Egg Test-chorioallantoic Membrane Research Articles

Doe guided chitosan based nano-ophthalmic preparation against fungal keratitis

A recent upsurge in ocular infections is a pointer towards an enhanced prevalence of ophthalmic disorders, posing challenges for researchers globally. The caveats of conventional therapeutics demand a specifically designed Ocular Drug Delivery System (ODDS) and hence the primary objective of the present work is a fabrication of a Design of Expert (DoE) guided Chitosan based Antifungal loaded Nanoparticles (CANs), as a locoregionally effective eye formulation/drops for fungal keratitis therapy. The purported formulation was prepared using High-Pressure Homogenisation technique and was critically characterized on various parameters to check their suitability as an ODDS. The optimized formulation has fruitfully yielded irregularly spherical particles in up to a size of 200 nm and a Poly-dispersity Index (PDI) of less than 0.2 nm. The optimised formulation has further showcased a high mucoadhesion capacity thereby, suggesting the greater retention of CANs on the mucous membrane of an eye with low ocular irritancy as highlighted using HET-CAM (Hen's Egg Chorioallantoic Membrane) test. The in-vitro drug release study across a dialysis membrane has indicated both diffusion as swelling controlled release pattern for an optimized formulation. The ex-vivo corneal permeation study on goat corneal tissues using a Franz-Diffusion cell also has indicated a steady increase in the permeation of drug with time for an optimized formulation. Further, the optimised formulation was found to be non-irritant and ocular safe in ex-vivo transcorneal toxicity studies on goat corneal tissues. In conclusion, the designing of a proposed nanosized formulation, offers a promising step towards the management of external ocular diseases with a positive attributes of high patient compliance, controlled drug delivery, prolonged drug precorneal residence time and enhanced ocular bioavailability. The optimized CANs could be further exploited as a potential ODDS.

Cytotoxic, genotoxic, oxidative, and irritant effects of zinc pyrithione in vitro

Zinc pyrithione is an organometallic compound with antimicrobial activity used in many industrial products. In this study, cytotoxicity (cell viability, lactate dehydrogenase release, proliferation rate, and mitotic index) and genotoxicity (sister chromatid exchange and micronucleus test) of zinc pyrithione on human lymphocytes were determined. Intracellular zinc concentrations were determined by inductively coupled plasma-optical emission spectrometry. Intracellular antioxidant/oxidant status was evaluated by total antioxidant capacity and total oxidative status assays. Its irritation potential was investigated using the hen’s egg test chorioallantoic membrane model. Up to the highest concentration of 1000 nmol/L, ZnPT did not cause genotoxicity and did not change the proliferation index. Above 500 nmol/L, zinc pyrithione caused an increase in lactate dehydrogenase, oxidative stress, and intracellular zinc levels, while decreasing cell viability and mitotic index level. At 1000 nmol/L, zinc pyrithione caused slight irritation. These results suggest that zinc pyrithione may exert toxic effects via increased oxidative stress on human cellular systems.

Investigating the ocular toxicity potential and therapeutic efficiency of in situ gel nanoemulsion formulations of brinzolamide.

Glaucoma is an ocular disease i.e. more common in older adults with elevated intraocular pressure and a serious threat to vision if it is not controlled. Due to the limitations regarding the conventional form of brinzolamide (Azopt®), two optimum formulations of in situ gel nanoemulsion were developed. To ensure the safety and efficacy of developed formulations for ocular drug delivery, the current study was designed. MTT assay was carried out on the human retinal pigmentation epithelial cells. To investigate the irritation potential of the chosen formulations, hen's egg test-chorioallantoic membrane as a borderline test between in vivo and in vitro methods has been done. The modified Draize method was utilized to evaluate eye tolerance against the selected formulations. Intraocular pressure was measured by applying the prepared formulations to the eyes of normotensive albino rabbits in order to assess the therapeutic efficacy. Based on MTT test, cell viability for NE-2 at 0.1% and NE-1 at 0.1 and 0.5% concentrations was acceptable. The results of the hen's egg test-chorioallantoic membrane test indicated no sign of vessel injury on the chorioallantoic membrane surface for both formulations. Also, during 24h, both formulations were well-tolerated by rabbit eyes. The pharmacodynamics effects of formulations had no difference or were even higher than that of suspension in case of adding lower concentration (0.5%) of brinzolamide to the formulations. With regard to the results of the mentioned methods, our advanced formulations were effective, safe, and well-tolerated, thus can be introduced as an appropriate vehicle for ocular delivery of brinzolamide.

