Concanavalin-A conjugated fine-multiple emulsion loaded with 6-mercaptopurine

Fine-multiple (water-in-oil-in-water) emulsions were prepared by two-step emulsification using sonication. They were coated with concanavalin-A (Con-A) by three methods. The one involving covalent coupling of Con-A to the multiple emulsion incorporated anchor was better compared with lipid derivatized Con-A anchoring or the glutaraldehyde-based cross-linking method, as shown by the faster rate of dextran-induced aggregation. The selected multiple emulsions were characterized by physical properties such as droplet size, encapsulation efficiency, and zeta potential. Stability parameters such as droplet size, creaming, leakage, and aggregation as a function of relative turbidity were monitored over a 1-month period, which revealed good stability of the formulations. The release profile of 6-mercaptopurine followed zero-order kinetics. Pharmacokinetic studies showed an increase in half-life and bioavailability from multiple emulsion formulations administered intravenously. There was prolonged retention of drug in various tissues of rats when treated with Con-A-coated multiple emulsion as compared with uncoated one. Our study demonstrates the suitability of fine-multiple emulsion for intravenous administration and the potential for prolonged retention of drugs and targeting in biological systems.     

LIPID EMULSIONS OF PALMITOYLRHIZOXIN: EFFECTS OF PARTICLE SIZE ON BLOOD DISPOSITIONS OF EMULSION LIPID AND INCORPORATED COMPOUND IN RATS

Emulsion formulations of various particle sizes for the highly lipophilic antitumour agent, RS-1541 (13-O-palmitoylrhizoxin), were prepared using dioctanoyldecanoyl-glycerol (ODO) as lipids and polyoxyethylene-(60)-hydrogenated castor oil (HCO-60) as a surfactant. These emulsions were evaluated as injectable drug carriers and compared with a colloidal solution. Both in vitro and in vivo after i.v. administration, RS-1541 was distributed into lipoproteins from the colloidal solution. When applied as emulsions of various particle sizes (124–419 nm) in vitro, RS-1541 was retained and stabilized within the emulsions. In the in vivo study, however, retention of RS-1541 in the emulsions after i.v. injection depended on their size. The small-particle emulsions (94–112 nm) resulted in long retention, and the large-particle emulsions (415–474 nm) led to short retention. Lipolysis rates of emulsion particles by lipoprotein lipase also depended on their size, indicating rapid lipolysis for small-particle emulsions (133 nm). However, the lipolysis was not such an extensive one, showing 10–30% release of capric acid from ODO within 6 h. Blood dispositions of capric acids approximately paralleled those of RS-1541 after i.v. injection of various particle size emulsions (130–368 nm) to rats, although relatively rapid eliminations of capric acids compared with RS-1541 were observed for the small-particle size emulsions (130 nm). These results suggest that when injected as emulsion formulations, the highly lipophilic antitumour agent, RS-1541, has behaviour similar to that of the emulsion particles in the body, which is dependent on the size of the latter. Thus, by properly selecting the particle size, lipid emulsions consisting of ODO and HCO-60 are expected to be effective and useful DDS carriers for RS-1541.     

Long-Circulating Multiple-Emulsion System for Improved Delivery of an Anticancer Agent

Stealth multiple emulsions of small size, containing 6-mercaptopurine, were prepared by incorporation of sphingomyelins (SM) and monosialogangliosides (GM1) in the oily phase and by coating with lipid-grafted polyethylene glycol (PEG-PC). The three types of "stealth" multiple emulsions were characterized for size distribution, zeta potential, viscosity, encapsulation efficiency, drug release, and stability. Drug disposition studies were performed with formulated multiple emulsions to assess "stealth" behavior. The tissue distribution studies were carried out with the PEG grafted multiple emulsion. The results suggest that PEG-PC-coated multiple emulsions are superior as prolonged release and extended bloodcirculating carriers compared to multiple emulsions bearing either SM or GM1.     

