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Objectives: Brain-derived neurotrophic factor (BDNF) plays an essential role in brain and metabolic health. The fact that higher concentrations are associated with improved cognitive performance has resulted in numerous intervention trials that aim at elevating BDNF levels. This systematic review provides an overview of the relation between various nutritional factors and BDNF concentrations in controlled human intervention studies. Methods: A systematic search in May 2020 identified 48 articles that examined the effects of dietary patterns or foods (n = 3), diets based on energy intake (n = 7), vitamins and minerals (n = 7), polyphenols (n = 11), long-chain omega-3 polyunsaturated fatty acids (n = 5), probiotics (n = 8), and miscellaneous food supplements (n = 7). Results: In particular, studies with dietary patterns or foods showed increased peripheral BDNF concentrations. There are also strong indications that polyphenols tend to have a positive effect on BDNF concentrations. Four of the 11 included studies with a polyphenol intervention showed a significant increase in BDNF concentrations, one study showed an increase but this was not statistically analyzed, and two studies showed a trend to an increase. Discussion: The two polyphenol classes, phenolic acids, and other phenolic compounds were responsible for the significant effects. No clear effect was found for the other dietary factors, which might also be related to whether serum or plasma was used for BDNF analysis. More work is needed to understand the relation between peripheral and central BDNF concentrations.

Several studies have shown that diets containing lower vitamin D than in the AIN-93G diet do not compromise bone structure, bone mineral density (BMD), and/or bone strength in male and female mice. This study determined if a diet containing low vitamin D from prepregnancy through to the end of lactation maintained these bone outcomes to a similar extent as a high vitamin D diet. Mice were fed an AIN-93G diet with 25 (LD diet) or 5000 (HD diet) IU vitamin D/kg diet from premating through to lactation (n = 15/group). Of the major structure outcomes, only cortical area fraction of the distal femur was lower (P, The study findings suggest that the level of bone-supporting nutrients such as vitamin D should be part of the conversation regarding a potential revision of the AIN-93 diets.

OBJECTIVES: The purpose of this study was to investigate the relationship between dietary Mg intake and concentrations of ionized magnesium (iMg(+2)) in whole blood and total Mg in serum. METHODS: We evaluated blood iMg(+2) concentration, the physiological active measure of Mg, as well as serum Mg concentration which is a more commonly used measure of body's status. The data study population were healthy participants (18–52 years; n = 23) who participated in three clinical visits in a magnesium supplement bioavailability study. All study participants were not taking Mg supplements within two weeks of study screening. Dietary Mg intake obtained from 3-day food records was compared with iMg(+2) in whole blood and total Mg in serum concentrations using Pearson's correlations. Ionized magnesium was measured with NOVA 8 (NOVA Biochemical, Waltham, MA), serum Mg was measured with atomic absorption spectroscopy, and dietary intake of Mg was assessed using the Nutrition Data System for Research (NDSR; University of Minnesota, MN). Results were reported as means (standard deviations) and percentages for continuous and categorical data, respectively. RESULTS: Mean iMg(+2) and serum Mg concentrations, and dietary Mg intake were 1.27 (0.07) mg/dL, 2.20 (5.0) mg/dL and 304.05 (128.77) mg/day, respectively. Results from the Pearson's correlation coefficient showed no significant association between dietary Mg intake vs. and serum Mg (r = −0.30, p = 0.23), nor was there a significant association between dietary Mg intake and iMg(+2) (r = −0.08, p = 0.74). CONCLUSIONS: Our findings suggest that iMg(+2) and serum Mg concentrations may not reflect dietary Mg intake in a significant manner. Further research is needed to identify objective and accurate biomarkers that reflect dietary Mg intake, which will enable a better understanding of Mg deficiency and its association with conditions such as cardiovascular diseases, hypertension, diabetes, and chronic kidney disease. FUNDING SOURCES: An unrestricted educational grant from New Capstone Inc., the manufacturer of ReMag®.

