Sodium pyruvate aerosol for regulating respiratory microecology and preparation method and application thereof

Abstract

The present invention belongs to the field of biomedicine, and specifically relates to a sodium pyruvate aerosol for regulating respiratory microecology, and a preparation method and application thereof. The sodium pyruvate aerosol comprises the following components in parts by weight: 0.05-0.1 parts of modified sodium pyruvate, 80-95 parts of propellant, and 5-10 parts of water for injection. The present invention detects the pharmacological properties of the above-mentioned aerosol, and the experimental results show that the aerosol of the present invention has the effects of anti-inflammatory and anti-fibrosis, reducing the abundance of pathogenic bacteria, and increasing the abundance of probiotics.

Description

Technical Field

[0001] The invention belongs to the field of biomedicine, and specifically relates to a sodium pyruvate aerosol for regulating respiratory microecology, and a preparation method and application thereof. Background Art

[0002] The human body is a huge ecosystem, with a large number of microorganisms attached to the surface of each organ. The microbial community ensures the normal nutritional metabolism and microecological stability of the body through interactions with host cells. For a long time, due to the physiological functions and characteristics of the respiratory tract, the regulatory role of its microbial flora has not received enough attention. Respiratory microecology refers to the microorganisms that normally colonize the human respiratory tract, which use the respiratory tract and its metabolites as their living environment, and the microbial community or the microbial community and the host constantly exchange substances, energy, and information, and finally form a relatively stable biological system. The respiratory microecology changes dynamically, and the components are constantly migrating, eliminating, and reproducing.

[0003] Changes in respiratory microecology are associated with a variety of diseases, such as chronic non-obstructive pulmonary disease and pulmonary fibrosis. Molyneaux et al. collected samples through bronchoscopy, used 16S rRNA gene real-time fluorescence quantitative PCR and pyrophosphate sequencing, and compared the alveolar lavage fluid specimens of patients with acute exacerbation of idiopathic pulmonary fibrosis and stable pulmonary fibrosis. They found that Campylobacter and Stenotrophomonas maltophilia increased in patients with acute exacerbation, while Veillonella was more common in stable patients. O'Dwyer et al. used a mouse pulmonary fibrosis model and found that changes in lung microecology were associated with local host inflammation in mice and could promote the progression of idiopathic pulmonary fibrosis. The above studies all indicate that changes in respiratory flora are closely related to the development of pulmonary fibrosis.

[0004] The 17 human clinical data submitted by Cellular Sciences Inc to the FDA found that inhaled sodium pyruvate can reduce lung inflammation and congestion in patients with chronic non-obstructive pulmonary disease, pulmonary fibrosis, cystic fibrosis, asthma, sinusitis, and influenza. And inhaled sodium pyruvate can also relieve symptoms associated with CoVID-19 patients with pulmonary fibrosis. Arofylline is a PDE4 inhibitor. Liu Pingxia et al. found that arofylline can relieve cough symptoms and reduce the use of bronchodilators. By summarizing a large amount of clinical data, it was found that although the above two drugs can be used to relieve the symptoms of pulmonary fibrosis, there are problems of poor efficacy and instability. Therefore, it is necessary to further explore the performance of drugs for the treatment of pulmonary fibrosis. And there is no report on sodium pyruvate aerosol for regulating respiratory microecology. Based on this, the present invention provides a sodium pyruvate aerosol for regulating respiratory microecology. Summary of the invention

[0005] The first object of the present invention is to provide a sodium pyruvate aerosol for regulating respiratory microecology, which has the effects of anti-inflammatory and anti-fibrosis, reducing the abundance of pathogenic bacteria, and increasing the abundance of probiotics.

[0006] The second purpose of the present invention is to provide a method for preparing a sodium pyruvate aerosol for regulating respiratory microecology, which is simple and easy to operate.

[0007] The third purpose of the present invention is to provide an application of sodium pyruvate aerosol for regulating respiratory microecology, which has broad application prospects.

[0008] In order to achieve the above object, the technical solution adopted by the present invention is:

[0009] A sodium pyruvate aerosol for regulating respiratory microecology comprises the following components in parts by weight: 0.05-0.1 parts of modified sodium pyruvate, 80-95 parts of propellant, and 5-10 parts of water for injection; the preparation process of the modified sodium pyruvate comprises the following steps:

[0010]

[0011] (1) reacting arofylline with ethyl 4-bromobutyrate under a catalyst condition, and after the reaction is completed, purifying the reaction solution to obtain intermediate 1;

[0012] (2) reacting the intermediate 1 obtained in step (1) with hydrazine hydrate. After the reaction is completed, the reaction solution is purified to obtain the intermediate 2;

[0013] (3) Under catalyst conditions, the intermediate 2 obtained in step (2) is reacted with a 2,3,4-trihydroxybenzaldehyde solution under stirring. After the reaction is completed, the reaction solution is purified to obtain the intermediate 3;