Enhancement of Chemical Stability and Dermal Delivery of Cordyceps militaris Extracts by Nanoemulsion.

This study aimed to develop nanoemulsions for enhancing chemical stability and dermal delivery of Cordyceps militaris extracts. C. militaris was extracted by maceration and infusion. The extracts were investigated for cordycepin, phenolic, and flavonoid content. The antioxidant activity was investigated by in vitro spectrophotometric methods. The irritation profile was investigated by hen’s egg-chorioallantoic membrane test. Nanoemulsions were developed using high-pressure homogenizer. C. militaris extract was incorporated into the nanoemulsion and investigated for safety, release profile, permeation, and skin retention. The results demonstrated that water extract (CW) contained the significantly highest content of cordycepin, phenolics, and flavonoids, which were responsible for antioxidant activity. CW was the most potent antioxidant. CW possessed comparable 2,2′-diphenyl-1-picrylhydrazyl radical scavenging activity and lipid peroxidation inhibition to l-ascorbic acid (96.9 ± 3.1%) and alpha-tocopherol (87.2 ± 1.0%). Consequently, ten mg/mL of CW was incorporated into nanoemulsions composing of sugar squalene, Tween® 85, and deionized water. Nanoemulsion, which had the smallest internal droplet size (157.1 ± 2.6 nm), enhanced the stability of CW, had no cytotoxicity effect and no skin irritation, released the most CW (0.9 ± 0.0% w/w after 24 h), and delivered the highest CW into the skin layer (33.5 ± 0.7% w/w). Therefore, nanoemulsion was suggested for enhancing the stability and dermal delivery of CW.

Micelles of Progesterone for Topical Eye Administration: Interspecies and Intertissues Differences in Ex Vivo Ocular Permeability.

Progesterone (PG) may provide protection to the retina during retinitis pigmentosa, but its topical ocular supply is hampered by PG poor aqueous solubility and low ocular bioavailability. The development of efficient topical ocular forms must face up to two relevant challenges: Protective barriers of the eyes and lack of validated ex vivo tests to predict drug permeability. The aims of this study were: (i) To design micelles using Pluronic F68 and Soluplus copolymers to overcome PG solubility and permeability; and (ii) to compare drug diffusion through the cornea and sclera of three animal species (rabbit, porcine, and bovine) to investigate interspecies differences. Micelles of Pluronic F68 (3–4 nm) and Soluplus (52–59 nm) increased PG solubility by one and two orders of magnitude, respectively and exhibited nearly a 100% encapsulation efficiency. Soluplus systems showed in situ gelling capability in contrast to the low viscosity Pluronic F68 micelles. The formulations successfully passed the hen’s egg-chorioallantoic membrane test (HET-CAM) test. PG penetration through rabbit cornea and sclera was faster than through porcine or bovine cornea, although the differences were also formulation-dependent. Porcine tissues showed intermediate permeability between rabbit and bovine. Soluplus micelles allowed greater PG accumulation in cornea and sclera whereas Pluronic F68 promoted a faster penetration of lower PG doses.

