CN113754662A

CN113754662A - RS-heptacyclic aldehyde, its synthesis, activity and application

Info

Publication number: CN113754662A
Application number: CN202010501811.5A
Authority: CN
Inventors: 赵明; 彭师奇; 侯梦雨
Original assignee: Capital Medical University
Current assignee: Capital Medical University
Priority date: 2020-06-04
Filing date: 2020-06-04
Publication date: 2021-12-07

Abstract

The invention discloses (2 'S, 5' S) -tetrahydropyrazines [1 ', 2': 1, 6) of the formula]And bis { (1R/S) - [ 1-oxoethyl-2-yl]-2,3,4, 9-tetrahydro-1H-pyridine [3,4-b ]]And indole } -1 ', 4' -dione (RS-heptacyclic aldehyde for short). Its preparing process, the activity of resisting venous thrombus and arterial thrombus, and the bleeding side effect are disclosed. Also discloses the activity of reducing the contents of blood coagulation factors X, P selectin and GPIIb/IIIa. Therefore, the invention discloses the application of the derivative in preparing anti-venous thrombosis drugs, anti-arterial thrombosis drugs, blood coagulation factor X antagonists, P-selectin antagonists and GPIIb/IIIa antagonists.

Description

RS-heptacyclic aldehyde, its synthesis, activity and application

Technical Field

The invention relates to (2 'S, 5' S) -tetrahydropyrazinyl [1 ', 2': 1,6] bis { (1R/S) - [ 1-oxoethyl-2-yl ] -2,3,4, 9-tetrahydro-1H-pyridine [3,4-b ] indole } -1 ', 4' -dione (RS-heptacyclal for short). Relates to its preparation method, its activity for resisting venous thrombosis and arterial thrombosis, and further relates to its bleeding side effect. Simultaneously relates to the activity of reducing the content of blood coagulation factors X, P selectin and GPIIb/IIIa. Therefore, the invention relates to the application of the derivative in preparing anti-venous thrombosis drugs, anti-arterial thrombosis drugs, blood coagulation factor X antagonists, P-selectin antagonists and GPIIb/IIIa antagonists. The invention belongs to the field of biological medicine.

Background

Both arterial and venous thrombosis have become diseases with a high incidence of morbidity and mortality. Coronary atherosclerotic heart disease is very severe in diseases associated with arterial thrombosis. In order to treat coronary heart disease, the patient needs to be placed with a stent. After the stent implantation operation, patients need to take anti-platelet medicines for anti-thrombosis treatment for a long time. In addition, patients undergoing mechanical heart valve replacement also need to take lifelong anticoagulation medications to prevent thrombus formation. Venous thrombotic diseases include deep vein thrombosis, which is the most common thrombotic disease, and pulmonary embolism, which has a high lethality rate. In 8 years from 2007 to 2016, the hospitalization rate of deep vein thrombosis patients in China is increased from 3.2/10 ten thousand to 17.5/10 ten thousand. The incidence of deep vein thrombosis continues to rise. The incidence of venous thrombosis also increases exponentially with the aging, and the negative influence of the venous thrombosis on the people in China is particularly serious as the people in China age gradually. Patients also need to take the oral anticoagulant warfarin or rivaroxaban after discharge for the purpose of treating venous thrombotic disease or preventing recurrence of deep vein thrombosis. However, both warfarin and rivaroxaban increase the risk of bleeding in patients. Therefore, the invention of the novel antithrombotic drug which has antithrombotic curative effect and can avoid bleeding side effect is very important.

The inventors have disclosed heptacyclic aldehydes of the formula (CN 110577530A). However, the heptacyclic aldehydes disclosed only inhibit venous thrombosis, but do not inhibit arterial thrombosis, blood coagulation factor X, P-selectin and GPIIb/IIIa. The inventors have conducted extensive studies and studies to overcome these disadvantages of the heptacyclic aldehydes disclosed, and have found that RR-heptacyclic aldehydes of the following formula inhibit not only venous thrombosis but also arterial thrombosis, blood coagulation factor X, P-selectin and GPIIb/IIIa. Based on these findings, the inventors have proposed the present invention.

