CN114751859A

CN114751859A - Cyclopentapyridine compound and preparation method and application thereof

Info

Publication number: CN114751859A
Application number: CN202210308419.8A
Authority: CN
Inventors: 王跃飞; 张潇予; 柴欣; 董雪媛; 孟宁; 崔英; 于海洋; 于卉娟; 张瑞虎
Original assignee: Tianjin University of Traditional Chinese Medicine
Current assignee: Tianjin University of Traditional Chinese Medicine
Priority date: 2022-03-23
Filing date: 2022-03-23
Publication date: 2022-07-15
Anticipated expiration: 2042-03-23
Also published as: CN114751859B

Abstract

The invention belongs to the technical field of chemical drugs, and particularly relates to a cyclopentapyridine compound as well as a preparation method and application thereof. The cyclopentapyridine compound is obtained by taking genipin and methylamine as raw materials, reacting under specific conditions, and then separating by column chromatography and liquid chromatography. Experiments prove that the compound has a remarkable inhibiting effect on proliferation of colon cancer cells and can be used for preparing antitumor drugs.

Description

Cyclopentapyridine compound and preparation method and application thereof

Technical Field

The invention belongs to the technical field of chemical drugs, and particularly relates to a cyclopentapyridine compound as well as a preparation method and application thereof.

Background

Colon cancer is a common malignant tumor which is primarily generated on the mucous epithelium of the colon, and the incidence rate of the colon cancer tends to rise year by year in China. The early stage of the colon cancer has no symptoms, the prognosis effect is good after treatment, and the patient has a longer survival period. After the colon cancer develops to the middle and late stage, symptoms such as hematochezia, abdominal pain and the like appear, and the cure rate and the survival time of the patient are obviously reduced. At present, the operation is a main method for treating colon cancer, and can completely cure part of patients or prolong the life cycle of the patients by combining radiotherapy, chemotherapy and targeted therapy.

At present, few medicines capable of effectively treating colon cancer have side effects of different degrees. Therefore, obtaining more compounds capable of being used for treating colon cancer has great significance for drug development and clinical treatment of colon cancer.

Disclosure of Invention

Aiming at the problems, the invention provides a cyclopentapyridine compound, a preparation method and an application thereof, wherein the compound can inhibit colon cancer cell proliferation and can be used for preparing antitumor drugs.

In order to achieve the purpose of the invention, the embodiment of the invention adopts the following technical scheme:

in a first aspect, an embodiment of the present invention provides a cyclopentapyridine compound, which has a structural formula shown in formula I:

experiments prove that the compound has a remarkable inhibiting effect on the proliferation of colon cancer cells. Reports related to the cyclopentapyridine compound are not found at present.

The invention also provides a preparation method of the cyclopentapyridine compound, which comprises the following steps:

a, dissolving genipin and methylamine in a non-nitrogen-containing buffer solution with the pH value of 6-8, and reacting at 30-80 ℃ for more than 4 hours to obtain a reaction solution;

and step B, adding the reaction solution into a small-pore resin chromatographic column, performing gradient elution by sequentially adopting ethanol-water solvents with volume ratios of 0:10, 1:9, 2:8, 3:7, 4:6, 5:5 and 6:4, and collecting eluent obtained by elution of the ethanol-water solvents with volume ratios of 5: 5-6: 4 to obtain the cyclopentapyridine compound.

The method comprises the steps of reacting genipin with methylamine in a specific solution at a specific temperature to obtain a reaction solution containing the cyclopentapyridine compound, and then separating the cyclopentapyridine compound from other compounds in the reaction solution through small-pore resin column chromatography to obtain the cyclopentapyridine compound in an eluent.

In combination with the first aspect, the preparation method may further include separating the eluate by preparative liquid chromatography, collecting the eluate fraction according to the peak appearance time of the chromatographic peak of the cyclopentapyridyl compound, concentrating, and drying to obtain a pure product of the cyclopentapyridyl compound in a specific eluate fraction.

