Temperature-sensitive gel composition for slowly releasing nitric oxide as well as preparation method and application of temperature-sensitive gel composition

Abstract

The invention provides a temperature-sensitive gel composition for slowly releasing nitric oxide as well as a preparation method and application thereof, and relates to the technical field of medical gels, the temperature-sensitive gel composition comprises the following components: a temperature-sensitive gel matrix, an NO donor composition, and an optional pH regulator and oxidant; wherein the temperature-sensitive gel matrix comprises the following components: a temperature-sensitive matrix, a phase-change temperature regulator and a solvent, after the temperature-sensitive gel preparation is used, NO gas can be slowly released, the release time can last for more than 12 hours, and the temperature-sensitive gel preparation has a remarkable curative effect in the aspect of long-acting bacteriostasis.

Description

Technical Field

[0001] The present invention relates to the technical field of medical gels, and in particular to a thermosensitive gel composition for slow-release of nitric oxide, a preparation method thereof, and an application thereof. Background Art

[0002] Nitric oxide (NO) is a bioactive gas. The generation pathway of NO in the human body is that nitric oxide synthase (NOS) catalyzes L-arginine (L-Arg) and molecular oxygen as substrates, catalyzes one of the two equivalent guanidyl nitrogens of L-Arg, and generates L-citrulline and releases a small molecule. During wound and mucosal repair, NO plays a variety of important roles:

[0003] NO can inhibit the aggregation and activation of inflammatory cells, reduce the release of inflammatory mediators, and thus alleviate the inflammatory response. It can promote the proliferation and migration of vascular endothelial cells, increase vascular permeability, and thus promote the formation of new blood vessels, which is crucial for wound healing. In addition, nitric oxide has direct antibacterial and antiviral effects, can inhibit bacterial growth and virus replication, and prevent infection. In terms of immune regulation, NO can regulate the functions of immune cells, including macrophages and lymphocytes, and promote immune balance at the wound site. Research shows that NO can promote the migration and proliferation of fibroblasts and keratinocytes, accelerate wound contraction and epithelialization, and also participate in the synthesis and remodeling of collagen, which plays an important role in the strength and functional recovery of wounds. In mucosal repair, NO promotes the repair of mucosal epithelium by regulating the proliferation and differentiation of mucosal cells.

[0004] Although NO has a variety of positive effects in wound and mucosal repair, its application also has some limitations: NO is a very active and unstable gas with a short half-life and is difficult to maintain an effective concentration in the body for a long time. High concentrations of NO may be toxic to human cells, especially under long-term exposure. Due to the gaseous nature of NO, its delivery and release are difficult to control and require special equipment and technologies. In local treatment, it is necessary to ensure that the concentration of NO can achieve the therapeutic effect without causing side effects due to systemic distribution.

[0005] In response to these problems, scholars at home and abroad have adopted various methods to control the slow release of nitric oxide in wounds, skin, and mucosae.

[0006] The patent technology with the publication number CN 103622917 A discloses a system and method for delayed generation of nitric oxide (NO) based on microencapsulated nitrite and acidified hydrogel, which requires specific storage conditions to maintain the activity of the microencapsulated reagent, which may limit its application in different environments. At the same time, controlling the release rate and amount of NO may be relatively complex, and it is necessary to precisely adjust the acidity of the hydrogel and the amount of reducing agent to ensure the effective release of NO.

[0007] Patent technology with publication number CN 104302175 A discloses a pharmaceutical composition that releases nitric oxide (NO) for local delivery of moisture-activated active pharmaceutical ingredients. The preparation of this pharmaceutical composition may involve complex production processes, resulting in relatively high production costs, which may limit its application in large-scale production. The patent mentions the concept of "activation volume", that is, only the volume of water that can moisten the surface of the microcapsules, which requires precise control of the amount of water in practical applications and may increase the operational complexity. At the same time, the gel matrix contains alcohols, oils, etc., and is not suitable for application in the fields of wounds and mucous membranes.

[0008] The above-mentioned patent technologies all use hydrogels as the release mechanism, but generally have problems such as the need for specific modified storage methods, precise adjustment of the amount of reactants, complex production processes and relatively high costs. At the same time, due to the harsh conditions required for releasing NO, some materials with poor biosafety are used, resulting in a narrow application range of the composition. In addition, ordinary gels have poor adhesion, are easy to flow, and stay at the affected area for a short time, resulting in a short drug efficacy time.

[0009] In view of this, the present invention is specifically proposed. Summary of the Invention

[0010] One of the purposes of the present invention is to provide a thermosensitive gel composition for sustained release of nitric oxide to solve at least one of the technical problems existing in the prior art. After the thermosensitive gel preparation is used, it can slowly release NO gas, and the release duration can last for more than 12 hours, showing significant efficacy in long-term antibacterial.

[0011] Another purpose of the present invention is to provide a preparation method of a thermosensitive gel composition for sustained release of nitric oxide.

[0012] The third purpose of the present invention is to provide an application of the thermosensitive gel composition for sustained release of nitric oxide or the preparation method in the preparation of thermosensitive gel preparations.

[0013] In order to achieve the above purposes of the present invention, the following technical solutions are specifically adopted:

[0014] In the first aspect, the present invention provides a thermosensitive gel composition for sustained release of nitric oxide, comprising the following components: a thermosensitive gel matrix, a NO donor composition, and optionally an acid-base regulator and an oxidant;

[0015] Among them, the components of the thermosensitive gel matrix include: a thermosensitive matrix, a phase transition temperature regulator, and a solvent.

[0016] Furthermore, the thermosensitive matrix includes at least one of chitosan, cellulose, poloxamer, poly(N-isopropylacrylamide), and PEG polyester block copolymer;

[0017] Preferably, the phase change temperature regulator comprises at least one of carbomer, acrylic acid polymers and their sodium salts, sodium carboxymethyl cellulose, sodium hyaluronate, sodium alginate, calcium alginate and alginate.

[0018] Preferably, the NO donor composition comprises at least one of inorganic nitrite donors, arginine donors, azodioxolium salts donors and S-nitrosothiols;

[0019] Preferably, the inorganic nitrite donor comprises at least one of lithium nitrite, sodium nitrite, potassium nitrite, cesium nitrite, rubidium nitrite, silver nitrite, magnesium nitrite, calcium nitrite, manganese nitrite and barium nitrite;

[0020] Preferably, the arginine donor comprises at least one of arginine and its derivatives;

[0021] Preferably, the arginine comprises at least one of L-arginine and L-arginine hydrochloride salts;

[0022] Preferably, the azodioxolium salt donors comprise at least one of piperazine, pyrrole, alkylamine or piperidine-derived dioxolium salts, 2-(N,N-diethylamino) dioxolium salts, diazeniumdiolate salts containing a glycol group and azodioxolium salts containing an O-N=N-O group;

[0023] Preferably, the S-nitrosothiols comprise thiol substances containing an -SNO group, and the thiol substances containing an -SNO group comprise at least one of S-nitroso-N-acetylpenicillamine and S-nitrosoglutathione;

[0024] Preferably, the acid-base regulator comprises a substance that can ionize H + or OH - in an aqueous solution;

[0025] Preferably, the acid-base regulator comprises at least one of ascorbic acid, citric acid, acetic acid, lactic acid, nicotinic acid, malic acid, tartaric acid, fumaric acid, mandelic acid, α-hydroxypropionic acid, β-hydroxypropionic acid, β-hydroxybutyric acid, gluconic acid, alginic acid, sodium ethylenediaminetetraacetate, sodium hydroxide, triethanolamine, sodium bicarbonate, arginine and disodium hydrogen phosphate;

[0026] Preferably, the oxidant comprises at least one of ozone, potassium permanganate, potassium perchlorate, hydrogen peroxide, sodium peroxide, urea peroxide, sodium percarbonate, tert-butyl hydroperoxide, peracetic acid, hypochlorous acid and inorganic nitrites;

[0027] Preferably, the solvent comprises water.

[0028] Further, the thermosensitive gel composition for slow-releasing nitric oxide comprises the following components in parts by weight:

[0029] 0.1 - 20 parts of NO donor composition and a thermosensitive gel matrix, wherein the thermosensitive gel matrix comprises 2 - 35 parts of a thermosensitive matrix, 0.5 - 5 parts of a phase transition temperature regulator, and 35 - 98 parts of water;

[0030] Preferably, the weight ratio of the thermosensitive matrix to the phase transition temperature regulator is 0.5 - 70:1.

