Novel drug delivery strategies for improving econazole antifungal action

Accepted Manuscript Title: Novel Drug Delivery Strategies for Improving Econazole Antifungal Action Author: Alireza Firooz Shohreh Nafisi Howard I. Maibach PII: DOI: Reference:

S0378-5173(15)30213-1 http://dx.doi.org/doi:10.1016/j.ijpharm.2015.09.015 IJP 15200

To appear in:

International Journal of Pharmaceutics

Received date: Revised date: Accepted date:

8-5-2015 10-9-2015 11-9-2015

Please cite this article as: Firooz, Alireza, Nafisi, Shohreh, Maibach, Howard I., Novel Drug Delivery Strategies for Improving Econazole Antifungal Action.International Journal of Pharmaceutics http://dx.doi.org/10.1016/j.ijpharm.2015.09.015 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Novel Drug Delivery Strategies for Improving Econazole Antifungal Action Alireza Firooz1, Shohreh Nafisi2,3* [email protected], Howard I. Maibach3*  [email protected] 1

Center for Research and Training in Skin Diseases and Leprosy, Tehran University of

Medical Sciences 2

Department of Chemistry, Central Tehran Branch, IAU, Tehran, Iran

3

Department of Dermatology, University of California, San Francisco, CA, USA

*

Correspondence authors. Tel.: (408)625 0046, (415)473 9693; fax: (415)673.

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Graphical abstract

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ABSTRACT Econazole is a commonly used azole antifungal in clinical treatment of superficial fungal infections. It is generally used as conventional cream and gel preparations under the brand names of Spectazole (United States), Ecostatin (Canada), Pevaryl (Western Europe). Treatment efficiency of antifungal drugs depends on their penetration through target layers of skin at effective concentrations. Econazole′s poor water solubility limits its bioavailability and antifungal effects. Therefore, formulation strategies have been examined for delivering econazole through targeted skin sites. The present overview focuses on novel nano-based formulation approaches used to improve econazole penetration through skin for treatment of superficial fungal infections. Keywords: Econazole; Nanoparticle; Percutaneous penetration; Drug Delivery

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1. Introduction Econazole (ECN; commonly used as nitrate salt) is an imidazole antifungal structurally related to another imidazole derivative, miconazole (Fig. 1) (Thienpont et al., 1975) and utilized to treat fungal infections such as tinea pedis and cruris, pityriasis versicolor (econazole cream, FDA Prescribing Information). Topical econazole is commercially available as cream and ointment. Econazole content in the formulation is 1% which can not be completely dissolved, thus, physical phase separation of the cream occurs due to econazole salting effect. Approximately, 90% of topically applied econazole nitrate (as the cream) remains on the skin surface (Heel et al., 1978). Econazole low solubility limits its concentration at the desired site of action and reduces permeation rate. It should be applied for several weeks (twice daily) till infection signs/symptoms on the skin completely disappear. Side effects such as irritation, redness, burning or itching have also been reported among 1 to 4% of patients (Heel et al., 1978). Thus, the conventional topical treatment is not appropriate in all cases and may be considered inconvenient by patients, which might affect compliance (Degreef and De Doncker, 1994). Topical antifungal therapy needs high drug amount at the site of action, that is, mainly stratum corneum while decreasing its systemic absorption that reduces unwanted side effects. Nanoparticle delivery to skin has attracted wide attention for topical antifungal therapies by enhanced skin penetration, as well as controlled and sustained release of encapsulated drug ingredients. By providing a drug depot and controlled release system at infection site, such formulations have potential to provide high local drug concentrations without significant systemic distribution. Colloidal drug carriers such as microemulsions, vesicular carriers (including liposomes, ethosomes, niosomes), lipidic and polymeric particulate carrier systems have been examined to enhance antifungals permeation. Review articles based on new alternative formulation approaches to improve penetration of antifungal drugs have been 4   

recently published (Kumar et al., 2014; Neubert, 2011; Benson, 2010; Yang et al., 2008; Güngör et al., 2013; Glujoy et al., 2014). Several nano-formulation strategies have been studied for delivering econazole through targeted skin sites (Canu et al., 2006; Passerini et al., 2009; Sanna et al., 2007; Sanna et al., 2010; Bachhav et al., 2011; Yordanov et al., 2012; Kumar et al., 2013). The purpose of this study is extracting and rationalizing data in nano-based formulations of fluconazole, while focuses on how nanoscale properties support, influence, or change significantly the knowledge of fluconazole and improve treatment of diseases. Recent studies are discussed and summarized in Table 1. 2. Penetration of Nanoparticles into Skin Several researchers have explained that drug penetration into skin is affected by the physicochemical properties of nanoparticles such as particle size, surface charge, surface area, solubility, effect of solvents, salt form of the drug and chemical composition (Liang et al., 2013; Labouta et al., 2011; Agharkar et al., 1976; Berge et al., 1977). Amongst these factors, nanparticles size and charge have been widely studied. Particle size is the most important property which influences skin penetration. Analysing nanoparticles at different sizes consisting of diverse materials with various surface properties has revealed that smaller size of nanoparticles are more likely to penetrate to skin compared with lager size. In addition, hair follicles are considered an important shunt route for nanoparticles. Particles of approximately 300–600 nm in size exhibit the deepest penetration into hair follicles (Patzelt et al., 2011) where they were stored significantly longer than in the stratum corneum (Lademann et al., 2011). Drug penetration can also be affected by the surface charge of nanoparticles. The results in the literature are contradictory. While some experiments report higher skin penetration potential of the positively charged submicron emulsions, other experiments show more skin 5   

penetration by negatively charged nanocompounds. Positively charged submicron emulsions exhibit more efficacy in promoting drug bioavailability due to droplets higher binding affinity to skin. Since the skin epithelial cells carry a negative charge on their surface and the presence of protein residues on the outer membrane of cells creates a negative charge on the surface of epithelial cells in various tissues including skin (Rojanasakul et al., 1992; Piemi et al., 1999; Yilmaz and Borchert, 2005). Piemi utilized stearylamine or deoxycholic acid (DCA) incorporated ECZ in positively and negatively charged submicron emulsions and concluded that negatively surface modified emulsions exhibited a significant influence on the drug absorption across skin (Piemi et al., 1999). Jung et al. 2006 indicated that cationic liposomes penetrate deeper into hair follicles than their anionic counterparts (Jung et al. 2006). However, Gillet et al., reported enhanced skin penetration using negatively charged liposomes of betamethasone and betamethasone dipropionate (Gillet et al., 2011). It was explained that negatively charged vesicles caused a higher flux than positively charge counterparts, which in turn could enhance drug localization in superficial skin strata (Sinico et al., 2005; Gillet et al., 2011). Using the salt forms of drugs is one of the important ways for improving percutaneous absorption when the physicochemical characteristics of the parent drug molecule are unsuitable. A large number of drugs which are being used for local applications are weak acid or bases and ionize under normal physiological conditions. Ionized molecules are generally not well absorbed by biologic membranes. Increased absorption followed by drug ion pairing and salt formation can be related to electrical neutrality and increased drug lipophilicity. Using different salt counterions cause significant changes in drug solubility, dissolution rate, and other pharmaceutically important properties which are essential for drug absorption (Agharkar et al., 1976; Berge et al., 1977). Piroxicam salt formation with monoethanolamine salt and diethanolamine improved the physicochemical properties and 6   

