Lignan compound extracted and separated from pepper leaves as well as preparation method and application of lignan compound

Abstract

The invention discloses lignan compounds extracted and separated from pepper leaves as well as a preparation method and application of the lignan compounds. The lignan compounds are pipernigumin A, pipernigumin B and pipernigumin C, and the lignan compounds are pipernigumin A, pipernigumin B and pipernigumin C; wherein the structural formulas of the pipernigrumin A, the pipernigrumin B and the pipernigrumin C are as follows: # imgabs0 #, and research finds that the pipernigrumin A, the pipernigrumin B and the pipernigrumin C have a remarkable inhibition effect on LPS (Lipopolysaccharide)-induced BV-2 microglial cell inflammation of mice and can be used as active ingredients to be applied to preparation of medicines or / and health-care products for resisting neuroinflammation or preventing and treating Parkinson's disease, so that a new treatment means is provided for treatment and prevention of neurodegenerative diseases.

Description

Technical Field

[0001] The invention relates to the field of natural product extraction, and in particular to a lignan compound extracted and separated from pepper leaves, and a preparation method and application thereof. Background Art

[0002] Parkinson's disease (PD) is an aging-related neurodegenerative disorder characterized by degeneration and loss of dopaminergic neurons in the substantia nigra and the deposition of Lewy bodies. Approximately 10 million people worldwide suffer from PD, and its prevalence increases significantly with age, reaching as high as 3% in people over 80 years of age. PD presents not only with motor symptoms such as resting tremor, muscle rigidity, and bradykinesia, but also with non-motor symptoms such as cognitive impairment and mood disorders, severely impacting patients' quality of life and imposing a heavy socioeconomic burden. Studies have shown that the pathogenesis of PD involves genetic, environmental, and endogenous factors, with microglia-mediated neuroinflammation playing a key role in disease progression. Microglia, the primary immune effector cells of the central nervous system, can be overactivated by stimuli such as lipopolysaccharide (LPS) under pathological conditions, releasing inflammatory mediators such as nitric oxide (NO), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α), leading to neuronal damage and dopaminergic neurodegeneration. Therefore, inhibiting microglia-mediated neuroinflammation has become an important strategy for PD treatment.

[0003] Pepper leaf, a plant of the Piper genus (Piper) in the Piperaceae family, is primarily found in Hainan and Yunnan, China. Modern research indicates that pepper leaf extracts possess antioxidant and antibacterial activities, but research on their anti-neuroinflammatory activity and related active ingredients has been inadequate. Currently, there are no reports demonstrating neuroprotective effects of pepper leaf compounds in a LPS-induced BV-2 microglial cell inflammation model. Therefore, the discovery and application of anti-inflammatory active ingredients from pepper leaf are of great scientific significance and potential economic value. Summary of the Invention

[0004] The first object of the present invention is to provide a lignan compound extracted and separated from pepper leaves, wherein the lignan compound is pipernigrumin A, pipernigrumin B and pipernigrumin C; wherein the structural formulas of pipernigrumin A, pipernigrumin B and pipernigrumin C are as follows:

[0005]

[0006] A second object of the present invention is to provide a method for extracting and isolating lignan compounds pipernigrumin A, pipernigrumin B and pipernigrumin C from pepper leaves, the method comprising the following steps:

[0007] Step S1: chopping pepper leaves and extracting them with 95% ethanol and 75% ethanol respectively under heating and refluxing, combining the multiple extracts, and concentrating under reduced pressure until there is no alcohol taste, thereby obtaining a total extract of pepper leaves; subjecting the total extract of pepper leaves to silica gel column chromatography, eluting with petroleum ether / dichloromethane / ethyl acetate / methanol as eluents in sequence, to obtain four fractions;

[0008] Step S2: The dichloromethane fraction was decolorized by MCI column chromatography and then separated by silica gel column chromatography using a petroleum ether-ethyl acetate system with a gradient elution ratio of 40:1, 20:1, 10:1, 5:1, 2:1, and 1:1, in sequence. The obtained fractions were analyzed by TLC and combined to obtain 9 fractions, namely Fr.A to Fr.I;

[0009] Step S3: The Fr.C fraction was separated and purified by silica gel column chromatography using a petroleum ether-ethyl acetate system with a gradient elution ratio of 15:1, 10:1, 5:1, 2:1, and 1:1, followed by TLC analysis and merging to obtain five fractions, namely Fr.C-1 to Fr.C-5.

[0010] Step S4: Fr.C-1 was subjected to semi-preparative high performance liquid chromatography with isocratic elution using acetonitrile-water as the mobile phase, and the acetonitrile content was 78% to obtain pipernigrumin A;

[0011] Step S5: Fr.C-3 was first isolated and purified by reverse-phase silica gel column chromatography using a methanol-water system with a gradient elution of 50%, 55%, 60%, 65%, 70%, 75%, 80%, and 85% methanol-water. After TLC analysis, the three fractions were combined to obtain Fr.C-3-1 to Fr.C-3-3. Fr.C-3-2 was isolated and purified by semi-preparative high performance liquid chromatography using methanol-water as the mobile phase by isocratic elution, with a methanol content of 90%, to obtain pipernigrumin B.

[0012] Step S6: Fr.F was first separated and purified by LH-20 gel column chromatography, eluting with a dichloromethane-methanol = 1:1 system. After TLC analysis, the four fractions were combined to obtain Fr.F-1 to Fr.F-4. The Fr.F-1 fraction was separated and purified by silica gel column chromatography, eluting with a petroleum ether-ethyl acetate system with a gradient of 6:1, 4:1, 2:1, and 1:1, to obtain pipernigrumin C.

[0013] As a preferred embodiment of the present invention, in step S1, dried pepper leaves are used and soaked with 95% ethanol at a solid-liquid ratio of 1:25 overnight, followed by heating, extraction, and reflux at 170°C for 4 hours. After the extract is cooled, the filtrate is filtered and recovered, and the extract is further heated, extracted, and refluxed at 170°C for 4 hours with 75% ethanol at a solid-liquid ratio of 1:25.

