Phenalenone compound as well as preparation method and application thereof

Abstract

The invention discloses a phenalenone compound as well as a preparation method and application thereof, and belongs to the technical field of active ingredient analysis of marine fungi. A series of phenalenone compounds with novel structures are extracted and separated from a fermentation culture of marine fungi Aspergillus flavipes GE2-6 by utilizing the polarity difference of the phenalenone compounds, and an in-vitro anti-HIV virus test shows that the phenalenone compounds provided by the invention have relatively good in-vitro anti-HIV virus activity, are low in cytotoxicity, and can be used for preparing an anti-HIV virus drug. The compound has a development prospect in the aspect of preparing anti-HIV (human immunodeficiency virus) medicines.

Description

[0001] This application is a divisional application of application number 202410134132.7, filed on January 31, 2024, entitled "A class of polyene compounds and their preparation methods and applications". Technical Field

[0002] This invention relates to the field of marine fungal active ingredient analysis technology, specifically to the analysis of active ingredients from Aspergillus flavus (… Aspergillus flavipes Finamane ketone compounds extracted from GE2-6 and their applications. Background Technology

[0003] Small molecule drugs for HIV (human immunodeficiency virus) play a crucial role in combating HIV infection. These drugs are developed to interfere with the viral life cycle, thereby preventing its replication and spread within the host. Nucleotide reverse transcriptase inhibitors, such as lamivudine and abacavir, effectively inhibit viral replication by inhibiting reverse transcriptase activity and preventing viral RNA synthesis. Non-nucleoside reverse transcriptase inhibitors, such as nipravirine, prevent viral genome synthesis by binding to reverse transcriptase and blocking its function. Protease inhibitors, such as ritonavir and retonavir, limit viral maturation by blocking viral proteases and hindering the generation of new viral particles. Integrase inhibitors, such as lavnatide, effectively inhibit viral integration by preventing viral DNA from inserting into the host cell's DNA. Although HIV infection remains a global health challenge, the continuous development and improvement of small molecule drugs provide powerful tools for managing and controlling the disease (Yang Weilin, Song Xian. Advances in clinical pharmacokinetic studies of HIV integrase inhibitors [J]. Modern Clinical Medicine, 2023, 49(05):382-386.).

[0004] Compared to terrestrial microorganisms, marine microorganisms are able to tolerate the extreme conditions unique to the ocean, such as high salinity, high pressure, low oxygen, and low light. This unique living environment leads to diversity in species, genetic composition, and ecological functions among marine microorganisms. The special characteristics of the marine environment, coupled with advancements in marine microbial resource acquisition technologies, have brought unprecedented opportunities for the research of natural medicinal compounds derived from marine microorganisms.

[0005] Marine fungi are a rich source of bioactive secondary metabolites. 70-80% of these metabolites possess biological activity, including small-molecule lactones; fungal toxins; novel substances with inhibitory activity against the central nervous system; 1-dodecyl alcohol, unsaturated hydrocarbons, acids, and esters; and lipopeptide antibiotics that can inhibit the synthesis of new target sites by fungal viruses acting on the fungal cell wall in plants and humans. Discovering natural products with specific structural types using marine fungi as raw materials is of great significance for the development of marine drugs.

[0006] Aspergillus fungi are found in both terrestrial and marine environments. Due to the diverse structure and bioactivity of some species within this genus, they have attracted considerable attention in the pharmaceutical field. Considering the biodiversity and metabolite diversity of this genus, Vadrapudi et al. established the Aspergillus Secondary Metabolite Database (A2MDB) in 2017. This database includes 807 non-repeating secondary metabolites from 675 Aspergillus species, as well as 100 metabolites targeting cellular targets. As of 2021, there are currently over 300 subspecies of *Aspergillus flavus*. Due to the structural and bioactivity diversity of its metabolites, this species is one of the most studied members of the *Aspergillus* genus. Secondary metabolites of *Aspergillus flavus* include cytochalasin, alkaloids, peptides, and polyketides (Chen Wenju. *Aspergillus flavus*, derived from the marine fungus *Aspergillus flavus*). Aspergillus flavipes Research on antibacterial components [D]. Harbin University of Commerce, 2020.).

