Herbicide formulation and enhanced composite microspheres

Abstract

A herbicidal formulation comprising a herbicidal active ingredient in aqueous solution and composite microspheres that enhance herbicidal efficacy against weeds in crops; wherein said composite microspheres, in powder form, are present in a concentration ranging from 2% to 100% w / w relative to the herbicide when combined with said aqueous solution, and wherein said composite microspheres include chitosan, with an average molecular weight in the range of 190,000 to 310,000 g / mol, and a crosslinking agent. These composite microspheres enhance herbicidal efficacy against weeds in crops.

Description

[0001] HERBICIDE FORMULATION AND ENHANCED COMPOSITE MICROSPHERES

[0002] Cross-reference to related Application

[0003] This application claims the benefit of U.S. Provisional Patent Application No. 62 / 621,881, filed on January 17, 2024, which is incorporated by reference herein in its entirety.

[0004] Field of the Invention

[0005] The present invention relates to the field of smart materials designed to improve the efficiency of chemicals used in agricultural processes. Specifically, the present invention pertains to a microstructured polymeric compound with tailored chemical composition, charge distribution, and advantageous properties for its application in the dosage and controlled release of agrochemicals for sustainable agricultural practices. In particular, it relates to herbicide formulations enhanced with composite microspheres. The present invention also includes scalable, high-conversion, verified, and validated methods for their preparation.

[0006] Background of the Invention

[0007] The use of fertilizers and pesticides has been a cornerstone in the development of modern agriculture, enabling improved crop yields to meet the growing global demand for food.

[0008] In general terms, agrochemicals are understood to encompass any substance or mixture of natural or synthetic substances used to prevent, eliminate, and / or control any pest, disease, or weed in agricultural activities. These substances are commonly referred to as pesticides or crop protection agents— also known as phytosanitary products— and include insecticides, herbicides, fungicides, acaricides, among others. This categorization of agrochemicals also includes substances aimed at providing elements that promote plant growth, commonly referred to as fertilizers and plant growth regulators.

[0009] Fertilizers and plant growth regulators are agrochemicals used to improve crop quality and growth. Fertilizers supply essential nutrients to plants, enhance root quality in soil, and facilitate faster and higher-quality plant growth and development. Plant growth regulators, on the other hand, are products that regulate plant growth, typically composed of plant hormones (phytohormones). Their main functions are to stimulate or inhibit the development of roots and aerial parts of the plant.

[0010] Herbicides are used to eliminate the main harmful plants affecting crops, commonly known as weeds. They are available in various types depending on their characteristics, such as application timing, the extent of their effect on the plant, or the specific phase in which they are applied.

[0011] Crops face numerous challenges, including insects, pests, diseases, and the toxicity associated with pesticides. Pests cause global crop losses of up to 50% in wheat and 80% in cotton. Other crops experiencing significant production losses include soybean (26- 29%), maize (31%), rice (37%), and potatoes (40%) [Oerke, 2006], Insect pests account for approximately 30% of crop losses [Vinutha, 2013], while weeds contribute to substantial losses of around 34%.

[0012] In addition to pests, fungal diseases also have a worldwide impact on crops. Fungi are responsible for approximately 70% of diseases affecting major agricultural crops [Agrios, 2005], including wheat, potatoes, cotton, tomatoes, peanuts, grapes, and cotton [Dhekney, 2005], The severity of the situation becomes evident when considering that the annual crop losses in the agricultural industry due to fungal diseases, both in the field and post-harvest, exceed $200 billion. In contrast, more than $600 million is spent annually on fungicides in the United States alone [Fernandez-Perez, 2000], Globally, approximately one-quarter of food crops are damaged by fungal toxins, such as aflatoxins, ergot toxins, Fusarium toxins, patulin, and tenuazonic acid [Razzaghi- Abyaneh, 2014],

[0013] Weeds compete with crops for light, water, nutrients, and space. This competition can reduce crop growth and affect its development (Segundo Congreso Argentine de Girasol, ASAGIR Workshop on Sunflower Weeds). Depending on the type and density of weeds, yield reductions ranging from 10% to 90% in extreme cases have been observed.

[0014] Fallowing, the practice of leaving a plot of land uncultivated for several months or years before replanting, often requires weed control between crop cycles. Poor management during fallow periods can lead to the emergence of herbicide-resistant weeds, such as Amaranthus spp. (redroot pigweed) and Lolium spp. (ryegrass), which pose significant challenges in the Pampa Humeda region of Argentina [Bedmar, 2002],

[0015] The impact of weed competition on soybean crop yield depends on the timing of weed emergence and the duration of interference. The following factors have been identified in this regard:

[0016] 1. Early Competition (First 4-6 Weeks): The most critical period is the initial growth stage (up to V4 or V5), during which soybean establishes its architecture and generates leaves for photosynthesis. Weed competition during this phase can reduce yields by 20-50% if not properly controlled.

[0017] 2. Interference from Problematic Weeds: Weeds such as Echinochloa spp. (barnyard grass) and Conyza spp. (horseweed) are particularly competitive. If inadequately managed, they can result in yield losses of up to 80% in cases of high infestation.

[0018] 3. Control Strategies: The use of pre-emergent and post-emergent residual herbicides is key to reducing the impact of weeds during sowing and crop growth. In areas with high pressure from resistant weeds, constant monitoring and the combination of different herbicide modes of action are essential.

[0019] In soybean cultivation, yield losses due to weed competition during fallow periods and early growth stages can range from 10% to 70%. In poorly managed fields or areas with resistant weeds, losses can exceed 90% if competition is not controlled in time. Weed control during fallow and early crop stages is therefore critical to maximizing soybean yield and preventing significant economic losses. A small improvement in weed control, from 85% to 90%, may seem minor in terms of percentage, but it can have a significant impact on agricultural yield [Green-Tracewicz, 2012],

[0020] Pesticides have been widely used to combat pests; however, their extensive application has had devastating effects on humans and other living organisms, with an increasing incidence of human poisoning.

[0021] The adverse effects of agrochemical use on human health can be described as follows: • Chronic Diseases: Long-term exposure to agrochemicals has been linked to chronic illnesses.

[0022] • Acute Health Problems: Accidental exposure to high doses can cause acute poisoning, with symptoms such as dizziness, vomiting, respiratory difficulties, and even death.

[0023] • Risks to Vulnerable Populations: Pregnant women and children are particularly susceptible to the neurotoxic and endocrine-disrupting effects of certain agrochemicals, potentially affecting physical and cognitive development [Islam, 2024],

[0024] The indiscriminate use of agrochemicals has disrupted natural balances, causing detrimental effects on various ecosystems, including:

[0025] • Soil Contamination: Reduced natural fertility, altered microbiota, and processes such as salinization or acidification.

[0026] • Water Contamination: Excessive application of agrochemicals can leach into rivers, lakes, and groundwater, leading to water pollution and phenomena such as eutrophication, where excess nutrients promote the proliferation of toxic algae, affecting water quality and aquatic life.

[0027] • Loss of Biodiversity: Excessive use of agrochemicals has led to resistance development in pests, weeds, and pathogens.

[0028] The report on the environmental and health impacts of pesticides and fertilizers, prepared in close collaboration with the Food and Agriculture Organization (FAO) and the World Health Organization (WHO), highlights the urgent need for transformative actions and improved management of pesticides and fertilizers as their global demand and use continue to rise,

[0029] (wedocs. unep.org / xmlui / bitstream / handle / 20.500.11822 / 38409 / pesticides. pdf).

[0030] There exists a pressing need to adopt sustainable practices that mitigate the negative impacts of excessive agrochemical use. In recent years, alternatives have been developed to complement traditional agriculture, aiming to reduce these adverse effects. One such approach is precision agriculture, which seeks to optimize agricultural practices by utilizing advanced technologies to collect, analyze, and interpret data on crops, soil, climate, and other factors to make more efficient decisions. The primary goal of precision agriculture is to maximize resource efficiency— such as water, fertilizers, and pesticides— while minimizing environmental impacts and enhancing productivity.

[0031] Nanotechnology has provided innovations and technologies that address these challenges by optimizing resource use. Nanotechnology is widely employed in modern agriculture to actualize the concept of precision agriculture [Duhan, 2017], It involves various materials, including the development of nano- and composite microspheres with one or more dimensions at the scale of 100 nm or less. Nanomaterials find extensive applications in plant protection, nutrition, and agricultural practices due to their unique properties, such as their small size and high surface-to-volume ratio [Auffan, 2009],

[0032] The development of controlled-release formulations (CRFs) is highly desirable to comply with international environmental and biodiversity regulations. The use of polymers for sustained release began in the early 1970s, offering greater efficacy for encapsulated components compared to commercial formulations. To enhance the value of traditional pesticides, many natural molecules, including carrageenan, galactomannans, chitosan, alginate, pectin, cellulose, gum arabic, guar gum, cashew gum, chitin, agar, tamarind seed polysaccharide, and starch, are frequently processed into nanomaterials for sustained and targeted release to specific sites of action for various agrochemicals [Salgueiro, 2013; Albuquerque, 2016; Sharma, 2019; Slade, 2019; Rashidipour, 2019; Elabasy, 2020],

[0033] Among the polymers commonly used in the state of the art, chitosan is particularly notable. Chitosan is a polysaccharide composed of repeating units of D-glucosamine and N-acetyl-D-glucosamine. It is the primary industrial derivative of chitin and can be obtained through the N-deacetylation of chitin. Chitosan is poorly soluble in water; however, when the degree of deacetylation reaches at least 50%, it dissolves as a polyelectrolyte in dilute aqueous acids [Robert, 1992], The degree of deacetylation and molecular weight of chitosan significantly influence its physicochemical and biological properties, such as solubility, hydrophobicity, crystallinity, and cellular response. In recent decades, chitosan has garnered growing interest in the agricultural sector. For instance, when dissolved in an acidic solution, chitosan activates its antimicrobial properties and stimulates plant defense mechanisms, providing the potential to become a new class of plant stress protectors [Bautista-Banos, 2006; Sahariah, 2017],

[0034] In the publication by Rychter [Rychter, 2019], chitosan microparticles encapsulating glyphosate were evaluated for their herbicidal effect and controlled-release properties.

[0035] Another approach to utilizing chitosan in various applications, particularly for agricultural purposes, involves forming three-dimensional structures through the crosslinking of chitosan molecules. These stable structures enable the nanoencapsulation of active components and are characterized by providing a more controlled release of the encapsulated component. Slow release, protection against degradation, and low solubility of the encapsulated insecticide are the most important characteristics of polymeric nanoformulations, making them the preferred choice for nanoencapsulation. This valuable information has paved the way for the development and practical application of polymeric nanoinsecticides with immense potential. Additionally, various types of nanoformulations, both polymer-based and non-polymer- based, have been proposed, including nanospheres, nanocapsules, nanogels, micelles, nanofibers, nanometals, and nanoemulsions, for the encapsulation of insecticides. Among these, nanocapsules are the most commonly used for the controlled release of insecticides. Recently, a novel concept of hybrid nanoformulations (nanoemulsion encapsulation or liposome coating) has been suggested for the controlled release of certain insecticides. However, the efficacy of this innovative approach needs to be tested across a broad range of insecticides [Das, 2014].

[0036] In the document by Grenha (2012) [Grenha, 2012], methods forthe preparation of nanostructured systems based on chitosan are described, primarily through emulsification, various types of coacervation, or slight modifications of both techniques. Specifically, the methods include emulsion droplet coalescence [Tokumitsu et al., 1999], solvent diffusion in emulsions [El-Shabouri, 2002], the reverse micelle method [Mitra et al., 2001], ionic gelation, polyelectrolyte complexation [Calvo et al., 1997; Sarmento et al., 2006], and desolvation [Tian & Groves, 1999], All these methods involve bottom-up manufacturing processes, which entail the assembly of molecules in solution to form defined structures [Chan & Kwok, 2011], In this context, composite nanomicro-spheres are obtained.

