The Tape-Stripping Technique as a Method for Drug Quantification in Skin

Quantification of drugs within the skin is essential for topical and transdermal delivery research. Over the last two decades, horizontal sectioning, consisting of tape stripping throughout the stratum corneum, has become one of the traditional investigative techniques. Tape stripping of human stratum corneum is widely used as a method for studying the kinetics and penetration depth of drugs. This paper shows the applications of the tape stripping technique to quantify drug penetration through the skin, underlining its versatile application in the area of topical and transdermal drugs.


INTRODUCTION
Tape stripping with adhesive tape is a widely accepted and used method to examine the localization and distribution of substances within the stratum corneum (SC) [1][2][3][4][5][6][7].This is a minimally invasive technique to sequentially remove SC by the repeated application of appropriate adhesive tapes [8].This technique can be used to investigate SC cohesion in vivo by quantifying the amount of SC removed [9].Today, weighing with precision balances is the most frequently used method to determine the amount of SC removed on a tape strip.The method is also used to provide information about the kinetics of transdermal drug delivery, offering an apparently easy and quite non-invasive methodology for skin tissue sampling, and is the basis of the FDA's so-called dermatopharmacokinetic (DPK) approach to the assessment of topical bioavailability and bioequivalence [10].However, validation and optimization of the procedure have not come quickly and the proposed guidance document has been withdrawn for re-evaluation.More recent work has addressed at least some of the important limitations of the DPK approach [11-13] and has proposed modifications in order to incorporate it into an improved protocol.
A number of excellent reviews that have been published contain detailed discussions concerning many aspects of the tape stripping technique [14][15][16].The present review shows an updated overview of the use of the tape stripping technique (TS) in the pharmaceutical field, specifically in the area of topical and transdermal drugs.This focus is justified due to the magnitude of the experimental data available with the use of this technique.The use of the tape stripping technique in experimental medicine and pharmaceutical sciences has a long history.

The skin
The skin is the largest organ of the body [17][18][19], accounting for more than 10% of body mass, and the one that enables the body to interact more intimately with its environment.Essentially, the skin consists of four layers: The SC, that is the outer layer of the skin (non-viable epidermis), and forms the rate-controlling barrier for diffusion for almost all compounds.It is composed of dead flattened, keratin-rich cells, the corneocytes.These dense cells are surrounded by a complex mixture of intercellular lipids, namely, ceramides, free fatty acids, cholesterol, and cholesterol sulphate.Their most important feature is that they are structured as ordered bilayer arrays [20].The predominant diffusional path for a molecule crossing the SC appears to be intercellular [21][22][23].The other layers are: the remaining layers of the epidermis (viable epidermis), the dermis, and the subcutaneous tissues (Figure 1).There are also several associated appendages: hair follicles, sweat ducts, apocrine glands and nails.In a general context, the skin's functions may be classified as protective, homeostasis maintaining functions, or sensing [25].
The skin became popular as a potential site for systemic drug delivery, on the one hand, because of the possibility of avoiding the problems of stomach emptying, pH effects, enzyme deactivation associated with gastrointestinal passage, and hepatic first-pass metabolism; and on the other hand, due to its capability to enable input control.

Tape stripping technique overview
The simplest method for reducing the barrier imposed by the SC is to remove it.Theoretically, an adhesive tape removes a layer of corneocytes.In vivo, removal of the SC by TS is performed by the repeated application of adhesive tapes to the skin's surface.In Figure 3 we can observe a detailed procedure of the tape stripping technique.It has been found that on the flexor surface of the forearm, about 30 tape strips are needed to strip off most of the horny layer [8].Multiple strips remove a substantial skin barrier, as evidenced by 20 to 25-fold increases in transepidermal water loss (TEWL) [38].Usually, the amount of SC removed by TS is not linearly proportional to the number of tapes removed [8].TS appears to be simple and easy to perform [39-40], however there are different parameters that can influence the quantity of SC removed by a piece of tape, and these include TS mode [38,41], skin hydration, cohesion between cells (which increases with SC's depth), the body site and inter-individual differences [9,42].The impact of these factors has been frequently investigated [8,[38][39][40][41][42][43][44].
After its description by Pinkus [8], TS has become a standard method in dermatological research [45].
This method can be used to obtain a more susceptible skin, e.g., prior to the application of an irritant [46] or an allergen [47][48][49].Similarly, TS is performed to induce a defined disruption of the water barrier, e.g., to evaluate the effect of a subsequently applied skin care product in barrier restoration [50].It may be also used to obtain cells for mycological culture [51,52] or to investigate SC quality [53].In dermatopharmacology, the SC barrier function [50,54] and the bioavailability and bioequivalence of topical drugs [39, [55][56][57] can be evaluated with the use of this technique [58,59].
Tape stripping appears to be simple and easy to perform.However, there are parameters which have to be defined, as they may change the outcome.Because various brands of tape differ in shape, surface area, composition and adhesive properties, the influence of the tape brand on the outcome seems apparent [38,41].Other parameters which influence the procedure can be subsumed in the intrinsic properties of the SC [38].Although these properties are often investigated, little is known about the anatomical sites (intrinsic factor) as well as the pressure with which the tape is applied on to the skin, the duration of pressure and the removal process (extrinsic factors) influencing SC removal.
In the case of bioequivalence studies, topical bioavailability can be estimated from the drug concentration within the SC, which is expected to be related to the drug concentration at the target site (i.e., usually viable epidermis or dermis) since the SC is the rate limiting barrier for percutaneous absorption.Similarly to the determination of the drug concentration in blood and/or urine as surrogate for the concentration at the target tissue, the determination of the drug concentration in the SC may serve as a surrogate for the concentration in the viable (epi-)dermis [58].A typical profile obtained from a skin permeation study with sodium naproxen is shown in figure 4 [7].TS, which enables the removal of the SC layer by layer, is a useful DPK technique for the assessment of drug amounts in SC as a function of time [59].

Applications of the Tape-stripping technique
Removal by TS of the outermost skin layer, the SC, has become a common practice in recent years [60,61].The determination of the kinetics and penetration depth of different kind of permeants by tracing the concentration profiles in SC, has been facilitated by the use of the virtually non-invasive method of SC stripping with adhesive tape [1, 2,5,43,44].For this reason, TS also offers the possibility of evaluating bioequivalence of topical dermatological dosage forms [5].DPK characterization of active drugs in human volunteers has been suggested to be able to replace comparative clinical trials as a means of documenting bioequivalence [59].Moreover, in vitro methods are encouraged by regulatory agencies regarding the provision of percutaneous absorption data for drugs, pesticides and cosmetics [62].All these points are emphasized in Table 1, which summarizes the research with the TS technique to determine the kinetics and penetration depth of permeants (drugs and toxic chemicals) [7,26,35,37,44,, in order to evaluate the factors that influence the physiology of the SC [42,44,105,106], to determine the composition of the SC [108], superficial infections in the skin [110,111], and evaluate skin regeneration [112,113], etc.

