Protection of dopamine towards autoxidation reaction by encapsulation into non-coated- or chitosan- or thiolated chitosan-coated-liposomes.

The aim of this work is to evaluate the potential of non-coated-, chitosan-(CS)- or chitosan-glutathione conjugate- (CS-GSH)-coated liposomes to protect the neurotransmitter Dopamine (DA) from the autoxidation reaction in neutral/alkaline conditions. This may be of interest in the development of nanotechnology-based approaches to improve Parkinson's disease treatment because decreased ROS production and reduced DA associated neurotoxicity are expected. For the mentioned purposes, DA-loaded vesicles were prepared by the Dried Reconstituted Vesicles (DRV) method, and were subsequently coated using solutions of polycations. As for the mean diameters of liposomes so prepared, the CS-GSH coated liposomes showed a significant decrease in size compared to the corresponding non-coated and CS-coated vesicles. The surface charge of DA-loaded non-coated liposomes was -10.8 mV, whereas the CS or CS-GSH coated vesicles showed a slightly positive ζ-potential. The capability of the herein studied vesicles to prevent DA autoxidation was evaluated by visual inspection, monitoring DA/lipid ratio as such and under stressed conditions. The results suggest that liposome formulations partially protect the neurotransmitter from the autoxidation reaction. In particular, the CS-GSH coated liposomes were more stable than the corresponding CS-coated and non-coated ones against the oxidative damage and were found to deliver the neurotransmitter in a sustained manner. Probably, this is due to the localization of the neurotransmitter in the core of the vesicles as indicated by XPS which confirmed the absence of the neurotransmitter on the surface of these vesicles.

Sustained release of intravitreal flurbiprofen from a novel drug-in-liposome-in-hydrogel formulation.

A novel Flurbiprofen (FLB)-in-liposome-in-hydrogel formulation was developed, as a method to sustain the release and increase the ocular bioavailability of FLB following intravitreal injection. For this, FLB loading into liposomes was optimized and liposomes were entrapped in thermosensitive hydrogels consisted of Pluronic F-127 (P). FLB solution, liposomes, and FLB dissolved in hydrogel were also used as control formulations. Actively loaded liposomes were found to be optimal for high FLB loading and small size, while in vitro studies revealed that P concentration of 18% (w/v) was best to retain the integrity of the hydrogel-dispersed liposome, compared to a 20% concentration. The in vitro release of FLB was significantly sustained when FLB-liposomes were dispersed in the hydrogel compared to hydrogel dissolved FLB, as well as the other control formulations. In vivo studies were carried out in pigmented rabbits which were injected through a 27G needle with 1mg/mL FLB in the different formulation-types. Ophthalmic examinations after intravitreal injection of all FLB formulations, revealed no evidence of inflammation, hemorrhage, uveitis or endophthalmitis. Pharmacokinetic analysis results confirm that the hybrid drug delivery system increases the bioavailability (by 1.9 times compared to solution), and extends the presence of the drug in the vitreous cavity, while liposome and hydrogel formulations demonstrate intermediate performance. Furthermore the hybrid system increases MRT of FLB in aqueous humor and retina/choroid tissues, compared to all the control formulations. Currently the potential therapeutic advances of FLB sustained release formulations for IVT administration are being evaluated.

Covalent immobilization of liposomes on plasma functionalized metallic surfaces.

A method was developed to functionalize biomedical metals with liposomes. The novelty of the method includes the plasma-functionalization of the metal surface with proper chemical groups to be used as anchor sites for the covalent immobilization of the liposomes. Stainless steel (SS-316) disks were processed in radiofrequency glow discharges fed with vapors of acrylic acid to coat them with thin adherent films characterized by surface carboxylic groups, where liposomes were covalently bound through the formation of amide bonds. For this, liposomes decorated with polyethylene glycol molecules bearing terminal amine-groups were prepared. After ensuring that the liposomes remain intact, under the conditions applying for immobilization; different attachment conditions were evaluated (incubation time, concentration of liposome dispersion) for optimization of the technique. Immobilization of calcein-entrapping liposomes was evaluated by monitoring the percent of calcein attached on the surfaces. Best results were obtained when liposome dispersions with 5mg/ml (liposomal lipid) concentration were incubated on each disk for 24h at 37°C. The method is proposed for developing drug-eluting biomedical materials or devices by using liposomes that have appropriate membrane compositions and are loaded with drugs or other bioactive agents.

Large-scale effects on resistivity index of porous media.

