Classifications

• • A—HUMAN NECESSITIES
  • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
  • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
  • A01N65/00—Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
  • A01N65/08—Magnoliopsida [dicotyledons]
  • A01N65/22—Lamiaceae or Labiatae [Mint family], e.g. thyme, rosemary, skullcap, selfheal, lavender, perilla, pennyroyal, peppermint or spearmint
• • A—HUMAN NECESSITIES
  • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
  • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
  • A01N65/00—Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
  • A01N65/08—Magnoliopsida [dicotyledons]
  • A01N65/10—Apiaceae or Umbelliferae [Carrot family], e.g. parsley, caraway, dill, lovage, fennel or snakebed
• • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
  • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
  • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
  • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
  • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

• the invention relates to natural and safe compounds that are useful as insecticides, bacteriocides, fungicides and nematicides.
• plants and plant extracts have long been used as medicaments, plants are also
• Essential oils are extracted from plant species by various extraction techniques, including steam distillation including the plants belonging to Labiatae and
• Kivanc (1989) have described in vitro activity of various Turkish spice extracts against food-
• borne fungi including Aspergillus spp., Penicillium spp., Rhizopas spp. and Mucor spp.
• Phytophthora capsici was better than the activities of the fungicides carbendazim and pentachlornitrobenzol. Fungitoxic components of the extracts were determined via thin layer chromatography to include carvacrol and thymol.
• T. spicata had the highest activity against the growth of test fungi, Akgul and Kivanc (1989) observed a negligible anti-microbial activity against food-borne fungi Aspergillus sp, . Mucor sp.,
• M. spicata and extracts of mentha-types have activity against 3 phytopathogens: Rynchosporium oryzae, Drechsbra spp. as well as Xanthomonas campestris.
• oils, as well as in formulations described herein vary between about 50-200 ppm, and have the
• the essential oils extracts can be used against plant foliar
• the extracts can be applied to soil as a methyl bromide replacement to kill nematodes, insects, pathogenic fungi and bacteria either alone in drip- watering systems, or together with solarization when applied in vapor form and/or by pouring
• these extracts have beneficial side effects: they can increase the concentration of beneficial microorganisms in soils, such as fluorescent pseudomonads.
• the extracts also have growth promotion effects on plants such as an increased germination rate, most probably due to
• Treated plants are
• the extracts have high thermostability, high volatile activity, a broad spectrum of activity
• Pinpinella anisum contains over 80% anethole. We have found that tr ⁇ «__>-anethole has fumigant activity that may match or exceed methyl bromide.
• IPM integrated pest management
• Figure 1 shows the activity of essential oil extracts from Thymbra spicata (100-400 ppm)
• Figure 2 shows the activity of 100- 1600 ppm essential oils (a mixture o ⁇ Thymbra spicata
• Figure 3 shows the activity of 100- 1600 ppm essential oils (a mixture of Thymbra spicata
• Figure 4 shows the activity of 100-400 ppm essential oils (a mixture of Thymbra spicata
• Figure 5 shows the activity of 100-400 ppm essential oils (a mixture of Thymbra spicata
• Figure 7 shows the activity of 100-400 ppm essential oils (a mixture of Thymbra spicata
• bacteria including Melodoigyne spp. and Phytophthora capsici.
• Figure 8 shows the activity of a bacterium (Pseudomonas fluorescens TR97) isolated
• Figure 9 shows the activity of a bacterium (Pseudomonas fluorescens TR97) isolated
• Figure 10 shows activity of the volatile phase of essential oils extracted from Thymbra
• oils were applied to the lids of the glass petri dishes. 1, Control; 2, 50 ⁇ l/L essential oil; 3, 40 ⁇ l/L essential oil; 4, 90 ⁇ l/L essential oil; 5, 60 ⁇ l/L essential oil; 6, 30 ⁇ l/L essential oil; 7, 80 ⁇ l/L essential oil; 8, 70 ⁇ l/L essential oil; 9, 20 ⁇ l/L essential oil.
