Gel for cultivating bovine gallstones in vivo and preparation method thereof

Abstract

The application discloses a kind of in-vivo cultivation of bovine calculus bezoar gel and preparation method thereof, it is related to in-vivo cultivation of bovine calculus bezoar technical field.A kind of in-vivo cultivation of bovine calculus bezoar gel, including following mass component: with mass percentage, 0.1%~5% drug depot, 8%~20% high molecular gel skeleton, 0.1%~2% crosslinking agent, 0.1%~2% crosslinking regulator, 50%~65% solvent, 10%~40% crosslinking agent solution;Drug depot exists in the form of β-glucuronidase sustained-release microsphere.The sponge-like porous gel prepared by the application can be injected into the gallbladder of a cow to form a sponge-like bovine calculus bezoar bed through crosslinking reaction, the bovine calculus bezoar bed can slowly release β-glucuronidase, promote the decomposition of conjugated bilirubin into free bilirubin, and finally the free bilirubin forms calcium bilirubinate, which, after combining with mucin, forms bovine calculus bezoar particles, and the particles are enriched by the bovine calculus bezoar bed to form bovine calculus bezoar with high yield and good quality.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of in-vivo cultivation of bovine bezoar, and particularly relates to a gel for in-vivo cultivation of bovine bezoar and a preparation method thereof. BACKGROUND

[0002] Bovine bezoar is a kind of precious traditional Chinese medicine, which has the effects of clearing heart, cooling liver, detoxifying and opening orifice, and is applied to the treatment of various difficult diseases, but the natural bovine bezoar resource is extremely scarce, which is difficult to meet the market demand, therefore, the artificial bovine bezoar technology is gradually developed.

[0003] Patent CN202411057451.9 a bovine bezoar cultivation temperature-sensitive gel and its preparation method and application discloses that N-isopropyl acrylamide, acrylamide and N,N-methylene bisacrylamide are used as raw materials to prepare polyacrylamide hydrogel, which is then injected into the gallbladder of a bovine to cultivate bovine bezoar in-vivo, the gel prepared by the scheme has temperature sensitivity and can quickly change into a viscous state capable of enriching bovine bezoar, but the gel prepared by the scheme has too fast degradation time, and when the degradation is completed, bovine bezoar small particles have not been formed, which increases the difficulty of collecting bovine bezoar.

[0004] Therefore, the present application provides a gel for in-vivo cultivation of bovine bezoar and a preparation method thereof to solve the problems mentioned in the background. SUMMARY

[0005] The present application aims to provide a gel for in-vivo cultivation of bovine bezoar and a preparation method thereof to solve the problems in the prior art.

[0006] To achieve the above-mentioned purpose, the present application provides the following technical scheme: a gel for in-vivo cultivation of bovine bezoar, comprising the following mass components: 0.1% to 5% of a drug depot, 8% to 20% of a high molecular gel matrix, 0.1% to 2% of a crosslinking agent, 0.1% to 2% of a crosslinking regulator, 50% to 65% of a solvent, and 10% to 40% of a crosslinking agent solution.

[0007] The drug depot exists in the form of beta-glucuronidase sustained-release microspheres.

[0008] A preparation method of the gel for in-vivo cultivation of bovine bezoar, comprising the following steps:

[0009] The beta-glucuronidase sustained-release microspheres, the high molecular gel matrix, the crosslinking agent, the crosslinking regulator and the solvent are mixed and stirred uniformly to obtain a gel emulsion, the crosslinking agent solution is added to the gel emulsion and stirred uniformly to obtain the gel for in-vivo cultivation of bovine bezoar.

[0010] Further, the gel for in-vivo cultivation of bovine bezoar is a sponge-like porous gel, which is referred to as a sponge-like gel hereinafter.

[0011] Further, the preparation method of the crosslinking agent solution is as follows:

[0012] The crosslinking agent and the solution are mixed in a mass ratio of 1: (9-99), uniformly stirred to obtain a crosslinking agent solution with a mass fraction of 1%-10%;

[0013] The solution is bovine bile.

[0014] Further, the high polymer gel skeleton is one or a mixture of several of polyacrylic acid, polyacrylic acid sodium salt, sodium polyacrylate, carbomer, sodium alginate, sodium carboxymethyl cellulose, and cross-linked sodium carboxymethyl cellulose.

