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As primary producers in aquatic ecosystems, microalgae function not only as a natural source of nourishment for several economically important aquatic species but also as reservoirs of bioactive molecules. Microalgae can secrete exosome-like nanoparticles that transport functional biomolecules, such as proteins and nucleic acids, into the extracellular milieu, thereby mediating intercellular signaling and eliciting ecological or biomedical responses. Although plant-derived exosome-like nanoparticles have attracted attention for their utility in drug delivery and dermatology, the functional properties of microalgae-derived nanoparticles-particularly from species extensively applied in aquaculture-remain inadequately characterized. In this study, exosome-like nanovesicles were isolated from Nannochloropsis oculata (N-ELNs), a microalgal species widely used in aquaculture, and their skin-whitening potential was evaluated using the B16-F10 mouse melanoma cell model. The highest N-ELN yield was observed during the adaptation, exponential, and stationary growth phases. Uptake analyses confirmed the efficient internalization of N-ELNs by B16-F10 cells. Cell counting kit-8 assays indicated that N-ELNs exhibited no cytotoxic effects on melanoma cells or normal human dermal fibroblasts (HFF-1). Scratch wound healing assays revealed that N-ELNs exerted no significant effect on cellular migration. In B16-F10 cells, N-ELNs suppressed tyrosinase activity by downregulating Mitf and its downstream genes Tyr and Tyrp1, resulting in a substantial reduction in melanin synthesis (p < 0.05). The inhibitory effects of N-ELNs on melanin production, tyrosinase activity, and gene expression of Tyr, Tyrp1, and Mitf were comparable to those of the positive control, arbutin. Collectively, these findings suggest that N. oculata exhibits promising skin-whitening properties, providing a novel perspective for clinical applications and supporting the high-value utilization of the microalgae aquaculture industry.