Analgesic and anti-inflammatory drugs
Arima et al. [63] investigated the effect of hydroxypropyl-P-cyclodextrin (HP-P-CD) on the cutaneous penetration and activation of ethyl 4biphenylyl acetate (EBA), a prodrug of the nonsteroidal anti-inflammatory drug 4biphenylylacetic acid (BPAA), from hydrophilic ointment, using hairless mouse skin in vitro.When the hydrophilic ointment containing a complex of EBA with HP-P-CD was applied to full-thickness skin, HP-P-CD facilitated the penetration of EBA into the skin.When the ointment containing the EBA:HP-P-CD complex was applied to the skin, the BPAA flux through the tape-stripped skin was greater than that through full-thickness skin, while the activation of the prodrug in the skin was slowed by TS.Their results suggest that the enhancing effect of HP-P-CD on the cutaneous penetration of EBA would be largely attributed to an increase in the effective concentration of EBA in the ointment.
Curdy et al. [35] administered piroxicam from a commercially available gel to human volunteers, both passively and under the application of an iontophoretic current.After treatment, the SC at the site of application was progressively tape-stripped and piroxicam transport into the membrane was assessed by UVanalysis of drug extracted from the tape-strips.Current application enhanced drug uptake into the SC, as indicated by both increased piroxicam concentrations in the horny layer and detectable concentrations at greater depths in the membrane.The total amount of drug recovered in the SC post-iontophoresis was significantly higher than that found following passive diffusion for each application time.
[7] determined the penetration of sodium naproxen, formulated in Pluronic F-127 gels containing Azone ® and Transcutol as penetration enhancers, through human skin in vivo by using the TS technique.It was found that the combination of Azone ® and Transcutol in PF-127 gels enhanced sodium naproxen penetration, with up to two-fold enhancement ratios compared with the formulation containing Transcutol only.These results were confirmed by TEWL and ATR-FTIR spectroscopy, suggesting a synergistic action for Azone ® and Transcutol ® .
Esposito et al. [64] produced and characterized monoleine (MO) dispersions as drug delivery systems for indomethacin.An in vitro diffusion study was conducted using Franz cells associated to SC epidermal membrane on cubosome dispersions viscosized by carbomer.In vivo studies based on skin reflectance spectrophotometry and TS were performed to better investigate the performance of cubosome as an indomethacin delivery system.
Indomethacin incorporated in viscosized MO dispersions exhibited a lower flux with respect to the analogous formulation containing the free drug in the aqueous phase and to the control formulation based on carbomer gel.Reflectance spectroscopy demonstrated that indomethacin incorporated into MO dispersions can be released in a prolonged fashion.TS experiments corroborated this finding.MO dispersions can be proposed as nanoparticulate systems able to control the percutaneous absorption of indomethacin Ganem-Quintanar et al.
[65] used naproxen-loaded nanoparticles to prepare, in a one-step process, unilaminar films of Eudragit E-100.Nanoparticle films and conventional films were characterized in vitro by drug release studies through a cellulose membrane using Franz-type cells, and in vivo by penetration experiments with the TS technique.Concerning in vivo penetration studies, no statistical differences were found for the amount of naproxen penetrated across the SC and the depth of penetration for the two films.
Herkenne et al. [66] investigated pig ear skin as a surrogate for human skin in the assessment of topical drug bioavailability by sequential TS of the SC.
Ex vivo experiments on isolated pig ears were compared with in vivo studies in human volunteers.Four formulations including ibuprofen in different propylene glycol (PG)-water mixtures (25:75, 50:50, 75:25 and 100:0), were compared.Derived DPK parameters characterizing the diffusion and partitioning of the drug in the SC ex vivo were consistent with those in vivo following a 30-minute application period.Furthermore, nonsteady-state ex vivo results could be used to predict the in vivo concentration profile of the drug across the SC when a formulation was administered for 3 h (i.e., close to steady-state).Taken together, the results obtained suggest that pig ear skin ex vivo is promising as a tool for topical formulation evaluation and optimization.
Hostýnek et al. [67] sheded light on the long-standing controversy on whether wearing copper bangles benefits patients suffering from inflammatory conditions such as arthritis.Sequential TS was performed on healthy volunteers to examine the diffusion of copper through human SC in vivo, following application of the metal as powder on the volar forearm for periods of up to 72 h.Exposure sites were stripped 20 times, and the strips were analyzed for metal content by inductively coupled plasmamass spectroscopy.
The results indicate that, in contact with skin, copper will oxidize and may penetrate the SC after forming an ion pair with skin exudates.The rate of reaction seems to depend on contact time and oxygen availability.A marked interindividual difference was observed in baseline values and amounts of copper absorbed.
Lodén et al. [68] compared the bioavailability of ketoprofen in a photostabilised gel formulation without photoprotection using a new DPK tape stripping model and an established ex vivo penetration method using human skin.Analyses of the SC showed that during the first 45 minutes, about 12 μg/cm 2 of ketoprofen were absorbed into the skin from the formulations.The area under the ketoprofen concentration-time curve (AUC 0-6 h ) for the photo-stabilised gel/transparent gel ratio was 73%.The rate of penetration of ketoprofen through isolated skin was approximately 0.2 μg/cm 2 h for both formulations.The ratio's AUC 0-36 h was 84%.Thus, the two methods did not disagree in terms of the relative efficacy of the two gels.The comparison of the amount of ketoprofen in the skin after 45 min with the amount penetrated through the excised skin during 36 h, suggests a change in the thermodynamic activity of ketoprofen during exposure.A supersaturated formulation may have been formed initially due to evaporation of ethanol.
Wagner Steffen et al. [69] studied the penetration kinetics of sesquiterpene lactones (SLs) in Arnica montana preparations; a stripping method with adhesive tape and pig skin as a model was used.For the determination of SLs in the stripped layers of the SC, a gas chromatography/mass spectrometry method was developed and validated.
The penetration behavior of one gel preparation and two ointment preparations was investigated.The SLs of all preparations showed a comparable penetration and permeation through the SC, in the uppermost layer of the skin.Interestingly, the gel preparation showed a decreased penetration rate over 4 h, whereas the penetration rate of ointments remained constant over time.Moreover, they could demonstrate that the total amount of SLs penetrated depends only on the kind of formulation and the SLs-content in the formulation, but not on SLs composition or the extraction agent used.
Wagner Heike et al. [70] carried out penetration experiments to investigate several incubation times with three different skin flaps using the Saarbruecken penetration model and the lipophilic model drug flufenamic acid.Drug distribution within SC was obtained by the TS technique, while the drug present in deep skin layers was determined by cryosectioning.In addition, for the lipophilic drug flufenamic acid, a direct linear correlation was found between SC/water partition coefficients and the drug amounts penetrated into the SC for all the time intervals tested.The authors concluded that SC/water-partition coefficients offer the possibility to predict drug amounts within the SC of different donor skin flaps, without a timeconsuming determination of the lipid composition of the SC.

