Natural bacteriostatic material and preparation method and application thereof

Abstract

The application relates to the field of food preservation, in particular to a natural bacteriostatic material and a preparation method and application thereof, and a preparation method of the natural bacteriostatic material, which comprises the following steps: mixing natural gelatin and dextran with deionized water, stirring, and freeze-drying to obtain a gelatin-dextran mixture powder, wherein the mass ratio of the natural gelatin to the dextran is 1-2:1-4, and the molecular weight of the dextran is 150-200 kDa; performing a glycosylation reaction on the gelatin-dextran mixture powder to obtain a gelatin-dextran conjugate; dissolving the gelatin-dextran conjugate in a buffer solution to obtain an emulsifier; and homogenizing edible essential oil, the emulsifier and deionized water, and performing ultrasonic emulsification to obtain the natural bacteriostatic material. The preparation method is simple, the conditions are simple, the prepared natural bacteriostatic material has good stability, and the shelf life of food can be prolonged.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of food preservation, in particular to a natural bacteriostatic material and a preparation method and application thereof. BACKGROUND

[0002] Food preservatives generally use substances with bacteriostatic effect, such as edible essential oils. However, the high volatility, thermal instability and obvious odor of edible essential oils limit their application in food preservation. Therefore, the prior art mixes emulsifiers or gelatin with edible essential oils to form emulsions, which improves the preservation effect of food preservatives to a certain extent. However, the stability of emulsifiers is poor, resulting in a short preservation period of food. Gelatin, a natural macromolecular compound derived from animal collagen, is widely used in the food industry due to its low price, good emulsifying properties, water retention, film forming properties and gel properties. Due to the amphiphilic nature of gelatin, it can spontaneously migrate to the interface layer of gas and water or oil and water, forming a protein film with good elasticity and high viscosity and extensive expansion. Through the orientation of hydrophobic groups with oil or gas phase and the orientation of hydrophilic groups with water phase, the surface tension between different phases is reduced, thereby playing an emulsifying role. However, during the preparation process, the triple helix structure of collagen protein is damaged to varying degrees due to the action of acid, alkali and high temperature, resulting in poor stability of gelatin when used in embedding and delivery systems. Moreover, gelatin tends to form large droplets when stabilizing emulsions, resulting in poor stability of emulsions and failing to fully meet the processing needs of food. Therefore, suitable modification techniques are needed to expand the application of gelatin in food preservation.

[0003] To expand the application of gelatin in the food industry, gelatin is usually modified to improve its function. Gelatin chemical modification is usually carried out through phosphorylation modification, phenolic modification, glycosylation modification, cationic modification, protein modification, etc. By the action of modification reagents on the carboxyl, hydroxyl, amino, carbonyl and sulfhydryl groups of gelatin, the molecular structure, steric hindrance, surface charge of gelatin are changed to improve the morphology, emulsifying properties of gelatin. In particular, glycosylation modification can help gelatin generate more stable properties and be applied in emulsions. Glycosylation reaction is simple to operate and has obvious effect without the need for adding other catalysts. The early stage of glycosylation reaction covalently combines proteins and reducing sugars to form glycoproteins. This process can be catalyzed by dry or wet method. Dry reaction is milder than wet reaction, the reaction conditions are more controllable, and the denaturation degree of protein is lower, which can make the glycosylation reaction proceed slowly.

[0004] Dextran is a high molecular weight neutral polysaccharide, which has the characteristics of low sweetness, low heat and the like. In addition, dextran also has the functions of enhancing human immunity, resisting radiation and inhibiting bacteria. Polysaccharide is a carbohydrate with different polymerization degrees composed of 10 or more than 10 monosaccharides, and has a large molecular weight. The molecular structure contains multiple hydroxyl structures, and the spatial folding is complex, and the main chain and side chain often exist at the same time. Polysaccharide has a certain degree of emulsifying ability, and has good hydrophilicity and high viscosity. The monosaccharide constituting the polysaccharide is connected by glycosidic bond (alpha-1, 4-glycosidic bond, beta-1, 4-glycosidic bond and alpha-1, 6-glycosidic bond), which can be connected into a straight chain or a branched chain. The structure determines the unique processing characteristics and rheological properties of polysaccharide, which can increase the viscosity and stability of the beverage and improve the water holding capacity and texture of the food. However, the glycosylation reaction activity of monosaccharide is strong, and by-products are easily produced by glycosylation reaction, and browning is obvious, so it is not suitable for emulsion.

[0005] The prior art scheme discloses a technical scheme for glycosylation modification of fish skin gelatin by using chitosan to improve the surface activity of fish skin gelatin. However, the wet glycosylation reaction is relatively violent, the denaturation degree of protein is too high, the grafting degree is low, the modification effect of gelatin is not good, and then the stability of the modified gelatin in food preservation is affected. SUMMARY

[0006] Therefore, the technical problem to be solved by the present application is to overcome the defects of unstable emulsification of the modified gelatin in the prior art, poor preservation effect of the prepared antibacterial material, and to provide a preparation method and application of a natural antibacterial material, improve the modification efficiency of gelatin, simplify the preparation steps of the natural antibacterial material, and form a natural antibacterial material with stable emulsifying properties.

