Research Project: Biobased Pesticide Discovery and Product Optimization and Enhancement from Medicinal and Aromatic Crops

Location: Natural Products Utilization Research

2024 Annual Report

Objectives
1. Discover bioactive pesticides (fungicidal and herbicidal) that enhance quality and marketability of small fruits and specialty crops. 1.1. Discover novel biofungicides and bioherbicides from crude extract screening and bioassay-directed isolation approaches. 1.2. Isolation and identification of antifungal metabolites from actinomycete fermentation for control of important fungal plant pathogens. 1.3. Investigation of plant pathogenic fungi for phytotoxic and antifungal activities and synthetic modification of the isolated compounds to gain insights into structure-activity relationships. 2. Determine plant growth and post-harvest processing conditions to enhance or optimize commercially important bioactive compound concentrations. 2.1. Target medicinal and aromatic plant growth and post-harvest processing conditions to enhance or optimize commercially important bioactive compound concentrations.

Approach
The long-term objectives of this project are to identify, chemically characterize, and develop new weed and plant pathogen managing biochemical biopesticides from natural sources and enhance or optimize commercially important bioactive compound concentrations. Plant and microbial extracts will be sourced using a variety of approaches and methods as evidenced through previous publications by the authors. These extracts will be subjected to in-house bioassays which include both antifungal and herbicidal based assays as described below. A bioassay-guided investigative approach will be performed on bioactive extracts in an effort to identify the specific natural compound(s) responsible for the activity of the extract(s). In-house high-throughput bioassays will be utilized to identify bioactive constituents with antifungal and herbicidal activities. A second component of this project pertains to natural products obtained from medicinal and aromatic crops which are low-volume high-value commodities that have numerous applications in various industries such as food, beverage, food supplement, flavor and fragrance, perfumery and cosmetics, pharmaceutical, aromatherapy, and various consumer products. Sourcing these and other natural compounds in an economical manner directly from plants is constantly evolving which creates an opportunity for small and mid-size farms.