Corticosteroids
The aim of Pellanda et al. [71] was to investigate the effect of i) dose and ii) application frequency on the penetration of triamcinolone acetonide (TACA) into human SC in vivo.The experiments were conducted on the forearms of 15 healthy volunteers, with i), single TACA doses (300 μg/cm 2 and 100 μg/cm 2 ), and ii) single (1 x 300 μg/cm 2 ) and multiple (3 x 100 μg/cm 2 ) TACA doses.SC samples were collected by TS after 0.5, 4 and 24 h (i) and after 4, 8 and 24 h (ii).In Experiment 1, TACA amounts within SC after application of 1 x 300 μg/cm 2 compared to 1 x 100 μg/cm 2 were only significantly different immediately after application, and were similar at 4 and 24 h.In ii), multiple applications of 3 x 100 μg/cm 2 yielded higher TACA amounts compared to a single application of 1 x 300 μg/cm 2 at 4 and 8 h.At 24 h, no difference was observed.In conclusion, by using this simple vehicle, considerable TACA amounts were retained within the SC, independently of dose and application frequency.

Disinfectants
Lboutounne et al. [72] investigated the sustained bactericidal activity of chlorhexidine base loaded poly(Є-caprolactone) nanocapsules against Staphylococcus epidermidis inoculated onto porcine ear skin.The antimicrobial activity of these colloidal carriers was evaluated (i) in vitro against eight strains of bacteria, and (ii) ex vivo against Staphylococcus epidermidis inoculated for 12 h onto porcine ear skin surface treated for 3 min either with 0.6% chlorhexidine base loaded or unloaded nanocapsules suspended in hydrogel, or 1% chlorhexidine digluconate aqueous solution.Chlorhexidine absorption into the SC was evaluated by the TS technique.The results showed that chlorhexidine nanocapsules in aqueous suspension with a 200-300 nm size and a positive charge exhibited similar minimum inhibitory concentrations against several bacteria, compared with chlorhexidine digluconate aqueous solution.Ex vivo, there was a significant reduction in the number of colony forming units from skin treated with chlorhexidine nanocapsules suspension for 3 min compared to chlorhexidine digluconate solution after an 8-h artificial contamination.Interestingly, nanocapsules were present in porcine hair follicles.Topical application of chlorhexidine base-loaded positively charged nanocapsules in an aqueous gel achieved a sustained release of bactericide against Staphylococcus epidermidis for at least 8 h.

Drugs for keratinization disorders
Fresno-Contreras et al. [73] designed an all-trans retinoic acid (RA) topical release system that modifies drug diffusion parameters in the vehicle and the skin in order to reduce systemic absorption and side-effects associated with the topical application of the drug to the skin.RA, either in free form or encapsulated in SC lipid liposomes, was included in hydrogels prepared with Carbopol ® UltrezTM 10 and hyaluronic acid.In vitro permeability experiments with [ 3 H]t-RA were carried out using a Franz-type diffusion cell in abdominal rat skin samples.Accumulation of the drug in the surface and skin layers was evaluated by both the TS technique and a dissection technique.The results show that RA encapsulation not only prolongs drug release, but also promotes drug retention in viable skin.At the same time, interaction between RA and hyaluronic acid has an obstructive effect on diffusion, which contributes to the formation of a reservoir.

Anesthetics
Padula et al. [74] studied the behavior of a skin bioadhesive film containing lidocaine, in vitro and in vivo.Film characterization included in vitro and in vivo drug transport studies with and without iontophoresis.The release rate was compared with a lidocaine commercial gel.The permeation kinetics across the skin was not linear, but the patch acted as a matrix controlling drug delivery.Additionally, permeation rate increased with drug loading.The in vivo experiments with TS indicated that the presence of water during film application is essential to achieve not only the proper adhesion, but also an effective accumulation.The application of an electric current to the patch can further increase the amount of drug accumulated in the SC.

Keratolytics
Bashir et al. [75] studied the keratolytic efficacy of topical preparations containing salicylic acid (SA) in humans by the TS technique, quantifying SC removal by protein analysis.In combination with TS, squamometry was used to evaluate the influence of SA on skin surface scaliness and desquamation.
Furthermore, skin barrier perturbation and skin irritability were recorded and related to the dermatopharmacological effect of the preparations.In contrast to squamometry, TS combined with protein analysis was sensitive in detecting the keratolytic effect of SA within hours of application.Importantly, whereas the pH of the preparations had only a minimal influence on efficacy, local dermatotoxicity was significantly increased at an acidic pH.
This indicates that the intent to increase the amount of free, non-dissociated SA is, in fact, counterproductive, as more acidic preparations resulted in skin irritation and barrier disruption.

Abdulmajed et al. [76] used a novel synthetic
technique to synthesize the co-drug retinyl ascorbate (RA-AsA) ester from all-trans-retinyl chloride (RA) and l-ascorbic acid (AsA) suspended in ethanol at low temperature.The flux and permeation coefficient were determined using heat separated human skin membrane, and skin penetration was determined by TS using fullthickness human skin.All experiments were performed in parallel with retinyl palmitate and ascorbyl palmitate.Overall, the data suggest the potential value of RA-AsA co-drug for treating damage to skin resulting from UV-induced production of free radicals.

Aquaporine-3
Hara et al. [77] showed that glycerol replacement corrects each of the defects in aquaporine-3 (AQP3)-null mice.SC water content, measured by skin conductance and 3 H 2 O accumulation, was 3-fold lower in AQP3-null vs. wild-type mice, but was similar after topical or systemic administration of glycerol in amounts that normalized glycerol content in the SC.Orally administered glycerol fully corrected reduced skin elasticity in AQP3-null mice, as measured by the kinetics of skin displacement after suction, and the delayed barrier recovery, as measured by TEWL after TS.The analysis of [ 14 C]glycerol kinetics indicated a reduced blood-to-SC transport of glycerol in AQP3-null mice, resulting in slowed lipid biosynthesis.These data provide functional evidence for a physiological role of glycerol transport by aquaglyceroporin, and indicate that glycerol is a major determinant of SC water retention and of mechanical and biosynthetic functions.Their findings establish a scientific basis for the >200 year old empirical practice of including glycerol in cosmetic and medicinal skin formulations.

Antiviral drugs
Morgan et al. [78] measured the contribution of SC barrier and microvascular perfusion in determining dermal tissue levels of two hydrophilic drugs (aciclovir and penciclovir) in vivo.Removal of the SC by TS resulted in a 1300-fold increase in penciclovir absorption and a 440-fold increase in aciclovir absorption, confirming that SC is the major barrier to hydrophilic drug absorption.

Anti-Varicella Zoster virus nucleoside
Jarvis et al. [79] determined the in-vitro dermal delivery of a new class of lipophilic, highly potent and uniquely selective anti-Varicella Zoster virus nucleoside (VZV) analogue compared with aciclovir.Three test compounds (Cf1698, Cf1743, and Cf1712) and acyclovir were formulated in propylene glycol/aqueous cream, and finite doses were applied to full-thickness pig ear skin for 48 hours in vertical Franz-type diffusion cells.Depth profiles were constructed following TS and membrane separation.All three test compounds reached the target basal epidermis in concentrations suggesting they would be highly efficacious in reducing viral load.Furthermore, the data showed that each of the test compounds would have a far superior performance than aciclovir.The dermatomal site of viral replication during secondary infection -the basal epidermis-was successfully targeted. .

