CN114014814A

CN114014814A - N-benzyl-4-phenylphthalazin-1-amine derivative, preparation method thereof and medicine for treating RECQL4 specific expression

Info

Publication number: CN114014814A
Application number: CN202111484091.7A
Authority: CN
Inventors: 赵雪梅; 张雪莲; 王桂玲; 王家玲; 王琢; 付琳琳; 狄伟华
Original assignee: Shandong First Medical University and Shandong Academy of Medical Sciences
Current assignee: Shandong First Medical University and Shandong Academy of Medical Sciences
Priority date: 2021-12-07
Filing date: 2021-12-07
Publication date: 2022-02-08
Anticipated expiration: 2041-12-07
Also published as: CN114014814B

Abstract

The invention belongs to the technical field of organic compounds, and particularly relates to an N-benzyl-4-phenylphthalazin-1-amine derivative, a preparation method thereof, and a liver cancer drug for treating RECQL4 specific expression. The invention provides an N-benzyl-4-phenylphthalazin-1-amine derivative which has a structural formula shown in formula I, wherein R in the formula I₁is-H or-OCH₃；R₂is-H, phenyl, substituted phenyl, alkyl, alkoxy, alkenyl or diphenylamino; r₃is-H, nitro or methyl; r₄is-H or phenyl; r₁、R₂、R₃And R₄Not being simultaneously-H. The N-benzyl-4-phenylphthalazine-1-amine derivative provided by the invention has a strong inhibition effect on the proliferation of liver cancer cells SNU-398 with high expression of RECQL 4. Book (I)The invention provides scientific experimental data for developing a novel medicine for resisting RECQL4 high-expression liver cancer and clinically and individually treating the RECQL4 high-expression liver cancer;

Description

N-benzyl-4-phenylphthalazin-1-amine derivative, preparation method thereof and medicine for treating RECQL4 specific expression

Technical Field

The invention belongs to the technical field of organic synthetic drugs, and particularly relates to an N-benzyl-4-phenylphthalazin-1-amine derivative, a preparation method thereof and a liver cancer drug for treating RECQL4 specific expression.

Background

Hepatocellular carcinoma (HCC) is a primary hepatic malignancy. Researches show that the causes of HCC are complex, the number of patients and the number of deaths are increased year by year, the main causes of HCC are different all around the world except ethnic and genetic backgrounds, and HCC can be caused by hepatitis virus, fatty liver, autoimmune liver disease, ingestion of fungal metabolite aflatoxin B1, DNA damage and the like. However, genomic instability is a major feature of cancer development, and RECQ helicase is one of the members of DNA helicase and plays an important role in maintaining gene stability. The RECQ helicase family includes five proteins, RECQ1, WRN, BLM, RECQL4, and RECQ. Abnormal low expression of RECQL4 is associated with cancer susceptibility and premature aging, and up-regulation of RECQL4 is associated with cancer development and metastasis. In vitro experiments show that RECQL4 can play a synergistic role with BLM, increase helicase activity of S phase of cell cycle, and increase nuclear genome stability. RECQL4 can regulate chromosome to perform normal function, improve mitochondrial stability, and maintain human gene in normal state. When RECQL4 is abnormally or abnormally expressed or deleted, the reactive oxygen species level in cells is obviously increased, and the oxidative damage repair capacity of mitochondrial DNA is greatly reduced, which indicates that human RECQL4 plays an important role in the DNA replication initiation process.

Therefore, in HCC patients with up-regulated RECQL4 expression level, silencing or inhibiting the function of RECQL4, inhibiting the proliferation of tumor cells and promoting the apoptosis of the tumor cells can be a novel anti-cancer treatment method, and the targeted control of the RECQL4 protein expression can play a role in treating liver cancer. At present, no report exists for screening the inhibitor for inhibiting the proliferation of the liver cancer cells by using RECQL4 as a target point.

Disclosure of Invention

In view of the above, the invention provides an N-benzyl-4-phenylphthalazin-1-amine derivative, a preparation method thereof and a liver cancer drug for treating RECQL4 specific expression, and the N-benzyl-4-phenylphthalazin-1-amine derivative provided by the invention has a strong inhibition effect on the proliferation of liver cancer cells with high expression of RECQL 4.

In order to solve the technical problem, the invention provides an N-benzyl-4-phenylphthalazin-1-amine derivative, which has a structural formula shown in formula I:

R₁is-H or-OCH₃；

R₂is-H, phenyl, substituted phenyl, alkyl, alkoxy, alkenyl or diphenylamino;

R₃is-H, nitro or methyl;

R₄is-H or phenyl;

R₁、R₂、R₃and R₄Not being simultaneously-H.

Preferably, the substituted phenyl group comprises phenoxy or

Preferably, the alkyl group comprises an alkyl group with 1-6 carbon atoms;

the alkoxy group includes an alkoxy group having 1 to 3 carbon atoms.

Preferably, the alkyl group comprises methyl, ethyl, propyl, tert-butyl, pentyl or hexyl;

the alkoxy group includes-OCH₃or-OC₃H₇；

The alkenyl group includes a vinyl group.

Preferably, the structure has a structure represented by any one of formulas 1 to 31:

the invention provides a preparation method of the N-benzyl-4-phenylphthalazin-1-amine derivative in the technical scheme, which comprises the following steps:

mixing a reactant 1, a reactant 2, a mixed solvent of soluble alkali, an organic solvent and water and an organic palladium catalyst, and carrying out a coupling reaction to obtain the N-benzyl-4-phenylphthalazine-1-amine derivative;

the reactant 1 is

Wherein R is₁is-H or-OCH₃；

The reactant 2 is

Wherein R is₂is-H, phenyl, substituted phenyl, alkyl, alkoxy, alkenyl or diphenylamino; r₃is-H, nitro or methyl; r₄is-H or phenyl; r₁、R₂、R₃And R₄Not being simultaneously-H.

Preferably, the molar ratio of the reactant 1 to the reactant 2 is 1: 1.1-1.7.

The soluble alkali comprises triethylamine, sodium hydroxide or sodium carbonate;

the molar ratio of the reactant 1 to the soluble alkali is 1: 2.8-3.2.

Preferably, the mixed solvent of the organic solvent and water comprises toluene, ethanol and water;

the organic palladium catalyst comprises palladium acetate or palladium tetratriphenylphosphine.

Preferably, the temperature of the coupling reaction is 100-110 ℃, and the time is 15-17 h.

The invention provides a liver cancer drug for treating RECQL4 specific expression, which comprises an active inhibitor and a drug carrier, wherein the active inhibitor comprises N-benzyl-4-phenylphthalazin-1-amine derivatives or N-benzyl-4-phenylphthalazin-1-amine;

the N-benzyl-4-phenylphthalazin-1-amine derivative is the N-benzyl-4-phenylphthalazin-1-amine derivative according to any one of claims 1 to 5 or the N-benzyl-4-phenylphthalazin-1-amine derivative prepared by the preparation method according to any one of claims 6 to 9.

The invention provides an N-benzyl-4-phenylphthalazin-1-amine derivative, which has a structural formula shown in a formula I:

R₁is-H or-OCH₃；R₂is-H, phenyl, substituted phenyl, alkyl, alkoxy, alkenyl or diphenylamino; r₃is-H, nitro or methyl; r₄is-H or phenyl; r₁、R₂、R₃And R₄Not being simultaneously-H. The N-benzyl-4-phenylphthalazine-1-amine derivative provided by the invention has a strong inhibition effect on the proliferation of liver cancer cells SNU-398 with high expression of RECQL 4. The invention provides scientific experimental data for developing a novel medicine for resisting RECQL4 high-expression liver cancer and clinically and individually treating the RECQL4 high-expression liver cancer.

