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Coadsorption of Human Milk Lactoferrin into the Dipalmitoylglycerolphosphatidylcholine Phospholipid Monolayer Spread at the Air/Water Interface

Fausto Miano ^*, Xiubo Zhao , Jian R. Lu  and Jeff Penfold 

^* Società Industria Farmaceutica Italiana SpA, Lavinaio (Catania), Italy; Biological Physics Group, School of Physics and Astronomy, The University of Manchester, Sackville Street Building, Manchester M60 1QD, United Kingdom; and ISIS Facility, Rutherford Appleton Laboratory, The Council for the Central Laboratory of the Research Councils, Chilton, Didcot OX11 0QZ, United Kingdom

Correspondence: Address reprint requests to Jian R. Lu, Tel.: 44-161-2003926; E-mail: j.lu{at}Manchester.ac.uk.

The coadsorption of human milk lactoferrin into a spread monolayer of dipalmitoylglycerol phosphatidylcholine (DPPC) at the air/water interface has been studied by neutron reflection. The system is a good model of the preocular tear film outer interface, which was the motivation for the study. The association of the protein with the surface was indicated by an increase of the surface pressure exerted by the DPPC monolayer. The extent of lactoferrin coadsorption was found to decrease with increasing surface pressure in the lipid monolayer, a trend consistent with the observation reported for other proteins, such as lysozyme and ß-lactoglobulin. The neutron reflectivity measurements were subsequently carried out at the three surface pressures of 8, 15, and 35 mN/m to examine the structure and composition of lactoferrin coadsorbed at the interface. Whereas the DPPC monolayer effectively prevented lactoferrin insertion at the high surface pressure, a measurable amount of lactoferrin was found at the air/water interface at the two lower surface pressures. At 15 mN/m it was difficult to identify the distribution of lactoferrin with respect to the DPPC monolayer, due to its relatively low adsorbed amount and much broader distribution. At the lowest surface pressure of 8 mN/m, the lactoferrin coadsorption was found to increase with time over the first few hours. After 5 h the distribution of the lactoferrin layer became similar to, though quantitatively lower than, that adsorbed in the absence of the DPPC monolayer. It is characterized by a top dense sublayer of 15 Å with a bottom diffuse sublayer of 60 Å, indicating structural unfolding induced by surface adsorption under these conditions.