Phenolic compound, preparation method thereof and application of phenolic compound in preparation of medicine for treating perimenopausal depression

Abstract

The invention relates to the technical field of natural pharmaceutical chemistry, in particular to a phenolic compound, a preparation method thereof and application of the phenolic compound to preparation of drugs for treating perimenopausal depression. The phenolic compound provided by the invention has a structure as shown in a formula I or a formula II, and data of the embodiment shows that the compound with the structure as shown in the formula I and the compound with the structure as shown in the formula II have better anti-PMD activity, can be used for preparing the medicine for treating perimenopausal depression, and fills the technical blank. # imgabs0 #

Description

Technical Field

[0001] The invention relates to the technical field of natural drug chemistry, and in particular to a phenolic compound and a preparation method thereof and application of the phenolic compound in preparing a drug for treating perimenopausal depression. Background Art

[0002] Perimenopausal depression (PMD) is a common and highly prevalent mental disorder in women worldwide, which seriously endangers physical and mental health. PMD is different from ordinary depression. The main causes of PMD are the functional decline of female ovaries and low estrogen levels. Conventional antidepressants have defects such as slow onset and poor clinical anti-PMD efficacy, while hormone replacement therapy (HRT) is also limited in clinical application due to its many adverse effects such as breast cancer. Therefore, it is particularly urgent to develop new, efficient and safe anti-PMD drugs. Although there is no special record of perimenopausal depression in ancient Chinese medical books, its clinical symptoms are similar to the traditional Chinese medical "depression syndrome", "visceral agitation", "lily disease" and "plum pit qi". According to traditional Chinese medicine, the basic pathogenesis of perimenopausal depression is kidney deficiency and liver depression, with kidney deficiency as the root and liver depression as the symptom, that is, kidney deficiency in women, coupled with liver qi stagnation, causes organ dysfunction, which leads to imbalance of yin and yang, disharmony of qi and blood, and loss of nourishment of the mind. Therefore, "tonifying the kidney, replenishing essence and benefiting marrow" is the basic principle of traditional Chinese medicine in treating "depression syndrome".

[0003] The traditional Chinese medicine for kidney tonification, epimedium, has estrogen-like effects. It not only improves HPA axis dysfunction, but also regulates the activity of aromatase CYP19, a key enzyme in estrogen synthesis, and promotes the synthesis of estrogen in human ovarian granulosa cells. Indran et al. found through a nude mouse model that compared with the synthetic estrogen ethinyl estradiol, epimedium extract has estrogen receptor (ER) selectivity, does not increase the risk of breast cancer, and has better safety. In addition, icariin can effectively increase the expression of hypothalamic hormone receptor ERα in PMD model rats, and activate the PI3K-AKT signaling pathway mediated by ERα, improve brain neurotransmitter disorders, and play an anti-PMD role. The classic prescription Erxian Decoction warms kidney yang, replenishes kidney essence, and regulates Chong and Ren meridians. It is mainly used to treat perimenopausal syndrome (PMS). It can upregulate the secretion of ovarian aromatase and increase the level of synthetic estradiol. Previous studies by our research group also showed that Erxian Decoction can effectively improve the depressive behavior of PMD model mice, upregulate the level of estradiol in the body, significantly improve the disorder of female endocrine hormones in model animals, increase the expression of estrogen receptors ERα and ERβ, and increase the expression of BDNF in the hippocampus of the brain. Miao Mingsan et al. reported that curculigoside can effectively improve the disorder of female endocrine hormones in perimenopausal depression model mice and increase the concentration levels of 5-HT and DA in the brain. Various studies have shown that treating perimenopausal depression from the perspective of liver and kidney can greatly improve the symptoms of perimenopausal depression in patients without increasing the risk of adverse reactions. Therefore, traditional Chinese medicine has great potential in treating PMD.