[0031] Further, the thermosensitive gel composition for slow-releasing nitric oxide further comprises 0.01 - 5 parts of an acid-base regulator;

[0032] Optionally, the components of the thermosensitive gel composition for slow-releasing nitric oxide include: 0.1 - 10 parts of NO donor composition, 0.1 - 2 parts of acid-base regulator, 2 - 25 parts of thermosensitive matrix, 0.5 - 5 parts of phase transition temperature regulator, and 50 - 98 parts of water;

[0033] Optionally, when the NO donor composition is nitrosothiols, its components are 0.1 - 5 parts;

[0034] Optionally, the components of the thermosensitive gel composition for slow-releasing nitric oxide include: 2 parts of S-nitrosoglutathione, 1 part of sodium hydroxide, 20 parts of thermosensitive matrix, 2 parts of phase transition temperature regulator, and 95 parts of water, wherein the thermosensitive matrix comprises poloxamer 407 and poloxamer 188, and the phase transition temperature regulator comprises carbomer and sodium alginate.

[0035] Further, the thermosensitive gel composition for slow-releasing nitric oxide further comprises 0.0001 - 15 parts of an oxidant;

[0036] Optionally, when the NO donor composition comprises sodium nitrite, the weight ratio of the sodium nitrite to the L-arginine donor is 0.25 - 4:1, wherein the L-arginine donor serves as an acid-base regulator;

[0037] Optionally, the components of the thermosensitive gel composition for slow-releasing nitric oxide include: 1 part of NO donor composition, 0.0004 parts of hydrogen peroxide, 10 parts of thermosensitive matrix, 3.5 parts of phase transition temperature regulator, and 95 parts of water, wherein the NO donor composition comprises 0.7 parts of sodium nitrite and 0.3 parts of L-arginine donor, the thermosensitive matrix comprises chitosan and glycerophosphate, and the phase transition temperature regulator comprises 2 parts of carbomer, 1 part of sodium alginate, and 0.5 part of sodium carboxymethylcellulose.

[0038] Further, when the temperature-sensitive matrix comprises poloxamer 407 and poloxamer 188, the weight ratio of poloxamer 407 to poloxamer 188 is 4 to 80:1;

[0039] Optionally, when the temperature-sensitive matrix comprises glycerophosphate or its salt, and chitosan, the weight ratio of glycerophosphate or its salt to chitosan is 1 to 5:1;

[0040] Optionally, when the phase transition temperature regulator comprises carbomer and sodium alginate, the weight ratio of carbomer to sodium alginate is 1 to 10:1;

[0041] Optionally, when the phase transition temperature regulator comprises carbomer, sodium alginate and sodium carboxymethylcellulose, the weight ratio of carbomer, sodium alginate to sodium carboxymethylcellulose is 0.5 to 50:0.5 to 5:1.

[0042] In a second aspect, the present invention provides a method for preparing the temperature-sensitive gel composition for sustained release of nitric oxide, comprising the following steps:

[0043] Mixing the formula amount of temperature-sensitive gel matrix, NO donor composition and optionally acid-base regulator and oxidant to prepare the temperature-sensitive gel composition for sustained release of nitric oxide.

[0044] Further, the method for preparing the temperature-sensitive gel composition for sustained release of nitric oxide comprises the following steps:

[0045] Adding the phase transition temperature regulator and the temperature-sensitive matrix to the NO donor composition solution in sequence, then adding the acid-base regulator or oxidant, and finally filling;

[0046] Preferably, adding the phase transition temperature regulator at a temperature of 50 to 80 °C and dissolving it sufficiently under the condition of a rotation speed of 30 to 400 rpm;

[0047] Preferably, adding the temperature-sensitive matrix under stirring conditions, and then placing it at a temperature of 0 to 4 °C until the temperature-sensitive matrix dissolves;

[0048] Preferably, adding the oxidant or acid-base regulator under stirring conditions, with the rotation speed of stirring being 10 - 20 rpm and the stirring time being 2 to 5 min.

[0049] Further, when the NO donor composition comprises sodium nitrite and arginine, the method for preparing the temperature-sensitive gel composition for sustained release of nitric oxide comprises the following steps: adding the phase transition temperature regulator, the temperature-sensitive matrix and arginine to the sodium nitrite solution in sequence, then adding the oxidant or acid-base regulator, and finally filling;

[0050] Preferably, the process of adding arginine includes: under stirring conditions, adding the arginine solution in batches within 2 - 5 min, with the stirring speed being 10 - 20 rpm and the stirring time being 10 - 30 min.

[0051] In a third aspect, the present invention provides an application of the thermosensitive gel composition for slow - releasing nitric oxide or the preparation method thereof in preparing a thermosensitive gel preparation.

[0052] Compared with the prior art, the present invention has the following beneficial effects:

[0053] The thermosensitive gel composition for slow - releasing nitric oxide provided by the present invention, the phase - change temperature regulator contained in the thermosensitive gel matrix in its components can adjust the phase - change temperature of the thermosensitive gel, and at the same time can also adjust the initial viscosity and consistency of the gel, which can serve as a spatial chamber for gas storage. The phase - change temperature regulator cooperates with the thermosensitive matrix to increase the gel viscosity, and by adjusting the usage amount of the phase - change temperature regulator, the release rate of NO gas is neither too fast nor too slow, improving the therapeutic effect of the thermosensitive gel preparation. After the thermosensitive gel composition is used, it can slowly release NO gas, and the release duration can last for more than 12 hours, showing significant efficacy in long - term antibacterial. Detailed implementation manners

[0054] Unless otherwise defined herein, scientific and technical terms used in conjunction with the present invention shall have the meanings commonly understood by those of ordinary skill in the art. The meaning and scope of the terms should be clear. However, in any case of potential ambiguity, the definitions provided herein shall prevail over any dictionary or extrinsic definition. In this application, unless otherwise specified, the use of "or" means "and / or". In addition, the use of the term "comprising" and other forms is non - restrictive.

[0055] The technical solutions of the present invention will be described clearly and completely below in conjunction with the embodiments. Obviously, the described embodiments are some, but not all, of the embodiments of the present invention. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0056] In a first aspect, the present invention provides a thermosensitive gel composition for slow - releasing nitric oxide, comprising the following components: a thermosensitive gel matrix, a NO donor composition, and optionally an acid - base regulator and an oxidant; wherein, the components of the thermosensitive gel matrix include: a thermosensitive matrix, a phase - change temperature regulator, and a solvent.

[0057] The thermosensitive gel composition for slow-release of nitric oxide provided by the present invention contains a phase-transition temperature regulator in the thermosensitive gel matrix of its components, which can adjust the phase-transition temperature of the thermosensitive gel and also adjust the initial viscosity and consistency of the gel. It can serve as a space chamber for gas storage. The phase-transition temperature regulator cooperates with the thermosensitive matrix to increase the gel viscosity, and by adjusting the usage amount of the phase-transition temperature regulator, the release rate of NO gas is neither too fast nor too slow, improving the therapeutic effect of the thermosensitive gel preparation. After the thermosensitive gel composition is used, it can slowly release NO gas, and the release duration can last for more than 12 hours, showing significant efficacy in long-term bacteriostasis.

[0058] Preferably, the solvent includes water.

[0059] In some preferred embodiments, the thermosensitive gel composition for slow-release of nitric oxide comprises the following components by weight:

[0060] 0.1 - 20 parts of NO donor composition and thermosensitive gel matrix, wherein the thermosensitive gel matrix comprises 2 - 35 parts of thermosensitive matrix, 0.5 - 5 parts of phase-transition temperature regulator and 35 - 98 parts of water.

[0061] In the thermosensitive gel composition for slow-release of nitric oxide, the addition amount of the NO donor composition is 0.1 - 20 parts, for example, it can be 0.1 part, 1 part, 5 parts, 15 parts, 20 parts, etc.;

[0062] In the thermosensitive gel composition for slow-release of nitric oxide, the addition amount of the thermosensitive matrix is 2 - 35 parts, for example, it can be 2 parts, 10 parts, 15 parts, 20 parts, 30 parts, 35 parts, etc.;

[0063] In the thermosensitive gel composition for slow-release of nitric oxide, the addition amount of the phase-transition temperature regulator is 0.5 - 5 parts, for example, it can be 0.5 part, 1 part, 2 parts, 3 parts, 4 parts, 5 parts, etc.

