CN109053732B

CN109053732B - Para-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative and preparation method and application thereof

Info

Publication number: CN109053732B
Application number: CN201811172929.7A
Authority: CN
Inventors: 王玮; 邓莉平; 吕峰平; 杜轶君
Original assignee: Shaoxing University Yuanpei College
Current assignee: Wuyuan Bencao (Shandong) Health Technology Co.,Ltd.
Priority date: 2018-10-09
Filing date: 2018-10-09
Publication date: 2020-06-23
Anticipated expiration: 2038-10-09
Also published as: CN109053732A

Abstract

The invention discloses a p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative and a preparation method and application thereof, wherein a pyrazole ring is introduced into an (E) -7-p-fluorobenzylidene-6, 7-indolizine-8 (5H) -one structure by using a1, 3-dipolar cycloaddition method, so that the p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative is synthesized. The p-fluoro substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative prepared by the invention has a strong tumor cell inhibition effect, and provides a basis for further application in the medical field.

Description

Para-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative and preparation method and application thereof

Technical Field

The invention belongs to the technical field of medicines, and particularly relates to a p-fluoro substituted pyridazinone-structure-containing spiro [ indolizine-pyrazoline ] derivative, and a preparation method and application thereof.

Background

(E) -7-p-fluorobenzylidene-6, 7-indolizin-8 (5H) -one having the following chemical formula:

wherein Ar is 4-FC₆H₄-。

The 1, 3-dipolar cycloaddition reaction is the most important method for synthesizing five-membered heterocyclic compounds with good regioselectivity and body selectivity, and is also a more active reaction in heterocyclic pharmaceutical chemistry research. In recent years, pyridazinone compounds become hot spots of research in the pharmaceutical field due to the unique physiological activity of pyridazinone compounds, and have strong effects of inhibiting cancer cell diffusion, resisting bacteria and viruses, reducing blood fat, reducing blood sugar, promoting metabolism and the like. Therefore, the heterocyclic compound has high synthetic value in terms of pharmacology and synthesis angle.

Pyrazole derivatives have attracted considerable attention as a class of useful intermediates and as a variety of pharmaceutical activities that they themselves exhibit. The indolizine skeleton is a common structural unit in bioactive alkaloids, and has remarkable anti-mitotic property or the effect of treating cardiovascular diseases and hypertension. Therefore, it is of great significance to research the influence of different heterocycles on pharmacological activity caused by aggregation in the same molecule.

Disclosure of Invention

The invention aims to provide a p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative, a preparation method and application thereof, and the p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative is prepared.

In order to achieve the purpose, the technical scheme of the invention is as follows:

a p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative has a chemical structural formula as follows:

wherein Ar is 4-FC₆H₄-。

The invention also provides a preparation method of the p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative, which comprises the following steps:

step 1, synthesizing a chromone aldehyde hydrazone derivative containing a pyridazinone structure;

step 2, synthesizing α -chloropyridazinone structure chromone aldehyde hydrazone derivatives;

and 3, adding (E) -7-p-fluorobenzylidene-6, 7-indolizinyl-8 (5H) -ketone and the chromone aldehyde hydrazone derivative with the α -chloropyridazinone structure into dichloromethane, dropwise adding a mixed solution of triethylamine and dichloromethane into the solution by using a constant-pressure dropping funnel, continuously stirring at normal temperature for 20-30 hours after the dropwise adding is finished, tracking by TLC (thin layer chromatography), removing the solvent by rotary evaporation after the reaction is completed, dissolving the residue left after the rotary evaporation, and performing chromatographic separation by using a silica gel column to obtain the spiro [ indolizine-pyrazoline ] derivative with the p-fluoro substituted pyridazinone structure.

Further, the mass ratio of the α -chloropyridazinone structure chromone aldehyde hydrazone derivative to the (E) -7-p-fluorobenzylidene-6, 7-indolizine-8 (5H) -one is 1.6: 1-2: 1.

Further, the volume ratio of triethylamine to dichloromethane is: 1: 10-1: 20.

further, the spreading agent adopted in the silica gel column chromatography separation is V_{(Ethyl acetate)}:V_{(Petroleum ether)}＝1:5。

The invention also provides application of the p-fluoro substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative in antitumor drugs.

