Polyelectrolyte-drug complexes of lambda carrageenan and basic drugs: relevance of particle size and moisture content on compaction and drug release behavior.

The interaction between polyelectrolytes (PE) and oppositely charged drugs (D) results in complexes (PE-D) that can be exploited in controlled release drug delivery systems. The aim of this work is to better understand the relevance of some preparative parameters such as moisture content and particle size on the performance of two PE-D complexes to be used in oral controlled release tablets. PE-D complexes containing diltiazem HCL (DTZ) or metoprolol tartrate (MTP) and lambda carrageenan were obtained at two particle size levels (<45 microm and 75-105 microm), maintained at different values of relative humidity (RH) (11, 52, 75, and 93%), and compressed. The tablets were characterized for porosity, hardness, moisture content, and contact angle. Drug release profiles were fitted to the Weibull equation, and a factorial design was used to understand the relevance of particle size and RH% on release rate as a function of medium pH. The results indicated that the hydrophobic character of the complex between PE and D depended on the drug and in the present case was more pronounced for DTZ than for MTP. This in turn affected the possible release mechanism and therefore the importance of particle size and RH%.

A novel dressing for the combined delivery of platelet lysate and vancomycin hydrochloride to chronic skin ulcers: Hyaluronic acid particles in alginate matrices.

The aim of the present work was to develop a medication allowing for the combined delivery of platelet lysate (PL) and an anti-infective model drug, vancomycin hydrochloride (VCM), to chronic skin ulcers. A simple method was set up for the preparation of hyaluronic acid (HA) core-shell particles, loaded with PL and coated with calcium alginate, embedded in a VCM containing alginate matrix. Two different CaCl2 concentrations were investigated to allow for HA/PL core-shell particle formation. The resulting dressings were characterized for mechanical and hydration properties and tested in vitro (on fibroblasts) and ex-vivo (on skin biopsies) for biological activity. They were found of sufficient mechanical strength to withstand packaging and handling stress and able to absorb a high amount of wound exudate and to form a protective gel on the lesion area. The CaCl2 concentration used for shell formation did not affect VCM release from the alginate matrix, but strongly modified the release of PGFAB (chosen as representative of growth factors present in PL) from HA particles. In vitro and ex vivo tests provided sufficient proof of concept of the ability of dressings to improve skin ulcers healing.

Alpha tocopherol loaded chitosan oleate nanoemulsions for wound healing. Evaluation on cell lines and ex vivo human biopsies, and stabilization in spray dried Trojan microparticles.

An amphiphilic chitosan salt, chitosan oleate (CS-OA), was previously proposed for the physical stabilization of lemongrass antimicrobial nanoemulsions (NE) through a mild spontaneous emulsification process. As both chitosan and oleic acid are described in the literature for their positive effects in wound healing, in the present study CS-OA has been proposed to encapsulate alpha tocopherol (αTph) in NEs aimed to skin wounds. A NE formulation was developed showing about 220 nm dimensions, 36% drug loading, and αTph concentration up to 1 mg/ml. Both CS-OA and αTph NE stimulated cell proliferation on keratinocytes and fibroblast cell cultures, and in ex vivo skin biopsies, suggesting the suitability of CS-OA and of the antioxidant agent for topical application in wound healing. αTph stability was further improved with respect of encapsulation, by spray drying the NE into a powder (up to about 90% αTph residual after 3 months). The spray drying process was optimized, to improve powder yield and αTph recovery, by a design of experiments approach. The powder obtained was easily re-suspended to deliver the NE and resulted able to completely release αTph.

Carvacrol/clay hybrids loaded into in situ gelling films.

