Single-dose non-steroidal anti-inflammatory drug eye drops

Abstract

The invention provides a single-dose non-steroidal anti-inflammatory drug eye drop. According to the present invention, the compound represented by the formula I is obtained through splitting, and the research results show that the free acid of the compound represented by the formula I has characteristics of low water solubility, poor physical and chemical stability, easy moisture absorption, stickiness, poor fluidity and the like; therefore, various salt forms of the compound shown in the formula I are further researched, tromethamine salt of the compound shown in the formula I is finally selected as an effective component from multiple aspects of physicochemical properties and safety, and the eye drops are prepared through a preparation means to be directly used for eye drop administration, so that the problem that when a prodrug is directly used, the dosage of the tromethamine salt is reduced is effectively solved. And the loxoprofen active metabolite can be used for enriching the application of the loxoprofen active metabolite in antipyretic, analgesic and anti-inflammatory medicines.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical preparations, specifically relating to a single-dose nonsteroidal anti-inflammatory eye drop. Background Technology

[0002] Nonsteroidal anti-inflammatory drugs (NSAIDs) have anti-inflammatory, anti-allergic, and analgesic effects. Topical application can prevent and treat inflammatory eye diseases caused by various factors, with fewer adverse reactions, and is safer than glucocorticoids in clinical use. Currently marketed NSAID eye drops include 0.1% pranoprofen eye drops, 0.1% diclofenac sodium eye drops, 0.5% and 0.4% ketorolac tromethorphan eye drops, 0.03% flurbiprofen sodium eye drops, 0.1% napafenamide, bromfenac sodium, and indomethacin eye drops, among others.

[0003] Pramoprolol eye drops, developed by Senju Pharmaceutical Co., Ltd., are marketed in 5ml:5mg multi-dose packaging, administered four times daily. Literature indicates an adverse reaction rate of 1.35%, with major adverse reactions including irritation, conjunctival hyperemia, itching, and eyelid redness or swelling. Bromfenac sodium eye drops are currently approved for marketing in China, available in single-dose packaging (0.4ml:0.4mg) and multi-dose packaging (5ml:5mg), administered twice daily. Literature indicates an adverse reaction rate of 1.87%, with major adverse reactions including corneal erosion, conjunctivitis, blepharitis, irritation, transient eye pain, superficial punctate keratitis, itching, corneal epithelial detachment, and eyelid heat. Diclofenac sodium eye drops are available in various dosage strengths, generally administered 4–6 times daily. At the American Cataract and Refractive Surgery Conference, 11 cases of corneal lysis following topical application of 0.5% diclofenac sodium eye drops were reported. Studies have also shown the effects of diclofenac sodium eye drops and its raw materials on the proliferation, migration, and damage of cultured human corneal epithelial cells. Furthermore, after using these eye drops, patients are prone to temporary burning, stinging, and tearing; a very small number may experience conjunctival congestion and blurred vision; and less than 3% of patients may experience systemic reactions such as fatigue, drowsiness, and nausea.

[0004] In summary, these commonly used NSAID eye drops have a variety of adverse reactions, and these corneal toxicities and ocular side effects may be closely related to the preservatives in the topical eye drops. Furthermore, NSAID eye drops should be used with caution in conditions such as diabetes, autoimmune diseases, ocular surface diseases, or early corneal lesions following eye surgery. Therefore, there is a great clinical need for new, safe, effective, highly penetrating, and stable topical nonsteroidal anti-inflammatory drugs (NSAIDs).

[0005] Loxoprofen sodium is the first propionate prodrug-type nonsteroidal anti-inflammatory drug (NSAID), originally developed by Daiichi Sankyo Co., Ltd. of Japan, and first marketed in Japan in 1986. This drug is a prodrug with no intrinsic activity. After oral administration, loxoprofen is absorbed as a free acid and biotransformed in the body into a hydroxyl metabolite with three chiral centers, theoretically generating eight stereoisomers. The pharmacologically active form is the trans-hydroxyl form (such as compound I).

[0006]

[0007] Its main characteristics are that peak plasma concentration is reached within 30 minutes after oral administration, with few toxic side effects and good clinical efficacy. Meanwhile, its indications are also very broad, showing good analgesic and anti-inflammatory effects for chronic rheumatoid arthritis, osteoarthritis, low back pain, frozen shoulder, cervicitis, and postoperative and traumatic conditions. Currently, the marketed drug is available in tablet, capsule, granule, patch, and gel forms. Guangzhou Aobo Pharmaceutical Technology Co., Ltd. has applied for a Class 2.2 new drug, loxoprofen sodium eye drops, and filed a new drug clinical trial application in February 2020 (acceptance number CXHL2000030), which has been granted implied clinical approval and is currently undergoing Phase II clinical trials. The patent application CN106880590A discloses a prodrug-type nonsteroidal anti-inflammatory drug eye drop and its preparation method, namely loxoprofen sodium eye drops. However, this eye drop contains an antibacterial agent, the presence of which can irritate the surface cells of the eye, directly affecting the composition of tears and altering the microenvironment of the ocular surface. This poses a significant potential risk, especially for eye drops that require frequent use. Furthermore, loxoprofen sodium needs to enter the systemic circulation through capillaries, be metabolized to produce active metabolites, and then return to the target site via the bloodstream to exert its effect. This results in low systemic exposure, slow elimination, and low bioavailability. Summary of the Invention

[0008] Currently, one of the problems facing eye drops is the low bioavailability (<5%) of the active substances. At the same time, when the prodrug loxoprofen is administered, only a portion of the prodrug is converted into active metabolites in the eye to exert its effect, further reducing bioavailability.

