Diversified strategies for improving iron bioavailability of maize
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Iron deficiency is the most common nutritional deficiency worldwide, affecting over 2 billion people, especially women and children. Populations consuming maize as a staple food are particularly prone to iron deficiency because of the low iron bioavailability (FeBA). The overall objective of this three-part study is to enhance FeBA in foods made of maize through novel fortification techniques, processing modifications, and genetic modifications.;The first study focused on the fortification of plant heme iron to improve FeBA in tortillas. Plants, particularly soy roots can produce heme iron as part of nitrogen fixation. Our objective was to test the effectiveness of soy root nodule (SRN) and purified soy hemoglobin (LHb) for FeBA using the in vitro Caco-2 cell model compared to bovine hemoglobin (BHb), and ferrous sulfate (FeSO4). When iron sources were tested alone, the FeBA values of LHb and BHb were 19% (P > 0.05) and 113% (P < 0.001) higher than FeSO4, respectively. However, when iron sources were used for fortification of maize tortillas (50 ppm), LHb and BHb showed similar FeBA, being 27% (P < 0.05) and 33% (P < 0.05) higher than FeSO 4. Our data suggest that heme iron from plants may be a novel value-added food fortificant with enhanced FeBA.;The second study evaluated the effects of maize processing on FeBA, with the intent to modify processing to enhance FeBA. Representative maize processing techniques were used: heating (porridge), fermentation (ogi), nixtamalization (tortillas), and decortication (arepas). The phytate content of maize products was significantly reduced by decortication (25.6%, p = 0.003) and nixtamalization (15%, p = 0.03), and iron content was reduced by decortication (29.1%, p = 0.002). The relative bioavailability (RBA, compared to 100% FeBA of porridge with FeSO4) of ogi was significantly higher than that of all other products when fortified with FeSO4 (p < 0.001) or reduced iron (p < 0.001). Addition of lactic acid (6 mg/g of maize) significantly increased iron solubility and increased FeBA by about 2-fold (p < 0.01), especially in tortillas. Consumer panel results showed that lactic acid addition did not affect the organoleptic characteristics of tortillas and arepas (p = 0.166 and 0.831, respectively). These results suggest that fermentation, or the addition of lactic acid to unfermented products, may improve FeBA of maize products. Lactic acid addition may be more feasible than using highly bioavailable but expensive fortificants.;The last study involved genetic modification by overexpression of Zea mays hemoglobin (ZmHb) in maize endosperm tissue. A first vector was developed by cloning a high expression 27gamma zein promoter from maize, and recombining it with a ZmHb gene fused to Green Fluorescent Protein (GFP), and nos terminator. Constructs with further modifications based on this first vector model were also produced: one without GFP and the other using 27gamma zein terminator. Maize callus was bombarded with the first vector, regenerated to plants which produced first generation (F1) seed. High GFP expressing seed, as determined by excitation fluorescence spectrophotometry, was shown to have 4.2-4.9 ug ZmHb/g maize based on immunoassay with ZmHb antibody. Heme pyrolle incorporation into the protein was shown by iron catalysed chemilluminescence. Preliminary studies on two high GFP expressing lines show that FeBA of ZmHb enhanced maize, determined by the Caco-2 cell assay, was 68--125% higher than the untransformed inbred maize variety, HiII (p < 0.01). Based on estimates from these assays, delivery of 1.6-2.5 mug heme iron per 100g serving of maize is possible. Our preliminary results in developing transgenic and intragenic maize suggest that overexpression of ZmHb in endosperm is feasible and may significantly enhance FeBA of maize by adding highly bioavailable heme iron.