The aim of the present work was the development of polymer films loaded with a carvacrol (CVR)/clay hybrid (HYBD) for the delivery of CRV in infected skin ulcer treatment. Different clays were considered: montmorrilonite, halloysite and palygorskite (PHC). CRV incorporation in PHC reduced its volatility. HYBD showed 20% w/w CRV loading capacity and was able to preserve CRV antioxidant properties. HYBD was characterized by improved antimicrobial properties against S. aureus and E. coli and cytocompatibility towards human fibroblasts with respect to pure CRV. Films were prepared by casting an aqueous dispersion containing poly(vinylalcohol) (PVA), poly(vinylpyrrolidone) (PVP), chitosan glutamate (CS), sericin and HYBD. Optimization of film composition was supported by a Design of Experiments (DoE) approach. In a screening phase, a full factorial design (FFD) was used and the following factors were investigated at two levels: PVA (12-14%w/w), PVP (2-4%w/w) and CS (0.134-0.5%w/w) concentrations. For the optimization phase, FFD was expanded to a "central composite design". The response variables considered were: elongation, tensile strength and buffer absorption of films, durability of the gels formed after film hydration. Upon hydration, the optimized film formed a viscoelastic gel able to protect the lesion area and to modulate CRV release.

Histological evaluation of buccal penetration enhancement properties of chitosan and trimethyl chitosan.

The aim of the present work was to compare the penetration enhancement properties of chitosan hydrochloride (HCS) both as a polymeric solution and as a nanoparticulate system with that of trimethyl chitosan hydrochloride (TMC) on buccal mucosa. The hydrophilic high molecular weight fluorescein isothiocyanate dextran (FD4; 4400 Da) was used as a macromolecule model. The mechanism involved in the HCS (solution and nanoparticles) and TMC solution penetration enhancement was investigated on pig buccal mucosa, characterized by having stratified epithelium and lacking in tight junctions. The permeation/penetration of FD4 and the change in morphology and histology of the mucosa after contact with the polymers were assessed: the experiments were performed ex-vivo by applying the formulations on excised porcine buccal tissue. For the morphology and the histology studies, the epithelial cell layers from freshly excised pig buccal mucosa were analysed with light microscopy by means of routine histopathology analysis (haematoxylin and eosin staining and Toluidine blue staining) and immunohistochemistry reactions. The organization of desmosomal junctions was assessed by means of an immunochemical reaction on desmosomes and transmission electron microscopy. Confocal laser scanning microscopy (CLSM) was used to find evidence of the location of FD4 in the tissue. Furthermore, the increase of the FD4 apparent permeability coefficient was quantified by means of Franz diffusion cells using isolated buccal epithelium to demonstrate the penetration enhancement properties of the polymer systems. Morphological analysis, performed by light microscopy, transmission electron microscopy and CLSM, suggests a similar mechanism of penetration enhancement for both HCS and TMC solutions and for HCS nanoparticles. Such a mechanism probably involves a repackaging of the epithelial cells up to the basal membrane and a partial disarrangement of desmosomes. The cell viability and the nuclear integrity indicated on the semi-thin section stained with Toluidine blue and by CLSM analysis, respectively, suggest that HCS as a polymer solution and a nanoparticulate system, and TMC polymer solution, do not cause cell damage. Trimethyl chitosan and chitosan nanoparticulate systems were able to increase FD4 permeation across buccal epithelium to a greater extent than the chitosan solution.

Palmitoyl Glycol Chitosan Micelles for Corneal Delivery of Cyclosporine.

Different substitution degrees of palmitoyl glycol chitosan (PGC), prepared according to the literature, were used to obtain polymeric micelles that have been assessed in comparison with Pluronic F127 micelles as possible carriers for poorly soluble drugs, such as cyclosporine A. Both PGC and Pluronic micelles were studied for their interactions with cell culture substrates. The least substituted and most hydrophilic derivative, PGC21 (approximately 5% substitution), showed a strong association with cyclosporine, more than tripling the colloidal concentration with respect to the saturated solution. It showed a greater ability to open Caco-2 tight junctions and to enhance the permeability of Caco-2 substrates with respect to micelles based on higher palmitoyl substitution, conceivably due to the lower modification of the chitosan chains. Permeation and penetration experiments were performed with PGC21 and Pluronic micelles on a rabbit corneal epithelial cell line (RCE) and on excised pig corneas. It was found that both PGC and Pluronic micelles could increase the permeation of the fluorescent probe rhodamine B through RCE cells by more than ten-fold. In RCE and in pig cornea, the micelles improved the penetration of both rhodamine and cyclosporine. For cyclosporine, the PGC21 micelles allowed penetration of approximately 1 µg/mg cyclosporine A in corneal tissue, demonstrating a potential for use in immunosuppression therapies.

