Soybean protein isolate / gelatin packaging film containing eucalyptus leaf essential oil nanoemulsion

Abstract

The invention discloses a soybean protein isolate / gelatin packaging film containing eucalyptus leaf essential oil nanoemulsion. The method comprises the following steps: firstly, embedding eucalyptus leaf essential oil by using a low-temperature ultrahigh-pressure continuous flow cell crusher to prepare eucalyptus leaf essential oil nanoemulsion, adding the eucalyptus leaf essential oil nanoemulsion into a soybean protein isolate / gelatin / glycerol mixed solution, magnetically stirring, and drying in a constant-temperature and constant-humidity box to obtain the eucalyptus leaf essential oil nanoemulsion / soybean protein isolate / gelatin packaging film. The eucalyptus leaf essential oil nanoemulsion is applied to the soybean protein isolate / gelatin composite membrane, the antibacterial and antioxidant effects of the eucalyptus leaf essential oil nanoemulsion are exerted, the eucalyptus leaf essential oil serves as a cross-linking agent, the internal structure of the membrane is more compact, the tensile strength and the elongation at break of the composite membrane are improved, the water vapor permeability and the oxygen permeability are reduced, and the service life of the membrane is prolonged. The synergistic advantage of the eucalyptus leaf essential oil is exerted, and the components have good compatibility, so that the membrane has better thermal stability. Cherry fresh-keeping experiments prove that the composite film has an obvious fresh-keeping effect on cherries.

Description

Technical Field

[0001] The invention belongs to the technical field of nano packaging films, and in particular relates to a soybean protein isolate / gelatin packaging film containing eucalyptus essential oil nanoemulsion. Background Art

[0002] Food packaging film acts as a protective barrier for food, which can protect food from adverse external environment. It can also extend the shelf life of food by reducing environmental pollution, thus making it better for storage and transportation.

[0003] Natural biopolymers such as proteins, polysaccharides and lipids are widely used in the field of food packaging because of their wide sources, safety, non-toxicity, good film-forming properties and degradability. Among them, protein is a polymer formed by multiple amino acids connected by peptide bonds. Its unique structure makes it have good film-forming properties. Soy protein isolate (SPI) is a plant protein with a compact and orderly structure, which lays a solid foundation for the formation of films with good gas barrier properties, but poor mechanical properties and high brittleness limit the application of soy protein isolate films. Gelatin (GE) is an animal protein with good flexibility. Studies have shown that plant protein and animal protein have good compatibility, and the miscibility of the two can make the protein molecules fully stretch. Soy protein isolate is filled between gelatin chains, so that the linear macromolecules of carboxyl and amine groups contained in gelatin interact with the amine and carboxyl groups on soy protein isolate to form a certain branched structure. The branches are entangled with each other, which increases the viscosity of the film-forming solution, changes the flexibility of the protein molecular chain, and increases the force between the molecular chains, especially the hydrogen bond effect. Therefore, the packaging film prepared with the above two proteins as the film matrix improves the mechanical properties while maintaining good film-forming properties, which can effectively improve the film performance.

[0004] However, although soy protein isolate and gelatin packaging films showed good performance, they lacked antioxidant and antimicrobial activities. Summary of the invention

[0005] In view of the above problems, the present invention provides a soy protein isolate / gelatin packaging film containing eucalyptus essential oil nanoemulsion. The present invention first uses a low-temperature ultra-high pressure continuous flow cell disruptor to embed eucalyptus essential oil to prepare eucalyptus essential oil nanoemulsion, and adds the eucalyptus essential oil nanoemulsion to the soy protein isolate / gelatin / glycerol mixed solution, and obtains the eucalyptus essential oil nanoemulsion / soy protein isolate / gelatin packaging film after magnetic stirring and drying in a constant temperature and humidity chamber. The present invention uses eucalyptus essential oil nanoemulsion for soy protein isolate / gelatin composite film, which not only exerts its own antibacterial and antioxidant effects, but also eucalyptus essential oil as a cross-linking agent, making the internal structure of the film more compact, improving the tensile strength and elongation at break of the composite film, and also reducing the water vapor permeability (WVP) and oxygen permeability, giving play to the synergistic advantages of eucalyptus essential oil, and each component has good compatibility, so that the film has better thermal stability. Cherry preservation experiments have proved that the composite film of the present invention has an obvious preservation effect on cherries.

[0006] To achieve the above-mentioned purpose, the present invention adopts the following technical scheme to achieve the purpose: a method for preparing a soy protein isolate / gelatin packaging film containing eucalyptus essential oil nanoemulsion, characterized in that:

[0007] S1: Preparation of eucalyptus essential oil nanoemulsion

[0008] The eucalyptus essential oil, an emulsifier and water are mixed to prepare a crude emulsion; and then a low temperature ultra-high pressure continuous flow cell disruptor is used to homogenize the mixture at a pressure of 1000-1200 bar (preferably 1092 bar) to prepare a eucalyptus essential oil nanoemulsion;

[0009] S2: Preparation of soy protein isolate and gelatin mixture

[0010] Prepare a soy protein isolate and gelatin mixed solution containing soy protein isolate, gelatin, glycerin and water; the mass ratio of the soy protein isolate to gelatin is 1:3.5-4.5;

[0011] S3: Preparation of soy protein isolate / gelatin packaging film containing eucalyptus essential oil nanoemulsion

[0012] The eucalyptus essential oil nanoemulsion is slowly added to the soy protein isolate and gelatin mixed solution and stirred evenly; then the mixture is poured into a plastic plate and dried in a constant temperature and humidity chamber to obtain a soy protein isolate / gelatin packaging film containing the eucalyptus essential oil nanoemulsion.

[0013] Preferably, in step S1, the content of eucalyptus essential oil is 1.5-1.8%, preferably 1.6%, of the total mass of the crude emulsion; preferably, the emulsifier is Tween-80; and the amount of Tween-80 is 40-60% of the total mass of the eucalyptus essential oil.

