Oleic acid, oleate

Figure 7.17 Hydrolysis of oleic anhydride catalyzed by spontaneously formed oleic acid vesicles at 40 °C, (a) during the first 3 h, and (b) during a long observation time. A vesicle suspension (10 ml in 0.2 M bicine buffer (pH 8.5)) was overlaid with 0.25 mmol oleic anhydride and 0.025 mmol oleic acid. The increase of the concentration of oleic acid/oleate is plotted as a function of reaction time. Initial concentration of oleic acid/oleate 0 mM ( ), 5 mM ( ), 10 mM (o), 20 mM ( ). For an initial oleic acid/oleate concentration of 20 mM, the concentration of oleic anhydride (A) present in the vesicles during the reaction is also plotted (b, right axis). (From Walde et al, 1994b.)...

Of all mentioned prebiotic membranogenic molecules, the ones that have gained more attention in the literature are long-chain fatty acids. In addition to their prebiotic relevance, these compounds are relatively simple from the structural point of view, and most of them are easily available. We will see in the next chapter that these vesicles have acquired a particular importance in the held of the origin of life. In fact, the hrst inveshgations on self-reproducing aqueous micelles and vesicles were carried out with caprylate (Bachmann et al, 1992) and most of the recent studies on vesicles involve vesicles from oleic acid/oleate (for simplicity we will refer to them as oleate vesicles). In this section, I would like to illustrate some of the basic properties of these surfactant aggregates. 

Figure 10.10 Transmission electron micrograph of ferritin entrapped in POPC liposomes (palmitoyloleoylphosphatidylcholine). Cryo-TEM micrographs of (a) ferritin-containing POPC liposomes prepared using the reverse-phase evaporation method, followed by a sizing down by extrusion through polycarbonate membranes with 100 nm pore diameters ([POPC] = 6.1 mM) and (b) the vesicle suspension obtained after addition of oleate to pre-formed POPC liposomes ([POPC] = 3 mM, [oleic acid - - oleate] = 3 mM). (Adapted from Berclaz et al, 2001a, b.)...

Figure 10.19 Effect of pre-added vesicles on the formation of oleic acid/oleate vesicles. Turbidity measured at 500 nm (1 cm path length) is plotted as a function of time, T=2°C. (a) 62 p,l of 80 mM aqueous sodium oleate was added to 2.438 ml of0.2Mbicine buffer, pH 8.8 ([oleic acid/oleate] = 2 mM). (b)62 p,l of 80mM aqueous sodium oleate was added to 2.438 ml of a 2 mM oleic acid/oleate 100 nm vesicle suspension (0.2 M bicine buffer, pH 8.8 ([oleic acid/oleate] = 4 mM). (c) Turbidity of 2 mM oleic acid/oleate 100 nm vesicles , (d) the same as (b), but using a 50 nm vesicle suspension, (e) Turbidity of 2 mM oleic acid/oleate 50 nm vesicles . (Modified from Blochiger etal, 1998.)...

Oleic acid/oleate vesicles containing the enzyme Q(3 replicase, the RNA template and the ribonucleotides. The water-insoluble oleic anhydride was added externally. 

Experiments have confirmed the idea that micelles as well as vesicles could grow autocatalytically (see [41] for a good overview). In a landmark paper Bachmann et al. [42] observed the formation of autocatalytically replicating micelles from sodium caprylate. The micelles could be converted into more stable vesiscles by pH change. Oleic acid/oleate vesicles can also mul-... 

Oleates Usually liquid preparations made by dissolving alkaloids in oleic acid. Oleate of Mercury, however, is an ointment-like product of mercuric oxide in oleic acid. Oleic acid was named by the pharmacist Chevreul after olives. ... 

Figure 3. Freeze-fracture electron microscopy analysis of the vesicle size distribution in the case of the spontaneous vesiculation of oleic acid/oleate. (A) Vesicles formed from the hydrolysis of 25 mM oleic anhydride (overall concentration) at 30 °C, yielding 50 mM oleic acid/oleate. (B) Vesicles extruded throughout 50 nm diameter filters. (C) Vesicles formed upon hydrolyzing 20 mM oleic anhydride (same conditions as in A) in the presence of pre-added extruded vesicles B—all in 0.2 M bicine buffer pH 8.5. For details see ref. 8.

All ingredients are present in the reaction mixture, which is added to a lipid film, and liposomes are prepared containing all macromolecules (enzymes and DNA or RNA templates) as well as all substrate molecules (nucleotides, for example). Consequently, this procedure has to be performed very quickly otherwise, the enzymatic reaction would mainly occur outside the liposomes and a distinction between product molecules synthesized inside from those produced outside and entrapped later would be difficult to draw. After the formation of liposomes, the enzymes outside the liposomes have to be inhibited by potent inhibitors (inhibitors that do their job even in the presence of substantial amounts of phospholipids) or the liposomal dispersion has to be treated by digestive enzymes. This strategy has basically been applied in the case of the RNA replication by QP replicase inside oleic acid/oleate liposomes" and in the case of the polymerase chain reaction (PCR) inside POPC or POPC/PS liposomes." In the former case, EDTA was added after the formation of the liposomes to inhibit the non-entrapped enzymes (and the kinetics was followed after addition of the EDTA molecules), in the latter case, the non-entrapped DNA template molecules were digested by DNase I before the temperature was raised to 95°C and the polymerization started. 

