Stable plant supporting formulation

Abstract

The present invention relates to a method of manufacture of a stable plant supporting formulation and the plant supporting formulation as produced by the method of manufacture as well as a method of any one or more of the following: (i) delivering one or more active compound(s) to a plant; (ii) improving the absorption of one or more active compound(s) in a plant; (iii) improving the distribution and translocation of one or more active compound(s) in a plant; (iv) releasing one or more active compound(s) to plant cells for use in the plants; and (v) improvement of energy metabolism in a plant thereby supporting the plant, the method comprising the use of the stable plant supporting formulation of the invention.

Description

[0001] STABLE PLANT SUPPORTING FORMULATION

[0002] FIELD OF APPLICATION OF THE INVENTION

[0003] The present invention relates to a method of manufacture of a stable plant supporting formulation. The invention additionally relates to a plant supporting formulation as produced by the method of manufacture. The stable plant supporting formulation may be for use in any one or more of the following applications: (i) delivering one or more active compound(s) to a plant; (ii) improving the absorption of one or more active compound(s) delivered to a plant by the stable plant supporting formulation; (iii) improving the distribution and translocation in a plant of one or more active compound(s) delivered to the plant by the stable plant supporting formulation; (iv) releasing an active compound delivered to a plant by the stable plant supporting formulation to plant cells; (v) and supporting a plant by improvement of energy metabolism of the plant.

[0004] Additionally, the invention relates to a method of any one or more of the following: (i) delivering one or more active compound(s) to a plant; (ii) improving the absorption of one or more active compound(s) in a plant; (iii) improving the distribution and translocation of one or more active compound(s) in a plant; (iv) releasing one or more active compound(s) to plant cells for use in the plants; and (v) improvement of energy metabolism in a plant thereby supporting the plant, the method comprising the use of the stable plant supporting formulation of the invention.

[0005] BACKGROUND TO THE INVENTION

[0006] The uptake of substances by plants and its subsequent translocation by plants are often improved using various additives or adjuvants as known in the art, including emulsions, oils and penetrants and collectively called ‘adjuvants’ for the purpose of this application. Such adjuvants need to be stable under extreme temperature and mobility conditions during various phases before final use: whilst in transport, when mixed as tank mixes on the field and when administered in a wide range of temperatures. Stability under these conditions is not a given, even if the product or adjuvant complies with normal agricultural stability testing.

[0007] The United States Environmental Protection Agency (EPA) regulates agricultural adjuvants under the Federal Insecticide, Fungicide, and Rodenticide Act (FIFRA) and stability data may be requested as part of the registration process for agricultural adjuvants, including information on physical, chemical, and biological stability. In Australia, agricultural adjuvants are regulated by the Australian Pesticides and Veterinary Medicines Authority (APVMA), which establishes guidelines and requirements for stability testing of agricultural adjuvants as part of the registration process. The European Chemicals Agency (ECHA) provides guidance on stability testing of biocidal products, including adjuvants, under the Biocidal Products Regulation (BPR) and stability data may be required as part of the product authorization process in the EU to ensure product quality, safety, and efficacy. In South Africa, agricultural adjuvants are regulated by the Department of Agriculture, Land Reform and Rural Development (DALRRD) under the Fertilizers, Farm Feeds, Agricultural Remedies, and Stock Remedies Act and stability testing of agricultural adjuvants is part of the registration process to demonstrate product quality and effectiveness.

[0008] The pesticide specifications of the World Health Organization I Food and Agriculture Organization of the United Nations (FAO / WHO) recommend testing of relevant product parameters before and after storage for 14 days at 54°C so any potential changes can be detected with an alternative time-temperature regime in certain circumstances. Herewith a typical accelerated stability test protocol prescribed in the pesticide manual of the FAO and WHO: For the stability test, the samples should be stored at 54°C ± 2°C for 14 days and analysed by an appropriate method at day 0, 7 and 14. The analyses are done to assess the shelf life of the formulations when exposed to factors such as temperature and humidity. Stability tests at elevated temperatures are designed to increase the rate of chemical degradation or physical change in a product in order to obtain information on the shelf life of a product in a shorter time than a real-time study. Accelerated testing involves extrapolations from higher to lower temperatures, and from shorter to longer storage periods.

[0009] Most of the stability requirements relate to shelf life and not so much to use conditions and for most agricultural chemical products, accelerated storage stability data is deemed sufficient, without the provision of real-time storage or use stability data. However, in practice and in many countries, products in drums (20L or 100L) are supplied to farmers on the back of open trucks after having been transported on dirt roads at 40°C. The interior temperatures of the containers can reach 70°C. In addition, product is shipped in 1000L flow-bins across the equator in choppy seas and dropped in harbours in temperatures below freezing.

[0010] The Applicant’s previous patent WO 2007 / 096833 discloses a plant supporting formulation, vehicle for the delivery and translocation of phytologically beneficial substances and compositions containing same. These formulations improved uptake of substances by plants, resulting in improved harvestable yield, yet the formulations prepared according to invention disclosure do not withstand extreme temperature and mobility conditions, as could be observed in terms of changes in morphology, consistency and colour of the formulations.

[0011] It is accordingly an object of the present invention to provide an improved plant supporting formulation which demonstrates increased stability, and which at least partially ameliorates or overcomes the disadvantages of the prior art.

