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Article

Mycomaterials from Agave Bagasse: A Valorization Strategy for Sustainable Tequila Packaging

by
Flavio A. de Anda-Rodríguez
1,
Mariana R. Corona-Ramírez
2,
Carlos D. Patiño-Arévalo
1,
Marco A. Zárate-Navarro
2,*,
Ana I. Zárate-Guzmán
1,* and
Luis A. Romero-Cano
1
1
Grupo de Investigación en Materiales y Fenómenos de Superficie, Departamento de Biotecnológicas y Ambientales, Universidad Autónoma de Guadalajara, Av. Patria 1201, Zapopan 45129, Jalisco, Mexico
2
Departamento de Biotecnológicas y Ambientales, Universidad Autónoma de Guadalajara, Av. Patria 1201, Zapopan 45129, Jalisco, Mexico
*
Authors to whom correspondence should be addressed.
Fermentation 2025, 11(9), 500; https://doi.org/10.3390/fermentation11090500 (registering DOI)
Submission received: 15 July 2025 / Revised: 21 August 2025 / Accepted: 21 August 2025 / Published: 26 August 2025
(This article belongs to the Special Issue Application of Fungi in Bioconversions and Mycoremediation)
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Versions Notes

Abstract

A sustainable strategy is proposed for the valorization of solid waste from the Tequila industry through the development of bio-packaging for Tequila bottles using mycelium from Ganoderma lucidum. The fungus was isolated from Bosque de la Primavera (Jalisco, Mexico) and cultivated on lignocellulosic substrates: agave bagasse and corn stover. These agricultural residues were dried, ground, and pasteurized to optimize their performance as growth media. Their structural integration before and after fermentation was evaluated using optical microscopy. The high cellulose and hemicellulose content of both substrates supported robust mycelial development, enabling the formation of moldable materials through solid-state fermentation. After growth, the mycelium colonized the substrate, forming a functional mold adapted to the geometry of a Tequila bottle prototype. The molded parts were dried to halt fungal activity, prevent fruiting, and stabilize the structure. Physical and mechanical characterization showed competitive performance with regard to bulk density (0.11 ± 0.1 g cm−3), water absorption (78.1 ± 4.2%), and high impact resistance (evaluated via Solidworks simulation). A life cycle assessment revealed that mycelium packaging has a significantly lower environmental impact than expanded polystyrene. The material supports circular economy principles within the Tequila production chain.

1. Introduction

The Agave–Tequila production chain, emblematic of Mexico’s cultural and economic heritage, is under increasing scrutiny due to its environmental impact, particularly regarding carbon emissions and solid waste generation. Recent studies indicate that producing one liter of Tequila (40% alcohol v/v) emits approximately 3 kg of CO2 eq, with 41% of this footprint attributed to the industrial transformation stage and up to 20% associated with packaging production, particularly due to the use of high-energy-demand materials such as glass, cardboard, and plastics [1].
In response to these environmental concerns, the 2016 Sustainability Strategy for the Agave–Tequila Production Chain, led by the Tequila Regulatory Council (CRT, by its acronym in Spanish), proposed a set of goals to be achieved by 2030. These include a 25% reduction in the carbon footprint per liter of Tequila, a 12% increase in the use of renewable energy, and an 80% treatment rate of process water [2]. To achieve these targets, various actions have been implemented, such as the transition to cleaner fuels [3,4] and the development of water treatment systems using agave bagasse materials [5,6,7,8,9].
However, a relatively unexplored but promising strategy is the valorization of agave bagasse as a precursor for biomaterials. The industry generates large amounts of agave bagasse, a lignocellulosic byproduct traditionally underutilized and usually discarded after sugar extraction during Tequila production [10]. This solid residue represents a significant waste, which contributes to environmental burdens and poses waste management challenges. Agave bagasse is rich in structural polysaccharides, with reported compositions of cellulose (30–45%), hemicellulose (20–30%), and lignin (15–25%), along with small amounts of extractives and ash [11]. These physicochemical characteristics, including a porous microstructure and high water absorption capacity, make it an attractive candidate as a substrate for fungal colonization and as a biological reinforcement agent in mycelium-based composite materials, which have gained interest for their low-energy fabrication, biodegradability, and potential to replace traditional packaging materials [12,13,14]. Despite growing interest in mycelium technology, its integration with agave-derived residues remains largely unaddressed in the context of the Tequila industry.
Recent advances in mycelium-based materials have demonstrated their applicability in creating lightweight, durable, and compostable packaging solutions [15,16]. These biofabricated materials are formed by growing fungal mycelium through lignocellulosic substrates under controlled conditions, producing a foam-like structure with tunable mechanical and thermal properties [17,18]. Mycelium materials have been developed for a wide range of applications, including paper, textiles, foams (for packaging, acoustic, and medical uses), vehicle parts, and electronics, with a growing number of patents reflecting their practical potential [19,20,21]. For instance, mycelium-based foam packaging is currently produced by Ecovative Design and Sealed Air and used by companies like Dell and IKEA for product packaging [22].
Hence, this research work presents the development of sustainable packaging prototypes composed of fungal mycelium grown on agave bagasse, with the specific goal of packaging Tequila bottles. This work aims not only to demonstrate the technical feasibility and environmental value of this bio-based alternative, but also to promote scientific dissemination and public awareness regarding the social and environmental responsibilities of the agave and Tequila production chain. By linking material innovation with the principles of the circular economy, this study contributes to the broader vision of a low-impact and socially responsible Tequila industry.