Several years ago, Chakrabarti et al. " described for the first time such a polymerization of RNA inside DMPC vesicles by polynucleotide phosphorylase. In these experiments, ADP was added externally to the enzyme-containing vesicles (this enzymatic reaction does not require template nucleic acids for initiation) and incubated at 23°C, the main phase transition temperature of DMPC. Because of these gel-to-liquid crystalline transitions of the bilayer, the vesicles could take up ADP from the external milieu, and the enzyme could produce poly (A). On the other hand, the enzyme could not leak out because of its size. Similar experiments were also carried out in our group by Walde et al. Here, the vesicles consisted of a single-chain amphiphile (oleic acid/oleate) that forms vesicles at a pH of about 8.0. ADP was shown to permeate across the oleic acid/oleate membrane and was incorporated inside the vesicles by polynucleotide phosphorylase to poly(A). 

K. Ruiz-Mrrazo, P. Stano, P. L. Luisi, Lysozyme effect on oleic acid/oleate vesicles, J. Lipos. Res., 2006, 16, 143-154. 

Evidence was found for a POPC matrix effect, in that the pre-added vesicles influenced the size distribution of the newly produced vesicles. The matrix effect leads to rapid formation of vesicle aggregates and control over the final size distribution, such that the mixed systems are more monodisperse than in the absence of the matrix, where the oleic acid vesicles grow in an uncontrolled way. This effect was more obvious at lower concentrations of oleic acid/oleate than at higher concentrations. 

A primary reason of interest and fascination is at the level of self-assembly and selforganization. This phenomenon, which we already know well for other structures, acquires in giant vesicles a particular dimension. One of the simplest giant vesicles was produced simply by dispersing oleic acid in water at pH 8.5 [1]. This was an oleic acid - oleate giant vesicle having a diameter of about 70 pm, as can be seen in Figure 1.1. 

In this example of spontaneous vesiculation, a total of about 10 oleic acid-oleate molecules are involved in the formation of one vesicle. It is self-assembly and self-organization in the highest form, and unprecedented in chemistry-unless we consider crystallization. It is useful to remember that this creation of order operates under thermodynamic control ordered structures are built by a process which is... 

One is the problem of reproducibility. Events witnessed with one giant vesicle are sometimes not easily observed again. For example, self-reproduction, budding and growth of oleic acid-oleate giant vesicles in one long scries of experiments [1], could be video recorded in real time, but it was not possible to repeat such behavior subsequently. A second drawback, already mentioned, is that the formation of giant vesicles seems to be restricted to a small class of surfactants. 

In contrast with phospholipid vesicles, the oleic acid-oleate vesicle system is characterized by a relatively high monomer solubility. The critical sodium oleate concentration for micelle formation (cmc) at about pH 10.5 is in the range 0.7-1.4mM [10] it has been reported that the monomer solubility at pH 7.4 is around 10-20 pM [11]. (For comparison, the monomer concentration of 1,2-dipalmitoyl-5n-glycero-3-phosphocholine (DPPC) is known to be aroimd 10 M [12].) Due to the relatively high monomer concentration, several physicochemical properties of oleic acid-oleate vesicles are different from vesicles made from phospholipids, such as phosphatidylcholines. The kinetics of lipid exchange is expected to be faster in the... 

In the present study, electron spin resonance (ESR) spectra of a fatty-acid spin probe (16-doxylstearic acid, see Figure 19.1) incorporated into an oleic acid-oleate system were measured in order to get insight into the pH-dependent aggregation properties of sodium oleate and oleic acid. Furthermore, the dilution-induced transformation of submicrometer-sized particles (micelles and/or vesicles) into giant oleic acid-oleate vesicles was investigated by electrophoretic light scattering measurements. 

Dynamic properties of oleic acid-oleate vesicles studied by ESR... 

In the case of pH 6.4, two lines were obviously overlapping at oleic acid-oleate concentrations below 15 mM. Based on the peak positions and shapes it is evident that the sharp peak corresponds to 16-DS in the aqueous medium, whereas the broader one most probably corresponds to 16-DS in bilayers (vesicles). Based on the observations that two peaks exist at pH 6.4, it is suggested that the exchange rate of the surfactant between the vesicles and the aqueous medium is slow with respect to the ESR time-scale. By increasing the concentration of oleic acid-oleate, the signal corresponding to the spin probe in the aqueous domain was reduced and the... 

Figure 19.3 ESR spectra of 16-DS in 25 mM oleic acid-oleate suspensions. The hypeifine coupling constant, aN, is plotted as a function of pH.

Based upon the above results, for the oleic acid-oleate system at intermediate pH (10.4-7.0) the scheme shown in Figure 19.5 is proposed, in which vesicles, micelles, and monomers always coexist, and the 16-DS probe molecules are distributed between vesicles, micelles, and the aqueous domain. Although the probe molecules can exchange position quickly (a rapid equilibrium between the monomers and the solubilized 16-DS), in the case of the bilayers (vesicles), the rate of exchange of 16-DS between the bilayers (vesicles) and the aqueous domain is considerably lower than for the micelles. 

Figure 19.4 Size distribution of oleic acid-oleate aggregates at pH 9.7 as determined by electrophoretic light scattering at a scattering angle of 90°.

Dilution-induced formation of oleic acid-oleate giant vesicles... 

As mentioned above, it has been found earlier [14] that extensive dilution of 80mM oleic acid-oleate suspensions prepared in 0.2 M bicine buffer (pH 8.5) increased the... 

Figure 19.5 Schematic representation of the oleic acid-oleate system at intermediate pH (10.4-7.0).

Figure 19.6 (a) Difference contrast light micrograph and (b) laser scanning confocal micrograph of giant oleic acid-oleate vesicles at pH = 8.5. 

Figure 19.7 Dilution-induced change in the size distribution of oleic acid-oleate vesicles at pH 8.5 (0.2 M bicine buffer) (a) nonextruded vesicles (b) vesicles extruded through polycarbonate membranes with a mean pore diameter of 0.2 pm.

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