Application of dihydroisocoumarin derivative extracted from Penicillium chrysogenum fermentation product in preparation of drug for preventing or treating drug-resistant bacterial infection

Abstract

The invention discloses an application of a dihydroisocoumarin derivative extracted from a penicillium chrysogenum fermentation product in preparation of a medicine for preventing or treating drug-resistant bacterial infection, and belongs to the technical field of medicines. According to the application, the dihydroisocoumarin derivative obtained by extracting secondary metabolites obtained by fermentation of penicillium chrysogenum is combined with antibiotics for use, so that the drug resistance of drug-resistant bacteria can be effectively overcome, and the drug-resistant bacteria can be effectively inhibited and killed; the compound has a wide application prospect in drugs for preventing or treating drug-resistant bacterium infection or diseases caused by the drug-resistant bacterium infection; particularly, the method for extracting the dihydroisocoumarin derivative through fermentation of the penicillium chrysogenum not only has the advantages that the strains are easy to obtain, but also has the advantages of being high in yield and mature in process, production can be remarkably achieved, the cost can be reduced, and the method is suitable for large-scale application and popularization.

Description

Technical Field

[0001] The invention belongs to the field of medical technology, and particularly relates to an application of a dihydroisocoumarin derivative extracted from a fermentation product of Penicillium chrysogenum in preparing a medicine for preventing or treating drug-resistant bacterial infection. Background Art

[0002] With the widespread use of antibiotics, bacterial resistance to antibiotics is increasing. However, due to the high cost of developing new antibiotics and the rapid rate at which bacteria develop resistance, current drug development cannot fully meet the clinical needs of antibiotics.

[0003] Penicillium Chrysogenum (P. chrysogenum) is an industrial strain that can produce penicillin, one of the most well-known antibiotics, which can be produced by fermentation of a variety of Penicillium. During the fermentation process, due to differences in raw materials, temperature and other factors, a variety of secondary metabolites can be provided, many of which have biological and pharmaceutical uses.

[0004] Isocoumarins are a class of secondary metabolites with diverse structures and multiple important pharmacological activities. They belong to the aminocoumarin antibiotics and are isomers of coumarin. Most of the common ones are dihydroisocoumarins, which are widely present in plants and microorganisms in nature. They are natural products with important antibacterial activity and can be isolated from Streptomyces snow white. It has a unique chemical structure and antibacterial mechanism, including the inhibitory function of bacterial DNA gyrase and eukaryotic DNA topoisomerase II, and has significant inhibitory and killing effects on a variety of bacteria and fungi. Its mechanism of action is to specifically target and inhibit the activity of the GyrB subunit, thereby interfering with DNA synthesis and ultimately affecting bacterial growth. Therefore, it has broad application prospects in the fields of medicine, agriculture, and environmental protection. Most natural dihydroisocoumarins are isolated from various terrestrial and marine natural sources, such as plants, marine organisms, mosses, insects, microorganisms, bacteria, and fungi (endophytes, soil, and marine fungi). For example, Slava Epstein et al. studied the use of marine fungi to prepare dihydroisocoumarin derivatives with plant growth inhibition.