OBJECTIVES: Investigate the impact of variable vitamin K (VK) intakes on the coagulation activities of four VK-dependent factors and clotting times, in warfarin-treated rats. METHODS: Male Wistar rats were randomly allocated to a AIN-93 based diet containing low (L: 80 mcg/kg/d), adequate (A: 750 mcg/kg/d) or enriched (E: 2000 mcg/kg/d) phylloquinone (K1) containing diet (n = 24/diet group). After one week, half the animals from each diet group were randomly allocated to receive 0.2 mg warfarin/kg/d through drinking water (W gp) or plain water (C gp), for 10 weeks. Coagulation activity (%) was assessed for factors II, VII, IX and X, and clotting times were based on prothrombin [PT (sec)] and activated thromboplastin times [APTT (sec)]. Measures were obtained at the end of the study and were conducted in the hospital clinical laboratory using standard procedures. Diet effects within C and W groups were investigated using one-way ANOVA and uncorrected Fisher post-hoc tests. RESULTS: Warfarin treatment resulted in significantly higher clotting times (PT and APTT) in all diet groups when compared to corresponding C groups (p

OBJECTIVES: Iron deficiency is one of the most common nutrient deficiencies worldwide. Adaptive thermogenesis plays a critical role for maintaining energy homeostasis. Currently, the role of dietary iron deficiency on adaptive thermogenesis is unknown. We hypothesized that dietary iron deficiency attenuates adaptive thermogenesis via dysfunctional iron metabolism in adipose tissue. METHODS: C57BL/6 male mice were fed a diet containing either adequate-iron (Cont: 35ppm) or deficient-iron (ID: 3ppm) throughout for 14 weeks and the last 10 weeks were fed a high-fat diet to induce obesity. Systemic iron status was evaluated by measuring serum ferritin, hemoglobin (Hb), and hematocrit (HCt). The iron content of adipose tissue was determined by ICP-MS spectrometer. The protein and gene expressions related to iron-handling and thermogenesis were measured in adipose tissue by Western blot analysis and qPCR. To assess adaptive thermogenic function, mice were exposed to cold temperature acutely (3 hour) or administrated with β3-adrenoceptor agonist (CL) for 5 days. The core body temperature and thermogenesis were determined by a rectal thermometer and infrared camera, respectively. RESULTS: The ID mice displayed a non-anemic iron deficiency with reduced serum ferritin (p

OBJECTIVES: To investigate the effects of vitamin K2 supplementation on the prevention and treatment of rheumatoid arthritis(RA). METHODS: Vitamin K2 and rheumatoid arthritis were searched in Pubmed from inception until December 2020. RESULTS: Study 1: Vitamin K2 inhibited the proliferation of mitogen-activated PBMCs of RA patients with an IC50 value of 3,288.47 ± 4,910.02 ng/mL. There was a significant correlation between IC50 values of vitamin K2 and patient-related factors of RA patients (p

OBJECTIVES: While the replacement of folic acid (FA) with the bioactive folate form 5-methyltetrahydrofolate (MTHF) is gaining popularity, a comparison of their effects during pregnancy is limited. In a comparison of maternal intakes of FA and MTHF at recommended or high (5X) doses in Wistar rat dams, we found that dams fed 5X MTHF gained > 70% more weight and ate 8% more food to 19-weeks post weaning (PW) than those fed the high FA diet (Nutrients. 2021;13:48). However, both high dose diets resulted in dysregulation of leptin and central energy regulatory systems. The objective of this study was to compare the effect of gestational FA and MTHF diets on metabolic response and leptin-dependent hypothalamic energy-regulatory genes in the brain of their female offspring. METHODS: Pregnant Wistar rats were fed an AIN-93G diet with 1X recommended (2mg/kg diet) or 5X FA or equimolar MTHF. At weaning, female offspring were fed a high-fat diet until 19-weeks PW. Body weight, food intake, adiposity, plasma leptin, and gene expression of leptin signaling and related candidate genes in the hypothalamus were measured. RESULTS: Similar to the mothers, the 5X-MTHF gestational diet resulted in offspring with higher weight (>15%, P 1.5-fold, P

