Effect of ethanol/propylene glycol on the in vitro percutaneous absorption of aspirin, biophysical changes and macroscopic barrier properties of the skin.

The effect of the solvent systems ethanol (EtOH), propylene glycol (PG) and combinations thereof was examined on the in vitro percutaneous absorption of the antithrombotic, aspirin, through porcine epidermis. Biophysical changes in the stratum corneum lipids were studied through the use of Fourier transform infrared (FTIR) spectroscopy. Macroscopic barrier properties of the epidermis were examined through the use of in vitro transepidermal water loss (TEWL). The flux of aspirin increased with increasing concentrations of EtOH in the solvent systems. The maximum flux of aspirin was achieved by 80% EtOH in combination with 20% PG beyond which (i.e. 100% EtOH) there was no increase in the flux. FTIR spectroscopic study was enacted in order to determine the biophysical properties of the stratum corneum when the solvents were applied. The FTIR spectra of the stratum corneum treated with 80% EtOH/20% PG showed a maximum decrease in absorbance for the asymmetric and symmetric C&z. sbnd;H peaks, which suggests a greater loss of the lipids in the stratum corneum layers. In vitro TEWL studies allowed an investigation into the macroscopic barrier integrity properties of the stratum corneum. The TEWL results indicated that each of the solvent systems significantly enhanced (P<0.05) in vitro TEWL in comparison to the control. In conclusion, 80% EtOH/20% PG enhanced the percutaneous absorption of aspirin by perturbing the macroscopic barrier integrity of the stratum corneum and through a loss of stratum corneum lipids. Copyright     

Effects of isopropanol-isopropyl myristate binary enhancers on in vitro transport of estradiol in human epidermis: a mechanistic evaluation

The purpose of this study was to mechanistically investigate effects of isopropanol (IPA)-isopropyl myristate (IPM) binary enhancers on transport of a model drug, estradiol (E2) in human epidermis (stratum corneum + viable epidermis) in vitro. The study was focused on use of the same IPA-IPM compositions on both sides of the skin ("symmetric" configuration) with saturated E2 (maximum thermodynamic activity). For E2 transport in all IPA-IPM compositions tested, stratum corneum still was the rate-limiting layer of human epidermis. The relative contributions to E2 flux enhancement were separated into the changes in solubility and diffusivity of E2 in stratum corneum. As a major factor, E2 solubility in stratum corneum was enhanced by 35 times with increasing IPA from neat IPM to neat IPA. E2 diffusivity in stratum corneum also played a significant role, which increased by 8 times from neat IPM to 50% IPA. Stratum corneum swelled more in IPM-rich region, decreased with increasing IPA, and even deswelled in neat IPA. IPA uptake correlated well to E2 solubility in stratum corneum; both linearly increased with increasing IPA. IPM uptake appeared to correlate to E2 diffusivity in stratum corneum; both maximized around 50% IPA.     

Magnetic resonance profiling of human skin in vivo using GARField magnets

It is shown that one-dimensional magnetic resonance imaging (MRI-profiling) of human forearm and side-of-hand skin in vivo is possible using GARField magnets. Strong profile contrast originating from differing molecular mobility is seen for stratum corneum and viable epidermis. The first in vivo spatially-resolved field-gradient measurements of water self-diffusivity, D, in the stratum corneum (2.0 x 10(-6) cm(2)/s) and viable epidermis (8.5 x 10(-6) cm(2)/s) are reported. Also reported are spatially resolved measurements of the (1)H spin-lattice relaxation time, T(1), the spin-spin relaxation time, T(2). It is further shown that the application of moisturizing agents to the skin noticeably affects the profiles. However, universal behavior is not seen as both signal increases and decreases are observed dependent on agent and volunteer.     

