Ploretin derivative and application thereof in medicine for treating non-alcoholic fatty liver disease

Abstract

The invention discloses a phloretin derivative and application thereof in a medicine for treating non-alcoholic fatty liver related to diabetes mellitus, relates to the technical field of medicines, and in particular relates to a compound shown in a formula (1) and activity of the compound for relieving the non-alcoholic fatty liver related to the diabetes mellitus metabolic syndrome. Pharmacological experiment results show that the compound shown in the formula (1) can significantly relieve hepatomegaly, reduce lipid droplets in liver tissues, recover the morphology of liver cells, reduce the content of total cholesterol, triglyceride and low-density lipoprotein in blood, reduce related inflammatory indexes and enhance antioxidant activity.

Description

Technical Field

[0001] The invention belongs to the field of medicine, and particularly relates to a phloretin derivative and its medical use. Background Art

[0002] Type 2 diabetes mellitus (T2DM) is a serious chronic metabolic disease with long-term hyperglycemia, characterized by insulin resistance and insufficient insulin secretion from pancreatic β cells leading to hyperglycemia. Patients with T2DM metabolic syndrome are at high risk of non-alcoholic fatty liver disease associated with diabetes metabolic syndrome. In addition to directly leading to decompensated cirrhosis, hepatocellular carcinoma and recurrence of transplanted liver, non-alcoholic fatty liver disease can also further participate in the onset of type 2 diabetes and atherosclerosis, forming a vicious cycle. Current treatment methods usually use hypoglycemic drugs to achieve the effect of reducing fat and weight, and control the further deterioration of fatty liver and diabetes. At present, some commonly used T2DM drugs (such as acarbose, metformin and thiazolidinediones) are used to treat type 2 diabetes, but these drugs have a wide range of side effects including hypoglycemia, stomach pain, diarrhea, flatulence, allergic reactions, etc.

[0003] Natural products usually have a wide range of biological activities due to their unique chemical structure. Among them, chalcone compounds are widely present in many plants, and studies have shown that chalcone compounds (including phloretin) have preventive and therapeutic effects on non-alcoholic fatty liver disease associated with type 2 diabetes metabolic syndrome. Its main pharmacological mechanism of action is related to its inhibition of lipid metabolism-related pathways and antioxidant activity. Summary of the invention

[0004] In view of the shortcomings of the prior art, the present invention aims to propose a new and more effective phloretin derivative for alleviating non-alcoholic fatty liver disease associated with diabetic metabolic syndrome, and further improve the effect by modifying the known compounds.

[0005] The first aspect of the present invention is to provide a compound, wherein the compound is a phloretin derivative represented by formula (1):

[0006]

[0007] Furthermore, in the compound, R is a C1-C10 saturated hydrocarbon group or a halogen;

[0008] Furthermore, the R is halogen, and further, the R is F.

[0009] The second aspect of the present invention is to provide a method for preparing the compound described in the first aspect, wherein the method is synthesized according to the following route:

[0010]

[0011] Furthermore, the specific steps of the method are:

[0012] 1) Using phloretin and 1-(fluorosulfonyl)-2,3-dimethyl-1H-imidazol-3-ium trifluoromethanesulfonate as raw materials, reacting at room temperature for 6 to 10 hours under the catalysis of triethylamine and acetonitrile to obtain a monosubstituted phloretin derivative;

[0013] 2) The product was then separated and purified by silica gel column chromatography using ethyl acetate / petroleum ether as a developing solvent.

[0014] Further, the molar ratio of phloretin to 1-(fluorosulfonyl)-2,3-dimethyl-1H-imidazol-3-ium trifluoromethanesulfonate is 1:1 to 1.5; preferably 1:1.3;

[0015] Furthermore, the product is obtained by rotary evaporation drying after the reaction in step 1);

[0016] Furthermore, in the step 2), the ratio of ethyl acetate to petroleum ether is 1:3-7 (v / v); preferably 1:5 (v / v).

[0017] The third aspect of the present invention is to provide a pharmaceutical composition for treating non-alcoholic fatty liver disease associated with diabetic metabolic syndrome, wherein the pharmaceutical composition comprises the compound described in the first aspect and a pharmaceutically acceptable carrier.