[0009] Based on this, the present invention first obtained the compound of formula I through resolution. Research revealed that the free acid of formula I itself has low water solubility, poor physicochemical stability, easy hygroscopicity and stickiness, and poor flowability. Therefore, various salt forms of formula I were further studied. Considering physicochemical properties and safety, the tromethamine salt of formula I was ultimately selected as the active ingredient. It was then formulated into eye drops for direct eye administration, effectively solving the problem of low bioavailability when prodrugs are directly administered, where only a portion is converted into active metabolites. This enriches the applications of loxoprofen's active metabolites in antipyretic, analgesic, and anti-inflammatory drugs.

[0010] The solution to achieve the above-mentioned objectives is:

[0011] This invention provides a single-dose nonsteroidal anti-inflammatory eye drop, comprising:

[0012] (a) A hydroxyl salt of compound I

[0013]

[0014] (b) Excipients for ophthalmic preparations;

[0015] The content of the compound tromethamine salt shown in Formula I is 0.05–1.0 wt%.

[0016] The excipients for the ophthalmic preparation include metal ion chelating agents, thickeners, osmotic pressure regulators, and pH regulators; the eye drops do not contain antibacterial agents.

[0017] According to an embodiment of the present invention, the metal ion complexing agent is selected from at least one of disodium edetate, calcium sodium edetate, dihydroxyethylglycine, ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid, penta(carboxymethyl)diethylenetriamine, aminotriacetic acid, sodium gluconate, sodium citrate, and tartaric acid; the content of the metal ion complexing agent is 0.01-0.1 wt%.

[0018] According to an embodiment of the present invention, the thickener is selected from at least one of sodium hyaluronate, hydroxypropyl methylcellulose, polysorbate 20, carbomer, xanthan gum, polyethylene glycol, polyvinyl alcohol, polyvinylpyrrolidone, sodium carboxymethyl cellulose, methylcellulose, carboxymethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, sodium polyacrylate, microcrystalline cellulose, povidone, glycerin, sodium alginate, poloxamer, dextran, and polycarboxyphene; the content of the thickener is 0.01–0.1 wt%.

[0019] According to an embodiment of the present invention, the osmotic pressure regulator is selected from at least one of sodium chloride, potassium chloride, boric acid, borax, glycerol, propylene glycol, mannitol, sodium sulfate, potassium sulfate, sodium nitrate, potassium nitrate, glucose, sorbitol, and sodium cromoglycate; the content of the osmotic pressure regulator is 0.5-2.0 wt%.

[0020] According to an embodiment of the present invention, the pH adjuster is selected from at least one of phosphoric acid, hydrochloric acid, sodium hydroxide, sodium citrate, citric acid, boric acid, borax, sodium acetate, and acetic acid; the pH adjuster is used to adjust the pH of the eye drops to 6.0-8.0; preferably 6.5-7.5.

[0021] According to an embodiment of the present invention, the osmotic pressure of the eye drops is 260–320 mOsmol / kg.

[0022] Those skilled in the art will understand that the osmotic pressure of tears is equal to that of serum, equivalent to the osmotic pressure of 0.9% sodium chloride (286 mOsm). The eye can tolerate an osmotic pressure range equivalent to 0.6–1.5% sodium chloride (approximately 200–450 mOsm), with the optimal range being 260–310 mOsm. The osmotic pressure of eye drops has a certain influence on corneal permeability. When the osmotic pressure of eye drops differs significantly from that of physiological saline, the irritant effect prompts tear secretion to dilute the medication, reducing corneal permeability. Therefore, eye drops should be prepared as a solution with an osmotic pressure equal to or close to that of tears. Hypertonic eye drops can cause the external ocular tissues to lose moisture, leading to dryness and discomfort. Hypotonic eye drops can cause the cells of the external ocular tissues to swell, resulting in irritation. Therefore, eye drops should be prepared as isotonic solutions. In this invention, it is preferable to control the osmotic pressure between 260 and 320 mOsm.

[0023] According to an embodiment of the present invention, the compound tromethamine salt of Formula I is in crystalline form.

[0024] This invention yielded crystalline form A of the tromethamine salt of Formula I. X-ray powder diffraction patterns obtained using Cu-Ka radiation showed characteristic diffraction peaks at 2θ values ​​of 4.7±0.2, 9.4±0.2, 9.8±0.2, 12.6±0.2, 16.6±0.2, 17.3±0.2, 18.6±0.2, 19.6±0.2, 20.4±0.2, 21.1±0.2, 23.4±0.2, and 23.7±0.2. The DSC pattern showed an endothermic peak at approximately 108.5 °C.

[0025] This invention yielded crystalline form B of the tromethamine salt of compound I. The X-ray powder diffraction pattern obtained using Cu-Ka radiation showed characteristic diffraction peaks at 2θ values ​​of 6.25±0.2, 8.77±0.2, 17.24±0.2, 17.97±0.2, 19.57±0.2, and 20.56±0.2. The DSC pattern showed an endothermic peak at approximately 112.6 °C.