[0004] Curculigo capitulata is a perennial herbaceous plant of the Curculigo family. Curculigo capitulata is used as medicine with its rhizome, which has the effect of warming yang and tonifying the kidney. The Chinese Materia Medica records that Curculigo capitulata can tonify the kidney and consolidate essence, and is mainly used to treat kidney deficiency cough and asthma, impotence and spermatorrhea, uterine cold and infertility, etc. It is also clearly recorded in the local Chinese medicine records of Sichuan, Yunnan, Guangxi and other places. Curculigo plants mainly contain two types of characteristic components: simple phenolic compounds represented by curculigoside and orcinol glucoside, and norlignan compounds with diarylpentane (Ph-5C-Ph) as the basic skeleton. Whether the phenolic compounds with pyrrolidin-2-one substituents in Curculigo capitulata have anti-PMD activity has not been reported so far. Summary of the invention

[0005] In order to solve the problems existing in the prior art, the present invention provides a phenolic compound and a preparation method thereof and application thereof in preparing a drug for treating perimenopausal depression. The phenolic compound provided by the present invention has anti-PMD activity and fills a technical gap.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0007] The present invention provides a phenolic compound having a structure shown in Formula I or Formula II:

[0008]

[0009] The present invention provides a method for preparing the phenolic compound described in the above technical solution, comprising the following steps:

[0010] The dried rhizome of Curculigo orchioides and an ethanol aqueous solution are mixed, and reflux extraction and concentration are performed in sequence to obtain an extract;

[0011] The extract is sequentially extracted with a weak acid aqueous solution and a weak alkali aqueous solution to obtain a weak alkali extract;

[0012] Extracting the weak base extract with ethyl acetate to obtain an ethyl acetate extract;

[0013] The ethyl acetate extract is subjected to a first silica gel column chromatography separation, and a first dichloromethane-methanol system is used for the first elution, and according to the elution order of the components, a Fr.2.1 fraction and a Fr.2.2 fraction are obtained;

[0014] The Fr.2.1 fraction is subjected to a second silica gel column chromatography separation, and a second dichloromethane-methanol system is used for a second elution. The obtained silica gel column chromatography eluate is subjected to a first dextran gel column chromatography, and methanol is used for elution. According to the order of component elution, three fractions Fr.2.1.1 to Fr.2.1.3 are obtained;

[0015] Sequentially subjecting the Fr.2.1.3 fraction to a second dextran gel column chromatography separation, a medium pressure preparative chromatography column separation, and a third silica gel column chromatography separation to obtain a phenolic compound having a structure represented by Formula I or Formula II;

[0016] During the second dextran gel column chromatography separation, the eluent is a mixture of dichloromethane and methanol;

[0017] When separating by medium-pressure preparative chromatographic column, the eluent is methanol-water solution;

[0018] During the third silica gel column chromatography, the eluent was dichloromethane-acetone system.

[0019] Preferably, the first elution is a gradient elution using a first dichloromethane-methanol system in which the volume ratio of dichloromethane to methanol varies from 50:1 to 10:1.

[0020] Preferably, the second elution is a gradient elution using a second dichloromethane-methanol system in which the volume ratio of dichloromethane to methanol varies from 20:1 to 1:1.

[0021] Preferably, the volume ratio of dichloromethane to methanol in the mixed solution of dichloromethane and methanol is 1:1.

[0022] Preferably, the volume ratio of dichloromethane to acetone in the dichloromethane-acetone system is 15:1.

[0023] Preferably, the volume concentration of the methanol aqueous solution is 30%.

[0024] Preferably, the weak acid aqueous solution includes a 3 wt. % hydrochloric acid aqueous solution or a 3 wt. % acetic acid aqueous solution.

[0025] Preferably, the weak alkali aqueous solution comprises a 3 wt. % sodium hydroxide aqueous solution.

[0026] The present invention also provides the use of the phenolic compound described in the above technical solution or the phenolic compound prepared by the above preparation method in the preparation of a drug for treating perimenopausal depression.

[0027] The present invention provides a phenolic compound having a structure shown in Formula I or Formula II. The data of the embodiments show that the compound having a structure shown in Formula I and the compound having a structure shown in Formula II provided by the present invention have good anti-PMD activity, can be used to prepare a drug for treating perimenopausal depression, and fills a technical gap. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 Schematic diagram of the chemical structure of compound 1 and compound 2;

[0029] Figure 2 Comparison of behavioral tests of mice in each group, where note: Sham is the sham operation group; Model is the model group; CF (Low) is the low-dose group of compound 1, CF (High) is the high-dose group of compound 2; CG (Low) is the low-dose group of compound 2, CG (High) is the high-dose group of compound 2; Positive is the positive control group. ###P<0.001,***P<0.001;

[0030] Figure 3 Comparison of serum estrogen levels in mice of each group; ###P<0.001, ***P<0.001;

