Pharmacokinetics and biodistribution study of self-assembled Gd-micelles demonstrating blood-pool contrast enhancement for MRI.

Magnetic resonance angiography (MRA) requires the use of contrast agents (CAs) to enable accurate diagnosis. There are currently no CAs on the market with appropriate pharmacokinetic (PK) parameters, namely long persistence in the blood, that can be easily used for MRA. We have recently synthesized amphiphilic building blocks loaded with gadolinium (Gd), which self-assemble into Gd-micelles in aqueous media, and have evaluated their potential as a blood-pool contrast agent (BPCA) in vivo. To assess the short and long term PK of Gd-micelles, the blood and organs of the mice were analyzed at t = 30 min, 1, 2, 3 h, 7, 14 and 21 days. Gd-DOTA was used as a control because it is the gold-standard CA for MRA despite its rapid clearance from the blood compartment. Gd-micelles circulated in the blood for more than 3 h postinjection whereas Gd-DOTA was eliminated less than half an hour postinjection. No side effects were observed in the mice up to the end of the study at 21 days and no accumulation of Gd was observed in the brain or bones. The Magnetic Resonance Imaging (MRI) parameters and the results of this in vivo study indicate the true BCPA properties of Gd-micelles and warrant further development.

Design, synthesis and in vitro evaluation of ß-glucuronidase-sensitive prodrug of 5-aminolevulinic acid for photodiagnosis of breast cancer cells.

Treatment of cancer cells by clinically approved hexyl ester of 5-aminolevulinic acid (ALA-Hex) induces accumulation of fluorescent porphyrins in tumors. This allows fluorescence photodiagnosis (PD) of bladder cancer by blue light illumination. However, PD of other cancers is hampered by acute toxicity of the compound limiting its use to local applications. We have designed and synthesized a new prodrug of ALA-Hex that tackles the stability-activity paradox of amino-modified 5-ALA prodrugs. The glucuronide prodrug Glu-ALA-Hex demonstrates excellent stability under physiological conditions and activation in the presence of the target enzyme. ß-glucuronidase-triggered release of 5-ALA is programmed to yield fluorescence in tumor environment with elevated ß-glucuronidase activity, a characteristic of many solid tumors. Glu-ALA-Hex produces similar levels of fluorescence as ALA-Hex in breast cancer MCF7 cells in vitro but with much lower non-specific cell toxicity.

Sonothrombolysis: the contribution of stable and inertial cavitation to clot lysis.

Microbubble-mediated sonothrombolysis (STL) is a remarkable approach to vascular occlusion therapy. However, STL remains a complex process with multiple interactions between clot, ultrasound (US), microbubbles (MB) and thrombolytic drug. The aim of this study was to evaluate the ability of combining US and MB to degrade fibrin and, more specifically, to assess the roles of both stable (SC) and inertial (IC) cavitation. Human blood clots containing radiolabeled fibrin were exposed to different combinations of recombinant tissue plasminogen activator (rtPA), US (1 MHz) and phospholipid MB. Three acoustic pressures were tested: 200, 350 and 1,300 kPa (peak-negative pressure). Clot lysis was assessed by diameter loss and release of radioactive fibrin degradation products. The combination rtPA + US + MB clearly revealed that IC (1,300 kPa) was able to enhance fibrin degradation significantly (66.3 ± 1.8%) compared with rtPA alone (51.7 ± 2.0%, p < 0.001). However, SC failed to enhance fibrin degradation at an acoustic pressure of 200 kPa. At 350 kPa, a synergistic effect between rtPA and US + MB was observed with an absolute increase of 6% compared to rtPA alone (p < 0.001). Conversely, without rtPA, the combination of US + MB was unable to degrade the fibrin network (0.3 ± 0.1%, p > 0.05 vs. control), but induced a distinct loss of red blood cells throughout the entire thickness of the clot, implying that MB were able to penetrate and cavitate inside the clot.

Tracking of antibody reduction fragments by capillary gel electrophoresis during the coupling to microparticles surface.

