Time-Dependent Differences in the Effects of Oleic Acid and Oleyl Alcohol on the Human Skin Barrier.

Oleic acid and oleyl alcohol are commonly used permeation and penetration enhancers to facilitate topical drug delivery. Here, we aimed to better understand the mechanism of their enhancing effects in terms of their interactions with the human skin barrier using diclofenac diethylamine (DIC-DEA), a nonsteroidal anti-inflammatory drug for topical pain management. Oleic acid promoted DIC-DEA permeation through ex vivo human skin more rapidly than oleyl alcohol (both applied at 0.75%) due to fluidization of stratum corneum lipids as revealed by infrared spectroscopy. After 12 h, the effect of these enhancers on DIC-DEA permeation leveled off, fluidization was no longer evident, and skin permeabilization was mainly due to the formation of fluid enhancer-rich domains. Contrary to oleyl alcohol, oleic acid adversely affected two indicators of the skin barrier integrity, transepidermal water loss and skin electrical impedance. The content of oleyl alcohol in the stratum corneum was lower than that of oleic acid (even 12 h after the enhancers were removed from the skin surface), but it caused higher DIC-DEA retention in both epidermis and dermis compared to oleic acid. The effects of oleyl alcohol and oleic acid on DIC-DEA permeation and retention in the skin were similar after a single and repeated application (4 doses every 12 h). Thus, oleyl alcohol offers several advantages over oleic acid for topical drug delivery.

Skin Barrier Fine Tuning through Low-Temperature Lipid Chain Transition.

The lipids in the mammalian stratum corneum (SC) adopt an unusually rigid arrangement to form a vital barrier preventing water loss and harmful environmental impacts. Just above the physiological temperature, a subset of barrier lipids undergoes a phase transition from a very tight orthorhombic to a looser hexagonal arrangement and vice versa. The purpose of this lipid transition in skin physiology is unknown. Permeability experiments on isolated human SC indicated that the transition affects the activation energy for a model compound that prefers lateral movement along lipid layers but not for water or a large polymer that would cross the SC through the pore pathway. The orthorhombic phase content of SC lipids, as determined by infrared spectroscopy, was also modulated by (de)hydration. Spontaneous rearrangement of human SC lipid monolayers into 10 nm higher multilamellar islets at 32-37 °C but not at room temperature was revealed by atomic force microscopy. Our findings add to our knowledge of fundamental skin physiology suggesting a fine temperature- and hydration-controlled switch from fluid lipids (required for lipid barrier assembly) to rigid and tightly packed lipids in the mature SC (necessary for the water and permeability barriers).

Proline, hydroxyproline, and pyrrolidone carboxylic acid derivatives as highly efficient but reversible transdermal permeation enhancers.

Overcoming the skin barrier properties efficiently, temporarily, and safely for successful transdermal drug delivery remains a challenge. We synthesized three series of potential skin permeation enhancers derived from natural amino acid derivatives proline, 4-hydroxyproline, and pyrrolidone carboxylic acid, which is a component of natural moisturizing factor. Permeation studies using in vitro human skin identified dodecyl prolinates with N-acetyl, propionyl, and butyryl chains (Pro2, Pro3, and Pro4, respectively) as potent enhancers for model drugs theophylline and diclofenac. The proline derivatives were generally more active than 4-hydroxyprolines and pyrrolidone carboxylic acid derivatives. Pro2-4 had acceptable in vitro toxicities on 3T3 fibroblast and HaCaT cell lines with IC50 values in tens of µM. Infrared spectroscopy using the human stratum corneum revealed that these enhancers preferentially interacted with the skin barrier lipids and decreased the overall chain order without causing lipid extraction, while their effects on the stratum corneum protein structures were negligible. The impacts of Pro3 and Pro4 on an in vitro transepidermal water loss and skin electrical impedance were fully reversible. Thus, proline derivatives Pro3 and Pro4 have an advantageous combination of high enhancing potency, low cellular toxicity, and reversible action, which is important for their potential in vivo use as the skin barrier would quickly recover after the drug/enhancer administration is terminated.

Transdermal Permeation and Skin Retention of Diclofenac and Etofenamate/Flufenamic Acid From Over-the-Counter Pain Relief Products.