The invention provides a p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative and a preparation method and application thereof, wherein a pyrazole ring is introduced into an (E) -7-p-fluorobenzylidene-6, 7-indolizine-8 (5H) -one structure by using a1, 3-dipolar cycloaddition method, so that the p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative is synthesized. The p-fluoro substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative prepared by the invention has a strong tumor cell inhibition effect, and provides a basis for further application in the medical field.

Drawings

FIG. 1 is a flow chart of a preparation method of compound 1 and compound 2 containing a pyridazinone structure according to the present invention;

FIG. 2 is a schematic diagram of the chemical structure of the compound 4 of the present invention.

Detailed Description

The technical solutions of the present invention are further described in detail below with reference to the drawings and examples, which should not be construed as limiting the present invention.

In recent years, pyridazinone compounds become hot spots of research in the pharmaceutical field due to the unique physiological activity of pyridazinone compounds, and have strong effects of inhibiting cancer cell diffusion, resisting bacteria and viruses, reducing blood fat, reducing blood sugar, promoting metabolism and the like. Pyrazole derivatives have attracted considerable attention as a class of useful intermediates and as a variety of pharmaceutical activities that they themselves exhibit. The indolizine skeleton is a common structural unit in bioactive alkaloids, and has remarkable anti-mitotic property or the effect of treating cardiovascular diseases and hypertension.

In order to better research the influence of different heterocycles on pharmacological activity caused by aggregation in the same molecule, the technical scheme synthesizes the spiro [ indolizine-pyrazoline ] derivative with a fluorine-substituted pyridazinone structure through 1, 3-dipolar cycloaddition reaction.

One embodiment of the present invention provides a p-fluoro substituted pyridazinone-containing spiro [ indolizine-pyrazoline ] derivative, which has a chemical structural formula as follows:

wherein: Ar-4-FC₆H₄-, Br is bromine.

The invention provides a preparation method of a p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative, which comprises the following steps:

step 1, synthesizing a chromone aldehyde hydrazone derivative containing a pyridazinone structure.

The invention synthesizes the chromone aldehyde hydrazone derivative containing a pyridazinone structure by a method of generating Schiff base by dehydration reaction of 6-bromo chromone with a formaldehyde group substituted at the 3-position and 6-oxo-1, 6-dihydropyridazine-3-hydrazine carbonate. The chromone aldehyde hydrazone derivative having a pyridazinone structure is also referred to as compound 1 hereinafter.

Example 1: adding 2mmol of 6-oxo-1, 6-dihydropyridazine-3-hydrazine carbonate into a flask containing 15mL of ethanol, refluxing and stirring in a boiling water bath until the mixture is dissolved, slowly dropwise adding 20mL of anhydrous ethanol solution in which 2mmol of 6-bromochromone substituted by 3-formaldehyde groups is dissolved, continuing refluxing and stirring in the boiling water bath for 1 hour, dropwise adding 10 drops of hydrochloric acid, and allowing a light yellow precipitate to appear. And (3) refluxing and stirring in a continuous boiling water bath for 5 hours, stopping the water bath, adding 20mL of distilled water, stirring, deepening the color of the light yellow precipitate, and performing suction filtration to obtain a yellowish red needle-shaped product. Washing with anhydrous ether for multiple times, recrystallizing with ethanol, and vacuum drying to obtain compound 1: chromone aldehyde hydrazone derivatives containing a pyridazinone structure.

And 2, synthesizing α -chloropyridazinone structure chromone aldehyde hydrazone derivatives.

Example 2 Compound 1(10.0mmol) was added to 12.5mL of DMF, and the mixture was completely dissolved under stirring at room temperature, the reaction was controlled to 35 ℃ or lower, 15.0mmol NCS was added in 5 to 10 portions, followed by TLC, and after completion of the reaction, the mixture was poured into water, extracted with diethyl ether for 3 times, the extracted solution was dried over anhydrous magnesium sulfate, and after removing the diethyl ether by rotary evaporation, the mixture was recrystallized from chloroform/petroleum ether to obtain a chromone aldehyde hydrazone derivative having a structure of α -chloropyridazinone, and a chromone aldehyde hydrazone derivative having a structure of α -chloropyridazinone was hereinafter also referred to as Compound 2.

It should be noted that many methods have been studied in the art for synthesizing

compounds

1 and 2, and this example only exemplifies specific examples used in a laboratory, and those skilled in the art can also prepare

compounds

1 and 2 by other methods.