Disclosure of Invention

In a first aspect of the invention there is provided (2 'S, 5' S) -tetrahydropyrazinyl [1 ', 2': 1,6] bis { (1R/S) - [ 1-oxoethyl-2-yl ] -2,3,4, 9-tetrahydro-1H-pyrido [3,4-b ] indolino } -1 ', 4' -dione (RS-heptacyclal for short) of the formula.

The second aspect of the present invention provides a process for producing RS-heptacyclic aldehyde, which comprises:

1) performing Pictet-Spengler condensation on L-tryptophan benzyl ester and 1,1,3, 3-tetramethoxypropane to generate (3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro-beta-carboline-3-carboxylic acid benzyl ester;

2) (3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro- β -carboline-3-carboxylic acid benzyl ester is converted to the corresponding (1R,3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro- β -carboline-3-carboxylic acid and (1S,3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro- β -carboline-3-carboxylic acid;

3) condensing (1R,3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro-beta-carboline-3-carboxylic acid and (1S,3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro-beta-carboline-3-carboxylic acid into RS-heptacyclic acetal serving as a precursor of RS-heptacyclic aldehyde;

4) the RS-heptacyclic acetal is converted into RS-heptacyclic aldehyde.

The third content of the present invention is to evaluate the anti-arterial thrombosis effect of RS-heptacyclic aldehyde.

The fourth aspect of the present invention is to evaluate the anti-venous thrombosis effect of RS-heptacyclic aldehyde.

The fifth aspect of the present invention is to evaluate bleeding risk of RS-heptacyclic aldehyde.

A sixth aspect of the present invention is to evaluate the effect of RS-heptacyclaldehyde on plasma P-selectin.

The seventh aspect of the present invention is to evaluate the effect of RS-heptacyclic aldehyde on plasma GPIIb/IIIa.

An eighth aspect of the present invention is to evaluate the effect of RS-heptacyclic aldehyde on plasma coagulation factor X.

Drawings

FIG. 1 is a synthetic route for RS-heptacyclic aldehydes. (i) Dichloromethane, trifluoroacetic acid, 1,1,3, 3-tetramethoxypropane; (ii) CH (CH)₃OH,Pd/C,H₂(ii) a (iii)2- (7-azabenzotriazole) -N, N' -tetramethyluronium hexafluorophosphate, triethylamine, anhydrous N, N-dimethylformamide; (iv) glacial acetic acid, H₂O。

Detailed Description

To further illustrate the invention, a series of examples are given below. These examples are purely illustrative and are intended to be a detailed description of the invention only and should not be taken as limiting the invention.

EXAMPLE 1 preparation of benzyl (3S) -1- (2, 2-Dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro-beta-carboline-3-carboxylate (1)

A mixed solution of 100mL of methylene chloride, 4.0mL of 1,1,3, 3-tetramethoxypropane and 3.0mL of trifluoroacetic acid was stirred for 40 minutes under ice bath, and then 4g (13.6mmol) of L-Trp-OBzl was added and stirred at 60 ℃ for 7 hours. At this time, thin layer chromatography (petroleum ether/ethyl acetate, 1/1) showed that L-Trp-OBzl disappeared, whereupon the reaction was terminated. The reaction mixture was concentrated under reduced pressure and the residual oil was dissolved in 80mL of ethyl acetate. The resulting solution was sequentially saturated with NaHCO₃Aqueous solution (30 mL. times.3) and saturated aqueous NaCl solution (30 mL. times.3). The combined ethyl acetate layer was added with anhydrous Na₂SO₄Drying for 6 h, filtration, concentration of the filtrate under reduced pressure and purification of the residue by column chromatography on silica gel (petroleum ether/ethyl acetate, 2/1) gave 3.26g (61%) of the title compound as a yellow oil. ESI-MS (M/e)393[ M-H]^-；¹HNMR(300MHz,DMSO-d₆)δ/ppm＝10.83(d,J＝14.4Hz,1H),7.43～7.22(m,7H),7.03(t,J＝6.9Hz,1H),6.95(t,J＝7.2Hz,1H),5.20(m,2H),4.69(m,1H),4.26(d,J＝9.6Hz,0.43H),4.16(m,0.56H),4.00(m,0.45H),3.75(m,0.57H),3.31(d,J＝5.1Hz,3H),3.26(d,J＝7.8Hz,3H),2.96(m,1H),2.85(m,0.49H),2.70(m,1H),2.42(m,0.53H),2.13(m,0.42H),1.82(m,1H)。