[0018] In a fourth aspect, the present invention further provides use of the compound described in the first aspect in the preparation of a drug for treating non-alcoholic fatty liver disease, preferably alcoholic fatty liver disease associated with diabetes.

[0019] On the other hand, the present invention provides the use of the above compounds in the preparation of drugs for treating diabetes-related fatty liver disease, preferably including the use of drugs for lowering blood sugar AST, ALT, TG, CHO, LDL, SOD, MDA and / or CAT, or increasing HDL levels. Preferably, the use of the above compounds in the preparation of drugs for increasing HDL levels in the blood.

[0020] Preferably, the above application includes reducing inflammatory damage in the blood, enhancing antioxidant levels, and / or reducing blood lipid levels simultaneously.

[0021] Preferably, the above application also includes application for alleviating inflammatory damage in liver and muscle.

[0022] Preferably, the above application is applied to diabetic patients or mammals, preferably model mice, and the model mice are db / db mice.

[0023] Compared with the prior art, the present invention has the following advantages:

[0024] The present invention obtains compound (1) by modifying phloretin, which can significantly enhance the effect of relieving non-alcoholic fatty liver disease associated with diabetic metabolic syndrome and has extremely high potential for resisting non-alcoholic fatty liver disease. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 Effects of compounds on liver organ indexes. Note: * represents: P<0.05, *** represents P<0.001.

[0026] Figure 2 Effects of compounds on HE staining of liver.

[0027] Figure 3 Effects of compounds on blood biochemical indices. Note: * represents: P<0.05, ** represents P<0.01, *** represents P<0.001, **** represents P<0.0001.

[0028] Figure 4 Effects of compounds on antioxidant indexes in muscle and liver. Note: * represents: P<0.05, ** represents P<0.01, *** represents P<0.001, **** represents P<0.0001. DETAILED DESCRIPTION

[0029] The following is a further description of the concept of the present invention and the technical effects produced in conjunction with specific embodiments, so as to fully understand the purpose, features and effects of the present invention. The methods are conventional methods unless otherwise specified. The materials can be obtained from public commercial channels unless otherwise specified. The illustrative embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute improper limitations of the present invention. It should be noted that the embodiments of the present invention and the features in the embodiments can be combined with each other without conflict.

[0030] Example 1: Preparation of phloretin derivatives

[0031] The compound shown in formula I was synthesized according to the following process:

[0032]

[0033] The specific preparation process is as follows: add 27.4 mg (0.1 mmol) of phloretin and 42 mg (0.13 mmol) of 1-(fluorosulfonyl)-2,3-dimethyl-1H-imidazol-3-ium trifluoromethanesulfonate to a 10 ml round-bottom flask, use a pipette to draw 1 ml of acetonitrile into the flask, then add 22 μl (0.16 mmol) of triethylamine, and stir at room temperature for 8 hours. After the reaction is completed, the reaction solution is dried and separated and purified by silica gel column chromatography using ethyl acetate / petroleum ether (1:5, v / v) as the developing solvent.

[0034] The physicochemical data of the obtained compound (1) and 1 H-NMR, 13 C-NMR, 19 The F NMR data are as follows:

[0035] Compound (1) 1 H NMR (500MHz, CDCl3) δ10.25 (s, 1H), 7.27 (d, J = 8.9Hz, 1H), 7.22–7.16 (m, 1H), 6. 34(s,1H),3.39(t,J=7.5Hz,1H),2.99(t,J=7.5Hz,1H),1.99(d,J=33.1Hz,1H); 13 C NMR (126MHz, CDCl3) δ205.05,163.03,153.85,148.46,142.06,130.44,120.86,109.64,100.80,45.93,29.26; 19 F NMR (471MHz,CDCl3)δ39.64,37.35; Molecular formula: C 15 H 12 F 2 O 9 S 2 ; Molecular weight: 438.3688.