[0037] The release systems resulting from bottom-up technologies typically exhibit size polydispersity [Wang et al., 2011], which in certain cases limits the utility of nanoparticles. Indeed, in polydisperse systems, it is assumed that larger nanoparticles may have greater active ingredient loading capacity, while smaller nanoparticles are expected to be more efficient in delivering agrochemicals to tissues or cells [Fan et al., 2012],

[0038] Chitosan microparticles and nanoparticles have been fabricated through chemical crosslinking with glutaraldehyde, glyoxal, and polyethylene glycol diglycidyl ether. Although these are highly effective crosslinking agents, they are not preferred due to their physiological toxicity. Chitosan, being polycationic in acidic media (pKa 6.5), can interact with negatively charged species, such as the crosslinker TPP (sodium tripolyphosphate) and sodium sulfate. This property can be exploited to prepare crosslinked chitosan nanoparticles. The interaction of chitosan with TPP leads to the formation of biocompatible crosslinked chitosan nanoparticles, which can be efficiently employed in the delivery of proteins, vaccines, and various other compounds. The crosslinking density, crystallinity, and hydrophilicity of crosslinked chitosan enable modulation of the release of encapsulated drugs and expand its range of potential applications in drug delivery [Bhumkar, 2006],

[0039] Chitosan, with its high degree of protonation in its amine groups, has the ability to form hydrogels in the presence of specific polyanions. This process, derived from inter- and intramolecular crosslinking mediated by anionic molecules [Janes et al., 2001; Terbojevich & Muzzarelli, 2009], is known as ionic gelation or polyelectrolyte complexation. It has been used to produce chitosan nanoparticles. This method involves ionic interaction between the positively charged amino groups of chitosan and the polyanion sodium tripolyphosphate (TPP), which acts as the crosslinking agent for chitosan. It is worth noting that the term ionic gelation is preferred when chitosan gelation is induced by small anionic molecules, such as phosphate, citrate, or sulfate. In contrast, polyelectrolyte complexation refers to cases where anionic macromolecules are used instead of small molecules [Bhattarai et al., 2010], The formation of nanoparticles via ionic gelation is described in the work of Carvalho [Carvalho et al., 2009], Several studies in the state of the art demonstrate crosslinked chitosan nanoparticles with TPP or other biopolymers containing an encapsulated active compound, in which their fungicidal or herbicidal activity is described depending on the encapsulated compound.

[0040] Additionally, several studies in the state of the art have combined chitosan nanoparticles loaded or encapsulated with herbicides. For instance, in the work of Dos Santos Silva and collaborators (2011), alginate / chitosan nanoparticles measuring 635 ± 12 nm with a zeta potential of -22.8 ± 2.3 mV and an association efficiency of 74.2% were prepared as an eco-friendly carrier system to encapsulate the herbicide Paraquat. The nanoparticles (NPs) conjugated with Paraquat improved the herbicide's release profile and its interaction with soil, demonstrating that this formulation could effectively reduce the adverse impacts of Paraquat [Dos Santos Silva, 2011],

[0041] In a similar approach, Grillo et al. (2014) carried out a comparable study using chitosan / tripolyphosphate (CSTPP) nanoparticles loaded with Paraquat. An encapsulation efficiency of 62.66 ± 0.77% was achieved, indicating a strong affinity between the CSTPP NPs and the active component of the herbicide. The herbicidal activity was evaluated in maize (Zea mays) and mustard (Brassica sp.). Both free and encapsulated forms of Paraquat caused flaccid foliar necrosis within 48 hours, a characteristic effect of this herbicide in both plants. In Z. mays, the use of the nanoherbicide resulted in more significant necrosis, likely due to better adhesion of the nanoparticles to the leaf, as their higher surface-to-volume ratio enhances interaction.

[0042] Finally, in a study by Butstraen and collaborators [Butstraen, 2014], microcapsules of chitosan and gum arabic crosslinked with TPP were loaded with commercial Miglyol 812 N. The microcapsules were obtained using the coacervation method, and different chitosan-to-gum arabic mass ratios of 1:4 and 1:5 were evaluated with TPP crosslinker in the presence of Miglyol during synthesis. The physicochemical parameters of the microparticles, their encapsulation capacity, and the zeta potential of the particles were assessed. This study aimed to identify optimal encapsulation conditions for Miglyol but did not describe or imply a correlation between TPP concentration and the zeta potential of the formed particles. The study suggests an optimal chitosan-to-gum arabic mass ratio of 1:4, while the present invention proposes a much broader range of 5:1 to 20:1. This broader range enables the herbicidal formulation of the present invention, which contains the composite microspheres, to be more readily redispersible in aqueous agrochemical solutions and prevents clogging of spray nozzles in field applications.

[0043] None of the prior art references disclose or suggest a composite microsphere like the one in the present invention, which does not encapsulate agrochemical actives but is instead added to a herbicidal active ingredient to enhance its efficacy. This enhancement allows for a reduction in the concentration of herbicide used while maintaining its effectiveness.

[0044] Furthermore, none of the prior art references demonstrate the regulation of surface charge or zeta potential of the composite microsphere by adjusting the concentration of the crosslinking agent (preferably TPP) during the synthesis process. Additionally, none of the prior art references suggest the use of gum, a preferred embodiment in the present invention, to improve the stability of the redispersion of chitosan-based composite microspheres when combined with a herbicidal solution.

[0045] The resuspension capacity of the microspheres described in the present invention is a crucial property for ensuring their agronomic application. Without this feature, their agglomeration and poor dispersion can clog the filters of spray nozzles in various types of applicators or sprayers.

[0046] Although the state of the art proposes the use of gums as dispersants during the synthesis process of microparticles made from natural polymers such as chitosan, there are no reports of using gums to enhance the stability of these particles during redispersion when added to pesticide formulations, including herbicides. The function of the gum in the present invention, in agronomic practice, is to prevent clogging of spray nozzles during agrochemical application processes.

[0047] The present invention provides a composite microsphere comprising chitosan and a crosslinking agent, and, in a preferred embodiment, a dispersing agent such as gum. The composite microsphere of the present invention can be synthesized by regulating its zeta potential through an adjustable concentration of TPP, which allows fine-tuning of the zeta potential in an aqueous solution. This distinctive feature of the present invention enables the design and selection of the desired composition, allowing it to be combined with various chemical compounds.

[0048] When formulated with a herbicidal active ingredient, the particle of the present invention enhances the effect of the herbicide.

[0049] The ability to adjust the zeta potential enables customization of the microspheres for different pH levels, salinity conditions, and soil types. This versatility allows the microspheres to be employed across a wide variety of crops and agricultural systems. The capacity to regulate the zeta potential through the concentration of the crosslinking agent suggests that the microspheres can control surface charge, influencing the release of active compounds (fertilizers, pesticides, or micronutrients) in aqueous solutions. Zeta potential control allows for the design of microspheres that selectively interact with specific surfaces or ions, adapting to the needs of crops and / or soil. This facilitates targeted delivery of active ingredients to specific action sites (plants, pathogens, or soil), thereby optimizing the use of agricultural inputs.

[0050] From an application perspective, these microspheres enable sustained and efficient delivery of agrochemicals and fertilizers, both chemical and biological, as well as phytosanitary products, reducing losses due to leaching or evaporation and increasing availability at biological targets (plants, fungi, insects). Additionally, the preferred components (chitosan, gum arable, and TPP) are biodegradable and non-toxic, making the microspheres environmentally friendly. As a result, they reduce the environmental impact compared to synthetic matrices, promoting sustainable agriculture and protecting soil biodiversity. Moreover, chitosan possesses antimicrobial properties that can protect plants against pathogens and fungal diseases.

[0051] The present invention provides composite microspheres that can be used as adjuvants in agrochemical formulations. Collectively, these properties make the composite microsphere an innovative and promising tool to enhance the efficiency of agricultural inputs, reduce environmental impact, and promote sustainability in modern agriculture.

[0052] The present invention distinguishes itself from related prior art by being combinable with the herbicide prior to its application, without requiring encapsulation during the particle synthesis process. Additionally, the agrochemical formulation of the present invention contains particles in concentrations that do not exhibit herbicidal activity on their own.

[0053] Compelling evidence has been demonstrated through laboratory and field trials of the agricultural application of the herbicidal formulation of the present invention, which combines a herbicide such as glyphosate with the composite microsphere described herein, showing enhanced herbicidal activity. These trials reveal that the chitosan:gum:TPP particles of the present invention are non-toxic and exhibit increased herbicidal activity of over 10%. Furthermore, they maintain the same herbicidal effect when applied at only 48% of the standard dose.

[0054] Thus, the present invention addresses the problems posed by the prior art by reducing agrochemical use with a biocompatible and biodegradable solution. It allows for the application of a lower dose of active ingredients, which are typically high-risk, toxic, and expensive commercial components.

[0055] Thus, the present invention addresses the problem identified in the prior art by reducing the use of agrochemicals through a harmless, safe, biocompatible, and biodegradable solution.

[0056] Brief Description of the Invention

[0057] The present invention relates to a composite microsphere designed to enhance the efficacy of herbicides, preferably for agronomic applications. The composite microsphere comprises chitosan, with an average molecular weight in the range of 190,000 to 310,000 g / mol, and a crosslinking agent, in the absence of a herbicidal active ingredient. The composite microsphere of the invention features an adjustable surface electric charge (zeta potential) in an aqueous medium, which is regulated by the concentration of the crosslinking agent. The chitosan-to-crosslinking agent mass ratio is 1:X, where X varies within a range of 0.08 to 1.5, and this ratio defines the zeta potential of the microsphere in an aqueous solution. Preferably, X varies within a range of 0.08 to 0.4, and most preferably, X is 0.08.

[0058] In a preferred embodiment, the crosslinking agent is selected from the group consisting of sodium tripolyphosphate (TPP), sodium hexametaphosphate, and their derivatives; most preferably, sodium tripolyphosphate (TPP). In another preferred embodiment, the present invention further comprises a dispersant that improves the redispersion of the microsphere when combined with a herbicide aqueous solution. The dispersant is selected from the group consisting of gum arabic, xanthan gum, guar gum, carrageenan, pectin, alginates, dextrins, cyclodextrins, maltodextrins, tragacanth gum, mesquite gum, polyethylene glycol, and their mixtures; preferably gum arabic. The chitosan-to-dispersant mass ratio is Y:l, where Y ranges from 5 to 20, meaning that the gum is incorporated into the microsphere of the invention at up to 20% of the chitosan mass.

[0059] A primary object of the present invention is a herbicidal formulation comprising a herbicidal active ingredient and the composite microspheres of the invention, which enhance the herbicidal efficacy on crops. This formulation is characterized in that the composite microsphere is added at a concentration ranging from 2% to 200% w / w relative to the herbicidal active ingredient, preferably between 2% and 100%.

[0060] In preferred embodiments of the present invention, the herbicidal active ingredient is selected from the group consisting of glyphosate, imazapyr, imazethapyr, atrazine, simazine, diuron, pendimethalin, trifluralin, fluazifop-p-butyl, quizalofop-p-ethyl, dicamba, paraquat, glufosinate, clethodim, haloxyfop, fomesafen, 2,4- dichlorophenoxyacetic acid, thiencarbazone-methyl, pyroxasulfone, ammonium glufosinate, imazamox, imazaquin, flurochloridone, clethodim, haloxyfop, lactofen, diflufenican, sulfentrazone, S-metolachlor, flumioxazin, pyroxasulfone, MCPA, saflufenacil, bromoxynil, their derivatives, and mixtures thereof; preferably glyphosate or glufosinate, between many others.

[0061] In a preferred embodiment of the herbicidal formulation, the composite microsphere comprises chitosan with an average molecular weight in the range of 190,000 to 310,000 g / mol, a dispersant, and a crosslinking agent, wherein the chitosan-to-crosslinking agent mass ratio is 1:X, where X varies within a range of 0.08 to 1.5 and defines the zeta potential in an aqueous solution of the microspheres. In one embodiment of the invention, the composite microspheres comprise chitosan, gum arabic, and sodium tripolyphosphate, in the absence of agrochemical actives. The composite microspheres of the invention exhibit a zeta potential in aqueous solution ranging from +30 mV to -30 mV, adjustable by the concentration of the crosslinking agent. At low crosslinking agent concentrations, such as 8% of the chitosan mass, the potential is approximately +30 mV. As the concentration of the crosslinking agent increases, the zeta potential decreases, reaching zero when the crosslinking agent accounts for approximately 40% of the chitosan mass. With further increases in crosslinking agent concentration, the zeta potential becomes more negative, reaching values around -30 mV at concentrations exceeding 80% of the chitosan mass.

[0062] In a preferred embodiment, the herbicidal formulation of the invention further comprises a co-adjuvant selected from the group consisting of adhesives, surfactants, stabilizers, antifoaming agents, pH buffers, chelating agents, their derivatives, and combinations thereof.

