each group remained in the slightly acidic to near-neutral range of 6.28-6.50, the crude protein content steadily increased from 31.5% to 36.2%, and the crude fat content remained within a relatively narrow range of 5.44%-5.64%.

[0076] 4. The impact of modified atmosphere packaging on the flavor and quality of ham Measurement method: (1) Sensory evaluation: A sensory analysis team consisting of 10 trained evaluators (5 men and 5 women, aged 25-40) was formed. Quantitative descriptive analysis (QDA) was used. The evaluation indicators included color, aroma, taste, texture and overall acceptability. A 10cm linear scale (0=none, 10=very strong) was used. (2) Electronic nose analysis: PEN3 type electronic nose is used, equipped with 10 metal oxide sensors. 5g of sample is placed in 20mL headspace vial, equilibrated at 40℃ for 20min, headspace volume is 10mL, and detection time is 120s. (3) Volatile flavor compounds (GC-MS): Headspace solid phase microextraction (HS-SPME) was used with HP-5MSUI column (30m×0.25mm×0.25μm). The qualitative analysis was performed by comparison with the NIST2020 standard library (matching degree >80%), and the semi-quantitative analysis was performed by internal standard method.

[0077] Sensory evaluation results such as Figure 10 The results showed that in October, the overall acceptability of both the high-concentration and low-concentration modified atmosphere groups reached 8.5 points, an improvement of 6.25% compared to the control group.

[0078] Ham Electronic Nose Analysis Results Figure 11 and Figure 12 As shown, GC-MS screening revealed 128 volatile compounds with an SI value greater than 90. Aldehydes were the main flavor contributors, accounting for 38.5%-55.2%. OAV analysis of key aroma active substances showed that 2,4-decadienal (E,E) (oily aroma), nonanal (citrus aroma), and 1-octen-3-ol (mushroom aroma) were the three substances with the highest OAV values. The OAV values ​​of the main aroma substances in the modified atmosphere treatment group remained at a high level.

[0079] 5. Effects of modified atmosphere treatment on the microbial community structure of ham Methods: Muscle samples were collected from the surface (0-2 cm) and deep (5-8 cm) layers of ham. High-throughput sequencing (16S rRNA gene V3-V4 region sequencing for bacteria, ITS1 region sequencing for fungi) was used to analyze the microbial community structure. β-diversity analysis was performed using QIIME v1.9.1, LEfSe analysis (LDA > 4) was used to screen for differentially expressed biomarkers between groups, and PICRUSt2 was used for functional gene pathway prediction.

[0080] The results are as follows Figure 13 As shown, the dominant bacterial phyla were Firmicutes, Proteobacteria, Bacteroides, and Actinobacteriota, which together accounted for over 85% of the total community relative abundance. With increasing CO2 concentration, the relative abundance of Firmicutes showed a significant upward trend, reaching 65.2% in the high CO2 treatment group. High CO2 treatment significantly enriched fermentation functional bacteria such as Tetragenococcus and Cobetia, while inhibiting potential putrefactive bacteria such as Pseudomonas and Acinetobacter.

[0081] 6. Effects of modified atmosphere treatment on the antioxidant activity of ham Assay methods: Ham protein hydrolysate was prepared by in vitro simulated gastrointestinal digestion, and peptide content was determined by OPA colorimetric method. ABTS free radical scavenging capacity, DPPH free radical scavenging capacity, and hydroxyl free radical scavenging rate were determined by spectrophotometry, with VC as a positive control, and IC50 was calculated. 50 Value (half-maximal inhibitory concentration).

[0082] The results are as follows Figure 14 , Figure 15 (Significance markers: ( ), Figure 16 (Significance markers: ( As shown in the figure, the antioxidant activity of the high-concentration modified atmosphere group was 15%-25% higher than that of the control group. 50 Value analysis shows that ABTS free radical scavenging IC 50 In October, the CO2 level in the high CO2 group decreased to 1.48 ± 0.04 mg / mL, and the DPPH free radical scavenging IC50 value was [value missing]. 50 The value decreased to 1.55±0.03 mg / mL, which was significantly lower than that of the control group.

[0083] 7. Conclusion This invention provides a staged controlled atmosphere technology method for controlling stratus fly pests, with the following main beneficial effects: (1) The staged modified atmosphere treatment showed a significant inhibitory effect on all stages of botfly. The high concentration group (35%-40% CO2) inhibited the emergence of botfly pupae by 99.6%, and the low concentration group (20%-35% CO2) also achieved an inhibition effect of 97.8%, indicating that the technology has excellent pest control capabilities.

