Non-polar lipid content and fatty acid (FA) composition of the commercial edible strain of Pleurotus ostreatus and the medicinal wild strain of Ganoderma australe were determined. A comparison of the FA profile was conducted between mycelium grown under optimum bioreactor conditions and naturally occurring fruit bodies in both species. Both strains contained unsaturated FA (UFA), amounting to 55–77% of total FA content, whereas the proportion of essential FA was contributive, permitting their consideration as potential food ingredients. Bioreactor process resulted in a significant total FA content increase accompanied with a considerable effect on ratios of nutritional interest (MUFA/SFA, PUFA/SFA, oleic/linoleic).