[0015] The crosslinking agent is one or a mixture of several of glycine aluminum, aluminum hydroxide, glycine aluminum hydroxide, aluminum chloride, aluminum citrate, synthetic aluminum silicate, and calcium chloride.

[0016] The crosslinking regulator is one or a mixture of several of tartaric acid, citric acid, phosphoric acid, malic acid, and edetic acid.

[0017] The solvent is glycerol.

[0018] Further, the β-glucuronidase sustained-release microspheres are prepared by the following process:

[0019] S1: PLGA (poly (lactic-co-glycolic acid)) is mixed with dichloromethane, vortexed and dissolved to obtain a mixed solution;

[0020] S2: β-glucuronidase is added to the mixed solution, vortexed and uniformly dispersed, polyvinyl alcohol is added, stirred, NaCl is added, and stirred and solidified to obtain β-glucuronidase sustained-release microspheres.

[0021] Further, in S1, the mass ratio of PLGA to dichloromethane is 1: (5-15).

[0022] Further, in S2, the mass ratio of the mixed solution, β-glucuronidase, polyvinyl alcohol, and NaCl is 100: (0.01-8): (700-1100): (900-1100).

[0023] Further, in S2, polyvinyl alcohol is added in the form of a solution with a mass fraction of 1%-4%;

[0024] In S2, NaCl is added in the form of a solution with a mass fraction of 1%-10%;

[0025] The temperature of the NaCl solution is 0°C-15°C.

[0026] Further, in S2, the process conditions for stirring and solidification are as follows: temperature 30°C-40°C, and time 1h-3h.

[0027] Compared with the prior art, the present application has the following advantages:

[0028] 1、The sponge-like gel prepared by the present application can cross-link in the bovine gallbladder to form a sponge-like gallstone bed, slowly release β-glucuronidase, promote the decomposition of conjugated bilirubin into free bilirubin, and finally the free bilirubin forms calcium bilirubinate, which, after combining with mucin, forms small gallstone particles that are enriched by the gallstone bed to form high-yield and high-quality gallstones.

[0029] 2、The sponge-like gel prepared by the present application has a porous structure and strong adsorption capacity, which is conducive to the precipitation of gallstone crystals, shortens the formation period of gallstones, and is conducive to the improvement of production efficiency.

[0030] 3、The gel emulsion prepared by the present application has a fast cross-linking speed, good injectability, and can quickly form a gel in the body. The viscoelastic response of the sponge-like gel shows that the elastic modulus (G') is greater than the viscous modulus (G"), and the gallstone bed can be kept for a long time without being washed away in rotating bile. The gallstone bed is implanted by injection, without surgery and incision, reducing the risk of bovine disease and being easy for breeders to operate, with little stress on the bovine body. BRIEF DESCRIPTION OF DRAWINGS

[0031] Figure 1 is a release curve diagram of β-glucuronidase in the gel for cultivating gallstones in vivo according to the present application;

[0032] Figure 2 is a scanning electron microscope image of the sponge-like gel prepared in Example 1 of the present application;

[0033] Figure 3 is a scanning electron microscope image of the sponge-like gel prepared in Example 2 of the present application;

[0034] Figure 4 is a scanning electron microscope image of the sponge-like gel prepared in Example 3 of the present application;

[0035] Figure 5 is a scanning electron microscope image of the sponge-like gel prepared in Example 4 of the present application;

[0036] Figure 6 is a scanning electron microscope image of the sponge-like gel prepared in Example 5 of the present application;

[0037] Figure 7 is a scanning electron microscope image of the sponge-like gel prepared in Example 6 of the present application;

[0038] Figure 8 is a scanning electron microscope image of the sponge-like gel prepared in Example 7 of the present application;

[0039] Figure 9 is a viscoelasticity curve diagram of the sponge-like gel prepared in Example 1 of the present application;

[0040] In which, 106022-01 is the sponge-like gel gel viscoelasticity curve of Example 6; 22012502 is the sponge-like gel gel viscoelasticity curve of Example 7;

[0041] The abscissa represents the oscillation frequency, and the ordinate represents the viscoelastic modulus. DETAILED DESCRIPTION

[0042] The technical solutions in the embodiments of the present application will be clearly and completely described below with reference to the drawings in the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by a person of ordinary skill in the art without creative labor fall within the scope of protection of the present application.

