Silvia Tampucci

Validation of bovine hoof slices as a model for infected human toenails: in vitro ciclopirox transungual permeation.

Background  Topical therapy has recently been proposed for treating onychomycosis and other nail disturbances. However, the clinical outcome may be limited by the difficulty of active ingredients effectively penetrating the nail plate. Bovine hoof membranes have been widely used to predict in vitro efficacy of drug products in nail diseases. Many studies have compared bovine hooves with human healthy nails, considering the difference between healthy and unhealthy nails to be negligible.

Span and Tween neutral and pH-sensitive vesicles: Characterization and in vitro skin permeation

The aim of this work was the preparation, characterization, and comparison of novel pH-sensitive nonphospholipid vesicles (niosomes) from two nonionic surfactants, with different hydrophilic-lipophilic balance values (Tween®20-TW20 = 16.7 and Span®60-SP60 = 4.7). Surfactants were mixed with cholesterol (CHOL) and its derivative, cholesteryl hemisuccinate (CHEMS), as a pH-sensitive molecule. Vesicles were characterized by dynamic light scattering, in order to evaluate their dimensions and vesicle stability, by ζ-potential measurements and by means of electronic microscopy after freeze-fracture. Ibuprofen (IBU) was used as the model drug, and high-performance liquid chromatography analyses were performed to evaluate drug-entrapment efficiency and release in a neutral, acidic environment. The influence of the vesicle composition on skin accumulation and transdermal permeation of IBU across excised hairless rat skin was investigated by using vertical Gummer diffusion cells. When niosomes with SP60 and CHEMS were prepared, there was a statistically significant increase of skin permeation of IBU, while TW20 niosomes did not show statistically significant differences in Papp values without the influence of the vesicle size and charge.

Solid lipid nanoparticles as promising tool for intraocular tobramycin delivery: Pharmacokinetic studies on rabbits

Eye drops are widely accepted as formulations for targeting the anterior segment notwithstanding their limitations in terms of bioavailability. The unique structure of the eye requires specially-designed formulations able to favor the pharmacokinetic profile of administered drugs, mainly minimizing the influence of ocular barriers. Nanotechnology-based delivery systems lead to significant technological and therapeutical advantages in ophthalmic therapy. The aim of the present study was to determine whether tobramycin as ion-pair incorporated in mucoadhesive Solid Lipid Nanoparticles (SLN) reaches the inner parts of the eye favoring drug activity. After technological characterization of the tobramycin entrapped SLN formulation (Tobra-SLN), a pharmacokinetic study in rabbits after topical instillation and intravenous administration of the formulation has been carried out. In addition, the intracellular activity of Tobra-SLN formulation against phagocytosed Pseudomonas aeruginosa was investigated. The SLN were spherical in shape, and showed a hydrodynamic diameter of about 80nm, a negative zeta potential (-25.7mV) with a polydispersity index of 0.15, representative of a colloidal dispersion with high quality, characterized by an unimodal relatively narrow size distribution. As demonstrated by FTIR and DSC, tobramycin ion-pair could be concentrated into lipid inner core of SLN, without interaction with the stearic acid, thus promoting a slow and constant drug release profile in the dissolution medium. Surprisingly, the drug concentration was significantly higher in all ocular tissues after ocular and intravenous administration of Tobra-SLN formulation with respect to reference formulations and only Tobra-SLN allowed the penetration of drug into retina. Furthermore, the use of Tobra-SLN resulted in both higher intraphagocytic antibiotic concentrations in polymorphonuclear granulocytes and greater bactericidal activity against intracellular Pseudomonas aeruginosa, probably due to the ability of Tobra-SLN to penetrate either into phagocytic cells, or alternatively to cross bacterial barrier. The present study broadens the knowledge on the use of SLN as carriers for ocular drug delivery to the posterior chamber and might open new avenues for treatment of ocular infections, representing a strategy to overcome the microbial resistance.

Liposomes as a potential ocular delivery system of distamycin A.

