2006 Annual Report 4d.Progress report. This report serves to document research conducted under a Specific Cooperative Agreement between ARS and Food Science at Cornell University. Additional details of research can be found in the report for the parent project 1907-42520-003-00D, "Understanding Soil-Plant-Human/Animal Food Systems and Nutrient Bioavailability to Improve Human Health." Our primary goal for this past year was to extend our work on the iron absorption enhancing properties of prebiotics with a focus on mechanisms underlying this enhancing effect. Anemic piglets at age 5 weeks were allocated to a standard corn-soy diet (control) or the same diet supplemented with 4% inulin. After 7 weeks on this diet, the animals were killed and sections of the small intestine and colon were quickly removed. Enterocytes were harvested by scraping with a glass slide and frozen in liquid nitrogen. Using primers designed to represent DMT-1, DcytB, ferroportin, and ZnT-1, RT-PCR analyses were performed to quantify the expression of the respective genes encoding for these proteins. Concentrations of m-RNA encoding for DMT-1, DcytB, ferroportin, and ZnT-1 in duodenal samples were significantly higher in the inulin group (p < 0.05). In colon samples, DMT-1 and ferroportin were significantly upregulated. These results suggest that up-regulation of genes encoding for iron and zinc membrane transporters is a possible mechanism whereby inulin enhances iron and possibly zinc absorption. A manuscript describing this work is nearly ready for submission to a journal. Portions of this research were funded by Harvest Plus. We also continued our work on the regulation of iron absorption from NaFeEDTA that we reported on last year. Specifically, we studied the uptake of iron from NaFeEDTA, FeSO4, and FeCl3 by Caco-2 cells in the presence or absence of ascorbic acid (AA) and bathophenanthroline disulfonic acid (BPDS). Uptake of iron from all 3 iron sources was similar in the absence of ascorbic acid. Adding ascorbic acid at a 5:1 molar excess compared to iron increased uptake by 5.4, 5.1, and 2.8-fold for FeSO4, FeCl3, and NaFeEDTA, respectively. The smaller effect of AA on uptake from NaFeEDTA may be related to the higher solubility of NaFeEDTA, and/or the strong binding affinity of EDTA for Fe3+. Adding BPDS to the media inhibited uptake from all three iron sources equally. BPDS binds Fe2+ but not Fe3+. These results suggest that uptake of iron from all 3 sources follows a common pathway, i.e. the iron is released from complexes or salts, reduced to Fe2+, and transported into the cell via DMT-1. Taken together, these results suggest that uptake of iron from NaFeEDTA by intestinal enterocytes is regulated similarly to uptake from iron salts. Portions of this work were supported by the USDA/NRI Competitive Grant Program (#58-1907-0-033). We also continued our studies designed to determine the effects of bread baking on the bioavailability of iron from flour fortified with elemental iron powders. Our hypothesis was that elemental iron powders get oxidized during bread baking to either ferrous or ferric iron, thereby enhancing the bioavailability of the elemental iron (it has been reported that the bioavailability of elemental iron powders to human subjects is approximately 50% that of FeSO4). We baked 3 large batches of bread, one using flour fortified with hydrogen reduced (HR) iron powder, one with unfortified flour, and a 3rd with flour fortified with FeSO4. The bread was dried and mixed into corn-based swine diets. For the diet containing the unfortified bread, hydrogen reduced iron was added to the diet during mixing, i.e. after baking. The diets were fed to anemic pigs in a hemoglobin repletion trail. Hemoglobin repletion efficiencies, an index of iron bioavailability, were 7.5%, 8.7%, and 18.7% for the pigs on the HR iron diet (iron added after baking), HR iron diet (iron added prior to baking), and FeSO4 diets, respectively. The difference between the 2 HR diets was not significant but iron bioavailability from the FeSO4 diet was significantly higher than from either of the HR diets. Therefore, our hypothesis was not supported. We conclude that bread baking does not enhance the bioavailability of elemental iron powders.