2005 Annual Report 1.What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? What does it matter? Important food crops do not provide enough bioavailable levels of several essential micronutrients (e.g., iron, zinc, selenium, vitamin A, etc.) to meet the needs of certain population groups dependent on them for their nutriture, which has resulted in micronutrient deficiencies occurring in over three billion people globally (mostly resource-poor women, infants and children). For example, worldwide there are now over 3 billion people deficient in iron and 200 million people are known to be vitamin A deficient. It is estimated that there may be as many zinc deficient people as there are iron deficient people because the primary sources of these nutrients are the same foods. However, currently there is no clinical test to determine the extent of zinc deficiency globally. Additionally, some important food crops (e.g., durum wheat grain) can contain potentially harmful levels of heavy metals (e.g., cadmium) that can depress the bioavailability of the essential micronutrients - iron and zinc, increasing the frequency of micronutrient malnutrition in at risk populations. Furthermore, unacceptable levels of heavy metals in food crops that exceed international limits set by the Codex Alimentarius Commission can restrict their sale in international markets. Thus, ways need to be found to increase the bioavailable levels of micronutrients, and lower the levels of harmful heavy in staple food crops to improve the health of people and maintain international markets for the sale of farmers' products. This project has three specific goals:. 1)increase the density of bioavailable Fe and Zn in edible portions of staple food crops through identifying the criteria needed for selecting for or engineering high yielding genotypes having these characteristics,. 2)identify factors (antinutrients, promoter substances, nutrient interactions) that affect trace mineral bioavailability in foods and ascertain the key mechanisms of action, and. 3)determine linkages between trace elements in edible portions of food crops and their forms in soil, with emphasis on the distribution of essential and potentially hazardous trace elements in soils and their accumulation in cereal grain elucidating the physiological functions of genes that confers low grain-Cd-accumulation to durum wheat. The research to be undertaken falls under National Program Codes 107 - Human Nutrition Requirements and Intake, and 108 - Food Composition and Food Safety addressing goals 3.1.2 and 3.1.3 as described in the National Program Action Plan. Specifically, these include: 3.1.2.2. Determine the mechanisms of plant accumulation of toxic amounts of heavy metals. 3.1.2.3. Develop management strategies, and assist plant breeders to produce crops with reduced toxic metal accumulation. 3.1.3.6. Use cell culture and animal models to determine the biological activity, mechanism(s) of action, and health effects of food components modified by classical breeding and biotechnology. 3.1.3.7. Develop in vitro methodologies that function as screening tools to measure nutrient bioavailability including biological systems (e.g., in vitro digestion/cell culture) and non-biological systems (e.g., mathematical modeling and algorithms). This research project empowers scientists with knowledge needed to understand the ways in which food crops control the accumulation of bioavailable micronutrients, health promoting factors and toxic metals. The knowledge obtained can be used to genetically modify food crops or change cultural practices in ways that enhance human nutrition and health protecting consumers, shielding farmers from trade restrictions being imposed by international regulatory bodies on plant food commodities and providing new crops with nutritional quality traits that can improve public health and increase small farm profitability in niche markets. 2.List the milestones (indicators of progress) from your Project Plan. Year 1 (FY 2004) Test and rank select Fe-dense genotypes of rice, wheat, maize, and beans for bioavailable Fe using an in vitro Caco-2 model. Identify factors affecting Fe bioavailability using an in vitro Caco-2 model. Develop an in vitro Caco-2 model for determining Zn bioavailability from plant foods. Establish and map the distribution of trace elements in soils of major durum production in North Dakota. Determine effects of field landscape location on trace elements in soils and durum grain in North Dakota. Determine the role of Cd-phytochelatin complexes on Cd accumulation in durum wheat grain. Determine the mechanisms for Cd compartmentalization in durum