Goat Cornea Research Articles

Design and evaluation of guar gum-based ofloxacin sustained release ocular insert

To prepare ocular inserts of ofloxacin using guar gum as a polymer for sustained delivery over a period of 24 h.Ofloxacin ocular inserts were prepared by the solvent casting method using guar gum in different proportions (0.5% w/v, 0.75% w/v and 1.0% w/v). The prepared formulations were evaluated for thickness, weight variation, percentage drug content, surface pH, folding endurance, percentage moisture absorption and loss, percentage swelling, mechanical strength and in vitro transcorneal permeation. In vitro transcorneal permeation study was performed on goat cornea using a modified Franz diffusion cell. The inserts were found to be of uniform thickness (ranging from 51.230 ± 0.385 μm to 109.275 ± 0.522 μm) and weight (1.720 ± 0.079 mg to 3.402 ± 0.105 mg). The % drug content in the inserts was found to vary between 95.450 ± 0.427% and 98.471 ± 0.225.The cumulative % drug releases from the formulation ranged from 38.19 to 75.21 over a period of 24 h. All the formulations followed a zero order release pattern. The in vitro transcorneal study revealed that an increase in concentration of the polymer slowed down the release of ofloxacin from the formulation.Ocular inserts using guar gum as a polymer were successfully prepared and can be effectively used for sustained ocular delivery over a period of 24 h.

Carboxymethyl tamarind kernel polysaccharide nanoparticles for ophthalmic drug delivery

Tropicamide-loaded carboxymethyl tamarind kernel polysaccharide (CMTKP) nanoparticles were prepared and evaluated for ocular delivery. Preparation of ionotropically gelled CMTKP nanoparticles was optimized employing three-levels, two-factor central composite design. Concentration of polymer and crosslinker had significant synergistic effect on particle size and % encapsulation efficiency. The optimal calculated parameters were concentrations of CMTKP 0.10% (w/v) and calcium chloride 0.11% (w/v). The optimized tropicamide-loaded CMTKP formulation showed ex vivo corneal permeation of tropicamide across isolated goat cornea comparable to its aqueous solution. Further, the mucoadhesive and non-irritant nature of CMTKP nanoparticles indicate their suitability as ocular delivery system.

Thiolated pectin nanoparticles: Preparation, characterization and ex vivo corneal permeation study

Abstract The objective of present study was to prepare thiolated pectin nanoparticles and to evaluate them for ocular delivery. Thiolated pectin nanoparticles were prepared by ionotropic-gelation technique using magnesium chloride as the ionic cross-linker and timolol maleate as the model drug. The results revealed that increasing the concentration of magnesium chloride results in significant increase in particle size, while % entrapment is decreased significantly by increase in the concentration of thiolated pectin. The optimal formulation having particle size of 237 nm and % entrapment of 94.6% was obtained at concentrations of thiolated pectin – 0.01% (w/v) and magnesium chloride – 0.01% (w/v). On comparative evaluation, thiolated pectin nanoparticulate formulation provided significantly higher ex vivo corneal permeation of timolol maleate across the excised goat cornea than the conventional aqueous solution.

Influence of acetazolamide loading on the (in vitro) performances of non-phospholipid-based cationic nanosized emulsion in comparison with phospholipid-based anionic and neutral-charged nanosized emulsions

Context: Acetazolamide (ACZM)-loaded anionic, cationic, and neutral-charged oil-in-water nanosized emulsions were prepared and compared with their mean droplet diameter, surface charge, entrapment efficiency, freeze–thaw cycling stability, in vitro drug release, and transcorneal permeation.Objective: The present study aims to determine the influence of ACZM loading on the performances of non-phospholipid-based cationic nanosized emulsion in comparison with phospholipid-based anionic and neutral-charged nanosized emulsions.Results and discussion: Regardless of charges, all of these emulsions exhibited a nanometer range mean particle diameter (240–443 nm) following autoclave sterilization. While the anionic and cationic emulsions did show high negative (−36.9 mV) and positive zeta potential (+41.4 mV) values, the neutral-charged emulsion did not. Presence of cryoprotectants (5% w/w sucrose + 5% w/w sorbitol) improved the stability of cationic emulsion to droplet aggregation during freeze–thaw cycling. The in vitro release kinetic behavior of drug exchange with physiological anions present in the simulated tear solution appears to be complex and difficult to characterize using mathematical fitting model equations. Augmentation in drug permeation through goat cornea, in vitro, was noticed for cationic emulsion.Conclusion: ACZM-loaded cationic nanosized emulsion could be suitable for topical application into eye to elicit better therapeutic effect in comparison with its anionic and neutral-charged emulsions.