Vaccines
The skin-associated lymphoid tissue, formed by powerful antigen-presenting cells (APCs), such as Langerhans cells (LCs), dermal dendritic cells (DCs), re-circulating T cells, and regional LNs, ensures the efficient presentation of antigen to immunocompetent cells and the induction of strong immune responses.LCs and dermal DCs commonly exist in the skin and are easy to target [80].The TS technique has been used to study the effect of oligodeoxynucleotides on the immune response [81] and expression of immune receptors [82].
Inoue et al. [81] examined the effect of CpG-oligodeoxynucleotide (CpG-ODN) on the immune response to an antigen applied to tapestripped mouse skin, by evaluating the production of cytokines and Ig isotypes.Confocal laser scanning microscopy revealed that the OVA (model antigen) and CpG-ODN easily penetrated the tape-stripped skin.Co-administration of CpG-ODN and OVA to the disrupted skin elicited an antigen-specific, Th1-predominant immune response, and enhanced the production of Th1type cytokines, IL-12 and IFN-γ.On the other hand, the production of a Th2-type cytokine, IL-4, was drastically suppressed.In terms of antigenspecific antibody production, the IgG2a level, which is regulated by IFN-γ, was increased by CpG-ODN, but IgE production regulated by IL-4 was suppressed.Furthermore, the administration of CpG-ODN through the skin drastically attenuated the production of IgE in mice experiencing IgE-type immune response.Administration of CpG-ODN through the skin may shift the immune response from a Th2 to a Th1-like response.
Continuing with their studies, Inoue et al.
[82] also demonstrated that TS induces the expression of toll-like receptor (TLR)-9 in the skin, and enhances the Th1-type immune response triggered by CpG-ODN administered through the tape-stripped skin.TS induces the expression of TLR-9 and tumor necrosis factor (TNF)-α in the skin, and CpG-ODN treatment through the tape-stripped skin enhances the migration of antigen-presenting cells to the draining lymph nodes.On the other hand, TLR-9 mRNA and TNF-α mRNA were not observed in the skin when CpG-ODN was injected intradermally, or in Th1-type immune response.The transdermal application of CpG-ODN with an antigen through the tape-stripped skin is an effective way to induce a Th1-type immune response, and is also a simple, cost-effective and needle-free vaccination system.

Other kind of permeants (Polyethylene glycols, 4-cyanophenol)
Ayala-Bravo et al. [83] investigated the effect of sucrose esters (sucrose oleate and sucrose laureate in water or Transcutol ® , TC) on the SC barrier properties in vivo, and examined the impact of these surfactant-like molecules on the in vivo percutaneous penetration of a model penetrant, 4hydroxybenzonitrile (4-cyanophenol, 4CP).The effect of the enhancers on 4CP penetration was monitored in vivo using ATR-FTIR spectroscopy in conjunction with TS of the treated site.A combination of sucrose esters (oleate or laureate) and TC is able to temporarily alter the SC barrier properties, thereby promoting 4CP penetration.
Results from TS experiments can be affected significantly by chemical diffusion into the SC during the time required to apply and remove all of the TSs, t TS (period of time required to completely remove the SC by tape stripping).For this reason, Reddy et al. [84] studied dermal absorption of 4CP in humans using TS experiments to assess the conditions under which diffusion alters TS results.Mathematical models were developed to assess the effects of diffusion on parameter estimation.In an experiment with t TS > t lag (i.e., the lag time for a chemical to cross the SC), the permeability coefficient for 4CP, Psc,v, calculated including t TS , was consistent with the values from the literature.When diffusion during stripping was not included in the model, Psc,v, was 70% smaller.Calculations show that chemical concentrations in TSs can be affected by diffusion during TS, but with t TS < 0.2 t lag and an exposure time > 0.3 t lag , TS concentrations are not significantly affected by t TS .
[85] developed a sensitive method for the determination of polyethylene glycols (PEGs) with different molecular weights (MW) in the human SC obtained by TS.The analysis is based on derivatization with pentafluoropropionic anhydride and gas chromatography-electron capture detection.The method showed to be suitable for studying permeability in normal and impaired skin with respect to MW in the range of 150-600 Da.
In order to obtain more data to assess the barrier function of uninvolved skin in atopic dermatitis (AD) patients, Jakasa et al. [86] determined the percutaneous penetration of PEGs of various molecular sizes in vivo in AD patients and control subjects using TS of the SC.The apparent diffusion coefficient of PEGs through atopic skin was twice as high as through normal skin, and decreased with increasing MW in both groups.The partition coefficient in the skin of AD patients was half of that for normal skin, but as for normal skin, there was no MW dependence.Although atopic skin exhibited an altered barrier with respect to diffusion and partitioning, the permeability coefficients were nearly the same for atopic and normal skin.The results support the assumption of an altered skin barrier in AD patients, even if the skin is visibly unaffected by the disease.
Tsai et al. [87] further investigated the dependence of permeability on MW with different forms of barrier disruption.A series of PEGs with a MW ranging from nearly 300 to over 1000 Da were used to study the effects of TS and sodium dodecyl sulphate (SDS) treatment on MW permeability profiles of mouse skin in vitro.The total penetration of PEG oligomers across control skin and tape-stripped skin and SDS-treated to different degrees of barrier disruption progressively decreased with increasing MW.Penetration enhancement relative to control skin was more prominent with larger molecules.The MW cut-off for skin penetration increased with the degree of barrier disruption, irrespectively of the treatment applied, and was 986 Da (TS) and 766 Da (SDS treatment) at TEWL levels in the range of 10-20 g/m 2 per h, compared with 414 Da for control skin.The results strongly suggest that, regardless of the form of barrier disruption applied, not only higher amounts, but also more varieties of chemicals (larger molecules), may penetrate into the skin in the presence of a compromised barrier compared with normal skin.

