Amikacin blood collection tube and preparation method thereof

Abstract

The application discloses an amikacin blood collection tube and a preparation method thereof, and relates to the technical field of blood collection tubes. The amikacin blood collection tube comprises a blood collection tube body, an anticoagulant in the tube body and vacuum for maintaining a blood collection negative pressure. The anticoagulant is an ethylenediamine tetraacetic acid (EDTA) salt, and the blood collection tube body further contains amikacin. The amikacin blood collection tube and the preparation method thereof introduce a specific concentration of amikacin as a depolymerizing agent in a conventional EDTA anticoagulation blood collection tube. The amikacin can effectively interfere with the conformational change of a platelet membrane surface GPIIb / IIIa receptor induced by EDTA and the subsequent activation and cross-linking process, thereby significantly inhibiting or eliminating the non-physiological abnormal aggregation of platelets in an in-vitro environment. This makes the blood collection tube maintain the high-efficiency anticoagulation performance of EDTA while fundamentally solving the counting deviation problem caused by EDTA-dependent pseudothrombocytopenia (EDTA-PTCP), and ensures the authenticity and accuracy of platelet detection results.

Description

Technical Field

[0001] This invention relates to the field of blood collection tube technology, specifically to an amikacin blood collection tube and its preparation method. Background Technology

[0002] In the field of clinical testing, the accurate collection and analysis of blood samples is a crucial cornerstone for disease diagnosis and treatment monitoring. Blood collection tubes, as key tools for blood preservation and transportation, directly determine the reliability of subsequent test results through their anticoagulant properties and ability to maintain the stability of blood components. Traditionally, ethylenediaminetetraacetic acid (EDTA) salts have been widely used in routine blood tests due to their potent anticoagulant effect, effectively preventing blood clotting and ensuring the quality of test samples. However, with the continuous improvement of testing accuracy requirements, a specific technical problem has gradually emerged: EDTA-dependent pseudothrombocytopenic purpura (EDTA-PTCP).

[0003] The core of the EDTA-PTCP phenomenon lies in the fact that EDTA can induce conformational changes in GPIIb / IIIa receptors on the platelet membrane surface under in vitro conditions, thereby triggering abnormal platelet activation and cross-linking, ultimately leading to non-physiological platelet aggregation. This process causes platelet counts to deviate significantly from the true values, severely affecting the accuracy of clinical diagnosis, especially in disease states requiring precise assessment of platelet counts, such as hemorrhagic diseases or pre-thrombotic states. Although some studies have attempted to circumvent this problem by adjusting blood collection conditions or using other anticoagulants, these methods often sacrifice the anticoagulant efficiency of EDTA or increase operational complexity, failing to fundamentally solve the counting bias problem caused by EDTA-PTCP. Therefore, improvements are needed. Summary of the Invention

[0004] The purpose of this invention is to provide an amikacin blood collection tube and its preparation method to solve the problem of platelet count deviation caused by EDTA-dependent pseudothrombocytopenia (EDTA-PTCP) in the prior art.

[0005] To achieve the above objectives, the present invention provides the following technical solution: an amikacin blood collection tube, comprising a blood collection tube body, an anticoagulant inside the tube body, and a vacuum to maintain negative pressure during blood collection, wherein the anticoagulant is ethylenediaminetetraacetic acid (EDTA) salt, and the blood collection tube body also contains amikacin.

[0006] Furthermore, the ethylenediaminetetraacetic acid (EDTA) salt is selected from one of EDTA-K2, EDTA-K3, or EDTA-Na2.

[0007] Furthermore, the amount of ethylenediaminetetraacetic acid (EDTA) salt added to the blood collection tube is 1.5-2.2 mg per milliliter of blood collection volume.

[0008] Furthermore, the concentration of amikacin added to the blood collection tube is 0.1-1.0 mg per milliliter of blood collection volume.

[0009] Furthermore, the concentration of amikacin added is 0.2-0.5 mg per milliliter of blood volume.

[0010] Furthermore, the ethylenediaminetetraacetic acid (EDTA) salt and the amikacin are in a dry solid form within the blood collection tube.

[0011] Furthermore, the dried solid form is a uniform film adhering to the inner wall or bottom of the blood collection tube.

[0012] A method for preparing an amikacin blood collection tube includes the following steps:

[0013] S1. Solution preparation: Dissolve ethylenediaminetetraacetic acid (EDTA) salt and amikacin in a solvent to prepare a mixed solution;

[0014] S2. Dispensing and drying: The mixed solution is dispensed into blood collection tubes and then dried to evaporate the solvent and form a solid state inside the tube.

[0015] S3. Sealing and Sterilization: Vacuum the dried blood collection tubes and seal the tube openings for sterilization.

[0016] Furthermore, in step S2, the drying process is freeze drying or low-temperature vacuum drying, with the drying temperature below 40°C.

[0017] Furthermore, in step S3, the sterilization process is gamma-ray irradiation sterilization.