Prediction of drug disposition kinetics in skin and plasma following topical administration

The development of a physically based pharmacokinetic model for percutaneous absorption is described. The simulation includes four first-order rate constants assigned the following significance: (a) absorption across the stratum corneum; (b) diffusion through the viable tissue; (c) a retardation process which retains penetrant in the stratum corneum (and hence provides a means to mathematically produce a "reservoir" effect, for example); and (d) uptake from the skin into the systemic circulation and subsequent elimination from the body. The kinetic equations of the model are solved and expressions are obtained for the concentration of penetrant within the stratum corneum (and available to subsequently partition into the viable epidermis) and the plasma concentration of the administered substance, as a function of time. Using example values for the four rate parameters, disposition profiles for the penetrant in skin and plasma were derived. The cases considered cover slow and fast stratum corneum penetrants, substances which are excreted rapidly or slowly from the body, and absorbing molecules with a variety of relative stratum corneum-viable tissue affinities. The results suggest a framework for the prediction of pharmaceutically and clinically relevant information following the topical administration of therapeutic agents for local or systemic effect.     

Transdermal Delivery of Molecules is Limited by Full Epidermis, Not Just Stratum Corneum

Purpose

Most methods to increase transdermal drug delivery focus on increasing stratum corneum permeability, without addressing the need to increase permeability of viable epidermis. Here, we assess the hypothesis that viable epidermis offers a significant permeability barrier that becomes rate limiting upon sufficient permeabilization of stratum corneum.

Methods

We tested this hypothesis by using calibrated microdermabrasion to selectively remove stratum corneum or full epidermis in pig and human skin, and then measuring skin permeability to a small molecule (sulforhodamine) and macromolecules (bovine serum albumin, insulin, inactivated influenza vaccine) in vitro.

Results

We found that removal of stratum corneum dramatically increased skin permeability to all compounds tested. However, removal of full epidermis increased skin permeability by another 1–2 orders of magnitude. We also studied the effects of removing skin tissue only from localized spots on the skin surface by covering skin with a mask containing 125-μm holes during tissue removal. Skin permeabilized in this less-invasive way showed similar results. This suggests that microdermabrasion of skin using a mask may provide an effective way to increase skin permeability.

Conclusions

We conclude that viable epidermis offers a significant permeability barrier that becomes rate limiting upon removal of stratum corneum.     

Epidermal cell culture model with tight stratum corneum as a tool for dermal gene delivery studies

The purpose of this study was to evaluate the feasibility of organotypic cultures of rat epidermal cells as a tool to study non-invasive dermal gene delivery. Also, a novel transfection method employing liposomal pre-treatment of stratum corneum (SC) was evaluated. Rat epidermal cells were cultured on Transwell tissue culture inserts and formation of stratum corneum barrier was evaluated in permeability studies with two model compounds. Transfections were performed with naked pCMV-SEAP2 plasmid and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP)/dioleyl-phosphatidylethanolamine (DOPE)/DNA lipoplexes. Naked DNA was administered on the stratum corneum of the cell culture model with or without prior treatment of the stratum corneum with DOTAP/DOPE liposomes. Transfection was evaluated non-invasively by monitoring concentrations of secreted alkaline phosphatase (SEAP) in the culture medium of the basolateral compartment at 24-h intervals. Transfection with lipoplexes produced significant gene expression in rat epidermal keratinocyte (REK) epidermal culture model. Likewise, delivery of naked DNA on stratum corneum after DOTAP/DOPE liposome pre-treatment produced gene expression. Naked DNA alone did not result in detectable gene expression. In dermal gene delivery studies REK epidermal culture model is a suitable tool that includes tight stratum corneum and allows transgene expression in viable epidermis and non-invasive sampling of secreted gene product in the basolateral compartment. Liposomal pre-treatment of the stratum corneum augments transfection of viable epidermis.     

