Smart active composite film with fresh-keeping and monitoring functions and preparation method and application thereof

Abstract

The application discloses an intelligent active composite film with fresh-keeping and monitoring functions and a preparation method and application thereof. The preparation method comprises the following steps: limiting and intercepting thymol by spirulina to obtain spirulina loaded with thymol; coating the spirulina loaded with thymol by carboxymethyl chitosan to prepare an intelligent fragrance micro-reactor with pH / humidity dual response characteristics; and performing film forming treatment on a film forming liquid containing konjac glucomannan, blueberry anthocyanin, the intelligent fragrance micro-reactor and water to prepare the intelligent active composite film with the fresh-keeping and monitoring functions. The intelligent active composite film can realize fresh-keeping through active anti-corrosion intervention, has the in-situ freshness real-time diagnosis function and has a good application prospect in novel green food packaging materials.

Description

Technical Field

[0001] This invention belongs to the field of fruit and vegetable preservation technology, specifically relating to an intelligent active composite film with preservation and monitoring functions, its preparation method, and its application. Background Technology

[0002] Fresh fruits and vegetables, such as strawberries, are highly susceptible to mechanical damage and infection by pathogens (such as botrytis cinerea) during post-harvest storage and cold chain logistics due to their high water content and various nutrients. This can lead to severe tissue softening, nutrient loss, and spoilage. Traditional passive barrier packaging is no longer sufficient to meet the modern food industry's demand for long-term preservation. In recent years, "active packaging" technology, which can actively regulate the internal microenvironment of packaging, has attracted much attention. Among them, thymol (THY), a natural polyphenol derived from plant essential oils, is widely considered an ideal alternative for preserving fruits and vegetables due to its excellent broad-spectrum gas-phase antibacterial and antioxidant activities. However, thymol's extremely high volatility and strong pungent odor make it prone to "burst release" in practical applications, which not only makes it difficult to maintain a long-term preservation concentration but may even negatively affect the flavor of food. Therefore, how to achieve precise targeted regulation and long-term sustained release of thymol is a key bottleneck that urgently needs to be overcome in the field of active packaging. Summary of the Invention

[0003] The main objective of this invention is to provide a smart active composite film with preservation and monitoring functions, its preparation method and application, so as to overcome the shortcomings of the prior art.

[0004] To achieve the aforementioned objectives, the technical solution adopted by this invention includes:

[0005] This invention provides a method for preparing a smart active composite film with preservation and monitoring functions, comprising:

[0006] Spirulina was used to restrict and retain thymol to obtain thymol-loaded Spirulina (SP-THY); wherein the mass ratio of Spirulina to thymol was 0.5:1 to 2:1.

[0007] A smart fragrance microreactor (SP@CMCS) with dual pH / humidity response characteristics was prepared by coating the thymol-loaded spirulina with carboxymethyl chitosan (CMCS); wherein the mass ratio of the thymol-loaded spirulina to carboxymethyl chitosan was 1:5 to 1:20.

[0008] Furthermore, a film-forming solution containing konjac glucomannan (KGM), blueberry anthocyanins (BA), a smart fragrance microreactor, and water is subjected to film-forming treatment to obtain a smart active composite film (BK-SP@CMCS) with preservation and monitoring functions; wherein the mass ratio of konjac glucomannan, blueberry anthocyanins, and the smart fragrance microreactor is 1:0.2:0.1~0.3.

[0009] The present invention also provides a smart active composite film with preservation and monitoring functions prepared by the aforementioned preparation method.

[0010] The embodiments of the present invention also provide the application of the aforementioned intelligent active composite film with preservation and monitoring functions in the field of fruit and vegetable preservation monitoring.

[0011] This invention also provides a method for preserving and monitoring fruits and vegetables, which includes: sealing and storing fruits or vegetables using the aforementioned intelligent active composite film with preservation and monitoring functions;

[0012] The storage environment is characterized by an ambient temperature of 20-25°C and a relative humidity of 70-80%.

[0013] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0014] (1) The present invention innovatively designed a dual-response intelligent fragrance microreactor, which effectively overcomes the bottleneck of traditional plant essential oils being volatile and prone to sudden release. This structure, through the confined retention of spirulina and electrostatic coating, endows the system with pH / humidity dual-response characteristics, enabling it to achieve intelligent slow release of thymol and maintain long-term freshness in the high-humidity and acidic microenvironment of fruit and vegetable spoilage.

[0015] (2) The intelligent active composite film in this invention integrates the dual functions of active anti-corrosion intervention and in-situ real-time visual diagnosis of freshness. The composite film can not only establish a long-lasting antibacterial and antioxidant barrier through microreactors, but also the anthocyanins inside it can undergo sensitive color changes when fruits and vegetables rot and produce acid, thus achieving non-destructive freshness monitoring.

[0016] (3) The present invention significantly improves the macroscopic water barrier function and comprehensive mechanical strength of the composite film. The micro-nano rough structure constructed on the membrane surface by the microreactor greatly enhances the hydrophobicity of the material. At the same time, the high-density three-dimensional hydrogen bond network formed by it and the matrix molecules significantly improves the tensile strength and elongation at break of the film.

[0017] (4) The present invention exhibits excellent multidimensional synergistic antibacterial efficacy and outstanding practical fruit and vegetable preservation effect. The film achieves broad-spectrum and efficient sterilization against typical foodborne pathogens. In practical applications, it can significantly delay the weight loss and quality decline of strawberries, successfully block the spoilage process and greatly extend the shelf life of fresh produce. Attached Figure Description

[0018] To more clearly illustrate the technical solutions in the embodiments of this application or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments recorded in this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0019] Figure 1 This is a graph showing the encapsulation rate of thymol-loaded spirulina prepared with different mass ratios of spirulina and thymol in Example 1 of the present invention.

