A contact lens care solution containing recombinant humanized collagen and a method of making the same

Abstract

The present application relates to the field of contact lens care technology, and particularly relates to a contact lens care solution containing recombinant humanized collagen and a preparation method thereof.The present application provides a contact lens care solution containing recombinant humanized collagen, which comprises the following components: recombinant humanized collagen type III, recombinant humanized collagen type IV, cationic hyaluronic acid sodium, pyridoxine hydrochloride, D-panthenol, N-acetyl peptides, poloxamer 188, stabilizer, buffer, preservative and bactericidal agent, osmotic pressure regulator and water.The care solution can effectively relieve the discomfort such as dry eyes, foreign body sensation and conjunctival hyperemia caused by long-term wearing of contact lenses, and simultaneously solves the problem of easy degradation and inactivation of recombinant humanized collagen in aqueous solution, and the product has excellent stability, is mild and non-irritating, and is suitable for long-term wearers.

Description

Technical Field

[0001] This invention relates to the field of contact lens care technology, and in particular to a contact lens care solution containing recombinant humanized collagen and its preparation method. Background Technology

[0002] Contact lenses offer advantages such as aesthetics, convenience, and good visual correction. However, long-term wear can lead to corneal epithelial hypoxia, shortened tear film breakup time, and micro-damage to the corneal epithelium. This can cause discomfort such as dry eyes, foreign body sensation, conjunctival congestion, and eye strain. In severe cases, it can induce bacterial or fungal keratitis and even lead to irreversible vision damage.

[0003] The core functions of contact lens care solutions are disinfection, cleaning, protein removal, moisturizing, and storage of lenses, directly determining the safety and comfort of wearing them. Commercially available ordinary care solutions can only meet basic disinfection and cleaning needs, lacking corneal barrier repair functions and failing to address the ocular surface damage caused by long-term lens wear; some care solutions with added moisturizing ingredients can only provide short-term lubrication and cannot fundamentally repair damaged corneal barriers.

[0004] Therefore, there is an urgent need for a contact lens care solution that has efficient disinfection, cleaning and protein removal, long-lasting moisturizing, corneal barrier repair functions, good ocular surface compatibility, and is non-irritating. Summary of the Invention

[0005] The purpose of this invention is to provide a contact lens care solution containing recombinant humanized collagen. This solution can effectively relieve discomfort such as dry eyes, foreign body sensation, and conjunctival congestion caused by long-term contact lens wear. At the same time, it solves the problem of easy degradation and inactivation of recombinant humanized collagen in aqueous solution. The product has excellent stability, is gentle and non-irritating, and is suitable for long-term wearers.

[0006] To achieve the above-mentioned objectives, the present invention provides the following technical solution: This invention provides a contact lens care solution containing recombinant humanized collagen, comprising the following components in parts by weight: 0.05-0.5 parts of recombinant type III humanized collagen, 0.02-0.2 parts of recombinant type IV humanized collagen, 0.5-1.8 parts of cationic sodium hyaluronate, 0.05-0.2 parts of pyridoxine hydrochloride, 0.5-2.0 parts of D-panthenol, 0.5-3.0 parts of N-acetylcarnosine, 0.5-3 parts of poloxamer 188, 4.5-10 parts of stabilizer, 3.3-5.3 parts of buffer, 0.1-0.4 parts of preservative and bactericide, 4-6 parts of osmotic pressure regulator, and 900-1000 parts of water.

[0007] Preferably, the stabilizer includes hydroxypropyl-β-cyclodextrin, trehalose, and L-arginine; The mass ratio of hydroxypropyl-β-cyclodextrin, trehalose, and L-arginine is 1~3:3~6:0.5~1.0.

[0008] Preferably, the buffer includes boric acid, borax, sodium dihydrogen phosphate, and disodium hydrogen phosphate; The mass ratio of boric acid, borax, sodium dihydrogen phosphate and disodium hydrogen phosphate is 2.5~4.0:0.5~1.2:0.2~0.5:0.1~0.3.

[0009] Preferably, the preservative and bactericide includes polyhexamethylene biguanide hydrochloride, polyquaternium-1, and disodium ethylenediaminetetraacetate; The mass ratio of polyhexamethylene biguanide hydrochloride, polyquaternium-1 and disodium ethylenediaminetetraacetate is 0.0005~0.002:0.005~0.015:0.1~0.3.

[0010] Preferably, the osmotic pressure regulator includes sodium chloride and potassium chloride; The mass ratio of sodium chloride to potassium chloride is 3.5~5.0:0.5~1.0.

[0011] The present invention also provides a method for preparing the aforementioned contact lens care solution, comprising the following steps: (1) Mix recombinant type III humanized collagen, recombinant type IV humanized collagen and hydroxypropyl-β-cyclodextrin at 2 to 4 times the total weight of collagen to obtain a pre-inclusion solution; (2) Mix 75-85% of the total volume of water with osmotic pressure regulator, buffer, L-arginine and disodium ethylenediaminetetraacetate to obtain the base solution; (3) Mix the base solution, the remaining hydroxypropyl-β-cyclodextrin, trehalose, pyridoxine hydrochloride, D-panthenol, N-acetylcarnosine and cationic sodium hyaluronate to obtain a mixed solution; (4) Mix the mixed solution with the pre-encapsulated solution, poloxamer 188, polyhexamethylene biguanide hydrochloride, polyquaternium-1, and the remaining water, filter, and obtain the contact lens care solution.

