CN115894969B

CN115894969B - Agarose-based hydrogel, preparation method and application thereof

Info

Publication number: CN115894969B
Application number: CN202211657349.3A
Authority: CN
Inventors: 邹圣灿; 林莎莎; 王宝群
Original assignee: Qingdao Chenland Marine Biotechnology Co ltd
Current assignee: Qingdao Chenland Marine Biotechnology Co ltd
Priority date: 2022-12-22
Filing date: 2022-12-22
Publication date: 2025-01-14
Anticipated expiration: 2042-12-22
Also published as: CN115894969A

Abstract

The invention discloses agarose-based hydrogel, a preparation method and application thereof, and belongs to the technical field of injection filling beauty. The agarose-based hydrogel is prepared by uniformly mixing a curing agent and a dispersing agent to prepare a curing system, and placing an agarose solution and the curing system in an alkaline solution for curing reaction, removing a film and cleaning to obtain the agarose-based hydrogel. According to the invention, the agarose is directly solidified to obtain the hydrophilic agarose-based hydrogel suitable for facial filling, so that the problems of difficult purification, obvious granular sensation and the like caused by using modified agarose microspheres are avoided, and the safety of the injection gel is improved. The injection gel prepared by the invention is uniform and fine, has no granular feel, has better cohesive force, can finely support skin and tissues, is not easy to generate movement and free, has no stimulation to the skin, has long local residence time, good plasticity and less side effect, and can realize long-acting filling.

Description

Agarose-based hydrogel, preparation method and application thereof

Technical Field

The invention belongs to the technical field of injection filling beauty, and particularly relates to agarose-based hydrogel, a preparation method and application thereof.

Background

Cosmetic medicine shows a erection in our country. Facial soft tissue filling is one of the current minimally invasive treatment approaches for treating facial volume tissue loss, contour changes and static wrinkles, and ideal soft tissue filling materials should have good biocompatibility, effectiveness and safety. Currently, facial soft tissue filling materials are mainly non-autologous tissue injection materials, such as Hyaluronic Acid (HA), polycaprolactone (PCL) and L-polylactic acid (PLLA), and filling materials derived from autologous tissues, such as fat grafts, platelet-rich plasma and the like. Although hyaluronic acid is still the most popular degradable injection product, its degradation period is short, requiring frequent multiple injections to maintain its filling effect. Therefore, finding an effective and safe tissue filling has been a continuing challenge for plastic and cosmetic surgery. In order to achieve the long-term filling effect, attempts have been made to use biodegradable materials such as polyvinyl alcohol (PVA), polymethyl methacrylate (PMMA), etc. as fillers for microspheres. Although the filling effect of these fillers is maintained for a significantly longer period of time, these materials remain in the body for a longer period of time, releasing harmful substances and thus causing a series of side reactions with high later safety risks.

Agarose is a biodegradable polymer, which has similarities to the extracellular matrix, and has led to a strong interest in its use in the field of dermal fillers. Like HA, agarose is a biocompatible polysaccharide with immediate and long lasting clinical results. However, it differs from HA in that it is stable in vivo and is capable of providing a long-lasting filling effect after tissue implantation.

In the prior art, agarose powder cannot be directly used as a raw material to prepare a hydrogel product suitable for facial filling, and the reason is that agarose is generally heated to more than 90 ℃ in water to be dissolved, and when the temperature is reduced to 35-40 ℃, a good semisolid gel is formed, the gel strength is high, and the agarose gel cannot be pushed and injected by a syringe, and is also the reason that agarose gel is generally only used as an electrophoresis support. Therefore, the current agarose filling products need to prepare agarose powder into agarose microspheres firstly and then take the agarose microspheres as raw materials to prepare hydrogel products suitable for facial filling, for example, in Chinese patent CN104774337A, a preparation method of agarose microsphere-containing crosslinked sodium hyaluronate gel for injection is provided.

The preparation methods of agarose microspheres include emulsification-solidification method, spray method, membrane emulsification method and microfluidic method, and the above methods have a series of problems, for example, the solvent consumption is high during the preparation, the complete removal or the particle size distribution of the prepared microspheres is too wide and the particles are not smooth, etc. Therefore, agarose microsphere is selected as raw material to prepare or mix and crosslink with sodium hyaluronate to prepare gel, and the defects of excessive addition of organic reagent, nonuniform particle size distribution, obvious granular feel and the like exist, so that the probability of inflammatory reaction is increased (bear sail. Preparation modification and application of the agarose microsphere).