EXAMPLE 2 preparation of (1R,3S) -and (1S,3S) -1- (2, 2-Dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro-beta-carboline-3-carboxylic acid (2-RS and 2-SS)

4.33g (11.0mmol) of (3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro- β -carboline-3-carboxylic acid benzyl ester (1) dissolved in 80mL of CH₃734mg Pd/C was added, stirring was continued, then hydrogen was added, stirring was carried out at room temperature for 12 hours, and TLC (petroleum ether/ethyl acetate, 1/1) showed disappearance of starting material 1. The reaction is then terminated, filtered, the filtrate is concentrated under reduced pressure, and the residue is dissolved in 10mL of purified water and separated by preparative high performance liquid chromatography (methanol/water, 1/4) to yield 2.34g (70%) (1R,3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro- β -carboline-3-carboxylic acid (2-RS) and 0.70g (21%) (1S,3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro- β -carboline-3-carboxylic acid (2-SS).

2-RS:ESI-FT-ICR-MS(m/e)305.14986[M+H]⁺；¹H NMR(800MHz,DMSO-d₆)δ/ppm＝10.86(s,1H),8.30(s,1H),7.40(d,J＝8.0Hz,1H),7.30(d,J＝8.0Hz,1H),7.05(t,J＝7.2Hz,1H),6.96(t,J＝7.2Hz,1H),4.73(m,1H),4.47(d,J＝8.0Hz,1H),3.77(m,1H),3.33(s,3H),3.32(s,3H),2.99(dd,J₁＝4.0Hz,J₂＝15.2Hz,1H),2.82(dd,J₁＝8.0Hz,J₂＝15.2Hz,1H),2.16(d,J＝12.8Hz,1H),2.05(m,1H)；

2-SS:ESI-FT-ICR-MS(m/e):305.15063[M+H]⁺；¹H NMR(800MHz,DMSO-d₆)δ/ppm＝10.90(s,1H),8.32(s,1H),7.41(d,J＝8.0Hz,1H),7.32(d,J＝8.0Hz,1H),7.05(t,J＝7.2Hz,1H),6.97(t,J＝7.2Hz,1H),4.68(s,1H),4.39(s,1H),3.51(m,1H),3.32(s,6H),3.04(d,J＝14.4Hz,1H),2.66(t,J＝12.8Hz,1H),2.47(m,1H),1.98(m,1H)。

EXAMPLE 3 preparation of (2 'S, 5' S) -Tetrahydropyrazine [1 ', 2': 1,6] and bis { (1R/S) - [ 1-dimethoxyethyl-2-yl ] -2,3,4, 9-tetrahydro-1H-pyrido [3,4-b ] indole } -1 ', 4' -dione (3, RS-heptacyclic acetal)

