Composition with pheromonal activity and its use in the control of deanonaceous fruit borers

Abstract

composition with pheromonal activity and its use in the control of the fruit borer of annonaceae the present invention refers to formulations with sexual pheromone activity, effective in the control of the fruit borer (cerconota anonella) of annonaceae (pine cone, soursop and atemoya), considered one of the most serious, due to the significant damage it causes to the culture. in a first embodiment, the composition of said formulation comprises: (a) the combination of methyl (z)-9-octadecenoate, (z)-9-octadecenal, (z)-9-octadecenol and, (b) a carrier agriculturally acceptable. the ratio between such components in the compositions of the invention is preferably 1:3:5, respectively. the present invention also relates to the method of controlling this borer using the formulation of this invention. such application covers the agriculture sector and can act in different regions where the culture of these fruits is being disseminated.

Description

Descriptive report for a patent of invention: COMPOSITION WITH PHEROMONAL ACTIVITY AND ITS USE IN THE CONTROL OF THE FRUIT BORER OF ANNONACEAE PLANTS Field of Invention

[001] The present invention relates to formulations with sex pheromone activity, effective in controlling the fruit borer (Cerconota anonella) of Annonaceae (sugar apple, soursop and atemoya), considered one of the most serious pests due to the significant damage it causes to the crop. In a first embodiment, the composition of said formulation comprises: (a) a combination of methyl (Z)-9-octadecanoate, (Z)-9-octadecenal, (Z)-9-octadecenol and, (b) an agriculturally acceptable carrier. The ratio between these components in the compositions of the invention is preferably 1:3:5, respectively. The present invention also relates to a method of controlling this borer through the use of the formulation of this invention. This application covers the agricultural sector and can be applied in different regions where the cultivation of these fruits is being disseminated.

[002] The present invention proposes to solve the problem faced by Annonaceae producers with regard to the attack by the pest in question, which has been causing serious damage to the commercial production of these fruits. Background of the Invention and Prior Art

[003] Several phytosanitary problems, especially pests, have discouraged the establishment of commercial plantations of Annonaceae (sugar apple, soursop, and atemoya), with the fruit borer, Cerconota anonella, standing out as one of the most serious due to the significant damage it causes to the crop. When attacked by this insect, the fruits exhibit pulp rot, and the affected part becomes hardened and blackened, reducing their commercial value or even rendering them unsuitable for sale or processing. Petition 870220103879, dated 09 / 11 / 2022, page 7 / 26 2 / 17 industrial. There are estimates that in each harvest the loss of commercial soursop pulp can reach percentages of up to 100%, due to the mass attack of this pest.

[004] From the point of view of the importance of Annonaceae cultivation, it is worth noting that it is not only the quantity of food produced that matters, but the quality of the harvested product is also important. The biological quality of food is reflected in its nutrients, and two important aspects must be considered by agriculture: the production of healthy plants and the obtaining of food without chemical agents that are potentially harmful to humans and the environment. Thus, losses caused by various diseases and a complex of pests on Annonaceae directly influence their marketing and production (ARAÚJO, JF 2007. Organomineral fertilization and liquid biofertilization in the production of sugar apple fruits (Annona squamosa L.) in the lower-middle São Francisco. Dissertation (Doctorate in Agronomy) - Department of Agronomic Sciences, São Paulo State University, Botucatu, Brazil, 115p.).Furthermore, in controlling such pests, the use of pesticides tends to be increasingly reduced due to the environmental impact that these substances have already caused and continue to cause. Restrictions on their use have been expanded in all countries, especially since the end of the last century.

[005] The fruit borer C. anonella can be found in Latin American countries, mainly in Venezuela. In Brazil it is quite widespread, having already been found in the states of São Paulo, Amazonas, Pará, Pernambuco, Bahia, Ceará, Mato Grosso, Rio de Janeiro, Alagoas and the Federal District (BRAGA-SOBRINHO, R. 2010. Potential for exploitation of annonaceae in northeastern Brazil. Embrapa Agroindústria Tropical, Fortaleza, Brazil, 18p.).