enhanced skin permeability of piroxicam (Cheong &Choi 2002). Ethanolamine salt of biphenylacetic acid (BPA), showed 7.2- and 5.4-fold higher skin permeation than the parent drug at pH 7.4 and 5.0, respectively (Pawar et al., 2015). 3. Econazole Nano-Topical Delivery Systems under Current Development 3.1. Solid Lipid nanoparticles (SLNs) and Nanostructured Lipid Carriers (NLCs) Development of lipid based nanoparticles for drug penetration and targeting in strata has been widely studied (Lombardi et al., 2005). SLNs which were first prepared in 1991, are colloidal lipid carriers, solid at body and room temperature (Müller et al., 1995). SLNs and NLCs have shown a great potential as vehicles for topical administration of active substances. They are the most exploited carrier systems for controlled delivery of pharmaceuticals and cosmetic ingredients (Muller et al., 2002; Souto et al., 2004). They offer lower toxicity compared with polymeric systems, and can be produced in large scale. Their specific characteristic makes them an interesting carrier system for optimized skin delivery (Schlupp et al., 2011; Muller et al., 2002; Kakadia and Conway, 2014). Nanostructured lipid carriers are the next generation of nanocarriers developed to overcome the potential limitations of solid lipid nanoparticles such as low drug loading, increase in particle size due to aggregation and leakage of drug during storage (Müller et al., 2000). NLCs are composed of highly ordered solid lipids matrix with high imperfections in crystal lattice, creating enough space for drug accommodation (Muller et al., 2002; Souto et al., 2004). Spherical SLNs and NLCs possess an average diameter between 10 to 1000 nanometers. Small size of the lipid particles ensures close contact to stratum corneum and can enhance drug amount penetrating into skin and allows controlled drug release (Wissing and Muller, 2001). Solid lipid cores, composed of physiological and biodegradable lipids are stabilized by surfactants or emulsifiers and can solubilize lipophilic molecules. Surfactants are used to reduce surface tension between two immiscible liquids followed by droplet disruption and also prevent re7   

coalescence of droplets by adsorbing on their surfaces, thus producing a stable solution and fabricate particles especially in emulsion type systems. They can also extend over particle surface and induce positive/negative charge. Using two or more surfactants gives higher coverage of particles, prohibit aggregation and decrease tensions. Greater zeta potential is obtained by mixture of surfactants (Rosen and Kunjappu, 2012). Selection of right surfactant or emulsifier for superficial formulations is extremely important. It significantly affects the extent and rate of drug absorption from skin. Surfactant structure should not change during nano-formation. The interaction of drug with surfactants may impact drug stability and release at the targeted point, then influence drug absorption and ability to permeate through skin. Careful selection and correct grade of surfactants can limit exposure to atmosphere, and minimize drug degradation upon oxidation. Surfactants frequently come in contact with skin in the form of topical formulations and may induce skin reactions such as irritant contact dermatitis and inflammation by damaging the barrier properties of stratum corneum, and denaturing proteins in epidermis and dermis. Some surfactants are recognized as having strong irritant potency whereas skin can much better withstand other surfactant molecules termed as “mild”. Mildness of surfactant is one of the important properties that should be considered in the design of a topical formulation in either pharmaceutical or cosmetic products (Imokawa, 1997). Tweens (o/w emulsifier) and Spans (w/o emulsifier) are a range of mild non-ionic surfactants which augment the formulation stability, flexibility and cause wider compability. They are stable in mild acids, alkalis, and electrolytes and do not react with ionic ingredients or actives and have a safe history of use). Canu et al, 2006 developed a controlled released SLN gel containing econazole nitrate (ECN) by adding gelling agent: Ginshicel Chinese brand (HPMC K100M: hydroxylpropyl methylcellulose), lipid carrier: Precirol ATO (PCR) (Glyceryl Palmitostearate) and emulsifying agent: Tween 80 (Polyoxyethylene (20) sorbitan monooleat), [SLN1(PCR: 5% 8   

w/w) and SLN2 (PCR:10% w/w); ECN: 1.0% w/w]. SLN1 and SLN2 encapsulation efficiency ranged 96-102%, respectively. Average size of the nanoparticles reported as 480 nm which was constant within 30 days. For all formulations, minor amount of drug was released after 2 hours from the gels. ECN release from the gel (formulation without SLN) was higher and more irregular compared with the gels which contained SLNs. ECN release rate from SLN gels was found to be related to the total lipids content of nanoparticles. Ex vivo drug penetration tests through the porcine stratum corneum indicated that release rate for SLN1 gel containing 5% w/w of lipid was twofold higher than SLN2, containing 10% Precirol. Therefore, higher lipid content led to the formation of lipid-enriched shell SLNs which caused slower release rate. Therefore, SLN formulated in gels suggested promising controlled release formulations for ECN topical delivery (Canu et al., 2006). Sanna et al., 2007 fabricated econazole-loaded SLNs for topical administration of econazole. SLNs-ECN were prepared by o/w high-shear homogenization method using different ratios of lipid/drug (SLN1:5/1, SLN2:10/1). The optimized formulation was incorporated into hydrogels. No aggregation was observed upon SLNs incorporation into hydrogels and the average sizes of the econazole-loaded nanoparticles remained substantially unchanged (156 nm for SLN1; 163 nm for SLN2). The white color gels containing SLN1and SLN2 exhibited good stability and properties of consistence and spreadability, regardless of the concentration of the lipids used. For permeation studies, stratum corneum of the adult pig ears was peeled off from both surfaces and then used. The ECN gel without SLN demonstrated a more rapid and irregular permeation profile compared with SLN1 and SLN2 gels (gels of drug encapsulated into SLNs). Enhanced penetration was observed for SLN1 gel with 43.3 µg.cm-2 of drug permeation after 9 hours compared with SLN2 with 26.8 µg.cm2

drug permeation. ECN release profile from SLN formulations was linear with square root of

time. Minor amount of ECN released from the gel within the first hour and the drug 9   

permeated through stratum corneum ranged from about 33 µg.cm−2 after 2 hours to 125 µg.cm−2 at the end of the test. Total drug released through stratum corneum after 24 hours from SLN1 gel was 124 µg.cm-2 which was markedly different from SLN2 gel (48 µg.cm-2). It can be concluded that higher lipid content in SLN2 formulations enhanced diffusional distance (Muller et al., 2002; Souto et al 2004) and consequently led to a significantly slower drug release (Wissing et al., 2004). In vivo human skin permeation studies using tape stripping method indicated that after 1 h of application, the amounts of drug penetrated into stratum corneum from SLN1 and SLN2 gels were 2.32 and 2.01 µg.cm2

respectively, which was significantly higher than that from ECN gel (1.56 µg.cm-2). After 3

hours of application, econazole recovered into stratum corneum for the SLN2 gel (10% of lipid material) (0.90 µg cm-2) was markedly lower than the ECN gel (1.84 µg.cm-2). No significant differences were detected between SLN1 and SLN2 formulations for both considered application times. Higher drug penetration after a shorter application time from the SLN gels was attributed to the smaller size of the lipid particles compared with the nonencapsulated drug in the reference gel; in fact, nanoparticles may act as a reservoir and localize in the upper skin by penetrating into follicular openings and sticking to the stratum corneum (Jenning et al., 2000). By the adhesion of particles on the skin surface, a lipidic film forms which reduces drug penetration into the outermost skin layer (Wissing et al., 2004). SLN2 gel with higher lipid content produced a lipid layer on skin surface particularly after 3 hours of application. Thus, the drug penetrated within the first hour could diffuse in deeper skin layers within 3 hours. Consequently, SLNs could provide a controlled release system of ECN compared with the conventional gel. The amount of ECN recovered into the upper skin layers from ECN-SLN gels depended on the application time and the composition of particles. SLNs with lower lipid content improved ECN penetration rate into the upper skin

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layer after a short period of application and were suggested as a good candidate for dermal delivery of drugs (Sanna et al., 2007). Effect of SLNs fatty alcohol chain lengths (C12–C18) on ECN skin permeability was studied by Sanna et al. 2010. Nanoparticle emulsions were synthesized by o/w high shear homogenization method using a mixture of Precirol and fatty alcohols as the lipid phase (A1: Lauryl alcohol; A2: Myristyl alcohol; A3: Cetyl alcohol; A4: Stearyl alcohol with 12, 14, 16, 18 chain lengths, respectively), then loaded with ECN. Encapsulation efficiency of different formulations ranged from 95 to 99%.