Preparation and evaluation of multiple emulsions water-in-oil-in-water (w/o/w) as delivery system for influenza virus antigens

In this study, investigations have been carried out to prepare adjuvant active delivery systems; multiple water-in-oil-in-water (w/o/w) emulsion formulations, containing influenza virus surface antigen Hemagglutinin (HA). A modified two-stage emulsification method has been used to prepare multiple emulsions. After improving multiple (w/o/w) emulsion formulations; F1: purified antigen solution (PAS)/soybean oil, HCO-40 and span 80/pluronic F-68, F2: PAS and HPbetaCD/soybean oil, HCO-40 and span 80/pluronic F-68, F3: PAS/squalane, HCO-40 and span 80/pluronic F-68, formulations were selected for the stability study that continued for a 3 month duration. To evaluate the stability of these formulations, microscopic observation, osmolarities of the internal and external aqueous phases, pH, globule size and viscosity were determined. SDS-PAGE (silver staining) was used to evaluate HA and the micro-bicinchoninic acid (mBCA) assay was used to determine the in vitro release of antigen from formulations. Immune responses of formulations were investigated in Wistar Albino rats and compared with the immune response raised against the conventional vaccine. These responses were detected with Hemagglutination Inhibition (HAI) assay. The results of this study demonstrated that HA was well entrapped in the multiple (w/o/w) emulsion formulations. Molecular weight and antigenicity of the entrapped HA were not affected by the emulsification procedure. These results suggest that multiple emulsion formulations entrapping influenza antigen may have potential for immunization studies as one of the vaccine delivery system with adjuvant properties.     

Effect of lyophilization on parenteral emulsions

The feasibility of preparing lyophilized anhydrous products, for reconstitution in to emulsion dosage forms was investigated. Stable soybean o/w emulsions were prepared using a mixture of lecithin and Span 20 as the emulsifiers. Two series of emulsions were prepared for this study, each containing a polyhydroxy alcohol as a consurfactant for particle size reduction. Increasing concentrations of glycerol (10-30% w/w) were added to one group of emulsions and propylene glycol (20-60% w/w) was added to the second group of emulsions. All formulations were found to have good particle size stability. The emulsion formulation containing 30% glycerol could be successfully lyophilized into an anhydrous product. Reconstitution of this lyophilized product resulted in an emulsion essentially similar to the original emulsion prior to lyophilization. This is because the mixture of the oil phase and 30% w/w glycerol formed a self-emulsifying system. All other emulsion formulations were not suitable for lyophilization. These formulations cracked during lyophilization, separating into an upper oil layer and a lower layer of the continuous phase. The formation of an upper oil layer prevented complete drying of these emulsions. The particle size of these lyophilized emulsions, when reconstituted with the external phase was greater than the emulsion particle size prior to lyophilization. But the change in particle size was less with increasing concentrations of polyhydroxy alcohols. These results indicate that emulsions can be lyophilized to prepare a product suitable for reconstitution to a parenteral emulsion dosage form provided the formulation is designed to withstand temperature and phase changes during the lyophilization process.     

Preparation and Evaluation of Multiple Emulsions Water-in-oil-in-water (w/o/w) as Delivery System for Influenza Virus Antigens

In this study, investigations have been carried out to prepare adjuvant active delivery systems; multiple water-in-oil-in-water (w/o/w) emulsion formulations, containing influenza virus surface antigen Hemagglutinin (HA). A modified two-stage emulsification method has been used to prepare multiple emulsions. After improving multiple (w/o/w) emulsion formulations; F1: purified antigen solution (PAS)/soybean oil, HCO-40 and span 80/pluronic F-68, F2: PAS and HPβCD/soybean oil, HCO-40 and span 80/pluronic F-68, F3: PAS/squalane, HCO-40 and span 80/pluronic F-68, formulations were selected for the stability study that continued for a 3 month duration. To evaluate the stability of these formulations, microscopic observation, osmolarities of the internal and external aqueous phases, pH, globule size and viscosity were determined. SDS-PAGE (silver staining) was used to evaluate HA and the micro-bicinchoninic acid (mBCA) assay was used to determine the in vitro release of antigen from formulations. Immune responses of formulations were investigated in Wistar Albino rats and compared with the immune response raised against the conventional vaccine. These responses were detected with Hemagglutination Inhibition (HAI) assay.

The results of this study demonstrated that HA was well entrapped in the multiple (w/o/w) emulsion formulations. Molecular weight and antigenicity of the entrapped HA were not affected by the emulsification procedure. These results suggest that multiple emulsion formulations entrapping influenza antigen may have potential for immunization studies as one of the vaccine delivery system with adjuvant properties.     