Due to their high specificity and efficiency, antibodies are ideal ligands for target-specific ultrasound contrast agents. The present study focuses on the chemical stability of antibodies during functionalisation with sulfosuccinimidyl-pyridyldithiopropionamidohexanoate (SPDP), a heterobifunctional linker, which exposes free thiol groups upon treatment with a reducing agent. Thiolated antibodies can then react with thiol-reactive group, such as maleimide present on the microbubble surface to form stable covalent complexes. The immunoglobulin structure relies on several intra- and inter-chain disulfide bridges which might be affected by reducing agents. A capillary electrophoresis-sodium dodecyl sulfate (CE-SDS) method with UV detection was applied to address the effect of the functionalisation process on the structural integrity of the antibodies and revealed that antibody disulfide bonds are prone to reduction as function of the reducing agents. Depending on the coupling conditions, various IgG fragments were identified reflecting different combinations between the light and heavy chains. Furthermore, two commonly used reducing agents, namely triscarboxyethylphosphine (TCEP) and 1,4-dithiothreitol (DTT) were compared under various preparation conditions. Results showed that reduction conditions based on DTT as a reducing agent under acidic pH were more appropriate to preserve intra- and inter-disulfide bridges of SPDP-modified antibodies.

Grafting of abciximab to a microbubble-based ultrasound contrast agent for targeting to platelets expressing GP IIb/IIIa - characterization and in vitro testing.

Abciximab-grafted ultrasound sensitive microbubbles were developed for the diagnosis of stroke. The antibody fragment abciximab, which binds to the GP IIb/IIIa and alpha v beta 3 receptors expressed by activated platelets, was chosen because most ischemic strokes are due to arterial thrombi that are mainly composed of platelets. The abciximab antibody fragment was activated by reduction of the disulfide bond for grafting on the microbubbles. The suspension was freeze-dried after the grafting was performed directly on the formed microbubbles. Quantification of the amounts of abciximab present on the surface of the microbubbles was assessed semi-quantitatively by flow cytometry, and quantitatively using radio-labeled abciximab. A protocol for human and rat platelets deposition and fixation was implemented and the expression of the GP IIb/IIIa receptor was validated by immunostaining. The abciximab-grafted microbubbles showed high static and dynamic binding to fixed platelets. Detection by ultrasonography of microbubbles bound on white and red clots gave higher signals compared to Sono Vue microbubbles.

Poly(D,L-lactide-co-glycolide) protein-loaded nanoparticles prepared by the double emulsion method--processing and formulation issues for enhanced entrapment efficiency.

Although extensive research in the field of biodegradable microparticles containing peptide or protein drugs has greatly advanced production know-how, the effects of critical parameters influencing successful drug entrapment have not yet been sufficiently investigated with nano-scaled carriers. This paper deals with the formulation and processing parameters of the w(1)/o/w(2) double emulsion method that can affect nanoparticle size and loading. Fluorescein isothiocyanate-labelled bovine serum albumin (FITC-BSA) was used as a model protein. Results showed that high FITC-BSA entrapment efficiencies were reached (>80%) when sonication was used for the two emulsification steps of the nanoparticle formation, independently of the mixing durations and intensities. By comparison, the use of a vortex mixer for obtaining the primary w(1)/o emulsion led to a rather poor entrapment efficiency (approximately 25%). Some inherent properties of the poly(D,L-lactic-co-glycolic acid) polymer, such as, for example, high molecular weight, high hydrophilicity or the presence of free carboxylic end groups, enhanced the drug entrapment efficiency. It was also demonstrated that a low nominal drug loading, a large volume of the inner w1 phase or the choice of methylene chloride instead of ethyl acetate as organic solvent favoured the drug entrapment, with entrapment efficiency values often reaching 100%. However, when using methylene chloride, the mean particle size was substantially increased, due to the presence of larger particles. Mean particle size increased also when the polymer concentration in the organic phase was increased.

In vitro and in vivo activities of verteporfin-loaded nanoparticles.