[0005] Wei Meiyan et al. studied the secondary metabolites of the endophytic fungus Microsphaeropsis sp. in the mangroves of South China Sea, and separated and obtained 8 dihydroisocoumarin monomer compounds from the ethyl acetate phase of 150L of the culture solution of the fungus, and the total yield of the 8 compounds was about 1.2mg / L; however, the fungus is not easy to obtain, the extraction efficiency of the metabolites is low, and it is not suitable for large-scale promotion and application. Chinese invention patent CN 110483467A provides a dihydroisocoumarin derivative and its preparation method and use. The invention uses Aspergillus pyrophorus with a preservation number of CCTCC No: M2014086 to obtain a fermentation product containing dihydroisocoumarin derivatives, then adds methanol to soak and ultrasonically filter, evaporate to dryness, and then extracts with ethyl acetate and evaporates to dryness to obtain a crude extract. The extract is subjected to reduced pressure silica gel column chromatography, reversed phase medium pressure column chromatography, and finally separated and purified by reversed phase high performance liquid chromatography, and applied to plant growth inhibition. Chinese invention patent CN103880824A discloses three new isocoumarin antibiotics, tanamycin B, C, and D, obtained by fermentation, extraction, and separation of Bacillus subtilis strains (Bacillus subtilis subsp. inaquosorum). The results show that tanamycin B, C, and D have inhibitory activity against Staphylococcus, including methicillin-sensitive Staphylococcus aureus (MSSA), methicillin-resistant Staphylococcus aureus (MRSA), methicillin-sensitive Staphylococcus epidermidis (MSSE), and methicillin-resistant Staphylococcus epidermidis (MRSE), and it is also expected to become a drug against Gram-positive bacteria such as Staphylococcus and its drug-resistant bacteria (MRSA) infection. The prior art uses different strains to extract isocoumarins of various structures, and the extraction yield is relatively low, which cannot meet the requirements of large-scale production.

[0006] Based on the current status of the prior art, the present invention provides a method for extracting antibacterial dihydroisocoumarins by fermenting Penicillium chrysogenum. The method has the characteristics of easy availability of strains and high yield. At the same time, the in vitro antibacterial effect of metabolites and their synergistic effect on common antibiotics are also studied, thereby playing a certain role in the prevention and treatment of common drug-resistant bacteria in clinical practice. Summary of the invention

[0007] In view of this, in order to overcome the shortcomings of the above-mentioned prior art, the present invention provides an application of dihydroisocoumarin derivatives extracted from fermentation products of Penicillium chrysogenum in the preparation of drugs for preventing or treating drug-resistant bacterial infections. By extracting and separating the metabolites of Penicillium chrysogenum (P. chrysogenum), dihydroisocoumarin derivatives with antibacterial activity and combinations thereof are provided, and by analyzing the activity inhibition of drug-resistant bacteria, the derivatives are applied to drugs for drug-resistant bacterial infections. In particular, the extraction method has the advantages of easy acquisition of strains and high yield.

[0008] In order to achieve the above object, the present invention provides an application of dihydroisocoumarin derivatives extracted from fermentation products of Penicillium chrysogenum in the preparation of drugs for preventing or treating drug-resistant bacterial infections.

[0009] Preferably, the dihydroisocoumarin derivative is obtained by extracting secondary metabolites after fermentation of Penicillium chrysogenum (P. chrysogenum) and separating them.

[0010] More preferably, the method for extracting the dihydroisocoumarin derivative comprises:

[0011] Step 1. Fermentation

[0012] Corn distillate was added to the fermentation medium of Penicillium chrysogenum as a fermentation precursor, the pH value was adjusted to 6.8, Penicillium chrysogenum was inoculated, and the culture was carried out in a constant temperature shaking incubator at 28°C for 10 days;

[0013] Step 2. Rough extraction

[0014] Adding ethyl acetate to the fermentation medium of Penicillium chrysogenum after fermentation in step 1, and obtaining a crude extract after filtering and evaporation;

[0015] Step 3. Purification

[0016] The crude extract is added to n-hexane-95% methanol, and the methanol phase is eluted by a n-hexane-ethyl acetate elution system. The obtained product is then eluted by n-hexane-ethyl acetate to obtain monomers of different dihydroisocoumarin derivatives.

[0017] Preferably, the structural formula of the dihydroisocoumarin derivative is as shown in formula (I) or formula (II):

[0018]

[0019] Preferably, the drug-resistant bacteria include at least one or more of Enterococcus, Staphylococcus aureus, Klebsiella, Acinetobacter baumannii, Pseudomonas aeruginosa and Escherichia coli.

[0020] Preferably, the drug also includes an antibiotic; the dihydroisocoumarin derivative and the antibiotic can inhibit the biological activity of drug-resistant bacteria through synergistic action.

