Fluorine-containing silazane-containing antiseptic paste and method for preparing the same

Abstract

The application provides a kind of fluorosilazole-containing bactericidal ointment and a preparation method thereof, and belongs to the technical field of pesticide bactericidal preparation.The bactericidal ointment of the application uses fluorosilazole and chitosan quaternary ammonium salt as the composite bactericidal active component, and is combined with a hydroxylated modified attapulgite slow-release carrier, a polyvinyl alcohol-carboxymethyl chitosan composite film-forming matrix, a compounded penetration enhancer and a composite thickening and moisturizing aid to construct a stable ointment system;Meanwhile, two novel evaluation formulas of synergistic bacteriostatic gain index SI and film-forming slow-release equivalent coefficient K are created, which accurately limit the synergistic performance and long-acting slow-release and wash-resistant performance of the product, so that the product has stable and excellent comprehensive protection effect.The application solves the technical problems of traditional fluorosilazole ointment, such as drug resistance, weak bactericidal synergism, fast release of active ingredients, poor rainwater wash-resistant ability, poor storage stability and short effective period, by using a vacuum load slow-release process combined with a gradient homogenization emulsification preparation process.

Description

Technical Field

[0001] This invention belongs to the field of pesticide fungicide technology, and more specifically, relates to a fungicide paste containing fluorosilazole and its preparation method. Background Technology

[0002] Flusilazole is a highly effective, broad-spectrum triazole systemic fungicide. It exerts its fungicidal effect by inhibiting ergosterol biosynthesis in pathogens and disrupting the integrity of fungal cell membranes. It exhibits excellent control efficacy against higher fungal diseases such as fruit tree canker, rot, and stem rot, and is widely used in the application of fungal pastes for the control of branch and trunk diseases in forest and fruit trees. Compared to spray formulations, fungicidal pastes can be directly applied to lesions on branches and trunks, offering strong retention and high targeting. They are effective in treating and protecting against hidden and persistent fungal diseases on branches and trunks, making them an important formulation for the green control of fruit tree diseases.

[0003] Currently available flusilazole fungicides still have many inherent technical defects, which seriously limit their effectiveness and service life in the field:

[0004] First, most existing ointments use a single flusilazole active system. Long-term single application can easily induce pathogens to develop drug resistance. The bactericidal spectrum is limited and the efficacy is severely reduced in the later stages. Conventional physical compounding methods have limited synergistic effects, lack quantitative control standards, and the compounding effect is highly random and has poor stability.

[0005] Secondly, traditional ointments mostly use ordinary adjuvant systems and lack slow-release loading structures and wash-resistant film-forming systems. The active ingredients are released quickly and there is a significant burst release phenomenon. Rainwater washes away the pesticide easily, and the field effect period is generally less than 2 to 3 months. Frequent reapplication is required, resulting in high labor costs and poor continuity of control.

[0006] Third, existing dry ointments generally suffer from insufficient stability of the ointment system. Long-term storage can easily lead to defects such as stratification, water separation, sedimentation of active ingredients, and high thermal decomposition rate, resulting in short product shelf life and poor batch stability.

[0007] Fourth, existing technologies lack a quantitative evaluation system for the "synergistic bactericidal ability and sustained-release erosion resistance" of ointments. Relying solely on single indicators such as conventional antibacterial rate and release time to evaluate product performance cannot accurately characterize the comprehensive protective efficacy of the formulation. Formula optimization lacks scientific basis, and there are bottlenecks in improving product performance.

[0008] In existing technologies, conventional flusilazole compound formulations are mostly simple physical mixtures, lacking suitable sustained-release loading and composite film-forming systems for trunk and branch coating scenarios. Furthermore, they lack dedicated synergistic quantification models and sustained-release performance evaluation models, resulting in a series of technical shortcomings such as insignificant synergistic effects, short duration of action, poor weather resistance, weak stability, and high risk of drug resistance. Therefore, developing a flusilazole-containing bactericidal ointment with significant synergistic antibacterial effects, long-lasting sustained release, resistance to rain washout, excellent storage stability, and quantifiable controllable product performance, along with its preparation method, has become an urgent technical problem to be solved in this field. Summary of the Invention

[0009] To address the technical problems of existing flusilazole-based dry bactericidal preparations, such as poor film-forming and wash-resistant properties, excessively rapid release rate of active ingredients, short duration of effect, easy induction of pathogen resistance by single agents, insignificant synergistic effect of compound formulations, and easy separation of layers and water separation in ointment form with poor storage stability, this invention provides a flusilazole-containing bactericidal ointment and its preparation method to solve the problems of insufficient long-term protective ability, weak synergistic effect of compound formulations, and poor stability of ointments in existing flusilazole preparations.

[0010] A bactericidal ointment containing flusilazole, comprising the following components by weight:

[0011] 5-12 parts of bactericidal active ingredient, 8-15 parts of loaded slow-release carrier, 10-20 parts of composite film-forming matrix, 2-5 parts of penetration enhancer, 1-3 parts of thickener, 3-8 parts of moisturizer, and deionized water to make up to 100 parts.