Detailed Description

R₁is-H or-OCH₃；

R₂is-H, phenyl, substituted phenyl, alkyl, alkoxy, alkenyl or diphenylamino;

R₃is-H, nitro or methyl;

R₄is-H or phenyl;

R₁、R₂、R₃and R₄Not being simultaneously-H.

In the present invention, said R₁is-H or-OCH₃Is preferably-OCH₃。

In the present invention, said R₂is-H, phenyl, substituted phenyl, alkyl, alkoxy, alkenyl or diphenylamino, preferably-H, alkyl, alkoxy or diphenylamino, more preferably alkyl. In the present invention, the substituted phenyl group preferably includes a phenoxy group or

More preferably

In the present invention, the alkyl group preferably includes an alkyl group having 1 to 6 carbon atoms, more preferably a methyl group, an ethyl group, a propyl group, a tert-butyl group, a pentyl group, or a hexyl group, and still more preferably a propyl group, a tert-butyl group, a pentyl group, or a hexyl group; the alkoxy group preferably comprises an alkoxy group with 1-3 carbon atoms, and is more preferably-OCH₃or-OC₃H₇Further preferred is-OC₃H₇. In the present invention, the alkenyl group preferably includes a vinyl group.

In the present invention, said R₃is-H, nitro or methyl, preferably-H or methyl, more preferably-H.

In the present invention, said R₄is-H or phenyl, preferably-H.

In the invention, the N-benzyl-4-phenylphthalazin-1-amine derivative preferably has a structure shown in any one of formulas 1-31:

the N-benzyl-4-phenylphthalazine-1-amine derivative provided by the invention can be combined with the active site amino acid of the ATPase structural domain of RECQL4, namely can interact with amino acids such as glutamic acid (Glu606), arginine (Arg804) and the like through hydrogen bonds; and can form a hydrophobic interaction force with amino acids such as valine (Va1772), proline (Pro532), leucine (Leu534) and methionine (Met776), so as to inhibit the activity of ATPase, indirectly influence the ATPase-mediated Helicase enzyme activity, inhibit the expression of RECQL4 protein, reduce or eliminate the function, influence the proliferation of tumor cells and play an anti-tumor effect.

The invention also provides a preparation method of the N-benzyl-4-phenylphthalazin-1-amine derivative in the technical scheme, which comprises the following steps:

mixing a mixed solvent of a reactant 1, a reactant 2, a soluble alkali, an organic solvent and water with an organic palladium catalyst to carry out a coupling reaction (hereinafter referred to as a first coupling reaction) to obtain the N-benzyl-4-phenylphthalazin-1-amine derivative;

the reactant 1 is

Wherein R is₁is-H or-OCH₃；

The reactant 2 is

In the present invention, when the reactant 1 is

When the reaction product 1 is prepared, the preparation method preferably comprises the following steps:

mixing phthalylhydrazine and a phosphoryl trichloro solution for substitution reaction to obtain an intermediate 1; what is needed isThe structural formula of the intermediate 1 is

The intermediate 1, benzylamine, and triethylamine are dissolved in tetrahydrofuran to perform a coupling reaction (hereinafter referred to as a second coupling reaction), thereby obtaining the reactant 1.

The method comprises the steps of mixing phthalhydrazide and a trichlorophosphoryl solution for substitution reaction to obtain an intermediate 1; the structural formula of the intermediate 1 is

In the invention, the phosphorus oxychloride solution is preferably 22 to 28% by mass, more preferably 24 to 26% by mass, and most preferably 25.2% by mass. In the invention, the molar ratio of the phthalhydrazide to the phosphoryl trichloride in the phosphoryl trichloride solution is preferably 0.5-1.5: 10-20, and more preferably 0.9-1.1: 14-16. In the present invention, the mixing is preferably: phthalhydrazide is added to a stirred solution of trichlorophosphoryl. The invention has no special requirements on the stirring, as long as the materials can be uniformly mixed. In the invention, the temperature of the substitution reaction is preferably 108-112 ℃, and more preferably 110 ℃; the time of the substitution reaction is preferably 0.8-1.2 h, and more preferably 1 h. The present invention preferably utilizes TLC to monitor whether the reaction is complete.

In the present invention, it is preferable that the substitution reaction further comprises: cooling the system after the substitution reaction to room temperature, and adding ice water for quenching; and filtering the quenched system. In the invention, the room temperature is preferably 20-30 ℃, and more preferably 23-25 ℃. In the present invention, the cooling method is not particularly limited as long as the cooling can be performed to a desired temperature. The amount of the ice water is not particularly limited, as long as quenching can be achieved. According to the invention, washing is preferably carried out in the filtering process, the solvent for washing is preferably deionized water, and the pH value of the washing liquid after washing is preferably neutral. In the present invention, the intermediate 1 is a white solid.

After the intermediate 1 is obtained, the intermediate 1, benzylamine and triethylamine are dissolved in tetrahydrofuran to carry out a second coupling reaction, so as to obtain the reactant 1. In the present invention, the molar ratio of the intermediate 1, benzylamine, and triethylamine is preferably 1:1.0 to 1.3:2.8 to 3.2, and more preferably 1:1.1: 3. The invention has no special requirement on the dosage of the tetrahydrofuran, as long as the intermediate 1, the benzylamine and the triethylamine can be completely dissolved. The present invention is not particularly limited as long as the dissolution can be completed. In the invention, the temperature of the second coupling reaction is preferably 98-102 ℃, and more preferably 100 ℃; the time of the second coupling reaction is preferably 3.8-4.2 h, and more preferably 4 h. The present invention preferably utilizes TLC to monitor whether the second coupling reaction is complete.

In the present invention, the second coupling reaction preferably further comprises:

mixing the solution after the second coupling reaction with water, and then extracting to obtain an organic phase;

and washing, drying and concentrating the organic phase in sequence, and then performing chromatographic separation to obtain the reactant 1.

The solution after the second coupling reaction is mixed with water and then extracted to obtain an organic phase. In the invention, the volume ratio of the solution after the second coupling reaction to water is preferably 1: 0.8-1.5, and more preferably 1: 0.95-1.1. In the present invention, the extraction solvent is preferably ethyl acetate or dichlorohexane, and more preferably ethyl acetate. The extraction is not particularly limited in the present invention and may be performed in a manner conventional in the art.

After an organic phase is obtained, the organic phase is washed, dried and concentrated in sequence and then subjected to chromatographic separation to obtain the reactant 1. In the present invention, the washing solvent is preferably a saturated sodium chloride solution. The method carries out interference emulsification through the washing, and the dosage of the sodium chloride solution is not specially limited as long as the interference emulsification can be realized. In the present invention, the drying is preferably performed using anhydrous sodium sulfate. In the present invention, the drying preferably further comprises filtering the dried product to obtain a filtrate. In the invention, the concentration is preferably rotary evaporation, and the temperature of the rotary evaporation is preferably 50-57 ℃, and more preferably 53-55 ℃. In the present invention, the time of the rotary evaporation is not particularly limited as long as the solvent can be removed. In the present invention, the chromatographic column for chromatographic separation is preferably a silica gel column; the eluent for chromatographic separation is preferably a mixture of petroleum ether and ethyl acetate, and the mass ratio of the petroleum ether to the ethyl acetate is preferably 14-16: 1, and more preferably 15: 1.

In the present invention, when the reactant 1 is

When the temperature of the water is higher than the set temperature,

the preparation method of (1) is preferably referred to the preparation method

Except that benzylamine was replaced with p-methoxybenzylamine.

