Preservation method of rape

Abstract

The invention belongs to the technical field of vegetable preservation, and particularly relates to a preservation method of rape. The invention provides a preservation method for harvested oilseed rape by a photosensitizer mediated photodynamic technology. The photosensitizer solution concentration, the illumination intensity and the illumination time with the best preservation effect are screened out through experiments, and the preservation effect of the photosensitizer mediated photodynamic technology on the harvested oilseed rape is achieved. Meanwhile, experiments prove that the antioxidant enzyme activity of the oilseed rape treated by the photosensitizer-mediated photodynamic technology provided by the invention can be remarkably improved, the decline speed of vitamin C and chlorophyll content can be effectively slowed down, the aging process of the harvested oilseed rape can be remarkably slowed down, the storage life of the harvested oilseed rape can be prolonged, and the oilseed rape yield can be improved. And a theoretical basis and technical support are provided for transportation and preservation of harvested oilseed rapes.

Description

Technical Field

[0001] This invention belongs to the field of vegetable preservation technology, specifically relating to a method for preserving rapeseed. Background Technology

[0002] rape( Brassica rapa var. parachinensis Rapeseed (Brassica oleracea var. chinensis), also known as Chinese cabbage or bok choy, belongs to the Brassica genus of the Brassicaceae family, subspecies of Brassica oleracea. It is rich in vitamin C, vitamin K, and beta-carotene, as well as minerals such as calcium and potassium. Its dietary fiber, glucosinolates, and flavonoids can enhance immunity and protect bone and cardiovascular health. Rapeseed is highly valued for its rich nutrition, high quality and yield, short cultivation cycle, and strong adaptability. However, rapeseed has a large leaf surface area, high water content, and tender tissue, making it prone to yellowing, wilting, and mold and rot after harvesting, leading to significant post-harvest losses. Therefore, effectively extending the storage and preservation period of rapeseed after harvesting and reducing its rot is a crucial issue that urgently needs to be addressed.

[0003] Currently, the main methods for preserving agricultural products in my country include physical preservation, chemical preservation, biological preservation, and intelligent and composite preservation. Patent document CN102934684A discloses a modified atmosphere storage method for marine-grade Shanghai bok choy. This modified atmosphere storage method proceeds in the following steps: 1) pre-cooling; 2) preservation in a preservation bag; 3) preparing gas parameters into the preservation bag: using a high-precision gas mixer, O2, CO2, and N2 gases are introduced into the preservation bag containing the Shanghai bok choy, with the following parameters: O2 concentration of 2%~15%, CO2 concentration of 8%~15%, and the remaining gas being N2; 4) cold storage: controlling the cold storage temperature at 0℃~5℃ and the relative humidity at 60%~90%. This method, through pre-cooling, packaging in preservation bags, and gas preparation within the bags, along with strict control of key processes such as temperature and humidity, extends the storage period of Shanghai bok choy to 45 days. Over 95% of the bok choy shows no signs of deterioration such as leaf dehydration or rotting, maintaining good edibility. This method is simple, low-cost, leaves no chemical residues during storage, and meets international standards.

[0004] Patent document CN107361129A discloses a modified atmosphere packaging method for preserving rapeseed. This method includes the following steps: selection, washing, draining, disinfection, bagging, vacuuming, gas filling, sealing, and storage. The gas filling step involves using a high-precision gas distributor to introduce O2, CO2, and N2 gases into the bags containing the rapeseed, with the following parameters: O2 concentration less than 2%, CO2 concentration greater than 20%, and the remainder being N2. This method solves the key technical problem of rapeseed's short shelf life and susceptibility to spoilage and yellowing, extending the shelf life from the original 3 days at room temperature to over 25 days, making it highly practical and scalable.

[0005] However, current domestic and international methods primarily employ physical preservation techniques such as low-temperature treatment, modified atmosphere packaging, and ozone treatment for vegetables. These methods aim to slow down vegetable respiration and inhibit microbial growth to achieve storage and preservation. However, physical preservation suffers from high equipment costs, and the preservation effect of a single physical preservation technology is limited. Photodynamic inactivation (PDI) is a novel non-thermal sterilization technology that uses light of a specific wavelength to excite photosensitizers (PSs) to generate reactive oxygen species, thereby inactivating microbial cells. Compared to thermal sterilization and other cold sterilization methods, this technology has multiple advantages: broad-spectrum bactericidal activity, effectively killing most Gram-positive and Gram-negative bacteria without easily inducing bacterial resistance; rapid sterilization speed, capable of inactivating a large number of bacteria in a short time; low energy consumption and environmental friendliness; furthermore, by screening and modifying photosensitizers and combining them with nanosphere technology, targeted sterilization can be achieved.

[0006] The unique molecular structure of natural phenolic acids possesses the potential to act as photosensitizers, while also exhibiting excellent antioxidant properties, capable of directly scavenging free radicals and delaying plant tissue senescence. Therefore, combining natural phenolic acids with photodynamic therapy (PDI) to construct a naturally mediated photodynamic preservation system and applying it to postharvest rapeseed storage to extend its shelf life is of significant research value. Summary of the Invention

[0007] To address the shortcomings of existing technologies, the present invention aims to provide a photosensitizer-mediated photodynamic therapy for postharvest rapeseed (… Brassica rapa var. parachinensis This invention describes a preservation method for rapeseed. It screens out the optimal photosensitizer solution concentration, light intensity, and light exposure time for best preservation effect, aiming to maximize the preservation effect of photosensitizer-mediated photodynamic technology on post-harvest rapeseed, ultimately providing a new technical means for the transportation and preservation of post-harvest rapeseed.