[0012] The bactericidal active component is composed of flusilazole and chitosan quaternary ammonium salt in a weight ratio of (4~7):1.

[0013] Preferably, the synergistic antifungal gain index SI of the fungicidal ointment against plant pathogenic fungi satisfies the following relationship:

[0014] In the formula EC 500 EC represents the half-maximal effective concentration (MCP) of flusilazole alone. 50m The concentration represents the half-maximal effective concentration after compounding, and W1 and W2 are the mass percentages of flusilazole and chitosan quaternary ammonium salt in the bactericidal active component, respectively.

[0015] Preferably, the loaded slow-release carrier is hydroxylated modified attapulgite with a particle size of 800-1200 mesh;

[0016] The film-forming sustained-release equivalent coefficient K of the bactericidal ointment satisfies the following relationship:

[0017] In the formula t 0.5The time it takes for the cumulative release rate of flusilazole to reach 50% is given by η, F is the erosion resistance retention rate of the membrane layer, M is the total proportion of flusilazole, and η is the cone penetration of the paste.

[0018] Preferably, the composite film-forming matrix is ​​composed of polyvinyl alcohol with a degree of hydrolysis of 87%~89% and carboxymethyl chitosan in a weight ratio of (3~5):1.

[0019] Preferably, the penetration enhancer is composed of laurocapram and propylene glycol in a weight ratio of 1:(2~4).

[0020] Preferably, the thickener is a mixture of xanthan gum and magnesium aluminum silicate in a weight ratio of 1:2, and the humectant is a mixture of glycerin and sorbitol in a weight ratio of 1:1.

[0021] Another technical problem to be solved by the present invention is to provide a method for preparing the aforementioned fluorosilazole-containing bactericidal ointment, comprising the following steps:

[0022] Preparation of S1 supported fluorosilazole micro powder: Fluorosilazole was dissolved in anhydrous ethanol, and the loaded slow-release carrier was added and mixed thoroughly. The mixture was impregnated and stirred for 2-3 hours under vacuum of -0.06~-0.08MPa and temperature of 45~55℃. After restoring to normal pressure, the mixture was aged for 1-2 hours, filtered, dried, ground and sieved to obtain supported fluorosilazole micro powder.

[0023] Preparation of S2 composite film-forming solution: Add the composite film-forming matrix to 60%~70% of the total amount of deionized water, heat to 65~75℃, keep warm and stir until completely dissolved to obtain a homogeneous film-forming solution.

[0024] S3 premixed slurry preparation: Chitosan quaternary ammonium salt, penetration promoter, and humectant are dissolved in the remaining deionized water, and then supported fluorosilazole micro powder is added and dispersed at high speed for 15~25 min to obtain a uniform premixed slurry;

[0025] S4 Homogenization to form paste: Add the film-forming adhesive to the premixed slurry and stir evenly. After adding the thickener, homogenize and emulsify at 50~60℃ for 20~30min. After cooling to room temperature, adjust the pH of the system to 5.5~6.5 to obtain the bactericidal paste.

[0026] Compared with the prior art, the present invention has the following beneficial effects:

[0027] In this invention, by directional compounding of flusilazole and chitosan quaternary ammonium salt, combined with precise control of the ratio using an original synergistic antibacterial gain index, a significant synergistic antibacterial effect is achieved, reducing the risk of pathogen resistance.

[0028] Modified attapulgite clay was used as a load carrier, and a dual system of "pore adsorption sustained release + membrane barrier controlled release" was constructed in combination with a composite film-forming matrix. The original film-forming sustained release equivalent coefficient was used to quantify the long-term protective ability and rain erosion resistance of the formulation.

[0029] The optimized adjuvant system improves the storage stability and coating application of the paste, while the matching vacuum loading-gradient homogenization process ensures the uniform dispersion of active ingredients and the stability of formulation quality. The overall solution has outstanding innovation and practical value in the prevention and control of tree branch diseases in fruit trees. Attached Figure Description

[0030] Figure 1 This is a flowchart illustrating the preparation process of the fluorosilazole-containing bactericidal ointment of the present invention. Figure 2 This is a schematic diagram of the microstructure of supported fluorosilazole micropowder. Figure 3 This is a schematic diagram of the dual-barrier sustained-release mechanism after the ointment is coated and formed into a film. Figure 4 A comparative bar chart showing the synergistic antibacterial gain index (SI) of each sample. Figure 5 The curves showing the cumulative release rate of fluorosilazole from each sample over time are shown. Figure 6 A comparison chart of the erosion resistance retention rates of the film layers for each sample. Figure 7 A comparison chart of the decomposition rates of thermal storage and freeze-thaw stability for each sample. Figure 8 A comparison chart showing the healing rate of lesions in the field 6 months after each sample was treated. Detailed Implementation

[0031] To make the technical solution, purpose, and beneficial effects of this invention clearer, the invention will be further described in detail below with reference to specific embodiments, comparative examples, and performance verification tests. It should be understood that the specific embodiments described herein are only for explaining the invention and are not intended to limit the scope of protection of the invention. All other embodiments obtained by those skilled in the art without creative effort based on the core concept of this invention are within the scope of protection of this invention.