[0014] (4) adding the intermediate 3 obtained in step (3), tert-butyldimethylsilyl chloride and imidazole to DMF, mixing and stirring for reaction. After the reaction is completed, the reaction solution is purified to obtain the intermediate 4;

[0015] (5) The intermediate 4 obtained in step (4) is added to DMF, copper iodide and samarium, and the mixture is stirred for 3-5 hours; water is added to the reaction solution to quench it, and then an organic phase is obtained by extraction; the organic phase is dried and concentrated, and then added to tetrahydrofuran, and then tetrabutylammonium fluoride is added, and the reaction is stirred. After the reaction is completed, the reaction solution is purified to obtain the intermediate 5;

[0016] (6) Adding sodium pyruvate and the intermediate 5 obtained in step (5) into a sodium hydroxide solution, stirring for reaction, and after completion of the reaction, purifying the reaction solution to obtain modified sodium pyruvate.

[0017] Furthermore, the catalyst in step (1) is potassium carbonate and benzyltriethylammonium chloride; the molar ratio of arofylline, ethyl 4-bromobutyrate, potassium carbonate and benzyltriethylammonium chloride is 1:1.5-2:2-2.5:0.5-1; the reaction temperature is 55-65°C and the reaction time is 15-17h.

[0018] Furthermore, in step (2), the molar ratio of the intermediate 1 to hydrazine hydrate is 1:4-6; the reaction temperature is 75-85°C, and the reaction time is 19-21h.

[0019] Furthermore, in step (3), the ratio of the intermediate 2 to 2,3,4-trihydroxybenzaldehyde and the catalyst is 1 mmol: 1-2 mmol: 0.04-0.06 mL; the catalyst is 37% concentrated hydrochloric acid; and the stirring reaction time is 70-74 h.

[0020] Furthermore, in step (4), the molar ratio of the intermediate 3, tert-butyldimethylsilyl chloride, and imidazole is 1:3.2-3.6:6-7, the stirring reaction temperature is 32-38°C, and the stirring time is 8-10h; in step (5), the molar ratio of the intermediate 4, copper iodide, samarium, and tetrabutylammonium fluoride is 1:0.1-0.12:2-2.2:4.5-5.0, and the stirring reaction time is 2-3h.

[0021] Furthermore, in step (6), the molar ratio of the intermediate 5, sodium pyruvate and sodium hydroxide is 1:2-2.5:1.8-22; and the stirring reaction time is 0.8-1.2h.

[0022] Furthermore, the propellant is tetrafluoroethane or heptafluoropropane.

[0023] The preparation method of the sodium pyruvate aerosol for regulating respiratory microecology comprises the following steps: compounding modified sodium pyruvate and water for injection according to a weight ratio, filling, and pressing into a propellant to obtain the aerosol.

[0024] Use of the above-mentioned sodium pyruvate aerosol for regulating respiratory microecology in the preparation of a drug for regulating respiratory microecology.

[0025] Compared with the prior art, the beneficial effects of the present invention are mainly:

[0026] The present invention provides a novel sodium pyruvate aerosol for regulating respiratory microecology, and the sodium pyruvate aerosol includes modified sodium pyruvate and other components. Modified sodium pyruvate is specifically prepared by subjecting arofylline to a substitution reaction with ethyl 4-bromobutyrate, and the intermediate 1 obtained undergoes nucleophilic substitution with hydrazine hydrate to generate an intermediate 2 containing a hydrazine group; intermediate 2 undergoes a condensation reaction with 2,3,4-trihydroxybenzaldehyde to generate a hydrazone (intermediate 3), and the hydroxyl group in intermediate 3 is protected to obtain intermediate 4; intermediate 4 reacts in the presence of copper iodide and samarium to obtain intermediate 5; intermediate 5 undergoes nucleophilic substitution with sodium pyruvate to obtain the final modified sodium pyruvate. The results of the study show that compared with the use of arofylline and sodium pyruvate alone, the aerosol prepared using the modified sodium pyruvate is:

[0027] (1) It can better improve the physical fitness of rats with pulmonary fibrosis model;

[0028] (2) It can more effectively reduce the expression of inflammatory factors IL-6, IL-1β, and TNF-α in the serum of rats with pulmonary fibrosis model, thereby exerting an anti-pulmonary fibrosis effect;

[0029] (3) It can more effectively reduce the lung coefficient of rats with pulmonary fibrosis model;

[0030] (4) It can more effectively reduce the hydroxyproline content in the lung tissue of rats with pulmonary fibrosis model and alleviate the degree of development of pulmonary fibrosis in model rats.

