Red light driven-spinach chlorophyll mediated novel coumarin and tetrahydroisoquinoline coupling method and application

Abstract

The invention provides a red light driven-spinach chlorophyll mediated novel coumarin and tetrahydroisoquinoline coupling method and application, a tetrahydroisoquinoline compound, a coumarin compound, a chlorophyll photosensitizer and a solvent are mixed according to a ratio, and an illumination reaction is performed in an air atmosphere; coupling the skeleton of the tetrahydroisoquinoline compound with the skeleton of the coumarin compound, and purifying to obtain the compound with both the coumarin skeleton and the tetrahydroisoquinoline skeleton. According to the red light driven-spinach chlorophyll mediated novel coumarin and tetrahydroisoquinoline coupling method and application, operation is easy and convenient, environment friendliness is achieved, and efficient coupling is achieved.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of chemical synthesis, in particular to a novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll and application. BACKGROUND

[0002] Coumarin and tetrahydroisoquinoline are two important medicinal skeleton structures, which are widely present in natural products and drug intermediates. Compounds based on such skeletons have attracted much attention in the field of drug research and development due to their significant pharmacological activities (such as anti-tumor, antibacterial, anti-inflammatory, etc.). In recent years, the modification of substituents, the synthesis of structural analogs and the structure-activity relationship of such compounds have become a hot topic. However, there are still challenges in efficiently constructing coumarin-tetrahydroisoquinoline hybrid molecules, and novel coupling strategies are needed to realize the precise connection of the two skeletons, thereby expanding their application value in medicinal chemistry.

[0003] The tetrahydroisoquinoline skeleton and the coumarin skeleton can generate a coumarin-isoquinoline hybrid molecule through certain chemical methods, and effective synthesis methods for generating a coumarin-tetrahydroisoquinoline hybrid molecule have been reported in the literature. However, traditional synthesis methods usually require the use of metal catalysts or toxic reagents, and may produce toxic by-products or result in high reaction costs. Existing methods have problems such as the need for oxidizing agents, high temperature and pressure conditions, and the possibility of producing toxic by-products or low reaction efficiency.

[0004] In recent years, although different functional group studies on each position of tetrahydroisoquinoline compounds and coumarin compounds have received extensive attention, there are still few reports on green and environmentally friendly photosensitizer catalysis. SUMMARY

[0005] The present application aims to provide a novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll and application, which significantly avoids problems such as high catalyst cost, harsh reaction conditions (such as anhydrous and anaerobic environment, high temperature, high pressure), environmental pollution, and complex operation.

[0006] To achieve the above-mentioned purpose, the present application provides a novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll, in which a tetrahydroisoquinoline compound, a coumarin compound, a chlorophyll photosensitizer and a solvent are mixed in proportion, and a photochemical reaction is carried out under an air atmosphere, so that the skeleton of the tetrahydroisoquinoline compound and the skeleton of the coumarin compound are coupled, and a compound with both a coumarin skeleton and a tetrahydroisoquinoline skeleton is obtained by purification.

[0007] Preferably, the molar ratio of the tetrahydroisoquinoline compound to the coumarin compound is 1:1-3.

[0008] Preferably, the molar amount of the chlorophyll photosensitizer is 2% to 5% of the molar amount of the tetrahydroisoquinoline compound.

[0009] Preferably, the ratio of the solvent to the tetrahydroisoquinoline compound is 0.1 mmol: 1-2 mL.

[0010] Preferably, the solvent is MeOH.

[0011] Preferably, the chlorophyll photosensitizer is spinach extract.

[0012] Preferably, the illumination uses 660~680nm red light.

[0013] Preferably, the reaction temperature of the photo-irradiation reaction is 25-30°C, and the reaction time is 12-24 hours.

[0014] Preferably, the general structural formula of the tetrahydroisoquinoline compound is: ; in, R1 is a C1-C16 alkyl, aromatic heterocyclic or... Any of the substituted aromatic rings; In this context, M1, M2, M3, M4, or M5 are each independently any one of H, F, Cl, Br, ester group, cyano group, sulfone group, nitro group, C1-C16 alkyl group, or C1-C16 alkyl group; The general structural formula of the coumarin compounds is: ; Among them, R2, R3, and R4 are each independently any one of H, O, and hydroxyl; R5, R6, R7, and R8 are each independently any one of H, F, Cl, Br, ester group, cyano group, sulfone group, nitro group, C1-C16 alkyl group, and C1-C16 alkyl group.

