Entrapment of Ketorolac Tromethamine in Polymeric Vehicle for Controlled Drug Delivery

The most common method for applying a drug in to the eye is to formulate the drug in the form of an eye drop, but this method is not considered ideal for ocular delivery of drug because of poor bioavailability arising from precorneal loss processes, this loss of drug from the precorneal area is a net effect of drainage, tear secretion and noncorneal absorption. Following the above lead we tried to improve the ocular bioavailability by increasing the corneal contact time and the feasible way was to formulate a drug with mucoadhesive/viscosity imparting agents. The adhesive strength of various polymers on corneal surface was studied with the help of self modifi ed Franz diffusion cell and freshly excised goat/bovine cornea. The polymers hydroxypropylmethylcellulose, carboxymethylcellulose sodium, Eudragit type E/RL/RS, Carbopol ETD 2020 and Carbopol 934 National Formulary were formulated with drug, ketorolac tromethamine. The adhesive strength of polymers on corneal surface and permeation characteristics of drug through cornea were investigated by using above said formulations. Eudragit type E/RL/RS did not show any improvement in mucoadhesion, but the formulations containing Carbopol ETD 2020 and Carbopol 934 national formulary showed good mucoadhesion on corneal surface in the concentration as low as 0.75%. The mucoadhesive strength was also evaluated using the combination of Carbopol acrylates/C 10-30 alkylacrylate with allylpentaerithrital and preservative benzalkonium chloride, which also resulted in good mucoadhesion with improved against S. aureus. Compounds 4b, 4c, 4d and 4j showed moderate activity against P. aeruginosa. Compounds 4b, 4c, 4d and 4j did not possess any encouraging activity against B. subtilis.


November -December 2009
Ketorolac tromethamine is a non-steroidal antiinfl ammatory drug (NSAID), received authorization for marketing in the beginning of the nineties for the short-term management of post-operative pains.The tromethamine salt enhances its water solubility and renders the drug fairly appropriate for parenteral administration [1] .Ketorolac is a derivative of pyrrolidine carboxylic acid and is structurally related to tolmetin and zomepirac.Ketorolac, like the other NSAIDs, has analgesic, antipyretic and antiinfl ammatory activities.These three actions share a common mechanism that is the inhibition of the cyclooxygenase, a critical enzyme involved in the biosynthesis of clinical mediators, like prostaglandins, prostacyclin and thromboxanes [2] .In animal models ketorolac has exhibited an analgesic/ antiinfl ammatory ratio greater than other NSAIDs [3] .For instance, weight by weight ketorolac proved to be 50 times more potent than naproxen in analgesia models but only 3 times more potent in infl ammation models [1] .This remarkable dissociation between analgesic and antiinflammatory effects provided the basis for the development of the drug as excellent antiinfl ammatory and analgesic in clinical settings [4] .Twenty to thirty milligrams (single dose) is effective in patients with moderate to severe postoperative and corneal cancer pain.Ketorolac containing ophthalmic preparations are used frequently to treat itchy eyes caused by allergies and also bring down swelling and redness (infl ammation) that can occur after cataract surgery.
During the last two decades, the development of novel mucoadhesive dosage forms has received considerable interest.The progress in this fi eld has recently been summarized in a number of reviews [5][6][7][8][9] .Entrapment of the drug inside a polymer has been an attractive concept.Mucoadhesive polymers may fulfi ll the desirable features of a prolonged residence time at the site of drug absorption owing to increased contact with the absorbing mucosa, resulting in a steep concentration gradient to favor drug absorption, and localization in specifi ed regions to improve and enhance the bioavailability of the drug [10] .Factors such as cross-linking status, ionic modifi cation, and salt formation can signifi cantly infl uence the ability of a material to show substantial mucoadhesion in an in vitro system [11,12] .Contrary to these factors, it has been postulated that positively charged polymeric hydrogels develop additional molecular forces by electrostatic interaction with negatively charged sugar moieties of the mucosal surface [13] .The mucoadhesive polymer should have a strong hydrogen bond-forming group.For example carboxylate or hydroxyl carry a strong anionic charge and have surface energy properties which favor easy spreading onto the mucus membrane and are nontoxic, non-absorbable, and non interacting with the drug.
In view of above information we attempted to evaluate the mucoadhesive properties of different polymers and in vitro trans-corneal permeation of ketorolac and formulation containing mucoadhesive polymer [14] .
All the reagents used for present study were of AR grade and instruments used were UV spectrophotometer (Shimadzu, North america), magnetic stirrer (Precision Industries Ltd., India, New Delhi), Digital Balance (Afcoset, Bangalore, India) and a pH meter (Control Dynamics, India).
A modifi ed Franz diffusion cell [15] consisting of 10 ml glass receptor along with a glass donor cell, which also contains an outlet assembly was used for the mucoadhesion permeability studies.A side arm was allowed for sampling of the receptor cell and the receptor chamber was surrounded by a water jacket through which water at 37º was circulated from a thermostat water bath.A Teflon coated magnetic bead was placed at the bottom of the receptor cell to ensure homogencity of the receptor solution.The entire cell was clamped over a magnetic stirrer.The donor compartment represented the conjunctival sac whereas the receiver compartments represented the anterior segment of the eye.Intravenous infusion set was used to control the flow of the saline bicarbonates ringer over cornea to mimic the fl ow of the tears.
Fresh whole goat and bovine eyeballs were collected *Address for correspondence E-mail: paliwalsarvesh@yahoo.com corneal permeation.Observations made in this study indicate the potentiality of the ophthalmic formulations containing mucoadhesive/viscosity imparting agents.

