A method for extending the shelf life of strawberries using silica nanomaterials

Abstract

This invention belongs to the field of fruit preservation technology and relates to a method for extending the shelf life of strawberries using silica nanomaterials. Addressing the problems of chemical reagent residues, high energy consumption of physical methods, and unstable effectiveness of biological methods in existing strawberry post-harvest preservation technologies, this invention provides a safe and efficient solution. The invention involves spraying a 100 mg / L aqueous solution of silica nanoparticles with a particle size of 40-80 nm onto the surface of freshly picked strawberries, at a dosage of 100-200 μL per fruit. These nanoparticles can target and inhibit the key rate-limiting enzyme gene for ethylene synthesis in strawberry fruits. ACS1 This invention reduces endogenous ethylene production at the source, effectively delaying fruit ripening and senescence, cell wall degradation, and microbial infection. The method is simple to operate, can be implemented at room temperature, and the silica nanomaterials used are non-toxic to plants, environmentally friendly, and harmless to human health. The preservation effect is significantly better than traditional chemical preservatives, providing a stable and environmentally friendly new technical approach for post-harvest strawberry preservation.

Description

Technical Field

[0001] This invention belongs to the field of preservation technology and relates to a new method for extending the shelf life of strawberries using silica nanomaterials. Background Technology

[0002] Strawberries have thin, delicate skins that are easily damaged by mechanical forces, microbial contamination, and ethylene-mediated ripening and senescence, leading to rapid softening and rotting. Post-harvest losses can reach 20-40%, severely hindering the large-scale development and economic benefits of the strawberry industry. Post-harvest preservation methods for strawberries mainly include chemical preservation, physical preservation, biological preservation, and novel nanomaterial preservation methods. Chemical preservation methods use chemical preservatives and ethylene inhibitors to regulate the senescence and decay process of the fruit and reduce microbial contamination.

[0003] Commonly used chemical reagents include 1-methylcyclopropene (1-MCP), potassium sorbate, and sodium benzoate. Among them, 1-MCP, as a highly efficient competitive inhibitor of ethylene receptors, can delay fruit ripening and senescence by blocking ethylene signal transduction and is widely used in strawberry preservation. However, it requires a closed fumigation process, which is cumbersome, and long-term use may affect the fruit flavor. Traditional chemical preservatives such as potassium sorbate can inhibit microbial growth and extend shelf life, but they are prone to leaving residues on the fruit surface. Excessive intake may pose potential health hazards to humans, and their stability is insufficient, making it difficult to achieve long-term preservation.

[0004] Physical preservation methods mainly include low-temperature storage, modified atmosphere packaging, radiation treatment, and plasma treatment. Their core principle is to inhibit fruit metabolism and microbial growth by regulating storage environment parameters. Low-temperature storage can effectively delay strawberry spoilage, but this technology relies on continuous refrigeration equipment, resulting in high energy consumption and operating costs. Furthermore, long-term low-temperature storage can easily lead to chilling injury in strawberries, affecting fruit flavor and quality. Simultaneously, low temperatures cannot completely inhibit ethylene synthesis, failing to fundamentally delay the fruit ripening and senescence process. Modified atmosphere packaging, by adjusting the oxygen and carbon dioxide concentrations in the storage environment, inhibits fruit respiration and ethylene release, extending shelf life. However, this technology is complex to operate, requires significant equipment investment, and gas concentrations are difficult to control precisely, easily leading to anaerobic fermentation in the fruit. While novel physical preservation technologies such as radiation treatment and plasma treatment have the advantage of leaving no chemical residues, their high equipment costs and technological barriers hinder industrial-scale promotion.

[0005] Biological preservation methods utilize natural bioactive substances (such as plant extracts, probiotics, and antimicrobial peptides) as their core, achieving strawberry preservation by inhibiting microbial proliferation and scavenging reactive oxygen species. For example, composite preservation films prepared using plant extracts such as quercetin and hesperidin can extend the shelf life of strawberries through antibacterial and antioxidant effects. However, these bioactive substances are easily degraded, leading to unstable effects and a relatively short preservation period, making it difficult to meet long-term preservation needs. Furthermore, biological preservation technology is costly and difficult to produce, limiting its practical application. Summary of the Invention