Assessment of the photoprotective potential and toxicity of Antarctic red macroalgae extracts from Curdiea racovitzae and Iridaea cordata for cosmetic use

Botanical and marine extracts have been used in cosmetic products as natural antioxidants or UV-absorbers to reduce the use of organic UV filters. However, despite the high UV exposure of Antarctic macroalgae that induces the production of photoprotective compounds, the use of marine macroalgae extracts for photoprotection has been little explored. Thus our objective was to assess the photoprotective potential and the toxicity of extracts from two red macroalgae Curdiea racovitzae and Iridaea cordata. Chemical characterization was based on the qualitative and quantitative analyses of mycosporine-like amino acids (MAAs) by high performance liquid chromatography coupled to mass spectrometry. Photoprotective properties were assessed by UV spectrophotometry and photostability assay. Antioxidant capacity was studied by DPPH and superoxide radical scavenging assays. UVB and UVA photoprotection was assessed in keratinocytes by cell viability analysis after UVB irradiation and quantification of intracellular reactive oxygen species (ROS) induced by UVA, respectively. Toxicity was estimated by cytotoxicity, phototoxicity and HET-CAM (Hen's egg test-chorioallantoic membrane) assays. Red algae extracts had high concentrations of MAAs, mainly consisted of palythine. The extracts absorbed in the UVB and UVA-II and were photostable. The C. racovitzae extract showed antioxidant activity against superoxide radicals. In addition, the C. racovitzae (1.0 and 0.5 mg/mL) and I. cordata (1.0 mg/mL) extracts had an UVB photoprotection similar to benzophenone-3, which was dependent on the total MAAs concentration. Besides, the C. racovitzae extract decreased ROS generation by 31 and 24% at 100 and 50 μg/mL, respectively. Furthermore, it was more effective in reducing ROS generation than quercetin and Helioguard 365®. Finally, both extracts were considered to be non-cytotoxic, non-phototoxic and non-irritant. Therefore, the C. racovitzae extract is a good candidate to be added to anti-aging and sunscreen formulations in order to protect against oxidative stress and to improve the absorption spectra of UV filters.

Skin irritation potential of cosmetic preservatives: An exposure-relevant study.

Preservatives represent one of the main causes of skin irritation and contact allergies. To comprehensively evaluate the skin irritation potential of phenoxyethanol, methylparaben, propylparaben, imidazolidinyl urea, and DMDM hydantoin under regulatory acceptable concentrations. A patch test and repeated open application test (ROAT) were applied to evaluate skin irritation in vivo. In vitro alternative methods consisting of the keratinocyte cytotoxicity assay, red blood cell (RBC) test, and hen's egg test-chorioallantoic membrane (HET-CAM) were performed to elucidate the mechanism of preservative-induced irritation responses. The patch test showed that all test substances showed a weak erythema response. Propylparaben had the highest occlusive irritancy potential in the patch test, owing to damage to the cell membrane. The two formaldehyde releasers showed noticeable skin irritation potential in the ROAT through their cytotoxicity to keratinocytes, while a visible response was observed after applying phenoxyethanol and the two parabens. No filtration was noticed in the in vivo tests, which might be attributed to the failure of subcutaneous vessel alteration by the preservatives. Commonly used cosmetic preservatives have minor skin irritation potential with mild erythema reaction under practical use, especially formaldehyde releasers and propylparaben.

In vivo PET/MRI Imaging of the Chorioallantoic Membrane

The Hen's Egg Test Chorioallantoic-Membrane (HET-CAM) of fertilized chick eggs represents a unique model for biomedical research. With its steadily increasing use, non-invasive in ovo imaging for longitudinal direct quantification of the biodistribution of compounds or monitoring of surrogate markers has been introduced. The full range of imaging methods have been applied to the HET-CAM model. From the current perspective, Magnetic Resonance Imaging (MRI), providing detailed anatomical and functional data, and Positron Emission Tomography (PET), providing excellent sensitivity, must be considered the most promising candidates for HET-CAM imaging. Especially by combining both techniques, the required sensitivity and anatomical localization of the signal source renders feasible. In the following a review of recent applications of MRI and PET for in ovo imaging with a special focus on techniques for imaging xenotransplanted tumors on the CAM will be provided.