0.304g (1.0mmol) of 2-RS and 0.304g (1.0mmol) of 2-SS are dissolved in 60mL of anhydrous N, N-Dimethylformamide (DMF), 0.57g (1.5mmol) of 2- (7-azabenzotriazole) -N, N, N ', N' -tetramethylurea hexafluorophosphate (HATu) and 0.17g (1.2mmol) of 1-hydroxybenzotriazole (HOBt) are added with stirring in an ice bath, then the reaction solution is adjusted to pH 9 with triethylamine and stirred at room temperature for 18 hours, TLC (petroleum ether/ethyl acetate, 1/1) shows disappearance of 2-RS and 2-SS. The reaction was terminated, the reaction was concentrated under reduced pressure, the residue was dissolved in 80mL ethyl acetate and successively saturated NaHCO₃Aqueous solution (40 mL. times.3), saturated aqueous NaCl solution (40 mL. times.3), 5% KHSO₄Aqueous wash (40 mL. times.3), saturated aqueous NaCl wash (40 mL. times.3), 5% NaHCO₃Aqueous solution (40 mL. times.3) and saturated aqueous NaCl solution (40 mL. times.3). The combined ethyl acetate phases were treated with anhydrous Na₂SO₄Drying for 8 hours, filtering, and concentrating the filtrate under reduced pressure. The resulting residue was purified by silica gel column chromatography (petroleum ether/ethyl acetate, 2/1; dichloromethane/methanol, 100/1) to give 372mg (65%) of the title compound as a pale yellow solid. ESI-MS (M/e):573[ M + H]⁺；

(c＝0.11,CH₃OH)；M.p.131～132℃；IR(cm^-1):3407.25,3304.95,2924.38,2833.52,2359.43,2331.09,1732.56,1650.90,1451.44,1423.38,1327.53,1260.39,1229.50,1189.91,1149.36,1119.31,1055.26,1008.09,970.34,883.18,840.39,739.53,682.19；¹H NMR(800MHz,DMSO-d₆)δ/ppm＝10.97(s,1H),10.95(s,1H),7.48(d,J＝7.8Hz,1H),7.43(d,J＝7.8Hz,1H),7.37(d,J＝8.1Hz,1H),7.33(d,J＝8.1Hz,1H),7.07(t,J＝7.5Hz,2H),6.98(m,2H),5.84(m,1H),5.21(t,J＝5.3Hz,1H),4.52(t,J＝5.5Hz,1H),4.47(dd,J₁＝11.2,J₂＝4.6,1H),4.36(dd,J₁＝15.6,J₂＝7.9,1H),4.17(m,1H),3.47(dd,J₁＝20.0,J₂＝11.4,1H),3.29(m,1H),3.27(s,3H),3.19(s,3H),3.12(s,3H),3.07(s,3H),2.84(m,1H),2.77(m,1H),2.53(m,1H),2.42(m,1H),2.17(m,2H)。

EXAMPLE 4 preparation of (2 'S, 5' S) -Tetrahydropyrazine [1 ', 2': 1,6] and bis { (1R/S) - [ 1-oxoethyl-2-yl ] -2,3,4, 9-tetrahydro-1H-pyridine [3,4-b ] indoline } -1 ', 4' -dione (4, RS-heptacyclal)

50mg (0.09mmol) of RS-heptacyclic acetal (3) was dissolved in 4mL of glacial acetic acid, and 0.4mL of ultrapure water was added dropwise with stirring, and the mixture was stirred at room temperature for 48 hours, whereupon no pale yellow solid precipitated from the reaction solution, and TLC (petroleum ether/ethyl acetate, 1/1) showed disappearance of 3 (RS-heptacyclic acetal). The reaction was terminated and the reaction solution was filtered to give 35mg (81%) of the title compound as a pale yellow solid. ESI-FT-ICR-MS (M/e):479.17224[ M-H]^-；

(c＝0.13,CH₃OH)；M.p.221～222℃；IR(cm^-1):3288.62,3062.04,2955.17,2900.75,1656.16,1453.90,1399.17,1369.86,1346.79,1299.72,1261.36,1203.55,1150.49,1117.04,1040.56,916.09,888.56,839.70,731.26,671.92；¹H NMR(800MHz,DMSO-d₆)δ/ppm＝11.06(s,1H),10.92(s,1H),9.78(t,J＝2.4Hz,1H),9.62(m,1H),7.49(d,J＝7.8Hz,1H),7.46(d,J＝7.8Hz,1H),7.38(d,J＝5.2Hz,1H),7.37(d,J＝5.2Hz,1H),7.10(t,J＝7.8Hz,2H),7.01(dd,J₁＝14.9Hz,J₂＝7.4Hz,2H),6.09(m,1H),5.48(m,1H),4.53(dd,J₁＝11.4Hz,J₂＝4.2Hz,1H),4.40(dd,J₁＝11.7Hz,J₂＝3.3Hz,1H),3.45(dd,J₁＝15.2Hz,J₂＝3.4Hz,1H),3.31(m,1H),3.17(m,1H),3.03(m,3H),2.86(m,2H)。

