CN1185243C - Tetrahydroxydiphenylethylene glycosides compounds for preventing and curing angiocardiopathy and cerebrovascular diseases - Google Patents

Abstract

The present invention relates to a novel compound of tetrahydroxystilbene glycosides for preventing and treating cardiovascular and cerebrovascular diseases, including 2,3,5,4′-tetrahydroxystilbene-2-O-(-2″-O-ferula Acyl)-β-D-glucoside, 2,3,5,4′-tetrahydroxystilbene-2-O-(-2″-O-p-coumaroyl)-β-D-glucoside, 2, 3,5,4′-tetrahydroxystilbene-2-O-(2″-O-acetyl)-β-D-glucoside, 2,3,5,4′-tetrahydroxystilbene-2 -O-(6″-O-α-D-glucose)-β-D-glucoside has strong and extensive cardiovascular and cerebrovascular activities, and can be used to prepare medicines and health food for preventing and treating cardiovascular and cerebrovascular diseases.

Description

A new stilbene glycoside compound for preventing and treating cardiovascular and cerebrovascular diseases

The invention relates to the technical field of medicine, and is a novel stilbene glycoside compound for preventing and treating cardiovascular and cerebrovascular diseases.

Atherosclerosis and the cardiovascular and cerebrovascular diseases caused by it, such as myocardial infarction and cerebral infarction, have now leapt to the list of the main causes of death in my country. It is generally believed that the current clinical treatment of the above diseases, especially atherosclerosis There is no ideal preventive drug yet.

The object of the present invention is to provide a new stilbene glycoside compound for preventing and treating cardiovascular and cerebrovascular diseases.

The chemical structure of the compound is

Wherein the _R1 and _R2 groups are respectively:

First group

R₂为OH _R1 is

_R2 is OH

2,3,5,4′-Tetrahydroxystilbene-2-O-(-2″-O-feruloyl)-β-D-glucoside (hereinafter referred to as compound 1)

Second Group

R₂为OH _R1 is

_R2 is OH

2.3,5,4′-Tetrahydroxystilbene-2-O-(-2″-O-p-coumaroyl)-β-D-glucoside (hereinafter referred to as compound 2)

The third group

_R1 is OH _R2 is _CH2COO

2,3,5,4′-Tetrahydroxystilbene-2-O-(2″-O-acetyl)-β-D-glucoside (hereinafter referred to as compound 3)

Fourth group

R ₁ =OH

2,3,5,4′-Tetrahydroxystilbene-2-O-(6″-O-α-D-glucose)-β-D-glucoside (hereinafter referred to as compound 4)

The present invention uses the tuber root (raw product or processed product) of Polygonum multiflorum Thunb. as raw material, and the specific preparation process is as follows: the dried root is pulverized to make medicinal material coarse powder, and 80% ethanol Soak for about 1 week, then extract by percolation with 80% ethanol as usual until the percolation liquid is almost colorless, combine the leachate and percolation liquid, concentrate under reduced pressure to obtain the extract, suspend the extract with 3 to 5 times the amount of water, press Routinely extract with petroleum ether, chloroform, ethyl acetate, and water-saturated n-butanol sequentially, pass the n-butanol extract through a macroporous resin column, and elute with water, 10%, 30%, 50%, 70%, and 95% ethanol ; 50% of the eluted fractions were collected and subjected to silica gel column chromatography, followed by chloroform/methanol (20:1, 15:1, 10:1, 7:1, 5:1, 3:1, 2:1, 1 : 1, 1: 2) solvent system elution, collect the elution site of chloroform/methanol (7: 1), and then repeat the column chromatography, first obtain the light brown crystal of compound 1, and then obtain the colorless crystal of compound 2; collect Chloroform/methanol (5:1) elution site, and then repeated column chromatography, the colorless crystal of compound 3 can be obtained; collect chloroform/methanol (3:1) elution site, pass through SephadexLH20 and ODS column, with 30% methanol After elution, the crude product of compound 4 can be obtained, which is prepared and purified by high-pressure liquid chromatography (HPLC) to obtain light brown crystals.

In order to further illustrate the application of the present invention in the field of medicine and health food, some pharmacological results are used to illustrate that platelets are an essential substance for thrombus formation, and their functional activation is a manifestation of a hypercoagulable state. Inhibition of platelet aggregation is important for antithrombotic However, substances with antithrombosis, protection of vascular endothelial cells, inhibition of proliferation of vascular smooth muscle cells, and anti-oxidation can antagonize the formation and development of atherosclerosis from multiple links, so substances with the above effects can be used in the preparation and prevention of atherosclerosis Drugs and health foods for cardiovascular and cerebrovascular diseases such as sclerosis and myocardial infarction and cerebral infarction caused by it have a wide range of uses.

