A method for preparing a calcite calcium carbonate nano-delivery platform

Abstract

The present invention discloses a method for preparing a calcite calcium carbonate nano-delivery platform. Copper ions and anti-tumor drugs are added to a calcium chloride (CaCl2) solution at the same time. No further treatment is required. The delivery platform can be prepared by utilizing carbon dioxide (CO2) generated by the decomposition of ammonium bicarbonate (NH4HCO3) in the external environment to diffuse into the reaction solution. The method is simple and does not require any post-treatment. Anti-tumor drugs and copper ions can be directly loaded into calcite calcium carbonate nanoparticles, and large-scale mass production can be achieved. The prepared nano-delivery platform has pH and GSH dual responsiveness and tumor targeting capabilities, thereby reducing the systemic toxic side effects of anti-tumor drugs and can also utilize Cu 2+ The reaction with the high content of GSH in the tumor produces a large amount of reactive oxygen, which further enhances the effect of tumor treatment. This method of constructing a nano drug delivery system is of great significance for achieving targeted treatment of tumors.

Description

Technical Field

[0001] The invention relates to the fields of material technology and biomedical engineering technology, and in particular to a method for preparing a calcite calcium carbonate nano-delivery platform for loading anti-tumor drugs and copper ions. Background Art

[0002] Malignant tumors (cancer) are diseases caused by malignant cell proliferation, and their probability of occurrence increases with the age of the individual.

[0003] Doxorubicin (DOX) is a traditional broad-spectrum chemotherapy drug that has been proven to have good anti-cancer effects. However, this anti-tumor drug does not have the ability to target tumors, will produce obvious toxic side effects, and has obvious killing ability on normal cells. The toxic side effects of this chemotherapy drug also increase the difficulty of tumor treatment. Copper ions also have certain anti-tumor effects. They can react with GSH in tumor cells to generate reactive oxygen species (ROS), thereby killing tumor cells. It has become an auxiliary choice for anti-tumor drugs to enhance the anti-tumor effect. In order to reduce the toxic side effects of anti-tumor drugs and achieve the simultaneous delivery of anti-tumor drugs and copper ions to improve the anti-tumor effect of anti-tumor drugs, the development of a targeted and responsive anti-tumor drug and copper ion co-delivery platform is currently a hot topic in tumor treatment.

[0004] After searching, the Chinese patent application number CN110302178A discloses a drug-loaded delivery nanoplatform and its preparation method and application. It mentions a scheme for preparing the nano-delivery platform by loading the gene miRNA21i and the anticancer drug DOX.

[0005] However, calcium carbonate nanomaterials are widely used in the delivery of antitumor drugs due to their low toxicity, good biocompatibility and pH responsiveness. The calcium carbonate carriers currently used to deliver antitumor drugs are mainly vaterite and amorphous calcium carbonate. The methods for preparing these two calcium carbonate carriers are relatively complex and require special environmental conditions. For example, the preparation of amorphous calcium carbonate generally requires vacuuming and synthesis under an external vacuum environment. At the same time, the process of loading drugs on these two carriers is a two-step process, that is, the first step is to synthesize calcium carbonate, and the second step is to use calcium carbonate to adsorb antitumor drugs. This post-adsorption process has a weak ability to bind antitumor drugs and is easy to fall off. This two-step synthesis further increases the complexity of the preparation process and the difficulty of subsequent large-scale production. In particular, the current calcium carbonate drug carrier mainly uses its pH responsiveness to achieve the release of drugs, and relative to the various obvious characteristics of the tumor microenvironment, such as a high GSH environment, calcium carbonate is not responsive, which seriously limits the application of calcium carbonate drug carriers in the field of drug delivery. Therefore, by improving the calcium carbonate synthesis method to achieve multiple responses to the tumor microenvironment, the practical application ability of calcium carbonate carriers can be further increased, and it is promoted to actual clinical application. Summary of the invention

[0006] The purpose of the present invention is to solve the defects in the prior art and to propose a method for preparing a calcite calcium carbonate nano-delivery platform.

