The orientational and hydrogen-bonding properties of water confined between lipid bilayers of a multilamellar vesicle of DPPC (dipalmitoyl-sn-glycero-3-phosphocholine) in D2O have been investigated using multiplex coherent anti-Stokes Raman (CARS) microscopy. Spatially resolved analysis of the complete OD- and CH-stretch vibrational spectral region (2200-3200 cm(-1)) reveals that part of the (confined) interlamellar water has a weaker hydrogen-bond network than bulk water. Polarization dependent studies show negligible orientational order of the interstitial water molecules, in contrast to earlier CARS studies. Possible reasons for this discrepancy are explained in terms of interference between lipid and water resonances. (c) 2006 Elsevier B.V. All rights reserved.