CN115382011A

CN115382011A - Skin injection filler gel and preparation process thereof

Info

Publication number: CN115382011A
Application number: CN202210710969.2A
Authority: CN
Inventors: 张雪峋
Original assignee: Yirui Biotechnology Suzhou Co ltd
Current assignee: Yirui Biotechnology Suzhou Co ltd
Priority date: 2021-07-01
Filing date: 2022-06-22
Publication date: 2022-11-25

Abstract

The invention discloses a skin injection filler gel and a preparation method thereof, wherein a hydrothermal synthesis process is adopted, phosphate, calcium salt and water for injection are used as reaction materials, and a hydrothermal reaction is carried out in a reaction kettle to obtain hydroxyapatite crystals; then uniformly mixing the prepared hydroxyapatite, suspending agent and anti-flocculation agent with water for injection, and putting the mixture into a reaction kettle for hydrothermal reaction to obtain hydroxyapatite filler gel, wherein the size of the hydroxyapatite particles is within the range of 10-80 mu m; a method of wet-grinding agglomerates of a hydroxyapatite filler gel by stirring the hydroxyapatite filler gel with a stirring device having a high shearing force, wherein the average particle diameter is 1 μm. The preparation method of the filler gel for skin injection has the advantages of high safety grade of used raw materials, simple process, low cost and better application prospect.

Description

Skin injection filler gel and preparation process thereof

Technical Field

The invention belongs to the technical field of cosmetic reshaping, and particularly relates to a hydroxyapatite filler gel for injection type cosmetic reshaping and a preparation process thereof.

Background

With the improvement of the quality of life, people not only pursue healthy life, but also pursue beautiful life. People seek a safe, effective and economical way to mitigate the signs of aging and change their appearance. The medical cosmetology is a series of treatments carried out by medical means to achieve the purposes of changing the external form and color of the human body, partially improving the physiological function of the human body and enhancing the external aesthetic feeling of the human body. With the development of modern medical beauty technology and the improvement of medical beauty requirements of people, minimally invasive beauty has become a modern medical beauty trend. As an important minimally invasive cosmetic means, the injection cosmetic technology refers to a medical cosmetic means which uses a percutaneous injection method to inject specific injection (such as various fillers) to a target position to achieve the effects of rejuvenation and beautification. It has the advantages of small wound, no pain, high safety, instant effect and the like, and is accepted by vast majority of amateurs and medical workers. Four classes of absorbable materials that have been approved for injectable fillers for soft tissue augmentation include: collagen, sodium hyaluronate, hydroxyapatite and polylactic acid. Wherein the hydroxyapatite (Calcium hydroxide, caHA, ca) ₁₀ (PO ₄ ) ₆ (OH) ₂ ) Is the main component of inorganic mineral composing the teeth and bones of organism, has excellent biodegradability and biocompatibility, is safe, nontoxic and harmless to human body, and the particles can stimulate the generation of endogenous collagen. In recent years, hydroxyapatite has been widely used as an injectable filler. The calcium hydroxyapatite particles similar to bones are uniformly suspended in suspending agents such as sodium carboxymethylcellulose to form gel, so that a new gel matrix is generated at an injection site, the maintenance time is more than 15 months or even longer, and the gel is finally metabolized normally by the body.

The existing hydroxyapatite preparation technology generally needs to prepare hydroxyapatite by a high-temperature sintering method, then uniformly mix the hydroxyapatite with other auxiliary materials, and finally carry out vacuum heating to 80-120 ℃ for sterilization treatment. The process faces three main problems in the industrial production process: 1) In order to ensure that the filler gel has no bubbles, the gel is easy to dehydrate in the vacuum degassing and filling process, so that a two-phase region is formed, and the product quality is seriously influenced; 2) In order to improve the stability of the filler gel, the dynamic viscosity of the gel is intentionally increased, so that the extrusion force is increased, the clinical injection is not facilitated, and if the viscosity is reduced, hydroxyapatite is easy to deposit, so that the product quality is seriously influenced; 3) High-temperature sintering is needed, the energy consumption is high, the process flow is complex, and the production period is long; 4) When the particles are mixed in the composition and not sufficiently stirred, it may be difficult to disperse particles having a small particle size, and particles having a large particle size may not be suitable for use of the composition.

Disclosure of Invention

The invention aims to provide a hydroxyapatite-containing injection filler gel which has excellent biocompatibility, durability and high moisturizing ability and can keep expected mechanical and rheological properties as a filler for cosmetic reshaping so as to achieve expected cosmetic reshaping effects on the basis of the prior art. Further, the present invention can easily disperse a composition containing hydroxyapatite having a large particle size and a composition containing hydroxyapatite having a small particle size in a liquid while suppressing damage to crystals of hydroxyapatite.

