Single-channel double-target weeding method for targeting branched chain amino acid biosynthesis

Abstract

The invention belongs to the technical field of agricultural chemicals, and relates to a single-channel double-target weeding method for targeting branched chain amino acid biosynthesis, in particular to a single-channel double-target weeding method realized by combined use of an acetolactate synthase inhibitor and a keto-alcohol acid reductoisomerase inhibitor. According to the weeding method, an AHAS herbicide and a KARI inhibitor are compounded to serve as active ingredients to achieve the purpose of synergistic interaction, and the mass percent of the AHAS herbicide and the mass percent of the KARI inhibitor in the active ingredients are 1%: 99%-99%: 1% respectively. When weeding is carried out according to a specific proportion, the inhibition rate of the rape root length is measured on a plate model, the synergistic multiple can reach 10.43, and when weeding activity evaluation is carried out on a potted model, the synergistic multiple can reach 6.79. AHAS herbicides, including all commercial herbicides targeting AHAS, especially mesosulfuron, nicosulfuron, penoxsulam, bispyribac-sodium, imazapyr and triafamone, and KARI inhibitors, including all possible KARI inhibitors, especially the following three compounds

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of agricultural chemicals, and relates to a single-channel double-target herbicidal method targeting branched-chain amino acid biosynthesis, in particular to a single-channel double-target herbicidal method achieving synergistic effect by jointly using acetolactate synthase inhibitors and ketol-acid reductoisomerase inhibitors. BACKGROUND

[0002]

[0003] Valine, leucine and isoleucine, three branched-chain amino acids, can only be synthesized in vivo by plants and microorganisms, and there is no such biochemical process in the mammalian body. Therefore, chemicals capable of inhibiting branched-chain amino acid biosynthesis can be used as herbicides or antibacterial drugs, and have biological safety to humans and livestock (McCourt JA, et al. Amino Acids, 2006, 31(2), 173-210). The biosynthetic pathway thereof is shown above. As can be seen, for the three branched-chain amino acids, although the complete biosynthetic pathways are different, they must all undergo the biological processes catalyzed by acetolactate synthase (AHAS), ketol-acid reductoisomerase (KARI) and dihydroxy acid dehydratase (DHAD). The biochemical reactions catalyzed by the three enzymes belong to the series relationship of upstream, midstream and downstream.

[0004] In fact, the first enzyme acetolactate synthase AHAS in this process is a very successful herbicide target. Herbicides targeting AHAS include sulfonylurea, imidazolinone, pyrimidinyl oxybenzoic acid and triazolopyrimidine sulfamide, etc. There are more than 60 commercialized varieties to date (Wang Jianguo, Journal of Pesticide Science, 2014, 16(4), 367-374). In particular, sulfonylurea, with a use amount of only 0.5-2 grams per mu, can efficiently achieve the herbicidal effect and protect the agricultural production. However, with the wide use of AHAS herbicides, weed resistance has gradually become an agricultural problem that cannot be ignored. Generally, as long as weeds have resistance to a certain herbicide, they soon have resistance to herbicides with the same mechanism to varying degrees (Yu Q, et al. Pest Manag. Sci., 2014, 70(9), 1340-1350). Studies have shown that weed resistance is divided into target resistance and non-target resistance, of which the proportion of target resistance is the largest.

[0005] Keto-aldonic acid reductase KARI is the second enzyme in the biosynthesis of branched-chain amino acids, downstream of AHAS, but so far no herbicide targeting KARI has been found to be as highly effective as AHAS inhibitors. Hoe 704, IpOHA and cyclopropane dicarboxylic acid (CPD) are representative compounds found internationally with relatively strong inhibitory activity on plant-derived KARI (Wang BL, et al., Bioorg. Med. Chem. Lett. 2017, 27(24), 5457-5462; Schulz A, et al., FEBS Lett. 1988, 238(2), 375-378; Lee YT, et al. Plant Sci. 2005, 168, 1035-1040.), but these inhibitors have weak herbicidal effects under pot conditions.

[0006]

[0007] Recently, Guddat and Wang Jianguo et al. found that a five-membered heterocyclic pyrimidinedione compound code (JK-5-67) has strong inhibition on plant-derived KARI, and has strong herbicidal effect on dicotyledonous weeds Amaranthus retroflexus and Brassica napus under pot test at a dose of 100 g / acre, whether stem and leaf treatment or soil treatment (Lin X, et al., Chem. Eur., 2025, 31, e202501158), providing an important starting point for further design and discovery of new types of KARI herbicides (Wang Jianguo et al., Chinese Invention Patent, CN119896232A). Wang Jianguo et al. further designed and synthesized a batch of heterocyclic pyrimidinedione compounds, which showed inhibition effect on plant KARI and had good pesticide and medical biological activity, among which compound SXW-1 had good herbicidal effect on representative broadleaf weeds Brassica napus and representative grass weeds Echinochloa crus-galli at a dose of 100 g / acre (Wang Jianguo et al., Chinese Invention Patent, CN120623197A).