Preferably, the chromatographic conditions of the preparative liquid chromatography are: the chromatographic column is a carbon-eighteen bonded silica gel chromatographic column, the mobile phase is 0.05-0.20% (v/v) formic acid water-methanol with the volume ratio of (40:60) - (60:40), and isocratic elution is carried out.

Under the chromatographic condition, the chromatographic column can be selected from Waters

C18 OBD^TMPrep Column with specification of 10 × 250mm and 5 μm; the mobile phase can be 0.1% (v/v) formic acid water-methanol with a volume ratio of 50: 50; the flow rate of isocratic elution can be selected from 4 mL/min; the sample size may be 2 mL.

With reference to the first aspect, the buffer solution is a potassium dihydrogen phosphate-sodium hydroxide buffer solution, a PBS buffer solution, or a sodium acetate-glacial acetic acid buffer solution.

In combination with the first aspect, the pH of the buffer solution is preferably 6.8.

In combination with the first aspect, the temperature of the reaction in step A is preferably 45 to 55 ℃.

In connection with the first aspect, the amount of each elution solvent used in step B may be selected from 3 column volumes to ensure separation of the target compound from other compounds and to obtain a better enrichment of the target compound.

In a second aspect, the embodiment of the invention also provides an application of the cyclopentapyridine compound in preparation of antitumor drugs.

Experiments prove that the compound can obviously inhibit the proliferation activity (P <0.05) of SW480 cells of a human colon cancer cell line and CT26 cells of a mouse colon cancer cell line, so that the compound can be applied to preparing antitumor drugs.

In combination with the second aspect, the anti-tumor drug is an anti-colon cancer drug.

In combination with the second aspect, the antitumor drug is a targeted preparation against colon cancer. The compound is prepared into a targeted preparation, so that the drug effect can be improved, and the influence on other tissues can be reduced.

Drawings

FIG. 1 is a drawing showing Genimethylamine B in example 1 of the present invention ¹H-NMR spectrum;

FIG. 2 is a drawing showing Genimethylamine B in example 1 of the present invention¹³C-NMR spectrum;

FIG. 3 is a drawing showing Genimethylamine B in example 1 of the present invention¹H-¹H COSY spectrogram;

FIG. 4 is the HSQC spectrum of Genimethylamine B in example 1 of the present invention;

FIG. 5 is HMBC chromatogram of Genimethylamine B in example 1 of the present invention;

FIG. 6 shows that Genimethylamine B inhibits the cell proliferation activity of the human colorectal cancer cell line SW480 in vitro in example 8 according to the present invention;

FIG. 7 shows that Genimethylamine B inhibits the cell proliferation activity of the mouse colorectal cancer cell line CT26 in vitro in example 8.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail with reference to the following embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

The incidence rate of colon cancer is high, the cure rate and survival rate of the middle and late stages are reduced, and the number of the medicines which can effectively treat colon cancer is small at present. The embodiment of the invention provides a cyclopentapyridine compound (named Genimethylamine B), which has a structural formula shown in a formula I:

experiments show that Genimethylamine B can inhibit the proliferation activity of SW480 cells of a human colon cancer cell line and CT26 cells of a mouse colon cancer cell line (P <0.05), and has important significance for treating colon cancer.

The embodiment of the invention also provides a preparation method of the compound, which comprises the following steps:

and step B, adding the reaction solution into a small-pore resin chromatographic column, performing gradient elution by sequentially adopting ethanol-water solvents with volume ratios of 0:10, 1:9, 2:8, 3:7, 4:6, 5:5 and 6:4, and collecting eluent obtained by elution of the ethanol-water solvents with the volume ratios of 5: 5-6: 4 to obtain Genimethylamine B in the eluent.

In the preparation method, the eluent obtained in the step B can be further purified by preparative liquid chromatography separation so as to obtain a pure product of Genimethylamine B. The purification method can select the following operations: separating the eluent obtained in the step B by preparative liquid chromatography, wherein the chromatographic column is a carbon-eighteen bonded silica gel chromatographic column, and the mobile phase is 0.05-0.20% (v/v) of formic acid water-methanol in a volume ratio of (40:60) - (60:40), and the likeAnd (5) eluting at a certain degree. Wherein the chromatographic column can be selected from Waters

C18OBD^TMPrep Column with specification of 10 × 250mm and 5 μm; the mobile phase can select 0.1% (v/v) formic acid water-methanol with the volume ratio of 50: 50; the flow rate for isocratic elution may be selected to be 4 mL/min. Under the condition, collecting the elution fraction for 12.0-12.5 min according to the peak-appearing time, concentrating and drying to obtain the pure Genimethylamine B product.