[0012] Preferably, in step (1), the mixing is a low-speed stirring mixture; the mixing speed is 20~40 rpm; the mixing time is 30~40 min; the mixing temperature is 20~30℃; and the mixing needs to be done in the dark.

[0013] Preferably, in step (2), the mixing is a low-speed stirring mixture; the mixing speed is 40~60 rpm; the mixing temperature is 25~35℃; and the pH of the base liquid is 7.2~7.4.

[0014] Preferably, in step (3), the mixing is a low-speed stirring mixture; the mixing speed is 40~60 rpm.

[0015] Preferably, in step (4), the mixing is a low-speed stirring mixture; the mixing speed is 40~60 rpm, and the mixing temperature is 30~35℃.

[0016] Beneficial effects: This invention provides a contact lens care solution containing recombinant humanized collagen. It utilizes a combination of recombinant type III and type IV humanized collagen to fundamentally alleviate problems such as corneal damage, tear film instability, dry eyes, and foreign body sensation caused by long-term wear. Hydroxypropyl-β-cyclodextrin, trehalose, and L-arginine are used as stabilizers. The addition of cationic sodium hyaluronate significantly prolongs the retention time of moisturizing ingredients, reduces friction between the lens and cornea, and improves wearing comfort. Pyridoxine hydrochloride, D-panthenol, and N-acetylcarnosine are combined to achieve synergistic anti-inflammatory, antioxidant, and soothing effects. A preservative system composed of polyhexamethylene biguanide hydrochloride, polyquaternium-1, and disodium EDTA is used, which work synergistically to achieve highly effective killing of common pathogens at extremely low concentrations. Furthermore, a dual buffer system (boric acid, borax, sodium dihydrogen phosphate, and disodium hydrogen phosphate) and osmotic pressure regulators (sodium chloride and potassium chloride) are used to maintain pH stability and osmotic pressure, reducing discomfort during wear. This invention also provides a preparation process for contact lens care solution. This process is simple, with controllable parameters, requires no complex equipment, and uses low-temperature, low-speed stirring throughout to maximize the preservation of the bioactivity of functional components. It exhibits good batch stability and is suitable for large-scale industrial production. The process uses hydroxypropyl-β-cyclodextrin and recombinant humanized collagen for pre-encapsulation, resulting in a stable system that forms a protective barrier for collagen molecules in advance, preventing damage to their activity in subsequent processes and solving the problem of easy inactivation of collagen. Detailed Implementation

[0017] This invention provides a contact lens care solution containing recombinant humanized collagen, comprising the following components in parts by weight: 0.05-0.5 parts of recombinant type III humanized collagen, preferably 0.2-0.35 parts, and more preferably 0.275 parts; The recombinant type IV humanized collagen is 0.02-0.2 parts, preferably 0.07-0.15 parts, and more preferably 0.11 parts; The amount of cationic sodium hyaluronate is 0.5 to 1.8 parts, preferably 0.9 to 1.4 parts, and more preferably 1.15 parts; The amount of pyridoxine hydrochloride is 0.05 to 0.2 parts, preferably 0.1 to 0.15 parts, and more preferably 0.125 parts; D-Panthenol 0.5~2.0 parts, preferably 1~1.5 parts, more preferably 1.25 parts; 0.5 to 3.0 parts of N-acetylcarnosine, preferably 1.2 to 2.3 parts, and more preferably 1.75 parts; Poloxamer 188: 0.5-3.0 parts, preferably 1.2-2.3 parts, more preferably 1.75 parts; The stabilizer is 4.5 to 10 parts, preferably 5.5 to 9 parts, and more preferably 7.25 parts; The buffer is 3.3 to 5.3 parts, preferably 3.8 to 4.8 parts, and more preferably 4.3 parts; The amount of preservative and bactericide is 0.1 to 0.4 parts, preferably 0.15 to 0.35 parts, and more preferably 0.25 parts; The osmotic pressure regulator is 4 to 6 parts, preferably 4.5 to 5.5 parts, and more preferably 5 parts; Water: 900-1000 parts, preferably 930-970 parts, and even more preferably 950 parts.

[0018] In this invention, type III collagen can promote the proliferation and migration of human corneal epithelial cells and accelerate the repair of epithelial wounds; type IV collagen can repair the corneal basement membrane and maintain the integrity of the corneal barrier. The two work together to achieve full-thickness repair from the epithelial layer to the basement membrane. The recombinant humanized collagen used is a fully human sequence, with no animal-derived components, no immunogenicity, no viral risk, high biosafety, and good batch stability. This invention uses cationic sodium hyaluronate instead of ordinary sodium hyaluronate. It carries a positive charge and can be adsorbed onto the negatively charged lens surface and corneal epithelium through electrostatic interaction, forming a uniform and stable moisturizing hydration film. This significantly prolongs the retention time of moisturizing ingredients, reduces friction between the lens and the cornea, and improves wearing comfort. In synergy with dual collagen, the dual effects of moisturizing and repair are superimposed, further relieving dry eyes when wearing the lens. Pyridoxine hydrochloride inhibits eye inflammation and relieves conjunctival congestion and eye fatigue; D-panthenol promotes epithelial healing and has excellent moisturizing properties; N-acetylcarnosine scavenges free radicals, resists glycation and oxidation, and relieves oxidative damage to the eyes; poloxamer 188 can effectively clean protein deposits and oil stains on lenses and improve lens hydrophilicity.