[0021] Preferably, when the structural formula of the dihydroisocoumarin derivative is a compound represented by formula (I) or formula (II), the dihydroisocoumarin derivative is combined with the antibiotic to form a dual pharmaceutical composition, and the mass ratio of the two is 1:1 to 3.

[0022] Preferably, when the structural formulas of the dihydroisocoumarin derivatives are the compound represented by formula (I) and the compound represented by formula (II), respectively, the dihydroisocoumarin derivatives are combined with the antibiotics to form a triple drug composition.

[0023] Preferably, the concentration ratio of the compound represented by formula (I) to the compound represented by formula (II) is 1:2 to 2:1.

[0024] Preferably, the ratio of the sum of the concentrations of the compound represented by formula (I) and the compound represented by formula (II) to the concentration of the antibiotic is 1:1-3.

[0025] Preferably, the antibiotics include any one of imipenem, imipenem, erythromycin, levofloxacin, vancomycin, colistin, amoxicillin, and ceftriaxone.

[0026] As one of the purposes of the invention, the present invention also provides a drug for preventing or treating drug-resistant bacterial infection, which at least comprises an antibiotic and a dihydroisocoumarin derivative as shown in formula (I).

[0027] Beneficial technical effects obtained by the present invention:

[0028] 1. The present invention uses the fermentation medium of Penicillium chrysogenum to extract antibacterial dihydroisocoumarins after 10 days of culture. Compared with the prior art using other relatively rare fungal fermentation or chemical synthesis methods to prepare antibacterial dihydroisocoumarins, it has the following advantages: on the one hand, the fermentation technology of Penicillium chrysogenum is more mature, and the strain is cheap, which can significantly reduce the cost of extracting metabolites; on the other hand, the extraction method of metabolites is simple, the process is mature, the types of metabolites are simple, the yield is high, and it is more suitable for large-scale promotion and application.

[0029] 2. By adopting the technical scheme of the present invention, the in vitro antibacterial effect of the metabolites extracted and fermented by Penicillium chrysogenum is determined, and the two antibacterial dihydroisocoumarins of the metabolites have certain antibacterial properties; especially when the antibacterial dihydroisocoumarins are used in combination with common clinical antibiotics, the drug resistance of drug-resistant bacteria can be effectively overcome, and the drug-resistant bacteria can be effectively inhibited and killed, which has broad application prospects in the prevention or treatment of drug-resistant bacterial infections or diseases caused by them.

[0030] 3 The technical scheme of the present invention is adopted to extract two monomer compounds from the metabolites of Penicillium chrysogenum (P. chrysogenum), which show good antibacterial activity against drug-resistant bacteria, and show significant antibacterial effects on enterococci, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Escherichia coli.

[0031] 4. The technical solution of the present invention is used in combination with the two metabolites to achieve a synergistic effect, which can increase the inhibitory effect of antibiotics on drug-resistant bacteria by up to 512 times. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 This is a flow chart for the extraction of antibacterial dihydroisocoumarin derivatives provided in Example 1 of the present invention.

[0033] Figure 2 This is the time-kill curve of the antibacterial dihydroisocoumarin 1 at the MIC concentration provided in Example 1 of the present invention against drug-resistant bacteria.

[0034] Figure 3 This is the time-killing curve of the antibacterial dihydroisocoumarin 2 at the MIC concentration provided in Example 1 of the present invention against drug-resistant bacteria. DETAILED DESCRIPTION

[0035] In order to make the purpose, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions in the embodiments of the present application will be clearly and completely described below in combination with the drawings in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, not all of the embodiments.

[0036] The invention provides an application of a dihydroisocoumarin derivative extracted from a fermentation product of Penicillium chrysogenum in preparing a medicine for preventing or treating drug-resistant bacterial infection.