• FIG. 11 shows the activity of the volatile phase of essential oils extracted from
• Thymbra spicata (as shown in the Figure as "thyme oil”) was mixed with 0.1 mL olive
• Figure 12 shows in vivo activity of 200 ppm essential oil from Thymbra spicata or copper
• Figure 13 shows the number of shoots exhibiting symptoms of fire blight disease in field
• Figure 14 shows the systemic activity of essential oils on Xanthomonas campestris pv.
• campestris in cabbage The soil was treated with 100-200 ppm essential oil in an aqueous
• campestris (1 x 10 5 CFU/mL). Control treatments were sprayed with 100 ppm Tween 20
• Figure 15 shows the contact activities of essential oil from Thymbra spicata emulsified
• Plants were spray inoculated with Xanthomonas campestris pv. campestris (I lO 5 CFU/mL).
• the control treatment was sprayed with 100 ppm Tween 20 (emulsifying agent) in water.
• Figure 16 shows the contact and systemic activities of essential oil from Thymbra spicata
• Plants were spray inoculated with Xanthomonas campestris pv. campestris (1 x 10 5 CFU/mL).
• Figure 17 shows the effect of essential oil from Thymbra spicata emulsified in water
• the inset illustrates the difference in leaf color between the control (leftmost) treatment and the essential oil treatments (the latter are a darker green).
• Figure 18 shows the toxicity of anethole vapors to the adults of T. confusum and S.
• Figure 19 shows the toxicity of anethole vapors to the eggs of T. confusum and E.
• the essentials oils covered in this application may be extracted from plant species
• Suitable plants may include specimens in the genera Thymbra, Satureja, Origanum, Corydothymus, Pinpinella and Foeniculum. Examples of plant
• Origanum majorana L
• Corydothymus capitatus L.
• Reichb. fil. Origanum vulgare (L) subsp.
• Origanum sriacum var. bevanii Letswart, Origanum onites (L), and Origanum
• the compounds identified from extracts of these plant species are: cis- anethole, trans-anethole, anisaldehyde, anis ketone, anisole, ⁇ -bisabolene, borneol, bornyl acetate, cadinene, camphene, camphor, ⁇ -3-carene, ⁇ -4-carene, carophyllene, carvone, carvacrol, ⁇ -
• caryophyllene cinnamic aldehyde, citral, citronellal, cineol, 1 ,8-cineole, -cymene, ?-cymene-8-
• the major active components are: carvacrol, thymol, cymene and anethole.
• product for commercial use may contain various individual plant extracts or combinations of
• T. spicata (10-90%), S. thymbra (10-90 %>) and Origanum spp. (5-30 %) can be mixed to provide optimal activity against bacteria, viruses and
• mixture containing more carvacrol and anethole is more effective against insects and nematodes.
• the invention also comprises chemically synthesized essential oil components which can
• Anethole may be synthesized relatively easily, for
• the LD 50 values of the components of the essential oils indicate that they are not highly toxic or teratogenic to humans and animals. Although concentrated oils are toxic to plants, and may cause a temporary painful inflammation on human skin, so far no
• Carvacrol, thymol and anethole content was increased by up to ten-fold of the original, wild-grown plants in four selected plant species, using various selection and conventional breeding techniques.
• anethole, carbacrol, and/or thymol would be expected to have more potent essential oil than
• the essential oil extracts can be used against plant diseases caused by a broad spectrum of fungi, bacteria and nematodes as well as insects, when applied in varying concentrations mixed with other oils, in vapor form, or sprayed as part of an aqueous emulsion in water as well
• the extracts can be applied to soil when embedded in
• perlite in granular form, powder form, or emulsified in water and applied via drip-watering systems.
• the extracts can also be used together with solarization when applied in vapor or any other form as well as by pouring or spraying aqueous emulsions around the growing area of plants in formulations indicated below.
• fungicidal in granular form, powder form, or emulsified in water and applied via drip-watering systems.
• the extracts can also be used together with solarization when applied in vapor or any other form as well as by pouring or spraying aqueous emulsions around the growing area of plants in formulations indicated below.
• these extracts have beneficial side effects: when used as a soil and/or
• foliar spray and/or soil application they can increase the concentration of beneficial microorganisms in soils, such as fluorescent pseudomonads (Pseudomonas fluorescens), and have growth promotion effects on plants (i.e. increased germination rate, foliage production and
• Essential oils as extracted are not soluble in water and are phytotoxic in undiluted oil
• oils have to be formulated in granules or powders or absorbed in a carrier, such as perlite or vermiculite, for soil applications and have to be emulsified in water for soil and foliar applications.
