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United States Department of Agriculture

Agricultural Research Service

Research Project: CHEMISTRY OF NATURAL PRODUCTS FOR PEST MANAGEMENT AND CROP DEVELOPMENT

Location: Natural Products Utilization Research

2009 Annual Report


1a.Objectives (from AD-416)
Objective.
1)Identify bioactive natural compounds for herbicidal, insecticidal, fungicidal, antimicrobial and nutraceutical use from Vaccinium species, allelopathic rice varieties, and other sources. Objective.
2)Determine and manipulate the biosynthetic pathways of bioactive compound(s) in blueberry and rice.


1b.Approach (from AD-416)
General procedures for isolation of secondary metabolites will be employed such as solvent extractions, distillations, column chromatography and thin layer chromatography. Identification of biologically active compounds will be done using spectroscopic methods (UV, IR, MS, NMR). Extracts, fractions, and pure compounds will be tested for phytotoxic, antifungal, algaecidal and/or termiticidal activities. Phytotoxicity, antifungal and algaecidal assays being carried out routinely at NPURU will be utilized. Activity against the Formosan subterranean termite will be performed at the USDA-ARS, Formosan Subterranean Research Unit, New Orleans, LA. Biosynthetic studies will be performed by feeding with 13C-labelled precursors and incorporation will be determined by 13C-NMR spectroscopy. Molecular biology experiments will be performed using standard protocols.


3.Progress Report
Stilbenes that were shown in our earlier studies to have strong inhibitory effect on growth of colon cancer cell HT-29 in vitro were synthesized in large scale and tested for inhibition of HT-29 xenograft tumor growth in severe combined immunodeficiency mice. Of the stilbenes tested in mice 3,5,4’-trimethoxystilbene (a naturally-occurring stilbene) showed strong activity; tumor weight was 21% lower and tumor volume was 45% lower compared to control animals. In another study, methylether analog of resveratrol were synthesized and evaluated for activity against prostate cancer in vitro. All stilbenes inhibited the growth of the prostate cancer cell line LNCaP and exerted their effect at different stages of the cell cycle. The natural compound 3,5,4’-trimethoxystilbene was the most inhibitory against prostate cancer cells.

A six-month rat feeding study to determine the effect of blueberry powder, blueberry skins and selected natural stilbenes in preventing colon cancer development has been completed. Analyses of relevant physiological markers and serum levels of the stilbenes are in progress.

The Eastern red cedar, Juniperus virginiana, a widely distributed species in the U.S. and parts of Canada, produces two important chemical products: the anticancer compound podophyllotoxin and essential oil. We evaluated variations in podophyllotoxin concentration in Eastern red cedar, across nine physiographic regions in Mississippi: Tishomingo Hills, Bluff Region, Blackland Prairie, Jackson Prairie, Delta, Pontotoc Ridge, Flatwoods, North Central Plateau, and Piney Woods. The concentration of podophyllotoxin in red cedar needles varied from 0.11% to 0.36%. The results demonstrated that J. viriginiana could be used as a consistent source of podophyllotoxin and that the collection of plant material for commercial extraction of podophyllotoxin can be done independent of the site of collection within the state.

It is well known that the major chemical components in the venom of the red imported fire ants, Solenopsis invicta Buren, are 2-methyl-6-alkyl or alkenyl piperidines. After a column chromatography fractionation of extracts from poison glands and whole worker bodies, we have obtained fractions containing a mixture of six piperideine alkaloids. We identified these piperideine alkaloids as 2-methyl-6-tridecenyl-6-piperideine, 2-methyl-6-tridecyl-6-piperideine, 2-methyl-6-pentadecenyl-6-piperideine, 2-methyl-6-pentadecyl-6-piperideine, 2-methyl-6-heptadecenyl-6-piperideine, and 2-methyl-6-heptadecyl-6-piperideine. It was the first time these piperideine alkaloids were reported in the red imported fire ant poison glands.


4.Accomplishments
1. Structure-activity Relationship Study on the Insect Repelling Compound Callicarpenal. In earlier work, callicarpenal was identified as a lead target for structure optimization studies due to its ability to repel mosquitoes (Aedes aegypti and Anopheles stephensi) and subsequently ticks (Ixodes scapularis and others). Due to the high interest by the private sector to develop this compound as a commercial insect repellent, it was necessary to understand how the structure of callicarpenal could be improved upon and or simplified while still maintaining effectiveness. A structure-activity relationship project which identified the functional groups required to maintain both mosquito repellency and toxicity of callicarpenal was completed. All modifications in this study targeted the C-12 aldehyde or the C-3 olefin functionalities or combinations thereof. Requirements for maintaining activity were identified. In addition, the X-ray crystal structure of callicarpenal was established for the first time. Not only was mosquito biting deterrency/repellency of analogs explored but also toxicity towards mosquitoes was established which had not been demonstrated previously. Analogs and isomers of callicarpenal have been identified which could potentially lead to cheaper active ingredients with the same effectiveness as callicarpenal, should the technology be licensed. In addition, the patent can potentially be strengthened and broadened with such data.

