pH-responsive κ-carrageenan/HPMC biodegradable films incorporated with roselle anthocyanin for freshness indication

Introduction

Food packaging is transforming from a simple barrier into an active and smart system that safeguards food while offering real-time data on product safety and quality. Smart packaging solutions that visually show freshness or decay can decrease food waste, enhance consumer confidence, and aid supply-chain choices (Boonsiriwit et al., 2021; Mathew, 2024).

Films made from biopolymers sourced from renewable polysaccharides and cellulose derivatives have garnered significant attention as eco-friendly substitutes for petroleum-based plastics due to their biodegradability, ability to form films, and safety for food contact (Mathew, 2024). Kappa-carrageenan, a sulfated galactan derived from red seaweeds, is a polysaccharide that provides excellent gelation, transparency, and oxygen barrier properties suitable for edible and biodegradable films. However, films made solely from pure κ-carrageenan tend to be brittle and vulnerable to moisture, necessitating formulation approaches like polymer blending to enhance their mechanical and barrier performance (Mathew, 2024).

Hydroxypropyl methylcellulose (HPMC) is a semi-synthetic cellulose ether commonly utilized in food and pharmaceutical coatings because of its superior film-forming properties, flexibility, and mechanical strength. Combining HPMC with polysaccharides like κ-carrageenan has demonstrated a synergistic enhancement of tensile strength, a decrease in brittleness, and the production of films that are better suited for packaging applications requiring mechanical integrity and handling (Boonsiriwit et al., 2021; Mu et al., 2025).

Anthocyanins stand out as one of the most promising natural pigments for visual freshness indicators due to their distinct, reversible color changes throughout the pH range commonly linked to food spoilage (acidic → neutral → basic), and they can be easily sourced from edible plants (Grajeda-Iglesias et al., 2016). Films made from anthocyanins have been utilized to signal volatile basic nitrogen (VBN) and additional spoilage indicators for meat, fish, and seafood by creating noticeable color changes that are easily understandable for both consumers and packers.

Roselle (Hibiscus sabdariffa) calyces are a rich, low-cost source of anthocyanins (mainly delphinidin and cyanidin derivatives) and possess antioxidant and antimicrobial traits that further support their use in active packaging formulations (Grajeda-Iglesias et al., 2016; Millinia et al., 2024). Recent studies have shown that anthocyanins from roselle can be effectively extracted, stabilized (through micro- and co-encapsulation), and integrated into polymer matrices to create halochromic films that are sensitively responsive to pH and volatile amines associated with spoilage (Huang et al., 2023; Millinia et al., 2024; Latiff et al., 2025).

While HPMC/κ-carrageenan films containing plant-derived anthocyanins have been reported, limited studies have explored the incorporation of roselle (Hibiscus sabdariffa) anthocyanins within this polymeric system. Roselle anthocyanins possess a distinct compositional profile, predominantly delphinidin- and cyanidin-based derivatives, with relatively higher intrinsic acidity and antioxidant capacity compared to commonly studied berry sources. These characteristics may influence polymer interactions, pH responsiveness, and overall film performance.Although the κ-carrageenan/HPMC blend provides a stable film-forming matrix due to complementary intermolecular interactions, anthocyanins function as active additives that can modify the polymer network through non-covalent interactions, potentially altering mechanical behavior. While previous studies (e.g., Mu et al., 2025) demonstrated the feasibility of berry-based anthocyanin indicator films, differences in anthocyanin source and structure may significantly affect compatibility and functionality, therebyhighlighting the need to investigate roselle-derived anthocyanins in this blended system.

Despite advances in anthocyanin-based intelligent films, challenges remain in achieving uniform anthocyanin dispersion, maintaining structural stability under humid conditions, and ensuring sensitive response to subtle pH variations associated with early-stage food spoilage. Therefore, the present study aims to develop a biodegradable and pH-responsive film by incorporating roselle-derived anthocyanins into a blended HPMC/κ-carrageenan matrix. The formulation was optimized to achieve a balanced combination of colorimetric sensitivity, acceptable mechanical integrity, and functional stability. By integrating natural pH indicators with edible biopolymers, this work seeks to contribute to the development of sustainable and application-oriented smart packaging systems suitable for real food monitoring.