The invention also aims to provide a preparation process of the hydroxyapatite-containing injection filler gel.

The technical scheme of the invention is as follows:

(1) Weighing phosphate and calcium salt, adding water for injection into the phosphate and the calcium salt, and uniformly mixing to obtain a solution A;

(2) Putting the uniformly mixed solution A into a reaction kettle, heating to 80-280 ℃, controlling the stirring speed in the kettle to be 80-800 r/min, and preserving heat for 0.5-10.0 h;

(3) After the reaction is finished, cooling the solution to room temperature to obtain hydroxyapatite crystals;

(4) Mixing the hydroxyapatite, the suspending agent and the deflocculating agent obtained in the step (3) with water for injection, and uniformly stirring to obtain a solution B;

(5) Putting the solution B into a reaction kettle, heating to 80-200 ℃, controlling the stirring speed in the kettle to be 80-800 r/min, and preserving heat for 0.5-5.0 h to obtain hydroxyapatite filler gel;

(6) The hydroxyapatite filler gel is wet-ground by stirring with a stirring device having a high shearing force, and the average particle diameter thereof is 1 μm.

In the step (1), the preferable material of the phosphate is one or more of diammonium hydrogen phosphate, dipotassium hydrogen phosphate trihydrate and sodium phosphate.

In the step (1), the preferable material of the calcium salt is one or more of calcium nitrate, calcium chloride, calcium bicarbonate, calcium lactate and calcium gluconate.

In the step (1), the ratio of the amounts of the calcium ion and phosphate ion substances in the solution a is 1.5 to 2.0, preferably 1.65 to 1.75. For example 1.67 or 1.7 or 1.73.

In the step (2), the hydrothermal synthesis reaction is conventional in the art, and is generally carried out in a reaction kettle. The hydrothermal synthesis reaction preferably includes a reaction temperature and a reaction time. The reaction temperature is preferably 80 to 180 ℃, more preferably 80 to 150 ℃. The reaction time is preferably 0.5 to 4.0 hours, more preferably 1.0 to 3.0 hours.

In the step (3), the particle size of the hydroxyapatite crystal is in the range of 1-80 μm, and the preferable particle size is 10-60 μm.

And (3) naturally cooling to obtain hydroxyapatite crystals.

In the step (4), the preferable material of the suspending agent is one or more of methylcellulose, ethylcellulose, hydroxyethyl cellulose, sodium carboxymethyl cellulose, peach gum, gelatin, chondroitin sulfate, sodium alginate and glycerin for injection.

In the step (4), the preferable material of the deflocculant is one or more of sodium citrate and sodium chloride.

In the step (4), the solution B contains 35-60% of water for injection by mass.

In the step (4), the suspending agent accounts for 15-30% of the solution B by mass.

In the step (4), the mass ratio of the deflocculant in the solution B is 0-8%.

In the step (4), the mass ratio of the hydroxyapatite to the solution B is preferably 1.

In the step (5), the reaction temperature is preferably 100 to 150 ℃, and more preferably 100 to 120 ℃. The reaction time is preferably 0.5 to 2.0 hours, more preferably 0.5 to 1.0 hour.

On the basis of the common knowledge in the field, the above preferred conditions can be combined randomly to obtain the preferred embodiments of the invention.

The reagents and raw materials used in the present invention are commercially available.

By adopting the technical scheme of the invention, the advantages are as follows:

(1) The preparation method of the filler gel for injection type beauty and plastic has the advantages of high safety grade of used raw materials, simple process, low cost, no need of high-temperature sintering in the preparation process and low energy consumption.

(2) The hydroxyapatite particles prepared by the method have controllable particle size range, uniform size and proper crystallinity, and are not easy to agglomerate.

(3) The filler gel for skin injection prepared by the invention realizes industrial vacuum filling without two-phase region by controlling the particle size of the filler and the dosage ratio of the suspending agent to the deflocculant, and can realize industrial large-scale production.

(4) The synthesis reaction of the invention is carried out in a reaction kettle, and has the sterilization function, so that the wet heat sterilization treatment is not needed, thereby avoiding the viscosity reduction, being beneficial to dispersing the hydroxyapatite in the gel for a long time and being easy for clinical injection.