In the preparation method, the mass ratio of genipin to methylamine is not limited, and the cyclopentapyridine compound can be prepared by the preparation method under any mass ratio. However, the preparation method does not limit the mass ratio of the raw materials, and does not represent that genipin and methylamine can completely participate in the reaction at any ratio, and the raw materials with large feeding amount can be remained in the reaction process. In order to ensure the yield of the product and reduce the waste of raw materials, the mass ratio of genipin to methylamine is preferably 8-12: 3, for example 10: 3.

The type of the buffer solution is not limited in the invention, and a non-nitrogen-containing common buffer solution can be used, such as potassium dihydrogen phosphate-sodium hydroxide buffer solution, PBS buffer solution or sodium acetate-glacial acetic acid buffer solution.

The pH of the buffer solution is preferably 6.8. Under otherwise identical conditions, the yield of the cyclopentapyridine compound can be improved when the pH is 6.8.

The reaction temperature of the preparation method is preferably 45-55 ℃, and the reaction time is 48 h. Under the preferable raw material mass ratio, reaction temperature, pH value and reaction time, more target products Genimethylamine B can be generated.

During the elution of the small-pore resin column chromatography, the amount of each elution solvent is 3 column volumes, so as to improve the separation effect of the Genimethylamine B from other compounds and enrich the Genimethylamine B. The small-pore resin chromatographic column can use CHP20/P120 small-pore resin as packing.

The embodiment of the present invention will be described below with reference to specific examples.

Example 1

The embodiment provides a cyclopentapyridine compound Genimethylamine B and a preparation method thereof.

The preparation method of the Genimethylamine B specifically comprises the following steps:

step A, precisely weighing 150mg of genipin and 45mg of methylamine respectively, placing the genipin and the methylamine respectively into 150mL beakers, and adding potassium dihydrogen phosphate-sodium hydroxide buffer solutions with the pH value of 6.8 respectively to fully dissolve the potassium dihydrogen phosphate-sodium hydroxide buffer solutions to prepare stock solutions with the concentration of 4.45mmol/L respectively; respectively transferring 150mL of the stock solutions, placing the stock solutions into a beaker, adding a potassium dihydrogen phosphate-sodium hydroxide buffer solution with the pH value of 6.8 to 900mL, shaking up, placing the mixture into a water bath at 50 ℃, reacting for 48 hours, and taking out the mixture to obtain a reaction solution;

step B, separating the reaction solution by using CHP20/P120 small-pore resin through column chromatography, performing gradient elution by using ethanol-water (0:10, 1:9, 2:8, 3:7, 4:6, 5:5 and 6:4) solvent systems (each elution system elutes 3 column volumes) to obtain a series of eluents, combining the eluents of ethanol-water (5: 5-6: 4), and concentrating for separation and purification of the preparative liquid chromatography;

Step C, taking a concentrated solution of an ethanol-water (5: 5-6: 4) eluent, and separating and purifying by adopting preparative liquid chromatography, wherein a chromatographic column is Waters

C18OBD^TMPrep Column (10 × 250mm, 5 μm), mobile phase 0.1% (v/v) formic acid-methanol (50:50), isocratic elution at a flow rate of 4mL/min, detection wavelength 290nm, sample introduction 2mL, collection of 12.0-12.5 min elution fraction according to peak time, concentration, and drying to obtain genimepimine B with a yield of 0.54% (yield: weight of genimepamine B ÷ total weight of genipin and methylamine × 100%, the same below), and purity of 99.2% (purity: fraction collected from preparation liquid phase is detected in hplc, peak area of genimepamine B ÷ sum of all hplc × 100%, the same below).