[0064] In the thermosensitive gel composition for slow-release of nitric oxide, the addition amount of water is 35 - 98 parts, for example, it can be 35 parts, 40 parts, 50 parts, 60 parts, 75 parts, 80 parts, 98 parts, etc.

[0065] In some preferred embodiments, the thermosensitive matrix comprises at least one of chitosan, cellulose, poloxamer, poly(N-isopropylacrylamide) (PNIPAM) and PEG polyester block copolymer;

[0066] Preferably, the thermosensitive matrix comprises poloxamer. Or, preferably, the matrix comprises chitosan.

[0067] Further preferably, the temperature-sensitive matrix comprises a combination of poloxamer 407 and poloxamer 188. Alternatively, further preferably, the temperature-sensitive matrix comprises a combination of chitosan and glycerophosphate or its salt.

[0068] In some preferred embodiments, when the temperature-sensitive matrix comprises poloxamer 407 and poloxamer 188, the weight ratio of poloxamer 407 to poloxamer 188 is 4-80:1, for example, it can be 4:1, 5:1, 10:1, 20:1, 30:1, 40:1, 50:1, 60:1, 70:1, 80:1, etc.

[0069] In the present invention, among the poloxamer family, the most commonly used temperature-sensitive gel materials are poloxamer 407 and poloxamer 188. Poloxamer 407 is the main gel matrix, which has gelling property within a suitable addition range, and the gelling temperature gradually decreases with the increase of its content. When the content is too low, it is difficult to form a gel even when the temperature rises. When the content is high, it can form a gel at a lower temperature, but the viscosity is relatively large, which is not conducive to gel preparation and large-scale production. Adding an appropriate amount of poloxamer 188 to poloxamer 407 can increase the gelling temperature. The greater the addition amount of poloxamer 188, the higher the gelling temperature. When the addition amount is too large, the gelling temperature is relatively high, and it is difficult to ensure that the gel is formed at room temperature or body temperature. An appropriate ratio of P407 to P188 can make the gel matrix have a gelling temperature matching the human body temperature range at an appropriate viscosity.

[0070] Optionally, when the temperature-sensitive matrix comprises glycerophosphate or its salt and chitosan, the weight ratio of glycerophosphate or its salt to chitosan is 1-5:1, for example, it can be 1:1, 2:1, 3:1, 4:1, 5:1, etc.

[0071] When the concentration of chitosan is too high, the solution will become viscous, lose fluidity, and form a jelly-like state, resulting in difficulty in spreading and operation during application. A high concentration of chitosan may lead to a decrease in the drug release rate because the high viscosity of chitosan will limit the diffusion and release of the drug. A low-concentration chitosan solution shows good fluidity in rheological tests, but may not form the required gel state in some applications. When the concentration of chitosan is too low, it may lead to insufficient gel strength and unable to provide the required mechanical support and stability.

[0072] Preferably, the phase change temperature regulator comprises at least one of carbomer, acrylic acid polymers and their sodium salts, sodium carboxymethyl cellulose, sodium hyaluronate, sodium alginate, calcium alginate, and alginates.

[0073] In the present invention, the phase change temperature regulator mainly functions to adjust the phase change temperature of the thermosensitive gel and simultaneously adjust the initial viscosity and consistency of the gel. It can serve as a spatial chamber for gas storage. If the addition amount is too large, the phase change temperature will decrease, the initial viscosity of the system will be too high, which is not conducive to use, and it will also cause the NO gas overflow rate to be slow and fail to achieve the therapeutic effect. If the addition amount is too small, it will not play a role in adjusting the phase change temperature, and the initial viscosity of the system is too thin to adhere to the damaged skin or mucous membrane, and it will also cause the NO release gas rate to be too fast and thus fail to achieve the ideal use effect.

[0074] Optionally, when the phase change temperature regulator includes carbomer and sodium alginate, the weight ratio of the carbomer to the sodium alginate is 1 to 10:1, for example, it can be 1:1, 5:1, 10:1, etc.

[0075] Optionally, when the phase change temperature regulator includes carbomer, sodium alginate and sodium carboxymethylcellulose, the weight ratio of the carbomer, the sodium alginate to the sodium carboxymethylcellulose is 0.5 to 50:0.5 to 5:1.

[0076] Among them, "0.5 to 50" can be, for example, 0.5, 1, 2, 10, 20, 30, 40, 50, etc.

[0077] Among them, "0.5 to 5" can be, for example, 0.5, 1, 2, 3, 4, 5, etc.

[0078] Preferably, the weight ratio of the thermosensitive matrix to the phase change temperature regulator is 0.5 to 70:1, for example, it can be 0.5:1, 1:1, 1.5:1, 2:1, 10:1, 20:1, 30:1, 40:1, 50:1, 60:1, 70:1, etc.

[0079] In the present invention, the appropriate addition of sodium alginate, carbomer, and sodium carboxymethylcellulose within a certain range can reduce the phase change temperature of the matrix and simultaneously increase its gel viscosity. This improved mechanical property may be due to the enhanced bonding strength of the polymer chains by the multiple interactions between poloxamer and polysaccharide molecules. The temperature regulation range of simply adding one of the phase change temperature regulators is limited, while when two or more are mixed and added, the temperature regulation range can be significantly increased and is greater than the sum of the temperature drops of the two. In addition, the cellulose-based thermosensitive hydrogel system also reduces the protein adsorption ability and enhances the cell adhesion ability, which is beneficial for the gel matrix to be used as a better delivery carrier for cell proliferation in damaged tissues.

[0080] In some preferred embodiments, the NO donor composition includes at least one of an inorganic nitrite donor, an arginine donor, an azodiglycolene salt donor, and a nitrosothiol.

[0081] In the present invention, the main function of the NO donor composition is to release NO and inhibit the growth of pathogenic bacteria. At the same time, it can also promote vasodilation, improve blood circulation, enhance blood flow, and promote tissue repair. If the addition amount is too high, the acidity or alkalinity of the system is too strong, which does not meet the physiological pH conditions, affects the NO release rate, and also affects the phase transition temperature, viscosity and other properties of the gel matrix. If the addition amount is too small, the release amount does not meet the standard and the beneficial effects cannot be achieved.

[0082] Preferably, the inorganic nitrite donor includes at least one of lithium nitrite, sodium nitrite, potassium nitrite, cesium nitrite, rubidium nitrite, silver nitrite, magnesium nitrite, calcium nitrite, manganese nitrite and barium nitrite; the inorganic nitrite donor is further preferably sodium nitrite and / or calcium nitrite.

[0083] Preferably, the arginine donor includes at least one of arginine and its derivatives; the arginine donor is preferably L-arginine or L-arginine hydrochloride.

[0084] Preferably, the azodolene salt donor includes at least one of piperazine, pyrrole, alkylamine or piperidine-derived azodolene salt, 2-(N,N-diethylamino) azodolate, azodiene containing a glycol group salt and azodolene containing an O-N=N-O group salt;

[0085] Preferably, the S-nitrosothiols include thiol substances containing an -SNO group, and the thiol substances containing an -SNO group include at least one of S-nitroso-N-acetylpenicillamine and S-nitrosoglutathione.

[0086] In some preferred embodiments, the thermosensitive gel composition for sustained release of nitric oxide further comprises 0.01 to 5 parts of an acid-base regulator, for example, it can be 0.01 part, 1 part, 2 parts, 3 parts, 4 parts, 5 parts.

[0087] In the present invention, the main functions of the acid-base regulator include providing reaction conditions for the donor and adjusting the pH of the gel preparation within the pH range suitable for the human body. If the addition amount is too high, a large amount of NO will be released rapidly in the early stage, and the duration will be short. It will also cause the system pH to be too large or too small, and at the same time, the gel cannot gel at human body temperature. If the addition amount is too small, the NO release amount is small and the release is slow.

[0088] Preferably, the acid-base regulator includes a substance that can ionize H + or OH - in an aqueous solution;

[0089] Preferably, the acid-base regulator includes at least one of ascorbic acid, citric acid, acetic acid, lactic acid, nicotinic acid, malic acid, tartaric acid, fumaric acid, mandelic acid, α-hydroxypropionic acid, β-hydroxypropionic acid, β-hydroxybutyric acid, gluconic acid, alginic acid, sodium ethylenediaminetetraacetate, sodium hydroxide, triethanolamine, sodium bicarbonate, arginine, and disodium hydrogen phosphate. The acid-base regulator is further preferably citric acid or sodium hydroxide.

