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consist of an aqueous core entrapped by one or more bilayers of natural and/or synthetic lipids. The size of a liposome ranges from some 20 nm up to several micrometers and they may be composed of one or several concentric membranes, each with a thickness of about 4 nm. Liposomes possess unique properties owing to the amphiphilic character of the lipids, which make them suitable for drug delivery. Drugs with widely varying lipophilicities can be encapsulated in liposomes either in the phospholipid bilayer, in the entrapped aqueous core or at the bilayer interface. Reformulation of drugs in liposomes has provided an opportunity to enhance the therapeutic indices of various agents mainly through the alteration of biodistribution. They are versatile drug carriers, which can be used to control retention of entrapped drugs in the presence of biological fluids, controlled vesicle residence in the systemic circulation or other compartments in the body and enhanced vesicle uptake by target cells. Liposomes composed of natural lipids are biodegradable, biologically inert, weakly immunogenic, produce no antigenic or pyrogenic reactions and possess limited intrinsic toxicity. Therefore, drugs encapsulated in liposomes are expected to be transported without rapid degradation and minimum side effects to the recipients. Because of their size, liposomes display some unique pharmacokinetic characteristics. Size, lamellarity, bilayer rigidity, charge and bilayer surface modifications determine the fate of liposomes on the shelf and in vivo. These include clearance via the reticuloendothelial system, which results in a relatively long systemic circulation time, and hepatic and splenic distribution. Furthermore, liposomes exhibit preferential extravasation and accumulation at the site of solid tumors due to increased endothelial permeability and reduced lymphatic drainage in these tissues, which has been defined as enhanced permeability and retention effect . Liposomal delivery is therefore a means to modify the pharmacokinetic and pharmacodynamic properties of therapeutic agents. Such modifications can, in some settings, improve the therapeutic efficacy of anticancer drugs and reduce or modulate their toxicity profile. For example, long circulating polyethylene glycol-coated liposomal formulation of doxorubicin has been shown to exhibit increased solid tumor accumulation due to the enhanced permeability and retention effect and decreased dose-limiting cardiac toxicity relative to the free drug Types of phospholipids used in the preparation of liposomes A phospholipid has two acyl chains linked to a headgroup by means of a glycerol-backbone. Figure shows the structural formula of a phospholipid, where R1 and R2 are saturated or unsaturated acyl chains and R3 is the polar head group. The polar head groups are used for classification, i.e. to distinguish between different phospholipids.
Figure The general structure of a phospholipid and the structure of Egg PC, DSPC(distearoyl PC) and DOPE(dioleoyl PC). Phosphatidylcholines or PC-lipids and PEs are the most widely used lipids in liposome work. PC-lipids are zwitterionic at all relevant pH1 and can therefore form lamellar structures independently of the pH in the solution. Purified egg PC, egg PG, soy PC, hydrogenated soy PC and sphingomyelin, phosphatidic acid , phosphatidyl inositol are natural phospholipids.Liposomes with PC, PG, PI and PA are negatively charged. Regarding synthetic phospholipid, DDPC, DLPC, DMPC, DPPC, DSPC, DOPC, POPC and DEPC are available for PC series. DMPG, DPPG, DSPG, POPG are available for PG series. DMPA, DPPA and DSPA are available for PA (phosphatidic acid) series. DMPE, DPPE, DSPE and DOPE are available for PE series. DOPS is available for PS series in stock. PEG or polyglycerin attached phospholipid (PEG phospholipid) are available for long circulating liposome formulation. Activated functional group attached phospholipid either PEG end or non PEG end are also available for immunoliposome or targeting use liposome.
