Water-insoluble microsphere and water-soluble freeze-dried fiber composition

Abstract

The invention provides a water-insoluble microsphere and water-soluble freeze-dried fiber composition, and relates to the technical field of biological materials. The composition is obtained by mixing water-insoluble microspheres and a water-soluble freeze-dried fiber aqueous solution according to a ratio of (0.005-1) g: 1 ml and then freeze-drying, and the water-soluble freeze-dried fibers are biomacromolecules which are gradually subjected to beta folding and self-assembly gelation after being dissolved in water; and the water-insoluble microspheres are bioabsorbable / degradable microspheres and bionon-absorbable / degradable microspheres. The defects in the prior art are overcome, the filling strength of the freeze-dried microspheres and the characteristics that the water-soluble freeze-dried fibers are good in dispersion and can be self-assembled and gelatinized are considered, and a good redissolving effect is achieved.

Description

Technical Field

[0001] The invention relates to the technical field of biomaterials, and in particular to a water-insoluble microsphere and water-soluble freeze-dried fiber composition. Background Art

[0002] Currently, the biopharmaceutical industry is developing rapidly, and many biomaterials can be prepared into microspheres and freeze-dried fibers for use in tissue filling and embolization therapy.

[0003] Due to the sensitivity of biomaterials to environmental factors such as water and temperature, many biomaterials are prepared into the form of freeze-dried microspheres or freeze-dried fibers for easy storage and transportation. When used, they are re-dissolved or compounded and dispersed in physiological saline before being applied to the human body.

[0004] The existing technology only has products that prepare biological materials into microspheres or freeze-dried fibers. For freeze-dried microspheres, excipients and dispersants such as mannitol are usually added to facilitate clinical use and better achieve dissolution and dispersion effects, but the compounding effect is usually poor, the dispersion is uneven, and the injection performance is poor. The freeze-dried fiber alone is usually a soluble freeze-dried fiber. After re-dissolving and applying it to the human body, it usually has the problem of insufficient filling support. Summary of the invention

[0005] In view of the shortcomings of the prior art, the present invention provides a water-insoluble microsphere and water-soluble freeze-dried fiber composition, which takes into account the filling strength of the freeze-dried microspheres and the good dispersion and self-assembly gelation of the water-soluble freeze-dried fibers, and has a good re-dissolution effect.

[0006] To achieve the above objectives, the technical solution of the present invention is implemented through the following technical solutions:

[0007] A water-insoluble microsphere and water-soluble freeze-dried fiber composition, wherein the composition is obtained by mixing the water-insoluble microsphere and the water-soluble freeze-dried fiber aqueous solution in a ratio of 0.005g-1g:1ml and then freeze-drying, wherein the mass fraction concentration of the water-soluble freeze-dried fiber aqueous solution is 0.01%-20%; the water-soluble freeze-dried fiber is a biomacromolecule that gradually undergoes beta folding and self-assembles gelation after being dissolved in water; the water-insoluble microspheres are bioabsorbable / degradable microspheres and bio-nonabsorbable / degradable microspheres.

[0008] Preferably, the water-soluble freeze-dried fiber is any one of water-soluble silk fibroin, myoglobin, hemoglobin, fibrinogen, and amyloid protein, or a mixture of two or more thereof.

[0009] Preferably, the method for preparing the water-soluble silk fibroin comprises the following steps:

[0010] S1-1, using silkworm cocoons / silk / spider silk as raw materials, and performing degumming treatment with sodium carbonate or sodium bicarbonate solution to obtain degummed silk fibers;

[0011] S1-2, dissolving the degummed silk fibers with a lithium bromide solution or a ternary solvent of calcium chloride-ethanol-water to obtain a degummed silk fiber solution;

[0012] S1-3. The degummed silk fiber solution is dialyzed using a dialysis bag to obtain a silk fibroin aqueous solution and the solution is adjusted to a desired concentration.

[0013] S1-4, freeze-drying the silk fibroin aqueous solution to obtain water-soluble silk fibroin.

[0014] Preferably, the concentration of the sodium carbonate or sodium bicarbonate solution in step S1-1 is 0.5-1.5 g / L, and the degumming method using the sodium carbonate solution is boiling degumming treatment for 0.5-3 h. If the degumming method using the sodium bicarbonate solution is 0.5-5 h of boiling degumming treatment.

