CN103381139A - In situ gel for astragalus polysaccharide injection and preparation method thereof - Google Patents

Abstract

The invention belongs to the field of pharmaceutical preparations for animals, and relates to an in-situ gel preparation for astragalus polysaccharide injection and a preparation method thereof. The invention uses water as a solvent, is made of traditional Chinese medicine astragalus extract-astragalus polysaccharide, in-situ gel matrix recognized in the field and other pharmacy necessary auxiliary materials, and clarifies the preparation method. The preparation of the present invention is a free-flowing liquid at room temperature, forms a solid or semi-solid gel after intramuscular injection or subcutaneous injection, has a long duration of action, and can be administered only once in a course of treatment. The preparation has the advantages of simple preparation, stable properties, convenient administration, definite curative effect, strong adaptability to animals and the like. The preparation of the invention has extensive pharmacological effects such as promoting antibody synthesis, enhancing body immunity, anti-virus, anti-tumor and anti-oxidation, and is suitable for preventing and treating animal diseases such as pigs, sheep, dogs, cats and chickens.

Description

In situ gel for injection of astragalus polysaccharide and preparation method thereof

technical field

The invention belongs to the field of pharmaceutical preparations for animals, and relates to an in-situ gel for astragalus polysaccharide injection with a suitable phase transition temperature and a preparation method thereof. the

technical background

Injectable in situ gel (injectable in situ gel) is to dissolve drugs and polymers in a suitable solvent, and inject them locally subcutaneously or intramuscularly. Under physiological conditions at the administration site, the polymer solidifies to form a semi-solid or solid drug storage. The library gel, and through the continuous degradation of the polymer, the drug is continuously released to achieve a sustained release and long-term effect. According to the gelation mechanism, it can be divided into temperature-sensitive gels, ion-sensitive gels, pH-sensitive gels, and light-sensitive gels. Temperature-sensitive in-situ gels are gelled by temperature changes, and can be more widely applied to many tissues, organs or cavities in the body, and the latter three require specific physiological conditions or external factors to trigger gelation. the

The temperature-sensitive in situ gel is a low-viscosity liquid before use (at ambient temperature), and gels when it absorbs heat at the physiological temperature of the body to form a soft, lubricated, transparent solid or semi-solid gel, mostly poloxamer matrix material. Poloxamer is an ABA block copolymer composed of polyoxyethylene (PEO) and polyoxypropylene (PPO). PEO is relatively hydrophilic and PPO is relatively lipophilic. At low temperature or low concentration, poloxamer exists in the form of monomer in aqueous solution; when the ambient temperature is higher than the critical micelle temperature and the poloxamer concentration reaches the critical micelle concentration, a hydrophobic PPO is formed as the core, and the hydrophilic Water-based PEO is spherical, rod-shaped or lamellar micelles with an outer shell; when the temperature rises to the phase transition temperature, the micelles are tightly packed to form a semi-solid gel. The gelation temperature decreases with the increase of the concentration of poloxamer 407 (P407) in the system, and increases with the increase of the concentration of poloxamer 188 (P188). Different gelation temperatures can be obtained by adjusting the ratio and concentration of P407 and P188. the

Injectable in situ gel not only overcomes the shortcomings of ordinary emulsions, liposomes, microspheres, etc., which are complicated to prepare, have large differences between batches, and are difficult to produce in large quantities, but also avoid the pain of traditional implants during surgical implantation. The injection-type temperature-sensitive in-situ gel has a simple preparation process, easy-to-control dosage, and good physiological compatibility, which can prolong the drug release cycle, reduce the frequency of administration, improve patient compliance, and improve economic benefits. the

Astragalus is a medicine for invigorating the middle and nourishing Qi, and Astragalus polysaccharide is its active component. It is a water-soluble heteropolysaccharide obtained by extracting, concentrating and purifying the dried root of Astragalus membranaceus or Astragalus membranaceus. It contains hexuronic acid, glucose, fructose, Components such as rhamnose, arabinose, galacturonic acid and glucuronic acid. Modern pharmacological studies have shown that astragalus polysaccharides have pharmacological effects such as enhancing the body's immunity, anti-virus, anti-tumor, anti-aging, anti-radiation, anti-stress, and anti-oxidation. the

Astragalus polysaccharides are used as immunomodulators, which can promote the development of animal thymus, spleen, lymph nodes, poultry supraluminal bursa and diffuse lymphoid tissue throughout the body; promote the proliferation of plasma cells in the spleen of animals, the proliferation and transformation of humoral immune lymphocytes, and the synthesis of antibodies. Cellular immunity level; improve the function of mononuclear macrophages, enhance their phagocytosis, increase the activity of natural killer cells, etc., thereby regulating the development of immune organs, specific immunity and non-specific immunity. the

Astragalus polysaccharides have the function of scavenging OH ^- and O ₂ ^- free radicals, increasing the activity of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and total antioxidant activity in immune organs to enhance animal immunity defense ability.