In order to solve the problems of complex process, obvious granular sensation, nonuniform grain diameter and the like of agarose microsphere gel, improve the seamless fusion of agarose and myobase fluid, ensure that the prepared hydrogel has a finer pushing effect and improves the safety and effectiveness of the agarose microsphere gel as a face filling product, the invention aims to take agarose powder as a raw material, overcome the prejudice existing in the prior art and explore the agarose hydrogel which is simpler, more convenient, effective and safer and the preparation process thereof.

Disclosure of Invention

The technical scheme of the invention is as follows:

The invention provides a preparation method of agarose-based hydrogel, which comprises the following steps:

And placing the agarose solution and the curing system in alkaline solution for curing reaction, removing the film and cleaning to obtain the agarose-based hydrogel.

In the preparation method, the curing agent is selected from one or more of epichlorohydrin, 1,2,7, 8-dicyclo-octane, divinyl sulfone and 1, 4-butanediol diglycidyl ether.

In the preparation method, the dispersing agent is selected from dimethyl sulfoxide or absolute ethyl alcohol.

In the preparation method, the curing agent accounts for 0.5-10% of the mass of the agarose solution, and the dispersing agent accounts for 1-30% of the mass of the agarose solution.

In the preparation method, the curing reaction condition is that the reaction temperature is 50-70 ℃ and the reaction time is 2-72 h.

In the preparation method, the alkaline solution is selected from sodium hydroxide solution, potassium hydroxide solution, calcium hydroxide solution and barium hydroxide solution, and the concentration of the alkaline solution is selected from 0.5-3M. The volume of the alkaline solution can be 0.1-10 times of that of the agarose solution.

In the preparation method, the agarose solution can be prepared by placing agarose powder into water, and dissolving for 15min to 5h at the temperature of 80 to 125 ℃ to obtain the agarose solution with the concentration of 0.5 to 20 percent.

The preparation method comprises the steps of putting a reactant obtained by a curing reaction into purified water, stirring and cleaning, filtering after cleaning to obtain agarose-based hydrogel, wherein the cleaning speed is 50-400 rpm/min, the cleaning time is 4-48 h, and cleaning liquid is changed every 2-6 h.

The present invention provides agarose-based hydrogels prepared by the above method.

The invention provides application of the agarose-based hydrogel in facial filling and cosmetology.

The invention provides application of the agarose-based hydrogel in preparing facial fillers.

The invention provides an agarose-based hydrogel facial filler which consists of a myobasal liquid and an agarose-based hydrogel, wherein the dosage ratio of the agarose-based hydrogel to the myobasal liquid is selected from 1:0.01-1:1, and g/mL. Preferably from 2:1 to 15:2, g: mL.

The muscle base solution can be one or more selected from phosphate buffer solution, physiological saline, hyaluronic acid, collagen, chitosan, amino acid, cellulose, and sodium alginate, wherein the concentration of the muscle base solution is 0.1-10%

The preparation method of the agarose-based hydrogel facial filler is simple, and the myobasal liquid is fully dispersed in the agarose-based hydrogel. After the preparation is completed, conventional filling sterilization can be performed.

The beneficial effects of the invention are as follows:

The invention overcomes the prejudice in the prior art, obtains the hydrophilic agarose-based hydrogel suitable for facial filling by directly solidifying agarose, avoids the problems of difficult purification, obvious granular feel and the like caused by using modified agarose microspheres, and improves the safety of the injection gel. The injection gel prepared by the invention is uniform and fine, has no granular feel, has better cohesive force, can finely support skin and tissues, is not easy to move and free, has no stimulation to the skin, has long local residence time, good plasticity and less side effect, and can realize long-acting filling. The invention has the advantages of easy control of process conditions, few operation steps and stable product quality, and is suitable for large-scale production.

Drawings

FIG. 1 is a rheology test result;

FIG. 2 is a graph showing the results of a push force test;

FIG. 3 is a tissue section, HE×20, after 1 week of implantation;

FIG. 4 is a tissue section, HE×20, 4 weeks after implantation;

FIG. 5 is a tissue section, HE×20, 30 weeks after implantation.