[0014] As preferred of the present invention, when the dichloromethane portion of the pepper leaves after decolorization treatment in step S2 is separated by a normal phase silica gel column, a normal pressure glass chromatography column is used: height 63 cm, inner diameter 12 cm, column temperature: room temperature, flow rate: 5 L / h, pressure: normal pressure; specific conditions are: petroleum ether - ethyl acetate = 40: 1 3BV, petroleum ether - ethyl acetate = 20: 1 3BV, petroleum ether - ethyl acetate = 10: 1 3BV, petroleum ether - ethyl acetate = 5: 1 3BV, petroleum ether - ethyl acetate = 2: 1 3BV, petroleum ether - ethyl acetate = 1: 1 3BV; a total of 9 fractions Fr.A to Fr.I are obtained, Fr.C is the last 1BV of petroleum ether - ethyl acetate = 10: 1 elution and the initial 1BV of petroleum ether - ethyl acetate = 5: 1 elution fraction, and the resulting fraction is concentrated under reduced pressure and evaporated to dryness, and Fr.F is the last 1BV of petroleum ether - ethyl acetate = 2: 1 elution fraction, and the resulting fraction is concentrated under reduced pressure and evaporated to dryness.

[0015] As a preferred embodiment of the present invention, when the fraction Fr.C in step S3 is separated by a normal phase silica gel chromatography column, a normal pressure glass chromatography column is used: height 58 cm, inner diameter 3.2 cm, column temperature: room temperature, flow rate: 1.2 L / h, pressure: normal pressure; specific conditions are: petroleum ether-ethyl acetate = 15:1 3BV, petroleum ether-ethyl acetate = 10:1 3BV, petroleum ether-ethyl acetate = 5:1 3BV, petroleum ether-ethyl acetate = 2:1 3BV, petroleum ether-ethyl acetate = 1:1 3BV; a total of 5 fractions Fr.C-1 to Fr.C-5 are obtained, Fr.C-1 is obtained by combining the elution fractions of petroleum ether-ethyl acetate = 15:1 and concentrating under reduced pressure to evaporate the solvent, and Fr.C-3 is obtained by combining the elution fractions of petroleum ether-ethyl acetate = 5:1 and concentrating under reduced pressure to evaporate the solvent.

[0016] As a preferred embodiment of the present invention, the semi-preparative HPLC column in step S4 is a GRACE Apollo C18 preparative chromatographic column with a column length of 250 mm, a diameter of 10 mm, a filler of 5 μm, a column temperature of room temperature, and a flow rate of 3 mL / min.

[0017] As a preferred embodiment of the present invention, when the step S5 fraction Fr.C-3 is separated by a reverse-phase silica gel chromatography column, a glass chromatography column is used: height 23 cm, inner diameter 1.8 cm, column temperature: room temperature, flow rate: 50 mL / h, pressure: pressurized; specific conditions are: methanol-water = 50% 1BV, methanol-water = 55% 1BV, methanol-water = 60% 2BV, methanol-water = 65% 2BV, methanol-water = 70% 2BV, methanol-water = 75% 3BV, methanol-water = 80% 3BV, methanol-water = 85% 2BV; the methanol-water (65%-75%) elution fractions are combined, and the solvent is recovered by concentration under reduced pressure to obtain fraction Fr.C-3-2.

[0018] As a preferred embodiment of the present invention, when the fraction Fr.F in step S6 is separated by an LH-20 gel chromatography column, a glass chromatography column is used: height 100 cm, inner diameter 1.9 cm, column temperature: room temperature, flow rate: 250 mL / h, pressure: normal pressure; specific conditions are: dichloromethane-methanol = 1:1 4BV, the 2BV fraction eluted with dichloromethane-methanol = 1:1 is combined, and the solvent is recovered after concentration under reduced pressure to obtain fraction Fr.F-1; when Fr.F-1 is separated by a normal phase silica gel chromatography column, a normal pressure glass chromatography column is used: height 30 cm, inner diameter 4.2 cm, column temperature: room temperature, flow rate: 280 mL / h, pressure: normal pressure; specific conditions are: petroleum ether-ethyl acetate = 6:11BV, petroleum ether-ethyl acetate = 4:1 8BV, petroleum ether-ethyl acetate = 2:1 5BV, petroleum ether-ethyl acetate = 1:1 The last 1BV fraction eluted with petroleum ether-ethyl acetate = 2:1 and the first 1BV fraction eluted with petroleum ether-ethyl acetate = 1:1 were combined and concentrated under reduced pressure to recover the solvent. The pipernigrumin C was obtained by repeated recrystallization with dichloromethane-methanol.

[0019] The present invention isolates the lignan compounds pipernigrumin A, pipernigrumin B, and pipernigrumin C from a pipernigrum leaf extract for the first time and discloses for the first time the neuroprotective activity of pipernigrumin A, pipernigrumin B, and pipernigrumin C in a LPS-induced BV-2 microglial inflammation model. Therefore, the pipernigrumin A, pipernigrumin B, and pipernigrumin C provided by the present invention can be used in the preparation of anti-neuroinflammatory drugs and / or health products.

[0020] The lignan compound pipernigrumin A separated from the pepper leaf extract of the present invention has neuroprotective activity and can be used in the preparation of medicines and / or health products for treating or preventing neurodegenerative diseases.

[0021] The lignan compound pipernigrumin B separated from the pepper leaf extract of the present invention has neuroprotective activity and can be used in the preparation of medicines and / or health products for treating or preventing neurodegenerative diseases.

[0022] The lignan compound pipernigrumin C separated from the pepper leaf extract of the present invention has neuroprotective activity and can be used in the preparation of medicines and / or health products for treating or preventing neurodegenerative diseases.

[0023] As a preferred embodiment of the present invention, the neurodegenerative diseases include Parkinson's disease, Alzheimer's disease, Huntington's disease, amyotrophic lateral sclerosis, and Lou Gehrig's disease.

[0024] The present invention also provides a drug for resisting neuroinflammation, wherein the active ingredient of the drug is any one of the above-mentioned lignan compounds pipernigrumin A, pipernigrumin B, and pipernigrumin C, or a mixture of any two or more thereof.

[0025] Advantages and beneficial effects of the present invention:

[0026] (1) The present invention is the first to extract and isolate the lignan compounds pipernigrumin A, pipernigrumin B and pipernigrumin C from pepper leaves, and discloses for the first time the neuroprotective activity of pipernigrumin A, pipernigrumin B and pipernigrumin C against the LPS-induced BV-2 microglial cell inflammation model.

[0027] (2) The lignan compounds pipernigrumin A, pipernigrumin B and pipernigrumin C extracted and separated from pepper leaves provided by the present invention have neuroprotective activity and can be used in the preparation of anti-neuroinflammatory drugs and / or health products, and can also be used in the preparation of drugs and / or health products for treating or preventing neurodegenerative diseases including Parkinson's disease, thereby providing a new therapeutic means for the treatment and prevention of neurodegenerative diseases.