[0007] In order to achieve the above object, the present invention adopts the following technical solutions:

[0008] A method for preparing a calcite calcium carbonate nano-delivery platform comprises the following steps:

[0009] Step 1: Dissolve 100-200 mg CaCl2, 5-25 mg CuCl2, and 1-10 mg DOX powder in 10-200 mL anhydrous ethanol solution, stir and mix continuously at a speed of 100-600 r / min, and place the mixed solution in a blue-capped bottle;

[0010] Step 2: Take 5-20g of NH4HCO3 solid and place it in a petri dish;

[0011] Step 3: Place the blue-cap bottle containing the mixed solution and the culture dish containing NH4HCO3 in a desiccator, and place the desiccator in an environment of 25-37°C to react for 10-24 hours. After the reaction is completed, a mixture for a calcite calcium carbonate nano-delivery platform loaded with anti-tumor drugs and copper ions is prepared;

[0012] Step 4: Use a centrifuge to separate the mixture formed in step 3, centrifuge at a certain speed and then vacuum dry at 30-60° C. to obtain a calcite calcium carbonate nano-delivery platform loaded with anti-tumor drugs and copper ions.

[0013] Further, the following steps are included:

[0014] Add 150 mg of CaCl2 powder into a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and stir continuously at 500 r / min to mix evenly and dissolve completely;

[0015] Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish. Place the blue-capped bottle (open) and the Petri dish containing NH4HCO3 (open) in a desiccator. Place the desiccator in a 37°C oven and react for 18 h.

[0016] After the reaction, centrifuge at 12000 r / min for 10 min and discard the supernatant;

[0017] The precipitate was washed twice with anhydrous ethanol and dried at 25°C to obtain CaCO3.

[0018] Further, the following steps are included:

[0019] Add 150 mg of CaCl2 powder and 10 mg of CuCl2 into a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and stir continuously at 500 r / min to mix evenly and completely dissolve;

[0020] Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish. Place the blue-capped bottle (open) and the Petri dish containing NH4HCO3 (open) in a desiccator. Place the desiccator in a 37°C oven and react for 18 h.

[0021] After the reaction was completed, the mixture was immediately centrifuged at 12000 r / min for 10 min at room temperature, and the supernatant was discarded;

[0022] The precipitate was washed twice with anhydrous ethanol and dried at 37 °C to obtain Cu@CaCO3

[0023] Further, the following steps are included:

[0024] Add 150 mg of CaCl2 powder and 4 mg of DOX into a blue-capped bottle containing 100 mL of anhydrous ethanol solution, stir continuously at 500 r / min to mix evenly, and use ultrasound to help dissolve;

[0025] Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish. Place the blue-capped bottle (open) and the Petri dish containing NH4HCO3 (open) in a desiccator. Place the desiccator in a 37°C oven and react for 18 h.

[0026] After the reaction was completed, the mixture was immediately centrifuged at 12000 r / min for 10 min at room temperature, and the supernatant was discarded;

[0027] The precipitate was washed twice with anhydrous ethanol and dried at 40 °C to obtain DOX@CaCO3

[0028] Further, the following steps are included:

[0029] 150 mg of CaCl2 powder, 10 mg of CuCl2, and 4 mg of DOX were added to a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and the mixture was stirred continuously at a speed of 500 r / min until uniform, and ultrasound was used to assist in dissolution;

[0030] Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish. Place the blue-capped bottle (open) and the Petri dish containing NH4HCO3 (open) in a desiccator. Place the desiccator in a 37°C oven and react for 18 h.

[0031] After the reaction was completed, the mixture was immediately centrifuged at 12000 r / min for 10 min at room temperature, and the supernatant was discarded;

[0032] The precipitate was washed twice with anhydrous ethanol and dried at 35 °C to obtain 10% Cu / DOX@CaCO3

[0033] Further, the following steps are included:

[0034] 150 mg of CaCl2 powder, 20 mg of CuCl2, and 4 mg of DOX were added to a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and the mixture was stirred continuously at a speed of 500 r / min until uniformly mixed, and ultrasound was used to assist in dissolution;

[0035] Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish. Place the blue-capped bottle (open) and the Petri dish containing NH4HCO3 (open) in a desiccator. Place the desiccator in a 37°C oven and react for 18 h.