[0008]

[0009] Since AHAS and KARI two targets catalyze two closely connected biochemical reactions respectively, the product of the biochemical reaction catalyzed by AHAS is the raw material of the biochemical reaction catalyzed by KARI, so theoretically if AHAS is inhibited, the raw material of the reaction catalyzed by KARI will be reduced, and at this time the KARI inhibitor can achieve the effect of inhibiting KARI with smaller dosage (such as Figure 4It was observed that IpOHA and metsulfuron-methyl produced a synergistic effect in the presence of mycobacterial inhibitors (Grandoni JA, et al., J. Antimicrob. Chemother. 1998, 42, 475−482.), but this synergistic effect against Mycobacterium tuberculosis was not observed with all AHAS inhibitors, and this was not explored in the field of herbicides. SUMMARY

[0010] The present application aims to provide a single-channel dual-target herbicidal method targeting branched-chain amino acid biosynthesis, in particular a single-channel dual-target herbicidal method realized by the combined use of acetolactate synthase inhibitors and ketol-acid reductoisomerase inhibitors, which realizes a good synergistic effect by mixing AHAS herbicides and KARI inhibitors.

[0011] The acetolactate synthase inhibitors used in the present application are all known commercial AHAS herbicides, including sulfonylurea, imidazolinone, pyrimidinyl oxybenzoic acid, triazolopyrimidine sulfonamide and triazolinone, etc.

[0012] The acetolactate synthase inhibitors mentioned in the present application can be any one of the following herbicides

[0013]

[0014]

[0015]

[0016] The acetolactate synthase inhibitors further preferred in the present application are mesosulfuron-methyl, nicosulfuron, penoxsulam, bensulfuron-methyl, nicosulfuron and flucarbazone.

[0017] The ketol-acid reductoisomerase inhibitors mentioned in the present application are all known KARI inhibitors.

[0018] The ketol-acid reductoisomerase inhibitors further preferred in the present application are one of the following three KARI inhibitors.

[0019]

[0020] The present application provides a herbicidal method, which comprises the acetyl-lactate synthase inhibitor and the ketol-acid reductoisomerase inhibitor as the active ingredients, and the mass percentage of the acetyl-lactate synthase inhibitor and the ketol-acid reductoisomerase inhibitor is 1:99 to 99:1. When the herbicidal method is used in a specific proportion, the inhibition rate of rape root length is determined in a plate model, and the synergistic effect multiple is up to 10.43. When the herbicidal activity is evaluated in a pot model, the synergistic effect multiple is up to 6.79.

[0021] The mass ratio of the acetyl-lactate synthase inhibitor and the ketol-acid reductoisomerase inhibitor is 20:1 to 1:20; preferably, the mass ratio of the acetyl-lactate synthase inhibitor and the ketol-acid reductoisomerase inhibitor is 10:1 to 1:10; further preferably, the mass ratio of the acetyl-lactate synthase inhibitor and the ketol-acid reductoisomerase inhibitor is 5:1 to 1:5.

[0022] In the plate model test, the addition of exogenous branched-chain amino acids significantly reduces the inhibition of rape root length until it is completely restored, indicating that the herbicidal method targets a single channel of branched-chain amino acid biosynthesis.

[0023] In the pot model test, the mixture of mesosulfuron-methyl and the KARI inhibitor shows safety to wheat under the condition of synergistic effect; in the pot model test, the mixture of penoxsulam and the KARI inhibitor shows safety to rice under the condition of synergistic effect; in the pot model test, the mixture of nicosulfuron and the KARI inhibitor shows safety to corn under the condition of synergistic effect.

[0024] The present application also provides a herbicide compound composition, which comprises the acetyl-lactate synthase inhibitor and the ketol-acid reductoisomerase inhibitor as the active ingredients, and the mass percentage of the acetyl-lactate synthase inhibitor and the ketol-acid reductoisomerase inhibitor is 1:99 to 99:1, and the compound composition comprises 1% to 99% by weight of the active ingredients, 99% to 1% by weight of solid or liquid adjuvant, and 0 to 25% by weight of surfactant.

[0025] The dosage form of the composition provided by the present application is emulsifiable concentrate, wettable powder, soluble powder, emulsion in water, microemulsion, aqueous solution, suspension, microcapsule or water dispersible granule. BRIEF DESCRIPTION OF DRAWINGS

[0026] Figure 1 is the branched-chain amino acid recovery experiment effect (corresponding to Table 26 of Example 4); Figure 2 is the toxicity regression formula of stem and leaf treatment for preventing and controlling amaranthus retroflexus when the ratio of penoxsulam and JK-5-67 is 2:3 (corresponding to Table 34 of Example 5); Figure 3 Safety test of mesosulfuron-methyl and KARI inhibitor on wheat (corresponding to Table 37 of Example 6); Figure 4 Schematic diagram of a single-channel dual-target herbicidal method targeting branched-chain amino acid biosynthesis. DETAILED DESCRIPTION

[0027] The essential characteristics of the present application can be embodied from the following examples, but these examples are only illustrative and not limiting to the present application.