2. Materials and Methods

2.1. Formulation and Growth Conditions of Mycelium-Based Biocomposites

To obtain mycelium-based biocomposite for sustainable packaging applications, agricultural residues were used as lignocellulosic substrates to support fungal growth and material formation. The selected fungal strain, Ganoderma lucidum, was isolated from the Bosque de la Primavera, located at the West of the Guadalajara Metropolitan Area, Jalisco, Mexico [23,24,25], and cultivated under controlled laboratory conditions.
The composite formulation consisted of 90% substrate and 10% fungal inoculum. The substrate was a mixture of agave bagasse (50%) and corn stover (50%), both of which are abundant agro-industrial by-products of the Tequila and maize production chains, respectively. These raw materials were subjected to a preconditioning process that included air-drying, grinding to particles with a maximum dimension of approximately 2.5 cm, and pasteurization at 65–80 °C to reduce microbial contamination and competition.
Once cooled, the pasteurized substrates were inoculated with homogenized mycelium and manually packed into custom-designed molds compatible with prototype Tequila bottle shapes. The inoculated molds were incubated under ambient temperature (24–28 °C) and high-humidity conditions (>80%), allowing for effective colonization over a period of 5 to 7 days. During this growth phase, adequate oxygen flow was ensured to promote uniform mycelial development throughout the matrix.
After full colonization and structural consolidation, the biocomposites were demolded and dried at temperatures of 45–60 °C to halt further biological activity and enhance mechanical stability. This post-processing step is critical to ensure durability and prepare the material for subsequent physical characterization and life cycle analysis.
Micrographs of the samples were obtained using a digital microscope (model MIC-600, 50×–1000×, Steren, Mexico City, Mexico) and an optical microscope (model B120A, 40×–1600×, United Scope LLC, Irvine, CA, USA) to document surface morphology and structural features.

2.2. Methodology for Design and Fabrication of 3D-Printed Mold for Mycelium-Based Packaging

To manufacture the mycelium-based biocomposite packaging, a custom mold was designed to fit the dimensions of a prototype Tequila bottle (50 mL). The mold consisted of two interlocking parts (base and lid) and was fabricated using fused deposition modeling (FDM) 3D-printing technology (Ultimaker S5, Ultimaker BV, Utrecht, The Netherlands). Polylactic acid (PLA) filament was selected as the printing material due to its rigidity, ease of use, and biodegradability. The design was modeled using SolidWorks 2024 (SP5, Dassault Systems, Waltham, Massachusetts, USA) and printed with a nozzle diameter of 0.4 mm and a layer height of 0.2 mm to ensure dimensional accuracy and smooth wall surfaces. Prior to inoculation, all tools and the interior of the 3D-printed mold were sanitized using 70% isopropyl alcohol to minimize microbial contamination.
The inoculated mixture was compacted into the mold cavity and gently sealed with the printed lid. To maintain humidity and airflow during the incubation phase, the entire mold was wrapped with plastic film, which was perforated with sterile needles. Moisture was preserved through daily mist spraying. The mold was incubated under ambient conditions (25 °C and >80% relative humidity) for 6 days, allowing full colonization and structural consolidation.
After this period, the composite was unmolded and dried at 50 °C for 12 h to deactivate fungal activity and improve the material’s mechanical stability. The use of PLA and 3D printing enabled a reusable, low-cost mold manufacturing process that facilitated iterative design and prototyping of sustainable packaging solutions.

2.3. Methodology for Physical and Mechanical Characterization of the Mycelium-Based Packaging

To evaluate the performance of the mycelium-based packaging developed from agave bagasse and corn stover, physical and mechanical tests were conducted in series following standard procedures.
The thermal behavior of the mycelium-based biocomposite packaging was assessed by placing square samples on a conventional laboratory hot plate preheated to 300 °C. Each sample was exposed to direct contact with the heated surface for 5 min. After exposure, visual changes such as surface charring, deformation, and structural integrity were documented. This test aimed to simulate extreme thermal conditions to assess the material’s resistance to high temperatures.
Bulk density (g/cm3) was calculated from the measured mass and volume of the dried biocomposite. Volume was determined based on the external dimensions of the packaging unit, while mass was measured using an analytical balance. This property provides insight into the material’s lightness and suitability for protective applications [26].
Water absorption (%) was assessed by submerging the dried samples in distilled water for 24 h at room temperature. The increase in weight was recorded and expressed as a percentage of the initial dry mass. This test evaluates the material’s susceptibility to moisture, an important factor for packaging performance under varying environmental conditions [27].
Impact resistance was evaluated according to ASTM D5276 standard [28], which involved a free-fall drop test from a height of 76 cm onto a hard surface. The test was conducted to simulate real-world handling and shipping conditions. The integrity of the mycelium-based packaging was visually inspected after the drop to assess its ability to protect the bottle-shaped cavity and maintain structural cohesion [29]. Additionally, a drop test simulation was performed in SolidWorks 2024 Simulation to evaluate the mechanical response of the mycelium-based packaging under impact conditions.
These characterization methods collectively provide a comprehensive evaluation of the material’s viability as a sustainable alternative to conventional packaging solutions such as expanded polystyrene.