[0032] I. Experimental Materials and Equipment:

[0033] (I) Main experimental materials:

[0034] Flusilazole technical grade: 95% active ingredient by mass, industrial grade, meeting the standards for pesticide technical grade products;

[0035] Chitosan quaternary ammonium salt (2-hydroxypropyltrimethylammonium chloride chitosan): Deacetylation degree ≥90%, cationic substitution degree 92%, weight average molecular weight 80,000, industrial grade;

[0036] Hydroxylated modified attapulgite (self-made): Natural attapulgite ore was crushed and passed through a 200-mesh sieve. A 30% hydrogen peroxide solution was added at a solid-liquid mass ratio of 1:10, and the mixture was stirred at 80℃ for 4 hours. After the reaction, the mixture was filtered and washed with deionized water until the filtrate was neutral. The filter cake was dried at 105℃ to constant weight and then crushed and classified by an air jet mill to obtain the product with the target particle size. The surface hydroxyl content was determined to be ≥1.2 mmol / g and the specific surface area was ≥180 m² / g by Boehm titration.

[0037] Polyvinyl alcohol: Model PVA1788, degree of alcoholysis 87%~89%, degree of polymerization 1700, industrial grade;

[0038] Carboxymethyl chitosan: Degree of deacetylation ≥85%, degree of carboxymethyl substitution 0.85, industrial grade;

[0039] Lauryl diazepine: pharmaceutical grade, content ≥99%;

[0040] Propylene glycol, glycerin, sorbitol: pharmaceutical grade;

[0041] Xanthan gum: food grade, molecular weight ≥ 2 million;

[0042] Magnesium aluminum silicate: pharmaceutical grade, colloidal grade;

[0043] Anhydrous ethanol, citric acid, triethanolamine: analytical grade, commercially available.

[0044] (II) Major Instruments and Equipment:

[0045] Vacuum reactor, planetary ball mill, high-speed disperser, high-shear homogenizer and emulsifier, cone penetration tester (compliant with GB / T269 standard), high performance liquid chromatograph (equipped with ultraviolet detector), constant temperature incubator, artificial rain erosion test device, dialysis bags (molecular weight cutoff 8000~14000Da), constant temperature shaker, pH meter.

[0046] II. Performance Parameters and Calculation Methods:

[0047] (a) Synergistic antibacterial gain index SI:

[0048] This invention defines a synergistic antibacterial gain index (SI) to quantify the synergistic antibacterial efficacy of flusilazole combined with chitosan quaternary ammonium salt. The calculation formula is as follows:

[0049] ;

[0050] Parameter substitution rules and unit explanations:

[0051] EC 500 The half-maximal effective concentration (MDC) of flusilazole when used alone against the target pathogen is expressed in mg / L. This concentration value should be directly substituted into the calculation.

[0052] EC 50m The half-maximal effective concentration (MCP) of the compound bactericidal active components of this invention when acting on the same target pathogens is expressed in mg / L. The concentration values ​​can be directly substituted into the calculation.

[0053] W1: The mass percentage of flusilazole in the bactericidal active ingredient, substituted in decimal form (e.g., if the mass percentage is 83.3%, then substitute 0.833).

[0054] W2: The mass percentage of chitosan quaternary ammonium salt in the bactericidal active component, substituted in decimal form (e.g., if the mass percentage is 16.7%, then substitute 0.167).

[0055] The index term 0.25 is the ratio weighting coefficient, used to weaken the excessive interference of the ratio on the index, so that the index core reflects the increase in toxicity. Judgment criteria: SI > 1.2 indicates a synergistic effect, and SI ≥ 1.8 indicates a significant synergistic effect.

[0056] (ii) Film-forming sustained-release equivalent coefficient K:

[0057] This invention defines a film-forming sustained-release equivalent coefficient K to comprehensively characterize the long-lasting protective capability of bactericidal ointments, coupling four dimensions: sustained-release rate, erosion resistance, drug loading, and ointment consistency. The calculation formula is as follows:

[0058] ;

[0059] Parameter substitution rules and unit explanations:

[0060] t 0.5 : The time required for the cumulative release rate of flusilazole to reach 50% after the ointment forms a film under constant temperature and static water release conditions at 25℃, in hours. Directly substitute the time value into the calculation.

[0061] F: Film mass retention rate after 12 hours of simulated rain rinsing, in percentage. Directly substitute the percentage value into the calculation (e.g., if the retention rate is 92.3%, substitute 92.3).

[0062] M: The mass percentage of the active ingredient flusilazole in the total mass of the ointment, expressed as a percentage. Directly substitute the percentage value into the calculation (e.g., if the percentage is 6.67%, then substitute 6.67).

