Improved biopesticides

Abstract

Disclosed are insecticides having polysulfanes as active agents and synthetically prepared or derived from garlic oil. The active agent is held as a dispersed oil phase in an aqueous continuous phase. A surfactant, typically comprising a carbohydrate fatty acid ester, is used to maintain the two-phase system. The emulsion droplet diameter and / or surface area of ​​the emulsion system are controlled to optimize the efficacy of the contained pesticide. Also disclosed are granules having an inert carrier matrix, such as diatomaceous earth or a cellulosic material, e.g., wood cellulose, loaded with a polysulfane-containing emulsion. The use of emulsion-containing granules in soil for controlled release of pesticides in the soil is also disclosed. The use of emulsion systems to promote breaking bud dormancy is also disclosed.

Description

[Technical Field]

[0001] FIELD OF THE INVENTION The present invention relates to improved formulations for delivering biopesticides, which are primarily derived from garlic oil and its synthetic analogues. Additionally, methods for making and using said formulations are disclosed. [Background technology]

[0002] Background of the Invention Given the continuing growth in the world population, the need for increased food production is becoming more urgent, which typically leads to more intensive farming practices with less crop rotation and less fallow land, which in turn increases the risk of infestation by pests adapted to specific crops, especially parasitic pests.

[0003] One crop to which this disclosure pertains, although not exclusively, is the potato. Potatoes can be affected by nematodes of the genus Globodera, commonly referred to as potato cyst nematodes (PCNs). Severe infestations can result in crop losses of up to 90%, and PCNs are estimated to cause losses of approximately £40,000,000 in the UK alone.

[0004] Many methods and products are available for controlling PCNs, but they have only been partially successful. First, as mentioned above, crop rotation is used, which usually means spacing susceptible crops at least six years apart. Alternatively, trap crops are used to attract pests and then eradicate them at the height of the infestation. This is a fairly expensive method to implement and is only relatively effective. Third, chemical agents, primarily carbamates, have been developed. Examples include aldicarb (sold under the brand name Temic) and oxamyl (sold under the brand name Vydate).

[0005] However, there are many problems with this control method. There is evidence that carbamates break down quickly over repeated short-term applications, resulting in reduced effectiveness. Secondly, legislation restricting pesticide use is on the rise, and it is expected that some pesticides will become illegal in the near future.

[0006] Therefore, there is both a need and a desire for pest control agents that are derived from or based on naturally occurring substances.

[0007] One such product is derived from garlic, such as garlic oil, or WO 08 / 5 Synthetic analogs of the active compounds in garlic, such as polysulfanes (also commonly referred to as polysulfides), are disclosed in US Pat. No. 9217. The main challenges to overcome with these compounds are timely and sustainable delivery of the active compound. One approach to addressing these criteria is to incorporate the active compound into granules, which are then eluted from the granules by rainwater or spray water. However, some synthetic analogs present a challenge: compounds containing sulfur chains with four or more atoms are relatively insoluble in water, making them difficult to elute.

[0008] Furthermore, typically only 15% of the product is soluble, the remainder being relatively strongly retained within the granules and unusable. Summary of the Invention [Problem to be solved by the invention]

[0009] The object of the present invention is to provide a biopesticide formulation and a means for its delivery that overcomes the above problems. [Means for solving the problem]

[0010] Summary of the Invention According to a first aspect, there is provided a biopesticide emulsion system having a first continuous phase comprising water and a second discontinuous phase comprising a biopesticide derived from concentrated liquid garlic juice, synthetic polysulfane or a mixture thereof, and including an emulsifier to maintain the two-phase system. Emulsions allow for more effective and efficient delivery of pesticides.

[0011] The diameter of the emulsion droplets of the discontinuous phase is preferably less than 2.4 μm, more preferably less than 2.0 μm, less than 1.5 μm, less than 1.3 μm, or about 1 μm. These specific diameters result in increased mortality when used against target pests. These specific diameters promote bud dormancy breaking when used on plants.

[0012] The specific surface area of ​​the emulsion is preferably 200,000 cm 2 / mL, more preferably 250,000 cm 2 / mL, greater than 300,000 cm 2 / mL or greater than 350,000 cm 2 / mL. These specific surface areas provide increased mortality when used against target pests. These specific surface areas promote bud dormancy breaking when used on plants.

[0013] The emulsifier is preferably an esterified carbohydrate. Particularly preferably, the carbohydrate is sorbitan. Particularly preferably, the sorbitan ester is polyethoxylated to increase the HLB value. A preferred emulsifier is Tween 20, which produces emulsions with the desired droplet size and / or specific surface area when using the above-mentioned pesticides.

[0014] Tween® 20 is an established product used in biological applications and will be familiar to those skilled in the art. It is a polyoxyethylene sorbitol ester that belongs to the polysorbate family. It is a nonionic detergent with a molecular weight of 1,225 daltons and contains 20 ethylene oxide units, one sorbitol, and one lauric acid as its primary fatty acid. The ethylene oxide subunits are responsible for the hydrophilicity of the surfactant, while the hydrocarbon chain provides a hydrophobic environment. Sorbitol forms the backbone ring to which the ethylene oxide polymer is attached.

[0015] In an alternative embodiment, the emulsifier is a sucrose ester, such as sucrose stearate SP 70. Preferably, the sucrose ester has a hydrophilic-lipophilic balance (HLB) value of at least 15 or at least 16.

