Background: Prosthetic candidiasis remains a significant clinical challenge, particularly due to the ability of Candida species to form resilient biofilms on dental prostheses, which limits the efficacy of conventional antifungal treatments. In this context, developing strategies to prevent or reduce biofilm formation is essential. Objectives This study investigates the antifungal and antibiofilm potential of a hydrogel formulation incorporating aminochalcone AM-35 as a candidate for the prevention and treatment of prosthetic candidiasis. Methods: To achieve this, experiments were conducted to determine the minimum inhibitory concentration (MIC) of aminochalcone AM-35 against Candida albicans and Candida tropicalis strains. AM-35 was incorporated into a hydrogel, which was subsequently tested on biofilms formed by these yeast species, both individually and in combination. The experimental disks were sterilized and incubated with C. albicans, C. tropicalis, and a mixture of both strains for 120 h to allow biofilm maturation. After contamination, the samples were divided into four experimental groups: Group 1: Hydrogel; Group 2: Hydrogel+AM-35; Group 3: Sodium hypochlorite (positive control); and Group 4: No treatment. The samples were then subjected to a sonication process to disaggregate the cells, which were then cultured on plates for colony-forming unit (CFU/mL) counts. The hydrogel’s toxicity was evaluated in vivo using the Galleria mellonella model. Results: The hydrogel formulation demonstrated significant antimicrobial activity, with an MIC of 7.8 μg/mL for C. albicans and 3.9 μg/mL for C. tropicalis. Treatment with the hydrogel at a concentration of 39 μg/mL resulted in a significant reduction in the formation and viability of mixed-species biofilms (p < 0.05). Additionally, the results indicated robust activity against C. albicans and C. tropicalis without presenting toxicity in the Galleria mellonella model. In conclusion, the hydrogel formulation exhibited effective antibiofilm activity, significantly reducing the microbial load. Conclusions: These findings open new possibilities for the development of alternative treatments for prosthetic candidiasis. The research suggests that the use of chalcone-based compounds may represent a promising approach in combating fungal infections in dentistry.
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