Pleuromutilin suspension and method for preparing same

Abstract

The present invention relates to a pleuromutilin suspension and a method for preparing the same. The pleuromutilin suspension contains 5-25 wt% pleuromutilin, 3-12 wt% surfactant, 0.5-2.5 wt% thickener, 2-6 wt% antifreeze agent, 0.3-0.5 wt% silicone antifoaming agent, and the balance is deionized water. It can be used to prevent and treat various agricultural diseases, such as rice bacterial leaf blight fungus, rice bacterial streak fungus, and citrus canker fungus.

Description

[Technical Field]

[0001] The present invention relates to the field of pesticide suspensions, and more particularly to a pesticide suspension of pleuromutilin for preventing and treating plant bacterial diseases. [Background technology]

[0002] Bacterial plant diseases are classified as the second-class plant disease, second only to viruses, and are characterized by rapid onset, widespread distribution, and difficulty in control. In recent years, due to changes in China's agricultural cultivation system and environmental climate, bacterial crop diseases such as rice bacterial leaf blight (Xoo), rice streak (Xoc), citrus canker (Xac), cabbage black rot, peach hole, Chinese cabbage soft rot, muskmelon angular spot, tobacco bacterial wilt, watermelon fruit rot, and tomato bacterial wilt have become increasingly prevalent in China. Chemical pesticides are the primary means of control for bacterial plant diseases, with copper compounds being the most commonly used. However, copper compounds have poor safety profiles and are prone to copper poisoning. Currently, there is a severe shortage of effective and safe chemical agents for prevention and treatment, resulting in significant losses in agricultural production.

[0003] Pleuromutilin suspension is a new type of fungicide independently developed by Guizhou University. It is characterized by high efficiency, low toxicity and low risk. It has excellent antibacterial activity against pathogens such as Xanthomonas oryzae (Xoo), Bacterial streak fungus (Xoc), Xanthomonas citri (Xac), Cabbage black rot fungus, Peach needle hole fungus, Chinese cabbage soft rot fungus, Muskmelon angular spot fungus, Tobacco bacterial wilt fungus, Watermelon fruit rot fungus, Tomato bacterial wilt fungus, Cucumber bacterial leaf blight fungus, Amorphophallus konjac bacterial leaf blight fungus, Grapevine canker fungus, Tomato canker fungus, Kiwi canker, Apple canker, Cucumber gray mold fungus, Pepper wilt fungus, Brassica napus, Wheat scab fungus, Potato late blight fungus and Blueberry root rot fungus. At a dosage of 200 μg / mL, it has excellent preventive effects against bacterial diseases such as rice bacterial leaf blight, rice bacterial streak, and citrus canker. Reference 1 describes the dissolution of commercially available trimmed pleuromutilin in DMSO, followed by dropwise addition to the medium to measure the growth inhibitory effect on Xanthomonas oryzae, Xanthomonas oryzae, and Phytophthora infestans at different final concentrations. However, the effects measured in the laboratory still require much research and verification before they can be applied in the field, and preparing an appropriate formulation is one of the challenges. Prior art documents Patent documents Patent Document 1: China Patent Publication No. CN114009443A Summary of the Invention

[0004] To solve the above-mentioned problems in the prior art, the present invention provides a 5-25% pleuromutilin aqueous suspension formulation, which contains the active ingredient pleuromutilin, a wetting and dispersing agent, a thickener, an antifreeze agent, an antifoaming agent, and water, where the weight ratios of each component are 5-25 wt%, 3-12 wt%, 0.5-2.5 wt%, 2-6 wt%, and 0.3-0.5 wt%, respectively, and water is added to make up 100%. Preferably, the weight ratio of pleuromutilin is 5-25 wt%, 5-20 wt%, 5-10 wt%, 10-20 wt%, or 10-25 wt%, and most preferably 10 wt% or 20 wt%.

[0005] The surfactants selected by the present application include dispersants or wetting agents, and are complexes of ionic or nonionic auxiliaries, such as polymeric amphiphilic anionic nonionic surfactant complexes, phosphate esters, lignin sulfonates, salts of phenolsulfonic acid or naphthalenesulfonic acid, salts of polyacrylic acid, sulfosuccinates, alkyl sulfonates, alkylaryl sulfonates, alkyl sulfates, lauryl ether sulfates, fatty alcohol sulfates, sulfonate formaldehyde condensates, sulfated hexadecane, sulfated heptadecane, sulfated octadecane, and sulfated fatty alcohol glycol ethers.

[0006] The antifoaming agent is a polydimethylsiloxane in a silicone antifoaming agent range of 0.3-0.5 wt%.

[0007] The antifreeze agent is an inorganic salt.

[0008] The thickener is one or both of magnesium aluminum silicate SF-40 and xanthan gum.

