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Smart Hydrogel Film of Bionanocellulose/Alginate/Polyethylene Glycol/Thermochromic Dye (BNC/ALG/PEG/TD) with Thermo-Responsive Behavior

Written By

Nurfarahin Zainal, Ku Marsilla Ku Ishak and Yazmin Bustami

Submitted: 09 October 2024 Reviewed: 24 October 2024 Published: 07 January 2025

DOI: 10.5772/intechopen.1008224

Application of Alginate - Based Smart Materials IntechOpen
Application of Alginate - Based Smart Materials Edited by Dr. Md. Abul Kalam Azad

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Application of Alginate - Based Smart Materials [Working Title]

Assistant Prof. Dr. Md. Abul Kalam Azad, Dr. Mireia Mallandrich Miret and Ph.D. Ariana Hudita

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Abstract

We aim to explore bionanocellulose/alginate/polyethylene glycol/thermochromic dye (BNC/ALG/PEG/TD) as a thermo-responsive hydrogel film. In this study, BNC was produced using Pichia kudriavzevii USM-YBP2. Then, the morphology of BNC was observed using transmission electron microscope (TEM). The formation of hydrogel film utilized the ionic crosslinking technique by mixing BNC, ALG, PEG, and thermochromic dye (TD) with Ca2+ ions. Then, the hydrogel film was evaluated for its swelling property, thermo-responsive behavior, and biodegradability. BNC exhibits aggregated fiber and clusters of spherical shape. BNC/ALG/PEG/TD formed a gray film, which indicates the successful integration of TD. Interestingly, it shows acceptable stability in wet and dry conditions, suggesting its robustness and stability. However, BNC/ALG/PEG/TD demonstrated low water content (16.6%) as compared to the control probably because of the increment in crosslink density or improvement of polymer-polymer interaction. The hydrogel film changes drastically to colorless at temperatures >30°C proving its thermo-responsive behavior. Interestingly, this hydrogel film also presented a reversible thermochromism property that enables the hydrogel to reverse back to its original color and morphology. However, it shows slow degradation and probably needs more than 2 weeks to fully degrade. In conclusion, BNC/ALG/PEG/TD can be potentially explored as a smart hydrogel film in a wide area of applications.

Keywords

  • smart hydrogel film
  • bionanocellulose
  • alginate
  • polyethylene glycol
  • thermo-responsive

1. Introduction

Hydrogels are fascinating materials made up of chains of hydrophilic polymers with a three-dimensional (3D) network [1, 2]. Their biocompatibility and environmental friendliness allow for extensive application and development of new hydrogel technology [2]. Smart hydrogels come into view as a result of ongoing scientific advancements and discoveries [3]. It is in demand due to its wide range of capabilities since it shows response to either physical, chemical, or biological stimuli. Research on hydrogel has changed from relatively straightforward, water-swollen macromolecular networks to hydrogels that may react to environmental stimuli like pH and temperature [4]. Scientists have shown increasing interest in creating temperature-responsive hydrogels that are both biodegradable and biocompatible, largely due to their impressive properties [5]. Thus, hydrogels that show response to temperature were widely studied and actively developed for various applications. In a recent study by Ma et al. [6], they elaborated that stimulus-responsive TEMPO-oxidized cellulose nanofibers/poly(N-isopropylacrylamide) (TOCN/PNIPAM) can be assembled into a flexible sensor and generate sensing signals when driven by temperature changes to achieve real-time feedback. Another study, [7] reported that poly(2-hydroxypropyl acrylate-co-acrylamide) (P(HPA-co-AM))/hydroxypropyl cellulose (HPC)/lithium chloride (LiCl) exhibits excellent thermo-responsiveness and provide ideas for the next generation of smart multifunctional electronic skin. Also, several studies reported the feasibility of using thermochromic composite hydrogel film for the fabrication of energy-saving smart windows [8, 9, 10, 11]. A study by He et al. [8] proposed that a thermochromic composite hydrogel film based on W-doped vanadium dioxide (W-VO2), poly N-isopropyl acrylamide (PNIPAM), and polyacrylamide (PAM) provides an effective energy-saving and multifunctional smart window. Furthermore, the introduction of thermochromic hydrogel as smart window materials is beneficial due to its excellent thermal response, high radiation-blocking efficiency, cost-effectiveness, biocompatibility, and good uniformity [10].