[0007] However, no literature has been found to date reporting the isolation of asperphenalenone-like compounds against HIV from Aspergillus flavus. Summary of the Invention

[0008] The purpose of this invention is to obtain Aspergillus flavus ( Aspergillus flavipes Natural active substances with medicinal value are extracted from them.

[0009] To achieve the above objectives, the present invention adopts the following technical solution: This invention is derived from Aspergillus flavus ( Aspergillus flavipes Two new finadenolide compounds were isolated from the fermentation products of GE2-6. After structural identification, the specific structural formulas of the two new compounds are shown in formulas (I)-(II), and they are named asperphenalenone O and asperphenalenone P, respectively.

[0010] (I); (II).

[0011] The Aspergillus flavus fungus ( Aspergillus flavipes GE2-6 is a large marine plant isolated from hydrothermal vent sediments on the seabed of Guishan Island, Taiwan Province, China. Its accession number is CCTCC NO: M 20211212.

[0012] This invention also provides a method for obtaining Aspergillus flavus fungus ( Aspergillus flavipes The method for separating and extracting the above-mentioned new compounds from the fermentation products of GE2-6 is described, but the preparation method of the above-mentioned compounds in this invention is not limited to this.

[0013] Specifically, the method for extracting the finadenolide compounds from fermentation products includes the following steps: (1) The Aspergillus flavus fungus with accession number CCTCC NO:M 20211212 ( Aspergillus flavipes After activation, GE2-6 was inoculated into liquid culture medium for fermentation. (2) After the fermentation culture is completed, the fermentation broth is separated. The fermentation broth is stirred with diatomaceous earth and then extracted by reflux with n-butanol to obtain the extract. (3) After concentrating the extract, normal phase silica gel column chromatography was performed. Gradient elution was performed with dichloromethane / methanol mixtures with volume ratios of 100:1, 50:1, 45:1, 40:1, 35:1, 30:1, 25:1, 20:1, 15:1, and 10:1. The fractions eluted by dichloromethane / methanol mixtures with volume ratios of 40:1 to 20:1 were collected and then separated by reverse phase silica gel column chromatography and high performance liquid chromatography to obtain the finadenolide compounds.

[0014] In step (1), the fungus Aspergillus flavus ( Aspergillus flavipes GE2-6 was used for fermentation culture.

[0015] Aspergillus flavus ( Aspergillus flavipes GE2-6 is a fungus, and can be fermented using conventional PDA liquid medium or malt extract medium. To increase yield and provide sufficient nutrients for microbial growth and metabolism, preferably, the liquid medium, per 1L volume, comprises the following ingredients: 1-5g starch, 10-20g wheat bran, 3-15g yeast extract, 1-8g KH2PO4, 0.1-0.8g MgSO4•7H2O, with the remainder being water. Alternatively, the liquid culture medium, per 1L volume, comprises the following ingredients: 200-600g potato, 2-10g peptone, 1-5g yeast extract, 5-20g glucose, with the remainder being water; the initial pH of the culture medium is 6.0-7.0. Alternatively, the liquid culture medium, per 1L volume, comprises the following ingredients: 10-40 g sucrose, 5-20 g corn flour, 1-4 g NaNO3, 1-4 g yeast extract, 0.2-0.8 g KH2PO4, 0.2-1 g MgSO4•7H2O, 0.2-1 g KCl, 0.001-0.005 g FeSO4, with the remainder being water; Alternatively, the liquid culture medium, per 1L volume, comprises the following ingredients: 20-30 g malt extract, 15-20 g glucose, 1-2 g casein peptone, with the remainder being water.

[0016] The fermentation culture conditions are static culture at 20-30℃ for 10-40 days. The static culture method refers to culture without shaking flasks.