Production yields ranged from 88 to 92%. A

formulation containing only Precirol and ECN was used as control. Ex vivo permeation studies of ECN from gel formulations were performed using porcine SC for 8 hours. The SC was cut and mounted on the bottom of a support (height 1.91 cm, diameter 2.28 cm), and around 200 mg of the gels were uniformly arranged on the surface of the skin. At the first hour of experiment, about 13 μg/cm2 of ECN was permeated from A4, while control, and A1–A3 gels released ECN at the second hour. At the end of the experiment, the amount of ECN permeated from control gel (39 μg/cm2) was almost equivalent to A3 and A4 gels and higher than A1 and A2 gels (24 and 34 μg/cm2, respectively). For all formulations, ECN permeation followed pseudo-first order kinetics and there was a linear relationship between the amounts of ECN released versus time with good correlation coefficients. Statistical analysis demonstrated that the ECN flux from the control gel was similar to A2–A4 gels and significantly higher than that of A1 formulation. An excellent linear relationship was observed between the ECN flux values against chain length of fatty alcohols. By increasing the fatty alcohols chain length from C12 to C18, the flux values increased.

The amount of

ECN penetration through skin by SLNs are strongly related to fatty acid chain length. Fatty acids can act as penetration enhancer of different drugs and have exhibited a parabolic relationship with alkyl chain length (Andega et al., 2001; Lee et al., 1993), with C10 and C12 11   

as the most effective (Sloan et al., 1998). Among the tested fatty alcohols, the cetyl and stearyl ones with 16 and 18 C atoms, respectively, were considered as the most efficient penetration enhancers, with potential application in drug delivery to skin. Long chain fatty alcohols may create less ordered solid lipid matrix and provide enough space for drug molecules, they may interact with densely packed lipids between extracellular spaces of SC and increase skin permeability (Sanna et al., 2010; Bunjes et al., 2000). Passerini et al 2009, compared ECN skin permeation of solid lipid microplates (SLMs) with solid lipid nanoparticles (SLNs). SLMs and SLNs were prepared at different lipid:drug weight ratios (5:1, 10:1, 12.5:1) by two production methods; spray congealing process and high-shear homogenization. Particle size of the SLMs and SLNs were 18–45 µm, 130–270 nm, respectively with drug encapsulation efficiency of 80–100%. SLMs and SLNs were incorporated into HPMC K100M hydrogels (hydroxylpropyl methylcellulose, Ginshicel Chinese brand). Epidermis was peeled off from ears skin of the adult domestic pigs and epidermal membrane was chosen for penetration studies. Permeation studies were carried out on SLMs or SLNs ECN gels and on non-encapsulated ECN gel. About 200 mg of each gel was uniformly spread on the surface of the skin (area = 4.08 cm2). The result of permeation studies demonstrated that the non-encapsulated ECN permeated the epidermis. The cumulative amount of ECN released from gel within the first hour was negligible (lag time 60 min); the total amount permeated through the epidermis ranged from about 32 µg/cm2 after 2 h to 124 µg/cm2 at the end of the test (24 hours), with a release rate of 25 µg/cm per h½. SLM gels exhibited similar ECN release rate and the same total amount of drug permeated after 24 hours. The release rate of ECN from SLN1, SLN2 or SLN3 gels showed that the cumulative amount permeated after 24 hours was not significantly different regardless of the amount of the lipid used. The lag times were between 60 and 97 min for all SLN gels. Comparison of the permeation results for SLN1 to SLN3 and nonencapsulated drug showed that SLN2 and 12   

SLN3 delayed ECN permeation through the skin (P < 0.05 and P < 0.01 after 9 hours; P < 0.001 and P < 0.05 after 24 hours, for SLN2 and SLN3, respectively). Minor difference in drug release rate and cumulative amount of ECN gels was observed after 24 hours from the particles with the same composition (SLM1 vs SLN1; SLM2 vs SLN2; SLM3 vs SLN3). As a result, ECN permeation profile through epidermis was influenced by the lipid content in SLN gel formulations. Lower lipid content led to more ECN permeation. Slower drug permeation from SLN2 and SLN3 gels caused enhanced diffusional distance related to higher lipid content, and greater affinity of the drug for the lipidic matrix. Strict adhesion between nano-sized SLNs may fix the drug into a formed film and lower diffusion through the stratum corneum. Lower skin adhesion and larger pores between the microparticles may improve accumulation of the released drug on the skin and cause more rapid permeation by SLMs. Comparing two synthesis methods demonstrated that spray congealing enabled production of ECN loaded SLMs with a suitable diameter for topical administration and highshear homogenization was proposed as a proper method for ECN-SLNs preparation. The present study demonstrated usefulness of SLNs as carriers for topical administration and suggested their potential for controlled drug delivery systems (Passerini et al., 2009). 3.2. Liposome Liposomes are self-closed structure vesicles composed of one or several concentric lipid bilayers with an aqueous phase inside and between lipid bilayers. They are often composed of phospholipid composition enriched in phosphatidylcholine and may also contain mixed lipid chains with surfactant properties such as egg phosphatidyl ethanolamine. They were first prepared in 1961, at the Babraham Institute, in Cambridge (Bangham & Horne 1964; Horne et al., 1963; Bangham et al., 1962). Size, charge and surface properties of liposomes can be easily tuned (Allen and Cullis, 2013; Torchilin, 2005; Torchilin, 2006). Liposomes

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can entrap hydrophilic compounds in their internal water part and hydrophobic compounds into the membrane. They are able to transfer entrapped and non-entrapped drugs into skin (Cevc, 2004; Imbert and Wickett, 1995; El Maghraby et al., 2008) and can act as carriers controlling the release of active ingredient. They can localize the drugs at the site of action, improve deposition within skin, reduce systemic absorption and minimizing drug side effects. They can also protect drugs from the inactivating effects of external conditions (El Maghraby et al., 2006). Current efforts in liposome dermal delivery focus around optimization procedures

and

novel

liposomic

compositions.

Transferosomes

and

ethoseomes,

ultradeformable or elastic vesicles, are the next generation of liposomes which follow the trans-epidermal water activity gradient in skin (Cevc et al., 1996; Touitou et al., 2000; Touitou et al., 2001). Transferosomes are composed of natural phospholipids like phosphatidyl choline (which self assemble into lipid bilayer) and a single chain surfactant which acts as an edge activator. The aqueous core is surrounded by the complex lipid bilayer and generates hydrophobic and hydrophilic moieties. Edge activator is a single chain surfactant that causes destabilization of the lipid bilayer thereby increasing the fluidity and elasticity of transferosomes and enables them to cross various transport barriers efficiently (Cevc et al., 1996; Cevc et al., 2002; Cevc, et al., 2003, Cevc and Gebauer, 2003; Cevc, 2004). Tranferosomes exhibit high drug entrapment efficiency which increases up to 90% for lipophilic drug (Sarmah et al., 2013). Comparing with other micelles, transferosomes possess a greater size and contain a water filled core which enables them to carry water as well as fatsoluble agent. They are more flexible and adaptable comparing with commonly used liposomes, but are chemically unstable due to predisposition to oxidative degradation and their formulations is expensive (Sarmah et al., 2013; Benson, 2006; Benson, 2010). Ethosomes are specific deformable vesicles which exhibit markedly higher transdermal flux in compare with liposomes. The main difference between liposomes and ethosomes is 14   