[0090] In some preferred embodiments, the components of the thermosensitive gel composition for sustained release of nitric oxide include: 0.1 to 10 parts of NO donor composition, 0.1 to 2 parts of acid-base regulator, 2 to 25 parts of thermosensitive matrix, 0.5 to 5 parts of phase transition temperature regulator, and 50 to 98 parts of water. Among them, the thermosensitive matrix is preferably a composition of poloxamer 407 and poloxamer 188 or a composition of chitosan and glycerophosphate, and the phase transition temperature regulator is preferably carbomer, sodium alginate, and sodium carboxymethylcellulose;

[0091] In the thermosensitive gel composition for sustained release of nitric oxide, the addition amount of the NO donor composition is 0.1 to 10 parts, for example, it can be 0.1 part, 1 part, 5 parts, 10 parts, etc.;

[0092] In the thermosensitive gel composition for sustained release of nitric oxide, the addition amount of the acid-base regulator is 0.1 to 2 parts, for example, it can be 0.1 part, 1 part, 1.5 parts, 2 parts, etc.;

[0093] In the thermosensitive gel composition for sustained release of nitric oxide, the addition amount of the thermosensitive matrix is 2 to 25 parts, for example, it can be 2 parts, 5 parts, 10 parts, 15 parts, 20 parts, 25 parts, etc.;

[0094] In the thermosensitive gel composition for sustained release of nitric oxide, the addition amount of the phase transition temperature regulator is 0.5 to 5 parts, for example, it can be 0.5 part, 1 part, 2 parts, 3 parts, 4 parts, 5 parts, etc.;

[0095] In the thermosensitive gel composition for sustained release of nitric oxide, the addition amount of the water is 50 to 98 parts, for example, it can be 50 parts, 60 parts, 70 parts, 80 parts, 90 parts, 98 parts, etc.

[0096] Optionally, when the NO donor composition is nitrosothiols, its components are 0.1 to 5 parts, for example, it can be 0.1 part, 1 part, 2 parts, 3 parts, 4 parts, 5 parts, etc.

[0097] Furthermore, the NO donor composition is GSNO;

[0098] Optionally, the components of the thermosensitive gel composition for slow-release of nitric oxide include: 2 parts of S-nitrosoglutathione (GSNO), 1 part of sodium hydroxide, 20 parts of thermosensitive matrix, 2 parts of phase transition temperature regulator, and 95 parts of water. Among them, the thermosensitive matrix includes poloxamer 407 and poloxamer 188, and the phase transition temperature regulator includes 1 part of carbomer and 1 part of sodium alginate. The gelation temperature of this gel preparation is 28 °C.

[0099] In some preferred embodiments, the thermosensitive gel composition for slow-release of nitric oxide further includes 0.0001 to 15 parts of an oxidant, such as 0.0001 part, 0.001 part, 0.01 part, 0.1 part, 1 part, 5 parts, 10 parts, 15 parts, etc.;

[0100] Preferably, the oxidant includes at least one of ozone, potassium permanganate, potassium perchlorate, hydrogen peroxide, sodium peroxide, carbamide peroxide, sodium percarbonate, tert-butyl hydroperoxide, peracetic acid, hypochlorous acid, and inorganic nitrite; the oxidant is preferably hydrogen peroxide.

[0101] Optionally, when the NO donor composition includes sodium nitrite, the weight ratio of sodium nitrite to the L-arginine donor (as an acid-base regulator) is 0.25 to 4:1, such as 0.25:1, 1:1, 2:1, 3:1, 4:1, etc.

[0102] In the present invention, L-arginine itself is an endogenous nitric oxide donor, which can release nitric oxide under the catalysis of NOS. At the same time, since L-arginine is a basic amino acid, its addition can play the role of a basic regulator and adjust the pH of the composition towards the alkaline direction. Therefore, when L-arginine is present in the composition, it can not only serve as a NO donor but also replace or partially replace the addition of the basic regulator in the composition. In the present invention, nitrite and arginine act synergistically to inhibit the growth of pathogenic bacteria and have a significant antibacterial effect. It can release nitric oxide for up to more than 3 days under physiological environment (pH, temperature) conditions.

[0103] Optionally, the components of the thermosensitive gel composition for slow-release of nitric oxide include: 1 part of NO donor composition, 0.0004 part of hydrogen peroxide, 10 parts of thermosensitive matrix, 3.5 parts of phase transition temperature regulator, and 95 parts of water. Among them, the NO donor composition includes 0.7 part of sodium nitrite and 0.3 part of L-arginine donor, the thermosensitive matrix includes chitosan and glycerophosphate, and the phase transition temperature regulator includes 2 parts of carbomer, 1 part of sodium alginate, and 0.5 part of sodium carboxymethylcellulose. The gelation temperature of this gel preparation is 32 °C.

[0104] Since the incorporation of conventional nitric oxide donors and acid-base regulators into thermosensitive gels can affect the phase transition temperature or viscosity of the thermosensitive gels due to reasons such as pH changes and ionic effects, thereby affecting the gel adhesion effect and usage effect; or the gel matrix components can affect the release rhythm of the NO donor due to reasons such as viscosity and pH. After the addition of the NO donor and acid-base regulator, it will affect the properties such as the phase transition temperature and viscosity of the original gel matrix, resulting in incompatibility with the human application scenario. In order to appropriately eliminate the influence while ensuring the release rhythm of NO, it is necessary to make adaptive designs and combinations of the types and ratios of the gel matrix, NO donor, and acid-base regulator to obtain a gel preparation that meets the human application scenario. When the nitric oxide release composition is incorporated into the thermosensitive gel in the present invention, it is not a product with all suitable properties obtained through conventional formulation or experiments, but through ingenious design and the combination of specific components, a product that meets the human application scenario is obtained.

[0105] By adopting the specific combination of several specific components, the present invention can minimize the influence of the NO donor on the gel matrix / the influence of the gel matrix on NO release, and then obtain a gel preparation with suitable viscosity, phase transition temperature, pH, and NO release rate. The raw materials used in the product composition of the present invention are all directly purchasable, without the need for operations such as molecular-level synthesis of the raw materials, and have more cost advantages.

[0106] The second aspect of the present invention provides a preparation method of the thermosensitive gel composition for slow-release nitric oxide, comprising the following steps:

[0107] Mix the formula amount of thermosensitive gel matrix, NO donor composition, and optionally acid-base regulator and oxidant to prepare the thermosensitive gel composition for slow-release nitric oxide.

[0108] In some preferred embodiments, the preparation method of the thermosensitive gel composition for slow-release nitric oxide comprises the following steps:

[0109] Add the phase transition temperature regulator and thermosensitive matrix to the NO donor composition solution in sequence, then add the acid-base regulator or oxidant, and finally perform filling;

[0110] Preferably, add the phase transition temperature regulator at a temperature of 50 - 80 °C and dissolve it fully under the condition of a rotation speed of 30 - 400 rpm;

[0111] Among them, "50 - 80 °C" can be, for example, 50 °C, 60 °C, 70 °C, 80 °C, etc.;

[0112] Among them, "30 - 400 rpm" can be, for example, 30 rpm, 50 rpm, 100 rpm, 150 rpm, 200 rpm, 250 rpm, 300 rpm, 350 rpm, 400 rpm, etc.

[0113] Preferably, the thermosensitive matrix is added under stirring conditions, and then placed at a temperature of 0 - 4 °C until the thermosensitive matrix dissolves;

[0114] Among them, "0 - 4 °C" can be, for example, 0 °C, 1 °C, 2 °C, 3 °C, 4 °C, etc.

[0115] Preferably, an oxidant or an acid-base regulator is added under stirring conditions, the stirring speed is 10 - 20 rpm, and the stirring time is 2 - 5 min.

[0116] Among them, "10 - 20 rpm" can be, for example, 10 rpm, 15 rpm, 20 rpm, etc.;

[0117] Among them, "2 - 5 min" can be, for example, 2 min, 3 min, 4 min, 5 min, etc.

[0118] In some preferred embodiments, when the NO donor composition includes sodium nitrite and arginine, the preparation method of the thermosensitive gel composition for sustained release of nitric oxide includes the following steps: sequentially adding a phase transition temperature regulator, a thermosensitive matrix, and arginine to a sodium nitrite solution, then adding an oxidant or an acid-base regulator, and finally filling.