[0020] Figure 2 This is a graph showing the loading results of thymol-loaded spirulina prepared with different mass ratios of spirulina and thymol in Example 1 of the present invention.

[0021] Figure 3 This is a graph showing the release rate of thymol from a series of intelligent fragrance microreactors prepared in Example 1 of this invention.

[0022] Figure 4 This is a statistical result graph of the DPPH free radical scavenging rate of different composite films in Example 1 of the present invention;

[0023] Figure 5 This is a statistical result graph of the ABTS free radical scavenging rate of different composite films in Example 1 of the present invention;

[0024] Figure 6 This is a statistical chart of the water contact angle (WCA) of different composite film surfaces in Embodiment 1 of the present invention;

[0025] Figure 7 These are Fourier transform infrared spectra of different composite thin films in Embodiment 1 of the present invention;

[0026] Figure 8 These are macroscopic thickness measurement diagrams of different composite films in Example 1 of the present invention;

[0027] Figure 9 This is a graph showing the elongation at break of different composite films in Example 1 of the present invention;

[0028] Figure 10 These are tensile results of different composite films in Embodiment 1 of the present invention;

[0029] Figure 11 This is a bar chart showing the antibacterial rate of representative colonies of Escherichia coli on agar plates after treatment with different composite thin-layer extracts in Example 1 of the present invention.

[0030] Figure 12This is a bar chart showing the quantitative antibacterial rate of representative colonies of Staphylococcus aureus on agar plates after treatment with different composite thin-layer extracts in Example 1 of the present invention.

[0031] Figure 13 These are optical photographs of the macroscopic phenotypic evolution of strawberries in the blank control group, KGM, BA-KGM and BK-SP@CMCS groups during storage at 25 ℃ on days 0, 2, 4 and 6 of Example 1 of this invention.

[0032] Figure 14 This is a graph showing the weight loss rate of strawberries in the blank control group, KGM, BA-KGM and BK-SP@CMCS groups during storage at 25 ℃ on days 0, 2, 4 and 6 in Example 1 of this invention.

[0033] Figure 15 This is a graph showing the firmness of strawberries in the blank control group, KGM, BA-KGM and BK-SP@CMCS groups during storage at 25 ℃ on days 0, 2, 4 and 6 in Example 1 of this invention.

[0034] Figure 16 The color difference L of strawberries in the blank control group, KGM, BA-KGM, and BK-SP@CMCS groups during storage at 25℃ on days 0, 2, 4, and 6 of Example 1 of this invention. * picture;

[0035] Figure 17 This is a graph showing the soluble solids (TSS) content of strawberries in the blank control group, KGM, BA-KGM and BK-SP@CMCS groups during storage at 25 ℃ on days 0, 2, 4 and 6 of Example 1 of this invention. Detailed Implementation

[0036] In view of the deficiencies of the prior art, the inventors of this case, through long-term research and extensive practice, have proposed the technical solution of this invention. The technical solution of this invention will be clearly and completely described below. Obviously, the described embodiments are only some, not all, of the embodiments of this invention. All other embodiments obtained by those skilled in the art based on the embodiments of this invention without creative effort are within the scope of protection of this invention.

[0037] Specifically, as one aspect of the technical solution of this invention, the method for preparing a smart active composite film with preservation and monitoring functions includes:

[0038] Spirulina was used to restrict and retain thymol to obtain thymol-loaded Spirulina (SP-THY); wherein the mass ratio of Spirulina to thymol was 0.5:1 to 2:1.

[0039] A smart fragrance microreactor (SP@CMCS) with dual pH / humidity response characteristics was prepared by coating the thymol-loaded spirulina with carboxymethyl chitosan (CMCS); wherein the mass ratio of the thymol-loaded spirulina to carboxymethyl chitosan was 1:5 to 1:20.

[0040] Furthermore, a film-forming solution containing konjac glucomannan (KGM), blueberry anthocyanins (BA), a smart fragrance microreactor, and water is subjected to film-forming treatment to obtain a smart active composite film (BK-SP@CMCS) with preservation and monitoring functions; wherein the mass ratio of konjac glucomannan, blueberry anthocyanins, and the smart fragrance microreactor is 1:0.2:0.1~0.3.

[0041] In some preferred embodiments, the method for preparing the spirulina includes:

[0042] Accurately prepare the modified BG11 medium: Dissolve the standard amount of BG11 medium powder in 1 L of ultrapure water, and add 13 g NaOH and 1 g NaCl to construct a strongly alkaline, high-salt microenvironment suitable for the growth and development of Spirulina. Inoculate the Spirulina seed culture in the logarithmic growth phase into the above medium at an appropriate inoculation rate, and then place it in a light-temperature incubator for static culture: temperature 25 ℃, light / dark cycle 12 h light / 12 h dark, light intensity 4700 lux. After the algal concentration reaches a plateau, transfer the algal culture to centrifuge tubes and centrifuge at high speed (2000 r / min, 10 min) to collect the bottom algal precipitate. Wash the algae three times with deionized water until the supernatant is neutral to completely remove residual culture medium salts from the surface. Then, pre-freeze the washed, purified algal sludge in an ultra-low temperature freezer at -80 ℃ for 12 h, and then freeze-dry it in a vacuum freeze dryer for 48 h. Collect the freeze-dried spirulina solid powder, grind it thoroughly in an agate mortar and sieve it to obtain spirulina (SP) carrier powder with a natural loose and porous microcavity structure, and store it in a desiccator in the dark for later use.