[0028] (3) Any one of pipernigrumin A, pipernigrumin B and pipernigrumin C provided by the present invention, or a mixture of two or more thereof, can be used as an active ingredient in an anti-neuroinflammatory drug. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 The MS spectrum of compound pipernigruminA(1);

[0030] Figure 2 For the compound pipernigruminA(1) 1 H-NMR nuclear magnetic resonance spectrum;

[0031] Figure 3 For the compound pipernigruminA(1) 13 C-NMR nuclear magnetic resonance spectrum;

[0032] Figure 4 is the HSQC NMR spectrum of compound pipernigruminA(1);

[0033] Figure 5 is the HMBC nuclear magnetic resonance spectrum of compound pipernigruminA(1);

[0034] Figure 6 For the compound pipernigruminA(1) 1 H- 1 H NOESY NMR spectrum;

[0035] Figure 7 is the ECD spectrum of compound pipernigruminA(1);

[0036] Figure 8 The MS spectrum of compound pipernigrumin B (2) is shown in FIG.

[0037] Figure 9 For the compound pipernigrumin B(2) 1 H-NMR nuclear magnetic resonance spectrum;

[0038] Figure 10 For the compound pipernigrumin B(2) 13 C-NMR nuclear magnetic resonance spectrum;

[0039] Figure 11 is the HSQC NMR spectrum of compound pipernigrumin B(2);

[0040] Figure 12 is the HMBC nuclear magnetic resonance spectrum of compound pipernigrumin B (2);

[0041] Figure 13 For the compound pipernigrumin B(2) 1 H- 1 H NOESY NMR spectrum;

[0042] Figure 14is the ECD spectrum of compound pipernigrumin B(2);

[0043] Figure 15 is the MS spectrum of compound pipernigrumin C;

[0044] Figure 16 For the compound pipernigrumin C(3) 1 H-NMR nuclear magnetic resonance spectrum;

[0045] Figure 17 For the compound pipernigrumin C(3) 13 C-NMR nuclear magnetic resonance spectrum;

[0046] Figure 18 is the HSQC NMR spectrum of compound pipernigrumin C(3);

[0047] Figure 19 is the HMBC NMR spectrum of compound pipernigrumin C(3);

[0048] Figure 20 For the compound pipernigrumin C(3) 1 H- 1 H NOESY NMR spectrum;

[0049] Figure 21 is the ECD spectrum of compound pipernigrumin C(3);

[0050] Figure 22 are the safe dosing concentrations of compounds pipernigruminA-C for BV-2 cells; wherein, (A) is the safe dosing concentration of pipernigruminA for BV-2 cells; (B) is the safe dosing concentration of pipernigrumin B for BV-2 cells; and (C) is the safe dosing concentration of pipernigrumin C for BV-2 cells. DETAILED DESCRIPTION

[0051] The present invention is not limited to the following embodiments, and specific implementation methods can be determined based on the technical solutions of the present invention and actual conditions. Unless otherwise specified, the various chemical reagents and chemicals mentioned in the present invention are all chemical reagents and chemicals commonly known in the prior art; unless otherwise specified, percentages in the present invention are all volume percentages; unless otherwise specified, solutions in the present invention are all aqueous solutions with water as the solvent, for example, ethanol solutions are ethanol-water solutions; normal temperature and room temperature in the present invention generally refer to temperatures between 15°C and 25°C, and are generally defined as 25°C.

[0052] The present invention will be further described below in conjunction with the embodiments:

[0053] 1. Reagents and Instruments

[0054] 1.1 Reagents and medicinal materials

[0055] Silicone GF 254 Thin layer plates (Qingdao Ocean Chemical Co., Ltd.); Sephadex LH-20 (GE Healthcare Bio-Science AB); MCI GEL (Mitsubishi Chemical Corporation, Japan); normal phase silica gel for column chromatography (60-80 mesh, Qingdao Ocean Chemical Co., Ltd.); normal phase silica gel for column chromatography (200-300 mesh, Qingdao Ocean Chemical Co., Ltd.); LiChroprep RP-18 (40-63 μm, Merck KGaA, Germany). Chromatographic methanol (Thermo Fisher Scientific, USA, chromatographic grade); chromatographic acetonitrile (Thermo Fisher Scientific, USA, chromatographic grade); methanol, ethanol, petroleum ether, dichloromethane, and ethyl acetate were all analytical grade (Beijing Modern Oriental Science and Technology Development Co., Ltd.). 0.25% trypsin (containing EDTA) (Gibco, USA); DMEM high-glucose culture medium (Shanghai Xiaopeng Biotechnology Co., Ltd.); fetal bovine serum (FBS) (Mengma Tianjin Biotechnology Co., Ltd.); penicillin-streptomycin mixture (Thermo Fisher Scientific, USA); dimethyl sulfoxide (DMSO) (Sigma, USA); MTT (Beijing Solebao Technology Co., Ltd.); LPS (Sigma-Aldrich, USA); L-NMMA (NG-monomethyl-L-arginine) (Biyuntian Biotechnology Co., Ltd.); nitric oxide detection kit (Pulile Gene Technology Co., Ltd.).

[0056] Pepper leaves were harvested from the Xinglong Tropical Botanical Garden in Wanning, Hainan Province. Dr. Wang Yu of the Institute of Tropical Biotechnology, Chinese Academy of Tropical Agricultural Sciences, identified them as fresh leaves of Pipernigrum L., a plant of the Piperaceae family (Piperaceae). The samples were naturally shade-dried and then chopped for later use. Samples of the original plant are currently stored in the Laboratory for Pharmacodynamic Substances Research at the School of Traditional Chinese Medicine, Capital Medical University.

[0057] 1.2 Cell lines

[0058] Mouse BV-2 microglial cells were purchased from the Chinese Academy of Medical Sciences.

