Bioactive agrichemical compositions and use thereof.

(54) Title: BIOACTIVE AGROCHEMICAL COMPOSITIONS AND USES OF THE SAME. (54) Title: BIOACTIVE AGRICHEMICAL COMPOSITIONS AND USE THEREOF.

(57) Summary

Concentrates and bioactive agrochemical compositions having improved bioactivity comprising metal-acid solutions are described.

(57) Abstract

Bioactive agrichemical concentrates and compositions having improved bioactivity comprising combinations of acid Solutions and conventional bioactive agrichemical actives or formulations.

Institute

Mexican Property

Industrial

<img file="MX336953B_D0001.tif" />

I KNOW

-k 3 &lt;»

PATENT TITLE NO. 336953

Headlines): AGION TECHNOLOGIES, INC.

Address: 60 Audubon Road, Wakefield, Massachusetts, 01880, USA

Name: BIOACTIVE AGROCHEMICAL COMPOSITIONS AND USES OF THE SAME. Classification: lnt.CI.8: A01N59 / 16; A01N59 / 20; A01P1 / 00

Inventors): JOSEPH J. CRUDDEN

REQUEST

Number:

MX / a / 2015/002941

International filing date:

May 2008

Divisional Patent Number: 328710

<td colspan="3">PRIORITY</td>

<td>Country:</td><td>Date:</td><td>Number:</td>

<td>US;</td><td>May 18, 2007</td><td> 60/930,913</td>

<td>T Validity: Twenty years</td><td></td><td></td>

Expiration Date »: November 19, 2028

The reference patent grants you can based on / articles 1, 2 ° section V, 6 ° section III, and 58 of the Industrial Property Law ® is &lt;'piafad IMbstnaL this patent has a validity of twenty years imp expendable, request nMntfNHnal and will be subject to the payment of the fee to maintain 'before the

In accordance with article 23 ddfja Law of the Pro¡ counted from the date of presentation of the rights. .

Whoever subscribes to this title does so on the basis imposed by the i

Industrial Property (Official Gazette of the Federation (DO- 2796/1991 «Bfonfiada al i

01/26/2004, 06/16/2005, 01/25/2006 j06 / 05 / 2009,06 / 01/2010, 06/18 ^ 1¾¾ ^ ¾¾¾ ^ ¾¾¾ ^ ¾ ^ 04/09/2012); articles 1, 3 »section V: subsection a), 4 'and 12 ° sections I and llfjdel the Regulations of the Mexican Institute of Industrial Reform (DOF 12/14/1999, r^s9V9Tff9.BrjJÍ (P7CT9li..l ° '^n?'^or1l7Wii0nⁱr) »eSéaete ^ fc ^ * fc« * &lt;ffrTr ¥ *ⁱTri'TT) r^t*^tJ* laugh^,TnrtÍT ^ iieilMrifleÍlClll.MJ Organic of the Mexican Institute of Industrial Property (DOF 12/27/1999, amended on 10/10/2002, 07/29/2004, 08/04/2004 and 09/13/2007); 1, 3 and 5 subsection a) of the Agreement that delegates powers to the Deputy Directors General, Coordinator, Divisional Directors, Holders of Regional Offices, Divisional Deputy Directors, Departmental Coordinators and other agents of the Mexican Institute of Industrial Property. (DOF 12/15/1999, amended on 02/04/2000, 07/29/2004, 08/04/2004 and 1.3 / 09 {2fÍQ7) .:

and 7 ° bis 2 of the Law of i / 10/1996, 26/12/1997, / 05/1999,

<img file="MX336953B_D0002.tif" />

Issue Date: February 3, 2016

DIVISIONAL DIRECTOR OF PATENTS

<img file="MX336953B_D0003.tif" />

NAHANNY CANAL REYES

Arenai No. 550, Press 1,

Coi. Puebb Santa María Tepepan. Xochífttíico. CP 16020,

Mexico City ϊ © I. (55) 53 34 07 0C¹ wuv.^¡ tmpi gob.arx

<img file="MX336953B_D0004.tif" />

<img file="MX336953B_D0005.tif" />

MX / 2016/23332 Cüs / ZoiJf / 2 ° tQl [

<img file="MX336953B_D0006.tif" />

BIOACTIVE AGROCHEMICAL COMPOSITIONS AND USES. FIE TAS-SAME-FIELD OF THE INVENTION

<img file="MX336953B_D0007.tif" />

The present invention relates to novel bioactive agrochemical compositions comprising a combination of a) an acidic solution having low levels of bioactive metal ion, alone or, preferably, in additional combination with one or more surfactants that can interact with the cell wall membranes of microorganisms, especially pathogenic microbes and b) a conventional agrochemical active, especially a fungicidal active, either as the pure active ingredient or as a formulated agrochemical composition. These bioactive agrochemical compositions can be used for any number of agrochemical applications that require the control, inhibition, and / or annihilation of microorganisms, especially fungi, bacteria, and / or protists, plant type, stramenophilic type, and fungus type.

BACKGROUND OF THE INVENTION

Bioactive materials to kill or inhibit the growth and / or proliferation / spread of bacteria,

IMPI

<img file="MX336953B_D0008.tif" />

Fungi, and other microorganisms have long been sought and used in society. Its uses date back several centuries, if not thousands of years. Initial applications have ranged from pharmaceutical or health-related applications to disinfectant and purification applications and more. The most recent applications include a large number of uses, with the largest use, by volume, being observed in the agricultural industry. Perhaps one of the earliest bioactive materials was metallic silver and, later, silver salts.

Although the initial bioactive agents were most often metals and simple metal salts, modern science and chemical synthesis have allowed the development and production of synthetic agents, most often organic and organometallic agents, for antibacterial, antifungal, and other similar applications. .

Indeed, for many applications, especially pharmaceutical applications, organic agents have, for the most part, overshadowed the use of inorganic bioactive agents. Although inorganic and organometallic materials continue to command a large part of the agrochemical business market, their use is limited due to their health and safety concerns, especially from an environmental perspective. Indeed, organic bioactive agents dominate a portion

MEXICAN INSTITUTE OF I “EQPiEDAD

INDUSTRIAL

<img file="MX336953B_D0009.tif" />

huge business of agrochemicals.

Despite the great success and immense volume / section of the market governed by pharmaceutical, antibacterial and organic agrochemical agents, they do not. they have arrived without costs and consequences. In all application areas, a marked and growing trend has emerged: specifically the manifestation and spread of resistance to such organic agents in most, if not all, microorganisms. Although this resistance is neither universal nor complete, it is growing and involves more and more organic agents. As its resistance grows, so does its apparent virulence. In this sense, there is knowledge of the increasing resistance of bacteria, especially pathogenic bacteria, to traditional pharmaceutical antibiotic agents and the subsequent appearance of what is commonly referred to as super-pests: pathogenic bacteria that show strong resistance to organic antibacterial and pharmaceutical agents. traditional.

And, be it a direct or indirect consequence of the appearance of super plagues and / or the increasing knowledge of the ease with which bacteria can spread combined with an increasing concern regarding potentially pandemic diseases such as

<img file="MX336953B_D0010.tif" />

<img file="MX336953B_D0011.tif" />

SARS and bird flu, the population has become more and more concerned with hygiene and general cleanliness. Consequently, there has been an immense proliferation and exponential growth in the indiscriminate and widespread use and application of cleaners and disinfectants containing organic antimicrobial agents, all in an effort to prevent exposure to bacteria and especially super plagues. However, this indiscriminate use of organic agents has come with, or at least presents the possibility of, a general increase in antimicrobial resistant organisms. By eradicating the weakest organisms, the strongest and most often the most harmful organisms remain.

A similar consequence has also manifested itself in the agricultural industry, especially in that portion regarding crop / food production. The widespread and repetitive use of organic biocidal agents, fungicides, antibacterials and the like, has led to the manifestation of less and less efficacy of them against the target diseases: an indicator of increasing resistance. Perhaps more alarming is the speed with which such resistance has begun to appear. For example, despite the great fanfare and promises behind the introduction of fungicides

<img file="MX336953B_D0012.tif" />

<img file="MX336953B_D0013.tif" />

of strobilurin in the mid-1990s, resistance was found only after a few years of use in some applications. Such an increasing trend is a bad sign for an industry in which less and less hectares are occupied to produce more and more crops to feed the ever-growing populations while those organisms and microorganisms responsible for attacking these crops become stronger and stronger. and increasingly resistant to traditional means of control.

Although resistance is certainly a major concern, the human and environmental cost associated with the widespread use of organic antimicrobial agents is perhaps even more worrisome. For more than half a century so far, more and more scientific literature has appeared that correlates long-term exposure (direct and indirect) and the use of such organic agrochemicals with various diseases and teratogenic, mutagenic, and other adverse health consequences. in animals, and more importantly, in the human population.

Perhaps the crux of this awareness is represented by the protest regarding the use of DDT and similar pesticide agents in the 1960s. In humans, such a correlation of birth defects, cancer, and other diseases with agrochemicals is especially puzzling to those

<img file="MX336953B_D0014.tif" />

<img file="MX336953B_D0015.tif" />

whose water supplies are or could be contaminated with said organic agents due to their solubility and / or solubility of their by-products and the long half-life times. Of course, drinking water is only one source of exposure: another source of exposure concern is inhalation of dust spread from fields, whimsical aerosols and / or particulate matter during aerial spraying or dusting, respectively, and exposure to workers' uniforms, who themselves are exposed in the fields or during application.

In an effort to move away from organic agents, more recent attention has again been focused on inorganic agents, including organometallic agents, because they tend not to show or do not have, or certainly less tend to result in bacteria. , resistant fungi and the like. However, this trend only reawakens the debates and concerns regarding the large-scale discharge of heavy metals into the environment. Although some efforts have recently focused on improving old, traditional inorganic agents, apparently much more effort, particularly in the non-agricultural area, has focused on more complex species and systems, in

<img file="MX336953B_D0016.tif" />

Essence, synthesized inorganic biocides such as zeolite, hydroxyapatite, and zirconium phosphate based antimicrobial ion exchange type antimicrobial agents. Other recent biocides include those based on electrolytically generated silver citrates; thiol-free specialized antimicrobial metal complexes; sulfuric acid / sulfate complexes prepared under elevated pressure and temperature; and the like. Although effective, they have limited applications and involve additional costs due to the complex and / or lengthy synthesis procedures by which they are prepared. The latter is of special concern in the agricultural industry where the balance between relative cost and performance often avoids the use of functionally very viable options. In this case, a few cents per hectare difference, even a difference of a fraction of a cent per hectare, can mean a world of difference in the acceptability and utility of a given agent.

Despite its inherent environmental and health problems, the use of natural inorganic agents, including manufactured / processed inorganic agents, has been pressured more and more by various environmental and conservation groups, as well as by health advocates such as a

b .. '

MEXICAN INSTITUTE V INDUSTRIAL PROPERTY

<img file="MX336953B_D0017.tif" />

favorable replacement for organic agents. While such pressure alone is unlikely to change the industry, the increasing resistance to organic agents combined with the higher and higher costs of synthetic organic agents is certainly having an impact:

not only in the agricultural industry but in general in all applications for such bioactive agents. However, as indicated above, the reintroduction of and / or increased use of inorganic agents only brings to the fore the same issues that at first made them relegate, specifically environmental toxicity and contamination and bioaccumulation. The concern is not only with the effects during application, but rather with the long-term effects associated with the continuous accumulation of these inorganic agents or their derivatives, especially metals, in the environment and in living organisms. Said accumulation belongs not only to the soils that are treated but also to the groundwater supplies that can be replenished from the treated fields.

There are also concerns, sometimes more or less, about the consequences of rainwater running off carrying metals into local streams and, again, downstream water supplies. Finally, this accumulation also occurs in the food chain, 'Ρ'.Ατί⁵*·

-LL 'WST / TUTOMSxl' '.

OF THE PiiOPiíbAü í- &lt;

'NDUSTSIAL and *

<img file="MX336953B_D0018.tif" />

finding higher and higher concentrations in those species in the higher order of the food chain. Finally, this affects the human supply chain as observed, for example with mercury and other metals in tuna and swordfish. Although many metals, at least at low exposure levels, have little or no effect on humans, their impact is much greater on marine life and other aquatic life, especially fish, which tends to be extremely sensitive to heavy metals. , like silver, which results in increased stress and, in situations of extreme exposure, widespread annihilation.

Consequently, as part of this reappearance of inorganic bioactive agents, there has been a significant increase in the money and time invested in research and development to address problems related to the use of such inorganic agents. A first point of attention has been with respect to making more concentrated materials that, it is hoped, allow the use of - less general materials. In the field of agrochemicals, one of the inorganic agents that receives the most attention, due to its high efficacy, is copper. Indeed, it is believed that copper could observe a multiple increase in use due to the fact that organic agents are excluded from use or

<img file="MX336953B_D0019.tif" />

ΪΓ Ό Τ

MEXICAN

PROPERTY? JE-'J3TRiAL

<img file="MX336953B_D0020.tif" />

farmers opt for more natural agents Cor. NCFAP data, in 1997 over 6,214 tons of copper were used in fungicide applications compared to 18,144 tons of synthetic fungicides at a treatment rate of about half that of copper. If all synthetic fungicides had to be supplanted by copper, this could have resulted in an increase of more than 36,288 tons of copper released into the environment. Now, ten years later, although the data is not available, it can only be assumed that the amounts are much higher. Furthermore, this is only a use of copper: copper and copper-based bioactive compositions are also used in other areas such as alcicides, etc. Regardless, it is evident that any significant change from synthetic fungicides to fungicides, copper algicides, etc. means an immense

<td rowspan="2">impact and</td><td colspan="2">release</td><td rowspan="2">coppermade Indian</td><td colspan="3">in the environment.</td>

<td>How</td><td>I know</td><td>previously,</td><td>the</td><td>recent</td>

<td>efforts</td><td>in</td><td colspan="2">Investigation</td><td>and development</td><td>with</td><td>fungicides</td>

<td colspan="2">inorganic it</td><td>have</td><td>in focus</td><td colspan="2">in the development</td><td>agent s</td>

Improved inorganics that produce better effects with less application. Indeed, -in August 2006, DuPont, one of the leading manufacturers of agrochemicals, especially copper-based fungicides, announced certain important advances, such as

<img file="MX336953B_D0021.tif" />

they described it, in copper fungicides, specifically its copper hydroxide based fungicide

Kocide 3000, promoting its ability to provide more antifungal action with less copper. However, its typical application rate is in the order of 4,077 grams of copper per hectare per application, with slightly lower rates, 815.4 grams per hectare, being allowed for certain applications.

Although it is certainly an improvement over conventional or traditional copper-based fungicides which are applied at almost 5.04 kg per hectare, it still means the intentional release of huge amounts of copper into the environment, even more so if environmentalists are successful in eliminate or prohibit the use of more and more organic agents.

Therefore, there is still a huge need for cost effective inorganic agrochemical agents that provide good antimicrobial, antifungal, antibacterial, etc. activity, without the problem of resistance build-up.

Therefore, there is a need for inorganic antimicrobial, antifungal, antibacterial, etc. agents that can be used universally, or almost universally, without cause for concern, or certainly with reduced concern regarding

<img file="MX336953B_D0022.tif" />

contamination and environmental toxicity, especially less than that which exists with current inorganic agents.

Similarly, there is a need for inorganic agents that are stable and easy to use, and that provide good short-term and preferably long-term efficacy compared to many of today's short-lived organic agents.

Additionally, there is a need for such inorganic agents that can be safely used in agricultural and horticultural applications, including soil and seed treatment, crop / food producing plants and trees, ornamental and flower plants and trees, feed and decoration grasses, and the like with minimal exposure problems.

Also, there is a need for inorganic bioactive agents that can be used in combination with active agents and inorganic and preferably conventional organic agrochemical compositions, especially antimicrobial, antifungal, antibacterial, antiprotista, etc. with · synergistic results; that, therefore, allow less general use of said assets.

Lastly, there is a need for bioactive agents that provide effective antimicrobial, antifungal, antiprotista, and / or antibacterial performance with release.

IMPI

MEXICAN INSTITUTE OF INDUSTRIAL LAUNDRY

<img file="MX336953B_D0023.tif" />

minimal inorganic metals into the environment

SUMMARY OF THE INVENTION

In accordance with one aspect of the present invention there are provided bioactive agrochemical concentrates in solid form comprising a) a conventional bioactive agrochemical active in solid form, either in its pure form or a formulated form and b) a solid bioactive acid composition comprising an acid, especially a weak or moderate acid, at least one source of at least one antimicrobial metal ion, and, optionally, although preferably, at least one surface active agent, especially at least one anionic, nonionic and / or amphoteric surface active agent, that affects or interacts with the cell wall membranes of microorganisms, especially pathogenic microbes, or the function thereof, said acid is present at least 40 percent by weight, preferably 40 to 80 percent, based on the total weight of the bioactive acid composition, and at a level representing at least a 2-fold molar excess relative to the antimicrobial metal ions in the source, said bioactive acid composition has a pH of less than 6, preferably approximately 1.5 to 5, when

<img file="MX336953B_D0024.tif" />

<img file="MX336953B_D0025.tif" />

it is diluted in a solvent, especially water, to a point where the amount of antimicrobial metal ion is 500 ppm or less in the case of a single ion or 1000 ppm or less in the case of multiple antimicrobial metal ions.

Generally speaking, the weight ratio of the conventional bioactive agrochemical to the bioactive acid composition is such that when the concentrate is diluted or lowered to the formulation to be applied, the amount of antimicrobial metal ion, based on the Antimicrobial metal ions of the bioactive acid composition, is 500 pprr. or less in the case of an individual antimicrobial ion or

1000 ppm or less in the case of multiple antimicrobial metal ions and the conventional bioactive agrochemical active is present at or near its conventional application amount or less. These solid concentrates can be lowered by dry mixing with diluents or solid fillers prior to application, or they can be converted to liquid using diluents or liquid fillers.

In accordance with a second aspect of the present invention, bioactive agrochemical concentrates in liquid form are provided comprising a) a conventional bioactive agrochemical active, either in its pure form or a formulated form, and b) a bioactive acid solution

<img file="MX336953B_D0026.tif" />

<img file="MX336953B_D0027.tif" />

aqueous or aqueous-based comprising a concentrated acid solution, especially a weak or moderate acid, at least one antimicrobial metal ion or antimicrobial metal ion source completely or partially dissolved in said acid solution and, optionally, although preferably, at least one surfactant, especially at least one anionic, nonionic and / or amphoteric surfactant, that affects or interacts with the cell wall membranes of microorganisms, especially pathogenic microbes, or their function, in which the concentration of the acid, based on the total weight for the bioactive acid solution, is at least approximately 40 percent by weight, preferably 40 to percent, of acid and the acid is present at a level that is at least a 2-fold molar excess relative to the antimicrobial metal ion (s), and in which, the pH of the bioactive acid solution is less than 6, preferably 1.5 to 5, when the concentrated bioactive acid solution is diluted in a solvent, especially water, to a point where the amount of antimicrobial metal ion is 500 ppm or less in the case of a single ion or 1000 ppm or less in the case of multiple antimicrobial metal ions. Preferably, the conventional agrochemical active dissolves in or is miscible with the

Ό τ .vi Λ ji MEXICAN INSTITUTE OF Property

INDUSTRIAL

<img file="MX336953B_D0028.tif" />

concentrated bioactive acid solution or in a solvent miscible with the bioactive acid solution. Alternatively, in cases where the conventional agrochemical active is insoluble in water or in the water-based solvent, it can be dissolved in a water-immiscible solvent and the concentrate exists as an emulsion or suspension. In general terms, the weight ratio of the conventional bioactive agrochemical to the bioactive acid composition is such that when the concentrate is diluted or lowered to the formulation to be applied, the amount of antimicrobial metal ion, based on the ions of antimicrobial metal of the bioactive acid composition, is 500 ppm or less in the case of a single antimicrobial ion or

1000 ppm or less in the case of multiple antimicrobial metal ions and the conventional bioactive agrochemical active is present at or near its conventional application amount or less. These liquid concentrates can be lowered by mixing with an appropriate solvent, particularly water or a water-based solvent, for application. Alternatively, the concentrate can be applied as such or in a diluted state to a solid absorbent material for application.

In accordance with a third embodiment of the present invention, agrochemical compositions are provided.

<img file="MX336953B_D0029.tif" />

<img file="MX336953B_D0030.tif" />

fluid particulate bioactives, for example, dusts, granules, powders, or combinations thereof, comprising a substantially homogeneous mixture of a) a conventional bioactive agrochemical in particle form and b) a solid carrier particle which has been treated with a bioactive acid solution, especially an aqueous or aqueous-based solution, which has a pH less than 6, preferably 1.5 to 5, and which comprises an acid, especially a weak or moderate acid, at least one antimicrobial metal ion source, and, optionally, though preferably, at least one surfactant, especially at least one anionic, nonionic and / or amphoteric surfactant, which affects or interacts with the cell wall membranes of microorganisms, especially pathogenic microbes, or their function, said acid present in a molar excess, relative to antimicrobial metal ions, in which the amount of antimicrobial metal ion is 500 ppm or less in the case of a single ion or 1000 ppm or less in the case of multiple antimicrobial metal ions. This embodiment also contemplates that the solid carrier material treated with the bioactive acid solution is the conventional bioactive agrochemical composition itself, for example, in cases where the bioactive active is present as an active

<img file="MX336953B_D0031.tif" />

Formulated in an appropriate absorbent or adsorbent vehicle or is present as an active granule and a solid filler or diluent. Generally speaking, the formulation of the bioactive agrochemical composition of the invention will be such that, when applied during use, the application rate or amount of the conventional bioactive agrochemical is at or near the conventional application amount or less and the amount applied antimicrobial metal ion or ions originating from the bioactive acid solution will be no more than approximately 1,235.5 grams per hectare, preferably not more than 617.8 grams per hectare.

In accordance with a fourth embodiment of the present invention, fluid bioactive particulate agrochemical compositions are provided, for example, powders (dusts), granules, powders, or combinations thereof, comprising a substantially homogeneous mixture of a) a conventional bioactive agrochemical in particle form and b) a bioactive acid composition in particle form, preferably a powder, said bioactive acid composition is prepared by i) forming an aqueous solution of an acid, especially a weak or moderate acid, at least one antimicrobial metal ion source, and, optionally, but preferably, at least one agent surfactant,

<img file="MX336953B_D0032.tif" />

<img file="MX336953B_D0033.tif" />

especially at least one anionic, nonionic and / or amphoteric surface active agent, which affects or interacts with the cell wall membranes of microorganisms, especially pathogenic microbes, or the function thereof, said acid present in a molar excess, with relative to antimicrobial metal ions, in which the amount of antimicrobial metal ion is 500 ppm or less in the case of a single ion or 1000 ppm or less in the case of multiple antimicrobial metal ions, ii) allowing the water to evaporate to form the solid bioactive acid and iii) if necessary, grinding or crushing the solid bioactive acid formed to form a powder. As with the previous embodiment, the formulation of the bioactive agrochemical composition of the invention will be such that, when applied during use, the application rate or amount of the conventional bioactive agrochemical is at or near the conventional application amount or less and the applied amount of the antimicrobial metal ion or ions originating from the bioactive acid solution shall be not more than approximately 1,235.5 grams per hectare, preferably not more than 617.8 grams per hectare.

In accordance with a fifth embodiment of the present invention, liquid bioactive agrochemical compositions are provided comprising a) a solvent or system

<img file="MX336953B_D0034.tif" />

of appropriate solvents, b) a conventional bioactive agrochemical active dissolved in said solvent or solvent system, c) an acid, especially a weak or moderate acid, d) at least one source of antimicrobial metal ion, and e) optionally, although preferably, at least one surfactant, especially at least one anionic, nonionic and / or amphoteric surfactant, that affects or interacts with the cell wall membranes of microorganisms, especially pathogenic microbes, or the function thereof, in which the acid and solvent solution has a pH less than 6, preferably

1.5 to 5, before, and preferably also after, the addition of the conventional bioactive active agent, said acid is present in a molar excess relative to the antimicrobial metal ions of the source (d) and in which the amount of antimicrobial metal ions attributed to the source (d) is 500 ppm or less in the case of a single ion or 1000 ppm or less in the case of multiple antimicrobial metal ions. More preferably, the solvent is water or a water-based solvent system, more preferred water, and the conventional bioactive active, acid, and antimicrobial metal ion source are all fully or substantially soluble or miscible in the solvent. In cases where the bioactive asset

<img file="MX336953B_D0035.tif" />

DS LA INSTITUTE

INjJ J.

<img file="MX336953B_D0036.tif" />

Conventional is not soluble or miscible in water or in a water-based solvent, it can be dissolved in an appropriate non-aqueous solvent, including a lipophilic solvent, and the metal source and acid are dissolved in water or a water-based solvent and the two solutions combine to form a suspension or emulsion. Generally speaking, this formulation of the bioactive agrochemical composition of the invention will be such that, when applied during use, the application rate or amount of the conventional bioactive agrochemical is at or near the conventional application amount or less and the amount applied of the antimicrobial metal ion or ions originating from the bioactive acid solution will be no more than approximately 1,235.5 grams per hectare, preferably not more than 617.8 grams per hectare.

In accordance with a sixth embodiment of the present invention there is provided a method for making a bioactive agrochemical concentrate in solid form comprising dry mixing a) a conventional bioactive agrochemical active in solid form, either in its pure form or in a formulated form , b) an acid, especially a weak or moderate acid, c) at least one source of at least one antimicrobial metal ion, and d) optionally, but preferably, at least one surfactant,

<img file="MX336953B_D0037.tif" />

especially at least one anionic, nonionic and / or amphoteric surfactant, which affects or interacts with the cell wall membranes of microorganisms, especially pathogenic microbes, or the function thereof, said acid being present in at least 40 percent by weight, based on the total weight of the acid and the antimicrobial metal ion source, and at a level that

<td>represent</td><td>by</td><td>least</td><td>a molar excess of</td><td> 2</td><td>times</td><td>with</td>

<td>relation to</td><td>the</td><td>ions of</td><td>antimicrobial metal</td><td>of</td><td colspan="2">the fountain,</td>

<td>the acid</td><td>this</td><td>Present</td><td>in an amount in</td><td>the</td><td>which</td><td>a</td>

<td colspan="2">solution that</td><td>prepare</td><td>adding the acid and</td><td>the</td><td>source</td><td>of</td>

Antimicrobial metal ions to water have a pH less than 6, preferably about 1.5 to 5, when diluted to a point where the amount of antimicrobial metal ion is 500 ppm or less in the case of a single ion or 1000 ppm or less for multiple antimicrobial metal ions. Preferably the acid, antimicrobial metal ion source and, if present, the surfactant are pre-mixed before combining with the conventional fungicidal active. More preferably, these ingredients are first dissolved in an appropriate volatile solvent, especially water or a water-based solvent, and then the solvent is allowed to evaporate to leave a cake which is then combined with or, preferably, first ^ ΡΙ

MEXICAN INSTITUTE OF THE FRO.'ISDAD INDUSTRIAL

<img file="MX336953B_D0038.tif" />

ground or crushed and then combined with the conventional bioactive active.

In accordance with a seventh embodiment of the present invention there is provided a method for making a liquid bioactive agrochemical concentrate, which method comprises forming a concentrated aqueous or aqueous-based acid solution of an acid, preferably a weak or rounded acid, in which the acid concentration is at least

40 wt%, dissolve in the concentrated acid solution at least one antimicrobial metal ion source and optionally, but preferably, at least one water soluble surfactant, preferably an anionic, nonionic surfactant, and / or amphoteric, and then dissolve in said concentrated bioactive acid solution a conventional bioactive agrochemical active which is soluble or miscible in said bioactive acid solution or, if not, it is dissolved in another suitable solvent and the two solutions combine to form an emulsion or suspension of the concentrate.

In accordance with an eighth embodiment of the present invention, a method is provided to prevent or inhibit the growth of plant pathogens, especially fungi, bacteria and / or protists, plant type, estramenophilic type and fungus type in agricultural applications,

<img file="MX336953B_D0039.tif" />

Ιί-

<img file="MX336953B_D0040.tif" />

<img file="MX336953B_D0041.tif" />

including horticultural, said method comprises applying to the seeds of the relevant crop or plant; to the soil in which the seed, crop, or plant is planted or is to be planted;

to the watery environment in which plants are growing; or to the matter of the plant itself, a fluid bi-active agrochemical composition in particle form comprising a substantially homogeneous mixture of a) a conventional bi-active agrochemical in particle form and b) a solid carrier particle which has been treated with an acid solution bioactive, especially an aqueous or water-based solution, which has a pH less than 6, preferably 1.5 to 5, and which comprises an acid, especially a weak or moderate acid, at least one antimicrobial metal ion source, and optionally, but preferably, at least one surfactant, especially at least one anionic, nonionic and / or amphoteric surfactant, that affects or interacts with the membranes of the cell wall of microorganisms, especially pathogenic microbes, or the function thereof, said acid present in a molar excess, relative to antimicrobial metal ions, in which the amount of antimicrobial metal ion is 500 ppm or less in the case of a single ion or 1000 ppm or less in the case of multiple antimicrobial metal ions. This modality also

<img file="MX336953B_D0042.tif" />

contemplates that the solid carrier material treated with the bioactive acid solution is the conventional bioactive agrochemical composition by itself, for example, in cases where the bioactive active is present as an active formulated in an appropriate absorbent or adsorbent vehicle or is present as an asset granule and a solid filler or diluent. More preferably, the composition according to this method also comprises at least one agent for adhering or increasing the adhesion of the bioactive agrochemical actives to the matter being treated. Generally speaking, the bioactive agrochemical composition of the invention is applied at a rate or amount at which the conventional bioactive agrochemical is at or near the conventional application amount or less and the applied amount of the antimicrobial metal ion or ions originating from the bioactive acid solution will be no more than approximately 1,235.5 grams per hectare, preferably no more than 617.8 grams per hectare.

In accordance with a ninth embodiment of the present invention, a method is provided to prevent or inhibit the growth of plant pathogens, especially plant-type, stramen-like and fungus-type fungi, bacteria and / or protists in agricultural applications.