2.4. Life Cicle Assesment (LCA)

The LCA study was performed using OpenLCA software (ver. 2.5.0, GreenDelta, Berlin, Germany), following the methodology reported by Gomez-Navarro et al. [5].

2.4.1. Goal and Scope

The LCA study is based on the ISO 14,044 standardized methodology [30]. The goal and scope of this step were to evaluate the environmental impact of a biocomposite production from agave bagasse and mycelium from the Ganoderma lucidum fungus. The system boundary followed a gate-to-gate approach, as detailed in Supplementary Figure S1. For the LCA, only the biocomposite production process was considered; the transportation and destination of the product and the original raw material processes were not included. The functional unit was defined as 1 kg of biocomposite obtained. According to the mass balance, 1 kg of biocomposite can produce 15.27 packages for several uses.

2.4.2. Inventory Analysis

The inventory analysis is shown in Supplementary Table S1. The input flows were energy, chemicals, water, and raw materials. Energy types were considered based on CFE (Comision Federal de Electricidad—México) data [31]. Energy was used mainly in the steps of sterilization, pasteurization, incubation, and dehydration. The only chemical used was potato dextrose agar for mycelium growth. The water consumed in all the steps of the process was considered to be fresh water, commonly sourced in Mexico. Agave bagasse, corn stover, and mycelium were the raw materials used for the biocomposite. The output included waste, emissions, wastewater, and material production (biocomposite).

2.4.3. Impact Assessment

The quantification of environmental impacts was performed using OpenLCA software, employing the ELCD 3.2 GreenDelta database. The ReCiPe 2016 midpoint (H) method was used, including 18 impact categories: fine particulate matter formation, fossil resource scarcity, freshwater ecotoxicity, freshwater eutrophication, global warming, human carcinogenic toxicity, human non-carcinogenic toxicity, ionizing radiation, land use, marine ecotoxicity, marine eutrophication, mineral resource scarcity, ozone formation-human health, ozone formation-terrestrial ecosystems, stratospheric ozone depletion, terrestrial acidification, terrestrial ecotoxicity, and, water consumption. This allowed a high level of detail and technical specificity.

2.4.4. Interpretation and Comparison Analysis

The most relevant steps and flows that most significantly contributed to environmental impacts were identified. Additionally, with the aim of comparing the biocomposite material with a commonly used material for packaging, the LCA of expandable polystyrene granulate (EPS) was performed using a predefined process in the ELCD GreenDelta database. This material was selected because density analysis indicated that almost the same quantity of EPS would be required to produce a package comparable in volume/functionality to the biocomposite. The functional unit (1 kg of product) was the same for both materials.

3. Results and Discussion

3.1. Growth Kinetics

The mycelial radial growth followed an exponential model: A m m 2   =   520.78 e 0.5226 t − 537.56 , with a high coefficient of determination (R2 = 0.9915), as shown in Figure 1. Over four days, the average colonization rate was 266.4 mm2/day, and the radial expansion rate was estimated at 4.37 mm/day. These values were derived from digital image analysis, which allowed time-resolved quantification of mycelial growth.
Image processing was performed using a MATLAB (ver. R2024b, MathWorks, Natick, MA, USA)-based image analysis routine, which enabled accurate segmentation and measurement of the colonized surface area. This approach is consistent with previously reported methods in macroscopic and hyphal-level image analysis, which have proven effective and accessible for characterizing fungal growth [32,33,34,35,36,37].
The resulting kinetic parameters provide information about the optimization of growth conditions in more complex geometries, such as the Tequila box mold [38]. In this context, the lignocellulosic substrate composed of agave bagasse and corn stover functions not only as a nutrient source but also as a structural matrix to shape the biocomposite [39].
Understanding fungal growth kinetics under room temperature conditions is essential for planning large-scale production where environmental control may be limited. Although a deeper optimization involving multiple variables is beyond the scope of this study, this basic kinetic evaluation serves as a foundational tool for production planning and process modeling [40,41,42].
The exponential model used here is particularly appropriate, as it describes growth in closed systems. Similar conditions have been reported in studies where sealed molds are used under batch processes, which emphasize maintaining the exponential phase to ensure optimal colonization [43]. The mycelial growth rate observed was consistent with previous reports using PDA medium under similar conditions. At day 4, the colony covered an estimated area of 3674.62 mm2, calculated from an exponential growth model with a coefficient of determination (R2) > 0.99. Comparable studies reported areas of 2212.88 mm2 for G. lucidum grown on PDA with varying malt concentrations at day 4 [44] and 3280.72 mm2 for mycelial growth in composite material applications [45]. The slightly higher value obtained here may be attributed to strain-specific growth potential and the nutrient-rich composition of the PDA medium used, which can enhance hyphal extension under optimal incubation conditions. This outcome remains within the expected range for G. lucidum, reflecting the rapid expansion typical of colonies on nutrient-rich agar media.