[0063] In a preferred embodiment of the herbicidal formulation, the herbicidal active ingredient is present at a concentration equal to or less than the recommended field application rate. This means that the herbicidal formulation of the invention enables a reduction in the amount of herbicide used in agriculture.

[0064] A particular feature of the herbicidal formulation is that the concentration of the composite microspheres ranges from 5% to 200% w / w relative to the herbicide, more preferably from 5% to 100%, and most preferably from 5% to 10%. The mixture of herbicide and composite microspheres is prepared in the field before application.

[0065] In one preferred embodiment, the herbicidal formulation of the present invention is an aqueous composition comprising a herbicidal active ingredient and composite microspheres packaged in containers, as the components are pre-mixed at the manufacturing facility and packaged together for sale and distribution.

[0066] In an alternative preferred embodiment, the herbicidal formulation of the present invention is an aqueous composition containing the herbicidal active ingredient and the composite microspheres in powder form, to be mixed before use.

[0067] Another object of the present invention is a method for preparing the herbicidal formulation of the invention, comprising the following steps: a) preparing an aqueous solution of the herbicidal active ingredient at the desired concentration; b) adding the dry composite microspheres at a concentration of up to 100% w / w relative to the herbicidal active ingredient.

[0068] In one embodiment, a co-adjuvant is also added to the formulation selected from the group consisting of an adhesive, a surfactant, a stabilizer, an antifoaming agent, a pH buffer, a sequestrant, their derivatives, and combinations thereof.

[0069] Another object of the present invention is a method for preventing or reducing the growth of undesired vegetation, which comprises applying the herbicidal formulation of the invention to a crop or soil prior to sowing. The crop is selected from the group consisting of cereals, oilseeds, forestry, fruit trees, trees, ornamentals, vegetables, wheat, barley, rye, triticale, oats, maize, sunflower, rice, soybean, peas, broad beans, beans, peanuts, rapeseed, kala, cotton, lentils, potatoes, sugar beets, sugarcane, grass, sorghum, canola, flax, legumes, tomatoes, peppers, squash, lettuce, chard, carrots, beets, radishes, apple trees, pear trees, plum trees, bananas, mangoes, citrus trees, walnut trees, almond trees, coffee, cocoa, vines, orange trees, lemon trees, grasses, alfalfa, forage crops, red clover, fescue, ryegrass, camellia, moringa, miscanthus, sweet corn, chamomile, lavender, mint, aloe vera, stevia, pepper, nutmeg, saffron, hemp, agave, jute, and moringa, between many others. The herbicidal formulation is applied at a concentration of herbicidal active ingredient lower than that indicated on the label and enhances herbicidal action by up to 70% compared to the herbicide's action without microspheres.

[0070] In conclusion, the present invention, in a preferred embodiment, comprises a herbicidal formulation that enhances the efficacy of a herbicidal active ingredient by at least 10%. This formulation comprises a herbicidal active ingredient and a composite microsphere, wherein the microsphere comprises chitosan and a crosslinking agent. The microsphere contains chitosan with an average molecular weight in the range of 190,000 to 310,000 g / mol, and the chitosan-to-crosslinking agent mass ratio is 1:X, where X varies between 0.08 and 1.5. The concentration of the composite microsphere ranges from 2% to 100% w / w relative to the herbicide in an aqueous phase. The composite microsphere further comprises a dispersing agent selected from the group consisting of gum arabic, xanthan gum, guar gum, carrageenan, pectin, alginates, dextrins, cyclodextrins, maltodextrins, tragacanth gum, mesquite gum, polyethylene glycol, and their mixtures; preferably gums. The composite microsphere has an adjustable surface electric charge or zeta potential modulated by the concentration of the crosslinking agent, with X ranging from 0.08 to 0.8. The crosslinking agent is TPP.

[0071] In a preferred embodiment of this herbicidal formulation, the herbicidal active ingredient is in an aqueous composition within a container, and the composite microsphere is in powder form, configured to be added to the container.

[0072] Another object of the present invention is a process for obtaining the composite microsphere, comprising the following steps: mixing acetic acid, chitosan, and water, and stirring; adding the dispersing agent and stirring; adding the solution of the crosslinking agent drop by drop; allowing the mixture to rest; and drying the resulting microspheres. Preferably, the process comprises the following steps: a. Dissolving chitosan in 1% v / v acetic acid under constant stirring at a temperature of

[0073] 20°C until no lumps are observed; b. Adding the dispersing agent and stirring until fully dissolved, achieving a chitosan-to- dispersing agent mass ratio of Y:l, where Y may vary from 5 to 20; c. Adding drop by drop to the mixture a solution of the crosslinking agent in a chitosan- to-crosslinking agent mass ratio of 1:X, where X may vary from 0.08 to 1.2, in the absence of a herbicidal active ingredient; d. Adjusting the pH to approximately 6.5; e. Drying the composite microspheres.

[0074] In a preferred embodiment, step e) involves lyophilization using a 220 V freeze dryer at -45°C and 10 Pa of pressure, followed by grinding with a mortar and sieving through a 100-mesh filter. Alternatively, step e) involves spray drying at an inlet temperature of 200°C.

[0075] Finally, a preferred embodiment of the process for obtaining the fungicidal formulation of the invention comprises the following steps: a. Preparing an aqueous fungicide solution; b. Adding dry composite microspheres to the aqueous fungicide solution.

[0076] Description of the Figures

[0077] Figure 1. Scanning Electron Microscopy (SEM) images of MCS 0.08-0.1, MCS 0.3-0.1, MCS 0.5-0.1, MCS 0.8-0.1, MCS 1.2-0.1, and MCS 1.5-0.1.

[0078] Figure 2. Transmission Electron Microscopy (TEM) images of MCS 0.08-0.1, MCS 0.3-0.1, MCS 0.8-0.1, and MCS 1.2-0.1.

[0079] Figure 3. Herbicidal activity assay of Glyphosate combined with MCS 0.8-0.1. Representative plants are shown for each treatment.

[0080] Figure 4. Herbicidal activity assay of Glyphosate combined with MCS 0.8-0.1. The bars represent the average fresh foliar biomass in grams for each treatment.

[0081] Figure 5. Herbicidal activity assay of Glyphosate combined with 5% MCS 0.08-0.1 and MCS 0.8-0.1 on foliar biomass in wheat plants grown for 14 days on plates. Plants were treated after 8 days of growth, and observations were made 6 days after application (6 DAA). Bar height represents the mean of each group, and error bars indicate standard error (n = 4).

[0082] Figure 6. Field trials of herbicidal activity of Glyphosate combined with MCS 0.8-0.1 on Cynodon dactylon (bermudagrass) weeds. Bar height represents the mean of each group (n = 3). Solid black bars correspond to measurements taken 7 days after application (7 DAA), and white bars correspond to measurements taken 14 days after application (14 DAA).

[0083] Figure 7. Field trials of herbicidal activity of Glyphosate combined with MCS 0.8-0.1 on Brassica rapa "elongated" weeds (Nobolza elongada). Bar height represents the mean of each group (n = 3). Solid black bars correspond to measurements taken 7 days after application (7 DAA), and white bars correspond to measurements taken 14 days after application (14 DAA).

[0084] Figure 8. Photograph showing clogged herbicide sprayer filters due to microspheres of the invention without gum. Figure 9. Scanning Electron Microscopy (SEM) innages of MCS 0.8 without gum (left) and with gum arable (right).

[0085] Detailed Description of the Invention

[0086] For the purposes of the present invention, the term "agrochemical" refers to any natural or synthetic substance or mixture of substances used to prevent, eliminate, and / or control pests, diseases, or weeds in agricultural activity, and / or to preserve or enhance soil fertility, and / or the quality and / or yield of crops.

[0087] In this document, such agrochemicals include fertilizers, plant growth regulators, biostimulants, pesticides, pest control agents, herbicides, fungicides, nematicides, insecticides, rodenticides, agricultural biocompounds, and phytosanitary products.

[0088] For the purposes of this invention, the term "adjuvant" or "co-adjuvant" or "agricultural adjuvant" refers to products of natural or synthetic origin that enhance or facilitate the action of an agrochemical (phytosanitary product, fertilizer, or biostimulant) by modifying certain characteristics of the solution and whose properties improve the activity of the agrochemical.

[0089] In this document, the term "herbicide" refers to a chemical or biological product, natural, artificial, or modified, used to control and / or eliminate undesired plants.

[0090] For the purposes of this document, the terms "undesired vegetation," "unwanted vegetation," "weed," "invasive plant," and "herbaceous plants" are used interchangeably.

[0091] In this document, the term "microsphere" refers to micrometric-sized particles that may include microparticles, microspherules, nanoparticles, or nanospheres, with various shapes, where the size of such microspheres is referred to as particle diameter or size.

[0092] For the purposes of this invention, the term "MCS" refers to the composite microspheres of the invention made from chitosan:gum:TPP.

[0093] The term "MCS0.08" refers to the composite microspheres of the invention made with chitosan:gum:TPP in a mass ratio of 1:0.1:0.08. The term "MCS0.3" refers to the composite microspheres of the invention made with chitosan:gum:TPP in a mass ratio of l:0.1:0.3.

[0094] The term "MCS0.5" refers to the composite microspheres of the invention made with chitosan:gum:TPP in a mass ratio of l:0.1:0.5.

[0095] The term "MCS0.8" refers to the composite microspheres of the invention made with chitosan:gum:TPP in a mass ratio of l:0.1:0.8.

[0096] The term "MCS1.2" refers to the composite microspheres of the invention made with chitosan:gum:TPP in a mass ratio of l:0.1:1.2.

[0097] In this document, "e.a. ha-1" refers to the amount of glyphosate acid equivalent applied per hectare of land. This is a commonly used unit for specifying doses in agricultural applications.

[0098] One of the significant challenges in agricultural activity is the excessive use of agrochemicals, which results in adverse effects on human health, the environment, and the emergence of resistant crops. The present invention provides a solution to the problems identified in the prior art.

[0099] This invention provides crosslinked composite microspheres based on chitosan, preferably spherical in shape, with a size ranging from 100 to 200 nanometers. These microspheres can aggregate into micrometric-sized structures and, when combined at low concentrations with a herbicide, exhibit a synergistic effect that enhances the herbicide's efficacy. This allows for the application of reduced doses while maintaining the same herbicidal effect.

[0100] A composite microsphere, the principal object of the present invention, reduces the use of agrochemicals in agriculture. It comprises chitosan, a crosslinking agent, and preferably a dispersing agent, wherein the microsphere exhibits a positive, neutral, or negative surface charge. This surface charge is adjustable through the concentration of the crosslinking agent within the microsphere.

[0101] The chitosan used in the present invention preferably has a degree of deacetylation of at least 90%. In preferred embodiments of the present invention, the crosslinking agent comprises TPP (tripolyphosphate) in a chitosan / crosslinking agent / dispersing agent weight ratio of 100 / 8 / 15 and 100 / 80 / 10.

[0102] The present invention provides hierarchical composite microspheres manufactured from chitosan with precisely defined and adjustable chemical composition, size, shape, and charge distribution. These microspheres act as co-formulators to reduce the dosage of herbicides in an agrochemical formulation. The microspheres exhibit a well-defined microstructure with an adjustable surface charge that can be positive, neutral, or negative.

[0103] The amount of crosslinking agent, preferably sodium tripolyphosphate, is strategically defined based on the desired charge distribution of the particle, ranging from 3% to 150% w / w relative to chitosan. The dispersing agent, generally a natural polymer, endows the microspheres with rapid dispersibility and enables their use across a broad range of salinity, pH, and temperature conditions. The dispersing agent is selected from the group consisting of gum arabic (and its derivatives), xanthan gum (and its derivatives), guar gum (and its derivatives), poly(ethylene glycol)-poly(propylene glycol) triblock copolymers, polyacrylamide derivatives, microcrystalline cellulose (and its derivatives), carboxymethylcellulose (and its derivatives). The dispersing agent is present at a concentration of up to 20% w / w relative to chitosan.

