To further knowledge of the biological activity of native neotropical fungal species, this study aimed to determine the chemical composition and microbiological activity of <i>Hornodermoporus martius</i>. Ethanol, hexane, diethyl ether, and ethyl acetate fractions and the water residue were analyzed and resulted in a total phenolic compound content between 13 and 63 mg of gallic acid equivalents per gram of crude extract. The total antioxidants ranged between 3 and 19 mg of ascorbic acid equivalents per gram of crude extract, and the percentage of antioxidant activity was determined to be between 6 and 25%. A preliminary profile of compounds is provided for the first time for the species; the results from the nonpolar fraction showcased the presence of saturated and unsaturated acids, fatty alcohol, sterols, and cis-vaccenic acid. Our findings also revealed antimicrobial properties from compounds within the hexane and diethyl ether fractions at concentrations of 1 mg mL<sup>-1</sup>, which inhibited the growth of certain gram-positive and gram-negative bacteria. For the first time in academic literature, our work analyzed and documented the chemical characteristics and microbial properties of <i>H. martius</i>, suggesting potential for medicinal applications.