[0059] 1.3 Instruments

[0060] N-2110 rotary evaporator (Eyela Co., Ltd., Japan); intelligent electric heating mantle (model: SXKW-10000ML, Linmao Technology Co., Ltd., Beijing), vacuum pump (Zhengzhou Great Wall Science and Technology Industry and Trade Co., Ltd.); ME235S 1 / 100000 electronic analytical balance (Sartorius Scientific Instrument Co., Ltd., Germany); LC-20AD semi-preparative high-performance liquid chromatograph (Shimadzu Corporation, Japan); GRACE Apollo preparative C18 column (10 mm × 250 mm, 5 μm) (GRACE, USA); Avance II 500 MHz nuclear magnetic resonance spectrometer (Bruker, Germany), UV-6100S ultraviolet spectrophotometer (Shimadzu Corporation, Japan), ThermoFisher FT-IR iS7 infrared spectrometer (Thermo Fisher, USA), Jasco 810 circular dichroism spectrometer (Jasco, Japan), Fourier cyclotron resonance high-resolution mass spectrometer (Bruker, Germany). ESCO AC2-4S1 ultra-clean biosafety cabinet (Thermo Fisher Scientific, USA); Model 371 carbon dioxide constant temperature incubator (Thermo Fisher Scientific, USA); MSL-3020 high pressure sterilizer (Sanyo Corporation, Japan); XDTD-8222 constant temperature blast drying oven (Shanghai Jinghong Laboratory Equipment Co., Ltd.); hemocytometer (Shanghai Qiujing Biochemical Company); BCD-254 refrigerator (Siemens, Germany); ULT-2186-4-v49 ultra-low temperature freezer (Thermo Fisher Scientific, USA); Primo Vert inverted microscope (Carl Zeiss, Germany); SpectraMax Plus 384 full-wavelength microplate reader (Molecular Devices, USA).

[0061] 2. Experimental Methods

[0062] 2.1 Extraction and separation of lignan compounds

[0063] Step S1: chopping dried pepper leaves (5.0 kg) and extracting them with 95% ethanol and 75% ethanol (solid-liquid ratio of 1:25) respectively, heating and refluxing for 4 hours, combining the multiple extracts, and concentrating under reduced pressure until there is no alcohol taste, thereby obtaining a total pepper leaf extract (862 g); subjecting the total pepper leaf extract to silica gel column chromatography, and eluting with petroleum ether / dichloromethane / ethyl acetate / methanol as eluents in sequence, to obtain a petroleum ether fraction (144 g), a dichloromethane fraction (96 g), an ethyl acetate fraction (67 g), and a methanol fraction (440 g).

[0064] Step S2: The dichloromethane fraction (96 g) decolorized by MCI column chromatography was dissolved in anhydrous ethanol and mixed with normal phase silica gel (60-80 mesh, 100 g). 3 kg (200-300 mesh) of normal phase silica gel was wet-packed into the column. The packing was tapped tightly to fill the column. Then, the sample-mixed normal phase silica gel from which the solvent had been evaporated was slowly added to the column. The developing solvent for column analysis was petroleum ether-ethyl acetate. Column separation system: atmospheric pressure glass chromatography column: height 63 cm, inner diameter 12 cm, column temperature: room temperature, flow rate: 5 L / h, pressure: atmospheric pressure; specific conditions: petroleum ether-ethyl acetate = 40:1 3BV, petroleum ether-ethyl acetate = 20:1 3BV, petroleum ether-ethyl acetate = 10:1 3BV, petroleum ether-ethyl acetate = 5:1 3BV, petroleum ether-ethyl acetate = 2:1 3BV, petroleum ether-ethyl acetate = 1:1 3BV; the obtained fractions were combined after TLC detection and concentrated under reduced pressure to recover the solvent, giving a total of 9 fractions Fr.A to Fr.I. Fr.C was obtained by combining the last 1BV of the fractions eluted with petroleum ether-ethyl acetate (10:1) and the first 1BV of the fractions eluted with petroleum ether-ethyl acetate (5:1), followed by concentration under reduced pressure to evaporate the solvent. Fr.F was obtained by combining the last 1BV of the fractions eluted with petroleum ether-ethyl acetate (2:1), followed by concentration under reduced pressure to evaporate the solvent.

[0065] Step S3: Dissolve Fr.C (3.4 g) in anhydrous ethanol and mix with normal phase silica gel (60-80 mesh, 4 g). Take 250 g (200-300 mesh) of normal phase silica gel and wet-pack it into the column. Tap the filler tightly and fill it solidly. Then, slowly add the mixed normal phase silica gel from which the solvent has been evaporated into the column. The developing solvent for column analysis is petroleum ether-ethyl acetate. Column separation system: atmospheric pressure glass chromatography column: height 58 cm, inner diameter 3.2 cm, column temperature: room temperature, flow rate: 1.2 L / h, pressure: atmospheric pressure; specific conditions: petroleum ether-ethyl acetate = 15:1 3BV, petroleum ether-ethyl acetate = 10:1 3BV, petroleum ether-ethyl acetate = 5:1 3BV, petroleum ether-ethyl acetate = 2:1 3BV, petroleum ether-ethyl acetate = 1:1 3BV; a total of 5 fractions Fr.C-1 to Fr.C-5 were obtained, Fr.C-1 was obtained by combining the petroleum ether-ethyl acetate (15:1) elution fractions and concentrating under reduced pressure to evaporate the solvent, and so on.

[0066] Step S4: Fr.C-1 (422 mg) was subjected to semi-preparative HPLC using acetonitrile-water (78%, 3 mL / min) as the mobile phase for isocratic elution to obtain pipernigrumin A (59 mg). Column separation system: GRACE Apollo C18 preparative column, 250 mm length, 10 mm diameter, 5 μm filler, column temperature: room temperature, DAD detection wavelengths: 210, 254, and 280 nm.

[0067] Step S5: Fr.C-3 is obtained by recovering the solvent by concentrating under reduced pressure the elution fraction of petroleum ether-ethyl acetate 5:1 of Fr.C. Fr.C-3 (1 g) is dissolved in anhydrous ethanol and mixed with RP-18 reverse-phase silica gel (1 g). 22 g of reverse-phase silica gel is wet-packed into a column, the filler is tapped and filled, and then the mixed reverse-phase silica gel from which the solvent has been evaporated is slowly added to the column. The developing solvent for column analysis conditions is petroleum ether-ethyl acetate. Column separation system: a glass chromatography column was used: 23 cm high and 1.8 cm inner diameter, column temperature: room temperature, flow rate: 50 mL / h, pressure: pressurized; specific conditions were: methanol-water = 50% 1BV, methanol-water = 55% 1BV, methanol-water = 60% 2BV, methanol-water = 65% 2BV, methanol-water = 70% 2BV, methanol-water = 75% 3BV, methanol-water = 80% 3BV, methanol-water = 85% 2BV; the methanol-water (65%-75%) elution fractions were combined, and the solvent was recovered by concentration under reduced pressure to obtain the fraction Fr.C-3-2. Fr.C-3-2 (700 mg) was purified by semi-preparative HPLC using methanol-water (90%, 3 mL / min) as the mobile phase for isocratic elution to obtain pipernigrumin B (17 mg); column separation system: GRACE Apollo C18 preparative column, column length 250 mm, diameter 10 mm, filler 5 μm, column temperature: room temperature, DAD detection wavelengths: 210, 254, 280 nm.

