<p></p><p></p><p><br></p><p></p><p>Vitamin A deficiency is the leading cause of childhood blindness affecting over 190 million preschool children around the world where the highest rates are found in Sub-Saharan Africa (1). The coexistence of this deficiency with shortfalls in iron and zinc has resulted in a shift in intervention strategies from single targeted approach to broader diet diversification. As a result, food-based strategies leveraging local nutrient-dense plants as natural fortificants have gained significant interest for their potential to simultaneously address multiple micronutrient, and in some instances macronutrient, deficiencies. However, the efficacy of such approach depends upon several factors including knowledge on the nutritional composition of native plant materials as well as strategies for their incorporation into staple consumer products. Additionally, there is lack of information on impact of concurrent introduction of mineral and provitamin A rich plants on the stability and bioavailability of each individual nutrients including changes in these factors over extended periods of exposure. This is a key point considering that many of these materials are reported to have potential inhibitors of carotenoid absorption (minerals, fiber and phenolics).</p><p></p><p>To address these research gaps, this dissertation focuses on three areas including 1) micronutrient, phytochemical and polysaccharide characterization of three commercially available native micronutrient dense African plant materials [Adansonia digitata (baobab), Moringa Oleifera (moringa) and Hibiscus Sabdariffa (hibiscus)] that have been targeted for use as natural iron fortificants; 2) determination of the impact of these materials on the bioaccessibility and intestinal uptake of provitamin A carotenoids from model composite cereal products and 3) assess the effect of longer term exposure to baobab and moringa on provitamin A carotenoid absorption and cellular differentiation biomarkers of human intestinal Caco-2 cells to better understand the potential impacts of extended exposure periods on long term micronutrient uptake. </p><p><br></p><p>Characterization of the plant fortificants focused on understanding both nutritive components and potential limiters of carotenoid bioavailability. Baobab, moringa and hibiscus all were found to contain key phytochemical and polysaccharide components that could be leveraged as nutritional and function ingredients. The relatively higher levels of lutein (57  4.6 g/g), zeaxanthin (11  0.1g/g) and -carotene (20  2 g/g) in moringa leaf powder support the notion that this plant material can be used as a source of provitamin A and non-provitamin A carotenoids. Phenolic analysis revealed the presence of substantial amounts of flavan-3-ols (1234  16 mg/100g) in baobab, anthocyanins (2001  56 mg/100g) in hibiscus, and flavonols (5352  139 mg/100g) in moringa leaf powder. Polysaccharide analysis demonstrated that the primary monosaccharide in baobab was found to be xyloglucan (47 %) which is in agreement with the tentative identification Xyloglucans (hemicellulosic polysaccharide) based on linkage analysis. Hibiscus was found to contain similar amounts of xylose (20%) and galactose (27%) supporting the presence of similar proportions of xyloglucans and pectic polysaccharides (type I, type II AG, RG I). The main monosaccharide in moringa was found to be galactose (36%) followed by glucose (23%) and linkage analysis revealed the presence of high proportions of pectic polysaccharides (type I, type II AG, RG I). These results provide insight into presence of potential enhancer or inhibitors of target micronutrient (provitamin A carotenoids or iron/zinc) bioavailability when used as functional and nutritional food ingredients.