1. Anti-platelet aggregation effect:

Blood was taken from the rabbit heart, platelet aggregation was induced by adenosine diphosphate, and the inhibitory rate of the compound was investigated. At 100 μM, the inhibitory rate of compound 1 was 86%, the inhibitory rate of compound 2 was 84%, and the inhibitory rate of compound 3 was 66%. %, the inhibition rate of compound 4 was 57%, indicating that

The new stilbene glycoside compound of the present invention has strong anti-platelet aggregation effect.

2. Inhibit the proliferation of vascular smooth muscle cells:

Taking the cultured bovine aortic smooth muscle cells as the experimental object, they were divided into 1 control group and 4 test groups. Compound 1, compound 2, compound 3 and compound 4 were added to the 4 test groups respectively, and the final concentration of each compound was 100 μM. , co-cultured for 48 hours, and investigated the inhibitory effect of each compound on the proliferation of smooth muscle cells. The inhibitory rate of compound 1 was 46%, the inhibitory rate of compound 2 was 54%, the inhibitory rate of compound 3 was 58%, and the inhibitory rate of compound 4 was 58%. 42%, indicating that the new stilbene glycoside compound of the present invention has a strong inhibitory effect on the proliferation of vascular smooth muscle cells.

3. Protective effect on vascular endothelial cell injury:

Taking the cultured bovine aortic endothelial cells as the experimental object, they were divided into a control group and 4 test groups, all of which were added with tumor necrosis factor-α (TNFα), and the 4 test groups were respectively added with compound 1, compound 2, compound 3, Compound 4, the final concentration of each compound was 100 μM. After co-incubating for 24 hours, the activity of lactate dehydrogenase (LDH) in the supernatant of the cells was measured and the cell morphology was observed; compound 1 reduced the activity of LDH by 58%, and compound 2 reduced the activity of LDH 46%, compound 3 reduced LDH activity by 63%, compound 4 reduced LDH activity by 35%, and the morphology of endothelial cells in each test group was normal, showing that the new compound of stilbene glycosides of the present invention has a strong effect on vascular endothelial cell damage. Protective effects.

4. Antioxidant effect:

30 Kunming mice were divided into normal group and 2 test groups. The 2 test groups were administrated with 5 mg/kg and 20 mg/kg of the active ingredients containing 80% of the total content of this class of compounds by intragastric administration respectively. After 3 days, the The neck was put to death, the whole brain homogenate was taken, and the lipid peroxide (LPO) value was measured, and the effective component (20mg/kg) reduced the LPO value in the normal mouse brain by 23%, indicating that the new stilbene glycoside compound of the present invention Has a significant antioxidant effect.

5. Acute toxicity test

50 mice of Kunming species were divided into 5 groups, and they were given 1.0g/kg, 2.0g/kg, 4.0g/kg, 6.0g/g, 1.0g/kg, 6.0g/g, 7.8g/kg, observe the general performance and the number of deaths of the mice in each dosage group within 7 days, and carry out anatomical and pathological examinations on the survival and dead mice. See abnormal performance, indicating that this type of compound has high safety.

Example: extracting and separating tetrahydroxystilbene glycosides from Polygonum multiflorum.

Grind 20 kg of dried root of Radix Polygoni Multiflori to make coarse powder of medicinal material, soak in 80% ethanol in a sufficient amount (more than 3 times the amount) for about 1 week, and then extract by percolation with 80% ethanol as usual until the percolation liquid is almost colorless, The leachate and percolation liquid were combined, concentrated under reduced pressure to obtain the extract, which was suspended in 3 to 5 times the amount of water, and extracted with petroleum ether, chloroform, ethyl acetate, and water-saturated n-butanol in sequence. Extract part of the macroporous resin with n-butanol, elute with water and 10%, 30%, 50%, 70%, 95% ethanol; collect 50% of the eluted components, concentrate under reduced pressure, and take 250g for silica gel column chromatography , sequentially eluted with chloroform/methanol (20:1, 15:1, 10:1, 7:1, 5:1, 3:1, 2:1, 1:1, 1:2) solvent system, and collected chloroform Chloroform/methanol (7:1) elution site, and then repeated column chromatography, first can obtain the light brown crystal of compound 1 (223mg), then obtain the colorless crystal of compound 2 (142mg); Collect chloroform/methanol (5:1 ) elution site, and then repeated column chromatography, the colorless crystals (433mg) of compound 3 can be obtained; the elution site of chloroform/methanol (3:1) is collected, passed through SephadexLH20 and ODS column, with 30% methanol elution, can The crude product of compound 4 was obtained, which was prepared and purified by HPLC to obtain light brown crystals (33 mg).

