Interaction of Oil and Lipids in Freestanding Lipid Bilayer Membranes Studied with Label-Free High-Throughput Wide-Field Second-Harmonic Microscopy

Abstract

The interaction of oils and lipids is relevant for membrane biochemistry since the cell uses bilayer membranes, lipid droplets, and oily substances in its metabolic cycle. In addition, a variety of model lipid membrane systems, such as freestanding horizontal membranes and droplet interface bilayers, are made using oil to facilitate membrane monolayer apposition. We characterize the behavior of excess oil inside horizontal freestanding lipid bilayers using different oils, focusing on hexadecane and squalene. Using a combination of second-harmonic (SH) and white-light imaging, we measure how oil redistributes within the membrane bilayer after formation. SH imaging shows that squalene forms a wider annulus compared with hexadecane, suggesting that there is a higher quantity of squalene remaining in the bilayer compared with hexadecane. Excess oil droplets that appear right after membrane formation are tracked with white-light microscopy. Hexadecane droplets move directionally to the edge of the membrane with diffusion constants similar to those of single lipids, whereas squalene oil droplets move randomly with lower diffusion speeds similar to lipid condensed domains and remain trapped in the center of the bilayer for ∼1-3 h. We discuss the observed differences in terms of different coupling mechanisms between the oil and lipid molecules induced by the different chemical structures of the oils.

Investigating the structural properties of hydrophobic solvent-rich lipid bilayers

“…As the diffusion coefficient is generally inversely proportional to molecular size (e.g. in Stokes-Einstein equation), our observation that individual oil molecules diffuse much faster than individual PLs is hence in agreement with the experimental measurement of Roke and coworkers 42 .…”

“…As we observed that hexadecane molecules do not occupy exclusively the space between monolayers, but can also extend between the lipid acyl chains (Figure 2B), we next characterized the molecular orientation of oil molecules, to quantify to what extent they interdigitate with DOPC acyl chains as previously suggested 42 . To do so, we computed the orientation of the oil molecules with respect to the membrane normal (Figure 3).…”

Fat inclusions strongly alter membrane mechanics

“…Despite contrasting chemistry, the two neutral lipids yielded similar elasticity. As for triolein, squalene may also adopt phospholipid-parallel or antiparallel orientations in the bilayer (46), depending on the surface tension, in a way that decreases the area expansion modulus.…”

A new approach for investigating the response of lipid membranes to electrocompression by coupling droplet mechanics and membrane biophysics

“…The maximum cholesterol mole fraction used in this work was 30%, ensuring cholesterol solubility with phospholipids and avoiding cholesterol precipitation [35,36]. Hexadecane (99%-Sigma-Aldrich) was used for the oil phase, as it has shown to enable the formation of relatively solvent-free model membranes in comparison to shorter-chain alkanes, such as decane [19,37]. royalsocietypublishing.org/journal/rsif J. R. Soc.…”

“…Measuring the changes in the membrane thickness under electrocompression provides information on the disjoining pressure and membrane mechanics (figure 2). It has been observed that the thickness properties and behaviour depend considerably on the oil medium [19,20,37,46]. As an example, figure 8 shows the changes in specific capacitance of three separate DIBs-submerged in three different oils-with respect to an increasing DC voltage.…”