Ocular formulation for dry eyes and cataract and preparation method thereof

Abstract

The present invention relates to an ocular formulation intended for the treatment of dry eyes and cataracts. The ocular formulation, particularly in the form of eye drop and in-situ gel, comprising gallic acid and quercetin. The invention further describes the method of preparation of ocular formulation.

Description

[0001] OCULAR FORMULATION FOR DRY EYES AND CATARACT AND PREPARATION METHOD THEREOF

[0002] PRIORITY:

[0003] This application claims the benefit of Indian application number 202421049292 dated 27th June 2024 entitled, ‘Ocular formulation for dry eyes and cataract and preparation method thereof, the contents of which are incorporated herein by reference.

[0004] TECHNICAL FIELD OF THE INVENTION

[0005] The present invention relates to an ocular formulation intended for the treatment of dry eyes and cataracts. The ocular formulation, particularly in the form of an eye drop and an in-situ gel formulation, comprising gallic acid and quercetin. The invention further describes the method of preparation of ocular formulation.

[0006] BACKGROUND OF THE INVENTION

[0007] Ocular conditions such as dry eyes and cataracts significantly impact the quality of life for millions of individuals worldwide. These conditions are particularly prevalent among the aging population, although they can affect individuals of all ages due to various etiological factors.

[0008] Dry eye disease, also known as keratoconjunctivitis sicca, is a multifactorial disorder of the tears and ocular surface that results in discomfort, visual disturbance, and tear film instability with potential damage to the ocular surface. It is associated with increased osmolarity of the tear film and inflammation of the ocular surface. The main symptoms include irritation and redness (patients often experience a persistent feeling of dryness, burning, or a gritty sensation in the eyes), fluctuating vision, particularly with prolonged visual tasks, is a common complaint, pain and discomfort.

[0009] The etiology of dry eyes can be broadly classified into two categories:

[0010] • Aqueous Tear-Deficient Dry Eye: Caused by a deficiency in tear production by the lacrimal glands.

[0011] • Evaporative Dry Eye: Resulting from excessive tear evaporation due to conditions affecting the eyelids or the meibomian glands. Common causes and risk factors include aging, hormonal changes, autoimmune diseases (like Sjogren’s syndrome), environmental factors (such as wind, smoke, or dry climates), and prolonged screen time.

[0012] The management of dry eyes involves various strategies aimed at increasing tear production, reducing tear evaporation, and addressing underlying inflammation. Common treatments include over-the-counter lubricating eye drops provide temporary relief but often need frequent reapplication, prescription eye drops containing corticosteroids or cyclosporine to reduce inflammation, lifestyle and environmental changes (increasing humidity, taking breaks during prolonged visual tasks, and avoiding smoke and wind) and despite these options, many patients experience only partial relief and must adhere to a frequent and sometimes inconvenient regimen of treatments.

[0013] Cataracts are characterized by the clouding of the eye's natural lens, leading to a gradual decline in vision. This condition is one of the leading causes of blindness globally. Key characteristics and symptoms of cataracts include blurry vision, sensitivity to light, difficulty with night vision and colors may appear less vibrant.

[0014] Cataracts typically develop as a result of aging, with the majority of cases occurring in individuals over the age of 60. However, other factors contributing to cataract formation include family history of cataracts can increase risk, elevated blood sugar levels can accelerate the development of cataracts, prolonged exposure to UV rays from the sun, smoking and alcohol consumption and ocular injuries or trauma to the eye can precipitate cataract formation. One of the major reasons is due to oxidative stress which oxidise lens cells proteins, damaging DNA and causing peroxidative damage. This leads to protein precipitation in the otherwise transparent lens, causing cataract.

[0015] Currently, the definitive treatment for cataracts is surgical removal of the clouded lens, which is then replaced with an artificial intraocular lens (IOL). Cataract surgery is generally safe and effective, but it is an invasive procedure with potential complications, such as infection, postoperative inflammation can occur, requiring additional treatment, secondary cataract, also known as posterior capsule opacification, can develop and may need laser treatment. There is also paucity of expert surgeons to meet the needs of demanding numbers of cataract surgeries. Given the invasive nature of cataract surgery and its associated risks, there is a strong interest in developing non-surgical treatments that can prevent, delay, or reduce the severity of cataracts.

