Ploroglucinol compounds Rxs-200 and Rxs-290 and pharmaceutical composition thereof

Abstract

The invention provides polycyclic polyisopentenyl acyl phloroglucinol derivatives Rxs-200 (compound 1) and Rxs-290 (2), a pharmaceutical composition containing the polycyclic polyisopentenyl acyl phloroglucinol derivatives, a preparation method of the polycyclic polyisopentenyl acyl phloroglucinol derivatives, and application of the polycyclic polyisopentenyl acyl phloroglucinol derivatives in pharmacy, and belongs to the technical field of medicines. The compounds 1 and 2 provided by the invention have obvious activity of inhibiting adipocyte differentiation, can be used for preparing medicines for treating obesity diseases and reducing lipid, and can be used for preparing medicines for treating obesity and / or metabolic diseases related to hyperlipidemia.

Description

Technical Field

[0001] The present invention belongs to the technical field of pharmaceuticals, and particularly relates to polycyclic polyprenylated acylphloroglucinol derivatives Rxs-200 (Compound 1) and Rxs-290 (Compound 2) isolated from Hypericum attenuatum, a pharmaceutical composition containing the same, a preparation method thereof, and their applications in the preparation of drugs for treating obesity and hyperlipidemia. Background Art

[0002] Obesity and hyperlipidemia are the most common lipid metabolism disorders in today's society. In addition to exogenous excessive intake of high-energy glycolipid components and low energy metabolism, the main pathophysiological forms of obesity are excessive white adipocytes and / or excessive lipid accumulation in adipocytes in the body. Obesity and hyperlipidemia are one of the inducing causes of many metabolic diseases (including type 2 diabetes, atherosclerosis, etc.). Obesity is accompanied by many complications, including type 2 diabetes, osteoarthritis, obstructive sleep apnea, cardiovascular diseases and various cancers. A large number of research data show that hyperlipidemia can directly harm human health and is an important risk factor for diseases such as stroke, coronary heart disease, and myocardial infarction. In addition, hyperlipidemia and obesity can also lead to fatty liver, cirrhosis, pancreatitis, and hyperuricemia.

[0003] Clinically used drugs for treating obesity and hyperlipidemia face problems such as many adverse reactions and obvious side effects. Treatment methods for obesity include control of daily diet, intervention in daily lifestyle, drug treatment, and weight loss surgery. Currently, the main drug used to improve obesity is orlistat, and its main side effect is flatulence, but other side effects such as nephrotoxicity, hepatotoxicity, kidney stones, and pancreatitis have also been reported. The mainstream drugs used clinically for lowering blood lipids are statins and niacin; they may affect the blood sugar, renal function, and liver function of patients, etc.

[0004] There are about 400 species of Hypericum in the world. There are about 55 species and 8 subspecies in China, which are distributed throughout the country, mainly in the southwestern region. Many plants of this genus have a long and extensive medicinal history among the people at home and abroad, such as Hypericum sampsonii, H. japonicum, etc. In recent years, it has been found that plants of this genus have activities such as anti-tumor, anti-viral, and antibacterial. Hypericum perforatum L. is a representative plant of this genus. It was used to treat central nervous system diseases in ancient Greece and has been used to treat depression in Germany for hundreds of years. Currently, various types of active ingredients have been discovered in Hypericum plants. Among them, prenylated acylphloroglucinols are their characteristic chemical components. Due to their novel, complex, variable, unique chemical structures and extensive biological activities, they have attracted extensive attention from scientists in the fields of pharmacy, chemistry, biology, etc. around the world.

[0005] Hypericum pseudohenryi, a shrub, is produced in western and southwestern Sichuan, northwestern and northeastern Yunnan. Its roots are used for acute and chronic icteric hepatitis, urinary tract infection stones, rheumatic pain, and traumatic injuries; its leaves are used externally for snake bites and knife and gunshot wounds. Literature reports that the chemical constituents of this plant are mainly prenylated acylphloroglucinols and flavonoid chemical constituents. However, so far, there have been no reports on polycyclic polyprenylated acylphloroglucinol derivatives Rxs-200 (Compound 1) and Rxs-290 (Compound 2) and their activities in the existing technologies. Summary of the Invention

[0006] The object of the present invention is to provide polycyclic polyprenylated acylphloroglucinol derivatives Rxs-200 (Compound 1) and Rxs-290 (Compound 2) isolated from Hypericum pseudohenryi (H. pseudohenryi), a pharmaceutical composition containing the same, a preparation method thereof, and their applications in the preparation of drugs for treating obesity diseases and lipid-lowering drugs, aiming at the above deficiencies existing in the prior art.