Materials and methods

Food-grade κ-carrageenan was obtained from Istore Direct Trading Pvt. Ltd. (Mumbai, India). Hydroxypropyl methylcellulose (HPMC) was purchased from Finar Chemicals Pvt. Ltd. (Gujarat, India). Shade-dried wild roselle (Hibiscus sabdariffa) powder was procured from Monks Bouffe LLP (Mumbai, India).The proximate composition of H. sabdariffa powder (per 100 g) was as follows: carbohydrates (62.13 g), protein (16.15 g), fat (1.97 g), and energy (315 kcal).

Extraction of roselle anthocyanins

Roselle anthocyanins (RA) were extracted according to the method reported by Giusti and Wrolstad (2001) with minor modifications. Briefly, 133.3 g of shade-dried H. sabdariffa powder was mixed with 2000 mL of 80% (v/v) ethanol. The pH of the extraction medium was adjusted to 2.0 using 1 M HCl to enhance anthocyanin stability and extraction efficiency. The mixture was heated at 50 °C in a thermostatically controlled water bath for 1 h and subsequently filtered through Whatman No. 1 filter paper.The filtrate was concentrated under reduced pressure using a rotary evaporator at 50 °C under dark conditions to prevent photo-degradation. The concentrated extract was further dried in a hot-air oven at 40 °C to obtain a semi-solid anthocyanin concentrate. The roselle anthocyanin extract was stored in amber-colored bottles at refrigerated conditions until further use.The extraction process was performed to obtain sufficient pigment for film incorporation; however, the total anthocyanin yield was not quantified in this study.

Preparation of film-forming solutions

Bioactive freshness-indicating films were prepared using κ-carrageenan and hydroxypropyl methylcellulose (HPMC) as film-forming polymers and sorbitol as a plasticizer, which was kept constant in all formulations. Roselle anthocyanin (RA) extract was incorporated as a natural pH-sensitive indicator at concentrations of 0%, 0.5%, 1.0%, 1.5%, and 2.0% (w/w) based on the dry polymer weight. Film-forming solutions were prepared by dissolving 8 g of κ-carrageenan and 3.2 g of HPMC (4:1, w/w) in 400 mL of distilled water at 80 °C for 1 h. After cooling to 50 °C, RA was added, and the mixtures were homogenized (5000 × g, 5 min) and degassed at 40 °C for 30 min. Aliquots (30 mL) were cast onto Petri dishes and dried at 30 °C for 24 h. The dried films were peeled and conditioned before characterization. The heating and drying conditions were carefully controlled to minimize possible thermal degradation of anthocyanins.The films were designated as (Figure 1) κ-C:Hm RA₀ (control film without anthocyanin), κ-C:Hm RA_0.5, κ-C:Hm RA₁, κ-C:Hm RA_1.5, and κ-C:Hm RA₂, corresponding to films containing 0, 0.5, 1.0, 1.5, and 2.0% (w/w) roselle anthocyanin, respectively.

Characterization of prepared films

Mechanical properties

Film thickness was measured using a digital thickness gauge (Asian Packaging 1686, India; accuracy ±0.01 mm). Measurements were taken at ten random locations per film, and mean values were reported.

Tensile strength (TS) and elongation at break (EAB) were determined according to ASTM D882-12 using a texture analyzer (Stable Micro Systems, Surrey, UK). Film strips (2 × 8 cm) were tested at an initial grip separation of 50 mm and a crosshead speed of 5 mm s⁻¹. TS was calculated as the ratio of maximum force to cross-sectional area, while EAB was expressed as the percentage increase in length at break relative to the initial length.

The elongation at break (%) of films was determined according to the reference method (ASTM, D882-12, 2012) using a texture analyzer (Stable Micro Systems, Surrey, UK). It was calculated by dividing the extension at the moment of specimen rupture by the initial length of the specimen and multiplying by 100.

Barrier properties

Film solubility in water (SW) was determined following Montoille et al. (2021). Dried film samples were immersed in distilled water for 6 h at room temperature with constant agitation, redried, and the solubility (%) was calculated based on weight loss.

