The Phytochemical Society of North America August 8–12, 2015 University of Illinois at Urbana-Champaign

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To Willard Airport (CMI) ST. MARY’S ROAD Savoy I Hotel and Conference Center Contents 2015 PSNA Meeting 2 Greetings! Organizing Committee 3 General Information Mark Berhow, USDA, ARS, Peoria, IL 3 Registration Desk Hours Franck Dayan, USDA, ARS, Oxford, MS 3 Meeting Venues Elvira de Mejia, University of Illinois at 4 Meeting Hotels Urbana-Champaign 4 Transportation David Gang, Washington State University 4 Parking Massuo Jorge Kato, University of Sao 4 Wi-Fi Access Paulo, Brazil 5 Emergency Numbers and Procedures Aruna Kilaru, East Tennessee State University 5 Moderators, Speakers, Poster Presenters, and Exhibitors Dhirendra Kumar, East Tennessee State University 5 Refreshments Argelia Lorence, Arkansas State 6 Meeting Agenda University 6 Saturday, August 8 Susan McCormick, USDA, ARS, Peoria, IL 6 Sunday, August 9 Shelly Nickols-Richardson, University of 7 Monday, August 10 Illinois at Urbana-Champaign 8 Tuesday, August 11 Dejan Nikolic, University of Illinois at 9 Wednesday, August 12 Chicago J. Fred Stevens, Oregon State University 10 Speaker Abstracts Lloyd Sumner, the Noble Foundation, 10 Symposium 1: Phytochemical and Plant Ardmore, OK “omics” Dorothea Tholl, Virginia Technical 15 Symposium 2: Phytochemical Lipids University and Metabolism Li Tian, University of California at Davis 19 Symposium 3: Phytochemical Biosynthesis De-Yu Xie, North Carolina State University 24 Symposium 4: Chemical Ecology I 30 Symposium 5: Chemical Ecology II 34 Symposium 6: Botanical Medicines 39 Symposium 7: Nutritional & Medicinal Phytochemistry 44 Poster Abstracts 74 Participant List

August 8–12, 2015 • University of Illinois at Urbana-Champaign 1 Greetings!

Welcome to the twin cities of Urbana and Founded in 1867 as a state-supported, land- Champaign, in the heart of east central Illinois. grant institution under the terms of the Morrill In case you’re wondering, they’re not identical Land Grant Act of 1862 with a threefold mission twins; their births were separated by about of teaching, research, and public service, the 30 years. Urbana dates back to 1822, when UIUC opened its doors to students in 1868 and William Tompkins built the first homestead on rapidly earned a reputation as an educational the site that 11 years later was officially platted institution of international stature. Today, as a new city. Champaign didn’t come into scholars and educators rank it among a select existence until 1852, when the Illinois Central group of the world’s great universities. For Railroad laid tracks two miles outside of Urbana more information, see the university website and residents who settled close by rejected (www.illinois.edu). annexation efforts and instead incorporated UIUC is a residential campus of classrooms, their own city a few years later. Despite the fact laboratories, libraries, residence halls, and that the two cities now share many services recreational and cultural facilities with 200 (as well as Wright Street, the official dividing major buildings on the central campus of line) and are cooperative and cordial with 1,783 acres. Nearby are the University’s 1,650- each other, referenda to merge the two into acre Willard Airport; Robert Allerton Park, the a single metropolis have been, since 1855, campus’s 1,768-acre nature and conference consistently and resoundingly defeated. The center; and 3,600 acres of agricultural land. An courthouse in Urbana, the permanent county additional 3,700 acres of farmland elsewhere in seat for Champaign County, was frequented Illinois are used by the College of Agricultural, in the 1840s and 1850s by a young lawyer Consumer, and Environmental Sciences as who practiced law in the 8th Judicial Circuit. In experimental fields. The University also owns 1854, this lawyer—named Abraham Lincoln— close to 1,000 acres of natural areas within delivered one of his first public speeches a 50-mile radius of the campus, held for the against slavery in the county courthouse in express purpose of conducting ecological downtown Urbana. research. About the University of Illinois at The University of Illinois and its various Urbana-Champaign campuses have had a long history of scientific exploration into the world of plants, both in The lives of the residents of Urbana and production and in biological and chemical Champaign were changed forever when, characterization. The Morrow Plots are the in 1867, the Illinois Industrial University, oldest agronomic experiment fields in the later to become the University of Illinois at United States which include the longest- Urbana-Champaign, was chartered. The term continuous corn plot in the world. In University of Illinois at Urbana-Champaign 1968, the Morrow Plots became a National is a comprehensive, major public university Historic Landmark. Some of the seminal work that is ranked among the best in the country.

2 The Phytochemical Society of North America General Information understanding on photosynthesis and carbon Registration Desk Hours dioxide fixation was done in part here including the work of the labs of Govindjee and W.L. The registration desk is located outside Illini Ogren. The University continues to feature a Rooms ABC on the main level of the Illini diverse array of agricultural-based research Union, except as noted. in the College of Agricultural, Consumer and Saturday, August 8 4:30–7:30 pm Environmental Sciences, one the hosts of this I Hotel and Conference Center—Lincoln Room meeting. Sunday, August 9 7:30 am–5:30 pm Today, UIUC is home to life scientists across Monday, August 10 the campus using new genomic tools, 7:30 am–5:30 pm computational methods and chemical analytical Tuesday, August 11 7:30 am–5:30 pm techniques to explore mechanisms of interest Wednesday, August 12 to phytochemists. In addition to departmental 7:30–11:00 am programs, the UIUC Institute for Genomic Your registration includes the conference Biology, is an international center for genomics program, the welcome reception on Saturday, research. Other unique campus research refreshment breaks Sunday-Wednesday, lunch resources include SoyFACE, a 60-acre facility Sunday-Tuesday, poster sessions on Sunday instrumental in documenting the chemical and Monday, and the banquet on Tuesday. A ecology of global climate change in agriculture, limited number of guest banquet tickets may the world’s largest public university library (with be available for purchase at the registration 24 million-plus items in its catalogue), and Blue desk during the conference. Waters, the petaflop supercomputer that is the fastest anywhere in the world on a university Meeting Venues campus), capable of over 13 quadrillion Illini Union calculations per second. With 22 Nobel Prize (Sunday–Wednesday) winners and 20 Pulitzer Prize winners among 1401 W. Green St. faculty and alumni, the University of Illinois is Urbana, IL 61801 the number one return on investment in the Phone: 217-333-4666 Midwest. union.illinois.edu I Hotel and Conference Center “The best way to predict the future is (Saturday’s welcome reception) to create it.” 1900 S. 1st St. —Abraham Lincoln Champaign, IL 61820 Phone: 217-819-5000 Welcome to Illinois! stayatthei.com May Berenbaum Alice Campbell Alumni Center Mark Berhow (Tuesday’s banquet) Elvira de Mejia 601 S. Lincoln Ave. Urbana, IL 61801 Phone: 217-333-1471 uiaa.org/alumnicenter

August 8–12, 2015 • University of Illinois at Urbana-Champaign 3 Meeting Hotels Terminal daily (217-352-4150 or greyhound. com). A few Amtrak trains serve Champaign- Illini Union Hotel Urbana via the Illinois Terminal in Champaign 1401 W. Green St. (amtrak.com). Urbana, IL 61801 Phone: 217-333-4666 Taxi Service union.illinois.edu Yellow Checker Cab, Phone: 217-355-3553 Orange Taxi, Phone: 217-363-1500 I Hotel Green Taxi, Phone: 217-721-5533 1900 S. 1st St. Quality Limo & Taxi, Phone: 217-552-7400 Champaign, IL 61820 Phone: 217-819-5000 Willard Airport stayatthei.com 11 Airport Rd., Savoy, IL 61822 Phone: 217-244-8618 Comfort Suites flycmi.com 2001 N. Lincoln Ave. Urbana, IL 61801 Parking Phone: 217-328-3500 comfortsuites.com/hotel-urbana-illinois-IL366 Parking is free and plentiful at the I Hotel and Conference Center and Comfort Suites. If you Transportation are staying at the Illini Union Hotel, you will be given a complimentary parking pass to park in Some area hotels have a complimentary shuttle their adjacent lot. For those staying elsewhere, that makes trips between the airport, the hotel, there is limited metered parking near the Illini and campus. Please call your hotel or check at Union and Alice Campbell Alumni Center. Be their front desk to arrange for a ride. sure to bring quarters to feed the meters or Champaign-Urbana MTD Bus System plan to pay by phone. If you are willing to walk Champaign-Urbana has a great public a few blocks, you may be able to find free transportation system, with several stops on parking in the residential area east of the Illini campus and near the conference hotels. The Union, just past Lincoln Avenue. 27 Air Bus runs between Willard Airport in Savoy and the Illini Union several times a day. Wi-Fi Access A standard ride costs $1, one-way, with free Wireless Internet access is free at the transfers from route to route. Only coins and $1 meeting venues. Please select the network bills accepted. Drivers can make change for up UIpublicWiFi, open your browser, and enter to $5 prior to 7:00 pm. Exact cash is required your contact information when prompted. after that time. Visit cumtd.com for maps and schedules, or call 217-384-8188. Other Bus and Train Services Peoria Charter coach buses depart from campus and the Illinois Terminal in Champaign daily (800-448-0572 or peoriacharter.com). Greyhound buses also depart from the Illinois

4 The Phytochemical Society of North America Emergency Numbers and Procedures Speakers Please consult the program ahead of time to Emergency (Police, Fire, or Ambulance): 911 confirm the time and location of your talk. Non-Emergency Arrive early at your session, and if you are University Police: 217-333-1216 planning to use slides, email them in advance Urbana Police: 217-384-2320 and also bring them on a USB stick. Find the Champaign Police: 217-351-4545 session moderator and identify yourself so that he or she is aware that you are present. Please Medical Assistance try to stay within your allotted time—it’s a Carle Foundation Hospital: 217-383-3311 courtesy to your audience and fellow speakers Christie Clinic: 217-366-1200 in your session. Provena Covenant Medical Center: 217-337- 2000 Poster Presenters The maximum size is the standard for scientific Tornado Preparedness posters, 4-feet by 4-feet. Landscape is East-central Illinois is prone to summer the typical orientation, but portrait is also thunderstorms; often conditions are conducive acceptable as long as it isn’t wider or taller than to tornadoes. If you hear the sirens go off, 4-feet. The display boards will be numbered a tornado warning is in effect. Take cover to indicate where to place your poster. Push immediately in the lowest floor of a building, pins will be provided to affix your poster to the and stay away from windows. board. Please put up your poster by 12:00 pm Fire Procedure on Sunday, and stand by your poster during If a fire alarm goes off, exit the building in your designated poster session. Any posters a calm and orderly manner. If you are on an remaining after Tuesday will be discarded. upper level floor, exit by the nearest stairwell. Poster Session I Do not use elevators. Odd-numbered posters (e.g. P1, P3, P5...) Sunday, August 9, 5:30–7:30 pm Moderators, Speakers, Poster Illini Room AB Presenters, and Exhibitors Poster Session II Moderators Even-numbered posters (e.g. P2, P4, P6...) If you are moderating a session, please arrive Monday, August 10, 5:30–7:30 pm a few minutes early to ensure that audio-visual Illini Room AB equipment is in place and functional. At each session, a student volunteer should be on Exhibitors hand to assist you. The speaker’s allotted time Exhibitors will be set up in Illini Room AB from includes time for questions. The moderator Sunday–Tuesday. should alert speakers when 3 minutes remain and again indicate when one minute remains. Refreshments After their allotted time, all speakers should Refreshments will be available in Illini Room AB be asked to leave the podium. If the speaker at the start of each day and during the break has used his/her entire allotted time, then the times. There are also a few eateries in the speaker cannot take questions. Moderators, Illini Union such as Jamba Juice and Espresso please announce this format at the beginning Royale (main level) and Einstein Bros. Bagels of the session, and stay on schedule. (lower level).

August 8–12, 2015 • University of Illinois at Urbana-Champaign 5 Meeting Agenda

All events take place in Illini Rooms ABC on the • 11:00 am | Fred Stevens, Oregon State main level of the Illini Union, except as noted. University. Shotgun metabolomics and lipidomics for mechanistic elucidation of the Saturday, August 8 anti-obesity effects of xanthohumol from hops (Humulus lupulus). 2:00–5:00 pm | Executive Committee Meeting, I Hotel and Conference Center—Lincoln Room • 11:20 am | Parnian Lak, Oregon State University. LC-MS/MS investigation of the 5:30–7:30 pm | Welcome Reception, I Hotel active phytochemicals in Centella asiatica. and Conference Center—Lincoln Room • 11:40 am | Marcos Soto Hernadez, Colegio Sunday, August 9 de Postgraduados. NMR-1H metabolomic analysis of Mexican races of avocado. Symposium 1: Phytochemical and Plant “omics” 12:00–1:30 pm | Lunch Chairs: Lloyd Sumner and Cecilia McIntosh Symposium 2: Phytochemical Lipids & • 8:00 am | Welcome by Robert Hauser, Dean, Metabolism College of ACES, University of Illinois at Chairs: Aruna Kilaru and Argelia Lorence Urbana-Champaign • 1:30 pm | Kent Chapman, Department of • 8:15 am | Miroslava Cuperlovic-Culf, National Biological Sciences, University of North Research Council. Cheminformatics and Texas. Metabolism and functions of N‐ metabolomics converge on wheat—fungal acylethanolamines in seedling development. pathogen interactions. • 2:15 pm | Pradeep Kachroo, Department • 9:00 am | Dean DellaPenna, Department of Plant Pathology, University of Kentucky. of Biochemistry and Molecular Biology, Chemical- and lipid-mediated systemic Michigan State University. Transorganellar acquired resistance in plants. complementation functionally demonstrates • 3:00 pm | Neish Award Speaker: Abe Koo, a new interface for the synthesis of non- Department Biochemistry, University of polar metabolites by membrane spanning Missouri. Metabolism of plant hormone pathways jasmonate: A sentinel for tissue damage and • 9:45 am | Elsevier Award Speaker Nicole master regulator of stress response. Clay, Department of Molecular, Cellular & • 3:35 pm | Break Developmental Biology, Yale University. Evolutionary toolkits for chemical innovation • 3:50 pm | Aruna Kilaru, East Tennessee State in land plants. University. Transcriptome analysis of avocado mesocarp reveals key genes necessary to • 10:30 am | Break improve oil yield. • 10:40 am | Lloyd Sumner, Noble Foundation. • 4:10 pm | Md Mahbubur Rahman, East Large-scale, computational and empirical Tennessee State University. Identification, UHPLC-MS-SPE-NMR annotation of plant and heterologous expression analysis of metabolomes. Avocado DGAT1 and DGAT2

6 The Phytochemical Society of North America • 4:30 pm | Reinhard Jetter, University of • 11:20 am | Li Tian, University of California, British Columbia. Wheat waxes revisited: Davis. Investigations on early steps of composition and biosynthesis. hydrolyzable tannin biosynthesis. • 4:50 pm | Samiddhi Senaratne, University • 11:40 am | Toshiaki Umezawa, Kyoto of Melbourne. Monoterpene acid glucose University. Structural modification and esters in Eucalyptus foliar oil glands. increase of lignin in Oryza sativa for biomass refinery. • 5:10 pm | Ebenezer Ajewole, University of Western Ontario/Agriculture and Agri-Food 12:00–1:30 pm | Lunch Canada. K+ Dependent and K+ independent Symposium 4: Chemical Ecology I Asparaginase from common Bean (Phaseolus Chairs: Dorothea Tholl and Massuo Kato vulgaris): mechanism of activation by K+. • 1:30 pm | May Berenbaum, Department of 5:30–7:30 pm | Poster Session I Entomology, University of Illinois. Honey Monday, August 10 bees as phytochemists: how pollen and nectar are converted into beebread and Symposium 3: Phytochemical Biosynthesis honey. Chairs: Li Tian and De-Yu Xie • 2:15 pm | Walter S. Leal, Department of • 8:00 am | Greetings Molecular and Cellular Biology, University of California at Davis. Mosquito odorant • 8:15 am | Xiao-Ya Chen, Institute of Plant receptor for the insect repellent DEET and Physiology and Ecology, Shanghai Institutes plant-derived semiochemicals. for Biological Sciences, Chinese Academy of Science. Regulation of sesquiterpene • 3:00 pm | Neish Award Speaker: Martha biosynthesis in plants. Vaughan, USDA, ARS, Peoria, IL. Climate change and crop phytochemical defenses: • 9:00 am | Joe Chappell, Department of Potential implications for food security and Plant & Soil Science, University of Kentucky. food safety. Valeriana officinalis as a novel platform for plant natural product drug discovery. • 3:35 pm | Break • 9:45 am | Cecilia McIntosh, mcintosc@etsu. • 3:50 pm | Katharina Schramm, University of edu, Department of Biological Sciences, East Utah. Metabolomics of juniper detoxification Tennessee State University. Structure and in a generalist and specialist mammalian function of flavonoid glucosyltransferases: herbivore. using a specific grapefruit enzyme as a • 4:10 pm | Felipe Christoff Wouters, Max model. Planck Institute for Chemical Ecology. • 10:30 am | Break Detoxification of maize chemical defenses by Lepidopteran herbivores. • 10:40 am | Hiroshi Maeda, University of Wisconsin-Madison. Diversification • 4:30 pm | Verena Jeschke, Max Planck of tyrosine biosynthetic pathways in Institute for Chemical Ecology. Toxicity of plants: Non-plastidic, tyrosine-insensitive glucosinolate-derived isothiocyanates to prephenate dehydrogenases in legumes. generalist-feeding caterpillars. • 11:00 am | Mark Lange, Washington State • 4:50 pm | De-Yu Xie, North Carolina State University. Evolution of chemical diversity in University. Gene-to-terpene landscapes in plants self-pollinating Artemisia annua.

August 8–12, 2015 • University of Illinois at Urbana-Champaign 7 • 5:10 pm | Michael Sullivan, US Dairy • 11:40 am | Mitchell Wise, Research Chemist Forage Research Center, ARS-USDA. USDA, Cereal Crops Research Unit. Field Engineering alfalfa to accumulate useful application of benzothiadiazole on oat: Effect caffeic acid derivatives and characterization on avenanthramide production and crown of hydroxycinnamoyl-CoA transferases from rust resistance. legumes. 12:00–1:30 pm | Lunch 5:30–7:30 pm | Poster Session II Symposium 6: Botanical Medicines Tuesday, August 11 Chairs: Dejan Nikoli and Fred Stevens • 1:30 pm | Richard van Breemen, Department Symposium 5: Chemical Ecology II of Medicinal Chemistry and Pharmacognosy, Chairs: Dhirendra Kumar and De-Yu Xie University of Illinois at Chicago. • 8:00 am | Greetings Development of safe and effective botanical dietary supplements for women’s health. • 8:15 am | Jonathan Gershenzon, Department of Biochemistry, Max Plank Institute for • 2:15 pm | Ehab Abourashed, Department Chemical Ecology. Metabolic tag in the of Pharmaceutical Sciences, Chicago State cornfield: Biosynthesis and activation University. The non-volatile constituents of of benzoxazinoids by corn plants and nutmeg—a key to solving an old riddle? deactivation by herbivores. • 3:00 pm | Neish Award Speaker: Philipp • 9:00 am | Eric Schmelz, Department of Zerbe, Dept of Plant Biology, University Biological Sciences, University of California of California, Davis. Elucidating modular at San Diego. Inducible small molecule diterpenoid metabolism in non-model defenses in maize: more than a few surprises. plant systems: From chemical diversity to biotechnology applications. • 9:45 am | Jyoti Shah, Department of Biological Sciences, University of North • 3:35 pm | Break Texas. Signaling function for an abietane • 3:50 pm | Foluso Oluwagbemiga Osunsanmi, diterpenoid in plant defense and University of Zululand. Antiplatelet development. aggregation and cytotoxicity activity of • 10:30 am | Break betulinic acid and its acetyl derivative from Melaleuca bracteates var. revolution gold. • 10:40 am | Franck Dayan, USDA/ARS, Oxford, MS. Sarmentine, a natural piper • 4:10 pm | Tristesse Burton, University amide herbicide with multiple mechanisms of of Illinois at Chicago. American Indian action. botanicals as possible alternatives to hormone therapy. • 11:00 am | Dimitre Ivanov, University of Western Ontario. The chemoattractant • 4:30 pm | Massuo Kato, University of Sao potential of ginsenosides in the ginseng - P. Paulo. Molecular diversity and evolution of irregulare pathosystem, Peperomia species. • 11:20 am | Chintamani Thapa, University of • 4:50 pm | Break Saskatchewan. Chemical interaction between 5:00–6:00 pm | Annual Meeting of the PSNA the cruciferous phytoalexin camalexin and Colletotrichum species.

8 The Phytochemical Society of North America 6:30–9:00 pm | Award Banquet, Alice Campbell Alumni Center Ballroom Welcome by Neal Merchen, Associate Dean for Research, College of ACES, University of Illinois at Urbana-Champaign

Wednesday, August 12 Symposium 7: Nutritional & Medicinal Phytochemistry Chairs: Elvira de Mejia and Shelly Nickols- Richardson • 8:00 am | Greetings • 8:15 am | Mary Ann Lila, Plants for Human Health Institute, North Carolina State University. Phytoactive chemicals: discovery is just at the “starting line” of the research. • 9:00 am | John (Jack) Juvik, Department of Crop Sciences, University of Illinois at Urbana-Champaign. Manipulation of glucosinolate biosynthesis, hydrolysis, and hydrolysis product anticancer bioactivity in broccoli (Brassica oleracea L. ssp.). • 9:45 am | Okiemute Rosa Johnson-Ajinwo, Keele University. Anti-malarial activities of Margaritaria discoidea and other Nigerian medicinal plants. • 10:15 am | Syeda Hussain, University of Sao Paulo. Phytochemical analysis for phenol and saponin in Brachiaria sp. • 10:45 am | Pavel Somavat, University of Illinois at Urbana-Champaign. Coproduct yield comparisons between purple corn, blue corn, and yellow dent corn for different milling processes. • 11:15 am | Richard Licayan, Rutgers University. Investigation of α-glucosidase inhibition properties, antioxidant activities, and cytotoxicity of Philippine herbal vines containing polyphenols.

August 8–12, 2015 • University of Illinois at Urbana-Champaign 9 Speaker Abstracts

Sunday, August 9, Morning approaches delivers fast, inexpensive way for molecular follow-up in treatment development Symposium 1: Phytochemical and and assessment as well as determination of Plant “omics” markers for targeted sensors development. Although current metabolomics approaches Chairs: Lloyd Sumner and Cecilia McIntosh can only view several hundreds of the Miroslava Cuperlovic-Culf estimated 200,000 compounds, information obtained Ph.D. in Biophysical about the concentration changes in this subset Chemistry at University of of metabolites in different conditions provides California, Santa Barbara valuable overview of distinct compound followed by post-doctoral classes and regions in the metabolic network. training in Biophysics with Prof. Metabolic changes are a significant part of Myer Bloom at University of plants’ response to pathogens. Recently British Columbia. Currently she is a Senior published studies have determined number Research Officer at National Research Council of resistance related metabolites as well as of Canada and Adjunct Professor of Chemistry important metabolic pathways involved in the in the department of Chemistry & Biochemistry response of plants to fusarium head blight at Universite de Moncton. Miroslava is and rust. Function of the majority of these particularly interested in metabolomics and metabolites in the resistance remains, however, computational analysis of biological system unknown. At the same time, a number of plant under different conditions and treatments. metabolites have been extensively studied for their health effect in mammalian cells. In this [S1-1] Cheminformatics and work we are combining results of metabolomics metabolomics converge on wheat– measurements in plant-pathogen interactions fungal pathogen interactions with cheminformatics analysis of properties of Presenter and Author: Miroslava Cuperlovic-Culf, molecules to describe major metabolic changes National Research Council, Canada inducing resistance in plants to fusarium head Pathogenic fungi are increasingly recognized blight and rust fungi. For known resistance as a foremost treat to plant, animal and related metabolites we are subsequently human health. Fungal pathogens are leading working on determining molecular functions to some of the most destructive diseases in and targets in either fungus or plant using plants including major damage to agricultural methods adopted from drug discovery crops such as rice, wheat and corn. In addition practices. to devastating plant yield, fungal toxins can Although function of the majority of resistance present a danger to animal and human health related metabolites remains unknown, several with many historical examples of fungal fungal proteins have been determined as toxins causing catastrophic losses of life. The possible targets. Out of these in this work development of fungal infection resistant we have explored histone deacetylases, plants, novel, environmentally friendly, anti- carbonic anhydrases, cytochrome P450 and fungal treatments as well as more informative lipoxygenases. All four groups of proteins methods for treatment follow-up is sorely have experimentally proven roles in fungal needed. Unbiased analysis of metabolites pathogenesis with number of plant and fungal provided by different metabolomics metabolites acting as their potent inhibitors.

10 The Phytochemical Society of North America Specific focus in the talk will be on the breed and engineer crops to provide balanced interaction of resistance related metabolites nutrition on a global scale. He was also a with these protein targets. Structures of these member of the Medicinal Plant Consortium; a four groups of proteins are conserved across multi institutional effort generating species and therefore can be inhibited by transcriptome and metabolome data from the same molecules. Roles of these groups medicinal plants for use in dissecting the of proteins in plant—fungus interaction were biosynthesis of plant-derived compounds of explored using computational modelling and medical significance. metabolomics analysis. Computational analysis was also performed to determine inhibitory [S1-2] Transorganellar complementation power of significantly over-concentrated functionally demonstrates a new metabolites against plant and fungal proteins. interface for the synthesis of non-polar Experimental testing has also been initiated metabolites by membrane spanning exploring the effect of inhibition of these pathways proteins on fungal metabolism and toxin Presenter and Author: Dean DellaPenna, production using NMR and FT-IR spectroscopy. Department of Biochemistry and Molecular Biology, Results of these studies will be presented. Michigan State University Plastids are subcellular factories that participate Dean DellaPenna is a MSU in the synthesis of a bewildering array of Foundation Professor and compounds, often by initiating biosynthetic University Distinguished pathways that are subsequently completed Professor in the Department of in other organelles. Such organelle-spanning Biochemistry and Molecular pathways require extensive exchange Biology at Michigan State of metabolites with the extraplastidic University. He earned a B.S. in environment, which for polar metabolites, Cellular Biology at Ohio University in 1984 and is handled by dozens of well-characterized a Ph.D. in Plant Physiology at the University of envelope membrane transporters. However, California at Davis in 1987. He was on the for the many thousands of plastid-synthesized faculty in the Plant Sciences Department at the nonpolar compounds synthesized by University of Arizona from 1990 to 1996 and membrane-spanning pathways, such then in the Biochemistry Department at the transporters have remained elusive. This talk University of Nevada, Reno before moving to will highlight recent data from an approach we Michigan State University in 2000. Dr. have termed transorganellar complementation DellaPenna received the Distinguished Faculty that functionally demonstrates enzymes in Award from MSU in 2008 and was elected a one organelle can directly access nonpolar Fellow of the American Association for metabolites from a companion organelle. We Advancement of Science (AAAS) in 2009. His propose a mechanism, based on hemifused- scientific interests lie at the interface of plant membranes at plastid:ER contact sites, that biochemistry and human health. He has allows enzymes in one organelle direct, pioneered genomics-enabled approaches to transporter-independent access to a range understand the synthesis and accumulation of of nonpolar compounds in both organelle essential dietary nutrients in plant tissues and membranes. Such an interface would facilitate he currently leads a consortium employing inter-organellar metabolism, allosteric genome wide association studies and deep regulation between organelles and allow the sequencing to understand and compare the observed rapid evolution of chemical diversity control of essential nutrient content and by membrane-spanning pathways to be composition in model plants and crops. He is a vocal proponent of using such knowledge to

August 8–12, 2015 • University of Illinois at Urbana-Champaign 11 uncoupled from coevolution with nonpolar Nicole K. Clay received her metabolite transporters. B.S. in Biology (1996) from Massachusetts Institute of [S1-3: Elsevier Award Speaker] Technology, and her Ph.D. in Evolutionary toolkits for chemical Biology (2005) from Yale innovation in land plants University. She received her Presenter and Author: Nicole Clay, Department of postdoctoral training in the Molecular, Cellular and Developmental Biology, Yale field of plant-microbe interactions (2005-2010) University. at Massachusetts General Hospital, an affiliate Plant secondary metabolites, both constitutive of the Harvard Medical School. In 2011, she and pathogen-inducible, have recently been joined the faculty at Yale University as an shown to be involved in controlling several Assistant Professor of Molecular, Cellular & evolutionary conserved plant innate immune Developmental Biology. Her research program responses, such as callose deposition and is focused on understanding how the function programmed cell death. Despite advances in and cell-surface abundance of immune our understanding of the contribution of plant receptors are regulated by the secretory and secondary metabolites to plant immunity, many endocytic pathways, and how the plant defense pathogen-inducible genes, pathways and network of specialized metabolic pathways is metabolites remain uncharacterized in even evolutionarily driven to diversification and the most-characterized model plant species. functionalization. Using expression profiling, co-expression analysis and untargeted metabolic profiling, [S1-4] Large-scale, Computational we have uncovered the complete biosynthetic and Empirical UHPLC-MS-SPE-NMR pathway to a previously unknown class of Annotation of Plant Metabolomes tryptophan-derived cyanogenic metabolites Presenter(s): Lloyd W. Sumner, Plant Biology in Arabidopsis. The enzymes catalyzing the Division, the Samuel Roberts Noble Foundation first committed steps of the novel cyanogenic Author(s): Lloyd W. Sumner, Feng Qiu, Dennis Fine, indole biosynthetic pathway and the ancestral Daniel Wherritt, Zhentian Lei, Plant Biology Division, camalexin biosynthetic pathway are encoded the Samuel Roberts Noble Foundation, 2510 Sam Noble Parkway, Ardmore, OK 73401, USA by paralogous cytochrome P450 genes, a finding that suggests that gene duplication and Integrated metabolomics is a revolutionary neofunctionalization of core pathway enzymes systems biology tool for understanding plant can serve as drivers of chemical diversity in metabolism and elucidating gene function. plant secondary metabolism. In addition, we Although the vast utility of metabolomics have identified two multiprotein regulatory is well documented in the literature, its full complexes involved in the production scientific promise has not yet been realized due of clade-specific phenylpropanoids and to multiple technical challenges. The number aromatic alkaloids in Arabidopsis. The stable one, grand challenge of metabolomics is the subnetworks of transcription factors at the core large-scale confident chemical identification of of these complexes may be highly conserved metabolites. To address this challenge, we have in distantly related plant species, a finding that developed powerful software entitled Plant suggests that the recruitment and remodeling Metabolite Annotation Toolbox’ (PlantMAT) of ancient gene regulatory networks during and are coupling it to a sophisticated ensemble land plant evolution can also serve as drivers composed of Ultrahigh pressure liquid of chemical diversity in plant secondary chromatography coupled to mass spectrometry metabolism. coupled to automated solid-phase extraction and NMR (UHPLC-MS-SPE-NMR) for the

12 The Phytochemical Society of North America large-scale systematic and biological directed about the potential benefits/risks of chronic annotation of plant metabolomes. exposure to XN in vivo. Even less is known about the underlying mechanisms of action. UHPLC-QTofMS/MS metabolite profiling The objective of this study is to identify the was performed on 80% aqueous methanol molecular targets of XN in obesity rodent extracts of Medicago truncatula, and the models that mimic metabolic syndrome in data processed using Bruker’s Compass humans. Zucker fa/fa rats and diet-induced DataAnalysis 4.1 for peak deconvolution and obese C57BL/6J mice, both accepted models formula prediction. Metabolite identifications of obesity and metabolic syndrome, were were first attempted through spectral matching treated for 6-12 weeks with XN at three dose with custom libraries generated with authentic levels (n=12-16/dose group). The XN-treated compounds. However, authentic compounds animals had significantly lower plasma glucose are not available for all metabolites; especially levels and smaller body weight gain compared plant natural products, and a large number of to the control groups, while food intake was the detected peaks could not be identified not affected by treatment. Metabolome and using spectral matching. Thus, the orthogonal lipidome profiles of plasma and liver tissue data was imported into PlantMAT and were recorded in addition to single endpoints structures for approximately 100 saponins of glucose and lipid homeostasis. Metabolites and polyphenolic glycosides were efficiently were identified using an in-house library of predicted. Ten of these were chosen for further >600 metabolites (IROA Technologies) and structural validation. These ten compounds by online database searching. We detected were isolated, purified and concentrated >5,000 metabolites by LC-QToF mass by mass directed UHPLC-MS-SPE. The spectrometry, of which >200 metabolites SPE isolated compounds were eluted with were identified by mass, isotope distribution, approximately 250 ul of deuterated methanol MS/MS fragmentation pattern, and when and 1D and 2D NMR spectra acquired. The standards were available, retention time. The NMR spectral data confirmed a 100% accuracy shotgun metabolomics and lipidomics analyses in the PlantMAT predicted structures. The showed a dose-dependent decrease of hepatic results demonstrated that the cumulative triglyceride content and metabolic products platforms allows for higher-throughput and of dysfunctional lipid metabolism (medium- high confidence metabolite identifications chain acylcarnitines, dicarboxy fatty acids, necessary for metabolome ‘sequencing’. hydroperoxy and hydroxy fatty acids). Taken together, the results indicate that xanthohumol [S1-5] Shotgun Metabolomics and improves beta-oxidation of fatty acids. These Lipidomics for Mechanistic Elucidation of anti-obesity effects are also consistent with our the Anti-obesity Effects of Xanthohumol preliminary results showing that XN inhibits from Hops (Humulus lupulus) de novo lipogenesis in differentiating 3T3-L1 Presenter(s): Jan Frederik Stevens, Linus Pauling adipocytes, determined by deuterium labeling Institute & College of Pharmacy and subsequent mass isotopomer distribution Author(s): J. Choi, C.L. Miranda, J.F. Stevens, Linus analysis (MIDA) of triglycerides. Pauling Institute & College of Pharmacy, Oregon State University, OR 97331, USA [S1-6] LC-MS/MS investigation of the Xanthohumol (XN) is a prenylated flavonoid active phytochemicals in Centella found in hops, beer, and in dietary asiatica supplements. There is substantial evidence Presenter(s): Parnian Lak, Department of Chemistry, in the literature that XN improves glucose Oregon State University, Corvallis, OR; Linus Pauling and lipid metabolism of mammalian cells in Institute, Oregon State University, Corvallis, OR; vitro, but relatively little is currently known