[0068] Step S6: Fr.F (17.1 g) was separated by LH-20 gel chromatography using a glass column: 100 cm high, 1.9 cm inner diameter, column temperature: room temperature, flow rate: 250 mL / h, pressure: atmospheric pressure; specific conditions: dichloromethane-methanol = 1:1 4 BV, the 2 BV fractions eluted with dichloromethane-methanol (1:1) were combined, and the solvent was recovered by vacuum concentration to obtain fraction Fr.F-1. Fr.F-1 (1.8 g) was dissolved in anhydrous ethanol and mixed with normal phase silica gel (60-80 mesh, 2 g). 100 g (200-300 mesh) of normal phase silica gel was wet-packed into the column, the packing was tapped to pack tightly, and then the solvent-free normal phase silica gel was slowly added to the column; the developing solvent for column analysis was petroleum ether-ethyl acetate. Column separation system: atmospheric pressure glass chromatography column: height 30 cm, inner diameter 4.2 cm, column temperature: room temperature, flow rate: 280 mL / h, pressure: atmospheric pressure; specific conditions: petroleum ether-ethyl acetate = 6:1 1BV, petroleum ether-ethyl acetate = 4:1 8BV, petroleum ether-ethyl acetate = 2:1 5BV, petroleum ether-ethyl acetate = 1:1 5BV; the last 1BV eluted with petroleum ether-ethyl acetate (2:1) and the first 1BV eluted with petroleum ether-ethyl acetate (1:1) were combined and concentrated under reduced pressure to recover the solvent, and pipernigrumin C (5 mg) was repeatedly recrystallized from dichloromethane-methanol.

[0069] The structures of the compounds pipernigrumin A, pipernigrumin B, and pipernigrumin C obtained in this example were analyzed by nuclear magnetic resonance spectroscopy (NMR), infrared spectroscopy (IR), ultraviolet spectroscopy (UV), electron circular dichroism (ECD), and high-resolution mass spectrometry (HR-ESI-MS). The structures of the new compounds were identified with the help of microspectral databases and SciFinder Scholar, and the structural formulas were determined as follows:

[0070]

[0071] The structural identification results are as follows:

[0072] Pipernigrumin A (1) is a colorless oil. UV (MeOH) λ max (logε)227(1.24),284(0.93)nm; IRν max 1514,1487,1439,1236,1138,1028,926cm -1 ; +18.0(c 0.05,MeOH); ECD(MeOH)λ max (Δε) 205 (-17.2), 218 (-1.0), 223 (-1.5), 286 (-1.0) nm. According to HR-ESI-MS m / z 423.1801 [M+Na] + (calculated value: 423.1783) and NMR data, the molecular formula was determined to be C 23 H 28 O6, with an unsaturation degree of 10. 1H NMR data combined with HSQC spectrum showed two groups of trisubstituted aromatic ring signals δ H 6.64 (1H, m), 6.78 (1H, d, J = 8.1 Hz), 6.70 (1H, m), 6.69 (1H, d, J = 1.0 Hz), 6.70 (1H, m), 6.63 (1H, m), 4 benzyl hydrogen signals δ H 2.53 (1H, dd, J = 5.2, 13.6 Hz), 2.73 (1H, m), 2.47 (1H, m), 2.75 (1H, m), 2 methine signals δ H 1.99 (1H, m), 2.41 (1H, m), 1 acetal hydrogen signal δ H 4.77 (1H, d, J = 4.6 Hz), 1 methylene signal δ H3.58 (1H, dd, J = 7.0, 8.3 Hz), 4.00 (1H, t, J = 8.2 Hz), 2 methoxy signals δ H 3.86 (3H, s), 3.87 (3H, s), 1 methylenedioxy signal δ H 5.92 (2H, d, J = 1.1 Hz), and one ethoxy signal δ H 3.37 (1H, m), 3.70 (1H, m), 1.20 (1H, t, J=7.1Hz). 13 C NMR data combined with HSQC spectrum showed one ethoxy carbon signal δ C 15.4, 62.9, 2 benzyl carbon signals δ C 33.8, 39.5, two methine carbon signals δ C 43.5, 52.4, two methoxy carbon signals δ C 56.0, 56.1, 1 methylene carbon signal δ C 72.4, 1 acetal carbon signal δ C 104.3, 1 methylenedioxy carbon signal δ C 100.9, 12 aromatic carbon signals δ C 149.1, 147.6, 147.6, 145.8, 135.1, 133.1, 121.8, 120.7, 112.0, 111.4, 109.5, 108.2 (Table 1). Further data analysis and literature comparison revealed that 1 had a high degree of similarity to the NMR data of the known compound rel-(8R,8'S,9R)-3,4-dimethoxy-3',4'-methylenodioxy-9α-ethoxy-lignans-8.8',9.O.9'. Compared with the known compound, their molecular formula and molecular weight were the same, with only the C-7(1:δ C 33.8; known compounds: δ C 38.6) and C-9(1:δ C 104.3; known compounds: δ C The chemical shifts of 109.2) were significantly different, suggesting that 1 is a configurational isomer of the known compound. HSQC and HMBC spectral analysis confirmed that both compounds have the same planar structure.

[0073] The larger coupling constants (J 8,9 =4.6Hz) indicates that H-8 and H-9 are on the same side, in cis configuration. NOESY spectrum shows that H-8 (δ H 1.99) and H-7'(δ H 2.47, δ H2.75) has a related point, and H-8'(δ H 2.41) and H-7(δ H 2.73) showed a correlation point, indicating that H-8 and H-8' were on opposite sides, in a trans configuration. This led to the deduction of the relative configuration of 1. By comparing the measured and calculated ECD curves, the structural formula of 1 was determined to be (8R,8'R,9S)-3,4-dimethoxy-3',4'-methylenodioxy-9-ethoxy-9,9'-epoxylignan, and the compound was named pipernigrumin A.

