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In this study, we used fluorescence microscopy to investigate the incorporation of droplets composed of triolein, trilinolein, trimyristolein, trieicosenoin, and cholesteryl arachidonate in the bilayers of giant unilamellar vesicles (GUVs) consisting of a mixture of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), and cholesterol. After incorporating the triacylglycerol droplets, the DOPC/DPPC/cholesterol (3:3:2) GUVs, which exhibited liquid-disordered (Ld) and liquid-ordered (Lo) phase separation, retained their phase-separated state. The triacylglycerol droplets predominantly partitioned in the Ld domains. To elucidate the basis of this preferential partitioning, we investigated the surface pressures of DOPC, DPPC, and cholesterol monolayers containing triolein at the air–water interface using a Langmuir trough. From these measurements, we determined the interfacial tension at the monolayer-covered triolein–water interface. The results showed that DOPC most effectively reduced the interfacial tension. Thus, the droplet sorting into the DOPC-enriched Ld domains likely arose from the difference in the abilities of the two phases to stabilize the droplet interface. In contrast, cholesteryl arachidonate had a profound effect on the bilayer phase behavior. Fluorescence images of the DOPC/DPPC/cholesterol (3:3:2) GUVs showed that the domain structures disappeared after droplet incorporation. Additionally, surface pressure measurements of DOPC/DPPC/cholesterol (3:3:2) monolayers containing cholesteryl arachidonate at the air–water interface suggested that cholesteryl arachidonate weakened the lipid–lipid interaction. The results indicate that the cholesteryl arachidonate molecules diffused across the bilayer to hinder bilayer phase separation.

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  This document is the Accepted Manuscript version of a Published Work that appeared in final form in Langmuir, copyright © 2025 American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see https://doi.org/10.1021/acs.langmuir.4c05063.

Keyword: Lipid droplets, triacylglycerols, giant unilamellar vesicles, GUVs

Date published: 2025-03-11

Publisher: American Chemical Society (ACS)

Journal:

• Langmuir (ISSN: 07437463) vol. 41 issue. 9

Funding:

• Japan Society for the Promotion of Science 22K05179

Manuscript type: Author's version (Accepted manuscript)

MDR DOI: https://doi.org/10.48505/nims.5365

First published URL: https://doi.org/10.1021/acs.langmuir.4c05063

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Updated at: 2026-02-26 12:30:10 +0900

Published on MDR: 2026-02-26 09:41:06 +0900