[0007] In one aspect, the present application provides a preparation method of a natural antibacterial material, comprising the following steps: S1, mixing natural gelatin and dextran with deionized water, stirring, freeze-drying to obtain a gelatin-dextran mixture powder, wherein the mass ratio of natural gelatin to dextran is 1-2:1-4, and the molecular weight of dextran is 150-200 kDa; S2, glycosylation reaction of the gelatin-dextran mixture powder to obtain a gelatin-dextran conjugate; S3, dissolving the gelatin-dextran conjugate in a buffer to obtain an emulsifier; S4, homogenizing the edible essential oil, the emulsifier and deionized water, and ultrasonic emulsification to obtain a natural antibacterial material.

[0008] In the step S1, the stirring mode is at least one of magnetic stirring, water bath stirring or high-speed stirring;

[0009] Optionally, the stirring rate is 100-150 rpm, and the stirring time is 2-4 h;

[0010] Optionally, the mass concentration of the gelatin and dextran mixture in the mixed solution is 5-8%.

[0011] In the step S1, the freeze-drying temperature is -35℃ to -70℃, the pressure is 0.2-0.8MPa, and the freeze-drying time is 24-28h.

[0012] In the step S1, after freeze-drying, the gelatin dextran mixture powder is 100-120 mesh.

[0013] In the step S2, in the glycosylation reaction, the relative humidity is 60-80%, the temperature is 70-90℃, and the holding time is 4-7 days.

[0014] In the step S3, the mass ratio of the gelatin dextran conjugate to the buffer is 0.8-1:40-50.

[0015] Optionally, the buffer is a phosphate buffer. The pH value of the phosphate buffer is 7, and the concentration of the phosphate buffer is 1%(w / w).

[0016] In the step S4, the addition amount of the edible essential oil is 4%-7% by volume of the natural antibacterial material, the addition amount of the emulsifier is 33%-36%, and the addition amount of the deionized water is 50%-60%.

[0017] In the step S4, the rotation speed of the homogenization treatment is 20000-24000rpm, and the homogenization time is 1-2min.

[0018] Optionally, the ultrasonic power of the ultrasonic emulsification is 190-210W, and the ultrasonic treatment time is 5-10min.

[0019] In another aspect, the application also provides a natural antibacterial material prepared by the preparation method of the natural antibacterial material.

[0020] The natural antibacterial material prepared by the preparation method of the natural antibacterial material can be applied to food preservation, and the specific application steps include: applying the natural antibacterial material to the surface of the food to be preserved, and placing it at room temperature for 2-4h, and sterile packaging.

[0021] Optionally, the mass ratio of the natural antibacterial material to the food to be preserved is 0.2-0.4:1.

[0022] The food to be preserved is at least one of meat, eggs, milk or seafood.

[0023] The technical scheme of the application has the following advantages:

[0024] 1. The preparation method of natural bacteriostatic material provided by the present application comprises the following steps: mixing natural gelatin and dextran with deionized water, stirring, and freeze-drying to obtain a gelatin-dextran mixture powder, wherein the mass ratio of natural gelatin to dextran is 1-2:1-4, and the molecular weight of dextran is 150-200 kDa; performing a glycosylation reaction on the gelatin-dextran mixture powder to obtain a gelatin-dextran conjugate; dissolving the gelatin-dextran conjugate in a buffer solution to obtain an emulsifier; and homogenizing edible essential oil, the emulsifier and deionized water, and ultrasonic emulsification to obtain the natural bacteriostatic material. The present application utilizes gelatin with amphiphilic properties, which is modified by dextran with good hydrophilicity and stability through glycosylation to simultaneously have the properties of proteins and saccharides, so that the solubility and hydrophilicity of gelatin can be effectively increased. Meanwhile, the modified gelatin is mixed with a buffer solution to prepare an emulsifier with good hydrophilicity and strong stability, by utilizing the property of gelatin that can spontaneously migrate to the interface layer of gas and water or oil and water. The edible essential oil is used as the oil phase core material of the natural bacteriostatic material, which is emulsified with the emulsifier formed by the modified gelatin and dextran to prepare a microcapsule nanoemulsion with strong stability, preservation and bacteriostasis.

[0025] In a colloidal solution, the presence of high molecules induces repulsive flocculation, which promotes colloidal aggregation and accelerates colloidal particle aggregation, which is often detrimental to the stability of the alternating system. With the increase of the concentration of high molecules, the repulsive flocculation ability is enhanced, which can further accelerate colloidal coalescence. When the concentration of high molecules is high to a certain extent, the thickening effect of high molecules plays a dominant role, slows down the coalescence of colloidal particles, and plays a role in stabilizing the colloidal system. Therefore, the present application limits the use of dextran with a molecular weight of 150-200 kDa and the mass ratio of natural gelatin to dextran in the preparation of the emulsifier is 1-2:1-4, so as to avoid excessive dextran content, increase the collision opportunity of gelatin and proteins, affect the grafting degree, and ensure that the grafting degree of the gelatin-dextran conjugate prepared remains within a certain range to improve the stability of the emulsifier. At the same time, the emulsifier prepared by the present application induces repulsive flocculation, which makes the emulsion droplets aggregate and quickly float up. The upper layer is milky white and the lower layer is transparent, and the boundary between the two layers is clear. The highly repulsive flocculation of the upper layer forms a network structure between the emulsion colloidal droplets and the droplets, which can form a natural bacteriostatic material with stable nanostructure.

