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Nile red nanosuspensions as investigative model to study the follicular targeting of drug nanocrystals
Abstract
The strategy of formulating poorly soluble actives as nanosuspension has been explored by more than a thousand research papers, with some medicinal products for oral and intravenous use having reached the market or advanced clinical trials. Interestingly, there is a limited number of reports of nanosuspensions formulated for dermal and transdermal drug delivery. In the present work, a nanocrystals suspension of the fluorescent, water-insoluble dye Nile Red, is prepared through a media milling technique and exploited to characterize the fate of the nanosuspended drug when applied on the skin. More in detail, the accumulation of Nile Red nanocrystals inside the hair follicles is evidenced by scanning electron microscopy, and the diffusion of drug molecules in the different skin layers is evaluated by confocal microscopy and skin permeation studies. Overall, the combination of the analytical techniques provide a description of the mechanisms underlying dermal accumulation, and transdermal penetration of a drug formulated as a nanosuspension.
... Pilot studies have been performed to unravel the intradermal fate of nanosuspensions (Corrias et al., 2017;Li et al., 2018;Vidlářová et al., 2016). Scanning electron microscopy (SEM) was used to identify drug nanoparticles in the dermal tissues (Corrias et al., 2017;Li et al., 2018). ...
... Pilot studies have been performed to unravel the intradermal fate of nanosuspensions (Corrias et al., 2017;Li et al., 2018;Vidlářová et al., 2016). Scanning electron microscopy (SEM) was used to identify drug nanoparticles in the dermal tissues (Corrias et al., 2017;Li et al., 2018). However, the sample process such as dehydration and dyeing/gold sputtering may confuse drug nanoparticles with backgrounds. ...
... Besides, SEM only provides static viewing and lacks the capability for dynamic monitoring. Another strategy depends on auto fluorescent drugs such as curcumin (CUR) and nile red (Corrias et al., 2017;Vidlářová et al., 2016). The intention is to track the transportation of nanosuspensions by the fluorescence of these drugs but fails to discriminate the intact drug particles from dissolved molecules. ...
Nanosuspensions have received much attention in enhanced transdermal delivery. However, the corresponding mechanisms have not been clarified. In particular, whether nanosuspensions can directly penetrate across the stratum corneum (SC) and what is the transdermal route for the enhanced penetration. Therefore, curcumin (CUR) was adopted in this study as a model drug, while an aggregation-caused quenching (ACQ) probe was physically embedded in CUR nanosuspensions, i.e., the CUR hybrid nanosuspensions (CUR-HNSs), for bioimaging. The ACQ properties enable identification of intact CUR-HNSs. The co-localization of particle and CUR signals was exploited to outline the translocation profiles of intact nanosuspensions as well as the cargoes. Three sizes of CUR-HNSs are prepared, which are spherical and amorphous. CUR is poor in transdermal transport even in propylene glycol solution, which was enhanced by nanosuspensions. Although 400 nm CUR-HNSs present higher steady state flux than 140 nm and 730 nm ones, the cumulative amount of permeated CUR is yet less than 2 % of the applied dose at 12 h. Co-localization of CUR and ACQ probe signals indicates that CUR-HNSs can infiltrate into the SC layer and accumulate in the hair follicles. The intact CUR-HNSs cannot enter into the skin. On the contrary, CUR molecules diffuse into the whole skin tissues following dissolution of CUR-HNSs in the SC and the hair follicles. In conclusion, nanosuspensions are advantageous for transdermal delivery of poor permeable drugs by filtrate into the SC and accumulate in hair follicles.
... They were compared to those with a mean size higher or lower than this range [13]. Although the correct mean size for the best transfollicular permeation or penetration is still under discussion [6], nanostructured systems are very promising, since they are generally smaller than the size of the follicular openings [14]. ...
... Several experiments have evaluated the affinity of nanocrystals with hair follicles. Corrias et al. [14] prepared a nanocrystal of water-insoluble dye, Nile Red, in combination with Polysorbate 80 or Poloxamer 188 as a stabilizer. The 230-nm nanocrystal was made using the media milling technique. ...
... The 230-nm nanocrystal was made using the media milling technique. The in vitro skin permeation studies have demonstrated the accumulation of Nile Red nanocrystal in the follicular duct [14]. Another hair follicle location study was performed with curcumin nanocrystal prepared by the smartCrystals process. ...
This chapter reviews the use of nanostructured systems for treating hair disorders. Hair is an intriguing organ, which involves complex and dynamic interactions, such as cell lifecycles and their impact from external and internal damage. Hair appearance reflects the social lifestyles and wellness of individuals, which causes concerns once this visual aspect is negatively affected. Understanding the primary sources of each disorder is essential for the best treatment, although several causes are still being studied. In the first section, we describe the structure of different parts of hair. The second section provides in-depth coverage of several types of nanostructured systems. Lastly, the third section reports on the main hair disorders and recent research based on nanostructured systems addressing these specific needs and treatments.
... More detailed investigations and systemic studies are required to prove the effect of humectant on hair follicle targeting. Corrias et al. [70] studied follicular targeting of fluorescent Nile red nanosuspension using scanning electron microscopy (SEM), which revealed that nanocrystals filled up the infundibulum and that the interior of hair follicle was surrounded by numerous nanocrystals. The distribution of curcumin nanocrystals within the porcine skin was studied using confocal laser scanning microscopy (CLSM), which revealed localization of curcumin in the hair follicle as compared to micron-sized curcumin particles [71]. ...
... Vidlá rová et al. [71] studied the distribution of curcumin nanocrystals within porcine skin using CLSM, showing pronounced penetration of nanosuspensions and gels comprising nanocrystals across the SC, epidermis up to its basal membrane and hair follicle. Corrias et al. [70] studied the diffusion of fluorescent Nile red nanosuspension into different skin layers using CLSM, demonstrating intense fluorescence even in lower skin layers at 8 h and 24 h indicating passive diffusion in skin as compared to untreated skin. Lohan et al. [50] investigated the penetration of dexamethasone nanocrystals into porcine skin using EPR spectroscopy, which showed 3-fold and 2.1-fold higher drug concentrations in the VE of intact skin and barrier-disrupted skin, respectively, over base cream formulation. ...
The barrier function of skin and the non-optimal physicochemical properties of drugs present challenges to the skin penetration of many drugs, motivating the development of novel drug-delivery systems. Recently, nanocrystal-based formulations have been investigated for topical drug delivery and have demonstrated improved skin penetration. This review highlights barriers in skin penetration, current techniques to improve topical delivery and the application of nanocrystals to conquer obstacles for topical delivery. Nanocrystals can improve delivery through the skin by mechanisms including the creation of a higher concentration gradient across skin resulting in increased passive diffusion, hair follicle targeting, formation of diffusional corona, and adhesion to skin. These mechanisms are of interest for formulation scientists seeking to develop products involving molecules that are ‘difficult-to-deliver’ topically.
... For visualization of the pores/microchannels produced by the DermaQ treatment, porcine skin samples were perforated as described in Section 2.7 and then stained for 3 min with a strongly colored nanocrystals suspension prepared with a water-insoluble fluorescent dye, Nile Red [21]. Following treatment, skin samples were immediately rinsed with saline to remove excessive dye. ...
... In this experiment, pig skin specimens were treated with a nile red nanosuspension after the first, second, third and fourth applications of the roller in the directions described in the figure (left panel). Nile red was chosen because it can be formulated as nanosuspensions, and it can be easily visualized due to its intense red color [21]. Thus, using this simple setup, it was possible to have a preliminary, qualitative information about the extent of nanocrystals accumulation in the skin pores created by the microneedle rollers. ...
Topical application of the anti-inflammatory drug diclofenac (DCF) reduces the severity of systemic unwanted effects compared to its oral administration. A number of transdermal formulations are available on the market and routinely used in clinical and home-care settings. However, the amount of DCF delivered across the skin remains limited and often insufficient, thus making the oral route still necessary for achieving sufficient drug concentration at the inflamed site. In attempting to improve the transdermal penetration, we explored the combined use of DCF nanosuspensions with a microneedle roller. Firstly, DCF nanosuspensions were prepared by a top-down media milling method and characterized by spectroscopic, thermal and electron microscopy analyses. Secondly, the pore-forming action of microneedle rollers on skin specimens (ex vivo) was described by imaging at different scales. Finally, DCF nanosuspensions were applied on newborn pig skin (in vitro) in combination with microneedles roller treatment, assessing the DCF penetration and distribution in the different skin layers. The relative contribution of microneedle length, nanosuspension stabilizer and application sequence could be identified by systemically varying these parameters.