In recent years, nanocellulose has been extensively used to improve the functionality of certain composites. In fact, it has been used widely to form multifunctional composites since it can be gelatinized to form a three-dimensional porous network structure due to a high concentration of hydroxyl groups [6]. There are 3 types of nanocellulose, namely, nanofibrillated cellulose (NFC), cellulose nanocrystals (CNC), and bacterial nanocellulose/bionanocellulose (BNC), depending on their yield methods, processing conditions, and properties [12, 13]. BNC has made an entry into commercial and medical applications upon its extensive utilization. Gluconacetobacter xylinus, which was previously known as Acetobacter xylinum, is the first bacterial species producing BNC [14]. Since then, the study of different groups of microorganisms for the biosynthesis of BNC has been widely conducted [15]. BNC has been produced by cultivating the respective microbial for a few days in an aqueous culture media containing suitable nutritional parameters which offers an optimum condition [16]. High elasticity and tensile strength are produced by the fibrillated network of the BNC, which results in very good mechanical characteristics [17]. Due to their distinctive physicochemical characteristics, such as hydrophilicity, chirality, biocompatibility, and biodegradability, these nanoparticles are widely regarded as the “green molecule” of the future [18].

In this study, we aim to produce hydrogel film using bionanocellulose (BNC), alginate (ALG), polyethylene glycol (PEG), and thermochromic dye (TD) that can behave as a thermo-responsive hydrogel. The biosynthesis of BNC was conducted using a static fermentation of indigenous yeast, Pichia kudriavzevii USM-YBP2, and further evaluated using TEM analysis. The physical morphology, swelling activity, thermo-responsive behavior, and biodegradable properties of BNC/ALG/PEG/TD hydrogel film are reported.

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2. Materials and methods

2.1 Materials

Glycerol 99.5% (MW 92.10 g/mol) and ethanol 95% (MW 46.07 g/mol) were purchased from R&M Chemicals (UK). Yeast extract (MW 319.193 g/mol), sodium alginate (MW 120 k–190 k g/mol), polyethylene glycol (6000), disodium phosphate (MW 141.96 g/mol), and calcium chloride dihydrate (MW 147.02 g/mol) were purchased from Sigma Aldrich (USA). The nutrient broth was purchased from Oxoid (UK), peptone (RM006) was purchased from HiMedia Laboratories (India), citric acid monohydrate (MW 210.14 g/mol) was purchased from BDH Laboratory (UK), molasses (MW 3.5 × 105–4.3 × 105 g/mol) was purchased from EMRO, Malaysia, and thermochromic black pigment dye powder (TD) was purchased from Hali Industry (China). Pichia kudriavzevii USM-YBP2 glycerol stock was obtained from Lab 207, School of Biological Sciences, Universiti Sains Malaysia, Penang, Malaysia.