[0026] The application adopts ultrasonic treatment of emulsifiers and edible essential oils, and when the concentration of the macromolecule is high enough and the revolution radius of the macromolecule is smaller than that of the colloidal particles of the dispersed phase, the gelatin macromolecule-induced repulsive flocculation produces the network structure between the colloidal particles, so that the emulsifiers and edible essential oils form a stable network structure at the nanometer level, and the natural bacteriostatic material reaches the nanometer level through ultrasonic emulsification, the smaller the particle size of the natural bacteriostatic material, the more uniform the particle size distribution, and the more stable the natural bacteriostatic material, and the ultrasonic wave makes the immiscible liquid in the system be crushed and uniformly dispersed, forms an emulsion with the surrounding liquid, improves the concentration of the dispersed phase of the bacteriostatic material, and further improves the stability of the natural bacteriostatic material.

[0027] 2. The preparation method of the natural bacteriostatic material provided by the application, the glycosylation reaction is a dry glycosylation, and the relative humidity is 60-80%, the temperature is 70-90 DEG C, and the time is 4-7 days. The application modifies the gelatin by adopting the dry glycosylation reaction, and limits the environmental humidity and temperature of the dry glycosylation reaction, so that the spatial structure of the gelatin becomes loose, the hydrophobic groups in the gelatin can play a role, the hydrophobic ability of the gelatin is enhanced, the balance of the hydrophilic and hydrophobic of the gelatin is improved, and the interfacial tension of the gelatin dextran conjugate formed is reduced, so that the emulsification performance of the emulsifier is better improved.

[0028] The application adopts the dry glycosylation reaction, can promote the spontaneous combination between the dextran and the gelatin, is safe and reliable, and is simple and convenient to operate.

[0029] 3. The natural bacteriostatic material provided by the application, the particle size is 35-60 nm. The natural bacteriostatic material prepared by the application is a nanoemulsion, has small particle size, can be stored for a long time, and has good stability.

[0030] 4. The preparation method of the natural bacteriostatic material provided by the application is simple, high in efficiency, good in emulsification and stability of the prepared bacteriostatic material, and uses edible essential oils as the oil phase of the emulsifier, so that other impurities and odors are less introduced, and the food pickling, seasoning and preservation effects are combined, the food shelf life is prolonged, and the food has good flavor. Meanwhile, the natural bacteriostatic material prepared by the application utilizes the thermal reversibility of the gelatin, is applied on the surface of the food to be preserved, especially meat, the natural bacteriostatic material covering the surface of the meat effectively slows down the water loss of the meat during the storage period, reduces the volatility, controls the release of the edible essential oils, prolongs the storage time of the active ingredients, improves the stability of the edible essential oils, and further prolongs the shelf life of the food. BRIEF DESCRIPTION OF DRAWINGS

[0031] In order to more clearly illustrate the technical solutions in the specific embodiments of the present application or the prior art, the drawings required to be used in the specific embodiments or prior art description will be briefly introduced below. Obviously, the drawings in the following description are some embodiments of the present application, and other drawings can also be obtained by those skilled in the art without creative effort on the basis of these drawings.

[0032] Figure 1 is the process flow chart of the preparation method of the natural bacteriostatic material in Embodiment 1 of the present application. DETAILED DESCRIPTION

[0033] The following examples are provided to better further understand the present application and are not limited to the best mode, and do not limit the content and protection scope of the present application. Any product identical or similar to the present application obtained by the inspiration of the present application or the combination of the present application with other prior art features falls within the protection scope of the present application.

[0034] The specific experimental steps or conditions not mentioned in the examples can be performed according to the conventional experimental steps described in the literature in the art or the operation or conditions. The reagents or instruments not mentioned by the manufacturer are conventional reagent products that can be obtained by market purchase.

[0035] The edible essential oil used in the embodiments of the present application is used as the oil phase of the natural bacteriostatic material and has a certain flavoring effect. The edible essential oil can be purchased on the market, and the brand and channel are not limited. It can also be extracted from raw materials, and the extraction method is not limited.