[0016] The level of emulsifier present is conveniently less than 8% w / w of the emulsion system, for example less than 6% w / w, or less than 5% w / w, or less than 3% w / w, particularly conveniently less than 2% w / w, more conveniently more than 0.25% w / w to ensure a good emulsion droplet size of the discontinuous phase and / or the desired specific surface area of ​​the emulsion system.

[0017] The percent w / w ratio of the continuous phase to the discontinuous phase is preferably greater than 20:1, particularly preferably greater than 30:1.

[0018] In one embodiment, there is provided an emulsion system as described above for promoting breaking bud dormancy.

[0019] According to a second aspect, there is provided a granular pesticide comprising a granular matrix, the granular matrix holding a biopesticide emulsion having a first continuous phase comprising water and a second discontinuous phase comprising a biopesticide derived from concentrated liquid garlic juice, synthetic polysulfone or a mixture thereof, and comprising an emulsifier to maintain the two-phase system.

[0020] The emulsion droplets of the discontinuous phase preferably have a diameter of less than 2.4 μm, less than 2.0 μm, less than 1.5 μm, less than 1.3 μm, or about 1 μm.

[0021] The specific surface area of ​​the biopesticide emulsion is preferably 200,000 cm 2 / mL, more preferably 250,000 cm 2 / mL, greater than 300,000 cm 2 / mL or greater than 350,000 cm 2 / mL. These specific surface areas provide increased mortality when used against target pests. These specific surface areas promote bud dormancy breaking when used on plants.

[0022] The emulsifier is preferably an esterified carbohydrate. Particularly preferably, the carbohydrate is sorbitan. Particularly preferably, the sorbitan ester is polyethoxylated to increase the HLB value. A preferred emulsifier is Tween 20, which, when used with the above-mentioned pesticides, can produce emulsions with the desired droplet size and / or specific surface area.

[0023] The level of emulsifier present is conveniently less than 8% w / w of the emulsion system, for example less than 6% w / w, or less than 5% w / w, or less than 3% w / w, particularly conveniently less than 2% w / w, more conveniently more than 0.25% w / w to ensure good emulsion droplet size and / or the desired specific surface area of ​​the discontinuous phase.

[0024] The percent w / w ratio of the continuous phase to the discontinuous phase is preferably greater than 20:1, particularly preferably greater than 30:1.

[0025] The emulsion is preferably present in a proportion of 0.5% w / w to 1.0% w / w relative to the particulate matrix.

[0026] The particulate matrix is ​​preferably selected from diatomaceous earth or alternatively from cellulosic materials, particularly preferably from wood cellulose.

[0027] According to a third aspect, there is provided a method for producing a granular biopesticide, comprising the steps of: - Producing emulsions of biopesticides in water; - Selection of granular matrix material; - Mixing the emulsion with a granular matrix material to absorb the emulsion into the matrix, wherein the biopesticide is derived from concentrated liquid garlic juice, synthetic polysulfane or a mixture thereof.

[0028] In some embodiments, preparing the emulsion comprises blending at a speed of at least 2000 rpm, at least 3000 rpm, at least 4000 rpm, at least 6000 rpm, or at least 8000 rpm; or about 3000 rpm, about 4000 rpm, about 6000 rpm, or about 8000 rpm.

[0029] The diameter of the emulsion droplets of the discontinuous phase is preferably less than 2.4 μm, more preferably less than 2 The particle size is less than 0.0 μm, less than 1.5 μm, less than 1.3 μm, or about 1 μm.

[0030] The specific surface area of ​​the biopesticide emulsion is preferably 200,000 cm 2 / mL or more, more preferably 250,000 cm 2 / mL or more, 300,000cm 2 / mL or more, or 350,000 cm 2 / mL or more. These specific surface areas provide increased mortality when used against target pests. These specific surface areas promote bud dormancy breaking when used on plants.

[0031] According to a fourth aspect, there is provided a use of a granular biopesticide composition, wherein the granular biopesticide composition comprises a granular matrix and an emulsified biopesticide absorbed within said matrix, wherein the biopesticide is derived from concentrated liquid garlic juice, synthetic polysulfane or a mixture thereof.

[0032] In one embodiment there is provided the use of a granular biopesticide as described above to promote breaking bud dormancy. The present specification is described with reference to figures illustrating results obtained in the preparation and efficacy of the disclosed invention. [Brief explanation of the drawings]

[0033] [Figure 1] Figure 1 shows the relationship between emulsion droplet size and mortality rate; [Figure 2] Figure 2 shows the effect of Tween™ concentration and mixing speed on the mortality of the emulsions produced; [Figure 3] Figure 3 shows the droplet size produced in the synthetic garlic oil emulsion system; [Figure 3a] Figure 3a shows the effect of homogenizer speed on emulsions formed from natural garlic using 1% Tween™ 20; [Figure 4] Figure 4 shows the effect of surfactant concentration on droplet size; [Figure 5] Figure 5 shows the effect of emulsification rate on mortality when synthetic garlic oil was used; [Figure 6] Figure 6 shows the average particle size of several emulsion systems. [Figure 7] Figure 7 shows the specific surface area of ​​several emulsion systems. [Figure 8] Figure 8 shows the mean mortality of nematodes after 1 hour of contact for the two emulsion systems at various dilutions. [Figure 9] FIG. 9 shows the average mortality of nematodes after 3 hours of contact with the two emulsions at various dilutions. [Figure 10] Figures 10 and 11 show the dissolution profiles of the prepared granules; [Figure 11] Figures 10 and 11 show the dissolution profiles of the prepared granules; [Figure 12] Figure 12 shows the mortality of nematodes using the granules of Figures 10 and 11; [Figure 13] Figure 13 shows the mortality of the elution fractions from the materials shown in Figure 10 ; [Figure 14] Figure 14 shows the concentration of sulfur-containing compounds during the elution cycle of synthetic polysulfide-containing granules; [Figure 15] Figure 15 shows the leaching pattern obtained from the soil sample containing the granules. DETAILED DESCRIPTION OF THE INVENTION

