A berberine derivative, its preparation method and application

Abstract

This invention belongs to the field of biomedical technology, and provides a berberine derivative, its preparation method, and its application. The invention involves reacting 3-cyclopentadienylpropionic acid, a first organic solvent, oxalyl chloride, and anhydrous DMF to obtain intermediate I; subjecting berberine hydrochloride to high-temperature negative pressure pyrolysis, followed by free radical substitution reaction with an alcohol to obtain intermediate II; and mixing intermediate II, a second organic solvent, intermediate I, and pyridine to obtain the berberine derivative. The berberine derivative of this invention possesses broad-spectrum antibacterial activity and high biosafety, exhibiting excellent antibacterial and bacteriostatic activity against drug-resistant Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli. It overcomes the limitations of traditional berberine drugs, becoming a new generation of antibacterial drugs, meeting the current clinical demand for antibiotic alternatives, and providing an innovative treatment solution for addressing bacterial resistance.

Description

Technical Field

[0001] This invention relates to the field of biomedicine, and in particular to a berberine derivative, its preparation method, and its application. Background Technology

[0002] With the increasing prevalence of bacterial resistance globally, the development of antimicrobial drugs faces severe challenges. The widespread use of traditional antibiotics has led to drug resistance in various pathogens, particularly the rapid spread of resistant strains in hospital settings, significantly reducing the effectiveness of clinical treatment. Existing antimicrobial drugs show decreased efficacy against novel resistant strains, making many common infectious diseases more difficult to control. Therefore, developing novel antimicrobial drugs has become an urgent task for the pharmaceutical industry and global public health. Berberine, a natural alkaloid widely found in traditional Chinese medicine, possesses various pharmacological activities, particularly showing significant effects in antibacterial, antiviral, and anti-inflammatory aspects, and holds promise as a potential antimicrobial drug. However, due to its poor water solubility, poor absorption, and rapid metabolism in the intestines, berberine achieves low effective concentrations in vivo, resulting in low bioavailability and greatly limiting its antimicrobial activity and application. Furthermore, berberine has a narrow antimicrobial spectrum, primarily targeting Gram-negative bacteria in the intestines, with weaker effects against Gram-positive bacteria and other bacteria. Therefore, the application of berberine in antimicrobial therapy remains significantly limited.

[0003] Therefore, there is an urgent need to provide a novel berberine derivative with broad-spectrum antibacterial activity, high bioavailability, effectiveness against drug-resistant strains, and low toxicity, in order to solve the problem of bacterial resistance in clinical treatment with existing antibacterial drugs. Summary of the Invention

[0004] In view of this, the present invention provides a berberine derivative, its preparation method and application, to solve the problem of bacterial resistance in clinical treatment.

[0005] To achieve the above objectives, the present invention adopts the following technical solution:

[0006] This invention provides a berberine derivative, the structural formula of which is as follows:

[0007]

[0008] This invention also provides a method for preparing berberine derivatives, comprising the following steps:

[0009] 1) 3-Cyclopentahexane propionic acid, the first organic solvent, oxalyl chloride, and anhydrous DMF were mixed and reacted to obtain intermediate I;

[0010] 2) Berberine hydrochloride was subjected to high-temperature negative pressure pyrolysis, and then mixed with alcohol to undergo free radical substitution reaction to obtain intermediate II;

[0011] 3) Intermediate II, the second organic solvent, intermediate I and pyridine are mixed and reacted to obtain the berberine derivative of formula I;

[0012] There is no specific order requirement for steps 1) and 2).

[0013] The structural formula of intermediate I is as follows:

[0014] The structural formula of intermediate II is as follows:

[0015] Preferably, the mass-to-volume ratio of 3-cyclopentadienylpropionic acid, the first organic solvent, oxalyl chloride, and anhydrous DMF in step 1) is 100 mg: 5-10 mL: 178 mg: 5-10 mL; the first organic solvent includes one or more of anhydrous dichloromethane, anhydrous dichloroethane, anhydrous diethyl ether, and anhydrous chloroform.

[0016] Preferably, the reaction temperature in step 1) is -20 to 0°C and the time is 1 to 2 hours.

[0017] Preferably, the mass-to-volume ratio of berberine hydrochloride and alcohol in step 2) is 1 g: 50-100 mL; the alcohol includes one or both of anhydrous ethanol and anhydrous methanol.