(5) The skin injection filler gel prepared by the invention has good biocompatibility, can be shaped as required at the initial stage of injection into subcutaneous tissues, can be used as a scaffold to promote fibroblast ingrowth and stimulate the generation of new collagen tissues, can form firm biological bonding, has long effect duration and has good application prospect.

Description of the drawings:

the accompanying drawings, which are included to provide a further understanding of the invention and are incorporated in and constitute a part of this specification, illustrate embodiments of the invention and together with the description serve to explain the invention and not to limit the invention. In the drawings:

FIG. 1 is an XRD pattern of hydroxyapatite crystals prepared in example 2;

FIG. 2 is an SEM photograph of the dried dermoadject filler gel prepared in example 2.

Detailed Description

Specific embodiment example 1:

331.65g of calcium chloride, 236.75g of diammonium phosphate and 1000.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 80 ℃, kept at the temperature for 4.0h, cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD to be hydroxyapatite. Then dispersing the precipitate into 500.00g of water for injection, adding 20.00g of methylcellulose, 30.00g of sodium carboxymethylcellulose, 200.00g of glycerol for injection (18.8%), and 10.00g of sodium citrate (0.752%), mixing and stirring uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 80 ℃, preserving heat for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out particle size test on the gel by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 30-60 mu m is 90%; cell culture experiment results show that the obtained solution has better biocompatibility; the cytotoxicity test detection is carried out by adopting an MTT method, the cell proliferation rate is more than 85 percent, and the obtained gel has no toxicity to cells.

Under the condition of a shear rate of 8.000-10.000 l/s at 25 ℃, the dynamic viscosity is 200000-250000 cP, the product is extruded by the aid of a needle with the thickness of 25G (0.26 mm inner diameter), the aid is 59.3N, the injection is smooth, and the product is not layered.

Specific embodiment example 2:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 1000.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 4.0h, then cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD and is known to be hydroxyapatite. And dispersing the precipitate into 580.00g of water for injection, adding 50.00g of sodium carboxymethylcellulose, 200.00g of glycerol for injection (15.99%) and 5.00g of sodium citrate (0.32%), mixing and stirring uniformly, pouring all the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 120 ℃, preserving the temperature for 2.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out particle size test on the sol by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 30-60 mu m is 85%; after drying the obtained gel, most of the particle diameters of the particles can be found to be more than 30 μm through SEM observation; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity test detection is carried out by adopting an MTT method, the cell proliferation rate is more than 88 percent, and the result shows that the prepared gel has no toxicity to cells.

The dynamic viscosity is 210000 cP-250000 cP at 25 ℃ under the condition that the shear rate is 8.000 l/s-10.000 l/s, accordingly, the product is extruded from a 25G (0.26 mm inner diameter) thick needle with boosting force, the corresponding boosting force is 62.3N, the injection is smooth, and the product is not layered.

Specific embodiment example 3:

331.65g of calcium chloride, 681.47g of sodium phosphate and 1000.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 100 ℃, kept at the temperature for 3.0h, then cooled to room temperature, and the precipitate obtained by filtration is analyzed by XRD to be hydroxyapatite. Then dispersing the precipitate into 677.5g of water for injection, adding 10.00g of methylcellulose, 50.00g of sodium carboxymethylcellulose, 180.00g of glycerol for injection (12.40%), and 5.00g of sodium citrate (0.258%), mixing and stirring uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving heat for 3.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out particle size test on the gel by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 40-60 mu m is 90%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity experiment detection is carried out by adopting an MTT method, the cell proliferation rate is over 90 percent, and the prepared gel has no toxicity to cells.

The dynamic viscosity is 230000 cP-250000 cP at 25 ℃ under the condition that the shear rate is 8.000 l/s-10.000 l/s, correspondingly, the product is extruded from a 25G (0.26 mm inner diameter) needle with the assistance of 64.8N, the injection is smooth, and the product is not layered.

Specific embodiment example 4:

490.28g of calcium nitrate, 409.14g of dipotassium phosphate trihydrate and 1200.00g of injection water are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 180 ℃, kept at the temperature for 0.5h, cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD to be hydroxyapatite. Then dispersing the precipitate into 736g of water for injection, adding 5.00g of hydroxyethyl cellulose, 40.00 g of sodium carboxymethylcellulose, 5.00g of gelatin, 150.00g (10.8%) of glycerol for injection and 6.00g (0.36%) of sodium chloride, mixing and stirring uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving heat for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out a particle size test on the gel by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 30-60 mu m is 80%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity experiment detection is carried out by adopting an MTT method, the cell proliferation rate is over 80 percent, and the prepared gel has no toxicity to cells.