<img file="MX336953B_D0043.tif" />

including horticultural, said method comprises applying to the seeds of the relevant crop or plant; to the soil in which the seed, crop, or plant is planted or is to be planted;

to the watery environment in which plants are growing; or to the plant material itself, a fluid bioactive particulate agrochemical composition comprising a substantially homogeneous mixture of a) a conventional bioactive particulate agrochemical and b) a bioactive acid particulate composition, preferably a powder , said bioactive acid composition is prepared by: i) formation of an aqueous solution of an acid, especially a weak or moderate acid, at least one source of antimicrobial metal ion, and optionally, but preferably, at least one surfactant, especially at least one agent anionic, nonionic and / or amphoteric surfactant, which affects or interacts with the cell wall membranes of microorganisms, especially pathogenic microbes, or the function thereof, said acid present in a molar excess, relative to antimicrobial metal ions, in which the amount of antimicrobial metal ion is 500 ppm or less in the case of a single ion or 1000 ppm or less in the case of multiple antimicrobial metal ions, ii) allowing the water evaporates to form the solid bioactive acid and iii) if ι :. .

<img file="MX336953B_D0044.tif" />

;?: íu; o?.; xxi¿; anus. CE LA? Yes: 5L / 2 if necessary, grind or crush the solid bioactive acid formed to form a powder. More preferably, the composition according to this method also comprises at least one agent for adhering or for increasing the adhesion of the bioactive agrochemical assets to the material being treated. As with the previous embodiment, the formulation of the bioactive agrochemical composition of the invention is such that, when applied during use, the application rate or amount of the conventional bioactive agrochemical is at or near the amount of conventional application or less and the applied amount of the antimicrobial metal ion or ions originating from the bioactive acid solution shall be not more than approximately 1,235.5 grams per hectare, preferably not more than 617.8 grams per hectare.

In accordance with a tenth embodiment of the present invention, a method is provided to prevent or inhibit the growth of phytopathogens, especially fungi, bacteria and / or plants, stramenophiols and mushroom-like protists in agricultural applications, including horticultural, said method comprises applying to seeds . the relevant crop or plant; to the soil in which the seed, crop, or plant is planted or is to be planted; to the watery environment in which plants are growing; or to the subject of the

<img file="MX336953B_D0045.tif" />

plant itself, liquid bioactive agrochemical compositions comprising a) an appropriate solvent or solvent system, b) a conventional bioactive agrochemical active dissolved in said solvent or solvent system, c) an acid, especially a weak or moderate acid, d) by at least one antimicrobial metal ion source, and e) optionally, but preferably, at least one surfactant, especially at least one anionic, nonionic and / or amphoteric surfactant, that affects or interacts with the cell wall membranes of microorganisms, especially pathogenic microbes, or the function thereof, in which the solution of the acid and solvent has a pH less than 6, preferably 1.5 to 5, before, and preferably also after, the addition of the conventional bioactive active, said acid present in a molar excess relative to the antimicrobial metal ions from the source (d) and in which the amount of antimicrobial metal ions attributed to the source (d) is 500 ppm or less in the case of an individual ion or 1000 ppm or less for multiple antimicrobial metal ions. Most preferably, the solvent is water or a water-based solvent system, most preferred water, and the conventional bioactive active, acid and antimicrobial metal ion source are all fully or

substantially

V &lt;CÍ, 'tí' ¿ij ¡% “¿yy ^ &lt;5

INSTITUTO KK'CAü :: ';&gt; DE LA PRO' · ':

INDUSTRIAL · soluble or miscible in the solvent. In cases where the conventional bioactive asset is not soluble or miscible in water or a water-based solvent, it can be dissolved in an appropriate nonaqueous solvent, including a lipophilic solvent, and the metal source and acid are dissolved in water or a water-based solvent and the two solutions combine to form a suspension or emulsion. Generally speaking, this formulation of the bioactive agrochemical composition of the invention will be such that, when applied during use, the application rate or amount of the conventional bioactive agrochemical is at or near the conventional application amount or less and the amount applied of the antimicrobial metal ion or ions originating from the bioactive acid solution will be no more than approximately 1,235.5 grams per hectare, preferably not more than 617.8 grams per hectare.

Since a key objective of the present invention is to reduce the amount of metals entering the environment, the preferred embodiments of each of the above embodiments will be such that the contribution of antimicrobial metal ions from the metal ion source antimicrobial will be not more than 300 ppm, more preferred not more than 50 ppm, in the case of a single antimicrobial metal ion and not more than 500 ppm of

IMPI

MEXICAN INSTITUTE OF THE PkC'i-TELMD INDUSTRIAL

<img file="MX336953B_D0046.tif" />

preference not greater than 150 ppm, in the case of multiple antimicrobial metal ions.

Since it is also an objective of the present invention to avoid the use of materials that induce phytotoxicity, the acids are preferably organic acids, more preferred carboxylic acids, and / or the bioactive acid solution, in a diluted state, has a pH greater than 2 and less than 6.

Finally, since it is also an object of the present invention to minimize or reduce the amount of conventional bioactive agrochemicals, especially fungicidal and antiprotistic agents for fungus and plant type protists, the present invention also, and especially, relates to synergistic combinations of antimicrobial-acid metal ion compositions and conventional bioactive agrochemical agents in which the amount of the latter is considerably less than, up to 25 percent less than, preferably up to 50 percent less than, the conventional amount needed to achieve the same result when used alone.

DETAILED DESCRIPTION OF THE INVENTION

The present invention encompasses many embodiments

<img file="MX336953B_D0047.tif" />

different, as indicated above, of which all have a significant degree of common characteristics and constitution. However, although the above describes each modality in its most general aspect, there are many aspects of each that are, in some modalities, common and others that are exclusive to their modalities. In its most fundamental sense, the invention pertains to compositions comprising a) one or more antimicrobial metal ions or ion sources in combination with an acid or in an acidic solution, optionally, which also includes one or more surfactants in combination with b) one or more conventional agrochemical active ingredients to combat or prevent the growth or proliferation of fungi, bacteria, viruses, and protists, plant type, stramenophilic type and fungus type, or formulated products that contain one or more of said assets. The first component (a) may exist as a solution or a mixture of the solid components and is generally referred to in the present invention as the bioactive acid solution and the bioactive acid composition, respectively, or together the solution or composition of bioactive acid. In general, and preferably, the bioactive acid solution or bioactive acid composition, especially the combination of the acid and the antimicrobial metal ion source, is prepared prior to

<img file="MX336953B_D0048.tif" />

addition of the conventional agrochemical active; however, it is also contemplated that the bioactive acid solution or bioactive acid composition, as appropriate, need not be pre-formed. These bioactive acid solutions and compositions, as well as the concentrates thereof, are discussed in greater detail in and are the subject of International Patent Application No. NÓ.PCT / US2008 / 006357 and in the North American patent application 12/154127, 12/154130 and 12/154132 of which both are titled Bioactive Agrichemical Compositions and Use Thereof and were filed the same day as this application with the same inventors than those of the present application, of which both are incorporated in the present invention in their entirety for reference.

For convenience in drafting and simplicity in reading this application, the word bioactive is intended to include agents that annihilate or prevent or inhibit the growth and / or proliferation of bacteria, fungi, viruses, and plant-type, estramenophilic, and fungal-type protists particularly those that attack or adversely affect, even from a purely aesthetic point of view, plants and trees, especially those associated with agricultural and horticultural products including food crops, feed crops, flowers, ornamentals, lawns, and the like, i.e., plant pathogens. Usually,

<img file="MX336953B_D0049.tif" />

The present invention is discussed in terms "" 3e fungicides and antifungal activity as well as fungicidal actives;

although it should be understood that it is not limited to them. Similarly, the word active, especially with reference to conventional bioactive agrochemical materials, refers to those compounds or compositions that are directly responsible for the bioefficacy of the bioactive agrochemical to attack bacteria, fungi, etc. agricultural or horticultural. A formulated active ingredient is one that, in addition to the active ingredient, also contains one or more other constituents that may or may not influence the bioefficacy of the active ingredient, but which are not in themselves directly responsible for the annihilation or prevention of growth. of the target microorganism.

The acids that can be used in the present invention can be solid or liquid in their natural state, but they easily dissolve in or are miscible with water or an aqueous base solvent or with the organic vehicle or diluent used for the particular application that will be faced. For example, if an oil or oil based fungicide is intended to be made, the acid must be soluble in or miscible with the oil of the bioactive composition. More preferably, in cases where a fungicide is used

<img file="MX336953B_D0050.tif" />

conventional oil-based or soluble / oil-miscible, for example, an aqueous environment, the conventional active dissolves in the organic solvent and the acid and the antimicrobial metal ion source in water and the bioactive composition of the present invention exists as an emulsion or suspension.

Exemplary acids include organic acids, especially carboxylic acids such as citric acid, valeric acid, itaconic acid, acetic acid, citriconic acid, lactic acid, malic acid, succinic acid, aldaric acid, malonic acid, propionic acid, malonic acid, maleic acid, salicylic acid, glutaric acid, tartaric acids, benzoic acid and the like, as well as mineral acids such as nitric acid, sulfuric acid, phosphoric acid, boric acid, and the like. Weaker or moderate acids such as aldaric, citric, malic, and lactic acids are preferred as opposed to moderate to strong mineral acids such as boric and phosphoric acids. However, strong acids can be used, especially strong mineral acids such as sulfuric acid or nitric acid; however, depending on the strength of the acid, it would be preferable to buffer the acid to avoid handling and usage problems, especially problems associated with the substrate to which it is being used.

INSTITUTE ? '

LA? AOFi £ PA0) INDUSTRIAL

<img file="MX336953B_D0051.tif" />

you will apply the bioactive composition. This is particularly important for bioactive agrochemical compositions that are to be applied to plants, animals, and crops or foodstuffs because the acid can damage the substrate, directly or indirectly. In plants, for example, there is considerable concern regarding phytotoxicity resulting from acid treatments, alone or in combination with metal compounds such as copper fungicides and the like. Therefore, although effective, it is more preferable to avoid mineral acids and, instead, to use carboxylic acids. Additionally, although some appropriate acids fall outside this range, it would be desirable for the pKa (in water at 25 ° C) of the acid to be greater than 0, preferably greater than 1, more preferred greater than 1.5.

Generally speaking, and despite the strong i

efficacy of phosphoric acid and nitric acid, it is preferred to use weak or moderate acids. This is especially desirable since it avoids the potential need for buffering agents, which are unwanted. Although it will be appreciated that other surfactants, fungicides, wetting agents, emulsifiers, and the like that can be added, and, depending on the end-use application, are likely to be added to the inventive compositions of the present invention, may have an effect. damper over

JLVl XA

MEXICAN INSTITUTE OF INDUSTRIAL PROPERTY

<img file="MX336953B_D0052.tif" />

the pH of such bioactive systems, this effect is not an intended or necessarily desirable effect. Indeed, it may be necessary to add more acid to the composition in order to maintain the required level of acidity.

As noted, acidity is critical to the efficacy of the bioactive agrochemicals of the present invention. Generally speaking, the pH of the bioactive acid solution or bioactive acid composition, prior to the addition of the conventional bioactive agrochemical and other auxiliary compounds, is less than 6, preferably about 1.5 to 5, and more preferably about about 2 to about 4, more preferred greater than 2. Most preferably, the fully formulated bioactive agrochemical compositions of the present invention also satisfy the above pH limitations when or when diluted to the concentration to be applied. In the case of evaluation or confirmation of the pH of the solid bioactive acid composition or of a solid bioactive agrochemical in accordance with the present invention, the bioactive acid composition or, as appropriate, the bioactive agrochemical is first dissolved in water until a concentration equivalent to that to which it will be applied during use, and the pH is measured.

The second critical aspect of concentration

<img file="MX336953B_D0053.tif" />

<img file="MX336953B_D0054.tif" />

"acid" refers to the molar equivalence with respect to the antimicrobial metal ions present in the composition of bioactive acid or bioactive acid solution. At a minimum, there should be a 2-fold molar excess, although preferably there is a molar-excess acid of at least 5-fold, and more preferred at least 10-fold. These levels are typically obtained by formulating bioactive acid solutions in which the acid concentration in the final dilute state of the bioactive composition is from about 0.01% to about 10%, preferably from about 0.1% to about 4% by weight of the solution. Higher concentrations, for example, up to 20% or more, may also be used, provided that the substrate to which the bioactive composition is to be applied is not affected by the higher acid content and / or the acid is a weak or weakly moderate acid. Certainly, higher concentrations can be used in the production of concentrates as discussed below.

The second critical component of bioactive compositions is the antimicrobial metal ion: more accurately its source of metal ion. Appropriate metal ions are selected from the group consisting of antimicrobial transition metal ions and metal ions

Μ mexican institute

FROM THE PKQPJEvAD ^^ saRff ^ lfía INtJUS »R1AL poor who have demonstrated antimicrobial bioefficacy. Preferred metal ions are selected from the group consisting of silver, copper, zinc, mercury, tin, gold, lead, iron, bismuth, cadmium, chromium and thallium ions or combinations of any two or more of the above. Most preferably, metal ions are selected from the group consisting of silver, copper, and zinc ions and combinations of any two or all of the three. Bioactive compositions in which at least two and preferably all three of said preferred ions are present are especially beneficial and preferred. In cases where multiple antimicrobial metal ions are present, each is present in a molar amount of 3 to 97 percent, preferably 9 to 91 percent, more preferred 20 to 80 percent. In this preferred embodiment, in which multiple metal ions are present, they are present in an equal amount in which no metal ion is more than 20 times, more preferably not more than times the amount of any other metal ion. metal. Particularly good results have been found in cases where each antimicrobial metal ion is present in an equal amount, by weight.

The metal ion is added to the acid solution or, as appropriate, to the acid, in the form of a compound, salt

<img file="MX336953B_D0055.tif" />

or complex source that easily releases ions or that otherwise dissociates in the acid solution or when the source and acid are dissolved in a solvent, especially water or a water-based solvent. Examples of organometallic salts and compounds that can suitably serve as the ion sources include the respective oxides, sulfides, carbonates, nitrates, phosphates, dihydrogen phosphates, sulfates, oxalates, quinolinolates, thiosulfates, sulfonates, phthalates, hydroxides, glycollates, and the like of antimicrobial metals as well as carboxylic acid salts thereof, especially simple carboxylates, such as citrates, benzoates, acetates, lactates, etc. of said antimicrobial metals.

Other salts such as halide salts and substituted halide salts, such as halides, hexafluroantimoniates, tetrafluroborates, and perchlorates of said antimicrobial metals can be used although these are less desirable since they tend to have slow and / or poor solubility, especially in water. Specific metal ion sources include, but are not limited to, silver nitrate, silver oxide, silver acetate, silver citrate, cupric oxide, copper hydroxide, cuprous oxide, copper oxychloride, cupric acetate, quinolinolate copper, copper citrate, zinc oxide, zinc citrate, and

<img file="MX336953B_D0056.tif" />

Similar .

It has been surprisingly discovered that some inorganic complexes can also serve as the source of the metal ion. Specifically, ion exchange type antimicrobial agents and solvent glass antimicrobial agents can be used in cases where the carrier matrix for said materials is soluble in the acid or dilute acid. For example, zeolites have been found to be readily soluble in concentrated citric acid. In this case the metal ion source or sources are added to the acid by mixing until the particles dissolve. These metal ion sources are also contemplated to dissolve only partially to provide a longer term source of the antimicrobial metal ion. Although these ion sources tend to dissolve in the dilute acid, to accelerate and / or increase the dissolution of the metal ion source, it is preferable to dissolve them in a concentrated acid solution, preferably one of a concentration of about 40% to 80%.

Appropriate ion exchange type agents include, but are not limited to, aluminosilicates, zeolites, hydroxyapatite, and zirconium phosphates, all of which are commercially available and / or fully described in the patent literature. For example, the yt rj- -wrPa! '\ T:

r-

<img file="MX336953B_D0057.tif" />

χ.Λ

INSTITU '

Gave .

Hydroxyapatite particles containing antimicrobial metal ion are described, for example, in patents.

US Nos. 5,009,898 and 5,268,174; zirconium phosphates containing antimicrobial metal ion are described in, for example, US Patent Nos. 4,025,608; 4,059,679;

5,296,238; 5,441,717 and 5,405,644 as well as in the Journal of Antibacterial and Antifungal Agents,

Vol. 22, No. 10, pp. 595-601, 1994; Alumina ilicates and zeolites containing antimicrobial metal ion are described in, for example, US Patent Nos.

, 911.8 98; 4,911,899; 4,938,955; 4,938,958; 4, 906.4 64; and

4,775,585, all of the aforementioned patents are incorporated into the present invention for reference in their entireties.

Appropriate soluble glasses include those described in, for example, US Patent No. 5,470,585, which is also incorporated in the present invention for reference in its entirety.

Although individual metal ion sources can be used, it is also desirable to use combinations of metal ion sources to provide a mixture of metal ions. In some cases, a single source can supply multiple metal ions. For example, preferred ion exchange type metal ion sources include the product

Agl10's AJ10D which contains both silver and

<img file="MX336953B_D0058.tif" />

<img file="MX336953B_D0059.tif" />

zinc and AglON's AC10D product which includes both silver and copper ions. More preferably, the metal ion sources are the easily soluble salts and compounds, as mentioned above, and most preferred the combination of such compounds with which solutions are prepared having equal or relatively equal concentrations of each of the silver, copper and zinc ions. Appropriate combinations include combinations of silver citrate, copper citrate, and zinc citrate as well as combinations of silver nitrate, copper sulfate, and zinc oxide.

The amount of the antimicrobial metal ion source to be incorporated into the acid solution or, as appropriate, to be combined with the acid is that sufficient to provide a concentration of approximately 1 ppm to approximately 500 ppm, preferably from about 1 ppm to about

300 ppm, more preferably from about 2 ppm to about 100 ppm, more preferably from about 5 to about 50 ppm of each antimicrobial metal ion, in the dilute bioactive acid or bioactive acid composition at its end-use concentration . In cases where metal ions and / or multiple metal ion sources are used to provide

<img file="MX336953B_D0060.tif" />

combinations of metal ions, the total concentration of the metal ions in the solutions should be from about 2 ppm to about 1000 ppm, preferably from about 2 ppm to about

500 ppm, more preferably from about 5 ppm to 300 ppm, more preferably from about 5 ppm to about 150 ppm, in the dilute bioactive acid solution or bioactive acid composition at its end-use concentration. Of course, higher levels can be used but are not necessary to provide proper bioefficacy and, very importantly, such higher use conflicts with the intended attempt to minimize the addition of metal to the environment. Therefore, to continue with this objective, it is preferable to use the minimum amount, or the least possible, for the desired application.

In agricultural and horticultural applications, phytotoxicity is of particular concern. Therefore, in accordance with the agricultural and horticultural applications of this invention, especially for application to seedlings and plants, the level of metals should be lower.

<td>that who</td><td>of</td><td>any</td><td>another way</td><td>could</td><td>cause</td>

<td>phytotoxicity.</td><td>Of</td><td>way</td><td>most preferred,</td><td>how</td><td>It indicated</td>

<td>previously,</td><td>the</td><td>objective</td><td>is to use a</td><td>level</td><td>ion</td>

<img file="MX336953B_D0061.tif" />

PI

MEXICAN INSTITUTE OF PROPERTY

INDUSTRIAL

<img file="MX336953B_D0062.tif" />

metal as low as is reasonably possible to nonetheless continue to provide the desired benefits, especially fungicidal, protistaced, and / or antimicrobial properties. This reason for concern is especially pertinent for those compositions that contain copper alone or in combination with one or more of the other metals and in a very special way, in cases where the conventional fungicide or bioactive active ingredient is a copper material. In this regard, it should be noted that the aforementioned limitations on antimicrobial metal ions refer only to those antimicrobial metal ions contributed by said one or more sources of antimicrobial metal ions associated with the bioactive acid solution or acid composition bioactive, and not to copper or any other antimicrobial metal or metal ions that could be contributed by the conventional bioactive agrochemical used in combination with bioactive acid solutions or bioactive acid compositions. Indeed, due to the synergies found with the present invention, the total content of said antimicrobial metals, especially copper, is probably less than what would be found with only the conventional bioactive agrochemical to achieve the same result.

Optionally, although preferably the

I '

INSTITUI /

Say LA I li ÜU

<img file="MX336953B_D0063.tif" />

bioactive acid solutions or bioactive acid compositions, and, in any event, the bioactive agrochemical compositions of the present invention include one or more surfactants, especially water soluble surfactants. Although good results have been achieved in weak and moderate acid bioactive acid solutions without the surfactants, the use of the surfactant should be and is generally preferred with such acids.

Also, although certain strong and very strong acids, especially mineral acids, do not justify the need for surfactants, for example, phosphoric acid, it is especially desirable, and in some cases necessary, for example, in cases where different bioefficacy is desired. of the short-term only, employing one or more surfactants. Especially preferred surfactants are those that affect or interact with the cell walls or membranes of microorganisms, especially pathogenic microbes, or their function. Appropriate surfactants include anionic, cationic, nonionic, and amphoteric surfactants (eg, zwi-terionics), especially those that are water soluble or show relatively good water solubility.

Preferably the surfactants are anionic, nonionic and / or amphoteric surfactants such as

K / ίΓ Ή) and AVI r A

EX INSTITUTE (PROPERTY CANO

INDUSTRIAL

<img file="MX336953B_D0064.tif" />

sulfonates, sulfates, sulfosuccinates, sarcosinates, mor.oglycerides and diglycerides, amine oxides, ether carboxylates, betaines, sulfobetaines, glycinates and the like. Generally speaking, cationic and nonionic surfactants having polyalkyl ether units, especially polyethylene oxide units, with degrees of polymerization of the alkylene ether unit of more than about 6 do not show the same level of effectiveness in providing synergy to bioactive compositions than other surfactants.

However, such surfactants can be used in combination with effective surfactants as long as they do not materially detract from or reduce the bio-efficacy of the compositions.

The surfactant is typically used in conventional amounts, that is, it is added to bioactive acid solutions or bioactive acid compositions in an amount with which the concentration of the surfactant in the dilute end-use state of the bioactive agrochemical compositions is consistent with its level of use in traditional fungicides. Generally speaking, the surfactant is present in an amount of from about 0.001% to about 3%, preferably from about 0.01% to about

<img file="MX336953B_D0065.tif" />

0.5%, by weight based on the total weight of the bioactive acid solution or bioactive acid composition in the diluted state. Although higher loads can be used, it is not necessary to manifest the desired synergy in bioefficacy. Generally speaking, in cases where the surfactant is basic in nature or one that is hydroized in water to form a basic solution, the amount should be minimized and / or the amount of acid increased to avoid too much neutralization of the bioactive acid solution.

Exemplary anionic surfactants and classes of anionic surfactants suitable for use in the practice of the present invention include:

alcohol sulfates; alcohol ether sulfates; alkylaryl ether sulfates; alkylaryl sulfonates such as alkylbenzenesulfonates and alkylnaphthalene sulfonates and salts thereof; alkylsulfonates; monophosphate or diphosphate esters of polyalkoxylated alkyl alcohols or alkylphenols; monosulfosuccinate or di-sulfosuccinate esters of C12-alkanols

C15 or C alkanols_i2 to polyalkoxylated C15; alcohol ether carboxylates; phenolic ether carboxylates; Polybasic acid esters of ethoxylated polyoxyalkylene glycols consisting of oxybutylene or the tetrahydrofuran residue;

sulfoalkylamides and salts thereof such as the condensation carboxylate salt of N-methyl-N-oleoyltaurate naphthalene v sodium alkyl ester sulfates;

polyoxyalkylene-alkylphenol; polyoxyalkylene alcohol carboxylate polyglucoside / alkenyl succinic anhydride products;

I rent; naphthalenesulfonates; formaldehyde condensates; alkylsulfonamides; sulfonated aliphatic polyesters; styrylphenyl alkoxylate ester sulfates; and styrylphenyl alkoxylate ester sulfonates and their corresponding sodium, potassium, calcium, magnesium, zinc, ammonium, alkylammonium, diethanolammonium, or triethanolammonium salts;

ligninsulfonic acid salts such as the sodium, potassium, magnesium, calcium or ammonium salt; polyarylphenol polyalkoxyether sulfates and polyallphenol polyalkoxyether phosphates; and sulfated alkylphenol ethoxylates and phosphated alkylphenol ethoxylates; sodium lauryl sulfate; sodium laurethsulfate; ammonium lauryl sulfate; ammonium laureth sulfate; sodium methylcocoyltaurate; lauroilsareos sodium inate; sodium cocoylsarcosinate; hydrolyzed coco-collagen potassium; TEA lauryl sulfate (triethanolamine); TEA laurethsulfate (triethanolamine); lauryl- or cocoyl-sarcosine;

disodium oleamide sulfosuccinate; disodium laureth-sulfosuccinate; disodium dioctylsulfosuccinate; Nmethyl-N-oleoyltaurate sodium salt; triestyrylphenol sulfate; ethoxylated ligninsulfonate; ethoxylated nonylphenol phosphate ester;

<img file="MX336953B_D0066.tif" />

<img file="MX336953B_D0067.tif" />

calcium alkylbenzenesulfonate; ethoxylated tridecyl alcohol phosphate ester; dialkylsulfosuccinates; perf luoroalkyl acids (C₆-Ci₈) phosphonic; perfluoroalkyl acids (C₆C₁₈] -phosphinic; perfluoroalkyl esters of C3-C20 carboxylic acids; alkenyl succinic acid diglucamides;

alkenylsuccinic acid alkoxylates; sodium dialkylsulfosuccinates; and alkenyl succinic acid alkyl polyglucosides.

Examples of amphoteric and cationic surfactants include alkylpolyglucosides; betaines;

sulfobetaines; glycinates; fatty acid alkanolamides of

Cg to C₁₈ and C fatty acid amine polyalkoxylates₈ to Ci₈;

alkyl (Cio-CRg) dimethylbenzylammonium chlorides; coconut alkyldimethylaminoacetic acids; C-fatty acid amine polyalkoxylate phosphate esters₈ a C₁₈;

alkylpolyglucosides (APG) that can be obtained from an acid catalyzed Fischer reaction of starch syrups or glucose with fatty acid alcohols, in particular C alcohols₈ to Ci₈, especially C-alkyl polyglucosides₈ to Cío YC₁₂ to C14 that have a degree of polymerization from 1.3 to 1.6, in particular 1.4 or 1.5.

Examples of nonionic surfactants and classes of nonionic surfactants include: polyarylphenol polyethoxyethers; polyethoxyethers from

<img file="MX336953B_D0068.tif" />

INSTITUTO MEXICANO de la pro? E: v :: INDUSTRIAL polyalquilfenol; polyglycol ether derivatives of saturated fatty acids; polyglycol ether derivatives of unsaturated fatty acids; polyglycol ether derivatives of aliphatic alcohols; polyglycol ether derivatives of cycloaliphatic alcohols; polyoxyethylene sorbitan fatty acid esters;

alkoxylated vegetable oils; alkoxylated acetylenic diols; polyalkoxylated alkylphenols; fatty acid alkoxylates; sorbitan alkoxylates; sorbitol esters;

C- alkyl- or alkenylpolyglucosides₈ to C22; polyalkoxy-styryl aryl ethers; alkylamine oxides; block copolymer ethers; polyalkoxylated fatty acid glyceride; polyalkylene glycol ethers; linear aliphatic or aromatic polyesters; organosilicones; polyarylphenols; sorbitol ester alkoxylates; and monoesters and diesters of ethylene glycol and mixtures thereof;

ethoxylated tristyrylphenol; ethoxylated fatty acid alcohol;

ethoxylated lauryl alcohol; ethoxylated castor oil; and ethoxylated nonylphenol; alkoxylated alcohols, amines or acids, mixtures thereof as well as mixtures thereof with diluents and solid carriers, in particular clathrates thereof with urea. The preferred alcohols, amines or alkoxylated acids are based on alkoxy units having 2 carbon atoms, being therefore a mixed ethoxylate, or 2 and 3 carbon atoms, being therefore a

<img file="MX336953B_D0069.tif" />

mixed ethoxylate / propoxylate, and having at least 5 alkoxy portions, suitably 5 to 25 alkoxy portions, preferably 5 to 20, particularly 5 to 15, in the alkoxy chain. The aliphatic portions of the amine or alkoxylated acid can be straight or branched chains of 9 to 24, preferably 12 to 20, carbon atoms. The alcohol portion of the alcohol alkoxylates as a rule is obtained from a Cg-Cis aliphatic alcohol, which can be unbranched or branched, especially mono-branched. Preferred alcohols are typically 50% by weight straight and 50% by weight branched alcohols.

As indicated above, the aforementioned surfactants can be used alone or in combination. Also, although not all of the surfactants mentioned above can provide the desired synergy when used alone with the metals and acids, depending on the ultimate end-use application for the bioactive agrochemical compositions of the present invention, they can nonetheless be used in combination with synergistic surfactants for their intended function. For example, some of the aforementioned surfactants may increase the dispersion of the assets in the solvent, especially the conventional bioactive agrochemical, or may increase the wetting

<img file="MX336953B_D0070.tif" />

of the substrate (eg, plants, seeds, soil) to which the inventive bioactive agrochemical compositions of the present invention are applied. All of these surfactants are well known and commercially available. Also, those skilled in the art, without undue experimentation, will readily appreciate which surfactants and / or combinations of surfactants, in addition to synergistic surfactants, can be used for the specific end-use application. Again, it is important that when additional surfactants are used for other purposes they do not interfere with or have minimal interference with the synergy that results from the desired surfactants, i.e. those that show synergy to provide antimicrobial activity, including antibacterial activity and / or antifungal, when used in combination with acid and metal ions. If there is any interference and the other surfactant is necessary or otherwise desired for the application, then its use should be minimized to produce the least adverse impact on synergy and / or attributes of the active components of the bioactive agrochemicals inventive of the present invention while manifesting the desired property for which it is to be used. Also, if there is a reason for aBM = 3XST! U lfl

IN5TI7UK

Four. P¹

and.

3.

<img file="MX336953B_D0071.tif" />

Concern with such interference, especially if the surfactants are used or will be used in an amount that can neutralize the acid of the bicactive compositions to leave them outside the claimed range, then these surfactants may still be added but not before now. of the application. In essence the bioactive compositions of the present inventions can be used as two or more part systems that are mixed when applied or when the diluted compositions are prepared, which are then applied immediately. More preferably, it is better to avoid the use of said surfactants or said amounts of the surfactants that may adversely affect the bioefreacia of the claimed compositions.