... With increasing interest for nanosuspensions formulation during early phases of drug development, a lot of effort is put to miniaturize production process (Frank and Boeck, 2016;Van Eerdenbrugh et al., 2008). One of the approaches is using cell disruptor device, with yttrium-stabilized zirconium beads as milling media (Corrias et al., 2017). ...
... 3.2.1. Particle size analysis, zeta potential measurements and stability study -Nanosuspensions of DK-I-56-1 stabilized by different steric stabilizers and their combinations were prepared by small scale media milling technique similar to protocol explained by Corrias et al. (2017). Parenteral route of administration limits number of stabilizers that can be used, so typical non-ionic stabilizers such as polysorbate 80 and poloxamers were chosen. ...
DK-I-56-1 (7-methoxy-2-(4-methoxy-d3-phenyl)-2,5-dihydro-3H-pyrazolo[4,3-c]quinolin-3-one), a recently developed deuterated pyrazoloquinolinone, has been recognized as a lead candidate for treatment of various neuropsychiatric disorders. During preclinical investigation of poorly water-soluble compounds such as DK-I-56-1, the application of nanotechnology could be advantageous due to improved safety and possibly increased bioavailability of nanosized formulation. DK-I-56-1 nanosuspensions stabilized by polysorbate 80, alone or in combination with poloxamers 188 i.e. 407 or d-α-tocopheryl polyethylene glycol 1000 succinate, were prepared using a small-scale media milling device. With particle size 208.7–250.6 nm and polydispersity index <0.250, selected nanodiseprsions were stable for three weeks. Pharmacokinetic and biodistribution studies following intraperitoneal administration of three types of formulation in mice indicated high plasma DK-I-56-1 levels after solution (10228.6 ± 1037.2 ngh/ml) and nanosuspension (6770.4 ± 770.7 ngh/ml) but not suspension administration (966.0 ± 58.1 ngh/ml). However, distribution of DK-I-56-1 after solution was heavily influenced by its composition, and brain availability of nanosuspension was superior to that of solution formulation. In spontaneous locomotor activity test, the expected hyperlocomotor effect was observed after nanosuspension administration, without compromising impact of the vehicle/excipients used. Therefore, nanonization of drug compound assembled with proper selection of stabilizers may seemingly contribute further thorough testing of DK-I-56-1 preclinical efficacy.
... To further elucidate the permeation mechanism of NLC-chitosan hydrogel beads, Nile red, a hydrophobic fluorescent probe, was encapsulated into the lipid phase of NLC. Nile red has been extensively employed to investigate the permeation of lipid-based nanocarriers, as it can visually demonstrate the permeation profile [46][47][48][49][50]. It is worth noting that the distribution of Nile red and quercetin in the skin might not be identical due to their distinct physicochemical properties, including solubility and logP. ...
The objective of the present study was to develop and evaluate NLC–chitosan hydrogel beads for topical administration. The feasibility of the preparation technology was verified by investigating various formulation factors and the impact of chitosan hydrogel beads on the NLC. The encapsulation efficiency of NLC–chitosan hydrogel beads was above 95% in optimized process conditions. The physical characterization of the NLC–chitosan hydrogel beads showed that the NLC was distributed within the network of the chitosan hydrogel beads. Furthermore, the incorporation of NLC into the chitosan hydrogel beads was related to the electrostatic interaction between the surface of the NLC and chitosan, which influenced the lipid ordering degree of the NLC and contributed to the stability. The stability studies showed that the retention rate of quercetin in the NLC–chitosan hydrogel beads was 88.63 ± 2.57% after 10 months of storage under natural daylight. An in vitro permeation study showed that NLC–chitosan hydrogel beads exhibited superior ability in enhancing skin permeation by hydrophobic active ingredients compared to the NLC and significantly increased skin accumulation. These studies demonstrated that the use of NLC–chitosan hydrogel beads might be a promising strategy for the delivery of hydrophobic active ingredients in topical administration.
... It decreases medication particle size to the nanoscale range and improves the solubility, rate of dissolution, and bioavailability of components that are not very water soluble. There are two ways to make nanosuspension: top-down and bottom-up, and they differ in terms of the manufacturing concept [16,17]. Top-down methodology used media milling and high-pressure homogenization to break up bigger drug particles, microparticles, and nanoparticles. ...
Angiotensin converting enzyme (ACE) overactivation is one of the primary causes of hypertension, which leads to cardiovascular disorders all over the world. In the scientific world, nanosuspension is a novel area of study that could offer an alternative treatment for active pharmaceuticals that are not well soluble in water. Since active compounds’ bioavailability is reduced by their poor solubility, there are eventually fewer applications. Drug solubility, dissolving rate, and bioavailability are improved by nanosuspension, which shrinks medication particle size into the nanoscale range and boosts the surface area to volume ratio of the drug. There is a need to prepare Rauvolfia serpentina’s nanosuspension in order to get around some of the major challenges that it faces because of its poor solubility and wide range of biological activities. Using the antisolvent precipitation approach, a nanosuspension of Rauvolfia serpentina was created with hydroxy propyl methyl cellulose (HPMC). Rouvolfia serpentina nanosuspensions were prepared using a design of expert (DOE) approach, which allowed for the evaluation of key process parameters. To get an optimal sample, the effects of stabilizer concentration and anti-solvent volume on particle size, zeta potential, and PdI using CCD-RSM were investigated. Using the substrate Hippuryl-histidyl-leucine, the in vitro ACE inhibitory potential was assessed. On human erythrocytes, the safety of nanosuspension was evaluated in vitro. The ideal value of independent variables was discovered to be 0.25% w/v in order to achieve the desired response. Using scanning electron microscopy, the morphology of optimized nanosuspension was discovered to be rod-shaped (SEM). Compared to nanoformulation, crude extract had higher ACE inhibitory potential (83.11%). Human erythrocytes were found to be unaffected by nano-sized particles.
... The region where the barriers are not fully developed and thus more permeable, which enables the passive diffusion of drug substances to the surrounding skin tissue. Corrias et al. developed a suspension of nile red nanocrystals (230 nm) and examined their fate when topically applied on newborn pig skin (stored at − 80 • C and pre-treated with a physiological solution before the experiments) (Corrias et al., 2017). They evidenced that the nile red nanocrystals were accumulated into the infundibular region of the HF and in the surrounding skin tissue (dermis and epidermis), which the authors hypothesized as passive diffusion of drug molecules from the HF into the different skin layers. ...
The importance of the hair follicle in the process of cutaneous drug penetration has been established since this skin appendage was recognized as an entry point for topically applied substances. A comprehensive review on the hair follicle as a target per se is here provided, exploring the current knowledge on both targeted regions and delivery systems that take advantage of this permeation route.
The follicular penetration is a complex process, whose effectiveness and efficiency strongly depends on a diversity of different factors, including follicular density and size, activity status of hair follicles and physicochemical properties of the topically applied substances. Nanocarriers represent a heterogeneous assembly of molecules organized into particles and they have revolutionized drug delivery in several areas of medicine, pharmacology and cosmetics. As they possess an inherent ability to use the follicular route, they are reviewed here having in perspective the hair follicle zones that they are able to reach as reported. In this way, a follicular road map for the different delivery systems was compiled to assist as a guiding tool for those that have interest in the development and/or application of such delivery systems for hair and skin treatment or care.
... In vitro skin delivery studies were performed under nonocclusive conditions using vertical Franz diffusion cells (diffusion area 0.785 cm 2 ) and newborn pig skin. Oneday-old Goland-Pietrain hybrid pigs (about 1.2 kg) were provided by a local slaughterhouse [20]. The skin (n = 6 per formulation), stored at −80 C, was pre-equilibrated in saline solution at 25 C for 12 hours and then sandwiched between donor and receptor cells. ...