[0018] Compared with existing technologies, the present invention provides an amikacin blood collection tube and its preparation method. By introducing a specific concentration of amikacin as a depolymerizing agent into a conventional EDTA anticoagulant blood collection tube, amikacin can effectively interfere with the conformational changes of GPIIb / IIIa receptors on the platelet membrane surface induced by EDTA and their subsequent activation and cross-linking processes, thereby significantly inhibiting or eliminating non-physiological abnormal platelet aggregation in vitro. This allows the blood collection tube to maintain the highly efficient anticoagulant properties of EDTA while fundamentally solving the counting bias problem caused by EDTA-dependent pseudothrombocytopenia (EDTA-PTCP), ensuring the authenticity and accuracy of platelet test results.

[0019] By pre-positioning EDTA salt and amikacin in a co-dried solid film within the blood collection tube, this solid complex dissolves rapidly and uniformly upon blood injection, achieving simultaneous and immediate onset of anticoagulant and depolymerization functions. This integrated dosage form design eliminates additional clinical procedures (such as adding depolymerizing agents), ensures precise dosage and stability of the active ingredient, and is fully compatible with existing fully automated blood cell analysis procedures, significantly improving the efficiency and standardization of clinical testing.

[0020] By optimizing and limiting the concentration of amikacin (0.1-1.0 mg / mL blood) and the drying process (e.g., freeze-drying, low-temperature vacuum drying), it was ensured that amikacin effectively depolymerized without interfering with other important parameters in routine blood tests (such as white blood cells, red blood cells, and hemoglobin), thus guaranteeing the specificity and safety of the testing system. This design balances the depolymerization effect with the overall integrity of the blood sample, enabling the blood collection tubes to fully meet the quality control requirements for all indicators in routine clinical blood tests while addressing specific interferences. Attached Figure Description

[0021] To more clearly illustrate the technical solutions in the embodiments of this application or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments recorded in this invention. For those skilled in the art, other drawings can be obtained based on these drawings.

[0022] Figure 1 The preparation process flowchart is provided for an embodiment of the present invention. Detailed Implementation

[0023] To enable those skilled in the art to better understand the technical solution of the present invention, the present invention will be further described in detail below with reference to the accompanying drawings.

[0024] As attached Figure 1 As shown:

[0025] Example 1:

[0026] This invention provides an amikacin blood collection tube and its preparation method, including materials and equipment:

[0027] Raw material: Dipotassium ethylenediaminetetraacetate (EDTA-K2), analytical grade, meeting medical standards.

[0028] Amikacin Sulfate, pharmaceutical raw material grade, purity ≥98.5%.

[0029] Sterile water for injection (WFI) meets the standards of the Chinese Pharmacopoeia.

[0030] 3mL transparent glass blood collection tube (preliminarily cleaned).

[0031] Rubber stopper and safety cap for blood collection tubes.

[0032] Reagent: 0.1 mol / L hydrochloric acid solution, used for pH adjustment.

[0033] A 0.1 mol / L sodium hydroxide solution is used for pH adjustment.

[0034] Main equipment: precision electronic balance (sensitivity: 0.01mg), magnetic stirrer and stir bar, pH meter, micropipettes and disposable sterile pipette tips, automatic dispensing system (or precision manual dispensing device), freeze dryer, vacuum sealing machine, cobalt-60 gamma ray irradiation sterilization device.

[0035] Designed for blood collection tubes with a standard blood collection volume of 3 mL.

[0036] Anticoagulant: EDTA-K2. The target anticoagulant concentration is 2.0 mg / mL of blood, therefore the amount added per tube is: 2.0 mg / mL × 3 mL = 6.0 mg.

[0037] Depolymerization agent: Amikacin (calculated as amikacin sulfate). The target concentration is 0.3 mg / mL of blood, therefore the amount added per tube is: 0.3 mg / mL × 3 mL = 0.9 mg.

[0038] Solvent: Sterile water for injection, 0.5 mL, to ensure complete dissolution of solid components and easy dispensing and drying of the solution.

[0039] Preparation process steps:

[0040] Step S1: Precise preparation of the mixed solution

[0041] In a sterile operating table (or a clean environment that meets GMP requirements), accurately weigh 6.0 mg EDTA-K2 and 0.9 mg amikacin sulfate powder using a precision electronic balance.

[0042] Transfer the weighed powder into a clean 2mL glass vial.

[0043] Add 0.5 mL of sterile water for injection to the vial using a micropipette.

[0044] Place the vial on a magnetic stirrer, add a small stir bar, and stir at 300 rpm for about 15 minutes until all the solids are completely dissolved and a clear solution is formed.

[0045] The pH of the mixed solution was measured using a calibrated pH meter. In this example, the initial pH was approximately 8.5. A 0.1 mol / L hydrochloric acid solution was added dropwise using a micropipette while stirring, until the pH was precisely adjusted to 7.2 ± 0.1. This pH value is close to the physiological pH of human blood, which is beneficial for stabilizing the blood condition after collection.

[0046] Label the prepared and pH-adjusted mixed solution for later use.

[0047] Step S2: Dispensing and freeze-drying

[0048] Using an automated dispensing system or a calibrated precision manual pipette, accurately and equally dispense all 0.5 mL of the mixed solution prepared in step S1 into the bottom of 100 3 mL glass blood collection tubes. Ensure that each tube contains 6.0 mg of EDTA-K2 and 0.9 mg of amikacin.

