A drug for treating neuroinflammation and its application

Abstract

This invention relates to a drug for treating neuroinflammation and its application, belonging to the field of pharmaceutical technology. This invention isolates and identifies novel terphenyl compounds from the acetone extract of *Thelephora ganbajun* Zang, and provides a method for the extraction and separation of these new compounds. The acetone extract is obtained by extracting dried fruiting bodies of *Thelephora ganbajun* Zang with acetone, and then separated by normal-phase silica gel column chromatography, reversed-phase silica gel (ODS) column chromatography, and high-performance liquid chromatography (HPLC). The novel compounds of this invention can serve as potential lead compounds for the preparation of drugs for treating neuroinflammation-related diseases.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and in particular to a drug for treating neuroinflammation and its application. Background Technology

[0002] Ganbajun (Thelephora ganbajun Zang), a precious and edible higher fungus, possesses various medicinal properties, including tonifying kidney essence, strengthening muscles and bones, invigorating the spleen and stomach, nourishing the liver and kidneys, promoting diuresis and lowering blood pressure, anti-fatigue, and anti-cancer effects. It is mainly produced in Yunnan Province, my country. Modern research indicates that Ganbajun extract contains multiple components, and pharmacological studies have shown that the chemical components in Ganbajun mainly exhibit antibacterial, antitumor, and antioxidant effects. Therefore, developing the types of compounds in Ganbajun extract and their pharmacological applications is a pressing technical issue that needs to be addressed in this field. Summary of the Invention

[0003] The purpose of this invention is to overcome the shortcomings of existing technologies and provide a drug for treating neuroinflammation and its application. This invention isolates and identifies novel terphenyl compounds from the acetone extract of *Thelephora ganbajun Zang*, and experimentally confirms that these compounds have therapeutic efficacy for neuroinflammation-related diseases.

[0004] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0005] In a first aspect, the present invention provides a terphenyl derivative having any of the following chemical structural formulas:

[0006]

[0007] Where R is hydrogen, hydroxyl, methoxy, benzoyloxy, phenylacetoxy, isopentenyl, alkyl, alkenyl, alkynyl, or substituted or unsubstituted aryl.

[0008] This invention isolates a terphenyl derivative compound from the dried fruiting body extract of *Bacteroides rubrum*. Studies have shown that this compound can inhibit inflammatory factors, improve behavior in a mouse model of depression, and has therapeutic effects on neuritis.

[0009] As a preferred embodiment of the present invention, the molecular structure of the terphenyl derivative having the chemical structural formula shown in formula (II) is as follows:

[0010]

[0011] Wherein, R1 is hydroxyl or methoxy; R2 is hydroxyl or methoxy; and R3 is hydrogen or phenylacetyl.

[0012] In a preferred embodiment of the 001 structural formula of the present invention, R1 is a hydroxyl group; R2 is a hydroxyl group; and R3 is a phenylacetyl group.

[0013] In a preferred embodiment of the 001 structural formula of the present invention, R1 is a hydroxyl group; R2 is a hydroxyl group; and R3 is hydrogen.

[0014] In a preferred embodiment of the 001 structural formula of the present invention, R1 is a methoxy group; R2 is a hydroxyl group; and R3 is hydrogen.

[0015] In a preferred embodiment of the 001 structural formula of the present invention, R1 is a hydroxyl group; R2 is a methoxy group; and R3 is hydrogen.

[0016] As a preferred embodiment of the present invention, the molecular structure of the terphenyl derivative having the chemical structural formula shown in formula (III) is as follows:

[0017]

[0018] Where R is hydrogen or phenylacetyl.

[0019] As a preferred embodiment of the present invention, the molecular structure of the terphenyl derivative having the chemical structural formula shown in formula (IV) is as follows:

[0020]

[0021] As a preferred embodiment of the present invention, the molecular structure of the terphenyl derivative having the chemical structural formula shown in formula (V) is as follows:

[0022]

[0023] Wherein, R1 is hydrogen or phenylacetyl; R2 is hydrogen or phenylacetyl.

[0024] In a preferred embodiment of the 004 structural formula of the present invention, R1 is hydrogen; R2 is phenylacetyl.

