Synergy between plant phenols and carotenoids in stabilizing lipid-bilayer membranes of giant unilamellar vesicles against oxidative destruction.

We have investigated the synergism between plant phenols and carotenoids in protecting the phosphatidylcholine (PC) membranes of giant unilamellar vesicles (GUVs) from oxidative destruction, for which chlorophyll-a (Chl-a) was used as a lipophilic photosensitizer. The effect was examined for seven different combinations of ß-carotene (ß-CAR) and plant phenols. The light-induced change in GUV morphology was monitored via conventional optical microscopy, and quantified by a dimensionless image-entropy parameter, ΔE. The ΔE-t time evolution profiles exhibiting successive lag phase, budding phase and ending phase could be accounted for by a Boltzmann model function. The length of the lag phase (LP in s) for the combination of syringic acid and ß-CAR was more than seven fold longer than for ß-CAR alone, and those for other different combinations followed the order: salicylic acid < vanillic acid < syringic acid > rutin > caffeic acid > quercetin > catechin, indicating that moderately reducing phenols appeared to be the most efficient membrane co-stabilizers. The same order held for the residual contents of ß-CAR in membranes after light-induced oxidative degradation as determined by resonance Raman spectroscopy. The dependence of LP on the reducing power of phenols coincided with the Marcus theory plot for the rate of electron transfer from phenols to the radical cation ß-CAR˙+ as a primary oxidative product, suggesting that the plant phenol regeneration of ß-CAR plays an important role in stabilizing the GUV membranes, as further supported by the involvement of CAR˙+ and the distinct shortening of its lifetime as shown by transient absorption spectroscopy.

Peroxyl radical induced membrane instability of giant unilamellar vesicles and anti-lipooxidation protection.

The present work is intended to investigate the morphological instability of lipid membrane induced by peroxyl radical (ROO•) and the underlying mechanism. To this end, the giant unilamellar vesicle (GUV) made from phosphatidylcholine was employed as a membrane model, and the azo compounds 2,2'-azobis(2,4-dimethylvaleronitrile) (AMVN) and 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) were used as the precursors of ROO•. Upon mild pyrolysis, the GUV immobilized in agarose gel was followed by conventional optical microscopy in real time, and the morphological variation was quantified by the image heterogeneity, perimeter and area all as a function of time for up to an hour. Lipid oxidation initiated from lipid phase with AMVN and from aqueous phase with AAPH led to different types of morphological changes, i.e. membrane coarsening and vesicle deformation/budding, respectively. Based on the compositional analysis of lipid oxidation products, we propose that ROO• as the primary radical initiator is responsible for the morphological changes of the GUV-AMVN while both ROO• and RO• are responsible for the morphological changes of the GUV-AAPH system. Lipophilic ß-carotene and amphipathic plant phenols as antioxidants are found to be able to stabilize the membrane integrity effectively, in corroboration with the proposed mechanisms for membrane destruction.

Spatial effects of photosensitization on morphology of giant unilamellar vesicles.

Singlet oxygen (1O2) formed through photosensitization may initiate oxidative destruction of biomembranes, however, the influence from the spatial organization of photosensitizers (PS) relative to membranes remains unclear. To clarify this issue, we loaded riboflavin 5'-(dihydrogen phosphate) monosodium (FMN-Na) as a hydrophilic PS into the lumen of halloysite nanotubes (HNTs), and attached the nanoassemblies (FMN-Na@HNTs), via Pickering effects, to the outer surfaces of giant unilamellar vesicles (GUVs) of phospholipids. We also prepared GUVs dopped with lumiflavin (LF) as a lipophilic PS having a 1O2 quantum yield comparable to FMN-Na. FMN-Na capsulated in HNT was characterized by a longer triplet excited state lifetime (12.1 µs) compared to FMN-Na free in solution (7.5 µs), and FMN-Na in both forms efficiently generated 1O2 upon illumination. The spatio-effects of PS on the photosensitized morphological changes of membranes were studied using conventional optical microscopy by monitoring GUV morphological changes. Upon light exposure (400-440 nm), the GUVs attached with FMN-Na@HNT merely experienced membrane deformation starting from the original spherical shape, ascribed to Type II photosensitization with 1O2 as oxidant. In contrast, photooxidation of LF dopped GUVs mainly led to membrane coarsening and budding assigned to Type I photosensitization. The spatial effects of PS on photosensitized morphological changes were related to the different lipid oxidation products generated through Type I and Type II photosensitized lipid oxidation.

Naturally occurring nanotube with surface modification as biocompatible, target-specific nanocarrier for cancer phototherapy.

Phototherapy has drawn increasing attention including the use of nanocarriers with high drug loading capacity and delivery efficacy for target-specific therapy. We have made use of naturally-occurring halloysite nanotubes (HNTs) to build a biomimetic nanocarrier platform for target-specific delivery of phototherapeutic agents. The HNTs were decorated with poly(sodium-p-styrenesulfonate) (PSS) to enhance the biocompatibility, and were further functionalized by lumen loading the type-II photosensitizer indocyanine green (ICG). The HNT-PSS-ICG nanocarrier, without further tethering targeting groups, was shown to associate with the membrane of giant unilamellar vesicles (GUVs) via Pickering effects. Application of HNT-PSS-ICG nanocarrier to human breast cancer cells gave rise to a cell mortality as high as 95%. The HNT-PSS-ICG nanocarrier was further coated with MDA-MB-436 cell membranes to endow it with targeting therapy performance against breast cancer, which was confirmed by in vivo experiments using breast cancer tumors in mice. The membrane-coated and biocompatible nanocarrier preferentially concentrated in the tumor tissue, and efficiently decreased the tumor volume by a combination of photodynamic and photothermal effects upon near-infrared light exposure. Our results demonstrate that the HNT-based nanocarrier by virtue of facial preparation and high loading capacity can be a promising candidate for membrane-targeting nanocarriers.