[0049] Place the pre-packaged blood collection tubes horizontally on the material tray of the freeze dryer, ensuring that the solution at the bottom of the tube is spread evenly.

[0050] Start the freeze-drying process:

[0051] Pre-freezing stage: The temperature of the freeze-drying chamber is rapidly reduced to -40°C and maintained at this temperature for 4 hours to allow the solution to completely solidify into solid ice.

[0052] First drying (sublimation) stage: Start the vacuum pump to reduce the pressure inside the chamber to below 10 Pa. Slowly raise the shelf temperature to -20°C and maintain this condition for 24 hours. During this process, the solid ice directly sublimates into water vapor and is removed.

[0053] Secondary drying (desorption) stage: The shelf temperature is slowly raised to 25°C and vacuum (<10Pa) is maintained for 6 hours to remove residual bound water.

[0054] After the drying process is complete, open the freeze-drying chamber under vacuum or nitrogen protection. Inspect each blood collection tube: a uniform, dense, smooth, white or off-white solid film is visible adhering to the bottom of the tube, without crystallization or uneven coloring. This film is firmly bonded to the tube wall and will not detach with slight vibration.

[0055] Step S3: Vacuuming, sealing and terminal sterilization

[0056] The dried blood collection tubes are then transferred to the vacuum packaging production line.

[0057] Using a vacuum sealing machine, a vacuum is drawn into the tube through the rubber stopper at the tube opening. The vacuum level is controlled at a preset negative pressure value that matches the 3mL blood collection volume (usually corresponding to an internal tube pressure of about -40kPa to -60kPa).

[0058] Under vacuum conditions, the aluminum-plastic composite cap is automatically pressed in to seal the tube opening.

[0059] The sealed blood collection tubes are packed into product turnover boxes and sent to the irradiation center.

[0060] The product was placed in a cobalt-60 gamma ray irradiation field and subjected to uniform irradiation at a dose of 25 kGy to ensure that the sterility assurance level (SAL ≤ 10) was achieved. -6 Gamma ray sterilization is a room-temperature process that does not cause thermal decomposition of EDTA or amikacin, thus maximizing the preservation of drug activity.

[0061] After sterilization, the product is sampled and tested for key quality attributes such as sterility, pH (after reconstitution), and fill volume variation. Once qualified, the finished amikacin-containing EDTA-K2 anticoagulant blood collection tube of this invention is obtained.

[0062] The products obtained through the above process have the following distinct characteristics:

[0063] Appearance: A uniform solid film is visible at the bottom of the blood collection tube.

[0064] Contents: Each blood collection tube contains 6.0 mg of EDTA-K2 and 0.9 mg of amikacin sulfate (equivalent to approximately 0.78 mg of amikacin bases).

[0065] Form: The anticoagulant and depolymerizing agent exist as a blended dry solid film.

[0066] Function: When in use, draw venous blood up to the 3mL mark. The blood comes into contact with the solid film at the bottom of the tube and quickly dissolves the EDTA-K2 and amikacin, achieving effective anticoagulation of the blood and simultaneously inhibiting platelet aggregation.

[0067] Example 2:

[0068] This embodiment is basically the same as the previous embodiment, except that four groups of blood collection tubes with different ratios (2A, 2B, 2C, 2D) are designed, respectively covering the lower and upper limits of the EDTA addition amount, the lower and upper limits of the amikacin addition concentration, and a preferred concentration point, to verify the feasibility of the parameter range. All preparation steps refer to the core process of Example 1, with adjustments only made to the formulation measurement.

[0069] Materials and equipment (basically the same as in Example 1; key differences are listed here):

[0070] Raw materials: In addition to EDTA-K2, add EDTA-K3 and EDTA-Na2.

[0071] Blood collection tube specifications: 2mL and 5mL blood collection tubes are used according to design requirements.

[0072] Main equipment: Same as in Example 1, but it is necessary to ensure that the balance and dispensing equipment are accurately adapted to tubes of different specifications.

[0073] The table below shows the detailed formulation design for the four samples (2A to 2D) in this embodiment:

[0074]

[0075] Calculation instructions:

[0076] Group 2A: EDTA-K3 dosage = 1.5 mg / mL × 2 mL = 3.0 mg; Amikacin dosage = 0.1 mg / mL × 2 mL = 0.2 mg.

[0077] Group 2B: EDTA-Na2 addition amount = 2.2mg / mL × 5mL = 11.0mg; Amikacin addition amount = 0.2mg / mL × 5mL = 1.0mg.

[0078] Group 2C: Amikacin dosage = 0.5 mg / mL × 3 mL = 1.5 mg.

[0079] Group 2D: EDTA-K2 dosage = 2.0 mg / mL × 2 mL = 4.0 mg; Amikacin dosage = 1.0 mg / mL × 2 mL = 2.0 mg.

[0080] Preparation process steps

[0081] The general preparation process is the same as in Example 1. The differences between the groups are explained in detail below:

[0082] Step S1: Precise preparation of the mixed solution (performed independently in four batches)

[0083] For each formulation, in separate clean containers, use a precision electronic balance to accurately weigh the corresponding type and mass of EDTA salt and amikacin sulfate according to the table above.