[0043] In the following detailed description,

[0044] PLGA, CAS No. 34346-01-5, item No. P879466-5g, from Macklin;

[0045] Sodium polyacrylate, CAS No. 9003-04-7, item No. S818399-2.5kg, from Macklin;

[0046] Polyacrylic acid, CAS No. 9003-01-4, item No. P815683-500g, from Macklin;

[0047] Beta-glucuronidase, CAS No. 9001-45-0, purity 1000u / mg, item No. TX21552-100KU, from Shanghai Yingxin Laboratory;

[0048] Sodium alginate, CAS No. 9005-38-3, item No. S100126-2.5kg, from aladdin;

[0049] Carbomer, CAS No. 54182-57-9, item No. S31102-250g, from Shanghai Yuan Ye Biology;

[0050] Sodium carboxymethyl cellulose, CAS No. 9004-32-4, viscosity: 10000mpa·s, from Anhui Shanhe;

[0051] Cross-linked sodium carboxymethyl cellulose, CAS No. 74811-65-7, model SD-711, from FMC;

[0052] Example 1: A preparation method of a gel for in-vivo cultivation of bovine calculus, comprising the following steps:

[0053] (1) Preparation of β-glucuronidase sustained-release microspheres:

[0054] S1: 10g of PLGA was mixed with 100g of dichloromethane, and vortexed to dissolve, to obtain a mixed solution; S2: 0.01g of β-glucuronidase was added to the mixed solution and vortexed to disperse uniformly, 800g of a 2% polyvinyl alcohol solution was added, stirred, 1000g of a 5% NaCl solution at 0°C was added, and stirred to solidify, to obtain β-glucuronidase sustained-release microspheres; the process conditions for stirring and solidification were: temperature 40°C, time 3h;

[0055] (2) Preparation of calcium chloride solution:

[0056] Calcium chloride and bovine bile were mixed in a mass ratio of 1:19, and stirred uniformly, to obtain a 5% calcium chloride solution;

[0057] (3) Preparation of sponge-like gel:

[0058] 2g of β-glucuronidase sustained-release microspheres, 15g of polyacrylic acid sodium, 1g of glycoxyaluminum, 1g of tartaric acid, and 80g of glycerol were mixed and stirred uniformly, to obtain a gel emulsion, 40g of a 5% calcium chloride solution was added to the gel emulsion and stirred uniformly, to obtain a sponge-like gel.

[0059] Example 2: A preparation method of a gel for cultivating bovine calculus in vivo, comprising the following steps:

[0060] (1) Preparation of β-glucuronidase sustained-release microspheres:

[0061] S1: 10g of PLGA was mixed with 100g of dichloromethane, and vortexed to dissolve, to obtain a mixed solution; S2: 0.01g of β-glucuronidase was added to the mixed solution and vortexed to disperse uniformly, 800g of a 2% polyvinyl alcohol solution was added, stirred, 1000g of a 5% NaCl solution at 0°C was added, and stirred to solidify, to obtain β-glucuronidase sustained-release microspheres; the process conditions for stirring and solidification were: temperature 40°C, time 3h;

[0062] (2) Preparation of calcium chloride solution:

[0063] Calcium chloride and bovine bile were mixed in a mass ratio of 1:19, and stirred uniformly, to obtain a 5% calcium chloride solution;

[0064] (3) Preparation of sponge-like gel:

[0065] Mix 1g of β-glucuronidase sustained-release microspheres, 10g of polyacrylic acid, 5g of sodium carboxymethyl cellulose, 0.5g of aluminum hydroxide, 0.5g of edetic acid, and 80g of glycerol, stir uniformly to obtain a gel emulsion, add 40g of 5% calcium chloride solution to the gel emulsion, stir uniformly to obtain a sponge-like gel.