Progress Report
This is the fourth year of research for this project. This report also includes progress made as part of the outgoing agreement with the National Center for Natural Products Research (NCNPR) at the University of Mississippi in Oxford, Mississippi. This will also be the second year to report on progress made as part of the new outgoing agreement with Marshall University. For objective 1, discover bioactive pesticides (fungicidal and herbicidal) that enhance quality and marketability of small fruits and specialty crops, milestones have been met and significant progress has been made. Anthracnose, caused by a fungal pathogen Colletotrichum fragariae is an economically important disease affecting strawberries globally, causing substantial losses in yield and fruit quality. The study was conducted to identify essential oils (EOs) with antifungal properties against C. fragariae and other plant pathogenic fungi. We identified 20 plant EOs with antifungal activity against anthracnose pathogens. Among these, patchouli EO (Pogostemon cablin) was further explored, and the major constituent’s patchouli alcohol exhibited antifungal activity against C. fragariae (isolate cf63) as well as other two pathogens C. gloeosporioides (another Colletotrichum species that cause anthracnose) and Botrytis cinerea (gray mold). Both the whole patchouli EOs and its major constituent, patchouli alcohol, can be used to manage anthracnose disease in strawberries. Another minor constituents of patchouli EO called pogostone was also identified and exhibited a strong antifungal activity against C. fragariae, C. gloeosporioides, and B. cinerea. Pogostone also exhibited a strong phytotoxicity and therefore can be used as a herbicide. The mode of action for both herbicidal and fungicidal is underway. In an effort to discover alternatives to chemical fungicides, our unit is exploring beyond small fruits crop to field crops like sugarbeet. Our unit was funded with a FY 2024 National Plant Disease Recovery System (NPDRS) proposal fund that was submitted to the office of National Programs (ONP). The proposal has been approved for funding in the amount of $25,730. This funding will support research aimed at identifying natural products (non-chemical methods) to control postharvest pathogens in sugarbeet, contributing to improved crop management and sustainability. Soybean is one of the most economically important crops worldwide. However, soybean yield can be substantially decreased by many diseases. Soybean genotypes could have different reactions to pathogen infection. As a first step toward investigating the biochemical basis of soybean resistance and susceptibility to disease, phytochemicals in the seeds of 52 soybean genotypes previously reported to have different reactions to diseases of soybean rust (SBR), Phomopsis seed decay (PSD), and purple seed stain (PSS) were analyzed. This is a collaborative project with Dr. Shuxiang Li from CGRU, ARS, USDA in Stoneville, Mississippi. For objective 2, determine plant growth and post-harvest processing conditions to enhance or optimize commercially important bioactive compound concentrations, milestones have been met and significant progress has been made. Yerba Santa (Eriodictyon californicum) stands as a testament to the rich botanical diversity and medicinal heritage of the western United States. Its leaves are a potentially rich source of natural flavonoids with possible medical value. In collaboration with NCNPR, University of Mississippi, we have isolated 20 novel compounds and many known compounds. The medicinal properties for prevention and treatment of inflammatory diseases and metabolic disorders, along with antifungal and phytotoxicity against plants are currently under investigation. Moreover, in collaboration with Scientific Ecological Services (a private company in Ontario, OR), we are leveraging chemical analysis and quality standards set forth by NPURU at ARS, USDA, to enhance flavonoid content through optimized cultivation practices. This partnership aims to identify superior varieties suitable for potential use in herbal supplements, thereby harnessing the full therapeutic potential of Yerba Santa. Beyond its culinary prowess, scientific research has increasingly recognized the potential of turmeric (Curcuma longa) and curcumin in promoting overall health and well-being. Through collaborative efforts with the NCNPR, University of Mississippi, Alabama A&M University, and industry partners (such as doTERRA International LLC), a unique cultivar of Curcuma has been identified. This cultivar has significantly higher levels of turmerones and curcuminoids, up to double the usual amounts. Furthermore, it contains a specific class of volatiles derived from Curcuma, which act as efflux inhibitors, enhancing the bioavailability of curcuminoids. Despite its significant nutritional and economic value, peanut (Arachis hypogaea L.) confronts challenges in post-harvest storage. Research indicates that storage induces the formation of deleterious oxidation products, impacting viability and growth, and posing health concerns. The oxidation products also substantially modify peanuts’ chemical composition, affecting flavor, shelf life, and nutritional value. However, the specific identities of oxidation products remain elusive, and the relationship between storage duration, oxidation product formation, and their effects on cultivation and nutritional quality is not well understood. Collaborating with an ARS researcher in Griffin, Georgia, our investigation unveiled a direct correlation between the storage duration of peanuts and the accumulation of oxidation products. In addition, the oxidation products have also been identified. University of Mississippi Non-Assisted Cooperative Agreement: Eight hundred eight plant samples were added to our plant repository this year – one of which was a sample identified and collected in Oxford, Mississippi, and over 2,500 natural product crude extracts, semi-purified fractions, and purified compounds were screened for biological activities against specific molecular targets and whole-cell systems. As part of our continuing effort in the search for anti-infective, cancer chemopreventive, and immunomodulatory/anti-inflammatory leads from natural sources, more than 40 compounds were isolated from 11 extracts, with three showing promising activity against drug-resistant Enterococci, Streptococci, and Staphylococci strains and two exhibiting activities against Plasmodium falciparum, sans cytotoxicity to mammalian cells. Also, 16 compounds were tested against a panel of transcription factors involved in cancer signaling pathways, of which 4 had potent anticancer activity. Characterization and development of immunomodulatory natural products. A major focus of this research program is the continued evaluation of the botanical Limnospira/Arthrospira (common name spirulina) for enhancing host immune resilience to respiratory viral infection. Ongoing efforts support the development of Immulina®, a patented Arthrospira extract that was previously discovered at the NCNPR (commercially available as a dietary supplement). An awarded NIH Botanical Dietary Supplements Research Center grant (2020-2025, umbdsrc.org) and Administrative Supplement has allowed the formation of a multi-disciplinary team of investigators to perform in depth chemical analysis and biological studies (both in vitro and in vivo animal studies) on this botanical product. Preclinical data from mouse studies indicate that Immulina® is most effective at alleviating illness caused by influenza when administered as a prophylactic (before infection) and as a prodromal supplement (soon after initial infection during the asymptomatic period). However, it is not effective as a therapeutic agent (post-infection during the symptomatic phase). Additionally, our team continued with human clinical biomarker-based studies on Immulina® during this period, and these studies will continue through the remainder of the funded Botanical Center grant. Marshall University Non-assisted Cooperative agreement: A Director for the Appalachian Natural Products Research Program (ANPRP) has been hired, and she will start in September 2024. Efforts are currently underway to hire technical staff to support the project. Additional supplies and equipment to support the ANPRP are in the process of being ordered in consultation with the Director. The primary investigator also visited the National Center for Natural Products Research in Oxford, Mississippi, for additional advice on advancing this project. Plant material collection is continuing in FY24. We have collected over 10 species of plants this growing season from southern West Virginia and are still collecting plant material during this growing season. We anticipate having 25+ species on hand for extraction by the end of the growing season. We are in contact with a West Virginia native plant association and local individuals knowledgeable in medicinal plants to help guide our selection of plants for study.