Opportunities offered by chitosan-based nanotechnology in mucosal/skin drug delivery.

Chitosan (CS) based nanocarriers have been extensively studied starting from early 90s. The unique properties of CS and in particular its capability to interact with various epithelia and its mucoadhesion potential have attracted many researchers. The mild preparation conditions of CS nanosystems offer the opportunities to load stress sensitive hydrophilic macromolecules such as proteins and genetic materials. Moreover CS nanosystems are able to protect their cargo from the environment (pH, enzymes). The safety issues related to this polymer seem in part overcome at least for CS as polymeric solution. For this reason skin and mucosae, in particular buccal and vaginal ones, seem the more promising administration routes with lower technical and regulatory challenges. Even if several papers focused on CS nanocarriers for skin and mucosal (buccal and vaginal) administration have been published, other work should be done aiming at optimizing CS nanocarriers in view of clinical applications.

Influence of mucin type on polymer-mucin rheological interactions.

There are numerous in vitro methods with which to investigate the mucoadhesive properties of polymers. One recent method is based on the measurement of rheological interactions between polymer and mucin, which implies the use of mucins isolated from the mucous tissue. The extraction and purification of glycoprotein fraction, which is responsible for rheological interaction, can modify the native structure of mucin or spoil it with exogenous substances. Therefore the particulars of the mucin employed (origin, purification grade, the effect of further treatments such as freezing or freeze-drying) are likely to be critical for the interaction. The aim of this work was to compare some commercial mucins of differing origin and grade of purification for their rheological interaction with well-known mucoadhesive polymers (polyacrylic acid and sodium carboxymethylcellulose). For polyacrylic acid, which is sensitive to ions, we found rheological interaction to be strongly influenced by mucin type. The removal of ions, with dialysis, improved the interaction. For sodium carboxymethylcellulose, which is less sensitive to ions, rheological interaction proved to be less dependent on mucin type and improved upon glycoprotein solubilization.

On the employment of lambda carrageenan in a matrix system. III. Optimization of a lambda carrageenan-HPMC hydrophilic matrix.

The lambda carrageenan/HPMC ratio in matrix tablets has been optimized in order to obtain pH-independent release profiles of chlorpheniramine maleate, a freely soluble drug. Release profiles in acidic (pH1.2) and neutral (pH 6.8) media were fitted according to the Weibull and the power law models. Model independent parameters(t50% and the percentage of drug released after 2 h) were also calculated. The Weibull parameters were found suitable to describe the dependence of the release profiles on matrix composition. Preparation and testing of the optimized formulation showed linear and pH-independent release profiles lasting about 24 h, in good accordance with the values predicted by the optimization procedure.

Characterization of chitosan hydrochloride--mucin rheological interaction: influence of polymer concentration and polymer:mucin weight ratio.

