Figure 7.4 Common lipid components of biological membranes

. For simplicity, all acyl chams axe shown as oleyl rescue, in vitro. It is considered the seminal work on the self-assembly of planar lipid bilayers [516,518,519,531,532]. Their research led them to the conclusion that a soap film in its final stages of thinning has a structure of a single bilayer, with the oily tails of detergent molecules pointing to the side of air, and the polar heads sandwiching a layer of water. Their experimental model drew on three centuries of observations, beginning with the work of Hooke. The membranes prepared by the method of Rudin's group became known as black lipid membranes (BLMs). Soon thereafter, vesicles with walls formed of lipid bilayers, called liposomes, were described by Bangham [533].

7.2.2 Black Lipid Membranes (BLMs)

Mueller et al. [516,531,532] described in 1961 that when a small quantity of a phospholipid (1-2% wt/vol n-alkane or squalene solution) was carefully placed over a small hole (0.5 mm diameter) in a thin sheet of teflon or polyethylene (10-25 mm thick), a thin film gradually forms at the center of the hole, with excess lipid flowing toward the perimeter (forming a "plateau-Gibbs border''). Eventually, the central film turns optically black as a single (5-nm-thick) bilayer lipid membrane (BLM) forms over the hole. Suitable lipids for the formation of a BLM are mostly isolated from natural sources, including phosphatidylcholine (PC), phosphatidyl-ethanolamine (PE), phosphatidylserine (PS), phosphatidylinositol (PI), and sphin-gomyelin (Sph). Such membranes have been viewed as useful models of the more complex natural membranes [516,532-544]. Figure 7.4 shows the most common membrane components. Sphingomyelin is an example of a broad class of sphingolipids, which include cerebrosides (carbohydrates attached to the head groups) and gangliosides (found in plasma membrane of nerve cells).

However, a serious drawback in working with BLMs as a model system is that they are extremely fragile (requiring a vibration-damping platform and a Faraday cage during measurements of electrical properties), and tedious to make [536-542]. That notwithstanding, Walter and Gutknecht [537] studied the permeation of a series of simple carboxylic acids across eggPC/decane BLMs. Intrinsic permeability coefficients, P0, were determined from tracer fluxes. A straight-line relationship was observed between log P0 and hexadecane-water partition coefficients, log Kp, for all but the smallest carboxylic acid (formic): log P0 = 0.90 log Kp + 0.87. Using a similar BLM system, Xiang and Anderson [538] studied the pH-dependent transport of a series of a-methylene-substituted homologs of p-toluic acid. They compared the eggPC/decane permeabilities to partition coefficients determined in octanol-, hexadecane-, hexadecene-, and 1,9-decadiene-water systems. The lowest correlation was found from comparisons to the octanol-water scale. With the hexadecane-water system, log P0 = 0.85 log Kp - 0.64 (r2 0.998), and with decadiene-water system, log P0 = 0.99 log Kp - 0.17 (r2 0.996). Corrections for the unstirred water layer were key to these analyses. Figure 7.5 shows the linear correlation between the logarithms of the permeability coefficients and the partition coefficients for the five lipid systems mentioned above.



Figure 7.5 Intrinsic permeabilities of ionizable acids versus oil-water partition coefficients.

7.2.3 Microfilters as Supports

Efforts to overcome the limitations of the fragile membranes (as delicate as soap bubbles) have evolved with the use of membrane supports, such as polycarbonate filters (straight-through pores) [543] or other more porous microfilters (sponge-like pore structure) [545-548].

Thompson et al. [543] explored the use of polycarbonate filters, and performed experiments to make the case that just single bilayer membranes formed in each of the straight-through pores. Several possible pore-filling situations were considered: lipid-solvent plug, lipid-solvent plug plus BLM, multilamellar BLM, and unila-mellar BLM. The key experiment in support of a single-bilayer disposition involved the use of amphotericin B (Fig. 7.6), which is an amphiphilic polyene zwitterionic molecule, not prone to permeate bilayers, but putatively forming tubular membrane-spanning oligomers if the molecules are first introduced from both sides of a bilayer, as indicated schematically in Fig. 7.6. Once a transmembrane oligomer forms, small ions, such as Na+ or K+, are able to permeate through the pore formed. The interpretation of the voltage-current curves measured supported such a single-bilayer membrane structure when polycarbonate microfilters are used as a scaffold support.

Cools and Janssen [545] studied the effect of background salt on the permeability of warfarin through octanol-impregnated membranes (Millipore ultrafiltration filters, VSWP, 0.025-p.m pores). At a pH where warfarin was in its ionized form, it was found that increasing background salt increased permeability (Fig. 7.7). This

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