[0025] In a preferred embodiment of the 004 structural formula of the present invention, R1 is phenylacetyl; R2 is hydrogen.

[0026] In a preferred embodiment of the 004 structural formula of the present invention, R1 is phenylacetyl; R2 is phenylacetyl.

[0027] Secondly, the present invention provides the use of the terphenyl derivatives described in the first aspect in the preparation of drugs for treating neuroinflammatory diseases.

[0028] Experiments have shown that the compounds of this invention can inhibit inflammatory factors, improve the behavior of a mouse model of depression, and have therapeutic effects on neuritis.

[0029] Thirdly, the present invention provides a method for preparing the terphenyl derivatives described in the first aspect, comprising the following steps:

[0030] S1 uses organic solvents to prepare the extract of *Ganba* bacteria;

[0031] S2 involves performing normal-phase chromatographic gradient elution separation on the extract of S1 to obtain components separated by different gradients; the eluent used for elution is a mixture of dichloromethane and methanol, with a volume ratio of dichloromethane to methanol of: dichloromethane:methanol = (100-0):(0-100);

[0032] S3 separates the components of S2 by reversed-phase chromatography gradient elution to obtain components separated by different gradients; the eluent used for elution is a mixture of methanol and water, and the volume ratio of methanol to water is: methanol:water = (40-100):(60-0);

[0033] S4 involves isocratic elution of the components from S3 using reversed-phase high-performance liquid chromatography, collecting the eluted components at 13.5 min or 29.8 min to obtain a natural furanolactone-type terphenylquinone compound.

[0034] As a preferred embodiment of the third aspect, the elution step S2 is as follows:

[0035] (1) Elute with an eluent of dichloromethane to methanol in a volume ratio of 100:0 to obtain component Fr.1;

[0036] (2) Elute with an eluent of dichloromethane to methanol in a volume ratio of 98:2 to obtain component Fr.2;

[0037] (3) Elute with an eluent of dichloromethane and methanol in a volume ratio of 95:5 to obtain component Fr.3;

[0038] (4) Elute with an eluent of dichloromethane to methanol in a volume ratio of 9:1 to obtain component Fr.4;

[0039] (5) Elute with an eluent of dichloromethane to methanol in a volume ratio of 8:2 to obtain component Fr.5;

[0040] (6) Elute with an eluent containing dichloromethane and methanol in a volume ratio of 1:1 to obtain component Fr.6;

[0041] (7) Elute with an eluent containing dichloromethane and methanol in a volume ratio of 0:100 to obtain component Fr.7.

[0042] In a preferred embodiment of the third aspect, the component eluted by reversed-phase chromatography in step S3 is Fr.3, and the elution steps are as follows:

[0043] (1) Elution was performed with a methanol to water volume ratio of 40:0 to obtain component Fr.31;

[0044] (2) Elution was performed with a methanol to water volume ratio of 60:40 to obtain component Fr.32;

[0045] (3) Elution was performed with an eluent of methanol and water in a volume ratio of 80:20 to obtain component Fr.33;

[0046] (4) Elute with a methanol to water volume ratio of 100:0 to obtain component Fr.34.

[0047] In a preferred embodiment of the third aspect, the component to be separated by reversed-phase high-performance liquid chromatography in step S4 is Fr.33, and the elution procedure satisfies the following conditions:

[0048] (1) The mobile phase is a mixture of methanol and water, and the volume ratio of methanol to water is: methanol:water = 70:30;

[0049] (2) Washing time: 0-35 min.

[0050] As a preferred embodiment of the third aspect.

[0051] The chemical structure of compound 1 in the eluting component at 29.8 min in S4 is a terphenyl derivative as shown in 002, where R is phenylacetyl.

[0052] The chemical structure of compound 2 of the effluent component in S4 at 13.5 min is a terphenyl derivative as shown in 002, where R is hydrogen.

[0053] The chemical structure of compound 3, which is the eluting component in S4 at 20.6 min, is a terphenyl derivative as shown in 003;

[0054] The chemical structure of compound 4, which is the component that elutes at 11.3 min in S4, is a terphenyl derivative as shown in 001, wherein R1 is a hydroxyl group; R2 is a hydroxyl group; and R3 is a phenylacetyl group.