Enhancement by Terpenes of 5-Fluorouracil Permeation through the Stratum Corneum: Model Solvent Approach

5-Fluorouracil permeates the stratum corneum through the intercellular pathway. 5-Fluorouracil is hydrophilic and, therefore, its partitioning from the aqueous region into the hydrocarbon interior of stratum corneum lipids is expected to be an important stage of its permeation and a target for some permeation enhancers. It has also been reported that complexation plays a role in the enhancement effect of some accelerants. These mechanisms have been investigated. For partitioning-permeation studies, isooctane was chosen as a model of the hydrocarbon interior of stratum corneum lipid bilayers and the effects of 26 different terpene enhancers on the solubility of 5-fluorouracil in isooctane were measured. Results were then compared with the effects of the same enhancers on the permeation of 5-fluorouracil through the epidermis in man. The stoichiometry of interaction of cineole and limonene with 5-fluorouracil were also studied to reveal possible complex formation. Solubility studies revealed good correlation between solubility and enhancement ratios for the majority of terpenes, indicating that one mechanism by which terpenes increase permeation of the stratum corneum by 5-fluorouracil is by improvement of partitioning. Stoichiometry studies showed that cineole can form 1:1 or higher complexes with 5-fluorouracil. With limonene, only a weak 1:1 complex was indicated. Data obtained using epidermis from man show that the enhancement effect of cineole toward 5-fluorouracil is much higher than that of limonene. These data reveal that terpenes might increase the permeation of 5-fluorouracil through the stratum corneum as a result of complex formation and a form of facilitated transport.     

Interaction of lipid nanoparticles with human epidermis and an organotypic cell culture model

Various lipid nanoparticle formulations were investigated with respect to (trans)dermal drug delivery with special regard to the mechanism of their effects on human and an organotypic cell culture epidermis. Potential alterations of stratum corneum lipid domains were studied using fluorescence assays with labeled liposomes and thermal analysis of isolated stratum corneum. Influences on the permeation of corticosterone were investigated and the occlusive properties of the nanoparticles were determined by measurements of the transepidermal water loss (TEWL). The penetration of a fluorescence dye was visualized by fluorescence microscopy of cross sections of human epidermis after incubation with cubic and solid lipid nanoparticles. Corticosterone permeation was limited when applied in matrix-type lipid nanoparticles (fat emulsion, smectic and solid lipid nanoparticles). An adhesion of solid lipid nanoparticles was clearly observed in thermal analysis as reflected by additional phase transitions probably caused by the nanoparticle matrix lipid. However, as for the other matrix-type nanoparticles, no distinct alterations of the phase transitions of the stratum corneum lipids were observed. Cubic nanoparticles led to the most predominant effect on skin permeation where the surface-active matrix lipid may act as penetration enhancer. An alteration of the stratum corneum lipids' thermal behavior as well as an interaction with fluorescence labeled liposomes was observed. Differences observed in permeation studies and thermal analysis of human and cell culture epidermis indicate that surface lipids, which are not present to the same extent in the cell culture model than in human epidermis, seem to play an important role.     

Skin Solubility Determines Maximum Transepidermal Flux for Similar Size Molecules

Purpose  The maximum flux of solutes penetrating the epidermis has been known to depend predominantly on solute molecular weight. Here we sought to establish the mechanistic dependence of maximum flux on other solute physicochemical parameters. Methods  Maximum fluxes, stratum corneum solubilities and estimated diffusivities through human epidermis were therefore determined for 10 phenols with similar molecular weights and hydrogen bonding but varying in lipophilicity. Results  Maximum flux and stratum corneum solubilities of the phenolic compounds both showed a bilinear dependence on octanol-water partition coefficient (P), with solutes having a maximum solubility in the stratum corneum when 2.7<log P<3.1. In contrast, lag times and diffusivities were relatively independent of P. Stratum corneum-water partition coefficients and epidermal permeability coefficients were consistent with previously reported data. Conclusion  A key finding is that the convex dependence of maximum flux on lipophilicity arises primarily from variations in stratum corneum solubility, and not from diffusional or partitioning barrier effects at the stratum corneum–viable epidermis interface for the more lipophilic phenols. Our data support a solute structure-skin transport model for aqueous solutions in which permeation rates depend on both partitioning and diffusivity: partitioning is related to P, and diffusivity to solute size and hydrogen bonding. (199 words)     