3.2. Growth Characterization of Ganoderma lucidum on Potato Dextrose Agar (PDA) Medium and Lignocellulosic Substrate (Agave Bagasse)

Figure 2a shows the agave bagasse used as substrate. The image reveals a multi-layered structure with well-defined, interconnected channels along the fibers. The fibers had an average length of 10 cm and a diameter of 108.00 ± 6.55 μm, consistent with previous studies that have characterized the highly porous and fibrous morphology of agave bagasse [5]. These features are favorable for fungal colonization, as they support extensive mycelial development and enhance oxygen availability [46]. Moreover, the digital microscopy image illustrates the surface texture of the agave fibers, highlighting the roughness and longitudinal arrangement of the vascular bundles, which contribute to the mechanical interlocking and structural cohesion within the composite. In addition, the elemental composition previously obtained via EDX analysis [5] showed high content of carbon (57.19 wt.%) and oxygen (37.49 wt.%), confirming the predominance of cellulose and hemicellulose, which are essential components for fungal growth and binding. Trace elements such as calcium, nitrogen, and magnesium were also detected; these may contribute to fungal metabolism. These characteristics position agave bagasse as a suitable and sustainable alternative to traditional conventional lignocellulosic substrates used in other mycelium-based composites.
Figure 2b shows the mycelial growth of Ganoderma lucidum on PDA (potato dextrose agar), displaying a dense, uniform white mycelium with a filamentous and cotton-like texture. The morphology and steady radial expansion observed, as seen in the insert of Figure 1, are typical of G. lucidum strains under laboratory conditions and are consistent with the ranges reported in previous studies [40,41,42,43]. No atypical growth patterns were detected in terms of rate or morphology, reinforcing the strain’s consistency with established references and supporting its selection for subsequent colonization assays on lignocellulosic substrates.
Figure 2c–f show the interaction between G. lucidum mycelium and the agave bagasse/corn stover substrate. The white hyphal network densely envelops the fibrous material, demonstrating not only extensive colonization but also effective adhesion to both the agave bagasse and corn stover. This robust attachment facilitates structural integration, essential for composite formation. The mycelial growth exhibits an intricate network of intertwined hyphae closely integrated with the agave bagasse and corn stover fibers and suggests good compatibility between the fungus and the substrate. This pattern is consistent with previously reported colonization behaviors of G. lucidum strains on lignocellulosic substrates [45]. These observations confirm the substrate’s suitability to support fungal proliferation and biointegration, providing satisfactory outcomes in line with the aims of this research.
Finally, the mycelium-based biocomposite composed of lignocellulosic biomass was exposed to 300 °C for 5 min to assess its thermal performance (Figure 2g–i). This temperature was selected to simulate potential accidental exposure to high heat, such as during a fire, and to evaluate the material’s thermal stability and charring behavior for safety purposes. Although not representative of normal use conditions, this stress test provided insight into the resilience of the biocomposite packaging. Results showed excellent structural stability, typical of lignocellulosic biocomposites [47], with minimal deformation or loss of integrity. The material’s resilience is attributed to its dense mycelial network and the natural binding properties of lignin, which promote rapid char formation during heating and are the main reasons for the popularity of these mycelium-based materials [13]. This char layer acts as a thermal barrier, protecting the underlying structure from significant degradation and demonstrating the material’s ability to withstand moderate thermal stress, which is typical of mycelium and mycelium composites [48]. These findings suggest potential applications requiring thermal insulation, making it suitable for Tequila bottles and comparable to the mycelium-based wine bottle packages already available [43]. The fractal growth pattern of mycelium, with its repeating, branching structures at multiple scales, contributes to this toughness by efficiently distributing stress throughout the material [49]. In this study, the thermal exposure was intended as a qualitative assay [50].
The results obtained indicate that the mycelium-based biocomposite based on Ganoderma lucidum, agave bagasse, and corn stover underwent thermal decomposition, including the breakdown of lignocellulosic components and the formation of a protective char layer. This layer played a crucial role in preserving the structural integrity of the material and acted as a thermal barrier, showcasing moderate fire-resistant properties. In the context of Tequila bottle packaging, this property is particularly relevant, as it may offer an additional layer of protection during storage or transportation under extreme conditions, such as accidental exposure to high temperatures.

3.3. Design and Fabrication of 3D-Printed Mold for Mycelium-Based Packaging

The design diagram and 3D renderings of the molds used for the fabrication of the mycelium-based packaging are presented in Figure 3 and Figure 4, respectively. The base (dimensions: 21.90 cm × 13.00 cm × 9.26 cm) includes an overhang that replicates the shape of the lid a standard 50 mL Tequila bottle prototype (Figure 3b). The top cover was designed with matching dimensions to fit snugly onto the base, enabling slight compression during the colonization phase, which supports mycelial binding and structural cohesion. Additionally, four ventilation holes (diameter: 0.55 cm) have been incorporated into the design (Figure 3a) to allow passive oxygen exchange during fungal growth.
Figure 4 shows the 3D-rendered model of the mold along with its assembled configuration. To maintain adequate humidity levels during incubation, the assembled mold was wrapped in plastic film. This film was perforated using sterile needles to allow gas exchange, and water was sprayed daily through the perforations to ensure a moist microenvironment. The incubation was carried out at room temperature (~25 °C) for six days, allowing complete colonization and visible white mycelial coverage on the substrate surface.
After the growth period, the fully colonized biocomposite was carefully demolded and subjected to thermal treatment at 50 °C for 12 h. This drying step served to halt further biological activity, stabilize the structure, and prepare the material for mechanical testing and subsequent applications.