[0084] Add the corresponding volume of sterile water for injection to each container (see table above) to ensure that all components are completely submerged.

[0085] Stir with a magnetic stirrer (300 rpm) until the solid is completely dissolved, forming a clear solution.

[0086] The pH value of each group of solutions was monitored using a pH meter. Since both EDTA salt and amikacin are water-soluble salts, the initial pH value of each group may vary slightly, but the final pH value of each group was precisely adjusted to 7.2±0.1 using 0.1mol / L hydrochloric acid or sodium hydroxide solution.

[0087] The four prepared solutions were labeled as 2A, 2B, 2C, and 2D, respectively, and were to be used later.

[0088] Step S2: Dispensing and freeze-drying

[0089] Packaged in groups:

[0090] Pack component 2A into a 2mL blood collection tube.

[0091] Pack 2B components into 5mL blood collection tubes.

[0092] Package component 2C into a 3mL blood collection tube.

[0093] Pack the 2D components into a 2mL blood collection tube.

[0094] When dispensing, ensure that the volume of solution obtained in each tube contains the total mass of the designed-added solids. For example, each tube in group 2A contains 0.3 mL of solution containing 3.0 mg EDTA-K3 and 0.2 mg amikacin sulfate.

[0095] Drying parameter adjustment:

[0096] All groups used the same freeze-drying process as in Example 1 (pre-freezing: -40℃ / 4h; first drying: -20℃ / 10Pa / 24h; second drying: 25℃ / 10Pa / 6h).

[0097] Since the solution loading volume of Group 2B (5mL tube) is slightly larger (0.8mL), the drying stage can be extended by 2-4 hours to ensure complete sublimation of water. Other groups should be processed according to the standard procedure.

[0098] After drying, the following inspections were conducted: a uniform, dry solid film formed at the bottom of the blood collection tubes in each group. The film in group 2D, due to its highest amikacin content, may have been slightly thicker or darker in color, but it remained uniformly adhered.

[0099] Step S3: Vacuuming, sealing and terminal sterilization

[0100] Group vacuuming: Based on the specifications of the blood collection tubes, different target negative pressure programs are set for the vacuum sealing machine so that the 2mL, 3mL, and 5mL tubes obtain a preset vacuum degree that matches their nominal blood collection volume.

[0101] Under the corresponding negative pressure, seal all pipe openings with aluminum-plastic composite caps.

[0102] All blood collection tubes from all groups were mixed together (because the terminal sterilization method is the same) and sent to the cobalt-60 gamma ray irradiation device.

[0103] Sterilize by uniform irradiation with a dose of 25 kGy.

[0104] After sterilization, samples are taken from each group for testing, including: sterility test, pH value (measured by reconstitution with the corresponding volume of water or buffer), and uniformity of contents (determined by HPLC or other methods to determine EDTA and amikacin content). All test results should meet the predetermined quality standards.

[0105] This embodiment successfully prepared four sets of blood collection tubes with different ratios:

[0106] Group 2A products demonstrated that the combination of the lowest concentration of EDTA (1.5 mg / mL) and the lowest concentration of amikacin (0.1 mg / mL) is feasible, with stable product form, supporting the lower limit of the range.

[0107] Group 2B products demonstrated that the combination of the highest concentration of EDTA (2.2 mg / mL) with a preferred concentration of amikacin (0.2 mg / mL) is feasible and suitable for large-volume (5 mL) blood collection tubes, supporting the upper limit of the range and the application of different EDTA salts.

[0108] Group 2C products: Demonstrates an embodiment using a preferred concentration of amikacin (0.5 mg / mL) in a standard 3 mL tube, which is within the preferred range.

[0109] Group 2D products: demonstrated that the combination regimen using the highest concentration of amikacin (1.0 mg / mL) is feasible, and the products still formed a qualified solid film, supporting the upper limit of the required range.

[0110] Example 3:

[0111] This embodiment is essentially the same as the previous embodiment, except that it uses a vacuum drying method at a mild temperature to directly remove water from the solution and form the desired solid film. This method avoids the freezing and thawing process and may be more suitable for production scenarios that are not sensitive to heat but have specific requirements for ice crystal morphology, or as an alternative to freeze-drying.

[0112] Raw materials: (same as in Example 1)

[0113] Dipotassium ethylenediaminetetraacetate (EDTA-K2), analytical grade.

[0114] Amikacin sulfate, pharmaceutical raw material grade.

[0115] Sterile water for injection (WFI).

[0116] 3mL transparent glass blood collection tube body, rubber stopper and safety cap.

[0117] Reagents: (same as in Example 1) 0.1 mol / L hydrochloric acid solution and sodium hydroxide solution.

[0118] Main equipment:

[0119] Precision electronic balance (sensitivity: 0.01mg), magnetic stirrer and stir bar, pH meter, micropipettes, automatic dispensing system, low-temperature vacuum drying oven (critical equipment, which must be able to accurately control the temperature in the range of room temperature to 60°C and the vacuum degree can reach below 10Pa), vacuum sealing machine, cobalt-60 gamma ray irradiation sterilization device.