Detailed Description

Other terms used in the present invention, unless otherwise indicated, generally have meanings commonly understood by those of ordinary skill in the art. The invention will be described in further detail below in connection with specific embodiments and with reference to the data. The following examples are intended to illustrate the invention and are not intended to limit the scope of the invention in any way.

Example 1

Preparation of agarose-based hydrogels:

5g of agarose powder was placed in a beaker, 100mL of purified water was added, and the mixture was dissolved at 121℃for 1 hour to obtain an agarose solution. 2g of epichlorohydrin and 15g of dimethyl sulfoxide are placed in a centrifuge tube to be uniformly mixed, so as to form a curing system. Adding the agarose solution and the curing system into 80mL of sodium hydroxide solution (1.2M), curing at 60 ℃ for 24 hours, taking out a reaction product after the reaction is finished, placing the reaction product into purified water for stirring and cleaning, wherein the stirring speed is 100rpm/min, changing water once every 4-8 hours, and cleaning for 24 hours to obtain the agarose-based hydrogel.

Preparing 20mL of phosphate buffer solution, adding the phosphate buffer solution into the 150g of agarose-based hydrogel, uniformly dispersing, filling the gel into a pre-filling syringe, and performing damp-heat sterilization to obtain the agarose-based hydrogel for injection and filling.

Example 2

Preparation of agarose-based hydrogels:

2g of agarose powder was placed in a beaker, 50mL of purified water was added, and the mixture was dissolved at 121℃for 30 hours to obtain an agarose solution. And placing 0.7g of epichlorohydrin and 10g of dimethyl sulfoxide into a centrifuge tube, and uniformly mixing to form a curing system. Adding the agarose solution and the curing system into 50mL of sodium hydroxide solution (0.7M), curing at 60 ℃ for 24 hours, taking out a reaction product after the reaction is finished, placing the reaction product into purified water for stirring and cleaning, wherein the stirring speed is 100rpm/min, changing water once every 4-8 hours, and cleaning for 24 hours to obtain the agarose-based hydrogel.

Preparing 50mL of 1% sodium hyaluronate solution, adding the sodium hyaluronate solution into the 100g of agarose-based hydrogel, uniformly dispersing, filling the gel into a pre-filling syringe, and performing damp-heat sterilization to obtain the agarose-based hydrogel for injection and filling.

Example 3

Preparation of agarose-based hydrogels:

10g of agarose powder was placed in a beaker, 200mL of purified water was added, and the mixture was dissolved at 121℃for 30 hours to obtain an agarose solution. 5g of epichlorohydrin and 50g of dimethyl sulfoxide are placed in a centrifuge tube to be uniformly mixed, so as to form a curing system. Adding the agarose solution and the curing system into 160mL of sodium hydroxide solution (1M), curing at 60 ℃ for 12 hours, taking out a reaction product after the reaction is finished, placing the reaction product into purified water for stirring and cleaning, changing water once every 4-8 hours at the stirring speed of 200rpm/min, and cleaning for 24 hours to obtain the agarose-based hydrogel.

Preparing 70mL of seaweed polysaccharide solution with the concentration of 2%, adding the seaweed polysaccharide solution into the 300g of agarose-based hydrogel, uniformly dispersing, filling the gel into a pre-filling syringe, and performing damp-heat sterilization to obtain the agarose-based hydrogel for injection filling.

Example 4

Preparation of agarose-based hydrogels:

5g of agarose powder was placed in a beaker, 200mL of purified water was added, and the mixture was dissolved at 100℃for 4 hours to obtain an agarose solution. 2g of 1, 4-butanediol diglycidyl ether and 10g of dimethyl sulfoxide are placed in a centrifuge tube and uniformly mixed to form a curing system. Adding the agarose solution and the solidification system into 100mL of sodium hydroxide solution (1M), solidifying and reacting for 6 hours at 55 ℃, taking out a reaction product after the reaction is finished, placing the reaction product into purified water, stirring and cleaning, wherein the stirring speed is 100rpm/min, changing water for two times, and cleaning for 6 hours altogether to obtain the agarose-based hydrogel.

Adding 2g of sodium hyaluronate, 2g of collagen and 1g of amino acid into 50mL of physiological saline, dissolving and stirring, adding into the 250g of agarose-based hydrogel after complete dissolution, dispersing uniformly, filling the gel into a pre-filling syringe, and performing damp-heat sterilization to obtain the agarose-based hydrogel for injection filling.

