Nanoemulsion-Loaded Hydrogels for Topical Administration of Pentyl Gallate.

The aim of this study is to describe the development of nanoemulsion-loaded hydrogels to deliver pentyl gallate (PG), a gallic acid n-alkyl ester, through the skin. PG is an antioxidant agent; however, it seems to be a promising agent for herpis labialis treatment. Aristoflex AVC® and chitosan were used as gelling agents for nanoemulsion thickening. The developed formulations presented suitable PG content (94.4-100.3% w/w), nanometric droplet sizes (162-297 nm), high zeta potentials, and a non-Newtonian pseudoplastic behavior. Both vehicles neither enhanced PG penetration nor delayed its release from the nanoemulsion. Formulations remained physically stable at 8°C during 3 months of storage.

Anti-inflammatory Effect from a Hydrogel Containing Nanoemulsified Copaiba oil (Copaifera multijuga Hayne).

Copaiba oil is used as a popular medicine in the Amazonian forest region, especially due to its anti-inflammatory properties. In this paper, we describe the formulation of hydrogel containing copaiba oil nanoemulsions (with positive and negative charges), its skin permeation, and its anti-inflammatory activity in two in vivo models: mouse ear edema and rat paw edema. Three hydrogels were tested (Carbopol®, hydroxyethylcellulose and chitosan), but only Carbopol® and hydroxyethylcellulose hydrogels presented good stability and did not interfere with the nanoemulsions droplet size and polydispersity index. In skin permeation assay, both formulations, positively charged nanoemulsion (PCN) and negatively charged nanoemulsion (NCN), presented a high retention in epidermis (9.76 ± 2.65 µg/g and 7.91 ± 2.46 µg/cm2, respectively) followed by a smaller retention in the dermis (2.43 ± 0.91 and 1.95 ± 0.56 µg/cm2, respectively). They also presented permeation to the receptor fluid (0.67 ± 0.22 and 1.80 ± 0.85 µg/cm2, respectively). In addition, anti-inflammatory effect was observed to NCN and PCN with edema inhibitions of 69 and 67% in mouse ear edema and 32 and 72% in rat paw edema, respectively. Histological cuts showed the decrease of inflammatory factors, such as dermis and epidermis hyperplasia and inflammatory cells infiltration, confirming the anti-inflammatory effect from both copaiba oil nanoemulsions incorporated in hydrogel.

Trypanocidal activity of the compounds present in Aniba canelilla oil against Trypanosoma evansi and its effects on viability of lymphocytes.

Aniba canelilla (H.B.K.) Mez, popularly known as "casca-preciosa" (precious bark), is a plant of the Lauraceae family, widely distributed in the Amazon region. Its major constituent is 1-nitro-2-phenylethane, a rare molecule in plants which is responsible for this plant's cinnamon scent. The present study aimed to report the chemical characterization of the oil extracted from Aniba canelilla using gas-chromatography/mass spectrometry and to assess its in vitro trypanocidal activity against Trypanosoma evansi, a prevalent haemoflagellate parasite that affects a broad range of mammal species in Africa, Asia and South America. The oil presented 1-nitro-2-phenylethane (83.68%) and methyleugenol (14.83%) as the two major components. The essential oil as well as both major compounds were shown to exert trypanocidal effect. Methyleugenol was slightly more active than 1-nitro-2-phenylethane. In vitro studies showed that the oil extracted from the stems of A. canelilla may be regarded as a potential natural treatment for trypanosomosis, once proven their in vivo action, may be an interesting alternative in the treatment of infected animals with T. evansi.

Solid dispersions enhance solubility, dissolution, and permeability of thalidomide.

