A study on the hydration property of lipid bilayer systems was performed with DSC and X-ray diffraction techniques for two sphingomyelins, a naturally occurring bovine brain sphingomyelin having a heterogeneous acyl chain and a semisynthetic sphingomyelin having the acyl chain of 16 saturated carbons. The number of differently bound interlamellar water molecules was estimated for the two sphingomyelin systems from a deconvolution analysis of the ice-melting DSC curves of varying water content. The estimated limiting, maximum number of freezable interlamellar water molecules for the semisynthetic sphingomyelin system was 5.5 H2O per molecule of lipid, which is close to a value (5 H2O/lipid) previously reported by us for a system of dipalmitoylphosphatidylcholine. However, for the brain-sphingomyelin system, a limiting hydration for this type of water was not reached even up to the water/lipid molar ratio similar to20. In this accord, the electron density profile analysis of X-ray lamellar diffraction patterns for the brain-SM system showed that the thickness of interlamellar water layer increases in proportion to the amount of added water. (C) 2003 Elsevier B.V. All rights reserved.