Astragalus polysaccharides can stimulate macrophages and T cells, induce a variety of cytokines, and promote the production of interleukins and endogenous interferons, so as to achieve the purpose of anti-virus. Veterinary clinical practice has proved that astragalus polysaccharides have a certain preventive effect on infectious bursal virus, Marek's virus, Newcastle disease virus, and infectious laryngotracheitis virus. the

Astragalus polysaccharides are widely used in livestock and poultry farming practice. For example, in the pig industry, astragalus polysaccharides are used as feed additives to inhibit harmful intestinal flora, reduce the incidence of diarrhea in piglets, enhance piglet immunity, improve the survival rate of weaned piglets, and promote piglet growth; intramuscular injection can enhance piglet immune function and significantly improve piglet immunity The antibody level of swine fever vaccine significantly increased the percentage of neutrophils in the peripheral blood of piglets and the transformation rate of lymphocytes. In pet clinics, appropriate amount of Huangmao polysaccharide can promote the transformation of canine lymphocytes and enhance the immune function of the body; dogs with parvovirus orally administered astragalus polysaccharide, the blood immunoglobulin (IgM, IgE), cyclic adenosine monophosphate (cAMP) content increased significantly, and Can induce interferon, enhance the body's disease resistance, and promote humoral immune function. Adding astragalus polysaccharide powder to dairy cow feed can significantly improve its lactation performance and reduce the incidence of recessive mastitis. the

At present, the dosage forms and specifications of astragalus polysaccharides approved for veterinary use are: ① Astragalus polysaccharide injection: 10ml: 0.2g; 10ml: 0.1g; 100ml: 1g; 50ml: 0.5g; 100ml: 5g (based on astragalus polysaccharide). ② Astragalus polysaccharide oral liquid: yellow to reddish brown liquid, each 200ml is equivalent to 300g of the original drug. ③Astragalus polysaccharide powder: each 1g contains astragalus polysaccharide not less than 450mg. the

The reported dosage forms of astragalus polysaccharide include its powder injection, liposome, microcapsule, etc., but no in situ gel dosage form for injection of astragalus polysaccharide has been reported. the

Astragalus polysaccharides have published patents related to dosage forms, including astragalus polysaccharide pellets, astragalus polysaccharide granules, astragalus polysaccharide liposome long-acting injection, compound astragalus polysaccharide injection for pigs, and compound astragalus polysaccharide injection for preventing and treating infectious hepatitis in dogs. See the report and published patent of astragalus polysaccharide in situ gel for injection. the

In a course of treatment, astragalus polysaccharides need to be injected multiple times, and animals need to be kept in place frequently, which is time-consuming and troublesome and can easily cause animal stress and aggravate the condition, and the obvious "peaks and valleys" of blood drug concentration are not conducive to the enhancement of the body's immunity by astragalus polysaccharides. Strength, anti-virus and anti-stress effects. When the astragalus polysaccharide is made into an injectable in situ gel, a single injection can maintain an effective and relatively stable blood concentration for a long time, and improve the prevention and treatment effect on diseases. Moreover, a single injection can reduce unfavorable factors such as animal stress, reduce the number of animal restraints, save prevention and control costs, and improve economic benefits. the

Contents of the invention

The purpose of the present invention is to provide an in-situ gel for injection of astragalus polysaccharides, aiming at the deficiency of astragalus polysaccharides in existing veterinary dosage forms. the

Another object of the present invention is to provide a method for preparing the astragalus polysaccharide in situ gel for injection. the

The purpose of the present invention can be achieved through the following technical solutions:

The in situ gel for injection of astragalus polysaccharide comprises the following components: 1.0-15.0 parts by weight of astragalus polysaccharide, 15.5-30.5 parts by weight of poloxamer 407, 0.5-10.5 parts by weight of poloxamer 188, 25.0-80.0 parts by weight of water, high 0.01-10.0 parts by weight of a molecular blocker, 0.01-2.0 parts by weight of a pH regulator, and 0.01-3.0 parts by weight of a preservative. the

The gel matrix is selected from any one or more of poloxamer 108, 124, 188, 237, 338, 407; preferably poloxamer 407 15.5-30.5 parts by weight, poloxamer 1880.5-10.5 parts by weight. the