[0019] In this invention, the stabilizer includes hydroxypropyl-β-cyclodextrin, trehalose, and L-arginine; The mass ratio of hydroxypropyl-β-cyclodextrin, trehalose, and L-arginine is 1~3:3~6:0.5~1.0, preferably 1.5~2.5:4~5:0.7~0.8, and more preferably 2:4.5:0.75.

[0020] Hydroxypropyl-β-cyclodextrin encapsulates collagen molecules through hydrophobic interactions, preventing damage from the external environment; trehalose, as a protein protectant, forms a hydration film on the surface of collagen molecules, maintaining their spatial conformation; L-arginine inhibits collagen aggregation and degradation, while regulating the pH of the system. The three work synergistically to significantly improve the stability of collagen aqueous solutions.

[0021] In this invention, the buffer includes boric acid, borax, sodium dihydrogen phosphate, and disodium hydrogen phosphate; The mass ratio of boric acid, borax, sodium dihydrogen phosphate, and disodium hydrogen phosphate is 2.5~4.0:0.5~1.2:0.2~0.5:0.1~0.3, preferably 3.0~3.5:0.7~1.0:0.3~0.4:0.15~0.25, and more preferably 3.25:0.85:0.35:0.2.

[0022] In this invention, the preservative and bactericide includes polyhexamethylene biguanide hydrochloride, polyquaternium-1, and disodium ethylenediaminetetraacetate; The mass ratio of polyhexamethylene biguanide hydrochloride, polyquaternium-1, and disodium ethylenediaminetetraacetate is 0.0005~0.002:0.005~0.015:0.1~0.3, preferably 0.001~0.0015:0.008~0.012:0.15~0.25, and more preferably 0.00125:0.01:0.2; It adopts a preservative system composed of polyhexamethylene biguanide hydrochloride, polyquaternium-1, and disodium EDTA. The three components work synergistically to achieve highly effective killing of common pathogens at extremely low concentrations. It avoids the use of high concentrations of preservatives, does not irritate the ocular mucosa, and does not damage the ocular barrier with long-term use. It is suitable for people with sensitive eyes and long-term wearers.

[0023] In this invention, the osmotic pressure regulator includes sodium chloride and potassium chloride; The mass ratio of sodium chloride to potassium chloride is 3.5~5.0:0.5~1.0, preferably 4.0~4.5:0.7~0.8, and more preferably 4.25:0.75.

[0024] The present invention also provides a method for preparing the aforementioned contact lens care solution, comprising the following steps: (1) Mix recombinant type III humanized collagen, recombinant type IV humanized collagen and hydroxypropyl-β-cyclodextrin at 2 to 4 times the total weight of collagen to obtain a pre-inclusion solution; (2) Mix 75-85% of the total volume of water with osmotic pressure regulator, buffer, L-arginine and disodium ethylenediaminetetraacetate to obtain the base solution; (3) Mix the base solution, the remaining hydroxypropyl-β-cyclodextrin, trehalose, pyridoxine hydrochloride, D-panthenol, N-acetylcarnosine and cationic sodium hyaluronate to obtain a mixed solution; (4) Mix the mixed solution with the pre-encapsulated solution, poloxamer 188, polyhexamethylene biguanide hydrochloride, polyquaternium-1, and the remaining water, filter, and obtain the contact lens care solution.

[0025] In this invention, in step (1), the mixing is a low-speed stirring mixing; the mixing speed is 20~40 rpm, preferably 25~35 rpm, and more preferably 30 rpm; the mixing time is 30~40 min, preferably 33~37 min, and more preferably 35 min; the mixing temperature is 20~30℃, preferably 23~27℃, and more preferably 25℃; the mixing needs to be done in the dark.

[0026] In this invention, in step (2), the mixing is a low-speed stirring mixing; the mixing speed is 40~60 rpm, preferably 45~55 rpm, and more preferably 50 rpm; the mixing temperature is 25~35℃, preferably 28~32℃, and more preferably 30℃; the pH of the base liquid is 7.2~7.4, preferably 7.3.

[0027] In this invention, in step (3), the mixing is a low-speed stirring mixture; the mixing is complete when the mixture is dissolved, and the mixing speed is 40~60 rpm, preferably 45~55 rpm, and more preferably 50 rpm; After mixing and dissolving in step (3), the mixture needs to be allowed to stand to remove bubbles, resulting in a mixed solution. In this invention, in step (4), the mixing is a low-speed stirring mixing until dissolved, the mixing speed is 40~60 rpm, preferably 45~55 rpm, more preferably 50 rpm, and the mixing temperature is 30~35℃, preferably 32~33℃, more preferably 32.5℃. The filtration method is as follows: filtration is performed using a 0.22μm polyethersulfone sterile filter membrane.