Under the condition that the shear rate is 10.000-15.000 l/s at 25 ℃, the dynamic viscosity is 170000-200000 cP, accordingly, the product is extruded from a needle with the thickness of 25G (0.26 mm inner diameter) by boosting, the boosting force is 43.2N, the injection is smooth, and the product is not layered.

Specific embodiment example 5:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 1200.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 2.0h, cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD to be known as hydroxyapatite. Then dispersing the precipitate into 1066.08g of water for injection, adding 50.00g of sodium carboxymethylcellulose, 200.00g of glycerol for injection (12.21%), 5.00g of sodium citrate (0.244%), stirring and mixing uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving the temperature for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out particle size test on the gel by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 20-60 mu m is 85%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity test detection is carried out by adopting an MTT method, the cell proliferation rate is more than 88 percent, and the result shows that the prepared gel has no toxicity to cells.

Under the condition of shear rate of 16.000-20.000 l/s at 25 ℃, the dynamic viscosity is 100000-150000 cP, the product is extruded by the aid of a needle with the thickness of 25G (0.26 mm inner diameter), the aid is 30.4N, the injection is smooth, and the product is not layered.

Specific embodiment 6:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 1500.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 2.0h, then cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD and is known to be hydroxyapatite. Then dispersing the precipitate into 1479.20g of water for injection, adding 50.00g of sodium carboxymethylcellulose, 200.00g of glycerol for injection (10.15%), 5.00g of sodium citrate (0.20%), stirring and mixing uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving the temperature for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out particle size test on the gel by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 10-60 mu m is 85%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity experiment detection is carried out by adopting an MTT method, and the cell proliferation rate is more than 88%, which indicates that the prepared gel has no toxicity to cells.

Under the condition that the shear rate is 23.000-28.000 l/s at 25 ℃, the dynamic viscosity is 100000-130000 cP, correspondingly, the product is extruded from a 25G (0.26 mm inner diameter) needle with the boosting force of 23.3N, the injection is smooth, and the product is not layered.

Specific embodiment example 7:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 2500.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 2.0h, then cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD and is known to be hydroxyapatite. Then dispersing the precipitate into 2115.86g of water for injection, adding 50.00g of sodium carboxymethylcellulose, 200.00g of glycerol for injection (8.07 percent), 5.00g of sodium citrate (0.16 percent), stirring and mixing uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving the temperature for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Using a laser particle size analyzer to carry out particle size test on the gel to obtain that the volume fraction of particles with the particle size of 10-60 mu m is 85%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity experiment detection is carried out by adopting an MTT method, and the cell proliferation rate is more than 88%, which indicates that the prepared gel has no toxicity to cells.

The dynamic viscosity is 40000cP to 53000cP under the condition of the shear rate of 32.000l/s to 35.000l/s at 25 ℃, the product is extruded by boosting from a needle head with the thickness of 25G (0.26 mm inner diameter), the boosting force is 18.2N, the injection is not smooth, and the product is slightly layered.

Specific embodiment example 8:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 2500.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 2.0h, then cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD and is known to be hydroxyapatite. And dispersing the precipitate into 2115.86g of water for injection, adding 50.00g of sodium carboxymethylcellulose and 200.00g of glycerol for injection (8.08%), stirring and mixing uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving heat for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Using a laser particle size analyzer to carry out particle size test on the gel to obtain that the volume fraction of particles with the particle size of 10-60 mu m is 85%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity test detection is carried out by adopting an MTT method, the cell proliferation rate is more than 88 percent, and the result shows that the prepared gel has no toxicity to cells.

The dynamic viscosity is 40000 cP-53000 cP under the condition that the shear rate is 32.000 l/s-35.000 l/s at 25 ℃, correspondingly, the product is extruded from a needle with the thickness of 25G (0.26 mm inner diameter) by boosting, the boosting force is 17.1N correspondingly, the injection is not smooth, and the product is obviously layered.

Specific embodiment example 9:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 1500.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 2.0h, then cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD and is known to be hydroxyapatite. And dispersing the precipitate into 1066.08g of water for injection, adding 50.00g of sodium carboxymethylcellulose and 200.00g of glycerol for injection (12.24%), stirring and mixing uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving heat for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Using a laser particle size analyzer to carry out particle size test on the gel to obtain that the volume fraction of particles with the particle size of 20-60 mu m is 85%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity experiment detection is carried out by adopting an MTT method, and the cell proliferation rate is more than 88%, which indicates that the prepared gel has no toxicity to cells.