[0026] According to an embodiment of the present invention, the preferred active ingredient in the eye drop formulation is compound I, tromethamine salt, crystal form B.

[0027] According to an embodiment of the present invention, the eye drops are individually packaged, and the volume of the container for each individually packaged item ranges from 0.4 to 0.8 mL.

[0028] According to a preferred embodiment of the present invention, it comprises:

[0029] Formula I compound aminobutadiene triol salt: 0.05–0.5 wt%;

[0030] Disodium edetate, a metal ion chelating agent: 0.01–0.1 wt%;

[0031] Thickener sodium hyaluronate: 0.01–0.1 wt%;

[0032] Sodium chloride, an osmotic pressure regulator: 0.5–2 wt%;

[0033] pH adjuster: Apply an appropriate amount to adjust the pH to 6.5–7.5;

[0034] The rest is water.

[0035] This invention also provides a method for preparing the above-mentioned single-dose nonsteroidal anti-inflammatory drug eye drops, comprising the following steps:

[0036] Step 1) Weigh out the prescribed amount of sodium hyaluronate, dissolve it thoroughly in purified water until it swells, and set aside.

[0037] Step 2) Weigh out the prescribed amounts of compound I, tromethamine salt, disodium edetate, sodium chloride, and pH adjuster and add them to the solution in Step 1). Stir until completely dissolved, adjust the pH value to 6.5-7.5, and adjust the osmolar concentration to 260-320 mOsmol / kg.

[0038] Step 3) Add purified water to 500ml, filter, dispense and seal, and autoclave at 121℃ for 15min;

[0039] Step 4) Fill and seal a single dose using a blow-fill-seal machine, with a volume of 0.4 to 0.8 ml. Cool to 4°C and maintain for 1 to 3 hours to obtain the product.

[0040] The present invention also provides the use of the single-dose nonsteroidal anti-inflammatory eye drops in the preparation of drugs for the prevention or treatment of ophthalmic diseases.

[0041] Furthermore, the ophthalmic diseases include at least one of blepharitis, conjunctivitis, keratitis, scleritis, superficial scleritis, iridocyclitis, and postoperative inflammation.

[0042] The eye drops of this invention have stable quality and good efficacy, making them an advantageous eye drops for treating eye inflammation.

[0043] The beneficial effects of this invention are as follows: Compared with the prior art, this invention uses the compound of Formula I, tromethamine salt, as the active ingredient directly as eye drops, effectively overcoming the problem of low bioavailability when the original drug is directly administered. Furthermore, after being placed under high temperature and strong light irradiation conditions, the pH value and content of the loxoprofen active metabolite tromethamine salt eye drops remain essentially unchanged, and the results of related substance tests all meet the limit requirements, indicating that the properties of this loxoprofen active metabolite tromethamine salt eye drops are stable. In a rat conjunctival edema inhibition test, this loxoprofen active metabolite tromethamine salt eye drops effectively inhibited the development of conjunctival edema, and its efficacy was basically consistent with that of the positive control drug, pranoprofen eye drops, showing good therapeutic effects. In addition, the single-dose eye drops provided by this invention do not contain antibacterial agents and are produced using autoclaving and aseptic filling processes, ensuring product sterility while avoiding the risk of microbial contamination during the use of multi-dose eye drops. In summary, the single-dose eye drops described in this invention are stable, safe, and have reliable efficacy. Attached Figure Description

[0044] Figure 1 This is the PXRD pattern of sodium salt crystal form A of compound I.

[0045] Figure 2 This is the DSC spectrum of sodium salt crystal form A of compound I.

[0046] Figure 3 This is the PXRD pattern of crystal form A of compound A, aminobutadiene triol salt, of formula I.

[0047] Figure 4 This is the DSC spectrum of crystal form A of compound A, aminobutadiene triol salt.

[0048] Figure 5 This is the PXRD pattern of crystal form B of compound I, aminobutadiene triol salt.

[0049] Figure 6 This is the DSC spectrum of crystal form B of compound I, aminobutadiene triol salt.

[0050] Figure 7H&E stained sections of rat conjunctival tissue (A is conjunctival tissue of rats in the blank control group, B is the model group, C is the pranoprofen eye drop group, D is the low-dose group of the test compound eye drops, and E is the high-dose group of the test compound eye drops). Detailed Implementation

[0051] To make the above-described objects, features, and advantages of this disclosure more apparent and understandable, the specific embodiments of this disclosure will be described in detail below with reference to the accompanying drawings and specific examples. Many specific details are set forth in the following description to provide a thorough understanding of this disclosure; however, this disclosure may be implemented in other ways different from those described herein, and those skilled in the art can make similar extensions without departing from the spirit of this disclosure. Therefore, this disclosure is not limited to the specific embodiments disclosed below.

[0052] Example 1

[0053] Preparation of Compound I

[0054] The preparation method of the compound of formula I of this invention is based on patent: ZL201610435178.8 A method for preparing loxoprofen active metabolites.