[0037] The invention adds corn distillate as a fermentation precursor to the fermentation medium of Penicillium chrysogenum and controls the pH to about 6.8, so that secondary metabolites can be extracted after the fermentation is completed. After separation, various antibacterial dihydroisocoumarin derivatives can be obtained, and in vitro antibacterial experiments are carried out on the various derivatives to determine their antibacterial properties.

[0038] The structural formula of the antibacterial dihydroisocoumarin derivative extracted by the technical solution of the present invention is shown in formula (I) or formula (II):

[0039]

[0040] The compounds shown in formula (I) and formula (II) have a certain degree of antibacterial activity and can produce obvious antibacterial effects on Gram-positive strains within 8 hours, while the effect on Gram-negative strains is slower, but can also produce a certain antibacterial effect within 24 hours.

[0041] As a preferred embodiment, when any one of the two antibacterial dihydroisocoumarin derivatives is used in combination with an antibiotic, the effect of the antibiotic can be enhanced by up to 8-64 times.

[0042] As a preferred embodiment, the triple use of two antibacterial dihydroisocoumarins and antibiotics can expand the antibacterial spectrum of some antibiotics and further increase the synergistic effect, which can enhance the effect of antibiotics by up to 512 times.

[0043] Therefore, the secondary metabolites produced by the fermentation of Penicillium chrysogenum in the present invention can be used in combination with various conventional antibiotics to exert a synergistic effect for the prevention and treatment of common clinical drug-resistant bacteria, thereby providing a new solution for the preparation of highly effective drugs for drug-resistant bacterial infections.

[0044] In particular, the extraction of secondary metabolites by fermentation of Penicillium chrysogenum adopted in the present invention has the advantages of simple method and relatively mature technology, which is different from the chemical synthesis and fungal fermentation methods of the prior art and has a wider significance for promotion and application.

[0045] The technical solution of the present invention is further described in detail below through specific embodiments.

[0046] Example 1

[0047] This embodiment provides a method for extracting dihydroisocoumarin derivatives, by extracting secondary metabolites from Penicillium chrysogenum fermentation, and obtaining a variety of antibacterial dihydroisocoumarin derivatives after separation. The corresponding in vitro antibacterial experiments are performed to determine their antibacterial properties. Specifically, the steps of extracting antibacterial dihydroisocoumarin include:

[0048] 1. Fermentation of Penicillium chrysogenum

[0049] See also Figure 1 , is a flow chart of the extraction of antimicrobial dihydroisocoumarins. A 1L flask containing 400mL of Penicillium chrysogenum fermentation medium was inoculated with 10 6 CFU Penicillium chrysogenum (P. chrysogenum, American Type Culture Collection ATCC10106), cultured in a constant temperature shaker at 28°C, 200 rpm for 10 days; the pH value of the fermentation medium of Penicillium chrysogenum is ∼6.8, and the composition is shown in Table 1. In Table 1, the corn steam liquor includes 400 g corn and 40 g rice, which are steamed and homogenized and then water is added to make the volume 1L.

[0050] Table 1 List of ingredients of fermentation medium for Penicillium chrysogenum

[0051] Element Content (g / L) glucose 2.0 lactose 25.0 Corn Evaporation 60.0 Calcium carbonate 5.0 Sodium phenylacetate 0.5 Sodium chloride 0.5

