In-situ preparation method of nano-silver-antibacterial peptide conjugate agar antibacterial film

Abstract

The invention discloses an in-situ preparation method of a nano-silver-antibacterial peptide conjugate agar antibacterial film. The in-situ preparation method comprises the following steps: dissolving pectin in an alkali solution to obtain a pectin alkali solution; adding nisin and silver salt into the pectin alkali solution, and mixing to obtain a colloidal solution; agar and an emulsifier are added into the colloidal solution, and the nano-silver-antibacterial peptide conjugate agar antibacterial film is obtained after mixing. Biomacromolecule pectin is used as a reducing agent and a stabilizing agent, pectin molecules contain a large number of carboxyl groups, the carboxyl groups can be ionized under the alkaline condition to form carboxyl anions, the reducing capacity of the pectin molecules is improved, silver ions can be reduced into silver atoms, and therefore synthesis of nano-silver is promoted; in addition, the alkaline condition can improve the reaction rate, improve the dispersity of the nano-silver and improve the stability of the nano-silver. The colloidal solution contains a nano silver-antibacterial peptide conjugate.

Description

Technical Field

[0001] The invention relates to the field of food industry, and in particular to an in-situ preparation method of a nano-silver-antimicrobial peptide conjugate agar antimicrobial film. Background Art

[0002] Traditional petroleum-based plastic materials and waste plastic materials have caused many environmental problems due to their difficult-to-degrade properties, and people's demand for environmentally friendly materials has greatly increased. Various biopolymers from natural resources are abundant, renewable, environmentally friendly and sustainable. They are eco-friendly materials and have been used in the development of biodegradable food packaging or edible films. Biopolymer-based packaging materials are made from natural materials such as carbohydrates, proteins and lipids. In particular, carbohydrate-based biopolymers have good colloidal properties and reasonable structures and are very ideal film-forming matrices. Agar is a linear polysaccharide extracted from red algae. It is composed of alternating β-(1,3)- and α-(1,4)-linked galactose residues and contains sulfated functional groups. It is one of the carbohydrate materials and is widely used as a gelling agent in the food industry. It is also used to prepare transparent and flexible biopolymer films. However, the weak mechanical properties and lack of antibacterial ability of biopolymer films limit their application in the packaging field.

[0003] In recent years, the use of various types of nanofillers, such as nanoclays, metals, or metal oxide nanomaterials as reinforcing fillers combined with thin films to improve the properties of thin films may be a promising solution.

[0004] Among nanofillers, silver nanoparticles (AgNPs) have been used to prepare antimicrobial active food packaging films due to their extensive and unique antimicrobial activity and high thermal stability. Rhim et al. (2013) prepared agar / AgNPs composite films and found that AgNPs were evenly distributed in the polymer matrix. The film properties were affected by the content of added silver nanoparticles, and showed strong inhibitory activity against both Gram-negative and Gram-positive pathogens. However, there are many limitations in the application of AgNPs in the food industry. For example, traditional physical and chemical preparation methods require high costs and toxic reagents, and there is an urgent need to improve antibacterial properties and biosafety. In order to overcome these problems, people choose to use biomacromolecules to synthesize AgNPs, which can effectively reduce costs and avoid the use of toxic reagents. Shankar et al. (2017) used amino acids as reducing agents and capping agents to synthesize AgNPs, and combined them with agar to prepare nanocomposite films. The experimental results show that the composite film containing AgNPs exhibits ultraviolet blocking properties and exhibits strong inhibitory effects on Lactobacillus monocytogenes and Escherichia coli. In addition, combining AgNPs with active organic molecules such as antimicrobial peptides to change their physical and chemical properties has a positive effect on improving antimicrobial properties and safety.

[0005] Nisin is a cationic antimicrobial peptide with multiple activities and is non-toxic to humans. It is often used as a food additive to control food spoilage microorganisms. Pandit et al. (2017) used lemongrass extract to synthesize AgNPs, which were coupled with Nisin to form nano-antimicrobial peptide conjugates, and finally the resulting nanoparticles were combined with agar to prepare a film. The experimental results showed that the nanosilver-antimicrobial peptide conjugate agar composite film showed higher antibacterial activity than the AgNPs agar film and the pure agar film. These studies show that the use of silver nanoparticles as nanofillers combined with a biopolymer matrix to prepare a film can improve the performance of the film and make it have the potential to become an antimicrobial packaging for food.

