Franz Cell Research Articles

Glyceryl Tristearate-Based Lipid Microparticles Loaded with the Tattoo Colorant, Acid Red 87: Colorant Retention Capacity in Excised Porcine Skin

Background: With the increasing diffusion of tattooing, the photolability of tattoo inks has become a critical issue, as available data indicated that several tattoo colorants are unstable under sunlight, generating potentially toxic photodegradation products. Therefore, it is desirable to enhance the photostability of coloring agents contained in tattoo inks. Aims: Lipid microparticles (LMs) highly loaded with Acid Red 87 (C.I. 45380), a colorant used in tattoo inks, were evaluated for their effect on the colorant photoinstability. In addition, the capacity of the LMs to retain the incorporated C.I. 45380 colorant after their intradermal administration in excised porcine skin was investigated. Methods: LMs loaded with C.I. 45380 were prepared using glyceryl tristearate as the lipidic material and phosphatidylcholine as the surfactant. Non-encapsulated C.I. 45380 or the colorant-loaded LMs were irradiated with a solar simulator for photodecomposition studies or introduced in the excised porcine skin mounted in Franz diffusion cells for stability evaluation in the dermal tissue. Results and Conclusion: The colorant content of the microparticles was 17.7%, and their size ranged from 25 to 170 μm. The light-induced degradation of C.I. 45380 was significantly decreased by its incorporation in the LMs from 20.2 ± 5.8% to 1.9 ± 2.1%. Moreover, after intradermal injection of free or microencapsulated C.I. 45380 in the excised pig skin, the LMs reduced by 93.7% (from 24.6 to 1.5%) the quantity of the colorant diffused and hence lost in the Franz cell receptor fluid. Hence, the LM carrier efficiently retained the entrapped C.I. 45380 following incubation in the dermal region of the isolated porcine skin, which is in favor of a long-lasting tattoo. Based on these data, the incorporation of C.I. 45380 in the LMs could represent a potentially useful strategy to reduce the photodecomposition of the tattoo colorant and its harmful interactions with the skin tissue.

Low-fluence laser-facilitated platelet-rich plasma permeation for treating MRSA-infected wound and photoaging of the skin

Platelet-rich plasma (PRP) is rich in cytokines and growth factors and is a novel approach for tissue regeneration. It can be used for skin rejuvenation but the large molecular size of the actives limits its topical application. In this study, low-fluence laser-facilitated PRP was delivered to evaluate its effect on absorption through the skin, infection-induced wound, and photoaging. The PRP permeation enhancement was compared for two ablative lasers: fractional (CO2) laser and fully-ablative (Er:YAG) laser. In the Franz cell experiment, pig skin was treated with lasers with superficial ablation followed by the application of recombinant cytokines, growth factors, or PRP. The transport of interferon (IFN)-γ and tumor necrosis factor (TNF)-α was negligible in intact skin and stratum corneum (SC)-stripped skin. Both lasers significantly elevated skin deposition of IFN-γ and TNF-α from PRP, and fully-ablative laser showed a higher penetration enhancement. A similar tendency was found for vascular endothelial growth factor and epidermal growth factor. Er:YAG laser-exposed skin displayed 1.8- and 3.9-fold higher skin deposition of platelet-derived growth factor (PDGF)-BB and transforming growth factor (TGF)-β1 from PRP, respectively. According to the confocal images, both laser interventions led to an extensive and deep distribution of IFN-γ and PDGF-BB in the skin. In the in vivo methicillin-resistant Staphylococcus aureus (MRSA) infection model, CO2 laser- and Er:YAG laser-assisted PRP delivery reduced bacterial load from 1.8 × 106 to 5.9 × 105 and 1.4 × 104 colony-forming units, respectively. The open wound induced by MRSA was closed by the laser-assisted PRP penetration. In the mouse photoaging model, elastin and collagen deposition were fully restored by combined PRP and full-ablative laser but not by PRP alone and PRP combined with fractional laser. Laser-facilitated PRP delivery even with a low fluence setting can be considered a promising strategy for treating some dermatological disorders.