[0052] 2. Extraction of Antibacterial Dihydroisocoumarin

[0053] Continue reading Figure 1The extraction of antibacterial dihydroisocoumarin includes: adding 500 mL of ethyl acetate to the culture medium after fermentation in the above step, extracting twice using an extraction flask, filtering and evaporating to obtain about 25 g of a crude extract, distributing it into n-hexane-95% methanol, and the methanol phase can be eluted by a n-hexane-ethyl acetate system (90:10 (S1), 50:50 (S2)) to obtain fractions S1 and S2, and fraction S1 can be eluted by n-hexane-ethyl acetate (9:1) to obtain antibacterial dihydroisocoumarin 1 (about 80.9 mg); fraction S2 can be eluted by n-hexane-ethyl acetate (6.5:3.5) to obtain antibacterial dihydroisocoumarin 2 (about 73.8 mg); both antibacterial dihydroisocoumarin 1 and antibacterial dihydroisocoumarin 2 are white amorphous powders. Obviously, the total extraction amount of antibacterial dihydroisocoumarin 1 and antibacterial dihydroisocoumarin 2 extracted in this example is greater than 150 mg / L, which is significantly higher than the total extraction amount (~1.2 mg / L) of the secondary metabolites of the endophytic fungus Microsphaeropsis sp. of the South China Sea mangroves in the prior art. For details, see Wei Meiyan et al., Journal of Sun Yat-sen University (Natural Science Edition), Vol. 49, No. 2.

[0054] Antimicrobial dihydroisocoumarin 1 1 H-NMR (DMSO, 400MHz): 1.35 (3H, d), 2.83 (1H, m), 3.08 (1H, m), 5.15 (1H, m), 7.53 (1H), 7.70 (1H), 12.74 (1H), 16.47 (1H).

[0055] Antibacterial dihydroisocoumarin 2 1 H-NMR (DMSO, 400MHz): 1.30 (3H, d), 2.29 (3H, s), 5.15 (1H, m), 5.37 (1H, d), 5.77 (1H), 6.84 (1H, d), 7.32 (1H, d), 16.47 (1H).

[0056] The structural formula of antibacterial dihydroisocoumarin 1 is:

[0057] The structural formula of antibacterial dihydroisocoumarin 2 is:

[0058] In vitro antibacterial test of dihydroisocoumarin

[0059] Including the determination of minimum inhibitory concentration (MIC), the determination of time-kill curve (Time-Kill curve), and the determination of minimum graded inhibitory concentration (FIC). Specifically, the test method includes:

[0060] (1) Minimum inhibitory concentration (MIC): The method in the Clinical Laboratory Standards Institute (CLSI)-M100 (broth microdilution method) was modified appropriately to determine the MIC of three antimicrobial dihydroisocoumarins against common clinical ESKAPE-resistant pathogens (Enterococcus, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Escherichia coli), with novobiocin as the control antibiotic, and repeated five times.

[0061] Among them, the drug-resistant pathogens used in this example are: Enterococcus: VRE, American Type Culture Collection ATCC29212; Staphylococcus aureus: MRSA, American Type Culture Collection ATCC BAA-1556; Klebsiella pneumoniae: KP, ATCC700603; Acinetobacter baumannii: DR-AB, bio-53272; Pseudomonas aeruginosa: PGO, American Type Culture Collection ATCC BAA-1556; Escherichia coli: EC, American Type Culture Collection ATCC8739. Novobiocin was purchased from Beijing Solaibao Technology Co., Ltd., item number N8310.

[0062] (2) Time-Kill curve: The cultured bacteria were diluted to the concentration required by the microdilution method. One portion was cultured normally (without the addition of antimicrobial dihydroisocoumarin) and the other portion was cultured with the antimicrobial dihydroisocoumarin added and the final concentration was adjusted to the MIC. The two were cultured simultaneously in a 96-well plate and the bacterial solution in each well was taken out at 8 culture time points (0, 2, 4, 6, 8, 12, 16, and 24 h) for 10 min. 0 -10 7 10 μL of the diluted sample was plated on LB agar plates and the colonies were counted after overnight culture.

[0063] (3) Minimum fractional inhibitory concentration (FIC): The antibacterial effects of antifungal drugs alone and in combination with antibacterial dihydroisocoumarins (final concentration: 1 / 4MIC) were measured on a 96-well plate using the microbroth chessboard method, and the fractional inhibitory concentration index (FIC index) was calculated. The FIC index value ≤ 0.5 indicates synergy, 0.5-2 indicates indifference or addition, and > 2 indicates antagonism. The FIC value calculation method is as follows, where MIC* is the concentration of a single component that has an inhibitory effect when used in combination.

