Addition of Whole Wheat Flour During Injera Fermentation Degrades Phytic Acid and Triples Iron Absorption from Fortified Tef in Young Women.

BACKGROUND: Iron deficiency is a major public health concern in Ethiopia, where the traditional diet is based on tef injera. Iron absorption from injera is low due to its high phytic acid (PA) content. OBJECTIVES: We investigated ways to increase iron absorption from FeSO4-fortified tef injera in normal-weight healthy women (aged 21-29 y). METHODS: Study A (n = 22) investigated the influence on fractional iron absorption (FIA) from FeSO4-fortified injera of 1) replacing 10% tef flour with whole wheat flour (a source of wheat phytase), or 2) adding an isolated phytase from Aspergillus niger. Study B (n = 18) investigated the influence on FIA of replacing FeSO4 in tef injera with different amounts of NaFeEDTA. In both studies, the iron fortificants were labeled with stable isotopes and FIA was calculated from erythrocyte incorporation of stable iron isotopes 14 d after administration. RESULTS: In study A, the median (IQR) FIA from the 100% tef injera meal was 1.5% (0.7-2.8%). This increased significantly (P < 0.05) to 5.3% (2.4-7.1%) on addition of 10% whole wheat flour, and to 3.6% (1.6-6.2%) on addition of A. niger phytase. PA content of the 3 meals was 0.62, 0.20, and 0.02 g/meal, respectively. In study B, the median (IQR) FIA from the 100% tef injera meal was 3.3% (1.1-4.4%) and did not change significantly (P > 0.05) on replacing 50% or 75% of FeSO4 with NaFeEDTA. CONCLUSIONS: FIA from tef injera by young women was very low. NaFeEDTA was ineffective at increasing iron absorption, presumably due to the relatively low EDTA:Fe molar ratios. Phytate degradation, however, greatly increased during tef fermentation on addition of native or isolated phytases. Replacing 10% tef with whole wheat flour during injera fermentation tripled FIA in young women and should be considered as a potential strategy to improve iron status in Ethiopia.

The Potential of Fermentation and Contamination of Teff by Soil to Influence Iron Intake and Bioavailability from Injera Flatbread.

The high phytic acid (PA) concentration in the diet based on teff injera is a likely contributing cause of iron deficiency in Ethiopia. We monitored PA during teff injera fermentation in 30 households in Debre Zeyit, Ethiopia and evaluated its influence on iron bioavailability, considering contaminant soil iron in teff flour. After fermentation (48h), mean PA concentration in injera batter decreased from 0.87 to 0.58 g/100 g dm (P < 0.001). Low phytase activity in teff flour (0.44 µmol phosphate/min/g) and a rapid drop in pH, indicated that PA degradation was driven by microbial phytases. The iron concentration in injera batter among the households ranged widely from 14.5-160.4 mg/100 g dm (mean: 34.7 mg/100 g dm) principally due to contamination with soil. Estimated intrinsic iron concentration of teff based on the strong correlation between total iron and aluminium concentrations (P < 0.001; aluminium concentrations in injera batter: 28.7-184.9 mg/100 g dm) was 4.4 mg/100 g dm, indicating that 86-97 % is extrinsic iron from soil. The median daily iron intakes from 3-day weighed food records in 10 young children were 18.9 mg/day including soil iron vs. 4.9 mg/day without soil iron (P < 0.01). The PA:iron molar ratios indicated low iron bioavailability from teff injera, particularly when soil iron was excluded. Traditional fermentation thus has a modest influence on PA levels and more complete degradation is needed to improve iron bioavailability. There is an urgent need to better understand the bioavailability of contamination iron from soil before considering national fortification or biofortification strategies in Ethiopia.

Mode of oral iron administration and the amount of iron habitually consumed do not affect iron absorption, systemic iron utilisation or zinc absorption in iron-sufficient infants: a randomised trial.

