USOO869 1256B2

(12) United States Patent (10) Patent No.: US 8,691.256 B2 Enan (45) Date of Patent: Apr. 8, 2014

(54) PEST CONTROL COMPOSITIONS AND (56) References Cited METHODS U.S. PATENT DOCUMENTS (75) Inventor: Essam Enan, Davis, CA (US) 4,320,113 A 3/1982 Kydonieus 4,943,435 A 7, 1990 Baker (73) Assignee: Tyratech, Inc., Melbourne, FL (US) 5,980,931 A 11, 1999 Fowler (*) Notice: Subject to any disclaimer, the term of this (Continued) patent is extended or adjusted under 35 U.S.C. 154(b) by 229 days. FOREIGN PATENT DOCUMENTS WO WOOO,05964 ck 2, 2000 (21) Appl. No.: 12/936,039 WO WO 2004/OO6968 1, 2004 (22) PCT Fled: Mar. 19, 2009 (Continued) OTHER PUBLICATIONS (86) PCT NO.: PCT/US2009/037.733 Appert-Collin, A., “Regulation of g protein-coupled receptor signal S371 (c)(1), ing by a-kinase anchoring proteins.” Recept. Signal Transduct. Res., (2), (4) Date: Jan. 28, 2011 vol. 26, p. 631-646, 2006. (87) PCT Pub. No.: WO2O09/117621 (Continued) PCT Pub. Date: Sep. 24, 2009 Primary Examiner — Neil Levy (74) Attorney, Agent, or Firm — Davis Wright Tremaine (65) Prior Publication Data LLP US 2011 FO1245O2A1 May 26, 2011 (57) ABSTRACT Related U.S. Application Data Embodiments of the present invention provide compositions for controlling a target pest including a first agent and a (60) Provisional application No. 61/070,137, filed on Mar. second agent comprising a pest control product or a signal 19, 2008, provisional application No. 61/043,084, cascade modulator, wherein the first agent and the second filed on Apr. 7, 2008, provisional application No. agent act synergistically to control the target pest. The first 61/048,477, filed on Apr. 28, 2008, provisional agent can be capable of interacting with a receptor in the application No. 61/087,140, filed on Aug. 7, 2008. target pest. The pest control product can have a first activity against the target pest when applied without the active agent (51) Int. C. and the compositions can have a second activity against the AOIN 25/32 (2006.01) target pest, and the second activity can be greater than the first (52) U.S. C. activity. Embodiments of the invention can include composi USPC ...... 424/406; 424/405; 424/745; 514/341: tions that modulate the signal cascade initiated by the binding 514/398; 514/552; 514/739 of ligands to, for example, cell surface receptors. Methods of (58) Field of Classification Search screening Such compositions are also disclosed. None See application file for complete search history. 6 Claims, 37 Drawing Sheets

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(56) References Cited Neitzel, K.L., “Cellular mechanisms that determine selective RGS protein regulation of G protein-coupled receptor signaling.” Semin U.S. PATENT DOCUMENTS Cell. Dev. Biol., vol. 17, p. 383-389, 2006. New, D.C., “Molecular mechanisms mediating the G protein 6,360,477 B1 3, 2002 Flashinski coupled receptor regulation of cell cycle progression.” J. Mol. Sig 6,610,254 B1 8, 2003 Furner naling, vol. 2, p. 2-16, 2007. 6,849,614 B1* 2/2005 Bessette et al...... 514/72 Premont, R.T., “Physiological roles of G protein-coupled receptor 2003/0060379 A1 3f2003 Souter et al...... 510,131 kinases and arrestins.” Annu. Rev Physiol., vol. 69, p. 511-534, 2007. 2003/0194454 A1* 10, 2003 Bessette et al...... 424,745 Sato, M., "Accessory proteins for G proteins; partners in signaling.” 2003. O1986.59 A1 10/2003 Hoffmann Annu. Rev. Pharmacol. Toxicol., vol. 46, p. 151-187, 2006. 2004/O185080 A1 9/2004 Hojo Saudou, et al., “Cloning and characterization of a Drosophila 2005, 0008714 A1 1/2005 Enan tyramine receptor.” The EMBO Journal, vol. 9(11), p. 3611-7, 1990. 2005/0214267 A1 9, 2005 Enan Schulte, G., “Novel aspects of G-protein-coupled receptor signal 2006/0121126 A1 6, 2006 McFadden ling—different ways to achieve specificity.” Acta Physiol., vol. 190, 2006/0263.403 A1 11/2006 Enan p. 33-38, 2007. 2007/0232495 A1 10/2007 Nappa Smrcka, A.V., “G protein beta gamma Subunits: Central mediators of 2007/0264297 A1 11/2007 Scialdone G protein-coupled receptor signaling.” Cell. Mol. LifeSci., vol. 65, p. 2007/0272.281 A1 11/2007 Pel 2191-2214, 2008. 2008.0020078 A1 1/2008 Enan Stewart, A.J., “Phospholipase C-eta Enzymes as Putative Protein 2008/0047312 A1 2/2008 Hill et al...... 71.21 Kinase C and Ca Signalling Components in Neuronal and 2008.0075796 A1 3/2008 Enan Neuroendocrine Tissues.” Neuroendocrinology, vol. 86, p. 243-248, 20080118585 A1 5/2008 Nouvel ...... 424f739 2007. Takeishi, Y., “Role of diacylglycerol kinase in cellular regulatory FOREIGN PATENT DOCUMENTS processes: a new regulator for cariomyocyte hypertrophy.” Pharmacol. Ther. vol. 115, p. 352-359, 2007. WO WO 2006/061803 6, 2006 Torrecilla, I., "Co-ordinated covalent modification of G-protein WO WO 2008.088827 T 2008 coupled receptors.” Curr. Pharm. Des., vol. 12, p. 1797-1808, 2006. WO WO 2009,117621 9, 2009 Von Nickisch-Rosenegk, et al., “Cloning of biogenic amine receptors from moths (Bombyx mori and Heliothis virescens ).” Insect OTHER PUBLICATIONS Biochem. Mol. Biol., vol. 26, p. 817 -827, 1996. Wolfe, B.L., "Clathrin-DependentMechanisms of G Protein-coupled Baxter, G.D., et al., “Isolation of a cDNA for an octopamine-like, Receptor Endocytosis.” Traffic, vol. 8, p. 462-470, 2007. G-protein coupled receptor from the cattle tick, Boophilus Xu, Z.O., “Regulation of G protein-coupled receptor trafficking.” microplus.” Insect Biochem. Mol. Biol., vol. 29, p. 461–467, 1999. Acta Physiol, vol. 190, p.39-45, 2007. Colby, S.R., "Calculating synergistic and antagonistic responses of Yang, W., “Mechanisms of regulation and function of Gprotein herbicide combinations.” Weeds, vol. 15:1, p. 20-22, 1967. coupled receptor kinases.” World J. Gastroenterol, vol. 12, p. 7753 Dalrymple, M.B., “G protein coupled receptor dimers: functional 7757, 2006. consequences, disease states and drug targets.” Pharmacaol. Ther. Yu, J.H., “Heterotrimeric G protein signaling and RGSs in Aspergil vol. 118, p. 359-371, 2008. lus nidulans.” J. Microbio. vol. 44, p. 145-154, 2006. Dong, C., “Regulation of G protein coupled receptor export traffick Genbank Accession No. AF343878, “Mamestra brassicae putative ing.” Biochem. Biophys. Acta, vol. 176, p. 853-870, 2007. octopamine receptor (OAR) mRNA, complete cods.” 2001 online Enan, E., et al., “Deltamethrin-induced thymus atrophy in male Retrieved on Nov. 19, 2012 Retrieved from: http://ww.ncbi.nlm. Balb/c mice.” Biochem. Pharmacol., vol. 51 No. 4, p. 447-454, 1995. nih.gov/nuccore/13173420. Gilchrist, A., “G-protein-coupled receptor pharmacology: examining Genbank Accession No. AJO07617. “Drosophila melanogaster mRNA for octopamine receptor, splice variant 1B, 1999 online the edges between theory and proof.” Curr. Opin. Drug Discov. Retrieved on Nov. 19, 2012 Retrieved from: http://www.ncbi.nlm. Devel., vol. 10, p. 446-451, 2007. nih.gov/nuccore/AJ007617. Grandy, D.K., “Trace amine-associated receptor 1—Family arche U.S. Appl. No. 61/043,084, filed Apr. 7, 2008, “Insect Control Meth type or iconoclast?” Pharmacol. Ther... vol. 116, p. 355-390, 2007. ods and Compositions Using Neonicotinoids'. Han, K.A., et al., “A Novel Octopamine Receptor with Preferential U.S. Appl. No. 61/048.477, filed Apr. 28, 2008, “Pest Control Com Expression in Drosophila Mushroom Bodies,” J. Neurosci., vol. 18. positions and Methods”. p. 3650-3658, 1998. U.S. Appl. No. 61/070,137, filed Mar. 19, 2008, “Insect and Parasite Jurado-Pueyo, M., “GRK2-dependent desensitization downstream Control Methods and Compositions”. of G proteins.” Recept. Signal Transduct. Res., vol. 28, p. 59-70, U.S. Appl. No. 61/087.140, filed Aug. 7, 2008, "Compositions and 2008. Methods for Increasing Pesticide Effectiveness”. Klaasse, E., “Internalization and desensitization of adenosine recep PCT International Preliminary Report on Patentability in corre tors.” Purinergic Signalling, vol. 4, p. 21-37, 2008. sponding International application No. PCT/US2009/037733, issued Ma, L., “Beta-arrestin signaling and regulation of transcription.” J. Sep. 21, 2010, 12 pages. Cell. Sci., vol. 120, p. 213-218, 2007. First Office Action of the State Intellectual Property Office, mailed in Milligan, G., “New aspects of g-protein-coupled receptor signalling related Chinese Application No. 200980118165.6, dated Nov. 1, and regulation.” Trends Endocrinol. Metab., vol. 9, p. 13-19, 1998. 2011. Nakahata, N., “Regulation of G Protein-coupled Receptor Function by Its Binding Proteins.” Yakugaku Zasshi, vol. 127, p. 3-14, 2007. * cited by examiner U.S. Patent Apr. 8, 2014 Sheet 1 of 37 US 8,691.256 B2