Synthesis and characterization of PVP microneedle patch using metal bioelectrodes for novel drug delivery system

In this study, we prepared a novel microneedle patch using metal bioelectrodes that provide excellent mechanical properties, drug-delivery efficiency, and biocompatibility at the same time. The biocompatible polyvinylpyrrolidone (PVP) based microneedle patch was prepared using gamma rays. The back side of the PVP based microneedle patch were coated with gold or silver as bioelectrodes using the thermal evaporation method to maximize and control the efficiency of drug release. The Au or Ag-coated microneedle patch has enough mechanical properties to successfully penetrate the skin. In addition, it was confirmed using a Franz cell that the amount of drug released from a drug-loaded microneedle patch was maximized by thermal or electrical stimulus. The drug-release efficiency increased more than 7.9 times compared to a flat PVP patch (without applying electricity) when 3 V was applied. In addition, when ~40 °C heat were applied, the drug-release efficiency of the Au- and Ag-coated PVP based microneedle patches improved about 5.3 times compared to a flat PVP patch at room temperature. These excellent properties of our metal-coated PVP based microneedle patches can be used not only in whitening or anti-wrinkling cosmetics, but also as medical drug-delivery systems to treat such as hypertension, diabetes, and arthritis.

The Potential of Caffeic Acid Lipid Nanoparticulate Systems for Skin Application: In Vitro Assays to Assess Delivery and Antioxidant Effect.

The object of this study is a comparison between solid lipid nanoparticles and ethosomes for caffeic acid delivery through the skin. Caffeic acid is a potent antioxidant molecule whose cutaneous administration is hampered by its low solubility and scarce stability. In order to improve its therapeutic potential, caffeic acid has been encapsulated within solid lipid nanoparticles and ethosomes. The effect of lipid matrix has been evaluated on the morphology and size distribution of solid lipid nanoparticles and ethosomes loaded with caffeic acid. Particularly, morphology has been investigated by cryogenic transmission electron microscopy and small angle X-ray scattering, while mean diameters have been evaluated by photon correlation spectroscopy. The antioxidant power has been evaluated by the 2,2-diphenyl-1-picrylhydrazyl methodology. The influence of the type of nanoparticulate system on caffeic acid diffusion has been evaluated by Franz cells associated to the nylon membrane, while to evaluate caffeic acid permeation through the skin, an amperometric study has been conducted, which was based on a porcine skin-covered oxygen electrode. This apparatus allows measuring the O2 concentration changes in the membrane induced by polyphenols and H2O2 reaction in the skin. The antioxidative reactions in the skin induced by caffeic acid administered by solid lipid nanoparticles or ethosomes have been evaluated. Franz cell results indicated that caffeic acid diffusion from ethosomes was 18-fold slower with respect to solid lipid nanoparticles. The amperometric method evidenced the transdermal delivery effect of ethosome, indicating an intense antioxidant activity of caffeic acid and a very low response in the case of SLN. Finally, an irritation patch test conducted on 20 human volunteers demonstrated that both ethosomes and solid lipid nanoparticles can be safely applied on the skin.

A bilayer microneedle for therapeutic peptide delivery towards the treatment of diabetes in db/db mice

Microneedles (MN) provide an alternative approach to hypodermic injection for the delivery of biomacromolecule in a minimally invasive manner. In this work, lixisenatide-loaded bilayer MNs were prepared using polyvinylpyrrolidone K29/32 (PVP K29/32) as the matrix material via a two-step molding method to avoid therapeutic wastes in the base layer. The MNs obtained in our study displayed good shapes and sufficient mechanical strength. After insertion into the rat skin, the needles of MNs fully dissolved in less than 3 min to release the therapeutic payloads. The Franz cell based transdermal permeation study revealed that the MNs could dramatically promote transdermal permeation efficiencies of the water-soluble rhodamine B and FITC-dextran (5 kDa). The skin insertion of MNs caused slight irritation and the skin recovered shortly. Also, the PVP K29/32-based MNs displayed minimum cytotoxicity against HaCaT cells and murine L929 cells. The in vivo pharmacodynamic study in db/db mice demonstrated that the MNs successfully delivered Lix and downregulated the blood glucose levels. Overall, the PVP K29/32-based bilayer fast-dissolving MNs offer a unique transdermal delivery system which can be further explored to a variety of biological therapeutics.

Formulation and evaluation of the novel herbal antibacterial gel to the treatment of cutaneous burn infections