[0036] Embodiment 1

[0037] The present embodiment provides a preparation method of a natural bacteriostatic material, and the specific steps and methods are as follows:

[0038] According to the process flow chart shown in Figure 1 10 g of tilapia skin gelatin and 10 g of dextran with a molecular weight of 150 kDa are mixed, deionized water is added, and the mixture is magnetically stirred at 20℃ and a rotation speed of 100 rpm for 3 h. A mixed solution with a mass concentration of 6% of the gelatin and dextran mixture is prepared, and the solution is frozen in a refrigerator at-40℃ for 24 h. The frozen solution is placed in a freeze dryer for freeze-drying, wherein the cold trap temperature is-40℃, the pressure is 0.22 MPa, and the freeze-drying time is 24 h. The freeze-dried mixture is sieved through a 120 mesh sieve and dried in a desiccator, wherein the desiccator is filled with saturated KBr solution at the bottom;

[0039] The gelatin dextran mixture powder is placed in a humidity 80%, temperature 70℃ artificial climate box for 6 days to obtain a gelatin dextran conjugate, and an emulsifier is configured according to a mass ratio of the gelatin dextran conjugate to the phosphate buffer of 1:50, wherein the concentration of the phosphate buffer is 1% (w / w), and the pH value of the phosphate buffer is 7;

[0040] The anise essential oil 0.4ml, star anise essential oil 0.5ml, pepper essential oil 5ml, chili essential oil 20ml, ginger essential oil 10ml, onion oil 2ml, lemon essential oil 0.8ml, litsea cubeba essential oil 0.5ml, and myrtle essential oil 2ml are mixed to prepare the edible essential oil;

[0041] The 7ml edible essential oil, 33ml emulsifier and 60ml deionized water are homogenized at a rotation speed of 20000rpm for 2min, and the mixture after homogenization is ultrasonically treated using an ultrasonic instrument with a frequency of 25kHz, an invasion depth of 25mm and a power of 200W for 5min to obtain the natural antibacterial material;

[0042] The natural antibacterial material is detected to have a particle size of 40-55nm.

[0043] The natural antibacterial material prepared in this embodiment is applied to the preservation of fresh tilapia, and the specific steps are as follows:

[0044] The fresh tilapia is washed and drained, and the natural antibacterial material is applied to the surface of the tilapia, and the tilapia is placed at 15℃ for 4h, wherein the mass ratio of the natural antibacterial material to the tilapia is 0.4:1, and the tilapia is vacuum-sterile packaged to obtain the preserved food, and the packaging bag is a food-grade high-temperature-resistant aluminum packaging bag.

[0045] Example 2

[0046] The present embodiment provides a preparation method of a natural antibacterial material, and the specific steps and methods are as follows:

[0047] 10g pigskin gelatin and 20g dextran with a molecular weight of 200kDa are mixed, deionized water is added, and the mixture is stirred in a water bath at 20℃ and a rotation speed of 150rpm for 2h to prepare a solution with a mass concentration of 5% of the gelatin and dextran mixture, and the solution is frozen in a refrigerator at-40℃ for 24h, and then the frozen solution is placed in a freeze dryer for freeze-drying, wherein the cold trap temperature is-35℃, the pressure is 0.8MPa, and the freeze-drying time is 28h, and then the freeze-dried mixture is sieved through a 120-mesh sieve and dried in a desiccator, wherein the desiccator is filled with saturated KBr solution at the bottom;

[0048] The gelatin dextran mixture powder is placed in a artificial climate chamber with relative humidity of 60% and temperature of 90℃ for 4 days to obtain a gelatin dextran conjugate, and an emulsifier is prepared according to a mass ratio of the gelatin dextran conjugate to the phosphate buffer of 0.8:50, wherein the concentration of the phosphate buffer is 1% (w / w), and the pH value of the phosphate buffer is 7;

[0049] The edible essential oil is prepared by mixing 1ml of geranium essential oil, 0.5ml of cassia essential oil, 0.5ml of star anise essential oil, 0.4ml of anise essential oil, 0.2ml of lemon essential oil, 0.3ml of pepper essential oil, 0.2ml of osmanthus essential oil and 0.5ml of Sichuan pepper essential oil;

[0050] The 4ml of edible essential oil is homogenized with 36ml of emulsifier and 50ml of deionized water at a rotation speed of 24000rpm for 1min, and then treated with an ultrasonic instrument at a frequency of 25kHz for 10min at an invasion depth of 25mm and a power of 190W to obtain a natural antibacterial material.

[0051] The natural antibacterial material has a particle size of 45-50nm.

[0052] The natural antibacterial material prepared in the embodiment is applied to the preservation of fresh five-flower meat, and the specific steps are as follows:

[0053] The fresh five-flower meat is washed and drained, and then the natural antibacterial material is applied to the surface of the five-flower meat at 20℃, and the mixture is placed at room temperature for 2h, wherein the mass ratio of the natural antibacterial material to the tilapia is 0.2:1, and the five-flower meat is vacuum-sterile packaged to obtain a preserved food, and the packaging bag is a food-grade high-temperature-resistant aluminum packaging bag.