[0015] Preferably, when lithium bromide solution is selected in step S1-2, the molar concentration of the lithium bromide solution is 5-12M, and when a ternary system of calcium chloride-ethanol-water is selected as solvent, the molar ratio of calcium chloride, ethanol and water in the system is 1:2:8.

[0016] Preferably, the bioabsorbable / degradable microspheres are any one of polylactide microspheres, aliphatic absorbable polyester microspheres, chitosan microspheres, gelatin microspheres, sodium alginate microspheres, silk fibroin microspheres, collagen microspheres, decellularized matrix microspheres, polyhydroxybutyrate microspheres, polyhydroxyalkanoate microspheres, and polycitric acid microspheres.

[0017] Preferably, the non-bioresorbable / degradable microspheres are any one of polymethyl methacrylate (PMMA) microspheres, polyvinyl alcohol (PVA) microspheres, and glass microspheres.

[0018] Preferably, the particle size of the water-insoluble microspheres is 0.05 microns to 1000 microns.

[0019] Preferably, the method for preparing the water-insoluble microspheres comprises the following steps:

[0020] S2-1, dispersing / dissolving the microsphere raw material in a dichloromethane solution to prepare a solution with a mass fraction of 10% of the microsphere raw material;

[0021] S2-2, adding the above solution to 0.5% PVA aqueous solution for dispersion and emulsification, and then volatilizing dichloromethane to solidify and form, and then air-drying / freeze-drying to prepare water-insoluble microspheres.

[0022] Preferably, the raw material of the microspheres in step S2-1 is any one of polylactide, aliphatic absorbable polyester substances, chitosan, gelatin, sodium alginate, silk fibroin, collagen, decellularized matrix, polyhydroxybutyrate, polyhydroxyalkanoates, polycitric acid, polymethyl methacrylate PMMA, polyvinyl alcohol PVA, and glass.

[0023] The present invention provides a water-insoluble microsphere and water-soluble freeze-dried fiber composition, which has the following advantages over the prior art:

[0024] The present invention provides a water-insoluble microsphere and water-soluble freeze-dried fiber composition, wherein the water-soluble freeze-dried fiber is a material that can undergo beta-folding self-assembly gelation under the induction of temperature, time and other conditions after re-dissolution to form a hydrogel / microgel function. The composition takes into account the filling strength of the freeze-dried microsphere and the characteristics of good dispersion and self-assembly gelation of the water-soluble freeze-dried fiber. The composition can fix and shape the freeze-dried microsphere after freeze-drying, which is convenient for storage. For freeze-dried microspheres and freeze-dried fibers prepared from absorbable materials, the influence of hydrolysis and degradation factors on product performance can be reduced in the freeze-dried state; when the product is re-dissolved for use, the microspheres have good dispersibility, and after being implanted into the human body, under the influence of temperature and time, the silk fibroin solution is beta-folded, and the solution undergoes self-assembly gelation, thereby further improving the filling effect and preventing the microspheres from migrating in human tissues. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 This is the SEM morphology of the composition of Example 1 of the present invention after freeze-drying;

[0026] Figure 2 This is a freeze-dried appearance diagram of the composition of Example 1 of the present invention;

[0027] Figure 3 This is a diagram showing the appearance of the composition of Example 1 of the present invention after reconstitution. DETAILED DESCRIPTION

[0028] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention is clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0029] Embodiment 1:

[0030] Preparation of water-insoluble microspheres and water-soluble freeze-dried fiber composition:

[0031] (1) Extracting silk fibroin from silkworm cocoons to obtain a silk fibroin solution:

[0032] 10g of silkworm cocoons were degummed with 500ml of 1.5g / L sodium carbonate solution for 3 hours to obtain about 6g of degummed silk fibers;

[0033] Take 2 g of degummed silk fiber and dissolve it in 50 ml of 8 M (mol / L) lithium bromide solution to obtain a lithium bromide solution of silk fibroin;

[0034] The lithium bromide solution was dialyzed for 8 hours using a dialysis bag with a molecular weight cutoff of 3000 Dalton to obtain a silk fibroin aqueous solution with a mass dispersion of 4%;

[0035] (2) Preparation of PDLLA racemic polylactic acid microspheres:

[0036] Using PVA as an emulsifier, a 10% dichloromethane solution of PDLLA racemic polylactic acid with a molecular weight of 10,000 is added to a 2% PVA aqueous solution, and the mixture is dispersed and emulsified at a speed of 1,000 r / min, and the dichloromethane is volatilized to solidify the mixture, and non-water-soluble dry PDLLA racemic polylactic acid microspheres are prepared by freeze drying.