OBJECTIVES: Thiamine acts as a coenzyme (prosthetic group) after converting to thiamine pyrophosphate in crucial metabolic enzymes of pyruvate dehydrogenase, α-ketoglutarate dehydrogenase and transketolase. Theoretically, thiamine should not be consumed by an increase in the number of substrates for those enzymes. However, thiamine is often described to be consumed by excessive carbohydrate intake. Such a possible misunderstanding is assessed in the relevant literature. METHODS: The original paper (Elmadfa, et al. Int J Vitam Nutr Res 2001), which seems to mislead the reader into considering an increase in thiamine requirement for excessive carbohydrate intake, and the relevant papers were reviewed. RESULTS: The original paper demonstrated that increases in carbohydrate intake from 55% to 65% and 75% of total energy caused decreases in plasma and urine concentrations of thiamine, while erythrocyte transketolase activity remained unchanged. This seems to misleadingly support the idea that excessive carbohydrate intake increases the thiamine requirement for carbohydrate metabolism. It could be interpreted as the decreases of thiamine concentrations were observed only during an adaptation period. It was reported that diabetic patients had low plasma thiamine levels in association with increased renal clearance and fractional excretion of thiamine, without a decrease in erythrocyte transketolase activity (Thornalley, et al. Diabetologia 2007). As a similar case of possible misinterpretation, a case report (Watson, et al. Ir J Med Sci 1981) demonstrated that glucose loading precipitated thiamine deficiency in malnourished patients with latent thiamine deficiency. CONCLUSIONS: From the relevant literature, it is inferred that excessive carbohydrate intake in people without hyperglycemia could lead to a small increase, if any, in urinary thiamine excretion, but not to an increase in thiamine requirement for glucose metabolism. Even in the academic field, it seems that thiamine is often misunderstood to be consumed by excessive carbohydrate intake. FUNDING SOURCES: None.

OBJECTIVES: High folic acid (FA) intake may be associated with adverse health outcomes, including colon cancer promotion. 5-methyltetrahydrofolate (5MTHF) has been proposed as a safer alternative form of folate supplementation. We compared the effects of FA and 5MTHF supplementation on the progression of aberrant crypt foci (ACF; earliest colon cancer precursor). METHODS: Male Sprague Dawley rats (n = 120) received a control diet (1mg FA or equimolar 5MTHF) at weaning and ACF were induced by azoxymethane. Six weeks post-induction, rats were randomized to the control or supplemental (10mg FA or equimolar 5MTHF) diets for 18 weeks. Plasma folate concentrations were assessed using microbiological assay and compared. Colorectal tumor incidence, multiplicity and burden (sum of tumor diameters) were determined and compared. RESULTS: 5MTHF resulted in higher plasma folate concentration compared to FA (p

OBJECTIVES: Cognitive decline is a major public health issue among older adults. Dietary supplement use is common among older adults, and a concern has been raised about high folic acid intake among those with vitamin B12 (vB12) deficiency and exacerbation of cognitive decline. We evaluated supplement intake and folate and vB12 concentrations among older adults with measured cognitive function. METHODS: We used NHANES 2011–2014 data on adults aged 60 + years (n = 3266) and estimated folic acid and vB12 intake from dietary supplements. We defined vB12 insufficiency as serum vB12 concentration ≤ 258 pmol/L and high folate concentrations as either serum folate ≥ 59 nmol/L or RBC folate ≥ 1609 nmol/L. Cognitive impairment was defined as a test score

OBJECTIVES: Calcium (Ca(2+)) is a vital micronutrient for many physiological functions with the greatest rate of accumulation occurring during the critical period of infancy. Previous work has demonstrated that molecular mechanisms of intestinal Ca(2+) absorption across the small intestine are significantly different in animal models of infants and adults to permit greater absorption early in life. The colon contributes to overall Ca(2+) balance in adults via transcellular, TRPV6 mediated and paracellular claudin-2 and -12 mediated pathways. Whether these same colonic pathways contribute to overall Ca(2+) absorption in infants is not known. Here we aimed to investigate the molecular details of Ca(2+) absorption across the large intestine in murine models of infancy relative to older mice. METHODS: Mice at 14 days (P14) were used as a model of suckling infants and mice at 2 months were employed to represent adult physiology. Wildtype and mutant mice with a non-functioning Trpv6 or deletions of Cldn2 or Cldn12 were used. Net (45)Ca(2+) flux (J(Ca)) and Ca(2+) permeability (P(Ca)) were measured in Ussing chambers. Gene expression was determined by real-time PCR. RESULTS: J(Ca) indicates net absorption across the colon at both P14 and 2 months. While gene expression of Trpv6 and S100g suggest greater cellular uptake of Ca(2+) into colonocytes at P14, net J(Ca) in vitro was not different than at 2 months. In contrast to previous work in mice at 2 months, TRPV6 does not mediate J(Ca) at P14. P(Ca) was 20% greater at P14 than 2 months, suggesting greater capacity for bidirectional diffusion of Ca(2+) down an electrochemical gradient in younger mice. In contrast to previous work in mice at 2 months, claudin-2 and claudin-12 do not mediate P(Ca) at P14 and, expression of Cldn2 and Cldn12 were significantly reduced in younger mice. CONCLUSIONS: These results improve our understanding of intestinal Ca(2+) handling during a critical age early in life. Future work is required to delineate molecular details under in vivo conditions of colonic Ca(2+) transport in infants. FUNDING SOURCES: This work was funded by grants from the Women and Children's Health Research Institute, which is supported by the Stollery Children's Hospital Foundation, and the National Sciences and Engineering Research Council to RTA, who is the Canada Research Chair in Renal Epithelial Transport Physiology.