related to their high alcohol content and composition. Combination of phospholipids and high ethanol content in ethosomes causes more flexibility and elasticity which permits ethosomes to squeeze themselves through skin pores, deeper distribution and penetration in the skin lipid bilayers. Ethosomal systems have exhibited much more efficient in delivering of hydrophilic and lipophilic drugs to skin in terms of quantity and depth than either conventional liposomes or hydro alcoholic solutions. They exhibit good storage stability, because of the presence of ethanol which provides a net negative surface charge, thus avoiding vesicles aggregation due to electrostatic repulsion (Jain et al., 2007; Dubey et al., 2007a; Dubey et al., 2007b). The first nano-carrier-econazole product for antimycotic therapy (Epi-Pevaryl C® Lipogel) marketed in 1988 was a liposome preparation produced by Cilag company in Switzerland (Naef, 1996; Muller et al., 1998; Muller et al., 2000). Schaller et al. 1999 compared the efficiency of two commercially available econazole formulations (econazole nitrate cream, econazole liposome gel) on an uninfected reconstructed human epidermis and human cutaneous candidiasis (CC) model. Treatment with liposomal formulation led to the reduced damage of epidermal barrier and decreased toxic effects of lower epidermis. Signs of Candida albicans (CA)-specific damage to CC were greatly reduced and damage to CA blastospores was more pronounced. Therefore, ECN-liposome gel distribution was more intense on stratum corneum surface and CA blastospores than two commercially econazole formulations (Schaller et al., 1999). Korting et al. 1997, examined econazole liposomal gel as a drug carrier system in a controlled, double-blinded trial. Treatment either encompassed once-daily application for 14 days of ECN liposome gel 1%, branded econazole cream 1 %, or a generic clotrimazole cream 1%. Higher cure rate was observed in econazole liposome gel treatment group. Tolerability was considered slightly better in econazole liposome gel group, compared with

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econazole cream and clotrimazole cream treatment groups. The results of this study revealed that econazole liposome gel might be slightly superior to the conventional preparation in terms of efficacy and tolerability (Korting et al., 1997). 3.3. Niosome Niosome, a non-ionic surfactant-based vesicle (200-250 nm) with microscopic lamellar structure can be synthesized by mixing cholesterol and single alkyl chain non-ionic surfactant with subsequent hydration in aqueous media. They have gained popularity in the field of topical and transdermal drug delivery because of their special characteristic features like increasing the penetration of active ingredients, and serving as local reservoir which provides a solubilizing matrix for hydrophilic and lipophilic drugs (Jain and Namdeo, 1996; Vanlerberghe and Morancias, 1996; Schreier and Bouwstra, 1994). Handjani-Vila et al., were first to fabricate the niosomes by hydration of mixture of cholestrol and a single alkyl chain non-ionic surfactant (Handjani-Vila et al., 1979). Structure and properties of niosomes depend on their synthesis method and composition of the bilayers. Cholesterol intercalation in the bilayers causes reduced entrapment volume and thus entrapment efficiency (Szoka and Papahadjopoulos, 1980). There exist differences between liposomes and niosomes. Niosomes are synthesized from uncharged single-chain surfactant and cholesterol, whereas liposomes are prepared from double-chain phospholipids (neutral or charged). Cholesterol concentration in niosomes is lower than liposomes. The phospholipid ingredients of liposomes are chemically unstable because of their predisposition to oxidative degradation. Drug entrapment efficiency of liposomes is lower than niosomes. Besides, liposomes are expensive and require particular storage and handling (Jadon et al., 2009; Choi and Maibach 2005). Kumar et al., 2013 prepared econazole-niosome gels utilizing different ratios of cholesterol/span 80 (A1:1/1, A2:1/2, A3:1/3, A4:1/4) by thin film hydration technique. For A1 to A4, the percentages of the entrapped drug were reported as 62, 71, 88, 98%, 16   

respectively and the drug released as 70, 77, 82, and 92% respectively. Highest entrapment percentage and desired ECN sustained release was observed for A4 with the highest content of span 80. A4 followed zero order release pattern as shown by the linearity plot of time versus drug release (Kumar et al., 2013). 3.4. Polymeric micelles An amphiphilic methoxy-poly (ethylene glycol)-hexyl substituted polylactide (MPEGhexPLA) block copolymers was utilized to prepare clotrimazole (CLZ), econazole (ECN) and fluconazole (FLZ) aqueous micelle solutions. ECN loaded MPEG-dihexPLA micelles were synthesized by two methods: co-solvent evaporation and co-solvent evaporation sonication. Hydrodynamic diameters of the azole loaded micelles ranged from 70 to 165 nm with corresponding number weighted diameters between 30 to 40 nm. Ex vivo penetration studies were performed using either full thickness porcine ears and human breast skin. Full thickness porcine skin samples were equilibrated in 0.9% NaCl solution for 30 min. Human skin samples were obtained shortly after surgery and were used after removing fatty tissue, then mounted on standard Franz diffusion cells (area=2.0±0.1 cm2). The lowest loading efficiency in MPEG-dihexPLA micelles was observed for the most hydrophobic drug, CLZ (20%) and the highest for ECN (98%). FLZ was successfully encapsulated into the different micelles, with drug contents between 250 and 268 mg/g and the loading efficiencies of 83 to 91%. ECN showed similar high drug loadings and an even higher incorporation efficiency in MPEG-dihexPLA micelles. Therefore, hydrophobic dihexPLA core caused enhanced storage stability of the micelle formulations. ECN was selected as optimized drug with the highest loading efficiency. ECN skin delivery was compared to Pevaryl® cream (1% w/w ECN), a marketed liposomal formulation for topical application. ECZ skin deposition experiments from the MPEG-dihexPLA micelle formulation and Pevaryl® using human skin after 6 hours indicated that ECZ deposition from the micelle formulation was almost 7.5 fold higher than 17   

Pevaryl® (11.3 and 1.5 μg/cm2, respectively). No ECZ permeation was observed across the human skin. Comparing the human skin and the porcine skin samples indicated equivalent ECZ deposition following application of Pevaryl®. However, a 2-fold difference in ECZ deposition from the MPEG-dihexPLA micelle formulation was observed between porcine and human skin. Since, in contrast to the porcine ear skin, the human breast skin used for these experiments was devoid of hair follicles. Thus, higher ECZ deposition observed using the MPEG-dihexPLA micelle formulation was related to the penetration of the smaller micelles through the porcine hair follicles. The hydrophobic MPEG-dihexPLA copolymers encapsulated FLZ and ECN with high loading efficiencies. Preparation method influenced the micelle properties in such a way that smaller micelles were produced by co-solvent evaporation sonication method. Significant increase in ECN skin deposition using MPEGdihexPLA micelles demonstrated the potential of these particles to improve dermal drug bioavailability. ECN incorporated with an efficiency of 98% demonstrated 7.5-fold delivery ®

improvement and significant penetration enhancement compared to Pevaryl cream (Glujoy et al., 2014; Bachhav et al., 2011). However, the authors have not reported any skin toxicological effects for the synthesized nanoparticle and it mandates specific toxicological studies prior to a wider implementation. Yordanov et al., 2012 investigated the effect of two preparation methods; nanoprecipitation and emulsion polymerization on physicochemical properties of econazoleloaded poly(butyl cyanoacrylate) nanoparticles. Three non-ionic colloidal stabilizers Pluronic® F-68 (poloxamer 188), Tween 80 (polysorbate 80 Polyoxyethylene (20) sorbitan monooleate and Gentran®; LMD® (dextran 40) were used to obtain various nanoparticles surface coatings. The average size of ECN-PBCA nanoparticles prepared by polymerization method in the presence of Dextran 40 and Poloxamer188 was 120 nm, while nanoparticles prepared in the presence of P80 were slightly smaller, ∼111 nm. Nanoprecipitation approach 18   