These long-circulating liposomes may also act as a reservoir for prolonged release of a therapeutic agent. POPC and DDPC are recommended for nasal delivery. As liposomes can solubilise lipophilic compounds. These include the development of remote drug loading methodologies based on pH or ionic gradient. large unilamellar vesicles (LUV) and large multilamellar vesicles (several bilayers oriented concentrically around an aqueous core) (MLV) or multivesicular vesicles (MVV) are differentiated. The introduction of positively or negatively charged lipids provides the liposomes with a surface charge. it is clear that liposomes provide an extremely flexible drug carrier modality with many potential applications and an impressive track record as a carrier system. while the unsaturated phosphatidylcholine species from natural sources (egg or soy bean phosphatidylcholine) give much more permeable and less stable bilayers. then ‘fragile’ liposomes with ‘fluid’ bilayers should be selected. Liposomes–Applications Liposomes can protect a drug against degradation (for example metabolic degradation). MLVs. For example. TargetingwithLiposomes Many approaches have been attempted to achieve targetable properties. glycoprotein bearing liposomes and natural and synthetic glycolipid containing liposomes. temperature sensitive liposomes for burst release in response to hyperthermia . If a fast pharmacological response isdesired. POPC and POPG are recommended for pulmonary delivery while DOPC. this solubilising potential can be used to inject poorly water-soluble. LUVs. Liposomeswith‘Homing’Devices . Conversely. Their characteristics depend on the manufacturing protocol and choice of bilayer components. Based on activity Development of liposomes as a drug carrier has been marked by a number of key innovations. DPPC. liposomes can protect the patient against side effects of the encapsulated drug. the choice of bilayer components determines the ‘rigidity’ (or ‘fluidity’) and the charge of the bilayer.For non injectable use. pH-sensitive liposomes for cytosolic drug delivery . ‘Stealth’ liposomes Coating liposomes with PEG reduces the rate of uptake by macrophages (‘stealth’ effect) and leads to a prolonged presence of liposomes in the circulation and consequently ample time for these liposomes to escape from the circulation through a leaky endothelium. as radiopharmaceuticals and radiodiagnostic carriers. Small unilamellar vesicles(meaning only one bilayer surrounds an aqueous core) (SUV). SUVs show a diameter of 20 to approximately 100 nm. including noncovalent association of cell specific antibodies with liposomes . Considering this list of applications and the existing literature. saturated phospholipids with long acyl chains such as dipalmitoylphosphatidylcholine form a rigid. In addition. covalent attachment of poly and monoclonal antibodies to the liposomes. and targeted liposomes for selective delivery to tumor cells or endothelium. rather impermeable bilayer structure. and MVVs range in size from a few hundred nanometers to several microns. What are the Different Types of Liposomes and How Are They Classified? Based on structure Liposomes are often distinguished according to their number of lamellae and size. Targeted liposomes can be used in cancer therapy. coating of liposomes with heat aggregated immunoglobulins M (IgM) . cationic liposomes for nucleic acid delivery. polyethylene glycol-coated long circulating liposomes . in delivery of drugs to lungs and in HIV treatment. lipophilic compounds intravenously.
ease of preparation.but other homing devices have been considered as well. compatibility with regulatory agencies. chemotherapies with limited biphasic solubility (e.g. reports have appeared on arginineglycine. However. polyethyleneglycol (PEG) immunoliposome attached antibodies at the distal end of PEG chain. in which the concentration in the liposome is directly proportional to the external concentration of the solution .e. The low trapping efficiencies of SUV systems largely stem from their low trapped volume (0. some important drugs exhibit significant lipophilic character which can result in rapid leakage from liposomal systems. plasmid. For example. In addition. the liposomal formulation is prepared with the chemotherapeutic agent of choice in the incubation mixture. Therefore. LUV or MLV) required for in vivo applications. Lipophilic and ampiphilic cytotoxic drugs. The free.An important consideration is how to make liposome uptake tissue or cell-specific. This prodrug is transferred selectively from the cellbound immunoliposomes into the target cell. Lipid molecules have to be introduced into an aqueous environment for the preparation of liposomes independent of liposome size and structure. the so called pendant type immunoliposomes. cost efficiency. Targeted liposomes should have ready access to the target site and should not be taken up by macrophages before encountering their target tissue or cells.aspartic acid (RGD) peptide-driven targeting of liposomes to endothelial cells in order to block angiogenesis. The presence of free PEG does not interfere with the binding of the terminally linked antibody to the antigen. drug/lipid ratio. 