[0028] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0029] In this invention, human corneal epithelial cells (HCEC) were purchased from Shanghai Yubo Biotechnology Co., Ltd., L929 mouse fibroblasts were purchased from Shanghai Xuanke Biotechnology Co., Ltd., and SPF-grade New Zealand rabbits were purchased from Qingdao Kangda Biotechnology Co., Ltd. All components of this invention are commercially available.

[0030] Example 1: Contact lens solution containing recombinant humanized collagen

[0031] (1) 0.2g of recombinant type III humanized collagen, 0.1g of recombinant type IV humanized collagen and 0.9g of hydroxypropyl-β-cyclodextrin were mixed at low speed for 35min at 25℃ and 30rpm in the dark to obtain a pre-inclusion solution; (2) Take 760g of water for injection, control the temperature at 30℃, and add 0.5g L-arginine, 3.5g sodium chloride, 1.0g potassium chloride, 2.5g boric acid, 1.2g borax, 0.2g sodium dihydrogen phosphate, 0.3g disodium hydrogen phosphate, and 0.1g disodium ethylenediaminetetraacetate in sequence at 50rpm. Stir and mix until completely dissolved, and the pH is 7.3 to obtain the base solution; (3) Add 2g hydroxypropyl-β-cyclodextrin, 3g trehalose, 0.2g pyridoxine hydrochloride, 0.5g D-panthenol, 3.0g N-acetylcarnosine and 0.5g cationic sodium hyaluronate to the base solution at a speed of 45 rpm, stir until completely dissolved, let stand to remove bubbles, and obtain a mixed solution. (4) At a temperature of 35℃ and a rotation speed of 60 rpm, add the pre-encapsulation solution to the mixed solution and mix until it is uniform and transparent without precipitation. Then add 1.5g poloxamer 188, 0.0005g polyhexamethylene biguanide hydrochloride, 0.015g polyquaternium-1, and 190g water for injection. Stir and mix until completely dissolved. Filter the solution using a 0.22μm polyethersulfone sterile filter membrane to obtain the contact lens care solution.

[0032] Example 2: Contact lens solution containing recombinant humanized collagen

[0033] (1) 0.05g of recombinant type III humanized collagen, 0.2g of recombinant type IV humanized collagen and 1.0g of hydroxypropyl-β-cyclodextrin were mixed at low speed for 40min at 20℃ and 20rpm in the dark to obtain a pre-inclusion solution; (2) Take 768g of water for injection, control the temperature at 25℃, and add 1.0g L-arginine, 4.0g sodium chloride, 0.8g potassium chloride, 3g boric acid, 1.0g borax, 0.4g sodium dihydrogen phosphate, 0.2g disodium hydrogen phosphate, and 0.2g disodium ethylenediaminetetraacetate in sequence at 40rpm. Stir and mix until completely dissolved, and the pH is 7.2 to obtain the base solution; (3) Add 1.5g hydroxypropyl-β-cyclodextrin, 4g trehalose, 0.05g pyridoxine hydrochloride, 2.0g D-panthenol, 1.0g N-acetylcarnosine and 1.0g cationic sodium hyaluronate to the base solution at 50 rpm, stir until completely dissolved, let stand to remove bubbles, and obtain a mixed solution. (4) At a temperature of 35℃ and a rotation speed of 40 rpm, add the pre-encapsulation solution to the mixed solution and mix until it is uniform and transparent without precipitation. Then add 3.0 g poloxamer 188, 0.001 g polyhexamethylene biguanide hydrochloride, 0.01 g polyquaternium-1 and 192 g water for injection. Stir and mix until completely dissolved. Filter with a 0.22 μm polyethersulfone sterile filter membrane to obtain the contact lens care solution.

[0034] Example 3: Contact lens solution containing recombinant humanized collagen

[0035] (1) 0.4g of recombinant type III humanized collagen, 0.15g of recombinant type IV humanized collagen and 1.1g of hydroxypropyl-β-cyclodextrin were mixed at low speed for 30min under the conditions of 30℃, 40rpm and protection from light to obtain a pre-inclusion solution; (2) Take 784g of water for injection, control the temperature at 35℃, and add 0.8g of L-arginine, 5.0g of sodium chloride, 0.5g of potassium chloride, 4.0g of boric acid, 0.5g of borax, 0.5g of sodium dihydrogen phosphate, 0.1g of disodium hydrogen phosphate and 0.3g of disodium ethylenediaminetetraacetate in sequence at 60rpm. Stir and mix until completely dissolved, and the pH is 7.4 to obtain the base solution; (3) Add 1g hydroxypropyl-β-cyclodextrin, 6g trehalose, 0.1g pyridoxine hydrochloride, 1.0g D-panthenol, 2.0g N-acetylcarnosine and 1.8g cationic sodium hyaluronate to the base solution at 60 rpm, stir until completely dissolved, let stand to remove bubbles, and obtain a mixed solution. (4) At a temperature of 35℃ and a rotation speed of 50 rpm, add the pre-encapsulation solution to the mixed solution and mix until it is uniform and transparent without precipitation. Then add 2.0 g poloxamer 188, 0.002 g polyhexamethylene biguanide hydrochloride, 0.005 g polyquaternium-1, and 196 g water for injection. Stir and mix until completely dissolved. Filter the solution using a 0.22 μm polyethersulfone sterile filter membrane to obtain the contact lens care solution.