[0012] SUMMARY OF THE INVENTION

[0013] “Vesicles” and “microsponges” mean, in terms of this invention, vesicles and microsponges as obtained using the method of WO 2007 / 096833. “Grobler patent” refers to PCT patent application WO 2007 / 096833, or a plant supporting formulation prepared as disclosed in WO 2007 / 096833.

[0014] According to a first aspect of the invention, there is provided a method for manufacturing a stabilised plant supporting formulation, the method comprising: i. introducing a gas into an aqueous solution in a pressure vessel under pressure to form an aqueous phase; ii. combining a first fatty acid-based component heated to from about 60 °C to about 70 °C and one or more surfactant component(s) heated to from about 100 °C to about 200 °C, or about 180 °C, followed by adding a second fatty acid-based component including a vitamin E derivative, to form an oil phase; iii. heating the aqueous phase of step i. to about 70 °C and then adding the heated oil phase of step ii. to the heated aqueous phase to a concentration of between about 3.2 % and 4 % (v / v of the final stabilised plant supporting formulation) of the oil phase of step ii.; iv. mixing the combined aqueous phase and oil phase of step iii. using a highspeed shearer wherein the combined components are subjected to at least three cycles of mixing and resting where the periods of mixing and resting is incrementally decreased in each cycle to form the stabilised plant supporting formulation; v. optionally subjecting the stabilised plant supporting formulation of step iv. to at least one cycle, or at least two cycles of heating to a first temperature of between about 65 and 70 °C and cooling to a second temperature of between about 15 and 23 °C; and vi. introducing a gas under pressure in a pressure vessel into the stabilised emulsion of step iv. or, when used, step v. until saturation for a period of about 72 hours to form the stabilised plant supporting formulation.

[0015] The method may further comprise a final step of bottling or encapsulating the stabilised plant supporting formulation in a protective coating. The aqueous solution may be purified water or unpurified water, including reverse osmosis purified water. Alternatively, the aqueous solution may be selected from a buffer having a pH of between about pH 3 to 9, growth media or even the water component of a plant.

[0016] The gas may be selected from nitrous oxide, carbon dioxide or carbon oxysulfide, all in the zwitterionic form. In a preferred embodiment of the invention, the gas is nitrous oxide.

[0017] The pressure in step i. and step vi may be about 2 kPa (2 bar) and may be introduced using either surface pressure or sparging. The gas may be introduced under pressure in step i. for a period of about 24 hours.

[0018] The fatty acid-based component may be an ethyl ester fatty acid-based component. The ethyl ester fatty acid may comprise about 1 to 5 % v / v of the fatty acid-based component.

[0019] The ethyl ester fatty acid component may be selected from any two or more sources that are combined, the sources comprising ethyl esters of soy oil, flax seed oil, chia seed oil, hemp seed oil, peppermint oil, rosemary oil, walnut oil, or fish oil. The ethyl ester fatty acid component chain length may range from Ci6 to C30. In particular, the combined ethyl ester fatty acid component chain length may range from C16 to Cis from a first source and from C20 and C22 from a second source.

[0020] A person skilled in the art would appreciate that the ethyl ester fatty acid source determines the structure and the characteristics of the emulsions formed by the method where the more unsaturated the fatty acid is, the more sponge-like the emulsion structure will be. The combined ethyl ester fatty acid component may have a concentration of about 3.8 % (v / v of the final plant supporting formulation). It is further to be appreciated that due to the length difference in fatty acid chain length between the two sources of ethyl ester fatty acids in the combined ethyl ester fatty acid component, the resulting emulsion includes two populations of sizes, one population in the nanometer-size range and a second population in the micro-size range as determined by particle size analysis on a Malvern sizer.

[0021] In one possible embodiment of the invention the first source of ethyl ester fatty acid may be a C16 and C18 Vitamin F ethyl ester (also known by the INCI name as ethyl linoleate (and) ethyl linolenate (and) ethyl oleate), an ethyl ester fatty acid obtained from soy. The second source of ethyl ester fatty acid in this possible embodiment of the invention may be a C20 and C22 long chain Vitamin F ethyl ester product that is manufactured through the extraction and ethyl esterification of a combination of linseed and flax oil. However, it is to be appreciated that alternative C20 and C22 long chain ethyl fatty acids may be obtained from fish oil, in particular the ethyl esters of decahexonoic acid and eicosapentanoic acid or a combination thereof. In this embodiment, the typical amount of each of the ethyl ester fatty acids is added to a produce a final concentration of about 2.7 % of the C16 and C18 ethyl ester fatty acids and about 0.1 % of the C20 and C22 ethyl ester fatty acids (v / v of the final plant supporting formulation).

[0022] The non-ionic surfactant may comprise one or more surfactant molecules in combination. For example, the ratio of the ethyl ester fatty acid combination component of step ii. to the non-ionic surfactant may range between about 2.4:1 and 3.8:1.

[0023] The non-ionic surfactant(s) may be selected from natural oils composed largely of ricinoleic acid-based oils with ethylene oxide. The surfactant(s) may be modified as to the extent of hydrogenation, ethylation and the addition of groups including polyethylene glycol. It is to be appreciated that a person skilled in the art may select a surfactant depending on a specific active compound to be delivered by the plant supporting formulation of the invention.