Due to the rapid development of antibiotic resistance, the strong need for alternative strategies to tackle this problem is inevitable. The objective of this study was to prepare and evaluate the antibacterial effects of a pharmaceutical gel containing herbal extracts including Lawsonia inermis (henna) and Matricaria chamomilla. Using hydroxypropyl methylcellulose (HPMC), carboxymethylcellulose (CMC), and propylene glycol (PEG), the pharmaceutical gel was formulated and the physical properties of the formulation were specified at 37 ± 2°C. The total phenolic content (TPC) of extracts was determined using the Folin–Ciocalteu method and expressed as mg gallaic acid (GA) per gram extract (Ex). The release of the polyphenol compounds from the optimum formulation was investigated using the Franz cell device. Eventually, the disc diffusion method was used to evaluate the antibacterial activity of optimum formulation against the two pathogenic bacteria strains Staphylococcus aureus and Pseudomonas aeruginosa. The results showed that the optimum formulation was stable at least for 3 months. The TPC of the aqueous extract of henna leaves, the hydroalcoholic extract of chamomile flowers, and the optimum formulation was 57.8, 181.08, and 202.75 mg GA/g Ex, respectively. Nearly 80% of the phenolic compounds in the optimum formulation were released over 4 h. The phenolic compounds have inhibitory effects on the growth of S. aureus and P. aeruginosa. On the basis of this finding, the formulation had excellent stability, viscosity, homogeneity, extrudability, and antibacterial activity which can be employed as a topical pharmaceutical gel in cutaneous burn infection treatment.

Dermal absorption of gallium antimonide in vitro and pro-inflammatory effects on human dermal fibroblasts

Gallium antimonide (GaSb) is a group III-V compound semiconductor with a comparatively narrow band gap energy (0.73 eV at 300 K) that allows efficient operation in the near-infrared region. This property may be useful in developing new biomedical instruments such as epidermal optoelectronic devices. The present study investigated the absorption of GaSb in pig skin in vitro for 24 h using Franz cells. A donor solution was prepared by soaking GaSb thin films in synthetic sweat. The results showed that both gallium and antimony penetrated the skin, and permeation and resorption occurred for gallium. Histopathological findings showed no inflammatory responses in pig skin exposed to GaSb for 24 h. Cytotoxicity was significantly elevated after 3 and 7 days, and pro-inflammatory cytokines and IL-8 levels were low after 1 and 3 days but elevated 7 days following the direct culturing of human dermal fibroblasts (HDF) on GaSb thin films. These results demonstrate that the short-term cytotoxicity and pro-inflammatory effect of GaSb on HDF were relatively low.

Evaluation of Electroosmotic Flow Promoted By a Porous Microneedle Array

A transdermal drug delivery (TDD) system has been an attractive method to delivery needed drug into our bodies directly with few side effect, and electroosmotic flow (EOF) is known as the method to enhance TDD and even the extraction of interstitial fluid (ISF) which contains biomarkers such as glucose. The limitation of these transdermal penetration / extraction arises from stratum corneum (SC), an outermost layer of skin. SC is highly resistive and functions as a barrier for harmful molecules, which makes it difficult to apply direct currents to the skin by mild and safe voltage and to deliver larger molecular drugs (>600 Da) into the skin1. The microneedle array (MNA) is one of the promising approaches to break through SC without pain, and solve problems of the electrical and the drug pass. Recently, we have realized organic polymer-based porous microneedles (PMN) made of poly(glycidyl methacrylate) (PGMA) which were prepared by the combination of molding process and the porogen method 2. In this study, we developed a PMN inducing the EOF for efficient TDD and extraction of ISF (Figure 1a). Furthermore, in order to enhance efficiency in the induction of the EOF, negatively-charged hydrogel was embedded in the porous channels of the PMN.The modified PMN was prepared by the following steps. Briefly, naked PMN was prepared by the step described in the report2. After that, the naked PMN chips were immersed in an aqueous solution containing 0.05 M of AMPS, 0.10 M of MBAAm, ammonium persulfate and tetramethylenediamine at 4 ℃ for more than 8 h, followed by thermal copolymerization at 70 ℃ for more than 8 h.First, we evaluated the property of water transport by EOF. A chip of PAMPS-modified PGMA was bound with side-by-side Franz cells with a horizontal capillary, and constant DC currents were applied by a source meter. McIlvaine buffer (pH 7.0) was used as the electrolyte. Flow velocity was calculated from the moving of the water surface in the capillary. The flow velocity of the EOF increased by the modification with AMPS (Figure 1b). The reason was that the negative charge was increased by the AMPS modification and that promoted the EOF.In order to evaluate the property of large molecule transport, the same system as water transport was employed as well. The Franz cells were poured with McIlvaine buffer (pH 6.0) and 0.75 mg/mL FITC-dextran (10,000 Da, pKa 6.4) was added to the donor chamber. We applied a constant DC current in direction from the donor side to the receptor side, and a 100 µL portion was collected from the receptor chamber every hour for analysis by a fluorescence spectrophotometer. From the result, we found that PAMPS significantly contributes to transport for the large molecule transport (Figure 1c). This might be due to dense polymer chains of PMAPS capturing large molecules such as dextran.Finally, the transdermal injection was demonstrated by enzymatic fructose / O2 battery into pig’s skin (Figure 1d). The injection of FITC-dextran at an anode was evaluated the PMN modified with 0.05 M AMPS and the cotton with 0.3 mL McIlvaine buffer (pH 6.0) containing 0.2 M D-fructose and 0.75 mg/mL FITC-dextran. The cross-sectional fluorescence image of the skin was observed after the application of the current around 0.2 mA/cm2 for 1 hour. These results showed that this novel PMN with negatively charged hydrogel can be a desirable tool for enhancing the injection of large molecules such as drugs and nutrients.Reference[1] Brown, M. B, Martin, G. P., Jones, S. A. & Akomeah, F. K. Dermal and transdermal drug delivery system: Current and future prospects. Drug Deliv. J. Deliv. Target. Ther. Agents 13, 175-187 (2006).[2] Liu, L., Kai, H., Nagamine, K., Ogawa, Y. & Nishizawa, M. Porous polymer microneedles with interconnecting microchannels for rapid fluid transport. RSC Adv. 6, 48630-48635 (2016). Figure 1