[0063] η: The penetration of the paste at 25℃, in mm. When calculating, divide the penetration gauge reading (unit: 0.1mm) by 10 and substitute it into the value (e.g., if the reading is 215, substitute it into 21.5).

[0064] Physical meaning of the formula: The comprehensive protective efficacy of the membrane layer under unit drug loading and unit consistency, considering both drug release duration and erosion resistance. A higher K value indicates superior long-term protective performance. Judgment criterion: K ≥ 120 indicates excellent long-term effectiveness and erosion resistance.

[0065] (III) Complete calculation example (taking Example 1 as an example):

[0066] SI value calculation and actual measured EC50 of flusilazole single agent against apple tree rot pathogen 500 =12.85mg / L;

[0067] Example 1 EC of compound components 50m =4.92mg / L; W1=5 / 6≈0.833, W2=1 / 6≈0.167;

[0068] Substituting into the formula: SI = (12.85 / 4.92) × (1 + 0.833 / 0.167) 0.25 ≈2.61×6 0.25 ≈2.61×1.565≈2.12;

[0069] K-value calculation, measured in Example 1, t 0.5 =216h, F=92.3%, M=6.67%, cone penetration reading 215 (0.1mm), i.e. η=21.5mm; Substituting into the formula: K=(216×92.3) / (6.67×21.5)=19936.8 / 143.405≈139.0.

[0070] III. Implementation Examples:

[0071] Example 1:

[0072] The fluorosilazole-containing bactericidal ointment of this embodiment comprises the following components by weight: 8 parts bactericidal active ingredient, 10 parts hydroxylated modified attapulgite, 15 parts composite film-forming matrix, 3 parts penetration enhancer, 2 parts thickener, 5 parts humectant, and deionized water to bring the total to 100 parts. The bactericidal active ingredient is composed of fluorosilazole and chitosan quaternary ammonium salt in a weight ratio of 5:1.

[0073] The composite film-forming matrix is ​​composed of PVA1788 and carboxymethyl chitosan in a weight ratio of 4:1; the penetration enhancer is composed of laurocapram and propylene glycol in a weight ratio of 1:3; the thickener is composed of xanthan gum and magnesium aluminum silicate in a weight ratio of 1:2; and the humectant is composed of glycerin and sorbitol in a weight ratio of 1:1.

[0074] The preparation method steps are as follows:

[0075] Preparation of S1 supported flusilazole micro powder: Weigh the flusilazole technical material according to the formula, dissolve it in anhydrous ethanol at a material-to-liquid ratio of 1g:9mL, and stir until completely dissolved; add hydroxylated modified attapulgite (1000 mesh), stir thoroughly, and then transfer to a vacuum reactor. Control the vacuum degree to -0.07MPa and the system temperature to 50℃, and impregnate and stir at 200r / min for 2.5h.

[0076] After shutting off the vacuum system and restoring normal pressure, let it stand at 25°C for 1.5 hours to age.

[0077] The filter cake was collected by vacuum filtration using a 0.45 μm organic filter membrane and placed in a forced-air drying oven at 55℃ and an air velocity of 0.5 m / s until the moisture content was ≤2%.

[0078] The dried product was transferred into a planetary ball mill, using zirconia balls as the grinding medium at a ball-to-material ratio of 3:1, and ground at 300 r / min for 30 min. The product was then passed through a 1000-mesh standard sieve to obtain supported fluorosilazole micro powder.

[0079] Preparation of S2 composite film-forming solution: Add each component of the composite film-forming matrix to 65% of deionized water, heat to 70℃, stir at 350r / min for 40min until completely dissolved and no visible colloids are obtained, thus obtaining a uniform film-forming solution.

[0080] S3 premixed slurry preparation: Add chitosan quaternary ammonium salt, penetration enhancer and humectant to the remaining deionized water and stir until completely dissolved;

[0081] Then add the supported fluorosilazole micro powder obtained in step S1, control the system temperature at 35℃, and disperse at a high speed of 2500r / min for 20min to obtain a uniform premixed slurry.

[0082] S4 Homogenization to form paste: Slowly add the film-forming adhesive to the premixed slurry, stir at low speed until uniform, then add the thickener; transfer to a high-shear homogenizing emulsifier, first stir at 400 r / min for 10 min to allow the thickener to fully swell, then heat to 55℃, and homogenize and emulsify at 1000 r / min for 25 min; after cooling to room temperature, slowly add a 10% (w / w) citric acid solution, monitor the pH value in real time, and adjust the pH of the system to 6.0 to obtain the bactericidal paste.

[0083] Example 2:

[0084] The fluorosilazole-containing bactericidal ointment of this embodiment has the following components by weight:

[0085] Five parts of bactericidal active ingredient, eight parts of hydroxylated modified attapulgite, ten parts of composite film-forming matrix, two parts of penetration enhancer, one part of thickener, three parts of humectant, and deionized water to make up to 100 parts.

[0086] The bactericidal active component is composed of flusilazole and chitosan quaternary ammonium salt in a weight ratio of 4:1.

