Intelligent Food Packaging Films Based on pH-Responsive Eugenol@ZIF-8/PVA-HACC with Enhanced Antimicrobial Activity

Abstract

Natural antibacterial food packaging materials endowed with environmental responsiveness are garnering substantial research interest in sustainable food preservation. This study reports the development of a pH-responsive antimicrobial composite film through encapsulation of eugenol—a natural phenolic compound—within zeolitic imidazolate framework-8 (ZIF-8). The engineered eugenol@ZIF-8 system demonstrated pH-dependent release characteristics, with cumulative release reaching 32.2% at pH 6 versus merely 0.61% at pH 7 over 4 h. Subsequent integration of this nanocarrier into a polyvinyl alcohol (PVA)/hydroxypropyltrimethyl ammonium chloride chitosan (HACC) matrix yielded a multifunctional composite film for active food packaging applications. The characterization of film revealed that while eugenol@ZIF-8 incorporation slightly compromised mechanical strength (tensile resistance decreased by 18.7%) and flexibility (elongation at break reduced to 54.3% of control), it significantly enhanced hydrophobicity (water contact angle increased to 92.5°) and thermal stability (decomposition temperature elevated by 34 °C). The composite film demonstrated synergistic antibacterial efficacy through the combined action of Zn²⁺ ions, ZIF-8 nanostructures, and eugenol, achieving 88% inhibition against E. coli. Practical validation through fresh noodle preservation trials confirmed the material’s effectiveness, with the optimized formulation (PVA-HACC-2% eugenol@ZIF-8, PHEZ2) extending shelf life by >5 days compared to conventional packaging. This work establishes a novel strategy for engineering intelligent ZIF-based packaging systems that respond to food spoilage microenvironments, offering significant potential for reducing food loss.

1. Introduction

Foodborne diseases caused by pathogenic microorganisms pose a serious threat to human health and safety. A WHO report indicated that 7.7% of the global population is threatened by foodborne diseases [1]. Appropriate food packaging contributes to alleviating this issue [2,3]. Antibacterial packaging with active agents can preserve food quality and minimize the effects of environmental factors like light, oxygen, and water vapor. More importantly, it effectively suppresses the growth and proliferation of microorganisms, thereby safeguarding food quality and safety [4]. In the food industry, such antimicrobial agents include organic compounds, like quaternary ammonium salts, halogenated compounds, phenols, chitosan, and chitin, or inorganic materials, including nanoparticles of metals and metal oxides [5]. Recently, developing natural product-based antimicrobial agents has represented an effective alternative to chemical antimicrobial agents, given concerns about microbial resistance and potential toxicity [6].

Novel bio-functional film materials with better film-forming and strong antibacterial properties have become a hot research direction in the field of food packaging [7]. Polyvinyl alcohol (PVA) is a non-toxic polymer with excellent hydrophilicity, biodegradability, film-forming ability, and tensile tear resistance that is commonly used in food packaging [8]. However, PVA has poor water resistance and gas barrier properties when used alone as a film, and it does not have antibacterial properties, which limits its applications. In order to improve these characteristics, chitosan, sodium alginate, etc., were blended with PVA, with PVA/chitosan composite films receiving tremendous amounts of attention [9]. Researchers have reported that PVA/chitosan-blended films could be used as antimicrobial packaging material for food preservation, including bread, fruits, etc. [9,10]. Modified chitosan, such as hydroxypropyltrimethyl ammonium chloride chitosan (HACC), often has the advantages of more stable antibacterial properties and better water solubility, and can form complexes with other materials through physical interactions, chemical bonding, and other methods [11]. Hence, in the present study, PVA and HACC were used as film-forming substrates to prepare antibacterial packaging films.

Eugenol can be applied in the food industry due to its broad-spectrum antibacterial and antioxidant properties [12]. However, the application scenarios of eugenol are limited due to its volatility, instability, and susceptibility to environmental factors. Environmental stimulus-responsive release may help improve this issue, and developing a delivery system that can intelligently regulate the release of bactericidal agents is very promising [13]. Several factors could be used to stimulate the release of antibacterial agents in active packaging, including temperature, humidity, light, magnetic fields, pH, etc. [14]. Microbial infection of food can alter the acidity and alkalinity of the microenvironment, and the pH-responsive release of antibacterial agents has significant practical significance in food preservation, especially for perishable foods [15,16]. Zeolitic imidazolate framework-8 (ZIF-8) is a porous metal–organic framework (MOF) composed of Zn²⁺ as a metal ion and 2-methylimidazole as a ligand [17]. ZIF-8 has a large pore size, high specific surface area, and excellent water and alkali stability. Furthermore, ZIF-8 can decompose under acidic conditions; therefore, it can serve as a carrier for acid-sensitive drug release [18,19]. Ref. [20] prepared pH-responsive ZIF-8-PAA-MB@AgNPs@Van-PEG composite nanomaterials, loaded with a broad-spectrum antibacterial agent (methylammonium blue, MB) for the treatment of bacterial infections, and found they were able to inhibit bacterial growth, such as Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus), and have good biocompatibility. The use of ZIF-8 in drug delivery systems has been relatively well studied, although there are few reports on its use as a food packaging material for the encapsulation and responsive release of natural products, such as eugenol.