[0043] In some preferred embodiments, the preparation method specifically includes: dissolving thymol in anhydrous ethanol to form an active core material stock solution, then adding spirulina and stirring the solution at room temperature and in the dark for 12-24 hours, followed by centrifugation, washing, and freeze-drying to obtain spirulina loaded with thymol.

[0044] Furthermore, the mass ratio of spirulina to thymol is 1:1 to 1.5:1.

[0045] Furthermore, the concentration of thymol in the active core material stock solution is 0.9~1.1 g / mL.

[0046] In some preferred embodiments, the preparation method specifically includes:

[0047] Spirulina loaded with thymol was dispersed in water to form a spirulina suspension loaded with thymol.

[0048] Carboxymethyl chitosan is dissolved in water to form an aqueous solution of carboxymethyl chitosan;

[0049] Furthermore, the spirulina suspension loaded with thymol was added dropwise to a carboxymethyl chitosan aqueous solution and stirred for 4-6 hours at room temperature and in the dark. After centrifugation, washing, vacuum freeze-drying, and grinding, an intelligent fragrance microreactor with dual pH / humidity response characteristics was obtained.

[0050] Furthermore, the mass ratio of the thymol-loaded spirulina to carboxymethyl chitosan is 1:15 to 1:20.

[0051] Furthermore, the intelligent fragrance microreactor has a core-shell structure.

[0052] In some preferred embodiments, the preparation method specifically includes:

[0053] Konjac glucomannan was dissolved in water to form a konjac glucomannan solution. Then, blueberry anthocyanins and a smart fragrance microreactor were added and thoroughly mixed and dispersed to form a film-forming solution.

[0054] Furthermore, the film-forming liquid is degassed, molded, and dried to obtain an intelligent active composite film with preservation and monitoring functions.

[0055] Furthermore, the concentration of the konjac glucomannan solution is 0.01 g / mL.

[0056] Furthermore, the drying process is carried out under light-protected conditions at 30~45°C.

[0057] Another aspect of the present invention provides a smart active composite film with preservation and monitoring functions prepared by the aforementioned preparation method.

[0058] Another aspect of the present invention provides the application of the aforementioned intelligent active composite film with preservation and monitoring functions in the field of fruit and vegetable preservation monitoring.

[0059] Another aspect of the present invention provides a method for preserving and monitoring fruits and vegetables, comprising: sealing and storing fruits or vegetables using the aforementioned intelligent active composite film with preservation and monitoring functions;

[0060] The storage environment is characterized by an ambient temperature of 20-25°C and a relative humidity of 70-80%.

[0061] The technical solution of the present invention will be further described in detail below with reference to several preferred embodiments and accompanying drawings. This embodiment is implemented on the premise of the technical solution of the invention, and provides detailed implementation methods and specific operation processes. However, the protection scope of the present invention is not limited to the following embodiments.

[0062] Unless otherwise specified, the experimental materials used in the examples below can be purchased from conventional biochemical reagent companies.

[0063] Example 1

[0064] (1) Preparation of Spirulina loaded with thymol (SP-THY)

[0065] The Spirulina used in this invention was obtained through a laboratory-developed culture system. A modified BG11 medium was accurately prepared: standard amounts of BG11 medium powder were dissolved in 1 L of ultrapure water, with an additional 13 g NaOH and 1 g NaCl added to create a highly alkaline, high-salt microenvironment suitable for Spirulina growth and development. Spirulina in the logarithmic growth phase was inoculated into the medium at an appropriate inoculation rate and then statically cultured in a light-controlled thermostat: temperature 25℃, light / dark cycle of 12 h light / 12 h dark, light intensity 4700 lux. Once the algal concentration reached a plateau, the algal solution was transferred to centrifuge tubes and centrifuged at high speed (2000 r / min, 10 min) to collect the bottom algal precipitate. The algal precipitate was washed three times with deionized water by centrifugation until the supernatant was neutral to completely remove residual culture medium salts from the surface. Subsequently, the washed pure spirulina mud was pre-frozen in an ultra-low temperature freezer at -80 ℃ for 12 h, and then freeze-dried in a vacuum freeze dryer for 48 h. The freeze-dried spirulina solid powder was collected, thoroughly ground in an agate mortar and sieved to obtain spirulina (SP) carrier powder with a natural loose porous microcavity structure, which was then stored in a desiccator in the dark for later use.

[0066] Accurately weigh a specific amount of thymol (THY) crystals and completely dissolve them in anhydrous ethanol to prepare an active core material stock solution with a concentration of 1 g / mL. Subsequently, according to experimental gradients of core-carrier mass ratios (SP:THY) of 0.5:1, 1:1, 1.5:1, and 2:1, the prepared SP porous carrier powder was added to the above solution sequentially. The mixture was continuously magnetically stirred for 24 h at room temperature and in the dark to allow THY to fully diffuse and be confined within the microcavity network of SP. After the reaction was completed, the precipitate was collected by centrifugation, and the free core material on the surface was rapidly eluted with an appropriate amount of low-concentration ethanol. The resulting product was then freeze-dried under vacuum to obtain primary microparticles (SP-THY) with different loading gradients.

[0067] Testing: Determination of Encapsulation Efficiency (EE) and Loading Capacity (LC)

[0068] Accurately weigh a certain mass of thymol standard, completely dissolve it in anhydrous ethanol, and then sequentially dilute it to prepare a series of THY standard solutions with concentration gradients. Using a UV-Vis spectrophotometer, measure the absorbance of each standard solution at the maximum absorption wavelength of THY, 274 nm. Plot a standard curve by plotting the mass concentration of the standard solution on the x-axis and the corresponding absorbance value on the y-axis, and obtain the linear regression equation to facilitate accurate quantification of the free THY concentration in the system.