[006] To date, studies conducted on this pest have been limited to describing the biology and reproductive behavior of this species. Petition 870220103879, dated 09 / 11 / 2022, page 8 / 263 / 17 (SILVA, EL, CARVALHO, CM, DO NASCIMENTO, RR, MENDONÇA, AL, SILVA, CE, GOLÇALVES, GB, FREITAS, MRT, SANTANA, AEG 2006. Reproductive behavior of the Annona fruit borer Cerconota anonella. Ethology, 112:971-976). Analogously to other lepidopteran species, communication between males and females of C. anonella is mediated by pheromones, and the perception of these signals by males triggers a series of behaviors expressed in a standard sequence. This sequence includes antennal movement, walking, wing vibration, takeoff, zigzag flight, locating the trail, flying against the wind following the pheromone trail, locating the pheromone source, landing on that source or in adjacent areas, walking and wing movement until finding the female. This is the long-distance response chain.The encounter with the female initiates another chain of short-distance responses, in which the male interacts with the female and may eventually lead to copulation (LIMA, ER, DELLA LUCIA, TMC Biodynamics of pheromones. 2001. In: Insect pheromones: Biology, chemistry and use in integrated pest management. VILELA, EF, DELLA LUCIA, TMC 2nd ed. Holos, Viçosa, Minas Gerais, Brazil, pp 13-25.

[007] The indiscriminate use of pesticides is still the most common method used by producers, which can pose risks to workers and consumers, as well as cause environmental damage. One practiced alternative is bagging the fruits while they are still small, in order to prevent pest attacks. This system is laborious and unproductive. Therefore, it is necessary to search for new methods to control pests, such as the use of pheromones, the subject of this invention.

[008] In general, the study of pest insect pheromones is primarily motivated by the economic losses caused by insects, which are usually accompanied by environmental and public health damage caused by the use of chemical pesticides. Therefore, this research fits within the principle of Integrated Pest Management (IPM), which can be defined Petition 870220103879, dated 09 / 11 / 2022, page 9 / 26 4 / 17 as a pest management system that links the environment and the population dynamics of the species, using all appropriate techniques and methods in a way that is as compatible as possible, keeping the pest population at levels below those capable of causing economic damage (WAQUIL, JM; VIANA, PA; CRUZ, I. 2006. Corn Cultivation. Production System. 2nd ed. Embrapa).

[009] In Brazil, up to the year 2009, 28 pheromones were registered for the control of insect pests in agriculture by the National Health Surveillance Agency (ANVISA, National Health Surveillance Agency. 2009. http: / / www.cnpma.embrapa.br / down_site / forum / 2009 / palestras / 2_ANVISA_Carlos_AIexandre.pdf accessed on 12 / 02 / 2014). These pheromones are used in monitoring, mass trapping and sexual confusion of pests. More than 90% of commercial pheromone-based products are used in monitoring, which consists of capturing males and / or females to detect population outbreaks, the time of emergence of adult insects, and to detect the presence, distribution, and abundance of insects (IBARRA, RTLB 2006. Monitoring of Leucoptera coffeella with sex pheromone traps. Doctoral thesis. Federal University of Viçosa. VIÇOSA, MINAS GERAIS).

[010] The use of pheromones in integrated pest management for crops of significant economic value has become widespread, especially in recent times. Thus, the study, characterization, and development of synthesis routes for the components of these pheromones have been substantially intensified.

[011] Several research groups have been working on identifying the components of pheromones and, to date, information on these components is available in several databases. For example, “The Pherobase” (available on the Web at http: / / www.pherobase.net / ) presents pheromones of insects belonging to the Oecophoridae family, to which the fruit borer of Annonaceae belongs, and have been elucidated to date. Although there are already Petition 870220103879, dated 09 / 11 / 2022, page 10 / 26 5 / 17 various pheromones identified and their composition elucidated, there is no reference to the pheromone of the species Cerconota anonella. There are also no patent applications to date for pheromonal formulations involving the fruit borer of Annonaceae, the subject of this patent. Some records relating to the Oecophopridae family can be found, but they refer to the application of gaseous fumigant pesticides such as sulfonyl fluoride (DE19633595C1; DE19651705C2) and its combination with an ester and carbon dioxide (DE19813894A1).