2.2 Methods

2.2.1 Production of BNC using Pichia kudriavzevii USM-YBP2

The production of BNC using P. kudriavzevii USM-YBP2 was adapted from Jasme et al. [19]. Initially, the yeast glycerol stock, P. kudriavzevii USM-YBP2, was thawed and activated by adding 1 mL of the glycerol stock into 50 mL of nutrient broth and was agitated at 200 rpm at 30°C for 24 hours. Then, 10 mL of activated yeast was inoculated into 1 L of Hestrin-Schramm broth (HSB) containing 0.5% (w/v) peptone, 0.5% (w/v) yeast extract, 0.27% (w/v) disodium phosphate, 0.115% (w/v) citric acid, and 0.5% (w/v) of 95% (v/v) ethanol. Following the inoculation, the flask was left on a bench for 7 days at room temperature under static fermentation conditions. The formation of pellicles on the air-liquid interphase was monitored. The morphology of P. kudriavzevii USM-YBP2 and pellicles was observed under a light microscope. BNC was centrifuged at 7871 x g for 10 minutes. The supernatant was discarded, and the remaining cell pellet was washed. This procedure was repeated three times to remove any excess medium. Then, the cell pellet was immersed and vortexed in 30 mL of 1 M NaOH solution for 15 minutes at 60°C to immobilize the yeast cells before it was subjected to centrifugation and washing. The obtained cell pellet was sonicated for 15 minutes and was incubated in a dialysis membrane for at least 3–5 days. Finally, the mixture was centrifuged and the supernatant was discarded. The cell pellet was stored at −40°C for at least 24 hours before the freeze-drying process. The obtained BNC powder was kept at room temperature (RT) until use.

2.2.2 Characterization of BNC using TEM analysis

BNC was analyzed using TEM at the Electron Microscopy Unit, School of Biological Sciences, Universiti Sains Malaysia, Penang, Malaysia. Firstly, a small amount of BNC powder was suspended in distilled water and was sonicated for 10 minutes. Then, the diluted aqueous suspension of the treated BNC was dried for a few minutes on the 400-mesh copper-coated grid. The morphology of BNC was captured using an FEI TECNAI G2 20 S-twin transmission electron microscope (TEM) operating at 200 kV.

2.3 Formulation of BNC/ALG/PEG/TD hydrogel film

To make thermo-responsive hydrogel film (BNC/ALG/PEG/TD), 0.17% (w/v) thermochromic dye (TD) solution was mixed with 4% (w/v) BNC, 8% (w/v) ALG, and 4% (w/v) PEG. Then, the mixture was added into a beaker containing 30 mL of distilled water and stirred until the mixture was completely homogenized. After that, 4 mL of the blending solution was slowly poured onto a Petri dish containing 4% CaCl2 solution, and the hydrogel film was allowed to crosslink for 5 minutes. Subsequently, the formed hydrogel film was washed and rinsed with dH2O to remove any excess of CaCl2. Finally, the formed BNC/ALG/PEG hydrogel film was air-dried for 12–18 hours at room temperature. The formed hydrogel film was stored at 4°C for further characterization and testing. For control, a hydrogel film without TD solution was prepared and denoted as BNC/ALG/PEG.

2.3.1 Swelling test

Firstly, the hydrogel film was air-dried at room temperature for 48 hours to obtain dried hydrogel. Then, it was immersed in 80 mL of distilled water. The swelling test was conducted for 5 days. The swollen hydrogel was weighed at 24, 48, 72, 96, and 120 hours. The filter paper was used to remove any excess water. The water content (Wr) of the hydrogel films and the equilibrium swelling ratio (eqsw) were calculated using Eqs. (1) and (2) [20].

Water contentWr=WS−WdWs×100%E1
eqsw=WSWdE2

where Ws denoted the swelling weight and Wd denoted the dry weight of the hydrogel film. The swelling studies were conducted in duplicates.

2.3.2 Testing of thermo-responsive behavior at different temperatures

BNC/ALG/PEG/TD hydrogel films were placed into glass Petri dishes and incubated overnight at 4°C, room temperature (RT), 30°C and 60°C. The thermo-responsive behavior of the respective hydrogel films was observed and recorded. In order to determine its reversible thermochromic property, the hydrogel film was initially heated until 100°C before cooling down by immersing the hydrogel film in dH2O for 5 days at RT. The color and physical morphology of hydrogel films after heating and cooling conditions were observed and recorded.

2.3.3 Biodegradability test

The biodegradability of hydrogel films was investigated by soil-burial test. The hydrogel film was introduced to a compost bed in a control environment; 30°C, pH 6.0–8.5, and moisture content of 60%. The biodegradation test was taken for 2 weeks. The changes in the hydrogel film were observed and recorded.