UV absorbers
Alvárez-Román et al. [88] determined whether encapsulation of lipophilic compounds in polymeric nanoparticles is able to improve topical delivery to the skin.The penetration of octyl methoxycinnamate (OMC) encapsulated in poly(Є-caprolactone) nanoparticles, into and across porcine ear skin in vitro, was investigated using TS.
Quantification of OMC in the skin using TS demonstrated that nanoparticulate encapsulation produced a 3.4-fold increase in the level of OMC within the SC.Nanoparticulate encapsulation of OMC increased its "availability" within the SC.
[26] prepared polymeric nanocapsules (NCs) containing OMC, and their in vivo distribution profile through the SC was determined by the TS technique.The penetration degree of OMC formulated in NCs was compared with that obtained for a nanoemulsion (NE) and a conventional oil-inwater (o/w) emulsion (EM).In vivo percutaneous penetration, evaluated by the TS technique, demonstrated that NE increased the extent of OMC penetration relative to the penetration achieved by NCs or EM.Likewise, OMC accumulation in the skin was significantly greater with NE than with EM or NCs.
Sarveiya et al. [89] developed a reverse HPLC assay to quantify four common sunscreen agents, namely, 2-hydroxy-4methoxybenzophenone, 2-ethylhexyl-pmethoxycinnamate, octylsalicylate and salicylic acid 3,3,5-trimethcyclohexyl ester in a range of biological matrices.This assay was further applied to study skin penetration and systemic absorption of sunscreen filters after topical application to human volunteers.The assay allows the analysis of sunscreen agents in biological fluids, including bovine serum albumin solution, plasma and urine, and in human epidermis by using the TS technique.The results from the preliminary clinical study demonstrate a significant penetration of all sunscreen agents into the skin.
[90] determined the amount of sunscreen present on the skin of people at the beach.The amounts of sunscreen applied to different body sites were quantitatively determined by TS.The actual amounts of sunscreen applied were compared with the COLIPA (European Cosmetic Toiletry and Perfumery Association) standard.Most volunteers had applied 10% or less of the COLIPA standard amount to all body sites assessed.
Mavon et al. [91] assessed the penetration of titanium dioxide (TiO 2 ) and methylene bis-benzotriazoyl tetramethylbutylphenol (MBBT), included in a broad-spectrum sunscreen formulation, into human skin in vivo, using the TS technique, and in vitro, using a compartmental approach.More than 90% of both sunscreens were recovered in the first 15 tape strippings.In addition, they have shown that the remaining 10% did not penetrate the viable tissue, but was localized in the furrows.Less than 0.1% of MBBT was detected in the receptor medium, and no TiO 2 was detected in the follicles, the viable epidermis or the dermis.Thus, this in vivo and in vitro penetration study showed an absence of TiO 2 penetration into viable skin layers through either transcorneal or transfollicular pathways, and a negligible transcutaneous absorption of MBBT.However, differences in distribution within the SC reinforced the need for a complementary approach, using a minimally invasive in vivo methodology and an in vitro compartmental analysis.This combination represents a welladapted method for testing the safety of topically applied sunscreen formulations in real-life conditions.

Fragances
In-vitro human skin permeation and distribution of geranyl nitrile (GN) were determined by Brian et al. [92] using epidermal membranes, following application in 70% ethanol, under non-occlusive conditions, at maximum in-use concentration (1%).Levels of GN in the epidermis (plus any remaining in SC after TS), filter paper membrane support, and receptor fluid were combined to produce a total absorbed dose value of 4.72±0.32%.The systemic exposure resulting from the use of GN as a fragrance ingredient, under unoccluded conditions, would be low based on the currently reported use levels.

Dyes
Teichmann et al. [93] developed a method to investigate the effectiveness of reservoir closure by different formulations.Patent Blue V in water was used as a model penetrant.Its penetration, with and without barrier cream treatment, was analyzed by TS in combination with UV/VIS spectroscopic measurements.The investigations showed that the SC represents a reservoir for topically applied Patent Blue V in water.Furthermore, the barrier investigations showed that vaseline and bees wax form a 100% barrier on the skin surface.The third barrier cream, containing waxes and surfactant, only partially showed a protective effect against the penetration of Patent Blue V in water.Strong inter-individual differences were observed for this barrier product.In conclusion, it was assumed that the application of barrier creams cannot replace other protective measures, and should be used to inhibit low-grade irritants or in combination with other protectants, or in body areas where other protective measures are not applicable.
Jacobi et al. [94] studied the penetration of highly hydrophilic (Patent blue V) and lipophilic (curcumin) dyes into the skin using pure oil (o) or water (w), and comparing them with an o/w emulsion.The penetration and localization of both dyes were investigated in vivo using TS and microscopy techniques.Differences in the distribution and the localization of both dyes within the SC were observed.These differences depend on the physicochemical properties of both the vehicles and the dyes.The vehicle appears to affect, in particular, the penetration pathways.
As we can observe, there is an ongoing search for the identification of testing methods to optimize topical dosage forms and to assess topical drug bioavailability.While in vitro screening continues to play an important role (and is relatively inexpensive and easy to use), regulatory approval of drug delivery systems to the skin, with few exceptions, requires clinical trials to be performed.For many drugs used topically, the problem remains unsolved, since an easily visualized pharmacodynamic response is not elicited.
As a consequence, various alternative techniques have been considered, of which SC tape-stripping is being given the greatest attention [7,26,35,.While the former is technically more challenging, the potential reward is a drug concentration-time profile in a compartment presumed to be in close communication with the site of action of most dermatological drugs.The latter, in contrast, offers an apparently easy and quite non-invasive methodology for skin tissue sampling, and is the basis of the FDA's so-called DPK.Validation and optimization of the procedure have not come quickly.The goal of the research described here is not only to contribute to further establishing the credibility of the tape stripping technique, but also to demonstrate that useful and relevant measurements can be made on a surrogate, ex vivo skin model.

2) Dermal absorption of toxic or irritant chemicals
The rate and extent of dermal absorption are important in the analysis of risk from dermal exposure to toxic chemicals, and for the development of topically applied drugs, barriers, insect repellents, and cosmetics, and the TS technique has been widely used to determine the penetration of these kind of substances [36,37,[95][96][97].
Mattorano et al. [36] developed and tested a simple, non-invasive dermal sampling technique on 22 human volunteers to estimate acute dermal exposure to jet fuel (JP-8).Two sites on the ventral surface of each forearm were exposed to 25 μl of JP-8, and the SC was sequentially tapestripped using an adhesive tape.The analysis of the first tape strips indicated that JP-8 was rapidly removed from the SC over the 20-min study period.On average, after 5 min of exposure, the first two tape strips removed 69.8% of the applied dose.The amount recovered with two tape strips decreased over time, to a recovery of 0.9% 20 min after exposure.By fitting a mixed-effect linear regression model to the TS data, the authors were able to accurately estimate the amount of JP-8 initially applied.This study indicates that naphthalene has a short retention time in the human SC and that the TS technique, if used within 20 min of initial exposure, can be used to reliably measures the amount of naphthalene initially present in the SC due to a single exposure to jet fuel.
Chao et al.
[37] presented a TS method for the removal and quantification of keratin from the SC for normalization of extracted concentrations of naphthalene (as a marker for jet fuel exposure) from 12 human volunteers before and after exposure to jet fuel (JP-8).Due to the potential for removal of variable amounts of squamous tissue from each tape strip sample, keratin was extracted and quantified using a modified Bradford method.Naphthalene was quantified in the sequential tape strips collected from the skin between 10 and 25 min after a single dose of JP-8 was initially applied.The penetration of jet fuel into the SC was demonstrated by the fact that the average mass of naphthalene recovered by a tape strip decreased with increasing exposure time and subsequent tape strips.The actual concentration of naphthalene (as a marker for JP-8 exposure) per unit of keratin in a tape-strip sample can be determined by using this method, and may prove necessary when measuring occupational exposures under field conditions.
Van der Merwe et al.
[95] described a physiologically based pharmacokinetic model developed to simulate the absorption of organophosphate pesticides, such as parathion, fenthion, and methyl parathion through porcine skin with flow-through cells.Three parameters were optimized based on experimental dermal absorption data, including solvent evaporation rate, diffusivity, and a mass transfer factor.Diffusion cell studies were conducted to validate the model under a variety of conditions, including different dose ranges (6.3-106.9μg/cm 2 for parathion; 0.8-23.6μg/cm 2 for fenthion; 1.6-39.3μg/cm 2 for methyl parathion), different solvents (ethanol, 2-propanol and acetone), different solvent volumes (5-120 μL for ethanol; 20-80 μL for 2-propanol and acetone), occlusion versus open to atmosphere dosing, and corneocyte removal by TS.The study demonstrated the utility of PBPK models for studying dermal absorption.The similarity between the overall shapes of the experimental and model-predicted flux/time curves and the successful simulation of altered system conditions for this series of small, lipophilic compounds, indicated that the absorption processes described in the model successfully simulated important aspects of dermal absorption in flow-through cells.These data have a direct relevance in the assessment of topical organophosphate pesticides' risk.
Jongh et al.
[96] studied whether sodium lauryl sulphate (SLS) penetration rate into the SC is related to an impairment of skin's water barrier function and inflammation.The penetration of SLS into the SC was assessed using a noninvasive TS procedure in 20 volunteers after a 4-h exposure to 1% SLS.Additionally, the effect of a 24-h exposure to 1% SLS on the skin water barrier function was assessed by measuring TEWL.A multiple regression analysis showed that the baseline TEWL, SC thickness and SLS penetration parameters K (SC ⁄water partition coefficient) and D clearly influenced the increase in TEWL after the 24-h irritation test.They found that variation in barrier impairment and inflammation of human skin depends on SLS' penetration rate, which was mainly determined by SC thickness.
Lundgren et al. [97] modified and tested a vacuuming sampler for removing particles from the skin.The sampler was compared with two other skin and surface exposure sampling techniques.These were based on surrogate skin (a patch sampler-adhesive tape on an optical cover glass) and a TS removal procedure.All three samplers measure the mass of dust on the skin.Dust containing starch was deposited onto the skin in a whole-body exposure chamber.Samples were taken from forearms and shoulders and analysed using optical microscopy.With the different sampling techniques, small differences in results were obtained.There was a good agreement between the vacuuming sampler and the TS technique.