[0120] Formula: For ease of comparison with Example 1, this example uses the exact same formula as Example 1:

[0121] Blood collection tube specifications: 3mL.

[0122] Dosage per tube: EDTA-K2 6.0 mg, Amikacin Sulfate 0.9 mg.

[0123] Solvent: 0.5 mL of sterile water for injection.

[0124] Target concentrations: EDTA-K2 2.0 mg / mL blood, amikacin 0.3 mg / mL blood.

[0125] Preparation process steps

[0126] Step S1: Precise preparation of the mixed solution

[0127] (This step is exactly the same as step S1 in Example 1 to ensure that the initial materials are the same.)

[0128] Accurately weigh 6.0 mg EDTA-K2 and 0.9 mg amikacin sulfate into a clean vial.

[0129] Add 0.5 mL of sterile water for injection.

[0130] Stir magnetically until completely dissolved.

[0131] The pH of the mixed solution was precisely adjusted to 7.2 ± 0.1 using a 0.1 mol / L hydrochloric acid solution.

[0132] Labeled solution, ready for dispensing.

[0133] Step S2: Dispensing and Low-Temperature Vacuum Drying (The core distinguishing step in this embodiment)

[0134] Dispensing: Using precision equipment, the prepared mixed solution is precisely dispensed into the bottom of 100 3mL blood collection tubes at a rate of 0.5mL per tube.

[0135] Preparation before drying: Place the dispensed blood collection tubes at an angle of approximately 30° on the special tray of the drying chamber. This operation aims to increase the spreading area of ​​the solution on the tube wall, forming a thinner liquid layer, thereby accelerating the drying rate and making the final solid film distribution more uniform.

[0136] Loading into the box: Quickly transfer the tray into the pre-cleaned low-temperature vacuum drying oven.

[0137] Setting up and running the drying program:

[0138] Stage 1 (Ambient Pressure Preheating): Close the drying oven door, but do not activate the vacuum. Set and stabilize the oven temperature at 30°C (this temperature must be strictly below 40°C, directly supporting claim 9). Maintain this temperature for 30 minutes to allow the solution temperature to rise uniformly, avoiding violent boiling and splashing during subsequent vacuuming.

[0139] Phase Two (Gradient Vacuum Drying): Start the vacuum pump.

[0140] First, the pressure inside the chamber is slowly reduced to approximately 5 kPa (50 mbar) at 30°C and maintained at this condition for 2 hours. During this stage, a large amount of free water is gently evaporated and removed.

[0141] Then, the vacuum level was further increased to <1 kPa (10 mbar) and maintained at 30°C for another 6 hours of drying.

[0142] Phase 3 (Deep Drying and Curing): Under high vacuum (<10 Pa), the temperature inside the chamber is slowly raised to 35°C and maintained at this temperature for 4 hours. This phase aims to remove residual bound water and allow EDTA-K2 and amikacin to form a stable blended solid.

[0143] Program completion and sampling: The total drying program lasts approximately 12.5 hours. After the program is completed, first fill the chamber with filtered dry nitrogen gas to atmospheric pressure, then open the chamber door.

[0144] Drying result inspection: The blood collection tube was removed for examination. A uniform, smooth, translucent to white dense solid film was visible on the lower part of the inclined inner wall of the tube (the original liquid surface area). The film was firmly attached, without cracks, bubbles, or crystalline precipitation, indicating that the solution was successfully dried and solidified under low-temperature vacuum conditions of 30-35℃, and the effective components were evenly distributed.

[0145] Step S3: Vacuuming, sealing and terminal sterilization

[0146] (This step is consistent with step S3 in Example 1 in principle and core parameters)

[0147] The dried blood collection tubes are then transferred to a vacuum sealing machine.

[0148] Vacuum the tube to the preset negative pressure value (approximately -40 to -60 kPa gauge pressure) of the 3 mL blood collection tube.

[0149] The gland is sealed under vacuum.

[0150] The sealed product was placed under a cobalt-60 source and sterilized by 25 kGy gamma irradiation.

[0151] After sterilization, samples are taken for sterility testing, pH testing (after reconstitution), and content uniformity testing. Those that pass the tests are considered finished products.

[0152] The blood collection tube product prepared using this embodiment:

[0153] Appearance and morphology: Similar to the product in Example 1, the tube contains a uniform, dry solid film. Under a microscope, the film morphology may appear denser, unlike the porous and loose structure that may result from freeze-drying, but the functional components are the same.

[0154] Key process parameters support: This embodiment clearly and specifically uses the "low temperature vacuum drying" method and strictly controls the drying temperature at 30-35℃ (clearly below 40℃), which provides direct and specific embodiment support for claim 9, proving that the drying process in this temperature range is feasible and effective.

[0155] Proof of method universality: This embodiment, together with Embodiment 1 (freeze-drying), demonstrates that the preparation method described in claim 8 is not limited to a single drying method. Whether using a "freeze-sublimation" freeze-drying process or a "low-temperature-evaporation" vacuum drying process, the inventive objective of converting a mixed solution of EDTA and amikacin into a stable solid film inside the tube can be achieved.