[0069] In the centrifugation and washing process for SP-THY preparation described above, all collected centrifugal supernatants were combined with low-concentration ethanol washing solution and transferred to a volumetric flask for final volume determination. An appropriate amount of the final volumetric supernatant was taken, and its absorbance was measured at 274 nm using a UV-Vis spectrophotometer. The absorbance was then substituted into the standard curve equation described above to calculate the total mass of free THY not retained by Spirulina in the solution (M). free Meanwhile, the total dry weight (M) of the primary microparticles (SP-THY) obtained after vacuum freeze-drying was accurately weighed using an analytical balance. total The encapsulation efficiency (EE) and loading capacity (LC) of the microreactor are calculated using the following formulas:

[0070]

[0071]

[0072] Where: M initial The total mass (mg) of thymol added to the initial system during preparation; M free The total mass (mg) of free thymol in the supernatant and eluent; M total The total dry weight (mg) of SP-THY powder collected after freeze-drying. Each sample group was tested in triplicate.

[0073] Conclusion: Encapsulation efficiency (EE) and loading capacity (LC) are the core indicators for evaluating the loading performance of microcapsules / microreactors, directly determining the effective release dose of active substances and economic benefits in subsequent composite films. To obtain the optimal loading system, the effects of different core-carrier ratios (spirulina (SP) to thymol (THY) mass ratios of 0.5:1, 1:1, 1.5:1, and 2:1, respectively) on the loading performance of microreactors were investigated.

[0074] like Figure 1As shown, the encapsulation efficiency of thymol increased significantly with the increase of the SP:THY mass ratio from 0.5:1 to 2:1. At a ratio of 0.5:1, the encapsulation efficiency (EE) was only about 37.4% due to excessive thymol and insufficient carrier space. When the mass ratio increased to 1:1, the EE jumped significantly to 86.2%. Further increasing the carrier ratio (1.5:1 and 2:1) further improved the encapsulation efficiency, which stabilized between 91% and 95%. This is because the increased Spirulina (SP) content provided thymol with more binding sites, a larger specific surface area, and more porous microcavities, effectively enhancing the physical confinement and retention of hydrophobic thymol molecules. However, the evolution of the loading (LC) showed a distinctly different trend of first increasing and then decreasing. Figure 2 At a SP:THY ratio of 1:1, the microreactor loading reached its peak (approximately 46.5%), significantly outperforming other groups. When the carrier ratio was further increased to 1.5:1 and 2:1, the LC significantly decreased to approximately 38.1% and 32.2%, respectively. This phenomenon is attributed to the "carrier dilution effect": although more spirulina can encapsulate more thymol, excessive empty or unsaturated spirulina substrate significantly increases the overall dry weight of the microreactor, resulting in a significant dilution of the proportion of effective active substances per unit mass of microreactor.

[0075] Considering both encapsulation efficiency and actual drug loading, when the SP:THY mass ratio is 1:1, the microreactor achieves the maximum loading (46.5%) while maintaining a high encapsulation rate (>86%), reaching the optimal balance of carrier space utilization. Therefore, subsequent experiments all used a 1:1 mass ratio to prepare thymol-loaded Spirulina microreactors (SP-THY), and on this basis, carboxymethyl chitosan (CMCS) coating and film preparation were carried out.

[0076] (2) Preparation of intelligent fragrance microreactor (spirulina-thymol@carboxymethyl chitosan, SP@CMCS):

[0077] To construct a smart, responsive, sustained-release shell, SP-THY powder was uniformly dispersed in deionized water at a specific ratio to form a suspension. Under vigorous mechanical stirring, the suspension was slowly added dropwise to a pre-prepared carboxymethyl chitosan (CMCS) aqueous solution at experimental gradients of SP-THY:CMCS coating ratios of 1:5, 1:10, 1:15, and 1:20. The mixture was stirred at a constant temperature and in the dark for 4–6 h to promote the spontaneous assembly of the negatively charged SP-THY core and the positively charged CMCS molecular chains through electrostatic attraction and hydrogen bonding to form a "core-shell" structure. Finally, the mixture was centrifuged and washed repeatedly with deionized water to remove free CMCS. The precipitate was then freeze-dried under vacuum and thoroughly ground to obtain the SP@CMCS smart fragrance microreactor powder (spirulina-thymol@carboxymethyl chitosan), which was sealed and protected from light for later use.

[0078] To evaluate the inhibitory effect of different carboxymethyl chitosan (CMCS) coating ratios on the initial burst release effect of thymol (THY), this experiment screened and optimized the coating process through in vitro release experiments. Equal amounts of SP@CMCS lyophilized powder with different coating ratios were accurately weighed and placed in blue-capped bottles containing a quantitative amount of PBS buffer. The samples were placed in a constant-temperature water bath shaker at 25 ℃ and a rotation speed of 150 r / min for light-protected shaking. After the initial 12 h, an appropriate amount of suspension was centrifuged at high speed (10000 r / min, 5 min), and the supernatant was collected. The absorbance of the supernatant was measured at 274 nm using a UV-Vis spectrophotometer, and the concentration of THY in the release medium was calculated by substituting the absorbance into the thymol standard curve. Based on the calculated ratio of THY release to the total THY loading in the sample, the THY release rate (%) of different coating systems was determined, thereby identifying the optimal electrostatic coating ratio.