[0036] Comparative Example 1: Basic Contact Lens Solution

[0037] (1) Take 760g of water for injection, control the temperature at 30℃, and add 0.5g L-arginine, 3.5g sodium chloride, 1.0g potassium chloride, 2.5g boric acid, 1.2g borax, 0.2g sodium dihydrogen phosphate, 0.3g disodium hydrogen phosphate, and 0.1g disodium ethylenediaminetetraacetate in sequence at 50rpm. Stir and mix until completely dissolved, and the pH is 7.3 to obtain the base solution; (2) Add 2.9g hydroxypropyl-β-cyclodextrin, 3g trehalose, 0.2g pyridoxine hydrochloride, 0.5g D-panthenol, 3.0g N-acetylcarnosine and 0.5g cationic sodium hyaluronate to the base solution at a speed of 45 rpm, stir until completely dissolved, let stand to remove bubbles, and obtain a mixed solution. (3) At a temperature of 35℃ and a rotation speed of 60rpm, add 1.5g poloxamer 188, 0.0005g polyhexamethylene biguanide hydrochloride, 0.015g polyquaternium-1 and 190g water for injection to the mixed solution, stir and mix until completely dissolved, and filter with a 0.22μm polyethersulfone sterile filter membrane to obtain contact lens care solution.

[0038] Comparative Example 2: Contact lens solution containing recombinant humanized collagen

[0039] Referring to Example 1, the difference from Example 1 is that no recombinant type IV humanized collagen is added, while the other components and preparation steps are the same as in Example 1.

[0040] Comparative Example 3: Contact lens solution containing recombinant humanized collagen

[0041] Referring to Example 1, the difference from Example 1 is that no recombinant type III humanized collagen is added, while the other components and preparation steps are the same as in Example 1.

[0042] Comparative Example 4: Contact lens solution containing recombinant humanized collagen

[0043] (1) Take 760g of water for injection, control the temperature at 30℃, and add 0.5g L-arginine, 3.5g sodium chloride, 1.0g potassium chloride, 2.5g boric acid, 1.2g borax, 0.2g sodium dihydrogen phosphate, 0.3g disodium hydrogen phosphate, and 0.1g disodium ethylenediaminetetraacetate in sequence at 50rpm. Stir and mix until completely dissolved, and the pH is 7.3 to obtain the base solution; (2) Add 2.9g hydroxypropyl-β-cyclodextrin, 3g trehalose, 0.2g pyridoxine hydrochloride, 0.5g D-panthenol, 3.0g N-acetylcarnosine, 0.5g cationic sodium hyaluronate, 0.2g recombinant type III humanized collagen and 0.1g recombinant type IV humanized collagen to the base solution at a speed of 45 rpm. Stir until completely dissolved, let stand to remove bubbles, and obtain a mixed solution. (3) At a temperature of 35℃ and a rotation speed of 60rpm, add 1.5g poloxamer 188, 0.0005g polyhexamethylene biguanide hydrochloride, 0.015g polyquaternium-1 and 190g water for injection to the mixed solution, stir and mix until completely dissolved, and filter with a 0.22μm polyethersulfone sterile filter membrane to obtain contact lens care solution.

[0044] Comparative Example 5: Contact lens care solution containing recombinant humanized collagen

[0045] (1) Take 760g of water for injection, control the temperature at 30℃, and add 0.5g L-arginine, 3.5g sodium chloride, 1.0g potassium chloride, 2.5g boric acid, 1.2g borax, 0.2g sodium dihydrogen phosphate, 0.3g disodium hydrogen phosphate, and 0.1g disodium ethylenediaminetetraacetate in sequence at 50rpm. Stir and mix until completely dissolved, and the pH is 7.3 to obtain the base solution; (2) Add 3g trehalose, 0.2g pyridoxine hydrochloride, 0.5g D-panthenol, 3.0g N-acetylcarnosine, 0.5g cationic sodium hyaluronate, 0.2g recombinant type III humanized collagen and 0.1g recombinant type IV humanized collagen to the base solution at a speed of 45 rpm. Stir until completely dissolved, let stand to remove bubbles, and obtain a mixed solution. (3) At a temperature of 35℃ and a rotation speed of 60rpm, add 1.5g poloxamer 188, 0.0005g polyhexamethylene biguanide hydrochloride, 0.015g polyquaternium-1 and 190g water for injection to the mixed solution, stir and mix until completely dissolved, and filter with a 0.22μm polyethersulfone sterile filter membrane to obtain contact lens care solution.

