The chemical composition and biological activity of three parts (rind, flesh and seeds) of pumpkin fruits (Cucurbita pepo L.) cultivated in Egypt were studied. Chemical analysis of fibre, protein, β-carotene, carbohydrates, minerals and fatty acids present in the rind, flesh, seeds and defatted seeds meal was conducted. Chemical, GC-MS and biological assays of organic extracts of the main fruit parts, rind and flesh established their unique constituents. Chromatographic purification of the extracts afforded triglyceride fatty acid mixture (1), tetrahydro-thiophene (2), linoleic acid (3), calotropoleanly ester (4), cholesterol (5) and 13(18)-oleanen-3-ol (6). GC-MS analysis of the extract's unpolar fraction revealed the existence of dodecane and tetradecane. Structures of the isolated compounds (1-6) were confirmed by NMR and EI-MS spectrometry. Antimicrobial, antiviral and antitumour activities of the fruit parts were discussed. The promising combined extract of rind and flesh was biologically studied for microbial and cytotoxic activities in comparison with the whole isolated components.