[0104] The herbicidal actives that can be incorporated into the present invention include, but are not limited to: glyphosate, imazapyr, imazethapyr, atrazine, simazine, diuron, pendimethalin, trifluralin, fluazifop-p-butyl, quizalofop-p-ethyl, dicamba, paraquat, glufosinate, clethodim, haloxyfop, fomesafen, 2,4-dichlorophenoxyacetic acid, thiencarbazone-methyl, pyroxasulfone, ammonium glufosinate, imazamox, imazaquin, flurochloridone, clethodim, haloxyfop, lactofen, diflufenican, sulfentrazone, S- metolachlor, flumioxazin, pyroxasulfone, MCPA, saflufenacil, bromoxynil, their derivatives, and mixtures thereof.

[0105] The composite microspheres of the present invention differ from those identified in the prior art by their adjustable physicochemical properties, particularly their zeta potential and charge distribution. The inventors have been able to adjust and define the zeta potential of the particles in the present invention based on the amount of TPP incorporated during synthesis.

[0106] Particles were developed with TPP content ranging from 0.03 to 1.5 per unit of chitosan, achieving an adjustable zeta potential in the range of +30 mV to -30 mV. This allows for the synthesis of particles with optimal physicochemical properties to be combined with a wide spectrum of agrochemicals.

[0107] In one embodiment of the present invention, the composition of the microspheres comprises a positive surface charge, and the amount of sodium tripolyphosphate is less than 40% w / w relative to chitosan.

[0108] In another embodiment of the present invention, the composition of the microspheres comprises a negative surface charge, and the amount of sodium tripolyphosphate is at least 50% w / w relative to chitosan.

[0109] In preferred embodiments, the microspheres of the present invention comprise medium molecular weight chitosan, in the range of 190,000 to 310,000 g / mol, with a degree of deacetylation greater than 90%.

[0110] The present invention also provides an agrochemical formulation, particularly a herbicidal formulation, containing the composite microspheres of the invention, combined with a herbicide. These composite microspheres, when incorporated at low concentrations, show no herbicidal activity per se but enhance the herbicidal effect, allowing the application of reduced doses— up to 52% less herbicide— while maintaining similar efficacy to the recommended doses of the standalone herbicide.

[0111] Compelling and surprising evidence has been observed that formulations of glyphosate combined with the composite microspheres of the invention, which constitute the formulation of the invention, exhibit herbicidal activity that is 15-21% higher than glyphosate applied alone in laboratory plant trials. It is noteworthy that the concentration of the composite microspheres used in the invention can range from 2% to 100% w / w relative to the herbicide's concentration. Excellent results have been achieved even within a narrow range of 5% to 10%. Field trials have demonstrated that the agrochemical formulation of the present invention, incorporating the composite microspheres at a ratio of 5% w / w relative to the active ingredient, enables a reduction in the application dose of glyphosate by up to 52%, while maintaining herbicidal efficacy similar to glyphosate applied alone at the recommended dose.

[0112] In this document, the formula for calculating the enhancement of a herbicidal active ingredient with the composite microspheres of the present invention is defined as follows:

[0113] Enhancement(%)=((Effect of the invention-Effect of herbicide alone)Effect of herbicide alone)xl00

[0114] In this case, the effect can be measured by herbicidal activity. However, it can also be assessed using other variables, such as yield, when the yield depends on the control exerted by the agrochemical in question.

[0115] Unlike similar particles identified in the prior art, which are characterized by encapsulating the agrochemical within the particle during the synthesis process, the composite microspheres of the present invention are combined with the active ingredient after synthesis, preferably just before field use. The most notable advantages include compatibility with a wide variety of agrochemicals and the enhancement of the agrochemical's effect when formulated together. By boosting efficiency, the invention allows for the recommendation of reduced application doses compared to commercially recommended doses for the product.

[0116] The fact that the composite microspheres of the invention are dried and combined at the time of application ensures a stable product under regular environmental conditions, with an extended shelf life. Being biodegradable, these composite microspheres do not accumulate and thus do not cause environmental damage. No cytotoxicity indices have been detected in human cells.

[0117] The present invention provides a manufacturing process for these composite microspheres, involving a physical gelation process with a strategically designed synthesis using three essential components: chitosan, a crosslinking agent, and a dispersing agent. The production process has been successfully scaled to a reaction volume of 20 liters. Throughout the process, the pH is maintained at 4.5 by adjusting with acetic acid, while stirring and temperature are kept constant at 130 rpm and 20°C, respectively. At the end of the process, the pH is adjusted to 6.5, and the microspheres are subjected to natural decantation. Subsequently, they are dried using spray drying and sieved through a 100- mesh filter.

[0118] The present invention also provides a synthesis method for obtaining the composite microsphere of the invention, comprising the following steps: a. Dissolving chitosan in 1% v / v acetic acid under constant stirring at a temperature of 20°C until no lumps are observed; b. Adding gum arable and stirring until fully dissolved, achieving a chitosan-to-gum arabic mass ratio of Y:l, where Y may vary from 5 to 20; c. Adding drop by drop to the mixture a TPP solution until reaching the desired TPP concentration, achieving a chitosan-to-TPP mass ratio of 1:X, where X may vary from 0.03 to 1.2, in the absence of a herbicidal active ingredient; d. Adjusting the pH to approximately 6.5; e. Drying the particles.

[0119] In a preferred embodiment of the invention:

[0120] • Step e) involves lyophilization using a 220 V freeze dryer at -45°C and 10 Pa of pressure, followed by grinding with a mortar and sieving through a 100-mesh filter; or

[0121] • Step e) involves spray drying at an inlet temperature of 200°C.

[0122] The present invention further provides a method for preventing the growth of undesired vegetation, comprising applying the herbicidal formulation of the invention to a crop or soil prior to sowing, wherein the crop is selected from the group consisting of cereals, trees, oilseeds, vegetables, fruits, ornamentals, chard (Beta vulgaris), peppers (Capsicum spp.), garlic and onions (Allium spp.), celery (Apium graveolens), eggplant (Solanum melongena), pumpkin (Cucurbita moschata), chayote (Sechium edule), cabbage (Brassica oleracea), spinach (Spinacia oleracea), green beans (Phaseolus vulgaris), lettuce (Lactuca sativa), corn (Zea mays), peanut (Arachis hypogaea), tomato (Solanum lycopersicum), cucumber (Cucumis sativus), okra (Hibiscus esculentus), radish (Raphanus sativus), beet (Beta vulgaris), carrot (Daucus carota), avocado (Persea americana), sugar apple (Annona squamosa), star apple (Chrysophyllum cainito), canistel (Pouteria campechiana), cherry (Malpighia punicifolia), custard apple (Annona reticulata), plum (Spondias dulcis), coconut (Cocos nucifera), papaya (Carica papaya), soursop (Annona muricata), guava (Psidium guajava), pomegranate (Punica granatum), lime (Citrus aurantifolia), lemon (Citrus limonum), apple (Malus domestica), mamey sapote (Calocarpum mammosum), mammee apple (Mammea americana), Spanish lime (Melicoccus bijugatus), mandarin (Citrus reticulata), mango (Mangifera indica), passion fruit (Passiflora laurifolia), watermelon (Citrullus lanatus), bitter orange (Citrus aurantium), sweet orange (Citrus sinensis), pear (Pyrus communis), pineapple (Ananas comosus), banana (Musa paradisiaca), plantain (Musa balbisiana), tamarind (Tamarindus indica), grapefruit (Citrus paradisi), pomelo (Citrus grandis), grape (Vitis vinifera), rice (Oryza sativa), coffee (Coffea arabica), sugarcane (Saccharum officinarum), cotton (Gossypium hirsutum), gherkin (Melothria guadalupensis), sweet potato (Ipomoea batatas), potato (Solanum tuberosum), Andean potato (Solanum phureja), golden berry (Physalis peruviana), cassava (Manihot esculenta), soybean (Glycine max), strawberries (Fragaria spp.), mulberries (Morusspp.), blackberries (Rubus spp.), palms of the family Aracaceae, oil palm (Elaeis guineensis), cacao (Theobroma cacao), tree tomato (Solanum betaceum), lulo (Solanum quitoense), chickpeas (Cicer arietinum), among other species.

[0123] The present invention is further described through the following examples, which are not to be considered as limiting the scope of the invention. The invention is not restricted by the illustrative examples provided below.

[0124] Examples

[0125] Example 1: Method for Synthesis and Production of Composite Microspheres of Chitosan: Gum Arabic: Tripolyphosphate (TPP) at Laboratory Scale The following materials were used for the synthesis of the composite microspheres: Chitosan (degree of deacetylation 95%, Mw 275,000), gum arabic (Mw 70,000), TPP, 1% acetic acid, 1 M sodium hydroxide, distilled water, 600 ml_ and 250 mL beakers, DragonLab mechanical stirrer, peristaltic pump, pH meter, and 100- and 450-mesh sieves.

[0126] Composite microspheres of chitosan, gum arabic, and TPP were obtained using the ionic gelation method in 0.6 and 20-lite r reactors, following this procedure: a) For a 600 mL reactor:

[0127] • Prepare a solution of 200 mL of 1% w / v acetic acid (10 mL / L) and add 4 g of chitosan under constant stirring at 1300 RPM with the DragonLab mechanical stirrer. Stir until the solution is completely dissolved, achieving a uniform, clear color without visible lumps, for 10 minutes at room temperature.

[0128] • Add 0.4 g of gum arabic (10% of chitosan weight). Stir for 10 minutes until fully dissolved.

[0129] • Prepare a 1.6% w / v aqueous solution of sodium tripolyphosphate.

[0130] • Using a peristaltic pump, begin adding the TPP solution drop by drop into the beaker until 200 mL of TPP is added to the reactor. Maintain stirring for 30 minutes after the addition is complete. Keep the reactor stirring at 1300 RPM.

[0131] • Measure and adjust the pH to 6.5 using 1 M sodium hydroxide.

[0132] • Centrifuge for 10 minutes at 1200 RPM and discard the supernatant.

[0133] • Freeze the composite microspheres in the freezer for 24 hours.

[0134] • Dry the composite microspheres using a PeetLab BK-FD10PT freeze dryer at 220 V, -18°C, and 10 Pa pressure.

[0135] • Grind and sieve the composite microspheres first through a 100-mesh sieve, then through a 450-mesh sieve. b) For a 20-liter reactor:

[0136] The same procedure was followed, but the material quantities were adjusted to the dimensions of the 20-liter reactor equipped with a DragonLab 4-paddle mechanical stirrer (Figmay), and the drying step was modified to spray drying as follows:

[0137] Briefly, in a 20-liter borosilicate glass reactor (Figmay), equipped with a 4-flat-paddle stirrer without heating, 5 L of 1% acetic acid (10 mL / L) and 100 g of chitosan (degree of deacetylation 95%, Mw 275,000) were added and stirred at 130 RPM for 10 minutes. The reactor temperature was maintained at 21 ± 2°C. A complete dissolution was achieved, with the solution appearing uniform, clear, and without visible lumps. Then, 15 g of gum arabic was added as a dispersing agent, and the mixture was stirred for 10 minutes.

[0138] In a second 10-liter reactor, a TPP solution (sodium tripolyphosphate) of 16 g / L concentration was prepared by adding 5 L of distilled water and 80 g of TPP and stirring until completely dissolved, for 5 minutes at room temperature.

[0139] The 20-liter reactor containing the chitosan solution was stirred at 130 RPM, and using two peristaltic pumps, the TPP solution was added drop by drop at a rate of 2 L / h. Once all the TPP was added, stirring was maintained for 30 minutes. The pH was then adjusted to 6.5 using 1 M NaOH.

[0140] The reactor was then emptied into a 20-liter container and left to rest overnight (12 hours) at room temperature. The supernatant liquid was discarded. Using a JISL LSD-48 benchtop spray dryer with a drying capacity of 1 L / h, the particles were dried at an inlet temperature of 200°C with a feed rate of 60% (600 mL / h). The particles were kept in suspension through stirring during the drying process.

[0141] Example 2: Production of Composite Microspheres of Chitosan with Variable Proportions of Gum Arabic and Tripolyphosphate

[0142] Composite microspheres were obtained according to the procedure described in Example 1, adjusting the amounts of gum arabic relative to the grams of chitosan and modifying the volumes of TPP added dropwise relative to the amount of chitosan. This produced composite microspheres with different mass ratios of Chitosan:Gum Arabi TPP. The following composite microspheres were prepared with mass ratios as shown in Table 1.