[0024] In one possible embodiment of the invention, the one or more non-ionic surfactant(s) is a pegylated, hydrogenated fatty acid, ricinoleic acid (also known by the INCI name as PEG-n-Hydrogenated Castor Oil). Optionally, the one or more non-ionic surfactant(s) comprise a second surfactant including a co-solvent in the form of: (i) a polysorbate, or a polyethylene glycol (PEG- 400) to a final concentration of 2 % (v / v of the final plant supporting formulation), or (ii) a fatty alcohol ethoxylate to a final concentration of 1 % (v / v of the final plant supporting formulation) may be added to the non-ionic surfactant of step ii.

[0025] The vitamin E derivative may be selected from dl-a-Tocopherol or a physiologically active isomer thereof, including tocotrienol (a more stable form of tocopherol). Typically, the vitamin E derivative is added to the oil phase to obtain a final concentration of about 0.25 % (v / v of the final plant supporting formulation).

[0026] The first temperature in step v. may be a temperature at least above the melting point of the first and second fatty acid-based components, and the second temperature may be room temperature. For example, the first temperature may be about 60 °C to 70 °C. The emulsion of step v. may be stirred.

[0027] For example, the mixing in step iv. may be performed by a first cycle of 2 minutes of mixing and 2 minutes rest; a second cycle of 1 minute mixing and 1 minute rest; followed by a third cycle of 0.5 minutes mixing and 0.5 minutes rest. However, a person skilled in the art would appreciate that should the volumes be upscaled (to greater than about 500 litres), the mixing and resting times may be extended.

[0028] According to a second aspect of the invention, there is provided a stabilised plant supporting formulation produced according to the method of the invention. The stabilised plant supporting formulation of the invention maintains substantially the same mean particle size and Zeta-potential after transport compared to before transport, contrary to the stabilised plant supporting formulation produced according to the Grobler patent. Furthermore, the plurality of vesicles and / or microsponges within the stabilised plant supporting formulation of the invention maintain a size of less than about 200 nm after transport.

[0029] The stabilised plant supporting formulation may comprise: I. a stabilised micro-emulsion comprising a dispersion of vesicles and / or microsponges of the oil phase in the aqueous phase; and

[0030] II. a gas dissolved in the oil phase.

[0031] The stabilised plant supporting formulation may be for use in any one or more of the following applications: a. delivering one or more compound(s) to a plant selected from one or more active compounds and / or one or more compounds beneficial for plant growth and / or health; b. improving the absorption of the one or more compound(s) delivered to the plant; c. improving distribution and translocation in a plant of the one or more compound(s); d. facilitating release of the one or more compound(s) delivered by the stabilised plant supporting formulation to a plant cell; or e. supporting plant growth and / or health by improvement of energy metabolism of the plant.

[0032] According to a third aspect of the invention, there is provided a method of treating a plant or supporting the growth and / or health of a plant, comprising any one or more of the following:

[0033] A. delivering one or more compound(s) to a plant selected from one or more active compounds and / or or one or more compound(s) beneficial for plant growth and / or health to a plant;

[0034] B. improving the absorption of the one or more compound(s) delivered to the plant;

[0035] C. improving the distribution and translocation in a plant of the one or more compound(s) delivered to the plant;

[0036] D. releasing to plant cells the one or more compound(s) delivered to the plant; or

[0037] E. supporting plant growth and / or health by improvement of energy metabolism of the plant, the method comprising the use of the stable plant, wherein the method comprises the use of the stabilised plant supporting formulation of the invention. The one or more compound(s) for delivery by the stabilised plant supporting formulation of the invention may be comprised of one or more active compound(s) used for treatment of plant conditions or diseases, phytologically beneficial substances, amines and polyamines, waxes, plant polymers, saccharides, natural compounds including amino acids, humic substances, algal extract, or a pyroligneous extract.

[0038] The stabilised plant supporting formulation of the invention may further comprise additives including tensoactives, additional surfactants, salicylic compounds, emulsifiers, stabilizers and preservatives.

[0039] The one or more compound(s) may be added to either or both of the oil phase or the aqueous phase, or to the final stabilised plant supporting formulation, depending on the physico-chemical properties and the lability of said substances and compound(s).

[0040] For example, the one or more compound(s), including phytologically beneficial substance(s) may comprise or be selected from the group consisting of a plant pesticide including a herbicide, fungicide, bactericide, insecticide, antiplant virus agent; a plant growth regulator selected from the products in the group consisting of,

[0041] 2-(1 -2-methylnaphthyl)acetamide; 2-(1 -2-methylnaphthyl)acetic acid; 2-(1 - naphthyl)acetamide; 2-(1 -naphthyl)acetic acid; 2,4-D (sodium salt); 3,5,6 TPA; 4-indol-