The final critical component of the bioactive compositions of the present invention is the conventional bioactive agrochemical component. Surprisingly, it has been found that these combinations typically provide a synergy relative to the bioefficacy of the conventional bioactive asset, allowing less to be used for the same, if not better, results. In particular, when used in combination with the bioactive acid solution or bioactive acid composition as indicated in the appended claims, the levels

<img file="MX336953B_D0072.tif" />

Previously ineffective of conventional bioactive actives, especially fungicidal actives, now turn out to be effective. Very importantly, these combinations frequently allow the same level of bioefficacy to be achieved with lower than conventional application rates or amounts of conventional bioactive agrochemical active. For example, it has been found that the addition of the bioactive acid solution or, as appropriate, bioactive acid composition, to a given amount of a conventional antifungal agent exhibits increased antifungal performance compared to the same amount of said antifungal agent. . Indeed, previously ineffective amounts of conventional agrochemical compositions can now be made effective by adding the bioactive acid solution or bioactive acid compositions. In addition to reducing the amount of conventional agrochemical assets that is necessary to achieve a given effect, and concurrently, reducing the amount of such assets that are released into the environment, it is believed that the combination also reduces the incidence of and / or the speed with which bio-resistance is manifested in the target organisms. Therefore, the commercial life expectancy of these and future assets is likely

<img file="MX336953B_D0073.tif" />

<img file="MX336953B_D0074.tif" />

Conventional agrochemicals are increased and the generation of super pests or resistant strains of bacteria, fungi, protists and the like is reduced or delayed.

As indicated above and indicated in the following examples, the present invention can also be applied to a wide variety of conventional bioactive agrochemicals, most especially fungicides. Examples of conventional bioactive agrochemicals, many of which have multiple applications as fungicides, bactericides, protistacids and the like, are described below in the present invention, some of the active ingredients are mentioned several times with different common names and / or chemical names include: AC 382042, acetochlor, alachlor, aldicarb, anilazine, asulam, atrazine, azoxystrobin, benalaxil, bendiocarb, benfuracarb, benomyl, bentiocarb, binapacryl, blasticidine S, borax, mixture of

Bordeaux, Bromacil, Bromuconazole, Bupirimate, Butachlor, Butam, Cadusafos, Calcium Cyanamide, Carpropamid, Captafol, Captan, Carbaryl, Carbendazim, Carbofuran, Carbon Disulfide, Carbohydrogen, Carboxin, CDAA, CDEA,

CDEC, CEPC, chlor-IPC, chloramben, corbenztiazon, chlorbromuron, chlordane, chlorfluazuron, chloridazon, chloropicrin, chlorothalonil, chlorotoluron, chloroxyphenidim (= chloroxuron), chlorprofam, clozolinate, copper acetate, acetoarsenate

<img file="MX336953B_D0075.tif" />

copper, copper arsenate, copper carbonate, basic, copper hydroxide, copper naphthenate, copper oleate, copper oxychloride, copper 8-quinolinolate, copper silicate, copper sulfate, basic copper sulfate, copper chromate and zinc, cyanozine, cycloheximide, cymoxanil, cipofuram, cyprodinil, dalapon, dazomet, decarbofuran, di-alato (= dialate), diazinon, dibrom, 1,2-dibromoethane, diclobenyl,

1,2-dichloroethane, dichloromethane, 1,2-dichloropropane, 1,3-dichloroporpene, dieldrin, diphenamid, dipterex, diuron, diclofluanid, diclone, dicloran, diclobutrazol, diclocimet, dimethomezine, difluconazole, difluconazole, diflumethorim, ditalimfos, ditianon, dodemorf, dodina, endosulfán, endrin, epoxiconazol, EPTC, etalfluralin, etilan (= ethyl-DDD), ethylene dibromide, ethylene dichloride, ethylene oxide, ethylmercury bromide, ethylmercury chloride, ethylmercury phosphate, edifenfos, ethaconazole, etirimol, etridiazole, famoxadone, fenapanil, fenamidone, fenarimol, fenbuconazole, fenfuram, fenhexamid, fenpiclonil, fenpropidin, fenpropimorf, fentinin, fentinin hydrochloride, fentin, hydrochloride fluquinconazole, flusilazol, flusulfamida, flutolanil, flutriafol, folpet, fuberidazol, furalaxil, furametpir, fenuron, ferbam, ferrous sulfate flucloralin folpel (= folpet) formaldehyde

I Μ. .Ρ I

INSTITUTO MEXICANO Kjpáflfrtí'J D2 PROPERTY V ¡MUUSTiílAL fostiazato, furmeciclox, gamma-BHC, gamma-cyhalothrin, gammaHCH, guanoctin (= guazatine), HCH, gamma-HCH, heptachlor, hexachlor, cyanide, hydrogen cyanide , imazapir, imazaquin, imazetapir, imazosulfuron, imidacloprid,

IPC, iprodiona, isonoruron, isoprocarb, isoproturon, isouron,

1KF-916, imazalil, iminoctadine, ipconazole, isoprothiolane, iprovalicarb, kasugamycin, KH-7281, kitazin P, kresoximmethyl, lactofen, lenacil, lime, linuron, mancobre, mancozeb, maneb, mercuric chloride , metalaxil, metalaxil-M, metam, metazaclor, metabenztiazuron, metam, methyl bromide, methylchloroform, isothiocyanate, methyl mercapto, methylmercaptofos, methylmercury benzoate, methylmercury, metiram, metobenzuron, metobromuron metolachlor, Smetolaclor, metosulam, metoxuron, metribuzin, mirex, molinate, monalide, monolinuron, monuron, MSMA, mepanipyrim, mepronil, metalaxil, metfuroxam, MON 65500, miclobutanil, nabamide, naphatic, naphthalic anhydride, nitralin, norflurazon, noruron, novaluron, necasozin, nickel dimethyldithiocarbamate, nitrotalisopropyl, nuarimol, ofurace, orizalin, oxadixil, oxina-copper, oxina-Cu, oxicarboxin, oxicarboxin, penconazole, pencicuron, fenazine oxide polyoxin D, poliram, probenazole prochloraz procymidione propamocarb

<img file="MX336953B_D0076.tif" />

propiconazole, propineb, pyrazophos, pyrifenox, pyrimethanil, pyroquilon, pyroxifur, PCNB, PCP, pebulate, pendimethalin, phenylmercuriurea, phenylmercury acetate, phenylmercuride nitrate, phosphorylphosphate, furate, picloram, pyrimicarb, potassium arsenate, potassium cyanate, potassium polysulfide, potassium thiocyanate, prochloraz, procymidone, profluralin, profam, propiconazole, propineb, prosulfalin, pyrazon, quinclorac, quinomethionate, quinoxifene, cincozen, spiroxamine, SSF-126,

SSF-129, streptomycin, strobilurins, sevin, siduron, simazine, sulfur, sulfuric acid, tebuconazole, terbacil, terraclor, thiabendazole, thiacloprid, thiameturon, thiobencarb, thiodan, thiodicarb, thiophanocarbyl, thioofanocarbyl, thiophanocarbyl, thiophanocarbonate, thiophanocarbyl, thiophanocarbyl, thiophanocarbonate, thiophanocarbonate, thiophanocarbonate, thioanopropyl, thioanoprotein, thioanopropyl thioanide, thioanopropyl thioanide, thioanopropyl thioanide. , tiram, thiabendazole, TMTD, toxaphene, tri-alate, thiuram, triadimefon, triadimenol, trialate, tribenuron, hydroxide, triflumuron, trifluralin tifluzamide triazoxide, triflumizol tecloftalam, technazene, tetraconazole, tolconazole tolylfluanid, triazbutyl, tricyclazole, tridemorf, trifloxystrobin, triforine, validamicin A, vinclozolin, vernolate, vinclozolin, zineb, 1,2-dichloropropane, 1,3-dichloropropene, XRD-563 and zarilamid. Further:

dithiocarbamates and their derivatives such as

<img file="MX336953B_D0077.tif" />

iron (III) dimethyldithiocarbamate, zinc dimethyldithiocarbamate, zinc ethylenebisdithiocarbamate, manganese and zinc ethylenebisdithiocarbamate, manganese and zinc ethylenediaminbis-dithiocarbamate, (N, N-ethylene dicarbonate, ammoniacal complex) Zinc N'-propylenebisdithiocarbamate), zinc (N, N '~ propylenebisdithiocarbamate), N, N' ~ polypropylenebis (thiocarbamoyl) disulfide;

• nitro derivatives, such as dinitro (1-methylheptyl) phenyl crotonate, 2-secbutyl-4,6-dinitrophenyl 3,3-dimethylacrylate, 2-sec-butyl-4,6-dinitrophenyl isopropyl carbonate, 5-nitro-isophthalate diisopropyl;

• heterocyclic substances, such as acetate

2-heptadecyl-2-imidazoline, 2,4-dichloro-6- (o-chloroanilino) -striazine, Ο, Ο-diethyl, phthalimidophosphonothioate, 5-amino-l [bis (dimethylamino) phosphinyl] -3-phenyl-l , 2,4-triazol, 2,3diciano-1,4-dithioanthraquinone, 2-thio-l, 3-dithiolo [4,5b] quinoxaline, methyl 1- (butylcarbamoyl) -2-benzimidazolcarbamate, 2-methoxycarbonylaminobenzimidazole, 2 - (2furyl) benzimidazole, 2- (4-thiazolyl) benzimidazole, N- (l, 1,2,2-tetrachloroethylthio) tetrahydroftalimide, Ntricloromethylthiotetrahydroftalimide, N-trichloromethylthiophthalimide; N-dichlorofluoromethylthio-N ', N'-dimethyl-Nphenylsulfodiamide, 5-ethoxy-3-trichloromethyl-1,2,3-thiadiazole,

Ύ -

<img file="MX336953B_D0078.tif" />

2-thiocyanatomethylthiobenzothiazole, l, 4-dichloro-2,5-dimethoxybenzene, 4- (2-chlorophenylhydrazone) -3-methyl-5isoxazolone, pyridin-2-thiol 1-oxide, 8-hydroxyquinoline or its copper salt, 2,3- dihydro-5-carboxanilido-6-methyl-l, 4-oxatiine, 2,3-dihydro-5-carboxanilido-6-methyl-l, 4-oxatiine

4,4-dioxide, 2-methyl-5,6-dihydro-4H-pyran-3-carboxanilide, 2-methylfuran-3-carboxanilide, 2,5-dimethylfuran-3-carboxanilide,

2.4.5- trimethylfuran-3-carboxanilide, N-cyclohexyl-N-methoxy 2.5- dimethylfuran-3-carboxamide, 2-methylbenzanilide, 2-iodobenzanilide, N-formyl-N-morpholin-2,2,2-trichloroethylacetal, piperazin- 1,4-di-ylbis-1- (2,2,2 trieloroeti1) formamide, l- (3,4-dichloroanilino) -1-formylamino2,2,2-trichloroethane, 2,6-dimethyl-N-tridecylmorpholine or its salts, 2,6-dimethyl-N-cyclododecylmorpholine or its salts, N— [3 - (p-tert-butylphenyl) -2-methylpropyl] -cis-2,6-dimethylmorpholine,

N- [3- (p-tert-Butylphenyl) -2-methylpropyl] piperidine, 1- [2 - (2,4-dichlorophenyl) -4-ethyl-l, 3-dioxolan-2-ylethyl] -1H-1,2,4 -triazole,

1- [2- (2,4-Dichlorophenyl) -4-n-propyl-l, 3-dioxolan-2-ylethyl] -lH1,2,4-triazole, N- (n-propyl) -N- (2 , 4,6-trichlorophenoxyethyl) -Ν 'imidazolylurea, 1- (4-chlorophenoxy) -3,3-dimethyl-l- (1H-1,2,4 triazol-l-yl) -2-butanone, l- ( 4-chlorophenoxy) -3,3-dimethyl-l (1H-1,2,4-triazol-l-yl) -2-butanol, (2RS, 3RS) -1- [3- (2-chlorophenyl) -2- ( 4-fluorophenyl) oxiran-2-ylmethyl] -1H-1,2,4-ta (2-chlorophenyl) -a- (4-chlorophenyl) -5-pyrimidinemethanol, 5-butyl20

2-Cimethylamino-4-hydroxy-6-methylpyrimidine chlorophenyl) -3-pyridinemethanol,

<img file="MX336953B_D0079.tif" />

bis (p1,2-bis (3-ethoxycarbonyl-2thioureido) benzene

1,2-bis (3-me toxicarbonyl-2thioureido) benzene;

• strobilurins such as methyl E-methoxyimino- [oi- (otolyloxy) -o-tolyl] acetate, E-2- {2- [6- (2-cyanophenoxy) pyrimidin-4-yloxy] phenyl} -3-methoxyacrylate methyl,

N-methyl-E-methoxyimino- [oi- (2-phenoxyphenyl)] acetamide, N-methylE-methoxyimino- [a- (2,5-dimethylphenoxy) -o-tolyl] acetamide, E-2 {2 - [( Methyl 2-trifluoromethylpyridyl-6-) oxymethyl] phenyl} -3-methoxyacrylate, (Ε, E) -methoximino- {2- [1- (3trifluoromethylphenyl) ethylidenamino-oxymethyl] phenyl} methyl acetate, N— (2 - {[ methyl- (4-chlorophenyl) -lH-pyrazolyl-oxymethyl} phenyl) -N-methoxycarbamate;

• anilinopyrimidines such as N- (4,6dimethylpyrimidin-2-yl) aniline, N- [4-methy1-6- (1-propinyl) pyrimidin-2-yl] aniline, N- [4-methy1-6-cyclopropylpyrimidin- 2yl] aniline;

• phenylpyrroles such as 4- [2,2-difluoro-l, 3benzodioxol-4yl) pyrro1-3-carbonitrile;

• cinnamamides such as 3- (4-chlorophenyl) -3- (3,4-dimethoxyphenyl) acryloylmorpholine;

• and a variety of fungicides such as dodecylguanidine acetate, 3- [3- (3,5-dimethyl-2-oxycyclohexyl) -262

<img file="MX336953B_D0080.tif" />

hydroxyethyl] glutarimide, hexachlorobenzene, methyl N- (2,6-dimethylphenyl) N- (2-furoyl) -DL-alaninate, DL-N- (2,6-dimethylphenyl) -N- (2'metoxiace ti 1 methyl ester ) alanine, N- (2,6-dimethylphenyl) -N-chloroacetyl-D, L2-aminobutyroiactone, DL-N- (2,6dime tilphenyl) -N- (phenylacetyl) alanine, 5-methyl-5- methyl ester vinyl-3- (3,5-dichlorophenyl) -2,4-dioxo-1,3-oxazolidine, 3- [3,5-dichlorophenyl (5-methyl-5-methoxymethyl] -1,3-oxazolidin-2,4- dione, 3- (3,5-dichlorophenyl) -1-isopropylcarbamoylhydantoin, N- (3,5-dichlorophenyl) -1,2-dimethylcyclopropan-1,2-dicarboximide, 2 cyano- [N- (ethylaminocarbonyl) 2-methoximino] acetamide, l— [2— (2,4-dichlorophenyl) pentyl] - 1H-1,2,4-triazole, 2,4difluoro-α- (1H-1,2,4-triazolyl-1-meth 1) benzhydryl, N- (3-chloro-2,6-dinitro-4-trifluoromethylphenyl) -5-trifluoromethyl-3-chloro-2-aminopyridine, 1 - ((bis (4-fluorophenyl) methylsilyl) methyl) -lH-l, 2,4-triazole.

Especially preferred fungicides include, in particular, those based on strobilurin and its derivatives as well as miclobutanil and mancozeb. When used in combination with the bioactive compositions of the present invention, a synergistic result is frequently observed in which efficacy is found where none previously existed and / or improved efficacy is found at the same application rates and, preferably,

<img file="MX336953B_D0081.tif" />

at lower application rates.

The bioactive agrochemical compositions according to the present invention can be used alone or, preferably and conveniently, they are used in combination with (typically as a mixture) one or more other compatible components or additives typical of agrochemical treatments and compositions including, for example, solid or liquid fillers or diluents, adjuvants, surfactants or equivalents, which are appropriate for the intended use and which are acceptable for use, from an environmental, health and safety as well as regulatory perspective, in the particular intended end use application. This is especially so for those applications in which bioactive compositions are to be used in agriculture: either as a soil, seed, or plant treatment or in edible treatment, before or after harvest.

Accordingly, the formulations may also contain other types of ingredients, such as protective colloids, adjuvants, binders, rain fixers, thickeners, thixotropic agents, penetrating agents, spray oils, stabilizers, antifreeze agents, defoaming agents, foaming agents, inhibitors. corrosion, dyes, or the like, as well as other active ingredients

<img file="MX336953B_D0082.tif" />

known to have pesticidal properties (in particular fungicidal, insecticidal, acaricidal or nematocidal properties) or, in the case of agricultural applications in the field, to have plant growth regulating properties.

The nature and quantity of the additives to be used in the bioactive agrochemical compositions of the present invention depend, in part, on the end use application and the way in which the composition is to be applied. Specifically, the bioactive compositions of the present invention may be in the form of and / or be manufactured as, for example, emulsion concentrates, solutions, oil-in-water emulsions, wettable powders, soluble powders, suspension concentrates, powders, granules, granules dispersible in water, microcapsules, gels, tablets and other types of formulation using well established procedures. The specific procedure typically includes thorough mixing and / or milling of the bioactive compositions with the other substances. The form of application such as spraying, spraying, dispersing, dusting, pouring and the like can be chosen based on the compositions to be applied, the desired objectives, and the given circumstances.

Although the typical definition of

ΙΜΡϊρ

D £ LA H'Op; ¿; i_AD P &lt;· INDUSTRIAL filler is an aggregate material with the primary purpose of adding volume. In the present application, filler materials typically have function and utility and generally refers to natural or synthetic, organic or inorganic components with which they are they combine the active components to facilitate their application, for example, on plants, seeds or the soil, for example as a vehicle. These fillers are generally inert and must be acceptable for the intended application, especially for agronomic uses, in particular for the treatment of plants.

The filler material can be solid, for example clays, natural or synthetic silicates, silica, resins, waxes, solid fertilizers (for example ammonium salts), natural soil minerals, such as kaolin, clay, talc, lime, carbonate of calcium, quartz, attapulguite, montmorillonite, bentonite or diatomaceous earth, or synthetic minerals, such as silica, alumina or silicates, in particular aluminum or magnesium silicates. Solid fillers that are appropriate for granules are as follows: natural crushed or crushed rocks such as calcites, marble, pumice, sepiolite or dolomite; synthetic granules of inorganic flours or

organic; granules of material

I to I

FROM LA rKCPISDAO WU A · 'inclstiíial ^ .ír.' · '' *

<img file="MX336953B_D0083.tif" />

organic such as sawdust, coconut husk, corn husks or leaves, or tobacco stalk; diatomaceous earth, tricalcium phosphate, powdered cork or adsorbent carbon black;

water soluble polymers, resins, waxes; or solid fertilizers. Said compositions may contain, if desired, one or more compatible agents such as wetting agents, dispersing agents, emulsifiers or colorants which, when these are solid, can also act as diluents. In cases where the additives are alkaline and are likely to increase the pH of the compositions, for example talc, lime, calcium carbonate, and marble, the amount in which they are added should not cause the pH to exceed the claimed ranges or additional acid must be added to maintain the desired pH.

Preferably, such materials should be completely avoided.

The fillers can also be liquids, for example: water, alcohols, in particular butanol or glycol, as well as ethers or esters thereof, in particular methyl glycol acetate; ketones, in particular acetone, cyclohexanone, methyl ethyl ketone, methyl isobutyl ketone or isophorone; petroleum fractions such as paraffinic or aromatic hydrocarbons, in particular xylenes or alkynaphthalenes; mineral or vegetable oils;

aliphatic chlorohydrocarbons, in particular trichloroethane or

<img file="MX336953B_D0084.tif" />

methylene chloride; aromatic chlorohydrocarbons, in particular chlorobenzenes; water-soluble or highly polar solvents such as dimethylformamide, dimethyl sulfoxide, N, N-dimethylacetamide or N-methylpyrrolidone; Noctylpyrrolidone, liquefied gases; or the like, whether these are applied separately or as a mixture.

As mentioned above, depending on the end use application, the bioactive agrochemical compositions or formulations of the invention may contain one or more additional surfactants (additional to the surfactant agent or agents that are optionally part of the bioactive acid solution or composition of bioactive acid) such as emulsifiers, dispersing agents, wetting agents and the like. These additional surfactants can be cationic, anionic, nonionic or amphoteric surfactants or mixtures of these surfactants. Among the surfactants that are used, for example, are polyacrylic acid salts, lignosulfonic acid salts, phenolsulfonic or naphthalenesulfonic acid salts, polycondensates of ethylene oxide with fatty acid alcohols or fatty acids or esters of fatty acid or fatty acid amines, substituted phenols (in particular alkylphenols or arylphenols), sulfosuccinic acid ester salts, taurine derivatives

INSTI? ' C-2 '

<img file="MX336953B_D0085.tif" />

(in particular alkyl taurates), phosphoric esters of alcohols or of polycondensates of ethylene oxide with phenols, fatty acid esters with polyols, or sulfate, sulfonate or phosphate functional derivatives of the above compounds as well as those surfactants described above in relation to to the synergistic surfactant for the bioactive composition. In this case, however, surfactants are generally present at much higher concentrations than necessary to show synergy with respect to the acid / metal combination. The presence of at least one additional surfactant is generally essential when the active materials and / or the inert filler material are insoluble or only very poorly soluble in water and when the filler material for said composition to be applied is water. For foliar applications, the choice of surfactants is often crucial to obtain good bioavailability of the material or active materials; therefore, preferably a combination of a hydrophilic surfactant (HLB&gt; 10) and a lipophilic surfactant (HLB &lt;5) is used.

In agricultural applications as well as applications where it is desired to fix the bioactive composition to a surface, the compositions typically have a

<img file="MX336953B_D0086.tif" />

binder, rain bra, or other adhesive-type components. Appropriate binders are well known and include, for example, water-soluble and water-dispersible film-forming polymers. Appropriate polymers have an average molecular weight of at least about 1,000 to about 100,000; more specifically at least about 5,000, up to about 100,000. Aqueous compositions generally contain from about 0.5% to about 10%, preferably from about

1.0% to about 5%, by weight of the composition of the binder, film-forming polymer, and the like. Suitable film-forming polymers include, but are not limited to: a) random and alkylene oxide block copolymers such as ethylene oxide-propylene oxide block copolymers (EO / PO block copolymers) including block copolymers both EO-PO-EO and

PO-EO-PO; random and block copolymers of ethylene oxide-butylene oxide, C2-C6 alkyl adducts of random and ethylene oxide-propylene oxide block copolymers, alkyl adducts of C₂-C₆ of block and random copolymers of ethylene oxide-butylene oxide; b) polyoxyethylene-polyoxypropylene monoalkyl ethers such as methyl ether, ethyl ether, propyl ether, ether

- V -¿λ,

MEXICAN INSTITUTE OF THE f'RüPírjXL)

INDUSTRIAL ^TO

<img file="MX336953B_D0087.tif" />

butyl or mixtures thereof; c) acetate copolymers

I of vinyl / -vinylpyrrolidone, d) alkylated vinylpyrrolidone copolymers, e) polyvinylpyrrolidone, and f) polyalkylene glycol including polypropylene glycols and polyethylene glycols. Specific examples of suitable polymers include Pluronic P103 (BASF) (EO-PO-EO block copolymer), Pluronic P65 (BASF) (block copolymer

EO-PO-EO), Pluronic P108 (BASF) (EO-POEO block copolymer), Vinamul 18160 (National Starch) (polyvinyl acetate),

Agrimer 30 (ISP) (polyvinylpyrrolidone), Agrimer VA7w (ISP) (vinyl acetate / vinylpyrrolidone copolymer), Agrimer

AL 10 (ISP) (alkylated vinylpyrrolidone copolymer), PEG

400 (Uniqema) (polyethylene glycol), Pluronic R25R2 (BASF) (PO-EO-PO block copolymer), Pluronic R31R1 (BASF) (PO-EO-PO block copolymer) and Witconol NS 500LQ (Witco) (butanol PO copolymer -EO).

Additional adhesive and adhesive type materials that can be used include carboxymethyl cellulose, or natural or synthetic polymers in the form of powders, granules or matrices, such as acacia, latex, polyvinylpyrrolidone, polyvinyl alcohol or polyvinyl acetate, can be used in the formulations, natural phospholipids, such as cephalins or lecithins, or synthetic phospholipids.

<img file="MX336953B_D0088.tif" />

It may also be desirable to thicken bioactive compositions and formulations, especially in cases where there is cause for concern that the composition will quickly exude or run off the substrate to which it is applied. Suitable thickeners include water-soluble polymers that exhibit pseudoplastic and / or thixotropic properties in an aqueous medium such as gum arabic, karaya gum, tragacanth gum, guar gum, locust bean gum, xanthan gum, carrageenan, alginate salt, casein, dextran. , pectin, agar, 2-hydroxyethyl starch, 2-aminoethyl starch, 2-hydroxyethyl cellulose, methyl cellulose, carboxymethyl cellulose salt, cellulose sulfate salt, polyacrylamide, alkali metal salts of maleic anhydride copolymers, alkali metal salts of poly (meth) acrylate, and the like. As suitable thickeners, including thixotropes, there may also be mentioned attapulguite-type clay, silica, fumed silica, carrageenan, croscarmellose sodium, furceleran, glycerol, hydroxypropylmethylcellulose, polystyrene, block copolymers of vinylpyrrolidone / styrene, hydroxypropylcellulose, hydroxypropylcellulose, and sodium carboxymethyl cellulose. Xanthan gum is preferred.

In the case of bioactive agrochemical compositions that freeze or could be frozen

IM^ÜT

FOR

MEXICAN INSTITUTE T ​​OF PROPERTY '&lt;INDUSTRIAL

<img file="MX336953B_D0089.tif" />

To or similar diethylene glycol, during storage or use, especially concentrates and aqueous and water-based solutions, it would be desirable to add antifreeze additives. Specific examples of suitable antifreeze include ethylene glycol, 1,2-propylene glycol, 1,3-propylene glycol, 1,2-butanediol, 1,3-butanediol, 1,4-butanediol, 1,4-pentanediol, 3-methyl-l, 5-pentanediol, 2 , 3-dimethyl-2,3-butanediol, trimethylolpropane, mannitol, sorbitol, glyceri, pentaerythritol, 1,4cyclohexandimethanol, xylenol, bisphenols such as bisphenol

In addition, ether alcohols such as triethylene glycol, tetraethylene glycol, polyoxyethylene glycols or polyoxypropylene glycols with a molecular weight of up to about 4000, diethylene glycol monomethyl ether, diethylene glycol ether, butylene glycol, monobutyl ether, triethylene glycol, butylene glycol ether tetraglycerol, pentaglycerol, hexaglycerol, heptaglycerol, octaglycerol and the like. As a particular subset of suitable antifreeze materials, there may be mentioned ethylene glycol, propylene glycol, and glycerin.

It is possible to use colorants such as inorganic pigments, such as, for example: iron oxides, titanium oxides, Prussian Blue; organic dyes,

<img file="MX336953B_D0090.tif" />

such as those of the metal alizarin, azo or phthalocyanine type; or of trace elements such as iron, manganese, boron, copper, cobalt, molybdenum or zinc salts. The use of such dyes allows it to be determined which areas and substrates, including plants, have been treated with the bioactive composition. Such marking is especially important for a variety of reasons and for a variety of applications. For example, the use of colorants in seed treatments can allow a quick visual determination of which seeds have been treated and which have not been treated. Similarly, in disinfectant applications, for example in biotechnology laboratories, microbiology laboratories, facilities for manufacturing and processing of food and / or pharmaceuticals and the like, the use of the colorant allows those who perform the cleaning operation to ensure that all surfaces are treated. In this case, for example, the material could be applied and allowed to sit for a short period before cleaning to leave the surface clean.

In addition, in aerial, launch or emission applications, it allows the driver or driver of the dispensing vehicle to see which areas have already been treated.

Although not all additives and adjuvants have been described above, those skilled in the art,

<img file="MX336953B_D0091.tif" />

particularly in the art relevant to the anticipated specific end-use application, you will certainly appreciate what other ingredients, additives and the like could or should be used for your application. The amount with which each additive is to be incorporated into the compositions, again, depends on the end use application and the application method and environment in which it is to be used.

Generally, however, the selection and quantity is that which is conventional for such additives in such applications. However, with the selection of any additives, it is important to ensure that they do not interfere with the bioactivity of the compositions of the present invention or that any such interference can be minimized to allow the greatest advantage to be taken from the compositions. Bioactives of the present invention. Those skilled in the art, based on the teachings set forth herein and the following examples, will appreciate where attention should be paid and, in any case, the same can be addressed by simple selection applications.

As stated above, it is important to avoid the use of conventional bioactive agrochemical assets as well as any other additives and components, including those of the types mentioned above, that

<img file="MX336953B_D0092.tif" />

interfere with or adversely affect the bioefficacy of the compositions in accordance with the present invention. In a very special way, it is important to avoid the use of those agrochemical active ingredients and other additives or compounds that are known or are likely to be irreversibly or strongly bound, bound, or complex with the antimicrobial metal ions in solution. Therefore, without wishing to be bound by theory, it is believed that retention of the ionic charge of the antimicrobial metal is important in maintaining bioefficacy.

For example, especially with regard to copper ions, it is better to avoid the use of ammonium salts such as ammonium sulfate, ammonium chloride, ammonium citrate, ammonium phosphate. To the extent that such materials are present or used, their use, or, more accurately, the amount thereof, should be minimized and / or increase the concentration of the metal ion to compensate for the loss of free ions in the compounds in solution.

The compositions of the present invention can be made using any known method of formulating agrochemical compositions, especially compositions of the antimicrobial and antifungal type. Generally speaking, whether a concentrate is made or ready-to-use bioactive agrochemical compositions

INST

<img file="MX336953B_D0093.tif" />

application of the present invention or if a liquid system or a solid system is made, the bioactive acid solution or, if applicable, the solid bioactive acid composition is prepared before the addition of a conventional bioactive agrochemical active or formulation.