[0009] Preferably, the dampness The wetting and dispersing agent is a polymeric amphiphilic anionic nonionic surfactant complex SC29, sodium xylene sulfonate HAS040, sodium naphthalene sulfonate formaldehyde condensate NNO, and any one of sodium p-toluene sulfonate, sodium methanesulfinate, or polynaphthalene formaldehyde sulfonate sodium salt in a weight ratio of 1:1. but More preferably, the wetting and dispersing agent is composed of a polymeric amphiphilic anionic nonionic surfactant complex SC29, sodium xylene sulfonate HAS040, sodium naphthalene sulfonate formaldehyde condensate NNO, and sodium p-toluene sulfonate in a weight ratio of 2:2:2:1.5. but It is composed of 2:2:2:1.5.

[0010] Preferably, the thickener is 2% magnesium aluminum silicate SF-40 or a combination of 1% magnesium aluminum silicate and 0.05% xanthan gum.

[0011] The pleuromutilin suspension for preventing and treating plant bacterial diseases is prepared by mixing the dispersant, wetting agent, antifreeze, and water according to the weight percentages, stirring and dissolving, adding pleuromutilin and thickener according to the weight percentages, and mixing uniformly under the action of a shear. The mixed slurry is then pumped into a grinder for grinding, adding silicone antifoaming agent according to the weight percentages, and conducting particle size tests every hour. When the D90 is approximately 6.0 μm, stirring is stopped and the suspension is filtered to obtain a 20% pleuromutilin suspension. The particle size D90 of the suspension is 5-8 μm.

[0012] The innovation of this patent is as follows: The main significance of the developed dosage form is that, through extensive screening of wetting and dispersing agents, a pleuromutilin suspension and a pleuromutilin wettable powder were prepared to effectively improve the preventive effect of the active pharmaceutical ingredient against plant bacterial diseases, and the pleuromutilin suspension prepared by this invention has the characteristics of good water dispersibility and favorable storage stability of the active pharmaceutical ingredient.

[0013] 1. Screening of wetting and dispersing agents The 5-25% pleuromutilin suspensions prepared by combining polymeric amphiphilic anionic nonionic surfactant complexes with different types of surfactants were tested for their water dispersibility and decomposition rate after thermal storage. The specific results are shown in Table 1. [Table 1] As shown in Table 1, after screening different types of auxiliary agents, it was found that the addition of sulfonate auxiliary agents was beneficial to the stability of the drug substance. The pleuromutilin suspension formulated with polymeric amphiphilic anionic nonionic surfactant complex SC29, sodium xylene sulfonate HAS040, naphthalenesulfonic acid sodium salt formaldehyde condensate NNO, and sodium p-toluenesulfonate, sodium methanesulfinate, or polynaphthaleneformaldehydesulfonate sodium salt had good dispersibility, and the drug substance decomposition rate after thermal storage was controlled to within 5%.

[0014] To screen for the optimal dosage of auxiliary agents such as sodium p-toluenesulfonate, polynaphthalene formaldehyde sulfonate sodium salt, and sodium methanesulfinate, three different contents of 0.5%, 1.5%, and 3.0% were set and screening was carried out. The specific results are shown in Table 2. [Table 2] JPEG0007799367000002.jpg76170 As shown in Table 2, the decomposition rate of pleuromutilin after thermal storage was less than 5% at three different dosages. The price of sodium methanesulfinate is relatively high, and its water dispersibility is average. The addition of polynaphthalene formaldehyde sulfonic acid sodium salt causes the suspension to turn yellow and affects its appearance. The addition of sodium p-toluenesulfonate not only improves the stability of the drug substance, but also has a relatively low price and a good appearance of the suspension. Therefore, taking into consideration the overall cost and properties, the preferred wetting and dispersing agent formulation is SC29:HAS040:NNO:sodium p-toluenesulfonate=2:2:2:1.5.

[0015] 2. Screening of thickeners Aqueous suspensions often suffer from storage physical stability problems such as increased granule aggregation, settling water, and thickened lumps. The formulation is prone to settling water after long-term storage. Increasing viscosity can improve settling stability. If the viscosity is too high, the product will be difficult to tilt and will adhere strongly to the wall, making processing, production, and use difficult. With appropriate viscosity, the formulation will have good stability and a high suspension rate. Based on relevant theory and taking into account the unstable chemical properties of pleuromutilin, this experiment did not change the types of auxiliary agents required for the formulation selected in the previous section, but instead changed the viscosity by selecting the amounts of magnesium aluminum silicate and xanthan gum used. However, because increased viscosity can reduce the autodispersibility of aqueous suspensions, increasing the amount of dispersant NNO was used to improve aqueous dispersibility. Specific results are shown in Table 3. [Table 3] As shown in Table 3, adding too much xanthan gum will have a certain impact on both the fluidity and water dispersibility of the formulation. Therefore, in situations where water dispersibility is not affected, 2% magnesium aluminum silicate is selected as a thickener, or 1% magnesium aluminum silicate and 0.05% xanthan gum are combined to improve the sedimentation stability of the suspension, while increasing the NNO content to increase the water dispersibility of the formulation.