[0055] Example 2

[0056] 1. Preparation of sodium salt crystal form A of compound I

[0057] Weigh 2.97 g (12 mmol) of compound I and 0.82 g (12 mmol) of sodium ethoxide, add 20 ml of ethanol, control the temperature at 35 °C, stir to dissolve, after the reaction is complete, concentrate under reduced pressure to 50% of the original volume under the condition of keeping warm, slowly add methyl tert-butyl ether until the cloud point appears, lower the temperature to 0-5 °C, filter, collect the solid, dry under reduced pressure at 45 °C to obtain sodium salt crystal form A of compound I.

[0058] This invention yielded sodium salt crystal form A of compound I, whose X-ray powder diffraction pattern obtained using Cu-Ka radiation exhibits characteristic diffraction peaks at 2θ values ​​of 8.1±0.2, 9.4±0.2, 12.9±0.2, 16.3±0.2, 16.8±0.2, 17.5±0.2, 18.8±0.2, 19.7±0.2, 21.7±0.2, 24.1±0.2, 24.8±0.2, and 27.9±0.2, and has essentially the characteristics shown in the attached figure. Figure 1 The X-ray powder diffraction pattern shown represents the characteristics; the DSC pattern shows endothermic peaks at approximately 69.6℃ and 181.6℃, as shown in the attached diagram. Figure 2 As shown.

[0059] 2. Preparation of amorphous α-form of sodium salt of compound I

[0060] Weigh 3.72 g (15 mmol) of compound I and 1 g (15 mmol) of sodium ethoxide, add 40 ml of isopropanol, control the temperature at 30 °C, stir to dissolve, after the reaction is complete, remove the solvent under reduced pressure, add 60 ml of n-heptane, stir at room temperature for 6-8 h, filter, collect the obtained product, dry under reduced pressure at 45 °C to obtain the amorphous α-sodium salt of compound I.

[0061] Example 3

[0062] 1. Preparation of crystal form A of compound I, aminobutadiene triol salt

[0063] Weigh 2g (8mmol) of compound I and 1g (8mmol) of tromethamine, add 10ml of anhydrous ethanol, control the temperature at 45℃, stir to dissolve, after the reaction is complete, concentrate under reduced pressure to remove the solvent, add 25-30ml of ethyl acetate, stir under the heat, and after complete solidification, cool to 0-5℃, filter, collect the solid, and dry under reduced pressure at 45℃ to obtain tromethamine salt crystal form A of compound I.

[0064] This invention yielded tromethamine salt crystal form A, and the X-ray powder diffraction pattern obtained using Cu-Ka radiation showed characteristic diffraction peaks at 2θ values ​​of 4.7±0.2, 9.4±0.2, 9.8±0.2, 12.6±0.2, 16.6±0.2, 17.3±0.2, 18.6±0.2, 19.6±0.2, 20.4±0.2, 21.1±0.2, 23.4±0.2, and 23.7±0.2; it possesses essentially the characteristics shown in the attached diagram. Figure 3 The X-ray powder diffraction pattern shown represents the characteristics; the DSC pattern shows an endothermic peak at approximately 108.5 °C, as shown in the attached image. Figure 4 As shown.

[0065] 2. Preparation of amorphous α-formula of aminobutadiene triol salt of Formula I

[0066] Weigh 2.7 g (10.8 mmol) of compound I and 1.3 g (10.8 mmol) of tromethamine, add 30 ml of anhydrous ethanol, control the temperature at 45–55 °C, stir to dissolve, after the reaction is complete, add 30 ml of ethyl acetate under the condition of keeping warm, quickly place in a cooling bath at -20–-10 °C, stir for 0.5–1 h, filter, add the solid to n-hexane and stir for 0.5 h, filter, collect the product, dry under reduced pressure at 45 °C to obtain the amorphous α-sodium salt of compound I.

[0067] 3. Preparation of crystal form B of aminobutadiene triol salt of Formula I

[0068] Weigh 4.9 g (19.7 mmol) of compound I and 2.4 g (19.7 mmol) of tromethamine, add 19.6 ml of anhydrous ethanol, stir and dissolve at 45–50 °C, react for 0.5 h, add 9.8 ml of ethyl acetate and a small amount of activated carbon, keep warm and stir at 45–60 °C for 0.5 h, filter, add 29.4 ml of ethyl acetate to the filtrate, gradually lower the temperature to 0–5 °C, keep warm and stir for 3.5–4 h, filter, slurry the filter cake with n-hexane at 0–10 °C for 1.5–2 h, filter by suction, and vacuum dry the filter cake at 45–50 °C for 10–12 h to obtain tromethamine salt crystal form B of compound I.

[0069] This invention yielded tromethamine salt crystal form B, and the X-ray powder diffraction pattern obtained using Cu-Ka radiation showed characteristic diffraction peaks at 2θ values ​​of 6.25±0.2, 8.77±0.2, 17.24±0.2, 17.97±0.2, 19.57±0.2, and 20.56±0.2. It possesses essentially the characteristics shown in the attached diagram. Figure 5 The X-ray powder diffraction pattern shown represents the characteristics; the DSC pattern shows an endothermic peak at approximately 112.6 °C, as shown in the attached image. Figure 6 As shown.

[0070] Table 1. Properties of sodium salts and tromethamine salts of Formula I

[0071]

[0072] Test Example 1: Solubility in water:

[0073] Weigh 20 mg each of the free acid of compound I, the sodium salt of compound I, and the tromethamine salt of compound I. Using the solubility test method in the general rules of the 2015 edition of the Chinese Pharmacopoeia, add each sample to an appropriate amount of water at 25°C. Shake vigorously for 30 seconds every 5 minutes and observe the dissolution over 30 minutes. Complete dissolution is considered complete when no visible solute particles remain. The results are shown in Table 2 below.