Thalidomide (THD) is a BCS class II drug with renewed and growing therapeutic applicability. Along with the low aqueous solubility, additional poor biopharmaceutical properties of the drug, i.e. chemical instability, high crystallinity, and polymorphism, lead to a slow and variable oral absorption. In this view, we developed solid dispersions (SDs) containing THD dispersed in different self-emulsifying carriers aiming at an enhanced absorption profile for the drug. THD was dispersed in lauroyl macrogol-32 glycerides (Gelucire® 44/14) and α-tocopherol polyethylene glycol succinate (Kolliphor® TPGS), in the presence or absence of the precipitation inhibitor polyvinylpyrrolidone K30 (PVP K30), by means of the solvent method. Physicochemical analysis revealed the formation of semicrystalline SDs. X-ray diffraction and infrared spectroscopy analyses suggest that the remaining crystalline fraction of the drug in the SDs did not undergo polymorphic transition. The impact of the solubility-enhancing formulations on the THD biopharmaceutical properties was evaluated by several in vitro techniques. The developed SDs were able to increase the apparent solubility of the drug (up to 2-3x the equilibrium solubility) for a least 4 h. Dissolution experiments (paddle method, 75 rpm) in different pHs showed that around 80% of drug dissolved after 120 min (versus 40% of pure crystalline drug). Additionally, we demonstrated the enhanced solubility obtained via SDs could be translated into increased flux in a parallel artificial membrane permeability assay (PAMPA). In summary, the results demonstrate that SDs could be considered an interesting and unexplored strategy to improve the biopharmaceutical properties of THD, since SDs of this important drug have yet to be reported.

Antifungal Activity of Aniba canelilla (Kunth) Mez Essential Oil and Its Main Compound 1-Nitro-2-Phenylethane against Dermatophytes.

The essential oil of Aniba canelilla (Kunth) Mez (EOAC), an Amazon plant composed of a rare nitro compound, has shown scientific evidence of antifungal activity but is still unexplored against dermatophytes. The antifungal susceptibility of EOAC and its main compound, 1-nitro-2-phenylethane (NP), was evaluated against dermatophytes (Trichophyton rubrum, T. mentagrophytes and Microsporum canis), evidencing antifungal activity with an inhibitory concentration lower than 256 µg/mL. The mechanism of action was also evaluated, and it is suggested that EOAC and NP have fungicidal action in the fungal membrane, since the antifungal activity occurs through a modification of the shape of the conidial structures of the fungus, showing the permeability of the intracellular content due to the visually observed plasmolysis and cytosolic extravasation through an osmotic process. These results suggest the essential oil and its main compound are promising plant-derived alternatives for treating ungual dermatophytosis.

Preparation, characterization, and in vitro intestinal permeability evaluation of thalidomide-hydroxypropyl-ß-cyclodextrin complexes.

Thalidomide is emerging as a therapeutic agent with renewed clinical importance, presenting anti-inflammatory, immunomodulatory, and antineoplasic properties. In this work, we studied the complexation of thalidomide with cyclodextrins as a strategy to circumvent the poor aqueous solubility of the drug. Thalidomide-hydroxypropyl-ß-cyclodextrin complexes were obtained by kneading method and were characterized by differential scanning calorimetry, powder X-ray diffractometry, and scanning electronic microscopy. The aqueous solubility and in vitro dissolution of thalidomide were significantly improved through the complexation. Physicochemical analysis of the complexes in solid state revealed a decreased crystallinity of the complexed drug in comparison with free thalidomide. Thalidomide was able to dissociate from the complexes and permeates across intestinal epithelial Caco-2 cells with a favorable high permeability profile equivalent to that of the free drug. In summary, the present results suggest that thalidomide-hydroxypropyl-ß-cyclodextrin complexes could be regarded as a promising strategy for improving the gastrointestinal absorption of thalidomide.

In vivo Acute Anti-Inflammatory Activity of Essential Oils: A Review.

In recent years, there has been a significant increase in the search for new therapeutic strategies for the treatment of inflammatory diseases. In this sense, natural products emerge as a potential source for the discovery of new drugs, with the research of the pharmacological properties of these products being very important. In addition to its function in plants (insect attraction and repellency), essential oils present pharmacological effects, such as antibacterial, antifungal, antimutagenic, antiviral, antiprotozoal, antioxidant, antidiabetic and anti-inflammatory properties. In this review, we describe the mostly used in vivo acute inflammatory experimental models and the studies showing the in vivo anti-inflammatory activity of essential oils. Essential oil from species from the Apiaceae, Asteraceae, Burseraceae, Boraginaceae, Cupressaceae, Euphorbiaceae, Fabaceae, Lamiaceae, Lauraceae, Myrtaceae, Piperaceae, Poaceae, Rutaceae, Verbenaceae and Zingiberaceae families were described as being anti-inflammatory in vivo. Five models of acute inflammation are commonly used to investigate the anti-inflammatory activity in vivo: ear and paw edema, pleurisy, peritonitis and the subcutaneous air pouch model. In addition to in vivo analysis, ex vivo and in vitro experiments are carried out to study the anti-inflammatory action of essential oils. The most commonly used model was paw edema, especially due to this model being easy to perform. In order to suggest or elucidate the mechanisms involved in the anti-inflammatory effect, many studies measured some inflammatory mediators, such as cytokines, COX-2 expression and the levels of PGE2, and NO, or evaluated the effect of essential oils or their major compounds on inflammation response directly induced by inflammatory mediators.

