Claudia Valenta

Design and in vitro evaluation of a novel bioadhesive vaginal drug delivery system for clotrimazole.

It was the purpose of this study to design and evaluate a new bioadhesive vaginal drug delivery system for clotrimazole. Chitosan, a cationic biopolymer derived by deacetylation of chitin, was modified by the introduction of thioglycolic acid (TGA). The modification was achieved by utilising a carbodiimide to link the carboxylic acid moieties of TGA covalently to the primary amino groups of chitosan. The amount of added carbodiimide was thereby varied, resulting in chitosan-TGA conjugates A and B with 160 microM (=micromol) and 280 microM thiol groups per gram polymer, respectively. In order to characterise the new polymers the water uptake, the disintegration behaviour, the bioadhesive properties utilising the rotating cylinder method, as well as the release of clotrimazole from tablets based on these derivatives were studied. The water uptake and cohesive properties of vaginal tablets consisting of these new conjugates could be significantly (p<0.05) improved. By adding clotrimazole the disintegration time of the conjugates was prolonged 1.6-fold for conjugate A and even 100-fold for conjugate B. Furthermore, the adhesion on vaginal mucosal tissue could be significantly improved. The addition of clotrimazole had also an impact on the adhesion time of chitosan-TGA conjugate B, which remained 26-times longer on vaginal mucosa than the corresponding unmodified polymer. The immobilisation of thiol groups guarantees a controlled drug release. Results of this study demonstrate that these new chitosan-TGA conjugates are very promising vehicles for the vaginal application of clotrimazole in treatment of mycotic infections.

pH, pKa and dermal delivery

The effect of pH on the permeation of ibuprofen and lignocaine through human skin has been modelled using a modification to the equation derived by Potts and Guy, which is normally applied to unionized entities. The results show that permeation is related to the distribution coefficient. The physicochemical properties have been predicted ab initio using commercially available software and compared to literature values. The approach is successful and shows that there is significant permeation of the ionized drugs through a lipophilic pathway, possibly as a result of ion pairing. Since the aqueous solubility of the ionized material is significantly higher than the unionized, the maximum flux through the skin may occur at a pH where ionization is high. Optimum topical or transdermal formulations may not therefore be for the free acid or free base.

Evaluation of novel soya-lecithin formulations for dermal use containing ketoprofen as a model drug

In this study soya-lecithin aggregates, prepared by a technique using compressed gas, are used to formulate new dermal preparations. Ketoprofen (KP), a nonsteroidal anti-inflammatory drug (NSAID) is included as a model drug. The technique offers the possibility of incorporating auxiliary agents, such as penetration enhancers, anti-irritants and moisturisers together with the drug in one process. Apparent partition coefficients for n-octanol-phosphate buffer were determined for each of the lecithin aggregates. In general, soya-lecithin improves the partition of KP into n-octanol. The resulting products were included in widely used hydrophilic and hydrophobic vehicles. After 24 h, the cumulative amount of drug released through an artificial membrane was higher from the hydrophilic gels (2.6-4.3 mg) and the hydrophobic creams (0.23-0.392 mg) than from the control preparations (control hydrogel: 1.3 mg; control hydrophobic cream: 0.141 mg). However, the cumulative amount released from the hydrophobic vehicles was generally lower than from the hydrophilic matrices. Cumulative amounts such as those released from the hydrophilic preparations can also be achieved using supersaturated formulations based solely on the drug-loaded lecithin aggregates and a suitable oily component (4.07 mg). Results from the diffusion studies using artificial membranes were confirmed by permeation studies using excised rat skin. The improvement in skin permeation is related to both the solubilising effect of the lecithin matrix and the penetration enhancing effect of lecithin itself. The novel soya-lecithin aggregates are promising candidates for new drug delivery systems in dermatology and cosmetology. Lecithin aggregates loaded with drugs are multifunctional carriers that also act as penetration enhancers.

Electron microscopy of nanoemulsions: an essential tool for characterisation and stability assessment.