[0046] Comparative Example 6: Contact Lens Solution Containing Recombinant Humanized Collagen

[0047] Referring to Example 1, the difference from Example 1 is that cationized sodium hyaluronate is replaced with sodium hyaluronate, while other components and preparation steps are the same as in Example 1.

[0048] Experiment Example 1 Stability Test

[0049] The contact lens solutions described in Examples 1-3 and Comparative Examples 1-6 were placed in an accelerated stability test chamber under the conditions of 40℃±2℃ and 75%±5% relative humidity, and stored in the dark for 6 months. Samples were taken at 0 months, 1 month, 3 months, and 6 months to test the appearance, pH fluctuation, and collagen content retention rate (determined by high performance liquid chromatography, with 0 months as 100%). The results are shown in Tables 1-2. Table 1 Results of accelerated stability tests

[0050] Table 2 Results of Collagen Content Retention Rate Measurement

[0051] The results showed that after 6 months of accelerated testing, Examples 1-3 of the present invention remained clear and transparent, with pH fluctuations all less than 0.1, demonstrating excellent system stability. Comparative Example 4, which did not use the pre-encapsulation process, and Comparative Example 5, which did not contain hydroxypropyl-β-cyclodextrin, showed obvious turbidity, protein precipitation, significantly increased pH fluctuations, and a substantial decrease in collagen retention rate, proving that the hydroxypropyl-β-cyclodextrin pre-encapsulation process can significantly inhibit protein aggregation and precipitation, and improve the storage stability of collagen in the system. Comparative Examples 2 and 3, which consisted of a single collagen component, had lower protein retention rates than the examples, proving that the combination of type III and type IV recombinant humanized collagen has a synergistic stabilizing effect.

[0052] Experiment Example 2: Determination of bactericidal performance

[0053] The bactericidal rate of the contact lens solutions described in Examples 1-3 and Comparative Examples 1-6 was determined with reference to GB 19192-2003 "Hygienic Requirements for Contact Lens Care Solutions". The results are shown in Table 3. Table 3 Results of sterilization rate determination

[0054] The results showed that the bactericidal rates of Examples 1-3 against the three pathogenic bacteria were all ≥99.98%, and the bactericidal rate against Candida albicans was ≥99.7%, which fully met the mandatory requirements of GB 19192-2003, and the broad-spectrum antibacterial performance was stable.

[0055] Experimental Example 3: Moisturizing and Wettability Determination

[0056] Lens water retention rate determination: Take a silicone hydrogel lens, blot dry the surface free water and weigh it (W0), immerse it in 5 mL of the contact lens care solution described in Examples 1-3 and Comparative Examples 1-6 respectively, and protect it from light at 37°C for 24 hours. Take it out, blot dry the surface free water and weigh it (W1); then place it in an environment of 37°C and 30% relative humidity for 4 hours and 8 hours respectively, and weigh it (W2, W3). Calculate the water retention rate at different times. Set 6 parallel samples for each group to verify the long-lasting moisturizing ability. The water retention rate (%) = (W2-W0) / (W1-W0)×100%; Contact angle measurement (°): The static water contact angle of the silicone hydrogel lens surface was measured using a contact angle measuring instrument after soaking for 24 hours. The smaller the contact angle, the better the hydrophilicity and wettability. Six parallel samples were set up for each group. Surface tension measurement (mN / m): The surface tension of the test sample was measured at 25℃ using the pendant drop method. The lower the surface tension, the better the wettability and the easier it is to spread on the lens surface. Three parallel samples were set for each group. The results are shown in Table 4. Table 4 Results of Moisturizing and Wetting Properties Tests

[0057] Results Analysis: The silicone hydrogel lenses of Examples 1-3 of this invention, after soaking, maintained a water retention rate of nearly 90% after 4 hours and over 77% after 8 hours, far superior to all comparative examples, demonstrating excellent long-lasting moisturizing ability and significantly alleviating dry eye symptoms caused by prolonged lens wear. The water contact angle of the lens surface in the examples was all below 35°, and the surface tension was as low as 34.76 mN / m, far lower than the comparative examples, proving that the care solution can significantly reduce the surface tension of the lens, improve hydrophilicity and wettability, and easily spread evenly on the lens surface to form a stable moisturizing hydration layer. The moisturizing effect of Comparative Example 6 (ordinary sodium hyaluronate) was significantly lower than that of Example 1, proving that cationic sodium hyaluronate can be better adsorbed onto the negatively charged silicone hydrogel lens surface through electrostatic interaction to form a long-lasting moisturizing layer, with moisturizing performance far superior to ordinary sodium hyaluronate. The examples of two collagen complexes showed significantly better moisturizing effects than the comparative examples of single collagen, proving that recombinant type III and type IV humanized collagen have a synergistic moisturizing effect.