[0006] To address the aforementioned problems, this invention provides a method for extending the shelf life of strawberries by inhibiting ethylene synthesis-related genes in plants using silica nanomaterials at room temperature. The SiO2NPs used in this method are nanoscale inorganic materials with various morphologies, characterized by numerous micropores, large specific surface area, stable effects, no phytotoxicity, no pollution, and no harm to human health. The SiO2NPs used in this invention have a diameter range of 40-80 nm, allowing them to completely penetrate the leaves through stomata and then enter plant cells. The high surface area to volume ratio of SiO2NPs enhances their reactivity and biochemical activity. After entering plant cells, they extend the shelf life of strawberries by enhancing the plant's own immunity and inhibiting the expression of the ACS1 gene in the fruit.

[0007] The specific technical solution involves spraying 100-200 μL (per fruit), 100 mg / L, and 40-80 nm diameter silica nanoparticle aqueous solution onto the surface of freshly picked strawberries.

[0008] The present invention has the following advantages: (1) The nano-silica particles (SiO2NPs) of this invention can act on the endogenous ethylene synthesis pathway in strawberry fruit under normal temperature conditions. ACS1 The gene-encoded ACC synthase is the rate-limiting enzyme, catalyzing the production of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) from S-adenosylmethionine (SAM). ACC is then catalyzed by ACC oxidase (ACO) to generate endogenous ethylene. SiO2NPs can target and inhibit strawberry fruit development. ACS1 By reducing the protein expression and bioenzyme activity of the rate-limiting enzyme ACC synthase from the source, and reducing the synthesis of ACC intermediates, the technical effect of extending the postharvest shelf life of strawberries is achieved. This effectively inhibits the generation and signal transduction of endogenous ethylene in strawberries, thereby delaying the ripening and senescence of strawberry fruits, cell wall degradation and microbial infection.

[0009] (2) SiO2NPs treatment of red strawberry and related genes for ethylene synthesis in strawberry ACS1The expression of these genes is suppressed, thereby reducing endogenous ethylene synthesis and delaying strawberry senescence and spoilage. SiO2NPs extend the shelf life of strawberries at room temperature primarily by inhibiting ethylene synthesis-related genes at the molecular level. ACS1 This method, described as a stable, non-toxic, environmentally friendly, and harmless strawberry preservation technique, offers a safe, efficient, and environmentally friendly new technological solution for post-harvest strawberry preservation. It has significant application potential in promoting the sustainable development of the strawberry industry, reducing post-harvest losses, and enhancing the industry's economic benefits. Attached Figure Description

[0010] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.

[0011] Figure 1 The images show the appearance phenotypes of strawberries treated with different preservatives in Experiment 1, used to compare the preservation time and preservation status of strawberries in each group. From top to bottom, they are: sterile water control group, SiO2NPs treatment group, 1-MCP treatment group, E2O2 treatment group, and SiO2NPs+1-MCP combined treatment group.

[0012] Figure 2 This is a statistical analysis of the number of days the strawberries in the control group and each treatment group remained fresh after harvest in Example 1.

[0013] Figure 3 The relative expression levels of the strawberry ethylene synthesis gene ACS1 in each group in Example 2 are shown. H2O is the control group. Detailed Implementation

[0014] The technical solutions in the embodiments of the invention are described clearly and completely 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.

[0015] Experimental Example 1 The silica nanoparticles (SiO2NPs) described in this invention can enter cells through the stomata of the fruit epidermis and induce a systemic immune response and related physiological regulation in the plant. This process has a certain response cycle. Therefore, this experiment selected the deep color-changing stage (when the colored area reaches 50-80%) before strawberry fruit harvest for pretreatment. The surface of the fruit treated with SiO2NPs was uniformly sprayed with 100 μL (per fruit, the same applies below) of a 100 mg / L SiO2NPs solution. All other experimental groups were sprayed with an equal volume of sterile water during the deep color-changing stage, with the spraying distance controlled at 15-20 cm, ensuring uniform coverage of the fruit surface without any residual liquid. After pretreatment, the strawberry plants in each group, including the SiO2NPs-treated group, were isolated and cultured in the same environment to prevent interference between groups. Subsequent storage experiments were conducted after the fruit reached the red ripening stage (completely red overall, with a colored area of ​​80-95%).

