Coagulation-promoting freeze-drying microsphere, preparation method thereof and application of coagulation-promoting freeze-drying microsphere in vacuum blood collection tube

Abstract

The invention discloses coagulation-promoting freeze-dried microspheres, a preparation method thereof and application of the coagulation-promoting freeze-dried microspheres in a vacuum blood collection tube, and belongs to the technical field of medical instruments. The coagulation-promoting freeze-dried microspheres comprise kaolin, coagulation-promoting powder, bovine serum albumin, a dispersing agent and an adhesive, the preparation method comprises the steps of raw material suspension preparation, liquid nitrogen drop bead freezing forming, vacuum freeze drying and microsphere screening. According to the preparation method, the coagulation accelerator is prepared into the form of the freeze-dried microspheres, so that the production process is simplified, the energy consumption is reduced, the biological activity of coagulation accelerating components is effectively protected, the coagulation accelerating effect is quicker, more stable and more uniform, and the product quality of the vacuum blood collection tube and the reliability of clinical examination results are remarkably improved.

Description

Technical Field

[0001] This invention belongs to the field of medical device technology, and particularly relates to a coagulation-promoting freeze-dried microsphere, its preparation method, and its application in vacuum blood collection tubes. Background Technology

[0002] Vacuum blood collection tubes are crucial instruments in clinical testing for the collection, transportation, and storage of blood samples. Their inner walls are typically coated with a coagulant to accelerate blood clotting, facilitating subsequent serum separation for biochemical analysis via centrifugation. Traditional coagulants (such as kaolin, diatomaceous earth, and thrombin) require pre-preparation as a suspension, which is then applied to the inner wall of the blood collection tube via spraying or direct addition, followed by high-temperature drying. This traditional process has several drawbacks: First, it is complex, time-consuming, labor-intensive, energy-intensive, and inefficient. Second, during high-temperature drying, the active ingredients in the coagulant (such as thrombin) are easily denatured and inactivated, affecting its coagulant effect and potentially leading to incomplete serum separation or inaccurate test results. Finally, the dried coagulant is firmly bound to the tube wall; if the tube is not thoroughly mixed after blood collection, it can lead to localized clotting or uneven distribution of the coagulant, affecting the sufficient contact between the blood and the coagulant, ultimately impacting the quality of the test results. Summary of the Invention

[0003] The purpose of this invention is to provide a coagulation-promoting freeze-dried microsphere, its preparation method, and its application in vacuum blood collection tubes, so as to solve the problems of complex production process, easily damaged coagulation accelerator activity, and unstable coagulation effect of traditional vacuum blood collection tubes.

[0004] To achieve the above objectives, the present invention adopts the following technical solution: A type of accelerator freeze-dried microsphere is composed of the following components: kaolin, accelerator powder, bovine serum albumin, dispersant and binder; the accelerator freeze-dried microsphere is prepared by mixing the above components in deionized water to form a suspension, then freezing it by drop beading to form a microsphere precursor, and then freeze-drying it under vacuum to produce freeze-dried microspheres with a diameter of 2.0 to 6.0 mm.

[0005] Preferably, the kaolin has a mesh size of 1250-10000 mesh, and its final effective concentration in blood is 0.01 mg / mL to 0.1 mg / mL. The coagulant is a mixture of rabbit brain powder and snake venom, and its final effective concentration in blood is 0.03 mg / mL to 0.3 mg / mL. The bovine serum albumin is the fifth component, and its mass-volume concentration in the suspension is 0.5% to 5.0%. The dispersant is one or both of Tween-80 and polyvinylpyrrolidone K30, and its total mass-volume concentration in the suspension is 0.2% to 1.0%. The binder is one or both of trehalose and chitosan, and its total mass-volume concentration in the suspension is 0.5% to 5.0%.