Preparation and evaluation of w/o/w type emulsions containing vancomycin

The objective of this contribution is to summarize the preparation and application of water-in-oil-in-water type multiple emulsions (w/o/w emulsions) entrapping vancomycin (VCM). Formulations of the emulsions (the composition of an oily phase or the type and concentrations of surfactants) and emulsification methods (a stirring method and a membrane method) or conditions (rotation rates, pore sizes of membrane or operation pressures) were evaluated in order to prepare stable w/o/w emulsions. The pharmaceutical properties of the w/o/w emulsions - particle sizes, viscosity, phase separation and drug entrapment efficiency were measured and evaluated. We prepared stable w/o/w emulsions with a particle size of about 3 micrometer and an entrapment efficiency of VCM of about 70%. When this emulsion was administered intravenously to rats, plasma concentrations of VCM were prolonged compared to the VCM solution alone. The results of this study show the potential of the w/o/w emulsions for several clinical applications as one of the drug delivery systems.     

Drug delivery to the ocular posterior segment using lipid emulsion via eye drop administration: Effect of emulsion formulations and surface modification

This work explored submicron-sized lipid emulsion as potential carriers for intraocular drug delivery to the posterior segment via eye drops. The effects of physicochemical properties of lipid emulsion on drug delivery were evaluated in vivo using mice. Different formulations of submicron-sized lipid emulsions were prepared using a high pressure homogenization system. Using coumairn-6 as a model drug and fluorescent marker, fluorescence could be observed in the retina after administration of the lipid emulsion. The fluorescence intensity observed after administration of medium chain triglycerides containing the same amount of coumarin-6 was much lower than that observed after administration of lipid emulsions. The inner oil property and phospholipid emulsifier did not affect the drug delivery efficiency to the retina. However, compared with unmodified emulsions, the fluorescence intensity in the retina increased by surface modification using a positive charge inducer and the functional polymers chitosan (CS) and poloxamer 407 (P407). CS-modified lipid emulsions could be electrostatically interacted with the eye surface. By its adhesive property, poloxamer 407, a surface modifier, possibly increased the lipid emulsion retention time on the eye surface. In conclusion, we suggested that surface-modified lipid emulsions could be promising vehicles of hydrophobic drug delivery to the ocular posterior segment.     

An investigation into the characteristics and drug release properties of multiple W/O/W emulsion systems containing low concentration of lipophilic polymeric emulsifier

Multiple W/O/W emulsions with high content of inner phase (Phi1=Phi2=0.8) were prepared using relatively low concentrations of lipophilic polymeric primary emulsifier, PEG 30-dipolyhydroxystearate, and diclofenac diethylamine (DDA) as a model drug. The investigated formulations were characterized and their stability over the time was evaluated by dynamic and oscillatory rheological measurements, microscopic analysis and in vitro drug release study. In vitro release profiles of the selected model drug were evaluated in terms of the effective diffusion coefficients and flux of the released drug. The multiple emulsion samples exhibited good stability during the ageing time. Concentration of the lipophilic primary emulsifier markedly affected rheological behaviour as well as the droplet size and in vitro drug release kinetics of the investigated systems. The multiple emulsion systems with highest concentration (2.4%, w/w) of the primary emulsifier had the lowest droplet size and the highest apparent viscosity and highest elastic characteristics. Drug release data indicated predominately diffusional drug release mechanism with sustained and prolonged drug release accomplished with 2.4% (w/w) of lipophilic emulsifier employed.     

Stability of parenteral nanoemulsions loaded with paclitaxel: the influence of lipid phase composition,drug concentration and storage temperature

Abstract

Paclitaxel was loaded into licensed parenteral nutrition nanoemulsions (Clinoleic® and Intralipid®) using bath sonication, and the stability of the formulations was investigated following storage for two weeks at room temperature or at 4?°C. In general, Clinoleic droplets were smaller than Intralipid droplets, being around 255 and 285?nm, respectively, for blank and freshly loaded emulsions. Regardless of storage temperature, the Clinoleic exhibited a very slight or no increase in droplet size upon storage, whilst the droplet size of the Intralipid emulsion increased significantly. The droplet size of both emulsions was minimally affected by paclitaxel concentration within the range of 0, 1, 3 and 6?mg/ml. The pH of both emulsions markedly decreased upon storage at room temperature, which was possibly attributed to the production of fatty acids resulting from phospholipid hydrolysis. However, at 4?°C, the pH of Clinoleic emulsion was unaffected by storage or paclitaxel concentration while the Intralipid emulsion demonstrated a trend for pH reduction. Both nanoemulsions had a negative zeta potential, with the Clinoleic formulations having the highest charge, possibly explaining the better size stability of this emulsion. Overall, this study has shown that paclitaxel was successfully loaded into clinically licensed parenteral emulsions and that Clinoleic showed greater stability than the Intralipid.     

Studies on the effect of pilocarpine incorporation into a submicron emulsion on the stability of the drug and the vehicle.

