Preparation method of antibacterial and absorbable medical suture based on bletilla striata polysaccharide

Abstract

The invention provides a preparation method of an antibacterial and absorbable medical suture based on bletilla striata polysaccharide. The preparation method comprises the following steps: weighing a deacetylated bletilla striata polysaccharide solution, polyethylene glycol and deacetylated bletilla striata polysaccharide microspheres, adding the weighed deacetylated bletilla striata polysaccharide solution, polyethylene glycol and deacetylated bletilla striata polysaccharide microspheres into deionized water, and mixing to obtain a wet spinning solution; spinning and solidifying the wet spinning solution to form filament fibers, and winding the filament fibers on a collecting roller; and performing post-treatment on the filament fibers to obtain the antibacterial absorbable medical suture based on the bletilla striata polysaccharide. According to the medical suture line, a deacetylated bletilla striata polysaccharide solution serves as a base material, deacetylated bletilla striata polysaccharide microspheres serve as a reinforcing phase, the mechanical strength and stability of a fiber aggregate are improved, meanwhile, the dispersity of fibers in the wet spinning process is improved, and the suture line has the extremely good antibacterial performance and can be used for preparing medical suture lines. The absorbable medical suture has a promoting effect on wound healing and repairing, and the mechanical strength meets the strength requirement of the absorbable medical suture 6-0 specification.

Description

Technical Field

[0001] The present invention relates to the technical field of medical suture, and particularly relates to a preparation method of an antibacterial absorbable medical suture based on bletilla striata polysaccharide. Background Art

[0002] Medical suture is a medical device commonly used in surgical operations to connect tissues, suture wounds and ligate blood vessels. Medical sutures are generally divided into two categories: absorbable and non-absorbable. Non-absorbable sutures cannot be absorbed and dissolved by the body; while absorbable sutures can be degraded and absorbed by the human body, so generally no additional suture removal is required. Existing absorbable sutures usually use synthetic materials such as polyester. The monomer fragments generated by the degradation of synthetic materials may cause swelling and local inflammation at the implantation site, and synthetic sutures generally have no promoting effect on wound healing and repair. According to different weaving processes, sutures can also be divided into two structures: single-strand and multi-strand (braided). The single-strand suture has a lower resistance when passing through tissues, is easier to fit, has no gaps and is not easy to hide bacteria, but the single-strand suture has the disadvantages of being more prone to breakage and difficult to tie knots; while the multi-strand (braided) suture is easier to tie knots than the single-strand suture, and has a higher tensile strength, but there are small gaps between the multi-strand materials of the multi-strand suture, providing a space for bacteria to lurk, resulting in a higher probability of infection. All in all, the most basic principle for selecting a suture is: try to use a suture that is thin, has a large tensile force, and has the least reaction to tissues.

[0003] Bletilla striata is a precious traditional Chinese medicine, which is widely used in the field of traditional Chinese medicine. Bletilla striata polysaccharide is neutral, its main component is glucomannan, which is non-toxic and harmless and can be completely absorbed by the human body; moreover, bletilla striata polysaccharide forms a protective film on the wound surface to isolate oxygen, so as to achieve the purpose of hindering the growth of staphylococcus and streptococcus. In addition, bletilla striata polysaccharide has a series of effects such as reducing blood coagulation time, reducing bleeding, and promoting the growth of granulation tissue, and can well promote wound healing. Therefore, bletilla striata can be applied to medical sutures. For example, Patent CN106729944A discloses a fibroin suture with analgesic and anti-inflammatory effects. The suture contains drugs such as bletilla striata powder and other components, and the obtained suture has analgesic and anti-inflammatory effects, but the composition of the suture is complex, and the content of the active ingredient in bletilla striata powder is low, so the efficacy of bletilla striata cannot be fully exerted, which may lead to unstable drug efficacy and limited duration. Patent CN208958880U discloses an absorbable surgical suture. The suture body is woven from absorbable silk thread, and the suture body is coated with a bletilla striata glue layer, which can reduce the trauma of patients, promote wound healing and prevent wound infection, but the preparation method of the suture is relatively complex, the coating layer is easy to fall off, and after the outer coating of bletilla striata glue layer degrades, it can no longer play its role.