[0028] Example 1. Preparation of compound XQ-2

[0029] The chemical reagents used in this example were purchased through the online store of Nankai University, and the methanol, ethanol, petroleum ether, ethyl acetate, dichloromethane, acetonitrile, dimethylbenzene, column chromatography silica gel, ammonia water, DMF were purchased from Guangda Service Department of Hedong District, Tianjin. Toluene, hydrochloric acid, acetone, methyl tert-butyl ether were purchased from Tianjin Chemical Reagent Supply and Sales Company. Hydroxylamine hydrochloride, sodium ethoxide were purchased from Shanghai Bide Technology Co., Ltd. Benzenesulfonyl chloride was purchased from Beijing Bailingwei Company. Various amides, urea raw materials were purchased from Shanghai Titan Reagent Co., Ltd.

[0030]

[0031] After 1 (5.0 g, 46.25 mmol) was dissolved in 12 mL of acetonitrile, 2 (9.3 mL, 50.88 mmol) was added, stirred and warmed to 80°C for 90 min, and after the generation of a new product was detected by TLC, it was concentrated under reduced pressure to obtain a black viscous liquid. After 10 mL of dimethylbenzene was added and mixed evenly with the product, it was warmed to 125°C and stirred for 12 hours. After the room temperature was restored, it was concentrated under reduced pressure to as little solvent as possible, the organic phase was collected and dissolved in acetone to form a homogeneous solution, dried with anhydrous magnesium sulfate (15 g), filtered and column chromatographed to obtain a light yellow solid powder 3 with a yield of 75%.

[0032]

[0033] 3 (1.0 g) was stirred with 6M hydrochloric acid (10 mL) and warmed to 95°C for 8 hours. After cooling to room temperature, it was adjusted to pH = 10 with 15% NaOH, the impurities were filtered off, and the maximum solid was precipitated by adjusting to 2M hydrochloric acid, washed, filtered under reduced pressure and dried to obtain white solid 4 with a yield of 63%.

[0034]

[0035] Dissolve 4 (5.88 mmol) in 10 mL of methanol, add dropwise dichlorosulfoxide (1.28 mL, 17.64 mmol), and heat to reflux for 12 hours. After the reaction is completed, cool and neutralize with 15% NaOH, and precipitate the solid. After filtration, wash with water and dry to obtain white solid 5 in a yield of 62%.

[0036]

[0037] Under argon protection, stir hydroxylamine hydrochloride (0.56 g, 8.2 mmol) and 30% sodium methoxide (1.6 mL, 8.2 mmol) in a methanol solution (5 mL) for 3 hours. Quickly filter to obtain a hydroxylamine solution. Under an ice bath, inject a mixture of 5 (0.5 g, 2.7 mmol) and methanol (3 mL) into the hydroxylamine solution, and stir for 24 hours. Obtain white solid 6 in a yield of 70%.

[0038] Stir 6 (0.49 g, 2.6 mmol) and benzenesulfonyl chloride (0.71 mL, 2.6 mmol) in 15% NaOH (8 mL) at room temperature for 4 hours to obtain a yellow liquid. Neutralize with 10% HCl to pH = 3, precipitate the solid, filter and dry to obtain white solid 7 in a yield of 49%.

[0039]

[0040] Stir 300 mg of 7 with 1 mol / L NaOH solution (4.5 mL) at 100°C for 3 minutes. After stopping heating, neutralize with 10% HCl solution to pH = 6-7 while hot, stir for 30 minutes, precipitate the solid, filter, wash and dry to obtain the target compound XQ-2 in a yield of 76%.

[0041] The physicochemical characterization data of XQ-2 are as follows:

[0042] Yellow solid, yield 65%, melting point > 250°C. 1 H NMR (400 MHz, DMSO-d6) δ 12.99 (s, 1H, OH), 11.83 (s, 1H, NH), 10.34 (s, 1H, CONH), 2.35 (s, 3H, CH3). 13 C NMR (101 MHz, DMSO-d6) δ 153.1, 151.1, 150.0, 146.2, 106.2, 14.7. High resolution mass spectrum (ESI) m / z calcd for C6H5N4O3 - [M-H] - 181.03671, found 181.03648.

[0043] The preparation method of compound JK-5-67 is described in the reference method (Wang Jianguo et al., Chinese Invention Patent, CN119896232A), and the preparation method of compound SXW-1 is described in the reference method (Wang Jianguo et al., Chinese Invention Patent, CN120623197A).

[0044] Example 2. Inhibition of rice KARI enzyme by compound XQ-2

[0045] The plant-derived KARI is from rice, with the cDNA code J013002N13. It was overexpressed using the commonly used *E. coli* BL21(DE3) system and purified by immobilized metal affinity chromatography (IMAC) to obtain pure KARI. The test was conducted at 30℃ in a 0.1M Tris-HCl buffer system (pH 8.0). The reaction solution contained 0.2 mM NADPH, 1 mM MgCl2, 0.1 mM acetyllactic acid, and different concentrations of small molecule inhibitors. During the experiment, the buffer solution was added to a 1 mL quartz tube, followed by 10 μL of rice KARI. The rate of NADPH disappearance was dynamically monitored. Its inhibition constant K... i The following formula can be used to calculate:

[0046] v = v o / (1+[I] / K i )

[0047] Among them, v o [I] represents the maximum catalytic reaction rate of rice KARI when it is not inhibited, [I] represents the concentration of the compound, and v represents the reaction rate. Table 1 lists the K values ​​of XQ-2 for rice KARI. i value.