Chitosan Coated PLA Nanoparticles for Ophthalmic Delivery: Characterization,<I>In-Vitro</I> and <I>In-Vivo</I> Study in Rabbit Eye

The objective of the present study is to develop a poly (D, L-lactic acid) (PLA) nano-carrier for topical ocular applications. PLA nanoparticles (PLA-NPs) with 5-fluorouracil were prepared using varying concentration and molecular weight of PLA to regulate the particle size. The dimension and shape of nanoparticles were verified by using dynamic light scattering (DLS), atomic force microscope (AFM) and scanning electron microscope (SEM). Ex-vivo permeation study was conducted by goat and rabbit excised cornea. In-vivo experiment was conducted in rabbit eye and 5-FU concentration was measured in aqueous and vitreous humor by HPLC. In-vitro experiments indicated a diffusion controlled release of 5-FU. No significant interaction was observed in between mucin and PLA NPs that measured in terms of viscosity change. Ex-vivo permeation was significantly higher with rabbit cornea as compared to goat cornea. PLA and CH-PLA DNPs showed increased level of 5-FU as comparison to 5-FU solution. In-vivo study showed significantly higher concentration in case of uncoated and CH coated PLA nanoparticles in rabbit eye as compared to free 5-FU solution. PLA nanoparticle was found non-irritant in nature by modified Draize test.

Functional characterization of L-tryptophan transport across mammalian cornea.

In last few years transporter targeted drug delivery has drawn attention of research to identify and explore various nutrient transport systems including amino acid transporters for better drug delivery. The aim of present research work is to investigate the transport characteristics of L-tryptophan (L-try) across goat cornea. Transport of L-try was investigated using a glass diffusion cell for effect of concentration, pH, presence of other amino acids or metabolic inhibitor or dipeptide and tripeptide. The amount of L-try transported increased as the pH of L-try aqueous solution increased from 5 to 9. Inhibition was observed in L-try transport in absence of sodium ions where L-try solution was made isotonic with dextrose. Amino acids like L-histidine, L-arginine, L-lysine (cationic), L-glutamic acid, L-aspartic acid (anionic), glycine and L-proline (neutral) inhibited the L-try transport as compared to control (L-try alone). In presence of sodium azide and Ouabain the inhibition in L-try transport across goat cornea was observed while no marked inhibition was observed on L-try transport across goat cornea in presence of aspartame and glutathione. The L-try transport was favored up to concentration 1% w/v and at higher pH in presence of sodium ions through excised goat cornea. Functional presence of a sodium dependent L-try transport system as inhibited by ouabain having affinity to cationic and neutral amino acid is evident on goat cornea. Keywords: Cornea; Amino acid; Tryptophan; Transport

Preparation and evaluation of nanoparticles of gum cordia, an anionic polysaccharide for ophthalmic delivery

The purpose of present investigation was to prepare nanoparticles using gum cordia as the polymer and to evaluate them for ophthalmic delivery of fluconazole. A w/o/w emulsion containing fluconazole and gum cordia in aqueous phase, methylene chloride as the oily phase, and di-octyl sodium sulfosuccinate and polyvinyl alcohol as the primary and secondary emulsifiers, respectively, was cross-linked by ionic gelation technique to produce fluconazole-loaded nanoreservoir system. The formulation of nanoparticles was optimized using response surface methodology. Multiple response simultaneous optimizations using the desirability approach were used to find optimal experimental conditions. The optimal conditions were found to be concentrations of gum cordia (0.85%, w/v), di-octyl sodium sulfosuccinate (9.07%, w/v) and fluconazole (6.06%, w/v). On comparison of the optimised nanosuspension formulation with commercial formulation, it was found to provide comparable in vitro corneal permeability of fluconazole across isolated goat cornea, indicating suitability of nanosuspension formulation in ophthalmic delivery of fluconazole.