[0007] In order to achieve the above object of the present invention, the present invention provides the following technical solutions:

[0008] The polycyclic polyprenylated acylphloroglucinol compounds 1 and 2 shown in the following structural formula:

[0009]

[0010] The present invention provides a preparation method of the above-mentioned compounds 1 and 2, including the following steps: Take the aerial parts of Hypericum pseudohenryi, crush them, soak and extract them three times with methanol at room temperature for two days each time, combine the extraction solutions, and obtain an extract after concentration under reduced pressure. The extract is mixed with silica gel of 100 - 200 meshes, and chloroform is used as the eluent. After passing through a normal-phase silica gel column, the chloroform part is obtained. The obtained chloroform fraction is then mixed with silica gel of 100 - 200 meshes, and gradient elution is carried out with petroleum ether - ethyl acetate (100:1 - 0:1). During the elution process, a thin-layer silica gel plate is used for detection, and after combining the same parts, three fractions (Fr.A–C) are obtained. Fr.A is subjected to MCI column chromatography, and gradient elution is carried out with methanol–water (80:20 - 100:0) to obtain nine fractions (Fr.A1–A9). Compound 1 is obtained after Fr.A3 is subjected to preparative HPLC. Fr.A7 is divided into ten segments (Fr.A7.1–7.10) after passing through an RP-18 column, and Compound 2 is obtained after Fr.A7.8 passes through a normal-phase silica gel column and semi-preparative HPLC.

[0011] The present invention provides a pharmaceutical composition having significant adipocyte differentiation inhibitory activity, which is composed of an active ingredient and a pharmaceutically acceptable excipient. The active ingredient is compound 1 and 2 described in the above technical solution or compound 1 and 2 prepared by the preparation method described in the above technical solution.

[0012] The preparation method of the pharmaceutical composition includes the following steps: First, use the preparation method of the polycyclic polyprenyl acylphloroglucinol derivatives 1 and 2 to obtain polycyclic polyprenyl acylphloroglucinol derivatives 1 and / or 2, and then add pharmaceutically acceptable excipients.

[0013] Use of the polycyclic polyprenyl acylphloroglucinol derivatives 1 and / or 2 or the pharmaceutical composition in the preparation of a drug for treating obesity diseases and / or in the preparation of a lipid-lowering drug.

[0014] Use of the polycyclic polyprenyl acylphloroglucinol derivatives 1 and / or 2 or the pharmaceutical composition in the preparation of a drug for treating metabolic diseases related to obesity and / or hyperlipidemia.

[0015] Use of the polycyclic polyprenyl acylphloroglucinol derivatives 1 and / or 2 or the pharmaceutical composition in the preparation of an adipocyte differentiation inhibitor.

[0016] An adipocyte differentiation inhibitor, which uses polycyclic polyprenyl acylphloroglucinol derivatives 1 and 2 as active ingredients.

[0017] Use of a combination of the polycyclic polyprenyl acylphloroglucinol derivatives 1 and / or 2 or a pharmaceutical composition containing the same as an active ingredient and other drugs in the preparation of a drug for treating metabolic diseases related to obesity and / or hyperlipidemia, in the preparation of a drug for treating obesity diseases and / or in the preparation of a lipid-lowering drug.

[0018] In the present invention, when the pharmaceutical composition is used for preparing a drug, the content of the composition in the drug is preferably 0.1-99%; in the pharmaceutical composition, the content of at least one of the polycyclic polyprenyl acylphloroglucinol derivatives 1 and 2 in the pharmaceutical composition is preferably 0.5-90%. The pharmaceutical composition of the present invention is preferably used in the form of a dosage per unit body weight.

[0019] In the present invention, preferably, the pharmaceutically acceptable excipients include one or several of a drug carrier, a surfactant, a buffering substance, a disintegrant, a binder, a filler, a lubricant, an excipient, a solubilizer, a flavoring agent, and a coloring agent.

[0020] Preferably, the prepared drug is preferably administered by two forms: injection (intravenous injection, intramuscular injection) and oral administration. The dosage forms of the drug for treating obesity diseases and reducing lipid include tablets, capsules, granules, infusion, pills, oral liquid preparations, injections or lyophilized powder injections.

