Nano-carrier systems: Strategies to overcome the mucus gel barrier.

The present review provides an overview of nanotechnology-based strategies to overcome various mucus gel barriers including the intestinal, nasal, ocular, vaginal, buccal and pulmonary mucus layer without destroying them. It focuses on the one hand on strategies to improve the mucus permeation behavior of particles and on the other hand on systems avoiding the back-diffusion of particles out of the mucus gel layer. Nanocarriers with improved mucus permeation behavior either exhibit a high density of positive and negative charges, bearing mucolytic enzymes such as papain and bromelain on their surface or display a slippery surface due to PEG-ylation. Furthermore, self-nanoemulsifying-drug-delivery-systems (SNEDDS) turned out to exhibit comparatively high mucus permeating properties. Strategies in order to avoid back-diffusion are based on thiolated polymers reacting to a higher extent with cysteine subunits of the mucus at pH 7 in deeper mucus regions than at pH 5 being prevalent in luminal mucus regions of the intestinal and vaginal mucosa. Furthermore, particles changing their zeta potential from negative to positive once they have reached the epithelium seem to be promising carriers. The summarized knowledge should provide a good starting point for further developments in this field.

Mucus permeating thiomer nanoparticles.

The aim of this study was to develop and evaluate a novel mucoadhesive drug delivery system based on thiolated poly(acrylic acid) nanoparticles exhibiting mucolytic properties to enhance particle diffusion into deeper mucus regions before adhesion. Mediated by a carbodiimide, cysteine and the mucolytic enzyme papain were covalently attached to poly(acrylic acid) via amide bond formation. The conjugates were co-precipitated with calcium chloride in order to obtain papain modified (PAA-pap) and thiolated nanoparticles (PAA-cys) as well as particles containing both conjugates (PAA-cys-pap). The nanoparticulate systems were characterized regarding particle size distribution and zeta potential. Particle transport was investigated by diffusion studies across intestinal mucus using two different techniques. Furthermore, mucoadhesive properties of all particles were evaluated via rheological measurements. Results demonstrated that all nanoparticles were in a size range of 158-214 nm and showed negative zeta potentials. Due to the presence of papain, the PAA-cys-pap particles were capable of cleaving mucoglycoprotein substructures and consequently exhibited a 2.0-fold higher penetration into the mucus layer in comparison with PAA-cys particles. Within the rheological studies, an 1.9-fold increase in mucoadhesion could be achieved for the nanoparticulate system based on thiolated PAA compared to papain modified particles (PAA-pap). Therefore, the newly developed particulate system (PAA-cys-pap) is characterized by mucoadhesive as well as mucolytic properties. The combination of both effects - mucus-permeating and mucoadhesive properties - might be a promising strategy for the development of oral drug delivery systems to overcome the mucus barrier and providing a prolonged residence time close to the absorption membrane.

Thiolated nanocarriers for oral delivery of hydrophilic macromolecular drugs.

It was the aim of this study to investigate the effect of unmodified as well as thiolated anionic poly(acrylic acid) (PAA) and cationic chitosan (CS) utilized in free-soluble form and as nanoparticulate system on the absorption of the hydrophilic compound FD4 across intestinal epithelial cell layer with and without a mucus layer. Modifications of these polymers were achieved by conjugation with cysteine to PAA (PAA-Cys) and thioglycolic acid to CS (CS-TGA). Particles were prepared via ionic gelation and characterized based on their amount of thiol groups, particle size and zeta potential. Effects on the cell layer concerning absorption enhancement, transepithelial electrical resistance (TEER) and cytotoxicity were investigated. Permeation enhancement was evaluated with respect to in vitro transport of FD4 across Caco-2 cells, while mucoadhesion was indirectly examined in terms of adsorption behaviour when cells were covered with a mucus layer. Lyophilized particles displayed around 1000 µmol/g of free thiol groups, particle sizes of less than 300 nm and a zeta potential of 18 mV (CS-TGA) and -14 mV (PAA-Cys). Cytotoxicity studies confirmed that all polymer samples were used at nontoxic concentrations (0.5% m/v). Permeation studies revealed that all thiolated formulations had pronounced effects on the paracellular permeability of mucus-free Caco-2 layers and enhanced the permeation of FD4 3.0- to 5.3-fold. Moreover, polymers administered as particles showed a higher permeation enhancement than their corresponding solutions. However, the absorption-enhancing effect of each thiolated formulation was significantly (p<0.05) reduced when cells were covered with mucus layer. In addition, all formulations were able to decrease the TEER of the cell layer significantly (p<0.05). Therefore, both thiolated polymers as nanoparticulate delivery systems represent a promising tool for the oral administration of hydrophilic macromolecules.

Development of a dosage form for accelerated release.

