Ganoderma lucidum has been used in oriental medicine for its contribution to vitality and longevity. None of them report about targeted antitumor activity against adenocarcinoma cells, or antioxidant and antiacetylcholinesterase activity for potential application in treatment of Alzheimer's and other neurodegenerative diseases. To date, there are a few studies available concerning supercritical carbon dioxide extraction of biologically active compounds from G. lucidum fruiting body. In our study, two stage extractions of biologically active compounds from G. lucidum were performed. First, supercritical carbon dioxide was used as extraction solvent. Next, the same material was used for hot water isolation of biologically active polysaccharides. Cytotoxicity, antioxidant and antiacetylcholinesterase activity were tested for all obtained extracts. Additionally, the effect of extraction process conditions on the biological activity of extracts was assessed. (C) 2014 Elsevier B.V. All rights reserved.