Designed Electroosmotic Polymer for an Anti-Drying Contact Lens Device

A smart contact lens (CL) is an attractive wearable device that could enable noninvasive monitoring of physiological information and augmented reality display in addition to vision correction. Whereas, the CL is one of the causes of dry eye syndrome due to more rapid moisture evaporation through the CLs than that from the normal tear film covered by a lipid layer. Conventional rehydration by regular eye drops takes time and efforts. The only example so far of a smart CL for maintaining the moisture is that of Lee et al., who developed a CL coated with single layer of graphene which decreases the evaporation. Hence, there is still a high demand to develop a CL equipped with an anti-dehydration function. We here designed a charge-fixed soft CL generating electroosmotic flow (EOF) under an electric field for maintaining the moisture of the lens (Figure 1a) [1]. The CL supplies tears via EOF from the tear meniscus (temporary tear reservoir) behind the lower eyelid. EOF is the motion of water induced by an applied electric field across a fluid conduit like a capillary tube or a porous material such as hydrogels. When the fluid conduit contains fixed electrical charges, the dominant electromigration of the mobile counter ions produces net water flow.The soft CL made of negatively charged hydrogel were fabricated by copolymerizing methacrylic acid (MA, 0 – 15 wt%), 2-hydroxyethyl methacrylate (HEMA) and methyl methacrylate (MMA). At First, we evaluated the efficiency of EOF generation for each of the 0.2 mm thick hydrogel films. DC currents of 1.0, 2.0, and 3.0 mA were applied to the films sandwiched by the side-by-side Franz cell with a horizontal capillary, and the flow velocity of EOF was calculated from the movement of the water surface in the horizontal capillary. These results indicate that a hydrogel with a high MA content is suitable for effective EOF generation in hydrogels. Secondly, compression tests were performed to the films to evaluate the indentation fracture toughness. The larger MA content tends to make the hydrogel become brittle and the hydrogel with 15 wt% MA was easily broken by folding. By considering both the results of the EOF strength and the fracture toughness, we mainly used the 10 wt% MA hydrogel in subsequent experiments. The water content and the conductance of the films were separately monitored at 100 kHz during the slow drying under 75% humidity. This result supports that the change in water content in the hydrogel film can be traced by the conductance measurement (Figure 1b). Finally, the EOF-based moisturization for the battery-mounted CL was demonstrated using an enzymatic fructose–oxygen fuel cell [2], in which the CL was put on a hemispherical jig (ocular model) equipped with printed Ag/AgCl electrodes for conductance measurements (Figure 1c). The lower end (≈3 mm) of the CL was dipped in a buffer solution containing D-fructose to imitate a tear meniscus. Figure 1d shows the observed changes in conductance (moisture) of CLs without EOF (gray, natural drying) and with upward EOF generated by an enzymatic fuel cell (red). The results demonstrated that applying an upward EOF through the CL reduced the moisture loss of the lens compared to natural drying. Since the demonstration was conducted at a relatively dry condition (40% humidity) for the hanging CLs, it can be expected that the moisturization would be more effective in situations where CLs are put onto eye. From these results, we believe that the proposed hydrogel will promise development of a contact lens devices with an anti-drying function.Reference[1] S. Kusama, K. Sato, S. Yoshida, and M. Nishizawa, “Self-Moisturizing Smart Contact Lens Employing Electroosmosis”, Adv. Mater. Technol. 5, 1, 1–9 (2020)[2] Y. Ogawa, K. Kato, T. Miyake, K. Nagamine, T. Ofuji, S. Yoshino, and M. Nishizawa, “Organic Transdermal Iontophoresis Patch with Built-in Biofuel Cell,” Adv. Healthc. Mater. 4, 506–510 (2015). Figure 1