[0119] Preferably, the addition process of arginine includes: under stirring conditions, adding the arginine solution in batches within 2 - 5 min, the stirring speed is 10 - 20 rpm, and the stirring time is 10 - 30 min;

[0120] Among them, "2 - 5 min" can be, for example, 2 min, 3 min, 4 min, 5 min, etc.;

[0121] Among them, "10 - 20 rpm" can be, for example, 10 rpm, 15 rpm, 20 rpm, etc.;

[0122] Among them, "10 - 30 min" can be, for example, 10 min, 20 min, 30 min, etc.

[0123] In an alternative embodiment of the present invention, more preferably, the preparation method of the thermosensitive gel composition for sustained release of nitric oxide (mainly for the preparation method of a single-phase gel preparation of sodium nitrite) specifically includes the following steps:

[0124] (1) Dissolution of NO donor: Accurately weigh sodium nitrite in the donor composition according to the corresponding formula amount and dissolve it in water;

[0125] (2) Dissolution of thermosensitive gel matrix: In the solution of (1), weigh and add phase change temperature regulators such as sodium alginate, sodium carboxymethylcellulose, and carbomer according to the corresponding formulation amounts, and dissolve them thoroughly under the conditions of a temperature of 50 - 80°C and a rotation speed of 30 - 400 rpm; then continuously stir and add poloxamer on this basis, and place it under low temperature conditions (0 - 4°C) until it is completely dissolved;

[0126] (3) Incorporation of arginine: Weigh arginine, dissolve it in water, and add it to (2) in batches (within 2 - 5 minutes) at a rotation speed of 10 - 20 rpm, and maintain the stirring time for 10 - 30 minutes;

[0127] (4) Incorporation of hydrogen peroxide: Weigh hydrogen peroxide and add it to (3) at a rotation speed of 10 - 20 rpm, and maintain the stirring time for 2 - 5 minutes.

[0128] (5) Transfer it to a gel filling machine for filling.

[0129] In the present invention, the dissolution of sodium alginate and sodium carboxymethylcellulose requires a certain temperature and shear force. If the temperature is too low and the stirring is too slow, it will lead to slow dissolution, aggregation, and even inability to dissolve; if the temperature is too high and the stirring is too fast, it will result in high energy consumption, molecular chain breakage, and reduced viscosity. The most commonly used preparation method for poloxamer is the cold method, that is, it is dissolved at a lower temperature and the solution is clear after dissolution. At a higher temperature, its viscosity is relatively large, which is not conducive to its complete dissolution. At the same time, the phase change temperature regulator needs to be dissolved first and then added to poloxamer. Otherwise, after poloxamer is completely dissolved, its viscosity is relatively large, and it is difficult for the phase change temperature regulator to continue to dissolve under normal temperature conditions. Arginine and sodium hydroxide are added to the above gel in batches, which can avoid excessive local viscosity, affect the uniformity of product quality, and also avoid premature reaction with the NO donor.

[0130] The third aspect of the present invention provides an application of the thermosensitive gel composition for slow - releasing nitric oxide or the preparation method as described above in the preparation of thermosensitive gel preparations.

[0131] The thermosensitive gel preparation provided by the present invention is a thermosensitive gel preparation for slow - releasing nitric oxide, which can slowly release NO under physiological pH and temperature conditions. The thermosensitive gel preparation includes a single - phase or two - phase system composed of a nitric oxide donor composition, a catalyst, a thermosensitive gel matrix, a phase change temperature regulator, and water.

[0132] After the thermosensitive gel preparation is used, it can slowly release NO gas, and the release duration can last for more than 12 hours, showing significant efficacy in long-term antibacterial. The thermosensitive gel matrix rich in nitric oxide will transform into a solid-like state at body temperature and adhere to the affected area, slowly releasing nitric oxide while forming a gel protective film barrier to prevent external pathogenic bacteria from entering, so that the wound or mucosa always maintains a microenvironment of nitric oxide at a certain concentration. The present invention also innovatively adds a phase change temperature regulator that is neither an inorganic salt nor an alcohol to regulate the thermosensitive performance of the gel, and at the same time adjusts the viscosity of the thermosensitive solution before gelation, making it easy to apply and use.

[0133] The thermosensitive gel preparation provided by the present invention has the following beneficial effects:

[0134] 1. Long-term slow release: After the gel preparation is used, it can slowly release NO gas, and the release duration can last for 12 hours and above, showing significant efficacy in long-term antibacterial;

[0135] 2. Temperature responsiveness: The gel preparation remains fluid at room temperature, facilitating application and use, while transforming into a semi-solid gel state at body temperature, enhancing the adhesion of the drug to the damaged skin or mucosa;

[0136] 3. NO donor versatility; The gel matrix can adapt to the nitric oxide release system, without pretreatment or activation, and rapid gelation at a temperature close to human body temperature makes the NO release rate more stable, with a long release time, and also avoids problems such as allergies and pain caused by excessive irritation due to high-concentration burst release;

[0137] 4. A phase change temperature regulator that is neither an inorganic salt nor an alcohol is added to the formula to regulate the thermosensitive performance of the gel, and at the same time adjusts the viscosity of the thermosensitive solution before gelation, making it easy to apply and use;

[0138] 5. Improve drug distribution: Since traditional gel preparations are unevenly distributed on mucous membranes or wounds, while the low-viscosity liquid state of the thermosensitive gel is conducive to the spread of the drug on mucous membranes or uneven wounds, solving the problem of uneven drug administration;

[0139] 6. Prolong the drug retention time: The thermosensitive gel quickly turns into a semi-solid and adheres to the mucous membrane, wound, and skin after reaching the gelation temperature, increasing the drug retention time and achieving the purpose of slow drug release.

[0140] The present invention will be further described below through examples. Unless otherwise specified, the materials in the examples are prepared according to existing methods or directly purchased from the market.

[0141] Example 1

[0142] This example provides a temperature-sensitive gel composition for slow-release of nitric oxide, which comprises the following components by weight: 2 parts of NO donor composition (GSNO), 1 part of acid-base regulator (sodium hydroxide), 20 parts of temperature-sensitive matrix (poloxamer 407 + poloxamer 188) (15 parts of poloxamer 407 and 5 parts of poloxamer 188), 2 parts of phase transition temperature regulator (carbomer + sodium alginate) (1 part of carbomer and 1 part of sodium alginate each), and 95 parts of water.

[0143] Example 2

[0144] This example provides a temperature-sensitive gel composition for slow-release of nitric oxide, which comprises the following components by weight: 5 parts of NO donor composition (GSNO), 2 parts of acid-base regulator (sodium hydroxide), 25 parts of temperature-sensitive matrix (poloxamer 407 + poloxamer 188) (20 parts of poloxamer 407 and 5 parts of poloxamer 188), 5 parts of phase transition temperature regulator (carbomer + sodium alginate) (4 parts of carbomer and 1 part of sodium alginate), and 98 parts of water.

[0145] Example 3

[0146] This example provides a temperature-sensitive gel composition for slow-release of nitric oxide, which comprises the following components by weight: 0.1 part of NO donor composition (GSNO), 0.1 part of acid-base regulator (sodium hydroxide), 20 parts of temperature-sensitive matrix (poloxamer 407 + poloxamer 188) (18 parts of poloxamer 407 and 2 parts of poloxamer 188), 0.5 part of phase transition temperature regulator (carbomer + sodium alginate) (0.45 part of carbomer and 0.05 part of sodium alginate), and 95 parts of water.

[0147] Example 4

[0148] This example provides a temperature-sensitive gel composition for slow-release of nitric oxide, which comprises the following components by weight: 5 parts of NO donor composition (GSNO), 5 parts of acid-base regulator (sodium hydroxide), 35 parts of temperature-sensitive matrix (poloxamer 407 + poloxamer 188) (30 parts of poloxamer 407 and 5 parts of poloxamer 188), 2 parts of phase transition temperature regulator (carbomer + sodium alginate) (1 part each), and 98 parts of water.

[0149] Example 5

[0150] This example provides a temperature-sensitive gel composition for slow-release of nitric oxide, which comprises the following components by weight: 5 parts of NO donor composition (GSNO), 2 parts of acid-base regulator (sodium hydroxide), 25 parts of temperature-sensitive matrix (poloxamer 407 + poloxamer 188) (20 parts of poloxamer 407 and 5 parts of poloxamer 188), 2 parts of phase transition temperature regulator (carbomer + sodium alginate) (1 part each), and 80 parts of water.