In the present study, the objective was to synthesize composite materials by encapsulating the eugenol into ZIF-8, with successful encapsulation verified through Fourier Transform Infrared Spectroscopy (FTIR) and Scanning Electron Microscopy (SEM). The pH-responsive release properties were determined by measuring the release amount of eugenol under different pH conditions. In order to improve the applicability of eugenol@ZIF-8, the prepared eugenol@ZIF-8 was mixed with a film-forming solution composed of PVA and HACC, and dried to form a composite film. The mechanical properties, rheological properties, and barrier properties of the composite film were measured and analyzed. The effect of eugenol@ZIF-8 on the water stability and hydrophobicity of the composite films was investigated. A detailed evaluation of the antibacterial properties of the composite film was conducted using the Gram-negative bacterium, E. coli, and the Gram-positive bacterium, S. aureus. Finally, the antimicrobial and freshness-keeping properties of the composite film were evaluated using storage experiments of fresh noodles. This work would provide a new method for the development of fresh food preservation films.

2. Results and Discussion

2.1. Characterization of eugenol@ZIF-8

As shown in Figure 1, under the preparation conditions of this study, the inclusion of eugenol showed no significant effect on the morphology of the metal–organic frameworks, and both demonstrated cruciate flower-like morphologies without significant change. This is consistent with the unchanged crystal structure of the XRD results. The SEM results are similar to the morphological results of the enzyme encapsulated in ZIF-8 reported by [21] and ε-polylysine@ ZIF-8 reported by [22].

The EE of eugenol in eugenol@ZIF-8 was evaluated by determining the OD value at 204 nm of eugenol in the supernatant and calculated using the standard curve, with the EE determined as 65.24%. DSC analysis was used to evaluate the thermal stability of the prepared materials in the present study. According to Figure 2a, the decomposition temperature of ZIF-8 was 257.61 °C, and that of eugenol@ZIF-8 was 296.67 °C, indicating that the addition of eugenol improved the thermal stability of the nanoparticles, also proving the successful encapsulation of eugenol in eugenol@ZIF-8.

Subsequently, FTIR spectra were determined to verify the encapsulation of eugenol and the internal molecular interactions of eugenol@ZIF-8. A comparison of FTIR spectra of ZIF-8 and eugenol@ZIF-8 is shown in Figure 2b. Two similar absorption peaks were detected at 758 cm⁻¹ and 683 cm⁻¹, demonstrating the stretching vibration peaks of the C-H bond. It can be seen that the concentration of the C-H bond in ZIF-8 was lower than that in eugenol@ZIF-8, which may be related to the addition of eugenol. The absorption peak of the C=O bond was detected at 1145 cm⁻¹ in the two samples, and the concentration of C=O in the two samples showed a similar trend to the variation of the C-H bond. The absorption peaks detected at 1418 cm⁻¹ and 2988 cm⁻¹ were caused by the stretching vibration of the C-C bond and the bending vibration of the O-H curved surface, respectively, and the vibration intensities of the two samples were almost the same [23]. The absorption band associated with the methoxy group in eugenol@ZIF-8 caused a wavelength shift in comparison to that of ZIF-8 (1100–1050 cm⁻¹), which confirmed a strong interaction between the methoxy group in eugenol and Zn²⁺ ions [24]. Furthermore, the absorption band at 1260–1150 cm⁻¹ in eugenol@ZIF-8 resulted from the -C-O-C- bond of eugenol [25], indicating that eugenol was successfully encapsulated in ZIF-8.

Also, the crystallinity of the synthesized ZIF-8 and eugenol@ZIF-8 was studied by XRD analysis (Figure 2c). The main characteristic peaks of the two samples were between 5 and 20°, the XRD morphology trends of the two samples were similar, the positions of the diffraction peaks were essentially the same, and there were no new diffraction peaks after the addition of eugenol, indicating that the introduction of eugenol had no effect on the crystal structure of ZIF-8 and had good biocompatibility. Similarly, the addition of ε-polylysine or riboflavin sodium did not change the crystal structure of ZIF-8 in the composite nanoparticles [22,26].