</p><p><br></p><p>Subsequently, the impact of mineral-rich baobab formulated at levels relevant for iron fortification on the bioaccessibility of provitamin A carotenoids (proVAC) from composite millet porridges containing dried carrot and mango was assessed using in vitro digestion. Proportions of millet flour and plant materials were dry blended to deliver ~25% of the RDA for vitamin A(VA) and iron(Fe) as follows: decorticated extruded millet (Senegalese Souna var.) (40-60%), dried proVA rich carrot and mango blend (30%), and dried Fe and ascorbic acid rich Adansonia digitata (baobab) (0-25%). While there were no significant differences in proVAC bioaccessibility from porridge formulations with 5 and 15% baobab (18.8+/-2.0 and 18.8±2.0% respectively) as compared to control containing no mineral-rich plant (23.8 +/- 1.2%), 25% baobab resulted in a significant decrease (p<0.05) in bioaccessibility of proVAC (13.3+/-1.6%). However, baobab inclusion did not impact intestinal uptake efficiency of provitamin A carotenoids by Caco-2 human intestinal cells</p><p><br></p><p>(3.3-3.6% -carotene and 3.7-4.5% for -carotene) across all formulation. These results suggest that any potential negative effects of baobab inclusion may be limited to food matrix interactions and digestion. This was confirmed in separate experiments that with experiments on baobab and carotenoid blends showing that digested baobab did not affect carotenoid absorption by Caco-2 cells. Overall these data support the notion that that modest inhibition of carotenoid bioaccessibility by baobab may not significantly limit carotenoid delivery from composite porridges. Furthermore, bioaccessible provitamin A content of a serving (200 g) of composite porridges can provide 27 - 48% of the RDA of vitamin A for children 1-3 years of age. </p><p><br></p><p>Finally, we evaluated the impact of long-term exposure to baobab and moringa digesta on Caco-2 cell differentiation biomarkers and provitamin A uptake to gain insight into how inclusion of these materials in to a daily diet may alter absorption and transport of nutrients or otherwise have potential negative effects on the intestine. Based on NMR analysis of intracellular metabolites in differentiating Caco-2 monolayers, significant alterations in specific osmotic pressure regulators, particularly glycerophosphocholine, taurine and myo-inositol were observed with repeated exposure to all treatment groups including the control (digested 0.9% saline solution). Changes in these metabolites levels have been linked with specific cellular function including protection against hyperosmotic stress and regulation of paracellular permeability of Caco-2 cells. Evaluation of carotenoid uptake comparing acute and acute on repeated exposure to treatment groups demonstrated that there was an overall significant reduction in carotenoid uptake with repeated exposure across all treatment groups including the control. Despite the reduction in carotenoid uptake, mRNA and protein levels of carotenoid transporters (CD-36, SR-B1 and FABP1) were not significantly altered with exposure through differentiation (except for SR-B1 protein levels). Decrease in SR-B1 levels may be due to bile acid accumulation from the digesta matrix which is known to regulate its own biosynthesis by a mechanism that involves the down-regulation SR-B1 expression to protect cells from cytotoxicity. Our results provide some insight into the impact of simulated gastrointestinal fluids alone on provitamin A uptake in this model system which are usually not taken into consideration in most Caco-2 cell studies. However, overall, these findings indicate that the introduction of baobab and moringa at levels relevant for delivery of meaningful levels of iron (15-23% RDA) should not have negative impacts on human intestinal function or carotenoid uptake over chronic use. </p><p>Taken together, our findings indicate that the three native Africa plant materials selected for investigation in these studies can be important sources of key micronutrients (iron, zinc and provitamin A carotenoids) and have potential as natural fortificants with application in staple foods such as cereal porridges. Incorporation of these plant materials, do not appear to negatively affect carotenoid bioavailability although there is a potential for their interaction during micellarization of carotenoids during normal digestion. While in vivo studies evaluating the bioavailability of provitamin A carotenoids from such composite formulations are required, these data support the further exploration of such natural fortification strategies in addressing micronutrient deficiencies in local African communities. </p><div><br></div><p><br></p>

No description available.