[0006] However, most of the current agar antibacterial film preparation processes are not in situ preparation, the preparation process is complicated, and the use of toxic reagents in the preparation of agar composite films with nano-antimicrobial peptide conjugates is still difficult to avoid; in addition, the antibacterial efficiency and mechanical properties of the existing in situ prepared agar antibacterial films are also poor, which is difficult to meet the requirements of the food industry. Summary of the invention

[0007] Based on this, the present invention proposes a method for in situ synthesis of nanosilver-antimicrobial peptide conjugate agar antibacterial film, with agar as the film-forming matrix and nanosilver-antimicrobial peptide conjugate as the nanofiller. No toxic reagents are required in the whole preparation process, and the prepared agar antibacterial film has high mechanical properties and antibacterial efficiency.

[0008] Part I:

[0009] An in-situ preparation method of a nanosilver-antimicrobial peptide conjugate agar antibacterial film comprises the following steps:

[0010] dissolving pectin in an alkaline solution to obtain a pectin alkaline solution;

[0011] adding nisin and silver salt to the pectin alkaline solution, and mixing to obtain a colloidal solution;

[0012] Agar and an emulsifier are added to the colloidal solution, and the nanosilver-antimicrobial peptide conjugate agar antibacterial film is obtained after mixing.

[0013] The present invention uses biological macromolecular pectin as a reducing agent and stabilizer. Pectin molecules contain a large number of carboxyl groups, which will be ionized under alkaline conditions to form carboxylic anions, increasing the reducing ability of pectin molecules, which is conducive to reducing silver ions into silver atoms, thereby promoting the synthesis of nanosilver; in addition, alkaline conditions can also increase the reaction rate, improve the dispersibility of nanosilver, and improve the stability of nanosilver. The colloidal solution contains nanosilver-antimicrobial peptide conjugates.

[0014] Emulsifiers can significantly reduce the interfacial tension between the colloidal solution and agar, allowing them to mix better and form a stable emulsion, thereby improving the stability of the film and improving the ductility, plasticity and tensile strength of the agar antibacterial film.

[0015] As a preferred solution, the method further comprises the following steps: after adding the agar and the emulsifier to the colloidal solution, the colloidal solution is assisted to dissolve for 10 to 30 minutes at a speed of 500 to 1000 r / min at 70 to 100° C. Through the assisted dissolution, the nanosilver-antimicrobial peptide conjugate is further dispersed, the uniformity and mechanical properties of the agar antimicrobial film are improved, and shrinkage, curling, and even holes after film formation are avoided.

[0016] As a preferred solution, the method further comprises the following steps: after auxiliary dissolution, a mixed solution is obtained, the mixed solution is placed in a mold, and dried at 30 to 60° C. for 1 to 4 hours to obtain a nanosilver-antimicrobial peptide conjugate agar antibacterial film. Drying can accelerate film formation, and the use of a mold can prepare a fixed shape of the agar antibacterial film.

[0017] As a preferred embodiment, the alkaline solution includes a sodium hydroxide alkaline solution; the sodium hydroxide content in the pectin alkaline solution is 0.5-5 g / L, and the pectin content is 0.1-0.5 g / L. If the sodium hydroxide content is too high or too low, the pH value of the entire reaction system will be higher or lower than the optimal reaction condition, which will result in the pectin being unable to be fully degraded, resulting in insufficient reducing ability and the inability to convert Ag into + If the pectin content is too low, the reducing ability will be insufficient and Ag cannot be reduced to nano silver. + Reduced to nano silver; if the pectin content is too high, it may affect the pectin and Ag + The contact between the nanoparticles and the nanostructured particles will reduce the reduction efficiency and may also affect the size and distribution uniformity of the nanosilver after reduction.

[0018] As a preferred embodiment, in the colloidal solution, the mass ratio of pectin:nisin:silver salt is 1:0.2-0.4:1-2, wherein the silver salt is calculated as silver ions.

