The biophysical mechanisms that underpin the interaction between microbubbles and cells in the context of ultrasound-mediated drug delivery are still poorly understood. To aid the identification of these mechanisms, giant unilamellar vesicles (GUVs) were used as cell models to quantify changes in membrane properties as a result of the interaction with ultrasound (1 MHz, 150 kPa, and 60 s continuous wave) and phospholipid-shelled microbubbles (DSPC-PEG40S 9:1 molar ratio), either alone or in combination. The spatial quantification of the vesicle lipid order was performed via spectral microscope imaging, by measuring the contours of generalized polarisation (GP) from the emission spectrum of c-Laurdan, a polarity-sensitive dye. Preliminary data show synergistic mechanical and chemical effects on membranes: ultrasound exposure and shear flow alone generally decrease the vesicle lipid packing, while exposures involving microbubbles reveal contrasting effects depending on the initial vesicle composition and acoustic regime. Results from the present mechanistic study provide an insight into the mechanisms of microbubble-membrane interactions, potentially benefitting the design of effective and predictable microbubble-based ultrasound treatments.