Lipids, polymerizable

While most vesicles are formed from double-tail amphiphiles such as lipids, they can also be made from some single chain fatty acids [73], surfactant-cosurfactant mixtures [71], and bola (two-headed) amphiphiles [74]. In addition to the more common spherical shells, tubular vesicles have been observed in DMPC-alcohol mixtures [70]. Polymerizable lipids allow photo- or chemical polymerization that can sometimes stabilize the vesicle [65] however, the structural change in the bilayer on polymerization can cause giant vesicles to bud into smaller shells [76]. Multivesicular liposomes are collections of hundreds of bilayer enclosed water-filled compartments that are suitable for localized drug delivery [77]. The structures of these water-in-water vesicles resemble those of foams (see Section XIV-7) with the polyhedral structure persisting down to molecular dimensions as shown in Fig. XV-11. 

Elbert R, Laschewsky A and Ringsdorf H 1985 Hydrophilic spacer groups in polymerizable lipids— formation of biomembrane models from bulk polymerized lipids J. Am. Ohem. Soc. 107 4134-41... 

Influence of subphase temperature, pH, and molecular structure of the lipids on their phase behavior can easily be studied by means of this method. The effect of chain length and structure of polymerizable and natural lecithins is illustrated in Figure 5. At 30°C distearoyllecithin is still fully in the condensed state (33), whereas butadiene lecithin (4), which carries the same numEer of C-atoms per alkyl chain, is already completely in the expanded state (34). Although diacetylene lecithin (6) bears 26 C-atoms per chain, it forms both an expanded and a condensed phase at 30°C. The reason for these marked differences is the disturbance of the packing of the hydrophobic side chains by the double and triple bonds of the polymerizable lipids. At 2°C, however, all three lecithins are in the condensed state. Chapman (27) reports about the surface pressure area isotherms of two homologs of (6) containing 23 and 25 C-atoms per chain. These compounds exhibit expanded phases even at subphase temperatures as low as 7°C. 

Figure 8. Formation of polymeric monolayers from polymerizable lipids (X =...

Mixtures of proteins, natural and polymerizable lipids can be transferred into liposomes and polymerized hereafter. Initial experiments have shown that even very complex proteins such as F F.-ATPase can be incorporated in polymeric liposomes by this method under retention of the activity of the protein (76). 

Polymerizable lipids can be incorporated into e.g. erythrocyte ghost cells by means of hemolysis and polymerized here-... 

Figure 15. Schematic of the build up of stable cell models via partial polymerization of the membrane. Key

The most complete insights into the behavior of mixtures of nonpolymeriz-able lipids have come from the phase diagrams of these systems. These data have provided important reference points for the polymerizable lipid systems described next, even though few phase diagrams have been reported for polymerizable lipid mixtures. In spite of this deficiency the polymerization studies have... 

Domain formation in binary mixtures of a polymerizable lipid and non-polymerizable lipid is well established for diacetylenic lipids. The rigid diacetylenic unit facilitates the formation of enriched domains in the condensed phase of monolayers or the solid-analogous phase of bilayers. Since diacetylenes polymerize most readily in solid-like states, most studies have focused on conditions that favor domain formation. Only in the case of a mixture of a charged diacetylenic lipid and a zwitterionic PC was phase separation not observed. Ringsdorf and coworkers first reported the polymerization of a phase-separated two-dimensional assembly in 1981 [33], Monolayer films were prepared from mixtures consisting of a diacetylenicPC (6) (Fig. 5) and a nonpolymerizable distearoyl PE (DSPE). 

If one intends to synthesize polymerizable lipids to build up membranes of high stability the polymerizable group may be introduced into different parts of the lipid molecule, i.e., into the hydrophilic head group or the alkyl chain (Fig. 4)7). 

All four types of polymerizable lipids shown in Fig. 4 have been realized synthetically. In this context, one need not attempt to reproduce mother nature slavishly (Fendler 8)). Kunitake 9) was able to show that simple molecules like dialkyldimethyl-ammonium salts also form bilayer assemblies. Fuhrhop 10) and Kunitake U) could accomplish the formation of monolayer liposomes with molecules containing only one alkyl chain and two hydrophilic head groups. Acryloylic and methacryloylic groups (type a and d, Table 1), as well as diacetylenic, butadienic, vinylic and maleic acid groups (type b and c), have been used as polymerizable moieties. A compilation of amphiphilic, photopolymerizable molecules is given in Table 1. 

Out of a variety of polymerizable lipids tested for possible use of bilayer formation, only three systems exhibited BLM lifetimes of more than a few minutes (Table 2 26)). These BLMs were characterized by measuring their resistance and capacitance (Table 2., see26> for details). The data obtained were comparable with values obtained with egg lecithin the most frequently used material for preparing BLMs. 