wheat roots. Year 2 (FY 2005) Test most promising genotypes of maize, wheat and beans using a pig model in collaboration with Drs. Xingen Lei and Dennis Miller (Cornell University) to verify Caco-2 cell model results. Continue to identify factors affecting Fe bioavailability using an in vitro Caco-2 model. Begin application of Zn bioavailability model to test Zn-dense genotypes of beans, rice, wheat and maize, or explore contingencies. Prepare technical bulletin of maps and data for trace elements in soils of northern North Dakota. Prepare manuscript comparing uptake and root-shoot Cd-phytochelatin partitioning in durum wheat. Begin investigations on PC enzymes and on studies of Cd-PC transport across the tonoplast of cortical root cells. Year 3 (FY 2006) Test most promising Fe and Zn dense genotypes of maize and wheat using a pig model validating data from the in vitro Caco-2 model. Use results on metal speciation studies to initiate identifying promising lines of staple food crops having high levels of bioavailable Fe. Develop protocal for in vito human trial testing the bioavailable Fe in the most promising bean genotype in collaboration with the Grand Forks Human Nutrition Research Center in Grand Forks, ND. Continue practical applications, or exploration of contingencies for the in vitro Caco-2 cell model for Zn bioavailability. Publish technical bulletin on trace elements in soils of ND, and evaluate the distribution of Soil-Zn in the region and prepare manuscript of findings. Investigate mechanisms controlling Cd re-translocation from durum wheat roots to shoots. Year 4 (FY 2007) Begin in vitro Caco-2 cell model testing of Fe bioavailability in sweet potato and cassava and begin human trial of iron and zinc bioavailability in elite genotypes of rice and beans. Summarize results of factors affecting iron bioavailability form staple plant foods and share data with the CGIAR cooperating Centers (IRRI, CIAT, CIMMYT, and IFPRI). Continue Human trial on bioavailable Fe and Zn in elite genotypes of beans. Continue practical applications of Zn bioavailability Caco-2 cell model and evaluate results. Initiate molecular phase of research on Cd accumulation in durum wheat grain and attempt to clone and confirm function of low-Cd gene in durum wheat. Year 5 (FY 2008) Begin human trials of bioavailable Fe in elite lines of rice in collaboration with the Grand Forks Human Nutrition Research Center, Grand Forks, ND. Publish the results concerning the identity of factors affecting Fe bioavailability from staple plant foods. Publish results of the bioavailability of Fe in rice, wheat, beans, and wheat. Report results of bioavailable Zn in rice, wheat, beans and maize to cooperating CGIAR Centers (IRRI, CIMMYT, CIAT and IFPRI) and publish papers concerning results. Use cloned low-Cd gene in wheat transformation system to develop a low-Cd durum wheat transformant for using in plant breeding programs. 4a.What was the single most significant accomplishment this past year? Dietary inulin improves iron bioavailability to weanling pigs. Supplemental inulin (4% if diet) was shown to enhance iron bioavailability to pigs fed a maize-soybean based diet. Low bioavailable dietary iron is thought to be responsible for about 50% of iron deficiency globally. Two experiments were performed using a pig model that showed that there was a significant increase in hemoglobin repletion efficiency in those pigs fed a 4% inulin diet compared to those fed a diet without added inulin. This research was carried out in cooperation with Cornell University. The Caco-2 cell model is being modified to test the effects of prebiotics (i.e., inulin) on Fe bioavailability to compare to the results obtained using the pig model. CRIS 1907-42520-003-07G, Bioavailability of Fe and Zn in Biofortified Crops; Milestone #1. 4b.List other significant accomplishments, if any. None. 4c.List any significant activities that support special target populations. None. 4d.Progress report. For in-house (“D”) projects: We have completed a preliminary study comparing gin vitro Caco-2 cell Fe bioavailability with a piglet model using white beans and red beans. Generally, the Fe bioavailability determined using the two models agree: however, some refinement in the piglet studies remain in regards to housing of the animals group size, and diet formulation. An experiment comparing low and high inulin wheat varieties on iron bioavailability is planned for August, 2005 using the piglet model in cooperation with Drs. Dennis Miller & Xingen Lei (Cornell University), Drs. Robin Graham and Julia Humphries (University of Adelaide). Previous HarvestPlus studies carried out by Drs. Miller