Structure Identification:

Compound 1: The molecular weight is 582, the molecular formula is C ₃₀ H ₃₀ O ₁₂ , its properties are light brown needle crystals (methanol), and the melting point range is 202-203°C; FAB-MSm/z: 582 (M ⁺ ), El-MSm /z: 244 (M ⁺ -glc), IRmax (KBr, cm ^-1 ): 3409 (OH), 1703 (C=O), 1603, 1515, 1463 (benzenering); ¹ H-NMR, ¹³ C-NMR See Table 1 for the data.

Compound 2: The molecular weight is 552, its properties are colorless needle crystals (methanol), and the melting point range is 197-198°C; FAB-MSm/z: 552 (M ⁺ ), El-MSm/z: 244 (M ⁺ -glc ), IRmax (KBr, cm ^-1 ): 3412 (OH), 1700 (C=O), 1606, 1515, 1465 (benzene ring); see Table 2 for ¹ H-NMR and ¹³ C-NMR data.

Compound 3: The molecular weight is 448, its properties are colorless needle crystals (methanol), and the melting point range is 173-174°C; IRmax (KBr) cm ^-1 : 3409 (OH), 1722 (C=O), 1606, 1515, 1463; FAB-MS: 448 (M ⁺ ), EI-MS m/z (%): 244 (100); see Table 3 for ¹ H-NMR and ¹³ C-NMR data.

Compound 4: The molecular weight is 568, its properties are light brown needle crystals (methanol), and its melting point range is 203-204°C. IRmax (KBr) cm ^-1 : 3409 (OH), 1606, 1515, 1463; FAB-MSm/z: 568 (M ⁺ ), 591 (M ⁺ +Na), 608 (M ⁺ +K), , EI- MSm/z (%): 244 (100); ¹ H-NMR (DMSO-d ₆ , TMS) δ: 4.41 (1H, d, J=7.8Hz, 1″-H), 4.71 (1H, d, J =3.6Hz, 1-H); 6.18(1H, d, J=2.7Hz, 4-H), 6.55(1H, d, J=2.7Hz, 6-H), 6.75(2H, m, 3′ -H, 5'-H), 7.39 (2H, m, 2'-H, 6'-H); 6.87 (1H, d, J=16.5Hz, β-H), 7.63 (1H, d, J= 16.5Hz, α-H); see Table 4 for ¹³ C-NMR data.

The symbols in the foregoing embodiments are represented as follows:

IR is infrared spectrum; FAB-MS is fast atom bombardment mass spectrometry; EI-MS is electron bombardment mass spectrometry; ¹ H-NMR is hydrogen nuclear magnetic resonance; ¹³ C-NMR is carbon nuclear magnetic resonance; CD ₃ OD is deuterated methanol; DMSO-d ₆ is deuterated dimethylsulfoxide.

The advantages of the present invention are: the source of raw materials of this type of compound is abundant, cheap, no toxic and side effects, the preparation process is relatively simple, and can be used as an oral dosage form or injection; it has the effects of significantly inhibiting the proliferation of vascular smooth muscle cells, protecting vascular endothelial cells, and anti-oxidation , which is beneficial to the prevention and treatment of cardiovascular and cerebrovascular diseases; it can be used to prepare drugs for preventing and treating cardiovascular and cerebrovascular diseases, and can also be used to prepare health food.

Table 1. 2,3,5,4′-Tetrahydroxystilbene-2-O-(-2″-O-feruloyl)-β-D-glucose

NMR data of glycosides (DMSO-d ₆ )

Hydrogen Chemical Shift (δ) Coupling Constant (Hz) Carbon Chemical Shift (δ)

Stilbene-4 6.22(d) 2.72 Stilbene-1 131.97s

6 6.48(d) 2.72 2 134.46s

2 ′, 6 ′ 7.38 (m) 3 150.10s

3 ′, 5 ′ 6.78 (m) 4 102.92D

α 7.21(d) 16.44 5 154.10s

β 6.88(d) 16.44 6 101.40d

Feruloyl-2 7.20(d) 1.84 1′ 128.88s

5 7.42(d) 8.31 2′, 6′ 127.93d

6 7.01(dd) 8.31, 1.84 3′, 5′ 116.26d

7 7.60(d) 15.18 4′ 157.25s

8 6.46(d) 15.18 α 120.02d

OCH ₃ 3.70(s) β 128.66d

β-Glucose-1″ 4.92(d) 8.05 Feruloyl-1 125.56s

2″ 5.06(dd) 8.05, 8.02 2 111.30d

3″ 3.58(m) 3 147.83s

4″ 3.40(m) 4 149.22s

5″ 3.22(m) 5 115.52d

6″ 3.60(m) 6 122.68d

7 145.21d

8 114.55d

COO 165.95s

OCH ₃ 55.62q

                                                                                                           