[0014] Preferably, the specific steps of step S2 are as follows:

[0015] (1) preparing a soy protein isolate solution with a concentration of 4.5-5.5%, adjusting the pH of the solution to 9.5-10.5 with alkali solution, and then heating and stirring in a water bath at 85-95° C. for 50-70 minutes;

[0016] (2) preparing a gelatin solution with a concentration of 4.5-5.5%, hydrating for 25-35 min, and then heating and stirring in a water bath at 45-55°C for 25-35 min;

[0017] (3) Mix the soy protein isolate solution and the gelatin solution (in a volume ratio of 1:3.5-4.5), add 20-30% glycerol (the content of glycerol is relative to the amount of dry matter), heat and stir in a water bath at 45-55° C. for 55-65 minutes, and obtain a soy protein isolate and gelatin mixed solution.

[0018] Further, the addition amount of the eucalyptus essential oil nanoemulsion is 10-30% (relative to the volume of the soy protein isolate and gelatin mixed solution). When the addition amount of the eucalyptus essential oil nanoemulsion is 20%, the thickness of the prepared eucalyptus essential oil nanoemulsion / soy protein isolate / gelatin film is 0.138±0.0068mm, and the tensile strength and elongation at break of the soy protein isolate / gelatin film are the largest. Compared with the soy protein isolate / gelatin film without the addition of eucalyptus essential oil nanoemulsion, the soy protein isolate / gelatin film containing the eucalyptus essential oil nanoemulsion has a continuously decreasing light transmittance, a continuously increasing opacity, a decreasing water vapor permeability and an increasing oxygen permeability at first, and wherein the water vapor permeability and the oxygen permeability are the lowest when the addition amount of the eucalyptus essential oil is 20%. When the addition amount of the eucalyptus essential oil is 20%, it has the best ultraviolet light blocking performance, can reduce lipid oxidation in food and extend the shelf life of food. As the amount of eucalyptus essential oil nanoemulsion added to the soy protein isolate / gelatin increases, the antibacterial zone continues to increase and the antibacterial performance continues to increase.

[0019] Eucalyptus essential oil is the secretion of the oil gland cells of the leaves of Eucalyptus genus of the Myrtaceae family. Its main components are terpenes and alcohol compounds. It is a natural antioxidant and antibacterial agent with broad-spectrum antioxidant and antibacterial properties. Adding eucalyptus essential oil to packaging film can effectively inhibit the growth of microorganisms and extend the shelf life of food. However, eucalyptus essential oil is volatile, easy to oxidize, and has poor stability. At the same time, the compatibility of essential oil directly added to the packaging film is poor, resulting in uneven film. Therefore, the application of essential oil as a natural food additive in food is subject to a certain degree of restriction.

[0020] Nanoemulsion refers to a colloidal dispersion system in which a liquid phase is dispersed in a second phase in the form of droplets. The particle size of nanoemulsion is between 10-200nm, which has the advantage of small particle size. At the same time, essential oil nanoemulsion can not only improve the stability of essential oils and reduce their pungent odor, but also control their release rate and prolong the action time. Therefore, adding essential oil nanoemulsion to the packaging film can give the film antioxidant and antibacterial properties while maintaining the film performance.

[0021] The present invention uses a low-temperature ultra-high-pressure continuous flow cell disruptor to embed eucalyptus essential oil to prepare a eucalyptus essential oil nanoemulsion. Compared with the traditional high-pressure homogenization technology, the technology uses high-pressure energy to release the sample instantly through a slit, and under the action of collision, cavitation and shearing effects, the eucalyptus essential oil mixed system is homogenized, dispersed and emulsified, and finally a stable eucalyptus essential oil nanoemulsion is formed. At the same time, the implementation of the process is carried out in a low-temperature circulating water bath, which can effectively maintain the activity of the effective components of the eucalyptus essential oil to the greatest extent.

[0022] Compared with the prior art, the technical effects of the present invention are:

[0023] 1. Eucalyptus essential oil is made into nanoemulsion for soybean protein isolate / gelatin composite film, which solves the problems of eucalyptus essential oil being easy to volatilize, easy to oxidize, poor stability and poor compatibility. In the process of preparing nanoemulsion, a low-temperature ultra-high-pressure continuous flow cell disruptor is used to homogenize, disperse and emulsify the eucalyptus essential oil mixed system, and finally form a stable eucalyptus essential oil nanoemulsion. At the same time, the implementation of this process is carried out in a low-temperature circulating water bath, which can effectively maintain the activity of the effective ingredients of eucalyptus essential oil to the greatest extent.

[0024] 2. The present invention uses eucalyptus essential oil nanoemulsion for soy protein isolate / gelatin composite film, which not only exerts its own antibacterial and antioxidant effects, but also eucalyptus essential oil nanoemulsion as a cross-linking agent, synergistically enhances the performance of the composite film with the soy protein isolate / gelatin composite film, so that the performance of the composite film is better, which is specifically reflected in the following aspects:

[0025] (1) The present invention adds an appropriate amount of eucalyptus essential oil, which serves as a cross-linking agent, to make the internal structure of the soy protein isolate / gelatin composite film more compact, thereby improving the tensile strength and elongation at break of the composite film and enhancing the physical properties of the film; since the internal structure of the film is more compact, the water vapor permeability (WVP) and oxygen permeability are also reduced; and the synergistic advantages of eucalyptus essential oil are brought into play;

[0026] (2) The composite film to which an appropriate amount of eucalyptus essential oil nanoemulsion is added according to the present invention has spherical structures on its surface and cross section, and at the same time, the composite film is uniform and dense as a whole, has no pores, and the eucalyptus essential oil nanoemulsion has good compatibility with the membrane matrix. At the same time, there is an intermolecular interaction between the eucalyptus essential oil nanoemulsion and the membrane matrix, so that the components have good compatibility, and the membrane has better thermal stability.

[0027] 3. The soy protein isolate / gelatin composite film containing eucalyptus essential oil nanoemulsion of the present invention reduces water vapor permeability and oxygen permeability, so that the packaging film has spontaneous gas-conditioning and moisture-barrier properties, and natural antibacterial substances (eucalyptus essential oil) are added to the film, and the two together give the film material excellent preservation effect. As a packaging material, it can contribute to the formation of high CO2 and low O2 in the packaging environment, reduce respiration, extend the storage period of fruits and vegetables, and reduce decay caused by microbial infection.