using D40 and P188 resulted in larger size of ECN-PBCA nanoparticles (∼230 nm). Utilization of P80 led to slightly smaller particles (∼217 nm). The colloidal stability of ECNPBCA depended on the preparation method and the type of the colloidal stabilizer. The most stable formulations were formed using P188 and P80 by nanoprecipitation method. The colloids were stable at 4 ◦C for at least 6 months without any observable change in their physical state. D40 formulations were unstable and D40-coated particles aggregated in a few days of storage at room temperature or at 4 ◦C. The particles synthesized by polymerization method were less stable than those prepared by nanoprecipitation and coagulated in few months of storage at 4 ◦C. Nanoprecipitation method caused sediment formation upon storage, but could redisperse upon sonication. Higher concentrations of stabilizers could lead to more stable colloids, but may increase the toxicity risks of the formulations which are intended for biomedical uses (Yordanov et al., 2012). 3.5. Silica Nanoparticles (MCM-41) MCM-41, a kind of silica nanoparticles (MSN) with meso-pores sizes between 2–50 nm was first developed by researchers at Mobil Oil Corporation (Reichinger 2007). They have recently examined for delivery of active payloads based on physical or chemical adsorption (Scalia et al., 2013; Berlier et al., 2013a; Berlier et al., 2013b; Nafisi et al., 2014). In an attempt to prepare a topical powder for the treatment of fungal infections, ECN was melted with mesoporous silicate (MCM-41) (drug/MCM-41: 1/3). Mesopore volumes had an average dimension of 3.25nm. The synthesized ECN-MCM-41 formulations exhibited significant improvement in dissolution rate and much higher release rate which reached a value of 11% after 6 hours, almost four times higher than that from commercial product. MCM-41 was found to be very hygroscopic with high adsorbent characteristics comparing with commercial formulation. Higher antifungal activity against Candida Albicans was observed compared with the commercial product. Econazole included in MCM-41 pores 19   

could maintain its physical state for 30 days at 408ºC in a desiccator over CaCl2 (Ambrogi, et al., 2010). 4. ECN-Nanoparticles Stability Employing of nanotechnology in dermatology has created drastic improvement in cosmeceutical products, skin pharmaceutical drugs and the treatment of various skin diseases but despite the advantages of drug nanoparticle, there are still drawbacks in nanoparticles production including complex manufacturing, nanotoxicity and stability problems. Stability is an important issue in increasing the safety and efficacy of drug products and needs significant attention during pharmaceutical product development. Major stability alterations may occur during manufacturing (high temperature and pressure), storage and shipping (Eerdenbrugh et al., 2008; Patravale et al., 2004; Shah et al., 2014). General stability issues related to nanosuspensions have been widely studied and grouped as physical and chemical stability. Sedimentation or creaming, agglomeration, crystal growth and change of crystallinity state are the signs of common physical stability changes. For instance, drug particle size and size distribution may change when nanosuspensions are intravenously administered; therefore, size changes need to be closely monitored. High pressure or temperature produced during manufacturing can cause the crystallinity change of drug particles. Storage and shipping of drug products may cause sedimentation, agglomeration and crystal growth (Wang et al., 2013; Wu et al., 2011).

Yet, the results of this literature survey for the prepared

complexes of ECN-nanoparticles show that the stability information has not completely evaluated or very limited information is reported, so it certainly mandates more evaluation. Canu et al., 2006 reported that the average size of the synthesized ECN nanoparticles gel were constant and didn’t change within 30 days (Canu et al., 2006). Sanna et al., 2007 described that the prepared ECN-SLN nanoparticles exhibited no aggregation upon incorporation into hydrogels (Sanna et al., 2007). Yordanov and co-workers noted that the 20   

stability of the polymeric system depended on the type of the stabilizers and the preparation method; the most stable formulations using P188, P80 and nanoprecipitation method were stable for 6 months at 4 ˚C (Yordanov et al., 2012). Stability study on the econazole embedded into MCM-41 pores showed that the physical state of the synthesized complex was stable for 30 days at 408ºC in a desiccator over CaCl2 (Ambrogi et al., 2010). 5. Concluding Remarks Different nano-formulation strategies have been examined to optimize the topical delivery of ezonazole. Nano delivery systems such as lipid nanoparticles, nanoparticles and nanovesicular systems have been examined to improve ECN delivery. SLNs and NLCs could act as a reservoir and localized ECZ in the upper skin by penetrating into follicular openings and stratum corneum and provided a controlled release system. ECN permeation across epidermis was affected by the lipid content and fatty alcohols chain length in the lipid nanoparticle formulations. NLCs can be considered as promising vehicles for econazole topical delivery due to their high drug loading, smaller particle size. For vesicular system, ECN-liposome gel distribution was more intense on stratum corneum surface and CA blastospores comparing with commercially econazole formulations. Niosomes are preferred carriers comparing with liposomes, since they are more stable with higher entrapment efficiency. Yet, the results of this literature survey show that the data related to improving the ECN efficacy are few and is insufficient and defining the appropriate nano-formulation for use of econazole is currently not possible. The experimental data are focused on mainly on lipid nanoparticles and there are a few data on the other nanoparticles. The relationship among NPs physicochemical properties, and econazole absorbency, localization, and biological responses are not well understood. Moreover, only a few short-term in vivo studies of ECNNPs are available while data following long-term dermal application are still to be awaited. 21   

Using novel nanoparticles such as polymeric nanoparticles mandates skin toxicological studies before performing more experiments. Most of the experiments are limited to in vitro or animal studies. Complementary research is needed to translate these findings into clinical arena. Whilst the different working groups have tried to formulate econazole in their special carriers, it is suggested that the capability of various nanoparticles and factors affecting the efficiency of econazole delivery to be studied and compared in equal experimental condition. Acknowledgement: The authors appreciate Center for Research and Training in Skin Diseases and Leprosy, Tehran University of Medical Sciences and Janus Knowledge Based Company and IAUCTB, Tehran, Iran for support of this work and Hamid Reza Rahimi for his valuable helps in data collection. Disclosure Statement No competing financial interests exist.

22   

References Agharkar S, Lindenbaum S, Higuchi T. 1976. Enhancement of solubility of drug salts by hydrophilic counterions: Properties of organic salts of an antimalarial drug. J. Pharm. Sci.65:747–749. Allen, T.M., Cullis, P.R., 2013. Liposomal drug delivery systems: From concept to clinical applications, Adv. Drug Deliv. Rev. 65, 36–48. Ambrogi, V., Perioli, L., Pagano, C., Marmottini, F., Moretti, M., Mizzi, F., Rossi, C., 2010. Econazole nitrate-loaded MCM-41 for an antifungal topical powder formulation. J. Pharm. Sci. 99, 4738-4745. Andega, S., Kanikkannan, N., Singh, M., 2001. Comparison of the effect of fatty alcohols on the permeation of melatonin between porcine and human skin, J. Cont. Rel. 77, 17–25. Bachhav, Y.G., Mondon, K., Kalia, Y.N., Gurny, R., Möller, M., 2011. Novel micelle formulations to increase cutaneous bioavailability of azole antifungals. J. Cont. Rel. 153, 126–132. Berge, S.M., Bighley, L.D., Monkhouse, D.C. 1977. Pharmaceutical salts. J. Pharm. Sci. 66, 1–19. Bangham, A.D., Horne, R.W., Glauert, A.M., Dingle, J.T., Lucy, J.A., 1962. Action of saponin on biological cell membranes. Nature 196, 952–955. Bangham, A.D., Horne, R.W., 1964. Negative staining of phospholipids and their structural modification by surface-Active agents as observed in the electron microscope. J. Mol. Biol. 8, 660–668. Benson, H.A., 2006. Transfersomes for transdermal drug delivery. Expert Opin. Drug Deliv. 3, 727-737.