and Lei with diets supplemented with 4% inulin, using a pig model, have shown significant affects of dietary inulin on improving iron bioavailability form a maize-soy-based diet. We are currently screening a core collection of U.S. wheats for inulin in collaboration with Dr. Mark Sorrells (Cornell University). Ten-fold differences in inulin levels in Australian wheat lines have been reported by our collaborators in Adelaide. No rice varieties were screened as our cooperators did not produce any that were of sufficient Fe concentration to warrant screening. Approximately 142 genotypes of maize were screened with the goal of using genetic mapping techniques to identify genetic markers that are related to Fe bioavailability. The results identified several markers that warrant further study. Ten lines of sweet potato and 3 lines of cassava were also screened for Fe bioavailability. The results indicated that compounds within orange flesh sweet potatoes may have a strong promotional effect on Fe bioavailability. Similar effects appear to be present in cassava, suggesting that breeding these two crops for higher Fe concentration could be an effective biofortification strategy. Alternatively, providing a fortified sweet potato product as a “first-food” or complementary food to infants could be a very effective vehicle for dietary Fe. We continue research on producing more of the meat factor isolates from fish muscle to allow further testing and characterization of the active compounds. This project has been significantly delayed in the past year due to personnel turnover at both our facility and at our collaborator in ERRC. We now have a postdoctoral associate performing mass spectral analysis of fractions and generating fractions for isolation and purification by Dr. Arland Hotchkiss at the ERRC. We have demonstrated that the inhibitory effects of phytate on Fe bioavailability can be partially reversed by the addition of meat or ascorbic acid. These promoters do not appear to reverse the inhibitory effect of polyphenolics on Fe bioavailability. A manuscript investigating the mechanisms of inhibition of Fe bioavailability by phytic acid and polyphenolics has been submitted to the J Agric Food Chem. Currently, we are isolating and identifying polyphenol inhibitors in colored seed coats of beans and trying to isolate and identify promoter substances in white bean seed coats and in orange flesh sweet potato and yellow cassava. Manuscripts summarizing the in vitro Fe bioavailability screening of beans, sweet potatoes, and cassava are in preparation. An in vitro Caco-2 cell model for Zn bioavailability has been pursued with reasonable success using metallothionein as a proxy for Zn bioavailability. Conditions similar to our model for Fe bioavailability can be used; however, one major obstacle has been identified as a limiting factor (i.e., the Caco-2 cell line does not show an increase in cell-Zn uptake unless the soluble Zn concentration in the digest is greater than 10 µM). This appears to be a physiological response of the Caco-2 cells. As this concentration of Zn is one that is likely to be fairly common in our in vitro digests of staple food crops, we researching ways to concentrate the digest-Zn to maintain the Zn concentration greater than 10 µM so that bioavailability comparisons are possible using staple food crops. Possibly, this can be done through centrifugation-evaporation techniques. This approach is now being tested. A publication on using metallothionein as a proxy for Zn bioavailability in Caco-2 cells is in preparation. Maps of the distribution of soil pH, cation exchange capacity, organic carbon content, concentrations of 11 chelate-extractable elements, and total concentrations of 27 elements have been prepared for an ARS technical bulletin on the composition of surface and subsurface soils in northern North Dakota. Statistical summaries and geochemical correlations have been tabulated, and frequency distribution histograms have been plotted for all characteristics. Results for the distribution of cadmium, zinc, and other elements in durum grain and soils from different landscape positions in 32 fields have been summarized and figures have been prepared showing concentrations as a function of elevation and downslope distance. Exploratory statistical analyses of relations among soil physical attributes, soil chemical attributes and grain composition have been completed, showing that while available forms of many elements increase in downslope positions, only a few elements in grain, including cadmium, increase concurrently. Manuscript is being prepared in cooperation with David Hopkins, NDSU. Manuscript