The aim of this work was to investigate total phenolic and flavonoid content, antioxidant properties and to assess antimicrobial activities of three different extracts from Polycephalomyces nipponicus TBRC 6537 mycelia. The 50% (v/v) aqueous methanol extract was found to have the highest total phenolic content, while methanol extract exhibited the highest total flavonoid content. Water and methanol extracts showed the strongest DPPH radical scavenging activity. In addition, water and 50% (v/v) aqueous methanol extracts had stronger ABTS radical scavenging activity than that of methanol extract. The antimicrobial activity of each extract was quantified by determining minimum inhibitory concentrations (MICs), minimum bactericidal concentrations (MBCs) and minimum fungicidal concentrations (MFC) against five human pathogenic bacteria and one human pathogenic fungus. The water extract was found to have the strongest activity against Escherichia coli, Staphylococcus epidermidis, Bacillus cereus and Candida albicans. Furthermore, aqueous methanol extract exhibited strongest activity against Staphylococcus aureus with the lowest MIC and MBC. The methanol extract had moderate antimicrobial activity against Pseudomonas aeruginosa, whereas the aqueous methanol and water extracts did not inhibit growth. This study has revealed that P. nipponicus TBRC 6537 mycelia extracts might be a useful source of natural antioxidant and antimicrobial compounds for the pharmaceutical industry.