FORMULATION AND EVALUATION OF MEFENAMIC ACID OINTMENT USING PENETRATION ENHANCERS

Objective: The aim of study was to formulate and evaluate Mefenamic acid ointment by the addition of penetration enhancer’s clove oil.
 Methods: 1%, 2% and 3% formulations of Mefenamic acid ointment formulated as per B. P, by melting hard paraffin 4.75g at 60 °C initially and to this 4.75 g wool fatwas incorporated, followed by addition of soft paraffin 80.75g and then adding Cetostearyl alcohol 4.75g and 1,2 and 3 ml clove oil by continuous stirring later on ointment being cooled at room temperature. These formulations were checked for consistency, Spreadability, homogeneity, PH, viscosity, skin irritation, drug content, UV absorbance, Differential scanning calorimetry (DSC) and XRD (X. ray diffraction) studies. In vitro pattern via using Franz cells besides with the use of dialysis cellulose membrane was done.
 Results: All the synthesized formulations illustrated fine physicochemical characteristics. SEM and XRD Studies expressed that there were no physicochemical incompatibilities among active ingredient (Mefenamic acid salt) and additives combined as drug permeation enhancers (clove oil).3% formulation showed maximum released 65.199%.
 Conclusion: In the present study, it was noted that clove oil can enhance the permeation of Mefenamic acid topical ointment.

Visualisation of H2O2 penetration through skin indicates importance to develop pathway-specific epidermal sensing

Elevated amounts of reactive oxygen species (ROS) including hydrogen peroxide (H2O2) are observed in the epidermis in different skin disorders. Thus, epidermal sensing of H2O2 should be useful to monitor the progression of skin pathologies. We have evaluated epidermal sensing of H2O2 in vitro, by visualising H2O2 permeation through the skin. Skin membranes were mounted in Franz cells, and a suspension of Prussian white microparticles was deposited on the stratum corneum face of the skin. Upon H2O2 permeation, Prussian white was oxidised to Prussian blue, resulting in a pattern of blue dots. Comparison of skin surface images with the dot patterns revealed that about 74% of the blue dots were associated with hair shafts. The degree of the Prussian white to Prussian blue conversion strongly correlated with the reciprocal resistance of the skin membranes. Together, the results demonstrate that hair follicles are the major pathways of H2O2 transdermal penetration. The study recommends that the development of H2O2 monitoring on skin should aim for pathway-specific epidermal sensing, allowing micrometre resolution to detect and quantify this ROS biomarker at hair follicles.Graphical abstract

Solvent Diffusion Method: An Effective Approach to Formulate Nanosponges Loaded with Naproxen Sodium

Objective: Solubility of naproxen sodium is limited. In conventional dosage form it causes different gastro intestinal problems. To overcome these difficulties naproxen sodium loaded nano sponges were designed.
 Methodology: Nanosponges were formulated by using emulsion solvent evaporation technique. To obtain dispersion of nanosponges, homogenization of active drug, with specified quantities of polyvinyl alcohol, dichloromethane, ethyl cellulose and distilled, water was done. Compatibility among excipients and active drug was checked by FTIR and results didn’t show any interaction between them. 11 trial formulations were tested for poly dispersity, zeta potential, particle size and viscosity.
 Results: Results showed all formulations except NS9, NS10 and NS11 were in nano range. Formulation NS1 to NS6 fall in category of “mid poly dispersity” and formulation NS7 to NS11 were in the category of “very poly dispersity”. Values of Zeta potential of all formulations were in negative range -0.106 to -9.75 mV. The value of viscosity of all formulations were 0.8872. NS2 and NS3 were selected for further testing like Franz cell diffusion study, stability testing and drug loading efficiency. In Franz cell diffusion study, drug release for NS2= 89.62%, for NS3= 89.10% at 50 minutes’ time. Stability studies performed for the 21 days, NS2 and NS3 revealed slight change in percentage drug content at 4°C and 25°C, and major changes were observed at 45°C temperature. Drug loading efficiency was found in NS2= 97.659 % and for NS3= 98.901%.
 Conclusion: Nanosponges formulations loaded with naproxen sodium have successfully been prepared.

