阅读说明：本技术 二苯并呋喃类化合物的降脂应用 (Lipid-lowering application of dibenzofuran compound ) 是由 肖朝江 姜北 陈威 杨丽 白惠荣 王影 郎利娟 董相 于 2021-10-25 设计创作，主要内容包括：本发明涉及二苯并呋喃类化合物在制备降脂产品中的应用,属于中药、天然药物制药领域。以本发明二苯并呋喃类化合物为活性成分的降脂药物。本发明拓展了二苯并呋喃类化合物的药用价值。(The invention relates to an application of dibenzofuran compounds in preparation of lipid-lowering products, belonging to the field of pharmacy of traditional Chinese medicines and natural medicines. The invention relates to a lipid-lowering drug which takes dibenzofuran compounds as active ingredients. The invention expands the medicinal value of the dibenzofuran compound.)

1. The dibenzofuran compound with the following structural formula,

。

2. a pharmaceutical composition comprising a therapeutically effective amount of a compound of any of claim 1 and a pharmaceutically acceptable carrier.

3. Lipid lowering agents containing a compound according to claim 1 and customary auxiliaries.

4. Use of a compound according to claim 1 for the preparation of a lipid lowering agent.

5. Use of a compound according to claim 1 for the manufacture of a medicament for the treatment of hyperlipidemia and obesity.

The technical field is as follows:

the invention belongs to the field of traditional Chinese medicine and natural medicine pharmacy, and particularly relates to a pharmaceutical composition taking dibenzofuran compounds as active ingredients and application thereof in lipid lowering agents.

Technical background:

dibenzofurans are mainly distributed in lichen plants, Rosaceae and Myrtaceae. The dibenzofuran compounds are reported to have anticancer function in literatureBiological activities (Love B E. Isolation and synthesis of polyoxygenated dibenzofurans stress biological activity) such as antibacterial, antiallergic, antiinflammatory, antioxidant, etc.Eur J Med Chem, 2015, 97: 377-387). So far, no reports of the lipid-lowering activity of the compounds are found.

The invention content is as follows:

the invention aims to provide a dibenzofuran compound, a pharmaceutical composition taking the dibenzofuran compound as an active ingredient, a preparation method of the dibenzofuran compound and an application of the dibenzofuran compound in preparing a lipid-lowering agent.

The above object of the present invention is achieved by the following technical solutions:

dibenzofuran compounds 1-9 represented by the following structures,

。

a method for preparing compounds 1-9 comprises collecting fruit or whole plant of Crataegus of Rosaceae, extracting with organic solvent chloroform or ethyl acetate or acetone or methanol or ethanol or water by cold soaking or hot reflux extraction, or extracting with the above organic solvent or water by cold soaking or reflux extraction and then extracting with ethyl acetate to obtain total extract, and performing repeated column chromatography to obtain compounds 1-9.

The process for the preparation of the compounds 1 to 9 according to the invention is more particularly carried out with:

a: extracting fruit or whole plant of Crataegus with acetone, methanol, ethanol, water, cold soaking or hot reflux to obtain total extract, extracting with ethyl acetate to obtain ethyl acetate extract, and repeatedly performing column chromatography to obtain compounds 1-9.

B: extracting coarse powder of fruit or whole plant of Crataegus with organic solvent (such as chloroform, methanol, ethanol, acetone, dichloromethane, etc.) by cold soaking or hot refluxing to obtain total extract, and repeatedly performing column chromatography to obtain compounds 1-9.

More specifically, the preparation method of the compounds 1-9 comprises the steps of drying fruits or whole plants of hawthorn plants in the shade, crushing the fruits or the whole plants to 30 meshes, leaching the dried fruits or the whole plants with 95% ethanol at room temperature for 3 times, each time for 24 hours, merging extracting solutions, concentrating the extracting solution under reduced pressure to obtain extract, suspending the extract with a proper amount of water, distributing the extract for several times with ethyl acetate to obtain an ethyl acetate extract, dissolving the extract with a proper amount of chloroform/acetone, mixing the extract with 80-100 meshes of silica gel, performing column chromatography with 200-300-mesh silica gel for coarse fractionation, performing gradient elution with 1: 0-0: 1 chloroform/acetone or 1: 0-0: 1 chloroform/methanol to obtain 8 main parts, performing silica gel column chromatography on the 1: 0 chlorine/propane part, the 9: 1 chlorine/propane part and the 8: 2 chlorine/propane part, performing gradient elution with 30: 1-1: 2 petroleum ether/acetone, obtaining 10 parts, and respectively carrying out repeated silica gel, RP-18 and Sephadex LH-20 column chromatography to respectively obtain compounds 1-9.