6-Mercaptopurine) are poorly incorporated into the liposome aqueous and lipid compartments . Another important consideration is the behaviour of liposomes in vivo which is very sensitive to vesicle size and composition. Liposome in gene delivery Cationic liposomes are considered to be a potential non-viral human gene delivery system. leading to improved trapping efficiencies.” is a more efficient and preferable strategy for liposomal drug loading. Another effective strategy is based on the selective binding of immunoliposomes that contain a lipophilic prodrug. and doxorubicin (ampiphilic). for example. it leaks into the cytoplasm. is not encapsulated in liposomes but complexed with cationic lipids by electrostatic interactions. Selection of an encapsulation protocol is largely dictated by concerns such as encapsulation efficiency. nowadays. In this method. In passive loading or entrapment. the prodrug is converted in the lysosome into the active drug. as lipid molecules themselves are poorly soluble in aqueous compartments. as well as liposome and drug stability. Plasmid liposome complexes are thought to enter the cells through fusion with the plasma or endosome membrane Techniques for Encapsulating Drugs in liposome vesicles The technique of drug entrapment must satisfy demands such as high trapping efficiency and reasonably long retention times. i. plasminogen-coated liposomes were designed to reach fibrin clots specifically (plasminogen has an affinity for fibrin) in order to deliver fibrinolytics. LUVs and MLVs can exhibit higher values (l-30 μl/Mmol lipid) and also can be prepared at higher lipid concentrations. Trapping efficiencies vary dramatically. drug loading through passive entrapment is less efficient for water soluble drugs. sterility. ranging from 1% or less for SUVs to as high as 88% for some MLVs.. this property is often limited by the type of vesicles (SUV.2-0. Although maximum trapping efficiencies are obviously desirable. such as methotrexate and cytarabine. for example the use of saccharide antennae (including galactose) to direct liposomes to hepatocytes. The negatively charged genetic material. Several ways of treating the lipids are known to support the hydration of these molecules.8 μl/mol lipid). Saccharide-directed targeting has also been described. More recently. These liposomes are usually composed of cationic lipid derivatives and a neutral phospholipid such as dioleoylphosphatidyl ethanolamine (DOPE). such as paclitaxel (highly lipophilic). also termed “remote loading. As expected. Liposomes can be loaded with drug either passively or actively. ease of scaleup. are loaded somewhat more efficiently because they partition stably in both the lipid membrane and internal compartment of the liposomes. stealth technology is often combined with attachment of a homing device to the terminal end of the PEG chain that is exposed to the aqueous medium leading to the formation of long-circulating immunoliposomes. Subsequently. nonentrapped drug is then washed away by gel-filtration or other dialysis method. Drug is then encapsulated into the liposome as the nanoparticle is formed. drug retention. Most work on liposome targeting has focused on antibodies or antibody fragments attached to the surface. Active entrapment. From the lysosome. a pH or ion gradient is created. which efficiently .
However. hydration with agitation. many different variations of this method have been developed differing in the organic solvents used for lipid solubilization. Passive trapping techniques Mechanical Methods Preparation by Film Methods. anionic. freeze-thawing and high pressure extrusion. and the way of film rehydration. The simplicity of this procedure has attracted widespread use. which range from 1% to 5%. In the end. which converts an MLV dispersion to an opalescent solution of vesicles ranging in size from about 0. this approach appears inappropriate for entrapment of many proteins and DNA due to ultrasonic degradation of these biological materials. A. Since then. Advantages of sonicated SUVs are that small homogeneous populations of vesicles are rapidly formed and virtually any lipid composition can be employed. Properties of lipid formulations can vary depending on the composition (cationic. In addition. such as doxorubicin and vincristine.05 pm. there exist several drawbacks in the technique when applied to drug encapsulation. Drug entrapment in SUVs SUVs have traditionally been prepared by sonication. with stable retention. The general steps of the procedure are formation of thin film of the lipids in dry state from organic solvents ( by rotary evaporator or hand-shaking) for hydration. the same preparation method can be used for all lipid vesicles regardless of composition. and neutral lipid species). These include the low trapping efficiencies obtained. . However. and sizing to a homogeneous distribution of vesicles by vortexing. efficiency and ability to actively load a liposomal nanoparticle depends on the individual chemotherapeutic drug characteristics and reaction conditions. the way of lipid drying.02 to 0. sonication.drives a molecule of choice across the lipid membrane leading to up to 100% loading efficiency of chemotherapeutic agents.