[0019] If the pectin content is too low, the reducing ability will be insufficient and Ag cannot be + Reduced to nano silver; if the pectin content is too high, it may affect the pectin and Ag +The contact between the silver ions and the pectin will reduce the reduction efficiency and may also affect the size and distribution uniformity of the reduced nanosilver. If there is too little nisin, the product will not have a good synergistic effect and will not be able to improve the antibacterial ability. On the contrary, if it is added in excess and dissolved in the system, it will hinder the pectin from reducing silver ions, and the ideal product cannot be obtained. If there is too little silver salt, the product will be too little, the pectin cannot be fully utilized, and the preparation efficiency is low; the more silver salt, the less it can be fully reduced, and the final system becomes a mixed solution of nanosilver and silver nitrate, and silver nitrate has certain toxic effects on the environment and the human body. In summary, it is crucial to choose the right ratio of the three, which is related to the efficient preparation of the product, the improvement of antibacterial ability and the avoidance of environmental pollution.

[0020] As a preferred embodiment, the amount of agar added is 1-4wt.% of the colloidal solution, and the amount of emulsifier added is 1-1.6wt.% of the colloidal solution. An agar mass fraction lower than 4% is conducive to film formation. If the mass fraction is too low, the mechanical strength of the film after drying is too low and the basic requirements of the film cannot be met. However, although the film with too high a mass fraction has high mechanical strength, its ductility and transparency will be greatly reduced. Selecting an agar addition amount of 1-4% by mass fraction can take into account mechanical strength, water solubility and other properties. The concentration range of the emulsifier can be selected in the same way.

[0021] As a preferred embodiment, the emulsifier includes glycerol. Glycerol as an emulsifier has the following advantages:

[0022] (1) Glycerol can remain stable under different environmental conditions and is not easy to evaporate or decompose, which helps the long-term stability of the emulsification system.

[0023] (2) Glycerol is considered non-toxic and harmless even if ingested in large quantities.

[0024] (3) Glycerol can be biodegraded and is environmentally friendly.

[0025] (4) Glycerol does not react with substances in the colloidal solution and is easy to mix with other ingredients, so the formula can be easily adjusted to meet specific emulsification requirements. In addition, compared with some other emulsifiers, glycerol has a lower cost and better cost-effectiveness.

[0026] As a preferred embodiment, the silver salt includes at least one of silver nitrate, silver chloride and silver sulfate. Silver nitrate is more preferred. Silver nitrate has low cost, good stability and good economic benefits in large-scale production. In addition, the reaction conditions of silver nitrate, such as temperature, time, solvent, protective agent type, reducing agent dosage, etc., are easy to control, and it is easy to prepare high-purity and well-dispersed nano-silver. The preparation process is not easy to generate by-products, reducing environmental pollution. Silver nitrate can also be combined with different reducing agents and protective agents to prepare nano-silver with different morphologies, such as spheres, triangles, hexagons, etc.

[0027] Beneficial effects of the present invention:

[0028] 1. A new method for preparing Nisin@pectin-based AgNPs agar antibacterial film in situ by using biomacromolecule pectin as a reducing agent and stabilizer, silver nitrate as a silver source, and combining nisin (Nisin) to greenly synthesize Nisin@pectin-based AgNPs (NP-AgNPs) colloidal solution and agar. The process is simple and efficient, avoiding the use of toxic reagents, and has good biocompatibility and biodegradability.

[0029] 2. The nanosilver-antimicrobial peptide conjugate agar antibacterial film prepared by the present invention has multiple biological activities and good antibacterial properties, ultraviolet blocking properties and mechanical properties. Therefore, it can be used as an efficient and low-cost food antibacterial packaging material, has broad application prospects, and provides a scientific basis for the research and development of food antibacterial packaging.

[0030] Part II:

[0031] A food package comprises the nano silver-antimicrobial peptide conjugate agar antimicrobial film prepared in the first part. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 The finished product (a) and UV-visible absorption spectrum (b) of the nanosilver-antimicrobial peptide conjugate agar antimicrobial film in Example 1;

[0033] Figure 2 The scanning electron microscope surface image (a) and cross-sectional image (b) of the nanosilver-antimicrobial peptide conjugate agar antibacterial film in Example 1;

[0034] Figure 3 This is the antibacterial performance test chart. DETAILED DESCRIPTION

[0035] An in-situ preparation method of a nanosilver-antimicrobial peptide conjugate agar antibacterial film comprises the following steps:

[0036] dissolving pectin in an alkaline solution to obtain a pectin alkaline solution;

[0037] adding nisin and silver salt to the pectin alkaline solution, and mixing to obtain a colloidal solution;

[0038] Agar and an emulsifier are added to the colloidal solution, which is sealed, and assisted to dissolve for 10 to 30 minutes at 70 to 100° C. and a rotation speed of 500 to 1000 r / min to obtain a mixed solution, which is placed in a mold and dried at 30 to 60° C. for 1 to 4 hours to obtain a nanosilver-antimicrobial peptide conjugate agar antibacterial film.