[012] In Brazil, there are already studies that demonstrate the efficiency of using synthetic pheromones, such as Rincoforol®, an aggregation pheromone for the coconut palm weevil. With regard to the order Lepidoptera, more specifically weevils, it is worth highlighting the pheromone of the weevil Diatraea flavipennella, recently discovered in Brazil, whose formulation has a patent application under number PI 0903698-9, developed by the chemical ecology group responsible for this product. This formulation has high efficiency under field conditions (Kalinová et al. B. Kalinova, RR Do Nascimento, M. Hoskovec, AL Mendonca, EL Silva, MRT De Freitas, CR Cabral-Jr, CE Silva, AEG Sant'Ana & A. Svatos. Identification of two components of the female sex pheromone of the sugarcane-borer Diatraea flavipennella (Lepidoptera: Crambidae. J. Appl. Entomol. 2012, Vol. 136, pp.203-211) when used in traps to capture males with the aim of reducing the species' population to levels below the economic damage threshold.

[013] Thus, the use of pheromones in behavioral pest control has been seen as an interesting alternative insofar as its environmental impact is minimal when compared to other forms of pest control. This possibility has contributed greatly to the knowledge of the reproductive habits of insects that attack important crops. More Petition 870220103879, dated 09 / 11 / 2022, page 11 / 26 6 / 17 That being said, knowledge of the components of insect pheromones has been important for the control of various pests.

[014] Given the importance of pheromones in the biocontrol of insect pests, several proposals for pheromone synthesis have been reported in published works and patent documents. Therefore, it is possible to say that the main known pheromone components are commercially available, or there are known synthesis routes for obtaining them, a fact that makes them potential products that can be applied in the form of specific and agriculturally acceptable formulations.

[015] The availability of pheromone components has allowed the development of compositions with pheromonal activity, some of which are found on the market. In document US 4632829, a sex pheromone composition is disclosed as a bait for the corn borer (southwestern corn borer, Diatraea grandiosella (Dyar)) consisting of 16-26% Z9-hexadecenal, 56-85% Z11-hexadecenal and 6-10% Z13-octadecenal. US patent 6355236 describes a slow-release pheromonal composition for this same species, packaged in a plastic container, comprising a liquid mixture of: (i) a sex pheromone which is an aldehyde of 10 to 18 carbon atoms and (ii) an aliphatic derivative selected from the group consisting of an aliphatic acetate, which has 2 to 4 fewer carbon atoms than the pheromone, and an aliphatic carboxylate ester which has 1 to 4 fewer carbon atoms than the pheromone.

[016] The advantage of pheromone monitoring is the rapid detection of the insect even at low population densities, unlike other sampling methods. Pheromone traps detect adults that are emerging or emigrating, and this can be a warning sign for making control decisions (WALL, C. 1989. Monitoring and spray timing. In: Insect pheromones in plant protection. JUTSUM, AR & GORDON, RFS Wiley, Chichester, UK., pp 39-66). Petition 870220103879, dated 09 / 11 / 2022, page 12 / 26 Ί / \Ί Summary of the Invention

[017] The present invention relates to formulations with sex pheromone activity, effective in controlling the fruit borer of Annonaceae (Cerconota anonella), soursop, custard apple, and atemoya. This pest is considered one of the most serious, due to the significant damage it causes to the crop. In a first embodiment, the composition of said formulation comprises: (a) the combination of methyl (Z)-9-octadecanoate, (Z)-9-octadecenal, (Z)-9-octadecenol and, (b) an agriculturally acceptable carrier. The ratio between such components in the compositions of the invention is preferably 1:3:5. The present invention also relates to the method of controlling this borer through the use of the formulation of this invention. Such application covers the agricultural sector and can be used in different regions where the cultivation of these fruits is being disseminated.

[018] The present invention proposes to solve the problem faced by Annonaceae producers with regard to the attack by the pest in question, which has been causing serious damage to the commercial production of these fruits. Detailed Description of the Invention

[019] The inventors of the present composition have identified the compounds (Z)-9-octadecanoate, (Z)-9-octadecenal and (Z)-9-octadecenol as the components of the ternary pheromone mixture of the female Cerconota annonella (LEPIDOPTERA: OECOPHORIDAE), also known as the fruit borer of Annonaceae.