The lipid-lowering agent comprises any one of compounds 1-9 and conventional auxiliary agents.

A pharmaceutical composition comprising a therapeutically effective amount of a compound of any one of compounds 1-9 and a pharmaceutically acceptable carrier.

The application of any one of the compounds 1-9 in preparing lipid-lowering agents.

The application of any compound in preparing medicines for treating hyperlipidemia and obesity.

The invention relates to a pharmaceutical composition for hyperlipidemia and obesity, which comprises any one of compounds 1-9 and a pharmaceutically acceptable carrier.

The pharmaceutically acceptable carrier in the pharmaceutical composition of the invention refers to a conventional pharmaceutical carrier in the pharmaceutical field. The compounds of the present invention may be administered in the form of compositions by oral, nasal, rectal or parenteral administration to a patient in need of such treatment. For oral administration, it can be made into conventional solid preparations such as tablet, powder, granule, capsule, etc., and liquid preparations such as oil suspension, syrup, elixir, etc.; for parenteral administration, it can be prepared into a solution for injection, etc. Preferred forms are tablets, capsules and injections.

Various dosage forms of the pharmaceutical composition of the present invention can be prepared according to conventional production methods in the pharmaceutical field. For example, the active ingredient may be combined with one or more carriers and then formulated into the desired dosage form.

The pharmaceutical compositions of the present invention preferably contain 0.1% to 99.5% by weight of the active ingredient, most preferably 0.5% to 95% by weight of the active ingredient.

The amount of the compound of the present invention to be administered may vary depending on the route of administration, age, body weight of the patient, type and severity of the disease to be treated, etc., and the daily dose may be 0.01 to 10 mg/kg body weight, preferably 0.1 to 5 mg/kg body weight. One or more administrations may be carried out.

The compounds of the present invention show better lipid-lowering activity.

The invention screens the lipid-lowering activity of the compounds 1-9, and the compounds show better lipid-lowering activity. In lipid lowering activity applications, compounds 1-9 are applied to a substrate or a population in an amount in the range of 1-1000 μ M, preferably 10-200 μ M, optionally in combination with a carrier and/or medium.

The specific implementation mode is as follows:

the following examples are provided to further illustrate the essence of the present invention, which will enable a person skilled in the art to more fully understand the invention, but are not intended to limit the invention in any way.

Example 1:

extraction, isolation and purification of compounds 1-9 of the invention:

drying branches and leaves (10 kg) of Yunnan fructus crataegi in the shade, pulverizing to 30 mesh, extracting with 95% ethanol at room temperature for 3 times (50L each time for 24 hr), mixing extractive solutions, and concentrating under reduced pressure to obtain extract (1.5 kg). Dissolving the extract with chloroform/acetone, adsorbing with a proper amount of 80-100 mesh silica gel, performing column chromatography segmentation coarse separation with 3.0 kg of 200-mesh silica gel and 300-mesh silica gel, performing gradient elution with chloroform/acetone (1: 0-0: 1) to obtain 8 main parts, performing silica gel column chromatography on the 1: 0 chloroform part, the 9: 1 chlorine/propyl part and the 8: 2 chlorine/propyl part, and performing gradient elution with 30: 1-1: 2 petroleum ether/acetone to obtain 8 parts. Wherein the first elution part is subjected to repeated silica gel, RP-18 and Sephadex LH-20 column chromatography to obtain compounds 1-4. Performing silica gel, RP-18 (methanol-water) and Sephadex LH-20 (chloroform-methanol 1: 1) column chromatography repeatedly on the second and third elution parts to obtain compounds 5-9.

