Liposomal ophthalmic drug delivery system. II. Dihydrostreptomycin sulfate

The carrier ability of liposomes for a model hydrophilic compound vas investigated in the rabbit eye. Dihydrostreptomycin sulfate was encapsulated in various types of liposomes, i.e. large and small uni- and multilamellar vesicles having either positive or neutral surface charge. An aqueous solution served as control preparation. Results indicated that liposomal encapsulation reduced the ocular drug con- centration. Addition of empty liposomes to the control solution did not alter drug levels in most of the ocular tissues. Among the liposomal preparations the large multi- and unilamellar vesicles provided higher drug concentration in all ocular tissue than the small unilamellar ones. Introduction of a positive charge on liposome surface enhanced liposome-conjunctiva interactions. The results suggest that liposomal encapsulation alters drug disposition in the eye lepending on the type of liposomes and the physicochemical properties of the encapsulated drug. In the case of the dihydrostreptomycin sulfate and possibly other hydrophilic drugs the liposomal encapsulation provides no advantages as far as drug delivery is concerned.     

Effect of preparation techniques on the properties of curcumin liposomes: characterization of size, release and cytotoxicity on a squamous oral carcinoma cell line

Curcumin, a chemopreventive agent, was incorporated into liposomes using different preparation techniques and characterized for parameters such as drug loading efficiency, size, in vitro release and in vitro cytotoxicity on a squamous carcinoma cell line. Liposomes were prepared with different methods - thin layer evaporation, ethanol injection and sonication methods, respectively, obtaining, multilamellar vesicles (MLVs) and small unilamellar vesicles (SUVs). The preparation techniques influenced the size, encapsulation efficiency, in vitro release and cytotoxicity profiles. Encapsulation efficiency increased with decrease in drug to lipid ratio in the following rank order - MLVs > SUVs > ethanol injection vesicles. In vitro release and in vitro cytotoxicity were a function of the size of vesicle, which varied depending on the preparation technique. Based on these results, it can be concluded that different liposomal formulations can be employed to achieve unique in vivo needs in cancer chemotherapy.     

Effect of preparation technique on the properties of liposomes encapsulating ketoprofen-cyclodextrin complexes aimed for transdermal delivery

The combined approach of cyclodextrin complexation and entrapment in liposomes was investigated in order to develop an effective topical formulation of ketoprofen. Equimolar complex of drug and hydroxypropyl-beta-cyclodextrin (HPbetaCyd) was added at different concentrations to the aqueous phase of liposomes consisting of phosphatidylcholine and cholesterol (60%/40%, w/w). Liposomes were prepared with different techniques, such as thin layer evaporation, freezing and thawing, extrusion through microporous membrane, and reverse phase evaporation method, obtaining, respectively, multi-lamellar vesicles (MLV), frozen and thawed MLV (FATMLV), small uni-lamellar vesicles (SUV) and large uni-lamellar vesicles (LUV). Size and morphology of the different types of liposomes were investigated by light scattering analysis, transmission electron microscopy, and confocal laser scanning microscopy, whereas drug entrapment efficiency was determined by dialysis experiments. Cyclodextrin complexation improved drug solubilization and allowed a strong improvement of its entrapment into the aqueous liposomal phase. Liposome preparation method and operating conditions clearly affected both liposome size and drug loading capacity. Encapsulation efficiency increased with increasing the complex concentration up to 10 mM, and was in the order MLV>LUV>SUV. An opposite behaviour was observed for FATMLV, probably due to the freezing phase required by such a preparation method, which reduced the complex solubility. Moreover, it was not possible to use higher complex concentrations, due to the destabilizing effect of cyclodextrins toward the liposomal membrane. Permeability studies of drug-HPbetaCyd complexes, directly in solution or incorporated in liposomes, performed across artificial membranes simulating the skin behaviour, highlighted, as expected, a prolonged release effect of liposomal formulations. Furthermore, the drug permeation rate depended on the vesicle characteristics and varied in the order: SUV>MLV=FATMLV>LUV. Therefore, the most suitable liposome preparation method can be suitably selected on the basis of drug encapsulation efficiency and/or desired drug release rate.     