[0066] Example 3: A preparation method of a gel for in-vivo cultivation of bovine calculus, comprising the following steps:

[0067] (1) Preparation of β-glucuronidase sustained-release microspheres:

[0068] S1: Mix 10g of PLGA with 100g of dichloromethane, vortex to dissolve to obtain a mixed solution; S2: add 0.01g of β-glucuronidase to the mixed solution, vortex to disperse uniformly, add 800g of 2% polyvinyl alcohol solution, stir, add 1000g of 5% NaCl solution at 0°C, stir and solidify to obtain β-glucuronidase sustained-release microspheres; the process conditions for stirring and solidification are: temperature 40°C, time 3h;

[0069] (2) Preparation of calcium chloride solution:

[0070] Mix calcium chloride and bovine bile at a mass ratio of 1:19, stir uniformly to obtain a 5% calcium chloride solution;

[0071] (3) Preparation of sponge-like gel:

[0072] Mix 5g of β-glucuronidase sustained-release microspheres, 15g of sodium polyacrylate, 5g of sodium alginate, 1.5g of aluminum glyceryl hydroxide, 0.5g of aluminum hydroxide, 1.5g of tartaric acid, 0.5g of citric acid, and 80g of glycerol, stir uniformly to obtain a gel emulsion, add 40g of 5% calcium chloride solution to the gel emulsion, stir uniformly to obtain a sponge-like gel.

[0073] Example 4: A preparation method of a gel for in-vivo cultivation of bovine calculus, comprising the following steps:

[0074] (1) Preparation of β-glucuronidase sustained-release microspheres:

[0075] S1: Mix 10g of PLGA with 100g of dichloromethane, vortex to dissolve to obtain a mixed solution; S2: add 0.01g of β-glucuronidase to the mixed solution, vortex to disperse uniformly, add 800g of 2% polyvinyl alcohol solution, stir, add 1000g of 5% NaCl solution at 0°C, stir and solidify to obtain β-glucuronidase sustained-release microspheres; the process conditions for stirring and solidification are: temperature 40°C, time 3h;

[0076] (2) Preparation of calcium chloride solution:

[0077] Mix calcium chloride and bovine bile at a mass ratio of 1:19, stir evenly, and obtain a 5% calcium chloride solution by mass fraction;

[0078] (3) Preparation of sponge-like gel:

[0079] Mix 0.5g of β-glucuronidase sustained-release microspheres, 10g of polyacrylic acid, 1g of cross-linked sodium carboxymethyl cellulose, 10.5g of aluminum glycinate, 0.5g of citric acid, and 80g of glycerol, stir evenly, and obtain a gel emulsion. Add 40g of 5% calcium chloride solution by mass fraction to the gel emulsion, stir evenly, and obtain a sponge-like gel.

[0080] Example 5: A preparation method of a gel for cultivating bovine calculus in vivo, comprising the following steps:

[0081] (1) Preparation of β-glucuronidase sustained-release microspheres:

[0082] S1: Mix 10g of PLGA with 100g of dichloromethane, vortex to dissolve, and obtain a mixed solution; S2: Add 0.01g of β-glucuronidase to the mixed solution, vortex to disperse evenly, add 800g of 2% polyvinyl alcohol solution by mass fraction, stir, add 1000g of 5% NaCl solution by mass fraction at 0°C, and stir to solidify, to obtain β-glucuronidase sustained-release microspheres; The process conditions for stirring and solidification are: temperature 40°C, time 3h;

[0083] (2) Preparation of calcium chloride solution:

[0084] Mix calcium chloride and bovine bile at a mass ratio of 1:19, stir evenly, and obtain a 5% calcium chloride solution by mass fraction;

[0085] (3) Preparation of sponge-like gel:

[0086] Mix 3g of β-glucuronidase sustained-release microspheres, 10g of polyacrylic acid sodium, 5g of sodium alginate, 5g of carbomer, 1g of aluminum citrate, 1g of citric acid, and 80g of glycerol, stir evenly, and obtain a gel emulsion. Add 40g of 5% calcium chloride solution by mass fraction to the gel emulsion, stir evenly, and obtain a sponge-like gel.

[0087] Example 6: A preparation method of a gel for cultivating bovine calculus in vivo, comprising the following steps:

[0088] (1) Preparation of β-glucuronidase sustained-release microspheres:

[0089] S1: 10 g of PLGA is mixed with 100 g of dichloromethane, and vortexed to dissolve, to obtain a mixed solution; S2: 0.01 g of β-glucuronidase is added to the mixed solution, vortexed to disperse uniformly, 800 g of a polyvinyl alcohol solution with a mass fraction of 2% is added, stirred, 1000 g of a NaCl solution with a mass fraction of 5% and at 0°C is added, and stirred to solidify, to obtain β-glucuronidase sustained-release microspheres; the process conditions for stirring and solidification are: a temperature of 40°C and a time of 3 h;

