We investigated the chemical and medicinal properties of methanolic and acetonic extracts of <i>Armillaria ostoyae</i> and the presence of heavy metals in its dry basidiocarps. The chemical content of extracts was analyzed with the HPLC-DAD-MS/MS method. According to our results, the most abundant mineral was potassium; the most abundant organic acid was malic acid; the most abundant carbohydrate was fructose, and the most abundant polyphenol was chlorogenic acid. The antimicrobial potential was evaluated using the microdilution assay, and the results ranged from 0.62 to 20 mg/mL. Antioxidant potential was studied by DPPH [half-maximal inhibitory concentration (IC<sub>50</sub>) of the methanolic extract was 619.67 μg/mL and of the acetonic extract was 533.65 μg/mL] and reducing power assays (the results ranged from 0.025 to 0.078 μg/mL). Total phenolic content was presented as gallic acid equivalent (methanolic extract, 6.12 mg GAE/g; acetonic extract, 3.99 mg GAE/g). The antidiabetic potential was explored by applying the α-amylase (the results ranged from 39.62 to 44.33%) and α-glucosidase assays (the results were in the range of 0.27-2.51%). The neuroprotective activity was asserted by the acetylcholinesterase inhibition assay (the results were in the range of 3.06-6.09%). The cytotoxic potential was investigated using the microtetrazolium assay, and the IC<sub>50</sub> values ranged from 221.96 to > 400 μg/mL. Heavy metal content of the dry basidiocarps was evaluated using the AAS method and iron was the most abundant metal. <i>A. ostoyae</i> is a conditionally edible mushroom, which was not studied thoroughly before, thus this research will provide valuable knowledge about this species.