Enhanced Transdermal Delivery of Concentrated Capsaicin from Chili Extract-Loaded Lipid Nanoparticles with Reduced Skin Irritation.

The aim of this study was to develop lipid-based nanoparticles that entrapped a high concentration of capsaicin (0.25%) from a capsicum oleoresin extract. The solid lipid nanoparticles (SLNs) and nanostructured lipid carriers (NLCs) were strategically fabricated to entrap capsaicin without a hazardous solvent. Optimized nanosize lipid particles with high capsaicin entrapment and loading capacity were achieved from pair-wise comparison of the solid lipid mixtures consisting of fatty esters and fatty alcohols, representing small and large crystal-structure molecules combined with a compatible liquid lipid and surfactants (crystallinity index = 3%). This report was focused on selectively captured capsaicin from oleoresin in amorphous chili extract-loaded NLCs with 85.27% ± 0.12% entrapment efficiency (EE) and 8.53% ± 0.01% loading capacity (LC). The particle size, polydispersity index, and zeta potential of chili extract-loaded NLCs were 148.50 ± 2.94 nm, 0.12 ± 0.03, and −29.58 ± 1.37 mV, respectively. The favorable zero-order kinetics that prolonged capsaicin release and the significantly faster transdermal penetration of the NLC attributed to the reduction in skin irritation of the concentrated capsaicin NLCs, as illustrated by the in vitro EpiDermTM three-dimensional human skin irritation test and hen’s egg test chorioallantoic membrane assay (HET-CAM).

Multi-Modal PET and MR Imaging in the Hen’s Egg Test-Chorioallantoic Membrane (HET-CAM) Model for Initial In Vivo Testing of Target-Specific Radioligands

The validation of novel target-specific radioligands requires animal experiments mostly using mice with xenografts. A pre-selection based on a simpler in vivo model would allow to reduce the number of animal experiments, in accordance with the 3Rs principles (reduction, replacement, refinement). In this respect, the chick embryo or hen’s egg test–chorioallantoic membrane (HET-CAM) model is of special interest, as it is not considered an animal until day 17. Thus, we evaluated the feasibility of quantitative analysis of target-specific radiotracer accumulation in xenografts using the HET-CAM model and combined positron emission tomography (PET) and magnetic resonance imaging (MRI). For proof-of-principle we used established prostate-specific membrane antigen (PSMA)-positive and PSMA-negative prostate cancer xenografts and the clinically widely used PSMA-specific PET-tracer [68Ga]Ga-PSMA-11. Tracer accumulation was quantified by PET and tumor volumes measured with MRI (n = 42). Moreover, gamma-counter analysis of radiotracer accumulation was done ex-vivo. A three- to five-fold higher ligand accumulation in the PSMA-positive tumors compared to the PSMA-negative tumors was demonstrated. This proof-of-principle study shows the general feasibility of the HET-CAM xenograft model for target-specific imaging with PET and MRI. The ultimate value for characterization of novel target-specific radioligands now has to be validated in comparison to mouse xenograft experiments.

Self-nanoemulsifying System Optimization for Higher Terconazole Solubilization and Non-Irritant Ocular Administration.