Example 2:

physical and spectral data for compounds 1-9 of the invention:

compound 1: pink crystals (chloroform).¹H-NMR (400 MHz, CDCl₃) δ: 7.84 (1H, dd, J = 7.5, 1.3 Hz, H-9), 7.54 (1H, brd, J = 8.2 Hz, H-6), 7.41 (1H, ddd, J = 8.2, 7.3, 1.3 Hz, H-7), 7.30 (1H, ddd, J = 7.5, 7.3, 0.8 Hz, H-8), 7.18 (1H, s, H-1), 5.74 (1H, s, 2-OH), 4.27 (3H, s, 4-OCH₃), 4.03 (3H, s, 3-OCH₃); ¹³C-NMR (100 MHz, CDCl₃) δ: 99.0 (d, C-1), 145.9 (s, C-2), 138.2 (s, C-3), 138.2 (s, C-4), 142.0 (s, C-4a), 156.6 (s, C-5a), 111.7 (d, C-6), 126.7 (d, C-7), 122.8 (d, C-8), 120.6 (d, C-9), 124.5 (s, C-9a), 120.8 (s, C-9b), 61.9 (q, 3-OCH₃), 61.1 (q, 4-OCH₃)。

Compound 2: a red solid.¹H-NMR (400 MHz, CDCl₃) δ: 8.07 (1H, dd, J = 7.5, 1.5 Hz, H-9), 7.54 (1H, brd, J = 8.2 Hz, H-6), 7.39 (1H, td-like, J = 7.8, 1.5 Hz, H-7), 7.32 (1H, td-like, J = 7.5, 0.9 Hz, H-8), 5.99 (1H, s, 1-OH), 4.11 (3H, s, 4-OCH₃), 4.03 (3H, s, 3-OCH₃), 3.97 (3H, s, 2-OCH₃); ¹³C-NMR (100 MHz, CDCl₃) δ: 139.6 (s, C-1), 135.6 (s, C-2), 145.2 (s, C-3), 132.1 (s, C-4), 145.2 (s, C-4a), 156.1 (s, C-5a), 111.3 (d, C-6), 126.1 (d, C-7), 123.1 (d, C-8), 122.6 (d, C-9), 123.7 (s, C-9a), 108.6 (s, C-9b), 62.0 (q, 2-OCH₃), 61.8 (q, 3-OCH₃), 61.8 (q, 4-OCH₃)。

Compound 3: a tan solid.¹H-NMR (400 MHz, CDCl₃) δ: 7.42 (1H, brd, J = 7.8 Hz, H-9), 7.23 (1H, dd, J = 8.0, 7.8 Hz, H-8), 7.15 (1H, s, H-1), 6.94 (7H, d, J = 8.0 Hz, H-7), 5.75 (1H, s, 2-OH), 4.28 (3H,s, 4-OCH₃), 4.05 (3H, s, 6-OCH₃), 4.03 (3H, s, 3-OCH₃); ¹³C-NMR (100 MHz, CDCl₃) δ: 98.9 (d, C-1), 146.0 (s, C-2), 138.2 (s, C-3), 138.4 (s, C-4), 142.1 (s, C-4a), 145.1 (s, C-5a), 145.7 (s, C-6), 109.2 (d, C-7), 123.5 (d, C-8), 112.7 (d, C-9), 126.1 (s, C-9a), 121.1 (s, C-9b), 61.9 (q, 3-OCH₃), 61.1 (q, 4-OCH₃), 56.3 (q, 6-OCH₃)。

Compound 4: a reddish brown oil.¹H-NMR (400 MHz, acetone-d ₆) δ: 7.81 (1H, s, 2-OH), 7.39 (1H, t, J = 8.4 Hz, H-7), 7.30 (1H, s, H-1), 7.20 (1H, dd, J = 8.4, 0.8 Hz, H-6), 6.90 (1H, dd, J = 8.4, 0.8 Hz, H-8), 4.18 (3H, s, 4-OCH₃), 4.05 (3H, s, 9-OCH₃), 3.92 (3H, s, 3-OCH₃); ¹³C-NMR (100 MHz, acetone-d ₆) δ: 103.5 (d, C-1), 148.9 (s, C-2), 140.4 (s, C-3), 139.9 (s, C-4), 142.5 (s, C-4a), 159.1 (s, C-5a), 105.6 (d, C-6), 129.1 (d, C-7), 105.5 (d, C-8), 157.3 (s, C-9), 115.0 (s, C-9a), 121.2 (s, C-9b), 62.3 (q, 3-OCH₃), 61.8 (q, 4-OCH₃), 56.8 (q, 9-OCH₃)。

