Abstract:
This invention relates to compositions of one or more compounds that incorporate one or more allyl sulfide, allyl disulfide or allyl polysulfide moieties, or one or more allyl sulfide, allyl disulfide or allyl polysulfide moieties and one or more hydroxyl groups, used in effective amount in formulations, including emulsions, to repel blood-feeding ectoparasitic arthropods, including mosquitoes, and to deter them from landing and feeding when applied to the skin, clothing or environment of animals, including humans. Said compounds can include, but are not limited to, 8-allyl-sulfanyloctan-1-ol. This invention also relates to compositions comprising one or more of said compounds in further combination with other arthropod repellent and deterrent compounds, including vanillin. These compounds may be formulated with inert ingredients to form a liquid, gel, paste, soap, spray, aerosol or powder.

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS 
       [0001]    This application claims priority to U.S. Provisional Patent Application No. 60/908481, filed 28 Mar. 2007 and entitled “NOVEL SEMIOCHEMICALS FOR REPELLING ECTOPARASITIC ARTHROPODS AND DETERRING THEIR LANDING AND FEEDING”, which is hereby incorporated herein by reference. 
     
    
     FIELD OF THE INVENTION 
       [0002]    This invention relates to compounds, compositions and methods for repelling blood-feeding ectoparasitic arthropods, and deterring their landing and feeding, by applying in one or more formulations compounds incorporating one or more allyl sulfide, allyl disulfide and/or allyl polysulfide moieties to the skin, clothing or environment of animals, including humans. It further relates to the group of repellent and deterrent compounds comprising allylsulfide aliphatic alcohols, to compositions comprising one or more allyl sulfide, allyl disulfide or allyl polysulfide moieties and one or more hydroxyl groups combined in a further composition with one or more additional compounds, and to formulations of said compositions as emulsions. 
       BACKGROUND OF THE INVENTION 
       [0003]    Haematophagous insects and certain other blood-feeding arthropods are ubiquitous ectoparasites of animals, including humans. In so doing, blood-feeding ectoparasitic arthropods constitute a major source of annoyance to humans and other animals, and are vectors of many microbial diseases, as well as those caused by viruses and virus-like disease agents (Harwood and James 1979). 
         [0004]    Blood-feeding arthropods that annoy man and animals through their biting and feeding activity, and often vector disease-causing pathogens, comprise members of numerous insect taxa, including, but not limited to: flies in the Families Culicidae, Tabanidae, Psychodidae, Simuliidae, Muscidae and Ceratopgonidae (Order Diptera), bugs in the Families Cimicidae and Reduviidae (Order Hemiptera), lice in the Orders Mallophaga and Anoplura, and fleas in the Order Siphonaptera, as well as non-insectan arthropods, particularly ticks and mites in the Order Acari (also known as Acarina). 
         [0005]    An example of a significant annoyance to humans and a major vector of disease-causing pathogens is the yellow fever mosquito,  Aedes aegypti  (Diptera: Culicidae), an exceptionally resilient blood-feeding species that breeds in any small container of water (Malavige et al. 2004). Adults are highly domesticated, typically resting indoors in dwellings, thus optimizing their opportunity to feed and vector pathogens that cause diseases such as yellow fever and dengue fever (Mackenzie et al. 2004; Malavige et al. 2004; Hill et al. 2005). Annually, 7.2 million humans become infected with yellow fever, and &gt;30,000 die from the disease. Moreover, 50-100 million humans are infected with dengue fever (500,000 with its henmorhagic form), resulting in approximately 24,000 deaths annually (Zanotto et al. 1996; Mairuhu et al. 2004). Other species of  Aedes,  as well as mosquitoes in other genera, particularly  Anopheles  and  Culex,  are also significant annoyance agents and vectors of disease-causing pathogens. 
         [0006]    The most effective protection against mosquitoes and other ectoparasitic arthropods, is to repel them from, or deter their landing and feeding on, potential hosts. Until recently, the most efficacious known “repellent” was N,N-diethyl-m-toluamide (DEET) (Fradin and Day 2002). There are concerns associated with N,N-diethyl-m-toluamide. It is a solvent for some plastics, paints, varnishes and synthetic fabrics (Trigg 1996; Badolo et al. 2004; Miot et al. 2004). When used alone it may attract rather than repel  A. aegypti.  Finally, products exceeding 30% N,N-diethyl-m-toluamide are not recommended for protection of children (Pest Management Regulatory Agency 2002). Thus, there is a strong need for alternatives to N,N-diethyl-m-toluamide. 
         [0007]    Research has led to several alternative repellents and deterrents to date (TABLE 1), some of them with efficacy equal to that of N,N-diethyl-m-toluamide (Barnard and Xue 2004). Many of these are natural compositions, and include essential oils from plants (cedar, rosemary, eucalyptus, andiroba, catnip, thyme, neem, clove, soybean) and grease or oils from animals. Active ingredients in some of these oils have been isolated and formulated in commercial products. OFF!® botanicals, for example, contain p-menthane-3,8-diol from lemon eucalyptus,  Eucalyptus maculata citriodon  (Beldock et al. 1997; Carroll and Loye 2006) as the active ingredient. Other new repellents for mosquitoes and other arthropods that are found in natural sources include: 2-undecanone (methyl nonyl ketone) from tomato plants (Roe 2002, 2004, 2007; Roe et al. 2006); tetrahydronootkatone (1,4,4a,5,6,7,8,10-octahydro-6-isopropyl-4,4a-dimethyl-2(1H)-naphthalenone) and 1,10-dihydronootkatone (1,4,4a,5,6,7,8,10-octahydro-6-isopropenyl-4,4a-dimethyl-2(1H)-naphthalenone) from yellow cedar (Zhu et al. 2005); and callicarpenal (13,14,15,16-tetranor-3-cleroden-12-al) and intermedeol [(4S,5S,7R, 10S)-eudesm-11-en-4-ol] from American beautyberry (Cantrell et al. 2005, 2006; Carroll et al. 2007). 
         [0000]    
       
         
               
             
               
             
           
               
                 TABLE 1 
               
               
                   
               
               
                 Names, ingredients, and formulations of 13 representative mosquito 
               
               
                 repellents. Concentration of ingredients as stated on product label. 
               
               
                 Names and ingredients 
               
               
                   
               
             
             
               
                   
               
             
          
           
               
                 Neem Aura:  Aloe vera,  extract of barberry, camomile, goldenseal, myrrh, 
               
               
                 neem, and thyme; oil of anise, cedary citronella, coconut, lavender, 
               
               
                 lemongrass, neem, orange, rhodiumwood. NeemAura Naturals, Inc., 
               
               
                 Alachua, FL 
               
               
                 GonE!:  Aloe vera,  camphor, menthol, oils of eucalyptus, lavender, 
               
               
                 rosemary, sage, and soybean. Aubrey Organics, Tampa, FL 
               
               
                 SunSwat: oils of bay, cedarwood, citronella, goldenseal, juniper, lavender, 
               
               
                 lemon peel, patchouli, pennyroyal, tea tree, and vetivert. Kiss My Face 
               
               
                 Corp., Gardiner, NY 
               
               
                 Natrapel: citronella (10%). Tender Corp., Littleton, NH 
               
               
                 Bygone: oils of canola, eucalyptus, peppermint, rosemary, and sweetbirch. 
               