[0074] The structures of the known compounds rel-(8R,8'S,9R)-3,4-dimethoxy-3',4'-methylenodioxy-9α-ethoxy-lignans-8.8',9.O.9' and pipernigrumin A, as well as the key HMBC and NOESY related signals of pipernigrumin A are as follows:

[0075]

[0076] Pipernigrumin B(2) is a colorless oil. UV(MeOH)λ max (logε)227(1.19),284(0.92)nm; IRν max 1513,1487,1441,1237,1033,922cm -1 ; +14.0(c 0.05,MeOH); ECD(MeOH)λ max (Δε) 208 (-10.2), 215 (-1.7), 218 (-2.1), 224 (-1.5), 231 (-2.0), 288 (-1.2) nm. HR-ESI-MS showed an m / z of 409.1637 [M+Na] + The quasi-molecular ion peak (calculated value: 409.1627) confirmed that its molecular formula was C 22 H 26 O6, unsaturation is 10.

[0077] Compound 2 1 The resonance signals shown in the H NMR spectrum can be attributed to two trisubstituted benzene rings, four benzyl hydrogens, two methine groups, one acetal hydrogen, one methylene group, one methoxy group and one methylenedioxy group connected to the benzene ring, and one ethoxy group. 13The C NMR spectrum gave carbon signals that completely corresponded to the above groups (Table 1). The above spectral data are similar to those of 1, indicating that it also has a monoepoxy lignan skeleton. Compared with 1, the NMR spectrum of 2 lacks a group of methoxy signals (2: δ H 3.88, δ C 55.9;1:δ H 3.86, 3.87, δ C 56.0, 56.1), but the molecular formula comparison results show that 2 to 1 only lacks one group of CH2 atoms (2:C 22 H 26 O6;1:C 23 H 28 O6), it is speculated that the hydroxyl group may replace one methoxy group in 2. In the HMBC spectrum, the methylenedioxy group (δ H 5.97) and C-3(δ C 147.7) are related, H-2(δ H 6.60) and C-6 (δ C 121.9) has relevant descriptions of methylenedioxy located at C-3 and C-4; methoxy (δ H 3.88) and C-3'(δ C 146.5) showed that the methoxy group was connected to the C-3' position. C The chemical shift of 144.0) is located downfield, suggesting that an oxygen-containing group is attached here. Combined with the aforementioned molecular formula comparison results, it is confirmed that a hydroxyl group is attached at the C-4' position. Thus, the planar structure of 2 is confirmed.

[0078] The smaller coupling constants (J 8,9 =1.5Hz) indicates that H-8 and H-9 are on opposite sides and in trans configuration. H 2.18) and H-10(δ H 3.74) further confirms the configuration. In the NOESY spectrum, H-8 (δ H 2.18) and H-7'(δ H 2.60) and H-8'(δ H 2.17) and H-7(δ HThe correlation points (2.47) indicate that H-8 and H-8' are on opposite sides, exhibiting a trans configuration. Based on the trans relationships of H-8 / H-9 and H-8 / H-8', the absolute configuration of 2 was inferred to be (8S,8'S,9S) or (8R,8'R,9R). ECD spectroscopy ultimately determined the structural formula of 2 to be (8S,8'S,9S)-3'-methoxy-4'-hydroxy-3,4-methylenedioxy-9-ethoxy-9,9'-epoxylignan, and the compound was named pipernigrumin B.

[0079] The key HMBC and NOESY related signals of pipernigrumin B are as follows:

[0080]

[0081] Pipernigrumin C(3) is a white amorphous solid. UV(MeOH)λ max (logε)227(1.18),284(0.84)nm; IRν max 3276,2931,2878,1414,1108,1039,995,924,676cm -1 ; -4.0(c 0.05,MeOH); ECD(MeOH)λ max (Δε)206(-15.4),221(-1.1),226(-1.4),286(-1.0)nm. HR-ESI-MS gives its quasi-molecular ion peak [M+Na] + m / z 469.1850 (calculated value: 469.1838), and its molecular formula was determined to be C 24 H 30 O8, unsaturation is 10.

[0082] Compound 3 1 The H NMR data are similar to those of 1, including two trisubstituted benzene rings, four benzyl hydrogens, two methine groups, one acetal hydrogen, one methylene group, two methoxy groups connected to the benzene ring, and one methylenedioxy group. 13 The C NMR spectrum gave carbon signals that completely corresponded to the above groups (Table 1). The above spectral data showed that 3 was a monoepoxy lignan compound. The most significant difference between 3 and 1 was that 3 lacked an ethoxy signal (1: δ H 3.42, 3.74, 1.22; δ C 15.4, 63.0), and one more glyceryl signal (δ H 3.51, 3.56, 3.60, 3.64, 3.76; δC 64.0, 70.4, 71.0). In the HMBC spectrum, H-10 (δ H 3.51), H-11(δ H 3.56) and H-12(δ H 3.60) and C-9(δ C 110.3) is related, indicating that the glyceryl group is connected to C-9 through the oxygen atom at C-10; the methylenedioxy group (δ H 5.92) and C-3(δ C 147.7) are related, H-2(δ H 6.54) and C-6(δ C 121.8) has relevant descriptions of methylenedioxy located at C-3 and C-4; methoxy (δ H 3.85) and C-3'(δ C 149.1) and methoxy (δ H 3.83) and C-4'(δ C 147.9) Correlation indicates that the two methoxy groups are attached at the C-3' and C-4' positions, respectively. In summary, the planar structure of 3 is confirmed.

[0083] In the NOESY spectrum, H-8(δ H 2.17) and H-11(δ H 3.64) have related signals, indicating that H-8 and H-9 are located on opposite sides and present a trans configuration; H-8 (δ H 2.17) and H-7'(δ H The correlation point (2.60) indicates that H-8 and H-8' are on opposite sides, exhibiting a trans configuration. Therefore, its absolute configuration was inferred to be (8R,8'R,9S) or (8S,8'S,9R). ECD spectroscopy ultimately determined the structural formula of 3 to be (8R,8'R,9S)-3',4'-dimethoxy-3,4-methylenodioxy-9-glycerol-9,9'-epoxylignan, and the compound was named pipernigrumin C.