Correlation between the Skin Permeation Profile of the Synthetic Sesquiterpene Compounds, Beta-Caryophyllene and Caryophyllene Oxide, and the Antiedematogenic Activity by Topical Application of Nanoemulgels.

Sesquiterpene compounds are applied as permeation promoters in topical formulations. However, studies exploring their impact on nanostructured systems, changes in permeation profile, and consequently, its biological activity are restricted. This study aimed to investigate the correlation between the skin permeation of the major sesquiterpenes, beta-caryophyllene, and caryophyllene oxide from the oleoresin of Copaifera multijuga, after delivery into topical nanoemulgels, and the in vivo antiedematogenic activity. First, ten nanoemulgels were prepared and characterized, and their in vitro permeation profile and in vivo anti-inflammatory activity were evaluated. In equivalent concentrations, ß-caryophyllene permeation was greater from oleoresin nanoemulgels, resulting in greater in vivo antiedematogenic activity. However, an inverse relationship was observed for caryophyllene oxide, which showed its favored permeation and better in vivo anti-inflammatory effect carried as an isolated compound in the nanoemulgels. These results suggest that the presence of similar compounds may interfere with the permeation profile when comparing the profiles of the compounds alone or when presented in oleoresin. Furthermore, the correlation results between the permeation profile and in vivo antiedematogenic activity corroborate the establishment of beta-caryophyllene as an essential compound for this pharmacological activity of C. multijuga oleoresin.

Formulating Bioactive Terpenes.

Terpenes are specialized metabolites mainly produced by plants and are highly bioactive [...].

Essential oils in nanostructured systems: Challenges in preparation and analytical methods.

Essential oils are natural products extracted from plants that present volatile and thermolabile characteristics. Essential oils have become products of interest in many fields, including the pharmaceutical, due to their medicinal properties. In recent years, the interest in the encapsulation of essential oils in nanometric systems for therapeutic approaches has risen and a number of studies have been published. This review intended to set a panorama on the research within this field through a data survey and identify the organic nanostructured systems, the preparation techniques and analytical quantification methods employed. Many techniques used to prepare nanosystems loaded with essential oils involve heating or solvent evaporation steps that may damage their composition. In this context, the quantification of essential oil on the final nanosystems is impaired. However, in more than half of the research papers, the quantification is ignored or an indirect quantification is performed, assuming no volatilisation upon formulation processes. Analytical methods used to assess essential oil encapsulation efficiency were discussed regarding their suitability.

Essential Oils and Isolated Terpenes in Nanosystems Designed for Topical Administration: A Review.

Essential oils are natural products with a complex composition. Terpenes are the most common class of chemical compounds present in essential oils. Terpenes and the essential oils containing them are widely used and investigated by their pharmacological properties and permeation-enhancing ability. However, many terpenes and essential oils are sensitive to environmental conditions, undergoing volatilization and chemical degradation. In order to overcome the chemical instability of some isolated terpenes and essential oils, the encapsulation of these compounds in nanostructured systems (polymeric, lipidic, or molecular complexes) has been employed. In addition, nanoencapsulation can be of interest for pharmaceutical applications due to its capacity to improve the bioavailability and allow the controlled release of drugs. Topical drug administration is a convenient and non-invasive administration route for both local and systemic drug delivery. The present review focuses on describing the current status of research concerning nanostructured delivery systems containing isolated terpenes and/or essential oils designed for topical administration and on discussing the use of terpenes and essential oils either for their biological activities or as permeation enhancers in pharmaceutic formulations.