Accomplishments
1. Glucose uptake enhancing property of capsiate from peppers. In support of a process ARS researchers at Oxford, Mississippi, patented two years ago to produce pure capsiate from peppers, we investigated the effects of a Capsicum annuum extract and pure capsiate on metabolic disorder important receptors. The findings from this study suggest that capsiate will enhance glucose uptake effect and prevent lipid accumulation. Additionally, capsiate could inhibit the adverse effect of blood glucose lowering drugs belonging to thiazolidinediones class without compromising their main effects. This is the first report to reveal the multiple nuclear receptors agonistic action and glucose uptake enhancing property of capsiate from C. annuum along with its antiadipogenic effect indicating its potential in preventing the undesired adipogenic effects of full PPAR gamma agonists such as the glitazone class of antidiabetic drugs. This study was published this year in the Journal of Agricultural Food and Chemistry.

2. Analytical method for detecting adulterated peppermint essential oil. Peppermint essential oil (EO) is extensively utilized in the food and beverage industry. Despite its wide acceptance and economic significance, peppermint EO stands out as one of the most commonly adulterated essential oils in the United States market. At the request of doTERRA International LLC, ARS researchers at Oxford, Mississippi, collaborated with the National Center for Natural Products Research (NCNPR) at the University of Mississippi to develop an integrated approach combining conventional and chiral gas chromatography–mass spectrometry with chemometrics for assessing the quality of peppermint EOs. Using this method, poor quality or adulterated peppermint EOs can be unambiguously identified in the commercial products sold in United States market. This study was published this year in the Journal of Agricultural Food and Chemistry.