comparing uptake and root-shoot Cd-phytochelatin partitioning has been prepared by Dr. Jon Hart and will be submitted to a journal. Past research indicates that phytochelatin accumulation in root tissues is not the major controlling factor affecting Cd accumulation in durum wheat grain. Experiments are planned to determine the nature of Cd unloading into vascular tissues in roots and re-translocation and unloading in shoots. Understanding these phenomena will allow the identification of the genes responsible for Cd accumulation in durum wheat grain. Knowing the genes controlling this process will allow for the development of low-Cd durum wheat grain for global markets. For Subordinate Projects: This report serves to document research conducted under a Trust Fund Type Cooperative Agreement between ARS and the CIAT (Centro Internacional de Agricultura Tropical) entitled “Biotechnology assisted development of iron & zinc dense beans to combat micronutrient deficiencies in E. Africa” CRIS 1907-42520-003-05T which terminated 02/14/2005. Additional details can be found in the report for the parent CRIS 1907-42520-003-00D (“Understanding Soil-Plant-Human/Animal Food Systems and Nutrient Bioavailability to Improve Human Health”). The research performed under this agreement provided in vitro Caco-2 cell model evidence that white beans are significantly superior sources of bioavailable iron compared to colored bean varieties (i.e., red, black or brown bean varieties). The amount of bioavailable iron in white beans is encouraging and suggests that increased consumption of iron dense varieties of white beans have the potential to alleviate iron deficiency anemia in target populations in developing countries. Much lower amounts of bioavailable iron were present in the colored bean varieties tested. However, some colored bean varieties showed significantly more bioavailable iron compared to other colored bean varieties which warrants further study. This report serves to document research conducted under a Trust Fund Type Cooperative Agreement between ARS and the IFPRI (International Food Policy Research Institute) entitled “Biofortified crops for improved human nutrition” CRIS 1907-42520-003-06T which terminated 06/30/2005 and was replaced by CRIS 1907-42520-003-08R. Additional details can be found in the report for the parent CRIS 1907-42520-003-00D (“Understanding Soil-Plant-Human/Animal Food Systems and Nutrient Bioavailability to Improve Human Health”). Eleven genotypes of rice, 19 genotypes of beans and 16 genotypes if wheat were screened for bioavailable Fe using an in vitro Caco-2 cell model. For rice, the screening studies found no promising genotypes having enhanced levels of bioavailable Fe. In wheat, several genotypes were identified as having enhanced Fe bioavailability. In bean, certain genotypes were found to have enhanced Fe bioavailability, which was associated with white seed-coat color. Very little bioavailable Fe was observed in all colored beans, and the addition of ascorbate did not enhance the bioavailable Fe from the colored bean samples. These results clearly indicate that polyphenolic compounds are the predominant inhibitors of Fe bioavailability in beans. Both kaempferol and quercitrin in colored bean seed coats were shown to be significant inhibitors of Fe bioavailability using the in vitro Caco-2 cell model. Relative to other staple food crops, the molar ratio of phytate to Fe in CIAT beans was low. Considerable variation in phytate levels was observed in beans, yet the Fe bioavailability was not correlated with phytate concentration. Our studies indicate that the effects of phytate are reversible by the addition of ascorbic acid and meat whereas the effects of polyphenolics are not reversible by ascorbate or meat additions. Specific amounts of ascorbate and fish needed to reverse the effects of phytate inhibition were studied; however, more detailed work on various food matrices and combinations of foods may be warranted depending on patterns of food consumption in a given targeted region. That orange-fleshed sweet potato greatly promoted added Fe bioavailability represents new and exciting findings. Ascorbate alone may not be responsible for the enhancement of added Fe bioavailability in the sweet potatoes. Possibly, increasing the sweet potato Fe density may make this crop a significant source of bioavailable Fe and may promote Fe bioavailability from other foods in the diet. This report serves to document research conducted under a Reimbursable Fund Type Cooperative Agreement between ARS, and the IFPRI and the CIAT entitled “Biofortified crops for improved human nutrition” CRIS 