[0039] The alkaline solution comprises a sodium hydroxide alkaline solution; the sodium hydroxide content in the pectin alkaline solution is 0.5-5 g / L, and the pectin content is 0.1-0.5 g / L.

[0040] In the colloidal solution, the mass ratio of pectin:nisin:silver salt is 1:0.2-0.4:1-2, wherein the silver salt is calculated as silver ions.

[0041] The amount of agar added is 1-4 wt.% of the colloidal solution, and the amount of emulsifier added is 1-1.6 wt.% of the colloidal solution.

[0042] The emulsifier includes glycerin, and the silver salt includes silver nitrate.

[0043] Example 1

[0044] An in-situ preparation method of a nanosilver-antimicrobial peptide conjugate agar antibacterial film comprises the following steps:

[0045] (1) Dissolve 0.2 g of pectin in 100 ml of 1 g / L sodium hydroxide alkaline solution to obtain a pectin alkaline solution.

[0046] (2) Add 50 mg of nisin and 0.425 g of silver nitrate to the pectin alkaline solution, and mix to obtain a colloidal solution.

[0047] (3) Add 2 g of agar and 1.2 g of glycerol to the colloidal solution, seal the solution, and place it in a constant temperature magnetic stirring water bath at 100° C. and 500 r / min to assist dissolution for 20 min to obtain a mixed solution. After complete dissolution, the solution has no obvious color change.

[0048] (4) placing the mixed solution in a mold, cooling and solidifying it, placing it in an electric heated air drying oven, drying it at 50° C. for 3 h, and demolding it to obtain a nanosilver-antimicrobial peptide conjugate agar antibacterial film.

[0049] Figure 1The finished product of the nanosilver-antimicrobial peptide conjugate agar antibacterial film in Example 1 (a) and the UV-visible absorption spectrum at 300-600nm (b); it can be seen that the film is smooth, uniform and transparent after drying and demoulding, and an absorption peak appears at 494nm, indicating the presence of silver nanoparticles in the film.

[0050] Figure 2 The scanning electron microscope surface image (a) and cross-sectional image (b) of the nanosilver-antimicrobial peptide conjugate agar antibacterial film in Example 1; as shown in the figure, silver nanoparticles are evenly distributed on the surface of the nanosilver-antimicrobial peptide conjugate agar antibacterial film, and the cross-section is tight and wrinkle-free, indicating that the film is successfully prepared in situ with excellent effect.

[0051] Example 2

[0052] This embodiment is substantially the same as embodiment 1, except that in step (3), the masses of agar added to the colloidal solution are 1 g, 3 g, and 4 g, respectively.

[0053] Example 3

[0054] This embodiment is substantially the same as embodiment 1, except that in step (3), the masses of glycerol added to the colloidal solution are 1.0 g, 1.4 g, and 1.6 g, respectively.

[0055] Example 4

[0056] This embodiment is substantially the same as embodiment 1, except that in step (3), the water bath temperatures are 70° C., 80° C., and 90° C., respectively.

[0057] Example 5

[0058] This embodiment is substantially the same as embodiment 1, except that in step (4), the drying temperatures are 30° C., 40° C., and 60° C., respectively.

[0059] Example 6

[0060] This embodiment is substantially the same as embodiment 1, except that in step (4), the drying time is 1 h, 2 h, and 4 h, respectively.

[0061] Comparative Example 1

[0062] The pectin base solution was replaced by pure water, and the other steps were exactly the same as in Example 1 to prepare a pure agar film.

[0063] Comparative Example 2

[0064] In step (2), no nisin was added, and the other steps were exactly the same as those in Example 1 to prepare the nanosilver agar antibacterial film.

[0065] Comparative Example 3

[0066] Antibacterial performance test

[0067] The nanosilver-antimicrobial peptide conjugate agar antibacterial film prepared in Example 1 and the nanosilver agar antibacterial film prepared in Comparative Example 2 were tested for minimum inhibitory concentration (MIC), and the specific steps were as follows:

[0068] (1) Weigh 10 mg of the nanosilver-antibacterial conjugate agar antibacterial film liquid prepared in Example 1 and the nanosilver agar antibacterial film prepared in Comparative Example 2 respectively for use.

