P32

22BG042, 23BG034

BL-13

Soft X-ray Absorption Spectroscopy of Phospholipid Films
Supported on Au Substrates by Different Casting
Techniques
Yuri Ohuraa, Kakuto Yoshiokab, Shogo Tendob, Akinobu Niozua,b, and Shin-ichi
Wadaa,b,caa

Faculty of Science, Hiroshima University, Higashi-Hiroshima 739-8526, Japan
Graduate School of Science and Engineering, Hiroshima University, Higashi-Hiroshima 739-8526, Japan
cHiroshima Synchrotron Radiation Center, Hiroshima University, Higashi-Hiroshima 739-0046, Japan

Keywords: Enter keywords. Linear polarization, XAS (X-ray absorption spectroscopy), NEXAFS (near edge X-ray absorption
fine structure), Spin-casting, Phospholipids, DPPC

Lipids are one of the major constituents of living organisms. In
particular, lipids self-assemble into oriented amphiphilic bilayers,which are the basic structure of the cell membranes. Lipid bilayers
not only play an important protective role in maintaining cells, butalso serve as sites for information transfer and protein reactions.
Therefore, understanding the properties of lipid membranes isfundamental to studying biological mechanisms in detail, and to thisend, attempts have been made to reproduce artificial cell membranes
by supporting lipids on metal substrates. Lipid membranes supported
on solid substrates (Figure 1), such as LB (Langmuir-Blodgett) andFIGURE 1. Molecular structure ofvesicle fusion methods, resemble natural cell membranes, and theDPPC phospholipids and schematic
information obtained are known to be biologically reliable. We havebilayer membrane.
found that even a drop of an organic solution of phospholipid
molecules onto a gold substrate can form a highly oriented multi lipid-bilayers. In this study, we preparedmulti lipid-bilayer membranes by spin-casting method of lipid solutions, aiming to create more uniformly
oriented bilayers and to evaluate the state of the bilayers.
DPPC shown in Figure 1 was used as the phospholipid and dissolved in chloroform to make a lipidsolution. The solution was dropped onto Au substrates and dried under a nitrogen stream to create a multi
lipid-bilayer membranes. The spin-casting method was also used to create a more uniform bilayer films onthe substrates.
The prepared lipid films were characterized by NEXAFS (near edge X-ray absorption fine structure)measurements at HiSOR BL13 and AFM (atomic force microscopy) measurements at the laboratory.
NEXAFS spectra were obtained by irradiating soft X-rays in the carbon K-edge region, and the emitted
electrons were detected as drain current. The polarization angle dependence of the NEXAFS spectra wasmeasured by changing the incidence angle of horizontally polarized X-rays on the sample surface, and theorientation angle of the organized lipid molecules was thereby determined to evaluate the differences in themultilayer films due to the different preparation methods.
In this study, soft X-ray absorption spectroscopy measurements were performed at different incident
angles of soft X-rays to investigate the orientation angles of lipid films prepared by drop and spin-casting
methods. Figure 2 shows NEXAFS spectra in the C K-edge region measured at different sample positions
for lipid films prepared by drop and the spectrum for the lipid film prepared by spin-casting. The obtainedNEXAFS spectra show characteristic resonant excitation peaks at specific energies and different spectral
shapes with respect to the incident angle of soft X-ray. In other words, both lipid membranes are highlyoriented, despite being multilayered.

Each spectrum can be fitted with a Gaussian function to extract each resonant excitation component.
From the area intensities of fitted functions, the orientation angle of the transition dipole moment at eachtransition was determined. As a result, there is a clear difference between the orientation angles of the lipid
films prepared by the drop and spin-casting methods. This is thought to be due to the difference in the method
used to prepare the lipid films and the difference in the thickness of the films.
From the Rydberg and σ*(C-C) transitions, the orientation angle of the hydrocarbon chains, which
determines the overall shape of the lipid molecules, can be derived specifically. In lipid films prepared by
the spin-casting method, the orientation angle is closer to 55 degrees, which is called the "magic angle",than in films prepared by drop-casting. Based on previous studies [1], it is thought that the orientation
ordering is reduced in lipid films obtained by spin-casting due to the random structure of some of thehydrocarbon chains. In addition, the NEXAFS spectra of the lipid films obtained by spin-casting are different
from those obtained by the drop-casting method in their detailed shape, and are in good agreement with thespectral shape of the single bilayer films in the previous study [1]. These results suggest that the spin-casted
lipid films are sufficiently thin to be considered single bilayer films, unlike the drop method, and that thedegree of freedom of the molecules constituting the films is increased in this ultra-thin state.

REFERENCES
1. M. Tabuse, A. Niozu, and S. Wada, HiSOR Activity Report 2021, 100-101 (2022).

2.5drop (center)

2.0
1.5
1.00.52040557090

θ：

0.0
280

285

290

295

Intensity / arb.units

2.5300305

310

drop (edge)

2.0
1.5
1.0

FIGURE 3. Typical result of Gaussian fitting obtainedfor drop-casting film at 55-degree incidence angle.

0.50.0280

285

290

295

2.5300305

310

spin-casting

2.0
1.5
1.0
0.50.0
280

285

290

295300305

310

Photon Energy / eV
FIGURE 2. Polarization dependent NEXAFS
spectra of (upper) center and (middle) edge positionsof DPPC drop-casting films and (c) spin-casting film
measured at C K-edge. θ: incident angle from thesurface, and therefore the angle of electric vector
from the surface normal.