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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

2010 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
This is the final report for research project no. 6408-41000-007-00D, a bridging project for 6408-41000-006-00D which terminated last year. Over the period covered by these two projects, significant results were realized. Objective 1 of the project (isolate and identify natural compounds with pest management and/or nutraceutical value) was fully met. Pterostilbene was identified in blueberries. In collaboration with ARS and university researchers, new activities for pterostilbene that benefit human health were discovered from both in vitro and in vivo (animal) studies. These include cholesterol lowering effect, ability to improve cognitive and motor functions in ageing, and cancer preventive property. Patent applications were filed for these biological activities, and two of these (the cholesterol lowering and the neuroprotective effects) were licensed. New dietary supplements containing pterostilbene now appear on the market. A further impact of these studies is the increase in blueberry sales and acreage attributed to the identification of pterostilbene in blueberries. Additionally, these studies have triggered new investigation by other researchers of the genetics of stilbenes in blueberries. Bioassay-guided fractionation of Callicarpa americana extract resulted in the isolation of callicarpenal, which was shown as a highly effective arthropod repellent with activity against the yellow fever mosquito vector, Aedes aegypti, the malaria mosquito vector, Anopheles stephensi, the Lyme disease deer tick vector, Ixodes scapularis, and two species of fire ants, Solenopsis (S.) invicta and S. richteri. Modification of the chemical structure of callicarpenal was performed in order to improve its activity. In addition, the X-ray crystal structure of callicarpenal was established for the first time. A patent application has been filed for the isolation and preparation of clerodanes (which include callicarpenal) as arthropod repellents. Confidentiality agreements are in place with companies that have shown interest to develop callicarpenal as a commercial insect repellent. Objective 2 of the research project (to determine the biosynthetic pathways and the genes involved in the biosynthesis of natural compounds) was also fully met. An O-methyltransferase gene from sorghum (Sorghum bicolor OMT3) capable of methylating resveratrol to produce pterostilbene was used in conjunction with a peanut stilbene synthase gene in a dual-transgene cassette approach in the transformation of Arabidopsis thaliana and Nicotiana tabacum. The levels of pterostilbene were increased in Arabidopsis and tobacco transformants. This study presents a technology to increase the plant production of pterostilbene. A patent has been issued by the U.S. Patent Office that includes a claim of a plant cell host into which a construct harboring the O-methyltransferase gene can be introduced, and expression of said construct results in the production of pterostilbene.


4.Accomplishments
1. Commercialization of Pterostilbene. Pterostilbene is a naturally-occurring phenolic compound, an analog of resveratrol, which is the well-known polyphenol in grapes and wine. Our studies have shown that pterostilbene is a more effective cholesterol lowering agent than resveratrol as far as activating a protein that plays a major role in lipid metabolism and transport. In another study, pterostilbene was shown to reverse memory and cognition deficits. For these activities, pterostilbene was licensed by a company (ChromaDex). Commercialization of pterostilbene under the tradename pTeroPure™ was launched in April 2010. pTeroPure™ pterostilbene is now on the market for use as a pure compound, or mixed with other natural compounds as dietary supplements. More pterostilbene products are expected to be on the market in the coming years.

2. Pterostilbene Shown to be Better than Resveratrol in Behavioral Tests Using Age-accelerated Mice Model. Based on earlier studies that demonstrated pterostilbene reverse cognitive deficits 19 month old rats, further studies were performed using an age-accelerated mouse model, which has characteristic learning and memory deficits at old age. In these studies two groups, one fed resveratrol and the other pterostilbene, were tested for cognitive function using the Passive Avoidance Test and the Radial Arm Water Maze test after 8 weeks of dietary supplementation. In both tests the pterostilbene group performed better than the resveratrol group. Examination of the brain tissues also showed the pterostilbene-fed animals to have higher levels of the protein superoxide dismutase (an important antioxidant defense in cells), with levels equivalent to the age-resistant control group. The pterostilbene group also showed higher levels in brain tissues of another protein called PPARalpha, which protects against brain insults. These studies provide further proof of the health benefit of pterostilbene in aging, using a relevant model. These studies will help boost the commercialization of pterostilbene.

3. Identification of Alantolactone and Isoalantolactone for the Control of Aedes (Ae.) aegypti, the Yellow Fever Mosquito. An Aedes aegypti larval toxicity bioassay was performed on compounds representing many classes of natural compounds. Among the compounds studied alantolactone and isoalantolactone showed potent larvicidal activity. Structural modifications were performed on both compounds for further structure-activity relationship study, and were evaluated for their toxicity against Ae. aegypti larvae and adults. None of the synthetic analogs of isoalantolactone were more active against the larvae than isoalantolactone itself. On the other hand, many of the analogs of alantolactone had grater larvicidal activity. The propyl amine analog of alantolactone was the most active adulticide. These compounds have the potential to be developed as agents for the control of the yellow fever mosquito.