[0074] Table 2. Solubility of compounds in water

[0075] name Solubility (as free acid) Pharmacopoeia solubility Formula I compound free acid Approximately 1.00 mg / mL Slightly soluble Sodium salt of Formula I Greater than 92.78 mg / mL Easily soluble Formula I compound aminobutane triol salt Greater than 67.88 mg / mL Easily soluble

[0076] Experimental Example 2: Accelerated Stability Study

[0077] The free acid of compound I, the sodium salt of compound I, and the tromethamine salt of compound I were subjected to an accelerated stability test for 6 months in a constant temperature and humidity chamber. The conditions were 40℃ / 75% relative humidity (RH). Samples were taken at 1, 2, 3, and 6 months, and the purity and impurities were tested by high performance liquid chromatography (HPLC), and the accelerated stability was characterized by PXRD. The results are shown in Table 3 below.

[0078] Table 3 Accelerated stability of compounds

[0079]

[0080]

[0081] Experimental Example 3: Hygroscopicity Study

[0082] Approximately 20 mg each of the free acid of compound I, the sodium salt of compound I, and the hygroscopicity of compound I was tested using a dynamic moisture adsorption (DVS) instrument. The results are shown in Table 4 below. The weight gain of the solids at 80% relative humidity was tested under a nitrogen atmosphere, with a relative humidity range of 0–80% RH and a temperature of 25°C. The hygroscopic characteristics and the definition of hygroscopic weight gain are as follows: deliquescence refers to the absorption of sufficient moisture to form a liquid; extremely hygroscopic means a weight gain of not less than 15%; hygroscopic means a weight gain of less than 15% but not less than 2%; slightly hygroscopic means a weight gain of less than 2% but not less than 0.2%; no or almost no hygroscopicity means a weight gain of less than 0.2%.

[0083] Table 4 Hygroscopicity of Compounds

[0084]

[0085] The above results indicate that, compared with the free acid, sodium salt, tromethamine salt, and polymorphs of compound I, the preferred active ingredient in the eye drop formulation of the present invention is tromethamine salt crystal form B of compound I, which exhibits superior performance in terms of stability and hygroscopicity.

[0086] Experimental Example 4: Acid-Base Destruction Test

[0087] Strong acid and strong base degradation tests were conducted on the crystal form B of compound I, tromethamine salt. The test results are shown in Table 5.

[0088] Table 5 shows the crystal form of compound I, aminobutyritol salt, and its acid-base degradation.

[0089]

[0090] Note: "ND" indicates not detected; "N / A" indicates not applicable.

[0091] In the forced degradation test of the active pharmaceutical ingredient, under oxidative conditions, this product produced approximately 0.06% loxoprofen acid, while other impurities showed no significant changes. Under high-temperature conditions, two relatively large unknown impurities, RRT0.63 and RRT0.66, were produced, amounting to approximately 0.16% and 0.27% respectively, with a slight increase in total impurities (approximately 0.4%), indicating relative sensitivity to high-temperature conditions. Under acidic and alkaline conditions, the number and amount of impurities did not increase significantly compared to the undegraded product, indicating that this product is relatively stable under acidic, alkaline, and light conditions.

[0092] Example 4

[0093] prescription Dosage Formula I compound aminobutane triol salt 0.223g (0.03wt% based on compound of formula I) Sodium hyaluronate 0.5g Sodium edetate 0.05g Sodium chloride 2.4g Sodium dihydrogen phosphate 2.3g Sodium hydrogen phosphate 2.4g Sodium hydroxide Appropriate amount Add water for injection to bring the volume to a final level. 500ml

[0094] Preparation process:

[0095] Weigh 0.5g of sodium hyaluronate and dissolve it fully in 400ml of purified water. Set aside. Weigh the prescribed amounts of compound I (aminobutyrate), disodium edetate, sodium chloride, and pH adjuster and add them to the solution from step 1). Stir until completely dissolved, adjust the pH to 7.0, and adjust the osmolar concentration to 260 mOsmol / kg. Add water for injection to a final volume of 500ml, filter, dispense, and seal. Autoclave at 121℃ for 15min. Fill and seal a single dose using a blow-fill-seal machine, filling 0.4-0.8ml at a temperature of 4℃ for 1-3 hours.

[0096] Example 5

[0097]

[0098]

[0099] Preparation process:

[0100] Weigh 0.5g of sodium hyaluronate and dissolve it fully in 400ml of purified water. Set aside. Weigh the prescribed amounts of compound I (aminobutyrate), disodium edetate, sodium chloride, and pH adjuster and add them to the solution from step 1). Stir until completely dissolved, adjust the pH to 7.5, and adjust the osmolar concentration to 280 mOsmol / kg. Add water for injection to a final volume of 500ml, filter, dispense, seal, and autoclave at 121℃ for 15min. Fill and seal a single dose using a blow-fill-seal machine, filling 0.4-0.8ml at a temperature of 4℃ for 1-3 hours to obtain the final product.