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Off US 8,691,256 B2 1. 2 PEST CONTROL COMPOSITIONS AND Common to most GPCRs is the cyclic process of signaling, METHODS desensitization, internalization, resensitization, and recycling to the plasma membrane. This cycle prevents cells from CROSS-REFERENCE TO RELATED undergoing excessive receptor stimulation or periods of pro APPLICATIONS 5 longed inactivity. Mechanisms for desensitization of GPCRs include receptor phosphorylation and Subsequent endocyto The present application is a U.S. National Stage entry sis, which removes the receptor-ligand complex from the cell under 35 U.S.C. S371 of International Application No. PCT/ Surface. As a result of this desensitization process, a common US2009/037733, filed Mar. 19, 2009, designating the United limitation of GPCR-targeted compositions is target organism States of America and published in English on Sep. 24, 2009, 10 tolerance or resistance, as receptor desensitization can mute which in turn claims priority to U.S. Application No. 61/070, their effectiveness. 137, filed Mar. 19, 2008, U.S. Application No. 61/043,084, filed Apr. 7, 2008, U.S. Application No. 61/048,477, filed Apr. SUMMARY 28, 2008, and U.S. Application No. 61/087,140, filed Aug. 7, 15 2008, each of which is incorporated herein by reference in its The present disclosure relates to embodiments of a com entirety. position for controlling a target pest comprising a first agent and a second agent, wherein the first agent has a first activity FIELD OF THE INVENTION against the target pest, the second agent has a second activity 2O against the target pest, the composition has a third activity The present invention relates to compositions and methods against the target pest, and the third activity reflects a syner related to controlling insects. The invention also relates to gistic interaction of the first agent and the second agent. compositions and methods for altering and/or disrupting G In a further aspect, the first agent comprises one or more protein-coupled receptor signaling and cycling resulting in compounds selected from the group consisting of amyl the disruption of normal biology or behavior in invertebrates. 25 butyrate, trans-anethole, anise star oil, black seed oil, citral, p-cymene, genistein, geraniol, geranyl acetate, isopropyl BACKGROUND OF THE INVENTION myristate, d-limonene, linalool, linallyl acetate, lilac flower oil, methyl salicylate, a-pinene, piperonal, piperonyl alcohol, While the first recorded use of chemicals to control pests tetrahydrolinalool, thyme oil, thyme oil white, thymol, tri dates back to 2500 BC, only in the last 60 years has chemical 30 ethylcitrate, Vanillin, and wintergreen oil. control has been widely used. Early pesticides included helle In a further aspect, the first agent is capable of interacting bore to control body lice, nicotine to control aphids, and with a receptor in the target pest. pyrithrin to control a wide variety of insects. Lead arsenate In a further aspect, the receptor is a G protein-coupled was first used in 1892 as an orchard spray, while at the same receptor. time it was discovered that a mixture of lime and copper 35 In a further aspect, the second agent is selected from the sulphate (Bordeaux mixture) controlled downy mildew, a group consisting of a pesticide, a fungicide, an herbicide, a fungal disease of grapes. nematicide, an insecticide, an acaricide, and a bacteriocide. The modern era of chemical pest control commenced dur In a further aspect, the second agent acts on a molecular ing World War II. For example, DDT played a major role in target other than the receptor. maintaining the health and welfare of soldiers who used it to 40 In a further aspect, the first agent is capable of interacting control body lice and mosquitoes. Further developments of with the receptor to trigger, alter, or disrupt a biological pesticides followed, and with their relatively low cost, ease of function related to the binding of the receptor with the first use, and effectiveness, they became the primary means of pest agent, and the second agent is capable of interacting with a control. Protection of crops, produce, animals, and humans non-receptor molecule or step associated with cycling of the over extended periods became possible with corresponding 45 receptor, to disrupt the cycling of the receptor. increases in food production and improved Standards of liv In a further aspect, the first activity persists for a first ing. period, the second activity persists for a second period, the Some modern pesticides are Sophisticated compounds that third activity persists for a third period, and the third period is are carefully researched to ensure they are effective against longer than either the first period or the second period. target organisms, generally safe to the environment, and can 50 In a further aspect, at least one of the first activity and the be used without undue hazard to users or consumers. Many of second activity is essentially Zero. these have been developed to target specific biochemical In a further aspect, the target pest is a species belonging to reactions within the target organism, e.g. an enzyme neces an animal order selected from the group consisting of Acari, sary for photosynthesis within a plant or a hormone required Anoplura, Araneae, Blattaria, Blattodea, Coleoptera, Collem for normal development in an insect. Thus, some modern 55 bola, Diptera, Grylloptera, Hemiptera, Heteroptera, chemicals are safer, more specific, and friendlier to the envi Homoptera, Hymenoptera, Isopoda, Isoptera, Lepidoptera, ronment than the older products they have replaced. Mantodea, Mallophaga, Neuroptera, Odonata, Orthoptera, Furthermore, G protein-coupled receptors (GPCRs) form Psocoptera, Rhabditida, Siphonaptera, Symphyla, Thysa one of the largest families of integral membrane receptors. nura, and Thysanoptera. GPCRs transduce information provided by extracellular 60 In a further aspect, the second agent is a diamide com stimuli into intracellular second messengers via their cou pound. pling to heterotrimeric G proteins and the Subsequent regu In a further aspect, the second agent is a spirocyclic phenyl lation of a variety of effector Systems. Therapeutic agents substituted tetronic acid pesticide. often target GPCRs because of their capability to bind In a further aspect, the second agent is selected from the ligands, hormones, and drugs with high specificity. Agonist 65 group consisting of fipronil, clothianidin, imidacloprid, activation of GPCRs also initiates processes that desensitize abamectin, forskolin, genistein, yohimbine, fluoxastrobin, GPCR responsiveness and their internalization. flubendiamide and spiromesifen. US 8,691,256 B2 3 4 In a further embodiment, the second agent comprises In a further aspect of this embodiment, the first agent fipronil. comprises isopropyl myristate, thyme oil white, and winter In a further aspect of this embodiment, the first agent green oil. comprises amylbutyrate and anise star oil. In a further aspect of this embodiment, isopropyl myristate In a further aspect of this embodiment, amylbutyrate is is present within a range of 30%-40%, thyme oil white is present within a range of 20%-30%, and anise star oil is present within a range of 15%-25%, and wintergreen oil is present within a range of 40%-60%. present within a range of 40%-50%. In a further aspect of this embodiment, the first agent In a further embodiment, the second agent comprises comprises geraniol, isopropyl myristate, and thyme oil white. clothianidin. 10 In a further aspect of this embodiment, the first agent In a further aspect of this embodiment, geraniol is present comprises d-limonene, thyme oil white, and lilac flower oil. within a range of 25%-35%, isopropyl myristate is present In a further aspect of this embodiment, d-limonene is within a range of 35%-45%, and thyme oil white is present present within a range of 25%-35%, thyme oil white is within a range of 25%-35%. present within a range of 25%-35%, and lilac flower oil is In a further aspect of this embodiment, the first agent 15 present within a range of 35%-50%. comprises p-cymene, linalool, a-pinene, and thymol. In a further embodiment, the second agent comprises In a further aspect of this embodiment, p-cymene is present yohimbine, forskolin, or genistein. within a range of 25%-35%, linalool is present within a range In a further aspect of this embodiment, the first agent of 5%-10%, a-pinene is present within a range of 2%-5%, and comprises isopropyl myristate and thyme oil white. thymol is present within a range of 30%-45%. In a further aspect of this embodiment, the first agent In a further aspect of this embodiment, the first agent additionally comprises geraniol. comprises isopropyl myristate, d-limonene, linalool, piper In a further aspect of this embodiment, geraniol is present onal, piperonyl alcohol, tetrahydrolinalool, and Vanillin. within a range of 25%-35%, isopropyl myristate is present In a further aspect of this embodiment, isopropyl myristate within a range of 35%-45%, and thyme oil white is present is present within a range of 15%-25%, d-limonene is present 25 within a range of 25%-35%. within a range of 5%-15%, linalool is present within a range In a further aspect of this embodiment, the first agent of 10%-20%, piperonal is present within a range of 15%- additionally comprises wintergreen oil. 25%, piperonyl alcohol is present within a range of 5%-15%, In a further aspect of this embodiment, wintergreen oil is tetrahydrolinalool is present within a range of 15%-25%, and present within a range of 40%-50%, isopropyl myristate is vanillin is present within a range of 0.5%-5%. 30 present within a range of 30%-40%, and thyme oil white is In a further embodiment, the second agent comprises imi present within a range of 15%-25%. dacloprid. In a further embodiment, the second agent comprises for In a further aspect of this embodiment, the first agent Skolin or genistein, and the first agent comprises t-anethole, comprises geraniol, isopropyl myristate, and thyme oil white. p-cymene, linalool, a-pinene, and thymol. In a further aspect of this embodiment, geraniol is present 35 In a further aspect of this embodiment, t-anethole is present within a range of 15%-35%, isopropyl myristate is present within a range of 15%-25%, p-cymene is present within a within a range of 35%-45%, and thyme oil white is present range of 0.5%-10%, linalool is present within a range of within a range of 25%-45%. 30%-45%, a-pinene is present within a range of 2%-10%, and In a further aspect of this embodiment, the first agent thymol is present within a range of 30%-45%. comprises wintergreen oil, isopropyl myristate, and thyme oil 40 In a further embodiment, the second agent comprises white. genistein. In a further aspect of this embodiment, wintergreen oil is In a further aspect of this embodiment, the first agent present within a range of 20%-60%, isopropyl myristate is comprises anise star oil. present within a range of 30%-40%, and thyme oil white is In a further aspect of this embodiment, the first agent present within a range of 2%-45%. 45 additionally comprises amylbutyrate. In a further aspect of this embodiment, the first agent In a further aspect of this embodiment, amyl butyrate is comprises p-cymene, linalool, a-pinene, and thymol. present within a range of 20%-30%, and anise star oil is In a further aspect of this embodiment, p-cymene is present present within a range of 40%-60%. within a range of 25%-35%, linalool is present within a range In a further embodiment, the second agent is an alkaloid. of 5%-10%, a-pinene is present within a range of 2%-5%, and 50 In a further aspect of this embodiment, the second agent is thymol is present within a range of 30%-45%. selected from the group consisting of caffeine, theobromine In a further aspect of this embodiment, the first agent and theophylline. comprises d-limonene and thyme oil white. In a further embodiment, the second agent is a flavonoid. In a further aspect of this embodiment, the first agent In a further aspect of this embodiment, the second agent is additionally comprises lilac flower oil. 55 selected from the group consisting of epicatechin, hesperidin, In a further aspect of this embodiment, d-limonene is kaempferol, naringin, nobiletin, proanthocyanidins, querce present within a range of 25%-35%, thyme oil white is tin, resveratrol, rutin, and tangeretin. present within a range of 25%-35%, and lilac flower oil is In a further embodiment, the second agent is an isoflavone. present within a range of 35%-50%. In a further aspect of this embodiment, the second agent is In a further aspect of this embodiment, the first agent 60 selected from the group consisting of daidzein, biochanin A, additionally comprises wintergreen oil. coumesterol, formononetin, and puerarin. In a further aspect of this embodiment, d-limonene is In a further embodiment, the second agent is a flavone, a present within a range of 50%-60%, thyme oil white is flavonol, a flavanone, a 3-hydroxyflavanone, a flavan-3-ol, or present within a range of 10%-20%, and wintergreen oil is an anthocyanidin. present within a range of 25%-40%. 65 In a further aspect of this embodiment, the second agent is In a further embodiment, the second agent comprises selected from the group consisting of apigenin, luteolin, abamectin. diosmin, flavoxate, fisetin, isorhamnetin, myricetin, pachy US 8,691,256 B2 5 6 podol, rhamnazin, morin, eriodictyol. hesperetin, homoeri and wherein the second agent is capable of interacting with a odictyol, maringenin, dihydrokaempferol, dihydroquercetin, non-receptor molecule or step associated with cycling of the catechin, epicatechin, epigallocatechin, cyanidin, delphini target receptor, to disrupt the cycling of the target receptor; din, malvidin, pelargonidin, peonidin, and petunidin. wherein the first and second agents in combination cooperate In a further embodiment, the second agent is an organosul to amplify the disrupted or altered function resulting from the fide. binding of the target receptor by the first agent, resulting in In a further aspect of this embodiment, the second agent is synergistic control of the pest. selected from the group consisting of allicin, glutathione, Further embodiments of the present invention provide indole-3-carbinol, an isothiocyanate, and Sulforaphane. compositions for controlling a target pest including a pest In a further embodiment, the second agent is a phenolic 10 control product and at least one active agent, wherein: the acid. active agent can be capable of interacting with a receptor in In a further aspect of this embodiment, the second agent is the target pest, the pest control product can have a first activity selected from the group consisting of capsaicin, ellagic acid, against the target pest when applied without the active agent gallic acid, roSmarinic acid, and tannic acid. and the compositions can have a second activity against the In a further embodiment, the secondagent is a phytosterol. 15 target pest, and the second activity can be greater than the first In a further aspect of this embodiment, the second agent is activity. The first and second activities can be quantified by selected from the group consisting of beta-sitosterol and measuring concentration of the pest control product effective Saponins. to control the target pest, and a concentration corresponding In a further embodiment, the second agent is selected from to the first activity can be higher than a concentration corre the group consisting of a damnacanthal, a digoxin and a sponding to the second activity. The first and second activities phytic acid. can be quantified by measuring disablement effect of the In a further embodiment, the target pest is a species belong target pest at a standard concentration of pest control product, ing to the order Rhabditida. and the compositions exhibit a greater disablement effect than In a further embodiment, the target pest is a roundworm. the pest control product applied without the active agent. The In a further embodiment, the target pest is a species belong 25 first activity can persist for a first period, the second activity ing to the order Diptera. can persist for a second period, and the second period can be In a further embodiment, the target pest is a species belong longer than the first period. The active agent can include a ing to an order selected from the group consisting of Acari, synergistic combination of at least two receptor ligands. The Diptera, Hemiptera, and Thysanoptera. second activity can reflect a synergistic interaction of the In a further embodiment, the target pest is a species belong 30 active agent and the pest control product. ing to an order selected from the group consisting of Blattaria The target pest can be selected from the group consisting of and Thysanoptera. a fungus, a plant, an animal, a moneran, and a protist. The A further embodiment provides a method of controlling a target pest can be an arthropod species, such as, for example, target pest, comprising administering an effective amount of an insect, an arachnid, or an arachnoid. The target pest can be a composition in accordance with one of the embodiments 35 a species belonging to an animal order selected from: Acari, described above to the target pest or a Substrate associated Anoplura, Araneae, Blattodea, Coleoptera, Collembola, with the target pest, thereby achieving synergistic pest con Diptera, Grylloptera, Heteroptera, Homoptera, trol. Hymenoptera, Isopoda, Isoptera, Lepidoptera, Mantodea, In a further aspect of this embodiment, the substrate is Mallophaga, Neuroptera, Odonata, Orthoptera, Psocoptera, selected from a group consisting of a crop, a plant, a surface, 40 Siphonaptera, Symphyla, Thysanura, and Thysanoptera. and a vertebrate animal. The pest control product can be a chlorphenoxy compound A method for controlling a target pest, comprising apply such as, for example, 2,4-D Amine and/or 2.4D IBE. Like ing to the target pest or to a substrate associated with the target wise, the pest control product can be a carbamate Such as, for pest a composition comprising a first agent and a second example, methomyl, carbofuran, carbaryl, BPMC, carbenda agent, wherein the first agent has a first activity against the 45 Zim, carbosulfan, captain hydrochloride, and/or cartap. The target pest, the second agent has a second activity against the pest control product can be an organophosphate Such as, for target pest, the composition has a third activity against the example, acephate, malathion, diazinon, chlorpy firos, target pest, and the third activity reflects a synergistic inter fenoxycab, edifenphos, febuconazole, chlorphenapyr, mag action of the first agent and the second agent, thereby result nesium phosphide, metamidophos, and/or fenitrothion. The ing in Synergistic pest control. 50 pest control product can be an organochlorine Such as, for A further embodiment provides a method of pest control example, DDT. DDE, and/or heptachlorepoxide. The pest comprising applying a composition to a target pest or to a control product can be a pyrethroid such as, for example, Substrate associated with a target pest, wherein the composi cypermethrin, cynmethylin+2,4-D IBE, lambdacyhalothrin, tion comprises a first agent comprising at least one receptor dazomet, cyfluthrin, betacy permethrin, pendimethlin, per ligand and a second agent comprising a pesticide, and 55 methrin, deltamethrin, bifenethrin, alphacypermethrin, fen wherein the pest control comprises affecting a physiological Valerate, propanil, and/or esfenvalerate. The pest control condition of the pest associated with a function of the pesti product can be a neonicotinoid such as, for example, thi cide while also affecting a function of the receptor associated omethoxam, fipronil, clothianidin, and/or imidacloprid. The with the receptor ligand, thereby resulting in Synergistic pest pest control product can include at least one of an avermectin, control. 60 abamectin, spinosad, fluxastrobin, and/or indoxacarb. The A further embodiment provides a method of pest control pest control product can be a botanical product such as, for comprising the steps of providing a target pest having at least example, rotenone, nicotine, caffeine, a pyrethrum, an essen one target GPCR receptor, contacting the target pest with a tial oil, and/or a fixed oil. The pest control product can be a composition comprising at least a first agent and a second fungicide, a nematicide, an insecticide, an acaricide, and/or a agent, wherein the first agent is capable of interacting with the 65 bactericide. target receptor to trigger, disrupt or alter a biological function The receptor can be a G protein-coupled receptor (GPCR), related to the binding of the target receptor with the first agent, such as a GPCR of the insect olfactory cascade, such as, for US 8,691,256 B2 7 8 example, a tyramine receptor, an olfactory receptor Ora3a, an Also provided in Some embodiments are pest-control com olfactory receptor Or83b and/or an octopamine receptor. positions exhibiting high potency against an invertebrate tar Binding of the receptor by an ingredient of the compositions get pest and low toxicity against a vertebrate animal, the can result in a change in intracellular level of cAMP and/or compositions including a synergistic combination of active calcium, wherein the change can be sufficient to permit con agents, wherein each active agent interacts with a molecular trol of the target pest. target with high affinity in the target pest and that can be Control can include a condition Such as, for example, kill absent form, or present with low affinity, from the vertebrate. ing, knockdown, repellency, interference with reproduction, The at least one active agent can be a ligand of a selected interference with feeding, and interference with a stage of a GPCR, and the at least one active agent is preferably not a 10 ligand of the selected GPCR. The high target potency and low life cycle of the target pest. vertebrate toxicity can be expressed as a ratio of LD50(target) Embodiments of the invention also include a crop pro versus LD50(vertebrate animal), and wherein the ratio can be tected by the compositions disclosed herein. less than 100:1. In addition, embodiments of the invention can include In some embodiments, the invention provides methods of compositions for controlling a target pest including a pest 15 pest control including contacting a target pest with a compo control product and at least one active agent, wherein: the sition as described herein, resulting in control of the pest. The active agent can include a ligand of a GPCR of a target pest, methods can include applying a composition to a target pest wherein binding of the ligand to the GPCR can cause a change or to a substrate associated with a target pest, wherein the in a level of cAMP or calcium that can permit control of the compositions can include a pesticide and an active agent target pest; the pest control product can have a first activity including at least one receptor ligand, and wherein the pest against the target pest, the active agent can have a second control can include affecting a physiological condition of the activity against the target pest, and the compositions can have pest associated with a function of the pesticide while also a third activity against the target pest; and the third activity affecting a function of the receptor associated with the recep can be greater than the first activity or the second activity. The tor ligand. The binding of the receptor by an ingredient of the active agent can include a synergistic combination of at least 25 compositions can result in a change in intracellular level of two GPCR ligands. The third activity can be indicative of cAMP and/or calcium, and wherein the change can be suffi synergy between the active agent and the pest control product. cient to permit control of the target pest. The pesticide can be In some embodiments, compositions can include at least two selected from a chlorphenoxy compound, a carbamate, an active ingredients, wherein at least one active ingredient organophosphate, an organochlorine, a pyrethroid, a neoni interacts with a G protein-coupled receptor (GPCR) of the 30 cotinoid, a botanical product, a fungicide, a nematicide, and pest and wherein at least one active ingredient does not inter insecticide, and acaracide, a bactericide. and an avermectin. act with the GPCR, and wherein the at least two active ingre The substrate can be, for example, a crop plant and/or a soil. dients in combination have a synergistic pest-control activity. The target pest can be, for example, a fungus, a plant, an The pest can be an insect and the GPCR can be associated animal, a moneran, or a protist. The use of the compositions with olfaction, and further the GPCR preferably can be absent 35 can permitan improvement of control of the pest as compared from vertebrate animals. The synergistic pest-control activity with use of the pesticide alone or the active agent alone. The can have a coefficient of synergy in excess of 1.5. The Syner improvement can include a synergistic interaction of the pest gistic pest-control activity can exceed additive effects of the control product with the active agent. The improvement can active ingredients, as measured by the Colby calculation of include an improved result with use of a substantially similar synergy. The GPCR can have a high affinity for the active 40 amount of the pest control product. The improved result can ingredient in a target organism and the GPCR can be absent or be at least one of increased killing of the target pest: can have a low affinity for the active ingredient in a non-target increased interference with reproduction by the target pest: organism. The non-target organism can be a vertebrate ani and prolonged effectiveness of the pest control product. The mal. In some embodiments, the target organism can be a improvement can include a Substantially similar result with plant, an animal, a fungus, a protist, or a moneran, and the 45 use of a Substantially lower amount of the pest control product non-target organism can be selected from a crop plant, a and/or the active agent. Use of the compositions permits an Vertebrate animal, and a non-pest invertebrate. agricultural improvement such as, for example, increased In some embodiments, the invention provides low-resis crop yield; reduced frequency of application of pest control tance pest-control compositions, including at least a first product; reduced phytotoxicity associated with the pesticide; active ingredient and a second active ingredient, wherein the 50 and reduced cost or increased value associated with at least first active ingredient interacts with a first molecular target one environmental factor. The environmental factor can under genetic control within a selected pest, and wherein the include, for example, air quality, water quality, Soil quality, second active ingredient interacts with a second molecular detectable pesticide residue, safety or comfort of workers: target under genetic control within the selected pest, and and a collateral effect on a non-target organism. wherein the ingredients in the compositions act together in a 55 Also provided are methods of developing a compositions complementary manner upon the target pest, and wherein for pest control, including: providing a cell line expressing at resistance to the compositions in an individual target pest least one of a tyramine receptor, an olfactory receptor Ora3a, requires two separate genetic lesions divergent from a non oran olfactory receptor Or83b, wherein binding of a ligand to resistant population of the pest. The first and second molecu any of the receptors causes a change in a level of intracellular lar targets can include two separate molecules encoded or 60 cAMP or calcium, and the change can be indicative of a controlled by separate genetic elements. The complementary potential for invertebrate pest control; contacting the cell with manner can include an additive effect of each agent acting a candidate ligand; detecting a change in the level of cAMP separately, or the complementary manner can include a syn and/or calcium in the cell; identifying the candidate ligand as ergistic effect as compared with each agent acting separately. an active compound for control of an invertebrate pest; and The first molecular target can be a GPCR, and the second 65 combining the active compound with a pesticide to form a molecular target is preferably not the same as the first molecu composition for pest control, wherein the pesticide does not lar target. bind to a receptor bound by the active compound, and wherein US 8,691,256 B2 10 a combined effect of the active compound and the pesticide target pest, including administering a pest-control composi can include an effect against a target pest that can be greater tion, the composition including at least a first active ingredi than the effect of either the active compound alone or the ent and a second active ingredient, wherein the first active pesticide alone. The compositions further can include a sec ingredient interacts with a first molecular target under genetic ond active compound capable of binding at least one of the control within a selected pest, and wherein the second active receptors. The active compounds can cooperate to cause a ingredient interacts with a second molecular target under synergistic change in the level of cAMP and/or calcium in the genetic control within the selected pest, and wherein the cell line and/or in a target pest. The combined effect of the ingredients in the composition act together in a complemen active compound and the pesticide can be synergistic. The tary manner upon the target pest, and wherein resistance to the combined effect can be determined by at least one condition 10 composition in an individual target pest requires two separate selected from the group consisting of killing, knockdown, genetic lesions divergent from a non-resistant population of repellency, interference with reproduction, interference with the pest. feeding, and interference with a stage of a life cycle of the Still other embodiments provide pest control compositions target pest. exemplified by the following: in combination, a blend of lilac Also provided are further methods of pest control, includ 15 flower oil (LFO), d-limonene, thyme oil, and further includ ing, providing a composition including at a first and a second ing a pesticide. The pesticide can be, for example, clothiani active ingredient, wherein the first active ingredient interacts din. The blend can include 10-80% LFO, 5-60% d-limonene, with a receptor of a target pest, and wherein the second active and 10-80% thyme oil. In other embodiments, the blend can ingredient can be a pesticide that does not interact with the include 20-60% LFO, 10-45% d-limonene, and 20-60% receptor of the first active ingredient; and contacting the pest thyme oil. In other embodiments, blend can include 42.6% with the compositions, wherein the contacting results in Syn w/w LFO, 27.35% w/w d-limonene, and 30.08% w/w thyme ergistic pest control. The compositions further can include a oil white. third active ingredient, wherein the third active ingredient In certain embodiments, the invention can include a interacts with a receptor of the target pest, and wherein at least method of pest control including the steps of providing a the first and third active ingredients in combination synergis 25 target pest having at least one target GPCR receptor, contact tically interact to permit control of the target pest. The first ing the target pest with a composition comprising at least a and third active ingredients can optionally bind the same first active agent and a second active agent, wherein the first receptor; in other embodiments, the first and third active active agent is capable of interacting with the target receptor ingredients do not bind the same receptor. The first, second, to trigger, disrupt or alter a biological function related to the and third active ingredients in combination can have a syner 30 binding of the target receptor with the first active agent, and gistic effect that can be greater than the effect of any single wherein the second active agent is capable of interacting with ingredient and can be also greater than the synergistic effect a non-receptor molecule or step associated with cycling of the of the first and third ingredients in combination. The receptor target receptor, to disrupt the signaling cascade of the target can be a GPCR such as, for example, a tyramine receptor, an receptor. In some embodiments the active agents in combi olfactory receptor Or43a, and an olfactory receptor Or83b. 35 nation can cooperate to amplify the disrupted or altered func The pest control can be associated with a receptor-activated tion resulting from the binding of the target receptor by the alteration in a level of cAMP and/or calcium within the pest. first active agent, resulting in control of the pest. In some The alteration can persist for at least about 60 seconds. embodiments of the invention, the composition can include a Also provided are other methods of pest control, including: third active agent, and the third active agent can be capable of providing a composition including at least two active ingre 40 interacting with a GPCR receptor in the target pest, and the dients, wherein at least one active ingredient interacts with a interaction can be complementary to the action of the first GPCR of a target pest, the composition produces a first level active agent. of at least one of intracellular calcium and cyclic AMP in a In some embodiments, the first and third active agents can cell expressing the GPCR on exposure to the cell, and the first interact with the same receptor, or different receptors. In some level can be higher than a second level produced when the cell 45 embodiments the complementarity of the first and third active can be contacted with any single active ingredient; and con agents can cause an additive effect of the active agents tacting the pest with the compositions, wherein the contacting together as compared with an effect of each active agent results in Synergistic pest control. Other embodiments pro separately. In some embodiments the complementarity of the vide methods for controlling a target pest including use of a first and third active agents can cause a synergistic effect of pest control compositions, the compositions including a pest 50 the active agents together as compared with an effect of each control product and at least one active agent, wherein: the active agent separately. In some embodiments, receptor active agent can include a ligand of a GPCR of a target pest, cycling can include at least one of receptor sensitization, wherein binding of the ligand to the GPCR causes a change in receptor desensitization, receptor recycling, ligand release, a level of cAMP or calcium that permits control of the target receptor phosphorylation, and receptor dephosphorylation. pest; the pest control product can have a first activity against 55 Embodiments of the invention can include a pest-control the target pest, the active agent can have a second activity composition that has a first active agent capable of disrupting against the target pest, and the compositions can have a third or altering a function of a receptor in a target pest, and a activity against the target pest, and the third activity can be second active agent capable of disrupting cycling of the greater than the first activity or the second activity. A further receptor, and the second active agent can act to amplify an method of pest control can include use of a pest control 60 effect of the first active agent. composition, wherein the composition can include at least Embodiments of the invention can include a method of two active ingredients, wherein at least one active ingredient making a pest control composition with the steps of provid interacts with a G protein-coupled receptor (GPCR) of the ing a target pest having at least one target receptor, contacting pest and wherein at least one active ingredient does not inter the target pest with a composition including at least a first act with the GPCR, and wherein the at least two active ingre 65 active agent and a second active agent, wherein the first active dients in combination have a synergistic pest-control activity. agent is capable of interacting with the target receptor to Other methods of pest control can permit low-resistance in a disrupt or alter a function related to normal activity of the US 8,691,256 B2 11 12 target receptor, and wherein the second active agentis capable FIG. 2 shows the binding of a ligand to a biogenic amine of interacting with a non-receptor molecule or step associated receptor, resulting in downstream signaling affecting certain with cycling of the target receptor, to disrupt the cycling of the physiological responses; target receptor. Some embodiments can include measuring FIG. 3 shows an insect control chemical, deltamethrin the effect of the composition upon the target pest and select (DM), affecting downstream signaling; ing the at least a first active agent based on the desired prop FIG. 4A shows a pesticidal effect against Aedes aegypti erties of the composition. Some embodiments of the inven caused by 1) a test composition; 2) clothianidin; and 3) a tion can include a third active agent, wherein the third active combination of a test composition and clothianidin; agent can be capable of interacting with a receptor in the FIG. 4B shows a pesticidal effect against Aedes aegypti target pest, and wherein the interaction can be complemen 10 caused by 1) a test composition; 2) clothianidin; and 3) a tary to the action of the first active agent. In some embodi combination of a test composition and clothianidin; ments, the first and third active agents can interact with a same FIG. 4C shows a pesticidal effect against Aedes aegypti receptor, or with a different receptor. In some embodiments caused by 1) a test composition; 2) imidacloprid; and 3) a the complementarity of the first and third active agents can combination of a test composition and imidacloprid; cause an additive effect of the active agents together as com 15 FIG. 4D shows a pesticidal effect against Drosophila sp. pared with an effect of each active agent separately. Likewise, caused by 1) a test composition; 2) imidacloprid; and 3) a the complementarity of the first and third active agents can combination of a test composition and imidacloprid; cause a synergistic effect of the active agents together as FIG. 5 shows a pesticidal effect against Aedes aegypti compared with an effect of each active agent separately. caused by 1) a test composition; 2) imidacloprid; and 3) a Embodiments of the invention can include a pest-control combination of a test composition and imidacloprid; composition that has an active agent capable of disrupting or FIG. 6A shows a pesticidal effect against Periplaneta altering cycling of a GPCR in a target pest, wherein the active americana caused by 1) a test composition; 2) clothianidin; agent acts to amplify an effect of a ligand binding the GPCR. and 3) a combination of a test composition and clothianidin; Further, the amplification can result in a prolonged intracel FIG. 6B shows a pesticidal effect against Periplaneta lular Ca2+ cascade as compared with the Ca2+ cascade that 25 americana caused by 1) a test composition; 2) imidacloprid; occurs when the receptor is bound without the presence of the and 3) a combination of a test composition and imidacloprid; active agent. In some embodiments, the amplification can FIG. 7 shows a pesticidal effect against bedbugs caused by result in a prolonged perturbation of intracellular cAMP level 1) a test composition; 2) pyrethrum; and 3) a combination of as compared with the perturbation in cAMP level that occurs a test composition and pyrethrum; when the receptor is bound without the presence of the active 30 FIG. 8A shows the nucleic acid sequence and the peptide agent. sequence of a Tyramine receptor; Embodiments of the invention can include a pest-control FIG. 8B shows the nucleic acid sequence and the peptide composition that has an active agent capable of disrupting or sequence of a Tyramine receptor; altering cycling of a GPCR in a target pest, wherein the active FIG. 9 shows dose dependent pesticidal effects (knock agent acts to attenuate an effect of a ligand binding the GPCR. 35 down and mortality) of imidacloprid against thrips overtime. Certain embodiments of the invention can include a FIG.10A shows a dose dependent pesticidal effect (knock method of pest control including the steps of providing a down) against thrips caused by Blend 11; target pest having at least one target GPCR receptor, contact FIG. 10B shows a dose dependent pesticidal effect (mor ing the target pest with a composition including at least a first tality) against thrips caused by Blend 11: active agent and a second active agent, wherein the first active 40 FIG. 10C shows a dose dependent pesticidal effect (knock agent can be capable of interacting with the target receptor to down) against thrips caused by Blend 8: trigger, disrupt or alter a biological function related to the FIG. 10D shows a dose dependent pesticidal effect (mor binding of the target receptor with the first active agent, and tality) against thrips caused by Blend 8. wherein the second active agent can be capable of interacting FIG. 11A shows shows a dose dependent pesticidal effect with a non-receptor molecule or step associated with the 45 (knockdown) against thrips caused by a composition contain biological pathway triggered, disrupted, or altered as a result ing the mixture of Blend 11 and imidacloprid; of the first active agents interacting with the target receptor, FIG. 11B shows shows a dose dependent pesticidal effect wherein the active agents in combination can cooperate to (mortality) against thrips caused by a composition containing amplify the disrupted or altered function resulting from the the mixture of Blend 11 and imidacloprid; binding of the target receptor by the first active agent, result 50 FIG. 11C shows a dose dependent pesticidal effect (knock ing in control of the pest. down) against thrips caused by a composition containing the In various embodiments, the composition can further mixture of Blend 8 and imidacloprid; include a third active agent, wherein the third active agent can FIG. 11D shows shows a dose dependent pesticidal effect be capable of interacting with a GPCR receptor in the target (mortality) against thrips caused by a composition containing pest, and wherein the interaction can be complementary to the 55 the mixture of Blend 8 and imidacloprid; action of the first active agent. Likewise, the first and third FIG. 12 shows dose dependent pesticidal effects (knock active agents can interact with a same receptor, or with dif down and mortality) of imidacloprid against thrips overtime. ferent receptors. Additionally, the complementarity of the FIG. 13 shows dose dependent pesticidal effects (knock first and third active agents can cause an additive effect of the down and mortality) of Blend 38 (labeled “B5049”) against active agents together as compared with an effect of each 60 thrips over time. active agent separately. FIG. 14 shows dose dependent pesticidal effects (knock down and mortality) against thrips over time caused by a BRIEF DESCRIPTION OF THE DRAWINGS composition containing the mixture of imidacloprid and Blend 38 (labeled “B5049” and “TT 1A). FIG. 1 shows a screening method using a transfected cell 65 FIG. 15 shows dose dependent pesticidal effect against lines expressing a receptor of interest, for example, a biogenic thrips over time caused by 1) water control or imidacloprid amine receptor, Such as, a TyR or an octopamine receptor, diluted in water; 2) a test composition; and 3) a combination US 8,691,256 B2 13 14 ofa test composition and imidacloprid. The test compositions FIG.22 shows fluorescence intensity curves corresponding are Blend 38 (labeled “B5049” and “TT 1A) and Blend 39 to intracellular calcium ion concentrations, with the curve (labeled “B5053” and “TT3C). corresponding to the compositions containing 1) clothianidin FIG.16A shows fluorescence intensity curves correspond alone, 2) a test composition alone, and 3) the mixture of ing to intracellular calcium ion concentrations, with the curve 5 clothianidin and the test composition. The test composition is corresponding to the compositions containing 1) imidaclo Blend 19. Clothianidin is labeled as AMP Agent. prid alone, 2) Blend 19 alone, and 3) the mixture of imida FIG. 23A shows fluorescence intensity curves correspond cloprid and Blend 19; ing to intracellular calcium ion concentrations, with the curve FIG. 16B shows fluorescence intensity curves correspond corresponding to the compositions containing 1) abamectin ing to intracellular calcium ion concentrations, with the curve 10 alone, 2) a test composition alone, and 3) the mixture of corresponding to the compositions containing 1) imidaclo abamectin and the test composition. The test composition is prid alone, 2) Blend 27 alone, and 3) the mixture of imida Blend 7. cloprid and Blend 27: FIG. 23B shows fluorescence intensity curves correspond FIG.16C shows fluorescence intensity curves correspond 15 ing to intracellular calcium ion concentrations, with the curve ing to intracellular calcium ion concentrations, with the curve corresponding to the compositions containing 1) imidaclo corresponding to the compositions containing 1) imidaclo prid alone, 2) a test composition alone, and 3) the mixture of prid alone, 2) Blend 75 alone, and 3) the mixture of imida imidacloprid and the test composition. The test composition cloprid and Blend 75. is Blend 7. FIG.17A shows fluorescence intensity curves correspond FIG.24A shows fluorescence intensity curves correspond ing to intracellular calcium ion concentrations, with the curve ing to intracellular calcium ion concentrations, with the curve corresponding to the compositions containing 1) imidaclo corresponding to the compositions containing 1) yohimbine prid alone, 2) Blend 7 alone, and 3) the mixture of imidaclo alone, 2) forskolin alone, and 3) genistein alone. prid and Blend 7: FIG.24B shows schematic molecular structures of yohim FIG. 17B shows fluorescence intensity curves correspond 25 bine, forskolin and genistein. ing to intracellular calcium ion concentrations, with the curve FIG. 25A shows fluorescence intensity curves correspond corresponding to the compositions containing 1) imidaclo ing to intracellular calcium ion concentrations, with the curve prid alone, 2) Blend 15 alone, and 3) the mixture of imida corresponding to the compositions containing 1) a test com cloprid and Blend 15; position alone, and 2) the mixture of yohimbine and the test FIG. 17C shows fluorescence intensity curves correspond 30 composition. The test composition is Blend 2, and labeled as ing to intracellular calcium ion concentrations, with the curve “B5028. corresponding to the compositions containing 1) imidaclo FIG.25B shows fluorescence intensity curves correspond prid alone, 2) Blend 11 alone, and 3) the mixture of imida ing to intracellular calcium ion concentrations, with the curve cloprid and Blend 11: corresponding to the compositions containing 1) a test com FIG. 17D shows fluorescence intensity curves correspond 35 position alone, and 2) the mixture of forskolin and the test ing to intracellular calcium ion concentrations, with the curve composition. The test composition is Blend 2, and labeled as corresponding to the compositions containing 1) imidaclo “B5028. prid alone, 2) Blend 11 alone, and 3) the mixture of imida FIG. 25C shows fluorescence intensity curves correspond cloprid and Blend 11. ing to intracellular calcium ion concentrations, with the curve FIG. 18A shows fluorescence intensity curves correspond 40 corresponding to the compositions containing 1) a test com ing to intracellular calcium ion concentrations, with the curve position alone, and 2) the mixture of genistein and the test corresponding to the compositions containing 1) imidaclo composition. The test composition is Blend 2, and labeled as prid alone, 2) Blend 38 alone, and 3) the mixture of imida “B5028. cloprid and Blend 38. Blend 38 is labeled as “B5049”. FIG. 26A shows fluorescence intensity curves correspond FIG. 18B shows fluorescence intensity curves correspond 45 ing to intracellular calcium ion concentrations, with the curve ing to intracellular calcium ion concentrations, with the curve corresponding to the compositions containing 1) a test com corresponding to the compositions containing 1) imidaclo position alone, and 2) the mixture of yohimbine and the test prid alone, 2) Blend 39 alone, and 3) the mixture of imida composition. The test composition is Blend 39, and labeled as cloprid and Blend 39. Blend 39 is labeled as “B5053”. “B5053. FIG. 19 shows fluorescence intensity curves corresponding 50 FIG. 26B shows fluorescence intensity curves correspond to intracellular calcium ion concentrations, with the curve ing to intracellular calcium ion concentrations, with the curve corresponding to the compositions containing 1) fipronil corresponding to the compositions containing 1) a test com alone, 2) a test composition alone, and 3) the mixture of position alone, and 2) the mixture of forskolin and the test fipronil and the test composition. The test composition is composition. The test composition is Blend 39, and labeled as Blend 19. 55 “B5053. FIG. 20 shows fluorescence intensity curves corresponding FIG. 26C shows fluorescence intensity curves correspond to intracellular calcium ion concentrations, with the curve ing to intracellular calcium ion concentrations, with the curve corresponding to the compositions containing 1) fipronil corresponding to the compositions containing 1) a test com alone, 2) a test composition alone, and 3) the mixture of position alone, and 2) the mixture of genistein and the test fipronil and the test composition. The test composition is 60 composition. The test composition is Blend 39, and labeled as Blend 42, and labeled as “AAT. “B5053. FIG.21 shows fluorescence intensity curves corresponding FIG. 27A shows fluorescence intensity curves correspond to intracellular calcium ion concentrations, with the curve ing to intracellular calcium ion concentrations, with the curve corresponding to the compositions containing 1) fipronil corresponding to the compositions containing 1) a test com alone, 2) a test composition alone, and 3) the mixture of 65 position alone, and 2) the mixture of genistein and the test fipronil and the test composition. The test composition is composition. The test composition is Blend 41, and labeled as Blend 39, and labeled as “B5053. AAL. US 8,691,256 B2 15 16 FIG. 27B shows fluorescence intensity curves correspond FIG.32 shows fluorescence intensity curves corresponding ing to intracellular calcium ion concentrations, with the curve to intracellular calcium ion concentrations, with the curve corresponding to the compositions containing 1) a test com corresponding to the composition containing the mixture of position alone, and 2) the mixture of genistein and the test fluoxastrobin and thyme oil indicated by triangles, the curve composition. The test compositionis Blend 42, and labeled as corresponding to the composition containing the thyme oil “AAT. alone indicated by Squares, and the curve corresponding to FIG. 28A shows fluorescence intensity curves correspond the composition containing fluoxastrobin alone indicated by ing to intracellular calcium ion concentrations, with the curve circles. corresponding to the compositions containing 1) a test com position alone, and 2) the mixture of yohimbine and the test 10 DESCRIPTION OF EXEMPLARY composition. The test composition is Blend 27, and labeled as EMBODIMENTS “B7001. Many previously known and commercialized products FIG. 28B shows fluorescence intensity curves correspond having sufficient pesticidal activity to be useful also have ing to intracellular calcium ion concentrations, with the curve 15 toxic or deleterious effects on mammals, fish, fowl, or other corresponding to the compositions containing 1) a test com non-target species. For example, common insecticides Such position alone, and 2) the mixture of forskolin (labeled “FK”) as organophosphorus compounds and carbamates inhibit the and the test composition. The test composition is Blend 27. activity of acetylcholinesterase in all classes of animals. and labeled as “B7001. Chlordimeformand related formamidines are knownto action FIG. 28C shows fluorescence intensity curves correspond insect octopamine receptors, but have been removed from the ing to intracellular calcium ion concentrations, with the curve market because of cardiotoxic potential in vertebrates and corresponding to the compositions containing 1) a test com carcinogenicity in animals and a varied effect on different position alone, and 2) the mixture of genistein and the test insects. composition. The test composition is Blend 27, and labeled as However, the deleterious effects of many pesticides can be “B7001. 25 mitigated by reducing the amount of pesticide that can be FIG. 29.A shows fluorescence intensity curves correspond applied to a given area to achieve the desired result. This ing to intracellular calcium ion concentrations, with the curve reduction can be achieved by combining the pesticidal com corresponding to the compositions containing 1) a test com pound or product with selected active ingredients. These position alone, and 2) the mixture of yohimbine and the test active ingredients can comprise, for example, plant essential composition. The test composition is Blend 58, and labeled as 30 oils, and the like. Combinations of selected active ingredients “B7002. with selected pesticidal compounds or products can reduce FIG.29B shows fluorescence intensity curves correspond the concentration of pesticide needed to achieve a net effi ing to intracellular calcium ion concentrations, with the curve ciency, and extend the useful life of existing synthetic pesti corresponding to the compositions containing 1) a test com cides. position alone, and 2) the mixture of forskolin (labeled “FK”) 35 The details of one or more embodiments of the invention and the test composition. The test composition is Blend 58, are provided. Modifications to embodiments described in this and labeled as “B7002. document, and other embodiments, will be evident to those of FIG. 29C shows fluorescence intensity curves correspond ordinary skill in the art after a study of the information pro ing to intracellular calcium ion concentrations, with the curve vided in this document. The information provided in this corresponding to the compositions containing 1) a test com 40 document, and particularly the specific details of the position alone, and 2) the mixture of genistein and the test described exemplary embodiments, is provided primarily for composition. The test composition is Blend 58, and labeled as clearness of understanding and no unnecessary limitations “B7002. are to be understood therefrom. FIG. 30A shows shows fluorescence intensity curves cor Embodiments of the invention are directed to methods of responding to intracellular calcium ion concentrations, with 45 screening compositions for pest control potential, composi the curve corresponding to the compositions containing 1) a tions for controlling pests, and methods for using these com test composition alone, and 2) the mixture of yohimbine and positions. the test composition. The test composition is anise oil. As used herein, "pests’ can mean any organism whose FIG. 30B shows shows fluorescence intensity curves cor existence it can be desirable to control. Pests can include, for responding to intracellular calcium ion concentrations, with 50 example, bacteria, cestodes, fungi, insects, nematodes, para the curve corresponding to the compositions containing 1) a sites, plants, and the like. test composition alone, and 2) the mixture of forskolin (la As used herein, "pesticidal can mean, for example, anti beled “FK”) and the test composition. The test composition is bacterial, antifungal, antiparasitic, herbicidal, insecticidal, anise oil. and the like. FIG.30C shows shows fluorescence intensity curves cor 55 Furthermore, G protein uncoupling in response to phos responding to intracellular calcium ion concentrations, with phorylation by both second messenger-dependent protein the curve corresponding to the compositions containing 1) a kinases and G protein-coupled receptor kinases (GRKS) leads test composition alone, and 2) the mixture of genistein and the to GPCR desensitization. GRK-mediated receptor phospho test composition. The test composition is anise oil. rylation promotes the binding of beta-arrestins, which in FIG.31 shows fluorescence intensity curves corresponding 60 addition to uncoupling receptors from heterotrimeric G pro to intracellular calcium ion concentrations, with the curve teins, also target many GPCRs for internalization inclathrin corresponding to the composition containing the mixture of coated vesicles. Beta-arrestin proteins play a dual role in imidacloprid and thyme oil indicated by triangles, the curve regulating GPCR responsiveness by contributing to both corresponding to the composition containing the thyme oil receptor desensitization and internalization. alone indicated by circles, and the curve corresponding to the 65 Following desensitization, GPCRs can be resensitized. composition containing imidacloprid alone indicated by GPCR sequestration to endosomes is thought to be the Squares. mechanism by which GRK-phosphorylated receptors are US 8,691,256 B2 17 18 dephosphorylated and resensitized. The identification of (2007) Role of diacylglycerol kinase in cellular regulatory beta-arrestins as GPCR trafficking molecules suggested that processes: a new regulator for cariomyocyte hypertrophy, beta-arrestins can be determinants for GPCR resensitization. Pharmacol Ther 115:352-359; Dalrymple (2008) G protein However, other cellular components also play pivotal roles in coupled receptor dimers: functional consequences, disease the de- and re-sensitization (D/R) process, including, for 5 states and drug targets, Pharmacol Ther 118:359-371; example, GRK, N-ethylmaleimide-sensitive factor (NSF), Jurado-Pueyo (2008) GRK2-dependent desensitization clathrin adaptor protein (AP-2 protein), protein phosphatases, downstream of G proteins, Recept Signal Transduct Res clathrin, and the like. In addition to these molecules, other 28:59-70; Milligan (1998) New aspects of g-protein-coupled moieties such as, for example, endoSomes, lysosomes, and receptor signalling and regulation, Trends Endocrinol Metab the like, also influence the D/R process. These various com 10 ponents of the D/R cycle provide opportunities to disrupt or 9:13-19. Each of the foregoing is incorporated by reference in alter GPCR “availability” to extracellular stimuli, and thus its entirety for its disclosure of the mechanisms and compo attenuate or intensify the effect of those extracellular stimuli nents of GPCR signaling, cycling, regulation, and the like. upon target organisms. Attenuation, achieved, for example, Embodiments of the invention can include a method to by inhibition of the resensitization process, or the like, can 15 disrupt or alter GPCR D/R by altering or disrupting the vari limit the effects of extracellular stimuli (such as, for example, ous signal cascades triggered through GPCR action. Certain UV exposure, toxins, or the like) on the GPCR signaling embodiments can disrupt or alter GPCRD/R in various ways, process. Intensifying a signal cascade, achieved, for example, including, for example, the application of compositions con by inhibition of the desensitization process, or the like, can taining active agents such as, for example, essential oils, and increase the effects of extracellular stimuli (such as, for the like. example, pharmaceuticals, insecticides, or the like) on the Screening of Compositions GPCR signaling process. In some embodiments of the invention, the screening Components of the GPCR signaling process have been the method for pest control potential can target a molecule of an object of significant study; opportunities and targets for dis insect olfactory receptor protein. In some embodiments of the ruption of the signaling process according to the present 25 invention, the screening method for pest control potential can invention are numerous. Discussions of the components of target an insect olfactory receptor protein. The insect olfac G-protein signaling are provided inYu (2006) Heterotrimeric tory system includes more than 60 identified olfactory recep G protein signaling and RGSS in Aspergillus nidulans, J tors. These receptors are generally members of a large family Microbiol 44:145-154; Dong (2007) Regulation of G protein of G protein coupled receptors (GPCRs). coupled receptor export trafficking, Biochim Biophy's Acta 30 As used herein, a “receptor is an entity on the cell mem 1768:853-870; Nakahata (2007) Regulation of G Protein brane or within the cell, cytoplasm, or cell nucleus that can coupled Receptor Function by Its Binding Proteins, Yakugaku bind to a specific molecule (a ligand), such as, for example, a Zasshi 127:3-14; Yang (2006) Mechanisms of regulation and neurotransmitter, hormone, or the like, and initiates the cel function of G-protein-coupled receptor kinases, World J. Gas lular response to the ligand. Ligand-induced changes in the troenterol 12:7753-7757: Ma (2007) Beta-arrestin signaling 35 behavior of receptor proteins can result in physiological and regulation of transcription: J Cell Sci 120:213-218: New changes that constitute the biological actions of the ligands. (2007) Molecular mechanisms mediating the G protein In accordance with the present disclosure, receptors such coupled receptor regulation of cell cycle progression, J Mol as G protein-coupled receptors may be classified on the basis Signaling 2:2-16; Klasse (2008) Internalization and desensi of binding affinity of the receptor to an active ingredient. This tization of adenosine receptors, Purinergic Signalling 4:21 40 may also be expressed as the binding affinity of the active 37; Stewart (2007) Phospholipase C-eta Enzymes as Putative ingredient for the receptor. The binding affinity of an active Protein Kinase C and Ca2+ Signalling Components in Neu ingredient for a receptor, or the binding affinity of a receptor ronal and Neuroendocrine Tissues, Neuroendocrinology for an active ingredient, may be measured in accordance with 86:243-248; Xu (2007) Regulation of G protein-coupled methods disclosed herein or methods known to those of skill receptor trafficking, Acta Physiol 190:39–45: Wolfe (2007) 45 in the art. As used in the present disclosure, a “low” affinity Clathrin-Dependent Mechanisms of G Protein-coupled indicates that a high concentration of the active ingredient Receptor Endocytosis, Traffic 8:462-470; Schulte (2007) relative to the receptor is required to maximally occupy the Novel aspects of G-protein-coupled receptor signalling—dif binding site of the receptor and trigger a physiological ferent ways to achieve specificity, Acta Physiol 190:33-38: response, while a “high affinity indicates that that a low Torrecilla (2006) Co-ordinated covalent modification of 50 concentration of the active ingredient relative to the receptor G-protein coupled receptors, Curr Pharm Des 12:1797-1808; is adequate to maximally occupy the binding site of the recep Neitzel (2006) Cellular mechanisms that determine selective tor and trigger a physiological response. A "high affinity RGS protein regulation of G protein-coupled receptor signal may correspond to, for example, an active ingredient concen ing, Semin Cell Dev Biol 17:383-389; Sato (2006) Accessory tration of two or more orders of magnitude less than the proteins for G proteins; partners in signaling, Annu Rev Phar 55 concentration of the receptor that is effective to trigger the macol Toxicol 46:151-187; Appert-Collin (2006) Regulation physiological response, while a “low” affinity may corre of g protein-coupled receptor signaling by a-kinase anchoring spond to an active ingredient concentration of one or more proteins. Recept Signal Transduct Res 26:631-46; Premont orders of magnitude greater than the concentration of the (2007) Physiological roles of G protein-coupled receptor receptor that is effective to trigger the physiological response. kinases and arrestins, Annu Rev Physiol 69:51 1-534; Smrcka 60 In Drosophila melanogaster, the olfactory receptors are (2008) G protein beta gamma subunits: Central mediators of located in two pairs of appendages located on the head of the G protein-coupled receptor signaling, Cell Mol Life Sci fly. The family of Drosophila chemoreceptors includes 65:2191-2214; Grandy (2007) Trace amine-associated recep approximately 62 odorant receptor (Or) and 68 gustatory tor 1—Family archetype or iconoclast?, Pharmacol Ther receptor (Gr) proteins, encoded by families of approximately 116:355-390; Gilchrist (2007) G-protein-coupled receptor 65 60 Or and 60 Grgenes through alternative splicing. Some of pharmacology: examining the edges between theory and these receptor proteins have been functionally characterized, proof. Curr Opin Drug Discov Devel 10:446-451; Takeishi while others have been identified by sequence homology to US 8,691,256 B2 19 20 other sequences but have not been fully characterized. Other In some embodiments of the invention, the cell used can be insects have similar olfactory receptor proteins. any cell capable of being transfected with and express a TyR. In certain embodiments, the insect olfactory receptor pro Examples of cells include, but are not limited to: insect cells, tein targeted by the screening or insect control method of the such as Drosophila Schneider cells, Drosophila Schneider 2 invention is the tyramine receptor (TyR). In additional cells (S2 cells), and Spodoptera frugiperda cells (e.g., Sf9 or embodiments, the insect olfactory receptor protein is the Sf21); or mammalian cells, such as Human Embryonic Kid insect olfactory receptor protein Or83b or Or43a. In addi ney cells (HEK-293 cells), African green monkey kidney tional embodiments, the targeted protein can be any of the fibroblast cells (COS-7 cells), HeLa Cells, and Human Kera insect olfactory protein receptors. tinocyte cells (HaCaT cells). Additionally, other components of the insect olfactory 10 The TyrR can be a full-length TyrR, a functional fragment receptor cascade can be targeted using the method of the of a TyrR, or a functional variant of a TyrR. A functional invention in order to identify useful insect control com fragment of a TyrR is a TyrR in which amino acid residues are pounds. Exemplary insect olfactory cascade components that deleted as compared to the reference polypeptide, i.e., full can be targeted by methods of the invention include but are 15 length TyrR, but where the remaining amino acid sequence not limited to serotonin receptor, Or22a, Or22b, GrSa, Gr21a, retains the binding affinity of the reference polypeptide for Gró1a, B-arrestin receptor, GRK2 receptor, and tyramine tyramine. A functional variant of a TyrR is a TyrR with amino B-hydroxylase receptor, and the like. acid insertions, amino acid deletions, or conservative amino With reference to FIG. 1, an exemplary screening method acid substitutions, that retains the binding affinity of the ref for identifying effective pest control compositions can make erence polypeptide for tyramine. A "conservative amino acid use of one or more transfected cell lines expressing a receptor substitution' is a substitution of an amino acid residue with a of interest, for example, a biogenic amine receptor, Such as, a functionally similar residue. Examples of conservative sub TyR or an octopamine receptor. stitutions can include, for example, the Substitution of one In some embodiments of the invention, isolated cell mem non-polar (hydrophobic) residue such as isoleucine, Valine, branes expressing the receptor of interest can be used in 25 leucine or methionine for another; the substitution of one competitive binding assays. Whole cells can be used to study polar (hydrophilic) residue for another such as between argi changes in signaling down-stream to the receptor, in response nine and lysine, between glutamine and asparagine, between to treatment with a test composition. glycine and serine; the Substitution of one basic residue Such Embodiments of the invention can utilize prokaryotic and as lysine, arginine or histidine for another, the Substitution of eukaryotic cells including, for example, bacterial cells, yeast 30 one acidic residue. Such as aspartic acid or glutamic acid for cells, fungal cells, insect cells, nematode cells, plant cells, another, and the like. A conservative amino acid Substitution animal cells, and the like. Suitable animal cells can include, can also include replacing a residue with a chemically deriva for example, HEK cells, HeLa cells, COS cells, U20S cells, tized residue, provided that the resulting polypeptide retains CHO-K1 cells, various primary mammalian cells, and the the binding affinity of the reference polypeptide for tyramine. like. An animal model expressing one or more conjugates of 35 Examples of TyrRs can include, for example: TyrRs, such as, an arrestin and a marker molecule, for example, throughout Drosophila melanogaster TyrR (GENBANKR) accession its tissues, within a particular organ or tissue type, or the like, number (GAN). CAA38565), Locusta migratoria TyrR can be used. (GAN: 25321), TyrRs of other invertebrates, TyrRs of The potential for insect control activity can be identified by nematodes, and the like. measuring the affinity of the test compositions for the recep 40 Exemplary Screening methods can include “positive' tor in the cell lines expressing a TyrR, Or83b, and/or Ora3a. screening, where, for example, compositions that bind a The potential for insect control activity can also be identified receptor of interest are selected. Exemplary screening meth by measuring the change in intracellular cAMP and/or Ca" ods can include “negative' screening, where, for example, in the cell lines expressing TyrR, Or83b, and/or Or43a fol compositions that bind a receptor of interest are rejected. An lowing treatment with the test compositions. The gene 45 exemplary method can include: Selecting a composition that sequences of the TyrR, the Or 83b receptor and the Or 43a binds a TyR. Another exemplary method can include: select receptor have substantial similarity between various insect ing a composition that binds a TyR and does not bind an species. As such, the Drosophila Schneider cell lines express octopamine receptor. ing these receptors can be used to screen for compositions In some embodiments of the invention, the efficacy of a test having insect control activity in various insect species. 50 composition can be determined by conducting studies with In some embodiments, a method of selecting a composition insects. For example, the efficacy of a test composition for for pesticidal use can include the following. A cell expressing repelling an insect can be studied using controlled experi a TyR is provided and is contacted with test compounds. The ments wherein insects are exposed to the test composition. In receptor binding affinity of the compounds is measured. At Some embodiments, the toxicity of a test composition against least one parameter selected from the following parameters is 55 an insect can be studied using controlled experiments measured: intracellular cAMP level, and intracellular Ca" wherein insects are exposed to the test composition. level. A first compound for the composition is identified, that Methods of screening compositions for insect control is capable of altering at least one of the parameters, and that activity are set forth in the following applications, each of has a high receptor binding affinity for the TyR; and a second which is incorporated in its entirety herein by reference: U.S. compound for the composition is identified, that is capable of 60 application Ser. No. 10/832,022, entitled COMPOSITIONS altering at least one of the parameters, and that has a low AND METHODS FOR CONTROLLING INSECTS; U.S. receptor binding affinity for the TyR. A composition is application Ser. No. 11/086,615, entitled COMPOSITIONS selected that includes the first and second compounds. In AND METHODS FOR CONTROLLING INSECTS Some embodiments, a composition is selected that includes RELATED TO THE OCTOPAMINE RECEPTOR; U.S. the first and second compounds and demonstrates an anti 65 application Ser. No. 1 1/365,426, entitled COMPOSITIONS parasitic effect that exceeds the anti-parasitic effect of any of AND METHODS FOR CONTROLLING INSECTS the compounds when used alone. INVOLVING THE TYRAMINE RECEPTOR; and U.S. US 8,691,256 B2 21 22 application Ser. No. 1 1/870,385, entitled COMPOSITIONS TABLE 1-continued AND METHODS FOR CONTROLLING INSECTS. Compositions for Pest Control PEST CONTROL CHEMICALS Embodiments of the invention can include a composition CAS Registry for controlling pests. Embodiments of the invention that Pest Control Chemical Number include a composition for controlling pests can include an CHLOZOLINATE pest control chemical or product. Embodiments of the inven CLOTHIANIDIN tion that include a composition for controlling pests can COPPER (DIFFERENT SALTS) include an active agent. COPPERFUNGICIDES In embodiments of the invention that include an active 10 CYAZOFAMID CYCLOPROPANECARBOXYLIC ACID, 2,2- 39515-40-7 agent, the active agent can be, for example, an agent that can DIMETHYL-3-(2-METHYL-1-PROPENYL)-, have a biological impact on an insect, Such as, for example, a CYANO(3-P ENOXYPHENYL)METHYL chemical, a compound, or the like. In embodiments of the ESTER invention that include an active agent, the active agent can be, CYFLUFENAMID 15 CYFLUTHRIN 68359-37-5 for example, one or more plant essential oils, or the like. The CYHALOFOPBUTYL 122008-85-9 plant essential oils, when combined, can have a synergistic CYHALOTH R N 68O85-85-8 effect. Embodiments can also can include a fixed oil, which is CYHALOTH R NK 91465-08-6 typically a non-volatile, non-scented plant oil. Additionally, CYHALOTH R N (lambda) 91465-08-6 CYHALOTH R N GAMMA 76703-62-3 in Some embodiments, these compositions can be made up of CYMOXANIL generally regarded as safe (GRAS) compounds. CYPERMETHRIN S2315-07-8 In embodiments of the invention that include at least one CYPROCONAZOLE pest control chemical, the at least one pest control chemical CYPRODINIL CYROMAZIN E 66215-27-8 can be selected from, for example, the pest control chemicals D-TRANS-ALLETHRIN 280S7-48-9 set forth in Table 1, or the like. DELTAMETHRIN (DECA-) 52918-63-5 25 AFENTHURON 80060-09-0 TABLE 1. AZINON 333-41-5 CHLOFENTHION 97-17-6 PEST CONTROL CHEMICALS CHLOFLUANID CLOCYMET CAS Registry CLOMEZINE Pest Control Chemical Number 30 CLORAN FENOCONAZOLE ABAMECTIN 71751-41-2 ETHOFENCARB ACEPHATE 30560-19-1 FLUBENZURON 3.5367-38-5 ACETAMIPRID 135410-20-7 FLUMETORIM ACETOCHLOR 34256-82-1 FENOCONAZOLE ACEQUINOCYL S7960-19-7 35 METHIRIMOL ACIBENZOLAR-S-METHYL METHOATE 60-51-5 ALACHLOR 15972-60-8 METHOMORPH ALDICARB 116-06-3 MOXYSTROBIN ALDIMORPH NICONAZOLE ALLETHRIN S84-79-2 NOCAP AMISULBROM SULFOTON 298-04-4 40 AMITRAZ 33O89-61-1 THIANON ANILAZINE ODEMORPH AZACONAZOLE ODINE AZOXYSTROBIN DFINPHOS BIFENTHRIN 826S7-04-3 NDOSULEAN 115-29-7 BENALAXYL NESTROBIN BENDIOCARB 22781-23-3 45 POXICONAZOLE BENTHIAVALICARB ESFENVALERATE 66230-04-4 BENODANIL ETHABOXAM BENOMYL ETHIRIMOL BIFENTHRIN 826S7-04-3 ETRIDIAZOLE BINAPACRYL FAMOXADONE BIORESMETHRIN 28434-01-7 50 FENBUCONAZOLE BIPHENYL BITERTANOL 122-14-5 BLASTICIDIN-S ENOXYCARB 72490-01-8 BOSCALID ENPROPATHRIN 3951S-41-8 BROMUCONAZOLE ENAMIDONE BUPIRIMATE 55 ENARIMOL CAPTAFOL ENEHEXAMID CAPTAN CARBENDAZIM 1563-66-2 CARBOFURAN CARBARYL 63-25-2 ENPROPIMORPH CARBENDAZIM ENTINACETATE CARBOXIN 60 NTINCHLORIDE NTINEHYDROXIDE CARPROPAMID CHLORDIMEFORM 61.64-98-3 S1630-58-1 CHLORFENVINFOS 470-90-6 CHLORONEB ERIMZONE CHLOROTHALONIL 1897-45-6 PRONIL 12OO68-37-3 CHLOROXURON 1982-47-4 65 FLUAZINAM CHLORPYRIFOS 2921-88-2 LUBENDIAMIDE 272451-65-7 US 8,691,256 B2 23 24 TABLE 1-continued TABLE 1-continued