[0151] Example 6

[0152] This example provides a thermosensitive gel composition for slow-release of nitric oxide, which comprises the following components by weight:

[0153] 1 part of NO donor composition (sodium nitrite + L-arginine donor) (0.7 part of sodium nitrite, 0.3 part of L-arginine donor);

[0154] 0.0004 part of oxidant (hydrogen peroxide);

[0155] 10 parts of matrix (chitosan + glycerophosphate) (2 parts of chitosan, 8 parts of glycerophosphate);

[0156] 3.5 parts of phase transition temperature regulator (carbomer + sodium alginate + sodium carboxymethylcellulose) (2 parts of carbomer, 1 part of sodium alginate, 0.5 part of sodium carboxymethylcellulose);

[0157] 95 parts of water.

[0158] Example 7

[0159] This example provides a thermosensitive gel composition for slow-release of nitric oxide, which comprises the following components by weight:

[0160] 1 part of NO donor composition (sodium nitrite + L-arginine donor) (0.2 part of sodium nitrite, 0.8 part of L-arginine donor);

[0161] 0.0001 part of oxidant (hydrogen peroxide);

[0162] 10 parts of matrix (chitosan + glycerophosphate) (5 parts of chitosan, 5 parts of glycerophosphate);

[0163] 3.5 parts of phase transition temperature regulator (carbomer + sodium alginate + sodium carboxymethylcellulose) (3.395 parts of carbomer, 0.035 part of sodium alginate, 0.07 part of sodium carboxymethylcellulose);

[0164] 95 parts of water.

[0165] Example 8

[0166] This example provides a thermosensitive gel composition for slow-release of nitric oxide, which comprises the following components by weight:

[0167] 1 part of NO donor composition (sodium nitrite + L-arginine donor) (0.8 part of sodium nitrite, 0.2 part of L-arginine donor);

[0168] 15 parts of oxidant (hydrogen peroxide);

[0169] Matrix (chitosan + glycerophosphate) 10 parts (chitosan 1.7 parts, glycerophosphate 8.3 parts);

[0170] Phase transition temperature regulator (carbomer + sodium alginate + sodium carboxymethylcellulose) 3.5 parts (carbomer 0.5 part, sodium alginate 2.5 parts, sodium carboxymethylcellulose 0.5 part);

[0171] Water 95 parts.

[0172] Example 9

[0173] This example provides a thermosensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is that the thermosensitive matrix uses poloxamer 407 and poloxamer 188. Poloxamer 407 is 19 parts and poloxamer 188 is 1 part, and the other components are the same as those in Example 6.

[0174] Example 10

[0175] This example provides a thermosensitive gel composition for slow-release of nitric oxide. The difference from Example 9 is that the thermosensitive matrix uses 15 parts of poloxamer 407 and 5 parts of poloxamer 188, and the weight ratio of the two is 3:1, and the other components are the same as those in Example 9.

[0176] Example 11

[0177] This example provides a thermosensitive gel composition for slow-release of nitric oxide. The difference from Example 9 is that the thermosensitive matrix uses 19.88 parts of poloxamer 407 and 0.12 part of poloxamer 188, and the weight ratio of the two is approximately 166:1, and the other components are the same as those in Example 9.

[0178] Example 12

[0179] This example provides a thermosensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is that in the thermosensitive matrix, glycerophosphate is 4 parts and chitosan is 6 parts, and the weight ratio of the two is approximately 0.67:1, and the other components are the same as those in Example 6.

[0180] Example 13

[0181] This example provides a thermosensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is that in the thermosensitive matrix, glycerophosphate is 8.5 parts and chitosan is 1.5 parts, and the weight ratio of the two is approximately 5.67:1, and the other components are the same as those in Example 6.

[0182] Example 14

[0183] This example provides a thermosensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is that:

[0184] Matrix (chitosan + glycerophosphate) 2.5 parts (chitosan 0.5 part, glycerophosphate 2 parts);

[0185] Phase transition temperature regulator (carbomer + sodium alginate + sodium carboxymethylcellulose) 5 parts (carbomer 2.9 parts, sodium alginate 1.4 parts, sodium carboxymethylcellulose 0.7 part);

[0186] The remaining components are the same as those in Example 6.

[0187] Example 15

[0188] This example provides a temperature-sensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is:

[0189] Matrix (chitosan + glycerophosphate) 35 parts (chitosan 7 parts, glycerophosphate 28 parts);

[0190] Phase transition temperature regulator (carbomer + sodium alginate + sodium carboxymethylcellulose) 0.5 part (carbomer 0.29 part, sodium alginate 0.14 part, sodium carboxymethylcellulose 0.07 part);

[0191] The remaining components are the same as those in Example 6.

[0192] Example 16

[0193] This example provides a temperature-sensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is:

[0194] Matrix (chitosan + glycerophosphate) 2 parts (chitosan 0.4 part, glycerophosphate 1.6 parts);

[0195] Phase transition temperature regulator (carbomer + sodium alginate + sodium carboxymethylcellulose) 5 parts (carbomer 2.9 parts, sodium alginate 1.4 parts, sodium carboxymethylcellulose 0.7 part);

[0196] The remaining components are the same as those in Example 6.

[0197] Example 17

[0198] This example provides a temperature-sensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is:

[0199] Matrix (chitosan + glycerophosphate) 35.5 parts (chitosan 7.1 parts, glycerophosphate 28.4 parts);

[0200] Phase transition temperature regulator (carbomer + sodium alginate + sodium carboxymethylcellulose) 0.5 part (carbomer 0.29 part, sodium alginate 0.14 part, sodium carboxymethylcellulose 0.07 part);

[0201] The remaining components are the same as those in Example 6.

[0202] Example 18

[0203] This example provides a thermosensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is: 0.1 part of sodium nitrite and 0.9 part of L-arginine donor;

[0204] The remaining components are the same as those in Example 6.

[0205] Example 19

[0206] This example provides a thermosensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is: 0.83 part of sodium nitrite and 0.17 part of L-arginine donor;

[0207] The remaining components are the same as those in Example 6.

[0208] Example 20

[0209] This example provides a thermosensitive gel composition for slow-release of nitric oxide. The difference from Example 6 is: the NO donor composition only contains 1 part of sodium nitrite, and the remaining components are the same as those in Example 6.

[0210] Examples 21 - 40

[0211] Examples 21 - 40 provide a preparation method of a thermosensitive gel composition for slow-release of nitric oxide, and respectively adopt the component formulations of the thermosensitive gel compositions in Examples 1 - 20, which specifically include the following steps:

[0212] (1) Dissolution of NO donor: Accurately weigh the NO donor composition according to the corresponding formulation amount, and dissolve it in water. If the NO donor composition includes a combination of sodium nitrite and L-arginine donor, first dissolve the sodium nitrite in the NO donor composition in water;

[0213] (2) Dissolution of gel matrix: In the solution of (1), weigh and add the phase transition temperature regulator according to the corresponding formulation amount, and make it fully dissolve at a temperature of 65°C and a rotation speed of 215 rpm; then continuously stir and add the thermosensitive matrix on this basis, and place it at a low temperature condition (0 - 4°C) until it is fully dissolved;

[0214] (3) If the NO donor composition includes a combination of sodium nitrite and L-arginine donor, then perform this step (3): Incorporation of arginine, weigh arginine, dissolve it in water, and add it to (2) in batches (within 2 - 5 minutes) at a rotation speed of 15 rpm, and keep stirring for 20 minutes; if the NO donor composition is only GSNO, then this step (3) is not required;

[0215] (4) Incorporation of oxidant hydrogen peroxide or acid-base regulator: When the components of the example contain the oxidant hydrogen peroxide, weigh the oxidant hydrogen peroxide and add it to the solution of the previous step, with a rotation speed of 15 rpm and a stirring time of 3.5 min maintained; alternatively, when the components of the example contain an acid-base regulator, weigh the acid-base regulator and add it to the solution of the previous step, with a rotation speed of 15 rpm and a stirring time of 3.5 min maintained.

[0216] (5) Transfer to a gel filling machine for filling.

[0217] The prepared gel product is stored in a sealed container.