3.4. Physical and Mechanical Characterization of the Mycelium-Based Packaging

Figure 5 illustrates the final appearance of the mycelium-based components. Each piece corresponds to a functional element of the proposed biodegradable packaging system. Figure 5a shows the top cover, which exhibits a homogeneous mycelial surface with well-defined hyphal structure and compactness. Some superficial discolorations and localized darkening were visible, probably due to minor inconsistencies in moisture distribution or localized microbial interaction. Nonetheless, the surface remained structurally coherent, with no signs of cracking or collapse.
Figure 5b presents the base, which includes a mold protrusion specifically designed to form a cavity shaped to fit a 50 mL tequila bottle prototype. This bottle-shaped indentation was successfully reproduced during colonization, resulting in a well-defined recess that allows the bottle to remain securely immobilized within the packaging. Dense mycelial growth was particularly evident in the recessed regions, indicating effective colonization even in complex geometries. Slight irregularities in pigmentation and surface texture were observed, possibly due to microenvironmental gradients in oxygen and humidity during incubation. These did not compromise the mechanical performance but indicate the importance of optimizing incubation uniformity for improved reproducibility.
Figure 5c,d depict a presentation-focused variant, intended for aesthetic product display. This component showed excellent edge fidelity and volume preservation. Its surface texture, while more porous and fibrous, is consistent with the natural aesthetic often associated with bio-based materials, making it suitable for display applications. These visual and structural attributes make it ideal for placing the packaging on product exhibition stands, with the purpose of promoting sustainability and circular economic practices within the Tequila industry.
Figure 6 illustrates the assembled prototype of the mycelium-based packaging, composed of the top and bottom parts previously described. This configuration was used in the final performance tests, including mechanical and physical evaluations. The assembled form demonstrates proper alignment and enclosure, with sufficient structural rigidity to immobilize the 50 mL Tequila bottle. A decorative ribbon was added to enhance the esthetic appeal and secure both components, reinforcing the packaging’s potential for premium product presentation.
Physical characterization of the assembled and dried components revealed a bulk density of 0.11 ± 0.01 g cm−3, confirming the lightweight nature of the composite, which is a desirable feature for sustainable packaging [51]. Additionally, the water absorption capacity of 78.1 ± 4.2% reflects the high hydrophilicity inherent to lignocellulosic substrates and fungal matrices [52]. While suitable for dry goods, this level of absorbency suggests that additional surface treatments or hydrophobic coatings may be necessary if the packaging is to be used in high-humidity environments or in contact with moisture-sensitive contents.
Despite minor surface irregularities, the final product retained its designed geometry and internal cavity, validating both the mold design and the substrate formulation. These findings highlight the feasibility of using fungal biocomposites in functional and marketable packaging solutions, while also indicating key areas for future optimization, such as environmental uniformity during growth and protection against moisture.
During the penetration and impact resistance tests, the mycelium-based packaging showed satisfactory structural integrity but exhibited surface fiber loss upon localized mechanical stress. This fraying or delamination effect was most evident at edges and high-pressure contact points, suggesting incomplete interfacial bonding between mycelial hyphae and larger lignocellulosic particles in the substrate. This behavior may be attributed to the coarse particle size or insufficient colonization in certain zones and indicates a potential area for improvement in compaction or growth conditions.
Additionally, during the drop test (Supplementary Video S1), the samples demonstrated an elastic response, evidenced by slight rebound upon impact. This behavior suggests that the composite exhibits low brittleness and moderate energy-absorption capacity. The ability to dissipate impact forces without catastrophic failure indicates potential for protective packaging applications; however, the observed rebound should be considered when designing for fragile contents, as it could affect product stability after collision [53].
These qualitative observations complement the quantitative measurements and provide insight into the material’s structural behavior under real-world mechanical stresses.
Finally, Figure 7 shows the interior of the packaging, highlighting its use for containing a bottle of Tequila. The design provides a precise and secure fit for the bottle, which is essential for product protection during handling and transportation, while also allowing elegant and stable display of the bottle. The way the bottle rests within the mycelium packaging emphasizes its contents, turning the packaging into an integral element of the product presentation.
Beyond its protective functionality, this prototype represents a powerful tool for scientific dissemination and a symbol of commitment to sustainability within the Tequila industry. The texture of the packaging is consistent with the natural esthetic associated with biobased materials, conferring a unique visual identity that resonates with values of sustainability and environmental respect. By presenting Tequila bottles within this innovative packaging, Tequila companies can tangibly communicate their interest and leadership in adopting circular economy practices and reducing their environmental footprint.
The packaging’s esthetics make it ideal for capturing consumer attention at points of sale and trade fairs, being visually distinctive. This prototype not only demonstrates a practical application for agave bagasse, a significant residue from Tequila production, but also becomes an effective means to educate the public about the potential of renewable materials and the future of responsible packaging.