Encapsulation of vinorelbine into cholesterol-polyethylene glycol coated vesicles: drug loading and pharmacokinetic studies

Objectives Pegylated liposome formulations of vinorelbine with prolonged circulation half‐life (t½) are desirable. However, DSPE‐PEG could affect vinorelbine loading into vesicles due to electrostatic interactions. To resolve this problem, chol‐PEG was used to prepare pegylated liposomal vinorelbine and the factors affecting drug loading and plasma pharmacokinetics were investigated. Methods Vinorelbine was loaded into liposomes using a novel triethylamine 5‐sulfosalicylate gradient. The effects of cholesterol and chol‐PEG on drug loading were investigated. Pharmacokinetic studies were performed in normal KunMing mice treated with different liposomal vinorelbine formulations. To clarify the effects of chol‐PEG on membrane permeability, drug release experiments were performed based on the fluorescence dequenching phenomenon of a fluorescence marker. Key findings In contrast to DSPE‐PEG, even at high PEG grafting density (～8.3 mol%), chol‐PEG had no effect on vinorelbine loading into HSPC/cholesterol (3 : 1, mass ratio) vesicles. However, for the formulations with low cholesterol content (HSPC/cholesterol 4 : 1), loading efficiency decreased with increasing chol‐PEG content. In vivo, the vinorelbine t½ of low cholesterol formulations decreased with increasing chol‐PEG content, but for high cholesterol liposomes, the maximum vinorelbine t½ was achieved at ～3 mol% chol‐PEG grafting density. The resulting vinorelbine circulation t½ was ～9.47 h, which was greater than that of non‐pegylated liposomes (～5.55 h). Drug release experiments revealed that chol‐PEG might induce membrane defects and concomitant release of entrapped marker, especially at high chol‐PEG density. Conclusions Through the investigation of the effects of chol‐PEG and cholesterol, an optimum pegylated liposomal vinorelbine formulation with prolonged t½ was achieved. In plasma, the membrane defect induced by chol‐PEG may counteract the long circulation characteristics that chol‐PEG afforded. When these two opposite effects reached equilibrium, the maximum vinorelbine t½ was achieved.     

Effect of formulation design and freeze-drying on properties of fluconazole multilamellar liposomes

Fluconazole-entrapped multilamellar liposomes were prepared using the thin-film hydration method. The effects of cholesterol molar ratio, charge-inducing agents, and α-tocopherol acetate on encapsulation efficiency values and in vitro drug release of multilamellar liposomes were studied. Freeze-dried liposomal products were prepared with or without cryoprotectants. Results showed that incorporation of stearylamine resulted in an increased entrapment of fluconazole, whereas incorporation of dicetyl phosphate decreased the drug entrapment efficiency. The incorporation of α-tocopherol acetate into fluconazole multilamellar liposomes resulted in the increase of entrapment efficiency of fluconazole liposomes. In vitro release studies revealed that incorporation of cholesterol into multilamellar liposomal formulations decreased drug permeability from formulations. Positively charged fluconazole multilamellar liposomes gave rise to a slow release rate compared to neutral liposomes whereas negatively charged fluconazole liposomes showed a rapid release rate. Physical stability studies showed that lyophilized cake of liposomes without cryoprotectants was compact and difficult to reconstitute compared to fluffy easily reconstituted cakes upon using cryoprotectants. Fluconazole retained in freeze-dried liposomes without cryoprotectants was 63.452% compared to 91.877% using three grams of trehalose as a cryoprotectant per gram lipid in positively charged multilamellar liposomes. Physical stability studies showed superior potentials of the lyophilized product after reconstitution in comparison with those of a solution product.     

Methods to increase the encapsulation efficiency for liposomal drugs

Liposomes as drug delivery systems--in comparison to the traditional dosage forms--offer the advantage of the targeted drug delivery and as a consequence, reduction of the side effects. In case of fluoroquinolone antibiotics, such as lomefloxacin, the liposomal encapsulation of the active ingredient can result in an enhancement of its therapeutic efficacy against intracellular bacteria. The aim to improve the liposomal encapsulation efficiency of drugs--which is one of the main factors influencing the therapeutic effect of vesicular dosage forms--is one of the important challenges in the field of pharmaceutical technology. In our experiments we prepared lomefloxacin containing multilamellar vesicles from various lipids using different hydrating solutions. We intended to study the effect of lipid composition, cholesterol content and surface charge of liposomes on the encapsulation efficiency of lomefloxacin. Our results can contribute to the rational design of fluoroquinolone containing liposomal drugs.     