[0015] Application of compounds with coumarin and tetrahydroisoquinoline skeletons prepared by a novel red light-driven, spinach chlorophyll-mediated coupling method in the treatment of colon cancer.

[0016] Therefore, the present invention employs the above-mentioned novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll, and its application, with the following technical effects: The method of this invention is not only simple to operate and mild, avoiding harsh conditions such as high temperature and high pressure, anhydrous and oxygen-free environments, but also does not require the use of precious metal catalysts, which significantly improves the economy and environmental friendliness of the reaction.

[0017] The method of this invention has a reaction yield of up to 83% or more, and the product is easy to separate and purify, ensuring the acquisition of high purity. Attached Figure Description

[0018] Figure 1 The proton spectrum of LWJ-3-1 in Example 1 of this invention; Figure 2 The carbon spectrum of LWJ-3-1 in Example 1 of this invention; Figure 3 The proton spectrum of LWJ-3-2 in Example 1 of this invention; Figure 4 The carbon spectrum of LWJ-3-2 in Example 1 of this invention; Figure 5 IC50 of compounds LWJ-3-1 and LWJ-3-2 in Caco-2 cells 50 . Detailed Implementation

[0019] The technical solution of the present invention will be further described below with reference to the accompanying drawings and embodiments.

[0020] Unless otherwise defined, the technical or scientific terms used in this invention shall have the ordinary meaning as understood by one of ordinary skill in the art to which this invention pertains.

[0021] The tetrahydroisoquinoline compound shown in Formula I was coupled with coumarin of Formula II under light and solvent conditions using spinach extract as a photosensitizer to synthesize a ketone compound containing a tetrahydroisoquinoline and coumarin skeleton, as shown in Formula III.

[0022] In Formula I: R1 is a C1-C16 alkyl, aromatic heterocyclic or... Any of the substituted aromatic rings; In this context, M1, M2, M3, M4, or M5 are each independently any one of H, F, Cl, Br, ester group, cyano group, sulfone group, nitro group, C1-C16 alkyl group, or C1-C16 alkyl group.

[0023] In Formula II: R2, R3, and R4 are each independently any one of H, O, or hydroxyl; R5, R6, R7, and R8 are each independently any one of H, F, Cl, Br, ester group, cyano group, sulfone group, nitro group, C1-C16 alkyl group, or C1-C16 alkyl group.

[0024] The preparation method of chlorophyll photosensitizer is as follows: Wash and dry fresh spinach, then trim off the stems and leaves. Mix the spinach leaves with an appropriate amount of calcium carbonate, add the mixture to a juicer, then add an appropriate amount of anhydrous ethanol. Juice the spinach and filter the juice.

[0025] Add an appropriate amount of water and saturated saline solution to the filtrate, and then extract with petroleum ether.

[0026] After extraction, the organic phase was taken, the solvent was removed under reduced pressure, and the residue was separated by column chromatography. A dark red band was collected to obtain the photosensitizer chlorophyll, hereinafter referred to as spinach extract photosensitizer PC.

[0027] The purification method is as follows: After the reaction was monitored by TLC until it was complete, the solvent was recovered under reduced pressure, and the solid was purified by petroleum ether / ethyl acetate (3:1) column chromatography to obtain a white solid.