SW(%)=Initial Dry Weight−Final Dry WeightInitial Dry Weight

Swelling behavior was evaluated as described by Norajit et al. (2010). Pre-weighed films were immersed in water at 25 °C for 2 min, surface-dried, and reweighed. The swelling ratio was calculated from weight gain.

Swelling Ratio=Final Weight−Initial WeightInitial Weight

Film samples (10 × 10 mm) were immersed in 1 N HCl and 1 N NaOH solutions at 25 °C. Visual changes were recorded after 24 h to assess stability under acidic and alkaline conditions.

Oil resistance was assessed following Hu et al. (2013) by sealing mineral oil–filled test tubes with films and observing oil penetration through the film onto filter paper.

Antioxidant analysis

Film samples (20 mg) were immersed in 100 µM DPPH methanolic solution and incubated in the dark for 1 h. Absorbance was measured at 517 nm, and antioxidant activity was expressed as percentage DPPH inhibition.

Structural properties

FT-IR spectra of films were recorded using an ATR-FTIR spectrometer (Bruker Alpha II) over a range of 4000–650 cm⁻¹ at a resolution of 2 cm⁻¹ to identify functional group interactions.

Thermal properties

Thermal stability was analyzed using a TGA Q50 analyzer (Shimadzu, Japan). Film samples (~6 mg) were heated from 20 to 600 °C at 10 °C min⁻¹ under a nitrogen atmosphere. Degradation behavior was evaluated from TGA and DTG curves.

Proximate analysis

Moisture content was determined by drying film samples at 105 °C for 24 h and calculating weight loss as a percentage of initial weight (Norajit et al., 2010).

Biodegradability

Biodegradation was assessed using the soil burial method (Martucci and Ruseckaite, 2009). Film samples were buried in moist soil and retrieved after 7, 14, and 21 days. Weight loss (%) was calculated to evaluate degradation.

Freshness indicator study

Fresh button mushrooms (Agaricus bisporus) were packaged with optimized biodegradable films (20 × 20 mm) and stored at 25 °C. Film color changes were visually monitored during storage to evaluate freshness indication.

Statistical analysis

All experiments were conducted in replicates (n = 5 or n = 10), and the results are presented as mean ± standard deviation (SD). Statistical differences among samples were evaluated using Duncan’s multiple range test, and differences were considered statistically significant at p < 0.05. Different superscript letters within the same column indicate significant differences.

Result and discussion

Mechanical properties

The average thickness of the κ-carrageenan:HPMC films incorporated with RA ranged from 0.047 to 0.076 mm (Table 1; Figure 2). Films containing 0.5% and 1.5% anthocyanin (κ-C:Hm RA_0.5 and κ-C:Hm RA_1.5) exhibited significantly higher thickness values (p < 0.05), whereas the film containing 1.0% anthocyanin (κ-C:Hm RA₁) showed the lowest thickness. Statistical analysis indicated that κ-C:Hm RA_0.5 and κ-C:Hm RA_1.5 were comparable to each other, while κ-C:Hm RA₀, κ-C:Hm RA₁, and κ-C:Hm RA₂ formed another statistically similar group. The observed variations in thickness may be attributed to differences in solid dispersion and intermolecular interactions between anthocyanin molecules and the polymer chains. At certain concentrations, anthocyanins may increase the effective solid content or promote polymer swelling, resulting in thicker films, whereas non-uniform dispersion at intermediate levels may lead to thinner structures. Similar thickness ranges and trends have been reported for polysaccharide-based films incorporated with plant-derived bioactive compounds (Terzioğlu et al., 2020; Prietto et al., 2017). These findings confirm that the thickness values obtained in the present work are consistent with those reported for biopolymer-based intelligent packaging films.

Table 1. Properties of κ-carrageenan films containing different HmRA concentrations.

Thickness measurements data were based on n = 10, whereas other properties data were based on n = 5. Values are mean ± SD with different letters indicating significance (p < 0.05).

