Location: Natural Products Utilization Research
2016 Annual Report
Objectives
Objective 1: Discover new insecticidal active ingredients that are based on natural products and that are safe with respect to occupational exposure and with respect to the environment, including non-target effects.
Subobjective 1.1: Discover novel arthropod repelling biopesticides from mosquito repelling folk remedies.
Subobjective 1.2: Discovery of fungal compounds and natural product analogs with activity against permethrin-resistant and susceptible mosquitoes.
Subobjective 1.3: Discover novel bioinsecticide active ingredients from crude plant and fungal extract screening and bioassay-directed fractionation.
Objective 2: Participate in development of new products based on green biopesticides.
Subobjective 2.1: Develop and optimize fatty acids as natural topical and spatial arthropod repellents.
Subobjective 2.2: Development and optimization of the chromene derivative 131-1 as a biopesticide.
Approach
A bioassay-directed investigative approach will be performed on bioactive extracts in efforts to discover bioinsecticides and repellents for use against arthropod pests. General procedures for isolation of active compounds from plants and microbes will be utilized. Solvent extractions, liquid-liquid partitioning, column chromatography and thin layer chromatography will be employed as needed. Extracts, fractions and pure compounds will be tested for insecticidal and insect repellent activity in assays being carried out routinely at collaborator's laboratories. Identification of active compounds will be done using spectroscopic methods including mass spectroscopy (EI, CI, and high resolution ESI), Nuclear magnetic resonance (one- and two-dimensional). Chemical modification will be performed on compound(s) identified as "highly active" to improve activity or bioavailability.
Progress Report
This is the first full year of research for Project No. 6060-32000-002-00D (Apr 1, 2015 - March 31, 2020) since it was newly created and approved. For objective 1, discovery of new insecticidal active ingredients that are based on natural products, milestones have been met and significant progress has been made. A plant commonly known as sweetgrass, which has documented use as an insect repellent by the Flatheads of Montana and Blackfoot of Alberta, has been evaluated for its insect repelling constituents. Phytol and coumarin were both determined to be responsible for the Aedes aegypti biting deterrency. Scientific evidence validates its traditional use as a biting insect deterrent. Options for development of constituents as mosquito control products are being explored.
The culture filtrate of a plant pathogenic fungus that infected Zinnia elegans and Hydrangea macrophylla was investigated for mosquitocidal constituents by bioassay guided isolation. The fungus responsible for the pathogenic effects on Zinnia elegans and Hydrangea macrophylla plants had been identified as Nigrospora spherica by molecular techniques. The mosquito adulticidal constituent in the culture filtrate was identified as phomalactone by spectroscopic techniques. Laboratory bioassays showed that phomalactone had larvicidal activity against permethrin susceptible and resistant Aedes (Ae.) aegypti larvae and topical adulticide activities on permethrin susceptible and resistant Aedes aegypti and Anopheles quadrimaculatus mosquitoes. Phomalactone was effective as a topical adulticide against the standard Orlando reference strain of Ae. aegypti. Activity against An. quadrimaculatus was also found.
Ammi visnaga seeds (Apiaceae) were extracted and analyzed for pesticidal activities. Two compounds visnagin and khellin were isolated that showed phytotoxic activity as well as mosquito larvicide activity. The phytotoxic activity of these constituents, visnagin and khellin, has been patented as part of a separate NP 306 research project in which both scientists on this project are also involved and co-inventors.
For objective 2, participate in development of new products based on green biopesticides, milestones have been met and significant progress has been made. Progress towards the development and optimization of 131-1 as a biopesticides has been made. We have attempted a series of synthetic routes to optimize the yield of 131-1. Our previous synthetic scheme was shown to be tedious and not very safe in our laboratory. We have now developed a synthetic scheme that is more efficient, convenient and most importantly produces the isomer of interest in greater than 80% yield. This method was safe, efficient and we could make the cyclized chromene aldehyde in the scale of 200 g at a time. Thus far we have produced 3.5 kilograms of the aldehyde analog which will be purified by an outside company as we do not have the chromatographic facilities to handle such large scale purification. The purified 131-1 will be sent to inhalation toxicity studies as part of the six-pack toxicology package needed for EPA registration and enhancement of private sector interest.