[0016] When the fruit reaches the red ripening stage, select Red Face strawberry plants with uniform growth and no pests or diseases. From these plants, select healthy and uniform fruits without mechanical damage and mark them. The test fruits are randomly divided into 5 groups, with 3 biological replicates in each group. The specific treatment methods for the strawberry fruits in each group after harvest are as follows: Blank control group: 100 μL of sterile water was sprayed evenly on the surface of the fruit; 1-MCP (1-methylcyclopropene) treatment group: 100 μL of sterile water was sprayed on the surface of the fruit, and 0.0028 g of 1-MCP was placed in a closed space for fumigation treatment; E202 (potassium sorbate) treatment group: 100 μL of potassium sorbate solution with a concentration of 1000 mg / L was sprayed evenly on the surface of the fruit; SiO2NPs treatment group: 100 μL of SiO2NPs solution with a concentration of 100 mg / L was sprayed evenly on the surface of the fruit; SiO2NPs+1-MCP composite treatment group: 100 μL of SiO2NPs solution with a concentration of 100 mg / L was sprayed on the surface of the fruit, and 0.0028 g of 1-MCP was placed in a closed space. The treatment was carried out by a combination of spraying and fumigation.

[0017] After processing, place all fruits in a container with a volume of 141.3 cm³. 3 The fruits were placed in sealed containers. Each group of fruits was arranged in a separate area, isolated from the others, and stored in a uniform environment at 22℃ and 65% relative humidity. During storage, the fruit condition was observed daily, with the appearance of rotten spots or mold mycelium on the fruit surface as the criterion for rotting. The time from freshness to the first occurrence of rot was recorded for each group of fruits to evaluate the preservation effect of different treatment methods. The results are shown in […]. Figure 1-2 .

[0018] Depend on Figure 1-2 It can be seen that the blank control group's fruits began to rot and mold after 3-4 days of storage; the 1-MCP treatment group and the potassium sorbate treatment group successively developed mold on the 6th-7th day of storage; the SiO2NPs single treatment group and the SiO2NPs and 1-MCP combined treatment group had the fruit mold starting time on the 8th-9th day of storage.

[0019] Experiments have shown that SiO2NPs can effectively delay the onset of mold on strawberry fruits, inhibit fruit senescence, and slow down the postharvest spoilage process. Compared with two conventional fruit and vegetable preservatives, 1-methylcyclopropene and potassium sorbate, SiO2NPs have a more prominent effect on improving the postharvest storage and preservation of Red Beauty strawberries.

[0020] Experimental Example 2 Red Beauty strawberry plants with uniform growth and free from pests and diseases were selected and randomly divided into a control group and an experimental group, with several plants in each group. Leaves of uniform size and shape were selected from both groups and marked to ensure the uniformity of the experimental materials. Spraying was uniformly carried out at 11:00 AM daily, when the stomatal opening of the strawberry leaves is at its maximum, facilitating the absorption of the foliar treatment solution.

[0021] Two days before the formal testing, foliar pretreatment was carried out: spraying was conducted using a sprayer, with the sprayer held perpendicular to the leaves and the spraying distance controlled at 15-20 cm to ensure uniform spraying of the pesticide solution; the spraying was terminated when fine water droplets formed on the leaf surface without any dripping, ensuring uniform dosage for each group and eliminating interference from uneven spraying on the experimental results. Specifically, 100 μL of sterile water was sprayed onto the abaxial surface of the leaves of the control group; and an equal volume of a 100 mg / L silica nanoparticle (SiO2NPs) solution was sprayed onto the abaxial surface of the leaves of the experimental group.

[0022] Two days after pretreatment, leaf samples were collected, and ethylene synthesis-related genes were detected and analyzed using real-time quantitative PCR (qPCR). ACS1 The relative expression level.

[0023] Unless otherwise specified, all methods used below are standard methods. Primers were provided by Shanghai Sangon Biotech Co., Ltd. RNA extraction kit, reverse transcription kit, qPCR kit, and Taq DNA polymerase were purchased from Beijing Solarbio Science & Technology Co., Ltd.