[0016] 3. Selection of antifoam dosage Many of the wetting and dispersing agents used in this application are sulfonates, which generate a large amount of bubbles during processing, affecting the polishing effect. Therefore, a silicone-based defoamer, such as polydimethylsiloxane, is usually added to the suspending agent. However, the amount of defoamer added should be appropriate; adding too much will reduce the surface tension of the system and make the layering phenomenon more pronounced. If too little is added, the defoaming effect will be insignificant. In this test, a general-purpose silicone-based defoamer was selected for testing, and the specific results are shown in Table 4. [Table 4] As shown in Table 4, if the amount of antifoaming agent used is small, the large amount of bubbles generated during the polishing process cannot be effectively resolved. If the amount used is kept to 0.3-0.5%, fewer bubbles are generated during the preparation process, which is advantageous for polishing.

[0017] 4. Selection of antifreeze Pesticide suspensions using water as a dispersion medium must maintain stability even under extremely cold and low-temperature conditions during storage, and typically require the addition of a certain amount of an appropriate antifreeze. Using glycerin as an antifreeze increases the viscosity of the system and reduces fluidity. Using ethylene glycol as an antifreeze improves formulation stability and fluidity, but increases the decomposition rate of the active pharmaceutical ingredient after hot storage. When sodium chloride is used as an antifreeze, very little precipitate appears at the bottom after cold storage, but this does not increase the degree of decomposition of the active pharmaceutical ingredient. Therefore, sodium chloride was selected as the optimal antifreeze.

[0018] 5. The effect of particle size on the stability of API after heat storage After determining the auxiliary agents, pleuromutilin aqueous suspensions with different particle sizes (Nos. 1-6) were prepared. After thermal storage, the content of the active ingredient was determined according to the pesticide thermal stability test method GB / T19136-2003. The decomposition rate of the active ingredient after thermal storage was also analyzed. The specific results are shown in Table 5. [Table 5] JPEG0007799367000005.jpg47170As shown in Figure 1 and Table 5, when the particle size of 20% pleuromutilin aqueous suspension prepared using the above additives is between 5 and 8 μm, it meets the stability requirements. This explains why, under the above conditions, the composition and particle size have a significant impact on the chemical stability of the drug substance in pleuromutilin aqueous suspension; the smaller the particle size, the better the chemical stability of the drug substance, but if it is too small, it is difficult to produce on an industrial scale.

[0019] 6. Determining the preferred formulation and preparation method [Table 6] JPEG0007799367000006.jpg57170

[0020] 7. 20% Pleuromutilin Suspension Control Item Index Standards In accordance with the national standard requirements for pesticide suspensions (GB / T19136-2003), the main properties of the suspension were tested, and are specifically shown in Table 7. The results show that under this formulation, the pleuromutilin aqueous suspension exhibits a milky white, uniform dispersion system, with relatively good dispersibility and suspension rate, and the drug substance is relatively stable after hot storage and stable at low temperatures. [Table 7] JPEG0007799367000007.jpg57170 [Brief explanation of the drawings]

[0021] [Figure 1] FIG. 1 is a particle size distribution diagram of a 20% pleuromutilin suspension. [Figure 2] 1 is a particle size distribution table for a 20% pleuromutilin suspension. [Figure 3] Protective activity of 20% pleuromutilin suspension at 200 μg / mL against rice bacterial leaf blight. [Figure 4] Therapeutic activity of 20% pleuromutilin suspension at 200 μg / mL against rice bacterial leaf blight. [Figure 5] Protective activity of 20% pleuromutilin suspension at 200 μg / mL against rice streak pathogen. [Figure 6]Therapeutic activity of 20% pleuromutilin suspension at 200 μg / mL against rice streak fungus. [Figure 7] Protective activity of 20% pleuromutilin suspension at 200 μg / mL against Xanthomonas campestris pv. citri. [Figure 8] Therapeutic activity of 20% pleuromutilin suspension at 200 μg / mL against Xanthomonas campestris pv. citri. DETAILED DESCRIPTION OF THE INVENTION

[0022] The present invention will be further explained by the following examples. It should be understood that the methods described in the examples of the present invention are merely for the purpose of illustrating the present invention, and are not intended to limit the present invention. Any simple improvements to the preparation methods of the present invention based on the concept of the present invention are within the scope of the present invention. All raw materials and solvents used in the examples are commercially available products of appropriate purity.