In order to obtain a novel ocular formulation with a potential for prolonging pilocarpine activity, the drug (2.0%) was incorporated into a submicron emulsion containing soya-bean oil and lecithin as emulgator. The effect of drug incorporation into the emulsion on its physical stability and on the other hand, the potential of the vehicle to reduce drug degradation at pH higher than 5.0 was studied. The pH was adjusted to 6.5 or 5.0 and the physicochemical stability of the formulations was observed. The mean diameter of oily particles in the resulting emulsions measured by a laser diffractometer was 0.6-0.7 micron and this was larger than in a drug-free emulsion where a 0.33 micron value was measured. The formulations were physically stable for 6 months at 4 degrees C, but progressing chemical degradation of pilocarpine was noted at pH 6.5. At that pH nearly 8% of pilocarpine was degraded to isopilocarpine and pilocarpic acid, both in the emulsion and in the solution. Thus, it may be concluded that pilocarpine in submicron emulsion is not protected against degradation. The presence of pilocarpine changes the physical stability of the vehicle since the formulation was easily destabilized during autoclaving or at room temperature. In the presence of higher concentration of lecithin (2.4%) or co-emulgators (poloxamer 2.0% or Tween 80 0.5%) the mean droplet size in the emulsions was the same as in a drug-free system. However the emulsions containing poloxamer were not stable during storage. Viscosity of pilocarpine emulsions can be increased by addition of methylcellulose or sodium carmellose (1.0%), but an intensive creaming occurs in these systems. Pilocarpine base is less suitable for emulsion preparation than hydrochloride salt, and emulsions prepared at pH 5.0 show the most satisfying stability.     

Fine Multiple Emulsions Bearing 6-Mercaptopurine: In Vitro and In Vivo Antitumor Studies

Fine multiple emulsions were prepared by a two-step emulsification using the sonication technique. 6-mercaptopurine (6-MP) was encapsulated in the internal aqueous phase. It was coated with a phospholipid derivative of polyethylene glycol (PEG-PC). The coated multiple emulsion was characterized for antitumor activity on the murine leukemia cell line L-1210 in vitro. A decreased uptake and cytotoxicity were observed for PEG-PC-coated multiple emulsion in vitro, compared with uncoated ones. The IC₅₀ (50% inhibitory concentrated) was also decreased 1.64 times. In vivo antitumor activity was recorded as mean survival time (MST) of mice injected intravenously or intraperitoneally with tumor cells or, treated with 6-MP formulations through similar routes. MST was increased upon treatment. The normal peritoneal cells remained unchanged while blood parameters were restored with multiple emulsion treatment to tumor-bearing mice. The efficacy of the formulation was mainly due to the slow release and effective delivery of the drug from the formulation, suggesting its potential use for the treatment of cancer.     

Fine Multiple Emulsions Bearing 6-Mercaptopurine: In Vitro and In Vivo Antitumor Studies

Fine multiple emulsions were prepared by a two-step emulsification using the sonication technique. 6-mercaptopurine (6-MP) was encapsulated in the internal aqueous phase. It was coated with a phospholipid derivative of polyethylene glycol (PEG-PC). The coated multiple emulsion was characterized for antitumor activity on the murine leukemia cell line L-1210 in vitro. A decreased uptake and cytotoxicity were observed for PEG-PC-coated multiple emulsion in vitro, compared with uncoated ones. The IC₅₀ (50% inhibitory concentrated) was also decreased 1.64 times. In vivo antitumor activity was recorded as mean survival time (MST) of mice injected intravenously or intraperitoneally with tumor cells or, treated with 6-MP formulations through similar routes. MST was increased upon treatment. The normal peritoneal cells remained unchanged while blood parameters were restored with multiple emulsion treatment to tumor-bearing mice. The efficacy of the formulation was mainly due to the slow release and effective delivery of the drug from the formulation, suggesting its potential use for the treatment of cancer.     

In vitro release, pharmacokinetic and tissue distribution studies of doxorubicin hydrochloride (Adriamycin HCl) encapsulated in lipiodolized w/o emulsions and w/o/w multiple emulsions.