2.2. Release Properties

To evaluate the pH-responsive release properties of the eugenol in eugenol@ZIF-8 nanoparticles, in vitro release experiments were conducted in PBS at pHs of 5.0, 6.0, and 7.0. The amount of eugenol released was measured using UV–vis absorption spectra of eugenol at 204 nm and calculated by a standard curve. As shown in Figure 3, in the acidic environment of pH 5 and 6, the release rate of eugenol was significantly greater than that of pH 7. After 4 h of release, the amount of eugenol released in a pH 5 environment was 28.7%, with 32.2% at pH 6, and only 0.61% in the neutral environment. The eugenol release amount slowed gradually and tended to reach a plateau after 12 h, with significant pH-responsive release behavior still maintained for eugenol@ZIF-8 throughout the whole process. The porosity of eugenol@ZIF-8 was inferred to be partially retained after eugenol encapsulation, consistent with the well-documented behavior of ZIF-8-based active agent composites. Previous studies have characterized ZIF-8 composites loaded with phenolic compounds via BET analysis, demonstrating that the encapsulation of hydrophobic phenolics into ZIF-8 pores did not destroy the inherent porous framework of ZIF-8. This preserved porosity is critical for ensuring the pH-responsive release of encapsulated active ingredients. These phenomena indicated that encapsulating eugenol with ZIF-8 as a delivery system could achieve a pH response and controllable release properties. The reason for this result may be that under acidic conditions, 2-methylimidazole was protonated, leading to the destruction of the coordination between zinc ions and the imidazole ring and the partial decomposition of zif8, resulting in a large amount of eugenol released [27]. Furthermore, the microenvironment of food storage could cause a decrease in environmental pH due to the respiration of microorganisms, such as carbon dioxide gas. In addition, the decomposition of food components, including glycogen, could also produce organic acids, leading to an acidic storage microenvironment [28]. Therefore, the prepared eugenol@ZIF-8 with pH-responsive release properties could contribute to achieving long-term antibacterial effects in food storage.

2.3. Characterization of the Composite Films

As shown in Figure 4a, the swelling ratio of the films decreased significantly with an increase in the amount of eugenol@ZIF-8 (p < 0.05). The swelling ratio reflected the water absorption and holding capacity of the films. The different swelling ratio of the films was affected by the composition and the hydrophilicity, and the reduction could be owing to the hydrophobic characteristic of ZIF-8 [29]. It was found that the WCA of the composite films increased with an increasing amount of eugenol@ZIF-8 (Figure 4b). The WCA of PHEZ0 was 53.24°, which was related to the hydrophilicity of PVA and HACC. Moreover, ZIF-8 was hydrophobic, and the WCA of the composite films increased with the addition of eugenol@ZIF-8, whereby the WCA values of PHEZ1 and PHEZ2 were 82.15° and 83.21°, respectively. The increase in WCA due to the hydrophobicity of ZIF-8 has also been reported in other literature [30,31]. As a food packaging material, the weaker the water absorption and water-holding capacity of a composite film, the easier it is to limit the water interaction between the food substrate and the film. The WVT results showed that after adding eugenol@ZIF-8 to the film, the amount of water vapor passing through increased significantly compared to the PVA-HACC film (Figure 4c). Due to the high hydrophobicity of ZIF-8 and its distribution throughout the entire film matrix, the insertion of eugenol@ZIF-8 into the film loosened the bonds between polysaccharide molecules, causing the network gaps in the film to become larger, increasing the volume of water vapor vacancy transport, and increasing the WVT. The higher WVT was conducive to the exchange of water vapor inside and outside the film, which is beneficial for food preservation, especially fresh food [22].

Visually, the inclusion of eugenol@ZIF-8 had little effect on the film’s appearance. However, the introduction of eugenol@ZIF-8 had a significant effect on the thickness of the film, with a range of 0.23~0.55 mm for PHEZ0–PHEZ2 (Figure 5a). Furthermore, eugenol@ZIF-8 significantly affected the mechanical properties of the film. The tensile strength of the PVA-HACC film was 2.37 MPa, and it decreased to 0.76 MPa with the addition of 2% eugenol@ZIF-8. There was a tight network structure between PVA and HACC, which was due to the formation of hydrogen bonds between the hydroxyl groups of PVA and the amino groups of HACC [9]. The introduction of eugenol@ZIF-8 could reduce the hydrogen bonding force. Moreover, due to the surface tension and electrostatic force, there was an accumulation of eugenol@ZIF-8 within the film, which also affected the continuity of the composite film, and eventually led to the reduction in tensile strength and resistance to damage [32].