[0031] In addition, after coupling sodium pyruvate and arofylline, they can reduce the abundance of pathogenic bacteria and increase the abundance of probiotics on the basis of combining the advantages of both, thereby regulating the respiratory microecology and fundamentally playing a role in treating lung diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 is the intermediate 1 in Example 1 of the present invention 1 HNMR spectra;

[0033] Figure 2 is the mass spectrum of the intermediate 1 in Example 1 of the present invention;

[0034] Figure 3 is the intermediate 2 in Example 1 of the present invention 1 HNMR spectra;

[0035] Figure 4 is the intermediate 3 in Example 1 of the present invention 1 HNMR spectra;

[0036] Figure 5 is the mass spectrum of intermediate 3 in Example 1 of the present invention;

[0037] Figure 6 is the intermediate 4 in Example 1 of the present invention 1 HNMR spectra;

[0038] Figure 7 is the mass spectrum of intermediate 4 in Example 1 of the present invention;

[0039] Figure 8 is the intermediate 5 in Example 1 of the present invention 1 HNMR spectra;

[0040] Fig. 9 The modified sodium pyruvate in Example 1 of the present invention 1 HNMR spectra;

[0041] Fig.10 is the mass spectrum of the modified sodium pyruvate in Example 1 of the present invention;

[0042] Fig.11 This is a result analysis diagram of the effect of the aerosol of the present invention on the body weight of rats with pulmonary fibrosis;

[0043] Fig.12 This is a result analysis diagram of the effect of the aerosol of the present invention on the TNF-α content in the serum of rats with pulmonary fibrosis;

[0044] Fig.13 This is a result analysis diagram of the effect of the aerosol of the present invention on the IL-6 content in the serum of rats with pulmonary fibrosis;

[0045] Fig.14 This is a result analysis diagram of the effect of the aerosol of the present invention on the IL-1β content in the serum of rats with pulmonary fibrosis;

[0046] Fig.15 This is a result analysis diagram of the effect of the aerosol of the present invention on the lung coefficient of rats with pulmonary fibrosis;

[0047] Fig.16 The figure is an analysis result of the effect of the aerosol of the present invention on the hydroxyproline content in the lung tissue of rats with pulmonary fibrosis. DETAILED DESCRIPTION

[0048] 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.

[0049] 1. Implementation

[0050] Example 1

[0051] Example 1 provides a sodium pyruvate aerosol for regulating respiratory microecology, which is composed of the following components in parts by weight: 0.06 parts of modified sodium pyruvate, 90 parts of tetrafluoroethane, and 5 parts of water for injection.

[0052] The preparation method of the modified sodium pyruvate is as follows:

[0053]

[0054] (1) Arofylline (10 mmol), ethyl 4-bromobutyrate (15 mmol), potassium carbonate (20 mmol), and benzyltriethylammonium chloride (5 mmol) were added to 100 mL of acetone in sequence and reacted at 60°C for 16 h. After the reaction, the reaction solution was filtered to remove the precipitate, and the filtrate was collected and concentrated and crystallized from ethanol to obtain intermediate 1;

[0055] The intermediate 1 1 HNMR Figure 1 As shown: (C 20 H 23 C1N 4 O 4 ,400MHz,DMSO- d6 )δ:0.86(t,3H),1.08(t,3H),1.55(m,2H),2.17(m,2H),2.33(m,2H),3.69(t,2H),4.02-4.05(m,4H),7.41(d,2H),7.60(d,2H),8.54(s,1H); The mass spectrum of the intermediate 1 is shown in Figure 2 As shown, [M+H] + m / z: 419.14, ESI-MS (m / z): 418.14; the above results confirmed that the obtained product was the target product.

[0056] (2) adding the intermediate 1 (10 mmol) obtained in step (1) and hydrazine hydrate (purity 85%, 50 mmol) to anhydrous ethanol (50 mL), and reacting at 80° C. for 20 h. After the reaction, cooling the reaction solution, filtering, washing with water, and recrystallizing to obtain intermediate 2;

[0057] The above intermediate 2 1 HNMR Figure 3 As shown: (C 18 H 21 C1N 6 O 4 ,400MHz,DMSO- d6)δ:0.86(t,3H),1.50-1.55(m,3H),2.05(m,2H),2.31(m,2H),3.41(s,2H),3.69(t,2H),4.05(t,2H),4.2(s,1H)7.41(d,2H),7.60(d,2H),8.54(s,1H); The above results confirmed that the obtained product was the target product.