The bioactive acid solution can be prepared in a number of conventional ways. For example, each component can be dissolved in the appropriate solvent, most notably water or a water-based solvent, and the solutions are combined in the appropriate proportions. To some degree, the sequence of the addition and whether or not a pre-concentrate of the acid is formed in the solvent depends on the solubility of the solids themselves. Preferably, the acid is initially dissolved in the appropriate solvent to the desired concentration. In the event that it is intended to form a concentrate, the amount of acid to be dissolved in the solvent must be such that the acid concentration is at least 40 percent and preferably 40 to 80 percent. The antimicrobial metal ion source (s) are then dissolved in the concentrated acidic solution. This method can also be used to prepare a non-concentrated bioactive agrochemical composition in which the rate at which the antimicrobial metal ion source (s) dissolves increases with

<img file="MX336953B_D0094.tif" />

higher acid concentrations. For example, as mentioned above, in cases where the metal ion source is an antimicrobial metal ion-containing ion exchange type agent, especially those whose core is a zeolite, the use of concentrated acids has been found easily dissolves zeolite. After this, the concentrated solution is simply diluted to the desired concentration after the solids dissolve.

In cases where there is difficulty in dissolving the antimicrobial metal source (s) in the concentrate or dilute acid solution, or the rate is undesirably slow, the antimicrobial metal ion source (s) may first be dissolved in water or other appropriate water-based solvent and this is combined with the acid solution formed. In this case, the preferred acid solution is of a higher concentration than intended in the bioactive acid solution to compensate for dilution after adding the dissolved antimicrobial metal ion source (s). Similarly, whether concentrates or end-use formulations are prepared, it would be desirable to make individual stock solutions of each of the components of the bioactive acid solution whose stock solutions are then combined in appropriate proportions. Again, the

<img file="MX336953B_D0095.tif" />

8 Concentration of each stock solution could be adjusted to compensate for dilution after they are combined. Clearly, to form concentrates, stock solutions are typically of a higher concentration than would otherwise be necessary if stock solutions are used to prepare the final end-use diluted formulations.

In each of the above instances, the solvent / solutions can be heated and preferably stirred to accelerate the dissolution of the solids in the liquid system. Also, although dissolving the antimicrobial metal ion source (s) is perhaps the simplest and most cost effective method of preparing bioactive acid solutions, these bioactive acid solutions can also be prepared, for example, electrolytically generating the metal ion in acid solutions as observed in Arata et. to the. (US

6,197,814; US 2003 / 0198689A1, US2003 / 0178374A1;

US2005 / 0245605A1 and US2006 / 0115440A1, all of which are incorporated in the present invention for reference in their entirety) or by elevated temperature and pressure as observed in Cummins et. to the. (US 7,192,618, incorporated in the present invention for reference).

Surfactants can be added to the

<img file="MX336953B_D0096.tif" />

9 _, __ bioactive acid solution or concentrate or can be added concurrently with or subsequent to the combination of the bioactive acid solution with the conventional bioactive agrochemical composition.

When you want to make a liquid bioactive acid solution concentrate, you can prepare the highly concentrated solution as discussed above or make a somewhat dilute form which is then further concentrated by allowing some of the solvent to evaporate. This is particularly beneficial in cases where the antimicrobial metal ion source (s) and / or surfactants and / or other constituents are not soluble in and / or insufficiently and / or rapidly dissolved in the acid solution .

Depending on the final form of the bioactive agrochemical composition of the invention, it would possibly be desirable to prepare a solid bioactive acid composition concentrate. These bioactive acid solid composition concentrates can also be made in a number of ways. For example, the antimicrobial metal ion source (s) and, if present, the surfactant, can be dry blended. Dry mixing is still possible even if the surfactant or one of the surfactants is a liquid since the amount

<img file="MX336953B_D0097.tif" />

<img file="MX336953B_D0098.tif" />

used is very low and will be adsorbed or absorbed by dry materials. The dry mixed materials can be used as such or preferably compacted to form granules. Alternatively, the solid bioactive acid composition concentrate can be formed by first preparing the aforementioned bioactive acid solution concentrate, using a volatile solvent, for example, water or a water-based solvent, and then allowing the solvent evaporate to leave solid material. As necessary, the solid material is then crushed or ground to form small particles, powder or granules, of the solid bioactive acid composition.

The bioactive acid solid composition concentrate can be used to form the bioactive acid solution or the bioactive acid composition to be combined with the conventional bioactive agrochemical. In the first case, the solid concentrate is dissolved in an appropriate solvent, especially water or a water-based solvent. Solid bioactive acid compositions can be prepared by dry blending the acid, the antimicrobial metal ion source (s) and the surfactant with a solid filler material or the aforementioned solid bioactive acid composition concentrate can be lowered or

<img file="MX336953B_D0099.tif" />

dilute with solid fillers. Alternatively, and preferably, the solid bioactive acid composition is prepared by treating a filler material with a bioactive acid solution. In this case the liquid bioactive acid solution is applied to or combined with the filler material, which is preferably in the form of a particle, and is adsorbed by and / or absorbed by the particles of the filler material. For example, a mist of the bioactive acid solution can be sprayed or a constant or intermittent stream of the bioactive acid solution can be poured onto the particles as they are spun, stirred, etc.

Given the high transportation costs and ease of dilution, it is more preferable and more cost effective to prepare concentrates, especially liquid concentrates, from the bioactive agrochemical compositions of the invention whose concentrates are then diluted or lowered at the time of application. . Liquid concentrates are prepared by dissolving the conventional bioactive agrochemical in the concentrated bioactive acid solution. Alternatively, in cases where the conventional bioactive agrochemical is soluble or miscible in the same solvent as that used for the bioactive acid solution or in a solvent miscible with it, the latter is

IMPI

MEXICAN INSTITUTE dí · ·, λ ??. Ο?:; ·; Όλο INDUSTRIAL

<img file="MX336953B_D0100.tif" />

first dissolve in said solvent and then said solution is combined with the concentrated bioactive acid solution. In cases where the conventional bioactive agrochemical is not soluble or miscible in the bioactive acid solution, the solvent in the bioactive acid solution, a miscible solvent with the bioactive acid solution, is dissolved in an appropriate solvent and then combined with the bioactive acid solution to form a suspension or emulsion.

These liquid bioactive agrochemical concentrates are then diluted or diluted with an appropriate solvent, especially water or a water-based solvent, to the desired concentration for application.

Alternatively, the bioactive agrochemical concentrates of the invention can be prepared in fluid particle form. In this case, the components of the solid acid concentrate and the conventional bioactive agrochemical active, such as the pure active or in a formulated concentrate, are all dry mixed with each other or, preferably, a dry mixture of the bioactive acid composition concentrate solid, preferably in granular form, dry mix with a solid concentrate, preferably in granular or powder form, of the conventional bioactive agrochemical active as the pure active or in a formulated concentrate.

<img file="MX336953B_D0101.tif" />

The solid bioactive agrochemical compositions of the present invention can be prepared using any number of ways including dilution, with a solid filler or diluent, of a solid bioactive agrochemical concentrate or the combination of a bioactive acid composition, a diluent, or a solid filler and a conventional solid or active bioactive agrochemical active formulated in its end-use concentration.

Alternatively, the solid bioactive agrochemical composition can be prepared by treating an absorbent and / or adsorbent filler material with a bioactive acid solution and combining this with a formulated active or conventional solid bioactive agrochemical active or treating a bioactive agrochemical formulated active or active. conventional absorbent and / or adsorbent in particulate form with a bioactive acid solution. This modality has the additional advantage that the amount or concentration of liquid acid solution applied to the adsorbent or absorbent vehicle or conventional bioactive formulated agrochemical active or active may be higher than would be applied in the liquid dilute state to allow for bioefficacy by installments. longer. In essence, the conventional active or treated vehicle serves as a reservoir for the bioactive components of the liquid acid solution.

<img file="MX336953B_D0102.tif" />

PI

Λ

I?⁴'STITUTO.&gt; .. * ·; · .jt? THE

INDU7¡ '? DAL

<img file="MX336953B_D0103.tif" />

The bioactive agrochemical compositions of the present invention have a multitude of agricultural and horticultural applications including as fungicides, bactericides, and / or agents to combat plant-type, estramenophilic-type, and fungus-type protists and can be applied to seeds, soils, plants, trees, and the like. These compositions show particular promise as fungicides, bactericides, and agents for fighting plant-type, estramenophilic, and mushroom-type protists due to their unique and surprising strong bioefficacy at extremely low levels of antimicrobial metals. For example, the use of these materials allows for effective application rates in which, for example, the amount of copper applied is in the order of grams per hectare, not kilograms as required with conventional copper and copper-based fungicides.

In addition to the marked bioefficacy at such low levels of antimicrobial metal ion, another attribute of the bioactive agrochemical compositions of the invention is that they do not have or have very little phytotoxicity. This is especially important because a bioeffective material that severely damages or annihilates the plant concurrent with the annihilation of the target organism is of little use unless you do not mind losing a crop and are more interested in

<img file="MX336953B_D0104.tif" />

IM F

INSTITUTE OF THE P

INDJ 1

<td>control the</td><td>target organism</td><td>before</td><td>this</td><td>I know</td><td>out</td><td>of</td>

<td>control. Other</td><td>important factor</td><td>and beneficial</td><td colspan="2">associated</td><td>with</td><td>the</td>

<td>compositions</td><td colspan="2">of the invention is the fact</td><td>of</td><td>than</td><td>these</td><td>not</td>

they induce and probably do not induce or are not associated with any resistance in the target organisms. This stands in stark contrast to the use of organic bioactive agrochemicals, especially fungicides and antibiotics, for which studies and commercial practice itself have shown a marked and increasing trend of resistance among target organisms, even within a few years or less than first time use. The development of such microorganisms resistant to bioactive agrochemicals, although currently troublesome, could lead to catastrophic results if neglected.

Typically, the application rate of the inventive bioactive agrochemical compositions of the present invention is such that the total amount of antimicrobial metal ions (as metal) originating from the dissolved antimicrobial metal ion source (s) applied per hectare is about 494.2 grams or less, preferably 247.10 grams or less, more preferably 123.6 grams or less, more preferred 49.42 grams or less. Of course, the specific application rate and, therefore, the total amount applied per hectare, may vary ·

<img file="MX336953B_D0105.tif" />

OF INDUSTRIAL PROPERTY

<img file="MX336953B_D0106.tif" />

from target organism to target organism, from one form to the other and from one application method to the other. Indeed, the appropriate rates may be such that the total metal ion (as metal) may be on the order of 12.35 grams per hectare, even on the order of fractions of one gram per hectare, perhaps as low as 1.24 grams per hectare. or even 0.02 grams per hectare. Although higher loads, greater than 494.2 grams per hectare, can provide even greater or faster bioefficacy, the balance between increased environmental, health and safety problems is generally not justified or typically not justified by the increase, often nominal increase , in bioefficacy.

The bioactive agrochemical compositions of the present invention can be applied to any number of agricultural crops, including horticultural crops, including ornamental plants, shrubs, and trees; plants for flowers;

fruit trees, vegetable crops; food crops; ornamental grasses and grass; etc. Example crops that are of particular interest due to their significant economic and food impact include soybeans, tomatoes, potatoes, apples, peanuts, grapes, almonds, sugar beets, and citrus. Diseases and microorganisms that are targeted by the

<img file="MX336953B_D0107.tif" />

Bioactive agrochemical compositions of the present invention include, but are not limited to citrus cancer; soybean rust; rust striated from the leaves, from the stem; leaf blights; early blights; late blights;

fire blight; leaf spots; powdery mildew; bacterial cancer; early rot; leaf blight by Alternaría;

leaf spot by Alternaria; leaf spot by Fabrea;

bacterial wilt; Pierce's disease; partial carbon (Karnal bunt); citrus greening;

potato wart; Agrobacterium turnefacíens; Clavibacter michiganensis; Pseudomonas syrinhea; Fusarium; Phytophthora infestans; I would alternate Solani; Erwinia amylovora; Botrytís cinérea; Xanthomonas vesicatoria; and the like. A more comprehensive list of specific pathogens and the cultures they attack is indicated in Tate-US 2 00 5/007 922 7A1, the contents of which are incorporated in the present invention for reference. Typically, the selection of the specific bioactive agrochemical composition to be used for a given target microorganism depends on the conventional bioactive agrochemical active used in the composition.

These compositions can be applied in any conventional manner, sprinkling, dusting, spreading, etc. Typically any given formulation can be applied consistently for the agrochemical component

IMP¹

MEXICAN INSTITUTE OF INDUSTRIAL PROPERTY

<img file="MX336953B_D0108.tif" />

specific conventional bioactive for the target crop and organism. Furthermore, it is anticipated that the bioactive acid solution or composition and the conventional bioactive agrochemical component can be applied concurrently or sequentially (essentially as a two-part system), generally within a few hours one after the other, preferably within one or two hours one after the other, particularly in cases where there is concern that the conventional agrochemical active or one or more components of the formulated active, may interfere with the performance of the bioactive acid solution or composition, for example by sequestering or adversely binding to antimicrobial metal ions. In this case, the bioactive acid solution or composition is applied first and the conventional bioactive agrochemical active or formulation is applied later. Without intending to be bound by theory, it is believed that sequential application allows the first applied bioactive component to make the target organism more susceptible to the second. This is particularly true in cases where the first applied bioactive component is the bioactive acid solution or composition. However, typically, especially for convenience and cost savings, the inventive agrochemical compositions of the invention are applied as an individual composition.

<img file="MX336953B_D0109.tif" />

<img file="MX336953B_D0110.tif" />

INETITV t Cj MEXICANO DE LA PROPÍtuAu

INDUSTRIAL

<img file="MX336953B_D0111.tif" />

Unlike disinfectants in which bioefficacy is measured in terms of logarithmic annihilation, particularly within specified time periods, the bioefficacy of the bioactive agrochemical compositions of the present invention is best represented or demonstrated by an increase in yields or reduction in crop losses. Even a 10% performance improvement can have a significant economic impact.

In essence, even an apparently minor reduction in the target organism or a modest inhibition in the growth or proliferation of the target organism can manifest an acceptable bioefficacy. Also, the duration of this effect does not need to be for life, for example, efficacy may last for a few days or more. Generally, and preferably, it would be desirable to observe a significant reduction, 25% or more, preferably 50% or more, in the growth or proliferation of the target organism over two or more, preferably four or more days. More preferably, it would be desirable to observe an 85% or more, more preferred 95% or more, reduction in growth or proliferation of the target organism over two or more, more preferred four or more days. Again, however, from a commercial perspective, the desired result is an increase, at least a

<img file="MX336953B_D0112.tif" />

10% increase, preferably at least a 30% increase, more preferred a 65% increase, in yield compared to untreated culture.

EXAMPLES

The following examples are presented as a demonstration of the bioefficacy of bioactive agrochemical compositions in accordance with the present invention as well as the unexpected synergy resulting from the use of bioactive acid solutions or bioactive acid compositions in combination with conventional bioactive agrochemical active ingredients. and formulated assets. These examples are only illustrative of the invention and should not be considered as limiting thereof. Those skilled in the art will recognize many variations that are within the scope of the invention and the scope of the claims.

Saccharomyces cerevisiae studies

A series of experiments (Examples 1,269 below) are performed to evaluate the performance of the individual components of the claimed bioactive compositions as well as various combinations thereof, including, the claimed compositions themselves, to

IMPIOUS

M DE LA PIL INSTITUTE

INDU.

suppress the growth of Saecharomyces cerevisiae (baker's yeast Fleishmann). Saccharomyces cerevisiae is selected as a test organism as it is generally accepted in the industry as an indicator or substitute organism for a wide variety of fungi. In each such experiment, the same general procedure is followed unless otherwise indicated.

Experimental detail

A culture medium is prepared by adding 10 grams of growth medium (Sabouraud's dextrose broth,

Difo, from BD of Franklin Lakes, NJ, USA) to 300 ml of distilled water. Fleishmann baker's yeast is then added to the culture medium while stirring using a magnetic stirrer until a uniform dispersion having an initial turbidity of between about 50 and 100 is obtained.

NTU as measured using a HF Instruments DRT 100B turbidity meter. Once the proper dispersion is obtained, 20 ml aliquots are then dispensed, with continuous mixing, into 40 ml borosilicate glass jars with Teflon lined lids (VWR International Cat. No. 15900004). The system / component to be evaluated is then added to the bottle and shaken intimately to ensure a suitable, substantially homogeneous mixture. Later

<img file="MX336953B_D0113.tif" />

<img file="MX336953B_D0114.tif" />

Determine the turbidity of each mixture and the bottle is transferred to an incubator at 30 ° C. Each bottle is periodically removed from the incubator and the mixture of the bottles is evaluated for turbidity: the specific times for said evaluation are as indicated in the discussion of the experiments and in the attached tables.

In each experiment, unless otherwise specified, 2 ml of aqueous solution containing or specified component bioactive system are added to 20 ml of yeast suspension and thoroughly mixed. Surfactants are typically added separately in a concentrated solution in water; however, the added volume is negligible: a fraction of a mi. For the sake of understanding the efficacy levels, the amounts or concentrations of the various components presented in each of the following tables and experiments are from the material diluted in the test bottle: not from the concentrate added to the bottle. Likewise, the concentrations presented are based on a total volume of 20 ml, not the volume of 22+ ml. Multiplying each of the listed concentrations by 0.9 (or 0.95 with those compositions using 1 ml aqueous solutions) provides a more accurate assessment of the concentrations of the various components tested, it is

<img file="MX336953B_D0115.tif" />

that is, a concentration of 5 ppm silver is actually closer to 4.5 ppm. Finally, for those vials to which no bioactive system or component thereof (controls) is added or that contain surfactants only, 2 ml of additional growth medium is added to ensure relative equivalent dilutions of the yeast.

In the following tables, the results are presented as the actual turbidity readings (NTU) with a sub-table presenting the change or delta in the values ​​of

NTU. Given the nature of the system, changes in turbidity are a reflection of the relative performance / bioefficacy of bioactive systems and their components. In some cases, a high level of bioactive material, especially the metal component, causes an immediate and relatively abrupt increase in optical density or turbidity. This is believed to be the result of lysis of at least a portion in the yeast cells themselves. Therefore, especially in those examples that have a high level of bioactive, it is equally, if not more, important to look at the change in turbidity of the turbidity results from either half an hour or an hour, if any, forward. , not from time zero.

<img file="MX336953B_D0116.tif" />

i

MEXICAN INSTITUTE OF THE PRO .'- iEPAO

INDUSTRIAL

<img file="MX336953B_D0117.tif" />

EXAMPLES 1-21

Acid concentration

A first series of experiments is carried out to evaluate the performance of various antimicrobial metals and combinations of these metals, with and without citric acid and with and without the anionic surfactant lauroylsarcosinate sodium. Each of the metals is added in the form of an aqueous solution of its citrate salts, specifically, silver citrate, copper citrate, and zinc citrate, or, in the case of Examples 16-19, as a mixture of all the three citrate salts (Mil). The specific formulations evaluated and the results of the resulting yeast growth study are shown in Tables 1 and 1A.

TABLE 1

<td rowspan="2">Example</td><td rowspan="2">Metal ion and quantity (ppm)</td><td rowspan="2">Acid citric (% in weigh)</td><td rowspan="2">Lauroil Sarcosinate sodium (% by weight)</td><td colspan="5">Turbides (NTU)</td>

<td>Weather zero</td><td>Weather IH</td><td>T 18 hours</td><td>T 24 hours</td><td>T 96 hours</td>

<td> 1</td><td>Ag 5 ppm</td><td> 0</td><td></td><td> 44.5</td><td> 59.6</td><td> 890</td><td> 932</td><td> 995</td>

<td> 2</td><td>Ag 5 ppm</td><td> 1</td><td></td><td> 47.5</td><td> 64</td><td> 882</td><td> 902</td><td> 1044</td>

<td> 3</td><td>Ag 5 ppm</td><td> 2</td><td></td><td> 50.9</td><td> 68.4</td><td> 881</td><td> 950</td><td> 1025</td>

<td> 4</td><td>Ag 5 ppm</td><td> 0</td><td> 0.005</td><td> 46.8</td><td> 51.5</td><td> 596</td><td> 677</td><td> 673</td>

<td> 5</td><td>Ag 5 ppm</td><td> 1</td><td> 0. 005</td><td> 59.4</td><td> 68.4</td><td> 85</td><td> 130</td><td> 854</td>

<td> 6</td><td>Ag 5 ppm</td><td> 2</td><td> 0.005</td><td> 70.9</td><td> 75</td><td> 85</td><td> 120</td><td> 880</td>

<td> 7</td><td>Zn 5 ppm</td><td> 0</td><td></td><td> 43.8</td><td> 64.5</td><td> 992</td><td> 993</td><td> 1051</td>

<td> 8</td><td>Zn 5 ppm</td><td> 1</td><td></td><td> 46.6</td><td> 66.5</td><td> 934</td><td> 962</td><td> 1027</td>

<td> 9</td><td>Zn 5 ppm</td><td> 2</td><td></td><td> 49.5</td><td> 71</td><td> 936</td><td> 1038</td><td> 1063</td>

<td> 10</td><td>Zn 5 ppm</td><td> 0</td><td> 0.005</td><td> 45.9</td><td> 63</td><td> 656</td><td> 747</td><td> 712</td>

<td> 11</td><td>Zn 5 ppm</td><td> 1</td><td> 0.005</td><td> 57</td><td> 71</td><td> 160</td><td> 223</td><td> 744</td>

<img file="MX336953B_D0118.tif" />

TABLE 1 (cont.)

<td rowspan="2">Example 12</td><td rowspan="2">Metal ion and quantity (ppm) Zn 5 ppm</td><td rowspan="2">Citric acid (% in weight) 2</td><td rowspan="2">Lauroil Sarcosinate of s odi o (% by weight) 0.005</td><td colspan="5">Turbidity (NTU)</td>

<td>Weather zero 73</td><td>Weather IH 76.5</td><td>T 18 hours 105</td><td>T 24 hours 119</td><td>T 96 hours 466</td>

<td> 13</td><td>Cu 5 ppm</td><td> 0</td><td></td><td> 45.6</td><td> 68</td><td> 940</td><td> 1021</td><td> 1100</td>

<td> 14</td><td>Cu 5 ppm</td><td> 1</td><td></td><td> 49</td><td> 72</td><td> 940</td><td> 1018</td><td> 1102</td>

<td> 15</td><td>Cu 5 ppm</td><td> 2</td><td></td><td> 49</td><td> 74</td><td> 900</td><td> 973</td><td> 1100</td>

<td> 16</td><td>MI 1</td><td> 0</td><td> 0. 005</td><td> 39</td><td> 44.5</td><td> 449</td><td> 575</td><td> 658</td>

<td> 17</td><td>MI 1</td><td> 1</td><td> 0. 005</td><td> 73.9</td><td> 87</td><td> 100</td><td> 105</td><td> 732</td>

<td> 18</td><td>MI 1</td><td> 2</td><td> 0. 005</td><td> 132</td><td> 137</td><td> 137</td><td> 137</td><td> 690</td>

<td> 19</td><td>MI 1</td><td> 1</td><td> 0.01</td><td> 74.5</td><td> 74.8</td><td> 87</td><td> 89</td><td> 116</td>

<td> 20</td><td>Control (without bioc ida)</td><td></td><td></td><td> 53.2</td><td> 69.4</td><td> 1031</td><td> 1085</td><td> 1122</td>

<td> 21</td><td>Control (without biocide)</td><td></td><td></td><td> 53.2</td><td> 78</td><td> 1101</td><td> 1093</td><td> 1128</td>

MI 1 a 4% citric acid solution containing 50 ppm of each of Ag, Cu and Zn per me which produces 5 ppm of each in the test flasks

TABLE 1A

<td rowspan="2">Example</td><td rowspan="2">Metal ion and quantity (ppm)</td><td rowspan="2">Acid citric (% by weight)</td><td rowspan="2">Lauroil Sodium sarcosinate (¾ by weight)</td><td colspan="4">Change in turbidity from To (delta NTU)</td>

<td>Weather zero</td><td>Weather IH</td><td>T 18 hours</td><td>T 24 hours</td>

<td> 1</td><td>Ag 5 ppm</td><td> 0</td><td></td><td> 15.1</td><td> 845. 5</td><td> 887.5</td><td> 950.5</td>

<td> 2</td><td>Ag 5 ppm</td><td> 1</td><td></td><td> 16. 5</td><td> 834. 5</td><td> 854.5</td><td> 996. 5</td>

<td> 3</td><td>Ag 5 ppm</td><td> 2</td><td></td><td> 17.5</td><td> 830. 1</td><td> 899.1</td><td> 974.1</td>

<td> 4</td><td>Ag 5 ppm</td><td> 0</td><td> 0. 005</td><td> 4.7</td><td> 549. 2</td><td> 630.2</td><td> 626.2</td>

<td> 5</td><td>Ag 5 ppm</td><td> 1</td><td> 0. 005</td><td> 9</td><td> 25.6</td><td> 70. 6</td><td> 7 94 . 6</td>

<td> 6</td><td>Ag 5 ppm</td><td> 2</td><td> 0. 005</td><td> 4 1</td><td> 14.1</td><td> 49. 1</td><td> 809.1</td>

<td> 7</td><td>Zn 5 ppm</td><td> 0</td><td></td><td> 20. 7</td><td> 948. 2</td><td> 94 9.2</td><td> 1007.2</td>

<td> 8</td><td>Zn 5 ppm</td><td> 1</td><td></td><td> 19. 9</td><td> 887. 4</td><td> 915.4</td><td> 980.4</td>

<td> 9</td><td>2n 5 ppm</td><td> 2</td><td></td><td> 21.5</td><td> 886. 5</td><td> 988.5</td><td> 1013.5</td>

<td> 10</td><td>Zn 5 ppm</td><td> 0</td><td> 0. 005</td><td> 17.1</td><td> 610. 1</td><td> 701.1</td><td> 666.1</td>

<td> 11</td><td>2n 5 ppm</td><td> 1</td><td> 0. 005</td><td> 14</td><td> 103</td><td> 166</td><td> 687</td>

<td> 12</td><td>Zn 5 ppm</td><td> 2</td><td> 0. 005</td><td> 3.5</td><td> 32</td><td> 46</td><td> 393</td>

<td> 13</td><td>Cu 5 ppm</td><td> 0</td><td></td><td> 22.4</td><td> 894.4</td><td> 975.4</td><td> 1054.4</td>

<td> 14</td><td>Cu 5 ppm</td><td> 1</td><td></td><td> 23</td><td> 8 91</td><td> 969</td><td> 1053</td>

<td> 15</td><td>Cu 5 ppm</td><td> 2</td><td></td><td> 25</td><td> 851</td><td> 924</td><td> 1051</td>

<td> 16</td><td>MI 1</td><td> 0</td><td> 0.005</td><td> 5.5</td><td> 410</td><td> 536</td><td> 619</td>

<img file="MX336953B_D0119.tif" />

<img file="MX336953B_D0120.tif" />

TABLE IA (cont.)

<td rowspan="2">Example</td><td rowspan="2">Metal ion and quantity (ppm)</td><td rowspan="2">Acid citric (% by weight)</td><td rowspan="2">Laur oil Sodium sarcosinate (% by weight)</td><td colspan="4">Turbidity change from Tq (delta NTU)</td>

<td>Weather zero</td><td>Weather IH</td><td>T 18 hours</td><td>T 24 hours</td>

<td> 17</td><td>MI 1</td><td> 1</td><td> 0. 005</td><td> 13. 1</td><td> 26.1</td><td> 31.1</td><td> 658. 1</td>

<td> 13</td><td>MI 1</td><td> 2</td><td> 0. 005</td><td> 5</td><td> 5</td><td> 5</td><td> 558</td>

<td> 19</td><td>MI 1</td><td> 1</td><td> 0.01</td><td> 0.3</td><td> 12.5</td><td> 14.5</td><td> 41.5</td>

<td> 20</td><td>Control (without Biocide)</td><td></td><td></td><td> 16. 2</td><td>977. δ</td><td> 1031.8</td><td> 1068.8</td>

<td> 21</td><td>With role (without Biocide)</td><td></td><td></td><td> 24.8</td><td> 1047.8</td><td> 1039.8</td><td> 1074.8</td>

MI 1 a 4% citric acid solution containing 50 ppm of each of Ag, Cu and Zn per me which produces 5 ppm of each in the test flasks

As seen in Tables 1 and IA, those formulations having both the acid and the anionic surfactant provide inhibition of marked yeast growth through at least the first 24-hour period, even with the lowest level of agent. anionic surfactant. Those samples with only the metal ion or the metal ion in combination with the acid have no appreciable effect on yeast growth.

Although some inhibition is also observed in those samples in which only the metal or metals and the surfactant are present, the inhibition is not appreciable. Instead, as noted, the additional presence of excess acid produces a marked and unexpected level of improvement. Finally, the formulation that has all three antimicrobial metal ions, plus the acid and agent

<img file="MX336953B_D0121.tif" />

Surfactant provided continues to show excellent yeast growth inhibition even at the 96 hour trial limit.

EXAMPLES 22 - 42

Surfactant evaluation

A similar series of experiments is again conducted to evaluate the performance of various combinations of the components of the bioactive compositions of the present invention as well as to demonstrate other anionic surfactants and surfactant combinations. The specific formulations evaluated and the results of yeast growth are presented in Tables 2 and 2A.