8-methoxypsoralen is the most common drug in psoralen plus ultraviolet light irradiation therapy for the treatment of severe psoriasis. Despite of the efficacy, its classic oral administration leads to several serious adverse effects. However, the topical psoralen application produces a drug skin accumulation lower than that obtained by oral administration, due to the drug low skin permeability. In this paper, 8-methoxypsoralen loaded Penetration Enhancer-containing Vesicles were prepared using soy phosphatidylcholine and the penetration enhancer Transcutol® (5% or 10%) and characterized in terms of size, polydispersity index, zeta potential and encapsulation efficiency. No statistically significant differences in both size (~135 nm) and encapsulation efficiency (~65%) were found for different Transcutol® concentration. Transdermal delivery study assessed by Franz diffusion cells, showed that the 8-methoxypsoralen mainly accumulated into the stratum corneum. Moreover, after Penetration Enhancer-containing Vesicles application, the drug recovered in this layer is almost double of that delivered by conventional liposomes, while no significant difference was found from the different Transcutol® concentrations. Finally, biocompatibility checked by an MTT assay, demonstrated that the incubation of human keratinocytes for 24 h with 8-methoxypsoralen loaded Penetration Enhancer-containing Vesicles did not significantly reduce cell viability.
... The region where the barriers are not fully developed and thus more permeable, which enables the passive diffusion of drug substances to the surrounding skin tissue. Corrias et al. developed a suspension of nile red nanocrystals (230 nm) and examined their fate when topically applied on newborn pig skin (stored at − 80 • C and pre-treated with a physiological solution before the experiments) (Corrias et al., 2017). They evidenced that the nile red nanocrystals were accumulated into the infundibular region of the HF and in the surrounding skin tissue (dermis and epidermis), which the authors hypothesized as passive diffusion of drug molecules from the HF into the different skin layers. ...
The follicular route is an important drug penetration pathway in any topical application, either concerning dermatological and cosmetic skin treatments or any transdermal administration regimen. Efficient transport into follicles will depend on drug inherent properties but also on the chosen vehicle. The main study goal was to compare several systems for the delivery to the hair bulb of two fluorescent molecules of different water affinities: the hydrophobic Nile Red and the quite similar but hydrophilic Nile Blue. Three common nanoparticle types were compared in terms of encapsulation efficiency and stability: liposomes, ethosomes and polymeric nanoparticles. A liquid serum-like formulation was also developed, adjusting the final ethanol amount to the type of dye to be solubilized. Then, this formulation and the nanoparticle systems that successfully passed characterization and stability stages were further studied on their ability to reach the bulb. The serum formulation was able to deliver, both drug models, to deeper follicular regions than nanoparticles. Attending to the envisioned zone target of the follicle, the simplest approach proved to be the best choice from all the systems tested in this work. Nonetheless, nanocarriers and the inherent complexity of their manufacturing processes may be justified under very specific requirements.
... 10 In the early 1990s, nanosuspension technology was developed to overcome low bioavailability of drugs showing poor water solubility, and was adopted in different pharmaceutical fields, for oral, parenteral, dermal, pulmonary and ocular administration. [11][12][13][14][15][16][17][18] Nanosuspensions are sub-micron colloidal dispersions of pure active particles (nanocrystals) stabilized by surfactants, polymers or a mixture of both. 19 Nanosuspensions can be prepared using two different approaches: the bottom-up and the top-down techniques or a combination of the two. ...
BACKGROUND
Low water solubility of pesticide requires formulations with high levels of stabilizers and organic solvents. Moreover, only 0.1% of the applied pesticides formulation reaches the target, while 99.9% spreads in the surrounding environment. Therefore, there is the need for more efficient and environmentally sustainable alternatives.
RESULTS
Zoxamide (ZO) nanosuspension was prepared through a media milling technique by using the stabilizer polysorbate 80. The thin and acicular crystals obtained, showed particle size of 227 nm, polydispersion index of 0.247 and zeta potential of −28 mV. Dimensional data and morphology of ZO nanocrystals alone, on tomato leaves and berries, were confirmed by scanning electron microscopy. The reduction in size for ZO crystals obtained after the milling process increased pesticide water solubility till 39.6 mg L⁻¹, about 1.6 the solubility obtained with a conventional commercial formulation. Field and dip contamination trials performed on tomato plants showed the nanosuspension's ability to increase ZO deposition and accumulation versus a coarse ZO suspension and commercial formulation, respectively.
CONCLUSIONS
The nanoformulation proposed, resulted in low cost and was easy to make. Moreover, the organic solvent‐free composition together with a low surfactant addition assured a minor environmental impact. Finally, the increased retention and deposition of the fungicide can reduce the amounts of ZO formulation applied to tomatoes. © 2021 Society of Chemical Industry.
... With the aim of fully understanding the performance of drug nanocrystals, their intracellular fate or in vivo fate have been explored by tracing nanocrystals with autofluorescence [27][28][29][30] or tracking fluorescently hybridized nanocrystals [31][32][33][34][35][36][37] . The fluorescent hybrid nanocrystals seem to be a promising tool with potential for tracking J o u r n a l P r e -p r o o f translocation of nanocrystals in biological environment. ...
In this study, self-discriminating hybrid nanocrystals was utilized to explore the biological fate of quercetin hybrid nanocrystals (QT-HNCs) with diameter around 280 nm (QT-HNCs-280) and 550 nm (QT-HNCs-550) following oral and intravenous administration and the contribution of integral nanocrystals to oral bioavailability enhancement of QT was estimated by comparing the absolute exposure of integral QT-HNCs and total QT in the liver. Results showed that QT-HNCs could reside in vivo as intact nanocrystals for as long as 48 h following oral and intravenous administration. A higher accumulation of integral QT-HNCs in liver and lung was observed for both oral and intravenous administration of QT-HNCs. The particle size affects the absorption and biodistribution of integral QT-HNCs and total QT. As compared to QT-HNCs-550, QT-HNCs-280 with smaller particle size is more easily absorbed, but dissolves faster in vivo, leading to higher distribution of QT (146.90 vs. 117.91 h·μg/mL) but lower accumulation of integral nanocrystals (6.8 2e10 vs. 15.27e10 μW/cm²) in liver following oral administration. Due to its slower dissolution and enhanced recognition by RES, QT-HNCs-550 with larger diameter shows higher liver distribution for both of QT (1015.80 h·μg/mL) and integral nanocrystals (259.63e10 W/cm²) than those of QT-HNCs-280 (673.82 & 77.66e10 W/cm²) following intravenous administration. The absolute exposure of integral QT-HNCs in liver following oral administration of QT-HNCs are 8.78% for QT-HNCs-280 and 5.88% for QT-HNCs-550, while the absolute exposure of total QT for QT-HNCs-280 and QT-HNCs-550 are 21.80% and 11.61%, respectively. Owing to imprecise quantification method, a surprisingly high contribution of integral QT-HNCs to oral bioavailability enhancement of QT (40.27% for QT-HNCs-280 and 50.65% for QT-HNCs-550) was obtained. These results revealed significant difference in absorption and biodistrbution between integral nanocrystals and overall drugs following oral and intravenous administration of QT-HNCs, and provided a meaningful reference for the contribution of integral nanocrystals to overall bioavailability enhancement.
... For sample preparation, a procedure mimicking wet ball milling was performed. Similar process has already been reported by Corrias et al. [22]. Briefly, coarse dispersions of curcumin were prepared using a rotor-stator homogenizer (UltraTurax, IKA, Staufen, Germany), and further nanonization was conducted in a Cell Disruptor Genie (Scientific Industries, USA), applying yttrium stabilized zirconium dioxide beads (Sigmund Lindner, Warmensteinach, Germany; diameter 0.1-0.2 ...
As the number of poorly soluble drugs is increasing, nanocrystals have become very interesting due to wide range of application possibilities. Curcuminwas used as a model active ingredient in this work. Even though it has many proven positive effects, due to its physicochemical issues, its possibilities have not been fully exploited. The goal of this work was to select optimal conditions for a top-down method for curcumin nanosuspension production, and to perform their comprehensive characterization applying complementary methodologies: dynamic light scattering, polarization and atomic force microscopy, thermal analysis, X-ray powder diffraction, antioxidant activity evaluation, release kinetics assessment, and screening of potential biological effects applying cell viability assays on normal human lung fibroblasts, human melanoma and human adenomacarcinoma cells. After 30 min of milling, nanosuspensions stabilized by polysorbate 80 and by its combinations with sucrose palmitate showed good stability, while curcumin crystal structure was unaltered. Obtained nanocrystals were well defined, with average diameter 120–170 nm and PDI of about 0.25, zeta potential was below −30 mV and pH~5 for all formulations. Nanodispersions exhibited high antioxidant potential and improved dissolution rate compared to the corresponding coarse dispersions. Although curcumin nanodispersions exhibited significant antiproliferative effect to each cancer cell line, the highest effect was towards adenocarcinoma cells.