[0028] Example 1 A novel red light-driven, spinach chlorophyll-mediated coupling method for coumarin and tetrahydroisoquinoline includes the following steps: In a reaction tube, phenyl-1,2,3,4-tetrahydroisoquinoline A1 (0.1 mmol, 20.9 mg, 1.0 equiv), 4-hydroxy-2H-chromen-2-one B1 (0.2 mmol, 32.4 mg, 2.0 equiv), spinach extract photosensitizer PC (2.0 mg, 2% mmol), and methanol (2.0 mL) were added sequentially. The reaction was carried out under air atmosphere at room temperature (25 °C) with stirring and irradiation by a 30 W red LED for 24 h. After the reaction was completed by TLC monitoring, the solvent was recovered under reduced pressure. The residue was separated by column chromatography to obtain a white solid C1: 4-hydroxy-3-(2-phenyl-1,2,3,4-tetrahydroisoquinoline-1-yl)-2H-chromen-2-one (2H-one) (30.7 mg), with a yield of 83%.

[0029] In coupling reactions, tetrahydroisoquinoline compounds are electron donors and the main source of the reactive active center, playing a major "driving role"; while coumarins are coupling acceptors, playing a "receiving" or "coupling backbone" role.

[0030] Example 2 The preparation method of Example 1 was used again, with changes to the reagent ratios and reaction conditions, specifically including: 2-Phenylacetyl-1,2,3,4-Tetrahydroisoquinoline: 0.1 mmol, 4-hydroxy-7-methylcoumarin: 0.2 mmol, chlorophyll photosensitizer (spinach extract): 2.0 mg (~2 mol%), solvent: methanol 2.0 mL. Light source: 680nm red LED, 30W; The reaction temperature was 25℃, and the reaction time was 24 hours. Result: The yield of the target product was 83%.

[0031] Example 3 Using the preparation method of Example 1 and the reaction conditions of Example 2, but with changes to the reagent ratios, the preparation was carried out again, specifically including: 2-Phenylacetyl-1,2,3,4-Tetrahydroisoquinoline: 0.1 mmol, 4-hydroxy-7-methylcoumarin: 0.2 mmol, chlorophyll photosensitizer: 0.5 mg (~0.5 mol%), methanol 2.0 mL; Experimental results showed that the yield of the target product was 36.7%.

[0032] Example 4 The preparation method of Example 1 was used, but the ratio of reagents and reaction conditions were changed to prepare the reagents again, specifically including: 2-Phenylacetyl-1,2,3,4-Tetrahydroisoquinoline: 0.1 mmol, 4-hydroxy-7-methylcoumarin: 0.2 mmol, chlorophyll photosensitizer: 0 mg (not added), methanol 2.0 mL, light source: none (dark environment), reaction temperature and reaction time are the same as in Example 2; Experimental results showed that almost no reaction occurred, with no coupling products or extremely low yields.

[0033] Example 5 Using the preparation method of Example 1 and the reaction conditions of Example 2, but with changes to the reagent ratios, the preparation was carried out again, specifically including: 2-Phenylacetyl-1,2,3,4-Tetrahydroisoquinoline: 0.1 mmol, 4-hydroxy-7-methylcoumarin: 0.1 mmol, chlorophyll photosensitizer: 2.0 mg, methanol 2.0 mL. After the reaction, the yield was 48.9%. The experimental results showed that the yield was 41.5%, the target product was less, and unreacted tetrahydroisoquinoline and byproducts were present.

[0034] Example 6 Using the preparation method of Example 1 and the reaction conditions of Example 2, but with changes to the reagent ratios, the preparation was carried out again, specifically including: 2-Phenylacetyl-1,2,3,4-Tetrahydroisoquinoline: 0.1 mmol, 4-hydroxy-7-methylcoumarin: 0.2 mmol, chlorophyll photosensitizer: 2.0 mg, methanol 2.0 mL, other components as in Example 2. Experimental results showed a yield of 83%, with a pure product, representing the optimal ratio.

[0035] Example 7 Using the preparation method of Example 1 and the reaction conditions of Example 2, but with changes to the reagent ratios, the preparation was carried out again, specifically including: 2-Phenylacetyl-1,2,3,4-Tetrahydroisoquinoline: 0.1 mmol, 4-hydroxy-7-methylcoumarin: 0.1 mmol, chlorophyll photosensitizer: 2.0 mg, methanol 2.0 mL. After the reaction, the yield was 48.9%. Excess coumarin caused side reactions and increased the complexity of column chromatography separation.