Accomplishments
1. Hierochloë odorata (L.) P. Beauv. (Poaceae), a plant commonly known as sweetgrass, has documented use as an insect repellent by the Flatheads of Montana and Blackfoot of Alberta. Both the Flatheads of Montana and Blackfoot of Alberta would use braided plant material in a sachet in clothing or burn them from one end as incense, air/clothing freshener, and insect repellent. This study evaluated the insect repellent properties of this plant using an in vitro mosquito feeding bioassay-directed approach to identify the compound(s) responsible for the observed activities. Evaluation of crude extracts produced from H. odorata revealed that the hydrodistillate had the highest level of mosquito biting deterrence. Fractionation of this extract followed by re-evaluation for mosquito biting deterrence produced many active fractions which were evaluated by spectroscopic techniques and determined to contain phytol, coumarin, and 2-methoxy-4-vinylphenol. Phytol and coumarin were both determined to be responsible for the Aedes aegypti biting deterrency. Scientific evidence reported here validates its traditional use as a biting insect deterrent.
2. Mosquitoes play a major role as vectors that transmit parasitic and viral diseases worldwide, especially in tropical and subtropical countries. Mosquito borne diseases not only affect humans but they also affect livestock in many parts of the world. They carry diseases that are lethal to dogs and horses. Dog heartworm disease (Dirofilaria immitis) is a parasitic disease spread through mosquitoes. This disease is not limited to dogs, but it can affect other animals and humans as well. Eastern equine encephalitis (EEE) and West Nile virus (WNV) are also mosquito borne diseases that affect the central nervous system of horses and cause severe complications and death. Emergence of resistance among mosquitoes to current pesticides has increased the importance of the search for alternate compounds that are effective and environmentally benign with diverse modes of actions than those that are commercially available. Aedes aegypti mosquitoes are the primary vector for transmission of Zika viral fever, yellow fever, dengue fever, and chikungunya. Mosquito control is currently the best strategy to prevent mosquito borne diseases. There are numerous approaches for control of potentially dangerous mosquito populations. These approaches include the use of adulticides (insecticides), larvicides, and, to a limited extent, the use of repellents. Our previous studies have shown the mosquito repellent activity of chromenes. We demonstrated larvicidal and adulticidal activity of chroman and chromene analogues against a permethrin susceptible laboratory strain as well as activity against a permethrin-resistant strain of Aedes aegypti.
None.
Review Publications
Xie, Q., Li, S.-X., Liao, D.-F., Wang, W., Tekwani, B., Huang, H.-Y., Ali, A., ur Rehman, J., Schrader, K.K, Duke, S.O., Cantrell, C.L., Wedge, D.E. 2015. Bio-pesticidal and antimicrobial coumarins from Angelica dahurica (Fisch. Ex Hoffm). Natural Product Communications. 2016;10(3):294-306.
Nakano, H., Seiji, K., Mamonov, L.K., Cantrell, C.L. 2016. 8-0-Acetyl-7-0-Methylgossypetin from Atraphaxis laetevirens. Chemistry of Natural Compounds. 52(1):127-129. doi: 10.1007/s10600-016-1567-x.
Bufalo, J., Cantrell, C.L., Jacob, M.R., Schrader, K.K., Tekwani, B.L., Kustova, T.S., Ali, A., Boaro, C.S.F. Antimicrobial and antileishmanial activities of diterpenoids isolated from the roots of Salvia deserta. Planta Medica. 2016;82:131-137.
Meepagala, K.M., Johnson, R.D., Techen, N., Wedge, D.E., Duke, S.O. 2015. Phomalactone from a phytopathogenic fungus infecting Zinnia elegans (Asteraceae) leaves. Journal of Chemical Ecology. 41:602-612.
Meepagala, K.M., Estep, A.S., Becnel, J.J. 2016. Larvicidal and adulticidal activity chroman and chromene analogues against susceptible and permethrin-resistant mosquito strains. Journal of Agricultural and Food Chemistry. 64(24):4914-4920.