[0028] 4. Cherry preservation experiments have shown that the soy protein isolate / gelatin composite film containing eucalyptus essential oil nanoemulsion provided by the present invention can well maintain the sensory quality of cherries, delay the corruption and deterioration of cherries during storage, and maintain a high vitamin C (V C ), total phenols and flavonoids, maintaining a high hardness and low weight loss rate, extending the shelf life of cherries, and having a more obvious preservation effect on cherries.

[0029] 5. The soy protein isolate / gelatin composite film containing eucalyptus essential oil nanoemulsion provided by the present invention is mainly made of soy protein isolate, gelatin and eucalyptus essential oil, all of which have good biodegradable properties and can be effectively decomposed after being discarded, and are environmentally friendly materials. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 is the effect of different concentrations of eucalyptus essential oil nanoemulsion on the transmittance of the composite film;

[0031] Figure 2 are scanning electron microscope images of composite films of eucalyptus essential oil nanoemulsions of different concentrations; wherein A, B, C, D, and E are plan views of SPI-GE-0%, SPI-GE-10%, SPI-GE-20%, SPI-GE-30%, and SPI-GE-40%, and a, b, c, d, and e are cross-sectional views of SPI-GE-0%, SPI-GE-10%, SPI-GE-20%, SPI-GE-30%, and SPI-GE-40%;

[0032] Figure 3 is the Fourier transform infrared spectra of the composite films of eucalyptus essential oil nanoemulsions with different concentrations;

[0033] Figure 4 is the TGA graph of the composite film of eucalyptus essential oil nanoemulsion with different concentrations;

[0034] Figure 5 is the DTG graph of the composite film of eucalyptus essential oil nanoemulsion with different concentrations;

[0035] Figure 6 The effect of different concentrations of eucalyptus essential oil nanoemulsion on the antibacterial effect of composite film;

[0036] Figure 7 is the effect of different film treatments on the appearance of cherries;

[0037] Figure 8 The effects of different film treatments on cherry firmness (A), decay rate (B), weight loss rate (C) and respiration rate (D);

[0038] Fig. 9 The effects of different film treatments on the content of vitamin C (A), total phenols (B) and flavonoids (C) in cherries. DETAILED DESCRIPTION

[0039] The technical solution of the present invention is described in detail below with reference to the accompanying drawings and embodiments.

[0040] The materials and instruments used in the present invention are as follows:

[0041] 1 Materials

[0042] Soy protein isolate (protein content greater than 90%), Tween-80: Shanghai Yuanye Biotechnology Co., Ltd.; gelatin, glycerol, sodium acetate trihydrate, anhydrous sodium carbonate, dipotassium hydrogen phosphate: Sinopharm Chemical Reagent Co., Ltd.; eucalyptus essential oil (content 99%): Ji'an Guoguang Spice Co., Ltd., also known as eucalyptus oil, obtained by steam distillation of eucalyptus branches and leaves; all reagents were analytically pure; polyvinyl pyrrolidone: Shanghai MacLean Biochemical Technology Co., Ltd.; potato dextrose agar: Qingdao High-tech Industrial Park Haibo Biotechnology Co., Ltd.

[0043] 2 Instruments

[0044] Water bath: Changzhou Guoyu Instrument Manufacturing Co., Ltd.; Constant temperature and humidity chamber: Shanghai Yiheng Scientific Instrument Co., Ltd.; Low temperature and ultrahigh pressure continuous flow cell disruptor: Guangzhou Jueneng Nano Biotechnology Co., Ltd.; Scanning electron microscope: Quanta 250, FEI Company, OR, USA; Fourier transform infrared spectrometer: Nicolet 5700, Thermo Fisher Scientific, MA, USA; Comprehensive thermal analyzer: Netzsch STA449C, Freistaat, Bayern, Germany; UV spectrophotometer: UV-2550, Shimadzu International Trading Co., Ltd., Shimadzu, Japan; composite film tensile machine: Jinan Languang Electromechanical Technology Co., Ltd.; precision analytical balance; desktop high-speed refrigerated centrifuge: Changsha High-tech Industrial Development Zone Xiangyi Centrifuge Instrument Co., Ltd.; vertical high-pressure steam sterilizer: Shanghai Shen'an Medical Equipment Factory; handheld refractometer: Hangzhou Delixi Group Co., Ltd.; texture analyzer: Shanghai Baosheng Industrial Development Co., Ltd.; portable O2 / CO2 gas analyzer: PBI-Dansensor, NJ, USA.

[0045] 3 Data processing

[0046] All experiments were repeated three times, and Duncan test and one-way analysis of variance were used. The results are expressed as mean ± standard deviation, and p < 0.05 was considered statistically significant. SPSS Statistics 25.0 (SPSS Inc., Chicago, IL, USA) was used for statistical analysis, and Origin 2021 software was used to fit and visualize the data.

[0047] Embodiment 1:

[0048] 1. Preparation method of soy protein isolate / gelatin composite film containing eucalyptus essential oil nanoemulsion and film thickness, mechanical properties, light transmittance, opacity, water vapor permeability and oxygen permeability.

[0049] 1. Preparation of eucalyptus essential oil nanoemulsion

[0050] 0.48 g of eucalyptus essential oil, 0.24 g of Tween-80 and 29.28 g of water were mixed to prepare a crude emulsion (the oil content was equivalent to 1.6% of the total mass, and the amount of Tween-80 was 1 / 2 of the oil), and then a low-temperature ultra-high pressure continuous flow cell disruptor was used to homogenize three times in a low-temperature circulating water bath of 4°C-6°C and a pressure of 1092 bar to prepare a nanoemulsion.

[0051] 2. Preparation of eucalyptus essential oil nanoemulsion soybean protein isolate gelatin composite film

[0052] (1) preparing a 5% soy protein isolate solution, adjusting the solution pH to 10 with 1 mol / L NaOH, and then heating and stirring in a 90° C. water bath for 60 min;

[0053] (2) Prepare a 5% gelatin solution, hydrate for 30 min, and then heat and stir in a 50°C water bath for 30 min;

[0054] (3) Mix the soy protein isolate solution and the gelatin solution (volume ratio of 1:4), add 25% glycerol (the content of glycerol is relative to the amount of dry matter), heat and stir in a 50°C water bath for 60 min; slowly add 0%, 10%, 20%, 30%, and 40% eucalyptus essential oil nanoemulsion (the content of nanoemulsion is v / v relative to the total volume of the membrane solution) into the membrane solution and stir for 30 min;

[0055] (4) Finally, pour 150 mL of the membrane solution into a plastic plate (22.5 × 22.5 cm 2 ) and placed in a constant temperature and humidity chamber at 45°C for drying for 12 h. The prepared composite film was peeled off the plate, placed in a sealed bag, and stored in the dark at room temperature for subsequent performance testing.