[0055] The chemical structure of compound 5, which is the component that flows out at 18.4 min in S4, is a terphenyl derivative as shown in 004, where R1 is hydrogen and R2 is phenylacetyl.

[0056] In a preferred embodiment of the third aspect, the organic solvent in S1 is acetone.

[0057] As a preferred embodiment of the third aspect, the preparation method of the *Gynostemma pentaphyllum* extract in S1 includes the following steps: soaking and extracting the dried fruiting body of *Gynostemma pentaphyllum* in acetone using an extraction method, and then concentrating the extract to obtain an acetone extract.

[0058] As a preferred embodiment of the third aspect, it further includes a step of identifying the components collected in step S4.

[0059] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0060] This invention isolates a natural furanolactone-type terphenylquinone compound from the dried fruiting body extract of *Bacteroides rubrum*. Studies have shown that this compound can inhibit inflammatory factors, improve behavior in a mouse model of depression, and has therapeutic effects on neuritis. Attached Figure Description

[0061] Figure 1 Compound 1 1 Schematic diagram of H-NMR spectrum;

[0062] Figure 2 Compound 1 13 Schematic diagram of C-NMR spectrum;

[0063] Figure 3 Schematic diagram of the HSQC spectrum of compound 1;

[0064] Figure 4 Schematic diagram of the HMBC spectrum of compound 1;

[0065] Figure 5 Compound 1 1 H- 1 Schematic diagram of H COSY spectrum;

[0066] Figure 6 Single-crystal diffraction pattern of compound 1;

[0067] Figure 7 Compound 2 1 Schematic diagram of H-NMR spectrum;

[0068] Figure 8 Compound 2 13 Schematic diagram of C-NMR spectrum;

[0069] Figure 9 Schematic diagram of the HSQC spectrum of compound 2;

[0070] Figure 10 Schematic diagram of the HMBC spectrum of compound 2;

[0071] Figure 11 Compound 2 1 H- 1 Schematic diagram of H COSY spectrum;

[0072] Figure 12 A schematic diagram of the single-crystal diffraction pattern of compound 2;

[0073] Figure 13 Compound 3 1 Schematic diagram of H-NMR spectrum;

[0074] Figure 14 Compound 3 13 Schematic diagram of C-NMR spectrum;

[0075] Figure 15 Schematic diagram of the HSQC spectrum of compound 3;

[0076] Figure 16 Schematic diagram of the HMBC spectrum of compound 3;

[0077] Figure 17 Compound 3 1 H- 1 Schematic diagram of H COSY spectrum;

[0078] Figure 18 Single-crystal diffraction pattern of compound 3;

[0079] Figure 19 Compound 4 1 Schematic diagram of H-NMR spectrum;

[0080] Figure 20 Compound 4 13 Schematic diagram of C-NMR spectrum;

[0081] Figure 21 Schematic diagram of the HSQC spectrum of compound 4;

[0082] Figure 22 Schematic diagram of the HMBC spectrum of compound 4;

[0083] Figure 23 Compound 4 1 H- 1 Schematic diagram of H COSY spectrum;

[0084] Figure 24 Single-crystal diffraction pattern of compound 4;

[0085] Figure 25 Compound 5 1 Schematic diagram of H-NMR spectrum;

[0086] Figure 26 Compound 5 13 Schematic diagram of C-NMR spectrum;

[0087] Figure 27 Schematic diagram of the HSQC spectrum of compound 5;

[0088] Figure 28 Schematic diagram of the HMBC spectrum of compound 5;

[0089] Figure 29Compound 5 1 H- 1 Schematic diagram of H COSY spectrum;

[0090] Figure 30 Schematic diagram of the effects of compounds 1-10 on the viability of BV2 microglia;

[0091] Figure 31 Schematic diagram of the effects of compounds 1-9 on LPS-induced release of IL-6, IL-1β and TNF-α from BV2 cells (Figure A: Effect of compounds 1-9 at 10 μM on LPS-induced release of pro-inflammatory factors IL-6, IL-1β and TNF-α from BV2 cells; Figure B: Effect of compounds 1-9 at 20 μM on LPS-induced release of pro-inflammatory factors IL-6, IL-1β and TNF-α from BV2 cells).