Key words: Corneal permeation, controlled drug delivery, goat eye, ketorolac tromethamine, polymeric vehicle
within an hour of slaughtering of the animals and the cornea were excised along with 2-4 mm of scleral tissue and washed carefully with cold (4°) normal saline.
The drug (500 mg) was dissolved in 100 ml of water to get 0.5% w/v solution.One millilitre of this solution was diluted with 2 ml of methanol and 22 ml of 0.1N HCl in a volumetric fl ask, then 1 ml of this solution was further diluted to 25 ml with 0.1N HCl in a volumetric fl ask to give the fi nal stock solution.From this solution various dilutions were made in order to get different concentration ranging from 0.32 µl/ml to 8 µl/ml and were used for the preparation of standard curve at 313 nm.
Eudragit E/RL/RS, Carbopol 934NF, Carbopol ETD 2020, carboxymethylcellulose sodium (CMC), hydroxypropylmethylcellulose (HPMC) were considered for the present study.Different concentrations of polymers were prepared in 0.8% saline solution and stirred continuously with the help of mechanical stirrer until homogenous dispersions were obtained [16] .After preparation of polymeric solution, ketorolac tromethamine was added and pH of the solution was adjusted to the desired level.Preservative benzalkonium chloride (BKC) (0.01%) and EDTA (0.01%) were also added to the above solution.
The freshly isolated and washed cornea was initially weighed and mounted between the donor and receptor chamber of the diffusion cell by sandwiching the scleral layer between the rims of the two chambers in such a way so that the epithelial surface faced the donor chamber.Bicarbonate ringer was fi lled in the receptor chamber.It included solution A (100 ml): NaCl-1.24g, KCl-0.071g, NaH 2 PO 4 -0.02 g, NaHCO 3 -0.49g and solution B (100 ml): CaCl 2 -0.023 g, MgCl 2 -0.031 g.Equal quantity of solution A and B were mixed to get bicarbonate ringer and allowed to equilibrate the tissue, any air bubble in the receptor chamber was removed by inverting the diffusion cell.The drug polymer solution was placed on the epithelial surface of the cornea through the donor chamber.Bicarbonate ringer solution of pH 7.2 was allowed to flow over the cornea at a flow rate of 12.5 µl/min from an I.V. infusion set.A sample of 1.0 ml was withdrawn from the receptor chamber at 30, 60, 90 and 120 min intervals and diluted with one ml of 0.1N HCl.The samples after dilution were analyzed for ketorolec content by measuring the absorbance at 313 nm in a UV spectrophotometer.The concentrations were determined with the help of a standard curve.
The corneas used for each permeation study [17,18] were evaluated for their hydration level.The wet cornea weight (Ww) was obtained.Each corneal sample was desiccated at 80º to a constant weight (Wd).The percent corneal hydration level (HL %) defi ned as [1-(Wd/Ww)]×100 were determined for cornea recovered after each permeation study [19] .
HPMC and CMC showed mild mucoadhesion at 1% (w/v) concentration.Lower concentration had no mucoadhesion.Eudragit E/RL/RS did not show any mucoadhesion at all.The result indicates that Carbopol ETD 2020 had best mucoadhesion properties on corneal surface in the concentration as low as 0.75 % followed by Carbopol 934NF.Mucoadhesion of Carbopol ETD 2020 and Carbopol 934NF were also studied at different formulation pH.The results indicate that at lower pH (4.5 and 6.0) both polymers have more mucoadhesion then at the pH 7.4.Though the viscosity of the polymeric dispersion was less at lower pH, the probable reason is that at lower pH the carboxylic group of polymer remains protonated and its binding capacity on the corneal surface increases due to hydrogen bonding.
Formulation with Carbopol ETD 2020 showed maximum permeation after 30 min which decreases afterwards, while formulation without Carbopol ETD 2020 showed increasing trend in permeation up to 60 min, but the total quantity of drug permeated was less then obtained with the formulation containing Carbopol ETD 2020.Results are given in Table 1.
The permeation of Carbopol 934 NF formulation was however less then obtained with formulation containing Carbopol ETD 2020.Formulation with and without Carbopol 934NF also showed no corneal damage as the hydration level was below 80%.Results are given in Table 2.
Addition of BKC to the Carbopol ETD 2020 formulation, increased the permeation after 90 and 120 min.Quantity permeated after 30 min was less than that obtained without BKC up to 120 min.On the other hand addition of EDTA to the formulation containing Carbopol ETD 2020 and BKC decreased     the permeation up to 120 min.Quantity permeated after 30 min was less then that obtained without BKC and EDTA.Corneal hydration with BKC and BKC+EDTA was in the physiological range.Results are given in Table 3. Addition of BKC and EDTA to the formulation containing Carbopol 934 NF had no signifi cant infl uence on permeation.Results are given in Table 4.
Thus ETD 2020 and BKC with and without EDTA were also tried in bovine cornea.Formulation with BKC Carbopol ETD 2020 showed maximum permeation after 60 min while formulation with BKC+EDTA and Carbopol ETD 2020 showed maximum permeation after 90 min.Results are given in Table 5.
The results of the present studies reveals that addition of mucoadhesive polymer like Carbopol ETD 2020 can signifi cantly improve mucoadhesion and corneal permeation, providing rationale behind inclusion of compatible, mucoadhesive polymers in controlled drug delivery systems for ophthalmic use.