In the present invention, the soluble base preferably comprises sodium hydroxide, sodium carbonate or triethylamine, more preferably sodium carbonate. In the invention, the organic solvent in the mixed solvent of the organic solvent and water preferably comprises toluene, ethanol and water, and the volume ratio of the toluene, the ethanol and the water is preferably 4.8-5.2: 1:1, and more preferably 5:1: 1.

In the present invention, the organopalladium catalyst preferably comprises palladium acetate or tetratriphenylphosphine palladium, more preferably tetratriphenylphosphine palladium. In the invention, the molar ratio of the reactant 1 to the reactant 2 is preferably 1: 1.1-1.7, more preferably 1: 1.3-1.7, and most preferably 1: 1.5; the molar ratio of the reactant 1 to the soluble alkali is preferably 1: 2.8-3.2, and more preferably 1: 3. The amount of the catalyst used in the present invention is not particularly limited as long as the catalyst can perform a catalytic function.

The mixing is not particularly limited in the present invention as long as it can be mixed uniformly. In the present invention, the first coupling reaction preferably further comprises: and introducing a protective gas into the reaction solution, wherein the protective gas preferably comprises nitrogen, inert gas or carbon dioxide, and the inert gas is preferably He gas, Ne gas or Ar gas. In the present invention, the shielding gas is more preferably nitrogen; the time for replacing nitrogen is preferably 8-12 min, and more preferably 10 min.

In the invention, the temperature of the first coupling reaction is preferably 100-110 ℃; the time of the first coupling reaction is preferably 15-17 h, and more preferably 16 h. In the present invention, the first coupling reaction is preferably carried out under a protective gas atmosphere; the protective gas preferably includes nitrogen, an inert gas, or carbon dioxide, and the inert gas is preferably He gas, Ne gas, or Ar gas. In the present invention, the shielding gas is more preferably nitrogen. In the present invention, the first coupling reaction is preferably accompanied by stirring; the rotating speed of the stirring is preferably 100-150 r/min, and more preferably 110-130 r/min. The present invention preferably utilizes TLC to monitor whether the first coupling reaction is complete.

In the present invention, the first coupling reaction preferably further comprises: and (3) performing rotary evaporation and post-treatment on the solution after the first coupling reaction in sequence. In the invention, the rotary evaporation temperature is preferably 50-57 ℃, and more preferably 53-55 ℃; the rotary evaporation time is preferably 10-15 min, and more preferably 10 min. The invention can remove partial solvent through the rotary evaporation, and is beneficial to the subsequent post-treatment. In the present invention, the post-treatment is preferably the same as the post-treatment after the second coupling reaction, and thus the detailed description thereof is omitted.

In one embodiment of the present invention, the reaction equation for preparing the N-benzyl-4-phenylphthalazin-1-amine derivative having formula I is shown as follows:

the invention also provides a liver cancer medicament for treating RECQL4 specific expression, which comprises an active inhibitor and a medicament carrier, wherein the active inhibitor comprises N-benzyl-4-phenylphthalazine-1-amine derivatives or N-benzyl-4-phenylphthalazine-1-amine;

the N-benzyl-4-phenylphthalazine-1-amine derivative is the N-benzyl-4-phenylphthalazine-1-amine derivative in the technical scheme or the N-benzyl-4-phenylphthalazine-1-amine derivative prepared by the preparation method in the technical scheme.

In the invention, the structural formula of the N-benzyl-4-phenylphthalazin-1-amine is shown as a formula 32:

the source of the N-benzyl-4-phenylphthalazin-1-amine is not specially limited, and the conventional commercial product is adopted. In the present invention, the preparation method of the N-benzyl-4-phenylphthalazin-1-amine is preferably the same as the preparation method of the N-benzyl-4-phenylphthalazin-1-amine derivative, except that R in the reactant 1₁is-H; reacting a reactant 2 with

Is replaced by

In order to further illustrate the present invention, the following embodiments are described in detail, but they should not be construed as limiting the scope of the present invention.

Example 1

Adding 1.68g (10mmol) of phthalhydrazide into 15mL of continuously stirred phosphorus oxychloride trichloro solution with the mass concentration of 98%, coupling reaction at 110 ℃ for 1h (TLC monitoring), cooling the reaction solution to 25 ℃, dropwise adding ice water, quenching, stirring, filtering, and washing with water to be neutral to obtain a white solid intermediate 1(1.85g, yield of 90%);

0.5mmol of intermediate 1, 0.55mmol of benzylamine and 1.5mmol of triethylamine are dissolved in 2mL of tetrahydrofuran and the coupling reaction is carried out for 4h at 100 ℃ (TLC monitoring); adding 2mL of water after the coupling reaction, adding ethyl acetate solvent for extraction, carrying out salt washing on the collected organic phase by using saturated sodium chloride solution, drying the organic phase by using anhydrous sodium sulfate, filtering to remove impurities, carrying out rotary evaporation concentration at 53 ℃, and feeding the organic phase by using a silica gel columnLine chromatography (eluent is a mixture of petroleum ether and ethyl acetate in a volume ratio of 15:1 to give 93.7mg of white solid reactant 1

(yield 69.4%);

adding 0.37mmol of reactant 1 and 0.555mmol of reactant

(wherein R is₂is-C₅H₁₁、R₃is-H, R₄-H), 1.11mmol of sodium carbonate was dissolved in 10mL of a mixed solvent containing toluene, ethanol and water (the volume ratio of toluene, ethanol and water was 5:1:1), and 5mg of tetrakistriphenylphosphine palladium was added to the solution; introducing nitrogen into the reaction solution for 10min, and then carrying out coupling reaction for 16h at 110 ℃ under the nitrogen atmosphere (TCL monitoring); performing rotary evaporation on the solution after the coupling reaction at 53 ℃ for 10min, mixing with 2mL of water, adding an ethyl acetate solvent for extraction, performing salt washing on the collected organic phase by using a saturated sodium chloride solution, drying the organic phase by using anhydrous sodium sulfate, filtering to remove impurities, performing rotary evaporation and concentration at 53 ℃, and performing chromatographic separation by using a silica gel column (the eluent is a mixture of petroleum ether and ethyl acetate with the volume ratio of 15:1 to obtain the product

(yield 38.48%).

Examples 2 to 15

The N-benzyl-4-phenylphthalazin-1-amine derivative was prepared as in example 1 except that R in reactant 2 was₂、R₃And R₄The substituents of (A) are set as in Table 2.

Examples 16 to 31

N-benzyl-4-phenylphthalazin-1-amine derivative was prepared as in example 1, except that benzylamine was replaced with p-methoxybenzylamine and R in reactant 2₂、R₃And R₄The substituents of (A) are set as in Table 2.

The binding site of the target protein of interest (RECQL4 protein) was docked with the derivatives prepared in examples 1-31. After docking is carried out by using Discovery Studio 2017R 2 software and Libdock scoring is given, the compounds with better combination can be virtually screened as the objects of the next research according to the scoring condition. The scoring condition will become a basis for designing the compound. The resulting derivatives and the target protein of interest (RECQL4 protein) docking scores are listed in table 1.

TABLE 1R in reactant 2 of examples 1-31₂、R₃And R₄The substituent and each derivative are butted and scored with target protein

In Table 1, -Ph is phenyl.

Subjecting the products obtained in examples 1 to 31 to hydrogen nuclear magnetic resonance spectroscopy (¹H NMR detection, nuclear magnetic resonance carbon Spectroscopy: (¹³C NMR) and high resolution mass spectrometry (electrospray ionization, HRMS (ESI +)) results are as follows.