[0036] like Figure 1 , Figure 2 , Figure 3 and Figure 4 As shown, the structural characterization data of the product of Example 1 are as follows: P1, 88 % yield, 1H NMR (600 MHz, Chloroform - d) δ7.52 (d, J = 8.1 Hz, 1H), 7.39 (t, J = 9.0 Hz, 3 H), 7.34 – 7.29 (m, 2H), 7.15 (dq, J = 14.5 , 7.2 Hz , 4 H ), 7.02 ( s , 1H ), 6.95 ( d, J =8.0 Hz, 1H ), 6.14 ( s, 1H ), 3.71 – 3.66 ( m, 1H ), 3.52 ( ddd, J = 17.0 ,12.4 , 5.2 Hz, 1H ), 3.24 (td, J = 12.3, 3.0 Hz, 1H), 2.94 (d, J = 16.4Hz, 1H), 2.35 (s, 3H). 13C NMR (151 MHz, cdcl3) δ 165.12, 164.04, 153.36, 148.52, 135.63, 132.70, 131.97, 129.64, 128.25, 127.44, 127.01, 127.00, 126.19, 123.61, 123.28, 122.37, 116.45, 116.44, 104.75, 77.24, 77.03, 76.82, 58.26, 54.96, 30.35, 14.11 ppm; High-resolution mass spectrometry calculated value: C14H19NO3+Na 351.1208, measured value: 351.1207. P2, 89 % yield, 1H NMR (600 MHz, Chloroform-d) 7.52(d, J = 8.1 Hz, 1H), 7.39 (t, J = 9.0 Hz, 3H), 7.34-7.29 (m, 2H),7.15 (dq, J = 14.6, 7.2 Hz, 4H ), 7.02 ( s, 1H ), 6.95 ( d, J = 8.0 Hz, 1H), 6.14 ( s, 1H ), 3.68 ( ddd, J = 12.0 , 5.2, 1.9 Hz, 1H ), 3.52 ( ddd, J =17.0, 12.4, 5.2 Hz, 1H ), 3.24 (td, J = 12.3, 3.0 Hz, 1H), 2.94 (d, J =16.4 Hz, 1H), 2.35 (s, 3H); 13C NMR (151 MHz, cdcl3) δ 165.20, 164.26, 153.49, 148.67 , 143.22 , 135.81 , 132.72 , 129.58 , 128.22 , 127.47 ,126.95 , 126.93 , 126.06 , 124.85 , 122.97 , 122.40 , 116.54 , 113.90, 103.99, 77.24, 77.03, 76.82, 58.17, 54.97, 30.39, 21.65 ppm; High-resolution mass spectrometry calculated value: C25H21NO3+H 343.1545, measured value: 343.1546. Drug testing was performed using the product obtained in Example 1. 1. Cell plating (1) Digestion, centrifugation, resuspension Take Caco-2 cells, aspirate the culture medium from the dish, rinse once with 3ml of PBS, add 1ml of trypsin, shake well and place in an incubator for 1min, remove and add 5ml of DMEM containing 10% FBS to complete the culture and stop digestion, pipette the cells with 1ml pipette, transfer to a 15ml centrifuge tube, centrifuge at 800rpm for 5min, and then add 3ml of complete culture to resuspend the cells depending on the cell volume.

[0037] (2) Cell counting After mixing by pipetting, add 20 μL of the suspension to a counting chamber and count three times using a computer counter to obtain the average cell concentration.

[0038] (3) Cell plating The cell concentration was calculated to be 5 × 10⁴ cells / ml. 100 µL of cell culture was added to each well of a 96-well plate and the plate was incubated for 24 h.

[0039] 2. Drug treatment Table 1. Reagents and dosages required for drug treatment

[0040] Dilute the compounds using DMEM according to the table above.

[0041] Remove the 96-well plate, aspirate the original culture medium from the wells, and add 100 µL of the above-mentioned culture medium containing different concentrations of the compound to each well, with 3 replicates for each concentration. Return to the incubator and incubate for 24 hours.

[0042] 3. Add CCK8 processing Remove the 96-well plate, add 10 µL of CCK8 to each well, and incubate at 37°C for 2 hours.