Concentration Dependency in Nicotine Skin Penetration Flux from Aqueous Solutions Reflects Vehicle Induced Changes in Nicotine Stratum Corneum Retention

Purpose

This study sought to understand the mechanism by which the steady state flux of nicotine across the human skin from aqueous solutions is markedly decreased at higher nicotine concentrations.

Methods

Nicotine’s steady state flux through human epidermis and its amount in the stratum corneum for a range of aqueous nicotine solutions was determined using Franz diffusion cells, with the nicotine analysed by high performance liquid chromatography (HPLC). Nicotine’s thermodynamic activity in the various solutions was estimated from its partial vapour pressure and stratum corneum hydration was determined using a corneometer. The amount of nicotine retained in the stratum corneum was estimated from the nicotine amount found in individual stratum corneum tape strips and a D-Squame determined weight for each strip.

Results

The observed steady state flux of nicotine across human epidermis was found to show a parabolic dependence on nicotine concentration, with the flux proportional to its thermodynamic activity up to a concentration of 48% w/w. The nicotine retention in the stratum corneum showed a similar dependency on concentration whereas the diffusivity of nicotine in the stratum corneum appeared to be concentration independent. This retention, in turn, could be estimated from the extent of stratum corneum hydration and the nicotine concentration in the applied solution and volume of water in the skin.

Conclusions

Nonlinear dependency of nicotine skin flux on its concentration results from a dehydration induced decrease in its stratum corneum retention at higher concentration and not dehydration induced changes nicotine diffusivity in the stratum corneum.     

Unexpected Clobetasol Propionate Profile in Human Stratum Corneum After Topical Application in Vitro

PURPOSE: The validity of using drug amount-depth profiles in stratum corneum to predict uptake of clobetasol propionate into stratum corneum and its transport into deeper skin layers was investigated. METHODS: In vitro diffusion experiments through human epidermis were carried out using Franz-type glass diffusion cells. A saturated solution of clobetasol propionate in 20% (V/V) aqueous propylene glycol was topically applied for 48 h. Steady state flux was calculated from the cumulative amount of drug permeated vs. time profile. Epidermal partitioning was conducted by applying a saturated drug solution to both sides of the epidermis and allowing time to equilibrate. The tape stripping technique was used to define drug concentration-depth profiles in stratum corneum for both the diffusion and equilibrium experiments. RESULTS: The concentration-depth profile of clobetasol propionate in stratum corneum for the diffusion experiment is biphasic. A logarithmic decline of the drug concentration over the first four to five tape strips flattens to a relatively constant low concentration level in deeper layers. The drug concentration-depth profile for the equilibrium studies displays a similar shape. CONCLUSIONS: The shape of the concentration-depth profile of clobetasol propionate is mainly because of the variable partitioning coefficient in different stratum corneum layers.     

Penetration enhancement of transdermal delivery--current permutations and limitations

In order to achieve enhanced topical drug delivery, it is necessary to make physical or biomolecular structural alterations to the stratum corneum by suitable techniques or by the use of specific chemical agents or drug carriers. The role of the chemical penetration enhancer is to reversibly alter the barrier properties of the stratum corneum by disruption of the membrane structures or by maximizing drug solubility within the skin. Alternatively, permeant delivery to the dermal vasculature using one of several physical methods to reduce diffusional resistance within the skin may be used to promote drug penetration. In the present article, we summarize the major facets of the diverse spectrum of penetration enhancement techniques that include modification of the stratum corneum, lipid-based delivery systems, drug/vehicle interactions, bypassing the stratum corneum, and electrical techniques of enhancement.     