By NMR¹H-NMR、¹³C-NMR、¹H-¹H COSY, HSQC and HMBC) technology for representing the chemical structure of Genimethylamine B,¹the H-NMR spectrum is shown in figure 1,¹³the C-NMR spectrum is shown in figure 2,¹H-¹the H COSY spectrum is shown in figure 3, the HSQC spectrum is shown in figure 4, and the HMBC spectrum is shown in figure 5.

¹The H-NMR spectrum has 7 groups of proton signals, including 4 groups of olefin proton signals at delta 7.00, 7.83, 8.22 and 9.00.¹³The C NMR spectrum showed 12 carbon signals, with δ 167.0 being the estercarbonyl carbon signal and 8 olefinic carbons at δ 133.2, 116.5, 135.2, 109.0, 143.3, 127.4, 120.9, 136.5. In HSQC spectra, 7 carbons were shown to correlate with the presence of proton signals, with δ 52.9 methoxy carbon and δ 46.6 aminomethyl carbon correlating with δ 3.94 and 4.12 proton signals, respectively, δ 184.3 aldehyde carbon correlating with δ 9.81 proton signals, and δ 133.2(C-3), 136.5(C-1), 143.3(C-7), 109(C-6)4 olefin carbons correlating with δ 8.22, 9.00, 7.83, 7.00 proton signals, respectively. ¹H-¹H COSY spectra show that there are correlations between H-6 and H-7, H-3 and H-1, and nitrogen methyl hydrogen atoms and H-3, H-1. In HMBC spectrum, the substitution position of nitrogen methyl can be determined by the correlation signal between nitrogen methyl carbon and H-1 and H-3, the position of aldehyde group can be determined by the correlation signal between aldehyde hydrogen and C-8 and C-9, and the information related to signals of H-3, carbonyl carbon, H-7 and C-5 is also displayed.

According to the NMR spectrum, the structural formula of the compound is shown as the formula I:

example 2

This example provides a preparation method of cyclopentapyridine compound genimethylethylamine B.

The method specifically comprises the following steps:

step A, respectively taking 120mg of genipin and 45mg of methylamine, precisely weighing, respectively placing the genipin and the methylamine into 150mL beakers, respectively adding potassium dihydrogen phosphate-sodium hydroxide buffer solutions with the pH value of 6.8 to fully dissolve the potassium dihydrogen phosphate-sodium hydroxide buffer solutions to prepare stock solutions with the concentrations of 3.5 and 4.4mmol/L respectively; respectively transferring 150mL of the stock solutions, placing the stock solutions into a beaker, adding a potassium dihydrogen phosphate-sodium hydroxide buffer solution with the pH value of 6.8 to 900mL, shaking up, placing the mixture into a water bath at 45 ℃, reacting for 60 hours, and taking out the mixture to obtain a reaction solution;

C18OBD^TMPrep Column (10 × 250mm, 5 μm), 0.2% (v/v) formic acid-methanol (60:40) as mobile phase, isocratically eluting at a flow rate of 4mL/min, with a detection wavelength of 290nm and a sample volume of 2mL, collecting the eluate fraction of 12.5-13.5 min according to the peak-out time, concentrating, and drying to obtain Genimethylamine B with a yield of 0.36% and a purity of 80.5%.

Example 3

The method specifically comprises the following steps:

step A, precisely weighing 180mg of genipin and 45mg of methylamine respectively, placing the genipin and the methylamine respectively into 150mL beakers, and adding potassium dihydrogen phosphate-sodium hydroxide buffer solutions with the pH value of 6.8 respectively to fully dissolve the potassium dihydrogen phosphate-sodium hydroxide buffer solutions to prepare stock solutions with the concentrations of 5.3 and 4.4mmol/L respectively; respectively transferring 150mL of the stock solutions, placing the stock solutions into a beaker, adding a potassium dihydrogen phosphate-sodium hydroxide buffer solution with the pH value of 6.8 to 900mL, shaking up, placing the mixture into a water bath at 55 ℃, reacting for 36h, and taking out the mixture to obtain a reaction solution;

C18OBD^TMPrep Column (10 × 250mm, 5 μm), 0.05% (v/v) formic acid-methanol (40:60) as mobile phase, isocratically eluting at a flow rate of 4mL/min, with a detection wavelength of 290nm and a sample volume of 2mL, collecting 10.5-12 min of elution fraction according to peak-out time, concentrating, and drying to obtain Genimethylamine B with a yield of 0.41% and a purity of 76.2%.