[0016] There is a pressing need for more effective, non-invasive, and easily administrable treatments for both dry eyes and cataracts. An ideal therapeutic should offer prolonged relief from dry eye symptoms and potentially delay or inhibit the progression of cataracts, reducing the need for surgical intervention. Such a formulation would significantly enhance patient quality of life, particularly for the elderly population who are most affected by these conditions.

[0017] An ocular formulation is a specialized pharmaceutical preparation designed specifically for administration to the eye. These formulations include a variety of dosage forms such as eye drops, ointments, gels, inserts, and contact lenses that contain active drug ingredients intended to treat or manage ocular conditions. The primary aim of ocular formulations is to deliver therapeutic agents effectively and safely to the anterior or posterior segments of the eye, ensuring optimal drug concentration at the site of action while minimizing systemic absorption and side effects.

[0018] Ocular formulations offer distinct advantages over other forms of treatment due to their direct application to the eye, enabling targeted delivery of therapeutic agents precisely where they're needed most. Unlike systemic treatments, which may cause unwanted side effects due to their circulation throughout the body, ocular formulations minimize systemic exposure, reducing the risk of adverse reactions. Additionally, these formulations often provide sustained release, ensuring a prolonged therapeutic effect with fewer administrations, thus enhancing patient compliance and convenience. Furthermore, the development of innovative delivery systems such as nanoparticles and hydrogels has revolutionized ocular therapy, offering improved drug penetration, bioavailability, and tissue targeting, ultimately leading to enhanced efficacy and better clinical outcomes for various ocular conditions.

[0019] Eye drops are a widely accepted and convenient method for delivering medications to the eye. The proposed eye drop formulation ensures that therapeutic agents are delivered directly to the ocular surface, providing immediate relief from dry eye symptoms and delivering antioxidants to the lens. The in-situ gel formulation transforms from a liquid to a gel upon contact with the eye. This transformation enhances the residence time of the therapeutic agents on the ocular surface, ensuring sustained release and prolonged therapeutic effects. The in-situ gel formulation can also improve patient compliance by reducing the frequency of administration required compared to conventional eye drops.

[0020] Indian Patent application 202121033801 discloses herbal ocular formulation of Saraca Asoca useful for the treatment and prophylaxis of cataract formation in diabetic patients as well as people exposed to UV rays without requiring eye surgery. The herbal ocular formulation of Saraca Asoca is also useful for the treatment of dry eyes and stress related eye injuries.

[0021] An article entitled, ‘Eyedrop Formulation and Evaluation of Quercetin - a component of Ginkgo biloba’ by James Brodie; Ben Davis; Lisa Turner; Shereen Nizari; M Francesca Cordeiro in Investigative Ophthalmology & Visual Science June 2015, discloses formulation of quercetin to enable topical drug delivery to the eye, thereby increasing the stability and bioavailability of the drug, and to evaluate its characteristics and suitability for future in vivo studies.

[0022] An article entitled, ‘ Suppression of Intracellular Reactive Oxygen Species in Human Corneal Epithelial Cells via the Combination of Quercetin Nanoparticles and Epigallocatechin Gallate and In Situ Thermosensitive Gel Formulation for Ocular Drug Delivery’ by Chuda Chittasupho, Taepin Junmahasathien, Jiratchaya Chalermmongkol et. al. in Pharmaceuticals 2021 discloses a thermosensitive gel as a delivery system for two antioxidant substances, namely, quercetin and epigallocatechin gallate

[0023] There is a critical need for new, effective, and non-invasive treatments that can provide sustained relief for dry eyes and potentially prevent or treat cataracts. An ideal therapeutic should offer prolonged relief i.e. reducing the frequency of application and improving patient compliance. Addressing both dry eye symptoms and the underlying pathophysiology of cataracts with minimizing side effects and avoiding the risks associated with surgical interventions. OBJECT OF THE INVENTION

[0024] Main object of the present invention is to provide an ocular formulation intended for the treatment of dry eyes and cataracts.

[0025] Another object of the present invention is to provide an ocular formulation, particularly in the form of an eye drop and an in-situ gel formulation, comprising gallic acid and quercetin.

[0026] Yet another object of the present invention is to provide a process for preparation of an ocular formulation, particularly eye drop and in-situ gel.

[0027] SUMMARY OF THE INVENTION

[0028] In an embodiment, the present invention relates to an ocular formulation comprising gallic acid and quercetin.

[0029] In an aspect of the embodiment, the formulation is in the form of eye drop.

[0030] In an aspect of the embodiment, the formulation is in the form of in-situ gel.