Example 1 test results:¹HNMR(400MHz,DMSO-d₆)δ8.44(d,J＝8.2Hz,1H),8.18(s,1H),7.95–7.81(m,3H),7.50(d,J＝8.0Hz,2H),7.43(d,J＝7.4Hz,2H),7.33(dd,J＝15.1,7.8Hz,4H),7.23(s,1H),4.86(d,J＝5.8Hz,2H),2.67(t,J＝7.6Hz,2H),1.65(s,2H),1.34(dd,J＝7.0,3.6Hz,4H),0.89(t,J＝6.9Hz,3H).

¹³C NMR(101MHz,DMSO-d₆)δ153.24,151.89,142.94,140.80,134.93,132.20,131.62,129.96,128.68,127.69,126.98,125.93,122.78,118.36,44.54,35.38,31.43,31.11,22.48,14.43.

HRMS (ESI +): molecular formula C₂₆H₂₇N₃，[M+H]⁺382.2285, found: 382.2289.

example 1 the derivative prepared has the formula

The name of the compound is N-benzyl-4- (4-pentylphenyl) phthalazine-1-amine, the yield is 38.4%, and the melting point is 129.7-130.1 ℃.

Example 2 test results:¹HNMR(400MHz,DMSO-d₆)δ8.43(d,J＝8.2Hz,1H),8.14(s,1H),7.89(dd,J＝29.1,3.8Hz,3H),7.52(d,J＝8.6Hz,2H),7.43(d,J＝7.3Hz,2H),7.32(t,J＝7.5Hz,2H),7.24(d,J＝7.3Hz,1H),7.08(d,J＝8.7Hz,2H),4.85(d,J＝5.7Hz,2H),4.02(t,J＝6.5Hz,2H),1.78(dd,J＝14.0,6.9Hz,2H),1.02(t,J＝7.4Hz,3H).

¹³C NMR(101MHz,DMSO-d₆)δ159.19,153.14,151.64,140.83,132.14,131.57,131.29,129.82,128.64,127.69,126.97,126.07,122.77,118.42,114.69,69.47,44.54,22.53,10.93.

HRMS (ESI +): molecular formula C₂₄H₂₃N₃O，[M+H]⁺370.1915, found: 370.1919.

example 2 the derivative prepared has the formula

The name is N-benzyl-4- (4- (propoxy) phenyl) phthalazine-1-amine, the yield is 38.05 percent, and the melting point is 128.3-129.2 ℃.

Example 3 test results:¹HNMR(400MHz,DMSO-d₆)δ8.43(d,J＝8.2Hz,1H),8.16(s,1H),7.92(s,1H),7.90-7.81(m,2H),7.57-7.46(m,4H),7.46–7.27(m,7H),7.38-7.29(m,3H),7.27-7.14(m,3H),5.20(s,2H),4.85(d,J＝5.8Hz,2H).

¹³C NMR(101MHz,DMSO-d₆)δ158.87,153.16,151.59,140.81,137.49,132.17,131.59,131.31,130.08,128.97,128.65,128.32,127.68,126.98,126.05,122.78,118.41,115.07,69.76,44.54.

HRMS(ESI+)：C₂₈H₂₃N₃O，[M+H]⁺418.1920, found: 418.1922.

example 3 the derivative prepared has the formula

The name of the compound is N-benzyl-4- (4- (benzyloxy) phenyl) phthalazine-1-amine, the yield is 35.55%, and the melting point is 168.2-169.1 ℃.

Example 4 test results:¹H NMR(400MHz,CDCl₃)δ8.03-7.98(d,1H),7.90-7.81(m,1H),7.80-7.71(m,2H),7.66(d,J＝8.6Hz,2H),7.49(d,J＝7.1Hz,2H),7.36(dd,J＝16.4,8.9Hz,4H),7.29(dd,J＝7.7,5.4Hz,1H),7.12(dd,J＝13.3,8.1Hz,5H),5.60(s,1H),4.95(s,2H).

¹³C NMR(101MHz,CDCl₃)δ157.86,139.06,131.38,131.09,129.87,128.72,128.46,127.54,126.80,123.64,120.92,119.35,118.44,46.24.

HRMS (ESI +): molecular formula C₂₇H₂₁N₃，[M+H]⁺388.1820, found: 388.1821.

example 4 the derivative prepared has the formula

Named 4- ([1,1' -biphenyl)]-4-yl) -N-benzylphthalazin-1-amine, the yield is 33.71%, and the melting point is 228.2-229.1 ℃.

Example 5 test results:¹HNMR(400MHz,CDCl₃)δ8.04(d,J＝8.2Hz,1H),7.86-7.81(m,1H),7.79-7.74(m,2H),7.70-7.66(m,2H),7.53-7.48(m,2H),7.41-7.34(m,4H),7.33-7.29(m,1H),7.18-7.07(m,5H),5.47(s,1H),4.96(s,2H).

¹³C NMR(101MHz,CDCl₃)δ157.85,156.88,152.74,139.07,131.90,131.36,131.06,129.87,128.75,128.49,127.59,126.84,123.63,120.69,119.34,118.45,46.30.

HRMS (ESI +): molecular formula C₂₇H₂₁N₃O，[M+H]⁺404.1764, found: 404.1765.

the structural formula of the derivative prepared in example 5 is

Is named as N-benzyl-4- (4-phenoxyphenyl) phthalazine-1-amine, the yield is 39.28 percent, the melting point is 112.2 to 113.1 DEG C。

Example 6 test results:¹HNMR(400MHz,CDCl₃)δ8.01(d,J＝7.7Hz,1H),7.92(d,J＝7.8Hz,1H),7.83-7.73(m,2H),7.67(d,J＝8.2Hz,2H),7.54(dd,J＝20.4,7.6Hz,4H),7.41-7.30(m,3H),6.81(dd,J＝17.6,10.9Hz,1H),5.86(d,J＝17.6Hz,1H),5.76-5.55(m,1H),5.37-5.29(m,1H),4.98(s,2H).

¹³C NMR(101MHz,CDCl₃)δ152.77,138.81,137.81,136.46,136.20,131.50,131.23,130.14,128.80,128.48,127.66,126.92,126.23,120.92,118.64,114.60,29.72.

HRMS (ESI +): molecular formula C₂₃H₁₉N₃，[M+H]⁺38.1664, found: 338.1666.

the structural formula of the derivative prepared in example 6 is

The name of the compound is N-benzyl-4- (4-vinyl phenyl) phthalazine-1-amine, the yield is 38.58%, and the melting point is 123.7-124.1 ℃.

Example 7 test results:¹HNMR(400MHz,CDCl₃)δ7.91(dd,J＝7.4,1.9Hz,1H),7.78(d,J＝7.9Hz,1H),7.72-7.60(m,2H),7.50(d,J＝8.0Hz,2H),7.44-7.38(m,2H),7.28(t,J＝7.3Hz,2H),7.22(dd,J＝9.7,7.6Hz,3H),5.46(s,1H),4.87(s,2H),2.36(s,3H).

¹³C NMR(101MHz,CDCl₃)δ153.32,152.66,139.07,138.36,134.07,131.22,131.00,129.85,129.06,128.74,128.49,127.57,127.02,126.38,120.67,118.59,46.29,21.38.

HRMS (ESI +): molecular formula C₂₂H₁₉N₃，[M+H]⁺326.1663, found: 326.1664.

the structural formula of the derivative prepared in example 7 is

The name is N-benzyl-4- (p-tolyl) phthalazin-1-amine, the yield is 36.63 percent, and the melting point is 158.2-158.9 ℃.