[0084] The key HMBC and NOESY related signals of pipernigrumin C are as follows:

[0085]

[0086] of pipernigrumin A, pipernigrumin B and pipernigrumin C 1 H and 13 C NMR data are shown in Table 1

[0087] Table 1 Pipernigrumin A-C 1 H (500 MHz, CDCl3) and 13 C NMR (125 MHz, CDCl3) data

[0088]

[0089]

[0090] 2.2 Neuroprotective activity of the new compounds pipernigrumin A, pipernigrumin B, and pipernigrumin C in the LPS-induced BV-2 microglial cell inflammation model

[0091] 2.2.1 Cell culture

[0092] Remove BV-2 cells from a -80°C freezer and quickly thaw in a 37°C water bath. Transfer cells to a centrifuge tube and add 4 mL of cell culture medium (DMEM containing 1% penicillin-streptomycin solution and 10% fetal bovine serum). Centrifuge at 1000 rpm for 3 minutes, discard the supernatant, resuspend the cells in 1 mL of cell culture medium, transfer them to a 60 mm cell culture dish, add 4 mL of cell culture medium, mix well, and culture in a cell incubator (37°C, 5% CO2). Change the culture medium every two days, observe cell morphology, and maintain the cells in the logarithmic growth phase until ready for use.

[0093] 2.2.2 Safe Dosing Concentration Screening

[0094] Take BV-2 cells in the logarithmic growth phase, discard the old culture medium, add 4 mL of new cell culture medium to make a single cell suspension, count the cells, and adjust the cell density to 2.5×10 5 / mL, 100μL cell suspension was added to each well of a 96-well plate for plating. After culturing for 24 hours, the old cell culture medium was discarded, and the control group was given 100μL of new cell culture medium. The drug-treated groups were added with different concentrations of pipernigruminA, pipernigrumin B and pipernigrumin C monomer compounds, and their concentrations were 2.5, 5, 5, 10, 20, and 40μg / mL, respectively. Three replicates were set for each concentration. After another 24 hours of incubation, MTT 10μL / well (5mg / mL) was added, and the cells were incubated in the dark for another 2.5 hours. The absorbance value at a wavelength of 570nm was read in a microplate reader. Cell viability = (OD 实验组 -OD 空白组 ) / (OD 对照组 -OD 空白组The drug concentration range in which the cell survival rate is above 80% is the safe drug administration concentration for BV-2 cells.

[0095] 2.2.3 Activity screening

[0096] Take BV-2 cells in the logarithmic growth phase, discard the old culture medium, add 4 mL of new cell culture medium to make a single cell suspension, count the cells, and adjust the cell density to 2.5×10 5 Cells were plated at 100 μL of cell suspension per well in a 96-well plate. After 24 hours of culture, the old cell culture medium was discarded and 100 μL of fresh cell culture medium was administered to the control and model groups. Different concentrations of pipernigrumin A, pipernigrumin B, and pipernigrumin C monomer compounds were added to the drug-treated groups, with concentrations of 1.25, 2.5, 5, 7.5, 10, and 15 μg / mL, respectively. Different concentrations of L-NMMA were added to the positive control group, with three replicates for each concentration. After 4 hours of culture, 1 μL of 1 mg / mL LPS solution was added to each well of all groups except the control group, and cultured for another 24 hours. The NO concentration in the cell supernatant was measured according to the instructions of the nitric oxide detection kit. NO production rate = NO concentration in the experimental group / NO concentration in the control group * 100%. The IC was calculated by fitting the NO production curve at different concentrations. 50 value.

[0097] 2.2.4 Results

[0098] The safe administration concentration of monomeric compounds in pepper leaf extract to BV-2 cells is shown in Figure 22 The results of neuroprotective activity screening on LPS-induced BV-2 cell inflammation model are shown in Table 2. As shown in Table 2, the IC values of pipernigrumin A, pipernigrumin B and pipernigrumin C in pepper leaves for inhibiting NO release are 50 The values were 7.28±0.25μg / mL, 11.51±0.34μg / mL, and 3.80±0.53μg / mL, respectively. It can be seen that the compounds pipernigrumin A, pipernigrumin B, and pipernigrumin C in pepper leaves have good neuroprotective activity against the LPS-induced BV-2 microglial cell inflammation model.

[0099] Table 2 Half-inhibitory concentration of pipernigrumin A-C on NO production in LPS-induced BV-2 cell inflammation model

[0100]

[0101]

[0102] The above results show that the new lignan compounds pipernigrumin A, pipernigrumin B and pipernigrumin C extracted and isolated from pepper leaves of the present invention have good neuroprotective activity, and the effect is better than that of the positive control group. Therefore, they can be used in the preparation of drugs and / or health products for preventing and treating neurodegenerative diseases (Parkinson's disease).

[0103] The above technical features constitute the embodiments of the present invention, which have strong adaptability and implementation effect. Non-essential technical features can be added or removed according to actual needs to meet the requirements of different situations.

Claims

1. A lignan compound extracted and separated from pepper leaves, characterized in that: The lignan compounds are pipernigrumin A, pipernigrumin B, and pipernigrumin C; wherein the structural formulas of pipernigrumin A, pipernigrumin B, and pipernigrumin C are as follows:

2. A method for extracting and separating lignan compounds pipernigrumin A, pipernigrumin B and pipernigrumin C from pepper leaves, characterized in that: The following steps are involved: Step S1: chopping pepper leaves and extracting them with 95% ethanol and 75% ethanol respectively under heating and refluxing, combining the multiple extracts, and concentrating under reduced pressure until there is no alcohol taste, thereby obtaining a total extract of pepper leaves; subjecting the total extract of pepper leaves to silica gel column chromatography, eluting with petroleum ether / dichloromethane / ethyl acetate / methanol as eluents in sequence, to obtain four fractions; Step S2: The dichloromethane fraction was decolorized by MCI column chromatography and then separated by silica gel column chromatography using a petroleum ether-ethyl acetate system with a gradient elution ratio of 40:1, 20:1, 10:1, 5:1, 2:1, and 1:1, in sequence. The obtained fractions were analyzed by TLC and combined to obtain 9 fractions, namely Fr.A to Fr.I; Step S3: The Fr.C fraction was separated and purified by silica gel column chromatography using a petroleum ether-ethyl acetate system with a gradient elution ratio of 15:1, 10:1, 5:1, 2:1, and 1:1, followed by TLC analysis and merging to obtain five fractions, namely Fr.C-1 to Fr.C-5. Step S4: Fr.C-1 was subjected to semi-preparative high performance liquid chromatography with isocratic elution using acetonitrile-water as the mobile phase, and the acetonitrile content was 78% to obtain pipernigrumin A; Step S5: Fr.C-3 was first isolated and purified by reverse-phase silica gel column chromatography using a methanol-water system with a gradient elution of 50%, 55%, 60%, 65%, 70%, 75%, 80%, and 85% methanol-water. After TLC analysis, the three fractions were combined to obtain Fr.C-3-1 to Fr.C-3-3. Fr.C-3-2 was isolated and purified by semi-preparative high performance liquid chromatography using methanol-water as the mobile phase by isocratic elution, with a methanol content of 90%, to obtain pipernigrumin B. Step S6: Fr.F was first separated and purified by LH-20 gel column chromatography, eluting with a dichloromethane-methanol = 1:1 system. After TLC analysis, the four fractions were combined to obtain Fr.F-1 to Fr.F-4. The Fr.F-1 fraction was separated and purified by silica gel column chromatography, eluting with a petroleum ether-ethyl acetate system with a gradient of 6:1, 4:1, 2:1, and 1:1, to obtain pipernigrumin C.