[0092] Figure 32 Schematic diagram of the effects of compounds 1-9 on the mRNA expression of IL-6, IL-1β and TNF-α in LPS-stimulated BV-2 cells;

[0093] Figure 33 Schematic diagram of how compound 2 inhibits LPS-mediated neuroinflammatory response in BV2 microglia by suppressing the JAK2 / STAT3 signaling pathway (Figure A: Band diagram of the effect of compound 2 at 20 μM on LPS-induced JAK2 / STAT3 pathway proteins in BV-2 cells; Figure B: Statistical graph of p-JAK2 / JAK2 protein expression; Figure C: Statistical graph of p-STAT3 / STAT3 protein expression; Figure D: Immunofluorescence detection of phosphorylation signal transduction and intracellular localization of p-STAT3).

[0094] Figure 34 A schematic diagram of the animal behavioral experiment of compound 2;

[0095] Figure 35 A schematic diagram of the chemical structure of compound 1;

[0096] Figure 36 A schematic diagram of the chemical structure of compound 2;

[0097] Figure 37 A schematic diagram of the chemical structure of compound 3;

[0098] Figure 38 A schematic diagram of the chemical structure of compound 4;

[0099] Figure 39 A schematic diagram of the chemical structure of compound 5. Detailed Implementation

[0100] To better illustrate the purpose, technical solution, and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments.

[0101] Example 1: Preparation of compounds 1-10

[0102] S1: Preparation of acetone extract:

[0103] The dried fruiting bodies of *Gynostemma pentaphyllum* were extracted three times with acetone (3L), each time for 24 hours. The extract was then concentrated to obtain the acetone extract for later use.

[0104] S2: Preliminary separation of acetone extract:

[0105] Elution column: Normal phase silica gel column (200-300 mesh)

[0106] Eluent: A mixture of dichloromethane and methanol. The elution gradient and elution components are shown in the table below:

[0107] Table 1

[0108] Dichloromethane (v) Methanol (v) Components 100％ 0 Fr.1 98％ 2％ Fr.2 95％ 5％ Fr.3 90％ 10％ Fr.4 80％ 20％ Fr.5 50％ 50％ Fr.6 0 100％ Fr.7

[0109] The specific steps are as follows:

[0110] The acetone extract was passed through a normal-phase silica gel column and eluted with dichloromethane and methanol (dichloromethane:methanol) at volume ratios of 100:0, 98:2, 95:5, 9:1, 8:2, 1:1, and 0:100, respectively. The extracts were combined under reduced pressure, the eluent was removed, and the extract was concentrated to obtain seven fractions: Fr.1 (dichloromethane-methanol 100:0 fraction), Fr.2 (dichloromethane-methanol 98:2 fraction), Fr.3 (dichloromethane-methanol 95:5 fraction), Fr.4 (dichloromethane-methanol 9:1 fraction), Fr.5 (dichloromethane-methanol 8:2 fraction), Fr.6 (dichloromethane-methanol 1:1 fraction), and Fr.7 (dichloromethane-methanol 0:100 fraction). These fractions were then prepared for use.

[0111] S3: Separation of component Fr.3:

[0112] Elution column: ODS column (reversed-phase column, 40-60 μm)

[0113] Eluent: Methanol-water system; elution gradient and elution components are shown in the table below:

[0114] Table 2

[0115] Methanol (v) Water (v) Components 40％ 60％ Fr.31 60％ 40％ Fr.32 80％ 20％ Fr.33 100％ 0 Fr.34

[0116] The specific steps are as follows:

[0117] Fraction Fr.3 was eluted using an ODS column (reversed-phase column) with a gradient of methanol-water system from 40:60 to 100:0, yielding four fractions: Fr.31 (methanol-water 40:60 fraction), Fr.32 (methanol-water 60:40 fraction), Fr.33 (methanol-water 80:20 fraction), and Fr.34 (methanol-water 100:0 fraction).

[0118] S4: Component Fr.33 is separated using reversed-phase high-performance liquid chromatography.