Example 5 evaluation of anti-arteriothrombotic Effect of RS-heptacyclic aldehyde

Experimental Material

Uratan (Ethyl carbamate, CAS:51-79-6, national pharmaceutical group Chemicals Co., Ltd.), Aspirin (CAS:50-78-2, Shanghai Allantin Biotech Co., Ltd.).

Laboratory animal

Male SD rats (200. + -.20 g) purchased from Experimental animals technology, Inc. of Weitongli, Beijing. The model is used for preparing a rat common carotid artery external jugular vein bypass circulation silk thread model during evaluation.

Dosage to be administered

The RS-heptacyclic aldehyde is suspended in 0.5 percent of CMC-Na, and the dosage is 0.1 mu mol/kg; the positive control aspirin was suspended in 0.5% CMC-Na at a dose of 167. mu. mol/kg; the negative control was 0.5% CMC-Na.

Experimental procedures

Rats were acclimatized and fasted for one day prior to surgery, and were gavaged at a dose of 0.3mL/100g body weight 30min prior to surgery using a double-blind sequential method. The polyethylene pipe is filled with physiological saline solution of heparin sodium, 20% of urethane solution is injected according to the weight of a rat to be completely anesthetized, the anesthetized rat is fixed on a rat board, the right common carotid artery is separated, an operation line is respectively penetrated into the proximal end and the distal end of the common artery, and the distal end is ligated. Then, the left external jugular vein of the rat is separated, and an operation line is respectively penetrated into the proximal end and the distal end of the vein, and the distal end is ligated. Carefully cutting an oblique opening on the exposed left external jugular vein, inserting the prepared tip of a circulation bypass pipeline into the oblique opening, fixing the vein and a polyethylene tube by using a surgical thread, pushing an accurate amount of heparin sodium physiological saline solution into the syringe through the other end, stopping bleeding by using an artery clamp at the proximal end of the right common carotid artery, cutting an oblique opening at a position not far away from the artery clamp, inserting the tip of the polyethylene tube into the proximal end of the artery oblique opening for fixing and loosening the artery clamp for circulation for 15 minutes, observing the breathing condition of a rat during circulation, and monitoring the blood circulation condition. After 15 minutes of circulation, the polyethylene tube at the venous end was cut open, the silk was removed with straight forceps, the filter paper was blotted to remove floating blood and weighed, and the wet weight of the thrombus was recorded. And counting the results by using a T test mode. The operation was performed alternately with four of each group. The experimental data are shown in table 1.

TABLE 1 treatment of arterial thrombotic heavies in rats

a) The ratio of P to CMC-Na is less than 0.01; n is 11.

The data in Table 1 show that RS-heptacyclaldehyde is effective in inhibiting arterial thrombosis in rats at an oral dose of 0.1. mu. mol/kg. Therefore, the invention has obvious technical effect.

Example 6 evaluation of anti-venous thrombosis Effect of RS-heptacyclaldehyde

Experimental Material

Uratan (Ethyl carbamate, CAS:51-79-6, national pharmaceutical group chemical Co., Ltd.), warfarin sodium (CAS:129-06-6, Bailingwei science and technology Co., Ltd.).

Laboratory animal

Male SD rats (200. + -.20 g) purchased from Experimental animals technology, Inc. of Weitongli, Beijing. For evaluation, the rat inferior vena cava ligation model was prepared.

Dosage to be administered

The RS-heptacyclic aldehyde is suspended in 0.5 percent of CMC-Na, and the dosage is 0.1 mu mol/kg; the positive control warfarin was suspended in 0.5% CMC-Na at a dose of 4.87. mu. mol/kg; the negative control was 0.5% CMC-Na.