A novel, simplified and stability-indicating high-throughput ultra-fast liquid chromatography method for the determination of rosmarinic acid in nanoemulsions, porcine skin and nasal mucosa.

Currently, there is an increasing interest on the development of topical formulations containing rosmarinic acid (RA) due to its well-documented antioxidant activity. This study aimed to develop and validate a stability-indicating ultra-fast liquid chromatography (UFLC) method for the determination of RA in nanoemulsions, porcine skin and nasal mucosa intended to be applied in permeation/retention studies and for development of topical nanoemulsions. Chromatographic separation was carried out using a C18 column packed with 2.6 µm particle size in isocratic conditions using as mobile phase water:acetonitrile (83:17, v/v), acidified with 0.1% trifluoracetic acid (v/v), with a total time of analysis of 3.5 min and detection at 330 nm. RA analysis was specific in the presence of both non-biological (blank nanoemulsion and receptor fluid) and biological matrices (porcine ear skin and porcine nasal mucosa). No interference of degradation products of RA was verified after different stress conditions such as acidic, alkaline, oxidative, light exposure (UV-A and UV-C) and thermal demonstrating the method stability-indicating property. The analytical (0.1-10.0 µg·mL-1) and bioanalytical (0.5-10.0 µg·mL-1) linearity was proved by analysis of the calibration curves of RA and no matrix effect was observed. The method was sensitive, precise and accurate, and showed recovery higher than 85%. The method was considered robust as evaluated by a Plackett-Burman experimental design. In the validated conditions, the RA was determined in the nanoemulsions obtained by spontaneous emulsification procedure (1.007 ±â€¯0.040 mg·mL-1), porcine ear skin (1.13 ±â€¯0.19 µg·cm-2) and nasal mucosa (22.46 ±â€¯3.99 µg·cm-2) after retention/permeation studies. Thus, a highly sensitive, simple, fast and stability-indicating method was developed for RA analysis during the development of topical nanoemulsions and bioanalytical assays in complex matrices.

Optimization, validation and application of headspace solid-phase microextraction gas chromatography for the determination of 1-nitro-2-phenylethane and methyleugenol from Aniba canelilla (H.B.K.) Mez essential oil in skin permeation samples.

Aniba canelilla (H.B.K.) Mez is an aromatic plant from the Amazon region whose essential oil has 1-nitro-2-phenylethane (NP) and methyleugenol (ME) as major compounds. Despite of the scientifically proven antifungal and anti-inflammatory activities for these compounds, there is no report up to date about the topical permeation or quantification of NP and ME on skin samples. The aim of this study was the validation of an optimized bioanalytical method by solid-phase microextraction in headspace mode in gas chromatograph with flame ionization detector (HS-SPME-GC-FID) for the determination of NP and ME from the oil in different samples from permeation study, such as porcine ear skin (PES) layers (stratum corneum, epidermis and dermis) and receptor fluid (RF). For this propose polydimethylsiloxane fibers (100 µm) were used and HS-SPME extraction condition consisted of 53 °C, 21 min, and 5% w.v-1 NaCl addition. The wide range of the calibration curve (2.08-207.87 µg mL-1 for NP and 0.40-40.41 µg mL-1 for ME), the presence of matrix interferences and the intrinsic characteristics of HS-SPME required a data linearization using Log10. Thereby, data and the gained results presented homoscedasticity, normalization of residues and adequate linearity (r2 > 0.99) and accuracy for both compounds. In order to verify the applicability of the validated method, the HS-SPME-GC-FID procedure was performed to determine the amount of NP and ME permeated and retained in samples after Franz diffusion cell study from different dosages (20, 100 and 200 µL) of A. canelilla oil. Compounds permeation showed a progressive increase and penetration dependence based on the dosage applied. Furthermore, retention was in order receptor fluid >> dermis >> epidermis >> stratum corneum for both compounds, suggesting NP and ME could penetrate deep tissue, probably due to the partition coefficient, mass, size, and solubility of these compounds. In conclusion, the proposed method by HS-SPME-GC-FID to quantify 1-nitro-2-phenylethane and methyleugenol from Aniba canelilla essential oil was able to determine selectively, precisely and accurately these main compounds in skin permeation samples.

