Hydrophobicity, UV resistance, and antioxidant properties of carnauba wax-reinforced CG bio-polymer film

2.1 Materials

The Carnauba wax (melting point: >83°C) was purchased from Macklin Co., Ltd. Chitosan was purchased from Shanghai Sigma-Aldrich Biochemical Technology Co., Ltd. The deacety-lation degree of chitosan ≥ 95%. Vanillin (melting point: 81°C, assay: 99.5%,) was purchased from Macklin Co., Ltd. Glycerol, tween 80, acetic acid and absolute ethanol (all analytical grade) were derived from Macklin Co., Ltd. Gelatin (glue strength ∼250 g Bloom) were purchased from Macklin (Shanghai) Co., Ltd.

2.2 Preparation of carnauba wax/chitosan/gelatin (CGVC) films

The CGV film (carnauba wax content is zero) was prepared by our previous work (21). Similarly, versatile carnauba wax/chitosan/gelatin bio-polymer film (CGVC) was prepared by solution blending method. Briefly, 0.212 g of chitosan was dissolved in a 1.5% v/v acetic acid aqueous solution and stirred in a water bath at 60°C to form a 20 wt% chitosan solution. It stood overnight at room temperature until the bubble disappeared. 0.100 g of carnauba wax and 5 mL of ethanol were mixed in a magnetic mixer for 3 h. 0.848 g of gelatin and 10 mL of deionized water were stirred for 30 min using a water bath set at 60°C to obtain a gelatin solution. The aforementioned chitosan solution, carnauba wax, gelatin solution, and tween-80 as surfactant (2 wt% of total mass) were added to a 100 mL triple neck flask and continuously stirred at 60°C for 1 h; then, the solution of vanillin and glycerol (10 and 30 wt% of total mass, respectively) was mixed, and the reaction was conducted for 3 h at 60°C at 200 rpm. The mixture was placed in a silicone mold measuring 140 × 20 mm and dried naturally for 48 h, until the weight of the film showed a minimal change. The CGVC films with different carnauba wax contents 5, 10, 15, and 20 wt% were successfully prepared and designated as CGVC₅, CGVC₁₀, CGVC₁₅, and CGVC₂₀, respectively.

2.3 Characterization of films

The prepared sample underwent complete drying at ambient temperature. An attenuated total reflectance infrared reflector was employed in the infrared spectroscopy analysis, using a Fourier transform infrared spectroscopy (FTIR) spectrophotometer (IRTracer-100, USA). The absorbance was recorded over a spectral range of 600–4,000 cm⁻¹, with 32 scans performed at a resolution of 0.09 cm⁻¹. Subsequently, an X-ray diffraction (XRD) analysis was performed using a SmartLab apparatus from Japan, which utilized nickel-filtered Cu-Kα radiation (λ = 1.5406 Å). The scanning range occurred from 5° to 90° at a rate of 10° per min. Finally, the cross-sections of CGV and CGVC films were observed with a scanning electron microscope (SEM) (FEI Magellan 400, USA). The surface of films was gold-coated with a sputtering tool at a current of 10 mA for 30 s.

2.4 Performance test

Detailed information about the performance testing in this study is given in the Supporting Information (S1).

3.1 Fabrication and characterization of CGVC bio-polymer film

Carnauba wax is insoluble in water, but dissolved in a hot ethanol solution. Hence, a homogenous system can be achieved through co-mixing at 60°C. However, in the cooling stage of film formation, carnauba wax is easy to migrate from the matrix phase to the surface layer and undergo secondary agglomeration, which has positive effects on the surface hydrophobic effect of film materials. However, the uniformity of the formed film was poor when the content of carnauba wax in the system exceeded 20 wt%. In addition, vanillin, as a crosslinking agent, can facilitate the formation of a three-dimensional network structure in films, stabilize carnauba wax, and significantly enhance the properties of CGVC bio-polymer films. The schematic diagram of film preparation is shown in Figure 1.

Figure 1

Schematic diagram of the film preparation.