[0006] Kaolin and coagulation-promoting powder, as coagulation-promoting components, can activate the activity of thrombin in the blood, thereby promoting blood coagulation. The dispersant promotes the uniform dispersion of kaolin in the suspension. The binder binds with kaolin to form a porous microsphere framework, increasing the mechanical strength and stability of the microspheres, preventing microsphere collapse and dispersion, and maintaining the spherical structure and porosity. Bovine serum albumin is a protein protectant that can form a protective film on the surface of thrombin in the coagulation-promoting powder, preventing denaturation, aggregation, or adsorption to the container surface during freeze-drying, preventing the microspheres from agglomerating and sticking together during preparation and storage, and simultaneously forming a glassy matrix during freeze-drying to protect thrombin activity from damage.

[0007] This invention also provides a method for preparing a coagulation-promoting microsphere vacuum blood collection tube, comprising the following steps: (1) Solution preparation: Weigh kaolin, coagulant powder, bovine serum albumin, dispersant and binder according to the proportion, add deionized water, stir to dissolve and make up to volume to prepare a uniform suspension; (2) Drop freezing: The prepared suspension is dropped into liquid nitrogen at a volume of 5-50 μL / drop through a drop device. The droplets freeze rapidly in the liquid nitrogen to form microsphere precursors; (3) Vacuum freeze drying: The microsphere precursor is transferred to a freeze dryer and subjected to programmed freeze drying to obtain dried coagulation-promoting microspheres; The vacuum freeze drying includes: a pre-freezing stage: temperature -80℃ to -40℃, time 2 to 6 hours; a sublimation drying stage: temperature -40℃ to 0℃, time 12 to 48 hours; and a desorption drying stage: temperature 0℃ to 60℃, time 2 to 12 hours. (4) Microsphere screening: The freeze-dried coagulation-promoting microspheres are sieved to select microspheres with a diameter of 2 to 6.0 mm; (5) Dispensing and packaging: The qualified coagulation microspheres are dispensed into test tubes that have been cleaned and treated on the inner wall. Each test tube contains at least one microsphere. Then, the test tubes are vacuumed and sealed with a stopper to obtain the coagulation microsphere vacuum blood collection tube.

[0008] Compared with the prior art, the present invention has the following beneficial effects: (1) Kaolin, coagulant powder, bovine serum albumin, dispersant and binder are premixed and freeze-dried microspheres are made in one step, eliminating the complicated steps such as spraying and multiple drying in the traditional process. The process is simple and easy to realize automated large-scale production, which significantly improves production efficiency and reduces labor and time costs.

[0009] (2) Vacuum freeze drying technology is used to replace the traditional high-temperature drying process, which greatly reduces energy consumption in the production process and meets the requirements of green manufacturing.

[0010] (3) The freeze-dried microspheres have a loose and porous structure, which can quickly dissolve and release the coagulation-promoting components when in contact with blood, ensuring that the coagulation-promoting agent is quickly and evenly distributed in the blood, avoiding coagulation problems caused by insufficient mixing.

[0011] (4) The vacuum freeze-drying process is carried out at low temperature, which maximizes the protection of the activity of bioactive substances such as thrombin in the coagulation powder, ensuring the stability and reliability of the coagulation effect, thereby improving the accuracy of clinical test results.

[0012] (5) Microspheres are in solid form and are easy to count and detect using visual or automated equipment, which effectively avoids the problems of reagent leakage or inaccurate addition that may occur in traditional liquid spraying, and improves the consistency and controllability of product quality. Attached Figure Description

[0013] Figure 1 This is a schematic diagram of the freeze-dried microspheres for promoting coagulation according to the present invention.

[0014] Figure 2 This is a schematic diagram of the product structure of the vacuum blood collection tube of the present invention.

[0015] Figure 3 This is a schematic diagram of the preparation method of the present invention. Detailed Implementation

[0016] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings of the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments.

[0017] Information on the equipment and reagents used in the examples: Bead-dropping machine: SCIENTZ-DQIR, Ningbo Xinzhi Freeze Drying Equipment Co., Ltd.

[0018] Freeze dryer: Scientz-50F / A, manufactured by Ningbo Xinzhi Freeze Drying Equipment Co., Ltd.