[0018] Preferably, the temperature of the high-temperature negative pressure pyrolysis reaction in step 2) is 200-300℃, the pressure is -0.01--0.05 MPa, and the time is 30-60 min; the time of the free radical substitution reaction is 3-5 min.

[0019] Preferably, in step 3), the mass-to-volume ratio of intermediate II, the second organic solvent, intermediate I, and pyridine is 100 mg: 5–10 mL: 0.2–0.4 mL: 0.2–0.3 mL; the second organic solvent includes one or more of anhydrous acetonitrile, anhydrous DMF, and anhydrous dioxane.

[0020] Preferably, the reaction temperature in step 3) is 20-30°C and the time is 1-2 hours.

[0021] The present invention also provides the application of the berberine derivative prepared by the above-mentioned method in the preparation of antibacterial drugs.

[0022] As can be seen from the above technical solution, compared with the prior art, the beneficial effects of the present invention are as follows:

[0023] This invention modifies the structure of berberine, resulting in a berberine derivative with broad-spectrum antibacterial activity and excellent antibacterial and bacteriostatic activity against methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, and Escherichia coli. Furthermore, compared to berberine (BBR) and currently first-line clinical drugs ampicillin and vancomycin, the berberine derivative of this invention exhibits superior antibacterial activity against MRSA, P. aeruginosa, and E. coli. Simultaneously, the berberine derivative of this invention has higher biocompatibility than berberine (BBR), better promoting wound healing in vitro caused by MRSA and improving the therapeutic effect on pneumonia caused by P. aeruginosa bacteria in vivo.

[0024] The berberine derivatives described in this invention can overcome the limitations of traditional berberine drugs, becoming a new generation of antibacterial drugs, meeting the current clinical demand for antibiotic alternatives, and providing innovative treatment solutions for solving the problem of bacterial resistance. Attached Figure Description

[0025] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.

[0026] Figure 1 This is a schematic diagram illustrating the wound healing effect of 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1 on MRSA-infected wounds. Figure 1 In the diagram, 'a' represents the wound healing process. Figure 1 In the diagram, b represents the wound closure trajectory.

[0027] Figure 2 This is a quantitative analysis diagram of the changes in wound area in the repair of MRSA-infected wounds by 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1;

[0028] Figure 3 This is a diagram showing the bacterial count in the wound of mice after MRSA infection in Experiment Example 2.

[0029] Figure 4 The graph shows the bacterial viability of 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1 on day 11 of MRSA-infected wound repair.

[0030] Figure 5The image shows the bacterial load in the lungs of mice infected with P. aeruginosa after treatment with 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1, where the vertical axis represents the bacterial load.

[0031] Figure 6 The graph shows the total protein concentration in the bronchoalveolar lavage fluid of mice infected with P. aeruginosa bacterial pneumonia after treatment with 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1.

[0032] Figure 7 The image shows the total cell count in the bronchoalveolar lavage fluid of mice infected with P. aeruginosa bacterial pneumonia after treatment with 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1.

[0033] Figure 8 The graph shows the change in the wet / dry weight ratio of the lungs of mice infected with P. aeruginosa bacterial pneumonia after treatment with 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1.

[0034] Figure 9 The image shows the alveolar structure of mice infected with P. aeruginosa bacterial pneumonia after treatment with 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1. Figure 9 In the diagram, 'a' represents an alveolar structure diagram with a scale of 50 μm. Figure 9 In the image, b is a magnified view of a, with a scale bar of 20 μm.

[0035] Figure 10 The image shows the degree of lung damage in mice infected with P. aeruginosa bacterial pneumonia after treatment with 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1.

[0036] Figure 11 The graph shows the changes in body weight of mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4.

[0037] Figure 12 Figure showing the heart weight of mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4;

[0038] Figure 13 The image shows the liver weight of mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4.

[0039] Figure 14 Figure showing the spleen weights of mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4;

[0040] Figure 15 Figure showing the lung weights of mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4;

[0041] Figure 16 The image shows the kidney weights of mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4.

[0042] Figure 17 The graph shows the changes in serum alanine aminotransferase (ALT) levels in mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4.

[0043] Figure 18 The graph shows the changes in serum aspartate aminotransferase (AST) levels in mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4.

[0044] Figure 19 The graph shows the changes in serum creatinine (Cr) levels in mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4.