OBJECTIVES: Zinc is an essential nutrient for diverse biological processes in the body. Cellular zinc homeostasis is established through differential expressions of the transmembrane zinc transporter proteins, ZnTs and ZIPs. The aims of the current studies were to elucidate the roles of cellular zinc in erythrocyte maturation, and to determine the zinc transporters essential to erythroid zinc homeostasis. METHODS: G1E-ER4 mouse cells were employed as an in vitro study model of terminal erythroid differentiation. A cell-impermeable zinc chelator, diethylenetriamine pentaacetate (DTPA), was used to limit extracellular zinc availability. For gene silencing, gene-specific siRNAs were introduced to cells via Nucleofection. Functional impacts of zinc and gene deficiency were assessed via ICP-MS-based metal quantitation, heme assays, and gene expression assays using RNA-seq, qPCR, and western analyses. RESULTS: G1E-ER4 cells featured a 1.7-fold increase in total cellular zinc contents after 48 h of differentiation. Restriction of zinc import by 50 µM of DTPA led to less red coloration and lower increases in mean corpuscular hemoglobin contents by development. The heme deficiency by DTPA was fully restored by the addition of equimolar zinc, and was not due to changes in cellular iron contents. Zinc-deficient G1E-ER4 cells differentiated with normal Alas2 and Hbb-b1 transcript responses, but less Alad and alpha-globin expressions. Among the 24 zinc transporter genes, Zip10 produced the most prominent response to zinc restriction in differentiating erythroid cells. ZIP10-deficient G1E-ER4 cells were less efficient than control cells in hemoglobin production under zinc restriction. ZIP10 deficiency alone had no effects on the molecular indices of red cell hemoglobinization. CONCLUSIONS: Our studies characterize zinc as a nutrient essential to normal erythroid maturation and heme synthesis. Moreover, we have identified a compensatory role of ZIP10 for erythroid zinc homeostasis during zinc restriction. Thus, poor zinc status and ZIP10 mutations might serve as potential risk factors and thus new therapeutic targets for anemia and other erythrocyte-related disorders. FUNDING SOURCES: Supported by CFANS Graduate Fellowship to JK, and the Allen Foundation, Inc. and USDA NIFA Hatch Funds to M-SR.

OBJECTIVES: The purpose of this secondary outcome analysis is to investigate whether small-quantity lipid-based nutrient supplements (SQ-LNS) alters lipid, protein or glycan composition, or the cholesterol efflux capacity (CEC), of high-density lipoprotein (HDL) particles in children in the International Lipid-Based Nutrient Supplements (iLiNS) DYAD trial in Ghana. METHODS: Plasma samples were obtained from a subcohort of 80 children at 18 months of age from the iLiNS-DYAD-Ghana trial. Mothers were randomized to either iron and folic acid (IFA) in pregnancy and 200 mg/d calcium for 6 months postpartum or SQ-LNS (pregnancy and 6 months postpartum). Children in the SQ-LNS group received SQ-LNS from 6 to 18 months while children in the IFA group did not receive supplements. HDL was isolated from plasma by sequential ultracentrifugation followed by size-exclusion chromatography. Assay of cholesterol efflux was performed in vitro, and glycoproteomic and lipidomic composition were analyzed by mass spectrometry. The primary analysis was a comparison of the effects of intervention groups on HDL lipidome, proteome, and CEC. In the exploratory analysis, we compared the enrichment of glycopeptides in measured HDL-associated proteins between groups. RESULTS: Mean (±SD) HDL CEC was higher among children in the SQ-LNS vs. IFA group (20.9 ± 4.1% vs. 19.4 ± 3.3%; one-tailed p = 0.038). We found no differences in HDL lipidomic or proteomic composition between groups. CONCLUSIONS: Prenatal and postnatal SQ-LNS may improve the CEC of child HDL particles. These improvements may have a potential impact on child health outcomes. FUNDING SOURCES: Supported by Bill & Melinda Gates Foundation grant to the University of California, Davis.