[0101] Example 6

[0102] prescription Dosage Formula I compound aminobutane triol salt 2.23 g (3.0 wt% based on compound of formula I) Sodium hyaluronate 0.5g Sodium edetate 0.05g Sodium chloride 2.4g Sodium dihydrogen phosphate 2.3g Sodium hydrogen phosphate 2.4g Sodium hydroxide Appropriate amount Add water for injection to bring the volume to a final level. 500ml

[0103] Preparation process:

[0104] Weigh 0.5g of sodium hyaluronate and dissolve it fully in 400ml of purified water. Set aside. Weigh the prescribed amounts of compound I (aminobutyrate), disodium edetate, sodium chloride, and pH adjuster and add them to the solution from step 1). Stir until completely dissolved, adjust the pH to 6.5, and adjust the osmolar concentration to 290 mOsmol / kg. Add water for injection to a final volume of 500ml, filter, dispense and seal, and autoclave at 121℃ for 15min. Fill and seal a single dose using a blow-fill-seal machine, filling 0.4-0.8ml each time. Cool to 4℃ and maintain for 1-3 hours to obtain the final product.

[0105] Comparative Example 1

[0106] prescription Dosage Sodium salt of Formula I 0.544 g (1.0 wt% based on compound of formula I) Sodium hyaluronate 0.5g Sodium edetate 0.05g Sodium chloride 2.4g Sodium dihydrogen phosphate 2.3g Sodium hydrogen phosphate 2.4g Sodium hydroxide Appropriate amount Add water for injection to bring the volume to a final level. 500ml

[0107] Preparation process:

[0108] Weigh 0.5g of sodium hyaluronate and dissolve it fully in 400ml of purified water. Set aside. Weigh the prescribed amounts of sodium salt of compound I (sodium salt crystal form A), disodium edetate, sodium chloride, and pH adjuster and add them to the solution in step 1). Stir until completely dissolved, adjust the pH to 7.5, and adjust the osmolar concentration to 280mOsmol / kg. Add water for injection to a final volume of 500ml, filter, dispense and seal, and autoclave at 121℃ for 15min. Fill and seal a single dose using a blow-fill-seal machine, with a filling volume of 0.4-0.8ml. Cool to 4℃ and maintain for 1-3h to obtain the final product.

[0109] Comparative Example 2

[0110] prescription Dosage Loxoprofen sodium 0.5g Sodium hyaluronate 0.5g Sodium edetate 0.05g Sodium chloride 2.4g Sodium dihydrogen phosphate 2.3g Sodium hydrogen phosphate 2.4g Sodium hydroxide Appropriate amount Add water for injection to bring the volume to a final level. 500ml

[0111] Preparation process:

[0112] Weigh 0.5g of sodium hyaluronate and dissolve it completely in 400ml of purified water. Set aside. Weigh the prescribed amounts of loxoprofen sodium, disodium edetate, sodium chloride, and pH adjuster and add them to the solution from step 1). Stir until completely dissolved, adjust the pH to 7.5, and adjust the osmolar concentration to 280 mOsmol / kg. Add water for injection to a final volume of 500ml, filter, dispense, seal, and autoclave at 121℃ for 15min. Fill and seal a single dose using a blow-fill-seal machine, filling 0.4-0.8ml at a temperature of 4℃ for 1-3 hours to obtain the final product.

[0113] Experimental Example 5

[0114] 1. Sample stability:

[0115] The influencing factors of the single-dose eye drops prepared in Examples 1-3 and Comparative Example 1 were studied. Temperature (40℃, 60℃) was measured for 10 and 30 days, and strong light (total illuminance not less than 1.2 × 10⁶ Lux·hr, near-ultraviolet energy not less than 200 W·hr / m²) was also measured. 2 The drug content, related substances, and pH value in the eye drops were measured under 6-day and 12-day conditions.

[0116] The stability test results of the single-dose eye drops prepared in Examples 1-3 and Comparative Example 1 are shown in Table 6.

[0117] Table 6 shows the stability test results of eye drops from Examples 1-3 and Comparative Example 1.

[0118]

[0119]

[0120] As shown in Table 2, the content and pH value of the single-dose Formula I compound tromethamine salt eye drops prepared in Examples 4-6 did not change significantly after being exposed to high temperatures of 40°C and 60°C, or after 12 days of strong light irradiation. The total impurities in the samples also met the limit requirements, indicating that the product is stable.

[0121] The sodium salt eye drops of Formula I prepared in Comparative Example 1 showed a decreasing trend in drug content under high temperature and strong light irradiation conditions, and a significant increase in related substances. Compared with the tromethamine salt eye drops of Formula I, this sample had poor stability.

[0122] 2. Pharmacokinetic Comparison Study

[0123] Thirty-six healthy New Zealand white rabbits were randomly divided into three groups: the treatment group of Example 2 and the treatment groups of Comparative Examples 1 and 2, with 12 rabbits in each group. The test eye drops were administered to each rabbit in each group at a dose of 50 μL, simultaneously in both eyes. At 20 min, 40 min, 1 h, 2 h, 4 h, and 6 h after administration, 50 μL of aqueous humor was collected from two rabbits in each group, and the content of the active metabolite (compound of Formula I) was detected by LC-MS.

[0124] Table 7. Content of active metabolites in rabbit aqueous humor of different eye drop treatment groups

[0125]

[0126] Table 8. Main pharmacokinetic parameters of different eye drops administered to the eyes of New Zealand rabbits.

