Sodium pyruvate sustained-release nanoformulation based on pH response and preparation method and application thereof

Abstract

The present invention belongs to the field of biomedicine, and specifically relates to a sodium pyruvate sustained-release nanoformulation based on pH response, and its preparation method and application. The preparation method of the nanoformulation: under the action of a catalyst, 2,3-dichlorobenzaldehyde, dopamine, and p-aminophenol are added to ethylene glycol dimethyl ether for substitution reaction to obtain compound 1; compound 1 and triethylamine are added to a phosphorus oxychloride solution, and stirred to react to obtain a compound with a phosphate group, an aldehyde group, and a catechol group; polylysine and aldehyde phenylboronic acid are reacted in a PBS buffer solution to obtain aldehyde phenylboronic acid-modified polylysine; sodium pyruvate is added to a solution of a compound with a phosphate group, an aldehyde group, and a catechol group to obtain a mixed solution, the mixed solution is homogenized, and a polylysine solution modified by aldehyde phenylboronic acid is added during the homogenization process, and a mixed solution is obtained after the homogenization is completed, and the mixed solution continues to be stirred and reacted, and the reaction solution is centrifuged, washed, and dried to obtain.

Description

Technical Field

[0001] The invention belongs to the field of biomedicine, and in particular relates to a sodium pyruvate sustained-release nano preparation based on pH response, and a preparation method and application thereof. Background Art

[0002] Chronic obstructive pulmonary disease (COPD) is a common chronic respiratory disease with high morbidity, disability and mortality. Its onset is related to factors such as exposure to environmental pollutants and individual genetic susceptibility. The clinical characteristics of COPD are persistent airflow limitation and corresponding respiratory symptoms, such as excessive respiratory mucus secretion, chronic bronchitis and emphysema, and a significant decline in lung function, which may even cause respiratory and circulatory failure and lead to death. Therefore, active treatment measures should be taken to reduce mortality. Many studies have shown that in addition to COPD, many immune-related pathological processes (such as acute asthma, tumors, inflammation and autoimmune diseases) will cause local acidification of lesion tissues and acidic microenvironments. Therefore, identifying lesion tissues through pH response can help improve drug efficacy and reduce toxic side effects.

[0003] Through in-depth research, John et al. found that exogenous sodium pyruvate can be used to treat chronic obstructive pulmonary disease. Although sodium pyruvate can show certain therapeutic effects, it still faces many limitations in practical applications. The most direct problem is that sodium pyruvate is often widely distributed throughout the body after systemic administration, resulting in insufficient drug concentration in the lesion area, while healthy tissues may be exposed to unnecessary drugs. In addition, the lack of drug stability, bioavailability and sustained release ability also limits its full efficacy.

[0004] Therefore, the present invention proposes a strategy of encapsulating sodium pyruvate into a pH-responsive carrier, and realizes the delivery of sodium pyruvate by constructing a carrier system that can respond and release drugs in an acidic environment. Summary of the invention

[0005] In order to overcome the shortcomings of the prior art, one of the purposes of the present invention is to provide a method for preparing a sodium pyruvate sustained-release nanoformulation based on pH response, which is simple and efficient.

[0006] The second object of the present invention is to provide a sodium pyruvate sustained-release nanoformulation based on pH response, which has good biocompatibility and encapsulation rate.

[0007] The third object of the present invention is to provide an application of a sodium pyruvate sustained-release nanoformulation based on pH response, which has a wide range of applications.

[0008] One of the purposes of the present invention is achieved by the following technical solution:

[0009] A method for preparing a pH-responsive sodium pyruvate sustained-release nanoformulation comprises the following steps:

[0010]

[0011] (1) In the presence of a catalyst, 2,3-dichlorobenzaldehyde, dopamine, and p-aminophenol are added to ethylene glycol dimethyl ether for substitution reaction to obtain compound 1; compound 1 and triethylamine are added to a phosphorus oxychloride solution, stirred for reaction for 5-10 minutes, and then the pH of the reaction solution is adjusted to 8.5-9.0, and stirring is continued at 70-80° C. for 8-10 hours. After the reaction is completed, the reaction solution is purified to obtain a compound with phosphate, aldehyde, and catechol groups;

[0012]