The aim of the present work was to investigate the influence of polymer concentration and polymer:mucin weight ratio on chitosan--mucin interaction, assessed by means of viscosimetric measurements. Two hydration media, distilled water and 0.1 M HCl, were used. Chitosan solutions were prepared at concentrations greater than the characteristic entanglement concentration and mixed with increasing amounts of porcine gastric mucin. Viscosity measurements were performed on the polymer--mucin mixtures and on polymer and mucin solutions having the same concentrations as in the mixtures. The flow curves were fitted according to a modified form of Cheng--Evans equation in order to obtain the relevant model parameters: low shear viscosity eta(0) and high shear viscosity eta(infinity), indexes of the sample structure at rest and upon high shear, respectively. The formation of chitosan--mucin interaction products was determined on the basis of the changes in eta(0) and eta(infinity) of the mixtures as a function of polymer:mucin weight ratio. Rheological synergism parameter was also calculated. The results obtained suggest that two different types of rheological interaction occur between chitosan and mucin in both media, depending on polymer concentration and polymer:mucin weight ratio: one is characterized by a minimum in viscosity and occurs at higher polymer:mucin weight ratio, the other one produces a positive rheological synergism and is observed in presence of an excess of mucin. Only the last one causes a 'strengthening' of the mucoadhesive interface and it is responsible for the mucoadhesive joint. This hypothesis is confirmed by tensile stress measurements performed on HCS solutions in presence of mucin dispersions at different concentrations.

Characterisation of particle properties and compaction behaviour of hydroxypropyl methylcellulose with different degrees of methoxy/hydroxypropyl substitution.

The particle characteristics and compaction behaviour of hydroxypropyl methylcellulose (HPMC) powders from two different suppliers were studied regarding effects of methoxy/hydroxypropyl substitution. Samples included Methocel K4M (low substitution ratio), E4M (medium) and F4M (high) and the corresponding substitution ratios from Metolose: 90 SH 4000, 60 SH 4000, and 65 SH 4000. Characterisation of the particle properties and compaction behaviour of the pure polymers suggested that reported differences in drug release behaviour of Methocel E4M compared with the other two powders may be related to the lower powder surface area, differing particle morphology and lower fragmentation propensity during compaction. In addition, compacts of Methocel E4M were weaker when tested in both axial and radial directions and had different porosity and elastic recovery properties. There were no differences between the polymers in degree of disorder, as evaluated by solid-state nuclear magnetic resonance spectroscopy. The different behaviour of Methocel E4M could, however, be related to the overall higher total degree of substitution of this polymer and in particular the high content of methoxy groups compared to the other polymers. The methoxy substituent is hydrophobic and may, when present in sufficiently high concentrations, change the particulate and mechanical properties of the powder, thus potentially affecting the compactability. The high content of methoxy groups might also decrease the development of inter- and intraparticulate hydrogen bonds during compaction, and suppress the actions of the hydrophilic hydroxypropyl groups, both of which could affect drug release.

Characterization of chitosan hydrochloride-mucin interaction by means of viscosimetric and turbidimetric measurements.

In the present work the interaction between chitosan hydrochloride (HCS) and two different types of mucin - one obtained from bovine submaxillary glands and the other from porcine stomach - was investigated. Two hydration media were tested: distilled water and 0. 1 M HCl. Intrinsic viscosity, which provides information about polymeric chain conformation, was assessed in both media for HCS and bovine submaxillary mucin. Changes in the specific viscosity of HCS-mucin mixtures were observed as a function of the polymer:mucin weight ratio. The formation of interaction products was indicated by a minimum in the specific viscosity. Such a minimum occurred at different polymer:mucin weight ratios depending on the hydration medium and mucin type. This suggested a different stoichiometry of interaction. Turbidimetric measurements were also effected in order to evidentiate the eventual precipitation of the polymer-mucin interaction products. While in distilled water the precipitation of the interaction product did occur, in acidic medium, although a minimum in specific viscosity was observed, no precipitate was formed. The two techniques employed, viscosimetric and turbidimetric, allowed us to investigate for both mucins the influence of hydration medium on the formation of the HCS-mucin interaction products and to conclude that a slightly acid-neutral pH favours the interaction between HCS and mucins.

Characterization of a diltiazem-lambda carrageenan complex.