[0054] Example 3

[0055] The embodiment provides a preparation method of a natural antibacterial material, and the specific steps and methods are as follows:

[0056] 10g of pigskin gelatin and 40g of dextran with a molecular weight of 170kDa are mixed, deionized water is added, and the mixture is magnetically stirred at 20℃ and a rotation speed of 120rpm for 4h to prepare a solution with a mass concentration of 8% of the gelatin and dextran mixture, and the solution is frozen in a refrigerator at-40℃ for 24h, and then placed in a freeze dryer for freeze-drying, wherein the cold trap temperature is-70℃, the pressure is 0.2MPa, and the freeze-drying time is 26h, and then the freeze-dried mixture is sieved through a 100-mesh sieve and dried in a desiccator, wherein the desiccator is filled with saturated KBr solution at the bottom;

[0057] The gelatin dextran mixture powder is placed in an artificial climate box with a relative humidity of 70% and a temperature of 80 DEG C for 7 days to obtain a gelatin dextran conjugate, and an emulsifier is prepared according to a mass ratio of the gelatin dextran conjugate to the phosphate buffer of 0.8:40, wherein the concentration of the phosphate buffer is 1% w / w;

[0058] The edible essential oil is prepared by mixing 15ml of ginger essential oil, 0.2ml of leaf essential oil, 1ml of star anise essential oil, 0.2ml of leaf essential oil, 0.5ml of fennel essential oil, 0.5ml of litsea cubeba essential oil, 0.3ml of pepper essential oil, 0.2ml of dried tangerine peel essential oil, 0.2ml of prickly ash essential oil and 0.5ml of chili essential oil;

[0059] The 6ml of edible essential oil is homogenized with 36ml of emulsifier and 58ml of deionized water at a rotation speed of 22000rpm for 1.5min, and the homogenized mixture is ultrasonically treated by using an ultrasonic instrument with a frequency of 25kHz, an invasion depth of 25mm and a power of 210W for 8min to obtain the natural bacteriostatic material;

[0060] The natural bacteriostatic material has a particle size of 45-60nm.

[0061] The natural bacteriostatic material prepared in the embodiment is applied to preservation of fresh pork knuckles, and the specific steps are as follows:

[0062] The fresh pork knuckles are washed and drained, the natural bacteriostatic material is applied to the surface of the pork knuckles, the mass ratio of the natural bacteriostatic material to the pork knuckles is 0.3:1, and meanwhile, 7g of edible salt, 3g of granulated sugar, 3g of cooking oil, 3g of Sprite and 2g of old soybean extract are additionally applied to the surface of each kilogram of the pork knuckles, and the pork knuckles are placed at 18 DEG C for 3h, and then the pork knuckles are vacuum-sterilized packaged to obtain the preserved food, and the packaging bag is a food-grade high-temperature-resistant aluminum packaging bag.

[0063] Embodiment 4

[0064] The embodiment of the present application provides a preparation method of a natural bacteriostatic material, and the specific steps and methods are as follows:

[0065] 20g of tilapia skin gelatin and 10g of dextran with a molecular weight of 150kDa are mixed, deionized water is added, and the solution with a mass concentration of 7% of gelatin and dextran is prepared by high-speed stirring at 20 DEG C and a rotation speed of 140rpm for 2.5h, and then the solution is frozen in a refrigerator at-40 DEG C for 24h, and then the frozen solution is placed in a freeze dryer for freeze-drying, wherein the cold trap temperature is-40 DEG C, the pressure is 0.60MPa, and the freeze-drying time is 25h, and then the freeze-dried mixture is sieved through a 120-mesh sieve and dried in a desiccator, to obtain a gelatin dextran mixture powder, wherein the desiccator is filled with saturated KBr solution at the bottom;

[0066] The gelatin dextran mixture powder is placed in an artificial climate chamber with relative humidity of 75% and temperature of 85℃ for 5 days to obtain a gelatin dextran conjugate, and the emulsifier is prepared according to a mass ratio of the gelatin dextran conjugate to the phosphate buffer of 1:40, wherein the concentration of the phosphate buffer is 1 w / w.

[0067] The anise essential oil 0.4ml, star anise essential oil 0.5ml, pepper essential oil 5ml, chili essential oil 20ml, ginger essential oil 10ml, onion oil 2ml, lemon essential oil 0.8ml, litsea cubeba essential oil 0.5ml, and myrtle essential oil 2ml are mixed to prepare the edible essential oil.

[0068] The 7ml edible essential oil, 36ml emulsifier, and 57ml deionized water are homogenized at a rotation speed of 23000rpm for 2min, and the mixture after homogenization is treated by ultrasonic waves with an ultrasonic instrument of 25kHz, an invasion depth of 25mm, and a power of 200W for 7min to obtain the natural antibacterial material.

[0069] The natural antibacterial material is detected to have a particle size of 35-45nm.

[0070] The natural antibacterial material prepared in the embodiment is applied to the preservation of fresh beef, and the specific steps are as follows:

[0071] The fresh beef is washed and drained, and the natural antibacterial material is applied to the surface of the beef at 25℃ for 2h, wherein the mass ratio of the natural antibacterial material to the beef is 0.4:1, and the beef is vacuum-sterile packaged to obtain the preserved food, and the packaging bag is a food-grade high-temperature-resistant aluminum packaging bag.