PEST CONTROL CHEMICALS PEST CONTROL CHEMICALS CAS Registry CAS Registry Pest Control Chemical Number Pest Control Chemical Number

FLUDIOXONIL HOPEHOROUSACID AND SALTS FLUMORPH HORATE 52645-53-1 FLUSILAZOLE HOSMET 298-02-2 FLUSULFAMIDE COXYSTROBIN FLUTRIAFOL 10 PERALIN FLUOPICOLIDE OLYOXIN FLUOXASTROBIN RALLETHRIN (ETOC) 23O31-36-9 FLUQUINCONAZOLE ROBENAZOLE (ALSO ANTIBACTERIAL FLUTOLANIL AND ANTIFUNGALACTIVITY) FOSETYL-AL ROCHLORAZ FOLPET 15 ROCYMIDONE FTHALIDE ROFENOFOS 41198-08-7 FUBERIDAZOLE ROPAMOCARB FURAMETPYR ROPICONAZOLE FURAL.AXYL ROPINEB GUAZATINE ROQUINAZID HEXACONAZOLE 67485-29-4 ROTHIOCARB HYDRAMETHYLNON ROTHIOCONAZOLE HYMEXAZOLE PYRACLOSTROBIN MAZALIL PYRAZOPHOS MIBENCONAZOLE 105827-78-9 PYRETHRUM 8003-34-7 MIDACLOPRID PYRIBUTICARB MINOCTADINE PYRIFENOX NDOXACARB 25 PYRIMETHANIL ODOCARB PYRIBENCARB PCONAZOLE PYROQUILON PROBENFOS (IBP) QUINTOZENE (PCNB) PRODINE QUINOXYFEN SOPROTHIOLANE RESMETHRIN 10453-86-8 SOTLANIL 30 SILITHIOFAM KASUGAMYCIN MECONAZOLE KRESOXIM-METHYL PINOSAD 131929-60-7 LAMBDA-CYHALOTHRIN 91465-08-6 ROMESIFEN 283,594-90-1 LUFENURON 1O3OSS-O7-8 ROXAMINE MALATHION 121-75-S TREPTOMYCIN MANCOZEB 35 PHUR MANDIPROPAMID UCONAZOLE MANEB E UFENOZIDE 11241O-23-8 MEPANIPYRIM LOFTHALAM (BACTERICIDE) MEPRONIL NAZENE (TCNB) METALAXYL EFLUTHRIN 79538-32-2 METALAXYL-M (=MEFENOXAM) TERBINAFINE METCONAZOLE 40 ETRACONAZOLE METHIDATHION 950-37-8 THIABENDAZOLE METHAMIDAPHOS 10265-92-6 TIADINIL (O.S.-Dimethylphosphoramidothiolate) THIFLUZAMIDE METHASULFOCARB HIOCYCLAM 3.1895-21-3 METHOMYL 16752-77-5 HIODICARB 59669-26-O METHYL PARATHION 298-OO-O 45 THIOPHANATE METIRAM THIOPHANATE-METHYLH METOMINOSTROBIN H AMETHOXAM 153719-23-4 METRAFENONE THIRAMH MINERAL OILS, ORGANIC OILS, TOLCLOFOS-METHYL POTASSIUM BICARBONATE, MATERIAL TOLYFLUANID OF BIOLOGICAL ORIGIN 50 TRALOMETHRIN 66841-25-6 MYCLOBUTANIL TRIADIMEFON NAFTIFINE TRIADIMIENOL NALED 300-76-5 TRIAZOXIDE NUARIMOL TRICYCLAZOLE OCTHILINONE TRIDEMORPH OFURACE 55 TRIFLOXYSTROBIN ORYSASTROBIN TRIFLUMIZOLE OXADIXYL FORINE OXAMYL 23135-22-0 TICONAZOLE OXOLINIC ACID VALIDAMYCIN OXPOCONAZOLE VALIPEHENALA OXYCARBOXIN VINCLOZOLIN OXYDEMETON METHYL 301-12-2 60 N,N-DIETHYL-3-METHYLBENZAMIDE 134-62-3 OXYTETRACYCLINE (DEET) PEFURAZOATE ZINEB PENCONAZOLE ZIRAM PENCYCURON ZOXAMIDE PENTHIOPYRAD PERMETHRIN 52645-53-1 65 PHENOTHRIN 26002-8O-2 Embodiments of the invention can include compounds Such as, for example, abamectin, allethrin, citronella oil, US 8,691,256 B2 25 26 IR3535(R) (3-N-butyl-N-acetyl-aminopropionic acid ethyl physiological responses. With reference to FIG. 3, insect ester), methyl nonyl ketone, metofluthrin, neem oil, nepeta control chemicals, such as deltamethrin (DM), can also affect lactone, oil of lemon eucalyptus, permethrin, picaridin, downstream signaling. As depicted in FIGS. 2 and 3, the p-menthane 3.8 diol, and the like. compounds or blends of plant origin and the insect control Embodiments of the present invention can include at least one insect control chemical, and at least one compound of a chemicals activate signaling in different manners. plant origin, or at least one blend of compounds of a plant In embodiments that include an insect control chemical, origin. With reference to FIG. 2, compounds of plant origin, the insect control chemical can include, for example, any Such as plant essential oils, can bind certain biogenic amine insect control chemical from the classes listed in the follow receptors, resulting in downstream signaling affecting certain ing table: TABLE 2

CLASSIFICATION OF INSECT CONTROL COMPOSITIONS Chemical Subgroup or exemplifying active Group Subgroup Primary target site of action ingredient Active ingredients 1: 1A Acetylcholine esterase Carbamates Aldicarb inhibitors Bendiocarb Carbaryl Carbofuran Methiocarb Methomyl Oxamyl Propoxur Thiodicarb 1B Organophosphates Acephate AZinphos-methyl Chlorpyrifos Chlorpyrifos methyl Coumaphos Diazinon Dichlorvos Dicrotophos Dimethoate Disulfoton Ethoprop Fenamiphos Fenthion Isofenphos Malathion Methamidophos Methidathion Methyl parathion Naled Oxydemeton methyl Phorate Profenofos Propetamphos Temephos Terbufos Tetrachlorvinphos Trichlorfon 2: 2A GABA-gated chloride Cyclodiene Endosulfan channel antagonists organochlorines Lindane 2B Fipronil Fipronil (phenylpyrazoles) 3 Sodium channel modulators Pyrethroids Alethrin d-cis-trans Alethrin d-trans Alethrin Bifenthrin Bioallethrin S cyclopentenyl Cyfluthrin Beta-Cyfluthrin Cypermethrin Zeta-Cypermethrin Cyphenothrin (1R)-trans isomers Deltamethrin Esfenvalerate Fenpropathrin Fenvalerate miprothrin Permethrin US 8,691,256 B2 27 28 TABLE 2-continued

CLASSIFICATION OF INSECT CONTROL COMPOSITIONS Chemical Subgroup or exemplifying active Group Subgroup Primary target site of action ingredient Active ingredients Phenothrin (1R)- trans-isomer Praethrin Resmethrin Tefluthrin Tetramethrin Trailomethrin Pyrethrins Pyrethrins (pyrethrum) Methoxychlor Methoxychlor 4: 4A Nicotinic acetylcholine Neonicotinoids Acetamiprid receptor agonistSantagonists Imidacloprid Thiamethoxam 4B Nicotine Nicotine 6 Chloride channel activators Avermectins, Abamectin Milbemycins 7: 7A Juvenile hormone mimics Juvenile hormone Hydroprene analogues Kinoprene Methoprene 7B Fenoxycarb Fenoxycarb 8:8 8A Compounds of unknown or Methyl bromide Methyl bromide non-specific mode of action and other alkyl (fumigants) halides 8B Chloropicrin Chloropicrin 8C Sulfuryl fluoride Sulfuryl fluoride 9* 9A Compounds of unknown or Cryolite Cryolite non-specific mode of action (selective feeding blockers) 0: 10A Compounds of unknown or Clofentezine Clofentezine non-specific mode of action Hexythiazox Hexythiazox 1OB (mite growth inhibitors) Etoxazole Etoxazole 1: 11A1 Microbial disruptors of insect B.t. var. israelensis B.t. war. israeinsis 11B1 midgut membranes (includes B.t. var. aizawai B.t. War. aizawaii 11B2 transgenic crops expressing B. t. war. kunstaki B.t. war. kunstaki B. t. toxins) 2: 12B Inhibitors of oxidative Organotin miticides Fentutatin oxide phosphorylation, disruptors of ATP formation (inhibitors of 12C ATP synthase) Propargite Propargite 5 Inhibitors of chitin Benzoylureas Diflubenzuron biosynthesis, type O, Hexaflumuron Leptodopteran Novaluron 7 Moulting disruptor, Dipteran Cyromazine Cyromazine 8:8 18A Ecclysone agonistS moulting Diacylhydrazines Halofenozide disruptors Methoxyfenozide Tebufenozide 18B Azadirachtin Azadirachtin 9 Octopaminergic agonists Amitraz Amitraz 20:8 2OA Mitochondrial complex III Hydramethylnon Hydramethylnon electron transport inhibitors (Coupling site II) 21 Mitochondrial complex I METI acaricides, Rotenone electron transport inhibitors Rotenone 22 Voltage-dependent sodium indoxacarb indoxacarb channel blockers 24: 24A Mitochondrial complex IV Aluminum phosphide Aluminum electron transport inhibitors phosphide 24C Phosphine Phosphine 25 Neuronal inhibitors (unknown Bilfenazate Bilfenazate mode of action) 27: 27A Synergists P450 monooxygenase Piperonyl butoxide inhibitors UN UNC Compounds with unknown Dicofol Dicofol UND mode of action** Pyridalyl Pyridalyl NS NSA Miscellaneous non-specific Borax Borax (multi-site) inhibitors US 8,691,256 B2 29 30 In some embodiments of the invention, the insect control In embodiments of the invention that include at least one chemical can include at least one of for example, an organo organochlorine compound, the organochlorine compound phosphate compound, a carbamate compound, a carbazate can be, for example, endosulfan, dicofil, or the like. compound, a neonicotinoid compound, an organochlorine In embodiments of the invention that include at least one compound, an organotin compound, an oxadiazine com organotin compound, the organotin compound can be, for pound, a pyridaZinone compound, a pyrethroid, a tetrazine example, hexakis, or the like. compound, or the like. In embodiments of the invention that include at least one oxadiazine compound, the oxadiazine compound can be, for In embodiments of the invention that include at least one example, indoxacarb, or the like. organophosphate compound, the organophosphate com In embodiments of the invention that include at least one pound can be, for example, azinphos-methyl, chlorpyrifos, 10 pyridaZinone compound, the pyridaZinone compound can be, diazinon, dimethoate, methidathion, phosmet, or the like. for example, pyridaben, or the like. In embodiments of the invention that include at least one In embodiments of the invention that include at least one carbamate compound, the carbamate compound can be, for pyrethroid, the pyrethroid can be, for example, esfenvalerate, example, methomyl, oxamyl, carbaryl, formetanate, hex fenpropathrin, permethrin, or the like. 15 In embodiments of the invention that include at least one ythiazox, or the like. tetrazine compound, the tetrazine compound can be, for In embodiments of the invention that include at least one example, clofentezine, or the like. carbazate compound, the carbazate compound can be, for Embodiments of the invention can include at least one example, bifenazate, or the like. insect control product; and at least one compound of a plant In embodiments of the invention that include at least one origin, or at least one blend of compounds of a plant origin. neonicotinoid compound, the neonicotinoid compound can The at least one insect control product can be selected from, be acetamiprid, imidacloprid, thiacloprid, thiomethoxam, or for example, the insect control products set forth in Table 4, or the like. the like. TABLE 3

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

ARCHER SOWP NICLOSAMIDE 2,4-D AMINE 6 2,4-D AMINE CHLOROPHENOXY COMPOUND LBS, USG 2,4-D AMINE 3.34 2,4-D AMINE CHLOROPHENOXY COMPOUND LBS, USG 2,4-D AMINE EC 2,4-D AMINE CHLOROPHENOXY COMPOUND 2,4-D ESTER 2,4-D IBE CHLOROPHENOXY COMPOUND 2,4-D ESTER 2,4-D IBE CHLOROPHENOXY COMPOUND 2,4-D ESTER 2,4-D IBE CHLOROPHENOXY COMPOUND 2,4-D ESTER 2,4-D IBE CHLOROPHENOXY COMPOUND 2,4-D ESTER 2,4-D IBE CHLOROPHENOXY COMPOUND 2,4-D GRANULES 2,4-D IBE CHLOROPHENOXY COMPOUND 2,4-D GRANULES 2,4-D IBE CHLOROPHENOXY COMPOUND S Star GENERAL ISOPROTHIOLANE ABATE SOOE TEBUFENOZIDE ABATE SG TEMEPHOS Access 2,4-d ESTER 2,4-D IBE CHLOROPHENOXY COMPOUND ACETAM7S SP ACEPHATE ORGANOPHOSPHATE ACROBAT SOWP DIMETHOMORPH ACROBATMZ DIMETHOMORPH MANCOZEB ACTARA 25 WG THIABENDAZOLE - O PHENOL ACTELLIC 2SEC PIPEROPHOS+ 2,4-D BE ACTIVO 22 SC ANILOFOS - ETHOYSULFRON ADER SEC CYPERMETHRIN PYRETHROID ADMIRE, SWP MAZAQUIN ADVANCEEC BUTACHLOR- MISCELLANEOUS PROPANIL ADVANTAGES G CARBOFURAN CARBAMATE ADVANTAGES G CARBOFURAN CARBAMATE AFALON SOWP LINDANE AGRIMEK1.8EC AVERMECTIN CHLORIDE CHANNELACTIVATOR AGRICOTEMZ 80 WP MANCOZEB DITEHIOCARBAMATE AGRISOLA-15OK POLYOXYETHYLENE DODECYLETEHER AGRISOLA-15OK POLYOXYETHYLENE SORBITANFATTY ACIDS AGRO CYPERMETHRIN PYRETHROID CYPERMETHRINSEC AGROPOINT CARTAP CARTAP 50 SP HYDROCHLORIDE AGROZEB 80 WP MANCOZEB DITEHIOCARBAMATE AL-1OOTS SETHOXYDIM US 8,691,256 B2 31 32 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

ALAKDAN 300 BPMC CHLOPYRIFOS ALIETTE 8OWP FOSETHYL-AL ALIETTE 800 WG FOSETHYL-AL ALMIX 20 WP METRIBUZIN AMBUSH SEC CYPERMETHRIN PYRETHROID AMDRO ANT BAIT HEXYTHIAZOX AMETREX 80 WP AMETRYNE MISCELLANEOUS AMETREX 80 WP AMETRYNE MISCELLANEOUS AMETRYNE 80 WP AMETRYNE MISCELLANEOUS AMISTAR2S SC AZOXYSTROBIN AMMO SEC CYPERMETHRIN PYRETHROID AMWAYAPSA 80 ALKYLARYL ALKOXYLATE-TALL OIL FATTY AC ANCOM BUTACHLOR BUTACHLOR MISCELLANEOUS 60 EC ANCOM CYPERMETHRIN PYRETHROID CYPERMETHRINSEC ANTRACOL 70 WG PROPICONAZOLE ANTRACOL 70 WP PROPINEB ANVILS SC HALOXYFOP-R- METHYLESTER PACHE 10 G CADUSAFOS PACHE 1 OO ME CADUSAFOS PACHE 1 OO ME CADUSAFOS PPLAUD 10 WP BUPROFESIN PRON 3.5 SD MCPA QUADIN 25 EC NICLOSAMIDE QUADIN 70 WP NICLOSAMIDE RGOLD 10 EC CINMETHYLIN RGOLD PLUS CYNMETHYLIN + 2,4-DIBE PYRETHROID RIES SUPER CYPERMETHRIN PYRETHROID ETHRINSEC RMOR THIOPHANATE METHYL RMURE 300 EC DIFECONAZOLE PROPICONAZOLE RNIS 2.5 EC LAMBDACYHALOTHRIN PYRETHROID RRIVO SEC CYPERMETHRIN PYRETHROID RROW SEC CYPERMETHRIN PYRETHROID ASCEND SOSC FIPRONIL ASSET 48 SL GLYPHOSATE MONOETHALONAMINE SALT ASSURE IIEC PYRIMETHANIL ATABRON SE CHLORFLUAZURON ATRAMET COMBI 80 AMETRYNE MISCELLANEOUS WP ATRAZINE ATRAZINE 80 WP ATRAZINE MISCELLANEOUS ATTACKSR CYPERMETHRIN PYRETHROID ATTAINM-8O MALATHION ORGANOPHOSPHATE AVANTECEC BUTACHLOR MISCELLANEOUS PROPANIL AVERMECTIN CHLORIDE CHANNELACTIVATOR YPERMETHRIN PYRETHROID BALEARSOOSC HLOROTHALONIL CHLORONITRILE t OXYSTROBIN BANKO 720 SC LOROTHALONIL MISCELLANEOUS BANKO 720 SC LOROTHALONIL MISCELLANEOUS BANKO 7SWP LOROTHALONIL MISCELLANEOUS BANNER 60 EC TACHLOR MISCELLANEOUS BANOLE OIL PARAFFINOIL BANOLE OIL 60 PARAFFINIC MINERAL L BASACRAN 48EC ENTAZONE BASAMID G AZOMET PYRETHROID BASTA 1S SL BBERRELICACID BASUDIN 40 WP AZINON ORGANOPHOSPHATE BASUDIN4OOEC AZINON ORGANOPHOSPHATE BASUDIN 600 EC AZINON ORGANOPHOSPHATE BAVISTINSO DF ARBAR Y L CARBAMATE BAYCOR3OOEC TERTANOL BAYLETON 25 WP i HIOPHANATE ETHYL BAYLUSCIDE 250 EC CLOSAMIDE US 8,691,256 B2 33 34 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

BAYLUSCIDE 50 WP NICLOSAMIDE ETHANOLAMINE SALT BAYLUSCIDE 70 WP NICLOSAMIDE ETHANOLAMINE SALT BAYONET 6% METALDEHYDE PELLETS BAY THROID 0125 EC CYFLUTHRIN Pyrethroid BAY THROID OSOEC CYFLUTHRIN Pyrethroid BAZZOKA CHLORPYFIROS Organophosphate + Carbamate BPMC BELEREXTABLET GIBBERRELICACID BELORAN 400 SL BENZOXONIUM CHLORIDE ENLATESOWPOD BENOMYL ENSUL 10 WP BENSULFURON METHYL ERDUGO SOWP NICLOSAMIDE ETHANOLAMINE SALT ERELEXTABLET GENERICNAME DA2SEC LAMBACYHALOTHRIN PYRETHROID FLEX 1 OTC FENTHRIN FLEX 1 OTC FENTHRIN FLEX 2.5 TC FENTHRIN FLEXTC FENTHRIN OACT WG PACLOBUTRAZOL ODAN3 G CARBUFORAN CARBAMATE OZEB MANCOZEB DITEHIOCARBAMATE OZEB 80 WP MANCOZEB DITEHIOCARBAMATE LADE 60 EC BUTACHLOR MISCELLANEOUS LINK 27 SEC CHLORPYFIROS ORGANOPHOSPHATE CYPERMETHRIN BLOCKADE 480 SL BENTAZONE BLUE COP 770 WP COPPERHYDROXIDE MISCELLANEOUS BOLT 50 SP CARTAP CARBAMATE BOOST SOOSC ACIBENZOLAR-S- METHYL BOXER SEC CYPERMETHRIN PYRETHROID BRAVO 720 FLO CHLOROTHALONIL MISCELLANEOUS BREAK-THRU POLYCARBOXYLIC ACID BRODAN 31.5 EC CHLORPYFIROS Organophosphate + Carbamate BPMC BROMO GAS METHOMYL BRONCHO GLYPHOSATE AMMONIUMSALT UGBUSTER SEC CYPERMETHRIN PYRETHROID ULLDOZER SOWP NICLOSAMIDE ULLET 5 EC CYPERMETHRIN PYRETHROID ULLSEYE CYPERMETHRIN PYRETHROID NSECTICIDE BUMPER 25 EC PROPICONAZOLE BURNDOWN 16OAS GLYPHOSATEDI AMMONIUMSALT BURNDOWN 16OAS GLYPHOSATE IPA BUSHWHACK SEC CYPERMETHRIN PYRETHROID BUTACHLOR 600 EC BUTACHLOR MISCELLANEOUS BUTATAF 60 E. BUTACHLOR MISCELLANEOUS CALIBER 70 WP NICLOSAMIDE CALIBER 70 WP NICLOSAMIDE CALIXIN 7SEC TRICLOPYR CAPTAN SOWP CAPTAN MISCELLANEOUS CAPTAN SOWP CAPTAN MISCELLANEOUS CAPTURE SEC CYPERMETHRIN PYRETHROID CARANCHO 2. SEC ETHOFENPROX CARBARYL 8SS CARBARYL CARBAMATE CARVIL SOEC BPMC CARBAMATE CASCADE 1 OWDC FLUFENOXURON CELCURE A(P)WOOD COPPER, CHROME, ARSENIC PRESERVE (CCA) CHAKU 2. SEC LAMBDACYHALOTHRIN CHAMPDP COPPERHYDROXIDE COPPER CHAMPIONWP CUPRICHYDROXIDE COPPER CHESS 25 WP PROPINEB CHESS SOWG PYMETROZINE CHDX 2. SEC BETACYPERMETHRIN PYRETHROID CHLORMITE TC CHLOROPYFIROS ORGANOPHOSPHATE US 8,691,256 B2 35 36 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

CHOPPER 85 S CARBARYL CARBAMATE CITRUS LUSTER 213 HIABENDAZOLE CIVIL 75 WP HLOROTHALONIL MISCELLANEOUS CLEANFIELDEC UTACHLOR PROPANIL MISCELLANEOUS CLEAR OUT 41 LYPHOSPATE IPA CLEAR OUT 41 PLUS LYPHOSPHATE IPA CLINCHER1OOEC YHALOFOP BUTYL COBRA 20 EC HLORPYFIROS ORGANOPHOSPHATE COMBATSEC YPERMETHRIN PYRETHROID COMMAND 3 ME COMMAND 3 ME COMMAND PLUS 600 EC COMPETE 75 SP ACEPHATE ORGANOPHOSPHATE COMPRO 600 EC CLOMAZONE - PROPANIL CONFIDOR1OOSL IMIDACLOPRID CONFIDOR 200 SL IMIDACLOPRID CONTRAZINE 80 WP ATRAZINE MISCELLANEOUS CONTRAZINE 80 WP ATRAZINE MISCELLANEOUS CONTROL 2SOEC NICLOSAMIDE CONTROL 70 WP NICLOSAMIDE ETHANOLAMINE SALT CORSAIR SEC PENDIMETHLIN PYRETHROID CORSAIR SEC PERMETHRIN PYRETHROID COSAVETDF SULFUR COTRINSEC CYPERMETHRIN PYRETHROID COTRINSEC CYPERMETHRIN PYRETHROID COUNTER1O G TEMEPHOS COZEB 80 WP MANCOZEB DITEHIOCARBAMATE CRUSHER2SOEC NICLOSAMIDE CRUSHER SOWP NICLOSAMIDE ETHANOLAMINE SALT CRUSHER 70 WP NICLOSAMIDE CULTAR2S SC OXYFLUORFEN CUPRAVIT OB 21 COPPER COPPER OXYCHLORIDE CURZATEM MANCOZEB DITEHIOCARBAMATE FUNGICIDE CYBEST SEC CYPERMETHRIN PYRETHROID CYCLONE SEC CYPERMETHRIN PYRETHROID CYMBUSH 5 EC CYPERMETHRIN PYRETHROID CYPER-5 CYPERMETHRIN PYRETHROID CYPERMETHRINSEC CYPERMETHRIN PYRETHROID CYPERTHRINSEC CYPERMETHRIN PYRETHROID CYPEX 50 EC CYPERMETHRIN PYRETHROID CYPROSEC CYPERMETHRIN PYRETHROID CYREN3OOEC HLORPYFIROS ORGANOPHOSPHATE DACINOL 278750 WP HLOROTHALONIL MISCELLANEOUS DACINOL 278775 WP HLOROTHALONIL MISCELLANEOUS DACONIL 72OSC HLOROTHALONIL MISCELLANEOUS

EADBOL CLOSAMIDE ECIDE 2.5 EC LTAMETHRIN ECIS 19/o SC LTAMETHRIN ECIS 2.5 EC LTAMETHRIN ECISM 2.5 EC LTAMETHRIN ECIS R LTAMETHRIN ECIS TAB LTAMETHRIN EFENSA 5 EC C PERMETHRIN ESCH MAGTOXIN LUFENURON EGESCH MAGNESSIUM PLATES STRIPS PHOSPHIDE EGESH PHOSTOXIN ALUMINUM RODENTICIDE PHOSPHIDE ELMARK2 SEC LTAMETHRIN PYRETHROID ETIA GAS EX-B A. UMINUM RODENTICIDE PHOSPHIDE DETIA GAS EX-T ALUMINUM RODENTICIDE PHOSPHIDE DETIAPHOSPHINE ALUMINUM RODENTICIDE PELLETS PHOSPHIDE ACARB SOEC BPMC CARBAMATE AFURAN 10 G CARBOFURAN CARBAMATE AFURAN3 G CARBOFURAN CARBAMATE AFURANSG CARBOFURAN CARBAMATE AGRANSG AZINON ORGANOPHOSPHATE AGRANSG AZINON ORGANOPHOSPHATE US 8,691,256 B2 37 38 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

AZINON 40 EC DIAZINON ORGANOPHOSPHATE AZINON 60 EC DIAZINON ORGANOPHOSPHATE AZINON 60 EC DIAZINON ORGANOPHOSPHATE AZINON 60 EC DIAZINON ORGANOPHOSPHATE AZINON 600 EC DIAZINON ORGANOPHOSPHATE AZOL 40 EC DIAZINON ORGANOPHOSPHATE AZOL 40 EC DIAZINON ORGANOPHOSPHATE AZOL 60 EC DIAZINON ORGANOPHOSPHATE AZOL 60 EC DIAZINON ORGANOPHOSPHATE CARE 37.5 WG DIAFENTHURON ORGANOPHOSPHATE FENOXYCAB CARZOL 20 SP FORMETHANATEHCL MOSOSP CARTAP HYDROCHLORIDE DIPEL WP BACILLUS PLANT ORIGIN THURINGIENSIS DIPTEREX 95 SP TRIBUTYLPOLYGLYCO ETHER EK 800 BUTACHLOR SAFENER THANE F-448 MANCOZEB THIOCARBAMATE THANE F-448 MANCOZEB THIOCARBAMATE THANEM-45 MANCOZEB THIOCARBAMATE THANEM-45 MANCOZEB THIOCARBAMATE THANEM-45 WP MANCOZEB THIOCARBAMATE THANEOS 600 MANCOZEB THIOCARBAMATE THANEOS-600 MANCOZEB THIOCARBAMATE UREX 80 WP DIURON REA UREX 80 WP DIURON REA URON 80 WP DIURON REA URON 80 WP DIURON REA URON 80 WP DIURON REA MA 3.34 LBS, USG 2,4-D AMINE HLOROPHENOXY COMPOUND RAGO 60 WP FLUFENACET REXELDIURON 80 DIURON U REA

REXELMALATHION MALATHION ORGANOPHOSPHATE EC REXEL SULFA 80 W SULFUR URSBAN CHLORPYRIFOS ORGANOPHOSPHATE YNAMEC AVERMECTIN CHLORIDE CHANNELACTIVATOR E ASY SEC CYPERMETHRIN PYRETHROID LTRA 20OSC CARBOFURAN CARBAMATE QUATION PRO 52.5 CYMOXANIL FAMOXADONE RASER 70 EC BUTACHLOR MISCELLANEOUS PROPANIL ETHREL 10 SL ETHEPHON

ETHRELPGR4.8% ETHEPHON ETROFOLAN SOWP ISAZOFOS EXPERT 20 EC CHLORPYFIROS ORGANOPHOSPHATE EXTREME SOSP ARTAP YDROCHLORIDE FASTAC 15 WDG LPHACYPERMETHRIN PYRETHROID FASTAC 2SOSC LPHACYPERMETHRIN PYRETHROID FASTACR HACYPERMETHRIN PYRETHROID - CARBAMATE . C FENOMD 225 EC Organophosphate + Pyrethroid

FLASH SEC Cy R I t N PYRETHROID FLIP 500 WP CLOSA FLIP 700 WP CLOSAMIDE S. THANOLAMINE SALT FOLICUR2SOEC SULPHUR FOLICUR 430 SC TEBUCONAZOLE FORWARD 700 EC BUTACHLOR MISCELLANEOUS PROPANIL FROWNCIDE SOSC F LUAZINAM FRUITONE CPA C HLOROPHENOXY P ROPIONIC ACID FUJI-ONE 40 EC ISOPROCARB FUMITOXIN ALUMINUM RODENTICIDE PHOSPHIDE FUNGAFLORSOL HYDRAMETHYLNON FUNGAFLOR75 SP IMAZALIL US 8,691,256 B2 39 40 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

FUNGITOX 70 WP THIOPHANATE METHYL FUNGURAN-OH COPPERHYDROXIDE MISCELLANEOUS FURADAN 10G. CARBARYL CARBAMATE FURADAN3 G CARBENDAZIM CARBAMATE FURADAN. 3G CARBOFURAN CARBAMATE FURADANS G CARBARYL CARBAMATE FURADANS G CARBOFURAN CARBAMATE FURUDAN 10 G CARBOSULFAN CARBAMATE FURUDAN3 G CARBOSULFAN CARBAMATE FURUDANSG CARBUFORAN CARBAMATE GALLANTSUPER HALOSULFURON METHYL GARLON 4 TRICHLOROFON GAROTEEC CHLORPYRIFOS - BPMC Organophosphate + Carbamate GAS 2SOEC NICLOSAMIDE GAUCHO 70 WS IMIDACLOPRID EM 2,4-D AMINE 2,4-D AMINE CHLOROPHENOXY COMPOUND EM 2.4-D ESTER 2,4-D IBE CHLOROPHENOXY COMPOUND EMATRAZINE ATRAZINE MISCELLANEOUS EMMALATEHION S7 MALATHION ORGANOPHOSPHATE

EMTRAK SOSP CARTAP HYDROCHLORIDE ESAPAX SOOFW AMETRYNE MISCELLANEOUS ESAPAX 80 WP AMETRYNE MISCELLANEOUS ESAPAX COMBI 80 AMETRYNE MISCELLANEOUS P ATRAZINE ESAPRIM 80 WP ATRAZINE MISCELLANEOUS LADIATOR75 WDG CHLORPYRIFOS ORGANOPHOSPHATE LYPHOMAX GLYPHOSATE IPA OAL 24 EC OXADIAZON OAL 24 EC OXYFLOURFEN RAMOXONE 20 AS PARAFINIC MINERAL OIL RASSEDGE THIOBENCARB RASSEDGE 800 EC THIOBENCARB + 2,4-D UARDIAN SEC CYPERMETHRIN PYRETHROID ALT BACILLUS PLANT ORIGIN THURINGIENSIS HEDONAL LIQ. SL 2,4-D AMINE CHLOROPHENOXY COMPOUND

BADOX 33 EC PENDIMETHALIN BIMAX PIRIMIPHOSMETHYL HERCULES 20 EC TRIADIMEFON HI-CONFILF 75 WP CHLOROTHALONIL MISCELLANEOUS HIDROCOB 77 WP COPPERHYDROXIDE MISCELLANEOUS HINOSAN3OOEC EDIFENPHOS Organophosphate HINOSAN SOEC EDIFENPHOS Organophosphate HI-PER SEC CYPERMETHRIN PYRETHROID HIT 250 EC NICLOSAMIDE HITWP NICLOSAMIDE ESTICK TRIAZOPHOS PCIDE. SOEC BPMC CARBAMATE PCIN SOEC BPMC CARBAMATE PKILL SOEC BPMC CARBAMATE STATHION2OEC TRIAZOPHOS DROX 77 WP COPPERHYDROXIDE COPPER DROXIDE SUPER COPERHYDROXIDE COPPER i7 WP DROXIDE SUPER COPPERHYDROXIDE COPPER 7 WP TOX 50 WP MICP HYWARX BROMACIL WEEDKILLER MAGE 1.5 LC IMAZALIL MPACT 2. SEC DELTAMETHRIN PYRETHROID NDAR 2F FENBUCONAZOLE ORGANOPHOSPHATE NDAR 2F FENBUCONAZOLE ORGANOPHOSPHATE NSECT PRO SOSP CARTAP HYDOCLORIDE NSECT PRO SOSP CARTAP HYDOCLORIDE NSTAR CARTAP HYDROCHLORIDE NVEST 10 WP CYCLOSULFAMURON US 8,691,256 B2 41 42 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

VADIURON 80 WP DIURON MISCELLANEOUS VAPYRITILINE 20 PE CHLORPYFIROS ORGANOPHOSPHATE MB WAZEB 80 WP MANCOZEB DITEHIOCARBAMATE KARATE 2.5 EC LAMBDACYHALOTHRIN PYRETHROID KARATE W ZEON LAMBDACYHALOTHRIN PYRETHROID TECHNOLOGY KARET 40 MANEB W, ZINC KARMEX DIURON MISCELLANEOUS WEEDKILLER KHOLUSCIDE 70 WP CLOSAMIDE ETHANOLAMINE SALT CK2SEC NICLOSAMIDE CK70 WP NICLOSAMIDE LLER SEC CYPERMETHRIN PYRETHROID LPES3 EC FENVALERATE PYRETHROID NG SEC CYPERMETHRIN PYRETHROID TALATRAZINE ATRAZINE MISCELLANEOUS TAL MANCOZEB ANCOZEB DITEHIOCARBAMATE TAL STRYKER SEC YPERMETHRIN PYRETHROID LEENUP 480 AS LYPHOSATE IPA LENUP 48OAS LYPHOSATE IPA LERAT WITH RODIFACOUM COUMARIN TRE LIK 700 EC UTACHLOR MISCELLANEOUS ROPANIL KNOCKOUT SEC YPERMETHRIN PYRETHROID KOCIDE 101 UPRICHYDROXIDE MISCELLANEOUS KOCIDEDF UPRICHYDROXIDE MISCELLANEOUS KOCIDEDF 2000 OPPERHYDROXIDE MISCELLANEOUS KOP-HYDROXIDE SO OPPERHYDROXIDE MISCELLANEOUS WP KOTETSU 10 SC CHLORPHENAPYR ORGANOPHOSPHATE KRISSEC LAMBDACYHALOTHRIN PYRETHROID KUH2AK2SEC NICLOSAMIDE KUH2AK7O WP NICLOSAMIDE KUMULUSDF ELEMENTAL SULFUR LANNATE 4.0 SP METHIOCARB LARVIN3SOFS THIOBENCARB + 2,4-DIBE LATRON B-1956 PHENTHOATE-BPMC LEAD CORP. 2,4-D 2,4-D AMINE CHLOROPHENOXY COMPOUND AMINE LEADCORP CARTAP CARTAP HYDROCHLORIDE LEADCORP MALATHION ORGANOPHOSPHATE MALATHION S7 EC LEADMARK3 EC FENVALERATE PYRETHROID LEADONILSOOSC CHLOROTHALONIL MISCELLANEOUS LEADREXTC CHLORPYFIROS ORGANOPHOSPHATE LEADTHREL 480 SL ETHEPON LEBAYCID SOEC FENTHION LECSPRO 44WP FENTRAZAMIDE - PYRETHROID PROPANIL LENTREKTC HLORPYRIFOS ORGANOPHOSPHATE LENTREKTC HLORPYRIFOS ORGANOPHOSPHATE LINDAFOR 7SF LAMBDACYHALOTHRIN ORGANOCHLORINE LONDAXWP BENSULFURON METHYL LORSBAN 3E C HLORPYRIFOS ORGANOPHOSPHATE LORSBAN 40 EC C HLORPYRIFOS ORGANOPHOSPHATE LUTENSOLA8 ALKYL POLYETHELENE GLYSOL ETHER LUV 2,4-D ESTER 2,4-D IBE CHLOROPHENOXY COMPOUND LUWMALATHION S7 MALATHION ORGANOPHOSPHATE EC MACHETES G BUTACHLOR MISCELLANEOUS MACHETEEC BUTACHLOR MISCELLANEOUS MACHETE EXPRESS BUTACHLOR MISCELLANEOUS MACHO BUTACHLOR MISCELLANEOUS MAGIK S90 EC CYPERMETHRIN PYRETHROID MAGNUMSEC CYPERMETHRIN PYRETHROID MAITHREL 10 PGR. ETHEPON MAITHREL 48 PGR. ETHEPON MALATHION S7 E MAGNESSIUM ORGANOPHOSPHATE PREMIUM PHOSPHIDE US 8,691,256 B2 43 44 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