[0058] (3) The intermediate 2 (10 mmol) obtained in step (2), 2,3,4-trihydroxybenzaldehyde (10 mmol), and 0.5 mL of concentrated hydrochloric acid (37%) were added to 100 mL of methanol, and the mixture was stirred at room temperature for 72 h. After the reaction, water was added to the reaction solution for precipitation, and the precipitate was filtered and recrystallized to obtain the intermediate 3;

[0059] The above intermediate 3 1 HNMR Figure 4 As shown: (C 25 H 25 C1N 6 O 7 ,400MHz,DMSO- d6 )δ:0.86(t,3H),1.55(m,2H),2.05(m,2H),2.31(t,2H),3.69(t,2H),4.05(t,2H),6.28(d,1H),7.11(d,1H),7.41(d,2H),7.60(d,2H),8.36(s,1H),8.54(s,1H),8.74(s,1H),9.49(s,1H),13.79(s,1H); The mass spectrum of the intermediate 3 is shown in Figure 5 Shown: [M+H] + m / z: 557.15, ESI-MS (m / z): 557.15; the above results confirmed that the obtained product was the target product.

[0060] (4) The intermediate 3 (10 mmol) obtained in step (3), tert-butyldimethylsilyl chloride (34 mmol) and imidazole (65 mmol) were added to DMF (15 mL) in sequence, stirred at 35 °C for 9 h, and extracted, dried and concentrated to obtain the intermediate 4;

[0061] The above intermediate 4 1 HNMR Figure 6 As shown: (C 43 H 67 C1N 6 O 7 Si 3 ,400MHz,DMSO- d6)δ:0.22(s,18H),0.86(t,3H),0.99(s,27H),1.51-1.55(m,3H),2.05(m,2H),2.31(t,2H),3.69(t,2H),4.05(t,2H),6.32(d,1H),7.11(d,1H),7.41(d,2H),7.60(d,2H),8.36(s,1H),8.54(s,1H); The mass spectrum of the intermediate 4 is shown in Figure 7 Shown: [M+H] + m / z: 899.41, ESI-MS (m / z): 899.41; the above results confirmed that the obtained product was the target product.

[0062] (5) The intermediate 4 (10 mmol) obtained in step (4) was mixed with copper iodide (1.1 mmol) and samarium (21 mmol) in DMF (20 mL) and stirred for 4 h. The reaction was quenched with water and the organic phase was washed with 1 M hydrochloric acid. The organic phase was dried and concentrated and then added to tetrahydrofuran (20 mL). 50 mL of 1 M tetrabutylammonium fluoride was added and stirred for 2 h. The mixture was extracted and concentrated and then purified to obtain the intermediate 5.

[0063] The above intermediate 5 1 HNMR Figure 8 As shown: (C 26 H 26 N 6 O 8 ,400MHz,DMSO- d6 )δ:0.86(t,3H),1.51-1.54(m,3H),2.05(m,2H),2.31(t,2H),3.69(t,2H),4.05(t,2H),6.28(d,1H),7.11(d,1H),7.43(d,2H),7.85(d,2H),8.36(s,1H),8.54(s,21H),8.74(s,1H),9.49(s,1H),9.90(s,1H),13.79(s,1H); The above results confirmed that the obtained product was the target product.

[0064] (6) Sodium hydroxide was dissolved in a mixed solvent of water and ethanol in a volume ratio of 1:1 to obtain 100 mL of a sodium hydroxide solution with a final concentration of 0.2 M, to which sodium pyruvate (20 mmol) and the intermediate 5 (10 mmol) obtained in step (5) were added, and the mixture was stirred at room temperature for 1 h. After the reaction was completed, the reaction solution was filtered, and the precipitate was collected and washed with a mixed solution of water and ethanol (the volume ratio of water to ethanol was 1:1), and then dried to obtain modified sodium pyruvate.

[0065] The modified sodium pyruvate 1 HNMR Fig. 9 As shown: (C 29 H 27 N 6 NaO 10 ,400MHz,DMSO- d6 )δ:0.86(t,3H),1.50-1.55(m,3H),2.05(m,2H),2.31(t,2H),3.69(t,2H),4.05(t,2H),6.28(d,1H),6.68(d,1H),7.11(d,1H),7.61(m,2H),7.71(m,1H),7.92(d,2H),8.40(s,1H),8.54(s,1H),8.74(s,1H),9.49(s,1H),13.79(s,1H); The mass spectrum of the above modified sodium pyruvate is shown in Fig.10 Shown: [M+H] + m / z: 643.17, ESI-MS (m / z): 643.17; the above results confirmed that the obtained product was the target product.

[0066] The preparation method of the above-mentioned sodium pyruvate aerosol for regulating respiratory microecology is as follows:

[0067] The modified sodium pyruvate and water for injection are compounded according to the above weight ratio, filled, and pressed into tetrafluoroethane to obtain the product.

[0068] Example 2

[0069] Example 2 provides a sodium pyruvate aerosol for regulating respiratory microecology, which is composed of the following components in parts by weight: 0.05 parts of modified sodium pyruvate, 95 parts of tetrafluoroethane, and 6 parts of water for injection.