Comparative example 1

Placing 5g of agarose powder in 150mL of purified water, dissolving at a high temperature of 121 ℃ for 30min, taking out, placing in a 65 ℃ water bath for heat preservation to obtain a water phase, placing 14g of span 80 in 300mL of liquid paraffin, stirring and mixing for 30min at 65 ℃ to obtain an oil phase, taking out after dissolving the water phase, adding the water phase into the oil phase, emulsifying and homogenizing for 10min, solidifying the homogenized solution for 1h at normal temperature, washing with ethanol after solidification, and carrying out suction filtration to obtain agarose microspheres. 20mL of 1% sodium hyaluronate solution was prepared, added to the 100g agarose microspheres described above, dispersed well, and then the gel was filled into a prefilled syringe and sterilized. The gel is agglomerated into a block shape after sterilization, and when the pushing force is detected, the phenomenon of bursting needles occurs, so that normal test cannot be performed, and the gel is not suitable for injection and cosmetic filling.

Comparative example 2

25G of agarose powder and 5g of non-crosslinked hyaluronic acid were added to 1000mL of phosphate buffer solution (pH 6.8), dissolved by stirring at 80℃for 30min, cooled to room temperature, filled and sterilized. After sterilization, the phenomenon of two-phase layering in the syringe is found, which indicates that the uniformity of the product is low and the syringe is not suitable for injection and cosmetic filling.

(One) viscoelasticity test

Rheological (viscoelasticity) tests were performed on agarose-based hydrogels prepared in examples 1-4 above. The rheological properties are related to the ability of the filler to withstand different types of deformation and forces when implanted in different areas and planes. The above disclosure of the rheological properties of the filling product will guide the clinician in choosing the ideal filling for each region of the face based on pathology and deformation forces acting on the region of interest. Thus, the choice of dermal fillers with the correct rheological characteristics is a key factor in achieving a natural appearance, long lasting desirable aesthetic effect.

The test conditions are as follows, the name of the instrument is rheometer, the test fixture is a 20mm plate on the upper plate, a 60mm plate on the lower plate, the gap is 1mm, and the test description is that the constant temperature 10 ℃ test is performed for amplitude scanning.

The test results are shown in FIG. 1:

The graph shows that the linear viscoelastic areas of the 4 groups of products are similar, the elastic modulus is about 500Pa, the viscous modulus is about 90Pa, the elastic modulus and the viscous modulus are moderate, the gel is smooth and soft, the shaping and pulling effects are obvious after injection, and the bulging feeling is avoided. It is known that the ratio between G 'and G "can reflect the change between the viscosity and elasticity of the gel, and it can be seen from the figure that the ratio of the loss modulus G'/the elastic modulus G" of the four sets of hydrogels is greater than 1, which indicates that the hydrogels prepared by the method of the invention exhibit good elastic properties, can be firmly retained at the injection site, maintain a firm current structural strength, and contribute to the maintenance of the shaping effect after injection.

(II) test of push force

The hydrogel products prepared in example 1, example 2, comparative example 1 and comparative example 2 were subjected to a push force test under the conditions of an instrument name of a universal material tester, a test temperature of room temperature, a test distance of setting the test distance to a full scale range, and a push speed of setting the push speed to 30mm/min.

The test results are shown in FIG. 2:

As can be seen from the graph, the pushing force of the gel in the embodiment 1 and the embodiment 2 is about 10N, meets the standard requirement of injection filling products, has good experience and shows excellent injectability, the pushing force of the gel in the comparative embodiment 1 is linearly reduced after being deformed by 10mm, which shows that the gel has poor fluidity and almost zero smoothness and is not suitable for injection filling in the test process, and the pushing result of the comparative embodiment 2 shows that the fluctuation of the pushing force range is large, which shows that the product continuity is low and the uniformity is poor and is also unfavorable for injection filling.