[0218] Example 41

[0219] This example provides a preparation method of a thermosensitive gel composition for slow-release nitric oxide. Using the component formula of the thermosensitive gel composition in Example 9, it specifically includes the following steps:

[0220] (1) Dissolution of NO donor: Accurately weigh sodium nitrite in the donor composition according to the corresponding formula amount and dissolve it in water;

[0221] (2) Dissolution of gel matrix: In the solution of (1), weigh sodium alginate, sodium carboxymethylcellulose, carbomer and other phase transition temperature regulators according to the corresponding formula amount and add them. Make it fully dissolve under the conditions of a temperature of 50 °C and a rotation speed of 400 rpm; then continuously stir and add poloxamer on this basis, and place it at a low temperature condition (0 - 4 °C) until it is fully dissolved;

[0222] (3) Incorporation of arginine: Weigh arginine, dissolve it in water, and add it to (2) in batches (within 2 - 5 minutes), with a rotation speed of 10 rpm and a stirring time of 30 min maintained;

[0223] (4) Incorporation of hydrogen peroxide: Weigh hydrogen peroxide and add it to (3), with a rotation speed of 10 rpm and a stirring time of 5 min maintained.

[0224] (5) Transfer to a gel filling machine for filling.

[0225] The prepared gel product is stored in a sealed container.

[0226] Example 42

[0227] This example provides a preparation method of a thermosensitive gel composition for slow-release nitric oxide. Using the component formula of the thermosensitive gel composition in Example 9, it specifically includes the following steps:

[0228] (1) Dissolution of NO donor: Accurately weigh sodium nitrite in the donor composition according to the corresponding formula amount and dissolve it in water;

[0229] (2) Gel matrix dissolution: In the solution of (1), according to the corresponding formula amount, weigh and add phase change temperature regulators such as sodium alginate, sodium carboxymethylcellulose, carbomer, etc., and dissolve them fully at a temperature of 80 °C and a rotation speed of 30 rpm; then continuously stir and add poloxamer on this basis, and place it at a low temperature condition (0 - 4 °C) until it is fully dissolved;

[0230] (3) Arginine incorporation: Weigh arginine, dissolve it in water, and add it to (2) in batches (within 2 - 5 minutes) at a rotation speed of 20 rpm, and keep stirring for 10 minutes;

[0231] (4) Hydrogen peroxide incorporation: Weigh hydrogen peroxide and add it to (3) at a rotation speed of 20 rpm, and keep stirring for 2 minutes.

[0232] (5) Transfer to a gel filling machine for filling.

[0233] The prepared gel product is stored in a sealed container.

[0234] Comparative Example 1

[0235] This comparative example provides a thermosensitive gel composition, which is different from Example 6 in that it does not contain a phase change temperature regulator. The other components are the same as those in Example 6.

[0236] Comparative Example 2

[0237] This comparative example provides a preparation method of a thermosensitive gel composition. Using the component formulation of Comparative Example 1, the preparation steps are the same as those in Example 26.

[0238] Test Example

[0239] Test samples: The thermosensitive gel compositions prepared in Examples 21 - 42 and Comparative Example 2 are used as samples for detection. The products prepared in Examples 21 - 42 are samples 1 - sample 22 respectively.

[0240] (1) Performance test 1: NO release amount test

[0241] Test method 1 - Test with a Thremo tester: After the samples are stored in a sealed container and aged for 6 months, take samples and test the NO release amount with a Thremo tester.

[0242] Test method 2 - Detection with a kit (Griess Reagent method): After the samples are stored in a sealed container and aged for 6 months, take the samples out of the sealed container and store them in an open environment for 72 hours, and then detect them with a nitric oxide kit (Griess Reagent method).

[0243] Name Product Number Brand Nitric Oxide Detection Kit S0021S Beyotime

[0244] Packing List:

[0245] Product Number Product Name Package S0021S-1 <![CDATA[1M NaNO2]]> 1ml S0021S-2 Griess Reagent l 25ml S0021S-3 Griess Reagent ll 25ml

[0246] Instructions for Use:

[0247] 1. Take out Griess Reagent I and II and allow them to return to room temperature.

[0248] 2. Dilute the standard (1 - 100 μM) with the solution used for the sample to be tested.

[0249] 3. Add the standard and the sample to the 96-well plate at 50 μl / well.

[0250] 4. Add room-temperature Griess Reagent I to each well at 50 μl / well.

[0251] 5. Add room-temperature Griess Reagent II to each well at 50 μl / well.

[0252] 6. Measure the absorbance at 540 nm and prepare a standard curve.

[0253] 7. Calculate the concentration of nitric oxide in the sample according to the standard curve.

[0254] (2) Performance Test 2: Gelation Temperature Test

[0255] Test Method: Place the gel sample in a test tube, seal it, and put it in an environment with controllable temperature. As the temperature changes, invert the test tube regularly and observe the flow state of the gel. When the temperature is lower than a certain temperature, the gel is liquid and will flow after the test tube is inverted; when the temperature is higher than a certain temperature, the gel becomes solid and will not flow after the test tube is inverted; this temperature is the gelation phase transition temperature.

[0256] (3) Performance Test 3: pH Test

[0257] Test Method: Randomly select 5 g of the gel, place it in a conical flask, inject 50 ml of distilled water, seal it with a rubber stopper, put it in a constant temperature water bath at a constant temperature of (60 °C ± 2 °C), shake it 5 times, swell for 10 minutes, cool to room temperature, and measure its pH.

[0258] (4) Performance Test 4: Antibacterial Effect Test of the Gel against Escherichia coli, Staphylococcus aureus, and Candida albicans.

[0259] Test Method: After the sample is stored in a sealed container and aged for 6 months, conduct the antibacterial effect test according to GB 15979-2024 Hygiene Standards for Disposable Sanitary Products.

[0260] The test results are shown in Table 1.

[0261] Table 1

[0262]

[0263]

[0264] The gelation temperature and pH of Sample 6 are both suitable for the human body's use environment. At the same time, the antibacterial rate is as high as 99.9%, showing antibacterial effects. When used as a vaginal gel preparation, the gelation temperature of 36°C enables the product to be stored under the vast majority of room temperature environmental conditions, and rapidly gelate at the vaginal temperature after application, adhering and remaining at the application site in the vagina, and then releasing nitric oxide for long-term antibacterial effects. The gelation temperatures of Sample 7 and Sample 8 are lower than that of Sample 6, but the corresponding products are also suitable for most storage conditions. Gelation may occur in advance under some hot room temperature environmental conditions, and such environmental conditions have requirements for storage conditions (refrigeration, low-temperature storage, etc.).

[0265] The gelation temperature of Sample 9 is suitable for the human body's use environment. At the same time, the antibacterial rate is as high as 99.9%, showing antibacterial effects. The pH of Sample 9 is slightly acidic, suitable for the vaginal use environment. The gelation temperature of Sample 10 is relatively high. When used as a vaginal gel preparation, it is difficult to gelate after application, affecting the product effect.

[0266] The gelation temperatures of Samples 11, 12, and 13 are relatively low, and they are prone to gelation at room temperature, having relatively high requirements for the storage environment and needing low-temperature storage.

[0267] The gelation temperature of Sample 14 is slightly higher than that of Sample 6. The antibacterial rate of Sample 15 is not as good as that of Sample 6, and it does not have antibacterial properties but has bacteriostatic properties.

[0268] The gelation temperature of Sample 16 is relatively high. When used as a vaginal gel preparation, it is difficult to gelate after application, affecting the product effect. The gelation temperature of Sample 17 is relatively low, and it is prone to gelation at room temperature, having relatively high requirements for the storage environment and needing low-temperature storage.

[0269] The pH of Sample 18 is slightly alkaline. When used as a vaginal gel preparation, there may be irritation. The antibacterial rates of Samples 19 and 20 are not as good as those of Sample 6 and Sample 18, and they do not have antibacterial properties but have bacteriostatic properties.

[0270] Meanwhile, by observing the NO release rhythms of Samples 6, 18, 19, and 20, it was found that: after Sample 6 was taken out of the closed container, within 30 min, its NO release amount remained at 50-100 ppb, and 72 hours after being taken out of the closed container, its NO release amount was measured to be 162.5 μM; after Sample 18 was taken out of the closed container, within 30 min, its NO release amount remained at 20-35 ppb, and 72 hours after being taken out of the closed container, its NO release amount was measured to be 76.5 μM; after Sample 19 was taken out of the closed container, within 30 min, its NO release amount remained at 20-50 ppb, and 72 hours after being taken out of the closed container, its NO release amount was measured to be 16.9 μM; after Sample 20 was taken out of the closed container, within 30 min, its NO release amount remained at 10-20 ppb, and no NO release was detected 72 hours after being taken out of the closed container. Under suitable component ratios, especially the suitable ratio of sodium nitrite and arginine, the gel composition can have a relatively high NO release at the initial stage of use, and at the same time, after 72 hours, it can still maintain an NO release at an effective antibacterial dose.