OBJECTIVES: The work was carried out to compare the quantity of vitamin C in two commonly consumed species of pineapple in Nigeria; Honey Queen Pineapple (indigenous) and Brazza imported from Republic of Benin. This is expedient because complaints of gastro-intestinal tract problems often emerge after consumption of Brazza specie. METHODS: Seven of both species were cleaned, peeled, flesh separated from hard core making seven flesh portions, seven hard core each. All flesh portions of each specie were blended separately into paste as well as the core. Four portions were the end result Brazza Pineapple Core (BRPC); Brazza Pineapple Flesh (BRPF); Honey Queen Pineapple Core (HQPC) and Honey Queen Pineapple Flesh (HQPF). Standard solutions of ascorbic acid was prepared by weighing 0.5g of standard ascorbic acid and dissolved in 10ml of 0.1 molar phosphoric acid solution to obtain a concentration. Hundred millimeters of each sample was poured in sterile tubes and centrifuged at 6000rpm for five minutes, poured into sterile sample bottles and filtered with muslin cloth to have clearer sample and stored in sterile sample bottles. 20ml of each prepared fruit juice samples solution was injected into the HPLC machine in triplicates. The flow rate was 0.8ml/min. The ascorbic acid was detected at 245nm using Agilent Variable Wavelength Detector. Linear curve was done by plotting concentration versus average detector response. RESULTS: The quantity of vitamin C in each sample was estimated from the standard curve calibration from the HPLC using the formular- Quantity = Curve Area-324.26 34.79. Vitamin C content in BRPC was 2.8ug/ml; BRPF 16ug/ml; HQPC 1.10ug/ml and HQPF 1.17ug/ml. Vitamin C concentration was derived by multiplying these results by 50. Amount of vitamin C per gram was 126.75 ug/g; 726.61 ug/g; 76.96 ug/g and 53.13 ug/g respectively. CONCLUSIONS: It has been proven that the Brazza specie from the neighbouring country contains more than 10 times higher than the Nigeria local specie. Nutritionist/dietitian advice should be sought before individuals who react negatively to high doses of vitamin C consume Brazza pineapple specie. FUNDING SOURCES: It is a self funded project.

OBJECTIVES: In vitro dissolution tests are valuable first-step tools in the development of bioavailable delivery systems, making it possible to assess the performance of novel technologies in releasing active ingredients through the amount dissolved in a dissolution medium. The objective of this study was to evaluate whether phosphatidyl choline-enriched lipid encapsulation releases the majority of vitamin C as ascorbic acid past the stomach in a standard in vitro dissolution procedure and to assess the release profile. METHODS: A novel phosphatidyl choline-enriched lipid encapsulation that is solid at room temperature was tested for dissolution in a standard dissolution apparatus according to compendial United States Pharmacopeia methods, as per Good Manufacturing Practices for dietary supplements. One serving (included 1000 mg ascorbic acid) was placed into vessels containing simulated gastric fluid (0.1 M HCl) for 120 minutes then changed to simulated intestinal fluid (buffered 2% sodium lauryl sulfate, pH 6.8) for an additional 360 minutes. Aliquots were tested for ascorbic acid concentration at 8 time points by titration. The data was compared with 1000 mg of regular ascorbic acid in a capsule format. RESULTS: The phosphatidyl choline-enriched lipid encapsulation released 36% of the vitamin C at 2 hours in the acid phase and released 56% at 3 hours, 67% at 4 hours, 85% at 6 hours and 98% at 8 hours in the intestinal phase. Regular vitamin C filled capsules released 100% of the vitamin C at 30 minutes. CONCLUSIONS: The dissolution results indicated that phosphatidyl choline-enriched lipid encapsulation can pass the stomach and release the majority of vitamin C in the small intestine. The encapsulation demonstrated a sustained and prolonged release of vitamin C over an 8 hour period. The release profile observed in this in vitro study suggests phosphatidyl choline-enriched lipid encapsulation may improve nutrient absorption and bioavailability which requires further testing including human clinical trials. FUNDING SOURCES: This study was supported by Lonza (Greenwood, SC) and Ritual (Natals Inc, Los Angeles, CA).