[0034] Detailed Description of the Invention The use of granules containing concentrated garlic oil as an active biopesticide is known in the art. When applied in soil to combat recognized pests, the granules can release the biopesticide—in the case of garlic oil, the active sulfur-containing compound—into the soil, where the active compound kills, weakens, or acts as a deterrent to the pests. The granules are most effective when exposed to water, either from natural rainfall or irrigation, with the water serving to transport the active compound from the granule into the soil. This is thought to be primarily due to dissolution of the active compound, although other studies have shown that the active compound is more readily absorbed by the soil. There may be a mechanism at work.

[0035] However, most of the active compound remains within the granules and exerts little effect. Furthermore, the release may be so slow and small that the effectiveness of the granules may be reduced.

[0036] Thus, the present invention acts to mobilize the active compound by emulsifying with water as the continuous phase prior to granulation. Furthermore, it has been found that the emulsification system results in improved efficacy of the biopesticide even when the biopesticide is used without granulation. It has also been found that the emulsification system enhances the effectiveness of the biopesticide for promoting breaking of bud dormancy when the biopesticide emulsion is used without granulation.

[0037] General preparation of emulsions The following is an example of a method for producing an emulsion. However, it will be recognized by those skilled in the art that other methodologies for preparing an emulsion can be used without departing from the scope of the present invention. For example, an emulsifier can be mixed with water and then a biopesticide can be added. Furthermore, the desired emulsion droplet size and / or the desired specific surface area of ​​the emulsion system can be obtained using any suitable means, such as homogenization or high shear mixing, for example, using a Silverson™ mixer.

[0038] An emulsifier solution was prepared by dissolving the emulsifier in the biopesticide at 58°C while stirring. The mixture was then poured into a small Waring cup and homogenized using a Waring blender to prepare the emulsion. A typical preparation cycle includes the following: blend (low speed) 15 seconds, rest 30 seconds, blend (high speed) 15 seconds, rest 30 seconds, blend (high speed) 20 seconds. In some embodiments, emulsion preparation includes at least one blending step at a speed of at least 2000 rpm, at least 3000 rpm, at least 4000 rpm, at least 6000 rpm, or at least 8000 rpm; or about 3000 rpm, about 4000 rpm, about 6000 rpm, or about 8000 rpm. The finished product was stored in a sealed vial until needed.

[0039] To measure particle size, approximately 0.5 ml was removed from each sample and diluted to 1.5 ml. The particle size distribution was then measured using a Beckmann Coulter LS13320 Laser Diffraction Particle Size Analyzer. It was determined.

[0040] The importance of the dispersed phase within the emulsion formed can be clearly seen in Figure 1. The results shown are from an in vitro experiment on emulsion mortality against nematodes. In the experiment, a test solution (0.9 ml) was dispensed into a small tube. Water (0.1 ml) containing 2-300 nematodes was added to the tube to bring the test volume to 1.0 ml. Tests were then carried out at various time intervals at room temperature: 50 μl aliquots of the test solution were removed from the solution to test for nematodes.

[0041] Figure 1 shows emulsions formed from concentrated garlic oil obtained from natural sources. The relationship between droplet size of the dispersed phase and its effect on nematode mortality can be clearly seen: the smaller the droplet size, the higher the mortality, and the higher the absolute value. The effect of garlic oil alone, without added emulsifier, is also shown (second column from the right, labeled "1.2 μm"). The background mortality under the test conditions without added garlic oil is shown in the last column on the far right of the results.

[0042] The effect of Tween™ concentration is shown in Figure 2 for synthetic garlic oil, whose active ingredient is a mixture of diallylpolysulfanes of different sulfur chain lengths. The emulsions for which data are shown in columns 3-7 of Figure 2 were prepared by blending at 8000 rpm. It can be seen that the bioactivity of systems containing 0.5-2.0% w / w Tween™ 20 is higher than that of the system containing 0.1% w / w Tween™ 20, and all are improved over the emulsion made using natural garlic oil (column labeled "clail").

[0043] Figure 2 also includes data for two samples of synthetic garlic oil emulsions prepared using 1% Tween™ 20 as a surfactant: one at a homogenizer speed of 8000 rpm (syn8000) (column 1 in Figure 2) and one at a homogenizer speed of 3000 rpm (syn3000) (column 2 in Figure 2).