[0064]

[0065] Analysis of in vitro antibacterial test results of antibacterial dihydroisocoumarin:

[0066] Minimum inhibitory concentration (MIC): The results of MIC determination by microbroth dilution method are shown in Table 2. Both extracted antibacterial dihydroisocoumarins have certain in vitro antibacterial effects, which are slightly weaker than that of neomycin.

[0067] Table 2 MIC (μg / mL) results of antibacterial dihydroisocoumarins

[0068]

[0069] Time-Kill curve: The time-kill curve of three antibacterial dihydroisocoumarins with MIC concentration (512μg / mL was used as the test concentration for MIC>512μg / mL) was measured against six resistant bacteria. The results showed that antibacterial dihydroisocoumarins can produce effects faster against Gram-positive strains, but slower against Gram-negative strains. However, they can have sufficient antibacterial effects within 24 hours.

[0070] Minimum graded inhibitory concentration (FIC) determination: When the final concentration of antibacterial dihydroisocoumarin is 1 / 4MIC, it is measured in combination with 2 times the gradient concentration of commonly used antibiotics to detect synergistic effects. The experimental results are shown in Table 3. Most commonly used antibiotics have synergistic effects with two antibacterial dihydroisocoumarins, and the rest are irrelevant or additive, with no antagonistic effects. In the synergistic combination, antibacterial dihydroisocoumarin at a concentration of 1 / 4MIC can increase the effect of antibiotics by 8-64 times, showing a good synergistic effect. When the two antibacterial dihydroisocoumarins are used in combination and not in combination with antibiotics, there is no significant enhancement of the antibacterial effect. The experimental results are shown in Table 4.

[0071] Table 3 FIC results of antibacterial dihydroisocoumarins and commonly used antibiotics

[0072]

[0073]

[0074]

[0075]

[0076] Table 4 Results of the combination of antibacterial dihydroisocoumarins

[0077]

[0078] Triple FIC effect determination:

[0079] The present invention conducts an in vitro antibacterial effect test on a triple antibacterial composition of the extracted antibacterial dihydroisocoumarin 1, the antibacterial dihydroisocoumarin 2 and conventional antibiotics. The method is to prepare 4 different proportions of antibacterial dihydroisocoumarin mixtures in advance and use them in combination with antibiotics. The results are shown in Table 5 below. Compared with the combination of a single antibacterial dihydroisocoumarin and an antibiotic, the triple combination can expand the antibacterial spectrum of some antibiotics and further increase the synergistic effect, and can enhance the effect of antibiotics by up to 512 times.

[0080] In Table 5, the antibacterial dihydroisocoumarin mixed scheme includes 4 combination schemes, including Combination 1: Antibacterial dihydroisocoumarin 1: Antibacterial dihydroisocoumarin 2 = 1 / 4MIC: 1 / 4MIC; Combination 2: Antibacterial dihydroisocoumarin 2 = 1 / 4MIC: 1 / 8MIC; Combination 3: Antibacterial dihydroisocoumarin 2 = 1 / 8MIC: 1 / 4MIC; Combination 4: Antibacterial dihydroisocoumarin 2 = 1 / 8MIC: 1 / 8MIC.

[0081] The two antibacterial dihydroisocoumarins are mixed and then used in combination with antibiotics in a mass ratio of (antibacterial dihydroisocoumarin 1+antibacterial dihydroisocoumarin 2) and used in combination with commonly used antibiotics in a gradient concentration of 2 times.