[0021] Compared with the prior art, the present invention has the following advantages:

[0022] 1. The present invention provides new polycyclic polyprenyl acylphloroglucinol compounds 1 and 2.

[0023] 2. The present invention provides a new preparation method for preparing new compounds 1 and 2. After extraction with methanol from Hypericum attenuatum, the target compounds with significant activity of inhibiting adipocyte differentiation can be obtained by using MCI column separation, normal-phase and reverse-phase column chromatography and high performance liquid chromatography separation techniques. The raw materials are easy to obtain and the operation is convenient, which is suitable for industrial production.

[0024] 3. The inhibitory activities of compounds 1 and 2 provided by the present invention on adipocytes enable the pharmaceutical composition with the compound as the active ingredient to be used for preparing drugs for treating obesity diseases and reducing lipid, and can be used for preparing adipocyte differentiation inhibitors, which has clinical application value and provides new drugs with good pharmacological effects for new drugs for treating obesity diseases and reducing lipid and adipocyte differentiation inhibitors.

[0025] 4. Compound 1 can significantly reduce the blood lipid level of obese mice, which is beneficial to improving lipid metabolism. Compared with the MDI group, compound 2 can significantly reduce the intracellular lipid accumulation, and shows a dose-dependent relationship. Compounds 1 and 2 have significant activities of inhibiting adipocyte differentiation, have significant activities of improving white fat differentiation and excessive lipid accumulation, and can be used for preparing drugs for improving obesity and related metabolic diseases of reducing lipid, and for preparing adipocyte differentiation inhibitors. Description of the Drawings

[0026] Figure 1 is the 1H NMR spectrum (600 MHz, CDCl3) of compound 1;

[0027] Figure 2 is the 13C NMR spectrum (150 MHz, CDCl3) of compound 1;

[0028] Figure 3 is the high-resolution mass spectrum of compound 1;

[0029] Figure 4 is the 1H NMR spectrum (600 MHz, CDCl3) of the methylation product a of the compound;

[0030] Figure 5 is the 13C NMR spectrum (150 MHz, CDCl3) of the methylation product a of the compound;

[0031] Figure 6 High-resolution mass spectrum of methylation product a of the compound;

[0032] Figure 7 1H NMR spectrum (600 MHz, CDCl3) of methylation product b of the compound;

[0033] Figure 8 13C NMR spectrum (150 MHz, CDCl3) of methylation product b of the compound;

[0034] Figure 9 High-resolution mass spectrum of methylation product b of the compound;

[0035] Figure 10 1H NMR spectrum (600 MHz, CDCl3) of compound 1;

[0036] Figure 11 13C NMR spectrum (150 MHz, CDCl3) of compound 1;

[0037] Figure 12 High-resolution mass spectrum of compound 1;

[0038] Figure 13 Schematic diagrams of the structures of compounds 1 and 2;

[0039] Figure 14 Methylation reaction of compound 1;

[0040] Figure 15 Compound 1 dose-dependently inhibits the differentiation and maturation of white adipocytes; (A) Representative images of adipocytes treated with different concentrations of 1 and stained with Oil Red O. (B) Quantitative results of Oil Red O staining. (C) IC 50 curve. (##p < 0.01 compared with the undifferentiated group (UND); *p < 0.05, **p < 0.01 compared with the MDI group; ns no significant difference, compared with the MDI group).

[0041] Figure 16Effect of compound 1 on body weight of obese mice. Obese C57BL / 6J male mice induced by high-fat diet (HFD) were treated with vehicle (1.5% DMSO), metformin (200 mg / kg / day), 1 (5 mg / kg / day or 20 mg / kg / day) for 9 weeks. The body weight of mice was monitored once a week. ND: normal diet, HFD: high-fat diet, Met: HFD + metformin (200 mg / kg), HFD+L: HFD + 1 (5 mg / kg), HFD+H: HFD + 1 (20 mg / kg). All values are expressed as mean ± standard deviation (n = 6 - 10). #P < 0.0001 compared with ND mice; *P < 0.05, **P < 0.05, ***P < 0.05, ****P < 0.05 compared with HFD mice.