Different metabolic pathways of supplemental and fortification Fe, or inhibition of Zn absorption by Fe, may explain adverse effects of supplemental Fe in Fe-sufficient infants. We determined whether the mode of oral Fe administration or the amount habitually consumed affects Fe absorption and systemic Fe utilisation in infants, and assessed the effects of these interventions on Zn absorption, Fe and Zn status, and growth. Fe-sufficient 6-month-old infants (n 72) were randomly assigned to receive 6·6 mg Fe/d from a high-Fe formula, 1·3 mg Fe/d from a low-Fe formula or 6·6 mg Fe/d from Fe drops and a formula with no added Fe for 45 d. Fractional Fe absorption, Fe utilisation and fractional Zn absorption were measured with oral (57Fe and 67Zn) and intravenous (58Fe and 70Zn) isotopes. Fe and Zn status, infection and growth were measured. At 45 d, Hb was 6·3 g/l higher in the high-Fe formula group compared with the Fe drops group, whereas serum ferritin was 34 and 35 % higher, respectively, and serum transferrin 0·1 g/l lower in the high-Fe formula and Fe drops groups compared with the low-Fe formula group (all P<0·05). No intervention effects were observed on Fe absorption, Fe utilisation, Zn absorption, other Fe status indices, plasma Zn or growth. We concluded that neither supplemental or fortification Fe nor the amount of Fe habitually consumed altered Fe absorption, Fe utilisation, Zn absorption, Zn status or growth in Fe-sufficient infants. Consumption of low-Fe formula as the only source of Fe was insufficient to maintain Fe stores.

Consuming Iron Biofortified Beans Increases Iron Status in Rwandan Women after 128 Days in a Randomized Controlled Feeding Trial.

BACKGROUND: Food-based strategies to reduce nutritional iron deficiency have not been universally successful. Biofortification has the potential to become a sustainable, inexpensive, and effective solution. OBJECTIVE: This randomized controlled trial was conducted to determine the efficacy of iron-biofortified beans (Fe-Beans) to improve iron status in Rwandan women. METHODS: A total of 195 women (aged 18-27 y) with serum ferritin <20 µg/L were randomly assigned to receive either Fe-Beans, with 86 mg Fe/kg, or standard unfortified beans (Control-Beans), with 50 mg Fe/kg, 2 times/d for 128 d in Huye, Rwanda. Iron status was assessed by hemoglobin, serum ferritin, soluble transferrin receptor (sTfR), and body iron (BI); inflammation was assessed by serum C-reactive protein (CRP) and serum α1-acid glycoprotein (AGP). Anthropometric measurements were performed at baseline and at end line. Random weekly serial sampling was used to collect blood during the middle 8 wk of the feeding trial. Mixed-effects regression analysis with repeated measurements was used to evaluate the effect of Fe-Beans compared with Control-Beans on iron biomarkers throughout the course of the study. RESULTS: At baseline, 86% of subjects were iron-deficient (serum ferritin <15 µg/L) and 37% were anemic (hemoglobin <120 g/L). Both groups consumed an average of 336 g wet beans/d. The Fe-Beans group consumed 14.5 ± 1.6 mg Fe/d from biofortified beans, whereas the Control-Beans group consumed 8.6 ± 0.8 mg Fe/d from standard beans (P < 0.05). Repeated-measures analyses showed significant time-by-treatment interactions for hemoglobin, log serum ferritin, and BI (P < 0.05). The Fe-Beans group had significantly greater increases in hemoglobin (3.8 g/L), log serum ferritin (0.1 log µg/L), and BI (0.5 mg/kg) than did controls after 128 d. For every 1 g Fe consumed from beans over the 128 study days, there was a significant 4.2-g/L increase in hemoglobin (P < 0.05). CONCLUSION: The consumption of iron-biofortified beans significantly improved iron status in Rwandan women. This trial was registered at clinicaltrials.gov as NCT01594359.

Phytic acid degrading lactic acid bacteria in tef-injera fermentation.

Ethiopian injera, a soft pancake, baked from fermented batter, is preferentially prepared from tef (Eragrostis tef) flour. The phytic acid (PA) content of tef is high and is only partly degraded during the fermentation step. PA chelates with iron and zinc in the human digestive tract and strongly inhibits their absorption. With the aim to formulate a starter culture that would substantially degrade PA during injera preparation, we assessed the potential of microorganisms isolated from Ethiopian household-tef fermentations to degrade PA. Lactic acid bacteria (LAB) were found to be among the dominating microorganisms. Seventy-six isolates from thirteen different tef fermentations were analyzed for phytase activity and thirteen different isolates of seven different species were detected to be positive in a phytase screening assay. In 20-mL model tef fermentations, out of these thirteen isolates, the use of Lactobacillus (L.) buchneri strain MF58 and Pediococcus pentosaceus strain MF35 resulted in lowest PA contents in the fermented tef of 41% and 42%, respectively of its initial content. In comparison 59% of PA remained when spontaneously fermented. Full scale tef fermentation (0.6L) and injera production using L. buchneri MF58 as culture additive decreased PA in cooked injera from 1.05 to 0.34±0.02 g/100 g, representing a degradation of 68% compared to 42% in injera from non-inoculated traditional fermentation. The visual appearance of the pancakes was similar. The final molar ratios of PA to iron of 4 and to zinc of 12 achieved with L. buchneri MF58 were decreased by ca. 50% compared to the traditional fermentation. In conclusion, selected LAB strains in tef fermentations can degrade PA, with L. buchneri MF58 displaying the highest PA degrading potential. The 68% PA degradation achieved by the application of L. buchneri MF58 would be expected to improve human zinc absorption from tef-injera, but further PA degradation is probably necessary if iron absorption has to be increased.