[0143] Table 1. Composite microspheres with mass ratios of gum arabic and TPP per one part of chitosan. Example 3: Production of Composite Microspheres of Chitosan with Sodium Hexametaphosphate (HMP) as Crosslinking Agent and Xanthan Gum and Polyethylene Glycol as Dispersing Agents

[0144] In a 20 L borosilicate glass reactor (brand Fignnay), made of borosilicate glass 3.3, with a 4-flat-paddle stirrer without heating, 5 L of a 15% citric acid solution (10 mL / L) and 100 g of chitosan (degree of deacetylation 92%, Mw 210,000) were added. The mixture was stirred at 150 RPM for 10 minutes, maintaining the reactor temperature at 21 ± 2 °C. A complete dissolution was achieved, resulting in a uniform, clear solution without visible lumps. Subsequently, 5 g of xanthan gum and 10 g of polyethylene glycol were added as dispersing agents, and the mixture was stirred for 10 minutes.

[0145] In a second 10 L reactor, a sodium hexametaphosphate (HMP) solution with a concentration of 25 g / L was prepared by adding 5 L of distilled water and 125 g of HMP, stirring until fully dissolved for 10 minutes at room temperature.

[0146] The 20 L reactor containing the chitosan solution was stirred at 150 RPM, and HMP was added drop by drop using a peristaltic pump at a rate of 0.5 L / h (10 hours), with stirring maintained for an additional 60 minutes after all the HMP had been added.

[0147] The reactor contents were then transferred to a 20 L container and allowed to rest overnight (12 hours) at room temperature. The supernatant was poured into another container, ensuring no material was lost during the process, and then centrifuged for 15 minutes at 1500 RPM using the "Rolco MOD.CM-2036" laboratory centrifuge. The samples were frozen for 24 hours at -24 °C. Using a benchtop multiple-collector laboratory freeze dryer, the samples were lyophilized for 72 hours. Finally, the samples were ground and sieved through a 450-mesh sieve.

[0148] Example 4: Determination of the Size and Morphology of Chitosan:Gum Arabic:Tripolyphosphate Composite Microspheres A comprehensive physicochemical characterization of the composite microspheres obtained according to the procedure in Example 1 and the chemical composition described in Example 2 was carried out to define their structure and properties.

[0149] Three techniques were used to study and define the size and morphology of the composite microspheres:

[0150] 1. Dynamic Light Scattering (DLS).

[0151] Three measurements, each lasting 60 seconds with 10-second intervals, were performed at 25 °C using a Malvern Zetasizer Nano S90. The Brownian motion of the composite microspheres in suspension was analyzed. The light scattered by the composite microspheres provides information about the diffusion coefficient, from which the hydrodynamic radius and the size distribution of the material can be determined.

[0152] Table 2. Physicochemical Parameters of Composite Microspheres (MCS) for Different TPP / CS Ratios, All Samples Containing 10% Gum Arabic.

[0153] 2. Transmission Electron Microscopy (TEM). This technique was performed using a JEOL JEM 2100 microscope with 200 kV and a B6La filament. Photographs were taken at magnifications of 50,000x or higher to analyze particle morphology and size.

[0154] 3. Scanning Electron Microscopy (SEM).

[0155] (FESEM) ZEISS Crossbeam 350 model. The sample was prepared with a Cr or Au coating. Images were taken at magnifications of 30x, 500x, 1500x, 6000x, 20,000x, and 40,000x. General appearance, particle size, and morphology were analyzed.

[0156] Characterization of particle sizes using DLS allows us to conclude that there are populations with different size distributions (Table 2). In many cases, up to three different hydrodynamic diameters can be observed, approximately in the ranges of 200- 300 nm, 600-1000 nm, and 2000-5000 nm.

[0157] When more precise techniques such as SEM and TEM are used to study particle morphology and size, it is observed that spherical particles with diameters of approximately 20-30 nm interact with each other to form larger aggregates, some amorphous and others spherical (Figures 1 and 2). These aggregates range from a few hundred nanometers to several micrometers, thus confirming the size distribution results obtained through DLS in the synthesis process.

[0158] Example 5: Determination of the Charge Distribution of Chitosan:Gum ArabicTripolyphosphate Composite Microspheres

[0159] To determine the Zeta Potential, three measurements were performed, each lasting 100 seconds with 10-second intervals, at 25°C, using a Horiba-SZ 100 device. The measurements were averaged, and the results are reported. The obtained results are shown in Table 2 for the different composite microspheres (MCS).

[0160] From the characterization results, it can be observed that composite microspheres with gum (10% w / w relative to chitosan) and a TPP / CS ratio between 0.08 and 0.3 inclusive (MCS0.08-0.1 and MCS0.3-0.1) exhibit positive Zeta Potential values, while those with a TPP / CS ratio between 0.5 and 1.5 (MCS0.5-0.1, MCS0.8-0.1, MCS1.2-0.1, and MCS1.5- 0.1) show negative values (Table 2). The inflection point appears to correspond to a TPP / CS ratio of 0.4, where the particle surface charge is near neutrality, as the Zeta Potential value is around 0, considering the measurement error (between ±5 and ±10 mV). These results demonstrate that increasing TPP concentrations increase the negative Zeta Potential of the particle, enabling the design of composite microspheres with the desired Zeta Potential based on TPP addition.

[0161] Example 6: Study of the Interaction Between Composite Microspheres and Glyphosate

[0162] Studies were conducted to evaluate the interaction capacity of the composite microspheres with different agrochemicals in aqueous solution and to determine the amount of active ingredient associated with the material.

[0163] Nuclear Magnetic Resonance (NMR).

[0164] Following the technique described in "Probing interactions by means of pulsed field gradient nuclear magnetic resonance spectroscopy" S. Cozzolino, et al. Magn. Reson. Chem. 2008, 46, S16-S23 DOI 10.1002 / mrc.2345, diffusion coefficients were calculated using the NMR technique. These coefficients can be used to determine the physicochemical association percentages of different analytes with composite microspheres (MCS).

[0165] The analyte and the particle have their own diffusion coefficients in a free state, reflecting their molecular weight and shape. However, when a complex forms, and the analyte and particle are strongly bound, they should have the same diffusion coefficient as they diffuse as a single molecular entity. In the case of weak or insignificant association, the diffusion coefficients of the analyte and particle remain unchanged. For other cases, assuming rapid exchange on the NMR timescale, the observed (measured) diffusion coefficient is a weighted average of the free and bound diffusion coefficients and can thus be used to calculate the bound fraction.

[0166] Diffusion coefficients were determined using a PFGSTE26 sequence on a Spinsolve80 Ultra from Magritek GmbH, with membrane strips placed in a 5 mm tube and operated at 26°C. For the study, MCSO.8-0.1 (with gum), MCS0.8-0 (without gum) samples from Example 2, and glyphosate were used to study the percentage of association between them. Components were added to a laboratory test tube with distilled water and manually stirred for 1 minute. Suspensions of the individual components and mixtures of each microparticle with glyphosate in a 1:5 mass ratio of glyphosate to microspheres were prepared.

[0167] In a second study, mixtures of glyphosate with the microspheres MCSO.08-0.1, MCSO.8- 0.1, and MCS1.2-0.1 from Example 2 were prepared in a 10:1 mass ratio of glyphosate to microspheres.

[0168] Table 3. Association Percentages of Glyphosate (Negative Charge) with Composite Microspheres(MCS). Percentages were obtained from the diffusion coefficients determined by the NMR technique.

[0169] This table highlights that the highest rate of association occurs with composite microspheres exhibiting a positive Zeta Potential, due to the negative charge of glyphosate. It also shows that the presence of gum reduces the association capacity between glyphosate and the microspheres, which acts as a limiting factor in the amount of gum that can be used.

[0170] Example 7: Herbicidal Activity Assay of Glyphosate Combined with Composite Microspheres (MCS) in Lettuce Plants (Lactuca sativa) To evaluate whether chitosan:gum arabic:TPP composite microspheres have a herbicidal potentiating effect when formulated with glyphosate, an assay was conducted applying different concentrations of commercial herbicide combined with composite microspheres to lettuce plants under the following experimental protocol:

[0171] Lettuce plants (Lactuca sativa var. mantecosas) were grown for two weeks in 7 x 9.5 x 4 cm pots, containing 25 g of commercial substrate Grow Mix Multipro per pot, under controlled conditions of light (16:8 photoperiod), temperature (25°C), and humidity (60% relative humidity). After 14 days of growth, the plants were divided into five experimental groups, each containing 10 plants (5 x 10), and treated via foliar spray with the treatments described in Table 4.

[0172] The application volume was 0.2 mL per plant. Both the quality and calibration of the spray were controlled using a hydrosensitive card. Spraying was performed with a plastic sprayer (PVC-EPA).

[0173] Table 4. Treatments to Evaluate the Herbicidal Efficacy of Different Concentrations of Glyphosate Combined with MCS 0.8-0.1.

[0174] To prepare the application solutions, an initial aqueous solution of 5 mg / ml of MCS was prepared. The required amount of MCS solution for glyphosate herbicide was calculated to maintain a weight ratio of 5% composite microspheres of the invention (MCS) over the acid equivalents (e.a.) of the active glyphosate present in the mixture at the specified ratios of composite microspheres. These ratios were expressed as 5% (w / e.a. glyphosate) or 10% (w / e.a. glyphosate) depending on the treatments. The application solutions were prepared in 15 or 50 ml Falcon tubes (Bio-Plast), depending on the required volume. Each formulation was agitated for 2 hours in an orbital shaker (Vicking, model M23) until the time of application.

[0175] Ten days after the application of each treatment (10 DDA), 10 plants from each experimental group were collected, and the fresh leaf biomass was quantified using an analytical balance (OHAUS, model Traveler TA302).

[0176] Figure 3 shows representative plants from each treatment, while Figure 4 presents the average weight (with standard deviation) of 10 tested plants for each treatment.

[0177] The application of the full herbicide dose (1 L ha-1glyphosate) resulted in a 62% reduction in fresh biomass compared to the water control (H2O). Plants treated with 0.5 L ha"1glyphosate showed a 30% reduction in biomass compared to the same control (Figure 4). In contrast, plants treated with 0.5 L ha"1glyphosate + 5% MCS 0.8-0.1 exhibited a 51% reduction in biomass relative to the control. This indicates that combining MCS 0.8-0.1 with the reduced glyphosate dose enhanced the herbicidal efficacy by 21% compared to the treatment with the same herbicide concentration without MCS 0.8-0.1.

[0178] In percentage terms, the use of the invention's formulation with 5% chitosan composite microspheres (MCS) enhanced glyphosate's herbicidal power by 70%. The percentage reduction in biomass increased from 30% without MCS 0.8-0.1 to 51% with MCS 0.8-0.1. The formula for calculating the enhancement is:

[0179] Enhancement (%)=((51-30 / 30)xl00

[0180] This document refers to this indicator of increased herbicidal efficacy as "enhancement." In this assay, the use of MCS 0.8-0.1 enhanced the herbicidal formulation by 70% compared to the herbicide alone without composite microspheres.

[0181] Control plants treated with 5% MCS 0.8-0.1 without glyphosate showed no visible phytotoxic effects or biomass reduction.

[0182] This assay demonstrates that adding MCS 0.8-0.1 at a 5% concentration (which has no herbicidal activity per se) increases herbicidal efficacy, allowing the application dose to be reduced by up to 50%. This is evidence of a surprising synergy that has never been previously suggested.

[0183] Example 8: Herbicidal Activity Assay of Glyphosate Combined with Composite Microspheres with Different TPP Concentrations on Wheat (Triticum aestivum) Used as a Target Plant (Simulating Weeds)

[0184] An evaluation was conducted to determine whether the surface charge (Zeta Potential) of chitosan composite microspheres differentially affects glyphosate efficacy on wheat plants grown on plates under controlled environmental conditions.

[0185] The following experimental design was used: wheat seeds were sterilized with 20% (v / v) bleach and evenly sown in sterile petri dishes (90 mm x 15 mm, Bio-Plast) containing filter paper moistened with 5 ml of sterile water. The seeds were germinated and grown for 8 days under controlled light, temperature, and humidity conditions.

[0186] After this period, each treatment was applied via foliar spray with the solutions described in Table 5. A PVC-EPA sprayer was used for application. The application volume for each solution was 0.8 ml per plate.

[0187] Table 5. Treatments to Evaluate the Potentiating Effect of Composite Microspheres MCS 0.8-0.1 and MCS 0.08-0.1 on Glyphosate Efficacy Applied to Wheat Plants.