Liposome encapsulation enhancement of methotrexate sensitivity in a transport resistant human leukemic cell line

A stable mutant of human leukemia CCRF/CEM cells has recently been isolated which is transport resistant for methotrexate (MTX). Encapsulation of MTX in cationic unilamellar liposomes increased the association of the drug 5-fold with the sensitive, and 50-fold with the resistant, cells as compared to the uptake of free drug. The liposome-mediated associations of MTX with sensitive and transport deficient cell lines were similar. Cytostatic studies demonstrated that liposome encapsulation increased MTX activity 4-fold towards the transport resistant cell line. The addition of cholesterol to the vesicles decreased their effectiveness. A 4-fold increase in drug sensitivity due to encapsulation may allow such transport resistant tumor cells to become responsive to chemotherapeutic doses of MTX which are currently feasible in human clinical protocols.     

Development and Characterization of a Liposome Preparation by a pH-Gradient Method

Abstract— A pH gradient across liposome bilayers was established in order to load a model drug (orciprenaline sulphate) into liposome vesicles. This method of liposome loading resulted in yields as high as 80–85% encapsulation. An eight-step process was designed to scale-up the process and was evaluated. In this process a diafiltration technique was successfully used to remove the excess orciprenaline sulphate present in the external medium. Finally, drug-loaded liposomes were lyophilized using lactose as an internal and external liposomal cryoprotectant. Five-month stability data for the liposomes is reported. An HPLC technique was used to determine the drug concentration and a laser light-scattering technique was employed to determine the liposome vesicle size and polydispersity factor. Liposomes prepared by the pH-gradient method showed high encapsulation efficiency. Upon storage at 2–8°C the vesicle size increased and encapsulation efficiency decreased with time. These phenomena are attributed to gradual fusion of liposomes and loss of drug to the extra-liposomal media.     

Effect of Neutral Liposomes on Corneal and Conjunctival Transport of Didanosine

The objective of this study is to determine the effect of various neutral liposomes on corneal and conjunctival permeability of didanosine (ddI), an antiviral drug. Multi-lamellar vesicles (MLVs), large unilamellar vesicles (LUVs), and sonicated multi-lamellar vesicles (SMLVs) encapsulating ddI (with trace quantities of 3HddI) were prepared using distearoyl phosphatidylcholine (DSPC), a neutral lipid. The liposomes contained 14C-cholesteryl oleate as a lipid tracer. Liposome formulations containing free and encapsulated drug (f + e) and those containing only encapsulated drug (e) of an equal quantity were compared with free drug in this study. The permeability studies were conducted in the mucosal to serosal direction across excised rabbit cornea and conjunctiva. The percent encapsulation of ddI in MLVs, LUVs, and SMLVs was 25.66 0.30, 26.56 0.57, and 19.41 0.30, respectively. The mean particle size of MLVs, LUVs, and SMLVs containingfree and encapsulated drug was 3058, 774, and 270 nm, respectively. With all liposome formulations tested, the percent uptake of lipid by tissues was higher compared to ddI uptake. While ddI permeated across the tissues, the lipid tracer did not permeate in detectable quantities.The SMLV(e) formulation was better than the SMLV(f + e) formulation with respect to initial flux and tissue uptake in both tissues and permeability across conjunctiva. In general, the permeability coefficient, initial flux, and tissue levels of ddI at the end of the transport study were lower in the presence of all liposome formulations compared to free drug. Thus, neutral liposomal encapsulation is not a suitable approach to enhance the corneal or conjunctival transport or uptake of ddI.     

Copper ion-mediated liposomal encapsulation of mitoxantrone: The role of anions in drug loading, retention and release

