WO2018037094A1

WO2018037094A1 - Fenazaquin and indoxacarb in a product for killing insects, especially mosquitoes

Info

Publication number: WO2018037094A1
Application number: PCT/EP2017/071377
Authority: WO
Inventors: Mikkel Vestergaard Frandsen; David Kim
Original assignee: Vestergaard Sa
Priority date: 2016-08-24
Filing date: 2017-08-24
Publication date: 2018-03-01

Abstract

Fenazaquin and Indoxacarbin a product and method against insects, especially mosquitoes for killing insects, especially adult mosquitoes, a substrate, for example fabric,is provided with Fenazaquin and Indoxacarb.

Description

FENAZAQUIN AND INDOXACARB IN A PRODUCT FOR KILLING INSECTS, ESPECIALLY

MOSQUITOES

FIELD OF THE INVENTION

The present invention relates to the use of Fenazaquin and Indoxacarb against insects, especially adult mosquitoes.

BACKGROUND OF THE INVENTION

In the fight against malaria, there is a steady trend towards new pesticide combi- nations, including the aim of fighting pesticidal resistance among mosquitoes. Accordingly, there is an on-going search for combinations of pesticidal agents that exhibit a synergistic effect, meaning that the combination of the active ingredients has a higher efficacy than the mere sum of the efficacies of the active ingredients. For example, if one of the ingredients has no effect, and the other ingredient has a moderate efficacy, a synergistic effect is proven for the combination of the pesticidal agents if the efficacy is higher than this moderate efficacy when the pesticidal agents are combined. Such synergistic effect has the advantage of reducing the necessary amounts and concentrations of the insecticides, which is a general desire, and it can yield a good measure to kill insects despite insecticidal re- sistance against specific insecticides or even classes of insecticides, as well as cross-resistance.

When using substrates, for example mosquito nets, the insecticides are either coated onto the net, typically called impregnation, or melt-blended into the poly- mer prior to extrusion into fibers, typically called incorporation, such that it is distributed inside and throughout the polymer material. The polymer material acts as storage for the incorporated insecticide, and the insecticide migrates from this storage to the surface of the substrate, for example the fibers. The migration is dependent on various factors, including the solubility of the insecticide in the pol- ymer, and is potentially regulated by migration promotors or inhibitors in order to achieve a long lasting efficacy. For regulation of migration of pesticidal agents in polymers, various methods have been proposed, although, there is not always a general agreement with respect to the effect of these means. For example, ethylene vinyl acetate (EVA) has been proposed in WO2003/063587 for promoting migration, whereas US6979455 explains that EVA retards migration. With respect to clay in the role of a migration controlling agent, kaolin has been proposed for increasing migration in WO2003/063587, whereas US3859121 proposes clay for retarding migration. For other migration controlling agents, similar discrepancies are found in the art. Thus, there is apparently no clear agreement in the art with respect to migration control.

International patent application WO2009/121580 assigned to Bayer Cropscience discloses a polypropylene or polyethylene polymer composition, for example for mosquito nets, which contains an organophosphate, pyrethroid, neonicotinoid or carbamate and an additive selected among sebacic esters, fatty acids, fatty acid esters, vegetable oils, esters of vegetable oils, alcohol alkoxylates and antioxidants. As examples of vegetable oils, there are mentioned sunflower oil, rapeseed oil, olive oil, castor oil, colza oil, maize kernel oil, cottonseed oil and soybean oil. Rapeseed oil is preferred. Among a list of numerous other possible additives, there are mentioned Indoxacarb and Fenazaquin, however, none of them exem- plified and not disclosed in combination.

The lack of disclosing the two pesticides Indoxacarb and Fenazaquin in combination can at least partly be attributed to the fact that they have very different modes of action. Fenazaquin is a quinazoline and belongs to the pesticide group 21 : MITOCHONDRIAL COMPLEX I ELECTRON TRANSPORT INHIBITORS, where the mode of action is a disruption of the biochemistry of insect mitochondria. Indoxacarb is a oxadiazine and belongs to the group 22A: VOLTAGE-DEPENDENT SODIUM CHANNEL BLOCKERS, where the mode of action is on nerves and muscles. Both are primarily used in agriculture.

Furthermore, once the insecticide is on the surface of the substrate, it should be prevented from crystallizing, typically called chalking, as this may lead to accelerated depletion from the storage. The above mentioned US6979455 by Ong et al. assigned to Microban Products discloses the problem of surface chalking on insec- ticidal polyethylene and proposes the addition of EVA or vinyl polymers.