[0031] Figure 4 Comparison of the neurotransmitter content in the hippocampus of mice in each group; ###P<0.001, ***P<0.001;

[0032] Figure 5 The expression of Erβ, TrKB and BDNF in the hippocampus of mice in each group; ###P<0.001, ***P<0.001. DETAILED DESCRIPTION

[0033] The present invention provides a phenolic compound having a structure shown in Formula I or Formula II:

[0034]

[0035] The present invention also provides a method for preparing the phenolic compound described in the above technical solution, comprising the following steps:

[0036] The dried rhizome of Curculigo orchioides and an ethanol aqueous solution are mixed, and reflux extraction and concentration are performed in sequence to obtain an extract;

[0037] The extract is sequentially extracted with a weak acid aqueous solution and a weak alkali aqueous solution to obtain a weak alkali extract;

[0038] Extracting the weak base extract with ethyl acetate to obtain an ethyl acetate extract;

[0039] The ethyl acetate extract is subjected to a first silica gel column chromatography separation, and a first dichloromethane-methanol system is used for the first elution, and according to the elution order of the components, a Fr.2.1 fraction and a Fr.2.2 fraction are obtained;

[0040] The Fr.2.1 fraction is subjected to a second silica gel column chromatography separation, and a second dichloromethane-methanol system is used for a second elution. The obtained silica gel column chromatography eluate is subjected to a first dextran gel column chromatography, and methanol is used for elution. According to the order of component elution, three fractions Fr.2.1.1 to Fr.2.1.3 are obtained;

[0041] Sequentially subjecting the Fr.2.1.3 fraction to a second dextran gel column chromatography separation, a medium pressure preparative chromatography column separation, and a third silica gel column chromatography separation to obtain a phenolic compound having a structure represented by Formula I or Formula II;

[0042] During the second dextran gel column chromatography separation, the eluent is a mixture of dichloromethane and methanol;

[0043] When separating by medium-pressure preparative chromatographic column, the eluent is methanol-water solution;

[0044] During the third silica gel column chromatography, the eluent was dichloromethane-acetone system.

[0045] The invention mixes the dried rhizome of Curculigo macrophylla with an ethanol aqueous solution, and sequentially performs reflux extraction and concentration to obtain an extract.

[0046] As an embodiment, the volume concentration of the ethanol aqueous solution is 95%.

[0047] As an implementation mode, the usage ratio of the dry rhizome of Curculigo orchioides to the ethanol aqueous solution is 6kg:30L.

[0048] As an implementation mode, the reflux extraction is performed twice, and the time of each extraction is 3 hours.

[0049] After obtaining the extract, the present invention extracts the extract sequentially with a weak acid aqueous solution and a weak alkali aqueous solution to obtain a weak alkali extract.

[0050] As an embodiment, the weak acid aqueous solution includes a 3wt.% hydrochloric acid aqueous solution or a 3wt.% acetic acid aqueous solution; as an embodiment, the weak base aqueous solution includes a 3wt.% sodium hydroxide aqueous solution.

[0051] After obtaining the weak base extract, the invention extracts the weak base extract with ethyl acetate to obtain an ethyl acetate extract.

[0052] As an embodiment, the extraction is performed three times.

[0053] After obtaining the ethyl acetate extract, the present invention performs a first silica gel column chromatography separation on the ethyl acetate extract, uses a first dichloromethane-methanol system for a first elution, and obtains the Fr.2.1 fraction and the Fr.2.2 fraction according to the elution order of the components.

[0054] As an embodiment, the first elution is a gradient elution using a first dichloromethane-methanol system in which the volume ratio of dichloromethane to methanol varies from 50:1 to 10:1.

[0055] After obtaining the Fr.2.1 fraction, the present invention performs a second silica gel column chromatography separation on the Fr.2.1 fraction, uses a second dichloromethane-methanol system for a second elution, and performs a first dextran gel column chromatography on the obtained silica gel column chromatography eluate, using methanol for elution, and obtains three fractions, Fr.2.1.1 to Fr.2.1.3, in the order of component elution.

[0056] As an embodiment, the second elution is a gradient elution using a second dichloromethane-methanol system in which the volume ratio of dichloromethane to methanol varies from 20:1 to 1:1.