[0045] Figure 20 The image shows the morphological changes in the liver and kidney tissues of mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine in Experiment Example 4. Detailed Implementation

[0046] This invention provides a berberine derivative, wherein the berberine derivative is 9-(3'-cyclopentylpropionyloxy)berberine, and its structural formula is as follows:

[0047]

[0048] This invention also provides a method for preparing berberine derivatives, comprising the following steps:

[0049] 1) 3-Cyclopentahexane propionic acid, the first organic solvent, oxalyl chloride, and anhydrous DMF were mixed and reacted to obtain intermediate I;

[0050] 2) Berberine hydrochloride was subjected to high-temperature negative pressure pyrolysis, and then mixed with alcohol to undergo free radical substitution reaction to obtain intermediate II;

[0051] 3) Intermediate II, the second organic solvent, intermediate I and pyridine are mixed and reacted to obtain the berberine derivative of formula I;

[0052] There is no specific order requirement for steps 1) and 2).

[0053] The structural formula of intermediate I is as follows:

[0054] The structural formula of intermediate II is as follows:

[0055] In this invention, the synthetic route for the berberine derivative is as follows:

[0056]

[0057] In this invention, the mass-to-volume ratio of 3-cyclopentadienylpropionic acid, the first organic solvent, oxalyl chloride and anhydrous DMF in step 1) is 100 mg: 5-10 mL: 178 mg: 5-10 mL, more preferably 100 mg: 5 mL: 178 mg: 5 mL.

[0058] In this invention, the first organic solvent in step 1) includes one or more of anhydrous dichloromethane, anhydrous dichloroethane, anhydrous diethyl ether, and anhydrous chloroform.

[0059] In this invention, the reaction temperature in step 1) is -20 to 0°C, preferably -18 to -1°C, more preferably -15 to -2°C; the reaction time is 1 to 2 hours, more preferably 1.5 hours.

[0060] In this invention, the intermediate I described in step 1) is oily.

[0061] In this invention, the mass-to-volume ratio of berberine hydrochloride and alcohol in step 2) is 1g:50-100mL, preferably 1g:55-85mL, and more preferably 1g:60-70mL; the alcohol includes one or both of anhydrous ethanol and anhydrous methanol, wherein the alcohol can provide hydroxyl groups for the reaction.

[0062] In this invention, the temperature of the high-temperature negative pressure pyrolysis reaction in step 2) is 200-300°C, preferably 220-280°C, and more preferably 230°C; the pressure of the high-temperature negative pressure pyrolysis reaction is -0.01--0.05 MPa, preferably -0.02--0.04 MPa, and more preferably -0.03 MPa; the time of the high-temperature negative pressure pyrolysis reaction is 30-60 min, preferably 35-55 min, and more preferably 40 min; wherein, the high-temperature negative pressure pyrolysis reaction can break the methyl bond of the benzene ring methoxy group.

[0063] In this invention, the free radical substitution reaction in step 2) takes 3 to 5 minutes, more preferably 4 minutes.

[0064] In this invention, the intermediate II mentioned in step 2) is a red solid.

[0065] In this invention, the mass-to-volume ratio of intermediate II, the second organic solvent, intermediate I and pyridine in step 3) is 100 mg: 5-10 mL: 0.2-0.4 mL: 0.2-0.3 mL, more preferably 100 mg: 10 mL: 0.4 mL: 0.3 mL.

[0066] In this invention, the second organic solvent in step 3) includes one or more of anhydrous acetonitrile, anhydrous DMF, and anhydrous dioxane.

[0067] In this invention, the reaction temperature in step 3) is 20-30°C, preferably 22-28°C, more preferably 25°C; the reaction time is 1-2 hours, more preferably 1.5 hours.

[0068] In this invention, after the reaction described in step 3) is completed, the product 9-(3'-cyclopentylpropionyloxy)berberine is further subjected to filtration and washing operations.

[0069] In this invention, the solvent used for washing preferably includes one or more of anhydrous dichloromethane, anhydrous ethyl acetate, and anhydrous diethyl ether; the number of washing cycles is preferably 2 to 5, more preferably 3.

[0070] In this invention, the 9-(3'-cyclopentylpropionyloxy)berberine is a yellow solid.

[0071] The present invention also provides the application of the berberine derivative prepared by the above-mentioned method in the preparation of antibacterial drugs.