               
                 Lakon Herbals, Inc., Montpelier, VT 
               
               
                 Bite Blocker: glycerin, lecithin, vanillin, oils of coconut, geranium, and 
               
               
                 soybean (2%). Homs, LLC, Clayton, NC 
               
               
                 BioUD8: undecanone (7.75%). Homs, LLC, Clayton, NC 
               
               
                 Skinsations: N,N-diethyl-m-toluamide (7%). Spectrum Corp., St. 
               
               
                 Louis, MO 
               
               
                 Off!: N,N-diethyl-m-toluamide (15%). S. C. Johnson &amp; Sons, 
               
               
                 Inc., Racine, WI 
               
               
                 Avon Skin-So-Soft Bug Guard plus IR3535: 3-(N-butyl-N-acetyl)-amino 
               
               
                 propionic acid, ethyl ester (7.5%). Avon Products, Inc., New York 
               
               
                 Autan Active Insect Repellent: 1-(1-methyl-propoxycarbonyl)-2- 
               
               
                 (2-hydroxy-ethyl)-piperidine (Picaridin KBR-3023) (10%). Bayer Ltd., 
               
               
                 Dublin, Ireland 
               
               
                 Repel: lemon eucalyptus insect repellent lotion. Oil of lemon 
               
               
                 eucalyptus (65% p-methane-3,8-diol [PMD]). Wisconsin Pharmacal Co., 
               
               
                 Inc., Jackson, WI 
               
               
                 MosquitoSafe: geraniol 25%, mineral oil 74%,  Aloe vera  1%. Naturale 
               
               
                 Ltd., Great Neck, NY 
               
               
                   
               
             
          
         
       
     
