Thymol derivatives and uses thereof

Abstract

The application discloses a thymol derivative as shown in formula I, wherein R is preferably 3-mercapto-1,2,4-triazole (1a), 3,4-dimethyl-1,2,4-triazole (1b), 1,2,3-triazole (1c), 3,5-dibromo-1,2,4-triazole (1d), 6-methyl-1H-benzo-1,2,3-triazole (1e), 3-methoxycarbonyl-1,2,4-triazole (1f), 6-chloro-1H-benzo-1,2,3-triazole (1g); the application also provides a pharmaceutical salt of the thymol derivative; the application also provides a preparation method of the thymol derivative, and provides the use of the thymol derivative and the pharmaceutical salt thereof as an antibacterial agent against fusarium oxysporum for preparing a pesticide.

Description

Technical Field

[0001] This invention belongs to the field of novel natural product-modified drug development technology, and particularly relates to a thymol derivative and its preparation method and application. Background Technology

[0002] Thymol, also known as thymol, is a natural product derived from the volatile oil of plants in the genus thyme. Its structure is as follows: As shown, it can be obtained by introducing isopropyl and methyl groups at the 2 and 5 positions of phenol, respectively. As a natural product, it has a wide range of uses. In the pharmaceutical field, it can be used as a topical medication to treat prickly heat, herpes, and pruritus; in agriculture, it can be used as an insecticide and antifungal agent to control plant diseases and pests; in animal husbandry, it can also be used as a feed additive to reduce animal diarrhea and enhance reproductive rates and lactation performance.

[0003] Fusarium oxysporum is a fungus belonging to the genus Fusarium in the class Deuteromycetes. It is a major pathogen causing wilt disease in plants, and can cause wilt in more than 150 kinds of plants, including cotton, tomatoes, bananas, watermelons, strawberries, cabbage, chickpeas, peas, alfalfa, lilies, etc.

[0004] Currently, the control of plant diseases caused by *Fusarium oxysporum* mainly falls into two categories: biological control and chemical control. Biological control generally utilizes microorganisms (bacteria, fungi, or actinomycetes) that produce antibiotics to inhibit the growth of *Fusarium oxysporum*. Although biological control has made some progress, it has largely not been scaled up, mainly due to unstable control effects in the field, lack of broad-spectrum efficacy, strict requirements for product production and storage, slower and less effective compared to chemical agents, and higher prices. Under current circumstances, chemical control remains the primary strategy for controlling *Fusarium oxysporum*. Chemical control primarily uses pesticides such as azoles, methyl esters, and amides. However, overall, the control effects of currently used chemical pesticides are not ideal, they easily lead to resistance, long-term use can cause soil acidification and heavy metal contamination, and they are relatively expensive. Therefore, it is essential to develop new, highly effective, low-toxicity, and inexpensive pesticides against *Fusarium oxysporum*.

[0005] This invention investigated the antifungal activity of thymol against Fusarium oxysporum and found that it has significant activity against Fusarium oxysporum, with an EC50 value of [missing information]. 50 The half-maximal inhibitory concentration (MCIC) is 40.1 μg / mL. However, this is still somewhat inferior to existing reported antibacterial agents. Therefore, this invention synthesizes a thymol derivative for use as an antifungal agent against Fusarium oxysporum in the preparation of pesticides, which are then used to control lily wilt disease. Summary of the Invention

[0006] One objective of this invention is to provide a series of thymol derivatives and their synthesis methods; another objective is to disclose the use of these derivatives as antifungal agents against Fusarium oxysporum in the preparation of pesticides. In some embodiments, the thymol derivative is a thymol ethyltriazole derivative.

[0007] On the one hand, the present invention provides thymol derivatives as shown in Formula I,

[0008]

[0009] Where R is

[0010] R1 and R2 are independently H, halogen, and C, respectively. m-n Alkyl, C x-y cycloalkyl or -COOR7;

[0011] R3 and R4 are each independently H, halogen, and C. m-n Alkyl, C x-y Cycloalkyl or -COOR7; or R3 and R4 together with the carbon atoms they are bonded to form a cycloalkyl group, said cyclic group being a five-membered or six-membered ring, said cyclic group being unsubstituted or optionally C m-n Alkyl and / or halogen substitution;