A higher proportion of iron-rich leafy vegetables in a typical Burkinabe maize meal does not increase the amount of iron absorbed in young women.

Food-to-food fortification can be a promising approach to improve the low dietary iron intake and bioavailability from monotonous diets based on a small number of staple plant foods. In Burkina Faso, the common diet consists of a thick, cereal-based paste consumed with sauces composed of mainly green leaves, such as amaranth and jute leaves. Increasing the quantity of leaves in the sauces substantially increases their iron concentration. To evaluate whether increasing the quantity of leaves in sauces would provide additional bioavailable iron, an iron absorption study in 18 young women was conducted in Zurich, Switzerland. Burkinabe composite test meals consisting of the maize paste tô accompanied by an iron-improved amaranth sauce, an iron-improved jute sauce, or a traditional amaranth sauce were provided as multiple meals twice a day for 2 consecutive days. Iron absorption was measured as erythrocyte incorporation of stable iron isotopes. Mean fractional iron absorption from maize paste consumed with an iron-improved amaranth sauce (4.9%) did not differ from the same meal consumed with an iron-improved jute sauce (4.9%; P = 0.9), resulting in a similar quantity of total iron absorbed (679 vs. 578 µg; P = 0.3). Mean fractional iron absorption from maize paste accompanied by a traditional amaranth sauce (7.4%) was significantly higher than that from the other 2 meal types (P < 0.05), but the quantity of total iron absorbed was similar (591 µg; P = 0.4 and 0.7, respectively). A food-to-food fortification approach based on an increase in leafy vegetables does not provide additional bioavailable iron, presumably due to the high phenolic compound concentration of the leaves tested. Alternative measures, such as adding iron absorption enhancers to the sauces, need to be investigated to improve iron nutrition from Burkinabe maize meals.

Sodium iron EDTA and ascorbic acid, but not polyphenol oxidase treatment, counteract the strong inhibitory effect of polyphenols from brown sorghum on the absorption of fortification iron in young women.

In addition to phytate, polyphenols (PP) might contribute to low Fe bioavailability from sorghum-based foods. To investigate the inhibitory effects of sorghum PP on Fe absorption and the potential enhancing effects of ascorbic acid (AA), NaFeEDTA and the PP oxidase enzyme laccase, we carried out three Fe absorption studies in fifty young women consuming dephytinised Fe-fortified test meals based on white and brown sorghum varieties with different PP concentrations. Fe absorption was measured as the incorporation of stable Fe isotopes into erythrocytes. In study 1, Fe absorption from meals with 17 mg PP (8·5%) was higher than that from meals with 73 mg PP (3·2%) and 167 mg PP (2·7%; P< 0·001). Fe absorption from meals containing 73 and 167 mg PP did not differ (P= 0·9). In study 2, Fe absorption from NaFeEDTA-fortified meals (167 mg PP) was higher than that from the same meals fortified with FeSO4 (4·6 v. 2·7%; P< 0·001), but still it was lower than that from FeSO4-fortified meals with 17 mg PP (10·7%; P< 0·001). In study 3, laccase treatment decreased the levels of PP from 167 to 42 mg, but it did not improve absorption compared with that from meals with 167 mg PP (4·8 v. 4·6%; P= 0·4), whereas adding AA increased absorption to 13·6% (P< 0·001). These findings suggest that PP from brown sorghum contribute to low Fe bioavailability from sorghum foods and that AA and, to a lesser extent, NaFeEDTA, but not laccase, have the potential to overcome the inhibitory effect of PP and improve Fe absorption from sorghum foods.

Total iron absorption by young women from iron-biofortified pearl millet composite meals is double that from regular millet meals but less than that from post-harvest iron-fortified millet meals.