Example 4

step A, precisely weighing 150mg of genipin and 45mg of methylamine respectively, placing the genipin and the methylamine respectively into 150mL beakers, and adding PBS buffer solution with the pH value of 7.35 respectively to fully dissolve the genipin and the methylamine so as to prepare stock solutions with the concentrations of 4.4mmol/L respectively; respectively transferring 150mL of the stock solutions, placing the stock solutions into a beaker, adding a PBS (phosphate buffer solution) with the pH value of 7.35 to 900mL, shaking up, placing the mixture into a water bath at 50 ℃, reacting for 48 hours, and taking out the mixture to obtain a reaction solution;

C18 OBD^TMPrep Column (10 × 250mm, 5 μm), 0.1% (v/v) formic acid water-methanol (50:50) as a mobile phase, isocratic elution at a flow rate of 4mL/min, a detection wavelength of 290nm and a sample size of 2mL, collecting an elution fraction of 12.0-12.5 min according to a peak time, concentrating and drying to obtain Genimethylamine B, wherein the yield is 0.48% and the purity is 97.6%.

Example 5

This example provides a cyclopentapyridine compound genimethylethylamine B and a method for preparing the same.

step A, precisely weighing 150mg of genipin and 45mg of methylamine respectively, placing the genipin and the methylamine into 150mL beakers respectively, and adding sodium acetate-glacial acetic acid buffer solutions with the pH value of 6.2 respectively to fully dissolve the sodium acetate-glacial acetic acid buffer solutions to prepare stock solutions with the concentrations of 4.4mmol/L respectively; respectively transferring 150mL of the stock solutions, placing the stock solutions into a beaker, adding a sodium acetate-glacial acetic acid buffer solution with the pH value of 6.2 to 900mL, shaking up, placing the mixture into a water bath at 50 ℃, reacting for 48 hours, and taking out the mixture to obtain a reaction solution;

step B, separating the reaction solution by using CHP20/P120 small-hole resin through column chromatography, performing gradient elution by using ethanol-water (0:10, 1:9, 2:8, 3:7, 4:6, 5:5 and 6:4) solvent systems (each elution system elutes 3 column volumes) to obtain a series of eluents, combining the eluents of ethanol-water (5: 5-6: 4), and concentrating for separation and purification of the preparative liquid chromatography;

C18OBD^TMPrep Column (10 × 250mm, 5 μm), 0.1% (v/v) formic acid water-methanol (50:50) as mobile phase, isocratic elution at flow rate of 4mL/min, detection wavelength of 290nm, sample injection amount of 2mL, collecting elution fraction of 12.0-12.5 min according to peak time, concentrating, drying to obtain Genimethylamine B with yield of 0.49%,the purity was 98.3%.

Example 6

step A, precisely weighing 150mg of genipin and 45mg of methylamine respectively, placing the genipin and the methylamine into 150mL beakers respectively, and adding sodium acetate-glacial acetic acid buffer solutions with the pH value of 6.0 respectively to fully dissolve the sodium acetate-glacial acetic acid buffer solutions to prepare stock solutions with the concentrations of 4.4mmol/L respectively; respectively transferring 150mL of the stock solutions, placing the stock solutions into a beaker, adding a sodium acetate-glacial acetic acid buffer solution with the pH value of 6.0 to 900mL, shaking up, placing the mixture into a water bath at 50 ℃, reacting for 48 hours, and taking out the mixture to obtain a reaction solution;

C18OBD^TMPrep Column (10 × 250mm, 5 μm), 0.1% (v/v) formic acid-methanol (50:50) as a mobile phase, isocratically eluting at a flow rate of 4mL/min, wherein the detection wavelength is 290nm, the sample volume is 2mL, eluting fractions of 12.0-12.5 min are collected according to the peak-out time, and the eluting fractions are concentrated and dried to obtain Genimethylamine B, wherein the yield is 0.47% and the purity is 99.0%.