[0271] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that they can still modify the technical solutions described in the foregoing embodiments, or perform equivalent replacements on some or all of the technical features; and these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A thermosensitive gel composition for slow release of nitric oxide, characterized in that, It comprises the following components: a thermosensitive gel matrix, an NO donor composition, and optionally an acid-base regulator and an oxidant; Among them, the components of the thermosensitive gel matrix include: a thermosensitive matrix, a phase transition temperature regulator, and a solvent.

2. The thermosensitive gel composition for slow-release nitric oxide according to claim 1, wherein The thermosensitive matrix includes at least one of chitosan, cellulose, poloxamer, poly(N-isopropylacrylamide), and PEG polyester block copolymer; Preferably, the phase transition temperature regulator includes at least one of carbomer, acrylic acid polymers and their sodium salts, sodium carboxymethyl cellulose, sodium hyaluronate, sodium alginate, calcium alginate, and alginate; Preferably, the NO donor composition includes at least one of inorganic nitrite donors, arginine donors, azodioxolium salts donors, and nitrosothiols; Preferably, the inorganic nitrite donor includes at least one of lithium nitrite, sodium nitrite, potassium nitrite, cesium nitrite, rubidium nitrite, silver nitrite, magnesium nitrite, calcium nitrite, manganese nitrite, and barium nitrite; Preferably, the arginine donor includes at least one of arginine and its derivatives; Preferably, the arginine includes at least one of L-arginine and L-arginine hydrochloride salts; Preferably, the azodioxolium salts donors include at least one of piperazine, pyrrole, alkylamine or piperidine-derived dioxolium salts, 2-(N,N-diethylamino) dioxolium salts, dioxolium salts containing a glycol group, and azodioxolium salts containing an O-N=N-O group; Preferably, the nitrosothiols include thiol substances containing an -SNO group, and the thiol substances containing an -SNO group include at least one of S-nitroso-N-acetylpenicillamine and S-nitrosoglutathione; Preferably, the acid-base regulator includes a substance that can ionize H + or OH - in an aqueous solution; Preferably, the acid-base regulator includes at least one of ascorbic acid, citric acid, acetic acid, lactic acid, nicotinic acid, malic acid, tartaric acid, fumaric mandelic acid, α-hydroxypropionic acid, β-hydroxypropionic acid, β-hydroxybutyric acid, gluconic acid, alginic acid, sodium ethylenediaminetetraacetate, sodium hydroxide, triethanolamine, sodium bicarbonate, arginine, and disodium hydrogen phosphate; Preferably, the oxidant includes at least one of ozone, potassium permanganate, potassium perchlorate, hydrogen peroxide, sodium peroxide, urea peroxide, sodium percarbonate, tert-butyl hydroperoxide, peracetic acid, hypochlorous acid, and inorganic nitrites; Preferably, the solvent includes water.

3. The thermosensitive gel composition for slow-release nitric oxide according to claim 2, wherein The thermosensitive gel composition for sustained release of nitric oxide comprises the following components by weight: 0.1 to 20 parts of an NO donor composition and a thermosensitive gel matrix, wherein the thermosensitive gel matrix includes 2 to 35 parts of a thermosensitive matrix, 0.5 to 5 parts of a phase transition temperature regulator, and 35 to 98 parts of water; Preferably, the weight ratio of the thermosensitive matrix to the phase transition temperature regulator is 0.5 to 70:

1.

4. The thermosensitive gel composition for slow-release nitric oxide according to claim 3, wherein The thermosensitive gel composition for sustained release of nitric oxide further includes 0.01 to 5 parts of an acid-base regulator; Optionally, the components of the thermosensitive gel composition for sustained release of nitric oxide include: 0.1 to 10 parts of an NO donor composition, 0.1 to 2 parts of an acid-base regulator, 2 to 25 parts of a thermosensitive matrix, 0.5 to 5 parts of a phase transition temperature regulator, and 50 to 98 parts of water; Optionally, when the NO donor composition is a nitrosothiol, its component is 0.1 to 5 parts; Optionally, the components of the thermosensitive gel composition for sustained release of nitric oxide include: 2 parts of S-nitrosoglutathione, 1 part of sodium hydroxide, 20 parts of thermosensitive matrix, 2 parts of phase transition temperature regulator, and 95 parts of water. Among them, the thermosensitive matrix includes poloxamer 407 and poloxamer 188, and the phase transition temperature regulator includes carbomer and sodium alginate.

5. The thermosensitive gel composition for slow-release nitric oxide according to claim 3, wherein The thermosensitive gel composition for sustained release of nitric oxide further includes 0.0001 to 15 parts of oxidant; Optionally, when the NO donor composition includes sodium nitrite, the weight ratio of sodium nitrite to the L-arginine donor is 0.25 to 4:1, where the L-arginine donor serves as an acid-base regulator; Optionally, the components of the thermosensitive gel composition for sustained release of nitric oxide include: 1 part of NO donor composition, 0.0004 parts of hydrogen peroxide, 10 parts of thermosensitive matrix, 3.5 parts of phase transition temperature regulator, and 95 parts of water. Among them, the NO donor composition includes 0.7 parts of sodium nitrite and 0.3 parts of L-arginine donor, the thermosensitive matrix includes chitosan and glycerophosphate, and the phase transition temperature regulator includes 2 parts of carbomer, 1 part of sodium alginate, and 0.5 parts of sodium carboxymethylcellulose.

6. The thermosensitive gel composition for slow-release nitric oxide according to claim 5, wherein When the thermosensitive matrix includes poloxamer 407 and poloxamer 188, the weight ratio of poloxamer 407 to poloxamer 188 is 4 to 80:1; Optionally, when the thermosensitive matrix includes glycerophosphate or its salt, and chitosan, the weight ratio of glycerophosphate or its salt to chitosan is 1 to 5:1; Optionally, when the phase transition temperature regulator includes carbomer and sodium alginate, the weight ratio of carbomer to sodium alginate is 1 to 10:1; Optionally, when the phase transition temperature regulator includes carbomer, sodium alginate, and sodium carboxymethylcellulose, the weight ratio of carbomer, sodium alginate, and sodium carboxymethylcellulose is 0.5 to 50:0.5 to 5:

1.

7. The preparation method of the thermosensitive gel composition for sustained release of nitric oxide according to any one of claims 1-6, characterized in that, Comprising the following steps: Mix the formulated amount of thermosensitive gel matrix, NO donor composition, and optionally acid-base regulator and oxidant to prepare the thermosensitive gel composition for sustained release of nitric oxide.

8. The preparation method according to claim 7, characterized in that, The preparation method of the thermosensitive gel composition for sustained release of nitric oxide comprises the following steps: Sequentially add the phase transition temperature regulator and the thermosensitive matrix to the NO donor composition solution, then add the acid-base regulator or oxidant, and finally perform filling; Preferably, add the phase transition temperature regulator at a temperature of 50 to 80 °C and dissolve it sufficiently under the condition of a rotation speed of 30 to 400 rpm; Preferably, add the thermosensitive matrix under stirring conditions, and then place it at a temperature of 0 to 4 °C until the thermosensitive matrix dissolves; Preferably, add the oxidant or acid-base regulator under stirring conditions, with a stirring speed of 10 - 20 rpm and a stirring time of 2 to 5 min.

9. The preparation method according to claim 8, characterized in that When the NO donor composition comprises sodium nitrite and arginine, the preparation method of the thermosensitive gel composition for sustained release of nitric oxide comprises the following steps: sequentially adding a phase transition temperature regulator, a thermosensitive matrix and arginine into a sodium nitrite solution, then adding an oxidant or an acid-base regulator, and finally performing filling; Preferably, the addition process of arginine comprises: under stirring conditions, adding an arginine solution in batches within 2 - 5 minutes, with the stirring speed being 10 - 20 rpm and the stirring time being 10 - 30 minutes.

10. Use of the thermosensitive gel composition for sustained release of nitric oxide according to claims 1 - 6 or the preparation method according to claims 7 - 9 in the preparation of a thermosensitive gel preparation.

Images