[0042] 3-ylbutyric acid; 6-benzyl adenine; alkoxylated fatty alkylamine polymer; alkylamine polymer; aminoethoxyvinylglycine hydrochloride; ammoniated nitrates; auxins; calcium arsenate; carbaryl; chlormequat chloride; chlorpropham; chlorthal-dimethyl; cloprop; cyanamide; daminozide; decan-1 -ol; dichlorprop; dichlorprop (2-butoxyethyl ester); dimethipin; dinocap; diquat dibromide; diuron; ethephon; fluazifop-p-butyl; gibberellins; glyphosate-isopropylamine; glyphosate-trimesium; haloxyfop-P-methyl; indolylacetic acid; maleic hydrazide; mepiquat chloride; methylcyclopropene; mineral oil; n-decanol; octan-1 -ol; paclobutrazole; paraquat dichloride; pendimethalin; prohexadione-calcium; salicylic acid, sodium chlorate; thidiazuron; trinexapac-ethyl; and uniconazole; a plant immune modulator; and a biostimulant selected from the group of products known as phytohormones, or any combination thereof. According to a fourth aspect of the invention there is provided a plant supporting formulation according to the invention comprising one or more compound(s) selected from one or more active compound(s) and / or or one or more compounds beneficial for plant growth and / or health to a plant.

[0043] According to a fifth aspect of the invention, there is provided a method of administration of the stabilised plant supporting formulation to a plant comprising a step of diluting the stabilised plant supporting formulation with a diluent solution suitable for administration to the plant which is typically water. The ratio of stabilised plant supporting formulation to the diluent solution may be about 1 :1 for stem application, about 1 :10 for ornamentals in open settings such as in the field or in greenhouses, and about 1 :5000 for hydroponic systems. It is to be appreciated that the dilution may vary depending on the method of administration, the type of cultivation (e.g. drip irrigation, foliar spraying by hand, tractor, or plane). Typically, between about 60 mL and 120 mL is administered per hectare for open field crops and controlled environments such as greenhouses.

[0044] In a case where one or more compound(s) selected from one or more active compounds and / or one or more compounds beneficial for plant growth and / or health are to be added to the final stabilised plant supporting formulation, the method of administration may further comprise a step of entrapment of the one or more compound(s) in the stabilised plant supporting formulation comprising:

[0045] (A) addition of the one or more compound(s) to the stabilised plant supporting formulation either prior to dilution or after dilution;

[0046] (B) mixing the stabilised plant supporting formulation and one or more compound(s);

[0047] (C) leaving the mixture of step (B) to stand for at least about 30 minutes to overnight for entrapment.

[0048] The mixing of step (B) can be performed at room or field temperature. The mixing of step (B) may further be performed by shaking or stirring, or by addition to a tank mixer on site. In the case of phytologically beneficial substances with large molecular weights such as peptides, the mixture of step (C) may be stored overnight at 4°C for entrapment.

[0049] The above and other characteristics, features and advantages of the present invention will become apparent from the following detailed description, taken in conjunction with the accompanying drawing which illustrate, by way of example, the principles of the invention. This description is given for the sake of example only, without limiting the scope of the invention. The reference figures quoted below refer to the attached drawings.

[0050] BRIEF DESCRIPTION OF THE DRAWINGS

[0051] Embodiments of the invention are described below, by way of non-limiting examples, and with reference to the accompanying drawings which:

[0052] Figure 1 graphically illustrates the particle size of products prepared according to the Grobler patent before (A) and after (B) transport;

[0053] Figure 2 graphically illustrates the particle size of products prepared according to a preferred embodiment of the present invention before (A) and after (B) transport;

[0054] Figure 3 graphically illustrates the particle size (A) and Zeta potential (B) of a product as prepared according to the Grobler patent before transport;

[0055] Figure 4 graphically illustrates the particle size (A) and Zeta potential (B) of a product as prepared according to the Grobler patent after transport;

[0056] Figure 5 graphically illustrates the particle size (A) and Zeta potential (B) of a product as prepared according to a preferred embodiment of the present invention before transport; and Figure 6 graphically illustrates the particle size (A) and Zeta potential (B) of a product as prepared according to a preferred embodiment of the present invention after transport.

[0057] The presently disclosed subject matter will now be described more fully hereinafter with reference to the accompanying Examples, in which representative embodiments are shown. The presently disclosed subject matter can, however, be embodied in different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the embodiments to those skilled in the art.

[0058] DESCRIPTION OF PREFERRED EMBODIMENTS OF THE INVENTION

[0059] Non-limiting examples of preferred embodiments of the invention are described in more detail below, with reference to Figures 1 to 6.

[0060] Here “Grobler patent” refers to a plant supporting formulation prepared as disclosed in WO 2007 / 096833.

[0061] Example 1 : Process for the manufacture of the plant supporting formulation of the present invention

[0062] In a preferred embodiment, a stable plant supporting formulation comprising an ethyl ester fatty acid-based component, a surfactant component, a vitamin E derivative, an aqueous phase and a gas phase is manufactured. To do so, a formulation according to the present invention is produced starting with the process as set out in the Grobler patent, namely:

[0063] Step 1. A pressure vessel is filled with a desired volume of the aqueous phase, typically reverse osmosis purified water, which is gassed with the indicated gas (in this example nitrous oxide, but it is to be appreciated that the same general procedure with minor modifications may be used when employing carbon dioxide or carbon oxysulfide; all in the zwitterionic form) at 2 kPa using either surface pressure or sparging in the following manner: The vessel is connected to a supply of the gas via a flow control valve and pressure regulator. The closed vessel is supplied with nitrous oxide at a pressure of 2 bar for a period of 24 hours. Although the aqueous phase can be water in unpurified or purified form, it may also be a buffer that can range from pH 3 to 9 depending on the active compounds to be delivered, growth media or even the water component of a plant.