Under the condition of shear rate of 16.000-20.000 l/s at 25 ℃, the dynamic viscosity is 100000-150000 cP, and accordingly, the product is extruded from a needle with the thickness of 25G (0.26 mm inner diameter) by boosting, the boosting force is 30.1N, the injection is smooth, but the product is layered.

Specific embodiment example 10:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 1000.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 2.0h, then cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD and is known to be hydroxyapatite. Then dispersing the precipitate into 329.09g of water for injection, adding 50.00g of sodium carboxymethylcellulose, 200.00g of glycerol for injection (19.06%), 5.00g of sodium citrate (0.379%), stirring and mixing uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving the temperature for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out particle size test on the gel by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 30-60 mu m is 85%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity test detection is carried out by adopting an MTT method, the cell proliferation rate is more than 88 percent, and the result shows that the prepared gel has no toxicity to cells.

Under the condition of a shear rate of 1.800 l/s-3.000 l/s at 25 ℃, the dynamic viscosity is 400000 cP-430000 cP, accordingly, the product is extruded from a needle with the thickness of 25G (0.26 mm inner diameter) by boosting, the boosting force is 85.1N, the injection is difficult, but the product is not layered.

Specific embodiment example 11:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 1000.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, the solution is stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 2.0h, then cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD and is known to be hydroxyapatite. Then dispersing the precipitate into 329.09g of injection water, adding 50.00g of sodium carboxymethylcellulose, 200.00g of injection glycerol (19.06%), 5.00g of sodium citrate (0.379%), stirring and mixing uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving the temperature for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out particle size test on the gel by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 30-60 mu m is 85%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity experiment detection is carried out by adopting an MTT method, and the cell proliferation rate is more than 88%, which indicates that the prepared gel has no toxicity to cells.

The dynamic viscosity is 320000 cP-400000 cP under the condition that the shear rate is 3.200 l/s-4.200 l/s at 25 ℃, the product is extruded by a needle with the thickness of 25G (0.26 mm inner diameter) with the aid of 76.6N, the injection is smooth, and the product is not layered.

Specific embodiment example 12:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 1000.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 2.0h, cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD to be known as hydroxyapatite. And then dispersing the precipitate into 580.00g of water for injection, adding 50.00g of sodium carboxymethylcellulose, 300.00g of glycerol for injection (21.06%), 5.00g of sodium citrate (0.36%), stirring and mixing uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving the temperature for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out particle size test on the gel by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 40-60 mu m is 80%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity experiment detection is carried out by adopting an MTT method, and the cell proliferation rate is more than 88%, which indicates that the prepared gel has no toxicity to cells.

The dynamic viscosity is 60000 cP-140000 cP at 25 ℃ under the condition that the shear rate is 4.000 l/s-6.000 l/s, and correspondingly, the product is extruded from a needle with the thickness of 25G (0.26 mm inner diameter) by boosting force, the corresponding boosting force is 21.5N, the injection is smooth, but the product can be seen to be layered.

Specific embodiment 13:

490.28g of calcium nitrate, 236.75g of diammonium phosphate and 1000.00g of water for injection are weighed, mixed and stirred uniformly, the solution is transferred into a reaction kettle, stirred at the speed of 100r/min, heated to 120 ℃, kept at the temperature for 2.0h, cooled to room temperature, and the precipitate obtained by filtering is analyzed by XRD to be known as hydroxyapatite. And dispersing the precipitate into 680.00g of water for injection, adding 50.00g of sodium carboxymethylcellulose and 200.00g of glycerol for injection (15.07%), stirring and mixing uniformly, pouring the solution into a reaction kettle, stirring the solution at the speed of 100r/min, heating to 100 ℃, preserving heat for 1.0h, and finally cooling to room temperature to obtain the hydroxyapatite gel. Carrying out particle size test on the gel by using a laser particle size analyzer to obtain that the volume fraction of particles with the particle size of 30-60 mu m is 84%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity test detection is carried out by adopting an MTT method, the cell proliferation rate is more than 88 percent, and the result shows that the prepared gel has no toxicity to cells.