Example 8 test results:¹HNMR(400MHz,CDCl₃)δ7.65(d,J＝8.2Hz,1H),7.61-7.55(m,2H),7.55-7.49(m,3H),7.49-7.45(m,2H),7.45-7.39(m,1H),7.39-7.34(m,2H),7.33-7.29(m,2H),7.23-7.17(m,2H),7.09-6.89(m,3H),5.41(s,1H),4.93(dd,J＝38.9,14.1Hz,2H).

¹³C NMR(101MHz,CDCl₃)δ154.15,152.91,141.92,141.13,139.10,135.70,131.46,130.80,129.90,129.33,129.05,128.79,128.56,127.91,127.61,126.96,126.58,120.28,117.86,46.33.

HRMS (ESI +): molecular formula C₂₇H₂₁N₃，[M+H]⁺388.1814, found: 388.1814.

the structural formula of the derivative prepared in example 8 is

Named 4- ([1,1' -biphenyl)]-2-yl) -N-benzylphthalazin-1-amine, with a yield of 32.1% and a melting point of 167.2-168.4 ℃.

Example 9 test results:¹HNMR(400MHz,CDCl₃)δ8.57(t,J＝1.8Hz,1H),8.39-8.29(m,1H),8.07(d,J＝7.7Hz,1H),7.98(d,J＝7.6Hz,1H),7.94–7.88(m,1H),7.88–7.79(m,2H),7.70(t,J＝8.0Hz,1H),7.53-7.44(m,2H),7.33(dt,J＝23.1,7.1Hz,3H),5.88(s,1H),4.99(s,3H).

¹³C NMR(101MHz,CDCl₃)δ153.02,150.88,148.23,138.60,135.93,132.07,131.66,129.57,128.80,128.39,127.71,125.83,124.82,123.44,121.32,118.46,46.25.

HRMS (ESI +): molecular formula C₂₁H₁₆N₄O₂，[M+H]⁺357.1355, found: 357.1357.

the structural formula of the derivative prepared in example 9 is

The name is N-benzyl-4- (3-nitrophenyl) phthalazine-1-amine, the yield is 37.91%, and the melting point is 153.9-154.3 ℃.

Example 10 test results:¹H NMR(400MHz,CDCl₃)δ7.91(dd,J＝7.4,1.9Hz,1H),7.78(d,J＝7.9Hz,1H),7.72-7.60(m,2H),7.50(d,J＝8.0Hz,2H),7.44-7.38(m,2H),7.28(t,J＝7.3Hz,5H),7.22(dd,J＝9.7,7.6Hz,1H),5.46(s,2H),4.87(s,3H),2.36(s,3H).

¹³C NMR(101MHz,CDCl₃)δ153.38,152.65,144.70,139.05,134.26,131.24,131.01,129.92,128.76,128.50,127.88,127.59,127.09,126.38,120.65,118.60,46.31,28.75,15.55.

HRMS (ESI +): molecular formula C₂₃H₂₁N₃，[M+H]⁺340.1817, found: 340.1817.

the structural formula of the derivative prepared in example 10 is

The name of the compound is N-benzyl-4- (4-ethyl phenyl) phthalazine-1-amine, the yield is 36.23%, and the melting point is 223.2-224.1 ℃.

Example 11 test results:¹H NMR(400MHz,CDCl₃)δ8.03-7.98(m,1H),7.91-7.85(m,1H),7.78-7.70(m,2H),7.60(d,J＝8.1Hz,2H),7.47(dd,J＝13.1,4.3Hz,2H),7.37-7.27(m,5H),5.64(s,1H),4.95(s,2H),2.75-2.64(m,1H),1.78-1.63(m,2H),0.99(t,J＝7.3Hz,3H).

¹³C NMR(101MHz,CDCl₃)δ153.32,152.68,143.16,139.08,134.24,131.24,131.03,129.83,128.70,128.47,127.52,127.04,126.38,120.82,118.65,46.23,37.92,24.53,13.91.

HRMS (ESI +): molecular formula C₂₄H₂₃N₃，[M+H]⁺354.1977, found: 354.1977.

the structural formula of the derivative prepared in example 11 is

The name of the compound is N-benzyl-4- (4-propylphenyl) phthalazine-1-amine, the yield is 39.22%, and the melting point is 198.9-199.3 ℃.

Example 12 test results:¹H NMR(400MHz,CDCl₃)δ8.01(dd,J＝7.1,2.2Hz,1H),7.85-7.74(m,3H),7.68-7.63(m,2H),7.51(d,J＝7.1Hz,2H),7.39(t,J＝7.3Hz,2H),7.33(d,J＝7.2Hz,1H),7.07–7.01(m,2H),5.49-5.31(m,1H),4.97(s,2H),3.90(s,3H).

¹³C NMR(101MHz,CDCl₃)δ159.95,152.59,139.08,131.20,130.99,128.77,128.51,127.61,127.03,120.59,113.83,77.29,77.03,76.71.

HRMS (ESI +): molecular formula C₂₂H₁₉N₃O，[M+H]⁺342.1643, found: 342.1644.

the structural formula of the derivative prepared in example 12 is

The name is N-benzyl-4- (4-methoxyphenyl) phthalazine-1-amine, the yield is 34.99 percent, and the melting point is 144.2-145.7 ℃.

Example 13 test results:¹HNMR(400MHz,CDCl₃)δ8.02-7.96(m,1H),7.90(d,J＝8.0Hz,1H),7.83-7.71(m,2H),7.52(t,J＝9.9Hz,3H),7.46(d,J＝7.6Hz,1H),7.38(dt,J＝10.5,7.5Hz,3H),7.30(dd,J＝9.6,7.4Hz,2H),5.92-5.55(m,1H),4.97(s,2H),2.44(s,3H).

¹³C NMR(101MHz,CDCl₃)δ153.44,152.69,138.94,138.12,136.71,131.37,131.14,130.62,129.32,128.75,128.48,128.16,127.59,127.03,126.39,120.85,118.64,46.26,21.51.

HRMS (ESI +): molecular formula C₂₂H₁₉N₃，[M+H]⁺326.1666, found: 326.1666.

the structural formula of the derivative prepared in example 13 is

The name is N-benzyl-4- (m-tolyl) phthalazine-1-amine, the yield is 35.71%, and the melting point is 205.7-206.7 ℃.

Example 14 test results:¹HNMR(400MHz,CDCl₃)δ8.12-8.06(m,1H),7.96(d,J＝7.0Hz,1H),7.83-7.75(m,2H),7.59-7.54(m,2H),7.54-7.48(m,2H),7.36(t,J＝7.2Hz,2H),7.33-7.26(m,6H),7.22-7.14(m,6H),7.09-7.02(m,2H),6.06-5.70(m,1H),4.97(s,2H).

¹³C NMR(101MHz,CDCl₃)δ152.46,148.39,147.54,131.50,131.25,130.80,129.37,128.76,128.46,127.62,127.10,124.84,123.28,122.79,118.85,29.71.

HRMS (ESI +): molecular formula C₃₃H₂₆N₄，[M+H]⁺479.2239, found: 479.2239.

the structural formula of the derivative prepared in example 14 is

The name of the compound is N-benzyl-4- (4- (diphenylamino) phenyl) phthalazine-1-amine, the yield is 30.34%, and the melting point is 249.0-249.7 ℃.

Example 15 test results:¹H NMR(400MHz,CDCl₃)δ8.04(dd,J＝7.0,2.3Hz,1H),7.85(d,J＝7.2Hz,1H),7.80-7.69(m,2H),7.64(d,J＝8.3Hz,2H),7.52(dd,J＝10.9,7.9Hz,4H),7.34(dt,J＝24.4,7.2Hz,3H),5.50(s,1H),4.97(s,4H),1.39(s,9H).