August 8–12, 2015 • University of Illinois at Urbana-Champaign 13 Department of Pharmaceutical Sciences, Oregon also been used to investigate the molecular State University, Corvallis, OR. characteristics of CAW phytochemicals Author(s): Parnian Lak1,2,5; Nora Gray3; Christopher inhibiting FAAH. Harris3; Jeff Morré1; Margeux Hunter3; Trevor 1 2 3,4 Shear ; William Bisson ; Joseph Quinn ; Amala [S1-7] NMR-1H metabolomic analysis of Soumyanath3; Jan F. Stevens2,5; Claudia Maier1,2 mexican races of avocado 1Department of Chemistry, Oregon State University, Corvallis, OR; 2Linus Pauling Institute, Oregon State Presenter(s): Marcos Soto-Hernandez, Colegio de University, Corvallis, OR; 3Department of Neurology, Postgraduados México Oregon Health and Science University, Portland, OR; Author(s): Emmanuel Ibarra-Estrada, Colegio de 4Department of Neurology and Parkinson’s Disease Postgraduados, postgrado en Fisiología Vegetal. Research Education and Clinical Care Center Alejandro Barrientos-Pliego, Universidad Autonoma (PADRECC), Portland Veterans Affairs Medical Chapingo Center, Portland; 5Department of Pharmaceutical Sciences, Oregon State University, Corvallis, OR The avocado is one of the fruits more important in México, and its consumption has increased Centella asiatica (CA) has been traditionally considerably. The variety ‘Hass’ is the most used for centuries to improve memory and traded and consumed around the world, cognition. We have previously shown that the and it is necessary to find news genotypes water extract of CA leaves (CAW) can enhance that compete in quality with Hass variety. cognitive function in the Tg2576 mouse model In México coexist several races and diverse of Aβ toxicity and in wild type old mice. Major species of Persea, which shows the diversity known constituents are triterpenoids and and importance of this crop. The aim of this caffeoylquinic acids. We have investigated the study was to analyze the close relations and underlying mechanisms of the effects of CAW differences in chemical composition in the on healthy aged mice brain, and the active pulp and skin of three races of avocado and compounds in CAW that contribute to this two species of Persea. A metabolomic analysis effect, by using liquid chromatography-tandem based on NMR-1H was done It was studied mass spectrometry (LC-MS/MS). So far we have three genotypes of the West Indians race, discovered that CAW increases brain levels of three of the guatemalan race, seven of the fatty acid amides, and inhibits fatty acid amide Mexican race, P. nubigena, P. schiedeana and hydrolase (FAAH). ‘Hass’. With the data it was done a component To isolate and characterize the active principal analysis and one discriminant analysis compounds, CAW was first subject to liquid- to determine the variability and the correlations liquid extraction. The fractions were tested for of the variables with the differences and close their bioactivity, and the butanol fraction was relations between the genotypes. In pulp, shown to contain all active phytochemicals. the results showed that the mexican race, the This fraction was further separated by silica guatemalan, P. nubigena and ‘Hass’ are very gel column chromatography and gel filtration similar, whereas the West Indians race and chromatography. In each step the fractions P. shiedeana were very different to those. In and partitions were analyzed by TLC/MS or skin, the grouping and separations between LC/MS for structural identification. In house genotypes were very similar. ‘Hass’, a breeding library, commercially available programs and between the guatemalan and mexican race, in on-line sources were used to comprehensively both pulp and skin was close to the mexican analyze the plant metabolites. By taking a race. The West Indians race in both structures multistep separation and analysis approach, a was characterized by its content of aminoacids few structural candidates have been identified and sugars, whereas the mexican genotypes and characterized for their inhibitory effects were distinguished by aromatic compounds. on FAAH. Computational methods have

14 The Phytochemical Society of North America Sunday, August 9, Afternoon Kent Chapman leads the Center for Plant Lipid Research Symposium 2: Phytochemical Lipids at the University of North Texas and Metabolism which focuses on basic and applied aspects of plant lipid Chairs: Aruna Kilaru and Argelia Lorence metabolism. Center scientists [S2-1] Metabolism and functions of use contemporary cellular, N-acylethanolamines in seedling biochemical, and molecular genetic development. approaches to understand how lipids influence Presenter: Kent Chapman, Department of Biological the growth and development of plants. Efforts Sciences, University of North Texas also contribute to the discovery of new Author(s): Kent D. Chapman1, Jantana products and uses for plant derived lipids and Keereetaweep1, Elison B. Blancaflor1,2 their potential public benefit. Kent Chapman 1University of North Texas, Center for Plant Lipid earned a B.A. degree in biology from Lycoming Research, Department of Biological Sciences, College in Williamsport, PA, and then traveled Denton, TX 76203 Tempe, AZ, where he earned a Ph.D. degree in 2Samuel Roberts Noble Foundation, Plant Biology botany (plant cell biology) from Arizona State Division, Ardmore, OK University. After completing his doctoral N-Acylethanolamines (NAEs) are fatty acid degree, Chapman was awarded a 2-year NSF derivatives conjugated to ethanolamine via an postdoctoral fellowship to study plant amide bond. In seeds of most plants including biochemistry at Louisiana State University in Arabidopsis thaliana, NAEs with linolenic (18:3) Baton Rouge, LA. In 1993, he accepted a and linoleic (18:2) acids together make up position as a tenure-track, Assistant Professor more than 70% of the total NAE pool, and they of Biochemistry at the University of North Texas both decline markedly with the progression of (UNT) in Denton, TX. During the last 20+ years, seed germination and seedling development. Chapman has developed an internationally- Evidence indicates that both NAE types recognized research program in plant are metabolized by specific lipoxygenase biochemistry and cell biology at UNT. Research (LOX) isoforms during the course of seedling in the Chapman laboratory is focused mostly in establishment, leading to an array of NAE the areas of cellular signaling pathways and oxylipin metabolites. Specific ethanolamide lipid storage. The Chapman lab has oxylipins of the NAEs are capable of inhibiting contributed more than 100 publications to the seedling development in a tissue-specific primary plant biology and biochemistry manner. Seedling sensitivity to growth literature, and new ideas about the inhibition by ethanolamide oxylipins overlaps a evolutionary conservation of lipid metabolism secondary dormancy stage induced by abscisic and function in eukaryotes have emerged from acid (ABA) that is believed to support survival these efforts. Chapman is co-inventor on six from stresses during seedling establishment. patents (issued or pending). In January, 2014, Collectively, our data suggest that the he took leave from UNT for one year to serve formation of ethanolamide oxylipins may as Program Director at the National Science participate in the coordinate control of seedling Foundation’s Division of Integrative Organismal establishment under unfavorable environmental Systems. Chapman currently holds the title of conditions in cooperation with phytohormone Regents Professor at the University of North and light signaling pathways. Texas and serves as Executive Editor for the journal, Progress in Lipid Research.

August 8–12, 2015 • University of Illinois at Urbana-Champaign 15 [S2-2] Chemical- and lipid-mediated Wisconsin-Madison. He was a Research systemic acquired resistance in plants. Associate with Professor Dan Klessig at the Presenter and Author: Pradeep Kachroo, Boyce Thompson Institute, NY until his Department of Plant Pathology, University of appointment as Assistant Professor at the Kentucky Department of Plant Pathology in September Systemic acquired resistance (SAR) is a highly 2002. desirable form of resistance that protects against a broad-spectrum of pathogens. [S2-3: Neish Award Speaker] Abe Koo, SAR involves the generation of a mobile Department Biochemistry, University of signal at the site of primary infection, which Missouri. Metabolism of plant hormone arms distal portions of a plant against jasmonate: A sentinel for tissue damage subsequent secondary infections. The last and master regulator of stress response. decade has witnessed considerable progress Abraham (Abe) Jeong-Kyu and a number of diverse chemical signals Koo did his undergraduate and contributing to SAR have been isolated and Master of Science studies at characterized. Among these, salicylic acid Korea University in South (SA) functions in parallel to nitric oxide (NO)- Korea. He moved to Michigan and reactive oxygen species (ROS)-derived State University (MSU) in 1999 signaling leading to SAR. Other chemical to do his PhD under Dr. John signals involved in SAR azelaic acid (AzA) and Ohlrogge where he studied lipid metabolism in glycerol-3-phosphate (G3P) function in the higher plants and wrote a dissertation on topics NO-ROS branch of the SAR pathway. The plant of fatty acid trafficking across the chloroplast galactolipids monogalactosyldiacylglycerol membranes. Upon receiving his PhD degree in (MGDG) and digalactosyldiacylglycerol 2004, he joined Dr. Gregg Howe’s group in (DGDG) are also required for SAR and of these Department of Energy-Plant Research DGDG contributes to plant NO as well as SA Laboratory (DOE-PRL) at MSU where he was biosynthesis. In contrast, MGDG regulates the first introduced to the field of jasmonate. biosynthesis of the SAR signals AzA and G3P. During this time he identified enzymes involved Interestingly, replacement of galactose sugar in JA biosynthesis and published papers on with glucose in DGDG is unable to restore SAR long-distance JA signaling. A Phytochemistry in dgd1 plants even though it does rescue review article that he had co-authored with his their morphological and fatty acid phenotypes. postdoctoral mentor was later awarded the Top These results suggest that MGDG and DGDG Five Most-Cited Papers Award (from 2009 are required at distinct steps and function through 2013) by the journal publisher. He was exclusively in their individual roles during the also awarded the Anton Lang Memorial Award induction of SAR. Relationship among recently for Research Excellence by the DOE-PRL (2010) identified mobile inducers of SAR will be and in the same year was promoted to research discussed. assistant professor in the DOE-PRL. In 2012, he Pradeep Kachroo conducted moved to University of Missouri to start his his Ph.D. research on independent group as tenure-track assistant transposable elements in the professor in the Division of Biochemistry. Since rice blast pathogen then he has been engaging in mentoring and Magnaporthe grisea. He leading his own research group as well as received his degree from participating in departmental teachings. He has Maharaja Sayajirao University of served in various committees and also served Baroda, India and from the University of as ad-hoc reviewer for multiple journals including Nature Chemical Biology, Plant Cell,

16 The Phytochemical Society of North America Plant Journal, Plant Physiology, and point of regulation of oil biosynthesis in oil- Phytochemistry. rich mesocarp tissue most likely occurs at the level of source and not sink. Overall, this study [S2-4] Transcriptome Analysis of provides a foundation for functional genomics Avocado Mesocarp Reveals Key Genes required to direct metabolic engineering efforts Necessary to Improve Oil Yield to enhance avocado oil yield. Presenter(s): Aruna Kilaru, East Tennessee State University [S2-5] Identification, and Heterologous Author(s): Xia Cao, Bayer CropSciences, Parker B. Expression Analysis of Avocado DGAT1 Dabbs, East Tennessee State University, Ha-Jung and DGAT2 Sung, East Tennessee State University, Mahbubur Presenter(s): Md Mahbubur Rahman, East Tennessee Md. Rahman, East Tennessee State University, State University at Johnson City Keithanne Mockaitis, Indiana University, John B. 1 3 1,2 Ohlrogge, Michigan State Univer Author(s): M. M. Rahman , J. Shockey , A. Kilaru 1Department of Biological Sciences, 2Department Avocado is an economically important of Biomedical Sciences, East Tennessee State crop with ~70% oil in its fruit tissue, which University, Johnson City, TN 37614, USA, 3USDA- is an essential component of human diet. ARS, New Orleans, LA 70124, USA The steady increase in global demand for The neutral lipid triacylglycerol (TAG) is the avocado production (9%/year) has drawn main storage lipid in plants. When stored in attention to the importance of understanding seeds, TAG provides the carbon and energy the genetic regulation of triacylglycerol source during germination. There is significant (TAG) accumulation. Using RNA-seq human nutritional demand for vegetable approach, mesocarp-specific regulation and oil, but its use in production of renewable biosynthesis of TAG in developing avocado biomaterials and fuels has intensified the need fruit was analyzed. During the period of TAG to increase oil production. In plants, the final accumulation in the mesocarp, an increased and committed step in TAG biosynthesis is expression was noted for genes mostly catalyzed by diacylglycerol acyltransferases associated with hexose metabolism in plastids, (DGAT) and/or a phospholipid: diacylglycerol including pyruvate kinase, relative to cytosol, acyltransferases (PDAT). Both DGAT and which is likely associated with the need for PDAT contribute to seed TAG biosynthesis higher pyruvate flux directed toward plastid in an independent or overlapping manner, fatty acid synthesis. A corresponding increase depending on the species. However, in in expression for plastidial fatty acid synthesis nonseed tissues such as mesocarp of avocado, genes was also noted but not for TAG assembly the regulation of TAG biosynthesis is not genes. Additionally, WRINKLED1 (WRI1), a well-studied. Based on the transcriptome data regulatory element typically associated with of Persea americana it is hypothesized that seed oil biosynthesis, was also highly expressed both DGAT and PDAT are likely to catalyze in oil-rich mesocarp of avocado, along with the conversion of diacylglycerol to TAG. In two other isoforms of WRI. Transcriptomics this study, putative DGAT1 and DGAT2 were also revealed that multiple acyltransferases that identified and comprehensive in silico analyses participate in rate-limiting step in TAG synthesis were conducted to determine the respective might be active concomitantly in mesocarp to start codons, full-length coding sequences, achieve higher levels of TAG accumulation. transmembrane domains, predicted protein Similar observations were previously made structures and phylogenetic relationships with with transcriptome analysis of oil-rich seed and other known DGATs. These data reveal that the non-seed tissues. Together these data suggest putative DGATs of a basal angiosperm species a ubiquitous role for WRI1 and that a major retain features that are conserved not only

August 8–12, 2015 • University of Illinois at Urbana-Champaign 17 among angiosperms but also other eukaryotes. [S2-7] Monoterpene acid glucose esters For further biochemical characterization, the in Eucalyptus foliar oil glands avocado DGATs were expressed in a TAG- Presenter(s): Samiddhi Lankani Senaratne, The deficient yeast strain and lipotoxicity rescue University of Melbourne, Australia assays were conducted. The complementation Author(s): Samiddhi L. Senaratne, Jason Q. D. of this yeast strain confirmed enzyme activity Goodger, Ian E. Woodrow, School of BioSciences, and supported the possible role of both The University of Melbourne, Victoria 3010, avocado DGATs in TAG biosynthesis. Future Australia studies will be focused on determining the Monoterpene acid glucose esters (MAGEs) substrate specificity of DGAT and its role, are a group of non-volatile plant secondary relative to PDATs in TAG biosynthesis in metabolites, composed of a monoterpene avocado mesocarp. acid esterified to glucopyranose. A growing number of MAGEs have been isolated from [S2-6] Wheat waxes revisited: different plant species, but they appear to composition and biosynthesis be particularly prevalent in trees of the genus Presenter(s): Reinhard Jetter, University of British Eucalyptus. Eucalyptus is rich in species Columbia diversity and eucalypt leaves are well known Author(s): R. Jetter, University of British Columbia, R. for the presence of a vast range of secondary Racovita, University of British Columbia, A. Aharoni, metabolites including their characteristic Weizmann Institute of Science, Z. Wang, Northwest terpene essential oils. Non-volatile resinous A&F University materials including MAGEs co-occur with the Plant leaves, stems, fruits and flowers are volatile essential oil components of eucalypts in covered by a skin consisting of the polyester sub-dermal embedded foliar secretory cavities, cutin and waxes. The latter are mixtures of commonly known as oil glands. Despite the homologous aliphatics that perform a number high species diversity and the predominance of crucial functions in protecting the plant of MAGEs, relatively few species have been against biotic and abiotic stress. Although studied in great detail, particularly with respect the waxes of various wheat cultivars had to the chemistry of the non-volatile compounds been analyzed before, many compounds in in their foliar oil glands. Our group conducted a the mixtures remained unidentified. Here, survey of 30 Eucalyptus species from subgenus comprehensive analysis of leaf and stem Symphyomyrtus and subgenus Eucalyptus waxes of wheat cv. Bethlehem revealed more to determine the presence and ubiquity of than 200 compounds, many of them with MAGEs in their leaf glands. A number of novel structures. Based on the characteristic known MAGEs were identified based on their isomer and homolog patterns, biosynthetic characteristic mass fragmentation patterns pathways leading to the formation of various using ESI-LCMS. In addition, several novel compound classes could be hypothesized. To MAGEs were also present in some species. test these models, diverse candidate genes The results show that MAGEs are the most were characterized through expression studies, abundant non-volatile compounds in eucalypts subcellular localization, mutant chemotyping in the subgenus Symphyomyrtus, but this and heterologous expression in various hosts. appears not to be the case in subgenus Several enzymes involved in different wax- Eucalyptus. The distribution pattern of MAGEs forming pathways were identified, and their in the two subgenera is likely to assist in the (partially redundant) roles will be discussed. identification of taxonomical relationships in the genus Eucalyptus. Moreover, the discovery of new non-volatiles in the genus may help inform biosynthetic studies and possibly open

18 The Phytochemical Society of North America up avenues for the development of novel a catalytic switch on enzyme’s dependence on natural products. K+ for activation, while activity was optimum at pH 7.5 for both enzymes. The extent of [S2-8] K+ Dependent and K+ conformational changes, stability and binding Independent Asparaginase from affinities of the enzyme as a result of the point Common Bean (Phaseolus vulgaris): mutation is also being investigated. This study Mechanism of Activation by K+ seeks to elucidate the mechanism of catalytic Presenter(s): Ebenezer Ajewole, University of activation of the plant-type asparaginase by Western Ontario K+. Author(s): Ebenezer Ajewole1,2, Agnieszka Pajak2, Mariusz Jaskolski3,4 and Frédéric Marsolais1,2 1Department of Biology, University of Western Ontario, London, ON, N6A 5B7, Canada 2Genomics Monday, August 10, Morning and Biotechnology, Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Symposium 3: Phytochemical Canada, London, ON, N5V 4T3, Canada Biosynthesis 3Center for Biocrystallographic Research, Institute of Bioorganic Chemistry, Polish Academy of Sciences, Chairs: Li Tian and De-Yu Xie Poznan, Poland 4Department of Crystallography, Faculty of Chemistry, A. Mickiewicz University, [S3-1] Regulation of sesquiterpene Poznan, Poland biosynthesis in plants. Asparagine is broadly known to be the major Presenter(s): Xiao-Ya Chen, Institute of Plant Physiology and Ecology, Shanghai Institutes for nitrogen (N) transport and storage form in Biological Sciences, Chinese Academy of Science most plants, especially legumes. Catabolism Author(s): Xiao-Ya Chen1,2, Chang-Qing Yang1 of L-asparagine provides N for protein 1 biosynthesis in developing plant tissues. The National Key Laboratory of Plant Molecular Genetics, Institute of Plant Physiology and Ecology, enzyme, L-asparaginase, is responsible for Shanghai Institutes for Biological Sciences, Chinese the catalytic deamidation of L-asparagine Academy of Sciences, Shanghai 200032, China into L-aspartate and ammonia, which is 2 Plant Science Research Center, Shanghai Chenshan necessary for further transformation into Botanical Garden, Shanghai 201602, China other amino acids. Previous studies showed Plant synthesizes a wealth of secondary that there are two isoforms of the hydrolytic metabolites, among which terpenoids form enzyme (potassium dependent - PvASPG1 the largest group. Sesquiterpenes, derived and potassium independent asparaginase - from the 15-carbon farnesyl diphosphate, PvASPG-T2) as a result of their K+ requirements play important roles in plant adaptability for activation. The crystal structure of the to environments, particularly in protecting potassium dependent asparaginase (PvASPG1) plants from herbivores and pathogens. We suggested that Ser118 in the activation loop are interested in sesquiterpene biosynthesis of PvASPG1 plays a critical role in coordinating in cotton (Gossypium spp.), Arabidopsis potassium for activation; however, this amino and some medicinal plants, and the role of acid residue is replaced by an isoleucine in phytochemicals in plant-insect interactions and PvASPG-T2. Reciprocal mutants of the enzymes co-evolution. were created by site directed mutagenesis and the effect of the amino acid residue substitution The phytohormone jasmonate (JA) plays on the kinetic parameters, optimum pH and an important role in regulating secondary K+ dependence of the wild type and mutant metabolism. In Arabidopsis the bHLH enzymes were studied, both in the presence transcription factor MYC2 is a key component and absence of potassium. There appears to be of JA signaling pathway. Interestingly, we

August 8–12, 2015 • University of Illinois at Urbana-Champaign 19 found that, in Arabidopsis inflorescence, MYC2 secondary metabolites in plant-insect integrates both gibberellin (GA) and JA signals interactions, plant-mediated RNAi technology in regulation of expression of terpene synthase for insect pest control, and cotton fiber genes, leading to inducible biosynthesis development with emphasis on key and release of volatile sesquiterpenes transcription factors. Chen was elected and monoterpenes. Another group of JA- member of Chinese Academy of Sciences in responsive transcription factors, AP2/ERF, 2005, member of The World the Academy of is also involved in regulating sesquiterpene Science (TWAS) in 2008, and received HLHL biosynthesis in Artemisia annua. Foundation Prize for Scientific and Technological Progress in 2008. He currently In addition to response to environmental serves as Editor-in-Chief for Science Bulletin factors, secondary metabolism also changes and Associate Editor for Plant Biotechnology during plant development and growth. Journal. Our recent study of Pogostemon cablin demonstrated that the miR156-targeted SPLs [S3-2] Valeriana officinalis as a novel directly regulate sesquiterpene synthase genes, platform for plant natural product drug linking the plant age cue to the elevated discovery. accumulation of patchouli oil in mature plant. Over-expression of MIR156 gene nearly Presenter: Joe Chappell, Department of Plant & Soil Science, University of Kentucky abolished and, on the contrary, expression of Authors: Santosh Kumar1, Scott Kinison1, Eric Nybo1, a miR156-resistant SPL gene promoted the Vincent Ricigliano2, Christopher Brooks2, Dianella G. biosynthesis of sesquiterpenes. This provides Howarth2, Joe Chappell1 a strategy to engineer plant for accelerated 1Dept. of Pharmaceutical Sciences, University of growth and enhanced production of secondary Kentucky, Lexington, KY metabolites simultaneously. 2Dept of Biological Sciences, St. John’s University, *We thank Zong-Xia Yu, Gao-Jie Hong and Ying-Bo Jamaica, NY, Mao for their contributions. Valerian is a nutraceutical preparation from Xiao-Ya Chen is Professor at the roots of Valeriana officinalis that is the Institute of Plant Physiology commonly recommended for relief of tension, and Ecology, Shanghai anxiety and insomnia. The greatest biological Institutes for Biological efficacy of valerian has been correlated with Sciences (SIBS), Chinese freshly harvested and carefully dried root Academy of Sciences. He preparations, and with the iridoid alkaloid and currently serves as President of sesquiterpene content of these preparations. Chinese Society of Plant Biology and Director The valepotriates are epoxyiridoid esters with of Shanghai Chenshan Plant Science Research the dominant species being valtrate. Because Center. He obtained B.Sc. (1982) from Nanjing of putative instability and water insolubility of University, China, and Ph.D. (1985) from the valepotriates, some investigations have Reading University, UK. Dr. Xiao-Ya Chen is suggested that the sesquiterpene compounds interested in plant secondary metabolism, are more important for the biological activity cotton biology and biotechnology. He has of valerian. To determine which chemical made important contributions in the components of V. officinalis are important for its investigation of biosynthesis of plant biological activities, we have developed several terpenoids (cotton, Arabidopsis and selected important capabilities and tools to support medicinal plants), transcription factors and this effort. First, we have developed and hormone signaling pathways regulating relied upon transcriptomic and metabolomic secondary metabolism, the role of plant resources to identify the genes encoding for

20 The Phytochemical Society of North America these unique biosynthetic capabilities. Second, the GT enzymes themselves have significant the methodology for genetic engineering hairy impact on the compounds found. We have root cultures of V. officinalis having diverse characterized several enzymes isolated from chemical profiles has been developed. Third, grapefruit leaf tissue and expressed from we have been working on the development grapefruit putative GT clones. One of the of a novel test platform for assessing the clones codes for a flavonol-specific O3- -GT, anxiolytic activity of the various engineered CpF3OGT. Unlike many flavonoid 3-O-GTs, this hairy root culture lines. Progress in all of these enzyme does not glucosylate anthocyanidins. areas will be presented. There are some reports of more promiscuous flavonoid GTs from other plants, including Joe Chappell’s laboratory is those that add sugars to multiple positions. dedicated to understanding the In order to study CpF3OGT structure and mechanisms that plants use to function, we have used bioinformatics and make the dizzying array of modeling approaches to identify residues for terpene/isoprenoid site-directed mutagenesis to test hypotheses compounds. For many years, related to acceptor substrate specificity and and like many laboratories, we regiospecificity. Over 30 single, double, focused our attention on how plants regulate triple, and more extensive mutants have been the biosynthesis of antimicrobial terpene-based synthesized. Characterization of the mutant phytoalexins. Our interests have expanded proteins is ongoing, however the majority have from there. Our work utilizes a wide range of been tested and experimental results used to experimental strategies including genetic refine structural models. engineering, structure-function comparisons of genes and proteins, and cross-comparisons Cecilia McIntosh earned her between many different plants and other Ph.D. from the University of organisms. South Florida in 1990 with Richard Mansell and was a [S3-3] Structure and function of postdoctoral research fellow flavonoid glucosyltransferases: using a with Dr. David Oliver at the specific grapefruit enzyme as a model. University of Idaho (1990-93). Presenter and Author: Cecilia A. McIntosh, She joined the faculty of the East Tennessee Department of Biological Sciences, School of State University Department of Biological Graduate Studies, and Institute for Quantitative Sciences in 1993 where she is now a full Biology, East Tennessee State University, Johnson professor. She served as a program director at City, TN 37614, USA the National Science Foundation from 2003-04, Flavonoids are a major group of ubiquitous and was named Dean of ETSU’s School of plant natural products and the majority are Graduate Studies in 2006. With a lifelong found natively in glycosylated forms. They interest in phytochemistry and plant natural have significant roles in the plant life cycle and products, her research program at ETSU has also have impact on consumer acceptance focused on different aspects of flavonoid and health properties of food crops and biosynthesis and metabolic regulation using processed products. Secondary product Citrus paradisi (grapefruit) as the model system. glycosyltransferases are key enzymes in the This research has been funded from the U.S. biosynthesis of the compounds that accumulate Department of Agriculture and the U.S. in plant tissues. Results from our studies to National Science Foundation. She has served elucidate the dynamics of flavonoid metabolism on multiple advisory boards including the and regulation in grapefruit revealed that, Arkansas ASSET external advisory board and in addition to other factors, properties of on the technical advisory board for the

August 8–12, 2015 • University of Illinois at Urbana-Champaign 21 Arkansas EPSCoR “P3” project from 2008-13. with recombinant enzyme characterization She is a founding fellow of ETSU’s Institute for identified a legume-specific monophyletic Quantitative Biology. In addition to her clade that contains PDH enzymes. Kinetic research, she is extensively involved in K-12 analyses revealed that soybean and Medicago student outreach and professional PDHs specifically use prephenate but not development for graduate students and faculty. arogenate substrate. Unlike previously- She has been active in the PSNA, serving on characterized plant ADHs, both legume PDHs several annual meeting committees, the awards were insensitive to Tyr regulation and localized committee, multiple terms on the advisory to the cytosol. Our study provides molecular board, and as treasurer. She was PSNA evidence for the diversification of the primary president during its 50th anniversary year. metabolic Tyr pathway, likely to support the chemodiversity that exists downstream of HPP [S3-4] Diversification of Tyrosine and Tyr in different plant species. Biosynthetic Pathways in Plants: Non-Plastidic, Tyrosine-Insensitive [S3-5] Evolution of chemical diversity in Prephenate Dehydrogenases in Legumes plants Presenter(s): Hiroshi A. Maeda, Department of Presenter and Author: Bernd Markus Lange, Botany, University of Wisconsin-Madison Washington State University Author(s): Hiroshi A. Maeda1, Craig Schenck1, Siyu The ability to sequester secondary (or 1 Chen specialized) metabolites and defense proteins 1Department of Botany, University of Wisconsin- in secretory structures was a critical adaptation Madison, WI 53706 USA. that has shaped plant-herbivore and plant- Tyrosine (Tyr) is an essential aromatic amino pathogen interactions for hundreds of millions acid synthesized de novo by plants and of years. Secretory structures in terrestrial plants microbes. Besides being a protein building appear to have first emerged as intracellular oil block, in plants Tyr and a Tyr-pathway bodies in liverworts. In vascular plants, internal intermediate 4-hydroxyphenylpyruvate (HPP) secretory structures, such as resin ducts and serve as key precursors of a diverse array laticifers, are usually found in conjunction with of natural products, such as cyanogenic vascular bundles, while subepidermal secretory glycosides, betalains, tocopherols (vitamin E), cavities and epidermal glandular trichomes plastoquinone, and isoquinoline alkaloids. Tyr generally have more complex tissue distribution biosynthesis can occur via two intermediates, patterns. An overview of the evolution of these arogenate and HPP. Arogenate dehydrogenase secretory structures, and the chemical diversity (ADH) and prephenate dehydrogenase (PDH) found within them, will be presented. Although are key enzymes for the arogenate and HPP particular emphasis is placed on describing the pathways, respectively, which are generally evolution of pathways leading to terpenoids, feedback regulated by Tyr. Most plants the emergence of other metabolites is assessed only have plastidic ADH activity and likely as well to outline metabolic capabilities of synthesize Tyr using the arogenate route within different plant lineages. Online resources to the plastids, whereas both ADH and PDH capture and interrogate plant chemical diversity activity was detected in some legume species. and biosynthetic pathways will be assessed as However, the genes encoding enzymes well. responsible for the plant PDH activity are Recent publications on the topic: unknown. Here we report identification and biochemical characterization of PDH enzymes • Turner G.W., Lange B.M. (2015) from soybean and Medicago. Comparative Ultrastructure of grapefruit secretory cavities genomics and phylogenetic analyses together and immunocytochemical localization of

22 The Phytochemical Society of North America (+)-limonene synthase. Int. J. Plant Sci., in candidate HT biosynthetic genes. Our results press. from transcriptome sequencing, pomegranate hairy root culture, and the biochemical • Lange B.M. (2015) The evolution of plant properties of candidate HT biosynthetic secretory structures and emergence of enzymes will be presented. terpenoid chemical diversity. Annu. Rev. Plant Biol. 66, 139-159. [S3-7] Structural modification and • Fischedick J.T., Johnson S.R., Ketchum increase of lignin in Oryza sativa for R.E.B., Croteau R.B., Lange B.M. (2015) biomass refinery NMR spectral search module for Spektraris, Presenter(s): Toshiaki Umezawa, Kyoto University an online resource for plant natural product Author(s): Yuri Takeda, Kyoto University, Taichi identification—taxane diterpenoids from Taxus Koshiba, Kyoto University, Yuki Tobimatsu, Kyoto x media cell suspension cultures as a case University, Masaomi Yamamura, Kyoto University, study. Phytochemistry 113, 87-95. Masahiro Sakamoto, Kyoto University, Toshiyuki Takano, Kyoto University, Shiro Suzuki, Kyoto • Johnson S.R., Lange B.M. (2015) Open-access University, Takefumi Hattor metabolomics databases for natural product Lignin has long been considered recalcitrant in research: present capabilities and future the polysaccharide utilization processes such potential. Frontiers Bioeng. Biotechnol. 3, 22. as pulping and bioethanol fermentation. Lignin is, however, a potential feedstock for aromatic [S3-6] Investigations on early steps of products. Hence, structural modification to hydrolyzable tannin biosynthesis produce lignins that are suitable for lignin Presenter(s): Li Tian, University of California, Davis utilization as well as higher lignin production Author(s): Li Tian, Nadia N. Ono, Xiaoqiong Qin and in biomass plant species are important Alexander E. Wilson, University of California, Davis breeding objectives. Herein, we aimed to Hydrolyzable tannins (HTs) are structurally develop transgenic rice plants which have diverse polyphenolic compounds derived lignin with modified structures by the use of from a shikimate pathway intermediate. This metabolic engineering of lignin biosynthesis. group of metabolites has been studied for their We produced a transgenic rice plant in function in plant-plant interactions, defensive which Oryza sativa ferulate 5-hydroxylase 1 role against insects and herbivores, and impact (OsF5H1 or OsCAld5H1) was downregulated on soil nutrient profiles. More recently, much by RNAi technique. The OsCAld5H1-knock attention has been given to these compounds down did not significantly alter the total lignin due to their health promoting activities in depositions but severely decreased syringyl humans. Although previous biochemical studies (S) lignin content in rice cell walls; reduced have outlined the biosynthetic steps leading S lignin levels in the transgenic lines were to the production of HTs, the biosynthetic compensated with increased guaiacyl (G) and genes and enzymes of the pathway have yet p-hydroxyphenyl (H) lignin levels. Lower S/G to be identified. We chose pomegranate as ratio contributes an increase of heating values our model system for interrogation of the of biomass, because G lignin has higher carbon HT biosynthetic pathway. Our transcriptomic content than S lignin. In addition, we produced sequencing of pomegranate fruit peel, a transgenic rice plants with increased lignin tissue enriched in HTs, revealed multiple content. We selected several genes which were candidate genes that are potentially involved predicted to be involved in lignin biosynthesis. in HT formation. We have also established These genes were overexpressed under the a pomegranate hairy root culture system in control of CaMV35S promoter in rice, and we our lab that allows genetic characterization of obtained transgenic rice T1 lines with about