[0013] (2) reacting polylysine and aldehyde phenylboronic acid in a molar ratio of 1:5-10 in a PBS buffer solution for 24-36 hours, dialyzing and freeze-drying the reaction solution to obtain aldehyde phenylboronic acid-modified polylysine;

[0014]

[0015] (3) Sodium pyruvate is added to the solution of the compound having phosphate, aldehyde and catechol groups obtained in step (1) to obtain a mixed solution, and the mixed solution is homogenized. During the homogenization process, a polylysine solution modified with aldehyde phenylboronic acid is added to obtain a mixed solution. The mixed solution is stirred and reacted continuously. After the reaction is completed, the reaction solution is centrifuged, and the precipitate is collected and washed and dried to obtain the product.

[0016] Furthermore, in step (1), the molar ratio of 2,3-dichlorobenzaldehyde, dopamine, p-aminophenol and catalyst is 1:1-1.5:1-1.5:0.01-0.03; the catalyst is a nitrogen heterocyclic carbene palladium complex; the temperature of the substitution reaction is 70-85°C and the time is 14-16h.

[0017] Furthermore, the nitrogen heterocyclic carbene palladium complex is selected from one of NHC-PD(II)-MP and NHC-PD(II)-IM.

[0018] Furthermore, in step (1), the molar ratio of compound 1 to phosphorus oxychloride and triethylamine is 1:0.3-0.5:1.5-2.

[0019] Furthermore, in step (3), the mass ratio of the sodium pyruvate, the compound with phosphate, aldehyde and catechol groups is (1-2):1, and the concentration of the aldehyde phenylboronic acid modified polylysine solution is 0.01-0.05 g / mL.

[0020] Furthermore, the dialysis in step (2) is performed using a dialysis bag with a molecular weight cutoff of 1000-2000Da.

[0021] Furthermore, in step (2), the dosage ratio of polylysine to PBS buffer solution is 1 g:10-20 mL; and the average molecular weight of the polylysine is 3600-4000 Da.

[0022] Furthermore, the pH of the PBS buffer solution is 7.4.

[0023] Furthermore, in step (3), the volume ratio of the mixed solution to the polylysine solution modified with aldehyde phenylboronic acid is 1:1-1.5, the homogenization time is 10-30 minutes, and the stirring reaction time is 48-55 hours.

[0024] The second object of the present invention is achieved by adopting the following technical solution:

[0025] The sodium pyruvate sustained-release nanoformulation based on pH response is prepared by using the preparation method of the sodium pyruvate sustained-release nanoformulation based on pH response.

[0026] The third object of the present invention is achieved by adopting the following technical solution:

[0027] The above-mentioned pH-responsive sodium pyruvate sustained-release nanoformulation is used for preparing medicine for treating chronic obstructive pulmonary disease.

[0028] Compared with the prior art, the present invention has the following beneficial effects:

[0029] 1. The present invention uses a compound with a phosphate group, an aldehyde group and a catechol group, and polylysine modified with aldehyde phenylboronic acid to encapsulate sodium pyruvate into nanoparticles, wherein the phenylboric acid on the polylysine modified with aldehyde phenylboronic acid forms a borate ester bond with the catechol on the compound with a phosphate group, an aldehyde group and a catechol group; the amino group on the polylysine modified with aldehyde phenylboronic acid forms a Schiff base bond with the aldehyde group on the compound with a phosphate group, an aldehyde group and a catechol group; the amino group on the polylysine modified with aldehyde phenylboronic acid can electrostatically interact with the phosphate group on the compound with a phosphate group, an aldehyde group and a catechol group, thereby forming three kinds of interactions. The drug sodium pyruvate is added during the cross-linking process, and finally the nanoparticles encapsulating sodium pyruvate are formed. Test data show that the encapsulation rate of the nanoparticles for sodium pyruvate is 93-96%.