Described macromolecule blocking agent is selected from polyvinyl alcohol, povidone, methyl cellulose, hydroxypropyl methyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, sodium carboxymethyl cellulose, Any one or more of carbomer, hyaluronic acid, xanthan gum, chitosan, carboxymethyl chitosan, sodium alginate, phospholipids, etc.; preferably povidone 1.0-10.0 parts by weight, or hydroxypropyl 0.02-0.8 parts by weight of methyl cellulose, or 0.05-0.8 parts by weight of sodium carboxymethyl cellulose. the

The pH regulator is selected from the group consisting of lactic acid, acetic acid, hydrochloric acid, citric acid and its sodium salt, tartaric acid and its sodium salt, sodium carbonate, sodium bicarbonate, dipotassium hydrogen phosphate and potassium dihydrogen phosphate, dihydrogen phosphate Any one or more of sodium and sodium dihydrogen phosphate, sodium hydroxide, monoethanolamine, etc.; preferably 0.01-0.5 parts by weight of sodium tartrate, or 0.01-0.5 parts by weight of sodium hydroxide. the

Described preservative is selected from parabens, chlorobutanol, benzyl alcohol, phenylethyl alcohol, chlorhexidine acetate, benzalkonium bromide, benzalkonium chloride, sodium benzoate, potassium sorbate, thimerosal, etc. One or more; preferably 0.2-0.5 parts by weight of sodium benzoate, or 0.5-1.2 parts by weight of benzyl alcohol, or 0.01-0.5 parts by weight of benzalkonium bromide. the

The preparation method of the in situ gel for injection of astragalus polysaccharide comprises the following steps: (a) taking an appropriate amount of astragalus polysaccharide raw material prepared by the method of water extraction and alcohol precipitation, and dissolving it with an appropriate amount of water for injection; (b) mixing the selected pH regulator and After the preservatives were dissolved with a small amount of water for injection, they were added to the above-mentioned astragalus polysaccharide solution; (c) the prescribed amount of gel matrix and the selected polymer blocker were sprinkled on the astragalus polysaccharide solution, and refrigerated at about 4°C for 24 hours to obtain Clear, lump-free, uniformly dispersed solution, filtered through a 0.45 μm or 0.22 μm microporous membrane; (d) dilute the solution obtained in (c) with water for injection, and stir evenly to obtain the product. the

The advantages of the present invention are:

The present invention utilizes the temperature-sensitive nature of the poloxamer aqueous solution and the combination of different types of poloxamers to prepare the astragalus polysaccharide in situ gel for injection with a suitable phase transition temperature, so that it exists in a liquid state at room temperature. When administered by injection or subcutaneous injection, a gel forms at the injection site. Compared with conventional astragalus polysaccharide injection, the prepared in-situ gel for astragalus polysaccharide injection releases slowly, has stable blood drug concentration, and greatly prolongs the duration of action; the polymer material used has good physiological compatibility and low irritation. The preparation process of the in-situ gel for astragalus polysaccharide injection is simple, the administration is convenient, the times of administration are reduced, and the effect of the astragalus polysaccharide can be better exerted.

Description of drawings

The accompanying drawing is the in vitro release curve of astragalus polysaccharide injection in situ gel. the

Detailed ways

Implementation example 1:

1% astragalus polysaccharide in situ gel for injection

prescription

Preparation method: (a) take an appropriate amount of astragalus polysaccharide raw material prepared by water extraction and alcohol precipitation method, and dissolve it with an appropriate amount of water for injection; (b) add sodium tartrate and sodium benzoate aqueous solution to the above-mentioned astragalus polysaccharide solution; (c) add the prescribed amount of condensed Sprinkle the gum base and sodium carboxymethyl cellulose on the water surface of the astragalus polysaccharide solution, refrigerate overnight at about 4°C to obtain a clear, lump-free, uniformly dispersed solution, and filter through a 0.45 μm or 0.22 μm microporous membrane; (d) Dilute (c) to volume with water for injection, stir evenly, and obtain. the

The prepared in situ gel was evaluated in vitro, including gelation temperature, gelation time, thermal reversibility, release rate, viscosity, pH value, etc. The measurement methods of each item are as follows, the measurement results are shown in Table 1, and the in vitro release results are shown in the accompanying drawings. the

Gel temperature measurement (test tube inversion method) Take 3-4ml of gel solution stored in the refrigerator into a test tube, and insert a thermometer into the gel solution. Place the test tube in a water bath (the liquid level of the water bath is 3cm higher than the gel solution in the test tube), and slowly raise the temperature at a rate of about 1°C every 1 to 2 minutes. Tilt the test tube by 90°, and observe the temperature at which the content does not flow, which is defined as the gelation temperature. Each sample was measured 3 times, and the average value was taken as the result. the

