In recent years, biosurfactants have attracted attention because of their low toxicity, high biodegradability, and good ecological acceptability. However, their production in submerged liquid culture is hampered by the severe foaming that occurs. Solid-state cultivation can avoid this problem. In the current work, we optimized the production of a rhamnolipid biosurfactant by Pseudomonas aeruginosa UFPEDA 614, grown on a solid medium impregnated with a solution containing glycerol. During the study, we increased the production of the biosurfactant over tenfold, with levels reaching 172 g of rhamnolipid per kilogram of dry initial substrate after 12 days. On the basis of the volume of impregnating solution added to the solid support, this yield is of the order of 46 g/L, which is comparable with the best results that have been obtained to date in submerged liquid cultivation. Our results suggest that there is a great potential for using solid-state cultivation for the production of rhamnolipids.