[0030] 2. The three interactions in the pH-responsive sodium pyruvate sustained-release nanoformulation of the present invention are all sensitive to pH, so it is possible to achieve sustained release of sodium pyruvate in the acidic microenvironment of the lungs, thereby prolonging the residence time of the drug in the lungs and improving the therapeutic effect of the drug. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 The compound with phosphate, aldehyde and catechol groups obtained in Example 1 of the present invention is 1 H-NMR spectrum;

[0032] Figure 2 is a SEM image of the pH-responsive sodium pyruvate sustained-release nanoformulation obtained in Example 1 of the present invention;

[0033] Figure 3 is a graph showing the biocompatibility of the nanoformulation with lung cells in Example 1;

[0034] Figure 4 This is a graph showing the biocompatibility of the nanoformulation in Example 1 with fibroblasts. DETAILED DESCRIPTION

[0035] The technical scheme of the present invention is further described below in conjunction with specific embodiments. However, it should be understood by those skilled in the art that the following examples are only used to illustrate the present invention and should not be regarded as limiting the present invention. The specific conditions not specified in the examples are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used, unless otherwise specified, are conventional products obtained through commercial channels.

[0036] 1. Implementation

[0037] Example 1

[0038] Provided is a method for preparing a pH-responsive sodium pyruvate sustained-release nanoformulation, the specific preparation method being as follows:

[0039] (1) Preparation of compounds with phosphate, aldehyde and catechol groups:

[0040] According to the molar ratio of 2,3-dichlorobenzaldehyde, dopamine, p-aminophenol and NHC-PD(II)-MP of 1:1:1:0.01, the above raw materials were added to ethylene glycol dimethyl ether, and the reaction was carried out at 80°C for 15 hours. After the reaction, the reaction solution was passed through a column and dried with Na2S04 to obtain compound 1. According to the molar ratio of compound 1 to phosphorus oxychloride and triethylamine of 1:0.4:1.7, the solution of compound 1 (the solvent is dichloromethane) was added dropwise to the phosphorus oxychloride solution (the solvent is dichloromethane) to obtain a mixture. liquid, then triethylamine solution (solvent is dichloromethane) is added dropwise to the above mixed solution, stirred for 8 minutes, quenched with water after the reaction is completed, the reaction solution is extracted with CH2Cl2, the organic phase is collected and dried with Na2S04 and concentrated, the viscous oil is placed in an ice bath for cooling, the pH is adjusted to 9 with 2 mol / L sodium hydroxide, stirred at 70°C for 8 hours, hot filtered, the filtrate is dried with Na2S04, concentrated and then recrystallized at 140°C to obtain a compound with phosphate, aldehyde and catechol groups;

[0041] The specific reaction formula is as follows:

[0042]

[0043] The NMR characterization results of the obtained compounds with phosphate, aldehyde and catechol groups are as follows: Figure 1 As shown:

[0044] 1 H-NMR (400MHz, DMSO)δ=2.94(t,2H),3.41(t,2H),6.53-6.58(m,2H),6.62(m,1H),6.80(m,1H),6.96(s,1H),7.02(m,1H),7.24-7.25(m,3H),7.57(d,2H),9.49(s,2H),10.37(s,1H),10.49(s,1H). The above results confirmed that the product obtained was the target product.

[0045] (2) Preparation of pH-responsive sodium pyruvate sustained-release nanoformulations:

[0046] Add polylysine (average molecular weight 3800Da) and aldehyde phenylboronic acid in a molar ratio of 1:8 to a PBS buffer solution with a pH of 7.4, wherein the amount ratio of polylysine to PBS buffer solution is 1g:15mL, and react for 30h. After the reaction is completed, the reaction solution is loaded into a dialysis bag with a molecular weight cutoff of 1500Da. After the dialysis is completed, the mixture in the dialysis bag is freeze-dried to obtain polylysine modified with aldehyde phenylboronic acid. The polylysine modified with aldehyde phenylboronic acid is dissolved in water to obtain a polylysine solution modified with aldehyde phenylboronic acid with a concentration of 0.03g / mL, which is set aside.

[0047] The specific reaction formula is as follows:

[0048]

[0049] According to the mass ratio of sodium pyruvate to the compound with phosphate group, aldehyde group and catechol group of 2:1, sodium pyruvate is added to the aqueous solution of the compound with phosphate group, aldehyde group and catechol group to obtain a mixed solution; the mixed solution is placed in a homogenizer, and the speed of the homogenizer is set to 5000 rpm, and the above-mentioned 0.03 g / mL aldehyde phenylboronic acid modified polylysine solution is slowly added during the homogenization process, the volume ratio of the mixed solution to the aldehyde phenylboronic acid modified polylysine solution is 1:1, the homogenization time is 20 minutes, and a mixed solution is obtained after the homogenization is completed. The mixed solution is transferred to a stirrer for reaction for 50 hours. After the reaction is completed, the reaction solution is centrifuged and the product is collected.

