Drug Permeation Studies Research Articles

Alginate/chitosan based bi-layer composite membrane as potential sustained-release wound dressing containing ciprofloxacin hydrochloride

Abstract The aims of this research were to develop and evaluate a novel ciprofloxacin hydrochloride loaded bi-layer composite membrane based on alginate and chitosan. In vitro antimicrobial activity, drug permeation study, morphology, cytotoxicity, primary skin irritation and in vivo pharmacodynamics were investigated. Results showed that the membranes could inhibit the growth of microorganisms for longer than 7 days. And there was no significant decrease in the metabolic activity of the Hacat fibroblasts cells were treated with the membranes. No edema and erythema were observed after administration of membranes on the rabbit skin after 14 days. Moreover, the results of pharmacodynamics showed that the membranes were more effective in improving the wound healing process. In conclusion, a novel bi-layer composite membrane was developed and results suggested that it could be exploited as sustained-release wound dressings.

COMPARATIVE ASSESSMENT OF CHLORPHENIRAMINE MALEATE PERMEATION THROUGH CELLULOSE ACETATE MEMBRANE AND SHEEP NASAL MUCOSA

The present research work was aimed to develop nasal mucoadhesive microspheres of chlorpheniramine maleate (CPM) & to compare its permeation rate through sheep nasal mucosa and cellulose acetate membrane. The impact of different concentrations of polymer and the dispersion media were evaluated for drug permeation, mucoadhesion, FT-IR and DSC studies. With the increase in polymer concentration, it was observed that the particle size and wall thickness of the microsphere increased; while the in-vitro drug permeation through sheep nasal mucosa significantly decreased when the volume of dispersion media was increased. Histopathological studies show the absence of epithelial cilia destruction. The permeation rate through sheep nasal mucosa was found to be 7.968 µg/hr/cm2, while through cellulose acetate (artificial) membrane showed 8.572 μg/hr/cm2. In conclusion, our results indicate that CPM released from chitosan microspheres penetrate well through sheep nasal mucosa, indicating suitability for nasal administration.

A comparative ex vivo drug permeation study of beta-blockers through porcine buccal mucosa

Apparent permeability coefficients (kp) of a series of beta-blockers: acebutolol, atenolol, labetalol, metoprolol, oxprenolol and propranolol, through porcine buccal mucosa were determined. The aim of the study was to determine the permeation parameters (apparent permeability coefficient, kp; flux, J; and lag time, TL) as a measure of the intrinsic permeability of porcine buccal mucosa to these drugs, in order to predict the efficacy of their possible administration through human buccal mucosa. A positive linear correlation was observed between the apparent permeability coefficient, kpand the partition coefficient, P. Oxprenolol and propranolol are the drugs that presented the highest values of kp: 0.3231×10(2) cm/h and 0.5666×10(2) cm/h, respectively. Multiple linear regression (MLR) using least square estimation was performed on the data set with logkpas dependent variable and the descriptors as predictor variables. The potential systemic capacity after a buccal administration was predicted by estimating the plasma concentrations at steady-stated (Css). Considering the entire process of permeation ex vivo, propranolol and oxprenolol would seem to be the best candidates for administration through the buccal mucosa.

A microdevice for parallelized pulmonary permeability studies

We describe a compartmentalized microdevice specifically designed to perform permeability studies across a model of lung barrier. Epithelial cell barriers were reproduced by culturing Calu-3 cells at the air-liquid interface (AIC) in 1 mm² microwells made from a perforated glass slide with an embedded porous membrane. We created a single basolateral reservoir for all microwells which eliminated the need to renew the growth medium during the culture growth phase. To perform drug permeability studies on confluent cell layers, the cell culture slide was aligned and joined to a collection platform consisting in 35 μL collection reservoirs connected at the top and bottom with microchannels. The integrity and functionality of the cell barriers were demonstrated by measurement of trans-epithelial electrical resistance (TEER), confocal imaging and permeability assays of ¹⁴C-sucrose. Micro-cell barriers were able to form confluent layers in 1 week, demonstrating a similar bioelectrical evolution as the Transwell systems used as controls. Tight junctions were observed throughout the cell-cell interfaces, and the low permeability coefficients of ¹⁴C-sucrose confirmed their functional presence, creating a primary barrier to the diffusion of solutes. This microdevice could facilitate the monitoring of biomolecule transport and the screening of formulations promoting their passage across the pulmonary barrier, in order to select candidates for pulmonary administration to patients.

