Blood glucose protection microsphere, preparation method thereof and application of blood glucose protection microsphere in vacuum blood collection tube

Abstract

The invention discloses a blood glucose protection microsphere, a preparation method thereof and application of the blood glucose protection microsphere in a vacuum blood collection tube, and belongs to the technical field of medical instruments. The blood glucose protection microsphere is prepared by the following steps: preparing a solution from an anticoagulant, an anti-glycolysis agent, an excipient and a protective agent in proportion, freezing the solution through liquid nitrogen drop beads to form a microsphere precursor, and performing vacuum freeze drying to prepare a freeze-dried microsphere with the diameter of 2.0-6.0 mm. According to the preparation method, various functional reagents are integrated into the microspheres at one time, the process is simple, and energy conservation and consumption reduction are achieved. The microspheres can be quickly dissolved and uniformly dispersed when being applied to the vacuum blood collection tube, glycolysis is effectively inhibited, hemolysis is prevented, the accuracy and reliability of blood glucose detection are improved, and meanwhile, product quality control is facilitated.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of medical devices, in particular to a blood glucose protection microsphere, a preparation method thereof and application thereof in a vacuum blood collection tube. BACKGROUND

[0002] Blood glucose detection is a key indicator for the clinical diagnosis of diabetes and related metabolic diseases. After blood is taken out of the body, blood cells continue to perform glycolysis, resulting in a decrease in the glucose concentration in the blood over time, thereby affecting the accuracy of the detection results. To inhibit this process, vacuum blood collection tubes with added blood glucose protection agents are commonly used in clinical practice.

[0003] The existing blood glucose protection agents for vacuum blood collection tubes usually contain anticoagulants (such as EDTA-K2, potassium oxalate, heparin) and anti-glycolysis agents (such as sodium fluoride, sodium iodoacetate). The traditional production process is to prepare the anticoagulant and the anti-glycolysis agent into liquid reagents, respectively, and add or spray them to the inner wall of the blood collection tube in sequence, and dry them separately. This step-by-step addition and multiple drying process has significant defects: the production process is complex, with many steps, long time consumption, and high labor cost. The energy consumption is high, and the multiple drying process consumes a large amount of energy. There is a risk of product quality, as the reagents are prone to hydrolysis during preparation and drying, which may cause hemolysis after blood collection; at the same time, the sprayed and dried reagents adhere tightly to the tube wall, which may not be mixed well when inverted, resulting in poor anticoagulation or anti-glycolysis effect. Quality control is difficult, as liquid addition may result in missing or less addition, which is difficult to detect visually. SUMMARY

[0004] The present application aims to provide a blood glucose protection microsphere, a preparation method thereof and application thereof in a vacuum blood collection tube, to solve the problems of complex production process, high energy consumption and easy hemolysis in the prior art.

[0005] To achieve the above-mentioned purpose, the present application adopts the following technical solutions: The present application provides a blood glucose protection microsphere, which is composed of an anticoagulant, an anti-glycolysis agent, an excipient and a protective agent, and is prepared by solution preparation, droplet freezing molding and vacuum freeze-drying to obtain freeze-dried microspheres with a diameter of 2.0-6.0mm.

[0006] The anticoagulant is one of EDTA-K2, potassium oxalate, or heparin; based on per milliliter of blood, the microspheres contain 1.5 mg to 2.5 mg of EDTA-K2, 1.5 mg to 2.5 mg of potassium oxalate, and 12 IU to 25 IU of heparin. The antiglycolytic agent is one of sodium fluoride or sodium iodoacetate; based on per milliliter of blood, the microspheres contain 1.5 mg to 2.5 mg of sodium fluoride and 0.5 mg to 2.5 mg of sodium iodoacetate. The excipient is one or more of trehalose, sucrose, and mannitol; based on the total mass of the raw materials, the excipient content is 2.0% to 10.0%. The protective agent is one or more of PEG, PVP, TW-80, and glycerin; based on the total mass of the raw materials, the protective agent content is 0.2% to 1.0%.