[0056] 3. Determination of composite film thickness and mechanical properties

[0057] The thickness of the composite film was measured with a digital micrometer with an accuracy of 0.001 mm. Three points were randomly marked on the surface of the composite film and their thickness was measured. The data were recorded and the average value was calculated. The tensile strength (TS) and elongation at break (EB) of the composite film were measured using a composite film tensile tester (MED-01, Jinan Languang Electromechanical Technology Co., China). The shape of the composite film was required to be a strip with a width of 15 mm and a length of 40 mm.

[0058] The results are shown in Table 1.

[0059] Table 1 Thickness, tensile strength (TS) and elongation at break (EB) of composite films

[0060]

[0061] Note: Values ​​are expressed as mean ± standard deviation. Different lowercase letters in the same column indicate statistically significant differences (p<0.05).

[0062] The thickness of the composite film is shown in Table 1. As the amount of eucalyptus essential oil nanoemulsion increases, the thickness of the composite film gradually increases. The increase in the thickness of the composite film is proportional to the increase in the volume concentration of the nanoemulsion, which may be due to the addition of eucalyptus essential oil nanoemulsion increasing the amount of solid matter in the composite film liquid.

[0063] Mechanical properties are a key factor in evaluating food packaging materials. Food packaging materials are required to have good mechanical properties in order to ensure the safety and integrity of food during transportation and storage. The mechanical properties (tensile strength and elongation at break) of the composite film are shown in Table 1. With the increase of eucalyptus essential oil nanoemulsion, the tensile strength and elongation at break of the composite film both show a trend of first increasing and then decreasing. When the addition amount is 20%, the composite film has the maximum tensile strength and elongation at break, which indicates that the eucalyptus essential oil nanoemulsion is evenly distributed in the composite film, making the molecules arranged more closely, resulting in a tighter composite film, and at this time it has the best mechanical properties. The reason for the subsequent decline may be that the stronger polymer chain-chain interaction is replaced by the weaker polymer-emulsion interaction, thereby weakening the continuity of the composite film matrix. In summary, the appropriate addition of eucalyptus essential oil nanoemulsion can effectively enhance the interaction between molecules and improve the mechanical properties of the composite film.

[0064] 4. Determination of water vapor transmission rate of composite films

[0065] The composite film was used to seal a glass bottle (5 cm in diameter) containing anhydrous calcium chloride (3 g) to achieve RH1 = 0% and weighed; the glass bottle was then placed in a desiccator containing a saturated K2SO4 solution (RH2 = 97%) and placed in an incubator at 25°C. The mass of the glass bottle was recorded every 24 hours and calculated according to the formula:

[0066]

[0067] Where WVTR is the change in mass of the glass bottle divided by the transfer area (m 2 ) and time (d), P = 3.1690 kPa, X is the average thickness of the composite membrane (μm).

[0068] The results are shown in Table 2.

[0069] Table 2. Water vapor transmission rate (WVP), oxygen transmission rate (OP), and opacity of composite films

[0070]

[0071]

[0072] Note: Values ​​are expressed as mean ± standard deviation. Different lowercase letters in the same column indicate statistically significant differences (p<0.05).

[0073] The water vapor permeability (WVP) of the composite film represents the parameter of water transfer between the composite film and the food, and is one of the key indicators for measuring the performance of the composite film. The results of the water vapor permeability in the composite films with different contents of eucalyptus essential oil nanoemulsions are shown in Table 2. The water vapor permeability of the composite film first decreases and then increases with the increase of the concentration of eucalyptus essential oil nanoemulsion. The soy protein isolate / gelatin films with SPI-GE-10%, SPI-GE-20%, and SPI-GE-30% are all lower than the blank group, and the lowest water vapor permeability is achieved when the concentration of eucalyptus essential oil nanoemulsion is 20%. This may be because eucalyptus essential oil is used as a cross-linking agent, and the -COOH in its molecules can undergo esterification reaction with the -OH of soy protein isolate and gelatin, making the structure of the film more compact and hindering the permeation of water vapor. The water vapor permeability of the experimental group containing 40% eucalyptus essential oil nanoemulsion is higher than that of the blank group. This is because the excessive eucalyptus essential oil nanoemulsion causes the structure of the composite film to change, which in turn leads to the hydrolysis of soy protein isolate and gelatin, resulting in an increase in the water vapor permeability of the composite film. Therefore, the addition amount of eucalyptus essential oil has a greater impact on the performance of the composite film.

[0074] 5. Determination of composite membrane oxygen permeability (OP): First, place 3g of deoxidizer in a weighing bottle, cover the bottle mouth with active membrane, seal it with a rubber band, and then weigh it. Then place the weighing bottle in a dryer with a certain humidity (RH=90%) for 48h, and measure the mass of the weighing bottle again. The calculation formula of OP is as follows:

[0075]

[0076] Where M is the mass change of the weighing bottle (g), h is the thickness of the film (cm), and S is the effective area of ​​the film (cm 2 ), T is the time interval (s), and P is the oxygen partial pressure difference on both sides of the membrane (Pa).

[0077] The oxygen permeability of packaging materials has a significant impact on food quality. After food is stored in packaging materials with high oxygen permeability for a period of time, the rate of oxidation increases and it is more likely to spoil. As shown in Table 2, with the increase in the concentration of eucalyptus essential oil nanoemulsion, the oxygen permeability first decreases and then increases. The SPI-GE-20% composite film has the lowest oxygen permeability. This may be due to the addition of eucalyptus essential oil nanoemulsion, which hinders the oxygen diffusion path and affects the interaction between soy protein isolate and gelatin molecules, increasing the number of intermolecular hydrogen bonds, thereby making the composite film tighter and reducing the oxygen permeability of the composite film. However, excessive eucalyptus essential oil nanoemulsion destroys the interaction of the composite film matrix, making it easier for oxygen molecules to penetrate, and ultimately leading to an increase in the oxygen permeability of the active film. In summary, an appropriate amount of eucalyptus essential oil nanoemulsion can reduce the oxygen permeability of the composite film.

