Homogenizer liposomes preparation

Weder, H. G., and Zumbuehl, O. (1984). The preparation of variably sized homogeneous liposomes for laboratory, clinical and... 

The dialysis method is well suited for the encapsulation of lipophilic drugs. The liposomes prepared by this method are usually homogeneous in size with good reproducibility, and the encapsulation condition is mild compared with other methods. The drawbacks include (i) low entrapment efLciency for hydrophilic molecules, (ii) the complete removal of residual detergent is impossible, (iii) the procedure is lengthy, and (iv) scale-up is difLcult. 

One of the major drawbacks of liposomes is related to their preparation methods [3,4]. Liposomes for topical delivery are prepared by the same classic methods widely described in the literature for preparation of these vesicles. The majority of the liposome preparation methods are complicated multistep processes. These methods include hydration of a dry lipid film, emulsification, reverse phase evaporation, freeze thaw processes, and solvent injection. Liposome preparation is followed by homogenization and separation of unentrapped drug by centrifugation, gel filtration, or dialysis. These techniques suffer from one or more drawbacks such as the use of solvents (sometimes pharmaceutically unacceptable), an additional sizing process to control the size distribution of final products (sonication, extrusion), multiple-step entrapment procedure for preparing drug-containing liposomes, and the need for special equipment. 

Liposome preparations have been shown to be suitable not only for studying special drug-membrane interaction effects in vitro but also for use as drug carriers. Various techniques have been developed and described to prepare homogeneous unil-... 

Brandi, M.M. Bachmann, D. Drechsler, M. Liposome preparation using high-pressure homogenizers. In Liposome Technology, 2nd Ed. Gregoriadis, G., Ed. CRC Press Boca Raton, 1993 1, 49-65. 

Liposomes made of pure phospholipids will not form at temperatures below T of the phospholipid. This temperature requirement is reduced to some extent, but not eliminated, by the addition of cholesterol (17). In some cases, it is recommended that liposome preparation be carried out at temperatures well above T of the vesicles. For instance, in the case of vesicles con-taining dipalmitoyl phosphatidylcholine (DPPC, T = 41°C), it has been suggested that the liposome preparation procedure be carried out at 10°C higher than the T at 51°C (18, 19). This is in order to make sure that all the phospholipids are dissolved in the suspension medium homogenously and have sufficient flexibility to align themselves in the structure of lipid vesicles. Following termination of the preparation procedure, usually nanoliposomes are allowed to anneal and stabilize for certain periods of time (e.g. 30-60 min), at a temperature above T, before storage. 

The following protocol, which is derived from the one-step liposome preparation technique, originally described in (12) is suited. A slurry is made from a single phospholipid or a freeze-dried cake of lipid-blend and buffer (drug/marker solution). The slurry is processed using an APV MicronLab 40 lab-scale homogenizers. A micro-fluidizer Ml 10 may be used as well. 

Brandi M, Bachmann D, Drechsler M, Bauer KH (1990) Liposome preparation by a new high pressure homogenizer GauUn Micron LAB 40. Drug Dev Ind Pharm 16(14) 2167-2191... 

The most commonly used method of liposome preparation (see Note 1) involves hydration of a lipid mixture in buffer, followed by extrusion through a press of some description. A number of devices are commercially available for this purpose, the most widely used being the Extruder from Lipex Biomembranes. Homogenous lipid mixtures can be prepared by drying the... 

Create liposomal vesicles using any established method (see Section 1) by mixing the lipid mixture with degassed, nitrogen-purged 10 mM Hepes, 0.15 M NaCl, pH 7, to obtain a final concentration of 5 mg/ml lipid in the aqueous buffer. Sonicate to emulsify the liposomal preparation. Remove diethylether by vacuum evaporation. Periodically mix by vortexing to maintain a homogeneous suspension of liposomes. 

When liposomes are prepared from a molecular mixture of lipid components it is important that all lipids be homogeneously dissolved in an organic solvent in order to obteiin bilayers with evenly distributed lipids after hydration. For example, the solubilities of phosphatidylcholine and cholesterol in chloroform are similar their solubility in benzene differs. Upon removal of benzene from the lipid solution an inhomogeneous lipid film is formed on the glass wall and... 

Schwendener, R. A., Asanger, M., and Weder, H. G. (1981). n-Alkylglucosides as detergents for the preparation of highly homogeneous bilayer liposomes of variable sizes (60-240 (p) applying defined rates of detergent removal by dialysis, Biochem. Biophys. Res. Commun.. 100, 1055-1062. 

Liposomes are formed due to the amphiphilic character of lipids which assemble into bilayers by the force of hydrophobic interaction. Similar assemblies of lipids form microspheres when neutral lipids, such as triglycerides, are dispersed with phospholipids. Liposomes are conventionally classified into three groups by their morphology, i.e., multilamellar vesicle (MLV), small unilamellar vesicle (SUV), and large unilamellar vesicle (LUV). This classification of liposomes is useful when liposomes are used as models for biomembranes. However, when liposomes are used as capsules for drugs, size and homogeneity of the liposomes are more important than the number of lamellars in a liposome. Therefore, "sized" liposomes are preferred. These are prepared by extrusion through a polycarbonate... 

LPDI nanoparticles are homogenous, self-forming spheres between 100 and 200 nm in diameter that are formed from the spontaneous rearrangement of a lipid bilayer around a polycation condensed DNA core. The LPDI particles (lipopolyplexes) have benefits over lipoplexes, which are composed of liposomes and DNA. Homogenous particles are formed during preparation and thus allow a more consistent production of particles, as required by the FDA for clinical use. The LPDI particles also have a lower toxicity associated with them as opposed to lipoplexes, which can generate severe systemic inflammatory responses, most likely to the increased DNA content on the surface of the particles. The internalization of DNA inside the LPDI also has a benefit of DNA protection. The DNA is not nearly as accessible to nuclease attack and mechanical stress. Therefore, a lower quantity of DNA is used because it is protected inside of the LPDI for delivery. 

Cationic lipids cannot be dissolved in water and form aggregates in aqueous solution, such as bilayers. To prepare a homogeneous reagent, in most cases liposomes were made from cationic lipids in a first step. When it is not possible to form stable lipid bilayers (i.e., liposomes) using a single lipid, then it may be necessary to combine the cationic lipid with one or more so-called helper lipids like cholesterol (Choi) (41) or 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine (DOPE) (42). 

A number of factors for DOTAP-cholesterol/DNA complex preparation including the DNA/liposome ratio, mild sonication, heating, and extrusion were found to be crucial for improved systemic delivery maximal gene expression was obtained when a homogeneous population of DNA/liposome complexes (200-450 nm) was used. Cryoelectron microscopy showed that the DNA was condensed on the interior of liposomes between two lipid bilayers in these formulations, a factor that was thought to be responsible for the high transfection efficiency in vivo and for the broad tissue distribution (150). 

Schwendener, R. A. (1986). The preparation of large volumes of homogeneous, sterile liposomes containing various lipophilic cytostatic drugs by the use of a capillary dialyBancer Drug Deliv., 3, 123-129. 

The methods for preparation of niosomes are similar and as complicated as those used for liposomes. One of the most frequently utilized techniques consists of the hydration of a mixture of the surfactant-lipid at elevated temperature followed by optional size reduction (by sonication, extrusion, homogenization, etc.) to obtain a homogeneous colloidal dispersion and separation of the unentrapped drug [36,40,41,52,55],... 

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