The main goal of this study was to develop a dispersed polymeric drug delivery system for verteporfin, suitable for intravenous administration and capable of improving its phototherapeutic index and minimizing the side effects. To achieve this objective, two types of verteporfin-loaded nanoparticles (167 and 370 nm in diameter) based on poly(D,L-lactide-co-glycolide) were prepared using the salting-out technique and were first tested on EMT-6 mammary tumor cells in comparison with an aqueous solution (DMSO/PBS). It was observed that small nanoparticles exhibited greater photocytotoxicity compared to large nanoparticles or DMSO/PBS, and the photocytotoxic efficiency was graded as small nanoparticles>DMSO/PBS>large nanoparticles. Furthermore, verteporfin, entrapped into small nanoparticles transferred to serum proteins more rapidly than when dissolved in DMSO/PBS. Drug clearance, measured by skin phototoxicity investigated in mice exposed to simulated sunlight 15 to 150 min after the injection of small nanoparticles was modest at early light exposure times with the small nanoparticles and diminished rapidly with later exposure times. Tumor bioassay results indicated that verteporfin incorporated into small nanoparticles effectively controlled tumor growth for 20 days in mice with early light irradiation times following drug administration.

Encapsulation of p-THPP into nanoparticles: cellular uptake, subcellular localization and effect of serum on photodynamic activity.

The cellular uptake, localization and efflux of meso-tetra-(4-hydroxyphenyl)porphyrin (p-THPP)-loaded nanoparticles have been studied in EMT-6 tumor cells. The effect of blood serum on photocytotoxicity has also been evaluated. Sub-130 nm nanoparticles based on poly(D,L-lactide-co-glycolide) (PLGA) (50:50 PLGA and 75:25 PLGA) and poly(D,L-lactide) (PLA) have been examined in comparison with free p-THPP. For all formulations tested, uptake of photosensitizer into cells was dependent on concentration, time and temperature. All nanoparticulate formulations accumulated within the cells to a greater extent relative to free drug. Indeed, the fluorescence intensities measured on EMT-6 cells treated with p-THPP-loaded nanoparticulate formulations were at least two-fold higher than those obtained with free dye. Furthermore, the highest accumulation level was found with PLGA nanoparticles. Fluorescence microscopy revealed that endocytosis is a major intracellular sequestration mechanism of these p-THPP formulations and that these were localized into early and late endosomes. The efflux study performed on both nonirradiated and irradiated cells indicated that free and p-THPP-loaded nanoparticles gradually escaped from EMT-6 cells as a function of time. This was more pronounced when cells were treated with nanoparticles and irradiated, reflecting important photodamage. It was also found that regardless of the nanoparticulate formulations tested, p-THPP photocytotoxicity was influenced by the concentration of the serum.

pH-Dependent dissolving nano- and microparticles for improved peroral delivery of a highly lipophilic compound in dogs.

RR01, a new highly lipophilic drug showing extremely low water solubility and poor oral bioavailability, has been incorporated into pH-dependent dissolving particles made of a poly(methacrylic acid-co-ethylacrylate) copolymer. The physicochemical properties of the particles were determined using laser-light-scattering techniques, scanning electron microscopy, high-performance liquid chromatography, and x-ray powder diffraction. Suspension of the free drug in a solution of hydroxypropylcellulose (reference formulation) and aqueous dispersions of pH-sensitive RR01-loaded nanoparticles or microparticles were administered orally to Beagle dogs according to a 2-block Latin square design (n = 6). Plasma samples were obtained over the course of 48 hours and analyzed by gas chromatography/mass spectrometry. The administration of the reference formulation resulted in a particularly high interindividual variability of pharmacokinetic parameters, with low exposure to compound RR01 (AUC0-48h of 6.5 microg x h/mL and coefficient of variation (CV) of 116%) and much higher Tmax, as compared to both pH-sensitive formulations. With respect to exposure and interindividual variability, nanoparticles were superior to microparticles (AUC0-48h of 27.1 microg x h/mL versus 17.7 microg x h/mL with CV of 19% and 40%, respectively), indicating that the particle size may play an important role in the absorption of compound RR01. The performance of pH-sensitive particles is attributed to their ability to release the drug selectively in the upper part of the intestine in a molecular or amorphous form. In conclusion, pH-dependent dissolving particles have a great potential as oral delivery systems for drugs with low water solubility and acceptable permeation properties.

