Use of a piperazine compound in the prevention and treatment of ralstonia solanacearum

Abstract

The application discloses application of a piperazine compound to prevention and treatment of Ralstonia solanacearum and belongs to the technical field of plant bacterial wilt prevention and treatment. The piperazine compound is 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinoline carboxylic acid. Research finds that the 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinoline carboxylic acid has strong inhibitory activity on Ralstonia solanacearum. According to the antibacterial experiment, the piperazine compound has an LC 50 of 1.079 μg / mL on Ralstonia solanacearum at 24 h, an inhibition rate of more than 98% at an effective concentration of 5 μg / mL at 24 h, and an inhibition rate of 100% at ≥7.5 μg / mL, and is a candidate compound for preventing and treating Ralstonia solanacearum at a low concentration to achieve 100% inhibition.

Description

Technical Field

[0001] This invention belongs to the field of plant bacterial wilt control technology, specifically relating to the application of a piperazine compound in the control of Ralstonia solanacearum. Background Technology

[0002] Ralstonia solanacearum ( Ralstonia solanacearum , Rso Bacterial wilt, also known as bacterial wilt fungus, is a widely distributed soil-borne pathogen that can cause bacterial wilt in more than 200 plant species from 50 families. It seriously damages important economic crops such as tomatoes, tobacco, potatoes, and bananas, and can even lead to crop failure. The direct economic losses caused by bacterial wilt worldwide exceed US$1 billion annually.

[0003] Currently, the main methods for controlling bacterial wilt of plants include agricultural control, biological control, and chemical control. Agricultural control measures such as crop rotation and soil improvement have limited effectiveness, while biological control is greatly affected by environmental factors and its effects are unstable. Although traditional fungicides can control the disease, their effectiveness against the soil-borne pathogen Ralstonia solanacearum is limited. Piperazine compounds are widely used in the pharmaceutical field due to their broad-spectrum antibacterial activity; however, their application against plant pathogens, especially Ralstonia solanacearum, has not been reported. Therefore, developing novel, highly effective, low-toxicity, and environmentally friendly piperazine compounds against Ralstonia solanacearum is of great significance for sustainable agricultural development. Summary of the Invention

[0004] The first objective of this invention is to provide an application of piperazine compounds in the prevention and control of Ralstonia solanacearum, and the second objective is to provide a bactericide for the prevention and control of Ralstonia solanacearum.

[0005] The first objective of this invention is achieved through the application of the piperazine compound in the control of Ralstonia solanacearum, wherein the piperazine compound is 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid, with the structural formula shown in Formula I:

[0006]

[0007] I;

[0008] The piperazine compound exhibited a LC50 spectral density against Ralstonia solanacearum at 24 h. 50 It was 1.079 μg / mL.

[0009] The second objective of this invention is achieved by the fact that the fungicide for controlling Ralstonia solanacearum uses 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid as the main active ingredient or one of the active ingredients.

[0010] This invention is the first to discover that the piperazine compound 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid exhibits strong inhibitory activity against Ralstonia solanacearum. Based on antibacterial experiments, the LC50 of the 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid described in this invention against Ralstonia solanacearum was [not specified in the original text]. 50 With a concentration of 1.079 μg / mL, the effective concentration of 5 μg / mL showed an inhibition rate of over 98% after 24 hours, and the inhibition rate reached 100% at concentrations of ≥7.5 μg / mL. This compound is a candidate compound for the control of Ralstonia solanacearum that can achieve 100% inhibition at low concentrations, providing new ideas and approaches for the development of novel plant pathogenic bacterial pesticides and showing broad application prospects. Attached Figure Description

[0011] Figure 1 The structural formula is 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid;

[0012] Figure 2 The results are calculated to determine the inhibitory effect of 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinoline carboxylic acid on Ralstonia solanacearum in the experimental example. Detailed Implementation

[0013] The present invention will be further described in detail below with reference to experimental examples and embodiments, but this does not limit the present invention in any way. Any modifications or improvements made based on the teachings of the present invention shall fall within the protection scope of the present invention.