3) Evaluation of factors that influence the physiology of the stratum corneum
Many factors are known to influence the physiology of the SC.In this way, increasing age is related to decreased skin thickness [98] and a variation in skin lipids [99] and flora [100].The anatomical region also influences lipid distribution [101, 102], microflora [103] and physical parameters such as TEWL [104].However, there are already a few studies reporting conflicting results for the effect of gender [105] on skin physiology, as well as for the effect of anatomical site, pressure, pressure duration and tape removal rate in skin physiology [42], or the effect of skin transport technology, as iontophoresis, on human skin [106].
Jacobi et al. [105] studied the effect of gender on the physiology of the SC.The physiological parameters: TEWL, pH value, hydration and sebum content were determined on the flexor forearms of 6 female and 6 male volunteers.In addition, SC samples, removed by TS, were studied for amount, spectroscopic properties, protein content, and mass.The skin of women was characterized by a significantly higher pH value (5.6±0.4)than that of men (4.3±0.4).Protein absorption was the only other parameter significantly dependent on gender.Both effects might be caused by differences in human biology, such as hormonal status.Therefore, volunteers' gender should be considered in dermatologic studies.
[43] investigated the influence of the procedures (anatomical site, pressure, pressure duration, tape removal rate) inherent to each stripping protocol on changes in skin physiology.A significant influence of all parameters on TEWL increase, as a function of tape strip number was observed.The fastest increase was demonstrated on the forehead, followed by the back and, lastly, the forearm.Rapid removal produced a protracted increase compared with slow removal.Pressure for 10s induced a faster increase in TEWL than 2s pressure.Likewise, pressure at 330 g cm -2 induced an earlier increase than pressure at 165 g cm -2 .Skin hydration was not influenced by the variables tested.
Van der Molen, et al.
[44] investigated the efficacy of TS in removing complete cell layers from the superficial part of the human SC.A histological section of skin that was tape-stripped 20 times, clearly showed non-stripped skin in the furrows, indicating persistent incomplete TS.Replicates of tape-stripped skin surface demonstrated that even after removing 40 tape strips, furrows were still present.They emphasize that the results from studies using the TS method have to be viewed from the perspective that cells on one tape strip of the SC may come from different layers, depending on the position of the tape strip in relation to furrow slope, and such results should be interpreted with considerable caution.
[106] investigated the local changes in the ultrastructure of human skin after iontophoresis in human skin in vitro and in vivo.Human dermatomed skin was subjected to passive diffusion for 6 hours, followed by nine hours of iontophoresis at 0.5mA/cm 2 .In addition, iontophoresis patches were applied to healthy volunteers for 3.5h with 0.5h of passive delivery followed by 3h of iontophoresis at a current density of 0.25mA/cm 2 .Subsequently, a series of TSs were performed, and were visualized by freeze-fracture transmission electron microscopy.In vitro/in vivo studies suggest that iontophoresis results in the formation of intercellular water pools, and in a weakening of the desmosomal structure only in the upper part of the SC.However, no changes in lipid organization were observed in vitro and in vivo at the 0.5 and 0.25mA/cm 2 current densities, respectively.Therefore, even at relatively high current densities, no drastic changes in the ultrastructure of the SC are observed.As far as structural changes in SC are concerned, iontophoresis is a safe method under the experimental conditions used.Weerheim et al. [107] established a suitable analytical method for the determination of local SC lipid composition.For this purpose, SC samples were collected by sequential stripping with Leukoplex tape in five healthy volunteers.Lipids were extracted with an ethyl acetate: methanol mixture (20:80) and separated by means of HPTLC.The results of this study revealed that the free fatty acid level is higher, and cholesterol and ceramide levels are lower in the uppermost SC layers.Levels remained unchanged in the underlying SC layers.In these layers, the ceramide level was about 60 wt % and free fatty acid and cholesterol levels were about 20 wt % each.Ceramides could be separated into seven different fractions, and the relative amounts of individual ceramide fractions did not significantly change with the SC depth.The method developed allowed to study the differences in the SC lipid profile in healthy and diseased human skin relative to the SC lipid organization and the skin barrier function in vivo.

Alberti et al.
[108] evaluated, using attenuated total reflectance Fourier transform infrared spectroscopy, the SC bioavailability of terbinafine (TBF) following topical treatment with four different formulations, based on a vehicle consisting of 50% ethanol and 50% isopropyl myristate.Three of these formulations included a percutaneous penetration enhancer: either 5% oleic acid, 10% 2-pyrrolidone or 1% urea.The SC concentration profile of TBF was measured by repeated infrared spectroscopic measurements while sequentially stripping off the layers of this barrier membrane with adhesive tape.TEWL measurements were also performed, to permit facile estimation of SC thickness.The SC concentration profiles of TBF were fitted to the appropriate solution of Fick's second law of diffusion.This analysis enabled the efficacies of the different formulations tested to be compared to the non-enhancer control.While it was found that the formulation containing 5% oleic acid significantly enhanced the SC availability of TBF, the other formulations did not improve the apparent drug delivery.Kalia et al. [109] determined whether a structurally heterogeneous biomembrane, human SC, behaved as a homogeneous barrier to water transport.Impedance spectra (IS) of the skin and measurements of the rate of TEWL were recorded sequentially in vivo in human subjects as layers of the SC were progressively removed by the serial application of adhesive tape strips.The lowfrequency impedance of skin was much more significantly affected by TS than the higher frequency values; removal of the outermost SC layer had the largest effect.In contrast, TEWL changed little as the outer SC layers were stripped off, but increased dramatically when 6-8 microns of the tissue had been removed.It follows that the two noninvasive techniques probe SC barrier integrity in somewhat different ways.After SC removal, recovery of barrier function, as assessed by increasing values of the low-frequency impedance, apparently proceeded faster than TEWL decreased to the pre-stripping control.The variation of TEWL as a function of SC removal behaved in a manner entirely consistent with a homogeneous barrier, thereby permitting the apparent SC diffusivity of water to be found.Skin impedance (low frequency) was correlated with the relative concentration of water within the SC, thus providing an in vivo probe for skin hydration.Finally, the SC permeability coefficient to water, as a function of SC thickness, was calculated and correlated with the corresponding values of skin admittance derived from IS.