It is very typical to state a large number of potential insecticides alone or in combination in a patent application without actually providing proof for its efficacy. Typically, it is a long way from the general proposal of a pesticide against a cer- tain type of insect until the insecticide actually finds its way to the market. For this reason, development in the field of efficient insecticides against mosquitoes is relatively slow despite a large number of proposals. As an example, for a polymer composition in fibrous pest control, including mosquito nets, a large variety of insecticides is mentioned in the patent applications WO2010/069697 assigned to BASF, US2003/0198659 by Hoffmann et al., and WO2006/12870 assigned to BASF, wherein the list of insecticides include Indoxa- carb as well as Fenazaquin, but not in combination. However, with respect to In- doxacarb against mosquitoes, the article of Ayesa Paul et al. "Evaluation of Novel Insecticides for Control of Dengue Vector Aedes aegypti (Diptera : Culicidae)" published in Journal of Medical Entomology, January 2006, concludes that Indoxacarb is moderately toxic to mosquito larvae but non-toxic to adult mosquitoes. This is in contradiction with prior art proposals for Indoxacarb as a useful insecticide against adult mosquitoes.

The fact that some pesticides, like Indoxacarb, are reported to work against mosquito larvae but not against adult mosquito reflects the different modes of action of pesticides in larvae as compared to flying insects. These findings are in line with experience that efficient acaricides against mites in agriculture are not necessarily useful against adult flies and adult mosquitoes. Thus, experimental results and practical experience from acaricides in agriculture is in general not transferable to insecticidal actions against adult mosquitoes. Therefore, in prac- tice, insecticidal and acaricidal science in agriculture and human malaria protection are typically very different fields with a low degree of overlap with respect of how insecticides are to be used and in which combinations and at what concentrations. It should be mentioned in general with respect to synergistic combinations of insecticides, that they typically should be applied together or at least subsequently within a short time interval. Various methods are discussed in the literature in order to fight mosquitoes or other insects. Substrates with various combinations of pesticidal agents are many-fold and include

- two agents distributed evenly in a coating on the substrate, for example as disclosed in WO01/37662 here combinations of various insecticides are proposed in combination with a pyrethroid;

- two agents evenly incorporated into a polymer of a substrate with migration to the surface, for example as disclosed in WO2010/015257, WO2011/124227 or WO2011/124228; differential migration control with different migration inhibitors inside the matrix is discussed in WO2003/063587, although in WO2009/003468, it is pointed out that different migration inhibitors influence each other such that a proper control is very difficult; - two agents incorporated in different parts of a fiber, for example as disclosed in WO2009/003468, such as in the core and shell of a bi-component fiber;

- two agents incorporated in different filaments combined into a single yarn, for example as disclosed in WO2009/003468;

- two agents in different yarns combined into a textile, for example as disclosed in WO2010/046348 or WO 2010/016561.

The large variety of proposed insecticides or combinations of insecticides against mosquitoes, the various proposals for means to control the release thereof, and the missing agreement on the effect of migration controllers reflect the ongoing race in the art to find further useful insecticidal systems. Accordingly, there is a steady need for improvements.

DESCRIPTION / SUMMARY OF THE INVENTION

It is therefore an objective of the invention to provide an improvement in the art. Especially, it is an objective to provide an alternative insecticidal product for counteracting insecticidal resistance in insects, particularly adult mosquitoes. It is also an objective to provide an alternative long lasting insecticidal product against insects, especially adult mosquitoes.

The objectives are achieved by an insecticidal substrate, for example mosquito net or wall lining, which has on its surface Fenazaquin (FNQ) and Indoxacarb (IDX). The objectives are also achieved by its use against insects, for example adult mosquitoes, and also by a method for production of the substrate and a method for killing insects, for example adult mosquitoes, as set forth in the following. It has been found experimentally that the combination of FNQ and IDX for some concentration ratios acts synergistically and for some other concentration ratios has an antagonistic effect on adult mosquitoes of the strain An. qradrimaculatus. This is a surprising finding, especially due to the very different mode of action as described above in the introduction.

Accordingly, when such type of adult mosquitoes are exposed to the combination of IDX and FNQ by contact with a substrate, it is advantageous to utilize the domain where a synergistic lethal effect is observed or an additive effect but avoid the domain where antagonistic effects are seen. Synergistic effects have been verified experimentally for concentration ratios of 1 : 1 and 1 :3 between IDX and FNQ, whereas a concentration ratio of 3 : 1 showed an antagonistic effect. Accordingly, for this mosquito strain, 3 : 1 is an approximate border between concentra- tion ratios that imply useful effects and non-useful effects when used against this specific mosquito strain. The antagonistic effect for a concentration ration of 3 and higher should be avoided. The border between the synergistic effect and the antagonistic effect is not sharp, as there is a region in between where an additive effect occurs, and it is reasonable to assume without explicit proof that an upper limit for the concentration ratio for synergistic effects is 2 : 1, possibly 1.5: 1. Although, no lower limit has been found experimentally, it is reasonable to assume without explicit proof that the lower limit is much smaller than 1 :3, for example 1 : 100 or even 1 : 100, however, for the lower limit having in mind that a certain concentration is necessary for the FNQ to have insecticidal efficacy.