[0048] Table 1. K-response of tested compounds to rice KARI i value

[0049] Compound Code Inhibition of KARI in rice K i values]]> XQ-2 85.4 μM SXW-1 20.1 μM Control drug JK-5-67 1.8 μM

[0050] Example 3. Synergistic effect experiment of plate model

[0051] A 5.6 cm diameter filter paper was placed in a 6 cm diameter petri dish, and 2 mL of a solution of the test compound of a certain concentration was added. Ten rapeseed seeds that had been soaked for 4 hours were then sown. After incubation in the dark at 28℃ for 72 hours, the radicle length was measured to detect the herbicidal activity of the compound. A blank control was set up. Plant height was measured. Analysis of variance was performed using DPS 18.10.

[0052] The data analysis method used is the co-toxicity coefficient method, as detailed below:

[0053] If the toxicological coefficient of commercial herbicide A is considered to be 100...

[0054]

[0055] Furthermore, if the toxicity difference between the two compounds is too large, the toxicity effect of the lower-activity inhibitor in the combination group is not considered. In this case, the theoretical toxicity number calculation of the combination group only refers to the one with higher toxicity. The reference data for discriminant synergy in the plate model is IC50. 50 and IC 65 When the synergistic ratio (SR) is less than 0.8, it is generally considered to be an antagonistic effect; when the SR is between 0.8 and 1.2, it is considered to have no significant effect (additive effect); when the SR is greater than 1.2, it is considered to be a synergistic effect.

[0056] The following table lists the inhibition and synergistic effects of different acetolactate synthase inhibitors (hereinafter referred to as AHAS herbicides) on rapeseed embryo root growth when combined with three ketool acid reductase inhibitors (hereinafter referred to as KARI inhibitors).

[0057] Table 2. Inhibition rate (%) of the combined use of JK-5-67 (B1) and mesosulfuron-methyl (A1) on the growth of rapeseed radicles.

[0058]

[0059] Table 3. Synergistic effect of JK-5-67 (B1) and mesosulfuron-methyl (A1) on the inhibition of rapeseed root length in a plate model.

[0060]

[0061] Table 4. Inhibition rate (%) of SXW-1 (B2) combined with mesosulfuron-methyl (A1) on the growth of rapeseed radicle.

[0062]

[0063] Table 5. Synergistic effect of SXW-1 (B2) and mesosulfuron-methyl (A1) on the inhibition of rapeseed root length in a plate model.

[0064]

[0065] Table 6. Inhibition rate (%) of XQ-2 (B3) combined with mesosulfuron-methyl (A1) on the growth of rapeseed radicle.

[0066]

[0067] Table 7. Synergistic effect of XQ-2 (B3) and mesosulfuron-methyl (A1) on the inhibition of rapeseed root length in a plate model.

[0068]

[0069] Table 8. Inhibition rate (%) of SXW-1 (B2) combined with nicosulfuron (A2) on the growth of rapeseed radicle.

[0070]

[0071] Table 9. Synergistic effect of SXW-1 (B2) and nicosulfuron (A2) on the inhibition of rapeseed root length in a plate model.

[0072]

[0073] Table 10. Inhibition rate (%) of XQ-2 (B3) and nicosulfuron (A2) on the growth of rapeseed radicles

[0074]

[0075] Table 11. Synergistic effect of XQ-2 (B3) and nicosulfuron (A2) on the inhibition of rapeseed root length in a plate model.

[0076]

[0077] Table 12. Inhibition rate (%) of rapeseed radicle growth by combination of JK-5-67 (B1) and penoxsulam (A3)

[0078] Table 13. Synergistic effect of JK-5-67 (B1) and penoxsulam (A3) on the inhibition of rapeseed root length in a plate model.

[0079]

[0080] Table 14. Inhibition rate (%) of SXW-1 (B2) combined with penoxsulam (A3) on rapeseed radicle growth.

[0081]

[0082] Table 15. Synergistic effect of SXW-1 (B2) and penoxsulam (A3) on the inhibition of rapeseed root length in a plate model.

[0083]

[0084]

[0085] Table 16. Inhibition rate (%) of XQ-2 (B3) combined with penoxsulam (A3) on rapeseed radicle growth.

[0086] Table 17. Synergistic effect of XQ-2 (B3) and penoxsulam (A3) on the inhibition of rapeseed root length in a plate model.

[0087]

[0088] Table 18. Inhibition rate (%) of rapeseed radicle growth by the combination of JK-5-67 (B1) and methomyl (A4).

[0089]

[0090] Table 19. Synergistic effect of JK-5-67 (B1) and fenbac (A4) on the inhibition of rapeseed root length in a plate model.

[0091]

[0092] Table 20. Inhibition rate (%) of the combination of JK-5-67 (B1) and bispyribac-sodium (A5) on the growth of rapeseed radicles.

[0093]

[0094]

[0095] Table 21. Synergistic effect of JK-5-67 (B1) and bispyribac-sodium (A5) on root growth inhibition in rapeseed using a plate model.

[0096]

[0097] Table 22. Inhibition rate (%) of the combined use of JK-5-67 (B1) and flufenoxuron (A6) on the growth of rapeseed radicles.