August 8–12, 2015 • University of Illinois at Urbana-Champaign 23 1.5-fold lignin contents in culms compared to The uniquely complex behavioral repertoire control wild-type plants, which was estimated of A. mellifera may compensate for this to contribute about 7% increase of heating deficit via a “social detoxification system,” value. This strategy may be applicable to lignin whereby collective behaviors may reduce upregulation in large-sized grass biomass the phytochemical complexity of the diet. plants, such as Sorghum, Miscanthus and Behavioral mechanisms that may reduce the Erianthus, which are suitable for direct fuel diversity and quantity of toxins requiring utilization of biomass. enzymatic transformation include selective foraging, toxin dilution by mixing nectars and pollens from multiple sources, maintenance of high hive temperatures to promote toxin Monday, August 10, Afternoon breakdown, enzymatic processing of honey, and culturing fermentative microbes to convert Symposium 4: Chemical Ecology I pollen into beebread. Remarkably, many Chairs: Dorothea Tholl and Massuo Kato of these mechanisms resemble processing techniques used by humans to reduce the toxic [S4-1] Honey bees as phytochemists: phytochemical content of plant-based foods. how pollen and nectar are converted into beebread and honey May Berenbaum, Ph.D. has been on the faculty of the Presenter and Author: May R. Berenbaum, Department of Entomology, University of Illinois at Department of Entomology at Urbana-Champaign, Urbana, IL 61801-3975 the University of Illinois at The western honey bee (Apis mellifera) and Urbana-Champaign since 1980, its congeners are unusual among pollinators serving as head since 1992 and in that they rarely consume raw pollen and as Swanlund Chair of nectar; rather, they process nectar into honey Entomology since 1996. She is known for and pollen into beebread, which are stored elucidating chemical mechanisms underlying in the hive and shared among nestmates. interactions between insects and their This processing allows honey bees to convert hostplants, including detoxification of natural raw materials into forms that can be stored and synthetic chemicals, and for applying in large quantities for long periods of time ecological principles in developing sustainable without spoilage. The perennial nature of management practices for natural and honey bee colonies necessitates collecting agricultural communities. Her research, pollen and nectar over many months from supported primarily by NSF and USDA, has a broad diversity of successively blooming produced over 230 refereed scientific flowers, exposing them to hundreds of publications and 35 book chapters. A member different phytochemicals. Moreover, by of the National Academy of Sciences, she has concentrating nectar to make honey, reducing chaired two National Research Council its water content from about 90 percent to committees, the Committee on the Future of 18-20 percent, honey bees also increase the Pesticides in U.S. Agriculture (2000) and the relative concentrations of phytochemicals Committee on the Status of Pollinators in North in their food. Despite the diversity of floral America (2007). Devoted to teaching and sources utilized to support the eusocial life fostering scientific literacy through formal and style and the concomitant exposure to a informal education, she has authored numerous tremendous diversity of phytochemicals, the magazine articles and six books about insects honey bee genome is markedly reduced in for the general public. She graduated summa terms of its inventory of detoxification genes. cum laude, with a B.S. degree and honors in biology, from Yale University in 1975 and

24 The Phytochemical Society of North America received a Ph.D. in ecology and evolutionary behavioral, and electrophysiological studies biology from Cornell University in 1980. mostly with the vector of West Nile virus in California, the southern house mosquito, [S4-2] Mosquito odorant receptor for the Culex quinquefasciatus. Here, we report on a insect repellent DEET and plant-derived DEET receptor, which is also sensitive to plant- semiochemicals. derived semiochemicals. Presenter and Author: Walter S. Leal, Department Walter Leal’s laboratory of Molecular and Cellular Biology, University of California, Davis, CA 95616 USA research is aimed at unraveling the molecular mechanisms that Insect repellents have been used since make the insect’s olfactory antiquity as prophylactic agents against system so sensitive and diseases transmitted by mosquitoes and other selective. Insect prominence arthropods, including malaria, dengue fever, among other animals is due in and encephalitis. They evolved from smoke large part to a key physiological element for generated by burning plants (e.g.: lemon their survival and reproduction—a refined gum) and topical applications of essential oils olfactory system. Olfaction is orchestrated at (e.g: lemon eucalyptus extract) into repellent various levels starting with reception of substances, including those isolated from odorants at the periphery, processing and plants (e.g.: p-menthane-3,8-diol, PMD) integration of olfactory and other sensory and a broad-spectrum synthetic repellent modalities in the brain, and ultimately N,N-dimethyl-3-methylbenzamide (DEET), translation of olfactory signal into behavior. which was discovered in the early 1940s from Thus, the cornerstone of a sophisticated a screening of thousands of compounds. olfactory system is the ability of the insect’s Thereafter, other synthetic repellents have peripheral system to selectively detect and been developed (e.g: IR3535, 3-(N-acetyl-N- rapidly inactivate minute amounts of odorants. butyl)aminopropionic acid ethyl ester), but Reception of odorants is mainly mediated by DEET remains the most widely used repellent three olfactory proteins, namely, odorant- substance world-wide. Molecular modeling led binding proteins (OBPs), odorant receptors to the development of picaridin ((RS)-sec-butyl (ORs), and odorant-degrading enzymes (ODEs). 2-(2-hydroxyethylpiperidine), but progress in OBPs are involved in the uptake, transport and drug design has been slow, in part because delivery of odorants to ORs. By using of the unknown mode of action. Although biochemical, electrophysiological, RNAi, and it is well established that DEET acts on the kinetic studies we have demonstrated that insect’s gustatory and olfactory systems, its OBPs are essential for the sensitivity of moth’s mode of action as an odorant was a matter of olfactory system. In collaborations with considerable debate, with two dichotomous structural biologists, including UC Davis hypotheses. One school suggests that DEET colleagues, we study the molecular modulates the activity of many odorant mechanisms of odorant binding, release, and receptors (ORs), whereas there is growing transport, including novel, pH-dependent evidence suggesting that repulsion is triggered conformational changes in moths and by excitatory responses of odorant receptor mosquitoes. By comparative kinetic studies of neurons and that OR are involved. As adult odorant degradation by recombinant and mosquito ORs remained elusive, it has been native enzymes we have demonstrated for the postulated that the ionotropic receptor IR40a first time that ODEs are involved in the fast could account for the widespread effect of inactivation of odorants. Using bioinformatics DEET olfactory repellency. To thoroughly test and molecular approaches coupled with the these hypotheses, we carried out molecular, Xenopus oocyte recording system we

August 8–12, 2015 • University of Illinois at Urbana-Champaign 25 investigate how ORs contribute to the verticillioides infection, maize plants grown at selectivity of the insect’s olfactory system. 2x[CO2] were capable of restricting pathogen proliferation to comparable levels as maize

[S4-3: Neish Award Speaker] Climate at 1x[CO2]. The adaptation of new disease change and crop phytochemical management strategies, such as priming, may defenses: Potential implications for food be necessary to improve the production and security and food safety. safety of the future maize crop at higher [CO2]. Presenter: Martha Vaughan, USDA, ARS, Peoria, IL Martha Vaughan is a molecular Authors: Martha M. Vaughan, Alisa Huffaker, Eric A. biologist with interdisciplinary Schmelz, Shawn Christensen, Leon Hartwell Allen, training in plant stress Peter E. A. Teal, USDA, ARS, Peoria, IL physiology, defense signaling, Elevated atmospheric carbon dioxide and secondary metabolism. Her concentration ([CO ]) increased maize 2 research focuses on how susceptibility to Fusarium verticillioides stalk climate affects crop-fungal rot. Even though the pathogen biomass pathogen interactions in a manner that accumulated to significantly higher levels at influences downstream mycotoxin production double ambient [CO ] (2x[CO ]), the projected 2 2 and grain contamination. She has expertise in [CO ] concentration to occur at the end 2 evaluating the response of plants to combined of this century, the quantity of fumonisin abiotic and biotic stress factors both above and mycotoxin contaminants were unaltered. belowground. Her work has led to the Combined effects of [CO ] and drought 2 identification, molecular characterization and on pathogen infection were contradictory; functional elucidation of novel defense drought reduced F. verticillioides biomass metabolites. She has contributed to the in maize stalks at ambient [CO ] (1x[CO ]), 2 2 understanding of how climate changes will but increased pathogen biomass in stems effect crop susceptibility to Fusarium at 2x[CO ]. In the absence of the host plant, 2 pathogens and mycotoxins. She has also been 2x[CO ] hindered F. verticillioides growth but 2 involved in the development of methods enhanced fuminison production on media. The necessary to conduct climate change research soluble carbohydrate, starch, protein and fatty including a plant culturing method that enables acid content of the maize stem tissues were long term plant growth at a consistent level of not significantly altered by 2x[CO ]. However, 2 drought. Through the evaluation of phenomena in response to F. verticillioides infection, the that naturally regulate mycotoxin production, accumulation of soluble carbohydrates, total such as phytochemicals, she seeks to identify proteins, fatty acids, MBOA and phytoalexins sustainable and climate resilient strategies to was absent or reduced in maize plants at eliminate mycotoxin contamination in grain and 2x[CO ]. Examination of early JA levels in 2 enhance food safety. response to F. verticillioides infection, indicated that the phytohormone defense response As a born and bred Hokie, Martha received both her BSc (2004) and PhD (2010) from maybe dampened at 2x[CO2]. There was a direct correlation between JA levels and the Virginia Tech. She then conducted postdoctoral levels of defense metabolites, and an inverse research with the USDA-ARS Chemistry Unit correlation with pathogen proliferations. The at the Center for Medical, Agricultural and attenuation of JA signaling and the reduction Veterinary Entomology in Gainesville FL. In of fumonisin produced per unit F. verticillioides 2013, she joined the Bacterial Foodborne biomass are potentially a result of limited free Pathogens & Mycology Research Team at the fatty acid substrate for oxylipin biosynthesis. National Center for Agricultural Utilization By stimulating the defense response prior to F. Research in Peoria, IL to pursue research on

26 The Phytochemical Society of North America the effects of climate change on mycotoxin acid conjugation. Our results suggest that contamination of grain crops. the specialist N. stephensi uses different detoxification pathways when feeding on its [S4-4] Metabolomics of juniper native juniper compared to N. lepida, whereas detoxification in a generalist and both woodrat species use the same pathways specialist mammalian herbivore. when feeding on the native host of N. lepida, J. Presenter(s): Katharina Schramm, University of Utah osteosperma. Author(s): Michele Skopec, Weber State University, James Cox, University of Utah, James Halpert, [S4-5] Detoxification of Maize Chemical University of Connecticut, Denise Dearing, Defenses by Lepidopteran Herbivores University of Utah Presenter(s): Felipe Christoff Wouters, Max Plank Herbivory is common in mammals, yet our Institute for Chemical Ecology understanding of the detoxification processes Author(s): Daniel Giddings Vassão, Michael Reichelt, used by mammals to biotransform plant Katrin Luck and Jonathan Gershenzon, Max Plank secondary compounds (PSC) is limited. Institute for Chemical Ecology, and Matthias Erb, Specialist herbivores are thought to have Universtity of Bern evolved unique detoxification mechanisms In order to avoid or overcome damage to process high levels of PSC in their diets. by herbivores, grasses (Poaceae) such as We explored this hypothesis by comparing rye, wheat, and maize rely on a family of the detoxification metabolite patterns of a indole-derived chemical defenses called specialist and a generalist herbivore from benzoxazinoids (BXDs). These compounds the genus Neotoma. Stephen’s woodrat, N. are stored in plant cells as stable glucosides stephensi, is a specialist on one-seeded juniper and, upon herbivory, are converted into toxic (Juniperus monosperma). The desert woodrat, aglucones by the action of plant glucosidases. N. lepida, is a generalist across its range; yet However, some herbivores evolved adaptations populations in the Great Basin often feed on that enable them to feed on well-defended a diet predominantly made up of juniper (J. plants, leading to the emergence of some osteosperma). While both woodrat species are species as serious agricultural pests. Although naturally exposed to high levels of terpenes BXDs biosynthesis in plants is well studied, little (the predominant class of PSC in juniper) the is known about their metabolism in insects. In terpene profiles and quantities differ between this context, we aim to better understand the the two juniper species. strategies used by lepidopteran herbivores to avoid toxicity from BXDs. Individuals from both woodrat species were fed diets of each juniper in a cross-over Comparing lepidopteran herbivores feeding design. Urine, collected over a 24h period, on maize, we observed that Spodoptera was extracted and analysed in an untargeted frugiperda, S. littoralis and S. exigua excrete metabolomics approach by both GC-MS the non-toxic DIMBOA-Glc (the most abundant and HPLC-MS/MS. When feeding on J. BXD in maize) in the frass. We performed in osteosperma, the native diet of N. lepida, vitro assays with gut tissue preparations and both woodrat species exhibited very similar confirmed that this compound is result of patterns of metabolites. On J. monosperma, enzymatic reglucosylation of DIMBOA by the native diet of N. stephensi, the profiles were insect. Surprisingly, the compound produced distinctly different between species. These by the insect is (2S)-DIMBOA-Glc, an epimer results suggest that the species used different of the plant-derived (2R)-DIMBOA-Glc. We metabolic pathways to biotransform J. demonstrate that plant glucosidases are not monosperma-PSC. Species-specific differences able to hydrolyze the insect metabolite, thus also existed with respect to glucuronic preventing it from exerting its toxicity after it

August 8–12, 2015 • University of Illinois at Urbana-Champaign 27 is transformed by the insect. Therefore, we leafworm) and Mamestra brassicae (cabbage identified the reglucosylation of DIMBOA moth) was investigated from hatching until with inversion of stereochemical configuration adult emergence while the larvae were reared as a detoxification mechanism employed by on Arabidopsis thaliana, accession Col-0, Spodoptera species. Moreover, we present our and three different glucosinolate-deficient ongoing work on identifying the UGT enzymes mutants. We found that both glucosinolate relevant for BXD detoxification inS. frugiperda, types negatively affected larval development based on a transcriptomic approach, and how when fed separately, but in combination their we plan to further explore BXD metabolism effect was significantly stronger. To our surprise, through feeding assays using radiolabeled larvae fed on glucosinolates gave rise to BXDs. Collectively, the elucidation of BXD heavier pupae and adults. Using artificial diets, metabolism in insects clarifies their coevolution we conducted detailed investigations on the with plants and provides targets for innovative effect of the aliphatic 4-methylsulfinylbutyl-ITC approaches in ecological and agricultural (sulforaphane) on the the biochemistry and research. metabolism of S. littoralis larvae. The most typical effect was the decrease of glutathione [S4-6] Toxicity of glucosinolate-derived in midgut tissue and hemolymph, likely isothiocyanates to generalist-feeding due to losses by conjugation to ITC during caterpillars detoxification. As a consequence, the levels Presenter(s): Verena Jeschke, MPI for Chemical of free amino acids were altered, with a sharp Ecology decrease in cysteine leading to reductions Author(s): Verena Jeschke, Emily E. Kearney, in protein content, but an increase in lipids. Jonathan Gershenzon and Daniel G. Vassão, MPI for To learn more about the mechanism of Chemical Ecology glucosinolate toxicity, we are now performing a Glucosinolates are major plant defense comprehensive study of the conjugation of ITC compounds in the order Brassicales (e.g. to the proteome in vivo. cabbage, broccoli, mustard). Upon tissue [1] Schramm K, Vassão DG, Reichelt M, Gershenzon damage caused e.g. by chewing herbivores, J, Wittstock U (2012) Metabolism of glucosinolate- the non-toxic parent glycosides are derived isothiocyanates to glutathione conjugates transformed into toxic isothiocyanates (ITCs) in generalist lepidopteran herbivores. Insect and other metabolites. Still, generalist-feeding Biochemistry and Molecular Biology 42, 174-182. insect herbivores sometimes feed successfully [S4-7] Gene-to-Terpene Landscapes in on glucosinolate-containing plants. Previous Self-pollinating Artemisia annua studies in our group revealed that a proportion of ITCs is metabolized to glutathione Presenter(s): De-Yu Xie, Department of Plant and Microbial Biology, North Carolina State University conjugates; yet, a large amount is excreted in Author(s): De-Yu Xie1, Dong-Ming Ma1, Fatima unmodified form [1]. These unmodified ITCs Alejos1, Zhilong Wang1, Liangjiang Wang2 and Ming- cause decreased rates of growth and delayed An Sun3 development. However, the mode of action 1Department of Plant and Microbial Biology, North of these nucleophilic and lipophilic toxins in Carolina State University, Raleigh, NC 27695, USA, caterpillars is poorly understood. 2Department of Genetics and Biochemistry, Clemson University, Clemson, SC 29634, USA, 3Virginia We compared the effects of Met- and Trp- Bioinformatics Institute, Virginia Tech, VI, USA derived glucosinolates, which lead principally Artemisia annua is an antimalarial medicinal to ITC and non-ITC hydrolysis products, plant that saves more than 100 million of respectively, on two generalist-feeding people annually. Here, we report to use caterpillars in the laboratory. The development an integrated approach of metabolomics, of larvae of Spodoptera littoralis (African cotton

28 The Phytochemical Society of North America transcriptomics and gene function analysis (HMT) in red clover responsible for phaselic to characterize gene-to-terpene scenarios in acid (caffeoyl-malate) accumulation, we different tissues of a self-pollinating variety. transferred this gene to alfalfa. Leaves of the Fifty-one identified terpenes were differentially resulting transgenic alfalfa accumulated mostly produced by leaves and flowers from lower p-coumaroyl and feruloyl-malate rather than to higher positions. 18,871 contigs were the desired caffeoyl derivative. Simultaneous assembled from sequences of six cDNA down-regulation of endogenous caffeoyl- libraries. Sequence mining identified forty- CoA O-methyl transferase (CCOMT) resulted seven genes that were mapped and integrated in increased accumulation of phaselic acid to networks composed of the artemisinin, non- in alfalfa leaves and even higher levels in amorphadiene sesquiterpene, monoterpene, stems, possibly due to the relatively higher triterpene, 2-C-methyl-D-erythritol 4-phosphate levels of phenylpropanoid pathway enzymes, (MEP) and mevalonate pathways. Manipulation particularly HCT and C3’H, in this tissue that of genes significantly altered both individual would normally be involved in monolignol pathway activities and genome-wide terpene biosynthesis. The levels of phaselic acid profiles. This study provides useful gene-to- produced in these plants should be sufficient terpene landscape data for rational metabolic to significantly impact post-harvest proteolysis. engineering of artemisinin. We are currently testing this via in vitro assays and small-scale ensiling experiments. We [S4-8] Engineering alfalfa to accumulate have also identified a gene from red clover useful caffeic acid derivatives and encoding a hydroxycinnamoyl-CoA transferase characterization of hydroxycinnamoyl- responsible for clovamide (caffeoyl-L-DOPA CoA transferases from legumes amide) production; a gene from bean encoding Presenter(s): Michael L. Sullivan, U.S. Dairy Forage a hydroxycinnamoyl-CoA transferase capable Research Center of transferring hydroxycinnamoyl moieties Author(s): Michael L. Sullivan, U.S. Dairy Forage to tetrahydroxyadipic acid isomers; and Research Center and Heather A. Green, U.S. Dairy hydroxycinnamoyl-CoA transferase activities Forage Research Center from bean and perennial peanut capable of Some forages crops, such as red clover, transferring hydroxycinnamoyl moieties to malic accumulate high levels of caffeic acid and tartaric acid, respectively. Characterization derivatives. Oxidation of these o-diphenols of these enzymes should provide insights into to o-quinones by endogenous polyphenol the structure/function relationships of this class oxidases (PPOs) and the subsequent reactions of BAHD transferases. of these quinones (probably with endogenous plant proteases) result in a significant reduction in post-harvest protein degradation. Reduced protein loss in forage crops should lead to more efficient nitrogen utilization in ruminant animal production systems, providing both economic and environmental benefits. One of the most commonly grown legume forage crops, alfalfa, does not accumulate PPO- utilizable phenolics. For this reason, we have been working to engineer biosynthetic pathways for accumulation of caffeic acid derivatives in this crop. Having identified a hydroxycinnamoyl-CoA:malate transferase

August 8–12, 2015 • University of Illinois at Urbana-Champaign 29 Tuesday, August 11, Morning callose deposition. This trade-off indicates a role for BXDs not only as direct toxins but also Symposium 5: Chemical Ecology II as signals for other defensive responses. Chairs: Dhirendra Kumar and De-Yu Xie Herbivores that feed readily on maize foliage must possess some type of metabolic or [S5-1] Metabolic tag in the cornfield: behavioral adaptation to avoid BXD toxicity, Biosynthesis and activation of which is based on hydrolysis of the glucose benzoxazinoids by corn plants and residue by a plant β-glucosidase and reaction deactivation by herbivores. of the resulting aglycone with nucleophilic Presenter: Jonathan Gershenzon, Department of targets in the herbivore. We investigated the Biochemistry, Max Planck Institute for Chemical metabolism of BXDs by the fall armyworm Ecology (Spodoptera frugiperda), a major pest of corn Authors: Vinzenz Handrick, Felipe C. Wouters, and other small grains in North and South Christelle A.M. Robert, Tobias G. Köllner, Daniel America. After BXDs are activated by hydrolysis Giddings Vassão, Georg Jander, Matthias Erb and Jonathan Gershenzon mediated by plant β-glucosidases, the Max Planck Institute for Chemical Ecology, Jena, armyworm larvae were found to re-glucosylate Germany them in their guts using a glucosyltransferase. Boyce Thompson Institute for Plant Research, Although the plant β-glucosidase also present Ithaca, New York in the gut could in theory rehydrolyze and reactivate the new glucoside, the herbivore A major group of phytochemicals in corn actually creates a glucoside of opposite and other grasses are the benzoxazinoids configuration to the plant compound which can (BXDs), a group of indole-derived, hydroxamic no longer be activated by the plant enzyme. acid derivatives that are produced in plants This metabolic trick appears to be the critical as glucosides. The best known of these adaptation allowing the fall armyworm to feed substances, DIMBOA, has a wide range of on maize without negative consequences. biological properties, including anti-insect, antibiotic and allelopathic activities. While After studying biology as an the core pathway of BXD biosynthesis is well- undergraduate at the University studied, little is known about the steps leading of California in Santa Cruz, to a number of derived structures whose Jonathan Gershenzon occurrence is induced by herbivore damage. received his PhD in botany We have discovered a number of genes from the University of Texas in involved in these later steps by quantitative 1984. From 1985 until 1997 he trait locus mapping and transcriptomic worked as a scientist at the Institute for methods, and characterized the corresponding Biological Chemistry, Washington State enzymes after heterologous expression. Among University in Pullman. Since 1997 he is a these are several O-methyltransferases and a Director and Scientific Member at the Max 2-oxoglutarate-dependent dioxygenase. Maize Planck Institute for Chemical Ecology in Jena, inbred lines that are naturally mutated in these Germany, where he heads the Department of genes generally show increased herbivore Biochemistry. He was appointed Honorary damage suggesting that the inducible, late Professor at Friedrich Schiller University Jena in pathway BXD metabolites have roles in 1999. herbivore defense. Curiously, an inactive copy Gershenzon studies the biochemistry of of one of these genes leads to plants that secondary plant metabolites, their mode are more susceptible to caterpillar damage, of action on herbivores, the regulation of but more resistant to aphids due to increased secondary metabolisms in plants and the

30 The Phytochemical Society of North America evolution of pathways. Most of the work in his Eric Schmelz completed his department focuses on two major groups of PhD within the Center for plant defenses: glucosinolates and terpenoids. Insect Science/Plant-Insect Interactions Group at the [S5-2] Inducible small molecule defenses University of Arizona, Tucson. in maize: more than a few surprises. He then carried out his Presenter and Author: Eric Schmelz, Department of postdoctoral and research Biological Sciences, University of California at San scientist studies at the USDA-Agricultural Diego Research Service located at the University of In response to pathogen and insect attack, Florida, Gainesville. He joined the Division of plants produce an array of small molecule Biological Sciences faculty at U.C. San Diego in biochemicals that either directly or indirectly July 2014. function in plant protection. Pathogen inducible transcript profiling in maize(Zea mays) reveals a [S5-3] Signaling function for an abietane large number of terpene synthases, cytochrome diterpenoid in plant defense and P450s, O-methyl transferases, proteases and development. numerous proteins of unknown function. Presenter and Author: Jyoti Shah, Department Long considered absent in maize, high of Biological Sciences, University of North Texas, concentrations of antimicrobial phytoalexins Denton, Texas are now known to predominate at the site of Abietane diterpenoids, which are major infection. As key biosynthetic precursors of constituents of conifer resins, have industrial zealexin class sesquiterpenoids, ZmTps6 and and medicinal applications. They are also ZmTps11 encode b-macrocarpene synthases found in flowering plants. However, their that are broadly elicited by fungi at both function in Angiosperms is poorly understood. the transcript and protein level. With similar We recently co-purified dehydroabietinal regulation, ZmAn2 encodes an ent-copalyl (DA) with a systemic acquire resistance (SAR) diphosphate synthase required for kauralexins, promoting activity that accumulates in the acidic diterpenoid phytoalexins. Outside of vascular sap of Arabidopsis thaliana leaves classical phytoalexin activities, zealexins and [1]. Further experiments demonstrated that kauralexins display inducible accumulation in DA itself was a very potent inducer of plant drying roots and are surprisingly associated immunity against pathogens. Local application with drought protection. Pathogen-induced of picomolar solutions of DA resulted in cyclopenten(an)one oxylipins with structural the systemic induction of salicylic acid (SA) parallels to phytohormones are an additional signaling, thereby conferring enhanced disease class of maize phytoalexins. In addition to fungal growth suppression, 9-lipoxygenase derived cyclopentenones are phytotoxic and display partially overlapping transcriptional activities with jasmonates suggesting additional roles as endogenous signals. Despite being among the worlds largest annually harvested crops, recent findings indicate that many key regulatory and multifunctional secondary metabolite pathways in poaceous species remain largely unknown.

August 8–12, 2015 • University of Illinois at Urbana-Champaign 31 resistance. In our efforts to further understand Foundation and the US Department of DA signaling, we identifiedArabidopsis Agriculture, has centered on signaling in plant thaliana mutants (ida) that are unresponsive to defense against pests, and the engineering of DA. These ida mutants are deficient in SAR. In defense signaling mechanisms for developing addition, several of these ida mutants exhibit a plants that are inherently resistant to pests. delayed flowering phenotype, thus suggesting that in addition to its role in promoting [S5-4] Sarmentine, a natural Piper amide systemic signaling associated with plant herbicide with multiple mechanisms of defense, DA may have a role in promoting action flowering. The effect of DA on flowering is Presenter(s): Franck E. Dayan, USDA-ARS Natural mediated through promoting expression of Products Utilization Research Unit FLOWERING LOCUS D (FLD) and FT, both Author(s): Franck E. Dayan and Daniel K. Owens, positive regulators of flowering, and the USDA-ARS Natural Products Utilization Research simultaneous repression of FLC, an inhibitor of Unit, and Ratnakar Asolkar and Louis Boddy, FLC expression. As opposed to flowering, the Marrone Bio Innovations effect of DA on SAR did not involve FLC and Sarmentine, 1-(1-pyrrolidinyl)-(2E,4E)-2,4- FT, but required FLD [2]. These results suggest decadien-1-one, is a natural product isolated that DA, a newly identified signaling molecule, from the fruits of Piper species. The compound has functions in plant defense as well as has a number of interesting biological development. We propose that DA promoted properties, including its broad-spectrum activity flowering and defense utilize common on weeds as a contact herbicide. Initial studies components leading to FLD expression, highlighted a similarity in response between beyond which the signaling pathway utilized for plants treated with sarmentine and herbicidal promoting SAR and flowering bifurcates [3]. soaps such as nonanoic acid (pelargonic acid). However, little was known about the References mechanism of action leading to the rapid Chaturvedi, R, Venables, B., Petros, R.A., Nalam, V., desiccation of foliage treated by sarmentine. In Li, M., Wang, X., Takemoto, L.J., and Shah, J. (2012) An abietane diterpenoid is a potent activator of cucumber cotyledon discs assays, sarmentine systemic acquired resistance. Plant J. 71, 161-172. induced rapid light-independent loss of Singh, V., Roy, S., Giri, M.K., Chaturvedi, R., membrane integrity at 100 µM or higher Chowdhury, Z., Shah, J., and Nandi, A.K. (2013) concentration, whereas 3 mM pelargonic acid Arabidopsis thaliana FLOWERING LOCUS D is was required for similar effect. Sarmentine was required for systemic acquired resistance. Mol. between 10 and 30 times more active than Plant-Microbe Interact. 26, 1079-1088. pelargonic acid on wild mustard, velvetleaf, Jyoti Shah was born in India, redroot pigweed and crabgrass. Additionally, where he studied Microbiology the potency of 30 µM sarmentine was greatly at University of Bombay stimulated by light, suggesting that this natural (Mumbai) (BSc 1983; MSc product may also interfere with photosynthetic 1985). He started his own processes. This was confirmed by observing a research group in the Division complete inhibition of photosynthetic electron of Biology, Kansas State transport at that concentration. Additional University after his PhD from University of Notre studies on isolated thylakoid membranes Dame (1991) and postdoctoral work at the confirmed that sarmentine acted as a PSII Waksman Institute, Rutgers University. He is inhibitor by competing for the binding site of currently Professor in Biological Sciences at the plastoquinone. This can be attributed in part to University of North Texas. His research, which structural similarities between herbicides like has been funded by the National Science diuron and sarmentine. While this mechanism

32 The Phytochemical Society of North America of action accounts for the light stimulation was evaluated in vivo, by monitoring the of the activity of sarmentine, it does not chlorophyll fluorescence parameterΦ NO account for its ability to destabilize membrane through non-invasive Chl fluorescence imaging in darkness. In this respect, sarmentine has in whole leaves of infected plants. Treatment some structural similarity to crotonoyl-CoA, of ginseng roots with a relatively high dose the substrate of enoyl-CoA reductase, a of ginsenosides prior to planting resulted in key enzyme in the early steps of fatty acid delayed infection by P. irregulare, of both one- synthesis. Inhibitors of this enzyme, such and two-year old ginseng plants. Meanwhile, as triclosan, cause rapid loss of membrane in vitro exposure of P. irregulare to pure integrity in the dark. Sarmentine inhibited the ginsenoside extracts and GSF enhanced and activity of enoyl-CoA reductase, with an I50app altered mycelial growth. While, these results of 18.3 µM. Therefore, the herbicidal activity of do not definitively show that ginsenosides act sarmentine appears to be a complex process as chemoattractants for P. irregulare, they do associated with multiple mechanisms of action. demonstrate that rhizosphere ginsenosides affect the growth pattern of P. irregulare [S5-5] The Chemoattractant Potential in vivo, which can affect the severity of its of Ginsenosides in the Ginseng - P. pathogenicity. irregulare Pathosystem Presenter(s): Dimitre A. Ivanov, Department of [S5-6] Chemical Interaction between the Biology and the Biotron, The University of Western cruciferous phytoalexin camalexin and Ontario Colletotrichum species Author(s): Ivanov, D.A., Georgakopolous, J.R.C., Presenter(s): Chintamani Thapa, Department of Bernards, M.A. chemistry, University of Saskatchewan, SK S7N 5C9, Department of Biology and the Biotron, The Canada University of Western Ontario, 1151 Richmond St. Author(s): Chintamani Thapa, M. Soledade C. N., London, ON, Canada, N6A 5B7 Pedras, Department of Chemistry, University of Ginsenosides, the triterpenoid saponins Saskatchewan, 110 Science Place, Saskatoon, SK, produced by American ginseng (Panax S7N 5C9, Canada quinquefolius L.), have been extensively To defend against pathogens, crucifers studied for their medicinal value, however their (e.g. canola, mustard, cabbage, rutabaga) function in the rhizosphere remains unknown. possess numerous pathways, some of which Similar to other saponins, ginsenosides, include de novo biosynthesis of defense possess antifungal properties against root and metabolites called phytoalexins. Phytoalexins non-root pathogenic fungal species. However, are antimicrobial compounds produced by growth of the ginseng root pathogen Pythium plants under biotic or abiotic stress, but irregulare, is stimulated when exposed to not present under non-stressed conditions. ginsenosides. Presently, the chemoattractant Colletotrichum species have been known to potential of ginsenosides for P. irregulare cause significant yield losses in various crops was evaluated through (1) an in vivo pot due to anthracnose, a fungal disease that experiment that monitored the pathogenicity infects a wide range of plant families, including of P. irregulare toward ginsenoside-treated crucifers. and -untreated one- and two- year old The objective of this project is to investigate ginseng plants and (2) by monitoring the the transformation of the cruciferous affects of a purified total ginsenoside extract phytoalexin camalexin by Colletotrichum (GSF) and pure ginsenosides (Rb1, Re and species. Firstly it was established that some F2) on the growth of the pathogen, in vitro. cruciferous phytoalexins are highly antifungal Disease severity and Time to Infection (TTI) against various Colletotrichum species. Next,