[0156] Example 4:

[0157] This embodiment is basically the same as the previous embodiment, except that it uses a rotary spray drying technology to directly and evenly spray the solution of the active ingredient onto the inner wall of the blood collection tube, and then dries it with a gentle airflow while rotating, forming a thin layer adhering to the entire inner wall. When using the blood collection tube to collect blood, the blood can come into contact with and mix with the anticoagulant and depolymerizing agent more quickly and evenly.

[0158] Raw materials: (same as in Example 1)

[0159] Dipotassium ethylenediaminetetraacetate (EDTA-K2), analytical grade.

[0160] Amikacin sulfate, pharmaceutical raw material grade.

[0161] Sterile water for injection (WFI).

[0162] 3mL transparent glass blood collection tube body, rubber stopper and safety cap.

[0163] Reagents: (same as in Example 1) 0.1 mol / L hydrochloric acid solution and sodium hydroxide solution.

[0164] Main equipment: precision electronic balance (sensitivity: 0.01mg), magnetic stirrer and stir bar, pH meter, dedicated rotary spray drying system (core equipment, which must include: a shaft that can fix and rotate blood collection tubes at a uniform speed, a precision atomizing spray head, a temperature-controlled air supply drying chamber, and a program control system), vacuum sealing machine, and cobalt-60 gamma ray irradiation sterilization device.

[0165] Formulation and solution preparation: In order to form a solution suitable for spraying, it is necessary to prepare a solution with a slightly higher viscosity and a higher solid content.

[0166] Blood collection tube specifications: 3mL.

[0167] Target dosage per tube: Same as in Example 1, i.e., 6.0 mg EDTA-K2 and 0.9 mg amikacin sulfate.

[0168] Preparation of spraying solution:

[0169] In a clean container, accurately weigh 60.0 mg EDTA-K2 and 9.0 mg amikacin sulfate (this is a batch of 10 vials, used to illustrate the mixing ratio).

[0170] Add only 1.0 mL of sterile water for injection (instead of 10 × 0.5 = 5 mL). This ratio increases the solution concentration, with a solid content of approximately 6.9% (w / v), which helps to form a film quickly after spraying.

[0171] Stir magnetically until completely dissolved; at this point, the solution will be slightly viscous.

[0172] The pH of the solution was precisely adjusted to 7.2 ± 0.1 using a 0.1 mol / L hydrochloric acid solution.

[0173] The prepared high-concentration mixed solution is transferred to the storage tank of the spraying system.

[0174] Preparation process steps:

[0175] Step S1: Pre-treatment and loading of blood collection tubes

[0176] Clean and siliconize the 3mL glass blood collection tube body (if necessary) to improve inner wall wettability and coating adhesion.

[0177] Secure each blood collection tube to the rotating shaft of the rotary spray drying system, ensuring that the tube axis coincides with the rotating shaft and that the tube opening faces the spray head.

[0178] Step S2: Rotary spraying and online drying (the core innovative step of this embodiment)

[0179] System startup: Start the system to stabilize the temperature inside the drying chamber at 40±2℃ and introduce low-humidity air (dew point temperature < -20℃) purified by a high-efficiency particulate air (HEPA) filter.

[0180] Rotation: Start the rotating shaft to make all blood collection tubes rotate at a constant speed of 200 rpm.

[0181] Precision spraying:

[0182] Start the precision spray head and spray the inner wall of each rotating blood collection tube according to the preset program.

[0183] Spraying parameters: spray rate 0.05 mL / tube, spraying time 3 seconds / tube, atomization pressure 0.2 MPa. Through program control, the amount of spraying liquid received by each tube is strictly consistent (containing 6.0 mg EDTA-K2 and 0.9 mg amikacin).

[0184] Under the action of centrifugal force, the atomized droplets evenly cover the entire inner wall, forming an extremely thin wet liquid film.

[0185] Online rotary drying: After spraying, the blood collection tubes continue to be dried in a 40℃, low-humidity hot air (wind speed 0.5m / s) environment.

[0186] Drying time: 8-10 minutes.

[0187] Online weight monitoring or optical monitoring confirmed that the liquid film on the inner wall of the tube had completely dried, forming a solid coating.

[0188] Cooling and unloading: Stop heating, continue to circulate room temperature dry air to cool for 1 minute, then stop rotating and remove the blood collection tube from the shaft.

[0189] Step S3: Vacuuming, sealing and terminal sterilization

[0190] Visual inspection: Inspect the inner wall of each blood collection tube under strong light. A qualified tube should exhibit an extremely uniform, transparent to semi-transparent, slightly glossy, continuous film adhering to the entire cylindrical inner wall, without drip marks, cracks, or uncovered blank spots.

[0191] Vacuum sealing: Transfer the qualified blood collection tubes to the vacuum sealing machine, evacuate to the standard 3mL tube negative pressure value and seal.

[0192] Terminal sterilization: Sterilization is performed using 25 kGy gamma ray irradiation. Due to the extremely thin and uniform coating, the irradiation dose can effectively penetrate, ensuring sterility.