[0017] Preferably, the fermentation culture temperature is 22-26°C. More preferably, the culture is carried out at 25°C for 20 days, under which the yield of the finadenolide compounds is the highest.

[0018] In step (2), the fermentation supernatant is obtained by separation, and the finadene compounds are extracted and separated from the fermentation supernatant.

[0019] In step (3), the separation and purification method is as follows: the extract is separated by normal-phase silica gel column chromatography, and the obtained fraction is further separated by reversed-phase silica gel column chromatography and high-performance liquid chromatography. Through multi-step separation and purification, finamazenone compounds with high purity can be obtained.

[0020] Preferably, the fractions are subjected to reversed-phase silica gel column chromatography, with gradient elution using methanol / water mixtures at volume ratios of 1:9, 2:8, 3:7, 4:6, 5:5, 6:4, 7:3, 8:2, and 9:1, eluting three times for each gradient, resulting in a total of 27 fractions, numbered sequentially from 1 to 27. The fractions numbered 20 to 24 are combined and eluted with a methanol / water mixture at a volume ratio of 80:20. The peaks with retention times of 35.6 minutes and 36.9 minutes, respectively, are recrystallized from cyclohexane to obtain compounds asperphenalenone O (structure of formula (I)) and asperphenalenone P (structure of formula (II)).

[0021] This invention demonstrates that the above method can be used to obtain *Aspergillus flavus* fungus (…). Aspergillus flavipes Finamane ketone compounds isolated from GE2-6 fermentation culture exhibit good anti-HIV activity. Therefore, this invention provides the application of these finamane ketone compounds in the preparation of anti-HIV drugs. This invention uses the compound's anti-Namalwa lymphoma cell activity as its cytotoxicity data to verify that it exhibits low toxicity to human cells while exerting anti-HIV drug effects, demonstrating its biocompatibility.

[0022] Specifically, the present invention provides the use of asperphenalenone O with the structural formula as shown in formula (I) or asperphenalenone P with the structural formula as shown in formula (II) in the preparation of drugs against HIV.

[0023] The beneficial effects of this invention are as follows: (1) This invention utilizes the polarity difference of finadene compounds to extract and separate two compounds with novel structures from the fermentation culture of marine fungi. The method is simple to operate, has a high extraction yield and high product purity, and is suitable for large-scale production.

[0024] (2) In vitro anti-HIV assays showed that the asperphenalenone O and asperphenalenone P compounds provided by this invention have good in vitro anti-HIV activity, IC50. 50 The values ​​were 6.1 µM and 4.6 µM, respectively. Further cytotoxicity tests showed that the asperphenalenone O and asperphenalenone P compounds provided by this invention have low cytotoxicity and are promising for the development of anti-HIV drugs. Attached Figure Description

[0025] Figure 1 This is the structural formula of the finadeninone compounds of the present invention.

[0026] Figure 2 For the compound asperphenalenone O 1 H NMR data (in CD3OD, 600 MHz).

[0027] Figure 3 For the compound asperphenalenone O 13 C NMR data (in CD3OD, 150 MHz).

[0028] Figure 4 For the compound asperphenalenone O 1 H- 1 H-COSY data (in CD3OD).

[0029] Figure 5 HSQC data (in CD3OD) for the compound asperphenalenone O.

[0030] Figure 6 HMBC data for the compound asperphenalenone O (in CD3OD).

[0031] Figure 7 NOESY data (in CD3OD) for the compound asperphenalenone O.

[0032] Figure 8Electronic circular dichroism chromatogram data for the compound asperphenalenone O (in CH3OH).

[0033] Figure 9 For the compound asperphenalenone P 1 H NMR data (in CD3OD, 600 MHz).

[0034] Figure 10 For the compound asperphenalenone P 13 C NMR data (in CD3OD, 150 MHz).

[0035] Figure 11 For the compound asperphenalenone P 1 H- 1 H-COSY data (in CD3OD).

[0036] Figure 12 HSQC data (in CD3OD) for compound asperphenalenone P.