The characterisation of pharmaceutical formulations by microscopic techniques is essential to obtain reliable data about the actual morphology of the system. Since the size range of colloidal drug delivery systems has long ago reached the lower end of the nanometer scale, classical light microscopy has been replaced by electron microscopy techniques which provide sufficient resolution for the visualisation of nano-sized structures. Indeed, the superior resolution and methodological versatility of electron microscopy has rendered this technique an indispensable tool for the analysis of nanoemulsions. Microscopic analysis of these lipid-based drug delivery systems with particle sizes in the lower submicron range provides critical information about the size, shape and internal structure of the emulsion droplets. Moreover, surfactant aggregates such as liposomes or multilamellar structures which remain unnoticed during particle size measurements can be detected in this fashion. This review provides a brief overview about both transmission electron microscopy (TEM) and scanning electron microscopy (SEM) techniques which have been employed to characterise nanoemulsions. Of special interest are sophisticated cryo techniques of sample preparation for both TEM and SEM which deliver high-quality images of nanoemulsions in their natural state. An overview about the instrumentation and sample preparation for all presented methods is given. Important practical aspects, sources of error and common artefacts as well as recent methodological advances are discussed. Selected examples of electron microscopic studies of nanoemulsions are presented to illustrate the potential of this technique to reveal detailed and specific information.

Lecithin based nanoemulsions: A comparative study of the influence of non-ionic surfactants and the cationic phytosphingosine on physicochemical behaviour and skin permeation

Charged drug delivery systems are interesting candidates for the delivery of drugs through skin. In the present study, it was possible to create negatively and positively charged oil/water nanoemulsions by using sucrose laureate and polysorbate 80 as non-ionic surfactants. The positively charged nanoemulsions were generated by adding cationic phytosphingosine (PS). The relationship between the physicochemical properties of the nanoemulsions was shown by particle size and zeta potential measurements. These properties were dependent on the type of non-ionic surfactant and the concentration of PS. Furthermore the cationic PS had a positive impact on the skin permeation rates (flux) of the incorporated model drugs fludrocortisone acetate and flumethasone pivalate. An enhancement factor between 1.1 and 1.5 was obtained in relation to the control. The interaction of pre-impregnated porcine skin with positively and negatively charged nanoemulsions was confirmed by DSC analysis. The generated DSC-curves showed a slight difference in the phase transition temperature assigned to the characteristic lipid transition. However, it was not possible to assign the effect to one of the ingredients in the multicomponent system.

Development and in vitro evaluation of a mucoadhesive vaginal delivery system for progesterone.

The purpose of the present study was to design a novel carrier system based on a mucoadhesive polymer exhibiting improved properties concerning drug delivery to the vaginal mucosa. This was reached by the covalent attachment of L-cysteine to commercially available polyacrylic acid (Carbopol 974P). Mediated by a carbodiimide, increasing amounts of L-cysteine were covalently linked to the polymer. The resulting thiolated polyacrylic acid conjugates (NaC974P-Cys) displayed between 24.8 and 45.8 micromol thiol groups per gram of polymer. Because of the formation of intra- and/or intermolecular disulfide bonds, the viscosity of an aqueous thiolated polymer gel (3%) increased about 50% at pH 7.0 within 1 h. In oscillatory rheological measurements, it was shown that this increase in viscosity is mainly due to the increase in elasticity. Tensile studies carried out on freshly excised cow vagina demonstrated a significant (P<0.05) increase in the total work of adhesion (TWA) compared to the unmodified polymer. An amount of 24.8 micromol thiol groups per gram of polymer resulted in a 1.45-fold increase in the TWA, whereas an amount of 45.8 micromol showed an even 2.28-fold increase. These improved mucoadhesive properties can be explained by the formation of disulfide bonds between the thiolated polymer and cysteine rich subdomaines of the mucus layer. The release rate of the model drug progesterone from tablets based on microcrystalline cellulose serving as the reference was approximately 1% per hour, whereas it was 0.58% per hour for the unmodified polymer (NaC974P) and 0.12% per hour for the thiolated polymer (NaC974P-Cys). Therefore, this thiolated polymer is a promising carrier for progesterone providing a prolonged residence time and a controlled drug release.

The dermal delivery of lignocaine: influence of ion pairing.