... studies and could be identified to represent efficient carriers for intrafollicular drug delivery [5][6][7]. However, until now only a few studies have investigated hair follicle targeting with nanocrystals [8][9][10][11]. A detailed study regarding the penetration depth or exact localisation of nanocrystals within the follicles and/or information on the influence of the vehicle in which the nanocrystals are formulated is not yet available. This information is of high importance, because within the hair follicle, key target sides at different depths could be identified, whose selective treatment enables the therapy of different follicular diseases [1]. ...
Nanocrystals are a universal formulation approach for improved drug delivery of poorly water-soluble drug substances. Besides oral application, also topical application of the nanocrystals is feasible, because the increased kinetic solubility of the nanocrystals results in an increased concentration gradient, thus fostering passive, dermal penetration. Nanocrystals are also promising for targeting drug substances into the hair follicle. After penetration into the hair follicle, the nanocrystals could form a depot from which the active is released into the hair follicle. Thus, leading to a long-lasting and very efficient dermal drug delivery. The efficacy of nanocrystals to penetrate the hair follicles and the influence of the vehicle in which the nanocrystals are suspended was not yet investigated. Therefore, in this study curcumin nanocrystals with a size of about 300 nm were produced and incorporated into gels with different properties. The efficacy to penetrate the hair follicles, as well as the passive, dermal penetration, was assessed on the ex-vivo pig ear model. Nanocrystals were efficiently taken up by the hair follicles and reached the lower part of the infundibulum. This region is optimal for efficient drug delivery because the barrier of the lower infundibulum is not fully developed and thus more permeable, which results in a less hindered passive diffusion of drug substances. The penetration efficacy of the nanocrystals into the hair follicles was not affected by the different types of vehicles, which represented either oleogels or hydrogels that varied in viscosity as well as in the type and the concentration of the gelling agent. All gels possessed a shear-thinning flow behavior and it is hypothesized that all gels fluidized during the skin massage, whereby leading to similarly low viscosities than the aqueous nanosuspension and thus to similar penetration results. The passive, dermal penetration of curcumin was different for the different gels and the main driving parameter leading to good passive diffusion was caused by good skin hydrating properties of the vehicle. The best passive penetration was achieved from hydrogels that contained a humectant. However, the addition of the humectant reduced the efficacy of the nanocrystals to penetrate the hair follicle. Data so far, therefore, suggest that hair follicle targeting with nanocrystals that are suspended in water or simple, shear-thinning gels is highly effective. However, the addition of other excipients, e.g. humectants, to these vehicles might cause changes in the penetration profiles. More research in this regard is needed to understand these observations in more detail.
... In vitro and in vivo studies have demonstrated that nanocrystals can increase dermal drug bioavailability by enhancing the dissolution rate and saturation solubility, thus, leading to an increased concentration gradient between dermal formulation and the skin and, consequently, an improved skin penetration of drugs [12,[17][18][19]. Moreover, the enhanced retention time of these systems on the skin, also through accumulation in the hair follicles [20], favours a prolonged and intimate interaction between drug and skin and promoting its therapeutic effect. ...
... In addition, poorly water soluble dyes, such as nile red could also be directly nanosized by WBM, which could be applied to study the nanocrystal penetration and accumulation in the hair follicles with the assistance of fluorescence imaging as well as microscopy technology (Corrias et al., 2017). ...
Drug nanosuspensions/nanocrystals have been recognized as one useful and successful approach for drug delivery. Drug nanocrystals could be further decorated to possess extended functions (such as controlled release) and designed for special in vivo applications (such as drug tracking), which make best use of the advantages of drug nanocrystals. A lot of novel and advanced size reduction methods have been invented recently for special drug deliveries. In addition, some novel downstream processes have been combined with nanosuspensions, which have highly broadened its application areas (such as targeting) besides traditional routes. A large number of recent research publication regarding as nanocrystals focuses on above mentioned aspects, which have widely attracted attention. This review will focus on the recent development of nanocrystals and give an overview of regarding modification of nanocrystal by some new approaches for tailor-made drug delivery.
... Many researchers worked on the enhancement of saturation solubility through the preparation of drug nanocrystals. , F et al.; 2017) Nile red nanocrystals reported to have 5 and 637 times higher saturation solubility than coarse suspension and bulk, respectively [53]. (Hatahet, T. et al.; 2016) Quercetin nanocrystals increased the saturation solubility up to 5.46 to 7.46 fold than the initial saturation solubility of 0.48±0.12 ...
Introduction
Nanocrystals constitutes of 100% drug and considered as a “new drug” by Food and Drug Administration. It is proven to be an effective alternative for topical delivery of drug with increased bioavailability. Recently formulation of the drug as nanocrystals has been accomplished for many drugs exhibiting low aqueous solubility, ineffective permeability or both in order to increase the dermal bioavailability.
Conclusion
In this review article, an effort was made to explain the role of nanocrystals in the dermal delivery of the drug which results in increased bioavailability and efficacy through enhancement of solubility, dissolution velocity, permeation and penetration. Recently dermal delivery of the drug as nanocrystals is a challenging method but explained by many researchers through their work. Preparation of drugs as nanocrystals might be a promising method of drug delivery to Class II and Class IV drugs of Biopharmaceutical Classification System. Drug nanocrystals can also be applied in cosmetics for effective results.
... Various methods have been reported for the preparation of organic fluorescent dye nanoparticles, including ball milling, 21 ion association, 22 selfassembly method, 23,24 micro-emulsion method, 25,26 laser fabrication, 27 vapor deposition, 28 supercritical fluid technology, 29 and antisolvent precipitation. 30,31 Among these techniques, antisolvent precipitation has a great prospect, because of its low cost, convenient processing, and easy industries scale-up. ...
Organic fluorescent dyes have attracted wide interest because of their high photoluminescence quantum efficiencies. However, there are several application limitations arising from their hydrophobicity, poor dispersity and large particle sizes. These problems can be improved by preparing nanoparticles with a small size. Herein, we present a continuous approach to efficiently prepare an aqueous nanodispersion of water‐insoluble organic fluorescent dye Nile red (NR) with monodispersed and uniform nanoparticles (35 nm) by high‐gravity antisolvent precipitation in a rotating packed bed (RPB). In contrast, NR nanodispersions prepared using a traditional batch stirred tank (ST) had a broad size distribution (20–150 nm). Due to its small size effect and good dispersity in water, the RPB nanodispersion displayed significantly increased saturation solubility and much stronger fluorescent intensity compared to raw NR, and was obviously superior to the ST counterpart. Furthermore, NR nanodispersions were mixed with ink to draw fluorescent patterns on paper for counterfeit labeling.
Skin disorders are the most common apprehension worldwide among different regions of the world. Topical route of administration offers benefits over other routes such as avoidance of first‐pass metabolism, low dose, longer residence time, and absence of off‐target delivery. Skin serves as a mechanical barrier for therapeutic delivery with selectively permeable essential molecules. Considering the structural complexity of skin, delivery of therapeutics at targeted site requires sophisticated method such as nanotechnology‐assisted therapeutic delivery. The roadmap for combinatorial approach of nanotechnology and skin therapeutics has proven significant in clinical and marketed products. Currently, various pharmaceutical aids such as nanocrystal (NCs), nanoparticles, nanoemulsion, nano‐micelles, nano lipidic carriers, and hybrid nanocarriers are currently in market. Among all the other nanocarriers, nanocrystal offers precedence over other nanocarriers due to its facile method of preparation, reproducibility, low excipient concentration, and high therapeutic loading capacity. The recent literature data suggest the breakthrough evolution of NCs in topical therapeutic delivery. The outcome of these interventions envisages the applicability of NCs for delivering molecules with compromised physicochemical characteristics such as solubility, stability, toxicity, and bioavailability concerns.