• the essential oil may be mixed with a carrier substance, such as a
• porous substance Suitable porous carrier substances include perlite and vermiculite. For these applications, at least about 0.5g of essential oil is combined with about 10-50g of the carrier
• Solarization may be conducted by covering the treated soil area with a transparent, impervious covering such as
• Plants can be transplanted into the soil immediately or several days after application of the essential oil treatment.
• essential oil concentrations may be about 10-1000 ppm. More preferably, the concentration of essential oil is about 100-1000 ppm.
• concentrates are atomized and applied over plants and/or soil in a storage area or greenhouse, for example.
• concentration of about 100-1000 ppm in air of the storage area is suitably
• the extracts obtained from the plants indicated above are also active, and being
• ticks and other pests when applied in vapor or aerosol form, in dust or granular formulations, diluted in carrier oils, extracted with solvents which dissolve essential oils or sprayed as aqueous
• Essential oil extracts also can be used, when applied as a vapor or in other forms, to
• oils poured on the soil around the plants via irrigation or pouring by hand), or directly sprayed
• Soil chemical fumigants and other treatments do not specifically target particular pests or pathogens. Therefore, they have negative effects on whole soil microfloral and microfaunal communities, including plant- and soil-beneficial organisms.
• the essential oils described in this application appear to work selectively. Detrimental organisms are targeted while
• organisms may influence plant disease resistance and growth either directly, or indirectly through
• antimicrobial activity to inhibit the growth of soil phytopathogens, either through the induction of plant defense responses, or through suppression of the pathogens via competition.
• Essential oils can easily be separated from water using a separatory funnel. Under commercial factory conditions, about 1 kg of dried plant
• material or seed is generally mixed with about 10 liter of water or extracted in plant material with
• Water and plant content may vary according to distillation technique.
• the plants were emulsified in water.
• a 1000 ppm emulsion about 1 mL essential oil containing extract(s) from one or several plant species was mixed with various concentrations
• Tween 20 or other commercial detergents (the optimal concentration is about 1 mL essential oil extract(s) dissolved in about 1 mL Tween 20) and added to about 1 L water.
• the water is acidic. Therefore, about 1 drop of concentrated hydrochloric acid per L of water was used to bring down the pH of the water to approximately 5.0.
• Soil tests Soil samples were obtained from infected fields in Kumluca, Antalya,
• Essential oils extracted from T. spicata and various mixtures of extracts were added in to the soil in an aqueous emulsion form by adding the emulsion to the irrigation water. Plots were covered
• Each container was inoculated with 2 mL zoospore suspension (1100 zoospores/mL) of P.
• the resulting concentrations of essential oil concentrations in the soil was 100, 200, 400 and/or 1,600 mg/kg (See Figure Legends).
• the containers were covered with airtight plastic film and incubated at 25°C for 5 days in a climate-controlled room. The film was then removed, and after 3 days of aeration, 15 pepper (Capsicum annum) seeds from a variety susceptible to P. capsici
• Bacteria (I x lO 5 CFU/mL) were incubated at 25°C
• NB nutrient broth
• test bacteria I x lO 5 CFU/mL were plated onto NA, and the plates
• the petri dishes were incubated for an additional 3 days before determining the amount of essential oil.
• MIC minimum inhibitory concentration
• the MIC was determined based on the equation of the regression analysis (Dimond et al., 1941).
• Essential oils extracted from plants can be emulsified in water and sprayed onto plants
• Essential oils can be used in vapor form, mixed with other oils, dissolved in solvents which dissolve essential oils, or in emulsion form to fog greenhouses or other buildings to kill
• Essential oils extracted from the plant species indicated above, alone or in combination, can be embedded into a carrier such as perlite or vermiculite as extracted in powder or granular
• essential oils were mixed with 10-50 g of perlite or vermiculite.
• the perlite or vermiculite was then sprinkled on the soil surface or mixed to a 5-10 cm depth into soil, using a commercial fertilizer applicator, to cover an area of one (1) square meter.