[020] In the studies that resulted in the composition of the invention, it was verified, in analyses of hexane extracts of virgin females of Cerconota anonella by two-dimensional Gas Chromatography coupled to Mass Spectrometry (GC x GC-TOF-MS), that the sex pheromone of this pest is constituted by a mixture of the three compounds mentioned above in the proportion of 1:3:5. Petition 870220103879, dated 09 / 11 / 2022, page 13 / 26 8 / 17

[021] Bioassays conducted in the laboratory demonstrated that the response of C. anonella males to this mixture is similar to that observed for virgin females and extracts of the pheromonal gland of virgin females. In other words, the mixture specified above is the sex pheromone of C. anonella. The use of this pheromone in the integrated management of this borer will enable its effective control, that is, the control of this pest in Annonaceae plantations, where this species has successfully established itself.

[022] The compositions of the invention may be in liquid, solid and semi-solid forms. In solid form, the composition may be in micronized form, in the form of granules, powder or any other suitable form. In liquid form, the compositions of the invention may be in the form of solutions, suspensions or gels.

[023] The compositions of the invention may include one or more additives to improve the stability of the composition, fillers, thickeners, solvents, binders. Examples of such additives include, but are not limited to, vegetable oils, such as olive oil, soybean oil, corn oil, sunflower oil, canola oil and combinations thereof. Examples of fillers include one or more mineral clays (e.g., attapulgite). Examples of solvents include ethyl alcohol, methyl alcohol, golden hydrocarbons, petroleum solvents and combinations thereof. Examples of binders include shellac gum, acrylic resins, epoxides, alkyds, polyurethanes, linseed oil and combinations thereof.Appetite stimulants may also be present in the compositions of the invention, stimulants which include, but are not limited to, cottonseed oil, phytol fatty acid esters, geranyl geraniol fatty acid esters, plant extracts, plant alcohols and combinations thereof.

[024] The compositions of the invention may contain one or more polymeric agents such as celluloses, proteins, casein, fluorocarbon-based polymers, hydrogenated resins, lignins, melamine, polyurethanes, vinyl polymers, such Petition 870220103879, dated 09 / 11 / 2022, p. 14 / 26 9 / 17 such as polyvinyl acetate (PVAC), polycarbonates, polyvinylidene dinitrile, polyamides, polyvinyl alcohol (PVA), polyamide aldehyde, polyvinyl aldehyde, polyesters, polyvinyl chloride (PVC), polyethylenes, polystyrenes, polyvinylidene, silicones, and combinations thereof. Examples of celluloses include, but are not limited to, methylcellulose, ethyl cellulose, cellulose acetate, cellulose acetate butyrate, cellulose acetate propionate, cellulose propionate, and combinations thereof.

[025] The compositions of the invention may also include one or more chemical insecticides such as cifluthrin, permethrin, cypermethrin, bifinthrin, fenvalerate, flucitrinate, azinfosmethyl, methyl parathion, malathion and others. The compositions of the invention may also include biological insecticides, such as natural toxins and pyrethrins, proteins from Bacillus thuringiensis and Beauveria bassiana.

[026] The compositions of the invention comprise the active and inert ingredients in an effective pheromonal quantity, that is, in a quantity sufficient to sexually attract males of the species C. anonella. Said effective pheromonal quantity corresponds to a ratio ranging from 1 to 10 (mg / mL) of active substances (pheromone components).

[027] The following examples refer to experiments for identification and characterization of the active components in the pheromonal mixture obtained from extracts of pheromone glands of Cerconota anonella. Example 1: Extraction from pheromone glands

[028] Ten females in call were placed in a freezer (-5°C) located in the Chemical Ecology Laboratory (IQB-UFAL) for 5 minutes, for immobilization purposes and to maintain the physiological conditions of the pheromone-producing gland. After this time, the females were removed from the freezer and, with the aid of a scalpel, the last abdominal segment of each female was cut and kept Petition 870220103879, dated 09 / 11 / 2022, page 15 / 26 10 / 17 in a glass container, where they remained in contact with 100 pL hexane for about 30 seconds. Throughout the entire extract preparation process, the container was kept immersed in ice to prevent solvent evaporation.