[0008] To achieve the above objectives, the present invention provides the following technical solution: This invention provides a method for preserving rapeseed, comprising the following steps: Step S1: Select rapeseed that is uniform in size, undamaged, fresh, and intact. Brassica rapa var. parachinensis After washing, air dry to get clean rapeseed; Step S2: Dissolve the photosensitizer in distilled water and stir until homogeneous to obtain the treatment solution; Step S3: Completely immerse the clean rapeseed treated in step S1 in the treatment solution prepared in step S2, irradiate it under blue light for 5-15 minutes, package it, and store it in the refrigerator.

[0009] Furthermore, the photosensitizer in step S2 is protocatechuic acid or caffeic acid.

[0010] Furthermore, when the photosensitizer is protocatechuic acid, the concentration of the protocatechuic acid treatment solution is 0.05~0.1 mg / L.

[0011] Furthermore, when the photosensitizer is caffeic acid, the concentration of the caffeic acid treatment solution is 0.01~0.075 mg / L.

[0012] Furthermore, the blue light irradiation conditions in step S3 are: light intensity of 30~50 mW / cm². 2 The distance from the light source should be 18-25 cm, the irradiation time should be 5-15 min, and the surface should be flipped over every 2 min.

[0013] Furthermore, step S3 also includes a coating treatment step, specifically, after light treatment, the rapeseed is soaked in the coating solution for 4-6 minutes, drained, and packaged to obtain the final product.

[0014] Furthermore, the preparation method of the coating liquid is as follows: Chitosan was dissolved in a 1% (v / v) acetic acid solution to prepare a chitosan solution; photosensitizer was added to distilled water to prepare a photosensitizer solution; the chitosan solution was added to the photosensitizer solution and stirred until homogeneous to obtain the final product.

[0015] Furthermore, the chitosan solution contains a chitosan concentration of 1-2% w / v.

[0016] Further, the photosensitizer is protocatechuic acid or caffeic acid. When the photosensitizer is protocatechuic acid, the concentration of the protocatechuic acid solution is 0.05~0.1 mg / L. When the photosensitizer is caffeic acid, the concentration of the caffeic acid solution is 0.01~0.075 mg / L.

[0017] Furthermore, the volume ratio of the photosensitizer solution to the chitosan solution is 1:(2~3).

[0018] Natural phenolic acids are ideal photosensitizers. Protocatechuic acid (molecular formula: (HO)2C6H3COOH, molecular weight: 154.12, CAS number: 99-50-3) and caffeic acid (chemical formula: C9H8O4, molecular weight: 180.157, CAS number: 331-39-5) possess excellent optical properties, low toxicity, and good antibacterial activity. Using protocatechuic acid or caffeic acid as photosensitizers to mediate photodynamic reactions to kill pathogens can effectively extend the storage period of rapeseed.

[0019] This invention screened the optimal concentration of photosensitizer, light intensity, and light exposure time. Simultaneously, experiments confirmed that protocatechuic acid-mediated photodynamic therapy has a good preservation effect on post-harvest rapeseed. It can effectively inhibit the yellowing rate of rapeseed flowers, protect the rapeseed cell membrane, and effectively slow down the decline in vitamin C and chlorophyll content during storage, reducing the loss of vitamin C and chlorophyll and thus effectively slowing down the aging process of post-harvest rapeseed. Furthermore, caffeic acid-mediated photodynamic therapy also has a good preservation effect on post-harvest rapeseed. It can effectively improve the antioxidant effect of post-harvest rapeseed, maintain the integrity and stability of cell membranes, effectively slow down the decline in vitamin C and chlorophyll content during storage, reducing the loss of vitamin C and chlorophyll and thus effectively extending the storage time of post-harvest rapeseed.

[0020] In addition, the inventors used a coating solution made by mixing photosensitizer and chitosan to soak rapeseed. After draining, a semi-permeable membrane can be formed on the surface of the rapeseed, creating a low O2 and high CO2 environment inside the rapeseed. This inhibits the respiration and metabolism of the rapeseed, reduces the transformation of substances within the rapeseed, and reduces the consumption of respiratory substrates. At the same time, the semi-permeable membrane also has good water retention properties, which can reduce water loss from the rapeseed and block direct contact between pathogens and host tissues or cells, thus reducing pathogen invasion. The photosensitizer inside the protective membrane also has antioxidant and free radical scavenging effects, which can further extend the storage period of the rapeseed.

[0021] In summary, compared with the prior art, the rapeseed preservation method provided by this invention has the advantages of simple operation and strong practicality. Applying photosensitizer-mediated photodynamic technology to the preservation of post-harvest rapeseed can effectively extend the storage period of post-harvest rapeseed and provide a new technical means for the transportation and preservation of post-harvest rapeseed. Attached Figure Description

[0022] Figure 1 The figure shows the effect of protocatechuic acid-mediated PDI on the activities of SOD (a), CAT (b), and POD (c) in rapeseed.

[0023] Figure 2 The graph shows the effect of protocatechuic acid-mediated PDI on the vitamin C content in rapeseed.

[0024] Figure 3 The graph shows the effect of protocatechuic acid-mediated PDI on the MDA content in rapeseed.

[0025] Figure 4 The effect of protocatechuic acid-mediated PDI on chlorophyll content in rapeseed.

[0026] Figure 5 The figure shows the effect of caffeic acid-mediated PDI on SOD activity in rapeseed.

[0027] Figure 6 The figure shows the effect of caffeic acid-mediated PDI on CAT activity in rapeseed.

[0028] Figure 7 The figure shows the effect of caffeic acid-mediated PDI on POD activity in rapeseed.

[0029] Figure 8 The graph shows the effect of caffeic acid-mediated PDI on vitamin C content in rapeseed.

[0030] Figure 9 The graph shows the effect of caffeic acid-mediated PDI on the MDA content in rapeseed.