Example 7

step A, precisely weighing 150mg of genipin and 45mg of methylamine respectively, placing the genipin and the methylamine respectively into 150mL beakers, and adding potassium dihydrogen phosphate-sodium hydroxide buffer solutions with the pH value of 8.0 respectively to fully dissolve the potassium dihydrogen phosphate-sodium hydroxide buffer solutions to prepare stock solutions with the concentration of 4.4mmol/L respectively; respectively transferring 150mL of the stock solutions, placing the stock solutions into a beaker, adding a potassium dihydrogen phosphate-sodium hydroxide buffer solution with the pH value of 8.0 to 900mL, shaking up, placing the mixture into a water bath at 50 ℃, reacting for 48 hours, and taking out the mixture to obtain a reaction solution;

C18OBD^TMPrep Column (10 × 250mm, 5 μm), 0.1% (v/v) formic acid-methanol (50:50) as a mobile phase, isocratically eluting at a flow rate of 4mL/min, wherein the detection wavelength is 290nm, the sample volume is 2mL, eluting fractions of 12.0-12.5 min are collected according to the peak-out time, and the eluting fractions are concentrated and dried to obtain Genimethylamine B, wherein the yield is 0.31% and the purity is 98.3%.

Example 8

3- (4, 5-dimethyl-2-thiazole) -2,5 diphenyl tetrazole bromide (MTT) can be reduced by dehydrogenase in mitochondria of living cells to generate a dark purple crystalline product formazan. The faster the cell proliferated, the higher the dehydrogenase activity and the more formazan was formed. Formazan can be completely dissolved into blue by DMSO, has strong absorption peak at 570nm wavelength, and can be used for evaluating living cell number and proliferation activity by detecting absorbance value at 570nm wavelength by enzyme labeling instrument. This example demonstrates the effect of genimethylethylamine B on inhibiting colon cancer cell proliferation activity by measuring cell viability using MTT colorimetry.

1. Reagent, instrument and consumable

(1) Reagent: MTT, Solarbio; DMSO, Solarbio; PBS buffer, Bai Si Nuo (Tianjin) Biotech limited.

(2) Instrument, consumable: model 450 microplate reader, shanghai burle corporation; 96-well plate, Nest.

2. Experimental method

Preparation of MTT working solution: adding 25mg MTT powder into 5mL PBS buffer solution to prepare 5mg/mL solution, subpackaging, and storing at-20 ℃ in a dark place.

The colon cancer cell line SW480 in the logarithmic growth phase was selected, the cell mass was digested into single cells with 0.25% (g/mL) of trypsin, digestion was stopped by adding three volumes of DMEM complete medium, centrifugation was performed at 1000rpm for 5min, and the supernatant was discarded. 5mL of DMEM complete medium is added, 10 mu L of cell suspension is sucked after uniform mixing, and cell liquid is injected into a counting plate along the edge of a cover slip to count the cells. Preparation of 4X 10 Using DMEM complete Medium⁴Cell suspension/mL, seeded in 96-well plates at 200. mu.L per well and placed in CO₂Culturing in an incubator for 24 h. Removing culture solution from a 96-well plate, and adding Genimethylamine B solutions with the concentrations of 10, 20, 30, 40 and 50 mu M respectively diluted by DMEM complete culture medium, wherein each concentration is provided with 10 multiple wells; meanwhile, a blank control group was set up. Continuously placing in CO₂After 24h incubation in an incubator the test drug was removed, 100. mu.L/well MTT solution (dissolved in DMEM complete medium at a concentration of 0.5mg/mL) was added and after incubation at 37 ℃ for 4h the crystalline dark purple product was observed microscopically, 100. mu.L/well DMSO was added to dissolve the dark purple crystals and the absorbance value (A) was measured at 570 nm.