Additionally, the films containing eugenol@ZIF-8 exhibited lower elongation-at-break values compared with the PVA-HACC film, ranging from 42% to 33.3% for PHEZ0–PHEZ2 (Figure 5b). The lower elongation-at-break values reflected the reduced flexibility of the film [33]. The results indicate that the introduction of eugenol@ZIF-8 has a certain negative effect on the mechanical behavior of the composite films. Further research is needed to improve these aspects using other methods.

2.4. Rheological Properties

The dynamic rheological behaviors of the film-forming solution with different concentrations of eugenol@ZIF-8 are shown in Figure 6. As can be observed, the storage modulus (G’) of the film-forming solution was higher than the loss modulus (G’’), and the G’ and G’’ of PVA-HACC and PVA-HACC-eugenol@ZIF-8 demonstrated an ascending order, which indicated that all film-forming solutions maintained an ideal elastic state over the applied frequency range [34,35]. Moreover, due to the addition of eugenol@ZIF-8, especially in the amounts of 1·% and 2%, the storage modulus of the film-forming solution increased significantly and showed an upward trend, indicating that eugenol@ZIF-8 may enhance the gel structure and improve the elastic properties of the system. Similar results were shown regarding the effect of Ag@MOF on the rheological properties of PVA/chitosan composite hydrogel and the effect of ZIF-8 on the storage modulus of konjac glucomannan/ZIF-8 hydrogel [36,37].

As shown in Figure 6d, the viscosity of the film-forming solution decreased with the increase in shear rate, exhibiting the shear-thinning behavior of non-Newtonian fluids. This result may be related to the fact that the network structure between the molecules of the film-forming solution was seriously damaged due to the increase in the shear rate during the shearing process, and the fracture rate was greater than the recombination rate, resulting in a decrease in flow resistance and viscosity between the molecular particles. In addition, the viscosity of the film-forming solution increased with the increase in eugenol@ZIF-8 content, which was similar to that of G’ and G’’. The result also indicated that the incorporation of eugenol@ZIF-8 did not significantly diminish the rheology behaviors of the composite films. Similarly, the addition of ZIF-8 did not show a significant effect on the rheological properties of ZIF-8@alginate solutions.

2.5. Antibacterial Activities

The antibacterial activities of the PVA-HACC films (PHEZ0, PHEZ1, and PHEZ2) were evaluated using the plate method, as shown in Figure 7a. Although the PVA-HACC film has a certain bacteriostatic effect due to the presence of HACC, the introduction of eugenol@ZIF-8 significantly improved the antibacterial efficiency. There were a large number of colonies of E. coli and S. aureus in the control group, while the colonies in the PVA-HACC (PHEZ0), PHEZ1, and PHEZ2 treatment groups gradually decreased compared with CK. The colony-forming units of E. coli were nearly 88% prevented in the PHEZ2 group compared with the CK. Moreover, both PVA-HACC and PHEZ2 showed higher inhibitory efficiency against the Gram-negative bacterium E. coli compared to the Gram-positive bacterium S. aureus. The reason for this phenomenon may be due to differences in cell wall composition between Gram-negative and Gram-positive bacteria [38]. The peptidoglycan layer on the cell wall of Gram-positive bacteria was thick and hard, while the lipopolysaccharide layer on the cell wall of Gram-negative bacteria was thinner [39]. Hence, E. coli may be more sensitive to eugenol and the cationic groups of HACC and more susceptible to cell wall rupture, thereby enhancing its antibacterial properties. Furthermore, the metal ions (Zn²⁺) released from ZIF-8 were able to impair cell membranes, which resulted in the cell death of bacteria [40,41]. As a result, the prepared composite film had a synergistic antibacterial effect on ZIF-8, HACC, and eugenol. Furthermore, the SEM images (Figure 7b) showed that compared to the untreated group, the appearance of E. coli and S. aureus underwent significant changes after treatment with PHEZ2. The surface roughness of the bacterial cells increased, wrinkles formed, and the bacterial structure was destroyed. These morphological changes indicate that the antibacterial agent PHEZ caused serious damage to E. coli and S. aureus.

Regarding the measurement of the growth of bacteria in liquid media, Figure 8 shows the effect of the film-forming solution on the bacterial growth kinetics of E. coli and S. aureus. The cell growth of the PHEZ0 group was slightly lower than that of the control group, indicating the antibacterial activity of HACC against the two bacteria. Also, the OD₆₀₀ value of E. coli was lower than that of S. aureus, indicating that the inhibition efficiency of PHEZ0 on Gram-negative bacteria was higher than that of Gram-positive bacteria. PHEZ1 and PHEZ2 were able to effectively inhibit the growth of E. coli and S. aureus within 16 h. Then, the OD₆₀₀ value of all groups increased again, while the OD₆₀₀ value of PHEZ2 was the lowest, followed by PHEZ1 at 24 h, which also verified the synergistic bacteriostatic effect of the composite film. Compared with the control group, PHEZ2 had an excellent inhibitory effect on E. coli and S. aureus, whereby its OD₆₀₀ value was reduced by 87.8% and 83.6%, respectively. The results demonstrate that the PHEZ film has excellent antibacterial activity, which is crucial in food preservation.