[0064] Step 2. The following fatty acid-based compositions are made up: the ethyl ester fatty acid component is heated to a maximum of 70°C since the double bonds in in unsaturated fatty acids have been found to be sensitive to temperatures exceeding 70°C. The ethyl ester fatty acid component originates from a combination of at least two sources, which may be soy oil, flax seed oil, chia seed oil, hemp seed oil, peppermint oil, rosemary oil, walnut oil, or fish oil. The fatty acids chain length may range from Ci6 to C30. The source determines the structure and the characteristics of the carriers formed - the more unsaturated the fatty acid is, the more sponge-like the structure will be. Ethyl esters of the fatty acids are widely available commercially. In a typical formulation, a total percentage of 3.8 % (v / v of the final plant supporting formulation) of the combination of at least two sources are used to obtain two populations of carriers - one in the nanometer size range and another population in the micro size range as determined by particle size analysis on a Malvern sizer.

[0065] In one possible embodiment of the invention, one such source of ethyl ester fatty acid is a C16 and C18 Vitamin F ethyl ester (also known by the INCI name as ethyl linoleate (and) ethyl linolenate (and) ethyl oleate), an ethyl ester fatty acid obtained from soy (CLR Chemicals Laboratorium Dr Kurt Richter GmbH of Berlin, Germany). The second source of ethyl ester fatty acid in this possible embodiment of the invention is a C20 and C22 long chain Vitamin F ethyl ester product that is manufactured through the extraction and ethyl esterification of a combination of linseed and flax oil (Biopher (Pty) Ltd). However, it is to be appreciated that alternative C20 and C22 long chain ethyl fatty acids may be obtained from fish oil, in particular the ethyl esters of decahexonoic acid and eicosapentanoic acid. In this embodiment, the typical amount of each of the ethyl ester fatty acids is added to a produce a final concentration of 2.7 % of the C16 and C18 ethyl ester fatty acids and 0.1 % of the C20 and C22 ethyl ester fatty acids (v / v of the final plant supporting formulation).

[0066] Step 3. A non-ionic surfactant composed of one or more molecules is heated to 180°C and immediately mixed with the heated ethyl ester fatty acid combination component of Step 2. The ratio of the ethyl ester fatty acid combination component of Step 2 to the non-ionic surfactant in the mixture generally ranges between 2.4:1 and 3.8:1 . The non-ionic surfactant may be selected from natural oils composed largely of ricinoleic acid-based oils with ethylene oxide. The surfactant may be modified as to the extent of hydrogenation, ethylation and the addition of groups such as polyethylene glycol. A range of such products is being marketed by BASF. The surfactant selection may relate to the specific active compound to be delivered by the carriers.

[0067] In one possible embodiment of the invention, the non-ionic surfactant is a pegylated, hydrogenated fatty acid, ricinoleic acid (also known by the INCI name as PEG-n- Hydrogenated Castor Oil).

[0068] Optionally, a second surfactant or co-solvent in the form of: (i) a polysorbate, or a polyethylene glycol (PEG-400) to a final concentration of 2 % (v / v of the final plant supporting formulation) or (ii) a fatty alcohol ethoxylate to a final concentration of 1 % (v / v of the final plant supporting formulation) may be added.

[0069] Step 4. dl-a-Tocopherol or alternatively tocotrienol (a more stable form of tocopherol) is immediately added to the mixture obtained in Step 4 to a final concentration of 0.25 % (v / v of the final plant supporting formulation) to form the oil phase.

[0070] Step 5. The mixture obtained in Step 4 (the oil phase) is then added to the heated aqueous phase of Step 1 and mixed with the aid of a high-speed shearer to a final concentration of between 3.2 % and 4 % (v / v of the final plant supporting formulation), depending on the specific use of the preparation. Cyclical mixing was performed, for example, 2 minutes of mixing, 2 minutes rest; 1 minute mixing, 1 minute rest; 0.5 minutes mixing, 0.5 minutes rest. Should the volumes be upscaled to very large volumes (>500 litres), the mixing and resting times may be extended. This constitutes the final formulation.

[0071] Step 6. The final formulation is cooled rapidly to room temperature (ideally between 15 and 23 °C). This formation may optionally be further subjected to one to two heating (to a maximum of 70°C) and cooling (to room temperature) cycles.

[0072] Step 7. The final formulation is saturated with the preferred gas as provided in Step 1 , for example, in one possible embodiment of the invention, nitrous oxide, for 72 hours. The final formulation is then ready for bottling.

[0073] Formulation for administration to plants

[0074] Step 8. A basic preparation of the final formulation may be diluted with water for administration to plants. For example, dilutions may be 1 :1 for stem application, 1 :10 for ornamentals in open settings, and 1 :5000 in hydroponic systems depending on the method of administration, the type of cultivation (e.g. drip irrigation, foliar spraying by hand, tractor, or plane). Between 60 mL and 120 mL is typically administered per hectare for open field crops and controlled environments such as greenhouses.

[0075] Stable particles of fairly homogeneous sizes ranging from 50nm to 50pm can be manufactured with ease using the method of the invention on a large scale. The size and shape of the particles can be reproducibly controlled.