Liposomes containing Distamycin A (DA) may be clinically useful in the treatment of ocular HSV infections, especially in acyclovir-resistant HSV keratitis. This study evaluated the in vitro and in vivo performance of a topical controlled release liposomal formulation containing DA (DA-Lipo) aimed at reducing the toxicity of the encapsulated active agent and improving drug uptake by ocular tissues. The bioavailability of DA in the tear fluid and the DA uptake into the cornea were increased after instillation of DA-Lipo in rabbits, reaching the DA corneal concentration corresponding to IC50 values against HSV without any sign of transcorneal permeation of drug. DA-Lipo was definitely less cytotoxic then plain DA in rabbit corneal epithelial cells. These results provide new insights into the correlation between the in vitro data and the drug kinetics following ocular applications of liposomal vesicles.

Optimization of skin permeation and distribution of ibuprofen by using nanostructures (coagels) based on alkyl vitamin C derivatives

In this investigation two vitamin C-based -6-O-ascorbic acid esters (ASC₁₂ and ASC₁₆), able to form liquid-crystal structures (coagels) was evaluated for their potential usefulness to promote the permeation and distribution of ibuprofen (IBU). Two coagel formulations and the same coagels added of polyethylene glycol (PEG-400) were assayed in comparison with a commercial product (Arfen®) by using hairless rat skin as model. The ASC₁₆ and ASC₁₂ derivatives gave rise to stable supramolecular assemblies in water and in water/PEG mixtures (coagels), allowing the solubilization of IBU (0.85%) and producing a IBU controlled release systems, as evidenced by the dynamic dialyse test: the n values were near 1.0, indicative of a linear kinetic, for all coagel formulations, except for the ASC₁₂PEG/C formulation (n=1.51). Our results evidenced the enhancement activity of coagels and the synergic effect of the combination with PEG: all coagels showed a higher amount of IBU permeated through the skin compared to commercial Arfen® with an enhancement factor of 52.94 and 21.53 for ASC₁₂PEG/C and ASC₁₆/C respectively. Otherwise, coagels formulations appeared to produce a low IBU depot in the skin and in the same order of magnitude in epidermis and derma, in spite of significant increase of IBU cutaneous permeation. The positive synergic effect of the coagel-PEG mixtures was demonstrated by the high amount of IBU accumulated in the upper skin layers. The effect of the coagels on the IBU skin permeation and distribution depending on their hydro-lipophilic character could allow a rational design and an optimization of topical formulations.

Skin permeation and distribution of two sunscreens: a comparison between reconstituted human skin and hairless rat skin

The aims of this work were (a) to develop a simple and reproducible procedure for percutaneous absorption and distribution tests of sunscreens using one human skin culture model (Epiderm™ 606; reconstructed epidermis, RE), (b) to compare the said model with rat skin (RS) in vitro and (c) to evaluate the effect of different formulations. The cutaneous permeation and distribution of two UV filters, ethylhexylmethoxycinnamate (MC80) and ethylhexyltriazone (T150), using 3 different vehicles were investigated. The permeation studies demonstrated that neither MC80 nor T150 permeated through both RS and RE in spite of different thicknesses of the 2 substrates. Distribution studies demonstrated that sectioning by cryomicrotome to obtain horizontal skin layers was suitable for both RS and RE (apart from its small thickness) with a good reproducibility of data. The amounts of sunscreens retained in the 2 substrates were in the same order of magnitude for all formulations with a greater depot in RS. Different distribution profiles of the tested formulations could be ascribed to the different lipid compositions of RE and RS. Since the physicochemical characteristics of RE are closer to those of human skin, the results obtained with reconstructed human skin models could be suitable to replace human skin in ‘in vitro testing’.

Ionic liquids as potential enhancers for transdermal drug delivery

The aim of this study was to verify the effect of several cyclic onium based ionic liquids (ILs), including mono- and dicationic derivatives of 1,4-diazabicyclo[2.2.2]octane (DABCO), a dialkyl morpholinium salt and a Brønsted acidic IL, as enhancers of the in vitro transdermal permeation and skin retention of diltiazem through and into hairless rat skin. The drug was used as both the hydrochloride salt (DZHCl) and the free base (DZB) to highlight the relationship between the enhancement effect and the physico-chemical characteristics of the active agent. Permeation tests were carried out using Gummer-type diffusion cells and excised rat skin with a 0.005M aqueous solution of diltiazem hydrochloride or diltiazem free base with and without the addition of 1% w/w ionic liquids. At the end of the permeation experiments with diltiazem hydrochloride, a suitable extraction procedure allowed for the determination of the drug content retained in the skin. Depending on the ionic liquid structure, a significant enhancement in diltiazem hydrochloride levels in the receiving phase was observed, and the transdermal permeation of the diltiazem free base was markedly increased by treatment with all of the ionic liquids. N-dodecyldabco bromide was the best enhancer for both salified and free base drug forms, even though it showed a certain toxicity. On the other hand, N-methyl-N-decylmorpholinium bromide showed a good balance between enhancer activity and cytotoxicity.