[0023] Example 1: Preparation of 20% Pleuromutilin Suspension 2.0 g of sodium xylenesulfonate HAS040, 2.0 g of naphthalenesulfonic acid sodium salt formaldehyde condensate NNO, 1.5 g of sodium p-toluenesulfonate, 4.0 g of sodium chloride, and 2.0 g of polymeric amphiphilic anionic nonionic surfactant complex SC29 were weighed into a beaker containing 66.1 g of secondary water and stirred to dissolve. 20.0 g of pleuromutilin and 2.0 g of magnesium aluminum silicate were added, and after shearing to homogenize, the mixture was placed in a grinder containing zirconium beads with a suspension volume to mass ratio of 1:1. 0.4 g of polydimethylsiloxane was added, and grinding was started in an ice-water bath. Particle size testing was performed every hour, and after approximately 4 hours, the particle size was tested using a laser particle sizer. When the particle size reached D90 of approximately 6 μm, stirring was stopped and the mixture was filtered to obtain a 20% pleuromutilin suspension.

[0024] Example 2: Preparation of 10% Pleuromutilin Suspension [Table 8] JPEG0007799367000008.jpg57170(1) 2.0 g of sodium xylene sulfonate HAS040, 2.0 g of sodium naphthalene sulfonate formaldehyde condensate NNO, 1.5 g of sodium p-toluenesulfonate, 4.0 g of sodium chloride, and 2.0 g of polymeric amphiphilic anionic nonionic surfactant complex SC29 were weighed into a beaker containing 66.1 g of secondary water, stirred, and dissolved, followed by 10 Add 0.0g of pleuromutilin and 2.0g of magnesium aluminum silicate, and after shear homogenization, place it in a grinder containing zirconium beads with a suspension volume mass ratio of 1:1, add 0.4g of silicone antifoaming agent polydimethylsiloxane, and begin grinding in an ice water bath. Test the particle size every hour, and after about 4 hours, test the particle size with a laser granulometer. When the D90 is about 6μm, stop stirring and obtain a 10% pleuromutilin suspension after filtration.

[0025] Example 3: Primary performance test of pleuromutilin aqueous suspension The key technical indicators of the pleuromutilin suspension of the present invention are measured in accordance with the provisions of China's national or industry standards for pesticides (GB / T14825-2006 Method for determining suspension rate of pesticides, GB / T19136-2003 Method for determining thermal storage stability of pesticides, GB / T19137-2003 Method for determining low-temperature stability of pesticides, HG / T2467.5-2003 Compilation specifications for product standards for pesticide suspensions). As shown in Table 5, when the particle size of the 20% pleuromutilin aqueous suspension prepared using the above additives was greater than 10 μm, the degradation rates of the drug substance after hot storage (54°C±2°C) were relatively high, above 20%. When the particle size of the drug substance in the suspension was between 5 and 8 μm, the degradation rate was approximately 5%-10%, meeting the stability requirements. This demonstrates that under the above formulation, the formulation and particle size have a significant impact on the chemical stability of the drug substance in the pleuromutilin aqueous suspension, and the smaller the particle size, the better the chemical stability of the drug substance. In accordance with the national standard requirements for pesticide suspensions (GB / T19136-2003), the main properties of the suspension were tested, as shown in Table 7. The results showed that with this formulation, the pleuromutilin aqueous suspension exhibited a milky white, uniform dispersion system, with relatively good dispersibility and suspension rate, and the decomposition rate of the active ingredient after heat storage was less than 10%.