[0076] Example 2: Typical preparation of a formulation containing a phytologically beneficial substance in the plant supporting formulation according to the invention as a component of a delivery vehicle

[0077] Step 1 One or more phytologically beneficial substances may be entrapped in the basic formulation or buffered formulation preparations described above, by thorough mixing of the desired substance into the formulation at room or field temperature as described in step 8 of Example 1. Mixing may occur by shaking or stirring, or by addition to tank mixers on farms. After mixing, active compounds are generally allowed to be entrapped into the basic formulation or buffered formulation for at least 30 minutes. In the case of phytologically beneficial substances with large molecular weights such as peptides, the mixed formulation is left overnight at 4°C for entrapment.

[0078] Phytologically beneficial substances and active compounds may be added to either the oil phase or the aqueous phase before the process of Example 1 , or to the final plant supporting formulation produced after the process of Example 1 , depending on the physico-chemical properties and the lability of said substances and active compounds. Additives, including amines and polyamines, waxes, plant polymers, saccharides, natural compounds such as amino acids, humic substances, algae extract, pyroligneous extract, tensoactives and surfactants, salicylic compounds, emulsifiers, stabilizers and preservatives may further be added to the finished mixture.

[0079] It is to be appreciated that the method of the invention may be adapted using food, medical and / or veterinary grade ethyl ester fatty acid-based component, surfactant component, vitamin E derivative, aqueous phase and gas phase to produce a formulation that is inherently beneficial for humans and animals. Additionally, in one possible example, an active compound may be combined with the ethyl ester fatty acid-based component and encapsulated in liquid capsules in the final formulation.

[0080] Example 3: Comparison - stabilities of the product produced according to the existing patent compared to the current method when transported to farmers

[0081] Despite very good repeatability of the formulations of the Grobler patent when subject to stability testing as prescribed by the FOA and WHO, it was subsequently determined by the applicant under conditions of field testing that the formulations of the Grobler patent were not able to maintain stability.

[0082] Field testing was performed by collection in the morning of an aliquot of 100 g of either the Grobler patent formulation, or the formulation of the present invention. Aliquots were collected in amber glass bottles from one of ten 25-liter HDPE containers filled with a basic mixture of the Grobler patent formulation or of the formulation of the invention and stored at room temperature, with no special conditions.

[0083] The HDPE containers with either a basic mixture of the Grobler patent formulation or of the formulation of the invention were then packed on the back of a small open truck and transported in a circular route over 500 km tar and dirt roads on a route mimicking delivery to farmers and back to the manufacturing site. A second 100 g aliquot from each of the Grobler patent formulation or the formulation of the present invention was then collected in the afternoon of the same day in a similar fashion from two of the containers; one of which was the same container from which the morning sample was collected, the other from a randomly selected container of the other 9 containers. The collected aliquots were analysed for their particle size, the morphology, their Zeta potential and pH before and after transport.

[0084] In the formulation prepared according to the Grobler patent, the nano-sized component of the plant supporting formulation having a size of > 200nm was completely lost during the transport process. There was also a substantial shift in the size of the micro-sized component of the plant supporting formulation. In contrast, there was very little change in the size of the components of the plant supporting formulation manufactured according to the present invention.

[0085] Differences in the specifications comparing the Grobler patent formulation and the formulation of the present invention are shown below in Table 1 and Figures 1 to 6.

[0086] Table 1 : Comparison between the plant supporting formulation specifications as manufactured in accordance with the Grobler patent or in accordance with the present invention.

Claims

CLAIMS1 . A method for manufacturing a stabilised plant supporting formulation, the method comprising: i. introducing a gas into an aqueous solution in a pressure vessel under pressure to form an aqueous phase; ii. combining a first fatty acid-based component heated to from about 60 °C to about 70 °C and one or more surfactant component(s) heated to from about 100 °C to about 200 °C, or about 180 °C, followed by adding a second fatty acid-based component including a vitamin E derivative, to form an oil phase; iii. heating the aqueous phase of step i. to about 70 °C and then adding the heated oil phase of step ii. to the heated aqueous phase to a concentration of between about 3.2 % and 4 % (v / v of the final stabilised plant supporting formulation) of the oil phase of step ii.; iv. mixing the combined aqueous phase and oil phase of step iii. using a highspeed shearer wherein the combined components are subjected to at least three cycles of mixing and resting where the periods of mixing and resting is incrementally decreased in each cycle to form the stabilised plant supporting formulation; v. optionally subjecting the stabilised plant supporting formulation of step iv. to at least one cycle, or at least two cycles of heating to a first temperature of between about 65 and 70 °C and cooling to a second temperature of between about 15 and 23 °C; and vi. introducing a gas under pressure in a pressure vessel into the stabilised emulsion of step iv. or, when used, step v. until until saturation for a period of about 72 hours to form the stabilised plant supporting formulation.

2. The method according to claim 1 further comprising a final step of bottling the stabilised plant supporting formulation or encapsulating the stabilised plant supporting formulation in a protective coating.

3. The method according to either claim 1 or 2, wherein the aqueous solution comprises purified water, including reverse osmosis purified water; unpurified water; a buffer, including a buffer having a pH of between about pH 3 to 9; a growth medium; or the water component of a plant.