[0042] Figure 17 Effect of compound 1 on blood lipid levels in obese mice; (A - D) Serum TC, TG, HDL-C and LDL-C levels after 9 weeks of administration; TC (total cholesterol), TG (triglyceride), HDL-C (high-density lipoprotein cholesterol) and LDL-C (low-density lipoprotein cholesterol); all values are expressed as mean ± standard deviation (n = 6 - 10). #P < 0.05, #P < 0.0001 compared with ND mice; *P < 0.05, **P < 0.05, ***P < 0.05, ****P < 0.05 compared with HFD mice. ns indicates no significance.

[0043] Figure 18 Compound 2 dose-dependently inhibits the differentiation and maturation of white adipocytes. (A) Representative images of adipocytes treated with different concentrations of 2 and stained with Oil Red O. (B) Quantitative results of Oil Red O staining. (C) IC 50 curve. (#p < 0.05 compared with the undifferentiated group (UND); *p < 0.05 compared with the MDI group; ns no significant difference, compared with the MDI group). Detailed implementation mode

[0044] The present invention provides polycyclic polyprenyl acylphloroglucinol derivatives Rxs-200 (compound 1) and Rxs-290 (compound 2), and the structural formulas are as Figure 13 shown.

[0045] The present invention provides a method for preparing the compounds 1 and 2 described in the above technical solution, comprising the following steps: taking the aerial part of Hypericum attenuatum, pulverizing it, soaking and extracting it three times with methanol at room temperature for two days each time, combining the extraction solutions, and obtaining an extract after concentration under reduced pressure. The extract is mixed with silica gel of 100 - 200 mesh, using chloroform as the eluent, and obtaining the chloroform fraction after passing through a normal-phase silica gel column. The obtained chloroform fraction is then mixed with silica gel of 100 - 200 mesh, and gradient elution is carried out with petroleum ether - ethyl acetate (100:1 - 0:1). During the elution process, thin-layer silica gel plates are used for detection, and three fractions (Fr.A - C) are obtained after combining the same parts. Fr.A is subjected to MCI column chromatography, and gradient elution is carried out with methanol - water (80:20 - 100:0) to obtain nine fractions (Fr.A1 - A9). Compound 1 is obtained after preparative HPLC of Fr.A3. Fr.A7 is divided into ten fractions (Fr.A7.1 - 7.10) after passing through an RP-18 column, and compound 2 is obtained after passing through a normal-phase silica gel column and semi-preparative HPLC of Fr.A7.8.

[0046] In the present invention, during the above silica gel column chromatography and MCI column separation processes, the volume of the eluent used has no special limitation and can be selected according to actual needs.

[0047] In the present invention, during the processes of performing silica gel column chromatography and separating through an MCI column, it is preferred to use thin-layer chromatography to view and fractionally collect to obtain multiple corresponding components (i.e., Fr.A - C components, Fr.A1 - A9 components).

[0048] The present invention provides a pharmaceutical composition having the activity of inhibiting adipocyte differentiation, which is composed of an active ingredient and a pharmaceutically acceptable excipient, and the active ingredient is at least one of the compounds 1 and 2 described in the above technical solution or the compounds obtained by the preparation method described in the above technical solution.

[0049] The present invention has no special limitation on the pharmaceutically acceptable excipient. Specifically, the pharmaceutically acceptable excipient preferably includes one or several of a drug carrier, a surfactant, a buffering substance, a disintegrant, a binder, a filler, a lubricant, an excipient, a solubilizer, a flavoring agent, and a coloring agent. The present invention has no special limitation on the specific types of the above excipients and can be selected according to actual needs.

[0050] The present invention provides the application of the pharmaceutical composition described in the above technical solution in the preparation of drugs for treating obesity diseases and reducing lipid levels.

[0051] The dosage form of the drug for treating obesity and reducing lipid is not particularly limited in the present invention, and can be selected according to actual needs. Specifically, it can be tablets, capsules, granules, instant granules, pills, oral liquid preparations, injections or lyophilized powder injections. The preparation method of the drug in different dosage forms is not particularly limited in the present invention, and the methods well-known to those skilled in the art can be used.

[0052] In the present invention, the dosage of the drug for treating obesity and reducing lipid will vary with the compound used, the mode of administration, the desired treatment and the indicated disorder. For example, for oral administration, the daily dosage of Compound 1 and 2 or their pharmaceutically acceptable salts can be in the range of 0.01 micrograms per kilogram of body weight (μg / kg) to 100 milligrams per kilogram of body weight (mg / kg).

[0053] The technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative work shall fall within the protection scope of the present invention.

