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UBC Theses and Dissertations

Synthesis of a model lipid and observation of behaviour of model liposomes by electrophoresis Song, Xu Chun


A model lipid consisting of a cholesterol base, tetraethoxy— spacer and glucuronic acid head group was synthesized. First, the head group was prepared by acetylation and esterification of glucuronolactone to produce methyl (1, 2,3,4-tetra-O— acetyl——D—glucopyran)uronate (j.) which was then brominated to produce methyl (2,3,4—tri-O—acetyl—--D—glucopyranosyl bromide)uronate (Z). The combination of the cholesterol part and tetra— ethoxy chain was made by reacting cholesteryl—p—toluenesulfonate and tetraethylene glycol, to produce 3—O—(i1-hydroxy—3, 6, 9—trioxaundecyl)cholest —5-ene (tetra-EC) (3). The above steps were carried out with methods reported previously. The coupling of the head group and tetra-EC employed a different method, which had been used by others in the coupling reaction of the same head group and cholesterol, by using silver oxide as the catalyst instead of silver carbonate. Methyl t3—O--(3,6,9—trioxaundecyl) cholest—5—en—3-yl—2,3, 4—tri—O—acetyl——D—glucopyranosid3uronate () was produced from the coupling reaction with a yield estimated to be — 50’!., higher than that of the reaction with silver carbonate as the catalyst. The final step was to remove the methyl group and the acetyl protecting groups on the head group by using excess MaCH in a specific solvent system and acidifying with HC1, to obtain crude 3—O—(3,6,9—trioxundecyl) cholest—5—en—3—yl——D—glucopyranosiduronic acid (). The crude acid product was primarily purified by adjusting the pH of the suspension of the acid in warn ethanol and water and the salt form was obtained. The salt product, (i), was precipitated pure from chloroform solution by addition of mixed ethyl acetate arid hexane. The product (tetra—ECG), which has a negative charge on the head group, was used with other lipids to prepare liposomes. The liposomes, which vary in poly(ethylene glycol) (PEG) chain density and charge location were made for the purpose of mimicking the glycocalyx region in actual biomembranes. Particle electrophoresis was used to measure the mobilities of the liposomes in the solutions with variation of pH (1.8—9.9), ionic strength (O.OO1M—O.1H) and PEG chain density (O-60’A by molar ratio). The classical theory for particle electrophoresis was applied to calculate the mobility and the apparent charge density of the liposoznes. The pKa of tetra—ECG was determined with a plot of the mobility of the tetra—ECG—containing liposome against pH on the surface of the particle. A numerical model, which has been developed as a computational program, was used to interpret the results of electrophoresis as a function of ionic strength, in terms of several parameters which describe the surface properties of the liposomes.

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