[0090] (2) Preparation of a calcium chloride solution:

[0091] Calcium chloride and bovine bile are mixed at a mass ratio of 1:19, and stirred uniformly, to obtain a calcium chloride solution with a mass fraction of 5%;

[0092] (3) Preparation of a sponge-like gel:

[0093] 4 g of β-glucuronidase sustained-release microspheres, 8 g of polyacrylic acid, 8 g of sodium alginate, 4 g of cross-linked sodium carboxymethyl cellulose, 1 g of glycoxyaluminum, 1 g of calcium chloride, 1 g of citric acid, 1 g of tartaric acid, and 100 g of glycerol are mixed and stirred uniformly, to obtain a gel emulsion; 50 g of a calcium chloride solution with a mass fraction of 5% is added to the gel emulsion, and stirred uniformly, to obtain a sponge-like gel.

[0094] Example 7: A preparation method of a gel for cultivating bovine calculus in vivo, comprising the following steps:

[0095] (1) Preparation of β-glucuronidase sustained-release microspheres:

[0096] S1: 10 g of PLGA is mixed with 100 g of dichloromethane, and vortexed to dissolve, to obtain a mixed solution; S2: 0.01 g of β-glucuronidase is added to the mixed solution, vortexed to disperse uniformly, 800 g of a polyvinyl alcohol solution with a mass fraction of 2% is added, stirred, 1000 g of a NaCl solution with a mass fraction of 5% and at 0°C is added, and stirred to solidify, to obtain β-glucuronidase sustained-release microspheres; the process conditions for stirring and solidification are: a temperature of 40°C and a time of 3 h;

[0097] (2) Preparation of a calcium chloride solution:

[0098] Calcium chloride and bovine bile are mixed at a mass ratio of 1:19, and stirred uniformly, to obtain a calcium chloride solution with a mass fraction of 5%;

[0099] (3) Preparation of a sponge-like gel:

[0100] 5 g of β-glucuronidase sustained-release microspheres, 10 g of sodium polyacrylate, 8 g of sodium alginate, 2 g of cross-linked sodium carboxymethyl cellulose, 1.5 g of aluminum chloride, 2 g of citric acid and 100 g of glycerol were mixed and stirred evenly to obtain a gel emulsion. 50 g of a 5% by mass calcium chloride solution was added to the gel emulsion and stirred evenly to obtain a sponge gel.

[0101] Example 8: Using Example 1 as a comparison, the mass ratio of each component in the β-glucuronidase sustained-release microspheres, the process parameters during preparation, and the mass fraction of the calcium chloride solution were adjusted, while the other conditions remained unchanged. The specific process is as follows:

[0102] (1) Preparation of β-glucuronidase sustained-release microspheres:

[0103] S1: 5 g of PLGA was mixed with 75 g of dichloromethane and vortexed to dissolve to obtain a mixed solution. S2: 5.2 g of β-glucuronidase was added to the mixed solution and vortexed to disperse it evenly. 650 g of a 4% polyvinyl alcohol solution was added and stirred. 700 g of a 5% NaCl solution at 10°C was added and stirred to obtain β-glucuronidase sustained-release microspheres. The stirring and curing process conditions were: temperature 35°C, time 2 h.

[0104] (2) Preparation of calcium chloride solution:

[0105] Calcium chloride and ox bile were mixed in a mass ratio of 1:9 and stirred evenly to obtain a 10% calcium chloride solution;

[0106] (3) Preparation of sponge gel:

[0107] 2 g of β-glucuronidase sustained-release microspheres, 15 g of sodium polyacrylate, 1 g of aluminum glycolate, 1 g of tartaric acid and 80 g of glycerol were mixed and stirred evenly to obtain a gel emulsion. 40 g of a 10% calcium chloride solution was added to the gel emulsion and stirred evenly to obtain a sponge gel.