Comparison of the In Vitro and Ex Vivo Permeation of Existing Topical Formulations Used in the Treatment of Facial Angiofibroma and Characterization of the Variations Observed.

Rapamycin has been used topically to treat facial angiofibromas associated with tuberous sclerosis for more than a decade. In the absence of a commercial form, a large number of formulations have been clinically tested. However, given the great heterogeneity of these studies, particularly with regard to the response criteria, it was difficult to know the impact and thus to compare the relevance of the formulations used. The objective of this work was therefore to evaluate the link between the diffusion of rapamycin and the physico-chemical characteristics of these different formulations on Strat-M® membranes as well as on human skin using Franz cells. Our results underline the importance of the type of vehicle used (hydrogel > cream > lipophilic ointment), the soluble state of rapamycin and its concentration close to saturation to ensure maximum thermodynamic activity. Thus, this is the first time that a comparative study of the different rapamycin formulations identified in the literature for the management of facial angiofibromas has been carried out using a pharmaceutical and biopharmaceutical approach. It highlights the important parameters to be considered in the development and optimization of topical rapamycin formulations with regard to cutaneous absorption for clinical efficacy.

Preparation and evaluation of kojic acid dipalmitate solid lipid nanoparticles

IntroductionNatural dissolution of kojic acid dipalmitate (KAD) in oil medium makes it difficult to load pharmacologic dose in a stable penetrative drug delivery system as topical dosage form. This study aimed to evaluate solid lipid nanoparticles (SLNs) as a drug delivery platform for the topical delivery KAD. MethodsSLNs were made with five different lipids and homogenization methods. The particle size was analyzed and the best lipid was chosen. Fourier-transform infrared spectroscopy, scanning electron microscopy, and differential scanning calorimetry were done. Then, KAD cream, KAD hydrogel, SLN-based KAD cream, and SLN-based KAD hydrogel were prepared and their release and permeation profile were evaluated on rat skin by Franz cell. ResultThe particle size of SLNs prepared with glycerol monostearate was about 70 nm. The optimum formulation was the combination with GMS 100 mg, KAD 10 mg, and ethanol/acetone in 2.5:1.5; all of which had the KAD entrapment efficiency of about 47%. The release of KAD powder was the fastest, and release from the SLN-based KAD hydrogel was slower than others. The KAD loaded in the SLN had the highest concentration in the stratum corneum, and KAD loaded in hydrogel formulation had the most concentration in the epidermis layer. ConclusionKAD can be loaded in the SLN with fair entrapment efficiency and particle size of <100 nm. KAD loaded in SLN-based cream made the maximum concentration of KAD in the epidermis compared with other formulations. Thus, SLN-based KAD cream is suggested as a proper topical formulation.

Biomimetic Artificial Membrane Permeability Assay over Franz Cell Apparatus Using BCS Model Drugs.

A major parameter controlling the extent and rate of oral drug absorption is permeability through the lipid bilayer of intestinal epithelial cells. Here, a biomimetic artificial membrane permeability assay (Franz–PAMPA Pampa) was validated using a Franz cells apparatus. Both high and low permeability drugs (metoprolol and mannitol, respectively) were used as external standards. Biomimetic properties of Franz–PAMPA were also characterized by electron paramagnetic resonance spectroscopy (EPR). Moreover, the permeation profile for eight Biopharmaceutic Classification System (BCS) model drugs cited in the FDA guidance and another six drugs (acyclovir, cimetidine, diclofenac, ibuprofen, piroxicam, and trimethoprim) were measured across Franz–PAMPA. Apparent permeability (Papp) Franz–PAMPA values were correlated with fraction of dose absorbed in humans (Fa%) from the literature. Papp in Caco-2 cells and Corti artificial membrane were likewise compared to Fa% to assess Franz–PAMPA performance. Mannitol and metoprolol Papp values across Franz–PAMPA were lower (3.20 × 10−7 and 1.61 × 10−5 cm/s, respectively) than those obtained across non-impregnated membrane (2.27 × 10−5 and 2.55 × 10−5 cm/s, respectively), confirming lipidic barrier resistivity. Performance of the Franz cell permeation apparatus using an artificial membrane showed acceptable log-linear correlation (R2 = 0.664) with Fa%, as seen for Papp in Caco-2 cells (R2 = 0.805). Data support the validation of the Franz–PAMPA method for use during the drug discovery process.