[0037] (3) Weighing 100 mg of water-insoluble microspheres and 1 ml of the above-mentioned silk fibroin aqueous solution respectively and mixing them evenly, the obtained mixed system was placed in a -20°C refrigerator for 24 hours, and then placed in a freeze dryer for freeze drying to obtain 140 mg of a water-insoluble microsphere and a water-soluble freeze-dried fiber composition;

[0038] The water-insoluble microspheres and the water-soluble freeze-dried fiber composition are re-dissolved in water and have good overall dispersibility.

[0039] Embodiment 2:

[0040] Preparation of water-insoluble microspheres and water-soluble freeze-dried fiber composition:

[0041] (1) extracting silk fibroin from silkworm cocoons to obtain a silk fibroin solution;

[0042] 10g of silkworm cocoons were degummed by boiling for 5 hours in 500ml of 1.5g / L sodium bicarbonate solution to obtain about 7g of degummed silk fibers;

[0043] Take 2 g of degummed silk fiber and dissolve it in 100 ml of a ternary system solvent of calcium chloride-ethanol-water with a molar ratio of 1:2:8 at a constant temperature of 70-80°C for 3 hours to obtain a fully dissolved and uniform calcium chloride-ethanol solution of silk fibroin.

[0044] The calcium chloride-ethanol solution of the silk fibroin was dialyzed for 24 hours using a dialysis bag with a molecular weight cutoff of 3000 Dalton to obtain a silk fibroin aqueous solution with a mass dispersion of 5%;

[0045] (2) Preparation of PLLA (Levorotatory Polylactic Acid) Microspheres:

[0046] Using PVA as an emulsifier, a 10% dichloromethane solution of PLLA (levorotatory polylactic acid) with a molecular weight of 50,000 is added to a 3% PVA aqueous solution, and the mixture is dispersed and emulsified at a speed of 2000 r / min, and the dichloromethane is volatilized to solidify the mixture, and water-insoluble dry PLLA (levorotatory polylactic acid) microspheres are prepared by air drying or freeze drying.

[0047] (3) 1 ml of 5% silk fibroin solution was mixed with 200 mg of PLLA (levorotatory polylactic acid) microspheres to obtain a mixed system. The mixed system was placed in a -20°C refrigerator for 24 hours and then freeze-dried in a freeze dryer to obtain 250 mg of a water-insoluble microsphere and a water-soluble freeze-dried fiber composition.

[0048] The water-insoluble microspheres and the water-soluble freeze-dried fiber composition are re-dissolved in water and have good overall dispersibility.

[0049] Embodiment 3:

[0050] Preparation of water-insoluble microspheres and water-soluble freeze-dried fiber composition:

[0051] (1) extracting silk fibroin from silkworm cocoons to obtain a silk fibroin solution;

[0052] 10 g of silkworm cocoons were degummed with 500 ml of 1.5 g / L sodium carbonate solution for 3 hours to obtain about 6 g of degummed silk fibers;

[0053] 2 g of degummed silk fibers were taken and dissolved in 50 ml of a lithium bromide solution having a molar concentration of 8 M (mol / L) to obtain a lithium bromide solution of silk fibroin.

[0054] The lithium bromide solution was dialyzed for 8 hours using a dialysis bag with a molecular weight cutoff of 3000 Dalton to obtain a silk fibroin aqueous solution with a mass dispersion of 4%.

[0055] (2) Preparation of PCL polycaprolactone microspheres:

[0056] Using PVA as an emulsifier, a 10% dichloromethane solution of PCL polycaprolactone with a molecular weight of 10,000 is added to a 0.5% PVA aqueous solution, and the mixture is dispersed and emulsified at a speed of 5,000 r / min, and the dichloromethane is volatilized to solidify the mixture, and water-insoluble PCL polycaprolactone microspheres are prepared by air drying or freeze drying.