MALATHION S7 EC MALATHION ORGANOPHOSPHATE MALATHION S7 EC MALATHION ORGANOPHOSPHATE MANAGER 80 WP MANCOZEB DITEHIOCAR BAMATE MANZATE 200 MANCOZEB DITEHIOCAR BAMATE FUNGICIDE MANZATE 7SDF MANCOZEB DITEHIOCAR MANZEB 80 WP MIPC MARSBYL 85 WP CARBARYL CARBAMAT MARVEL SEC CYPERMETHRIN PYRET THROI MASO 70 WP NICLOSAMIDE MASTER2SEC LAMBDACYHALOTHRIN PYRET THROI MASTRADIURON 80 DIURON MISCELLANEOUS C ATADOR 60 SC METAMIDOPHOS ORGANOPHOSPHATE ATCHOSOEC LINURON UREA ATON SEC CYPERMETHRIN PYRET THROID EBROM METHYL BROMIDE - CHLOROPICRIN EGARIFOS 20 EC CHLORPYFIROS ORGANOPHOSPHATE EGATHRINSEC CYPERMETHRIN PYRET THROID ELODY DUO IPRODIONE ESUROL SOWP METHAMIDOPHOS ETA BAIT METALDEHYDE ETA BAIT 6% METALAXYL-m- ELLETS MANCOZEB ETABROM METHYL BROMIDE - CHLOROPICRIN CROTHIOLDF MIC 20 F MIC 20 F TEBUFENOZIDE NER SOSP HYDROCHLORIDE PCIN SOWP METSULFURON METHYL CHLORIMURON ETH MIRACLE AMINE 2,4-D AMINE CHLOROPHENOXY COMPOUND MIRAL 3 G IPROVALICARB PROPINEB MOCAP 10 G ETHOPROP MODEL SEC CYPERMETHRIN PYRETHROID MOLUXIDE 2SOEC NICLOSAMIDE MOSPHILAN3 EC ACETAMIPRID NABU-S QUIZALOFOP-P-ETHYL NEMACUR1O G PERMETHRIN - Zn NEMACUR400 EC PHENAMIPHOS NEMATHORIN 1.0 G. FOZTEHIAZATE NET 50 WP NICLOSAMIDE ETHANOLAMINE SALT NICLOSM NICLOSAMIDE NISSORUN SEC HEXAFLUMURON NOBLITE 60 WG FENAMIDONE - MANCOZEB NOMINEE 100 SC SPYRIBAC SODIUM NOMINEE 100 SC B SPYRIBAC SODIUM NORDOX 50 WP C OPPEROXIDE COPPER NURELLED C HLORPYFIROS ORGANOPHOSPHATE CYPERMETHRIN NUVACRON 300 SCW M. ZETHYLENE SDITHIOCARBAMATE NYDREL 100 ETHEPHON NYDREL 480 ETHEPHON OCHO SWP CARBARYL CARBAMATE OMEGA 45 EC PRETILACHLOR FENCLORIM ONECIDE 15 EC FLUAZIFOP-P-BUTYL ORTHENEACETAM ACEPHATE ORGANOPHOSPHATE 75 SP ORTHENE 75 SP ACEPHATE ORGANOPHOSPHATE OXYCHLOR 85 WP COPPER OXYCHLORIDE PADAN SOSP CAPTAN CARBAMATE HYDROCHLORIDE PADAN SOSP CARTAP HYDROCHLORIDE PARAFUNGUS 80 WP MANCOZEB DITEHIOCARBAMATE PARAKUHOL 2SOEC NICLOSAMIDE PARAPESTD 400 EC DIAZINON ORGANOPHOSPHATE US 8,691,256 B2 45 46 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

PARAULOD 3OOEC CHLORPYFIROS ORGANOPHOSPHATE PARTNER 40 DF CARFENTRAZONE ETHYL PARTNER 40 DF CARFENTRAZONE ETHYL PASSPORT SOOSC CHLOROTHALONIL MISCELLANEOUS PENNANT PHENAMIPHOS PERFEK 31.5 EC HLORPYRIFOS Organophosphate + Carbamate PMC PERFEKTHION 40 EC METHOATE PERMIT 10 WP LYSOPHATE IPA PESTMASTER PERMETHRIN PYRETHROID PILARICH, SOO G, LFP HLOROTHALONIL MISCELLANEOUS PILARZEB 80 WP ANCOZEB DITEHIOCARBAMATE PIPSET 35 WP NOSULFURCN PEROPHOS PISTOL SOWP CLOSAMIDE ETHANOLAMINE PISTOL SOWP NICLOSAMIDE ETHANOLAMINE SALT PLANTERS MALATHION ORGANOPHOSPHATE MALATHION S7 EC POLIDO 2. SEC ETHOFENPROX PORSANAIL ETALDEHYDE POSSE2OOSC ARBOSULFAN CARBAMATE POWER LYPHOSATE IPA POWER SUPRATECH LYPHOSATEDI :MMONIUMSALT PREDATOREC HLORPYFIROS ORGANOPHOSPHATE PREDATOR PLUS HLORPYFIROS ORGANOPHOSPHATE YPERMETHRIN PREKILL 330 PARAQUAT DICHLORIDE REMISE 20OSC IMIDACLOPRID REMIUMSEC CYPERMETHRIN PYRETHROID REVENT 77 WP COPPERHYDROXIDE COPPER REVICUR-N PROFENOFOS ROCIN 25 WP BUFROFESIN ROCURE SOWP BENOMYL ROPLANT PROPAMOCARB ROVADO SUPRA OSO IMIDACLOPRID

ROVIN 85 WP CARBARYL CARBAMATE UNISEHSSEC CYPERMETHRIN PYRET THROID PYRITILENE 20 PE CHLOPYFIROS ORGANOPHOSPHATE MB PYTOX 10 EC PERMETHRIN PYRET THROID QUICKPHOS (ROUND ALUMINUM RODENTICIDE TAB) PHOSPHIDE RACUMINDUST COUMATETRALYL RADISSON MANCOZEB DITEHIOCARBAMATE MANCOZEB 80 WP RADOR 262. SEC CHLORPYFIROS Organophosphate + Pyrethroid BETACYFLUTHRIN RAFT 800 WG NICLOSAMIDE ETHANOLAMINE SALT RAPIDO SEC CYPERMETHRIN PYRETHROID RATKILZINC WARFARIN PHOSPHIDE80% BAIT RATOXINP TRISILOXANE ALKOXYLATE--ALLYL ETHOXYLA RECRUIT II HEXACONAZOLE REDEEM 80 WP MANCOZEB DITEHIOCARBAMATE RED-OUT 80 WP MANCOZEB DITEHIOCARBAMATE REGENT 0.3 GR FIPRONIL REV 800 WP MANCOZEB DITEHIOCARBAMATE RICESTAREC FENOXAPROPP-ETHYL RIDOMIL GOLD MZ 68 METALAXYL WP MANCOZEB RIDOMIL MZS8 WP METALAXYL RILOF SOOEC PICLORAM + 2,4-D RIPCORD 2SEC CYPERMETHRIN PYRETHROID ROBODAX 2SEC NICLOSAMIDE ROGUEEC BUTACHLOR+ 2,4-D RONSTAR2SEC OXADIARGYL US 8,691,256 B2 47 48 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

RONSTAR 2G OXADIAZON ROUND-UP BIOSORB GLYPHOSATE ISOPROPYLAMINE SALT ROUNDUPEW GLYPHOSATE IPA ROUND-UP MAX GLUFOSINATE AMMONIUM ROVER CHLOROTHALONIL MISCELLANEOUS ROVRAL SOWP INDOXACARB ROVRAL AQUAFLO IPRODIONE SOSC ROYAL CARTAP CARTAP CARBAMATE ROYANIL 75 WP LOROTHALONIL MISCELLANEOUS SABEDONG SEC PERMETHRIN PYRETHROID SAMURAI 6OEC ACHLOR MISCELLANEOUS SANAFURAN3 G OSULEAN CARBAMATE SANAZOLE 2SOEC s CONAZOLE SAPROLEC FLUMIZOLE SATURN 60 EC AMETOXAM SATURND OBENCARB + 2,4-D SATURNS OBENCARB THIOCARBAMATE SAVIOR 80 WP iNCOZEB DITEHIOCARBAMATE SCOPE 70 WP M OPHANATE THYL SCORE 2SOEC ENOCONAZOLE SELECRON SOOEC C HLORAZ MN LECT 120 EC LETHODIM LECT 120 EC LETHODIM NCOR 70 WP ETHYL BROMIDE - HLOROPICRIN SENTINEL 75 WP HLOROTHALONIL MISCELLANEOUS SERVWEL 2,4-D 2,4-D AMINE CHLOROPHENOXY COMPOUND AMINE SERVWEL MALATHION ORGANOPHOSPHATE MALATHION S7 EC SERVWEL MANCOZEB DITEHIOCARBAMATE MANCOZEB 80 WP RVWELTKO SOEC CYPERMETHRIN PYRETHROID RV WEL 2,4-D 2,4-D IBE CHLOROPHENOXY COMPOUND RANULES VIN SOWP CARBUFORAN CARBAMATE VIN 85 WP CARBUFORAN CARBAMATE ERPA SEC CYPERMETHRIN PYRETHROID HIELD HLOROTHALONIL MISCELLANEOUS HOTGUNM ANCOZEB DITEHIOCARBAMATE CO 250 EC FENOCONAZOLE GA 300 EC HLORPYRIFOS ORGANOPHOSPHATE GANEX 600 SC YMETROZINE GMA LYPHOSPHATE IPA LWET 4.08 RISILOXANE LKOXYLATE--ALLYL ETHOXYLA SILWET 4.08 FORINE SLASH LYPHOSATE IPA SMART 480 LYPHOSATE IPA SMART 480 LYPHOSATE IPA SMASH SEC Y PE RMETHRIN PYRETHROID SNAIL CHAMP 2SEC CLOSAMIDE SNAIL OUT SOWP CLOSAMIDE SNAILKIL 6% P METALDEHYDE SNIPER SEC CYPERMETHRIN PYRETHROID SOFIT 300 EC POLYOXYETHYLENE SORBITANTFATTY ACIDS SOLIGNUMBROWN PERMETHRIN PYRETHROID SOLIGNUM PERMETHRIN PYRETHROID COLORLESS SOLNET SOOEC PRETILACHLOR SONIC 60 EC BUTACHLOR MISCELLANEOUS SPECTRA SEC CYPERMETHRIN PYRETHROID SPEED 25 EC MONOCROTOPHOS SPEED SOWP NICLOSAMIDE SPEEDEX POLYETHERPOLYMETHYLSILOXANE COPOLYME STAM LV-10 PROPAMOCARB HCL STAR SEC CYPERMETHRIN PYRETHROID US 8,691,256 B2 49 50 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

STEADFASTTC ALPHACYPERMETHRIN PYRETHROID STEWARD WDG MIDACLOPRID CYFLUTHRIN STIMUKIL FLY BAIT METHOMYL STINGRAY S.625 DELTAMETHRIN, BUPROFEZIN STIX 480 EC CARBUFORAN CARBAMATE STOP 6% PELLETS METALDEHYDE STORMWAXW. FLOCOUMAFEN BITREX SUCCESS SORBITAN NATURALYTE 25 SC MONOOLATE(SB), POLYOXYETHYL UMI-ALPHA2. SEC ESFENVALERATE PYRETHROID UMI-ALPHA2. SEC ESFENVALERATE PYRETHROID UMI-ALPHA2. SEC ESFENVALERATE PYRETHROID UMICIDIN FENVALERATE PYRETHROID UMICIDIN3 EC FENVALERATE PYRETHROID UMICIDIN3 EC FENVALERATE PYRETHROID UMI-EIGHT DINICONAZOLE UMITHION4O WDP FENITROTHION ORGANOPHOSPHATE UMITHION SOEC FENITROTHION ORGANOPHOSPHATE UMITHION SOEC FENITHROTHION ORGANOPHOSPHATE UMITHION SOEC FENITROTHION ORGANOPHOSPHATE UNRICE 15 WDG ETHOXYSULFURON UNSPRAY 8N PAECILOMYCES LILACINUS STRAIN 2S1 S UPER BLUE 85 WP COPPER COPPER OXYCHLORIDE SUPREME SEC CYPERMETHRIN PYRETHROID SUPREMOEC BPMC CHLORPYFIROS SURE 2SOEC NICLOSAMID SUREKILL 70 WP NICLOSAMID SURFACTANTA-100 POLYETHER POLYMETHYLSILOXANE COPOLYM SURFACTANTA-100 POLYOXYETHYLENE DODECYLETHER SURFIX BETAPINENE OLYMER SWEEP HIOPHANAT E ETHYL SWIPE 25 EC CLOSAMID SWIPESOWP CLOSAMID TAMARON 600 SL ETALDEHYDE TAMEX 360 EC UTRALIN TARGET 2.5 EC CLOSAMID TARGET 25 EC CLOSAMID TECTO 45 FW ETRAMETHYLTHIURAM SULPHIDE TEGAO75 EC RIDEMORPH TELONE II CHCHLOROPROPENE TERMEX 48EC HLORPYFIROS ORGANOPHOSPHATE RMIDOR 2. SEC PRONIL RMINATOR2SEC PYRETHROID

ERMITE-X CHLORPYFIROS ORGANOPHOSPHATE TERRAGUARD 48EC CHLORPYFIROS ORGANOPHOSPHATE THESIS 2. SEC DELTAMETHRIN PYRETHROID THIRAM 80 WG TETRAMETHYLTHIURAM DISULPHIDE THYLATE 8OWG T TIGER2S SC N CLOSAMIDE TILT 250 EC C TIMBER GUARD C ERMETHRIN - Zn CLEAR TIMBER GUARD PYRETHROID MEDIUMBROWN TOP 70 WP

TOPNOTCH THIODICARB TOPSIN-M 70 WP THIOPHANATE METHYL TOPSTAR 60 EC OXADIARGYL TORDON 101 PHTHALIC GLYCEROLALKYL MIXTURE TORNADO 6OEC BUTACHLOR - PROPANIL MISCELLANEOUS TORNADO 6OEC BUTACHLOR MISCELLANEOUS PROPANIL US 8,691,256 B2 51 52 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

TORO BUTACHLOR MISCELLANEOUS PROPANIL TORPEDO SEC CYPERMETHRIN PYRETHROID TRAMEX COMBI 80 AMETRYNE MISCELLANEOUS WP ATRAZINE TRANZEB 4SSFC MANCOZEB DITEHIOCARBAMATE TRANZEB 80 WP MANCOZEB DITEHIOCARBAMATE TRAP 70 WP NICLOSAMIDE EBON 10 EC ETHOFENPROX EBON 10 EC ETHOFENPROX : EBON 10 EW ETHOFENPROX EFIC 20 WP ETHOFENPROX FMINE 30 WP TRIFLOXYSTROBIN GARD 75 WP CYROMAZINE M50 WP LINURON NEB 80 WP MANCOZEB - CYMOXANIL TRIO SOWP PROCEHLORAZ TRIPLEX 5 OEC CYPERMETHRIN PYRETHROID TROJAN31.SEC CHLORPYFIROS - BPMC TWISTER 70 EC BUTACHLOR MISCELLANEOUS PROPANIL TWISTEREC BUTACHLOR - PROPANIL MISCELLANEOUS LTIMOEC 200 NICLOSAMIDE LTIMOEC 225 NICLOSAMIDE PROOT 60 EC BUTACHLOR MISCELLANEOUS ECTRON 10 EW ETHOFENPROX ECTRON2O WP ETHOFENPROX EGETOX SOSP CARTAP CARBAMATE ERTIMEC AVERMECTIN CHLORIDE CHANNELACTIVATOR EXTER 300 EC CHLORPYFIROS ORGANOPHOSPHATE NDEXPLUS PHENTHOATE SOCOL SOWP NICLOSAMIDE TAL BLUE 85 WP COPPER COPPER OXYCHLORIDE TIGRANBLUE S8 COPPER COPPER WP OXYCHLORIDE WITIGRANBLUE S8 COPPER COPPER WP OXYCHLORIDE WONDOZEB 42 SC MANCOZEB DITEHIOCARBAMATE VONDOZEB 75 DF MANCOZEB DITEHIOCARBAMATE WONDOZEBL MANEB WONDOZEB PLUS MANCOZEB DITEHIOCARBAMATE WALLOP 70 WP NICLOSAMIDE WARRIOR 31.5 CHLORPYRIFOS ORGANOPHOSPHATE - CARBAMATE BPMC WAZARY 10 FL FENVALERATE PYRETHROID WAZARY 10 FL FENVALERATE THROID WEAPON SEC CYPERMETHRIN THROID WEDKILL 2,4-D 2,4-D IBE CHLOROPHENOXY COMPOUND WEEDER 60 E BUTACHLOR MISCELLANEOUS WEEDTROL 40 EC 2,4-D IBE CHLOROPHENOXY COMPOUND WEISERATRAZINE 80 ATRAZINE 3,5-TRIAZINE WP WEISSERATRAZINE ATRAZINE 3,5-TRIAZINE 80 WP WEISSER CYPERMETHRIN PYRETHROID CYPERMETHRINSEC WHIP-S 120 EW FENOXAPROPP-ETHYL WHIP-S 75 EW FENOXAPROPP-ETHYL WINNER SEC CYPERMETHRIN PYRETHRO WIPERSEC CYPERMETHRIN PYRETHRO R WOLMAN CCA-C COPPER, CHROME, ARSENIC (CCA) XENTARIWDG BACILLUS PLANT ORIGIN THURINGIENSIS X-PHOS 20 EC CHLORPYFIROS ORGANOPHOSPHATE X-PHOS 40 EC CHLORPYFIROS ORGANOPHOSPHATE X-RAT 190 P WARFARIN XTRAGRO 10 LS ETHEPHON XTRAGRO 240 PGR. ETHEPHON XTRAGRO 480 PGR. ETHEPHON ZACARB 85 WP CARBARYL CARBAMATE ZACK SOWP MIPC ZECTRIC 6% PELLETS METALDEHYDE ZEPHYR AVERMECTIN CHLORIDE CHANNELACTIVATOR US 8,691,256 B2 53 54 TABLE 3-continued

INSECT CONTROL PRODUCTS

Brand Name Generic name Classification

ZINCPHOSPHIDE 80 ZINCPHOSPHIDE DP ZOOMSEC CYPERMETHRIN PYRETHROID

10 Embodiments of the invention can include at least one nary ammonium herbicides, thiocarbamate herbicides, thio biologically-based insecticide, such as, for example, abam carbonate herbicides, thiourea herbicides, triazine herbicides, ectin, proteins and/or spores derived from Bacillus thurinien chlorotriazine herbicides, methoxytriazine herbicides, meth sis, spinosad, or the like. ylthiotriazine herbicides, triazinone herbicides, triazole her bicides, triazolopyrimidine herbicides, uracilherbicides, urea Embodiments of the invention can include at least one 15 herbicides, phenylurea herbicides, sulfonylurea herbicides, insect growth regulator, Such as, for example, etoxazol, meth pyrimidinylsulfonylurea herbicides, triazinylsulfonylurea oxyfenozide, pyriproxyfen, or the like. herbicides, thiadiazolylurea herbicides, unclassified herbi Embodiments of the invention can include at least one oil, cides, and the like. such as, for example, “Superior oil.” highly-refined oils, and Embodiments of the invention can include a fungicidal the like. chemical or product. In some embodiments, these fungicidal Embodiments of the invention can include at least one chemicals can include, for example, aliphatic nitrogen fungi pheromone, Such as, for example, Codling moth pheromone, cides, amide fungicides, acylamino acid fungicides, anilide Oriental fruit moth pheromone, and the like. fungicides, benzanilide fungicides, furanilide fungicides Sul Embodiments of the invention can include a herbicidal fonanilide fungicides, benzamide fungicides, furamide fun chemical or product. In some embodiments, these herbicidal 25 gicides, phenylsulfamide fungicides, Sulfonamide fungi chemicals can include, for example, amide herbicides, anilide cides, valinamide fungicides, antibiotic fungicides, herbicides, arylalanine herbicides, chloroacetanilide herbi strobilurin fungicides, aromatic fungicides, benzimidazole cides, sulfonanilide herbicides, sulfonamide herbicides, fungicides, benzimidazole precursor fungicides, benzothiaz thioamide herbicides, antibiotic herbicides, aromatic acid ole fungicides, bridged diphenyl fungicides, carbamate fun herbicides, benzoic acid herbicides, pyrimidinyloxybenzoic 30 gicides, benzimidazolylcarbamate fungicides, carbanilate acid herbicides, pyrimidinylthiobenzoic acid herbicides, fungicides, conazole fungicides, copper fungicides, dicar phthalic acid herbicides, picolinic acid herbicides, quinolin boximide fungicides, dichlorophenyl dicarboximide fungi ecarboxylic acid herbicides, arsenical herbicides, benzoylcy cides, phthalimide fungicides, dinitrophenol fungicides, clohexanedione herbicides, benzofuranylalkylsulfonate her dithiocarbamate fungicides, imidazole fungicides, inorganic bicides, benzothiazole herbicides, carbamate herbicides, 35 fungicides, mercury fungicides, morpholine fungicides, carbanilate herbicides, cyclohexene oxime herbicides, cyclo organophosphorus fungicides, organotin fungicides, oxathin propylisoxazole herbicides, dicarboximide herbicides, dini fungicides, oxazole fungicides, polysulfide fungicides, pyra troaniline herbicides, dinitrophenol herbicides, diphenyl Zole fungicides, pyridine fungicides, pyrimidine fungicides, ether herbicides, nitrophenyl ether herbicides, dithiocarbam pyrrole fungicides, quinoline fungicides, quinone fungicides, ate herbicides, halogenated aliphatic herbicides, imidazoli 40 quinoxaline fungicides, thiazole fungicides, thiazolidine fun none herbicides, inorganic herbicides, nitrile herbicides, gicides, thiocarbamate fungicides, thiophene fungicides, tri organophosphorus herbicides, oxadiazolone herbicides, phe azine fungicides, triazole fungicides, urea fungicides, unclas noxy herbicides, phenoxyacetic herbicides, phenoxybutyric sified fungicides, and the like. herbicides, phenoxypropionic herbicides, aryloxyphenox In embodiments of the invention that include at least one ypropionic herbicides, phenylenediamine herbicides, pyra 45 compound or chemical of a plant origin, the at least one Zole herbicides, benzoylpyrazole herbicides, phenylpyrazole compound or chemical of a plant origin can include, for herbicides, pyridazine herbicides, pyridazinone herbicides, example, any of the compounds or chemicals listed in table 4. pyridine herbicides, pyrimidinediamine herbicides, quater or the like: TABLE 4

COMPOUNDS OF PLANT ORIGIN

TANETHOLE CORN OIL LILACFLOWER PIPERONAL ALLYL SULFIDE B-COSTOL OIL (LFO) PIPERONYL ALLYLTRISULFIDE CRYPTONE LIME OIL PIPERONYL ALLYL-DISULFIDE CUMINOIL D-LIMONENE ACETATE ARTEMISIA CURZERENONE LINALOOL PIPERONYL ALCOHOLACETATE P-CYMENE LINALYL ALCOHOL BENZALDEHYDE DAVANONE ACETATE PIPERONYL BENZOIC ACID DIALLYL LINALYL AMINE BENZYLACETATE TETRASULFIDE ANTHRANILATE PRENAL BENZYLALCOHOL DIETHYL LINDESTRENE PULEGONE BERG AMOTENE PHTHALATE LINDENOL QUININE B-BISABOLENE DIHYDROPYROCURZERENONE LINSEED OIL ROSEMARY OIL BISABOLENE OXIDE DIEHYDROTAGENTONE METHYL-ALLYL- SABINENE A-BISABOLOL BETA-ELEMENE TRISULFIDE SABINYL BISABOLOLOXIDE GAMMA- MENTHOL ACETATE BISOBOLOLOXIDE B ELEMENE MENTHONE SAFFLOWER OIL BORNYLACETATE ELMOL 2-METHOXY A-SANTALENE US 8,691,256 B2 55 56 TABLE 4-continued

COMPOUNDS OF PLANT ORIGIN

B-BOURBONENE ESTRAGOLE FURANODIENE SANTALOL BLACK SEED OIL 2-ETHYL-2- MENTHYL SATIVEN (BSO) HEXEN-1-OL ACETATE A-SELINENE A-CADINOL EUGENOL METHYL SESAME OIL CAMPHENE EUGENOL CINNAMATE B A-CAMPHOLENE ACETATE METHYL. CITRATE SESQUPHELANDRENE A-CAMPHOLENE A-FARNESENE METHYLDI- SILICONE FLUID ALDEHYDE (Z.E)-A- HYDROJASMONATE SODIUMLAURYL CAMPHOR FARNESENE MENTHYL SULFATE CARVACROL E-B-FARNESENE SALICYLATE SOYBEAN OIL D-CARVONE FENCEHONE MINERAL OIL SPATHULENOL L-CARVONE FURANODIENE MUSKAMBRETTE TAGETONE CARYOPHYLLENE A-1,3-DIENE MYRCENE TANGERINE OIL OXIDE FURANOEUDESM MYRTENAL A-TERPINENE TRANS- A-1,4-DIENE NERALDIMETHYL TERPINENE900 CARYOPHYLLENE FURANO ACETATE A-TERPINEOL CASTOR OIL GERMACRA NEROLIDOL A-TERPINOLENE CEDAR OIL 10(15)-DIENE-6- NONANONE GAMMA CHAMAZULENE ONE GAMMA- TERPINEOL 1,8-CINEOLE FURANOSESQUITERPENE NONALACTONE A-TERPINYL CINNAMALDEHYDE GARLIC OIL OIL OF ACETATE CINNAMYL GERANIOL PENNYROYAL 2-TERT-BUTYL-P- ALCOHOL GERANIOL OILIVE OIL QUINONE CINNAMON OIL ACETATE ORANGE SWEET A-TEHUJONE CITRALA GERANIAL OIL THYME OIL CITRALB GERMACRENED 1-OCTANOL THYMOL ISOPROPYL GERMACRENEB E OCIMIENONE THYMYLMETHYL CITRATE GRAPEFRUIT OIL Z, OCIMIENONE ETHER CITRONELLAL A-GURUNENE 3-OCTANONE GAMMA CITRONELLA OIL A-HUMULENE OCIMIENE UNDECALACTONE CITRONELLOL A-IONONE OCTYLACETATE VALERIC CITRONELLYL B-IONONE PEANUT OIL ANHYDRIDE ACETATE ISOBORNEOL PERILLYL WANILLIN CITRONELLYL ISOFURANOGERM ALCOHOL TRANS FORMATE ACRENE PEPPERMINT OIL VERBENOL CLOVE OIL ISO-MENTHONE A- CIS-VERBENOL A-COPAENE ISO-PULEGONE PHELLANDRENE VERBENONE CORNMINT OIL JASMONE WHITE MINERAL LECITHIN PHELLANDRENE OIL LEMON OIL PHENETHYL YOMOGI LEMON GRASS PROPRIONATE ALCOHOL OIL PHENYL ZINGIBERENE ACETALDEHYDE A-PINENE B-PINENE PINE OIL TRANS PINOCARVEOL