Effect of formulation on properties, stability, carvacrol release and antimicrobial activity of carvacrol emulsions

The structural design of essential oil emulsions can be exploited to modulate their antimicrobial activity, through the effect that the main formulation parameters (oil phase composition and type of emulsifier) have on the release of encapsulated antimicrobial compounds. In this work, different emulsions containing carvacrol, selected as model essential oil component, were characterized in terms of emulsions size, stability, and carvacrol release and solubilization, determined in Franz cells, and tested for minimum inhibitory and microbicidal concentration against P. fluorescens, S. epidermidis, and S. cerevisiae.The results showed that carvacrol fraction in the oil phase significantly affected oil viscosity, density, and O/W interfacial tension. Carvacrol solubilization in the aqueous phase, in equilibrium with the oil mixture, increased with the concentration of carvacrol in the oil phase and with the presence of an emulsifier/stabilizer in the aqueous phase. However, when encapsulated in emulsions carvacrol solubilization exhibited a weak dependence on carvacrol fraction in oil phase because part of the emulsifier/stabilizer was adsorbed at the O/W interface. Higher carvacrol solubilization was observed for WPM Pickering emulsions, followed by WPI and T80 emulsions. The antimicrobial activity was proportional to carvacrol solubilization, suggesting that emulsion droplets act as micrometric tanks for carvacrol, which is steadily released over time in the aqueous phase. The high carvacrol solubilization in the aqueous phase at higher carvacrol fractions in the oil phase (≥75% w/w) was also responsible for lower T80 and WPI emulsion stability because of coalescence, whereas all WPM emulsions exhibited signs of flocculation.

Transcutol® P Containing SLNs for Improving 8-Methoxypsoralen Skin Delivery.

Topical psoralens plus ultraviolet A radiation (PUVA) therapy consists in the topical application of 8-methoxypsoralen (8-MOP) followed by the skin irradiation with ultraviolet A radiation. The employment of classical 8-MOP vehicles in topical PUVA therapy is associated with poor skin deposition and weak skin permeability of psoralens, thus requiring frequent drug administration. The aim of the present work was to formulate solid lipid nanoparticles (SLNs) able to increase the skin permeation of 8-MOP. For this purpose, the penetration enhancer Transcutol® P (TRC) was added to the SLN formulation. SLNs were characterized with respect to size, polydispersity index, zeta potential, entrapment efficiency, morphology, stability, and biocompatibility. Finally, 8-MOP skin diffusion and distribution within the skin layers was investigated using Franz cells and newborn pig skin. Freshly prepared nanoparticles showed spherical shape, mean diameters ranging between 120 and 133 nm, a fairly narrow size distribution, highly negative ζ potential values, and high entrapment efficiency. Empty and loaded formulations were almost stable over 30 days. In vitro penetration and permeation studies demonstrated a greater 8-MOP accumulation in each skin layer after SLN TRC 2% and TRC 4% application than that after SLN TRC 0% application. Finally, the results of experiments on 3T3 fibroblasts showed that the incorporation of TRC into SLNs could enhance the cellular uptake of nanoparticles, but it did not increase their cytotoxicity.

Co-electrospraying technology as a novel approach for dry powder inhalation formulation of montelukast and budesonide for pulmonary co-delivery.

In the current study electrospraying methodology was used for particle engineering of montelukast and budesonide to prepare a combined inhalable dry powder formulation applicable as a smart regimen in asthma treatment. For this, electrospraying was carried out using different solvents and drug concentrations. No carrier was added for the formulation of montelukast-budesonide combination as montelukast played the role of both active ingredient and carrier. Scanning electron microscopy, particle size analysis, gas chromatography, powder X-ray diffraction, Fourier transform infrared spectroscopy, and differential scanning calorimetry were used to evaluate the physicochemical properties of the produced drug particles. In vitro drug deposition pattern was assessed using next generation impactor, and the dissolution profile of the selected formulations was characterized via modified diffusion franz cell method. The FPF value for the co-electrosprayed carrier free formulation of montelukast-budesonide was 38% with a significantly enhanced dissolution rate for budesonide compared to the budesonide alone formulations. The pharmacological effects of hypothesized combined formulation was assessed by measuring its power to inhibit the production of reactive oxygen species in human normal lung cells. The results showed that the combination of montelukast and budesonide can exert a synergistic effect. The findings in the current study emphasize that using montelukast as a carrier for budesonide not only has greatly improved the aerosolization behavior and dissolution rate of budesonide but also has resulted in synergistic pharmacological effects, indicating the suitability of this combination as an anti-asthmatic therapeutic.

Deformable liposomes as enhancer of caffeine penetration through human skin in a Franz diffusion cell test.