[0054] Example 1

[0055] Preparation of polycyclic polyprenyl acylphloroglucinol derivatives Rxs-200 (Compound 1) and Rxs-290 (Compound 2):

[0056] The present invention provides a method for preparing polycyclic polyprenyl acylphloroglucinol derivatives Rxs-200 (Compound 1) and Rxs-290 (Compound 2), which includes the following steps: Take the aerial parts (75 kg) of Hypericum pseudohenryi, crush them, soak and extract three times with methanol at room temperature for two days each time, combine the extraction solutions, and concentrate under reduced pressure to obtain an extract. The extract is mixed with silica gel of 100-200 mesh, and chloroform is used as the eluent. After passing through a normal-phase silica gel column, the chloroform fraction is obtained. The obtained chloroform fraction is then mixed with silica gel of 100-200 mesh, and gradient elution is carried out with petroleum ether-ethyl acetate (100:1-0:1). Thin-layer silica gel plates are used for detection during the elution process, and three fractions (Fr.A–C) are obtained after combining the same parts. Fr.A is subjected to MCI column chromatography, and gradient elution is carried out with methanol–water (80:20-100:0) to obtain nine fractions (Fr.A1–A9). Compound 1 (1.0 g) is obtained after Fr.A3 is subjected to preparative HPLC. Fr.A7 is divided into ten segments (Fr.A7.1–7.10) after passing through an RP-18 column, and Compound 2 (16.3 mg) is obtained after Fr.A7.8 passes through a normal-phase silica gel column and semi-preparative HPLC.

[0057] Structure analysis of Compounds 1 and 2 (characterization diagrams are as Figures 1 to 14 shown) is as follows:

[0058] Compound 1, a colorless oil. From its high-resolution mass spectrum (m / z 535.3800 [M–H] – ) and carbon spectrum, its molecular formula can be deduced as C 35 H 52 O4, with an unsaturation degree of 10. The IR spectrum shows absorption bands of hydroxyl (3426 cm –1 ) and carbonyl (1729 cm –1 ). Whether it is the signals in the 1 H NMR spectrum or the 13 C NMR spectrum of this compound appear in pairs, which is caused by the common 1,3-diketone enol tautomerism in the structure of PPAP derivatives. Due to the severe overlap of the signals in the high-field regions of the hydrogen spectrum and carbon spectrum of this compound, many signals are difficult to assign in the HSQC spectrum, so it is very difficult to completely analyze the structure through such spectra. Therefore, the methylation derivatives (a and b) of 1 were prepared using TMS-CHN2, successfully blocking the tautomeric part in its structure and obtaining their single spectra ( Figure 11 ).

[0059] In the 1 H NMR spectrum of a, an isopropyl signal (δ H 2.75, sept, J = 6.8 Hz; 1.12, d, J = 6.8 Hz; 1.09, d, J = 6.8 Hz), four olefinic proton signals (δ H 4.77, t, J = 7.8 Hz; 5.03, t, J = 7.8 Hz; 5.00, t, J = 7.8 Hz; 5.08, t, J = 7.8 Hz), nine singlet methyl signals (δ H 0.50 - 1.70) and a methoxy signal (δ H 3.68, s) can be clearly observed. 13 The C C NMR spectrum shows 35 carbon resonance signals. Except for one isobutyryl group, four isopentenyl groups and one methoxy group, the remaining 11 carbon atoms are assigned to two methyl groups, one methylene group, one methine group and seven quaternary carbons (one non-conjugated carbonyl and one α,β-unsaturated ketone). Careful analysis of the 1D NMR data reveals that Compound a has the core skeleton signals of B-type bridged-ring PPAP: δ C 71.7 (C-1); 196.0 (C-2); 125.2 (C-3); 172.3 (C-4); 58.0 (C-5); 40.3 (C-6); 42.1 (C-7); 47.1 (C-8); 207.9 (C-9).