Besides pH gradient, other transmembrane gradients such as metal ion gradient could be also employed to load drugs into liposomes. In pH gradient method, anions have an important role since they could form specific aggregates with drugs, and then affect drug release kinetics from vesicles. To explore the role of anions in metal ion gradient method, copper ion-mediated mitoxantrone (MIT) loading was investigated systematically. When empty liposomes exhibiting a transmembrane copper ion gradient (300 mM) were mixed with MIT in a molar ratio of 0.2:1, after 5 min incubation at 60 degrees C, >95% MIT could be loaded into vesicles and the encapsulation was stable, regardless of the kinds of anions and initial intraliposomal pH values. The encapsulation ratio decreased with increased MIT/lipid molar ratio. But even when the molar ratio increased to 0.4, >90% encapsulation could still be achieved. In the presence of nigericin and ammonium, the drug loading profiles were affected to different degree with respect to both drug loading rate and encapsulation ratio. Relative to CuSO(4)-containing systems, CuCl(2) mediated MIT loading was unstable. Both nigericin and ammonium could alter the absorption spectra of liposomal MITs loaded with CuSO(4) gradient. In vitro release studies were performed in glucose/histidine buffer and in 50% human plasma using a dialysis method. In both of release media, CuCl(2)-containing vesicles displayed rapid release kinetics in comparison with CuSO(4) systems; and during the experiment period, MIT was lost from the vesicles continuously. When the formulations were injected into BDF1 mice at a dose of 4 mg/kg, all the liposomal formulations exhibited enhanced blood circulation time, with half-life values of 6.8-7.2h, significantly compared to the rapid clearance of free-MIT. In L1210 ascitic model, CuCl(2) formulation was more therapeutically active than CuSO(4) formulation. At a dose of 6 mg/kg, the treatment with CuCl(2) formulation resulted in a median survival time of 21 days, considerably larger than that of CuSO(4) groups (15 days). Based on these data, it was concluded that during the drug loading process, a dynamic transmembrane pH gradient is generated and intraliposomal pH might affect the complexation manner in which Cu(2+) binds MIT. Owing to the presence of pH gradient, after the accumulation within vesicles, a part of MIT will be protonated and precipitated by sulfate. Accordingly, the aggregation status of MIT inside CuSO(4) system was more complicated than that in CuCl(2) vesicles. The difference in physical status of MIT aggregates affects not only the drug release rate, but also their therapeutic effects.     

Encapsulation,Stability and In-vitro Release Characteristics of Liposomal Formulations of Colchicine

The severe toxicity and low therapeutic index of colchicine limit its therapeutic use. Encapsulation in liposomes might reduce these toxic effects. The objective of this study was to determine the factors influencing encapsulation of colchicine in liposomes and to optimize the encapsulation parameters. Colchicine was encapsulated in multilamellar liposomes and large unilamellar liposomes prepared using various phospholipids. The effects of method of preparation, type of vesicle, charge, and concentration of cholesterol on encapsulation of colchicine in liposomes were investigated. Also, stability of colchicine under stress conditions and at various temperatures, and in-vitro release characteristics were determined. A significant difference in encapsulation of colchicine in multilamellar liposomes was observed when prepared by two different methods. Induction of charge on the liposome surface increased encapsulation of colchicine in multilamellar liposomes, but did not affect large unilamellar liposomes. The liposome preparations could withstand simulated transport conditions and frequent changes in temperature. Particle size and concentration of colchicine did not change significantly during storage at various temperatures for six months. In order to retain encapsulated colchicine in liposomes, storage at or below room temperature was found to be suitable. In-vitro release of colchicine from large unilamellar liposomes was biphasic and was influenced by two rate-limiting barriers, the dialysis membrane and the liposome bi-layers. For optimum encapsulation and stability of colchicine liposomes were prepared from a mixture of 1,2-distearoyl-sn-glycero-3-phosphocholine, cholesterol and either stearylamine or dicetyl phosphate.     

Increasing bioavailability of silymarin using a buccal liposomal delivery system: preparation and experimental design investigation

Silymarin is a natural lipotropic agent of low bioavailability from oral products. The aim of our study is to prepare buccal liposomal delivery system of silymarin with higher bioavailability. The effect of lecithin:cholesterol molar ratio on the percentage drug encapsulated was investigated. The influence of fluctuating the amount of added drug was also determined. The effect of additives such as positive charge inducer, negative charge inducer and surfactants was studied using two different 2(3) full factorial designs. Furthermore, additives used to optimize liposomal product were also investigated for their optimal concentrations, release properties and in vitro permeation and absorption through chicken cheek pouch. Optimal liposomal encapsulation efficiency was found at 7:4 lecithin to cholesterol molar ratio. A decrease in entrapment efficiency with increasing cholesterol content was observed. Tween 20 or Tween 80 beyond 0.5 molar ratio decreased the entrapment efficiency. Positively charged liposomes showed superior entrapment efficiency over neutral and negatively charged liposomes. Release studies as well as permeation and absorption studies showed that hybrid liposomes prepared according to formula 3 containing lecithin, cholesterol, stearyl amine and Tween 20 in 9:1:1:0.5 molar ratio, respectively, gave the best drug absorption and permeation. It showed steady state permeation through chicken cheek pouch for 6h. This is expected to improve the bioavailability of silymarin in the developed liposomal buccal delivery system, as the results show an increase in drug penetration compared to free drug powder.     