reduced pressure induces the spontaneous formation of LUVs. therefore. Although this technique was used initially to produce LUVs. Numerous techniques. degradation of proteins and DNA is negligible.An important feature of the French press vesicle (FPV) technique is that proteins do not appear to be significantly affected during the process as they are in sonication . Upon addition of water to the dehydrated material. Drug entrapment in ML Vs Since the efflux of water-soluble drugs from liposomes is often dictated by their membrane permeability. An interesting observation is that FPVs appear to retain entrapped solutes significantly longer than do SUVs produced by sonication or detergent removal. Encapsulation efficiencies approaching 50% for MLVs can also be obtained by dehydration-rehydration procedures where SUVs are dried in the presence of the desired agent by lyophilization or other methods. In addition. A disadvantage of solvent based liposomes is that some macromolecules are deactivated by this procedure .056 to 0. Drug entrapment in LUVs Solvent evaporation/vaporization techniques have gamed widespread popularity for entrapping drugs in LUVs. An attractive feature of this technique is that vesicles composed of virtually any lipid composition which adopts liquid crystalline lamellar systems can be obtained within a very short period of time (<30 min). These procedures have been shown to yield trapping efficiencies between 30 and 45% for a wide range of solutes including proteins. Under appropriate conditions. These increased trapping efficiencies are achieved by increasing the aqueous trapped volume of the vesicles and utilizing high lipid concentrations. have been developed to yield multilamellar preparations which entrap higher proportions of the drug. However.( membrane extrusion procedure) These pressures allow the preparation of homogeneous LUVs with average diameters ranging from 0. DNA and RNA.Under appropriate conditions. MLVs can exhibit maximal drug retention due to the number of lamellae the agent must cross to reach the vesicle exterior. These vesicles exhibit improved trapping efficiencies over sonicated systems and the average diameter of the liposomes can be controlled to some extent by varying the initial detergent/lipid ratio. More recently. they produce MLVs which exhibit stable solute retention . MLVs can be produced employing solvent evaporation techniques.2 pm. Important features of the freeze thaw and dehydrate-rehydrate processes. forming MLVs which entrap a significant amount of the original solutes. extrusion renders the preparation sterile and scale-up would appear to be straightforward. Major problems typically experienced with MLVs are the low aqueous trapped volumes and trapping efficiencies obtained for traditional MLV dispersions. SUVs can also be produced employing detergent removal techniques . the vesicles display optimal retention of entrapped solutes. studies have shown the multiple freeze-thaw steps completed directly on MLV dispersions induce a remarkable morphological change in the liposomes and results in a 10-fold or greater increase in the aqueous trapped volumes. Recent reports have described the production of homogeneously sized LUVs by extrusion of MLVs through polycarbonate filters of defined pore size under moderate pressures (2800 lb/in*). the multilayers swell. in addition to the large trapping efficiencies are that they appear applicable to a broad spectrum of biologically active agents. under proper conditions MLVs can be prepared in this manner to efficiently encapsulated numerous solutes without significantly affecting the biological activity of more sensitive agents such as DNA and proteins. Lipids dissolved in organic solvent are mixed with an aqueous phase (containing the agent to be trapped) and removal of the solvent by heat and/or. The resulting preparations tend to be quite heterogeneous in size and the extent of multilamellar character appears to vary for different procedures.05 μm (depending on lipid composition) by passing a lipid suspension under extreme pressures through a small outlet orifice. problems arise due to difficulties in removing residual detergent The French pressure cell produces fairly homogeneous SUVs of a size 0. Trapping efficiencies up to 25% can be achieved although this value decreases somewhat with increasing molecular weight of the trapped solute . Freezing and thawing SUVs results in a dramatic increase in vesicle size and trapped volume.