[0004] There are currently related technical solutions for preparing Bletilla striata polysaccharide fibers using Bletilla striata polysaccharide as the main raw material. For example, Xiang Yanli's master's thesis "Spinning of Bletilla striata Polysaccharide Fibers and Regulation of Their Water Solubility" from Wuhan Textile University discloses the preparation of Bletilla striata fibers using Bletilla striata polysaccharide as the main raw material. The obtained Bletilla striata fibers have an uneven and non-smooth surface, with a breaking strength of 0.319 - 0.446 cN / dtex and an initial modulus of 5.508 - 7.265 cN / dtex. These Bletilla striata fibers have a relatively high modulus and large stiffness, resulting in poor weaving performance. Therefore, Bletilla striata / polyethylene glycol fibers were also prepared in this thesis. By adding polyethylene glycol, the softness of the Bletilla striata fibers was improved, thereby enhancing their weaving performance. However, this also reduces the strength of the Bletilla striata fibers, making them less suitable for wound suturing as a medical suture. Summary of the Invention

[0005] The present invention provides a method for preparing an antibacterial and absorbable medical suture based on Bletilla striata polysaccharide. This medical suture has excellent antibacterial properties, promotes the healing and repair of wounds, and its mechanical strength meets the strength requirements of the absorbable medical suture size 6-0 specification.

[0006] To solve the above technical problems, the technical solution of the present invention is as follows:

[0007] A method for preparing an antibacterial and absorbable medical suture based on Bletilla striata polysaccharide, comprising the following steps:

[0008] S1. Weigh deacetylated Bletilla striata polysaccharide solution, polyethylene glycol, and deacetylated Bletilla striata polysaccharide microspheres, and add them to deionized water to mix and obtain a wet spinning stock solution; the solid content of the wet spinning stock solution is 16% - 20%, the solid mass ratio of the deacetylated Bletilla striata polysaccharide solution to the deacetylated Bletilla striata polysaccharide microspheres is 1:(0.25 - 4), and the mass of polyethylene glycol is 10% - 20% of the total solid mass of the deacetylated Bletilla striata polysaccharide solution and the deacetylated Bletilla striata polysaccharide microspheres;

[0009] S2. The wet spinning stock solution is spun through a spinneret and coagulated to form filament fibers, which are wound around a collecting roller;

[0010] S3. The filament fibers are post-treated to obtain an antibacterial and absorbable medical suture based on Bletilla striata polysaccharide.

[0011] Preferably, in step S1, the deacetylated Bletilla striata polysaccharide solution and polyethylene glycol are first added to deionized water and stirred evenly, and then the deacetylated Bletilla striata polysaccharide microspheres are added; the deacetylated Bletilla striata polysaccharide solution serves as the matrix material to form the main part of the medical suture. Mixing it well with polyethylene glycol first can better exert the effect of polyethylene glycol in making the matrix material softer; while the deacetylated Bletilla striata polysaccharide microspheres, as the reinforcing material and the dispersed phase, are added later to reduce the consumption of polyethylene glycol.

[0012] Preferably, in step S1, the particle size of the deacetylated Bletilla striata polysaccharide microspheres is 40 - 200 μm.

[0013] Preferably, in step S1, the molecular weight of polyethylene glycol is 4000 - 8000.

[0014] Preferably, in step S2, when spinning, the extrusion rate of the wet spinning dope is 20 - 25 mL / h, the coagulation bath used for coagulation is absolute ethanol, and the collection rate of the filament fiber is 0.5 - 0.8 m / min.

[0015] Preferably, the post - treatment step in step S3 is: remove the filament fiber and the collection roller together and soak them in an absolute ethanol solution, then take them out and perform washing, drying, and disinfection and sterilization treatments in sequence.

[0016] Preferably, the soaking time in the absolute ethanol solution is 2 - 8 h, and the drying temperature is 40 - 60 °C.

[0017] Preferably, the specific preparation method of the deacetylated Bletilla striata polysaccharide solution in step S1 is: using deionized water as a solvent, preparing a 5 - 10 wt% Bletilla striata polysaccharide solution, then adding a 0.1 - 0.5 mol / L NaOH solution and stirring evenly, and the volume ratio of the NaOH solution to the Bletilla striata polysaccharide solution is 1:6.