Physical and chemical Properties

The physical and chemical properties of the agarose-based hydrogel prepared in example 1 were tested and compared with those of agarose gel prepared using agarose microspheres in example 1 of patent CN115260545A (reference example), as shown in table 1:

TABLE 1

As can be seen from Table 1, the two hydrogels were greatly different in shape, and the hydrogels prepared with agarose microspheres were translucent milky and viscous, whereas the hydrogels prepared directly with agarose powder were transparent and elastic, so that they were strong in deformation resistance and not easy to deform. Therefore, the agarose-based hydrogel prepared by the invention is more suitable for the parts such as chin, nose bridge and the like which need lasting plasticity. In the aspect of particle size, the invention realizes that the hydrogel is directly prepared by taking agarose powder as a raw material, so the hydrogel does not contain granular components, has fine texture and no granular feel, and shows more excellent cohesive force, and the injection is smoother, the filling is more natural and the degradation is more uniform. In terms of curing agent residue, the hydrogel provided by the invention does not detect epichlorohydrin residue, has higher safety, and does not cause inflammatory reaction after in vivo injection. In terms of pushing force, the pushing force of the two hydrogels meets the requirements of ergonomics and standards, so that an operator can perform better pushing operation. The hydrogel of the present invention has a degree of crosslinking of 2.1% and less than 10.22% of the reference example, indicating excellent plasticity, low degree of crosslinking (degree of modification) reflects high safety in application, and can reduce allergy rate.

Comparative example 3

Effect of curing reaction temperature on agarose-based hydrogels:

5g of agarose powder was placed in a beaker, 100mL of purified water was added, and the mixture was dissolved at 121℃for 1 hour to obtain an agarose solution. 2g of epichlorohydrin and 15g of dimethyl sulfoxide are placed in a centrifuge tube to be uniformly mixed, so as to form a curing system. Adding agarose solution and solidifying system into 80mL sodium hydroxide solution (1.2M), solidifying at 37 deg.C, 38 deg.C, 39 deg.C and 40 deg.C for 24 hr, taking out reaction product after the reaction is completed, finding that all solidification is failed, making the reaction product in centrifuge tube not form jelly gel, still making it be liquid with high fluidity, placing it into purified water, completely dispersing, and making it not be cleaned and separated.

Comparative example 4

Effect of the type of solidifying agent on agarose-based hydrogels:

2g of agarose powder was placed in a beaker, 50mL of purified water was added, and the mixture was dissolved at 121℃for 30 hours to obtain an agarose solution. 0.5g of adipic acid dihydrazide, 1g of carbodiimide and 10g of dimethyl sulfoxide are placed in a centrifuge tube to be uniformly mixed, so as to form a curing system. Adding the agarose solution and the solidifying system into 50mL of sodium hydroxide solution (0.7M), solidifying at 60 ℃ for 24 hours, taking out a reaction product after the reaction is finished, placing the reaction product into purified water, stirring and cleaning, wherein the stirring speed is 100rpm/min, changing water once every 4-8 hours, and cleaning for 24 hours.

Preparing 50mL of 1% sodium hyaluronate solution, adding the sodium hyaluronate solution into 100g of agarose-based hydrogel, dispersing uniformly, filling the gel into a prefilled syringe, carrying out damp-heat sterilization, finding that the product is hydrated, losing the gel characteristics, and solidifying and hardening the hydrated gel after the temperature is reduced to room temperature, so that the gel cannot be injected. The reason for this is that adipic Acid Dihydrazide (ADH) and carbodiimide (EDC) are crosslinking agents commonly used for carboxyl sites, and there is no carboxyl site in the agarose molecular structure, so that the carboxyl crosslinking method is not suitable for preparing agarose gel, and the crosslinking effect is poor.

Comparative example 5

Effect of curing agent amount on agarose-based hydrogels:

5g of agarose powder was placed in a beaker, 200mL of purified water was added, and the mixture was dissolved at 100℃for 4 hours to obtain an agarose solution. 0.8g of 1, 4-butanediol diglycidyl ether and 10g of dimethyl sulfoxide are placed in a centrifuge tube and uniformly mixed to form a curing system. The agarose solution and the solidification system are added into 100mL of sodium hydroxide solution (1M), solidification reaction is carried out for 6 hours at 55 ℃, after the reaction is finished, the reaction product is taken out, no gel is formed, and the reaction system is yellow water-shaped liquid. After the mixture is cooled to room temperature, the fluidity is still strong, and the properties are not changed. Thus, under this preparation parameter, the support force and viscoelasticity required for filling the injection product cannot be obtained, and the gel cannot be used as an injection filling gel.

