Partitioning of a variety of nonionic and acidic organic compounds was examined in aqueous dextran–poly(ethylene glycol) (PEG) two‐phase systems of different ionic composition at pH 2.00 and 7.40. Relative hydrophobicity of the compounds was estimated in terms of equivalent numbers of methylene units. Effect of the ionic composition and pH of aqueous media on the relative hydrophobicity of organic compounds was established and described quantitatively. The results obtained indicate that the ionic composition affects the relative hydrophobicity of ionized compounds and has no effect on that of nonionic compounds and ionizable compounds in the undissociated state. Ionization of a given ionic group seems to bring about a constant change in the relative hydrophobicity of a compound. This result supports the applicability of the additivity principle to the relative hydrophobicity estimates obtained with the aqueous two‐phase partitioning technique. © 2001 Wiley‐Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 90:1366–1374, 2001