The hair follicle (HF) is a multicellular complex structure of the skin that contains a reservoir of multipotent stem cells. Traditional hair repair methods such as drug therapies, hair transplantation, and stem cell therapy have limitations. Advances in nanotechnology offer new approaches for HF regeneration, including controlled drug release and HF-specific targeting. Until recently, embryogenesis was thought to be the only mechanism for forming hair follicles. However, in recent years, the phenomenon of wound-induced hair neogenesis (WIHN) or de novo HF regeneration has gained attention as it can occur under certain conditions in wound beds. This review covers HF-specific targeting strategies, with particular emphasis on currently used nanotechnology-based strategies for both hair loss-related diseases and HF regeneration. HF regeneration is discussed in several modalities: modulation of the hair cycle, stimulation of progenitor cells and signaling pathways, tissue engineering, WIHN, and gene therapy. The HF has been identified as an ideal target for nanotechnology-based strategies for hair regeneration. However, some regulatory challenges may delay the development of HF regeneration nanotechnology based-strategies, which will be lastly discussed.
Particle size is a key parameter to determine the capacity of nanoparticles to overcome the skin barrier; however, such effect and the possible mechanism remain only partially understood for nanosuspensions. In this work, we examined the skin delivery performance of andrographolide nanosuspensions (AG-NS) ranging in diameter from 250 nm to 1000 nm and analyzed the role of particle size in influencing their ability of skin penetration. The AG-NS with particle sizes of about 250 nm (AG-NS250), 450 nm (AG-NS450), and 1000 nm (AG-NS1000) were successfully prepared by ultrasonic dispersion method and characterized by transmission electron microscopy. The drug release and penetration via the intact and barrier-removed skin were compared by the Franz cell method, and the related mechanisms were probed using laser scanning confocal microscopy (LSCM) via visualization of penetration routes and histopathological study via observation of structural change of the skin. Our finding revealed that drug retention in the skin or its sub-layers was increased with the reduction of particle size, and the drug permeability through the skin also exhibited an obvious dependence on the particle size from 250 nm to 1000 nm. The linear relationship between the in vitro drug release and ex vivo permeation through the intact skin was well established among different preparations and in each preparation, indicating the skin permeation of the drug was mainly determined by the release process. The LSCM indicated that all these nanosuspensions could deliver the drug into the intercellular lipid space, as well as block the hair follicle in the skin, where a similar size dependence was also observed. The histopathological investigation showed that the formulations could make the stratum corneum of the skin loose and swelling without severe irritation. In conclusion, the reduction of particle size of nanosuspension would facilitate topical drug retention mainly via the modulation of drug release.
Vaccine hesitancy and the occurrence of elusive variants necessitate further treatment options for coronavirus disease 2019 (COVID-19). Accumulated evidence indicates that clinically used hypertensive drugs, angiotensin receptor blockers (ARBs), may benefit patients by mitigating disease severity and/or viral propagation. However, current clinical formulations administered orally pose systemic safety concerns and likely require a very high dose to achieve the desired therapeutic window in the lung. To address these limitations, we have developed a nanosuspension formulation of an ARB, entirely based on clinically approved materials, for inhaled treatment of COVID-19. We confirmed in vitro that our formulation exhibits physiological stability, inherent drug activity, and inhibitory effect against SARV-CoV-2 replication. Our formulation also demonstrates excellent lung pharmacokinetics and acceptable tolerability in rodents and/or nonhuman primates following direct administration into the lung. Thus, we are currently pursuing clinical development of our formulation for its uses in patients with COVID-19 or other respiratory infections.
Pomalidomide (POM) is an FDA-approved immunomodulatory imide drug (IMiDs) an it is effectively used in the treatment of multiple myeloma. IMiDs are analogs of the drug thalidomide and they have been repurposed for the treatment of several diseases such as psoriatic arthritis and Kaposi Sarcoma. In recent years, IMiDs have been also evaluated as a new treatment for neurological disorders with an inflammatory and neuroinflammatory component. POM draws particular interest for its potent anti-TNF-α activity at significantly lower concentrations than the parent compound thalidomide. However, POM’s low water solubility underpins its low gastrointestinal permeability resulting in irregular and poor absorption. The purpose of this work was to prepare a POM nanocrystal-based formulation that could efficiently improve POM’s plasma and brain concentration after intraperitoneal injection. POM nanocrystals prepared as a nanosuspension by the media milling method showed a mean diameter of 219 nm and a polydispersity index of 0.21. POM’s nanocrystal solubility value (22.97 µg/mL) in phosphate buffer was about 1.58 folds higher than the POM raw powder. Finally, in vivo studies conducted in adult Male Sprague-Dawley rats indicated that POM nanocrystal ensured higher and longer-lasting drug levels in plasma and brain when compared with POM coarse suspension.
Oxybutynin (OXY) is the most common drug to treat overactive bladder (OAB) syndrome. Transdermal administration is a more ideal route replacing oral administration to resolve problems of low bioavailability and severe side effects. However, commercial transdermal products of OXY frequently cause skin irritation and low permeation efficiency arising discontinued medication. Here, oxybutynin nanosuspension (OXY-NS) and its gel preparation (OXY-NG) were constructed to resolve these issues. In vitro permeation test and in vivo pharmacokinetics study confirmed that OXY-NG significantly enhanced the transdermal permeation of OXY, about 4-fold and 3-fold higher than oxybutynin coarse suspension (OXY-CG) respectively and in vitro retention test certified that OXY-NG increased OXY concentration especially in viable epidermis (VE) and Dermis (about 3 times that of OXY-CG), consequently improving the bioavailability. Skin irritation assay demonstrated that OXY-NG would not trigger skin adverse effects. In addition, selectively blocking hair follicles test evidenced that hair follicles pathway played an important role in OXY-NS transdermal delivery. In general, by virtue of excellent drug loading, low toxicity and ease of scale-up, OXY-NG is a promising strategy to ameliorate skin permeation of insoluble OXY for better transdermal treatment for OAB, hence increasing its bioavailability, reducing adverse effects and achieving good patient compliance.
Androgenetic alopecia (AGA) affects physical and mental health with limited therapeutic options. Novel materials and delivery methods have considerable potential to improve the current paradigm of treatment. In this study, we used a novel plant nanoparticle of safflower oil body (SOB) loaded with human fibroblast growth factor 10 (hFGF10) to target hair follicles and accelerate hair regeneration in AGA mice with few adverse effects. Our data revealed that the average particle size of SOB-hFGF10 was 226.73 ± 9.98 nm, with a spherical and uniform structure, and that SOB-hFGF10 was quicker to preferentially penetrate into hair follicles than hFGF2 alone. Using a mouse model of AGA, SOB-hFGF10 was found to significantly improve hair regeneration without any significant toxicity. Furthermore, SOB-hFGF10 inhibited dihydrotestosterone (DHT)-induced TNF-α, IL-1β, and IL-6 overproduction in macrophages in relation to hair follicle microinflammation, thereby enhancing the proliferation of dermal papilla cells. Overall, this study provides an applicable therapeutic method through targeting hair follicles and reducing microinflammation to accelerate hair regeneration in AGA.
Curcumin has shown a potential extraordinary activity as an add-on ingredient in asthma treatment, due to its immunomodulatory and anti-inflammatory mechanism of action. However, its low water solubility and bioavailability lead to a poor therapeutic effect, which can be overcome by its formulation as nanocrystals. The aim of this study was to prepare a multicomponent formulation for the delivery of curcumin (CUR) and beclomethasone dipropionate (BDP) into the lungs as water-based nanosuspensions (NS). Single component formulations (CUR-NS, BDP-NS) and a multicomponent formulation (CUR+BDP-NS) were prepared through a wet ball media milling technique, using P188 as a non-toxic stabilizer. Characterization was carried out in terms of size, size distribution, zeta potential, nanocrystals morphology, and solid-state properties. Moreover, the inhalation delivery efficiency was studied with Next Generation Impactor (NGI, Apparatus E Ph. Eu). CUR-NS was optimized and showed a long-term stability and improved nanocrystals apparent solubility. The three formulations exhibited a nanocrystal mean diameter in the range of 200-240 nm and a homogenous particle size distribution. Aggregation or sedimentation phenomena were not observed in the multicomponent formulation on 90 days storage at room temperature. Finally, the nebulization tests of the three samples showed optimal aerodynamic parameters and MMAD < 5 µm.