[0127] Group Tmax(h) AUC(μg*h*ml-1) MRT(h) Example 2 2 2854.51 2.87 Comparative Example 1 1 2300.95 2.82 Comparative Example 2 1 1033.47 2.43

[0128] Compared to Comparative Example 2 eye drops, both Example 2 and Comparative Example 1 eye drops used different salt forms of compound 1, and their concentrations in rabbit eyes were significantly higher than those of Comparative Example 2 loxoprofen sodium eye drops at the same concentration. Loxoprofen sodium is a prodrug that can be bioconverted in vivo into a hydroxyl metabolite with three chiral centers, theoretically generating eight stereoisomers, of which only compound 1 has pharmacological activity. Therefore, the conversion efficiency of loxoprofen sodium to the active metabolite in vivo is low, and the bioavailability of Comparative Example 2 eye drops is lower than that of Example 2 and Comparative Example 1 eye drops. Furthermore, the concentration and residence time of Example 2 eye drops in rabbit eyes were slightly higher than those of Comparative Example 1 eye drops.

[0129] In summary, the embodiments of the present invention directly prepare the tromethamine salt of loxoprofen sodium active metabolite (compound of formula 1) into eye drops through formulation methods, without the need for in vivo conversion, effectively overcoming the problem of low bioavailability and showing greater advantages.

[0130] 3. Pharmacodynamic testing

[0131] To evaluate the anti-inflammatory effect of the eye drops containing the compound of the present invention on conjunctivitis, the inhibitory effect of the eye drops on edema formation in a rat model of arachidonic acid-induced conjunctivitis was studied. Arachidonic acid was instilled into the eyes of rats to create a membrane. The swelling rate of the conjunctiva in the model group (Group B) was calculated based on the weight of the conjunctiva in the control group and the model group. The test compound eye drops were the tromethamine salt of compound I prepared in Examples 2 and 3, designated as the low-dose group (Group D) and the high-dose group (Group E), respectively. The positive control group used commercially available pranoprofen eye drops (Senju Pharmaceutical Co., Ltd., Group C). It should be noted that the edema formation inhibition rate was calculated based on the edema weight of the control group (Group A) and the edema weight of the test compound treatment group.

[0132] Thirteen healthy male SD rats were randomly divided into five groups. Except for the blank control group (group A) and the model group (group B), which each had two rats, the low-dose group (group D) and the high-dose group (group E) of compound I tromethamine salt eye drops, and the pranoprofen eye drops group (group C) each had three rats.

[0133] Physiological saline containing 5% arachidonic acid was dripped into the lower eyelid conjunctival sac of each rat in the model group and the experimental group (10 μL / eye) to establish the model. The blank control group was dripped with the same volume of physiological saline.

[0134] Fifteen minutes after modeling, 10 μL of physiological saline was instilled into each eye of the blank control group and the model group; in the test compound group, 10 μL of low-dose and high-dose Formula I compound tromethamine salt eye drops were instilled into each eye, and 10 μL of 0.1% pranoprofen eye drops were instilled into each eye of the pranoprofen eye drop group.

[0135] One hour after modeling, the rats were euthanized, and the bulbar conjunctiva was harvested, weighed, and the swelling rate or inhibition rate was calculated. The bulbar conjunctiva was fixed in tissue fixative and then subjected to H&E staining.

[0136] Experimental results showed that the test compound eye drops had a significant inhibitory effect on the experimental acute conjunctival edema model induced by arachidonic acid. Compared with pranoprofen eye drops, the high-dose group of test compound eye drops showed a higher inhibitory effect and better therapeutic efficacy.

[0137] Table 9. Inhibitory effect of the test compound eye drops on arachidonic acid-induced experimental acute conjunctival edema.

[0138] Group Swelling rate / inhibition rate Blank control group (Group A) — Model Group (Group B) 24.16％ Pranoprofen eye drops group (Group C) 22.11％ Low-dose group (Group D) of the test compound eye drops 20.43％ High-dose group (Group E) of the test compound eye drops 30.75％

[0139] Note: Inhibition rate (%) = (mB group - mC / D / E group) / mB group × 100%; Swelling rate (%) = (mB group - mA group) / mA group × 100%.

[0140] Experimental conclusions: Arachidonic acid induced conjunctivitis in rats. Compared with the blank control group, the model group showed significant inflammatory infiltration and severe tissue damage in the conjunctival tissue. The pranoprofen eye drop group showed inflammatory infiltration, but the tissue structure remained relatively intact. The low-dose group of the tested compound eye drops showed inflammatory infiltration with relatively intact tissue structure, while the high-dose group showed significantly improved inflammatory infiltration and intact tissue structure.

[0141] 4. Eye irritation studies

[0142] Healthy New Zealand white rabbits were examined under a slit lamp 24 hours before the test to ensure there were no eye lesions before the test began. The left and right eyes of the rabbits were used as self-controls, with the right eye being given the test compound (pranoprofen eye drops, low dose of test compound eye drops, and high dose of test compound eye drops).

[0143] The repeated-dose irritation test was conducted by administering 50 μl of the drug three times a day for seven consecutive days, with the left eye receiving the same amount of physiological saline at the same frequency as a control.

