Preparation method and application of bis(4-pyrazolone) methane compounds
A simplified synthetic method was used to prepare bis(4-pyrazolinone)methane compounds, which solved the problem of insufficient research on succinate dehydrogenase promoters and achieved low-cost and efficient preparation and promotion of edible fungi growth.
Patent Information
- Application Number
- CN202310361488.X
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-04-06
- Publication Date
- 2026-01-30
- Estimated Expiration
- 2043-04-06
AI Technical Summary
There is insufficient research on succinate dehydrogenase promoters. Traditional methods for synthesizing bis(4-pyrazolone)methane compounds require many raw materials and have harsh reaction conditions, which are difficult to meet the needs of agricultural production.
Bis(4-pyrazolone)methane compounds were prepared by heating and refluxing compounds A and B in an organic mixed solvent, filtering, and then allowing the solvent to evaporate naturally. Compound A is a 4-acylpyrazolone, and B is benzoyl hydrazine or its derivative. The solvent is a mixture of dimethyl sulfoxide and 95% methanol or ethanol in a ratio of 5:1 or 1:5.
A low-cost, high-yield preparation of bis(4-pyrazolinone)methane compounds was achieved, which can promote the mycelial growth of edible fungi and activate succinate dehydrogenase activity, and can be used as a potential growth regulator for edible fungi.
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Abstract
Description
Technical Field
[0001] This invention belongs to the field of compound synthesis, specifically relating to a method for preparing bis(4-pyrazolinone)methane compounds and their application as succinate dehydrogenase promoters. Background Technology
[0002] Succinate dehydrogenase is a crucial link between oxidative phosphorylation and electron transport, providing electrons to the respiratory chain of eukaryotic mitochondria and various prokaryotic cells for both aerobic and energy-producing processes. Its activity is generally used as an indicator of the progress of the tricarboxylic acid cycle. The succinate dehydrogenase complex is the smallest complex in the mitochondrial respiratory chain, capable of transferring electrons from succinate to quinone conjugates. Targeted molecular design against succinate dehydrogenase can achieve specific regulation of respiration, further leading to the development of growth promoters for crops or inhibitors for pathogens, ultimately promoting increased agricultural yields and income.
[0003] Research on succinate dehydrogenase inhibitors is currently extensive. Since 2009, succinate dehydrogenase inhibitors have gradually developed into the third largest class of fungicides after methoxyacrylates and triazoles. To date, 18 varieties have entered the market or are under development. Research on these succinate dehydrogenase inhibitors reveals that they generally possess the following four characteristics: first, they have aromatic or heterocyclic structures, especially pyrazole heterocycles; second, they all contain amide structures; third, most contain fluorine atoms; and fourth, they have low molecular symmetry. Conversely, research on succinate dehydrogenase promoters is still in its early stages, and no particularly high-activity lead compounds have yet been discovered. Therefore, it is necessary to develop a succinate dehydrogenase promoter to improve the quality and efficiency of agricultural production.
[0004] Currently, there are not many reports on the synthesis methods of bis(4-pyrazolone)methane compounds. The main methods include: one is the reaction of 5-methyl-6-phenyl-1,2,4-triazine-4-oxide and 1,3,5-pyrazolone in dimethyl sulfoxide (DMSO) in the presence of triethylamine, phosphorus oxychloride, and 2-formamide to synthesize bis(4-pyrazolone)methane. However, this method requires many raw materials and harsh reaction conditions, which is not conducive to production. Therefore, it is necessary to develop a simpler synthetic method. Summary of the Invention
[0005] 1. A method for preparing a bis(4-pyrazolinone)methane compound. The preparation method is as follows: a mixture of compound A and compound B is added to an organic mixed solvent, heated under reflux at a certain temperature for 4-6 hours, then filtered while hot, and the solvent is allowed to evaporate naturally at room temperature to obtain the final target product C.