Cancer remains a significant health issue worldwide. The most common group of chemotherapeutic agents are small-molecule drugs, which often are associated with toxic side-effects and non-specific delivery, leading to limited therapeutic effect. This paper describes the development of a targeted drug delivery system based on lipid nanoparticles for cancer therapy. The lipid nanoparticles consist of a lipid core conjugated to an albumin stealth coating and targeting antibodies through thiol chemistry synthesized utilizing a one-step method. Applying the developed method, lipid nanoparticles with diameters down to 87 nm, capable of encapsulating small molecule compounds were synthesized. Cellular uptake studies of the lipid nanoparticles loaded with the model drug Nile red demonstrated that stealth-coating reduced non-specific cell uptake by up to a 1000-fold compared to free drug. Moreover, antibody-conjugation led to a significant cellular retargeting. Finally, it was shown that the lipid nanoparticles undergo cellular uptake through the endocytic pathway. The lipid nanoparticles are simple to synthesize, stabile in serum and have the potential to be versatile targeted towards receptors selectively expressed by diseased cells using antibodies. Thus, the system may reduce the toxic side-effects of cancer drugs while improving their delivery to cancer cells, increasing the therapeutic effect.
Data on dermal penetration of nanoparticles (NPs) was reviewed with the goal to establish a worst-case dermal penetration value for NPs. To this aim, the main focus was on studies providing quantitative dermal penetration data (29 studies). In vivo dermal penetration studies and ex vivo studies based on skin explants were included. These studies used NPs with different compositions, dimensions, and shapes. The overall results showed that skin is an efficient barrier for NPs, indistinctly of their properties. However, some studies reported that a small percentage of the applied NP dose penetrated the skin surface and reached deeper skin layers. The integrity of the skin layer and the product formulation were more critical determinants of dermal penetration than the NP properties. Most quantitative studies were based on elemental analysis such that it cannot be concluded if detected levels are attributable to a dissolved fraction or to the penetration of particles as such. Results of qualitative imaging studies suggest that at least a fraction of the levels reported in quantitative studies could be due to particle penetration. Altogether, based on the data compiled, we propose that 1% could be used as a worst-case dermal penetration value for nanoparticles within the boundaries of the properties of those included in our analysis.
In this study, beclomethasone dipropionate nanocrystals were formulated using an antisolvent sonoprecipitation technique. The aim of this study was to assess the ability of these nanocrystals to actively accumulate in skin layers for the treatment of atopic dermatitis with minimal systemic levels. A three-factor, three-level central composite design was employed to optimize the formulation parameters. The following formulation variables were adopted: stabilizer concentration, antisolvent volume, and stabilizer type. The nanocrystals were assessed for particle size, zeta potential and polydispersity index. Saturation solubility and ex vivo studies were carried out on the optimized formula (PN1). The saturation solubility of PN1 was approximately 745.5 times that of raw BDP in water. Ex vivo studies showed that the calculated Local Accumulation Efficiency (LAC) in the case of PN1 was 6.20, which was significantly higher (p < 0.05) than that of the brand formula, with a LAC ratio of 0.25. Our results confirmed the enhancement of drug deposition, ascertaining the successful use of nanocrystals for the topical delivery of poorly soluble drugs.
The search for some effective and non-invasive delivery modules to transport the therapeutic molecules across skin has led to the discovery of a number of nanocarriers (viz: liposomes, ethosomes, dendrimers, etc.) in last few decades. However, available literature suggests that these delivery modules are facing several issues including poor stability, low encapsulation efficiency, and scale-up hurdles. Recently carbon nanotubes (CNTs) emerged as a versatile tool to deliver the therapeutics across the skin. Superior stability, high loading capacity, well-developed synthesis protocol as well as ease of scale-up are some of the reason for growing interest in CNTs. CNTs have a unique physical architect, a large surface area with unique surface chemistry that can be tailored for vivid biomedical applications. CNTs have been thus largely engaged in the development of transdermal systems such as tuneable hydrogels, programmable nonporous membranes, electroresponsive skin modalities, protein channel mimetic platforms, reverse iontophoresis, microneedles, and dermal buckypapers. In addition, CNTs were also employed in the development of RNA interference (RNAi) based therapeutics for correcting the defective dermal genes. This review expounds the state-of-art synthesis methodologies, skin penetration mechanism, drug liberation profile, loading potential, characterization techniques, transdermal applications along with a summary on patent/regulatory status and future scope of CNT based skin therapeutics.
Liposomes had been widely used for drug delivery in the past. In this study, five different liposomes were used as a follicular delivery system in pig ear skin. The liposomes mainly differed in their sphere diameter, lipid composition, and surface charge. A novel class of liposomes being amphoteric in their charge behavior are compared to established anionic and cationic liposomes. Two different fluorescent dyes, hydrophilic carboxyfluoresceine or lipophilic curcumin, were enclosed in the liposomes and used as model drugs. The fluorescent dyes were also applied in a standard formulation for reference. The penetration depth of the dyes was measured by laser scanning microscopy in histological sections. One hour, 3, 5, and 7 days after application, biopsies were taken and the penetration depth into the hair follicle was measured in longitudinal sections. The liposomes showed a higher penetration depth compared to the standard formulation. The relative penetration depth of the dyes, applied in the standard formulation, averaged 30% of the full follicle length during the whole observation period, whereas the liposomal formulations penetrated considerably deeper into the hair follicles. Amphoteric and cationic liposomes reached an average relative penetration depth of approximately 70% of the full hair follicle length.
The penetration and storage behavior of dye-containing nanoparticles (diameter 320 nm) into the hair follicles was investigated. The results were compared to the findings obtained with the same amount of dye in the non-particle form. In the first part of the experiments, the penetration of the dye into the hair follicles was investigated in vitro on porcine skin, which is an appropriate model for human tissue. It was found that the nanoparticles penetrate much deeper into the hair follicles than the dye in the non-particle form, if a massage had been applied. Without massage, similar results were obtained for both formulations. Subsequently, the storage behavior of both formulations in the hair follicles was analyzed in vivo on human skin by differential stripping. Using the same application protocol, the nanoparticles were stored in the hair follicles up to 10 days, while the non-particle form could be detected only up to 4 days. Taking into consideration the surface structure of the hair follicles, it was assumed that the movement of the hairs may act as a pumping mechanism pushing the nanoparticles deep into the hair follicles.
Inflammatory disorders of the skin pose particular therapeutic challenges due to complex structural and functional alterations of the skin barrier. Penetration of several anti-inflammatory drugs is particularly problematic in psoriasis, a common dermatitis condition with epidermal hyperplasia and hyperkeratosis. Here, we tested in vivo dermal penetration and biological effects of dendritic core-multishell-nanocarriers (CMS) in a murine skin model of psoriasis and compared it to healthy skin. In both groups, CMS exclusively localized to the stratum corneum of the epidermis with only very sporadic uptake by Langerhans cells. Furthermore, penetration into the viable epidermis of nile red as a model for lipophilic compounds was enhanced by CMS. CMS proved fully biocompatible in several in vitro assays and on normal and psoriatic mouse skin. The observations support the concept of CMS as promising candidates for drug delivery in inflammatory hyperkeratotic skin disorders in vivo.
In this work a diclofenac acid nanosuspension formulation was produced as a novel approach for the treatment of skin inflammation. Drug nanocrystals, prepared by the wet media milling technique and stabilized using Poloxamer 188, were characterized by different techniques: scanning electron microscopy, differential scanning calorimetry, X-ray powder diffractometry, Fourier transform infrared spectroscopy and photon correlation spectroscopy. The ability of nanocrystals to improve dermal drug bioavailability was investigated ex vivo by using Franz diffusion vertical cells and mouse skin, in comparison with both diclofenac acid coarse suspensions and a commercial formulation. The topical anti-inflammatory activity of the drug nanosuspension was assessed in vivo by testing its effect compared to common inflammatory endpoints: i.e. the inhibition of chemically induced oedema and leucocyte infiltration (reflected in myeloperoxidase activity). Following the milling procedure, diclofenac nanocrystals exhibited a mean diameter of approximately 279 nm, a low polydispersity index (∼ 0.17) and maintained the same polymorphic form of the starting bulk powder. When the drug nanosuspension was applied on the mouse skin it produced a higher accumulation of diclofenac in the skin compared to both the coarse suspensions and the commercial formulation, as demonstrated by ex vivo transdermal delivery experiments. Moreover, the nanosuspension provided an in vivo oedema inhibition of 50%, which was not statistically different from the commercial formulation. On the contrary, the nanosuspension showed a higher inhibition of myeloperoxidase activity in the damaged tissue (86%) than the commercial formulation (16%).