Example 3 adding 1mmol of the compound 3((E) -7-p-fluorobenzylidene-6, 7-indolizin-8 (5H) -one) and 1.6mmol of the compound 2 (the chromone aldehyde hydrazone derivative of the α -chloropyridazinone structure) to 20mL of dichloromethane, dropping a mixed solution of 0.5mL of triethylamine and 5mL of dichloromethane to the solution with a constant pressure dropping funnel, continuing stirring at room temperature for 24 hours after dropping, tracking by TLC, removing the solvent by rotary evaporation after the reaction is completed, dissolving 3mL of ethyl acetate for standby, and finally dissolving with V_{(Ethyl acetate)}:V_{(Petroleum ether)}Silica gel column chromatography 1:5 gave compound 4.

It should be noted that the above dichloromethane is a reaction solvent for dissolving the

compounds

2 and 3, and the present invention is not limited to a specific amount of dichloromethane.

In the step, the mass ratio of the chromone aldehyde hydrazone derivative with the α -chloropyridazinone structure to the (E) -7-p-fluorobenzylidene-6, 7-indolizine-8 (5H) -one is 1.6: 1-2: 1, and the volume ratio of triethylamine to dichloromethane in the mixed solution of triethylamine and dichloromethane is 1: 10-1: 20.

Example 4 adding 1mmol of the compound 3((E) -7-p-fluorobenzylidene-6, 7-indolizin-8 (5H) -one) and 2mmol of the compound 2 (the chromone aldehyde hydrazone derivative of the α -chloropyridazinone structure) to 20mL of dichloromethane, dropping a mixed solution of 0.5mL of triethylamine and 10mL of dichloromethane to the solution with a constant pressure dropping funnel, continuing stirring at room temperature for 24 hours after dropping, tracking by TLC, removing the solvent by rotary evaporation after the reaction is completed, dissolving 3mL of ethyl acetate for standby, and finally dissolving with V_{(Ethyl acetate)}:V_{(Petroleum ether)}Silica gel column chromatography 1:5 gave compound 4.

In examples 3, 4, ethyl acetate was used to dissolve the residue remaining from the rotary evaporation, to facilitate removal, and to facilitate addition to the top of the column. It will be readily understood that the residue may be dissolved with a solvent other than ethyl acetate, for example, ethanol, acetone may also be used.

Preferably, the expanding agent adopted in the silica gel column chromatography separation is V_{(Ethyl acetate)}:V_{(Petroleum ether)}1: 5. Of course, other proportions of extender may be used, e.g. V_{(Ethyl acetate)}:V_{(Petroleum ether)}＝1:6。

The experimental data are as follows: a p-fluoro substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative (compound 4) is a pale yellow powder, yield 37.4%, melting point: 215 ℃ and 216 ℃, and the nuclear magnetic hydrogen spectrum data and the element analysis data are as follows:

¹H NMR(DMSO-d₆)δ:10.17(s,1H,NHN),7.32-7.19(m,9H,Ar-H),6.88(t,J＝8.0,1H),6.58(s,1H),6.50(s,1H,C＝C-H),6.32(d,J＝4.0,1H),4.53(t,J＝12.5,1H),4.47(s,1H,C＝C-H),4.05(d,J＝12.5,1H),2.43(t,J＝14.0,1H),1.69(d,J＝14.5,1H),

IR(KBr)v/cm^-13429(N-C＝O),3103(ArH),1770(C＝O,pyridazinone),1664(C＝O),1617,1582,1464(C＝N,C＝C),1255(C-O-C),1610(chromone-ring),655(C-F)

m/e:627(100.0％)

Anal.calcd.for C₃₀H₁₉BrFN₅O₅：C,57.33；H,3.07；N,11.12。

wherein pyridazinone is pyridazinone and chromone-ring is chromone mother ring.

In this example, MTT method is used to determine the in vitro inhibitory effect of compound 4 on different tumor strains, and the results of the determination of the anti-tumor activity of the fluorine-substituted pyridazinone-structure-containing spiro [ indolizine-pyrazoline ] derivative are as follows:

dissolving compound 4 in DMSO, diluting, inoculating tumor cells KB (oral cancer cells), SGC7901 (gastric cancer cells), HL-60 (leukemia cells), Bel-7402 (human liver cancer cells), HO8901 (ovarian cancer cells), and ECA109 (intestinal cancer cells) into a 96-well plate at 4000/200 μ L/well, adding compound 2 μ L per well to the final concentration of 12.0 μ M, 6.0 μ M, 3.0 μ M, 1.5 μ M, and mixing at 37 deg.C and 5% CO₂The cells were incubated in an incubator for 72 hours, with DMSO (1%) as a blank control. After 72 hours, MTT was added to a final concentration of 0.25mg/mL and the mixture was left at 37 ℃ with 5% CO₂After 4 hours in the cell incubator, the solvent was blotted, 100. mu.l DMSO was added to each well, absorbance (OD value) was measured at 570nm with an enzyme-linked immunosorbent assay, and the data obtained was used to calculate IC₅₀The value is obtained. Selecting compounds with high inhibitory activity, and determining the influence of different action times of the compounds at different concentrations on the human tumor cell cycle and apoptosis.