Topical pain relief products differ in the type of drug, concentration, and formulation. All these factors influence the drug transit through the skin barrier, and its eventual retention in the skin as a reservoir for subsequent release. In addition, the drug potency can be different, which is important for the product efficacy. We studied here ex vivo human skin permeation and retention of five over-the-counter NSAID gels containing 2.32% diclofenac (DIC) and 5-10% etofenamate (ETF). The potency of the permeated/retained drug amounts were compared using a composite parameter, the Index of Relative Topical Anti-inflammatory Activity (IRTAA), which is calculated as the product of the skin permeation/retention and the drug relative potency. The IRTAAs of the DIC gel were 94-667-fold higher and 72-208-fold higher for transdermal delivery and skin retention, respectively, than IRTAAs of the ETF gels. These superior IRTAAs indicate that DIC delivered by this topical formulation would achieve a higher bioactivity and would form a potent drug reservoir relevant for its subsequent long-lasting release.

Permeation enhancers in transdermal drug delivery: benefits and limitations.

Introduction: Transdermal drug delivery has several clinical benefits over conventional routes of drug administration. To open the transdermal route for a wider range of drugs, including macromolecules, numerous physical and chemical techniques to overcome the natural low skin permeability have been developed.Areas covered: This review focuses on permeation enhancers (penetration enhancers, percutaneous absorption promoters or accelerants), which are chemicals that increase drug flux through the skin barrier. First, skin components, drug permeation pathways, and drug properties are introduced. Next, we discuss properties of enhancers, their various classifications, structure-activity relationships, mechanisms of action, reversibility and toxicity, biodegradable enhancers, and synergistic enhancer combinations.Expert opinion: Overcoming the remarkable skin barrier properties in an efficient, temporary and safe manner remains a challenge. High permeation-enhancing potency has long been perceived to be associated with toxicity and irritation potential of such compounds, which has limited their further development. In addition, the complexity of enhancer interactions with skin, formulation and drug, along with their vast chemical diversity hampered understanding of their mechanisms of action. The recent development in the field revealed highly potent yet safe enhancers or enhancer combinations, which suggest that enhancer-aided transdermal drug delivery has yet to reach its full potential.

Esters of terpene alcohols as highly potent, reversible, and low toxic skin penetration enhancers.

Skin penetration/permeation enhancers are compounds that improve (trans)dermal drug delivery. We designed hybrid terpene-amino acid enhancers by conjugating natural terpenes (citronellol, geraniol, nerol, farnesol, linalool, perillyl alcohol, menthol, borneol, carveol) or cinnamyl alcohol with 6-(dimethylamino)hexanoic acid through a biodegradable ester linker. The compounds were screened for their ability to increase the delivery of theophylline and hydrocortisone through and into human skin ex vivo. The citronellyl, bornyl and cinnamyl esters showed exceptional permeation-enhancing properties (enhancement ratios up to 82) while having low cellular toxicities. The barrier function of enhancer-treated skin (assessed by transepidermal water loss and electrical impedance) recovered within 24 h. Infrared spectroscopy suggested that these esters fluidized the stratum corneum lipids. Furthermore, the citronellyl ester increased the epidermal concentration of topically applied cidofovir, which is a potent antiviral and anticancer drug, by 15-fold. In conclusion, citronellyl 6-(dimethylamino)hexanoate is an outstanding enhancer with an advantageous combination of properties, which may improve the delivery of drugs that have a limited ability to cross biological barriers.

Fluorescent Penetration Enhancers Reveal Complex Interactions among the Enhancer, Drug, Solvent, and Skin.