Effect of pH and Formulation Variables on In Vitro Transcorneal Permeability of Flurbiprofen: A Technical Note

Most of the topically applied drugs have to enter the eye through the corneal route and therefore epithelial and stromal layer of the cornea act as barrier to transcorneal permeability of both hydrophilic and lipophilic drugs respectively. Paracellular and transcellular pathways are the main two routes for corneal drug transport. Lipophilic drugs cross the cornea through transcellular pathway while the hydrophilic drugs cross through paracellular pathway (1,2). It has been reported based on studies involving different series of compounds that the fraction absorbed through corneal route varies from less than 1% for hydrophilic drugs to 7% for lipophilic drugs (1–3). Corneal epithelium and stroma exert varying degree of resistance to penetration depending on the nature of the drug (4). A corneal permeability coefficient of the order of 0.1 × 10−5 to 4.0 × 10−5 cm/s can be considered a measure of efficient corneal drug permeation (5). Corneal permeability increases when the corneal integrity is compromised by the usage of high concentration of certain formulation excipients like, preservatives and chelating agents (5). Rabbit cornea or eye is most commonly used for in vitro and in vivo studies on ocular drug delivery. An investigation on the mammalian corneal epithelium in different species including human, rabbit and pig revealed morphological similarity of corneal cell layers (6). Nevertheless dissimilarities do exist between these species in terms of response of the eye to irritation and trauma (7). Rabbit eyes were found to be more sensitive to irritant than other species. Reer and his coworkers have suggested that rabbit cornea is not always the model of choice and the great morphological uniformity of mammalian cornea allow a different model like goat cornea to be chosen for in vitro permeation studies (8). Goat corneal membrane has been used for in vitro transcorneal permeability studies of ketrolac tromethamine from aqueous drops, (9) oil based drops and ointments (10). Effect of preservatives on in vitro transcorneal permeability of ibuprofen and flurbiprofen across goat cornea has also been reported (11). Taking the above reports in view, goat cornea was chosen for in vitro permeation studies. A practical advantage of goat cornea is its easy availability so that larger number of experiments could be performed in a shorter time period. The present study was aimed to investigate the effect of various formulation variables in ophthalmic formulation on the in vitro transcorneal permeability of flurbiprofen in excised goat cornea using modified Franz diffusion cell. Effect of pH of the buffer, pH of the unbuffered solution and formulation excipients like, preservatives and chelating agents on the trans-corneal permeability of the drug was investigated. To support transcorneal permeability data, solubility studied were carried out in buffered and unbuffered systems. Also investigated was the transcorneal permeability of flurbiprofen from various 5% w/v gels made of viscosity enhancing polymers and from various vegetable oil vehicles like, olive oil, linseed oil and sesame oil. All the animal experiment protocols were approved by Institutional Animal Ethics Committee, BITS, Pilani, India.

Effect of Formulation Parameters on Corneal Permeability of Ofloxacin

Influence of pH, buffer, preservative, tonicity and viscosity modifiers on in vitrotranscorneal permeation of ofloxacin was studied using excised goat cornea. Permeation studies were carried out by putting 1 ml of test formulation on the cornea (0.78cm²) mounted between the donor and receptor compartments of an all glass modified Franz diffusion cell and measuring ofloxacin concentration in the receptor (containing bicarbonate ringer under stirring at 35°C) by spectrophotometry at 288 nm, at various time intervals up to 120 min. Buffering the formulation with phosphate buffer or use of combination of methyl and propyl paraben or benzalkonium chloride or combination of benzalkonium chloride and disodium edetate as preservative provided significant increase in the apparent corneal permeability coefficient of ofloxacin. While the use of phenyl mercuric acetate as preservative or tonicity adjustment with mannitol showed a significant decrease in apparent corneal permeability of ofloxacin. Raising the pH of test formulation from 6.4 to 7.2 or addition of hydroxyl-propyl-β-cyclodextrin or use of viscosity modifier had no significant effect on apparent corneal permeability of drug.

Transcorneal permeation of ciprofloxacin and diclofenac from marketed eye drops

The purpose of this research was to evaluate the in vitro permeation characteristics of various marketed eye drops of ciprofloxacin (0.3% w/v aqueous solution) and diclofenac (0.1% w/v aqueous solution) through isolated goat cornea. Effect of these drugs on isolated goat eye lenses was also evaluated. Permeation studies were conducted by putting 1 ml of formulation on the cornea fixed between the donor and receptor compartments of an all glass modified Franz diffusion cell and monitoring ciprofloxacin and diclofenac concentration in the receptor (containing normal saline or bicarbonate ringer solution under continuous stirring at 37±2°) spectrophotometrically at their respective absorption maxima, after 120 min. Paired isolated goat lenses (i.e. of same animal) were used to evaluate the effect of these drugs at selected concentrations against oxidative stress (1 mM hydrogen peroxide solution). After 24 h of incubation at 37°, the lens treated with test solution (hydrogen peroxide+drug in bicarbonate ringer solution) was estimated for soluble protein content and compared with control (only hydrogen peroxide). Among marketed eye drops of ciprofloxacin, Joxin (Jawa Pharmaceuticals) showed maximum in vitro transcorneal permeation (0.558%) while I-Gesic (Centaur Pharmaceuticals) showed maximum % in vitro permeation or in vitro ocular availability among diclofenac eye drops after 120 min of permeation. The soluble protein content estimation studies revealed that these drugs at selected concentrations (permeated after 120 min.) had no deleterious effect on eye lenses rather possessed protective effect, since all formulation showed more soluble protein content when compared with control.