Stable liposomes can be prepared with less than 100% entrapment efficiency from pro-liposomes ( dried lipid film formation on finely divided particulate solids like sodium chloride. Removal of solvent film results in unilamellar vesicle with high entrapment efficiency.000 psi) through a 5 μm orifice where two streams of liquids collide each other at right angles at high velocity. SPLV is different from MLV-REVs in that they lack a large aqueous core . Methods Based on Detergent Removal In this group of liposome preparation procedures. and extrusion methods are all based on preformed vesicles. are replaced by an aqueous solution.0 to pH 11. Stable plurilamellar vesiclesThese vesicles are formed by water-in-organic phase dispersion with an excess of lipid followed by drying under continued bath sonication with an intermittent stream of nitrogen. thus forming liposomes with high encapsulation rates of hydrophilic as well as lipid phase soluble substances. Microemulsification methodMicro fluidizer is used to prepare small MLVs from concentrated lipid dispersion. In addition. Methods Based on Replacement of Organic Solvents by Aqueous Media The liposome preparation methods described in this section have in common that organic solvents. This replacement is either performed by injection of the lipid carrying organic solution into the aqueous phase—the injection methods—or by stepwise addition of aqueous phase to the organic phase. Freeze thawing. Compositionally homogeneous dispersions of liposomes are formed by sudden precipitation of lipid mixture in aqueous buffer and fast and efficient removal of organic solvent( not necessarily a volatile one). they do not require organic solvents or detergents and they display extended stabilities by storage in the frozen or dried state. such as bile salts or alkylglycosides.characteristics. There is equilibrium between the detergent molecules in the aqueous phase and the lipid . detergents. It pumps the fluid at very high pressure (10. freeze drying. the emulsification methods. namely. the majority of entrapped aqueous medium being located in the compartment in between adjacent lamellae. are used for the solubilization of lipids in micellar systems. This process is very suitable for encapsulation of water-soluble materials and for samples with 20% of lipids in the formulation and the encapsulation efficiency is 70%. sorbitol or other polysaccharides. are based on the replacement of a water-immiscible solvent by an aqueous phase. Reverse phase evapopration vesicles The droplets are formed by bath sonication of the mixture of the two phases and drying of the emulsion into a semisolid gel in a rotary evaporator under reduced pressure. In contrast to lipids. detergents are highly soluble in both aqueous and organic media. they are relatively simple. Mechanical agitation later allows collapse of water droplets and lipid monolayers of adjacent vesicles form the outer leaflet of the bilayer of a large unilamellar vesicle. The two aqueous compartments are separated by a pair of phospholipid monolayers across a thin film of organic solvent. Rapid solvent exchange vesicles The lipid mixture is quickly transferred between an essentially pure solvent environment and a pure aqueous environment. in particular ethanol—the proliposome-liposome method.5-3. pH induced vesiculation This method is used to transform MLVs to Luvs using a change in the pH of the dispersion (from pH 2. either water miscible or immiscible. Double emulsion vesicles The organic solution containing water droplet is introduced into excess aqueous medium by mechanical dispersion forming w/o/w emulsion and forming multicompartment vesicles .0) for a very short period of < 2minsthus avoiding the use of sonication or high pressure. the reverse-phase evaporation method and the double emulsion technique.
Alterations in the lipid composition of liposomes have been used to enhance the encapsulation efficiency and decrease the release rates of these agents through ionic interactions between the drug and charged lipid components. reduced leakage of the encapsulated compounds. Active trapping techniques At one extreme are agents which are virtually insoluble in water and can be incorporated into the lipid bilayer during vesicle formation. Characterisation of liposomes The liposomes produced by different techniques have different physicochemical characteristics which may influence their in vitro properties such as sterility. liposome formulations for this class of drugs vary dramatically from one agent to the next.environment of the micelle. At the other extreme are watersoluble materials which interact with the polar headgroup of phospholipids and are sequestered. Unilamellar vesicles are formed spontaneously on removal of detergent from preformed mixed micelles containing phospholipid. the most frequently applied method for membrane protein reconstitution involves the cosolubilization of membrane proteins and phospholipids. immunoliposomes virosomes and cationic liposomes. shape. These materials are generally treated as lipids themselves. B. stability as well as in vivo properties like PK and PD. To date. If the analytical procedure cannot distinguish the . and the lipids used. particularly with cardiolipin. phase response and transitional behavior . this property poses a major difficulty. surface morphology or topology. thin-layer chromatography (TLC)) when the analytical procedure can distinguish the desired lipids from possible impurities. The size and shape of the resulting vesicles are depending on the chemical nature of the detergent. Ficoll density gradient. vesicle fusion measurements and drug release profiles. Drug loading in response to ion gradients offers a more general means for encapsulating agents which are amphiphilic cations. An understanding of phase transitions and fluidity of phospholipid membranes is important in the manufacture