[0070] The preparation method of the modified sodium pyruvate is as follows:

[0071]

[0072] (1) Arofylline (10 mmol), ethyl 4-bromobutyrate (20 mmol), potassium carbonate (25 mmol), and benzyltriethylammonium chloride (10 mmol) were added to 110 mL of acetone in sequence and reacted at 55°C for 17 h. After the reaction, the reaction solution was filtered to remove the precipitate, and the filtrate was collected and concentrated and crystallized from ethanol to obtain intermediate 1;

[0073] (2) The intermediate 1 (10 mmol) obtained in step (1) and hydrazine hydrate (purity 85%, 40 mmol) were added to anhydrous ethanol (50 mL), and the mixture was reacted at 85° C. for 19 h. After the reaction was completed, the reaction solution was cooled and filtered, washed with water, and recrystallized to obtain the intermediate 2;

[0074] (3) The intermediate 2 (10 mmol) obtained in step (2), 2,3,4-trihydroxybenzaldehyde (15 mmol), and 0.4 mL of concentrated hydrochloric acid (37%) were added to 100 mL of methanol, and the mixture was stirred at room temperature for 74 h. After the reaction, water was added to the reaction solution for precipitation, and the precipitate was filtered and recrystallized to obtain the intermediate 3;

[0075] (4) The intermediate 3 (10 mmol) obtained in step (3), tert-butyldimethylsilyl chloride (32 mmol) and imidazole (60 mmol) were added to DMF (15 mL) in sequence, stirred at 32 °C for 10 h, and extracted, dried and concentrated to obtain the intermediate 4;

[0076] (5) The intermediate 4 (10 mmol) obtained in step (4) was mixed with copper iodide (1 mmol) and samarium (20 mmol) in DMF (20 mL) and stirred for 3 h. The reaction was quenched with water and the organic phase was washed with 1 M hydrochloric acid. The organic phase was dried and concentrated and then added to tetrahydrofuran (20 mL). 45 mL of 1 M tetrabutylammonium fluoride was added to the tetrahydrofuran and stirred for 2.5 h. The mixture was extracted and concentrated and then purified to obtain the intermediate 5.

[0077] (6) Sodium hydroxide was dissolved in a mixed solvent of water and ethanol in a volume ratio of 1:1 to obtain 100 mL of a sodium hydroxide solution with a final concentration of 0.22 M, to which sodium pyruvate (25 mmol) and the intermediate 5 (10 mmol) obtained in step (5) were added, and the mixture was stirred at room temperature for 1.2 h. After the reaction was completed, the reaction solution was filtered, and the precipitate was collected and washed with a mixed solution of water and ethanol (the volume ratio of water to ethanol was 1:1), and then dried to obtain modified sodium pyruvate.

[0078] The preparation method of the above-mentioned sodium pyruvate aerosol for regulating respiratory microecology is as follows:

[0079] The modified sodium pyruvate and water for injection are compounded according to the above weight ratio, filled, and pressed into tetrafluoroethane to obtain the product.

[0080] Example 3

[0081] Example 3 provides a sodium pyruvate aerosol for regulating respiratory microecology, which is composed of the following components in parts by weight: 0.1 parts of modified sodium pyruvate, 80 parts of heptafluoropropane, and 10 parts of water for injection.

[0082] The preparation method of the modified sodium pyruvate is as follows:

[0083]

[0084] (1) Arofylline (10 mmol), ethyl 4-bromobutyrate (18 mmol), potassium carbonate (22 mmol), and benzyltriethylammonium chloride (8 mmol) were added to 90 mL of acetone in sequence and reacted at 65°C for 15 h. After the reaction, the reaction solution was filtered to remove the precipitate, and the filtrate was collected and concentrated and crystallized from ethanol to obtain intermediate 1;

[0085] (2) The intermediate 1 (10 mmol) obtained in step (1) and hydrazine hydrate (purity 85%, 60 mmol) were added to anhydrous ethanol (50 mL), and the mixture was reacted at 75° C. for 21 h. After the reaction was completed, the reaction solution was cooled and filtered, washed with water, and recrystallized to obtain the intermediate 2;

[0086] (3) The intermediate 2 (10 mmol) obtained in step (2), 2,3,4-trihydroxybenzaldehyde (20 mmol), and 0.6 mL of concentrated hydrochloric acid (37%) were added to 100 mL of methanol, and the mixture was stirred at room temperature for 70 h. After the reaction, water was added to the reaction solution for precipitation, and the precipitate was filtered and recrystallized to obtain the intermediate 3;

[0087] (4) The intermediate 3 (10 mmol) obtained in step (3), tert-butyldimethylsilyl chloride (36 mmol) and imidazole (70 mmol) were added to DMF (15 mL) in sequence, stirred at 38 °C for 8 h, and extracted, dried and concentrated to obtain the intermediate 4;

[0088] (5) The intermediate 4 (10 mmol) obtained in step (4) was mixed with copper iodide (1.2 mmol) and samarium (22 mmol) in DMF (20 mL) and stirred for 5 h. The reaction was quenched with water and the organic phase was washed with 1 M hydrochloric acid. The organic phase was dried and concentrated and then added to tetrahydrofuran (20 mL). 45 mL of 1 M tetrabutylammonium fluoride was added to the tetrahydrofuran and stirred for 3 h. The mixture was extracted, concentrated and purified to obtain the intermediate 5.