[0044] The particle size distribution of emulsions prepared using synthetic oils is more variable than that prepared using natural garlic oil. Figure 3 illustrates this point for syn3000 and syn8000 emulsions prepared as described above. The droplet size of the dispersed phase was measured, and the results are shown in Figure 3. As can be seen, the droplet size at 3000 rpm is much larger than that produced at 8000 rpm. This is in contrast to emulsions formed from natural garlic oil, which showed little effect of homogenizer speed when using 1% Tween™ 20, as shown in Table 1 below and Figure 3a. Without being bound by theory, this may be due to the fact that natural garlic oil contains compounds (such as carbohydrates) that act as surfactants themselves.

[0045] [Table 1]

[0046] Furthermore, Tween™ 20 is effective in producing emulsions at various concentrations. Figure 4 shows the droplet size distribution of emulsions prepared using 0.1% w / w to 2% w / w Tween™ 20. Figure 4 shows that the droplet size distribution remains relatively constant between 0.5 and 2.0% w / w Tween™. However, the position of the maximum peak shifts, and the average droplet diameter actually decreases, especially when 0.1% w / w Tween™ 20 is used.

[0047] When producing emulsions, it is important to pay attention to the hydrophilic / lipophilic balance (HLB) of the surfactant. In general, lipophilic surfactants with low HLB values ​​tend to be more efficient in producing water-in-oil emulsions, while hydrophilic surfactants tend to be more efficient in producing oil-in-water emulsions.

[0048] The importance of the physical structure of the active compound is again shown in Figure 5. 3% w / w synthetic garlic oil was emulsified using sucrose stearate SP70 (S. Black) with an HLB value of 15 as the surfactant at various mixing speeds. It is anticipated that other sucrose esters, including mono-, di-, and tri-esters, may also be applicable as emulsifiers, and this will be apparent to those skilled in the art. Additionally, other fatty acids, such as lauric acid, myristic acid, palmitic acid, etc., may also be applicable as emulsifiers. The resulting emulsion was tested as a nematicide. For comparison, an emulsion containing processed natural garlic oil (also referred to herein as clail or clail021) was also used (CL00021). The results are shown in Figure 5. Bioassays showed that at low mixing speeds, the activity against nematodes was relatively localized. However, above 4000 rpm, a stepwise improvement was observed, with the effect reaching nearly 100% after 24 hours. Considering the above effects in relation to emulsification conditions and droplet size, it is likely that droplets of an effective size were produced by high-speed mixing.

[0049] The relationship between droplet diameter, surface area and biological activity was investigated, and the results are shown in Figures 6 to 9.

[0050] The control was Nemguard SC (CLAIL0021). Nemguard SC is a concentrated garlic juice containing approximately 35% w / w water, approximately 55% w / w complex carbohydrates, approximately 3% w / w garlic oil (polysulfide oil), and approximately 7% w / w protein, electrolytes, and lipids derived from the garlic plant. Nemguard SC was blended at approximately 4000 rpm.

[0051] Two test emulsions were prepared: PK03 (Nemguard SUGO (polysorbate)) and PK03 SE (Nemguard SEGO (also containing sucrose esters)). The PK03 emulsion consisted of approximately 91% w / w water, approximately 6% w / w polysorbate 20, and approximately 3% w / w garlic oil. The PK03 emulsion was blended in a high-shear mixer at four different speeds: 2000 rpm, 4000 rpm, 6000 rpm, and 8000 rpm. The PK03 SE emulsion consisted of approximately 90% w / w water, approximately 6% w / w polysorbate 20, approximately 3% w / w garlic oil, and approximately 1% w / w sucrose esters. The PK03 SE emulsion was mixed in a high-shear mixer at approximately 8000 rpm. All PK03 and PK03 SE emulsions were highly stable, as demonstrated by the consistency of particle size distribution at various dilutions. A total of five test samples were investigated.

[0052] The particle size and specific surface area of ​​emulsions were measured. The measurement process requires the introduction of the test substance into a volume of water within an optical device through which a light beam passes. The scattering pattern indicates the size / surface area of ​​the droplets in solution. The dilution level within the test cell is estimated to be approximately 0.2-0.5% v / v in water.

[0053] Figure 6 shows that all preparations of PK03 emulsions had mean particle sizes at least 40% smaller than Nemguard SC prepared at 4000 rpm. The data also show that particle size decreases with increasing mixing speed, with a significant inflexion point between 6000 and 8000 rpm, where particle size becomes even smaller. It is hypothesized that extraneous components of natural garlic juice (homogenized proteins, dissolved electrolytes, etc.) limit the amount of emulsification that can occur and therefore the particle size achievable with natural garlic juice.

[0054] Figure 7 shows the specific surface area of ​​the emulsions. It can be seen that the droplet surface area increases inversely with droplet diameter. The highest surface area resulted from mixing at 8000 rpm (corresponding to the smallest particle size). All PK03 emulsions had significantly greater droplet surface area than the Nemguard SC control.

[0055] The surface area / volume ratio is given by 3 / r, where r is the radius of the sphere (droplet), and the radius is half the diameter. Therefore, when r is 1, 0.75, 0.5, 0.25, and 0.1, the surface area / volume ratio changes to 3, 4, 6, 12, and 30. Therefore, a droplet with a radius of 0.5 μm (diameter of 1 μm) has twice the surface area / volume ratio of a droplet with a radius of 1 μm (diameter of 2 μm).