[0188] Initially, an aqueous solution of 5 mg / ml MCS was prepared. The required amount of MCS solution (depending on the case) was combined with the tested doses of glyphosate. The solutions were placed in an orbital shaker (Vicking, model M23) for 2 hours. Six days after the treatments began (6 DDA), the plants were harvested and weighed on an analytical balance (OHAUS, model Traveler TA302) to quantify the fresh biomass of the aerial parts. The foliar biomass per plant was estimated (Figure 5).

[0189] Plants treated with 1 L ha"1glyphosate showed a 28% (w / w) reduction in dry foliar biomass compared to the water control (H2O) (Figure 5). In the case of plants treated with 5% MCS 0.08-0.1 + 1 L ha"1glyphosate, the biomass reduction was 40.56% (w / w) relative to the same control (H2O). This means that, in percentage terms, the use of the invention's formulation with 5% chitosan composite microspheres increased glyphosate's herbicidal efficacy by 45%. The percentage reduction in biomass increased from 28% (w / w) without MCS 0.08-0.1 to 40.56% (w / w) with MCS 0.08-0.1.

[0190] Enhancement formula:

[0191] Enhancement (%)=((40.56-28) / 28)xl00

[0192] This document refers to this indicator of increased herbicidal efficacy as "enhancement." In this assay, the use of MCS 0.08-0.1 enhanced the herbicidal formulation by 45% compared to the herbicidal active ingredient alone, without the addition of composite microspheres (MCS).

[0193] Additionally, in plants treated with 5% MCS 0.8-0.1 + 1 L ha"1glyphosate, the biomass reduction was 32.24% (w / w) relative to the H2O control. This indicates that the combination of 5% MCS 0.08-0.1 with 1 L ha"1glyphosate showed a 12.2% (w / w) increase in herbicidal efficacy compared to the same glyphosate dose without the addition of MCS. Meanwhile, the increase in glyphosate efficacy by adding 5% MCS 0.8- 0.1 was 3.9% (w / w) (Figure 5).

[0194] In percentage terms, the use of the invention's formulation with 5% chitosan composite microspheres increased glyphosate's herbicidal efficacy by 15%. The percentage reduction in biomass increased from 32.24% (w / w) without MCS 0.08 to 40.56% (w / w) with MCS 0.8-0.1:

[0195] Enhancement (%)=((32.24-28) / 28)xl00

[0196] This document refers to this indicator of increased herbicidal efficacy as "enhancement." In this assay, the use of MCS 0.8 provided the invention's herbicidal formulation with a 15% enhancement. Example 9: Field Trial of Herbicidal Activity of Glyphosate Combined with Composite Microspheres for Different Weeds

[0197] Independent field trials were conducted in various locations across the Argentine Republic. In all trials, glyphosate was formulated at different concentrations, always combined with MCS 0.8-0.1 composite microspheres dried using the spray method. Different weeds were tested, and measurements were taken on different days posttreatment.

[0198] The following general protocol was used for the trials:

[0199] The trials were conducted during the summer (2023 / 2024) in Argentina. The herbicidal activity of glyphosate on chemical fallow was evaluated. The design consisted of microplots measuring 3 meters in width by 8 meters in length, with three randomized repetitions. Different solutions were applied to the fallow using a CO2backpack sprayer at constant pressure.

[0200] Table 6 describes the different treatments evaluated. Treatments were assigned completely randomly to represent each experimental group equitably (n = 3). The percentage of weed control was quantified on different days after the treatments were applied (DDA). To determine the percentage of weed control, the control percentage of a treatment was estimated relative to the untreated weed treatment. These determinations were performed periodically following EWRS (European Weed Research Society) guidelines.

[0201] Monthly and cumulative rainfall during the campaign year were recorded.

[0202] Table 6. Treatments to Evaluate the Efficacy of Glyphosate Combined with Composite Microspheres MCS 0.8-0.1.

[0203] 1- Locality: Adolfo Gonzales Chaves

[0204] Field Location: 38°09'39.93"S / 59°45'42.70"W

[0205] Application Conditions: Applied using a CO2backpack sprayer with a 2- meter boom, flat fan nozzle, applying 110 L / ha of spray volume at 2 bar pressure.

[0206] Weed Tested: Cynodon dactylon (Gramon)

[0207] In Figure 6, the average results of n=3 for each treatment are shown, displaying the percentage of control relative to an untreated plot at two post-treatment intervals, 7 and 14 days. Treatments with glyphosate at a full dose of 1500 e.a. ha"1combined with composite microspheres (5% and 10% w / e.a.) showed slightly higher control percentages than glyphosate at the same dose alone. Additionally, treatments with 900 e.a. ha"1glyphosate combined with composite microspheres (5% and 10% w / e.a.) (treatments 4 and 5, respectively) achieved control percentages similar to those of the recommended glyphosate dose (treatment 1).

[0208] Weed Tested: Brassica rapa ("elongated" Nobolza)

[0209] In Figure 7, the average results of n=3 for each treatment are shown, displaying the percentage of control relative to an untreated plot at two post-treatment intervals, 7 and 14 days. Treatments 4, 5, 6, and 7 showed herbicidal control percentages similar to those of 1500 e.a. ha-1glyphosate alone (treatment 1).

[0210] Both trials demonstrated that the combination of composite microspheres with the herbicide allows at least a 40% reduction in application dose, as the combination enhances the herbicide's efficacy to match that of the highest doses tested.

[0211] 2- Locality: Tres Arroyos

[0212] Field Location: -38.602085°, -59.960860°

[0213] Application Conditions: Constant pressure backpack sprayer equipped with a carbon fiber side boom with four hollow cone nozzles spaced 52 cm apart, applying a flow rate of 100 L / ha at 3 bar pressure.

[0214] Weed Tested: Datura ferox (Chamico)

[0215] Glyphosate was applied as indicated on the label: 1500 e.a. ha-1glyphosate

[0216] With the following field applications of the invention's formulation under the same conditions:

[0217] • 900 e.a. ha-1glyphosate + 5% MCS 0.8

[0218] At 19 days post-treatment, the same herbicidal effect was observed, indicating that 40% less glyphosate can be applied while maintaining the same herbicidal potency.

[0219] • 900 e.a. ha-1glyphosate + 10% w / w MCS 0.8

[0220] At 19 and 28 days post-treatment, a 15% enhancement of the herbicidal effect was observed for the invention's formulation compared to glyphosate applied conventionally (#1).

[0221] • 1500 e.a. ha-1glyphosate + 10% w / w MCS 0.8

[0222] At 19 days post-treatment, a 40% enhancement of herbicidal power was achieved, and at 28 days, a 15% enhancement was noted.

[0223] Weed Tested: Chenopodium album (Quinoa) Glyphosate was applied as indicated on the label: 1500 e.a. ha1glyphosate

[0224] With the following field applications of the invention's formulation under the same conditions:

[0225] • 720 e.a. ha-1glyphosate + 5% w / w MCS 0.8

[0226] At 7 days post-treatment, a 5% enhancement of herbicidal effect was observed with a glyphosate dose that was 48% lower than the labeled application. This demonstrates that 52% less glyphosate can be applied while achieving a higher herbicidal potency.

[0227] • 1500 e.a. ha-1glyphosate + 10% w / w MCS 0.8

[0228] At 7 days post-treatment, a 10% enhancement of herbicidal powerwas achieved.

[0229] 3- Locality: 9 de Julio

[0230] Field Location: 35°23'12.86"S; 61°0'41.64"W

[0231] Application Conditions: CO2manual constant-pressure backpack sprayer with a boom equipped with four nozzles spaced 52 cm apart, using hollow cone 80015 nozzles, with a flow rate of 100 L / ha and 2.5 bar pressure.

[0232] Weed Tested: Eleusyne sp.

[0233] Glyphosate was applied as indicated on the label: 1500 e.a. ha-1glyphosate

[0234] With the following field applications of the invention's formulation under the same conditions:

[0235] • 900 e.a. ha-1glyphosate + 5% w / w MCS 0.8

[0236] • 900 e.a. ha-1glyphosate + 10% w / w MCS 0.8

[0237] At 14 days post-treatment, a 24% enhancement of herbicidal effect was observed with a glyphosate dose that was 60% of the labeled application. This demonstrates that 40% less glyphosate can be applied while achieving higher herbicidal potency. The treatment with 1500 e.a. ha-1glyphosate resulted in 63% weed control (Eleusine indica) at 21 days post-treatment (21 DDA) in chemical fallow. Adding 5% (w / e.a. glyphosate) MCS 0.8 at this herbicide concentration resulted in 72% weed control at 21 DDA, representing a 9% increase in glyphosate efficacy (Figure 11). Furthermore, the addition of 5% (w / e.a. glyphosate) MCS 0.8 to a reduced glyphosate dose (40% less than the full dose) achieved 70% weed control, demonstrating that the addition of MCS allows glyphosate application to be reduced by at least 40% under field conditions.

[0238] Example 10: Field Trial of Glyphosate and 2,4-D Choline Salt Herbicidal Activity Combined with Composite Microspheres on Weeds

[0239] Independent field trials were conducted in various locations across the Argentine Republic. In all trials, glyphosate and / or 2,4-D choline salt were formulated at different concentrations, always combined with MCS 0.8-0.15 composite microspheres dried using the spray method. The trials targeted Conyza sumatrensis (Rama Negra), and measurements were taken on different days post-treatment.

[0240] The following general protocol was used for the trials: The trials were conducted during the summer in Argentina. The herbicidal activity of glyphosate on chemical fallow was evaluated. The design consisted of micro-plots measuring 3 meters in width by 8 meters in length, with three randomized repetitions. Different solutions were applied to the fallow using a constant-pressure CO2backpack sprayer.

[0241] Table 7 describes the different treatments evaluated. Treatments were assigned completely randomly to represent each experimental group equitably (n = 3). The percentage of weed control was quantified on different days after treatment application (DDA). To determine the percentage of weed control, the control percentage of a treatment was estimated relative to the untreated weed treatment. These determinations were performed periodically following EWRS (European Weed Research Society) guidelines.

[0242] Monthly and cumulative rainfall during the campaign year were recorded. Table 7. Treatments to Evaluate Herbicidal Activity in Glyphosate Formulations and Glyphosate Combined with 2,4-D Choline Salt (2,4-D) and Composite Microspheres MCS 0.8-0.15.

[0243] Locality: Tres Arroyos

[0244] Field Location: -38.388612; -60.346111

[0245] Application Conditions: Manual backpack sprayer with a 2-meter working width, equipped with TeeJet 80015 nozzles at a constant pressure of 2.5 bar using CO2, with an application volume of 126 L / ha. After application, visual control measurements were conducted at 10, 20, 30, and 50 days after application (DDA). Data were subjected to variance analysis, and means were compared using Fisher's LSD test (p<0.05).

[0246] Weed Tested: Conyza sumatrensis (Rama Negra) Table 8 presents the average control percentage values for the four treatments at 10, 20, 30, and 50 DDA.

[0247] Table 8. Control of Rama Negra by Glyphosate Formulations and Glyphosate Combined with 2,4-D Choline Salt (2,4-D) and Composite Microspheres MCS 0.8-0.15.

[0248] This demonstrates the residual power of the present invention.

[0249] Example 11: Field Trial to Evaluate the Herbicidal Efficacy of Glufosinate Ammonium Combined with Composite Microspheres on the Weed Rama Negra

[0250] A field trial was conducted to evaluate the herbicidal effect of Glufosinate Ammonium at various concentrations, either alone or combined with composite microspheres MCS 0.8-0.15 dried using the spray method. The trials targeted Conyza sumatrensis (Rama Negra), and measurements were taken on different days post-treatment.

[0251] The following general protocol was used for the trials: The trials were conducted during the summer in Argentina. The herbicidal activity of Glufosinate Ammonium on chemical fallow was evaluated. The design consisted of micro-plots measuring 3 meters in width by 8 meters in length, with three randomized repetitions. Different solutions were applied to the fallow using a constant-pressure CO2backpack sprayer.

[0252] Table 9 describes the different treatments evaluated. Treatments were assigned completely randomly to represent each experimental group equitably (n = 3). The percentage of weed control was quantified on different days after treatment application (DDA). To determine the percentage of weed control, the control percentage of a treatment was estimated relative to the untreated weed treatment. These determinations were performed periodically following EWRS (European Weed Research Society) guidelines.

[0253] Monthly and cumulative rainfall during the campaign year were recorded.

