Application of 3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno)propionamide in the preparation of tyrosinase inhibitors

Abstract

The present invention belongs to the field of pharmacy, and specifically relates to a 3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno) propionamide compound and its application in the preparation of tyrosinase inhibitors. The compound of the present invention is prepared by a metal-free catalytic olefin azidation selenation method, is a red liquid, and has a molecular formula of C 17 H 14 ClN5OSe.

Description

[0001] This application is a divisional application with application number 202110512784.6, application date May 11, 2021, and the invention name at the time of application is “3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno)propanamide”. Technical Field

[0002] The present invention relates to the field of pharmacy, and in particular to a 3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno) propionamide compound and application thereof. Background Art

[0003] Organic azide compounds have a wide range of applications in the fields of fine chemicals and pharmaceutical industries. The azide group is not only a good functional group for organic synthesis transformation, but also a functional group in drugs. The carbon-N3 bond is an important synthetic method that provides a method for introducing nitrogen atoms into various organic molecules. Therefore, how to introduce N3 into organic molecules has always been a hot topic for organic chemists. Therefore, there is a constant need to develop sustainable and practical methods to form carbon-N3 bonds in synthetic organic chemistry. On the other hand, nitrogen-containing heterocycles are key skeleton structures for many drug molecules, especially the structurally diverse quinoline-2,4-diones, which are widely present in many natural products, drugs and agrochemicals. Although some methods for constructing quinoline-2,4-diones have been reported, metal-free radical tandem carbocyclization reactions are undoubtedly one of the best options for obtaining atom and step economy and thus obtaining these complex molecules.

[0004] Organoselenium compounds are important intermediates in synthetic chemistry and exist as core structural skeletons in a considerable number of biologically active drug molecules. Organoselenium compounds have a wide range of applications in organic synthesis, medicine, organic materials, etc. Therefore, the development of green, novel, efficient and practical methods for synthesizing organoselenium compounds has attracted widespread attention from chemists. Summary of the invention

[0005] The technical problem to be solved by the present invention is to provide a 3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno) propionamide compound and its application.

[0006] The technical solution of the present invention to solve the above technical problems is as follows:

[0007] One of the purposes of the present invention is to provide a compound 3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno)propionamide, whose chemical structure 2 is as follows:

[0008]

[0009] The preparation method of the above compound is as follows: using N-(3-chloro-2-cyanophenyl)methacrylamide (1) as a substrate, the reaction of (PhSe)2 and PhI(OAc)2 pre-activated TMSN3 in DMSO effectively obtains the azidation-selenation product of the olefin through an azidation-selenation process. After the reaction is completed by shaking for 12 hours, the mixture is quenched with H2O and extracted with CH2Cl2, and then the organic solvent is concentrated in vacuo, and the residue is purified by flash column chromatography using petroleum ether and ethyl acetate as eluents to obtain 3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno)propionamide (2).

[0010] Further, the reaction equation is:

[0011]

[0012] Furthermore, the compound 3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno)propionamide (2) is used in the preparation of anti-tumor drugs.

[0013] Furthermore, the compound 3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno)propionamide (2) is used in the preparation of tyrosinase inhibitor drugs and cosmetics.

[0014] The beneficial effects of the present invention are: a method for azidation and selenization of olefins without metal catalysis, and application of the obtained new compound in the preparation of anti-tumor and tyrosinase inhibitor drugs and cosmetics.

[0015] 3-Azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno)propionamide (2) Structure analysis:

[0016] like Figure 1-3 As shown, the compound 2 of the present invention 1 H NMR, 13 The structure of the compound was known from C NMR and HR-ESI-MS spectra. Specifically, compound 2 was a red liquid with Rf (petroleum ether:ethyl acetate) = 0.42. 1 H NMR (600MHz, CDCl3): δ8.79(s,1H),8.27(dd,J=8.5,0.5Hz,1H),7.60-7.58(m,2H),7.50(t,J=8.3Hz,1H),7.42-7.39 (m,1H),7.32-7.30(m,2H),7.23(dd,J=8.1,0.8Hz,1H),3.92(d,J=12.6Hz,1H),3.69(d,J=12.6Hz,1H),1.69(s,3H).13 C NMR (150MHz, CDCl3): δ170.76,142.11,137.59,136.73,134.51,130.07,129.48 ,125.21,124.80,118.60,113.76,103.24,58.04,51.18,22.45.HRMS(ESI)calcd for C 17 H 14 ClN5OSe[M+Na] + :441.9944,found:441.9944. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 Compound 2 1 H NMR spectrum;

[0018] Figure 2 Compound 2 13 C NMR spectrum;

[0019] Figure 3 HR-ESI-MS spectrum of compound 2. DETAILED DESCRIPTION

[0020] The present invention is further described below by way of examples, so that those skilled in the art can more fully understand the present invention, but the present invention is not limited in any way.