[0079] Conclusion: To further delay the burst release effect of thymol and impart environmental responsiveness to the system, the selected SP-THY microreactor was electrostatically coated with carboxymethyl chitosan. The effect of the coating process on the release rate was evaluated, and the successful construction of the SP@CMCS composite system was verified by multidimensional spectroscopy.

[0080] As an outer shell material, the concentration of CMCS directly determines the density and thickness of the coating layer, thus affecting the release kinetics of the core material. For example... Figure 3As shown, the release rate of THY exhibited a significant, phased decreasing trend as the mass ratio of SP-THY to CMCS increased from 1:5 to 1:20. At ratios of 1:5 and 1:10, due to insufficient shell material, a complete coating network could not be formed, resulting in release rates as high as 67.8% and 41.5%, respectively. When the mass ratio was further increased to 1:15, the release rate of THY decreased significantly and stabilized between approximately 13% and 15%; further increasing the CMCS ratio to 1:20 did not result in a significant difference in release rate. This indicates that at a ratio of 1:15, CMCS formed a complete and dense physical barrier on the surface of SP-THY, effectively suppressing the initial burst release of thymol. Therefore, subsequent characterizations were performed using SP@CMCS particles prepared at a ratio of 1:15.

[0081] (3) Preparation of intelligent active composite film (anthocyanin-konjac glucomannan / spirulina-thymol@carboxymethyl chitosan, BK-SP@CMCS):

[0082] A series of smart composite films were prepared under light-protected conditions. First, konjac glucomannan (KGM) was slowly dissolved in deionized water to prepare a homogeneous film-forming solution of 0.01 g / mL. After vacuum degassing, the solution was quantitatively poured into a flat mold and dried at a constant temperature of 45 °C to obtain a KGM membrane (konjac glucomannan membrane). Based on this, 0.2 g of blueberry anthocyanin (BA) was fully dissolved in 100 mL of the above KGM solution, and a BA-KGM membrane (anthocyanin-konjac glucomannan membrane) was prepared using the same drying and film-forming process. To further construct the BK-SP@CMCS smart composite film, 0.1 g, 0.2 g, and 0.3 g of dried SP@CMCS microreactor powder were accurately added to each 100 mL of uniformly mixed BA-KGM membrane solution, respectively, and homogeneously dispersed by vigorous mechanical stirring and short-term ice bath ultrasonication. The mixed membrane solution was then degassed, placed in a mold, and dried at 45 ℃ in the dark, and labeled as follows: BK-SP@CMCS-1 (anthocyanin-konjac glucomannan / spirulina-thymol@carboxymethyl chitosan-1), BK-SP@CMCS-2 (anthocyanin-konjac glucomannan / spirulina-thymol@carboxymethyl chitosan-2), and BK-SP@CMCS-3 (anthocyanin-konjac glucomannan / spirulina-thymol@carboxymethyl chitosan-3). After all the prepared films were peeled off, they were uniformly placed in a desiccator with 50% relative humidity for equilibration and later use.

[0083] a. In vitro antioxidant activity assay

[0084] To evaluate the antioxidant efficacy of the composite film, quantitatively cut film samples were immersed in an appropriate amount of solvent to prepare an extract. This extract was then thoroughly mixed with a standard concentration of DPPH alcohol solution and an ABTS free radical working solution, reacting in the dark for 30 min and 6 min, respectively. The absorbance of the reaction system was measured at 517 nm and 734 nm wavelengths using a UV-Vis spectrophotometer (UV-2600, Shimadzu Corporation, Japan). The scavenging rates of the film against the two types of free radicals were calculated based on these values. Each group was measured independently in triplicate, and the average value was taken.

[0085] Conclusion: Lipid peroxidation and reactive oxygen species (ROS) accumulation are the dominant factors leading to browning and nutritional degradation in fruits. Based on this, the antioxidant efficiency of the membrane was systematically evaluated using DPPH and ABTS free radical scavenging experiments. Figure 4 and 5 KGM membranes (konjac glucomannan membranes) lack effective electron donor structures and have extremely low antioxidant activity. BA-KGM membranes (anthocyanin-konjac glucomannan membranes) exhibit significantly enhanced antioxidant capacity, primarily attributed to the abundant phenolic hydroxyl groups in the blueberry anthocyanin molecule, which can rapidly quench free radical chain reactions by donating hydrogen atoms.

[0086] In the BK-SP@CMCS system, the antioxidant capacity of the film achieved a qualitative leap. With increasing microreactor concentration, the film's scavenging rates for both DPPH and ABTS free radicals exceeded the 90% threshold. This superior antioxidant barrier stems from multiple synergistic effects. The confined encapsulation effect of the porous microcavities in SP@CMCS greatly protects the structural integrity and chemical activity of thymol during film formation. Simultaneously, the phenolic hydroxyl groups of thymol and the antioxidant sites of anthocyanins produce a significant pharmacodynamic synergistic effect. This continuous and efficient free radical scavenging mechanism provides a solid biochemical guarantee for inhibiting oxidative stress within the packaging microenvironment and significantly extending the shelf life of fruits.

[0087] b. Physicochemical properties and structural characterization of composite thin films

[0088] The surface hydrophilicity and hydrophobicity of packaging films were assessed at room temperature using a contact angle meter (DSA100, Krüss GmbH, Germany) via the seated drop method. A 2 μL droplet of deionized water was accurately aspirated using a microsyringe and placed onto a smooth film surface. After the droplet stabilized, its profile was captured using a high-resolution camera, and the water contact angle (WCA) was calculated using the instrument's built-in software. Three parallel measurements were performed at randomly selected locations for each sample, and the average value was taken.