[0056] In biology, a larger surface area to volume ratio is generally more efficient than a smaller one. This is due to the amount of cell membrane relative to the cell's volume. The more plasma membrane available for transporting substances into and out of the cell, the more efficiently the cell can perform certain functions that require transport of substances across the membrane. The same principle applies to the translocation of active substances in the various formulations tested herein into the cytoplasm of target organisms and the expression of their cytotoxic effects.

[0057] Two of the PK03 test emulsions (Nemguard-SUGO and Nemguard-SEGO, blended at 4000 rpm) were tested for their bioactivity against nematodes. The emulsions were diluted with water to 0.05%, 0.1%, 0.2%, and 0.4% and used in bioassays with nematodes.

[0058] Figure 8 shows the average mortality of nematodes after 1 hour of contact, and Figure 9 shows the average mortality of nematodes after 3 hours of contact. Figure 8 shows that PK03 emulsion has a more rapid cytotoxic effect at 1 hour, starting from the 0.2% dilution. The presence of sucrose esters produces an even greater effect (Nemguard-SEGO).

[0059] At 3 hours of contact (Figure 9), both PK03 emulsions demonstrated 100% mortality at a 0.2% dilution. At the same concentration, the Nemguard SC control at a 0.2% v / v dilution demonstrated only less than 70% mortality. These results indicate that the PK03 formulations are more active at lower dilutions than the Nemguard SC control.

[0060] The apparent improvement in biological activity of PK03 compared to NEMguard SC appears to be correlated with the smaller droplet diameters and increased droplet surface area generated in the PK03 formulation by the emulsification process. The natural emulsifiers in Nemguard SC may be able to provide similar stability to PK03 emulsions, but may not be able to generate the small droplet sizes seen in PK03 emulsions, potentially delivering more active substance to the target and increasing biological activity.

[0061] The emulsions produced herein can be incorporated into solid dosage forms such as granules, offering advantages over purely liquid spray systems.

[0062] For example, unlike liquid spray systems, the granules themselves are not washed away by water and remain active in the soil for a long period of time. Secondly, the granule composition can be adjusted to provide a release rate suitable for a specific active ingredient and target pest. Surprisingly, it has been found that the above emulsions retain their activity even when incorporated into granules. Furthermore, the supply of active compounds improves the performance of granules containing the same amount of non-emulsified active substances.

[0063] In summary, the emulsion is prepared as described above. The emulsion is then mixed into the granular carrier matrix by sequentially adding the emulsion to the carrier matrix in a coating pan until the desired amount of emulsion is added and well-formed pellets are obtained. The ratio of emulsion to carrier matrix varies depending on the carrier matrix used. For example, when wood flour is used as the carrier matrix, the % w / w mix ratio can be 45:55 emulsion:wood flour. For diatomaceous earth carriers, a ratio of 25:75 is typical.

[0064] A typical granular matrix is ​​wood flour with a granule size of 1.5-2.5 mm. However, the carrier matrix can be selected from wood flour, diatomaceous earth, or Biodac™, which is primarily composed of calcium carbonate (14-20%) and kaolin. It is a cellulosic material (47-53%) containing clay (28-34%) and contains titanium dioxide (less than 1%).

[0065] While garlic oil has been successfully incorporated into granules, the percentage of active ingredient released by water flushing is typically less than 15% of the total amount contained in the granules, even after repeated dissolution. Again, without being bound by theory, it is believed that the polysulfides remaining in the granules tend to be of long sulfur chain length, are fairly tightly bound within the granules, and are relatively insoluble in water. Therefore, the purpose of including an emulsion of the active is to improve the overall release rate. In general, results have shown a 10- to 20-fold increase in the release of active polysulfides from granules prepared from emulsions.

[0066] This is illustrated by the results in Figures 10 and 11 below. As elsewhere, DAS in the specification stands for diallylsulfane, and the numbers indicate the S chain length in the molecule. In these experiments, the release profile of polysulfides from granules was investigated. Granules were compared and water was passed through them repeatedly.

[0067] In the experiment, the granules were mixed in equal weight ratios with Celite 545 (a silica carrier made from diatomaceous earth calcined with flux), and the mixture was loaded into a syringe barrel. A layer of glass wool precented the bottom of the syringe barrel to prevent the granules from falling out of the syringe hub. Once the mixture was loaded, more glass wool was inserted on top. Water in 15 ml aliquots was eluted through the barrel (test cell), and fractions were collected for bioassay. The first aliquot was not usually collected due to absorption.

[0068] The eluted water was analyzed by HPLC to determine the relative amounts and types of polysulfides released into the water.

[0069] The granules used in Figure 10 were single-core wood cellulose granules, with water as the continuous phase and emulsified processed garlic oil (prepared from whole plant extract) as the dispersed phase. 2% Tween™ 20 was added as an emulsifier to emulsify the garlic oil.

[0070] The granules used in Figure 11 utilize a liquid matrix containing 25% w / w sucrose and 5% w / w polysulfide. These granules were formulated to investigate the effect of using non-esterified carbohydrates.

[0071] Comparing Figures 10 and 11 reveals a 10-fold increase in polysulfide release from granules formed using emulsion. These results are reflected in the nematicidal effects of eluate fractions collected from cycle 2 of each of the above granule types, as shown in Figure 12. In these experiments, bioassays were performed by transferring 0.9 ml of cycle 2 eluate to Eppindorf tubes. Next, water (100 μl) containing 250–350 fresh nematodes was added to the Eppindorf tubes, which were then closed and gently mixed. Immediately afterward, and every hour thereafter, 50 μl aliquots were removed and nematode mortality was measured. A water control was also included.