2.3.4 Characterization of hydrogel film using SEM analysis

The selected hydrogel films (before and after the degradation process) were analyzed using SEM at the Electron Microscopy Unit, School of Biological Sciences, Universiti Sains Malaysia, Penang, Malaysia. The samples were sputter coated with a thin layer of palladium or gold alloy to improve surface conductivity and tilted 30° for better observation. The surface views of the hydrogel films were taken at a magnification of 100x–10,000x.

2.4 Statistical analysis

A student’s T-test was calculated to compare means between two treatment groups. Statistical significance was defined as a p < 0.05. The data are presented as the mean ± standard deviation (SD) of multiple replicates.

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3. Result and discussion

3.1 Production of Pichia kudriavzevii USM-YBP2

Pichia kudriavzevii USM-YBP2 presented an oval or elliptical shape under a light microscope, as seen in Figure 1. Furthermore, the majority of the cells become clustered and in proximity with one another as indicative of budding behavior and is aligned with our previous study in Low et al. [18]. The formation of pellicles was observed at the liquid-air interface of the Hestrin-Schramm broth (HSB) and appeared as an interwoven network of fiber. As reported by Lahiri et al. [21], a visible, three-dimensional (3D), and interconnected reticular pellicle is seen at the medium interface during static fermentation of BNC. This physical trait is a strong indication that the obligate aerobic isolates are generating cellulose [22]. A study by Ul-Islam et al. [23] reported that the entrapped carbon dioxide (CO2) produced by the bacterial metabolism causes the BNC biofilm to float on the top of the medium. It was also speculated that the obligate aerobes, microorganisms embedded within the cellulose matrix, use oxygen for cell growth while utilizing sugars to build cellulosic biofilms or pellicles [24]. According to Campano et al. [25], bacteria gather at the surface and produce BNC in layers as the amount of oxygen drops. This is because microorganisms producing cellulose predominantly consume dissolved oxygen during microbial cellulose production.

Figure 1.

Morphology observation of P. kudriavzevii USM-YBP2 under a light microscope with 1000X magnification (left image) and pellicle production on the surface of HSB after 7 days of incubation (right image).

Transmission electron microscopy (TEM) was used to observe the morphology of BNC produced in static fermentation (Figure 2). It can be noticed that BNC shows a densely aggregated fiber network and resembles long needles. Likewise, El-Naggar et al. [26] stated that BNC produced during static culture developed needle-like particles with sizes ranging from 30 to 40 nm. According to Cai and Kim [27], BNC differs from plant cellulose in structure due to a different synthesis procedure in which bacteria synthesize fine sub-elementary cellulose fibrils that are extruded from terminal enzyme complexes into the culture medium during cultivation. Moreover, other types of morphology were also observed. A cluster of spherical shapes BNC was observed in the TEM image and is in agreement with our previous study [18]. Similarly, a study conducted by Zhong et al., Chawla et al., and Trilokesh and Uppuluri [28, 29, 30] also reported the formation of spherical CNC (SCNC), and it is a common morphology for BNC produced from fruit sources.

Figure 2.

TEM image of BNC produced using Pichia kudriavzevii USM-YBP2. A thin and highly aggregated fiber network of BNC (blue arrow) and clustered spherical shapes of BNC (red arrow) were observed under 40,000X magnification.

3.2 Formulation and characterization of thermo-responsive hydrogel film

In Figure 3, a distinct hydrogel film color can be observed for the control and thermo-responsive hydrogel. The control hydrogel (BNC/ALG/PEG) formed as a translucent sheet of film with a hint of brown color that is due to the addition of BNC. While BNC/ALG/PEG/TD hydrogel formed as a gray film, which indicates the successful integration of TD within the BNC/ALG/PEG hydrogel network. Furthermore, TD was blending well in the BNC/ALG/PEG hydrogel as the gray color was evenly distributed. Interestingly, both hydrogel films show acceptable stability in both wet and dry conditions (Figure 3A and B). The formed hydrogel films were not only capable of absorbing a large amount of water, but they also could withstand high temperatures. This property is important since thermo-responsive hydrogel uses temperature as the external stimulus. Thus, it is necessary to ensure that the hydrogel shows good mechanical strength and is stable at different temperatures, from low to high temperatures. There is no doubt that the integration of BNC helps to improve the mechanical properties and form a robust hydrogel film. A study by Wang et al. [31] reported that nanocellulose-based hydrogels show mechanical stability and create intertwined molecular chains to stabilize rich composite spaces. In a common condition, alginate-hydrogel is not stable and easily disintegrates with the presence of monovalent cations such as Na+ [32]. Therefore, the addition of BNC is instrumental and helps to improve the mechanical properties of alginate-hydrogel.