[029] C. anonella caterpillars were removed from fruits using entomological tweezers, transferred to transparent polystyrene (plastic) Petri dishes (90 x 15 mm), and kept on an artificial diet until pupae and adults were obtained for use in experiments: females were used to obtain extracts and bioassays, while males were used in bioassays employing extracts from females and synthetic compounds. The pupae were sexed, and the insects were kept in glass cages (14.8 cm x 30 cm x 30 cm) in a climate-controlled room with an inverted photoperiod of 12 hours of photophase and 12 hours of scotophase. During this period, cotton soaked in a sucrose solution (10%) was offered to the sex-separated insects as food, which was changed every two days. Example 2: Analysis of extracts by one-dimensional gas chromatography coupled to mass spectrometry (GC-MS) and two-dimensional gas chromatography coupled to a time-of-flight mass spectrometry detector (GC-GC-TOF / MS)

[030] For gas chromatography analyses, abdominal gland extracts from female C. anonella were concentrated to 10 pL (1 female-equivalent). 1 pL aliquots of the extracts were injected into a Gas Chromatograph with Flame Ionization Detector (GC / FID), QP2010 plus, using an RTX-5 column (30 m, 0.25 mm internal diameter; 0.25 µm film diameter; Restek) and nitrogen as carrier gas. The extracts were analyzed using a program with an initial oven temperature of 50°C and a heating rate of 9°C / min up to 240°C, with a "splitless" injection mode and injector and FID detector operating at temperatures of 200°C and 250°C, respectively. Using the results obtained from the GC-FID analyses as parameters, further analyses were performed. Petition 870220103879, dated 09 / 11 / 2022, page 16 / 26 11 / 17 in a Gas Chromatograph coupled to a Mass Spectrometer (GC / MS), QP2010 plus, with 70 eV electron impact ionization (Figure 16) using an RTX-5 column (30 m, 0.25 mm film diameter; Shimadzu) using Helium as carrier gas. The analyses of the gland extracts were performed with a program in which the initial oven temperature was 50°C with a heating rate of 9°C / min up to 240°C, "splitless" injection mode, with a DIC injector and detector at temperatures of 200°C and 250°C, respectively. The compounds present in the abdominal gland extract were identified by comparative study of mass spectra from the NISTO8 library and by calculating the retention index (Kovats index) using the database available at www.pherobase.com.To calculate the Kovats index, co-injection of the abdominal gland extract with a homologous series of n-alkanes (Sigma-Aldrich C7-C30) was performed, and to quantify it, 1 pL of 3-penten-2-one was added to the gland extraction samples as an internal standard, at a concentration of 0.17109 mg / mL.

[031] GC-GC / TOF / MS investigation of C. anonella extracts revealed a range of compounds, which were selected and tested in bioassays to verify biological activity. Table 1 describes the compounds identified in the abdominal extracts of female C. anonella that showed biological activity. Their identities were confirmed by injection of synthetic standards prepared in the laboratory. In addition to these, (Z)-9-Octadecanol and (Z)-9-octadecanal were also tested, as they are unsaturated derivatives of their unsaturated equivalents. Although present in the pheromonal mixture, the concentrations of these compounds are below the GC-MS detection limit. Thus, for the bioassays, their concentrations were established by taking the concentrations of their saturated analogues as a standard. Table 1: Chemical characterization of the abdominal gland extract of C. anonella. Retention time Compound Concentration (mg / mL) Petition 870220103879, dated 09 / 11 / 2022, page 17 / 26 12 / 1 / (minutes) 20305 Octadecanol 0.001593 20542 Octadecanal 0.014425 24706 Methyl (Z)-9-octadecenoate 0.006813 25246 Methyl octadecanoate 0.012527 28109 Octanoic acid 0.012491 Source: authors Example 3: Bioassays for determining the biological activity of compounds under laboratory conditions.