[0037] Figure 13 HMBC data (in CD3OD) for compound asperphenalenone P.

[0038] Figure 14 NOESY data (in CD3OD) for the compound asperphenalenone P.

[0039] Figure 15 Electronic circular dichroism chromatogram data (in CH3OH) for the compound asperphenalenone P. Detailed Implementation

[0040] The present invention will be further described below with reference to specific embodiments. These embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Any modifications or substitutions made to the methods, steps, or conditions of the present invention without departing from the spirit and essence of the invention are within the scope of the invention.

[0041] Unless otherwise specified, the experimental methods used in the following examples are conventional methods; the materials and reagents used are commercially available unless otherwise specified.

[0042] Example 1: Fungal Isolation Large marine plants were collected from seabed sediments at the hydrothermal vent of Guishan Island, Taiwan Province, China. After the samples were brought back to the laboratory, they were first rinsed three times with sterile seawater to remove non-attached microorganisms. The plants were then placed in centrifuge tubes, and a small amount of seawater was added. The mixture was vortexed for 10 min, and the suspension after removing the algae was centrifuged for 20 min (5000 r / min). The supernatant was discarded. The precipitate was resuspended in a small amount of sterile seawater, and 0.1 mL was spread on Martin's medium (containing 8 U / L gentamicin) plates. After incubation at 20°C for 10 days, single colonies were picked, purified by streak plating, and then transferred to slant plates for storage at 4°C for later use.

[0043] Example 2: Identification of Aspergillus fungi The isolated fungus was cultured on a PDA, and the 18S rDNA gene sequence of the strain was determined. The 18S rDNA sequence of the strain is shown in SEQ ID No. 1.

[0044] Based on the morphological characteristics and 18S rDNA sequence analysis, the strain was identified as belonging to the genus *Aspergillus*. Aspergillus (sp.) fungus. It was named *Aspergillus flavus*. Aspergillus flavipes GE2-6 was deposited at the China Center for Type Culture Collection (CCTCC) on September 24, 2021, at Wuhan University, Wuhan, China, with accession number CCTCC NO:M 20211212, and was identified as viable on October 9, 2021.

[0045] Example 3: Fermentation culture of Aspergillus fungi The Aspergillus flavus isolated and identified in Example 2 ( Aspergillus flavipes After activation, GE2-6 was made into a spore suspension, which was then inoculated into the culture medium and statically fermented at 25°C for 20 days.

[0046] The culture medium formula is as follows: starch 3g, wheat bran 14g, yeast extract 6g, KH2PO4 5g, MgSO4·7H2O 0.4g, and water 1000mL.

[0047] Example 4: Fermentation culture of Aspergillus fungi The Aspergillus flavus isolated and identified in Example 2 ( Aspergillus flavipes After activation, GE2-6 was made into a spore suspension, which was then inoculated into the culture medium and statically fermented at 24°C for 20 days.

[0048] The culture medium formula is as follows: 400g potato, 6g peptone, 2g yeast extract, 10g glucose, and 1000mL water; the initial pH of the culture medium is 6.5.

[0049] Example 5: Fermentation culture of Aspergillus fungi The Aspergillus flavus isolated and identified in Example 2 ( Aspergillus flavipes After activation, GE2-6 was made into a spore suspension, which was then inoculated into the culture medium and statically fermented at 25°C for 20 days.

[0050] The culture medium formula is as follows: 25g sucrose, 10g corn flour, 2g NaNO3, 2g yeast extract, 0.5g KH2PO4, 0.5g MgSO4·7H2O, 0.5g KCl, 0.001g FeSO4, and 1000mL water.

[0051] Example 6: Fermentation culture of Aspergillus fungi The Aspergillus flavus isolated and identified in Example 2 ( Aspergillus flavipes After activation, GE2-6 was made into a spore suspension, which was then inoculated into the culture medium and statically fermented at 25°C for 20 days.