2.6. Application of the Composite Film in Fresh Noodle Preservation

Fresh noodles are the product of wheat processing and are favored by consumers because of their freshness, good taste, and high nutritional value. However, due to their high moisture content and abundant carbon source, they are highly susceptible to microbial invasion, which can affect storage quality and shelf life [42]. Generally speaking, the TPC of 10⁶ CFU/g was considered the boundary point for the spoilage and deterioration of fresh noodles. The initial TPC of fresh noodles was nearly 10³ CFU/g, which is similar to a previous study [16,43]. For the control group, the TPC of fresh noodles on the second day had exceeded 10⁶, indicating that the shelf life was less than 2 days. However, the group of PHEZ0, PHEZ1, and PHEZ2 all extended the shelf life of noodles due to the antibacterial activities. As shown in Figure 9, compared to PHEZ0, which can extend the shelf life of noodles to 3 days, the introduction of eugenol@ZIF-8 (PHEZ1, PHEZ2) could extend the shelf life to more than 5 days. This indicates that the introduction of eugenol@ZIF-8 enhanced the antibacterial activity, which is consistent with the results of in vitro antibacterial experiments.

Compared with the control group, there were significant differences in the TPC of the treatment group. Specifically, on the fifth day, there was a significant difference in the TPC of PHEZ1 and PHEZ2, which had not been observed on previous days. This may be related to the content of eugenol released by the composite membrane in the noodle storage microenvironment. Ref. [44] studied the pH changes during the fresh noodles storage, and the results showed that the pH values of fresh noodles decreased to 6 on day 2. With the growth of microorganisms and the decomposition of nutrients, the microenvironment of noodle storage is prone to acidity. According to the release properties, the PHEZ composite film exhibited an acid-responsive release property; thus, after 2 days of storage, PHEZ2 released a greater amount of eugenol, resulting in a more pronounced antibacterial effect.

Color serves as a critical determinant of noodle quality, and consumers typically exhibit a preference for fresh noodles that possess a visually appealing appearance [45]. In color assessment experiments, the L* value was employed to measure the brightness of fresh moist noodles, with higher L* values corresponding to samples with greater luminosity and visual appeal. As shown in Figure 9b, while the noodles demonstrated a marked reduction in the L* value by day 5, the environment of the PHEZ2 film significantly delayed the degree of darkening. This decline in L* value could be attributed to the results of the TPC, as an increase in TPC may lead to surface darkening of the noodles and a corresponding decrease in L* value. The b* value increased as storage time increased for nearly all groups, suggesting an increase in the yellowness of the noodles. The increase in b* values for noodles was delayed when packaged by PHEZ films, especially the PHEZ2 film, which may be related to properties such as antibacterial properties and water permeability. Similar phenomena have also been reported in other fresh noodle packaging films [46]. Therefore, the prepared acid-sensitive antibacterial composite film (PHEZ2) could help extend the shelf life of fresh noodles and improve the quality of noodles to some extent during storage.

3. Materials and Methods

3.1. Materials

Eugenol with a concentration of 99% was purchased from Aladdin Chemistry Co., Ltd. (Shanghai, China). Zinc nitrate hexahydrate, 2-methylimidazole, methanol, and glycerin of analytical grade were obtained from Macklin Biochemical Technology Co. (Shanghai, China). Chitosan quaternary ammonium salt (HACC), with a degree of substitution of 95%, and Tween-80 were purchased from Macklin Biochemical Technology Co. (Shanghai, China). Phosphate Buffer solution (PBS) was obtained from Yida Biochemical Technology Co. (Quanzhou, China). Polyvinyl alcohol (PVA) was purchased from Sino pharm Reagent Co. LTD (Shanghai, China). Luria–Bertani (LB) broth medium and nutrient agar medium were purchased from Hope-bio technology LTD (Qingdao, China). E. coli and S. aureus strains were preserved in our laboratory and used after activation. The experimental water was ultrapure water.