• the surface of the soil was then
• chemicals such as materials known to induce systemic disease resistance in plants, including
• Plants can be planted or transplanted into the soil immediately or several days after application according to
• Essential oils diluted in other oils, dissolved in solvents which dissolve essential oils, or
• fertilizers to irrigation water at about 10-1000 ppm concentrations, optimal concentrations generally range from about 100-200 ppm), and also directly to plants in drip- water irrigated
• First irrigation can be made before
• Example 3 Storage Applications: The essential oils extracted from the plants indicated above, and the vapors of these oils,
• Essential oils may also be used to kill storage pests and pathogens. Fogging, as indicated above, increases the activity of the oils due to a higher distribution rate to a larger area. Essential oils may also be used to kill storage pests and pathogens. Fogging, as indicated above, increases the activity of the oils due to a higher distribution rate to a larger area. Essential oils may also be used to kill storage pests and pathogens. Fogging, as indicated above, increases the activity of the oils due to a higher distribution rate to a larger area. Essential oils may also be used to kill storage pests and pathogens. Fogging, as indicated above, increases the activity of the oils due to a higher distribution rate to a larger area. Essential oils may also be used to kill storage pests and pathogens. Fogging, as indicated above, increases the activity of the oils due to a higher distribution rate to a larger area. Essential oils may also be used to kill storage pests and pathogens. Fogging, as indicated above, increases the activity of the oils due to a higher distribution rate to
• vapors i.e., from heated extracts or mixed in paint (preferably, an oil-based paint). Heating is not required for vaporization, however, heating improves vaporization. Bombs
• anethole is highly effective for this application and may be extracted from plants or is easily synthesized.
• Essential oils can also be used as mixed with liquid paraffin as in any form indicated above for protecting
• the test bacteria than contact with the essential oil.
• the essential oil in the volatile phase effect tests showed an MIC ranging from 41 ⁇ g/mL to 684 ⁇ g/mL (Table 4).
• insects i.e. ants, house flies, spiders, mites, fleas, mosquitoes, termites, ticks, etc. They also
• insects can be applied to skin in a cream or spray form to deter insects such as mosquitoes and ticks.
• oils to treat honeybee parasites are used in the form of aqueous emulsions, diluted in other oils,
• Rhizoctonia solani Sclerotinia sclerotium and Phytophthora capsici
• Thymbra spicata var. spicata showed the best activity against P. capsici, the agent of pepper blight, both in greenhouse and in field studies. So far there is no
• Phytophthora species are a major pathogen of peppers and many other crops in many areas of
• the essential oils reduced the population of beneficial soil fungi and bacteria up to 40%>, while
• Aqueous emulsion application may be repeated every 15 days to increase the protection level
• bacterial plant pathogens including: Erwinia amylovora, E. carotovora pv. carotovora,
• MIC Minimum inhibitory concentrations
• essential oils in media ranged from 200-400 ⁇ g/mL against all bacteria tested.
• MIC of the volatile phase of essential oils were ranged from 40-650 ⁇ g/mL of air, indicating that volatile
• insects when used at a 100-200 ⁇ l/L air concentration within 1-6 days.
• insecticides when used at a 100-200 ⁇ l/L air concentration within 1-6 days.
• Essential oils also have high activity in vapor form and/or
• Extracts of plant species naturally grown in Turkey are found to be potent anti-fungal
• Example 8 The Effect of Essential Oils from Origanum spp. against Xanthomortus axonopodis pv. vesicatoria
• bactericidal effect living bacteria were not detected at any point up to three days following the
• Essential oil also had a bactericidal effect at concentrations of 1,000
• Control suspensions reached stationary phase 24 hr after inoculation.
• the essential oil also demonstrated antibacterial activity at relatively low concentrations
• extracts emulsified in water as described above, at concentrations ranging from 100- 1000 ⁇ g/mL
• Plants were inoculated with a bacterial suspension (100 CFU/mL) and the
• the antibacterial activity can further be increased by using
• T. spicata and various mixtures of essential oils had higher activity compared to Origanum spp.