It should be pointed out that there is a likelihood that similar synergistic effects are valid for some mosquito strains, however, the synergistic concentrations ratios may vary for other mosquito strains; and for some other strains, there may be no synergistic effect at all.

For example, in order to kill mosquitoes, an insecticidal fabric is provided and the insect, especially a mosquito, is caused contacting the yarn of the fabric while both FNQ and IDX are on the surface of the matrix such that FNQ and IDX are transferred both to the insect, especially a mosquito, by the contact.

In some embodiments, the surface of the substrate is coated with an impregnation formulation comprising IDX, FNQ, and an impregnation binder, and IDX and FNQ are caused migrating to the surface of the impregnation formulation. Alternatively, the substrate is a thermoplastic polymer, and the IDX and FNQ are mixed into the molten polymer, which is then extruded or molded as part of the production of the substrate, and IDX and FNQ are caused migrating to the surface of the polymer.

In alternative embodiments, the yarn of the fabric is a thread with intertwined first and second types of filaments. The first type of filaments has incorporated FNQ but not IDX and the second type of filaments has incorporated IDX but not FNQ. Alternatively, the first type of filaments has incorporated FNQ but not IDX and the second type of filaments has incorporated IDX as well as FNQ in order to provide a high total amount of FNQ without substantially influencing the physical properties of the filaments by high FNQ concentration in the polymer. The intertwined filaments imply that a mosquito contacting the thread will simultaneously pick up FNQ and IDX from the different types of filaments in the thread. Further filament types are, optionally, added, for example containing further active ingredients or being provided for sake of increased stability.

In further alternative embodiments, the fabric comprises a first type of yarn and a second type of yarn, the first type having incorporated FNQ but not IDX and the second type having incorporated IDX but not FNQ. Alternatively, the first type of yarn has incorporated FNQ but not IDX and the second type of yarn has incorporated IDX as well as FNQ in order to provide a high total amount of FNQ without substantially influencing the physical properties of the yarn by high FNQ concentration in the polymer. The two types of yarn are then interwoven, knitted togeth- er, or combined to a non-woven fabric as a compressed non-ordered mix of yarn. The type of yarn is optionally a monofilament or a multifilament or an intertwined yarn or a combination thereof.

The substrate, for example, the yarn advantageously is made of a thermoplastic polymer. Options include polyester (polyethylene terephthalate), polyamide, or polyolefin. Useful polyolefins include polyethylene and polypropylene, especially with respect to incorporation, whereas polyester and polypropylene are especially useful for impregnation. For polyethylene, especially for incorporation, monofilaments are suitable, especially for mosquito nets, although, also multifilaments can be used. For polypropylene and polyester, especially when used for impregnation, typically, multifilaments are used, for example 10-100 filaments. A useful yarn for mosquito nets has a weight of 50-100 Denier.

For incorporation, the substrate is typically made by extrusion of the thermo- plastic polymer, especially in the case of the substrate being a yarn, for example as part of a fabric, although, also molding can be used as a production technique in some cases, where the substrate is not a yarn. The method comprises melting the polymer, mixing IDX and FNQ into the molten polymer and extruding or molding the polymer as part of the production of the substrate and causing migration of IDX and FNQ to the surface of the substrate.

For example, in the extruded polymer, the concentration of FNQ is in the range of 0.5 and 5 wt%, optionally between 0.5 and 2 wt%, and the concentration of IDX is in the range of 0.5 and 5 g wt%, optionally between 0.5 and 2 wt%. However, the concentrations adjusted for the ratio to yield a synergistic effect.

For coating of the substrate, the IDX and FNQ are provided in an impregnation formulation that comprises a binder. For example, the impregnation formulation is provided by mixing a first intermediate formulation, and a second intermediate formulation with the addition of a polymeric binder, for example containing silicone or acrylate or urethane or a combination thereof. For example, the first intermediate formulation comprises IDX and water, optionally not FNQ, and the second intermediate formulation comprises FNQ, optionally not IDX.