August 8–12, 2015 • University of Illinois at Urbana-Champaign 33 investigation of the metabolism of camalexin in greenhouse studies. Avenanthramides using fungal cultures of C. higginsianum, are phenolic antioxidants demonstrating C. dematium and C. lentis (previously C. several nutraceutical effects in laboratory truncatum) and chemical analyses (HPLC and experiments including: inhibition of nuclear LC-ESI-MS) has demonstrated that camalexin factor kappa beta activation, suppression of was slowly transformed by all species. The the aberrant Wnt/b-catenin signaling pathway chemical structures of the biotransformation in HeLa cells and activation of anti-oxidant products were established after isolation and response element (ARE) associated genes in spectroscopic analyses (NMR, HRMS, UV, IR) vitro. Avenanthramides are also reported to of each compound. The structures of new function as phytoalexins in response to Crown metabolites were confirmed by chemical Rust infection. Presented here are the results syntheses. The antifungal activity of the of application of BTH under field conditions. metabolic products of camalexin established Field trials were conducted in 2013 using that they are less antifungal than camalexin. four application rates on two cultivars of oat Furthermore, camalexin was able to induce the and in 2014 using three application rates production of the fungal metabolite chrysogine on four oat cultivars. The results showed a in C. higginsianum. Altogether, relative to subtle but demonstrable effect of increased cruciferous pathogens, the transformations avenanthramide production in vegetative of camalexin by Colletotrichum species are tissue, mature grain and enhanced resistance substantially slower (e.g. 12 hours vs. 8 days). to Crown Rust infection. Gene expression Hence, based on these results, it is suggested of pathogenesis related protein-1 (PR-1), that Colletotrichum species, unlike cruciferous phenylalanine ammonia lyase (PAL) and pathogens (e.g. Alternaria brassicicola), use hydroxycinnamoyl-CoA:hydroxyanthranilate non-specific enzymes (“house-keeping” hydroxycinnamoyl-CoA transferase (HHT) was enzymes) to metabolize camalexin. These also monitored by qRT-PCR. Both PR-1 and PAL results will be presented and compared with expression were demonstrably up-regulated reported work on the metabolism of camalexin in the field trials, HHT was not. However, by other fungal pathogens. greenhouse experiments showed all three genes to be elicited with 68, 9, and 3 fold [S5-7] Field Application of increases, respectively. Benzothiadiazole on Oat: Effect on Avenanthramide Production and Crown Rust Resistance Presenter(s): Mitchell L. Wise, USDA, ARS, Cereal Tuesday, August 11, Afternoon Crops Research Unit Author(s): Mitchell L. Wise1, Marcus Vinje1, Shawn Symposium 6: Botanical Medicines Conley2 Chairs: Fred Stevens and Dejan Nikoli 1USDA, ARS, Cereal Crops Research Unit, 502 Walnut St., Madison, WI. 53726, USA [S6-1] Development of safe and effective 2Department of Agronomy, University of Wisconsin, botanical dietary supplements for 1575 Linden Dr, Madison, WI. 53706, USA women’s health. Plant defense activators such as Presenter and Author: Richard B. van Breemen, benzothiadiazole (BTH) are known to elicit UIC/NIH Center for Botanical Dietary Supplements Research, University of Illinois College of Pharmacy, the biosynthesis of plant phytoalexins. In 833 S. Wood St., Chicago, IL 60612 oat, BTH treatment has been shown to up- regulate avenanthramide production in To ensure a safe and effective product, both the vegetative tissue and filling grain botanical dietary supplements can be

34 The Phytochemical Society of North America developed in a manner analogous to Catherine Fenselau) in 1985. He carried out pharmaceuticals that involves identification of post-doctoral research in laser desorption mass mechanisms of action and active constituents, spectrometry at Johns Hopkins with Robert chemical standardization based on the active Cotter before joining North Carolina State compounds, biological standardization based University as assistant professor of Chemistry on activity, preclinical evaluation of toxicity and Biotechnology. Until 2010, he served as and potential for drug-botanical interactions, the first Editor-in-Chief of Combinatorial metabolism of active compounds, and finally, Chemistry & High Throughput Screening and is clinical studies of safety and efficacy. Unlike currently on the editorial board of Assay and pharmaceuticals, botanical also require Drug Development Technologies and a authentication, which will be discussed in Regional Editor of Biomedical Chromatography. view of the current controversy regarding Prof. van Breemen received a University Scholar DNA barcoding vs. chemical fingerprinting faculty award from the University of Illinois, a for finished products. Completing these steps 2010 Expert Methods Panel award from the enable the translation of botanicals from the AOAC International for his work on analytical field to safe human use as dietary supplements. methods for dietary supplements, and the 2008 Focused since 1999 on botanicals used by Harvey W. Wiley Award from the AOAC menopausal women as alternatives to hormone International. His research concerns the therapy, the UIC Botanical Center has carried discovery and development of natural products out this systematic approach for a series of as chemoprevention agents and the botanicals including red clover (Trifolium investigation of botanical dietary supplements pratense), black cohosh (Actea racemosa), and as alternatives to hormone therapy for hops (Humulus lupulus) culminating in Phase menopausal women. His translational research I and II clinical trials. Our current investigation has involved phase I and phase II clinical trials of licorice species will be highlighted with a of lycopene for the prevention of prostate focus on safety studies such as metabolism cancer in men and studies of the safety and of active compounds and potential for drug- efficacy of botanical dietary supplements for botanical interactions. Supported by NIH grant the management of menopausal symptoms in P50 AT000155 from the NIH Office of Dietary women. Prof. van Breemen has published over Supplements and the National Center for 300 papers and book chapters concerning Complementary and Integrative Health natural products, botanical dietary supplements and the use of mass spectrometry for drug Richard B. van Breemen is the discovery and development from natural Matthias C. Lu Collegiate product sources. Professor of Pharmacy and Professor of Medicinal [S6-2] The non-volatile constituents of Chemistry and Pharmacognosy nutmeg—a key to solving an old riddle? at the University of Illinois Presenter: Ehab A. Abourashed1 College of Pharmacy at Authors: Ehab A. Abourashed1 and Abir T. El-Alfy1 1 Chicago. He serves as Director of the UIC/NIH Department of Pharmaceutical Sciences, College of Pharmacy, Chicago State University, Center for Botanical Dietary Supplements Chicago, IL Research and leads the Mass Spectrometry, Nutmeg, the kernel of Myristica fragrans Metabolomics and Proteomics Facility for the (Myristicaceae), is a kitchen spice and a University of Illinois Cancer Center. Prof. van common ingredient of many food products. Breemen received his B.A. in chemistry from The characteristic taste and aroma of Oberlin College in 1980 and Ph.D. in nutmeg is mainly attributed to its essential Pharmacology and Experimental Therapeutics oil rich in terpenes and phenylpropenoids. from the Johns Hopkins University (with Nutmeg has also been reputed for its CNS

August 8–12, 2015 • University of Illinois at Urbana-Champaign 35 activity and is often classified as a natural Ehab A. Abourashed, M.S., hallucinogenic agent. Due to its ease of Ph.D. is a graduate of the accessibility, nutmeg is often used within Faculty of Pharmacy, Cairo confined communities as a cheap substitute University. He received an M.S. for marijuana and narcotic agents. Moreover, in Organic Chemistry from the sporadic reports have been published on University of Tennessee and a the anxiolytic/anxiogenic, antidepressant Ph.D. in Pharmacognosy from and anti-inflammatory effects of nutmeg the University of Mississippi. He is currently preparations. Based on earlier speculations, Associate Professor at the Chicago State the CNS effects of nutmeg were attributed University College of Pharmacy, which he to biotransformation of myristicin, the major joined in 2009 during its second year of constituent of its volatile oil, to amphetamine- inception, and Adjunct Research Associate like metabolites after consumption by abusers. Professor at the National Center for Natural Recent studies, however, failed to identify any Products Research, University of Mississippi. amphetamine-like metabolites in test animals Prior to Chicago State University, Dr. or human volunteers. On the other hand, Abourashed held other positions in academia, recent evaluation in our laboratory of HPLC- as Research Scientist at the University of fingerprinted extracts further established the Mississippi National Center for Natural psychoactivity of nutmeg in the mouse tetrad Products Research and Associate Professor at assay. Subsequent in vitro evaluation of the King Saud University College of Pharmacy. In same extracts against an extended panel of the private sector, he was Senior Scientist at CNS receptors, including cannabinoid type GlaxoSmithKline, New Jersey and Director of 1 & 2 receptors, did not identify any specific Quality Assurance at ElSohly Labs, Mississippi. target.\. Due to the apparently contradictory reports, i.e. lack of coherent chemical and Dr. Abourashed has more than 40 peer- molecular evidence alongside of significant reviewed publications, is the co-author of one in vivo activity, the phytopharmacology of textbook and is an editorial board member nutmeg had to be revisited in an attempt of the journal ‘Antioxidants’. His research to resolve this issue. Following a systematic interests and publications are focused on and reproducible phytochemical procedure, isolation, structure elucidation and microbial the major non-volatile constituents of transformation of biologically active natural nutmeg were isolated, characterized and products; application of HPLC and LC-MS in evaluated in vitro against novel targets that phytochemical, environmental and biochemical significantly modulate neurotransmission. analysis; and phytopharmacology of herbal Twelve compounds were identified and the medicines. Exposing professional pharmacy majority had a lignan skeleton with one new students to natural products drug discovery compound reported for the first time. A through investigating traditional medicinal number of the isolated compounds showed plants with psychoactive properties and significant activity against the newly identified antioxidant activities is the main focus of Dr. targets and may thus contribute towards the Abourashed’s current research activities. reputed psychoactivity of nutmeg. These compounds may have therapeutic potential in various psychological conditions including substance abuse withdrawal. Phytochemical and pharmacological details of the ‘road map’ leading to our proposed solution to the riddle of nutmeg will be the focus of this presentation.

36 The Phytochemical Society of North America [S6-3: Neish Award Speaker] Elucidating Philipp Zerbe is an Assistant modular diterpenoid metabolism in Professor at the Department of non-model plant systems: From chemical Plant Biology, University of diversity to biotechnology applications. California at Davis. His research Presenter and Author: Philipp Zerbe, Department interests focus on the of Plant Biology, University of California, Davis, CA evolutionary diversification of 95616 USA terpenoid metabolism in Plants are nature’s master chemists; they various medicinal and other economically produce a myriad of natural products that are relevant plants, and the development of tools essential for life on the planet. Mankind has for the production of terpenoid bioproducts utilized this chemical diversity for millennia with human benefit. Dr. Zerbe received his PhD for ceremonial purposes, as flavors, and to from the Ruhr-University Bochum, Germany cure various diseases. Among these bioactive (2007) under mentorship of Professor Elmar metabolites, terpenoids form the largest Weiler, studying the structural and functional and most diverse class with a wide range of interrelations of key enzymes in jasmonate ecological functions and industrial uses. We biosynthesis. Prior to his position at the established deep transcriptome resources for University of California, Davis, Dr. Zerbe worked more than a dozen plant species that produce as a Postdoctoral Fellow and later Research terpenoid metabolites of economic importance, Associate with Professor Jörg Bohlmann at the such as clary sage, balsam fir, horehound, Michael Smith Laboratories, University of British and maize. To decipher the corresponding Columbia (Vancouver, Canada). Here, his biosynthetic genes, enzymes and pathways research focused on the discovery of across various species, we developed a gene specialized diterpene pathways in coniferous discovery strategy that integrates tissue- trees and medicinal plants as part of two specific metabolite and transcriptome analysis Genome Canada funded research consortia with phylogeny-guided gene identification (SMarTForests, PhytoMetaSyn). and efficientin vitro and in vivo enzyme characterization. This approach resulted in the [S6-4] Antiplatelet aggregation and discovery of more than 60 diterpene synthases cytotoxicity activity of betulinic acid and several hundred cytochrome P450- and its acetyl derivative from Melaleuca dependent monooxygenases, as key enzymes bracteates var. revolution gold. in generating terpenoid metabolic diversity. Presenter(s): Foluso Oluwagbemiga Osunsanmi, Functional characterization revealed numerous University of Zululand at South Africa novel diterpene synthase functions as part of Author(s): Oluwagbemiga S. Soyinbe, University dynamic modular pathways, where catalytically of Zululand and Idiat. B. Ogunyinka , University distinct enzymes may function in different of Zululand and Rebamang A. Mosa and Andy R.Opoku, University of Zululand at South Africa. combinations to enhance chemical diversity. Following nature’s lead, we developed proof- Platelet dysfunctions are implicated in of-concept yeast expression platforms for cardiovascular diseases. Management of several diterpenoids through combinatorial abnormal Platelet aggregations with natural expression of functionally distinct diterpenoid products is a promising approach to the pathway genes. treatment of cardiovascular diseases. In this study, betulinic acid (BA) and its acetyl derivative (3-β acetylbetulinic acid) (BAA) from Melaleuca bracteata var. revolution gold were investigated for their antiplatelet aggregation and cytotoxicity. BA was

August 8–12, 2015 • University of Illinois at Urbana-Champaign 37 isolated from Melaleuca bracetata by column numerous American Indian plants that lack chromatography and some portion of BA scientific studies on their safety and efficacy. were used to synthesize BAA. The antiplatelet This talk will discuss the pharmacognostic aggregation activity of the compounds were process to identify if a subset of American separately evaluated on collagen, ADP, Indian botanicals can serve as alternatives thrombin and epinephrine to induce rat to HT while also protecting women from platelet aggregations. The MTT cytotoxicity hormone-dependent cancers. This project assay was used to determine the cytotoxic is expected to find novel and effective drug effect of the compounds against human leads for menopausal symptoms, cancer, and embryonic kidney (HEK293) and hepatocellular inflammation while also supporting American carcinoma (HEPG2) cell lines. BA and BAA Indian traditional knowledge. exhibited significant (p<0.05) dose dependent Fifteen native plants were selected to antiplatelet aggregation activity. BA and BAA examine estrogenic, chemopreventive, showed the highest platelet aggregation and anti-inflammatory potential through inhibition on epinephrine induced platelet bioassay-guided fractionation. Amorpha aggregation with IC50 values 0.78 mg/ml and canescens Pursh. (Fabaceae)—leadplant, 0.85 mg/ml respectively. BA and BAA showed Echinocystis lobata (Michx.) Torr. & A. less cytotoxicity effect on both HEK293 cell Gray (Cucurbitaceae)—wild cucumber, and (IC50 1027 µg/ml and 1051 µg/ml respectively) Silphium perfoliatum L. (Asteraceae)—cup and HEPG2 cells (IC50 448 mg/ml and 672 plant inflorescent tissue was selected for mg/ml respectively). The results suggest that initial screenings based on American Indian the compounds could serve as template for ethnobotany and published biological data. synthesis of new antiplatelet drugs. Leadplant MeOH extract was selected as the best candidate for bioassay-guided [S6-5] American Indian botanicals fractionation due to high anti-estrogenic as possible alternatives to hormone activity in the Ishikawa cell-based assay and therapy. anti-inflammatory activity in the Griess assay. Presenter: Tristesse Burton, University of Illinois at Cup plant and Lespedeza capitata Mixch. Chicago (Fabaceae) roundhead lespedeza (another Authors: Tristesse Burton1, Tareisha Dunlap1, Huali species within the 15 species) are being Dong1, Guannan Li1, Judy Bolton1, Djaja Soejarto1, and Richard B. van Breemen1 pursued as alternatives due to significant anti- estrogenic and estrogenic activity respectively. 1UIC/NIH Center for Botanical Dietary Supplements Research, Department of Medicinal Chemistry and Pharmacognosy, University of Illinois at Chicago, [S6-6] Molecular diversity and Evolution Chicago, IL 60612, USA of Peperomia species Although hormone therapy (HT) remains the Presenter(s): Massuo J. Kato, University of São Paulo standard treatment for managing menopausal Author(s): M.J. Kato1, L.Y.Valero-Gutiérrez1, M.M. symptoms, many women seek alternatives such Martins, M.C.R. Oliveira-Junior1, V. Bueno1, L.F. 1 2 2 as botanical dietary supplements since HT has Yamaguchi , A.P.M. Egydio , A. Salatino , M.L. Salatino2 been associated with increased risks of breast 1Department of Chemistry, University of São Paulo, cancer, coronary heart disease and stroke. Av. Prof. Lineu Prestes, 748, 05508-000, São Paulo, The leading botanicals that women take for SP Brazil menopause are black cohosh and red clover, 2Department of Botany, Institute of Biosciences, which were historically used by American Universidade de São Paulo, Rua do Matão, 277, Indians. Although these two botanicals have 05429-970, São Paulo, SP, Brazil been investigated extensively, there are still

38 The Phytochemical Society of North America One remarkable aspect among basal Wednesday, August 12, Morning Angiosperm is the diversification observed in some families. While some families such as Symposium 7: Nutritional & Medicinal Myristicaceae (e.g. nutmeg—Myristica fragrans) Phytochemistry contain nearly 440 species (20 genera), Chairs Elvira de Mejia and Shelly Nickols-Richardson Piperaceae (e.g. black pepper—Piper nigrum) has about 3600 species (5 genera). The family [S7-1] Phytoactive chemicals: discovery Piperaceae contains the two major genera is just at the “starting line” of the Piper and Peperomia with approximately 2000 research. and 1600 species, respectively. Although both Presenter and Author: Mary Ann Lila, Plants for genera are mostly Neotropical, phytochemical Human Health Institute, North Carolina State studies on Peperomia are scarce as compared University to Piper species. Thus, the major aim of this Research on the bioactive properties of work was to determine the phylogenetic phytochemicals, whether from edible food relationship among Brazilian Peperomia species crops or in herbals used in supplements (matK, ITS, and CHS as markers) as well as their or medicines, should generally include a chemical profiles obtained by means of HPLC- three-pronged approach: 1) Biodiscovery, DAD, LC-ESI-MS, NMR, and GC-MS data. The 2) Bioefficacy, and 3) Effective Delivery to phylogenetic trees provided by ITS and matK Consumers. Biodiscovery includes rigorous sequences were congruent with that from CHS. examination of the health-protective The Hierarchical Cluster Analysis (HCA) and constituents in well-recognized fruits and Discriminant Analysis of Principal Components vegetables, as well as exploration of novel (DAPC) of crude extracts obtained from leaves resources which may have historic incidence of indicated some trends with the largest group use by some localized cultures, but, may not encompassing species containing lignoids such have been examined in a scientific context. as P. blanda (tetrahydrofurans) and P. glabella Bioefficacy involves comprehensive testing (peperomins), while the second group contains of natural products/phytoactive chemicals the species P. tetraphylla (amides) and P. to decipher their unique biological activities trineura (polyketides). Due to the occurrence of against chronic human diseases, to discover orsellinic acid derivatives in several species, the mechanisms of action, and/or to determine corresponding OAS (orsellinic acid synthase) effective dosages relevant to human was investigated in the plants and in the 27 interventions. Bioefficacy can be investigated endophytic fungi isolated from them. Each at the lab bench in chemical bioassays, in vitro, fungi amplified for at least one fungal PKS in animal studies, or (as the gold standard) excepting the strain POS1 (an endophyte in human clinical trials. But unless we as from stems of P. oreophila) while the OAS researchers can follow through with that last was confirmed in several isolates. Although prong of the research agenda—i.e. effective the orsellinic acid was not detected in the delivery to consumers—everything else may be extracts of these endophytes growing in PDB end up as just an academic exercise. We need culture media, sequencing of the amplified to make sure that efficacious natural product fragment for the PUS3 fungi with 60% of discoveries can be deliverable to consumers in homology with the OAS from Streptomyces a format that they will willingly embrace, and viridochromogenes do not preclude the chose to adopt for their own health benefit. hypothesis of horizontal transfer from fungi to Despite rigorous informational and educational the plants. campaigns, consumers have largely shown reluctance and/or chronic inability even to incorporate adequate phytoactive-rich fruit and

August 8–12, 2015 • University of Illinois at Urbana-Champaign 39 vegetable produce into routine diets. In order globe, allowing consumers to make proactive, to address this disconnect, a novel strategy was responsible dietary choices that benefit their developed to concentrate immunoprotective own, and their families’ health. Dr. Lila is fruit and vegetable phytochemicals for currently a co-Director of an ambitious Plant delivery in convenient, and highly bioavailable Pathway Elucidation Project (P2EP) which functional food formats. Phytoactives synergizes the talents from 7 UNC system from candidate fruits and vegetables were universities, campus industry partners, and captured and concentrated into lightweight, partnering local 2 and 4-year colleges. portable protein-dense ingredients. Recent projects include a Bill&Melinda Gates Sensory panels confirmed the favorable Foundation Grand Exploration Challenges organoleptic properties of the ingredient, and project in Zambia, an NIH NIDDKD project recommended wider applications to counteract on functional food innovations, the Medicines the negative trends of Western diets. Most for Malaria Venture which examines promising recently, the phytoactive-protein chimeric plant-derived chemicals with potent anti- ingredients have been incorporated into snack malarial efficacy; a major blueberry genome food products with direct utility for meals in sequencing initiative using state-of-the-art transit, and even for humanitarian aid efforts in NextGen sequencing capacity, which focuses undernourished populations. Simultaneously, on the genes relevant to health-protective the complexing of phytoactive-protein particles properties in the fruits; and a USDA program addresses structural and formulation challenges on tribal resources and STEM education in (e.g. bar hardening, thermal degradation, American Indian/Alaska Native communities, or ingredient separations) that are current the health protective properties of traditionally- challenges in the industries. used medicinal planta, and the threats imposed Mary Ann Lila is Director of by climate change. the Plants for Human Health Lila was formerly Director (2006-2008) of ACES Institute, North Carolina State Global Connect (the international arm of the University, North Carolina College of ACES, University of Illinois) and Research Campus. She holds Associate Director of the nationally acclaimed the endowed David H. Functional Foods for Health Program (1997- Murdock Chair, and is a 2000) at the University of Illinois. Dr. Lila Professor in the Department of Food, has been honored with the Paul A. Funk Bioprocessing, and Nutrition Sciences. Through Scholarship Recognition Award (the premier ground-breaking, transdisciplinary discovery research award in the College of ACES, and outreach, the team of faculty at the Plants University of Illinois), the Spitze Professorial for Human Health Institute (PHHI) pioneers a Career Excellence Award, the Faculty Award for dramatic shift in the way the American public Excellence in Research, the University Scholar views and uses plant food crops—not merely as Award, the Amoco Award for Excellence a source of nutrients and flavorful calories, but in Undergraduate Instruction, and the Lilly as a powerful resource for components that Endowment Teaching Fellowship. Dr. Lila has protect and enhance human health. Integrated ongoing research projects in Egypt, Central research in metabolomics, biochemistry, Asia, Oceania, Mexico, Ecuador, Chile, and pharmacogenomics, breeding and postharvest Africa, and is Vice President of the Global attributes are aimed at development and Institute for BioExploration (GIBEX). In 1999, promotion of mainstream fruit and vegetable Dr. Lila won a Fulbright Senior Scholarship to produce with enhanced health benefits,and conduct research and outreach in New Zealand, introduction of new or underappreciated crops and returns to Australasia at least once/year. and products from various sites throughout the

40 The Phytochemical Society of North America [S7-2] Manipulation of glucosinolate marker-assisted introgression of beneficial biosynthesis, hydrolysis, and hydrolysis alleles for improved quality, yield, and other product anticancer bioactivity in broccoli traits into elite germplasm for commercial (Brassica oleracea L. ssp.). release. Most recently he is working in Presenter and Author: John A. Juvik, Department collaboration with Kraft Foods on a project of Crop Sciences, University of Illinois at Urbana- investigating the feasibility of using pigments Champaign from corn as natural colorants in processed Epidemiological studies have long suggested foods and beverages. Professor Juvik has that an inverse relationship exists between published over 100 peer-reviewed journal the consumption of Brassica vegetables and articles and has advised 36 M.S. and Ph.D. the induction of cancer. This response is students during his tenure at the University of associated with the hydrolysis of endogenous Illinois. glucosinolates generating indolyl and isothiocyanate products that display anti- [S7-3] Anti-Malarial Activities of carcinogenic bioactivities. Using a population Margaritaria discoidea and Other of broccoli F2:3 lines varying in glucosinolate Nigerian Medicinal Plants composition and concentration in floret tissue Presenter(s): Okiemute Rosa Johnson-Ajinwo, our lab has identified hydrolysis products and Institute for Science and Technology in Medicine, key genes associated with quinone reductase Keele University, UK. (QR) activity, an in vitro biomarker for anti- Author(s): Okiemute Rosa Johnson-Ajinwo1, 2, Imran 1 1 cancer activity. In another study, preharvest Ullah, Christopher Lee , Haima Raman , Nicolas Ellerby,1 Paul Horrocks1 and Wen-Wu Li1 spray treatment of the plant signal transduction 1 compound, methyl jasmonate (MeJA) and Institute for Science and Technology in Medicine, Keele University, UK postharvest treatment of 1-methylcyclopropene 2Faculty of Pharmaceutical Sciences, University of (1-MCP) was found to increase floret QR activity Port Harcourt, Nigeria and prolong broccoli postharvest quality. We previously evaluated in vitro anti-cancer activity John (Jack) A. Juvik, is the of a Nigeria medicinal plant—Margaritaria discoidea Director of Graduate Studies (Baill.) G.L.Webster [1]. The aim of current study for the Department of Crop is to investigate crude extracts of ten medicinal Sciences and the Director of plants including M. discoidea for their anti-malarial the Illinois Plant Breeding activities and identification of potential bioactive Center at the University of compounds. Illinois. Dr. Juvik’s research program has focused on breeding vegetables Methods: Ten selected medicinal plants for improved flavor, insect and disease (Anchomanes difformis (Blume) Engl., Mucuna resistance and for enhanced levels of pruriens var. pruriens, Bombax buonopozense phytochemicals associated with health P. Beauv, Millettia thonningii (Schum & Thonn.) promotion. He has investigated transformation Baker, Corchorus olitorus L., Margaritaria strategies and tissue culture ploidy level discoidea (Baill.) G.L. Webster, Moringa oleifera manipulation in Brassica vegetables and Lam., Newbouldia laevis (P. Beauv.) Seem., Miscanthus. His program has pioneered the use Fleurya sp and Platycerium bifurcatum (Cav.) of DNA markers for the creation of linkage C. Chr.) were sourced from a bio reserve in maps of tomatoes, sweet corn, broccoli and Nigeria. Extracts of the plants were screened more recently for the bioenergy crop, for anti-malarial activities on plasmodium Miscanthus. These maps have been used for falciparum. Following bioassay-guided the identification of favorable QTLs and for fractionation, potential bioactive compounds from M. discoidea were identified by gas

August 8–12, 2015 • University of Illinois at Urbana-Champaign 41 chromatography mass spectrometry following N/ ha) combined with two cutting heights (10 trimethylsilylation derivation. Their anti-malarial and 20 cm (BD) and 15 and 30 (BB) and were activity was confirmed and a preliminary applied according to complete randomized structural modification was undertaken. design in a 2x4x4 factorial arrangement, and season was considered a repeated measure. Results: Preliminary screening of extracts Sampling was done after every 28 days for from the ten plants disclosed that the organic phytochemical tests (saponin (foam test) and extracts of M. discoidea had the most phenols (ferric chloride). Foaming index varied promising anti-malarial activities. The potential only with season (P= 0.001) in BB, and lowest bioactive compounds in M. discoidea are values were reported in spring (58.9±2.06%). securinine (IC50 of 11.3 µM) and gallic acid However, in BD there was a significant (IC50 of 26.8 µM) against P. falciparum interaction between all factors (P=0.0006) Conclusions: M. discoidea possesses anti- on foam stability. There were no differences malarial activity against the intraerythrocytic between seasons in non-fertilized swards and stage of development. This is the first report of those receiving 450 kg N/ha, while lowest the anti-malarial activity of this plant. Securinine values were reported on treatments 300/20, and gallic acid appear the main anti-malarial 300/10 and 150/20, respectively in autumn, compounds in this plant. winter and spring. There was a significant Reference: [1] Okiemute Rosa Johnson-Ajinwo, interaction between all factors for phenols in Alan Richardson, Wen-Wu Li. Cytotoxic effects BD and BB (P<0.0001 for both species). In BD, of stem bark extracts and pure compounds from regardless cutting heights,highest values were Margaritaria discoidea on human ovarian cancer cell observed during summer and spring for all lines. Phytomedicine 2015, 22, 1-4. nitrogen levels. While, in BB the highest phenol concentration occurred in summer and autumn [S7-4] Phytochemical analysis for phenol or during summer and spring for all nitrogen and saponin in Brachiaria sp. levels on plant cutting heights of 15 cm and Presenter(s): Syeda Maryam Hussain (university of 30 cm, respectively. Overall, for both species, sao paulo) increasing on nitrogen fertilization tends to Author(s): Syeda Maryam Hussain, University of Sao decrease phenols concentration regardless the Paulo at Pirassununga, SP, Brazil. cutting height. However, variations on saponin Valdo Rodrigues Herling, University of Sao Paulo at seems more related to season of the year. Pirassununga, SP, Brazil. Lilian Elgalise Techio Pereira, University of Sao Paulo [S7-5] Coproduct yield comparisons at Pirassununga, SP, Brazil. between purple corn, blue corn and Plants secondary metabolites (PSM) can act as yellow dent corn for different milling a toxin or detoxifying agents, depending on processes their origin and nature. Tannins and saponins Presenter(s): Pavel Somavat, University of Illinois at are PSM found in Brachiaria sp., although the Urbana-Champaign former in negligible quantity, while the later in Author(s): Pavel Somavat, Qian Li, Elvira Gonzalez higher concentration. In order to comprehend de Mejia, Vijay Singh, University of Illinois at how sward management affects PSM, the Urbana-Champaign concentration of tannins and saponins were Purple corn (Zea mays L.) and blue corn (Zea determined in Brachiaria decumbens cv. mays var. saccharata) are varieties of colored Basilisk (BD) and Brachiaria brizantha cv. Xaraés corn rich in anthocyanins. Studies have shown (BB) in summer, autumn, winter and spring. that anthocyanins possess anti-oxidant/anti- Sward managements comprised four levels of carcinogenic properties and help fight obesity, nitrogen fertilization (0, 150, 300 and 450 kg tissue injury and inflammation. Various products

42 The Phytochemical Society of North America made from anthocyanin rich colored corn are The prevalence of diabetes mellitus is rising already available in market. In this study, we globally. The World Health Organization fractionated purple and blue corn using wet- (WHO), reports that this disease is one of the milling, dry-milling and dry-grind processes at 1 leading causes of deaths in the Philippines kg scale with three replications and compared and is responsible for about five percent of various coproduct yields against traditional global deaths. Diabetes can be divided into yellow dent corn (Zea mays var. indentata). In two types. In Type I, the pancreas does not wet-milling process, starch yield was largest produce sufficient insulin and in Type II, the for yellow dent corn 70.1% (d.b.) while it was insulin that is produced, does not enter the 63.3 and 61.5% (d.b.) for purple and blue corn. cells properly. Treatment of diabetes is done Gluten solids yield was largest in blue corn by injecting insulin and/or changing the diet. 14.2% (d.b.) while it was 13.8 and 10.5% (d.b.) Providing insulin can generate an elevation for purple and yellow dent corn. Fiber yields of postprandial hyperglycemia (PPHG) that were 10.9, 10.6 and 8.1% (d.b.) for purple, causes a spike in the production of glucose. blue and yellow dent corn, respectively. In Decreasing the level of PPHG may be dry-milling process, yellow dent corn yielded accomplished by inhibiting a carbohydrate- greatest amount of total endosperm fraction hydrolyzing enzyme like α-glucosidase. This 84.9% (d.b.), total endosperm yield for purple study, with funding by the USAID, investigated and blue corn were 80.1 and 77.0% (d.b.), leaf polyphenols content, alpha glucosidase respectively. Pericarp yields for blue, purple inhibition, antioxidant, and cytotoxic activities and yellow dent corn were 12.5, 9.9 and 6.9% of four Philippine herbal vines including (d.b.), respectively. In dry-grind process, final Merremia peltata, Ipomoea cairica, Ipomoea ethanol concentration was 17.1% (v/v) for lacunosa, and Ipomoea pes-caprae. The yellow dent corn while ethanol concentration antioxidant activity was determined by ABTS, for purple corn and blue corn were 14.5 and FRAP, DPPH, Hydroxyl radical, and Superoxide 14.4% (v/v), respectively. DDGS yield was radical inhibition assays. Cytotoxicity of the leaf greatest for purple corn 41.6% (d.b.) and extract was evaluated in terms of LC50 using yields were 38.0 and 32.9% (d.b.) for blue and brine shrimp lethality assay. Results revealed yellow dent corn, respectively. Purple corn that I. lacunosa had the highest polyphenolic and blue corn can potentially be used in all content (71.54 mg GAE per 100 gram dry the three milling processes, yield differences weight). HPLC/UV/MS analysis indicated that being offset by health promoting properties the four vines contained a large number of of various coproducts recovered. However, polyphenols and the majority of the peaks further techno-economical analysis of the three were identified as flavonoids.Ipomoea cairica processes is required. showed the highest antioxidant capacity in five methods used for radical scavenging [S7-6] Investigation of α-Glucosidase assays. The M. peltata, I. cairica, I. lacunosa, Inhibition Properties, Antioxidant and I. pes-caprae extracts demonstrated good

Activities, and Cytotoxicity of Philippine α-glucosidase inhibition with IC50 values of 500, Herbal Vines Containing Polyphenols 250, 234, and 62.5 μg/mL, respectively. Results Presenter(s): Richard I. Licayan, Rutgers the State from the cytotoxicity screens showed that the University of New Jersey extracts of M. peltata, I. cairica, I. lacunosa, Author(s): Thomas Villani, Rutgers the State and I. pes-caprae were potent against the University of New Jersey, Romeo M. Del Rosario, brine shrimp with LC50 values of 29.66, 61.54, Mindanao University of Science and Technology, 96.08, and 30.70 μg/mL, respectively. These Philippines, James E. Simon, Rutgers the State plants appear to be rich sources of natural University of New Jersey antioxidants and may be useful in treatment of diabetes mellitus.