• Example 10 The Activity of Essential Oils from Thymbra spicata against Fire Blight
• Fire blight disease caused by Erwinia amylovora, of is one of the most damaging disease
• Example 11 Determination of Activity of Essential Oils from Thymbra spicata against Xanthomonas campestris pv. campestris in Cabbage
• Example 12 Activity of Essential Oils against Carmine Spider Mite (Tetranycus cinnabarinus) in Pepper Pepper plants were treated with 100, 200 and 500 ppm concentrations of essential oil
• Emulsions of essential oils have a very high contact activity, as well as a volatile phase activity, against small insects, i.e. Drosophila, spiders, mosquitoes, sugar ants and aphids.
• An emulsion containing about 100-1000 ppm essential oil will kill over 50%> of the sampled
• insects within 0.5-3 minutes after spraying are insects within 0.5-3 minutes after spraying.
• the antifungal and antibacterial activity of the essential oils is derived from their ability
• Essential oils can be used against Phytophthora fragaria and nematodes infesting
• T. confusum were reared on a mixture of wheat flour, bran and
• yeast E. kuehniella were reared on ground wheat, and S. oryzae were reared on wheat grains.
• Jraws-anethole (Sigma) used in the tests was of 99% purity.
• E. kuehniella E. kuehniella confusum and S. oryzae and larvae (13-16 days old) of E. kuehniella were exposed to anethole in small nylon gauze bags containing rearing food. Twenty insects were placed in each bag to make one replicate. Three replicates for each dose and exposure time combination were taken.
• a set of cloning plates consisted of a bottom plate with 60 microwells and a cover plate which had 60 holes drilled over the
• test chambers were 650 mL glass jars with screw-top lids. Anethole, diluted in
• acetone was applied on a blotting paper strip which measured 3 x 8 cm.
• the blotting paper was attached to the lower side of the jar's lid with adhesive tape.
• anethole/L air were required at varying exposure periods to achieve 100%> mortality in the adults of S. oryzae and T confusum, and the larvae of E. kuehniella, respectively.
• Anethole was also toxic to the eggs of T. confusum and E. kuehniella. A concentration
• Anethole apparently was more toxic than its parent compound, essential oil of anise, against the species tested. For instance,
• Tribolium castaneum (Herbst) and Sitophilus zeamais Motschulsky .
• Sitophilus zeamais Motschulsky Tribolium castaneum (Herbst) and Sitophilus zeamais Motschulsky .
• bromide recommended for treatment of various commodities are 25, 35, and 40 g/m 3 , respectively, at 24 hours (Anonymous,
• Anethole was reported to have contact toxicity against the eggs, larvae, and adults of J. confusum and the adults of S. zeamais and exhibited a repellent effect against the adults of J. confusum (Ho et al., 1997). Anethole was also shown to be toxic and totally inhibit the reproductive activity of a serious fruit pest, Ceratitus capitata Wied.(the
• Example 14 Mixtures of Essential Oils from T. spicata, P. anisum and F. vulgare Are
• a mixture of essential oils was prepared from in the following proportions: 40%> T. spicata, 10%> F. vulgare, 10%o O. ssp., 30% S. thymbra. The mixture was formulated in
• compositions comprising 10-50%> essential oils in olive oil.
• Application of the essential oil composition was effective in repelling mosquitoes for 3-4 hours. No bites were received in the time period tested. Olive oil controls were not effective in repelling mosquitoes. The 50%>
• composition was the most effective, but slight burning sensation was reported. A 30%>
• composition formulated in a cream was effective for repelling mosquitoes for 1.5 to 2 hours.
• This preservative may be replaced with a combination of 1 g/kg methyl paraben and 0.5g/kg
• Karapinar, M. The effects of citrus oils and some spices on growth and aflatoxin production by Aspergillus parasiticus NRRL 2999. Int. J. Fd. Microbiol. 2, 239-245, 1985.
• Kivanc, M., A. Akgul Mould growth on black table olives and prevention by sorbic acid, methyl-eugenol and spice essential oil.
• Academic 34 369-373, 1990
• Tripathi S. C, S. P. Singh and S. Dube. 1986. Studies on antifungal properties of essential oil of Trachyspermum ammi (L.) Sprague. J. Phytopath. 116, 113-120.

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