Dehydrated lipids

Proposed more than 20 years ago, the stalk intermediate—a highly curved lipid stmcture— provides the most plausible description of the initial fusion stage currently available. The related stalk-pore mechanism (23-25) of fusion is viewed favorably by most researchers. It shows the close relation between fusion and the transition from lamellar into bilayer cubic and hexagonal phases (see Fig. 4 in the section entitled Formation of nonlamellar phases in membrane lipids ). Studies on the rhombohedral phase formed in partially dehydrated lipids provide another insight into the possible structure of fusion stalks (26). 

Madin, K.A.C. Crowe, J.H. (1975). Anhydrobiosis in nematodes carbohydrate and lipid metabolism during dehydration. Journal of Experimental Zoology, 193, 335-41. 

A method resulting in improved encapsulation of aqueous phase by MLV is the so-called dehydration-rehydration procedure (Kirby and Gregoriadis, 1984 Shew and Deamer, 1985). The lipid (usually preformed liposomes) is dried (by either lyophilization or evaporation) in the presence of the aqueous solute to be entrapped, thus forming a mixed film with solute trapped between layers. Subsequent gradual rehydration with a minimum of aqueous phase leads to the formation of MLV with a high entrapment of the aqueous solutes added. 

Figure 1.7 Typical zero-order and corresponding second-derivative electronic absorption spectra of ethanol-reconstituted lipid/chloroform extracts of autoxidized model polyunsaturated fatty-acid compounds and inflammatory synovial fluid obtained after (1) reduction with NaBH4 and (2) dehydration with alcoholic H2S04- (a) Methyl linoleate subsequent to autoxidation in air at ambient temperature for a period of 72 h (—), or exposure to a Fenton reaction system containing EDTA (5.75 x 10 mol/dm ), H2O2 (1.14 X 10 mol/dm ) and Fe(ll) (5.75 x IO mol/dm ) as an aqueous suspension (—) (b) as (a) but with methyl linolenate (c) untreated rheumatoid knee-joint synovial fluid.

Several processes often occur in lipids, including oxidation, hydration, dehydration, decarboxylation, esterification, aromatization, hydrolysis, hydrogenation and polymerization. In fact, the chemistry of these materials can be affected, for example, by heat (anthropogenic transformations), humidity, pH, and microbial attacks. 

Harrigan, P.R., Madden, T.D., and Cullis, P.R. (1990) Protection of liposomes during dehydration or freezing. Chem. Phys. Lipids 52, 139-149. 

Foradada and Estelrich [3.63] studied the encapsulation of thioguanine (TG) in three types of liposomes produced by extrusion, ethanol injection and dehydration-rehydration vesicles. The entrapment has been examined at three different concentrations (1, 0.1 and 0.01 mM) and at three different pH values (4.7,7.4 and 9.2). The dehydration-rehydration vesicles were found to be the optimum method to encapsulate TG, independent of the pH value. At pH 4.7, 12 mmol/mol of lipid were entrapped, while with the other methods a maximum of 3 mmol/mol of lipid has been achieved. The authors related this behavior to the formation of hydrogen bridges between the TG and the liposomes. 

Castner cell, 22 767, 768-769, 772 Cast nickel-beryllium alloys, 3 657t, 658t Castor oil, 2 82 9 143 10 817, 820-822 cosmetically useful lipid, 7 833t in cosmetic molded sticks, 7 840t in defoamer formulations, 3 237-238 dehydrated, 9 150... 

To obtain tissue preparations whose constituents were maintained as closely as possible to their state in vivo, the material had to be fixed, i.e. the enzymes inactivated so that cell structures were instantaneously preserved, an almost unattainable ideal. Formalin was the favored fixative, but others (e.g. picric acid), were also employed. Different methods of fixation caused sections to have different appearances. Further artifacts were introduced because of the need to dehydrate the preparations so that they could be stained by dyes, many of which were lipid-soluble organic molecules. Paraffin wax was used to impregnate the fixed, dehydrated material. The block of tissue was then sectioned, originally by hand with a cut-throat razor, and later by a mechanical microtome. The sections were stained and mounted in balsam for examination. Hematoxylin (basophilic) and eosin (acidophilic) (H and E staining) were the commonest stains, giving blue nuclei and pink cytoplasm. Eosinophils in the blood were recognized in this way. 

The continuous availability of trillions of independent microreactors greatly multiplied the initial mixture of extraterrestrial organics and hydrothermal vent-produced chemicals into a rich variety of adsorbed and transformed materials, including lipids, amphiphiles, chiral metal complexes, amino add polymers, and nudeo-tide bases. Production and chiral amplification of polypeptides and other polymeric molecules would be induced by exposure of absorbed amino adds and organics to dehydration/rehydration cydes promoted by heat-flows beneath a sea-level hydro-thermal field or by sporadic subaerial exposure of near-shore vents and surfaces. In this environment the e.e. of chiral amino adds could have provided the ligands required for any metal centers capable of catalyzing enantiomeric dominance. The auto-amplification of a small e.e. of i-amino adds, whether extraterrestrially delivered or fluctuationally induced, thus becomes conceptually reasonable. 

Jansson et al. [189] used the conventional approach of blending the solid particles with solvent after which an aliquot was taken to determine the volatile compounds (e.g., phenols and chlorobenzenes). A second fraction was taken after the lipid removal for determination of compounds sensitive to concentrated sulfuric acid. The bulk lipids were removed by oxidative dehydration with Si02 /H2S04 and further cleaned-up with GPC. The chloroparaffins were isolated at this stage. Separation on silica isolated the OCPs, and the organochlorines and organobromines were finally fractionated on active charcoal. 

The dehydration-rehydration procedure is characterized by its mildness and is thus compatible with most labile materials. The amounts of lipids and vaccine materials described below are typical for the preparations made for animal work described here but could be scaled up or down depending on the number of animals and the amount of antigens used. 

Entrapment of plasmid DNA and/or protein into liposomes entails the preparation of a lipid film from which multilamellar vesicles and, eventually, small unilamellar vesicles (SUVs) are produced. SUVs are then mixed with the plasmid DNA and/or protein destined for entrapment and dehydrated. The dry cake is subsequently broken up and rehydrated to generate multilamellar dehydration-rehydration vesicles (DRV) containing the plasmid DNA and/or protein. On centrifugation, liposome-entrapped vaccines are separated from nonentrapped materials. When required, the DRV are reduced in size by microfluidization in the presence or absence of nonentrapped materials or by employing an alternative method (7) of DRV production, which utilizes sucrose (see below). 

Quantitative entrapment of vaccines into small (up to about 200 nm diameter) liposomes in the absence of microfluidization (which can damage DNA and other labile materials when extensive) can be carried out by a novel one-step method (7) as follows SUVs (e.g., cationic) prepared as in section Preparation of Small Unilamellar Vesicles are mixed with sucrose to give a range of sucrose-to-lipid weight/weight ratio of 1.0 to 5.0 and the appropriate amount of plasmid DNA (e.g., 10-500 pg) and/or protein (e.g., up to 1 mg). The mixture is then rapidly frozen and subjected to dehydration by freeze-drying, followed by rehydration as in section Preparation of Vaccine-Containing Dehydration-Rehydration Vesicles. ... 

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