TABLE 2

<td rowspan="2">Example</td><td rowspan="2">Me ta1 citrates (ppm) in citric acid to 0.4%</td><td rowspan="2">Surfactant (% in weigh)</td><td colspan="5">Turbidity (NTU)</td>

<td>Weather zero</td><td>T 1 Hour</td><td>T 18 hours</td><td>T 24 hours</td><td>T 96 hours</td>

<td> 22</td><td>Copper 5 ppm</td><td></td><td> 103</td><td> 114</td><td> 410</td><td> 4 63</td><td> 588</td>

<td> 23</td><td>Zinc 5 ppm</td><td></td><td> 103</td><td> 118</td><td> 475</td><td> 488</td><td> 589</td>

<td> 24</td><td>Silver 5 ppm</td><td></td><td> 155</td><td> 168</td><td> 181</td><td> 190</td><td> 670</td>

<td> 25</td><td>Copper 5 ppm</td><td>NaLS 0.005</td><td> 145</td><td> 146</td><td> 157</td><td> 160</td><td> 149</td>

<td> 26</td><td>Copper 5 ppm</td><td>SLS 0.005</td><td> 119</td><td> 128</td><td> 252</td><td> 326</td><td> 502</td>

<td> 27</td><td>Copper 5 ppm</td><td>NaLS 0.005: SLS 0.005</td><td> 145</td><td> 144</td><td> 156</td><td> 154</td><td> 157</td>

<td> 28</td><td>Zinc 5 ppm</td><td>NaLS 0.005</td><td> 148</td><td> 156</td><td> 157</td><td> 157</td><td> 157</td>

<td> 29</td><td>Zinc 5 ppm</td><td>SLS 0.005</td><td> 126</td><td> 134</td><td> 217</td><td> 234</td><td> 539</td>

<img file="MX336953B_D0122.tif" />

<img file="MX336953B_D0123.tif" />

TABLE 2 (cont.)

<td rowspan="2">E] emplo 30</td><td rowspan="2">Metal citrates (ppm) in citric acid to 0.4% Zinc 5 ppm</td><td rowspan="2">Surfactant (% by weight) NaLS 0.005: SLS 0.005</td><td colspan="5">Turbidity (NTU)</td>

<td>Weather zero 155</td><td>T 1 Hour 155</td><td>T 18 hours 157</td><td>T 24 hours 157</td><td>T 96 hours 158</td>

<td> 31</td><td>Silver 5 ppm</td><td>NaLS 0.005</td><td> 170</td><td> 170</td><td> 184</td><td> 184</td><td> 180</td>

<td> 32</td><td>Silver 5 ppm</td><td>SLS 0.005</td><td> 177</td><td> 177</td><td> 193</td><td> 196</td><td> 196</td>

<td> 33</td><td>Silver 5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> 193</td><td> 190</td><td> 198</td><td> 199</td><td> 199</td>

<td> 34</td><td>Copper 2.5 ppm: Zinc 2.5 ppm</td><td></td><td> 99</td><td> 109</td><td> 498</td><td> 510</td><td> 614</td>

<td> 35</td><td>Copper 2.5 ppm: Silver 2.5 ppm</td><td></td><td> 128</td><td> 152</td><td> 424</td><td> 530</td><td> 727</td>

<td> 36</td><td>Zinc 2.5 ppm: Silver 2.5 ppm</td><td></td><td> 128</td><td> 151</td><td> 541</td><td> 621</td><td> 720</td>

<td> 37</td><td>Control 1 (without biocide)</td><td></td><td> 91</td><td> 114</td><td> 5 60</td><td> 580</td><td> 754</td>

<td> 38</td><td>Control 2 (without biocide)</td><td></td><td> 91</td><td> 114</td><td> 5 63</td><td> 584</td><td> 726</td>

<td> 39</td><td>Charge 2.5 ppm: Zinc 2.5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> 192</td><td> 180</td><td> 193</td><td> 193</td><td> 193</td>

<td> 40</td><td>Copper 2.5 ppm: Silver 2.5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> 181</td><td> 204</td><td> 205</td><td> 206</td><td> 206</td>

<td> 41</td><td>Zinc 2.5 ppm: Silver 2.5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> 194</td><td> 193</td><td> 212</td><td> 212</td><td> 212</td>

<td> 42</td><td>Copper 2.5 ppm: Silver 2.5 ppm: Zinc 2.5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> 193</td><td> 193</td><td> 199</td><td> 200</td><td> 205</td>

* NaLS = sodium lauroylsarcosinate, SLS = sodium iauryl sulfate

TABLE 2A

<td rowspan="2">Example</td><td rowspan="2">Metal citrates (ppm) in 0.4% citric acid</td><td rowspan="2">Surfactant (% by weight)</td><td>Change in</td><td colspan="2">turbidity from To NTU)</td><td>(delta</td>

<td>T 1 hour</td><td>T 18 hours</td><td>T 24 hours</td><td>T 96 Hours</td>

<td> 22</td><td>Copper 5 ppm</td><td></td><td> 11</td><td> 307</td><td> 360</td><td> 485</td>

<td> 23</td><td>Zinc 5 ppm</td><td></td><td> 15</td><td> 372</td><td> 385</td><td> 486</td>

<td> 24</td><td>Silver 5 ppm</td><td></td><td> 13</td><td> 26</td><td> 35</td><td> 515</td>

<td> 25</td><td>Copper 5 ppm</td><td>0. 005 NaLS</td><td> 1</td><td> 12</td><td> 15</td><td> 4</td>

<td> 26</td><td>Copper 5 ppm</td><td>0.005 SLS</td><td> 9</td><td> 133</td><td> 207</td><td> 383</td>

<td> 27</td><td>Copper 5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> -1</td><td> 11</td><td> 9</td><td> 12</td>

<td> 28</td><td>Zinc 5 ppm</td><td>NaLS 0.005</td><td> 8</td><td> 9</td><td> 9</td><td> 9</td>

<td> 29</td><td>Zinc 5 ppm</td><td>SLS 0.005</td><td> 8</td><td> 91</td><td> 108</td><td> 413</td>

<td> 30</td><td>Zinc 5 ppm</td><td>NaLS 0.005: SLS 0.005</td><td> 0</td><td> 2</td><td> 2</td><td> 3</td>

<td> 31</td><td>Silver 5 ppm</td><td>NaLS 0.005</td><td> 0</td><td> 14</td><td> 14</td><td> 10</td>

<td> 32</td><td>Silver 5 ppm</td><td>0. 005 of SLS</td><td> 0</td><td> 16</td><td> 19</td><td> 19</td>

<td> 33</td><td>Silver 5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> -3</td><td> 5</td><td> 6</td><td> 6</td>

<img file="MX336953B_D0124.tif" />

TABLE 2A (cont.)

<td rowspan="2">Example</td><td rowspan="2">Metal citrates (ppm) in 0.4% citric acid</td><td rowspan="2">Surfactant (% by weight)</td><td colspan="4">Change in turbidity from To (delta NTU)</td>

<td>T 1 hour</td><td>T 18 hours</td><td>T 24 hours</td><td>T 96 Hours</td>

<td> 34</td><td>Copper 2.5 ppm: Zinc 2.5 ppm</td><td></td><td> 10</td><td> 399</td><td> 411</td><td> 515</td>

<td> 35</td><td>Copper 2.5 ppm: Silver 2.5 ppm</td><td></td><td> 24</td><td> 296</td><td> 402</td><td> 599</td>

<td> 36</td><td>Zinc 2.5 ppm: Silver 2.5 ppm</td><td></td><td> 23</td><td> 413</td><td> 493</td><td> 5 92</td>

<td> 37</td><td>Control 1 (without biocide)</td><td></td><td> 23</td><td> 469</td><td> 489</td><td> 6 63</td>

<td> 38</td><td>Control 2 (without biocide)</td><td></td><td> 23</td><td> 472</td><td> 493</td><td> 635</td>

<td> 39</td><td>Copper 2.5 ppm: Zinc 2.5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> -12</td><td> 1</td><td> 1</td><td> 1</td>

<td> 40</td><td>Copper 2.5 ppm: Silver 2.5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> 23</td><td> 24</td><td> 25</td><td> 25</td>

<td> 41</td><td>Zinc 2.5 ppm: Silver 2.5 ppm</td><td>NaLS 0.005: 0.005 from SLS</td><td> -1</td><td> 18</td><td> 18</td><td> 18</td>

<td> 42</td><td>Copper 2.5 ppm: Silver 2.5 ppm: Zinc 2.5 ppm</td><td>NaLS 0.005: SLS 0.005</td><td> 0</td><td> 6</td><td> 7</td><td> 12</td>

Again, the importance of all three constituents is evident from the results shown in Tables 2 and 2A. These results also confirm that even a low excess acid content, in this case 0.4%, provides excellent inhibition in yeast growth over 96 hours. The slightly less than ideal results shown in Examples 26 and 29 suggest some variation between anionic surfactants, at least with sodium lauryl sulfate (SLS), with zinc and copper ions. However, the results are still significantly better than without a surfactant and suggest a possible synergy with two. Also, due to the easier solubility of SLS, in

100

T PT

MEXICAN INSTITUTE Dli THE PROPERTY

INDUSTRIAL

<img file="MX336953B_D0125.tif" />

Compared to sodium lauroylsarcosinate (NaLS), the presence of SLS helps to improve and / or increase the solubility of NaLS under acidic conditions.

EXAMPLES 43-57

Low concentration evaluation

A series of experiments are again conducted to evaluate the performance of various combinations of the components of the bioactive compositions of the present invention, this time focusing on the impact of low concentrations of the components and their combinations.

In this set of experiments, 1 ml aqueous solutions of the bioactive / citric acid components are added to the 20 ml bottles. The specific formulations evaluated and the results of yeast growth are presented in Tables 3 and 3A.

TABLE 3

<td rowspan="2">Example</td><td rowspan="2">Metal bioactive *</td><td rowspan="2">Acid citric (% by weight)</td><td rowspan="2">Surfactant (% by weight) **</td><td colspan="5">Turbidity (NTU)</td>

<td>DO (To)</td><td>DO (Tlhr)</td><td>DO (T18)</td><td>DO (T24)</td><td>DO (T48)</td>

<td> 43</td><td></td><td></td><td>NaLS 0.01</td><td> 43</td><td> 45</td><td> 550</td><td> 613</td><td> 521</td>

<td> 44</td><td></td><td></td><td>NaLS 0.02</td><td> 43</td><td> 40</td><td> 4 60</td><td> 524</td><td> 624</td>

<td> 45</td><td></td><td></td><td>SLS 0.01</td><td> 43</td><td> 47</td><td> 675</td><td> 728</td><td> 758</td>

<td> 46</td><td></td><td></td><td>SLS 0.02</td><td> 37</td><td> 42</td><td> 4 95</td><td> 610</td><td> 605</td>

<img file="MX336953B_D0126.tif" />

101

<img file="MX336953B_D0127.tif" />

TABLE 3 (cont.)

<td rowspan="2">Example</td><td rowspan="2">Metal bioactive *</td><td rowspan="2">Citric acid (% in weight)</td><td rowspan="2">Surfactant (% by weight) **</td><td colspan="5">Turbidity (NTU)</td>

<td>DO (To)</td><td>DO (Tlhr)</td><td>DO (T18)</td><td>DO (T24)</td><td>DO (T48)</td>

<td> 47</td><td></td><td></td><td>0.01 from NaLS / 0.01 of SLS</td><td> 40</td><td> 41</td><td> 370</td><td> 466</td><td> 580</td>

<td> 48</td><td></td><td></td><td>0.005 from NaLS / 0. 005 from SLS</td><td> 43</td><td> 47</td><td> 630</td><td> 696</td><td> 728</td>

<td> 49</td><td></td><td> 0.05</td><td></td><td> 42</td><td> 46</td><td> 835</td><td> 920</td><td> 878</td>

<td> 50</td><td></td><td> 0.1</td><td></td><td> 38</td><td> 44</td><td> 780</td><td> 8 64</td><td> 852</td>

<td> 51</td><td>MI 1</td><td> 0. 2</td><td></td><td> 50</td><td> 62</td><td> 809</td><td> 8 91</td><td> 915</td>

<td> 52</td><td>MI 1</td><td> 0. 2</td><td>NaLS 0.01</td><td> 64</td><td> 63</td><td> 67</td><td> 68</td><td> 69</td>

<td> 53</td><td>MI 1</td><td> 0.2</td><td>NaLS 0.02</td><td> 61</td><td> 65</td><td> 300</td><td> 569</td><td> 1039</td>

<td> 54</td><td>MI 1</td><td> 0. 2</td><td>0.005 from SLS NaLS / 0.005</td><td> 60</td><td> 63</td><td> 62</td><td> 63</td><td> 73</td>

<td> 55</td><td>MI 1</td><td> 0.2</td><td>0.01 from NaLS / 0.01 of SLS</td><td> 85</td><td> 76</td><td> 76</td><td> 79</td><td> 79</td>

<td> 56</td><td>Control 1</td><td></td><td></td><td> 43</td><td> 51</td><td> 9 60</td><td> 997</td><td> 939</td>

<td> 57</td><td>Control 2</td><td></td><td></td><td> 43</td><td> 51</td><td> 8 90</td><td> 986</td><td> 887</td>

* MI 1 a 4¾ citric acid solution containing 50 ppm each of Ag, Cu, and Zn by me which produces ~ 5 ppm each in the test flasks ** NaLS = sodium lauroylsarcosinate, SLS = lauryl sulfate of sodium

TABLE 3A

<td rowspan="2">Example</td><td rowspan="2">Me so bioactive *</td><td rowspan="2">Citric acid (% by weight)</td><td rowspan="2">Surfactant (% in weight)**</td><td colspan="4">Change in turbidity from To (delta NTU)</td>

<td>DO (Tlhr)</td><td>DO (T18)</td><td>DO (T24)</td><td>DO (T48)</td>

<td> 43</td><td></td><td></td><td>NaLS 0.01</td><td> 2</td><td> 507</td><td> 570</td><td> 478</td>

<td> 44</td><td></td><td></td><td>NaLS 0.02</td><td> -3</td><td> 417</td><td> 481</td><td> 581</td>

<td> 45</td><td></td><td></td><td>SLS 0.01</td><td> 4</td><td> 632</td><td> 685</td><td> 715</td>

<td> 46</td><td></td><td></td><td>SLS 0.02</td><td> 5</td><td> 458</td><td> 573</td><td> 568</td>

<td> 47</td><td></td><td></td><td>NaLS 0.01 / 0.01 NaLS SLS</td><td> 1</td><td> 330</td><td> 426</td><td> 540</td>

<td> 48</td><td></td><td></td><td>NaLS / 0 0.005. SLS 005</td><td> 4</td><td> 587</td><td> 653</td><td> 683</td>

<td> 49</td><td></td><td> 0. 05</td><td></td><td> 4</td><td> 793</td><td> 878</td><td> 836</td>

<td> 50</td><td></td><td> 0. 1</td><td></td><td> 6</td><td> 742</td><td> 826</td><td> 814</td>

102

TABLE 3A (eont.)

<img file="MX336953B_D0128.tif" />

<td rowspan="2">Example</td><td rowspan="2">Metal bioactive *</td><td rowspan="2">Acid citric (% by weight)</td><td rowspan="2">Tensive agent (% in weight)**</td><td colspan="4">Change in turbidity from Tq (delta NTU)</td>

<td>DO (Tlhr)</td><td>DO (T18)</td><td>DO (T24)</td><td>DO (T48)</td>

<td> 51</td><td>MI 1</td><td> 0. 2</td><td></td><td> 12</td><td> 759</td><td> 841</td><td> 865</td>

<td> 52</td><td>MI 1</td><td> 0.2</td><td>NaLS 0.01</td><td> -1</td><td> 3</td><td> 4</td><td> 5</td>

<td> 53</td><td>MI 1</td><td> 0. 2</td><td>NaLS 0.02</td><td> 4</td><td> 239</td><td> 508</td><td> 978</td>

<td> 54</td><td>MI 1</td><td> 0. 2</td><td>NaLS 0.005 / SLS 0.005</td><td> 3</td><td> 2</td><td> 3</td><td> 13</td>

<td> 55</td><td>MI 1</td><td> 0. 2</td><td>0.01 from SLS NaLS / 0.01</td><td> -9</td><td> -9</td><td> -6</td><td> -6</td>

<td> 56</td><td>Control 1</td><td></td><td></td><td> 8</td><td> 917</td><td> 954</td><td> 896</td>

<td> 57</td><td>Control 2</td><td></td><td></td><td> 8</td><td> 847</td><td> 943</td><td> 844</td>

* MI 1 a 4% citric acid solution containing 50 ppm of each of Ag, Cu and Zn p mi which produces * 5 ppm of each in the test flasks ** NaLS = sodium lauroylsarcosinate, SLS = sodium lauryl sulfate

As seen in Tables 3 and 3A, again the combination of bioactive metal ions, citric acid, and anionic surfactant demonstrates marked inhibition in yeast growth compared to individual components, even at low excess acid concentrations and surfactant. However, once again, surfactants appear to have a marginal inhibitory effect, compared to controls, by themselves, the inhibition is negligible compared to that of systems according to the present invention.

<img file="MX336953B_D0129.tif" />

<img file="MX336953B_D0130.tif" />

103

EXAMPLES 58-71

Ion exchange metal ion source

A metal citrate solution is prepared by adding approximately 4 grams of citric acid to approximately 8 grams of water and mixing until completely dissolved. After this, 0.1 grams of each of the two ion exchange type antimicrobial agents, AglON AC10D antimicrobial agents and

AglON AK10D from AglON Technologies of Wakefield, MA, USA, to the concentrated citric acid solution with stirring until the antimicrobial agents are completely dissolved.

Then approximately 92 grams of water are added to provide a 4% citric acid solution having 0.1% by weight of AC10D and 0.1% by weight of

AK10D. AglON's AK10D contains approximately 5.0% by weight of silver and approximately 13% by weight of zinc and AC10D of

AglON contains approximately 6.0% by weight of copper and approximately 3.5% by weight of silver. Various amounts of the citric acid solution thus formed are then added to the test bottles to provide a silver content in the test bottles of approximately 1.25 ppm, 2.5 ppm, 5.0 ppm, and 10 ppm. Additionally, different surfactants and combinations of

<img file="MX336953B_D0131.tif" />

Surfactants to some vials to demonstrate the effect of different metal and acid contents on bioefficacy with and without surfactants. The specific formulations evaluated and the results of yeast growth are presented in Tables 4 and 4A.

TABLE 4

<td rowspan="2">Ex-</td><td rowspan="2">Cone of Ag ppm</td><td rowspan="2">Surfactant (% by weight) *</td><td colspan="6">Turbidity (NTU)</td>

<td>DO (T 0)</td><td>DO (Tlhr)</td><td>DO (Tl8hr)</td><td>DO (T24hr)</td><td>DO (T44hr)</td><td>DO (T120HR)</td>

<td> 58</td><td> 1.25</td><td></td><td> 108</td><td> 128</td><td> 913</td><td> 880</td><td> 954</td><td> 1136</td>

<td> 59</td><td> 2. 5</td><td></td><td> 127</td><td> 157</td><td> 8 65</td><td> 8 90</td><td> 941</td><td> 1024</td>

<td>SW</td><td> 5</td><td></td><td> 176</td><td> 199</td><td> 229</td><td> 227</td><td> 234</td><td> 721</td>

<td> 61</td><td> 10</td><td></td><td> 168</td><td> 173</td><td> 191</td><td> 191</td><td> 190</td><td> 180</td>

<td> 62</td><td> 1.25</td><td>0.005 from NaLS</td><td> 143</td><td> 158</td><td> 240</td><td> 5 60</td><td> 843</td><td> 708</td>

<td> 63</td><td> 2. 5</td><td>0.005 from NaLS</td><td> 180</td><td> 179</td><td> 2 04</td><td> 210</td><td> 729</td><td> 843</td>

<td> 64</td><td> 5</td><td>0.005 from NaLS</td><td> 194</td><td> 201</td><td> 222</td><td> 221</td><td> 227</td><td> 227</td>

<td> 65</td><td> 1.25</td><td>0.005 from SLS</td><td> 136</td><td> 167</td><td> 953</td><td> 930</td><td> 973</td><td> 1132</td>

<td> 66</td><td> 2. 5</td><td>0.005 from SLS</td><td> 201</td><td> 212</td><td> 880</td><td> 880</td><td> 967</td><td> 1145</td>

<td> 61</td><td> 5</td><td>0.005 from SLS</td><td> 248</td><td> 247</td><td> 272</td><td> 272</td><td> 296</td><td> 297</td>

<td> 68</td><td> 1.25</td><td>NaLS 0.0025 / SLS 0.0025</td><td> 166</td><td> 180</td><td> 343</td><td> 730</td><td> 957</td><td> 986</td>

<td> 69</td><td> 2. 5</td><td>NaLS 0.0025 / SLS 0.0025</td><td> 215</td><td> 217</td><td> 235</td><td> 239</td><td> 759</td><td> 940</td>

<td> 70</td><td> 5</td><td>NaLS 0.0025 / SLS 0.0025</td><td> 235</td><td> 235</td><td> 257</td><td> 255</td><td> 259</td><td> 268</td>

<td> 71</td><td>Control</td><td></td><td> 101</td><td> 125</td><td> 1050</td><td> 1050</td><td> 1040</td><td> 1183</td>

NaLS = sodium lauroylsarcosinate, SLS = sodium lauryl sulfate

<img file="MX336953B_D0132.tif" />

105

<img file="MX336953B_D0133.tif" />

TABLE 4A

<td rowspan="2">Ex.</td><td rowspan="2">Ag Conc. ppm</td><td colspan="2">Agent surfactant</td><td colspan="5">Change in Turbidity (delta NTU)</td>

<td>(% in</td><td>weight)*</td><td>DO (Tlhr)</td><td>DO (T18hr)</td><td>DO (T24hr)</td><td>DO (T44hr)</td><td>DO (T120HR)</td>

<td> 58</td><td> 1.25</td><td colspan="2"></td><td> 20</td><td> 805</td><td> 772</td><td> 846</td><td> 1028</td>

<td> 59</td><td> 2.5</td><td colspan="2"></td><td> 30</td><td> 738</td><td> 763</td><td> 814</td><td> 8 97</td>

<td> 60</td><td> 5</td><td colspan="2"></td><td> 23</td><td> 53</td><td> 51</td><td> 58</td><td> 545</td>

<td> 61</td><td> 10</td><td colspan="2"></td><td> 5</td><td> 23</td><td> 23</td><td> 22</td><td> 12</td>

<td> 62</td><td> 1. 25</td><td> 0.005</td><td>by NaLS</td><td> 15</td><td> 97</td><td> 417</td><td> 700</td><td> 5 65</td>

<td> 63</td><td> 2.5</td><td> 0.005</td><td>by NaLS</td><td> -1</td><td> 24</td><td> 30</td><td> 549</td><td> 663</td>

<td> 64</td><td> 5</td><td> 0.005</td><td>by NaLS</td><td> 7</td><td> 28</td><td> 27</td><td> 33</td><td> 33</td>

<td> 65</td><td> 1.25</td><td> 0.005</td><td>from SLS</td><td> 31</td><td> 817</td><td> 794</td><td> 837</td><td> 996</td>

<td> 66</td><td> 2.5</td><td> 0.005</td><td>from SLS</td><td> 11</td><td> 679</td><td> 67 9</td><td> 766</td><td> 944</td>

<td> 67</td><td> 5</td><td> 0.005</td><td>from SLS</td><td> -1</td><td> 24</td><td> 24</td><td> 48</td><td> 49</td>

<td> 68</td><td> 1. 25</td><td>0.0025 NaLS / 0 from SLS</td><td>of . 0025</td><td> 14</td><td> 177</td><td> 564</td><td> 791</td><td> 820</td>

<td> 69</td><td> 2.5</td><td>0.0025 SLS NaLS / 0</td><td>of . 0025</td><td> 2</td><td> 20</td><td> 24</td><td> 544</td><td> 725</td>

<td> 70</td><td> 5</td><td>0.0025 SLS NaLS / 0</td><td>of . 002 5</td><td> 0</td><td> 22</td><td> 20</td><td> 24</td><td> 33</td>

<td> 71</td><td>Control</td><td colspan="2"></td><td> 24</td><td> 949</td><td> 94 9</td><td> 939</td><td> 1082</td>

* NaLS = sodium lauroylsaicosinate, SLS = sodium lauryl sulfate

As seen in Tables 4 and 4A, the compositions according to the present invention provide marked inhibition in yeast growth.

Although Example 61 contains the highest concentration of metal ions (10 ppm silver, 7 ppm copper, and 15.3 ppm zinc), it shows good inhibition of yeast growth,

The highest degree of efficacy comes with the concomitant increase in the release of these metals into the

106

<img file="MX336953B_D0134.tif" />

environment. This becomes especially important in cases where bioactive materials are to be used in or near marine and / or agricultural applications. Therefore, although high concentrations of metal, especially silver, may provide better bioefficacy, they also accelerate the impact on aquatic environments. On the other hand, as shown in those examples using antimicrobial metal-containing acid solutions with the anionic surfactant, especially sodium lauroylsarcosinate, alone or in combination with sodium lauryl sulfate, the same and even better yeast inhibition is obtained with less than half, even less than a quarter, of the metal ion concentrations. Furthermore, these results show that by adjusting the level of the surfactant, the level of metal ion can be further reduced while still providing marked inhibition of the fungus.

Also surprising about this example is the discovery that citric acid can dissolve antimicrobial zeolite particles. This discovery presents other means by which the compositions of the invention can be made as well as a number of alternative applications for such materials not otherwise possible with zeolites in their solid form.

<img file="MX336953B_D0135.tif" />

EXAMPLES 72-79

Metal concentration ·

A concentrated bioactive system (MI2) is prepared for this study, comprising a 16% aqueous citric acid solution that has dissolved in the same silver citrate, copper citrate and zinc citrate, each added in an amount to provide 200 ppm of each metal, along with

0.25% sodium lauroylsarcosinate and 0.32% sodium lauryl sulfate. Various amounts of this system are added to the test flasks to also assess the impact of yeast inhibiting metal concentration. An additional example is prepared that also includes a nonionic surfactant, Tween 20 (polyoxyethylene (20) sorbitan monolaurate), an emulsifier to evaluate its impact on performance. The specific formulations evaluated and the results are presented in Tables 5 and 5A.

TABLE 5

<td rowspan="2">Example</td><td rowspan="2">MY 2* aggregate (me)</td><td rowspan="2">Concentration of each metal (ppm)</td><td colspan="6">Turbidity (NTU)</td>

<td>T 0</td><td>T 18</td><td>T 22</td><td>T 24</td><td>T 64</td><td>T 82</td>

<td> 72</td><td> 0</td><td> 0</td><td> 63</td><td> 920</td><td> 980</td><td> 964</td><td> 1020</td><td> 1050</td>

<td> 73</td><td> 0. 1</td><td> 1</td><td> 81</td><td> 608</td><td> 722</td><td> 820</td><td> 1077</td><td> 1062</td>

<td> 74</td><td> 0.25</td><td> 2.5</td><td> 111</td><td> 126</td><td> 142</td><td> 160</td><td> 752</td><td> 810</td>

<td> 75</td><td> 0. 5</td><td> 5</td><td> 145</td><td> 198</td><td> 208</td><td> 208</td><td> 205</td><td> 203</td>

<img file="MX336953B_D0136.tif" />

108

TABLE 5 (cont.)

<img file="MX336953B_D0137.tif" />

<td rowspan="2">Example</td><td rowspan="2">M12 * aggregate (me)</td><td rowspan="2">Concentration of each metal (ppm)</td><td colspan="6">Turbidity (NTU)</td>

<td>T 0</td><td>T 18</td><td>T22</td><td>T 24</td><td>T 64</td><td>T 82</td>

<td> 76</td><td> 1. 0</td><td> 10</td><td> 483</td><td> 410</td><td> 395</td><td> 369</td><td> 320</td><td> 300</td>

<td> 77</td><td> 2.0</td><td> 20</td><td> 1295</td><td> 820</td><td> 714</td><td> 660</td><td> 399</td><td> 2 64</td>

<td> 78</td><td> 3. 0</td><td> 30 </td><td> 1435</td><td> 766</td><td> 620</td><td> 555</td><td> 340</td><td> 340</td>

<td> 79</td><td> 0.5^T</td><td> 5</td><td> 141</td><td> 240</td><td> 405</td><td> 600</td><td> 1116</td><td> 1129</td>

* KI2 a 16% citric acid solution containing 200 ppm each of Ag, Cu and Zn per my + this formulation also contains 0.1% by weight of Tween 20 a nonionic surfactant

TABLE 5A

<td rowspan="2">Example</td><td rowspan="2">MY 2* aggregate (me</td><td rowspan="2">Concentration of each metal (ppm)</td><td colspan="5">Change in turbidity (delta NTU)</td>

<td>T18-T0</td><td>T22-T0</td><td>T24-T0</td><td>T64-T0</td><td>T82-T0</td>

<td> 72</td><td> 0</td><td> 0</td><td> 857</td><td> 917</td><td> 901</td><td> 957</td><td> 987</td>

<td> 73</td><td> 0.1</td><td> 1</td><td> 527</td><td> 641</td><td> 739</td><td> 996</td><td> 981</td>

<td> 74</td><td> 0.25</td><td> 2.5</td><td> 15</td><td> 31</td><td> 49</td><td> 641</td><td> 699</td>

<td> 75</td><td> 0.5</td><td> 5</td><td> 53</td><td> 63</td><td> 63</td><td> 60</td><td> 58</td>

<td> 76</td><td> 1.0</td><td> 10</td><td> -73</td><td> -88</td><td> -114</td><td> -163</td><td> -183</td>

<td> 77</td><td> 2 0</td><td> 20</td><td> -475</td><td> -581</td><td> - 635</td><td> -896</td><td> -1031</td>

<td> 78</td><td> 3.0</td><td> 30</td><td> -669</td><td> -815</td><td> -880</td><td> -1095</td><td> -1095</td>

<td> 79</td><td> 0.5⁺</td><td> 5</td><td> 108</td><td> 2 64</td><td> 459</td><td> 975</td><td> 988</td>

* MI2 a 16% citric acid solution containing 200 ppm each of Ag, Cu and Zn per my + this formulation also contains 0.1% by weight of Tween 20 a nonionic surfactant

As seen in Tables 5 and 5A, high concentrations of metals dramatically inhibit, if not completely stop, the growth of

109

IMPI

MEXICAN INSTITUTE OF THE KOMSOAD

INDUSTRIAL

<img file="MX336953B_D0138.tif" />

yeast. The solutions of Examples 76, 77 and 78 containing ultra-high metal content appear to destroy yeast cells, showing what appears to be rapid yeast denaturation by adding the bioactive material to the test flasks. It is possible that the high initial turbidity reflects so much that this arises from the addition of the bioactive materials by themselves as well as the destruction of the yeast cells.

Regardless, the results show that marked inhibition is also obtained at much lower concentrations of the metal in the presence of the excess acid and the surfactant. Indeed, only 15 ppm of metals (5 ppm of each) provide excellent inhibition through the hours and beyond.