[0012] R8 and R9 are independently H, halogen, and C. m-n Alkyl, C x-y cycloalkyl or -COOR7;

[0013] R5 and R6 are independently H, halogen, and C. m-n Alkyl, C x-y Cycloalkyl groups or -COOR7; or R5 and R6 together with the nitrogen and carbon atoms they are bonded to form heterocyclic groups, said heterocyclic groups being five-membered or six-membered rings, said cyclic groups being unsubstituted or optionally C-shaped. m-n Alkyl and / or halogen-substituted; the heterocyclic group is a group with only one heteroatom;

[0014] R7 is C m-n Alkyl or C x-y cycloalkyl;

[0015] In some implementations, the C m-n Alkyl is C 1-10 Alkyl, the C x-y Cycloalkyl is C 3-8 cycloalkyl;

[0016] In some implementations, the C m-n Alkyl and C x-yThe cycloalkyl group is either unsubstituted or optionally substituted with a halogen.

[0017] In some implementations, the C m-n Alkyl is C 1-6 Alkyl, the C x-y Cycloalkyl is C 5-8 Cycloalkyl.

[0018] In some implementations, the C m-n The alkyl group is methyl, ethyl, or propyl; the C x-y Cycloalkyl is C 5-6 Cycloalkyl.

[0019] In some embodiments, R1 and R2 are not both -COOR7; in other embodiments, R1 and R2 are not a combination of cycloalkyl and -COOR7.

[0020] In some embodiments, R3 and R4 are not both -COOR7; in other embodiments, R3 and R4 are not a combination of cycloalkyl and -COOR7.

[0021] In some embodiments, R5 and R6 are not both -COOR7; in other embodiments, R5 and R6 are not a combination of cycloalkyl and -COOR7.

[0022] In some embodiments, R8 and R9 are not both -COOR7; in other embodiments, R8 and R9 are not a combination of cycloalkyl and -COOR7.

[0023] "alkyl" refers to a saturated aliphatic group that has a straight or branched chain of carbon atoms.

[0024] "Halogen" refers to fluorine, chlorine, bromine, or iodine. "Halogenated" refers to fluorinated, chlorinated, bromine, or iodinated substances.

[0025] Unless otherwise stated, “optionally substituted” means that a given group may consist only of hydrogen substituents with available valences.

[0026] "Cycloalkyl" refers to a non-aromatic saturated cyclic group.

[0027] "Heteroatoms" are atoms that are not carbon or hydrogen atoms.

[0028] In some embodiments, R8 and R9 are not a combination of cycloalkyl and -COOR7, nor are they simultaneously -COOR7; and / or, R3 and R4 are not a combination of cycloalkyl and -COOR7, nor are they simultaneously -COOR7; and / or, R5 and R6 are not a combination of cycloalkyl and -COOR7, nor are they simultaneously -COOR7.

[0029] In some specific embodiments, the thymol derivative is compound 1a-1g, wherein R is 3-mercapto-1,2,4-triazole (1a), 3,4-dimethyl-1,2,4-triazole (1b), 1,2,3-triazole (1c), 3,5-dibromo-1,2,4-triazole (1d), 6-methyl-1H-benzo-1,2,3-triazole (1e), 3-methoxyformyl-1,2,4-triazole (1f), or 6-chloro-1H-benzo-1,2,3-triazole (1g), and the structure of R is shown in formula 1a-1g:

[0030]

[0031] On the one hand, the present invention also provides pharmaceutically acceptable salts of thymol derivatives as described above. Pharmaceutically acceptable salts refer to salts of the compounds of the present invention that are pharmaceutically acceptable and possess the desired pharmacological activity. Such salts include acid addition salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc.; or acid addition salts formed with organic acids such as acetic acid, propionic acid, hexanoic acid, heptanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, o-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, etc. Ethylenesulfonic acid, 1,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, p-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, p-toluenesulfonic acid, camphorsulfonic acid, 4-methylbicyclo[2.2.2]oct-2-ene-1-carboxylic acid, glucoheponic acid, 4,4'-methylenebis(3-hydroxy-2-ene-1-carboxylic acid), 3-phenylpropionic acid, trimethylacetic acid, tert-butylacetic acid, trifluoroacetic acid, lauryl sulfate, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, mucoconic acid, etc.