Experimental procedures

Rats were acclimatized and fasted for one day prior to surgery and were gavaged at a dose of 0.3mL/100g body weight. Anesthesia was performed with a 20% urethane solution administered intraperitoneally 30 minutes after administration and 2 minutes before surgery. Fixing the rat on a rat fixing plate, preparing skin on the abdomen, sterilizing, opening the abdominal cavity along the leucorrhea line, and opening to expose one corner of the liver until the opening is about 4cm long. The organs such as small intestine in the abdominal cavity were removed and wrapped with gauze soaked with normal saline. Blunt separating connective tissues around blood vessels, exposing inferior vena cava and branches thereof, peeling off the abdominal aorta and the inferior vena cava below the left renal vein, ligating the inferior vena cava at the junction of the inferior vena cava and the left renal vein by using a suture soaked by normal saline, moving organs such as intestines back to the abdominal cavity according to anatomical positions, suturing the abdominal cavity layer by using the suture, and then placing the rat in an environment at 25-28 ℃ for circulation for 4 hours. Then, the patient is subjected to ether anesthesia, the abdominal cavity is opened, the branch vessels of the inferior vena cava are ligated one by one, the inferior vena cava with the length of 2cm is extracted from the ligation site at the junction of the inferior vena cava and the left renal vein, and the thrombus is extracted from the inferior vena cava. Blood was reserved as a sample for the determination of coagulation factor X, P-selectin and GPIIb/IIIa. The thrombus was weighed and the results were counted using the T-test. The operation was performed alternately with four of each group. The experimental data are shown in table 2.

TABLE 2 treatment of venous thrombosis in rats

a) Compared with 0.5 percent of CMC-Na, P is less than 0.01; n is 9

The data in Table 2 show that RS-heptacyclaldehyde effectively inhibits venous thrombosis in rats at an oral dose of 0.1. mu. mol/kg. Therefore, the invention has obvious technical effect.

Example 7 evaluation of bleeding side Risk of RS-heptacyclaldehyde

The international normalized ratio was automatically calculated by a semi-automatic coagulometer (model TS6000, MD Pacific). The specific test method is that 3.6 mL/rat whole blood collected in example 6 is placed in a centrifuge tube filled with 0.4mL sodium citrate solution (3.8%), centrifuged for 15 minutes at 2500g, then Platelet Poor Plasma (PPP) at the upper layer is sucked, 100 μ L PPP is added into a test cup, a test magnetic bead is added, a sample to be tested is pre-warmed in a pre-warm area of an instrument for 180 seconds, 200 μ L plasma prothrombin time test solution (kit goods number: 20-7011, MD Pacific) is added, the instrument starts to automatically detect to obtain the plasma prothrombin time, the international standard ratio is automatically calculated, the statistical result is carried out by using the T test mode, and the experimental data are shown in Table 3.

TABLE 3 International normalized ratio of treated rats

a) The ratio of P to CMC-Na is less than 0.01; n is 8

The data in Table 3 show that RS-heptacyclaldehyde effectively inhibits rat INR at an oral dose of 0.1. mu. mol/kg. The RS-heptacyclic aldehyde has no bleeding risk, and the invention has obvious technical effect.

Example 8 evaluation of the Effect of RS-heptacyclic aldehyde on P-selectin levels in the plasma of intravenous rats