The tensile strength of κ-carrageenan/HPMC films plasticized with sorbitol and incorporating increasing concentrations of roselle anthocyanin was evaluated according to ASTM D882.As shown in Table 1; Figure 2, tensile strength decreased significantly with increasing anthocyanin concentration (p < 0.05). The control film (κ-C:Hm RA₀) exhibited the highest tensile strength (43.5 ± 0.26 MPa), indicating strong intermolecular interactions and a compact polymer network. In contrast, the film containing 2.0% anthocyanin (κ-C:Hm RA₂) showed the lowest tensile strength (11.5 ± 0.10 MPa). Statistical analysis confirmed that all treatments differed significantly from one another, demonstrating a clear concentration-dependenteffect. The reduction in tensile strength may be attributed to structural modifications within the polymer matrix induced by anthocyanin incorporation. Anthocyanins possess multiple hydroxyl groups capable of forming hydrogen bonds with κ-carrageenan and HPMC chains, thereby participating in non-covalent intermolecular interactions. However, at elevated concentrations, excessive anthocyanin incorporation may disturb polymer chain packing density and reduce intermolecular cohesion, resulting in a less compact and mechanically weaker network. Such structural heterogeneity compromises stress distribution within the film, leading to decreased tensile resistance. Similar reductions in tensile strength have been widely reported in starch-, carrageenan-, and chitosan-based films containing natural pigments or extracts (Wang et al., 2021; Fonseca-García et al., 2021; Liu et al., 2022). Wang et al. (2021) observed a significant decrease in tensile strength of cornstarch-based films incorporated with plant extracts, attributing the effect to weakened polymer–polymer interactions. Likewise, Fonseca-García et al. (2021) reported reduced tensile strength in starch–chitosan composite films containing functional additives, which was associated with structural heterogeneity in the film network. Comparable trends were also reported by Liu et al. (2022) for anthocyanin-loaded intelligent films, supporting the mechanical behavior observed in the present study. Although increasing roselle anthocyanin concentration negatively affected tensile strength, films containing lower and moderate anthocyanin levels maintained acceptable mechanical integrity, making them suitable for intelligent food packaging applications where responsiveness and visual indication are prioritized.

Elongation at break represents the flexibility of films, defined as the percentage increase in length before failure. As shown in Table 1; Figure 2, the elongation at break of κ-C:Hm films decreased significantly with increasing roselle anthocyanin (RA) content (p < 0.05). The control film (κ-C:Hm RA₀) exhibited the highest flexibility (18.20 ± 0.03%), while the highest RA load (κ-C:Hm RA₂) showed the lowest elongation (6.06 ± 0.02%). Statistical analysis indicated significant differences among most treatments, with κ-C:Hm RA_1.5 and RA₂ being statistically similar. The observed reduction in elongation at break with increasing anthocyanin concentration may be due to restricted polymer chain mobility and increased rigidity induced by anthocyanin–polymer interactions, which limit plastic deformation. Similar decreases in flexibility with higher concentrations of anthocyanins or phenolic extracts have been reported in other biopolymer systems. For instance, colorimetric films based on polyvinyl alcohol incorporated with anthocyanins from purple tomato demonstrated a significant decrease in elongation at break as the anthocyanin content increased, attributed to tighter intermolecular bonding and reduced chain mobility in the film matrix (Qi and Li, 2024). In other studies of anthocyanin-based films, incorporation of anthocyanin compounds led to changes in mechanical properties that included reductions in elongation at break, confirming the trend observed here, where increased bioactive loading reduced film flexibility (Chen et al., 2023).

