Location: Natural Products Utilization Research2022 Annual Report
1. Develop biopesticides to improve control of arthropods. 1.1. Discover novel bioinsecticide active ingredients from crude plant extract screening and bioassay-directed fractionation. 1.2. Discovery of fungal compounds with activity against permethrin-resistant and susceptible mosquitoes. 2. Develop biorepellents to improve control of arthropods. 2.1. Discover novel arthropod repelling biopesticides from mosquito repelling folk remedies and plant essential oils. 2.2. Development and optimization of the chromene derivative 131-1 as a biopesticide. 3. Develop natural product synthetic analogs for management and control of arthropods. 3.1. Use synthetic methods to develop and optimize fatty acids as natural topical and spatial arthropod repellents. 3.2. 131-1 Development into a commercial mosquito repelling product.
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.
For Objective 1, develop biopesticides to improve control of arthropods, milestones have been met and significant progress has been made. An infected mosquito from the genus Aedes, such as Aedes aegypti or Aedes albopictus has become the world's most influential factor for morbidity and death each year. This study describes the isolation and identification of six secondary metabolites from a soil-derived isolate of Streptomyces distallicus using the GNPS (Global Natural Product Social Molecular Networking) platform, ToF-DART HRMS exact mass determinations, and spectroscopic characterization via bioassay-guided isolation approaches. In addition, these metabolites were evaluated for Ae. aegypti mosquito larvicidal activity and antifungal activity against Colletotricum sp. Two isomeric compounds, aureothin and allo-aureothin, showed larvicidal activity against Ae. aegypti with LC50 values of 1.5 and 3.1 ppm for 24 h post-treatment, respectively, and 3.8 and 7.4 ppm for 48 h post-treatment, respectively. The crude extract of S. distallicus demonstrated potent larvicidal activity with LC50 values of 1.46 and 1.2 ppm for 24 and 48 h post-treatment, respectively. Deoxyaureothin, a furan ring reduced form of aureothin, showed no activity against Ae. aegypti. Thus, the study discovered that S. distallicus and crude extracts thereof are promising sources of potential microbial larvicides. A review article was published as a result of the congressional funds that pass through NPURU to the University of Mississippi. Briefly, natural products (NPs) have a long history as sources of compounds for crop protection. Perhaps a more important role for NPs has been as models and inspiration for the discovery and development of synthetic crop protection compounds. NPs and their synthetic mimics account for 18% of all crop protection compounds, whereas another 38% of all crop protection compounds have a NP that could have served as a model. Because NPs are often complex molecules, have limited availability, or possess structural features that constrain their suitability for use in agricultural settings, a key element in NP-inspired compounds is the simplification of the NP structure to provide a synthetically accessible molecule that possesses the physicochemical properties needed for use in crop protection. We reviewed a series of examples of NP mimics that demonstrate the structural or synthetic simplification of NPs as a guide for the discovery of future NP-inspired agrochemicals focused on fungicides, herbicides, and insecticides. The genus Salvia includes around 900 species distributed throughout the world and is the largest genus in the economically and medicinally important plants of the Lamiaceae family. The secondary compounds of Salvia species have been extensively studied for numerous biological activities. The genus Salvia contains several allelopathic species and extracts from three of these species, Salvia miltiorrhiza, S. hispanica and S. columbariae, that were tested in this study for phytotoxicity. Although there are several reports of phytotoxins from Salvia, there are no reports of phytotoxins against higher plants from Salvia miltiorrhiza. The abietane diterpenes tanshinone I and tanshinone IIA were identified as the compounds responsible for phytotoxicity. Neither compound was toxic to mosquito (Aedes aegypti) at 125 ppm (ca. 0.435 mM). For Objective 2, develop biorepellents to improve control of arthropods., milestones have been met and significant progress has been made. Curcuma longa L. belongs to family Zingiberaceae produced in warm wet regions of the world. Commonly known as turmeric, C. longa is commonly used in Indian folk medicines. Turmeric is especially popular because of its attractive culinary, cosmetic, and medicinal uses. This is an essential spice used in many dishes for good color and aroma. Turmeric contains odoriferous oils and the rhizomes have been reported to possess biological actives including the toxicity and repellency against Ae. aegypti. Essential oils of Curcuma longa rhizome and leaf were evaluated for their repellency and toxicity against imported fire ants. The samples of essential oils used in this study were reported to have ar-turmerone constituents of 36.9%, and 24.9 % respectively. Ar-turmerone was the major compound of the rhizome oil (36.9%) and leaf oil (24.9%) essential oils. Essential oils of C. longa rhizome and leaf showed significantly higher repellency than DEET. Both the essential oils tested in this study showed very high repellency and appears to have a potential for field use. Whiteflies are a common arthropod pest in vegetable crops, and they attack horticultural plants in both fields and the environment. This research focused on a wild watermelon species, Citrullus ecirrhosus, that were offered resistance against the Bemisia tabaci MEAM1. The isolation was performed by liquid extraction, hydrodistillation and/or steam distillation from frozen Citrullus ecirrhosus, fresh Citrullus ecirrhosus plant material, and/or fruit. The biological evaluation was delayed multiple times as samples have been provided to an ARS researcher in Charleston, South Carolina. For Objective 3, develop natural product synthetic analogs for management and control of arthropods, milestones have been met and