Example 6 Whole blood of rats collected 4.0 mL/mouse was placed in a centrifuge tube containing 0.4mL of sodium citrate solution (3.8%), centrifuged at 1000g for 15 minutes, and then Platelet Poor Plasma (PPP) was aspirated. mu.L of each PPP sample and standard was added to each well of the test plate, incubated at 37 ℃ for 2h after plating with the cover plate membrane, then all the liquid in the plate was poured off, the plate was not washed, 100. mu.L of Biotin solution was added directly to each well, and incubated at 37 ℃ for 1 hour after plating with a new cover plate membrane. Then, the plate liquid was discarded, 200. mu.L of the washing solution was added to each well, and the washing solution was discarded after standing for 2min and the 96-well plate was patted dry, and this was repeated 3 times. Then 100. mu.L of HRP-avidin per well was added and incubated at 37 ℃ for another 1 hour after coating with a new coversheet membrane. After incubation, the plate washing operation was repeated 5 times to remove the liquid from each well. Adding 90 mu L of 3,3',5,5' -tetramethyl benzidine into each well, covering a new cover plate membrane, incubating for 20 minutes at 37 ℃ in the dark, adding 50 mu L of stop solution into each well, and rapidly reading the OD value of each well by using an enzyme-linked immunosorbent assay at the dual wavelengths of 450nm and 540 nm. And subtracting the OD value read at 540nm from the OD value read at 450nm, drawing a standard curve by taking the OD value as a vertical coordinate and the concentration of the P-selectin standard substance as a horizontal coordinate, and calculating the content of the P-selectin in the sample to be detected from the standard curve according to the OD value. The differences in P-selectin content of each group were analyzed using the one-way analysis of variance (ANOVA one-way, LSD) method in SPSS 22.0, and the experimental data are shown in Table 4.

TABLE 4 content of P-selectin in plasma of treated rats

a) The ratio of P to CMC-Na is less than 0.01; n is 5

The data in Table 4 show that RS-heptacyclaldehyde is effective in reducing plasma P-selectin levels in rats at an oral dose of 0.1. mu. mol/kg. Therefore, the invention has obvious technical effect.

Example 9 evaluation of the Effect of RS-heptacyclic aldehyde on the GPIIb/IIIa content in the plasma of phlebothrombotic rat

Example 6 Whole blood of rats collected 4.0 mL/mouse was placed in a centrifuge tube containing 0.4mL of sodium citrate solution (3.8%), centrifuged at 1000g for 15 minutes, and then Platelet Poor Plasma (PPP) was aspirated. mu.L of each PPP sample and standard was added to each well of the test plate, incubated at 37 ℃ for 2 hours after plating with the cover plate membrane, after which all the liquid in the plate was poured off, the plate was not washed, 100. mu.L of Biotin solution was added directly to each well, and incubated at 37 ℃ for 1 hour after plating with a new cover plate membrane. Then, the plate liquid was discarded, 200. mu.L of the washing solution was added to each well, and the washing solution was discarded after standing for 2 minutes and the 96-well plate was patted dry, and this was repeated 3 times. Then 100. mu.L of HRP-avidin was added to each well and coated with a new cover film and incubated at 37 ℃ for another 1 hour. After incubation, the plate washing operation was repeated 5 times to remove the liquid from each well. mu.L of 3,3',5,5' -tetramethylbenzidine was added to each well, covered with a new cover film and incubated at 37 ℃ for 20 minutes in the dark. Then 50 mul of stop solution is added into each hole, and the OD value of each hole is rapidly read by using a microplate reader under the dual-wavelength of 450nm and 540 nm. And subtracting the OD value read at 540nm from the OD value read at 450nm, drawing a standard curve by taking the OD value as a vertical coordinate and the concentration of the GPIIb/IIIa standard substance as a horizontal coordinate, and calculating the content of GPIIb/IIIa in the sample to be detected according to the OD value of the GPIIb/IIIa standard substance from the standard curve. The difference of GPIIb/IIIa content of each group is analyzed by using a single-factor analysis of variance (ANOVA one-way, LSD) method in SPSS 22.0, and the experimental data are shown in Table 5.

TABLE 5 GPIIb/IIIa content in treated rat plasma

a) The ratio of P to CMC-Na is less than 0.01; n is 5

The data in Table 5 show that RS-heptacyclic aldehyde is effective in reducing plasma GPIIb/IIIa in rats at 0.1. mu. mol/kg oral dose. Therefore, the invention has obvious technical effect.