Barrier properties

The water solubility values of κ-C: Hm films incorporated with different concentrations of RA are presented in Table 1; Figure 2. The control film (κ-C: Hm RA₀) exhibited the highest solubility (80.06 ± 0.27%), while the film containing 2.0% RA (κ-C: Hm RA₂) showed the lowest solubility (70.27 ± 0.14%). Statistical analysis revealed that κ-C: Hm RA₀ and κ-C: Hm RA_0.5 were not significantly different from each other (p > 0.05), whereas κ-C: Hm RA_1.5 and κ-C: Hm RA₂ formed a statistically similar group. The progressive decrease in water solubility with increasing anthocyanin concentration may be attributed to enhanced intermolecular interactions between anthocyanin molecules and the polymer matrix, which reduces the availability of free hydrophilic groups for water binding. In the control film, the absence of anthocyanin allows κ-carrageenan and HPMC to interact freely with water, resulting in higher solubility. In contrast, anthocyanin incorporation may promote hydrogen bonding and partial crosslinking within the polymer network, thereby reducing film dissolution. Similar reductions in water solubility upon incorporation of plant extracts or phenolic compounds have been reported for starch- and polysaccharide-based films. Fonseca-García et al. (2021) observed decreased solubility in starch and chitosan composite films containing functional additives due to increased matrix cohesion. Likewise, Wang et al. (2021) reported a reduction in water solubility of cornstarch-based films incorporated with natural plant extracts, attributing the effect to strengthened intermolecular interactions and reduced polymer leaching. Comparable trends were also reported by Prietto et al. (2017) for anthocyanin-based pH-sensitive films, supporting the results obtained in the present study. The observed solubility behavior indicates that roselle anthocyanin incorporation enhances the structural integrity of κ-C: Hm films while reducing their water solubility, which may be advantageous for applications requiring moderate water resistance in intelligent food packaging systems.

The swelling ratio values of κ-carrageenan: HPMC films containing different concentrations of RA are presented in Table 1; Figure 2. The control film (κ-C: Hm RA₀) exhibited the highest swelling ratio (6.55 ± 0.09), while a progressive reduction in swelling was observed with increasing RA concentration. The lowest swelling ratio was recorded for κ-C: Hm RA₂ (1.85 ± 0.02). Statistical analysis indicated that κ-C: Hm RA_1.5 and κ-C: Hm RA₂ were not significantly different from each other (p > 0.05), whereas all other treatments differed significantly. The high swelling behavior of the control film can be attributed to the hydrophilic nature of κ-carrageenan and HPMC, which contain abundant hydroxyl and sulfate groups capable of forming hydrogen bonds with water molecules. The reduction in swelling ratio following anthocyanin incorporation suggests increased intermolecular interactions between anthocyanin molecules and the polymer chains, leading to reduced polymer relaxation and restricted water penetration into the matrix. Similar reductions in swelling behavior upon incorporation of plant-derived phenolic compounds have been reported in polysaccharide-based films. Maran et al. (2013) observed a decrease in the swelling ratio of biopolymer films due to enhanced structural cohesion and a reduced availability of hydrophilic sites. Comparable trends were also reported for anthocyanin-based pH-sensitive films, where increasing anthocyanin concentration resulted in lower water uptake and swelling due to strengthened polymer networks (Prietto et al., 2017; Wang et al., 2021). The reduction in swelling ratio with increasing roselle anthocyanin concentration indicates enhanced water resistance and improved structural stability of the films, which is desirable for their application as freshness-indicating materials in humid food packaging environments.

The κ-C:Hm RA films exhibited notable stability over a wide pH range (pH 2–12), retaining their structural integrity in both acidic and alkaline environments, as shown in Figure 3. While the films remained largely intact across the pH range tested, minor disintegration was observed at elevated pH levels. This behavior may be attributed to partial disruption of hydrogen bonding and other intermolecular interactions within the polymer matrix, particularly as hydroxyl and sulfate groups become ionized under alkaline conditions, weakening cohesion and structural integrity. Similar pHresponsive behaviors have been reported by Avila et al. (2022) in anthocyanin-based carrageenan films, where colorimetric and structural changes were observed as pH shifted from acidic to basic conditions due to alterations in anthocyanin molecular species and polymeranthocyanin interactions (e.g., carrageenan films with natural anthocyanin additives showed distinct responses to pH changes). Although total pigment yield was not determined, the retained color intensity and clear pH-responsive behavior of the films indicate that anthocyanins remained functionally stable during film formation.In addition, hydroxypropyl methylcellulose/carrageenan blends containing anthocyanins demonstrated notable pH sensitivity and maintained film integrity over a broad pH range, supporting the use of such blends for pH-responsive food packaging applications (Sun et al., 2019).