Taguchi design for optimization and development of antibacterial drug-loaded PLGA nanoparticles

This research report was to develop Cefixime loaded polylactide-co-glycolide (PLGA) nanoparticles using modified precipitation method. TEM analysis indicated formation of well-formed, smooth, spherical nanoparticles with no aggregates whereas XRD recommended dispersion of drug in PLGA carrier system in amorphous form. The polymer and stabilizer concentration and organic to aqueous ratio were found to be significant factors for nanoparticles and their optimization using Taguchi design (L9). The design formulations showed entrapment efficiency (EE), particle size and poly-dispersity index (PDI) ranging 68.31±1.74%, 159.8–157.7nm and 0.126–0.149, respectively indicated small and stable nanoparticles with good homogeneity and encapsulation. The design optimized formulation drug release and permeation studies demonstrated that it is four times sustained release behavior and 1.74 times better permeation than free drug. The result of microbiological assay also suggested that optimized formulation has significant antibacterial activity against intracellular multidrug resistance (MDR) of Salmonella typhi.

Comparative study of four immortalized human brain capillary endothelial cell lines, hCMEC/D3, hBMEC, TY10, and BB19, and optimization of culture conditions, for an in vitro blood–brain barrier model for drug permeability studies

BackgroundReliable human in vitro blood–brain barrier (BBB) models suitable for high-throughput screening are urgently needed in early drug discovery and development for assessing the ability of promising bioactive compounds to overcome the BBB. To establish an improved human in vitro BBB model, we compared four currently available and well characterized immortalized human brain capillary endothelial cell lines, hCMEC/D3, hBMEC, TY10, and BB19, with respect to barrier tightness and paracellular permeability. Co-culture systems using immortalized human astrocytes (SVG-A cell line) and immortalized human pericytes (HBPCT cell line) were designed with the aim of positively influencing barrier tightness.MethodsTight junction (TJ) formation was assessed by transendothelial electrical resistance (TEER) measurements using a conventional epithelial voltohmmeter (EVOM) and an automated CellZscope system which records TEER and cell layer capacitance (CCL) in real-time.Paracellular permeability was assessed using two fluorescent marker compounds with low BBB penetration (sodium fluorescein (Na-F) and lucifer yellow (LY)). Conditions were optimized for each endothelial cell line by screening a series of 24-well tissue culture inserts from different providers. For hBMEC cells, further optimization was carried out by varying coating material, coating procedure, cell seeding density, and growth media composition. Biochemical characterization of cell type-specific transmembrane adherens junction protein VE-cadherin and of TJ proteins ZO-1 and claudin-5 were carried out for each endothelial cell line. In addition, immunostaining for ZO-1 in hBMEC cell line was performed.ResultsThe four cell lines all expressed the endothelial cell type-specific adherens junction protein VE-cadherin. The TJ protein ZO-1 was expressed in hCMEC/D3 and in hBMEC cells. ZO-1 expression could be confirmed in hBMEC cells by immunocytochemical staining. Claudin-5 expression was detected in hCMEC/D3, TY10, and at a very low level in hBMEC cells. Highest TEER values and lowest paracellular permeability for Na-F and LY were obtained with mono-cultures of hBMEC cell line when cultivated on 24-well tissue culture inserts from Greiner Bio-one® (transparent PET membrane, 3.0 μm pore size). In co-culture models with SVG-A and HBPCT cells, no increase of TEER could be observed, suggesting that none of the investigated endothelial cell lines responded positively to stimuli from immortalized astrocytic or pericytic cells.ConclusionsUnder the conditions examined in our experiments, hBMEC proved to be the most suitable human cell line for an in vitro BBB model concerning barrier tightness in a 24-well mono-culture system intended for higher throughput. This BBB model is being validated with several compounds (known to cross or not to cross the BBB), and will potentially be selected for the assessment of BBB permeation of bioactive natural products.