[032] The biological activity of the other identified components was tested in preliminary bioassays under the same conditions described here, but was not found to be attractive to C. anonella and was therefore disregarded. The bioassays were performed in a glass arena (14.8 cm x 30 cm x 20.3 cm) with a perforated lid, placed in a dark room under the following conditions: temperature of 21.46 ± 0.73°C; relative humidity of 84.00 ± 3.87% and a photoperiod of 12L:12D. Observations were made during the scotophase using a 15-watt red lamp. The identified compounds were synthesized in the laboratory and diluted in hexane at concentrations of 0.1 mg / mL and 1 mg / mL. Aliquots (10 pL) of each solution were applied to the filter paper disc (Whatman No. 01, φ20 mm). After the solvent evaporated, the discs were placed in the glass container.A virgin male (1-3 days old) was introduced into the arena and its behavior, as well as the time it took to reach the pheromone source, was recorded for 10 minutes.

[033] All experiments were conducted using 10 (ten) replicates per treatment and the following variables, based on the courtship behavior exhibited by males, were considered and quantified: (1) wing vibration, (2) short flight Petition 870220103879, dated 09 / 11 / 2022, page 18 / 26 13 / 17 and fast, (3) touch on the source and (4) copulation attempt. Bioassays with live virgin females hanging in a polyethylene cage served as a control treatment and hexane-impregnated paper discs, as well as clean discs, were also tested serving as control and blank treatments, respectively. Table 2 presents the comparison of the mean values ​​of each treatment tested in relation to the control treatment (live females). Table 2. Averages obtained in laboratory tests using the identified components of the Cerconota anonella pheromone. Treatments: Agitation, wings, direct flight, odor source, touch odor source. 1: OAL 0.1mg / mL 1.30BC* 0.20C 0.20AB 2: OM 0.1mg / mL 3.10BC 1.00BC 0.00B 3: OOI 0.1mg / mL 1.90BC 0.20C 0.00B 4: AO*** 0.1mg / mL 3.50BC 2.30ABC 0.40AB 5: OAL 1mg / mL 3.80BC 0.30C 0.00B 6: OM 1mg / mL 0.90BC 0.10C 0.00B 7: OOI 1mg / mL 6.50BC 0.10C 0.00B 8: OA 1mg / mL 3.90BC 1.10BC 0.1 OAB 9: Mixture 1r* 17.20AB 1.00BC 0.1 OAB 10: Mixture 2 18.40AB 2.70ABC 0.1 OAB 11: Virgin female 9.60a 3.1 OAB 0.40AB 12: White m 0.20C 0.00C 0.00C 13: Hexanolv 0.00C 0.00C 0.00C 14: Mixture 3V 19.60AB 4.90A 0.60AB 15: Mixture 4VI 11.40BC 0.60BC 0.00B 16: Mixture 5VH 5.40BC 0.90BC 0.90A 17: Mixture 6VI 6.60BC 0.40BC 0.00B Source: authors *Means with different letters differ from each other by Tukey's test at a 5% probability level (p<0.05). **I-Quaternary mixture of saturated compounds [1(OM):3 (OAL):5(Ool):5(OA)]-1mg / mL, II-Quaternary mixture of saturated compounds [1(OM):3 (OAL):5(OOI):5(OA)]-0.1mg / mL, III-Clean paper disc, IV-Hexane-impregnated paper disc, V-Ternary mixture of unsaturated compounds [1(OMI):3 (OALI):5(OOI)]-1mg / mL, VI-Ternary mixture of Petition 870220103879, dated 09 / 11 / 2022, page 19 / 26 14 / 17 unsaturated compounds [1(OMI):3 (OALI):5(OOII)]-0.1mg / mL, VII-Ternary mixture of saturated compounds [1(OM):3 (OAL):5(Ool):]-1mg / mL, VIII- Ternary mixture of saturated compounds [1(OM):3 (OAL):5(Ool):]-0.1mg / mL. **OM: Methyl octadecanoate, OALOctadecanal, AO: Octanoic acid, OMI: Methyl (Z)-9-octadecenoate, OALI: (Z)-9-octadecenal, OOII: (Z)-9-octadecenoL