23   

Benson, H.A., 2010. Elastic liposomes for topical and transdermal drug delivery. Methods Mol. Biol. 605, 77-86. Berlier, G., Gastaldi, L., Ugazio, E., Miletto, I., Iliade, P., Sapino, S., 2013a. Stabilization of quercetin flavonoid in MCM-41 mesoporous silica: Positive effect of surface functionalization. J. Colloid Interface Sci. 393,109-18. Berlier, G., Gastaldi, L., Sapino, S., Miletto, I., Bottinelli, E., Chirio, D., Ugazio, E. 2013b. MCM-41 as a useful vector for rutin topical formulations: Synthesis, characterization and testing. Int. J. Pharm. 457, 177-186. Bunjes, H., Koch, M.H.J., Westesen, K., 2000. Effect of particle size on colloidal solid triglycerides. Langmuir 16, 5234–5241. Canu, G., Sanna, V., Gavini, E., Cossu, M., Rassu, G., Giunchedi, P., 2006. Ex vivo cutaneous penetration of econazole nitrate from SLN incorporated in hydrophilic gel, In: Sardinia Chem, giornata di studio dedicata alla chimica organica delle molecole biologicamente attive, 5 giugno, Cagliari, Italia. Cevc, G., Blume, G., Schatzlein, A., Gebauer, D., Paul, A., 1996. The skin: a pathway for the systemic treatment with patches and lipid based agent carriers. Adv. Drug Deliv. 18, 349-378. Cevc, G., Schätzlein, A., Richardsen, H., 2002. Ultradeformable lipid vesicles can penetrate the skin and other semi-permeable barriers intact. Evidence from double label CLSM experiments and direct size measurements. Biochim. Biophys. Acta. 1564, 21–30. Cevc, G., Schätzlein, A., Richardsen, H., Vierl U., 2003. Overcoming semi-permeable barriers, such as the skin, with ultradeformable mixed lipid vesicles, Transfersomes, liposomes or mixed lipid micelles. Langmuir 19, 10753–10763.

24   

Cevc, G., Gebauer, D., 2003. Hydration-driven transport of deformable lipid vesicles through fine pores and the skin barrier. Biophys. J. 84, 1010–1024. Cevc, G. 2004. Lipid vesicles and other colloids as drug carriers on the skin. Adv. Drug Deliv. Rev. 56, 675–711. Cheong, H.A., Choi, H.K. 2002. Enhanced percutaneous absorption of piroxicam via salt formation with ethanolamines, Pharm. Res. 19, 1375-1380. Choi, M.J., Maibach, H.I., 2005. Liposomes and niosomes as topical drug delivery systems. Skin Pharmacol Physiol. 18, 209–219. Degreef, H.J., De Doncker, P.R.G., 1994. Current therapy of dermatophytosis. J. Am. Acad. Dermatol. 31, S25-S30. Dubey, V., Mishra, D., Jain, N.K. 2007a. Melatonin loaded ethanolic liposomes: delivery. Eur. J. Pharm. Biopharm. 67, 398-405. Dubey, V., Mishra, D., Dutta, T., Nahar, M., Saraf, D.K., Jain, N.K., 2007b. Dermal and transdermal delivery of an anti-psoriatic agent via ethanolic liposomes. J. Cont. Rel. 123, 148-154. Econazole

Cream,

FDA

Prescribing

Information,

Available

at

[http://www.accessdata.fda.gov/drugsatfda_docs/label/2013/205175s000lbl.pdf]. El Maghraby, G.M., Williams, A.C., Barry, B.W., 2006. Can drug bearing liposomes penetrate intact skin? J. Pharm. Pharmacol. 58, 415–429. El Maghraby, G.M., Barry, B.W., Williams, A.C., 2008, Liposomes and skin: From drug delivery to model membranes. J. Pharm. Pharmacol. 58, 415–429. Eerdenbrugh, B.V., Mooter, G.V.D. Augustijns, P., 2008. Top-down production of drug nanocrystals: Nanosuspension stabilization, miniaturization and transformation into solid products. Int. J. Pharm. 364, 64–75.

25   

Gillet, A., Compère, P., Lecomte, F., Hubert, P., Ducat, E., Evrard, B., Piel, G., 2011. Liposome surface charge influence on skin penetration behaviour. Int. J. Pharm. 411, 223-231. Glujoy, M., Salerno, C., Bregni, C., Carlucci, A.D., 2014. Percutaneous drug delivery systems for improving antifungal therapy effectiveness: A Review. Int. J. Pharm. Pharm. Sci. 6, 8-16. Güngör, S., Erdal, M.S., Aksu, B., 2013. New formulation strategies in topical antifungal therapy. J. Cosmet. Dermatol. Sci. Appl. 3, 56-65. Heel, R.C., Brogden, R.N., Speight, T.M., Avery, G.S., 1978. Econazole: a review of its antifungal activity and therapeutic efficacy. Drugs 16, 177–201. Handjani-Vila, R.M., Rlbier, A., Rondot, B., Vanlerberghe, G., 1979. Dispersion of lamellar phases of non ionic lipids in cosmetic products. Int. J. Cosmetic Sci. 1, 303-314. Horne, R. W., Bangham, A. D., Whittaker, V. P., 1963. Negatively stained lipoprotein membranes. Nature 200, 1340. Imokawa, G., Surfactant mildness, in: Rieger, M.M., Rhein L.D., (Eds.), Surfactants in cosmetics. Surfactant Science Series, Marcel Dekker, New York, 1997, 427–471. Imbert, D., Wickett, R.R., 1995. Topical delivery with liposomes. Cosmet. Toiletries 110, 32–46. Jadon, P.S., Gajbhiye, V., Jadon, R.S., Gajbhiye, K.R., Ganesh, N. 2009. Enhanced oral bioavailability of griseofulvin via niosomes. AAPS. Pharm. Sci. Tech. 10, 1186–1192. Jain, N.K., Namdeo, A., 1996. Niosomes as Drug carriers. Ind. J. Pharm. Sci. 58, 41-46. Jain, S., Tiwary, A.K., Sapra, B., Jain, N.K., 2007. Formulation and evaluation of ethosomes for transdermal delivery of lamivudine. AAPS. Pharm. Sci. Tech. 8, 1-9.

26   

Jenning, V., Gysler, A., Schafer-Korting, M., Gohla, S. H. 2000. Vitamin A loaded solid lipid nanoparticles for topical use: occlusive properties and drug targeting to the upper skin. Eur. J. Pharm. Biopharm. 49, 211–218. Jung, S., Otberg, N., Thiede, G., Richter, H., Sterry, W., Panzner, S., Lademann, J. 2006. Innovative liposomes as a transfollicular drug delivery system: penetration into porcine hair follicles. J. Invest. Dermatol. 126, 1728–1732. Kakadia, P.G., Conway, B.R., 2014. Solid lipid nanoparticles: A potential approach for dermal drug delivery, Am. J. Pharmacol. Sci. 2, 1-7. Korting, H.C., Klövekorn, W., Klövekorn, G., 1997. Comparative efficacy and tolerability of econazole liposomal gel 1%, branded econazole conventional cream 1% and generic clotrimazole cream 1% in tinea pedis. Clin. Drug Invest. 14, 286-293. Kumar, Y.P., Kumar, K.V., Shekar, R.R., Ravi, M., Kishore, V.S., 2013. Formulation and evaluation of econazole niosomes. Sch. Acad. J. Pharm. 2, 315-318. Kumar, J.R., Muralidharan, S., Parasuraman, S., 2014. Antifungal agents: New approach for novel delivery systems. J. Pharm. Sci. Res. 6, 229-235. Labouta, H.I., El-Khordagui, L.K., Kraus. T., Schneider, M., 2011. Mechanism and determinants of nanoparticle penetration through human skin, Nanoscale, 3, 4989-4999. Lademann, J., Richter, H., Schanzer, S., Knorr, F., Meinke, M., Sterry, W., Patzelt, A., 2011. Penetration and storage of particles in human skin: Perspectives and safety aspects, Eur. J. Pharm. Biopharm. 77, 465–468. Lee, C., Uchida, T., Noguchi, E., Kim, N., Goto, S., 1993. Skin permeation enhancement of Tegafur by ethanol-panasate 800 or ethanol–water binary vehicle and combined effect of fatty acids and fatty alcohol. J. Pharm. Sci. 82, 1155–1159.