<i>In Vitro</i> Corneal Permeation of Diclofenac from Oil Drops

In vitro transcorneal permeation of diclofenac from oil drops was studied using freshly excised goat cornea. The maximum apparent corneal permeability coefficient (Papp) was obtained with 0.2% (w/v) diclofenac drops in sesame oil followed by safflower oil, while formulation in castor oil provided minimal Papp. The addition of benzyl alcohol, a preservative, in oil drops, increased the Papp value of diclofenac. Partition experiments indicated increased partitioning of diclofenac in the aqueous phase in the presence of benzyl alcohol, and the same could be responsible for the benzyl alcohol-induced increase in Papp. The solubility of diclofenac was higher in castor, arachis, and sunflower oil. But drug permeation from 0.5-1.0% (w/v) diclofenac drops in castor oil or 0.5% (w/v) drops in arachis /sunflower oil was less than that observed with 0.2% (w/v) drops in sesame oil. Thus diclofenac 0.2% (w/v) drops in sesame oil containing 0.5% (v/v) benzyl alcohol provides maximum Papp. The formulation increased corneal hydration indicating corneal damage. Since corneal hydration is less than 83% the damage appears to be reversible. The saturation solubility of diclofenac in sesame oil at 4 degrees C is 0.33% (w/v). Hence diclofenac 0.2% (w/v) solution in sesame oil will not precipitate at 4 degrees C and therefore the chances of crystallization of diclofenac from the formulation due to climatic change leading to physical instability appear to be remote.

Effect of Formulation Factors on <i>In Vitro</i> Permeation of Diclofenac from Experimental and Marketed Aqueous Eye Drops through Excised Goat Cornea

The effect of formulation factors on permeation of diclofenac from some experimental and marketed aqueous eye drops through excised goat cornea was evaluated. Raising the pH of formulation from 6.0 to 8.0 or diclofenac concentration from 0.05 to 0.15% (w/v) or adjusting tonicity with mannitol or addition of viscolizing agent decreased apparent permeability coefficient (Papp). Formulation (pH 7.4) containing sodium metabisulfite or EDTA or combination of methyl and propyl paraben showed significantly (p<0.05) higher Papp whereas benzalkonium chloride (BAC) had no effect and sorbic acid (SA) had reduced permeation. Surprisingly marketed drops containing BAC or SA, showed significantly (p<0.05) higher Papp and decreased in the order of Difen>Voveran>NSAID>Dicol>Diclolab. Lower pH (7.1-7.3) and surface tension of drops indicating presence of surfactant, could mediate increased permeation and presence of buffer could cause irritation on in vivo instillation. The marketed formulations showed corneal hydration >83% suggesting corneal damaging potential.

Effect of formulation factors on in vitro transcorneal permeation of gatifloxacin from aqueous drops

The purpose of this research was to optimize the formulation factors for maximum in vitro permeation of gatifloxacin from aqueous drops through excised goat cornea and to evaluate the permeation characteristics of drug from selected marketed eyedrop formulations. Permeation studies were conducted by putting 1 mL of formulation on the cornea (0.67 cm(2)) fixed between the donor and receptor compartments of an all-glass modified Franz diffusion cell and measuring gatifloxacin concentration in the receptor (containing normal saline under stirring) by spectrophotometry at 291.5 nm, after 120 minutes. Raising the drug concentration of the drops increased the drug permeation but decreased the percent permeation and the in vitro ocular availability. Raising the pH of the formulation from pH 5 to 7.2 increased both the drug permeation and the in vitro ocular availability. Eyedrops containing benzalkonium chloride (BAK; 0.01% wt/vol) and disodium edetate (EDTA; 0.01% wt/vol) showed maximum permeation, followed by Zymar, BAK (0.01% wt/vol), Gatilox, Gatiquin, and Gate (statistically significant P < .05 compared with control). In vitro titration of the formulations with 0.1N NaOH indicated the presence of a buffer in Zymar (pH 6) and Gate (pH 5.8), which may cause irritation and induce lacrimation, resulting in reduced ocular availability in vivo. Thus, formulation with BAK and EDTA, which is unbuffered, has a better likelihood of being absorbed in vivo. The BAK-EDTA formulation significantly (P < .05) increased the permeation of gatifloxacin through paired excised corneas of goat, sheep, and buffalo, compared with the control formulation. The goat cornea showed the greatest increase in permeation, followed by the sheep and buffalo corneas.