Liposomal ophthalmic drug delivery system I. Triamcinolone acetonide

The potential of liposomes as an ophthalmic drug delivery system was investigated in the rabbit. Triamcinolone acetonide as a model compound for lipophilic drugs was encapsulated into large multilamellar vesicles and instilled into the eye. A suspension form served as a control preparation. Drug distribution was determined at various time intervals. Compared to the suspension, the liposomal form produced significantly higher drug levels in the ocular tissues up to 5 h after the drug administration. The results suggest that liposomal encapsulation of the drug may be a superior drug delivery system for ocular therapy.     

A targeted liposome delivery system for combretastatin A4: formulation optimization through drug loading and in vitro release studies

Efficient liposomal therapeutics require high drug loading and low leakage. The objective of this study is to develop a targeted liposome delivery system for combretastatin A4 (CA4), a novel antivascular agent, with high loading and stable drug encapsulation. Liposomes composed of hydrogenated soybean phosphatidylcholine (HSPC), cholesterol, and distearoyl phosphoethanolamine-PEG-2000 conjugate (DSPE-PEG) were prepared by the lipid film hydration and extrusion process. Cyclic arginine-glycine-aspartic acid (RGD) peptides with affinity for alphav beta3-integrins overexpressed on tumor vascular endothelial cells were coupled to the distal end of polyethylene glycol (PEG) on the liposomes sterically stabilized with PEG (non-targeted liposomes; LCLs). Effect of lipid concentration, drug-to-lipid ratio, cholesterol, and DSPE-PEG content in the formulation on CA4 loading and its release from the liposomes was studied. Total liposomal CA4 levels obtained increased with increasing lipid concentration in the formulation. As the drug-to-lipid ratio increased from 10:100 to 20:100, total drug in the liposome formulation increased from 1.05+/-0.11 mg/mL to 1.55+/-0.13 mg/mL, respectively. When the drug-to-lipid ratio was further raised to 40:100, the total drug in liposome formulation did not increase, but the amount of free drug increased significantly, thereby decreasing the percent of entrapped drug. Increasing cholesterol content in the formulation decreased drug loading. In vitro drug leakage from the liposomes increased with increase in drug-to-lipid ratio or DSPE-PEG content in the formulation; whereas increasing cholesterol content of the formulation up to 30 mol-percent, decreased CA4 leakage from the liposomes. Ligand coupling to the liposome surface increased drug leakage as a function of ligand density. Optimized liposome formulation with 100 mM lipid concentration, 20:100 drug-to-lipid ratio, 30 mol-percent cholesterol, 4 mol-percent DSPE-PEG, and 1 mol-percent DSPE-PEG-maleimide content yielded 1.77+/-0.14 mg/mL liposomal CA4 with 85.70+/-1.71% of this being entrapped in the liposomes. These liposomes, with measured size of 123.84+/-41.23 nm, released no significant amount of the encapsulated drug over 48 h at 37 degrees C.     

5-Fluorouracil in vesicular phospholipid gels for anticancer treatment: entrapment and release properties