[0018] Preferably, the Bletilla striata polysaccharide solution is prepared from Bletilla striata polysaccharide powder and deionized water. The preparation method of the Bletilla striata polysaccharide powder is: place the dried Bletilla striata rhizome in deionized water for heating and soaking, collect the Bletilla striata polysaccharide concentrate; add absolute ethanol to the Bletilla striata polysaccharide concentrate, and obtain the Bletilla striata polysaccharide powder through filtration, washing, and drying steps.

[0019] Preferably, the concentration of the collected Bletilla striata polysaccharide concentrate is 4 - 5 wt%; the volume ratio of absolute ethanol to the Bletilla striata polysaccharide concentrate is 3:1 - 4:1, and the drying temperature is 80 °C.

[0020] Preferably, the preparation method of the deacetylated Bletilla striata polysaccharide microspheres in step S1 is: prepare a mixed oil phase with liquid paraffin and 3% Span80 at a mass ratio of (10 - 15):1; add the deacetylated Bletilla striata polysaccharide solution to the mixed oil phase according to an oil - to - water ratio of (5 - 10):1. In a 40 - 50 °C constant temperature water bath, first emulsify for 15 - 20 min, then add 5 - 10 mL of ethylenediamine cross - linker at a dropping rate of 1 - 3 drops / s. After the cross - linker is dropped, continue the reaction for 40 - 50 min, then add 40 - 50 mL of isopropanol for curing for 30 - 35 min. After the reaction is completed, cool to room temperature, then centrifuge to separate the oil phase and the microspheres, and then wash with an organic solvent and deionized water until there is no oil phase on the surface of the microspheres. Freeze - dry the microspheres to obtain the deacetylated Bletilla striata polysaccharide microspheres.

[0021] Compared with the prior art, the beneficial effects of the present invention are as follows: (1) The medical suture provided by the present invention uses deacetylated bletilla striata polysaccharide solution as the main raw material, and polyethylene glycol and deacetylated bletilla striata polysaccharide microspheres are added thereto, significantly improving the mechanical properties of the suture. The prepared suture meets the strength requirements of the absorbable medical suture size 6-0. Moreover, the suture has excellent antibacterial properties and promotes the healing and repair of wounds.

[0022] (2) In the present invention, deacetylated bletilla striata polysaccharide solution is used as the matrix material to form the main part of the medical suture, and deacetylated bletilla striata polysaccharide microspheres are used as the reinforcing phase. The microspheres are interconnected to form a three-dimensional network structure, and bletilla striata fibers penetrate through this network. This network structure can effectively restrict the movement of the fibers, enhance the interaction between the fibers, enable the fibers to better cooperate and deform when stressed, improve the mechanical strength and stability of the fiber aggregate. At the same time, the addition of deacetylated bletilla striata polysaccharide microspheres can also improve the dispersion of the fibers during the preparation of the suture by wet spinning, and improve the uniformity and consistency of the fibers. In addition, the addition of polyethylene glycol can improve the softness of the bletilla striata fibers, which is beneficial to the use of the medical suture in actual operation. Although polyethylene glycol will reduce the strength of the bletilla striata fibers to a certain extent, the addition of deacetylated bletilla striata polysaccharide microspheres improves the overall mechanical strength.

[0023] (3) In the present invention, the acetylation reaction of sodium hydroxide is used to remove the acetyl groups on the surface of the glucomannan macromolecule of bletilla striata, so that the polysaccharide molecules undergo conformational changes and aggregate to form a hydrophobic structure, avoiding the glucomannan from combining with water molecules, swelling and dissolving in water, resulting in a short residence time and poor effect of bletilla striata polysaccharide at the wound. By adjusting the proportion of sodium hydroxide, the degradation rate of bletilla striata polysaccharide is adjusted to meet the degradation time of medical sutures under different requirements.

[0024] (4) When preparing the deacetylated bletilla striata polysaccharide microspheres in the present invention, ethylenediamine reacts with the hydroxyl groups and other groups of bletilla striata polysaccharide to form a stable three-dimensional network structure. This covalent bond can withstand the action of molecular thermal motion in the solution and prevent the molecular chains of the microspheres from dispersing and dissolving. At the same time, the bletilla striata polysaccharide molecule contains a large number of hydroxyl groups, and hydrogen bonds can be formed between molecules. In the microspheres, the molecular chains are closely packed and there are many hydrogen bonds. A large number of hydrogen bonds inside the microspheres bind the molecular chains to each other, ensuring the stability of the microspheres. Description of the Drawings

[0025] Figure 1 is the antibacterial rate of the sutures prepared in Examples 1-3 against Escherichia coli;

[0026] Figure 2 is the antibacterial rate of the sutures prepared in Examples 1-3 against Staphylococcus aureus. Detailed Embodiments

[0027] The present invention will be further described below in conjunction with the accompanying drawings and embodiments.