Formulation approaches to improving the delivery of an antiviral drug with activity against seasonal flu

The main objective of the present study was to develop formulations of noscapine hydrochloride hydrate with enhanced solubility and bioavailability using co-solvent- and cyclodextrin-based approaches. Different combinations of co-solvents, which were selected on the basis of high-throughput solubility screening, were subjected to in vitro intestinal drug permeability studies conducted with Ussing chambers. Vitamin E tocopherol polyethylene glycol succinate and propylene glycol based co-solvent formulations provided the maximum permeability coefficient for the drug. Inclusion complexes of the drug were prepared using hydroxypropyl-β-cyclodextrin and sulphobutylether cyclodextrins. Pharmacokinetic studies were carried out in male Sprague–Dawley rats for the selected formulations. The relative bioavailabilities of the drug with the co-solvent- and cyclodextrin-based formulations were found to be similar.

Development, characterization & in vitro skin permeation of rutin ethosomes as a novel vesicular carrier

Objective : The aim of the present study was to formulate and characterize rutin ethosomes and to evaluate the rutin ethosomes to enhance its skin permeation rate as compared to parent drug, rutin. Materials and Methods : Rutin ethosomes were formulated by cold method and optimized using 2 3 factorial design by design expert software (version 8.0). The optimized rutin ethosomes were characterized for particle size distribution, vesicular shape using scanning electron microscopy, zeta potential, entrapment efficiency and short term stability studies. A specially designed franz diffusion cell was used for the percutaneous absorption studies of optimized rutin ethosomes using pig ear skin and the results were analysed using DD solver 8 software. Results and Discussion : The entrapment efficiency of optimized ethosomes prepared using phospholipid concentration of 2.5 g with 30 ml ethanol for 12.5 minutes, was found to be maximum i.e. 73.9 %. Lipid vesicular system of rutin exhibited a negative zeta potential of -46.0 mV indicating a high degree of stability for rutin ethosomes. In vitro drug permeation studies of pure rutin and rutin ethosomes suggested that rutin ethosomes were better able to permeate across pig ear skin at the end of 120 minutes than pure rutin. Therefore, rutin ethosomes were better able to facilitate permeation of rutin deeply into the pig ear skin when compared with pure rutin. Conclusion : Rutin ethosomal formulation has tremendous potential to serve as a topical drug delivery system due to its enhanced rate of permeation.

Nuts ’n’ guts: transport of food allergens across the intestinal epithelium

The increase in the incidence of food allergy is a growing problem for the western world. This review will focus on the findings from several macromolecular epithelial transport experiments and drug permeability studies to provide a recent comprehension of food allergen intestinal epithelial cell transport and the allergen-epithelial relationship. Specifically, this review will aim to answer whether allergens can permeate the intestinal barrier directly via intestinal epithelial cells, and whether this mode of transport affects downstream immune reactions. By improving our understanding of the interactions which take place during exposure of food allergens with the intestinal epithelium, we can begin to understand whether the epithelial barrier plays a major role in the allergic sensitization process rather than simply restricting the entry of allergens to the underlying lamina propria.

Development of solid dispersions of artemisinin for transdermal delivery

Solid dispersions of the poorly soluble drug artemisinin were developed using polymer blends of polyvinylpyrrolidone (PVP) and polyethylene glycol (PEG) with the aim of enhancing solubility and in vitro permeation of artemisinin through skin. Formulations were characterised using a combination of molecular dynamics (MD) simulations, differential scanning calorimetry (DSC), X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FT-IR). Solubility of artemisinin was determined in two solvents: de-ionised water and phosphate buffered saline (PBS; pH 7.4), while in vitro drug permeation studies were carried out using rabbit skin as a model membrane. MD simulations revealed miscibility between the drug and polymers. DSC confirmed the molecular dispersion of the drug in the polymer blend. Decrease in crystallinity of artemisinin with respect to polymer content and the absence of specific drug-polymer interactions were confirmed using XRD and FT-IR, respectively. The solubility of artemisinin was dramatically enhanced for the solid dispersions, as was the permeation of artemisinin from saturated solid-dispersion vehicles relative to that from saturated solutions of the pure drug. The study suggests that high energy solid forms of artemisinin could possibly enable transdermal delivery of artemisinin.