According to the infrared spectrum of the CGV film (Figure 2a), the obvious absorption peak observed at 3,280 cm⁻¹ is attributed to the stretching vibration of −OH, and the peak at 1,640 cm⁻¹ is attributed to the stretching vibration of C═O (22), and the absorption peak at 1,030 cm⁻¹ is related to the interaction between the −OH component in glycerol and the functional groups present in chitosan and gelatin (23). From the infrared spectrum of the wax, it can be inferred that the peaks formed at 2,920 and 2,850 cm⁻¹ are attributed to the methylene asymmetry and symmetric stretching vibration of the fatty acid chain, respectively. Compared with the CGVC film, only two new peaks were added, located at 2,920 and 2,850 cm⁻¹, respectively, while the remaining peaks aligned with the absorption peak of CGV. This indicates that the addition of carnauba wax did not affect the hydrogen bonding interaction between gelatin and chitosan, as well as the formation of Schiff bonds between vanillin and gelatin/chitosan (24).

Figure 2

(a) FTIR and (b) XRD spectra.

It can be seen from the XRD spectra of CGV and CGVC films (Figure 2b) that CGV showed a broad diffraction peak at 20.3°, which is related to the lower crystallinity of CG. The CGVC film, on the other hand, exhibits new sharp and strong peaks at 21.2° and 23.6°, respectively, based on retaining the broad diffraction peak at 20.3°, which is related to carnauba wax (25). This indicates that the crystal structure of the film was not destroyed after adding carnauba wax, which was further confirmed by SEM analysis results.

From the image of the surface of CGVC film, a large number of uniformly dispersed spherical structures were observed, indicating that carnauba wax is evenly distributed on the surface of CG matrix (Figure 3a). The cross-sectional images showed that the carnauba wax left a pore structure in the diameter range of 10–80 μm (Figure 3b). These results explain from a microscopic perspective that the particle size and distribution of wax affect hydrophobicity and mechanical properties of the film.

Figure 3

(a) SEM images of the surface of CGVC film and (b) the cross-section of CGVC film.

3.2 Mechanical and thermal performance

Although CG has good film formation, its toughness is poor. In order to enhance its toughness, plasticizer is often added to the system or other bio-polymer materials for the formation of the composite film. In this study, TS and breaking elongation (EB) of the film were tested according to international organization for standardization 527-2 (26). TS values reduced from 29 to 10 MPa as the amount of carnauba wax increased, while EB continued to increase significantly from 16% to 38% (Figure 4a and b). This strength and toughness can meet the requirements for materials used as packaging films. The reason for this phenomenon is mainly attributed to wax plasticity (27). Carnauba wax mainly contains more than 80% aliphatic esters and cassia bark acid diesters, which improves the flexibility of the film. It is also possible that the presence of carnauba wax weakens the intermolecular forces, especially hydrogen bonds, thereby reducing the TS.

Figure 4

(a) TS and EB of CGVC films, (b) stress–strain plot of CGVC films, and (c) TGA and DTG thermograms of the CGVC film.

Thermogravimetric analysis (TGA) of CGVC and thermogravimetric differential curve (DTG) show the multistep weight loss (Figure 4c). The initial stage of weight change in the film takes place between 80°C and 120°C, primarily resulting from the evaporation of unbound water. The loss in the second stage is in the range from 180°C to 220°C. At this stage, the weight loss of the film is due to the decomposition of plasticizer (glycerol) (28,29). The reduction of weight in the third stage occurs between 270°C and 400°C, which corresponds to the thermal degradation of chitosan, gelatin, and wax (30). The thermal decomposition temperature of chitosan and gelatin is 270–320°C and 280–370°C, respectively. The improvement in thermal stability includes two reasons: the improvement of crosslinking on thermal stability and the influence of carnauba wax. Carnauba wax can promote the crosslinking reaction between chitosan and gelatin, forming a more stable three-dimensional network structure, thus improving the thermal stability of the film.

3.3 UV–visible light transmittance

Packaging materials outdoors need to have significant UV rays protection capabilities to avoid excessive UV damage to fruits or food. As shown in the results (Figure 5), the transmittance of CG film is low in the range of 200–300 nm, but high in the range of 300–400 nm. For CGV films, the introduction of vanillin significantly reduces the UV transmittance of the film (31). Furthermore, the introduction of carnauba wax did not weaken the UV-blocking performance of the film, and all CGVC films have zero UV transmittance in the range of 200–400 nm. It is worth noting that CGVC films completely prevent short wavelength blue light at 410 nm from harming human health (32). Meanwhile, carnauba wax is dispersed inside the matrix as a ball, greatly reducing the transmittance to visible light. This value of CG film reduced at 600 nm from about 82–28% with the introduction of 5 wt% carnauba wax.

