This section presents the methodology and results of the development and characterization of alginate-based conductive hydrogels incorporating different carbonaceous materials. The aim was to evaluate their suitability for printing and sensing applications through a comparative analysis of drying behavior, electrical conductivity, rheological response, and extrusion performance.
The selected carbon additives, PCO1000C (activated carbon), Vulcan V3 (carbon black), and natural graphite, were chosen for their high conductivity, surface area, and chemical stability [
21]. Sodium alginate was used as the hydrogel matrix due to its biocompatibility and ionic crosslinking ability, which support structural integrity and tunable mechanical properties [
22]. Their combination enables the formulation of printable and stable hydrogels with enhanced functionality for sensor applications [
23].
2.1. Electrical Characterization of Starting Materials
Electrical characterization allowed understanding the behavior of these materials when exposed to electric fields and evaluating their potential in electrochemical applications. In the specific case of alginate and carbonaceous materials, electrical characterization provided valuable information on their electrical conductivity.
Table 1 shows the electrical conductivity measurements at room temperature of the studied carbonaceous elements and sodium alginate, used in their original form and without undergoing any hydration process and at constant pressure.
The table above shows that the samples had the following decreasing order of electrical conductivity: graphite ˃ Vulcan V3 > PCO1000C ˃ alginate. Graphite, as expected, exhibited the highest electrical conductivity (6.12 (Ω·cm)
−1). This high conductivity is attributed to the unique arrangement of carbon atoms in its crystalline structure. All other materials showed a significant reduction in electrical conductivity. Specifically, Vulcan V3 exhibited an electrical conductivity of 2.21 (Ω·cm)
−1. This reduction in conductivity compared with graphite may be due to its porous structure and the presence of impurities [
24]. Overall,
Table 1 reveals a wide range of electrical conductivities among the materials studied.
PCO1000C and sodium alginate offered the lowest conductivities. Understanding these properties is crucial for the proper selection of materials for various applications. Carbonaceous materials such as activated carbon and graphite are used in supercapacitor electrodes because of their high surface area, excellent electrical conductivity, and stability [
25].
This difference in intrinsic conductivity was expected to influence the overall electrical behavior of the composite hydrogels, especially under mechanical stress or postdrying conditions.
2.2. Results of Hydrogel Preparation and Drying Treatments
The integration of carbonaceous materials into alginate-based hydrogels aims to enhance their electrical conductivity, mechanical integrity, and suitability for 3D printing. Selecting the appropriate formulation parameters, particularly alginate concentration and moisture content, is essential for achieving reproducible and functional constructs suitable for sensor applications.
Alginate concentrations of 1%, 1.5%, and 2% (
w/
v) were tested in combination with 8% (
w/
w) carbonaceous additives, including PCO1000C (activated carbon), Vulcan V3 (carbon black), and natural graphite. The hydrogel preparation process is illustrated in
Figure 1, and the nomenclature used for the different formulations according to their drying treatment is summarized in
Table 2.
Increasing alginate concentration typically results in a denser polymeric network, which may reduce porosity and ionic transport but improves mechanical strength and print fidelity. The 2% alginate formulation demonstrated the best balance between printability and structural robustness. These findings are in line with previous studies that report alginate concentrations ranging from 1–5% for biomedical uses [
22], 0.1–1% for food industry applications [
26], and up to 10% for industrial-grade hydrogels [
27].
Moreover, ionic crosslinking also contributes to the final stiffness of the hydrogels. Calcium ions, for example, can create rigid structures even at relatively low alginate concentrations (1–3%), while other divalent cations such as magnesium may require higher polymer contents to achieve similar mechanical properties [
28].
In parallel, drying strategies were assessed as a critical factor influencing the flow behavior and functional performance of the hydrogels. Three conditions were tested: oven drying at 50 °C, drying in open containers at room temperature, and drying in containers with perforated closures. Proper moisture control proved essential for modulating viscosity, mechanical resistance, and the formation of conductive pathways, especially relevant for applications involving extrusion-based printing and sensor response.
By systematically evaluating the interaction between alginate content, carbon phase, and drying conditions, a rational formulation was identified that supports extrusion, maintains mechanical stability, and enhances conductivity, as further discussed in the following sections.
Effect of Drying Conditions on Hydrogel Properties
The drying method significantly influenced the final properties of the carbon-loaded hydrogels, particularly with respect to moisture retention, printability, and structural stability. The percentage of water loss measured under different drying conditions is shown in
Table 3.
Oven drying at 50 °C was the most effective strategy, yielding the highest water loss in all formulations. This condition promotes uniform and accelerated moisture removal, which favors the formation of a denser matrix and facilitates conductive pathway restoration. In contrast, drying in perforated containers resulted in lower water loss, likely due to slower evaporation and partial moisture retention, while open-container drying showed intermediate behavior.