[0072] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0073] Example 1

[0074] 1) Add 3-cyclopentadienylpropionic acid (100 mg) to 5 mL of anhydrous CH2Cl2, stir at 0 °C for 5 min, then add oxaloyl chloride (178 mg) and anhydrous DMF (5 mL), react at 0 °C for 1 h, and then evaporate to dryness using a vacuum rotary evaporator to obtain oily intermediate I.

[0075] 2) Berberine hydrochloride (100 mg) was placed in a round-bottom flask and stirred for 30 min under negative pressure (-0.01 MPa) at 200 °C. After the reaction was completed, it was cooled to room temperature and 5 mL of anhydrous ethanol was added to mix and dissolve the mixture for free radical substitution reaction for 4 min. The mixture was then concentrated using a vacuum rotary evaporator and purified by column chromatography to obtain red solid intermediate II.

[0076] 3) Dissolve 50 mg of intermediate II in 5 mL of anhydrous acetonitrile, transfer the solution to a round-bottom flask containing 0.2 mL of intermediate I, add 3 drops of pyridine, react at 25 °C for 1 h, filter, and wash the precipitate three times with anhydrous dichloromethane solvent to obtain the yellow solid product 9-(3'-cyclopentylpropionyloxy)berberine.

[0077] The NMR data for 9-(3'-cyclopentylpropionyloxy)berberine are as follows:

[0078] 1 H NMR (400MHz, DMSO-d) 6) δ10.00(s,1H),9.06(s,1H),8.27(d,J=9.3Hz,1H),8.21(d,J=9.2Hz,1H),7. 81(s,1H),7.09(s,1H),6.17(s,2H),4.98(t,J=5.9Hz,2H),4.02(s,3H),3.22 (t,J=6.1Hz,2H),2.88(t,J=7.6Hz,2H),1.98–1.89(m,1H),1.87–1.79(m,2H) ,1.79–1.72(m,2H),1.67–1.59(m,2H),1.58–1.49(m,2H),1.23–1.13(m,2H).

[0079] 13 C NMR(101MHz,DMSO-d6)δ171.20,150.86,150.45,148.19,144.93,138.55,134.10,133.42,131.33,127.15, 126.36,121.66,121.08,120.85,108.91,106.03,102.62,57.72,55.73,33.05,32.44,30.76,26.67,25.23.

[0080] Experimental Example 1

[0081] The minimum inhibitory concentration (MIC) of 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1 was determined. The specific determination steps are as follows:

[0082] In fresh MH broth, overnight cultures of methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, and Escherichia coli were inoculated at a bacterial-to-MH broth ratio of 1%, resulting in three cultures containing different bacteria. These cultures were then incubated at 37°C and 200 rpm until mid-exponential growth was achieved. Subsequently, all three cultures were diluted to 7.5 × 10⁻⁶. 5CFU / mL was prepared for use. Next, 25.6 mg / mL of 9-(3'-cyclopentylpropionyloxy)berberine solution was serially diluted twofold in 96-well plates with fresh MH broth, with each well containing 100 μL. Then, 100 μL of bacterial suspension (one of the three cultures containing different bacteria) was added to each well, gently shaken to mix, and incubated at 37°C horizontally for 12 h. Finally, the presence of indicator bacteria was visually observed at the bottom of the wells. The lowest concentration with no indicator bacteria growth was taken as the minimum inhibitory concentration (MIC) of 9-(3'-cyclopentylpropionyloxy)berberine. In addition, the same experimental procedure was used to determine the MIC of berberine, ampicillin, and vancomycin. The experimental results are shown in Table 1.

[0083] Table 1 Minimum Inhibitory Concentration

[0084]

[0085]

[0086] As shown in Table 1 above, 9-(3'-cyclopentylpropionyloxy)berberine exhibits significantly better antibacterial activity against drug-resistant Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli than berberine and the currently first-line clinical drugs ampicillin and vancomycin.