[0031] Figure 10 Figure showing the effect of caffeic acid-mediated PDI on chlorophyll content in rapeseed. Detailed Implementation

[0032] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0033] Example 1: Investigating the effect of protocatechuic acid-mediated photodynamic therapy on the preservation of postharvest rapeseed. 1. Experimental Samples: Select fresh, uniform-sized, undamaged, and intact rapeseed. Rinse with tap water and allow to air dry naturally with a fan and windows open. Select 6 uniform-sized, fresh, and intact rapeseed plants as one group, with 3 replicates per group. Protocatechuic acid (purity ≥97%) was purchased from Shanghai Yuanye Biotechnology Co., Ltd. Vitamin C assay kit, catalase (CAT) assay kit, peroxidase (POD) assay kit, and total superoxide dismutase (T-SOD) assay kit were purchased from Nanjing Jiancheng Bioengineering Institute. Plant malondialdehyde (MDA) assay kit was purchased from Shanghai Yuanye Biotechnology Co., Ltd.

[0034] 2. Experimental methods: 2.1 Optimal parameter screening for protocatechuic acid-mediated photodynamic therapy: (1) Design a single-factor experimental treatment for rapeseed: First, protocatechuic acid treatment solutions of 0 mg / L, 0.025 mg / L, 0.05 mg / L, 0.075 mg / L, 0.1 mg / L, and 0.125 mg / L were prepared. The washed rapeseed samples were then immersed in the protocatechuic acid treatment solutions of different concentrations and then stored in a refrigerator at 4 ℃ in the dark. The samples were observed on days 1, 3, 5, and 7.

[0035] Secondly, a light intensity of 30 mW / cm was selected. 2 The light source was used to irradiate the plants for 0, 5, 10, 15, 20, and 25 minutes, respectively. The plants were then stored in a refrigerator at 4 ℃ in the dark and observed on days 1, 3, 5, and 7. The plants were turned over every 2 minutes during irradiation.

[0036] Then, the illumination time was selected as 10 min, and the light intensity was set at 0, 10, 20, 30, 40, and 50 mW / cm². 2 The pretreated samples were irradiated with different light intensities, stored in a refrigerator at 4 ℃ in the dark, and observed on days 1, 3, 5, and 7, with the samples turned over every 2 minutes.

[0037] (2) Design a three-factor, three-level orthogonal experiment: The optimal protocatechuic acid concentrations were determined to be 0.05 mg / L, 0.075 mg / L, and 0.1 mg / L, and the optimal light intensities were 30, 40, and 50 mW / cm², respectively, based on single-factor experiments. 2 An orthogonal experiment with three factors and three levels was conducted with light exposure times of 5, 10, and 15 minutes.

[0038] (3) Verification experiment: Based on the orthogonal experimental results, the specific parameters are: protocatechuic acid concentration 0.075 mg / mL, under LED blue light (light intensity 50 mW / cm²). 2 Irradiation was performed for 5 minutes to verify the results. Subsequent experimental groups were as follows: Protocatechuic acid treatment group (L-P+): 0.075 mg / mL protocatechuic acid; Protocatechuic acid plus photodynamic therapy group (L+P-): 0.075 mg / mL protocatechuic acid, light intensity 50 mW / cm². 2 Irradiation for 5 minutes; and a blank control group (LP-).

[0039] 2.2 Determination of antioxidant enzyme activity in rapeseed: The activities of antioxidant enzymes (SOD, POD, CAT) were measured using a kit. The groups were divided into protocatechuic acid treatment group (L-P+), protocatechuic acid plus photodynamic treatment group (L+P-), and blank control group (LP-). The enzyme activities of these three groups were measured.

[0040] 2.3 Determination of Vitamin C Content in Rapeseed: The changes in vitamin C content in rapeseed were determined using a kit. The groups were divided into a protocatechuic acid treatment group (L-P+), a protocatechuic acid plus photodynamic therapy treatment group (L+P-), and a blank control group (LP-). The vitamin C content of these three groups was measured.

[0041] 2.4 Determination of lipid peroxidation in rapeseed cell membranes: The level of cell membrane lipid peroxidation was characterized by measuring the MDA content in rapeseed using a kit. The groups were divided into protocatechuic acid treatment group (L-P+), protocatechuic acid plus photodynamic therapy treatment group (L+P-), and blank control group (LP-). The MDA content of these three groups was measured.

[0042] 2.5 Determination of chlorophyll content in rapeseed: The determination of chlorophyll content in rapeseed was performed according to the "Experimental Methods of Food Preservation Technology". Rapeseed samples were placed in a mortar that had been pre-frozen overnight at low temperature and ground on ice. A small amount of quartz sand (0.3 g) and calcium carbonate powder (0.2 g) were added, followed by 3 mL of 95% ethanol. The mixture was quickly homogenized, and another 3 mL of 95% ethanol was added. The homogenate was then rapidly homogenized until the tissue turned white. The homogenate was quickly transferred three times with 6 mL of 95% ethanol into glass centrifuge tubes, then into 25 mL volumetric flasks. The flasks were brought to a final volume with 95% ethanol, the caps were tightened, and the mixture was shaken repeatedly. The mixture was then incubated in a biochemical incubator for 1 hour. After incubation, the mixture was transferred to 15 mL plastic centrifuge tubes and centrifuged at 10 ℃ and 8000 r / min for 15 min. Immediately after centrifuging, approximately 10 mL of the supernatant was carefully transferred to a clean centrifuge tube using a pipette next to the centrifuge (if plant tissue is accidentally transferred, the process of re-centrifuging and re-collecting must be repeated, as this step greatly affects the test results). Before the experiment, turn on the air conditioner in the UV chamber and maintain a temperature of 25 °C for at least 1 hour. Turn on the UV spectrophotometer and preheat it. Click on baseline calibration under no-load conditions, then select the spectral function, set the detection wavelength, and set the method. Add the reference solution (95% ethanol) to two cuvettes, then click zeroing. Remove the cuvettes from the sample cell and load them with samples one by one for testing; no further zeroing is required. Measure the absorbance at 663 nm and 645 nm, using 95% ethanol as a blank control. Store the test solutions away from light; measure one solution at a time before moving on to the next.