[0078] 6. Determination of optical properties of composite films

[0079] The composite film was cut into a 1 cm × 4 cm rectangle and the absorbance of the composite film was measured at 600 nm. The opacity is equal to the ratio of absorbance to thickness. -1 ) is calculated according to the following equation:

[0080]

[0081] Where T is the thickness of the film, in mm.

[0082] Opacity is an important parameter of food packaging, which affects the acceptance of consumers. The opacity of the composite film is shown in Table 2. As the content of eucalyptus essential oil nanoemulsion increases, the opacity of the composite film increases continuously. When the concentration of eucalyptus essential oil is 40%, the composite film has the highest opacity. It may be due to the strong molecular binding between eucalyptus essential oil nanoemulsion and soy protein isolate and gelatin, which improves the opacity of the composite film.

[0083] The composite film was cut into a 1 cm × 4 cm rectangle and placed in a cuvette so that it was attached to the light side of the cuvette. The composite film was scanned in the wavelength range of 200-800 nm using an ultraviolet spectrophotometer (UV / Vis) to determine the transmittance of the composite film.

[0084] The transmittance of the composite film in the range of 200-800nm ​​is as follows Figure 1 As shown. In the visible light wavelength range (400-800nm), as the wavelength increases, the transmittance of the composite film is significantly lower than that of the blank control group, which may be related to the highly entangled network structure formed by the cross-linked soy protein isolate and gelatin by eucalyptus essential oil nanoemulsion, which hinders the transmission of light. In the ultraviolet light wavelength range (200-400nm), the transmittance of all cross-linked composite films is lower than that of the blank control group. This result shows that the addition of eucalyptus essential oil nanoemulsion gives the composite film excellent ultraviolet blocking properties, which can reduce lipid oxidation in food and extend the shelf life of food. Among them, in the range of 200-350nm, when the addition amount of eucalyptus essential oil nanoemulsion is 20%, the transmittance of the composite film is significantly lower than that of other groups, indicating that SPI-GE-20% has the best ultraviolet blocking properties.

[0085] 2. Scanning electron microscopy, Fourier transform infrared spectroscopy, thermal stability and antibacterial properties of soy protein isolate gelatin composite films containing eucalyptus essential oil nanoemulsion

[0086] 1. Microscopic morphology of eucalyptus essential oil nanoemulsion composite film

[0087] The composite film was cut into a 0.5 cm × 1.0 cm rectangle and adhered to the sample stage with a conductive adhesive. After the sample was subsequently sprayed with gold, the surface and cross-section of the prepared composite film were analyzed by scanning electron microscopy.

[0088] Scanning electron microscopy can directly reflect the microstructural characteristics of the composite film. The morphological characteristics of the surface and cross-section of the composite film with different addition amounts of eucalyptus essential oil nanoemulsion are as follows: Figure 2 As shown. The surface and cross section of the soy protein isolate / gelatin composite membrane are uniform, dense and free of pores, indicating that the two matrices have good compatibility. The addition of eucalyptus essential oil nanoemulsion leads to the appearance of spherical structures on the surface of the composite membrane. This shows that the eucalyptus essential oil nanoemulsion was successfully loaded into the composite membrane, and spherical holes also appeared in the cross section of the composite membrane. It is worth noting that Figure 2 The surface image of SPI-GE-20% shown in Cc has no obvious spherical particles and impurities, which may be due to the uniform distribution of droplets and few flocculation and agglomeration problems. Eucalyptus essential oil nanoemulsion particles were observed in the cross-sectional image, indicating that eucalyptus essential oil was successfully encapsulated in the composite membrane matrix. Therefore, the appropriate amount of eucalyptus essential oil nanoemulsion has good compatibility with the membrane matrix and can improve the performance of the composite membrane.

[0089] 2. Fourier transform infrared spectroscopy (FTIR) analysis of eucalyptus essential oil nanoemulsion composite film

[0090] The FTIR spectrum of the composite membrane was measured by KBr compression method to analyze the changes in the structure and functional groups of the composite membrane. The measured wave number range was 4000-500cm -1 , resolution 4cm -1 .

[0091] FTIR is a commonly used technique to examine the chemical composition of active membranes. Figure 3 The chemical bonds of the packaging film and the interactions between molecules in the film are depicted, and the molecular structure characteristics of the active film are intuitively displayed. -1 The absorption peak at 2930 cm is attributed to the free vibration of -OH within and between molecules. -1 The characteristic peak observed at 1652 cm -1 The characteristic peak of hydrogen bond bending vibration observed at 1045 cm-1 is attributed to the interaction between -OH in the emulsion and the membrane matrix material. The stretching coupling of CC and CO bonds occurs at 1045 cm-1. -1This is the characteristic peak of the carbohydrate functional groups in the composite film matrix. Overall, the peaks of the composite films with and without latex addition are similar, and no new characteristic peaks appear between the two. The difference in peak intensity may be attributed to the interaction between the composite film matrix and the latex. The lack of obvious differences may be because the lower latex content in the active film is not enough to cause significant changes in the packaging film.

[0092] 3. Analysis of thermal stability of eucalyptus essential oil nanoemulsion composite film

[0093] The thermal stability of the composite membranes was analyzed using a comprehensive thermal analyzer (Netzsch STA449C, Freistaat, Bayern, Germany) with a temperature range of 0-800°C, a heating rate of 10°C / min, and a gas atmosphere of N2.