[0098] Table 23. Synergistic effect of JK-5-67 (B1) and flufenoxuron (A6) on the inhibition of rapeseed root length in a plate model.

[0099]

[0100] Table 24. Inhibition rate (%) of SXW-1 (B2) combined with flufenoxuron (A6) on rapeseed radicle growth.

[0101]

[0102] Table 25. Synergistic effect of SXW-1 (B2) and flufenoxuron (A6) on the inhibition of rapeseed root length in a plate model.

[0103]

[0104]

[0105] The results in the table above show that the AHAS herbicide and KARI inhibitor used in the examples, when combined in specific ratios, can achieve synergistic effects. In particular, when penoxsulam (A3) and SXW-1 (B2) are combined in a 1:1 ratio, the IC50... 50 The synergistic effect can reach 10.43.

[0106] Example 4. Branched-chain amino acid recovery experiment

[0107] Given that both AHAS herbicides and KARI inhibitors act on the branched-chain amino acid biosynthesis pathway, in order to clarify whether the combined treatment still targets this pathway rather than generating a new mechanism of action, this study verified the pathway specificity through an exogenous addition experiment of branched-chain amino acids (valine, leucine, and isoleucine).

[0108] Test drugs: penoxsulam (A3), JK-5-67 (B1), leucine, isoleucine, valine

[0109] Test method:

[0110] (1) Set up a blank group; two single-dose groups, A3 and B1; a compound group A3:B1=1:4; a control group and a recovery group.

[0111] (2) Preparation of drug solution: Dilute the two stock solutions from 10000 mg / L to 10 mg / L.

[0112] A3 single-dose group: 200 μL of stock solution was added to 9.8 mL of emulsion water to obtain an initial dose of 0.2 μg / mL, and then it was serially diluted in a 5-fold gradient (2 mL of super solution was added to 8 mL of emulsion water);

[0113] B1 single-agent group: 24 μL of secondary mother liquor was added to 12 mL of emulsified water to obtain a primary concentration of 2 μg / mL, and then serially diluted twice to 0.125 μg / mL (6 mL of primary mother liquor was added to 6 mL of emulsified water);

[0114] Compound group: The labeled concentration is the mixed concentration of two solutions. Taking 1:4 as an example, take 40μL of A3 stock solution and 160μL of B1 stock solution, add 9.8mL of emulsified water to obtain an initial dose with a mixed concentration of 0.2μg / mL, and then dilute stepwise in a 5-fold gradient.

[0115] Recovery group emulsified water: Based on the preparation of normal emulsified water, add three amino acids to it at a concentration of 0.5mM and mix evenly.

[0116] Table 26. Inhibition rate (%) of branched-chain amino acid recovery experiment on rapeseed radicle growth

[0117]

[0118] Exogenous compensation experiments using branched-chain amino acids (isoleucine, leucine, and valine, 0.5 mM each) revealed that the synergistic toxicity effect of the herbicide combination was manifested in the compensation group as a significant reduction in seedling root growth inhibition rate (recovery rate ≥60%). At the highest dose of 0.1 mg / L, the inhibition rate significantly decreased from 79.3% to 16.4%, with a recovery rate of 79.3%. At lower doses, complete recovery was observed, and the physiological damage phenotype was completely eliminated. These results validate that the synergistic mechanism of AHAS herbicide and KARI inhibitor still operates on the branched-chain amino acid biosynthesis pathway.

[0119] Example 5. Synergistic effect experiment of potted plant model

[0120] The bonsai model uses ED as required. 50 ED 80 ED 65 ED 90 and ED 40 Different indicators are used to describe the weeding effect. (I) Synergistic effect of KARI inhibitors combined with mesosulfuron-methyl (A1)

[0121] 1. Experimental materials:

[0122] Seeds used in the test: Jointed goatgra seeds were collected by the test team. The foliar treatment was performed on seedlings at the two-leaf to three-leaf stage, while the soil treatment was performed by direct sowing of seeds.

[0123] Test reagents: Mesosulfuron technical grade (A1), JK-5-67 (B1), SXW-1 (B2)

[0124] 2. Experimental Design:

[0125] (1) Ratio setting: The synergistic ratio in the plate model is A1:B1 = 1:4 and 3:2. Since the total amount of medicine used under the 1:4 configuration is significantly more than the total amount of medicine used under the 3:2 ratio, 3:2 is regarded as the optimal ratio, and compounding is carried out at the pot level around this ratio.

[0126] (2) Preparation of drug solution: Dilute the stock solution from 10000 mg / L to 1000 mg / L

[0127] Three replicates (6 cups in total), using 0.1m 2 The tray was used for spraying, with a total spray volume of 4 mL.

[0128] For single-dose group A1: 30 g / ha, the preparation method is to take 300 μL of stock solution containing 1 mg / 1 mL and add 3.7 mL of emulsified water. The same method applies to lower concentrations.

[0129] Compound group: Taking a 1:1 ratio as an example, take 300 μL of each of the two stock solutions at 1000 mg / L and add 3.4 mL of emulsified water. This is to fix the content of AHAS herbicide and compound it with different proportions of KARI inhibitors. Detailed dosage settings and fresh weight inhibition results are shown in Table 27.