1907-42520-003-08R replacing CRIS 1907-42520-003-06T. Additional details can be found in the report for the parent CRIS 1907-42520-003-00D (“Understanding Soil-Plant-Human/Animal Food Systems and Nutrient Bioavailability to Improve Human Health”). Research is in progress to isolate and identify the inhibitor and promoter substances of iron bioavailability in white and colored beans. To date two polyphenolic compounds (quercitrin and kaempferol isolated and identified in colored bean hulls but not in white bean hulls) have been shown to greatly inhibit iron bioavailability using an in vitro Caco-2 cell model. Additionally, the promoter substances in orange fleshed sweet potato and yellow cassava are also being studies to identify these promoter substances and their mechanisms of action. Resent research suggests that ¿-carotene and ascorbate may be responsible for promoting iron bioavailability from orange fleshed sweet potato and yellow cassava, and their mechanism of action my be through their ability to reduce ferric iron to ferrous iron. Human trails are planned with Drs. Janet Hunt and Gerald Combs, Jr. at the Grand Forks Human Nutrition Research Center in North Dakota to determine the bioavailable amounts of iron in white and colored beans to validate the Caco-2 cell iron bioavailability model that can be used to screen iron-dense staple food crop genotypes (rice, wheat, maize, bean, cassava and sweet potato) for bioavailable iron and to test the interactions of diet ingredients on iron bioavailability. This report serves to document research conducted under a General Assistance Type Cooperative Agreement between ARS and the Department of Food Science at Cornell University entitled “Bioavailability of Fe and Zn in biofortified crops” CRIS 1907-42520-003-07G which terminated 08/28/2005. Additional details can be found in the report for the parent CRIS 1907-42520-003-00D (“Understanding Soil-Plant-Human/Animal Food Systems and Nutrient Bioavailability to Improve Human Health”). Experiments have been initiated to determine the bioavailability of iron in two wheat varieties that differ in their inulin levels (one high and one low inulin wheat variety) using a pig model in cooperation with Drs. Dennis Miller (Department of Food Science) and Xingen Lei (Dept. Animal Science) at Cornell University. Previous pig model studies, carried out in 2004 and early 2005, showed that pigs fed a maize-soy based diet supplemented with 4% inulin absorbed significantly more iron from the diet (as determined by iron incorporation into hemoglobin; hemoglobin repletion efficiency) compared to pigs fed the same diet not supplemented with inulin. Studies are also under way to compare iron bioavailability from colored and white beans fed to pigs with the same bean varieties screened for iron bioavailability using an in vitro Caco-2 cell model in order to further validate the Caco-2 cell model. This report serves to document research conducted under a Reimbursable Specific Cooperative Agreement between ARS and the Department of Crop and Soil Science, Cornell University. Additional details can be found in the report for the parent CRIS 1907-42520-003-00D (“Understanding Soil-Plant-Human/Animal Food Systems and Nutrient Bioavailability to Improve Human Health”). There has been little activity or expenditure of funds under this cooperative agreement during the FY2005. During the summer months of 2005FY, a post doctoral research scientist has been finalizing maps of total and available elements in soils of northern North Dakota for a technical bulletin for farmers, agricultural advisors, and researchers. Completion and publication of this bulletin is expected in the coming fiscal year. In addition, revisions requested by GEODERMA have been completed on a manuscript comparing four geostatistical methods for handling skewed geographic information for soil zinc in northern North Dakota. This study has demonstrated benefits also from concurrent use of correlated characteristics of soil pH and organic carbon content in predicting soil zinc by cokriging. Collaborating in this research were Drs. Jay Wu and Steve DeGloria (Cornell University), and David Hopkins (NDSU). Technical assistance was provided by Dr. Michael Rutzke (Research Support Specialist). This report serves to document research conducted under a Reimbursable Specific Cooperative Agreement between ARS and the Department of Food Science, Cornell University entitled “Improving the nutritional quality of staple foods, food ingredients and food products” CRIS 1907-4520-003-02S. Additional details can be found in the report for the parent CRIS 1907-42520-003-00D (“Understanding