[0094] Thermogravimetric analysis was used to investigate the effect of eucalyptus essential oil nanoemulsion on the thermal stability of the composite film. The thermogravimetric curves (TGA) of the composite film containing different concentrations of eucalyptus essential oil nanoemulsion are shown in Figure 2. Figure 4 As shown, it reflects the change of the weight of the composite film with temperature. The thermal decomposition of the composite film can be divided into three stages. The first weight loss stage is at 89.05-187.27℃, which may be related to the evaporation of free water in the composite film and the loss of volatile components. The second stage of weight loss occurs at around 206.74-288.24℃, which is mainly due to the loss of plasticizer glycerol molecules in the composite film and the degradation of macromolecular substances such as proteins and polysaccharides. The third stage of weight loss is in the range of 294.04-450.13℃, and the mass loss in this stage may be caused by polymer carbonization. It is worth noting that the mass loss of all composite films with eucalyptus essential oil nanoemulsion is significantly lower than that of the blank film, indicating that the addition of eucalyptus essential oil improves the thermal stability of the composite film. When the temperature is higher than 400℃, the composite film with eucalyptus essential oil nanoemulsion has a higher residual mass than the composite film without eucalyptus essential oil nanoemulsion, which indicates that the addition of eucalyptus essential oil nanoemulsion hinders the thermal degradation and decomposition of the components in the composite film material, thereby improving the thermal stability of the composite film. Thermogravimetric analysis (DTG) results are as follows: Figure 5 As shown in the figure, three peaks were observed in the composite film near 90℃, 250℃, and 320℃, which is consistent with the results of TGA. In summary, the addition of eucalyptus essential oil nanoemulsion reduced the degradation of SPI and improved the performance of SPI. When the temperature is below 200℃, the composite film can maintain good thermal stability, which is sufficient to protect food for daily use in most cases.

[0095] 4. Analysis of antibacterial properties of eucalyptus essential oil nanoemulsion composite film

[0096] The agar diffusion method was used to detect the inhibitory effect of the composite membrane on Escherichia coli and Staphylococcus aureus. Take 100 μL of bacterial solution (live bacteria content 10 7 CFU / mL) was evenly spread on the surface of the agar medium, and the membrane was cut into small pieces with a 6mm diameter perforator. These small discs were placed in the center of the medium coated with the bacterial solution. The entire device was incubated in a constant temperature and humidity chamber at 37°C for 2 days. The diameter of the inhibition zone was measured with a vernier caliper.

[0097] The agar diffusion method was used to test the antibacterial effect of the nanoemulsion composite film with different contents of eucalyptus essential oil on Staphylococcus aureus and Escherichia coli. Figure 6 As shown in the figure, the SPI / GE composite film without eucalyptus essential oil nanoemulsion has no obvious antibacterial effect on the two bacteria. When eucalyptus essential oil nanoemulsion is added to the composite film, the diameter of the inhibition zone increases with the increase of the content of active substances added. When the addition amount of eucalyptus essential oil nanoemulsion is 40%, the inhibition zone for Escherichia coli and Staphylococcus aureus is the largest, reaching 12.6mm and 14.7mm respectively. The antibacterial effect of the composite film is mainly related to the released eucalyptus essential oil, whose active ingredients can destroy the cell membrane and cytoplasmic contents of bacteria and inhibit bacterial growth. Compared with Escherichia coli, the composite film has stronger inhibitory activity against Staphylococcus aureus.

[0098] 3. Analysis of the effect of eucalyptus essential oil nanoemulsion composite film on cherry preservation

[0099] Cherries of similar size were selected, and the SPI-GE-20% composite film with better comprehensive performance, the composite film without eucalyptus essential oil nanoemulsion, and the commercially available PE film were used for packaging test. At the same time, unpackaged cherries were set as blank controls, marked as emulsion film group, pure film group, PE group and blank group respectively. The practicality of the preservation effect of eucalyptus essential oil nanoemulsion soy protein isolate / gelatin composite film was evaluated. Cherries were stored at a constant 10°C and 50% relative humidity. The soluble solids (TSS), hardness, total phenols, flavonoids, headspace gas and V were measured every three days. C In addition, cherries of similar size were selected for the determination of weight loss and decay rate, and were also divided into four groups: emulsion membrane group, pure membrane group, PE group and blank group, with three parallel groups in each group, 7 sampling points were set, and 20 cherries were placed in each group.

[0100] 1. Cherry epigenetic analysis

[0101] The cherries were photographed and recorded every 3 days for a total of 21 days.

[0102] The appearance of cherries directly affects consumers' purchasing decisions. The effects of different composite film treatments on the appearance of cherries are as follows: Figure 7As shown. During storage, the appearance, color, taste, and smell of cherries changed, and the sensory performance of all cherries gradually decreased. Among the samples in each group, the fruit and stalk of the blank group shrank fastest and to a serious extent. The reason is that there is no packaging to resist the influence of the external environment, which leads to rapid loss of water in the cherries. PE plastic wrap has excellent barrier properties and fully blocks air and water vapor. However, the water vapor produced by the respiration of cherries cannot be discharged into the air. As the storage time increases, moisture accumulates and the cherries quickly rot and deteriorate. The cherries in the pure film group still have good color at 12 days. By the 15th day, the cherries have become soft in large quantities and mold colonies appear. This group of packaging films has a certain hygroscopicity, which can discharge internal water vapor, keep the surface of cherries dry, and delay the growth of colonies; however, compared with PE film, its barrier properties are poor, and soy protein isolate and gelatin themselves have certain nutrients. Therefore, after the mycelium growth appears on the cherries, the colonies multiply rapidly. Among all the samples, the sensory performance of cherries treated with SPI-GE-20% composite film was the best. After 15 days of storage, the appearance of cherries in this group was still relatively intact. The cherries in this group not only maintained a good taste and smell, but also had better appearance quality than other treatment groups. The cherries that rotted on the 18th day had a smaller rot area and did not spread rapidly on the 21st day. This may be because the eucalyptus essential oil in the emulsion continued to play an antibacterial role and inhibited the growth of bacterial colonies. In short, SPI-GE-20% is conducive to maintaining the quality of cherries and extending their shelf life.

[0103] 2. Analysis of cherry hardness

[0104] Ten cherries were randomly selected from each treatment group, and the hardness of the cherries was measured using a texture analyzer. The test conditions were: measuring speed of 1 mm / min and compression distance of 3 mm.