Finally, the addition of the Tween surfactant appears to be antagonistic to the action of the bioactive systems of the present invention resulting in a reduction in the level of yeast inhibition. However, this composition (Example 79) exhibits moderate yeast inhibition over 24 hours. Depending on the specific end-use application contemplated, it is apparent that routine preliminary evaluations must be made before formulating with the various additives to establish their impact on the inventive systems of the present invention.

<img file="MX336953B_D0139.tif" />

MEXICAN INSTITUTE DS LA PT.OI'IECAD

INDUSTRIAL

<img file="MX336953B_D0140.tif" />

110

EXAMPLES 80-95

Bioactive synergy

A series of experiments are carried out in which the possible synergies between the compositions of the invention and other bioactive materials as well as between said other bioactive materials including a fungicide, an antimicrobial agent and a disinfectant are evaluated. The bioactive system of the invention used in this set of experiments (MI3) is an aqueous citric acid solution at

4% containing 50 ppm of silver, 50 ppm of copper and 50 ppm of zinc.

The fungicide evaluated is Mancozeb Fluid with Zinc from Bonide Products, Inc. of Oniskany, NY, USA, a commercially formulated fungicide containing 37% by weight of mancozeb.

Although the specific formulation of the Mancozeb product is registered, as a commercial formulation it may also contain some surfactants to allow application to plants for efficacy. Mancozeb is an insoluble, dispersible powder that increases the turbidity of the liquids to which it is added. However, in a separate evaluation, not reproduced herein, Mancozeb was found to be able to control or inhibit yeast growth at a concentration of 1.23 x 10

UTO MEXICANO £ THE PROPERTY

INDUSTRIAL

<img file="MX336953B_D0141.tif" />

111 ___ .

approximately. The tag indicates its use at a rate of 2.6 x

10’³.

The antimicrobial asset evaluated is AglON AC10D, an antimicrobial zeolite additive available from AglON

Technologies, Inc., of Wakefield, MA, USA, which, as noted above, contains 6.0 wt% copper and 3.5 wt% silver. In a separate dilution evaluation, not reproduced herein, an aqueous suspension of AC10D was found to show some control or inhibition of yeast at a concentration of approximately 6.25xl0 "⁴.

Finally, the disinfectant evaluated is AglON

SilverClene 24, a disinfecting material based on an aqueous solution of silver citrate electrolytically generated 30 ppm silver), also distributed by AglON

Technologies, Inc. Although registered, this product and its manufacture are believed to be described in Arata - US

6,583,176, which is incorporated in the present invention for reference in its entirety.

The aforementioned materials as well as various combinations thereof are evaluated to establish their efficacy in stopping or inhibiting the growth of yeast.

The specific formulations analyzed and the yeast inhibition results obtained with them are presented in Tables 6 and 6A.

TABLE 6

<img file="MX336953B_D0142.tif" />

THE

OR

THE

YOU

TABLE 6A

<img file="MX336953B_D0143.tif" />

THE

LT)

OR

ΙΜ

114

INSTITUTO MEXICa DE LA PHO: 'EE

INDUSTRIAL

<img file="MX336953B_D0144.tif" />

The results presented in Tables 6 and 6A demonstrate a marked synergy between inventive compositions in accordance with the present invention and commercial fungicides and antimicrobial agents.

Specifically, for example, a comparison of the results for Examples 80, 81, and 82 demonstrates that combining low amounts of the metal ions, citric acid, and fungicide provides excellent antifungal performance. Although these formulations are observed to have no additional surfactant, the commercial fungicide itself contains surfactants that work in combination with the metal ions and citric acid to provide the benefits due to such a combination as currently claimed. These results show that excellent antifungal activity, as measured by inhibition of yeast growth, can be obtained with less than 10% of the amount of fungicide necessary to inhibit yeast growth by simply adding low levels of the acid and the metal ions. As seen from Examples 91, 92 and 93, a similar synergy is shown for the compositions of the invention in combination with a conventional inorganic antimicrobial agent. In this case too, less than 10% of the amount of the antimicrobial agent needed when iMPi

MEXICAN INSTITUTE -Pódq

Say THE PROPERTY lV

INDUSTRIAL

115 _ used alone, provides good antimicrobial performance when combined with low levels of bioactive composition in accordance with the present invention. However, the replacement of the SilverClene 24 product by the inventive composition of the present invention, Examples 89 and

90, provides no apparent benefit despite the relatively high silver content.

Finally, in Example 86, ammonia is added to a portion of the MI3 solution until the solution reaches a pH of 6. Then 2 ml of this buffered solution is used in the experiment. This example indicates the importance of the low pH of the compositions according to the present invention to provide desirable performance.

EXAMPLES 96-107

Synergy with Immunox

A similar study is performed for synergy between the bioactive compositions of with the present invention and a second fungicide, commercial fungus containing miclobutanil available from Spectrum Brands United Division of Madison, WI, USA. As a formulation it is also expected that it has some content to assess conformity

Immunox, 1.55%,

Commercial, agent industries

<img file="MX336953B_D0145.tif" />

<img file="MX336953B_D0146.tif" />

116

<img file="MX336953B_D0147.tif" />

surfactants. The bioactive composition used in this experiment is the concentrated bioactive system (MI2) produced in Examples 72-79 above. The specific dilutions of each one and the results obtained with them are presented in Table 7.

TABLE 7

<td colspan="3">Dilution ratio</td><td rowspan="2">T zero</td><td rowspan="2">DO T 1.5</td><td rowspan="2">T 18</td><td rowspan="2">DO T 68</td><td rowspan="2">Delta 68</td>

<td>Example</td><td>Immunox</td><td>MY 2</td>

<td> 96</td><td></td><td> 1:80</td><td> 150</td><td> 152</td><td></td><td> 832</td><td> 682</td>

<td> 97</td><td></td><td> 1: 200</td><td> 106</td><td> 112</td><td></td><td> 980</td><td> 874</td>

<td> 98</td><td> 1:64</td><td></td><td> 97</td><td> 107</td><td> 1043</td><td></td><td></td>

<td> . 99</td><td> 1:12 8</td><td></td><td> 111</td><td> 113</td><td> 1126</td><td></td><td></td>

<td> 100</td><td> 1:256</td><td></td><td> 84</td><td> 131</td><td></td><td> 1170</td><td> 1086</td>

<td> 101</td><td> 1:512</td><td></td><td> 81</td><td> 140</td><td></td><td> 12 4 0</td><td> 1159</td>

<td> 102</td><td> 1:256</td><td> 1:80</td><td> 138</td><td> 141</td><td></td><td> 268</td><td> 130</td>

<td> 103</td><td> 1:256</td><td> 1: 200</td><td> 102</td><td> 114</td><td></td><td> 1037</td><td> 935</td>

<td> 104</td><td> 1:512</td><td> 1:80</td><td> 138</td><td> 140</td><td></td><td> 292</td><td> 154</td>

<td> 105</td><td> 1:512</td><td> 1: 200</td><td> 97</td><td> 110</td><td></td><td> 1031</td><td> 934</td>

<td> 106</td><td>Control 1</td><td></td><td> 86</td><td> 175</td><td></td><td> 754</td><td> 668</td>

<td> 107</td><td>Control 2</td><td></td><td> 87</td><td> 176</td><td></td><td> 1180</td><td> 1093</td>

As indicated in Table 7, none of the test bottles containing the low levels of each of the bioactive compositions or the Immunox dilution provide antifungal activity throughout the entire 96 hour period analyzed. Likewise, neither the 1: 128 dilution (example

<img file="MX336953B_D0148.tif" />

117

<img file="MX336953B_D0149.tif" />

99) nor does the 1:64 dilution (example 98) of Immunox provide any measure of efficacy, even in the shortest 18-hour trial period, despite the fact that the manufacturer generally recommends a 1:64 dilution. Similarly, Examples 103 and 105 that have a 1: 200 dilution of the bioactive composition (~ lppm for each metal, 0.08% citric acid, 0.00125 for NaLS, and 0.0016 for

SLS) in combination with the two Immunox dilutions cannot demonstrate bioefficacy while the combinations of both Immunox dilutions with a slightly higher level, 1:80 dilution, of the bioactive composition (~ 2.5 ppm of each metal, 0.2% of citric acid, 0.003 of NaLS and 0.004 of SLS) demonstrate bioefficacy. This demonstrates a synergy between the two compositions since the 1:80 dilution alone cannot show bioefficacy throughout the entire period analyzed.

EXAMPLES 108-126

Metal fountains

A series of experiments is carried out using different metal salts as the metal ion sources.

In this case, sufficient amounts of silver nitrate, copper sulfate, and zinc oxide are added to a solution of

<img file="MX336953B_D0150.tif" />

118

<img file="MX336953B_D0151.tif" />

5% aqueous citric acid to provide 31.75 ppm silver,

12.5 ppm of copper and 40.17 ppm of zinc. Different amounts of this stock concentrate solution (MI4) are added to the test bottles to evaluate the efficacy. The specific formulations, including the resulting ppm of each metal in the test flask, as well as the results thereof to inhibit yeast growth are as presented in Tables 8 and 8A.

TABLE 8

<td rowspan="2">Axis.</td><td rowspan="2">Volume of MI4 ag watered</td><td colspan="3">Metal concentration</td><td colspan="2">Surfactant% (w / w)</td><td colspan="7">Turbidity (NTU)</td>

<td>Ppm of Ag</td><td>Ppm of Cu</td><td>Ppm of Zn</td><td>NaLS</td><td>SDS</td><td>T zero</td><td>T 2</td><td>T 18</td><td>T 28</td><td>T 44</td><td>T 48</td><td>T 68</td>

<td> 108</td><td> 0. 5</td><td> 0.79</td><td> 0. 31</td><td> 1.00</td><td></td><td></td><td> 81</td><td> 129</td><td> 950</td><td> 1046</td><td> 1046</td><td> 1046</td><td> 1054</td>

<td> 109</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td></td><td></td><td> 85</td><td> 135</td><td> 950</td><td> 997</td><td> 1055</td><td> 990</td><td> 1023</td>

<td> 110</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td></td><td></td><td> 112</td><td> 158</td><td> 916</td><td> 930</td><td> 960</td><td> 930</td><td> 970</td>

<td> 111</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td></td><td></td><td> 126</td><td> 158</td><td> 760</td><td> 799</td><td> 810</td><td> 83 0</td><td> 844</td>

<td> 112</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td> 0.005</td><td></td><td> 140</td><td> 143</td><td> 179</td><td> 307</td><td> 919</td><td> 93 6</td><td> 980</td>

<td> 113</td><td> 1</td><td> 1. 59</td><td> 0.63</td><td> 2.01</td><td> 0.005</td><td></td><td> 140</td><td> 137</td><td> 143</td><td> 152</td><td> 279</td><td> 306</td><td> 468</td>

<td> 114</td><td> 2</td><td> 3. 18</td><td> 1.25</td><td> 4.02</td><td> 0.005</td><td></td><td> 180</td><td> 174</td><td> 174</td><td> 177</td><td> 24 4</td><td> 25 2</td><td> 28 2</td>

<td> 115</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td> 0.005</td><td></td><td> 187</td><td> 185</td><td> 184</td><td> 18 4</td><td> 18 4</td><td> 18 4</td><td> 272</td>

<td> 116</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td></td><td> 0.00 5</td><td> 83</td><td> 132</td><td> 948</td><td> 1054</td><td> 1066</td><td> 1078</td><td> 1097</td>

<td> 117</td><td> 1</td><td> 1. 59</td><td> 0.63</td><td> 2.01</td><td></td><td> 0.00 5</td><td> 97</td><td> 138</td><td> 911</td><td> 1003</td><td> 1100</td><td> 1060</td><td> 1075</td>

<td> 118</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td></td><td> 0.005</td><td> 116</td><td> 147</td><td> 746</td><td> 90 7</td><td> 970</td><td> 1001</td><td> 1006</td>

<td> 119</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td></td><td> 0.005</td><td> 124</td><td> 156</td><td> 504</td><td> 701</td><td> 840</td><td> 868</td><td> 916</td>

<td> 120</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td> 0.0025</td><td> 0.0025</td><td> 140</td><td> 140</td><td> 250</td><td> 64 0</td><td> 1065</td><td> 1088</td><td> 1133</td>

<td> 121</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td> 0.0025</td><td> 0.0025</td><td> 149</td><td> 149</td><td> 160</td><td> 256</td><td> 930</td><td> 901</td><td> 1014</td>

<td> 122</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td> 0.0025</td><td> 0.0025</td><td> 164</td><td> 177</td><td> 174</td><td> 174</td><td> 291</td><td> 459</td><td> 804</td>

<td> 123</td><td> 3</td><td> 4. 76</td><td> 1.88</td><td> 6.03</td><td> 0.0025</td><td> 0.0025</td><td> 176</td><td> 179</td><td> 177</td><td> 181</td><td> 32 0</td><td> 445</td><td> 738</td>

<td> 124</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td> 0. 01</td><td></td><td> 162</td><td> 162</td><td> 162</td><td> 163</td><td> 163</td><td> 164</td><td> 164</td>

<td> 125</td><td>0.8S</td><td> 1.37</td><td> 0.54</td><td> 1.73</td><td> 0. 01</td><td></td><td> 150</td><td> 140</td><td> 140</td><td> 14 0</td><td> 186</td><td> 20 8</td><td> 254</td>

<td> 126</td><td></td><td></td><td></td><td></td><td></td><td></td><td> 78</td><td> 113</td><td> 877</td><td> 856</td><td> 878</td><td> 865</td><td> 898</td>

119

<img file="MX336953B_D0152.tif" />

INSTITUTE mu-ü-auD £ LA PRGPÍFiV.f iNOUSTRIAi

TABLE 8A

<td rowspan="2">Axis.</td><td rowspan="2">M14 volume n added</td><td colspan="3">Metal concentration</td><td colspan="2">Surfactant% (w / w)</td><td colspan="6">Change in turbidity (delta NTU)</td>

<td>Ppm of Ag</td><td>Ppm of Cu</td><td>Ppm of Zn</td><td>NaLS</td><td>SDS</td><td>Delta T2-T0</td><td>D T18- T0</td><td>D T26- T0</td><td>D T44- TO</td><td>D T48- T0</td><td>D T68- T0</td>

<td> 108</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td></td><td></td><td> 48</td><td> 869</td><td> 965</td><td> 965</td><td> 965</td><td> 973</td>

<td> 109</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td></td><td></td><td> 51</td><td> 865</td><td> 912</td><td> 970</td><td> 905</td><td> 938</td>

<td> 110</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td></td><td></td><td> 48</td><td> 804</td><td> 818</td><td> 848</td><td> 818</td><td> 858</td>

<td> 111</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td></td><td></td><td> 32</td><td> 624</td><td> 67 3</td><td> 68 4</td><td> 704</td><td> 718</td>

<td> 112</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td> 0.005</td><td></td><td> 3</td><td> 39</td><td> 167</td><td> 779</td><td> 796</td><td> 840</td>

<td> 113</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td> 0.005</td><td></td><td> -3</td><td> 3</td><td> 12</td><td> 139</td><td> 166</td><td> 328</td>

<td> 114</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td> 0.005</td><td></td><td> -8</td><td> -6</td><td> -3</td><td> 64</td><td> 72</td><td> 102</td>

<td>lis</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td> 0.005</td><td></td><td> -2</td><td> -3</td><td> -3</td><td> -3</td><td> -3</td><td> 85</td>

<td> 116</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td></td><td> 0.005</td><td> 49</td><td> 865</td><td> 971</td><td> 98 3</td><td> 995</td><td> 1014</td>

<td> 117</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td></td><td> 0.005</td><td> 39</td><td> 814</td><td> 906</td><td> 1003</td><td> 963</td><td> 978</td>

<td> 118</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td></td><td> 0.005</td><td> 31</td><td> 630</td><td> 791</td><td> 854</td><td> 885</td><td> 890</td>

<td> 119</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td></td><td> 0.00 5</td><td> 32</td><td> 380</td><td> 577</td><td> 716</td><td> 744</td><td> 792</td>

<td> 120</td><td> 0. 5</td><td> 0,79</td><td> 0.31</td><td> 1.00</td><td> 0.0025</td><td> 0.0025</td><td> 0</td><td> 110</td><td> 500</td><td> 925</td><td> 948</td><td> 993</td>

<td> 121</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td> 0.0025</td><td> 0.0025</td><td> 0</td><td> 11</td><td> 107</td><td> 781</td><td> 752</td><td> 865</td>

<td> 122</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td> 0.0025</td><td> 0.0025</td><td> 13</td><td> 10</td><td> 10</td><td> 127</td><td> 295</td><td> 540</td>

<td> 123</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td> 0.0025</td><td> 0.0025</td><td> 3</td><td> 1</td><td> 5</td><td> 144</td><td> 269</td><td> 560</td>

<td> 124</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td> 0. 01</td><td></td><td> 0</td><td> 0</td><td> 1</td><td> 1</td><td> 2</td><td> 2</td>

<td> 125</td><td> 0.86</td><td> 1.37</td><td> 0.54</td><td> 1.73</td><td> 0. 01</td><td></td><td> -10</td><td> -10</td><td> -10</td><td> 36</td><td> 58</td><td> 104</td>

<td> 126</td><td></td><td></td><td></td><td></td><td></td><td></td><td> 35</td><td> 799</td><td> 78 8</td><td> 800</td><td> 787</td><td> 820</td>

The results shown in Tables 8 and 8A demonstrate that the selection of the metal ion source is not critical as long as it is readily soluble and soluble to the degree necessary to provide the desired level of metal ion concentration in the solution.

Furthermore, the results demonstrate bioefficacy even at extremely low metal and acid contents. Although, the

120

<img file="MX336953B_D0153.tif" />

Efficacy is of relatively short duration at the lowest concentrations; long-term bioefficacy is found with only minor adjustments in the relative concentration of the necessary components. Also, depending on the final end-use application, such short-term antifungal efficacy may be sufficient; therefore, allowing any environmental contamination from the general application of these materials to be minimized.

The results also suggest that sodium lauryl sulfate may be ineffective on its own in promoting the bioefficacy of the bioactive compositions of the present invention. However, its presence may be desirable in cases where the effective surfactant is not readily soluble in the aqueous system. On the other hand, their presence or the presence of similar surfactants may not be important in cases where attempts are made to produce non-aqueous systems. For example, systems that are applied as an emulsion in water or as an oil that can be spread in an aqueous medium to which these are applied, for example, a paddy, might need surfactants that are less hydrophilic and more lipophilic.

i ?.

12.1 institute me? :: CANO OF INDUSTRIAL PROPERTY

<img file="MX336953B_D0154.tif" />

EXAMPLES 127-143

Lactic acid

A series of experiments similar to the above are carried out except that citric acid is replaced by lactic acid. Therefore, the bioactive composition (MI5) comprises sufficient amounts of silver nitrate, copper sulfate, and zinc oxide dissolved in a 5% aqueous lactic acid solution to provide 31.75 ppm silver, 12.5 ppm copper, and 40.17 ppm zinc. The specific formulations analyzed and the results obtained with them are as presented in Tables 9 and 9A.

TABLE 9

<td rowspan="2">Example</td><td rowspan="2">Volume of MI5 ag watered</td><td colspan="3">Concentration of metals</td><td colspan="2">Surfactant (w / w)%</td><td colspan="5">Turbidity (NTU)</td>

<td>Ppm of Ag</td><td>Ppm of CU</td><td>Ppm of Zn</td><td>NaLS</td><td>SDS</td><td>T zero</td><td>YOU</td><td>IT 8</td><td>T24</td><td>T44</td>

<td> 127</td><td> 0.5</td><td> 0.79</td><td> 0.31</td><td> 1. 00</td><td></td><td></td><td> 107</td><td> 130</td><td> 1000</td><td>lili</td><td> 1001</td>

<td> 128</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2. 01</td><td></td><td></td><td> 109</td><td> 130</td><td> 1006</td><td> 1021</td><td> 1016</td>

<td> 129</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td></td><td></td><td> 14 8</td><td> 154</td><td> 970</td><td> 995</td><td> 1014</td>

<td> 130</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6. 03</td><td></td><td></td><td> 178</td><td> 202</td><td> 914</td><td> 925</td><td> 990</td>

<td> 131</td><td> 0.5</td><td> 0.79</td><td> 0.31</td><td> 1. 00</td><td> 0.005</td><td></td><td> 134</td><td> 170</td><td> 30 0</td><td> 454</td><td> 923</td>

<td> 132</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2. 01</td><td> 0.005</td><td></td><td> 153</td><td> 169</td><td> 200</td><td> 227</td><td> 292</td>

<td> 133</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4. 02</td><td> 0.00 5</td><td></td><td> 218</td><td> 217</td><td> 20 7</td><td> 204</td><td> 228</td>

<td> 134</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6. 03</td><td> 0.005</td><td></td><td> 222</td><td> 22 3</td><td> 222</td><td> 215</td><td> 227</td>

<td> 135</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1. 00</td><td></td><td> 0. 005</td><td> 120</td><td> 14 5</td><td> 1074</td><td>lili</td><td> 1079</td>

<td> 136</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2. 01</td><td></td><td> 0. 005</td><td> 14 0</td><td> 156</td><td> 1050</td><td> 1092</td><td> 1110</td>

<td> 137</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td></td><td> 0. 005</td><td> 179</td><td> 19 3</td><td> 94 5</td><td> 1031</td><td> 1080</td>

<img file="MX336953B_D0155.tif" />

<img file="MX336953B_D0156.tif" />

<img file="MX336953B_D0157.tif" />

122

TABLE 9 (cont.)

<td rowspan="2">Example</td><td rowspan="2">Volume from MI5 ag watered</td><td colspan="3">Concentration of you tell me</td><td colspan="2">Surfactant% (w / w)</td><td colspan="5">Turbidity (NTU)</td>

<td>Ppm of Ag</td><td>Ppm of CU</td><td>Ppm of Zn</td><td>NaLS</td><td>SDS</td><td>T zero</td><td>YOU</td><td>T18</td><td>T24</td><td>T44</td>

<td> 138</td><td> 3</td><td> 4. 76</td><td> 1.88</td><td> 6. 03</td><td></td><td> 0.005</td><td> 223</td><td> 239</td><td> 690</td><td> 977</td><td> 1180</td>

<td> 139</td><td> 0.5</td><td> 0.79</td><td> 0. 31</td><td> 1. 00</td><td> 0.0025</td><td> 0.0025</td><td> 143</td><td> 151</td><td> 884</td><td> 968</td><td> 1170</td>

<td> 140</td><td> 1</td><td> 1. 59</td><td> 0.63</td><td> 2. 01</td><td> 0.0025</td><td> 0.0025</td><td> 175</td><td> 175</td><td> 237</td><td> 330</td><td> 1110</td>

<td> 141</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td> 0.0025</td><td> 0.0025</td><td> 210</td><td> 214</td><td> 207</td><td> 223</td><td> 730</td>

<td> 142</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6. 03</td><td> 0.0025</td><td> 0.0025</td><td> 24 0</td><td> 240</td><td> 228</td><td> 228</td><td> 475</td>

<td> 143</td><td>Control</td><td></td><td></td><td></td><td></td><td></td><td> 100</td><td> 139</td><td> 1175</td><td> 1163</td><td> 1170</td>

TABLE 9A

<td rowspan="2">Axis.</td><td rowspan="2">Volume n of MI5 ag watered</td><td colspan="3">Metal concentration</td><td colspan="2">Ag surfactant% (w / w)</td><td colspan="3">Change in turbidity (delta NTU)</td><td>(NTU)</td>

<td>Ppm of Ag</td><td>Ppm of CU</td><td>Ppm of Zn</td><td>NaLS</td><td>SDS</td><td>D T1-T0</td><td>D T18-T10</td><td>DT24-T0</td><td>DT44-T0</td>

<td> 127</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td></td><td></td><td> 23</td><td> 893</td><td> 1004</td><td> 894</td>

<td> 123</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td></td><td></td><td> 21</td><td> 897</td><td> 912</td><td> 907</td>

<td> 129</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td></td><td></td><td> 8</td><td> 822</td><td> 847</td><td> 866</td>

<td> 133</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td></td><td></td><td> 24</td><td> 736</td><td> 747</td><td> 812</td>

<td> 131</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td> 0.005</td><td></td><td> 36</td><td> 166</td><td> 320</td><td> 789</td>

<td> 132</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td> 0.005</td><td></td><td> 16</td><td> 47</td><td> 74</td><td> 139</td>

<td> 133</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td> 0.005</td><td></td><td> -1</td><td> -11</td><td> -14</td><td> 10</td>

<td> 134</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td> 0.005</td><td></td><td> 1</td><td> 0</td><td> *7</td><td> 5</td>

<td> 135</td><td> 0. 5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td></td><td> 0.005</td><td> 25</td><td> 954</td><td> 991</td><td> 959</td>

<td> 136</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td></td><td> 0.005</td><td> 16</td><td> 910</td><td> 952</td><td> 970</td>

<td> 137</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td></td><td> 0.00 5</td><td> 14</td><td> 766</td><td> 852</td><td> 901</td>

<td> 138</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td></td><td> 0.005</td><td> 16</td><td> 467</td><td> 754</td><td> 957</td>

<td> 139</td><td> 0.5</td><td> 0.79</td><td> 0.31</td><td> 1.00</td><td> 0.0025</td><td> 0.0025</td><td> 8</td><td> 741</td><td> 825</td><td> 1027</td>

<td> 140</td><td> 1</td><td> 1.59</td><td> 0.63</td><td> 2.01</td><td> 0.0025</td><td> 0.0025</td><td> 0</td><td> 62</td><td> 155</td><td> 935</td>

<td> 141</td><td> 2</td><td> 3.18</td><td> 1.25</td><td> 4.02</td><td> 0.0025</td><td> 0.0025</td><td> 4</td><td> -3</td><td> 13</td><td> 520</td>

<td> 142</td><td> 3</td><td> 4.76</td><td> 1.88</td><td> 6.03</td><td> 0.0025</td><td> 0.0025</td><td> 0</td><td> -12</td><td> -12</td><td> 23 5</td>

<td> 143</td><td>Control</td><td></td><td></td><td></td><td></td><td></td><td> 39</td><td> 1075</td><td> 1063</td><td> 1070</td>

<img file="MX336953B_D0158.tif" />

123

<img file="MX336953B_D0159.tif" />

The

9A, mimic experiments extrapolate to results such as those shown in those found in the set indicating that the invention acids of similar characteristics.

tables 9 and prior to you can

EXAMPLES 144-156

Phosphoric acid

Two are prepared

<td>evaluation in</td><td>the</td><td>which</td>

<td>phosphoric. In</td><td>the</td><td>first,</td>

<td colspan="2">copper citrate and</td><td>citrate &lt;</td>

<td>phosphoric acid</td><td>to the</td><td>16% for</td>

<td colspan="2">prepare a second</td><td>solution</td>

<td>silver sulfate</td><td>of</td><td>copper and</td>

Stock solutions for the acid used is acid. Silver citrate, zinc are added to an aqueous solution to provide 200 ppm of each metal. Reserve using zinc oxide nitrate, again in the 16% phosphoric acid solution to provide 200 ppm of each metal. Both compositions also contain 0.32% surfactant, either as an individual surfactant or as a 50:50 mixture. Specific formulations and results of their efficacy in controlling yeast growth are as presented in Tables and 10A.

TABLE 10

<td rowspan="8">P Hey s Ñ 0) T3 -you £ you you</td><td>T 96</td><td> 1106</td><td> 154</td><td> 169</td><td> 1162</td><td> 168</td><td>σ-ι "you</td><td>CO co "you</td><td> 185</td><td> 188</td><td> 206</td><td>'T co "you</td><td> 215</td><td> 988</td>

<td>T 72</td><td> 1146</td><td> 162</td><td> 172</td><td> 778</td><td> 170</td><td>σ » ΟΊ «—1</td><td> 192</td><td> 197</td><td> 201</td><td>"you CÑ</td><td> 208</td><td> 232</td><td>LL8</td>

<td>CO Hey</td><td> 1094</td><td> 163</td><td> 177</td><td> 174</td><td> 174</td><td> 204</td><td> 203</td><td> 205</td><td> 208</td><td> 229</td><td> 213</td><td> 238</td><td> 913</td>

<td>CÑ AND-</td><td>CD = T OR I — 1</td><td> 160</td><td> 172</td><td> 172</td><td> 17 5</td><td> 196</td><td>co σ » «—I</td><td> 200</td><td>«T or CÑ</td><td> 223</td><td> 216</td><td> 232</td><td> 938</td>

<td>T 24</td><td> 400</td><td> 166</td><td> 178</td><td> 179</td><td> 180</td><td> 197</td><td> 200</td><td> 216</td><td> 200</td><td> 229</td><td> 222</td><td>r- xT CÑ</td><td>ΟΊ CTi the</td>

<td>T 18</td><td> 300</td><td> 166</td><td>CO r— "you</td><td>ΟΊ Γ- "you</td><td> 18 0</td><td> 197</td><td> 200</td><td> 216</td><td> 200</td><td> 229</td><td> 226</td><td> 250</td><td>Γ- ΟΟ 'T</td>

<td>T 1 hour</td><td>vT CO 5-)</td><td> 180</td><td> 193</td><td>CO co "you</td><td>CO Ch "you</td><td> 219</td><td>00 "you Cñ</td><td>TT m CÑ</td><td> 237</td><td> 263</td><td>co Γ- CÑ</td><td> 272</td><td>00 the</td>

<td>T zero</td><td> 123</td><td>σι Ch i — 1</td><td> 211</td><td> 168</td><td> 209</td><td> 228</td><td>CO CÑ CÑ</td><td> 258</td><td> 253</td><td> 285</td><td> 280</td><td> 283</td><td>CÑ the</td>

<td colspan="2">&amp; &amp; OR &gt; •you 4J OR &lt;0 0 (0 C 0) 4J Q) 4-1 you 0) σ &lt;</td><td></td><td></td><td></td><td></td><td></td><td></td><td>SLS 0.05</td><td>NaLS 0. 05</td><td>0.05 SLS / 0.05 NaLS</td><td>SLS 0.05</td><td>NaLS 0.05</td><td>0.05 SLS / 0.05 NaLS</td><td></td>

<td colspan="2">«Η 5 a</td><td>THE CÑ</td><td>the</td><td>or «~ I</td><td>m CÑ</td><td>to go"</td><td>OR "you</td><td>the</td><td>io</td><td>io</td><td>to go"</td><td>the</td><td>the</td><td>THE</td>

<td colspan="2">Metal fountain</td><td>Citrate salts *</td><td>Citrate salts *</td><td>Citrate salts *</td><td>OR you CÑ 'T OR w you or co i</td><td>OR you tS3 'T OR w you υ co one</td><td>or you tS) 'T OR ω you υ co one</td><td>Citrate salts *</td><td>Citrate salts *</td><td>Citrate salts *</td><td>OR you M OR 3 υ co one</td><td>OR you Cñ 'T OR CO 3 υ co one</td><td>OR you N OR 3 OR co one</td><td>Control</td>

<td colspan="2">Example</td><td>&lt;= r "you</td><td> 145</td><td> 14 6</td><td> 147</td><td> 148</td><td> 14 9</td><td> 150</td><td> 151</td><td> 152</td><td> 153</td><td>THE "you</td><td> 155</td><td> 15 6</td>

THE

You (0 β

σ α&gt;

&gt;

• Η you (0 ο

g (Ο

Ti c0

Or you

N

Q)

Ό

OR

4-J ¢ 0

Li

4-&gt;

•uncle

&gt; 1 you

OR

Φ

You

4-&gt;

with you

4-&gt;

Ή o

&lt;

Ό o

4-1 (0 you

4-1 • you u

<img file="MX336953B_D0160.tif" />

LO «—I

TABLE 10A

<img file="MX336953B_D0161.tif" />

THE

IMPI

MEXICAN INSTITUTE OF INDUSTRIAL PROPERTY

<img file="MX336953B_D0162.tif" />

I

126

<img file="MX336953B_D0163.tif" />

Results such as those shown in Tables 10 and 10A suggest that the surfactant may not be critical in those compositions in which the excess acid is a strong to moderate acid, such as phosphoric acid.