[0060] From the correlations of H2-17 with C-1 / C-2 / C-8 and H2-27 with C-6 / C-7 / C-8 in the HMBC spectrum, it can be seen that the two isopentenyl groups are respectively connected to C-1 and C-7 positions. Then, through 1 H- 1 the correlations of H2-34 / H2-35 / H-36 in the H- 1 H and 13 COSY spectrum, the correlations of Me-25 with C-34, H2-22 with C-4 / C-5 / C-6 / C-9, and 4-OCH3 with C-4 in the HMBC spectrum, it can be determined that the geranyl group and the methoxy group are respectively connected to C-5 and C-4 positions. From the NOE correlations of H-23 / H2-26, it can be seen that the C-23 / C-24 double bond is trans. Then, through the correlations of Me-33 / H-6b, Me-33 / H2-27, and H-6b / H-22a, the isopentenyl groups at C-1 and C-7 positions and the geranyl group at C-5 position are all in the β-orientation. Similarly, from the correlation of 2-OCH3 with C-2 in the HMBC spectrum of compound b, it can be seen that the methoxy group is connected to C-2 position. Based on the two derivatives of 1, its structure is finally determined, and its

[0061] Table 1 The 1 H-NMR and 13 C-NMR data of compound 1 (CDCl3)

[0062]

[0063]

[0064] Compound 2, colorless oil. According to the molecular ion peak given by the high-resolution mass spectrum and 13 the 35 C NMR spectrum, its molecular formula can be deduced as C 52 H H 1.16, d, J = 6.8 Hz; 1.17, d, J = 6.8 Hz; 2.20, sept, J = 6.8 Hz). Through 1 H- 1The correlations of H-12 / H-11 / H-13 in the H COSY spectrum and the correlations of Me-12 / Me-13 with C-10 in the HMBC spectrum can be further confirmed. According to the correlations of H-6b / Me-37, H-32 / Me-38, H-6a / H-33 and Me-20 / H-6a in the ROESY spectrum, the orientation of H-32 can be assigned as α, and the orientations of H-18 and H-33 can be assigned as β.

[0065] Table 2 of Compound 2 1 1H-NMR and 13 13C-NMR data (CDCl3)

[0066]

[0067]

[0068] Physical and chemical properties and structural data of Compound 1 and its methylation products a and b:

[0069] Compound 1: Colorless oil; (c 0.236, MeOH); UV (MeOH) λ max (logε) 274 (3.82), 255 (3.74), 238 (3.84), 220 (3.81), 195 (4.32) nm; IR (KBr) ν max 3426, 2974, 2932, 1729, 1660, 1626, 1450, 1378 cm -1 .

[0070] Compound a: Colorless oil; (c 0.140, MeOH); UV (MeOH) λ max (logε) 271 (3.81), 248 (3.77), 195 (4.32) nm; IR (KBr) ν max 3428, 2974, 2932, 1725, 1641, 1592, 1450, 1378 cm -1 .

[0071] Compound b: Colorless oil; (c 0.103, MeOH); UV (MeOH) λ max (logε) 271 (3.68), 248 (3.59), 195 (4.40) nm; IR (KBr) ν max 3435, 2969, 2923, 1728, 1648, 1582, 1445, 1378 cm -1 .

[0072] Physicochemical properties and structural data of Compound 2:

[0073] Compound 2: colorless oil; (c 0.157, MeOH); UV (MeOH) λ max (logε) 195 (4.29) nm; IR (KBr) ν max 2968, 2925, 1718, 1462, 1384 cm -1 .

[0074] Example 2:

[0075] The inhibitory activity of the polycyclic polyprenyl acylphloroglucinol derivative Rxs-200 (Compound 1) of the present invention against adipocytes, its therapeutic activity for obesity and lipid-lowering activity, and the experimental methods and results of the inhibitory activity of the polycyclic polyprenyl acylphloroglucinol derivative Rxs-290 (Compound 2) against adipocytes are as follows:

[0076] The activity of Compound 1 in the present invention to inhibit white adipocyte differentiation and intracellular lipid accumulation:

[0077] 1. Take Compound 1 and dissolve it in high-glucose DMEM medium containing 10% FBS to prepare drug solutions with concentrations of 0.25 μM, 0.5 μM, 1 μM, 2 μM, and 4 μM respectively.

[0078] 2. Inoculate 3T3-L1 preadipocytes with good growth status in a 12-well plate. After the cells are confluent, starve for 2 days, and then divide them into an undifferentiated blank group (UND), a model group (MDI), a positive drug group (LiCl: 20 mM), and groups given different concentrations of Compound 1.

[0079] 3. Stain the cells with Oil Red O, take pictures, and quantify the intracellular lipid content. The results show that a large amount of lipid accumulation occurs in normally differentiated white adipocytes (MDI); compared with the MDI group, Compound 1 can significantly reduce intracellular lipid accumulation, and shows a dose-dependent relationship. To better evaluate the effect of Compound 1 in inhibiting the differentiation and maturation of white adipocytes, the drug administration concentration was increased, and the S curve of Compound 1 inhibiting white adipocyte differentiation was fitted, in which the half-maximal inhibitory concentration (IC 50 ) was 1.644 μM ( Figure 15 ).