[0057] After obtaining the Fr.2.1.3 fraction, the present invention sequentially subjects the Fr.2.1.3 fraction to a second dextran gel column chromatography separation, a medium pressure preparative chromatography column separation and a third silica gel column chromatography to obtain a phenolic compound of the structure shown in Formula I or Formula II.

[0058] As an embodiment, during the second dextran gel column chromatography separation, the eluent is a mixture of dichloromethane and methanol; the volume ratio of dichloromethane to methanol in the mixture of dichloromethane and methanol is 1:1.

[0059] As an embodiment, during separation using a medium-pressure preparative chromatography column, the eluent is a methanol-water solution, and the volume concentration of the methanol-water solution is 30%.

[0060] As an embodiment, during the third silica gel column chromatography, the eluent is a dichloromethane-acetone system, and the volume ratio of dichloromethane to acetone in the dichloromethane-acetone system is 15:1.

[0061] The preparation method of the compound having the structure shown in Formula I and Formula II provided by the present invention is simple, easy to operate, environmentally friendly and material-saving.

[0062] The present invention also provides the use of the phenolic compound described in the above technical solution or the phenolic compound prepared by the preparation method described in the above technical solution in the preparation of a drug for treating perimenopausal depression.

[0063] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0064] Example 1

[0065] The dried rhizomes of Curculigo orchioides (about 6 kg) were extracted twice with 30 L of 95 vol.% ethanol aqueous solution at 60°C by reflux extraction in a multifunctional vacuum extraction and concentration tank for 3 hours each time. The extracts were combined and distilled under reduced pressure until no extract remained, to obtain about 1250 g of ethanol extract of Curculigo orchioides.

[0066] The obtained extract was extracted with 1 L of 3 wt. % hydrochloric acid aqueous solution, and the extract was mixed with 1 L of 3 wt. % sodium hydroxide aqueous solution, followed by extraction with ethyl acetate three times. The extract was concentrated to obtain 36 g of an extract.

[0067] 36 g of the ethyl acetate extract was separated by silica gel column chromatography and eluted with a dichloromethane-methanol system (the volume ratio of dichloromethane to methanol was changed from 50:1→1:1) to obtain Fr.2.1 fraction and Fr.2.2 fraction.

[0068] 10gFr.2.1 fraction was separated by silica gel column chromatography and eluted with dichloromethane-methanol system (the volume ratio of dichloromethane and methanol changed from 20:1→1:1) to obtain eluate; the obtained eluate was separated by Sephadex LH-20 column chromatography and eluted with methanol to obtain Fr.2.1.1 fraction, Fr.2.1.2 fraction and Fr.2.1.3 fraction.

[0069] The 1.5 g Fr.2.1.3 fraction was separated by Sephadex LH-20 column chromatography and eluted with a mixture of dichloromethane and methanol (the volume ratio of dichloromethane to methanol was 1:1). The eluate was separated by medium pressure preparative chromatography (ODS column separation and eluted with 30 vol.% methanol aqueous solution. The eluate was separated by silica gel column chromatography and eluted with a dichloromethane-acetone system (the volume ratio of dichloromethane to acetone was 15:1) to obtain 410 mg of compound 1 with the structure shown in Formula I and 380 mg of compound 2 with the structure shown in Formula II.

[0070] 1. Detection:

[0071] The present invention performs ultraviolet spectrum detection, infrared spectrum detection, nuclear magnetic resonance hydrogen spectrum detection, carbon spectrum detection and high-resolution mass spectrum detection on compound 1 and compound 2.

[0072] The data of compound 1 are characterized as: amorphous powder; -29.32(c 0.1,MeOH); UV(MeOH)λ max (logε)208(3.09),284(2.46)nm; IRν max :3393.0,2678.1,1646.5,1593.2,1427.0,1372.7,1280.6,1160.7,1060.9,984.9,824.5,641.0,589.0,514.8cm -1 ; 1 H (600 MHz, MeOH), 13 C (175MHz, MeOH) NMR data, see Table 1; (+)-HRESIMS m / z 230.0826 [M+Na] + (calcd.forC 11 H 13 NNaO3230.0793).