Purpose: Eye drops’ formulations of poorly water-soluble drugs, offer the advantage of crossing the lipophilic cornea, but their limited aqueous solubility may lead to low ocular bioavailability limiting their therapeutic uses. Terconazole (TZ) is an antifungal drug with low aqueous solubility, restricting its application in ocular fungal infection. Thus, the aim of the work in this study is to enhance TZ solubilization, permitting better ocular permeation and higher bioavailability. To achieve this goal, different self-nanoemulsifying systems (SNESs) were prepared using different oils, surfactants and co-surfactants. Methods: Ternary phase diagrams were constructed to identify self nano-emulsification regions for each oil system examined; either Labrafil® M2125CS or Capryol™ 90. TZ saturated solubility in the different formulated systems were measured and systems showing highest potential for TZ solubilization were selected. The optimized systems were chosen based on their globule size, polydispersity index, self-emulsification characteristics. Finally, TZ release as well as the irritation effect via Hen’s Egg test-chorioallantoic membrane (HET-CAM test) of the optimized system was observed in vitro. Results: The optimized system was formulated using 20% w/w Labrafil® M2125 CS, 50% w/w Tween® 80 and 30% w/w Transcutol® HP. Oil globules showed size range of 15.13 nm and self-emulsification time of 12.80 seconds. The system released 100% of the drug within half an hour compared to 2 hours in case of TZ-suspension. Finally, HET-CAM test showed non-irritating response and normal vascularization of the chorioallantoic membrane. Conclusion: The formulated SNES could be a promising approach to enhance ocular efficacy of TZ.

Delineating penetration enhancer-enriched liquid crystalline nanostructures as novel platforms for improved ophthalmic delivery.

Liquid crystalline nanostructures (LCNs), for instance cubosomes, have been widely used as a promising carrier for drug delivery through the last few years. To date, the ophthalmic application of these platforms was not well explored, and the effect of integrating penetration enhancers (PEs) into LCNs has not been investigated yet. Hence, the present work aimed coupling novel PEs into glyceryl monooleate-based cubosomes for ocular administration. Various enhancers viz, free fatty acids (oleic and linoleic acids), natural terpenes (D-limonene and cineole), medium-chain triglycerides (Captex® 1000 and Captex® 8000), mono-/di-glycerides (Capmul® MCM, Capmul® PG-8, and Capmul® PG-12) were tested at different amounts. The morphology of the formed LCNs was investigated using transmission electron microscopy (TEM). The crystallinity and thermal behavior studies were also conducted. The ocular safety of optimized formulae was tested via hen's egg test-chorioallantoic membrane (HET-CAM), rabbit eye Draize test, and histopathological examinations of ocular tissues. Confocal laser scanning microscopy (CLSM) was utilized to assess the enhanced permeation of fluorescently-labeled LCNs across corneal layers. The acceptable formulations exhibited relatively homogenous particle nano-sizes ranging from 139.26±3.68 to 590.56±24.86nm carrying negative surface charges. TEM images, X-ray patterns and DSC thermograms demonstrated the influential effect of PEs in developing altered crystalline structures. The ocular compatibility of optimized LCNs was confirmed. The corneal distribution using CLSM proved the disseminated fluorescence intensity of LCNs enriched with oleic acid, Captex® 8000 and Capmul® MCM. Selected LCNs showed good physical stability upon storage and lyophilization. The results demonstrated the efficiency of tailored PE-modified LCNs in enhancing the ocular transport with no evidence of any irritation potential, and hence suggested their prospective applicability in ophthalmic drug delivery.

Synthesis, Characterization, Self-Assembly, and Irritation Studies of Polyglyceryl-10 Caprylates.