OBJECTIVES: To develop a compartmental model which describes vitamin B-6 metabolism in the blood and tissues of the mouse under a variety of circumstances and to develop a computer application which can utilize the data to display the movement of tracer through the various compartments. METHODS: A circulation model was developed. Literature data from vitamin B-6 feeding studies and tracer experiments were used to develop an appropriate compartmental model using the SAAM II program (The Epsilon Group, Charlottesville, VA). A web-based application transforms the mass and flux data into intuitive and interactive graphical illustrations. RESULTS: Describing the interconversions between pyridoxine, pyridoxine 5’-phosphate, pyridoxal, pyridoxal 5’-phosphate, pyridoxamine, pyridoxamine 5’-phosphate and 4-pyridoxic acid in multiple tissues required 231 compartments. The largest amount of data deals with liver and brain. The model includes less detailed information on plasma, erythrocytes, gut, bone, muscle, heart, kidney, skin, adipose tissue and lung. The model includes adjustments to food intake, water intake, cardiac output, binding sites and Vmax values for enzymes based on the specified body weight of experimental animals. We did not include growth curves at this time. The model uses two parallel systems to monitor the steady state of endogenous metabolites as well as following tracer administration. Binding mechanisms are included to provide conservation of vitamin B-6 when intake is reduced. The model provides reasonable agreement with literature data on various vitamin B-6 intakes as well as oral and intravenous administration of tracer. It also reveals some areas which need clarification. For example, we have not found any detailed analysis of vitamin B-6 metabolites in mouse urine. There is little pyridoxic acid in plasma or urine suggesting that pyridoxic acid may not be the primary end product for vitamin B-6 in the mouse. The visualization application shows changes in the content of all 231 compartments over time illustrating the value of such computer applications in the interpretation of large, complex models. CONCLUSIONS: This model facilitates the simulation of various dietary and physiological conditions on vitamin B-6 metabolism in mice. We hope to adapt it to rats, pigs and humans. FUNDING SOURCES: Purdue University Fort Wayne.

OBJECTIVES: Gestational diabetes (GDM) is associated with adverse outcomes in both mother and child. Women with diabetes and obesity (BMI ≥ 30kg/m(2)) are recommended to take more folic acid than recommended for healthy women, to prevent birth defects. However, little is known about the effects of high folic acid supplementation in GDM. The objective of this study was to determine if folic acid supplementation in a mouse model of GDM affects maternal and fetal metabolic health. METHODS: Female (C57BL/6J) mice were fed from weaning a control diet (10% kcal fat; control dams) or western diet (45% kcal fat; western dams; model of GDM) with (10mg/kg diet) or without (2mg/kg diet) supplemental folic acid (SFA). Dams were fed for 13 weeks prior to breeding. Physiological assessments of glucose homeostasis were conducted before breeding. Tissues were collected at gestational day 18.5 RESULTS: Western dams had greater adiposity (p

OBJECTIVES: The prenatal period is a critical time for fetal development, programming the offspring's later-life health in response to the postnatal environment. We have shown that a high maternal choline diet programs long-term energy regulation leading to higher food intake and weight-gain in mature rat offspring fed a normal fat diet. However, the offspring's response to an obesogenic post-weaning diet has not been described. We aim to elucidate the interaction between the choline content of the gestational diet (GD) and fat content of the post-weaning diet (PWD) on male Wistar rat offspring's long-term metabolic phenotype. METHODS: Pregnant Wistar rats were fed an AIN-93G diet with either recommended choline (RC, 1g/kg diet choline bitartrate) or high choline (HC, 2.5-fold). Male pups were weaned to either a normal (10%) fat (RC-NF and HC-NF) or a high (45%) fat (RC-HF and HC-HF) diet for 17 weeks. Dependent measures were body weight, food intake, visceral adiposity, plasma glucoregulatory hormones and triglycerides, and plasma and hepatic free fatty acids (FFAs). Data were analyzed with 2-way ANOVA for main effects of GD and PWD and their interaction. Measures with significant interaction effects were followed by a Student's T-test comparing groups stratified by PWD. RESULTS: HC-HF offspring had lower body weight (7%, P