         [0008]    Many of the patented repellents and deterrents for arthropods are compositions. These are of two types: 1) compositions comprising a single active ingredient formulated with one or more inert ingredients that serve as a carrier or stabilizer, and 2) compositions of two or more active ingredients that provide an additive or synergistic effect on efficacy of the composition over that provided by any of the components alone. Compositions of the second type usually also have inert ingredients as formulants. 
         [0009]    Examples of the first type of composition include: p-menthane-3,8-diol in ethylene/vinyl acetate co-polymer (Sikinami et al. 1991); N,N-diethyl-m-toluamide formulated in a liquefiable powder (Nichols 1993); garlic juice in filtered water (McKenzie 1995); carane-3,4-diol in a cellulose matrix (Ishiwateri 1999); and N,N-diethyl-n-toluamide formulated as an emulsion (Ross 2003). 
         [0010]    Compounds used as additives with other repellents include, but are not limited to: vanillin, 1,8-cineole, linalool, citronellal, citronellol, camphor, menthone, isomenthone, menthol, borneol, isomenthol, α-terpineol, cis- and trans-piperitol, nerol, neral, cinnamaldehyde, cumin aldehyde, geraniol, geraniol, thymol, bornyl acetate, menthyl acetate, cumin alcohol, geranyl formate, geranyl acetate, caryophyllene, and cis-cinnamyl acetate. As indicated in some of the following examples, repellent additives are often combined with N,N-diethyl-m-toluamide to improve its efficacy. 
         [0011]    Specific examples of the second type of composition include: 1,2,3,4-tetrahydro-β-naphthol and 2-phenyl cyclohexanol (Pijoan and Jachowski 1950); N,N-diethyl-m-toluamide and halobenzoylproprionate (1977, Dec. 20, U.S. Pat. No. 4,064,268); N,N-diethyl-m-toluamide, citral and citronella oil (Hautmaim 1979); oils of citronella, cedar, wintergreen and pennyroyal in an oleic acid carrier (Sherwood and Sherwood 1992); N,N-diethyl-m-toluamide and N-alkyl neotridecanamide (Polefka et al. 1997); p-menthane-3,8-diol, citronella, geraniol and α-terpineol (Beldock et al. 1997); geraniol, citronellol and nerol (Butler 2001); and N,N-diethyl-m-toluamide and dihydronepalactone (Hallahan 2007). 
         [0012]    Vanillin has been shown to improve the repellence of N,N-diethyl-m-toluamide against black flies (Retnakaran 1984), and extracted oils of four species of plants (of 11 tested) against mosquitoes (Tuetun et al. 2005; Choochote et al. 2007). On the other hand, Fried et al. (2007) teach that “vanillin may be added as a stabilizer” in combination with a number of essential plant oils, but do not report any increase in repellence against flies and mosquitoes. 
         [0013]    Garlic,  Allium sativum,  has well known antibacterial, antihelminthic and antitumor properties (Block 1992). It is also used with uncertain efficacy and understanding as an insect repellent. Bassett (1998) claims repellence of mosquitoes with a composition of garlic juice and hot pepper sauce, but does not reveal the contribution of each of these components. McKenzie (1995) describes a simple method of combining garlic juice and water to make a composition for repelling insects from fruit and vegetable plants. Arand and Arand (2002, 2003) improved on the methodology of McKenzie (1995), by developing a method of combining a measured amount of extract of a garlic puree with an inert carrier to form a composition of known concentration that is then added to a more conventional pesticide to improve its efficacy in a maimer that “is not totally understood at this time.” Similarly, Anderson and Brock (1998) claim that spraying dilute garlic juice on a grassy area can repel mosquitoes for many months, for “reasons not wholly understood”. Arand and Arand (2002, 2003) report improvements in the preparation of garlic extract and the use of said extract in composition with known insecticides. 
         [0014]    Consumption of garlic as a means of repelling mosquitoes is widely practiced (Moore et al. 2006), but with no proven efficacy (Rajan et al. 2005). Weisler (1989) reports that administration of a 1:20 composition of aneurine (Vitamin B 1 ) and garlic oil in the diet of domesticated animals can protect them from ingestion by fleas and ticks. However, in a test with flea-infested dogs, neither component was effective alone, and Weisler (1989) did not disclose whether either aneurine or some component of garlic oil actually was present in the skin of the test dogs. Therefore, the actual role of both components is uncertain. Moreover, Weisler (1989) erroneously teaches that allyl sulfide is the same as garlic oil, when in fact garlic oil is a complex mixture of many compounds (see  FIG. 1 ). 
         [0015]    Topical application of pure garlic oil can also be used to repel mosquitoes. Such an application provided 70 minutes of protection against  A. aegypti  (Trongtokit et al. 2005). Similarly, garlic oil (1%) formulated in petroleum jelly and beeswax provided 8 hours of protection against  Culex fatigans  (Bhuyan et al. 1974). 
         [0016]    No systematic experimental study has been done to determine the identity of potentially bio-active compounds in garlic and garlic oil. Therefore, the ingredients therein that express repellence and deterrence to blood-feeding insects and other blood-feeding arthropods are unknown. 
       SUMMARY OF THE INVENTION 
       [0017]    In general terms this invention pertains to compounds found in garlic, or synthetic compounds of related structure, that incorporate one or more of allyl sulfide, allyl disulfide, and/or allyl polysulfide moieties, and one or more hydroxyl groups, and are used to repel blood-feeding ectoparasitic arthropods or deter their landing and feeding. Said blood-feeding arthropods can include, but are not limited to, ticks and mites in the Order Acari (also known as Acarina) and insects in the Orders Mallophaga, Anoplura, Siphonaptera, Hemiptera (Families Cimicidae and Reduviidae), and Diptera (Families Culicidae, Tabanidae, Psychodidae, Simuliidae, Muscidae and Ceratopogonidae). The dipteran insects in the family Culicidae can include, but are not limited to, species in the genera  Aedes, Culex, Anopheles, Chagasia, Bironella, Culiseta, Ochlerotatus, Psorophora, Toxorhynchites, Mansonia,  and  Coquillettidia.    
         [0018]    In specific terms said blood-feeding arthropod repellent and deterrent compounds can include, but are not limited to: methylallyl disulfide; [3H]-1,2-dithiolane; diallyl disulfide; methylallyl trisulfide; [4H]-1,2,3-trithiin; diallyl trisulfide; 5-methyl-1,2,3,4-tetrathiane; methylallyl tetrasulfide; [5H]-1,2,3,4-tetrathiepine; diallyl tetrasulfide; 4,5,9,10-tetrathiatrideca-1,12-diene; 6-methyl-4,5,8,9-tetrathiadodeca-1,11-diene; 2-(2,3-dithia-5-hexenyl)-3,4-dihydro-2(H)-thiopyran; 3-(2,3-dithia-5-hexenyl)-3,4-dihydro-2(H)-thiopyran; 1,2-bis-(1,2-dithia-4-pentenyl)-benzene; (E)-4,5-dithia-1,7-octadien-1-yl-benzene; trans-distyryldisulfide; 4,8-dithiaundeca-1,10-diene; 4,11-dithiatetradeca- 1,13-diene; 4,13-dithiahexadeca-1,15-diene; and 4,12-dithiapentadeca-1,14-diene. 
         [0019]    Additional blood-feeding ectoparasitic arthropod repellent and deterrent compounds can be saturated or unsaturated primary, secondary or tertiary alcohols with an allylsulfide, allyl disulfide or allyl polysulfide moiety, including, but not limited to: 9-allylsulfanylnonan-1-ol, 9-allylsulfanylnonan-2-ol, 8-allylsulfanyloctan-1-ol, 8-allylsulfanyloctan-2-ol, 8-allylsulfanyloctan-3-ol, 8-allylsulfanyloctan-4-ol, 7-allylsulfanylheptan-1-ol, 7-allylsulfanylheptan-2-ol, 6-allylsulfanylhexan-1-ol, and 6-allylsulfanylhexan-2-ol. 
         [0020]    In a further aspect, this invention pertains to compositions comprising one or more compounds with one or more allyl sulfide, allyl disulfide, or allyl polysulfide moieties, and/or one or more hydroxyl groups, in combination with one or more additional blood-feeding arthropod repellent and deterrent compounds selected from the group including, but not limited to, the following: vanillin; 1,8-cineole; linalool; citronellal; citronellol; camphor; menthone; isomenthone; menthol; borneol; isomenthol; α-teipineol; cis- and trans-piperitol; nerol; neral; cinnamaldehyde; cumin aldehyde; geraniol; geranial; thymol; bornyl acetate; menthyl acetate, cumin alcohol; geranyl formate; geranyl acetate; caryophyllene; cis-cinnamyl acetate, N,N-diethyl-m-toluamide, p-menthane-3,8-diol, 2-undecanone, tetrahydronootkatone, 1,10-dihydronootkatone, callicaipenal, and intermedeol. 
         [0021]    In a final aspect, this invention pertains to methods of repelling blood feeding ectoparasitic arthropods, by applying said compositions of one or more compounds that incorporate one or more allyl sulfide, allyl disulfide, or allyl polysulfide moieties, and one or more hydroxyl groups, alone or in further compositions with one or more of said additional blood-feeding arthropod repellent and deterrent compounds, in effective amounts ranging from 1 nanogram to 100 milligrams per cm 2  of surface area in formulations, including emulsions, as a liquid, gel, paste, soap, spray, aerosol or powder to the skin, clothing or environment of an animal. Said animals can be an amphibian, reptile, bird or mammal, including a human. Said environment of an animal can include, but is not limited to, bedding, furniture, dwellings, vehicles and plants. 
     
    
     