Iron biofortification of pearl millet (Pennisetum glaucum) is a promising approach to combat iron deficiency (ID) in the millet-consuming communities of developing countries. To evaluate the potential of iron-biofortified millet to provide additional bioavailable iron compared with regular millet and post-harvest iron-fortified millet, an iron absorption study was conducted in 20 Beninese women with marginal iron status. Composite test meals consisting of millet paste based on regular-iron, iron-biofortified, or post-harvest iron-fortified pearl millet flour accompanied by a leafy vegetable sauce or an okra sauce were fed as multiple meals for 5 d. Iron absorption was measured as erythrocyte incorporation of stable iron isotopes. Fractional iron absorption from test meals based on regular-iron millet (7.5%) did not differ from iron-biofortified millet meals (7.5%; P = 1.0), resulting in a higher quantity of total iron absorbed from the meals based on iron-biofortified millet (1125 vs. 527 µg; P < 0.0001). Fractional iron absorption from post-harvest iron-fortified millet meals (10.4%) was higher than from regular-iron and iron-biofortified millet meals (P < 0.05 and P < 0.01, respectively), resulting in a higher quantity of total iron absorbed from the post-harvest iron-fortified millet meals (1500 µg; P < 0.0001 and P < 0.05, respectively). Results indicate that consumption of iron-biofortified millet would double the amount of iron absorbed and, although fractional absorption of iron from biofortification is less than that from fortification, iron-biofortified millet should be highly effective in combatting ID in millet-consuming populations.

Iron bioavailability from a lipid-based complementary food fortificant mixed with millet porridge can be optimized by adding phytase and ascorbic acid but not by using a mixture of ferrous sulfate and sodium iron EDTA.

Home fortification with lipid-based nutrient supplements (LNSs) is a promising approach to improve bioavailable iron and energy intake of young children in developing countries. To optimize iron bioavailability from an LNS named complementary food fortificant (CFF), 3 stable isotope studies were conducted in 52 young Beninese children. Test meals consisted of millet porridge mixed with CFF and ascorbic acid (AA). Study 1 compared iron absorption from FeSO4-fortifed meals with meals fortified with a mixture of FeSO4 and NaFeEDTA. Study 2 compared iron absorption from FeSO4-fortifed meals without or with extra AA. Study 3 compared iron absorption from FeSO4-fortified meals with meals containing phytase added prior to consumption, once without or once with extra AA. Iron absorption was measured as erythrocyte incorporation of stable isotopes. In study 1, iron absorption from FeSO4 (8.4%) was higher than that from the mixture of NaFeEDTA and FeSO4 (5.9%; P < 0.05). In study 2, the extra AA increased absorption (11.6%) compared with the standard AA concentration (7.3%; P < 0.001). In study 3, absorption from meals containing phytase without or with extra AA (15.8 and 19.9%, respectively) increased compared with meals without phytase (8.0%; P < 0.001). The addition of extra AA to meals containing phytase increased absorption compared with the test meals containing phytase without extra AA (P < 0.05). These findings suggest that phytase and AA, and especially a combination of the two, but not a mixture of FeSO4 and NaFeEDTA would be useful strategies to increase iron bioavailability from a CFF mixed with cereal porridge.

Afebrile Plasmodium falciparum parasitemia decreases absorption of fortification iron but does not affect systemic iron utilization: a double stable-isotope study in young Beninese women.

BACKGROUND: Iron deficiency anemia (IDA) affects many young women in sub-Saharan Africa. Its etiology is multifactorial, but the major cause is low dietary iron bioavailability exacerbated by parasitic infections such as malaria. OBJECTIVE: We investigated whether asymptomatic Plasmodium falciparum parasitemia in Beninese women would impair absorption of dietary iron or utilization of circulating iron. DESIGN: Iron absorption and utilization from an iron-fortified sorghum-based meal were estimated by using oral and intravenous isotope labels in 23 afebrile women with a positive malaria smear (asexual P. falciparum parasitemia; > 500 parasites/µL blood). The women were studied while infected, treated, and then restudied 10 d after treatment. Iron status, hepcidin, and inflammation indexes were measured before and after treatment. RESULTS: Treatment reduced low-grade inflammation, as reflected by decreases in serum ferritin, C-reactive protein, interleukin-6, interleukin-8, and interleukin-10 (P < 0.05); this was accompanied by a reduction in median serum hepcidin of ≈ 50%, from 2.7 to 1.4 nmol/L (P < 0.005). Treatment decreased serum erythropoietin and growth differentiation factor 15 (P < 0.05). Clearance of parasitemia increased geometric mean dietary iron absorption (from 10.2% to 17.6%; P = 0.008) but did not affect systemic iron utilization (85.0% compared with 83.1%; NS). CONCLUSIONS: Dietary iron absorption is reduced by ≈ 40% in asymptomatic P. falciparum parasitemia, likely because of low-grade inflammation and its modulation of circulating hepcidin. Because asymptomatic parasitemia has a protracted course and is very common in malarial areas, this effect may contribute to IDA and blunt the efficacy of iron supplementation and fortification programs. This trial was registered at clinicaltrials.gov as NCT01108939.