Over the past two decades nanosizing technology has become one of the most successful formulation approaches for improving the bioavailability of poorly soluble drugs, which show a low oral absorption due to low dissolution velocity. Nanocrystals are nanoparticles of pure drug, without any matrix material, with an average diameter below 1 µm (typically in the range of 200-500 nm), which can be prepared in both water and non-water media as colloidal nanosuspensions stabilized using surfactants or polymers. The reduction of the drug particle diameter below 1 µm increases the dissolution velocity by increasing the particle surface and decreasing the diffusion layer thickness. Nanosuspension production processes involve two different approaches such as bottom-up (drug nanocrystal precipitation) and top-down (drug particle disintegration) technologies or a combination of two. Within these main approaches, a variety of possible techniques to achieve particle size reduction have been proposed by different research groups from both industry and academia. Even though nanosuspensions formulations have been especially studied for oral and parenteral administration, nanocrystals have showed a great potential also for topical delivery through alternative routes such as dermal, pulmonary and ocular route. The purpose of this review is to describe the main technologies used to produce nanosuspensions as well as to explore the most significant results and progresses obtained by application of drug nanocrystal formulations through topical routes.
In this work, nanocrystal formulations, containing two different diclofenac acid crystal forms, were developed with the aim to improve dermal drug bioavailability. Nanosuspensions were obtaining using wet media milling technique and were characterized in terms of size distribution, morphology, zeta potential, differential scanning calorimetry and X-ray powder diffractometry. The ability of the nanocrystals to improve dermal drug bioavailability was investigated in vitro using Franz diffusion vertical cells and newborn pig skin, in comparison with diclofenac acid coarse suspensions and a commercial topical formulation containing diclofenac sodium. Nanocrystals exhibited a mean diameter ranging between 279 and 315 nm and a PI lower than 0.25, as shown by PCS measurements. The XRDP and DSC analysis clearly indicated that the preparation process did not modify the diclofenac polymorphic forms. In vitro transdermal delivery experiments showed an improved skin deposition and permeation of the nanocrystals compared to coarse suspensions and diclofenac sodium commercial topical formulation. These results highlight the fundamental role of the crystal size on drug solubility and, thus, on the ability of a poorly soluble drug to cross the skin and accumulate in the deeper skin layers.
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Industrial concentrates of hesperidin nanocrystals (5.0% nominal concentration) were produced applying the smartCrystal(®) combination technology of wet bead milling and subsequent high pressure homogenization. Stabilization was performed by Kolliphor(®) P 188, preservation by Euxyl PE 9010 and glycerol. Physical and chemical stability were monitored over 1.5 years of storage at 4-6°C. The size of the bulk population stayed unchanged with about 250nm (photon correlation spectroscopy). Absence of crystal growth by Ostwald ripening and absence of agglomerates were shown by laser diffraction (LD) and light microscopy. The LD diameter 90% was still 0.7μm after 1.5 years. Despite the large surface of the nanosuspension in contact with the water phase, the chemical content proved also stable, only a reduction by 0.15% from 5.70% to 5.55% content was observed. The nanocrystals kept their crystalline state unchanged as shown by X-ray diffraction. The saturation solubility of the nanosuspension was more than triple compared to the raw drug powder in water. The data show the availability of a stable hesperidin concentrate as intermediate for industry to produce dermal formulations.
Copyright © 2014. Published by Elsevier B.V.
After use in oral pharmaceutical products, nanocrystals are meanwhile applied to improve the dermal penetration of cosmetic actives (e.g., rutin, hesperidin) and of drugs. By now, nanocrystals are only dermally applied made from poorly soluble actives. The novel concept is to formulate nanocrystals also from medium soluble actives, and to apply a dermal formulation containing additionally nanocrystals. The nanocrystals should act as fast dissolving depot, increase saturation solubility and especially accumulate in the hair follicles, to further increase skin penetration. Caffeine was used as model compound with relevance to market products, and a particular process was developed for the production of caffeine nanocrystals to overcome the supersaturation related effect of crystal growth and fiber formation - typical with medium soluble compounds. It is based on low energy milling (pearl milling) in combination with low dielectric constant dispersion media (water-ethanol or ethanol-propylene glycol mixtures) and optimal stabilizers. Most successful was Carbopol(®) 981 (e.g., 20% caffeine in ethanol-propylene glycol 3:7 with 2% Carbopol, w/w). Nanocrystals with varied sizes can now be produced in a controlled process, e.g., 660nm (optimal for hair follicle accumulation) to 250nm (optimal for fast dissolution). The short term test proved stability over 2 months of the present formulation being sufficient to perform in vivo testing of the novel concept.
The aims of this work were to improve cutaneous targeting and photostability of tretinoin by using nanosuspension formulation. Tretinoin is a drug widely used in the topical treatment of various dermatological diseases. The tretinoin nanosuspension was prepared by precipitation method and then characterized by photo correlation spectroscopy for mean size and size distribution, and by transmission electron microscopy for morphological studies. An oil in water tretinoin nanoemulsion was also prepared and used as a control. Dermal and transdermal delivery of both tretinoin nanosuspension and nanoemulsion were tested in vitro by using Franz diffusion cells and newborn pig skin. Photodegradation studies were carried out by UV irradiation (1h, λ=366nm) of the tretinoin nanosuspension in comparison with the nanoemulsion and a methanolic solution of the drug. During 8h percutaneous experiments, the nanosuspesion was able to localize the drug into the pig skin with a very low transdermal drug delivery, whereas the nanoemulsion greatly improved drug permeation. UV irradiation of the nanosuspension showed a great improvement of tretinoin stability in comparison with both controls. Overall results show that nanosuspension might be a useful formulation for improving tretinoin dermal delivery and stability.
The effects of both particle shape and size on the dissolution rate of sparingly soluble micro-particles have been studied. Griseofulvin, three different qualities of barium sulphate, oxazepam and glibenclamide were chosen as the model substances. As these materials differed in both particle size (from 1.8 to 3.8 μm) and particle shape (from rounded, isodiametric to flaky or irregular particles), different combinations of particle size and shape were used to investigate their combined effect on the dissolution rate. The surface specific dissolution rate was determined, using a Coulter counter technique. The ratio of this value to the equilibrium solubility was calculated for all materials and its relation to the shape and size of the micro-particles was investigated. The results obtained indicated that the dissolution rates of sparingly soluble drugs are related to the particle shape as well as to the particle size. For particles of the same size, the dissolution rate decreased as the level of flakiness and irregularity increased. This phenomenon can be explained by an increase in the average hydrodynamic boundary layer thickness as the particles become more irregular. Therefore, the product of the surface shape factor and mean particle diameter was used to discuss the combined effect of these factors on the dissolution rates of sparingly soluble drugs.
Nanosuspensions are nanosized colloidal dispersion systems stabilized by surfactants, polymers or a combination of both. Due to nanosizing results in the creation of new interfaces and in a positive Gibbs free energy change, nanosuspension is a thermodynamically unstable system with tendency of agglomeration or crystal growth. Despite the nanosuspensions technology has been extensively investigated, stability issue is still the limitation and shortcoming for its application on pharmaceutical industrial. Furthermore, the knowledge on empirical relationship between the stabilizer efficacy and stability of the nanosuspension is relatively deficiency. This review especially focused on the stability issue of nanosuspensions in drug delivery to retrieve the state art of the nanosuspensions. Therefore, the main contents of present review including the presentation of nanosuspensions instable, the method and guideline for section and optimizing stabilizers, the approaches for enhancing stability, as well as the other influencing factors on the stability of the prepared nanosuspensions. For a given drug candidate having a set of properties, this article could be used as a reference in making educated choice of stabilizer and in optimizing operation parameters for nanosuspensions formulation, rather than a trial and error approach that is being practiced currently.