[0021] Example 1

[0022] At room temperature, N-(3-chloro-2-cyanophenyl)methacrylamide (1) (0.3 mmol, 66 mg) and azidotrimethylsilane TMSN3 (93%, 3.0 equiv, 0.9 mmol, 127 μL) pre-activated by diphenyl diselenide (PhSe)2 (0.3 mmol, 93.6 mg) and iodophenyl diethyl ester PhI(OAc)2 (0.45 mmol, 145 mg) were added to a reaction tube in DMSO (2 mL). The reaction tube was then stirred at room temperature for 12 hours until the starting material was completely consumed as monitored by TLC analysis. After the reaction was completed, the mixture was quenched with H2O (15 mL) and extracted with CH2Cl2 (3×5 mL). Then, the organic solvent was concentrated in vacuo. The residue was purified by flash column chromatography using petroleum ether and ethyl acetate as eluents, Rf = 0.42 to obtain 3-azido-N-(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno)propionamide (2), yield: 55%, 69 mg. The reaction equation is:

[0023]

[0024] Example 2

[0025] 1. Tumor cell culture: Human liver cancer cells (HepG-2), human lung cancer cells (A549), human breast cancer cells (MCF-7), human gastric cancer cells (SGC-7901), and human ovarian cancer cells (SKOV-3) were cultured in RPMI 1640 medium containing 10% fetal bovine serum and 1% penicillin-streptomycin. Trypsin was used for digestion and subculture, and cells in the logarithmic growth phase were selected for experiments.

[0026] 2. Drug preparation: Dissolve the drug in DMSO and prepare a 1 mg / mL stock solution, which is stored at -80°C for future use. Dilute the prepared culture medium to the required concentration for the experiment (DMSO final concentration <0.01%) before the experiment.

[0027] 3. Cell proliferation inhibition assay

[0028] The MTT colorimetric method was used to detect the inhibitory effect of compound 2 on the proliferation of various tumor cells. 100 μL of tumor cells in the logarithmic growth phase were inoculated in a 96-well plate, with a cell volume of approximately 1×10 5 / mL. After incubation for 24 hours, 10 μL of compound 2 with a final concentration of 1.25 μg / mL, 2.5 μg / mL, 5 μg / mL, 10 μg / mL, and 20 μg / mL was added to each well, and an equal volume of RPMI 1640 medium was added to the negative control group. Three parallel wells were set up for each group. The 96-well plate was placed in a 37°C constant temperature CO2 incubator for 48 hours, 20 μL of 0.5% MTT solution was added to each well, and the incubation continued in the incubator for 4-6 hours. The supernatant was discarded, 100 μL of DMSO was added to each well, and after sufficient shaking, the absorbance value (OD value) was measured at 570 nm, and the half inhibitory concentration (IC 50 ), and the experiment was repeated 3 times.

[0029] The calculation formula of the inhibition rate of compound 2 on tumor cell proliferation is:

[0030] Proliferation inhibition rate (%) = (1-OD value of experimental group / OD value of blank control group) * 100%

[0031] 4. Experimental Results

[0032] The experimental results are shown in Table 1. After 72 hours of action of different concentrations of compound 2 on human liver cancer HepG-2 cells, human lung cancer A549 cells, human breast cancer MCF-7 cells, human gastric cancer SGC-7901 cells, and human ovarian cancer SKOV-3 cells, the proliferation of the above tumor cells was inhibited to varying degrees. With the increase of drug concentration, the proliferation inhibition rate of each tumor cell gradually increased, showing concentration dependence.

[0033] Table 1 Inhibitory effect of compound 2 on proliferation of different tumor cells

[0034]

[0035] Embodiment 3:

[0036] Determination of tyrosinase inhibitory activity

[0037] 1. Preparation of sample solution and reference solution: Compound 2 and positive reference substances citric acid and arbutin were prepared into 2 mg / mL stock solution with phosphate buffer-dimethyl sulfoxide (PBS-DMSO) mixture, and then diluted into 1, 0.5, 0.25, and 0.125 mg / mL solutions in sequence.

[0038] 2. Take 80 μL of sample solutions of different concentrations and citric acid and arbutin solutions, add 280 μL PBS (pH 6.8) and 80 μL of 0.1 mg / mL L-tyrosine solution in turn, mix well, and react in a 25°C water bath for 10 minutes. Then add 40 μL tyrosinase (100 U / mg) and mix well. After reacting in a 25°C water bath for 10 minutes, take out and use a UV spectrophotometer to measure the absorbance at 475 nm. Using each blank control as a reference, calculate the inhibition rate of the sample solution and citric acid and arbutin solutions on tyrosinase, and calculate the half inhibition concentration (IC) according to the dose-effect curve. 50 ).

[0039] Inhibition rate (%) = (A0-A1) / A0×100%

[0040] Wherein: A0 is the absorbance of the standard control, that is, the absorbance of the standard control solution composed of PBS buffer, L-tyrosine solution and tyrosinase; A1 is the absorbance of the sample solution or citric acid, arbutin.

[0041] 3. The test results are shown in Table 2. The inhibitory effects of compound 2, citric acid and arbutin on tyrosinase increase with the increase of concentration. IC 50 The value is 0.865mg / mL, and the IC of citric acid 50 The value is 0.813mg / mL, and the IC of arbutin 50 The value was 0.485 mg / mL. The inhibitory effect of compound 2 on tyrosinase was comparable to that of the positive control citric acid, and slightly worse than that of the positive control arbutin.

[0042] Table 2 Tyrosinase inhibition of compound 2, citric acid and arbutin

[0043]

[0044] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. 3-Azido- N -(3-chloro-2-cyanophenyl)-2-methyl-2-(phenylseleno)propionamide compound in the preparation of anti-tumor drugs, characterized in that: The tumor is selected from human liver cancer cells (HepG-2) and human lung cancer cells (A549); the compound is shown in the structural formula 2: 。

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