OBJECTIVES: Chronic inflammation and selenium deficiency are common in persons infected with the human immunodeficiency virus (HIV). Both C-reactive protein, a pro-inflammatory biomarker, and selenium deficiency have been associated with accelerated disease progression and poorer survival among persons infected with HIV-infection. However, research on the role of serum selenium levels in inflammation is limited. We assessed the association between serum selenium levels and C-reactive protein (CRP) concentrations in a cohort of persons infected with HIV. METHODS: A cross-sectional survey was conducted among 233 HIV-infected persons (124 men and 109 women) aged 18–60 years who were residing in Kathmandu, Nepal. Regular serum CRP and selenium concentrations were measured by the latex agglutination turbidimetric and the atomic absorption method, respectively. Relationships were assessed using multiple linear regression analysis to adjust for potential confounding factors including sociodemographic, smoking, body mass index, chronic diseases, CD4(+) T-cell count, and anti-retroviral therapy. RESULTS: The geometric means of serum selenium and CRP concentrations were 9.65 µg/dL and 1.43 mg/L, respectively. Mean serum CRP concentration was significantly decreased with increasing serum selenium concentration across selenium tertiles (P for trend = 0.019), with mean serum CRP concentration in the highest tertile of serum selenium concentration was 40.8% lower than that in the lowest tertile. We found a significant inverse relation between log selenium and log CRP concentrations (beta for 1 unit change in log selenium; β = −1.01, p = 0.06). CONCLUSIONS: Serum selenium levels may be inversely associated with serum CRP concentrations in persons infected with HIV. Further prospective study to confirm the role of serum selenium levels in inflammation among persons infected with HIV is warranted. FUNDING SOURCES: University of Massachusetts Amherst.

OBJECTIVES: Bioequivalence of vitamin E and folate in a single oral dose multivitamin (MVI) gummy or tablet. METHODS: This crossover clinical trial involved healthy adults randomized to either gummy or tablet MVI containing vitamin E (VE – 279 IU gummy; 239 IU tablet) and folate (1860 µg gummy; 1877 µg tablet) as a single dose in Phase 1 with blood samples collected at pre-dose and 0.5-, 1-, 2-, 4-, 6-, 8-, 9-, 10-, 24-, and 48-hours after dosing, followed by a 2-week washout period. In Phase 2, participants crossed over to receive MVI in the form not previously given, with blood draws at the same timepoints. Maximum Concentration (Cmax), Time of Maximum Concentration (Tmax), and Area Under the Curve (AUC) were calculated for each subject for both vitamin forms. Bioequivalence for AUC, Tmax, and Cmax was defined as a 90% confidence interval (CI) for the gummy to tablet comparator ratio of the geometric mean between 80% and 125%. RESULTS: Nineteen participants completed the study. Both gummy and tablet demonstrated absorption for both vitamins. The ratio of geometric means demonstrated bioequivalence between gummy and tablet for both VE and folate with both AUC and Cmax. While bioequivalence was also demonstrated for VE with Tmax, folate absorption peaked earlier in the gummy group (Tmax 1.89 hrs) than the tablet group (Tmax 4.00 hrs), shifting Cmax values in the gummy group to earlier timepoints than in the tablet group without affecting total absorption AUC. These results were consistent with a previous pilot study. CONCLUSIONS: Overall, under the conditions of this study, both gummy and tablet showed similar absorption of vitamin E and folate. FUNDING SOURCES: Church & Dwight Co., Inc.