[0087] The composite film-forming matrix is ​​composed of PVA1788 and carboxymethyl chitosan in a weight ratio of 3:1.

[0088] The penetration enhancer is composed of laurocapram and propylene glycol in a weight ratio of 1:2; the composition of the other additives is the same as in Example 1.

[0089] The carrier particle size is 800 mesh.

[0090] The preparation method steps are as follows:

[0091] S1 material-to-liquid ratio 1g:8mL, vacuum degree -0.06MPa, temperature 45℃, soaking and stirring for 2h, aging at 25℃ for 1h, drying at 50℃, and grinding through an 800-mesh sieve;

[0092] S2 deionized water accounts for 60% of the total volume. Heat to 65℃ and stir at 300r / min for 30min.

[0093] S3 high-speed dispersion speed 2000r / min, time 15min, system temperature 30℃;

[0094] S4 is first stirred at 300 r / min for 10 min to swell, then homogenized and emulsified at 50℃ and 800 r / min for 20 min;

[0095] Adjust the pH to 5.5 with a 10% citric acid solution. The remaining procedures are the same as in Example 1.

[0096] Example 3:

[0097] The fluorosilazole-containing bactericidal ointment of this embodiment has the following components by weight:

[0098] 12 parts of bactericidal active ingredient, 15 parts of hydroxylated modified attapulgite, 20 parts of composite film-forming matrix, 5 parts of penetration enhancer, 3 parts of thickener, 8 parts of humectant, and deionized water to make up to 100 parts.

[0099] The bactericidal active component is composed of flusilazole and chitosan quaternary ammonium salt in a weight ratio of 7:1; the composite film-forming matrix is ​​composed of PVA1788 and carboxymethyl chitosan in a weight ratio of 5:1; the penetration promoter is composed of laurocapram and propylene glycol in a weight ratio of 1:4; the composition of the other additives is the same as in Example 1; the carrier particle size is 1200 mesh.

[0100] The preparation method steps are as follows:

[0101] S1 material-to-liquid ratio 1g:12mL, vacuum degree -0.08MPa, temperature 55℃, soaking and stirring for 3h, aging at 25℃ for 2h, drying at 60℃, and grinding through a 1200 mesh sieve;

[0102] S2 deionized water accounts for 70% of the total volume. Heat to 75℃ and stir at 400r / min for 45min.

[0103] S3 high-speed dispersion speed 3000r / min, time 25min, system temperature 40℃;

[0104] S4 is first stirred at 500 rpm for 10 minutes to swell, then homogenized and emulsified at 60°C and 1200 rpm for 30 minutes; the pH is adjusted to 6.5 with 10% citric acid solution. The remaining operations are the same as in Example 1.

[0105] Example 4 (pH boundary and regulator verification):

[0106] The formulation in this embodiment is completely identical to that in Example 1, except that: in step S4, a 10% (w / w) triethanolamine solution is used to adjust the pH of the system to 5.5. The rest of the preparation operations are the same as in Example 1, which is used to verify the feasibility of the lower limit of the pH range and the alkaline regulator in the claims.

[0107] Example 5 (Verification of the boundary of the additive ratio):

[0108] The only difference between the formulation in this embodiment and that in Example 1 is that the penetration enhancer is composed of laurocapram and propylene glycol in a weight ratio of 1:4, and the composite film-forming matrix is ​​composed of a weight ratio of 5:1; the remaining components and preparation operations are the same as in Example 1, and are used to verify the performance at the end of the range of auxiliary agent ratios in the claims.

[0109] IV. Comparative Examples:

[0110] Comparative Example 1 (Single-component active ingredient control):

[0111] The only difference from Example 1 is that the bactericidal active component is a single fluorosilazole, the effective ingredient dosage is 8 parts (the same as the total active component mass in Example 1), and no chitosan quaternary ammonium salt is added; the dosage of other components and the preparation steps are completely consistent with Example 1.

[0112] Comparative Example 2 (Control without Loaded Sustained-Release Carrier):

[0113] The only difference from Example 1 is that no hydroxylated modified attapulgite is added, and the flusilazole technical material is directly dispersed at high speed together with other adjuvants in step S3; the dosage of other components and preparation steps are completely consistent with Example 1.

[0114] Comparative Example 3 (Single Film-Forming Matrix Control):

[0115] The only difference from Example 1 is that the composite film-forming matrix is ​​replaced with a single PVA1788, the total amount is still 15 parts, and no carboxymethyl chitosan is added; the other components and preparation steps are completely consistent with Example 1.

[0116] Comparative Example 4 (Comparison with similar commercially available products):

[0117] Commercially available conventional flusilazole dry bactericidal ointment was selected as a prior art control. Its main components are: 5 parts of flusilazole active ingredient, 2 parts of xanthan gum, 10 parts of petrolatum, and organic solvents and additives to make up to 100 parts.

[0118] Comparative Example 5 (Unmodified attapulgite soil control):

[0119] The only difference from Example 1 is that unmodified ordinary attapulgite (1000 mesh) is used instead of hydroxylated modified attapulgite. All other components and preparation steps are completely consistent with Example 1.