[034] The data in Table 2 demonstrate that the chemical components, when tested individually, showed similar responses in male C. anonella, however, these responses are significantly lower when compared to the treatments of saturated and unsaturated mixtures and live females, for all variables tested (p<0.05). Regarding these latter treatments, the mixture of unsaturated compounds, as well as its proportion, was chosen based on the identification analyses of its saturated analogues. It is possible to verify that the biological activity of both mixtures (saturated ternary and unsaturated ternary) were those that elicited a more effective response for the studied variables related to the behaviors exhibited by the males, when compared to the female (control). The presence of octanoic acid inhibited the response of the males for all variables.Structurally, this compound is similar to biosynthetic precursors, consistent with literature reports that consider the absence of biosynthetic precursors in the composition of lepidopteran sex pheromones (SUBCHEV M. et al. 2000. 1-methylethyl octanoate, a new lepidopteran sex pheromone from the bagworm, Megalophanes viciella. J. Chem. Ecol., 26(2):487-495.; MILLAR, JG et al. 2008. (9Z)-9,13-Tetradecadien-11-ynal, the sex pheromone of the avocado seed moth, Stenoma catenifer. (6Z,9Z,12Z)-6,9,12-Octadecatriene and (3Z,6Z,9Z,12Z)-3,6,9,12-icosatetraene, the novel sex pyromones produced by moths). emeralds. Composition of the Diamondback Moth, Plutella xylostella (L.) in Korea J. Asia-Pacific Entomol. Therefore, it can be considered that a. Petition 870220103879, dated 09 / 11 / 2022, p. 20 / 26 15 / 17 The presence of OA may negatively interfere with the intensity of the response to the mixture of compounds that constitutes the sex pheromone of C. anonella. Regarding the concentrations tested, an increase in response was observed with an increase in the concentration of the mixture. Thus, the concentration of 1 mg / mL was considered more efficient in attracting males for all variables and treatments studied.

[035] Based on these results, it was possible to conclude that the mixtures containing the saturated and unsaturated compounds, without the presence of octanoic acid and at a concentration of 1 mg / mL, both in the proportions of 1:3:5, were as efficient as live females in attracting males, under laboratory conditions. Example 4: Bioassays for determining the biological activity of compounds under field conditions.

[036] In all field bioassays, rubber septa (0.7 cm⁰), which served as substrate, were impregnated with the test sample one day (1) before the field experiments were carried out. 0.2 mg of 2,6-di-tert-butyl-p-cresol (BHT) was added to the samples. For impregnation, the treatments were placed in contact with the rubber septum inside eppendorf-type tubes, which were closed and kept in a freezer (-5°C) until their use, when they were transported, in a thermal box, to the test areas.

[037] Bioassays were conducted in an Annona muricata orchard. The relative humidity was approximately 80% and the average temperature was 27 ± 0.3°C. An area of ​​1 ha was chosen, with a spacing of 4 m x 4 m between trees approximately 3 m high, and the traps were distributed in the middle third of each tree, equidistant at least 8 meters apart, in alternating rows and plants. Each trap contained 1 (one) cage (40 mm x 65 mm height) containing the rubber septum impregnated with the treatments. The volume of the impregnated sample was 500 µL, at a concentration of 1 mg / ml. Petition 870220103879, dated 09 / 11 / 2022, page 21 / 26 16 / 17

[038] Five (5) traps were distributed per treatment, for a total of forty-five (45), namely: (1) HPLC grade hexane (control); (2) Ternary mixture of saturated compounds [1 (methyl octadecanoate):3 (octadecanal):5 (octadecanol)] at a concentration of 1 mg / mL; (3) Ternary mixture of unsaturated compounds [1(Z-9-methyl octadecenoate):3 (Z-9-octadecanal):5(Z-9-octadecenol)] at a concentration of 1 mg / mL; (4) Live female (control); (5) Untreated trap (control).

[039] Thirty-five (35) repetitions were performed and the observed variable was the number of males captured per trap (NMC). Subsequently, an analysis of variance (ANOVA) was performed using Tukey's test at a 5% probability level (p<0.05) to verify significant differences between the analyzed factors and their interactions. Every seven (7) days, readings were taken of the number of males captured per treatment and the traps were re-randomized.