[0028] Embodiment 1

[0029] This embodiment provides a preparation method of a medical suture based on antibacterial and absorbable Bletilla striata polysaccharide, comprising the following steps:

[0030] S1. Weigh deacetylated Bletilla striata polysaccharide solution, polyethylene glycol, and deacetylated Bletilla striata polysaccharide microspheres, and add them to deionized water to mix and obtain a wet spinning stock solution; specifically, add deacetylated Bletilla striata polysaccharide solution and polyethylene glycol to deionized water, stir evenly, and then add deacetylated Bletilla striata polysaccharide microspheres. The solid content of the wet spinning stock solution is 18%, the solid mass ratio of the deacetylated Bletilla striata polysaccharide solution to the deacetylated Bletilla striata polysaccharide microspheres is 1:0.25, and the mass of polyethylene glycol is 20% of the total solid mass of the deacetylated Bletilla striata polysaccharide solution and the deacetylated Bletilla striata polysaccharide microspheres; preferably, the particle size of the deacetylated Bletilla striata polysaccharide microspheres is about 80 μm. Preferably, the molecular weight of polyethylene glycol is 6000.

[0031] Specifically, the preparation method of the above-mentioned deacetylated Bletilla striata polysaccharide solution is as follows: Take dry Bletilla striata rhizomes, wash to remove surface impurities and pollutants, place them in deionized water, and the mass ratio of Bletilla striata rhizomes to deionized water is 1:5. After standing for 12 h, heat and soak at a constant temperature of 70 °C, and the concentration of the collected Bletilla striata polysaccharide concentrate is 5 wt%; let the Bletilla striata polysaccharide concentrate stand and cool, add anhydrous ethanol (the volume ratio of anhydrous ethanol to the Bletilla striata polysaccharide concentrate is 3:1) to the concentrated filtrate and stir continuously. Filter the standing solution with gauze to obtain a white precipitate; spread the white precipitate flat in a tray and place it in an oven at 80 °C to dry and remove the residual anhydrous ethanol to obtain Bletilla striata polysaccharide powder; take the extracted Bletilla striata polysaccharide powder, use deionized water as a solvent to prepare a 5 wt% Bletilla striata polysaccharide solution, add 0.2 mol / L NaOH solution and stir evenly, and the volume ratio of the NaOH solution to the Bletilla striata polysaccharide solution is 1:6, then the deacetylated Bletilla striata polysaccharide solution is obtained.

[0032] Specifically, the weighed polyethylene glycol is a polyethylene glycol solution, which is prepared with deionized water as a solvent, and the concentration of the polyethylene glycol solution is 4 wt%.

[0033] Specifically, the preparation method of the above-mentioned deacetylated bletilla striata polysaccharide microspheres is as follows: Prepare a mixed oil phase with liquid paraffin and 3% Span80 at a mass ratio of 10:1; Add the deacetylated bletilla striata polysaccharide solution prepared previously to the mixed oil phase according to an oil-water ratio of 10:1. First, emulsify for 15 min in a constant temperature water bath at 45 °C and a rotation speed of 470 rpm, then add 5 mL of ethylenediamine cross-linking agent at a dropping rate of 2 s / drop. After the cross-linking agent is added dropwise, continue the reaction for 45 min, then add 50 mL of isopropanol to solidify for 30 min. After the reaction is completed, cool to room temperature, and then centrifuge (rotation speed 4000 rpm) to separate the oil phase and the microspheres. Then wash with petroleum ether, isopropanol, and deionized water in sequence until there is no oil phase on the surface of the microspheres. Place them at -20 °C for pre-freezing, and finally freeze-dry to obtain deacetylated bletilla striata polysaccharide microspheres with a particle size of about 80 μm.

[0034] S2. Place the wet spinning stock solution in a syringe. The pushing rate of the wet spinning stock solution is 23 mL / h. Extrude it from the spinneret and enter the coagulation bath to form filament fibers. The used coagulation bath is anhydrous ethanol. The filament fibers are wound on the collecting roller under the traction of the roller, and the collecting rate of the filament fibers is 0.6 m / min.