[0119] Chromatographic conditions: ODS column (reversed-phase column 40-60μm)

[0120] Eluent: Methanol-water system; elution gradient and elution components are shown in the table below:

[0121] Table 3

[0122]

[0123] The specific steps are as follows:

[0124] The following compound was obtained by eluting fraction Fr.33 with methanol-water (70:30) in preparative reversed-phase high-performance liquid chromatography:

[0125] Compound 1 (7.2 mg, t) R 29.8 min), its chemical structural formula is as follows Figure 35 As shown, its single-crystal diffraction pattern is as follows: Figure 6 As shown;

[0126] The name of compound 1 is: 4,7-bis(4-hydroxyphenyl)-2,6-dioxo-3-phenyl-2,6-dihydrobenzofuran-5-yl

[0127] 2-phenylacetate.

[0128] Compound 2 (4.6 mg, t) R 13.5 min), its chemical structural formula is as follows Figure 36 As shown, its single-crystal diffraction pattern is as follows: Figure 12 As shown;

[0129] The name of compound 2 is:

[0130] 5-hydroxy-4,7-bis(4-hydroxyphenyl)-3-phenylbenzofuran-2,6-dione.

[0131] Compound 3 (1.1 mg, t) R 20.6 min), its chemical structural formula is as follows Figure 37 As shown, its single-crystal diffraction pattern is as follows: Figure 18 As shown;

[0132] The name of compound 3 is:

[0133] 2,5,6-trihydroxy-4,7-bis(4-hydroxyphenyl)-2-methylbenzofuran-3(2H)-one.

[0134] Compound 4 (6.0 mg, t) R 11.3 min), its chemical structural formula is as follows Figure 38 As shown, R1 is hydroxyl or methoxy; R2 is hydroxyl or methoxy; R3 is hydrogen or phenylacetyl; preferably, R1 is hydroxyl; R2 is hydroxyl; R3 is phenylacetyl, and its single-crystal diffraction pattern is shown below. Figure 24 As shown;

[0135] The name of compound 4 is:

[0136] 4,4",5'-trihydroxy-3',6'-dioxo-3',6'-dihydro-[1,1':4',1"-terphenyl]-2'-yl

[0137] 2-phenylacetate.

[0138] Compound 5 (2.3 mg, t) R 18.4 min), its chemical structural formula is as follows Figure 39 As shown, R1 is hydrogen or phenylacetyl; R2 is hydrogen or phenylacetyl; preferably, R1 is hydrogen; R2 is phenylacetyl. ；

[0139] The name of compound 5 is: 2,7,8-trihydroxy-3-(4-hydroxyphenyl)dibenzo[b,d]furan-1,4-diyl bis(2-phenylacetate).

[0140] S5: Component Fr.4 is separated using reversed-phase high-performance liquid chromatography.

[0141] Chromatographic conditions: ODS column (reversed-phase column 40-60μm)

[0142] Eluent: Methanol-water system; elution gradient and elution components are shown in the table below:

[0143] Table 4

[0144] Methanol (v) Water (v) Components 40％ 60％ Fr.41 60％ 40％ Fr.42 80％ 20％ Fr.43 100％ 0 Fr.44

[0145] The specific steps are as follows:

[0146] Fraction Fr.4 was eluted using an ODS column (reversed-phase column) with a gradient elution system of methanol-water 40:60–100:0 to obtain four fractions: Fr.41 (methanol-water 40:60 fraction), Fr.42 (methanol-water 60:40 fraction), Fr.43 (methanol-water 80:20 fraction), and Fr.44 (methanol-water 100:0 fraction).

[0147] S6: Component Fr.42 is separated using reversed-phase high-performance liquid chromatography.

[0148] Chromatographic conditions: ODS column (reversed-phase column 40-60μm)

[0149] Eluent: Methanol-water system; elution gradient and elution components are shown in the table below:

[0150] Table 5

[0151]

[0152] The specific steps are as follows:

[0153] The following compound was obtained by eluting fraction Fr.42 with methanol-water (70:30) in preparative reversed-phase high-performance liquid chromatography:

[0154] Compound 6(t) R =28.1min, 3mg)

[0155] Compound 8(t) R =33.5min, 7.2mg)

[0156] Compound 9(t) R =37.2min, 5mg)

[0157] Compound 7(t) R =41.0 min, 6 mg)

[0158] Compound 10(t) R =44.5 min, 3 mg).