[0024] Step 1, Total RNA Extraction and Quality Assay: Intact leaf samples were cut and placed in 50 mL centrifuge tubes, then immediately flash-frozen in liquid nitrogen to effectively inhibit RNA degradation. Total RNA was extracted from the leaves using an RNA extraction kit. The extracted products were analyzed using agarose gel electrophoresis and a NanoDrop spectrophotometer to assess RNA integrity, concentration, and purity, ensuring the sample quality met the requirements for subsequent experiments.

[0025] Step 2, reverse transcription synthesis: The total RNA that has passed the test is reverse transcribed using a reverse transcription kit to synthesize complementary DNA (cDNA), which is then used as a reaction template for real-time quantitative PCR (qPCR).

[0026] Step 3, Real-time quantitative PCR amplification: Prepare the qPCR reaction system by adding upstream and downstream primers, the above-mentioned cDNA template, and Taq DNA polymerase to the system, and carry out the amplification reaction. Monitor the changes in fluorescence signal in real time during the experiment.

[0027] qPCR target gene primer sequences: qFaACS1-F 5'-AAGAGGTTGGAAGCAAGGCA-3', SEQ ID No. 1; qFaACS1-R 5'-CTGGTTGGGGGCAATGAAAAGA-3', SEQ ID No. 2; qPCR internal reference gene primer sequences: qFaACTIN-F 5'-GGGCAGAAAGATGCTTATGTCGG-3', SEQ ID No. 3; qFaACTIN-R 5'-GGGCAACAGAAGTCATTGTAGAAG-3', SEQ ID No. 4; Reaction procedure: 95℃ pre-denaturation for 5 minutes; 94℃ denaturation for 30 seconds; 55℃ annealing for 30 seconds; 72℃ extension for 1 minute; 35 cycles; 72℃ extension for 10 minutes. After the reaction, the data were analyzed.

[0028] Step four: Analyze the data obtained from real-time quantitative PCR detection and calculate the ΔCt value of the target gene relative to the internal reference gene in each sample.

[0029] Step 5: Calculate the average ΔCt of all samples in the control group; subtract the average ΔCt of the control group from the ΔCt value of a single sample in the experimental group to obtain the ΔCt value of each sample. This index reflects the difference in gene expression between the experimental group and the control group.

[0030] Step 6: Calculate the relative gene expression levels of each sample in the experimental group relative to the control group using the 2ΔΔCt method, and then calculate the arithmetic mean of the relative expression levels of each sample in the experimental group.

[0031] Step 7: Use GraphPad Prism 8.0.2 software to create bar charts to visually compare the relative expression levels of the target gene in the control and experimental groups. The experimental results are shown below. Figure 3 .

[0032] Depend on Figure 3 It can be seen that the genes in the control group and the experimental group plants are different. ACS1 The expression levels of these substances differed significantly, with the experimental group showing the highest levels. ACS1 The expression level of SiO2NPs was 1.96 times lower than that of the control group, indicating that SiO2NPs inhibited the expression of strawberry. ACS1 Gene expression levels.

[0033] According to Experiment 2, SiO2NPs inhibited genes related to ethylene synthesis in strawberries. ACS1 The expression of [the substance] inhibits the synthesis and release of endogenous ethylene. Combined with Experiment 1, it can be seen that SiO2NPs can effectively delay the senescence and spoilage of strawberry fruits, thereby extending the post-harvest shelf life of strawberries.

[0034] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for extending the shelf life of strawberries using nano-silica materials, characterized in that, Spraying freshly picked strawberries with an aqueous solution of silica nanoparticles inhibits genes related to ethylene synthesis. ACS1 The expression.

2. The method for extending the shelf life of strawberries using nano-silica material according to claim 1, characterized in that, The concentration of the aqueous solution of the silica nanoparticles is 100 mg / L.

3. The method for extending the shelf life of strawberries using nano-silica material according to claim 2, characterized in that, The application rate of the silica nanoparticle aqueous solution is 100-200 μL per fruit.

4. The method for extending the shelf life of strawberries using nano-silica material according to claim 1, characterized in that, The particle size of the silica nanoparticles is 40-80 nm.

5. The method for extending the shelf life of strawberries using nano-silica material according to claim 1, characterized in that, The strawberry variety is the Red Face strawberry.

6. The application of nano-silica in the method of any one of claims 1-5 in the field of extending shelf life.

7. The nano-silica in the method according to any one of claims 1-5 in inhibiting gene... ACS1 Application in expression.

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