Integrity of Membrane Structures in Giant Unilamellar Vesicles as Assay for Antioxidants and Prooxidants

• Published in: Analytical chemistry (Washington) 2018-02, Vol.90 (3), p.2126-2133
• Main Authors: Liu, Xiao-Chen, Du, Hui-Hui, Fu, Li-Min, Han, Rui-Min, Wang, Peng, Ai, Xi-Cheng, Zhang, Jian-Ping, Skibsted, Leif H
• Format: Article
• Language: English
• Subjects: Antioxidants; Antioxidants - chemistry; beta Carotene - chemistry; Bioassays; Chemistry; Chlorophyll; Chlorophyll A - chemistry; Chlorophyll A - radiation effects; Curve fitting; Destruction; Diffusion; Entropy; Fluorescence; Glycine max - chemistry; Integrity; Kinetics; Lag phase; Lecithin; Light; Membrane structures; Membranes; Microscopy; Morphology; Optical microscopy; Oxidative Stress - radiation effects; Parameter sensitivity; Phosphatidylcholine; Phosphatidylcholines - chemistry; Photosensitizing Agents - chemistry; Photosensitizing Agents - radiation effects; Singlet Oxygen - chemistry; Soybeans; Stability; Unilamellar Liposomes - chemistry; Unilamellar Liposomes - radiation effects; β-Carotene
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/acs.analchem.7b04383

We have attempted to evaluate, on the basis of optical microscopy for a single giant unilamellar vesicle (GUV), the potency of antioxidants in protecting GUV membranes from oxidative destruction. Photosensitized membrane budding of GUVs prepared from soybean phosphatidylcholine with chlorophyll a (Chl a) and β-carotene (β-Car) as photosensitizer and protector, respectively, were followed by microscopic imaging. A dimensionless entropy parameter, ΔE, as derived from the time-resolved microscopic images, was employed to describe the evolution of morphological variation of GUVs. As an indication of membrane instability, the budding process showed three successive temporal regimes as a common feature: a lag phase prior to the initiation of budding characterized by LP (in s), a budding phase when ΔE increased with a rate of k ΔE (in s–1), and an ending phase with morphology stabilized at a constant ΔE end (dimensionless). We show that the phase-associated parameters can be objectively obtained by fitting the ΔE–t kinetics curves to a Boltzmann function and that all of the parameters are rather sensitive to β-Car concentration. As for the efficacy of these parameters in quantifying the protection potency of β-Car, k ΔE is shown to be most sensitive for β-Car in a concentration regime of biological significance of