The test compounds of different concentrations were coarse-screened in 96-well plates and IC was calculated from the resulting inhibition₅₀Values, results are given in the table below.

TABLE 1

In Table 1, spiro [ indolizine-pyrazoline ] having a p-fluoro-substituted pyridazinone structure is shown]IC of derivative (Compound 4) against six tumor cell lines₅₀The values indicate that the compounds have strong tumor cell inhibiting effects on KB (oral cancer cells), SGC7901 (gastric cancer cells), HL-60 (leukemia cells) and Bel-7402 (human hepatoma cells), and further indicate that the compounds have strong tumor cell inhibiting effects on KB (oral cancer cells), SGC7901 (gastric cancer cells), HL-60 (leukemia cells) and Bel-7402 (human hepatoma cells)Provides a basis for the application of the medicine field.

The above embodiments are only for illustrating the technical solution of the present invention and not for limiting the same, and those skilled in the art can make various corresponding changes and modifications according to the present invention without departing from the spirit and the essence of the present invention, but these corresponding changes and modifications should fall within the protection scope of the appended claims.

Claims

1. The p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative is characterized in that the chemical structural formula of the p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative is as follows:

wherein Ar is 4-FC₆H₄-。

2. A preparation method of a p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative is characterized in that the preparation method of the p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative comprises the following steps:

step 1, synthesizing a chromone aldehyde hydrazone derivative containing a pyridazinone structure by adopting 6-bromochromone with a formaldehyde group substitution at the 3-position and 6-oxo-1, 6-dihydropyridazine-3-hydrazine carbonate, wherein the chromone aldehyde hydrazone derivative containing the pyridazinone structure has the following chemical structural formula:

step 2, synthesizing α -chloropyridazinone structure chromone aldehyde hydrazone derivative, wherein the α -chloropyridazinone structure chromone aldehyde hydrazone derivative has the following chemical structural formula:

step 3, adding (E) -7-p-fluorobenzylidene-6, 7-indolizine-8 (5H) -ketone and a chromone aldehyde hydrazone derivative with a α -chloropyridazinone structure into dichloromethane, dropwise adding a mixed solution of triethylamine and dichloromethane into the solution by using a constant-pressure dropping funnel, continuously stirring at normal temperature for 20-30 hours after the dropwise adding is finished, tracking by TLC (thin layer chromatography), removing the solvent by rotary evaporation after the reaction is completed, dissolving residues left after the rotary evaporation, and performing chromatographic separation by using a silica gel column to obtain a spiro [ indolizine-pyrazoline ] derivative with a p-fluoro substituted pyridazinone structure;

the chemical structural formula of the p-fluorine substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] derivative is as follows:

wherein Ar is 4-FC₆H₄-。

3. The method for preparing a p-fluoro substituted pyridazinone-structure-containing spiro [ indolizine-pyrazoline ] derivative according to claim 2, wherein the ratio of the amounts of the α -chloropyridazone-structure chromonaldehyde hydrazone derivative and the (E) -7-p-fluorobenzylidene-6, 7-indolizine-8 (5H) -one is 1.6: 1 to 2: 1.

4. The method for preparing a p-fluoro-substituted pyridazinone-structure-containing spiro [ indolizine-pyrazoline ] derivative according to claim 2, wherein the ratio of triethylamine to dichloromethane by volume is: 1: 10-1: 20.

5. the para-fluoro substituted pyridazinone structure-containing spiro [ indolizine-pyrazoline ] according to claim 2]The preparation method of the derivative is characterized in that the expanding agent adopted in the silica gel column chromatography separation is V_{(Ethyl acetate)}:V_{(Petroleum ether)}＝1:5。

6. The use of the p-fluoro substituted pyridazinone-structure-containing spiro [ indolizine-pyrazoline ] derivative according to claim 1 in the preparation of an antitumor drug.