       DESCRIPTION OF THE DRAWINGS 
         [0022]      FIG. 1  illustrates flame ionization detector (FID) and electroantennograhic detector (EAD: male or female  Aedes aegypti  antennae) responses to aliquots of the low boiling temperature portion of garlic oil extract. Chromatography: Hewlett Packard (HP) gas chromatograph equipped with a fused silica column (30 m×0.32 mm ID) coated with DB-5; injector and detector temperature: 250° C.; temperature program: 50° C. (held for 1 min, 10° C. per min to 160° C., then 20° C. per min to 300° C. The molecular structure and name of Compounds 1-10 that elicited responses from antennae are depicted above the GC-EAD trace. Antennal stimulatory components 11, 12, 13 and 14 (see TABLE 2) elute later than 13.5 min in the gas chromatogram. Trace components 11-14 are detectable by FID and EAD particularly after heating garlic oil in open Petri dishes at 40° C. for 12 hours, thus enhancing their absolute and relative abundance. 
           [0023]      FIG. 2  illustrates a four-step, two-pot synthetic pathway to 8-allylsulfanyloctan-1-ol. 
           [0024]      FIG. 3  shows the duration of protection of a 100 cm 2  area of an exposed human forearm from bites by  Aedes aegypti  caused by 5%, 10% and 25% formulations of 8-allylsulfanyloctaii-1-ol in mineral oil applied to the skin at a dose of 1.5 mg per cm 2 , as opposed to no protection provided by mineral oil alone. 
           [0025]      FIG. 4  shows the duration of protection of a 100 cm 2  area of an exposed human forearm from bites by  Anopheles gambiae  caused by 5% and 10% formulations of 8-allylsulfanyloctan-1-ol in mineral oil applied to the skin at a dose of 1.5 mg per cm 2 , as opposed to negligible protection provided by mineral oil alone. 
           [0026]      FIG. 5  shows the duration of protection of a 100 cm 2  area of an exposed human forearm from bites by  Culex quinquefasciatus  caused by 2% and 10% formulations of 8-allylsulfanyloctan-1-ol in mineral oil applied to the skin at a dose of 1.5 mg per cm 2 , as opposed to negligible protection provided by mineral oil alone.* In each of the three replicates in Experiment 11, there was complete protection against bites when the experiment was terminated after eight hours. 
           [0027]      FIG. 6  shows the duration of protection of a 100 cm 2  area of an exposed human forearm from bites by  Anopheles gambiae  caused by a 2% formulation of 8-allylsulfanyloctan-1-ol in mineral oil applied at a dose of 1.5 mg per cm 2 , compared to the duration of protection caused by a 2% formulation of N,N-diethyl-m-toluamide (DEET) applied at the same dose. 
           [0028]      FIG. 7  shows the duration of protection of a 100 cm 2  area of an exposed human forearm from bites by  Aedes aegypti  caused by a composition of 8-allylsulfanyloctan-1-ol (6.7%) in Emulsion No. 1, vanillin (6.7%) in Emulsion No. 1, both vanillin (6.7%) and 8-allylsulfanyloctan-1-ol (6.7%) in Emulsion No. 1, and by Emulsion No. 1 alone. All compositions applied to the skin at 1.5 mg per cm 2 . 
           [0029]      FIG. 8  shows the duration of protection of a 100 cm 2  area of an exposed human forearm from bites by  Anopheles gambiae  caused by a composition of 8-allylsulfanyl-octan-1-ol (2.0%) in Emulsion No. 1, vanillin (2.0%) in Emulsion No. 1, both vanillin (2.0%) and 8-allylsulfanyloctan-1-ol (2.0%) in Emulsion No. 1, and by Emulsion No. 1 alone. All compositions applied to the skin at 1.5 mg per cm 2 .* In two of the three replicates in Experiment 20, there was complete protection against bites when the experiment was terminated after eight hours. 
           [0030]      FIG. 9  shows the duration of protection of a 100 cm 2  area of an exposed human forearm from bites by  Culex quinquefasciatus  caused by a composition of 8-allylsulfanyl-octan-1-ol (2.0%) in Emulsion No. 1, vanillin (2.0%) in Emulsion No. 1, both vanillin (2.0%) and 8-allylsulfanyloctan-1-ol (2.0%) in Emulsion No. 1, and by Emulsion No. 1 alone. All compositions applied to the skin at 1.5 mg per cm 2 . 
           [0031]      FIG. 10  shows the duration of protection of a 100 cm 2  area of an exposed human forearm from bites by  Aedes aegypti  caused by a formulation of N,N-diethyl-m-toluamide (DEET)(6.7%) in ethanol in comparison to a composition of Emulsion No. 1 with vanillin (6.7%) and 8-allylsulfanyloctan-1-ol (6.7%). Both formulations applied to the skin at 1.5 mg per cm 2 . 
           [0032]      FIG. 11  shows the duration of protection of a 100 cm 2  area of an exposed human forearm from bites by  Aedes aegypti  caused by a formulation of N,N-diethyl-m-toluamide (DEET) (10.4%) in ethanol in comparison to a composition of Emulsion No. 2 with vanillin (7.9%) and 8-allylsulfanyloctan-1-ol (10.4%). Both compositions applied to the skin at 1.5 mg per cm 2 . 
       
    
    