Optionally, a spin finish agent is added to the impregnation formulation prior to coating, for example a surfactant. Advantageously, a part or all of the FNQ are provided on a microporous or na- noporous support. Useful types of support micro-particles with submicron pore sizes are disclosed in various patents by Amcol International Corporation and Amcol Health and Beauty Solutions Inc. In experiments, a specific type of porous support micro-particles were used which are sold by Amcol International Corporation under the brand name ES-4™. This type of porous particle is produced from methacrylic and methyl-ethylene glycol- bismethacrylate copolymer with the molecular formula C10H 14O4.C5H8O2)x and CAS# 25777-71-3. These microporous particles have been found experimentally as being a good migration control agent for FNQ.

Alternatively, other support particles could potentially be used, for example nano clay. Examples of nano-clays are attapulgite and montmorillonite. As an alternative to clay, micro-particles or nano-particles are used carriers in some embodi- ments. For example, a carrier is of the type of ground natural minerals or synthetic material, including silica, alumina and silicates. Examples of carriers include kaolin, talc, chalk, quartz, carbon black, diatomaceous earth, calcite, marble, pumice, sepiolite and dolomite. For example, the FNQ is present in the coating as a first portion and a second portion, the first portion being provided in particular form in the coating and the second portion being provided in dissolved form on a supporting micro-particle in the coating. Optionally, the production of the substrate comprises coating with an impregnation formulation containing FNQ and IDX, wherein the impregnation formulation is provided by dividing the FNQ into a first portion and a second portion, providing a first aqueous intermediate formulation with the first portion and a second intermediate formulation with the second portion, adding the IDX to the first and/or second intermediate formulation, and mixing the first and the second intermediate formulation with the addition of an impregnation binder to form the impregnation formulation. The first intermediate formulation is provided by mixing the first portion of FNQ with water.

For example, in an impregnation formulation, the concentration of FNQ is in the range of 1 and 10 wt%, the concentration of IDX is in the range of 1 and 10 wt%, however within the synergistic ratio domain as explained above.

For example, the weight ratio between the total FNQ and support particles is in the range of 3 to 15. For example, the weight ratio between the total FNQ and the polymeric binder is in the range of 2 to 6. As an example, 30 part IDX and 17.5 parts FNQ are suspended in water for the first intermediate formulation. For the second intermediate formulation 5 parts microporous support particles are suspended in water and mixed with 17.5 parts FNQ. The first and the second intermediate formulation are then mixed and a polymeric binder added as well as water for the final impregnation formulation. For example, there is added an amount of water to reach 1000 parts in total.

The idea behind mixing part of the FNQ into the water suspension with the IDX and part of it loaded on the porous micro-particle as a support is as follows: the FNQ mixed with the IDX provides a fast availability of the FNQ and the IDX on the surface of the impregnated/coated substrate, whereas the support-loaded FNQ is released slowly and yields a long term effect, keeping the ratio between FNQ and IDX within the synergistic regime.

In some successful experiments for mosquito nets, the final concentration of the FNQ and IDX on the dried mosquito net were 9 wt% and 8 wt% relatively to the total weight of the net, respectively. Exemplary useful ranges are expected to be 5 to 20 wt% for each of FNQ and IDX. All percentages herein given are in term of weight percentages relatively to the total weight of the resin composition or total weight of the impregnation composition, also denoted wt%. It is for sake of convenience pointed out that the measure of Denier is the weight of the yarn/filament per meter multiplied by 9000.

SHORT DESCRIPTION OF THE DRAWINGS

The invention will be explained in more detail with reference to the figures, in which

FIG. 1 is a chart illustrating the efficacy of FNQ and INX;

FIG. 2 is an electron micrograph of a knitted mosquito net coated with an impreg- nation formulation containing ES-4 microporous particles that are loaded with FNQ, where a) is an overview image and b) is an enlarged part of FIG. 2a.

DETAILED DESCRIPTION / PREFERRED EMBODIMENT

In the following, the term "wt%" is used for percentages in term of weight. Further, the following terms are used

FNQ= Fenazaquin,

IDX=Indoxacarb,

The combination of Indoxacarb (IDX) and Fenazaquin (FNQ) was tested in various experiments. In a first experiment, the general efficacy of combinations of IDX and FNQ was tested . In the second experiment, IDX and FNQ were melt- incorporated in a Polyethylene (PE) monofilament which was used for production of a mosquito net to which mosquitoes were exposed. In a third experiment, IDX and FNQ were part of an impregnation formulation used to coat a polypropylene (PP) multifilament yarn of a mosquito net.