[0072] Comparative Example 1

[0073] The comparative example provides a preparation method of an antibacterial material, and the specific steps and parameters are the same as those of the embodiment 1 of the application, and the difference lies in that in the preparation of the gelatin dextran mixture, 10g of tilapia skin gelatin is mixed with 10g of dextran with a molecular weight of 70kDa.

[0074] The natural antibacterial material prepared in the comparative example is applied to the preservation of fresh tilapia, and the specific steps are as in the embodiment 1.

[0075] Comparative Example 2

[0076] The comparative example provides a preparation method of an antibacterial material, and the specific steps and parameters are the same as those of the embodiment 1 of the application, and the difference lies in that in the preparation of the gelatin dextran mixture, 10g of tilapia skin gelatin is mixed with 10g of dextran with a molecular weight of 300kDa.

[0077] The natural antibacterial material prepared in the comparative example is applied to the preservation of fresh tilapia, and the specific steps are as in the embodiment 1.

[0078] Comparative Example 3

[0079] The present comparative example provides a preparation method of the bacteriostatic material, the specific steps and parameters are the same as those of Example 1 of the present application, and the difference lies in that, in the preparation of the gelatin chitosan mixture, 10 g of chitosan is dissolved in dilute acetic acid with a concentration of 1%, and then the pH is adjusted to 7.0 with sodium hydroxide solution, and 10 g of tilapia fish skin gelatin is mixed with the chitosan solution.

[0080] The natural bacteriostatic material prepared in the present comparative example is applied to the preservation of fresh tilapia, and the specific steps are as in Example 1.

[0081] Comparative Example 4

[0082] The present comparative example provides a preparation method of the bacteriostatic material, the specific steps and parameters are the same as those of Example 1 of the present application, and the difference lies in that, in the preparation of the gelatin chitosan mixture, 10 g of chitosan is dissolved in dilute acetic acid with a concentration of 1%, and then the pH is adjusted to 7.0 with sodium hydroxide solution, and 10 g of tilapia fish skin gelatin is mixed with the chitosan solution.

[0083] The natural bacteriostatic material prepared in the present comparative example is applied to the preservation of fresh tilapia, and the specific steps are as in Example 1.

[0084] Comparative Example 5

[0085] The present comparative example provides a preparation method of the bacteriostatic material, the specific steps and parameters are the same as those of Example 1 of the present application, and the difference lies in that the gelatin dextran conjugate is prepared by wet Maillard reaction:

[0086] 10 g of tilapia fish skin gelatin is mixed with 10 g of dextran with a molecular weight of 150 kDa, deionized water is added, and the mixture is magnetically stirred at 20°C and a rotation speed of 100 rpm for 3 h. A mixed solution with a mass concentration of 6% of the gelatin and dextran mixture is prepared, the pH of the mixed solution is adjusted to 10, and then the mixed solution is placed in a water bath at 60°C for 100 min, followed by ice bath for 10 min, and then frozen at -20°C for 6-12 h, followed by freeze-drying, wherein the cold trap temperature is -40°C, the pressure is 0.22 MPa, and the freeze-drying time is 24 h. The freeze-dried mixture is sieved through a 120 mesh sieve and dried in a desiccator to obtain a gelatin dextran conjugate.

[0087] The remaining steps are the same as those of Example 1.

[0088] Comparative Example 6

[0089] The present comparative example provides a preparation method of the bacteriostatic material, the specific steps and parameters are the same as those of Example 1 of the present application, and the difference lies in that it does not contain edible essential oil.

[0090] Experimental Example 1

[0091] The grafting degree of the gelatin dextran conjugates prepared in Examples 1-3 and Comparative Examples 1-6 was detected, and the emulsifying activity index, emulsifying stability index of the natural bacteriostatic materials prepared in Examples 1-3 and Comparative Examples 1-6, and the stability of the natural bacteriostatic materials in salt ions and heat treatment were detected, and the detection results are shown in Table 1, and the detection methods are as follows:

[0092] The detection method of the grafting degree of the gelatin dextran conjugate: the o-phthaldehyde (OPA) reagent was prepared on site, and the preparation method was as follows: 40 mg of OPA, 1 ml of methanol, 25 ml of 0.1 M sodium tetraborate buffer (pH 9.3), 100 μL of β-mercaptoethanol and 2.5 ml of 20% (w / v) SDS were mixed thoroughly, and then deionized water was added to 50 ml. 4 ml of OPA reagent was added to 200 μL of 0.4% sample solution to be detected, and the mixture was incubated at 35°C for 2 minutes. The absorbance at 340 nm was measured by ultraviolet spectrophotometry. Under the same conditions, distilled water was used instead of the sample as a control group.

[0093] The grafting degree (GD) (%) = [(A0-At) / A0] x 100%, wherein A0 is the absorbance of the sample before reaction (0 h), and At is the absorbance of the sample after reaction (th).

[0094] The detection method of the emulsifying activity index and emulsifying stability index of the natural bacteriostatic material: 100 μL of the natural bacteriostatic material prepared in Examples 1-3 and Comparative Examples 1-6 was diluted 100 times with 0.1% (w / v) SDS solution, and the absorbance was measured at 500 nm.