[0057] (3) 1 ml of 4% silk fibroin solution was mixed evenly with 300 mg of microspheres, and the mixture was placed in a -20°C refrigerator for 24 hours, and then placed in a freeze dryer for freeze drying to obtain 340 mg of a water-insoluble microsphere and water-soluble freeze-dried fiber composition.

[0058] The water-insoluble microspheres and the water-soluble freeze-dried fiber composition are re-dissolved in water and have good overall dispersibility.

[0059] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A water-insoluble microsphere and water-soluble freeze-dried fiber composition, characterized in that: The composition is obtained by mixing water-insoluble microspheres and a water-soluble freeze-dried fiber aqueous solution in a ratio of 0.005g-1g:1ml and then freeze-drying, wherein the mass fraction concentration of the water-soluble freeze-dried fiber aqueous solution is 0.01%-20%; The water-soluble freeze-dried fibers are biomacromolecules that gradually beta-fold and self-assemble into gel after being dissolved in water; the water-insoluble microspheres are bioabsorbable / degradable microspheres and bio-insoluble / degradable microspheres.

2. The composition according to claim 1, characterized in that: The water-soluble freeze-dried fiber is any one of water-soluble silk fibroin, myoglobin, hemoglobin, fibrinogen, and amyloid protein, or a mixture of two or more thereof.

3. The composition according to claim 1, characterized in that: The preparation method of the water-soluble silk fibroin comprises the following steps: S1-1, using silkworm cocoons / silk / spider silk as raw materials, and performing degumming treatment with sodium carbonate or sodium bicarbonate solution to obtain degummed silk fibers; S1-2, dissolving the degummed silk fibers with a lithium bromide solution or a ternary solvent of calcium chloride-ethanol-water to obtain a degummed silk fiber solution; S1-3. The degummed silk fiber solution is dialyzed using a dialysis bag to obtain a silk fibroin aqueous solution and the solution is adjusted to a desired concentration. S1-4, freeze-drying the silk fibroin aqueous solution to obtain water-soluble silk fibroin.

4. The composition according to claim 3, characterized in that: The concentration of the sodium carbonate or sodium bicarbonate solution in step S1-1 is 0.5-1.5 g / L, and the degumming method using the sodium carbonate solution is boiling degumming for 0.5-3 hours. If the degumming method using the sodium bicarbonate solution is 0.5-5 hours of boiling degumming.

5. The composition according to claim 3, characterized in that: When lithium bromide solution is selected in step S1-2, the molar concentration of the lithium bromide solution is 5-12M. When calcium chloride-ethanol-water ternary solvent is selected, the molar ratio of calcium chloride, ethanol and water in the system is 1:2:

8.

6. The composition according to claim 1, characterized in that: The bioabsorbable / degradable microspheres are any one of polylactide microspheres, aliphatic absorbable polyester microspheres, chitosan microspheres, gelatin microspheres, sodium alginate microspheres, silk fibroin microspheres, collagen microspheres, decellularized matrix microspheres, polyhydroxybutyrate microspheres, polyhydroxyalkanoate microspheres, and polycitric acid microspheres.

7. The composition according to claim 1, characterized in that: The bio-non-absorbable / degradable microspheres are any one of polymethyl methacrylate (PMMA) microspheres, polyvinyl alcohol (PVA) microspheres, and glass microspheres.

8. The composition according to claim 1, characterized in that: The particle size of the water-insoluble microspheres is 0.05 microns to 1000 microns.

9. The composition according to claim 1, characterized in that: The preparation method of the water-insoluble microspheres comprises the following steps: S2-1, dispersing / dissolving the microsphere raw material in a dichloromethane solution to prepare a solution with a mass fraction of 10% of the microsphere raw material; S2-2, adding the above solution to 0.5% PVA aqueous solution for dispersion and emulsification, and then volatilizing dichloromethane to solidify and form, and then air-drying / freeze-drying to prepare water-insoluble microspheres.

10. The composition according to claim 9, characterized in that: The raw material of the microspheres in step S2-1 is any one of polylactide, aliphatic absorbable polyester substances, chitosan, gelatin, sodium alginate, silk fibroin, collagen, decellularized matrix, polyhydroxybutyrate, polyhydroxyalkanoates, polycitric acid, polymethyl methacrylate PMMA, polyvinyl alcohol PVA, and glass.

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