¹³C NMR(101MHz,CDCl₃)δ153.32,152.66,151.53,139.08,134.00,131.20,131.00,129.63,128.74,128.51,127.57,127.13,126.36,125.33,120.66,118.60,46.31,34.75,31.40.

HRMS (ESI +): molecular formula C₂₅H₂₃N₃，[M+H]⁺368.2120, found: 368.2120.

example 15 the derivative prepared has the formula

The name of the compound is N-benzyl-4- (4- (tert-butyl) phenyl) phthalazine-1-amine, the yield is 34.06%, and the melting point is 165.1-166.1 ℃.

Example 16 test results:¹H NMR(400MHz,DMSO-d₆)δ8.49(d,J＝8.1Hz,1H),8.19(t,J＝5.8Hz,1H),8.01-7.83(m,3H),7.66(dd,J＝8.0,1.5Hz,2H),7.62-7.54(m,3H),7.43(d,J＝8.6Hz,2H),6.95(d,J＝8.6Hz,2H),4.84(d,J＝5.8Hz,2H),3.78(s,3H).

¹³C NMR(101MHz,DMSO-d₆)δ158.58,153.25,151.86,131.74,130.59,130.04,129.76,129.15,128.81,126.44,125.99,122.88,114.06,113.61,55.50,44.08.

HRMS (ESI +): molecular formula C₂₂H₁₉N₃O，[M+H]⁺342.1610, found: 342.1613.

the structural formula of the derivative prepared in example 16 is

The name is N- (4-methoxybenzyl) -4-phenylphthalazine-1-amine, the yield is 43.32%, and the melting point is 136.2-136.6 ℃.

Example 17 test results:¹HNMR(400MHz,CDCl₃)δ8.01(dd,J＝7.2,2.1Hz,1H),7.86(t,J＝5.5Hz,1H),7.79-7.71(m,2H),7.60(d,J＝8.0Hz,2H),7.43(d,J＝8.6Hz,2H),7.33(d,J＝8.0Hz,2H),6.90(d,J＝8.6Hz,2H),5.76-5.36(m,1H),4.88(s,2H),3.81(s,3H),2.76–2.56(m,2H),1.75-1.65(m,2H),1.37(tt,J＝8.7,4.2Hz,4H),0.99-0.83(m,3H).

¹³C NMR(101MHz,CDCl₃)δ159.15,153.27,152.61,143.51,134.02,131.32,131.03,129.85,128.46,127.13,126.42,120.84,118.72,114.13,55.35,45.82,35.81,31.57,31.13,22.59,14.08.

HRMS (ESI +): molecular formula C₂₇H₂₉N₃O，[M+H]⁺412.2395, found: 412.2397.

the structural formula of the derivative prepared in example 17 is

The name is N- (4-methoxybenzyl) -4- (4-pentylphenyl) phthalazin-1-amine, the yield is 38.01 percent, and the melting point is 87.2-87.9 ℃.

Example 18 test results:¹H NMR(400MHz,DMSO-d₆)δ8.41(d,J＝8.2Hz,1H),8.09(s,1H),7.94–7.80(m,3H),7.52(d,J＝8.6Hz,2H),7.36(d,J＝8.5Hz,2H),7.08(d,J＝8.6Hz,2H),6.89(d,J＝8.6Hz,2H),4.77(d,J＝5.6Hz,2H),4.02(t,J＝6.5Hz,2H),3.72(s,2H),1.78(d,J＝7.1Hz,2H),1.02(t,J＝7.4Hz,3H).

¹³C NMR(101MHz,DMSO-d₆)δ158.62,158.37,151.62,132.31,132.04,131.38,130.77,129.73,129.16,127.76,119.37,118.75,115.18,114.78,114.08,113.60,69.51,56.17,55.51,22.52,10.91.

HRMS (ESI +): molecular formula C₂₅H₂₅N₃O₂，[M+H]⁺400.2023, found: 400.2025.

example 18 the derivative prepared has the formula

The name is N- (4-methoxybenzyl) -4- (4-propoxyphenyl) phthalazin-1-amine, the yield is 41.29%, and the melting point is 139.3-140.0 ℃.

Example 19 test results:¹H NMR(400MHz,CDCl₃)δ8.04-7.91(m,2H),7.75(ddd,J＝14.3,7.2,1.4Hz,2H),7.65-7.57(m,2H),7.48(d,J＝7.4Hz,2H),7.45-7.39(m,4H),7.34(dd,J＝6.1,3.5Hz,1H),7.15-7.03(m,2H),6.88(dd,J＝9.1,2.4Hz,2H),5.99–5.69(m,1H),5.14(s,2H),4.89(s,2H),3.80(s,3H).

¹³C NMR(101MHz,CDCl₃)δ159.17,152.98,152.64,136.87,132.11,131.45,131.21,130.90,129.82,129.42,128.66,128.06,127.55,127.01,126.42,121.17,118.84,114.77,114.10,70.12,55.34,45.71.

HRMS (ESI +): molecular formula C₂₉H₂₅N₃O₂，[M+H]⁺448.2030, found: 448.2030.

the structural formula of the derivative prepared in example 19 is

The name of the compound is 4- (4- (benzyloxy) phenyl) -N- (4-methoxybenzyl) phthalazin-1-amine, the yield is 24.97%, and the melting point is 192.1-192.5 ℃.

Example 20 test results:¹HNMR(400MHz,CDCl₃)δ8.02-7.96(m,1H),7.80(s,1H),7.75-7.65(m,6H),7.64-7.58(m,2H),7.48-7.36(m,4H),7.33(s,1H),6.84(d,J＝8.5Hz,2H),5.47(s,1H),4.84(s,2H),3.74(s,3H).

¹³C NMR(400MHz,CDCl₃)δ159.17,152.89,141.35,140.73,131.39,131.12,130.37,129.87,128.87,127.52,127.20,127.01,118.61,114.15,55.35,45.80.

HRMS (ESI +): molecular formula C₂₈H₂₃N₃O，[M+H]⁺418.1923, found: 418.1923.

example 20 the derivative prepared has the formula

Named 4- ([1,1' -biphenyl)]-4-yl) -N- (4-methoxybenzyl) phthalazin-1-amine, with a yield of 26.1% and a melting point of 260.2-260.6 ℃.

Example 21 test results:¹H NMR(400MHz,DMSO-d₆)δ8.43(d,J＝8.2Hz,1H),8.13(t,J＝5.6Hz,1H),7.97-7.83(m,3H),7.62(d,J＝8.5Hz,2H),7.46(t,J＝7.9Hz,2H),7.37(d,J＝8.4Hz,2H),7.26–7.10(m,5H),6.89(d,J＝8.5Hz,2H),4.78(d,J＝5.7Hz,2H),3.72(s,3H).

¹³C NMR(101MHz,DMSO-d₆)δ158.58,156.70,153.20,151.25,132.35,131.78,130.58,129.79,129.15,128.34,124.37,119.65,119.03,118.52,114.06,113.58,55.50,44.08.

HRMS (ESI +): molecular formula C₂₈H₂₃N₃O₂，[M+H]⁺434.1872, found: 434.1872.

the structural formula of the derivative prepared in example 21 is

The name is N- (4-methoxybenzyl) -4- (4-phenoxyphenyl) phthalazine-1-amine, the yield is 30.38%, and the melting point is 134.0-134.5 ℃.

Example 22 test results:¹H NMR(400MHz,CDCl₃),δ8.43(d,J＝8.2Hz,1H),8.16(s,1H),7.96–7.89(m,1H),7.85(d,J＝6.2Hz,2H),7.61(dt,J＝12.7,6.4Hz,4H),7.37(d,J＝8.4Hz,2H),6.89(d,J＝8.3Hz,2H),6.86–6.80(m,1H),5.96(d,J＝17.7Hz,1H),5.36(d,J＝11.0Hz,1H),4.78(d,J＝5.6Hz,2H),3.72(s,3H).