     DETAILED DESCRIPTION OF THE INVENTION 
       [0033]    Past research and practice has demonstrated that garlic,  Allium sativum,  and preparations therefrom, can be repellent to mosquitoes and other blood-feeding ectoparasitic arthropods. One study claims that allyl sulfide in combination with Vitamin B 1  causes repellence of fleas when ingested by dogs, but neither Vitamin B 1  nor garlic oil (erroneously assumed to be composed solely of allyl sulfide) was effective alone, and repellence caused directly by the blend was never demonstrated. 
         [0034]    In contrast to this prior art, we have discovered unexpectedly that repellence of blood-feeding ectoparasitic arthropods, and deterrence of landing and feeding by such arthropods, is imparted by the presence of one or more allyl sulfide, allyl disulfide or allyl polysulfide moieties in various compounds found in garlic oil. We have further discovered unexpectedly that synthetic molecules not found in garlic oil, but incorporating one or more of these moieties in their molecular structure, can have repellent and deterrent properties in excess of those imparted by compounds that occur naturally in garlic or garlic oil. Of particular interest for repellent and deterrent properties is the family of compounds comprising aliphatic alcohols with an allylsulfide moiety, including the novel compound 8-allylsulfanyloctan-1-ol. In accordance with these discoveries, it is an object of this invention to provide methods and compositions that may be used in protecting animals, including humans, from vectored disease-causing pathogens, and from biting and annoyance caused by mosquitoes and other blood-feeding ectoparasitic arthropods. 
       EXAMPLE 1 
     Experimental insects 
       [0035]    The black-eyed Liverpool strain of  Aedes. aegyti  was obtained from Dr. Carl Lowenberger, Simon Fraser University (SFU). Insects were reared under standardized conditions (60-70% relative humidity, 26-28° C., 14 h light: 10 h dark photoperiod) in SFU&#39;s insectary. Neonate larvae that hatched in glass dishes of sterilized hypoxic water were transferred to trays of distilled water provisioned with Nutrafin® Basix Staple Food fish diet. Pupae were collected daily and separated by sex, and 15 females and 10 males were placed in a paper cup (7.5 cm diameter, 8.5 cm high) with a mesh lid. Emergent adults were fed a 10% (w/v) sucrose solution via braided cotton dental rolls. Arm-fed gravid females were offered water-containing paper cups, lined with paper-towel as an oviposition substrate. 
         [0036]    The Ifakara strain of  Anopheles gambiae  was obtained from Dr. Bernard Roitberg, SFU. Insects were reared under standardized conditions (see above) in SFU&#39;s insectary. Neonate larvae that hatched in glass dishes of distilled water were transferred to trays (250-500 larvae/tray) of distilled water, and provisioned with fish diet ad libitum (see above). Pupae were collected and separated daily and placed in a paper cup with a mesh lid (30-45 pupae/cup). Emergent adults were fed a 10% (w/v) sucrose solution via braided cotton dental rolls. Ann-fed gravid females were offered a water-containing dish (9 cm diameter) with moistened filter paper as an oviposition substrate. 
         [0037]      Culex quinquefasciatus  were obtained from Erin Vrzal of the United States Department of Agriculture (USDA), Gainesville, Fla. Insects were reared in SFU&#39;s quarantine facility at 40-50% relative humidity, 25-27° C., and a photoperiod of 14 h light and 10 h dark. Neonate larvae that hatched in glass dishes of distilled water were transferred to trays (250-500 larvae/tray) of distilled water, and provisioned with fish diet ad libitum (see above). Pupae were collected and separated daily and placed in a paper cup with a mesh lid (30-45 pupae/cup). Emergent adults were fed a 10% (w/v) sucrose solution via braided cotton dental rolls. Ann-fed gravid females were offered a water-containing dish for oviposition. 
       EXAMPLE 2 
     General Bioassay Procedure 
       [0038]    Candidate repellents and deterrents were bioassayed according to a modified protocol from the World Health Organization (1996). At least 1 hour prior to each bioassay, 75 host-seeking non blood-fed, nulliparous, 5- to 8-day-old female  Aedes aegypti, Anopheles gambiae  or  Culex quinquefasciatus  were placed into a wood-framed cage (26.5 cm on each side and 42.5 cm high) with a wooden floor, screened mesh sides and top, and a clear acrylic front fitted with a cotton stockinette sleeve (10 cm diameter). The test subject&#39;s arm was covered with an elbow-length polyethylene glove with an excised patch (16.6 cm long, 6 cm wide) to expose the ventral forearm of the test subject. Candidate deterrents were formulated in mineral (paraffin) oil and applied to the exposed forearm 5 min prior to inserting the arm into the cage. The inserted arm remained in the cage for 3 min every 30 min. Prior to each 3-min bioassay period, the hand of the untreated arm was inserted into the cage to ascertain that it received 10 bites within 30 sec as an indication of “biting pressure”. 
         [0039]    The bioassay was terminated when the treated arm received ≧2 bites in one 3-min bioassay period or one bite in each of two consecutive bioassay periods. The time elapsed from experiment initiation to first bite was recorded as deterrent failure or complete protection time. Percentage repellency at the time the deterrent failed to protect the exposed forearm was calculated by the equation (C−T)/C×100, where C and T represent the numbers of mosquitoes landing on and/or biting the control and treatment arm, respectively (Tawatsin et al. 2001). 
         [0040]    N,N-diethyl-m-toluamide formulated in ethanol at a corresponding dose served as a positive control, and mineral oil by itself served as a negative control. On each day, only one candidate compound was tested, ensuring that any residual material in the chamber had disappeared before the next bioassay. 
       EXAMPLE 3 
     Repellence and Deterrence of Garlic Oil and Identification of Bioactive Ingredients 
       [0041]    Garlic oil ( Allium sativum —Mexico; Clearwater Soap Works, Box 1775 RR1, Clearwater, BC V0E 1N0, Canada) was formulated in mineral oil and tested using the general bioassay procedure described in EXAMPLE 2 at a dose of 0.1 mg per cm 2  of arm surface. It expressed repellence and deterrence for ˜30 min. 
         [0042]    Aliquots of garlic oil extracts were then subjected to coupled gas chromatographic-electroantennographic detection (GC-EAD) analysis. Fourteen components (10 shown in  FIG. 1 ) elicited responses from female or male mosquito antennae. Some of these components were isolated by high-performance liquid chromatography (HPLC) for identification by nuclear magnetic resonance spectroscopy (NMR). Other components were identified (compounds 11-14 in TABLE 2) by coupled GC-mass spectrometry and by retention index calculations. Assignment of molecular structure for an antennal stimulatory constituent was confirmed by comparing its GC retention time and mass spectrum with that of an authentic standard that was purchased or synthesized. Four of the 10 components in  FIG. 1  were tentatively identified by comparing their mass spectra and retention indices with those reported in the literature (Block et al. 1998). 
         [0043]    Some of the 10 components in  FIG. 1  were bioassayed singly (see general bioassay procedure) and in various combinations, all of which were as deterrent to mosquitoes as garlic oil. 
       Mosquito-Deterrent Fractions of Garlic Oil 
       [0044]    To determine the most deterrent component(s) in garlic oil, it was fractionated by HPLC into four fractions, each containing one or more of the EAD-active components (compounds 1-10 in  FIG. 1  and 11-14 in TABLE 2). Each fraction was then bioassayed and shown to be similarly effective in repelling and deterring mosquitoes. 
         [0045]    Components with high molecular weight (&gt;250 daltons) are advantageous in that they: 1) dissipate slowly from treated surfaces; 2) provide long-lasting protection against mosquitoes and 3) convey little, if any, offensive smell. The relative abundance of such components was enhanced by treating garlic oil under vacuum, thus stripping away lower-boiling components. In GC-EAD analyses of the high-boiling residue, four components elicited responses from mosquito antennae. They were identified as 4,5,9,10-tetrathiatrideca-1,12-diene, 6-methyl-4,5,8,9-tetrathiadodeca-1,11-diene, 2-(2,3-dithia-5-hexenyl)-3,4-dihydro-2(H)-thiopyran and 3-(2,3-dithia-5-hexenyl)-3,4-dihydro-2(H)-thiopyran (Block et al. 1998) (Compounds 11-14 in TABLE 2), and shown to have a strong repellent and deterrent effect in bioassays. 
       EXAMPLE 4 
     Relationship Between Molecular Structure and Deterrence 
       [0046]    To determine the part of the molecule responsible for deterrence, we synthesized compounds 15-24 (TABLE 2). Synthesis of 8-allylsulfanyloctan-1-ol (Compound 24 in TABLE 2) is described as an example of the preparation of new semiochemical repellents (see  FIG. 2 ). 
         [0047]    1,8-Octanediol (A) was purchased from Alfa Aesar and was converted in a 1-step synthesis to 8-bromo-1-octanol (B, 60-70% yield) by continuous liquid-liquid extraction with n-heptane and 48% aqueous hydrobromic acid. By maintaining the aqueous level at ambient temperature, alcohol B was produced with &gt;99% purity. Warming the aqueous layer to 50-55° C. accelerated the reaction by &gt;10 times but increased the by-product 1,8-dibromooctane from 0.6% to 1.8-2.0%. After removal of heptane in vacuo, alcohol B was used as is. 
         [0048]    Alcohol B (9.7 g, 46.4 mmol) was stirred with thiourea (4.0 g, 52.6 mmol) in 95% ethanol (150 mL). The mixture was refluxed for 6 h to allow formation of the isothiuronium salt C that was not isolated. To this mixture, 5.6 g of KOH pellets were added in one portion. After another 2.5 h of reflux, the mixture was cooled to room temperature and, without isolating thio-alcohol D or its potassium salt, 6 ml of allyl chloride (73 mmol) were added in one portion. After stirring the reaction mixture overnight, water (150 mL) and a 1:1 mixture of ether/hexane (200 mL) were added. Products were extracted, and the organic phase was washed with water and brine, and dried (anh. MgSO 4 ). Solvents were removed in vacuo, and the crude reaction mixture was filtered through silica (25 g), using in sequence hexane and a 1:1 mixture of hexane-ether as eluents to remove non-polar impurities, such as diallyldisulfane, and to obtain desired 8-allylsulfanyl-octan-1-ol (E). The yield of E (&gt;99% pure based on gas chromatography) was 8.1 g (40.0 mmol, 86.2%). The following mass spectrometric fragmentation ions [m/z (relative abundance)] of E were obtained: 203 (M+1, 28), 202 (M, 53), 143 (50), 142 (17), 131 (53), 101 (23), 87 (100), 85 (26), 81 (24), 79 (15), 74 (66), 73 (23), 69 (32), 68 (17), 67 (73), 59 (28), 55 (51), 53 (15), 47 (15), 45 (48), 41 (91). Nuclear magnetic resonance (NMR) data were as follows:  1 H NMR (600 MHz, CD 3 CN): δ 1.20-1.40 (m, 8H), 1.45 (m, 2H), 1.53 (m, 2H), 2.43 (m, 2H), 3.11 (dt, J=7.2, 0.9 Hz, 2H), 3.46 (dt, J=6.6, 5.4 Hz, 2H), 5.05 (tdd, J=10.0, 1.8, 0.9 Hz, 1H) 5.08 (m, 1H), 5.78 (tdd, J=17.0, 10.0, 7.2 Hz, 1H).  13 C NMR (CD 3 CN): δ 26.9, 29.8, 30.2, 30.3, 30.4, 31.4, 33.9, 35.2, 62.8, 117.3, 136.4. 
         [0049]    In bioassays, neither compound 20 (the sulfur atoms replaced by an oxygen atoms) nor compound 21 (containing no hetero atoms) had any repellence or deterrence. Furthermore, taking other results in TABLE 2 and in  FIG. 1  into account, there is compelling evidence that repellence and deterrence is expressed by molecules with one or more allyl sulfide, allyl disulfide, or allyl polysulfide moieties. Further repellence and deterrence can be obtained by combining in the same compound one or more allyl sulfide, allyl disulfide or allyl polysilfide moieties with one or more hydroxyl moieties. 
         [0050]    A compound with commercial appeal should be 1) odorless, 2) stable, 3) easy and inexpensive to synthesize, 4) non-toxic, and 5) deterrent for a long time. Compound 24 (TABLE 2) appears to meet all these requirements. It has almost no detectable odor and it is very potent, even at a low dose. 
         [0000]    
       