[0043] The chlorophyll concentration in the extract is: In the formula: cT -- chlorophyll concentration in the extract, mg / L W -- Fresh weight of the sample, in grams N -- Dilution factor.

[0044] 3. Results and Analysis: 3.1 Optimal parameter screening results for protocatechuic acid-mediated photodynamic therapy: The optimal preservation parameters for rapeseed using protocatechuic acid-mediated photodynamic therapy (PDPT) were determined. The primary and secondary factors influencing rapeseed storage were protocatechuic acid concentration > light intensity > light duration. The optimal preservation parameters for rapeseed using protocatechuic acid-mediated PPT were: rapeseed immersed in a protocatechuic acid treatment solution with a concentration of 0.075 mg / L, and irradiated with a light intensity of 50 mW / cm². 2 The illumination time was 5 minutes.

[0045] 3.2 Results of the determination of antioxidant enzyme activity in rapeseed: The results of the determination of antioxidant enzyme activity in rapeseed are as follows: Figure 1 As shown. Figure 1 The figure shows the effect of protocatechuic acid-mediated PDI on the activities of SOD(a), CAT(b), and POD(c) in rapeseed. Reactive oxygen species (ROS) increase with the storage time of rapeseed. SOD, CAT, and POD are the main antioxidant enzymes; therefore, it is necessary to increase the activity of antioxidant enzymes to reduce ROS accumulation. The effect of protocatechuic acid-mediated PDI on SOD(a), CAT(b), and POD(c) in rapeseed is shown in the figure. Figure 1 As shown, the SOD activity of the three rapeseed samples generally showed a decreasing trend during storage. The LP- storage group showed significant differences starting on day 1, while the L-P+ and L+P+ storage groups showed significant differences starting on day 3 (P < 0.01). The CAT activity of the L-P+ and L+P+ storage groups was consistently higher than that of the LP- storage group during storage. POD activity showed a trend of first increasing and then decreasing during storage, reaching its peak on day 5. These results indicate that protocatechuic acid-mediated PDI can effectively increase the activity of major antioxidant enzymes and reduce ROS accumulation during storage, thus serving as a means to slow down rapeseed senescence.

[0046] 3.3 Results of Vitamin C Content Determination in Rapeseed: The results of the vitamin C content determination of rapeseed are as follows: Figure 2 As shown. Figure 2 This figure shows the effect of protocatechuic acid-mediated PDI on vitamin C content in rapeseed. Vitamin C is a highly sensitive reducing agent; during storage, vitamin C in rapeseed is easily oxidized by reactive oxygen species (ROS), leading to a decrease in vitamin C content. Figure 2It was found that the vitamin C content in rapeseed generally showed a continuous decreasing trend with the extension of storage time. After 5 days of storage, the vitamin C content in the L+P+ storage group was 59.405 mg / g, and the vitamin C content in the LP- storage group was 41.089 mg / g, showing a highly significant difference between the two (P < 0.001). During storage, the vitamin C content of the three groups gradually decreased, but the decrease in the L+P+ storage group was relatively slow. This is related to the protocatechuic acid-mediated PDI (Precipitation Intake Discharge). PDI treatment has a protective effect on vitamin C, effectively slowing down the rate of vitamin C decrease during storage and reducing vitamin C loss.

[0047] 3.4 Results of the determination of lipid peroxidation index in rapeseed cell membranes: The results of the determination of lipid peroxidation index in rapeseed cell membranes are as follows: Figure 3 As shown. Figure 3 This graph shows the effect of protocatechuic acid-mediated PDI on MDA content in rapeseed. During storage, rapeseed cells produce large amounts of reactive oxygen species (ROS). ROS are byproducts of aerobic metabolism, including superoxide anions and hydrocarbon free radicals. These ROS induce peroxidation of unsaturated acids in rapeseed cell membranes or organelle membranes, generating lipid free radicals. These free radicals can further induce membrane lipid peroxidation, leading to increased cell membrane permeability, accelerated cell membrane damage, and ultimately, the decay of rapeseed tissue. Malondialdehyde (MDA) is one of the main products of membrane lipid peroxidation, reflecting the degree of cell membrane lipid peroxidation and the extent of cell membrane damage. Figure 3 It was found that the MDA content of the three rapeseed samples increased during storage, with significant differences starting on day 5 (P < 0.01). The membrane lipid peroxidation level in the LP- storage group was significantly higher than that in the L-P+ and L+P+ storage groups. The MDA content in the L+P+ storage group was the lowest. This is likely because PDI, induced by protocatechuic acid, binds to ROS, reducing damage to rapeseed cells and regulating the redox balance of rapeseed during storage. Compared to the L+P+ storage group, the MDA content in the L-P+ storage group was higher, which is likely due to the antioxidant effect of protocatechuic acid itself, effectively reducing damage to rapeseed. Therefore, by comparing the MDA content among the three groups, it can be concluded that protocatechuic acid-mediated PDI plays an important protective role in rapeseed cell membranes.

[0048] 3.5 Results of chlorophyll content determination in rapeseed: The results of the chlorophyll content determination of rapeseed are as follows: Figure 4 As shown. Figure 4 This is a graph showing the effect of protocatechuic acid-mediated PDI on chlorophyll content in rapeseed. From... Figure 4It can be seen that the chlorophyll content showed a continuous decreasing trend during storage. On the 5th day of storage, the chlorophyll a content in the L+P+ storage group was 6.5326 mg / L, while the chlorophyll a content in the LP- storage group was 5.5433 mg / L, showing a significant difference between the two (P < 0.05). The chlorophyll b content in the L+P+ storage group was significantly higher than that in the LP- storage group, indicating that the L+P+ storage group was more conducive to the storage of rapeseed and alleviated the yellowing rate of rapeseed.