The lipiodolized w/o emulsion or w/o/w multiple emulsion containing Doxorubicin hydrochloride (1; Adriamycin HCl) with different emulsifiers was prepared to evaluate in vitro sustained-release behavior, pharmacokinetic and tissue distribution function in Sprague Dawley (SD) rats. The results of dissolution indicate that the release of 1 was significantly sustained for both emulsions when HCO-60 (polyoxyethylene (60) hydrogenated castor oil) was used as an emulsifier. The serum concentration of 1 was reduced and prolonged for both emulsions with the increase of HCO-60. The C(max) level was lowered and T(max) value was delayed after administration of w/o emulsions with higher HCO-60 concentration. The apparent terminal half-life for 1 released from some emulsions with higher concentration of HCO-60 was 3-folds higher than that of the 1 solution. The clearance of some w/o or w/o/w ADR emulsions also decreased with the increase of HCO-60. Not only the concentration of 1 in heart and kidney decreased significantly after the administration of w/o emulsions with the higher concentration of HCO-60, but also the hepatic concentration of 1 was higher and increased with HCO-60 concentration. The hepatic 1 level became lower after administration of w/o/w multiple emulsions with the increase of HCO-60; however, the concentration of 1 in heart, lung and spleen increased somewhat. The results indicate that lipiodol and HCO-60 seemed to play an important role in the prolongation and selective retention of w/o emulsion or w/o/w multiple emulsion, in vitro and in vivo.     

Multiple emulsions containing rifampicin

A stable multiple emulsion containing rifampicin in the internal aqueous phase was prepared by the incorporation of additives in both aqueous and oily phases. The formulation and process variables were optimized for primary and secondary emulsification. Drug release studies were performed on selected multiple emulsions to observe the effect of dilution. The release data were fitted to the Higuchi equation for slab and spherical geometry and effective diffusion coefficients were calculated. Stability studies for three months revealed good stability of the multiple emulsions with respect to creaming, phase separation, viscosity change, drug leakage, change in droplet size upon storage and exposure to osmotic and shear stress. The in vivo studies performed with stable multiple emulsions administered orally in human volunteers showed prolonged plasma drug levels. The multiple emulsions were found suitable for an improved tuberculosis therapy.     

Pharmacokinetics of Vancomycin after Intravenous Administration of a W/O/W Emulsion to Rats

A stable water-in-oil-in-water multiple emulsion (w/o/w emulsion) was prepared, and its potential for drug delivery was evaluated. W/o/w emulsions were prepared using a Lipiodol Ultra-Fluid(r) and isopropyl myristate oil mixture for the oily phase and vancomycin (VCM) for the entrapped drug. The surfactants, HCO-40 (5% [w/v]) and Pluronic F-88 were dissolved in the oily phase and the external aqueous phase, respectively. Resultant w/o/w emulsions were evaluated for entrapment efficiency, particle size, viscosity, drug release in vitro, and disposition kinetics of the drug and the w/o/w emulsion in vivo. The particle size of the w/o/w emulsion decreased with an increase in the concentration of F-88 in the external aqueous phase and stirring speed at the second emulsification stage (the smallest being 2.9 ± 1.5 μm). Entrapment efficiency of VCM in the w/o/w emulsion decreased with an increase in the concentration of F-88 (the maximum being 65.3 ± 5.4%). VCM release from the w/o/w emulsion was prolonged and tended to be slower with an increase in the particle size of the emulsion. After intravenous administration, significant differences in pharmacokinetic parameters, such as k₂₁, k_elβ, AUC₀-±, MRT₀-₆, and MRT₀-±, were observed between the VCM solution and the w/o/w emulsion-entrapped drug. When the w/o/w emulsion with Sudan II in the oily phase was administered intravenously, the emulsion accumulated in the lung at first (the highest value was observed just after administration) and then in the liver (the highest value was observed at 60 min). The w/o/w emulsion prepared in this study is expected to be a possible carrier for the prolonged release of water-soluble drugs after intravenous administration.     

Multiphase flow microfluidics for the production of single or multiple emulsions for drug delivery

Considerable effort has been directed towards developing novel drug delivery systems. Microfluidics, capable of generating monodisperse single and multiple emulsion droplets, executing precise control and operations on these droplets, is a powerful tool for fabricating complex systems (microparticles, microcapsules, microgels) with uniform size, narrow size distribution and desired properties, which have great potential in drug delivery applications. This review presents an overview of the state-of-the-art multiphase flow microfluidics for the production of single emulsions or multiple emulsions for drug delivery. The review starts with a brief introduction of the approaches for making single and multiple emulsions, followed by presentation of some potential drug delivery systems (microparticles, microcapsules and microgels) fabricated in microfluidic devices using single or multiple emulsions as templates. The design principles, manufacturing processes and properties of these drug delivery systems are also discussed and compared. Furthermore, drug encapsulation and drug release (including passive and active controlled release) are provided and compared highlighting some key findings and insights. Finally, site-targeting delivery using multiphase flow microfluidics is also briefly introduced.     