[0089] In-situ non-destructive testing of the chemical bonding state of the thin film was performed using a Fourier transform infrared spectrometer (Nicolet iS50, Thermo Fisher Scientific, USA) equipped with an attenuated total reflectance (ATR) accessory. The film to be tested was tightly bonded to the surface of an ATR crystal and moderate pressure was applied at 4000–500 cm⁻¹. -1 Within the wavenumber range at 4 cm -1 The system was scanned 32 times at a resolution, with the baseline subtracted from the air background, to analyze the hydrogen bonding and intermolecular interactions between the components.

[0090] Using a high-precision digital micrometer, six different test points were randomly selected on each film to measure the thickness, and the average value was used for mechanical calculations. Subsequently, the film was cut into standard strips (e.g., 50 mm × 10 mm), and a room temperature tensile test was conducted using a computer-controlled electronic universal testing machine (TA.XT Plus, Stable Micro Systems, UK) at an initial gauge length of 30 mm and a tensile rate of 1 mm / s. The system automatically recorded and calculated the tensile strength (TS, MPa) and elongation at break (EB, %) of the film.

[0091] Conclusion: High humidity environments rapidly accelerate the spoilage of fresh agricultural products; therefore, excellent surface hydrophobicity is a core indicator for evaluating the water barrier function of packaging materials. The interfacial wetting kinetics of the film were quantitatively determined using the water contact angle (WCA). Figure 6 It is known that the KGM membrane (konjac glucomannan membrane) has a WCA of only 59.91 ± 1.44° because its sugar ring backbone is rich in a large number of strongly hydrophilic hydroxyl groups, which are very easy to form hydrogen bonds with water molecules. The incorporation of BA slightly increases the WCA to 64.98 ± 0.56° by consuming some of the free hydroxyl groups.

[0092] As the loading of the SP@CMCS microreactor continued to increase, the hydrophobic properties of the system exhibited a significant stepwise leap, with the WCA of the BK-SP@CMCS-3 (anthocyanin-konjac glucomannan / spirulina-thymol@carboxymethyl chitosan-3) group reaching a peak of 74.85 ± 0.78°. This dramatic change in interfacial wettability is dominated by a dual chemical and physical mechanism: at the chemical level, the core material thymol contains strongly hydrophobic isopropyl and benzene ring structures, which reduces the free energy of the material surface at the molecular level; at the physical level, as shown by the SEM results, the spiral microstructure of SP@CMCS constructs a micro-nano rough composite interface on the membrane surface similar to the "lotus effect." According to the Cassie-Baxter wetting model, this rough structure can effectively trap air, significantly reducing the actual contact area between water molecules and the film surface, thereby achieving excellent macroscopic water-blocking performance.

[0093] Fourier transform infrared spectroscopy (e.g.) Figure 7 This revealed complex non-covalent interactions between the components. Compared to the pure components, the composite film showed better performance at 3200–3400 cm⁻¹. -1 The stretching vibration peaks of hydroxyl (-OH) and amino (-NH2) groups in the region show significant broadening accompanied by wavenumber shifts, while the peak at 1055 cm⁻¹... -1 The characteristic peaks near the COC glycosidic bond underwent morphological evolution. This confirms that a highly dense three-dimensional hydrogen bond network is constructed between the polysaccharide backbone of KGM (konjac glucomannan), the phenolic hydroxyl groups of BA (blueberry anthocyanins), and the amino / carboxyl groups of CMCS (carboxymethyl chitosan).

[0094] In the actual stress environment of food cold chain packaging, the film needs to possess both sufficient rigidity (resistance to deformation) and toughness (absorption of impact energy). For example... Figure 8 As shown, the film thickness exhibits a significant stepwise increase with the increase of microreactor loading, reflecting the physical filling effect of solid particles on the free volume of the polymer network.

[0095] In terms of mechanical behavior, the KGM membrane exhibits elongation at break (EB) and tensile strength (TS) as low as 2.17% and 5.89 MPa, respectively (e.g., Figure 9 and Figure 10The composite film exhibits typical hard and brittle characteristics. After introducing BA and integrating SP@CMCS, the mechanical properties of the composite film achieved a synergistic leap, with EB and TS significantly increasing to 3.94% and 13.37 MPa, respectively. The intrinsic mechanism of this mechanical strengthening stems from two aspects: firstly, the rigid SP@CMCS microparticles, as active components, are uniformly dispersed in the continuous phase, effectively passivating the microcrack tips during stress and hindering rapid crack propagation; secondly, thanks to the high-density hydrogen bonds and electrostatic network confirmed by FT-IR, when the material is subjected to external tensile forces, these reversible non-covalent bonds can dissipate a large amount of strain energy through a fracture-reorganization mechanism, achieving efficient stress transfer between interfaces, thereby endowing the composite film with excellent comprehensive mechanical properties of strengthening and toughening.

[0096] c. Evaluation of the antibacterial properties of the composite film

[0097] To quantitatively evaluate the in vitro antibacterial efficacy of the composite film against typical foodborne pathogens such as Escherichia coli and Staphylococcus aureus, this experiment strictly employed the plate count method. Equal-area film samples, pre-sterilized on both sides by ultraviolet light, were completely immersed in an initial concentration adjusted to approximately 1.0 × 10⁻⁶. 6 A fresh bacterial suspension in logarithmic phase (CFU / mL) was incubated at 37 °C with shaking in a constant-temperature shaker for 12 h. Subsequently, an appropriate amount of the co-cultured bacterial suspension was aseptically serially diluted tenfold. 100 μL of each appropriate dilution was accurately measured and evenly spread onto a nutrient agar plate. After incubation at 37 °C upside down for 24 h, the total number of viable colonies (CFU) was counted, and the quantitative antibacterial rate of the membrane was calculated by comparing it with a blank control group without a membrane.