[0014] The application of the piperazine compound of the present invention in the control of Ralstonia solanacearum, wherein the piperazine compound is 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid, and its structural formula is shown in Formula I:

[0015]

[0016] I;

[0017] The piperazine compound exhibited a LC50 spectral density against Ralstonia solanacearum at 24 h. 50 It was 1.079 μg / mL.

[0018] The fungicide for controlling Ralstonia solanacearum described in this invention uses 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid as the main active ingredient or one of the active ingredients.

[0019] Furthermore, the effective concentration of 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid in the bactericide is ≥5 μg / mL.

[0020] Furthermore, the effective concentration of 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid in the bactericide is ≥7.5 μg / mL.

[0021] Furthermore, the bactericide is available in the form of an aqueous solution, an emulsion, a wettable powder, a soluble powder, a soluble granule, a suspension, a tablet, or an effervescent granule.

[0022] The piperazine compound 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid (hereinafter referred to as piperazine compound A) described in this invention is derived from a drug-like diversity compound library containing 50,000 compounds developed by Shanghai Taoshu Biotechnology Co., Ltd., with Library ID F6660-2932, Catalog Number HY-Q05753, CASNumber 2379995-76-1, and a relative molecular mass of 386.377.

[0023] This invention uses piperazine compound A as the research object and conducts the following experiments:

[0024] Experimental example to detect the antibacterial activity of piperazine compound A against Ralstonia solanacearum

[0025] 1. Preparation of experimental reagents

[0026] 1-1. Weigh piperazine compound A and dissolve it in 100% DMSO. Shake or sonicate until completely dissolved to obtain the base solution. Dilute the base solution with pure DMSO to obtain experimental test solutions with piperazine compound A concentrations of 10 μg / mL, 25 μg / mL, 50 μg / mL, 75 μg / mL, 100 μg / mL, 250 μg / mL, 500 μg / mL, 750 μg / mL, and 1000 μg / mL.

[0027] 1-2. Use 100% DMSO as the negative control solution;

[0028] 1-3. Prepare NB liquid culture medium by dissolving 5g of peptone, 3g of beef extract, and 5g of sodium chloride in purified water and bringing the volume to 1L. Then sterilize at high temperature to obtain NB liquid culture medium. Use NB liquid culture medium as blank control solution.

[0029] 1-4. Based on the ratio of NB liquid culture medium prepared in step 1-3, add 15g of agar, sterilize at high temperature to obtain NA solid culture plate, and set aside.

[0030] 2. Preparation of Ralstonia solanacearum for the experiment

[0031] Ralstonia solanacearum was streaked onto the NA solid culture plate obtained in steps 1-4 and placed in an incubator at 28°C for overnight static culture. Single colonies from the NA solid culture plate were picked and inoculated into the NB liquid culture medium obtained in steps 1-3. The culture was then shaken in an incubator at 28°C for 48 hours. Finally, the culture was diluted 100 times by volume with sterilized NB liquid culture medium to obtain the Ralstonia solanacearum bacterial suspension for the experiment, which was then set aside.

[0032] 3. Experimental Methods

[0033] The experiment was conducted in a sterile 96-well plate using the microdilution method.

[0034] Experimental group: 1 μL of the test drug solution of each piperazine compound A concentration from step 1-1 was added sequentially to each well of a 96-well plate. Then, 99 μL of the experimental Ralstonia solanacearum bacterial suspension from step 2 was added to each well. After mixing, the plate was incubated at 28°C. The growth of Ralstonia solanacearum in each well was observed and the OD was measured after 24 hours. 600 At this point, the final concentrations of piperazine compound A in each well were 0.1 μg / mL, 0.25 μg / mL, 0.50 μg / mL, 0.75 μg / mL, 1.0 μg / mL, 2.5 μg / mL, 5.0 μg / mL, 7.5 μg / mL, and 10.0 μg / mL, respectively.