Inhibition of liver metastasis by all-trans retinoic acid incorporated into O/W emulsions in mice

All-trans retinoic acid (ATRA) was incorporated into lipid emulsions in an attempt to alter its distribution characteristics and improve its inhibition of liver cancer metastasis. Lipid emulsions composed of egg phosphatidylcholine, cholesterol, and soybean oil were the optimized carriers for ATRA delivery, as shown by the submicron particle size and high incorporation efficiency. The particle size and zeta potential of ATRA incorporated into emulsions were about 133 nm and -11 mV, respectively. In vitro drug release study demonstrated that the release of ATRA from emulsions was sustained in the absence and present of bovine serum albumin, suggesting that ATRA was stable when incorporated in emulsions. After intravenous administration in mice, [3H]cholesteryl hexadecyl ether incorporated into emulsion, which is the inherent distribution of emulsions, accumulated gradually mainly in the liver. The blood concentration and hepatic accumulation of [3H]ATRA incorporated into emulsion was significantly higher than that of serum dissolving [3H]ATRA, which represent the original distribution characteristic of free ATRA. In a murine liver metastasis model by colon adenocarcinoma, the liver metastasis number and liver weight were significantly reduced and the survival time of mice was prolonged following intravenous injection of ATRA incorporated into emulsions.     

Stability of total nutrient admixtures using various intravenous fat emulsions

The stability of four lipid emulsions with amino acids and dextrose in total nutrient admixtures (TNAs) was studied. The admixtures were divided into three groups. In group 1, 24 admixtures representing 20 different combinations of Liposyn II (safflower oil-soybean oil fat emulsion) with various manufacturers' amino acids (FreAmine III, Travasol, Novamine, Nephramine, and RenAmin) were tested. In group 2, 19 TNAs representing 14 combinations containing soybean-oil emulsions (Intralipid, Travamulsion, and Soyacal) and Aminosyn II amino acids were studied. In group 3, 14 TNAs representing 9 combinations containing the above soybean oil emulsions and Aminosyn II with Electrolytes were tested. Both 10% and 20% concentrations of fat emulsion, various amino acid concentrations ranging from 5.4% to 11.4%, and dextrose injections of 10, 20, 40, 50, and 70% were used. The admixtures were compounded in an ethylene vinyl acetate container. The mixing sequence involved transfer of fat emulsion to the empty container, followed by amino acids and dextrose. One of two electrolyte and trace metal profiles was added to each core admixture after compounding. Multivitamins were added just before the 24-hour room-temperature (25 +/- 4 degrees C) storage. Admixtures were tested initially and after one day at room temperature or nine days at 5 degrees C plus one day at room-temperature storage. Measurements of pH, emulsion particle size, osmolality, and zeta potential (electrostatic surface charge of lipid particles) were made after visual inspection of each admixture. In general, the TNAs retained a uniform, milk-like appearance under both storage conditions. The values of pH, zeta potential, particle size, and osmolality remained essentially unchanged throughout the study. Under the conditions of this study, the TNA formulations tested are stable for up to 10 days.     

Enhanced mucosal and systemic immune response with squalane oil-containing multiple emulsions upon intranasal and oral administration in mice

The objective of this study was to develop and evaluate squalane oil-containing water-in-oil-in-water (W/O/W) multiple emulsion for mucosal administration of ovalbumin (OVA) as a model candidate vaccine in BALB/c mice. Control and optimized OVA-containing W/O/W emulsion (OVA-Emul) and chitosan-modified W/O/W emulsion (OVA-Emul-Chi) formulations were administered intranasally and orally at an OVA dose of 100 mug. The mucosal and systemic immune responses were evaluated after the first and second immunization. The OVA-Emul formulations resulted in higher immunoglobulin-G (IgG) and immunoglobulin-A (IgA) responses as compared with aqueous solution. In addition, significant IgG and IgA responses were observed after the second immunization dose using the emulsions with both routes of administration. Intranasal vaccination was more effective in generating the systemic OVA-specific IgG response than the mucosal OVA-specific IgA response. Oral immunizations, on the other hand, showed a much higher systemic IgG and mucosal IgA responses as compared with the nasally treated groups. The results of this study show that squalane oil-containing W/O/W multiple emulsion formulations can significantly enhance the local and systemic immune responses, especially after oral administration, and may be adopted as a better alternative in mucosal delivery of prophylactic and therapeutic vaccines.