[040] Table 3 shows the averages obtained in the capture of males using the most efficient formulations in the laboratory, tested under field conditions. Table 3: Averages obtained in field trials using pheromonal mixtures of Cerconota anonella. Treatments Number of males captured per trap (Mean ± SE) Control (Hexane) 0.00 ± 0.00 aA Mixture 1* 6.30 ± 0.52 bB Mixture 2** 8.10 ± 0.52 cB Female 8.57 ± 0.52 cB Blank 0.0 ± 0.00 aA Source: authors Different lowercase letters between columns are statistically significant according to Tukey's test (p<0.05). Petition 870220103879, dated 09 / 11 / 2022, page 22 / 26 17 / 17 Different capitalizations between columns are statistically significant, in relation to the control, according to Dunnett's test (p<0.05). *Ternary mixture of saturated compounds [1 (methyl octadecanoate):3 (octadecanal):5 (octadecanol)] at a concentration of 1 mg / mL. ** Ternary mixture of unsaturated compounds [1 (Z-9-octadecenoate methyl):3 (Z-9-octadecanal):5(Z-9-octadecenol)] at a concentration of 1 mg / mL.

[041] Thus, it is possible to verify that despite exerting attractiveness, the mixture containing saturated compounds presents a lower average capture rate than that for the mixture of unsaturated compounds and live female. In these last two cases, the efficiency of the mixture of unsaturated compounds represented by the compounds (Z)-9-octadecenoate methyl, (Z)-9-octadecanal and (Z)-9-octadecenol in the proportion of 1:3:5, at a concentration of 1 mg / mL, is observed, which exerted an attractiveness equivalent to that represented by the live female, without a statistically significant difference (p<0.05).

Claims

1- Composition with pheromonal activity for the control of the fruit borer (Cerconota anonella) of Annonaceae characterized by comprising a combination of (a) methyl (Z)9-octadecanoate, (Z)-9-octadecenal, (Z)-9-octadecenol and (b) an agriculturally acceptable carrier. 2- Composition according to claim 1, characterized in that the ratio between the components in combination (a) of the invention is preferably 1:3:

5. 3- Composition according to claim 1, characterized by being in liquid, solid or semi-solid form. 4- Composition according to claim 1, characterized by comprising an effective pheromonal quantity corresponding to a ratio ranging from 1 to 10 (mg / mL) of active substances (pheromone components). 5- Composition according to claim 1, characterized by the solid form being micronized, in granules or in powder form. 6- Composition according to claim 1, characterized by the liquid form being a solution, suspension or gel. 7- Composition according to claim 1, characterized by containing additives to improve the stability of the composition, fillers, thickeners, solvents, binders, such as: vegetable oils, such as olive oil, soybean oil, corn oil, sunflower oil, canola oil and their combinations; fillers include one or more mineral clays (e.g., attapulgite). Examples of solvents include ethyl alcohol, methyl alcohol, chlorinated hydrocarbons, petroleum solvents and their combinations. Examples of binders include shellac gum, acrylic resins, epoxides, alkyds, polyurethanes, linseed oil and their combinations. 8- Composition according to claim 1, characterized by comprising phagostimulants that include, but are not limited to, cottonseed oil, phytol fatty acid esters, geranyl geraniol fatty acid esters, plant extracts, plant alcohols and combinations thereof. 9- Composition according to claim 1, characterized by comprising one or more polymeric agents such as celluloses, proteins, casein, fluorocarbon-based polymers, hydrogenated resins, lignins, melamine, polyurethanes, vinyl polymers such as polyvinyl acetate (PVAC), polycarbonates, polyvinylidene dinitrile, polyamides, polyvinyl alcohol (PVA), polyamide aldehyde, polyvinyl aldehyde, polyesters, polyvinyl chloride (PVC), polyethylenes, polystyrenes, polyvinylidene, silicones, and combinations thereof. 10- Composition according to claim 1, characterized by comprising one or more chemical insecticides such as ciflutrin, permethrin, cypermethrin, bifintrin, fenvalerate, flucitrinate, azinfosmethyl, methyl parathion, malathion and others. 11- Composition according to claim 1, characterized by comprising biological insecticides, such as natural toxins and pyrethrins, proteins from Bacillus thuringiensis and Beauveria bassiana. 12- Composition according to claims 1 to 11, characterized by being for the control of the fruit borer (Cerconota anonella) of Annonaceae. 13- A method for controlling the fruit borer (Cerconota anonella) of Annonaceae characterized by using the composition of claims 1 to 12 inserted in traps distributed in the plantations to capture males of this insect.