Vesicular phospholipid gels (VPG), i.e. highly concentrated liposomal dispersions, are suitable for entrapping substances such as anticancer drugs with particular high encapsulation efficiencies (EE). We prepared different formulations of VPG with 30% (w/w) lipid containing 5-fluorouracil (5-FU) by high pressure homogenization and analysed their EE and drug release. Using mixtures of hydrogenated soy phosphatidylcholine and cholesterol with molar ratios ranging from 55/45 to 75/25, a decreasing amount of cholesterol correlated with an increasing EE, which is probably due to a reduced amount of smaller vesicles and number of lamellae. Using a 5-FU solution of pH 8.6 for VPG preparation, an EE of approximately 40% was found after redispersion of the gel to a liposomal dispersion and separation of free drug from liposomal drug by size exclusion chromatography. The reduced EE for preparations with lower pH values was attributed to a fast initial drug release due to the increased drug lipophilicity below the pK(a) value of 8. After redispersion of a VPG of pH 8.0, an initially faster release of about a third of the entrapped drug was found during the first 20 min, followed by stable entrapment over many hours. The rapid initial release may be due to the portion of liposomes smaller than 40 nm in diameter, determined by photon correlation spectroscopy. Cryo electron microscopic pictures show a lentil-like shape of these small liposomes. The membrane defects on the edges are probably the reason for the very high initial drug release rate. The half-life time of the release of 5-FU from intact FU-VPG at both pH 7.4 and 8.0 was found to be in the order of 4-5 h and the kinetics are typical for matrix-controlled drug diffusion. The in vitro data of 5-FU loaded VPG suggest their applicability as implants with controlled release properties or, after redispersion, as intravenously injected liposomal formulations.     

Pharmacological and local toxicity studies of a liposomal formulation for the novel local anaesthetic ropivacaine

This study reports an investigation of the pharmacological activity, cytotoxicity and local effects of a liposomal formulation of the novel local anaesthetic ropivacaine (RVC) compared with its plain solution. RVC was encapsulated into large unilamellar vesicles (LUVs) composed of egg phosphatidylcholine, cholesterol and alpha-tocopherol (4:3:0.07, mole %). Particle size, partition coefficient determination and in-vitro release studies were used to characterize the encapsulation process. Cytotoxicity was evaluated by the tetrazolium reduction test using sciatic nerve Schwann cells in culture. Local anaesthetic activity was assessed by mouse sciatic and rat infraorbital nerve blockades. Histological analysis was performed to verify the myotoxic effects evoked by RVC formulations. Plain (RVC(PLAIN)) and liposomal RVC (RVC(LUV)) samples were tested at 0.125%, 0.25% and 0.5% concentrations. Vesicle size distribution showed liposomal populations of 370 and 130 nm (85 and 15%, respectively), without changes after RVC encapsulation. The partition coefficient value was 132 +/- 26 and in-vitro release assays revealed a decrease in RVC release rate (1.5 fold, P < 0.001) from liposomes. RVC(LUV) presented reduced cytotoxicity (P < 0.001) when compared with RVC(PLAIN). Treatment with RVC(LUV) increased the duration (P < 0.001) and intensity of the analgesic effects either on sciatic nerve blockade (1.4-1.6 fold) and infraorbital nerve blockade tests (1.5 fold), in relation to RVC(PLAIN). Regarding histological analysis, no morphological tissue changes were detected in the area of injection and sparse inflammatory cells were observed in only one of the animals treated with RVC(PLAIN) or RVC(luv) at 0.5%. Despite the differences between these preclinical studies and clinical conditions, we suggest RVC(LUV) as a potential new formulation, since RVC is a new and safe local anaesthetic agent.     

Long circulating liposomes of 2',3'-dideoxyinosine: Formulation and stability

2',3'-Dideoxyinosine (ddI), an anti-human immunodeficiency virus (HIV) agent, was encapsulated in liposomes. The influence of the phospholipid/cholesterol ratio, concentration of phospholipid (PL), and chain length of PL on the encapsulation of ddI in multilamellar vesicles (MLVs), frozen and thawed multilamellar vesicles (FAT MLVs), and large unilamellar vesicles (LUVs) was studied. An optimum formulation was then selected to prepare long circulating liposomes. Stability studies at 4, 25, and 37°C and under certain stress conditions were performed. Release characteristics in phosphate buffer (pH 7.4) at 37°C were studied. Results show an increase in encapsulation efficiency (EE) with increasing amounts of cholesterol, a decrease in EE and increase in encapsulation yield (EY) with increasing concentrations of PL, and an increase in EE with increases in PL chain length, in both MLVs and LUVs. Freezing and thawing of MLVs had no influence on EE at a PL concentration of 10 mg/mL but increased EE at higher concentrations of PL. Various stability tests showed the formulation to be stable to leakage of entrapped drug when stored at 4, 25, and 37°C for 6 months, when subjected to mechanical stress, and on exposure to human serum. The release studies indicated that 70% of ddI was released over a period of 72 h.     

