Because liposomes containing fluoroalkylated phospholipids are being developed for in vivo drug delivery, the structure and interactive properties of several fluoroalkylated glycerophosphocholines (PCs) were investigated by x-ray diffraction/osmotic stress, dipole potential, and hydrophobic ion binding measurements. The lipids included PCs with highly fluorinated tails on both alkyl chains and PCs with one hydrocarbon chain and one fluoroalkylated chain. Electron density profiles showed high electron density peaks in the center of the bilayer corresponding to the fluorine atoms. The height and width of these high density peaks varied systematically, depending on the number of fluorines and their position on the alkyl chains, and on whether the bilayer was in the gel or liquid crystalline phase. Wide-angle diffraction showed that in both gel and liquid crystalline bilayers the distance between adjacent alkyl chains was greater in fluoroalkylated PCs than in analogous hydrocarbon PCs. For interbilayer separations of less than about 8 A, pressure-distance relations for fluoroalkylated PCs were similar to those previously obtained from PC bilayers with hydrocarbon chains. However, for bilayer separations greater than 8A, the total repulsive pressure depended on whether the fluoroalkylated PC was in a gel or liquid-crystalline phase. We argue that these pressure-distance relations contain contributions from both hydration and entropic repulsive pressures. Dipole potentials ranged from -680 mV for PCs with both chains fluoroalkylated to -180 mV for PCs with one chain fluoroalkylated, compared to +415 mV for egg PC. The change in dipole potential as a function of subphase concentration of tetraphenyl-boron was much larger for egg PC than for fluorinated PC monolayers, indicating that the fluorine atoms modified the binding of this hydrophobic anion. Thus, compared to conventional liposomes, liposomes made from fluoroalkylated PCs have different binding properties, which may be relevant to their use as drug carriers.

The syntheses of various fluorocarbon/Ruorocarbon and fluorocarbon/hydrocarbon rac-l,2-and 1,3-di-0-alk(en)ylglycerophosphocholines and rac-l,2-di-O-alkylglycerophosphoethanolamines (see Fig. 2), which may be used as components for drug-carrier and delivery systems, are described together with some results concerning their biological tolerance. They were obtained by phosphorylation of perfluoroalkylated ruc-di-0 -alk(en)ylglycerols using POCI,, then condensation with choline tosylate or N-Boc-ethanolamine (2-[(tert -butoxy)carbonylaminolethanol) followed by Boc-deprotection (Schemes 6-8). The flnorocarbon/fluorocarbon 1,2-di-0 -alkylglycerols were prepared by 0-alkylation of ruc-1-0-benzylglycerol using perfluoroalkylated mesylates, then hydrogenolysis for benzyl deprotection (Scheme 1 ). The two different hydrophobic chains in the mixed fluorocarbon/ fluorocarbon and fluorocarbon/hydrocarbon 1,2-di-O-alk(en)ylglycerols were introduced starting from 1,2-0 -isopropylidene-then 0 -trityl-protected glycerols or from 1,3-0 -benzylidene-glycerol (Schemes 3 and 4). The perfluoroalkylated 0-alkenylglycerols were obtained by 0-alkylation of a glycerol derivative using an o-unsaturated alkenyl reagent, the perfluoroalkyl segment being connected onto the double bond in a subsequent step (Schemes 1 and 3 ) . The perfluoroalkylated symmetrical and mixed 1,3-di-0 -alkylglycerols were synthesized by displacement of the C1-atom in epichlorohydrin by perfluoroalkylated alcohols, then catalytic (SnCI.,) opening of the oxirane ring of the resulting alkyl glycidyl ethers in neat alcohols (Scheme 5 ) . When injected intravenously into mice, acute maximum tolerated doses higher than 1500 and 2000 mg/kg body weight were observed for the fluorinated glycerophosphocholines, indicating a very promising in vivo tolerance.

The stability (with respect to encapsulated carboxyfluorescein release) of liposomes made from various fluorocarbon 1,2-or 1,3-di-O-alkylglycerophosphocholines (ether-connected) and their membrane permeability have been investigated in buffer and in human serum. Membranes and liposomes, whether formulated with fluorocarbon/fluorocarbon or mixed fluorocarbon/hydrocarbon, 1,2- or 1,3-di-O-alkylglycerophospholipids, display lower permeability coefficients and are able to retain more efficiently encapsulated CF, even when incubated in human serum, than any of their conventional counterparts. These fluorinated liposomes are as stable as the first generation of liposomes formulated with their fluorocarbon ester-connected 1,2-di-O-acylglycerophosphocholine analogs. These results further confirm that a fluorinated intramembranar layer reduces the permeability of membranes (more significantly when they are in a fluid state), protects them from the destabilizing effects of serum components and increases even the stability of the fluorinated liposomes whose membranes are in the gel state when incubated in human serum. The impact of the modular structure of the fluorinated phospholipids (number of fluorocarbon chains, ether vs. ester bond, 1,2- vs. 1,3-isomer, etc...) and structure/permeability/ stability relationships are also presented.