1,4-dioxane should be less than or equal to 10 ppm in finished cosmetic products according to the recommendation of the Scientific Committee on Consumer Safety, but it is often generated as a by-product during the manufacturing process of poly(ethylene glycol) (PEG)-based derivatives. In order to avoid the possible risk caused by 1,4-dioxane, it might be a good choice for preparing cosmetic ingredients by using polyglycerin (PG) instead of PEG as a hydrophilic segment. In the present study, polyglyceryl-10 caprylates were synthesized by the esterification reaction between polyglycerin-10 and caprylic acid. FTIR and 1H NMR were utilized to confirm the chemical structures of the obtained polyglyceryl-10 caprylates. Light transmittance was availed to investigate the water solubility of polyglyceryl-10 caprylates. The self-assembly behavior, size, and size distribution of polyglyceryl-10 caprylates were investigated by dynamic light scattering. The makeup cleansing effect was also evaluated by in vitro and in vivo methods. Irritation was evaluated by hen’s egg test-chorioallantoic membrane assay (HET-CAM). Results showed that polyglyceryl-10 monocaprylate could self-assemble into nanoparticles in the water at the concentration range of 2.5–10 wt% with a transparent appearance. The diameter of formed nanoparticles was around 100 nm with a narrow particle size distribution around 0.1 at the concentration of 2.5 wt% or 5 wt%. Polyglyceryl-10 monocaprylate exhibited good removal effect against makeup and excellent removal efficacy against pen eyeliner. The irritation of polyglyceryl-10 monocaprylate evaluated by HET-CAM at the concentration of 4 wt% was moderate irritant (irritation score = 8.4), which was lower than that of PEG-6 caprylic/capric glycerides (severe irritant, irritation score = 14.1). Therefore, polyglyceryl-10 monocaprylate might be a promising cosmetic ingredient for transparent makeup removing water.

The Chemical Composition, Antioxidant and Anti-Tyrosinase Activities, and Irritation Properties of Sripanya Punica granatum Peel Extract

This is the first study to investigate the effect of ethanol concentrations on the phenolic content and biological activities of Sripanya Punica granatum peel extract. Various concentrations of ethanol, (20 to 80% v/v), were used in the maceration process. Extracts were investigated for total phenolic content by the Folin–Ciocalteu method, antioxidant activities by 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2’-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assay, and anti-tyrosinase activity by a spectrophotometric method. The chemical composition was investigated using high-performance liquid chromatography. The irritation of the extracts was determined by Hen’s egg test–chorioallantoic membrane assay. The results demonstrated that E60, which was extracted using 60% v/v ethanol, had the highest yield (66.23 ± 0.38% w/w), the highest total phenolic content (490.42 ± 7.20 μg GAE/g), and the highest antioxidant and anti-tyrosinase activities. IC50 values on DPPH• and ABTS•+ inhibition were 10.97 ± 0.97 and 6.48 ± 0.06 µg/mL, respectively. The 60% v/v ethanol extract (E60) possessed strong tyrosinase inhibition with an IC50 value of 0.10 ± 0.13 µg/mL, which is more potent than kojic acid. The major constituents were punicalagin (4.05 ± 0.26% w/w) and ellagic acid (0.63 ± 0.04% w/w). E60 was safe for topical application since it did not induce irritation. Therefore, we suggest that E60 is a promising agent for further product development.

Diphenyl Diselenide and Clotrimazole Co-loaded into Eudragit® RS 100 Nanocapsules Formulation Has Superior Antioxidant Potential and Promising Anti-candida Activity

In the current study, nanocapsules (NC) formulations containing a co-load of clotrimazole (C), a highly prescribed antifungal drug, and diphenyl diselenide [(PhSe)2], an organoselenium compound with a promising scope of pharmacological actions, were prepared. Formulations were characterized as well as the potential toxicity, antioxidant action, and antifungal effect were assessed using in vitro techniques. The NCs were prepared employing Eudragit® RS 100 as polymeric wall and medium chain triglycerides or virgin coconut oil (CO) as core. All NC suspensions had pH around acid range, compound content close to theoretical value (1 mg/mL/drug), average diameter in nanometric range, positive values of zeta potential as well as high encapsulation efficacy and mucoadhesive property. Physicochemical stability was performed over a 30-day period and showed no modification in the aforementioned parameters to all samples. Preliminary screening of toxicological potential performed by the hen’s egg test chorioallantoic membrane technique classified the formulations as non-irritant. The DPPH radical assay revealed that nanoencapsulated compounds had superior antioxidant action in comparison to their free forms (concentration range tested 1.0-25.0 µg/mL). Importantly, the formulation composed of CO and containing C and (PhSe)2 showed the highest antioxidant potential and was selected for further investigation regarding antifungal effect against some Candida spp strains. Results of in vitro antifungal assay demonstrated that the C and (PhSe)2 co-encapsulation had a minimum inhibitory concentration (MIC) values around 60. Thus, our study supplies additional data about advantages achieved by encapsulating active compounds.