The estimation of humidity in the unsaturated zone of soils and NAPL saturation in contaminated aquifers may be based on the interpretation of electrical resistivity index logs. In the present work, concepts of the theory of the two-phase flow in pore networks are employed to interpret the form of the equilibrium and dynamic resistivity index curves of large porous samples. A resistivity cell is constructed to measure the capillary and electrical properties of large samples of unconsolidated porous media. The drainage capillary pressure and resistivity index curves of a sand column are measured by using the micropore membrane (porous plate) method, where a 0.5% wt/vol NaCl aqueous solution is displaced by n-dodecane. The dynamic resistivity index curves are measured by using the continuous injection technique for various orientations of the sand column. Based on concepts of the two-phase flow theory, concerning the dominant displacement growth pattern in a pore network and arising from the cooperative effects of capillary, buoyancy, and viscous forces, approximate relationships are developed for the resistivity index and saturation exponent as functions of the water saturation. The saturation exponent decreases as the displacement advances and the fluid distribution across the sand column tends to be homogenized after oil breakthrough. Both the resistivity index and saturation exponent increase as the displacement pattern tends to become compact and stable. In the destabilized flow pattern, as the Bond number decreases, the resistivity index may increase respectably within a narrow range of values of the Bond number. This happens when the thickness of the unstable capillary finger exceeds the lateral dimension of the porous sample and becomes a fractal percolation cluster. The saturation exponent becomes almost constant and independent of water saturation only over the destabilized displacement pattern at high values of the Bond number.

The effect of pH on the electrophoretic behaviour of a new class of liposomes: arsonoliposomes.

Herein we report the effect of pH on the surface charge of a new class of liposomes: arsonoliposomes. Plain or mixed arsonoliposomes with cholesterol (Chol) and distearoyl-phosphatidylcholine (DSPC) in 1:1 molar ratio were prepared with lauryl-(C12), myristoyl-(C14) and palmitoyl-(C16) acyl side chain arsonolipids. The one step hydration method was used for vesicle preparation and zeta potential measurements were performed in the pH range from 3 to 9. The results revealed that these lipids hold a negative surface charge at all pH values investigated. The presence of cholesterol in 1:1 molar ratio results in higher zeta potential compared with plain arsonoliposomes with the exception of palmitoyl-(C16) acyl chain arsonolipids in neutral and slightly basic pH values. Oppositely, the DSPC (1:1 molar ratio) containing arsonoliposomes had lower values of zeta potential compared with plain arsonoliposomes. Concluding, the experimental results reveal that zeta potential of arsonoliposomes is indeed modified when the vesicles are incubated in environments with different acidity. In most cases these changes are in accordance with the ionization pattern of the arsonolipid headgroup, while some peculiar deviations may be connected with the known difference in the structure between some of the vesicle types studied.

Effect of copolymer composition on the physicochemical characteristics, in vitro stability, and biodistribution of PLGA-mPEG nanoparticles.

The physicochemical properties, the colloidal stability in vitro and the biodistribution properties in mice of different PLGA-mPEG nanoparticle compositions were investigated. The nanoparticles were prepared by a precipitation-solvent evaporation technique. The physical characteristics and the colloidal stability of the PLGA-mPEG nanoparticles were significantly influenced by the composition of the PLGA-mPEG copolymer used to prepare the nanoparticles. PLGA-mPEG nanoparticles prepared from copolymers having relatively high mPEG/PLGA ratios were smaller and less stable than those prepared from copolymers having relatively low mPEG/PLGA ratios. All PLGA-mPEG nanoparticle compositions exhibited prolonged residence in blood, compared to the conventional PLGA nanoparticles. The composition of the PLGA-mPEG copolymer affected significantly the blood residence time and the biodistribution of the PLGA-mPEG nanoparticles in liver, spleen and bones. The in vivo behavior of the different PLGA-mPEG nanoparticle compositions did not appear to correlate with their in vitro stability. Optimum mPEG/PLGA ratios appeared to exist leading to long blood circulation times of the PLGA-mPEG nanoparticles. This may be associated with the effects of the mPEG/PLGA ratio on the density of PEG on the surface of the nanoparticles and on the size of the nanoparticles.

PLGA-mPEG nanoparticles of cisplatin: in vitro nanoparticle degradation, in vitro drug release and in vivo drug residence in blood properties.

The in vitro nanoparticle degradation, in vitro drug release and in vivo drug residence in blood properties of PLGA-mPEG nanoparticles of cisplatin were investigated. The nanoparticles were prepared by a double emulsion method and characterized with regard to their morphology, size, zeta potential and drug loading. The rate of in vitro degradation of the PLGA-mPEG nanoparticles in PBS (pH 7.4) depended on their composition, increasing when the mPEG content (mPEG:PLGA ratio) of the nanoparticles increased. Sustained cisplatin release over several hours from the PLGA-mPEG nanoparticles in vitro (PBS) was observed. The composition of the nanoparticles affected drug release: the rate of release increased when the mPEG content of the nanoparticles increased. Within the range of drug loadings investigated, the drug loading of the nanoparticles did not have any significant effect on drug release. The loading efficiency was low and needs improvement in order to obtain PLGA-mPEG nanoparticles with a satisfactory cisplatin content for therapeutic application. The i.v. administration of PLGA-mPEG nanoparticles of cisplatin in BALB/c mice resulted in prolonged cisplatin residence in systemic blood circulation. The results appear to justify further investigation of the suitability of the PLGA-mPEG nanoparticles for the controlled i.v. delivery and/or targeting of cisplatin.