3.2. Synthesis of ZIF-8 and eugenol@ZIF-8

The synthesis of ZIF-8 and eugenol@ZIF-8 was based on a previously reported method with some modifications [47]. Zinc nitrate hexahydrate (150 mg) was dissolved in 5 mL of deionized water, 2-methylimidazole (330 mg) was dissolved in 10 mL of methanol, and eugenol (30 mg) was dissolved in 2 mL of methanol. These three solutions were ultrasonically treated at 300 W for 10 min to ensure adequate dissolution. The eugenol solution and zinc nitrate hexahydrate solution were added sequentially to the 2-methylimidazole solution and stirred magnetically for 15 min. During the stirring process, the solution changed from transparent to milky white and cloudy. The resultant mixture was centrifuged at 12,000 rpm for 15 min, washed three times with methanol, and then dried at 50 °C overnight to obtain eugenol@ZIF-8. ZIF-8 was prepared using the same method, except that eugenol was not added to the mixture.

3.3. Characterization of Materials

3.3.1. SEM

The morphology of ZIF-8 and eugenol@ZIF-8 was examined using SEM (TM3000, Hitachi, Tokyo, Japan), and the samples were fixed on a brass stub using adhesive and sprayed with gold prior to the test [31].

3.3.2. FTIR

FTIR analysis was used to verify the combination of eugenol and ZIF-8 within the spectral range of 400–4000 cm⁻¹ (Thermo Scientific Nicolet iN 10, Madison, WI, USA). Appropriate amounts of ZIF-8 and eugenol@ZIF-8 nanoparticles were taken as samples, which were thoroughly mixed with KBr and then scanned using a spectrometer [29].

3.3.3. DSC

The thermal stability of the sample was analyzed using a differential scanning calorimeter (DSC). ZIF-8 and eugenol@ZIF8 nanoparticles were sealed in an aluminum pot, and the samples were analyzed at a flow rate of 25 mL/min, a scanning temperature range of 25–400 °C, and a scanning speed of 5 °C/min under a nitrogen atmosphere (DSC25, TA instruments, New Castle, DE, USA) [41].

3.3.4. XRD

The crystal structures of ZIF-8 and eugenol@ZIF-8 were determined using X-ray diffraction (XRD) with Co Kα radiation in the 2θ range of 5–60°, and a scan speed of 5°/min (Bruker D8 advance, Karlsruhe, Germany).

3.3.5. Encapsulation Efficiency

The encapsulation efficiency (EE, %) was evaluated by the absorbance method [31]. The wash supernatant in the preparation process of eugenol@ZIF-8 was collected, the absorbance of the collected supernatant at 204 nm was determined, the content of eugenol was determined by a standard curve (prepared by dissolving eugenol in methanol), and the EE was calculated using the following formula:

$$\mathrm{E}\mathrm{E}\%={\displaystyle \frac{m_{eugenol}-m_{eugenol in supernatant}}{m_{eugenol}}}\times 100\%$$

(1)

where EE% is the encapsulation efficiency and $m_{eugenol}$ is the weight of eugenol used in the preparation process.

3.4. Evaluation of Release Properties

The pH-responsive release properties of eugenol were evaluated according to a previously reported procedure [48]. Specifically, 10 mg of eugenol@ZIF-8 was dispersed in 20 mL of PBS (pH = 5, 6, 7). The solutions were subjected to agitation at 37 °C (200 rpm), 1 mL of the release supernatant was taken out at a certain interval, and 1 mL of fresh PBS solution was replenished. The OD value at 204 nm was determined by an ultraviolet spectrophotometer (UV-1780, SHIMADZU Corporation, Kyoto, Japan). The release rate was calculated based on the standard curve of eugenol.

3.5. Preparation and Characterization of Composite Films

The preparation of composite films was achieved by compounding the film-forming solution (PVA and HACC) with different proportions of eugenol@ZIF-8, followed by drying. PVA (1.8 g) was swollen in 28.2 g of water for 15 min and kept in a water bath at 90 °C for 2 h to form a completely transparent solution. HACC (0.1 g) was dissolved in 100 mL of water under magnetic stirring. The PVA solution (6% w/v) and the HACC solution (0.1% w/v) were mixed thoroughly at a ratio of 4:1 to prepare the film-forming solution. Subsequently, glycerin (1% w/v) and Tween-80 (0.1% w/v) were added to the mixed solution as plasticizers and stabilizers. Different concentrations of eugenol@ZIF-8 (0% w/v, 0.5% w/v, 1% w/v, 2% w/v) were added to the film-forming solution and dispersed evenly after stirring. The film-forming solutions of each group were subjected to ultrasonic treatment (300 w for 10 min) to ensure uniform dispersion of the components and removal of excess bubbles. Then, the film-forming solution (10 mL) was poured into an acrylic mold (5 cm × 5 cm) and vacuum dried at 40 °C for 5 h.