Development of thermosensitive in situ gel nanoemulsions for ocular delivery of acyclovir

The aim of the current study was to formulate thermosensitive in situ gel nanoemulsions (NEs) containing acyclovir (ACV) for ocular drug delivery and evaluate their ex vivo corneal permeation ability and ocular irritation possibility. ACV loaded NEs were prepared by low energy method. Physicochemical properties, drug release, and stability studies of in situ gel NEs were investigated. The optimized formulation was chosen for excised bovine corneal permeation, hen's egg test-chorioallantoic membrane and modified Draize tests to evaluate its ocular permeation and irritation. The optimum in situ gel NE consisting Triacetin, Transcutol®P, Poloxamer 407, and Poloxamer188 had the mean droplet size and polydispersity index of 28 nm and 0.38 respectively. Moreover, physicochemical properties such as pH, viscosity, osmolality, and refractive index were acceptable for ocular administration. This formulation showed a sustained drug release pattern compared to the control solution. Based on the permeation study results, drug permeation of optimum formulation was about 2.8-fold higher than the control solution. Irritation tests illustrated that the optimum in situ gel NE could be well tolerated by the eye. In conclusion, these results demonstrated that this new ACV formulation could be considered as a novel ophthalmic delivery system for the treatment of ocular viral infections.

The proteolytic fraction from Vasconcellea cundinamarcensis accelerates wound healing after corneal chemical burn in rabbits.

Chemical ocular burns are among the most frequently eye-related injuries, which require immediate and intensive evaluation and care since they may lead to potential complications such as superinfection, corneal perforation, and blindness.Vasconcellea cundinamarcensis, a species from Caricaceae family, contains highly active proteolytic enzymes in its latex that show healing activity in animal models bearing lesions of different etiologies. We evaluate the ocular toxicity of the proteolytic fraction from V. cundinamarcensis (P1G10) by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and Hen's Egg Test-Chorioallantoic Membrane test. The corneal healing property of P1G10 was studied by the ethanol-chemical burn in the rabbit's eyes. P1G10 is safe for ocular administration, except when administrated at 10μg/mL. P1G10 at 1μg/mL accelerates the corneal re-epithelization achieving complete wound closure after 72h of chemical burn. Also, P1G10 modulated the inflammatory response and controlled the arrangement of collagen fibers in the stroma, demonstrating its potential corneal healing properties. Our work was the first one to evaluate the ophthalmic application of P1G10. Here we demonstrated that P1G10 is suitable for ocular administration and it has a promising corneal healing activity which may emerge as a new pharmacological tool to the development of a new drug for ocular surface chemical injuries in the future.

Synthesis, characterization and toxicity assessment of a new polymeric nanoparticle, l-glutamic acid-g-p(HEMA)

The toxic effects of poly(HEMA)-based polymeric nanoparticles must be analyzed before their biomedical applications as drug delivery systems. The aim of the study was to characterize and evaluate the toxicity for its biocompatibility of a newly synthesized l-glutamic acid-g-p(HEMA) polymeric nanoparticle The nanoparticle was synthesized with surfactant-free emulsion polymerization and grafting techniques. Grafting efficiency was estimated at 58%. The nanoparticle shape was verified as nearly spherical by scanning electron microscopy. Atomic force microscopy images showed a rough surface topography. The nanoparticle had an average size of ~194.6 nm on zeta analysis, and the zeta potential value was −18 mV. Fourier transformed infrared spectroscopy revealed spectra from 750 to 4000 cm−1 and characteristic peaks of stretching bands. The swelling ratio was 46%. With 24-h exposure, p(HEMA) and l-glutamic acid-g-p(HEMA) did not have cytotoxic effects on a human bronchial epithelial cell line (16HBE) and human monocyte cell line by water-soluble tetrazolium salt 1 (WST-1) assay and lactate dehydrogenase assay (LDH). It did not show genotoxic potential by comet assay and did not have mutagenic effects on Salmonella typhimurium TA98, TA100, TA1535 and TA1537 strains by Ames test. The nanoparticle at 160 μg/ml showed 2% hemolytic activity on erythrocytes. On cell migration assay, the percentage closure difference between exposed and control cells was estimated at 21%. We found no irritation effect on Hen's egg test-chorioallantoic membrane test. We determined that the polymeric nanoparticle l-glutamic acid-g-p(HEMA) was biocompatible and has potential for use in a drug delivery system.