[0050] The specific reaction formula is as follows:

[0051]

[0052] This embodiment also provides a pH-responsive sodium pyruvate sustained-release nanoformulation, which is prepared by the above-mentioned preparation method, and its SEM image is as follows: Figure 2 shown.

[0053] Example 2

[0054] Provided is a method for preparing a pH-responsive sodium pyruvate sustained-release nanoformulation, the specific preparation method being as follows:

[0055] (1) Preparation of compounds with phosphate, aldehyde and catechol groups:

[0056] According to the molar ratio of 2,3-dichlorobenzaldehyde, dopamine, p-aminophenol and NHC-PD(II)-IM of 1:1.2:1.2:0.02, the above raw materials were added to ethylene glycol dimethyl ether, and the reaction was carried out at 70°C for 16 hours. After the reaction, the reaction solution was passed through a column and dried with Na2S04 to obtain compound 1. According to the molar ratio of compound 1 to phosphorus oxychloride and triethylamine of 1:0.3:1.5, the solution of compound 1 (the solvent is dichloromethane) was added dropwise to the phosphorus oxychloride solution (the solvent is dichloromethane) to obtain a mixed solution, and then the triethylamine was added to the solution. Ethylamine solution (solvent: dichloromethane) was added dropwise to the above mixed solution and stirred for 10 minutes. After the reaction was completed, the reaction was quenched with water, the reaction solution was extracted with CH2Cl2, the organic phase was collected and dried with Na2S04 and concentrated, the viscous oil was cooled in an ice bath, the pH was adjusted to 9 with 2 mol / L sodium hydroxide, stirred at 70°C for 8 hours, and filtered hot. The filtrate was dried and concentrated with Na2S04 and then recrystallized at 140°C to obtain a compound with phosphate, aldehyde and catechol groups. The specific reaction formula is the same as that in Example 1.

[0057] (2) Preparation of pH-responsive sodium pyruvate sustained-release nanoformulations:

[0058] Polylysine (average molecular weight of 3600Da) and aldehyde phenylboronic acid in a molar ratio of 1:5 are added to a PBS buffer solution with a pH of 7.4, wherein the amount ratio of polylysine to PBS buffer solution is 1g:10mL, and the reaction is carried out for 36 hours. After the reaction is completed, the reaction solution is loaded into a dialysis bag with a molecular weight cutoff of 1500Da. After the dialysis is completed, the mixture in the dialysis bag is freeze-dried to obtain aldehyde phenylboronic acid-modified polylysine, and the aldehyde phenylboronic acid-modified polylysine is dissolved in water to obtain a aldehyde phenylboronic acid-modified polylysine solution with a concentration of 0.01g / mL; the specific reaction formula is the same as that in Example 1.

[0059] According to the mass ratio of sodium pyruvate, compounds with phosphate, aldehyde and catechol groups of 1.5:1, sodium pyruvate is added to the aqueous solution of compounds with phosphate, aldehyde and catechol groups to obtain a mixed solution, the mixed solution is placed in a homogenizer, and the speed of the homogenizer is set to 5000rpm, and the above-mentioned concentration of 0.01g / mL of aldehyde phenylboronic acid-modified polylysine solution is slowly added during the homogenization process, the volume ratio of the mixed solution to the aldehyde phenylboronic acid-modified polylysine solution is 1:1.5, the homogenization time is 30min, and a mixed solution is obtained after the homogenization is completed, and the mixed solution is transferred to a stirrer for reaction for 55h. After the reaction is completed, the reaction solution is centrifuged to obtain. The specific reaction formula is the same as in Example 1.

[0060] This embodiment also provides a pH-responsive sodium pyruvate sustained-release nanoformulation, which is prepared using the above-mentioned preparation method.