[0007] This invention also provides a method for preparing the blood glucose-protecting microspheres, comprising the following steps: Solution preparation: Dissolve the anticoagulant, antiglycation agent, excipient and preservative in deionized water in proportion and stir until homogeneous; Bead formation: The above solution is dripped into liquid nitrogen using a bead-dropping machine and frozen to form microsphere precursors; Freeze-drying: The microsphere precursor is freeze-dried in a freeze dryer under vacuum to obtain freeze-dried microspheres; Screening and packaging: Microspheres with a particle size of 2.0 to 6.0 mm are screened out.

[0008] The blood glucose-protecting microspheres are dispensed into test tubes to make vacuum blood collection tubes. The number of microspheres added is 1 to 5, or can be precisely added according to the nominal blood collection volume of the vacuum blood collection tube.

[0009] The present invention has the following beneficial effects: (1) The anticoagulant and antiglycation components are mixed at once and made into freeze-dried microspheres, avoiding the multiple addition and drying steps in the traditional process, which greatly simplifies the production process, shortens the production cycle, and significantly reduces energy consumption.

[0010] (2) By adding a protective agent and using a mild freeze-drying process, the hydrolysis of reagents was effectively inhibited, the risk of hemolysis of samples was significantly reduced, and the accuracy of detection was improved.

[0011] (3) The freeze-dried microspheres have a loose and porous structure, which can dissolve quickly and disperse evenly after contact with blood, ensuring the immediacy and uniformity of the anticoagulation and antiglycation effects, and overcoming the problem of poor mixing that may exist in traditional spraying reagents.

[0012] (4) Solid microspheres are easy to count and judge by visual inspection system, which effectively avoids the problem of missing or insufficient addition of liquid reagents, and improves the reliability and consistency of the product.

[0013] (5) By adjusting the number of microspheres or solution concentration, the vacuum blood collection tube with different blood collection volume can be flexibly adapted. BRIEF DESCRIPTION OF DRAWINGS

[0014] Figure 1 The preparation process flow chart of the emergency biochemical microspheres.

[0015] Figure 2 The structure schematic diagram of the emergency biochemical microspheres.

[0016] Figure 3 The structure schematic diagram of the vacuum blood collection tube using the microspheres. DETAILED DESCRIPTION

[0017] The technical solutions in the embodiments of the present application will be clearly and completely described below with reference to the drawings in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments of the present application.

[0018] The information of the equipment and reagents used in the embodiments: Droplet machine: SCIENTZ-DQIR, produced by Ningbo Xinzhi Freeze-drying Equipment Co., Ltd.

[0019] Freeze dryer: Scientz-50F / A, produced by Ningbo Xinzhi Freeze-drying Equipment Co., Ltd.

[0020] EDTA-K2: batch number 20241125, produced by Hubei Xindesheng Material Technology Co., Ltd.

[0021] Sodium fluoride: batch number 241218E1, produced by Xilong Scientific Co., Ltd.

[0022] Potassium oxalate: batch number XL230323, produced by Xilong Scientific Co., Ltd.

[0023] Sodium iodoacetate: batch number 20150206, produced by Nanjing Aodofuni Biological Technology Co., Ltd.

[0024] Sodium heparin: batch number XH241215, produced by Wujiang Hehe Biochemical Products Co., Ltd.

[0025] Trehalose: batch number P3119481, produced by Adamas-beta.

[0026] Sucrose: batch number 180308, produced by Xilong Chemical Co., Ltd.

[0027] Mannitol: batch number 20150206, produced by Shanghai Ruili Spectrum Biological Technology Co., Ltd.

[0028] PEG-2000: batch number 20200522, produced by Shanghai Huishi Laboratory Equipment Co., Ltd.

[0029] PVP (K30): batch number 210107, produced by Xilong Scientific Co., Ltd.

[0030] Glycerol: batch number 250117E1, produced by Xilong Scientific Co., Ltd.