Additional compounds and chemicals of a plant origin that 45 In embodiments of the invention that include at least one can be used in accordance with embodiments of the present blend of compounds of a plant origin, the compounds of plant invention are set forth in the following applications, each of origin can be tested for their precise chemical composition which is incorporated in its entirety herein by reference: U.S. using, for example, High-Pressure Liquid Chromatography application Ser. No. 10/832,022, entitled COMPOSITIONS (HPLC), Mass Spectrometry (MS), gas chromatography, or AND METHODS FOR CONTROLLING INSECTS; U.S. 50 the like. application Ser. No. 11/086,615, entitled COMPOSITIONS The term “about' or “approximately’ means within an AND METHODS FOR CONTROLLING INSECTS acceptable error range for the particular value as determined RELATED TO THE OCTOPAMINE RECEPTOR; U.S. by one of ordinary skill in the art, which will dependin part on application Ser. No. 1 1/365,426, entitled COMPOSITIONS how the value is measured or determined, i.e., the limitations AND METHODS FOR CONTROLLING INSECTS 55 of the measurement system, i.e., the degree of precision INVOLVING THE TYRAMINE RECEPTOR; and U.S. required for a particular purpose. Such as a pharmaceutical application Ser. No. 1 1/870,385, entitled COMPOSITIONS formulation. For example, “about can mean within 1 or more AND METHODS FOR CONTROLLING INSECTS. than 1 Standard deviations, per the practice in the art. Alter In certain embodiments, it can be desirable to include a natively, “about can mean a range of up to 20%, preferably naturally-occurring version or a synthetic version of a com 60 up to 10%, more preferably up to 5%, and more preferably pound. For example, in certain embodiments it can be desir still up to 1% of a given value. Alternatively, particularly with able to include a synthetic lime oil that can be obtained respect to biological systems or processes, the term can mean commercially. In certain exemplary compositions, it can be within an order of magnitude, preferably within 5-fold, and desirable to include a compound that is designated as meeting more preferably within 2-fold, of a value. Where particular Food Chemical Codex (FCC), for example, Geraniol Fine 65 values are described in the application and claims, unless FCC or Tetrahydrolinalool FCC, which compounds can also otherwise stated the term “about” meaning within an accept be obtained commercially. able error range for the particular value should be assumed. US 8,691,256 B2 57 58 The term “substantially, as used herein, means at least agents, vitamins, tanning agents, plant extracts, anti-inflam about 80%, preferably at least about 90%, more preferably at matory agents, anti-oxidants, radical scavenging agents, ret least about 99%, for example at least about 99.9%. In some inoids, alpha-hydroxy acids, emollients, antiseptics, antibiot embodiments, the term 'substantially can mean completely, ics, antibacterial agents, antihistamines, and the like, and can or about 100%. be present in an amount effective for achieving the therapeu In embodiments of the invention that include at least one tic or cosmetic result desired. blend of compounds of a plant origin, the at least one blend of In some embodiments, compositions of this invention can compounds can include at least two compounds. For include one or more materials that can function as an antioxi example, in an exemplary embodiment, the at least one blend dant, Such as reducing agents and free radical scavengers. of compounds can include LFO and Black Seed Oil (BSO). 10 Suitable materials that can function as an antioxidant can Other exemplary embodiments include the blends of com include, for example: acetyl cysteine, ascorbic acid, t-butyl pounds set forth on pages 71-120 of WIPO Publication No. hydroquinone, cysteine, diamylhydroquinone, erythorbic WO/2008/088827, published on Jul. 24, 2008. acid, ferulic acid, hydroquinone, p-hydroxyanisole, hydroxy In certain embodiments wherein the composition includes lamine Sulfate, magnesium ascorbate, magnesium ascorbyl LFO, one or more of the following compounds can be sub 15 phosphate, octocrylene, phloroglucinol, potassium ascorbyl stituted for the LFO: Tetrahydrolinalool, Ethyl Linalool, tocopheryl phosphate, potassium sulfite, rutin, Sodium ascor Heliotropine, Hedion, Hercolyn D, and Triethyl Citrate. In bate, sodium sulfite, sodiumthloglycolate, thiodiglycol, thio certain embodiments wherein the composition includes LFO, diglycolamide, thioglycolic acid, thiosalicylic acid, toco a blend of the following compounds can be substituted for the pherol, tocopheryl acetate, tocopheryl linoleate, tris LFO: Isopropyl myristate, Tetrahydrolinalool FCC, Linalool, (nonylphenyl)phosphite, and the like. Geraniol Fine FCC, Piperonal (aldehyde), and Vanillin. Embodiments of the invention can also include one or more In certain embodiments wherein the composition includes materials that can function as a chelating agent to complex LFO, a blend of the following compounds can be substituted with metallic ions. This action can help to inactivate the for the LFO: Isopropyl myristate, Tetrahydrolinalool, Lina metallic ions for the purpose of preventing their adverse lool, Geraniol, Piperonal (aldehyde), Vanillin, Methyl Sali 25 effects on the stability or appearance of a formulated compo cylate, and D-limonene. sition. Chelating agents Suitable for use in an embodiment of In certain embodiments wherein the composition includes this invention can include, for example, aminotrimethylene BSO, one or more of the following compounds can be sub phosphonic acid, beta-alanine diacetic acid, calcium diso stituted for the BSO: alpha-thujene: alpha-pinene; beta dium EDTA, citric acid, cyclodextrin, cyclohexanediamine pinene; p-cymene; limonene; and tert-butyl-p-benzoquinone. 30 tetraacetic acid, diammonium citrate, diammonium EDTA, In certain exemplary embodiments wherein the composi dipotassium EDTA, disodium azacycloheptane diphospho tion includes Thyme Oil, one or more of the following com nate, disodium EDTA, disodium pyrophosphate. EDTA (eth pounds can be substituted for the Thyme Oil: thymol, C-thu ylene diamine tetraacetic acid), gluconic acid, HEDTA (hy jone; C-pinene, camphene, B-pinene, p-cymene, C-terpinene, droxyethyl ethylene diamine triacetic acid), methyl linalool, borneol, B-caryophyllene, and carvacrol. 35 cyclodextrin, pentapotassium triphosphate, pentasodium Compounds used to prepare the exemplary compositions aminotrimethylene phosphonate, pentasodium triphosphate, of the present invention can be obtained from commercial pentetic acid, phytic acid, potassium citrate, potassium glu SOUCS. conate, sodium citrate, sodium diethylenetriamine pentam In some embodiments of the compositions, it can be desir ethylene phosphonate, Sodium dihydroxyethylglycinate, able to include compounds each having a purity of about 60%, 40 Sodium gluconate, sodium metaphosphate, sodium metasili 65%, 70%, 75%, 80%, 85%, 90%, or 95%. For example, in cate, sodium phytate, triethanolamine ("TEA)-EDTA, TEA Some embodiments of the compositions that include geraniol, polyphosphate, tetrahydroxypropyl ethylenediamine, tet it can be desirable to include a geraniol that is at least about rapotassium pyrophosphate, tetrasodium EDTA, tetrasodium 60%, 85% or 95% pure. In some embodiments, it can be pyrophosphate, tripotassium EDTA, trisodium EDTA, triso desirable to include a specific type of geraniol. For example, 45 dium HEDTA, trisodium phosphate, and the like. in some embodiments, the compositions can include: Embodiments of the invention can also include one or more geraniol 60, geraniol 85, or geraniol 95. When geraniol is materials that can function as a humectant. A humectant is obtained as geraniol 60, geraniol 85, or geraniol 95, then forty added to a composition to retard moisture loss during use, percent, fifteen percent, or five percent of the oil can be Nerol. which effect is accomplished, in general, by the presence Nerol is a monoterpene (CoHO), that can be extracted from 50 therein of hygroscopic materials. attar of roses, oil of orange blossoms and oil of lavender. In some embodiments, each compound can make up Embodiments of the present invention can include art between about 1% to about 99%, by weight (wit/wt %) or by recognised ingredients normally used in Such formulations. volume (vol/vol%), of the composition. For example, one These ingredients can include, for example, antifoaming composition of the present invention comprises about 2% agents, anti-microbial agents, anti-oxidants, anti-redeposi 55 alpha-Pinene and about 98% D-limonene. As used herein, tion agents, bleaches, colorants, emulsifiers, enzymes, fats, percent amounts, by weight or by Volume, of compounds are fluorescent materials, fungicides, hydrotropes, moisturisers, to be understood as referring to relative amounts of the com optical brighteners, perfume carriers, perfume, preservatives, pounds. As such, for example, a composition including 7% proteins, silicones, soil release agents, solubilisers, Sugar linalool, 35% thymol, 4% alpha-pinene, 30% para-cymene, derivatives, Sun screens, Surfactants, vitamins waxes, and the 60 and 24% soybean oil (vol/vol%) can be said to include a ratio like. of 7 to 35 to 4 to 30 to 24 linalool, thymol, alpha-pinene, In certain embodiments, embodiments of the present para-cymene, and soybean oil, respectively (by Volume). As invention can also contain other adjuvants or modifiers such Such, if one compound is removed from the composition, or as one or more therapeutically or cosmetically active ingre additional compounds or other ingredients are added to the dients. Exemplary therapeutic or cosmetically active ingredi 65 composition, it is contemplated that the remaining com ents useful in the compositions of the invention can include, pounds can be provided in the same relative amounts. For for example, fungicides, Sunscreening agents, Sunblocking example, if soybean oil were removed from the exemplary US 8,691,256 B2 59 60 composition, the resulting composition would include 7 to 35 In Formula 4, A is expressed as % effect of the blend, X, to 4 to 40 linalool, thymol, alpha-pinene, and para-cymene, is expressed as % effect of the comparison composition (X). respectively (by Volume). This resulting composition would and C is expressed as % (wit/wt) or % (vol/vol) concentration include 9.21% linalool, 46.05% thymol. 5.26% alpha-pinene, of the comparison composition in the blend. and 39.48% para-cymene (vol/vol%). For another example, 5 In some embodiments of the invention, a coefficient of if safflower oil were added to the original composition to yield synergy of 1.1, 1.2, 1.3, 1.4, or 1.5 can be substantial and a final composition containing 40% (vol/vol) safflower oil, commercially desirable. In other embodiments, the coeffi then the resulting composition would include 4.2% linalool, cient of synergy can be from about 1.6 to about 5, including 21% thymol, 2.4% alpha-pinene, 18% para-cymene, 14.4% but not limited to 1.8, 2.0, 2.5, 3.0, 3.5, 4.0, and 4.5. In other soybean oil, and 40% safflower oil (vol/vol%). One having 10 embodiments, the coefficient of synergy can be from about 5 ordinary skill in the art would understand that volume per to 50, including but not limited to 10, 15, 20, 25, 30, 35, 40, centages are easily converted to weight percentages based the and 45. In other embodiments, the coefficient of synergy can known or measured specific gravity of the Substance. be from about 50 to about 500, or more, including but not Surprisingly, by combining certain insect control chemi limited to 50, 75, 100, 125, 150, 200, 250, 300,350, 400, and cals, and compounds or blends of the present invention, insect 15 450. Any coefficient of synergy above 500 is also contem control activity of the resulting compositions can be plated within embodiments of the present invention. enhanced, i.e., a synergistic effect on insect control activity is Given that a broad range of synergies can be found in achieved when a certain chemical or chemicals, and a certain various embodiments of the invention, it is expressly noted compound or compounds are combined. In other words, the that a coefficient of synergy can be described as being compositions including certain combinations of at least one 'greater than a given number and therefore not necessarily chemical, and at least one compound or at least one blend of limited to being within the bounds of a range having a lower compounds can have an enhanced ability to control insects, as and an upper numerical limit. Likewise, in some embodi compared to each of the chemicals or compounds taken alone. ments of the invention, certain low synergy coefficients, or In embodiments of the present invention, “synergy can lower ends of ranges, are expressly excluded. Accordingly, in refer to any Substantial enhancement, in a combination of at 25 Some embodiments, synergy can be expressed as being least two ingredients, of a measurable effect, when compared “greater than a given number that constitutes a lower limit of with the effect of one active ingredient alone, or when com synergy for Such an embodiment. For example, in some pared with the effect of the complete combination minus at embodiments, the synergy coefficient is equal to or greater least one ingredient. Synergy is a specific feature of a com than 25; in Such an embodiment, all synergy coefficients bination of ingredients, and is above any background level of 30 below 25, even though substantial, are expressly excluded. enhancement that would be due solely to, e.g., additive effects Compositions containing combinations of certain chemi of any random combination of ingredients. Effects include cals and compounds can be tested for synergistic effect on but are not limited to: repellant effect of the composition; insect control activity by comparing the effect of a particular pesticidal effect of the composition; perturbation of a cell combination of at least one chemical, and at least one com message or cell signal Such as, e.g., calcium, cyclic-AMP, and 35 pound or at least one blend of compounds, to the effect of the the like; and diminution of activity or downstream effects of individual chemical(s) and compound(s). Additional infor a molecular target. mation related to making a synergy determination can be In various embodiments, a Substantial enhancement can be found in the Examples set forth in this document. expressed as a coefficient of synergy, wherein the coefficient Exemplary methods that can be used to determine the is a ratio of the measured effect of the complete blend, divided 40 synergistic effect of a particular composition are set forth in by the effect of a comparison composition, typically a single the following applications, each of which is incorporated in ingredient or a Subset of ingredients found in the complete its entirety herein by reference: U.S. application Ser. No. blend. In some embodiments, the Synergy coefficient can be 10/832,022, entitled COMPOSITIONS AND METHODS adjusted for differences in concentration of the complete FOR CONTROLLING INSECTS; U.S. application Ser. No. blend and the comparison composition. 45 11/086,615, entitled COMPOSITIONS AND METHODS Synergy coefficients can be calculated as follows. An activ FOR CONTROLLING INSECTS RELATED TO THE ity ratio (R) can be calculated by dividing the '% effect of the OCTOPAMINE RECEPTOR: U.S. application Ser. No. composition (A) by the '% effect of the comparison compo 11/365,426, entitled COMPOSITIONS AND METHODS sition (X), as follows: FOR CONTROLLING INSECTS INVOLVING THE 50 TYRAMINE RECEPTOR; and U.S. application Ser. No. R-A/X, Formula 1 1 1/870,385, entitled COMPOSITIONS AND METHODS A concentration adjustment factor (F) can be calculated FOR CONTROLLING INSECTS. based on the concentration (C), i.e., 9% (wit/wt) or % (vol/ In some embodiments, synergy or synergistic effect asso Vol), of the comparison composition in the composition, as ciated with a composition can be determined using calcula follows: 55 tions similar to those described in Colby, S. R. “Calculating synergistic and antagonistic responses of herbicide combina F=100/C, Formula 2 tions. Weeds (1967) 15:1, pp. 20-22, which is incorporated herein by this reference. In this regard, the following formula The synergy coefficient (S) can then be calculated by mul can be used to express an expected% effect (E) of a compo tiplying the activity ratio (R) and the concentration adjust 60 sition including two compounds, Compound X and Com ment factor (F), as follows: pound Y: Formula 3 E=X-Y-(X*Y/100) Formula 5 As such, the synergy coefficient (S) can also by calculated, In Formula 5, X is the measured actual % effect of Com as follows: 65 pound X in the composition, and Y is the measured actual % effect of Compound Y of the composition. The expected '% S-(A/X)(100). C. Formula 4 effect (E) of the composition is then compared to a measured US 8,691,256 B2 61 62 actual % effect (A) of the composition. If the actual % effect Furthermore, embodiments of the invention can include a (A) that is measured differs from the expected% effect (E) as method for screening a composition for indirect GPCR calculated by the formula, then the difference is due to an desensitization inhibitory activity. In certain embodiments of interaction of the compounds. Thus, the composition has the invention, an indication that the test composition has synergy (a positive interaction of the compounds) when A.E. indirect GPCR desensitization inhibitory activity can be Further, there is a negative interaction (antagonism) when apparent when a test composition has GPCR desensitization AE'. resensitization inhibitory activity with respect to each of the Compositions containing two or more compounds in cer two or more different GPCRs. In certain embodiments of the tain ratios or relative amounts can be tested for a synergistic invention, indications of resensitization inhibition can effect by comparing the pesticidal effect of a particular com 65 include a reduced response to extracellular stimuli. Such as, position of compounds to the pesticidal effect of a component for example, a reduction in GPCR recycling from the plasma the composition. membrane to the cells interior and back to the plasma mem US 8,691,256 B2 63 64 brane, or the like. Another indication can be an altered period nist for desensitization), or the like, then determining whether for the GPCR regulated activation of the Ca" cascade or the the composition has GPCR desensitization inhibitory activity cAMP levels in the organism. with respect to the first GPCR and with respect to the second In an embodiment of the invention, one cell can be used to GPCR. In such embodiments, an indication of GPCR desen screen a test composition for indirect GPCR desensitization 5 sitization inhibitory activity with respect to the first GPCR inhibitory activity. In such an embodiment, the cell can and an indication of GPCR desensitization inhibitory activity express two or more GPCRs that are different from each other with respect to the second GPCR can be distinguished by Such that a detection method can be used for determining using, for example, a different conjugate for the determina whether there is an indication that a test composition has tion of GPCR desensitization inhibitory activity of the com GPCR desensitization inhibitory activity with respect to each 10 positions with respect to the different GPCRs, or the like. For of the different GPCRs. example, a cell can include a first conjugate comprising a first In some embodiments of the invention, a multi-well format GPCR and a first marker molecule and a second conjugate can be used to screen a test composition for indirect GPCR comprising a second GPCR and a second marker molecule. In desensitization inhibitory activity. In some embodiments, Such an embodiment, it can be possible to expose the cell to each well of the plate can contain at least one cell that includes 15 the test composition, the agonist for the first GPCR (if needed a GPCR, and the assay can include adding a compound in an for desensitization), and the agonist for the second GPCR (if amount known to activate that GPCR, and thus affect intrac needed for desensitization) simultaneously or non-simulta ellular Ca" levels, to each well. In some embodiments, at neously, and determine whether the composition has GPCR least one test compound can also be added to each well. In desensitization inhibitory activity with respect to the first some embodiments, Ca" level can be tested at various time 20 GPCR and the Second GPCR. points after adding the at least one test compound. In certain Detection for each of the items/events discussed above can embodiments, time points used for testing intracellular Ca" be conducted, for example, at one point in time, over a period level can extend beyond the time points where an increase in of time, at two or more points in time for comparison (e.g., Ca" level can be seen without the presence of the at least one before and after exposure to a test composition), or the like. test compound. In some embodiments, methods of the inven- 25 An indication of GPCR desensitization inhibitory activity can tion can identify compounds that prolong agonist effect on be determined by, for example, detecting one or more of the GPCRs. In some embodiments of the invention, cAMP levels items or events discussed above in a cell exposed to the test can be evaluated to gauge the effect of the at least one test composition and comparing the results to those obtained by compound on GPCR response. detecting for the same item or event in a control cell, by In some embodiments of the invention, a multi-well format 30 comparing the results to a predetermined value, or the like. can be used to screen a test composition for indirect GPCR Embodiments of the invention can utilize prokaryotic and desensitization inhibitory activity. In some embodiments, eukaryotic cells including, for example, bacterial cells, yeast each well of the plate can contain at least one cell that includes cells, fungal cells, insect cells, nematode cells, plant cells, a GPCR, and the assay can include adding a compound in an animal cells, and the like. Suitable animal cells can include, amountless than that required to activate that GPCR, and thus 35 for example, HEK cells, HeLa cells, COS cells, U20S cells, affect intracellular Ca" levels, to each well. In some embodi CHO-K1 cells, various primary mammalian cells, and the ments, at least one test compound can also be added to each like. An animal model expressing one or more conjugates of well. In some embodiments, Ca" level can be tested at vari an arrestin and a marker molecule, for example, throughout ous time points after adding the at least one test compound. In its tissues, within a particular organ or tissue type, or the like, certain embodiments, time points used for testing intracellu- 40 can be used. lar Ca level can extend beyond the time points where an Embodiments of the invention can utilize at least one cell increase in Ca level can not be seen without the presence of that expresses, for example, a known GPCR, a variety of the at least one test compound. In some embodiments, meth known GPCRs, an unknown GPCR, a variety of unknown ods of the invention can identify compounds that enhance GPCRs, a modified GPCR, a variety of modified GPCRs, and agonist effect on GPCRs. In some embodiments of the inven- 45 the like. The at least one cell can, for example, naturally tion, cAMP levels can be evaluated to gauge the effect of the express the GPCRs, can be genetically engineered to express at least one test compound on GPCR response. the GPCRs at varying levels of expression, can be genetically In some embodiments of the invention, a cell used in the engineered to inducibly express the GPCRs, or the like. method can also include at least one conjugate comprising a In certain embodiments of the invention, the at least one marker molecule and a protein associated with the GPCR 50 cell can comprise one or more conjugates of a marker mol desensitization pathway of one or more of the GPCRs that are ecule and a protein associated with the GPCR desensitization being evaluated. The conjugate can indicate, through the use pathway. For example, one or more of the cells can comprise of the marker molecule, GPCR desensitization inhibitory a conjugate of an arrestin protein and a marker molecule, or a activity of a test composition with respect to each of the conjugate of a GPCR and a marker molecule, or the like. GPCRs that are being used to screen the test composition. The 55 For certain embodiments of the invention, marker mol conjugate can comprise, for example, an arrestin protein and ecules that can be used as a conjugate can include, for a marker molecule, a GPCR and a marker molecule, or the example, molecules that are detectable by spectroscopic, like. In one embodiment, the cell can comprise a conjugate of photochemical, radioactivity, biochemical, immunochemi an arrestin protein and a marker molecule as well as a conju cal, colorimetric, electrical, and optical means, including, for gate of a GPCR and a marker molecule. 60 example, bioluminescence, phosphorescence, fluorescence, In some embodiments of the invention, two or more differ and the like. Marker molecules can be, for example, biologi ent GPCRs that require agonist for desensitization, or are cally compatible molecules, and the like. Suitable marker constitutively desensitized, can be used. In general. Such molecules can include, for example, radioisotopes, epitope methods can comprise exposing the cell to, for example, a test tags, affinity labels, enzymes, fluorescent groups, chemilumi composition, to an agonist for the first GPCR (when the first 65 nescent groups, and the like. In some embodiments of the GPCR requires agonist for desensitization), and to an agonist invention, the marker molecules are optically detectable, for the second GPCR (when the second GPCR requires ago including, for example, optically detectable proteins, such US 8,691,256 B2 65 66 that they can be excited chemically, mechanically, electri with the composition. In some embodiments, repellant effect cally, or radioactively to emit fluorescence, phosphorescence, is an effect wherein at least about 90% of insects are repelled or bioluminescence. Optically detectable marker molecules away from a host or area that has been treated with the can include, for example, beta-galactosidase, firefly composition. luciferase, bacterialluciferase, fluorescein, Texas Red, horse “Pesticidal effect” is an effect wherein treatment with a radish peroxidase, alkaline phosphatase, rhodamine-conju composition causes at least about 1% of the insects to die. In gated antibody, and the like. In other embodiments, the opti this regard, an LC to LCoo (lethal concentration) or an LD cally detectable marker molecules can be inherently to LDoo (lethal dose) of a composition will cause a pesticidal fluorescent molecules, such as fluorescent proteins, includ effect. In some embodiments, the pesticidal effect is an effect ing, for example, Green Fluorescent Protein (GFP), and the 10 wherein treatment with a composition causes at least about like. 5% of the exposed insects to die. In some embodiments, the In certain embodiments of the invention, all forms of arres pesticidal effect is an effect wherein treatment with a compo tin, both naturally occurring and engineered variants, includ sition causes at least about 10% of the exposed insects to die. ing, for example, visual arrestin, beta-arrestin 1, beta-arrestin In some embodiments, the pesticidal effect is an effect 2, and the like, can be used. Confocal microscopy can be used 15 wherein treatment with a composition causes at least about to identify such protein-protein interaction and also to study 25% of the insects to die. In some embodiments the pesticidal the trafficking of the protein complex. effect is an effect wherein treatment with a composition In some embodiments, the cell can be transfected with causes at least about 50% of the exposed insects to die. In DNA so that the conjugate of arrestin and a marker molecule some embodiments the pesticidal effect is an effect wherein can be produced within the cell. treatment with a composition causes at least about 75% of the GPCRs used in embodiments of the invention can also be exposed insects to die. In some embodiments the pesticidal conjugated with a marker molecule. In some embodiments, effect is an effect wherein treatment with a composition the carboxyl-terminus of the GPCR can be conjugated with a causes at least about 90% of the exposed insects to die. marker molecule. A carboxyl-terminal tail conjugated or “Disablement' is an effect wherein insects are mobility attached to a marker molecule can be used in a carboxyl 25 impaired such that their mobility is reduced as compared to terminal tail exchange to provide a modified GPCR. insects that have not been exposed to the composition. In In some embodiments of the invention, the GPCRs can be some embodiments, disablement is an effect wherein at least antibody-labeled, for example with an antibody conjugated to about 75% of insects are mobility-impaired such that their an immunofluorescence molecule, or the like, or the GPCRs mobility is reduced as compared to insects that have not been can be conjugated with, for example, a luminescent donor or 30 exposed to the composition. In some embodiments, disable the like. In some embodiments, the GPCRs can be conjugated ment is an effect wherein at least about 90% of insects are with, for example luciferase, Renilla luciferase, or the like. mobility-impaired such that their mobility is reduced as com Embodiments of the invention can be used to evaluate the pared to insects that have not been exposed to the composi effect of a test compound on GPCRR/D by measuring intra tion. In some embodiments, disablement can be caused by a cellular second messenger generation. Intracellular effectors 35 disabling effect at the cellular or whole-organism level. can include, for example, cAMP, cyclic GMP calcium, phos Embodiments of the invention can be used to control para phatidylinositol, a hydrogen ion, an ion transport molecule, sites. As used herein, the term "parasite' includes parasites, and the like. Additionally, enzymes such as, for example, Such as but not limited to, protozoa, including intestinal pro adenylyl cyclase, phosphodiesterase, phospholipase C, pro tozoa, tissue protozoa, and blood protozoa. Examples of tein kinase, phospholipase A, and the like, can be measured 40 intestinal protozoa include, but are not limited to: Entamoeba to gauge the effects of test compounds on GPCR R/D. hystolytica, Giardia lamblia, Cryptosporidium muris, and Controlling Pests Cryptosporidium parvum. Examples of tissue protozoa Embodiments of the invention can be used to control insect include, but are not limited to: Trypanosomatida gambiense, species belonging to orders Acari. Anoplura, Araneae, Blat Trypanosomatida rhodesiense, Trypanosomatida crusi, todea, Coleoptera, Collembola, Diptera, Grylloptera, Het 45 Leishmania mexicana, Leishmania braziliensis, Leishmania eroptera, Homoptera, Hymenoptera, Isopoda, Isoptera, Lepi tropica, Leishmania donovani, Toxoplasma gondii, and Tri doptera, Mantodea, Mallophaga, Neuroptera, Odonata, chomonas vaginalis. Examples of blood protozoa include, but Orthoptera, Psocoptera, Siphonaptera, Symphyla, Thysa are not limited to Plasmodium vivax, Plasmodium ovale, nura, and Thysanoptera. Plasmodium malariae, and Plasmodium falciparum. Histo Embodiments of the present invention can be used to con 50 monas meleagridis is yet another example of a protozoan trol, for example, the insects set forth on pages 123-195 of parasite. WIPO Publication No. WO/2008/088827, published on Jul. As used herein, the term “parasite' further includes, but is 24, 2008. not limited to: helminthes or parasitic worms, including For purposes of simplicity, the term “insect' shall be used nematodes (round worms) and platyhelminthes (flat worms). throughout this application; however, it should be understood 55 Examples of nematodes include, but are not limited to: animal that the term “insect” refers, not only to insects, but also to and plant nematodes of the adenophorea class, such as the arachnids, larvae, and like invertebrates. Also for purposes of intestinal nematode Trichuris trichiura (whipworm) and the this application, the term “insect control' shall refer to having plant nematode Trichodorus obtusus (stubby-root nema a repellant effect, a pesticidal effect, or both. tode); intestinal nematodes of the secementea class, such as “Target pest” refers to the organism that is the subject of the 60 Ascaris lumbricoides, Enterobius vermicularis (pinworm), insect control effort. Ancylostoma duodenale (hookworm), Necator americanus “Repellant effect” is an effect wherein more insects are (hookworm), and Strongyloides Stercoralis; and tissue nema repelled away from a host or area that has been treated with todes of the secementea class, such as Wuchereria bancrofti the composition than a control host or area that has not been (Filaria bancrofti) and Dracunculus medinensis (Guinea treated with the composition. In some embodiments, repel 65 worm). Examples of plathyeminthes include, but are not lim lant effect is an effect wherein at least about 75% of insects ited to: Trematodes (flukes), including blood flukes, such as are repelled away from a host or area that has been treated Schistosoma mansoni (intestinal Schistosomiasis), Schisto US 8,691,256 B2 67 68 Soma haematobium, and Schistosoma japonicum; liver Another exemplary dispenser of a system of the present flukes, such as Fasciola hepatica, and Fasciola gigantica; invention can deliver an insect control composition to a intestinal flukes, such as Heterophyes heterophyes; and lung desired area. The dispenser can include a sealed pouch that flukes such as Paragonimus westermani. Examples of plath can be constructed from a material that is impermeable to the eminthes further include, but are not limited to: Cestodes insect control composition, for example, a metallic foil, a (tapeworms), including Taenia solium, Taenia saginata, polymeric material, or the like. The pouch can define a Vol Hymenolepis mana, and Echinococcus granulosus. ume for holding the insect control composition. The compo Furthermore, the term “parasite' further includes, but is sition can be provided in a material disposed within the Vol not limited to those organisms and classes of organisms set ume of the pouch, for example, a sponge, a cloth Saturated 10 with the material, or the like. When it becomes desirable to forth on pages 196-205 of WIPO Publication No. WO/2008/ place the exemplary system into use, the pouch can be 088827, published on Jul. 24, 2008, or the like. unsealed, exposing the composition for release into the atmo Embodiments of the invention can be used to prevent or sphere or for application to a desired area. treat the parasite hosts set forth on pages 205-232 of WIPO In certain embodiments the insect control composition is Publication No. WO/2008/088827, published on Jul. 24, 15 provided in a saturated cloth within the pouch, which can be 2008, or the like. used to apply the control composition a desired area. For Embodiments of the invention can be used to treat crops in example, a desired area can be an animal. Such as a human, a order to limit or prevent insect infestation. The types of crops domestic animal, Surfaces within a dwelling, an outdoor liv that can be treated can include, for example, those listed on ing area, or the like. pages 232-239 of WIPO Publication No. WO/2008/088827, In certain embodiments, the dispenser can further include a published on Jul. 24, 2008, or the like. hook, allowing the pouch and exposed control composition to In certain embodiments of the invention, an area can be be hung in a desired location, Such as in a closet or a pantry. treated with a composition of the present invention, for In certain embodiments, a method of the present invention example, by using a spray formulation, Such as an aerosol or can deliver insectan control composition to a desired area. In a pump spray, or a burning formulation, such as a candle or a 25 certain embodiments, a dispenser used with the method can piece of incense containing the composition, or the like. In be constructed from a substantially planar, integral piece of certain embodiments of the invention, an area can be treated, material, having a first side that is coated with control com for example, via aerial delivery, by truck-mounted equip position, and a second side that is not coated with control ment, or the like. Ofcourse, various treatment methods can be composition. The integral piece of material can be folded and used without departing from the spirit and scope of the 30 sealed such that the side coated with the control composition present invention. For example, compositions can be com is contained within the Volume defined by the sealed pouch. prised in household products, for example, hard surface When the pouch is unsealed, the side that is coated with cleaners, and the like. control composition is exposed. The Substantially planar An exemplary dispenser of a system of the present inven piece of material can be placed in a desired location to deliver tion can deliveran pest control composition to the atmosphere 35 control composition to the atmosphere, or to crawling insects in a continuous manner over a period of time. The exemplary that walk across the material. dispenser can include a reservoir for holding a pest control Another exemplary dispenser of a system of the present composition, and a wick for drawing the composition from invention can deliver an insect control composition to a the reservoir and releasing the insect control composition into desired area. The control composition can be incorporated the atmosphere. The reservoir can be constructed from a 40 into an appropriate material. In certain embodiments, the material that is impermeable to the pest control composition, composition-containing material can be a material that is for example, appropriate glass, ceramic, or polymeric mate capable of controlling the release rate of the control compo rials can be used. The reservoir can include an aperture, which sition, i.e., controlled-release material, allowing the control can be sealed or unsealed, as desired. When the exemplary composition to be released into the atmosphere at a desired system of the present invention is not in use, the aperture can 45 rate that can be adjusted by providing controlled-release be sealed to prevent the release of the pest control composi material having appropriate specifications. The controlled tion into the atmosphere. It may be desirable, for example, to release material can be constructed from an appropriate poly seal the aperture when the exemplary system is being stored mer. In other embodiments the composition-containing mate or transported. When the system is in use, the aperture is rial does not allow the control composition to be released into unsealed, such that the wick can draw the pest control com 50 the atmosphere, but rather retains the control composition. An position from the reservoir, and release the control composi optional casing that is impermeable to the insect control com tion through the aperture into the atmosphere. position can be provided to hold the composition-containing In certain embodiments of the invention, the rate of release material until the system is ready for use. When the system is of the composition can be controlled, for example, by making ready for use, the casing can be peeled away, exposing the adjustments to the wick of the dispenser. For example, the 55 composition-containing material. The composition-contain Surface area of the wick that is exposed to the atmosphere can ing material can be placed in a desired location to deliver be altered. Generally, the greater the exposed surface area, the control composition to crawling insects that walk across the greater the rate of release of the pest control composition. In material, or to deliver the control composition to the atmo this regard, in certain embodiments, the dispenser can include sphere when a controlled-release material is used, e.g., con multiple wicks and the reservoir can include multiple aper 60 trol flying insects. tures through which the insect control composition can be In certain embodiments, the composition-containing mate released into the atmosphere. As another example, the wick rial can have a substantially planar design, appropriate for can be constructed from a particular material that draws the positioning adjacent a mattress for controlling bedbugs, e.g., pest control composition from the reservoir and releases it Cimex lectularius. A Substantially planar design can also be into the environment at a desired rate. Such as, for example, a 65 used, for example, as or with a picnic table cloth. In certain wick made of wood, a wick made of a synthetic fiber, or the embodiments, the composition-containing material can be like. used as ground cover for a garden bed or adjacent crop plants US 8,691,256 B2 69 70 to control weeds. In certain embodiments, the composition Embodiments of the invention can result in agricultural containing material can take the shape of a bag, and could be improvements, such as, for example, increased crop yield, used for trash collection, while controlling insect commonly reduced frequency of application of pest control product, attracted to household garbage or other trash. reduced phytotoxicity associated with the pesticide, reduced Another exemplary dispenser of a system of the present cost or increased value associated with at least one environ invention can be a Substantially dry sheet containing the con mental factor, and the like. trol composition, which control composition can be applied to a desired location upon exposing the cloth to water or an In embodiments of the invention that can reduce the cost of, aqueous liquid, e.g., perspiration. In certain embodiments, or increase the value associated with at least one environmen the dry sheet containing the control composition can dissolve 10 tal factor, the environmental factor can include, for example, into a cream or gel when exposed to water or an aqueous air quality, water quality, Soil quality, detectable pesticide liquid, which can then be applied to a desired area. For residue, safety or comfort of workers, collateral effect on a example, a desired area can be an animal. Such as a human, a non-target organism, and the like. domestic animal, or another animal. Embodiments of the present invention can be used to con The following references are incorporated herein by this 15 trol pests by either treating a host directly, or treating an area reference: U.S. Pat. No. 6,610,254 to Furner et al., issued where the host will be located. For purposes of this applica Aug. 26, 2003, entitled “Dual Function Dispenser U.S. Pat. tion, host is defined as a plant, human or other animal. The No. 6,360,477 to Flashinski et al., issued Mar. 26, 2002, host can be treated, for example, directly by using a cream or entitled “Insect Control Pouch, U.S. Pat. No. 5,980,931 to spray formulation, that can be applied externally or topically, Fowler et al., issued Nov. 9, 1999, entitled “Cleansing Prod 20 when appropriate in light of the specific composition being ucts Having a Substantially Dry Substrate. U.S. Pat. No. used, e.g., to the skin of a human. A composition can be 4.320,113 to Kydonieus, issued Mar. 16, 1982, entitled “Pro applied to the host, for example, in the case of a human, using cess for Controlling Cockroaches and Other Crawling formulations of a variety of personal products or cosmetics Insects. U.S. Pat. No. 4,943,435 to Baker et al., issued Jul. for use on the skin or hair. For example, any of the following 24, 1990, entitled “Prolonged Activity Nicotine Patch.” 25 can be used, when appropriate in light of the specific compo United States Patent Publication No. 2004/0185080 to Hojo, sition being used: fragrances, colorants, pigments, dyes, etal, entitled “Sustained Release Dispenser Comprising Two colognes, skin creams, skin lotions, deodorants, talcs, bath or More Sex Pheromone Substances and a Pest Control oils, Soaps, shampoos, hair conditioners and styling agents. Method, PCT Publication No. WO/2006/061803 to Firmen The present invention is further illustrated by the following ich, etal, entitled “A Device for Dispensing a Volatile Liquid 30 examples. and Method for its Activation, and PCT Publication No. WO/2004/006968 to Firmenich, et al., entitled “A Device for Dispensing Active Volatile Liquid.” EXAMPLES Treatment can include, for example, use of a oil-based formulation, a water-based formulation, a residual formula 35 Test compositions are provided, including a first agent tion, and the like. In some embodiments, combinations of comprising a blend selected from Table 5 (below) and a formulations can be employed to achieve the benefits of dif second agent comprising a pest control chemical or a syner ferent formulation types. gist. TABLE 5

BLENDS CAS Registry Compounds Number low % high % Blend 1 LFO 4 30 D-Limonene S989-27-5 8 99 Thyme Oil White 8007-46-3 O. 2O Blend 65 8 99 Blend 2 D-Limonene S989-27-5 9 99 Thyme Oil White 8007-46-3 O. 2O Linalool Coeur 78-70-6 O. 4 Tetrahydrolinalool 78-69-3 O. 5 Vanillin 121-33-5 O.O6 O.3 Sopropyl myristate 110-27-0 O. 5 Piperonal (aldehyde) Heliotropine 120-57-O O. 5 Blend 66 8 99 Geraniol Fine FCC 106-24-1 O. 4 Triethyl Citrate 77-93-0 O. 5 Blend 3 D-Limonene S989-27-5 45 99 Thyme Oil White 8007-46-3 O. 10 Blend 66 5 30 Blend 63 O. 10 Blend 4 LFO 30 99 BSO 977O17-84-7 15 99 Blend 5 BSO 977O17-84-7 15 99 Linalool Coeur 78-70-6 6 40 Tetrahydrolinalool 78-69-3 8 45 Vanillin 121-33-5 O.1 5 Sopropyl myristate 110-27-0 10 55 Piperonal (aldehyde) Heliotropine 120-57-O O.1 2O Geraniol Fine FCC 106-24-1 O.1 25 US 8,691,256 B2 71 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high% Blend 6 D-Limonene S989-27-5 O.1 25 BSO 977O17-84-7 15 85 Linaloo Coeur 78-70-6 O.1 25 Tetrahydrolinalool 78-69-3 O.1 25 Vanillin 121-33-5 O.1 3 Isopropyl myristate 110-27-0 O.1 30 Piperonal (aldehyde) Heliotropine 120-57-O O.1 10 Geraniol Fine FCC 106-24-1 O.1 15 Methyl Salicylate 98% Nat 119-36-8 8 70 Blend 7 Thyme Oil White 8007-46-3 15 90 Wintergreen Oil 68917-75-9 15 99 Vanillin 121-33-5 O.1 4 Isopropyl myristate 110-27-0 2O 99 Blend 8 D-Limonene S989-27-5 2O 99 Thyme Oil White 8007-46-3 O.1 25 Wintergreen Oil 68917-75-9 25 99 Blend 9 LFO 6 40 D-Limonene S989-27-5 25 99 Thyme Oil White 8007-46-3 5 30 Linaloo Coeur 78-70-6 O.1 3 Citral S392-40-5 O.1 2O gamma-terpinene 99-85-4 O.1 2O Alpha-Pinene, 98% 80-56-8 O.1 5 alpha-Terpineol 98-55-S O.1 15 Terpinolene 586-62-9 O.1 15 Para-Cymene 99-87-6 O.1 5 LinallylAcetate 115-95-7 O.1 6 Beta Pinene 127-91-3 O.1 6 Camphor Dextro 464-49-3 O.OS O.3 Terpinene 4 OL S62-74-3 O.OS O.3 Alpha Terpinene 99-86-5 O.1 6 Borneol L 507-70-0 O.1 3 Camphene 79-92-5 O.1 2 Decanal 112-31-2 O.O6 O.3 Dodecanal 112-54-9 O.O6 O.3 Fenchol Alpha 512-13-0 O.OOS O.1 GeranylAcetate 105-87-3 O.O6 O.3 Isoborneo 124-76-5 O.08 1 2-Methyl 1,3-cyclohexadiene 30640-46-1, O.08 1 1888-90-0 Myrcene 123-35-3 O.1 3 Nonanal 124-19-6 O.OOS O.08 Octanal 124-13-0 O.OOS O.2 Tocopherol Gamma (TENOX (R) 54-28-4 O.OOS O.08 Blend 10 D-Limonene S989-27-5 O.1 25 Thyme Oil White 8007-46-3 O.1 25 Blend 65 40 99 Linaloo Coeur 78-70-6 O.1 6 Tetrahydrolinalool 78-69-3 O.1 8 Vanillin 21-33-5 O.08 O.6 Isopropyl myristate 10-27-0 O.1 8 Piperonal (aldehyde) Heliotropine 20-57-O O.1 8 Geraniol Fine FCC 06-24-1 O.1 4 Triethyl Citrate 77-93-0 O.1 8 Blend 11 Thyme Oil White 8007-46-3 3 65 Wintergreen Oil 68917-75-9 15 99 sopropyl myristate 10-27-0 2O 99 Blend 12 D-Limonene S989-27-5 5 30 Linalool Coeur 78-70-6 8 40 Tetrahydrolinalool 78-69-3 15 99 Vanillin 21-33-5 O.1 8 sopropyl myristate 10-27-0 15 85 Piperonal (aldehyde)Heliotropine 20-57-O 5 30 Geraniol Fine FCC 06-24-1 5 30 Blend 13 D-Limonene S989-27-5 5 30 Geraniol Fine FCC 06-24-1 5 30 Blend 62 50 99 Blend 14 D-Limonene S989-27-5 5 30 Blend 72 55 99 Blend 15 D-Limonene S989-27-5 5 30 Linalool Coeur 78-70-6 10 55 Tetrahydrolinalool 78-69-3 10 65 Vanillin 121-33-5 O.1 4 sopropyl myristate 110-27-0 10 60 US 8,691,256 B2 73 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high% Piperonal (aldehyde)Heliotropine 120-57-O 10 65 Piperonyl Alcohol 495-76-1 O. 25 Blend 16 D-Limonene S989-27-5 5 30 BSO 977O17-84-7 15 8O Linalool Coeur 78-70-6 5 30 Tetrahydrolinalool 78-69-3 6 35 Vanillin 121-33-5 O. 4 Mineral Oil White (USP) 8.042-47-5 8 45 sopropyl myristate 110-27-0 8 45 Piperonal (aldehyde)Heliotropine 120-57-O O. 15 Geraniol Fine FCC 106-24-1 O. 2O Blend 17 D-Limonene S989-27-5 10 99 Linalool Coeur 78-70-6 O. 10 Tetrahydrolinalool 78-69-3 O. 10 Vanillin 121-33-5 O.08 O6 sopropyl myristate 110-27-0 O. 10 Piperonal (aldehyde)Heliotropine 120-57-O O. 10 Piperonyl Alcohol 495-76-1 O. 5 Blend 66 10 99 Blend 18 Linaloo Coeur 78-70-6 O. 15 Tetrahydrolinalool 78-69-3 O. 2O Vanillin 121-33-5 O. 2 sopropyl myristate 110-27-0 O. 2O Piperonal (aldehyde)Heliotropine 120-57-O O. 2O Piperonyl Alcohol 495-76-1 O. 10 Blend 66 40 99 Blend 19 LFO 2O 99 D-Limonene S989-27-5 15 85 Thyme Oil White 8007-46-3 15 90 Blend 20 D-Limonene S989-27-5 15 85 Thyme Oil White 8007-46-3 15 95 Blend 63 2O 99 Blend 21 D-Limonene S989-27-5 15 85 Thyme Oil White 8007-46-3 15 90 Linalool Coeur 78-70-6 O. 15 Tetrahydrolinalool 78-69-3 O. 25 Vanillin 121-33-5 O. 2 sopropyl myristate 110-27-0 O. 25 Piperonal (aldehyde)Heliotropine 120-57-O O. 25 Geraniol Fine FCC 106-24-1 O. 10 Triethyl Citrate 77-93-0 O. 25 Blend 22 Phenyl Ethyl Propionate 2O 99 Methyl Salicylate 2O 99 Blend 43 15 85 Blend 23 D-Limonene S989-27-5 O. 10 Thyme Oil White 8007-46-3 O. 15 Benzyl Alcohol 100-51-6 8 50 sopar M 64742-47-8 10 65 Water 7732-18-5 25 99 Blend 63 O. 15 Stock 10% SLS Solution O. 10 Blend 24 D-Limonene S989-27-5 O. 10 Thyme Oil White 8007-46-3 O. 15 Linalool Coeur 78-70-6 O. 3 Tetrahydrolinalool 78-69-3 O. 4 Vanillin 121-33-5 O.OS O.3 sopropyl myristate 110-27-0 O. 4 Piperonal (aldehyde)Heliotropine 120-57-O O. 4 Geraniol Fine FCC 106-24-1 O. 2 Triethyl Citrate 77-93-0 O. 4 Benzyl Alcohol 100-51-6 8 50 sopar M 64742-47-8 10 65 Water 7732-18-5 25 99 Stock 10% SLS Solution O. 10 Blend 25 D-Limonene S989-27-5 6 40 Thyme Oil White 8007-46-3 8 45 Benzyl Alcohol 100-51-6 30 99 Blend 63 10 55 Blend 26 LFO O.1 25 D-Limonene S989-27-5 8 99 Thyme Oil White 8007-46-3 O.1 2O Blend 66 8 99 Blend 27 Linaloo Coeur 78-70-6 O.1 2O Soy Bean Oil 8O1 6-70-4 10 70 Thymol (crystal) 89-83-8 2O 99 US 8,691,256 B2 75 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high% Alpha-Pinene, 98% 80-56-8 O. 10 Para-Cymene 99-87-6 15 85 Blend 28 Linaloo Coeur 78-70-6 O. 25 Thymol (crystal) 89-83-8 25 99 Alpha-Pinene, 98% 80-56-8 O. 15 Para-Cymene 99-87-6 2O 99 Blend 29 D-Limonene S989-27-5 O. 25 Thyme Oil White 8007-46-3 O. 30 Blend 65 35 99 Linaloo Coeur 78-70-6 O. 8 Tetrahydrolinalool 78-69-3 O. 10 Vanillin 121-33-5 O.08 1 Isopropyl myristate 110-27-0 O. 10 Piperonal (aldehyde) Heliotropine 120-57-O O. 5 Geraniol Fine FCC 106-24-1 O. 5 Blend 30 D-Limonene S989-27-5 15 85 Thyme Oil White 8007-46-3 O. 15 Methyl Salicylate 35 99 Blend 31 Thyme Oil White 8007-46-3 O. 5 Wintergreen Oil 68917-75-9 O. 8 sopropyl myristate 110-27-0 O. 6 Span 80 1338-43-8 O. 2 sopar M 64742-47-8 8 45 Water 7732-18-5 40 99 Bifenthrin 83657-04-3 O.OOS O.2 Blend 32 Castor Oil hydrogenated - PEO40 30 99 Lemon Grass Oil - India 10 70 Blend 1 10 70 Blend 33 LFO 8 50 D-Limonene S989-27-5 35 99 Thyme Oil White 8007-46-3 6 35 BSO 977O17-84-7 O.1 15 Blend 34 D-Limonene S989-27-5 O.1 25 Thyme Oil White 8007-46-3 O.1 30 Blend 65 30 99 Linalool Coeur 78-70-6 O.1 5 Tetrahydrolinalool 78-69-3 O.1 8 Vanillin 21-33-5 O.O6 O.S sopropyl myristate 10-27-0 O.1 8 Piperonal (aldehyde)Heliotropine 20-57-O O.1 8 Geraniol Fine FCC 06-24-1 O.1 4 Triethyl Citrate 77-93-0 O.1 8 Sopar M 64742-47-8 8 40 Blend 35 sopropyl myristate 10-27-0 2O 99 Wintergreen Oil 25 99 Blend 68 10 60 Blend 36 Wintergreen Oil 68917-75-9 25 99 sopropyl myristate 10-27-0 2O 99 Thyme Oil Red 8007-46-3 10 60 Blend 37 Wintergreen Oil 68917-75-9 25 99 Vanillin 21-33-5 O.O6 O.3 sopropyl myristate 10-27-0 2O 99 Thyme Oil Red 8007-46-3 10 60 Blend 38 Thyme Oil White 8007-46-3 15 95 sopropyl myristate 10-27-0 25 99 Geraniol Fine FCC 06-24-1 10 70 Blend 39 sopropyl myristate 10-27-0 25 99 Geraniol Fine FCC 06-24-1 10 70 Thyme Oil White 8007-46-3 2O 99 Blend 40 Orange Terpenes 68647-72-3 O.1 25 Blend 68 O.1 30 Blend 69 35 99 Blend 71 6 40 Blend 41 Linalool Coeur 78-70-6 10 70 Amyl Butyrate 540-18-1 10 70 Anise Star Oil 30 99 Blend 42 Thyme Oil White 8007-46-3 15 75 Amyl Butyrate 540-18-1 10 70 Anise Star Oil 30 99 Blend 43 Tetrahydrolinalool 78-69-3 10 70 Vanillin 121-33-5 O.1 4 Hercolyn D 8OSO-15-S O.1 15 Isopropyl myristate 110-27-0 8 45 Piperonal (aldehyde) Heliotropine 120-57-O O.1 25 Ethyl Linalool 10339-55-6 10 70 US 8,691,256 B2 77 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high%

