This study reports the development and characterization of chitosan-based nanoparticles (CTS-NPs) for the co-delivery of clotrimazole (CLZ) and bioactive compounds derived from Enteromorpha prolifera (algal extract, AE) to enhance antifungal efficacy against drug-resistant Clavispora lusitaniae. The dual-loaded nanoparticles (CTS-CLZ-AE-NPs) exhibited an encapsulation efficiency of 65 % for CLZ and 81 % for AE, with respective loading capacities of 13 % and 20.25 %. Structural and compositional analyses using FTIR and SEM confirmed successful encapsulation and notable morphological changes. GC–MS analysis identified key antimicrobial constituents in AE, including phenolics, terpenoids, and fatty acids. In vitro, antifungal assays demonstrated that CTS-CLZ-AE-NPs achieved the highest fungicidal activity, with a minimum inhibitory concentration (MIC) of 2 mg/mL and an 85.99 % reduction in biofilm formation at 4 × MIC. SEM analysis further revealed significant morphological damage in treated fungal cells. These findings highlight the synergistic potential of CLZ and AE in a nanochitosan platform and underscore its promise as an effective alternative strategy against antifungal-resistant pathogens. Further in vivo evaluation is warranted to establish clinical applicability.