[0031] In another embodiment, the present invention relates to an ocular formulation in the form of eye drop comprising, a) 0.06 % w / v of Gallic acid; b) 0.01 % w / v of Quercetin; c) 0.1 % w / v of Hydroxypropyl Methylcellulose (HPMC) as viscosity enhancer; d) 0.1 % w / v of Polysorbate as solubilizer; e) 0.01 % w / v of Benzalkonium Chloride as preservative; f) 0.1 % w / v of Sodium Bisulphite as antioxidant; g) 0.1 % w / v of Mannitol as coolant; h) 1 % w / v of Propylene Glycol; and i) Citrate Buffer pH 5; wherein the percentages are with respect to total weight of the formulation.

[0032] In another aspect of the embodiment, the process for preparation of an ocular formulation in the form of eye drop comprises the steps of: a) Dissolving gallic acid in water to form stock solution I; b) Adding Quercetin to polysorbate, propylene glycol and water to obtain stock solution II; c) Adding hydroxypropyl methylcellulose (HPMC) and mannitol to citrate buffer pH 5 to obtain solution III; d) Mixing stock solution I, II and III to form solution IV; e) Adding benzalkonium chloride and sodium bisulphite to solution IV to form an eye drop formulation.

[0033] In yet another embodiment, the present invention relates to an ocular formulation in the form of in-situ gel comprising, a) 0.06 % w / w of Gallic acid; b) 0.01 % w / w of Quercetin; c) 24 % w / w of Pol oxamers as gel forming agents; d) 0.1 % w / w of Hydroxypropyl Methylcellulose (HPMC) as viscosity enhancer; e) 0.1 % w / w of Polysorbate as solubilizer; f) 0.01 % w / w of Benzalkonium Chloride as preservative; g) 0.1 % w / w of Sodium Bisulphite as antioxidant; h) 0.1 % w / w of Mannitol as coolant; i) 1 % w / w of Propylene Glycol; and j) Citrate Buffer pH 5; wherein the percentages are with respect to total weight of the formulation.

[0034] In another aspect of the embodiment, the pol oxamers are pol oxamer 188 and pol oxamer 407 in a weight ratio of 3 :8.

[0035] In another aspect of the embodiment, the in-situ gel is a temperature dependent in-situ gel formulation.

[0036] In another aspect of the embodiment, the process for preparation of an ocular formulation in the form of in-situ gel comprises the steps of: a) Dissolving gallic acid in water to form stock solution I; b) Adding Quercetin to polysorbate, propylene glycol and water to obtain stock solution II; c) Dissolving poloxamers and hydroxypropyl methylcellulose (HPMC) in citrate buffer pH 5 to obtain gel base; d) Adding stock solution I, II and mannitol, benzalkonium chloride and sodium bisulphite to gel base to form an in-situ gel formulation.

[0037] BRIEF DESCRIPTION OF THE DRAWINGS

[0038] The accompanying drawings are included to provide a further understanding of the present invention and are incorporated in and constitute a part of this specification. The drawings illustrate exemplary embodiments of the present invention and, together with the description, serve to explain the principles of the present disclosure.

[0039] • Figure 1 : Drug release profile of eye drop formulation

[0040] • Figure 2: Representation of pre-gelation and post-gelation Viscosity data

[0041] • Figure 3 : Drug release profile of in-situ gel

[0042] • Figure 4: DPPH assay of Compound A and B

[0043] • Figure 5: DPPH assay of Eye drop and In-situ gel

[0044] • Figure 6: HET-CAM assay test observation

[0045] • Figure 7: Ex-vivo anti-cataract Preventive activity

[0046] • Figure 8: Ex-vivo anti-cataract Therapeutic activity

[0047] DESCRIPTION OF THE INVENTION

[0048] The present invention relates to an ocular formulation intended for the treatment of dry eyes and cataracts. The ocular formulation, particularly in the form of an eye drop and an in-situ gel formulation, comprising gallic acid and quercetin. The invention further describes the method of preparation of ocular formulation.

[0049] The term "comprising", which is synonymous with "including", "containing", or "characterized by" here is defined as being inclusive or open-ended, and does not exclude additional, unrecited elements or method steps, unless the context clearly requires otherwise.