Compound 5: a brown solid.¹H-NMR (400 MHz, acetone-d ₆) δ: 9.03 (1H, s, 1-OH), 8.58 (1H, s, 6-OH), 7.57 (1H, dd, J = 7.8, 1.1 Hz, H-9), 7.16 (1H, t, J = 7.8 Hz, H-8), 6.95 (1H, dd, J = 7.8, 1.1 Hz, H-7), 4.05 (3H, s, 4-OCH₃), 3.98 (3H, s, 2-OCH₃), 3.85 (3H, s, 3-OCH₃); ¹³C-NMR (100 MHz, acetone-d ₆) δ: 141.6 (s, C-1), 137.2 (s, C-2), 146.9 (s, C-3), 132.6 (s, C-4), 145.9 (s, C-4a), 145.1 (s, C-5a), 143.2 (s, C-6), 113.7 (d, C-7), 124.6 (d, C-8), 114.2 (d, C-9), 126.5 (s, C-9a), 110.2 (s, C-9b), 61.8 (q, 2-OCH₃), 61.8 (q, 3-OCH₃), 61.6 (q, 4-OCH₃)。

Compound 6: a tan solid.¹H-NMR (400 MHz, acetone-d ₆) δ: 7.53 (1H, t, J = 8.4 Hz, H-9), 7.15 (1H, t, J = 8.4 Hz, H-8), 6.99 (1H, d, J = 8.4 Hz, H-7), 4.13 (3H, s, 4-OCH₃), 4.01 (3H, s, 1-OCH₃), 3.96 (3H, s, 3-OCH₃); ¹³C-NMR (100 MHz, acetone-d ₆) δ: 137.9 (s, C-1), 141.7 (s, C-2), 140.2 (s, C-3), 136.2 (s, C-4), 142.1 (s, C-4a), 145.6 (s, C-5a), 144.2 (s, C-6), 114.5 (d, C-7), 124.6 (d, C-8), 113.8 (d, C-9), 125.8 (s, C-9a), 115.4 (s, C-9b), 61.9 (q, 3-OCH₃), 61.3 (q, 4-OCH₃), 60.8 (q, 1-OCH₃)。

Compound 7: a tan solid.¹H-NMR (400 MHz, acetone-d ₆) δ: 7.44 (1H, d, J = 8.5 Hz, H-9), 6.89 (1H, d, J = 8.5 Hz, H-8), 7.10 (1H, s, H-1), 4.22 (3H, s, 4-OCH₃), 4.13 (3H, s, 6-OCH₃), 3.90 (3H, s, 6-OCH₃); ¹³C-NMR (100 MHz, acetone-d ₆) δ: 100.1 (d, C-1), 148.3 (s, C-2), 139.5 (s, C-3), 139.7 (s, C-4), 142.3 (s, C-4a), 148.2 (s, C-5a), 133.9 (s, C-6), 149.3 (s, C-7), 113.3 (d, C-8), 115.2 (d, C-9), 119.4 (s, C-9a), 122.1 (s, C-9b), 61.8(q, 3-OCH₃), 61.3 (q, 4-OCH₃), 61.2 (q, 6-OCH₃)。

Compound 8: a pale yellow solid.¹H-NMR (400 MHz, acetone-d ₆) δ: 8.11 (1H, s, 8-OH), 7.89 (1H, s, 2-OH), 7.26 (1H, s, H-1), 7.17 (1H, d, J = 8.7 Hz, H-6), 7.01 (1H, d, J = 8.7 Hz, H-7), 4.16 (3H, s, 4-OCH₃), 4.01 (3H, s, 9-OCH₃), 3.92 (3H, s, 3-OCH₃); ¹³C-NMR (100 MHz, acetone-d ₆) δ: 102.5 (d, C-1), 148.1 (s, C-2), 140.3 (s, C-3), 139.4 (s, C-4), 143.0 (s, C-4a), 151.8 (s, C-5a), 107.5 (d, C-6), 116.6 (d, C-7), 146.1 (s, C-8), 142.3 (s, C-9), 119.0 (s, C-9a), 120.1 (s, C-9b), 61.7 (q, 3-OCH₃), 61.1 (q, 4-OCH₃), 60.8 (q, 9-OCH₃)。