Determination of gelation time The in situ gel solution was placed at 25°C for 0.5h, placed in a test tube preheated to 37°C and kept warm, and the phase transition time was recorded. the

Thermal reversibility determination Heat the gel to a specific temperature (30, 35, 40, 45, 50, 55, 60, 70°C), and then slowly cool it to room temperature, which counts as one heating cycle, and check until the gel is no longer Thermosensitivity or composition changes, if repeated 10 times still has thermosensitivity, recorded as the number of cycles > 10. the

Determination of release rate Precisely draw 10ml of the in-situ gel preparation, place it in a pre-weighed flat-bottomed graduated test tube, and then weigh it. Place the test tube in a constant temperature water bath shaker at 37.3±0.2°C to balance for 10 minutes, so that the polymer solution can completely form a gel. Carefully add 5ml of buffer solution (pH7.4, containing 0.5% SDS, used as a release medium) preheated at 37°C, and shake in a constant temperature water bath (95 times/min). , 5, 8, 12, 24, 48, 60, 72, 84, 96h immediately pour out all the release medium, dry the inner and outer surfaces of the container with filter paper, quickly weigh and record, and then put it back into the constant temperature water bath shaker to balance After 10 minutes, add 5ml of release medium. Repeat this operation until the end of the test. the

Viscosity Measurement The viscosity of the gel at room temperature (25° C.) was measured using a rotary viscometer. the

pH Determination The pH value of the in situ gel solution at room temperature was measured with a pH meter. the

Implementation example 2:

2% astragalus polysaccharide in situ gel for injection

prescription

Preparation method: (a) take an appropriate amount of astragalus polysaccharide raw material prepared by water extraction and alcohol precipitation method, and dissolve it with an appropriate amount of water for injection; (b) add aqueous sodium benzoate solution to the above-mentioned astragalus polysaccharide solution; (c) mix prescription amount of gel matrix and polysaccharide Sprinkle vitamin D on the water surface of astragalus polysaccharide solution, refrigerate overnight at about 4°C to obtain a clear, lump-free, uniformly dispersed solution, and filter through a 0.45 μm or 0.22 μm microporous membrane; (d) mix (c) with water for injection Constant volume, stir evenly, that is. The in vitro performance evaluation method is the same as in Implementation 1, and the results are shown in Table 1, and the in vitro release results are shown in the accompanying drawings. the

Implementation example 3:

5% astragalus polysaccharide in situ gel for injection

prescription

Preparation method: (a) take an appropriate amount of astragalus polysaccharide raw material prepared by the method of water extraction and alcohol precipitation, and dissolve it with an appropriate amount of water for injection; (b) soak the carbomer with an appropriate amount of water for injection, and adjust it with benzalkonium bromide and sodium hydroxide aqueous solution. Dissolve and add to the above-mentioned astragalus polysaccharide solution; (c) Sprinkle the prescribed amount of gel matrix on the water surface of the astragalus polysaccharide solution, and refrigerate overnight at about 4°C to obtain a clear, lump-free, uniformly dispersed solution, 0.45 μm or 0.22 μm microporous membrane filtration; (d) dilute (c) with water for injection, and stir evenly to obtain. The in vitro performance evaluation method is the same as in Implementation 1, and the results are shown in Table 1, and the in vitro release results are shown in the accompanying drawings. the

Implementation example 4:

8% astragalus polysaccharide in situ gel for injection

prescription

Preparation method: (a) take an appropriate amount of astragalus polysaccharide raw material prepared by the method of water extraction and alcohol precipitation, and dissolve it with an appropriate amount of water for injection; (b) add benzyl alcohol, sodium benzoate and sodium tartrate aqueous solution to the above-mentioned astragalus polysaccharide solution; (c) add the prescription Sprinkle a certain amount of gel matrix and sodium carboxymethyl cellulose on the water surface of the astragalus polysaccharide solution, refrigerate overnight at about 4°C to obtain a clear, lump-free, uniformly dispersed solution, and filter through a 0.45 μm or 0.22 μm microporous membrane; (d) Dilute (c) to volume with water for injection, stir evenly, and obtain. The in vitro evaluation method is the same as in Implementation 1, and the results are shown in Table 1, and the in vitro release results are shown in the accompanying drawings. the

Implementation example 5:

10% astragalus polysaccharide in situ gel for injection

prescription

Preparation method: (a) take an appropriate amount of astragalus polysaccharide raw material prepared by water extraction and alcohol precipitation method, and dissolve it with an appropriate amount of water for injection; (b) add benzalkonium bromide and sodium tartrate aqueous solution to the above-mentioned astragalus polysaccharide solution; (c) add the prescription amount Sprinkle the gel matrix and sodium carboxymethyl cellulose on the water surface of the astragalus polysaccharide solution, refrigerate overnight at about 4°C to obtain a clear, lump-free, and uniformly dispersed solution, and filter through a 0.45 μm or 0.22 μm microporous membrane; ( d) Dilute (c) to volume with water for injection, stir evenly, and obtain. the

The specific preparation method and in vitro evaluation method are the same as in Implementation 1. The results are shown in Table 1, and the in vitro release results are shown in the accompanying drawings. the

Table 1 In vitro performance evaluation of astragalus polysaccharide injection in situ gel

Claims (6)

1. Prepare an in situ gel for injection of astragalus polysaccharide with a suitable phase transition temperature, which is characterized in that it comprises the following components: 1.0-15.0 parts by weight of astragalus polysaccharide, 15.5-30.5 parts by weight of poloxamer 407, and 15.5-30.5 parts by weight of poloxamer 0.5-10.5 parts by weight of Mu 188, 25.0-80.0 parts by weight of water, 0.01-10.0 parts by weight of polymer blocker, 0.01-2.0 parts by weight of pH regulator, and 0.01-3.0 parts by weight of preservative. The gel has an appropriate phase transition temperature, can exist in a liquid state at room temperature, and forms a solid or semi-solid gel in the body after administration. 2. The astragalus polysaccharide in situ gel for injection with a suitable phase transition temperature according to claim 1, characterized in that the temperature-sensitive gel matrix in the preparation is selected from Poloxamer 407 and Poloxamer 188 , wherein Poloxamer 407 accounts for 15.5-30.5 parts by weight, and Poloxamer 188 accounts for 0.5-10.5 parts by weight. Adjusting the amount of the two can control the phase transition temperature of the preparation to be between 32°C and 37°C. 3. A kind of astragalus polysaccharide in situ gel for injection with suitable phase transition temperature according to claim 1, characterized in that the polymer blocker is selected from polyvinyl alcohol, povidone, methyl Cellulose, hydroxypropyl methylcellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, sodium carboxymethyl cellulose, carbomer, hyaluronic acid, xanthan gum, chitosan, carboxymethyl shell Any one or more of polysaccharides, sodium alginate, phospholipids, etc.; preferably povidone 1.0-10.0 parts by weight, or hydroxypropyl methylcellulose 0.02-0.8 parts by weight, or sodium carboxymethylcellulose 0.05- 0.8 parts by weight. 4. A kind of astragalus polysaccharide in situ gel for injection with suitable phase transition temperature according to claim 1, characterized in that the pH regulator in the preparation is selected from lactic acid, acetic acid, hydrochloric acid, citric acid and Any one or more of sodium salt, tartaric acid and its sodium salt, sodium carbonate, sodium bicarbonate, dipotassium hydrogen phosphate and potassium dihydrogen phosphate, disodium hydrogen phosphate and sodium dihydrogen phosphate, sodium hydroxide, monoethanolamine, etc. ; Preferably 0.01-0.5 parts by weight of sodium tartrate, or 0.01-0.5 parts by weight of sodium hydroxide. 5. A kind of astragalus polysaccharide in situ gel for injection with suitable phase transition temperature according to claim 1, characterized in that the preservatives in the preparation are selected from parabens, chlorobutanol, benzene Any one or more of methanol, phenylethyl alcohol, chlorhexidine acetate, benzalkonium bromide, benzalkonium chloride, sodium benzoate, potassium sorbate, thimerosal, etc.; preferably 0.5-1.2 parts by weight of benzyl alcohol, or 0.2 parts by weight of sodium benzoate -0.5 parts by weight, or 0.01-0.5 parts by weight of benzalkonium bromide. 6. A method for preparing an astragalus polysaccharide in situ gel for injection with a suitable phase transition temperature according to claim 1, characterized in that it comprises the following steps: (a) taking the astragalus polysaccharide bulk drug prepared by water extraction and alcohol precipitation Appropriate amount, dissolve with appropriate amount of water for injection; (b) dissolve the selected pH regulator and preservative with a small amount of water for injection respectively, and then add them to the above-mentioned astragalus polysaccharide solution; Sprinkle the molecular blocker on the astragalus polysaccharide solution, and refrigerate at about 4°C for 24 hours to obtain a clear, lump-free, and uniformly dispersed solution, which is filtered through a 0.45 μm or 0.22 μm microporous membrane; (d) the solution obtained in (c) Dilute to volume with water for injection, stir evenly, and get ready.

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