EXAMPLES 157-166

Nitric acid

<td>For</td><td>further demonstrate</td><td>the</td><td>scope</td><td>of the</td>

<td>compositions</td><td>bioactive, it is used</td><td>a</td><td>acid</td><td>mineral</td>

<td>relatively</td><td>strong, nitric acid,</td><td>how</td><td colspan="2">the component</td>

acid. A stock solution is prepared by combining 78.7 mg of silver nitrate, 62.2 mg of zinc oxide and 200 mg of copper sulfate with 20 ml of purified water and 1.5 g of concentrated nitric acid (68%) under constant stirring. Once the solids dissolve, additional purified water is added to obtain a volume of 250. As prepared, this mixture contains approximately 200 ppm of each metal, as calculated. The pH is measured and found to be 1.66. The mixture is then divided into three aliquots of approximately equal volume. One aliquot is set aside and the other two are subjected to pH adjustment with ammonium hydroxide. The amount of ammonium hydroxide added is the

127

JL x _r. v,

INSTITUTO MEXIC / Λ'Ο i ** —ζ · * 7 '- &lt;β CE LA PROPIOD / D Úr

INDUSIRL'.L necessary to bring the pH of the first aliquot up to 2.55 and of the second aliquot up to 3.63.

Each solution is then evaluated, with and without surfactants, to assess its bioefficacy in inhibiting yeast growth. The amount of each of the three aliquots added to the 20 ml bottle of yeast suspension is indicated in Table 11 along with the amount of surfactant added, where indicated. The surfactant used is a 50:50 mixture of sodium lauryl sulfate and sodium lauroylsarcosinate. The specific formulations analyzed and the results thereof are presented in Table 11. As can be seen from Table 11, the combination of metal and acid does not provide any inhibition at the levels analyzed. However, when the surfactant is added, bioefficacy manifests itself even at the lowest metal / acid concentration.

TABLE 11

Nitric acid

<td rowspan="2">Axis.</td><td rowspan="2">Vol. Of MI 6 aggregate</td><td rowspan="2">Metals (ppm)</td><td rowspan="2">Surfactant% (w / w)</td><td rowspan="2">PH</td><td colspan="5">Turbidity / Change in Turbidity</td>

<td>T0</td><td>T18</td><td>Τ18-Ϊ0</td><td>T42</td><td>T42-T0</td>

<td> 157</td><td> 0. 5</td><td> 5</td><td></td><td> 1. 66</td><td> 69</td><td> 1243</td><td> 1174</td><td> 1133</td><td> 1064</td>

<td> 158</td><td> 0. 5</td><td> 5</td><td></td><td> 2.55</td><td> 67</td><td> 1245</td><td> 1178</td><td> 1133</td><td> 1066</td>

<td> 159</td><td> 0. 5</td><td> 5</td><td></td><td> 3. 63</td><td> 69</td><td> 1243</td><td> 1174</td><td> 1150</td><td> 1081</td>

<img file="MX336953B_D0164.tif" />

<img file="MX336953B_D0165.tif" />

128

TABLE 11 (cont.)

<td rowspan="2">Axis.</td><td rowspan="2">Vol. Of MI 6 aggregate</td><td rowspan="2">Metals (ppm)</td><td rowspan="2">Surfactant% (w / w)</td><td rowspan="2">pH</td><td colspan="3">Turbidity / Change in</td><td colspan="2">Turbidity</td>

<td>T0</td><td>T18</td><td>T18-T0</td><td>T42</td><td>T42-T0</td>

<td> 160</td><td> • 1</td><td> 10</td><td></td><td> 1.66</td><td> 65</td><td> 976</td><td> 911</td><td> 1162</td><td> 1097</td>

<td> 161</td><td> 1</td><td> 10</td><td></td><td> 2.55</td><td> 66</td><td> 1012</td><td> 946</td><td> 1186</td><td> 1120</td>

<td> 162</td><td> 1</td><td> 10</td><td></td><td> 3.63</td><td> 67</td><td> 1036</td><td> 969</td><td> 1166</td><td> 1099</td>

<td> 163</td><td> 0. 5</td><td> 5</td><td> 0. 05</td><td> 1.66</td><td> 61</td><td> 55</td><td> -6</td><td> 58</td><td> -3</td>

<td> 164</td><td> 0. 5</td><td> 5</td><td> 0. 05</td><td> 2.55</td><td> 62</td><td> 53</td><td> -9</td><td> 55</td><td> -7</td>

<td> 165</td><td> 0. 5</td><td> 5</td><td> 0. 05</td><td> 3.63</td><td> 60</td><td> 57</td><td> -3</td><td> 52</td><td> -8</td>

<td> 166</td><td> 0</td><td></td><td></td><td></td><td> 67</td><td> 1255</td><td> 1188</td><td> 1212</td><td> 1145</td>

EXAMPLES 167-222

Surfactant evaluation

A series of experiments is conducted to select various surfactants for efficacy in accordance with the present invention. Surfactants are evaluated as a pure additive (0 ppm metal) or in combination with either 1 ml or 2 ml of a 4% citric acid solution containing 50 ppm each of copper, silver and zinc. With the addition of 1 ml of the citric acid solution, the yeast suspension test bottle contains approximately 0.2% citric acid and approximately 2.5 ppm of each metal. With the addition of 2 ml of the citric acid solution, the acid is approximately 0.4% and the metals are each present

129

<img file="MX336953B_D0166.tif" />

at about 5 ppm in test bottles. Each surfactant is evaluated at a concentration of approximately

0.05% by weight. Controls are also evaluated with and without metals.

The specific surfactants evaluated as well as the formulations of each test composition along with the results thereof are indicated in Table 12.

As seen in Table 12, the benefits of the present invention are obtained with a wide range of surfactant materials. Especially preferred surfactants are those that are free of or substantially free of ethylene oxide repeating units and / or have moderate to low molecular weights. Notwithstanding the foregoing, good results are reported to be obtained with the product Pluronic L62, a polyethylene oxide containing surfactant, when used in combination with the lowest level of acid and metals. The higher acid level is believed to have affected the stability of this material, and possibly similar materials.

TABLE 12

<img file="MX336953B_D0167.tif" />

THE

LQ τ — I

OR

TABLE 12 (Cont)

<td>= T CM Jj σ. 3</td><td>CN Ν 'do υ η Ν 'Γ * · Cl</td><td> 803 932 960</td><td>co&gt; σ- P— CN Γ ~ \ £&gt; CO CO</td><td> 588 1221 8</td><td> 1119</td><td>&gt; Ν ' σι</td><td> 1101</td><td> 688</td><td>i — 1 Ν ' OR Rh</td>

<td> 734 1034 1060</td><td> 651 922 939</td><td> 742 988 1007</td><td> 1030 775 936</td><td> ¡ 726 ¡ 1002 ³</td><td> 1110</td><td> 1086</td><td> 1096 !</td><td> 1035</td><td> 1076 ^:</td>

<td> 1078 1078 1085</td><td> 966 962 911</td><td> 977 1038 1010</td><td> 1061 1024 975</td><td> 850 558 3</td><td> 1094</td><td> 979</td><td> 1053</td><td> 987</td><td> 1034</td>

<td>O IA LT&gt; * 00 CN</td><td> 2 65 2 58</td><td> 957 112 368</td><td> 1020 840 911</td><td> 290 308 5</td><td> 1113</td><td>Ν ' r- οο</td><td>T CO or ι-1</td><td> 806</td><td> 1008</td>

<td>„CO Ν 'LO 3 O CN 5-1 5-1</td><td> 541 876 1128</td><td> 809 1192 1319</td><td> 730 719 104 8</td><td> 64 2 1323 237</td><td> 1177</td><td> 1041</td><td> 1233</td><td> 962</td><td> : 1193</td>

<td>s SP</td><td> 700 1014 1128</td><td> 948 1248 1366</td><td> 1087 867 1105</td><td> 782 1104 232</td><td> 1168</td><td> 1180</td><td> 1228</td><td> 1128</td><td> 1228</td>

<td> 1128 1164 1300</td><td> 1015 1054 1100</td><td> 1183 1296 138 9</td><td> 1118 1116 1144</td><td> 906 680 232</td><td> 1152</td><td> 1073</td><td> 1185</td><td> 1080</td><td>CO CO Rh Rh</td>

<td>O CN O Ν 'Cl 00 CTi rH Ν '</td><td>Ν ' m cn</td><td> 1163 372 725</td><td> 1077 932 1080</td><td>co O m Ν 'rH ΓΟ CO Ν' CN</td><td> 1171</td><td> 896</td><td> 1196</td><td> 1001</td><td>1160 l !</td>

<td> 50 106 215</td><td>σι cu e- σ,</td><td> 206 260 359</td><td> 57 92 169</td><td> 56 102 229</td><td>CO THE</td><td>Ν '</td><td> 132</td><td>CO ΟΊ</td><td> 152</td>

<td> 0 2.5 5</td><td> 0 2.5 5</td><td> 0 2.5 5</td><td> 0 2.5 5</td><td> 0 2.5 5</td><td>or</td><td>or</td><td>OR</td><td>the CN</td><td>a</td>

<td>Non-ionic</td><td>Non-ionic</td><td>Non-ionic</td><td>Non-ionic</td><td>non-ionic</td><td></td><td></td><td></td><td></td><td></td>

<td>Dow Chemical</td><td>Dow Chemical</td><td>Hunisman Chemical</td><td></td><td>Cognis</td><td></td><td></td><td></td><td></td><td></td>

<td>Ethoxylate alcohol secondary rami ficado</td><td>Ethoxylate alcohol secondary rami ficado</td><td>Alcohol ethoxylated from C1-C14</td><td>polyoxyethylene (20) sorbitan monolaurate</td><td>Rent polyglucoside</td><td></td><td></td><td></td><td></td><td></td>

<td>Tergitol TMN6</td><td>Tergitol TMN3</td><td>Sulfonic TDA3B</td><td>Tween 20</td><td>Pose 2000</td><td>Control</td><td>Control (2.5 ppm)</td><td>Control (5 ppm)</td><td>Metals Control</td><td>Metals Control</td>

IΜ PI

MEXICAN INSTITUTE OF INDUSTRIAL PROPERTY

<img file="MX336953B_D0168.tif" />

THE

OR

THE

132

<img file="MX336953B_D0169.tif" />

EXAMPLES 223 -236

Comparison with strobilurin

A series of experiments is carried out in order to evaluate the comparative performance of the bioactive compositions of the present invention and various commercial strobilurin-based fungicides. Two bioactive formulations are used. The first, MI2, comprises a 16% aqueous citric acid solution that has silver citrate, copper citrate, and zinc citrate dissolved in it, each added in an amount to provide 200 ppm of each metal, along with 0.25 % of sodium lauroylsarcosinate and 0.32% of sodium lauryl sulfate, as indicated above. The second, MI7, comprises a 160: 1 dilution of a 16% aqueous phosphoric acid solution that has dissolved in the same silver citrate, copper citrate, and zinc citrate, each added in an amount to provide 200 ppm of each metal in the phosphoric acid solution. Each fungicide is evaluated at different levels.

The specific formulations analyzed and the results obtained with them are presented in Tables 13 and

13A.

I Μ ΡI institute ^^ ..- τ == ίΓ? \

133

TABLE 13

<td rowspan="2">Example</td><td rowspan="2">Fungicide</td><td rowspan="2">Vol. Aggregate</td><td colspan="5">Turbidity (NTU)</td>

<td>T0</td><td>YOU</td><td>T 18</td><td>T 26</td><td>T 50</td>

<td> 223</td><td>Quadris³</td><td> 1</td><td> 384</td><td> 393</td><td> 1066</td><td> 1139</td><td> 1134</td>

<td> 224</td><td></td><td> 2</td><td> 767</td><td> 772</td><td> 1264</td><td> 1311</td><td> 1315</td>

<td> 225</td><td></td><td> 5</td><td> 1332</td><td> 1332</td><td> 1364</td><td> 1377</td><td> 1376</td>

<td> 226</td><td>Flint¹³</td><td> 1</td><td> 418</td><td> 424</td><td> 1115</td><td> 1208</td><td> 1234</td>

<td> 227</td><td></td><td> 2</td><td> 718</td><td> 708</td><td> 1141</td><td> 1299</td><td> 1327</td>

<td> 228</td><td></td><td> 5</td><td> 1210</td><td> 1210</td><td> 1270</td><td> 1265</td><td> 1245</td>

<td> 229</td><td>Headline^c</td><td> 1</td><td> 232</td><td> 225</td><td> 961</td><td> 1114</td><td> 1137</td>

<td> 230</td><td></td><td> 2</td><td> 387</td><td> 391</td><td> 1066</td><td> 1134</td><td> 1199</td>

<td> 231</td><td></td><td> 5</td><td> 717</td><td> 747</td><td> 1178</td><td> 1222</td><td> 1241</td>

<td> 232</td><td>MY 2</td><td> 0.5</td><td> 128</td><td> 129</td><td> 154</td><td> 177</td><td> 174</td>

<td> 233</td><td>M12</td><td> 1</td><td> 414</td><td> 384</td><td> 366</td><td> 366</td><td> 352</td>

<td> 234</td><td>MI 7</td><td> 0.5</td><td> 249</td><td> 244</td><td> 248</td><td> 248</td><td> 242</td>

<td> 235</td><td>MI 7</td><td> 1</td><td> 311</td><td> 302</td><td> 283</td><td> 283</td><td> 277</td>

<td> 236</td><td>Control</td><td></td><td> 67</td><td> 68</td><td> 793</td><td> 871</td><td> 904</td>

a - Quadris fungicide from Syngenta Crop Prctections, Inc. of Greensboro, NC, USA.

b - Flint fungicide from Bayer CropScience LP of Research Triangle Park, NC, USA. c ~ Headline of BASF Corporation of Research Triangle Park, NC, USA.

TABLE 13A

<td rowspan="2">Example</td><td rowspan="2">Fungicide</td><td rowspan="2">Vol. Aggregate</td><td colspan="3">Change in turbidity (delta NTU)</td>

<td>T18-T1</td><td>T26-T1</td><td>T50-T1</td>

<td> 223</td><td>Quadris^to</td><td> 1</td><td> 673</td><td> 746</td><td> 741</td>

<td> 224</td><td></td><td> 2</td><td> 492</td><td> 539</td><td> 543</td>

<td> 225</td><td></td><td> 5</td><td> 32</td><td> 45</td><td> 44</td>

<td> 226</td><td>Flint¹³</td><td> 1</td><td> 691</td><td> 784</td><td> 810</td>

<td> 227</td><td></td><td> 2</td><td> 433</td><td> 591</td><td> 619</td>

<td> 228</td><td></td><td> 5</td><td> 60</td><td> 55</td><td> 35</td>

<td> 229</td><td>Headline^c</td><td> 1</td><td> 736</td><td> 889</td><td> 912</td>

<td> 230</td><td></td><td> 2</td><td> 675</td><td> 743</td><td> 808</td>

<td> 231</td><td></td><td> 5</td><td> 431</td><td> 475</td><td> 494</td>

<td> 232</td><td>MY 2</td><td> 0.5</td><td> 25</td><td> 48</td><td> 45</td>

<td> 233</td><td>MY 2</td><td> 1</td><td> -18</td><td> -18</td><td> -32</td>

<td> 234</td><td>M17</td><td> 0.5</td><td> 4</td><td> 4</td><td> -2</td>

<td> 235</td><td>MI 7</td><td> 1</td><td> -19</td><td> -19</td><td> -25</td>

<td> 236</td><td>Control</td><td></td><td> 725</td><td> 803</td><td> 836</td>

a - Quadris fungicide from Syngenta Crop Protections, Inc. of Greensboro, NC, USA. b - Flint fungicide from Bayer CropScience LP of Research Triangle Park, NC, USA.

c- Headline of BASF Corporation of Research Triangle Park, NC, USA.

<img file="MX336953B_D0170.tif" />

L&gt; t INDUSTRIAL PROPERTY

<img file="MX336953B_D0171.tif" />

As seen in Tables 13 and 13A, the bioactive compositions of the present invention provide marked inhibition of yeast growth, even at the lowest concentrations, ~ 5 ppm of each metal ion. On the other hand, all but two of the tested strobilurin-based fungicide formulations were unable to demonstrate any significant bioefficacy against yeast over the time period analyzed. The two formulations that provide good inhibition are at comparatively high loads.

EXAMPLES 237-250

Strobilurin synergy

In view of the poor prior performance of strobilurins in general, a series of experiments is conducted in order to assess the potential synergy between the bioactive compositions of the present invention and the prior commercial strobilurin-based fungicides. The compositions used are the same as those used in the previous set of examples. The specific formulations analyzed and the results obtained with them are presented in Tables 14 and 14A.

<img file="MX336953B_D0172.tif" />

135

<img file="MX336953B_D0173.tif" />

TABLE 14

<td rowspan="2">Axis.</td><td rowspan="2">Bioactive</td><td rowspan="2">Vol. Aggregate</td><td rowspan="2">Fungicide*</td><td rowspan="2">Vol. Aggregate</td><td colspan="5">Turbidity (NTU)</td>

<td>T0</td><td>YOU</td><td>T18</td><td>T24</td><td>T96</td>

<td> 237</td><td>MY 2</td><td> 0.25</td><td>Q</td><td> 1</td><td> 552</td><td> 554</td><td> 544</td><td> 670</td><td> 1315</td>

<td> 238</td><td>MY 2</td><td> 0.25</td><td>Q</td><td> 2</td><td> 896</td><td> 894</td><td> 868</td><td> 891</td><td> 1470</td>

<td> 239</td><td>MY 2</td><td> 0. 5</td><td>Q</td><td> 1</td><td> 588</td><td> 578</td><td> 564</td><td> 5 64</td><td> 608</td>

<td> 240</td><td>MY 2</td><td> 0.25</td><td>F</td><td> 1</td><td> 578</td><td> 599</td><td> 568</td><td> 568</td><td> 1320</td>

<td> 241</td><td>MY 2</td><td> 0.25</td><td>F</td><td> 2</td><td> 900</td><td> 900</td><td> 886</td><td> 836</td><td> 1330</td>

<td> 242</td><td>MY 2</td><td> 0.25</td><td>H</td><td> 1</td><td> 436</td><td> 433</td><td> 454</td><td> 454</td><td> 1312</td>

<td> 243</td><td>MY 2</td><td> 0.25</td><td>H</td><td> 2</td><td> 611</td><td> 637</td><td> 667</td><td> 632</td><td> 1302</td>

<td> 244</td><td>MI 7</td><td> 0.25</td><td>Q</td><td> 1</td><td> 558</td><td> 574</td><td> 640</td><td> 668</td><td> 1273</td>

<td> 245</td><td>MI 7</td><td> 0.25</td><td>F</td><td> 1</td><td> 517</td><td> 560</td><td> 990</td><td> 1197</td><td> 1396</td>

<td> 246</td><td>MI 7</td><td> 0.25</td><td>H</td><td> 1</td><td> 465</td><td> 476</td><td> 605</td><td> 587</td><td> 12 90</td>

<td> 247</td><td>Control</td><td></td><td> -</td><td></td><td> 93</td><td> 101</td><td> 901</td><td> 938</td><td> 1075</td>

<td> 248</td><td>MY 2</td><td> 0. 5</td><td></td><td></td><td> 499</td><td> 440</td><td> 390</td><td> 390</td><td> 373</td>

<td> 249</td><td>MY 2</td><td> 0.25</td><td></td><td></td><td> 182</td><td> 179</td><td> 175</td><td> 176</td><td> 1122</td>

<td> 250</td><td>MY 2</td><td> 0. 5</td><td></td><td></td><td> 262</td><td> 260</td><td> 260</td><td> 275</td><td> 275</td>

a) Q - fungicide. Quadris from Syngenta Crop Protections, Inc. of Greensboro, NC, USA.

F - Flint fungicide from Bayer CropScience LP from Research Triangle Park, NC, USA.

H- Headline of BASF Corporation of Research Triangle Park, NC, USA.

TABLE 14A

<td rowspan="2">Axis.</td><td rowspan="2">Bioactive</td><td rowspan="2">Vol. Aggregate</td><td rowspan="2">Fungicide*</td><td rowspan="2">Vol. Aggregate</td><td colspan="3">Change in turbidity (delta NTU)</td>

<td>T18-T1</td><td>T24-T1</td><td>T96-T1</td>

<td> 237</td><td>MY 2</td><td> 0.25</td><td>Q</td><td> 1</td><td> -10</td><td> 116</td><td> 761</td>

<td> 238</td><td>MY 2</td><td> 0.25</td><td>Q</td><td> 2</td><td> -26</td><td> -3</td><td> 576</td>

<td> 239</td><td>MY 2</td><td> 0.5</td><td>Q</td><td> 1</td><td> -14</td><td> -14</td><td> 30</td>

<td> 240</td><td>MY 2</td><td> 0.25</td><td>F</td><td> 1</td><td> -31</td><td> -31</td><td> 721</td>

<td> 241</td><td>MY 2</td><td> 0.25</td><td>F</td><td> 2</td><td> -14</td><td> -14</td><td> 430</td>

<td> 242</td><td>MY 2</td><td> 0.25</td><td>H</td><td> 1</td><td> 21</td><td> 21</td><td> 879</td>

136

TABLE 14A (cont.)

IM F

INSTITUTO MEXIC DE LA FROÍ-'E INDüST

YEAR

RL

<img file="MX336953B_D0174.tif" />

<td rowspan="2">Axis.</td><td rowspan="2">Bioactive</td><td rowspan="2">Vol. Aggregate</td><td rowspan="2">Fungicide*</td><td rowspan="2">Vol. Aggregate</td><td colspan="3">Change in turbidity (delta NTU)</td>

<td>T18-T1</td><td>T24-T1</td><td>T96-T1</td>

<td> 243</td><td>MY 2</td><td> 0.25</td><td>H</td><td> 2</td><td> 30</td><td> -5</td><td> 665</td>

<td> 244</td><td>MI 7</td><td> 0.25</td><td>Q</td><td> 1</td><td> 66</td><td> 94</td><td> 699</td>

<td> 245</td><td>MI 7</td><td> 0.25</td><td>F</td><td> 1</td><td> 430</td><td> 637</td><td> 836</td>

<td> 246</td><td>MI 7</td><td> 0.25</td><td>H</td><td> 1</td><td> 129</td><td> 111</td><td> 814</td>

<td> 247</td><td>Control</td><td></td><td> -</td><td></td><td> 800</td><td> 885</td><td> 97 4</td>

<td> 248</td><td>MY 2</td><td> 0.5</td><td></td><td></td><td> -50</td><td> -50</td><td> -67</td>

<td> 249</td><td>MY 2</td><td> 0.25</td><td></td><td></td><td> -4</td><td> -3</td><td> 943</td>

<td> 250</td><td>MY 2</td><td> 0.5</td><td></td><td></td><td> 0</td><td> 15</td><td> 15</td>

to)

Q - Quadris fungicide from Syngenta Crop Protections, Inc. of Greensboro F - Flint fungicide from Bayer CropScience LP of Research Triangle Park,

NC, USA NC, USA

H- BASF Headline

As a combination of invention with synergy in products

Research Triangle Park Corporation,

NC, USA shows in Tables 14 the bioactive compositions of the strobilurin products which even the highest levels and 14A, the present one produces a low of the analyzed strobilurins, they produce a significant inhibition in the growth of yeast, even when these products appear to increase yeast growth when used alone, as shown in Tables 13 and

13A.

137 ^p i 7 ///

MEXICAN INSTITUTE OF INDUSTRIAL PROPERTY

EXAMPLES 251-259

Copper / zinc study

A series of experiments is performed to demonstrate the bioefficacy of binary metal systems compared to the ternary system used in most other examples. In this case, a MI2 solution is compared with a similar composition that contains 300 ppm copper and 300 ppm zinc (that is, an aqueous solution of citric acid at

16% having dissolved in the same copper citrate and zinc citrate, each added in an amount to provide 300 ppm of each metal, along with 0.25% sodium lauroylsarcosinate and 0.32% sodium lauryl sulfate). The two bioactive compositions are evaluated at different loads to establish their bioefficacy. The specific formulations analyzed and the results obtained with them are presented in Tables 15 and 15A.

As seen in Tables 15 and 15A, both binary (copper / zinc-Cu / Zn) and ternary piate / copper / zinc MI2 antimicrobial compositions demonstrate comparable bioefficacy in inhibiting yeast growth.

<img file="MX336953B_D0175.tif" />

138

TABLE 15

<td rowspan="2">Example</td><td colspan="2">Composition (gm)</td><td colspan="5"></td>

<td>Cu / Zn</td><td>MY 2</td><td>T0</td><td>YOU</td><td>T18</td><td>T24</td><td>T46</td>

<td> 251</td><td> 1</td><td></td><td> 776</td><td> 586</td><td> 468</td><td> 463</td><td> 436</td>

<td> 252</td><td> 0.5</td><td></td><td> 292</td><td> 269</td><td> 250</td><td> 250</td><td> 245</td>

<td> 253</td><td> 0.2</td><td></td><td> 147</td><td> 162</td><td> 772</td><td> 1055</td><td> 1075</td>

<td> 254</td><td> 0.1</td><td></td><td> 93</td><td> 125</td><td> 1076</td><td> 1070</td><td> 1036</td>

<td> 255</td><td>Control</td><td></td><td> 66</td><td> 127</td><td> 1020</td><td> 1012</td><td> 1137</td>

<td> 256</td><td></td><td> 1</td><td> 830</td><td> 633</td><td> 547</td><td> 522</td><td> 500</td>

<td> 257</td><td></td><td> 0.5</td><td> 335</td><td> 320</td><td> 292</td><td> 302</td><td> 284</td>

<td> 258</td><td></td><td> 0.2</td><td> 152</td><td> 178</td><td> 512</td><td> 1064</td><td> 1098</td>

<td> 259</td><td></td><td> 0.1</td><td> 90</td><td> 136</td><td> 1083</td><td> 1087</td><td> 1067</td>

TABLE 15A

<td rowspan="2">Example</td><td colspan="2">Composition (gm)</td><td colspan="4"></td>

<td>Cu / Zn</td><td>MY 2</td><td>T1-T0</td><td>T18-T0</td><td>T24-T0</td><td>T46-T0</td>

<td> 251</td><td> 1</td><td></td><td> -190</td><td> -118</td><td> -5</td><td> -27</td>

<td> 252</td><td> 0.5</td><td></td><td> -23</td><td> -19</td><td> 0</td><td> -5</td>

<td> 253</td><td> 0.2</td><td></td><td> 15</td><td> 610</td><td> 283</td><td> 20</td>

<td> 254</td><td> 0.1</td><td></td><td> 32</td><td> 951</td><td> -6</td><td> -34</td>

<td> 255</td><td>Control</td><td></td><td> 61</td><td> 893</td><td> -8</td><td> 125</td>

<td> 256</td><td></td><td> 1</td><td> -197</td><td> -86</td><td> -25</td><td> -22</td>

<td> 257</td><td></td><td> 0.5</td><td> -15</td><td> -2 8</td><td> 10</td><td> -18</td>

<td> 258</td><td></td><td> 0.2</td><td> 26</td><td> 334</td><td> 552</td><td> 34</td>

<td> 259</td><td></td><td> 0.1</td><td> 46</td><td> 947</td><td> 4</td><td> -20</td>

EXAMPLES 260-269

Synergy with Mancozeb

<td>I know</td><td>make a</td><td>additional series</td><td>of</td><td>experiments</td><td>for</td>

<td>evaluate the</td><td>bioefficacy,</td><td>especially</td><td>the</td><td>synergy of</td><td>the</td>

<td>composition</td><td>agrochemical</td><td colspan="3">bioactive containing Mancozeb</td><td>(a</td>

<img file="MX336953B_D0176.tif" />

ethylene bisdithiocarbamate) and the bioactive acid solution MI2 (MI2). The specific formulations analyzed and the results obtained with them are presented in Tables 16 and 16A.