[0193] Quality inspection: Sampling is performed for sterility testing. Special methods (such as rinsing the entire inner wall of the tube with a quantitative solvent and then measuring the solution content) are used to verify the uniformity of EDTA and amikacin content; the results should meet the specified standards.

[0194] The blood collection tube product prepared through this embodiment has the following characteristics:

[0195] Unique form: The active ingredient exists as a uniform, dry film adhering to the entire inner wall of the blood collection tube, which is completely different from the form deposited at the bottom of the tube.

[0196] Process specificity: This morphology was achieved through a specific process combining spin coating and online hot air drying, demonstrating the achievable path of a "uniform film".

[0197] Potential performance advantages: Due to the large and extremely thin coating area, the blood comes into full contact with the coating during the blood collection process, the active ingredients dissolve quickly, the mixing efficiency is high, and platelet aggregation caused by delayed or improper mixing can be minimized.

[0198] Comparative example:

[0199] The product of this invention was compared with that of the conventional EDTA-K2 anticoagulant blood collection tube prepared and used without amikacin, under the same experimental conditions.

[0200] Materials and equipment: (Highly consistent with Example 1 to ensure experimental comparability)

[0201] Raw materials: Dipotassium ethylenediaminetetraacetate (EDTA-K2, analytical grade), sterile water for injection (WFI).

[0202] Consumables: 3mL transparent glass blood collection tube, rubber stopper and safety cap.

[0203] Equipment: Precision electronic balance, magnetic stirrer, pH meter, micropipettes, automatic dispensing system, freeze dryer, vacuum sealing machine, cobalt-60 gamma ray irradiation sterilization device.

[0204] formula:

[0205] Blood collection tube specifications: 3mL.

[0206] Dosage per tube: 6.0 mg EDTA-K2 only.

[0207] Solvent: 0.5 mL of sterile water for injection.

[0208] Target concentration: EDTA-K2 2.0 mg / mL blood.

[0209] Note: This comparative design excludes amikacin and only includes the most commonly used EDTA-K2 anticoagulant in clinical blood routine tests, representing the closest existing technology.

[0210] Preparation process steps: (The preparation process is exactly the same as in Example 1 to eliminate the influence of process differences on the results)

[0211] Step S1: Solution preparation

[0212] In a clean environment, accurately weigh 6.0 mg of EDTA-K2 powder using a precision electronic balance.

[0213] Transfer the powder to a clean vial and add 0.5 mL of sterile water for injection.

[0214] Stir magnetically until completely dissolved.

[0215] The pH of the solution was precisely adjusted to 7.2 ± 0.1 using a 0.1 mol / L hydrochloric acid solution.

[0216] Step S2: Dispensing and freeze-drying

[0217] Dispense the prepared EDTA solution precisely into the bottom of each 3mL blood collection tube at a rate of 0.5mL per tube.

[0218] The freeze-drying procedure is exactly the same as in Example 1:

[0219] Pre-freeze: -40℃, 4 hours.

[0220] Single drying: -20℃, 10Pa, 24 hours.

[0221] Secondary drying: 25℃, 10Pa, 6 hours.

[0222] After drying, a uniform EDTA-K2 dry solid film is formed at the bottom of the tube.

[0223] Step S3: Vacuuming, sealing and terminal sterilization

[0224] Evacuate the drying tube to a standard negative pressure value matching the 3mL blood collection and seal it.

[0225] Sterilization was performed using cobalt-60 gamma irradiation at a dose of 25 kGy.

[0226] After sterilization, samples are taken for sterility inspection and pH testing. Once qualified, they become the comparative product (conventional EDTA-K2 blood collection tubes).

[0227] Comparative experiment: Verification of platelet counting effect

[0228] This experiment aims to simulate a clinical EDTA-PTCP scenario and quantitatively and qualitatively compare the performance differences between the product of this invention and the comparative product.

[0229] Experimental samples:

[0230] Source: Five patients (volunteers, informed consent) who had previously exhibited typical EDTA-PTCP phenomena in routine blood tests were identified through clinical laboratory screening. Diagnostic criteria: A decrease of more than 50% in platelet count (PLT) after blood collection using standard EDTA tubes compared to immediate testing or sodium citrate tube testing, with numerous platelet aggregates visible on microscopic examination of blood smears.

[0231] Blood collection requirements: The blood collection must be completed by the same experienced nurse, using the same lancet, and within the same time period.

[0232] Experimental grouping and methods:

[0233] Experimental group: Blood collection tubes prepared according to Example 1 of this invention (containing EDTA-K2 2.0 mg / mL + amikacin 0.3 mg / mL).

[0234] Control group: Conventional EDTA-K2 blood collection tubes prepared according to this comparative example (containing only EDTA-K2 2.0 mg / mL).

[0235] For each patient, two blood samples were collected simultaneously, one for the experimental group and one for the control group, with the order of blood collection being random.

[0236] Detection and observation:

[0237] After blood collection, both groups of samples were left to stand under the same conditions (room temperature, 22±2℃).

[0238] Time points: Tests were performed at 30 minutes (T1) and 120 minutes (T2) after blood collection. T1 simulated the routine testing time, and T2 was used to observe whether aggregation intensified over time.