The skin injection must meet the following conditions for normal use:

under the condition of a shear rate of 4.000-6.000 l/s at 25 ℃, the dynamic viscosity is 60000-140000 cP, and accordingly, the product is extruded from a needle with the thickness of 25G (0.26 mm inner diameter) by assistance, the assistance force is 21.5N, the injection is smooth, and the product is visually layered. In order to verify the usage amounts of water for injection, a suspending agent and a deflocculant in the preparation process of injection products, and on the basis that the ratio of the amounts of calcium ions and phosphate ions in specific embodiment 2 is 1.67, specific embodiment 5 to specific embodiment 13 are carried out, and for convenience of comparison, the experimental results are plotted in the following table:

as can be seen from the data in embodiments 1 to 4, when the ratio of the amounts of the calcium ion and phosphate ion species in embodiment 2 is about 1.67, the volume fraction of particles having a particle diameter of 30 to 60 μm obtained by cooling the reaction solution is 85%; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity experiment detection is carried out by adopting an MTT method, the cell proliferation rate is more than 88%, in the specific embodiment 3, the using amount of phosphate is greatly increased, and the volume fraction of particles with the particle size of 40-60 mu m is 90% by carrying out the particle size test on the gel by using a laser particle size analyzer; cell culture experiment results show that the obtained gel has better biocompatibility; the cytotoxicity test detection is carried out by adopting an MTT method, the cell proliferation rate is over 90 percent, all indexes of the skin injection filler gel prepared by the test are slightly improved but not greatly improved than those of the concrete embodiment 2, when the quantity ratio of calcium ions to phosphate radical ions in the concrete embodiment 4 is about 1.44, all indexes of the skin injection filler gel prepared by the test are slightly reduced than those of the concrete embodiment 2, so that the preparation process related to the invention can reach a more optimized range when the quantity ratio of the calcium ions to the phosphate radical ions is 1.5-2.0 in terms of finished product performance and raw material input, and the peak value is reached when the quantity ratio of the calcium ions to the phosphate radical ions is 1.65-1.75.

As can be seen from the specific embodiments 1 to 7, the mass fraction of the water for injection is in the range of 30% to 60%. When the proportion of the water for injection is too much, the boosting force is too small, the product is slightly layered, and the normal use is influenced; when the proportion of water for injection is small, the dynamic viscosity is too high, the boosting force is too high, and the normal injection process is influenced. Comparing the specific embodiment 2 and the specific embodiment 11, it can be seen that when the mass fraction of the water for injection is reduced from 15.99% to 4.7%, the viscosity of the product is too high, the injection is difficult, and the use standard is not met.

As can be seen from the specific embodiments 6 to 9, when the injection water and the suspending agent are in large dosage, the demixing is easily caused without adding the deflocculating agent, and the use standard is not met. However, it is clear from the specific examples 4 and 13 that the deflocculant may not be used when the water for injection and the suspending agent are properly formulated.

Comparing the above experimental results, it can be known that the content of the suspending agent and the water for injection can obviously change the dynamic viscosity of the injection product in the using process and has direct influence on the injectability of the product, when the content is too high, the dynamic viscosity of the product is too small, the pushing force during injection is too small, and the product is layered, and at this time, the deflocculant can be properly added to avoid the product layering phenomenon.

According to the hydroxyapatite particle of the present another embodiment, by calcining the hydroxyapatite particle (for example, heating at 800 ℃ for 1 hour), the crystallinity of the particle increases, and the agglomerate of the plurality of primary particles is melted by heating, thereby becoming stronger and more stable. In particular, it is preferable to perform firing by a dispersion firing method, and firing using a flux inhibitor can obtain apatite nanoparticles having high crystallinity and being less likely to aggregate.

By using such calcined apatite particles, a filler having a high effect of stimulating fibroblasts and promoting collagen production can be obtained. That is, by converting hydroxyapatite particles into calcined hydroxyapatite, there is a remarkable effect of improving collagen production promoting effect as compared with that of uncalcined hydroxyapatite. Furthermore, calcined hydroxyapatite has a higher crystallinity and a lower solubility in the body than amorphous hydroxyapatite. Therefore, since the biological activity can be maintained in the living body for a long period of time, it is likely that the collagen production-promoting effect is exhibited for a long period of time.

Calcined hydroxyapatite particles are obtained by calcining amorphous hydroxyapatite. Specifically, for example, a calcined hydroxyapatite can be obtained by calcination by a dispersion calcination method. Further, highly crystalline hydroxyapatite particles having high crystallinity of the hydroxyapatite particles are preferably used.