[0026] Example 4: In vitro test of pesticide efficacy In this study, the turbidity method was used to evaluate the inhibitory EC of ten species of bacteria, namely, bacterial leaf blight fungus of rice (Xoo), bacterial streak fungus of rice (Xoc), bacterial canker fungus of citrus (Xac), cabbage black rot fungus, peach hole fungus, Chinese cabbage soft rot fungus, muskmelon angular spot fungus, tobacco bacterial wilt fungus, watermelon fruit rot fungus, and tomato bacterial wilt fungus, by using formulations such as pleuromutilin suspension. 50 The values ​​were measured. 1. Test materials: Gun heads (5 mL, 1 mL, 200 μL), 15 mL centrifuge tubes, secondary water, 2 mL EP tubes, and NB medium must all be sterilized. 2. NB medium: 3 g of beef peptone, 5 g of peptone, 1 g of yeast powder, 10 g of glucose, and 1 L of secondary water were dispensed into test tubes (4 mL each), and the pH was adjusted to 7.0-7.2. 3, Operation steps: (1) Weigh out a certain amount of the formulation to be measured and dissolve it in the corresponding volume (200 μL) of DMSO. Prepare a 5% solution, dilute it to the required concentration by the ratio, and make up to 4 mL with 0.1% Tween solution. (2) Using a pipette, aspirate 1 mL of the Tween mixed solution and add it to a test tube containing 4 mL of culture medium. (3) 200 μL of the mixed solution (culture medium) is aspirated using a pipette, and the OD value is measured and recorded in a 96-well plate. (4) Inject 50 μL of bacterial suspension of the pathogens of rice bacterial leaf blight (Xoo), rice bacterial streak (Xoc), citrus canker (Xac), cabbage black rot, peach needle hole, Chinese cabbage soft rot, muskmelon bacterial angular spot, tobacco bacterial wilt, watermelon fruit rot, and tomato bacterial wilt into each test tube. (5) Place in a shaker and set the shaker to 28°C and 180 rpm. (6) After 24 to 36 hours (the OD value of CK is 0.6 to 0.8 (indicating that the bacteria we created are not contaminated and are growing normally)), measure and record the OD value, and calculate the inhibition rate. Calibrated OD value = OD value of bacterial-containing medium - OD value of sterile medium Inhibition rate % = (OD value of bacterial solution in control medium after correction - OD value of bacterial-containing medium after correction) / OD value of bacterial solution in control medium after correction * 100 4. Activity data 20% Pleuromutilin suspension exhibits excellent antibacterial activity against ten types of bacterial diseases, including rice bacterial leaf blight (Xoo), and its EC 50 All values ​​were superior to the comparators 20% thiadiazole zinc suspension, 20% thiadiazole copper suspension, and 95% pleuromutilin drug substance. [Table 9] As can be seen from Table 9, the 20% pleuromutilin suspension and 98% pleuromutilin drug substance of the present invention have an EC 50 The values ​​were 0.79 μg / mL and 1.33 μg / mL, respectively. 50 The activity of this drug was superior to that of the control drugs 20% thiadiazole zinc suspension (13μg / mL), 20% thiadiazole copper suspension (76.27μg / mL), 12% chongshenmycin powder (85.16μg / mL) and 98% pleuromutilin drug substance (1.33μg / mL). [Table 10] As can be seen from Table 10, the 20% pleuromutilin suspension and 98% pleuromutilin drug substance of the present invention have an EC 50 The EC values ​​of the 20% pleuromutilin suspension of the present invention were 0.65 μg / mL and 0.88 μg / mL, respectively. 50 All of the activity values ​​were superior to those of the control drugs 20% thiadiazole zinc suspension (29.11 μg / mL), 20% thiadiazole copper suspension (77.30 μg / mL), 12% chongshenmycin powder (60.75 μg / mL), and 98% pleuromutilin drug substance (1.04 μg / mL). [Table 11] As can be seen from Table 11, the EC of the 20% pleuromutilin suspension of the present invention and the 95% pleuromutilin drug substance against Xanthomonas campestris pv. citri 50 The EC values ​​of the 20% pleuromutilin suspension of the present invention were 1.13 μg / mL and 2.87 μg / mL, respectively. 50 The activity of each compound was superior to that of the control drugs 20% thiadiazole zinc suspension (41.97μg / mL), 20% thiadiazole copper suspension (104.89μg / mL), 12% chongshenmycin powder (108.51μg / mL), and 98% pleuromutilin drug substance (2.87μg / mL). [Table 12] JPEG0007799367000012.jpg42170As can be seen from Table 12, the EC of the 20% pleuromutilin suspension of the present invention and the 98% pleuromutilin drug substance against Black Rot Fungus of Cabbage 50 The values ​​were 11.69 μg / mL and 14.48 μg / mL, respectively. 50 The activity values ​​were all superior to those of the control drugs 20% thiadiazole zinc suspension (55.41μg / mL), 20% thiadiazole copper suspension (82.27μg / mL), 12% chengshengmycin powder (92.61μg / mL) and 98% pleuromutilin drug substance (14.48μg / mL). [Table 13] As can be seen from Table 13, the EC of the 20% pleuromutilin suspension of the present invention and the 98% pleuromutilin drug substance against peach borer fungus was 50 The EC values ​​of the 20% pleuromutilin suspension of the present invention were 16.93 μg / mL and 24.80 μg / mL, respectively. 50 The activity of each compound was superior to that of the control drugs 20% thiadiazole zinc suspension (82.54μg / mL), 20% thiadiazole copper suspension (263.87μg / mL), 12% chongshenmycin powder (146.78μg / mL) and 98% pleuromutilin drug substance (24.80μg / mL). [Table 14] JPEG0007799367000014.jpg42170As can be seen from Table 14, the EC of the 20% pleuromutilin suspension of the present invention and the 98% pleuromutilin drug substance against Chinese cabbage soft rot fungus 50 The values ​​were 18.50 μg / mL and 21.30 μg / mL, respectively. 50 The activity was superior to that of the control drugs 20% thiadiazole zinc suspension (30.56μg / mL), 20% thiadiazole copper suspension (131.32μg / mL), 12% chengshengmycin powder (136.50μg / mL) and 98% pleuromutilin drug substance (28.89μg / mL). [Table 15] JPEG0007799367000015.jpg42170As can be seen from Table 15, the EC of the 20% pleuromutilin suspension of the present invention and the 98% pleuromutilin drug substance against the muskmelon angular spot pathogen 50 The values ​​were 40.43 μg / mL and 52.17 μg / mL, respectively. 50 The activity values ​​were all superior to those of the control drugs 20% thiadiazole zinc suspension (65.41μg / mL), 20% thiadiazole copper suspension (156.40μg / mL), 12% chongshenmycin powder (135.42μg / mL) and 98% pleuromutilin drug substance (52.17μg / mL). [Table 16] As can be seen from Table 16, the EC of the 20% pleuromutilin suspension of the present invention and the 98% pleuromutilin drug substance against tobacco bacterial wilt fungus was 50 The EC values ​​of the 20% pleuromutilin suspension of the present invention were 27.03 μg / mL and 33.31 μg / mL, respectively. 50 The activity was superior to that of the control drugs 20% thiadiazole zinc suspension (50.50μg / mL), 20% thiadiazole copper suspension (101.23μg / mL), 12% chengshengmycin powder (122.95μg / mL) and 98% pleuromutilin drug substance (33.31μg / mL). [Table 17] JPEG0007799367000017.jpg42170As can be seen from Table 17, the EC of the 20% pleuromutilin suspension of the present invention and the 98% pleuromutilin drug substance against watermelon fruit rot fungus 50 The values ​​were 56.50 μg / mL and 60.99 μg / mL, respectively. 50 The activity of these values ​​was superior to that of the control drugs 20% thiadiazole zinc suspension (88.81 μg / mL), 20% thiadiazole copper suspension (97.46 μg / mL), 12% chongshenmycin powder (110.33 μg / mL), and 98% pleuromutilin drug substance (60.99 μg / mL). [Table 18] As can be seen from Table 18, the EC of the 20% pleuromutilin suspension of the present invention and the 98% pleuromutilin drug substance against bacterial wilt of tomato 50 The values ​​were 22.62 μg / mL and 43.29 μg / mL, respectively. 50 The activity of all values ​​was superior to that of the control drugs 20% thiadiazole zinc suspension (88.13 μg / mL), 20% thiadiazole copper suspension (141.25 μg / mL), and 98% pleuromutilin drug substance (43.29 μg / mL).