Soil-Plant-Human/Animal Food Systems and Nutrient Bioavailability to Improve Human Health”). This CRIS was terminated on 08/31/05. Experiments were conducted to determine the effects of bread baking on the chemical form of iron fortificants. Our hypothesis was that elemental iron powders get oxidized during bread baking to either ferrous or ferric iron. If this is true, then elemental iron powders should have comparable bioavailability to iron salts such as ferrous sulfate. Our hypothesis was only partially supported. We found that only about 1/3 of the elemental iron is oxidized during bread baking and that the bioavailability of elemental iron powders added to bread are only about 50% as bioavailable as iron from ferrous sulfate. Further, we extended our work on the regulation of iron absorption from NaFeEDTA that we reported on last year. The objectives of this study were to compare iron uptake from radiolabeled ferrous sulfate, ferrous ascorbate, ferrous bisglycinate, ferric chloride, and ferric EDTA by Caco-2 cells with our without other added divalent metal cations (divalent cobalt and manganese compete with iron for uptake via the DMT-1 pathway). We hypothesized that manganese and cobalt would inhibit uptake of iron from ferrous sulfate, ferrous ascobate, and ferric chloride which follow the DMT-1 pathway into the cell, but not uptake from NaFeEDTA or ferrous bisglycinate since these stable complexes would be taken up intact via a paracellular or other pathway. Our results showed that cobalt and manganese did inhibit uptake from all the salts and complexes we tested, suggesting that iron from the two chelates follow the same uptake pathway as iron from the salts. This information is significant as it demonstrates that iron absorption from NaFeEDTA is regulated similar to forms of Fe that are known to be safe for fortification. This research could influence how this iron compound is used in food fortification. This report serves to document research conducted under a Trust Fund Type Cooperative Agreement between ARS and Kraft Foods entitled “Nutritional quality of fortified foods and their contribution to diets of developed regions” CRIS 1907-42520-003-03T. Additional details can be found in the report for the parent CRIS 1907-42520-003-00D (“Understanding Soil-Plant-Human/Animal Food Systems and Nutrient Bioavailability to Improve Human Health”). Due to its chemical and organoleptic properties, encapsulated micronized ferric phosphate (MFP) may be a suitable form of Fe fortificant for process cheese. Using the in vitro digestion/Caco-2 cell culture model, iron availability from encapsulated MFP was evaluated in process cheese, and in the combination of the MFP plus a rice, fish and vegetable meal typical of Southeast Asia. The results indicate that Fe from the MFP cheese was moderately available and was enhanced by the presence of ascorbic acid in the meal, thus it represents a promising form of fortification for this food product. Various forms of encapsulated NaFeEDTA were also evaluated in a snack food such as chocolate chip cookies. In general, NaFeEDTA was found to be a good Fe fortificant in the cookies, however the chocolate chips were found to be strong inhibitors of Fe availability. Although the cookies are considered a snack food, the addition of NaFeEDTA to the food does appear to increase its nutritional value by providing more bioavailable Fe. 5.Describe the major accomplishments over the life of the project, including their predicted or actual impact. Micronutrient malnutrition afflicts over 3 billion people worldwide especially resource-poor women, infants and children in both developing and developed countries. Iron deficiency alone affects over 2 billion people worldwide. Staple food crops (whole grains and edible legume seeds) contain substances (e.g., phytate and polyphenolics) that inhibit the utilization of iron and other micronutrients in these crops to people that are dependent on these foods for sustenance and are thought to be primarily responsible for about 50% of iron deficiency in the world. Ways need to be found to counteract the inhibitory effects of these staple food crop substances on micronutrient bioavailability. We have been studying the potential beneficial effects of certain prebiotics on iron bioavailability from staple food crops. The non-digestible carbohydrate, inulin (a fructooligosaccharide) is such a prebiotic that stimulates the proliferation of certain beneficial bacteria (i.e., Bifidobacteria and Lactobacilli) in the hind gut (i.e., colon). We hypothesized that the fermentation of inulin in the hind gut would enhance iron bioavailability in the presence of inhibitory