[0049] Example 2: Investigating the effect of caffeic acid-mediated photodynamic therapy on the preservation of postharvest rapeseed. 1. Experimental Samples: Select fresh, uniform-sized, undamaged, and intact rapeseed. Rinse with tap water and allow to air dry naturally with a fan and windows open. Select two uniformly sized, fresh, and intact rapeseed plants as one group, with three replicates per group. Caffeic acid (purchased from Shanghai Yuanye Biotechnology Co., Ltd.). Total superoxide dismutase (SOD) assay kit, malondialdehyde (MDA) content assay kit, peroxidase (POD) activity assay kit, and catalase (CAT) assay kit were purchased from Nanjing Jiancheng Bioengineering Institute.

[0050] 2. Experimental methods: 2.1 Optimal parameter selection for caffeic acid-mediated photodynamic therapy: (1) Design a single-factor experimental treatment for rapeseed: First, caffeic acid treatment solutions of 0 mg / L, 0.01 mg / L, 0.025 mg / L, 0.05 mg / L, and 0.075 mg / L were prepared. The washed rapeseed samples were then soaked in the caffeic acid treatment solutions of different concentrations and then stored in a refrigerator at 4 ℃ in the dark. The samples were observed on days 1, 3, 5, and 7.

[0051] Secondly, a light intensity of 30 mW / cm was selected. 2 The light source was used to irradiate the plants for 0, 5, 10, 15, 20, and 25 minutes, respectively. The plants were then stored in a refrigerator at 4 ℃ in the dark and observed on days 1, 3, 5, and 7. The plants were turned over every 2 minutes during irradiation.

[0052] Then, the illumination time was selected as 10 min, and the light intensity was set at 0, 10, 20, 30, 40, and 50 mW / cm². 2 The pretreated samples were irradiated with different light intensities, stored in a refrigerator at 4 ℃ in the dark, and observed on days 1, 3, 5, and 7, with the samples turned over every 2 minutes.

[0053] (2) Design a three-factor, three-level orthogonal experiment: The optimal caffeic acid concentrations were determined to be 0.01 mg / L, 0.025 mg / L, and 0.05 mg / L, and the optimal light intensities were 30, 40, and 50 mW / cm², respectively, based on single-factor experiments. 2 An orthogonal experiment with three factors and three levels was conducted with light exposure times of 5, 10, and 15 minutes.

[0054] (3) Verification experiment: Based on the orthogonal experimental results, the specific parameters are: caffeic acid concentration 0.05 mg / mL, under LED blue light (illuminance 50 mW / cm²). 2 Irradiated for 10 minutes to verify the results. Subsequent experimental groups were as follows: Caffeic acid plus photodynamic therapy (CAI) group: 0.05 mg / mL caffeic acid, light intensity 50 mW / cm². 2 Irradiate for 10 minutes; Caffeic acid treatment group (CA group): 0.05 mg / mL caffeic acid; and control group (T group).

[0055] 2.2 Determination of antioxidant enzyme activity in rapeseed: The activity of antioxidant enzymes (SOD, POD, CAT) was measured using a kit. The groups were divided into caffeic acid plus photodynamic treatment group (CAI group), caffeic acid treatment group (CA group), and control group (T group). The enzyme activity of these three groups was measured.

[0056] 2.3 Determination of Vitamin C Content in Rapeseed: The changes in vitamin C content in rapeseed were determined using a kit. The groups were divided into three groups: caffeic acid plus photodynamic treatment group (CAI group), caffeic acid treatment group (CA group), and control group (T group). The vitamin C content of these three groups was measured.

[0057] 2.4 Determination of lipid peroxidation in rapeseed cell membranes: The level of cell membrane lipid peroxidation was characterized by measuring the MDA content in rapeseed using a kit. The groups were divided into three groups: caffeic acid plus photodynamic therapy group (CAI group), caffeic acid treatment group (CA group), and control group (T group). The MDA content of these three groups was measured.

[0058] 2.5 Determination of chlorophyll content in rapeseed: The determination of chlorophyll content in rapeseed was performed according to the "Experimental Methods of Food Preservation Technology". Rapeseed samples were placed in a mortar that had been pre-frozen overnight at low temperature and ground on ice. A small amount of quartz sand (0.3 g) and calcium carbonate powder (0.2 g) were added, followed by 3 mL of 95% ethanol. The mixture was quickly homogenized, and another 3 mL of 95% ethanol was added. The homogenate was then rapidly homogenized until the tissue turned white. The homogenate was quickly transferred three times with 6 mL of 95% ethanol into glass centrifuge tubes, then into 25 mL volumetric flasks. The flasks were brought to a final volume with 95% ethanol, the caps were tightened, and the mixture was shaken repeatedly. The mixture was then incubated in a biochemical incubator for 1 hour. After incubation, the mixture was transferred to 15 mL plastic centrifuge tubes and centrifuged at 10 ℃ and 8000 r / min for 15 min. Immediately after centrifuging, approximately 10 mL of the supernatant was carefully transferred to a clean centrifuge tube using a pipette next to the centrifuge (if plant tissue is accidentally transferred, the process of re-centrifuging and re-collecting must be repeated, as this step greatly affects the test results). Before the experiment, turn on the air conditioner in the UV chamber and maintain a temperature of 25 °C for at least 1 hour. Turn on the UV spectrophotometer and preheat it. Click on baseline calibration under no-load conditions, then select the spectral function, set the detection wavelength, and set the method. Add the reference solution (95% ethanol) to two cuvettes, then click zeroing. Remove the cuvettes from the sample cell and load them with samples one by one for testing; no further zeroing is required. Measure the absorbance at 663 nm and 645 nm, using 95% ethanol as a blank control. Store the test solutions away from light; measure one solution at a time before moving on to the next.