4. The method according to claim 3, wherein the aqueous solution is reverse osmosis purified water.

5. The method according to any one of the preceding claims, wherein the gas is selected from nitrous oxide, carbon dioxide or carbon oxysulfide, all in the zwitterionic form.

6. The method according to any one of the preceding claims, wherein the gas is nitrous oxide.

7. The method according to any one of the preceding claims, wherein the pressure in step i. and step vi. is about 2 kPa (2 bar).

8. The method according to any one of the preceding claims, wherein the pressure in step i. is introduced under pressure for a period of about 24 hours.

9. The method according to any one of the preceding claims, wherein the fatty acidbased component is an ethyl ester fatty acid-based component.

10. The method according to claim 9, where in the ethyl ester fatty acid comprises about 1 to 5 % v / v of the fatty acid-based component.11 . The method according to either claim 9 or 10, wherein the ethyl ester fatty acid component is selected from the group comprising any two or more ethyl ester fatty acid sources that are combined, the sources comprising ethyl esters of soyoil, flax seed oil, chia seed oil, hemp seed oil, peppermint oil, rosemary oil, walnut oil, or fish oil.

12. The method according to any one of claims 9 to 11 , wherein the ethyl ester fatty acid component chain length ranges from Ci6 to C30.

13. The method according to any one of claims 9 to 12, wherein the ethyl ester fatty acid component chain length ranges from C16 to Cis from a first ethyl ester fatty acid source, and from C20 and C22 from a second ethyl ester fatty acid source.

14. The method according to any one of claims 9 to 13, wherein the combined ethyl ester fatty acid component has a concentration of about 3.8 % (v / v) of the final plant supporting formulation.

15. The method according to either claim 13 or 14, wherein the first source of ethyl ester fatty acid is a C16 and C18 Vitamin F ethyl ester obtained from soy and the second source of ethyl ester fatty acid is a C20 and C22 long chain Vitamin F ethyl ester product manufactured through the extraction and ethyl esterification of a combination of linseed and flax oil.

16. The method according to either claim 13 or 14, wherein the first source of ethyl ester fatty acid is a C16 and C18 Vitamin F ethyl ester obtained from soy and the second source of ethyl ester fatty acid is a C20 and C22 long chain ethyl fatty acid obtained from fish oil, selected from the ethyl esters of decahexonoic acid and eicosapentanoic acid or a combination thereof.

17. The method according to any one of claims 13 to 16, wherein the first and second sources of ethyl ester fatty acid are added to a produce a final concentration of about 2.7 % of the C16 and C18 ethyl ester fatty acids and about 0.1 % of the C20 and C22 ethyl ester fatty acids (v / v) of the final plant supporting formulation.

18. The method according to any one of the preceding claims, wherein one or more non-ionic surfactant component(s) are one or more surfactant molecule(s), optionally two non-ionic surfactant components, selected from the group comprising natural oils composed largely of ricinoleic acid-based oils with ethylene oxide, and including a surfactant modified as to the extent of hydrogenation, ethylation and the addition of groups; and including polyethylene glycol; or a polysorbate; or a polyethylene glycol (PEG-400); or a fatty alcohol ethoxylate; or any combination thereof.

19. The method according to claim 18, wherein the one or more non-ionic surfactant component(s) is a pegylated, hydrogenated fatty acid, ricinoleic acid also known as PEG-n-Hydrogenated Castor Oil.

20. The method according to claim 18, wherein a first of the one or more non-ionic surfactant component(s) is selected from the group comprising natural oils composed largely of ricinoleic acid-based oils with ethylene oxide, and including a surfactant modified as to the extent of hydrogenation, ethylation and the addition of groups and including polyethylene glycol or any combination thereof; and a second of the one or more non-ionic surfactant component(s) is selected from the group comprising a polysorbate, a polyethylene glycol (PEG-400), or a fatty alcohol ethoxylate, or any combination thereof.

21. The method according to any one of the preceding claims, wherein the ratio of first fatty acid-based component of step ii. to the one or more non-ionic surfactant component(s) range between about 2.4:1 and 3.8:1 .

22. The method according to any one of the preceding claims, wherein the vitamin E derivative is selected from dl-a-Tocopherol or a physiologically active isomer thereof, including tocotrienol.

23. The method according to any one of the preceding claims, wherein the vitamin E derivative is added to form the oil phase in step ii. to obtain a final concentration of vitamin E derivative of about 0.25 % (v / v) of the final plant supporting formulation.

24. The method according to any one of the preceding claims, wherein the mixing in step iv. is performed by a first cycle of at least 2 minutes of mixing and at least 2 minutes rest; a second cycle of at least 1 minute mixing and at least 1 minute rest; followed by a third cycle of at least 0.5 minutes mixing and at least 0.5 minutes rest.

25. A stabilised plant supporting formulation produced according to any one of claims 1 to 24.

26. The stabilised plant supporting formulation according to claim 25 having a plurality of vesicles and / or microsponges within the stabilised plant supporting formulation with a size of less than about 200 nm.

27. The stabilised plant supporting formulation according to either claim 25 or 26, comprising:I. a stabilised micro-emulsion comprising a dispersion of vesicles and / or microsponges of the oil phase in the aqueous phase; andII. a gas dissolved in the oil phase.