When packaging oily or oil-containing foods, effective resistance to oil permeation is essential to maintain product quality and prevent leakage. In this study, the oil barrier performance of κ-C: Hm films was evaluated by inverting oil-filled test tubes over filter paper covered with the film and incubating at 25 °C for 24 h. The results showed no observable oil penetration, indicating that the developed films possess effective oil barrier characteristics. The ability of polysaccharide-based films to resist oil permeation has been attributed to their hydrophilic polymeric matrix, which tends to reject non-polar oil molecules, thereby limiting oil diffusion through the film network. Similar observations have been reported for edible films based on polysaccharides such as κ-carrageenan, whey protein isolate, and other hydrophilic biopolymers, which exhibited low oil permeability due to the incompatibility between hydrophilic polymers and non-polar oil phases (Wang et al., 2007; Kokkuvayil Ramadas et al., 2024). In addition, studies on carrageenan composite films reinforced with essential oils or natural extracts have indicated that incorporation of hydrophobic moieties can further enhance oil barrier properties by increasing tortuosity for polar and non-polar permeants within the film matrix (da Rosa et al., 2020; Prietto et al., 2017). These findings support the effective oil barrier function observed in the κ-C: Hm films developed in the current work, underscoring their potential utility in packaging high-fat or oil-rich food products.

Antioxidant activity

The antioxidant activity of κ-C: Hm films incorporated with varying concentrations of RA is presented in Table 1; Figure 2. The control film (κ-C: Hm RA₀) showed zero DPPH inhibition, confirming the absence of inherent radical scavenging ability in the base polymer matrix. Antioxidant activity increased significantly with RA concentration (p < 0.05), with κ-C: Hm RA_0.5, κ-C: Hm RA₁, and κ-C: Hm RA_1.5 exhibiting 64.67 ± 0.88%, 68.19 ± 1.42%, and 70.42 ± 1.05% inhibition, respectively. The highest antioxidant activity was observed in κ-C: Hm RA₂ (80.63 ± 1.26%), indicating enhanced radical scavenging with higher anthocyanin content. The increased DPPH radical scavenging activity found in anthocyanin-enriched films is in alignment with recent studies on biopolymer-based active packaging systems, which showed that adding phenolic-rich natural extracts to carrageenan HPMC matrices significantly enhanced antioxidant capacity because phenolic hydroxyl groups can donate hydrogen to DPPH radicals (Kokkuvayil Ramadas et al., 2024). Similarly, carrageenan TiO₂-doped films with sweet potato peel anthocyanins exhibited strong antioxidant activity in DPPH and ABTS assays, reflecting the synergistic effects of anthocyanins and nanofillers (Riahi et al., 2024). Anthocyanin-rich smart films based on Mexican petunia anthocyanins also demonstrated improved free radical scavenging relative to films without extract (Kanphai et al., 2025). These findings substantiate the present observation that the antioxidant capacity of biopolymer films increases proportionally with anthocyanin content, enhancing their functional properties as active packaging materials.

