A novel, salting-out type of protein purification method was described by B. H. Odegaard, P. C. Anderson, and R. E. Lovrien (J. Appl. Biochem. 6, 156, 1984) that included the use of the water-miscible organic solvent, tert-butanol, which forms a separate phase when layered on 20-40% relative saturation ammonium sulfate solutions. Of proteins dissolved in the salt solutions, fractions can be precipitated which, contrary to the sedimentation found in classical salting-out, form a protein disc or midlayer between the organic and aqueous phases, Various inorganic salts, KCl, MgCl2, CaCl2, Na2SO4, MgSO4, (NH4)(2)SO4, applied at equimolar concentrations (1.5 mol.dm(-3)) were found to affect the partitioning of crude neutral proteinase solution differently. The amount of precipitated material absorbing at 280 nm exhibited proportionality to the interfacial tension of the 1.5 mol.dm(-3) salt solutions equilibrated with 30 vol % tert-butanol as well as to the density of the lower phase. Results obtained with a purified fraction showed a trend essentially similar to the case when original crude enzyme solution was applied.