[0105] Hardness is one of the important indicators reflecting the quality of cherries. A decrease in hardness will seriously reduce the edible quality of the fruit. Figure 8 A shows the hardness changes of cherry fruits with different treatments during the entire storage period. As can be seen from the figure, the hardness of cherries after harvest tends to decrease with the increase of storage time. This is because the cell wall of cherry fruits contains more protopectin at the beginning of storage, which has a higher hardness. As the fruit matures, the protopectin transforms into water-soluble pectin and dissolves in the cell solution of the fruit, resulting in a decrease in hardness. In addition, the water in the fruit has a supporting effect on the cells. A large amount of water loss will cause the texture of the fruit to soften and the hardness to decrease. During the entire storage process, the hardness of cherries in the emulsion film treatment group decreased more slowly than that in the PE treatment group, the pure film treatment group and the blank control group. At the end of storage (21 days), the four groups of data decreased by 76.03%, 84.67%, 85.80% and 88.40% respectively compared with the initial hardness of the fruit, indicating that the eucalyptus essential oil nanoemulsion composite film can effectively delay the decline in fruit hardness during cherry storage.

[0106] 3. Analysis of cherry rot rate

[0107] The rot rate is an important indicator for evaluating the preservation effect of cherries. In this study, the rot rate of cherries was determined by identifying fruits that were one-third damaged, soft and rotten, or had mold growth as rotten fruits. The rot rate of cherries was calculated using the following formula:

[0108]

[0109] like Figure 8 As shown in Figure B, the rot rate of cherries treated with different films at 10°C showed an overall upward trend with the extension of storage time. On the 6th day of storage, the cherries treated with blank and PE cling film rotted first, while the cherries treated with pure film group and emulsion film group rotted on the 12th and 18th days, respectively. This shows that both the SPI-GE-20% composite film and the composite film without eucalyptus essential oil nanoemulsion have played a role in extending the shelf life of cherries, and their effect is better than that of commercially available PE cling film. Among them, the emulsion film group rotted the latest and had a low rot rate, which may be due to the antibacterial effect of the essential oil, which reduced the probability of rot and deterioration caused by pathogen infection, thereby inhibiting the increase in the rot rate of the fruit. This shows that the eucalyptus essential oil nanoemulsion composite film treatment can better inhibit the rot of cherries and has a good storage and preservation effect.

[0110] 4. Analysis of cherry weight loss rate

[0111] The cherries were weighed and recorded every three days, and the weight loss rate of the cherries was calculated using the following formula:

[0112]

[0113] Where m0 is the initial weight of the cherry, m t is the weight of cherries on the sampling day.

[0114] After being picked, cherries are very likely to lose water and wilt due to respiration and transpiration during storage, resulting in loss of nutrients and weight loss. Figure 8 C shows the changes in weight loss of cherries treated with four different methods during storage. Throughout the test, the weight loss of all samples increased with the extension of storage time. The lower permeability of the composite film inhibits the occurrence of cherry transpiration during post-harvest storage. SPI-GE-20% composite film can effectively reduce water evaporation during storage and reduce weight loss.

[0115] 5. Analysis of Cherry Respiration Rate

[0116] The respiration rate of cherries was determined using the static closed method. 20 cherry fruits were placed in a respiration bottle, and the bottle cap was treated with a sealing film to maintain a closed environment. After the respiration bottle was stored at 4±1℃ for 6 hours, the concentration of CO2 in the respiration bottle was determined using a portable O2 / CO2 gas analyzer. The respiration rate of cherries was calculated according to the following formula:

[0117]

[0118] Where △C is the change in the volume fraction of CO2 in the breathing bottle (%), △V is the volume of gas in the closed container (L), m is the mass of cherries (g), and t is the storage time (h).

[0119] Respiration is an important metabolic process that maintains the life activities of fruits and vegetables. In this process, complex organic matter is slowly decomposed into simple organic matter under the action of enzymes, accompanied by the generation of energy. Therefore, the respiration rate is an important reference for the maturity, aging and even corruption of fruits and vegetables. Figure 8 D shows the changes in the respiration rate of cherries in different treatment groups during storage. As the storage time increases, the respiration rate of cherries in different treatment groups continues to increase, which will lead to a gradual decrease in organic matter in cherry fruits, continuous aging of cherries, and a gradual decrease in storage resistance. After the sixth day of storage, the respiration rate of cherries treated with the eucalyptus essential oil emulsion film with low O2 permeability was significantly lower than that of other treatment groups (p<0.05), and the respiration rate of cherries treated with the PE film and the composite film without eucalyptus essential oil was close, and the respiration rate of cherries treated with the blank film was the strongest. This may be because the eucalyptus essential oil released by the eucalyptus essential oil emulsion film has antioxidant properties, which can inhibit the respiration of cherries and delay the aging of cherries. In addition, the eucalyptus essential oil emulsion film has a low O2 permeability, which can maintain a high CO2 and low O2 gas environment and inhibit respiration. In summary, the eucalyptus essential oil emulsion film can reduce the respiration rate of cherries, thereby helping to maintain the good quality of cherries and extend the shelf life.

[0120] 6. Cherry V C Analysis

[0121] Weigh 1 g of fruit, add 9 mL of 0.9% saline, grind into homogenate under ice bath, centrifuge at 4°C, 2500 r / min for 10 min, collect the supernatant (10% homogenate supernatant) for V C Determination of content.

[0122] Use "Vitamin C (V C ) / Ascorbic acid (ASA) assay kit" to determine the V C content.

[0123] V CIt is an important antioxidant ingredient in cherries, is closely related to slowing down the aging and deterioration of fruits, and is one of the important indicators for evaluating fruit quality. Fig. 9 A is the V of cherry fruits in different treatment groups during storage C As can be seen from the figure, with the extension of storage time, the V C The content showed a trend of first decreasing and then increasing. Among them, the cherry V treated with SPI-GE-20% composite film C The content was the highest in all treatment groups, which may be related to the antioxidant activity of eucalyptus essential oil nanoemulsion on the inhibition of oxidative stress during cherry storage.

[0124] 7. Analysis of total phenol content in cherry

[0125] Weigh 1.0 g of fruit, add 30 mL of 80% (v / v) ethanol solution, grind into a homogenate under ice bath conditions, centrifuge at 4°C, 5000 r / min for 10 min, and collect the supernatant for the determination of total phenolic content.