[0036] After the reaction was completed, the mixture was immediately centrifuged at 12000 r / min for 10 min at room temperature, and the supernatant was discarded;

[0037] The precipitate was washed twice with anhydrous ethanol and dried at 35 °C to obtain 20% Cu / DOX@CaCO3.

[0038] Compared with the prior art, the present invention has the following beneficial effects:

[0039] The method is simple and does not require any post-processing. Anti-tumor drugs and copper ions can be directly loaded into calcite calcium carbonate nanoparticles, which can achieve large-scale mass production.

[0040] The nanodelivery platform prepared by this method not only has the pH responsiveness of the currently known calcium carbonate carriers but also has special GSH responsiveness, which is a special property brought about by this synthesis method, enabling it to achieve the ability to target tumor cells.

[0041] In addition, the nanodelivery platform utilizes the dual responsiveness of pH and GSH to achieve the simultaneous targeted delivery of DOX and copper ions into tumor cells, which can solve the toxic side effects of anti-tumor drugs and achieve the ability to enhance the therapeutic effect of tumors. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] The accompanying drawings are used to provide further understanding of the present invention and constitute a part of the specification. They are used to explain the present invention together with the embodiments of the present invention and do not constitute a limitation of the present invention.

[0043] Figure 1 Schematic diagram of the synthesis of different calcite calcium carbonate nano-delivery platforms in Examples 1-5 of the present invention, in which: (a) CaCO3, (b) Cu@CaCO3,

[0044] (c) 10% Cu@CaCO3, (d) 10% Cu / DOX@CaCO3, (e) 20% Cu / DOX@CaCO3;

[0045] Figure 2 1-5 are XRD data diagrams of different calcite calcium carbonate nano-delivery platforms synthesized in Examples 1-5 of the present invention, in which: (a) CaCO3, (b) Cu@CaCO3,

[0046] (c) 10% Cu@CaCO3, (d) 10% Cu / DOX@CaCO3, (e) 20% Cu / DOX@CaCO3;

[0047] Figure 3 It is a transmission electron microscopy image of the synthesized calcite calcium carbonate nano-delivery platform loaded with DOX and copper ions (20% Cu / DOX@CaCO3) in an embodiment of the present invention;

[0048] Figure 4 It is a mapping diagram of the synthesized calcite calcium carbonate nano-delivery platform loaded with DOX and copper ions (20% Cu / DOX@CaCO3) in the embodiment of the present invention;

[0049] Figure 5 Schematic diagram of pH responsiveness test of the synthesized DOX and copper ion loaded calcite calcium carbonate nano-delivery platform (20% Cu / DOX@CaCO3) in an embodiment of the present invention;

[0050] Figure 6Schematic diagram of the GSH responsiveness test of the synthesized calcite calcium carbonate nano-delivery platform loaded with DOX and copper ions (20% Cu / DOX@CaCO3) in the embodiment of the present invention;

[0051] Figure 7 It is a schematic diagram showing that the synthesized calcite calcium carbonate nano-delivery platform loaded with DOX and copper ions (20% Cu / DOX@CaCO3) in the embodiment of the present invention enters tumor cells at different time points;

[0052] Figure 8 The synthesized calcite calcium carbonate nano-delivery platform loaded with DOX and copper ions (20% Cu / DOX@CaCO3) in the embodiment of the present invention treated the Ca2+ in tumor cells for different time periods. 2+ Schematic diagram of

[0053] Fig. 9 Schematic diagram of ROS generation in tumor cells after being treated for different time periods with the synthesized calcite calcium carbonate nano-delivery platform loaded with DOX and copper ions (20% Cu / DOX@CaCO3) in an embodiment of the present invention;

[0054] Fig.10 Schematic diagram of the anti-tumor effect of the synthesized DOX and copper ion-loaded calcite calcium carbonate nano-delivery platform (20% Cu / DOX@CaCO3) in an example of the present invention. DETAILED DESCRIPTION

[0055] The technical solutions in the embodiments of the present invention will be described clearly and completely below in conjunction with the drawings in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all the embodiments.