[0035] S3. Remove the filament fibers and the collecting roller together and soak them in an anhydrous ethanol solution for 8 h. Then take them out and wash, dry at 60 °C, and sterilize them in sequence to obtain the antibacterial absorbable medical suture based on bletilla striata polysaccharide.

[0036] Examples 2 - 5

[0037] The preparation methods of the antibacterial absorbable medical sutures provided in Examples 2 - 5 are basically the same as those in Example 1, except that the solid mass ratio of the deacetylated bletilla striata polysaccharide solution to the deacetylated bletilla striata polysaccharide microspheres in the wet spinning stock solution is different. The solid mass ratios of the deacetylated bletilla striata polysaccharide solution to the deacetylated bletilla striata polysaccharide microspheres in Examples 2 - 5 are 3:2, 1:1, 2:3, and 1:4 respectively.

[0038] Comparative Example 1

[0039] The preparation method of the medical suture prepared in Comparative Example 1 is basically the same as that in Example 1, except that no deacetylated bletilla striata polysaccharide microspheres are added in Comparative Example 1. The wet spinning stock solution in Comparative Example 1 is obtained by mixing deacetylated bletilla striata polysaccharide and polyethylene glycol in deionized water. The solid content of the wet spinning stock solution is 18%, and the mass of polyethylene glycol is 20% of the total solid mass of the deacetylated bletilla striata polysaccharide solution.

[0040] Performance Test

[0041] 1. Mechanical Properties

[0042] Using an Instron material testing machine, a simple knot was tied in the middle of the suture. The two ends of the suture were fixed to the two fixed clamps of the material testing machine, with the knot located in the middle of the two fixed clamps. The stretching speed was set at 300 mm / min according to the standard, the test gauge length was 130 mm, and the material testing machine applied tension until the suture broke. The maximum tension value was recorded as the single value of the breaking strength. The test results are shown in the following table.

[0043]

[0044]

[0045] As can be seen from the results in the above table, the breaking strength and breaking elongation of the sutures prepared in Examples 1-5 are greater than those of the suture in Comparative Example 1. This is because in Examples 1-5, deacetylated Bletilla striata polysaccharide microspheres were added to the wet spinning dope. As a filler particle reinforcing material, the deacetylated Bletilla striata polysaccharide microspheres significantly improved the mechanical properties of the deacetylated Bletilla striata polysaccharide sutures, enhancing the toughness and tensile strength of the sutures and providing sufficient mechanical strength for wound suturing. However, when the amount of deacetylated Bletilla striata polysaccharide microspheres added is relatively large, it will also affect the mechanical properties of the suture.

[0046] 2. Antibacterial property

[0047] The sutures prepared in Examples 1-3 were tested for antibacterial activity against Escherichia coli and Staphylococcus aureus. The test method was through the inhibition zone test, and then the data obtained from the experiment was plotted through simulation calculation. The test results of the sutures prepared in Examples 1-3 against Escherichia coli and Staphylococcus aureus are shown respectively as Figure 1 and Figure 2 shown. As can be seen from the figures, the sutures containing deacetylated Bletilla striata polysaccharide have good antibacterial effects against Escherichia coli and Staphylococcus aureus.

[0048] The above examples are only used to illustrate the technical solutions of the present invention and not to limit them. Although the present invention has been described in detail with reference to the preferred embodiments, those of ordinary skill in the art should understand that the technical solutions of the present invention can be modified or equivalently replaced without departing from the spirit and scope of the technical solutions of the present invention.

Claims

1. A method for preparing an antibacterial absorbable medical suture based on Bletilla striata polysaccharide, characterized in that The following steps are involved: S1. Weigh deacetylated Bletilla striata polysaccharide solution, polyethylene glycol, and deacetylated Bletilla striata polysaccharide microspheres, add them into deionized water and mix them to obtain a wet spinning stock solution; the solid content of the wet spinning stock solution is 16%-20%, the solid mass ratio of the deacetylated Bletilla striata polysaccharide solution to the deacetylated Bletilla striata polysaccharide microspheres is 1:(0.25-4), and the mass of the polyethylene glycol is 10%-20% of the total solid mass of the deacetylated Bletilla striata polysaccharide solution and the deacetylated Bletilla striata polysaccharide microspheres; S2, the wet spinning solution is spun and coagulated to form filament fibers, and the filament fibers are wound on a collecting roller; S3. After post-treatment, the filament fibers are used to obtain antibacterial and absorbable medical sutures based on Bletilla striata polysaccharide.