[0087] Experimental Example 2

[0088] The therapeutic effect of 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1 on wound repair in MRSA infection was determined. The specific determination steps are as follows:

[0089] Six- to eight-week-old male ICR mice were housed for one week to allow them to acclimatize. Afterward, they were randomly assigned to five groups (n=7 mice per group): a control group, a negative control group (blank gel group), a 0.5 μg / mL 9-(3'-cyclopentylpropionyloxy)berberine treatment group (low-dose group containing gel I), a 1 μg / mL 9-(3'-cyclopentylpropionyloxy)berberine treatment group (medium-dose group containing gel I), and a 2 μg / mL 9-(3'-cyclopentylpropionyloxy)berberine treatment group (high-dose group containing gel I). Based on mouse weight, mice were anesthetized by intraperitoneal injection of 1% sodium pentobarbital solution at a dose of 0.1 mL / 10 g. Then, a 2×2 cm piloerection was performed on the mid-posterior back of the mice using a shaver. 2 Remove hair from the affected area, then apply hair removal cream with a cotton swab, leave for 10 minutes, and clean with a cotton ball moistened with water. Place the mouse on its side, and use tweezers to symmetrically pull apart the hairless skin. Use a 6mm diameter skin biopsy device to make a puncture; apply 2×10⁻⁶ hair removal samples with a cotton swab. 8CFU / mL MRSA bacterial suspension was evenly applied to the wound for inoculation; 1 hour later, all groups except the control group received the corresponding treatment; the healing status of the mouse skin wound was photographed and recorded every two days.

[0090] During treatment, the bacterial count in the wound was also measured. The specific steps were as follows: 1. 50 mg of skin around the wound was cut and weighed into a 1.5 mL centrifuge tube, and 1 mL of PBS buffer was added to homogenize the skin tissue; 2. After homogenization, the tissue was serially diluted with PBS buffer, and then 40 μL was transferred to a TSB agar plate and spread with an L-shaped spreader; 3. The plate was placed in a 37°C oven and incubated for 12 h before being counted.

[0091] The experimental results of this example are as follows: Figures 1-4 As shown, Figure 1 This is a schematic diagram illustrating the wound healing effect of 9-(3'-cyclopentylpropionyloxy)berberine on MRSA-infected wounds within 11 days. Figure 1 In the diagram, 'a' represents the wound healing process. Figure 1 In the diagram, 'b' represents the wound closure trajectory. From... Figure 1 As can be seen, the wound area of ​​all five groups gradually decreased with the increase of post-traumatic time. However, the wound shrinkage rate of the control group and the blank gel group was significantly slower than that of the 9-(3'-cyclopentylpropionyloxy)berberine administration group from day 1. Moreover, the wound repair speed of the three administration groups was positively correlated with the treatment dose of 9-(3'-cyclopentylpropionyloxy)berberine.

[0092] Figure 2 A quantitative analysis graph of wound area changes, from Figure 2 As can be seen, on day 11, the wound healing rate of the control group was 30.18±1.62%, the wound healing rate of the blank gel group was 60.21±2.01%, the wound healing rate of the low-dose gel group containing 1 was 75.49±0.87%, the wound healing rate of the medium-dose gel group containing 1 was 83.09±2.67%, and the wound healing rate of the high-dose gel group containing 1 was 91.62±3.01%. This indicates that the administration of 9-(3'-cyclopentylpropionyloxy)berberine has the function of promoting the repair of MRSA / ATCC33591 infected wounds.

[0093] Figure 3 The results of the measurement of bacterial content in the wound during treatment are from... Figure 3As can be seen, on the 11th day after the trauma, the bacterial colonies on the skin of the control group basically filled the entire culture dish, with a colony formation rate of 100%; the colony formation rate of the skin of the blank gel group was 62%; the colony formation rate of the skin of the low-dose gel group containing I was 37%; the colony formation rate of the skin of the medium-dose gel group containing I was 15%; and the colony formation rate of the skin of the high-dose gel group containing I was 8%. Figure 4 The survival rate of each bacterial group is plotted on day 11 after the trauma. Figure 4 As can be seen, the bacterial survival ability in the 9-(3'-cyclopentylpropionyloxy)berberine administration group was weak, and the bacterial survival ability decreased with the increase of the 9-(3'-cyclopentylpropionyloxy)berberine dosage. In summary, the 9-(3'-cyclopentylpropionyloxy)berberine described in this invention has excellent antibacterial properties.