Design, synthesis, and evaluation of novel 2-substituted 1,4-benzenediol library as antimicrobial agents against clinically relevant pathogens

Development of new antimicrobial agents, capable of combating resistant and multidrug-resistant fungal and bacterial clinical strains, is necessary. This study presents the synthesis and antimicrobial screening of 42 2-substituted-1,4-benzenediols, being 10 novel compounds. In total, 23 compounds showed activity against fungi and/or bacteria. Benzenediol compounds 2, 5, 6, 8, 11, and 12 demonstrated broad spectrum antimicrobial actions, including resistant and multidrug-resistant species of dermatophytes (Trichophyton mentagrophytes), Candida spp. and the ESKAPE panel of bacteria. Minimum inhibitory concentrations of these compounds for fungi and bacterial strains ranged from 25 to 50 µg/ml and 8–128 µg/ml, respectively. The antifungal mechanism of action is related to the fungal cell wall of dermatophytes and membrane disruption to dermatophytes and yeasts, in the presence of compound 8. Specific structural changes, such as widespread thinning along the hyphae and yeast lysis, were observed by scanning electron microscopy. The effects of compound 8 on cell viability are dose-dependent; however they did not cause genotoxicity and mutagenicity in human leukocyte cells nor haemolysis. Moreover, the compounds were identified as nonirritant by the ex-vivo Hen’s egg test-chorioallantoic membrane (HET-CAM). The furan-1,4-benzenediol compound 5 showed in vivo efficacy to combat S. aureus infection using embryonated chicken eggs. Therefore, the compounds 8, and 5 are promising as hits for the development of new antimicrobial drugs with reduced toxicity.

Ocular Tolerability and Efficacy of Ebastine Colloidal Formulation in Allergic Conjunctivitis

Allergic conjunctivitis (AC), defined by ocular itching, hyperemia, lacrimation and edema, impairs the quality of life across the globe. Ebastine is available as an oral antihistamine formula, such as tablets and syrup, for allergic disorders. Topical antihistamines are preferred over oral agents since their direct application at the site of action results in rapid onset and superior efficacy with less systemic side effects. The objective of the present work was to evaluate the antiallergic potential of optimized ebastine (1% w/v) colloidal ocular formulation by performing in vitro study like hen's egg chorioallantoic membrane test (HET-CAM) for tolerability and in vivo efficacy study in ovalbumin (OA)-induced allergic conjunctivitis (AC) with acute ocular irritation study. Eye scratching behavior and edema were evaluated after topical antigen challenge. Edema was scored at periodic interval after the instillation of ovalbumin followed by histopathology. The results showed that ebastine (1% w/v) colloidal ocular formulation was effective in inhibiting symptoms of eye inflammation induced by ovalbumin. Further, the study indicated that said formulation has a quick onset and the duration of effect sufficient to provide relief from symptoms for 24 hr. Ocular irritation by HET-CAM assay showed that the developed formulation does not cause any irritation to the blood vessels. Acute ocular irritation test was performed using rabbits and results showed that developed formulation was non-irritant to the eye. The present study revealed that the ocular ebastine formulation could offer a novel therapeutic opportunity against IgE-mediated allergic conjunctivitis.