3.6. Determination of Rheological Properties

The rheological properties, such as the storage modulus and loss modulus, are key parameters for evaluating the physical properties of film-forming solutions. The rheological properties of film-forming solutions were investigated via stress scanning and frequency scanning in a rotating rheometer (HR10, TA instruments, USA). The film-forming solution containing different concentrations of eugenol@ZIF-8 (0%, 0.5%, 1%, and 2%) was dripped onto the platform, the residual pressure was relaxed, and the spilled solution was scraped off. The parameters of the rheometer were set as follows: a plate diameter of 40 mm, a gap of 250 mm, and a temperature of 25 °C. Then, the storage modulus, loss modulus, and apparent viscosity of the solution were determined.

3.7. Mechanical Properties of Composite Films

The thickness of the composite film was measured using a spiral micrometer (Deli, Ningbo, China). The thickness was measured at 8 random locations from 3 pieces of film, and the average value was taken as the film thickness. The mechanical properties of the composite films, including tensile strength and elongation at break, were measured by a texture analyzer (TA.XTC-18, Shanghai Baosheng instrument, Shanghai, China). The load was 30 kg, and the film was cut to a width of 10 mm and a length of 50 mm and fixed on the test grips. The initial gripper clearance was set to 30 mm, and the crosshead speed was maintained at 50 mm/min [31].

3.8. Swelling Ratio, Water Contact Angle, and Water Vapor Transmittance

The moisture sensitivity was evaluated by measuring the swelling ratio and water contact angle (WCA). In the swelling ratio measurement [49], one piece of the composite film was weighed and then immersed in 25 mL of deionized water for 24 h at room temperature. Then, the composite film was removed from the deionized water, the excess water on the surface was removed by the filter paper, and the film was weighed again. The swelling ratio of the composite film was calculated as follows:

$$\mathrm{s}\mathrm{w}\mathrm{e}\mathrm{l}\mathrm{l}\mathrm{i}\mathrm{n}\mathrm{g} \mathrm{r}\mathrm{a}\mathrm{t}\mathrm{i}\mathrm{o}={\displaystyle \frac{W_s-W_i}{W_i}}\times 100\%$$

(2)

where $W_i$ is the initial weight of the dry sample and $W_s$ is the weight of the film in a swollen state.

The WCA of the prepared composite film was measured using a contact angle meter (Dataphysics OCA20, Filderstadt, Germany). A drop of deionized water was dropped vertically on the surface of the film, and images were obtained for further analysis. The sample to be tested was kept flat and dry.

The moisture permeability was evaluated according to the water vapor transmittance (WVT). The WVT of the composite films was determined by the gravimetric method with slight modification based on the standard method of GB/T1037–2021 [50]. Anhydrous calcium chloride was added to the bottle at a distance of 1 cm from the bottle mouth, and the bottle mouth was sealed with the prepared composite film containing petroleum jelly. The diameter of the mouth of the bottle was 1 cm. The flask was sealed in a dryer, and then the saturated sodium chloride solution was poured into the bottom of the dryer to achieve controlled conditions of relative humidity (69%) and temperature (23 °C). After 16 h of equilibration, it was weighed as the initial mass. Finally, the test bottle was placed in a constant humidity chamber, and the interval between the two weight measurements was 24 h. The equilibrium state was achieved when the mass difference between the two weights was less than 5 wt%.

The WVT of each sample was obtained from the average of three specimens. The WVT was calculated using the following formula:

$$\mathrm{W}\mathrm{V}\mathrm{T}={\displaystyle \frac{∆m\times d}{A\times t}}$$

(3)

where $∆m$ (g) is the mass difference between two consecutive measurements within the time of t at the equilibrium state, $d$ (cm) is the thickness of the film, $A$ (cm²) is the test area, and $t$ is the time interval between two consecutive measurements.

3.9. Evaluation of Antibacterial Activities

Gram-negative bacteria (E. coli) and Gram-positive bacteria (S. aureus) were used to evaluate the antibacterial efficiency of the composite films according to a previously reported method [51] with slight modifications. Single colonies of E. coli and S. aureus were selected from LB solid agar plates, inoculated in 50 mL of LB liquid medium, and shaken overnight in a biochemical incubator at 37 °C and 180 rpm. The bacterial density was adjusted to 10⁶ CFU/mL, and the bacterial solution was mixed with the film-forming solution to achieve film-forming solutions (PHEZ0, PHEZ1, and PHEZ2) with a concentration of 1 g/L and incubated at 37 °C for 12 h. In the experiment, the film-forming solution was replaced by pure water as the control group (CK). The treated bacterial solution was fold serially diluted, and 50 μL of the bacterial suspension was spread on the prepared LB nutrient agar medium. The colonies were counted after incubation at 37 °C for 12 h. In addition, 1 g/L of the film-forming solution was mixed with bacteria solution, and during the cultivation, the optical density (OD600) of the solution was measured regularly to determine the bacterial growth kinetics. Before the experiment, all dry samples underwent UV sterilization treatment.

