Multicomponent insulin-containing microparticles are prepared by layer-by-layer assembly of dextran sulfate and chitosan on the core of protein-polyanion complex with or without protease inhibitors. Oral bioavailability of the encapsulated insulin is improved due to the cumulative effect of each component. A physico-chemical study shows that the particle design allows adjustment of the pH-dependent profile of the insulin release, as well as mucoadhesive properties and Ca2+ binding ability of the microparticles. Supplementing the microparticles with 2–3% protease inhibitors fully prevents proteolysis of human insulin. The pharmacological effect of microencapsulated insulin in doses 50–100 IU kg−1 is demonstrated in chronic experiments after oral administration to diabetic rats fed ad libitum.