[0036] Example 2: Effect of compound (1) on liver organ index of non-alcoholic fatty liver disease associated with diabetic metabolic syndrome

[0037] We used mouse gavage experiments to evaluate the effects of compounds on non-alcoholic fatty liver disease associated with diabetic metabolic syndrome in vivo. 5-6 week-old db / db mice and C57 / BL mice were used as control groups. They were adapted to the experimental environment for one week before the formal experiment. Animal experiments were conducted in a specific sterile environment (SPF). This study was approved by the Laboratory Animal Management and Welfare Ethics Committee of Beijing Huayuan Times Technology Co., Ltd. (Ethics Review Approval No.: HYSD2023-04). The db / db mice were randomly divided into a diabetes model control group, an acarbose group, and a compound (1) group, with 8 mice in each group. Compound (1) and acarbose were dissolved in saline containing 10% DMSO. The compound (1) group was gavaged at a dose of 20 mg / kg and was labeled as the compound (1) group. The acarbose group was gavaged at a dose of 20 mg / kg and was labeled as the acarbose group. The normal control group and the diabetes model group received equal amounts of saline gavage, respectively; each was labeled as the control group and the model group. During the four-week oral administration period, all mice had free access to food and water, and their body weight and water intake were recorded.

[0038] After administration, mice were anesthetized for orbital blood collection. The collected blood was centrifuged at a low temperature of 3500r / min, and the serum was separated after 10 minutes and dispensed into cryopreservation tubes. After blood collection, the mice were euthanized by cervical dislocation and dissected to obtain liver and muscle tissue samples. Any blood on the tissue was washed with saline, and then filter paper was used to remove excess water on the surface. The organ index was calculated based on this. A portion of the liver tissue sample was taken and fixed in a sample tube containing 4% paraformaldehyde, while the other portion was stored in a cryotube.

[0039] The results are as follows Figure 1 As shown, the liver volume of mice in the model group increased significantly, while the liver volume of mice in the compound (1) group tended to decrease (P<0.05). There was no significant difference in the positive drug group.

[0040] Example 3: Effects of Compounds on HE Staining of Liver

[0041] HE staining: The tissue is trimmed, cut, placed in a dehydration box, and dehydrated in sequence with 75% to 100% alcohol. Subsequently, the tissue is immersed in wax and processed using an embedding machine. After dressing, it is sliced ​​using a paraffin slicer. The resulting slices are mounted on slides and dried for later use. These slices are HE stained according to the following main steps: (1) treated with methanol and xylene, followed by rinsing with distilled water; (2) continuous staining with Harry's hematoxylin, differentiation with 1% hydrochloric acid ethanol, blue restoration with 0.6% ammonia water, and regular washing with distilled water; (3) eosin staining lasts for about 3-5 minutes; (4) dehydration and transparency are achieved through different concentrations of alcohol and xylene solutions before sealing with glue; (5) observation under a microscope with the aid of photography.

[0042] The results are as follows Figure 2 As shown, HE staining histological examination showed significant differences between the model group and the other treatment groups. The hepatocytes in the model group were severely swollen, and there were obvious lipid droplets in the cytoplasm. The nucleus was compressed and deformed to one side, the cells were arranged in disorder, and the hepatic sinusoids were enlarged. Compared with the model group, the hepatocytes treated with acarbose showed improved deformability, fewer lipid droplet vacuoles, regular cell arrangement, and reduced fatty degeneration. Similarly, compared with the acarbose-treated group, the cell types in the compound (1) group were arranged more orderly and had fewer vacuoles.

[0043] Example 4 Effects of Compounds on Blood Biochemical Indices

[0044] Blood biochemical indexes: The kits made in Nanjing were used to quantitatively measure the levels of serum triglyceride (TG), cholesterol (TC), high-density lipoprotein (HDL-C), low-density lipoprotein (LDL-C), alanine aminotransferase (ALT), aspartate aminotransferase (AST), glycosylated serum protein (GSP), lipopolysaccharide (LPS), and malondialdehyde (MDA) in each group of rats. At the same time, catalase activity (CAT), reduced glutathione level (GSH) and superoxide dismutase activity (T-SOD) were measured. For details, please refer to the instructions of the kit.