Capability of arsonolipids to transport divalent cations: a Pressman cell study.

The ability of the newly synthesized arsonolipids (2,3-diacyloxyprophlarsonic acids) to transport cations was studied using the Pressman cell. Experimental results demonstrate that arsonolipids are much more efficient carriers of Ca(2+) and Mg(2+) than natural phosphatidic acid in the Pressman cell experiments. The ability of arsonolipids to transfer Ca(2+) is affected by the lipid side chain length in the order: C(12)>>C(14) approximately C(16). Ca(2+) is transferred faster than Mg(2+), suggesting that the latter is more tightly bound to the arsonolipids. The transfer kinetic curves are parabolic for C(12), while initially linear with a tendency to reach a steady state for C(14) and C(16), when the pH in the donor compartment was 8.3. The transport kinetics for both ions studied were best fitted by an equation derived from saturation kinetics that apply in reversible chemical reactions. The ion transfer rates increased as the pH in the donor compartment decreased.

Preparation and properties of arsonolipid containing liposomes.

Arsonolipids are analogs of phosphonolipids which have a chemically versatile head group. In preliminary cell culture studies, liposomes composed solely of arsonolipids or of phosholipid-arsonolipid mixtures, demonstrate a specific toxicity against cancer cells (Gortzi et al., unpublished results). The possibility of using such formulations as an alternative of arsenic trioxide with or without combination of other cytostatic agents (encapsulated in their aqueous interior) prompted the investigation of their physicochemical characteristics. Herein we compared the characteristics of arsonolipid containing vesicles with different lipid compositions. Experimental results and morphological observations reveal that non-sonicated formulations have different structures and stability (when both membrane integrity and aggregation are taken into account) depending on the acyl chain length of the arsonolipid. When phospholipids and especially cholesterol are included in their membranes almost all arsonolipids studied produce more stable vesicles. An interesting aspect of these arsonolipid containing vesicles is also their negative surface charge, which may be modulated by mixing phospholipids with arsonolipids. Sonicated vesicles have smaller sizes and profoundly higher stability, especially when containing cholesterol and phosphatidylcholine mixed with arsonolipids. The only exception is that of the arsonolipid with the C(12) acyl chain which was observed to produce long tubes which break down to cubes by sonication. In conclusion, these initial studies demonstrate that sonicated vesicles composed of arsonolipid and phospholipid mixtures mixed with cholesterol posses the stability required to be used as an arsonolipid delivery system. In addition, although cryo-electron microscopy demonstrated that the sonicated vesicles are elliptical in shape, their encapsulation efficiency is not significantly lower than sonicated phospholipid liposomes. Thereby, these vesicles may be also used for the delivery of other drug molecules which can be sufficiently retained in their aqueous interior.

Effect of preparative variables on the properties of poly(dl-lactide-co-glycolide)-methoxypoly(ethyleneglycol) copolymers related to their application in controlled drug delivery.

The effect of certain preparative variables, such as the composition of the feed, the reaction time and the reaction temperature, on the properties of prepared poly(dl-lactide-co-glycolide)-methoxypoly(ethyleneg lycol) (PLGA-mPEG) copolymers and on the yield of the reaction was investigated. The results with regard the molecular weight and yield were discussed in relation to a polymerization mechanism proposed recently (Du et al., 1995. Macromolecules 28, 2124-2132). The higher the PEG content of the feed the lower the molecular weight of the copolymer and the yield of the reaction. The breadth of the molecular weight distribution decreased initially with time, but appeared to stabilize later at low values. Both the ethylene oxide content and the lactide to glycolide molar ratio in the copolymer depended on the reaction temperature and varied with the reaction time. PLGA and mPEG appeared to be partially miscible, and copolymers containing approximately 40% mol or higher ethylene oxide exhibited crystallinity.

A model system for the investigation of urinary stone formation.

OBJECTIVE: To develop an in vitro model system for the investigation of calcium oxalate urinary stones under conditions of spontaneous precipitation. MATERIALS AND METHODS: Using a calcium-ion selective electrode and automatic titrator, a test solution was kept supersaturated with calcium oxalate and the rate of crystallization measured. RESULTS: During the spontaneous precipitation at constant supersaturation at 37 degrees C, the induction times required for the precipitation of calcium oxalate monohydrate were inversely proportional to the supersaturation of the solution. No transient phases were identified and the interfacial energy determined from kinetic analysis was 28.4 mJ/m2. The rates of precipitation showed a first-order dependence on the degree of supersaturation and were in good agreement with those reported for the in vivo formation of calcium oxalate monohydrate stones. CONCLUSION: This experimental model system allows precise measurements of the kinetics of calcium oxalate monohydrate. From the dependence of the rates of precipitation on supersaturation, a mechanism controlled by surface diffusion is suggested.