[0027] Example 5: Efficacy test of potted rice bacterial leaf blight live pesticide This time, the leaf cutting method was used to measure the protective and curative effects of the pleuromutilin formulation of the present invention at a concentration of 200 μg / mL on live potted rice plants infected with bacterial leaf blight. Therapeutic activity: Using scissors dipped in a rice leaf blight fungus solution, a 1 / 3 cm section of the leaf tip was cut off from the rice plant. After 24 hours, a 20% pleuromutilin suspension and a 98% pleuromutilin drug substance of the present invention were selected and dissolved in DMSO. A 200 μg / mL concentration of the drug was then prepared in a 0.1% Tween-80 solution. This drug was sprayed evenly onto the rice leaves until droplets fell. A control was also set up with pure water without the drug. Ten to fifteen leaves were treated for each rice plant. Fourteen days after drug administration, the length of the lesions on the rice leaves was measured, and the preventive and therapeutic effect was calculated based on the length of the lesions. [Number 1] JPEG0007799367000019.jpg20170 Protective activity: A 20% pleuromutilin suspension and a 98% pleuromutilin drug substance of the present invention were selected and dissolved in 200 μL of DMSO. Each was then mixed with a 0.1% Tween-80 solution to a concentration of 200 μg / mL. The mixture was sprayed evenly onto rice leaves until droplets formed. After 24 hours, the leaves were inoculated with the rice bacterial leaf blight pathogen, which was then cut with scissors. The bacterial solution was placed in the apical third of the rice leaf. A control was also prepared using pure water without the agent. Each treatment involved 10-15 leaves, and the treatment was repeated three times. 14 days after the agent administration, the length of the lesions on the rice leaves was measured, and the preventive and therapeutic effects were calculated based on the length of the lesions. [Number 2] JPEG0007799367000020.jpg20170[Table 19] JPEG0007799367000021.jpg43170Table 19 and Figure 2 show that at a concentration of 200 μg / mL, the therapeutic activities of the 20% pleuromutilin suspension of the present invention and the 98% pleuromutilin drug substance are 62.8% and 59.9%, respectively, of which the activity of the 20% pleuromutilin suspension of the present invention (62.8%) is superior to that of the 98% pleuromutilin drug substance (59.9%), the 20% thiazole zinc suspension (56.0%), the 20% thiadiazole copper suspension (44.8%), and the 12% Zhongshenmycin wettable powder. [Table 20] JPEG0007799367000022.jpg43170Table 20 and Figure 3 show that at a concentration of 200 μg / mL, the therapeutic activities of the 20% pleuromutilin suspension of the present invention and the 98% pleuromutilin drug substance are 50.5% and 48.7%, respectively. Among them, the activity of the 20% pleuromutilin suspension of the present invention (50.5%) is superior to that of the 98% pleuromutilin drug substance (48.7%), the 20% thiazole zinc suspension (47.7%), the 20% thiadiazole copper suspension (37.6%), and the 12% zhongshenmycin wettable powder (21.3%).