[0059] The chlorophyll concentration in the extract is: In the formula: cT -- chlorophyll concentration in the extract, mg / L W -- Fresh weight of the sample, in grams N -- Dilution factor.

[0060] 3. Results and Analysis: 3.1 Optimal parameter screening results for caffeic acid-mediated photodynamic therapy: The optimal preservation parameters for rapeseed using caffeic acid-mediated photodynamic therapy (PDPT) were determined. The primary and secondary factors influencing rapeseed storage were caffeic acid concentration > light intensity > light duration. The optimal preservation parameters for rapeseed using Caffeic acid-mediated PPT were: rapeseed immersed in a 0.05 mg / L caffeic acid treatment solution, with an irradiation intensity of 50 mW / cm². 2 The illumination time was 10 minutes.

[0061] 3.2 Results of the determination of antioxidant enzyme activity in rapeseed: The results of the determination of antioxidant enzyme activity in rapeseed are as follows: Figures 5-7 As shown. Figure 5 The figure shows the effect of caffeic acid-mediated PDI on SOD activity in rapeseed. Figure 6 The figure shows the effect of caffeic acid-mediated PDI on CAT activity in rapeseed. Figure 7 The figure shows the effect of caffeic acid-mediated PDI on POD activity in rapeseed.

[0062] 3.2.1 The effect of caffeic acid-mediated PDI on SOD activity in rapeseed is as follows: Figure 5 As shown: Superoxide dismutase (SOD) acts as a scavenger of superoxide anion free radicals (O2) in plants. - The first key line of defense in rapeseed is its activity level, which is an important indicator for measuring the strength of antioxidant capacity and aging process of post-harvest vegetables. During post-harvest storage, rapeseed experiences an imbalance in normal physiological metabolism, resulting in the continuous production of reactive oxygen species (ROS). When ROS accumulates beyond a certain threshold, it leads to cell structure damage, ultimately manifesting as yellowing, wilting due to dehydration, and even rotting. Figure 5 As shown, throughout the storage period, the SOD activity of all experimental groups showed a common trend of first increasing and then decreasing. In the early stages of storage, rapeseed was subjected to strong oxidative stress, and its own antioxidant system was rapidly activated. However, rapeseed's own defense capabilities are limited, and excessive ROS caused damage or degradation of enzyme proteins, resulting in the inability to maintain SOD activity. Therefore, the SOD activity of group T peaked on day 1 and then decreased sharply. Groups CA and CAI also showed a decreasing trend, but groups CA and CAI were consistently significantly higher than group T (P<0.001). The rate of decrease in group CAI was relatively gradual after day 3, and its SOD activity was higher than that of group CA in the later stages of storage. Caffeic acid itself has antioxidant properties and can protect SOD activity. However, when caffeic acid is combined with photodynamic therapy, its antioxidant effect is enhanced. While delaying the decline of SOD activity in the later stages of storage, it maintains a more durable and efficient ROS scavenging capacity, which has a positive effect on the postharvest physiological and biochemical reactions of rapeseed.

[0063] 3.2.2 The effect of caffeic acid-mediated PDI on CAT activity in rapeseed is as follows: Figure 6 As shown: Catalase (CAT) catalyzes the decomposition of H2O2 into O2 and H2O to eliminate the toxic effects of H2O2. Changes in CAT activity are related to the ability of cells to avoid oxidative damage. During storage, the CAT activity of each treatment group generally showed a trend of first increasing and then decreasing, with significant differences in peak activity and maintenance capacity among different treatment groups (P<0.05). In group T, CAT activity reached its peak on day 1 and then continuously decreased. This was because the rapid accumulation of H2O2 in rapeseed during the early stages of storage induced an upregulation of CAT activity, but the antioxidant system was overloaded, leading to CAT activity depletion. In contrast, the CA group showed a more gradual decline, with a higher peak activity and a longer peak duration than group T. Caffeic acid, as an antioxidant, helped CAT to some extent to remove peroxides and maintain CAT activity levels. The CAT activity in the CAI group was significantly higher than that in the first two groups (P<0.001), with the highest peak level. In particular, its activity showed a significant advantage on days 3, 5, and 7, which were 1.33 times, 1.87 times, and 2.16 times that of the T group, respectively. This indicates that caffeic acid acts as both an antioxidant and a photosensitizer, and under the action of photodynamic therapy, it significantly enhances and maintains the CAT activity of postharvest rapeseed, effectively reducing oxidative damage to rapeseed, thereby delaying senescence and yellowing, and achieving a preservation effect.

[0064] 3.2.3. The effect of caffeic acid-mediated PDI on the POD activity of rapeseed is as follows: Figure 7 As shown: Peroxidase (POD) is a redox enzyme in plants that can synergistically scavenge H2O2 with caffeic acid (CAT) to reduce oxidative damage. With prolonged storage, the POD activity of all three treatment groups initially increased and then decreased, with no significant difference in the time of peak activity (P>0.05), but significant differences in peak activity (P<0.001), in the order of CAI group > CA group > T group. During storage, the CAI group maintained the highest POD activity, followed by the CA group, while the T group had the lowest POD activity. This is because caffeic acid, as an exogenous antioxidant, can directly scavenge reactive oxygen species (ROS), maintaining a high level of POD activity. Under photodynamic therapy, the scavenging ability of caffeic acid is enhanced, thus the POD activity level of the CAI group is higher than that of the CA group. This indicates that caffeic acid-mediated photodynamic therapy is more effective in preserving rapeseed than caffeic acid treatment alone.