Carbamazepine parenteral nanoemulsions prepared by spontaneous emulsification process.

Carbamazepine (CBZ), a widely used anticonvulsant drug, is a poorly soluble drug with no parenteral treatment available for patients. This study was aimed at developing a nanoemulsion for CBZ intravenous delivery. The spontaneous emulsification method was used to prepare different formulations containing 2mg/mL CBZ. Likewise, a 2(2) full factorial experimental design was applied to study the influence of two independent variables (type of oil and type of lipophilic emulsifier) on emulsion physicochemical characteristics. The nanoemulsions were evaluated concerning droplet size, zeta potential, viscosity, drug content and association to oily phase. The formulation, which presented the best characteristics required for intravenous administration was selected and refined with respect to the lipophilic emulsifier content (increase from 5% to 6% of soy lecithin). This formulation was characterized and kept its properties in a satisfactory range over the evaluated period (3 months), i.e. droplet size around 150 nm, drug content around 95% and zeta potential around -40 mV. The transmission electron microscopy revealed emulsion droplets almost spherical in shape with an amorphous core, whereas the in vitro release profile assessed by dialysis bags demonstrated a release kinetics square root time dependent, with 95% of ca. having been released within 11h.

Pentyl Gallate Nanoemulsions as Potential Topical Treatment of Herpes Labialis.

Previous studies have demonstrated the antiherpes activity of pentyl gallate (PG), suggesting that it could be a promising candidate for the topical treatment of human herpes labialis. PG low aqueous solubility represents a major drawback to its incorporation in topical dosage forms. Hence, the feasibility of incorporating PG into nanoemulsions, the ability to penetrate the skin, to inhibit herpes simplex virus (HSV)-1 replication, and to cause dermal sensitization or toxicity were evaluated. Oil/water nanoemulsions containing 0.5% PG were prepared by spontaneous emulsification. The in vitro PG distribution into porcine ear skin after topical application of nanoemulsions was assessed, and the in vitro antiviral activity against HSV-1 replication was evaluated. Acute dermal toxicity and risk of dermal sensitization were evaluated in rat model. Nanoemulsions presented nanometric particle size (from 124.8 to 143.7 nm), high zeta potential (from -50.1 to -66.1 mV), loading efficiency above 99%, and adequate stability during 12 months. All formulations presented anti-HSV-1 activity. PG was able to reach deeper into the dermis more efficiently from the nanoemulsion F4. This formulation as well as PG were considered safe for topical use. Nanoemulsions seem to be a safe and effective approach for topically delivering PG in the treatment of human herpes labialis infection.

Determination of ß-caryophyllene skin permeation/retention from crude copaiba oil (Copaifera multijuga Hayne) and respective oil-based nanoemulsion using a novel HS-GC/MS method.

Copaiba oil is largely used in the Amazonian region for the treatment of inflammation, and recent studies demonstrated that one of the major components of the oil, ß-caryophyllene (CAR), is a potent anti-inflammatory. The nanoemulsification of this oleoresin, which has unctuous character, converts it in a more acceptable hydrophilic formulation and may improve CAR penetration through the skin due to the small droplet size and the high contact surface afforded by the nanoemulsions. This paper describes the validation of a novel, sensitive, practical and solvent free method that uses gas chromatography in headspace mode coupled with mass spectrometry to evaluate the skin permeation/retention of CAR from the crude copaiba oil and its nanoemulsion. Our results show that the bioanalytic method was fully validated, demonstrating linearity (r(2)>0.99), specificity (no peaks co-eluting with CAR retention time), precision (RSD<15%) and accuracy (recovery>90%) within the accepted parameters and that the copaiba oil nanoemulsion presented a better skin penetration compared to the crude oil, with CAR achieving the most profound layer of the skin, the dermis.

A versatile, stability-indicating and high-throughput ultra-fast liquid chromatography method for the determination of isoflavone aglycones in soybeans, topical formulations, and permeation assays.