Figure 3.

The wet and dry conditions of (A) BNC/PEG/ALG hydrogel film and (B) BNC/PEG/ALG/TD hydrogel film.

Based on Figure 4, BNC/ALG/PEG/TD reached swelling equilibrium faster (48 h) than the control, BNC/ALG/PEG (72 h). However, it exhibits significantly low water content (16.6%) as compared to the control (22.5%) (p < 0.05). Accordingly, we suggested that the inclusion of TD might affect the hydrogel network as depicted by low water content and equilibrium swelling ratio (1.2). When the crosslink density of the hydrogel increases, swelling decreases, and vice versa [33]. Besides, it is also speculated that the addition of TD might improve the polymer-polymer interaction within the hydrogel network. According to Ghosh and Katiyar [20], the combination of biopolymers has boosted the polymer-polymer contact and further created a hydrogel material with customized features, with polymer-polymer interaction dominating the water-polymer interaction. They also recommended that the polymer-polymer interactions between molecules be improved, which the polymers linked themselves by covalent bonding, hence resulting in disregarding water interactions between the molecules. This property might be causing lower water absorption due to a stronger link between the polymer.

Figure 4.

Comparison of swelling activity between BNC/ALG/PEG/TD and control, BNC/ALG/PEG hydrogel films depict different water content, Wr, and swelling equilibrium ratio, eqsw.

3.3 Thermo-responsive behavior

Figure 5 presents the thermo-responsive behavior of the BNC/ALG/PEG/TD hydrogel film after being treated at different temperatures. It can be seen that at 4°C, the color of the hydrogel film was gray and slightly changed to a lighter gray color at room temperature (RT). Notably, as the temperature increased to 30°C and 60°C, the hydrogel film was colorless and appeared as translucent hydrogel films, similar to the control, BNC/ALG/PEG, in Figure 3A. In the present study, BNC/ALG/PEG/TD hydrogel film remains its original gray color at temperatures <30°C, while changing drastically to colorless at temperatures >30°C. This result clearly shows the thermochromism characteristic of the BNC/ALG/PEG/TD hydrogel film in response to the temperature change. A study conducted by Yan et al. [34] elaborated that thermochromic hydrogel film based on photonic nanochains (TRPCF-PNC) exhibited a distinctive and rapid optical change behavior in response to temperatures ranging from 26 to 44°C.

Figure 5.

BNC/ALG/PEG/TD hydrogel film displayed thermo-responsive behavior. At temperatures <30°C; 4°C and RT, hydrogel films appeared as dark and light gray colors, respectively (A). At temperatures >30°C; 30°C and 60°C, hydrogel films appeared as translucent hydrogel films similar to the control, BNC/ALG/PEG.

While the BNC/ALG/PEG/TD hydrogel film successfully displayed thermo-responsive behavior, it is important to determine its reversible thermochromic properties. As expected, the hydrogel film was transformed from its original gray color to colorless when heated to 100°C (Figure 6). It also can be seen that the film was dehydrated and showed slight deformation, especially in its peripheral area. It is known that hydrogel is rich in water content; thus, exposing hydrogel to high temperatures can cause water evaporation, which contributes to hydrogel dehydration and deformation. Interestingly, the hydrogel film was recovered to its original form when cooling at RT for five consecutive days. The hydrogel film transformed back to a gray color and was rehydrated. Nevertheless, the BNC/ALG/PEG/TD hydrogel film proposed in this study is a promising reversible thermochromic hydrogel, evidenced by its capability to regulate based on the external stimulus, and is found attractive for the fabrication of medical devices, sensors, actuators, soft robotics, and smart windows [6, 11]. The hydrogel actuator is related to its actuated behavior that can absorb and release water. Thus, a thermosensitive hydrogel actuator can be prepared by manipulating the water movement that can be achieved by changing the temperature [6]. Ideally, the solar light getting into the building can be manipulated by the utilization of thermochromic smart windows since it can respond to the atmospheric temperature automatically without additional energy consumption [11]. In the most recent study by Zhong et al. [35], they reported that hydroxypropyl cellulose (HPC) hydrogel smart windows show a significant energy-saving potential compared to ordinary clear glass windows.