Changes in electrophysiological properties of rat skin with age

The age-related changes in the electrical and physiological properties of the skin were examined in rats at the ages of 5, 10, 21, 90, and 180 d. The resistance of the stratum corneum, the resistance of the viable skin (epidermis and dermis), and the capacitance of the stratum corneum were analyzed from skin impedance data using an equivalent circuit. With development and aging, the resistance of the stratum corneum and the viable skin increased, whereas the capacitance of the stratum corneum decreased. Physiological characteristics such as the thickness of skin strata and the content of lipid and water in the stratum corneum were also measured. The lipid content in the stratum corneum was constant at all ages. The water content in the stratum corneum decreased, and the thickness of skin strata increased with age. Comparison between electrical data and physiological properties suggested that the increase in the resistance of the stratum corneum with aging is primarily caused by the decrease in the water content and that the capacitance of the stratum corneum and the resistance of the viable skin depend on age-related increases in the thickness of skin strata. In conclusion, the age dependency of cutaneous electrical properties may affect the permeation profile of drugs through the skin, and impedance analysis can be used to estimate age-related changes in transdermal drug delivery.     

Human skin penetration of sunscreen nanoparticles: in-vitro assessment of a novel micronized zinc oxide formulation

The extent to which topically applied solid nanoparticles can penetrate the stratum corneum and access the underlying viable epidermis and the rest of the body is a great potential safety concern. Therefore, human epidermal penetration of a novel, transparent, nanoparticulate zinc oxide sunscreen formulation was determined using Franz-type diffusion cells, 24-hour exposure and an electron microscopy to verify the location of nanoparticles in exposed membranes. Less than 0.03% of the applied zinc content penetrated the epidermis (not significantly more than the zinc detected in receptor phase following application of a placebo formulation). No particles could be detected in the lower stratum corneum or viable epidermis by electron microscopy, suggesting that minimal nanoparticle penetration occurs through the human epidermis.     

Mechanism of urea effect on percutaneous absorption of clonidine

The urea effect on skin permeation of clonidine was investigated to reduce a log time and to increase a permeability. ICR mouse skin and human skin were used and were assumed to be a two-layer membrane consisted of stratum corneum and viable epidermis. The urea acted as a skin denaturant and humectant in the whole epidermis. Also it enhanced the skin permeability of clonidine about 3.5 times. On the other hand, it enhanced the skin permeability by acting as a humectant in the viable epidermis. But the urea effect on the whole epidermis was shown to be greater than that on the viable epidermis. Therefore, it was found that the effect of urea was greater on the stratum corneum than the viable epidermis. Variation of enhancing effect according to the concentration of urea was not found in the range of 1% to 20%.     

Protection against dryness of facial skin: a rational approach

Facial skin is unique in that it gets far more exposure to the external environment than skin on other areas of the body and paradoxically, because it contains the thinnest epidermis and stratum corneum, especially on the eye lid. Environmental attacks contribute to drying of facial skin and damage to the stratum corneum. In recent years, there has been an explosion of new products that contain ingredients that claim to benefit facial skin and protect against environmental damage. This review critically examines the scientific basis, rationale and evidence for inclusion of these ingredients in products for protection of facial skin.     

Effects of etretinate in epidermal metabolism and histology after abrasio of superficial layers. Mouse tail assay