2. The method for preparing the antibacterial absorbable medical suture based on Bletilla striata polysaccharide according to claim 1, characterized in that: In step S1, deacetylated Bletilla striata polysaccharide solution and polyethylene glycol are first added to deionized water, stirred evenly, and then deacetylated Bletilla striata polysaccharide microspheres are added.

3. The method for preparing the antibacterial absorbable medical suture based on Bletilla striata polysaccharide according to claim 1, characterized in that: The particle size of the deacetylated Bletilla striata polysaccharide microspheres in step S1 is 40-200 μm.

4. The method for preparing the antibacterial absorbable medical suture based on Bletilla striata polysaccharide according to claim 1, characterized in that: The molecular weight of the polyethylene glycol in step S1 is 4000-8000.

5. The method for preparing the antibacterial absorbable medical suture based on Bletilla striata polysaccharide according to claim 1, characterized in that: During spinning in step S2, the wet spinning solution ejection rate is 20-25 mL / h, the coagulation bath used during coagulation is anhydrous ethanol, and the filament fiber collection rate is 0.5-0.8 m / min.

6. The method for preparing the antibacterial absorbable medical suture based on Bletilla striata polysaccharide according to claim 1, characterized in that: The post-processing step in step S3 is: taking down the filament fibers and the collecting roller together and soaking them in an anhydrous ethanol solution, and then taking them out and washing, drying and disinfecting them in sequence.

7. The method for preparing the antibacterial absorbable medical suture based on Bletilla striata polysaccharide according to claim 1, characterized in that: The specific preparation method of the deacetylated Bletilla striata polysaccharide solution in step S1 is: using deionized water as solvent, preparing a 5-10wt% Bletilla striata polysaccharide solution, then adding 0.1-0.5mol / L NaOH solution and stirring evenly, the volume ratio of NaOH solution to Bletilla striata polysaccharide solution is 1:

6.

8. The method for preparing the antibacterial absorbable medical suture based on Bletilla striata polysaccharide according to claim 7, characterized in that: The bletilla striata polysaccharide solution is prepared from bletilla striata polysaccharide powder and deionized water. The preparation method of bletilla striata polysaccharide powder is as follows: placing dried bletilla striata stem pieces in deionized water for heating and soaking, collecting bletilla striata polysaccharide concentrated solution; adding anhydrous ethanol to the bletilla striata polysaccharide concentrated solution, and obtaining bletilla striata polysaccharide powder through filtering, washing and drying steps.

9. The method for preparing the antibacterial absorbable medical suture based on Bletilla striata polysaccharide according to claim 8, characterized in that: The concentration of the collected bletilla striata polysaccharide concentrated solution is 4-5wt%; the volume ratio of anhydrous ethanol to the bletilla striata polysaccharide concentrated solution is 3:1-4:1, and the drying temperature is 80°C.

10. The method for preparing the antibacterial absorbable medical suture based on Bletilla striata polysaccharide according to claim 7, characterized in that: The preparation method of deacetylated bletilla striata polysaccharide microspheres in step S1 is as follows: a mixed oil phase is prepared with liquid paraffin and 3% Span80 in a mass ratio of (10-15):1; the deacetylated bletilla striata polysaccharide solution is added to the mixed oil phase in an oil-water ratio of (5-10):1, and the mixture is emulsified for 15-20 minutes in a constant temperature water bath at 40-50°C, and then 5-10 mL of ethylenediamine crosslinking agent is added at a dripping rate of 1-3s / drop, and after the dripping of the crosslinking agent is completed, the reaction is continued for 40-50 minutes, and then 40-50 mL of isopropanol is added for solidification for 30-35 minutes. After the reaction is completed, the mixture is cooled to room temperature, and then centrifuged to separate the oil phase and the microspheres, and then washed with an organic solvent and deionized water until there is no oil phase on the surface of the microspheres, and the microspheres are freeze-dried to obtain deacetylated bletilla striata polysaccharide microspheres.

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