Figure 6.

The reversible thermochromic property of BNC/ALG/PEG/TD hydrogel film after heating at 100°C and cooling at RT for five consecutive days.

The biodegradability of BNC/ALG/PEG/TD hydrogel film was investigated under composting environments [moisture content: 60%, 30°C, pH 6.0–8.5]. As presented in Figure 7, the physical morphology of the hydrogel film shows minimal damage, with most of the film remaining intact even after 2 weeks of composting. However, when observed under SEM, the porous hydrogel film was not visible, and it might be due to the dissolution of the network chain, and the hydrogel film started to lose its structure. A study by Wang et al. [36] elaborated that hybrid injectable hydrogels consisting of alginate, gelatin, and nanocrystalline cellulose (NCC) have superior mechanical properties resulting from interpenetrated networks with reinforced nanomaterials and good stability against degradation. In fact, alginate-based hydrogel exhibits a remarkably slow degradation rate, as discussed in many studies [36, 37, 38]. In the area of bioprinting and tissue engineering applications, alginate shows limitations since mammalian cells do not produce enzymes capable of degrading the polymer, leading to a slow degradation process that is primarily controlled by the outward flux of ions into the surrounding environment [38]. Therefore, it is sufficient to mention that the hydrogel film in this study shows slow degradation under the composting environment, as it probably needs more than 2 weeks to fully degrade and might be useful in certain areas of applications.

Figure 7.

Observation of the physical morphology and the close-up SEM image of BNC/ALG/PEG/TD hydrogel film (A) before and (B) after 2 weeks of composting.

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4. Conclusions

In conclusion, BNC was successfully produced from Pichia kudriavzevii USM-YBP2 by the presence of pellicles at the liquid-air interface of the medium. The morphologies of formed BNC were observed as aggregated fiber and clusters of spherical shape. The addition of BNC is instrumental in improving the mechanical properties of hydrogel film since it shows acceptable stability in wet and dry conditions. The successful integration of thermochromic dye (TD) into the BNC/ALG/PEG network can be easily seen as it formed a gray color film as compared to the control, BNC/ALG/PEG, which formed a translucent film. BNC/ALG/PEG/TD hydrogel film exhibits significantly low water content (16.6%) as compared to the control (22.5%), which is probably because of the increment in crosslink density or improvement of polymer-polymer interaction. Most importantly, BNC/ALG/PEG/TD presented as an excellent thermo-responsive hydrogel as it shows response at different temperatures and displayed a drastic change from gray to colorless when the temperature was >30°C. It also exhibits reversible thermochromic properties as it can be recovered to its original color and morphology when heated to 100°C and cooled at RT. Furthermore, this thermo-responsive hydrogel film shows slow degradation under the composting environment and probably needs more than 2 weeks to fully degrade. Overall, BNC/ALG/PEG/TD is a robust thermo-responsive hydrogel that can be explored in different applications, such as for the fabrication of medical devices, sensors, actuators, soft robotics, and smart windows.

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Written By

Nurfarahin Zainal, Ku Marsilla Ku Ishak and Yazmin Bustami

Submitted: 09 October 2024 Reviewed: 24 October 2024 Published: 07 January 2025

© The Author(s). Licensee IntechOpen. This content is distributed under the terms of the Creative Commons 4.0 International License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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