[0073] The data of compound 2 are characterized as: amorphous powder; -29.32(c 0.1,MeOH); UV(MeOH)λ max (logε)208(3.09),284(2.46)nm; IRν max :3393.0,2678.1,1646.5,1593.2,1427.0,1372.7,1280.6,1160.7,1060.9,984.9,824.5,641.0,589.0,514.8cm -1 ; 1 H (600 MHz, MeOH), 13C (175MHz, MeOH) NMR data, see Table 1; (+)-HRESIMS m / z 230.0826 [M+Na] + (calcd.forC 11 H 13 NNaO3230.0793).

[0074] Table 1 H NMR and C NMR data of compound 1 and compound 2 in CD3OD

[0075]

[0076]

[0077] 2. Therapeutic effects of compound 1 and compound 2 on PMD model mice

[0078] 2.1. Experimental animals

[0079] SPF female ICR mice; 8 weeks old; weight 20-25 g; provided by: Shanghai Slake Laboratory Animal Co., Ltd.; Animal Ethics Certificate Number: LLSC20220879.

[0080] 2.2. Preparation of bilateral ovariectomy (OVX) model

[0081] The mice were anesthetized with isoflurane, fixed in a supine position, and the skin was prepared. The skin from the lowest rib edge to the sacrum of the rat's back was fully disinfected with iodine, and then a longitudinal incision with a long diameter of about 2 cm was made. The skin and fascia were cut layer by layer, the waist muscle layer was dissected, and the mouse cauliflower-like ovaries were found and pulled out of the fat tissue. The ovaries were removed after ligation along the distal end of the cervix to stop bleeding. After the operation, the remaining tissues were returned to the abdominal cavity, the muscles and skin were sutured and fully disinfected with iodine, and the same operation was performed on both sides. Penicillin was injected intramuscularly at a dose of 200,000 U / mouse to prevent infection. The sham operation group opened the abdominal cavity after anesthesia, found the ovaries but did not remove them, only the fat mass of the same size near them was removed, and sutured in the same way. Other surgical operations were the same as those in the surgical group.

[0082] The mice were randomly divided into 7 groups, with 8 mice in each group: sham operation group, model group, positive control group, low-dose and high-dose groups of compound 1, and low-dose and high-dose groups of compound 2.

[0083] 2.3. Establishment of the chronic unpredictable mild stress (CUMS) model

[0084] On the 8th day after surgery, rats in each group were subjected to CUMS modeling. Stimulation included: all mice except the sham operation group were subjected to CUMS stimulation, and were housed in single cages. The stimulation included fasting for 24 hours, water deprivation for 24 hours, moist bedding for 24 hours, tilting the cage for 12 hours, swimming in ice water for 5 minutes, shaking the cage for 15 minutes, ultrasonic stimulation for 1 hour, and reversing black and white. One stimulus was randomly selected every day so that the mice could not anticipate the occurrence of the stimulus to avoid adaptation. Each stimulus was applied 2 to 3 times on average, and the same stimulus was not repeated in the same mouse within two days for a total of 21 days.

[0085] Mice in the low-dose group (8.65 mg / kg) of compound 1, high-dose group (70 mg / kg) of compound 1, low-dose group (8.65 mg / kg) of compound 2 and high-dose group (70 mg / kg) of compound 2 were gavaged with corresponding doses of compounds 1 h before CUMS intervention, once a day; mice in the sham operation group and the model group were given an equal volume (1 mL / 100 g) of 0.9% NaCl solution; mice in the positive control group were given 0.21 g / mL fluoxetine hydrochloride co-estradiol suspension, and all were given drugs continuously for 21 days.

[0086] 2.4. Evaluation of depressive behavior

[0087] Behavioral measurements were performed after 21 days of treatment.

[0088] 2.4.1. Forced swim test (FST)

[0089] The mice were placed in a cylindrical transparent container with a height of 50 cm and a diameter of 20 cm. Each mouse was tested for 6 minutes, with the first 2 minutes as adaptation time and the last 4 minutes to observe and record the duration of the mouse floating on the water surface (slight movements were allowed to maintain oral and nasal breathing). The water was changed after each test.

[0090] 2.4.2. Tail suspension test (TST)

[0091] Tie one end of the suspension rope to 2 cm from the mouse's tail, hang the mouse in the air (about 50 cm from the ground), and secure it with tape to prevent it from falling off due to struggling during the experiment. After 1 min of adaptation, record the duration of immobility of the mouse in the hanging state within 5 min.