PURPOSE: It was the aim of this study to develop an oral capsule delivery system capable of rapidly ejecting the incorporated payload in the small intestine. METHODS: The capsule consists of four parts: a reaction mixture comprising of a basic and a corresponding acidic component, a plunger necessary to separate the reaction mixture from the inserted ingredients, capsule cap and body (made out of ethylcellulose (EC)), where at the bottom of the body a semipermeable filter membrane is mounted. As soon as water permeates through the membrane, the reaction mixture dissolves and carbon dioxide (CO2) is released resulting in a high speed liberation of inserted compounds onto the intestinal mucosa. Several filter membranes were investigated regarding water influx, capillary force and water retention capacity. CO2 release of sodium hydrogen carbonate (NaHCO3) was examined in presence of several acidic components in different morphological forms (powder, lyophilisate and granule) and the amount of CO2 liberation out of prepared capsules was determined. Furthermore, release of enteric coated capsules was tested within 0.1M HCl and 100mM phosphate buffer pH 6.8. RESULTS: The rank order regarding membrane permeability was determined to be: cellulose acetate with a pore diameter of 12-15 µm>4-12 µm cellulose acetate>8 µm cellulose nitrate>8-12 µm cellulose acetate. NaHCO3 in combination with tartaric acid in form of a granule could be identified as the most promising reaction mixture with the highest amount of released CO2 compared to all other reaction mixture combinations. Stability of enteric coated capsules in HCl and a spontaneous release in phosphate puffer could be demonstrated within in vitro release studies. CONCLUSION: In light of these results, the developed releasing system seems to be a promising tool for an accelerated delivery of several incorporated excipients.

Liposomes coated with thiolated chitosan enhance oral peptide delivery to rats.

The aim of the present study was the in vivo evaluation of thiomer-coated liposomes for an oral application of peptides. For this purpose, salmon calcitonin was chosen as a model drug and encapsulated within liposomes. Subsequently, the drug loaded liposomes were coated with either chitosan-thioglycolic acid (CS-TGA) or an S-protected version of the same polymer (CS-TGA-MNA), leading to an increase in the particle size of about 500 nm and an increase in the zeta potential from approximately -40 mV to a maximum value of about +44 mV, depending on the polymer. Coated liposomes were demonstrated to effectively penetrate the intestinal mucus layer where they came in close contact with the underlying epithelium. To investigate the permeation enhancing properties of the coated liposomes ex vivo, we monitored the transport of fluoresceinisothiocyanate-labeled salmon calcitonin (FITC-sCT) through rat small intestine. Liposomes coated with CS-TGA-MNA showed the highest effect, leading to a 3.8-fold increase in the uptake of FITC-sCT versus the buffer control. In vivo evaluation of the different formulations was carried out by the oral application of 40 µg of sCT per rat, either encapsulated within uncoated liposomes, CS-TGA-coated liposomes or CS-TGA-MNA-coated liposomes, or given as a solution serving as negative control. The blood calcium level was monitored over a time period of 24h. The highest reduction in the blood calcium level, to a minimum of 65% of the initial value after 6h, was achieved for CS-TGA-MNA-coated liposomes. Comparing the areas above curves (AAC) of the blood calcium levels, CS-TGA-MNA-coated liposomes led to an 8.2-fold increase compared to the free sCT solution if applied orally in the same concentration. According to these results, liposomes coated with S-protected thiomers have demonstrated to be highly valuable carriers for enhancing the oral bioavailability of salmon calcitonin.

Thiomer-coated liposomes harbor permeation enhancing and efflux pump inhibitory properties.

An ideal oral drug carrier should facilitate drug delivery to the gastrointestinal tract and its absorption into the systemic circulation. To meet these requirements, we developed a thiomer-coated liposomal delivery system composed of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and a maleimide-functionalized lipid, to which chitosan-thioglycolic acid (CS-TGA) was covalently coupled. In addition to conventional 77 kDa CS-TGA (CS-TGA77), we tested the 150 kDa homologue (CS-TGA150) as well as an S-protected version of this polymer (CS-TGA150-MNA), in which some of the free SH-groups are conjugated with 6-mercaptonicotinamide to protect them from oxidation. Coupling of CS-TGA to the liposomal surface led to an increase in the particle size of at least 150 nm and an increase in the zeta potential from approximately -33 mV to a maximum of about +36 mV, depending on the polymer. As revealed by fluorescence dequenching the formulations have a storage stability of at least two weeks without releasing any encapsulated compounds. In simulated gastric fluid, the system was shown to be stable over 24 h, while in simulated intestinal fluid, a slow, sustained release of encapsulated compounds was observed. According to our experiments, thiomer-coated liposomes did not induce immunogenic reactions after an oral administration to mice. To evaluate the permeation enhancing and efflux pump inhibiting properties of CS-TGA coated liposomes we monitored the transport of fluoresceinisothiocyanate-dextran (FD(4)) and rhodamine-123 (Rho-123), respectively, through rat small intestine. Permeation studies showed a 2.8-fold higher permeation of FD(4) in the presence of CS-TGA77 coated liposomes and an even 4-fold higher permeation in the presence of CSA-TGA150-MNA coated liposomes. The latter also performed best when we evaluated P-glycoprotein inhibiting properties by monitoring the transport of Rho-123, revealing a 4.2-fold enhancement respective to the buffer control. Taken together, thiomer-coated liposomes were shown to protect encapsulated drugs in the stomach, slowly release them in the small intestine and enhance their absorption through the intestinal tissue by opening tight junctions and inhibiting efflux pumps.