[0095] The emulsifying activity index (EAI) is calculated as follows,

[0096] EAI (m 2 / g) = (2 x 2.303 x A0 x DF) / (c x θ x 10000), wherein DF is the dilution factor (the dilution multiple is reasonable when the absorbance measured at 500 nm by ultraviolet spectrophotometry after the solution is diluted with SDS is between 0.2 and 0.8, and in this experimental example, DF = 100), c is the protein concentration in the gelatin, θ is the proportion of the oil phase in the emulsion, and in this experimental example, the proportion of the oil phase in the emulsion is the ratio of the total volume of essential oil to the volume of the natural bacteriostatic material, A0 is the absorbance of the emulsion at 500 nm at 0 minutes, and the protein concentration in the gelatin is measured by the BCA detection method;

[0097] The emulsifying stability index (ESI) is calculated as follows,

[0098] ESI (minimum) = A0 / (A0-A10) x 10, wherein A0 and A10 are the absorbance of the emulsion at 500 nm after 0 min and 10 min;

[0099] Method for detecting the stability of the natural bacteriostatic material in salt ions: the natural bacteriostatic material prepared in Examples 1-3 and Comparative Examples 1-6 and NaCl solutions with different concentrations (0, 100, 300 and 500 mmol / L) were mixed in a ratio of 1:1 (v / v) respectively, and then placed at room temperature for 2 h. The particle size of the emulsion was measured by a Zeta potential analyzer to evaluate the effect of ion concentration on the stability of the emulsion.

[0100] Method for detecting the stability of the natural bacteriostatic material in heat treatment: 10 mL of nanoemulsion was taken from the natural bacteriostatic material prepared in Examples 1-3 and Comparative Examples 1-6 and sealed in a test tube, and then placed in an incubator at 25℃, 37℃ and 72℃ respectively for 24 hours. The particle size of the emulsion was measured to evaluate the effect of temperature change on the stability of the emulsion. The particle size of the natural bacteriostatic material with different salt ion concentrations was determined by a Zeta potential analyzer.

[0101] Experimental Example 2

[0102] The volatile base nitrogen and total bacterial count of the fresh-keeping food obtained from Examples 1-3 and Comparative Examples 1-6 were detected on the 5th, 10th, 15th, 20th and 25th day of storage, and the detection results are shown in Table 2, and the detection methods are as follows,

[0103] Method for detecting volatile base nitrogen: according to GB5009.228-2016 "Determination of volatile base nitrogen in food", automatic Kjeldahl nitrogen determination method was selected to determine the volatile base nitrogen of meat sample

[0104] Method for detecting total bacterial count: the sample was determined according to GB4789.2-2022 "Food microbiology test determination of total bacterial count", and the plate pouring method was used for counting determination.

[0105] Table 1 Results of grafting degree of gelatin dextran conjugate, emulsifying property and stability of natural bacteriostatic material prepared in Examples 1-3 and Comparative Examples 1-6

[0106]

[0107] As shown in Table 1, the natural bacteriostatic material prepared in Examples 1-3 is superior to the natural bacteriostatic material prepared in Comparative Examples 1-5 in terms of grafting degree, emulsion stability, salt ion stability and heat treatment stability. The emulsion activity index of the natural bacteriostatic material prepared in Example 1 is increased by 5.33%, 6.71%, 6.64%, 8.54% and 8.2% respectively compared with Comparative Examples 1-5, indicating that the content of dextran plays a certain role in the emulsifying property of the natural bacteriostatic material. In the examples of the present application, the mass ratio of gelatin to dextran is 1-2:1-4, and the molecular weight of dextran is 150-200 kDa. In particular, when the mass ratio of gelatin to dextran is 1:1, the natural bacteriostatic material prepared has the best emulsifying performance. The emulsion activity and emulsion stability of the natural bacteriostatic material prepared in Example 1 and Comparative Example 4 are compared. The dextran used in the present application to modify the gelatin has a better modification effect than chitosan, and the natural bacteriostatic material prepared has better emulsion stability and activity. However, Comparative Example 6 does not add edible essential oil, so it cannot form an emulsion, but only a mixture of Maillard reaction gel and sugar, and cannot form oil-in-water microcapsules, so other parameters cannot be detected.

[0108] The smaller the size of the nanoemulsion droplets and the more uniform the size distribution, the more stable the nanoemulsion. The size of the emulsion droplets is affected by various factors. Different emulsification methods provide different energy, which significantly affects the size and distribution of the emulsion droplets. Generally, the greater the energy provided, the smaller the size of the emulsion droplets, the more uniform the energy, and the more uniform the size distribution of the emulsion droplets. The stability of the natural bacteriostatic material prepared in Examples 1-3 and Comparative Examples 1-5 is detected under different salt concentrations and without temperature treatment. In the heat treatment detection, when the temperature is 40-70℃ for 30 min, the natural bacteriostatic material prepared in Examples 1-3 has good stability, while the natural bacteriostatic material prepared in Comparative Examples 1-5 has a layering phenomenon at 50-70℃ for 30 min. With the increase of salt ion concentration, the size of the natural bacteriostatic material prepared in Examples 1-3 decreases first and then increases, and the size of the natural bacteriostatic material prepared in Comparative Examples 1-5 is larger than that of Examples 1-3, proving that the ultrasonic emulsification method used in the examples of the present application can significantly improve the stability of the natural bacteriostatic material. Therefore, the natural bacteriostatic material prepared by the method of the examples of the present application can reduce the layering phenomenon of the natural bacteriostatic material under certain salt concentration and temperature treatment conditions, and improve the stability of the natural bacteriostatic material.