[0061] Example 3

[0062] Provided is a method for preparing a pH-responsive sodium pyruvate sustained-release nanoformulation, the specific preparation method being as follows:

[0063] (1) Preparation of compounds with phosphate, aldehyde and catechol groups:

[0064] According to the molar ratio of 2,3-dichlorobenzaldehyde, dopamine, p-aminophenol and NHC-PD(II)-MP of 1:1.5:1.5:0.03, the above raw materials were added to ethylene glycol dimethyl ether, and the reaction was carried out at 85°C for 14 hours. After the reaction, the reaction solution was passed through a column and dried with Na2S04 to obtain compound 1. According to the molar ratio of compound 1 to phosphorus oxychloride and triethylamine of 1:0.5:2, the solution of compound 1 (the solvent is dichloromethane) was added dropwise to the phosphorus oxychloride solution (the solvent is dichloromethane) to obtain a mixed solution, and then triethylamine was added to the mixture. Amine solution (solvent: dichloromethane) was added dropwise to the above mixed solution and stirred for 5 minutes. After the reaction was completed, the mixture was quenched with water. The reaction solution was extracted with CH2Cl2, the organic phase was collected and dried over Na2S04 and concentrated. The viscous oil was cooled in an ice bath, the pH was adjusted to 9 with 2 mol / L sodium hydroxide, stirred at 70°C for 8 hours, and filtered hot. The filtrate was dried over Na2S04, concentrated, and then recrystallized at 140°C to obtain a compound with phosphate, aldehyde and catechol groups. The specific reaction formula is as follows: the same as Example 1.

[0065] (2) Preparation of pH-responsive sodium pyruvate sustained-release nanoformulations:

[0066] Polylysine (average molecular weight of 4000Da) and aldehyde phenylboronic acid in a molar ratio of 1:10 are added to a PBS buffer solution with a pH of 7.4, wherein the amount ratio of polylysine to PBS buffer solution is 1g:20mL, and the reaction is carried out for 24 hours. After the reaction is completed, the reaction solution is loaded into a dialysis bag with a molecular weight cutoff of 1500Da. After the dialysis is completed, the mixture in the dialysis bag is freeze-dried to obtain polylysine modified with aldehyde phenylboronic acid. The polylysine modified with aldehyde phenylboronic acid is dissolved in water to obtain a polylysine solution modified with aldehyde phenylboronic acid with a concentration of 0.05g / mL. The specific reaction formula is as follows, which is the same as that in Example 1.

[0067] According to the mass ratio of sodium pyruvate, the compound with phosphate, aldehyde and catechol groups of 1:1, sodium pyruvate is added to the aqueous solution of the compound with phosphate, aldehyde and catechol groups to obtain a mixed solution, the mixed solution is placed in a homogenizer, and the speed of the homogenizer is set to 5000rpm, and the above-mentioned concentration of 0.05g / mL of aldehyde phenylboronic acid modified polylysine solution is slowly added during the homogenization process, the volume ratio of the mixed solution to the aldehyde phenylboronic acid modified polylysine solution is 1:1, the homogenization time is 10min, and a mixed solution is obtained after the homogenization is completed, and the mixed solution is transferred to a stirrer for reaction for 48h. After the reaction is completed, the reaction solution is centrifuged to obtain. The specific reaction formula is the same as in Example 1.

[0068] This embodiment also provides a pH-responsive sodium pyruvate sustained-release nanoformulation, which is prepared using the above-mentioned preparation method.

[0069] (II) Comparative Example

[0070] Comparative Example 1

[0071] Comparative Example 1 uses polylysine instead of polylysine modified with aldehyde phenylboronic acid, and the rest is the same as Example 1.

[0072] (III) Experimental examples

[0073] Experimental Example 1

[0074] Experimental Example 1 The biocompatibility of the nanoparticle preparation of Example 1 was tested, and the testing process was as follows:

[0075] I. Lung cell biocompatibility:

[0076] 100 μL of HULEC-5a lung cells (1×10 5 / mL), the culture medium in the 96-well plate is DMEM complete culture medium, the 96-well plate is placed in a cell culture incubator at 37°C and 5% CO2 for 12 hours, and then the cell culture medium in each well is gently aspirated, 200μL of DMEM complete culture medium containing the nanoformulation of Example 1 at a concentration of 50μg / mL is added to the 96-well plate, and the culture is continued in the cell culture incubator for 24 hours, the 96-well plate is taken out, and the cell morphology is observed under a microscope. The present invention uses DMEM serum-free culture medium as a blank control and DMEM complete culture medium supplemented with 5% phenol as a positive control. The results are as follows Figure 3 shown.