TABLE 16

<td rowspan="2">Example</td><td colspan="2">Composition (gm)</td><td rowspan="2">T0</td><td rowspan="2">T2</td><td rowspan="2">T18</td><td rowspan="2">T24</td><td rowspan="2">T44</td>

<td>Mancozeb</td><td>MY 2</td>

<td> 260</td><td> 0.5</td><td></td><td> 934</td><td> 976</td><td> 1220</td><td> 1095</td><td> 1091</td>

<td> 261</td><td> 0 . 4</td><td></td><td> 780</td><td> 859</td><td> 1021</td><td> 982</td><td> 1052</td>

<td> 2 62</td><td> 0.3</td><td></td><td> 624</td><td> 717</td><td> 12 0 9</td><td> 1067</td><td> 1113</td>

<td> 2 63</td><td> 0.2</td><td></td><td> 392</td><td> 489</td><td> 1035</td><td> 933</td><td> 1073</td>

<td> 264</td><td></td><td> 0.2</td><td> 57</td><td> 55</td><td> 54</td><td> 72</td><td> 756</td>

<td> 265</td><td> 0.5</td><td> 0.2</td><td> 930</td><td> 897</td><td> 864</td><td> 839</td><td> 788</td>

<td> 2 66</td><td> 0.4</td><td> 0.2</td><td> 727</td><td> 709</td><td> 684</td><td> 664</td><td> 591</td>

<td> 267</td><td> 0.3</td><td> 0.2</td><td> 537</td><td> 555</td><td> 535</td><td> 509</td><td> 460</td>

<td> 268 269</td><td>0.2 Control</td><td> 0.2</td><td> 370 23</td><td> 369 106</td><td> 370 935</td><td> 343 824</td><td> 331 917</td>

TABLE 16A

<td>Example</td><td colspan="2">Composition (gm)</td><td></td><td></td><td></td><td></td>

<td></td><td>Mancozeb</td><td>MY 2</td><td>T2-T0</td><td>T18-T0</td><td>T24-T0</td><td>T44-T0</td>

<td> 260</td><td> 0.5</td><td></td><td> 42</td><td> 286</td><td> 161</td><td> 157</td>

<td> 261</td><td> 0.4</td><td></td><td> 79</td><td> 241</td><td> 202</td><td> 272</td>

<td> 2 62</td><td> 0.3</td><td></td><td> 93</td><td> 585</td><td> 443</td><td> 489</td>

<td> 263</td><td> 0.2</td><td></td><td> 97</td><td> 643</td><td> 541</td><td> 681</td>

<td> 264</td><td></td><td> 0.2</td><td> -2</td><td> -3</td><td> 15</td><td> 699</td>

<td> 265</td><td> 0.5</td><td> 0.2</td><td> -33</td><td> -66</td><td> -91</td><td> -142</td>

<td> 266</td><td> 0.4</td><td> 0.2</td><td> -18</td><td> -43</td><td> -63</td><td> -136</td>

<td> 267</td><td> 0.3</td><td> 0.2</td><td> 18</td><td> -2</td><td> -28</td><td> -77</td>

<td> 268</td><td> 0.2</td><td> 0.2</td><td> -1</td><td> 0</td><td> -27</td><td> -39</td>

<td> 269</td><td>Control</td><td></td><td> 83</td><td> 912</td><td> 801</td><td> 894</td>

140

<img file="MX336953B_D0177.tif" />

<td></td><td>How I know</td><td>observe in</td><td>the</td><td>Tables 16 and</td><td>16A, the mancozeb</td>

<td>if</td><td>same is</td><td>ineffective z</td><td>to</td><td>the levels</td><td>analyzed. The</td>

<td>solution</td><td>acid</td><td>bioactive</td><td>by</td><td colspan="2">itself provides bioefficacy</td>

<td>modest,</td><td>despite</td><td colspan="3">from the very low level of</td><td>metal ions</td>

antimicrobial; however, the appropriate bioefficacy seems to have disappeared after 44 hours. In sharp contrast, the combination of the two, at all levels of mancozeb, demonstrates excellent bioefficacy, even after 44 hours.

EXAMPLES 270-285

Antibacterial study

A series of experiments are performed to evaluate the performance of the individual components of the claimed bioactive compositions as well as various combinations thereof, including, the claimed compositions themselves, to suppress the growth of various bacteria. Escherichía coli (E. coli},

Pseudomonas aerugínosa (P. aeruginosa) and Staphylococcus aureus (S. aureus) as test organisms because they are generally accepted in the industry as indicator organisms for a wide variety of bacteria.

Two different methodologies are evaluated, one that analyzes the effectiveness in a growth broth medium and the other that

141

<img file="MX336953B_D0178.tif" />

analyzes inhibition in inoculated growth media.

EXAMPLES 270-281

In the first set of experiments, a growth medium is prepared by adding 10 grams of nutrient medium (Sabouraud Difeo dextrose broth from Franklin BD

Lakes, NJ, USA) to 300 ml of distilled water. The 20 ml aliquots of the growth medium are dispensed in sterile 40 ml borosilicate glass bottles with Teflon lined lids (VWR International Cat. No. 15900-004). The vials are inoculated with the bacteria using a sterile microbiological loop and the vials are then incubated at

37 ° C. After a bioactive composition according to the invention is added to certain bottles, the bioactive composition is (MI2), as described above, which comprises a 16% aqueous citric acid solution having dissolved in the same silver citrate, citrate copper and zinc citrate, each added in an amount to provide 200 ppm of each metal, along with

0.25% sodium lauroylsarcosinate and 0.32% sodium lauryl sulfate. The turbidity of each mixture is then determined and the bottle is transferred to an incubator at 30 ° C. The turbidity measurements are carried out as in the

<img file="MX336953B_D0179.tif" />

n ¿^ rjTUTO MEXICANO s NDDíTRíAL PROPERTY

<img file="MX336953B_D0180.tif" />

2 yeast mentioned above. Each bottle is periodically removed from the incubator and the mixture in the bottles is analyzed for turbidity. The specific formulations analyzed, the time for each turbidity evaluation, and the results obtained with them are as indicated in Table 17.

As with the yeast study, the concentration of metals refers to the approximate amount of each metal, copper, silver, and zinc. The concentrations do not compensate with respect to the volume of MI2 added: therefore, the concentrations presented are based on a total volume of 20 ml.

TABLE 17

<td rowspan="2">Example</td><td rowspan="2">Bacterium</td><td rowspan="2">MY 2 (me)</td><td rowspan="2">Metals ppm</td><td colspan="5">Time (hours)</td>

<td>T0</td><td>T0.5</td><td>T18</td><td>T24</td><td>T96</td>

<td> 294</td><td>E col i</td><td> 0</td><td> 0</td><td> 15.3</td><td> 16</td><td> 119</td><td> 136</td><td> 264</td>

<td> 2 95</td><td></td><td> 0.5</td><td> 5</td><td> 131</td><td> 135.3</td><td> 165</td><td> 162</td><td> 162</td>

<td> 296</td><td></td><td> 1</td><td> 10</td><td> 445</td><td> 454</td><td> 481</td><td> 480</td><td> 480</td>

<td> 297</td><td></td><td> 2</td><td> 20</td><td> 1039</td><td> 1080</td><td> 1135</td><td> 1140</td><td> 1009</td>

<td> 298</td><td>P. aerugínosa</td><td> 0</td><td> 0</td><td> 35.8</td><td> 37.8</td><td> 158</td><td> 383</td><td> 436</td>

<td> 299</td><td></td><td> 0.5</td><td> 5</td><td> 197</td><td> 207</td><td> 250</td><td> 2 62</td><td> 261</td>

<td> 300</td><td></td><td> 1</td><td> 10</td><td> 705</td><td> 735</td><td> 782</td><td> 808</td><td> 807</td>

<td> 301</td><td></td><td> 2</td><td> 20</td><td> 1011</td><td> 1057</td><td> 1121</td><td> 1159</td><td> 1146</td>

<td> 302</td><td>S. aureus</td><td> 0</td><td> 0</td><td> 46</td><td> 45</td><td> 148</td><td> 184</td><td> 406</td>

<td> 303</td><td></td><td> 0.5</td><td> 5</td><td> 215</td><td> 163</td><td> 173</td><td> 183</td><td> 184</td>

<td> 304</td><td></td><td> 1</td><td> 10</td><td> 643</td><td> 494</td><td> 326</td><td> 309</td><td> 276</td>

<td> 305</td><td></td><td> 2</td><td> 20</td><td> 1203</td><td> 1032</td><td> 595</td><td> 525</td><td> 281</td>

143

<img file="MX336953B_D0181.tif" />

<td></td><td>How</td><td>I know</td><td>watch</td><td>in table T7 “</td><td>exists</td><td>a</td>

<td colspan="3">short increment</td><td>term in</td><td>turbidity. Due</td><td>why not</td><td>I know</td>

<td>anticipates</td><td>than</td><td>I know</td><td>beech</td><td>manifested no</td><td colspan="2">increase</td>

Significant in such a short period of time, the initial increase in turbidity is believed to be the result of protein denaturation in the broth and / or bacterial proteins. Regardless, the longer term results show excellent bacterial inhibition with the compositions according to the present invention.

EXAMPLE 282

In this experiment, six sterile 25mm coverslips are placed in separate sterile Petri dishes of

10C x 15 mm and two of each are inoculated with 100 μΐ of one of three TSB broths: each broth contains one of E. coli, P.

aeruginosa and S. aureus that have been incubated for 48-54 hours. In order to attach the inoculum to the coverslips, the Petri dishes are placed on a low temperature heating rack for approximately 5 minutes. One of each of the inoculated Petri dishes is set aside as positive controls. The other is sprayed with 4 sprays of a 4: 1 dilution of the MI2 bioactive compositions. After 2-3 minutes the coverslips and the? I

144

INSTITUTE '/ r · D £ L \ F

INDUS

<img file="MX336953B_D0182.tif" />

Liquid contents of each Petri dish are aseptically transferred into separate vials containing my TSB and incubated at 37 ° C for 24 hours. Negative controls are prepared by placing non-inoculated sterile coverslips within 20 ml of TSB and are also incubated. After 24 hours, no growth is observed with the negative controls or with those inoculated coverslips that were sprayed with the bioactive composition of the present invention. Visible growth is observed in two of the positive controls (i.e. those vials containing the inoculated coverslips that were not

<td>sprinkled):</td><td>positive control</td><td>for</td><td>P.</td><td>aeruginous</td><td>not</td>

<td colspan="2">presents visible growth. I know</td><td>believe</td><td>than</td><td>the lack</td><td>of</td>

<td>increase</td><td colspan="2">visible in the latter is</td><td>the</td><td>Outcome</td><td>of</td>

overheat the inoculum during the fixation step.

EXAMPLE 283

In this experiment, two soy-trypticase agar (TSA) plates are inoculated with 500 μΐ of one of three TSB broths for a total of 6 inoculated plates: each broth contains one of E. coli, P. aeruginosa and S. aureus which were incubated for 48-54 hours. The inoculum is spread evenly across the surface of the plate with a sterile loop. I know

<img file="MX336953B_D0183.tif" />

<img file="MX336953B_D0184.tif" />

145

<img file="MX336953B_D0185.tif" />

Place a 15mm diameter disc of filter paper previously dipped in a 4: 1 dilution of the MI2 bioactive composition in the center of one of each set of inoculated plates and all plates are placed in an incubator at

37 ° C for 24 hours. The uninoculated control plates are also placed in the incubator.

After 24 hours, visible growth is observed. Bacterial growth is not observed in the uninoculated plates. Growth is observed in all inoculated plates; however, on the plates on which the treated filter paper was placed, no growth is observed on or near the filter paper. Each treated filter paper disc exhibits a clear zone of bacterial growth inhibition.

EXAMPLE 284

In this experiment, two soy-trypticase agar (TSA) plates are inoculated with 500 μΐ of one of three TSB broths for a total of 6 inoculated plates: each broth contains one of E. coli, P. aerugínosa and S. aureus which were incubated for 48-54 hours. The inoculum is spread evenly across the surface of the plate with a sterile loop. Then one of each of the inoculated plates is sprayed,

<img file="MX336953B_D0186.tif" />

MEXICAN PROPERTY INSTITUTE

INDUSTRIAL

<img file="MX336953B_D0187.tif" />

6 . __ approximately 24 times, with the 4: 1 dilution of the bioactive composition MI2. The inoculated plates plus a set of uninoculated control plate plates are placed in an incubator at 37 ° C for 24 hours.

After 24 hours, visible growth is observed in the inoculated but untreated plates, whereas no bacterial growth is observed in the uninoculated plates or in the inoculated plates that were sprayed with the diluted bioactive composition.

EXAMPLE 285

Bacterial minimum inhibitory concentration (MIC) study

A study is carried out to determine the minimum inhibitory concentration (MIC) of the acid solution

MI2, that is, 200 ppm of each of the silver, copper and zinc metals (see examples 72-79). Three different bacteria are evaluated, Clavíbacter michiganese, Pseudomonas syríngae and Erwínía amylovora, each in a different growth medium appropriate for said bacterium, in specific agar / brain infusion broth, agar / nutrient broth, and nutrient agar / broth with glucose, respectively. To perform the test, three sets of 10 test tubes are prepared, one set for each

147

INSTITUTE V * '

DE U FPf i »'J

In ¡x &lt;j. ¿Ik «Aí.

<img file="MX336953B_D0188.tif" />

bacteria, and labeled 1 through 10. 0.5 ml of the appropriate broth is placed in each of test tubes 2 through 10.

Then 0.5 ml of the MI2 solution is added to each of test tubes 1 and 2. Then 0.5 ml of the contents of test tube 2 are transferred to test tube 3 and then 0.5 ml of test tube 3 to the tube Test 4 and so on until reaching test tube 9. 0.5 ml is discarded for test tube 9. Then 0.5 ml of suspension of each bacterium to be analyzed is added to each of the 10 tubes for said series and the tubes they are incubated for 24 hours at 26 ° C.

Because the acid solution causes considerable clouding of the tubes to which it is added, macroscopic evaluation is not possible. Instead, each tube is subcultured on corresponding agar plates. The observed growth is as indicated in Table 18 (a + indicates visible growth and a no growth).

TABLE 18

<td>Test tube</td><td> 1</td><td> 2</td><td> 3</td><td> 4</td><td> 5</td><td> 6</td><td> 7</td><td> 8</td><td> 9</td><td> 10</td>

<td>Concentration of metals * (ppm)</td><td> 200</td><td> 50</td><td> 25</td><td> 12.5</td><td> 6.75</td><td> 3.125</td><td> 1.56</td><td> 0.782</td><td> 0.391</td><td> 0.195</td>

<td>C. michíganese</td><td> —</td><td></td><td></td><td> -</td><td> —</td><td></td><td> +</td><td> +</td><td> +</td><td> +</td>

<td>P. syxíngae</td><td> —</td><td></td><td></td><td> -</td><td> —</td><td> +</td><td> +</td><td> +</td><td> +</td><td> +</td>

<td>E. amylovora</td><td> —</td><td> —</td><td></td><td> -</td><td> —</td><td> —</td><td> +</td><td> +</td><td> +</td><td> +</td>

* - concentration of each metal, the total metal content is 3 times the number presented

<img file="MX336953B_D0189.tif" />

MEXICAN PROPERTY INSTITUTE

INDUSTRIAL

<img file="MX336953B_D0190.tif" />

¹⁴⁸ __ - Based on the results presented in Table 18, the MIC of MI2 is 3,125 ppm for C. michíganese and for E. amylovora and 6.75 ppm for P. syringae. The bioefficacy of such low levels is anticipated to show synergy when combined with conventional fungicides / bactericides for such target organisms.

EXAMPLE 286

Alternary leaf spot

To demonstrate the efficacy of bioactive compositions on live plants, a comparative study comparing the efficacy of a bioactive composition in accordance with the present invention with two commercial products, Eagle 40WP, a fungicide based on miclobutanil (40% by weight), is conducted. available from Dow AgroSciences LLC of Indianapolis, IN, USA, and Scala SC, a pyrimethanil-based fungicide (54.6% by weight) available from Bayer

CropScience LP of Research Triangle Park, NC, USA Additional evaluations are performed to analyze the potential for synergy between the bioactive compositions of the invention and the Eagle 40WP product. The bioactive composition according to the present invention comprises a 16% aqueous citric acid solution having dissolved

149

<img file="MX336953B_D0191.tif" />

in the same silver citrate, copper citrate, and zinc citrate in an amount that provides 200 ppm of each metal in the solution, 0.25% of sodium lauroylsarcosinate and 0.32% of sodium lauryl sulfate (MI6). This solution is diluted at 40: 1 and 20: 1 rates for application to plants providing a solution containing ~ 5 ppm and ~ 10 ppm of each metal as sprayed.

Rooted cuttings of Pittosporum tobíra Wheeleri are sown in standard 10.16-era pots containing Sunshine Mix No. 1 and fertilized with 1/2 teaspoon Osmocote Plus 15-9-12. Plants are placed in a greenhouse heated with poly and shading cloth that cover the top and sides and are irrigated by overflow as needed. After 44 days, the plants are treated with the various antifungal treatments - 12 plants are treated with each treatment.

After this, the plants are placed in individual clear plastic bags (high humidity) in the greenhouse for the duration of the study. Plants are watered from below using a reflux and overflow bench to ensure that no water is applied to their leaves during the test. The plants are subsequently inoculated by spraying with a spore suspension from an Alternaría pittospori culture mixed with

150 ,«=\

MEXICAN INSTITUTE OF PROPERTY

INDUSTRY!

sterilized water 4 days after the initial treatment. Treatments are reapplied 7 days and 17 days after inoculation. All treatments are applied by spraying until the surfaces of the plant leaves are completely wet (they begin to drain).

Two sets of plants are used as positive and negative controls: the first set is treated with water only (Treatment A) and is not inoculated. The second set is also treated with water only, but is also inoculated concurrently with the others. The specific formulations for each of the treatments are as indicated in Table 19.

TABLE 19

<td>Treatment</td><td>Composition</td><td>Dilution</td>

<td>TO</td><td>Water - not inoculated</td><td></td>

<td>B</td><td>Water - inoculated</td><td></td>

<td>C</td><td>MI 6</td><td>6.25 ml / 250 mi water</td>

<td>D</td><td>MI 6</td><td>12.5 ml / 250 my water</td>

<td>AND</td><td>MI6 / Eagle 40WP</td><td>6.25 ml / 250 ml water / 0.11 g / 1 water</td>

<td>F</td><td>MI6 / Eagle 40WP</td><td>6.25 ml / 250 ml water / 0.22 g / 1 water</td>

<td>G</td><td>MI6 / Eagle 40WP</td><td>12.5 ml / 250 ml water / 0.11 g / 1 water</td>

<td>H</td><td>Eagle 40WP</td><td>0.11 * g / 1 of water</td>

<td>I</td><td>Eagle 40WP</td><td>0.22 g / 1 water</td>

<td>J</td><td>Scala</td><td>0.67 * g / 1 of 'water</td>

* 'manufacturer recommended application rates

151

<img file="MX336953B_D0192.tif" />

Six days after the second treatment, the plants are evaluated for leaf spot by Alternaria by visual inspection. The results of the leaf spot evaluation are as presented in Table 20. As seen in Table 20, plants treated with the lowest concentration of the bioactive composition (with ~ 5 ppm of each metal ion - Treatment C ) continue to show a nearly 50% drop in leaf spot formation.

Double the bioactive composition (~ 10 ppm of each metal ion - Treatment D) reduces leaf spot by about 75%. Somewhat similar results are found with the two dilutions of commercial fungicide.

Eagle 40WP in which the lowest concentration (Treatment H) reduces leaf spot by approximately

30% while the highest concentration (Treatment I) reduces leaf spot by 80%. Combining the two marked improvements provided with, oddly enough, the combination of the two lower concentrations that provide almost complete inhibition of leaf spot manifestation. The other commercial fungicide Scala SC does not provide inhibition and appears to promote the manifestation of leaf spot.

<img file="MX336953B_D0193.tif" />

<img file="MX336953B_D0194.tif" />

152

TABLE 20

<td rowspan="2">Treatment</td><td colspan="5">Nc</td><td>. of</td><td colspan="6">plant</td><td rowspan="2">Half</td>

<td> 1</td><td> 2</td><td> 3</td><td> 4</td><td> 5</td><td> 6</td><td> 7</td><td> 8</td><td> 9</td><td> 10</td><td> 11</td><td> 12</td>

<td>TO</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0.0</td>

<td>B</td><td> 4</td><td> 5</td><td> 0</td><td> 15</td><td> 35</td><td> 20</td><td> 40</td><td> 15</td><td> 10</td><td> 25</td><td> 30</td><td> 20</td><td> 18.2</td>

<td>C</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 5</td><td> 35</td><td> 35</td><td> 40</td><td> 0</td><td> 0</td><td> 0</td><td> 9.6</td>

<td>D</td><td> 0</td><td> 1</td><td> 0</td><td> 0</td><td> 5</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 5</td><td> 10</td><td> 30</td><td> 4.2</td>

<td>AND</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 1</td><td> 0</td><td> 0</td><td> 5</td><td> 0</td><td> 0</td><td> 0</td><td> 0.5</td>

<td>F</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 25</td><td> 0</td><td> 0</td><td> 5</td><td> 10</td><td> 0</td><td> 3.3</td>

<td>G</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 15</td><td> 10</td><td> 0</td><td> 0</td><td> 0</td><td> 2.1</td>

<td>H</td><td> 0</td><td>or 1</td><td> 0</td><td> 5</td><td> 10</td><td> 0</td><td> 30</td><td> 35</td><td> 40</td><td> 10</td><td> 10</td><td> 15</td><td> 12.9</td>

<td>I</td><td> 2</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 0</td><td> 10</td><td> 25</td><td> 5</td><td> 0</td><td> 0</td><td> 0</td><td> 3.5</td>

<td>J</td><td> 25</td><td> 25</td><td> 10</td><td> 5</td><td> 15</td><td> 25</td><td> 30</td><td> 0</td><td> 40</td><td> 40</td><td> 40</td><td> 20</td><td> 22.9</td>

Eleven days after the last treatment, the severity of the disease is again evaluated. However, due to the number of spots that make it impossible to give a numerical evaluation, the severity of the disease is recorded using the following scale: 1 - 1 - no disease, 2 - mild, 3 - moderate, 4 - severe to 5 -dead .

The results are presented in Table 21.

TABLE 21

<td rowspan="2">Treatment</td><td colspan="12">Plant No.</td><td rowspan="2">Half</td>

<td> 1</td><td> 2</td><td> 3</td><td> 4</td><td> 5</td><td> 6</td><td> 7</td><td> 8</td><td> 9</td><td> 10</td><td> 11</td><td> 12</td>

<td>TO</td><td> 2</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td>1.1a</td>

<td>B</td><td> 2.5</td><td> 2 .5</td><td> 1</td><td> 4</td><td> 3.5</td><td> 3</td><td> 4</td><td> 2</td><td> 2</td><td> 3</td><td> 3</td><td> 3.5</td><td>2.8c</td>

<td>C</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 2</td><td> 2</td><td> 2.5</td><td> 2.5</td><td> 2</td><td> 2</td><td> 1</td><td>1.6a</td>

<td>D</td><td> 1</td><td> 1</td><td> 2</td><td> 1</td><td> 2</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 2</td><td> 2.5</td><td>1.4a</td>

<td>AND</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 2</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 2</td><td>1.2a</td>

153

<img file="MX336953B_D0195.tif" />

TABLE 21 (cont.)

<td rowspan="2">Treatment</td><td colspan="12">Plant No.</td><td rowspan="2">Half</td>

<td> 1</td><td> 2</td><td> 3</td><td> 4</td><td> 5</td><td> 6</td><td> 7</td><td> 8</td><td> 9</td><td> 10</td><td> 11</td><td> 12</td>

<td>F</td><td> 1</td><td> 2</td><td> 2</td><td> 1</td><td> 1</td><td> 1</td><td> 2</td><td> 1</td><td> 1</td><td> 1</td><td> 2</td><td> 1</td><td>1.3a</td>

<td>G</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 1</td><td> 2</td><td> 2</td><td> 2</td><td> 1</td><td> 1</td><td> 1</td><td>1.2a</td>

<td>H</td><td> 2</td><td> 2</td><td> 1</td><td> 2.5</td><td> 2</td><td> 2</td><td> 2.5</td><td> 3</td><td> 3</td><td> 2</td><td> 2.5</td><td> 2.5</td><td>2.2b</td>

<td>I</td><td> 2.5</td><td> 1</td><td> 1</td><td> 1</td><td> 2</td><td> 2</td><td> 2</td><td> 2</td><td> 1</td><td> 2</td><td> 2</td><td> 2</td><td>1.7a</td>

<td>J</td><td> 3.5</td><td> 4</td><td> 3</td><td> 3</td><td> 3</td><td> 4</td><td> 3.5</td><td> 2.5</td><td> 4</td><td> 4</td><td> 4</td><td> 4</td><td>3.5d</td>

As shown in Table 21, the bioactive compositions according to the present invention provide excellent protection against leaf spot, in which plants treated at the highest level and in combination with the commercial fungicide Eagle 40WP show almost the same level of disease than those that were not inoculated. In contrast, the Eagle product alone, even at the recommended application rate, was shown to be less effective than the bioactive composition. Finally, the Scala product again failed to show any efficacy, and in fact, proved to be more damaging. Plants treated with the Scala product are suspected of exhibiting both leaf spot disease and phytotoxicity. None of the plants treated with the bioactive composition or the commercial Eagle fungicide show evidence of phytotoxicity.

<img file="MX336953B_D0196.tif" />

1'

154

<img file="MX336953B_D0197.tif" />

EXAMPLE 287

Fire Blight on Wild Apple Trees

A study of the bioefficacy of the various agrochemical compositions, including bioactive acid solutions and mixtures thereof with an antibiotic agrochemical, is evaluated to assess their bioefficacy against fire blight in wild apple trees. The study is carried out on 5-year-old Snow Drift wild apple trees, in which each of the compositions is applied to ten trees in two sublots of five trees, at four times.

<td>of</td><td>flowering -day 1,</td><td>a</td><td>second</td>

<td>in</td><td>on day 4, a third in</td><td>the</td><td>day 11 and</td>

<td>in</td><td>day 19. The trees</td><td>I know</td><td>inoculate</td>

<td>to</td><td>a concentration of 4 x</td><td> 10⁶</td><td>cells</td>

by me on day 1, followed by drying the treatment, repeating the inoculation on day 13. The evaluation for fire blight is completed in 100 flowers for each of the batches on day 12, day 19 and day 27. Additionally, on day 63 the incidence of outbreak infection and the length of cancers are evaluated. The results are shown in Table 22.

Test compositions are prepared by forming solutions of the bioactive agents. Two are evaluated

155

IMPI ¢ 31

MEXICAN INSTITUTE OF PROPERTY

INDU5T ILLAL

<img file="MX336953B_D0198.tif" />

Different dilutions of the bioactive MI2, the first uses ml / 1 of water (5 ppm of each metal) and the second uses 50 ml / 1 of water (10 ppm of each metal). A streptomycin product (17% concentrate) is diluted at a rate of 58.72 g / 1 (200 ppm). Finally, copper hydroxide is diluted at a rate of 119.8 g / 1. Each of the compositions and mixtures are sprayed at a rate of 467.6 liters per hectare. The specific tests and the results obtained with them are shown in Table 22. The results are presented as the average of the counts.

TABLE 22

<td>Bioactive</td><td>Rate (per hectare)</td><td colspan="3">Incidence on flowers</td><td>Incidence in buds</td><td>Size of the cancer (cm)</td>

<td>Day</td><td></td><td> 12</td><td> 19</td><td> 27</td><td> 63</td><td> 63</td>

<td>M12</td><td>11.67 1 / ha</td><td> 0.2</td><td> 5.5</td><td> 16.5</td><td> 8.8</td><td> 40.0</td>

<td>MY 2</td><td>23.34 1 / ha</td><td> 1.7</td><td> 2.2</td><td> 6.0</td><td> 4.8</td><td> 19.8</td>

<td>Copper hydroxide^b</td><td>5.61 kg / ha</td><td> 3.1</td><td> 2.2</td><td> 9.0</td><td> 1.0</td><td> 0.0</td>

<td>Streptomycin^c</td><td>0.55 kg / ha</td><td> 1.8</td><td> 4.8</td><td> 3.09</td><td> 1.2</td><td> 0.0</td>

<td>M12 + Streptoiriicina</td><td>11.67 / ha 0.55 kg / ha</td><td> 1.3</td><td> 1.7</td><td> 4.3</td><td> 1.0</td><td> 26.0</td>

<td>Control (untreated)</td><td></td><td> 7.0</td><td> 8.5</td><td> 31.7</td><td> 7.6</td><td> 22.4</td>

a- 16% w / w aqueous citric acid solution that has dissolved in the same silver citrate, copper citrate and zinc citrate, each added in an amount to provide 200 ppm metal decade, along with 0.25% lauroylsarcosinate sodium and 0.32% sodium lauryl sulfate.

b- concentrated copper hydroxide 53% w / w c- streptomycin 17% w / w

As seen in Table 22, all tested compositions show fewer infected flowers in

<img file="MX336953B_D0199.tif" />

156

<img file="MX336953B_D0200.tif" />

comparison with control. The combination of the MI2 acid solution with streptomycin performs better than any of the bioactive alone. A significant rate response is observed between the two MI2 compositions:

the 10 ppm solution performs significantly better than the 5 ppm solution. The incidence of outbreak blight is greatly reduced for the combination of copper hydroxide, streptomycin, and streptomycin / MI2 and a modest improvement with the higher MI2 solution.

Only streptomycin and copper hydroxide appear to prevent cancer, although measurable cancers are seen with all compositions containing bioactive acid solution. All in all, these results show a marked benefit from bioactive acid solutions alone or in combination with streptomycin. More importantly, it should be noted that even at the highest concentration of the MI2 composition, the 10 ppm of each metal, a total of 30 ppm of metals, pale in comparison to the more than 50,000 ppm of copper in the solution. copper hydroxide. In essence, each application of the copper hydroxide releases almost 1000 g of copper into the environment compared to less than 2 grams or any metal released by the higher concentration MI2 composition.

Although the present invention has been described with

IMPI

<img file="MX336953B_D0201.tif" />

157 With respect to the above specific embodiments and examples, it should be appreciated that other embodiments utilizing the concept of the present invention are possible without departing from the scope of the invention. The present invention is defined by the claimed elements and any and all modifications, variations, or equivalents that fall within the scope and field of the underlying principles.

<img file="MX336953B_D0202.tif" />

<img file="MX336953B_D0203.tif" />

<img file="MX336953B_D0204.tif" />

158