[0031] Example 1 Solution preparation: EDTA-K2 10.0 g, sodium fluoride 10.0 g, sucrose 5.0 g, PVP (K30) 0.5 g were weighed and placed in a triangular flask, about 80 mL of deionized water was added, stirred and dissolved, and heated if necessary to aid dissolution. After cooling to room temperature, it was transferred to a 100 mL volumetric flask, diluted to the mark, shaken and used.

[0032] Droplet formation: the above solution was dropped into liquid nitrogen using a droplet machine. Parameter setting: droplet volume 20 μL / drop, liquid outlet speed 1.5 seconds / drop, rotation speed 144000 rpm. The droplets were instantly frozen in liquid nitrogen to form microsphere precursors.

[0033] Freeze-drying: the microsphere precursors were transferred to a freeze-drier. Program: pre-freezing-50℃ / 2h; sublimation drying:-30℃ / 15h, -15℃ / 1.5h, 0℃ / 1.5h; desorption drying: 10℃ / 1h, 20℃ / 1h.

[0034] Screening and sub-packaging: the dried microspheres were screened, and microspheres with a diameter of 2.5-3.5 mm were selected. Each 12x75 mm test tube was filled with 2 microspheres, vacuumed to 2 mL, and then plugged to prepare a vacuum blood collection tube for collecting 2 mL of blood.

[0035] Example 2 Solution preparation: potassium oxalate 10.0 g, sodium fluoride 10.0 g, mannitol 5.0 g, PVP (K30) 0.5 g were accurately weighed, and 100 mL solution was prepared according to the method of Example 1.

[0036] Droplet formation and freeze-drying: the steps were the same as in Example 1.

[0037] Screening and sub-packaging: microspheres with a diameter of 2.5-3.5 mm were screened, and each test tube was filled with 2 microspheres, vacuumed to 2 mL, and then plugged.

[0038] Example 3 Solution preparation: EDTA-K2 20.0 g, sodium iodoacetate 2.5 g, sucrose 5.0 g, PEG-2000 0.5 g were accurately weighed, and 100 mL solution was prepared according to the method of Example 1.

[0039] Beading and lyophilization: the same as Example 1.

[0040] Screening and packaging: the microspheres with a diameter of 2.5-3.5 mm were screened, and one microsphere was put into each test tube, which was vacuumed to 2 mL and then sealed.

[0041] Example 4 Solution preparation: 0.94 g of sodium heparin (160 IU / mg), 5.0 g of sodium iodoacetate, 8.0 g of sucrose and 0.5 g of glycerol were accurately weighed, and a 100 mL solution was prepared according to the method of Example 1.

[0042] Beading and lyophilization: the same as Example 1.

[0043] Screening and packaging: the microspheres with a diameter of 2.5-3.5 mm were screened, and one microsphere was put into each 12x100 mm test tube, which was vacuumed to 4 mL and then sealed, so as to prepare a vacuum blood collection tube for collecting 4 mL of blood.

[0044] Example 5 Solution preparation: 10.0 g of EDTA-K2, 10.0 g of sodium fluoride, 2.0 g of trehalose, 0.3 g of glycerol and 0.3 g of TW-800 were accurately weighed, and a 100 mL solution was prepared according to the method of Example 1.

[0045] Beading and lyophilization: the same as Example 1.

[0046] Screening and packaging: the microspheres with a diameter of 2.5-3.5 mm were screened, and one microsphere was put into each test tube, which was vacuumed to 2 mL and then sealed.

[0047] To verify the effectiveness of the product of the present example, three healthy volunteers were recruited, who were fasted for more than 10 hours, and blood was collected using the vacuum blood collection tubes prepared according to Examples 1-5 above and a commercially available vacuum blood collection tube produced by a traditional process (batch number: 25010341, as a control group). The blood glucose value of each sample was detected immediately after blood collection, and hemolysis was observed; then the samples were placed in an environment of 25°C for 4 hours and an environment of 2-8°C for 24 hours, respectively, and the blood glucose value was detected again and hemolysis was observed. The detection results are shown in Tables 1 and 2.