[0065] 3.3 Results of Vitamin C Content Determination in Rapeseed: The results of the vitamin C content determination of rapeseed are as follows: Figure 8 As shown. Figure 8The graph shows the effect of caffeic acid-mediated PDI on vitamin C content in rapeseed. Vitamin C is an important antioxidant, and its content is one of the key indicators for evaluating the nutritional quality and antioxidant capacity of post-harvest vegetables. During storage, the vitamin C content decreased in all groups. The vitamin C content in groups CA and CAI was consistently significantly higher than that in group T (P<0.001), while there was no significant difference in vitamin C content between groups CA and CAI (P>0.05). Post-harvest rapeseed continues to respire and accumulates a large amount of reactive oxygen species (ROS) during storage. Vitamin C, as a major antioxidant, is largely consumed to remove harmful substances, leading to a deterioration in the nutritional quality of rapeseed. Caffeic acid itself has antioxidant properties and can directly react with ROS, reducing the consumption of vitamin C in rapeseed tissues. Under certain light intensity, the light source also has the effect of scavenging ROS, exhibiting a synergistic effect with caffeic acid. Therefore, the vitamin C content in group CAI was higher than that in the other two groups, indicating that caffeic acid, under light treatment, exhibits a strong vitamin C retention capacity, effectively delaying the decline in the nutritional quality of post-harvest rapeseed.

[0066] 3.4 Results of the determination of lipid peroxidation index in rapeseed cell membranes: The results of the determination of lipid peroxidation index in rapeseed cell membranes are as follows: Figure 9 As shown. Figure 9 This graph shows the effect of caffeic acid-mediated PDI on MDA content in rapeseed. Malondialdehyde (MDA) is the end product of membrane lipid peroxidation, and its content is a key indicator for measuring the degree of damage to the cell membrane system. Figure 9 As shown, the MDA content in all treatment groups increased throughout the storage process, with significant differences in the MDA accumulation rate and level among the treatment groups. Group T had the highest MDA content and the fastest growth rate, indicating severe oxidative damage to rapeseed leaf cells. Membrane lipid peroxidation led to increased cell membrane permeability and cell structure destruction. Group CA maintained a low MDA content; caffeic acid molecules can directly scavenge reactive oxygen species (ROS), mitigating oxidative damage to the membrane system to some extent and thus delaying rapeseed yellowing and wilting symptoms. Group CAI showed the best effect in inhibiting MDA accumulation, with its MDA content significantly lower than the other two groups (P<0.001). Caffeic acid, as a photosensitizer, was activated under specific light conditions, enhancing its antioxidant efficacy, effectively inhibiting membrane lipid peroxidation, and maintaining cell membrane integrity and stability. In conclusion, this demonstrates that caffeic acid and photodynamic therapy have a synergistic effect on postharvest rapeseed storage and preservation.

[0067] 3.5 Results of chlorophyll content determination in rapeseed: The results of the chlorophyll content determination of rapeseed are as follows: Figure 10 As shown. Figure 10The figure shows the effect of caffeic acid-mediated PDI on chlorophyll content in rapeseed. Rapeseed is rich in chlorophyll, and its content can reflect the degree of chlorosis. Chlorophyll includes chlorophyll a and chlorophyll b. During the 7-day storage period, the contents of chlorophyll a, chlorophyll b, and total chlorophyll in all treatment groups showed a decreasing trend. Since group T received no treatment, reactive oxygen species (ROS) accumulated in the rapeseed tissue, damaging the chloroplast membrane structure and causing severe yellowing of the leaves. Therefore, the contents of chlorophyll a, chlorophyll b, and total chlorophyll decreased significantly, and the chlorophyll content level was the lowest. The contents of chlorophyll a, chlorophyll b, and total chlorophyll in group CA were higher than those in group T (P<0.001), while those in group CAI were significantly higher than those in the other two groups (P<0.001). Caffeic acid molecules can effectively scavenge ROS to reduce the degree of oxidative damage to the chloroplast membrane. Caffeic acid enhances the antioxidant effect under photodynamic therapy and delays its enzymatic degradation process, thereby achieving excellent greening effect. On day 7, the total chlorophyll content of group T was 0.38 mg / g fresh weight, while the total chlorophyll content of groups CA and CAI was 47.95% and 66.96% higher than that of group T, respectively. In summary, caffeic acid-mediated photodynamic therapy is the most effective way to delay chlorophyll degradation and maintain the chlorophyll content and good leaf quality of rapeseed.

[0068] Example 3: A method for preserving rapeseed Step S1: Select uniform-sized, undamaged, fresh and intact rapeseed, wash it clean, and air dry it to obtain clean rapeseed; Step S2: Dissolve protocatechuic acid in distilled water, stir well, and prepare a protocatechuic acid solution with a concentration of 0.075 mg / L to obtain the treatment solution; Step S3: Completely immerse the cleaned rapeseed treated in step S1 in the treatment solution prepared in step S2, and place it under a light intensity of 50 mW / cm². 2 Irradiate the product under LED blue light at a distance of 20 cm from the light source for 5 minutes, flipping it over every 2 minutes, then package and refrigerate.

[0069] Example 4: A method for preserving rapeseed Step S1: Select uniform-sized, undamaged, fresh and intact rapeseed, wash it clean, and air dry it to obtain clean rapeseed; Step S2: Dissolve caffeic acid in distilled water, stir well, and prepare a caffeic acid solution with a concentration of 0.05 mg / L to obtain the treatment solution; Step S3: Completely immerse the cleaned rapeseed treated in step S1 in the treatment solution prepared in step S2, and place it under a light intensity of 50 mW / cm². 2 Irradiate the product under LED blue light at a distance of 20 cm from the light source for 10 minutes, flipping it over every 2 minutes, then package and refrigerate.