and application of liposomes. Since a large number of commonly used drugs are amphiphilic molecules. This positively charged anthracycline derivative interacts with negatively charged phospholipids. For synthetic lipids such as DMPC and semisynthetic lipids. Biological evaluation helps in determining safety and efficacy of the preparation in vivo. lamellarity. since the phase behaviour of a liposomal membrane determines such properties as permeability. The amount of hydrophobic drug that can be introduced in a liposome is therefore highly dependent on packing restrictions in the lipid bilayer and. Some of the methods used to remove the untrapped drug are dialysis. Active loading methods have the following advantages over passive encapsulation techniques: high encapsulation efficiency. Additionally. Liposomal fusion with cells and with intracellular contents has been a major area of research in the case of fusogenic liposomes. Removal of unentrapped drugs from liposomes It is easier to remove the encapsulated drug in case of MLVs than in the case of SUV or LUVs. and several investigators have utilized this association to efficiently entrap adriamycin into liposomes and increase drug-retention. being mixed homogeneously with the lipid component prior to vesicle hydration step. A good example is the amphiphilic anticancer drug adriamycin.g. their concentration. the assay and impurity tests can be done by comparison with the reference standard (e. detergents can also be removed by adsorption to hydrophobic resins or cyclodextrins. Physical characterization includes study of vesicle size.. Between these two extremes are amphiphilic agents which are often the most difficult to retain inside liposomes as they can rapidly permeate through lipid bilayers. MLVs because of their large size settle at the bottom forming a pellet when centrifuged at high speed and drug remains in the supernatant. like pH sensitive liposomes. Common procedures of detergent removal from the mixed micelles are dilution. gel chromatography etc. aggregation for the role of liposomes in triggered drug release or stimulus-mediated fusion of liposomal constituents with target cells and protein binding. gel chromatography and dialysis through hollow fibers or through membrane filters. as a result.by the liposomes. Chemical characterization includes studies establishing purity and potency of various liposomal constituents. minicolumn centrifugation. fusion.
positional specificity of acyl side chains. Therefore. the lipid composition (e. remains substantially in the encapsulated form d ratio of unencapsulated to encapsulated drug substance remains constant . then assays capable of confirming the fatty acid composition and positional specificity should be used. Stability in vivo covers the stability aspects once the formulation is administered via various routes to the biological fluids like in blood (serum) when given intravenously and in GI tract when given orally. aggregation. High temperature stability studies are not undertaken for liposomal preparations since they can dramatically alter the nature of the interfacial film especially if the phase transition temperature is reached. tests for physical parameters should be developed to assess the integrity and size of the liposomes. Liposome drug products should be evaluated for stability of the encapsulated drug substance as well as stability of the lipids that compose the liposomal bilayer. stable liposomes might not be formed. Other examples of parameters that can be critical to the performance of the lipid are the amount of phosphatidylglycerol or phosphatidylserine in a lecithin preparation. small unilamellar vesicles are more susceptible to size changes than are multilamellar vesicles. Also. Stability testing of unloaded liposomes (i. For natural lipid mixtures such as egg lecithin. percentage of each lipid and fatty acid. Liposomes are susceptible to fusion. The liposome is considered stable in vivo if. when in circulation. and leakage of the encapsulated drug substance during storage. For instance. A wide variation in the storage temperature often leads to a change in the fundamental nature of the system. Based on the nature of lipid or lipid mixtures. including the liposome itself in vitro and in vivo. For instance. rug substance. degree of fatty acid unsaturation) should be specified in some circumstances. Stress testing of liposome drug products and unloaded liposomes may be warranted to demonstrate possible degradation or other reaction processes unique to the liposomes. while both saturated and unsaturated lipids are subject to hydrolysis to form lysolipids and free fatty acids. Therefore.e. over the time course of the single-dose study. If the data indicate that this is a critical factor. tests should be developed to evaluate the chemical stability of the lipids in the liposome drug product.g.. acceptance criteria for the degree of fatty acid unsaturation should be included in the specifications. if the degree of un saturation of the fatty acid side chains is too high. liposomes to be combined with a drug substance before use) should also be performed. the type of lipids in the bilayer or the encapsulated drug substance may affect fusion of the liposomes or leakage of drug substance from the liposome. The storage of liposomal preparations should be strictly defined and controlled. the specifications should be sufficient to ensure that the lipid can perform adequately in the liposome drug product and conform to impurity limits. Stability studies should address both physical and chemical stability of the liposome drug product. Lipids with unsaturated fatty acids are subject to oxidative degradation.. The physical stability of liposome drug products is a function of the integrity and the size distribution of the lipid vesicles.Desired lipids from impurities.
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