Among the different formulations, Vulcan V3-based hydrogels exhibited the highest dehydration range across all conditions (86.37–92.05%), indicating a network more prone to moisture loss. Graphite-based hydrogels showed greater stability (84.00–85.90%), suggesting lower permeability and more consistent water retention. PCO1000C formulations displayed the greatest variability (85.68–94.90%), reflecting a strong dependence on the drying condition, likely due to its porous microstructure and surface chemistry.
These differences confirm that both the type of carbon additive and the drying method play key roles in defining the hydrogel’s functional characteristics. As reported in previous studies [
29,
30], drying control is essential for balancing print fidelity, mechanical strength, and electrical performance in conductive hydrogel systems.
2.3. Results of the Characterization of Hydrogel–Carbon Composites
The characterization of a hydrogel mixture with carbonaceous materials is a fundamental step in understanding its structure, properties and possible applications. This characterization allowed evaluating the effectiveness of the incorporation of carbonaceous materials in the hydrogel, as well as determining the final properties of the composite material.
2.3.1. Results of Electrical Conductivity Tests
Following the preparation of the hydrogel formulations, an initial qualitative screening of electrical conductivity was performed in the liquid state to verify their potential suitability for sensor applications. Each hydrogel sample was placed inside a cuvette integrated into a custom-designed testing device (
Figure 2), which applied a low-voltage electric potential across the sample. The successful activation of an LED indicated that the hydrogel exhibited sufficient conductivity to complete the circuit.
All tested hydrogel formulations, regardless of the carbonaceous additive used, successfully conducted electricity in their hydrated state, demonstrating baseline electrical functionality under standard conditions.
To evaluate their performance under realistic postprinting conditions, the electrical conductivity of each hydrogel was further assessed after a 48 h drying period, simulating the state of the material following extrusion and initial setting. Conductivity was measured under incremental mechanical pressure, reflecting practical use conditions in pressure-sensitive or contact-based sensors.
The results showed that conductivity improved notably with increased drying intensity, especially after oven drying at 50 °C, where the electrical performance approached that of the pristine carbonaceous fillers.
This behavior reflects the dual role of water in the hydrogel matrix. On one hand, it facilitates ion transport by solvating ionic species and contributes to the flexibility of the polymer network; on the other, it impedes efficient electron transfer between conductive particles by increasing the interparticle distance and preventing the formation of continuous percolation pathways. As previously reported by Su et al. (2024) [
31], hydrated hydrogels rely primarily on ionic conductivity, and only upon dehydration does the electron-conductive mechanism emerge, driven by particle contact and matrix contraction.
Part of the applied stress is dissipated through polymer chain relaxation and water-mediated flow, while in drier hydrogels, the matrix exhibits more elastic behavior, directly transmitting compressive forces to the conductive phase without significant polymer rearrangement. This elastic response has been previously reported in dual-mode conductive hydrogels, where water loss leads to a solid-like state that promotes stress transfer through the polymer network rather than deformation [
31].
Conversely, in drier samples, the polymer matrix becomes mechanically stiffer, reducing internal reorganization and allowing pressure to compact the carbon particles more effectively. This leads to decreased interparticle distances and increased formation of direct conductive bridges, promoting enhanced electron hopping and tunneling mechanisms. The observed linear conductivity gain with pressure in these samples reflected dominant percolation by mechanical densification rather than ionic contribution.
Therefore, water content critically modulates the interplay between polymer mobility and particle contact, determining whether conductivity arises predominantly from ionic mobility, matrix reconfiguration, or particle compaction under load.
In these formulations, the absence of residual water minimized matrix reorganization, and mechanical densification became the dominant mechanism enhancing conductivity.
As shown in
Figure 3,
Figure 4 and
Figure 5, a notable improvement in conductivity was observed in all formulations subjected to more intense drying, particularly oven drying at 50 °C. In these cases, the electrical conductivity values approached those of the original carbon materials, suggesting effective restoration of conductive pathways. In contrast, the samples dried in perforated or open vessels retained more water and exhibited lower conductivity, likely because of incomplete moisture removal and less favorable particle–particle interactions.
This pressure-dependent conductivity suggests a possible application in pressure- or strain-sensitive devices, where compaction of the material translates into measurable electrical changes.
In summary, water content plays a dual role in determining electrical conductivity: it affects both the packing density of carbonaceous particles and the structural flexibility of the polymer matrix under pressure [
32]. Proper drying control is therefore essential to optimize electrical performance in conductive hydrogels designed for 3D-printed sensor applications.