[0058] Experimental Example 4: Compatibility Test

[0059] Silicone hydrogel lenses were divided into 9 groups of 10 lenses each. Each group was immersed in 5 mL of the contact lens care solution described in Examples 1-3 and Comparative Examples 1-6. The lenses were immersed continuously at 37°C in the dark for 30 days, and the test samples were replaced every 3 days to simulate daily use. The diameter, base curve, oxygen permeability coefficient Dk / t, and equilibrium water content of the lenses were measured before and after immersion. The rate of change of parameters was calculated. Each group had 10 parallel samples. The experimental results are shown in Table 5. Table 5 Compatibility test results

[0060] The results showed that after continuous immersion of the silicone hydrogel lenses in Examples 1-3 of the present invention for 30 days, the change rates of lens diameter, base curve, oxygen permeability coefficient Dk / t, and water content were all less than 0.5%. The lenses showed no deformation, no turbidity, and no parameter exceeding the standard, demonstrating excellent compatibility with silicone hydrogel lenses and not affecting the lens's fit, core oxygen supply safety, or wearing comfort. The change rates of lens parameters in Comparative Examples 4 and 5 were significantly higher than those in the Examples, especially the difference in the change rate of oxygen permeability coefficient, proving that unencapsulated collagen aggregates and precipitates on the lens surface, blocking the lens's oxygen permeability channels and seriously affecting the lens's core safety performance, further verifying the necessity of the pre-encapsulation process.

[0061] Experimental Example 5: Assay on the Repair Activity of Corneal Epithelial Cells

[0062] Cell proliferation experiment: HCEC cells in logarithmic growth phase were used, and 5 × 10⁻⁶ cells were cultured together. 3Cells were seeded per well in 96-well plates and cultured for 24 hours. Then, DMEM medium containing 10% (v / v) of the contact lens care solutions described in Examples 1-3 and Comparative Examples 1-6 was added, while an equal volume of serum-free DMEM was added to the blank control group. Each group had 6 replicates. After 48 hours of culture, cell viability was determined using the CCK-8 assay, with the blank control group as 100%. Cell scratch repair assay: HCEC cells were cultured to a confluence of over 90%, and uniform scratches were made using a sterile pipette tip. After rinsing with PBS, serum-free DMEM containing 10% (v / v) of the contact lens care solutions described in Examples 1-3 and Comparative Examples 1-6 was added. An equal volume of serum-free DMEM was added to the blank control group. Each group had 3 replicates. Photos were taken at 0h and 24h, and the scratch healing rate was calculated using ImageJ software. The results are shown in Table 6. Table 6. Effects of contact lens solution on HCEC cell proliferation and scratch repair.

[0063] The results showed that Examples 1-3 of the present invention could significantly promote HCEC cell proliferation, with cell survival rates exceeding 150% and a 24-hour scratch healing rate approaching 80%, far superior to the blank control group and all comparative examples, demonstrating excellent corneal epithelial repair activity and accelerating the healing of corneal micro-damage caused by contact lens wear. The repair effects of comparative examples 2 and 3, which consisted of a single collagen component, were significantly lower than those of the examples, proving that the combination of recombinant type III collagen (promoting epithelial cell proliferation and repair) and type IV collagen (maintaining the integrity of the corneal basement membrane barrier) has a synergistic repair effect, achieving full-thickness corneal epithelial repair. The repair activity of comparative examples 4 and 5 decreased significantly, proving that the pre-encapsulation process is the core key to maintaining the biological activity of collagen.

[0064] Experiment Example 6 Safety Test

[0065] Cytotoxicity: The MTT assay was used to detect cytotoxicity in L929 mouse fibroblasts in logarithmic growth phase. Cells were incubated at a concentration of 1×10⁻⁶ cells / cell. 4Each well was seeded with 100 cells / well in a 96-well plate. After culturing for 24 hours, DMEM medium containing 10% or 20% (v / v) of the contact lens care solutions described in Examples 1-3 and Comparative Examples 1-6 was added. An equal volume of complete medium was added to the blank control group, and 20% dimethyl sulfoxide (DMSO) was added to the positive control group. Each group had 6 replicates. After culturing for 24 hours, MTT solution was added and incubated for another 4 hours. The culture medium was then discarded, and DMSO was added to dissolve the formazan crystals. The absorbance was measured at 490 nm using an ELISA reader, and the relative cell proliferation rate (RGR) was calculated. Toxicity was graded according to GB / T16886.5-2017 Medical Device Biological Evaluation Standard: Grade 0: RGR ≥ 100% - no cytotoxicity; Grade 1: 75% ≤ RGR < 100% - very mild toxicity; Grade 2: 50% ≤ RGR < 75% - mild toxicity; Grade 3: 25% ≤ RGR < 50% - moderate toxicity; Grade 4: 1% ≤ RGR < 25% - severe toxicity; Grade 5: RGR < 1% - very severe toxicity. The results are shown in Table 7. Table 7. Relative cell proliferation rate and toxicity grading results

[0066] The results showed that, at concentrations of 10% and 20%, the relative cell proliferation rate of Examples 1-3 of the present invention exceeded 90%, the toxicity grade was 1, which met the GB / T 16886.5-2017 standard for biological evaluation of medical devices, with no obvious cytotoxicity and excellent biocompatibility.