Review Publications
Mizuno, C.S., Rimando, A.M., Duke, S.O. 2010. Phytotoxic Activity of Quinone and Resorcinolic Lipid Derivatives. Journal of Agricultural and Food Chemistry. 58:4353-4355.

Mizuno, C.S., Rimando, A.M. 2009. Blueberries and Metabolic Syndrome. Silpakorn University Open Journal Systems. 3(2):7-17.

Cantrell, C.L., Wei Pridgeon, Y., Fronczek, F.R., Becnel, J.J. 2010. Structure-Activity Relationship Studies on Natural Eremophilanes from Inula helenium as Toxicants Against Aedes aegypti Larvae and Adults. Chemistry and Biodiversity. 7:1681-1697.

Chen, J., Cantrell, C.L., Shang, H., Rojas, M.G. 2009. Piperideine Alkaloids from the Poison Gland of the Red Imported Fire Ant (Hymenoptera: Formicidae). Journal of Agricultural and Food Chemistry. Available: http://pubs.acs.org

Kim, H., Bartley, G.E., Rimando, A.M., Yokoyama, W.H. 2010. Hepatic Gene Expression Related to Lower Plasma Cholesterol in Hamsters Fed High Fat Diets Supplemented with Blueberry Pomace and Extract. Journal of Agricultural and Food Chemistry. 58 (7), pp 3984–3991. DOI: 10.1021/jf903230s.

Hoel, D., Pridgeon, J.W., Bernier, U.R., Chauhan, K., Meepagala, K., Cantrell, C. 2010. Departments of Defense and Agriculture team up to develop new insecticides for mosquito control. Wing Beats. 21(1):29-34.

Mikstacka, R., Rimando, A.M., Ignatowicz, E. 2010. Antioxidant Effect of trans-Resveratrol, Pterostilbene, Quercetin and Their Combinations in Human Erythrocytes In Vitro. Plant Foods for Human Nutrition. 65:57-63.

Paul, S., Decastro, A., Lee, H., Smolarek, A.K., So, J., Simi, B., Wang, C., Zhou, R., Rimando, A.M., Suh, N. 2010. Dietary Intake of Pterostilbene, a Constituent of Blueberries, Inhibits the B-catenin/p65 Downstream Signaling Pathway and Colon Carcinogenesis in Rats. Carcinogenesis. 31(7):1272-1278.

Vougogiannopoulou, K., Fokialakis, N., Aligiannis, N., Cantrell, C.L., Skaltsounis, A. 2010. The Raputindoles: Novel Cyclopentyl Bisindole Alkaloids from Raputia simulans. Organic Letters. 12(9):1908-1911.

Zheljazkov, V.D., Cantrell, C.L., Astatkie, T., Ebelhar, M. 2010. Productivity, Oil Content and Composition of Two Spearmint Species in Mississippi. Agronomy Journal. 102(1):129-133.

Zheljazkov, V.D., Cantrell, C.L., Astatkie, T. 2010. Study on Japanese Cornmint in Mississippi. Agronomy Journal. 102(2):697-703.

Zheljazkov, V.D., Cantrell, C.L., Astatkie, T., Ebelhar, W.M. 2009. Peppermint Productivity and Oil Composition as a Function of Nitrogen, Growth Stage and Harvest Time. Agronomy Journal. 102(1):124-128.

Zheljazkov, V.D., Cerven, V., Cantrell, C.L., Ebelhar, W.M., Horgan, T. 2009. Effect of Nitrogen, Location and Harvesting Stage on Peppermint Productivity, Oil Content and Oil Composition. HortScience. 44(5):1267-1270.

Paul, S., Mizuno, C.S., Lee, H., Zheng, X., Chajkowisk, S., Rimoldi, J.M., Conney, A., Suh, N., Rimando, A.M. 2010. In vitro and In vivo Studies on Stilbene Analogs as Potential Treatment Agents for Colon Cancer. European Journal of Medicinal Chemistry. 45:3702-3708.

Fiore, M., Rimando, A.M., Andolfi, A., Evidente, A. 2010. A New GC-MS Method for the Analysis of Ascaulitoxin, Its Aglycone and 4-Aminoproline from Culture Filtrates of Ascochyta caulina. Analytical Methods. 2:159-163.

Last Modified: 8/30/2014
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