Modulation of liposome matrix by cholesterol for efficient localization of photosensitizer. A fluorescence quenching study on chlorophyll extract from spinach leaves

Abstract

To develop potential biomimetic solar energy conversion devices, egg phosphatidylcholine (EPC) liposomes were prepared with extracts from spinach leaves as the photosensitizer. Fluorescence energy transfer studies using these liposomes established that the extract consists of an organized assembly of components such as chlorophyll a (Chl a), chlorophyll b (Chl b), pheophytin, etc. For homogeneous solutions in CHCl 3, the spectroscopic properties of the extract are as follows: Chl a, λ ab=430 nm (Soret) and 668 nm (Q), λ fl=670 nm; Chl b, λ ab=460 nm (Soret), λ fl = 650 nm; in liposomes: Chl a, λ ab = 420 nm, λ fl = 678 nm, λ exc = 420 nm; Chl b, λ ab = 460 nm, λ fl = 678 nm, λ exc=470 nm (monitored at 678 nm). A fluorescence quenching technique using anthraquinone-1,5-disulphonate (1,5-AQDS 2−) as the electron acceptor (A) and ethylenediaminetetraacetic acid (EDTA) as the sacrificial electron donor (D), was employed to study the localization of the photosensitizer in the liposome matrix on modulation with cholesterol. The Stern—Volmer quenching constants K sv for 1,5-AQDS 2− are nearly diffusion controlled in homogeneous solutions: K sv= 1100 M −1 in CH 3COCH 3-H 2O (60:40) and K sv = 1420 M −1 in CH 3CN-H 2O (60:40). In unmodified liposomes with A in the outer pool and D in the inner pool, the value decreases to 540 M −1; this value is reduced further on addition of 10% cholesterol, remains constant up to 30% cholesterol and decreases sharply on addition of 50% cholesterol. A reverse situation is observed when A is located in the inner pool and D is added gradually to the outer pool: a low value is obtained for 0% cholesterol, which increases slightly at 10% cholesterol, remains constant up to 30% cholesterol and increases sharply at 50% cholesterol. These changes indicate the movement of Chl a into the interior of the liposome and follow the changes in structure and size of EPC liposomes on addition of cholesterol as discussed by Huang and Mason. The distribution of Chl a within these modified unilamellar liposomes is a function of the physicochemical properties of the matrix and the dimensions of the liposome. Our study indicates that for constant performance, EPC liposomes containing 15% cholesterol may be prepared to improve the structural fluidity of the liposome, to obtain appropriate localization of the photosensitizer and to optimize the efficiency of quenching and stability of the system.