[0050] Unless defined otherwise, all technical as well as scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention pertains. The term "a" and "an" refers to one or to more than one (i.e., to at least one) of the grammatical object of the article. The information provided in this document, and particularly the specific details of the described exemplary aspects, is provided primarily for clearness of understanding and no unnecessary limitations are to be understood from there. As used herein, the term "about" means that the numerical value is approximate and small variations would not significantly affect the practice of the disclosed embodiments. Where a numerical limitation is used, unless indicated otherwise by the context, "about" means the numerical value can vary by ±10% and remain within the scope of the disclosed embodiments.

[0051] As used herein, the terms "treat," or "treatment" mean both therapeutic treatment or prophylactic or preventative measures wherein the object is to prevent or slow down (lessen) an undesired physiological condition, disorder or disease, or obtain beneficial or desired clinical results. For purposes of this invention, beneficial or desired clinical results include, but are not limited to, alleviation of symptoms; diminishment of extent of condition, disorder or disease; stabilized (i.e., not worsening) state of condition, disorder or disease; delay in onset or slowing of condition, disorder or disease progression; amelioration of the condition, disorder or disease state or remission (whether partial or total), whether detectable or undetectable; an amelioration of at least one measurable physical parameter, not necessarily discernible by the patient; or enhancement or improvement of condition, disorder or disease.

[0052] As used herein, the term “ocular formulation” refers to a specialized pharmaceutical preparation designed specifically for administration to the eye. These formulations include a variety of dosage forms such as eye drops, ointments, gels, inserts, and contact lenses that contain active drug ingredients intended to treat or manage ocular conditions. The primary aim of ocular formulations is to deliver therapeutic agents effectively and safely to the anterior or posterior segments of the eye, ensuring optimal drug concentration at the site of action while minimizing systemic absorption and side effects.

[0053] As used herein, the term “eye drop formulation” refers to a liquid formulation specifically designed for administration to the eye. They are a common and convenient method for delivering medications directly to the ocular surface, providing immediate relief from symptoms and targeted treatment for various eye conditions such as dry eyes, infections, allergies, and cataracts.

[0054] As used herein, the term “In-situ gels” refers to innovative ocular formulations that transition from a liquid to a gel upon contact with the eye. This transformation can be triggered by changes in temperature, pH, or ionic concentration. In-situ gels provide sustained and prolonged drug release, making them particularly useful for chronic conditions like dry eyes and cataracts.

[0055] Although the invention has been described with reference to specific embodiments, this description is not meant to be construed in a limiting sense. Various modifications of the disclosed embodiments, as well as alternate embodiments of the invention, will become apparent to persons skilled in the art upon reference to the description of the invention. It is therefore contemplated that such modifications can be made without departing from the spirit or scope of the present invention as defined.

[0056] Main embodiment of the present invention relates to an ocular formulation comprising gallic acid and quercetin.

[0057] Gallic acid is a powerful antioxidant that scavenges free radicals and protects cells from oxidative stress. Gallic acid works by neutralizing reactive oxygen species (ROS) and reducing oxidative damage to the cornea and lens. This helps maintain the integrity of the ocular surface and lens transparency, thereby improving vision and comfort for individuals with dry eyes and cataracts.

[0058] Quercetin is a natural flavonol antioxidant found in various plant and dietary sources. Quercetin exerts its effects in ocular formulations by scavenging free radicals and reducing oxidative stress, which is critical in preventing damage to the cornea and lens. Its antiinflammatory properties help reduce ocular surface inflammation, improving symptoms of dry eyes and enhancing overall ocular health. Furthermore, by preventing protein aggregation, quercetin helps maintain lens clarity, thus playing a role in cataract prevention.

[0059] The combination of gallic acid and quercetin in ocular formulations provides a synergistic effect, enhancing the overall therapeutic potential for treating dry eyes and cataracts. The dual antioxidant and anti-inflammatory properties of these compounds work together to provide comprehensive protection and treatment for ocular tissues.

[0060] In an aspect of the embodiment, the formulation is in the form of eye drop. In an aspect of the embodiment, the formulation is in the form of in-situ gel.

[0061] In another embodiment, the present invention relates to an ocular formulation in the form of eye drop comprising, a) 0.06 % w / v of Gallic acid; b) 0.01 % w / v of Quercetin; c) 0.1 % w / v of Hydroxypropyl Methylcellulose (HPMC) as viscosity enhancer; d) 0.1 % w / v of Polysorbate as solubilizer; e) 0.01 % w / v of Benzalkonium Chloride as preservative; f) 0.1 % w / v of Sodium Bisulphite as antioxidant; g) 0.1 % w / v of Mannitol as coolant; h) 1 % w / v of Propylene Glycol; and i) Citrate Buffer pH 5; wherein the percentages are with respect to total weight of the formulation.

