| Summary: | There is an urgent need for new substances to overcome current challenges in the health sciences. Marine fungi are known producers of numerous compounds, but the manipulation of growth conditions for optimal compound production can be laborious and time-consuming. In Portugal, despite its very long coastline, there are only a few studies on marine fungi. From a collection of Portuguese marine fungi, we screened for antimicrobial, antioxidant, enzymatic, and cytotoxic activities. Mycelia aqueous extracts, obtained by high pressure-assisted extraction, and methanolic extracts of culture media showed high antioxidant, antimicrobial, and cytotoxic activities. The mycelium extracts of <i>Cladosporium rubrum</i> showed higher antioxidant potential compared to extracts from other fungi. Mycelia and culture media extracts of <i>Aspergillus affinis</i> and <i>Penicillium lusitanum</i> inhibited the growth of <i>Staphylococcus aureus</i>, <i>Kocuria rhizophila</i>, <i>Enterococcus faecalis</i>, <i>Escherichia coli</i>, <i>Klebsiella pneumoniae</i>, and <i>Pseudomonas aeruginosa</i>, including multiresistant strains. <i>Penicillium lusitanum</i> and <i>Trichoderma aestuarinum</i> inhibited the growth of clinical strains of <i>Candida albicans</i>, <i>C. glabrata</i>, <i>C. parapsilosis</i>, and <i>C. tropicalis</i>. All extracts from culture media were cytotoxic to Vero cells. Sea salt induced alterations in the mycelium’s chemical composition, leading to different activity profiles.
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