[0072] The results shown in Figure 12 show mortality versus time. Granules prepared with polysulfide emulsion (B28) were found to have a significantly increased effect on nematode mortality, presumably related to higher release.

[0073] As further evidence of the effectiveness of the eluates from granules prepared from the emulsified material, the nematicidal effect of the first five eluates (detailed in Figure 10) is shown in Figure 13. In this experiment, the eluates were in contact with nematodes for 18 hours.

[0074] In further experiments, granules were prepared using synthetic polysulfide oil emulsified in water with sucrose ester (SP70) at levels of 3% and 4% by weight of polysulfide oil. The dissolution pattern of this compound is shown in Figure 14.

[0075] The effectiveness of granular carriers for emulsifying active materials in soil was also investigated. Emulsions were formed using a moderate sheer of concentrated garlic oil in water with 1% w / w Tween™ 20 as an added surfactant. The resulting emulsion was sprayed onto the granule manufacturing process to produce the carrier.

[0076] The resulting granules were mixed with soil and leached once daily for 15 days. Sequential fractions were collected and analyzed for polysulfide concentration. The leaching profile is shown in Figure 15. As can be seen, almost all of the released polysulfides appeared by the end of the fourth day. In vitro testing of the granules with Celite™ 545 demonstrated high direct nematicidal activity against helminthic nematodes (J2s). Furthermore, polysulfide concentrations in cycles 2-4 were more than 10 times higher than those of commercial granules containing concentrated garlic oil, demonstrating demonstrable activity in soil.

[0077] The field test was conducted on behalf of the applicant by a contract testing agent accredited by ORETO (Official Recognition of Efficacy Testing Organizations and facilities). It was carried out in a typical silty soil of the Shah province.

[0078] An emulsion according to the present disclosure was prepared using natural garlic extract. Additionally, an emulsion according to the present disclosure was prepared containing a synthetic formulation having the same weight ratio of polysulfide oil composition as 12L of natural garlic oil product of natural garlic extract. This preparation consisted of water, polysorbate 20, and polysulfide oil mixed into the emulsion. Equal areas of potato infested with nematodes were treated as follows: - control (no spraying); - Device control (water injector); - Natural garlic emulsion through an injector at a dose of 6 L / ha; - Natural garlic emulsion through an injector at a dose of 12 L / ha; - Natural garlic emulsion through injectors at a dose of 24 L / ha; - 12 L / ha of synthetic emulsion administered through an injector; - Nemathorin® (Syngenta), a commercially available registered nematicide containing fosthiazate for the control of potato cyst nematodes, was applied at the recommended dose.

[0079] result The total yield recovered from the fields and the final average PCN population (measured as eggs / gram of soil) are shown in Table 2 below.

[0080] [Table 2]

[0081] These results demonstrate the effectiveness of the emulsions of the present disclosure: applications of 24 L of natural emulsion and 12 L of synthetic emulsion increased yield by comparable amounts compared to the untreated control, the device control, and the commercial nematicide Nemathorin®.

[0082] Furthermore, application of 24 L of natural emulsion and 12 L of synthetic emulsion reduced the final population of PCNs at comparable doses compared to untreated and device controls, thus demonstrating nematicidal efficacy.

[0083] These results indicate that synthetic emulsions may exhibit greater efficacy than natural products in a dose-to-dose comparison.

[0084] It has also been surprisingly discovered that the biopesticide emulsions described herein are effective in promoting the breaking of bud dormancy.

[0085] Bud dormancy is a process in which plant meristems become largely inactive. Its purpose is to enable plants to withstand harsh environmental conditions. Various types of environmental stimuli can induce bud dormancy. Similarly, various types of environmental stimuli, such as changes in temperature and day length, can break bud dormancy. However, climate change has affected bud dormancy and its breaking. In particular, warming in autumn, winter, and spring affects bud dormancy breaking, affecting the timing and intensity of growth resumption and flowering progression.

[0086] The effect of PK03 (Nature Identical Product (NIP)) emulsion in altering germination was investigated in comparison with the untreated control, and the results are shown in Table 3. The PK03 emulsion had the same composition as above.

[0087] [Table 3]

[0088] All of the above applications were made on February 1, 2023, 60 days before expected emergence. Results showed that at all NIP doses, the number of open shoots was significantly higher in the advanced BBCH (11–13) compared with the untreated control. BBCH (BBCG) is a scale used to identify plant phenological development stages.

[0089] With an application of 5 L / ha, approximately 35% of open shoots emerged in BBCH11-13 compared with 15.5% in the untreated control.

[0090] The biochemical processes involved in breaking bud dormancy are highly complex and are only just beginning to be elucidated in detail. However, one of the key factors in initiating and subsequently releasing plant bud dormancy is the level and duration of oxidative stress in bud cells and tissues, and how this affects mitochondrial activity (the cellular powerhouse). The concentration of low-molecular-weight thiols, particularly glutathione (GSH), is thought to play a key role in maintaining plant tissues in a dormant state, as GSH is a key regulator of cellular redox potential. As the GSH pool decreases over time (under winter conditions), oxidative stress gradually develops, possibly reaching a critical redox potential, triggering increased metabolic activity and mitochondrial activation to generate energy to break dormancy and initiate shoot growth.