The mouse tail assay is suitable for the histological detection of altered differentiation of the epidermis. In the present study, the stratum corneum of the tail epidermis of female mice (NMRI) was removed by means of fine sandpaper. One group of mice was treated with 2 mg of ethyl (all-trans)-9-(4-methoxy-2,3,6-trimethylphenyl)-3,7-dimethyl- 2,4,6,8-nonatetraenoate (etretinate)/kg body weight daily. The untreated group served as control. After removal of the stratum corneum epidermal hyperplasia and formation of a stratum granulosum was found in the controls (peak on the third day). The alterations declined from the fourth day onwards. The histidine incorporation into protein was increased from days 4 to 6. A similar reaction sequence was observed in the animals treated with etretinate. In addition to the hyperplasia, there was an enlargement of the cells (hypertrophy). The control group showed a significant elevation of leucine incorporation and a reduction of histidine incorporation into protein on the first day after removal of the stratum corneum. The incorporation rate of thymidine triphosphate into DNA peaked on the second day. The rise of leucine incorporation into protein and the increase of thymidine triphosphate incorporation into DNA on the second day after removal of the stratum corneum could not be detected in the animals treated with etretinate. In these animals histidine metabolism could not be influenced by etretinate. In conclusion, etretinate greatly influences the alteration of basal metabolism caused by the removal of the stratum corneum.(ABSTRACT TRUNCATED AT 250 WORDS)     

Percutaneous Absorption of Benzoic Acid Across Human Skin. I. In Vitro Experiments and Mathematical Modeling

The percutaneous absorption of benzole acid across human skin in vitro was experimentally and mathematically modeled. Skin partition coefficients were measured over a range of benzoic acid concentrations in both saline and distilled water. The permeation of benzoic acid was measured across isolated stratum corneum, stratum corneum and epidermis, and split-thickness skin. These experiments demonstrated that the stratum corneum was the rate-limiting barrier and that the flux is proportional to the concentration of the undissociated species. The permeation data were analyzed with a comprehensive non-steady-state mathematical model of diffusion across skin. Two adjustable parameters, the effective skin thickness and diffusivity, were fit to the permeation data by nonlinear regression.     

A New Method for Estimating Dermal Absorption from Chemical Exposure: 2. Effect of Molecular Weight and Octanol-Water Partitioning

A new method for estimating dermal absorption including the effects of exposure time and chemistry is described generally in Part 1 of this series. This method accounts for the larger absorption rates during the initial exposure period as well as the hydrophilic barrier which the viable epidermis presents to lipophilic chemicals. A key parameter in this procedure, the ratio of the stratum corneum and epidermis permeabilities (B) depends on molecular weight and octanol-water partitioning. Several approaches for approximating B and its affect on the dermal absorption prediction are discussed here. Generally, the parameter B is only important for highly lipophilic chemicals which also have relatively small molecular weights. When B is important, the recommended prediction for B is based on the Potts and Guy correlation for human stratum corneum permeability.     

Binding of primaquine to epidermal membranes and keratin

The localisation of primaquine was studied within epidermal membranes following the application of a topical dose. A depth profile was constructed by tape-stripping human epidermis following permeation of a 70 mgml(-1) solution of primaquine in Miglyol 840. Comparative binding studies of primaquine were carried out on isolated human stratum corneum and whole epidermis, using normal and delipidised tissue. An additional study was undertaken using bovine keratin powder as a model of human keratin. The depth profile showed that primaquine decreased with depth and decreasing keratin content, and the total primaquine recovered (15.5 mgcm(-2)) was 300 x the amount of extractable lipid. Binding to delipidised skin was saturable, whereas binding to normal skin was unsaturable, reflecting the high miscibility of drug in the lipid domains as opposed to a finite, but large number of binding sites on the corneocytes. Binding was greater for stratum corneum than stratum corneum plus viable epidermis, probably due to greater accessibility of corneocytes keratin. Binding was dose dependent, although binding to delipidised skin was far greater than to normal skin, demonstrating that primaquine had an affinity for lipoidal regions and an even higher affinity for the proteinaceous domains of the stratum corneum. This was supported by high saturable levels of primaquine binding to bovine horn keratin. The results indicated extensive binding to corneocyte keratin has a significant effect on reservoir formation and the permeability of primaquine across human skin. It is speculated that the large amount of keratin presented at the skin surface may be an evolutionary protective process for the sequestration of ingressing molecules.