[0120] Comparative Example 6 (Comparison of process without vacuum loading):

[0121] The only difference from Example 1 is that the vacuum impregnation loading process in step S1 is omitted, and the flusilazole technical material is directly added to the system after physical mixing with attapulgite; the remaining components and preparation steps are completely consistent with Example 1.

[0122] V. Performance Testing Methods:

[0123] 1. Determination of antibacterial activity and SI value:

[0124] The target pathogen was *Valsamali*, the causal agent of apple tree rot. The mycelial growth rate method was used for determination.

[0125] The test sample was serially diluted with sterile water to five concentration gradients, and mixed with PDA medium that had been melted and cooled to 50°C at a volume ratio of 1:9. The mixture was then poured into plates to prepare drug-containing medium. Each concentration was replicated three times, with PDA plates containing an equal amount of sterile water serving as blank controls.

[0126] Mycelial cakes (7 days old, with uniform edge growth) with a diameter of 5 mm were punched using a puncher. The mycelial side was placed face down in the center of a drug-containing plate and incubated in a 25℃ incubator in the dark for 72 hours. The diameter of colonies on each plate was measured using the cross-cross method, and the mycelial growth inhibition rate was calculated. The EC50 was calculated using the probability unit regression method in SPSS software. 50 The SI index is calculated using the aforementioned formula.

[0127] 2. Sustained-release performance and t 0.5 Measurement:

[0128] Static dialysis method:

[0129] Accurately weigh 0.5g of the ointment to be tested, and evenly coat it on a clean glass plate of 10cm×10cm with a coating thickness of 0.2mm. Allow it to air dry at room temperature to form a film.

[0130] The glass plate was transferred to a stoppered conical flask containing 200 mL of pH 7.0 phosphate buffer and placed in a 25°C shaker at 100 rpm. 2 mL samples were taken at each of the set time points, and an equal volume of buffer solution was added simultaneously. The flusilazole content in the solution was determined by high-performance liquid chromatography (HPLC). Chromatographic conditions: C18 column (250 mm × 4.6 mm, 5 μm), mobile phase: methanol.

[0131] Water ratio 75:25 (V / V), flow rate 1.0 mL / min, detection wavelength 280 nm, column temperature 30 °C, injection volume 20 μL. The cumulative release rate was calculated, and the time t for the cumulative release rate to reach 50% was obtained by fitting the release curve. 0.5 .

[0132] 3. Determination of membrane erosion resistance and F-value:

[0133] A self-made simulated rainwater rinsing device was used. The test paste was evenly coated onto a 150mm × 70mm simulated bark specimen (with the same roughness as apple bark), at a rate of 0.1 g / cm². After drying at room temperature to form a film, the initial mass was measured. The specimen was fixed at a 45° angle below the rinsing device, with a nozzle height of 30cm, water pressure of 0.05MPa, and a spray flow rate of 0.1L / min. Rinsing was performed continuously for 12 hours. After rinsing, the specimen was removed and dried at 60℃ to constant weight. The remaining film mass was measured, and the film mass retention rate F was calculated.

[0134] 4. Cone penetration η measurement:

[0135] Referring to GB / T269-1991 "Determination of Cone Penetration of Lubricating Grease and Petroleum Grease", a full-size standard cone was used, and the cone penetration value was measured under constant temperature conditions of 25℃. The value was read after 5 seconds, and the unit was 0.1 mm.

[0136] 5. Storage stability test:

[0137] Thermal storage stability: The ointment to be tested was sealed in a glass container and stored in a constant temperature chamber at 54℃±2℃ for 14 days. After being taken out and restored to room temperature, the appearance was observed to see if there was stratification, water separation, or clumping. The content of flusilazole before and after thermal storage was determined, and the decomposition rate of the active ingredient was calculated.

[0138] Freeze-thaw stability: The sample was frozen at -10℃ for 18 hours and then thawed at 25℃ for 6 hours as one cycle. A total of 5 cycles were performed, and the appearance changes were observed and the decomposition rate of the active ingredients was measured.

[0139] 6. Field efficacy determination:

[0140] The experiment was conducted in a Fuji apple orchard with 10-year-old trees that had a history of severe canker disease. Canker lesions of uniform severity were selected, and the diseased bark was scraped down to healthy tissue. The test paste was then evenly applied to the lesions and a 2cm layer of healthy bark around them, with a thickness of approximately 1mm. Each treatment was replicated with 3 trees and 5 lesions per tree. A water treatment served as a blank control. The healing of the lesions was assessed at 3 and 6 months after application, and the healing rate was calculated.