Topical Formulations Containing Finasteride. Part I: In Vitro Permeation/Penetration Study and In Vivo Pharmacokinetics in Hairless Rat

In hair follicle (Hf) cells, the type-2 5-α-reductase enzyme, implicated in androgenetic alopecia, is selectively inhibited by finasteride (FNS). Because an effective topical formulation to deliver FNS to Hf is currently unavailable, this investigation aimed at evaluating in vitro FNS skin permeation and retention through and into hairless rat and human abdominal skin. Four hydroxypropyl chitosan (HPCH)-based formulations (P-08-012, P-08-016, P-08-063, and P-08-064) and one anhydrous formulation without HPCH (P-10-008) were tested. The pharmacokinetics in plasma and skin after application of P-08-016 or P-10-008 on dorsal rat skin with single and repeated doses was investigated. P-08-016 performed the best in driving FNS to the reticular dermis without producing a high transdermal flux. Neither the in vivo single nor the repeated dose experiments produced plasma levels of FNS and no differences were found between formulations concerning skin retention. No increase in the amount of drug retained in the skin was obtained with the repeated dose experiment. In conclusion, the HPCH-based formulation P-08-016 might represent an alternative to systemic therapy for its ability to promote a cutaneous depot of FNS in the region of hair bulbs, minimizing systemic absorption even after repeated treatments.

In vitro evaluation of some parameters involved in mucoadhesion of aqueous polymeric dispersions.

Abstract Context: The mucoadhesive formulations are constantly developing due to their relevance in the drug delivery to various districts of the organism. Objective: The purpose of this study was to find a direct link between physicochemical properties of the polymers and their adhesive ability in order to offer guidelines for the development of mucoadhesive semisolid formulations. Materials and methods: Twelve polymers were dispersed in water and characterized with regard to their mucoadhesiveness, apparent viscosity, contact angle on solid surface, and hydrodynamic diameter of their molecules. The adhesive properties were related to the other measured parameters. Results and discussion: The data seem to indicate the existence of an optimal value of viscosity, around 5–6 Pa s, to obtain the highest mucoadhesiveness of the polymeric dispersions. Regarding the molecular sizes, the best mucoadhesive performances seem to be given from polymers with a hydrodynamic diameter lower than 350–400 nm. In any case, the ability to wet the surface by the polymeric dispersion seems to play an essential role in bioadhesion process, capable of strongly limiting the phenomenon. Conclusions: Performing simple in vitro measurements, it seems possible to identify the best polymeric concentration to obtain a semisolid formulation with good mucoadhesive properties.

Topical Formulations Containing Finasteride. Part II: Determination of Finasteride Penetration into Hair Follicles using the Differential Stripping Technique

The differential stripping technique consists of a tape-stripping phase followed by a cyanoacrylate biopsy. This technique not only allows the quantification of drug retained in the stratum corneum (SC) and in the hair follicles but also differentiates transepidermal from transfollicular penetration. Our study aimed at both validating the differential stripping procedure on hairless rat skin and assessing the role of the hair follicle in the cutaneous penetration of finasteride (FNS) after application of two experimental formulations for 6 or 24 h: P-08-016, a hydroxypropyl chitosan (HPCH)-based formulation and P-10-008, an anhydrous formulation devoid of HPCH. Microscopic and histological evaluation showed that after 15 tape strips both the SC and the viable epidermis were completely removed. A subsequent cyanoacrylate skin surface biopsy led to the removal of the infundibula content. The largest amounts of FNS were found in the epidermis and in the appendages after application of P-08-016, regardless of the time from application. In contrast, smaller and statistically significant amounts of FNS were recovered with P-10-008 6 h after application, compared with that at 24 h. In conclusion, the differential stripping technique allowed determination of the amount of FNS localized in different skin districts, focusing particularly on the follicular contribution.