[0108] Example 9: Using Example 1 as a comparison, the mass ratio of each component in the β-glucuronidase sustained-release microspheres, the process parameters during preparation, and the mass fraction of the calcium chloride solution were adjusted, while the other conditions remained unchanged. The specific process is as follows:

[0109] (1) Preparation of β-glucuronidase sustained-release microspheres:

[0110] S1: 10 g of PLGA was mixed with 50 g of dichloromethane, vortexed to dissolve, to obtain a mixed solution; S2: 2.8 g of β-glucuronidase was added to the mixed solution, vortexed to disperse uniformly, 660 g of a 1% by mass polyvinyl alcohol solution was added, stirred, 540 g of a 10% by mass NaCl solution at 15°C was added, stirred and solidified, to obtain β-glucuronidase sustained-release microspheres; the process conditions for stirring and solidification were: temperature 30°C, time 1 h;

[0111] (2) Preparation of calcium chloride solution:

[0112] Calcium chloride and bovine bile were mixed in a mass ratio of 1:99, stirred uniformly, to obtain a 1% by mass calcium chloride solution;

[0113] (3) Preparation of sponge-like gel:

[0114] 2 g of β-glucuronidase sustained-release microspheres, 15 g of polyacrylic acid sodium, 1 g of aluminum hydroxyglycerate, 1 g of tartaric acid and 80 g of glycerol were mixed, stirred uniformly, to obtain a gel emulsion, 40 g of a 1% by mass calcium chloride solution was added to the gel emulsion, stirred uniformly, to obtain a sponge-like gel.

[0115] Comparative Example 1: Example 1 was used as a comparison, β-glucuronidase sustained-release microspheres were not prepared, β-glucuronidase was directly blended with polyacrylic acid sodium, aluminum hydroxyglycerate, tartaric acid and glycerol, and the other conditions were unchanged.

[0116] Comparative Example 2: Example 1 was used as a comparison, a calcium chloride solution was not prepared, calcium chloride was directly blended with β-glucuronidase sustained-release microspheres, polyacrylic acid sodium, aluminum hydroxyglycerate, tartaric acid and glycerol, and the other conditions were unchanged.

[0117] Experiment:

[0118] In-vitro release experiment: 10 g of the sponge-like gel obtained in the examples and comparative examples was added to 250 mL of fresh bovine bile solution, placed in a 37°C constant-temperature water bath shaker, set to a shaking speed of 50 r / min, the in-vitro release was determined, samples were taken at regular time intervals every day, after 15 days of release, the release curve of β-glucuronidase in the sponge-like gel was obtained by using the MicroBCA method under an enzyme marker;

[0119] Surface morphology of the sponge-like gel: 10 g of the sponge-like gel obtained in the examples and comparative examples was taken, the surface morphology was observed by using a scanning electron microscope and was recorded;

[0120] Crosslinking time and gel formation morphology: 10 g of the gel emulsion obtained in the examples and comparative examples was taken, a 1 mm needle and a 10 ml needle tube were used, the gel emulsion was injected into 5% calcium chloride bile in a rotating state, the crosslinking time and state were observed and recorded;

[0121] Rheological properties of sponge-like gel: Take the sponge-like gel obtained in Example 6, Example 7, use Thermo Scientific Rheometer (Model: HAAKE MARS 40), the rotor type is flat plate, the rotor model is P35 / Ti-02190347, the detection gap is 1 mm, the detection temperature is 32℃, the scanning mode is Osc oscillation frequency scanning, the detection setting is control shear stress (CS) mode, the shear stress τ = 200 Pa, the oscillation frequency variable range f = 100 Hz ~ 0.1 Hz, 8 data are collected in each order of variable; start the measurement program, evenly coat the sample on the rheometer measurement table, lower the rotor, scrape the edge, and detect the viscoelasticity curve;

[0122] As shown in Figure 9 , the viscoelastic response is that the elastic modulus (G') is greater than the viscous modulus (G"), mainly showing elastic properties, and will not be washed away in the rotating bile.

[0123] The following table is the performance test results of the sponge-like gel;

[0124]

[0125] According to the data in the above table, the following conclusions can be drawn:

[0126] The gels obtained in Examples 1-9 are all sponge porous structures, have strong adsorption capacity, and are beneficial to the enrichment of small particles of bovine gallstones;

[0127] The in vitro release rate of β-glucuronidase in the sponge-like gels obtained in Example 3, Example 6, and Example 7 is relatively slow, which is more conducive to the production of bovine gallstone particles when cultivating bovine gallstones in vivo;

[0128] In Example 6 and Example 7, the viscoelastic response of the sponge-like gel shows that the elastic modulus (G') is greater than the viscous modulus (G"), mainly showing elastic properties;