[0091] Polysulfides in PK03 are known to react with GSH-depleting concentrations to generate oxidative stress, which is thought to affect mitochondrial activity in a way that breaks bud dormancy.

[0092] Aspects The following are non-limiting aspects of the present disclosure.

[0093] 1. An emulsion system having a first continuous phase comprising water and a second discontinuous phase comprising a biopesticide derived from concentrated liquid garlic juice, synthetic polysulfone or mixtures thereof, and including an emulsifier to maintain the two-phase system. 2. The emulsion system of aspect 1, wherein the emulsion droplets of the discontinuous phase have a diameter of less than 2.4 μm. 3. The emulsion system of embodiment 2, wherein the diameter is less than 2.0 μm, less than 1.5 μm, less than 1.3 μm, or about 1 μm. 4. The emulsion has a viscosity of 200,000 cm 2 10. The emulsion system of any preceding embodiment, having a specific surface area of ​​greater than 1 / mL. 5. The specific surface area is 250,000 cm 2 / mL, greater than 300,000 cm 2 / mL or greater than 350,000 cm 2 5. The emulsion system of embodiment 4, wherein the emulsion system has a solubility of greater than 1 / mL. 6. The emulsion system of any preceding aspect, wherein the emulsifier is an esterified carbohydrate. 7. The emulsion system of aspect 6, wherein the carbohydrate is sorbitan. 8. The emulsion system of aspect 7, wherein the sorbitan ester is polyethoxylated to increase the HLB value. 9. The emulsion system of any one of aspects 1-5, wherein the emulsifier is Tween® 20. 10. The emulsion system of any preceding aspect, wherein the emulsifier is present at less than 3% w / w, or less than 2% w / w of the emulsion system. 11. The emulsion system of any preceding aspect, wherein the emulsifier is present at greater than 0.25% w / w. 12. The emulsion system of any preceding embodiment, wherein the percent w / w ratio of continuous to discontinuous phase is greater than 20:1, optionally said ratio is greater than 30:1. 13. An emulsion system according to any preceding aspect for use as a biopesticide. 14. The emulsion system of any preceding aspect for promoting breaking bud dormancy. 15. A granular composition comprising a granular matrix maintaining an emulsion having a first continuous phase comprising water and a second discontinuous phase comprising a biopesticide derived from concentrated liquid garlic juice, synthetic polysulfone or mixtures thereof, the granular composition comprising an emulsifier to maintain the two-phase system. 16. The granular composition of aspect 15, having a diameter of less than 2.4 μm. 17. The granular composition of embodiment 16, having a diameter of less than 2.0 μm, less than 1.5 μm, less than 1.3 μm, or about 1 μm. 18. The biopesticide emulsion has a viscosity of 200,000 cm 2 18. The granular composition according to any one of aspects 15 to 17, having a specific surface area of ​​greater than 1 / mL. 19. The specific surface area is 250,000 cm 2 / mL, greater than 300,000 cm 2 / mL or greater than 350,000 cm 2 19. The granular composition of embodiment 18, wherein the granular composition has a molecular weight of greater than 1 / mL. 20. The granular composition of any one of aspects 15 to 19, wherein the emulsifier is an esterified carbohydrate. 21. The granular composition of aspect 20, wherein the carbohydrate is sorbitan. 22. The granular composition according to aspect 21, wherein the sorbitan ester is polyethoxylated to increase the HLB value. 23. The granular composition of any one of aspects 15-19, wherein the emulsifier is Tween™ 20. 24. A granular composition according to any one of aspects 15 to 23, wherein the emulsifier in the emulsion is present at less than 3% w / w or less than 2% w / w of the emulsion. 25. The granular composition of any one of aspects 15 to 24, wherein the emulsifier is present at greater than 0.25% w / w. 26. The granular composition of any one of aspects 15-25, wherein the percent w / w ratio of continuous to discontinuous phase of the emulsion is greater than 20:1, or greater than 30:1. 27. A granular composition according to any one of aspects 15 to 26, wherein the emulsion is present in a proportion of 0.5 to 1.0% w / w relative to the granular matrix. 28. The granular composition of any one of aspects 15-27, wherein the granular matrix is ​​selected from diatomaceous earth or a cellulosic material, preferably, the granular matrix is ​​wood cellulose. 29. The granular composition of any one of aspects 15 to 28 for use as a pesticide. 30. The following process: - the process of producing an emulsion of biopesticides in water; - selecting a granular matrix material; and - mixing the emulsion with the particulate matrix material and absorbing the emulsion into the matrix; A method for producing a granular composition comprising: A method wherein said biopesticide is derived from concentrated liquid garlic juice, synthetic polysulfane or a mixture thereof. 31. The method of embodiment 29, wherein preparing the emulsion comprises blending at a speed of at least 2000 rpm, at least 3000 rpm, at least 4000 rpm, at least 6000 rpm, or at least 8000 rpm; or about 3000 rpm, about 4000 rpm, about 6000 rpm, or about 8000 rpm. 32. Use of a granular composition, said granular composition comprising a granular matrix and an emulsified biopesticide absorbed within said matrix, said biopesticide being derived from concentrated liquid garlic juice, synthetic polysulfane or a mixture thereof. 33. The use according to aspect 32 as a biopesticide.