[0006] 2. The method for preparing a bis(4-pyrazolone)methane compound as described in claim 1, characterized in that: compound A is a 4-acylpyrazolone, and compound B is benzoyl hydrazine or its derivative. The general structural formulas of compounds A, B, and target product C are as follows:
[0007]
[0008] R1, R2, R3, R4, and R5 are any one of alkyl, aryl, heterocyclic, or hydrogen atoms, respectively.
[0009] 3. The method for preparing a bis(4-pyrazolinone)methane compound as described in claim 1, characterized in that: the molar ratio of compound A to compound B is 1:1.
[0010] 4. The method for preparing a bis(4-pyrazolone)methane compound as described in claim 1, characterized in that: the organic mixed solvent consists of two components, one of which is dimethyl sulfoxide (DMSO), and the other is one of 95% methanol and 95% ethanol.
[0011] 5. The method for preparing a bis(4-pyrazolone)methane compound as described in claim 3, characterized in that: the volume ratio of DMSO to 95% methanol in the organic mixed solvent is 5:1.
[0012] 6. The method for preparing a bis(4-pyrazolone)methane compound as described in claim 3, characterized in that: the volume ratio of DMSO to 95% ethanol in the organic mixed solvent is 5:1.
[0013] 7. The application of the final target product according to claim 1 as a mycelial growth promoter. Its characteristic is that a portion of the target compound exhibits significant growth-promoting activity against *Pleurotus ostreatus* mycelia.
[0014] 8. The application of the final target product according to claim 1 as a succinate dehydrogenase promoter. Its characteristic is that a portion of the target compound has a significant promoting effect on the succinate dehydrogenase activity of the stipe of *Pleurotus ostreatus*.
[0015] The method for preparing bis(4-pyrazolone)methane compounds provided by this invention requires fewer raw materials, has simpler synthesis steps, milder reaction conditions, and higher yields compared to traditional methods. These compounds have the ability to increase the growth rate of edible fungi mycelia and activate the activity of mycelial succinate dehydrogenase, and can be used as potential growth regulators for edible fungi. Detailed Implementation
[0016] The present invention will be further described in detail below through specific embodiments. However, this should not be construed as limiting the scope of the present invention to the following embodiments; all technologies implemented based on the above-described content of the present invention fall within the scope of the present invention. In the following embodiments, the melting point is not corrected and the yield is not optimized.
[0017] Implementation Scheme 1: Synthesis of bis-(1-phenyl-3-methyl-pyrazolone-5)methane (ligand L)
[0018] 0.270 g (1.0 mmol) of 1-phenyl-3-methyl-4-trifluoroacetylpyrazolone-5 and 0.143 g (1.0 mmol) of benzoylhydrazine were dissolved separately in a suitable amount of mixed solvent (V(95% Ethanol):V(DMSO)) = 5:1. The two solutions were mixed with stirring, and then stirring was continued for half an hour. The mixture was then refluxed in a water bath at 90 °C for 4 hours. The mixture was filtered while hot, and the solvent was slowly evaporated from the filtrate at room temperature. Finally, orange-yellow needle-like crystals were obtained, which was the target product. Yield 58.3%, mp 185.0~185.3 °C, Anal.Calc.For C 21 H 18 N4O2: C, 64.62; H, 4.62; N, 14.36%. Found: C, 64.70; H, 4.53; N, 14.33%. FT-IR (KBr, cm -1 ): 3428 (m, OH), 3066 (w, phenyl-H), 1703 (w, C=O), 1620 (s, C=C), 1548 (s, δ N-H ), 1492(s), 1328(s), 754(s).UV-Vis(nm), ν max :258(0.89),290.5(0.46),344(0.46),446.5(0.82). 1 HNMR (CDCl3, 400Hz) δ: 7.3~7.9 (m, 12H), 2.38 (s, 6H).