[0062] In another aspect of the embodiment, the process for preparation of an ocular formulation in the form of eye drop comprises the steps of: a) Dissolving gallic acid in water to form stock solution I; b) Adding Quercetin to polysorbate, propylene glycol and water to obtain stock solution II; c) Adding hydroxypropyl methylcellulose (HPMC) and mannitol to citrate buffer pH 5 to obtain solution III; d) Mixing stock solution I, II and III to form solution IV; e) Adding benzalkonium chloride and sodium bisulphite to solution IV to form an eye drop formulation.

[0063] Hydroxypropyl Methylcellulose (HPMC) is a versatile polymer widely used as a viscosity enhancer in ocular formulations. It increases the viscosity of eye drops and in-situ gels, improving the retention time of the therapeutic agents on the ocular surface. This prolonged contact enhances the effectiveness of the medication by allowing sustained release and better absorption, ultimately providing longer-lasting relief from symptoms and improved treatment outcomes.

[0064] Polysorbate commercially available as Tween 80, is a non-ionic surfactant widely used as a solubilizer in pharmaceutical formulations, including ocular solutions. It enhances the solubility of poorly water-soluble drugs, improving their bioavailability and ensuring a homogeneous mixture. In ocular formulations, Polysorbate 80 helps to disperse active ingredients uniformly, enhancing the stability and effectiveness of the medication.

[0065] Benzalkonium chloride (BAK) is a widely used preservative in ocular formulations due to its strong antimicrobial properties. It effectively prevents the growth of bacteria, fungi, and viruses in eye drops, ensuring the sterility and safety of the product throughout its shelf life. BAK is highly effective at low concentrations, making it a popular choice for preserving ophthalmic solutions without compromising patient safety.

[0066] Sodium bisulphite acts as an antioxidant in ocular formulations by protecting the active pharmaceutical ingredients (APIs) from oxidative degradation. It effectively scavenges free radicals and reactive oxygen species, thereby maintaining the stability and potency of the drugs over time. This is particularly important in eye drops and in-situ gels, where oxidative damage could compromise the efficacy and safety of the treatment. By preventing oxidation, sodium bisulphite helps to extend the shelf life of ocular products and ensures consistent therapeutic benefits for patients.

[0067] In yet another embodiment, the present invention relates to an ocular formulation in the form of in-situ gel comprising, a) 0.06 % w / w of Gallic acid; b) 0.01 % w / w of Quercetin; c) 24 % w / w of Pol oxamers as gel forming agents; d) 0.1 % w / w of Hydroxypropyl Methylcellulose (HPMC) as viscosity enhancer; e) 0.1 % w / w of Polysorbate as solubilizer; f) 0.01 % w / w of Benzalkonium Chloride as preservative; g) 0.1 % w / w of Sodium Bisulphite as antioxidant; h) 0.1 % w / w of Mannitol as coolant; i) 1 % w / w of Propylene Glycol; and j) Citrate Buffer pH 5; wherein the percentages are with respect to total weight of the formulation.

[0068] In another aspect of the embodiment, the pol oxamers are pol oxamer 188 and pol oxamer 407 in a weight ratio of 3 :8. Pol oxamers, including Pol oxamer 188 and Pol oxamer 407, are triblock copolymers widely used in pharmaceutical formulations for their surfactant and thermoreversible gelation properties. In ocular applications, they enhance the solubility and stability of drugs while providing a gel-forming capability that transforms from liquid to gel at body temperature. This characteristic is particularly valuable in in-situ gels, where Poloxamers help prolong the residence time of the medication on the ocular surface, improving drug bioavailability and therapeutic efficacy.

[0069] In another aspect of the embodiment, the in-situ gel is a temperature dependent in-situ gel formulation.

[0070] Temperature-dependent in-situ gel formulations remain in a liquid state at room temperature and transform into a gel upon contact with the eye's body temperature. This transformation enhances the retention time and sustained release of the drug, improving therapeutic efficacy and patient compliance in ocular treatments.

[0071] In another aspect of the embodiment, the process for preparation of an ocular formulation in the form of in-situ gel comprises the steps of: a) Dissolving gallic acid in water to form stock solution I; b) Adding Quercetin to polysorbate, propylene glycol and water to obtain stock solution II; c) Dissolving poloxamers and hydroxypropyl methylcellulose (HPMC) in citrate buffer pH 5 to obtain gel base; d) Adding stock solution I, II and mannitol, benzalkonium chloride and sodium bisulphite to gel base to form an in-situ gel formulation.