[0052] The culture medium formula is as follows: 25g malt extract powder, 15g glucose, 1.5g casein peptone, and 1000mL water.

[0053] Example 7: Preparation of finadeninone compounds Aspergillus flavus ( Aspergillus flavipes After fermentation of GE2-6, 5L of fermentation broth was taken, centrifuged, and the supernatant was collected to obtain the fermentation broth. The fermentation broth was concentrated, mixed with 10g of diatomaceous earth, refluxed with 1L of n-butanol, and subjected to normal-phase silica gel column chromatography (200-300 mesh, 1kg; silica gel column size L 50 mm, Ø 12 mm). Gradient elution was performed with dichloromethane / methanol mixtures and methanol at volume ratios of 100:1, 50:1, 45:1, 40:1, 35:1, 30:1, 25:1, 20:1, 15:1, and 10:1, respectively. The fractions eluted from the dichloromethane / methanol mixtures at volume ratios of 40:1 to 20:1 were collected.

[0054] The fraction was subjected to reversed-phase silica gel column chromatography with methanol / water (1:9-9:1) as the eluent. Gradient elution was performed sequentially with methanol / water mixtures of volume ratios of 1:9, 2:8, 3:7, 4:6, 5:5, 6:4, 7:3, 8:2, and 9:1, with 1.5 L eluted per gradient. Fractions were collected in 0.5 L increments using an automated collector, resulting in 27 fractions, which were numbered sequentially from 1 to 27. Based on the similarity of the distillate components, 15 sub-components were merged, resulting in sub-components named 5-1 to 5-15. Specifically, 5-1: 1-3; 5-2: 4; 5-3: 5; 5-4: 6-9; 5-5: 10-11; 5-6: 12; 5-7: 13-14; 5-8: 15; 5-9: 16; 5-10: 17; 5-11: 18; 5-12: 19; 5-13: 20-24; 5-14: 25-26; 5-15: 27.

[0055] Subsequently, high performance liquid chromatography was used for separation. The peaks of subfraction 5-13 with retention times of 35.6 min and 36.9 min, respectively, under the eluent of a methanol / water mixture with a volume ratio of 80%, were recrystallized from cyclohexane to form compounds I and II.

[0056] Example 8: Structural Identification of Finamane Ketones The purity of the synthesized compounds was determined by HPLC. Samples with a purity greater than 98% were subjected to structural identification using mass spectrometry and nuclear magnetic resonance (NMR). NMR was performed using a Bruker AVANCE DRX-600 NMR spectrometer with TMS as an internal standard. High-resolution mass spectrometry (FTIRMS) was performed using a Bruker Apex Spectrometer. Electrospray ionization mass spectrometry (ESI-MS) was performed using a Bruker Esquire. 3000 plus Spectrometer measurement.

[0057] According to the one-dimensional NMR of compound I (see Table 1), Figure 2-3 According to the results of mass spectrometry analysis, the molecular formula of compound I is C1. 35 H 48 O9. Results of analysis by two-dimensional NMR and electron circular dichroism spectroscopy (see...) Figure 4-8 The structure of the compound was confirmed. Through database and literature review, compound I was identified as a new compound, asperphenalenone O. The structure is as follows: Figure 1 As shown in (I).

[0058] Table 1. NMR data of compound I According to the one-dimensional NMR of compound II (see Table 2), Figure 9-10 According to the results of mass spectrometry analysis, the molecular formula of compound II is C2. 35 H 48 O9. Results of analysis by two-dimensional NMR and electron circular dichroism spectroscopy (see...) Figure 11-15 The structure of the compound was confirmed, and through database and literature review, compound II was identified as a new compound, asperphenalenone P. The structure is as follows: Figure 1 As shown in (II).