Radiopaque liposomes: effect of formulation conditions on encapsulation efficiency

Liposomes containing sodium ioxitalamate were prepared by sonication. Suitable amounts of purified soybean phosphatidylcholine and cholesterol were used at various molar ratios. Stearylamine or dicetylphosphate were added to this lipid composition when charged liposomes were required. After sonication and removal of unencapsulated solute, this manufacturing process yielded small multilamellar vesicles as confirmed by electron microscopy. These liposomes did not exhibit a narrow range of size distribution; the mean particle size varied from 135 to 145 nm. With respect to the efficiency of encapsulation, two parameters were distinguishable: the volume of encapsulated aqueous space per unit of lipid weight and the percentage of the contrast agent added that became encapsulated in the liposomes. Investigation of the preparative parameters revealed that increased molar ratios of cholesterol yielded higher aqueous volume and iodine contents in the liposomes, which were attributed to a reduction of the liposome permeability to the contrast agent. However, the inclusion of cholesterol into the bilayer liposomal membrane was limited, probably by solubility restrictions. Negatively and positively charged liposomes had higher rates of encapsulation than did neutral liposomes. This result was expected since efficient encapsulation of polar compounds requires formation of large aqueous spaces within the vesicles per mole of lipids. Increase of the lipid fractions at a constant, reduced the aqueous volume entrapped per millimole of lipid and, consequently, the iodine content in the liposomes. However, an increase in the initial sodium ioxitalamate concentration diminished the aqueous volume entrapped in the liposomes but increased the iodine content.     

Niosomes as carriers for tretinoin. I. Preparation and properties

Tretinoin-loaded niosomes were prepared from polyoxyethylene (4) lauryl ether, sorbitan esters and a commercial mixture of octyl/decyl polyglucosides, in the presence of cholesterol and dicetyl phosphate. Liposomes made of hydrogenated and non-hydrogenated phosphatidylcholine were also prepared as a comparison reference. A study was made of the influence of vesicle composition and preparation method on the vesicle structure (MLV, LUV, SUV), size distribution, entrapment efficiency and in vitro release of incorporated tretinoin. Results showed that in the presence of cholesterol all the amphiphiles used were able to form stable vesicle dispersions with or without tretinoin. Vesicle sizes were dependent on the preparation method, bilayer composition and drug load. Multilamellar (MLV) vesicles were larger than extruded (LUV) and sonicated (SUV) vesicles while drug-loaded vesicles were generally smaller than empty ones. Entrapment efficiencies of tretinoin were always very high especially for multilamellar (91-99%) and extruded (88-98%) vesicles. The in vitro release of tretinoin from the prepared vesicular formulations was studied using the vertical Franz diffusion cells. The rate of drug release through a Silastic membrane from a liposomal and niosomal tretinoin dispersion was generally faster than from a tretinoin solution. Release data showed that tretinoin delivery is mainly affected by the vesicular structure and that tretinoin delivery increased from MLVs to LUVs to SUVs.     

Ciprofloxacin Encapsulation Into Giant Unilamellar Vesicles: Membrane Binding and Release

This study aimed at investigating some respects of binding and interaction between water-soluble drugs and liposomal carrier systems depending on their size and lamellarity. As model substance, ciprofloxacin hydrochloride (CPFX) was incorporated into giant unilamellar vesicles (GUVs) to study their CPFX encapsulation/binding capacity. To characterize molecular interactions of various CPFX microspecies with lipid bilayer, zeta potential and electron paramagnetic resonance (EPR) spectroscopy measurements were performed. The increase of the zeta potential at pH 5.4 but no change at pH 7.2 was interpreted in terms of the CPFX microspecies' distribution at the two pH values. EPR observations showed an increased fluidity because of CPFX binding to GUVs. We worked out and applied a three-compartment dialysis model to separately determine the rate of drug diffusion through the liposomal membrane. Equilibrium dialysis showed (a) different permeation of CPFX through the membranes of GUVs and multilamellar vesicles (MLVs), with characteristic half-lives of 54.4 and 18.1 h, respectively; and (b) increased retention of CPFX in case of GUVs with released amounts of 70% compared with about 97% in case of MLVs. Our results may provide further details for efficient design of liposomal formulations incorporating water-soluble drugs. © 2012 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 102:694–705, 2013