[0072] The foregoing examples are illustrative embodiments and are merely exemplary. A person skilled in the art may make variations and modifications without deviating from the spirit and scope of the invention. All such modifications and variations are intended to be included within the scope of the claims.

[0073] EXEMPLARY EMBODIMENTS OF THE INVENTION

[0074] Example 1- Eye drop formulation according to the invention

[0075]

[0076] Qs - Quantity sufficient

[0077] Table 1 - Batches of Eye drop formulation

[0078] • Phosphate buffer saline was incompatible with the active pharmaceutical ingredients (APIs), causing formulation discoloration (turning green after one week).

[0079] • Citrate buffer at pH 5 was identified as the optimal choice, effectively preventing oxidation and maintaining API stability. • Sodium bisulphite was selected as the antioxidant after trials showed it prevented discoloration, unlike sodium metabisulfite and sodium thiosulfate.

[0080] • Tween 80 was chosen as a solubilizer, benzalkonium chloride as a preservative, mannitol to maintain isotonicity, and HPMC as a viscosity enhancer.

[0081] Manufacturing process of Eye drop formulation -

[0082] 1) The required quantity of the drug (API) was weighed accurately in an Eppendorf tube.

[0083] 2) Stock of gallic acid solution was prepared in water while quercetin is insoluble in water but soluble in tween 80, PG, and water, hence the stock solution is prepared in the optimized solvent mixture.

[0084] 3) Prepared the required quantity of citrate buffer in filtered water. Added HPMC and vortex it, followed by the addition of mannitol.

[0085] 4) Mix BKC and antioxidants into the mixture.

[0086] 5) Lastly, add gallic acid and quercetin from stock solution in their respective concentrations and vortexed the mixture.

[0087] 6) The formulation was then sterilized by membrane filtration between two burners with a 0.22 pm filter paper.

[0088] 7) The formulation was put into sterile vials and sealed with a sterile cap.

[0089] 8) The sterilized formulation was then stored in a refrigerator (2-8 °C) until further usage.

[0090] Characterization of Eye drop formulation -

[0091] • Determination of pH - The pH of the eye drop was found to be 5 ± 0.3.

[0092] • Clarity Test - The eye drop was clear and free from foreign particles.

[0093] • Viscosity - The viscosity of the eye drop was found to be 3.7 ± 0.5 cp.

[0094] • Refractive index - RI was found to be 1.35.

[0095] • Percent drug content

[0096] A slight difference in drug content was observed between the filtered and unfiltered eye drops containing quercetin, which can be attributed to its low solubility. The filtered eye drops showed a drug content of 80% for quercetin, while the gallic acid content in the filtered eye drops was 96% while without filtered eye drop shows 96% of quercetin and 95% of gallic acid when it was compared with the standard drug solution. • Isotonicity

[0097] There was no any shrinking or swelling observed in case of the eye drop formulation so formulation was isotonic.

[0098] • In-vitro Release eye drop

[0099] It was shown that good results than the drug solution (gallic acid and quercetin in water) In- vitro drug release of eye drop formulation shows 99 % of both the drug release up to the 1 hr as shown in fig 1.

[0100] Table 2 - In-vitro-drug release of eye drop

[0101] • Stability studies

[0102] The stability of a 3 -month eye drop formulation was demonstrated to be stable when stored in a refrigerator. Additionally, it was determined that the formulation remains stable for a period of 1 month under accelerated conditions of 30°C and 60% relative humidity. However, it was observed that the stability of the formulation was unstable when exposed to higher temperatures of 40°C and a relative humidity of 75%.

[0103] Example 2 - In-situ gel formulation according to the invention

[0104] Table 4 - Formulation batches of In-situ gel formulation

[0105] • Various polymers, namely gellan gum (F3), sodium alginate (F2) and HPMC, were tested to form a gel for the ion-activated in-situ gel type. However, no gel formation was observed with this specific gel type.

[0106] • Carbopol 980 with HPMC (Fl) also showed no gel formation.

[0107] • Poloxamer was found to be a more appropriate polymer for the formation of in-situ gel due to its ability to significantly improve the solubility of quercetin.

[0108] Manufacturing process of In-situ gel formulation -

[0109] 1) Poloxamer (P188 and P407) and hydroxypropyl methylcellulose (HPMC) were selected as the gel-forming agents.