Compound 9: a red solid.¹H-NMR (400 MHz, CDCl₃) δ: 7.10 (1H, s, H-1), 6.85 (1H, d, J = 2.2 Hz, H-9), 6.56 (1H, d, J = 2.2 Hz, H-7), 4.26 (3H, s, 4-OCH₃), 4.02 (3H, s, 6-OCH₃), 4.00 (3H, s, 8-OCH₃), 3.89 (3H, s, 3-OCH₃); ¹³C-NMR (100 MHz, CDCl₃) δ: 99.2 (d, C-1), 145.9 (s, C-2), 138.2 (s, C-3), 138.4 (s, C-4), 142.5 (s, C-4a), 145.8 (s, C-5a), 140.8 (s, C-6), 98.7 (d, C-7), 156.7 (s, C-8), 94.2 (d, C-9), 125.6 (s, C-9a), 121.3 (s, C-9b), 61.8 (q, 3-OCH₃), 61.0 (q, 4-OCH₃), 56.3(q, 6-OCH₃), 56.1 (q, 8-OCH₃)。

Example 3:

lipid-lowering activity assay of the compounds of the invention:

HepG2 cells were cultured at 1.5X 10⁴The number of seeds/well was inoculated in a 96-well plate and incubated at 37 ℃ in 5% CO₂When the cell abundance reached more than 90% by culturing in the environment, the old culture solution was replaced with a fresh culture solution containing 0.6 mM oleic acid and 0.3 mM sodium palmitate (oleic acid and sodium palmitate were bound by 40% bovine serum albumin). After 24 h incubation, 20. mu.L of the extract solution was added to each well of the test group to a final concentration of 0.5 mg/mL, and 20. mu.L of the corresponding vehicle was added to each well of the blank and model groups. After continuing culturing for 24 h, the culture solution in the wells was carefully removed, washed once with Phosphate Buffered Saline (PBS), and 60 μ L of 10% formaldehyde solution was added to each well. After fixation for 30 min, the fixative solution is removed, PBS is washed once, and 60 muL of 0.4 mg/mL oil red O working solution is added into each hole for dyeing. After 60 min, the staining solution was removed, washed 3 times with PBS, and placed under an inverted microscope to observe the staining of intracellular lipid droplets. Finally, 200. mu.L of isopropanol is added into each hole, and after full dissolution, the OD value of each group under 510 nm is measured by an enzyme-labeling instrument. The fat reduction rate is calculated according to the following formula: fat reduction rate% = [ 1- (test group OD value-blank group OD value)/(model group OD value-blank group OD value) ]] × 100。

The activity data are shown in Table 1.

TABLE 1 data for lipid lowering Activity of Compounds 1-9 (mean ±)s, n = 4）

Example 4:

and (3) tablet preparation: 1-910 mg, 180 mg of lactose, 55 mg of starch, 5 mg of magnesium stearate of any one of the compounds obtained in examples 1 and 2;

the preparation method comprises the following steps: the compound, lactose and starch were mixed, uniformly moistened with propylene glycol, the moistened mixture was sieved and dried, and sieved again, magnesium stearate was added, and the mixture was then tabletted to a weight of 250 mg per tablet, with a compound content of 10 mg.

Example 5:

an ampoule agent: 1-92 mg of any one of the compounds obtained in examples 1 and 2;

the preparation method comprises the following steps: any of the compounds 1 to 9 obtained in examples 1 and 2 was dissolved in 3 mL of propylene glycol, and the resulting solution was filtered and aseptically filled in an ampoule.

Example 6:

and (3) capsule preparation: 1-910 mg of any one of the compounds obtained in examples 1 and 2, 187 mg of lactose, 3 mg of magnesium stearate;

the preparation method comprises the following steps: mixing the compound with adjuvants, sieving, mixing, and encapsulating the obtained mixture into hard gelatin capsules with 200 mg of active ingredient content of 10 mg in each capsule.