[0094] Various modifications to the embodiments of the invention described herein will be readily apparent to those skilled in the art, and such modifications are intended to be included within the scope defined in the appended claims.

Claims

1. A biopesticide emulsion system having a first continuous phase comprising water and a second discontinuous phase comprising a biopesticide derived from concentrated liquid garlic juice, synthetic polysulfane or a mixture thereof, and including an emulsifier to maintain the two-phase system.

2. 10. The emulsion system of claim 1, wherein the discontinuous phase emulsion droplets have a diameter of less than 2.4 μm.

3. 3. The emulsion system of claim 2, wherein the diameter is less than 2.0 μm, less than 1.5 μm, less than 1.3 μm, or about 1 μm.

4. The emulsion has a viscosity of 200,000 cm ２ 4. The emulsion system according to claim 1, having a specific surface area of ​​greater than 1 / mL.

5. The specific surface area is 250,000 cm ２ / mL, greater than 300,000 cm ２ / mL or greater than 350,000 cm ２ 5. The emulsion system of claim 4, wherein the emulsion system has a viscosity of greater than 1000 saturations per mL.

6. An emulsion system according to any one of claims 1 to 5, wherein the emulsifier is an esterified carbohydrate.

7. 7. The emulsion system of claim 6, wherein the carbohydrate is sorbitan.

8. 8. The emulsion system of claim 7, wherein the sorbitan ester is polyethoxylated to increase its HLB value.

9. 6. An emulsion system according to any one of claims 1 to 5, wherein the emulsifier is Tween 20®.

10. 10. An emulsion system according to any preceding claim, wherein the emulsifier is present at less than 3% w / w, or less than 2% w / w of the emulsion system.

11. 11. An emulsion system according to any one of claims 1 to 10, wherein the emulsifier is present at greater than 0.25% w / w.

12. 12. An emulsion system according to any one of claims 1 to 11, wherein the percent w / w ratio of continuous to discontinuous phase is greater than 20:1, optionally the ratio is greater than 30:

1.

13. An emulsion system according to any one of claims 1 to 12 for promoting breaking of bud dormancy.

14. A granular pesticide comprising a granular matrix, the granular matrix holding a biopesticide emulsion having a first continuous phase comprising water and a second discontinuous phase comprising a biopesticide derived from concentrated liquid garlic juice, synthetic polysulfane or a mixture thereof, the granular pesticide comprising an emulsifier to maintain the two-phase system.

15. 15. The granular pesticide according to claim 14, having a diameter of less than 2.4 μm.

16. The diameter is less than 2.0 μm, less than 1.5 μm, less than 1.3 μm, or about 1 μm. Item 16. The granular pesticide according to Item 15.

17. The biopesticide emulsion has a viscosity of 200,000 cm ２ The granular pesticide according to any one of claims 14 to 16, having a specific surface area of ​​more than 1 / mL.

18. The specific surface area is 250,000 cm ２ / mL, greater than 300,000 cm ２ / mL or greater than 350,000 cm ２ The granular pesticide according to claim 17, wherein the granular pesticide has a water content of greater than 1 / mL.

19. The granular pesticide according to any one of claims 14 to 18, wherein the emulsifier is an esterified carbohydrate.

20. 20. The granular pesticide according to claim 19, wherein the carbohydrate is sorbitan.

21. 21. The granular pesticide according to claim 20, wherein the sorbitan ester is polyethoxylated to increase its HLB value.

22. The granular pesticide according to any one of claims 14 to 18, wherein the emulsifier is Tween (trademark) 20.

23. 23. A granular pesticide according to any one of claims 14 to 22, wherein the emulsifier in the emulsion is present in an amount of less than 3% w / w or less than 2% w / w of the emulsion.

24. A granular pesticide according to any one of claims 14 to 23, wherein the emulsifier is present in an amount of more than 0.25% w / w.

25. 25. A granular pesticide according to any one of claims 14 to 24, wherein the percent w / w ratio of continuous to discontinuous phase of the emulsion is greater than 20:1 or greater than 30:

1.

26. 26. The granular pesticide according to any one of claims 14 to 25, wherein the emulsion is present in a ratio of 0.5 to 1.0% w / w relative to the granular matrix.

27. 27. A granular pesticide according to any one of claims 14 to 26, wherein the granular matrix is ​​selected from diatomaceous earth or cellulosic materials, preferably the granular matrix is ​​wood cellulose.

28. 1. A method for producing a granular biopesticide, wherein the biopesticide is derived from concentrated liquid garlic juice, synthetic polysulfane or a mixture thereof, the method comprising the steps of: - preparing an emulsion of the biopesticide in water; - selecting a particulate matrix material; and - mixing the emulsion with the particulate matrix material and absorbing the emulsion into the matrix.

29. 30. The method of claim 28, wherein producing the emulsion comprises blending at a speed of at least 2000 rpm, at least 3000 rpm, at least 4000 rpm, at least 6000 rpm, or at least 8000 rpm; or about 3000 rpm, about 4000 rpm, about 6000 rpm, or about 8000 rpm.

30. 1. Use of a granular biopesticide composition, the granular biopesticide composition comprising a granular matrix and an emulsified biopesticide absorbed within the matrix, the biopesticide being concentrated. Use derived from concentrated liquid garlic juice, synthetic polysulfones or mixtures thereof.

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