[0129] In Comparative Example 1, β-glucuronidase is directly blended with sodium polyacrylate, aluminum glyceryl hydroxy, tartaric acid and glycerol, and the in vitro release rate of β-glucuronidase is fast, which is not conducive to the production of bovine gallstone particles;

[0130] In Comparative Example 2, calcium chloride is directly blended with β-glucuronidase sustained-release microspheres, sodium polyacrylate, aluminum glyceryl hydroxy, tartaric acid and glycerol, and the in vitro release rate of β-glucuronidase is fast, and the obtained product is a flocculent particle, and the gelation state is poor;

[0131] In summary, the gels obtained in Example 3, Example 6 and Example 7 have short crosslinking time, and will not be washed away in the rotating bile, have good injectability and can form gel quickly in vivo, and have stronger enrichment of small particles of bovine gallstone.

[0132] It will be obvious to a person skilled in the art that, without departing from the scope of the present application, the application can be implemented in other particular forms. The examples are therefore to be considered as being illustrative and not restrictive, the scope of the application being defined by the claims appended hereto rather than that which precedes it, and it is intended to embrace all technical equivalents which fall within the meaning and range of equivalents of the essential characteristics based on this patent. No figure reference in the claims should be considered as limiting the claims concerned.

Claims

1. A gel for cultivating bezoar in vivo, characterized in that: The invention comprises the following mass components: by mass percentage, 0.1% to 5% drug reservoir, 8% to 20% polymer gel skeleton, 0.1% to 2% cross-linking agent, 0.1% to 2% cross-linking regulator, 50% to 65% solvent, and 10% to 40% cross-linking agent solution; The drug reservoir is in the form of β-glucuronidase sustained-release microspheres; The polymer gel skeleton is one or a mixture of polyacrylic acid, polyacrylic acid sodium salt, sodium polyacrylate, carbomer, sodium alginate, sodium carboxymethyl cellulose, and cross-linked sodium carboxymethyl cellulose; The cross-linking agent is one or a mixture of aluminum glycinate, aluminum hydroxide, aluminum glycolate, aluminum chloride, aluminum citrate, synthetic aluminum silicate, and calcium chloride; The cross-linking regulator is one or a mixture of tartaric acid, citric acid, phosphoric acid, malic acid, and edetic acid; The solvent is glycerol; The cross-linking agent solution is prepared by mixing the cross-linking agent and ox bile in a mass ratio of 1: (9-99).

2. A method for preparing a gel for cultivating bovine gallstones in vivo, characterized by: The following steps are involved: The beta-glucuronidase sustained-release microspheres, a polymer gel skeleton, a crosslinking agent, a crosslinking regulator and a solvent are mixed and stirred evenly to obtain a gel emulsion. The crosslinking agent solution is added to the gel emulsion and stirred evenly to obtain a gel for cultivating bezoar in vivo.

3. The method for preparing a gel for in vivo cultivation of bezoar according to claim 2, wherein: The β-glucuronidase sustained-release microspheres are prepared by the following process: S1: PLGA was mixed with dichloromethane and vortexed to dissolve to obtain a mixed solution; S2: Add β-glucuronidase to the mixed solution, vortex to disperse evenly, add polyvinyl alcohol, stir, add NaCl, stir to solidify, and obtain β-glucuronidase sustained-release microspheres.

4. The method for preparing a gel for in vivo cultivation of bezoar according to claim 3, wherein: In S1, the mass ratio of PLGA to dichloromethane was 1:(5-15).

5. The method of preparing a gel for in-vivo cultivation of bovine gallstones as claimed in claim 3, wherein: In S2, the mass ratio of the mixed solution, β-glucuronidase, polyvinyl alcohol and NaCl is 100: (0.01~8): (700~1100): (900~1100).

6. The method of preparing a gel for in-vivo cultivation of bovine gallstones as claimed in claim 2, wherein: The preparation method of the cross-linking agent solution is as follows: The cross-linking agent and ox bile are mixed in a mass ratio of 1: (9-99), stirred evenly, and a cross-linking agent solution with a mass fraction of 1%-10% is obtained.

7. The method of preparing a gel for in-vivo cultivation of bovine gallstones as claimed in claim 3, wherein: In S2, polyvinyl alcohol is added in the form of a solution with a mass fraction of 1% to 4%; In S2, NaCl is added in the form of a solution with a mass fraction of 1% to 10%.

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