[0126] The total phenol content was determined by the Folin phenol method. Take 0.4 mL of supernatant, add 2 mL of 10% (v / v) Folin phenol solution, shake well and let stand in the dark for 5 minutes, then add 1.8 mL of 7.5% (w / v) Na2CO3 solution, shake well and let stand in the dark for 1 hour, and measure the absorbance at 765 nm. Use standard gallic acid solution as the standard curve (y=0.01758+0.05129x). Repeat the determination three times. The results are calculated according to the following formula:

[0127]

[0128] Wherein, C is the mass of gallic acid obtained according to the standard curve, μg; V is the total volume of the sample extract, ml; V1 is the volume of the extract taken during the determination, ml; M is the mass of the fruit, g.

[0129] Total phenolics refer to the collection of all phenolics contained in fruits and vegetables, which are easily lost due to oxidation, which is closely related to their nutritional value. Fig. 9 B shows the changes in total phenolic content of cherries treated with four different methods during storage. The total phenolic content of cherries in each treatment group showed a rapid downward trend in the early stage of storage, and the change gradually slowed down in the later stage. Both the pure film and emulsion film treatment groups played a role in delaying the decline of total phenolic content in the fruit. After 6 days of storage, the total phenolic content of cherries treated with SPI-GE-20% composite film was significantly higher than that of other groups. This result shows that SPI-GE-20% composite film treatment can promote the accumulation of phenolic substances in cherries.

[0130] 8. Analysis of flavonoid content in cherry

[0131] Weigh 1.0 g of fruit, add 30 mL of 80% (v / v) ethanol solution, grind into a homogenate under ice bath conditions, centrifuge at 4°C and 5000 r / min for 10 min, and collect the supernatant for the determination of flavonoid content.

[0132] The flavonoid content was determined by ultraviolet spectrophotometry. Take 1 mL of supernatant, add 0.3 mL of 5% (w / v) NaNO2 solution, shake well and let stand in the dark for 5 min, then add 0.3 mL (w / v) 10% AlCl3 solution, shake well and let stand in the dark for 5 min, then add 2 mL of 1 mol L -1 Add 3.4 mL of distilled water to the NaOH solution, shake well, let stand in the dark for 1 hour, and measure the absorbance at 510 nm. Use the standard rutin solution as the standard curve. Repeat the measurement three times. The results are calculated according to the following formula:

[0133]

[0134] Wherein, C is the mass of gallic acid obtained according to the standard curve, μg; V is the total volume of the sample extract, ml; V1 is the volume of the extract taken during the determination, ml; M is the mass of the fruit, g.

[0135] Flavonoid compounds are important nutrients in cherries. They have antioxidant, anti-inflammatory, antibacterial, tumor cell proliferation inhibition and immune regulation functions. Their content is an important indicator for evaluating the nutritional value of cherries. Fig. 9 C shows the changes in flavonoid content of cherry fruits in different treatment groups during storage. As can be seen from the figure, with the extension of storage time, the flavonoid content of cherry fruits shows a trend of first decreasing and then increasing. The later increase may be due to the biological stress of cherries caused by microbial reproduction, which in turn induced the production of secondary metabolites and caused the increase of total phenolic flavonoid content. Among them, the flavonoid content of cherries treated with the emulsion film group was the highest among all treatment groups, which shows that the SPI-GE-20% composite film treatment can inhibit the post-harvest metabolic activity of cherries, delay the ripening of fruits, and thus extend the shelf life. The total phenol and flavonoid content in cherries are two important indicators for measuring the flavor quality of fruits. The SPI-GE-20% composite film maintains a higher level of total phenols and flavonoids, which can delay the loss of cherry flavor and achieve the effect of preservation.

Claims

1. A method for preparing a soy protein isolate / gelatin packaging film containing eucalyptus essential oil nanoemulsion, characterized in that: S1: Preparation of eucalyptus essential oil nanoemulsion The eucalyptus essential oil, an emulsifier and water are mixed to prepare a crude emulsion; and then a low temperature ultrahigh pressure continuous flow cell disruptor is used to homogenize the mixture at a pressure of 1000-1200 bar to prepare a eucalyptus essential oil nanoemulsion; S2: Preparation of soy protein isolate and gelatin mixture Prepare a soy protein isolate and gelatin mixed solution containing soy protein isolate, gelatin, glycerin and water; the mass ratio of the soy protein isolate to gelatin is 1:3.5-4.5; S3: Preparation of soy protein isolate / gelatin packaging film containing eucalyptus essential oil nanoemulsion The eucalyptus essential oil nanoemulsion is slowly added into the soy protein isolate and gelatin mixed solution and stirred evenly; then the mixture is poured into a plastic plate, stirred, and dried to obtain a soy protein isolate / gelatin packaging film containing the eucalyptus essential oil nanoemulsion.

2. The preparation method according to claim 1, characterized in that: In the step S1, the content of eucalyptus essential oil is 1.5-1.8% of the total mass of the crude emulsion.

3. The preparation method according to claim 1, characterized in that: The emulsifier is Tween-80; the dosage of Tween-80 is 40-60% of the total mass of the eucalyptus essential oil.

4. The preparation method according to claim 2, characterized in that: The specific steps of step S2 are as follows: (1) preparing a soy protein isolate solution with a concentration of 4.5-5.5%, adjusting the pH of the solution to 9.5-10.5 with alkali solution, and then heating and stirring in a water bath at 85-95° C. for 50-70 minutes; (2) preparing a gelatin solution with a concentration of 4.5-5.5%, hydrating for 25-35 min, and then heating and stirring in a water bath at 45-55°C for 25-35 min; (3) Mixing the soy protein isolate solution and the gelatin solution, adding 20-30% glycerol, heating and stirring in a water bath at 45-55° C. for 55-65 minutes, and obtaining a soy protein isolate and gelatin mixed solution; the glycerol content is the amount relative to the dry matter of the soy protein isolate and gelatin mixed solution.

5. The preparation method according to claim 4, characterized in that: The addition amount of the eucalyptus essential oil nanoemulsion is 10-30% relative to the volume of the soy protein isolate and gelatin mixed solution.

6. The preparation method according to claim 5, characterized in that: The added amount of the eucalyptus essential oil nanoemulsion is 20%.

7. The soy protein isolate / gelatin packaging film containing eucalyptus essential oil nanoemulsion prepared by the preparation method described in any one of claims 1-6.

8. Application of the soy protein isolate / gelatin packaging film containing eucalyptus essential oil nanoemulsion according to claim 7 in food antibacterial, antioxidant and fresh-keeping properties.

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