Ocular hazards of the colors used during the festival-of-colors (Holi) in India—Malachite green toxicity

The objective of this study was to evaluate the nature of the colors used and their toxicity to the eye upon exposure to them during celebration of Holi (our festival-of-colors). Color powders and formulations were procured at random in and around Delhi during the festival. The green/bluish-green colors reported with the higher incidence of ocular toxicity were subjected for further evaluation. Eyewash fluid collected from the patients exposed to the colors was also subjected for analysis. This study was further extended to evaluate the corneal penetration of malachite green using goat cornea in perfusion chamber. In 16/18 color samples collected, malachite green or 4-[(4-dimethylaminophenyl)-phenyl-methyl]- N, N-dimethyl-aniline was detected at different concentrations. In the eyewash fluid of four patients, HPLC estimation confirmed the presence of malachite green at concentrations of 1.3, 0.18, 3.5 and 5.4 μg in 250 ml which was responsible for its reported toxicity. The in vitro trans-corneal penetration studies did not show any detectable amount of malachite green in effluent fluid— in vitro tissue retention studies revealed that increasing the contact time increases tissue concentration. After 2 min of exposure, the tissue concentration was significantly higher. To conclude, malachite green was extensively used in our festival of Holi and has caused severe ocular irritation with epithelial defect upon exposure, though it did not penetrate through the cornea—further in vitro and in vivo studies are required on colors used in Holi.

Effect of formulation factors on in vitro permeation of moxifloxacin from aqueous drops through excised goat, sheep, and buffalo corneas.

The purpose of this investigation was to evaluate the effect of formulation factors on in vitro permeation of moxifloxacin from aqueous drop through freshly excised goat, sheep, and buffalo corneas. Aqueous isotonic ophthalmic solutions of moxifloxacin hydrochloride of different concentrations (pH 7.2) or 0.5% (wt/vol) solutions of different pH or 0.5% solutions (pH 7.2) containing different preservatives were made. Permeation characteristics of drug were evaluated by putting 1 mL formulation on freshly excised cornea (0.50 cm2) fixed between donor and receptor compartments of an all-glass modified Franz diffusion cell and measuring the drug permeated in the receptor (containing 10 mL bicarbonate ringer at 37°C under stirring) by spectrophotometry at 291 nm, after 120 minutes. Statistical analysis was done by one-way analysis of variance (ANOVA) followed by Dunnett's test. Increase in drug concentration in the formulation resulted in an increase in the quantity permeated but a decrease in percentage permeation. Increase in pH of the solution from 5.5 to 7.2 increased drug permeation, indicating pH-dependent transport. Compared with control formulation, moxifloxacin 0.5% (wt/vol) solution (pH 7.2) containing disodium edetate (EDTA) (0.01% wt/vol) produced significantly (P<.05) higher permeation with all the corneas. Formulation with benzyl alcohol significantly (P<.05) increased permeation with buffalo cornea compared with its control. Presence of benzalkonium chloride (BAK) (0.01% wt/vol) and EDTA (0.01% wt/vol) in the formulation increased permeation to the maximum with all the corneas. The results suggest that moxifloxacin 0.5% ophthalmic solution (pH 7.2) containing BAK (0.01%) and EDTA (0.01%) provides increased in vitro ocular availability through goat, sheep, and buffalo corneas.

A histochemical comparison of human corneal stromal glycoconjugates with eight other species

Frozen sections of human, calf, rabbit, rat, cat, dog, goat, lamb, and hog corneas were stained with various lectins using an avidin-biotin-peroxidase complex to study glycoconjugates of stromal matrix. Staining of the stromal matrix and keratocytes with an alpha-galactose-specific lectin, Griffonia simplicifolia I (GSA-I) was species-dependent. The stromal matrices of cat, dog, and hog corneas invariably reacted intensely with this lectin, whereas those of the human, calf, rabbit, rat, and lamb did not react. A positive reaction with GSA-I could be abolished in each instance by preincubation of the sections with alpha-galactosidase. The stromal matrices and keratocytes of all nine species reacted positively with wheat germ agglutinin, concanavalin A, and Ricinus communis agglutinin but did not react with soybean agglutinin. Results of this study may help select an appropriate animal model for further investigate human corneal stromal glycoconjugates.