3. The method according to claim 2, characterized in that In step S1, dried pepper leaves are soaked in 95% ethanol at a solid-liquid ratio of 1:25 overnight, then heated at 170° C. for extraction and reflux for 4 hours. After the extract is cooled, the filtrate is filtered and recovered, and the extract is further heated at 170° C. for extraction and reflux for 4 hours with 75% ethanol at a solid-liquid ratio of 1:

25.

4. The method according to claim 2, characterized in that When the dichloromethane portion of the pepper leaf after the decolorization treatment in step S2 is separated by a normal phase silica gel column, a normal pressure glass chromatography column is used: height 63 cm, inner diameter 12 cm, column temperature: room temperature, flow rate: 5 L / h, pressure: normal pressure; specific conditions are: petroleum ether-ethyl acetate = 40:1 3BV, petroleum ether-ethyl acetate = 20:1 3BV, petroleum ether-ethyl acetate = 10:1 3BV, petroleum ether-ethyl acetate = 5:1 3BV, petroleum ether-ethyl acetate = 2:1 3BV, petroleum ether-ethyl acetate = 1:1 3BV; a total of 9 fractions Fr.A to Fr.I are obtained, Fr.C is the result of merging the last 1BV of the flow fraction eluted with petroleum ether-ethyl acetate = 10:1 and the initial 1BV of the flow fraction eluted with petroleum ether-ethyl acetate = 5:1, and concentrating under reduced pressure to evaporate the solvent; Fr.F is the result of merging the last 1BV of the flow fraction eluted with petroleum ether-ethyl acetate = 2:1, and concentrating under reduced pressure to evaporate the solvent.

5. The method according to claim 4, characterized in that When the fraction Fr.C in step S3 was separated by normal phase silica gel chromatography, a normal pressure glass chromatography column was used: height 58 cm, inner diameter 3.2 cm, column temperature: room temperature, flow rate: 1.2 L / h, pressure: atmospheric pressure; specific conditions were: petroleum ether:ethyl acetate = 15:1 3BV, petroleum ether:ethyl acetate = 10:1 3BV, petroleum ether:ethyl acetate = 5:1 3BV, petroleum ether:ethyl acetate = 2:1 3BV, petroleum ether:ethyl acetate = 1:1 3BV; a total of five fractions Fr.C-1 to Fr.C-5 were obtained, Fr.C-1 was obtained by combining the fractions eluted with petroleum ether:ethyl acetate = 15:1 and concentrating under reduced pressure to evaporate the solvent, and Fr.C-3 was obtained by combining the fractions eluted with petroleum ether:ethyl acetate = 5:1 and concentrating under reduced pressure to evaporate the solvent; In step S6, when fraction Fr.F is separated by LH-20 gel chromatography column, a glass chromatography column is used: height 100 cm, inner diameter 1.9 cm, column temperature: room temperature, flow rate: 250 mL / h, pressure: normal pressure; specific conditions are: dichloromethane-methanol = 1:1 4BV, the 2BV fractions eluted with dichloromethane-methanol = 1:1 are combined, and the solvent is recovered after concentration under reduced pressure to obtain fraction Fr.F-1; when Fr.F-1 is separated by normal phase silica gel chromatography column, a normal pressure glass chromatography column is used: height 30 cm, inner diameter 4.2 cm, column temperature: room temperature, flow rate: 280 mL / h, pressure: normal pressure; specific conditions are: petroleum ether-ethyl acetate = 6:1 1BV, petroleum ether-ethyl acetate = 4:1 8BV, petroleum ether-ethyl acetate = 2:1 5BV, petroleum ether-ethyl acetate = 1:1 The last 1BV fraction eluted with petroleum ether-ethyl acetate = 2:1 and the first 1BV fraction eluted with petroleum ether-ethyl acetate = 1:1 were combined and concentrated under reduced pressure to recover the solvent. The pipernigrumin C was obtained by repeated recrystallization with dichloromethane-methanol.

6. The method according to claim 5, characterized in that Step S4: Semi-preparative HPLC chromatography column: GRACEApollo C18 preparative chromatography column, column length 250 mm, diameter 10 mm, filler 5 μm, column temperature: room temperature, flow rate: 3 mL / min; When the fraction Fr.C-3 in step S5 is separated by reverse-phase silica gel chromatography, a glass chromatography column is used: height 23 cm, inner diameter 1.8 cm, column temperature: room temperature, flow rate: 50 mL / h, pressure: pressurized; specific conditions are: methanol-water = 50% 1BV, methanol-water = 55% 1BV, methanol-water = 60% 2BV, methanol-water = 65% 2BV, methanol-water = 70% 2BV, methanol-water = 75% 3BV, methanol-water = 80% 3BV, methanol-water = 85% 2BV; the elution fractions with methanol-water ratio of 65%-75% are combined, and the solvent is recovered by concentration under reduced pressure to obtain fraction Fr.C-3-2.

7. Use of the lignan compound pipernigrumin A, pipernigrumin B or pipernigrumin C according to claim 1 in the preparation of anti-neuroinflammatory drugs and / or health products.

8. Use of the lignan compound pipernigrumin A, pipernigrumin B or pipernigrumin C according to claim 1 in the preparation of a medicine or / and health product for treating or preventing neurodegenerative diseases.

9. The use according to claim 8, characterized in that The neurodegenerative diseases include Parkinson's disease, Alzheimer's disease, Huntington's disease, amyotrophic lateral sclerosis, and Lou Gehrig's disease.

10. A drug for anti-neuroinflammatory, characterized in that: The active ingredient in the medicine is any one of the lignan compounds pipernigrumin A, pipernigrumin B, and pipernigrumin C described in claim 1, or a mixture of any two or more thereof.

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