August 8–12, 2015 • University of Illinois at Urbana-Champaign 43 Poster Abstracts

Poster Session I are also tolerant to abiotic stresses including Odd-numbered posters (e.g. P1, P3, P5...) salt, cold, and heat. The downside of manual phenotyping is that it is time consuming, Sunday, August 9, 5:30–7:30 pm low throughput, subjective, and limited to Illini Room AB the resolution of the human eye. On the other hand, high throughput phenotyping Poster Session II technologies are accurate, non-destructive, Even-numbered posters (e.g. P2, P4, P6...) and more sensitive, allowing the detection of Monday, August 10, 5:30–7:30 pm subtle phenotypes. We have used a Scanalyzer Illini Room AB HTS system to phenotype our high AsA Arabidopsis lines with visible, fluorescence, [P1] Phenomics Approaches to Elucidate and near infrared cameras. We have shown that the Role of the Various Ascorbate by using this approach, high AsA lines grow Pathways to Abiotic Stress Tolerance in faster, accumulate more biomass, and display Arabidopsis healthier chlorophyll fluorescence and water Presenter(s): Lucia Acosta-Gamboa, Arkansas content profiles than controls. By studying Biosciences Institute and Department of Molecular abiotic stress in a high throughout fashion using Biosciences optimized protocols, we have also shown that Author(s): Zachary Campbell, Arkansas Biosciences these high AsA lines are tolerant to salt and Institute, Arkansas State University drought stresses. Our ongoing experiments Raquel Torres, Arkansas Biosciences Institute, aim to dissect the contribution of the various Arkansas State University AsA pathways to abiotic stress tolerance. Lindsay Mull, Arkansas Biosciences Institute, Arkansas State University [P2] Isolation of flavonoids from Delonix Argelia Lorence, Arkansas Biosciences Institute and elata and determination of its rutin Department of Chemistry and Physics, Arkansas content using capillary electrophoresis State University Presenter(s): Areej Altaweel, Pharmacognosy L-Ascorbic acid (AsA, vitamin C) is a key Department, College of Pharmacy, King Saud antioxidant and enzyme cofactor in plants. University, P.O. Box 22452 Riyadh 11495, Saudi Ascorbate controls cell division, affects cell Arabia expansion, and plays an important role in Author(s): Ghada Ahmed Fawzy1,2, Shagufta 1 1,3 1 modulating plant senescence. It also protects Perveen , Hadir. M. Maher , Nourah. Z. Al-Zoman , 1 plants against reactive oxygen species Mona. M. Al-Shehri 1 that are produced in response to abiotic Pharmacognosy Department, 3Department of Pharmaceutical Chemistry, College of Pharmacy, and biotic stresses. Biosynthesis of AsA in King Saud University, P.O. Box 22452 Riyadh 11495, plants is carried out by a complex metabolic Saudi Arabia network involving D-mannose/L-galactose, 2Pharmacognosy Department, Faculty of Pharmacy, D-galacturonate, L-gulose, and myo-inositol Cairo University, Cairo 11562, Egypt as main precursors. We have previously shown 3Department of Pharmaceutical Analytical Chemistry, by manual phenotyping that Arabidopsis lines Faculty of Pharmacy, Alexandria University, El- over-expressing enzymes in the myo-inositol Messalah, Alexandria 21521, Egypt pathway have elevated AsA, accumulate more Delonix elata (L.) Gamble is an important, biomass of both aerial and root tissues and traditionally used plant in Saudi Arabia. It is

44 The Phytochemical Society of North America used to relieve rheumatic pain, flatulence lipid transfer proteins (PLTPs) are a group of and the seeds are employed as purgatives. highly conserved proteins found in tissues The aim of the present study was to isolate of higher plants primarily responsible for the chemical constituents of the n-butanol fraction shuttling of phospholipids and other fatty (BF) of D. elata and to find out, by capillary acid groups between cell membranes. Studies electrophoresis, percentage of rutin present also reveal that PLTPs play a very important in this BF. Three quercetin glycosides and role in the defense against phytopathogens. one kaempferol rutinoside were isolated from Previous studies using the Arabidopsis T-DNA the BF of aerial parts of D. elata; namely, insertion knockout plants were shown to be Quercetin 3-O-rutinoside-7-O-glucoside (1), defective in the growth phenotype as well as Quercetin 3,7-diglucoside (2), Rutin(3) and its basal resistance to pathogens. Based on Kaempferol 3-O-rutinoside (4). Rutin, an active this interesting finding and in order to fully constituent has been reported to possess delineate the role of SBIP-470 in plants, we are good pharmacological as well as therapeutic generating transgenic tobacco RNAi lines that potentials. A sensitive and rapid procedure for are silenced in SBIP-470 expression. SBIP-470 quantitative determination of rutin by capillary overexpressing transgenic tobacco lines using electrophoresis was developed and the content inducible system is also being generated. The of RUT was found to be 7.349 mg/gm, relative role of SBIP-470 in inducing SAR is currently to n-butanol fraction and 18.373 mg%, relative being investigated using Arabidopsis T-DNA to the dry powder of D. elata. The method knockout lines and in the SBIP-470 RNAi could be recommended for approval and use in tobacco transgenic lines when available. the pharmaceutical and food industries. Defense gene expression studies under biotic and abiotic stresses (drought and salinity) in the [P3] Characterization of SBIP-470, a RNAi-silenced plants compromised in SBIP-470 Family 1 Lipid Transfer Protein and its expression will also be carried out. In order to Role in Plant Stress Signaling biochemically characterize SBIP-470 in terms Presenter(s): Timothy Audam, East Tennessee of its ability to transfer and bind to plant lipids, State University, Johnson City, TN-37604 lipid binding and transfer assays will also be done. Fully understanding the role of SBIP-470 Author(s): Timothy Audam, East Tennessee in stress response of plants can lead to the State University, Johnson City, TN-37604; development of disease resistant plants which Dr. Dhirendra Kumar, East Tennessee State can have a direct impact on food production University, Johnson City, TN-37604, Danda globally. Chapagai, Department of Biological Sciences, University of South Carolina, Columbia, SC- [P4] Exploring the Transcriptome of the 29208 Glucosinolate/Myrosinase System in SBIP-470 is a tobacco SABP2-interacting Broccoli (Brassica oleracea var. italica) protein. Nucleotide sequence analysis of Presenter(s): Talon M. Becker, University of Illinois at SBIP470 clone obtained from the yeast two- Urbana-Champaign hybrid screening shows that it is a putative Author(s): Kang Mo Ku, University of Illinois at lipid transfer protein that belongs to nsLTP1 Urbana-Champaign subfamily. SABP2 is a methyl esterase like Elizabeth H. Jeffery, University of Illinois at Urbana- tobacco enzyme that catalyzes the conversion Champaign of methyl salicylate; an important lipid mobile John A. Juvik, University of Illinois at Urbana- signal to Salicylic Acid (SA). This is a critical Champaign step in the development of a systemic acquired The glucosinolate/myrosinase system is a plant resistance (SAR) against pathogens. Plant defense system found in the Brassicaceae

August 8–12, 2015 • University of Illinois at Urbana-Champaign 45 family, which includes the genera of Brassica Objective and Arabidopsis. This system has been linked The scientific and traditional data revealed primarily to plant defense against insect that Phyllanthus niruri (Euphorbiaceae) (PN), herbivory and pathogen infection, but also Andrographis paniculata (Acanthaceae) (AP) has been shown to affect human health. and Piper longum (Piperaceae) (PL) have Several glucosinolate hydrolysis products, their antioxidant and hepatoprotective role. the active compounds created from the In the present study, we have investigated reaction of glucosinolates and the enzyme, the synergistic effect of these herbs against myrosinase, have been shown to be bioactive alcohol and HFD (High Fat Diet) induce in humans. Specifically, the ingestion of these hepatotoxicity in rats. Further mechanistic study compounds can increase the transcription was conducted to establish the mechanism rates, and consequently the enzymatic activity, of the developed formulation with respect to of certain detoxification enzymes in the human Hemeoxygenase-1 (HO-1) which involve in the body. This has led to these compounds hepatoprotective effect. being described as having cancer-preventive/ Material and method chemopreventive activity, making the ability to The hepatoprotective activity of three herbal accurately manipulate this system an important extracts combinations Phyllanthus niruri breeding objective in the Brassica vegetable (PN), Andrographis paniculata (AP) and Piper market. In this research, we show the effect of longum (PL) were first studied in in-vitro on spraying the stress elicitor compound, methyl HepG-2 cell lines by using ethanol (100mM) jasmonate (MeJA), on the transcriptome of and then in-vivo activity was explored again the glucosinolate/ myrosinase system as well with 40% Alcohol 2ml/100g and HFD induce as final glucosinolate content and measurable hepatotoxicity in Wistar rats. Oxidative stress cancer-preventive bioactivity. In addition, parameters, liver enzymes, Lipid profile and we have explored the feasibility of using histopathological and mechanistic study were transcriptomic data of genes from this system assessed in liver homogenate. The body weight to build predictive models for final phenotypes, and urine analysis was done on 7th, 14th, 21st such as glucosinolate levels or measureable day. cancer-preventive bioactivity. If viable, this type of transcriptomic selection could be used in Results breeding programs focused on manipulating The herbal combinations with different ratios these traits. (AP: PN: PL, 1:1:1, 2:2:1, 3:1:1) showed significant hepatoprotective effect using in vitro [P5] Synergistic hepatoprotective effect (HepG-2) and in vivo models. Further herbal of Phyllanthus niruri and Andrographis combination with equal ratio group showed paniculata extracts in the presence of significant hepatoprotective effect as compare Piper longum in wistar rats. to other groups. The herbal combination group Presenter(s): Onkar Bedi,Department of showed significant reduction in oxidative stress Pharmacology, ISF College of Pharmacy, Moga parameters and improved antioxidant and liver (Punjab, INDIA) enzymes level. Author(s): Puneet Kumar,Department of Pharmacology, ISF College of Pharmacy, Moga Conclusion (Punjab, INDIA) and Vinod Gauttam,Department of The present studies suggest that the Pharmacognosy, ISF College of Pharmacy, Moga combination of herbal drugs showed (Punjab, INDIA) synergistic hepatoprotective effect by antioxidant and induction of HO-1 mechanism. The combination of these three herbal drugs provide base for their clinical implications.

46 The Phytochemical Society of North America [P6] Reduction of Total Glucosinolates in conversion further reduced GLS up to 97.8 and Canola Meal via Thermal Treatment and 97.5% in solid-state and submerged fungal Fungal Bioprocessing incubation, respectively. Optimized trials will be Presenter(s): Mark Berhow, USDA ARS NCAUR analyzed for toxic GLS breakdown compounds. Author(s): Jason R. Croat, Biology & Microbiology Department, South Dakota State University, [P7] GC-MS and NMR Metabolomic Brookings, SD 57007, USA, Mark Berhow, USDA, Profiling with Solid-state NMR Analysis Agricultural Research Service, National Center of FHT-RNAi Silenced Potato Periderm for Agricultural Utilization Research; Peoria, Tissues IL 61604, USA, Bishnu Karki, Agricultural & Biosystems Engineering Department, South Dakota Presenter(s): Qing Cai, City College of New State University, Brookings, SD 57007, USA, York, CUNY Graduate Center and Institute for Kasiviswanathan Muthukumarappan, Agricultural Macromolecular Assemblies, & Biosystems Engineering Department, South Author(s): Qing Cai, City College of New York; Dakota State University, Brookings, SD 57007, USA, Wenlin Huang, City College of New York; Olga and William R. Gibbons, Biology & Microbiology Serra, University of Girona, Campus Montilivi Mercè Department, South Dakota State University, Figueras, University of Girona, Campus Montilivi Brookings, SD 57007, USA Marisa Molinas, University of Girona, Campus Montilivi; Ruth E. Stark, City College of New York On a worldwide basis, canola (Brassica napus) meal is second only to soybean meal Suberin is a biopolyester constituent of as a protein source for livestock. A general specialized plant periderm tissues formed limitation of Brassica spp. meals is the within the phellem cell walls. Suberin and presence of glucosinolates (GLS). GLS and waxes of the periderm layer act to prevent the enzyme myrosinase are compartmentally water diffusion, mechanical breakdown and stored separately in the plant. Upon disruption pathogenic invasion in plants. Ferulic esters of plant tissues, myrosinase cleaves glucose are considered to be the most important from GLS and releases toxic compounds such linkage between aliphatic and aromatic as nitriles, thiocyanates, and isothiocyanates suberin domains and also linked with cell-wall to defend the plant. For this reason canola polysaccharides. The potato gene FHT (fatty was bred to contain lower levels of GLS and ω-hydroxyacid/fatty alcohol hydroxycinnamoyl erucic acid. However feed inclusion rates are transferase) esterifies ferulic acid to suberin. still limited to 25-30% of livestock diets. The Diminished levels of feruloyl transferase activity objective of this research was to develop a have been associated with lowered amounts microbial process to metabolize GLS and the of feruloyl esters of fatty acids in both suberin- resulting breakdown products into non-toxic associated waxes and suberin breakdown components, to enable higher inclusion levels products derived from transesterification. FHT in livestock rations. An additional objective was knockdown by RNAi silencing is accompanied to boost the protein levels and digestibility. by increases in periderm water permeability Cold pressed and hexane extracted canola but unaltered periderm ultrastructure. meals were autoclaved (thermal treatment) To investigate how this potato suberin then processed using several metabolically biosynthesis pathway is affected in FHT- diverse fungal cultures. Aurobasidium pullulans, RNAi silenced potato cultivars, metabolite Trichoderma reesei, Fusarium venenatum, profiles for non-polar periderm tissue extracts Pichia kudriavzevii, and Mucor circinelloides were examined by complementary GC- are grown in solid-state and submerged culture MS and solution NMR measurements, and at 50 and 10% solid loading rate, respectively. solid suspensions after polar and nonpolar Total GLS levels were reduced up to 65.5 and extractions were assessed by solid-state 13C 79.8% by thermal treatments while microbial NMR. Metabolomic analyses (PCA and OPLS- DA) showed consistency among biological

August 8–12, 2015 • University of Illinois at Urbana-Champaign 47 replicates and discrimination between 20:4 acts as an endocannabinoid ligand FHT-RNAi and wild type samples, allowing and mediates a multitude of physiological identification of 25 out of 31 biomarkers responses. This unique NAE type, NAE 20:4 is including alkanes, α,ω-fatty diacids, fatty acids/ hypothesized to effect development, growth alcohols, methyl esters and glyceryl esters. and cellular organization of P. patens. To The relative numbers of each major carbon determine the role of NAEs in moss type in the solid residue were estimated from development, NAE content and composition in quantitatively reliable DPMAS 13C NMR protonema, early and late gametophyte stages, spectra. The FHT-RNAi cultivar has oxygenated and sporophytes, will be quantified from their aromatic-to-(CH2)n and oxygenated-aliphatic- total lipid extracts, using selective lipidomics. to-(CH2)n ratios that exceed the values for wild The effects of anandamide on growth will be type cultivars by ~60 and 80%, respectively. studied by culturing moss in the presence of These compositional trends indicate an exogenous NAE 20:4 in a dose-dependent enhanced hydrophilic-hydrophobic balance manner. Temporal changes in growth patterns in FHT-RNAi suberin samples, consistent with will be determined by the evaluation of digital augmented water permeability and reduced images using ImageJ tool. The effect of relative amounts of fatty acid and fatty alcohol- anandamide on cytoskeletal organization will containing polymeric products deduced be visualized by immunostaining the phyllodes from suberin breakdown. The solid-state exposed to NAE 20:4 and observing them results complement our GC-MS data showing under confocal microscope. Preliminary results large amounts of fatty acid and fatty alcohol indicate that the endogenous NAE content metabolites accumulated in non-polar extracts and composition is variable, depending on from FHT-RNAi native periderms. the developmental stage and that NAE 20:4 is a potent negative regulator of moss growth. [P8] Effects of Anandamide on More detailed studies are expected to provide Development, Growth and Cellular novel insights into the role NAEs, specifically Organization of Physcomitrella patens NAE 20:4 might play in mediating growth and Presenter(s): Jedaidah Chilufya, East Tennessee development of seedless plants. State University Author(s): Shiva Devaiah and Aruna Kilaru, East [P9] Isolation of new phenolics with Tennessee State University. anticancer and antioxidant potential Mosses are bryophytes with a simple cellular from roots of Rhodiola imbricata and organization and distinctive growth stages. their quantification by validated HPLC With their unique lipid profile, most mosses are method also tolerant to various stressors. A ubiquitous Presenter(s): Alka Choudhary, aDepartment class of bioactive fatty acid ethanolamides in of Natural Products, National Institute of eukaryotes called N-acylethanolamines (NAEs) Pharmaceutical Education and Research, Sector-67, also occurs in the moss Physcomitrella patens. S.A.S. Nagar, 160062, Punjab, India Unlike in higher plants, where saturated and Author(s): Alka Choudhary, Sunil Kumar Surapaneni, unsaturated NAE types are limited to those Kulbhushan Tikoo, and Inder Pal Singh, National with acyl chains 12C to 18C, P. patens also Institute of Pharmaceutical Education and Research, contains anandamide, NAE 20:4. In higher Sector-67, S.A.S. Nagar, 160062, Punjab, India. plants, NAEs are most abundant in desiccated Raj Kumar, Division of Phytochemistry, Defence seeds and mediate plant growth, development, Institute of High Altitude research (DRDO), C/o 56 APO, Leh-Ladakh, 901205, J&K, India cellular organization and response to stress, in an abscisic acid (ABA)-dependent or Several plants of genus Rhodiola (family independent manner. In mammals, NAE Crassulaceae) have been widely used in

48 The Phytochemical Society of North America traditional/modern medicine for their ability and malaria. Extracts from leaves, seeds, fruits to enhance physical endurance and to treat and roots of this plant have shown to possess impotence, fatigue, gastrointestinal, cardiac biological activities in vitro, however the and central nervous system disorders. One such bioactive constituents of these extracts have species with cytoprotective/radioprotective not been elucidated. Furthermore, the seeds of properties is Rhodiola imbricata. Aqueous this plant are the source for the natural pigment extract of R. imbricata rhizome has shown “annatto dye” which is commonly used in the anti-cancer activities, which might be useful in food industry. Previously, we established hairy leukemia cancer treatment. The proliferation root cultures of annatto to produce and identify of K-562 cell line was significantly decreased potential bioactive compounds from this after incubation with aqueous extract of R. species. This work led to the characterization imbricata due to induction of apoptosis and of candidate chemicals with anti-malarial cell cycle arrest at G2/M phase. Inspite of activity in vitro, including ishwarane, tocotrienol several biological studies, no phytochemical and stigmasterol (Molecules 2014, 19:756- studies have been reported for this plant. As 766). To increase the levels of these bioactive a part of our ongoing programme to isolate compounds, the hairy root cultures of annatto and characterize natural products, we planned were treated with different combinations of phytochemical investigations on R. imbricata elicitors, including methyl jasmonate and in search of new molecules from nature. In the cyclodextrin, for different time periods. HPLC present study, phytochemical investigations analyses showed that several compounds were led to isolation of several phenolic glycosides induced and excreted into the culture medium. (including three new compounds) along with Further characterization of these compounds is simple phenolics from the roots of R. imbricata. underway. We propose that hairy root cultures The structures of isolated molecules were of annatto can be developed as a sustainable established by NMR spectroscopy (1D and 2D source for the production and discovery of NMR) and Mass spectrometry (ESI HRMS). The bioactive compounds with distinct bioactivities. compounds were evaluated for antioxidant activity in various antioxidant assays and [P11] Black bean (Phaseolus vulgaris anticancer activity against A549 and MCF7 cell L.) anthocyanins: extraction, thermal lines. the percentage of isolated molecules was stability and shelf-life at different pHs determined in the extract by a validated HPLC and temperatures method. Presenter(s): Luis Mojica, University of Illinois Author(s): Luis Mojica, University of Illinois, Andy [P10] Application of annatto hairy Tan, University of Illinois, Mark Berhow, United roots for the production of bioactive States Department of Agriculture, Agricultural compounds for human health Research Service, and Elvira Gonzalez de Mejia, University of Illinois Presenter(s): Jarrod Creameans, Arkansas Biosciences Institute The aim of this research was to optimize the Author(s): Jarrod Creameans, Lingling Fang, anthocyanins extraction from black bean Denzel McGregory, and Luis Nopo-Olazabal, coats and evaluate their thermal and shelf-life Arkansas Biosciences Institute, Fabricio Medina- stability at different pHs and temperatures. Bolivar Arkansas Biosciences Institute and Dept. of Anthocyanins were extracted from black bean Biological Sciences, Arkansas State University coats using different ethanol concentrations, Annatto (Bixa orellana) is a tropical plant native temperatures and solid-to-liquid ratios. to South America which is currently used as a Anthocyanin solutions were adjusted to traditional medicine to treat multiple diseases different pHs according to commercially including skin infections, respiratory problems available beverages (pH 2.5, 3.0, 3.5, 4.3).

August 8–12, 2015 • University of Illinois at Urbana-Champaign 49 From the black bean coats, it was possible enables the confident assignment of to obtain a 7.1% dry powder yield of elemental compositions to ions and facilitates anthocyanins. Optimal extraction conditions identification of known compounds and begins were ethanol 24%, solid-to-liquid ratio 1:40 structure elucidation of unknowns. In order and 29 ˚C (P < 0.0001). Bean anthocyanins to collect more comprehensive metabolomic were more stable at pH 2.5 and 4 ˚C (89.6%) profiles in a single analysis we have developed in comparison to 22 ˚C (19.3%), with a half-life a combined ESI-APCI ion source coupled to of 273 and 56 days, respectively. Anthocyanins a HRAM LC-MS system capable of polarity presented the lowest degradation rate value at switching. We applied this technique for 70 ˚C (k = 0.061/h) and pH 3.0, and the highest the metabolomic profiling of five species half-life values (11 h). Energy of activation of grasses: Arundinella hirta, Capillipedium ranged from 55.9 to 84.8 kJ/mol, having spicigerum, Coelorachis tuberculosa, higher values the solutions with lower pHs. Ischaemum hubbard and Miscanthus junceus. The highest Q10 change was observed at pH 2.5 with a value of 2.99 for 70 to 80 °C, while [P13] Biomimetic Preparation of the lowest Q10 was found at pH 4.3 (1.18) for Halogenated Norcoclaurine Analogs via 80 to 90 ˚C change. Three anthocyanins were Tyrosinase for Directed Biosynthesis In identified by their mass ions as delphinidin Benzylisoquinoline Alkaloid Producing 3-O-glucoside (M+ = 465.1), petunidin Cells 3- O- glucoside (M+ = 479.1) and malvidin Presenter(s): Jordan Fauser, DePaul University 3-O-glucoside (M+ = 493.1). Black beans are a Author(s): Jordan E Fauser1, Sean O Crowe1, Realino good source of anthocyanins with potential to Gurdiel1, Greg Gillespie1, Michael J Sislow1, Justin J be used as food colorants. Maresh1 1Chemistry, DePaul University, 2320 North Kenmore [P12] Development of a Combined Avenue, Chicago, IL 60614 Electrospray Ionization/Atmospheric We present a low-cost, one-pot, aqueous Pressure Chemical Ionization, Polarity synthesis of halogenated norcoclaurine Switching High Resolution-Accurate analogs, as potential substrates for precursor Mass Liquid Chromatography-Mass directed biosynthesis. Using a cross linking Spectrometry Method and Comparative technique we form an insoluble aggregate of Metabolomics of Five Grass Species tyrosinase from plant and fungal sources such Presenter(s): Bradley S. Evans, Donald Danforth as sweet potato, avocado, and white button Plant Science Center mushroom. This aggregate catalyzes oxidation Author(s): Elizabeth A. Kellogg, Donald Danforth of halogenated tyramine to halogenated Plant Science Center dopamine, a reaction with an inherently The two most widely utilized ionization unfavorable equilibrium. High-yield turnover is source types in liquid chromatography- achieved by coupling dopamine production to mass spectrometry (LC-MS) are electrospray Pictet-Spengler condensation with 4-hydroxy- ionization (ESI) and atmospheric pressure phenylacetaldehyde to generate halogenated chemical ionization (APCI). Each can be used analogs of the tetrahydroisoquinoline in either positive or negative ionization modes. norcoclaurine. The resulting norcoclaurine The ESI+, ESI-, APCI+ and APCI- modes of analogs may be isolated by extraction or used operation provide complementary information directly from the reaction mixture for precursor- about the molecular composition of a sample, directed biosynthesis in plant cell cultures yet if all these modes are used for analysis it is to produce halogenated benzylisoquinoline in series rather than parallel. High resolution alkaloids. To the best of our knowledge, this accurate mass (HRAM) mass spectrometry work is the first to successfully utilize tyrosinase

50 The Phytochemical Society of North America for high-yield production of unnatural catechol [P15] Identification and quantification products. of bioactive compounds from Panax ginseng using HPLC-CAD [P14] Gas chromatography-based Presenter(s): Rajarshi Ghosh, Middle Tennessee myrosinase activity and QTL mapping in State University broccoli (Brassica oleracea L. var. italica) Author(s): Rajarshi Ghosh, Anthony Farone, Elliot Presenter(s): Alicia M. Gardner, University of Illinois Altman and Paul Kline, Middle Tennessee State at Urbana-Champain University Author(s): Alicia M. Gardner, University of Illinois Panax ginseng is one of the most popular at Urbana-Champaign, Kang Mo Ku, University and widely used medicinal plants worldwide. of Illinois at Urbana-Champaign, Allan F. Brown, North Carolina State University, and John A. Juvik, Ginseng roots have been shown to have broad University of Illinois at Urbana-Champaign spectrum pharmacological effects on the immune system, cardiovascular system, and Vegetables in the Brassica genus have been central nervous system. The ginsenosides and shown to contain a number of compounds polysaccharides have been credited to be the that exhibit pharmacological activity. This active ingredients responsible for the various research elucidates genetic loci involved in pharmacological effects of ginseng. With the the production of glucosinolate hydrolysis increasing amount of ginseng products in the products, which are associated with anti-cancer market, it is essential to have proper quality bioactivity in broccoli (Brassica oleracea L. control procedures to standardize the presence var. italica). The myrosinase (thioglucosidase of various ginsenosides and polysaccharides EC 3.2.1.147) activities of 104 F2:3 families in adequate quantities. There is a need for from the mapping population BNC x VI158, new and simple methodologies for detection grown in 2009 and 2010, were determined and quantification of these compounds. In by the quantification of sinigrin and benzyl the present study, an attempt has been made glucosinolate (exogenous glucosinolate) to identify and quantify the various bioactive hydrolysis products on a gas chromatography- compounds in ginseng using High performance flame ionization detector system. QTL analysis liquid chromatography (HPLC) coupled with of the data was conducted in the software charged aerosol detector (CAD). CAD offers package MapQTL 5 (Van Ooijen, 2004) using an attractive alternative detection technique a dense linkage map with 590 mapped SNP for various compounds as it does not depend marker loci, covering an estimated 96% of the on the spectral properties of the analyte. B. oleracea genome. This map was developed This feature is especially helpful in detecting by Brown et al. (2014) using a 60K Illumina SNP compounds such as carbohydrates which array of Brassica napus, and is anchored to the have poor UV absorptivity. Quantification genomic sequence of the rapid cycling Brassica of compounds can be performed based on oleracea TO1000. The results of this study will calibration curve of a single standard. A Waters provide useful information for the identification XBridge amide column with gradient elution of genes involved in the chemopreventative was used to separate the compounds. The activity of broccoli and support the breeding detected compounds were identified based on of Brassica cultivars that can serve as functional retention times of respective standards. foods. [P16] Synthesis of 1,3,4-oxadiazole derivatives as potential and selective COX-2 inhibitors Presenter(s): Jagdeep Grover, Department of Natural Products, National Institute of

August 8–12, 2015 • University of Illinois at Urbana-Champaign 51 Pharmaceutical Education and Research, Sector-67, [P17] Metabolomic Profiling and S.A.S. Nagar-160062, Punjab Thioacidolysis Analysis of FHT-RNAi Author(s): Jagdeep Grover, Bhatt Nirav, Vivek Silenced Potato Periderm Tissues Kumar, Neeraj Patel, M. Elizabeth Sobhia, K. K. Presenter(s): Liqing Jin, City College of New Bhutani, Sanjay M. Jachak York, CUNY Graduate Center and Institute for National Institute of Pharmaceutical Education and Macromolecular Assemblies Research, Sector-67, S.A.S. Nagar-160062, Punjab, Author(s): Liqing Jin, City College of New York, India Wenlin Huang, City College of New York, Olga Cyclooxygenase (COX) catalyzes the Serra, University of Girona, Campus Montilivi, Joan conversion of arachidonic acid to inflammatory Rigau, Centre for Research in Agricultural Genomics mediators such as prostaglandins (PGs), (CRAG), Consorci CSIC-IRTA-UAB-UB, Campus de Bellaterra UAB, Mercè Figueras, University of prostacyclins and thromboxanes. Ongoing Girona, Campus Montilivi, Marisa Molinas, University safety concerns pertaining to the use of non- of Girona, Campus Montilivi, Ruth E. Stark, City selective NSAIDs have spurred development College of New York of coxibs with improved safety profile.1 As a The potato (Solanum tuberosum) is an part of our continued efforts to discover new important staple crop. Potato tubers are COX inhibitors2,3, a novel series of 2,5-biaryl- protected from dehydration and pathogens by 1,3,4-oxadiazoles have been synthesized a covering peel (periderm) impregnated with and evaluated for in vitro COX-1 and COX-2 suberin, a complex cross-linked heteropolymer inhibition, in vivo anti-inflammatory potential that contains both polyaliphatic and lignin-like and cytotoxicity evaluation in J774A.1 and aromatic domains. Current models describing RAW 264.7 cell lines. Almost all the compounds the macromolecular structure of suberin assume displayed selective COX-2 inhibition with SI that ferulic acid cross-links both domains as it range from 30-100. Compounds with nitro may form carboxyl esters bonds with aliphatic and tert-butyl substituents emerged as most monomers and non-ester radical coupled potent with IC50 value in the range of 0.81- bonds with phenolics. Ferulic acid also links by 0.89 µM against COX-2. The same compounds ester bonds to glycans and acts in cross-linking also exhibited superior anti-inflammatory polysaccharides and lignins. Fatty alcohol/ω- activity (50-59% inhibition of edema at hydroxyacid hydroxycinnamoyl transferase 150 µmol/Kg) in comparison to celecoxib. (FHT) is a BADH acyltransferase responsible Molecular docking studies further predicted of the synthesis of akyl-ferulates necessary for the binding orientation of ligands at the active suberin biosynthesis. Periderm from tubers site of enzyme. Cytotoxicity study of potent deficient in FHT showed a significant decrease compounds revealed the cell viability of more in suberin ferulate esters and an increase in than 90% in the said cell lines. The present soluble phenolics. This periderm also showed study suggests the significance of bulky changes in the texture and water permeability substituent like methylsulfonyl and tert-butyl but, surprisingly, the ultrastructure of the as a pharmacophoric feature for selective suberin is not altered. To further investigate inhibition of COX-2 enzyme. the consequences of FHT silencing in potato References periderm, polar extracts of native tuber 1. Blobaum, A. L.; Marnett, L. J. J. Med. Chem. periderm have been analyzed using Liquid 2007, 50, 1425-1441. Chromatography-Mass Spectrometry (LC-MS), 2. Grover, J.; Kumar, V.; Singh, V.; Bairwa K.; Sobhia, revealing significantly enhanced amounts of six M.E.; Jachak, S.M. Eur. J. Med. Chem. 2014, 80, identified phenolic amines including feruloyl 47-56. tyramine, feruloyl putrescine and caffeoyl 3. Grover, J.; Kumar, V.; Sobhia, M.E.; Jachak, S.M. putrescine, which are derived from feruloyl- Bioorg. Med. Chem. Lett. 2014, 24, 4638-4642. CoA. The insoluble part of suberin has also

52 The Phytochemical Society of North America been subjected to thioacidolysis and analyzed alone, relative to blank controls. S. agrili were by Gas Chromatography-Flame Ionization also significantly attracted to a stem with a larva Detector (GC-FID), showing that FHT-RNAi and leaf tissue together, relative to leaf tissue aromatic suberin contains larger amounts of alone. In contrast, naïve S. floridanus were guaiacyl units, which are also derived from attracted to a larva within a stem, but not to feruloyl-CoA. These metabolite and polymer any other component. Thus, naïve S. agrili and changes for FHT-RNAi periderms and the S. floridanus appear to employ different host- associated fate of feruloyl-CoA in the suberin location strategies. Because S. floridanus attack biosynthetic pathway suggest that FHT-RNAi multiple species of Agrilus, host-associated silencing disfavors alkyl ferulate biosynthesis cues perceived during emergence from their and favors an alternative disposition of the natal hosts may improve attraction to emerald feruloyl-CoA substrate. ash borer larvae. Subsequent experience may elicit attraction to plant foliage, and thereby [P18] Responses of two parasitoids facilitate location of host habitat. In contrast of the emerald ash borer, Agrilus naïve S. agrili may learn additional host- planipennis Fairmaire, the introduced associated cues during oviposition. Further Spathius agrili Yang, and native Spathius understanding of host-location behavior floridanus Ashmead, to volatile host- may improve the utility of these parasitoids associated cues for biological control, both by suggesting Presenter(s): Todd David Johnson, University strategies for pre-release conditioning, and of Wisconsin-Madison (where the work was by providing tools for assessing post-release conducted); establishment. University of Illinois Urbana-Champaign (current affiliation) [P19] Design, Synthesis, Drug-Likeness Author(s): Todd David Johnson (as above), Jonathan and Anti-ovarian Cancer Activity of P. Lelito, USDA APHIS PPQ, EAB Unit, and Kenneth F. Raffa, University of Wisconsin-Madison Thymoquinone Analogues Presenter(s): Okiemute Rosa Johnson-Ajinwo, The introduced specialist parasitoid Spathius Institute for Science and Technology in Medicine, agrili Yang (Hymenoptera: Braconidae) has Keele University, UK. been released as a biological control agent Author(s): Okiemute Rosa Johnson-Ajinwo1,2, Alan of the emerald ash borer, Agrilus planipennis Richardson1, and Wen-Wu Li1 Fairmaire. Spathius floridanus Ashmead 1Institute for Science and Technology in Medicine, (Hymenoptera: Braconidae) a native generalist Keele University, UK parasitoid, opportunistically attacks A. 2Faculty of Pharmaceutical Sciences, University of planipennis and may contribute to control Port Harcourt, Nigeria of the insect. We currently know little about Cancer-related deaths claim about 20,000 the host location behaviors of these wasps. lives daily. Thymoquinone (TQ), a bioactive We evaluated responses of female wasps to compound isolated from Nigella sativa has components of this host complex, Fraxinus demonstrated in vitro moderate activities pennslyvanica (green ash) stem tissue, F. against breast, colon, ovarian, pancreatic and pennslyvanica foliage, and an A. planipennis prostate cancers, including leukaemia, but larva within a stem. Experiments were the compound is poorly soluble.1 Recently, conducted in Y-tube olfactometer bioassays, thymquinone has been shown to block the using wasps reared on A. planipennis larvae pSer/pThr recognition by Polo-like-kinase feeding in F. pennslyvanica twigs at USDA 1 Polo-Box-Domain at the atomic level. In APHIS in Brighton, MI. Naïve S. agrili were this work, the synthesis of TQ analogues via attracted to the entire complex, and leaf tissue structure-based design, cytotoxicity, solubility,