[0141] VI. Test Results and Analysis:

[0142] The core performance test results of each embodiment and comparative example are shown in the table below:

[0143] Table 1 Comparison of core performance parameters of each sample:

[0144] Example 1 2.12 216 92.3 6.67 215 139.0 1.82 2.35 89.2 Example 2 1.95 140 88.7 4.00 248 125.2 2.15 2.78 83.5 Example 3 2.31 350 95.1 10.5 196 161.7 1.57 2.01 93.7 Example 4 2.09 212 91.8 6.67 218 136.1 1.90 2.42 88.5 Example 5 2.10 209 91.5 6.67 217 134.7 1.87 2.39 88.1 Comparative Example 1 1.00 200 91.5 8.00 212 107.9 2.03 2.64 71.4 Comparative Example 2 2.07 72 76.4 6.67 227 36.3 3.68 4.52 52.8 Comparative Example 3 2.09 154 72.1 6.67 208 80.1 2.41 3.10 65.3 Comparative Example 4 0.98 48 53.2 5.00 260 19.6 4.25 5.87 41.6 Comparative Example 5 2.05 128 85.6 6.67 221 74.2 2.76 3.41 70.9 Comparative Example 6 2.06 112 83.7 6.67 224 63.0 3.02 3.75 67.2

[0145] Synergistic antibacterial effect: The SI values ​​of Examples 1-5 were all ≥1.8, achieving a significant synergistic effect, far exceeding the single-agent control (Comparative Example 1) and commercially available products (Comparative Example 4). This demonstrates that when flusilazole and chitosan quaternary ammonium salt are combined according to the ratio of this invention, the antibacterial activity is significantly enhanced through the synergistic effect of different target sites, reducing the dosage of active ingredient and delaying the development of drug resistance in pathogens. The SI values ​​of Examples 4 and 5 are close to those of Example 1, proving that the synergistic effect is stable within the ratio and pH range specified in the claims.

[0146] Sustained-release and erosion resistance: The K values ​​of Examples 1-5 are all ≥120, significantly higher than all comparative examples. In Comparative Example 2, without a carrier, the release rate is significantly accelerated, and the K value drops sharply; the erosion resistance of the single film-forming matrix in Comparative Example 3 is significantly reduced; Comparative Examples 5 and 6 verify the key roles of hydroxylation modification and vacuum loading processes. Unmodified carriers cannot firmly load drugs through hydrogen bonds, and without vacuum impregnation, drugs cannot easily enter the carrier pores, both of which lead to a significant decrease in sustained-release capacity. This demonstrates that the dual system of "hydroxylated carrier pore adsorption + composite film-forming barrier controlled release" of this invention is the core to achieving long-lasting efficacy, and its effect is significantly better than conventional techniques.

[0147] Storage stability: The thermal storage and freeze-thaw decomposition rates of the examples were both below 3%, with no stratification, water separation, or clumping. The stability was significantly better than the comparative examples lacking a carrier or film-forming system. This demonstrates that the composite thickening system and loading structure of the present invention effectively improve the suspension stability and physical stability of the paste.

[0148] Field efficacy: In the examples, the lesion healing rate after 6 months of application all exceeded 83%, which was much higher than that of the control groups and consistent with the results of the indoor performance test. This verifies the long-term therapeutic and protective effect of the ointment in the natural field environment and has practical application value.

[0149] Mechanism of action:

[0150] Synergistic antibacterial mechanism: Flusilazole exerts its bactericidal effect by inhibiting ergosterol biosynthesis in pathogens and disrupting cell membrane structure; chitosan quaternary ammonium salt binds to the negatively charged groups of the pathogen cell wall through its cationic groups, disrupting cell membrane permeability and coagulating intracellular substances. Since the two drugs target different points, their combination achieves multi-target synergistic attack, enhancing the antibacterial effect while reducing the selectivity pressure of single agents and delaying the development of drug resistance in pathogens.

[0151] Sustained-release and erosion-resistant mechanism: The abundant hydroxyl groups on the surface of hydroxylated modified attapulgite can form hydrogen bonds with flusilazole molecules, loading the drug into the carrier channels and achieving slow release through diffusion through the channels; The composite film-forming system formed by polyvinyl alcohol and carboxymethyl chitosan can form a dense and flexible three-dimensional network film on the bark surface, which not only acts as a physical barrier to prevent rainwater erosion, but also regulates drug release through the swelling rate of the film, achieving dual controlled release of "carrier adsorption + film barrier" and significantly prolonging the duration of effect.

[0152] The stabilization mechanism of the paste: Xanthan gum and magnesium aluminum silicate are combined to form a complex thickening system, which builds a thixotropic network structure, improves the suspension stability of the paste, and avoids the sedimentation and stratification of the active ingredients; the moisturizing system of glycerin and sorbitol maintains the moisture balance of the paste, prevents drying and water separation during storage, and improves the storage stability and application spreadability of the product.

[0153] The embodiments of the present invention are given for illustrative and descriptive purposes only, and are not intended to be exhaustive or to limit the invention to the forms disclosed. Many modifications and variations will be apparent to those skilled in the art. The embodiments were chosen and described in order to better illustrate the principles and practical application of the invention, and to enable those skilled in the art to understand the invention and to design various embodiments with various modifications suitable for a particular purpose.