Drop Test Simulation

To evaluate the mechanical behavior of the mycelium-based packaging under drop conditions, a dynamic drop test simulation was carried out using SolidWorks Simulation software. The 3D model of the packaging was designed in SolidWorks based on the actual dimensions of the mold used (21.90 cm × 13.00 cm × 9.26 cm), accurately replicating the internal volume corresponding to a standard 50 mL Tequila bottle.
The explicit drop simulation module was employed, setting an impact height of 762 mm onto a rigid surface to represent a common scenario during product handling. The packaging material was defined using the following mechanical properties: elastic modulus: 1.5 MPa [54], Poisson’s ratio: 0.3 [54], shear modulus: 0.577 MPa (calculated assuming linear elastic and isotropic behavior as an approximation), density: 110 kg/m3 (data from present study), tensile strength: 0.20 a 0.87 MPa (average: 0.535 MPa) [55], compressive strength: 0.25 to 1.87 MPa (average: 1.06 MPa) [55], and compressive yield strength: 47.5 kPa [55]. A contact condition with friction was defined between the ground and the package, as well as a refined solid tetrahedral mesh to ensure accurate resolution in the regions of highest deformation. The analysis considered a total impact duration of 0.0025 s.
Figure 8 presents the results of the drop test simulation for the packaging. Figure 8a shows the total displacement (URES), reaching a maximum value of approximately 2.4 mm at the lower corners of the package, corresponding to the initial point of contact with the ground. This global deformation reflects an elastic response of the material, without structural collapse. In Figure 8b, the distribution of equivalent strain (ESTRN) is shown. The highest strain values were in the front corner regions, where a slight concentration of stress was also observed, consistent with the main energy entry points during impact. The maximum strain was on the order of 4%, indicating that the material absorbed the impact without exceeding its structural capacity. Finally, Figure 8c presents the distribution of von Mises equivalent stress, reaching a maximum of approximately 13.17 kPa, which is well below the material’s compressive yield strength (47.5 kPa). This confirms that the package remained within the elastic region during the impact, with recovery capacity and no permanent damage.
These results are consistent with experimental observations, where a slight rebound of the material was reported after impact, characteristic of an elastic response with low stiffness. Surface friction and fiber detachment phenomena were also observed in areas of intense contact, suggesting that the interparticle bonding between lignocellulosic components could be improved through increased compaction or adjustments in colonization time.
The results demonstrate that the mycelium-based packaging exhibits adequate impact absorption capacity, with limited deformation, making it a promising candidate for biodegradable packaging applications for fragile products such as alcoholic beverages.

3.5. Lyfe Cycle Assesment

Figure 9 and Supplementary Table S1 show the LCA of biocomposite production. In the case of biocomposte material, the main impact categories were scarcity of fossil resources, global warming, and water consumption. Energy consumption contributes to fossil fuel scarcity with 8.43 kg oil eq, mainly through the use of energy from natural gas and oil, with contributions of 89% and 11%, respectively. Mycelium incubation accounts for the total CO2 emissions attributed to fungal respiration during this stage (0.027 kg CO2 eq), based on the average respiration rate reported in the literature [56]. In the case of water consumption, the mixed step is the main contribution to this category with a water requirement of 0.02 m3. Hazardous organic waste has no impact on any category due to the main waste being microbiological residues from culture medium; the biological risk is not significant. Final disposal is the main management of this waste. The results obtained for the biocomposite material were compared with the LCA results for EPS. In the fossil resource scarcity category, EPS showed a value of 1.39 kg oil eq (Figure 7a), while the biocomposite reached 8.43 kg oil eq. This indicates that the biocomposite makes a sixfold higher contribution to this category. This difference is mainly attributed to the fact that the energy matrix in our country relies heavily on fossil fuels (oil and natural gas), whereas the EPS data were based on a process powered by cleaner energy sources. The global warming impact of EPS was estimated to be 2.77 kg CO2 eq (Figure 9b), being 100 times greater than the contribution of biocomposite; this represents an important advantage in the use of this green material. Finally, regarding water consumption (Figure 9c), EPS requires around 0.011 m3 per 1 kg of material, while for the biocomposite, 0.001 m3 of water per kg is needed, representing less impact in the water consumption category. The rest of the impact categories are listed in Supplementary Table S2. It is evidenced that EPS has a higher contribution to environmental impact than the biocomposite. For example, in the case of toxicity, EPS production has a considerable contribution to terrestrial toxicity (0.841 kg 1,4-DCB), terrestrial acidification (5.7 × 10−3 kg SO2 eq), and ozone formation (3.8 × 10−4 kg Nox eq). This effect is mainly due to the use of chemicals and petroleum having significant impact on ecosystem quality, climate change, and human health [57].
While this study primarily focused on technical feasibility and environmental performance through LCA, a holistic understanding of sustainable materials, particularly for consumer products, inherently demands consideration of social acceptance. Mycomaterials, despite their significant ecological advantages, often face challenges related to their aesthetic appearance and intrinsic odor—factors that critically influence design quality and user experience. As highlighted by Bonenberg et al. [58], a notable disparity exists between the acceptance of ecological materials in professional design (e.g., by architects) and their broader adoption in domestic or consumer settings. This disconnect underscores a critical barrier to the widespread implementation of novel bio-based solutions. Acknowledging these factors is crucial for a realistic assessment of the material’s market readiness and for delineating future research. Therefore, future work will focus on improving the sensory and aesthetic appeal of the mycelium-based packaging. This includes investigating methods for odor mitigation, such as optimized drying processes and substrate pre-treatments, as well as exploring the potential for surface enhancements, like laser engraving, or the incorporation of natural fragrances to address the comprehensive consumer experience beyond mere odor masking.