[0056] Reference Figure 1-10 , a method for preparing a calcite calcium carbonate nano-delivery platform comprises the following steps:

[0057] Step 1: Dissolve 100-200 mg CaCl2, 5-25 mg CuCl2, and 1-10 mg DOX powder in 10-200 mL anhydrous ethanol solution, stir and mix continuously at a speed of 100-600 r / min, and place the mixed solution in a blue-capped bottle;

[0058] Step 2: Take 5-20g of NH4HCO3 solid and place it in a petri dish;

[0059] Step 3: Place the blue-cap bottle containing the mixed solution and the culture dish containing NH4HCO3 in a desiccator, and place the desiccator in an environment of 25-37°C to react for 10-24 hours. After the reaction is completed, a mixture for a calcite calcium carbonate nano-delivery platform loaded with anti-tumor drugs and copper ions is prepared;

[0060] Step 4: Use a centrifuge to separate the mixture formed in step 3, centrifuge at a certain speed and then vacuum dry at 30-60° C. to obtain a calcite calcium carbonate nano-delivery platform loaded with anti-tumor drugs and copper ions.

[0061] In this embodiment, the formation principle of preparing a calcite calcium carbonate nano-delivery platform for loading anti-tumor drugs and copper ions is described below.

[0062] Reaction mixture:

[0063] CaCl2(aq)→Ca 2+ +2Cl -

[0064] CuCl2(aq)→Cu 2+ +2Cl -

[0065] DOX

[0066] External CO2 gas diffusion:

[0067] NH4HCO3(s)→CO2(aq)

[0068] CO2(aq)+2OH - →CO3 2- +H2O

[0069] Formation of calcite calcium carbonate nano-delivery platform loaded with anti-tumor drugs and copper ions:

[0070] Ca 2+ +Cu 2+ +DOX+CO3 2- →Cu / DOX@CaCO3(s)

[0071] In order to better understand the technical solution of the present application, it is further described below in conjunction with embodiments.

[0072] Example 1: Synthesis of Calcite Calcium Carbonate (CaCO3)

[0073] Add 150 mg of CaCl2 powder to a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and stir continuously at 500 r / min to mix evenly and dissolve completely. Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish. Place the blue-capped bottle (open) and the Petri dish (open) containing NH4HCO3 in a desiccator, and place the desiccator in a 37°C oven to react for 18 hours. After the reaction is completed, centrifuge at 12000 r / min for 10 minutes and discard the supernatant. Wash the precipitate twice with anhydrous ethanol and dry it at 25°C to obtain calcite calcium carbonate. The product color is as follows: Figure 1 As shown in (a), the appearance of the synthesized calcite calcium carbonate sample is white. The product XRD data is as follows Figure 2 As shown in (a), the characteristic peaks at 23.03°, 29.39°, 35.93°, 39.41°, 43.08°, and 47.47° indicate that the product is a calcite crystal.

[0074] Example 2: Synthesis of Cu@CaCO3

[0075] Add 150 mg of CaCl2 powder and 10 mg of CuCl2 to a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and stir continuously at 500 r / min to mix evenly and completely dissolve. Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish. Place the blue-capped bottle (open) and the Petri dish containing NH4HCO3 (open) in a desiccator, and place the desiccator in a 37°C oven to react for 18 hours. After the reaction is completed, immediately centrifuge at 12000 r / min at room temperature for 10 minutes, and discard the supernatant. Wash the precipitate twice with anhydrous ethanol, and dry it at 37°C to obtain Cu@CaCO3. The product color is as follows Figure 1 As shown in (b), the appearance of the synthesized calcium carbonate sample is blue, which indicates that the copper ions have entered the calcium carbonate and achieved the loading of copper ions. Figure 2 As shown in (b), the characteristic peaks of 23.04°, 29.39°, 35.93°, 39.41°, 43.08°, and 47.47° indicate that the product is a calcite crystal.