[0098] Conclusion: From the quantitative statistical bar chart of antibacterial rate (e.g. Figure 11 and Figure 12 In the studies, it was clearly observed that the KGM membrane (konjac glucomannan membrane) group had almost no antibacterial ability, and the surface of the culture dish was covered with dense pathogenic colonies, which is consistent with its characteristic as a natural polysaccharide that is easily utilized by microorganisms as a carbon source. After the introduction of BA, the BA-KGM membrane (anthocyanin-konjac glucomannan membrane) group showed moderate antibacterial activity, and produced a certain degree of inhibition against both strains. However, after integrating the SP@CMCS intelligent slow-release microreactor, the BK-SP@CMCS-3 (anthocyanin-konjac glucomannan / spirulina-thymol@carboxymethyl chitosan-3) group showed extremely excellent broad-spectrum bactericidal efficacy, and the inhibition rate against E. coli and S. aureus both showed a stepwise increase.

[0099] The superior antibacterial performance of this composite system is not due to a single effect, but rather stems from a multi-dimensional synergistic biochemical mechanism involving physical adsorption, membrane penetration, and metabolic interference constructed from multiple components. Firstly, the polymer CMCS in the microreactor shell has a high density of protonated amino groups (-NH3) on its molecular chains. + SP@CMCS can strongly electrostatically target and adsorb onto the negatively charged surface of bacteria, thereby blocking the exchange of nutrients between bacteria and the external environment. Secondly, THY, the core antibacterial agent, is released responsively by SP@CMCS. Its highly lipophilic isopropyl and benzene ring structure allows it to easily penetrate the bacterial cell wall and seamlessly insert into the phospholipid bilayer of the cell membrane. This insertion drastically disrupts lipid arrangement, destroys cell membrane permeability and transmembrane proton kinetics, leading to a massive leakage of key intracellular substances and ultimately causing irreversible bacterial lysis and death. Furthermore, the polyphenolic structure in the BA molecule can inactivate key extracellular bacterial enzymes through hydrogen bonding. The synergistic effect of these three mechanisms, combined with the long-lasting sustained-release properties of gas-phase THY, provides a powerful packaging defense against the spread of microorganisms that cause fruit and vegetable spoilage.

[0100] d. Experiments on strawberry preservation and determination of quality indicators

[0101] Fresh strawberries used in this experiment were purchased from a fruit and fresh produce supermarket in Hefei, Anhui Province. Fruits of uniform size, without mechanical damage, and of consistent ripeness were selected. After washing with pure water and air-drying, they were randomly divided into a blank control group and groups packaged with KGM film (konjac glucomannan film), BA-KGM film (anthocyanin-konjac glucomannan film), and BK-SP@CMCS-3 (anthocyanin-konjac glucomannan / spirulina-thymol@carboxymethyl chitosan-3). All were stored in a 25 ℃ constant temperature and humidity chamber for 6 days. Macroscopic decay and browning processes of the strawberries were photographed in situ on days 0, 2, 4, and 6 of storage using a high-resolution digital camera.

[0102] Using a precision analytical balance, the strawberry samples in each group were weighed at the set monitoring time points. The cumulative weight loss rate was obtained by calculating the percentage of the weight difference before and after storage relative to the initial weight.

[0103] The surface reflectance colorimetry of randomly selected strawberry sections at the equator was measured using a colorimeter, with particular emphasis on recording the L value, which represents brightness / lightness. * The value was used to quantitatively assess the degree of gloss degradation and browning of the fruit peel. Subsequently, a texture analyzer equipped with a P / 2 cylindrical metal probe was used to perform a puncture test on the equatorial part of the strawberry, and the maximum yield force when the probe penetrated the pulp was recorded as the fruit hardness (g).

[0104] Strawberry pulp was randomly cut from each group, placed in a mortar and thoroughly homogenized, then squeezed and filtered through multiple layers of defatted gauze to obtain clear juice. An appropriate amount of juice was dropped onto the detection prism surface of a digital handheld refractometer, and the mass fraction of soluble solids (TSS) was read after the instrument automatically compensated for temperature stabilization.

[0105] Conclusion: Strawberries have high post-harvest water content and are highly susceptible to infection by pathogens, leading to soft rot. For example... Figure 13 As shown, during storage at 25 ℃, the fruit in the blank control group (unpackaged) developed extensive mold, softened, and released juice by day 4, losing its commercial value. While the KGM and BA-KGM film-packaged groups showed slight delays in the early stages, obvious rotten patches still appeared by day 6. In contrast, the BK-SP@CMCS-3 group maintained a bright red and plump healthy phenotype throughout the entire 6-day storage period, without significant mold or collapse, demonstrating that the thymol (THY) released slowly by the microreactor and the dense film matrix formed a highly efficient synergistic preservation barrier. Simultaneously, the smart film exhibited a sensitive microenvironment-responsive color-changing function (e.g., Figure 13 (Right side of the middle section) As strawberries respire and decay, volatile acidic and alkaline gases such as CO2 accumulate inside the packaging, altering the pH value of the microenvironment and causing a visible color change in the BA-KGM and BK-SP@CMCS films. This optical gradient closely matches the macroscopic decay process of the fruit, successfully enabling real-time, non-destructive visual monitoring of strawberry freshness.