4. Conclusions

This study demonstrated the technical and environmental feasibility of producing mycelium-based packaging using agave bagasse and corn stover as lignocellulosic substrates. The biocomposites developed with Ganoderma lucidum showed promising physical and mechanical properties, including low density, high water absorption, and elastic impact response, making them suitable for protective, biodegradable packaging of Tequila bottles. The life cycle assessment confirmed a significantly lower environmental impact compared with expanded polystyrene, particularly in terms of carbon footprint and water consumption, aligning with circular economy principles in the agave–Tequila production chain. The proposed strategy contributes to the circular economy of the Tequila industry, the main generator of agave bagasse.
However, some limitations remain. The high water absorption suggests the need for additional surface treatments, and partial fiber detachment during impact indicates opportunities for improving internal cohesion through better substrate compaction or colonization conditions. Future work should focus on enhancing moisture resistance, optimizing substrate composition, and scaling the process under real-world conditions. The integration of material innovation with sustainable design offers a compelling strategy for reducing the environmental footprint of traditional industries.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/fermentation11090500/s1, Supplementary Video S1.: https://youtu.be/KJ8nEg96ydE accessed on 20 August 2025, Drop test of mycelium-based packaging from agave bagasse: a valorization approach for sustainable Tequila packaging. Supplementary Figure S1. Production process of the biocomposite for mycelium-based packaging. The dotted red line represents the boundaries of the system for a gate-to-gate life cycle assessment. Supplementary Table S1. Inventory analysis for LCA. Supplementary Table S2. Comparison of LCA of biocomposite packaging vs. expanded polystyrene granulate.

Author Contributions

Conceptualization, F.A.d.A.-R. and L.A.R.-C.; methodology, L.A.R.-C., M.A.Z.-N. and A.I.Z.-G.; software, M.R.C.-R., C.D.P.-A., M.A.Z.-N. and A.I.Z.-G.; validation, M.R.C.-R. and C.D.P.-A.; formal analysis, M.A.Z.-N., A.I.Z.-G. and L.A.R.-C.; investigation, F.A.d.A.-R.; resources, M.A.Z.-N. and L.A.R.-C.; data curation, M.A.Z.-N., A.I.Z.-G. and L.A.R.-C.; writing—original draft preparation, all authors; writing—review and editing, all authors; visualization, M.A.Z.-N., A.I.Z.-G. and L.A.R.-C.; supervision, M.A.Z.-N., A.I.Z.-G. and L.A.R.-C.; project administration, M.A.Z.-N., A.I.Z.-G. and L.A.R.-C.; funding acquisition, M.A.Z.-N. All authors have read and agreed to the published version of the manuscript.

Funding

This research was partially funded by COECyTJAL, through FODECIJAL-11178 project for the acquisition of the computer equipment used for the kinetic study and drop test simulation, and the optical microscope used for the qualitative inspection following thermal exposure.

Data Availability Statement

The data that support the findings of this study are available from the corresponding author upon reasonable request.

Acknowledgments

M.R.C.-R. thanks to the Universidad Autónoma de Guadalajara for the financial support provided. C.D.P.-A., Thanks to the Universidad Autónoma de Guadalajara for the support through the Honoris causa scholarship. The authors thank Arantza Villa-González, Camila S. Gómez-Navarro, and Mariana García-Aceves for the support received during the drop tests (both experimental and computer-assisted simulation).

Conflicts of Interest

The authors declare no conflicts of interest.