[0254] Locality: Bragado, Field Location: -35.08043°S; -60.33043°W Application Conditions: Manual backpack sprayer with a 2-meter working width, equipped with TeeJet 80015 nozzles at a constant pressure of 2.5 bar using CO2, with an application volume of 126 L / ha. After application, visual control measurements were conducted at 7 and 25 days after application (DDA). Data were subjected to variance analysis, and means were compared using Fisher's LSD test (p<0.05).

[0255] Weed Tested: Conyza sumatrensis (Rama Negra)

[0256] Table 8. Control of Rama Negra by Different Formulations of Glufosinate Ammonium Combined with Composite Microspheres MCS 0.8-0.15. % Control for the Different Treatments Tested.

[0257] The incorporation of the microspheres in this trial, maintaining a weight-to-weight ratio of microspheres to the mass of active ingredient in each mixture (i.e., w / w MCS / Glufosinate Ammonium), demonstrated greater efficacy, faster weed burn-down, and lower weed regrowth rates. The treatment that included the composite microspheres of the present invention increased efficacy compared to the Glufosinate Ammonium treatment, even when applied at a lower dose than the commercially recommended full dose (2 L / ha).

[0258] Example 12: Field Trial of Herbicidal Activity of Glyphosate + 2,4-D Choline Salt Combined with Composite Microspheres on Weeds

[0259] Treatment 3 from Table 7 of Example 10 was applied, consisting of Glyphosate + 2,4-D Choline Salt in equal parts, along with 5% and 10% of the composite microspheres of the invention defined by Trial 4 of Table 1 in Example 2, specifically MCS 0.8-0 microspheres with a mass ratio of chitosan:gum:TPP of l:0:0.8, i.e., in the absence of gum, using a portable backpack sprayer. It was observed that the filters of the spray nozzles became clogged by particles, as shown in Figure 8. This was due to the low redispersibility of the microspheres of the invention in the absence of gum.

[0260] In subsequent similar trials using microspheres from Trial 4 of Table 1 in Example 2, MCS 0.8-0.1, which contain a gum concentration of 10% relative to the mass of chitosan, the spray nozzles did not clog, and it was not necessary to clean their filters.

[0261] Figure 9 demonstrates that a gum concentration between 5% and 20% improves the formation of the microspheres of the invention.

[0262] References

[0263] 1. OERKE E-C. Crop losses to pests. The Journal of Agricultural Science. 2006;144(l):31-43. doi:10.1017 / S0021859605005708 .

[0264] 2. Fisher, M., Henk, D., Briggs, C. et al. Emerging fungal threats to animal, plant and ecosystem health. Nature 484, 186-194 (2012). https : / / doi. org / 10.1038 / naturel0947

[0265] 3. Agrios, G.N. Introduction to Plant Pathology, 5th ed.; Elsevier Academic Press Publication: New York, NY, USA, 2005; p. 922.

[0266] 4. Vinutha, J.S.; Bhagat, D.; Bakthavatsalam, N. Nanotechnology in the management of polyphagous pest Helicoverpaarmigera. J. Acad. Ind. Res. 2013, 1, 606-608.

[0267] 5. Dhekney, S.A.; Li, Z.T.; Van Aman, M.; Dutt, M.; Tattersail, J.; Kelley, K.T.; Gray, D.J. Genetic transformation of embryogenic cultures and recovery of transgenic plants in Vitis vinifera, Vitis rotundifolia and Vitis hybrids. In Proceedings of the International Symposium on Biotechnology of Temperate Fruit Crops and Tropical Species, Daytona Beach, FL, USA, 10-14 October 2005; Volume 738, pp. 743-748. 6. Fernandez-Perez, M.; Gonzalez-Pradas, E.; Villafranca-Sdnchez, M.; Flores-Cespedes, F. Mobility of isoproturon from an alginate- bentonite controlled release formulation in layered soil. Chemosphere 2000, 41, 1495- 1501.

[0268] 7. Razzaghi-Abyaneh M, Chang P-K, Shams-Ghahfarokhi M and Rai M (2014) Global health issues of aflatoxins in food and agriculture: challenges and opportunities. Front. Microbiol. 5:420. doi: 10.3389 / fmicb.2014.00420

[0269] 8. BEDMAR, E, EYHERABIDE, G. H., SATORRE, E.H. 2002. Bases para el manejo de malezas. Bases para el manejo del maiz, el girasol y la soja. 273- 311. CREA.

[0270] 9. Green-Tracewicz E, Page ER, Swanton CJ. Light Quality and the Critical Period for Weed Control in Soybean. WeedScience. 2012;60(l):86-91. doi: 10.1614 / WS-D-l 1 -00072.1

[0271] 10. Islam MM, Megharaj M, Asaduzzaman M and Sudharsanam A (2024) Editorial: Agrochemicals in agricultural and non-agricultural settings: fate, distribution, and potential human and environmental health hazards. Front. Sustain. FoodSyst. 8:1461954. doi: 10.3389 / fsufs.2024.1461954

[0272] 11. Duhan JS, Kumar R, Kumar N, Kaur P, Nehra K, Duhan S. Nanotechnology: The new perspective in precision agriculture. Biotechnol Rep (Amst). 2017 May 24;15:ll-23. doi: 10.1016 / j.btre.2017.03.002. PMID: 28603692; PMCID: PMC5454086.

[0273] 12. Auffan, M., Rose, J., Bottero, JY. et al. Towards a definition of inorganic nanoparticles from an environmental, health and safety perspective. Nature Nanotech 4, 634-641 (2009). https: / / doi.org / 10.1038 / nnano.2009.242

[0274] 13. Kumar, S.; Bhanjana, G.; Sharma, A.; Sidhu, M.C.; Dilbaghi, N. Synthesis, characterization and on-field evaluation of pesticideloaded sodium alginate nanoparticles. Carbohydr. Polym. 2014, 101, 1061-1067.

[0275] 14. Faizan, M.; Rajput, V.D.; Al-Khuraif, A. A.; Arshad, M.; Minkina, T.; Sushkova, S.; Yu, F. Effect of Foliar Fertigation of Chitosan Nanoparticles on Cadmium Accumulation and Toxicity in Solanumlycopersicum. Biology 2021, 10, 666.

[0276] 15. Campos, E.V.R.; De Oliveira, J.L.; Da Silva, C.M.G.; Pascoli,M.; Pasquoto, T.; Lima, R.; Abhilash, P.C.; Fraceto, L.F.U. Polymeric and solid lipid nanoparticles for sustained release of carbendazim and tebuconazole in agricultural applications. Sci. Rep. 2015, 5, 13809.

[0277] 16. Salgueiro, A.M.; Daniel-da-Silva, A.L.; Fateixa, S.; Trindade, T. - Carrageenan hydrogel nanocomposites with release behaviour mediated by morphological distinct Au nanofillers. Carbohydr. Polym. 2013, 91, 100-109.

[0278] 17. Albuquerque, P.; Coelho, L.C.; Teixeira, J. A.; Carneiro-da-Cunha, M.G. Approaches in biotechnological applications of natural polymers. AIMS Mol. Sci. 2016, 3, 386-425.

[0279] 18. Sharma, A.; Sood, K.; Kaur, J.; Khatri, M. Agrochemical-loaded biocompatible chitosan nanoparticles for insect pest management. Biocatal. Agric. Biotechnol. 2019, 18, 101079.

[0280] 19. Slade, G.G.; Dourado, S.M., Jr.; Oliveira, R.J.D.; Moreto, J. A. Developing a mathematical model for the controlled release over time of sulfentrazone herbicide from a biodegradable polymer. Mater. Res. 2019, 22.

[0281] 20. Rashidipour, M.; Maleki, A.; Kordi, S.; Birjandi, M.; Pajouhi, N.; Mohammadi, E.; Rasoulian, R.; Davari, B. Pectin / chitosan / tripolyphosphate nanoparticles: Efficient carriers for reducing soil sorption, cytotoxicity, and mutagenicity of paraquat and enhancing its herbicide activity. J. Agric. FoodChem. 2019, 67, 5736-5745.

[0282] 21. Elabasy, A.; Shoaib, A.; Waqas, M.; Shi, Z.; Jiang, M. Cellulose nanocrystals loaded with thiamethoxam: Fabrication, characterization, and evaluation of insecticidal activity against Phenacoccussolenopsis Tinsley (Hemiptera: Pseudococcidae). Nanomaterials 2020, 10, 788.

[0283] 22. Roberts, G. A. F. Chitin Chemistry; Macmillan Press: London, 1992.; 64. 23. Bautista-Ba~nos, S.; Hernandez-Lauzardo, A. N.; Velazquez-del Valle, M. G.; Hernandez-Lopez, M.; AitBarka, E.; Bosquez- Molina, E.; Wilson, C.

[0284] L. Chitosan as a potential natural compound to control pre and postharvest diseases of horticultural commodities. CropProt. 2006, 25, 108-118. DOI: 10.1016 / j.cropro. 2005.03.010.

[0285] 24. Sahariah, P.; Masson, M. Antimicrobial chitosan and chitosan derivatives: a review of the structure-activity relationship. Biomacromolecules 2017, 18, 3846—3868. DOI: 10.1021 / acs.biomac. 7b01058.

[0286] 25. Francesconi, S.; Steiner, B.; Buerstmayr, H.; Lemmens, M.; Sulyok,

[0287] M.; Balestra, G.M. Chitosan Hydrochloride Decreases Fusariumgraminearum Growth and Virulence and Boosts Growth, Development and Systemic Acquired Resistance in Two Durum Wheat Genotypes. Molecules2020, 25, 4752. https: / / doi.org / 10.3390 / molecules25204752.

[0288] 26. Rychter P. Chitosan / glyphosate formulation as a potential, environmental friendly herbicide with prolonged activity. J EnvironSciHealth B. 2019;54(8):681-692. doi: 10.1080 / 03601234.2019.1632644. Epub 2019 Aug 12. PMID: 31403392.

[0289] 27. Das, R. K., Sarma, S. J., Brar, S. K., &Verma, M. (2014). Nanoformulation of insecticides-Novel products. J BiofertilBiopestici, 5(1), el20.

[0290] 28. Grenha A. Chitosan nanoparticles: a survey of preparation methods. J Drug Target. 2012 May;20(4):291-300. doi:

[0291] 10.3109 / 1061186X.2011.654121. Epub 2012 Feb 2. PMID: 22296336.

[0292] 29. Tokumitsu H, Ichikawa H, Fukumori Y, Block LH. (1999a). Preparation of gadopentetic acid-loaded chitosan microparticles for gadolinium neutron-capture therapy of cancer by a novel emulsion-droplet coalescence technique. ChemPharm Bull, 47, 838-842.

[0293] 30. El-Shabouri MH. (2002). Positively charged nanoparticles for improving the oral bioavailability of cyclosporin-A. Inti Pharm, 249, 101-108. 31. Mitra S, Gaur U, Ghosh PC, Maitra AN. (2001). Tumour targeted delivery of encapsulated dextran-doxorubicin conjugate using chitosan nanoparticles as carrier. J Control Release, 74, 317-323.

[0294] 32. Calvo P, Remunan-Lopez C, Vila-Jato JL, Alonso MJ. (1997). Novel hydrophilic chitosan-polyethylene oxide nanoparticles as protein carriers. J AppIPolymSci, 63, 125-132.

[0295] 33. Sarmento B, Martins S, Ribeiro A, Veiga F, Neufeld R, Ferreira D. (2006). Development and comparison of different nanoparticulate polyelectrolyte complexes as insulin carriers. Int J Peptide Res Ther, 12, 131-138.

[0296] 34. Tian XX, Groves MJ. (1999). Formulation and biological activity of antineoplastic proteoglycans derived from Mycobacterium vaccae in chitosan nanoparticles. J PharmPharmacol, 51, 151-157.

[0297] 35. Chan HK, Kwok PC. (2011). Production methods for nanodrug particles using the bottom-up approach. AdvDrugDelivRev, 63, 406-416.

[0298] 36. Wang J, Byrne JD, Napier ME, DeSimone JM. (2011). More effective nanomedicines through particle design. Small, 7, 1919-1931.

[0299] 37. Fan W, Yan IV, Xu Z, Ni H. (2012). Formation mechanism of monodisperse, low molecular weight chitosan nanoparticles by ionic gelation technique. Colloids Surf B Biointerfaces, 90, 21-27. Fan W, Yan W, Xu Z, Ni H.