[0019] Implementation Plan 2: Measurement of Mycelial Growth Rate of Pleurotus ostreatus
[0020] Ligand L and its copper complex (L-Cu) and zinc complex (L-Zn) were dissolved in DMSO to prepare solutions with concentrations of 0.5 mg / ml, 1.0 mg / ml, and 2.0 mg / ml, respectively. 6.0 ml of each DMSO solution was mixed with 594 ml of PDA medium while hot, sterilized at 121°C for 0.5 h, poured onto plates, and inoculated with pre-activated *Pleurotus ostreatus* mycelium. The plates were then incubated at 22°C. Plaque diameters were measured on the fifth and tenth days of incubation, and mycelial growth rate and inhibition rate were calculated. The results are shown in the table below.
[0021] Table 1. Growth rate and inhibition rate (%) of *Pleurotus ostreatus* mycelium.
[0022]
[0023] The data in the table show that the inhibition rate of ligand L at a concentration of 2 mg / mL on the mycelium of Pleurotus ostreatus is negative, indicating that it does not inhibit the growth of the mycelium, but rather promotes it.
[0024] Implementation Plan 3: Assay of Succinate Dehydrogenase Activity in Mushrooms of the White Jade Plant
[0025] A modified MTT assay was used. The stipes of *Pleurotus ostreatus* were divided into 0.5g segments. In a 25mL centrifuge tube, 0.5g of stipe, 5mL of distilled water, 0.2mL of DMSO solution, and 0.4mL of 2.5mg / mL MTT solution were added sequentially. The centrifuge tube was incubated at 35°C in the dark for 1 hour. The solution was then removed, leaving only the stipe segments. 2mL of DMSO was added to the centrifuge tube, and the tube was shaken for 1.5 hours until the stipe segments returned to their white color. The stipe segments were then removed, and the absorbance of the DMSO solution was measured at 490nm. Trial measurements were performed, with a blank obtained without the compound solution. The activation rate of succinate dehydrogenase was calculated using the following formula:
[0026]
[0027] Table 2. Activation rate (%) of succinate dehydrogenase in the stem of *Agaricus blazei*.
[0028]
[0029]
[0030] The data in the table show that ligand L at a concentration of 2 mg / mL activated succinate dehydrogenase in the stalk of *Agaricus bisporus* by 91%, indicating that it has a significant activating effect on succinate dehydrogenase in the stalk of *Agaricus bisporus*.
Claims
1. A method for preparing bis(4-pyrazolone) methane compounds, which comprises adding a mixture of 1-phenyl-3-methyl-4-trifluoroacetyl pyrazolone-5 and benzoyl hydrazine in an organic mixed solvent, heating to reflux at a certain temperature for 4-6 hours, then filtering while hot, and allowing the filtrate to evaporate the solvent naturally at room temperature to obtain the product bis-(1-phenyl-3-methyl-pyrazolone-5) methane.
2. The method for preparing a bis(4-pyrazolone)methane compound as described in claim 1, characterized in that: The molar ratio of 1-phenyl-3-methyl-4-trifluoroacetyl pyrazolone-5 to benzoyl hydrazine is 1:
1.
3. The method for preparing a bis(4-pyrazolinone)methane compound as described in claim 1, characterized in that: The organic mixed solvent consists of two components, one of which is dimethyl sulfoxide (DMSO), and the other is one of 95% methanol and 95% ethanol.
4. The method for preparing a bis(4-pyrazolone)methane compound as described in claim 3, characterized in that: The volume ratio of DMSO to 95% methanol in the organic mixed solvent is 5:
1.
5. The method for preparing a bis(4-pyrazolinone)methane compound as described in claim 3, characterized in that: The volume ratio of DMSO to 95% ethanol in the organic mixed solvent is 5:
1.
6. Use of the end product of claim 1 as a mycelium growth promoter, characterized in that Some of the target compounds have obvious growth-promoting activity on the mycelium of White Jade Fungus.
7. Use of the end product of claim 1 as a succinate dehydrogenase promoter in the preparation of succinic acid, characterized in that Some of the target compounds have obvious promoting effect on the succinate dehydrogenase activity of the White Jade Fungus stipe.