         
               
             
               
               
               
               
               
             
           
               
                 TABLE 2 
               
             
             
               
                   
               
               
                 Failure time and percent repellency in experiments with  Aedes aegypti  testing 
               
               
                 garlic oil semiochemicals 11-14, synthetic analogues (Compounds 15-24), and N,N- 
               
               
                 diethyl-m-toluamide (DEET) (Compound 25). 
               
             
          
           
               
                   
                   
                   
                 Failure 
                   
               
               
                   
                   
                 Dosage 
                 time 
                 Percent 
               
               
                   
                 Substance a   
                 mg/cm 2   
                 (min) b   
                 repellence c   
               
               
                   
               
               
                 11 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.1&gt;0.1  
                  067.7 
                 8592 
               
               
                   
               
               
                 12 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.10.5 
                  0102  
                 8797 
               
               
                   
               
               
                 13,14 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.1 
                 35 
                 93 
               
               
                   
               
               
                 15 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.1 
                 34 
                 92 
               
               
                   
               
               
                 16 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.10.5 
                 3536 
                 8992 
               
               
                   
               
               
                 17 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.10.5 
                  036 
                 8997 
               
               
                   
               
               
                 18 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.5 
                 67 
                 93 
               
               
                   
               
               
                 19 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.10.5 
                  0102  
                 9397 
               
               
                   
               
               
                 20 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.1 
                  0 
                 81 
               
               
                   
               
               
                 21 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                 0.1 
                  0 
                 50 
               
               
                   
               
               
                 22 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                  0.05 
                 33 
                 — 
               
               
                   
               
               
                 23 
                 
                   
                             
                     
                         
                         
                     
                   
                 
                  0.05 
                 99 
                 — 
               
               
                   
               
               
                 24 d   
                 
                   
                             
                     
                         
                         
                     
                   
                 
               
               
                   
               
               
                 25 e   
                 
                   
                             
                     
                         
                         
                     
                   
                 
               
               
                   
               
               
                   a Compound names as follows: 11 = 4,5,9,10-tetrathiatrideca-1,12-diene; 12 = 6-methyl-4,5,8,9-tetrathiadodeca-1,11-diene; 13 = 2-(2,3-dithia-5-hexenyl)-3,4-dihydro-2(H)-thiopyran; 14 = 3-(2,3-dithia-5-hexenyl)-3,4-dihydro-2(H)-thiopyran; 15 = 1,2-bis-(1,2-dithia-4-pentenyl)-benzene; 16 = (E)-4,5-dithia-1,7-octadien-1-yl-benzene; 17 = 4,8-dithiaundeca-1,10-diene; 18 = 4,11-dithiatetradeca-1,13-diene; 19 = 4,13-dithiahexadeca-1,15-diene; 20 = 1,10-bis-allyloxy-decane; 21 = octadeca-1,17-diene: 22 = 9-allylsulfanylnonan-1-ol; 23 = 8-allylsulfanyloctan-2-ol; 24 = 8-allylsulfanyloctan-1-ol. 
               
               
                   b A failure time of zero indicates immediate failure (i.e. biting occurred), even though the percent repellence may be high. 
               
               
                   c See EXAMPLE 2 for method of calculating percent repellence. 
               
               
                   d For bioassay result see EXAMPLES 5-7 
               
               
                   e For bioassay result see EXAMPLE 8 
               
             
          
         
       