NOSE TO BRAIN DELIVERY OF ZIPRASIDONE MICROEMULSION: DESIGEN AND CHARACTERIZATION

Nasal administration offers an interesting promising alternative technique to the parenteral route for achieving systemic drug effects. Now a day many drugs have better systemic bioavailability through nasal route as compared to oral administration. Microemulsion is dispersed, macroscopically homogenous thermodynamically stable, optically transparent, single phase system, formed by spontaneous solubilisation of two immiscible liquid, in the presence of surfactants. Mic roemulsion offers number of advantages than drug formu lation for nasal drug delivery. Ziprasidone is lipophilic, benz - isothiazoyl - piperazine derivative, used in the treatment of schizophrenia, mania and mixed states associated with bipolar disorder. The mi croemulsion was prepared by precipitation method. The characterization of microemulsion formulation includes determination of viscosity, pH, drug content, refractive, conductivity measurement, zeta potential analysis and particle size analysis . In - vitro dr ug permeation studies were done b y using Keshary Diffusion cell. All ME formulations were shown fast drug permeat ion 99.50 % for ME 2 within 1 h and 99.40 % for ME 6 within 100 minutes since the ME 2 which is optimized on the basis of in - vitro study. Nasal mucoadhesive drug delivery system is designed with an aim to target the drug and to maintain the dosage form at its absorptio n site for a rapid onset of time. This will result in the enhancement of the absorption of the drug, which will in turn reduce the presystemic metabolism; increase the bioavailability of the drug, initiate rapid onset of action and thus will decrease the dosing frequency and dose r elated side effects of the drug . For this purpose in microemulsion system is preferred over conventional solution system.

Re-evaluation of the role of P-glycoprotein in in vitro drug permeability studies with the bovine brain microvessel endothelial cells

1. Currently available in vitro blood-brain barrier models all have recognized restrictions. In addition to leakiness, inconsistent data about P-glycoprotein mediated efflux limit the attractiveness of the primary bovine brain microvessel endothelial cells (BBMECs). Therefore, we re-evaluated the role of P-glycoprotein mediated efflux with two culture conditions in BBMECs for prediction of drug permeability of potential P-glycoprotein substrates.2. BBMECs were monocultured on filters on petri dishes and on filter inserts, and expression and localization of P-glycoprotein were compared by using western blot and confocal microscopy, respectively. The functionality of P-glycoprotein was assessed by using cellular uptake, calcein-AM and bidirectional transport assays.3. P-glycoprotein expression was higher in BBMECs cultured on filter inserts decreasing the permeability of digoxin and paclitaxel, but not the permeability of vinblastine. However, the monocultured BBMECs were not able to demonstrate efflux in the bidirectional transport assays. Under certain culture conditions, occludin may not be correctly located, perhaps explaining in part the leakiness of BBMECs.4. In conclusion, BBMECs, despite possessing a functional P-glycoprotein, under certain culture conditions may not be a suitable in vitro model for the bidirectional transport assays and for predicting the permeability of drugs and xenobiotics that are potential P-glycoprotein substrates.