[0028] Example 6: Pesticide efficacy test on live potted rice streak bacterial disease When the concentration is 200μg / mL, the osmotic pressure method is used to measure the protective effect of 20% pleuromutilin suspension against rice bacterial streak disease in live potted plants. Protective activity: Using an analytical balance, accurately weigh out 20% pleuromutilin suspension, 98% pleuromutilin bulk drug, 10% pleuromutilin wettable powder, and control agent into 1.5mL centrifuge tubes. Add 200μL of DMSO to each and dissolve. Each was mixed with 0.1% Tween-20 solution to a concentration of 200μg / mL. Spray evenly onto rice leaves until droplets formed. After 24 hours, fill a syringe without a needle with the fungal solution and inject it into the rice plants approximately one-third of the way from the leaf tip. Simultaneously, set aside a control of pure water without the fungus. Each test was repeated three times on 10-15 leaves. 14 days after application, measure the length of the lesions on the rice leaves, and calculate the disease index and preventive efficacy. Preventive treatment effect = (lesion length of negative control group - lesion length of treatment group) / lesion length of negative control group × 100% Therapeutic activity: The fungal solution was filled into a needleless syringe and injected into rice plants approximately one-third of the way from the leaf tip. 24 hours later, using an analytical balance, 20% pleuromutilin suspension, 98% pleuromutilin bulk drug, 10% pleuromutilin wettable powder, and control agent were accurately measured into 1.5mL centrifuge tubes. Each was dissolved with 200μL of DMSO. Each was mixed with a 0.1% Tween-20 solution to a concentration of 200μg / mL. The mixture was sprayed evenly onto rice leaves until droplets fell. A control was also set up with untreated water. 10-15 leaves were treated for each treatment, and the treatment was repeated three times. 14 days after treatment, the length of the lesions on the rice leaves was measured, and the disease index and preventive efficacy were calculated. Preventive treatment effect = (lesion length of negative control group - lesion length of treatment group) / lesion length of negative control group × 100% [Table 21] 4, at a concentration of 200 μg / mL, the therapeutic activity of the 20% pleuromutilin suspension of the present invention is 44.1%, of which the activity against rice bacterial streak of the 20% pleuromutilin suspension of the present invention (44.1%) is superior to that of the 98% pleuromutilin drug substance (41.8%), the 20% thiazole zinc suspension (38.5%), the 20% thiadiazole copper suspension (34.4%), and the 12% zhongshenmycin wettable powder (18.7%). [Table 22] 5, at a concentration of 200 μg / mL, the therapeutic activity of the 20% pleuromutilin suspension of the present invention is 45.3%. The activity of the 20% pleuromutilin suspension of the present invention (45.3%) against rice bacterial streak is superior to that of the 98% pleuromutilin drug substance (44.0%), the 20% thiazole zinc suspension (42.5%), the 20% thiadiazole copper suspension (30.0%), and the 12% Zhongshenmycin wettable powder (19.3%).