[0045] The results are as follows Figure 3 As shown, in order to further study the effect of compound (1) administration on the liver of diabetic mice, we analyzed the inflammatory enzymes and fat content in the blood. The results showed that compared with the model group, the ALT, AST, TG, CHO, LDL, and MDA levels of mice in the compound (1) and acarbose groups were significantly reduced. Compared with the model group, compound (1) reduced the levels of ALT, AST, TG, CHO, LDL and MDA by 16.9%, 31.5%, 17.4%, 12.7%, 21.0% and 23.1%, respectively (as shown in Table 1 below). There was no significant difference between compound (1) and acarbose in these indicators. At the same time, the levels of HDL, T-SOD, GSH and CAT in mice in the compound (1) group increased by 22.1%, 20.1%, 29.8% and 32.3%, respectively. Similarly, there was no significant difference between compound (1) and acarbose in these indicators. The above results indicate that both the compound (1) group and the acarbose group can reduce inflammatory damage in the blood of diabetic mice, enhance antioxidant levels, and reduce blood lipid levels.

[0046] Table 1

[0047]

[0048] Example 5 Effects of Compounds on Muscle and Liver Antioxidant Indexes

[0049] Also according to the instructions of Nanjing Jiancheng kit, the levels of TG, TC, MDA, CAT, GSH and SOD in mouse liver and muscle were quantified.

[0050] The results are as follows Figure 4As shown, compared with the model group, compound (1) reduced liver MDA and muscle MDA levels by 23.05% and 40.45%, respectively. At the same time, liver T-SOD, liver GSH, liver CAT, muscle T-SOD, muscle GSH and muscle CAT levels increased by 57.2%, 34.75%, 30.58%, 47.52%, 25.84% and 28.85%, respectively (Table 2). There was no significant difference between compound (1) and acarbose in these indicators. The above results show that both the compound (1) group and the acarbose group can reduce inflammatory damage in the liver and muscles of diabetic mice.

[0051] Table 2

[0052]

[0053]

[0054] The embodiments described above are only some embodiments of the present invention, rather than all embodiments. Based on the embodiments of the present invention, other embodiments obtained by those skilled in the art without creative work shall all fall within the scope of protection of the present invention.

Claims

1. A compound, wherein the compound is a phloretin derivative represented by formula (1): It is characterized in that The R is F.

2. A method for preparing the compound according to claim 1, characterized in that: The method described is synthesized according to the following route:

3. The method according to claim 2, characterized in that The specific steps of the method are: 1) Using phloretin and 1-(fluorosulfonyl)-2,3-dimethyl-1H-imidazol-3-ium trifluoromethanesulfonate as raw materials, reacting at room temperature for 6 to 10 hours under the catalysis of triethylamine and acetonitrile to obtain a monosubstituted phloretin derivative; 2) The product was then separated and purified by silica gel column chromatography using ethyl acetate / petroleum ether as a developing solvent.

4. The method according to claim 3, characterized in that The molar ratio of phloretin to 1-(fluorosulfonyl)-2,3-dimethyl-1H-imidazol-3-ium trifluoromethanesulfonate is 1:1 to 1.

5.

5. A pharmaceutical composition for treating non-alcoholic fatty liver disease associated with diabetic metabolic syndrome, characterized in that: The pharmaceutical composition comprises the compound according to claim 1 and a pharmaceutically acceptable carrier.

6. Use of the compound according to claim 1 or the compound prepared by the method according to any one of claims 2 to 4 for non-therapeutic purposes in vitro.

7. Use of the compound according to claim 1 or the compound prepared by the method according to any one of claims 2 to 4 in the preparation of a drug for treating non-alcoholic fatty liver disease associated with diabetic metabolic syndrome.

8. Use of the compound according to claim 1 or the compound prepared by the method according to any one of claims 2 to 4 in the preparation of a drug for lowering blood sugar AST, ALT, TG, CHO, LDL, SOD, MDA and / or CAT, or increasing HDL levels.

9. Use of the compound according to claim 1 or the compound prepared by the method according to any one of claims 2 to 4 in the preparation of a drug for increasing HDL levels.

10. The use according to any one of claims 7 to 9, characterized in that: The uses include use in reducing inflammatory damage in the liver and muscles of diabetic patients.

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