[0109] The grafting degree of the gelatin dextran conjugate prepared in Examples 1-3 is 20-30%, and the grafting degree of Comparative Examples 1-5 is 10-20%. It can be seen that the dextran with smaller molecular weight has higher reactivity due to shorter polysaccharide chain and smaller steric hindrance, and is more easily penetrated into the folding structure of the protein. Therefore, under the same conditions, the Maillard reaction can occur more quickly to generate intermediate products and colored substances.

[0110] Table 2 Preservation effect of the natural antibacterial material prepared in Examples 1-3 and Comparative Examples 1-6

[0111]

[0112] It can be seen from Table 2 that the volatile basic nitrogen of the natural antibacterial material prepared in Examples 1-3 is still less than 30 mg / 100 g when the preservation time is 20 days, and the total number of colonies is still less than 6.5 lg(cfu / g) when the preservation time is 10 days. The volatile basic nitrogen of the natural antibacterial material prepared in Comparative Examples 1-6 is greater than 30 mg / 100 g when the preservation time is 20 days, and the total number of colonies is greater than 6.5 lg(cfu / g) when the preservation time is 10 days. In particular, Comparative Example 6, due to the absence of edible essential oil, the antibacterial material lacks an oil phase, and the raw material cannot be prepared into a microcapsule nanosolution, and cannot be homogenized and ultrasonically emulsified, resulting in the inability to form a stable nanoemulsion, which causes poor antibacterial effect. It can be seen that the natural antibacterial material prepared in the present application has good stability and can prolong the preservation period of food.

[0113] Obviously, the above examples are only examples for clearly illustrating, but not limit the embodiments. For those skilled in the art, other different forms of changes or variations can be made on the basis of the above description. Here, all the embodiments need not and cannot be exhausted. The obvious changes or variations derived therefrom are still within the protection scope of the present application.

Claims

1. A method for preparing a natural antibacterial material, characterized in that, Includes the following steps: S1, mix natural gelatin and dextran with deionized water, stir, and freeze-dry to obtain gelatin-dextran mixture powder, wherein the mass ratio of natural gelatin to dextran is 1-2:1-4, and the molecular weight of dextran is 150-200kDa; S2, Glycosylation reaction of gelatin-dextran mixture powder to obtain gelatin-dextran conjugate. During the glycosylation reaction, the relative humidity is 60-80%, the temperature is 70-90℃, and the reaction is maintained for 4-7 days. S3, dissolve the gelatin dextran conjugate in buffer solution to obtain an emulsifier; S4. The edible essential oil, the above-mentioned emulsifier and deionized water are homogenized and ultrasonically emulsified to obtain a natural antibacterial material.

2. The method for preparing the natural antibacterial material according to claim 1, characterized in that, The stirring method in step S1 is at least one of magnetic stirring, water bath stirring, or high-speed stirring; The stirring speed is 100-150 rpm, and the stirring time is 2-4 hours. The mass concentration of the gelatin and dextran mixture in the mixed solution is 5-8%.

3. The method for preparing the natural antibacterial material according to claim 1, characterized in that, In step S1, after freeze-drying, the gelatin-dextran mixture powder is 100-120 mesh.

4. The method for preparing the natural antibacterial material according to claim 1, characterized in that, In step S3, the mass ratio of the gelatin dextran conjugate to the buffer solution is 0.8-1:40-50; The buffer solution is a phosphate buffer; And / or, in step S4, the amount of edible essential oil added is 4%-7% based on the volume of the natural antibacterial material, the amount of emulsifier added is 33%-36%, and the amount of deionized water added is 50%-60%.

5. The method for preparing the natural antibacterial material according to claim 4, characterized in that, In step S4, the homogenization speed is 20000-24000 rpm, and the homogenization time is 1-2 min. The ultrasonic power for ultrasonic emulsification is 190-210W, and the ultrasonic treatment time is 5-10 minutes.

6. A natural antibacterial material prepared by the method of any one of claims 1-5, characterized in that, The particle size of the natural antibacterial material is 35-60 nm.

7. The application of a natural antibacterial material prepared by the method of any one of claims 1-5 or the natural antibacterial material of claim 6 in food preservation.

8. The application of the natural antibacterial material according to claim 7 in food preservation, characterized in that, Apply natural antibacterial materials to the surface of the food to be preserved, let it stand at 15-25℃ for 2-4 hours, and then aseptically package it. The mass ratio of natural antibacterial materials to food to be preserved is 0.2-0.4:1.

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