[0144] After each application of medication, observe the rabbit's eyes for symptoms of eye irritation such as frequent blinking, photophobia, and tearing, and score the eye irritation according to the Draize eye irritation scoring principle.

[0145] Table 10 Irritation rating of rabbit eyes

[0146] Group congestion edema secretions Total score Stimulation intensity Pranoprofen eye drops set 0 0 0 0 Non-irritating Low-dose group of test compound eye drops 0 0 0 0 Non-irritating High-dose group of test compound eye drops 0 0 0 0 Non-irritating

[0147] According to the experimental results, no obvious irritation symptoms were observed in any of the drug administration groups, indicating that the eye drops prepared in this invention have simple components, do not contain antibacterial agents or stabilizers, and do not irritate rabbit eyes with repeated administration, demonstrating good biocompatibility.

[0148] Scoring criteria:

[0149]

[0150]

[0151] Eye irritation response scoring standards

[0152] Score 0-1 2-3 4-6 7-9 evaluate Non-irritating Mild stimulation moderate stimulation Intensity stimulation

Claims

1. A single-dose nonsteroidal anti-inflammatory eye drop, characterized by comprising, It comprises: (a) a compound of Formula I tromethamine salt (b) an ophthalmic preparation adjuvant; The content of the compound of Formula I tromethamine salt is 0.01-1.0wt%; The ophthalmic preparation adjuvant includes a metal ion complexing agent, a thickening agent, an osmotic pressure regulator, a pH regulator; the eye drops do not contain a bacteriostatic agent.

2. The eye drop according to claim 1, characterized by, The metal ion complexing agent is at least one selected from the group consisting of edetate disodium, calcium disodium edetate, dihydroxyethyl glycine, ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetraacetic acid and penta(carboxymethyl)diethylenetriamine, amino triacetic acid, sodium gluconate, sodium citrate, tartaric acid; the content of the metal ion complexing agent is 0.01-0.1wt%.

3. The eye drop according to claim 1, characterized by, The thickening agent is at least one selected from the group consisting of sodium hyaluronate, hypromellose, polysorbate 20, carbomer, xanthan gum, polyethylene glycol, polyvinyl alcohol, polyvinylpyrrolidone, sodium carboxymethyl cellulose, methyl cellulose, carboxymethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, sodium polyacrylate, microcrystalline cellulose, povidone, glycerol, sodium alginate, poloxamer, dextran, polycarbophil; the content of the thickening agent is 0.01-0.1wt%.

4. The eye drop according to claim 1, characterized by, The osmotic pressure regulator is at least one selected from the group consisting of sodium chloride, potassium chloride, boric acid, borax, glycerol, propylene glycol, mannitol, sodium sulfate, potassium sulfate, sodium nitrate, potassium nitrate, glucose, sorbitol, sodium cromoglicate; the content of the osmotic pressure regulator is 0.5-2.0wt%.

5. The eye drop according to claim 1, wherein The pH regulator is at least one selected from the group consisting of phosphoric acid, hydrochloric acid, sodium hydroxide, sodium citrate, citric acid, boric acid, borax, sodium acetate, acetic acid; the pH regulator is used to adjust the pH of the eye drops to 6.0-8.0; preferably 6.5-7.

5.

6. The eye drop of claim 1, wherein The compound of Formula I tromethamine salt is in a crystalline form, preferably the compound of Formula I tromethamine salt Form B, the X-ray powder diffraction pattern using Cu-Ka radiation has characteristic diffraction peaks at 2θ values of 6.25±0.2, 8.77±0.2, 17.24±0.2, 17.97±0.2, 19.57±0.2, 20.56±0.2, and the DSC pattern has an endothermic peak at about 112.6℃.

7. The eye drop of claim 1, wherein The eye drops are packaged independently, and the volume of the container of the independent package ranges from 0.4 to 0.8 mL per package.

8. The eye drop of claim 1, wherein It comprises: The compound of Formula I tromethamine salt: 0.05-0.5wt%; The metal ion complexing agent edetate disodium: 0.01-0.1wt%; The thickening agent sodium hyaluronate: 0.01-0.1wt%; The osmotic pressure regulator sodium chloride: 0.5-2wt%; The pH regulator: appropriate amount, adjust the pH to 6.5-7.5; The rest is water.

9. The method for preparing the single-dose nonsteroidal anti-inflammatory eye drops as described in claim 8, characterized in that, It comprises the following steps: Step 1) Weigh the prescribed amount of sodium hyaluronate, fully swell and dissolve in purified water, and prepare for use; Step 2) Weigh the prescribed amount of the compound of Formula I tromethamine salt, edetate disodium, sodium chloride, and pH regulator into the solution of step 1), stir until completely dissolved, adjust the pH value to 6.5-7.5, and adjust the osmotic pressure molarity to 260-320 mOsmol / kg; Step 3) add purified water to 500ml, filter, seal, sterilize at 121℃ high pressure steam for 15min; Step 4) carry out single dose filling and sealing in a "blow-filling-seal" integrated machine, the filling amount is 0.4-0.8ml, cool to 4℃ and keep for 1-3h, and then obtain.

10. The use of the single dose non-steroidal anti-inflammatory eye drops according to any one of claims 1-8 in the preparation of a medicine for preventing or treating ophthalmic diseases.

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