[0130] Table 27. Herbicidal effect (%) of mesosulfuron-methyl (A1) combined with KARI inhibitor on jointed goatgrass.

[0131]

[0132] Tables 28 and 29 list the synergistic effects of mesosulfuron-methyl with different KARI inhibitors during potted plant treatment.

[0133] Table 28. Synergistic effect of mesosulfuron-methyl (A1) and KARI inhibitors on jointed goatgrass (stem and leaf treatment)

[0134]

[0135]

[0136] Table 29. Synergistic effect of mesosulfuron-methyl (A1) and KARI inhibitors on jointed goatgrass (soil treatment)

[0137]

[0138] It can be seen that, in pot cultivation, mesosulfuron-methyl (A1) and KARI inhibitors also exhibit synergistic effects, especially in foliar treatment, where mesosulfuron-methyl (A1) and JK-5-67 (B1) at a 4:1 ratio showed the best effect on efflux. 80 The synergistic effect factor is 3.33.

[0139] (ii) Synergistic effect of KARI inhibitors combined with penoxsulam (A3)

[0140] 1. Experimental materials:

[0141] Test plant: Barnyard grass (collected from Taitou Town, Jinghai District, Tianjin). The foliar treatment was performed on seedlings at the two-leaf to two-leaf stage with a central bud, while the soil treatment involved direct sowing of germinated seeds.

[0142] Tested pesticides: penoxsulam technical (A3), JK-5-67 (B1).

[0143] 2. Experimental Design:

[0144] (1) Ratio setting: Since the combination of the three KARI inhibitors and penoxsulam (A3) had a synergistic effect in the plate experiment at different ratios, four different ratios of A3:B1 were set in the pot experiment: 1:4; 2:3; 3:2; 4:2.

[0145] (2) Preparation of drug solution: Dilute the mother solution from 10000 mg / L to 1000 mg / L.

[0146] For the single-dose group: the preparation method for 15 g / ha is to take 150 μL of stock solution and add 3.85 mL of emulsified water.

[0147] Compound group: Taking a 1:4 ratio of 15 g / ha as an example, the preparation method is to take 150 μL of A3 stock solution and 600 μL of B1 stock solution, mix them, and then add 3.25 mL of emulsified water. The dosage settings and fresh weight inhibition results are shown in Table 30.

[0148] Table 30. Herbicidal efficacy (%) of penflusulfonamide (A3) combined with JK-5-67 (B1) against barnyardgrass.

[0149]

[0150] Tables 31 and 32 list the synergistic effects of mesosulfuron-methyl with different KARI inhibitors in potted plants.

[0151] Table 31. Synergistic effect of penoxsulam (A3) and JK-5-67 (B1) on barnyardgrass (foliar treatment)

[0152]

[0153] Table 32. Synergistic effect of penflusulfonamide (A3) and JK-5-67 (B1) on barnyardgrass (soil treatment)

[0154]

[0155] Similarly, a synergistic effect study was conducted on penoxsulam (A3) in combination with a KARI inhibitor against amaranth. The dosage settings and fresh weight inhibition effects are shown in Table 33, and the synergistic effect results are shown in Table 34.

[0156] Table 33. Herbicidal effect (%) of KARI inhibitor combined with penoxsulam (A3) on antibranch amaranth.

[0157]

[0158] Table 34. Synergistic effect of KARI inhibitor combined with penoxsulam (A3) on anti-branch amaranth

[0159]

[0160]

[0161] It can be seen that, in pot cultivation, penoxsulam (A3) and the KARI inhibitor also exhibit a synergistic effect. The synergistic effect is particularly pronounced when controlling amaranth retroflexus; when the ratio of penoxsulam (A3) to JK-5-67 (B1) is 2:3, its ED50 is significantly enhanced. 90 The synergistic effect can reach 6.79.

[0162] (iii) Synergistic effect of KARI inhibitors combined with nicosulfuron-velvetleaf

[0163] 1. Test materials:

[0164] The tested plant was jute. For the stem and leaf treatment, seedlings at the two to three leaf stage were selected. For the soil treatment, germinating seeds were sown.

[0165] Test reagents: Nicosulfuron technical grade (A2), JK-5-67 (B1), SXW-1 (B2)

[0166] 2. Experimental Design:

[0167] (1) Dosage setting: Since no significant synergistic ratio was found in the plate experiment, the pot experiment was conducted according to the principle of reducing the dosage, and three ratios of A2+B were set: 1:1; 3:2; 4:1, for a total of six groups.

[0168] (2) Preparation of the drug solution: The concentration of the stock solution is 10000 mg / L;

[0169] Three replicates (6 cups), using 0.1m 2 The total amount of pesticide sprayed was 4 mL;

[0170] Single-agent group: Taking 90g / ha as an example, the preparation method is to add 3.9mL of emulsified water to 90μL of A2 stock solution;

[0171] Compound group: Taking 90g / ha at a 1:1 ratio as an example, the preparation method is to mix 90μL of A2 stock solution and 90μL of B stock solution, and add 3.8mL of emulsified water. Detailed dosage settings and fresh weight inhibition results are shown in Table 35. Synergistic effects are shown in Table 36.