6) Determination of superficial infections and viruses
Topical infections due to S. aureus and S. pyogenes are clinically relevant and cause a variety of serious symptoms, including toxic shock syndrome and skin lesions [110], that can progress to sepsis and systemic shock if they are left untreated [111].These bacterial species are also the most common causes of impetigo in humans [111].The TS technique has offered the possibility of studying superficial infections on the skin [112], as well as viruses in skin tumors [113].
Kugelberg et al. [112] presented a new animal model for the purpose of studying superficial infections.In this model, an infection is established by disruption of the skin barrier by partial removal of the epidermal layer by TS and subsequent application of the pathogens Staphylococcus aureus and Streptococcus pyogenes.The infection and the infection route were purely topical.Thus, the present model is considered more biologically relevant for the study of superficial skin infections in mice and humans.Established topical antibiotic treatments are shown to be effective.The procedures involved in the model are simple, a feature that increases throughput and reproducibility.This new model should be applicable to the evaluation of novel antimicrobial treatments for superficial infections caused by S. aureus and S. pyogenes.

Forslund et al. [113] investigated 229
immunocompetent patients tested for human papilloma virus (HPV) DNA in swab samples collected on top of skin tumors and in biopsies of the same tumors, obtained after stripping with tape to remove superficial layers.HPV DNA was detected on top of 69% of the lesions, and in 12% of the stripped biopsies.A difference was seen for the four types of tumors studied.Seborrheic keratosis had 79% HPV positivity on top of lesions versus 19% in biopsies; actinic keratosis had 83% HPV positivity on top of lesions versus 11% in biopsies; basal cell carcinoma had 63% on top of lesions versus 8% in biopsies; and squamous cell carcinoma had 58% on top of lesions versus 19% in biopsies.HPV DNA is common in superficial layers of lesions, but is not necessarily present in tumors.

7) Skin regeneration
Malminen et al. [114] investigated the expression of tight junction components during reepithelialization of suction blisters and regeneration of the corneal layer after TS.Suction blisters were induced in eight healthy volunteers, and skin biopsies were taken 4 or 6 days afterwards.The restoration of epidermal barrier function was evaluated by measuring water evaporation (WE) from the wound area.TS was performed on three volunteers to remove the corneal layer.Prior to the biopsies, WE from the blister wounds was markedly elevated compared with normal skin.In the epidermis surrounding the blister, occludin and ZO-1 (membraneassociated guanylate kinase homologue protein family) were expressed in the granular cell layer only.In the hyperproliferative zone adjacent to the border of the blister, the expression of ZO-1 was redistributed into several spinous cell layers, while occludin expression was restricted to the upper epidermis.In the leading edge of migrating keratinocytes, both proteins were expressed solely in the most superficial layer of keratinocytes.Double labelling for ZO-1 and involucrin showed expression of both proteins in the same layers of hyperproliferative keratinocytes, while the expression patterns were clearly different in migrating keratinocytes.Tight junctions of regenerating epidermis may provide a functional barrier prior to regeneration of the corneal layer.Sekkat et al. [115] developed an in vitro model for the developing skin of the premature neonate.Barriers of different levels of efficiency were produced by differential tape-stripping of the SC from the skin of excised porcine ears, and were characterized by measurements of TEWL.In this way, it was possible to express the recorded TEWL as a function of percentage SC thickness (F), generating the following relationship: TEWL = 2.7+41.exp[-0.028.F].These data were then compared to previously published in vivo measurements of TEWL obtained from a population of premature neonates at various postconceptional ages (PCA).The former showed a remarkably parallel relationship to that found in vitro with the porcine skin model, namely TEWL = 3.3+41 exp [-0.026.(PCA-160)].Therefore, it can be suggested that the empirically adjusted PCA (i.e., PCA-160) has a close correlation with the developing thickness of the neonate's SC.Consequently, porcine skin in vitro, tape-stripped to a particular level, can provide a barrier corresponding to a specific degree of neonate maturation, and thus, can serve as a useful tool to explore whether transdermal drug delivery in this unique patient population may be beneficial.
Zhai et al. [116] used an in vivo human model to define the irritation potential of a topical agent after partial removal of the SC by cellophane TS.The tape was applied to and removed approximately 50 times (mean, 50.0 +/-16.7)from each test site on the volar aspect of the forearm.TEWL was measured before and daily for 5 days.The TEWL values at baseline after stripping represented the point of maximal stripping barrier disruption.The barrier disruption and irritation potential were assessed with TEWL measurements.The results showed that the model topical agent had no adverse effect on barrier repair, i.e. did not interfere with TEWL normalization.This model provides a method for the prediction, with exaggerated sensitivity, of chemical irritation and proclivity to enhance or retard water barrier repair.They believe that the model may predict the response of low irritation materials and may be more sensitive than patch testing on normal skin, particularly for products to be used on certain areas, e.g. the face, anus, etc., or even mucous membranes.

Conclusions
The quantification of drugs within the skin is essential for topical and transdermal delivery research.Over the last two decades, horizontal sectioning, consisting of TS, has been the traditional investigative technique.
Many in vivo methods for measuring dermal absorption of chemicals are invasive (e.g., blood sampling) or slow (e.g., urine samples collected for extended periods).TS of the outermost skin layer, the SC, is a fast and relatively non-invasive technique for measuring the rate and extent of dermal absorption.Tape stripping data have been used to estimate permeability coefficients and partition coefficients, SC mass, barrier function, drug reservoir from in vivo dermal exposures, and even to explain the SC physiology.TS has also been proposed as a method for evaluating the bioequivalence of topical dermatological dosage forms.DPK characterization of the penetration of active drugs in human volunteers has been suggested to be able to replace comparative clinical trials as means of documenting bioequivalence.It is suggested that DPK assessment of drug concentrations in the SC is comparable to blood/urine measurements of systemically administered drugs, where the concentration of a drug in the SC is expected to relate to its concentrations in viable tissue.Shortcontact DPK experiments can be used to obtain diffusion and partitioning parameters that may subsequently be able to predict drug penetration into the SC following longer application periods.Although tape stripping is widely used to determine dermal absorption through the SC, several factors can influence the actual technique.Recent reviews on this topic provide updated and additional insights (117,118).The investigation of variations in skin condition (dry versus moist skin, skin defects, etc.) to determine their potential impact on the sampling method is warranted.For these reasons, the tape stripping technique requires further development.