[0032] Pharmaceutically acceptable salts also include base addition salts, which can be formed when the acidic protons present can react with inorganic or organic bases. Acceptable inorganic bases include sodium hydroxide, sodium carbonate, potassium hydroxide, aluminum hydroxide, and calcium hydroxide. Acceptable organic bases include ethanolamine, diethanolamine, triethanolamine, aminobutanetriol, α-methylglucosamine, etc.

[0033] On the one hand, the present invention provides a method for obtaining thymol derivatives as described above, the method comprising the steps (such as...) Figure 1 )include:

[0034] (1) Thymol (compound 2) undergoes a substitution reaction with dibromoethane in the presence of a base to give thymol oxyethyl bromide (compound 3);

[0035] (2) Compound 3 was reacted with triazole or triazole derivative (compound 4) under reflux in the presence of carbonate to prepare thymol derivative (compound 1) as shown in Formula 1.

[0036] In some specific embodiments, the triazole or its derivative is

[0037] R1, R2, R3, R4, R5, R6, R8, and R9 are as described above.

[0038] Figure 1 In this document, number 1 refers to compound 1, namely the thymol derivative described herein; number 2 refers to compound 2, namely thymol; number 3 refers to compound 3, namely thymol oxyethyl bromide; and number 4 refers to compound 4, namely triazole or its derivative.

[0039] In some specific embodiments, the triazole or its derivative is compound 4a-4g, and the resulting thymol derivative is a compound as shown in Formula 1, wherein R is as shown in 1a-1g.

[0040]

[0041] In some specific embodiments, the alkali mentioned in step (1) is sodium hydroxide or potassium hydroxide; preferably, the alkali is sodium hydroxide.

[0042] In some specific embodiments, the carbonate in step (2) is sodium carbonate or potassium carbonate; preferably, the carbonate is potassium carbonate.

[0043] On the one hand, the present invention also provides the application of the above-mentioned thymol derivatives in inhibiting Fusarium oxysporum.

[0044] Specifically, the application is in the preparation of antimicrobial agents against Fusarium oxysporum;

[0045] Specifically, the application can be to use the above-mentioned thymol derivative as an active ingredient to prepare a pesticide, which is used for the prevention and control of lily wilt disease.

[0046] On one hand, the present invention provides a method for inhibiting Fusarium oxysporum, wherein the method involves contacting the Fusarium oxysporum with the aforementioned thymol derivative.

[0047] The thymol derivatives provided by this invention have the following advantages: In in vitro bioactivity tests, the thymol derivatives provided by this invention have anti-Fusarium oxysporum activity, wherein the inhibition rate of compound 1a is significantly higher than that of thymol. Attached Figure Description

[0048] Figure 1This is the preparation process of thymol derivatives;

[0049] Figure 2 Compound 1 exhibits anti-Fusarium oxysporum activity. Detailed Implementation

[0050] The present invention is described through the following specific embodiments, through which the present invention can be better understood, but the scope of the present invention is not limited to these embodiments.

[0051] Example 1: Preparation of thymol oxyethyl bromide (compound 3)

[0052] Specific synthesis method: Thymol (10 g, 67 mmol) was added to a 500 mL three-necked flask, followed by slow addition of 150 mL of pure water. The mixture was stirred, and then sodium hydroxide (NaOH, 4 g, 100 mmol), tetrabutylammonium bromide (TBAB, 2.2 g, 6.7 mmol), and potassium iodide (KI, 1.1 g, 6.7 mmol) were added sequentially. The mixture was heated until completely dissolved. When the temperature reached 90 °C, 1,2-dibromoethane (12 mL, 134 mmol) was slowly added to the reaction solution, maintaining the temperature at approximately 100 °C. Stirring was continued for 12 h. The reaction was monitored by thin-layer chromatography (TLC). After the reaction was complete, the mixture was filtered, and the oily liquid was collected. The solution was then washed three times with 2% NaOH aqueous solution and saturated brine, respectively. Finally, it was extracted with ethyl acetate. The collected organic layer solution was dried over anhydrous sodium sulfate. The filtrate was concentrated and separated by column chromatography using petroleum ether / ethyl acetate (V / V = 40 / 1) as the eluent to obtain compound 3.