substances from staple plant foods by making the hind gut an absorptive surface for iron absorption, and by the action of hindgut microorganism on degrading the inhibitors. In two weanling pig experiments, maize-soybean diets (containing 0.8% phytate) supplemented with 4% inulin significantly improved iron bioavailability (as measured by hemoglobin repletion efficiency) compared to the same diets without inulin added. Current research is focused on the effects of inulin in wheat grain on iron bioavailability, and on the mechanisms of action of inulin on iron bioavailability. This research was performed in cooperation with Drs. Dennis Miller (Department of Food Science) and Xingen Lei (Department of Animal Science) at Cornell University. Milestone 2a. Continue to identify factors affecting Fe bioavailability. ARS National Program Action Plan 107 [Objective 3.1.3, Nutritious Plant and Animal Products (Components 6 & 7)]. NP 108; Action Plan Component. Year 2 (FY2005) milestone -. 6.What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end-user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? Results on the bioavailability of Fe in promising genotypes of bean, wheat, maize, sweet potato and cassava using an in vitro Caco-2 cell model were provided to cooperating CGIAR Centers (i.e., CIMMYT, CIAT & CIP). Results were provided as a ranked list of genotypes in order of bioavailable Fe normalized using a standard control genotype. These results will be used by the Centers to advance genotypes in their breeding programs to improve the bioavailable Fe density in the diets of resource-poor people dependent on these staples for their nutriture globally. 7.List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). Presented a keynote address entitled “Nutritious food systems for subsistence farmers” at the Micronutrients in South and South East Asia International Workshop on Agricultural Strategies to Reduce Micronutrient Problems in Mountains and Other Marginal Areas of South and South East Asia held in Kathmandu, Nepal Sept. 8-10, 2004. Presented an overview of bioavailability and breeding objectives for the HarvestPlus Wheat Biofortification Meeting held in Istanbul, Turkey, Sept. 14-16, 2004 and chaired the session on Screening Staple Food Crops for Bioavailable Micronutrients. Presented the closing address at the 2004 Southern Plant Nutrient Management Conference titled “Crop micronutrient nutrition: food and feed quality impacts with reflections on opportunities for the southern U.S.” held in Olive Branch, Mississippi. He also gave a banquet speech on “Linking Agriculture to Human Health” at that meeting. Scientist was scheduled to give an opening address entitled “Agriculture: the real nexus for enhancing bioavailable micronutrients in food crops” at the International Symposium on Trace Elements and Health – Trace Elements in Agroecosystems and Health held in Hangzhou, China, Oct. 10-13, 2004. Because of delays in getting a visa to travel to China, he could not attend but his presentation was given for him by Dr. Zed Rengel (University of W. Australia) for him. Helped organize and gave a presentation entitled “Agriculture, nutrition and health: challenges and opportunities” at the Harvesting Health: Farming for More Nutritious Foods Through Agricultural Technologies: HarvestPlus Biofortification Challenge Program session held at the 2004 ASA-CSSA-SSSA International Annual Meetings in Seattle, WA on Nov. 1, 2004. Co-organizer and co-chair of a symposium on Managing Trace Element Bioavailability held during the VIIth Conference of the International Society for Trace Element Research in Humans held in Bangkok, Thailand from Nov. 7-12, 2004. He also gave a presentation entitled “Breeding plants with bioavailable trace elements”. Invited presentation entitled “Breeding objectives research under HarvestPlus” at the HarvestPlus – China Organization and Research Symposium held in Beijing, China Nov. 15, 2004. Opening keynote address entitled “Nutrition security: fertilizing crops for nutritious food” at the opening session of the 30th International Fertilizer Industry Association (IFA) Enlarged Council Meeting held in Santiago, Chile, Dec. 1-3, 2004. Invited presentation entitled “Harvesting health: the HarvestPlus biofortification challenge program” at the Agricultural Science Gordon Research Conference held in Ventura, CA, Feb. 13-18, 2005. Plenary presentation entitled “Linkages between trace elements in food crops and human health” at the 8th International Conference on the Biogeochemistry of Trace Elements held in Adelaide, Australia, April 3-7, 2005. He also was co-organizer of a symposium on Micronutrient