Surfactant-rich biocompatible microemulsions as effective carriers of methylxanthine drugs

Under the perspective of their potential pharmaceutical applications, new surfactant-rich biocompatible microemulsions were prepared and structurally characterized to be used as novel carriers of theophylline and theobromine. The existence of single phase regions was investigated in pseudo-ternary phase diagrams of three different microemulsion systems: (1) isopropyl palmitate/Triton X-100/water+propylene glycol, (2) Miglyol 818/Triton X-100/water+propylene glycol and (3) isopropyl palmitate/Triton X-100+Span 20/water+propylene glycol. Electrical conductivity measurements indicated the formation of water-in-oil (w/o) structures. Interfacial properties of the microemulsions were studied by electron paramagnetic resonance (EPR) spectroscopy employing the nitroxide spin probe 5-doxylstearic acid (5-DSA). The determined order parameter and wobbling angle showed that the system composition influences the membrane flexibility whereas drug incorporation resulted in more flexible membranes. Particle size measurements were performed using dynamic light scattering (DLS) showing that addition of the drugs resulted in the formulation of larger aqueous droplets. To evaluate the ability of the proposed microemulsions to serve as carriers of bioactive compounds for topical administration, in vitro permeation studies were carried out using the Franz type diffusion cells and a model membrane. The nature of the oil and the surfactant used for the construction of the microemulsions affected the phase behavior, the size and also the interfacial properties of both free and loaded systems. Drug permeation studies revealed the effectiveness of the proposed formulations as carriers of theophylline and theobromine.

Antioxidant activity, lipophilicity and extractability of polyphenols from pig skin – development of analytical methods for skin permeation studies

Permeation of polyphenols through the stratum corneum barrier is a precondition for the protective action of polyphenols against oxidative skin damage. Prior to in vitro skin permeation experiments, we developed a method for the quantification of polyphenols in pig skin, including organic solvent extraction and HPLC analysis. Catechine hydrate, epigallocatechin gallate, trans-resveratrol, quercetin, rutin and protocatechuic acid were chosen for this study as representatives of phenolics with different lipophilicity and molecular weight. The antioxidative activities of polyphenols as well as their octanol-water partition coefficients at different pH values were determined. Extraction of polyphenols from pig skin was optimized by variation of solvent composition, homogenization intensity and time, as well as partial exclusion of oxygen during extraction. The highest recovery rates could be reached by extraction with the methanol-water mixture (90:10, v/v), containing 0.2 g/L l-ascorbic acid, after the cryo-milling for 4 min. Recoveries of 72% for total phenolics, 96% for quercetin and protocatechuic acid, 90% for rutin and 74% for trans-resveratrol, were achieved. These extraction parameters will be selected for the polyphenol extraction from pig skin for further in vitro drug permeation studies.

Dose-Dependent Targeted Suppression of P-glycoprotein Expression and Function in Caco-2 Cells

The efflux transporter P-glycoprotein (Pgp), encoded by the ABCB1 gene, decreases the bioavailability of a wide range of orally administered drugs. Drug permeability studies using the in vitro Caco-2 cell model commonly rely on small molecule modulators to estimate the contribution of Pgp to drug efflux. The use of such modulators may be limited by their interactions with other membrane transporters. RNA interference, a tool allowing for the specific degradation of a target gene’s mRNA, has emerged as a technique to study gene expression and function. This manuscript describes the use of chemically modified small interfering RNA (siRNA) for a dose-dependent suppression of ABCB1 in Caco-2 cells and the subsequent drug permeability assay. We transfected Caco-2 cells while in suspension with chemically modified synthetic siRNA–lipid complexes and then seeded the cells on polycarbonate semipermeable supports. Once the monolayer of Caco-2 cells formed tight junctions and expressed brush border enzymes, we determined the dose-dependent suppression of the ABCB1 gene using RT-qPCR. We measured the duration of silencing at the optimal siRNA dose by Western blot for Pgp protein. The utility of this in vitro model was determined by performing bidirectional transport studies using a well-established substrate for Pgp, rhodamine 123. A single 4 h transfection of the Caco-2 cells with ≥100 nM siRNA reduced the expression of ABCB1 mRNA by >85% at day five in culture. The time-course study revealed that the single transfection reduces Pgp protein levels for 9 days in culture. This magnitude of silencing was sufficient to reduce the efflux of rhodamine 123 as measured by the apparent permeability coefficient and intracellular accumulation. In this study, we demonstrate the dose-dependent, targeted degradation of Pgp in Caco-2 cells as a new model for assessing drug efflux from enterocytes. The dose-dependent nature of the Pgp silencing in this study offers significant improvements over other approaches to creating a Caco-2 model with suppressed ABCB1 expression. We envision that this technique, in conjunction with better small molecule inhibitors, will provide a useful tool for future drug permeability studies.