August 8–12, 2015 • University of Illinois at Urbana-Champaign 53 drug-likeness and structure-activity relationship The present study was conducted to evaluate (SAR) investigation were carried out. the hepatoprotective and antioxidative potential of hydroalcoholic extract of Glycosmis Methods: Eleven TQ analogues were pentaphylla leaves (GpE) and chloroform, synthesised via reduction and amine addition ethyl acetate & n-butanol fractions of GpE. to quinone, while nine analogues were The toxicant CCl4 (1 ml/kg in olive oil; sc) was procured from Sigma Aldrich. The in vitro administered on 4th and 5th day of study to Sulforhodamine B assay and Trypan blue induce oxidative hepatotoxicity in rats. The method were used for the evaluation of the hepatoprotection of the test drugs where cytotoxic activities of TQ and its analogues on compared with the reference drug silymarin three ovarian cancer cell lines; A2780, OVCAR8 (50 mg/kg). The pre-treatment of rats with and A2780CIS. The in vitro Selective index higher dose of GpE (400 mg/kg) and EAf (SI) was determined using the human ovarian (150 mg/kg) for 7 days produced a significant epithelial cells (HOE). Phosphate buffer was (p<0.05) dose dependent hepatoprotective employed in the aqueous solubility study. and antioxidative affect evident by decreased ChemBioDraw software was used to estimate levels of serum (AST, ALT, ALP & TB) and the log P values of the compounds. tissue TBARS in addition to increased levels Results/Conclusion: TQ analogues disclosed of serum (TP & ALB) and reduced tissue GSH. a structure-activity relationship. Three of the The histological examination provided the analogues showed significant cytotoxic activity supportive evidences that strongly demonstrate and were twice-more potent than TQ. The antioxidative affect on hepatocytes and solubility data validated the Log P values restoring their normal functional ability. The estimated by the software. An evaluation of present study scientifically validates the free the drug-likeness of the analogues showed radical-scavenging property to be one of the that ‘Lipinski rule of 5’ was obeyed; justifying mechanisms of hepatoprotection and justify their potentials for further research as possible the traditional medicinal claims attributed to clinical agents. Glycosmis pentaphylla leaves. Further detailed studies are in progress to elucidate bioactive References: 1. R. Schneider-Stock, I. H. constituent/s and their concentration in the Fakhoury, A. M. Zaki, C. O. El-Baba, H. U. Gali- plant. Muhtasib. Thymoquinone: fifty years of success in the battle against cancer models. Drug [P21] In-vitro Cytotoxicity Activity of Discov Today 2014, 19, 18. Eclipta alba Linn Against A-549, HEp-2 and HeLa Cell Lines [P20] In-vivo Hepatoprotective and Antioxidative Activity of Glycosmis Presenter(s): Sonia Kamboj, Dept. of Biology, Satluj Public School, Sirsa Pentaphylla Leaves Against CCl4- Induced Oxidative Hepatocellular Author(s): Sonia Kamboj, Department of Biology, Satluj Public School, Sirsa, Haryana, India-125055 Damage and Uttam Chand Gupta, Department of Presenter(s): Pradeep Kamboj, JCDM College of Microbiology, Hans PG College, Jaipur, Rajasthan, Pharmacy India-302001 Author(s): Pradeep Kamboj, Department of The present study was aimed to evaluate the Pharmacognosy, JCDM College of Pharmacy, Sirsa, in-vitro cytotoxicity activity of ethanolic extract Haryana, India-125055 of Eclipta alba aerial parts (EaE) against three Ajudhia N. Kalia, Department of Pharmacognosy, human cancer cell lines namely lung (A- 549), Sri Sai College of Pharmacy, Pathankot, Punjab, India-145001 liver (Hep-2) and Cervix (HeLa) cell lines. The cytotoxicity potential of EaE was evaluated

54 The Phytochemical Society of North America by SRB and MTT assays and the percentage glucosylate anthocyanidins, it did glucosylate viability of the cell lines were carried out flavanones and flavones in addition to using Trypan blue dye exclusion method. The flavonols. This is significant because flavanones EaE extract showed maximum percentage of and flavonols do not contain a 3-OH group. growth inhibition against A-549 cell line; where HPLC was performed to identify the reaction as in case of Hep-2 cell lines no cytotoxicity products. Early results indicated that the activity was observed. In concentration range mutant protein glucosylates naringenin at 7-OH of 1000 to 31.25 (µg/ml), EaE significantly position forming prunin. Product identification produced cytotoxicity effect on A-549 and with other substrates is in progress. Results are HeLa cell lines with CTC50 values 44.60 and being used to revisit and refine the structure 51.30 in SRB and MTT assays, respectively. model. Structural and functional analysis of Therefore, the results of present study flavonoid GTs may contribute to custom design scientifically validate the use ofEclipta alba of GTs for the synthesis of novel glucosides by as an anticancer agent in traditional system of changing glucosylation patterns medicine, and provide potential information for its further in-vivo and phytochemical studies. [P23] The identification of biochemical pathway genes from non-model plant [P22] Structural and functional analysis species of Grapefruit Cp-3-O-GT mutant P145T. Presenter(s): Matthew B. Kilgore, Donald Danforth Presenter(s): Sangam Kandel, East Tennessee State Plant Science Center University, Johnson City, TN Author(s): Megan M. Augustin, Donald Danforth Author(s): Sangam Kandel, Sarah Khaja, Shivakumar Plant Science Center Devaiah, McIntosh A. Cecilia, East Tennessee State Gregory D. May, National Center for Genome University, Johnson City Resources Flavonoids are a class of secondary John A. Crow, National Center for Genome metabolites, the majority of which are present Resources in glucosylated form. Glucosyltransferases are Toni M. Kutchan, Donald Danforth Plant Science the enzymes that mediate glucosylation by Center transferring glucose from a high energy sugar Secondary metabolites are often restricted donor to the acceptor substrates. Our study in their distribution to different groups of focuses on the structural and functional analysis organisms. For this reason, attempts to study of a flavonol-specific 3-O-glucosyltransferase these often useful and interesting forms of (Cp-3-O-GT) clone from Citrus paradisi that metabolism require an ability to work in a has been characterized previously in our lab. diversity of non-model species. Methods for Multiple sequence alignment and homology gene discovery with low investment and high modeling was done to identify candidate efficiency are needed to effectively identify residues for mutation. Cp-3-O-GT was modeled the biosynthetic genes in these diverse with a flavonoid 3-O-GT from Vitis vinifera pathways. During this work, a workflow for (VvGT) that can glucosylate both flavonols efficiently identifying biosynthetic genes is and anthocyanidins. We identified a proline developed and applied to Amaryllidaceae residue at position 145 of Cp-3-O-GT that alkaloid biosynthesis. Genes discovered corresponded to a threonine residue in VvGT during this work include a cytochrome and designed a Cp-3-O-GT- P145T mutant P450 capable of phenol-phenol coupling to test the hypothesis that that mutation of 4’-O-methylnorbelladine to noroxomaritidine, proline by threonine in Cp-3-O-GT could a norbelladine 4’-O-methyltransferase alter substrate or regiospecificity of Cp-3-O- (NpN4OMT) and a short chain dehydrogenase/ GT. While the mutant P145T enzyme did not reductase capable of forming norbelladine

August 8–12, 2015 • University of Illinois at Urbana-Champaign 55 from tyramine and 3,4-dihydroxybenzaldehyde. used to identify reaction products. An enzyme These enzymatic discoveries support the kinetics study will show the extent of any future application of this workflow to other biochemical change in function as a result of biosynthetic pathways and organisms. this mutation; results will then be incorporated into a refined protein model. [P24] The Effect of R382w Mutation on C. Paradisi Flavonol Specific [P25] SABP2, A Tobacco Methyl 3-O-Glucosyltransferase Salicylate Esterase has Role in Presenter(s): Kathleen King, East Tennessee State Modulating Nonhost Resistance University Presenter(s): Dhirendra Kumar, East Tennessee State Author(s): Shivakumar Devaiah, East Tennessee University State University and Cecilia McIntosh, East Author(s): MD Imdadul Haq, East Tennessee State Tennessee State University University Flavonoids are a class of plant metabolites Pavan Chigurupati, East Tennessee State University with C6-C3-C6 structure responsible for many Nonhost resistance is a type of broad-spectrum biological functions, including coloration and resistance exhibited by a given plant species to defense. Citrus paradisi, grapefruit, contains a most strains of a pathogen which are generally wide variety of flavonoids which are grouped pathogenic to other plant species. Studies by the extent of modification, examples of show that gene-for-gene resistance activates which are flavonols, flavones, and flavanones. SA-signaling but if it also activated during A major modification is the addition of glucose nonhost resistance is not clear. SABP2, a methyl by glucosyltransferases (GTs) to stabilize salicylate esterase is a critical component in the structure and provide ease of transport. SA-signaling pathway in tobacco plants, it also This process can be highly substrate and has a role in nonhost resistance. To study this, regiospecific. With Cp3OGT, glucose is added SABP2-silenced tobacco plants were infected at the 3-hydroxy position. This 3GT only with tobacco nonhost pathogen, Pseudomonas accepts flavonols as its substrate; however, syringae (P.s) pv. phaseolicola (Psp) and its a Vitis vinifera (grape) 3-GT can accept both growth was monitored. Data analysis revealed flavonols and anthocyanidins. Homology slightly enhanced growth of Psp suggesting modeling using the crystallized structure of SABP2 plays a minor role in inducing nonhost the V. vinifera GT predicted sites of amino resistance against Psp in tobacco plants. acids that could influence substrate binding RT-PCR analysis of SABP2 transcripts upon site. The 382 position was of particular interest Psp infection showed that its transcript level with arginine in C. paradisi and tryptophan increased with time in both control plants in V. vinifera. This change is hypothesized to as well as in SABP2-silenced tobacco plants cause a shift in substrate specificity of the suggesting the involvement of SABP2 in Cp3OGT to accept anthocyanidins as well nonhost resistance. To fully suppress the SABP2 as flavonols. Site-directed mutagenesis was activity, SABP2-silenced plants were further performed to form the R382W mutant Cp3OGT treated with 2,2,2,2’-tetra-fluoroacetophenone and transformed into yeast for expression. (tetra-FA), a known inhibitor of SABP2 prior to Western blot determined the optimal protein the inoculations with Psp. The tetra-FA treated induction period for the cells, after which the SABP2-silenced plants showed compromised cells were broken to extract the recombinant resistance to Psp suggesting a role for SABP2 mutant protein. Purification of the R382W 3GT in nonhost resistance. This observation was allowed for enzyme analysis to be performed supported by gene expression analysis of by measuring the incorporation of radioactive tetra-FA treated plants. The expression of other glucose into the reaction product. HPLC will be nonhost marker genes such as CDM1 and HIN1

56 The Phytochemical Society of North America was also monitored in P.s. tabaci (Pst, virulent/ heptanal, 2-octanone could be observed host pathogen on tobacco), P.s. phaseolicola only in aroma rice varieties, while 14 volatiles and P.s. tomato (Psp and Psto, avirulent/ such as 3,7,11-trimethyl-1-dodecanol and nonhost pathogens on tobacco), and TMV 1,3-dimethyl-Benzene could be observed only infected control and SABP2-silenced tobacco in non-aroma rice variety. Two tested aroma plants. This revealed no significant difference varieties even showed different volatile profiles which suggest that these two genes are probably due to the difference in flavor- activated in SABP2-SA independent manner. inducing genes introduced during the breeding Overall results suggest that SABP2-mediated program. SA signaling has a role in nonhost resistance in tobacco plants. Understanding the mechanism [P27] Sulfur-containing amino acids of nonhost resistance is likely to help in enhanced the antioxidant activity of developing broad-spectrum disease-resistant peptides present in common bean crop plants. (Phaseolus vulgaris L.) non-digestible fraction [P26] Characterization of Volatile Presenter(s): Diego Luna-Vital, University of Illinois at Compounds in Grains of Newly Urbana-Champaign Developed Aroma Rice Varieties by Author(s): Diego Luna-Vital, University of Illinois at using HS-SPME-GCMS Urbana-Champaign; Michael Kolman, University of Presenter(s): Young-Sang Lee, Soonchunhyang Illinois at Urbana-Champaign; Guadalupe Loarca- University Pina, Universidad Autonoma de Queretaro and Elvira de Mejia, University of Illinois at Urbana- 1 1 Author(s): M M Chayan. Mahmud , E.J. Kim , Y. H. Champaign Cho2, Y.J. Park3, Y.H. Kim1, and Y.S. Lee1 The objective of this research was to assess 1Dept. of Medical Biotechnology, Soonchunhyang Univ. Asan, South Korea, 2Seedpia Inc. Suwon, the antioxidant potential of five pure South Korea, 3Dept. of Plant Resources, Kongju synthesized peptides (GLTSK, LSGNK, Nat’l Univ., Yesan, South Korea GEGSGA, MTEEY and MPACGSS) originally To characterize flavor property of newly identified from common bean non-digestible developed aroma rice varieties (cv. fraction (NDF). The in vitro evaluations were Gahyangchal-1-ho and cv. GoldenQueen-3- performed using CCD-33Co normal colon ho), brown rice powders were transferred into a cells treated simultaneously with hydrogen headspace vial and solid phase microextraction peroxide (H2O2) and the bean peptides for (SPME) combined with a gas chromatography 4 h. Intracellular reactive oxygen species mass spectrometry (GCMS) were used for the (ROS) were detected with the cell permeant analysis of volatile compounds and comparison reagent 2’,7’ –dichlorofluorescein diacetate to non-aroma variety (cv. Chucheong). Based (DCFDA), a fluorogenic dye that measures upon NIST mass spectral library total 64 volatile several ROS activities. The cytoprotective compounds consisting of 28 hydrocarbons, effect of the peptides on CCD-33Co cells 10 alcohols, 7 ketones, 4 aldehydes, 3 esters, exposed to H2O2 was determined treating and some bases etc. could be identified in all the cells simultaneously with H2O2 and the 3 tested varieties. Among those 64 identified, peptides for 24 h and evaluating cell viability. 17 volatiles such as hexanal and 2-pentyl- Also, superoxide (SO), hydroxyl (HO) and nitric furan were found commonly in both aroma oxide (NO) radicals scavenging activity and and non-aroma rice varieties suggesting the effect on linoleic acid oxidation of the these compounds as general volatiles in rice peptides were assessed. The peptides reduced grain. However, 23 volatiles such as 2-acetyl- the intracellular concentration of ROS ranging 1-pyrroline (responsible for popcorn flavor), from 18.2% to 72.1% in CCD-33Co cells.

August 8–12, 2015 • University of Illinois at Urbana-Champaign 57 Besides, the peptides increased the viability a monophyletic clade that is sister to a (68.2% to 124.8%) of CCD-33Co cells in cystine lyase clade. Detailed biochemical comparison when they were treated with H2O2 characterization showed that the recombinant only (21.3%). Furthermore, it was observed enzymes of both TAT1 and TAT2 converted that peptides had scavenging properties with Tyr to HPP, as previously reported, but also radical inhibition over NO (16.1% to 73.8%), used various keto acceptors and other SO (9.5% to 64.6%) and HO (4.9% to 56.0%) amino acids such as phenylalanine (Phe) and radicals. Finally, the peptides promoted a methionine (Met). The tat1 and tat2 single delay in linoleic acid oxidation ranging from and double mutants of A. thaliana exhibited 0.0066 to 0.426 AU/h (absorbance units are only partial reduction in total TAT activity, proportional to oxidation level), compared to increased Tyr levels, and partially reduced the control (0.516 AU/h). The number of sulfur- levels of tocopherol and plastoquinone. GFP containing amino acids significantly correlated localization and subcellular fractionation with all the antioxidant activities tested; studies further showed that TAT1 and TAT2 as NO (r=0.95), SO (r=0.95) and HO (r=0.69), well as subsequent HPP dioxygenase (HPPD) the reduction on linoleic acid oxidation (r=- are located outside of the plastids. These 0.86), and the in vitro determinations of ROS results indicate that TAT1 and TAT2 partially decreased (r=0.66) and cytoprotective effect contribute to the conversion of Tyr to HPP (r=0.70). In general, MPACGSS was the most outside the plastids, leading to tocopherol and potent peptide followed by MTEEY in all the plastoquinone biosynthesis. However, TAT1 evaluations, correlating with the presence and TAT2 likely have additional functions and of sulfur-containing amino acids methionine also yet to be identified aminotransferases and cysteine, known to provide antioxidant having TAT activity exist in both cytosol and properties to peptides. plastids in A. thaliana. The study highlights biochemical and genetic complexity of Tyr [P28] Biochemical and genetic catabolism leading to the formation of natural characterization of tyrosine products in plants. aminotransferases in Arabidopsis thaliana [P29] Solgen 40, the high Genistin/ Presenter(s): Hiroshi A Maeda, University of Genistein soy concentrate prevents Wisconsin-Madison bone mineral loss during menopause in a Author(s): Minmin Wang, Kyoko Today, Hiroshi A. dose concentrate manner preventing the Maeda, University of Wisconsin-Madison oxidized bone microenvironment. In plants, tyrosine (Tyr) serves as a key precursor Presenter(s): Jose Angel Marañon, Tradichem of various plant natural products (e.g., Innovation Center tocochromanols, plastoquinone, ubiquinone, Author(s): Lucia de los Santos, Cristina Lozano, rosmarinic acid, morphine alkaloids), which are Lorena Martíne-CampesinoTradichem Innovation crucial for plant environmental responses and Center. c/ Faraday 7. LAB 02. Parque Científico de Madrid. Campus de Cantoblanco. 28049- of pharmaceutical or nutritional importance Madrid (SPAIN), Ernesto Caballero-Garrido Dep. to human. Tyrosine aminotransferases (TAT) Neurosurgery. University of New Mexico. 1100 catalyze the reversible transamination between Yale Blvd. 87131. Alburquerque. NM. USA, Tyr and 4-hydroxyphenylpyruvate (HPP), the Fernando Galán-Estella Química Analítica y Análisis key entry reaction to many of the Tyr-derived Instrumental. Universidad Autónoma de Madrid. compounds. Here we report phylogenetic, Campus de Cantoblanco. 28049- Madrid (SPAIN) biochemical, genetic, and subcellular Estrogen deficiency accelerates the effects of localization analyses of TAT enzymes from aging on bone by decreasing defense against Arabidopsis thalian. TAT1 and TAT2 formed oxidative stress (OS). Genistin/genistein

58 The Phytochemical Society of North America and daidzin/daidzein, the most common gradually. The resulting absorption data comes phytoestrogens from soy isoflavones seems to as a sigmoidal curve from which melting protect the adult skeleton against bone loss temperature can be determined when half of by slowing the rate of bone remodeling and the DNA has denatured. The current study by maintaining a focal balance between bone aims to assess DNA intercalating activities formation and resorption. Recent studies report of four pure bioactive isoquinoline alkaloids: that reactive oxygen species (ROS) may play a sanguinarine, berberine, allocryptopine role in postmenopausal bone loss by creating and chelerythrine which were isolated from a more oxidized bone microenvironment and Chelidonium majus (Papaveraceae) by increase the intracellular concentration of repeated silica gel column chromatography, the antioxidant glutathione in bone prevents recrystallization and preparative thin bone loss during estrogen deficiency in mice. layer chromatography (TLC). The isolated For understanding how soy isoflavones can compounds were identified by comparing their prevent the bone damage by ROS and help physical properties and mass spectra with those in the maintenance of glutathione levels we of the published data. The results showed that have studied the capacity of genistin/genistein, sanguiarine is the most active intercalating daizin/daidzein for blocking a common ROS as agent with Tm value of 83.55 ± 0.49 followed DPPH radical, by a novel HPLC-DPPH-DAD in by berberine, chelerythrine and allocryptopine vitro test. Our results has revealed that the high with Tm values 62.58 ± 0.47, 51.38 ± 0.37 and genistin/genistein soy concentrate SOLGEN 50.94 ± 0.65, respectively, relative to 49.78 ± 40 blocks ROS species in a dose dependent 1.05 of bacteriophage DNA alone and 86.09 ± manner. In the other hand soy concentrates 0.5 for ethidium bromide as a positive control. containing high daidzin/daidzein just partially blocks ROS and the increase of dadizin/ [P31] Improving stability of bean daidzein concentration do not increase the anthocyanins by β-cyclodextrin in a block of ROS. Therefore, the administration sport beverage of high dosages of SOLGEN 40 and other Presenter(s): Luis Mojica, University of Illinois at high genistin/genistein soy concentrates Urbana-Champaign seems to be a safe an efficient strategy during Author(s): Luis Mojica, University of Illinois at menopause for preventing bone mineral loss Urbana-Champaign; Y., Aguilera, Universidad by diminishing the intracellular concentration of Autónoma de Madrid; M.A. Berhow, United States reactive oxygen species Department of Agriculture; María A Martín-Cabrejas, Universidad Autonoma de Madrid; and E. González de Mejia, University of Illinois at Urbana-Champaign [P30] DNA Intercalating Alkaloids Isolated From Chelidonium Majus The objective was to investigate the color (Papaveraceae) stability of common bean anthocyanins from two black bean cultivars (Otomi and Idaho) Presenter(s): Tamer Mohamed, Institute of Pharmacy and Molecular Biotechnology, Department of co-pigmented with β-cyclodextrin in a sport Biology, University of Heidelberg, Germany beverage, as potential replacements for Author(s): Wink Michael, Institute of Pharmacy and synthetic colorants. The model beverage Molecular Biotechnology, Department of Biology, system was exposed to white fluorescent light University of Heidelberg, Germany and stored at 32 ºC for 10 days. Antioxidant DNA intercalating agents increase the stability capacity, total anthocyanins, and individual of DNA which can be demonstrated by anthocyanin characterization (LC-ESI-MS) were measuring the melting temperature (Tm). Tm performed. Cultivar Idaho bean exhibited can be determined in a spectrophotometer the highest antioxidant capacity (325.1 µM in which the cell temperature is increased Trolox/g dry coat) and presented the highest

August 8–12, 2015 • University of Illinois at Urbana-Champaign 59 concentration of anthocyanins (183.0 mg/100 best identify and annotate small molecules g dry coat). For both cultivars, petunidin based on mass spectrometric data. In this 3-O-glucoside (1.2-37.8 mg/100 g dry coat), contest participants are asked to solve the delphinidin 3-O-glucoside (55.8-60.0 mg/100 structures of unknown compounds based on g dry coat), petunidin (48.5-85.3 mg/100 g a set of high resolution MS and MS/MS data dry coat) and malvidin 3-O-glucoside (5.1- provided by the organizers. The third round of 14.5 mg/100 g dry coat) were found. The CASMI was organized by an international team degradation reaction of anthocyanins in the of researchers from the United Kingdom and presence of β-cyclodextrin followed the first- the United States. The challenges were divided order kinetics (6.2 and 5.1 d-1 in Otomi and into four groups: endogenous metabolites, Idaho cultivars, respectively). A similar behavior natural products, synthetic chemicals and occurred in the absence of β-cyclodextrin unknown compounds. In addition to high (4.5 and 3.9 d-1 in Otomi and Idaho cultivars, resolution MS and MS/MS data, additional respectively). The half-life (t1/2) of anthocyanins information such as chromatographic data was increased significantly (p < 0.05) with the and sample origin was also provided. Mass addition of 2% β-cyclodextrin (8 vs. 13 days). spectrometric data were acquired on different Moreover, in the co-pigmented sport beverage types of instruments including qTOF, Orbitrap, the color difference values (ΔE*) were higher IT-TOF and FTMS. The contest consisted (43.7 and 47.1 in Otomi and Idaho cultivars, of two categories: best molecular formula respectively) than the no co-pigmented (Category 1) and best structure (Category 2). (34.4 and 30.2 in Otomi and Idaho cultivars, Seven participants took part in the contest. respectively) under the same experimental The winner in the category 1 correctly solved conditions. Thus, data demonstrated that bean 34 out of 42 challenges; the winner in the anthocyanins have the potential to be used as Category 2 correctly solved 23 out of 42 natural pigments in beverages stabilized by challenges. Natural products proved to be the β-cyclodextrin. most challenging group of compounds with the smallest number of correct solutions submitted. [P32] CASMI 2014: Challenges, solutions and results [P33] Molecular Identification of Presenter(s): Dejan Nikolic, University of Illinois at Tuliposide B-Converting Enzyme: a Chicago Lactone-Forming Carboxylesterase from Author(s): Warwick Dunn,School of Biosciences, the Pollen of Tulip University of Birmingham, Birmingham, United Presenter(s): Taiji Nomura, Toyama Prefectural Kingdom University Lloyd W. Sumner,The Samuel Roberts Noble Author(s): Taiji Nomura, Toyama Prefectural Foundation, Ardmore, OK University; Martin Jones, School of Biosciences, University of Shinjiro Ogita, Toyama Prefectural University; Birmingham, Birmingham, United Kingdom Yasuo Kato, Toyama Prefectural University Emma Schymanski,Eawag: Swiss Federal Institute of Aquatic Science and Technology, Dübendorf, 6-Tuliposides A (PosA) and B (PosB), which are Switzerland the major secondary metabolites in tulip (Tulipa Steffen Neuman, Leibnitz Institute of Plant gesneriana), are enzymatically converted to the Biochemistry, Halle, Germany antimicrobial lactonized aglycons, tulipalins The Critical Assessment of Small Molecule A (PaA) and B (PaB), respectively. We recently Identification (CASMI) contest, inaugurated identified a PosA-converting enzyme (TCEA) in 2012, was designed as the medium for as the first reported member of the lactone- researchers to exchange ideas as to how to forming carboxylesterases, Herein, we report the identification of another lactone-forming

60 The Phytochemical Society of North America carboxylesterase, PosB-converting enzyme SBIP68, a tobacco protein was found was (TCEB), which preferentially reacts with PosB to found to interact with SABP2 in a two-hybrid give PaB. This enzyme was isolated from tulip screening. SABP2 is an important component pollen, which showed high PosB-converting of the salicylic acid pathogen response activity. Purified TCEB exhibited greater activity pathway in tobacco and other plants. Based towards PosB than PosA, which was contrary on bioinformatics, SBIP68 was predicted to that of the TCEA. Novel cDNA (TgTCEB1) to be a putative UDP-glucose: flavonoid encoding the TCEB was isolated from tulip glucosyltransferase, having a GT1_Gtf_like pollen following peptide sequencing of the conserved domain as well as the 44-amino purified TCEB. TgTCEB1 belonged to the acid PSPG Box characteristic of Family 1 carboxylesterase family of alpha/beta-hydrolase glycosyltransferases (UGTs). Glucosides are fold and was approximately 50% identical a ubiquitous class of secondary metabolites to the TgTCEA polypeptides. Functional involved in roles ranging from the protection of characterization of the recombinant enzyme plants against pathogens and herbivory to the verified that TgTCEB1 catalyzed the conversion physical appearance of plants, transportation of PosB to PaB with an activity comparable of metals, symbiotic agents between plants with the native TCEB. RT-qPCR analysis of and microorganisms, and acting as sexual each part of plant revealed that TgTCEB1 hormones. Glucosyltransferases transfer transcripts were limited almost exclusively to glucose molecules from activated glucose the pollen, despite the presence of significant donors like UDP-glucose to potential levels of PosB-converting activity in several aglycones, producing the corresponding other tissues (e.g., roots), which suggested the glucosides. The full length SBIP68 gene was presence of a TgTCEB1 isozyme gene being cloned and expressed heterologously in expressed in such tissues. Immunostaining of E. coli as well as in Pichia pastoris. Purified the anther cross-section using anti-TgTCEB1 recombinant protein from Pichia was further polyclonal antibody verified that TgTCEB1 used for biochemical analyses. A total of was specifically expressed in the pollen grains, fourteen different substrates have been but not in the anther cells. N-Terminal transit tested so far, including flavonoids and simple peptide of TgTCEB1 was shown to function phenolics. It showed relatively high activity as plastid-targeted signal. Taken together, with several flavonoids known to have roles in these results indicate that mature TgTCEB1 is stress signaling. Biochemical analyses using specifically localized in plastids of pollen grains. simple phenolics as aglycones is in progress. Interestingly, PosB, the substrate of TgTCEB1, Research is also ongoing to determine the accumulated on the pollen surface, but not in effect of SBIP68’s interaction with SABP2 on its the intracellular spaces of pollen grains. This biochemical activity. Earlier studies on the gene sequestration system of the enzyme (TgTCEB1) expression analyses of SBIP68 showed that it is from its substrate (PosB) represents a unique likely modulated upon viral pathogen infection. chemical defense mechanism of the pollen of Studying SBIP68 will help to improve our tulip. general understanding of how plants respond to pathogen attacks. [P34] Characterization of SBIP68 a Putative Tobacco Glucosyltransferase and Its Role in Plant Defense Presenter(s): Abdulkareem Odesina, East Tennessee State University Author(s): Abdulkareem Odesina and Dhirendra Kumar, East Tennessee State University

August 8–12, 2015 • University of Illinois at Urbana-Champaign 61 [P35] Peptides from beans increased and (25%) applied directly to mature 3T3- insulin secretion, reduced oxidative L1 cells. In addition, treatments improved stress and inhibited lipid accumulation in insulin-induced glucose uptake (64%) in insulin in vitro models resistant adipocytes. This study demonstrates Presenter(s): Miguel Eduardo Oseguera Toledo, that protein hydrolysates and their peptide University of Illinois at Urbana-Champaign/ fractions, even after gastrointestinal digestion, Universidad Autonoma de Queretaro improved insulin secretion and protected Author(s): Miguel E. Oseguera-Toledo, Universtity pancreatic cells against oxidative stress, of Illinois at Urbana-Champaign/Universidad inhibited lipid accumulation and improved Autónoma de Querétaro: Silvia Amaya- glucose uptake in adipocytes. Llano,Universidad Autónoma de Querétaro; Elvira González de Mejía, Universtity of Illinois at Urbana- Champaign [P36] A Survey of Anthocyanin Concentration and Composition in The objective was to evaluate the effect of Diverse Maize Germplasm de-hulled hard-to-cook (HTC) bean bioactive Presenter(s): Michael Paulsmeyer, University of peptide fractions on important proteins Illinois at Urbana-Champaign for type-2 diabetes, oxidative stress, lipid Author(s): Laura Chatham, University of Illinois at accumulation and glucose uptake in in vitro Urbana-Champaign models. Protein isolates from two common Kang-Mo Ku, University of Illinois at Urbana- bean varieties, pinto Durango and Negro 8025 Champaign beans, were hydrolyzed (120 min) with either Leslie West, University of Illinois at Urbana- alcalase® or bromelain and then peptides < 1 Champaign kDa were fractionated using an ultrafiltration Megan West, Kraft Foods, Glenview, IL membrane system. The hydrolysate and Charlotte Allen, Kraft Foods, Glenview, IL peptide fraction < 1 kDa were further hydrolyzed with pepsin-pancreatin simulating a Jack Juvik, University of Illinois at Urbana- Champaign gastrointestinal digestion. To evaluate the role of the peptides as insulin secretagogues and Increasing consumer demand for more antioxidants, iNS-1E pancreatic beta-cells were natural ingredients has spurred interest in used. Pinto-bromelain peptide fractions < 1 the replacement of artificial dyes in foods kDa were able to increase glucose-stimulated and beverages with naturally occurring insulin secretion by 20-150 µIU insulin/mL in a phytopigments. In this study, our lab used dose-dependent manner (1 μg/mL and 100 μg/ HPLC to screen the anthocyanin content of mL, respectively) in iNS-1E cells; Pinto-alcalase over 375 accessions of maize from diverse peptide fractions < 1 kDa increased the relative backgrounds. Anthocyanin concentrations protein expression of IGF-II, IGFBP-2 and 3 and in our collection ranged from less than 1 reduced VEGF. Oxidative stress and reactive to 2547 mg anthocyanins per kg whole oxygen species (ROS) in basal conditions did corn. After HPLC analysis, each sample not affect cell viability however decreased ROS was categorized based on the location and production (70%). Pre-treatment of iNS-1E cells abundance of various anthocyanin structural with hydrolysates or their fractions (1–100 μg/ types. Our findings showed that pericarp ml) for 20 h significantly reduced cell damage tissue produces more pigment than aleurone induced by 100 μM tert-butylhydroperoxide tissue on average and pericarp tissue has (t-BOOH) for 2 h, up to 70% for complete the unique ability to produce anthocyanins hydrolysate of pinto Durango-alcalase (100 condensed with flavonols. Aleurone tissue μg/ml). The effect of peptides reduced was found to be important as a source of high intracellular fat accumulation (28%) when ratios of malonylglucoside anthocyanins. In applied during 3T3-L1 adipocyte differentiation general, cyanidin derivatives were the most

62 The Phytochemical Society of North America common anthocyanin types, but we have remain largely untested. We take advantage found 43 accessions segregating for the purple of recent technological advances for the aleurone1 (pr1) gene that produce primarily generation of spectral data from crude plant pelargonidin types of anthocyanins. Finally, we extracts to characterize phytochemical diversity are now characterizing a new phenotype that, in a suite of co-occurring plants in the tropical to our knowledge, has never been documented genus, Piper (Piperaceae). In combination with before in maize. Several accessions in our 20 years of data on Piper associated insects, collection are unable to produce anthocyanins we found that phytochemical diversity has a containing acylated glycosides. The information direct and positive effect on the diversity of collected in this study represents the most herbivores, but also reduces overall herbivore comprehensive screen of colored maize lines damage. Elevated chemical diversity is to date, providing a valuable tool for the associated with more specialized assemblages production of high anthocyanin yielding corn of herbivores, and the cascading positive effect lines. of phytochemistry on herbivore enemies is stronger as herbivore diet breadth decreases. [P37] Phytochemical diversity drives It is clear from these results that high tropical plant-insect community diversity phytochemical diversity not only enhances the Presenter(s): Lora A. Richards, University of Nevada diversity of plant-associated insects, but also Reno contributes to the ecological predominance of Author(s): Lee A. Dyer, Biology Department, specialized insect herbivores. University of Nevada Reno, Reno, NV 89557 Matt L. Forister, Biology Department, University of [P38] P450 from woad is involved in Nevada Reno, Reno, NV 89557 formation of indigo and indirubin in Angela M. Smilanich,Biology Department, University recombinant Escherichia coli of Nevada Reno, Reno, NV 89557 Presenter(s): Laxmi Sagwan-Barkdoll, Southern Craig D. Dodson, Chemistry Department, University Illinois University Carbondale of Nevada Reno, Reno, NV 89557 Author(s): Laxmi Sagwan-Barkdoll, Parminder Michael D. Leonard,Chemistry Department, Multani and Aldwin Anterola, Southern Illinois University of Nevada Reno, Reno, NV 89557 University Carbondale Chris S. Jeffrey, Chemistry Department, University of The plant Isatis tinctoria, commonly known Nevada Reno, Reno, NV 89557 as woad, is a natural source of the blue dye What are the ecological causes and indigo and its structural isomer indirubin, which consequences of variation in phytochemical is considered to be the active ingredient in diversity within and between plant taxa? antileukemic Chinese herbal preparations. Despite decades of natural products Oxidation of indole can produce indigo and discovery by organic chemists and research indirubin, but the enzymes that catalyze this by chemical ecologists, our understanding of reaction in plants are unknown. We cloned phytochemically-mediated ecological processes a P450 gene from I. tinctoria and expressed in natural communities has been restricted it in E. coli, which then produced a water- to either broad classes of compounds or a insoluble blue precipitate. Extraction of these small number of well-characterized molecules. bacterial cultures with chloroform and dimethyl Until now, very few studies have assessed formamide, followed by HPLC and UV-Vis the ecological causes or consequences of spectrophotometry, showed the presence of quantified phytochemical diversity across both indigo and indirubin. Indigo production taxa in natural systems. Consequently, was increased approximately 3-fold when the hypotheses that attempt to explain variation P450 is coexpressed with isatin hydrolase (from in phytochemical diversity among plant taxa Pseudomonas), an enzyme that degrades isatin