Hedione 24851-98-7 O.1 2O Triethyl Citrate 77-93-0 5 30 Dipropylene glycol (DPG) 246-770-3 O.1 25 Blend 44 Blend 63 25 99 Thyme Oil White 30 99 Blend 45 Linalool coeur 78-70-6 O.1 2O Tetrahydrolinalool 78-69-3 O.1 25 Vanillin 121-33-5 O.1 2 sopropyl myristate 110-27-0 O.1 30 Piperonal (aldehyde)Heliotropine 120-57-O O.1 30 Geraniol Fine FCC 106-24-1 O.1 15 Triethylcitrate 77-93-0 O.1 30 Thyme Oil White 30 99 Blend 46 Phenyl Ethyl Propionate O 55 Benzyl Alcohol 100-51-6 30 99 Methyl Salicylate O 55 Blend 43 8 40 Blend 47 Thyme Oil White 8007-46-3 5 75 Amyl Butyrate 540-18-1 O 70 Anise Star Oil 30 99 Genistein O.OOS O.1 Blend 48 Linalool coeur 78-70-6 O 70 Amyl Butyrate 540-18-1 O 70 Anise Star Oil 30 99 Thyme Oil White O.OOS O.1 Blend 49 LFO O 70 BSO 977O17-84-7 O 70 Benzyl Alcoho 100-51-6 30 99 Blend SO sopropyl myristate 110-27-0 O 70 Wintergreen oi 5 90 Thyme oil white 8 40 Myristicin 5 99 Blend S1 sopropyl myristate 110-27-0 5 8O Wintergreen oi 5 95 sopropyl alcohol 67-63-0 O.1 10 Thyme oil white 8 40 Myristicin 5 75 Blend 52 Isopropyl myristate 110-27-0 2O 99 Wintergreen oi 25 99 Thyme oil white O 60 Genistein O.OO1 O.1 Blend 53 sopropyl myristate 110-27-0 2O 99 Wintergreen oi 2O 99 sopropyl alcohol 67-63-0 5 30 Thyme oil white 8 50 Genistein O.OO1 O.1 Blend 54 Isopropyl myristate 110-27-0 O 70 Wintergreen oi 5 90 Thyme oil white 8 40 Genistein O.OO1 O.1 Myristicin 5 99 Blend 55 Mineral oil white 8.042-47-5 2O 99 Wintergreen oi 25 99 Thyme oil white O 60 Blend 56 Mineral oil white 8.042-47-5 O 50 Wintergreen oi O 65 Thyme oil white 5 30 Benzaldehyde 30 99 Blend 57 Mineral oil white 8.042-47-5 O 55 Wintergreen oi O 65 Thyme oil white 5 30 Genistein 5 75 Benzaldehyde 5 8O Blend 58 Linaloo Coeur 78-70-6 4 65 Thymol (crystal) 89-83-8 2O 99 Alpha-Pinene, 98% 80-56-8 1 10 Para-Cymene 99-87-6 1 55 Trans-Anethole 418O-23-8 10 55 Blend 59 Linaloo Coeur 78-70-6 O.1 30 Thymol (crystal) 89-83-8 25 99 Alpha-Pinene, 98% 80-56-8 O.1 30 Para-Cymene 99-87-6 15 99 Blend 60 Soy Bean Oil 8O1 6-70-4 15 75 Alpha-Pinene, 98% 80-56-8 O.1 10 Para-Cymene 99-87-6 15 85 US 8,691,256 B2 79 TABLE 5-continued

BLENDS CAS Registry Compounds Number ow % high% LinallylAcetate 15-95-7 O.1 2O Thymol acetate S28-79-0 2O 99 Blend 61 Alpha-Pinene, 98% 80-56-8 O.1 30 Para-Cymene 99-87-6 O 55 LinallylAcetate 15-95-7 O 70 Thymol acetate S28-79-0 30 99 Blend 62 Linaloo Coeur 78-70-6 O 60 Tetrahydrolinalool 78-69-3 O 70 Vanillin 21-33-5 O.1 8 sopropyl myristate 10-27-0 5 90 Piperonal (aldehyde)Heliotropine 20-57-O 5 30 Geraniol Fine FCC 06-24-1 8 40 Blend 63 Linaloo Coeur 78-70-6 8 40 Tetrahydrolinalool 78-69-3 O 55 Vanillin 21-33-5 O.1 4 sopropyl myristate 10-27-0 O 55 Piperonal (aldehyde)Heliotropine 20-57-O O 55 Geraniol Fine FCC 06-24-1 5 30 Triethyl Citrate 77-93-0 O 55 Blend 64 Linaloo Coeur 78-70-6 O 60 Tetrahydrolinalool 78-69-3 O 70 Vanillin 21-33-5 O. 4 sopropyl myristate 10-27-0 O 70 Piperonal (aldehyde)Heliotropine 20-57-O O 70 Piperonyl Alcohol 495-76-1 O. 30 Blend 65 D-Limonene S989-27-5 25 99 Linalool Coeur 78-70-6 O. 4 Citral S392-40-5 5 30 gamma-terpinene 99-85-4 5 30 Alpha-Pinene, 98% 80-56-8 O. 6 alpha-Terpineol 98-55-S O. 2O Terpinolene 586-62-9 O. 2O Para-Cymene 99-87-6 O. 5 LinallylAcetate 115-95-7 O. 8 Beta Pinene 127-91-3 O. 10 Camphor Dextro 464-49-3 O.O6 O.3 Terpinene 4 OL S62-74-3 O.O6 O.3 Alpha Terpinene 99-86-5 O. 10 Borneol L 507-70-0 O. 5 Camphene 79-92-5 O. 2 Decanal 112-31-2 O.08 O6 Dodecanal 112-54-9 O.O6 O.3 Fenchol Alpha 512-13-0 O.OO1 O.1 GeranylAcetate 105-87-3 O.08 O6 Isoborneo 124-76-5 O.1 2 2-Methyl 1,3-cyclohexadiene 30640-46-1, O.1 2 1888-90-0 Myrcene 123-35-3 O.1 4 Nonanal 124-19-6 O.OO1 O.1 Octanal 124-13-0 O.OS O.2 Tocopherol Gamma (TENOX (R) 54-28-4 O.OO1 O.1 Blend 66 D-Limonene S989-27-5 30 99 Linalool Coeur 78-70-6 O.1 5 gamma-terpinene 99-85-4 6 40 Alpha-Pinene, 98% 80-56-8 O.1 8 Terpinolene 586-62-9 O.1 25 Para-Cymene 99-87-6 O.1 6 LinallylAcetate 115-95-7 O.1 10 Beta Pinene 127-91-3 O.1 10 Camphor Dextro 464-49-3 O.1 10 Terpinene 4 OL S62-74-3 O.O6 O.3 Alpha Terpinene 99-86-5 O.08 O6 Borneol L 507-70-0 O.1 5 Camphene 79-92-5 O.1 3 Decanal 112-31-2 O.08 O6 Dodecanal 112-54-9 O.08 O6 Fenchol Alpha 512-13-0 O.OO1 O.1 GeranylAcetate 105-87-3 O.08 O6 Soborneo 124-76-5 O.1 2 2-Methyl 1,3-cyclohexadiene 30640-46-1, O.1 2 1888-90-0 Myrcene 123-35-3 O.1 5 Nonanal 124-19-6 O.OO1 O.2 Octanal 124-13-0 O.OS O.3 Tocopherol Gamma (TENOX (R) 54-28-4 O.OO1 O.2 US 8,691,256 B2 81 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high% Blend 67 D-Limonene S989-27-5 2O 99 Linalool Coeur 78-70-6 5 30 Alpha-Pinene, 98% 80-56-8 O. 15 Terpinolene 586-62-9 5 30 Para-Cymene 99-87-6 5 30 LinallylAcetate 115-95-7 O. 15 Beta Pinene 127-91-3 O. 15 Alpha Terpinene 99-86-5 O. 15 Camphene 79-92-5 O. 2O Myrcene 123-35-3 O. 30 Blend 68 D-Limonene S989-27-5 O.08 1 Thyme Oil Red 8007-46-3 O. 4 Thymol (crystal) 89-83-8 30 99 alpha-Terpineol 98-55-S O. 6 Para-Cymene 99-87-6 10 60 LinallylAcetate 115-95-7 O. 5 Caryophyllene-B 87-44-5 O. 10 Borneol L 507-70-0 O. 6 Myrcene 123-35-3 O. 4 Tea Tree Oil O. 6 Cypress Oil O. 10 Peppermint Terpenes 8006-90-4 O. 30 Linalool 90 O. 10 Blend 69 D-Limonene S989-27-5 30 99 Citral S392-40-5 O. 25 gamma-terpinene 99-85-4 5 30 Alpha-Pinene, 98% 80-56-8 O. 5 alpha-Terpineol 98-55-S O. 15 Terpinolene 586-62-9 O. 2O Lime Distilled Oil O.O6 O.3 Lime Expressed Oil O.O6 O.3 Linallyl Acetate 115-95-7 O. 6 Caryophyllene-B 87-44-5 O.O6 O.3 Beta Pinene 127-91-3 O. 8 Terpinene 4 OL S62-74-3 O.OOS O.2 Alpha Terpinene 99-86-5 O. 6 Borneol L 507-70-0 O. 5 Camphene 79-92-5 O. 2 GeranylAcetate 105-87-3 O.08 O6 Soborneo 124-76-5 O.O6 O.3 Linalool 90 O. 3 Camphor Gum O.OOS O.2 Aldehyde C-10 O.OOS O.2 Aldehyde C-12 O.O6 O.3 Blend 70 Eugenol 97-53-0 O.OO3 O.1 Eucalyptol (1.8 Cineole) O.OS O.3 Methyl Salicylate 60 99.9 Linalool 90 O.OS O.3 Ethyl Salicylate O.OS O.3 Blend 71 Tetrahydrolinalool 78-69-3 6 35 Hercolyn D 8OSO-15-S O.1 25 sopropyl myristate 110-27-0 O.1 2O Piperonal (aldehyde)Heliotropine 120-57-O 5 30 Ethyl Linalool 10339-55-6 5 30 Triethyl Citrate 77-93-0 O.1 30 Dipropylene glycol (DPG) 246-770-3 5 30 Cinnamic Alcohol 104-54-1 O.1 5 Eugenol 97-53-0 O.1 5 Phenyl Ethyl Alcohol 60-12-8 10 65 so Eugenol O.08 1 Methyl Dihydrojasmonate 5 30 Blend 72 Linaloo Coeur 78-70-6 8 40 Tetrahydrolinalool 78-69-3 10 70 Vanillin 121-33-5 O.1 8 sopropyl myristate 110-27-0 15 85 Piperonal (aldehyde)Heliotropine 120-57-O 5 30 Piperonyl Alcohol 495-76-1 5 30 Geraniol Fine FCC 106-24-1 5 30 Blend 73 Blend 11 50 99 Stock 10% SLS Solution 5 30 Blend 74 Polyglycerol-4-oleate 9007-48-1 O.1 3 Lecithin 8002-43-5 O.08 O6 Water 7732-18-5 5 30 Blend 11 50 99 US 8,691,256 B2 83 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high% Blend 75 Potassium Sorbate 590-OO-1 or O.1 4 24634-61-5 Xanthan Gum 11138-66-2 O.08 1 Water 7732-18-5 45 99 Blend 74 10 50 Blend 76 Potassium Sorbate 590-OO-1 or O.1 2 24634-61-5 Polyglycerol-4-oleate 9007-48-1 O.1 2 Xanthan Gum 11138-66-2 O.08 1 Lecithin 8002-43-5 O.O6 O.3 Water 7732-18-5 2O 99 Blend 11 15 99 Blend 77 Thyme Oil White 8007-46-3 O.1 25 Wintergreen Oil 68917-75-9 2 55 sopropyl myristate 110-27-0 1 40 Potassium Sorbate 590-OO-1 or O.O6 O.3 24634-61-5 Polyglycerol-4-oleate 9007-48-1 O.1 2 Xanthan Gum 11138-66-2 O.08 1 Lecithin 8002-43-5 O.O6 O.3 Water 7732-18-5 2O 99 Blend 78 Polyglycerol-4-oleate 9007-48-1 O.1 3 Lecithin 8002-43-5 O.08 O.6 Water 7732-18-5 5 30 Blend 11 50 99 Blend 79 Water 7732-18-5 O.1 2O Blend 74 40 99 Stock 2.5% Xanthan-1% Ksorbate 6 40 Blend 80 Water 7732-18-5 O.1 10 Blend 78 45 99 Stock 2.5% Xanthan-1% Ksorbate 6 40 Blend 81 Potassium Sorbate 590-OO-1 or O.1 4 24634-61-5 Xanthan Gum 11138-66-2 O.08 1 Water 7732-18-5 45 99 Blend 78 10 50 Blend 82 Blend 1 O.1 8 Water 60 99 Blend 83 Polyglycerol-4-oleate 9007-48-1 O.1 3 Lecithin 8002-43-5 O.08 O.6 Water 7732-18-5 5 30 Blend 11 50 99 Blend 84 Potassium Sorbate 590-OO-1 or O.1 4 24634-61-5 Xanthan Gum 11138-66-2 O.08 1 Water 7732-18-5 45 99 Blend 83 10 50 Blend 85 Citroneia Oil 106-22-9 O.08 O.6 Carbopol 940 9003-01-4) O.08 O.6 BHT (butylated hydroxytoluene) 128-37-0 O.O6 O.3 Water 7732-18-5 30 99 Emulsifying Wax 67762-27-0, 8 40 90OS-67-8 Light Liquid Paraffin 8O12-95-1 O.1 10 White Soft Paraffin 8009-03-8 O.1 25 Sodium Metabisulphate 7681-57-4) O.08 1 Propylene Glycol 57-55-6 O.1 6 Methyl Paraben 99-76-3) O.08 O.6 Propyl Paraben 94-13-3) O.OOS O.2 Cresmer RH40 hydrogenated castor 61791-12-6 O.1 15 oil Triethanolamine 102-71-6) O.08 O.6 Vitamin E. Acetate 58-95-7 O.OO2 O.08 Disodium EDTA 139-33-3 O.OOS O.2 Blend 1 O.1 15 Blend 86 Span 80 1338-43-8 O.OOS O.2 Sodium Benzoate S32-32-1 O.08 O.6 Isopar M 64742-47-8 15 85 A46 Propellant 8 45 Water 7732-18-5 25 99 Isopropyl alcohol 67-63-0 O.1 5 Blend 8 6 40 Blend 87 Isopar M 64742-47-8 30 99 A46 Propellant 2O 99 US 8,691,256 B2 85 86 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high% sopropyl alcohol 67-63-0 10 Blend 25 2O Blend 88 sopar M 64742-47-8 99 A46 Propellant 99 Bifenthrin 83657-04-3 O.2 sopropyl alcohol 67-63-0 10 Blend 25 2O Blend 89 sopar M 64742-47-8 99 A46 Propellant 99 Blend 20 2O Blend 90 Potassium Sorbate 590-OO-1 or O.3 24634-61-5 Polyglycerol-4-oleate 9007-48-1 Xanthan Gum 11138-66-2 Lecithin 8002-43-5 O.1 Water 7732-18-5 99 sopropyl alcohol 67-63-0 Blend 35 45 Blend 91 Potassium Sorbate 590-OO-1 or O.3 24634-61-5 Polyglycerol-4-oleate 9007-48-1 O.08 Xanthan Gum 11138-66-2 O.08 Lecithin 8002-43-5 O.OO3 O.1 Water 7732-18-5 99 Blend 35 40 Blend 92 sopropyl myristate 110-27-0 10 Geraniol Fine FCC 106-24-1 Potassium Sorbate 590-OO-1 or . 24634-61-5 Polyglycerol-4-oleate 9007-48-1 Xanthan Gum 11138-66-2 Lecithin 8002-43-5 O.2 7732-18-5 5 99 10 sopropyl alcohol 67-63-0 Blend 93 Wintergreen Oil 68917-75-9 15 sopropyl myristate 110-27-0 10 Thyme Oil Red 8007-46-3 Stock 0.3% SLS-0.1% Xanthan Soln 99 Blend 94 Stock 0.3% SLS & 0.1% Xanthan 99 Soln Blend 38 15 Blend 95 Lecithin, Soya. 8O3O-76-0 O6 Polyglycerol-4-oleate 9007-48-1 7732-18-5 30 Blend 11 99 Blend 96 Thyme Oil White 8007-46-3 99 sopropyl myristate 110-27-0 95 Lecithin, Soya. 8O3O-76-0 O6 Polyglycerol-4-oleate 9007-48-1 Water 7732-18-5 30 Wintergreen Oil 65 Blend 97 Lecithin, Soya. 8O3O-76-0 O.3 Polyglycerol-4-oleate 9007-48-1 Water 7732-18-5 30 Blend 7 99 Blend 98 Thyme Oil White 8007-46-3 55 Wintergreen Oil 68917-75-9 99 Vanillin 121-33-5 sopropyl myristate 110-27-0 90 Lecithin, Soya. 8O3O-76-0 O.3 Polyglycerol-4-oleate 9007-48-1 Water 7732-18-5 30 Blend 99 Polyglycerol-4-oleate 9007-48-1 Water 7732-18-5 25 Blend 11 99 Blend 100 Thyme Oil White 8007-46-3 99 sopropyl myristate 110-27-0 95 Polyglycerol-4-oleate 9007-48-1 Water 7732-18-5 25 Wintergreen Oil 65 Blend 101 Potassium Sorbate 590-OO-1 or O.3 24634-61-5 Polyglycerol-4-oleate 9007-48-1 Xanthan Gum 11138-66-2 O.08 US 8,691,256 B2 87 88 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high%

Water 7732-18-5 50 99 Blend 97 6 35 Blend 102 D-Limonene S989-27-5 O.1 15 Thyme Oil White 8007-46-3 O.1 Lecithin, Soya. 8O3O-76-0 O.OO1 O.04 Potassium Sorbate 590-OO-1 or O.O6 O.3 24634-61-5 Polyglycerol-4-oleate 9007-48-1 O.1 Xanthan Gum 11138-66-2 O.08 Water 7732-18-5 50 99 Wintergreen Oil O.1 10 Blend 103 Potassium Sorbate 590-OO-1 or O.O6 O.3 24634-61-5 11138-66-2 O.08 7732-18-5 50 99 Blend 95 6 35 Blend 104 Thyme Oil White 8007-46-3 O.1 10 sopropyl myristate 110-27-0 O.1 10 Lecithin, Soya. 8O3O-76-0 O.OO2 O.08 Potassium Sorbate 590-OO-1 or O.O6 O.3 24634-61-5 Polyglycerol-4-oleate 9007-48-1 O.O6 O.3 Xanthan Gum 11138-66-2 O.08 Water 7732-18-5 55 99 Wintergreen Oil O.1 Blend 105 Potassium Sorbate 590-OO-1 or O.O6 O.3 24634-61-5 11138-66-2 O.08 7732-18-5 50 99 6 35 Blend 106 Thyme Oil White 8007-46-3 O.1 10 Wintergreen Oil 68917-75-9 O.1 sopropyl myristate 110-27-0 O.1 10 Potassium Sorbate 590-OO-1 or O.O6 O.3 24634-61-5 Polyglycerol-4-oleate 9007-48-1 O.08 Xanthan Gum 11138-66-2 O.08 Water 7732-18-5 55 99 Blend 107 Potassium Sorbate 590-OO-1 or O. 24634-61-5 Xanthan Gum 11138-66-2 O. Water 7732-18-5 60 99 Blend 108 Sodium Benzoate S32-32-1 O. Water 7732-18-5 60 99 Blend 109 Span 80 1338-43-8 O. Tween 80 O. Isopar M 64742-47-8 8 40 Water 7732-18-5 35 99 Blend 8 O. 10 2% Sodium Benzoate 6 35 Blend 110 D-Limonene S989-27-5 O. Thyme Oil White 8007-46-3 O. Wintergreen Oil 68917-75-9 O. Span 80 1338-43-8 O. Tween 80 O. Sodium Benzoate S32-32-1 O.08 Isopar M 64742-47-8 8 40 Water 7732-18-5 40 99 Blend 111 Propellent A70 10 65 Blend 109 45 99 Blend 112 D-Limonene S989-27-5 O.1 Thyme Oil White 8007-46-3 O.08 Wintergreen Oil 68917-75-9 O.1 Span 80 1338-43-8 O.1 Tween 80 O.1 Sodium Benzoate S32-32-1 O.08 Isopar M 64742-47-8 6 35 Water 7732-18-5 35 99 Propellent A70 10 65 Blend 113 Sodium Lauryl Sulfate 151-21-3 5 30 Water 7732-18-5 55 99 Blend 114 Sodium Lauryl Sulfate 151-21-3 O.08 Xanthan Gum 11138-66-2 O.O6 O.3 Water 7732-18-5 60 99.9 US 8,691,256 B2 89 90 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high% Blend 115 Citroneia Oil 106-22-9 O.08 O.6 Carbopol 940 9003-01-4) O.08 O.6 BHT (butylated hydroxytoluene) 128-37-0 O.O6 O.3 Water 7732-18-5 30 99 Emulsifying Wax 67762-27-0, 8 40 90OS-67-8 Light Liquid Paraffin 8O12-95-1 O.1 10 White Soft Paraffin 8009-03-8 O.1 25 Sodium Metabisulphate 7681-57-4) O.08 1 Propylene Glycol 57-55-6 O.1 6 Cresmer RH40 hydrogenated castor 61791-12-6 O.1 15 oil Triethanolamine 102-71-6) O.08 O.6 Vitamin E. Acetate 58-95-7 O.OO2 O.08 Disodium EDTA 139-33-3 O.OOS O.2 Blend 1 O.1 15 Blend 116 Water 7732-18-5 2O 99 Blend 75 35 99 Blend 117 D-Limonene S989-27-5 O.1 10 Thyme Oil White 8007-46-3 O.1 15 Benzyl Alcohol 100-51-6 8 50 Isopar M 64742-47-8 10 65 Water 7732-18-5 25 99 Bifenthrin 83657-04-3 O.OOS O.2 Blend 63 O. 15 Stock 10% SLS Solution O. 10 Blend 118 Thyme Oil White 8007-46-3 O. 2 Wintergreen Oil 68917-75-9 O. 3 Isopropyl myristate 110-27-0 O. 3 Sodium Lauryl Sulfate 151-21-3 O.OO2 O.08 Water 7732-18-5 60 99 Blend 119 Thyme Oil White 8007-46-3 O. 4 Wintergreen Oil 68917-75-9 O. 8 Isopropyl myristate 110-27-0 O. 5 AgSorb clay carrier 60 99 Blend 120 Thyme Oil White 8007-46-3 O. 4 Wintergreen Oil 68917-75-9 O. 8 Isopropyl myristate 110-27-0 O. 5 DG Lite 60 99 Blend 121 D-Limonene S989-27-5 15 75 Thyme Oil White 8007-46-3 O. 4 Linaloo Coeur 78-70-6 O.08 O.6 Tetrahydrolinalool 78-69-3 O.08 O.6 Vanillin 121-33-5 O.OO2 O.08 Isopropyl myristate 110-27-0 O.08 O.6 Piperonal (aldehyde) Heliotropine 120-57-O O.08 O.6 Blend 66 O.1 10 Geraniol 60 106-24-1 O.O6 O.3 Triethyl Citrate 77-93-0 O.08 O.6 Water 7732-18-5 35 99 Stock 10% SLS Solution O.1 10 Blend 122 Miracle Gro (Sterile) 60 99 Blend 11 O.1 15 Blend 123 Thyme Oil White 8007-46-3 15 75 Amyl Butyrate 540-18-1 15 75 Anise Star Oil 30 99 Genistein O.OO1 O.1 Blend 124 Linaloo Coeur O.1 2O Tetrahydrolinalool O.1 25 Vanillin O.1 2 sopropyl myristate O.1 30 Piperonal (aldehyde) Heliotropine O.1 30 Geraniol Fine FCC O.1 15 Triethyl Citrate O.1 30 Thyme Oil White 30 99 Blend 125 D-Limonene S989-27-5 5 30 Linalool Coeur 78-70-6 8 40 Tetrahydrolinalool 78-69-3 15 75 Vanillin 121-33-5 O.1 8 sopropyl myristate 110-27-0 15 85 Piperonal (aldehyde) 120-57-O 5 30 Geraniol 60 5 30 Blend 126 D-Limonene S989-27-5 45 99 Thyme Oil White 8007-46-3 O.1 10 Linalool Coeur 78-70-6 O.1 2 US 8,691,256 B2 92 TABLE 5-continued

BLENDS CAS Registry Compounds Number low % high% Tetrahydrolinalool 78-69-3 O.1 3 Vanillin 121-33-5 O.OOS O.2 Isopropyl myristate 110-27-0 O.1 Piperonal (aldehyde) Heliotropine 120-57-O O.1 Blend 66 5 Geraniol 60 O.1 Triethyl Citrate 77-93-0 O.1

The foregoingTable 5 provides exemplary combinations of 15 range, and in the fifth column indicating a value at the high ingredients for useful blends in accordance with the inven end of Such exemplary range. The provided ranges are exem tion. In many cases a particular ingredient is listed very spe plary; other useful ranges exist and are expressly within the cifically such as, for example, with reference to a CAS num Scope of certain embodiments on the invention. Namely, ber and/or particular modifiers of the basic name of the ingredient. Such specific listings are non-limiting examples other high and low amounts defining other useful ranges of types of ingredients, and similar ingredients (such as, for and/or amounts of the listed ingredients, can include 1%, 2%, example, with different CAS numbers and/or variant forms of 5%, 10%, 15%, 20%, 25%, 40%, 50%,60%, 75%, 85%,95%, the type of ingredient) can be substituted within the scope of 110%, 125%, 150%, 175%, 200%, 250%, 300%, 400%, certain embodiments of the invention. 500%, 750%, 900%, or 1000% of the amount listed as the low The foregoingTable 5 also provides an examplary range of amount and/or the high amount, with the caveat that the amounts of each ingredient expressed as a weight/weight relative percentage of any given ingredient cannot exceed percentage of the listed blend. The exemplary range for each 99.99% of the total blend of ingredients. ingredient in each blend is provided as a number in the fourth Furthermore, other blends useful in accordance with the column indicating a value at the low end of Such exemplary present invention are shown in the following table. TABLE 6 Exemplified Ingredients Exemplified form % Range 1 % Range 2 % Range 3 % Range 4 % (w.fw) Example 1 - Ingredient Family 1

Linalool Linaloo Coeur O.66% 19.80% 3.30% 9.90% 4.95% 8.25% 5.94% 7.26% 6.60% Base Oil Soy Bean Oil 2.40% 72.00% 2.00%. 36.00% 18.00%. 30.00%. 21.60%. 26.40% 24.00% Thymol Thymol (crystal) 3.72% 99.00% 8.60% 55.80%. 27.90% 46.50%. 33.48%. 40.92% 37.20% Pinene Alpha-Pinene, 98% O.38% 11.40% 1.90% 5.70% 2.85% 4.75% 3.42% 4.18% 3.80% Cymene Para-Cymene 2.84% 85.17% 4.20%. 42.59%. 21.29%. 35.49%. 25.55%. 31.23% 28.39% Example 2 - Ingredient Family 2 Thyme Oil Thyme Oil White 2.06% 61.80% O.30%. 30.90% 15.45%. 25.75%. 18.54%. 22.66% 20.60% Wintergreen Oil Wintergreen Oil 4.51% 99.00%. 22.55%. 67.65%. 33.83% 56.38%. 40.59% 49.61% 45.10% Isopropyl myristate Isopropyl myristate 3.43% 99.00% 7.15% S1.45%. 25.73%. 42.88% 30.87%. 37.73% 34.30% Example 3 - Ingredient Family 3 Thyme Oil Thyme Oil White 2.48% 74.25% 2.38%. 37.13%. 18.56%. 30.94%. 22.28%. 27.23% 24.75% Amyl Butyrate Amyl Butyrate 2.30% 69.12% 1.52%. 34.56%. 17.28%. 28.80%. 20.74%. 25.34% 23.04% Anise Star Oil Anise Star Oil S.22% 99.00%. 26.11% 78.32%. 39.16% 65.26%. 46.99% S7.43% 52.21% Example 4 - Ingredient Family 4 Thyme Oil Thyme Oil White 2.48% 74.25% 2.38%. 37.13%. 18.56%. 30.94%. 22.28%. 27.23% 24.75% Amyl Butyrate Amyl Butyrate 2.30% 69.12% 1.52%. 34.56%. 17.28%. 28.80%. 20.74%. 25.34% 23.04% Anise Star Oil Anise Star Oil S.22% 99.00%. 26.10% 78.30%. 39.15%. 65.25% 46.98% S7.42% 52.20% Isoflavone Genistein O.OO190 S.00% O.005% O.02% O.OO8% 0.01.2% 0.009% 0.01.1% O.01% Example 5 - Ingredient Family 5 Thyme Oil Thyme Oil White 2.05% 61.50%, 10.25%. 30.75%. 15.38%. 25.63%. 18.45%. 22.SS90 20.50% Wintergreen Oil Wintergreen Oil 4.50% 99.00%. 22.50% 67.50%. 33.75% 56.25% 40.50%. 49.50% 45.00% Vanillin Vanillin O.11% S.00% 0.55% 1.65% 0.83%, 1.38% 0.99%. 1.21% 1.10% Isopropyl myristate Isopropyl myristate 3.34% 99.00% 16.70% SO.10% 25.05% 41.75%. 30.06%. 36.74% 33.40% Example 6 - Ingredient Family 6

Limonene D-Limonene S.63% 99.00%. 28.15% 84.45%. 42.23% 70.38% SO.67%. 61.93% 56.30% Thyme Oil Thyme Oil White 1.24% 37.14% 6.19%. 18.57% 9.29%. 15.48%. 11.14%. 13.62% 12.38% Wintergreen Oil Wintergreen Oil 3.13% 93.96%. 15.66%. 46.98%. 23.49%. 39.15%. 28.19%. 34.45% 31.32% Example 7 - Ingredient Family 7

Potassium Sorbate Potassium Sorbate O.10% S.00% O.S.0% 1.50% 0.75%. 1.25% O.90% 1.10% 1.00% Xanthan Gum Xanthan Gum O.O.3% S.00% O.14% O.42% 0.21% 0.35% 0.25% O.31% O.28% Water Water 8.18% 99.00%, 40.91% 99.00% 61.37% 99.00% 73.64% 90.00% 81.82% Blend 74 Blend 74 1.69% SO.7% 8.45%. 25.35%. 12.68%. 21.13%. 15.21%. 18.59% 16.90% US 8,691,256 B2 93 94 TABLE 6-continued Exemplified Ingredients Exemplified form % Range 1 % Range 2 % Range 3 % Range 4 % (w.fw) Example 8 - Ingredient Family 8

Isopropyl myristate Isopropyl myristate 4.84% 99.00% 24.18% 72.53% 36.26% 60.44% 43.52% S3.19% 48.35% Geraniol Geranio Fine FCC 1.50% 44.94% 7.49%. 22.47% 11.24% 18.73% 13.48% 16.48% 14.98% Thyme Oil Thyme Oil White 3.67% 99.00% 18.34% SS.01% 27.50% 45.84% 33.00% 40.34% 36.67% Example 9 - Ingredient Family 9