In the present paper the interaction between lambda carrageenan, a natural sulphated polysaccharide, and diltiazem HCl, a Ca channel blocking agent, was studied. Dialysis equilibria were performed to quantify the binding capacity of lambda carrageenan for diltiazem. The relevance of the interaction to hydrophilic matrix systems was confirmed: a relationship was found between the binding capacity and the release profiles of matrix tablets containing a fixed amount of drug and different percentages of lambda carrageenan. Dialysis equilibria in buffered media showed that the interaction is quite insensitive to the pH of the medium (in the range 1.8-6.8), while it is reduced by increasing ionic strength; this behaviour is in line with the importance of ionic bonds in diltiazem-carrageenan interaction. On the basis of the calculated binding capacity, the complex was prepared, dried and milled. A preliminary characterization of the diltiazem-carrageenan complex in the solid state was effected by means of X-ray and DSC analysis. The amount of drug going into solution from the complex was not significantly affected by the pH of the medium (in the range 1.8-6.8), while it was increased by increasing ionic strength.

Characterization of rheological and mucoadhesive properties of three grades of chitosan hydrochloride.

In the present study the rheological and mucoadhesive properties of three viscosity grades of a cationic polymer, chitosan, were investigated to assess the suitability of this polymer for gastroadhesive formulations. The influence of the pH, ionic strength and temperature of the hydration medium on the rheological properties was studied. The mucoadhesive performance was assessed in vitro by means of rheological synergism and tensile stress testing. A correlation was found between rheological synergism and tensile parameters (force of detachment and work of adhesion), which proves that both methods can correctly predict the mucoadhesive performance of the polymers examined. It was found that the interaction of chitosan with mucin decreases on increasing molecular weight.

Factorial analysis of the influence of dissolution medium on drug release from carrageenan-diltiazem complexes.

This research studied the influence of buffer composition, pH, and ionic strength on the release of diltiazem hydrochloride from a complex of the drug with lambda carrageenan. Two viscosity grades of carrageenan were also compared. A factorial analysis was used to evaluate the influence of individual variables and their interactions. Both the complex solubility, measured as the drug concentration in equilibrium with the solid complex, and the drug release rate from constant surface area were considered. The increase of ionic strength significantly increased complex solubility in all the buffer systems. A significant effect of polymer grade on complex solubility was evidenced only in phosphate buffer with a pH of 6.8, indicating lower solubility of the complex when higher polymer molecular weight was involved. In most cases, drug release rate decreased when high polymer grade was involved in the complex. Ionic strength did not always have a significant effect on drug release rate and was quantitatively less important than for solubility. Ionic strength especially affected the drug release profiles. At higher ionic strength drug release was no longer constant, but decreased with time, probably because of lower polymer solubility.

Rheological and mechanical properties of pharmaceutical gels. Part II: Medicated systems: relevance to hydration properties and drug release.

Aim of this work was to evaluate the influence of rheological and mechanical properties of medicated gels containing two viscosity grades of either sodium carboxymethylcellulose or polyacrylic acid on hydration properties and drug release. The two polymers were chosen since they produce different semi-solid systems: polymeric colloidal solutions in the case of sodium carboxymethylcellulose and true gels in the case of polyacrylic acid. Salicylic acid and sodium salicylate were used as model drugs for sodium carboxymethylcellulose solutions, benzoic acid and sodium benzoate were employed for polyacrylic acid gels. The relationship previously found (Part I of this study) between the viscoelastic (tg delta) and the gel strength parameters in non-medicated gelified vehicles also exists in medicated gels: the higher the tg delta, the lower the gel strength. The results obtained on isoviscous samples (either colloidal solutions or true gels), containing two viscosity grades of the same polymer, demonstrate that an increase in tg delta and the concomitant decrease in the mechanical resistance of the sample always accompany an improvement in hydration properties. The different rheo-mechanical and hydration properties entrain different release profiles only when drugs are suspended in the vehicle (i.e. in the case of the free acids here considered). In these cases, the limiting step for drug release is likely the preliminary drug dissolution, which is in turn affected by the hydration propensity of the vehicle.