Skin penetration/permeation enhancers facilitate drug delivery through the skin barrier. However, the specific mechanisms that govern the enhancer interactions with the skin, drug, and donor solvent are not fully understood. We designed and synthesized fluorescent-labeled enhancers by attaching 7-nitrobenzo[c][1,2,5]oxadiazol-4-yl (NBD) groups to 6-aminohexanoic acid esters. These NBD esters (applied at a 1% concentration) enhanced the permeation of the model drugs theophylline and hydrocortisone through human skin in vitro up to 6.6- and 3.9-times, respectively. The enhancement effects were strongly affected by the ester chain length (C8-C12) and the polarity of the donor solvent. Using high-performance liquid chromatography with fluorescence detection, no NBD esters were detected in the acceptor buffer, but their hydrolysis product, NBD acid, was detected, whereas both acid and esters were found in the skin. The enhancer hydrolysis occurred in the lower stratum corneum and epidermis; more hydrophilic NBD acid, which is an inactive enhancer, penetrated deeper. This illustrates the principle of biodegradable enhancers. The enhancer concentrations in the skin depended not only on the enhancer chain length and the donor solvent, but also on the drug used. Thus, the drug, when coapplied with the enhancer, modulates the enhancer penetration into the skin and, consequently, its effect. Finally, active (NBD-C8 ester) and inactive (NBD acid) enhancers were visualized in human skin by confocal laser scanning microscopy. Both compounds were found mostly in the stratum corneum intercellular spaces, suggesting that although both are located within the skin barrier lipids, only the active ester is able to effectively interact with the lipids, which was proved by infrared spectroscopy of enhancer-treated stratum corneum. This proof-of-concept study illustrates the use of fluorescent enhancers to obtain insight into the skin penetration/permeation process; interactions among the enhancer, drug, solvent, and skin; and enhancer metabolism.

Permeability and microstructure of cholesterol-depleted skin lipid membranes and human stratum corneum.

Cholesterol (Chol) is one of the major skin barrier lipids. The physiological level of Chol in the stratum corneum (SC) appears to exceed its miscibility with other barrier lipids, as some Chol is phase separated. Chol synthesis is essential for epidermal homeostasis, yet the role of these Chol domains in SC permeability is unknown. We investigated the impact of Chol depletion on the permeability properties and microstructure of model membranes and human SC. X-ray powder diffraction of membranes constructed from isolated human skin ceramides or synthetic ceramides confirmed that only approximately half of the normal Chol amount can be incorporated in either long or short periodicity lamellar phases. The long periodicity lipid arrangement persisted even in the absence of Chol. Infrared spectroscopy suggested that Chol had negligible effects on the lipid chain order and packing at physiological skin temperature. Chol depletion of the model membranes or isolated human SC did not compromise the barrier function to water and two model permeants. On the contrary, the membrane with the Chol content reduced to 40% of the normal value, where no separated Chol was observed, was significantly less permeable than the control. Thus, a 0.4:1:1 M ratio of Chol/ceramides/fatty acids appears sufficient for skin lipids to limit water loss and prevent the entry of environmental substances. We speculate that the SC Chol domains may have roles in the skin other than barrier function.

Galactosyl Pentadecene Reversibly Enhances Transdermal and Topical Drug Delivery.

PURPOSE: To study new skin penetration/permeation enhancers based on amphiphilic galactose derivatives. METHODS: Two series of alkyl and alkenyl galactosides were synthesized and evaluated for their enhancing effect on transdermal/topical delivery of theophylline (TH), hydrocortisone (HC) and cidofovir (CDV), reversibility of their effects on transepidermal water loss (TEWL) and skin impedance, interaction with the stratum corneum using infrared spectroscopy, and cytotoxicity on keratinocytes and fibroblasts. RESULTS: Initial evaluation identified 1-(α-D-galactopyranosyl)-(2E)-pentadec-2-ene A15 as a highly potent enhancer - it increased TH and HC flux through human skin 8.5 and 5 times, respectively. Compound A15 increased the epidermal concentration of a potent antiviral CDV 7 times over that reached by control and Span 20 (an established sugar-based enhancer). Infrared spectroscopy of human stratum corneum indicated interaction of A15 with skin barrier lipids but not proteins. These effects of A15 on the skin barrier were reversible (both TEWL and skin impedance returned to baseline values within 24 h after A15 had been removed from skin). In vitro toxicity of A15 on HaCaT keratinocytes and 3T3 fibroblasts was acceptable, with IC50 values over 60 µM. CONCLUSIONS: Galactosyl pentadecene A15 is a potent enhancer with low toxicity and reversible action.

SAMPA: A free software tool for skin and membrane permeation data analysis.

