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Proc Natl Acad Sci U S A. 2003 Aug 19;100(17):9826-30. Epub 2003 Aug 6.

Ordering of water molecules between phospholipid bilayers visualized by coherent anti-Stokes Raman scattering microscopy.

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  • 1Department of Chemistry and Chemical Biology, Division of Engineering and Applied Sciences, Harvard University, 12 Oxford Street, Cambridge, MA 02138, USA.


We demonstrate ordered orientation of the hydration water at the surface of phospholipid bilayers by use of coherent anti-Stokes Raman scattering (CARS) microscopy, a highly sensitive vibrational imaging method recently developed. We investigated negatively charged POPS (1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-l-serine) and neutral POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) multilamellar onions dispersed in deuterated dodecane. The imaging contrast based on the CARS signal from the H2O stretching vibration shows a clear dependence on the excitation field polarization. Our results provide direct experimental evidence that water molecules close to the phospholipid bilayer surface are ordered with the symmetry axis along the direction normal to the bilayer. Moreover, the amount of ordered water molecules depends on the lipid polar group. The spectral profile for the inter-lamellar water shows that the water molecules bound to the bilayer surface are less hydrogen-bonded and exhibit a higher vibrational frequency than bulk water.

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