[0043] 4. Read the OD value using an ELISA reader. Remove the 96-well plate, place it in a microplate reader, and read the OD value at a wavelength of 450 nm.

[0044] The inhibition rate is calculated based on the OD value.

[0045] This experiment investigated the in vitro activity of compounds LWJ-3-1 and LWJ-3-2, using inhibition rate as an indicator to evaluate their inhibitory effects on target enzymes or cells. The results showed (e.g.) Figure 5 As shown in the figure, both compounds exhibited dose-dependent inhibitory properties, with IC50 values ​​of 39.42 μM and 41.05 μM, respectively, indicating that they can effectively exert inhibitory effects within the micromolar concentration range. These results suggest that the LWJ-3 series compounds synthesized by the red light-driven, spinach chlorophyll-mediated coupling of coumarin and tetrahydroisoquinoline skeletons of this invention have the potential to be further developed as lead compounds into drugs for treating related diseases, especially showing promising applications in regulating pathological processes related to this target. Further optimization of the molecular structure can improve its selectivity and affinity, and in vivo experiments can be used to verify its efficacy and safety, providing a scientific basis for the development of novel therapeutic drugs.

[0046] Therefore, the present invention employs the above-mentioned novel coumarin and tetrahydroisoquinoline coupling method and application driven by red light and mediated by spinach chlorophyll, which is simple to operate, environmentally friendly, and highly efficient in coupling.

[0047] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and not to limit them. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can still be made to the technical solutions of the present invention, and these modifications or equivalent substitutions cannot cause the modified technical solutions to deviate from the spirit and scope of the technical solutions of the present invention.

Claims

1. A novel red light-driven, spinach chlorophyll-mediated coupling method for coumarin and tetrahydroisoquinoline, characterized in that, Tetrahydroisoquinoline compound, coumarin compound, chlorophyll photosensitizer and solvent were mixed in a certain proportion and subjected to light reaction in an air atmosphere, so that the skeleton of tetrahydroisoquinoline compound and coumarin compound were coupled together, and the compound was purified to obtain a compound that has both coumarin skeleton and tetrahydroisoquinoline skeleton.

2. The novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll according to claim 1, characterized in that, The molar ratio of the tetrahydroisoquinoline compound to the coumarin compound is 1:1 to 3.

3. The novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll according to claim 1, characterized in that, The molar amount of the chlorophyll photosensitizer is 2% to 5% of the molar amount of the tetrahydroisoquinoline compound.

4. The novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll according to claim 1, characterized in that, The ratio of solvent to tetrahydroisoquinoline compound is 0.1 mmol: 1-2 mL.

5. The novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll according to claim 1, characterized in that, The solvent is MeOH.

6. The novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll according to claim 1, characterized in that, The chlorophyll photosensitizer is spinach extract.

7. The novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll according to claim 1, characterized in that, The illumination uses 660-680nm red light.

8. The novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll according to claim 1, characterized in that, The reaction temperature of the photo-induced reaction is 25–30°C, and the reaction time is 12–24 h.

9. The novel coumarin and tetrahydroisoquinoline coupling method driven by red light and mediated by spinach chlorophyll according to claim 1, characterized in that, The general structural formula of the tetrahydroisoquinoline compounds is: ； in, R1 is a C1-C16 alkyl, aromatic heterocyclic or... Any of the substituted aromatic rings; In this context, M1, M2, M3, M4, or M5 are each independently any one of H, F, Cl, Br, ester group, cyano group, sulfone group, nitro group, C1-C16 alkyl group, or C1-C16 alkyl group; The general structural formula of the coumarin compounds is: ； Among them, R2, R3, and R4 are each independently any one of H, O, and hydroxyl; R5, R6, R7, and R8 are each independently any one of H, F, Cl, Br, ester group, cyano group, sulfone group, nitro group, C1-C16 alkyl group, and C1-C16 alkyl group.

10. The application of compounds with coumarin and tetrahydroisoquinoline skeletons prepared by the novel coumarin and tetrahydroisoquinoline coupling method mediated by red light-driven spinach chlorophyll as described in any one of claims 1-9 in the treatment of colon cancer.

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