2.3.2. Results of Rheological Characterization
Preliminary rheological tests focused on stiffness, flow behavior, and viscosity revealed that alginate concentrations significantly influenced the extrudability and mechanical properties of the hydrogels. Formulations with alginate concentrations either below or above 2% (w/v) were found to be unsuitable for extrusion-based printing, as they exhibited either insufficient structural integrity or excessive viscosity that hindered flow through standard nozzles.
Based on these observations, the working concentration was set at 2% alginate, which provided a suitable balance between mechanical stability and flow behavior for 3D printing applications. The final compositions of the hydrogels containing different carbonaceous materials are summarized in
Table 4.
The hydrogel formulations were prepared following the indicated procedure, and their rheological behavior was evaluated by measuring the shear viscosity as a function of shear rate at 50 °C (see
Figure 6).
The hydrogel composed solely of alginate exhibited the lowest initial viscosity. Its viscosity decreased only slightly with increasing shear rate, in line with prior reports in the literature, and it showed the lowest initial viscosity [
33,
34].
The results clearly indicated pseudoplastic (shear-thinning) behavior for all formulations, characterized by a decrease in viscosity with increasing shear rate [
35]. This behavior is typical of polymer-based hydrogels and is desirable for extrusion applications, as it facilitates flow under pressure while maintaining shape fidelity after deposition [
36,
37,
38].
Among the tested samples, the formulations containing PCO1000C and Vulcan V3 exhibited higher initial viscosities, which may lead to increased flow resistance through the nozzle. It is important to note that extrusion performance is influenced not only by viscosity but by applied pressure and nozzle geometry. These factors must be considered when selecting materials for bioprinting processes.
In comparison with previous studies on alginate-based hydrogels reinforced with conductive additives, our formulations exhibited similar pseudoplastic behavior, yet with significant differences in initial viscosity and flow resistance depending on the nature of the incorporated carbonaceous additive. For instance, Serafin et al. [
15] reported formulations containing carbon nanofibers (CNFs) with very high initial viscosities, which could lead to challenges during the printing process, including filament breakage and low geometric fidelity. In contrast, the formulations shown in
Figure 6 maintained a pseudoplastic rheological profile without reaching such high viscosity levels. Notably, the graphite-based hydrogel exhibited the lowest initial viscosity among the tested samples, resulting in a lower extrusion pressure requirement and greater stability during the printing process. Moreover, unlike other approaches that require complex chemical modifications or copolymerization steps, such as those involving PVA/PANI hydrogels [
13], the formulations presented in
Table 4 were prepared by simple physical blending of alginate and carbon-based materials, which facilitates their fabrication and may enhance their biocompatibility for biomedical applications.
2.3.3. Determining Optimum Printing Pressure
Following the rheological analysis and the evaluation of available bioprinting nozzle geometries [
39,
40], tests were performed to determine the optimal extrusion pressure required for each hydrogel formulation [
41].
The geometry of the nozzle plays a fundamental role in extrusion-based printing, as it affects not only the quality and resolution of the printed structure but the flow dynamics of the material. The nozzle influences material distribution, reduces the risk of clogging, improves interlayer adhesion, and determines the surface finish of the printed construct. To identify the most suitable configuration, various nozzle types were tested, including standard cylindrical, angled orifice, oval, and rectangular nozzles.
Among these, the standard G20 rectangular nozzle demonstrated the best performance when used with the alginate–graphite hydrogel. This configuration provided a good balance between resolution and extrusion force, while maintaining print stability. In contrast, the other hydrogels (containing PCO1000C or Vulcan V3) were not extrudable under standard conditions because of excessive viscosity and high resistance to flow.
Once the nozzle type was selected, the extrusion force was quantified for each formulation using a custom-built testing system (
Figure 7). The hydrogel was loaded into a syringe connected to the selected nozzle, and a controlled displacement force was applied by a load cell. The force required for the hydrogel to begin flowing was recorded and is presented in
Table 5.
The relationship between applied force and extrusion behavior was analyzed by identifying the inflection point of the force–displacement curve, which corresponds to the minimum pressure needed to initiate continuous flow without overextrusion or deformation [
42,
43]. For the alginate–graphite hydrogel, this threshold was reached at approximately 0.008 kN, indicating that it possessed the necessary flow characteristics for precise and stable printing.
In contrast, the PCO1000C and Vulcan V3 formulations required forces close to 0.98 kN and 0.91 kN, respectively, values too high for practical bioprinting under standard conditions.
These results confirmed that only the graphite-based hydrogel formulation exhibited adequate extrudability under moderate pressure, making it the most suitable candidate for 3D-printed conductive hydrogel applications.
2.3.4. Determination of 3D Printing Capability
Based on the previous extrusion and rheological analyses, the alginate–graphite hydrogel formulation was selected for experimental 3D printing trials because of its favorable printability, moderate viscosity, and stable flow behavior.