The lower limit of the firing temperature is more preferably 500 ℃ or higher. If the firing temperature is less than 500 ℃, firing may be insufficient. On the other hand, the upper limit of the firing temperature is more preferably 1800 ℃ or less, still more preferably 1250 ℃ or less, and particularly preferably 1200 ℃ or less. If the firing temperature is higher than 1800 ℃, the hydroxyapatite may be decomposed. Therefore, by setting the firing temperature within the above range, hydroxyapatite which is difficult to dissolve in the living body (high crystallinity) can be produced. The firing time is not particularly limited and may be set as appropriate. Further, the particles may be fused to each other by firing, but in this case, the fired particles may be crushed and used.

Whether or not the hydroxyapatite particles are calcined may be determined by the crystallinity of the particles. The crystallinity of the hydroxyapatite particles may be measured by X-ray diffraction (XRD). It can be said that the narrower the half-peak width of the peak showing each crystal plane, the higher the crystallinity. Specifically, in the calcined hydroxyapatite particles according to the present embodiment, the half-value width of the peak in the 2 θ =32 ° (300) plane in the X-ray diffraction (CuK α ray) is preferably 0.8 or less (more preferably 0.5 or less). Less), highly crystalline hydroxyapatite particles.

Next, the undiluted solution is stirred by applying high shear forces to the agglomerates to wet-pulverize the pulverized hydroxyapatite agglomerates. In addition, "shear force is applied to the agglomerates from multiple directions" herein does not mean a complicated stress occurring in an ordinary mill or the like, applied to the inside of an apparatus for the agglomerates, nor does it mean a complicated stress of the agglomerates. A device is shown that is capable of applying a plurality of different stresses (stresses from different directions) when viewing the system. More specifically, examples of the stirring device for applying a high shearing force include a wet jet mill, a planetary rotary mill, a high-speed homogenizer, and the like, and it is particularly preferable to use a planetary rotary mill.

By using such a pulverization step, the shearing force is sufficient to pulverize the agglomerates under wet conditions, the dispersibility of the pulverized particles in the liquid is improved, and the decrease in crystallinity can be prevented. The comminution results in calcined hydroxyapatite particles. This effect can be further enhanced by making the pulverization method (pulverization apparatus) as described above appropriate. In addition, in the case of the planetary rotary mill, since the equipment used in the mill can be made of a material having high safety such as zirconia, it is suitably used for applications having high safety such as subcutaneous injections.

The treatment conditions in these pulverization steps are not particularly limited, and for example, in the case of a planetary rotary mill, the rotation speed is

At a rotational speed of

The treatment time is

6000. And (3) minutes. More specifically, the wet treatment was performed at rotation speeds of 800rpm and 1600rpm for 180 minutes. By adjusting the pulverization conditions (for example, prolonging the treatment time), the pulverization efficiency can be improved. Further, when the beads are used as a pulverization step as in a planetary rotary mill, the pulverization efficiency can be improved by increasing the weight of the beads to be charged.

The beads used in the planetary rotary mill are not particularly limited as long as the hardness thereof is higher than that of the calcined hydroxyapatite, and are, for example, menou beads (mohs hardness of 8) and zirconia beads (modified). Mohs hardness of 11), tungsten carbide beads (mohs hardness of 12), alumina beads (mohs hardness of 12), and the like. Here, the modified mohs hardness of the calcined hydroxyapatite is usually 5 (that is, the modified mohs hardness of the beads used in the pulverization production method of the present invention is usually more than 5).

Furthermore, it has been found that the crushing efficiency is improved by reducing the bead diameter. From such a viewpoint, for example, the bead diameter is preferably 15mm or less, more preferably 5mm or less, and particularly preferably 0.5mm or less. The lower limit of the bead diameter is not particularly limited and may be appropriately set in accordance with the viscosity of the drug, but if the bead diameter is set to be too small, it may be difficult to remove the bead from the high-viscosity agent. For example, the bead diameter is preferably 0.4 μm or more.

When the pulverization method is a planetary rotary mill, the beads are removed from the composition obtained in the pulverization step. When the beads are removed, the beads may be removed appropriately, but they may also be filtered with a filter having a diameter smaller than that of the beads. Furthermore, beads can also be removed by centrifugation.

By the pulverization production method, a composition containing calcined hydroxyapatite particles having an average particle diameter of 9000nm or less can be obtained. Further, according to the pulverization production method, the variation in crystallinity of the calcined hydroxyapatite particles before and after pulverization is small, and the destruction of crystals is suppressed. That is, by the grinding production method, a calcined hydroxyapatite-containing composition having a small particle size and high crystallinity can be easily obtained from a calcined hydroxyapatite-containing aggregate composition having a large particle size.

