Biosurfactants are amphipathic molecules produced by plants, animals, and microorganisms, that present emulsifying properties and may act reducing surface and interfacial tensions. When compared to synthetic surfactants, these biological analogues have high biodegradability potential, and may be produced from renewable raw materials within overall biotechnological processes involving low generation of residues. The production and application of microbial surfactants have been recently considered in several industrial sectors, as these low toxicity versatile compounds find applications in food, pharmaceutical, cosmetic, and petrochemical products, in nanotechnology and agriculture, and in the bioremediation of xenobiotic-contaminated areas. Herein, the main conceptual aspects and physicochemical properties, as well as the classifications of biosurfactants according to their origin and their chemical structures, are addressed. The production of microbial biosurfactants through sustainable processes are also described, with particular focus on new applications and on the increasing relevance of such bioproducts for the sustainable development of modern society.