[0159] S7: Identification of compounds 1-5:

[0160] Nuclear magnetic resonance (NMR) spectra: proton NMR, carbon NMR, and two-dimensional NMR (HMBC, HSQC). 1 H- 1 The determination was performed using H COSY (high-resolution mass spectrometry), high-resolution mass spectrometry, infrared spectroscopy, ultraviolet spectroscopy, circular dichroism spectroscopy, and single-crystal X-ray diffraction. The results are shown in the table below. Figure 1-29 :

[0161] Table 6 Compounds 1-5 11H NMR (150MHz) NMR data (J in Hz).

[0162]

[0163]

[0164] Measured in DMSO-d6.

[0165] Table 7 Compounds 1-5 13 C NMR (150MHz) NMR data (δi n ppm).

[0166] No. Compound 1 Compound 2 Compound 3 Compound 4 Compound 5 1 119.6,C 120.0,C 123.2,C 121.3,C 122.9,C 2,6 132.1,CH 131.9,CH 131.7, CH 131.7, CH 131.1,CH 3,5 114.6, CH 114.0,CH 114.7, CH 114.4,CH 115.3,CH 4 158.1,C 157.8,C 156.8,C 157.0,C 157.2,C 1' 135.0,C ND 113.5,C ND 125.3,C 2' 178.1,C ND 130.3,C ND 133.9,C 3' 142.7,C ND 137.6,C ND 145.8,C 4' 115.3,C ND 110.9,C 114.8,C 114.5,C 5' 136.8,C ND 164.3,C ND 150.1,C 6' 154.7,C ND 115.1,C ND 157.6,C 1” 119.5,C 120.0,C 122.2,C 121.3,C 115.5,C 2” 130.0,CH 130.9, CH 131.5, CH 131.7, CH 146.7,C 3” 115.1,CH 114.9, CH 114.2,CH 114.4,CH 98.9,CH 4” 157.9,C 156.4,C 156.4,C 156.8,C 143.1,C 5” 115.1,CH 114.9,CH 114.2,CH 114.4,CH 142.0,C 6” 130.0,CH 130.9, CH 131.5, CH 131.7, CH 114.3,CH 1”' 167.1,C 167.7,C 196.4,C 169.5,C 170.6,C 2”' 131.2,C 131.9,C 103.1,C <![CDATA[40.7,CH2]]> <![CDATA[40.5,CH2]]> 3”' 128.2,C 129.8,C <![CDATA[22.3,CH3]]> 133.2,C 134.1,C 4”',8”' 128.4,CH 129.8, CH 129.6, CH 129.9, CH 5”',7”' 127.0,CH 127.0,CH 128.5, CH 128.8, CH 6”' 129.3,CH 130.9, CH 127.1,CH 127.4, CH 1”” 169.5,C 170.1,C 2”” <![CDATA[39.5,CH2]]> <![CDATA[40.5,CH2]]> 3”” 133.3,C 134.1,C 4””,8”” 129.4, CH 129.7, CH 5””,7”” 127.3, CH 128.6, CH 6”” 130.0,CH 127.2,CH

[0167] Measured in DMSO-d6.

[0168] Example 2: Investigation of the anti-neuroinflammatory activity of compounds 1-10

[0169] BV2 cells (a mouse microglial cell line) were cultured in DMEM medium containing 10% fetal bovine serum at 37°C and 5% CO2. Cells up to passage 10 were used for all experiments. The compound was dissolved in DMSO to obtain a 10 mM stock solution, and LPS was diluted with PBS to a concentration of 1 mg / mL. BV2 cells (2 × 10⁻⁶ cells) were then cultured. 6 BV-2 cells were seeded overnight in 96-well plates with LPS (100 ng / mL), pretreated with different concentrations of the compound for 1 hour, and then co-cultured with LPS (100 ng / mL) for 24 hours. Cell suspensions or supernatants were collected for analysis. Cell viability was determined using a CCK8 assay kit (E-CK-A362, Elabscience). BV-2 cells were seeded at 2 × 10⁻⁶ ng / mL. 5 Cells were seeded at a density of 10 cells / well in 96-well plates. Results showed that compounds 1-9 did not exhibit significant cytotoxicity against BV-2 microglia at concentrations up to 20 μM. Figure 30 ).