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Fermentation 11 00500 g001
Figure 1. Digital image analysis for mycelium growth in potato dextrose agar (PDA) medium, kinetic characterization (fitted to y   =   a e b t + c model, R 2   =   0.9915 ). Images (a–e) depict the inoculation at time 0 days until the lag phase, at t = 6 days. Below, images (a’–e’) depict the digital contrast mask. The area at t = 0 days was subtracted to compute only the growth area.
Figure 1. Digital image analysis for mycelium growth in potato dextrose agar (PDA) medium, kinetic characterization (fitted to y   =   a e b t + c model, R 2   =   0.9915 ). Images (a–e) depict the inoculation at time 0 days until the lag phase, at t = 6 days. Below, images (a’–e’) depict the digital contrast mask. The area at t = 0 days was subtracted to compute only the growth area.
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Figure 2. Digital photographs of (a) Agave bagasse (substrate), (b) Ganoderma lucidum, (c–f) biocomposite (Agave bagasse/corn stover + G. lucidum). Additionally, thermal exposure and response of mycelium-lignocellulose biocomposites obtained with an optical microscope at 5× g magnification: (g) untreated control showing intact hyphal networks (0 min.); (h) moderately heated (2–3 min.) specimen exhibiting hyphal shrinkage; (i) charred structure (4–5 min.).
Figure 2. Digital photographs of (a) Agave bagasse (substrate), (b) Ganoderma lucidum, (c–f) biocomposite (Agave bagasse/corn stover + G. lucidum). Additionally, thermal exposure and response of mycelium-lignocellulose biocomposites obtained with an optical microscope at 5× g magnification: (g) untreated control showing intact hyphal networks (0 min.); (h) moderately heated (2–3 min.) specimen exhibiting hyphal shrinkage; (i) charred structure (4–5 min.).
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Figure 3. Technical diagram of the custom-made mold for the manufacturing of mycelium-based Tequila bottle packaging: (a) top cover, and (b) base. All units in cm.
Figure 3. Technical diagram of the custom-made mold for the manufacturing of mycelium-based Tequila bottle packaging: (a) top cover, and (b) base. All units in cm.
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Figure 4. 3D render of the mold designed for the manufacturing of mycelium-based Tequila bottle packaging. (a–c) represent the packaging assembly sequence.
Figure 4. 3D render of the mold designed for the manufacturing of mycelium-based Tequila bottle packaging. (a–c) represent the packaging assembly sequence.
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Figure 5. Mycelium-based packaging components after colonization and drying. (a) Top cover, (b) molded base, (c,d) top and side view of packaging or bottle display stand.
Figure 5. Mycelium-based packaging components after colonization and drying. (a) Top cover, (b) molded base, (c,d) top and side view of packaging or bottle display stand.
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Figure 6. Final prototype of the mycelium-based packaging assembled with the top (lid) and bottom (base) components. (a) front right, (b) top, and (c) side view of the prototype.
Figure 6. Final prototype of the mycelium-based packaging assembled with the top (lid) and bottom (base) components. (a) front right, (b) top, and (c) side view of the prototype.
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Figure 7. Final prototype of the mycelium-based packaging showing the interior design and the display of the Tequila bottle. Views: (a) front left, (b) front, and (c) front right of the prototype.
Figure 7. Final prototype of the mycelium-based packaging showing the interior design and the display of the Tequila bottle. Views: (a) front left, (b) front, and (c) front right of the prototype.
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Figure 8. Drop test simulation of the developed mycellium-based packaging, in SolidWorks: (a) total displacement distribution (URES), (b) equivalent strain (ESTRN), and (c) von Mises stress during impact from 762 mm, in accordance with standard drop test protocols. Note. Plot step: 25; time: 6.22269 microseconds; deformation scale: 1.
Figure 8. Drop test simulation of the developed mycellium-based packaging, in SolidWorks: (a) total displacement distribution (URES), (b) equivalent strain (ESTRN), and (c) von Mises stress during impact from 762 mm, in accordance with standard drop test protocols. Note. Plot step: 25; time: 6.22269 microseconds; deformation scale: 1.
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Figure 9. Environmental impact in terms of (a) fossil fuel scarcity (kg oil eq.), (b) global warming (kg CO2 eq), and (c) water consumption (m3).
Figure 9. Environmental impact in terms of (a) fossil fuel scarcity (kg oil eq.), (b) global warming (kg CO2 eq), and (c) water consumption (m3).
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de Anda-Rodríguez, F.A.; Corona-Ramírez, M.R.; Patiño-Arévalo, C.D.; Zárate-Navarro, M.A.; Zárate-Guzmán, A.I.; Romero-Cano, L.A. Mycomaterials from Agave Bagasse: A Valorization Strategy for Sustainable Tequila Packaging. Fermentation 2025, 11, 500. https://doi.org/10.3390/fermentation11090500

AMA Style

de Anda-Rodríguez FA, Corona-Ramírez MR, Patiño-Arévalo CD, Zárate-Navarro MA, Zárate-Guzmán AI, Romero-Cano LA. Mycomaterials from Agave Bagasse: A Valorization Strategy for Sustainable Tequila Packaging. Fermentation. 2025; 11(9):500. https://doi.org/10.3390/fermentation11090500

Chicago/Turabian Style

de Anda-Rodríguez, Flavio A., Mariana R. Corona-Ramírez, Carlos D. Patiño-Arévalo, Marco A. Zárate-Navarro, Ana I. Zárate-Guzmán, and Luis A. Romero-Cano. 2025. "Mycomaterials from Agave Bagasse: A Valorization Strategy for Sustainable Tequila Packaging" Fermentation 11, no. 9: 500. https://doi.org/10.3390/fermentation11090500

APA Style

de Anda-Rodríguez, F. A., Corona-Ramírez, M. R., Patiño-Arévalo, C. D., Zárate-Navarro, M. A., Zárate-Guzmán, A. I., & Romero-Cano, L. A. (2025). Mycomaterials from Agave Bagasse: A Valorization Strategy for Sustainable Tequila Packaging. Fermentation, 11(9), 500. https://doi.org/10.3390/fermentation11090500

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