In transdermal drug delivery systems, it is always a challenge to achieve stable and prolonged high permeation rates across the skin since the concentrations of the drug dissolved in the matrix have to be high in order to maintain zero order release kinetics. Several attempts have been reported to improve the permeability of poorly soluble drug compounds using supersaturated systems. However, due to thermodynamic challenges, there was a high tendency for the drug to nucleate immediately after formulating or even during storage. The present study focuses on the efficiency of nanoparticles and influence of different concentrations of solubilizer such as vitamin E TPGS (D-a-tocopheryl polyethylene glycol 1000 succinate) to improve the permeation rate through the skin. Effects of several formulation factors were studied on the nanosuspension systems using ibuprofen as a model drug. The overall permeation enhancement process through the skin was influenced mostly by the solubilizer and also by the size of nanoparticles. The gel formulation developed with vitamin E TPGS + HPMC nanosuspension, consequently represent a promising approach aiming to improve the permeability performance of a poorly water soluble drug candidate.
The aims of this study were to investigate the effect of the oil content on the physicochemical properties of NLC and to elucidate the potential of NLC for skin targeting. The obtained results showed that an increase in the oil content did not affect the mean particle size of NLC but impacted on the zeta potential. The inner structure of NLC was influenced by the increasing proportion of oil towards the less ordered structure as confirmed by differential scanning calorimetry (DSC) and X-ray diffraction (XRD), particularly for the higher medium chain triglycerides (MCT) loading. The data from proton nuclear magnetic resonance (1H NMR) revealed that cetyl palmitate nanoparticles did not completely recrystallize after cooling down to room temperature. 1H NMR and DSC results indicate that MCT molecules were restricted in the NLC as compared to the nanoemulsions (NE). Nile red distribution and penetration into skin from NLC were pronounced as compared to NE and dependent on the MCT loading. The deep penetration and high amount of Nile red were related to the occlusion factor. Moreover, the epidermal targeting was achieved by NLC applications, particularly those containing 5% MCT (NLC-5) depending on the amount of MCT loading.
A basic problem of poorly soluble drugs is often an insufficient bioavailability. To allow the i.v. injection of these drugs, they were formulated as nanosuspensions by high pressure homogenization. The effect of the production parameters pressure and cycle number on the mean particle size and on the polydispersity of the nanosuspension was investigated with special attention to contamination by microparticles — the limiting factor for i.v. injection. Properties of the nanosuspensions are increased saturation solubility Cs and dissolution rate dc/dt. These phenomena are explained using the Prandtl and the Ostwald–Freundlich equations. These properties promote the dissolution of the nanosuspensions in the blood after i.v. injection. The size distribution obtained and the use of an APV Gaulin homogenizer (FDA approved for parenterals) lead to a pharmaceutical product considered acceptable by the regulatory authorities.
Hair follicles represent interesting target sites for topically applied substances such as topical vaccinations or agents used in the field of regenerative medicine. In recent years, it could be shown that particles penetrate very effectively into the hair follicles. In the present study, the influence of particle size on the follicular penetration depths was examined. The penetration depths of two different types of particles sized 122 to 1000 nm were determined in vitro on porcine skin. The results revealed that the particles of medium size (643 and 646 nm, respectively) penetrated deeper into the porcine hair follicles than smaller or larger particles. It was concluded that by varying the particle size, different sites within the porcine hair follicle can be targeted selectively. For the human terminal hair follicle, the situation can be expected to be similar due to a similar size ratio of the hair follicles.
Coated titanium dioxide (TiO2) microparticles are commonly used as UV filter substances in commercial sunscreen products. The penetration of these microparticles into the horny layer and the orifice of the hair follicle was investigated. The distribution of the microparticles in the horny layer was analyzed using the method of tape stripping in combination with spectroscopic measurements. Deeper layers of the stratum corneum were devoid of TiO(2) even after repetitive application of sunscreen preparation when analyzing interfollicular areas. Only in the areas of the pilosebaceous orifices could microparticles be identified. The penetration of TiO(2) was investigated in histological skin sections. A biopsy was taken from a skin area from which the horny layer had been removed by tape stripping. In isolated areas, a penetration of coated TiO2 into the open part of the follicle was observed. The amount of TiO2 found in a given follicle was less than 1% of the applied total amount of sunscreens. A penetration of microparticles into viable skin tissue could not be detected.
To clarify the effect of the surface charge of liposomes on percutaneous absorption, the permeation of liposomal drugs through rat skin was investigated in vitro and in vivo. Liposomes were prepared using egg yolk lecithin (EPC, phase transition temperature, -15 to -17 degrees C), cholesterol and dicetylphosphate (DP) or stearylamine (SA) (10:1:1, mol/mol). Also examined was the penetration behavior of positively and negatively charged liposomes, using a fluorescent probe (Nile Red). The in vitro penetration rate of melatonin (MT) entrapped in negatively charged liposomes was higher than that of positively charged ones (p<0.05). When the percutaneous absorption of ethosuximide (ES) encapsulated was estimated in vivo, the absorption of ES from negatively charged liposomes was slightly higher than that from positively charged liposomes. Additionally, the absorption of ES from both types of liposomes was superior to that from the lipid mixtures consisting of the same composition as the vesicles. The percutaneous absorption of betahistine (BH) from a gel formulation containing negatively charged liposomes of BH was much more than that from the formulation with positively charged ones, with 2-fold higher AUC (p<0.05). Histological studies revealed that the negatively charged liposomes diffused to the dermis and the lower portion of hair follicles through the stratum corneum and the follicles much faster than the positive vesicles at the initial time stage after application. Thus, the rapid penetration of negatively charged liposomes would contribute to the increased permeation of drugs through the skin.
The influence of liposome composition, size, lamellarity and charge on the (trans)dermal delivery of tretinoin (TRA) was studied. For this purpose we studied both multilamellar (MLV) or unilamellar (UV) liposomes. Positively or negatively charged liposomes were obtained using either hydrogenated (Phospholipon90H) or non-hydrogenated soy phosphatidylcholine (Phospholipon90) and cholesterol, in combination with stearylamine or dicetylphosphate. Liposomal formulations were characterized by transmission electron microscopy (TEM) and optical and light polarized microscopy for vesicle formation and morphology, and by dynamic laser light scattering for size distribution. In order to obtain more information about the stability and the thermodynamic activity of the liposomal tretinoin, TRA diffusion through a lipophilic membrane was investigated. The effect of the vesicular incorporation of tretinoin on its accumulation into the newborn pig skin was also studied. The experiments were performed in vitro using Franz cells in occlusive conditions and were compared to three different controls. The tretinoin amount delivered through and accumulated in the several skin layers was detected by HPLC. Furthermore, TEM in combination with osmium tetroxide was used to visualize the skin structure after the liposomal administration. Overall obtained results showed that liposomes may be an interesting carrier for tretinoin in skin disease treatment, when appropriate formulations are used. In particular, negatively charged liposomes strongly improved newborn pig skin hydration and TRA retention, though no evidence of intact vesicle penetration was found.
Poorly water-soluble compounds are difficult to develop as drug products using conventional formulation techniques and are frequently abandoned early in discovery. In the present study, the melt emulsification method traditionally used to prepare solid lipid nanoparticles was adapted to produce drug nanosuspensions. The method was evaluated in comparison with the well known solvent diffusion process for ibuprofen as a model drug. Control of the preparation variables (stabilizers, drug content, homogenization procedure and cooling conditions) allowed formation of nanosuspensions with diameters less than 100 nm. The major advantage of the melt emulsification method over the solvent diffusion method is the avoidance of organic solvents during production, although the mean particle size is slightly greater. The combination of Tween 80 and PVP K25 as stabilizers yields nanosuspensions with the smallest average particle size. The formulation of ibuprofen as a nanosuspension, either in the form of lyophilized powder or granules, was very successful in enhancing dissolution rate, more than 65% of the drug being dissolved in the first 10 min compared to less than 15% of the micronized drug. The increase in in vitro dissolution rate may favourably affect bioavailability and improve safety for the patient by decreasing gastric irritancy.