Figure 5

UV and visible light transmittance.

3.4 Antioxidant activity

One of the important features of high-quality packaging film is their ability to antioxidant activity. Owing to the fact that a large portion of the chemical composition of carnauba wax is saturated compounds, they are less susceptible to traditional self-oxidation reactions that primarily attacks double bonds, and therefore, they often maintain their oxidative stability. In the experiment, the absorbance of 2,2-diphenyl-1-picrylhydrazyl (DPPH) stock solution, as well as solutions soaked in CGV and CGVC films, was tested separately using the method described in the previous study (33), and the results are shown in Figure 6a. As the carnauba wax content increases, the absorbance at 517 nm shows a decreasing trend. Subsequently, Figure 6b shows the results of further evaluating the antioxidant capacity of CGVC in terms of changes in absorbance at 517 nm. Due to unsaturated bonds in the CGV film being attacked by free radicals to form more stable radicals, its ability to scavenge DPPH free radicals is relatively low (34). With the increase of a large amount of saturated components in carnauba wax, the antioxidant capacity against DPPH is enhanced. The maximum scavenging activity can reach 69.07%. The carnauba wax contains natural antioxidant components such as gallic acid, catechin, and chlorogenic acid, which is the main reason for improving antioxidant performance of the film (35).

Figure 6

(a) Absorbance of the solutions soaked in CGV and CGVC film and (b) DPPH scavenging capacity of CGVC films.

3.5 Water resistance

Low expansion rate, water solubility (WS), and excellent waterproofing have always been the goals pursued in the research and development of biopolymer films. The determination of WS reflects the ability of the film to maintain its integrity when in contact with wet food, which is related to the durability of the film and the safety of food. However, the vast majority of bio-polymers are highly soluble in water (36), which restricts their waterproof properties and is an important factor limiting the application of bio-polymer films. The prepared CGVC film did not dissolve visibly in water after 24 h, which may be responsible for the formation of stable crosslinking network. Moreover, the swelling ratio (SR) of the film reduced significantly with carnauba wax increasing from 5 to 20 wt% (Figure 7a). As the carnauba wax content increases, the WS of the film has a slightly decreasing trend, which may be due to the hydrophobicity of wax molecules (37).

Figure 7

(a) SR and WS (b) WCA of CGVC films.

Wet-proof and waterproof resistance are the most important properties of packaging film. The waterproof properties of the CG film are inadequate because of the presence of numerous hydrophilic groups (38). As shown in Figure 7b, the water contact angle (WCA) of CGVC films was improved to different degrees compared with that of CGV films, indicating that the carnauba wax improved the hydrophobic capacity of the films. With the increasing contents of carnauba wax, the WCA exhibited a trend of rising at first and then decreasing. WCA was increased to 124° when 10 wt% carnauba wax was added to the system. The improvement in hydrophobicity is mainly related to the hydrophobicity of wax components, while the three-dimensional network formed by crosslinking reduces the interaction between bio-polymers and water molecules. And further increasing the amount of carnauba wax, the WCA began to decrease. The reason for this phenomenon may be that the excess carnauba wax is not evenly dispersed in the matrix, leading to the decrease in hydrophobicity caused by agglomeration (2).

Obtaining packaging films with excellent performance is currently the focus of research and development for bio-polymer films. Table 1 lists the relevant research works on CG films, some of which have obtained excellent results in terms of antioxidant and anti-UV properties, but have poor waterproof performance. Some studies mainly focus on waterproofing, but there is room for improvement in terms of UV resistance performance. Here, a novel and simple preparation method has been developed, which can obtain carnauba wax/chitosan/gelatin films with good hydrophobicity, excellent UV barrier, and antioxidant properties. However, after adding carnauba wax to films, although the hydrophobicity was improved, it did not achieve the superhydrophobic effect. This may be related to the distribution and morphology of carnauba wax. According to SEM images, on the surface of the film, wax particles did not form a uniform protrusion structure similar to lotus leaves. Moreover, the presence of wax particles inside the film may also affect the hydrophobic effect of the film to some extent due to compatibility issues.