Research Outcome Author (Ref.)-Year
Effect of Azone ® and Transcutol ® on skin permeation of sodium naproxen formulated in PF-127 gels.
The combination of Azone ® and Transcutol ® in PF-127 gels enhanced sodium naproxen penetration, with up to two-fold enhancement ratios compared with the formulation containing Transcutol ® only.
Administration of piroxicam from a commercially available gel to human volunteers, both passively and under the application of an iontophoretic current.
The total amount of drug recovered in the SC post-iontophoresis by TS was significantly higher than that found following passive diffusion for each application time.
The enhancing effect of HP-P-CD on the cutaneous penetration of EBA would be largely attributable to an increase in the effective concentration of EBA in the ointment.Reflectance spectroscopy demonstrated that indomethacin incorporated into MO dispersions can be released in a prolonged fashion.TS experiments corroborated this finding.
In vivo penetration studies showed no statistical differences for the penetrated amount of naproxen across the SC and the depth of penetration for the two films.The films formulated from nanoparticle dispersions were shown to be effective for the transdermal administration of naproxen.

Ganem-Quintanar et al. (65), 2006
Investigation of pig ear skin as a surrogate for human skin in the assessment of topical drug bioavailability by sequential TS of the SC.
Pig ear skin ex vivo is promising as a tool for topical formulation evaluation and optimization.

Herkenne et al. (66), 2006
Examination of the diffusion of copper through human SC in vivo following application of the metal as powder on the volar forearm for periods of up to 72 h.
Copper will oxidize and may penetrate the stratum corneum after forming an ion pair with skin exudates.The rate of reaction seems to depend on contact time and oxygen availability.A marked Design of an all-trans retinoic acid (RA) topical release system that modifies drug diffusion parameters in the vehicle and the skin, in order to reduce the systemic absorption and side-effects associated with the topical application of the drug to the skin.
RA encapsulation not only prolongs drug release, but also promotes drug retention in viable skin.

Fresno-Contreras et al. (73), 2005
Behaviour of a skin bioadhesive film containing lidocaine in vitro and in vivo.
In vivo experiments with TS indicated that the presence of water during film application is essential to achieve not only the proper adhesion, but also an effective accumulation.

Padula et al. (74), 2003
Keratolytic efficacy of topical preparations containing salicylic acid in humans by TS, and quantification of SC removal by protein analysis.TS combined with protein analysis was sensitive in detecting the keratolytic effect of salicylic acid within hours of application.
The data suggest the potential value of RA-AsA co-drug for treating damage to skin resulting from UV-induced production of free radicals.
The findings establish a scientific basis for the >200-yr-old empirical practice of including glycerol in cosmetic and medicinal skin formulations due to its influence on water retention and the mechanical and biosynthetic functions of the SC.The method showed to be suitable for studying permeability in normal and impaired skin with respect to MW in the range of 150-600 Da.Jakasa et al. (85, 86), 2004Jakasa et al. (85, 86), , 2007 Dependence of permeability on molecular weight with different forms of barrier disruption.
Irrespectively of the form of barrier disruption, not only higher amounts, but also more varieties of chemicals (larger molecules) may penetrate into the skin in the presence of a compromised barrier, compared with normal skin.A preliminary clinical study demonstrates a significant penetration of all sunscreen agents into the skin, as well as of oxybenzone and its metabolites across the skin.

Sarveiya et al. (89), 2004
Amount of sunscreen present on the skin of people at the beach.
The best protected areas were the upper arm and décolleté, but even in these areas, most volunteers had applied only 10% of the COLIPA standard amount.

Lademann et al (90), 2004
Penetration of titanium dioxide (TiO 2 ) and methylene bisbenzotriazoyltetramethylbutylphenol (MBBT), included in a broad-spectrum sunscreen formulation, into human skin in vivo, using the TS method, and in vitro, using a compartmental approach.
In vivo and in vitro penetration studies showed an absence of TiO  In vitro model for the developing skin of the premature neonate.
Porcine skin, in vitro, tape-stripped to a particular level, can provide a barrier corresponding to a specific degree of neonate maturation, and thus, can serve as a useful tool to explore whether transdermal drug delivery in this unique patient population may be beneficial.

Sekkat et al. (113), 2002
An in vivo human model was utilized to define the irritation potential of a topical agent after partial removal of the stratum corneum by cellophane tape stripping This model provides a method for the prediction, with exaggerated sensitivity, of chemical irritation and proclivity to enhance or retard water barrier repair.

Figure 2 .
Figure 2. Processes of percutaneous absorption and transdermal delivery.

Figure 3 .Figure 4 .
Figure 3.The tape stripping technique procedure Hostýnek et al.(67), 2006 inter-individual difference was observed in baseline values and the amounts of copper absorbed.Comparison of the bioavailability of ketoprofen in a photostabilised gel formulation without photoprotection using a new dermatopharmacokinetic TS model and an established ex vivo penetration method using human skin.The comparison of the amount of ketoprofen in the skin after 45 min with the amount penetrated through the excised skin during 36 h, suggests a change in the thermodynamic activity of ketoprofen during exposure.Lodén et al. (68), 2004Penetration kinetics of SLs (sesquiterpene lactones) in Arnica montana preparations, by using a stripping method with adhesive tape and pig skin as a model.Gel preparation showed a decrease in penetration rate, whereas the penetration rate of ointments remained constant over time.The total amount of SLs penetrated depends only on the kind of formulation and the SLscontent, but not on SLs composition or on the extraction agent used.Wagner Steffen et al. (69), 2006 Penetration experiments investigating several incubation times with three different skin flaps, using the Saarbruecken penetration model and the lipophilic model drug flufenamic acid.A direct linear correlation was found between the SC/water partition coefficients and the drug amounts penetrated into the SC for all time intervals tested.Wagner Heike et al. (70), 2002 Effect of dose and application frequency on the penetration of triamcinolone acetonide (TACA) into human SC in vivo.Considerable TACA amounts were retained within the SC, independently of dose and application frequency.Pallenda et al. (71), 2006 Sustained bactericidal activity of chlorhexidine base loaded poly(Єcaprolactone) nanocapsules against Staphylococcus epidermidis inoculated onto porcine ear skin.Topical application of chlorhexidine base-loaded positively charged nanocapsules in an aqueous gel achieved a sustained release of bactericide against Staphylococcus epidermidis for at least 8 h.Lboutounne et al. (72), 2002

Forslund
regeneration of the corneal layer after TS.Tight junctions of regenerating epidermis may provide a functional barrier prior to regeneration of the corneal layer.Malminen et al. (112), 2003

Determination of superficial infections and viruses
Kugelberg et al. (110), 2005Immunocompetent patients were tested for human papilloma virus (HPV) DNA in swab samples collected on top of skin tumors and in biopsies of the same tumors, obtained after stripping with tape to remove superficial layers.HPV DNA is common in superficial layers of lesions, but is not necessarily present in tumors.