Example 10 evaluation of the Effect of RS-heptacyclaldehyde on the level of blood coagulation factor X in the plasma of phlebothrombotic rat

Example 6 Whole blood of rats collected 4.0 mL/mouse was placed in a centrifuge tube containing 0.4mL of sodium citrate solution (3.8%), centrifuged at 1000g for 15 minutes, and the supernatant Platelet Poor Plasma (PPP) was aspirated. The method comprises the following steps of taking a PPP sample of 6 rats after administration for each experimental group, diluting the PPP sample by 200 times with a sample diluent in a kit, adding 50 mu L of a sample to be detected or a standard substance into each hole in a test plate, simultaneously adding 50 mu L of HRP-avidin into each hole, covering a cover plate membrane, incubating at 37 ℃ for 1 hour, then discarding liquid in the plate, adding 200 mu L of washing liquid into each hole, standing for 2 minutes, discarding the washing liquid, drying a 96-hole plate, and repeating the steps for 5 times. Then, 90. mu.L of 3,3',5,5' -tetramethylbenzidine is added into each well, a new cover plate membrane is covered on the well, the well is incubated for 20 minutes in the dark at 37 ℃, 50. mu.L of stop solution is added into each well after incubation, and the OD value of each well is rapidly read by using an enzyme-linked immunosorbent assay at the dual-wavelength of 450nm and 540 nm. And subtracting the OD value read at 540nm from the OD value read at 450nm, drawing a standard curve by taking the OD value as a vertical coordinate and the FXa standard substance concentration as a horizontal coordinate, and calculating the FXa content in the sample to be detected from the standard curve according to the OD value. The difference of FXa content of each group was analyzed by the method of T Test (T-Test) in SPSS 22.0, and the experimental data are shown in FIG. 6.

TABLE 6 treatment of FXa content in rat plasma

a) The ratio P to 0.5 percent CMC-Na is less than 0.01, and the ratio P to warfarin is more than 0.05; n is 5

The data in Table 6 show that RS-heptacyclic aldehyde is effective in reducing plasma FXa levels in rats at 0.1. mu. mol/kg oral dose, which is not significantly different from warfarin group at 4.87. mu. mol/kg oral dose, indicating that RS-heptacyclic aldehyde is at least 48.7 times as active as warfarin in reducing plasma FXa levels in rats. Therefore, the invention has obvious technical effect.

It should be noted that the above summary and the detailed description are intended to demonstrate the practical application of the technical solutions provided by the present invention, and should not be construed as limiting the scope of the present invention. Various modifications, equivalent substitutions, or improvements may be made by those skilled in the art within the spirit and principles of the invention.

Claims

1. An RS-heptacyclic aldehyde compound having the formula:

2. the method of synthesizing an RS-heptacyclic aldehyde compound according to claim 1, comprising the steps of:

1) carrying out Picture t-Spengler condensation on L-tryptophan benzyl ester and 1,1,3, 3-tetramethoxypropane to generate (3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro-beta-carboline-3-benzyl carboxylate;

3) condensation of (1R,3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro- β -carboline-3-carboxylic acid with (1S,3S) -1- (2, 2-dimethoxyethyl-2-yl) -2,3,4, 9-tetrahydro- β -carboline-3-carboxylic acid to the RS-heptacyclic aldehyde precursor RS-heptacyclic acetal according to claim 1;

4) the conversion of the RS-heptacyclic acetal to the RS-heptacyclic aldehyde compound of claim 1.

3. Use of the RS-heptacyclic aldehyde compound of claim 1 in the preparation of an anti-arterial thrombosis medicament.

4. Use of the RS-heptacyclic aldehyde compound of claim 1 in the preparation of an anti-venous thrombosis medicament.

5. Use of the RS-heptacyclic aldehyde compound of claim 1 for the preparation of a factor X antagonist.

6. Use of the RS-heptacyclic aldehyde compound of claim 1 for the preparation of a P-selectin antagonist.

7. Use of the heptacyclic aldehyde compound of claim 1 for the preparation of a GPIIb/IIIa antagonist.