<p>Successful adoption of biofortified orange maize in developing countries requires careful consideration of factors across the chain from farm to fork. This includes consideration of post-harvest storage conditions optimal for the retention of both proviatamin A carotenoids and cooking quality critical to consumers. In these considerations, identification of economical storage methods is critical considering the limitations within specific countries that biofortified maize is being disseminated. To address these points, this dissertation research focused on evaluation of the utility of the Purdue Improved Crop Storage (PICS) bags as a post-harvest storage solution for biofortified maize. The specific focus of this research was to monitor retention of provitamin A and other carotenoids in two biofortified maize genotypes (OPVI and OPVII) as well as storage effect on flour functionality. Finally, a preliminary assessment of the impacts of storage on carotenoid bioaccessibility was completed to begin to translate findings to practice.</p><p>Maize grain from 2016 harvest was stored at ambient conditions for eight months in either PICS bags with or without an O₂ scavenger, (PICS-oxy) and (PICS-noxy), respectively and compared to storage in common polypropylene woven bags (control). After 4 months of storage carotenoid content was significantly higher (p<0.05) in PICS-oxy compared to PICS-noxy and woven bags demonstrating the importance of entrapped oxygen on maize carotenoid degradation. Furthermore, differences in carotenoid stability between maize genotypes were observed with OPVI having higher retention than OPVII. After 8 months, carotenoid retention remained dependent on storage bag and genotype with retention being greater in PICS-oxy and PICS-noxy compared to woven bags. However, final levels after 8 months were more similar between storage methods. Overall, oxygen content and genotype were found to be determining factors in the effectiveness of PICS to mitigate carotenoid degradation during post-harvest storage of maize.</p><p>While reducing the rate of carotenoid degradation during postharvest storage of biofortified maize is important, success of biofortified maize is also dependent on consumer adoption of these grains and their performance in traditional food preparation. Assessment of the rheological and functional properties of these two biofortified maize genotypes as a function of post-harvest storage was completed to assess the impact of post-harvest storage in PICS bags on flour functionality and rheological properties for the two biofortified orange maize genotypes and a control white maize genotype. Flour pasting profiles were assessed initially and at 4 and 8 months. After 8 month storage in woven and PICS bag, OPVI and OPVII produced porridges with similar viscosities to their initial viscosities regardless of postharvest storage type. White maize viscosities progressively decreased with storage and were significantly lower (p<0.05) in woven compared to PICS storage. Sequestration of oxygen (PICS-oxy) had modest but significant effects (p<0.05) on key pasting parameters including peak and final viscosities. These results suggest that oxygen sequestration has a critical effect on final flour functionality. DTT treatment partially restored flour pasting profiles suggesting disulfide linkages may modify pasting profiles of flour. There was also an increase in free ferrulic and <i>p</i>-coumaric acids during storage which may have contributed to observed decreases in porridge viscosities. Evidence of this was found through Raman spectroscopy with spectral intensity at both 478cm^-1 and 2911cm^-1 decreasing with storage suggesting the potential for structural changes induced by storage on starch polymer. While storage in PICS bags does not seem to adversely affect flour functionality it may provide some additional economic benefit resulting from requiring proportionally less flour to achieve similar final viscosities as flour from woven bag stored grains. </p><p>Finally, the effect of postharvest storage on bioaccessibility of carotenoids was explored using experimental wet cooked porridges made from ‘fresh’ and stored grains using an established three stage in-<i>vitro</i> digestion model. Relative carotenoid bioaccessibility (% micellarization) was generally higher in less viscous porridge made from grains stored in woven bags compared to porridge from initial or PICS bags stored grains suggesting that higher viscosity might partly explain lower relative bioaccessibility in porridge from grains stored in PICS bags. Absolute carotenoid bioaccessibility from experimental porridge was dependent on carotenoid species and storage system. Extrapolation of relative bioaccessibility (%) to absolute bioaccessibility (µg/g flour) suggests that fresh grains and their corresponding porridges would provide more absolute bioaccessible carotenoids compared to stored grains despite some improvement in relative accessibility. As such, storage losses remain the main factor impacting total available carotenoids and should continue to be an area of focus for future mitigation. With the potential to minimize post-harvest losses, improve carotenoid retention and provide a product with improved cooking performance, PICS bags do appear to offer a viable storage alternative to improve both food and nutrition security in developing countries.</p><p></p>