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Natural adjuvants have recently garnered interest in the field of vaccinology as their immunostimulatory effects. In this study, we aimed to investigate the potential use of Peyssonnelia caulifera (PC), a marine alga, as a natural adjuvant for an inactivated split A/Puerto Rico/8/1934 H1N1 influenza vaccine (sPR8) in a murine model. We administered PC-adjuvanted vaccines to a murine model via intramuscular prime and boost vaccinations, and subsequently analyzed the induced immunological responses, particularly the production of antigen-specific IgG1 and IgG2a antibodies, memory T and B cell responses, and the protective efficacy against a lethal viral infection. PC extract significantly bolstered the vaccine efficacy, demonstrating balanced Th1/Th2 responses, increased memory T and B cell activities, and improved protection against viral infection. Notably, within 3 days post-vaccination, the PC adjuvant stimulated activation markers on dendritic cells (DCs) and macrophages at the inguinal lymph nodes (ILN), emphasizing its immunostimulatory capabilities. Furthermore, the safety profile of PC was confirmed, showing minimal local inflammation and no significant adverse effects post-vaccination. These findings contribute to our understanding of the immunomodulatory properties of natural adjuvants and suggest the promising roles of natural adjuvants in the development of more effective vaccines for infectious diseases.Supplementary InformationThe online version contains supplementary material available at 10.1038/s41598-024-76736-9.