[0094] Experimental Example 3

[0095] The therapeutic effect of 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1 on bacterial pneumonia caused by P. aeruginosa was determined. The specific determination steps are as follows:

[0096] Six- to eight-week-old male ICR mice were fed for one week until they adapted to their environment. They were then randomly assigned to five groups (n=7 mice per group): a control group, a Pseudomonas aeruginosa infection group (referred to as the Pseudomonas aeruginosa model group), a 50 μg / mL 9-(3'-cyclopentylpropionyloxy)berberine treatment group (referred to as the low-dose group I), a 100 μg / mL 9-(3'-cyclopentylpropionyloxy)berberine treatment group (referred to as the medium-dose group I), and a 200 μg / mL 9-(3'-cyclopentylpropionyloxy)berberine treatment group (referred to as the high-dose group I). Based on mouse body weight, mice were anesthetized by intraperitoneal injection of 1% sodium pentobarbital solution at a dose of 0.1 mL / 10 g. The trachea was then exposed. Control group mice were injected intratracheally with 50 μL of 1×PBS solution. Mice in the Pseudomonas aeruginosa model group and treatment groups (low-dose I, medium-dose I, and high-dose I) were injected intratracheally with 50 μL of 1×PBS solution. 6 CFU / mL of Pseudomonas aeruginosa bacterial suspension (resuspended in 50 μL 1×PBS) was used to keep mice upright for 2 min to allow the bacteria to enter the lungs smoothly, and the wound was sutured. 2 h later, the treatment group was injected intraperitoneally with the corresponding concentration of 9-(3'-cyclopentylpropionyloxy)berberine solution. 24 h after bacterial infection, the mice were anesthetized and sacrificed, and serum, bronchoalveolar lavage fluid and lung tissue were collected and stored in a -80℃ freezer.

[0097] The collected serum, bronchoalveolar lavage fluid, and lung tissue were tested, and the experimental results are as follows: Figures 5-10 As shown. Among them, Figure 5The graph shows the bacterial load in the lungs of mice infected with P. aeruginosa bacterial pneumonia after treatment in the control group, the P. aeruginosa model group, and the three treatment groups. Figure 5 As can be seen, compared with the Pseudomonas aeruginosa model group, the three treatment groups can significantly reduce the amount of bacteria in the lungs; Figure 6 and Figure 7 The figures show the total protein concentration and total cell count in the bronchoalveolar lavage fluid of mice with acute lung injury induced by Pseudomonas aeruginosa, respectively. Figures 6-7 As can be seen from the data, the 9-(3'-cyclopentylpropionyloxy)berberine treatment group was able to significantly inhibit the progression of inflammatory pathologies such as inflammatory cell infiltration in bronchoalveolar lavage fluid; Figure 8 A graph showing the wet / dry weight ratio of mouse lungs, from... Figure 8 As can be seen, the 9-(3'-cyclopentylpropionyloxy)berberine treatment group also reduced pulmonary edema. Figure 9 This is a diagram of the results of a pathological experiment. Figure 9 As can be seen, the alveolar structure of the lung tissue of normal control mice was intact and without obvious lesions, while the lung tissue morphology of mice in the Pseudomonas aeruginosa model group was significantly altered, with obvious thickening of the alveolar walls and a large number of inflammatory cell infiltrations. After treatment with 9-(3'-cyclopentylpropionyloxy)berberine, the lung tissue morphology of mice was significantly improved, and the alveolar wall thickness showed a concentration-dependent thinning. In the high-concentration treatment group (high-dose I administration group), the lung tissue morphology was basically restored to a level that was no different from that of normal control mice. Figure 10 The experimental results regarding the degree of lung injury, from Figure 10 It can be clearly seen that 24 hours after bacterial infection, there was a significant difference between the Pseudomonas aeruginosa model group and the normal control group (***p<0.001, where * indicates significant significance, more * indicates stronger significance, and ns indicates no significant difference), indicating that the model was successful; after treatment with 9-(3'-cyclopentylpropionoxy) drug, the degree of lung injury in the treatment group mice was much lower than that in the Pseudomonas aeruginosa model group, and the degree of lung injury was inversely proportional to the drug dose.

[0098] Experiment Example 4

[0099] The biosafety of 9-(3'-cyclopentylpropionyloxy)berberine prepared in Example 1 was evaluated, and the specific steps are as follows:

[0100] 1. Select mice with similar weight, sex, and health status, and randomly divide them into four groups (n=10 mice per group). The groups were: the administration group (30 mg / kg) receiving 100 μL of 9-(3'-cyclopentylpropionyloxy)berberine; the administration group (60 mg / kg) receiving 100 μL of 9-(3'-cyclopentylpropionyloxy)berberine; the administration group (120 mg / kg) receiving 100 μL of 9-(3'-cyclopentylpropionyloxy)berberine; and the control group receiving 100 μL of physiological saline. 2. Monitor the clinical symptoms, toxicity, and mortality of the four groups of mice for 24 hours. 3. Observe and record the weight and physical condition of the mice every day until the 7th day. On the 7th day, the mice were euthanized and their organs (heart, liver, spleen, lungs and kidneys) were removed.