[0172] Table 35. Herbicidal effect (%) of KARI inhibitor combined with nicosulfuron (A2) on velvetleaf.

[0173]

[0174] Table 36. Synergistic effect of KARI inhibitor combined with nicosulfuron (A2) on velvetleaf.

[0175]

[0176]

[0177] It can be seen that, in potted plant mode, nicosulfuron (A2) and KARI inhibitors also exhibit a synergistic effect. When the ratio of nicosulfuron (A2) to SXW-1 (B2) is 4:1, its ED... 40 The synergistic effect can reach 3.41.

[0178] Example 6. Crop safety study of the combination of KARI inhibitor and AHAS herbicide

[0179] Safety tests were conducted on wheat (the main crop for which mesosulfuron (A1) is mainly used), rice (the main crop for which penoxsulam (A3) is mainly used), and maize (the main crop for which nicosulfuron (A2) is mainly used).

[0180] (a) Safety tests of wheat

[0181] To clarify whether the combination of AHAS inhibitor mesosulfuron (A1) and KARI inhibitor, while having a synergistic effect, would affect crop safety, a greenhouse pot experiment was conducted to study the dose-response relationship of different ratios and application methods on wheat (Triticum aestivum L). The details are as follows.

[0182] 1. Experimental materials:

[0183] Test seeds: Wheat: Yu Nong variety. Foliar treatment was performed by spraying wheat seedlings at the two-leaf to three-leaf stage. Soil treatment was performed by pre-emergence spraying of germinating seeds after sowing.

[0184] Test reagents: Mesosulfuron technical grade (A1), JK-5-67 (B1), SXW-1 (B2)

[0185] 2. Experimental Design:

[0186] (1) Proportion setting: Based on the synergistic effect results, the proportions of the compound groups were set as follows: A1:B1 = 3:2; 4:1, A1:B2 = 1:1; 3:2. Three groups were replicated (6 cups), and the pesticide was sprayed using a 0.1m2 tray. The total spraying volume was 4mL.

[0187] (2) Preparation of drug solution: Dilute the stock solution from 10000 mg / L to 1000 mg / L

[0188] Under normal circumstances, the application rate of mesosulfuron-methyl (A1) in wheat fields is 7.5-15 g / ha. Therefore, three concentrations of 7.5 g / ha, 15 g / ha and 30 g / ha were set in the safety test.

[0189] For single-dose group A1 (30 g / ha), the preparation method is to take 300 μL of a 1000 mg / L stock solution and add 3.7 mL of emulsified water. The same method applies to lower concentrations.

[0190] Compound group: Taking 3:2 as an example, take 300μL of 1mg / 1mL A1 stock solution and 200μL of B, then add 3.5mL of emulsified water to obtain a mixed solution with A1 content of 30g / ha and a ratio of 3:2.

[0191] 3. Data Survey and Statistical Analysis

[0192] After 21 days, the effects of the drug combination on the crop were evaluated based on the drug dosage and the fresh weight of the aboveground parts of the plant.

[0193] The fresh weight inhibition is shown in Table 37.

[0194] Table 37. Inhibition rate (%) of wheat by combination of mesosulfuron-methyl (A1) and KARI inhibitor.

[0195]

[0196] It can be seen that, under the experimental conditions, when the AHAS herbicide mesosulfuron (A1) and two KARI inhibitors were combined in different proportions, mesosulfuron (A1) showed weaker inhibition of wheat when used alone, indicating that this weeding method has advantages in terms of safety.

[0197] (ii) Safety tests on rice

[0198] To clarify whether the combination of the AHAS inhibitor penoxsulam (A3) and the KARI inhibitor, under synergistic conditions, would affect crop safety, a greenhouse pot experiment was conducted to study the dose-response relationship of different ratios and application methods on the growth of rice (Oryza sativa L). The details are as follows.

[0199] 1. Experimental materials:

[0200] The tested plant was rice, specifically the variety Hanyou 73 indica rice. Foliar treatment involved spraying seedlings at the two-leaf to three-leaf stage, while soil treatment involved pre-emergence spraying of germinating seeds after sowing (10 plants per cup for both treatments). The soil used for planting was a mixed nutrient soil.

[0201] Test reagents: penoxsulam technical grade (A3), JK-5-67 (B1), SXW-1 (B2)

[0202] 2. Experimental Design:

[0203] (1) Dosage setting: Based on the above test results and adhering to the principle of reducing medication, three dosages were set in this group of tests: A3:B = 1:1; 3:2; 4:1. Similarly, for KARI single dose, we also set a dosage of 60g / ha for safety testing.

[0204] (2) Preparation of medicine solution: The concentration of the mother liquor is 10000 mg / L.

[0205] Three replicates (6 cups), using 0.1m 2 The tray was used for spraying, with a total spray volume of 4 mL.

[0206] Single-dose group: Taking 60g / ha as an example, the preparation method is to add 3.9mL of emulsified water to 60μL of stock solution.

[0207] Compound group: Taking 60g / ha at a 1:1 ratio as an example, the preparation method is to add 60μL of A3 stock solution and 60μL of B stock solution to 3.9mL of emulsified water.