[0035] Negative control group: Add 1 μL of the negative control group solution from steps 1-2 to a 96-well plate, then add 99 μL of the *Ralstonia solanacearum* bacterial suspension from step 2. Mix well and incubate at 28°C. Observe the growth of *Ralstonia solanacearum* and measure the OD value after 24 hours. 600 ;

[0036] Blank control group: Add 99 μL of the blank control solution from steps 1-3 to a 96-well plate, then add 1 μL of the negative control solution from steps 1-2, mix well, and incubate at 28°C. Measure the OD at 24 h. 600 ;

[0037] Each group had 3 replicates, OD 600 The value is taken as the average of three replicates; OD is measured. 600To quantify the inhibitory effect of different concentrations of the piperazine compound A on Ralstonia solanacearum, and to analyze the relationship between inhibitory activity and concentration, the inhibition rate was calculated using the following formula: Inhibition rate % = [(average OD value of negative control group - average OD value of experimental group) / (average OD value of negative control group - average OD value of blank control group)] × 100.

[0038] 4. Experimental Results:

[0039] The antibacterial activity of piperazine compound A against Ralstonia solanacearum is shown in Table 1 and... Figure 2 As shown; the mean OD of the negative control group was measured. 600 The mean OD value was 0.430, and the mean OD value of the blank control group was 0.430. 600 The value was 0.267; combined with the average OD of the experimental group in Table 1... 600 The inhibition rate (%) corresponding to the concentration of the test drug solution in each experimental group was obtained from the inhibition rate calculation formula in Table 1.

[0040] Table 1. Inhibitory effect of piperazine compound A on Ralstonia solanacearum

[0041]

[0042] Results analysis: Combining Figure 2 The LC50 of piperazine compound A against Ralstonia solanacearum was determined at 24 h. 50 The effective concentration was 1.079 μg / mL; Table 1 clearly shows that the inhibition rate of 5 μg / mL reached over 98% after 24 h, and the inhibition rate reached 100% at ≥7.5 μg / mL; indicating that 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid has good antibacterial activity against Ralstonia solanacearum and can be used for the prevention and control of Ralstonia solanacearum.

[0043] The following are examples of the preparation of fungicides for controlling Ralstonia solanacearum.

[0044] Example 1

[0045] The effective concentration of 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid in the aqueous bactericide is 6 μg / mL.

[0046] Example 2

[0047] The effective concentration of 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid in the bactericide emulsion is 7.5 μg / mL.

[0048] Example 3

[0049] The effective concentration of 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid in the wettable powder of the bactericide is 6.5 μg / mL.

[0050] Example 4

[0051] The effective concentration of 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid in the bactericide suspension is 5 μg / mL.

[0052] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments.

Claims

1. The application of a piperazine compound in the control of Ralstonia solanacearum, characterized in that, The piperazine compound is 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid, and its structural formula is shown in Formula I: I； The piperazine compound exhibited a LC50 spectral density against Ralstonia solanacearum at 24 h. 50 It was 1.079 μg / mL.

2. A fungicide for controlling Ralstonia solanacearum, characterized in that, The bactericide uses 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid as described in claim 1 as its main active ingredient.

3. The bactericide according to claim 2, characterized in that, The effective concentration of 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid in the bactericide is ≥5 μg / mL.

4. The bactericide according to claim 3, characterized in that, The effective concentration of 7-[4-(2-cyanoacetyl)-1-piperazinyl]-1-ethyl-6-fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid in the bactericide is ≥7.5 μg / mL.

5. The bactericide according to claim 2 or 3, characterized in that, The bactericide is available in the form of an aqueous solution, an emulsion, a wettable powder, a soluble powder, a soluble granule, a suspension, a tablet, or an effervescent granule.

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