Deficiencies in Global Crop Production at that conference and gave a presentation entitled “Trace element bioavailability: an important determinant in human health” at symposium. Invited presentation entitled “Farming for health: fertilizer technologies to improve human health” to the executives and technical staff at the Rio Tinto Borax fertilizer company’s headquarters in Valencia, CA, May 17, 2005. Invited presentation titled “Biofortification: breeding staple food crops for better human health” at the Cornell Institute of Food Science Symposium on Functional Foods, Bioactive Compounds and Human Health held in Ithaca, NY, May 22-24, 2005. Invited presentation entitled “Bioavailable iron – the determinant in the iron nutritional value of food crops” at the 2005 In Vitro Biology Meeting held in Baltimore, MD, June 5-7, 2005. One focus of Dr. Welch’s research was reported in Feedinfo News Service Scientific Reviews May, 2005 in an article entitled “Biofortification – a sustainable agricultural approach to addressing micronutrient malnutrition”. Available from URL: http://www.feedinfo.com. Invited paper to the HarvestPlus Maize Crop Meeting in Sete Lagoas, Brazil from Aug. 10-12, 2005 entitled "Essential Concepts of Screening Staple Food Crops for Fe and Zn Bioavailability". Review Publications Welch, R.M., Graham, R.D. 2005. Agriculture: the real nexus for enhancing bioavailable micronutrients in food crops. Journal of Trace Elements in Medicine and Biology. 18:299-307. Yeung, C., Miller, D., Welch, R.M., Glahn, R.P. 2005. Prebiotics and iron bioavailability - is there a connection?¿. Journal of Food Science. 70:1288-1292. Chaney, R.L., Reeves, P.G., Ryan, J.A., Simmons, R.W., Welch, R.M., Angle, J.S. 2004. An improved understanding of soil cd risk to humans and low cost methods to remediate soil cd risks. Biometals. 17(5):549-553. Liu, C., Glahn, R.P., Liu, R.H. 2004. Assessment of carotenoid bioavailability of whole foods using a caco-2 cell culture model coupled with an in vitro digestion. Journal of Agricultural and Food Chemistry. 52:4330-4337. Yeung, C.K., Glahn, R.P., Miller, D. 2004. Inhibition of iron uptake from iron salts and chelates by divalent metal cations in intestinal epithelial cells. Journal of Agricultural and Food Chemistry. Available: http://pubs.acs.org/journals/jafcau/. Yeung, C.K., Zhu, L., Glahn, R.P., Miller, D. 2004. Iron absorption from nafeedta is down-regulated in iron-loaded rats. Journal of Nutrition. 134:2270-2274. Etcheverry, P., Wallingford, J.C., Miller, D.D., Glahn, R.P. 2004. Calcium, zinc and iron bioavailabilities from a commercial human milk fortifier: a comparison study. Journal of Dairy Science. 87:3629-3637. Etcheverry, P., Wallingford, J., Miller, D., Glahn, R.P. 2005. The effect of calcium salts, ascorbic acid and peptic ph on calcium, zinc and iron bioavailabilities from fortified human milk. International Journal for Vitamin and Nutrition Research. 75:171-178. Yeung, C., Miller, D.D., Zhiqiang, C., Glahn, R.P. 2005. Bioavailability of elemental iron powders in bread assessed with an in vitro digestion/caco-2 cell culture model. Journal of Food Science. 70:5199-5203. Wortley, G., Leusner, S., Good, C., Gugger, E., Glahn, R.P. 2004. Iron availability of a fortified processed wheat cereal: a comparison of 14 fe forms using an in vitro digestion/caco-2 model. British Journal of Nutrition. 93:65-71. Oikeh, S.O., Menkir, A., Maziya-Dixon, B., Welch, R.M., Glahn, R.P. 2004. Assessment of iron bioavailability from 20 elite late-maturing tropical maize varieties using an in vitro digestion/caco-2 cell model. Journal of the Science of Food and Agriculture. 84:1202-1206. Welch, R.M., House, W.A., Monasterio, I., Cheng, Z. 2005. Potential for improving bioavailable zinc in wheat grain (triticum sp.) through plant breeding. Journal of Agricultural and Food Chemistry. 53:2176-2180. Hart, J.J., Welch, R.M., Norvell, W.A., Clarke, J.M., Kochian, L.V. 2005. Zinc effects on cadmium accumulation and partitioning in near-isogenic lines of durum wheat that differ in grain cadmium concentration. New Phytologist. 167:391-401. Li, L., Lyi, S.M., Heller, L., Rutzke, M., Welch, R.M., Kochian, L.V. 2005. Molecular and biochemical characterization of the selenocysteine se-methyltransferase gene and se-methylselenocysteine synthesis in broccoli (brassica oleracea var. italica). Plant Physiology. 138:409-420. Yeung, C., Miller, D., Hurrell, R.F., Lynch, S., Bothwell, T., Cori, H., Hertrampf, E., Kratky, Z., Rodenstein, M., Glahn, R.P. 2004. Enhancing the absorption of fortification iron: a sustain task force report. International Journal for Vitamin and Nutrition Research. 74:387-401.