[0089] (6) Sodium hydroxide was dissolved in a mixed solvent of water and ethanol in a volume ratio of 1:1 to obtain 100 mL of a sodium hydroxide solution with a final concentration of 0.18 M, to which sodium pyruvate (25 mmol) and the intermediate 5 (10 mmol) obtained in step (5) were added, and the mixture was stirred at room temperature for 0.8 h. After the reaction was completed, the reaction solution was filtered, and the precipitate was collected and washed with a mixed solution of water and ethanol (the volume ratio of water to ethanol was 1:1), and then dried to obtain modified sodium pyruvate.

[0090] The preparation method of the above-mentioned sodium pyruvate aerosol for regulating respiratory microecology is as follows:

[0091] The modified sodium pyruvate and water for injection are compounded according to the above weight ratio, filled, and pressed into heptafluoropropane to obtain the product.

[0092] (II) Comparative Example

[0093] Comparative Example 1

[0094] Comparative Example 1 provides a drug, arofylline original drug.

[0095] Comparative Example 2

[0096] Comparative Example 2 provides a sodium pyruvate aerosol, which is composed of the following components in parts by weight: 0.06 parts of sodium pyruvate, 90 parts of tetrafluoroethane, and 5 parts of water for injection.

[0097] (III) Experimental examples

[0098] Experimental Example 1 Animal Experiment

[0099] 1. Animal grouping, pulmonary fibrosis rat modeling, and drug administration

[0100] SD rats, weighing 210-220g, a total of 70 were randomly divided into 7 groups, with a male-female ratio of 1:1 in each group, and adaptive feeding for 5 days, one of which was a blank control group. Each group of mice was intraperitoneally injected with 4% chloral hydrate, with an injection volume of 10mL / kg, and the rats in each group were anesthetized and fixed in a supine position, the neck hair was removed, and the trachea was exposed by a longitudinal incision. Except for the blank control group, the other groups of rats used a syringe to pierce the trachea 1cm through the gap between the two tracheal cartilage rings toward the heart end, and bleomycin (dose of 5mg / kg) was injected into the trachea after no resistance was withdrawn. The blank control group was injected with an equal volume of normal saline. After the injection, the rats in each group were rotated upright for 3min to evenly distribute the drug in the two lungs, and each layer of tissue was sutured.

[0101] 24 hours after modeling, the rats in the blank group and the model group were gavaged with an equal amount of normal saline, the rats in Example 1-3 were given 0.6 mg / kg of each preparation by atomization (i.e., the amount of modified sodium pyruvate in the aerosol was 0.6 mg / kg), the rats in the comparative example 1 group were gavaged with 0.6 mg / kg of arofylline, and the rats in the comparative example 2 group were given 0.6 mg / kg of the preparation by atomization (i.e., the amount of sodium pyruvate in the aerosol was 0.6 mg / kg), and the administration was continued for 28 days.

[0102] 2. Detection indicators and methods

[0103] 2.1 Weight

[0104] The body weight of rats in each group was weighed on the day of administration and recorded as the initial body weight. The body weight of rats in each group was weighed again every 7 days. The experimental results are the average values ​​of rats in each group. Fig.11 , Table 1.

[0105] 2.2 Serum inflammatory factor levels

[0106] On the 28th day, whole blood was collected from the eyeballs of mice in each group, and the levels of IL-6, IL-1β, and TNF-α in serum were detected using ELISA kits. The experimental results are the average values ​​of each group of rats. The results are shown in Figure 12-14 .

[0107] 2.3 Lung coefficient

[0108] On the 14th and 28th days, 5 rats in each group were killed, lung tissues were removed and weighed, and the lung coefficient was calculated. Lung coefficient = lung wet weight (mg) / body weight (mg) × 100%. The experimental results are the average values ​​of rats in each group. The results are shown in Fig.15 .

[0109] 2.4 Hydroxyproline content in lung tissue

[0110] The remaining 5 rats in each group were killed 28 days after intragastric administration, and the hydroxyproline content in the lung tissue was detected using a hydroxyproline content detection kit. The experimental results are the average values ​​of the rats in each group. The results are shown in Fig.16 .