2 ″ 73.96D

|

4 ″ 69.74D

5 ″ 76.98D

6″ 60.57t

Table 2. 2,3,5,4′-Tetrahydroxystilbene-2-O-(-2″-O-p-coumaroyl)-β-D-glucoside

NMR data (CD ₃ OD)

Hydrogen Chemical Shift (δ) Coupling Constant (Hz) Carbon Chemical Shift (δ)

Stilbene-4 6.23(d) 2.86 Stilbene-1 133.06s

6 6.55(d) 2.86 2 136.46s

2 ′, 6 ′ 7.39 (m) 3 152.02s

3 ′, 5 ′ 6.74 (m) 4 103.86d

α 7.18(d) 16.26 5 155.88s

β 6.93(d) 16.26 6 103.41d

Coumaroyl-2,6 7.19(m) 1′ 131.73s

3, 5 6.74(m) 2′, 6′ 129.41d

7 7.58(d) 15.85 3′, 5′ 116.59d

8 6.32(d) 15.85 4′ 158.71s

β-glucose-1″ 4.93(d) 8.18 α 120.95d

2″ 5.19(dd) 8.18, 8.12 β 130.94d

                                                                       

4″ 3.60(m) 2.6 131.28d

5″ 3.35(m) 3.5 116.76d

6″ 3.80(m) 4 161.25s

7 147.46d

8 114.86d

COO 168.62s

                                                                                           

2 ″ 75.31D

|

4″ 71.07d

5 ″ 78.49d

6″ 62.01t

Table 3. 2,3,5,4′-Tetrahydroxystilbene-2-O-(2″-O-acetyl)-β-D-glucoside

NMR data of (DMSO-d ₆ )

Hydrogen Chemical Shift (δ) Coupling Constant (Hz) Carbon Chemical Shift (δ)

Stilbene-4 6.20(d) 2.6 Stilbene-1 131.9s

6 6.56(d) 2.6 2 2 136.4s

2′, 6′ 7.34(m) 3 150.6s

3 ′, 5 ′ 6.72 (m) 4 102.5D

α 7.59(d) 16.5 5 154.8s

β 6.90(d) 16.5 6 100.9d

3-OH 9.02(s) 1′ 128.5s

5-OH 9.29(s) 2′, 6′ 127.8d

4′-OH 9.64 (s) 3 ′, 5 ′ 115.5D

Acetyl-CH ₃ 1.78(s) 4′ 157.2s

β-glucose-1″ 4.47(d) 7.6 α 120.5d

2″ 3.32(m) β 128.3d

                                                                                                           

4″ 3.35(m) CH ₃ 20.4q

                                                                                         

6″ 4.20(d) 10.2 2″ 73.9d

4.11(m) 3″ 74.0d

4″ 69.7d

5″ 75.8d

6″ 64.0t

Table 4. 2,3,5,4′-Tetrahydroxystilbene-2-O-(6″-O-α-D-glucose)-β-D-glucoside

¹³ CNMR data (DMSO-d ₆ )

Carbon Chemical Shift (δ) Carbon Chemical Shift (δ)

Stilbene-1 131.87s β-glucose-1″ 106.71d

2 136.28s 2″ 73.36d

                                                                                

4 102.79d 4″ 69.31d

5 154.71s 5″ 76.22d

6 101.15d 6″ 65.92t

                                                                                       

                                                                                                                                                                                                                          will 72.19 d

3 ′, 5 ′ 115.66d 3 75.16d

4′ 157.24s 4 70.20d

|

β 6 60.84t

Claims (2)

1, a kind of tetrahydroxystilbene glycosides compound, its structure is:

R wherein ₁And R ₂Group is respectively:

First group

R ₁For R ₂Be OH

Second group

R ₁For R ₂Be OH

The 3rd group

R ₁Be OH R ₂Be CH ₃COO

The 4th group

R ₁Be OH R ₂For

2, the application of the described compound of claim 1 in preparation control cardiovascular and cerebrovascular diseases medicament or protective foods.