[0059] Table 2. NMR data of compound II Example 9: Analysis of the anti-HIV activity of compounds asperphenalenone O and asperphenalenone P (1) Packaging of HIV pseudovirus 293T cells were seeded in 6-well plates and transfected when the cell confluence reached 50%–60%. 100 μL of Opti-MEM was added to a 1.5 mL EP tube, along with the backbone packaging and reporter plasmid pNL-4.3-Luc, and the VSV-substituted HIV envelope plasmid pCDNA3.1-VSV-G. Transfection reagent was then added, and the mixture was incubated at room temperature for 15 min. The mixture was then added to the 6-well plates. After 6 h, the medium was aspirated and replaced with fresh DMEM complete medium. After 48 h, the supernatant was collected into a 1.5 mL EP tube, centrifuged at 5000 × g for 10 min at 4 °C, and the supernatant was transferred to a new EP tube and stored at -80 °C for later use.

[0060] (2) HIV pseudovirus infection inhibition experiment 293FT cells were seeded in 96-well plates. When the cell density approached 90%, HIV pseudoviruses were prepared for infection experiments. Asperphenalenone O and asperphenalenone P were added to the cells at different concentrations to influence the entire viral infection process. A blank control group (M) without viral infection and a viral infection control group (C) without any samples were set up. After 48 hours of culture, the chemiluminescence value was detected using a firefly luciferase reporter gene assay kit. The inhibition rate and IC50 were calculated. 50 value.

[0061] (3) Experimental Results Table 3. HIV virus inhibition rate and IC50 of the compounds 50 value Initial screening data showed that asperphenalenone O and asperphenalenone P exhibited inhibition rates of over 75% against HIV pseudovirus infection at a concentration of 30 µM, with asperphenalenone P showing an inhibition rate of 93.5% ± 0.9%. Further rescreening of these compounds revealed that their half-maximal inhibitory concentrations (IC50) were significantly higher than the reported values. 50 The values ​​were 6.1 ± 0.9 and 4.6 ± 1.1 µM, respectively, indicating that compounds asperphenalenone O and asperphenalenone P have good in vitro anti-HIV pseudovirus infection activity.

[0062] Example 10: Cytotoxicity analysis of finadene compounds (1) Reagent preparation 0.4% SRB solution: Weigh 0.8g SRB, dissolve in 200mL 1% acetic acid, and store at room temperature.

[0063] 50% TCA solution: Weigh 50g of TCA, add water to make up to 100mL, and store at 4℃.

[0064] 10mM Tris-base solution: Weigh 0.6057g Tris-base, add water to a final volume of 500mL, pH 10.5, and store at 4℃.

[0065] (2) Experimental apparatus CO2 incubator (Thermo), mini shaker (Kylin-Bell Lab instruments), microplate reader (MD, M5 model).

[0066] (3) Method Namalwa cells in logarithmic growth phase were selected, and after trypsin digestion, the cell concentration was adjusted to 2 × 10⁶ cells / mL using RPMI 1640 medium containing 10% fetal bovine serum. 4Cells were seeded at a rate of 190 μL / well in a 96-well plate and cultured at 37°C with 5% CO2 for 24 h. For drug treatment wells, 10 μL of sample solution was added (final concentration: 5 μg / mL for the compound; 50 μg / mL for the mixture). Positive control wells were treated with 5-FU at a final concentration of 5 μg / mL. Control wells were treated with culture medium containing an equal volume of solvent and cultured at 37°C with 5% CO2 for 3 days. The culture medium was discarded, and 100 μL of 50% TCA (pre-chilled at 4°C) was gently added to fix the cells. The cells were incubated for 5 min, then transferred to 4°C for 1 h. The fixative was discarded, and the cells were washed 5 times with distilled water to remove TCA and air-dried for 1 h. 80 μL of 0.4% SRB solution was added to each well, and staining was performed at room temperature for 30 min. The staining solution was discarded, and the cells were washed 5 times with 1% acetic acid to thoroughly remove unbound SRB and air-dried. Dissolve in 150 μL of 10 mM Tris-base (pH 10.5) and shake on a micro-shaker for 5 min. Measure the OD510 nm value using an M5 microplate reader.