Rheological and mechanical properties of pharmaceutical gels. Part I: Non-medicated systems.

Non-medicated gel systems based on polymers of different chemical nature and source have been characterized for rheological as well as for gel strength properties: a synthetic product, polyacrylic acid and a natural product, scleroglucan, both known to produce true gels, and two semi-synthetic materials, sodium carboxymethylcellulose and hydroxypropylmethylcellulose, both known to originate polymeric colloidal solutions, have been chosen. In particular, the rheological analysis involved the comparison of the different polymers on the basis either of viscosity (constant rate test) or of viscoelastic measurements (oscillation test). Subsequently the possible relationships between either viscosity or viscoelastic parameters (G', G", tg delta) and the gel strength parameter have been investigated. The mechanical resistance of the pharmaceutical gels examined didn't depend on the viscosity but rather on the viscoelasticity of the systems. In particular, low tg delta values were always accompanied by high gel strength values. A rank order correlation was found between the tg delta values obtained for the various samples at different frequency values and the relevant gel strength parameter: this suggests that mechanical resistance measurements, similarly to viscoelastic ones, give information on the structural properties of the samples and therefore represent a suitable tool in the optimization of gel formulations.

Rheological properties and diffusion dissolution behaviour of hydrophilic polymers.

In a mathematical model that has been recently proposed to predict drug release from polyvinyl alcohol matrices, polymer dissolution is described as a chain disentanglement process and characterized by a threshold polymer concentration. In present work viscosity measurements were employed for defining the characteristic entanglement concentration of polymer solutions. The viscosity of aqueous polymer solutions of differing concentrations was measured over a wide range of shear rate using a rotational viscometer. The flow curves were fitted according to an asymptotic model and from the best fit equation the relevant viscosity parameters were obtained. The relationships between polymer concentration and viscosity parameters were also examined. The critical concentration at which an abrupt change of viscosity properties occurred was identified and related to the molecular chain disentanglement conditions.

A computer-aided simulation approach in the development of a prolonged release formulation.

Prolonged medication with diltiazem has proved advantageous in the treatment of coronary insufficiency and arterial hypertension. Consistently, a number of extended release formulations, based on different retardation mechanisms, have been proposed. In the present work two prolonged release oral formulations containing diltiazem, one intended for twice-a-day and one for once-a-day administration, were tested for in vitro behaviour; the in vitro release test had been opportunely validated using an in vitro-in vivo correlation approach. On the basis of in vitro release profiles, simulations were effected, using a computer program previously developed, in order to generate the plasma levels that could be expected on single dosing of the two formulations. The two formulations were then tested in vivo and the measured plasma profiles were compared with those predicted from in vitro data. For both formulations, a good agreement was found between the measured and the simulated plasma levels, thus demonstrating the usefulness of the simulation approach in the formulative development.

[Particulate contamination of amino acid solutions for parenteral use]. / Studio sulla contaminazione particellare di soluzioni di aminoacidi per uso parenterale.

Three industrial batches of a 10% (w/v) amino acid solution have been checked for particulate contamination. The batches differed in preparative technology and/or type of additives. In particular, one batch was prepared under nitrogen, filled under vacuum and with the addition of sodium metabisulfite and cysteine. The other two batches were simply prepared under nitrogen and differed in the presence or not of cysteine and sodium metabisulfite. Accelerated stability tests were performed (40, 50, e 60 degrees C) and controls were effected both at room (25 degrees C) and refrigerator (6 degrees C) temperature. The results show that the preparation under nitrogen and the presence of cysteine result in an improved chemical stability and allow the particulate contamination level to be maintained within the limits of the Italian Pharmacopoeia. In particular, the batch prepared under nitrogen and filled under vacuum is characterized by a particulate contamination level which remains within the more restrictive limits of the British Pharmacopoeia for the entire period of the stability studies. Moreover, the particulate contamination control performed at 6 degrees C has proved to be a useful predictive tool with regards to batch quality evaluation.