[0076] Example 3: Synthesis of DOX@CaCO3

[0077] Add 150 mg of CaCl2 powder and 4 mg of DOX to a blue-capped bottle containing 100 mL of anhydrous ethanol solution, stir continuously at 500 r / min to mix evenly, and use ultrasound to help dissolve. Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish, place the blue-capped bottle (open) and the Petri dish containing NH4HCO3 (open) in a desiccator, and place the desiccator in a 37°C oven to react for 18 hours. After the reaction is completed, immediately centrifuge at 12000 r / min at room temperature for 10 minutes, and discard the supernatant. Wash the precipitate twice with anhydrous ethanol, and dry it at 40°C to obtain DOX@CaCO3. The product color is as follows Figure 1 As shown in (c), the appearance of the synthesized calcium carbonate sample is red, which indicates that DOX has entered the interior of the calcium carbonate and achieved the loading of DOX. The XRD data of the product are shown in Figure 2 As shown in (c), the characteristic peaks of 23.04°, 29.39°, 35.93°, 39.41°, 43.08°, and 47.47° indicate that the product is a calcite crystal.

[0078] Example 4: Synthesis of 10% Cu / DOX@CaCO3

[0079] Add 150 mg of CaCl2 powder, 10 mg of CuCl2, and 4 mg of DOX to a blue-capped bottle containing 100 mL of anhydrous ethanol solution, stir continuously at 500 r / min to mix evenly, and use ultrasound to help dissolve. Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish, put the blue-capped bottle (open) and the Petri dish containing NH4HCO3 (open) into a desiccator, and place the desiccator in a 37°C oven to react for 18 hours. After the reaction is completed, immediately centrifuge at 12000 r / min at room temperature for 10 minutes, and discard the supernatant. Wash the precipitate twice with anhydrous ethanol, and dry it at 35°C to obtain 10% Cu / DOX@CaCO3. The color of the product is as follows Figure 1 As shown in (d), the appearance of the synthesized calcium carbonate sample is light gray, which indicates that DOX and copper ions have entered the interior of the calcium carbonate, achieving the loading of DOX and copper ions. Figure 2 As shown in (d), the characteristic peaks of 23.04°, 29.39°, 35.93°, 39.41°, 43.08°, and 47.47° indicate that the product is a calcite crystal.

[0080] Example 5: Synthesis of 20% Cu / DOX@CaCO3

[0081] Add 150 mg of CaCl2 powder, 20 mg of CuCl2, and 4 mg of DOX to a blue-capped bottle containing 100 mL of anhydrous ethanol solution, stir continuously at 500 r / min to mix evenly, and use ultrasound to help dissolve. Weigh 5 g of NH4HCO3 powder and place it in a 10 cm Petri dish, put the blue-capped bottle (open) and the Petri dish containing NH4HCO3 (open) into a desiccator, and put the desiccator in a 37°C oven to react for 18 hours. After the reaction is completed, immediately centrifuge at 12000 r / min at room temperature for 10 minutes, and discard the supernatant. Wash the precipitate twice with anhydrous ethanol, and dry it at 35°C to obtain 20% Cu / DOX@CaCO3. The color of the product is as follows Figure 1 As shown in (e), the appearance of the synthesized calcium carbonate sample is dark purple, which indicates that DOX and copper ions have entered the interior of the calcium carbonate, achieving the loading of DOX and copper ions. The XRD data of the product are shown in Figure 2 As shown in (e), the characteristic peaks of 23.41°, 29.76°, 36.29°, 39.78°, 43.53°, and 47.92° indicate that the product is a calcite crystal form. By comparing the products of implementation cases 1 to 5, it can be clearly found that these peaks have shifted significantly, which indicates that DOX and copper ions have entered the interior of the calcium carbonate and have a certain impact on the crystal form of the calcium carbonate, resulting in the synthesized calcite calcium carbonate nano-delivery platform having certain special properties. Figure 3 This is the TEM image of the product. It can be seen from the image that the size of the synthesized calcite calcium carbonate nanoplatform is between 100 and 200 nm, and is mainly in the form of blocky particles. Figure 4 This is the mapping diagram of the product. It can be seen from the figure that individual nanoparticles are composed of different elements. The presence of Ca, C and O elements indicates that the main body of the synthesized product is calcium carbonate. The presence of N and Cu elements indicates that DOX and copper ions are doped into the calcium carbonate, realizing the loading of DOX and copper ions. Based on this, a calcite calcium carbonate nano-delivery platform with high loading of anti-tumor drugs DOX and copper ions was successfully constructed.