Although, the fabric in the experiments was a mosquito net, the formulations and principles described herein do also apply to other types of fabrics, knitted, woven or non-woven, for example a wall lining hanging on the wall for killing insects that settle on the surface of such wall lining or a fencing protecting areas against flying insects, in particular flies and adult mosquitoes. The first experiment - synergistic and antagonistic efficacies

In this first experiments, glass vials were coated with a solution of IDX and/or FNQ in an acetone/triton mix with 0.5 wt% Dimethyl sulfoxide (DMSO) added. Mosquitoes of the species An. qradrimaculatus were brought in contact with the coated glass surface for testing insecticidal efficacy. Average EC50 (half maximal effective concentration) values based on multiple determinations were obtained. Deltamethrin was used for positive control, and the negative control was the carrier (acetone/triton mix + 0.5% DMSO).

TABLE 1

Taking offset in the approximate EC50 values of 18 ppm for FNQ and 8 ppm for IDX, Combination studies were conducted for FNQ and IDX using Loewe's ap- proach. For the Loewe approach, see, Loewe S (1928) "Die quantitativen probleme der pharmakologie", published in Ergebn Physiol 27:47-187; and Loewe S (1953) "The problem of synergism and antagonism of combined drugs" published in Arzneimittelforschung 3 : 285-290. A newer reference is Ronald J. Tallari- da (2006) "An Overview of Drug Combination Analysis with Isobolograms", pub- lished in JPET 319: 1-7, 2006.

Combination Index (CI) values were calculated at EC50, EC75, and EC90.

TABLE 2

Compound CombinaCone A Cone B CI at CI at CI at

Ratio

tion (ppm) (ppm) EC50 EC75 EC90

Indoxa-

3 : 1 75 25 270 486 904 carb: Fenazaquin

Indoxa-

1 : 1 50 50 0.44 0.20 O.IO carb: Fenazaquin

Indoxa-

1 : 3 25 75 0.59 0.32 0.20 carb: Fenazaquin As it appears from the table, the CI values were found to be less than 1 for ratios 1 : 1 and 1 : 3, suggesting synergistic effects, whereas antagonistic effects were found for the ratio of 3 : 1 between IDX and FNQ, seeing that the CI was much larger than 1. Accordingly, 3 : 1 is an upper limit for useful effects when used against this specific mosquito strain. The border between the synergistic effect and the antagonistic effect is not sharp, as there is a region in between where an additive effect occurs, and it is reasonable to assume without explicit proof that an upper limit for the concentration ratio for synergistic effects is 2 : 1, possibly 1.5 : 1. A lower limit for the synergistic effect, was not established, however, it can be reasonably be assumed that the effect is synergistic for much smaller ratios, as long as the concentration of Fenazaquin is sufficient for inducing an effect in the mosquito.

It should be pointed out that there is a likelihood that similar synergistic effects are valid for some other mosquito strains whereas different ratios may apply for other mosquito strains; and for some other strains, there may be no synergistic effect at all.

The two insecticides are of very different nature and have different targets in the mosquito, why at best, an additive effect was expected. Therefore, the finding of the synergistic effect was utmost surprising. Also surprising was the change of the efficacy from a synergistic domain to an antagonistic domain dependent on the concentration ratio.

The findings of the synergistic effect resulted in the decision to use FNQ and IDX in equal concentrations in further experiments where the efficacy of the combination of IDX and FNQ was tested against adult mosquitoes when the combination was used on mosquito nets produced according to procedures suitable for mass production.

The second experiment - Incorporation into mosquito net yarn

In this second experiment, IDX and/or FNQ were mixed into molten PE containing 90wt% HDPE and 10wt% LDPE, which was then extruded to a 100 Denier PE monofilament. The concentrations were 1.5wt% for IDX and 1.5wt% for FNQ. The resulting monofilaments were then knitted into a mosquito net, and mosquitoes of the type An. Gambiae Tiassale were exposed to the active agents for 3 minutes by contact with the mosquito net showing high knock down as well as killing efficacy.

The following terms are used in the graph :

TD24, TD48, TD72: Total death rate after 24, 48, or 72 hours, respectively.

FNQ: FNQ melt-incorporated in PE without IND and without SB. The concentration of FNQ in PE is 1.78 wt%

IDX: IDX melt-incorporated in PE but not FNQ or SB. The concentration of IDX in PE is 1.29 wt%

IDX+FNQ: IDX and FNQ and MESB melt-incorporated in PE. The concentration of IDX and FNQ were 1.8 wt% and 1.2 wt%, respectively.