[0111] 3. Test results

[0112] 3.1 Effect of the aerosol of the present invention on the body weight of rats with pulmonary fibrosis

[0113] Table 1

[0114]

[0115] observe Fig.11 As shown in Table 1, compared with the blank control group, the body weight of the rats in the model group decreased significantly on the 21st day. Compared with the model group, the body weight of the rats in Example 1-3 groups increased significantly. The above results show that the aerosol prepared by the present invention can improve the physique of rats with pulmonary fibrosis model. Compared with the comparative example 1-2 group, the body weight of the rats in Example 1-3 groups increased significantly. The above results show that compared with the use of arofylline and sodium pyruvate alone, the aerosol prepared by using the modified sodium pyruvate of the present application can effectively alleviate the effect of pulmonary fibrosis on rats.

[0116] 3.2 Effect of the aerosol of the present invention on the levels of inflammatory factors in the serum of rats with pulmonary fibrosis

[0117] observe Figure 12-14It can be seen that compared with the blank control group, the expression of inflammatory factors in the serum of the model group increased significantly. Compared with the model group, the expression of inflammatory factors in the serum of Example 1-3 groups was significantly reduced. Compared with the comparative example 1-2 group, the expression of inflammatory factors in the serum of Example 1-3 groups was lower. From the above, it can be seen that the aerosol of the present invention can effectively reduce the expression of inflammatory factors IL-6, IL-1β, and TNF-α in serum, thereby playing an anti-pulmonary fibrosis role.

[0118] 3.3 Effect of the aerosol of the present invention on the lung coefficient of rats with pulmonary fibrosis

[0119] observe Fig.15 It can be seen that the lung coefficient of the model group rats increased significantly compared with the blank control group. After 14d and 28d of administration, the lung coefficient of the rats in Example 1-3 groups was significantly reduced compared with the model group. The above results show that the aerosol of the present invention can effectively improve the pulmonary fibrosis of rats. It can be seen from the above that the aerosol of the present invention can effectively alleviate the development of pulmonary fibrosis in model rats.

[0120] 3.4 Effect of the aerosol of the present invention on the hydroxyproline content in lung tissue of rats with pulmonary fibrosis

[0121] The hydroxyproline content in lung tissue is one of the important indicators for judging the degree of lung fibrosis. Fig.16 It can be seen that the hydroxyproline content in the lung tissue of the model group rats was significantly increased compared with the blank control group. The hydroxyproline content in the lung tissue of the rats in Example 1-3 groups was significantly reduced compared with the model group. The above results show that the aerosol of the present invention can effectively alleviate the development of pulmonary fibrosis in model rats.

[0122] Experimental Example 2 Clinical Experiment

[0123] 1. Experimental subjects, grouping, and drug administration

[0124] According to the patients' medical history and clinically relevant treatment, the present invention included a total of 50 patients with acute exacerbation of COPD and randomly divided them into 2 groups. One group was a blank control group, and the patients did not use the aerosol of Example 1 after discharge; the other group started to use the aerosol of Example 1 once a day after discharge, and were followed up for a total of 6 months.

[0125] 2. Detection indicators and methods

[0126] The TGuide S96 magnetic bead lavage fluid genomic DNA extraction kit was used to extract nucleic acids, and the 16srRNA detection technology was used to detect the extract. The primers were designed as follows: the bacterial universal primer 16s was used to amplify the V3+V4 region of the sample 16srDNA gene; the amplification region of the sample fungus was the ITS1 region, and the primer design is shown in Table 2:

[0127] Table 2

[0128]

[0129] The bacterial species and abundance were confirmed by comparing with known sequences in the database. The results are shown in Table 3-4.

[0130] 3. Test results

[0131] The effects of the aerosol of the present invention on the changes in airway bacterial colonies are shown in Tables 3-4.

[0132] Table 3. Bacterial abundance at the gate level before and after medication

[0133]

[0134] Table 4 Genera-level bacterial abundance before and after medication

[0135]

[0136] It can be seen from Table 3 that the top 10 bacterial phyla in abundance in the alveolar lavage fluid before and after the treatment with the aerosol of the present invention are the same, among which Proteobacteria (Proteobacteria), Cynaobacteria (Cyanobacteria), Bacteroidetes (Bacteroidetes), Actinobacteria (Actinomycetes), Fusobacteria (Fusobacteria), Firmicutes (Firmicutes) abundance accounted for 94%, and after drug use Cynaobacteria (Cyanobacteria), Actinobacteria (Actinomycetes), Firmicutes (Firmicutes) increased in abundance, Proteobacteria (Proteobacteria), Bacteroidetes (Bacteroidetes) decreased in abundance.

[0137] It can be seen from Table 4 that the top 10 bacterial genera in abundance in the alveolar lavage fluid before and after treatment with the aerosol of the present invention are the same. Haemophilus (Haemophilus spp.) decreased significantly, while Streptococcus (Streptococcus), Lactobacillus (Lactobacillus spp.) increased in abundance.