[0077] II. Fibroblast Biocompatibility:

[0078] 100 μL of L929 fibroblasts (1×10 5 / mL), the culture medium in the 96-well plate is DMEM complete culture medium, the 96-well plate is placed in a cell culture incubator at 37°C and 5% CO2 for 12 hours, and then the cell culture medium in each well is gently aspirated, and 200 μL of DMEM complete culture medium containing 50 μg / mL of the nanoparticle preparation of Example 1 is added to the 96-well plate and continued to be cultured in the cell culture incubator for 24 hours, and the 96-well plate is taken out and the cell morphology is observed under a microscope. The present invention uses DMEM serum-free culture medium as a blank control and DMEM complete culture medium supplemented with 5% phenol as a positive control. The results are as follows Figure 4 shown.

[0079] Figure 3 This is a graph showing the biocompatibility of the nanoformulation in Example 1 with lung cells. Figure 4 This is the result of the biocompatibility of the nanoparticle preparation and fibroblasts in Example 1. Figure 3-4It is known that there is no significant difference in the fluorescence intensity of cells in the field of view. That is, the nanoparticle preparation prepared in Example 1 has good biocompatibility.

[0080] Experimental Example 2

[0081] Experimental Example 2 The encapsulation efficiency of the sodium pyruvate sustained-release nanoformulations of Examples 1-3 and Comparative Example 1 was tested, and the testing process was as follows:

[0082] Select 255nm as the detection wavelength of sodium pyruvate, determine its content by UV method, and establish the standard curve of sodium pyruvate;

[0083] In step (2) of preparing the sodium pyruvate sustained-release nanoformulation, 1 mL of the reaction solution after the reaction of Examples 1-3 and Comparative Example 1 (before centrifugation) was taken, and the volume of the supernatant was measured after centrifugation. The absorbance was substituted into the standard curve to determine the content of free sodium pyruvate in the supernatant, and the encapsulation rate of sodium pyruvate was calculated. The calculation formula was: encapsulation rate = free sodium pyruvate content in the supernatant / initial amount of sodium pyruvate added × 100%. For specific results, see Table 1.

[0084] Table 1

[0085]

[0086] It can be seen from Table 1 that the sodium pyruvate nano sustained-release preparation prepared in the present invention has a higher encapsulation rate.

[0087] Experimental Example 3

[0088] Experimental Example 3 The effects of the sodium pyruvate preparations of Examples 1-3 and Comparative Example 1 were tested, and the testing process was as follows:

[0089] Phosphate buffer solutions with pH values ​​of 5.6, 6.5, and 7.4 were prepared respectively, 50 mg of the sodium pyruvate preparations of Examples 1-3 and Comparative Example 1 were placed in dialysis bags, and the dialysis bags were placed in 50 mL of phosphate buffer solutions with different pH values, respectively. The drugs were released in vitro under constant temperature of 37° C. with magnetic stirring. Samples were taken once at 2 h, 4 h, 8 h, 12 h, and 14 h after release, and 5 mL of the solution was taken each time to detect the absorbance value of sodium pyruvate. At the same time, 5 mL of fresh phosphate buffer solution was added. The absorbance at an absorption wavelength of 255 nm was tested using an ultraviolet spectrophotometer. According to the detected absorbance value of sodium pyruvate, the cumulative percentage of drug release under different pH conditions was calculated. The results are shown in Table 2.

[0090] Table 2

[0091]

[0092] It can be seen from Table 2 that the cumulative release of sodium pyruvate has obvious pH sensitivity. The stronger the acidity, the greater the cumulative release of sodium pyruvate in the same time. This is because the phenylboric acid on the polylysine modified by the aldehyde phenylboronic acid forms a borate ester bond with the catechol on the compound with phosphate, aldehyde and catechol groups; the amino group on the polylysine modified by the aldehyde phenylboronic acid forms a Schiff base bond with the aldehyde group on the compound with phosphate, aldehyde and catechol groups; the amino group on the polylysine modified by the aldehyde phenylboronic acid can interact electrostatically with the phosphate on the compound with phosphate, aldehyde and catechol groups, thereby forming three interactions, and the above three interactions are all sensitive to pH.

