Preparation method of chitosan/silk fibroin microspheres for thrombin immobilization

A technology of thrombin and chitosan, which is applied in the field of medical materials, can solve the problems of poor mechanical properties of silk fibroin and can not achieve the expected effect, and achieve the effect of ensuring the efficiency of vitality recovery and improving the stability of the enzyme

Inactive Publication Date: 2017-05-17
GUANGDONG OCEAN UNIVERSITY
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Problems solved by technology

However, silk fibroin has relatively poor mechanical properties

Method used

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  • Preparation method of chitosan/silk fibroin microspheres for thrombin immobilization
  • Preparation method of chitosan/silk fibroin microspheres for thrombin immobilization

Examples

Experimental program
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Effect test

Embodiment 1

[0025] 1. Preparation of silk fibroin solution

[0026] Take a certain amount of cocoon slices and put them into 0.5% sodium carbonate solution at 98±2°C, the bath ratio is 1:50, and the time is 30 minutes. Take out the degummed silk and rinse it with tap water. Then repeat the operation once more, take out the degummed silk, first rinse it with tap water, then rinse it twice with deionized water, and dry it in an oven at 40°C to obtain degummed silk fibroin fiber.

[0027] Put the obtained silk fiber into CaCl with a bath ratio of 1:10 2 :H 2 O:C 2 h 5 OH=1:8:2 (molar ratio), heating and dissolving at 70°C, and cooling to obtain a brown-yellow silk fibroin solution. Put it into a dialysis bag with a carrying capacity of 8000-14000, dialyze with tap water for two days first, and then dialyze with deionized water for one day. Then, after filtering to remove impurities in the solution, store it in a reagent bottle and put it in a refrigerator for later use.

[0028] 2. Pre...

Embodiment 7

[0036] Add 5 mL each of 100 U / mL heparin sodium solution to the system of Examples 1 to 6, and after 30 minutes of adsorption, centrifuge at 20°C and r3000; take the supernatant and set the volume to 100 mL; take 5.0 mL of each solution in a test tube, Then add 1.0mL barbiturate solution and 1.0mL azure dye solution; the blank group is 5.0mL distilled water plus 1.0mL barbiturate solution and 1.0mL azure dye solution; measure the absorbance at 505nm wavelength, and measure the titer of heparin sodium , to determine its immobilizing enzyme effect.

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Abstract

The invention discloses a preparation method of chitosan/silk fibroin microspheres for thrombin immobilization. The method comprises steps as follows: S1, degummed fibroin fiber is added to an ethanol solution containing calcium chloride to be dissolved, and a fibroin solution is obtained after dialysis and filtration; S2, chitosan is dissolved in an acetic acid solution, the fibroin solution in S1 and a CuSO4 solution are added, and a mixed solution is obtained; S3, the mixed solution in S2 is added to an alkaline ethanol solution, hollow beads are prepared, EDTA is added, complex Cu2<+> is removed through stirring, dehydration, cleaning and drying are performed after filtration, and chitosan/fibroin beads are obtained; S4, chitosan/fibroin beads in S3 are swelled, a thrombin solution and a buffer solution are added, and a glutaraldehyde solution is added for an immobilized enzyme reaction. Chitosan and fibroin are used as carriers for immobilized enzyme, the enzyme stability can be improved, the immobilization activity recovery efficiency is guaranteed, and the method has great actual significance.

Description

technical field [0001] The invention belongs to the technical field of medical materials, and more specifically relates to a preparation method of chitosan / silk fibroin microspheres immobilized with thrombin. Background technique [0002] Thrombin is now widely used in medical research. It is a good hemostatic drug, but due to its poor stability, it has the problem of reduced effectiveness or even inactivation when used. Some researchers are committed to improving the immobilization effect of thrombin, and they hope to find out suitable carriers and storage methods for enzymes to prolong their storage time. In some literatures and reports, it can be seen that the current immobilization method of thrombin includes adding stabilizers glycine and gelatin; exploring the immobilization carrier of enzyme drugs, such as recoagulation sponge, water-soluble chitosan, sodium alginate microcapsules and others immobilization method. [0003] Chitosan is often used as a carrier. It has...

Claims

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Application Information

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IPC IPC(8): A61K38/48A61K9/51A61K47/42A61K47/36A61P7/04
CPCA61K9/5161A61K9/5169A61K38/4833C12Y304/21005
Inventor 苗艳丽李泳谷长生胡章邓春梅陈绍红吴湛霞
Owner GUANGDONG OCEAN UNIVERSITY
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