The colon cancer cell line CT26 was treated in the same manner and prepared in RPMI 1640 complete medium at 4X 10⁴Cell suspension of cell/mL, seeded in 96-well plates at 200. mu.L per well and placed in CO₂Culturing in an incubator for 24 h. Removing culture solution from a 96-well plate, adding Genimethylamine B solution with concentration gradients of 5, 10, 15, 20, 25 and 30 mu M diluted by RPMI 1640 complete culture medium, and setting 10 multiple wells for each concentration; meanwhile, a blank control group was established. Continuously placing in CO₂After 24h incubation in the incubator, the test drug was removed and 100. mu.L/well MTT solution (RPMI) was added1640 complete medium dissolution, concentration 0.5mg/mL), incubation at 37 ℃ for 4h and microscopic observation of the crystalline dark purple product, adding 100. mu.L/well DMSO to dissolve the dark purple crystals, and detecting the absorbance value (A) at a wavelength of 570 nm.

The cell viability of colon cancer cells was calculated as follows:

cell viability/% ═ a_{Testing drugs}/A_{Blank space}×100％

3. Statistical analysis

The study data were analyzed using Graphpad Prism 8 statistical software. The difference between the blank control group and the experimental group was analyzed by one-way ANOVA.

4. Results of the experiment

The results show that: compared with a blank control group, the Genimethylamine B obviously inhibits the proliferation activity of SW480 cells (P <0.05) of the human colon cancer cell line and CT26 cells (P <0.001) of the mouse colon cancer cell line, and has a certain dose-dependent relationship, and the results are shown in FIGS. 6 and 7. In the figure, C is a blank control group; p <0.05, P < 0.001.

The above description is intended to be illustrative of the preferred embodiment of the present invention and should not be taken as limiting the invention, but rather, the invention is intended to cover all modifications, equivalents, and alternatives falling within the spirit and scope of the invention.

Claims

1. A cyclopentapyridine compound is characterized in that the structural formula is shown as formula I:

2. a method for preparing the cyclopentapyridine compound of claim 1, comprising the steps of:

and step B, adding the reaction solution into a small-pore resin chromatographic column, performing gradient elution by sequentially adopting ethanol-water solvents with volume ratios of 0:10, 1:9, 2:8, 3:7, 4:6, 5:5 and 6:4, and collecting eluent obtained by elution of the ethanol-water solvents with the volume ratios of 5: 5-6: 4 to obtain the cyclopentapyridine compound.

3. The method according to claim 2, wherein the method further comprises separating the eluate by preparative liquid chromatography, collecting the eluate fraction according to the peak appearance time of the cyclopentapyridyl compound, concentrating, and drying to obtain the pure cyclopentapyridyl compound.

4. The method for preparing cyclopentapyridines according to claim 3, wherein the chromatographic conditions of the preparative liquid chromatography are: the chromatographic column is a carbon-eighteen bonded silica gel chromatographic column, the mobile phase is 0.05-0.20% (v/v) formic acid water-methanol with the volume ratio of (40:60) - (60:40), and isocratic elution is carried out.

5. The method of claim 2, wherein the chromatography column is Waters

C18 OBD^TMPrep Column with specification of 10 × 250mm and 5 μm; and/or

The mobile phase is 0.1% (v/v) formic acid water-methanol with a volume ratio of 50: 50; and/or

Isocratic elution is carried out at the flow rate of 4 mL/min; and/or

The sample volume was 2 mL.

6. The method for preparing a cyclopentapyridine compound of claim 2, wherein the buffer solution is potassium dihydrogen phosphate-sodium hydroxide buffer solution, PBS buffer solution, or sodium acetate-glacial acetic acid buffer solution; and/or

The pH of the buffer solution was 6.8.

7. The method for preparing cyclopentapyridine compounds of claim 2, wherein the reaction temperature in step a is 45-55 ℃; and/or

The amount of each eluting solvent used in step B was 3 column volumes.

8. The use of the cyclopentapyridines of claim 1 in the preparation of an anti-tumor medicament.

9. The use of claim 8, wherein the anti-neoplastic drug is an anti-colon cancer drug.

10. The use of claim 9, wherein the anti-neoplastic drug is a targeted agent against colon cancer.