For SEM observations, the bacterial suspensions (E. coli and S. aureus) were combined with the PHEZ2 dispersions and then incubated at 37 °C for 4 h. Bacterial cells were collected via centrifugation and processed through a series of steps. Initially, the collected bacterial pellets were washed and fixed with 2.5% glutaraldehyde at 4 °C for 2 h. Subsequently, they underwent dehydration using a graded ethanol series (30%, 50%, 70%, 90%, and 100% ethanol), with each ethanol concentration applied for 15 min. Finally, the samples were lyophilized under vacuum to prepare them for SEM observation.

3.10. Effect of Composite Films on Shelf Life of Fresh Noodles

3.10.1. Microbiological Analysis

In order to evaluate the antimicrobial activities of the composite films in food storage, a simulated storage experiment using fresh noodles was carried out. The fresh noodle samples were prepared according to the method described by [16]. The resultant noodles were 1.2 mm in width and 0.9 mm in thickness. The same weight of fresh noodles (20 g) was wrapped in different composite films and then stored at 25 °C and 55% RH. The commercial PE (Polyethylene) films served as the control group. The fresh noodle samples were removed regularly to determine the total plate counts (TPCs) by the method in GB/T 4789.2–2016 [52] to evaluate the shelf life of fresh noodles. Specifically, the samples were incubated in sterile saline for 3 min via a homogenizer. Then, the dilutions of the incubation solution were mixed with nutrient broth agar and transferred to the incubator at 37 °C and 90% relative humidity. The TPCs were determined after 48 h.

3.10.2. Fresh Noodle Color Measurements

The color of fresh noodles was analyzed using a colorimeter (CR-400, Konica Minolta Holdings, Tokyo, Japan), and the corresponding L* and b* values were recorded.

The hardness of the noodles was evaluated by placing three parallel noodle strips on a testing surface. Hardness measurements were performed using a P36/R probe (TA.XTC-18, Shanghai Baosheng instrument, Shanghai, China), which compressed the noodles to a 75% strain level. The testing procedure comprised three stages—pre-test, test, and post-test—conducted at a constant speed of 1.0 mm/s. For the tensile strength assessment, a noodle segment approximately 10 cm in length was attached to an A/SPR probe and stretched until fracture. The peak force recorded during the stretching process was defined as the tensile strength. The test was performed at a speed of 2.0 mm/s with a trigger force of 5.0 g, matching the parameters used in the hardness test [53]. Each measurement was repeated eight times to ensure reproducibility and reliability of the results.

3.11. Statistical Analysis

In the present study, each experiment was repeated three times, and the experimental data was expressed as means ± standard deviations (SDs). IBM SPSS 20 was employed for the ANOVA analysis, and ORIGIN 18.0 was utilized for data visualization. Duncan’s test was performed to identify significant differences, and p-values < 0.05 were considered statistically significant.

4. Conclusions

In summary, novel eugenol@ZIF-8 with pH-responsive release and antimicrobial activity was synthesized in the present study. The successful encapsulation of eugenol was confirmed by FTIR, DSC, and XRD. The eugenol@ZIF-8 exhibited significant pH-responsive release characteristics, with 32.2% eugenol released within 4 h at pH 6, compared to only 0.61% released at pH 7. The composite film formed by mixing eugenol@ZIF-8 with PVA/HACC was designed to compensate for the shortcomings of a single component, and the PHEZ2 composite film showed enhanced hydrophobicity and stability. However, the film containing eugenol@ZIF-8 displayed slightly reduced damage resistance and flexibility. Crucially, the prepared composite film PHEZ2 demonstrated an excellent synergistic antibacterial effect of Zn²⁺, ZIF-8, and eugenol. The composite film showed better antibacterial efficiency against Gram-negative bacteria compared to Gram-positive bacteria, and the inhibition rate on E. coli reached 88%. In addition, the antibacterial effect of the composite film was verified by the fresh noodles’ preservation, and compared with untreated noodles, the TPC of fresh noodles indicated that the PHEZ2 composite film could extend the shelf life of noodles by more than 5 days. The present study introduces an approach for utilizing the natural agent eugenol encapsulated in MOFs with pH-responsive properties in food packaging films. While this study demonstrates effective short-term pH-responsive antimicrobial activity, future research should investigate the long-term storage stability of the composite films under various environmental conditions and expand the inhibitory effect of this pH-responsive system to a broader range of microorganisms to fully assess their commercial applicability.