[0106] To further elucidate the preservation mechanism of the BK-SP@CMCS-3 system, the physicochemical and microbiological indicators of strawberries during storage were quantitatively measured. With prolonged storage, strawberries continuously underwent water transpiration and tissue degradation. For example... Figure 14 and Figure 15 As shown, the blank control group experienced severe weight loss and rapid softening; while the BK-SP@CMCS-3 group, thanks to the excellent water-blocking barrier effect of the film and the inhibition of pectinase metabolism by THY, effectively locked in tissue moisture, minimizing the rate of fruit firmness decline.

[0107] Surface color (L) * ) and soluble solids (TSS) directly reflect the browning and nutrient consumption of fruit. The blank control group showed L in the later stages of storage * Both TSS and TSS decreased sharply, while the BK-SP@CMCS-3 group effectively inhibited the respiratory peak and oxidative stress (e.g., TSS) in strawberries. Figure 16 and Figure 17 This significantly slows down the metabolic consumption of substrates such as sugars, maintaining the bright color of the fruit. Microbial proliferation is the core cause of strawberry spoilage.

[0108] In this invention, carboxymethyl chitosan (CMCS) is used as a smart shell to coat spirulina loaded with thymol, constructing a core-shell microreactor with dual pH / humidity response characteristics. This microreactor is then organically combined with a visual sensing matrix constructed from konjac glucomannan and blueberry anthocyanins, forming a synergistic incentive mechanism of "real-time monitoring-active intervention". On the one hand, the thymol released by the microreactor and the anthocyanins in the matrix produce a strong pharmacodynamic synergy in terms of antioxidant and antibacterial functions, jointly and efficiently quenching free radicals in the packaging microenvironment and inhibiting pathogens. On the other hand, when fruits and vegetables rot and produce acidic metabolites and a high-humidity environment, the visual matrix emits an intuitive color warning through the structural transformation of anthocyanins, while the smart microreactor undergoes simultaneous protonation swelling of its shell after receiving homologous stimulation, precisely triggering the accelerated release of thymol. This achieves a high degree of spatiotemporal unity and synergistic effect between sensing warning and active preservation.

[0109] In addition, the inventors of this case also conducted experiments with other raw materials, process operations, and process conditions described in this specification, referring to the aforementioned embodiments, and obtained relatively ideal results in all cases.

[0110] It should be understood that the technical solutions of the present invention are not limited to the specific embodiments described above. Any technical modifications made to the technical solutions of the present invention without departing from the spirit and scope of the claims are within the scope of protection of the present invention.

Claims

1. A method for preparing a smart active composite film with preservation and monitoring functions, characterized in that, include: Thymol was dissolved in anhydrous ethanol to form an active core material stock solution. Spirulina was then added and stirred and reacted at room temperature and in the dark for 12-24 hours. The solution was then centrifuged, washed, and freeze-dried to obtain spirulina loaded with thymol. The mass ratio of spirulina to thymol was 1:1 to 2:

1. A smart fragrance microreactor with dual pH / humidity response characteristics was prepared by coating the thymol-loaded spirulina with carboxymethyl chitosan; wherein the mass ratio of the thymol-loaded spirulina to carboxymethyl chitosan was 1:15 to 1:

20. Furthermore, a film-forming solution containing konjac glucomannan, blueberry anthocyanins, a smart fragrance microreactor, and water is subjected to film-forming treatment to obtain a smart active composite film with preservation and monitoring functions; wherein, the mass ratio of konjac glucomannan, blueberry anthocyanins, and the smart fragrance microreactor is 1:0.2:0.1~0.

3.

2. The preparation method according to claim 1, characterized in that: The mass ratio of spirulina to thymol is 1:1 to 1.5:1; And / or, the concentration of thymol in the active core material stock solution is 0.9~1.1 g / mL.

3. The preparation method according to claim 1, characterized in that, Specifically, it includes: Spirulina loaded with thymol was dispersed in water to form a spirulina suspension loaded with thymol. Carboxymethyl chitosan is dissolved in water to form an aqueous solution of carboxymethyl chitosan; Furthermore, the spirulina suspension loaded with thymol was added dropwise to a carboxymethyl chitosan aqueous solution and stirred for 4-6 hours at room temperature and in the dark. After centrifugation, washing, vacuum freeze-drying, and grinding, an intelligent fragrance microreactor with dual pH / humidity response characteristics was obtained.

4. The preparation method according to claim 3, characterized in that: The intelligent fragrance microreactor has a core-shell structure.

5. The preparation method according to claim 1, characterized in that, Specifically, it includes: Konjac glucomannan was dissolved in water to form a konjac glucomannan solution. Then, blueberry anthocyanins and a smart fragrance microreactor were added and thoroughly mixed and dispersed to form a film-forming solution. Furthermore, the film-forming liquid is degassed, molded, and dried to obtain an intelligent active composite film with preservation and monitoring functions.

6. The preparation method according to claim 5, characterized in that: The concentration of the konjac glucomannan solution was 0.01 g / mL.

7. The preparation method according to claim 5, characterized in that: The drying process was carried out under light-protected conditions at 30-45°C.

8. A smart active composite film with preservation and monitoring functions prepared by any one of claims 1-7.

9. The application of the intelligent active composite film with preservation and monitoring functions as described in claim 8 in the field of fruit and vegetable preservation monitoring.

10. A method for preserving and monitoring fruits and vegetables, characterized in that, include: The intelligent active composite film with preservation and monitoring functions described in claim 8 is used to seal and store fruits or vegetables. The storage environment is characterized by an ambient temperature of 20-25°C and a relative humidity of 70-80%.

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