[0070] Example 5: A method for preserving rapeseed Step S1: Select uniform-sized, undamaged, fresh and intact rapeseed, wash it clean, and air dry it to obtain clean rapeseed; Step S2: Dissolve protocatechuic acid in distilled water, stir well, and prepare a protocatechuic acid solution with a concentration of 0.075 mg / L to obtain the treatment solution; Step S3: Completely immerse the cleaned rapeseed treated in step S1 in the treatment solution prepared in step S2, and place it under a light intensity of 50 mW / cm². 2 Under LED blue light, at a distance of 20 cm from the light source, irradiate for 5 minutes, turning the rapeseed over every 2 minutes. Then, soak the rapeseed in the coating solution for 5 minutes, drain, package, and refrigerate to obtain the product. The preparation method of the coating solution is as follows: Chitosan was dissolved in a 1% (v / v) acetic acid solution to prepare a 1.5% (w / v) chitosan solution. Protocatechuic acid was added to distilled water to prepare a 0.075 mg / L protocatechuic acid solution. The chitosan solution was then added to the protocatechuic acid solution at a volume ratio of 2:1, and the mixture was stirred until homogeneous.

[0071] Example 6: A method for preserving rapeseed Step S1: Select uniform-sized, undamaged, fresh and intact rapeseed, wash it clean, and air dry it to obtain clean rapeseed; Step S2: Dissolve caffeic acid in distilled water, stir well, and prepare a caffeic acid solution with a concentration of 0.05 mg / L to obtain the treatment solution; Step S3: Completely immerse the cleaned rapeseed treated in step S1 in the treatment solution prepared in step S2, and place it under a light intensity of 50 mW / cm². 2 Under LED blue light, at a distance of 20 cm from the light source, irradiate for 10 minutes, turning the rapeseed over every 2 minutes. Then, soak the rapeseed in the coating solution for 5 minutes, drain, package, and refrigerate to obtain the final product. The preparation method of the coating solution is as follows: Chitosan was dissolved in a 1% (v / v) acetic acid solution to prepare a 1.5% (w / v) chitosan solution; caffeic acid was added to distilled water to prepare a 0.05 mg / L protocatechuic acid solution; the chitosan solution was added to the caffeic acid solution at a volume ratio of 2:1, and the mixture was stirred until homogeneous to obtain the final product.

[0072] Example 7: Preservation effect test of rapeseed 1. Experimental Method: Observe and record the number of days it took for rapeseed leaves stored under cold storage in Examples 3, 4, 5, and 6 to yellow to grade 2. Six rapeseed plants were counted in each group, with three replicates. Specifically, the outer leaves of the rapeseed were divided into sections, and based on the proportion of yellowed tissue to the total leaf area, they were classified into grades 0-4: Grade 0: no yellowing; Grade 1: yellowed area not exceeding 25%; Grade 2: yellowed area 25%-50%; Grade 3: yellowed area 50%-75%; Grade 4: rotten area 75%-100%.

[0073] 2. Experimental Results: The experimental results are shown in Table 1.

[0074] Table 1. Preservation effect of rapeseed As shown in Table 1, the rapeseed preservation method provided by the present invention can significantly extend the storage period of rapeseed.

Claims

1. A method for preserving rapeseed, characterized in that, Includes the following steps: Step S1: Select uniform-sized, undamaged, fresh and intact rapeseed, wash it clean, and air dry it to obtain clean rapeseed; Step S2: Dissolve the photosensitizer in distilled water and stir until homogeneous to obtain the treatment solution; Step S3: Completely immerse the clean rapeseed treated in step S1 in the treatment solution prepared in step S2, irradiate it under blue light for 5-15 minutes, package it, and store it in the refrigerator.

2. The method for preserving rapeseed as described in claim 1, characterized in that, The photosensitizer in step S2 is protocatechuic acid or caffeic acid.

3. The method for preserving rapeseed as described in claim 2, characterized in that, When the photosensitizer is protocatechuic acid, the concentration of the protocatechuic acid treatment solution is 0.05~0.1 mg / L.

4. The method for preserving rapeseed as described in claim 2, characterized in that, When the photosensitizer is caffeic acid, the concentration of the caffeic acid treatment solution is 0.01~0.075 mg / L.

5. The method for preserving rapeseed as described in claim 1, characterized in that, The blue light irradiation conditions in step S3 are: light intensity of 30~50 mW / cm². 2 The distance from the light source should be 18-25 cm, the irradiation time should be 5-15 min, and the surface should be flipped over every 2 min.

6. The method for preserving rapeseed as described in claim 1, characterized in that, Step S3 also includes a coating treatment step, which involves soaking the rapeseed in a coating solution for 4-6 minutes after light treatment, draining it, and packaging it to obtain the final product.

7. The method for preserving rapeseed as described in claim 6, characterized in that, The preparation method of the coating liquid is as follows: Chitosan was dissolved in a 1% (v / v) acetic acid solution to prepare a chitosan solution; photosensitizer was added to distilled water to prepare a photosensitizer solution; the chitosan solution was added to the photosensitizer solution and stirred until homogeneous to obtain the final product.

8. The method for preserving rapeseed as described in claim 7, characterized in that, The chitosan solution contains a chitosan concentration of 1-2% w / v.

9. The method for preserving rapeseed as described in claim 7, characterized in that, The photosensitizer is protocatechuic acid or caffeic acid. When the photosensitizer is protocatechuic acid, the concentration of the protocatechuic acid solution is 0.05~0.1 mg / L. When the photosensitizer is caffeic acid, the concentration of the caffeic acid solution is 0.01~0.075 mg / L.

10. The method for preserving rapeseed as described in claim 7, characterized in that, The volume ratio of the photosensitizer solution to the chitosan solution is 1:(2~3).

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