Inhibition of HIV-1 in cell culture by oligonucleotide-loaded nanoparticles.

PURPOSE: To investigate the potential use of polymeric nanoparticles for the delivery of antisense oligonucleotides in HIV-1-infected cell cultures. METHODS: Phosphorothioate oligonucleotides were encapsulated into poly (D,L-lactic acid) nanoparticles. Two models of infected cells were used to test the ability of nanoparticles to deliver them. HeLa P4-2 CD4+ cells, stably transfected with the beta-galactosidase reporter gene, were first used to evaluate the activity of the oligonucleotides on a single-round infection cycle. The acutely infected lymphoid CEM cells were then used to evaluate the inhibition of the viral production of HIV-1 by the oligonucleotides. RESULTS: The addition to infected CEM cells of nanoparticles containing gag antisense oligonucleotides in the nanomolar range led to strong inhibition of the viral production in a concentration-dependent manner. Similar results were previously observed in HeLa P4-2 CD4+ cells. Nanoparticle-entrapped random-order gag oligonucleotides had similar effects on reverse transcription. However, the reverse transcriptase activity of infected cells treated with nanomolar concentrations of free antisense and random oligonucleotides was not affected. CONCLUSIONS: These results suggest that poly (D,L-lactic acid) nanoparticles may have great potential as an efficient delivery system for oligonucleotides in HIV natural target cells, i.e., lymphocytic cells.

Comparison of two methods of encapsulation of an oligonucleotide into poly(D,L-lactic acid) particles.

The aim of this study was to compare two methods to encapsulate a 25-mer-phosphorothioate oligonucleotide (ODN) into poly(D,L-lactic acid) (PLA) particles. Antisense oligonucleotides belong to a new therapeutic class especially attractive for the treatment of cancers and viral diseases. The development of these new drugs suffers, however, from poor stability in biological media and very low cellular uptake. Polymeric particulate systems display interesting features for ODN delivery. ODN are highly hydrophilic and most encapsulation methods are inappropriate for such molecules. Using poly(D,L-lactide) polymer, two methods of encapsulation were compared. First, a double emulsion technique was used to prepare nano- and microparticles. Secondly, the ODN was combined with a quaternary ammonium, the cethyltrimethyl-ammonium bromide (CTAB), to enhance the hydrophobicity of the molecule before entrapment by the emulsification-diffusion method. Both methods led to the formation of individualized and spherical particles loaded with a significant amount of ODN. Similar entrapment efficiencies were obtained for the nanoparticles prepared by both methods (approx. 27% of the theoretical loading) whereas 45% of entrapment efficiency was observed for the microparticles. Seventy five percent of the ODN were released in 60 min with the particles prepared by the emulsification-diffusion method, whereas only 7% were released in 60 h when using the double emulsion method. A viability test on U-937 cells showed better survival rates with the particles prepared by the double emulsion technique. The results suggest that the location of the ODN in the polymeric matrix is affected by the encapsulation method. Particles containing CTAB appeared more toxic than the ones obtained by the double emulsion technique, however, these particles can still be used for antisense activity since high oligonucleotide loading can be achieved.

Freeze-drying and lyopreservation of diblock and triblock poly(lactic acid)-poly(ethylene oxide) (PLA-PEO) copolymer nanoparticles.