Determination of lamivudine and zidovudine permeability using a different ex vivo method in Franz cells

IntroductionThe major processes that control the absorption of orally administered drugs are dissolution and gastrointestinal permeation. These processes depend on two main properties: solubility and permeability. Based on these characteristics, the Biopharmaceutical Classification System (BCS) was proposed as a tool to assist in biowaiver and bioavailability prediction of drugs. MethodsThe purpose of the present study was to evaluate the permeability of lamivudine (3TC) and zidovudine (AZT) using a different ex vivo method in Franz cells. A segment of jejunum was inserted in a Franz cells apparatus, in order to assess drug permeability in the apical–basolateral (A–B) and basolateral–apical (B–A) directions. Each drug was added to the donor chamber, collected from the acceptor chamber and analyzed by HPLC. Fluorescein (FLU) and metoprolol (METO) were used as low and high permeability markers, respectively. ResultsThe apparent permeability (Papp) results for the A–B direction were: Papp FLU A–B=0.54×10−4cm·s−1, Papp METO A–B=7.99×10−4cm·s−1, Papp 3TC A–B=4.58×10−4cm·s−1 and Papp AZT A–B=5.34×10−4cm·s−1. For the B–A direction, the Papp results were: Papp FLU B–A=0.56×10−4cm·s−1, Papp METO B–A=0.25×10−4cm·s−1, Papp 3TC B–A=0.24×10−4cm·s−1 and Papp AZT B–A=0.19×10−4cm·s−1. DiscussionFor the A–B direction, the Papp results of fluorescein and metoprolol show low and high permeability, respectively, indicating that the membranes were appropriate for permeability studies. For the A–B direction, the Papp results of 3TC and AZT suggest that these antiretroviral drugs have permeability values close to metoprolol. Nevertheless, for the B–A direction the Papp results do not suggest efflux mechanism for any of the drugs. Thereby, the different ex vivo methods using Franz cells can be successfully applied in drug permeability studies, in particular for drug biopharmaceutical classification.

Novel surface modified solid lipid nanoparticles as intranasal carriers for ropinirole hydrochloride: application of factorial design approach

Present investigation deals with intranasal delivery of ropinirole hydrochloride (ROPI HCl), loaded in solid lipid nanoparticles (SLNs). Prime objectives of this experiment are avoidance of hepatic first pass metabolism and to improve therapeutic efficacy in the treatment of Parkinson’s disease. SLNs were fabricated by emulsification-solvent diffusion technique. A 32-factorial design approach has been employed to assess the influence of two independent variables, namely Pluronic F-68 and stearylamine concentration on particle size, ζ-potential and entrapment efficiency of prepared SLNs. Prepared samples were further evaluated for in vitro drug diffusion, ex vivo drug permeation, histopathological and stability studies. Differential scanning calorimetry analysis revealed the encapsulation of amorphous form of drug into lipid matrix, while scanning electron microscopy studies indicated the spherical shape. Fabricated SLNs had shown no severe signs of damage on integrity of nasal mucosa. Release pattern of prepared drug-loaded sample was best fitted to zero-order kinetic model with non-Fickian super case II diffusion mechanism. In vivo pharmacodynamic studies were carried out to compare therapeutic efficacy of prepared nasal formulation against marketed oral formulation. Results of analysis of variance demonstrated the significance of suggested model. Three-dimensional response surface plots and regression equations confirmed the corresponding influence of selected independent variables on measured responses. Our findings suggested the feasibility of investigated system for intranasal administration.