[0067] Irritation test: Forty healthy SPF-grade New Zealand rabbits were randomly divided into 10 groups of 4 rabbits each. Self-control was used, with 0.1 mL of the test sample instilled into the conjunctival sac of the left eye and 0.1 mL of sterile saline in the right eye. The eyelids were closed for 10 seconds. Damage to the cornea, iris, and conjunctiva of the rabbits was observed using a slit-lamp microscope at 1 h, 24 h, 48 h, and 72 h after administration. The intensity of irritation was determined according to the Draize eye irritation rating scale: a total score of 0-0.5 indicated no irritation, 0.6-2.9 indicated mild irritation, and 3 or higher indicated significant irritation. The results are shown in Table 8. Table 8. Average score results of rabbit eye irritation test over 72 hours

[0068] The results showed that the total eye stimulation score of Examples 1-3 of the present invention was less than 0.5 over 72 hours. There was no damage to the cornea or iris, and no obvious abnormalities such as conjunctival hyperemia, edema, or increased secretions. The samples were non-irritating and had excellent ocular safety.

[0069] As can be seen from the above embodiments, the present invention provides a contact lens care solution containing recombinant humanized collagen and its preparation method. This care solution can effectively relieve discomfort such as dry eyes, foreign body sensation, and conjunctival congestion caused by long-term wear of contact lenses. At the same time, it solves the problem that recombinant humanized collagen is easily degraded and inactivated in aqueous solution. The product has excellent stability, is mild and non-irritating, and is suitable for long-term wearers.

[0070] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A contact lens care solution comprising recombinant humanized collagen, characterized in that, The product comprises the following components in parts by weight: 0.05-0.5 parts of recombinant type III humanized collagen, 0.02-0.2 parts of recombinant type IV humanized collagen, 0.5-1.8 parts of cationic sodium hyaluronate, 0.05-0.2 parts of pyridoxine hydrochloride, 0.5-2.0 parts of D-panthenol, 0.5-3.0 parts of N-acetylcarnosine, 0.5-3.0 parts of poloxamer 188, 4.5-10 parts of stabilizer, 3.3-5.3 parts of buffer, 0.1-0.4 parts of preservative and bactericide, 4-6 parts of osmotic pressure regulator, and 900-1000 parts of water.

2. The contact lens care solution of claim 1, wherein, The stabilizers include hydroxypropyl-β-cyclodextrin, trehalose, and L-arginine; The mass ratio of hydroxypropyl-β-cyclodextrin, trehalose, and L-arginine is 1~3:3~6:0.5~1.

0.

3. The contact lens care solution of claim 1, wherein, The buffer includes boric acid, borax, sodium dihydrogen phosphate, and disodium hydrogen phosphate; The mass ratio of boric acid, borax, sodium dihydrogen phosphate and disodium hydrogen phosphate is 2.5~4.0:0.5~1.2:0.2~0.5:0.1~0.

3.

4. The contact lens solution according to claim 1, characterized in that, The preservative and bactericide includes polyhexamethylene biguanide hydrochloride, polyquaternium-1, and disodium ethylenediaminetetraacetate. The mass ratio of polyhexamethylene biguanide hydrochloride, polyquaternium-1 and disodium ethylenediaminetetraacetate is 0.0005~0.002:0.005~0.015:0.1~0.

3.

5. The contact lens solution according to claim 1, characterized in that, The osmotic pressure regulator includes sodium chloride and potassium chloride; The mass ratio of sodium chloride to potassium chloride is 3.5~5.0:0.5~1.

0.

6. The method for preparing the contact lens care solution according to any one of claims 1 to 5, characterized in that, Includes the following steps: (1) Mix recombinant type III humanized collagen, recombinant type IV humanized collagen and hydroxypropyl-β-cyclodextrin at 2 to 4 times the total weight of collagen to obtain a pre-inclusion solution; (2) Mix 75-85% of the total volume of water with osmotic pressure regulator, buffer, L-arginine and disodium ethylenediaminetetraacetate to obtain the base solution; (3) Mix the base solution, the remaining hydroxypropyl-β-cyclodextrin, trehalose, pyridoxine hydrochloride, D-panthenol, N-acetylcarnosine and cationic sodium hyaluronate to obtain a mixed solution; (4) Mix the mixed solution with the pre-encapsulated solution, poloxamer 188, polyhexamethylene biguanide hydrochloride, polyquaternium-1, and the remaining water, filter, and obtain the contact lens care solution.

7. The preparation method according to claim 6, characterized in that, In step (1), the mixing is a low-speed stirring mixture; the mixing speed is 20~40 rpm; the mixing time is 30~40 min; the mixing temperature is 20~30℃; the mixing needs to be done in the dark.

8. The preparation method according to claim 6, characterized in that, In step (2), the mixing is a low-speed stirring mixture; the mixing speed is 40~60 rpm; the mixing temperature is 25~35℃; and the pH of the base liquid is 7.2~7.

4.

9. The preparation method according to claim 6, characterized in that, In step (3), the mixing is a low-speed stirring mixture; the mixing speed is 40~60 rpm.

10. The preparation method according to claim 6, characterized in that, In step (4), the mixing is a low-speed stirring mixture; the mixing speed is 40~60 rpm, and the mixing temperature is 30~35℃.

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