[0170] This study investigated whether compounds 1-9 could inhibit LPS-induced release of pro-inflammatory cytokines in BV2 cells. BV2 cells were treated with different concentrations (10 μM and 20 μM) of the compounds for 24 hours, with and without co-treatment with 100 ng / ml LPS, and the levels of inflammatory factors in the culture supernatant were then measured. The results showed that without compounds 1-9, LPS increased the release of IL-6, IL-1β, and TNF-α from BV2 cells; the addition of compounds 1-9 significantly inhibited LPS-induced release of pro-inflammatory cytokines. Figure 31 and Figure 32Further research revealed that compound 2 can reduce the production of pro-inflammatory cytokines by inhibiting the activation of the JAK2 / STAT3 signaling pathway. Figure 33 ).

[0171] Example 3: Animal behavioral experiments with compound 2

[0172] Among the various pathophysiological mechanisms of depression, mounting evidence suggests that neuroinflammation is a major contributing factor. Neuroinflammation refers to inflammatory processes within the nervous system, characterized by complex multicellular immune responses in the central nervous system (CNS) triggered by acute or chronic pathological stimuli. Microglial-mediated neuroinflammation has now been revealed to play a crucial role in the development of depressive-like behaviors.

[0173] Establishment of an LPS-induced depression model: A mouse depression model was established using LPS. Six-week-old C57BL / 6J mice were purchased and housed in an environment with a constant temperature of 22±1℃ and relative humidity of 55±5%, with ample food and water provided. The light cycle was 12 hours of light / 12 hours of darkness (lighting was turned on at 8:00 AM daily). The mice were allowed to acclimatize for 5 consecutive days to eliminate transport and environmental transition stress. Mice were randomly assigned to three experimental units (n=15 per unit): a control group (Ctrl group), a model group (LPS group), and a treatment group (LPS + compound 2 group). The control group received only saline injection and gavage. The treatment group received compound 2 (5 mg / kg) once daily by gavage for one week. From day 6 onwards, the model and treatment groups received LPS (L2880, Sigma) dissolved in saline and administered intraperitoneally (1 mg / kg) for two consecutive days. Behavioral tests were performed 24 hours after the last LPS injection. After the test, the mice were euthanized, and the tissues were collected and stored at -80°C for subsequent experiments.

[0174] Experimental treatment group: The above model mice were injected with 5 mg / kg of compound 2.

[0175] Sucrose Preference Test (SPT): A two-bottle free choice paradigm was used. Mice were given an acclimatization diet of 1% sucrose solution for 3 days prior to the experiment. After randomization, they were fasted and deprived of water for 24 hours on the third day after administration. The following day, they were provided with two bottles of liquid (tap water and 1% sucrose solution) for free drinking. The bottles were switched every 12 hours to eliminate the influence of positional preference. The formula for calculating the sucrose preference rate is:

[0176] Sucrose preference rate (%) = Total liquid consumption (water + sucrose) / Sucrose solution consumption × 100

[0177] Forced swimming test (FST): Mice were placed in a transparent plexiglass tank (70cm high, 30cm in diameter) filled with clean water at 23±1℃ (30cm deep). Behavioral videos were recorded for 6 minutes, and the immobility time (s) for the last 4 minutes was analyzed. Immobility was defined as static floating or minimal movement only maintaining the head above the water surface; horizontal paddling was swimming, and vertical push-off was climbing.

[0178] Suspended Tail Test (TST): The mouse's tail tip was suspended 1 cm above the ground using medical tape. After 2 minutes of adaptation, the time (s) of immobility for the next 4 minutes was recorded to quantify the degree of behavioral despair.