In FIG. 1, the results are compared to the total death rate of this mosquito when exposed to PermaNet® 2.0 (PN2.0) and Permanet® 3.0 (PN3.0). PermaNet® 2.0 is a mosquito net impregnated with deltamethrin and Permanet® 3.0 is a mosquito net in which yarn deltamethrin is incorporated, and which also has piperonyl butoxide (PBO) incorporated as a synergist in order to kill pyrethroid- resistant mosquitoes. This is reflected in the total death rate (TD) in the two right triple columns for PN3.0 as compared to PN2.0, the latter not comprising a synergist. These two brands are state-of-the-art and used generally for comparison with other nets, especially by the World Health Organization (WHO).

The following concusions were drawn from the results illustrated in FIG. 1. The incorporation of FNQ alone, as illustrated in the first column, did not result in a high efficacy. The low but not negligable efficacy proves that FNQ migrates to the surface to some extent, but the mosquitoes are not deadly affected. From the second column with IDX only in the polymer, it appears that IDX as compared to FNQ has a much higher efficacy in this mosquito strain. This is in line with the results in tabel 1 where a lower concentration of IDX relatively to FNQ resulted in a higher death TD24 rate. From the third column in FIG. 1, illustrating the efficacy of the combination of IDX and FNQ, it is seen that it results in a surprisingly strong synergistic efficacy, more than the additive effect.

When comparing the effect to the mosquito net Permanet ® 3.0 (PN3.0), only the combination of IDX and FNQ reached the same efficacy, and it was even higher than the efficacy of Permanet® 2.0 (PN2.0). Accordingly, this combination constitutes a proper substitution of PN3.0, which is highly interesting in case of pyrethroid resistance. The third experiment - Impregnation of a mosquito net

In a third series of experiments, the insect killing efficacy of Indoxacarb (IDX) and Fenazaquin (FNQ) was tested when used in a coating of fibers, in particular fibers in a mosquito net. Also, the wash resistance was tested.

An impregnation formulation was made which included a first intermediate formulation, a second intermediate formulation, and a polymeric binder, which were mixed together.

The concentration of IDX and FNQ in the final formulation was 30 grams per liter and 35 grams per liter, respectively. Thus, the final formulation had a concentration ratio between IDX and FNQ of 30: 35 which is equal to 1 : 1.2. This is in the synergistic domain, which experimentally in Experiment 1 was verified for the range between 1 : 1 and 1 : 3.

The FNQ was fine milled to a particle size dominantly in the region of 2-5 microns, although, the milling also provided a small portion of even smaller and larger particles.

In some experiments, the FNQ particle size distribution after milling comprised particles with a diameter of between 0.4 microns and 20 microns, having a mean of 4.9 microns and a median of 3.8 medians. 10% of the particles had a diameter of less than 1.0 micron, and 25% a diameter of less than 1.7 microns.

For the IDX, the particle size was larger, the size distribution having a mean of 13 microns and a median of 8 microns. 10% of the particles had a diameter of less than 1.4 micron, and 25% a diameter of less than 3.1 microns. The larger size of the particles was intentional in order to promote loading of FNQ into porous sup- port particles as compared to loading IDX into or onto the support particles.

For the first intermediate formulation, half of the FNQ and all of the IDX were mixed in water suspension. The amount of water in the suspension was adjusted to yield the final concentration of 30 g/l and 35 g/l, respectively. However, the amount of water and the corresponding concentration of IDX and FNQ in the final formulation is not an important factor, as the water is evaporated in the final step of the coating process. Important is the ratio between the IDX and the FNQ. For the second intermediate formulation, the other half of the FNQ was loaded onto porous support particle of the type ES-4, which were added at a weight ratio of 1 :3 for FNQ: ES-4. The water-dissolved portion of FNQ is loaded into the ES-4 particles through the pores and also onto the surface of the ES-4 particles. The smallest of the fine-milled FNQ particles, which have sub-micron size, are loaded into and onto the ES-4 particles, whereas the larger micron-sized are potentially loaded onto the ES-4 particles, as they do not fit through the pores.