27   

Liang, X.W., Xu, Z.P., Grice, J., Zvyagin, A.V., Roberts, M.S., Liu, X. 2013. Penetration of nanoparticles into human skin . Curr. Pharm. Des. 19, 6353-6366. Lombardi, B.S., Regehly, M., Sivaramakrishnan, R., Mehnert, W., Korting, H.C., Danker, K., Röder, B., Kramer, K.D., Schäfer-Korting., M. 2005. Lipid nanoparticles for skin penetration enhancement-correlation to drug localization within the particle matrix as determined by fluorescence and parelectric spectroscopy. J. Cont. Rel. 110, 151–163. Müller, R.H., Radtke, M., Wissing, S.A., 2002. Solid lipid nanoparticles (SLN) and nanostructured lipid carriers (NLC) in cosmetic and dermatological preparations. Adv. Drug Deliv. Rev. 1, 54 Suppl 1, S131-S155. Müller, R.H., Mehnert, W., Lucks, J.S., Schwarz, C., Zur MuÈhlen, A., Weyhers, H., Freitas, C., RuÈhl, D., 1995. Solid lipid nanoparticles (SLN)-an alternative colloidal carrier system for controlled drug delivery. Eur. J. Pharm. Biopharm. 41, 62-69. Müller, R.H., Jenning, V., Mader, K., Lippacher, A., 2000. Solid-liquid (semi-solid) lipid particles and method of producing highly concentrated lipid particle dispersions. German patent application 199 45 203.2. Müller, R.H., Hildebrand, G.E., (Hrsg) 1998. Pharmazeutische Technologie: Moderne Arzneiformen, Lehrbuch für Studierende der Pharmazie - Nachschlagewerk für Apotheker

in

Offizin,

Krankenhaus

und

Forschung,

Wissenschaftliche

Verlagsgesellschaft Stuttgart, 2. erweiterte Aufl. 471 S. Müller, R.H., Mäder, K., Gohla, S., 2000. Solid lipid nanoparticles (SLN) for controlled drug delivery - a review of the state of the art. Eur. J. Pharm. Biopharm. 50, 161-177. Naeff, R., 1996. Feasibility of topical liposome drugs produced on an industrial scale. Adv. Drug Deliv. Rev. 18, 343–347. 28   

Nafisi, S., Schafer-Korting, M., Maibach, H.I., 2015. Perspectives on percutaneous penetration: Silica nanoparticles, Nanotoxicology, 9, 643-657. Neubert, R.H.H., 2011. Potentials of new nanocarriers for dermal and transdermal drug delivery. Eur. J. Pharm. Biopharm. 77, 1-2. Passerini, N., Gavini, E., Albertini, B., Rassu, G., Di Sabatino, M., Sanna, V., Giunchedi, P., Rodriguez, L., 2009. Evaluation of solid lipid microparticles produced by spray congealing for topical application of econazole nitrate. J. Pharm. Pharmacol. 61, 559567. Patravale, V.B., Date, A.A., Kulkarni, R.M. 2004. Nanosuspensions a promising drug delivery strategy. J. Pharm. Pharmacol. 56, 827–840. Patzelt, A., Richter, H., Knorr, F., Schäfer, U., Lehr, C.M., L. Dähne, L., Sterry, W., Lademann, J. 2011. Selective follicular targeting by modification of the particle sizes, J. Cont. Rel. 150, 45-48. Pawar, V., Naik, P., Giridhar R., Yadav M.R. 2015. Enhancing skin permeation of biphenylacetic acid (BPA) using salt formation with organic and alkali metal bases. Sci. Pharm. 83, 191–205. Piemi, M.P.Y., Korner, D., Benita, S., Marty, J.P., 1999. Positively and negatively charged submicron emulsion for enhanced topical delivery of antifungal drugs. J. Cont. Rel. 58, 177–187. Reichinger, M., Poröse Silikate mit hierarchischer Porenstruktur: Synthese von mikro/mesoporösem MCM-41 und MCM-48 Materialien aus zeolithischen Baueinheiten des MFI-Gerüststrukturtyps, Dissertation Ruhr-Universität Bochum. Rojanasakul, Y., Wang, L.Y., Bhat, M., Glover, D.D., Malanga, C.J., Ma, J.K. 1992. The transport barrier of epithelia – a comparative study on membrane-permeability and charge selectivity in the rabbit. Pharm. Res. 9, 1029-1034. 29   

Rosen, M.J., Kunjappu, J.T., 2012. Surfactants and interfacial phenomena, 4th edition 2012, Wiley & Sons, New Jersey. Sanna, V., Gavini, E., Cossu, M., Rassu, G., Giunchedi, P., 2007. Solid lipid nanoparticles (SLN) as carriers for the topical delivery of econazole nitrate: in vitro characterization, ex vivo and in vivo studies. J. Pharm. Pharmacol. 59, 1057-1064. Sanna, V., Caria, G., Mariani, A., 2010. Effect of lipid nanoparticles containing fatty alcohols having different chain length on the ex vivo skin permeability of econazole nitrate. Powder Technol. 201, 32–36. Sarmah, P.J., Kalita, B., Sharma, A.K., 2013. Transferosomes based transdermal drug delivery: An overview. Int. J. Adv. Pharm. Res. 4, 2555-2563. Scalia, S., Franceschinis, E., Bertelli, D., Iannuccelli, V., 2013. Comparative evaluation of the effect of permeation enhancers, lipid nanoparticles and colloidal silica on in vivo human skin penetration of quercetin. Skin Pharmacol. Physiol. 26, 57-67. Schaller, M., Preidel, H., Januschke, E., Korting, H.C., 1999. Light and electron microscopic findings in a model of human cutaneous Candidosis based on reconstructed human epidermis following the topical application of different econazole formulations. J. Drug Target, 6, 361-372. Schlupp, P., Blaschke, T., Kramer, K.D., Höltje, H.D., Mehnert, W., Korting, M.S., 2011. Drug release and skin penetration from solid lipid nanoparticles and a base cream: a systematic approach from a comparison of three glucocorticoids. Skin Pharmacol. Physiol. 24, 199-209. Schreier, H., Bouwstra, J., 1994. Liposomes and niosomes as drug carriers: dermal and transdermal drug delivery. J. Cont. Rel. 30, 1–15. Shah, R., Eldridge, D., Palombo, E., Harding, I., 2014. Lipid nanoparticles: Production, characterization and stability, Springer. 30   

Sinico, C., Manconi, M., Peppi, M., Lai, F., Valenti, D., Fadda, A.M., 2005. Liposomes as carriers for dermal delivery of tretinoin: In vitro evaluation of drug permeation and vesicle–skin interaction. J. Cont. Rel. 103, 123–136. Sloan, K.B., Beall, H.D., Taylor, H.E. 1998. Transdermal delivery of theophilline from alcohol vehicles. Int. J. Pharm. 171, 185–193. Souto, E.B., Wissing, S.A., Barbosa, C.M., Müller, R.H., 2004. Development of a controlled release formulation based on SLN and NLC for topical clotrimazole delivery. Int. J. Pharm. 278, 71–77. Szoka, F., Jr., Papahadjopoulos, D., 1980. Comparative properties and methods of preparation of lipid vesicles (liposomes). Annu. Rev. Biophys. Bioeng. 9, 467–508. Thienpont, D., Van Cutsem, J., Van Nueten, J.M., Niemegeers, C.J., Marsboom, R., 1975. Bilogical

and

toxicological

properties

of

econazole,

a

broad-spectrum

antimycotic. Arzneimittel-Forschung. 25, 224–230. Torchilin, V.P., 2005. Recent advances with liposomes as pharmaceutical carriers. Nat. Rev. Drug Discov. 4, 145-160. Torchilin, V.P., 2006. Multifunctional nanocarriers. Adv. Drug Deliv. Rev. 58, 1532–1555. Touitou, E., Dayan, N., Bergelson, L., Godin, B., Eliaz, M., 2000. Ethosomes-novel vesicular carriers for enhanced delivery: characterization and skin penetration properties J. Cont. Rel. 65, 403-418. Touitou, E., Godin, B., Dayan, N., 2001. Intracellular delivery mediated by ethosomal carrier. J. Biomater. 22, 3055-3059. Vanlerberghe, G., Morancias, J.L., Niosomes in perspective, S.T.P. Pharma Sci. 1996, 6, 511.