[0239] Detection method:

[0240] Fully automated blood cell analyzer test: Using the same instrument (such as the Sysmex XN series) and under the same settings, platelet count (PLT) is measured.

[0241] Blood smear microscopy: Blood smears are prepared immediately after each instrument test, stained with Wright-Giemsa, and examined by two experienced laboratory technicians under oil immersion (1000x) in a double-blind manner, examining at least 20 fields of view to assess platelet aggregation status. Aggregation degree grading:

[0242] Grade 0: No aggregation, platelets are evenly distributed.

[0243] Level 1: Occasionally small clusters of 2-3 platelets are observed.

[0244] Grade 2: Multiple moderate aggregates of 3-10 platelets are visible.

[0245] Level 3: Large aggregates or patches of more than 10 platelets are visible.

[0246] Experimental data recording:

[0247]

[0248] Data Analysis and Conclusions

[0249] Platelet count:

[0250] The PLT values ​​in the control group (without amikacin) were generally severely low (38-112×10). 9 The value of / L is far below the normal reference range, and it shows a further decreasing trend over time (T2).

[0251] The PLT values ​​in the experimental group (including amikacin) returned to normal or near-normal range (172-245×10). 9 The results ( / L) were stable at different time points with minimal variation.

[0252] Statistical analysis: Paired-samples t-test was used to compare the PLT values ​​of the two groups at T1, and the difference was statistically significant (P<0.001).

[0253] Morphological observation:

[0254] The control group's blood smears showed a large number of (grade 2-3) platelet aggregates, which was the direct cause of the instrument's low count.

[0255] In the blood smears of the experimental group, platelet aggregation was significantly inhibited, and under the microscope, scattered individual platelets (grade 0-1) were the main feature, with only occasional very small aggregates.

[0256] in conclusion:

[0257] This comparative model (standard EDTA tubes) accurately reproduces the clinical problems of EDTA-PTCP: leading to pseudo-, severe, and progressive thrombocytopenia.

[0258] Under the same conditions described above, the product of this invention (EDTA tube containing amikacin) can effectively and stably prevent or significantly reduce EDTA-induced platelet aggregation, thereby restoring the platelet count to its true level.

[0259] This comparative experiment, with solid data and intuitive morphological evidence, demonstrates that adding a specific concentration (0.3 mg / mL) of amikacin to EDTA anticoagulant blood collection tubes produces unexpected technical effects and solves the long-standing EDTA-PTCP problem in clinical testing.

[0260] The foregoing has only described certain exemplary embodiments of the present invention by way of illustration. Undoubtedly, those skilled in the art can modify the described embodiments in various ways without departing from the spirit and scope of the present invention. Therefore, the foregoing drawings and descriptions are illustrative in nature and should not be construed as limiting the scope of protection of the claims of the present invention.

Claims

1. An amikacin blood collection tube, comprising a blood collection tube body, an anticoagulant within the tube body, and a vacuum to maintain negative pressure during blood collection, characterized in that, The anticoagulant is ethylenediaminetetraacetic acid (EDTA) salt, and the blood collection tube also contains amikacin.

2. The amikacin blood collection tube according to claim 1, characterized in that, The ethylenediaminetetraacetic acid (EDTA) salt is selected from one of EDTA-K2, EDTA-K3, or EDTA-Na2.

3. The amikacin blood collection tube according to claim 1, characterized in that, The amount of EDTA salt added to the blood collection tube is 1.5-2.2 mg per milliliter of blood collection volume.

4. An amikacin blood collection tube according to claim 1, characterized in that, The concentration of amikacin added to the blood collection tube is 0.1-1.0 mg per milliliter of blood collection volume.

5. An amikacin blood collection tube according to claim 4, characterized in that, The concentration of amikacin added is 0.2-0.5 mg per milliliter of blood volume.

6. An amikacin blood collection tube according to claim 1, characterized in that, The ethylenediaminetetraacetic acid (EDTA) salt and the amikacin are in a dry solid form inside the blood collection tube.

7. An amikacin blood collection tube according to claim 6, characterized in that, The dried solid form is a uniform thin film adhering to the inner wall or bottom of the blood collection tube.

8. A method for preparing an amikacin blood collection tube, applicable to the amikacin blood collection tube according to any one of claims 1 to 7, characterized in that, Includes the following steps: S1. Solution preparation: Dissolve ethylenediaminetetraacetic acid (EDTA) salt and amikacin in a solvent to prepare a mixed solution; S2. Dispensing and drying: The mixed solution is dispensed into blood collection tubes and then dried to evaporate the solvent and form a solid state inside the tube. S3. Sealing and Sterilization: Vacuum the dried blood collection tubes and seal the tube openings for sterilization.

9. The method for preparing an amikacin blood collection tube according to claim 8, characterized in that, In step S2, the drying process is freeze drying or low-temperature vacuum drying, with the drying temperature below 40°C.

10. The method for preparing an amikacin blood collection tube according to claim 8, characterized in that, In step S3, the sterilization process is gamma-ray irradiation sterilization.

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