The average particle diameter of the hydroxyapatite particles after calcination in the composition obtained by the pulverization production method may be 1000nm or less, 500nm or less, 250nm or less, 125nm or less, and the average particle diameter of the primary particles may be 1nm or less. It will be appreciated that it may be crushed to 50nm or less.

By using the pulverization production method, the crystallinity of the calcined hydroxyapatite particles in the obtained composition can be maintained. That is, the half-value widths a of the peaks of the calcined hydroxyapatite particles (calcined hydroxyapatite particle composition) and the calcined hydroxyapatite particles after the grinding step are within the half-value range of B, and a/B may be 0.9 or more, and may be set to 0.95 or more, with respect to the peak measured by X-ray diffraction (XRD).

After the subcutaneous injection of the present embodiment, first, the hydrogel is absorbed and decomposed. Since the calcined hydroxyapatite particles according to this embodiment have a smaller particle size, the calcined hydroxyapatite particles are released. Collagen production is then promoted by stimulating fibroblasts with calcined hydroxyapatite particles. As described above, since its size is difficult to be recognized as a foreign substance in an organism, it shows an effect of hardly inducing macrophages, and since it has high crystallinity, biological activity can be maintained. Can be existed in organism for a long time. As a result, collagen-rich tissue remains subcutaneous.

On the other hand, when the average particle size of the hydroxyapatite particles is larger than that of the present embodiment, the apatite passively acts. The surface of the apatite particles is gradually covered with collagen under the skin due to biocompatibility, and the apatite is phagocytized by macrophages and covers the collagen layer of the surface due to the size recognized as a foreign matter, and the wrinkle removal effect is continued.

The above-mentioned embodiments only express several embodiments of the present invention, and the description thereof is specific and detailed, but not to be understood as limiting the scope of the present invention. It should be noted that various changes and modifications can be made by those skilled in the art without departing from the spirit of the invention, and these changes and modifications are all within the scope of the invention. Therefore, the protection scope of the present patent shall be subject to the appended claims.

Claims

1. A preparation method of filler gel for skin injection comprises the following steps:

weighing phosphate and calcium salt, adding water for injection into the phosphate and the calcium salt, and uniformly mixing to obtain a solution A;

putting the solution A into a reaction kettle, heating to 80-280 ℃, controlling the stirring speed in the kettle to be 80-800 r/min, and preserving heat for 0.5-10.0 h;

after the reaction is finished, cooling the solution to room temperature to obtain hydroxyapatite crystals;

mixing the obtained hydroxyapatite, suspending agent and anti-flocculation agent with water for injection, and stirring uniformly to obtain solution B;

putting the solution B into a reaction kettle, heating to 80-200 ℃, controlling the stirring speed in the kettle to be 80-800 r/min, and preserving heat for 0.5-5.0 h to obtain hydroxyapatite filler gel;

the hydroxyapatite filler gel is wet-ground by stirring with a stirring device having a high shearing force, and the average particle diameter thereof is 1 μm.

2. The method of preparing a dermal injection filler gel according to claim 1, wherein: the phosphate is one or more of diammonium hydrogen phosphate, dipotassium hydrogen phosphate trihydrate and sodium phosphate.

3. The method of preparing a dermal injection filler gel according to claim 1, wherein: the suspending agent is one or more of methylcellulose, ethyl cellulose, hydroxyethyl cellulose, sodium carboxymethylcellulose, peach gum, gelatin, chondroitin sulfate, sodium alginate, and injectable glycerol.

4. The method of preparing a filler gel for dermal injection according to claim 1, wherein: the calcium salt is one or more of calcium nitrate, calcium chloride, calcium bicarbonate, calcium lactate and calcium gluconate.

5. The method of preparing a dermal injection filler according to claim 1, wherein: the deflocculant is one or more of sodium citrate and sodium chloride.

6. The method of preparing a dermal injection filler according to claim 1, wherein: the ratio of the amount of calcium ion to phosphate ion in the solution A is 1.5-2.0.

7. The method of preparing a dermal injection filler according to claim 1, wherein: the injection water in the solution B accounts for 35-60% by mass.

8. The method of preparing a dermal injection filler according to claim 1, wherein: the mass ratio of the suspending agent in the solution B is 15-20%.

9. The method of preparing a dermal injection filler according to claim 1, wherein: the mass ratio of the deflocculant in the solution B is 0-2%.

10. The method of preparing a dermal injection filler according to claim 1, wherein: the grain size of the calcium hydroxyapatite crystal is within the range of 10-80 μm, and the cooling mode of the solution after heat preservation is natural cooling.