[0082] Table 5 Triple FIC results of antibacterial dihydroisocoumarins and commonly used antibiotics

[0083]

[0084]

[0085]

[0086]

[0087]

[0088]

[0089]

[0090] It should be noted that the present invention has tested the antibacterial activity of the minimum inhibitory concentration (MIC) on the above-mentioned monomer compounds and the combined use of monomer compounds. The test results show that the monomer compounds have antibacterial activity, and the inhibitory effects on different strains are significantly different, but the antibacterial activity is significantly lower than the antibacterial effect of antibiotics in the prior art, for example, the antibacterial activity of antibacterial dihydroisocoumarin 1 and antibacterial dihydroisocoumarin 2 is significantly lower than that of neomycin. Furthermore, the present invention also studies the monomer compounds for combined use, showing obvious antibacterial activity, indicating that the compounds play a synergistic role when used in combination, and using them as a composition in antibacterial drugs can achieve obvious technical effects.

[0091] The above are only preferred embodiments of the present invention, which do not limit the protection scope of the present invention. For those skilled in the art, the present invention may have various modifications and changes. Within the spirit and principle of the present invention, changes, modifications, replacements, integrations and parameter changes to these embodiments by conventional substitutions or capable of achieving the same functions without departing from the principles and spirit of the present invention all fall within the protection scope of the present invention.

Claims

1. Application of dihydroisocoumarin derivatives extracted from fermentation products of Penicillium chrysogenum in the preparation of drugs for preventing or treating drug-resistant bacterial infections.

2. The use according to claim 1, characterized in that: The dihydroisocoumarin derivatives are obtained by extracting secondary metabolites from Penicillium chrysogenum P. chrysogenum after fermentation and then separating the secondary metabolites.

3. The use according to claim 1, characterized in that: The extraction method of the dihydroisocoumarin derivative comprises: Step 1. Fermentation Corn distillate was added to the fermentation medium of Penicillium chrysogenum as a fermentation precursor, the pH value was adjusted to 6.8, Penicillium chrysogenum was inoculated, and the culture was carried out in a constant temperature shaking incubator at 28°C for 10 days; Step 2. Rough extraction Adding ethyl acetate to the fermentation medium of Penicillium chrysogenum after fermentation in step 1, and obtaining a crude extract after filtering and evaporation; Step 3. Fractionation The crude extract is added to n-hexane-95% methanol, and the methanol phase is eluted with a n-hexane-ethyl acetate elution system. The obtained fraction is then eluted with n-hexane-ethyl acetate to obtain monomers of different dihydroisocoumarin derivatives.

4. The use according to claim 1, characterized in that The structural formula of the dihydroisocoumarin derivative is shown in formula (I) or formula (II):

5. The use according to claim 1, characterized in that: The drug-resistant bacteria include at least one or more of enterococci, Staphylococcus aureus, Klebsiella, Acinetobacter baumannii, Pseudomonas aeruginosa, and Escherichia coli.

6. The use according to any one of claims 1 to 5, characterized in that: The medicine also includes antibiotics; the dihydroisocoumarin derivative and the antibiotics can inhibit the biological activity of drug-resistant bacteria through synergistic action.

7. The use according to claim 6, characterized in that: When the structural formula of the dihydroisocoumarin derivative is a compound represented by formula (I) or formula (II), the dihydroisocoumarin derivative is combined with the antibiotic to form a dual drug composition, and the mass ratio of the two is 1:1 to 3.

8. The use according to claim 6, characterized in that: When the structural formulas of the dihydroisocoumarin derivative are respectively the compound represented by formula (I) and the compound represented by formula (II), the dihydroisocoumarin derivative is combined with the antibiotic to form a triple drug composition; The mass ratio of the compound represented by formula (I) to the compound represented by formula (II) is 1:2 to 2:1; The mass ratio of the sum of the mass of the compound represented by formula (I) and the compound represented by formula (II) to the mass of the antibiotic is 1:1-3.

9. The use according to claim 6, characterized in that: The antibiotics include any one of imipenem, imipenem, erythromycin, levofloxacin, vancomycin, colistin, amoxicillin, and ceftriaxone.

10. A drug for preventing or treating drug-resistant bacterial infection, characterized in that: It at least comprises an antibiotic and a dihydroisocoumarin derivative as shown in formula (I) or formula (II).

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