[0092] 2.4.3. Open field test (OFT)

[0093] An open-field reaction box (80 cm × 80 cm × 40 cm) with black walls was used. Its bottom surface consisted of 25 blocks of equal area, and the camera was placed directly above the open-field reaction box. During the experiment, the mice were placed in the central grid at the bottom of the box, and the camera was used to record the activities of the mice within 3 minutes. After each test, the reaction box was wiped with 75% ethanol to avoid the "predecessor effect". The video was analyzed using SuperMaze V2.0 software, and the total distance of the mouse's horizontal movement (Path length) was used as the evaluation index.

[0094] 2.4.4 Behavioral measurement results

[0095] The immobility time of mice in the model group in the forced swimming and tail suspension tests was significantly longer than that in the sham operation group ( Figure 2 AB, ##P<0.01, ###P<0.001, ***P<0.001), indicating that the model was successfully established; the immobility time of mice in the high-dose and low-dose groups of compound 1, high-dose and low-dose groups of compound 2, and the positive control group in the forced swimming and tail suspension tests was shortened to varying degrees. In the open field test, the path length of mice in the model group was significantly shorter than that in the sham operation group, and the path length of mice in the high-dose and low-dose groups of compound 1, high-dose and low-dose groups of compound 2, and the positive control group was significantly increased compared with the model group ( Figure 2 (C) The results of forced swimming test, tail suspension test and open field test showed that compound 1 and compound 2 had an improving effect on PMD.

[0096] 3.5 Determination of hormone levels in serum of PMD mice

[0097] After the behavioral experiment, blood was collected from the orbits of rats in each group. The whole blood was centrifuged (at 3000 rpm) for 15 min after standing at room temperature for 4 h. The supernatant was collected as serum, which was stored at -80 °C for standby use and thawed at 4 °C before use. The enzyme-linked immunosorbent assay (ELISA) method was used to detect the levels of estradiol (E2), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) in the serum of mice in each group. The detection steps were carried out according to the instructions of the kit.

[0098] Compared with the normal control group, the serum E2 content of mice in the model control group was significantly reduced (P<0.001), and the LH and FSH contents were significantly increased; compared with the model, the serum E2 content of mice in the high-dose and low-dose groups of compound 1, the high-dose and low-dose groups of compound 2 and the positive control group was significantly increased (P<0.001), and the LH and FSH contents were significantly decreased. Figure 3 .

[0099] 2.6. Determination of neurotransmitter levels in serum of PMD mice

[0100] The levels of dopamine (DA), norepinephrine (NE), and 5-hydroxytryptamine (5-HT) in the serum obtained in 2.5 were detected by ELISA. Compared with the normal control group, the serum DA, NE, and 5-HT levels of mice in the model control group were significantly reduced (P<0.001); compared with the model, the serum DA, NE, and 5-HT levels of mice in the high- and low-dose groups of compound 1, the high- and low-dose groups of compound 2, and the positive control group were significantly increased (P<0.001). Figure 4 .

[0101] 2.7. Study on the anti-PMD mechanism of action of compounds

[0102] 2.7.1. Tissue protein extraction

[0103] After the behavioral experiment, mice in each group were killed by cervical dislocation, and the hippocampal tissue was quickly separated on ice, frozen in liquid nitrogen, and stored in a -80℃ refrigerator for later use. Take the tissue block and add an appropriate volume of RIPA lysis buffer, and homogenize it with an automatic grinder. Centrifuge at 4℃, 12000rpm for 10min, and take the supernatant. The protein concentration was measured using the BCA kit. Add 5× Loading Buffer to dilute it to 1×, vortex mix, and place in a 100℃ metal bath for 5min, and store at -20℃ for later use.

[0104] 2.7.2.Western Blot

[0105] Prepare 10% SDS-PAGE gel separation gel and concentrated gel, load the sample, and continue electrophoresis at a constant voltage of 80V for about 30 minutes. After the sample enters the separation gel with bromophenol blue, adjust the voltage to 120V, and determine the electrophoresis time according to the molecular weight of the protein. After the electrophoresis, refer to the molecular weight shown by the marker, cut the gel in the required protein range, and transfer the protein to the PVDF membrane at a constant current of 250mA. After the transfer, 5% skim milk powder (TBST is prepared with TBS buffer containing 0.1% Tween-20, and blocked at room temperature for 1h. After the blocking, the PVDF membrane was incubated with the primary antibody at 4°C overnight. The next day, the membrane was washed 3 times with TBST, incubated with the secondary antibody (1:9000) at room temperature for 1h, and washed 3 times with TBST. The PVDF membrane was covered with ECL luminescent liquid, and then placed in a molecular gel imaging system for development and image acquisition. The bands were analyzed using Image Lab 5.1 software. The expression level of the target protein is expressed by its grayscale ratio to the internal reference GAPDH.