[0069] (2) The two antibacterial films in (1) were mixed with 0.75 mL of bacterial suspension (10 7 CFU / mL), and 18 mL of sterilized LB broth were mixed evenly.

[0070] At this time, the concentration of nanosilver-antimicrobial peptide conjugate agar antibacterial film liquid and nanosilver agar antibacterial film in the whole system accounted for 533.3 μg / mL.

[0071] (3) (2) was diluted in a gradient manner to obtain mixed systems with membrane liquid concentrations of 266.7, 133.3, 66.7, 33.3, and 16.7 μg / mL, respectively. Escherichia coli and Staphylococcus aureus were added to the mixed systems of each concentration, and all samples were incubated at 37°C for 24 hours.

[0072] (4) Samples were taken every 3 hours on a clean bench, and the OD value at 600 nm was measured by an ELISA reader to draw a growth curve.

[0073] The experimental results show that the nanosilver agar antibacterial film prepared in Comparative Example 2 can completely inhibit the growth of Escherichia coli and Staphylococcus aureus at a concentration of 533.3 μg / mL, and the nanosilver-antimicrobial peptide conjugate agar antibacterial film prepared in Example 1 can completely inhibit the growth of Escherichia coli and Staphylococcus aureus at concentrations of 16.7 μg / mL and 33.3 μg / mL, respectively, which are 32 and 16 times higher than the antibacterial performance of Comparative Example 2, respectively.

[0074] Comparative Example 4

[0075] Antibacterial performance test

[0076] The nanosilver-antimicrobial peptide conjugate agar antibacterial film (0.05 g) prepared in Example 1 was used to test the antibacterial performance of Escherichia coli and Staphylococcus aureus by coating the plate.

[0077] First, the agar antibacterial film was sterilized by ultraviolet irradiation for 1 hour, and then immersed in 500 μL of bacterial suspension (10 7CFU / mL) in a glass bottle and gently shake (200r / min) at 37°C for 1 hour. Then take out 150μL of bacterial suspension and inoculate it on an agar plate. After incubation at 37°C for 18 hours, the bacterial reproduction is detected. The experiment without agar antibacterial film treatment is used as a control.

[0078] The results are as follows Figure 3 As shown, the growth status of Escherichia coli (a) and Staphylococcus aureus (c) without treatment with the nanosilver-antimicrobial peptide conjugate agar antibacterial film in Example 1, and the growth status of Escherichia coli (b) and Staphylococcus aureus (d) after treatment with the nanosilver-antimicrobial peptide conjugate agar antibacterial film in Example 1. It can be seen that after treatment with the agar antibacterial film, the growth of bacteria is significantly inhibited, indicating that the prepared nanosilver-antimicrobial peptide conjugate agar antibacterial film has good antibacterial ability.

[0079] Comparative Example 5

[0080] Various performance tests

[0081] The nanosilver-antimicrobial peptide conjugate agar antibacterial film prepared in Example 1 and the pure agar film prepared in Comparative Example 1 were tested, wherein the test standards were GB1037-1988 and GB1040-79.

[0082] Table 1 Performance tests of Example 1 and Comparative Example 1

[0083]

[0084] The results are shown in Table 1. The thicknesses of the nanosilver-antimicrobial peptide conjugate agar antibacterial film obtained in Example 1 and the pure agar film obtained in Comparative Example 1 are 26.5 μm and 12.9 μm, respectively. This is due to the addition of nanofillers in the nanosilver-antimicrobial peptide conjugate agar antibacterial film, which increases the total solid content.

[0085] The tensile strengths of the nanosilver-antimicrobial peptide conjugate agar antibacterial film obtained in Example 1 and the pure agar film obtained in Comparative Example 1 were 16.1 MPa and 16.8 MPa, respectively. The tensile strength of the agar antibacterial film was slightly reduced, which may be because the embedding of the nanosilver-antimicrobial peptide conjugate in the agar antibacterial film reduced the interactions within and between the agar molecular chains, resulting in the discontinuity of the polymer network.