Skin and membrane permeation experiments comprise an important step in the development of a transdermal or topical formulation or toxicological risk assessment. The standard method for analyzing these data relies on the linear part of a permeation profile. However, it is difficult to objectively determine when the profile becomes linear, or the experiment duration may be insufficient to reach a maximum or steady state. Here, we present a software tool for Skin And Membrane Permeation data Analysis, SAMPA, that is easy to use and overcomes several of these difficulties. The SAMPA method and software have been validated on in vitro and in vivo permeation data on human, pig and rat skin and model stratum corneum lipid membranes using compounds that range from highly lipophilic polycyclic aromatic hydrocarbons to highly hydrophilic antiviral drug, with and without two permeation enhancers. The SAMPA performance was compared with the standard method using a linear part of the permeation profile and a complex mathematical model. SAMPA is a user-friendly, open-source software tool for analyzing the data obtained from skin and membrane permeation experiments. It runs on a Microsoft Windows platform and is freely available as a Supporting file to this article.

Dodecyl Amino Glucoside Enhances Transdermal and Topical Drug Delivery via Reversible Interaction with Skin Barrier Lipids.

PURPOSE: Skin permeation/penetration enhancers are substances that enable drug delivery through or into the skin. METHODS: To search for new enhancers with high but reversible activity and acceptable toxicity, we synthesized a series of D-glucose derivatives, both hydrophilic and amphiphilic. RESULTS: Initial evaluation of the ability of these sugar derivatives to increase permeation and penetration of theophylline through/into human skin compared with a control (no enhancer) or sorbitan monolaurate (Span 20; positive control) revealed dodecyl 6-amino-6-deoxy-α-D-glucopyranoside 5 as a promising enhancer. Furthermore, this amino sugar 5 increased epidermal concentration of a highly hydrophilic antiviral cidofovir by a factor of 7. The effect of compound 5 on skin electrical impedance suggested its direct interaction with the skin barrier. Infrared spectroscopy of isolated stratum corneum revealed no effect of enhancer 5 on the stratum corneum proteins but an overall decrease in the lipid chain order. The enhancer showed acceptable toxicity on HaCaT keratinocyte and 3T3 fibroblast cell lines. Finally, transepidermal water loss returned to baseline values after enhancer 5 had been removed from the skin. CONCLUSIONS: Compound 5, a dodecyl amino glucoside, is a promising enhancer that acts through a reversible interaction with the stratum corneum lipids.

Transdermal delivery and cutaneous targeting of antivirals using a penetration enhancer and lysolipid prodrugs.

PURPOSE: In this work, we investigate prodrug and enhancer approaches for transdermal and topical delivery of antiviral drugs belonging to the 2,6-diaminopurine acyclic nucleoside phosphonate (ANP) group. Our question was whether we can differentiate between transdermal and topical delivery, i.e., to control the delivery of a given drug towards either systemic absorption or retention in the skin. METHODS: The in vitro transdermal delivery and skin concentrations of seven antivirals, including (R)- and (S)-9-[2-(phosphonomethoxy)propyl]-2,6-diaminopurine (PMPDAP), (S)-9-[3-hydroxy-2-(phosphonomethoxy)propyl]-2,6-diaminopurine ((S)-HPMPDAP), its 8-aza analog, and their cyclic and hexadecyloxypropyl (HDP) prodrugs, was investigated with and without the penetration enhancer dodecyl-6-(dimethylamino)hexanoate (DDAK) using human skin. RESULTS: The ability of ANPs to cross the human skin barrier was very low (0.5-1.4 nmol/cm(2)/h), and the majority of the compounds were found in the stratum corneum, the uppermost skin layer. The combination of antivirals and the penetration enhancer DDAK proved to be a viable approach for transdermal delivery, especially in case of (R)-PMPDAP, an anti-HIV effective drug (30.2 ± 2.3 nmol/cm(2)/h). On the other hand, lysophospholipid-like HDP prodrugs, e.g., HDP-(S)-HPMPDAP, reached high concentrations in viable epidermis without significant systemic absorption. CONCLUSIONS: By using penetration enhancers or lysolipid prodrugs, it is possible to effectively target systemic diseases by the transdermal route or to target cutaneous pathologies by topical delivery.