[0179] The results showed that compound 2 reversed the LPS-induced decrease in sucrose consumption. Figure 34 A), and reduced the stationary time of TST and FST ( Figure 34 B-34C) further confirms that compound 2 has the potential to alleviate LPS-induced neuroinflammation and can be used to treat neuroinflammation.

[0180] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A terphenyl derivative, characterized in that, The terphenyl derivatives have any of the following chemical structural formulas: Wherein, R1 is a hydroxyl group; R2 is a hydroxyl group; and R3 is a phenylacetyl group.

2. The use of the terphenyl derivatives as described in claim 1 in the preparation of drugs for treating neuroinflammatory diseases.

3. The application of terphenyl derivatives in the preparation of drugs for treating neuroinflammatory diseases, characterized in that, The terphenyl derivatives have the following chemical structural formula: Where R is hydrogen.

4. A method for preparing a terphenyl derivative, characterized in that, Includes the following steps: S1 uses acetone to prepare the extract of *Ganba* bacteria; S2 involves performing normal-phase gradient elution separation on the extract from S1 to obtain components separated by different gradients; the eluent used for elution is a mixture of dichloromethane and methanol, with a volume ratio of dichloromethane to methanol of (100-0):(0-100). Specifically, the elution step in step S2 is as follows: (1) Elute with an eluent of dichloromethane to methanol in a volume ratio of 100:0 to obtain component Fr.1; (2) Elute with an eluent of dichloromethane to methanol in a volume ratio of 98:2 to obtain component Fr.2; (3) Elute with an eluent of dichloromethane to methanol in a volume ratio of 95:5 to obtain component Fr.3; (4) Elute with an eluent of dichloromethane to methanol in a volume ratio of 9:1 to obtain component Fr.4; (5) Elute with an eluent of dichloromethane to methanol in a volume ratio of 8:2 to obtain component Fr.5; (6) Elute with an eluent of dichloromethane and methanol in a volume ratio of 1:1 to obtain component Fr.6; (7) Elute with an eluent of dichloromethane to methanol in a volume ratio of 0:100 to obtain component Fr.7; S3 separates the Fr.3 component of S2 by reversed-phase chromatography gradient elution to obtain components separated by different gradients; the eluent used for elution is a mixture of methanol and water, and the volume ratio of methanol to water is: methanol:water = (40-100):(60-0); Specifically, the elution step in step S3 is as follows: (1) Elution with a methanol to water volume ratio of 40:0 yielded component Fr.31; (2) Elution with a methanol to water volume ratio of 60:40 yielded component Fr.32; (3) Elute with a methanol to water eluent in a volume ratio of 80:20 to obtain component Fr.33; (4) Elute with an eluent of methanol and water in a volume ratio of 100:0 to obtain component Fr.34; S4 involves isocratic elution of the Fr.33 fraction from S3 using reversed-phase high-performance liquid chromatography (RP-HPLC). The eluent used is a mixture of methanol and water, with a volume ratio of methanol to water of 70:

30. The fractions eluted at 11.3 min, 13.5 min, 18.4 min, 20.6 min, or 29.8 min are collected to obtain a natural furanolactone-type terphenylquinone compound. Among them, compound 1 of the eluting component at 29.8 min in S4 has the chemical structure shown below; Compound 2, the component that elutes at 13.5 min in S4, has the following chemical structure; Compound 3, the component that elutes at 20.6 min in S4, has the following chemical structure; Compound 4, the component that elutes at 11.3 min in S4, has the following chemical structural formula; Wherein, R1 is a hydroxyl group; R2 is a hydroxyl group; and R3 is a phenylacetyl group. Compound 5, the component that elutes at 18.4 min in S4, has the following chemical structural formula; Where R1 is hydrogen and R2 is phenylacetyl.

5. The preparation method according to claim 4, characterized in that, The component to be separated by reversed-phase high-performance liquid chromatography in step S4 is Fr.33, and the elution time is 0-35 min.

6. The preparation method according to claim 4, characterized in that, The preparation method of the *Gnaphalium affine* extract in S1 includes the following steps: The dried fruiting bodies of *Gynostemma pentaphyllum* were extracted by soaking in acetone using an immersion extraction method, and the acetone extract was obtained after concentration.

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