[0208] 3. Data Survey and Statistical Analysis

[0209] After 21 days, the effects of the drug combination on the crop were evaluated based on the drug dosage and the fresh weight of the aboveground parts of the plant.

[0210] The fresh weight inhibition is shown in Table 38.

[0211] Table 38. Inhibition rate (%) of rice by stem and leaf treatment with KARI inhibitor and penoxsulam (A3)

[0212]

[0213] It can be seen that, under the experimental conditions, when the AHAS herbicide penoxsulam (A3) and two KARI inhibitors were combined in different proportions, penoxsulam (A3) showed weaker inhibition on rice when used alone, indicating that this weeding method has an advantage in terms of safety.

[0214] (iii) Safety tests on corn

[0215] To clarify whether the combination of the AHAS inhibitor nicosulfuron (A2) and the KARI inhibitor, under synergistic conditions, would affect crop safety, a greenhouse pot experiment was conducted to study the dose-response relationship of different ratios and application methods on the growth of maize (Zeamays L.), as detailed below.

[0216] 1. Experimental materials:

[0217] Test plant: Maize, variety Xindan 66. Foliar treatment was performed by spraying seedlings at the two-leaf stage to the two-leaf-one-heart stage, and soil treatment was performed by pre-emergence spraying after sowing germinating seeds (5 plants per cup for both treatments), and the plants were planted in large cups.

[0218] Test reagents: Nicosulfuron technical grade (A2), JK-5-67 (B1), SXW-1 (B2), XQ-2 (B3).

[0219] 2. Experimental Design:

[0220] (1) Dosage setting: Based on the above experimental results and the principle of reducing medication, the dosage setting was A2:B = 4:1, and a single-dose group was set in this group of experiments.

[0221] (2) Preparation of medicine solution: The concentration of the mother liquor is 10000 mg / L.

[0222] Three replicates (6 cups), using 0.1m 2 The tray was used for spraying, with a total spray volume of 4 mL.

[0223] Single-agent group: Taking 90g / ha as an example, the preparation method is to add 3.9mL of emulsified water to 90μL of stock solution.

[0224] Compound group: Taking 90g / ha as an example, the preparation method is to add 90μL of A2 stock solution and 90μL of B stock solution to 3.8mL of emulsified water.

[0225] After 21 days, the effects of the drug combination on the crop were evaluated based on the drug dosage and the fresh weight of the aboveground parts of the plant.

[0226] 3. Data Survey and Statistical Analysis

[0227] The fresh weight inhibition is shown in Table 39.

[0228] Table 39. Inhibition rate (%) of maize by combination of KARI inhibitor and nicosulfuron (A2)

[0229]

[0230] It can be seen that, under the experimental conditions, the combination of AHAS herbicides nicosulfuron (A2) and JK-5-67 (B1) in a 4:1 ratio showed weaker inhibition of corn growth compared to nicosulfuron (A2) alone, indicating that this weeding method has an advantage in terms of safety.

Claims

1. A single-channel dual-target weeding method targeting branched-chain amino acid biosynthesis, which achieves synergistic weeding effect by combining acetolactate synthase inhibitor and ketool acid reductase inhibitor.

2. The weeding method according to claim 1, characterized in that... The acetolactate synthase inhibitor is 3. The weeding method according to claim 1, characterized in that... The acetolactate synthase inhibitor is 4. The weeding method according to claim 1, characterized in that... The ketool acid reductase inhibitor is 5. The weeding method according to claim 1, wherein the above-mentioned acetolactate synthase inhibitor and ketool acid reductase inhibitor are compounded as active ingredients, and the active ingredients contain acetolactate synthase inhibitor and ketool acid reductase inhibitor in a mass percentage of 1%:99% to 99%:1%.

6. The weeding method according to claim 1, which uses a combination of mesosulfuron-methyl and ketohydric acid reductase inhibitor, can achieve synergistic effect in weed control while being safe for wheat.

7. The weeding method according to claim 1, which uses penoxsulam and ketoxate reductase inhibitor in combination, can achieve synergistic effect of weeding while being safe for rice.

8. The weeding method according to claim 1, which uses nicosulfuron and ketoxate reductase inhibitor in combination, can achieve synergistic effect in weeding while being safe for corn.

9. A herbicide compound composition comprising an acetolactate synthase inhibitor and a ketoolactate reductase inhibitor as active ingredients, wherein the active ingredients comprise acetolactate synthase inhibitor and ketoolactate reductase inhibitor in a mass percentage ratio of 1%:99% to 99%:1%, the compound composition comprising 1% to 99% by weight of the active ingredient, 99% to 1% by weight of a solid or liquid adjuvant, and a surfactant selected from 0 to 25% by weight.

10. A herbicide synergistic composition comprising an acetolactate synthase inhibitor and a ketoolactate reductase inhibitor as active ingredients, wherein the mass ratio of the acetolactate synthase inhibitor to the ketoolactate reductase inhibitor is 20:1 to 1:

20.

11. The herbicide compound composition according to claim 9, characterized in that... Its dosage forms include emulsifiable concentrates, wettable powders, soluble powders, water emulsions, microemulsions, aqueous solutions, suspensions, microcapsules, or water-dispersible granules.

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