[0086] The elongation at break of the nanosilver-antimicrobial peptide conjugate agar antibacterial film obtained in Example 1 and the pure agar film obtained in Comparative Example 1 were 47.2% and 13.1%, respectively. The elongation at break of the agar antibacterial film increased significantly, which is due to the presence of nanofillers causing plastic deformation of the matrix, such as shear bands and cavitation growth.

[0087] The water solubility of the nanosilver-antimicrobial peptide conjugate agar antibacterial film obtained in Example 1 and the pure agar film obtained in Comparative Example 1 are 16.4% and 13.5%, respectively. Water solubility can reflect the degradable properties of the agar film to a certain extent. The water solubility of the nanosilver-antimicrobial peptide conjugate agar antibacterial film is higher because the presence of nanofillers in the agar antibacterial film makes the surface rougher, and the rough surface has more pores, which leads to more water absorption during immersion.

[0088] The transmittance (T280) of the nanosilver-antimicrobial peptide conjugate agar antibacterial film obtained in Example 1 and the pure agar film obtained in Comparative Example 1 at 280nm is 3.1% and 56.6%, respectively. The enhanced ultraviolet blocking performance is due to the uniform dispersion of the nanofiller in the agar antibacterial film. These particles can effectively scatter and absorb ultraviolet rays, thereby improving the ultraviolet blocking ability of the agar antibacterial film. In addition, the nanofiller has a small size, a large specific surface area, and a larger contact area with ultraviolet rays, which also helps to absorb and scatter ultraviolet rays.

[0089] The above-mentioned embodiments only express several implementation methods of the present invention, and the description is relatively specific and detailed, but it cannot be understood as limiting the scope of the invention patent. For ordinary technicians in this field, several modifications and improvements can be made without departing from the concept of the present invention, which all belong to the protection scope of the present invention.

Claims

1. A method for preparing an in-situ nanosilver-antimicrobial peptide conjugate agar antibacterial film, characterized in that: The steps include: dissolving pectin in an alkaline solution to obtain a pectin alkaline solution; adding nisin and silver salt to the pectin alkaline solution, and mixing to obtain a colloidal solution; Agar and an emulsifier are added to the colloidal solution, and the nanosilver-antimicrobial peptide conjugate agar antibacterial film is obtained after mixing.

2. An in-situ preparation method of a nanosilver-antimicrobial peptide conjugate agar antibacterial film according to claim 1, characterized in that: The following steps are also included: After the agar and the emulsifier are added to the colloidal solution and mixed, the mixture is assisted to dissolve at 70-100° C. and a rotation speed of 500-1000 r / min for 10-30 minutes.

3. The in-situ preparation method of a nano-silver-antimicrobial peptide conjugate agar antibacterial film according to claim 2, characterized in that: The following steps are also included: After auxiliary dissolution, a mixed solution is obtained, and the mixed solution is placed in a mold and dried at 30 to 60° C. for 1 to 4 hours to obtain a nanosilver-antimicrobial peptide conjugate agar antibacterial film.

4. The in-situ preparation method of a nanosilver-antimicrobial peptide conjugate agar antibacterial film according to claim 1, characterized in that: The sodium hydroxide content in the pectin alkali solution is 0.5-5 g / L, and the pectin content is 0.1-0.5 g / L.

5. The in-situ preparation method of a nano-silver-antimicrobial peptide conjugate agar antibacterial film according to claim 4, characterized in that: In the colloidal solution, the mass ratio of pectin:nisin:silver salt is 1:0.2-0.4:1-2, wherein the silver salt is calculated as silver ions.

6. According to the in-situ preparation method of the nanosilver-antimicrobial peptide conjugate agar antibacterial film as claimed in claim 1, the amount of agar added is 1-4wt.% of the colloidal solution, and the amount of emulsifier added is 1-1.6wt.% of the colloidal solution.

7. According to the in-situ preparation method of a nanosilver-antimicrobial peptide conjugate agar antibacterial film according to claim 1, the emulsifier comprises glycerol.

8. According to the in-situ preparation method of a nano-silver-antimicrobial peptide conjugate agar antibacterial film according to claim 1, the silver salt comprises at least one of silver nitrate, silver chloride and silver sulfate.

9. A food package, characterized in that: The invention comprises the nano silver-antimicrobial peptide conjugate agar antimicrobial film prepared according to any one of claims 1 to 8.

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