There is a growing interest in the pharmaceutical field concerning isoflavones topical delivery systems, especially with regard to their skin care properties and antiherpetic activity. In this context, the present work describes an ultra-fast liquid chromatography method (UFLC) for determining daidzein, glycitein, and genistein in different matrices during the development of topical systems containing isoflavone aglycones (IA) obtained from soybeans. The method showed to be specific, precise, accurate, and linear (0.1 to 5 µg mL(-1)) for IA determination in soybean acid extract, IA-rich fraction obtained after the purification process, IA loaded-nanoemulsions, and topical hydrogel, as well as for permeation/retention assays in porcine skin and porcine esophageal mucosa. The matrix effect was determined for all complex matrices, demonstrating low effect during the analysis. The stability indicating UFLC method was verified by submitting IA to acidic, alkaline, oxidative, and thermal stress conditions, and no interference of degradation products was detected during analysis. Mass spectrometry was performed to show the main compounds produced after acid hydrolysis of soybeans, as well as suggest the main degradation products formed after stress conditions. Besides the IA, hydroxymethylfurfural and ethoxymethylfurfural were produced and identified after acid hydrolysis of the soybean extract and well separated by the UFLC method. The method's robustness was confirmed using the Plackett-Burman experimental design. Therefore, the new method affords fast IA analysis during routine processes, extract purification, products development, and bioanalytical assays.

Influence of beta-cyclodextrin complexation on carbamazepine release from hydroxypropyl methylcellulose matrix tablets.

The in vitro release profiles of carbamazepine and beta-cyclodextrin either complexed or simply mixed and subsequently incorporated in hydrophilic matrix tablets containing 15 or 30% hydroxypropyl methylcellulose were evaluated. Solubility studies revealed a linear relationship between the increase in carbamazepine solubility and the increase in beta-cyclodextrin concentration. Drying methods (spray-drying and freeze-drying) were used to obtain carbamazepine/beta-cyclodextrin solid complexes in order to prepare tablets. The results demonstrated that matrix tablets containing carbamazepine/beta-cyclodextrin solid complexes displayed faster carbamazepine and beta-cyclodextrin release compared to that containing simple physical mixture. Gelling and matrix formation was impaired in formulation containing 15% hydroxypropyl methylcellulose and spray-dried complex. The comparison of spray-drying and freeze-drying revealed no significant influence of both drying methods on carbamazepine and beta-cyclodextrin dissolution rate when carbamazepine/beta-cyclodextrin complexes were incorporated in 30% hydroxypropyl methylcellulose matrix tablets. The results point to the possibility of modulating carbamazepine release using a hydroxypropyl methylcellulose matrix associated to the drug complexed with beta-cyclodextrin.

Mathematical evaluation of in vitro release profiles of hydroxypropylmethylcellulose matrix tablets containing carbamazepine associated to beta-cyclodextrin.

The release kinetics of carbamazepine (CBZ) either complexed or physically mixed with beta-cyclodextrin (betaCD) from hydroxypropylmethylcellulose (HPMC) matrix tablets was investigated using different mathematical equations. The model-dependent approach was compared to the utilization of fit factors. Notwithstanding difference (f1) and similarity (f2) factors allowed the differentiation of the formulations containing CBZ complexed with betaCD from the one containing a simple physical mixture of CBZ and betaCD. The Weibull model was more useful for comparing the release profiles. Weibull parameters were more sensitive to the differences between the two release kinetic data.

Bioavailability of carbamazepine:beta-cyclodextrin complex in beagle dogs from hydroxypropylmethylcellulose matrix tablets.

The bioavailability of a carbamazepine:beta-cyclodextrin (CBZ:betaCD) complex from hydroxypropylmethylcellulose (HPMC) matrix tablets was evaluated in beagle dogs. A solubility study demonstrated the improvement of CBZ aqueous solubility by adding increasing amounts of betaCD. The 1:1 CBZ:betaCD molar ratio was chosen to produce the complex, which was obtained by spray-drying. Matrix tablets were prepared by direct compressing either a CBZ:betaCD complex or a physical mixture of both substances with HPMC. Both matrix formulations displayed a controlled-release profile when compared to the reference formulation (Tegretol CR 200). CBZ presented a significantly higher bioavailability from matrix tablets containing the CBZ:betaCD complex than that obtained from Tegretol CR 200). Although a high inter-subject variability was observed, the results pointed to the feasibility of using betaCD in order to modulate CBZ release and absorption, as well as to reduce the drug dosage maintaining the same plasma levels.