In this study, the formulation and process parameters that determine successful production and long-term stability of freeze-dried poly(lactic acid) (PLA) nanoparticles with "hairy-like" poly(ethylene oxide) (PEO) surfaces were investigated. Nanoparticles with grafted (covalently bound) PEO coatings were produced by the salting-out method from blends of PLA and PLA-PEO diblock or triblock copolymers. PLA nanoparticles with physically adsorbed PEO were also produced. The redispersibility of the nanoparticles after freeze-drying under various conditions was assessed. The surface of the nanoparticles was characterized and classified in terms of "brush" and "loop" conformations. Upon freeze-drying, it appeared that the presence of PEO at the nanoparticle surface could severely impair the redispersibility of the particles, especially in the PEO-grafted systems. This effect was shown to be related to the amount and molecular weight of PEO in the various formulations. In most cases, particle aggregation was prevented by use of trehalose as lyoprotective agent. Increasing the concentration of particles in the suspension to be freeze-dried was shown to induce much less damage to the nanoparticles, and freezing the suspension at a very low temperature (-196 degrees C) was found to further improve the lyoprotective effect. Most of the lyoprotected nanoparticles remained stable for at least 12 weeks at 4 and -25 degrees C. The production and preservation of freeze-dried PLA-PEO diblock and triblock copolymer nanoparticles is feasible under optimized lyoprotective conditions.

Cellular uptake of PEO surface-modified nanoparticles: evaluation of nanoparticles made of PLA:PEO diblock and triblock copolymers.

Nanoparticles with either physically adsorbed or covalently bound poly(ethylene oxide) (PEO) coatings were produced from various combinations of poly(lactic acid) (PLA) and diblock or triblock copolymers of PLA and PEO. The particles were produced by the salting-out process and purified by the cross-flow filtration technique. The amount of PEO at the nanoparticle surface, as well as the residual amount of emulsifier poly(vinyl alcohol) were assessed, with a good correlation with expected values. Stability of the nanoparticulate suspensions was studied at 4 degrees C and after freezing under various conditions for up to 6 months. The nanoparticle redispersibility after storage was related to the thermal behavior of the PEO coatings. The in vitro cellular uptake of the different types of nanoparticles was compared by flow cytometry after incubation with human monocytes in serum and in plasma. The influence of the PEO molecular weight and surface density on the particle uptake was especially marked for the diblock and triblock copolymer formulations, with a decrease in uptake of up to 65% with one of the diblock copolymer formulations. Nanoparticles made of triblock copolymer with short PEO chains at their surface in the postulated "loop conformation" proved to be as resistant to cellular uptake as nanoparticles made of diblock copolymers with PEO chains in the "brush conformation".

Oral bioavailability of a poorly water soluble HIV-1 protease inhibitor incorporated into pH-sensitive particles: effect of the particle size and nutritional state.

The new chemical entity CGP 70726, a very poorly water-soluble HIV-1 protease inhibitor, was incorporated into pH-sensitive nanoparticles and microparticles made of the poly(methacrylic acid-co-ethylacrylate) copolymer Eudragit((R)) L100-55. The particles were characterized in terms of morphology, size distribution, drug loading, production yield and dispersion state of the drug inside the polymeric matrices. Aqueous dispersions of the particles were administered orally to Beagle dogs against a suspension of free drug (control formulation) all at a dose of 100 mg/kg. Oral administration was conducted in the absence and presence of food. Plasma concentrations and pharmacokinetic parameters were determined within 8 h post-dose. While no measurable absorption of the drug resulted after administration of the control formulation, substantial systemic exposure to the compound was obtained with both kinds of pH-sensitive formulations. The selective release of CGP 70726 in a highly dispersed/amorphous state and creation of high concentrations close to its absorption site was thought to account for this positive result. The largest areas under the plasma concentration-time curve (AUC) were obtained in the fasted state, with slightly better performance of the microparticles over the nanoparticles, in both nutritional states (7.8+/-1.5 versus 5.8+/-0. 8 micromol.h/l in the fasted state; 4.4+/-1.4 versus 2.00+/-0.5 micromol.h/l in the fed state). With these results, the potential of pH-sensitive particles for the oral delivery of HIV-1 protease inhibitors with low water solubility was confirmed.