2. Discovery of 3,5,4'-trimethoxystilbene for prevention of colon cancer. Colon cancer is the third leading cause of death due to cancer in the U.S.A. One of the approaches to controlling colon cancer is to prevent it before it develops into a malignant state. Several studies have shown prevention of colon cancer in animal experiments and have been validated in clinical trials. A number of animal studies have demonstrated prevention of colon cancer by food constituents. In a collaborative project, we have shown that the natural compound 3,5,4'-trimethoxystilbene inhibited the growth of colon cancer cells in vitro, and had greater inhibitory effect than similar compounds resveratrol and pterostilbene. Therefore the activity of 3,5,4'-trimethoxystilbene was investigated in mice and we found it to inhibit the growth of colon cancer cells injected in the hind flank of the animals; the tumor weight was 21% lower and tumor volume was 45% lower in animals treated with this compound compared to control animals. In another collaborative study, 3,5,4'-trimethoxystilbene was also demonstrated to inhibit growth of prostate cancer cells in vitro. These studies provide basis for the development plants/crops as sources of this nutraceutical.


6.Technology Transfer

Number of the New/Active MTAs (providing only)2
Number of New Commercial Licenses Executed1
Number of Other Technology Transfer1

Review Publications
Rimando, A.M., Suh, N. 2008. Biological/Chemopreventive Activity of Stilbenes and Their Effect on Colon Cancer. Planta Medica. 74:1635-1643.

Parajuli, P., Joshee, N., Rimando, A.M., Mittal, S., Yadav, A. 2009. In vitro Anti-Tumor Mechanisms of Various Scutellaria Extracts and Constituent Flavonoids. Planta Medica. 75:41-48.

Joshee, N., Rimando, A.M., Parajuli, P., Rawat, G.S., Yadav, A.K. 2009. Investigating Two Medicinal Scutellaria Species of Himalayan Origin. In Advances in Agriculture, Environment and Health: Fruits, Vegetables, Animals and Biomedical Research. Singh, S.B.; Chaurasia, O.P.; Yadav, A.; Rimando, A.M.; Terrill, T.H. (Eds.), Satish Serial Publishing House, Delhi, India. PP. 347-356.

Gawde, A.J., Zheljazkov, V.D., Maddox, V., Cantrell, C.L. 2009. Bioprospection of Eastern Red Cedar from Nine Physiographic Regions in Mississippi. Industrial Crops and Products. 30:59-64.

Joseph, J.A., Fisher, D.R., Cheng, V., Rimando, A.M., Shukitt Hale, B. 2008. Cellular and Behavioral Effects of Stilbene Resveratrol Analogs: Implications for Reducing the Deleterious Effects of Aging. Journal of Agriculture and Food Chemistry.56(22)10544-10551.

Lage, M., Cantrell, C.L. 2009. Quantification of Saffron (Crocus sativus L.) Metabolites Crocins, Picrocrocin and Safranal for Quality Determination of the Spice Grown under Different Environmental Moroccan Conditions. Scientia Horticultureae. 121:366-373.

Paul, S., Rimando, A.M., Lee, H., Ji, Y., Reddy, B.S., Suh, N. 2009. Anti-Inflammatory Action of Pterostilbene is Mediated Through the p38 Mitogen-Activated Protein Kinase Pathway in Colon Cancer Cells. Cancer Prevention Research. 2(7):650-657.

Ryabushkina, N., Gemedzhieva, N., Kobaisy, M., Cantrell, C.L. 2008. Brief Review of Kazakhstan Flora and Use of its Wild Species. Asian and Australasian Journal of Plant Science and Biotechnology. 2(2):64-71.

Stutte, G.W., Eraso, I., Rimando, A.M. 2008. Carbon Dioxide Enrichment Enhances Growth and Flavanoid Content of Two Scutellaria Species. Journal of the American Society for Horticultural Science. 133(5):631-638.

Wilson, M.A., Rimando, A.M., Wolkow, C.A. 2008. Methoxylation Enhances Stilbene Bioactivity in Caenorhabditis elegans. Biomed Central (BMC) Pharmacology. 8:15.

Cantrell, C.L., Klun, J.A., Pridgeon, Y.W., Becnel, J.J., Green Iii, S., Fronczek, F.R. 2009. Structure-Activity Relationship Studies on the Mosquito Toxicity and Biting Deterrency of Callicarpenal Derivatives. Chemistry and Biodiversity. 6:447-458.

Last Modified: 4/19/2014
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