     
       EXAMPLE 5 
     Repellence and Deterrence of 8-allylsulfanyloctan-1-ol against  Aedes aegypti    
       [0051]    To determine the repellence and deterrence of 8-allylsulfanyloctan-1-ol against  Aedes aegypti,  a 5%, 10% or 25% formulation of 8-allylsulfanyloctan-1-ol in mineral oil was applied in Experiments 1-3 at a dose of 1.5 mg (total composition) per cm 2  to the skin of the test person, and was bioassayed according to the protocol described under EXAMPLE 2. In Experiment 4, mineral oil by itself served as a negative control and was bioassayed at the same dose (1.5 mg per cm 2 ) as in Experiments 1-3. Each of Experiments 1-4 was replicated four times. 
         [0052]    In Experiments 1, 2 and 3, 5%, 10% and 25% formulations of 8-allylsulfanyl-octan-1-ol in mineral oil provided protection from bites by  Aedes egypti  on average for 52 min, 157 min and 305 min, respectively ( FIG. 3 ). In Experiment 4, mineral oil by itself failed to provide any protection from bites ( FIG. 3 ). 
       EXAMPLE 6 
     Repellence and Deterrence of 8-allylsulfanyloctan-1-ol against  Anopheles gambiae    
       [0053]    To determine the repellence and deterrence of 8-allylsulfanyloctan-1-ol against  Anopheles gambiae,  a 2%, 5% or a 10% formulation of 8-allylsulfanyloctan-1-ol in mineral oil was applied in Experiments 5-7 at a dose of 1.5 mg per cm 2  to the skin of the test person, and was bioassayed according to the protocol described under EXAMPLE 2. In Experiment 8, mineral oil by itself served as a negative control and was tested at the same dose (1.5 mg per cm 2 ) as in Experiments 5-7. Each of Experiments 5-8 was replicated 4 times. 
         [0054]    In Experiments 5, 6 and 7, 2%, 5% and 10% formulations of 8-allylsulfanyloctan-1-ol in mineral oil provided protection from bites by  Anopheles gambiae  on average for 297 min, 314 min and 487 min, respectively ( FIG. 4 ). In Experiment 8, mineral oil by itself failed to provide appreciable protection ( FIG. 4 ). 
       EXAMPLE 7 
     Repellence and Deterrence of 8-allylsulfanyloctan-1-ol against  Culex quinquefasciatus    
       [0055]    To determine the repellence and deterrence of 8-allylsulfanyloctan-1-ol against  Culex quinquefasciatus,  a 2% and 5% or a 10% formulation of 8-allylsulfanyloctan-1-ol in mineral oil was applied in Experiments 9-11 at a dose of 1.5 mg per cm 2  to the skin of the test person, and was bioassayed according to the protocol described under EXAMPLE 2. In Experiment 12, mineral oil by itself served as a negative control and was tested at the same dose (1.5 mg per cm 2 ) as in Experiments 9 and 10. Each of Experiments 9-11 was replicated 4 times. 
         [0056]    In Experiments 9, 10 and 11, 2%, 5% and 10% formulations of 8-allylsulfanyloctan-1-ol in mineral oil provided protection from bites by  Culex quinquefasciatus  on average for 198 min, 319 min, and 495 min, respectively ( FIG. 5 ). In each of the three replicates in Experiment 11, there was complete protection against bites when the experiment was terminated after eight hours. Thus, the protection time is even greater than illustrated conservatively in  FIG. 5 . In Experiment 12, mineral oil by itself failed to provide appreciable protection ( FIG. 5 ). 
       EXAMPLE 8 
     Comparison of Repellence and Deterrence caused by N,N-diethyl-m-toluaniide and by 8-allylsulfanyloctan-1-ol against  Anopheles gambiae    
       [0057]    To be able to compare the repellence and deterrence caused N,N-diethyl-m-toluamide (DEET), and by 8-allylsulfanyloctan-1-ol, Experiment 13 tested a 2% formulation of N,N-diethyl-m-toluamiide in ethanol (the best formulant for this compound) for protection from bites by  Anopheles gambiae.  A dose of 1.5 mg per cm 2  was applied to the skin of the test person and bioassayed according to the protocol described under EXAMPLE 2. 
         [0058]    In Experiment 13, a 2% formulation of N,N-diethyl-m-toluamide provided protection from bites by  Anopheles gambiae  on average for 66 min ( FIG. 6 ). Unexpectedly, and in contrast, the duration of protection provided by N,N-diethyl-m-toluamide was only one sixth of the 297 min protection provided by a 2% formulation of 8-allylsulfanyl-octan-1-ol in Experiment 5 ( FIG. 6 ; see also  FIG. 4 ). 
       EXAMPLE 9 
     Formulation of Emulsion No. 1 
       [0059]    The components, amounts and proportions of Emulsion No. 1 were: distilled water 0.35 g, 23%; light paraffin oil (EMD Chemicals) 0.53 g, 35%; glycerol (Anachemia) 0.27g, 18%; soy lecithin (Xenex Labs) 0.21 g, 14%; 8-allylsulfanyloctan-1-ol 2.0-6.7% (if added); ethanol 0.05 mL (quickly evaporating and not remaining as part of the emulsion); vanillin (BDH Laboratory Chemicals) 2.0-6.7% (if added). According to amounts of the active ingredients 8-allylsulfanyloctan-1-ol and vanillin, the percentage of other constituents was slightly adjusted accordingly. 
         [0060]    A vessel was charged with water, glycerol, paraffin oil, soy lecithin, and 8-allylsulfanyl-octan-1-ol (if added) in the above order and mixed to homogeneity after each addition. If vanillin was added, a second vessel was charged with ethanol and vanillin, stirring the mixture until vanillin was completely dissolved; the vanillin solution was then added to the first vessel and vortexed for several minutes to achieve homogeneity. If vanillin was not added, ethanol was added to the first vessel and vortexed for several minutes to achieve homogeneity. 
       EXAMPLE 10 
     Formulation of Emulsion No. 2 
       [0061]    The components, amounts and proportions of Emulsion No. 2 were: Structure ZEA (hydroxypropyl cellulose) (National Starch and Chemical Co.) 0.16 g, 4%; distilled water 2.56 g, 63%; glycerol (Sigma-Aldrich) 0.21 g, 5%; Stepanquat ML [methyl sulfate quaternary ammonium salt of the esterification of oleic acid with N,N,N′,N′-tetrakis(2-hydroxypropyl) ethylene-diamine] (Stepan Co.) 0.12 g, 3%; isopropanol (Anachemia) 0.21 g, 5%; Lipocol L (Lipo Chemicals Inc.) 0.06 g, 2%; 8-allylsulfanyloctan-1-ol 0.41 g, 10%; and vanillin (Sigma-Aldrich) 0.32 g, 8%. 
         [0062]    Structure ZEA was charged into a vessel, water was added, and the components were mixed to homogeneity. A second vessel was charged with the glycerol, the Stepanquat ML, the isopropanol, the Lipocol L, 8-allylsulfanyloctan-1-ol, and vanillin in the above order. After each addition, the resulting mixture was stirred for several minutes. The water/Structure ZEA homogenate was then added slowly to the rapidly mixing organic mixture over several minutes. After mixing was completed, the gross emulsion was homogenized @ 30,000 rpm for several minutes. 
       EXAMPLE 11 
     Test of Emulsion No. 1 Compositions with 8-allylsulfanyloctan-1-ol and Vanillin against  Aedes aegypti    