[0080] The activity of Compound 1 in the present invention to treat obesity and lipid-lowering activity:

[0081] The male C57BL / 6J mice used in the experiment were approximately 6 weeks old, purchased from the Experimental Animal Center of Yunnan University, and subsequent animal feeding and animal experiments were carried out at this center. The animals were housed in a temperature-controlled (23 ± 3 °C) and humidity-controlled (50 ± 10%) constant temperature and humidity ventilation system with a 12 h light / 12 h dark cycle. After 2 weeks of adaptive feeding after purchasing the mice, the 8-week-old mice were freely fed with a high-fat diet (composition: 60% fat, 20% carbohydrates, 20% protein) and drinking water containing 20% g / v fructose. After 6 weeks of feeding, there were significant changes in the body weight and serum biochemical indices of the mice fed with a high-fat diet (HFD) compared with those fed with a normal diet. All HFD-induced mice were randomly divided into 4 groups (n = 6 - 10), namely the HFD group, the HFD + L (1, 5 mg / kg / d) group, the HFD + H group (1, 20 mg / kg / d) group, and the HFD + Met (metformin, 200 mg / kg / d) group. The control group (n = 10) was fed with normal feed and water. The mice were gavaged (metformin) / intraperitoneally injected (1) with the solvent or drug every day from the 15th week to the 24th week. The body weight of the mice was monitored weekly during the treatment period.

[0082] The administration was carried out for 9 weeks. Weight monitoring found that after 9 weeks of administration, compared with the HFD group, the weight gain of the mice in the 1 treatment group was significantly inhibited ( Figure 16 ). The results indicate that compound 1 can effectively control the weight gain of obese mice induced by a high-fat diet and effectively inhibit the further development of obesity.

[0083] The blood lipid levels of the mice in each group were detected 2 h after administration. The results showed that compared with the HFD group and the metformin group, the administration of compound 1 could reduce the levels of TC, TG, HDL-C, and LDL-C in obese mice ( Figure 17 ). The results indicate that compound 1 can significantly reduce the blood lipid levels of obese mice, which is beneficial to improving lipid metabolism.

[0084] Compound 2 in the present invention inhibits the activity of white adipocyte differentiation and intracellular lipid accumulation:

[0085] 1. Take compound 2 and dissolve it in high-glucose DMEM medium containing 10% FBS to prepare drug solutions with concentrations of 0.78125 μM, 1.5625 μM, 3.125 μM, 6.25 μM, 12.5 μM, and 50 μM respectively.

[0086] 2. Inoculate the well-grown 3T3-L1 preadipocytes into a 12-well plate. After the cells are confluent, starve for 2 days and then divide them into an undifferentiated blank group (UND), a model group (MDI), a positive drug group (LiCl: 20 mM), and groups given different concentrations of compound 2.

[0087] 3. Stain the cells with Oil Red O, take pictures, and quantify the intracellular lipid content. The results show that a large amount of lipid accumulation is present in normally differentiated white adipocytes (MDI); compared with the MDI group, Compound 2 can significantly reduce intracellular lipid accumulation in a dose-dependent manner. Fit the S curve of Compound 2 inhibiting white adipocyte differentiation, and the half-maximal inhibitory concentration (IC 50 ) is 12.85 μM( Figure 18 ).

[0088] Formulation Examples 1-7:

[0089] In the following formulation examples, conventional reagents are selected and formulation preparation is carried out according to existing conventional methods. These formulation examples only demonstrate that at least one of the compounds 1 and 2 described in the present invention can be prepared into different formulations, and specific reagents and operations are not specifically limited:

[0090] 1. Dissolve at least one of Compounds 1 and 2 in DMSO, add water for injection according to conventional methods, filter it precisely, and fill and sterilize it to make an injection solution, and the concentration of the injection solution is 0.5-5 mg / mL.

[0091] 2. Dissolve at least one of Compounds 1 and 2 in DMSO, dissolve it in sterile water for injection, stir to dissolve it, filter it with a sterile suction filter funnel, then filter it precisely aseptically, dispense it into ampoules, freeze-dry it at low temperature and seal it aseptically to obtain a powder injection.