31   

Wang, Y., Zheng, Y., Zhang, L., Wang, Q., Zhang, D., 2013. Stability of nanosuspensions in drug delivery. J. Cont. Rel. 172, 1126-1141. Wissing, S.A., Kayser, O., Muller, R.H. 2004. Solid lipid nanoparticles for parenteral drug delivery. Adv. Drug Deliv. Rev. 56, 1257–1272. Wissing, S.A., Muller R.H., 2001. A novel sunscreen system based on tocopherol acetate incorporated into solid lipid nanoparticles. Int. J. Cosm. Sci. 23, 233-243. Wu, L., Zhang, J., Watanabe. W., 2011. Physical and chemical stability of drug nanoparticles. Adv. Drug Deliv. Rev. 63, 456–469. Yang, W., Wiederhold, N.P., Williams R.O. 3rd., 2008. Drug delivery strategies for improved azole antifungal action. Expert Opin. Drug Deliv. 5, 1199-1216. Yilmaz, E., Borchert, H.H., 2005. Design of a phytosphingosine-containing, positivelycharged nanoemulsion as a colloidal carrier system for dermal application of ceramides. Eur. J. Pharm. Biopharm. 60, 91–98. Yordanov, G., 2012. Influence of the preparation method on the physicochemical properties of econazole-loaded poly (butyl cyanoacrylate) colloidal nanoparticles. Colloid Surf. A: Physicochem. Eng. Asp. 413, 260-265.

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Figure Captions

Figure 1: Chemical structure of econazole, miconazole and isoconazole

33   

Tables Table 1: Summary of the novel formulation approaches of econazole using nanoparticles Research Question SLN-ECZ dispersions (480 nm): SLN1(PCR: 5% w/w), SLN2 (PCR:10% w/w), gelling agent: HPMCK100M 2.0% w/w, lipid carrier: Precirol ATO (PCR), emulsifying agent: Tween 80, ECZ final Conc:1.0% w/w - encapsulation efficiency - permeation study - stability ECZ-SLN: lipid/drug (5/1, 10/1), incorporated into hydrogels (150 nm) - encapsulation efficiency - release study - permeation study ECZ-SLN: different chain length (C12– C18) of saturated fatty alcohols (200 nm) - encapsulation efficiency - permeation study SLMs (18–45 µm), SLNs (130–270 nm). lipid:drug (5:1, 10:1, 12.5:1) lipidic carrier: Precirol ATO 5

Characterizati on Method modified USP apparatus

Experiments

Results

Ref

ex-vivo: porcine skin

- encapsulation efficiency: 96-102% - SLN1 release rate two fold higher than SLN2 - higher lipid content led to slower release rate - stable over 30 days

Canu et al, 2006

- SEM - differential scanning calorimetry - dissolution apparatus

ex-vivo: porcine skin (adult pig ears); human skin in-vivo: human (inner forearm) ex vivo: porcine skin

- SLN could control drug release through SC - ECZ release rate depended on NPs lipid content - SLN (5:1) improved ECZ penetration into upper skin - SLN increased ECZ penetration through SC after 1 h, improved drug diffusion in deeper skin layers after 3 h compared with conventional gel

Sanna et al 2007

- encapsulation efficiency; 95- 98% - no direct relationship between lipid composition of NPs and drug loading - drug flux increased as the alcohol chain length increased - spray congealing, good method for SLMs-ECZ production - high-shear homogenization, good method for SLNs-

Sanna et al. 2010

- differential scanning calorimetry -SEM - X-Ray photoelectron

ex-vivo: pig skin

34   

Passerin i et al 2009

SLMs, SLNs incorporated into HPMC K 100M hydrogels; Production methods: spray congealing process, high-shear homogenization - encapsulation efficiency - permeation study Comparing ECZ nitrate cream and ECZ liposome gel on uninfected RHE, CC - skin morphological alterations

microscopy - dissolution apparatus

- light microscopy - electron microscopy

Comparing treatment by ECZ-liposome gel%, branded ECZ cream 1 % and generic clotrimazole cream 1 % in one controlled, double-blinded trial ECZ-niosomes gel Preparation method: thin Film hydration; cholesterol/span80 (1/1, 1/2, 1/3, 1/4; A1 to A4 orderly) - ECZ-loaded poly(butyl cyanoacrylate) NPs ; stabilizers: poloxamer 188, polysorbate 80, dextran 40 - effect of preparation method on NPs

- SEM - Light Scattering

ECN preparation - encapsulation efficiency: 80-100% - permeation rate depended on particle size, lipid amount - SLNs can act as carrier - SLMs can be used for controlled delivery of drugs to skin - Uninfected RHE - infected RHE by Candida Albicans - human cutaneous candidosis (CC)

ECZ-liposome gel - reduced epidermal barrier damage - reduced toxic effects of lower epidermis - active agent reached to deeper parts of skin - reduced signs of CA-specific damage to CC - pronounced damage to CA blastospores - more intense distribution on SC & CA blastospores

in vivo: human skin

- higher cure rate for ECZ-liposome gel treatment group Korting - tolerability slightly better in ECZ-liposome gel group et al., compared with ECZ cream, clotrimazole cream treatment 1997 groups

-

A4 showed - highest entrapment percentage - desired sustained release - zero order release pattern -ECZ-PBCA NPs; - polymerization method: 120 nm (using D40 and P188) ∼111 nm (using P80) 35 

 

Schaller et al., 1999

Kumar et al., 2013 Yordano v et al, 2012

size Preparation methods: nanoprecipitation, emulsion polymerization

- nanoprecipitation method: 230 nm (using D40 and P188); ∼217 nm (using P80)

- drug content - drug loading

Fabrication of CLZ, ECZ, FLZ using MPEG-dihexPLA, MPEG-monohexPLAECZ, MPEG-PLA (70-165 nm) - ECZ-MPEGdihexPLA micelles (40 nm) compared with Pevaryl® cream (1% w/w ECZ) - loading efficiency - drug deposition - skin deposition

- TEM - CLSM - Franz diffusion cell

- ex-vivo: porcine skin, human skin

- colloidal stability depended on preparation method, and stabilizer - most stable colloids formed with P188 &P80 by nanoprecipitation method - D40 formulations were unstable, polymerization method caused less stable particles - higher stabilizer Conc gave more stable colloids - CLZ (most hydrophobic): lowest loading efficiency (20%) - ECZ loading efficiency: 98.3% in MPEG- dihexPLA micelles - high incorporation efficiencies for ECZ, FLZ hydrophobic MPEG-dihexPLA - ECZ deposition in porcine skin and human skin following 6 h application using MPEG-dihexPLA micelles : orderly >13, 7.5-fold higher than Pevaryl® cream

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Bachnav et al., 2011