[0053] Compound 3, a colorless oily liquid. 1 HNMR (600M, CDCl3) δppm: 7.12 (d, J = 7.7Hz, 1H), 6.79 (d, J = 7.7Hz, 1H), 4.30 (t, J = 6 .1Hz,2H),3.67(t,J=6.1Hz,2H),3.33(m,1H),2.33(s,3H),1.23(d,J=6.7Hz,6H).

[0054] Example 2: General method for preparing thymol derivatives (compound 1)

[0055] Thymoloxyethyl bromide (compound 3, 500 mg, 2.07 mmol) was added to a 50 mL reaction flask, followed by 25 mL of acetonitrile. The mixture was stirred, then potassium carbonate (572 mg, 4.14 mmol) was added, and finally 3-mercapto-1,2,4-triazole (316 mg, 3.12 mmol) was added. The mixture was heated to reflux at 80 °C for 12 h. After the reaction was monitored by thin-layer chromatography (TLC), the precipitate was filtered, washed three times with 50 mL of acetonitrile, and the filtrate was concentrated under reduced pressure. The precipitate was then purified by column chromatography using petroleum ether / ethyl acetate (V / V = 1 / 1) as the eluent to obtain the target product 1a. The synthesis methods for compounds 1b-1g are the same.

[0056] The physicochemical data of compound 1a-1g are as follows:

[0057] Compound 1a, a white powdery solid. 1 HNMR (600M, CDCl3) δppm: 8.24 (s, 1H), 7.13 (d, J = 7.6Hz, 1H), 6.80 (d, J = 7.6Hz, 1H), 6.71 (s, 1H), 4.30 (t ,J=6.4Hz,2H),3.60(t,J=6.4Hz,2H),3.29(m,1H),2.33(s,3H),2.22(s,1H,-NH),1.23(d,J=7.1Hz,6H); 13 CNMR (150M, CDCl3) δppm: 155.5, 155.3, 136.3, 136.2, 134.1, 126.0, 121.7, 112.5, 66.8, 31.8, 26.5, 22.7, 22.7, 21.2.

[0058] Compound 1b is a white powdery solid. 1 HNMR (600M, CDCl3) δppm: 7.06 (d, J = 7.7Hz, 1H), 6.75 (d, J = 7.7Hz, 1H), 6.58 (s, 1H) 4.38 (t, J = 6.4Hz, 2 H),4.31(t,J=6.4Hz,2H),3.06(m,1H),2.47(s,3H),2.31(s,3H),2.28(s,3H),1.10(d,J=6.9Hz,6H); 13 CNMR (150M, CDCl3) δppm: 159.7, 155.1, 153.1, 136.4, 133.9, 125.9, 122.0, 112.4, 66.3, 47.7, 26.3, 22.7, 22.7, 21.2, 13.7, 11.7.

[0059] Compound 1c is a colorless, oily liquid.1 HNMR (600M, CDCl3) δppm: 7.65 (s, 2H), 7.09 (d, J = 7.7Hz, 1H), 6.78 (d, J = 7.7Hz, 1H), 6.67 (s, 1H) ,4.88(t,J=5.6Hz,2H),4.50(t,J=5.6Hz,2H),3.16(m,1H),2.33(s,3H),1.11(d,J=7.0Hz,6H).; 13 CNMR (150M, CDCl3) δppm: 155.3, 136.3, 134.4, 134.4, 126.1, 121.9, 112.5, 66.0, 54.2, 26.5, 22.7, 22.7, 21.3.

[0060] Compound 1d, colorless needle crystals (methanol), 1 HNMR (600M, CDCl3) δppm: 7.08 (d, J = 7.7Hz, 1H), 6.78 (d, J = 7.7Hz, 1H), 6.60 (s, 1H), 4.57 ( t,J=5.1Hz,2H),4.36(t,J=5.1Hz,2H),3.29(m,1H),2.31(s,3H),1.10(d,J=6.9Hz,6H).; 13 CNMR (150M, CDCl3) δppm: 154.6, 140.4, 136.2, 134.0, 130.6, 125.9, 122.0, 112.0, 65.0, 49.4, 26.2, 22.5, 22.5, 21.1.