[0101] Blood and biochemical tests were performed on the extracted heart, liver, spleen, lungs, and kidneys. The experimental results are as follows: Figures 11-19 As shown. Among them, Figure 11 This is a graph showing the weight monitoring of mice, from... Figure 11 As can be seen, compared with the control group, the treatment with the specified dose did not affect the weight of the mice; Figures 12-16 The images show the weights of the heart, liver, spleen, lung, and kidneys of mice that received different doses of 9-(3'-cyclopentylpropionyloxy)berberine. Figures 12-16 As can be seen, 9-(3'-cyclopentylpropionyloxy)berberine treatment had no significant effect on the organ weight (heart, lung, liver, spleen, and kidney) of mice. Figures 17-19 The images show the monitoring levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and creatinine (Cr) in mouse serum, respectively. Figures 17-19 As can be seen, treatment with low, medium, and high concentrations of 9-(3'-cyclopentylpropionyloxy)berberine did not alter serum ALT, AST, and Cr levels, indicating that 9-(3'-cyclopentylpropionyloxy)berberine treatment had no significant adverse effects on liver and kidney function. Furthermore, we used H&E staining to observe morphological changes in mouse liver and kidney tissues, such as... Figure 20 As shown, treatment of mice with a high dose of 9-(3'-cyclopentylpropionyloxy)berberine (120 mg / kg) did not result in significant changes in organ morphology. In conclusion, the 9-(3'-cyclopentylpropionyloxy)berberine described in this invention exhibits high biocompatibility.

[0102] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A berberine derivative, characterized in that, The structural formula of the berberine derivative is as follows: 。 2. The method for preparing a berberine derivative according to claim 1, characterized in that, Includes the following steps: 1) 3-Cyclopentylpropionic acid, the first organic solvent, oxalyl chloride, and anhydrous DMF were mixed and reacted to obtain intermediate I; 2) Berberine hydrochloride was subjected to high-temperature negative pressure pyrolysis, and then mixed with alcohol to undergo free radical substitution reaction to obtain intermediate II; 3) Intermediate II, the second organic solvent, intermediate I and pyridine are mixed and reacted to obtain the berberine derivative of claim 1; There is no specific order requirement for steps 1) and 2). The structural formula of intermediate I is as follows: ; The structural formula of intermediate II is as follows: .

3. The method for preparing a berberine derivative according to claim 2, characterized in that, The mass-to-volume ratio of 3-cyclopentylpropionic acid, the first organic solvent, oxalyl chloride, and anhydrous DMF in step 1) is 100 mg: 5~10 mL: 178 mg: 5~10 mL; The first organic solvent includes one or more of anhydrous dichloromethane, anhydrous dichloroethane, anhydrous diethyl ether, and anhydrous chloroform.

4. The method for preparing a berberine derivative according to claim 3, characterized in that, The reaction in step 1) is carried out at a temperature of -20 to 0°C for 1 to 2 hours.

5. A method for preparing a berberine derivative according to any one of claims 2 to 4, characterized in that, In step 2), the mass-to-volume ratio of berberine hydrochloride to alcohol is 1 g: 50-100 mL. The alcohol includes one or both of anhydrous ethanol and anhydrous methanol.

6. The method for preparing a berberine derivative according to claim 5, characterized in that, The high-temperature negative pressure pyrolysis reaction described in step 2) has a temperature of 200~300℃, a pressure of -0.01~-0.05Mpa, and a time of 30~60min; The free radical substitution reaction takes 3-5 minutes.

7. The method for preparing a berberine derivative according to claim 6, characterized in that, In step 3), the mass-to-volume ratio of intermediate II, the second organic solvent, intermediate I, and pyridine is 100 mg: 5-10 mL: 0.2-0.4 mL: 0.2-0.3 mL. The second organic solvent includes one or more of anhydrous acetonitrile, anhydrous DMF, and anhydrous dioxane.

8. The method for preparing a berberine derivative according to claim 7, characterized in that, The reaction in step 3) is carried out at a temperature of 20-30°C for 1-2 hours.

9. The use of the berberine derivative according to claim 1 in the preparation of antibacterial drugs, characterized in that, The bacteria in the antibacterial drugs include drug-resistant Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli.

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