August 8–12, 2015 • University of Illinois at Urbana-Champaign 63 to isatic acid. Meanwhile, indirubin production identification revealed that the S20G+T21S was improved by supplementation with 500 mutant protein widened the substrate and mg/l of 2-oxindole and 100 mg/l of isatin. We regio-specificity of CP3OGT. Docking analysis now propose that P450 genes are involved revealed that H154 and Q87 could be involved in the biosynthesis of indigo precursors and in orienting the ligand molecules within the indirubin in plants. acceptor binding site. H363, S20, and S150 were also found to make close contact with the [P39] Structure-function investigations 7-OH, 4-OH and 3’-OH groups, respectively. of site directed mutants of Citrus paradisi flavonol specific 3 O [P40] The Effects of Coconut Water as glucosyltransferase—Impact of an Undefined Media Amendment on the mutations of serine, histidine and Growth of Panax Quinquefolius Calli. glutamine residues Presenter(s): Shannon Smith, Middle Tennessee Presenter(s): Preethi Sathanantham, East Tennessee State University State University at Johnson City Author(s): Shannon Smith, Middle Tennessee State Author(s): ShivaKumar Devaiah, East Tennessee University State University at Johnson City and Cecilia Dr. John Dubois, Middle Tennessee State University McIntosh, East Tennessee State University at Dr. Elliot Altman, Middle Tennessee State University Johnson City Panax quinquefolius, or American ginseng, has Glucosyltransferases (GTs) are enzymes been harvested and cultivated for generations. that enable transfer of glucose from an Research is ongoing to illuminate the activated donor (UDP-glucose) to the relationship between secondary metabolites acceptor substrates. A flavonol specific present and physical responses to the glucosyltransferase cloned from Citrus paradisi metabolites in humans. Due to the recalcitrance has strict substrate and regiospecificity of the plant to tissue culture, research into (Cp3OGT). The amino acid sequence of the optimization of quality tissue production Cp3OGT was aligned with a purported for scientific analysis has been undertaken. anthocyanin GT from Clitorea ternatea and Calli were initiated for experimentation, and a GT from Vitis vinifera that can glucosylate initial results pointed to the beneficial affects both flavonols and anthocyanidins. Using certain media amendments had on tissue homology modeling to identify candidate growth. A study was performed to determine regions followed by site directed mutagenesis, if the promising media mixture has a long term three double mutations of Cp3OGT were statistically significant effect on the growth made. Biochemical analysis of the three of calli. Samples of calli were weighed and mutant proteins was performed. S20G+T21S plated on media containing the undefined protein retained activity similar to the wildtype nutrient supplement (+S media), and media (WT- Kmapp-80 µM; Vmax = 16.5 pkat/µg, without the undefined nutrient supplement Mutant- Kmapp-83 µM; Vmax -11 pkat/µg) but (-S media). Differences in growth rate, overall the mutant was more thermostable compared mass, phenolic production, and culture count to the WT and this mutation broadened its were tracked over a six month period, with substrate acceptance to include the flavanone, cultures passaged once every four weeks. naringenin. S290C+S319A mutant protein Data were statistically analyzed via Sigma retained 40% activity relative to wildtype, had Stat. Results suggest the media supplement an optimum pH shift, but had no change in does affect the calli, giving an initial boost substrate specificity (Kmapp-18 µM; Vmax-0.5 in growth with effects tapering off over time. pkat/µg). H154Y+Q87I protein was inactive The initial differences were sufficient to result with every class of flavonoid tested. Product in the variance in mass increase between

64 The Phytochemical Society of North America the two media types. This information has is influenced by various biotic and abiotic been used to better optimize tissue growth stress factors which implicates SIR2 plant protocols, supplying tissue for chemical analysis stress signaling. Transgenic tobacco plant with and experiments to optimize metabolite altered expression (RNAi-mediated silencing production. and inducible overexpression of SBIP-428) is being developed which will be used to study [P41] Characterization of a Tobacco SIR2 its role in stress signaling in tobacco plants. like Enzyme for its Role in Plant Stress Understanding the role of SBIP-428 is likely Signaling to implicate SABP2/SA in regulating various Presenter(s): Bal Krishna Chand Thakuri, East plant stress signaling pathways via acetylation/ Tennessee State University deacetylation. Author(s): Mackenzie Davenport1, East Tennessee State University [P42] Probing the Volatile Terpenome Md Imdadul Haq, East Tennessee State University of Switchgrass Roots in Interaction with Dhirendra Kumar, East Tennessee State University Rhizobacteria SBIP-428 is a SABP2-interacting tobacco Presenter(s): Dorothea Tholl, Department of Biological Sciences, Virginia Tech protein identified in yeast two-hybrid screen. It shows high similarity to SIR2 (Silent Author(s): Andrew Muchlinski, Department of Biological Sciences, Virginia Tech, 432 Latham Hall, Information Regulator 2) like proteins. SIR2 220 Agquad Lane, Blacksburg, VA 24061, USA enzymes exhibit NAD+-dependent deacetylase Feng Chen, Department of Plant Sciences, activity by catalyzing the deacetylation of University of Tennessee, 2431 Joe Johnson Dr., post-translationally acetylated lysine residue Knoxville, TN 37996, USA in histones as well as non-histone proteins. Richard Rodrigues, Department of Horticulture, SABP2 is critical for inducing systemic Virginia Tech, 301 Latham Hall, 220 Agquad Lane, acquired resistance (SAR) and local resistance Blacksburg, VA 24061, USA in tobacco and other plants. By its esterase Mark Williams, Department of Horticulture, activity, SABP2 catalyzes the conversion of Virginia Tech, 301 Latham Hall, 220 Agquad Lane, methyl salicylate to salicylic acid (SA), a plant Blacksburg, VA 24061, USA hormone which plays an important role in plant Dorothea Tholl, Department of Biological Sciences, defense system. Arabidopsis SRT2 (AtSRT2) Virginia Tech, 409 Latham Hall, 220 Agquad Lane, negatively regulates the basal defenses by Blacksburg, VA 24061, USA inhibiting the biosynthesis of SA with no Plant roots sustain intimate relationships with visible effect in growth phenotype, indicating diverse microbiota that contribute to plant its possible role in plant immunity. A large productivity and resilience. The plant traits number of proteins/enzymes implicated that define the composition of root microbial in various metabolic pathways present in communities by “habitat” filtering are poorly cytoplasm and various organelles in plants understood. Volatile organic compounds and other organisms have been found to be (VOCs) such as terpenoids affect colonization lysine acetylated. SBIP-428 was cloned from by epiphytic bacteria in aboveground tissues tobacco leaves and heterologously expressed (Junker and Tholl, 2013). However, the role of in E. coli. Interestingly the lyse residue of VOCs in interaction of roots with epiphytic and recombinant SBIP-428 expressed in E.coli was endophytic microbes is largely unknown. We found to be in acetylated, which opens up the investigate the function of volatile terpenoids possibility of its post-translational regulatory as host-specific chemo-selective factors in the role in the expression of the various non- rhizosphere by using the native prairie grass, histone protein present in plant cell including switchgrass (Panicum virgatum), as a model defense related proteins. Expression of SIR2 system. Root volatile profiling of multiple

August 8–12, 2015 • University of Illinois at Urbana-Champaign 65 switchgrass cultivars demonstrated qualitative NAEs that makes it relevant to elucidate the and quantitative variation of several terpenoid conserved metabolic pathway of NAEs in crop compounds. By contrast, the monoterpenoids, species such as tomato. It is hypothesized camphor and borneol, were highly conserved that there is a functional FAAH in tomato among cultivars suggesting an important role that hydrolyzes NAEs. To test this hypothesis, for these VOCs in belowground interactions. AtFAAH was used as a template to identify Camphor is known for its antimicrobial and putative FAAH sequences in tomato, using insect repellent effects but also serves as BLASTX. Six SlFAAH sequences with the the exclusive carbon source of soil-borne conserved amidase signature sequence and microbes. The accumulation of camphor the catalytic triad, formed by Lys205, Ser281, was stimulated by inoculation of switchgrass and Ser305 in AtFAAH, were identified. seedlings with nitrogen-fixing rhizobacteria, Phylogenetic analysis of putative SlFAAH which supports the role of this compound as a homologs and other FAAH family proteins potential chemical mediator in the rhizosphere. (Arabidopsis, rice and moss), using CLUSTALW, Following a root transcriptome analysis, revealed the two sequences that are closely we are currently identifying root-expressed related to the functionally characterized terpene synthase genes to subsequently AtFAAH1. Using molecular visualization generate terpenoid (camphor) biosynthetic system (PyMOL), protein structures of putative mutants and analyze associated changes of SlFAAH1and 2 were predicted and compared their root microbiomes. These studies will help with AtFAAH; both sequences showed similar define VOC-based niches of microbes in the domain structure to AtFAAH, with minor rhizosphere. differences in spatial arrangement. For further biochemical characterization, full-length Junker R. and D. Tholl (2013). Volatile Organic coding sequence of SlFAAH1 and SlFAAH2 Compound Mediated Interactions at the Plant- were isolated and cloned into a heterologous Microbe Interface. Journal of Chemical Ecology expression system. The expressed protein 39: 810-825. will be characterized for its hydrolytic activity against radiolabelled NAE substrates. [P43] Identification and In-silico Analysis Furthermore, transcript levels for SlFAAH1 and of Fatty Acid Amide Hydrolases in SlFAAH2 will be quantified and correlated with Tomato the NAE levels in various tissues to predict Presenter(s): Vijay Tiwari, East Tennessee State their role in tissue-specific NAE hydrolysis. University Together, these molecular and biochemical Author(s): Vijay Tiwari, Derek Stuffle and Aruna characterization studies in tomato are expected Kilaru, East Tennessee State University to further validate the conserved nature of NAE N-acylethanolamines (NAEs) are a metabolic pathway in plants. family of signaling lipids derived from a minor membrane lipid constituent [P44] Biosynthesis Enhancement and N-acylphosphatidylethanolamine (NAPE). Purification of Bioactive Prenylated In Arabidopsis, NAE mediates physiological Stilbenoids from Hairy Root Cultures functions such as seedling growth, flowering, of Peanut by High Performance and response to stress via abscisic acid (ABA) Countercurrent Chromatography –dependent and –independent signaling Presenter(s): Christopher Aaron Tollett, Arkansas pathways. The function of NAEs is terminated State University by a highly conserved fatty acid amide Author(s): Carson Day, Arkansas State University hydrolase (FAAH). Studies in model plant Tianhong Yang, Arkansas State University Luis Arabidopsis showed the significant role of

66 The Phytochemical Society of North America Nopo-Olazabal, Arkansas State University and Service, Elvira Gonzalez de Mejia, University of Fabricio Medina-Bolivar, Arkansas State University Illinois at Urbana-Champaign, Erika Garcia Chavez, Stilbenoids are polyphenolic compounds University of San Luis Potosi, and Celia Aradillas Garcia, University of San Luis Potosi found in plants like peanuts and grapes which have a wide range of biological effects Low-grade inflammation has been widely and potential benefits to human health. related with metabolic disorders and chronic Among the peanut stilbenoids is arachidin-1, diseases, such as insulin resistance, obesity, a prenylated analog of piceatannol which type 2 diabetes, cardiovascular diseases has shown higher antioxidant activity and and cancer. The tea prepared with Calea cytotoxicity in different cancer cell lines and urticifolia leaves is used traditionally like a potentially higher metabolic stability than remedy to treat gastric ulcers, diabetes and other stilbenoids. In order to produce this and inflammatory processes by the Xi’uy ancient other prenylated stilbenoids, peanut hairy native community of San Luis Potosi, Mexico. root cultures were co-treated with multiple The objective of this study was to characterize elicitors for different periods of time. Ethyl the phenolic compounds of Calea urticifolia acetate extracts were prepared from the culture lyophilized aqueous extract (CuAqE) and medium of the elicited cultures and analyzed evaluate in vitro its anti-inflammatory and anti- by HPLC. In addition to previously described oxidant properties in lipopolysaccharide (LPS)- stilbenoids, potentially novel stilbenoids stimulated RAW 264.7 macrophages. Phenolic were also detected in the culture medium. To compounds of CuAqE were characterized further study the biological activity of these by LC-ESI-MS. RAW 264.7 macrophages compounds, they are being purified using high were treated with CuAqE (10, 25, 50, 75 performance counter current chromatography and 100 mg/mL) and stimulated with LPS. (HPCCC). Initially, the distribution constant of Nitric oxide (NO) release, inducible nitric the desired compounds is determined using a oxide synthase, prostaglandin E2 production, solvent system composed of different volumes cyclooxygenasa-2, p65, p50 and reactive of hexane, ethyl acetate, methanol and water. oxygen species (ROS) were measured. A mix of Pooled extracts from the culture medium caffeoylquinic acid derivatives and flavonoid- are then separated by HPCCC and fractions glucoside was found in CuAqE. NO (5.3, corresponding to the various compounds 17.9, 57.4, 90.2 and 103.2% respectively) and are analyzed by HPLC to assess purity. Our ROS (94.4, 99.3, 108.1, 108.4 and 108.1% preliminary studies have shown that the levels respectively) production were significantly of prenylated stilbenoids can be increased (P<0.05) inhibited in a dose-response manner several fold by co-treatments with different in LPS-stimulated RAW 264.7 macrophages. elicitors and that HPCCC can be used to obtain Therefore, phenolic compounds in CuAqE selected prenylated stilbenoids with high levels could be responsible to inhibit LPS-induced of purity. inflammation through ROS/NO reduction. In conclusion, results support the traditional [P45] Calea urticifolia lyophilized knowledge of Calea uritifolia tea such as aqueous extract inhibits an anti-inflammatory agent and suggested lipopolysaccharide-induced inflammation antioxidant properties. in RAW 264.7 macrophages. [P46] Sub-chronic oral toxicity of Calea Presenter(s): Maria Lucina Torres Rodriguez, University of San Luis Potosí urticifolia lyophilized aqueous extract in Author(s): Maria Lucina Torres Rodriguez, University Wistar rats. of San Luis Potosí, Mark Berhow, United State Presenter(s): Maria Lucina Torres Rodríguez, Department Agriculture, Agricurtural Research University of San Luis Potosi

August 8–12, 2015 • University of Illinois at Urbana-Champaign 67 Author(s): Maria Lucina Torres Rodríguez, University [P47] High-throughput Phenotyping of of San Luis Potosi and Erika Garcia Chavez, Rice Lines Within a Rice Diversity Panel University of San Luis Potosi to Determine Salinity Tolerance Calea urticifolia is a shrub of the Asteraceae Presenter(s): Nathan Sullivan Tripod, Research family and is used by the Xi’uy ethnic group Assistant , Arkansas Bioscience Institute of San Luis Potosi, Mexico. The tea prepared Author(s): Nathan Tripod1, Zachary Campbell1, with the leaves is used such as remedy to Malachi Campbell2, Harkamal Walia2, Argelia treat gastric ulcers, diabetes and inflammatory Lorence1,3 processes. However, toxicity studies of aqueous 1Arkansas Biosciences Institute, Arkansas State extract have not been reported. Therefore, University, 504 University Loop, Jonesboro, AR the objective of this research was to evaluate 72401, Craighead County; sub chronic toxicity in vivo of Calea urticifolia 2University of Nebraska, 1400 R Street, Lincoln, NE, lyophilized aqueous extract (CuAqE) as part of 68588, Lancaster County; its pre-clinical studies. Male and female young 3Department of Chemistry and Physics, P.O. Box adult Wistar rats (180-200 g) were treated 419, State University, AR, 72467, Craighead County with 0.5, 50 and 500 mg/kg of CuAqE by Rice is currently one of the most important gavage during 28 days. Mortality incidence, crops for global food security. A significant clinical signs, body-weight gain changes, portion of optimal rice producing areas are food and water consumption were registered prone to climate change and rising sea levels. every day. Organ weights, biochemical (urea, This is causing the soil in these rice producing creatinine, alanine transaminase and aspartate areas to become increasingly saline over time. transaminase) and hematological (white blood Eventually the lands become too saline to cells, red blood cells, platelets, hematocrit, produce the crops and become unutilized. haemoglobin, mean corpuscular volume, mean In order to keep meeting rice demand there corpuscular haemoglobin) parameters were is a need to develop new rice varieties that measured at the end of study. Sub chronic oral can thrive under saline conditions. This work administration of CuAqE during 28 days did is a collaborative effort between the Walia not produce both mortality and clinical signs. Laboratory at the University of Nebraska Food and water consumption, organ weights, Lincoln and the Lorence Group at Arkansas biochemical and hematological parameters State University. The Walia team has been showed not-significant changes in both male studying the salt tolerance of a rice diversity and female rats (P > 0.05). Body weight gain panel at two developmental stages that was significantly (P < 0.05) lower than control are particularly susceptible to this stress in group (distilled water), when was administered greenhouse experiments. Based on their 50 mg/kg at 21 and 28 day (28 ± 25 % and results, a selected group of rice lines including 44 ± 25 % respectively) and 500 mg/kg at 7, salt tolerant and salt sensitive types as well as 14, 21 and 28 day(95 ± 66 %, 64 ± 45 %, 59 ± positive and negative controls were sent to the 31 %, 64 ± 19 % respectively) in female rats. Lorence team to study their response to salt Eventually, results of this study support that stress at the early vegetative growth stage in CuAqE has a low grade of toxicity. Therefore, greenhouse experiments. We have utilized a the traditional dose (0.55 mg CuAqE/kg body powerful high throughput phenotyping system, weight) of Calea urticifolia tea can be safely the Scanalyzer HTS, in order to acquire digital consumed. images using visible and florescence cameras to quantify the tolerance to salt of over 85 rice accessions grown in hydroponic conditions. In this work we will present our progress on

68 The Phytochemical Society of North America the identification and characterization of salt Author(s): Neil P.J. Price, Karl E. Vermilion, Fred J. tolerant rice lines. Eller and Steven F. Vaughn, USDA, ARS, NCAUR, 1815 N. University St., Peoria, IL 61604 [P48] Isolation of bioactive compounds A novel arabinogalactan polysaccharide of Piper holdridgeanum identified to (FGP) is described that is produced in large cause inhibition of yeast cell growth quantities from the cut stems of Frost grape Presenter(s): Federico Urbano Muñoz, University of (Vitis riparia Michx.). The sugar composition Nevada Reno consists of L-arabinofuranose (L-Araf, 55.2 %) Author(s): F. Urbano-Muñoz, Christopher .S. Jeffrey, and D-galactopyranose (D-Galp 30.1%), with Lora .A. Richards, and Ian .S. Wallace, University of smaller components of D-xylose (11.2 %), Nevada Reno D-mannose (3.5%), and glucuronic acid (GlcA, Natural products isolated from an array of ~2%), the latter linked via a galactosyl residue. organisms have served as templates for Permethylation identified 3-linked Galp medicine and many other applications. Piper residues, some substituted at the 2-position is a phytochemically diverse genus, which with Galp and Manp, terminal Araf and Xylp, has not fully been chemically explored. One and internal 3-substituted Arap. NMR identified major challenge lies in identifying bioactive βGalp and three αAraf spin systems, in a molecules from crude extracts. To identify Araf-α1,3-Araf-α1,2-Araf-α1,2-Galp structural possible bioactive compounds found in a motif. Diffusion-orientated NMR showed that species of Piper, a series of assays were the FGP has a molecular weight of 1 - 10 MDa. performed to determine which species crude Unlike gum arabic, the FGP does not contain extract demonstrated growth inhibition on a hydroxyproline-rich protein. FGP forms Arabidopsis thaliana Col-0 root length, and stable gels at >15 % w/v, and solutions at cells of Saccharomyces cerevisiae S288c,and lower concentrations (1 - 12% w/v) are viscous, Escherichia coli DH5α. We combined these and are excellent emulsifiiers of flavoring screens with NMR based metabolomics oils (grapefruit, clove, and lemongrass), analysis to determine possible candidates giving stable emulsions for ≥72 hours. Lower for further isolation of bioactive compounds. concentrations (0.1% w/v) were less viscous, yet From the 31 species screened in triplicate still gave stable grapefruit oil/water emulsions. we found high inhibition levels of cell These properties indicate that FGP is a β-1,3- growth of S. cerevisiae S288c grown in the linked arabinogalactan with potential as a gum presence of Piper holdridgeanum extract. arabic replacement in the food and beverage In further investigations of these species we industries. found that P. holdridgeanum produces two sesquiterpenes and two unknown phytol [P50] Evaluation of the Cytotoxic isomers. Future studies will focus on employing Activity of Origanum Vulgare bioassay guided fractionation to identify Presenter(s): Tibebe Woldemariam, California active metabolites and more comprehensive Northstate College of Pharmacy bioassays of isolated compounds to determine Author(s): Tibebe Woldemariam and david Pearson, if synergistic effects are resulting in enhanced California Northstate College of Pharmacy bioactivity. Herbs used for flavoring and scents have been shown to be a rich source of phytochemicals [P49] A novel emulsion-forming and are getting much attention as a potential arabinogalactan gum from the stems of source of cancer chemoprotective agents. Frost grape (Vitis riparia Michx.) Strong epidemiological evidence also suggests Presenter(s): Steven F. Vaughn, USDA, ARS, NCAUR that regular ingestion of herbs and spices can reduce the risk of cancer. The culinary herb

August 8–12, 2015 • University of Illinois at Urbana-Champaign 69 oregano Origanum vulgare for example has Most of pathway genes are upregulated been demonstrated to have antimicrobrial to form an engineered pathway toward activity along anti-inflammatory and wound anthocyanins. Eleven anthocyanin molecules healing activity. Oregano was also reported to are detected from extracts of engineered contain anti prostate cancer activity. plants. One anthocyanin molecule accounts for approximately 98% of total anthocyanins. In the present study, Carvacrol derived from HPLC-qTOF-MS/MS analysis identifies Oregano methanolic extract was isolated and that the dominant anthocyanin is cyanidin assessed for possible in vitro cytotoxic activity 3-O-rutinoside. This result implies a preferred against a panel of pancreatic cell lines and effect of PAP1 regulation molecular dominance non-small cell lung cancer cell line A549 and of anthocyanins in dark tobacco plants. PANC-10. Bioactivity-guided fractionation of the [P52] Metabolic diversity during methanol extract afforded several fractions ontogeny of Piper species and compounds which showed significant Presenter(s): Lydia Fumiko Yamaguchi, University of reductions in cell viability. The major São Paulo compound, carvacrol, was found to be Author(s): Lydia Fumiko Yamaguchi, University of significantly active against both lung and São Paulo; Anderson M. Gaia, University of São pancreatic cell lines. The results of these Paulo; Christopher S. Jeffrey, University of Reno; studies including the bioassay-guided Massuo J. Kato, University of São Paulo fractionation of the methanolic extracts, Tropical forests comprise almost 5% of Earth isolation of carvacrol and cytotoxic activity of surface and with estimates of at least 3 million the fractions and compounds will be presented. of organisms. Astonishingly only about a sixth of these are known to science. The Piper These outcomes highlight the prominence of species have pantropical distribution and the use of herbs and spices as possible sources considered as model system to understand for prevention and treatment of cancer. the intricate net of ecological interactions in tropical ecosystems. The investigation of the [P51] Anthocyanin biosynthesis in red- dynamics in secondary metabolism during colored dark tobacco plants activated by ontogeny in plants would provide essential PAP1/MYB75 clues for understanding their interactions Presenter(s): Xianzhi He, Department of Plant and with herbivores. Ten species of Piper were Microbial Biology, North Carolina State University, analysed and three groups of chemical NC 27695, USA, profile were observed; for species producing Author(s): De-Yu Xie, Department of Plant and amides in seedling stages (P. tuberculatum, Microbial Biology, North Carolina State University, NC 27695, USA, [email protected] P. amalago and P. reticulatum) no differences in the composition were observed during PAP1/MYB75 is master regulator controlling the development. In the second group (P. anthocyanin biosynthesis in Arabidopsis gaudichaudianum, P. solmsianum, P. regnellii, thaliana. Its regulatory function requires P. caldense and P. hemmendorfii) allylphenols transcription factor partner, such as TT8/ were detected as major compounds in the bHLH and TTG1/WD40. Although PAP1 can seedling leaves while adult leaves contained activate anthocyanin pathway, anthocyanin lignans, neolignans or prenylated benzoic molecule diversities depend upon plant acids. Additionally, in a third group (P. species. In this study, we introduce PAP1 into richardiaefolium and P. permucronatum) amides two dark tobacco varieties. The anthocyanin were observed as major compounds in the pathway is activated by PAP1 overexpression.

70 The Phytochemical Society of North America seedling leaves instead of lignans or flavonoids show that suspension cultured cells contain found in the adult leaves (Fig. 1). Such disparity certain catalytic activities to convert catechin in the metabolic profile between different and epicatechin. We will continue red cell stages could be related to several different suspension culture to characterize whether the adaptive strategy to increase the survival rate unknown catalytic activities are associated with of the seedlings. PA biosynthesis.

[P53] Suspension culture of transgenic [P54] Low-Temperature Conditioning red tobacco cells to understand of “Seed” Cloves Induces Changes in proanthocyanidin biosynthesis Protein Profile, Enhances Expression Presenter(s): Seyit Yuzuak, North Carolina State of Phenolic Metabolism Genes and University Anthocyanin Content in ‘Coreano’ Author(s): Seyit Yuzuak and De-Yu Xie, North Garlic (Allium sativum) during Plant Carolina State University Development. Proanthocyanidins (PAs) are synthesized in Presenter: Miguel David Dufoo-Hurtado, the fruits, leaves, barks and seeds of many Universidad Autónoma de Querétaro, University of plants. They are oligomers or polymers of Illinois at Urbana-Champaign 1,2 flavonoids resulting from polymerization of Authors: Miguel David Dufoo-Hurtado , Karla 1 3 flavan-3-ol units (e.g., epicatechin, catechin) Guadalupe Zavala-Gutiérrez , Cong-Mei Cao , Luis Cisneros-Zevallos3, Ana Paulina Barba de la Rosa4, and characterized by diverse structures. José Ángel Huerta-Ocampo4,5, Edmundo Mercado- PAs can prevent plants from pathogens and Silva1 radiation-caused damages. PAs and their 1Departamento de Investigación y Posgrado en monomers are also strong antioxidant nutrients Alimentos, Universidad Autónoma de Querétaro, having multiple beneficial effects to human Centro Universitario S/N Las Campanas health, such as protective activities against 2Department of Natural Resources and cardiovascular, cancer and Alzheimer’s diseases. Environmental Sciences, University of Illinois at In food products, PAs give an astringent Urbana-Champaign flavor. To date, the biosynthetic pathway of 3Department of Horticultural Sciences, Texas A&M PAs has gained intensive understanding from University phenylalanine through leucoanthocyanidins 4División de Biología Molecular, Instituto Potosino to flavan-3-ols. Two late pathways, the LAR de Investigación Científica y Tecnológica (Leucoanthocyanidin reductase) and the ANR 5Programa Cátedras CONACYT, Centro de (Anthocyanidin reductase) pathways, have Investigación en Alimentación y Desarrollo A.C. been identified from leucoanthocyanidins to Low-temperature conditioning of garlic “seed” flavan-3-ols. The ANR pathway has allowed cloves at 5 °C for 5 weeks accelerated the the success of metabolic engineering of PAs development of the crop cycle, decreased in anthocyanin-producing cells of plants. plant growth, and increased the synthesis However, mechanisms of PA polymerization of phenolic compounds and anthocyanins remain completely unknown. In this report, in outer scale leaves of the bulbs at harvest we show establishment of suspension of red time, leading to 3-fold content increase transgenic tobacco cells. Four to six liters compared with those conditioned at room of suspension culture were developed to temperature. Cold conditioning of “seed” obtain red cells with a high production of cloves also altered the anthocyanin profile anthocyanins culture. Different methods are during bulb development and at harvest. Two tested to understand enzymatic activities new anthocyanins are reported for the first time of red cells. Our preliminary experiments in garlic. The high phenolic and anthocyanin

August 8–12, 2015 • University of Illinois at Urbana-Champaign 71 contents in bulbs of cloves conditioned at 5 °C mice fed a 60% high fat diet for ten weeks. for 5 weeks were preceded by overexpression C57BL/6j mice were randomized into six groups of putative genes of the phenolic metabolism and allowed to drink (ad libitum) an alcohol- [6-fold for phenylalanine ammonia lyase free blackberry-blueberry beverage (AFFB), (PAL)] and anthocyanin synthesis [1-fold for three doses of a phenolic extract (PAE) from UDP-sugar:flavonoid 3-O-glycosyltransferase AFFB at 1 (0.1 X), or 9 (1X) or 19 (2X) mg ANC/ (UFGT)] compared with those conditioned at kg BW/d anthocyanin (ANC) concentrations, room temperature. Results also revealed that sitagliptin (hypoglycemic positive control), or low-temperature conditioning of garlic “seed” water (negative control). Changes in expression cloves causes alterations in the accumulation of mice genes were analyzed using an Illumina of proteins involved in cellular growth, MouseWG-6 v2.0 Expression BeadChip. As antioxidative/oxidative state, macromolecules determined in the heatmaps, there was a trend transport, protein folding and transcription to shift the differentially expressed genes (DEG) regulation process. Thus, low-temperature profile with the water and sitagliptin groups conditioning affected protein biosynthesis having similar patterns, but different from AFFB and quality control system, photosynthesis, (1 DEG), 0.1X (29 DEGs), 1X (202 DEGs) and photorespiration, energy production, and 2X (123 DEGs). The top up-regulated genes for carbohydrate and nucleotide metabolism. 0.1X, 1X and 2X were Acot1 (FC=7.42), REEP1 These processes can work cooperatively to (FC= 3.84) and REEP2 (FC=3.88) respectively, establish a new cellular homeostasis that while the top down-regulated genes were might be related with the physiological and GPER (FC=-1.90), GCNT1 (FC=-2.69) and biochemical changes observed during cold RPRM (FC=-2.53) respectively. These genes conditioning and plant development. are related to fatty acid oxidation, cell surface proteins expression and cell proliferation. The [P55] Phenolics in alcohol-free fermented most significantly affected pathways caused blueberry-blackberry beverage by PAE included amino acid degradation caused differential gene expression in and biosynthesis such as methionine, glycine pancreatic tissue of C57BL/6J mice fed and cysteine, amino acids known to have high-fat diet a negative impact in insulin secretion. On Presenter: Diego Luna-Vital, University of Illinois at the other hand, some genes with important Urbana-Champaign roles in carbohydrate metabolism such as Authors: D.A. Luna-Vital1,2, M. Johnson3, E. de ME1, ME2, PGD, GALK1, GPD1 and GPD2 Mejia1,3 were down regulated as a response to PAE 1Department of Food Science and Human Nutrition, treatment (FDR=0.3). Additionally, treatments University of Illinois at Urbana-Champaign, 228 caused a significant decrease of differential ERML, 1201 W Gregory Drive, Urbana, IL 61801, gene expression related to concentration of USA glucose, according to IPA analysis (Z < - 2). 2Research and Graduate Studies in Food Science, Results suggested that in the pancreas, PAE School of Chemistry, Universidad Autónoma de contributed to the antidiabetic effect of the Querétaro, Centro Universitario Cerro de las AFFB modulating genes related to glucose Campanas S/N. Querétaro, Qro 76010, Mexico. homeostasis and amino acid metabolism. 3Division of Nutritional Sciences, University of Illinois at Urbana-Champaign The aim of this research was to determine the potential of phenolic compounds from a fermented blackberry-blueberry beverage to modulate the pancreatic gene expression in