The ability to accurately print hydrogel-based structures with defined geometry is essential for applications in flexible sensors and other soft electronic platforms. To evaluate this capability, test cylindrical structures were fabricated using a modified 3D press for hydrogel extrusion, operating with the parameters listed in
Table 6.
The hydrogel formulations were extruded prior to CaCl2 crosslinking. Because of the low aspect ratio and planar geometry of the printed discs, structural collapse was not a concern, and the native viscosity of the ink was sufficient to preserve the shape during deposition. Final stabilization was achieved by CaCl2 application after printing.
As shown in
Figure 8A, the hydrogel was successfully extruded and printed into self-supporting cylindrical structures, indicating good shape fidelity and adequate material cohesion. To further evaluate the mechanical performance of the printed hydrogels, compressive tests were carried out on the printed cylinders before and after ionic crosslinking with calcium chloride.
Compression tests were conducted at 28%, 57%, and 75% strain, and both the force required for deformation and the compressive strength after crosslinking were recorded (
Table 7). The experimental setup is illustrated in
Figure 8B.
These results demonstrate that the printed alginate–graphite hydrogel structures exhibited a clear increase in mechanical resistance with higher compressive strain and that ionic crosslinking significantly enhanced their load-bearing capacity. After crosslinking, the resistance to compression increased from 0.056 N at 28% strain to 0.99 N at 75% strain (an almost 18-fold improvement), highlighting the hydrogel’s ability to maintain structural cohesion under mechanical stress.
In practical terms, even a lightweight printed hydrogel structure (0.35 g) was able to withstand more than 150 times its own weight (0.056 N) at just 28% deformation. This indicates that the hydrogel is not only printable and self-supporting but mechanically robust enough to support additional printed layers, which is essential for multilayer fabrication in 3D printing.
The combination of dimensional fidelity, compressive strength, and postprinting mechanical enhancement via crosslinking confirms the potential of this formulation for wearable and implantable soft sensors, particularly in applications requiring tactile response, pressure sensitivity, or structural resilience in dynamic environments.
2.3.5. Results of the Cytotoxicity Assay
Cell viability in the presence of graphite extracts at different concentrations, as well as after direct contact with printed alginate–graphite hydrogel discs, was assessed by MTS assay.
Figure 9 shows that exposure of cells to graphite extract showed a concentration-dependent trend, with minimum cell viability observed with 100% extract and a gradual increase in viability with decreasing extract ratio. In particular, mean viability values increased progressively in the groups treated with 75%, 50%, and 25% extract, reaching levels comparable to the control at 50% dilution. This trend suggests a negative dose–response relationship between extract concentration and cell survival, coinciding with results obtained in previous studies [
44]. In addition, other studies have demonstrated the toxicity of compounds such as nanographite and nanoparticles [
45], which, because of its size (10–100 nm), could pass through the 0.22 um filter, exerting a cytotoxic effect on the cells in contact with the extract [
46].
At 50% dilution and above, no significant differences were observed with respect to the negative control, indicating that the toxicity induced by free graphite was substantially reduced below this concentration.
In contrast, direct contact with the alginate–graphite hydrogel discs did not induce significant alterations in cell viability compared with the control, which was around 100%. This behavior can be attributed to the high adsorption capacity of the graphite powder, which can sequester components present in the culture medium, such as phenol red [
47], vitamins [
48], amino acids [
49] o factors present in fetal bovine serum [
50].
Furthermore, studies have also indicated that graphite was able to adsorb both synthetic and organic dyes [
51], as well as industrial effluents containing compounds such as phenol [
52].
The loss of color of the medium observed after incubation with graphite reinforces this hypothesis and suggests an alteration in the composition of the culture medium that compromises the chemical balance of the cellular environment, which could have contributed to the decrease in viability.
In contrast, no change in the staining of the culture medium and no decrease in cell viability were observed when the alginate–graphite discs were placed in contact with the cell culture. Other studies have shown that coating carbonaceous compounds with sodium alginate reduced toxicity and improved compatibility [
53], This may suggest that when graphite is immobilized within an alginate matrix, its interaction with the medium is more limited, and therefore, its cytotoxic effects are less severe.
This finding is in line with previous work highlighting the importance of the immobilization of nanomaterials or carbonaceous particles in polymeric matrices to preserve biocompatibility [
54,
55]. Thus, the immobilization of graphite in a hydrogel network could act as a physical and chemical barrier, preventing both direct contact with the cells and the alteration of the components of the medium.
These results reinforce the potential of alginate–graphite hydrogel as a biocompatible support for sensor applications and suggest that the presentation form of graphite is a critical factor for its biosafety.