Structural properties

The FTIR spectra (Figure 4) displayed distinct absorption bands linked to the functional groups of the κ-C: Hm RA film constituents. The control film (κ-C: Hm RA₀) showed a wide absorption band near 3324 cm⁻¹, linked to O–H stretching vibrations of polysaccharides, suggesting considerable intra and intermolecular hydrogen bonding. The band around 2924 cm⁻¹ is related to C–H asymmetric stretching, whereas the peak at approximately 1640 cm⁻¹ is linked to bound water in the polymer matrix (Kraisit et al., 2017; Liew et al., 2017; Jairam et al., 2025). Absorption bands validated the distinctive sulfate ester groups of κ-carrageenan at around 1218, 1155, and 844 cm⁻¹, aligning with described FTIR characteristics of κ-carrageenan-derived films (Qin et al., 2024; Zhong et al., 2025; Mu et al., 2025). The peak in the 920–845 cm⁻¹ range signifies the presence of 3,6-anhydrogalactose units, crucial for the formation of κ-carrageenan gels. With the addition of roselle anthocyanin (κ-C: Hm RA_0.5–RA₂), the FTIR spectra preserved the key structural characteristics of the base polymers, suggesting strong compatibility (Ren et al., 2025; Zhong et al., 2025; Mu et al., 2025). A minor change and widening of the O–H stretching band (≈3356–3328 cm⁻¹) indicated improved hydrogen bonding between the hydroxyl groups of the polymer and the phenolic parts of anthocyanins. The appearance and enhancement of bands between 1630–1640 cm⁻¹ were linked to aromatic C=C stretching vibrations of anthocyanins, verifying their effective integration into the film matrix. Extra absorptions found in the 1730–1785 cm⁻¹ range within RA-containing films relate to C=O stretching vibrations tied to anthocyanin structures and lesser ester or carboxylic functionalities (Ren et al., 2025; Zhong et al., 2025; Zhao et al., 2022). The sulfate ester bands of κ-carrageenan (≈1215–846 cm⁻¹) remained noticeable in all formulations, suggesting that the essential carrageenan structure was maintained following the addition of anthocyanins. Bands in the range of 1030–1060 cm⁻¹ were attributed to C–O–C and C–O stretching vibrations from glycosidic linkages, indicating the preservation of the polysaccharide structure. At higher RA concentrations, weak absorptions around 690–730 cm⁻¹ were observed, corresponding to aromatic ring deformations of anthocyanins, further supporting their presence within the polymer matrix (Kraisit et al., 2017; Zhong et al., 2025; Zhao et al., 2022). The FTIR results confirm the successful incorporation of roselle anthocyanin into κ-carrageenan/HPMC films without disrupting the primary polymer structure, while indicating molecular interactions mainly governed by hydrogen bonding and aromatic–polysaccharide interactions. Recent studies on carrageenan-anthocyanin films similarly report preserved backbones with shifted O-H bands due to hydrogen bonding. These features align with pH-responsive edible films using Hibiscus extracts, showing intensified aromatic peaks post incorporation (Latiff et al., 2025; Zhong et al., 2025; Ren et al., 2025).

Thermal properties

The TGA profile (Figure 5) revealed three main weight-loss stages from ambient temperature to 600 °C, consistent with multi-step degradation typical of polysaccharide-based films. The first weight loss (~15.1%) between 100–200 °C was primarily due to moisture evaporation and disruption of hydrogen-bonding networks within the hydrophilic film matrix, as reported in carrageenan/HPMC systems (κCHM), where water loss occurs below ~100–140 °C before polymer decomposition begins (Guo et al., 2022; Kanphai et al., 2025).The second significant mass loss (~19.7%) between 200–300 °C corresponds to decomposition of plasticizers (e.g., sorbitol/glycerol) and partial breakdown of weaker polymer segments. Similar thermal transitions have been observed in carrageenan films, where decomposition of glycerol and side chains occurs in the 130–230 °C range (Bhatia et al., 2024). The third degradation stage, from 300 to 450 °C, is attributed to the main chain scission and complete breakdown of the biopolymer backbone, consistent with earlier carrageenan-based film studies, which show polymer decomposition above ~230 °C with residual char at higher temperatures (Bhatia et al., 2024; Kanphai et al., 2025).The multi-step degradation pattern confirms the complex thermal behavior of biopolymer films, influenced by moisture loss, plasticizer volatilization, and polysaccharide backbone degradation.

The developed roselle-anthocyanin films showed improved color responsiveness, acceptable mechanical strength, and stable thermal behavior, indicating successful integration of bioactive compounds into the polymer matrix. Similar findings were reported by Mu et al. (2025), who found that anthocyanin-based films exhibited enhanced pH sensitivity due to hydrogen-bonding interactions with the polymer network. While Mu et al. (2025) observed a slight reduction in thermal stability at higher pigment levels, the present study maintained relatively stable thermal properties, possibly due to stronger interactions among κ-carrageenan, HPMC, and roselle anthocyanins. These results confirm that the developed films perform competitively with recent intelligent packaging systems.