Design, development and permeation studies of nebivolol hydrochloride from novel matrix type transdermal patches

Background:Nebivolol hydrochloride is a third generation β-blocker with highly selective β1-receptor antagonist with antihypertensive properties having plasma half life of 10 h and 12% oral bioavailability. The aim of the present investigation was to form matrix type transdermal patches containing Nebivolol hydrochloride to avoid its extensive hepatic first pass metabolism, lesser side effect and increase bioavailability of drug.Materials and Methods:Matrix type transdermal patches containing Nebivolol hydrochloride were prepared using EudragitRS100, HPMC K100M (2:8) polymers by solvent evaporation technique. Aluminum foil was used as a backing membrane. Polyethylene glycol (PEG) 400 was used as plasticizer and Dimethyl sulfoxide (DMSO) was used as a penetration enhancer. Drug polymer interactions determined by FTIR and standard calibration curve of Nebivolol hydrochloride were determined by using UV estimation.Result:The systems were evaluated physicochemical parameters and drug present in the patches was determined by scanning electron microscopy. All prepared formulations indicated good physical stability. In vitro drug permeation studies of formulations were performed by using Franz diffusion cells using abdomen skin of Wistar albino rat. Result showed best in vitro skin permeation through rat skin as compared to all other formulations prepared with hydrophilic polymer containing permeation enhancer.Conclusions:It was observed that the formulation containing HPMC: EudragitRS100 (8:2) showed ideal higuchi release kinetics. On the basis of in vitro drug release through skin permeation performance, Formulation F1 was found to be better than other formulations and it was selected as the optimized formulation.

Development of O/W nanoemulsions for ophthalmic administration of timolol

After an initial screening of ingredients and production methods, nanoemulsions for ocular administration of timolol containing the drug as maleate (TM) or as ion-pair with AOT (TM/AOT) were prepared. The physico-chemical characterization of nanoemulsions, regarding mean diameter, pH, zeta potential, osmolarity, viscosity and surface tension, underlined their feasibility to be instilled into the eyes. Single components and emulsions were tested ex vivo on rabbit corneas to evaluate corneal irritation, that was measured according to opacity test. A marked decrease in corneal opacity was observed using the drug formulated in nanoemulsions rather than in aqueous solutions. Drug permeation and accumulation studies were performed on excised rabbit corneas. An increase in drug permeation through and accumulation into the corneas were observed using TM-AOT compared to TM due to an increase of lipophilicity of the drug as ion-pair. The introduction of chitosan (a positive charged mucoadhesive polymer) into emulsions allowed to increase TM permeation probably due to the interaction of chitosan with corneal epithelial cells.

Establishment of a simplified in vitro porcine blood–brain barrier model with high transendothelial electrical resistance

Good in vitro blood–brain barrier (BBB) models that mimic the in vivo BBB phenotype are essential for studies on BBB functionality and for initial screening in drug discovery programmes, as many potential therapeutic drug candidates have poor BBB permeation. Difficulties associated with the availability of human brain tissue, coupled with the time and cost associated with using animals for this kind of research have led to the development of non-human cell culture models. However, most BBB models display a low transendothelial electrical resistance (TEER), which is a measure of the tightness of the BBB. To address these issues we have established and optimised a robust, simple to use in vitro BBB model using porcine brain endothelial cells (PBECs). The PBEC model gives high TEER without the need for co-culture with astrocytes (up to 1300Ωcm2 with a mean TEER of ∼800Ωcm2) with well organised tight junctions as shown by immunostaining for occludin and claudin-5. Functional assays confirmed the presence of high levels of alkaline phosphatase (ALP), and presence of the efflux transporter, P-glycoprotein (P-gp, ABCB1). Presence of the breast cancer resistance protein (BCRP, ABCG2) was confirmed by TaqMan real-time RT-PCR assay. Real-time RT-PCR assays for BCRP, occludin and claudin-5 demonstrated no significant differences between batches of PBECs, and also between primary and passage 1 PBECs. A permeability screen of 10 compounds demonstrated the usefulness of the model as a tool for drug permeability studies. Qualitative and quantitative results from this study confirm that this in vitro porcine BBB model is reliable and robust; it is also simpler to generate than most other BBB models.This article is part of a Special Issue entitled Electrical Synapses.