The current study evaluated the antioxidant, α-amylase inhibitory and anti-Staphylococcus aureus activities of the fruiting bodies of Ganoderma lucidum. Methanolic extract of G. lucidum (MEGL) was investigated for its in vitro antioxidant activity using 1,1-diphenyl 2-picrylhydrazyl (DPPH) assay and EC 50 value was determined. α-Amylase inhibitory and anti-S. aureus activities were carried out by conventional methods. The mushroom extract showed maximum free radical scavenging activity at a dose of 500 µg/mL with EC 50 value of 22.1399 µg/mL. EC 50 value for the inhibition of α-amylase activity was found 22.6011 µg/mL. G. lucidum extracts exhibited a concentration-dependent Anti-S. aureus activity. The vast differences of this inhibition between lower and higher doses were assessed significantly (p < 0.05). This study shows that the fruiting body of G. lucidum has moderate antioxidant, α-amylase inhibitory, and anti-S. aureus activities. Thus this mushroom may be the alternative plant source for the antioxidant, anti-diabetic and antibacterial agents and further in vivo studies and isolation of bioactive compounds are required.