© 2021, Pleiades Publishing, Ltd.Abstract: The present study was designed to determine the antioxidant potential of extracellular and intracellular metabolites of 130 macro/microfungi isolated from different parts of Turkey. To decide precisely and accurately, firstly, the sample ratio in a 2,2-diphenyl-1-picrylhydrazyl (DPPH) reaction mixture, culture medium, cultivation mode and extraction method of antioxidants from biomass were optimized. After optimization studies, a screening study was performed under the conditions of; potato malt peptone medium; ten days at 25°C under static cultivation mode; sample : DPPH ratio as 0.1 : 2.9 (3.33%); and microwave application for extraction of antioxidants from biomass. In the screening stage, culture liquid extracts of the vast majority of the isolates (80.77%) exhibited higher scavenging activity value than mycelial biomass extracts. Among 130 fungal isolates, six isolates (Epicoccum nigrum, Suillus collitinus, Hypomyces chrysospermus, Omphalotus olearius, Inonotus hispidus, and Pleurotus eryngii) and one isolate (Omphalotus olearius) were selected for further investigation of their extracellular and intracellular antioxidant metabolites, respectively. The selected fungal isolates had scavenging activities ranging from 50.35 to 100.00% and had phenolics ranging from 0.80 to 4.62 gallic acid equivalent mg/mL. After time-dependent DPPH radical-scavenging activity and extraction yield studies, the selected isolates were evaluated for their antioxidant activity using six different methods. As a result, Hypomyces chrysospermus was determined as the best isolate, according to high capabilities in all assay methods, except for lipid peroxidation, which was moderate. Mycelial biomass extract of the Omphalotus olearius gave impressive results for ABTS radical-scavenging activity, β-carotene bleaching and inhibition of lipid peroxidation.