[0019] Kaolin (batch number 20240826), Shandong Xiya Chemical Co., Ltd.

[0020] Coagulant powder (batch number 20250117), Hubei Xindesheng Materials Technology Co., Ltd.

[0021] Trehalose (batch number P3119481), Adamas-beta.

[0022] Chitosan (batch number 20241117), Shanghai Lindi Chemical Co., Ltd.

[0023] Tween-80 (batch number 20150206), Shanghai Rayleigh Biotechnology Co., Ltd.

[0024] Polyvinylpyrrolidone K30 (batch number 240322D1), Xilong Chemical Co., Ltd.

[0025] Bovine serum albumin (batch number 210107), Shanghai Rayleigh Biotechnology Co., Ltd.

[0026] Example 1 like Figure 3 As shown, to prepare a 100mL suspension: weigh 1.0g kaolin, 0.6g coagulant powder, 0.5g Tween-80, 0.5g polyvinylpyrrolidone K30, 1.0g bovine serum albumin, and 0.5g trehalose, place them in an Erlenmeyer flask, add about 80mL of deionized water, stir to dissolve, cool to room temperature, and then bring the volume to 100mL.

[0027] The suspension was dropped into liquid nitrogen using a dropper to freeze into microspheres at a drop volume of 20 μL / drop and a drop rate of 1.5 s / drop. The microsphere precursor was then transferred to a lyophilizer and lyophilized according to the following procedure: pre-freezing (-50℃, 2 h), sublimation drying I (-30℃, 15 h), sublimation drying II (-15℃, 1.5 h), sublimation drying III (0℃, 1.5 h), desorption drying I (10℃, 1 h), and desorption drying II (20℃, 1 h). After lyophilization, lyophilized microspheres with diameters of 2.5 mm to 3.5 mm were selected. Figure 1 As shown.

[0028] One lyophilized microsphere was dispensed into a 12×100mm test tube, a 5mL vacuum was drawn, and the tube was stoppered to prepare a vacuum blood collection tube for collecting 5mL of blood. Figure 2 As shown.

[0029] Calculations show that the concentration of kaolin in the blood in the blood collection tube prepared in this embodiment is 0.04 mg / mL, and the concentration of coagulant is 0.024 mg / mL.

[0030] Example 2 The method is basically the same as in Example 1, except that the adhesive trehalose is replaced with an equal amount of chitosan (0.5g). Calculations show that the concentration of kaolin in the blood in the blood collection tube prepared in this embodiment is 0.04 mg / mL, and the concentration of coagulant is 0.024 mg / mL.

[0031] Example 3 Prepare 100 mL suspension: Weigh 2.0 g kaolin, 1.0 g coagulant powder, 0.5 g Tween-80, 0.5 g polyvinylpyrrolidone K30, 0.5 g bovine serum albumin, and 0.5 g chitosan. Subsequent dropper, lyophilization, screening, and dispensing steps are the same as in Example 1. A vacuum blood collection tube for collecting 5 mL of blood is obtained, with a kaolin concentration of 0.08 mg / mL and a coagulant powder concentration of 0.04 mg / mL.

[0032] Example 4 Prepare 100 mL suspension: Weigh 0.9 g kaolin, 0.6 g coagulant powder, 0.5 g polyvinylpyrrolidone K30, 1.0 g bovine serum albumin, and 0.5 g chitosan (without Tween-80). Subsequent dropper, lyophilization, and screening steps are the same as in Example 1. Dispense one microsphere into a 12 × 75 mm test tube, draw a 3 mL vacuum, and stopper to obtain a vacuum blood collection tube for collecting 3 mL of blood. The calculated concentration of kaolin is 0.06 mg / mL, and the concentration of coagulant powder is 0.04 mg / mL.