¹³C NMR(400MHz,CDCl₃)δ159.14,152.80,137.74,136.49,131.32,131.05,130.15,129.88,126.83,126.23,120.72,118.53,114.54,114.13,55.36,45.79,29.72.

HRMS (ESI +): molecular formula C₂₄H₂₁N₃O，[M+H]⁺368.1760, found: 368.1762.

the structural formula of the derivative prepared in example 22 is

The name is N- (4-methoxybenzyl) -4- (4-vinyl phenyl) phthalazine-1-amine, the yield is 34.79%, and the melting point is 142.1-142.7 ℃.

Example 23 test results:¹HNMR(400MHz,CDCl₃)δ8.03(d,J＝7.4Hz,1H),7.98-7.90(m,1H),7.78-7.68(m,2H),7.51(dd,J＝24.7,4.7Hz,2H),7.40(d,J＝8.6Hz,2H),7.29(d,J＝7.9Hz,2H),6.83(d,J＝8.6Hz,2H),6.15(s,1H),4.88(s,2H),3.77(s,3H),2.43(s,3H).

¹³C NMR(101MHz,CDCl₃)δ159.01,153.04,152.57,138.51,133.67,131.51,131.26,130.85,129.78,129.08,126.95,126.43,121.47,118.91,114.01,55.31,45.57,21.38.

HRMS (ESI +): molecular formula C₂₃H₂₁N₃O，[M+H]⁺356.1766, found: 356.1766.

the structural formula of the derivative prepared in example 23 is

The name is N- (4-methoxybenzyl) -4- (p-tolyl) phthalazin-1-amine, the yield is 36.92%, and the melting point is 147.2-147.9 ℃.

Example 24 test results:¹H NMR(400MHz,CDCl₃)δ7.65(d,J＝8.2Hz,1H),7.61-7.55(m,2H),7.55-7.49(m,3H),7.49-7.45(m,4H),7.45-7.39(m,1H),7.39-7.34(m,2H),7.33-7.29(m,2H),7.23-7.17(m,2H),7.09-6.89(m,3H),5.41(s,1H),4.93(dd,J＝38.9,14.1Hz,2H).

¹³C NMR(101MHz,CDCl₃)δ159.17,153.98,152.70,141.87,141.00,135.39,131.36,131.33,129.86,129.17,127.89,127.54,126.96,126.57,120.56,117.97,114.15,55.36,45.77.

HRMS (ESI +): molecular formula C₂₈H₂₃N₃O，[M+H]⁺418.1925, found: 418.1925.

the structural formula of the derivative prepared in example 24 is

Named 4- ([1,1' -biphenyl)]-2-yl) -N- (4-methoxybenzyl) phthalazin-1-amine, with a yield of 34.85% and a melting point of 123.2-123.7 ℃.

Example 25 test results:¹H NMR(400MHz,CDCl₃)δ8.04-7.98(m,1H),7.84-7.78(m,1H),7.78-7.69(m,2H),7.66–7.60(m,2H),7.46-7.41(m,2H),7.35(d,J＝8.2Hz,2H),6.93–6.87(m,2H),5.37(s,1H),4.88(d,J＝3.4Hz,2H),3.81(s,3H),2.75(q,J＝7.6Hz,2H),1.36–1.29(m,3H).

¹³C NMR(101MHz,CDCl₃)δ159.07,153.24,152.72,144.63,134.35,131.17,130.92,129.88,127.87,126.97,126.35,120.71,118.59,114.08,55.33,45.74,28.75,15.56.

HRMS (ESI +): molecular formula C₂₄H₂₃N₃O，[M+H]⁺370.1923, found: 370.1924.

the structural formula of the derivative prepared in example 25 is

The name is 4- (4-ethyl phenyl) -N- (4-methoxy benzyl) phthalazine-1-amine, the yield is 39.54 percent, and the melting point is 176.4-176.7 ℃.

Example 26 test results:¹HNMR(400MHz,CDCl₃)δ8.05-7.93(m,1H),7.88(dd,J＝5.6,3.1Hz,1H),7.79-7.68(m,2H),7.66-7.56(m,2H),7.48-7.37(m,2H),7.33(t,J＝12.7Hz,2H),6.86(d,J＝8.3Hz,2H),5.65(s,1H),4.87(s,2H),3.78(s,3H),2.76-2.60(m,2H),1.83-1.59(m,2H),0.99(t,J＝7.3Hz,3H).

HRMS (ESI +): molecular formula C₂₅H₂₅N₃O，[M+H]⁺384.2075, found: 384.2075.

example 26 the derivative prepared has the formula

The name is N- (4-methoxybenzyl) -4- (4-propylphenyl) phthalazin-1-amine, the yield is 34.94%, and the melting point is 138.2-139.7 ℃.

Example 27 test results:^.1H NMR(400MHz,CDCl₃)δ8.05-7.95(m,1H),7.83(dd,J＝6.7,2.5Hz,1H),7.79-7.69(m,2H),7.69-7.59(m,2H),7.43(d,J＝8.6Hz,2H),7.09-7.01(m,2H),6.94-6.87(m,2H),5.47–5.31(m,1H),4.88(s,2H),3.89(s,3H),3.81(s,3H).

¹³C NMR(400MHz,CDCl₃)δ159.93,159.12,152.91,152.58,131.38,129.86,129.46,126.97,126.39,120.70,118.65,114.12,113.82,55.37,45.78.

HRMS (ESI +): molecular formula C₂₃H₂₁N₃O₂，[M+H]⁺372.1716, found: 372.1716.

the structural formula of the derivative prepared in example 27 is

The name is N- (4-methoxybenzyl) -4- (4-methoxyphenyl) phthalazine-1-amine, the yield is 32%, and the melting point is 96.4-97.1 ℃.

Example 28 test results:¹HNMR(400MHz,CDCl₃)δ7.97(d,J＝8.3Hz,1H),7.92-7.84(m,1H),7.81-7.68(m,2H),7.53(d,J＝11.2Hz,1H),7.41(qd,J＝12.1,6.2Hz,4H),7.28(d,J＝7.5Hz,1H),6.89(d,J＝7.1Hz,2H),5.86-5.40(m,1H),4.88(s,2H),3.80(s,3H),2.44(s,3H).

¹³C NMR(101MHz,CDCl₃)δ159.10,153.32,152.69,138.10,136.78,131.29,131.06,130.62,129.86,129.27,128.14,127.01,126.36,120.84,118.62,114.09,55.34,45.72,21.52.

HRMS (ESI +): molecular formula C₂₃H₂₁N₃O，[M+H]⁺356.1765, found: 356.1765.

example 28 the derivative prepared has the formula

The name is N- (4-methoxybenzyl) -4- (m-tolyl) phthalazin-1-amine, the yield is 40.48%, and the melting point is 221.1-221.7 ℃.

Example 29 test results:¹HNMR(400MHz,CDCl₃)δ8.12-8.05(m,1H),7.94(s,1H),7.77(dd,J＝9.2,5.0Hz,2H),7.56(d,J＝8.4Hz,2H),7.42(t,J＝11.3Hz,2H),7.28(dd,J＝12.3,4.9Hz,3H),7.15(ddd,J＝21.3,8.5,2.7Hz,6H),7.06(t,J＝7.3Hz,2H),6.88(d,J＝7.3Hz,2H),6.02-5.65(m,1H),4.89(s,2H),3.80(d,J＝1.0Hz,3H).