Pseudolatex preparation using a novel emulsion-diffusion process involving direct displacement of partially water-miscible solvents by distillation.

Pseudolatexes were obtained by a new process based on an emulsification-diffusion technique involving partially water-miscible solvents. The preparation method consisted of emulsifying an organic solution of polymer (saturated with water) in an aqueous solution of a stabilizing agent (saturated with solvent) using conventional stirrers, followed by direct solvent distillation. The technique relies on the rapid displacement of the solvent from the internal into the external phase which thereby provokes polymer aggregation. Nanoparticle formation is believed to occur because rapid solvent diffusion produces regions of local supersaturation near the interface, and nanoparticles are formed due to the ensuing interfacial phase transformations and polymer aggregation that occur in these interfacial domains. Using this method, it was possible to prepare pseudolatexes of biodegradable and non-biodegradable polymers such as poly(D,L-lactic acid) and poly(epsilon-caprolactone), Eudragit E, cellulose acetate phthalate, cellulose acetate trimellitate using ethyl acetate or 2-butanone as partially water-miscible solvents and poly(vinyl alcohol) or poloxamer 407 as stabilizing agent. A transition from nano- to microparticles was observed at high polymer concentrations. At concentrations above 30% w/v of Eudragit E in ethyl acetate or cellulose acetate phthalate in 2-butanone only microparticles were obtained. This behaviour was attributed to decreased transport of polymer molecules into the aqueous phase.

Formulation and lyoprotection of poly(lactic acid-co-ethylene oxide) nanoparticles: influence on physical stability and in vitro cell uptake.

PURPOSE: To investigate the feasibility of producing freeze-dried poly(ethylene oxide) (PEO)-surface modified nanoparticles and to study their ability to avoid the mononuclear phagocytic system (MPS), as a function of the PEO chain length and surface density. METHODS: The nanoparticles were produced by the salting-out method using blends of poly(D,L-lactic acid) (PLA) and poly(D,L-lactic acidco-ethylene oxide) (PLA-PEO) copolymers. The nanoparticles were purified by cross-flow filtration and freeze-dried as such or with variable amounts of trehalose as a lyoprotectant. The redispersibility of the particles was determined immediately after freeze-drying and after 12 months of storage at -25 degrees C. The uptake of the nanoparticles by human monocytes was studied in vitro by flow cytometry. RESULTS: PLA-PEO nanoparticles could be produced from all the polymeric blends used. Particle aggregation after freeze-drying was shown to be directly related to the presence of PEO. Whereas this problem could be circumvented by use of trehalose, subsequent aggregation was shown to occur during storage. These phenomena were possibly related to the specific thermal behaviours of PEO and trehalose. In cell studies, a clear relationship between the PEO content and the decrease of uptake was demonstrated. CONCLUSIONS: The rational design of freeze-dried PEO-surface modified nanoparticles with potential MPS avoidance ability is feasible by using the polymer blends approach combined with appropriate lyoprotection and optimal storage conditions.

Uptake of oligonucleotide-loaded nanoparticles in prostatic cancer cells and their intracellular localization.

The development of antisense biotechnology is dependent, in part, on creating improved methods for delivering oligonucleotides to cells. In this study, we investigated a colloidal system (nanoparticles (NP) of poly (D,L) lactic acid) that affects the intracellular delivery of oligonucleotides. We have examined the intracellular compartmentalization in DU145 cells of fluorescein labeled phosphorothioate oligonucleotides, both in the free state and when loaded into NP. Fluorescent oligonucleotides were incubated for 18 h with DU145 cells and the mean intracellular fluorescence was determined by flow cytometry. After the addition of monensin, an increase in signal intensity was observed, indicating that free oligonucleotides were resident in an acidic intracellular environment, whereas oligonucleotides from the NP did not reside in an acidic compartment. Free and NP loaded with oligonucleotides effluxed from DU145 cells from two intracellular compartments. This preliminary report indicates that colloidal carriers such as NP could prove to be useful in affecting intracellular trafficking of oligonucleotides in DU145 and in other cells.