[0033] Example 5 Prepare 100 mL suspension: Weigh 0.9 g kaolin, 0.6 g coagulant powder, 0.4 g polyvinylpyrrolidone K30, 0.4 g Tween-80, 2.0 g bovine serum albumin, 0.3 g trehalose, and 0.3 g chitosan. Adjust the droplet volume to 40 μL / drop. After lyophilization, screen for microspheres with a diameter of 3.0 mm to 4.0 mm. Dispense into 12 × 100 mm test tubes, draw a 5 mL vacuum, and prepare vacuum blood collection tubes for collecting 5 mL of blood. The calculated concentration of kaolin is 0.072 mg / mL, and the concentration of coagulant powder is 0.048 mg / mL.

[0034] To verify the effectiveness of the product in this embodiment, six healthy volunteers were recruited to collect blood using vacuum blood collection tubes prepared in Examples 1-5 above and commercially available vacuum blood collection tubes produced using conventional processes (batch number: 25010713, used as a control group). Six tubes were collected from each volunteer, for a total of 36 samples. All samples were mixed and centrifuged under the same conditions, and the supernatant serum was sent to a third-party testing laboratory for analysis of over 40 biochemical indicators, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood glucose (GLU), and total cholesterol (TCHO).

[0035] The test results show that in all test items, the deviation between the test results of each embodiment group and the control group of the present invention is less than 10%, with no statistically significant difference. This proves that the provided coagulation-promoting freeze-dried microsphere vacuum blood collection tube has the same performance as traditional products while ensuring the accuracy of test results, and has advantages in production process, energy consumption and quality control.

[0036] The above description is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A type of freeze-dried microsphere that promotes coagulation, characterized in that, It is made from raw materials including kaolin, coagulant powder, bovine serum albumin, dispersant and binder; the coagulant freeze-dried microspheres are freeze-dried microspheres with a diameter of 2.0 to 6.0 mm, which are made by preparing the raw materials into a suspension, and then freezing and vacuum freeze-drying them.

2. The freeze-dried microspheres for accelerating coagulation according to claim 1, characterized in that, The kaolin has a mesh size of 1250~10000 mesh, and its final effective concentration in blood is 0.01mg / mL~0.1mg / mL.

3. The freeze-dried microspheres for accelerating coagulation according to claim 1, characterized in that, The coagulant powder is a mixture of rabbit brain powder and snake venom, and its final effective concentration in the blood is 0.03 mg / mL to 0.3 mg / mL.

4. The freeze-dried microspheres for accelerating coagulation according to claim 1, characterized in that, The bovine serum albumin concentration in the suspension is 0.5% to 5.0% by mass and volume.

5. The lyophilized microspheres for accelerating coagulation according to claim 1, characterized in that, The dispersant is one or both of Tween-80 and polyvinylpyrrolidone K30, and its total mass-volume concentration in the suspension is 0.2% to 1.0%.

6. The freeze-dried microspheres for accelerating coagulation according to claim 1, characterized in that, The adhesive is one or both of trehalose and chitosan, and its total mass-volume concentration in the suspension is 0.5% to 5.0%.

7. A method for preparing accelerated freeze-dried microspheres according to any one of claims 1 to 6, characterized in that, Includes the following steps: Step 1: Dissolve kaolin, coagulant powder, bovine serum albumin, dispersant and binder in deionized water to prepare a suspension; Step 2: The suspension is dropped into liquid nitrogen for droplet freezing to form microsphere precursors; Step 3: The microsphere precursor is subjected to vacuum freeze-drying to obtain the coagulation-promoting microspheres.

8. The method for preparing lyophilized microspheres according to claim 7, characterized in that, In step 2, the drop volume is 5–50 μL / drop.

9. The method for preparing lyophilized microspheres according to claim 7, characterized in that, In step 3, the vacuum freeze-drying includes: Pre-freezing stage: temperature -80℃ to -40℃, time 2 to 6 hours; Sublimation drying stage: temperature -40℃~0℃, time 12~48 hours; Analysis and drying stage: temperature is 0℃~60℃, time is 2~12 hours.

10. A coagulation-promoting microsphere vacuum blood collection tube, characterized in that, The test tube includes a test tube and at least one lyophilized microsphere as described in any one of claims 1 to 6 sealed inside the test tube, wherein the test tube is in a vacuum state.

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