The two intermediate formulations were mixed together, and a polymeric binder formulation added prior to coating of the fabric, for example the mosquito net. The polymeric binder can also be added into the water prior to mixing. The binder potentially results in further loading of FNQ into and onto the ES-4 particles. In the mix, some of the dissolved IDX may also be loaded into the ES-4 particles. The addition of porous support particles for the FNQ is optional. As an alternative to the EN-4 particles, the support particles are clay particles, for example mont- morillonite, which is a 2-to-l layered smectite clay mineral with a plate-like structure. For example, such porous particles are marketed by Nanocor®. Important is the fact that the support particles, such as the ES-4 or the nano-clay is provided at a concentration to take up the FNQ as a storage buffer for a long term release effect. The support improves the wash resistance and the insecticidal synergistic lifetime of the product. If no support, such as ES-4 or nano-clay, is provided, the synergistic effect will also be achieved, however, the wash resistance of the impregnated fabric is less, and the general synergistic insecticidal lifetime of the impregnated product is shorter when washed repeatedly. The idea behind mixing part, for example half, of the FNQ in particular form into the water suspension with the particular IDX and loading the remaining portion, for example half, of the FNQ on the support is, on the one hand, quickly available FNQ from the FNQ micro-particles in the coating, and, on the other hand, a retain mechanism on the FNQ by the support for a long term FNQ supply. The particular FNQ mixed with the IDX provides a fast availability of the FNQ and the IDX on the surface of the impregnated/coated substrate. The content of FNQ relatively to the IDX keeps the ratio FNQ: IDX within the synergistic regime. However, with each wash, the FNQ is washed more out of the binder than the IDX due to the higher water solubility. Thus, after a few washes, the concentration of FNQ particles in the coating is reduced to an extent such that the ratio cannot be maintained within the synergistic regime, as the reduction of FNQ from the binder is more pronounced than the loss of the IDX from the binder. Seeing that with increasing number of washes, the ratio of the FNQ and IDX concentration in form of particles changes, there is a need for a secondary supply of FNQ in order to keep the ratio within the synergistic regime. For this reason, the FNQ is provided on the support particles. The dissolved FNQ release from the support is less pronounced when washed, which ensures a long term wash-resistant effect. However, with in- creased washing, the IDX concentration has been reduced in the coating, such that slow the wash-resistant release of dissolved FNQ from the support particles is sufficient to maintain the long term synergistic ratio between FNQ and IDX. This way, the time frame immediately after impregnation is covered as well as the time frame on a long term scale after repeated washing. When the FNQ from the particular FNQ in the coating is getting exhausted, the FNQ on the support is still present for gradual released such that the FNQ on the surface can be continuously replenished, especially after washing.

For the polymeric binder, various options are available, and numerous examples can be found in the prior art on emulsion polymers that are suitable for coating on a substrate, especially fabrics. Examples are acrylate binders and silicone binders or mixtures thereof. The polymeric binder formulation used in the experiment was a mix of a silicone binder and acrylate binder of the type PS14195 and PS12617 as available from Goulsten Technologies, however, other types of silicone and acrylate binders apply as well. In the experiment, the concentration of the polymer of the binder was 12 g/l, however, this can be varied without affecting the synergistic effect, as the synergistic effect depends primarily on the ratio of the concentration of FNQ and IDX and not on the concentration itself. The sili- cone/acrylate binder of the type PS14195/12617 was provided as a mix of equal concentration.

Addition of small amounts of wetting agent, anti-foam agents or dispersants can also be considered as well as spin finish agents. The final formulation was used for coating/impregnation of a mosquito net made from 750 Denier polypropylene multifilament yarn with 36 filaments. The formulation was transferred to the net from a bath by so-called kiss rollers, which are partly inserted into the bath and transfer the formulation from the bath to the top of the roller over which the fabric is transported in contact with the roller. Calendar rollers were used to squeeze the fabric to adjust the amount of the formulation on the dried net to approximately 9 wt% FNQ and 8 wt% IDX relatively to the total weight of the dried net.

Electron microscope images of the knitted polypropylene multifilament net are shown in FIG. la and lb, the latter being an enlarged section of FIG. la. The particle size was in the range of 20-35 microns The impregnated nets were used for an insecticidal 3 minutes exposure to mosquitoes of a laboratory mosquito strain A.gambiae (Akron) in a WHOPES cone test. The total death rates after 1 hour were 90%, and after 24 hours (TD24) and after 48 hours (TD48) were 100%. As long lasting insecticidal nets (LLIN) are washed from time to time when in real use over long periods, the WHOPES has provided guidelines with requirements of wash resistance of up to 20 washes. In order to test the wash resistance of the impregnation, the impregnated nets were washed. The washing procedure was performed in accordance with the standard washing procedure as described and requested by WHOPES. Net samples (25 cm x 25 cm) are introduced individually into beakers containing 0.5 I deionized water, with 2 g/l soap (pH) added and fully dissolved just before washing. The beakers are introduced into a water bath at 30 °C and shaken for 10 min at 155 movements per minute. The samples are then removed, rinsed twice for 10 min in clean, deionized water under the same shak- ing conditions as above, dried at room temperature and stored at 30 °C in the dark between washes. The washes were conducted as rapid sequences of 5 washes per day, and tested 1 day after the last wash with the pyrethroid resistant strain of Akron. After washing 15 times and 20 times, respectively, a similar cone test was performed . The TD24 rate decreased to 80% in both cases, and the TD48 rates were 95% and 87%, respectively. Thus, the impregnation performed well for killing mosquitoes even if the net was washed 20 times.