[0110] 2) A citrate buffer solution was prepared. The poloxamer (Pl 88 and P407) and HPMC were dissolved in the citrate buffer solution (10 ml in 50 ml falcon) and vortexed.

[0111] 3) The falcon with the gel formulation component were placed in a refrigerator (2 to 8 °C) for 24 hours. Refrigerating the formulation aids in the gelation process, allowing to form gel.

[0112] 4) The poloxamer and HPMC underwent a sol-to-gel phase transition, resulting in the formation of an in-situ gel.

[0113] 5) Added respective excipients and APIs into it make weight up to 20 gm.

[0114] 6) After that formulated in-situ gel was taken for the evaluation. Characterization of In-situ gel formulation -

[0115] • Determination of visual appearance and clarity

[0116] In-situ gel was found to be clear and free from foreign particles.

[0117] . pH - The pH of in-situ gel was found to be 5 ± 0.5

[0118] • Percentage of drug content

[0119] The initial drug content of gallic acid in the in-situ gel formulation was determined to be 95% before the filtration process. Subsequently, after filtration, the drug content slightly decreased to 94%. Similarly, for quercetin, the initial drug content in the gel formulation was found to be 98% prior to filtration, which then decreased to 90% after the filtration step.

[0120] • Viscosity Study

[0121] When we checked the viscosity at different revolution per minute (RPMs) from 5 to 100, it was found that there is increase in the viscosity in case of post gelation at 37 °C as compared to the pre-gelation. (Refer figure 2)

[0122] • In-situ gelling capacity

[0123] Gel formation was observed, and it was determined by placing 1 ml of the sample into an Eppendorf and putting it in the water bath, and maintaining the temperature at 37± 10C.

[0124] • In-vitro drug permeation studies

[0125] In citrate buffer gallic acid show approx. 95 % of drug release and quercetin shows only 9 % of release it was due to higher solubility in poloxamer it would produce complexation with drug it shows the lower release but as shown in fig 3. It was sufficient for the activity as per the antioxidant activity shown by the gallic acid and quercetin in DPPH and ABTS assay only 1 to 2% of quercetin release was sufficient for the antioxidant activity.

[0126] Table 5 - In-vitro drug permeation studies of in-situ gel • In-vitro antioxidant potential analysis - DPPH (2,2-DiphenyI-l-picryl hydrazyl Radical Scavenging Capacity) Assay

[0127] The inhibitory concentration (IC50) of eye drops and in-situ gel was found to be 10.34002 pl and 3.522026 pl, respectively. (Refer figure 4 and 5)

[0128] From the DPPH assay, it was found that Compound A and Compound B showed excellent antioxidants properties in comparison with ascorbic acid. Also, both Eye drop and In-situ Gel containing optimized combination of both Compound A and Compound B showed synergism of antioxidant activity as compared to individual Compound A and Compound B. Based on the amount of Compound B present in eye drop and in-situ gel, a 6-fold and 15-fold increase in DPPH scavenging activity is observed respectively.

[0129] Table 6 - Result of Anti-oxidant activity

[0130] • Stability Study

[0131] In-situ gel shows good one month stability in freezing condition (long term condition) than the accelerated condition.

[0132] Table 7 - In-situ gel stability study

[0133] • In-vitro irritation assay- Hen's eggs chorioallantoic membrane (HET-CAM) assay

[0134] HET-CAM assay was performed to check the irritation potential of the ophthalmic formulation. The assay was carried out using 4 test samples namely- 0.1N NaOH, 0.9% NaCl, eye drop and in-situ gel. After addition of each sample chorioallantoic membrane was observed for a period of 300 seconds and the time in seconds at which haemorrhage, vascular lysis and protein coagulation occurred was noted. (Refer figure 6)

[0135] Table 8 - Calculation of HET-CAM irritation score (IS)

[0136] • Ex-vivo model for cataract using goat lens assay

[0137] Cataract was induced ex-vivo in isolated goat lenses with the following parameter, time of exposure and distance between lens and light source is optimized: a) Product: Osram (ULTRA VITALUX 300 W 230 V E27) b) UV Wavelength: 300 nm (within the range 280-320 nm) c) Medium for lens holding: Artificial Aq. Humour d) Energy of the light: 8 kJ / m2 UV-B radiation e) Incubation period: 2-3 days

[0138] Lenses were divided into 14 sub-groups culminating into 2 major groups i.e.:

[0139] A. Preventive group

[0140] In this group, the lenses were treated before inducing the cataract. (Refer figure 7)

[0141] Table 9 - Ex-vivo anti-cataract Preventive activity B. Therapeutic group

[0142] In this group, the lenses were treated after the induction of cataract. (Refer figure 8)

[0143] Table 10 - Ex-vivo anti-cataract Therapeutic activity

[0144] • Dry eye syndrome

[0145] Ophthalmic formulations were evaluated for their activity on the dry eye disorder on the nine rabbits divided into three groups with three rabbits in each group i.e., control group, treatment group of Eye drop and In-situ gel, where one eye (left eye) of each rabbit were serve as control and the other (right eye) were receive treatment. Dry eye was induced by the topical installation of 0.2% benzalkonium chloride (BKC), twice daily for 20 days. As expected, tear secretion calculated by Schirmer test and Tear breakup time was reduced after installation of 0.2% BKC. Also, an inherent tendency to cure experimentally induced dry eyes with 0.2% BKC was observed during the experiment.

[0146] Table 11 - Result of Schirmer test pre and post treatment

[0147] Table 12 - Result of Tear breakup test pre and post treatment

[0148] It will be understood that various modifications may be made to the aspects disclosed herein. Therefore, the above description should not be construed as limiting, but merely as exemplifications of preferred embodiments. Other arrangements and methods may be implemented by those skilled in the art without departing from the scope of this invention.

Claims

We Claim:

1. An ocular formulation comprising gallic acid and quercetin.

2. The ocular formulation as claimed in claim 1, wherein the formulation is in the form of eye drop.

3. The ocular formulation as claimed in claim 1, wherein the formulation is in the form of in- si tu gel.

4. An ocular formulation in the form of eye drop comprising, a) 0.06 % w / v of Gallic acid; b) 0.01 % w / v of Quercetin; c) 0.1 % w / v of Hydroxypropyl Methylcellulose (HPMC) as viscosity enhancer; d) 0.1 % w / v of Polysorbate as solubilizer; e) 0.01 % w / v of Benzalkonium Chloride as preservative; f) 0.1 % w / v of Sodium Bisulphite as antioxidant; g) 0.1 % w / v of Mannitol as coolant; h) 1 % w / v of Propylene Glycol; and i) Citrate Buffer pH 5;Wherein the percentages are with respect to total weight of the formulation.

5. The ocular formulation as claimed in claim 4, wherein the process for preparation of an ocular formulation in the form of eye drop comprises the steps of: a) Dissolving gallic acid in water to form stock solution I; b) Adding Quercetin to polysorbate, propylene glycol and water to obtain stock solution II; c) Adding hydroxypropyl methylcellulose (HPMC) and mannitol to citrate buffer pH 5 to obtain solution III; d) Mixing stock solution I, II and III to form solution IV; e) Adding benzalkonium chloride and sodium bisulphite to solution IV to form an eye drop formulation.

6. An ocular formulation in the form of in-situ gel comprising, a) 0.06 % w / w of Gallic acid; b) 0.01 % w / w of Quercetin; c) 24 % w / w of Pol oxamers as gel forming agents; d) 0.1 % w / w of Hydroxypropyl Methylcellulose (HPMC) as viscosity enhancer; e) 0.1 % w / w of Polysorbate as solubilizer; f) 0.01 % w / w of Benzalkonium Chloride as preservative;g) 0.1 % w / w of Sodium Bisulphite as antioxidant; h) 0.1 % w / w of Mannitol as coolant; i) 1 % w / w of Propylene Glycol; and j) Citrate Buffer pH 5;Wherein the percentages are with respect to total weight of the formulation.

7. The ocular formulation as claimed in claim 6, wherein the pol oxamers are pol oxamer 188 and pol oxamer 407 in a weight ratio of 3 :8.

8. The ocular formulation as claimed in claim 6, wherein the in-situ gel is a temperature dependent in-situ gel formulation.

9. The ocular formulation as claimed in claim 6, wherein the process for preparation of an ocular formulation in the form of in-situ gel comprises the steps of: a) Dissolving gallic acid in water to form stock solution I; b) Adding Quercetin to polysorbate, propylene glycol and water to obtain stock solution ii; c) Dissolving poloxamers and hydroxypropyl methylcellulose (HPMC) in citrate buffer pH 5 to obtain gel base; d) Adding stock solution I, II and mannitol, benzalkonium chloride and sodium bisulphite to gel base to form an in-situ gel formulation.

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