Structure And Digestion Stability Of Liposomes And Their Formation Mechanism

Liposomes, formed by amphiphilic phospholipid, are delivery systems usually used in pharmaceutics, food and cosmetics. Liposomal stability is one of the most important factors in their application. However, liposomal structure are easy effected by O2, light, acid and enzyme during storage or digestion, which seviously restric their application. In addition, liposome formation mechanism, which is fundamental and the core of liposome research, still has no finally systematic conclusion. This project mainly focused on:(1) Blank liposomes were labled by fluorescent probe or encapsulated by model protein, lactoferrin, and their in vitro digetion stability was estimated; (2) In order to improve physical-chemical and digestion stability of liposomes, a polyelectrolyte delivery system (PDS) was prepared by layer-by-layer self-assembly based on chitosan (CH)-alginate (AL)-coated nanoliposomes; (3) Liposome formation mechanism was investigated using a dynamic high pressure microfluidization (DHPM), and the relationship between liposomal structure and thermodynamic stability was studied as well. The results were as follows.Fluorescence probe (calcein) labled crude liposomes and nanoliposomes, respectively, formed both from a milk fat globule membrane (MFGM) phospholipid fraction and from soybean phospholipid were prepared by thin layer dispersion and dynamic high pressure microfluidization methods. Their physical and chemical properties (average diameter, zeta potential, microstructure, lipolysis, and membrane permeability) were little influenced in simulated gatric fluid (SGF) containing pepsin. However, the liposomes exhibited lower stability in simulated intestinal fluid (SIF). Besides, liposomes loaded with positively charged lactoferrin were prepared from MFGM phospholipids using the thin layer dispersion method. The entrapment efficiency of lactoferrin encapsulated in the liposomes was about 46%. The entrapped lactoferrin remained unchanged as a function of time and pepsin concentration when the liposome samples were digested in SGF. However, in SIF, the entrapped lactoferrin was more susceptible to hydrolysis by the protease in pancreatin, as shown by changes in the average diameter, free fatty acid release and membrane structure of the liposomes.The optimized formular of PDS, of which CH concentration was 0.6% and AL concentration was 0.5%, was obtained based on the average diameter, surface charge, CH coating efficiency and AL sedimentation efficiency. The average diameter and zeta potential of PDS were about 330 nm and-15 mV, respectively, with medium chain fatty acids (MCFAs) encapsulation efficiency 67%. Besides, morphology and Fourier Transform Infrared Spectroscopy (FTIR) observation confirmed PDS has been successfully coated by CH and AL. The physical stability studies (pH and heat treatment at 70℃) indicated that the average diameter and surface charge of PDS altered more significantly that those of nanoliposomes. However, the core (nanoliposome) was protected by the out-layered polymers from damage. The apperence, chromatic aberration, value of malondialdehyde of PDS changed less than those of nanoliposomes after heat teatment for 48 h. Besides, the average diameter of PDS was remarkly affected by onic strength. Further enzymic digestion stability studies demonstrated that PDS could better resist lipolytic degradation and facilitate lower level of encapsulated component release in SIF (fit pseudo-first-order kinetic model), and the PDS release less MCFAs than nanoliposomes (fit Ritger-Peppas model). This work suggested that deposition of polyelectrolyte on the surface of nanoliposomes can improve physical-chemical stability and digestion stability of PDS under enzymic conditions.The properties (average diameter, surface charge, surface tention and relative turbidity), structure (morphology, FTIR, membrane integrity and permeability) and thermodynamic stability (long-term storage(180 d), physical stability (sonication and centrifugation) and chemical stability (pH and ionic strength)) of liposomes before and after treating with DHPM were studied. Based on the physical-chemical properties and structure of both crude liposomes and nanoliposomes, three liposomal formation mechanism models were obtained. Firstly, liposomes are formed by swelling and self-assembly when no energy or little energy is input. Secondly, liposomes are mainly formed by budding after inputting proper energy. Thirdly, when the liposomes are input high energy, they can be disrupt and then bend and self-closed to smaller liposomes. In addition, according to the changes of liposomal structure and thermodynamic stability of crude liposomes and nanoliposomes, their relationships were speculated. One is the preparation method, which affects the arrangement of liposomes and the conformations of membrane. The other is the conditions. The pH and ion concentration in liposome solution could alter the angle of polar head-groups of phospholipids and membrane surface, which change the surface charge of liposomes. Thus, the thermodynamic stability is affected by the preparation method and conditions.