[0061] Compound 1e, a white, lumpy solid. 1 HNMR(600M, CDCl3)δppm:7.74(dd,J=8.7,0.8Hz,1H),7.60(s,1H),7.21(d,J=8.8,1H),7.03(d,J=7.7Hz,1H),6.74(d,J=7.8Hz, 1H),6.66(ds,1H),5.10(t,J=5.6Hz,2H),4.62(t,J=5.6Hz,2H),3.07(m,1H),2.49(s,3H),2.29(s,3H),0.99(d,J=6.9Hz,6H).; 13CNMR (150M, CDCl3) δppm:155.5,145.5,143.6,136.9,136.7,134.9,129.7,1 26.5,122.4,117.8,116.7,113.0,66.7,56.1,26.8,23.0,23.0,22.5,21.7..

[0062] Compound 1f, a white, lumpy solid. 1 HNMR (600M, CDCl3) δppm: 8.00 (s, 1H), 7.04 (d, J = 8.0 Hz, 1H), 6.74 (d, J = 8.0, 1H), 6.59 (m, 1H), 5.11 (t ,J=5.3Hz,2H),4.37(t,J=5.3Hz,2H),4.01(s,3H),3.00(m,1H),2.29(s,3H),1.06(d,J=6.9Hz,6H).; 13 CNMR (150M, CDCl3) δppm: 158.8, 155.4, 151.4, 145.1, 136.8, 134.5, 126.4, 122.3, 112.5, 66.4, 53.5, 50.7, 26.8, 23.1, 23.1, 21.7.

[0063] 1g of the compound, a colorless oily liquid. 1 HNMR (600M, CDCl3) δppm: 7.87 (s, 1H), 7.81 (d, J = 9.1Hz, 1H), 7.34 (d, J = 9.0, 1H), 7.04 (d, J = 7.7Hz, 1H), 6.75 (d, J = 7.7H z,1H),6.66(s,1H),5.12(t,J=5.5Hz,2H),4.63(t,J=5.5Hz,2H),3.04(m,1H),2.30(s,3H),0.98(dt,J=7.0,1.4Hz,6H); 13 CNMR (150M, CDCl3) δppm: 154.8, 144.6, 142.8, 136.1, 134.2, 132.0, 127. 8,125.9,121.8,119.0,116.9,112.3,65.8,55.9,26.2,22.3,22.3,21.1.

[0064] Example 3: Assay of the anti-Fusarium oxysporum activity of compound 1

[0065] Anti-Fusarium oxysporum activity was determined using the toxic culture medium method.

[0066] (1) Test strains

[0067] Fusarium oxysporum was purchased from Beijing Beina Chuanglian Biotechnology Research Institute.

[0068] (2) Preparation of the sample solution to be tested:

[0069] After completely dissolving 5 mg of sample in 500 μL of DMSO, 9500 μL of sterile water was added to obtain a 500 μg / mL sample solution.

[0070] (3) Measurement method

[0071] Add 10 mL of the diluted sample solution to 90 mL of PDA (60-70℃) medium. After the medium cools and solidifies, use a 5 mm diameter sterile punch to collect *Fusarium oxysporum* mycelial pellets (5-6 days growth period). Inoculate the 5 mm pellets into the center of the PDA medium containing the sample solution; this is the sample treatment group. Use PDA medium without the sample solution as the blank control group. After inoculation, incubate at 30℃ for 4-5 days, measure the colony diameter, and calculate the inhibition rate.

[0072]

[0073] d c d0: Colony diameter in the blank control group; d0: Diameter of the mycelial cake; d s : Colony diameter of sample treatment group

[0074] (4) Results of the anti-Fusarium oxysporum activity of compound 1

[0075] Results of the antifungal activity of compound 1 and thymol against Fusarium oxysporum are shown in [the table below]. Figure 2 And Table 1. It can be seen that...

[0076] Table 1. Antifungal activity of compound 1 against Fusarium oxysporum

[0077]

[0078]

Claims

1. A thymol derivative, characterized in that, The structural formula of the thymol derivative is: 。 2. The application of thymol derivatives, characterized in that, The use of the thymol derivative of claim 1 in the preparation of an antibacterial agent against Fusarium oxysporum.

3. The application of thymol derivatives, characterized in that, The use of the thymol derivative of claim 1 in the preparation of pesticides, wherein the pesticide is used for the control of lily wilt disease, and the thymol derivative is the active ingredient.

Images