28. The stabilised plant supporting formulation according to any one of claims 25 to 27 for use in any one or more of the following applications: a. delivering one or more compound(s) selected from the group comprising one or more active compounds and / or one or more compounds beneficial for plant growth and / or health to a plant; b. improving the absorption of the one or more compound(s) delivered to the plant;c. improving distribution and translocation in a plant of the one or more compound(s); d. facilitating release of the one or more compound(s) delivered by the stabilised plant supporting formulation to a plant cell; or e. supporting plant growth and / or health by improvement of energy metabolism of the plant.

29. The stabilised plant supporting formulation according to any one of claims 25 to 28, further comprising additives selected from tensoactives, additional surfactants, salicylic compounds, emulsifiers, stabilizers or preservatives.

30. A method of treating a plant or supporting the growth and / or health of a plant, comprising any one or more of the following:A. delivering one or more compound(s) to a plant selected from one or more active compounds and / or or one or more compound(s) beneficial for plant growth and / or health to a plant;B. improving the absorption of the one or more compound(s) delivered to the plant;C. improving the distribution and translocation in a plant of the one or more compound(s) delivered to the plant;D. releasing to plant cells the one or more compound(s) delivered to the plant; orE. supporting plant growth and / or health by improvement of energy metabolism of the plant, the method comprising the use of the stable plant, wherein the method comprises the use of the stabilised plant supporting formulation of any one of claims 25 to 29.

31. The method according to claim 30, wherein the one or more compound(s) are comprised of one or more active compound(s) used for treatment of plant conditions or diseases, phytologically beneficial substances, amines and polyamines, waxes, plant polymers, saccharides, natural compounds including amino acids, humic substances, algal extract, or a pyroligneous extract.

32. The method according to either claim 30 or 31 , wherein the one or more compound(s) are added to the oil phase or the aqueous phase, or both the oil phase and the aqueous phase, or to the final stabilised plant supporting formulation.

33. The method according to any one of claims 30 to 32, wherein the one or more compound(s), including phytologically beneficial substance(s) comprise or are selected from the group consisting of: a plant pesticide including a herbicide, fungicide, bactericide, insecticide, antiplant virus agent; a plant growth regulator selected from the products in the group consisting of, 2-(1 -2- methylnaphthyl)acetamide; 2-(1 -2-methylnaphthyl)acetic acid; 2-(1 - naphthyl)acetamide; 2-(1 -naphthyl)acetic acid; 2,4-D (sodium salt); 3,5,6 TPA; 4-indol-3-ylbutyric acid; 6-benzyl adenine; alkoxylated fatty alkylamine polymer; alkylamine polymer; aminoethoxyvinylglycine hydrochloride; ammoniated nitrates; auxins; calcium arsenate; carbaryl; chlormequat chloride; chlorpropham; chlorthal-dimethyl; cloprop; cyanamide; daminozide; decan-1 -ol; dichlorprop; dichlorprop (2-butoxyethyl ester); dimethipin; dinocap; diquat dibromide; diuron; ethephon; fluazifop-p-butyl; gibberellins; glyphosate- isopropylamine; glyphosate-trimesium; haloxyfop-P-methyl; indolylacetic acid; maleic hydrazide; mepiquat chloride; methylcyclopropene; mineral oil; n- decanol; octan-1 -ol; paclobutrazole; paraquat dichloride; pendimethalin; prohexadione-calcium; salicylic acid, sodium chlorate; thidiazuron; trinexapac- ethyl; and uniconazole; a plant immune modulator; and a biostimulant comprising the group of products known as phytohormones, or any combination thereof.

34. A stabilised plant supporting formulation according to any one of claims 25 to 29, comprising one or more compound(s) selected from the group comprising: one or more active compound(s), and / or or one or more compounds beneficial for plant growth and / or health to a plant.

35. A method of administration of the stabilised plant supporting formulation according to any one of claims 25 to 29 or claim 34 to a plant comprising a step of diluting the stabilised plant supporting formulation with a diluent solution suitable for administration to the plant, including water.

36. The method according to claim 35, wherein the ratio of stabilised plant supporting formulation to the diluent solution is about 1 :1 for stem application, about 1 :10 for ornamentals in open settings including in the field or in greenhouses, and about 1 :5000 for hydroponic systems.

37. The method according to either claim 35 or 36, wherein where the one or more compound(s) selected from one or more active compounds and / or one or more compounds beneficial for plant growth and / or health are added to the final stabilised plant supporting formulation, the method of administration further comprises a step of entrapment of the one or more compound(s) in the stabilised plant supporting formulation comprising:(aa) addition of the one or more compound(s) to the stabilised plant supporting formulation either prior to dilution or after dilution;(bb) mixing the stabilised plant supporting formulation and one or more compound(s);(cc) leaving the mixture of step (bb) to stand for at least about 30 minutes to overnight for entrapment.

38. The method according to claim 37, wherein where the mixing of step (bb) is performed at room or field temperature, including by shaking or stirring, or by addition to a tank mixer on site.

39. The method according to claim 37, wherein where the one or more compound(s) are phytologically beneficial substances with large molecular weights including peptides, step (cc) includes storing the mixture of (bb) overnight at 4°C for entrapment.

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