[0029] Example 7: Citrus canker fungus live potted plant pesticide efficacy test When the concentration is 200 μg / mL, the needle puncture method is used to test the protective and curative effects of the pleuromutilin suspension formulation of the present invention on potted live plants against citrus canker. protective activity Accurately weigh out 18.0 mg of 20% pleuromutilin suspension using an analytical balance, add 200 μL of DMSO, dissolve each, and mix with 0.1% Tween-80 solution to a concentration of 200 μg / mL. Spray evenly onto rice leaves until droplets fall. After 24 hours, inoculate with a disposable syringe containing Xanthomonas campestris pv. citri. Prick 18 small holes per leaf and inject Xac suspension (OD ). 595 A new filter paper soaked in chlorine-free water (=0.6) was applied to the wound for 24 hours. The filter paper containing the drug was discarded, and a control of pure water without any drug was also prepared. 10-15 leaves were treated for each treatment, and the treatment was repeated three times. 21 days after drug administration, the disease status of the citrus leaves was measured, and the preventive treatment was calculated based on the disease index. [Number 3] JPEG0007799367000025.jpg20170 Therapeutic activity The leaves were inoculated with a disposable syringe containing Xanthomonas campestris pv. citri, and 18 small holes were punctured in each leaf, followed by the injection of a Xanthomonas campestris suspension (OD ). 595 After applying a new filter paper soaked in DMSO (=0.6) to the wound surface for 24 hours, accurately measure 18.0 milligrams of 20% pleuromutilin suspension using an analytical balance, dissolve each in DMSO, and then prepare a 200 μg / mL solution with 0.1% Tween-80 solution. This is sprayed evenly onto the citrus leaves until droplets fall. At the same time, a control of pure water without any drug is used. 10 to 15 leaves are treated for each citrus plant. 21 After 3 days, the disease status of the citrus leaves is measured, and the preventive and therapeutic effects are calculated based on the disease index. [Number 4] JPEG0007799367000026.jpg20170[Table 23] JPEG0007799367000027.jpg42170Table 23 and Figure 7 show that at a concentration of 200 μg / mL, the protective activity of the 20% pleuromutilin suspension of the present invention is 48.7%, of which the activity against citrus canker of the 20% pleuromutilin suspension of the present invention (48.7%) is superior to that of the 98% pleuromutilin drug substance (44.9%), the 20% thiazole zinc suspension (45.8%), the 20% thiadiazole copper suspension (39.7%), and the 12% Zhongshenmycin wettable powder (20.1%). [Table 24] JPEG0007799367000028.jpg47170From Table 24 and Figure 8, it can be seen that at a concentration of 200 μg / mL, the protective activity of the 20% pleuromutilin suspension of the present invention is 50.8%, of which the activity against citrus canker of the 20% pleuromutilin suspension of the present invention (50.8%) is superior to that of the 98% pleuromutilin drug substance (47.9%), the 20% thiazole zinc suspension (48.2%), the 20% thiadiazole copper suspension (36.5%), and the 12% zhongshenmycin wettable powder (37.4%).

Claims

1. A suspension of pleuromutilin, comprising: A pleuromutilin suspension comprising 5-25 wt% pleuromutilin, 3-12 wt% wetting and dispersing agent, 0.5-2.5 wt% thickener, 2-6 wt% antifreeze, and 0.3-0.5 wt% silicone antifoaming agent, the balance being deionized water; the particle size of the suspension is 5-8 μm; the wetting and dispersing agent consists of a polymeric amphiphilic anionic nonionic surfactant complex SC29, sodium xylene sulfonate HAS040, sodium naphthalene sulfonate formaldehyde condensate NNO, and sodium p-toluenesulfonate in a weight ratio of 2:2:2:1.5; the thickener is selected from one or both of magnesium aluminum silicate SF-40 and xanthan gum; the antifreeze is selected from sodium chloride; and the silicone antifoaming agent is selected from polydimethylsiloxane.

2. The method for preparing a pleuromutilin suspension according to claim 1, characterized in that the weight percent of wetting and dispersing agent, antifreeze and deionized water are taken, stirred and dissolved, then the weight percent of pleuromutilin and thickener are added, and the mixture is uniformly mixed under the action of a shear machine. The mixed slurry is then pumped into a polishing machine for polishing, and the weight percent of silicone antifoaming agent is added. Particle size tests are conducted every hour, and when D90 becomes smaller than 8 μm, stirring is stopped and the mixture is filtered to obtain a 5-25% pleuromutilin suspension.

3. Use of the suspension according to claim 1 in the field of controlling agricultural pests, 2. The use of the suspension according to claim 1 in controlling agricultural pests, wherein the agricultural pest is at least one of Xanthomonas oryzae (Xoo), Xanthomonas oryzae (Xoc), Xanthomonas citri (Xac), Cabbage black rot fungus, Peach needle hole fungus, Chinese cabbage soft rot fungus, Muskmelon angular spot fungus, Tobacco bacterial wilt fungus, Watermelon fruit rot fungus, and Tomato bacterial wilt fungus.

4. A method for controlling agricultural pests, comprising:

10. A method for controlling agricultural pests, comprising applying the suspension according to claim 1 to harmful substances or their living environment, the harmful substances being at least one of bacterial leaf blight fungus (Xoo), bacterial streak fungus (Xoc), bacterial canker fungus (Xac), cabbage black rot fungus, peach hole fungus, Chinese cabbage soft rot fungus, muskmelon angular spot fungus, tobacco bacterial wilt fungus, watermelon fruit rot fungus, and tomato bacterial wilt fungus.

5. 10. A method for protecting plants from agricultural pests, comprising the step of contacting agricultural pests with the suspension of claim 1, 1. A method for protecting plants from agricultural pests, wherein the agricultural pest is at least one of bacterial leaf blight fungus (Xoo), bacterial rice streak fungus (Xoc), bacterial canker fungus (Xac), cabbage black rot fungus, peach hole fungus, Chinese cabbage soft rot fungus, muskmelon bacterial angular spot fungus, tobacco bacterial wilt fungus, watermelon fruit rot fungus, and tomato bacterial wilt fungus.

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