Based on the preparation method, the dosage of the curing agent is continuously adjusted to 0.5g, 0.6g and 0.7g, and the curing reaction is carried out for 6 hours. After the completion of the reaction, the reaction system was found to be a yellow aqueous liquid as well, and failed to form a gel. After the mixture is cooled to room temperature, the fluidity is still strong, and the properties are not changed. This means that if the agarose gel which is suitable for facial filling is to be prepared directly from agarose powder, the amount of the solidifying agent to be used needs to be increased.

Application example 1

In vivo implant degradation assay:

The injectable gel prepared in example 1 was subjected to a rabbit in vivo implantation degradation test. Before the test, rabbit hair on two sides of the rabbit vertebra is cut off, intravenous injection anesthesia is selected or not according to animal reaction conditions, the skin of an operation area is disinfected by iodine tincture and 75% ethanol according to the requirement of a surgical routine operation, 4 implantation points are respectively selected at equal distances at the positions of about 2.5cm on two sides of the rabbit vertebra, each point is spaced by 2.5cm, and 0.2mL of sample is implanted into each point. After implantation, animals are sacrificed at 1 week, 4 weeks and 30 weeks respectively, skins at two sides of the spine are separated, subcutaneous implanted materials are exposed, the size, shape, the existence of suppuration and secretion of surrounding tissues and the like of an implanted sample are observed, specimens of surrounding tissue parts containing the materials are taken, tissues which are wrapped around the specimens and are about 0.5 cm to 1cm are cut, 10% formalin solution is used for fixation, dehydration and transparency, paraffin embedding, continuous sections are carried out, the thickness is 5 mu m, hematoxylin-eosin staining is carried out, and pathological conditions of the tissues are observed under a microscope.

The test results are shown in fig. 3-5:

the pathological results of implantation for 1 week (figure 3) and 4 weeks (figure 4) show that no obvious abnormality is found around the sample, which indicates that the injection gel has better compatibility with surrounding tissues and shows good biocompatibility and tissue adaptability. The sections implanted for 30 weeks (fig. 5) were observed to find that there was still a sample present in the body, indicating that the hydrogel products of the present invention have excellent in vivo degradability and long-lasting filling properties.

The above description is only a preferred embodiment of the present invention, and is not intended to limit the invention in any way, and any person skilled in the art may make modifications or alterations to the disclosed technical content to the equivalent embodiments. However, any simple modification, equivalent variation and variation of the above embodiments according to the technical substance of the present invention still fall within the protection scope of the technical solution of the present invention.

Claims

1. A method for preparing an agarose-based hydrogel, characterized in that the steps are as follows:

The curing agent and the dispersant are mixed evenly to prepare a curing system; the agarose solution and the curing system are placed in an alkaline solution to perform a curing reaction, and the film is removed and cleaned to obtain an agarose-based hydrogel;

The curing agent is selected from one or more of epichlorohydrin, 1,2,7,8-diepoxyoctane, and 1,4-butanediol diglycidyl ether;

The dispersant is selected from dimethyl sulfoxide or anhydrous ethanol;

The solidifying agent accounts for 0.5-10% of the mass of the agarose solution; the dispersing agent accounts for 1-30% of the mass of the agarose solution;

The curing reaction conditions are: reaction temperature is 50-70°C, reaction time is 2-72h;

The preparation method of the agarose solution is as follows: agarose powder is placed in water, and dissolved at 80-125° C. for 15 min-5 h to obtain an agarose solution with a concentration of 0.5-20%.

2. method according to claim 1, is characterized in that, the alkaline solution is selected from sodium hydroxide solution, potassium hydroxide solution, calcium hydroxide solution, barium hydroxide solution, and the concentration of the alkaline solution is selected from 0.5~3M.

3. An agarose-based hydrogel prepared by the method according to any one of claims 1 to 2.

4. Use of the agarose-based hydrogel according to claim 3 in the preparation of facial fillers.

5. An agarose-based hydrogel facial filler, characterized in that it is composed of a base solution and the agarose-based hydrogel according to claim 3; wherein the amount ratio of the agarose-based hydrogel to the base solution is selected from 1:0.01 to 1:1, g:mL.

6. The facial filler according to claim 5, characterized in that the base fluid is selected from one or more of phosphate buffer, physiological saline, hyaluronic acid, collagen, chitosan, amino acids, cellulose, and sodium alginate; wherein the concentration of the base fluid is selected from 0.1-10%.