Claims

1. A bactericidal ointment containing flusilazole, characterized in that, By weight, it includes the following components: 5-12 parts of bactericidal active ingredient, 8-15 parts of loaded slow-release carrier, 10-20 parts of composite film-forming matrix, 2-5 parts of penetration enhancer, 1-3 parts of thickener, 3-8 parts of moisturizer, and deionized water to make up to 100 parts. The bactericidal active component is composed of flusilazole and chitosan quaternary ammonium salt in a weight ratio of (4~7):1; The synergistic antifungal gain index SI of the bactericidal ointment against plant pathogenic fungi satisfies the following relationship: In the formula, EC 500 The half-maximal effective concentration (MDF) of flusilazole when used alone against the target plant pathogenic fungus is given in mg / L. EC 50m The half-maximal effective concentration (MCP) of the fungicidal active components when they are combined and act on the same target plant pathogenic fungi, expressed in mg / L. W1 represents the mass percentage of flusilazole in the bactericidal active ingredient; W2 represents the mass percentage of chitosan quaternary ammonium salt in the bactericidal active component.

2. The bactericidal ointment containing fluorosilazole according to claim 1, characterized in that, The loaded slow-release carrier is hydroxylated modified attapulgite with a particle size of 800~1200 mesh; The film-forming sustained-release equivalent coefficient K of the bactericidal ointment satisfies the following relationship: In the formula, t 0.5 The time required for the cumulative release rate of flusilazole to reach 50% after the ointment forms a film under constant temperature static water immersion conditions at 25℃, expressed in hours; F represents the film mass retention rate after 12 hours of simulated rainwater washing following film formation, expressed in % (%). M represents the percentage of flusilazole in the total mass of the ointment; η is the cone penetration of the paste at 25℃, in units of 0.1 mm.

3. The fluorosilazole-containing bactericidal ointment according to claim 1, characterized in that, The composite film-forming matrix is ​​composed of polyvinyl alcohol with a degree of alcoholysis of 87%~89% and carboxymethyl chitosan in a weight ratio of (3~5):

1.

4. The fluorosilazole-containing bactericidal ointment according to claim 1, characterized in that, The penetration enhancer is composed of laurocapram and propylene glycol in a weight ratio of 1:(2~4).

5. The fluorosilazole-containing bactericidal ointment according to claim 1, characterized in that, The thickener is a mixture of xanthan gum and magnesium aluminum silicate in a weight ratio of 1:

2. The moisturizer is a mixture of glycerin and sorbitol in a weight ratio of 1:

1.

6. A method for preparing a bactericidal ointment containing fluorosilazole as described in any one of claims 1 to 5, characterized in that, Includes the following steps: Preparation of S1 supported flusilazole micro powder: Flusilazole was dissolved in anhydrous ethanol, and the supported slow-release carrier was added and mixed thoroughly. The mixture was impregnated and stirred for 2-3 hours under vacuum of -0.06 to -0.08 MPa and temperature of 45 to 55℃. After restoring to normal pressure, the mixture was aged for 1-2 hours. The filter cake was dried at 50 to 60℃ until the moisture content was ≤2%. The mixture was then ground and sieved to obtain supported flusilazole micro powder. Preparation of S2 composite film-forming solution: Add each component of the composite film-forming matrix to 60%~70% of the total amount of deionized water, heat to 65~75℃, keep warm and stir until completely dissolved to obtain a homogeneous film-forming solution. S3 premixed slurry preparation: Chitosan quaternary ammonium salt, penetration enhancer, and humectant are added to the remaining deionized water and stirred until completely dissolved; then the supported fluorosilazole micro powder is added and dispersed at high speed at 2000~3000 r / min for 15~25 min to obtain a uniform premixed slurry; S4 Homogenization to form paste: Add the film-forming adhesive to the premixed slurry, stir evenly at low speed, add thickener, transfer to homogenizer, and homogenize and emulsify at 800-1200 r / min for 20-30 min at 50-60℃; after cooling to room temperature, adjust the pH value of the system to 5.5-6.5 to obtain the bactericidal paste.

7. The preparation method according to claim 6, characterized in that, In step S1, the ratio of flusilazole to anhydrous ethanol is 1 g: (8~12) mL; after grinding, it is passed through an 800-mesh sieve to obtain supported flusilazole micro powder.

8. The preparation method according to claim 6, characterized in that, In step S2, the stirring speed is 300~400 r / min and the stirring time is 30~45 min, until there are no undissolved clumps visible to the naked eye in the glue solution.

9. The preparation method according to claim 6, characterized in that, In step S3, the system temperature is controlled at 30~40℃ during the high-speed dispersion process.

10. The preparation method according to claim 6, characterized in that, In step S4, the pH value is adjusted using a 10% citric acid solution or triethanolamine. The homogenization emulsification adopts a gradient speed process: first, stir at a low speed of 300~500 r / min for 10 min to allow the thickener to fully swell, and then increase to a high speed of 800~1200 r / min for homogenization.

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