[0300] 38. Bhumkar DR, Pokharkar VB. Studies on effect of pH on crosslinking of chitosan with sodium tripolyphosphate: a technical note. AAPS PharmSciTech. 2006 Jun 2;7(2):E50. doi: 10.1208 / pt070250. PMID: 16796367; PM CID: PMC2750277.

[0301] 39. Janes KA, Calvo P, Alonso MJ. (2001). Polysaccharide colloidal particles as delivery systems for macromolecules. AdvDrugDelivRev, 47, 83-97.

[0302] 40. Terbojevich M, Muzzarelli RAA. (2009). Chitosan. In: Phillips GO, Williams P, eds. Handbook of Hydrocolloids. Cambridge: Woodhead Publishing Ltd., 367-378. 41. Bhattarai N, Gunn J, Zhang M. (2010). Chitosan-based hydrogels for controlled, localized drug delivery. AdvDrugDelivRev, 62, 83-99.

[0303] 42. Bodmeier R, Chen HG, Paeratakul O. (1989). A novel approach to the oral delivery of micro- or nanoparticles. Pharm Res, 6, 413-417.

[0304] 43. Carvalho EL, Grenha A, Remunan-Lopez C, Alonso MJ, Seijo B. (2009). Mucosaldelivery of liposome-chitosannanoparticlecomplexes. MethEnzymol, 465, 289-312.

[0305] 44. Kumar, R.; Nain, V.; Duhan, J.S. An Ecological Approach to Control Pathogens of Lycopersicon esculentum L. by Slow Release of Mancozeb from Biopolymeric Conjugated Nanoparticles. J. Xenobiot. 2022, 12, 329-343. https: / / doi.ora / 10.3390 / ioxl2040023.

[0306] 45. Silva Mdos S, Cocenza DS, Grillo R, de Melo NF, Tonello PS, de Oliveira LC, Cassimiro DL, Rosa AH, Fraceto LF. Paraguat-loaded alginate / chitosan nanoparticles: preparation, characterization and soil sorption studies. J Hazard Mater. 2011 Jun 15;190(l-3):366-74. doi: 10.1016 / j.jhazmat.2011.03.057. Epub 2011 Mar 23. PMID: 21493003.

[0307] 46. Grillo R, Pereira AE, Nishisaka CS, de Lima R, Oehlke K, Greiner R, Fraceto LF. Chitosan / tripolyphosphate nanoparticles loaded with paraquat herbicide: an environmentally safer alternative for weed control. J Hazard Mater. 2014 Aug 15;278:163-71. doi: 10.1016 / j.jhazmat.2014.05.079. Epub 2014 Jun 11. PMID: 24968252.

[0308] 47. Costa, Marcia &Rabelo, Kissila& Ferreira, Ivana & Cruz, Mauricio. (2021). Sodium alginate / chitosan / glyphosate superabsorbent bio-foam as a release system for herbicide. Journal of AppliedPolymerScience. 139. 51776. 10.1002 / app.51776.

[0309] 48. Butstraen C, Salatin F. Preparation of microcapsules by complex coacervation of gum Arabic and chitosan. CarbohydrPolym. 2014 Jan;99:608-16. doi: 10.1016 / j.carbpol.2013.09.006. Epub 2013 Sep 8. PMID: 24274550.

Claims

Claims1. A herbicidal formulation comprising a herbicidal active ingredient in an aqueous solution and composite microspheres that enhance herbicidal efficacy against weeds in crops, wherein said composite microspheres, in powder form, are present in a concentration ranging from 2% to 200% w / w relative to the herbicidal active ingredient when combined with the aqueous solution, and wherein said composite microspheres include chitosan with an average molecular weight ranging from 190,000 to 310,000 g / mol and a cross-linking agent.

2. The herbicidal formulation according to claim 1, wherein the herbicidal active ingredient is selected from the group consisting of Glyphosate, Imazapyr, Imazethapyr, Atrazine, Simazine, Diuron, Pendimethalin, Trifluralin, Fluazifop-p- butyl, Quizalofop-p-ethyl, Dicamba, Paraquat, Glufosinate, Clethodim, Haloxyfop, Fomesafen, 2,4-Dichlorophenoxyacetic Acid, Thiencarbazone- methyl, Pyraclostrobin, Ammonium Glufosinate, Imazamox, Imazaquin, Flurochloridone, Clethodim, Haloxyfop, Lactofen, Thiencarbazone-methyl, Diflufenican, Sulfentrazone, S-metolachlor, Flumioxazin, Pyroxasulfone, MCPA, Saflufenacil, Bromoxynil, their derivatives, and their mixtures.

3. The herbicidal formulation according to claim 1, wherein the herbicidal active ingredient is Glyphosate.

4. The herbicidal formulation according to claim 1, wherein the herbicidal active ingredient is Glufosinate.

5. The herbicidal formulation according to claim 1, wherein the composite microspheres comprise chitosan with an average molecular weight ranging from 190,000 to 310,000 g / mol, a dispersant, and a cross-linking agent, and have a zeta potential ranging from +30 to -30 in the aqueous solution, determined by the chitosan-to-cross-linking agent mass ratio of 1:X, where X ranges from 0.08 to 1.5.

6. The herbicidal formulation according to claim 1, wherein the composite microspheres comprise chitosan, gum arable, and tripolyphosphate in the absence of agrochemical actives.

7. The herbicidal formulation according to claim 1, further comprising a co-adjuvant selected from the group consisting of an adhesive, a surfactant, a stabilizer, an antifoaming agent, a pH buffer, a sequestrant, their derivatives, and combinations thereof.

8. The herbicidal formulation according to claim 1, wherein the herbicidal active ingredient is at a concentration equal to or less than the field application rate indicated on the product label for the herbicide alone.

9. The herbicidal formulation according to claim 1, wherein the concentration of the composite microspheres ranges from 5% to 100% w / w relative to the herbicidal active ingredient.

10. The herbicidal formulation according to claim 1, wherein the concentration of the composite microspheres ranges from 5% to 10% w / w relative to the herbicidal active ingredient.

11. The herbicidal formulation according to claim 1, wherein the mixture of the herbicidal active ingredient and composite microspheres is prepared in the field prior to use.

12. The herbicidal formulation according to claim 1, wherein the mixture of the herbicidal active ingredient and composite microspheres is contained in packaging intended for commercialization and distribution.

13. A method for obtaining the herbicidal formulation of claim 1, comprising the steps of: a) Preparing an aqueous solution of the herbicidal active ingredient at the desired concentration; and b) Adding dry composite microspheres in a concentration of up to 100% by weight relative to the herbicidal active ingredient and stirring.

14. The method of claim 13, further comprising the addition of a co-adjuvant.

15. A method for preventing or reducing the growth of undesired vegetation, comprising applying the herbicidal formulation of claim 1 to a crop or soil prior to planting.

16. The method of claim 15, wherein the crop is selected from the group consisting of cereals, oilseeds, forest crops, fruits, trees, ornamentals, vegetables, wheat, barley, rye, triticale, oats, corn, sunflower, rice, soybean, pea, bean, peanut, canola, kale, cotton, lentils, potato, sugar beet, sugarcane, grass, sorghum, flax, legumes, tomato, pepper, squash, lettuce, chard, carrot, radish, apple, pear, plum, banana, mango, citrus, walnut, almond, coffee, cacao, grapevine, and others.

17. The method of claim 15, wherein the herbicidal formulation is applied at a herbicidal active ingredient concentration lower than the rate indicated on the product label.

18. The method of claim 15, wherein the herbicidal action is enhanced by up to 70% compared to the herbicide without composite microspheres.

19. A herbicidal formulation enhancing the efficacy of a herbicidal active ingredient by at least 10%, comprising a herbicidal active ingredient and a composite microsphere; wherein the composite microsphere includes chitosan with an average molecular weight ranging from 190,000 to 310,000 g / mol and a crosslinking agent, with a chitosan-to-cross-linking agent mass ratio of 1:X, where X ranges from 0.08 to 1.5; and wherein the concentration of the composite microsphere ranges from 2% to 100% w / w relative to the herbicidal active ingredient in an aqueous phase.

20. The formulation of claim 1 or 19, wherein the composite microsphere further comprises a dispersant selected from the group consisting of gum arabic, xanthan gum, guar gum, carrageenan, pectin, alginates, dextrins, cyclodextrins, maltodextrins, tragacanth gum, mesquite gum, and mixtures thereof.

21. The formulation of claim 20, wherein the composite microsphere comprises a surface electric charge or zeta potential adjustable by the concentration of thecross-linking agent, with X ranging from 0.08 to 0.8, and wherein the cross-linking agent is TPP.

22. The formulation of claim 19, wherein the herbicidal active ingredient is in an aqueous composition in a container, and the composite microsphere is a powder added to the container.

23. The formulation of claim 1 or 19, wherein the composite microsphere comprises a cross-linking agent selected from the group consisting of sodium tripolyphosphate (TPP), sodium hexametaphosphate, and their derivatives.

24. The formulation of claim 1 or 19, wherein the composite microsphere comprises a cross-linking agent, which is sodium tripolyphosphate (TPP).

25. The formulation of claim 1 or 19, wherein the composite microsphere further comprises a dispersant that enhances the redispersibility of the microsphere when combined with an aqueous herbicide solution, wherein the dispersant is selected from the group consisting of gum arabic, xanthan gum, guar gum, carrageenan, pectin, alginates, dextrins, cyclodextrins, maltodextrins, tragacanth gum, mesquite gum, polyethylene glycol, and mixtures thereof, and wherein the dispersant comprises a chitosan-to-dispersant mass ratio of Y:l, where Y ranges from 5 to 20.

26. The formulation of claim 25, wherein the dispersant is gum arabic.

27. A composite microsphere that enhances herbicidal efficacy against weeds in crops for preparing the formulations of claim 1 or 19, comprising chitosan with an average molecular weight ranging from 190,000 to 310,000 g / mol and a crosslinking agent, in the absence of a herbicidal active ingredient.

28. The composite microsphere of claim 27, comprising a surface electric charge (zeta potential) in an aqueous medium adjustable by the concentration of the cross-linking agent, and wherein the chitosan-to-cross-linking agent mass ratio is 1:X, where X ranges from 0.08 to 1.5, defining the zeta potential in an aqueous solution of the microsphere.

29. The composite microsphere of claim 27, wherein the cross-linking agent is selected from the group consisting of sodium tripolyphosphate (TPP), sodium hexametaphosphate, and their derivatives.

30. The composite microsphere of claim 27, wherein the cross-linking agent is sodium tripolyphosphate (TPP).

31. The composite microsphere of claim 27, further comprising a dispersant that enhances the redispersibility of the microsphere when combined with an aqueous herbicide solution, wherein the dispersant is selected from the group consisting of gum arabic, xanthan gum, guar gum, carrageenan, pectin, alginates, dextrins, cyclodextrins, maltodextrins, tragacanth gum, mesquite gum, polyethylene glycol, and mixtures thereof.

32. The composite microsphere of claim 31, wherein the dispersant comprises a chitosan-to-dispersant mass ratio of Y:l, where Y ranges from 5 to 20.

33. The composite microsphere of claim 31, wherein the dispersant is gum arabic.

34. A process for obtaining the composite microsphere of claim 27, comprising the steps of: a) Mixing acetic acid, chitosan, and water and stirring; b) Adding the dispersant and stirring; c) Adding the cross-linking agent solution dropwise; d) Allowing the mixture to rest; and e) Drying the obtained microspheres.

35. The process of claim 34, comprising the steps of: a) Dissolving chitosan in 1% v / v acetic acid with constant stirring at a temperature of 20°C until no lumps are observed; b) Adding dispersant and stirring until completely dissolved to achieve a chitosan-to-dispersant mass ratio of Y:l, where Y ranges from 5 to 20; c) Adding a cross-linking agent solution dropwise to the mixture, at a chitosan-to-cross- linking agent mass ratio of 1:X, where X ranges from 0.08 to 1.2, in the absence of a herbicidal active ingredient; d) Adjusting the pH to approximately 6.5; and e) Drying the composite microspheres.

36. The process of claim 35, characterized in that said step e) involves freeze-drying using a 220 V lyophilizer at -45°C and 10 Pa pressure, followed by grinding witha mortar and sieving through a 100-mesh filter to obtain the composite microspheres.

37. The process of claim 34, characterized in that said step e) involves spray drying at an inlet temperature of 200°C.

Images