       [0063]    To compare the repellence and deterrence against  Aedes aegypti  caused by 8-allylsulfanyloctan-1-ol as a single active ingredient and by 8-allylsulfanyloctan-1-ol in combination with vanillin as a second active ingredient, we tested a 6.7% emulsion of vanillin (Experiment 14), a 6.7% emulsion of 8-allylsulfanyloctan-1-ol (Experiment 15), and a 13.4% emulsion of 8-allylsulfanyloctan-1-ol plus vanillin (1:1 ratio) (Experiment 16). 8-Allylsulfanyloctan-1-ol and vanillin were formulated in Emulsion No. 1 (see EXAMPLE 9). In each of Experiments 13-15, test stimuli were applied at a dose of 1.5 mg per cm 2  to the skin of the test person, and were bioassayed according to the protocol described under EXAMPLE 2. In Experiment 17, Emulsion No. 1 served as a positive control and was tested at the same dose (1.5 mg per cm 2 ) as in Experiments 14-16. 
         [0064]    In Experiments 14, 15 and 16, vanillin, 8-allylsulfanyloctan-1-ol, and 8-allylsulfanyl-octan-1-ol plus vanillin provided protection from bites by  Aedes aegypti  on average for 44 min, 77 min and 330 min, respectively ( FIG. 7 ). These results indicate that there is an unexpected synergistic interaction between 8-allylsulfanyloctan-1-ol and vanillin. In Experiment 17, Emulsion No. 1 composition by itself failed to provide any protection ( FIG. 7 ). 
       EXAMPLE 12 
     Test of Emulsion No. 1 Compositions with 8-allylsulfanyloctan-1-ol and Vanillin against  Anopheles gambiae    
       [0065]    To compare the repellence and deterrence against  Anopheles gambiae  caused by 8-allyl-sulfanyloctan-1-ol as a single active ingredient and by 8-allylsulfanyloctaii-1-ol in combination with vanillin as a second active ingredient, we tested a 6.7% emulsion of vanillin (Experiment 18), a 6.7-% emulsion of 8-allylsulfanyl-octan-1-ol (Experiment 19) and a 13.4% emulsion of 8-allylsulfanyloctan-1-ol plus vanillin (1:1 ratio) (Experiment 20). 8-Allylsulfanyloctan-1-ol and vanillin were formulated in Emulsion No. 1 (see EXAMPLE 9). In each of Experiments 18-20, test stimuli were applied at a dose of 1.5 mg per cm 2  to the skin of the test person, and were bioassayed according to the protocol described under EXAMPLE 2. In Experiment 21, Emulsion No. 1 served as a positive control and was tested at the same dose (1.5 mg per cm2) as in experiments 18-20. Each of Experiments 18-21 was replicated three times. 
         [0066]    In Experiments 18-21, vanillin, 8-allylsulfanyloctan-1-ol and 8-allylsulfanyloctan-1-ol plus vanillin provided protection from bites by  Anopheles gambiae  on average for 110 min, 369 min and 462 min, respectively ( FIG. 8 ). In two of the three replicates in Experiment 20, there was complete and unexpected protection against bites for eight hours, after which the experiment was terminated. Thus, the protection time is even greater than illustrated conservatively in  FIG. 8 . The results indicate that there is an interactive effect between 8-allylsulfanyloctan-1-ol and vanillin. In Experiment 21, Emulsion No. 1 by itself failed to provide appreciable protection ( FIG. 8 ). 
       EXAMPLE 13 
     Test of Emulsion No. 1 Compositions with 8-allylsulfanyloctan-1-ol and Vanillin against  Culex quinquefasciatus    
       [0067]    To compare the repellence and deterrence against  Culex quinquefasciatus  caused by 8-allylsulfanyloctan-1-ol as a single active ingredient and by 8-allylsulfanyloctan-1-ol in combination with vanillin as a second active ingredient, we tested a 6.7% emulsion of vanillin (Experiment 22), a 6.7% emulsion of 8-allylsulfanyloctan-1-ol (Experiment 23) and a 13.4% emulsion of 8-allylsulfanyloctan-1-ol plus vanillin (1:1 ratio) (Experiment 24). 8-Allylsulfanyloctan-1-ol and vanillin were formulated in Emulsion No. 1 (see EXAMPLE 9). In each of Experiments 21-24, test stimuli were applied at a dose of 1.5 mg per cm 2  to the skin of the test person, and were bioassayed according to the protocol described under EXAMPLE 2. In Experiment 25, Emulsion No. 1 served as a positive control and was tested at the same dose (1.5 mg per cm 2 ) as in Experiments 22-24. Each of Experiments 22-25 was replicated three times. 
         [0068]    In Experiments 22-24, vanillin, 8-allylsulfanyloctan-1-ol and 8-allylsulfanyloctan-1-ol plus vanillin provided protection from bites by  Culex quinquefasciatus  on average for 88 min, 242 min and 286 min, respectively ( FIG. 9 ). These results indicate that there is an interactive effect between 8-allylsulfanyloctan-1-ol and vanillin. In Experiment 25, Emulsion No. 1 by itself failed to provide any protection ( FIG. 9 ). 
       EXAMPLE 14 
     Comparison of Deterrence against  Aedes aegypti  caused by N,N-diethyl-m-toluamide in Ethanol and by 8-allylsulfanyloctan-1-ol Plus Vanillin in Compositions Formulated in Emulsions No. 1 or No. 2 
       [0069]    To be able to compare the repellence and deterrence caused by N,N-diethyl-m-toluamide and by 8-allylsulfanyloctan-1-ol plus vanillin in Emulsion No. 1 (EXAMPLE 9), Experiment 26 tested a 6.7% formulation of N,N-diethyl-m-toluamide in ethanol (the best formulant for DEET) for protection from bites by  Aedes egypti.  In each replicate, a dose of 1.5 mg per cm was applied to the skin of the test person and bioassayed according to the protocol described under EXAMPLE 2. 
         [0070]    In Experiment 26, N,N-diethyl-m-toluamide provided protection from bites by  Aedes aegypti  on average for 197 min ( FIG. 10 ). 8-Allylsulfanyloctan- 1-ol (6.7%) plus vanillin (6.7%) in Emulsion No. 1 provided protection on average for 333 min ( FIG. 10 ). 
         [0071]    To be able to further compare the deterrence caused by N,N-diethyl-m-toluamide with that caused by 8-allylsulfanyloctan-1-ol plus vanillin, Experiment 27 tested a 10.4% formulation of N,N-diethyl-m-toluamide in ethanol, and Experiment 28 tested 8-allylsulfanyloctan-1-ol (10.4%) plus vanillin (7.9%) in Emulsion No. 2 (EXAMPLE 10). In each replicate of both experiments, a dose of 1.5 mg per cm was applied to the skin of the test person and bioassayed according to the protocol described under EXAMPLE 2. 
         [0072]    In Experiment 28, allylsulfanyloctan-1-ol plus vanillin in Emulsion No. 2 provided protection from bites by  Aedes aegypti  on average for 333 min, considerably longer than the 231 min on average provided by N,N-diethyl-m-toluamide in Experiment 27 ( FIG. 11 ). 
         [0073]    As will be apparent to those skilled in the art in the light of the foregoing disclosure, many alterations and modifications are possible in the practice of this invention without departing from the spirit or scope thereof. Accordingly, the scope of the invention is to be construed in accordance with the substance defined by the following claims. 
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