¹³C NMR(400MHz,CDCl₃)δ159.01,153.04,152.57,138.51,133.67,131.51,131.26,130.85,129.78,129.08,126.95,126.43,121.47,118.91,114.01,55.31,45.57,21.38.

HRMS (ESI +): molecular formula C₃₄H₂₈N₄O，[M+H]⁺510.2377, found: 510.2377.

the structural formula of the derivative prepared in example 29 is

The name is 4- (4- (diphenylamino) phenyl) -N- (4-methoxybenzyl) phthalazine-1-amine, the yield is 40.48%, and the melting point is 264.0-264.5 ℃.

Example 30 test results:¹H NMR(400MHz,CDCl₃)δ8.03(dd,J＝6.9,2.4Hz,1H),7.83(d,J＝7.9Hz,1H),7.78–7.70(m,2H),7.64(d,J＝8.4Hz,2H),7.55–7.50(m,2H),7.43(d,J＝8.5Hz,2H),6.90(d,J＝8.5Hz,2H),5.43(s,1H),4.89(s,2H),3.81(s,3H),1.39(s,9H).

¹³C NMR(101MHz,CDCl₃)δ159.12,153.24,152.64,151.51,134.01,131.36,129.88,129.63,127.09,126.35,125.32,120.67,118.60,114.12,55.35,45.80,34.75,31.39.

HRMS (ESI +): molecular formula C₂₆H₂₇N₃O，[M+H]⁺398.2225, found: 398.2225.

the structural formula of the derivative prepared in example 30 is

The name is 4- (4- (tert-butyl) phenyl) -N- (4-methoxybenzyl) phthalazine-1-amine, the yield is 40.48%, and the melting point is 239.1-240.6 ℃.

Example 31 test results:¹H NMR(400MHz,CDCl₃)δ7.94(s,1H),7.69(dd,J＝12.7,10.1Hz,3H),7.54(d,J＝7.4Hz,2H),7.36(d,J＝7.8Hz,2H),7.25(d,J＝7.5Hz,2H),6.84(d,J＝7.7Hz,2H),5.28(s,1H),4.82(s,2H),3.74(s,3H),2.63(t,J＝7.7Hz,2H),1.60(d,J＝7.4Hz,2H),1.26(s,6H),0.83(t,J＝5.9Hz,3H).

¹³C NMR(101MHz,CDCl₃)δ159.11,152.95,151.90,144.01,132.40,131.95,129.97,128.48,127.26,126.62,114.06,55.28,35.84,31.75,31.37,29.05,22.64,14.13.

HRMS (ESI +): molecular formula C₂₈H₃₁N₃O，[M+H]⁺425.2467, found: 425.2467.

the structural formula of the derivative prepared in example 31 is

Named 4- (4-hexylphenyl) -N- (4-methoxybenzyl) phthalazin-1-amine as a yellow oily liquid in 34.48% yield.

Method for detecting influence of derivative on cell proliferation capacity by using CCK-8 kit

The CCK-8 kit can evaluate the proliferation activity of cells by detecting the absorbance at the wavelength of 450 nm. SNU398 and LO2 cells were digested to prepare a cell suspension, which was diluted with medium to a density of about 5X 10³One well, 100. mu.l of cell fluid was added to each well, and a blank group and a negative control group were set. Weighing about 1.0000mg of the derivative, placing in 0.5mL EP tube, and preparing into 5 × 10 with DMSO (dimethyl sulfoxide)^-2Mother liquor of M, -20 deg.CAnd (4) storing.

The 31 compound stock solution was diluted to a final concentration of 100. mu. mol/L using medium. The 96-well plate stock medium was aspirated off, and 100. mu.L each of 31 compound dilutions were added, and three duplicate wells were set in parallel. Culturing in a cell culture box for 48 h. A10% CCK-8 test solution was prepared, 100. mu.L of each well was added to a 96-well plate after the solution was aspirated, and after 2 hours of incubation, OD at a wavelength of 450nm was measured to calculate the Cell survival rate (Cell Viability) and the inhibition rate, and the results are shown in Table 2.

TABLE 2 results of preliminary screening of the influence of the derivatives prepared in examples 1 to 31 and N-benzyl-4-phenylphthalazin-1-amine on cell proliferation Activity

In the preliminary screening results, compounds with an inhibitory rate of more than 80% were selected, and cell-level rescreening was performed by stepwise dilution with concentration gradients so that the final concentrations of the compounds in the cell matrix were 100. mu. mol/L, 20. mu. mol/L, 4. mu. mol/L, 0.8. mu. mol/L, and 0.16. mu. mol/L. Analyzing the data to determine IC₅₀Value, measure the ability of a compound to intervene in the proliferation of tumor cells, the greater the ability to intervene, the IC₅₀The lower the value, the resulting IC₅₀The values are listed in table 3.

TABLE 3 rescreening results of the Effect of derivatives on cell proliferation Activity

The results in tables 2 and 3 show that the derivatives prepared in examples 1 to 31 and N-benzyl-4-phenylphthalazin-1-amine have a high inhibitory effect on a liver cancer cell strain SNU398, and particularly, example 27 has a good selectivity on liver cancer cells with high expression of RECQL 4.

Although the present invention has been described in detail with reference to the above embodiments, it is only a part of the embodiments of the present invention, not all of the embodiments, and other embodiments can be obtained without inventive step according to the embodiments, and the embodiments are within the scope of the present invention.

Claims

1. An N-benzyl-4-phenylphthalazin-1-amine derivative having the formula shown in formula I:

R₁is-H or-OCH₃；

R₂is-H, phenyl, substituted phenyl, alkyl, alkoxy, alkenyl or diphenylamino;

R₃is-H, nitro or methyl;

R₄is-H or phenyl;

R₁、R₂、R₃and R₄Not being simultaneously-H.

2. The N-benzyl-4-phenylphthalazin-1-amine derivative of claim 1 wherein said substituted phenyl group comprises phenoxy or

3. The N-benzyl-4-phenylphthalazin-1-amine derivative according to claim 1, wherein the alkyl group comprises an alkyl group having 1 to 6 carbon atoms;

the alkoxy group includes an alkoxy group having 1 to 3 carbon atoms.

4. The N-benzyl-4-phenylphthalazin-1-amine derivative of claim 3 wherein said alkyl group comprises methyl, ethyl, propyl, tert-butyl, pentyl or hexyl;

the alkoxy group includes-OCH₃or-OC₃H₇；

The alkenyl group includes a vinyl group.

5. The N-benzyl-4-phenylphthalazin-1-amine derivative according to claim 1, having a structure represented by any one of formulas 1 to 31:

6. a process for the preparation of N-benzyl-4-phenylphthalazin-1-amine derivatives as claimed in any one of claims 1 to 5, comprising the steps of:

the reactant 1 is

Wherein R is₁is-H or-OCH₃；

The reactant 2 is

7. The preparation method according to claim 6, wherein the molar ratio of the reactant 1 to the reactant 2 is 1: 1.1-1.7.

the molar ratio of the reactant 1 to the soluble alkali is 1: 2.8-3.2.

8. The production method according to claim 6, wherein the mixed solvent of the organic solvent and water comprises toluene, ethanol and water;

9. The preparation method according to claim 6, wherein the temperature of the coupling reaction is 100-110 ℃, and the time of the coupling reaction is 15-17 h.

10. A medicine for treating liver cancer specifically expressed by RECQL4 comprises an activity inhibitor and a medicine carrier, wherein the activity inhibitor comprises N-benzyl-4-phenylphthalazin-1-amine derivatives or N-benzyl-4-phenylphthalazin-1-amine;