significant progress has been made in the previous year to meet this objectives milestone. There is a need for the development of natural product-based insect repellents and biting deterrents to manage important disease vectors such as Aedes aegypti, which transmits viral pathogens to humans, including yellow fever and dengue, both of which can cause severe human morbidity and mortality. Prior work by our group has focused on investigations of traditional insect repelling remedies used by indigenous groups as a source of natural product-based chemical solutions for managing disease vectors. These investigations lead to successful research on the Indian folk remedy plant, Jatropha curcas, as well as an investigation of breadfruit, Artocarpus altilis, which is used throughout Oceania. The usage of these plants in controlling biting mosquitoes was attributed to the presence of fatty acids and to a smaller extent to the presence of triglycerides. We report here an extension of our research and understanding of the role triglycerides may play in biting deterrency. Analogs of triglycerides are being investigated to understand their role in biting deterrency as well as repellency of both mosquitoes and fire ants. In search of biopesticides we have explored plants belonging to Rutaceae family. We have been very successful in finding highly active mosquitocidal compounds and their synthetic analogs. During this year we have been working on various plant parts (roots, leaves and fruits) of Poncirus trifoliata a member in the Rutaceae family also known as hardy orange or trifoliate orange. We found that the ethyl acetate and hexane extracts showed larvicidal and topical mosquitocidal activities but the extracts were not discriminant between insecticide resistant and susceptible mosquitoes on adulticide activity. When tested for larvicide activity, the ethyl acetate extract and the hexane extracts showed higher activities towards the insecticide resistant Orlado strain Ae. Aegypti larvae. Bioassay guided fractionation led to the isolation and identification of four pyanocoumarins Xanthoxyleti, dentatin and nordentatin and ponfolin, We have also carried out isovalaryl, acetate and benzoate analogs of nordentatin. Several of these analogs have been reported to possess anticancer activity in the literature. At present these isolated and synthesized derivatives are being tested on dose dependent larvicide assay at USDA, ARS Gainesville laboratory. We have been investigating phytopathogens as sources of biopesticides. We were able to isolate, identify and evaluate phytopathogens belonging to various genera such as Phoma, Diaporthe,Curvularia, Didymella, Pantoea, and Ascochyta. We successfully isolated and synthesized analogs of active principles from some of these pathogens belonging to isochromenes, macrolides and lactones. We isolated a fungus growing in Ipomoea aquatica plants in Oxford, Mississippi. The ethyl acetate extract of PDB (Potato dextrose broth) culture broth has showed larvicidal activity. We are carrying out large scale growth of this fungus and bioassay guided isolation of active metabolites. In our previous work on plants belonging to the Rutaceae family, we were able to isolate and identify a chromene amide. Based on this compound we were able to synthesize potent chromene derivatives that have higher activity and three times longer duration of protection than DEET against mosquito bites. These analogs have now shown activity against Fall army worms, and we are synthesizing various analogs to be tested against fire ants and house flies. Plumbagin is a naphthoquinone derivative that is well known for its biological activity as an insecticide. It is also commonly found in natural sources, including the plant genus Plumbago, the carnivorous plant genera Drosera, and Nepenthes. This study is a synthesis of naphthoquinone derivatives and evaluates their larvicidal activity. In addition, molecular docking studies performed with two Ae. aegypti proteins, an odor-binding protein, and a saliva protein, show that the novel naphthoquinone derivatives possess a reliable binding affinity. Twenty different naphthoquinone derivatives have been synthesized, and the potential biological effects of these compounds are being evaluated.
1. Streptomyces distallicus as a potent microbial biolarvicide. Mosquitoes are widely prevalent species of vectors capable of spreading diseases such as malaria, dengue fever, filariasis, yellow fever, etc. Mosquitos also affect humans by causing localized angioedema as well as skin and systemic allergic reactions. A relatively new threat, the Zika virus, is transmitted to humans via an infected mosquito from the genus Aedes, such as Ae. aegypti or Ae. albopictus. Consequently, infected mosquitos from the genus Aedes have become one of the world's most influential contributors to human morbidity and death. Microbial insecticides are currently a principal portion of the biopesticide industry. To explore new biopesticides with activity against Ae. aegypti, Streptomyces distallicus was investigated by ARS researchers in Oxford, Mississippi. The present study evaluated S. distallicus for the discovery of larvicidal compounds against Ae. aegypti for the first time. We reported the isolation and characterization of six compounds from S. distallicus, as well as their larvicidal activity against Ae. aegypti. Thus, the study discovered that S. distallicus and crude extracts thereof are promising sources of potential microbial larvicides.
Mccomic, S.E., Meepagala, K.M., Swale, D. 2021. Characterization of toxicological and neurophysiological effects of natural product based chromenes to Fall Armyworm, Spodoptera frugiperda. Journal of Economic Entomology. https://doi.org/10.1093/jee/toab172.
Da Cruz-Silva, C.T., Cantrell, C.L., Nobrega, L.P., Ali, A., Duke, S. 2021. Bioassay-guided isolation of phytotoxins from three salvia species. Allelopathy Journal. 54(1):13-24. https://doi.org/10.26651/allelo.j/2021-54-1-1344.
Sparks, T.C., Duke, S.O. 2021. Structure simplification of natural products as a lead generation approach in crop protection compound discovery. Journal of Agricultural and Food Chemistry. 69(30):8324-8346. https://doi.org/10.1021/acs.jafc.1c02616.