Claims

1. A method of killing insects, the method comprising providing a non-living substrate with Indoxacarb, IDX, and Fenazaquin, FNQ, on its surface and exposing the insect to the Indoxacarb and Fenazaquin by contact with the surface.

2. A method according to claim 1, wherein the insect is an adult mosquito.

3. A method according claim 2, the method comprising providing the IDX and the FNQ on the surface with a concentration ratio IDX: FNQ between the concentration of IDX and the concentration of FNQ, adjusting the concentration ratio IDX: FNQ for synergistic or additive lethal action on the adult mosquito but not antagonistic action, and exposing the adult mosquito to the IDX and FNQ in this synergistic concentration ratio by contact with the surface.

4. A method according to claim 2, wherein the concentration ratio of IDX: FNQ is less than 2 : 1.

5. A method according to claim 2, wherein the concentration ratio of IDX: FNQ is less than 1 : 1.

6. A method according to any preceding claim, the method comprising coating the surface of the substrate with an impregnation formulation comprising IDX, FNQ and an impregnation binder and causing migration of IDX and FNQ to the surface of the impregnation formulation; or wherein substrate is a thermoplastic polymer and the method comprises melting the polymer, mixing IDX and FNQ into the molten polymer and extruding or molding the polymer as part of the production of the substrate and causing migration of IDX and FNQ to the surface of the polymer.

7. A method according to any preceding claim, comprising providing the substrate as a fabric.

8. A substrate that is non-living and comprises a surface for contact with an in- sect, the surface comprising a combination of Indoxacarb, IDX, and Fenazaquin,

FNQ.

9. A substrate according to claim 8, wherein the IDX and FNQ are provided on the surface with a concentration ratio IDX: FNQ between the concentration of IDX and the concentration of FNQ for synergistic or additive lethal but not antagonistic effect on adult mosquitoes.

10. A substrate according to claim 9, wherein the weight concentration ratio IDX: FNQ is less than 2 : 1.

11. A substrate for a method according to claim 9, wherein the weight con- centration ratio IDX: FNQ is less than 1 : 1.

12. A substrate according to anyone of the claims 8-11, wherein the substrate is a thermoplastic polymer having melt-incorporated therein IDX and FNQ and being configured for migration of IDX and FNQ to the surface of the polymer.

13. A substrate according to claim 12, wherein concentration of FNQ in the polymer is in the range of 0.5 and 5 wt%.

14. A substrate according to claim 12 or 13, wherein, and the concentration of IDX is in the range of 0.5 and 5 g wt%.

15. A substrate according to anyone of the claims 12-14 , wherein the substrate is an extruded polylolefin monofilament yarn.

16. A substrate according to anyone of the claims 8-11, the substrate comprising a coating forming a surface on the substrate, the coating comprising the combination of IDX and FNQ.

17. A substrate according to claim 16, wherein the FNQ is present in the coating as a first portion and a second portion, the first portion being provided in particular form and the second portion being provided in dissolved form on a supporting micro-particle.

18. A substrate according to claim 16 or 17, wherein the substrate is a multi- filament polypropylene yarn with 50-100 Denier, the multifilament yarn comprising 10-100 monofilaments.

19. A substrate according to claim 15 or 18, wherein the yarn is part of a fabric.

20. A substrate according to claim 19, wherein the fabric is a mosquito net.

21. A method for producing a substrate according to claim 16, wherein the method comprises providing a substrate and coating it with an impregnation formulation containing FNQ and IDX; wherein the impregnation formulation is provided by dividing the FNQ into a first portion and a second portion, providing a first aqueous intermediate formulation with the first portion and a second intermediate formulation with the second portion, adding the IDX to the first or second intermediate formulation, and mixing the first and the second intermediate formulation with the addition of an impregnation binder to form the impregnation formulation; wherein the first intermediate formulation is provided by mixing the first portion of FNQ with water.

22. A method for producing a substrate according to claim 21, wherein the second intermediate formulation is provided by adding porous micro-particles as support for the FNQ in or on the micro-particles.

23. A method according to claim 22, wherein the porous micro-particles are of the type ES-4™ marketed by Amcol, CAS# 25777-71-3, the micro-particles having a size in the range of 10-50 microns.

24. A method according to anyone of the claims 21-23, wherein the concentration ratio between the concentration of IDX and the concentration of FNQ in the impregnation formulation is 2 : 1.

25. A method according to claim 24, wherein the concentration ratio between the concentration of IDX and the concentration of FNQ in the impregnation formulation is less than 1 : 1.