[0093] Under the same pH conditions, compared with Comparative Example 1, the cumulative percentage of in vitro drug release of the sustained-release nanoformulations of Examples 1-3 is higher. That is, the sustained-release nanoformulations prepared by the present invention have obvious pH sensitivity. Further analysis shows that the nano sustained-release preparation prepared by Comparative Example 1 using polylysine instead of polylysine modified with aldehyde phenylboronic acid is slightly less sensitive to pH.

[0094] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit them. The basic principles and main features of the present invention have been described above with specific implementation schemes. On the basis of the present invention, some modifications or replacements may be made thereto, but these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of protection claimed by the present invention.

Claims

1. A method for preparing a pH-responsive sodium pyruvate sustained-release nanoformulation, characterized in that: The following steps are involved: (1) In the presence of a catalyst, 2,3-dichlorobenzaldehyde, dopamine, and p-aminophenol are added to ethylene glycol dimethyl ether for substitution reaction to obtain compound 1; compound 1 and triethylamine are added to a phosphorus oxychloride solution, stirred for reaction for 5-10 minutes, and then the pH of the reaction solution is adjusted to 8.5-9.0, and stirring is continued at 70-80° C. for 8-10 hours. After the reaction is completed, the reaction solution is purified to obtain a compound with phosphate, aldehyde, and catechol groups; (2) reacting polylysine and aldehyde phenylboronic acid in a molar ratio of 1:5-10 in a PBS buffer solution for 24-36 hours, dialyzing and freeze-drying the reaction solution to obtain aldehyde phenylboronic acid-modified polylysine; (3) Sodium pyruvate is added to the solution of the compound having phosphate, aldehyde and catechol groups obtained in step (1) to obtain a mixed solution, and the mixed solution is homogenized. During the homogenization process, the polylysine solution modified with aldehyde phenylboronic acid obtained in step (2) is added to obtain a mixed solution. After the homogenization, the mixed solution is stirred to react. After the reaction is completed, the reaction solution is centrifuged, and the precipitate is collected and washed and dried to obtain the product.

2. The method for preparing the pH-responsive sodium pyruvate sustained-release nanoformulation according to claim 1, characterized in that: The molar ratio of 2,3-dichlorobenzaldehyde, dopamine, p-aminophenol and catalyst in step (1) is 1:1-1.5:1-1.5:0.01-0.03; the catalyst is a nitrogen heterocyclic carbene palladium complex; the temperature of the substitution reaction is 70-85°C and the time is 14-16h.

3. The method for preparing the pH-responsive sodium pyruvate sustained-release nanoformulation according to claim 2, characterized in that: The nitrogen heterocyclic carbene palladium complex is selected from one of NHC-PD(II)-MP and NHC-PD(II)-IM.

4. The method for preparing the pH-responsive sodium pyruvate sustained-release nanoformulation according to claim 1, characterized in that: In step (1), the molar ratio of compound 1 to phosphorus oxychloride and triethylamine is 1:0.3-0.5:1.5-2.

5. The method for preparing the pH-responsive sodium pyruvate sustained-release nanoformulation according to claim 1, characterized in that: The mass ratio of the sodium pyruvate, the compound with phosphate, aldehyde and catechol groups in step (3) is (1-2):1, and the concentration of the aldehyde phenylboronic acid modified polylysine solution is 0.01-0.05 g / mL.

6. The method for preparing the pH-responsive sodium pyruvate sustained-release nanoformulation according to claim 1, characterized in that: The dosage ratio of the polylysine and PBS buffer solution in step (2) is 1 g:10-20 mL; the average molecular weight of the polylysine is 3600-4000 Da.

7. The method for preparing the pH-responsive sodium pyruvate sustained-release nanoformulation according to claim 1, characterized in that: In step (3), the volume ratio of the mixed solution to the polylysine solution modified with aldehyde phenylboronic acid is 1:1-1.5, the homogenization time is 10-30 minutes, and the stirring reaction time is 48-55 hours.

8. A pH-responsive sodium pyruvate sustained-release nanoformulation, characterized in that: The preparation method is described in any one of claims 1 to 7.

9. The use of the pH-responsive sodium pyruvate sustained-release nanoformulation according to claim 8, characterized in that: Used for preparing medicines for treating chronic obstructive pulmonary disease.

Images