[0082] Example 6: pH and GSH responsiveness of 20% Cu / DOX@CaCO3 nanodelivery platform

[0083] 2 mg 20% ​​Cu / DOX@CaCO3 was dispersed in 2 mL PBS with pH 5.5 and 7.4, respectively. Samples were taken at different time points of 0.5 h, 1 h, 3 h, 6 h, 9 h, 12 h, and 24 h to measure the release of DOX. Figure 5As shown in the figure, as the pH value decreases, more DOX is released, indicating that the calcite calcium carbonate nano-delivery platform loaded with DOX and copper ions has good pH responsiveness. 3mg20%Cu / DOX@CaCO3 was dispersed in PBS and PBS+10mM GSH solution at pH=7.4, and samples were taken at different time points of 0.5h, 1h, 3h, 6h, 9h, 12h, and 24h to measure the release of DOX. Figure 6 As shown in the figure, DOX can be released quickly in the presence of GSH. This shows that the calcite calcium carbonate nano-delivery platform loaded with DOX and copper ions has good GSH responsiveness. This unique GSH responsive release is a special property of the nano-delivery platform synthesized by this method. The pH and GSH responsiveness experiments show that this synthesized nano-delivery platform has a certain targeting ability for tumor cells and can deliver DOX and copper ions to tumor cells very well.

[0084] Example 7: Study on the entry of 20% Cu / DOX@CaCO3 into tumor cells and drug release in tumor cells

[0085] This example mainly studies the synthetic nano-delivery platform 20% Cu / DOX@CaCO3 entering tumor cells and releasing drugs in tumor cells. The synthesized 20% Cu / DOX@CaCO3 was co-cultured with tumor cells and observed with laser confocal microscopy at different time points ( Figure 7 At the same time, after the synthesized 20% Cu / DOX@CaCO3 was co-cultured with tumor cells, the intracellular calcium release in living cells was detected using Fluo-4AM calcium ion fluorescent probe ( Figure 8 ). This indicates that the synthesized nano drug delivery platform 20% Cu / DOX@CaCO3 can be well engulfed by tumor cells, enter into tumor cells, and can be quickly decomposed in tumor cells to release DOX and copper ions.

[0086] Example 8: Anti-tumor effect

[0087] The synthesized 20% Cu / DOX@CaCO3 was co-cultured with tumor cells, and the ROS generation was detected at different time points ( Fig. 9 ). As time goes by, ROS increases, which indicates that the nano-delivery platform can release copper ions in tumor cells very well. Copper ions can produce ROS in tumor cells that have obvious killing ability to tumor cells. At the same time, CCK-8 was used to verify the killing effect of the synthesized nano-drug delivery platform on tumor cells ( Fig.10), by comparing with other treatment groups, it can be seen that the synthesized 20% Cu / DOX@CaCO3 showed the maximum inhibition rate on tumor cells, which also shows that the nano-delivery platform has a good anti-tumor effect and can kill tumor cells.

[0088] From the above, it can be seen that compared with the direct treatment of traditional chemotherapy drugs, the nano-delivery platform prepared by the method of the present application has the dual responsiveness of pH and GSH and the ability to target tumors, thereby reducing the systemic toxic side effects of anti-tumor drugs and can also utilize Cu 2+ The reaction with the high content of GSH in the tumor produces a large amount of reactive oxygen, which further enhances the effect of tumor treatment. This method of constructing a nano drug delivery system is of great significance for achieving targeted treatment of tumors.