Review Publications
Liang, W., Wang, Q., Min, L., Han, L., Cantrell, C.L., Bajsa Hirschel, J.N., Duke, S.0., Ye, P., Liu, X. 2023. Synthesis, herbicidal activity and in silico analysis of novel pyrido[2,3-d]pyrimidine compounds. Molecules. https://doi.org/10.3390/molecules28217363.
Chae, H., Cantrell, C.L., Khan, I.A., Jarret, R.L., Khan, S.I. 2023. Capsiate-rich fraction of Capsicum annuum induces muscular glucose uptake, ameliorates rosiglitazone-induced adipogenesis, and exhibits activation of NRs regulating multiple signaling pathways. Journal of Agricultural and Food Chemistry. https://doi.org/10.1021/acs.jafc.3c06148?urlappend=%3Fref%3DPDF&jav=VoR&rel=cite-as.
Semerdjieva, I., Cantrell, C.L., Zheljazkov, V.D., Radoukova, T., Koleva-Valkova, L.H., Astatkie, T., Kacaniova, M., Borisova, D. 2023. Chemical profile, antioxidant and antimicrobial activity of Pinus heldreichii Christ. distributed in Bulgaria. Heliyon. 10:1-15. https://doi.org/10.1016/j.heliyon.2023.e22967.
Tan, L., He, Y., Li, S., Deng, J., Avula, B., Zhang, J., Nirmal, P.D., Solis-Sainz, J., Wang, M. 2023. Proximate composition and nutritional analysis of selected bananas cultivated in Hainan, China. Journal of Food Composition and Analysis. https://doi.org/10.1016/j.jfca.2023.105798.
Sun, N., Min, L., Sun, Z., Zhai, Z., Bajsa Hirschel, J.N., Wei, Z., Hua, X., Cantrell, C.L., Xu, H., Duke, S.O., Liu, X. 2024. Novel pyrazole acyl(thio)urea derivatives containing a biphenyl scaffold as potential succinate dehydrogenase inhibitors: Design, synthesis, fungicidal activity and SAR. Journal of Agricultural and Food Chemistry. 72:2512-2525. https://doi.org/10.1021/acs.jafc.3c07735.
Sun, X., Yu, C., Li-Jing, M., Cantrell, C.L., Hua, X., Sun, N., Liu, X. 2023. Discovery of highly efficient novel antifungal leads targeting succinate dehydrogenase: pyrazole-4-carboxamide derivatives with an N-phenyl substituted amide fragment. Journal of Agricultural and Food Chemistry. 71:19312-19323. https://doi.org/10.1021/acs.jafc.3c04842.
Adams, S.J., Lee, J., Avula, B., Katragunta, K., Wang, M., Parveen, I., Techen, N., Chittiboyina, A., Khab, I. 2023. Investigation of morpho-anatomical, molecular, GC/QToF, and LC/QToF characterization of Salvia mellifera (black sage), Saliva apiana (white sage) and their varieties. South African Journal of Botany. 164:77-90. https://doi.org/10.1016/j.sajb.2023.11.038.
Ribeiro, V., Bajsa Hirschel, J.N., Tamang, P., Meepagala, K.M., Duke, S. 2023. Antifungal and phytotoxic activities of isolated compounds from Helietta parvifolia stems. Molecules. 28(23):7930. https://doi.org/10.3390/molecules28237930.
Tamang, P., Upadhaya, A., Paudel, P., Meepagala, K.M., Cantrell, C.L. 2024. Mining Biosynthetic Gene Clusters of Pseudomonas vancouverensis Utilizing Whole Genome Sequencing. Microorganisms. 12:1-17. https://doi.org/10.3390/microorganisms12030548.
Paudel, P., Pandey, P., Paris, J.J., Ashpole, N.M., Mahdi, F., Tian, J., Lee, J., Wang, M., Xu, M., Chittiboyina, A.G., Khan, I.A., Ross, S.A., Li, X. 2023. Cannabinoid receptor type II ligands from sandalwood oil and synthetic alpha-santalol derivatives. Journal of Natural Products. https://doi.org/10.1021/acs.jnatprod.3c00282.
Moreau, T., Denning, S., Byrne, P., Volk, G.M. 2023. Climate change impacts agricultural productivity and food security. In: Volk, G.M., Moreau, T.L., Byrne, P.F., editors. Conserving and Using Climate-Ready Plant Collections. Fort Collins, Colorado: Colorado State University. Available: https://colostate.pressbooks.pub/climatereadyplantcollections/chapter/agricultural-productivity-and-food-security/
Cantrell, C.L., Travaini, M., Bajsa Hirschel, J.N., Svendsen, L.D., Reichley, A.C., Sosa, G.M., Kim, S., Tamang, P., Meepagala, K.M., Duke, S.O. 2023. Synthesis, Herbicidal Activity, and Structure-Activity Relationships of O-Alkyl Analogues of Khellin and Visnagin. Journal of Agriculture and Food Chemistry. 71:14593-14603. https://doi.org/10.1021/acs.jafc.3c03254.
Duke, S.O. 2023. Why are there no widely successful microbial bioherbicides for weed management in crops?. Pest Management Science. 80:56-94. https://doi.org/10.1002/ps.7595.
Aydogan, F., Boga, M., Khan, S.I., Zulfiqar, F., Khan, I.A., Ali, Z. 2022. Phytochemical investigation of Teucrium pruinosum and biological potential assessment of the isolated diterpenoids. Biochemical Systematics and Ecology. 105:1-4. https://doi.org/10.1016/j.bse.2022.104545.
Haron, M.H., Zhang, J., Chittiboyina, A.G., Khan, I.A., Pugh, N.D. 2023. Validation of a Toll-like receptor (TLR)2/TLR1 activation assay for biological standardization of Arthrospira/Limnospira immune-enhancing potency. Journal of Dietary Supplements. https://doi.org/10.1080/19390211.2023.2263566.
Parveen, I., Wang, M., Zhao, J., Zhu, Y., Chittiboyina, A.G., Khan, I.A., Pan, Z. 2024. Identification and functional characterization of oxidosqualene cyclases from medicinal plant Hoodia gordonii. Plants. 13(2):231. https://doi.org/10.3390/plants13020231.
Soltani, A., Ospanov, M., Ibrahim, Z.M., Bajsa Hirschel, J.N., Cantrell, C.L., Cizdziel, J.V., Khan, I.A., Ibrahim, M.A. 2024. Menthalactone from Mentha piperita L., a Monocot-Selective Bioherbicide. International Journal of Plant Biology. 15:293-303. https://doi.org/10.3390/ijpb15020025.
Akins, N.S., Mohammed, S.F., Pandey, P., Kim, S., Mahdi, F., Khan, M.H., Moss, E., Worth, C.J., Keanne, M.M., Chittiboyina, A.G., Doerksen, R.J., Paris, J.J., Le, H.V. 2023. Alleviation of cocaine withdrawal and pertinent interactions between salvinorin-based antagonists and kappa opioid receptor. ACS Chemical Neuroscience. https://doi.org/10.1021/acschemneuro.2c00806.
Khan, M.H., Mahdi, F., Penformis, P., Akins, N.S., Hossain, M., Kim, S., Sulochana, S.P., Adam, A.T., Tran, T.D., Tan, C., Claudio, P., Paris, J.J., Hoang, L.V. 2022. Synthesis and biological evaluation of tert-butyl ester and ethyl ester prodrugs of L-gamma-methyleneglutamic acid amides for cancer. Bioorganic and Medicinal Chemistry. https://doi.org/10.1016/j.bmc.2022.117137.
Thoma, J., Cantrell, C.L., Tamang, P., Zheljazkov, V.D. 2023. Determining the optimum mixture of three essential oils for potato sprout suppression at room temperature storage. Frontiers in Plant Science. https://doi.org/10.3389/fpls.2023.1199117.
Wang, M., Lee, J., Zhao, J., Lee, J., Chatterjee, S., Chittiboyina, A.G., Ali, Z., Khan, I.A. 2023. Comprehensive quality assessment of peppermint oils and commercial products: An integrated approach involving conventional and chiral GC/MS coupled with chemometrics. Journal of Chromatography B. 1232:123953. https://doi.org/10.1016/j.jchromb.2023.123953.