[0092] 3. Add at least one of Compounds 1-2 to an excipient in a mass ratio of 9:1 to make a powder.

[0093] 4. Add at least one of Compounds 1 and 2 to an excipient in a mass ratio of 5:1 to granulate and press tablets.

[0094] 5. Make an oral liquid according to the conventional preparation method of oral liquid with at least one of Compounds 1 and 2.

[0095] 6. Add at least one of Compounds 1 and 2 to an excipient in a mass ratio of 5:1 to make a capsule.

[0096] 7. Add at least one of Compounds 1 and 2 to an excipient in a mass ratio of 5:1 to make a granule.

[0097] The above are only the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and refinements can be made, and these improvements and refinements should also be regarded as the protection scope of the present invention.

Claims

1. The polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and 2 shown in the following structural formula, 2. The polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and 2 represented by the structural formula as claimed in claim 1, characterized in that: It is a compound having an isopentenyl heteroaromatized phloroglucinol core.

3. A method for preparing the polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and 2 represented by the structural formula according to claim 1 or 2, characterized in that: The method comprises the following steps: taking the above-ground part of the northern rice flower, crushing it, soaking it in methanol at room temperature for extraction three times, each time for two days, combining the extracts, and concentrating it under reduced pressure to obtain an extract, mixing the extract with 100-200 mesh silica gel, using chloroform as an eluent, passing through a forward silica gel column to obtain a chloroform part, mixing the obtained chloroform section with 100-200 mesh silica gel, gradient eluting with 100:1-0:1 petroleum ether-ethyl acetate, using a thin layer silica gel plate for detection during the elution process, and combining the phases. After the same part, three sections Fr.A–C were obtained; Fr.A was chromatographed on an MCI column with a gradient elution of methanol-water from 80:20 to 100:0 to obtain nine fractions Fr.A1–A9; Fr.A3 was subjected to preparative HPLC to obtain a polycyclic polyisopentenyl acyl pyrogallol derivative 1; Fr.A7 was divided into ten sections Fr.A7.1–7.10 after being subjected to an RP-18 column, and Fr.A7.8 was subjected to a normal silica gel column and semi-preparative HPLC to obtain compound 2.

4. A pharmaceutical composition, characterized in that It consists of active ingredients and pharmaceutically acceptable excipients, wherein the active ingredients are polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and / or 2 represented by the structural formula described in claim 1 or 2, or polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and / or 2 prepared by the preparation method described in claim 3.

5. A pharmaceutical composition for treating obesity and reducing lipids, characterized in that: It consists of active ingredients and pharmaceutically acceptable excipients, wherein the active ingredients are polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and / or 2 represented by the structural formula described in claim 1 or 2, or polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and / or 2 obtained by the preparation method described in claim 3.

6. The method for preparing the pharmaceutical composition according to claim 4 or 5, characterized in that: The method comprises the following steps: firstly obtaining the polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and / or 2 by the preparation method described in claim 3, and then adding pharmaceutically acceptable excipients.

7. Use of the polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and / or 2 represented by the structural formula according to claim 1 or 2 or the pharmaceutical composition according to any one of claims 4 or 5 in the preparation of drugs for treating obesity and / or in the preparation of lipid-lowering drugs.

8. Use of the polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and / or 2 represented by the structural formula according to claim 1 or 2 or the pharmaceutical composition according to any one of claims 4 or 5 in the preparation of drugs for treating metabolic diseases related to obesity and / or hyperlipidemia.

9. Use of the polycyclic polyisopentenyl acyl phloroglucinol derivative 1 and / or 2 represented by the structural formula according to claim 1 or 2 or the pharmaceutical composition according to any one of claims 4 or 5 in the preparation of an adipocyte differentiation inhibitor.

10. An adipocyte differentiation inhibitor, characterized in that The polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and 2 represented by the structural formula described in claim 1 or 2 are used as active ingredients.

11. Use of the polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and / or 2 represented by the structural formula of claim 1 or 2 or the polycyclic polyisopentenyl acyl phloroglucinol derivatives 1 and / or 2 prepared by the preparation method of claim 3 as active ingredients, or a pharmaceutical composition containing the same, in combination with other drugs in the preparation of drugs for treating obesity and / or metabolic diseases related to hyperlipidemia, in the preparation of drugs for treating obesity and / or in the preparation of lipid-lowering drugs.

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