Proximate analysis

The moisture content of the κ-C: Hm RA0 film was the lowest (12.17 ± 0.63%), whereas κ-C: Hm RA2 exhibited the highest MC (15.09 ± 0.35%) shown in Table 1; Figure 3. The observed increase in moisture with RA incorporation is consistent with previously reported trends in hydrophilic polysaccharide films, where higher plasticizer or extract content increases the water-holding capacity of the polymer network. Similar MC values (~12–15%) have been reported in carrageenan and carrageenan-based biopolymer films, indicating hydrophilic matrix behavior and absorption of ambient moisture (Kristó et al., 2024; Ramli et al., 2024).

Although the moisture-related properties were evaluated, the absence of water vapor permeability (WVP) measurements limits a comprehensive interpretation of the film’s moisture-barrier performance. WVP is a critical parameter for understanding mass transfer behavior and predicting real-world packaging functionality under different humidity conditions. Therefore, future studies should include WVP analysis to provide a more complete assessment of barrier efficiency and to better correlate structural modifications with functional performance.

Biodegradable analysis of films

Biodegradation behavior of films shown in Figures 6, 7 depends on physicochemical composition, crystallinity, and polymer interactions affecting microbial access and water penetration. Hydrophilic polysaccharide films like κ-C: Hm RA degrade faster in soil burial due to moisture absorption and microbial hydrolysis. In this study, weight loss was monitored over soil burial time (Martiny et al., 2020; Zhou et al., 2022; Kokkuvayil Ramadas et al., 2024).​ As shown in Figures 5, 6, κ-C: Hm RA2 film displayed continuous weight loss over 21 days, with linear degradation from polysaccharide susceptibility to soil enzymes. Similar linear trends occur in carrageenan/cellulose films under soil burial (Zhou et al., 2022; Kokkuvayil Ramadas et al., 2024).​ The pattern suggests near-complete degradation within ~2 months, aligning with polysaccharide film reports showing full breakdown in 7–30 days. Recent carrageenan composites confirm 70–100% weight loss in 20–30 days (Martiny et al., 2020; Kokkuvayil Ramadas et al., 2024; Eelager et al., 2026).

Freshness monitoring of button mushrooms using indicator film

The practical applicability of the κ-C:Hm RA₂ freshness-indicating film was evaluated on button mushrooms (Agaricus bisporus), selected for their high respiration rate and rapid spoilage at 25 °C. The optimized film was chosen for its superior antioxidant activity and pH-sensitive color response (Remedio and Parada Quinayá, 2024; Kuswandi et al., 2024; Ren et al., 2025). As shown in Table 2, mushrooms stayed fresh with firm texture up to 8 h, with the film retaining red color. After 16 h, package moisture rose from transpiration, but no spoilage odor appeared and film change was minimal. At 19–24 h, mushroom discoloration and off-odors coincided with film darkening (Remedio and Parada Quinayá, 2024; Kuswandi et al., 2024; Zhan et al., 2024). Beyond 28 h, strong odors and deterioration matched complete red-to-black film transition. Full spoilage occurred at 32–36 h, confirmed by black film. This color shift reflects roselle anthocyanin response to spoilage volatiles raising pH. Recent anthocyanin films on mushrooms show comparable red-to-dark transitions for real-time monitoring (Liu et al., 2022; Remedio and Parada Quinayá, 2024; Kumar et al., 2025).

Table 2. Observations of packed mushroom with κ-C: Hm RA2 film at 25°C.

Conclusion

This study successfully developed a biodegradable, freshness-indicating film by incorporating roselle anthocyanin into a κ-carrageenan and HPMC polymer matrix. The formulated films exhibited reliable mechanical strength, satisfactory barrier properties, and strong pH-responsive color changes, along with enhanced antioxidant activity proportional to anthocyanin concentration. Structural and thermal analyses confirmed good compatibility and stability of the composite system, while soil burial tests demonstrated complete biodegradation within two months. The optimized film effectively indicated spoilage of button mushrooms through a clear visual color transition, highlighting its practical applicability. Overall, the developed film offers a sustainable, consumer-friendly intelligent packaging solution with strong potential for monitoring the freshness of perishable foods and reducing food waste.

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Footnotes

Edited by: Dipendra Kumar Mahato, Deakin Univeristy, Australia

Reviewed by: Myleidi Vera, University of Concepcion, Chile

Abimbola Ige, University of Minnesota Duluth, United States