Aqueous dispersions of polymerizable lipids and surfactants can be polymerized by UV irradiation (Fig. 18). In the case of diacetylenic lipids the transition from monomeric to polymeric bilayers can be observed visually and spectroscopically. This was first discussed by Hub, 9) and Chapman 20). As in monomolecular layers (3.2.2) short irradiation brings about the blue conformation of the poly(diacetylene) chain. In contrast, upon prolonged irradiation or upon heating blue vesicles above the phase transition temperature of the monomeric hydrated lipid the red form of the polymer is formed 23,120). The visible spectra of the red form in monolayers and liposomes are qualitatively identical (Fig. 19). 

Fig. 18. Scheme of formation of polymeric liposomes from polymerizable lipids 19-33)... 

Whether polymerized model membrane systems are too rigid for showing a phase transition strongly depends on the type of polymerizable lipid used for the preparation of the membrane. Especially in the case of diacetylenic lipids a loss of phase transi tion can be expected due to the formation of the rigid fully conjugated polymer backbone 20) (Scheme 1). This assumption is confirmed by DSC measurements with the diacetylenic sulfolipid (22). Figure 25 illustrates the phase transition behavior of (22) as a function of the polymerization time. The pure monomeric liposomes show a transition temperature of 53 °C, where they turn from the gel state into the liquid-crystalline state 24). During polymerization a decrease in phase transition enthalpy indicates a restricted mobility of the polymerized hydrocarbon core. Moreover, the phase transition eventually disappears after complete polymerization of the monomer 24). 

First, a mixture of synthetic or natural phospholipids, polymerizable lipids, and proteins can be converted to liposomes and then be polymerized. Second, polymerizable lipids are introduced into e.g. erythrocyte ghost cells by controlled hemolysis and subsequent polymerization as described by Zimmermann et al.61). This hemolysis technique is based on a reversible dielectric breakdown of the cell membrane. Dielectric breakdown provides a third possible path to the production of bi omembrane models. Zimmermann et al. could show that under certain conditions cells can be fused with other cells or liposomes61). Thus, lipids from artificial liposomes could be incorporated into a cell membrane. A fourth approach has been published by Chapman et al.20). Bacterial cells incorporate polymerizable diacetylene fatty acids into their membrane lipids. The diacetylene units can be photopolymerized in vivo. The investigations discussed in more detail below are based on approaches 1. and 3. 

What reasons are there for mixing polymerizable lipids with natural ones Polymerized membrane systems, especially those based on diacetylenic lipids, have proven to be excessively rigid and to show no phase transition. Addition of natural lipids could help to retain a certain membrane mobility even in the polymerized state, with almost unaffected stability. Furthermore, natural lipids can provide a suitable environment for the incorporation of membrane proteins into polymerizable membranes (see 4.2.3). Besides this, enzymatic hydrolysis of the natural membrane component can be used for selectively opening up a vesicle in order to release entrapped substances in a defined manner (see 4.2.2). Therefore, it is interesting to learn about the miscibility of polymerizable and natural lipids and also about the polymerization behavior of these mixtures. Investigations on this subject have thus far focused on mixtures of natural lipids with polymerizable lipids carrying diacetylene moieties. 

Provided the components are completely miscible and hexagonally packed in a mixed film below a molar ratio of 0.25 of diacetylenic lipid, each of the 6 nearest neighbors of a polymerizable lipid molecule is a nonpolymerizable natural lipid. Due to the low lateral diffusion rate in the condensed phase diacetylene polymerization should either become impossible or at least proceed at a considerably lower rate. 

In contrast, unchanged polymerization rates for all molar ratios of a polymerizable lipid indicate the formation of monomer islands and thus complete immiscibility. 

Since cholesterol is an important component of many biological membranes mixtures of polymerizable lipids with this sterol are of great interest. In mixed monolayers of natural lipids with cholesterol a pronounced condensation effect , i.e. a reduction of the mean area per molecule of phospholipid is observed68. This influence of cholesterol on diacetylenic lecithin (18, n = 12), however, is not very significant (Fig. 32). Photopolymerization indicates phase separation in this system. Apparently due to the large hydrophobic interactions between the long hydrocarbon chains of... 

The action of phospholipase A2 on mixed monolayers of natural and polymerizable lipids can be measured under constant surface pressure by the contraction of the monolayer as a function of time as depicted schematically in Fig. 39. It turns out that the chief parameter influencing the enzymatic activity is the miscibility of the lipid components and not the fact whether the film is polymerized or not. In mixed and demixed membranes the enzyme is able to hydrolyze the natural lipid component, but with considerable differences in the hydrolizing rate (Fig. 40). A pure dilauroyllecithin (DLPC) monolayer is completely hydrolyzed in a few minutes after injecting the enzyme... 

The preparation of giant liposomes79) made it possible to apply electrical field-induced fusion to the fusion of vesicles made from natural and polymerizable lipids. Since in dielectrophoresis the net force pulling vesicles towards higher field intensities is proportional to the volume of the particle, only large vesicles can be aligned in parallel to the field lines between the electrodes. 

Natural lipids used for fusion experiments were mainly phospholipids with different chain lengths and their mixtures with cholesterol. As polymerizable lipids, butadienic derivatives with a phosphatidylcholine (19) and a dimethylammonium head group (26) were used in the fusion experiments. 

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