[0067] (4) Calculation of results Tumor cell growth inhibition rate (%) = (OD) 对照 -OD 药物 ) / (OD 对照 -OD 空白 )×100% (5) Experimental Results Table 4. Namalwa cell survival rate of the compound The cytotoxicity of asperphenalenone O and asperphenalenone P to Namalwa cell line at a concentration of 20 µM was further determined. The data showed that the survival rate of the positive control drug 5-Fu to Namalwa cell line was 11.2%, and the survival rate of the positive control drug As2O3 to Namalwa cell line was 5.6%. All compounds showed low cytotoxicity to Namalwa cell line.

Claims

1. A class of finadeninone compounds, characterized in that, The structural formulas of the finadenophenone compounds are shown in formula (I) or formula (II). (AND); (II)。 2. The method for preparing finadeninone compounds as described in claim 1, characterized in that, Includes the following steps: (1) The Aspergillus flavus fungus with accession number CCTCC NO:M 20211212 ( Aspergillus flavipes After activation, GE2-6 was inoculated into liquid culture medium for fermentation. (2) After the fermentation culture is completed, the fermentation broth is separated. The fermentation broth is stirred with diatomaceous earth and then extracted by reflux with n-butanol to obtain the extract. (3) After concentrating the extract, normal phase silica gel column chromatography was performed. Gradient elution was performed with dichloromethane / methanol mixtures with volume ratios of 100:1, 50:1, 45:1, 40:1, 35:1, 30:1, 25:1, 20:1, 15:1, and 10:

1. The fractions eluted by dichloromethane / methanol mixtures with volume ratios of 40:1 to 20:1 were collected and then separated by reverse phase silica gel column chromatography and high performance liquid chromatography to obtain the finadenolide compounds.

3. The preparation method according to claim 2, characterized in that, In step (1), the liquid culture medium comprises the following raw materials in 1L volume: starch 1-5g, wheat bran 10-20g, yeast extract 3-15g, KH2PO4 1-8g, MgSO4•7H2O 0.1-0.8g, and the remainder is water.

4. The preparation method according to claim 2, characterized in that, In step (1), the liquid culture medium, in 1L volume, includes the following raw materials: 200-600g of potato, 2-10g of peptone, 1-5g of yeast extract, 5-20g of glucose, and the remainder is water; the initial pH of the culture medium is 6.0-7.

0.

5. The preparation method according to claim 2, characterized in that, In step (1), the liquid culture medium, in 1L volume, comprises the following raw materials: 10-40 g sucrose, 5-20 g corn flour, 1-4 g NaNO3, 1-4 g yeast extract, 0.2-0.8 g KH2PO4, 0.2-1 g MgSO4•7H2O, 0.2-1 g KCl, 0.001-0.005 g FeSO4, with the remainder being water.

6. The preparation method according to claim 2, characterized in that, In step (1), the liquid culture medium comprises the following raw materials in 1L volume: 20-30 g malt extract powder, 15-20 g glucose, 1-2 g casein peptone, and the remainder is water.

7. The preparation method according to claim 2, characterized in that, In step (1), the fermentation culture conditions are static culture at 20-30℃ for 10-40 days.

8. The preparation method according to claim 2, characterized in that, In step (3), the fractions are subjected to reversed-phase silica gel column chromatography, with gradient elution using methanol / water mixtures of volume ratios of 1:9, 2:8, 3:7, 4:6, 5:5, 6:4, 7:3, 8:2, and 9:1, eluting three times for each gradient. The collected fractions are numbered sequentially from 1 to 27. Then, high-performance liquid chromatography is used for separation. The fractions numbered 20 to 24 are combined and eluented with a methanol / water mixture of volume ratio of 80:

20. The peaks with retention times of 35.6 minutes and 36.9 minutes, respectively, are recrystallized from cyclohexane to obtain compounds asperphenalenone O with structural formula (I) and asperphenalenone P with structural formula (II).

9. Application of finadene compounds with structural formulas as shown in formula (I) or (II) in the preparation of drugs against HIV.

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