[0084] (2) Modified atmosphere treatment has little impact on the physicochemical quality of ham. The key physicochemical indicators such as water activity, pH value, crude protein content, and crude fat content of each treatment group were not significantly different from those of the control group, indicating that the staged modified atmosphere technology can maintain the normal physicochemical properties of ham while effectively controlling pests.

[0085] (3) The effect of modified atmosphere treatment on the overall flavor profile of ham is controllable. A total of 128 volatile flavor compounds were identified by GC-MS. The OAV values ​​of key aroma active substances remained at a high level, and the modified atmosphere treatment group still maintained the typical flavor characteristics of Yunnan dry-cured ham.

[0086] (4) High-concentration modified atmosphere treatment significantly enhanced the antioxidant activity of ham. The DPPH, ABTS free radical scavenging capacity and hydroxyl free radical scavenging capacity were increased by 15-25% compared with the control group, giving the product higher nutritional and health value.

[0087] This invention successfully established a staged controlled atmosphere pest control technology system suitable for dry-cured ham, achieving multiple goals of "killing pests without harming the fermentation process, preserving quality and increasing efficiency".

[0088] The above description is merely a specific embodiment of this application, but the scope of protection of this application is not limited thereto. Any changes or substitutions that can be easily conceived by those skilled in the art within the scope of the technology disclosed in this application should be included within the scope of protection of this application.

Claims

1. A method for staged controlled atmosphere packaging to control cheese fly pests in traditional dry-cured ham, characterized in that, Based on the fermentation stage of the ham and / or the life history of the cheese fly, multiple independent modified atmosphere processing cycles are performed at multiple predetermined time points. In this process, at least one treatment cycle uses a first CO2 concentration range, and at least one treatment cycle uses a second CO2 concentration range that is higher than the first range.

2. The method for controlling cheese fly in traditional dry-cured ham using staged modified atmosphere packaging according to claim 1, characterized in that, The first CO2 concentration range is 20%-35%.

3. The method for staged controlled atmosphere packaging for controlling cheese fly pests in traditional dry-cured ham according to claim 1, characterized in that, The second CO2 concentration range is 35%-40%.

4. The method for staged controlled atmosphere packaging for controlling cheese fly pests in traditional dry-cured ham according to claim 1, characterized in that, The modified atmosphere processing cycle includes: Store the ham in a sealed environment; Introduce CO2 or a mixture of CO2 and N2 to bring the CO2 concentration in the environment to a predetermined target value; Maintain at a predetermined temperature and relative humidity for a predetermined time.

5. The method for staged controlled atmosphere packaging for controlling cheese fly pests in traditional dry-cured ham according to claim 4, characterized in that, During the middle stage of ham fermentation, the second CO2 concentration range is used, while the first CO2 concentration range is used during the early and late stages of fermentation.

6. The method for staged controlled atmosphere packaging for controlling cheese fly pests in traditional dry-cured ham according to claim 4, characterized in that, The conditions for the controlled atmosphere treatment are: a predetermined temperature of 25±1℃, a relative humidity of 75±5%, and a treatment time of 5-10 days.

7. The method for staged controlled atmosphere packaging for controlling cheese fly pests in traditional dry-cured ham according to claim 4, characterized in that, In the modified atmosphere treatment, the O2 concentration of the mixed gas is less than 1%.

8. The method for staged controlled atmosphere packaging for controlling cheese fly pests in traditional dry-cured ham according to claim 6, characterized in that, After the controlled atmosphere treatment is completed, there is also a 0-10 day in-store settling period.

9. The method for staged controlled atmosphere packaging for controlling cheese fly pests in traditional dry-cured ham according to claim 8, characterized in that, After the controlled atmosphere treatment is completed, the process of restoring the natural air environment includes the slow release of CO2 at a depressurization rate of less than 0.01 MPa / min.

10. A controlled atmosphere fermentation system for dry-cured ham for implementing the method of any one of claims 1-9, characterized in that, include: At least one sealed controlled atmosphere unit; Gas supply and distribution unit; Environmental control and monitoring unit; And a control unit configured to perform the staged modified atmosphere processing procedure.

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