[0138] As shown in Tables 3-4, after using the aerosol of the present invention, the proportion of Firmicutes increased significantly, and the proportion of Streptococcus and Lactobacillus at the genus level also increased; the proportion of Proteobacteria decreased significantly, and the proportion of Haemophilus at the genus level decreased significantly. In summary, the aerosol of the present invention can reduce the abundance of pathogenic bacteria and increase the abundance of probiotics.

[0139] 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 sodium pyruvate aerosol for regulating respiratory microecology, characterized in that: The method comprises the following components in parts by weight: 0.05-0.1 parts of modified sodium pyruvate, 80-95 parts of propellant, and 5-10 parts of water for injection; The preparation process of the modified sodium pyruvate comprises the following steps: (1) reacting arofylline with ethyl 4-bromobutyrate under a catalyst condition, and after the reaction is completed, purifying the reaction solution to obtain intermediate 1; (2) reacting the intermediate 1 obtained in step (1) with hydrazine hydrate, and after the reaction is completed, purifying the reaction solution to obtain the intermediate 2; (3) Under catalyst conditions, the intermediate 2 obtained in step (2) is stirred and reacted with a 2,3,4-trihydroxybenzaldehyde solution. After the reaction is completed, the reaction solution is purified to obtain an intermediate 3; (4) adding the intermediate 3 obtained in step (3), tert-butyldimethylsilyl chloride and imidazole to DMF, mixing and stirring for reaction. After the reaction is completed, the reaction solution is purified to obtain the intermediate 4; (5) The intermediate 4 obtained in step (4) is added to DMF, copper iodide and samarium, and the mixture is stirred for 3-5 hours; water is added to the reaction solution to quench it, and then an organic phase is obtained by extraction; the organic phase is dried and concentrated, and then added to tetrahydrofuran, and then tetrabutylammonium fluoride is added, and the reaction is stirred. After the reaction is completed, the reaction solution is purified to obtain the intermediate 5; (6) Adding sodium pyruvate and the intermediate 5 obtained in step (5) into a sodium hydroxide solution, stirring the mixture for reaction, and after the reaction is completed, purifying the reaction solution to obtain modified sodium pyruvate.

2. The sodium pyruvate aerosol for regulating respiratory microecology according to claim 1, characterized in that: The catalyst in step (1) is potassium carbonate and benzyltriethylammonium chloride; the molar ratio of arofylline, ethyl 4-bromobutyrate, potassium carbonate and benzyltriethylammonium chloride is 1:1.5-2:2-2.5:0.5-1; the reaction temperature is 55-65°C and the reaction time is 15-17h.

3. The sodium pyruvate aerosol for regulating respiratory microecology according to claim 1, characterized in that: In step (2), the molar ratio of the intermediate 1 to hydrazine hydrate is 1:4-6; the reaction temperature is 75-85°C and the reaction time is 19-21h.

4. The sodium pyruvate aerosol for regulating respiratory microecology according to claim 1, characterized in that: In step (3), the ratio of the intermediate 2 to 2,3,4-trihydroxybenzaldehyde and the catalyst is 1 mmol: 1-2 mmol: 0.04-0.06 mL; the catalyst is 37% concentrated hydrochloric acid; and the stirring reaction time is 70-74 h.

5. The sodium pyruvate aerosol for regulating respiratory microecology according to claim 1, characterized in that: In step (4), the molar ratio of the intermediate 3, tert-butyldimethylsilyl chloride, and imidazole is 1:3.2-3.6:6-7, the stirring reaction temperature is 32-38°C, and the stirring time is 8-10h; in step (5), the molar ratio of the intermediate 4, copper iodide, samarium, and tetrabutylammonium fluoride is 1:0.1-0.12:2-2.2:4.5-5.0, and the stirring reaction time is 2-3h.

6. The sodium pyruvate aerosol for regulating respiratory microecology according to claim 1, characterized in that: In step (6), the molar ratio of the intermediate 5, sodium pyruvate and sodium hydroxide is 1:2-2.5:1.8-2.2; and the stirring reaction time is 0.8-1.2h.

7. The sodium pyruvate aerosol for regulating respiratory microecology according to claim 1, characterized in that: The propellant is tetrafluoroethane or heptafluoropropane.

8. The method for preparing the sodium pyruvate aerosol for regulating respiratory microecology according to any one of claims 1 to 7, characterized in that: The following steps are involved: The modified sodium pyruvate and water for injection are compounded according to the weight ratio, filled, and pressed into the propellant to obtain the product.

9. Use of the sodium pyruvate aerosol for regulating respiratory microecology according to claim 1 in the preparation of a medicament for treating pulmonary fibrosis.

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