Limonene D-Limonene O.99% 29.70% 4.95%. 14.85% 7.43% 12.38% 8.91% 10.89% 9.90% Linalool Linalool Coeur 1.41% 42.42% 7.07% 21.21% 10.61% 17.68% 12.73% 15.55% 14.14% Tetrahydrolinalool Tetrahydrolinalool 2.43% 72.87% 12.15%. 36.44% 18.22% 30.36% 21.86% 26.72% 24.29% Vanillin Vanillin O.25% 7.44% 1.24% 3.72% 1.86% 3.10% 2.23% 2.73% 2.48% Sopropyl myristate Sopropyl myristate 2.89% 86.76% 14.46%. 43.38% 21.69% 36.15% 26.03% 31.81% 28.92% Piperonal Piperonal (aldehyde) 1.00% 29.91% 4.99%. 14.96% 7.48% 12.46% 8.97% 10.97% 9.97% Geraniol Geranio Fine FCC 1.03% 30.90% S.15%. 15.45% 7.73% 12.88% 9.27% 11.33% 10.30% Example 10- Ingredient Family 10

Limonene D-Limonene 2.85% 85.38% 14.23%. 42.69% 21.35% 35.58% 25.61% 31.31% 28.46% Thyme Oil Thyme Oil White 3.13% 93.87% 15.65% 46.94% 23.47% 39.11% 28.16% 34.42% 31.29% Blend 63 Blend 63 4.03% 99.00% 20.13% 60.38% 30.19% S.O.31% 36.23% 44.28% 40.25% Example 11 - Ingredient Family 11

Limonene D-Limonene O.96% 28.89% 4.82%. 14.45% 7.22% 12.04% 8.67% 10.59% 9.63% BSD BSD 2.67% 79.98% 13.33%. 39.99% 20.00% 33.33% 23.99% 29.33% 26.66% Linalool Linalool Coeur O.98% 29.46% 4.91%. 14.73% 7.37% 12.28% 8.84% 10.80% 9.82% Tetrahydrolinalool Tetrahydrolinalool 1.18% 35.43% 5.91%. 17.72% 8.86% 14.76% 10.63% 12.99% 11.81% Vanillin Vanillin O.12% S.00% O.60% 1.80% O.90% 1.50% 1.08% 1.32% 1.20% Base oil Mineral Oil White USP 1.50% 44.91% 7.49%. 22.46% 11.23% 18.71% 13.47% 16.47% 14.97% Sopropyl myristate Sopropyl myristate 1.45% 43.62% 7.27% 21.81% 10.91% 18.18% 13.09% 15.99% 14.54% Piperonal Piperonal (aldehyde) O.49% 14.55% 2.43% 7.28% 3.64% 6.06% 4.37% S.34% 4.85% Geraniol Geranio Fine FCC O.65% 19.53% 3.26% 9.77% 4.88% 8.14% 5.86% 7.16% 6.51% Example 12 - Ingredient Family 12

Thyme Oil Thyme Oil White 4.19% 99.00% 20.93% 62.79% 31.40% 52.33% 37.67% 46.05% 4.1.86% Isopropyl myristate Isopropyl myristate 3.83% 99.00% 19.17% S7.51% 28.76% 47.93% 34.51% 42.17% 38.34% Geraniol Geranio Fine FCC 1.98% 59.40% 9.90%. 29.70% 14.85% 24.75% 17.82% 21.78% 19.80% Example 13 - Ingredient Family 13

Linalool Linaloo Coeur 2.34% 70.14% 11.69%. 35.07% 17.54% 29.23% 21.04% 25.72% 23.38% Amyl Butyrate Amyl Butyrate 2.35% 70.38% 11.73%. 35.19% 17.60% 29.33% 21.11% 25.81% 23.46% Anise Star Oil Anise Star Oil 5.32% 99.00% 26.58% 79.74% 39.87% 66.45% 47.84% 58.48% 53.16% Example 14 - Ingredient Family 14

Linalool Linaloo Coeur 3.74% 99.00% 18.72%. 56.16% 28.08% 46.80% 33.70% 41.18% 37.44% Thymol Thymol 3.67% 99.00% 18.36% SS.08% 27.54% 45.90% 33.05% 40.39% 36.72% Pinene Alpha-pinene, 98% O.47% 13.98% 2.33% 6.99% 3.50% 5.83% 4.19% S.13% 4.66% Cymene Para-Cymene O.19% S.61% O.94% 2.81% 1.40% 2.34% 1.68% 2.06% 1.87% Anethole Trans-Anethole 1.93% 57.93% 9.66%. 28.97% 14.48% 24.14% 17.38% 21.24% 19.31% Example 15 - Ingredient Family 15

Limonene D-Limonene 2.74% 82.05% 13.68%. 41.03% 20.51% 34.19% 24.62% 30.09% 27.35% Thyme Oi Thyme Oil White 3.01% 90.24% 15.04% 45.12% 22.56% 37.60% 27.07% 33.09% 30.08% Lilac Flower Oi Lilac Flower Oil 4.26% 99.00% 21.30% 63.90% 31.95% 53.25% 38.34% 46.86% 42.57% Example 16 - Ingredient Family 16

Thyme Oi Thyme Oil White 3.82% 99.00% 19.11% S7.32% 28.66% 47.76% 34.39% 42.03% 38.21% Wintergreen Oi Wintergreen Oi 2.48% 74.37% 12.40%. 37.19% 18.59% 30.99% 22.31% 27.27% 24.79% Isopropyl Myristate Isopropyl Myristate 3.59% 99.00% 17.95%. 53.84% 26.92% 44.86% 32.30% 39.48% 35.89% vanillin Vanillin O.11% S.00% O.S.6% 1.67% O.83% 1.39% 1.00% 1.22% 1.11% Example 17 - Ingredient Family 17

Wintergreen Oi Wintergreen Oi 2.48% 74.46% 12.41%. 37.23% 18.62% 31.03% 22.34% 27.30% 24.82% Isopropyl Myristate Isopropyl Myristate 3.59% 99.00% 17.97% S3.91% 26.96% 44.93% 32.35% 39.53% 35.94% Thyme Oi Thyme Oil White 3.92% 99.00% 19.62%. 58.86% 29.43% 49.05% 35.32% 43.16% 39.24% Example 18 - Ingredient Family 18

Thyme Oi Thyme Oil White O.46% 13.8% 2.30% 6.90% 3.45% 5.75% 4.14% S.06% 4.60% Wintergreen Oi Wintergreen Oi 5.78% 99.00% 28.90% 86.70% 43.35% 72.25% S2.02% 63.58% 57.80% Isopropyl Myristate Isopropyl Myristate 3.76% 99.00% 18.80% 56.40% 28.20% 47.00% 33.84% 41.36% 37.60% Example 19 - Ingredient Family 19

Thyme Oi Thyme Oil White 3.16% 94.71% 15.79% 47.36% 23.68% 39.46% 28.41% 34.73% 31.57% Isopropyl myristate Isopropyl myristate 3.86% 99.00% 19.28% S7.84% 28.92% 48.20% 34.70% 42.42% 38.56% Wintergreen Oi Wintergreen Oi 2.99% 89.61% 14.94%. 44.81% 22.40% 37.34% 26.88% 32.86% 29.87% US 8,691,256 B2 95 96 TABLE 6-continued Exemplified Ingredients Exemplified form % Range 1 % Range 2 % Range 3 % Range 4 % (w.fw) Example 20 - Ingredient Family 20 Thyme Oil Thyme Oil White 2.06% 61.80%. 10.30%. 30.90% 15.45%. 25.75%. 18.54%. 22.66%. 20.60% Isopropyl myristate Isopropyl myristate 3.43% 99.00% 17.15% S1.45%. 25.73%. 42.88% 30.87%. 37.73% 34.30% Geraniol Geranio Fine FCC 4.51% 99.00%. 22.SS90 67.65%. 33.83%. 56.38%. 40.59% 49.61%. 45.10% Example 21 - Ingredient Family 21 Thyme Oil Thyme Oil White 1.24%. 37.14% 6.19%. 18.57% 9.29%. 15.48%. 11.14%. 13.62%. 12.38% Wintergreen Oil Wintergreen Oil 3.13% 93.96%. 15.66%. 46.98%. 23.49%. 39.15%. 28.19%. 34.45%. 31.32% Limonene D-Limonene S.63% 99.00%. 28.15% 84.45%. 42.23% 70.38% SO.67%. 61.93% 56.30% Example 22 - Ingredient Family 22

LFO LFO S.O1% 99.00%. 25.07% 75.20%. 37.60% 62.66%. 45.12% SS.14% SO.13% BSO (Black Seed Oil) BSO 4.99% 99.00% 24.94% 74.81% 37.40% 62.34%. 44.88% 54.86%. 49.87% Example 23 - Ingredient Family 23

LFO LFO 8.01% 99.00%. 40.05% 99.00% 60.07% 99.00% 72.08% 88.10% 80.09% BSO (Black Seed Oil) BSO 1.99% S9.73% 9.96%. 29.87%. 14.93%. 24.89%. 17.92%. 21.90%. 19.91%

Example 1 TABLE 7-continued Pesticidal Effect on Culex quinquefasciatus - Mosquitoes - 25 # Added to # Dead after % The effect of compositions, and their individual ingredi- Chamber 4 hours Mortality ents, on the mortality of insects is tested. Multiple plexiglass Composition 50 50 100% chambers are used. A treatment chamber is provided for each (Pyrethrum and Blend 4) composition and ingredient that is tested, and the chambers are sprayed (aerosol spray) evenly on all Surfaces with the 30 composition or ingredient being tested. A control chamber is Example 2 provided that is not treated. Southern house mosquitoes, Culex quinquefasciatus, are Synergistic Compositions as Indicated by TyR obtained as test organisms. Multiple laboratory-cultured, is Binding Inhibition Sucrose-fed female mosquitoes aged about 2-5 days are released into the glass chambers prior to the spraying of When the chemical(s) and compound(s) are combined tO aerosol. The discharge rate (gm/second) of each can of aero- provide the EEE the pesent invention, ther th sol to be tested is predetermined. Based on the dosage synergistic effect. The efficacy for insect control and the - 0 synergistic effect of compositions can be predicted and dem required,he gl an estimatedhamb time of spray of aerosol is discharged 40 onstrated in a variety of manners, for example, a competition 1nto the glass chamber. - binding assay can be used. With reference to Table 8, the Knockdown of mosquitoes is observed at indicated 1nter- percent TyrR binding inhibition affected by the following vals up to about 20 minutes. After about 20 minutes, all agents was determined using a competition binding assay: the mosquitoes are collected and placed in cylindrical polyethyl- natural ligand, Tyramine (TA); Blend 7 (exemplified in Ingre ene containers with 10% sucrose pads. Mortality is observed 45 dient Family 5 of Table 6); Blend 12 (exemplified in Ingredi 4 hours post-treatment. The mortality value is based on a ent Family 9 of Table 6); DM; Pyrethrum:90:1 Blend 7+DM: combination of dead and moribund mosquitoes over the total 9:1 Blend 7+Pyrethrum: 90:1 Blend 12+DM; and 9:1 Blend number of mosquitoes initially released. 12+Pyrethrum. The data from an exemplary study is shown in Table 7. The study tested: (1) a composition comprising Pyrethrum and 50 TABLE 8 Blend 4; (2) Pyrethrum; (3) BSO; and (4) LFO (IFF Inc., Hazlet, N.J.). The percent mortality of the mosquitoes treated Agent % TyrR Binding Inhibition with the composition was 100%, compared to 60% for BSO Tyramine (TA) 75 alone, 80% for LFO alone, 90% for Pyrethrum alone, and 0% Blend 7 30 for the non-treated control. Blend 4 is exemplified in Ingre- 55 Red 12 dient Family 23 in Table 6. Pyret 5 90:1 Blend 7+ DM 50 TABLE 7 9:1 Blend 7 + Pyrethrum 60 90:1 Blend 12 + DM 60 Mosquitoes 60 9:1 Blend 12 + Pyrethrum 60

# Added to # Dead after % Chamber 4 hours Mortality One example of an Synergistic effect shown by this study is 0. as follows: the insect control chemical, Pyrethrum, only has a Sye so o 5%TyrRbinding inhibition, and Blend 7 only has a 30%TyrR LFO 50 40 80% 65 binding inhibition; however, when Pyrethrum and Blend 7 are Pyrethrum 50 45 90% combined, the TyrR binding inhibition increases to 60%, approaching that of the natural ligand. US 8,691,256 B2 97 98 Example 3 with 2.5% Blend 19 resulted in 90% KD at 30 seconds post treatment. The composition including Blend 19 and CL was Pesticidal Effect Against Blattella germanica shown to be effective and was shown to have a synergistic effect. With reference to Table 9, the pesticidal effect against 5 Similarly, with reference to FIG. 4D, the pesticidal effect Blattella germanica (German cockroaches) was determined against Drosophila sp. was determined for Blend 19 (labeled for DM, Blend 12 (exemplified in Ingredient Family 9 of “HL1) and the composition including Imidacloprid and Table 6), and the composition including deltamethrin (DM) Blend 19. Treatment with Imidacloprid alone at 50 ppm and Blend 12. Treatment with DMalone resulted in an aver resulted in 0% KD of the target insect at 30 seconds post age knock down (KD) of the insects in 120 sec, and 100% 10 treatment, while treatment with 2.5% Blend 19 alone also killing of the insects in 15 minutes. Treatment with Blend 12 resulted in 0% KD of the target insect at 30 seconds post alone resulted in an average KD of the insects in 20 sec, and treatment. However treatment with Imidacloprid at 50 ppm 100% killing of the insects in 5 minutes. A synergistic effect combined with 2.5% Blend 19 resulted in 70% KD at 30 was shown for the combination treatment that resulted in an seconds post-treatment. The composition including Blend 19 average KD of the insects in 5 sec, and 100% killing of the 15 and CL was shown to be effective and was shown to have a insects in 55 seconds. The composition including Blend 12 synergistic effect. and DM was shown to be effective and was shown to have a synergistic effect. Additionally, the above-described meth Example 5 ods, including competition receptor binding assays, assess ments of changes in cAMP, and assessments of changes in Pesticidal Effect Against Aedes aegypti Ca", are confirmed to be effective at predicting and demon strating the synergistic effect of and the efficacy of the com With reference to FIG. 5, the pesticidal effect against Aedes position. aegypti was determined for Blend 11 (labeled “B5028, exemplified in Ingredient Family 2 in Table 6) and the com TABLE 9 25 position including Imidacloprid (labeled “I”) and B5028. Treatment with Imidacloprid alone at 500 ppm resulted in no Efficacy of DM and Blend 12 against German cockroaches KD of the target insect, and treatment with B5028 at 5% Bioactivity showed 10% KD of the target. However treatment with Imi dacloprid at 500 ppm combined with B5028 at 5% resulted in Chemicals KD 100% Kill 30 100% KD. The composition including B5028 and imidaclo DM (0.037 mg/cm) 120 sec 15 min prid was shown to be effective and was shown to have a (17 ptl of 16.99% formulated DM) synergistic effect. Blend 12 20 sec 5 min (1.9 mg/cm) Example 6 Composition (1.9 mg/cm) 5 sec SS Sec 35 (1 part DM:9 parts Blend 12 (v/v)) Comparison of Pesticidal Effects Similarly, with reference to Table 10, the pesticidal effect Example 4 against German cockroaches was determined for delta 40 methrin (DM), Blend 7 (exemplified in Ingredient Family 5 of Pesticidal Effect Against Aedes aegypti Table 6), and the composition including DM and Blend 7. Treatment with DM alone resulted in an average KD of the With reference to FIG. 4A, the pesticidal effect against insects in 140 sec, and 100% killing of the insects in 12 Aedes aegypti was determined for Blend 19 (labeled “HL1, minutes. Treatment with Blend 7 alone resulted in an average exemplified in Ingredient Family 15 in Table 6) and the com 45 KD of the insects in 10 sec, and 100% killing of the insects in position including clothianidin (CL) and Blend 19. Treatment 45 seconds. A synergistic effect was shown for the combina with CL alone at 500 ppm resulted in no KD of the target tion treatment that results in an average KD of the insects in 5 insect, however treatment with CL at 167 ppm combined with sec, and 100% killing of the insects in 17 seconds. The com 2.5% Blend 19 resulted in 100% KD. The composition position including Blend 7 and DM was shown to be effective including Blend 19 and CL was shown to be effective and was 50 and was shown to have a synergistic effect. The above-de shown to have a synergistic effect. scribed methods, including competition receptor binding Similarly, with reference to FIG. 4B, the pesticidal effect assays, assessments of changes in cAMP, and assessments of against Aedes aegypti was determined for Blend 19 (labeled changes in Ca", were confirmed to be effective at predicting “HL1) and the composition including CL and Blend 19. and demonstrating the Synergistic effect of and the efficacy of Treatment with CL alone at 250 ppm resulted in no KD of the 55 the composition. target insect, however treatment with CL at 167 ppm com bined with 2.5% Blend 19 resulted in 100% KD. The com TABLE 10 position including Blend 19 and CL was shown to be effective and was shown to have a synergistic effect. Efficacy of DM and Blend 7 against German cockroaches Similarly, with reference to FIG. 4C, the pesticidal effect 60 against Aedes aegypti was determined for Blend 19 (labeled Bioactivity “HL1) and the composition including Imidacloprid and Chemicals KD 100% Kill Blend 19. Treatment with Imidacloprid alone at 250 ppm DM (0.037 mg/cm) 140 sec 12 min resulted in 20% KD of the target insect at 30 seconds post (17 pil of 16.99% formulated DM) treatment, while treatment with 2.5% Blend 19 alone resulted 65 Blend 7 10 sec 45 sec in 40% KD of the target insect at 30 seconds post-treatment. (3.8 mg/cm) However treatment with Imidacloprid at 250 ppm combined US 8,691,256 B2 99 100 TABLE 10-continued Blend 19 and Imidacloprid was shown to be effective and was shown to have a synergistic effect. Efficacy of DM and Blend 7 against German cockroaches Bioactivity Example 9 5 Chemicals KD 100% Kill Pesticidal Effect Against Bed Bugs Composition (3.8 mg/cm) 5 sec 17 sec (1 part DM:99 parts Blend 7 (v/v)) Turning now to FIG.7 showing the pesticidal effect against bed bugs expressed as percent mortality as a function of time, 10 the 1:1 ratio composition was shown to have a synergistic effect, when compared to the pesticidal effect of Blend 12 Example 7 (labeled as “CL-4, exemplified in Ingredient Family 9 in Table 6) or Pyrethrum alone. The pyrethrum alone did not Comparison of Pesticidal Effects achieve higher than about 30% mortality, and Blend 12 alone 15 did not achieve higher than about 80% mortality. However, With reference to Table 11, the pesticidal effect against the 1:1 ratio composition including Blend 12 and Pyrethrum Darkling Beetles was determined for Pyrethrum, Blend 12 resulted in 100% mortality, as early as about 30 minutes after (exemplified in Ingredient Family 9 of Table 6), and the treatment, and had a residual effect lasting up to about 24 composition including Pyrethrum and Blend 12. hours after treatment.

TABLE 11 Example 10 Efficacy of Pyrethrum and Blend 12 against Darkling Beetles Pesticidal Effects of a Combination of Blend 11 with % Mortality after Application Imidacloprid Against Thrips by direct Spray to Darkling Beetle 25 Thrips were exposed to three different groups of composi Test Material Day 1 Day 4 Day 8 Day 12 tions. Both knockdown (KD) and mortality were measured at Vehicle Control O O% O O% S 7% S 7% indicated time intervals. Knockdown of target pests is mea (Water) Sured in a plexilglass chamber at indicated intervals. Mortal 4% Blend 12 155% 40 - 13%. 55 10% 800% 30 ity of target pests is measured in cylindrical polyethylene 4% Pyrethrum O O% 10 10% 20 19%. 30.28% 2% Blend 12 25 13% 45 17% 80 - 14%. 100 O%* containers with 10% sucrose pads. The mortality value is and 2% based on a combination of dead and moribund pests over the Pyrethrum total number of pests initially released. The KD and mortality Values displayed are the mean plus or minus the standard deviation for 4 replicates of 10 measurements were made at 5 minutes, 30 minutes, 1 hour, 2 insects each, except vehicle control-(2 replicates of 10 insects each), 35 hours, 4 hours and 24 hours post treatment. **Significantly greater than all other values for mortality (P<0.001, 2 tail student t Test) With reference to FIG. 9, the first group of compositions The synergistic effect can be altered by changing the spe contained imidacloprid at 1 ppm, 10 ppm, 20 ppm, 50 ppm cific combinations of ingredients or changing the specific and 100 ppm. Treatment with imidacloprid alone at 1 ppm or ratios of ingredients. 10 ppm resulted in no KD of the thrips till the end of the 40 experiment at 24 hours post treatment. Treatment with imi dacloprid alone at 20 ppm resulted in 12% KD of the thrips at Example 8 24 hours post treatment. Treatment with imidacloprid alone at 50 ppm resulted in 100% KD of the thrips at 24 hours post Pesticidal Effect Against Periplaneta americana treatment. Treatment with imidacloprid alone at 100 ppm 45 resulted in 10%, 20% and 80% KD of the thrips at 2 hours, 4 With reference to FIG. 6A, the pesticidal effect against hours and 24 hours post treatment, respectively. Moreover, Periplaneta americana was determined for Blend 19 (labeled treatment with imidacloprid alone at 1 ppm, 10 ppm or 20 “HL1, exemplified in Ingredient Family 15 in Table 6) and ppm resulted in no mortality of the thrips till the end of the the composition including clothianidin (CL) and Blend 19. experiment at 24 hours post treatment. Treatment with imi Treatment with CL alone at 0.05% resulted in no mortality of 50 dacloprid alone at 50 ppm resulted in 50% mortality of the the targetinsect at 30 minutes post-treatment, while treatment thrips at 24 hours post treatment. Treatment with imidaclo with Blend 19 at 5% resulted in 60% target mortality 30 prid alone at 100 ppm resulted in 10%, 20% and 40% mor minutes post-treatment. However treatment with CL at 0.05% tality of the thrips at 2 hours, 4 hours and 24 hours post combined with 5% Blend 19 resulted in 100% mortality 30 treatment, respectively. minutes post-treatment. The composition including Blend 19 55 The composition of Blend 11 is exemplified in Ingredient and CL was shown to be effective and was shown to have a Family 17 in Table 6. synergistic effect. With reference to FIG. 10A, the second group of compo With reference to FIG. 6B, the pesticidal effect against sitions contained Blend 11 at 0.01%, 0.1%, 0.2%, 0.5% and Periplaneta americana was determined for Blend 19 (labeled 1% by volume. Treatment with Blend 11 alone at 0.1% by “HL1) and the composition including Imidacloprid and 60 volume resulted in no KD of the thrips till the end of the Blend 19. Treatment with Imidacloprid alone (at 0.05%, experiment at 24 hours post treatment. Treatment with Blend 0.033%, and 0.01%) resulted in no mortality of the target 11 alone at 0.01%, 0.2%, 0.5% and 1% by volume resulted in insect at 30 minutes post-treatment, while treatment with 12%, 20%, 36% and 50% KD of the thrips at 24 hours post Blend 19 at 5% resulted in 60% target mortality 30 minutes treatment, respectively. Moreover, as shown in FIG. 10B, post-treatment. However treatment with Imidacloprid at 65 treatment with Blend 11 alone at 0.1% by volume resulted in 0.033% combined with 5% Blend 19 resulted in 90% mortal no mortality of the thrips till the end of the experiment at 24 ity 30 minutes post-treatment. The composition including hours post treatment. Treatment with Blend 11 alone at US 8,691,256 B2 101 102 0.01%, 0.2%, 0.5% and 1% by volume resulted in 12%, 20%, As shown in FIG. 11C, treatment with the composition 36% and 37% mortality of the thrips at 24 hours post treat with the mixture of Blend 8 at 0.01% by volume and imida ment, respectively. cloprid resulted in no KD of the thrips till the end of the With reference to FIGS. 11A and B, the third group of experiment at 24 hours post treatment. Treatment with the compositions contained a mixture of Blend 11 and imidaclo 5 composition with the mixture of Blend 8 at 0.1% by volume prid at 100:1 by volume. The concentration of Blend 11 was and imidacloprid resulted in 17%, 17% and 70% KD of the 0.01%, 0.1%, 0.2%, 0.5% and 1% by volume, respectively. thrips at 2 hours, 4 hours and 24 hours post treatment, respec As shown in FIG. 11A, treatment with the composition tively. Treatment with the composition with the mixture of with the mixture of Blend 11 at 0.01% and 0.1% by volume Blend 8 at 0.2% by volume and imidacloprid resulted in 8% and imidacloprid resulted in 40% and 22% KD of the thrips at 10 and 43% KD of the thrips at 4 hours and 24 hours post 24 hours post-treatment, respectively. Treatment with the treatment, respectively. Treatment with the composition with the mixture of Blend 8 at 0.5% by volume and imidacloprid composition with the mixture of Blend 11 at 0.2% by volume resulted in 60% KD of the thrips at 1 hour post treatment, and and imidacloprid resulted in 20%, 20%, 50%, 60% and 70% 100% KD between 2 hours and 24 hours post treatment, KD of the thrips at 30 minutes, 1 hours, 2 hours, 4 hours and 15 respectively. Treatment with the composition with the mix 24 hours post treatment, respectively. Treatment with the ture of Blend 8 at 1% by volume and imidacloprid resulted in composition with the mixture of Blend 11 at 0.5% by volume 100% KD between 5 minutes and 4 hours post treatment, and imidacloprid resulted in 18% and 50% KD of the thrips at respectively, and 90% KD at 24 hours post treatment. More 5 minutes and 30 minutes post treatment, respectively, and over, as shown in FIG. 11D treatment with the composition 100% KD between 1 hour and 24 hours post treatment, with the mixture of Blend 8 at 0.01% by volume and imida respectively. Treatment with the composition with the mix cloprid resulted in no mortality of the thrips till the end of the ture of Blend 11 at 1% by volume and imidacloprid resulted experiment at 24 hours post treatment. Treatment with the in 100% KD between 5 minutes and 24 hours post treatment. composition with the mixture of Blend 8 at 0.1% by volume Moreover, as shown in FIG. 11B, treatment with the compo and imidacloprid resulted in 8% and 40% mortality of the sition with the mixture of Blend 11 at 0.01% and 0.1% by 25 thrips at 4 hours and 24 hours post treatment, respectively. volume and imidacloprid resulted in 20% and 10% mortality Treatment with the composition with the mixture of Blend 8 of the thrips at 24 hours post-treatment, respectively. Treat at 0.2% by volume and imidacloprid resulted in 28% mortal ment with the composition with the mixture of Blend 11 at ity of the thrips at 24 hours post treatment. Treatment with the 0.2% by volume and imidacloprid resulted in 20% and 30% composition with the mixture of Blend 8 at 0.5% by volume mortality of the thrips at 4 hours and 24 hours post-treatment, 30 and imidacloprid resulted in 70%, 70% and 100% mortality respectively. Treatment with the composition with the mix of the thrips at 2 hours, 4 hours and 24 hours post treatment, ture of Blend 11 at 0.5% by volume and imidacloprid resulted respectively. Treatment with the composition with the mix in 100% mortality of the thrips between 1 hour and 24 hours ture of Blend 8 at 1% by volume and imidacloprid resulted in post treatment. Treatment with the composition with the mix 44%. 44%, 40% and 68% mortality of the thrips at 1 hour, 2 ture of Blend 11 at 1% by volume and imidacloprid resulted 35 hours, 4 hours and 24 hours post treatment, respectively. in 80% mortality of the thrips between 1 hours and 4 hours These results demonstrate that the combination of imida post treatment, and 90% mortality at 24 hours post treatment. cloprid and Blend 8 is effective and has a synergistic effect. These results demonstrate that the combination of imida cloprid and Blend 11 is effective and has a synergistic effect. Example 12 40 Example 11 Pesticidal Effects of a Combination of Blend 38 with Imidacloprid Against Thrips Pesticidal Effects of a Combination of Blend 8 with Imidacloprid Against Thrips Thrips were exposed to three different groups of composi 45 tions. Both knockdown (KD) and mortality were measured as Thrips were exposed to three different groups of composi described in Example 10 at 1 hour, 2 hours, 4 hours and 24 tions. Both knockdown (KD) and mortality were measured as hours post treatment. Blend 38 (label “B5049” and “TT1A) described in Example 10 at 5 minutes, 30 minutes, 1 hour, 2 is exemplified in Ingredient Family 12 in Table 6. hours, 4 hours and 24 hours post treatment. The composition With reference to FIG. 12, the first group of compositions of Blend 8 is exemplified in Ingredient Family 21 in Table 6. 50 contained imidacloprid at 1 ppm, 10 ppm, 20 ppm, and 50 With reference to FIG. 9, the first group of compositions ppm. Treatment with imidacloprid alone at 1 ppm or 10 ppm contained imidacloprid at 1 ppm, 10 ppm, 20 ppm, 50 ppm resulted in no KD of the thrips till the end of the experiment and 100 ppm, as discussed in Example 10. at 24 hours post treatment. Treatment with imidacloprid alone With reference to FIG. 10C, the second group of compo at 20 ppm resulted in 12% KD of the thrips at 24 hours post sitions contained Blend 8 at 0.01%, 0.1%, 0.2%, 0.5% and 1% 55 treatment. Treatment with imidacloprid alone at 50 ppm by volume. Treatment with Blend 8 alone at 0.2% by volume resulted in 100% KD of the thrips at 24 hours post treatment. resulted in 20% KD of the thrips at 24 hours post-treatment. Moreover, treatment with imidacloprid alone at 1 ppm, 10 Treatment with Blend 8 alone at 0.01%, 0.1%, 0.5% and 1% ppm or 20 ppm resulted in no mortality of the thrips till the by volume resulted in no KD of the thrips till the end of the end of the experiment at 24 hours post treatment. Treatment experiment at 24 hours post treatment, respectively. More 60 with imidacloprid alone at 50 ppm resulted in 50% mortality over, as shown in FIG.10D treatment with Blend 8 alone at all of the thrips at 24 hours post treatment. concentrations tested resulted in no mortality of the thrips till With reference to FIG. 13, the second group of composi the end of the experiment at 24 hours post treatment. tions contained Blend 38 (labeled “B5049) at 0.01%, 0.10%, With reference to FIGS. 11C and D, the third group of 0.20%, and 0.50% by volume. Treatment with Blend 38 alone compositions contained a mixture of Blend 8 and imidaclo 65 at 0.10% by volume resulted in no KD of the thrips till the end prid at 100:1 by volume. The concentration of Blend 8 was of the experiment at 24 hours post treatment. Treatment with 0.01%, 0.1%, 0.2%, 0.5% and 1% by volume, respectively. Blend 38 alone at 0.01%, 0.20% and 0.50% by volume US 8,691,256 B2 103 104 resulted in 12%, 20% and 37% KD of the thrips at 24 hours 39 (labeled “B5053” and “TT3C', exemplified in Ingredient post treatment, respectively. Moreover, treatment with Blend Family 8 in Table 6) at 1% by volume resulted in 86% 38 alone at 0.10% by volume resulted in no mortality of the decrease in aphid number on the plants (86 Percent Control as thripstill the end of the experiment at 24 hours post treatment. shown in FIG. 15). Treatment with Blend 38 alone at 0.01%, 0.20% and 0.50% by volume resulted in 12%, 20% and 37% mortality of the These results demonstrate that imidacloprid and Blends 38 thrips at 24 hours post treatment, respectively. and 39 are effective and have synergistic effects. With reference to FIG. 14, the third group of compositions contained a mixture of Blend 38 (label “B5049” and “TT Example 14 1A) and imidacloprid at 100:1 by volume. The concentration 10 of Blend 38 was 0.01%, 0.1%, 0.2%, and 0.5% by volume, respectively. The KD and mortality measurements were done Pesticidal Effect Against C. Elegans at 1 hour, 4 hours and 24 hours post treatment. As shown in FIG. 14, treatment with the composition with C. elegans were exposed to two different groups of com the mixture of Blend 38 (label “B5049” and “TT 1A) at 15 positions before the mortality of C. elegans was measured. 0.01% by volume and imidacloprid resulted in 40%. KD of the thrips at 24 hours post treatment. Treatment with the compo With reference to Table 12, the first group of compositions sition with the mixture of Blend 38 at 0.1% by volume and contained Blend 27 alone at LC50 and applied to C. elegans. imidacloprid resulted in 22% KD of the thrips at 24 hours post The treatment killed 50% C. elegans. The second group of treatment. Treatment with the composition with the mixture composition contained Blend 27 (exemplified in Ingredient of Blend 38 at 0.2% by volume and imidacloprid resulted in Family 1 in Table 6) at LC50 combined with fipronil at 5 ppm 20%, 60% and 70% KD of the thrips at 1 hour, 4 hours and 24 and applied to C. elegans. The treatment killed 93%. C. hours post treatment, respectively. Treatment with the com elegans. The result demonstrates that fipronil and Blend 27 position with the mixture of Blend 38 at 0.5% by volume and are effective and have synergistic effects. imidacloprid resulted in 100% KD of the thrips between 1 25 Blends listed in Table 12, alone and in combination with hour and 24 hours post treatment, respectively. Moreover, fipronil, were tested in the same fashion. treatment with the composition with the mixture of Blend 38 at 0.01% by volume and imidacloprid resulted in 20% mor TABLE 12 tality of the thrips at 24 hours post treatment. Treatment with the composition with the mixture of Blend 38 at 0.1% by 30 Pesticidal Effects of Blends with and without Fipronil volume and imidacloprid resulted in 11% mortality of the thrips at 24 hours post treatment. Treatment with the compo Fipronil sition with the mixture of Blend 38 at 0.2% by volume and Concen- Kill 90 Celegans imidacloprid resulted in 20% and 30% mortality of the thrips Exemplified tration by wo at 4 hours and 24 hours post treatment, respectively. Treat 35 in Table 6 LCSO Volume Fipronil w Fipronil ment with the composition with the mixture of Blend 38 at 0.5% by volume and imidacloprid resulted in 100% mortality Blend 27 Ingredient 150 ppm 5 ppm SO% 93% of the thrips between 1 hour and 24 hours post treatment, Family 1 respectively. Blend 39 Ingredient 424 ppm 5 ppm 43% 779, These results demonstrate that the combination of imida 40 Family 8 Blend 42 Ingredient 269 ppm 5 ppm 48% 100% cloprid and Blend 38 is effective and has a synergistic effect. Family 3 Blend 12 Ingredient 519 ppm 5 ppm SO% 85% Example 13 Family 9