Highly loaded nanoparticulate carrier using an hydrophobic antisense oligonucleotide complex.

Antisense oligonucleotides, and particularly those with phosphorothioate backbones, have emerged as potential gene specific therapeutic agents and are currently undergoing evaluation in clinical trials for a variety of diseases. In the area of HIV-1 therapeutics, targeting of oligonucleotides to infected cells, such as macrophages, would be highly desirable. The present study was designed to prepare and characterize oligonucleotide-loaded nanoparticles for this purpose. Due to their hydrophilic characteristics, oligonucleotides are difficult to entrap in polymeric particles. Here, the oligonucleotides were first complexed with cetyltrimethylammonium bromide. The oligonucleotide-loaded nanoparticles were prepared by the emulsification-diffusion method and subsequently purified. In comparison with previous studies, a high oligonucleotide-loading was achieved; 2.5, 5 and 10% oligonucleotide loading were assessed. If the initial oligonucleotide content was 4%, this method produced a final oligonucleotide loading of 1.9% with an entrapment efficiency of 47%. The integrity of the oligonucleotide and of the polymer, in the final freeze-dried product, was retained.

Preparation and characterization of nanocapsules from preformed polymers by a new process based on emulsification-diffusion technique.

PURPOSE: The aim of this study was to investigate whether biodegradable nanocapsules could be obtained by the emulsification-diffusion technique. METHODS: This technique consists of emulsifying an organic solution containing an oil, a polymer, and a drug in an aqueous solution of a stabilizing agent. The subsequent addition of water to the system induces solvent diffusion into the external phase, resulting in the formation of colloidal particles. Nanoparticles obtained in this way were characterized by their particle size, zeta potential, isopycnic density and drug entrapment. The shape, surface and structure of the nanocapsules were evaluated by freeze fracture scanning electron microscopy (SEM) and by atomic force microscopy (AFM). RESULTS: Density gradient centrifugation confirmed the formation of nanocapsules. The density was found to be intermediate between those of nanoemulsions and nanospheres. The existence of a unique density band indicated high yields. Nanocapsule density was a function of the original oil/polymer ratio, revealing that the polymer content and, consequently, the wall thickness, can be controlled by this method. SEM and AFM showed the presence of capsular structures with smooth homogeneous walls. The versatility and effectiveness of the method were demonstrated using different lipophilic drug/oil core/wall polymer/partially water-miscible solvent systems. The mechanism of nanocapsule formation was explained as a chemical instability (diffusion stranding) generated during diffusion. CONCLUSIONS: This study demonstrated that the emulsification-diffusion technique enables the preparation of nanocapsules in a simple, efficient, reproducible and versatile manner.

Comparative pharmacokinetic study of a floating multiple-unit capsule, a high-density multiple-unit capsule and an immediate-release tablet containing 25 mg atenolol.

The purpose of this study was to evaluate the possible advantages of floating and high-density dosage forms and their influence on pharmacokinetic parameters. Atenolol was chosen as a model drug because of its poor absorption in the lower gastrointestinal tract. Three formulations containing 25 mg atenolol, a floating multiple-unit capsule, a high-density multiple-unit capsule, and an immediate-release tablet were compared with respect to estimated pharmacokinetic parameters. The two multiple-unit dosage forms were composed of compressed minitablets and had sustained release properties. The bioavailability of the two gastroretentive preparations with sustained release characteristics was significantly decreased when compared to the immediate-release tablet. The floating minitablets seemed to be retained longer in the stomach than the high-density dosage form. The first atenolol concentration detectable in the plasma and the time to peak Tmax were delayed for the floating dosage form. For the parameters Cmax and AUC 0-infinity, the lower limit of the 90% confidence interval was outside the bioequivalence range (0.80-1.25). This study showed that it was not possible to increase the bioavailability of a poorly absorbed drug such as atenolol using gastroretentive formulations. Atenolol absorption was delayed and the maximum plasma concentration was diminished.