[0106] 2.7.3 Experimental results

[0107] The results showed that compared with the sham operation group, the TrKB, BDNF, and Erα in the Model group were significantly decreased (p<0.05 or p<0.01). After drug intervention, the high-dose and low-dose groups of compound 1, the high-dose and low-dose groups of compound 2, and the positive control group could significantly upregulate the expression of TrkB, BDNF, and ERβ (p<0.05). Figure 5 .

[0108] From the above experimental results, it can be seen that compounds 1 and 2 significantly improve the depressive-like behavior induced by OVX-CUMS by regulating the expression of TrKB, BDNF, and Erβ in the hippocampus, increasing serum estradiol, reducing serum follicle-stimulating hormone (FSH) and luteinizing hormone (LH), and promoting the release of neurotransmitters DA, NE, and 5-HT.

[0109] Although the above embodiment describes the present invention in detail, it is only a part of the embodiments of the present invention, not all of the embodiments. Other embodiments can be obtained based on this embodiment without creativity, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A phenolic compound, characterized in that It has the structure shown in Formula I or Formula II:

2. The method for preparing a phenolic compound according to claim 1, characterized in that: The following steps are involved: The dried rhizome of Curculigo orchioides and an ethanol aqueous solution are mixed, and reflux extraction and concentration are performed in sequence to obtain an extract; The extract is sequentially extracted with a weak acid aqueous solution and a weak alkali aqueous solution to obtain a weak alkali extract; Extracting the weak base extract with ethyl acetate to obtain an ethyl acetate extract; The ethyl acetate extract is subjected to a first silica gel column chromatography separation, and a first dichloromethane-methanol system is used for the first elution, and according to the elution order of the components, a Fr.2.1 fraction and a Fr.2.2 fraction are obtained; The Fr.2.1 fraction is subjected to a second silica gel column chromatography separation, and a second dichloromethane-methanol system is used for a second elution. The obtained silica gel column chromatography eluate is subjected to a first dextran gel column chromatography, and methanol is used for elution. According to the order of component elution, three fractions Fr.2.1.1 to Fr.2.1.3 are obtained; Sequentially subjecting the Fr.2.1.3 fraction to a second dextran gel column chromatography separation, a medium pressure preparative chromatography column separation, and a third silica gel column chromatography separation to obtain a phenolic compound having a structure represented by Formula I or Formula II; During the second dextran gel column chromatography separation, the eluent is a mixture of dichloromethane and methanol; When separating by medium-pressure preparative chromatographic column, the eluent is methanol-water solution; During the third silica gel column chromatography, the eluent was dichloromethane-acetone system.

3. The preparation method according to claim 2, characterized in that: The first elution is a gradient elution using a first dichloromethane-methanol system in which the volume ratio of dichloromethane to methanol is changed from 50:1 to 10:

1.

4. The preparation method according to claim 2, characterized in that: The second elution is a gradient elution using a second dichloromethane-methanol system in which the volume ratio of dichloromethane to methanol changes from 20:1 to 1:

1.

5. The preparation method according to claim 2, characterized in that: The volume ratio of dichloromethane to methanol in the mixed liquid of dichloromethane and methanol is 1:

1.

6. The preparation method according to claim 2, characterized in that: The volume ratio of dichloromethane to acetone in the dichloromethane-acetone system is 15:

1.

7. The preparation method according to claim 2, characterized in that: The volume concentration of the methanol aqueous solution is 30%.

8. The preparation method according to claim 2, characterized in that: The weak acid aqueous solution includes a 3 wt. % hydrochloric acid aqueous solution or a 3 wt. % acetic acid aqueous solution.

9. The preparation method according to claim 2, characterized in that: The weak alkali aqueous solution includes a 3 wt. % sodium hydroxide aqueous solution.

10. Use of the phenolic compound according to claim 1 or the phenolic compound prepared by the preparation method according to any one of claims 2 to 9 in the preparation of a drug for treating perimenopausal depression.

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