Pesitcideal Effect Against Adult Green Peach Aphids 45 The results demonstrate that the combinations of fipronil Adult green peach aphids were observed and counted on and these blends are effective and have synergistic effects. the plants in a greenhouse. Then different compositions were sprayed directly onto the plants with aphids. Three days after the composition was applied, aphids were counted again to 50 Example 15 evaluate the pesticidal effects of the composition. With reference to FIG. 15, water and imidacloprid in water at 100 ppm were used as control. No changes in aphid num Pesticidal Effect Against Drosophila bers were observed in these two groups. With reference to FIG. 15, treatment with Blend 38 (label 55 Drosophila were exposed to two different groups of com “B5049” and “TT 1A, exemplified in Ingredient Family 12 positions before the mortality of drosophila was measured. in Table 6) alone at 1% by volume resulted in 33% decrease in With reference to Table 13, the first group of compositions aphid number on the plants (33 Percent Control as shown in contained Blend 39 (exemplified in Ingredient Family 8 in FIG. 15). Treatment with Blend 39 (label “B5053” and “TT Table 6) alone at LD50 and applied to drosophila. The treat 3C, exemplified in Ingredient Family 8 in Table 6) alone at 60 ment killed 50% drosophila. The second group of composi 1% by volume also resulted in 33% decrease in aphid number on the plants (33 Percent Control as shown in FIG. 15). tions contained Blend 39 at LD50 combined with fipronil at 5 With reference to FIG. 15, treatment with the composition ppm and applied to drosophila. The treatment killed 95% containing the mixture of Blend 38 (labeled “B5049” and drosophila. The result demonstrates that the combination of “TT 1A) at 1% by volume and 100 ppm imidacloprid 65 fipronil and Blend 39 is effective and has a synergistic effect. resulted in complete elimination of aphids on the plants (100 Blends listed in Table 13, alone and in combination with Percent Control as shown in FIG. 15). Treatment with Blend fipronil, were tested in the same fashion. US 8,691,256 B2 105 106 TABLE 13 GCCATCGGCAGATCAGATCCTG 3. and 3. oligonucleotide: 5 taatctagaTCAATTCAGGCCCA Pesticidal Effects of Blends with and without Fipronil GAAGTCGCTTG 3'. Capitalized letters match the tyramine Fipronil receptor sequence. An added Kozak sequence (Grosmaitre, Concen- Kill 90 Celegans 5 X., Jacquin-Joly, E., 2001 Mamestra brassicae putative octo pamine receptor (OAR) mRNA, complete cds. NCBI direct Exemplified tration by wo submission, Accession AF43878) is indicated by underlined in Table 6 LDSO Volume Fipronil w Fipronil nucleotides. The 5' oligonucleotide also contains an EcoRI Blend 39 Ingredient 75 ppm 5 ppm SO% 95% site and the 3' oligonucleotide a Xba I site. The PCR is Family 8 10 performed using Vent polymerase (New England Biolabs) Blend 42 Ingredient 46 ppm 5 ppm 41% 100% with the following conditions: about 95°C., about 5 min for Family 3 about 1 cycle; about 95°C., about 30 sec; and about 70° C., about 90 sec for about 40 cycles and about 70° C., about 10 These results demonstrate that these combinations of min for about 1 cycle. fipronil and the blends are effective and have synergistic 15 The PCR product is digested with EcoRI and Xba I, effects. subcloned into pcDNA 3 (Invitrogen) and sequenced on both strands by automated DNA sequencing (Vanderbilt Cancer Example 16 Center). When this open reading frame is translated to pro tein, it is found to correctly match the published tyramine Pesticidal Effect Against Drosphila melanogaster receptor sequence (Saudou, et al., The EMBO Journal vol 9 no 1, 6-617). For expression in Drosophila Schneider cells, With reference to Table 14, the pesticidal effect against the TyrR ORF is excised from pcDNA3 and inserted into drosphila melanogaster was determined for Blend 19 and the pAC5.1/V5-His(B) pAc5(B) using the Eco RI and Xba I composition including fipnoril and Blend 19. The composi restriction sites. tion of Blend 19 is also exemplified in Ingredient Family 15 of 25 For transfection, Drosophila Schneider cells are stably Table 6. Each composition was tested on 30 drosphila mela transfected with pAc5(B)-TyrRORF using the calcium phos nogaster by spraying directly onto the flies. Treatment with phate-DNA coprecipitation protocol as described by Invitro acetone as control at 0.5ul/fly killed 2 flies, fipnoril alone at gen Drosophila Expression System (DES) manual. The pre 20 ppm killed 12 flies, and Blend 19 alone at 0.5ug/fly killed cipitation protocol is the same for either transient or stable 10 flies. However, treatment with fipronil at 20 ppm com 30 transfection except for the use of an antibiotic resistant plas bined with Blend 19 at 0.5ug/fly killed all of the 30 flies. The mid for stable transfection. At least about ten clones of stably composition including Blend 19 and fipronil was shown to be transfected cells are selected and separately propagated. effective and was shown to have a synergistic effect. Stable clones expressing the receptors are selected by whole cell binding/uptake using H-tyramine. For this assay, cells TABLE 1.4 35 are washed and collected in insect saline (170 mM NaCl, 6 mMKC1, 2 mM NaHCO, 17 mM glucose, 6 mMNaH2PO, Toxicity of Blend 19 with and without Fipronil against Drosphila Melanogaster 2 mM CaCl2, and 4 mM MgCl). About 3 million cells in about 1 mL insect saline are incubated with about 4 nM # of Flies H-tyramine at about 23° C. for about 5 minutes. Cells are Chemical Dose (amount fly) Killed 40 centrifuged for about 30 seconds and the binding solution is Control (acetone) 0.5 ul 2.30 aspirated. The cell pellets are washed with about 500 uL Fipronil 20 ppm 12.30 insect saline and the cells are resuspended and transferred to Blend 19 0.5 g. 10.30 scintillation fluid. Nonspecific binding is determined by Blend 19 + Fipronil 0.5 g Blend 19 + 20 ppm fipronil 30.30 including about 50 uM unlabeled-tyramine in the reaction. 45 Binding is quantified counting radioactivity using a using a Liquid Scintillation B-counter (Beckman, Model LS1801). Example 17 B. Selection of Clones Having the Highest Level of Func tionally Active Tyramine Receptor Protein. Preparation of Stably Transfected Schneider Cell Tyramine receptor binding/uptake is performed to deter Lines with Tyramine Receptor (TyrR) mine which of the transfected clones have the highest levels of functionally active tyramine receptor protein. There are A. PCR Amplification and Subcloning Drosophila mela about 10 clonal lines for tyramine receptor and about 2 p.Ac nogaster Tyramine Receptor. (B) for control. H-tyramine (about 4 nM/reaction) is used as Tyramine receptor is amplified from Drosophila melano a tracer, with and without about 50 uMunlabeled tyramine as gaster head cDNA phage library GH that is obtained through 55 a specific competitor. For this assay, cells are grown in plates the Berkeley Drosophila Genome Project (Baumann, A., and are collected in about 3 ml of medium for cell counting 1999, Drosophila melanogaster mRNA for octopamine and the number of cells is adjusted to about 3x10 cells/ml. receptor, splice variant 1B NCBI direct submission, Acces About two pAcB clones are used in parallel as controls. About sion AJO07617). The nucleic acid sequence and the peptide 1 ml cell Suspension is used per reaction. Based on specific sequence of TyrR are set forth in FIGS. 8A and 8B. Phage 60 binding, about 3 clones express a high levelofactive tyramine DNA is purified from this library using a liquid culture lysate. receptor protein. The clone having the highest specific (Baxter, et al., 1999, Insect Biochem Mol Biol 29, 461–467). tyramine receptor binding (about 90%), is selected for further Briefly, oligonucleotides that are used to amplify the open studies. The selected clone is propagated and stored in liquid reading frame of the Drosophila tyramine receptor (TyrR) nitrogen. Aliquot of the selected clone are grown for whole (Han, et al., 1998, J Neurosci 18, 3650-3658; von Nickisch 65 cell binding and for plasma membrane preparation for kinetic Rosenegk, et al., 1996. Insect Biochem Mol Biol 26, 817 and screening studies. The control pAcB does not demon 827) consist of the 5' oligonucleotide: 5' gcc.gaatticgccaccAT strate any specific binding for the tyramine receptor. US 8,691,256 B2 107 108 C. Efficacy of Schneider Cells Transfected with Tyramine washed one time with about 4 ml ice cold Tris buffer and air Receptor for Screening Compositions for Tyramine Receptor dried before retained radioactivity is measured using LSC. Interaction. Binding data is analyzed by curve fitting (GraphPad software, Cells transfected with the tyramine receptor (about 1x10' Prism). cells/ml) are cultured in each wellofa multi-well plate. About 5 In a saturation binding curve of H-tyramine (H-TA) to 24 hours after plating the cells, the medium is withdrawn and membranes prepared from Schneider cells expressing replaced with about 1 ml insect saline (about 23C). Different tyramine receptor, H-tyramine has a high affinity to tyramine concentrations of H-tyramine (about 0.1-10 nM) are added receptor in the stably transfected cells with p AcB-TyrR with with and without about 10 Munlabeled tyramine and incu K determined to be about 1.257nM and B determined to bated at room temperature (RT). After about a 20 minute 10 incubation, the reaction is stopped by rapid aspiration of the be about 0.679 pmol/mg protein. saline and at least one wash with about 2 ml insect saline In inhibition binding of H-tyramine (H-TA) to mem (about 23C). Cells are solubilized in about 300 u10.3MNaOH branes prepared from Schneider cells expressing tyramine for about 20 min at RT. Solubilized cells are transferred into receptor in the presence and absence of various concentra about 4 ml Liquid Scintillation Solution (LSS) and vigor 15 tions of unlabeled tyramine (TA), the ECs and the K, for ously vortexed for about 30 sec before counting the radioac tyramine against its receptor in Schneider cells expressing tivity using a Liquid Scintillation B-counter (Beckman, tyramine receptor are about 0.331 uM and 0.127 uM, respec Model LS1801) (LSC). tively. Receptor specific binding data is expressed as fimol specific In order to determine the pharmacological profile of binding per 1x10° cells and measured as a function of tyramine receptor (TyrR), the ability of a number of putative H-tyramine concentration. Specific binding values are cal Drosophila neurotransmitters to displace H-tyramine (H- culated as the difference between values in the absence of and TA) binding from membranes expressing tyramine receptor is values in the presence of about 10 uM unlabeled tyramine. tested. In inhibition binding of H-Tyramine to membranes The maximum specific binding occurs at about 5 nM prepared from Schneider cells expressing tyramine receptor H-tyramine. Untransfected cells do not respond to tyramine 25 in the presence and absence of different concentrations of at concentrations as high as about 100 uM. unlabeled ligands (including Tyramine (TA), Octopamine To study the kinetics of the tyramine receptor in stably (OA), Dopamine (DA), and Serotonin (SE)), tyramine dis transfected cells with p AcB-TyrR, crude membrane fractions plays the highest affinity (K, of about 0.127 uM, ECso of about are prepared from the transfected cells and used to calculate 0.305 uM) for the Drosophila TyrR. Octopamine, dopamine the equilibrium dissociation constant (K), Maximum Bind 30 and serotonin were less efficient than tyramine at displacing ing Capacity (B), equilibrium inhibitor dissociation con H-tyramine binding. stant (K) and ECs (effective concentration at which binding With respect to the K, and ECs of the ligands, the rank is inhibited by 50%). A preliminary study to determine the order of potency is aS follows: optimum concentration of membrane protein for receptor tyramine octopamine dopaminesserotonin, showing the binding activity is performed. In this study, different concen 35 trations of protein (about 10-50 ug/reaction) are incubated in likelihood that the stably transfected Schneider cells are about 1 ml binding buffer (50 mM Tris, pH 7.4, 5 mMMgCl, expressing a functionally active tyramine receptor. and 2 mMascorbic acid). The reaction is initiated by the AS Such, Schneider cells expressing tyramine receptor are addition of about 5 nM H-tyramine with and without about effective as a model for studies and Screening for composi 10 uM unlabeled tyramine. After about 1 hr incubation at 40 tions that interact with the tyramine receptor. room temperature, reactions are terminated by filtration through GF/C filters (VWR), which have been previously Example 18 soaked in about 0.3% polyethyleneimine (PEI). The filters are washed one time with about 4 ml ice cold Tris buffer and air In vitro Calcium Mobilization Measurement dried before the retained radioactivity is measured using 45 LSC. Binding data is analyzed by curve fitting (GraphPad Intracellular calcium ion concentrations (Cal.) are mea software, Prism). The data demonstrates no differences sured by using the acetoxymethyl (AM) ester of the fluores between about 10, 20, 30 and 50 ug protein/reaction in cent indicator fura-2 (Enan, et al., Biochem. Pharmacol. Vol tyramine receptor specific binding. Therefore, about 10 ug 51, 447-454). Cells expressing the tyramine receptor are protein/reaction is used. 50 grown under standard conditions. A cell Suspension is pre To determine B, and K values for tyramine receptor pared in assay buffer (140 mMNaCI, 10 mM HEPES, 10 mM (TyrR) in membranes expressing TyrR, Saturation binding glucose, 5 mMKC1, 1 mM CaCl2, 1 mM MgCl) and the cell experiments are performed. Briefly, about 10 ug protein is number is adjusted to about 2x10 cells per ml. Briefly, about incubated with H-tyramine at a range of concentrations 1.0 ml cell suspension (about 2x10 cells) is incubated with (about 0.2-20 nM). Binding data is analyzed by curve fitting 55 (GraphPad software, Prism) and the K for tyramine binding about 511M fura 2/AM for about 30 min at about 28°C. After to its receptor is determined. incubation, the cells are pelleted at about 3700 rpm for about To determine the affinities of several ligands for TyrR, 10 sec at room temperature and then resuspended in about 1.5 increasing concentration of several compounds are tested for ml assay buffer. Ca"1, changes are analyzed in a spectro? their ability to inhibit binding of about 2nMH-tyramine. For 60 luorometer in the presence and absence of test chemicals. both Saturation and inhibition assays total and non-specific Excitation wave lengths are about 340 nm (generated by binding is determined in the absence and presence of about 10 Cat-bound fura-2) and about 380 nm (corresponding to uMunlabeled-tyramine, respectively. Receptor binding reac Cat-free fura-2). The fluorescence intensity is monitored at tions are incubated for about 1 hour at room temperature (RT) an emission wave length of about 510 nm. No absorbance of in restricted light. Reactions are terminated by filtration 65 fluorescence artifacts are observed with any of the com through GF/C filters (VWR), which have been previously pounds used. The ratio of about 340/380 nm is calculated and soaked in about 0.3% polyethyleneimine (PEI). The filters are plotted as a function of time. US 8,691,256 B2 109 110 Example 19 Blends listed in the Table 15, alone and in combination with imidacloprid, were tested in the same fashion. In vitro Cyclic AMP (cAMP) Measurement TABLE 1.5 Cells are grown on dishes and the media changed the day 5 Blend Imidacloprid before the treatment. When cells are approximately 95% con Exemplified Concentration Concentration Results fluent, media is aspirated and the cells are washed one time in Table 6 by Volume by volume Shown with about 5 mL of about 27°C. insect saline (170 mMNaCl, 6.0 mM KC1, 2.0 mM NaHCO, 17.0 mM glucose, 6.0 mM Blend 19 ngredient O.01% 20 ppm FIG. 16A 10 Family 15 NaH2PO, 2.0 mM.CaCl, 4.0 mMMgCl; pH 7.0). About 20 Blend 27 ngredient O.01% 20 ppm FIG. 16B mL of insect saline is added, and cells are harvested by gentle Family 1 scraping. An aliquot of the cells is counted by hemocytom Blend 75 ngredient O.01% 20 ppm FIG.16C Family 7 eter, and the cells are then centrifuged for about 5 minutes at Blend 7 ngredient O.S9/o 20 ppm FIG. 17A about 1000 RPM. Cells are resuspended to give about 3x10 Family 16 cells per mL. IBMX is added to about 200 mu.M. Then about 15 Blend 11 ngredient O.S9/o 20 ppm FIG. 17B 1 mL of cell Suspension is aliquoted for treatment. Forskolin Family 17 Blend 11 ngredient O.S9/o 20 ppm FIG. 17C (cAMP inducing agent), tyramine or different composition Family 18 candidates are added, and the cells are incubated at about 27° Blend 38 ngredient O.S9/o 20 ppm FIG. 18A C. for about 10 minutes. Family 12 Treated cells are centrifuged at about 13000 g for about 10 seconds. The solution is aspirated and about 1 mL of about -20°C. 70% ethanol is added. The cell pellet is disrupted by Blend 75 used in this example is exemplified in Ingredient Vortexing and the samples placed at about -20°C. overnight. Family 7, and contains (wit/wt) 1.00% potassium sorbate, Following the ethanol extraction, cellular debris is pelleted by 0.28% xanthan gum, 81.82% water, 0.04% D-limonene, centrifugation at about 13000 g for about 5 minutes. The 25 0.17% thyme oil white, 8.62% thymol (crystal), 0.33% alpha Supernatant is transferred to a tube and lyophilized to dryness terpineol. 3.37% para-cymene, 0.25% linallyl acetate, 0.67% in a rotary speed-vac. The resulting extract is resuspended in Caryophyllene-B, 0.33% Borneol L, 0.16% myrcene, 0.33% about 100 .mu.LTE and used for the cAMP assay. tea tree oil, 0.48% cypress oil, 1.64% peppermint terpenes, The cAMP assay is based on competition binding between and 0.52% linalool 90. endogenous cAMP and H-cAMP to a cAMP binding pro 30 tein. The H-cAMP Biotrak system (Amersham Biosciences) Synergistic effects were observed for all the compositions is used for this assay as per the manufacturers instructions. containing the mixture of imidacloprid and each blend shown Briefly, about 50 .mu.L of the cellular extract is incubated in Table 15. The results are shown in FIGS. 16A-C, 17A-C with about 50 mu.LH-cAMP and about 100 .mu.L. cAMP and 18A. binding protein in an ice bath for about 2-4 hours. Charcoal 35 These combinations of ingredients, when applied to a pest (about 100 .mu.L) is then added and the solution centrifuged expressing the tyramine receptor, also act synergistically to for about 3 minutes at about 4.degree. C. About 200 .mu.L of control the pest. the reaction mixture is removed and levels of sup.3H-cAMP are determined by Scintillation counting. Levels of endog enous cAMP from the cells are calculated using a standard 40 Example 21 curve with cold cAMP ranging from about 0 to 16 pmol per reaction. Synergistic Effects of a Combination of a Blend and Example 20 Fipronil on In vitro Calcium Mobilization 45 Synergistic Effects of a Combination of a Blend and A Schneider cell line was produced that expressed a cell Imidacloprid on In vitro Calcium Mobilization Surface tyramine receptor of Drosophila melanogaster, as described above. Cells of this line were exposed to three A Schneider cell line was produced that expressed a cell different compositions. The first composition contained Surface tyramine receptor of Drosophila melanogaster, as 50 fipronil at 20 ppm. The second solution contained Blend 19 at described above. Cells of this line were exposed to three 0.1% by volume. The third composition contained a mixture different compositions. The first composition contained imi offipronil at 20 ppm and Blend 19 at 0.1% by volume. Blend dacloprid at 20 ppm. The second solution contained Blend 19 19 is exemplified in Ingredient Family 15 in Table 6. The at 0.01% by volume. The third composition contained a mix results of this procedure are shown in FIG. 19 as fluorescence ture of imidacloprid at 20 ppm and Blend 19 at 0.01% by 55 intensity curves corresponding to intracellular calcium ion volume. The results of this procedure are shown in FIG.16A concentrations. as fluorescence intensity curves corresponding to intracellu lar calcium ion concentrations. As shown in FIG. 19, the composition containing the mix As shown in FIG. 16A, the composition containing the ture offipronil and Blend 19 exhibited (1) a much higher peak mixture of imidacloprid and Blend 19 exhibited (1) a much 60 intensity, (2) a higher V, per second, and (3) a much higher higher peak intensity, (2) a higher V, per second, and (3) a intensity at each time point between 25 second and 120 sec higher intensity at each time point between 30 second and 120 onds (end of the calcium measurement) than the composi seconds (end of the calcium measurement) than the compo tions containing either of the ingredients alone. This demon sitions containing either of the ingredients alone. This dem strates that fipronil and Blend 19 act synergistically in this cell onstrates that imidacloprid and Blend 19 act synergistically in 65 system to affect intracellular calcium ion concentrations. this cell system to affect intracellular calcium ion concentra Blends listed in the Table 16, alone and in combination tions. with fipronil, were tested in the same fashion. US 8,691,256 B2 111 112 TABLE 16 described above. Cells of this line were exposed to three different compositions. The first composition contained Blend Fipronil clothianidin (labeled “AMPAgent”) at 0.01% by volume. The Exemplified Concentration Concentration Results second solution contained Blend 19 at 5% by volume. The in Table 6 by Volume by Volume Shown third composition contained a mixture of clothianidin at Blend 19 Ingredient O.1% 20 ppm FIG. 19 0.01% by volume and Blend 19 at 0.1% by volume. Blend 19 Family 15 Blend 42 Ingredient O.1% 2.5 ppm FIG. 20 is exemplified in Ingredient Family 15 in Table 6. The results Family 3 of this procedure are shown in FIG.22 as fluorescence inten Blend 39 Ingredient O.1% 1 ppm FIG. 21 sity curves corresponding to intracellular calcium ion con Family 8 10 centrations. As shown in FIG. 22, the composition containing the mix These results demonstrate that fipronil and the blends act ture offipronil and Blend 19 exhibited (1) a much higher peak synergistically in this cell system to affect intracellular cal intensity, (2) a higher Viper second, and (3) a much higher cium ion concentrations. These combinations of ingredients, intensity at each time point between 20 second and 120 sec when applied to a pest expressing the tyramine receptor, also 15 onds (end of the calcium measurement) than the composi act synergistically to control the pest. tions containing either of the ingredients alone. This demon strates that clothianidin and Blend 19 act synergistically in Example 22 this cell system to affect intracellular calcium ion concentra tions. Synergistic Effects of a Combination of a Blend and This combination of ingredients, when applied to a pest Fipronil on In vitro Calcium and Camp expressing the tyramine receptor, also acts Synergistically to A Schneider cell line was produced that expressed a cell control the pest. Surface tyramine receptor of Drosophila melanogaster, as Example 24 described above. Cells of this line were exposed to two dif 25 ferent compositions. The first composition contained fipronil at 1 ppm. The second solution contained a mixture offipronil Synergistic Effects of a Combination of Blend 7 and at 1 ppm and Blend 27 at 0.1% by volume. The results of this Abamectin on In vitro Calcium Mobilization procedure are shown in Table 17 as changes in intracellular calcium concentrations and changes in intracellular cAMP 30 A Schneider cell line was produced that expressed a cell concentrations induced by the mixture compared to fipronil Surface tyramine receptor of Drosophila melanogaster, as alone. As shown in Table 17 the composition containing the mix described above. Cells of this line were exposed to three ture offipronil and Blend 27 exhibited greater changes in both different compositions. The first composition contained intracellular calcium level and intracellular cAMP level. This abamectin at 20 ppm. The second solution contained Blend 7 demonstrates that fipronil and Blend 27 act synergistically in 35 at 0.01% by volume. The third composition contained a mix this cell system to affect intracellular calcium ion concentra ture of abamectin at 20 ppm and Blend 7 at 0.01% by volume. tion and intracellular cAMP concentration. The composition of Blend 7 was exemplified in Ingredient Blends listed in the Table 17, alone and in combination Family 5 of Table 6. Blend 7 is exemplified in Ingredient with fipronil, were tested in the same fashion. Family 5 in Table 6. The results of this procedure are shown TABLE 17 Blend Fipronil Ca"),96 Change cAMP,96 Change Exemplified Concentration Concentration as Compared as Compared in Table 6 by Volume by Volume to Fipronil Alone to Fipronil Alone Blend 27 Ingredient O.1% 1 ppm 150% 13.0% Family 1 Blend 39 Ingredient O.1% 1 ppm 18.0% 1.45% Family 8 Blend 42 Ingredient O.1% 1 ppm 33.3% 1.70% Family 3 Blend 12 Ingredient O.1% 1 ppm 16.7% -25% Family 9

These results demonstrate that fipronil and the blends act in FIG. 23A as fluorescence intensity curves corresponding to synergistically in this cell system to affect intracellular cal 55 intracellular calcium ion concentrations. cium ion concentrations and cAMP concencentrations. These As shown in FIG. 23A, the composition containing the combinations of ingredients, when applied to a pest express mixture of imidacloprid and Blend 7 exhibited (1) a much ing the tyramine receptor, also act synergistically to control higher peak intensity, (2) a higher V. per second, and (3) a the pest. much higher intensity at each time point between 25 second 60 and 120 seconds (end of the calcium measurement) than the Example 23 compositions containing either of the ingredients alone. This demonstrates that imidacloprid and Blend 7 act synergisti Synergistic Effects of a Combination of Blend 19 cally in this cell system to affect intracellular calcium ion and Clothianidin on In vitro Calcium Mobilization concentrations. 65 This combination of ingredients, when applied to a pest A Schneider cell line was produced that expressed a cell expressing the tyramine receptor, also acts Synergistically to Surface tyramine receptor of Drosophila melanogaster, as control the pest. US 8,691,256 B2 113 114 Example 25 are shown in FIG. 23B as fluorescence intensity curves cor responding to intracellular calcium ion concentrations. Synergistic Effects of a Combination of Blend As shown in FIG. 23B, the intracellular calcium concen 1 1/Blend 39 and Vohimbine/Forskolin/Genistein on tration profiles were nearly identical for the Blend 7 alone and In vitro Calcium Mobilization 5 Blend 7 with imidacloprid. This demonstrates that imidaclo prid and Blend 7 do not act synergistically in this cell system A Schneider cell line was produced that expressed a cell to affect intracellular calcium ion concentrations. Surface tyramine receptor of Drosophila melanogaster, as Blends listed in the Table 18, alone and in combination described above. Cells of this line were exposed to three with imidacloprid, were tested in the same fashion. different compositions. With reference to FIG. 24A, the first 10 composition contained yohimbine at 100 ppm. With refer TABLE 1.8 ence to FIG. 25A, the second solution contained Blend 11 at Blend Imidacloprid 0.1 mg/ml by Volume, and the third composition contained a Exemplified Concentration Concentration Results in Table 6 by Volume by Volume Shown mixture of yohimbine at 100 ppm and Blend 11 at 0.1% by 15 volume. Blend 11 was labeled as “B5028” and exemplified in Blend 7 Ingredient O.S9/o 20 ppm FIG. 23B Ingredient Family 2 of Table 6. The results of this procedure Family 5 Blend 11 Ingredient O.S9/o 20 ppm FIG. 17D are shown in FIGS. 24A and 25A as fluorescence intensity Family 2 curves corresponding to intracellular calcium ion concentra Blend 39 Ingredient O.S9/o 20 ppm FIG. 18B tions. Family 8 As shown in FIG. 25A, the composition containing the mixture of yohimbine and Blend 11 exhibited (1) a higher As shown in FIGS. 23B, 17D and 18B, the intracellular peak intensity, (2) a higher V, per second, and (3) a higher calcium concentration profiles were nearly identical for the intensity at each time point between 25 second and 120 sec Blends 11 and 39 alone and each blend with imidacloprid. onds (end of the calcium measurement) than the composi 25 This demonstrates that imidacloprid and Blends 7, 11 and 39 tions containing either of the ingredients alone. This demon do not act synergistically in this cell system to affect intrac strates that yohimbine and Blend 11 act synergistically in this ellular calcium ion concentrations. cell system to affect intracellular calcium ion concentrations. The synergistic effects of the combinations of Blend 11 at Example 27 0.1 mg/ml by volume with forskolin at 100 ppm and with 30 genistein at 100 ppm were also tested in the same fashion, as Negligible Effects of a Combination of Blend 27 and shown in FIG. 25. Yohimbine/Forskolin/Genistein on In vitro Calcium Similarly, the synergistic effects of the combinations of Mobilization Blend 39 at 0.1 mg/ml by volume with forskolin at 100 ppm and with genistein at 100 ppm were tested in the same fashion, 35 A Schneider cell line was produced that expressed a cell as shown in FIG. 26. Blend 39 was labeled as “B5053 and Surface tyramine receptor of Drosophila melanogaster, as exemplified in Ingredient Family 8 of Table 6. described above. Cells of this line were exposed to three Similarly, the synergistic effects of the combinations of different compositions. The first composition contained Blend 41 at 0.1 mg/ml with genistein at 100 ppm were tested yohimbine at 100 ppm. The second solution contained Blend in the same fashion, as shown in FIG. 27. Blend 41 was 40 27 at 0.1% by volume. The third composition contained a labeled as AAL and exemplified in Ingredient Family 13 of mixture of yohimbine at 100 ppm and Blend 27 at 0.1% by Table 6. volume. Blend 27 was labeled as “B7001” and exemplified in Furthermore, the synergistic effects of the combinations of Ingredient Family 1 of Table 6. The results of this procedure Blend 42 at 0.1 mg/ml with genistein at 100 ppm were tested are shown in FIG. 28 as fluorescence intensity curves corre in the same fashion, as shown in FIG. 27. Blend 42 was 45 sponding to intracellular calcium ion concentrations. labeled as AAT and exemplified in Ingredient Family 3 of As shown in FIG. 28, the intracellular calcium concentra Table 6. tion profiles for the Blend 27 alone and Blend 27 with yohim The results demonstrate that these combinations of ingre bine were nearly identical. This demonstrates that yohimbine dients act synergistically in this cell system to affect intrac and Blend 27 do not act synergistically in this cell system to ellular calcium ion concentrations. These combinations of 50 affect intracellular calcium ion concentrations. ingredients, when applied to a pest expressing the tyramine Similarly, the combination of Blend 27 at 0.1% by volume receptor, also act synergistically to control the pest. with forskolin at 100 ppm and with genistein at 100 ppm were tested in the same fashion, as shown in FIG. 28. This demon Example 26 strates that Blend 27 do not act synergistically with yohim 55 bine, forskolin or genistein in this cell System to affect intra Negligible Effects of a Combination of a Blend and cellular calcium ion concentrations. Imidacloprid on In vitro Calcium Mobilization Example 28 A Schneider cell line was produced that expressed a cell Surface tyramine receptor of Drosophila melanogaster, as 60 Inhibitive Effects of a Combination of Blend 58 and described above. Cells of this line were exposed to three Yohimbine/Forskolin/Genistein on In vitro Calcium different compositions. The first composition contained imi Mobilization dacloprid at 20 ppm. The second solution contained Blend 7 at 0.01% by volume. The third composition contained a mix A Schneider cell line was produced that expressed a cell ture of imidacloprid at 20 ppm and Blend 7 at 0.01% by 65 Surface tyramine receptor of Drosophila melanogaster, as volume. The composition of Blend 7 was exemplified in described above. Cells of this line were exposed to three Ingredient Family 5 of Table 6. The results of this procedure different compositions. The first composition contained US 8,691,256 B2 115 116 yohimbine at 100 ppm. The second solution contained Blend composition containing the mixture of imidacloprid and 58 at 0.1% by volume. The third composition contained a thyme oil is indicated by triangles, the curve corresponding to mixture of yohimbine at 100 ppm and Blend 58 at 0.1% by the composition containing the thyme oil alone is indicated volume. Blend 58 was labeled as “B7002 and exemplified in by circles, and the curve corresponding to the composition Ingredient Family 14 of Table 6. The results of this procedure containing imidacloprid alone is indicated by Squares. These are shown in FIG. 29 as fluorescence intensity curves corre curves may be obtained by the following method. sponding to intracellular calcium ion concentrations. As shown in FIG. 31, the composition containing the mix As shown in FIG. 29, the composition containing the mix ture of imidacloprid and thyme oil exhibited a much higher ture of yohimbine and Blend 58 exhibited (1) a lower peak peak intensity and V per second than the compositions intensity, (2) a lowerV per second, and (3) a lower inten 10 containing either of the ingredients alone. This demonstrates sity at each time point between 25 second and 100 seconds. that imidacloprid and thyme oil act synergistically in this cell This demonstrates that yohimbine and Blend 58 do not act system to affect intracellular calcium ion concentrations. synergistically in this cell system to affect intracellular cal This combination of ingredients, when applied to a pest cium ion concentrations. expressing the tyramine receptor, also acts Synergistically to Similarly, the combination of Blend 58 at 0.1% by volume 15 control the pest. with forskolin at 100 ppm and with genistein at 100 ppm were tested in the same fashion, as shown in FIG. 29. The results Example 31 demonstrate that Blend 58 does not act synergistically with forskolin or genistein in this cell system to affect intracellular In vitro Calcium Mobilization Effects of a calcium ion concentrations. Combination of Thyme Oil and Fluoxastrobin Example 29 A Schneider cell line was produced that expressed a cell Surface tyramine receptor of Drosophila melanogaster, as Inhibitive Effects of a Combination of Anise Oil and described above. Cells of this line were exposed to three Yohimbine/Forskolin/Genistein on In vitro Calcium 25 different compositions. The first composition contained flu Mobilization oxastrobin at 1 mg/ml. The second solution contained thyme oil at 1 mg/ml. The third composition contained an approxi A Schneider cell line was produced that expressed a cell mately 50/50 mixture of fluoxastrobin and thyme oil, with the Surface tyramine receptor of Drosophila melanogaster, as mixture contained at a concentration of 1 mg/ml. The results described above. Cells of this line were exposed to three 30 of this screening procedure are shown in FIG. 10 as fluores different compositions. The first composition contained cence intensity curves corresponding to intracellular calcium yohimbine at 100 ppm. The second solution contained anise ion concentrations. In FIG. 10, the curve corresponding to the oil at 0.1% by volume. The third composition contained a composition containing the mixture of fluoxastrobin and mixture of yohimbine at 100 ppm and anise oil at 0.1% by thyme oil is indicated by triangles, the curve corresponding to volume. The results of this procedure are shown in FIG.30A 35 the composition containing the thyme oil alone is indicated as fluorescence intensity curves corresponding to intracellu by Squares, and the curve corresponding to the composition lar calcium ion concentrations. containing fluoxastrobin alone is indicated by circles. These As shown in FIG. 30A, the composition containing the curves may be obtained by the method described above. mixture of fipronil and anise oil exhibited (1) a lower peak As shown in FIG. 32, the composition containing the mix intensity, (2) a lowerV per second, and (3) a lower inten 40 ture of fluoxastrobin and thyme oil exhibited a much higher sity at each time point between 35 second and 120 seconds. peak intensity and V per second than the compositions This demonstrates that yohimbine and anise oil do not act containing either of the ingredients alone. This demonstrates synergistically in this cell system to affect intracellular cal that fluoxastrobin and thyme oil act synergistically in this cell cium ion concentrations. system to affect intracellular calcium ion concentrations. Similarly, the combination of anise oil at 0.1% by volume 45 This combination of ingredients, when applied to a pest with forskolin at 100 ppm and with genistein at 100 ppm were expressing the tyramine receptor, also acts Synergistically to tested in the same fashion, as shown in FIG. 30. The results control the pest. demonstrate that anise oil does not act synergistically with One of ordinary skill in the art will recognize that modifi forskolin or genistein in this cell system to affect intracellular cations and variations are possible without departing from the calcium ion concentrations. 50 teachings of the invention. This description, and particularly the specific details of the exemplary embodiments disclosed, Example 30 is provided primarily for clearness of understanding and no unnecessary limitations are to be understood therefrom, for In vitro Calcium Mobilization Effects of a modifications and other embodiments will become evident to Combination of Thyme Oil and Imidacloprid 55 those skilled in the art upon reading this disclosure and can be made without departing from the spirit or scope of the A Schneider cell line was produced that expressed a cell claimed invention. Surface tyramine receptor of Drosophila melanogaster, as What is claimed is: described above. Cells of this line were exposed to three 1. A composition for controlling a target pest comprising a different compositions. The first composition contained imi 60 first agent and a second agent, wherein the first agent has a dacloprid at 1 mg/ml. The second solution contained thyme first activity against the target pest, the second agent has a oil at 1 mg/ml. The third composition contained an approxi second activity against the target pest, the composition has a mately 50/50 mixture of imidacloprid and thyme oil, with the third activity against the target pest, and the third activity mixture contained at a concentration of 1 mg/ml. The results reflects a synergistic interaction of the first agent and the of this screening procedure are shown in FIG. 31 as fluores 65 Second agent; cence intensity curves corresponding to intracellular calcium wherein the first agent comprises geraniol, isopropyl ion concentrations. In FIG.31, the curve corresponding to the myristate, and thyme oil white; and US 8,691,256 B2 117 118 wherein the second agent comprises imidacloprid. the target pest or a Substrate associated with the target pest, 2. The composition of claim 1, wherein the first agent is thereby achieving synergistic pest control. capable of interacting with a G-protein coupled receptor in the target pest, and wherein the second agent acts on a 6. A method of pest control comprising the steps of molecular target other than the receptor. providing a target pest having at least one target G-protein 3. The composition of claim 1, wherein the first activity coupled receptor, contacting the target pest with the persists for a first period, the second activity persists for a composition of claim 1 comprising at least a first agent second period, the third activity persists for a third period, and and a second agent, wherein the first agent is capable of the third period is longer than either the first period or the interacting with the target receptor to trigger, disrupt or second period. 4. The composition of claim 1, wherein the target pest is a 10 alter a biological function related to the binding of the species belonging to an animal order selected from the group target receptor with the first agent, and wherein the sec consisting of Acari, Anoplura, Araneae, Blattaria, Blattodea, ond agent is capable of interacting with a non-receptor Coleoptera, Collembola, Diptera, Grylloptera, Hemiptera, molecule or step associated with cycling of the target Heteroptera, Homoptera, Hymenoptera, Isopoda, Isoptera, receptor, to disrupt the cycling of the target receptor; Lepidoptera, Mantodea, Mallophaga, Neuroptera, Odonata, 15 wherein the first and second agents in combination Orthoptera, Psocoptera, Rhabditida, Siphonaptera, Sym cooperate to amplify the disrupted or altered function phyla, Thysanura, and Thysanontera. resulting from the binding of the target receptor by the 5. A method of controlling a target pest, comprising admin first agent, resulting in Synergistic control of the pest. istering an effective amount of the composition of claim 1 to k k k k k UNITED STATES PATENT AND TRADEMARK OFFICE CERTIFICATE OF CORRECTION

PATENT NO. : 8,691,256 B2 Page 1 of 1 APPLICATION NO. : 12/936039 DATED : April 8, 2014 INVENTOR(S) : ESSam Enan

It is certified that error appears in the above-identified patent and that said Letters Patent is hereby corrected as shown below:

On the Title Page:

The first or sole Notice should read --

Subject to any disclaimer, the term of this patent is extended or adjusted under 35 U.S.C. 154(b) by 358 days.

Signed and Sealed this Twenty-ninth Day of September, 2015 74-4-04- 2% 4 Michelle K. Lee Director of the United States Patent and Trademark Office