72 The Phytochemical Society of North America [P56] Effect of sulfur dioxide and lactic and Saturation were not significantly correlated acid during corn steeping process on to any of the variables measured. This study extraction of anthocyanins from purple proposes an economical and novel procedure corn pericarp to enhance total anthocyanin extraction from Presenter: Qian Li, Department of Food Science and corn pericarp. Human Nutrition, University of Illinois at Urbana- Champaign [P57] Phytohormones production during Qian Li1, Pavan Somavat2, Vijay Singh2, Elvira floral transition of soybean 1 Gonzalez de Mejia Presenter: Prem L. Bhalla, The University of 1Department of Food Science and Human Nutrition, Melbourne University of Illinois at Urbana-Champaign, IL, Authors: Prem L. Bhalla, Chui E. Wong, Mohan B. 61801, United States Singh, The University of Melbourne 2Department of Agricultural and Biological Soybean is a major legume crop and a Engineering, University of Illinois at Urbana- Champaign, IL, 61801, United States significant source of dietary proteins and edible oil. Flowering process governs seed set Purple corn is rich in natural pigments such and hence affects crop productivity. Soybean as anthocyanins, which can potentially be is a short-day plant grown broadly across used as an alternative to artificial colorants in the latitude but with each cultivar having a food. Anthocyanins can be extracted during narrow range of the north to south adaptation. corn processing, which usually generates This geographic adaptation of soybean is anthocyanin-rich coproducts. Our previous data likely a result of diversity associated with a shows that steeping water from wet-milling large number of genes regulating soybean process contains 79.1% of total anthocyanins flowering behavior. We used an integrated of the whole corn. Therefore, the objective of bioinformatic and experimental approach this study was to optimize the extraction of to understanding the molecular control of anthocyanin from purple corn pericarp steeped soybean flower initiation. RNA-Seq analysis using common wet-milling chemicals, sodium of shoot apical meristem and leaf undergoing metabisulfite (Na S O ) and lactic acid (L.A.). 2 2 5 floral transition revealed phytohormones Two thousand ppm SO treated steeping water 2 might play predominant roles in the short-day- contained total anthocyanin (TA) equivalent to induced floral initiation process. ABA level was 20.5 ± 1.5 mg cyanidin-3-glucoside (C3G)/g measured using an immunoassay at the shoot pericarp, which was significantly higher than apical meristem showed a significant increase only water for steeping (7.1 ± 0.6 mg C3G/g in ABA during floral transition. Further, our pericarp, p < 0.05). Lactic acid did not change data indicated a major reprogramming events TA concentration. But the combination of SO 2 in leaves and the shoot apical meristem that and lactic acid significantly increased TA up point toward phytohormones gibberellins and to 22.9 ± 0.2 mg C3G/g pericarp (p < 0.05). cytokinin as key regulators in the production of HPLC revealed that this greater extraction was flowering signal(s) in leaves. associated with increased C3G and decreased condensed forms of anthocyanins as compared to only using water. Chroma was highly correlated to the concentration of anthocyanins (r = 0.999) and tannin (r = 0.994) and Hue angle

August 8–12, 2015 • University of Illinois at Urbana-Champaign 73 Participant List (as of 7/22/15)

Abourashed, Ehab Audam, Timothy Chicago State University East Tennessee State University Department of Pharmaceutical Sciences 811 Magnolie Ext, 9501 S. King Drive / DH 206 Apartment 47 Chicago, IL 60628 USA Johnson City, TN 37614 USA 773-821-2159 423-676 8382 [email protected] [email protected] Acosta-Gamboa, Lucia Becker, Talon Arkansas State Univeristy University of Illinois at Urbana-Champaign PO Box 1144 1102 S Goodwin State University, AR 72467 USA Turner Hall 501-415-6810 Urbana, IL 61801 USA [email protected] 815-973-5734 [email protected] Ajewole, Ebenezer University of Western Ontario / Agriculture and Berenbaum, May Agri-Food Canada University of Illinois at Urbana-Champaign 1394 Basswood Rd 320 Morrill Hall London, ON N5V3T3 Canada 505 S Goodwin 519-494-9819 Urbana, IL 6801-3795 USA [email protected] 217-333-7784 [email protected] Alexander, Molly Plant Powered Production (P3) Center Berhow, Mark Biological Sciences/Chemistry and Physics USDA/ARS/NCAUR P.O. Box 639 1815 N. University St. State University, AR 72467 USA Peoria, IL 61604 USA 870-680-4344 309-681-6347 [email protected] [email protected] Allen, Charlotte Burton, Tristesse Kraft Foods University of Illinois at Chicago 801 Waukegan Rd 833 South Wood Street Glenview, IL 60025 USA Chicago, IL 60612 USA 847-968-4565 773-593-5779 [email protected] [email protected] Altaweel, Areej Cai, Qing King Saud University City University of New York–City College Riyadh 160 Convent Ave. Riyadh, Kingdom of Saudi Arabia 11447 Saudi New York, NY 10031 USA Arabia 212-650-8798 00966504773399 [email protected] [email protected]

74 The Phytochemical Society of North America Chapman, Kent Clay, Nicole University of North Texas Yale University Denton, TX 76203-5017 USA KBT 734 940-300-6961 266 Whitney Ave [email protected] New Haven, CT 06511 USA 203-595-1837 Chappell, Joe [email protected] University of Kentucky 789 S. Limestone St. Creameans, Jarrod Lexington, KY 40536 USA Arkansas Biosciences Institute/Arkansas State 859-218-0775 University [email protected] 811 Strawfloor Dr. Jonesboro, AR 72401 USA Chatham, Laura 870-530-6801 University of Illinois at Urbana-Champaign [email protected] 1201 W. Gregory Dr. 311 ERML Cuperlovic-Culf, Miroslava Urbana, IL 61801 USA National Research Council 309-838-9413 100 des Aboiteaux St. [email protected] Moncton, NB E1A 7R1 Canada 506-861-0952 Chen, Xiao-Ya [email protected] Shanghai Institute of Plant Physiology and Ecology, SIBS, CAS Dayan, Franck 300 Fenglin Road USDA/ARS Shanghai, China P.O. Box 1848 862154924033 University, MS 38677 USA [email protected] 662-915-1039 [email protected] Chilufya, Jedaidah East Tennessee State University de Mejia, Elvira 1301 Seminole Drive Apt 53B University of Illinois at Urbana-Champaign Johnson City, TN 37604 USA 228 ERML 770-330-7438 1201 W. Gregory Dr. [email protected] Urbana, IL 61801 USA 217-244-3196 Choudhary, Alka [email protected] NIPER S.A.S Nagar C-312, Department of Natural Prodcuts DellaPenna, Dean Mohali 160062 India Michigan State University 946-394-5544 East Lansing, MI 48824 USA [email protected] 517-432-9284 [email protected] Christoff Wouters, Felipe Max Planck Institute for Chemical Ecology Erlandson, Petra Hans-Knoell-Str 8 Alkemist Labs (distributor for Extrasynthese) Jena 07745 Germany 1260 Logan Ave #B2 +491634107926 Costa Mesa, CA 92626 USA [email protected] 714-754-4372 x 225 [email protected]

August 8–12, 2015 • University of Illinois at Urbana-Champaign 75 Evans, Bradley Jacobs, Tom Donald Danforth Plant Science Center PerkinElmer Inc. 975 North Warson Road 2000 York Rd. Saint Louis, MO 63132 USA Suite 132 314-587-1464 Oak Brook, IL 60523 USA [email protected] 847-420-6061 [email protected] Fauser, Jordan DePaul University Jeschke, Verena 3921 N.Hamilton Ave Max Planck Institute Chemical Ecology Chicago, IL 60618 USA Hans-Knöll-Str. 8 713-614-9656 Jena 07745 Germany [email protected] +49 3641 571327 [email protected] Gardner, Alicia University of Illinois at Urbana-Champaign Jetter, Reinhard 1201 W. Gregory Dr. 307 ERML MC-051 University of British Columbia Urbana, IL 61801 USA 6270 University Blvd. 309-444-0052 Vancouver, BC V6S1Z4 Canada [email protected] (001)6048222477 [email protected] Gershenzon, Jonathan Max Planck Institute for Chemical Ecology Jin, Liqing Hans-Knoell Strasse 8 City College of CUNY Jena D-07745 Germany 160 Convent Avenue +49 3641 571300 New York, NY 10031 USA [email protected] 212-650-8798 [email protected] Ghosh, Rajarshi Middle Tennessee State University Jinks, Angie 1301 E Main St. Thermo Scientific Murfreesboro, TN 37130 USA 355 River Oaks Parkway 850-776-4599 San Jose, CA 95134 USA [email protected] 408-499-4093 [email protected] Grover, Jagdeep National Institute of Pharmaceutical Education and Johnson, Todd Research University of Illinois at Urbana-Champaign NIPER S.A.S Nagar 320 Morrill Hall, 505 S. Goodwin Ave. Mohali 160062 India Urbana, IL 61801 USA 946-550-8115 610-984-5636 [email protected] [email protected] Ivanov, Dimitre Johnson-Ajinwo, Okiemute Rosa University of Western Ontario Keele University 1151 Richmond St. N. Newcastle-Under-Lyme, Staffordshire ST5 5BG London, ON N6A-5B7 Canada United Kingdom (519) 661-2111 86468 +447741755812 [email protected] [email protected]

76 The Phytochemical Society of North America Juvik, John Kilaru, Aruna University of Illinois at Urbana-Champaign East Tennessee State University 1201 W. Gregory 424 Headtown Rd 307 E.R. Madigan Lab Jonesborough, TN 37659 USA Urbana, IL 61801 USA 423-439-6931 217-333-1966 [email protected] [email protected] Kilgore, Matthew Kachroo, Pradeep Donald Danforth Plant Science Center University of Kentucky 975 N. Warson Rd. 1405 Veterans Drive St. Louis, MO 63132 USA Lexington, KY 40546 USA 859-200-1384 859-218-0729 [email protected] [email protected] King, Kathleen Kage, David East Tennessee State University Thermo Fisher Scientific 201 Lee Carter Dr. 1201 Wiley Road Johnson City, TN 37601 USA Suite 160 423-767-1152 Schaumburg, IL 60173 USA [email protected] 408-234-0285 Koo, Abraham [email protected] University of Missouri Kamboj, Pradeep 127 Schweitzer Hall JCDM College of Pharmacy Columbia, MO 65201 USA No. 81 Barnala Rd 573-882-9227 Sirsa, Haryana 125055 India [email protected] 735-700-0002 Kumar, Dhirendra [email protected] East Tennessee State University KAMBOJ, Sonia 423 Brown Hall, Box 70703 Satluj Public School Johnson City, TN 37614 USA Barnala Road 423-788-0143 Sirsa, 125055 India [email protected] 735-700-0077 Lak, Parnian [email protected] Oregon State University Kandel, Sangam 445 LPI East Tennessee State University Corvallis, OR 97333 USA 807 University Parkway 541-908-1823 Johnson City, TN 37614 USA [email protected] 423-439-6572 Lange, Mark [email protected] Washington State University Kato, Massuo IBC/Clark Hall University of São Paulo 100 Dairy Road Av. Prof. Lineu Prestes, 748 Pullman, WA 99164 USA São Paulo, 05508-000 Brazil 509-335-3863 +55 11 30913813 [email protected] [email protected]

August 8–12, 2015 • University of Illinois at Urbana-Champaign 77 Leal, Walter Luna-Vital, Diego University of California-Davis University of Illinois at Urbana-Champaign Department of Molecular and Cellular Biology 217-819-2715 1 Shields Avenue [email protected] Davis, CA 95616 USA Maeda, Hiroshi 530-752-7755 University of Wisconsin-Madison [email protected] 430 Lincoln Dr. Lee, Young-Sang Madison, WI 53706 USA Soonchunhyang University 608-262-5833 646 Shinchang-myon Eupnae-ri [email protected] ASAN, Chungnam 336-745 Republic of Korea Maresh, Justin +82-41-530-1287 DePaul University [email protected] 1110 W. Belden Ave. Li, Qian (Grace) Suite 100 University of Illinois at Urbana Champaign Chicago, IL 60614 USA 1201 W. Gregory Dr. 773-600-6058 228 ERML [email protected] Urbana, IL 61801 USA McIntosh, Cecilia 217-220-3502 East Tennessee State University [email protected] School of Graduate Studies LICAYAN, RICHARD Box 70720 New Use Agriculture & Natural Plant Products Johnson City, TN 37614 USA Program, Rutgers University 423-439-6147 59 Dudley Road, Foran Hall [email protected] New Brunswick, NJ 08901 USA Mojica, Luis 732-524-4855 University of Illinois at Urbana-Champaign [email protected] 1201 W. Gregory Dr. Lila, Mary Ann 228 ERML MC-051 North Carolina State University Urbana, IL 61801 USA Plants for Human Health Institute 312-208-4756 600 Laureate Way [email protected] Kannapolis, NC 28081 USA Nikolic, Dejan 704-250-5407 University of Illinois at Chicago [email protected] 833 S. Wood Street Long, Michael Chicago, IL 60612 USA 503 Chinkapin Court 312-413-5867 Schenectady, NY 12303 USA [email protected] 518-396-0770 Nomura, Taiji [email protected] Toyama Prefectural University Lorence, Argelia 5180 Kurokawa Arkansas State University Imizu, Toyama 939-0398 Japan PO Box 639 +81-766-56-7500 x 548 State University, AR 72467 USA [email protected] 870-680-4322 [email protected]

78 The Phytochemical Society of North America Odesina, Abdulkareem Sathanantham, Preethi East Tennessee State University East Tennessee State University 1415 Colony Park Dr. Apt. 9 807 University Parkway Johnson City, TN 37604 USA Johnson City, TN 37614 USA 423-557-9137 423-439-6572 [email protected] [email protected] Oseguera, Miguel Schmelz, Eric University of Illinois at Urbana-Champaign University of California, San Diego 228 E R Madigan Laboratory 9500 Gilman Drive, #MC0116 1201 W. Gregory Dr. Muir Biology Bldg, Rm 3254 Urbana, IL 61801 USA La Jolla, CA 92093 USA 217-244-3196 858-534-3946 [email protected] [email protected] Owens, Daniel Schramm, Katharina USDA-ARS University of Utah P.O. Box 1848 257 S 1400 E University, MS 38677 USA Salt Lake City, UT 84112 USA 662-701-9044 801-585-1324 [email protected] [email protected] Paulsmeyer, Michael Senaratne, Samiddhi University of Illinois at Urbana-Champaign The University of Melbourne 2101 W. Gregory Ave. School of BioSciences Room 311 Victoria 3010 Australia Urbana, IL 61801 USA +61424832662 217-671-3720 [email protected] [email protected] Shah, Jyoti Rahman, Md Mahbubur University of North Texas East Tennessee State University 1155 Union Circle, #305220 423-444-2428 Life Sciences Building B [email protected] Denton, TX 76203 USA 940-565-3535 Riggins, Chance [email protected] University of Illinois at Urbana-Champaign 260 ERML Smith, Shannon 1201 West Gregory Drive Middle Tennessee State University Urbana, IL 61801 USA 865-389-1575 217-300-0563 [email protected] [email protected] Soto-Hernandez, Marcos Rowe, Tim Colegio de Postgraduados PerkinElmer Carretera Mexico-Texcoco Km 36.5 2000 York Rd. Texcoco, Estado de Mexico 56230 Mexico OakBrook, IL 60523 USA 00525558045900 x 1361 312-661-2182 [email protected] [email protected] Sagwan, Laxmi Southern Illinois University 618-559-7817 [email protected]

August 8–12, 2015 • University of Illinois at Urbana-Champaign 79 Stevens, Fred Tokuhisa, Jim Oregon State University Virginia Tech 435 Linus Pauling Science Center Derring Biol. Sci. /3030 Corvallis, OR 97331 USA 1405 Perry St. 541-737-9534 Blacksburg, VA 24061 USA [email protected] 540-394-1545 [email protected] Sullivan, Michael US Dairy Forage Reserch Center, ARS-USDA Tollett, Christopher 1925 Linden Drive Arkansas Biosciences Institute/Arkansas State Madison, WI 53706 USA University 608-890-0046 4000 Legends Cove [email protected] Jonesboro, AR 72401 USA 870-557-6641 Sumner, Lloyd [email protected] Noble Foundation 2510 Sam Noble Pkwy Torres, Lucina Ardmore, OK 73401 USA University of Illinois at Urbana-Champaign 580-224-6710 1201 W. Gregory Drive [email protected] 228 ERML Urbana, IL 61801 USA Thakuri, Bal Krishna 217-244-3198 East Tennessee State University [email protected] 400 JL Seehorn Jr Rd Post Box 10510 Tripod, Nathan Johnson City, TN 37614 USA Arkansas Bioscience Institute 423-741-2367 504 University Loop [email protected] Jonesboro, AR 72401 USA 870-450-0846 Thapa, Chintamani [email protected] University of Saskatchewan 165-110 Science Place Umezawa, Toshiaki Saskatoon, SK S7N 5C9 Canada Research Institute for Sustainable Humanosphere, 306-241-1147 Kyoto University [email protected] Gokasho Ujji, 611-0011 Japan Tholl, Dorothea +81-774-38-3625 Virginia Tech [email protected] 409 Latham Hall, 220 Agquad Lane Blacksburg, VA 24061 USA van Breemen, Richard 540-231-4567 University of Illinois at Chicago [email protected] 833 S. Wood Street, M/C 781 Chicago, IL 60612 USA Tian, Li 312-996-9353 University of California, Davis [email protected] Mail Stop 3 Davis, CA 95616 USA Vaughan, Martha 530-752-0940 USDA ARS [email protected] 1815 N. University St. Peoria, IL 61604 USA Tiwari, Vijay 309-681-6295 East Tennessee State University [email protected] 423-741-9008 [email protected]

80 The Phytochemical Society of North America Vaughn, Steve Yamaguchi, Lydia Fumiko USDA/ARS/NCAUR USP 1815 N. University St. Av Prof Lineu Prestes, 748 Sala 1115 Peoria, IL 61604 USA Sâo Paulo, 05508-000 Brazil 309-681-6344 1130913813 [email protected] [email protected] Walker, Larry Yang, Changqing The University of Mississippi School of Pharmacy Shanghai Insititute of Plant Physiology and Ecology, National Center for Natural Products SIBS, CAS P.O. Box 1848 300 Fenglin Road 1014 Thad Cochran Research Center Shanghai, Shanghai 200032 China University, MS 38677-1848 USA 86 21 54924034 662-915-8859 [email protected] [email protected] Yuzuak, Seyit West, Leslie North Carolina State University University of Illinois at Urbana-Champaign 205-886-2781 Edward R. Madigan Laboratory [email protected] 1201 W. Gregory Ave., Urbana, IL 61801 USA Zerbe, Philipp 847-702-7941 University of California, Davis [email protected] 1002 Life Sciences West, Megan One Shields Avenue Kraft Foods Davis, CA 95616 USA 801 Waukegan Rd. 530-754-9652 Glenview, IL 60025 USA [email protected] [email protected] Wilhite, Thomas Shimadzu Scientific Instruments 2055 Army Trail Rd Ste 106 Addison, IL 60101 USA 800-792-1992 [email protected] Wise, Mitchell USDA, ARS, MWA, Cereal Crops Research Unit 502 Walnut Street Madison, WI 53726 USA 608-262-9242 [email protected] Xie, De-Yu North Carolina State University Raleigh, NC 27695 USA 919-515-2129 [email protected]

August 8–12, 2015 • University of Illinois at Urbana-Champaign 81 Notes

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84 The Phytochemical Society of North America Illini Union

ADA Access Art Gallery Colonial Room Event Services Hotel President’s Unisex Front Desk Lounge Elevators Bathroom

North Lounge Pine Lounge Room 104

Courtyard Cafe Espresso Royale

Stage ADA ADA Access Access M Illini Room W A

Illini Room U of I B Credit Union Jamba Juice Illini Room W C M Quad Shop Elevators Quad Cones Elevator A La Mode ATMs

SW SE South Lounge Foyer Foyer Tech Zone South Patio

ADA Access N W Women’s Restroom M Men’s Restroom Stairs MAIN LEVEL Illini Union Meeting Sponsors The Phytochemical Society of North America (PSNA) University of Illinois at Urbana-Champaign College of Agricultural, Consumer and Environmental Sciences (ACES) Department of Crop Sciences Department of Food Science and Human Nutrition (FSHN) Alkemist Labs (distributor for Extrasynthese) Bruker Kraft Foods Mahmoud A. ElSohly, Ph.D. PerkinElmer Inc. Arkansas Center for Plant Powered Production (P3) Shimadzu Scientific Instruments Thermo Fisher Scientific Inc. The University of Mississippi National Center for Natural Products Research

PROGRAM ADDENDUM (as of 8/5/15)

The following posters have been added to the program (odd numbers present on Sunday, even numbers present on Monday):

[P58] Genistein reduced colon cancer metastasis to liver through regulating the expression of metastasis-related genes in a mouse xenograft model Qian Li1, Stephane Lezmi2, Danni Chen1, Hong Chen1. 1Food Science and Human Nutrition, 2Pathobiology, University of Illinois at Urbana-Champaign, Urbana, IL

This study aimed at investigating the effect of genistein (Gen) on liver metastases derived from colon cancer and its molecular mechanisms. Male athymic mice were randomized to three dietary groups, which were western diet (W; calories%, fat 39%, carb 50%), W diet supplemented with 100 ppm Gen (GL) or 500 ppm Gen (GH) . Animals were fed with assigned diets for 16 weeks before they were intra-splenically injected with metastatic human colon cancer cells SW620 and sacrificed 8 weeks later. GL and GH groups have much lower liver metastasis rate (MR), which were 36% and 17%, respectively, compared to W group with MR of 55%. Mean metastases size of W group (4.7 mm2) is larger than GL (2.1 mm2) and GH (2.0 mm2) groups. To understand the molecular mechanism of the anti-metastatic function of Gen, the expression of multiple metastasis-related genes were determined by real-time PCR. Importantly, mRNA of NDRG1 gene was greatly decreased by 50% and 60% in GL (p=0.005) and GH group (p=0.001) compared to W group. As NDRG1 has been reported as a biomarker for metastasis and poor prognosis in hepatocellular carcinoma, the down- regulation of NDRG1 by Gen supplementation was corresponding to decreased liver metastasis. In summary, our study indicated that dietary supplementation of Gen suppressed colon cancer metastasis, which is associated with altered expression of metastasis-related genes by genistein.

[P59] Role of Acyl-lipid Desaturase 1 in Priming for Cold Acclimation in Arabidopsis Mingjie Chen1,2 and Jay J. Thelen2 1Haixia Institute of Science and Technology, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, P. R. China. 2Division of Biochemistry, Christopher S. Bond Life Sciences Center, University of Missouri, Columbia, Missouri 65211, USA

Membrane fluidity change has long been suggested as the primary mechanism plants adapt to cold stress. To further study this process, acyl- lipid/CoA desaturase 1 (ADS1; EC 1.14.99) was characterized using genetic, cellular, and biochemical approaches. Plants containing T-DNA insertion alleles of ads1 all appear similar to wild type (WT) under either standard growth conditions or low temperature, but displayed enhanced freezing tolerance after cold acclimation. Fatty acid composition analysis demonstrated that ads1 mutant plants grown at 23°C have 20% lower 18:1 content compared to WT. Lipidomics analysis revealed that 34C- species of monogalactosyl diacylglycerol (MGDG) content in ads1 mutants were 4.0% lower compared to WT. Lipid positional analysis identified that the sn-2 position of MGDG had higher 18:0 and 10% lower 18:1 fatty acid content compared to WT. Metabolomics profiling identified that the concentrations of some carbohydrates, amino acids and secondary metabolites were higher in ads1 mutants than in WT post cold acclimation. The cytosolic calcium content in ads1 mutant plants also was 1-fold higher than that of WT in response to cold shock. Subcellular localization of C- and N-terminal enhanced-fluorescence-fusion proteins indicated that ADS1 localizes exclusively to chloroplasts. These observations suggested that ADS1 encodes a plastid 18:1Δ9 desaturase specific to the sn-2 position of MGDG and is involved in cold perception and acclimation response.

[P60] Phytochemical characterization and antioxidant activity of different whole freeze dried berries and cherries with potential effect for type II diabetes. Boris Nemzer1,2,, Jing Yuan Wang1, Elvira de Mejia2 1FutureCeuticals, Inc., Momence, IL 60954, USA 2Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA

Type II diabetes is becoming more prevalent in the United States with about 8.3% of the population having this metabolic disorder. Berries are considered super fruits because of their natural antioxidants and have been investigated for their potential health benefit to manage diabetes. Objective: This study aimed to provide phytochemical composition, antioxidant capacity (AC) and evaluation of freeze dried whole elderberry, bilberry, blueberry (wild and cultivated), blackberry, black currant, tart cherry, sweet cherry, raspberry, cranberry, and strawberry for future biomedical functions in chronic diseases such as type II diabetes. Methods: Total anthocyanins (TA) content was determined by UV/Vis spectrophotometry by pH differential method and results were expressed as cyaniding 3-glucoside equivalent. The anthocyanins composition was provided by HPLC. The total polyphenols (TP) content was determined by spectrophotometry using Folin-C reagent and results were expressed as milligram of gallic acid equivalent per gram of dry sample. AC was measured by ORAC (oxygen radical absorbance capacity) assay. Results were expressed as micromoles of Trolox equivalent per gram of sample. α –Glucosidase, α- Amylase and DPP-IV methods for enzymatic inhibition activity were performed. Absorbance was read at 405 nm using Synergy2 Multi-well plate reader. Results were expressed as percent inhibition relative to diabetes drug acarbose as the positive control. Results: Thirteen anthocyanins were identified by HPLC and the highest total anthocyanins content was found in bilberry (2.89%) followed by elderberry (1.63%). TP ranged from 50.2 ± 0.14 mg/g in elderberry to 12.2 ± 0.05 mg/g in sweet cherry. Blackberry and bilberry are other good sources of high TP content (44.1± 0.08 and 41.3± 0.05 mg/g respectively). Elderberry had the much highest level of AC (1399 umole TE/g) compared to all other berries and cherries due to the highest level of polyphenols. The ORAC values were positively correlated to the total anthocyanins and polyphenol content. α-Glucosidase varied from 20.84± 3.46% inhibition/mg for whole elderberry to 104.12 ± 0.49 % inhibition for whole raspberry; α-Amylase varied from 35.64± 7.72 % for whole elderberry to 99.05 ± 1.53% inhibition for whole raspberry; DPP-IV varied from 43.74± 8.86% inhibition/mg for whole strawberry to 102.08 ± 1.02 % inhibition for whole elderberry. Conclusion: The results indicated that elderberry had the highest AC and TP content and higher DPP-IV inhibition activity. The formulation of berries blend with elderberry, raspberry and others have potential benefit for oxidative stress and diabetes prevention. PROGRAM ADDENDUM (as of 8/7/15)

 Cancellation: [S6-4] Tuesday at 3:50 pm / Presenter: Foluso Oluwagbemiga Osunsanmi  Change: [S7-3] will be presented Tuesday at 3:50 pm instead of Wednesday at 9:45 am / Presenter: Okiemute Rosa Johnson-Ajinwo  Cancellation: [S7-4] Wednesday at 10:15 am / Presenter: Syeda Maryam Hussain  Change: [S7-5] & [S7-6] will shift up to 9:45 am & 10:15 am on Wednesday / Presenters: Pavel Somavat & Richard Licayan

The following posters have been added to the program (odd numbers present on Sunday, even numbers present on Monday):

[P58] Genistein reduced colon cancer metastasis to liver through regulating the expression of metastasis-related genes in a mouse xenograft model Qian Li1, Stephane Lezmi2, Danni Chen1, Hong Chen1 1Food Science and Human Nutrition, 2Pathobiology, University of Illinois at Urbana-Champaign, Urbana, IL

This study aimed at investigating the effect of genistein (Gen) on liver metastases derived from colon cancer and its molecular mechanisms. Male athymic mice were randomized to three dietary groups, which were western diet (W; calories%, fat 39%, carb 50%), W diet supplemented with 100 ppm Gen (GL) or 500 ppm Gen (GH) . Animals were fed with assigned diets for 16 weeks before they were intra-splenically injected with metastatic human colon cancer cells SW620 and sacrificed 8 weeks later. GL and GH groups have much lower liver metastasis rate (MR), which were 36% and 17%, respectively, compared to W group with MR of 55%. Mean metastases size of W group (4.7 mm2) is larger than GL (2.1 mm2) and GH (2.0 mm2) groups. To understand the molecular mechanism of the anti-metastatic function of Gen, the expression of multiple metastasis-related genes were determined by real-time PCR. Importantly, mRNA of NDRG1 gene was greatly decreased by 50% and 60% in GL (p=0.005) and GH group (p=0.001) compared to W group. As NDRG1 has been reported as a biomarker for metastasis and poor prognosis in hepatocellular carcinoma, the down- regulation of NDRG1 by Gen supplementation was corresponding to decreased liver metastasis. In summary, our study indicated that dietary supplementation of Gen suppressed colon cancer metastasis, which is associated with altered expression of metastasis-related genes by genistein.

[P59] Role of Acyl-lipid Desaturase 1 in Priming for Cold Acclimation in Arabidopsis Mingjie Chen1,2 and Jay J. Thelen2 1Haixia Institute of Science and Technology, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, P. R. China 2Division of Biochemistry, Christopher S. Bond Life Sciences Center, University of Missouri, Columbia, Missouri 65211, USA

Membrane fluidity change has long been suggested as the primary mechanism plants adapt to cold stress. To further study this process, acyl- lipid/CoA desaturase 1 (ADS1; EC 1.14.99) was characterized using genetic, cellular, and biochemical approaches. Plants containing T-DNA insertion alleles of ads1 all appear similar to wild type (WT) under either standard growth conditions or low temperature, but displayed enhanced freezing tolerance after cold acclimation. Fatty acid composition analysis demonstrated that ads1 mutant plants grown at 23°C have 20% lower 18:1 content compared to WT. Lipidomics analysis revealed that 34C- species of monogalactosyl diacylglycerol (MGDG) content in ads1 mutants were 4.0% lower compared to WT. Lipid positional analysis identified that the sn-2 position of MGDG had higher 18:0 and 10% lower 18:1 fatty acid content compared to WT. Metabolomics profiling identified that the concentrations of some carbohydrates, amino acids and secondary metabolites were higher in ads1 mutants than in WT post cold acclimation. The cytosolic calcium content in ads1 mutant plants also was 1-fold higher than that of WT in response to cold shock. Subcellular localization of C- and N-terminal enhanced-fluorescence-fusion proteins indicated that ADS1 localizes exclusively to chloroplasts. These observations suggested that ADS1 encodes a plastid 18:1Δ9 desaturase specific to the sn-2 position of MGDG and is involved in cold perception and acclimation response.

[P60] Phytochemical characterization and antioxidant activity of different whole freeze dried berries and cherries with potential effect for type II diabetes. Boris Nemzer1,2,, Jing Yuan Wang1, Elvira de Mejia2 1FutureCeuticals, Inc., Momence, IL 60954, USA 2Department of Food Science and Human Nutrition, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA

Type II diabetes is becoming more prevalent in the United States with about 8.3% of the population having this metabolic disorder. Berries are considered super fruits because of their natural antioxidants and have been investigated for their potential health benefit to manage diabetes. Objective: This study aimed to provide phytochemical composition, antioxidant capacity (AC) and evaluation of freeze dried whole elderberry, bilberry, blueberry (wild and cultivated), blackberry, black currant, tart cherry, sweet cherry, raspberry, cranberry, and strawberry for future biomedical functions in chronic diseases such as type II diabetes. Methods: Total anthocyanins (TA) content was determined by UV/Vis spectrophotometry by pH differential method and results were expressed as cyaniding 3-glucoside equivalent. The anthocyanins composition was provided by HPLC. The total polyphenols (TP) content was determined by spectrophotometry using Folin-C reagent and results were expressed as milligram of gallic acid equivalent per gram of dry sample. AC was measured by ORAC (oxygen radical absorbance capacity) assay. Results were expressed as micromoles of Trolox equivalent per gram of sample. α –Glucosidase, α- Amylase and DPP-IV methods for enzymatic inhibition activity were performed. Absorbance was read at 405 nm using Synergy2 Multi-well plate reader. Results were expressed as percent inhibition relative to diabetes drug acarbose as the positive control. Results: Thirteen anthocyanins were identified by HPLC and the highest total anthocyanins content was found in bilberry (2.89%) followed by elderberry (1.63%). TP ranged from 50.2 ± 0.14 mg/g in elderberry to 12.2 ± 0.05 mg/g in sweet cherry. Blackberry and bilberry are other good sources of high TP content (44.1± 0.08 and 41.3± 0.05 mg/g respectively). Elderberry had the much highest level of AC (1399 umole TE/g) compared to all other berries and cherries due to the highest level of polyphenols. The ORAC values were positively correlated to the total anthocyanins and polyphenol content. α-Glucosidase varied from 20.84± 3.46% inhibition/mg for whole elderberry to 104.12 ± 0.49 % inhibition for whole raspberry; α-Amylase varied from 35.64± 7.72 % for whole elderberry to 99.05 ± 1.53% inhibition for whole raspberry; DPP-IV varied from 43.74± 8.86% inhibition/mg for whole strawberry to 102.08 ± 1.02 % inhibition for whole elderberry. Conclusion: The results indicated that elderberry had the highest AC and TP content and higher DPP-IV inhibition activity. The formulation of berries blend with elderberry, raspberry and others have potential benefit for oxidative stress and diabetes prevention.

[P61] Facultative Mutualism Between the Navel Orangeworm (Amyelois transitella) and Aspergillus flavus Daniel S. Bush1, Joel P. Siegel2, and May R. Berenbaum1 1University of Illinois at Urbana-Champaign, 2USDA, Agricultural Research Service, Parlier, CA

The highly aflatoxigenic fungal species Aspergillus flavus is closely associated with the navel orangeworm, Amyelois transitella Walker (Lepidoptera: Pyralidae), which attacks damaged or overripe tree nuts and fruits. Recent work suggests that this association is a facultative mutualism; A. flavus opportunistically infects nuts damaged by navel orangeworm, for example, and is phoretic on mobile life stages of the insect, while navel orangeworm larvae and adults are attracted to fungal volatiles. We now provide evidence of a nutritional component of this apparent mutualism, in which A. flavus detoxifies phytochemicals in host plants and increases the quality of the caterpillar's diet, while A. transitella provides additional nitrogen-rich substrate (in the form of frass) for A. flavus. Growth rates for A. transitella and A. flavus were measured separately and together on a diet containing almond meal, in the presence and absence of the furanocoumarin xanthotoxin (which occurs in some fruit hosts of A. transitella). In the absence of xanthotoxin, each species grows faster in the presence of the other than they do in isolation. A. transitella reaches adulthood 33% more quickly in the presence of A. flavus, while the fungus covers its medium 43% faster in the presence of A. transitella. In addition, xanthotoxin in the diet reduces the growth rate of A. transitella, but only when the fungus is absent. Ascomycete fungi, including Aspergillus species, are known to metabolize xanthotoxin and other furanocoumarins, and we suggest that this activity is highly beneficial to insect mutualist partners. These data are indicative of an ectosymbiotic mutualism between the fungus and the lepidopteran, an ecological relationship that may be more widespread than hitherto suspected and that has important implications for management of A. transitella in almonds and other economically important host plants.