[0089] The above description is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can make equivalent replacements or changes according to the technical scheme and inventive concept of the present invention within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.

Claims

1. A method for preparing a calcite calcium carbonate nano-delivery platform, characterized in that: The following steps are involved: Add 150 mg of CaCl2 powder into a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and stir continuously at 500 r / min to mix evenly and dissolve completely; Weigh 5g of NH4HCO3 powder and place it in a 10cm Petri dish. Place the blue-capped bottle and the Petri dish containing NH4HCO3 in a desiccator. Place the desiccator in a 37℃ oven to react for 18h. After the reaction, centrifuge at 12000 r / min for 10 min and discard the supernatant; The precipitate was washed twice with anhydrous ethanol and dried at 25°C to obtain CaCO3.

2. A method for preparing a calcite calcium carbonate nano-delivery platform, characterized in that: The following steps are involved: Add 150 mg of CaCl2 powder and 10 mg of CuCl2 into a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and stir continuously at 500 r / min to mix evenly and completely dissolve; Weigh 5g of NH4HCO3 powder and place it in a 10cm Petri dish. Place the blue-capped bottle and the Petri dish containing NH4HCO3 in a desiccator. Place the desiccator in a 37℃ oven to react for 18h. After the reaction was completed, the mixture was immediately centrifuged at 12000 r / min for 10 min at room temperature and the supernatant was discarded; The precipitate was washed twice with anhydrous ethanol and dried at 37 °C to obtain Cu@CaCO3.

3. A method for preparing a calcite calcium carbonate nano-delivery platform, characterized in that: The following steps are involved: Add 150 mg of CaCl2 powder and 4 mg of DOX into a blue-capped bottle containing 100 mL of anhydrous ethanol solution, stir continuously at 500 r / min to mix evenly, and use ultrasound to help dissolve; Weigh 5g of NH4HCO3 powder and place it in a 10cm Petri dish. Place the blue-capped bottle and the Petri dish containing NH4HCO3 in a desiccator. Place the desiccator in a 37℃ oven to react for 18h. After the reaction was completed, the mixture was immediately centrifuged at 12000 r / min for 10 min at room temperature and the supernatant was discarded; The precipitate was washed twice with anhydrous ethanol and dried at 40 °C to obtain DOX@CaCO3.

4. A method for preparing a calcite calcium carbonate nano-delivery platform, characterized in that: The following steps are involved: 150 mg of CaCl2 powder, 10 mg of CuCl2, and 4 mg of DOX were added to a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and the mixture was stirred continuously at a speed of 500 r / min until uniformly mixed, and ultrasound was used to assist in dissolution; Weigh 5g of NH4HCO3 powder and place it in a 10cm Petri dish. Place the blue-capped bottle and the Petri dish containing NH4HCO3 in a desiccator. Place the desiccator in a 37℃ oven to react for 18h. After the reaction was completed, the mixture was immediately centrifuged at 12000 r / min for 10 min at room temperature and the supernatant was discarded; The precipitate was washed twice with anhydrous ethanol and dried at 35 °C to obtain 10% Cu / DOX@CaCO3.

5. A method for preparing a calcite calcium carbonate nano-delivery platform, characterized in that: The following steps are involved: 150 mg of CaCl2 powder, 20 mg of CuCl2, and 4 mg of DOX were added to a blue-capped bottle containing 100 mL of anhydrous ethanol solution, and the mixture was stirred continuously at a speed of 500 r / min until uniformly mixed, and ultrasound was used to assist in dissolution; Weigh 5g of NH4HCO3 powder and place it in a 10cm Petri dish. Place the blue-capped bottle and the Petri dish containing NH4HCO3 in a desiccator. Place the desiccator in a 37℃ oven to react for 18h. After the reaction was completed, the mixture was immediately centrifuged at 12000 r / min for 10 min at room temperature and the supernatant was discarded; The precipitate was washed twice with anhydrous ethanol and dried at 35 °C to obtain 20% Cu / DOX@CaCO3.

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