The permeation of hydrophilic molecules through the skin is still a challenge due to the barrier posed by stratum corneum, the outermost layer of the skin. Liposomes have frequently been used as carriers for different types of drugs and may also function as permeation enhancers. Propylene glycol has also been used as an edge activator in liposomes to increase the permeation. The aim of this work was to prepare liposomes containing an edge activator and loaded with caffeine to evaluate the potential of caffeine reaching the deeper layers in the skin. The formulations were prepared by a top-down process using high-pressure homogenization at 20000 psi for 10min. They were characterized by size, polydispersity index (PI), zeta potential (ZP), pH, caffeine content and encapsulation efficiency (EE%) on preparation (time zero) and after 30days. Cytotoxicity of blank and loaded liposomes was assessed by MTT proliferation assay with a normal keratinocyte cell line (HaCaT). In vitro permeation tests were performed with human skin in Franz cells over 24h, and caffeine concentration was determined in the skin surface, stratum corneum, dermo-epidermal fraction and receptor medium by HPLC. The caffeine liposomes with (DL-Caf) or without propylene glycol (CL-Caf) showed, respectively, mean size 94.5 and 95.4nm, PI 0.48 and 0.42, ZP+1.3 and+18.1mV and caffeine content of 78.57 and 80.13%. IC50 values of caffeine in DL-Caf (3.59 v/v %) and CL-Caf (3.65 v/v %) were not significantly different from conventional blank liposome (3.27 v/v %). The DL-Caf formulation presented the best capability to enhance the caffeine permeation through the skin, resulting 1.94-folds higher than caffeine solution. Furthermore, the caffeine flux from DL-Caf was 1.56- and 3.05-folds higher than caffeine solution and CL-Caf, respectively. On the other hand, CL-Caf showed the lowest caffeine penetration revealing the importance of edge activator to aid hydrophilic drug penetration to all skin layers. The DL-Caf formulation tested was able to improve the permeation of caffeine through the stratum corneum and dermo-epidermal layers, suggesting that this delivery system may be effective for deep skin delivery of hydrophilic drugs.

Topical Delivery of Nivolumab, a Therapeutic Antibody, by Fractional Laser and Pneumatic Injection.

PD-L1 is a tumor ligand that binds to the PD-1 receptor on immune cells, thereby inhibiting the antitumor immune response. The antibody nivolumab is a PD-1 inhibitor, Food and Drug Administration approved for systemic treatment of several aggressive cancer types. Topically applied nivolumab may hold potential as a future strategy to treat keratinocyte cancer, but its molecular properties preclude unassisted topical uptake. The aim of this study was to investigate uptake and biodistribution of topically delivered nivolumab, assisted by two physical enhancement techniques with different delivery kinetics; ablative fractional laser (AFL) and electronically controlled pneumatic injection (EPI). In vitro porcine skin was exposed to CO2 AFL (20 mJ/mb, 5% density), followed by passive diffusion of nivolumab in a Franz cell (1 mg/ml, 18 hours, n = 6) or treated with EPI (4 bar) for immediate delivery of nivolumab (1 mg/ml, 10 minutes, n = 6). The resulting nivolumab skin concentrations were quantified by enzyme-linked immunosorbent assay (ELISA) at three skin depths (100, 500, and 1500 µm), comparing the uptake from assisted delivery with intact skin. Biodistribution of nivolumab in the skin for all interventions was visualized by laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) and fluorescence microscopy. Delivery of nivolumab by AFL-assisted passive diffusion and immediate EPI both resulted in significantly enhanced uptake of nivolumab in all skin depths compared with intact skin (P < 0.05). WithAFL, nivolumab concentrations reached 86.3 µg/cm3 (100 µm), 105.8 µg/cm3 (500 µm), and 19.3 µg/cm3 (1500 µm), corresponding to 2-10% of the applied concentration, with the highest deposition in the mid dermis. Immediate EPI delivered 429.4 µg/cm3 (100 µm), 584.9 µg/cm3 (500 µm), and 295.9 µg/cm3 (1500 µm) into the skin, corresponding to 29-58% of the applied nivolumab concentration. From qualitative visualization of the biodistribution, it appeared that nivolumab distributed in a horizontal and continuous homogenous band in the upper and mid dermis through AFL-exposed skin, whereas EPI-delivery showed a deep focal deposition extending into the deep dermis. AFL-assisted passive diffusion and immediate EPI-assisted delivery show the potential to deliver therapeutic antibodies locally. Future in vivo and pharmacokinetic studies would reveal the full potential for topical antibody delivery by energy-based devices. Lasers Surg. Med. © 2020 Wiley Periodicals LLC.