[0048] Table 1: Detection results after the samples were placed at 25°C for 4 hours

[0049] As shown in Table 1, there is no obvious difference between the detection results of the control group and the examples after the samples are placed at 25℃ for 4 hours, and the blood glucose reduction rate of all the examples is lower than that of the control group, and the control group has slight hemolysis, and all the examples have no hemolysis. It is shown that the blood glucose protection microspheres of the vacuum blood collection tube of the application are slightly better than the control group (using the conventional spraying and drying process) in protecting the blood glucose in the sample under the condition of being placed at 25℃ for 4 hours.

[0050] Table 2: Detection results of samples placed at 2℃-8℃ for 24 hours

[0051] As shown in Table 2, there is no obvious difference between the detection results of the control group and the examples after the samples are placed at 2℃-8℃ for 24 hours, and the blood glucose reduction rate of all the examples is lower than that of the control group, and the control group has slight hemolysis after being placed at 2℃-8℃ for 24 hours, and all the examples have no hemolysis. It is shown that the blood glucose protection microspheres of the vacuum blood collection tube of the application are slightly better than the control group (using the conventional spraying and drying process) in protecting the blood glucose in the sample under the condition of being placed at 2℃-8℃ for 24 hours.

[0052] The above description is only the preferred embodiment of the application, but the protection scope of the application is not limited to this. Any skilled person in the art can make equivalent replacement, change or modification according to the technical solution and the inventive concept of the application within the technical range disclosed by the application, which should be covered in the protection scope of the application.

Claims

1. A glycemic protection microsphere, characterized in that, The microspheres are prepared by dissolving, bead freezing and vacuum freeze drying of raw materials including an anticoagulant, an anti-glycolysis agent, an excipient and a protective agent, and have a diameter of 2.0-6.0 mm.

2. The blood glucose protection microspheres according to claim 1, characterized in that, The anticoagulant is one of EDTA-K2, potassium oxalate or heparin; the content of EDTA-K2 in the microspheres is 1.5-2.5 mg per ml of blood, the content of potassium oxalate is 1.5-2.5 mg per ml of blood, and the content of heparin is 12-25 IU per ml of blood.

3. The blood glucose protection microspheres according to claim 1, wherein, The anti-glycolysis agent is one of sodium fluoride or sodium iodoacetate; the content of sodium fluoride in the microspheres is 1.5-2.5 mg per ml of blood, and the content of sodium iodoacetate is 0.5-2.5 mg per ml of blood.

4. The blood glucose protection microspheres according to claim 1, wherein, The excipient is one or more of trehalose, sucrose and mannitol; the content of the excipient is 2.0-10.0% based on the total mass of the raw materials.

5. The blood glucose protection microspheres according to claim 1, wherein, The protective agent is one or more of PEG, PVP, TW-80 and glycerol; the content of the protective agent is 0.2-1.0% based on the total mass of the raw materials.

6. A method of preparing the blood glucose protective microspheres of any one of claims 1 to 5, characterized in that, The method comprises the following steps: S1, solution preparation: dissolving the anticoagulant, anti-glycolysis agent, excipient and protective agent in deionized water, stirring and mixing uniformly to obtain a mixed solution; S2, bead forming: dropping the mixed solution obtained in step S1 into liquid nitrogen through a bead machine, and rapidly freezing the droplets to form microsphere precursors; S3, vacuum freeze drying: vacuum freeze drying the microsphere precursors obtained in step S2 to obtain the blood glucose protection microspheres.

7. The method of claim 6, wherein, In step S2, the volume of the droplets is 5-50 μL / drop, and the liquid discharge speed is 1-3 seconds / drop.

8. The method of claim 6, wherein, In step S3, the vacuum freeze drying comprises: a pre-freezing stage at a temperature of -80- -40℃ for 2-6 hours; a sublimation drying stage at a temperature of -40-0℃ for 12-48 hours; and an analytical drying stage at a temperature of 0-60℃ for 2-12 hours.

9. A vacuum blood collection tube, characterized by, It comprises a test tube and at least one blood glucose protection microsphere according to any one of claims 1-5 placed in the test tube, and the test tube is in a vacuum state.

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