The purpose of the present study was to determine the significance of ion pairing on the permeation of lignocaine. Results of diffusion studies through polydimethylsiloxane (PDMS) at different pH values 4. 0, 6.0, 7.0, 8.0 indicated that lignocaine hydrochloride (L-HCl) flux significantly increased with the amount of unionized base. In order to see if similar results could be obtained using human skin, permeation runs were performed with human skin at pH of 4.0, 5.5 and 7.0. These values were chosen to simulate an appropriate range of physiological conditions. Results of the experiments with human epidermis showed increasing L-HCl flux with increasing pH, confirming the trends seen with PDMS membranes. A linear relationship was found between the apparent partition coefficient and the steady state flux. Further experiments were conducted at donor pH 4.0 to minimise the contribution of the unionized species. Although an excess of different ions such as nitrate, mesylate and bromide increased the apparent partition coefficient, the steady state flux was not significantly increased. The steady state lignocaine flux was increased up to 2.45-fold using different counter ions. The highest flux was measured from lignocaine morpholinopropane sulfonate (L-mps). It is possible to enhance the flux of salts across lipophilic membranes by using an ion pair approach. The degree to which this is possible depends on the lipophilicity of the counter ion, the medium in which the ion pair forms, and the ionic strength.

In vitro vs. in vivo tape stripping: Validation of the porcine ear model and penetration assessment of novel sucrose stearate emulsions

Porcine ear skin is frequently used as a substitute for human skin in dermatological research and is especially useful for tape stripping experiments where the penetration of active substances into the uppermost skin layers is investigated. However, certain differences between the surface properties of these skin types exist, and reports on the comparability of tape stripping data obtained in vitro using porcine ear skin and data obtained in vivo on human forearm skin are scarce. Thus, we performed comparative tape stripping experiments in which the skin penetration of curcumin and fluorescein sodium from conventional microemulsions and hydrogels was investigated. In this context, the skin penetration potential of novel semi-solid macroemulsions and fluid nanoemulsions based on sucrose stearate was evaluated as well. The removed corneocytes were quantified by NIR-densitometry using recent correlation data for human and porcine proteins. The trends observed for the skin penetration into porcine ear skin were highly representative for the in vivo situation on human skin, confirming that the porcine ear is an excellent in vitro model for tape stripping experiments. Moreover, the validity of the NIR-densitometric approach for the quantification of both human and porcine stratum corneum proteins was confirmed in this study for the first time.

Lecithin-based nanoemulsions

Lipid-based emulsions with particle sizes in the submicron range have been extensively investigated as vehicles for various active agents during the last decades. This review summarizes the current knowledge about nanoemulsions with a special focus on nanoemulsions comprising lecithin as the main emulsifier. A short introduction is given on the origin of lecithin-based nanoscale emulsions and the properties of these metastable systems. The differences between nanoemulsions and microemulsion phases are highlighted and recent discrepancies in terminology are discussed. Furthermore, the peculiarities of lecithin as emulsifying agent are presented. The production and optimization of nanoemulsions as well as the controversy concerning the different proposed production methods is another important aspect that is covered in detail. Moreover, general information about the stability and characterization of lecithin-based nanoemulsions is included. The last part of the review deals with current applications and research focuses with emphasis on the application of lecithin-based nanoemulsions on skin.

Novel Bioadhesive Chitosan-EDTA Conjugate Protects Leucine Enkephalin from Degradation by Aminopeptidase N

AbstractPurpose. To develop a novel bioadhesive polymer that protects peptide drugs from luminal degradation by aminopeptidase N and to evaluate the system in vitro on porcine mucosa. Methods. EDTA was covalently bound to chitosan in order to combine the bioadhesive properties of the polymer with the well known capacity of EDTA to complexe metal ions which are essential for the enzymatic activity of proteases. The inhibitory effect of this polymer conjugate was evaluated by using leucine enkephalin (Leu enkephalin) as a model drug. The degree of Leu enkephalin degradation caused by aminopeptidase N (EC 3.4.11.2), as well as porcine mucosa, in the presence of the polymer conjugate, was quantified by HPLC analysis. Results. The chitosan-EDTA conjugate is capable of binding 2.01 ± 0.12 mmole of zinc per gram of polymer at pH 6.5 (n = 3; ± S.D.). As zinc is an essential co-factor for aminopeptidase N, enzyme activity (48 mU/ml) could be completely inhibited under the use of 1.0% chitosan-EDTA conjugate. The inhibitory effect of 1.0% chitosan-EDTA conjugate on the degradation of Leu enkephalin on porcine mucosa within 3 h at 37°C was even 2.9-fold higher than that of a recently developed zinc complexing bacitracin-poly(acrylic acid) conjugate of the same concentration. The novel polymer conjugate is more bioadhesive than unmodified chitosan and is easily hydratable in water and basic aqueous solutions exhibiting quick swelling properties. Conclusions. The bioadhesive polymer conjugate described here seems to be a useful tool in overcoming enzymatic degradation by aminopeptidase N.