OBJECTIVES: The 2013 Palestinian Micronutrient Survey(1) reported high and comparable 1(st) trimester prevalences of micronutrient deficiencies in the Gaza Strip and West Bank: 23.6% and 21.3% for iron, 67.9% and 49.6% for zinc, 11.4% and 8.8% for vitamin A, 27.9% and 19.1% for B(12), 78.6% and 66.7% for vitamin D and 17.5% and 13.2% for vitamin E, respectively. Rates were generally higher among gravida in their 2(nd)-3(rd) trimesters. Interim, clinic-based, anemia rates(2) in the Gaza Strip and West Bank of 32% and 19% in the first trimester and 71% and 38% in the 2(nd) and 3(rd) trimesters, respectively, coupled with food insecurity, dietary inadequacy, civil conflict and stresses from the COVID-19 pandemic, suggest micronutrient deficiencies persist as a public health burden in the State of Palestine. To replace current iron-folic acid (IFA) with a multiple micronutrient supplement (MMS) providing a Recommended Dietary Allowance of 15 essential vitamins and minerals as standard of antenatal care (ANC) in UNWRA clinics and hot spots serving pregnant women in the Gaza (n = 22) in 2021–2 and West Bank (n = 44) in 2022–3. METHODS: MMS is planned to start in Gaza in the Fall of 2021, where UNRWA antenatal services reach ∼38,000 pregnant women with IFA each year; ∼97% of whom attended ANC ≥ 4 times(2). Implementation will follow a randomized, step-wedge procedure whereby MMS will start in the 1(st) 11 clinics, and six months later, the 2(nd) group of 11 clinics, providing a design for monitoring and comparing the new MMS to existing IFA programs during an initial 6–8 month period. RESULTS: Outcomes will include indicators of adoption, acceptability, coverage, adherence, fidelity, cost-efficiencies and, as a routine clinical outcome, late pregnancy anemia. CONCLUSIONS: Maternal micronutrient deficiencies are common in Palestine(3), meriting replacing IFA with MMS. This research protocol will evaluate implementation in the Gaza Strip to provide guidance for launching and improving antenatal MMS delivery throughout the UNRWA health system. FUNDING SOURCES: UNWRA, Sight and Life, Kirk Humanitarian, Vitamin Angels Alliance, Ministry of Foreign Affairs of Japan, Palestine Ministry of Health.

OBJECTIVES: Prebiotic galacto-oligosaccharides (GOS) have shown promising results in enhancing the absorption of oral supplemental iron in form of ferrous fumarate (FeFum) in iron depleted adults. The mechanism behind this effect is unknown. We aimed to assess the absorption profile of FeFum given with and without GOS using stable isotope appearance curves (SIACs). METHODS: We conducted a controlled, single-center, prospective cross-over trial in iron depleted women between Sept and Nov 2019 at the Laboratory of Human Nutrition, ETH Zurich, Switzerland. All women (n = 11; median SF = 15.2 µg/L) received two supplemental iron doses of 14 mg in form of FeFum containing 6 mg of isotopic tracers ((57)Fe or (58)Fe). The supplemental iron doses were administered in random order on d 1 and 15 after an overnight fast. The supplement combinations were: 1) (57)FeFum + 300 ml water; and 2) (58)FeFum + 15 g GOS + 300 ml water. Eleven blood samples were collected up to 24 h after administration of the iron supplements to determine serum iron appearance. Fractional iron absorption (FIA) of the isotopic tracers was calculated in whole blood collected 14 days after administration of each of the supplemental iron doses on the basis of the shift in iron isotope ratios according to the principle of isotope dilution. RESULTS: There was a trend towards increased FIA and total absolute AUC of the SIAC from FeFum given with 15 g GOS compared to without (P = 0.064; P = 0.080 respectively). Estimated timepoints of the peak serum isotope concentration were not different (P = 0.096). Relative SF and relative bioavailability correlated significantly (P = 0.037). In subjects with a lower SF the increase in bioavailability of iron when given with GOS was more pronounced compared to participants with higher SF. CONCLUSIONS: Adding 15 g GOS to FeFum did not result in a different absorption profile compared to without GOS. The enhancing effect of GOS seems to depend on iron status, even though overall the improvement of iron absorption when given with 15 g GOS did not reach significance, probably due to the small sample size. Since women with a low iron status can benefit the most from an increase in iron bioavailability, the use of GOS for the improvement of iron bioavailability from FeFum in women with very low iron stores is promising. FUNDING SOURCES: Innosuisse, Swiss Innovation Agency, in collaboration with Antistress AG - Burgerstein Vitamine.

The placenta transports iron to the fetus to support growth and development. Mice are commonly used to study placental iron transport. The subcellular locations of transferrin receptor and ferroportin 1 in iron-transporting cells in the mouse placenta have not been directly assessed. Using immunogold electron microscopy, we determined that transferrin receptor is concentrated on the intracellular vesicles in syncytiotrophoblast I while ferroportin 1 is on the basal membrane of syncytiotrophoblast II. Fluorescent imaging of maternally injected transferrin iron in the placentas collected at 6 time points postinjection (n = 1–3 animals/time point) showed that transferrin iron was taken up and metabolized within syncytiotrophoblast I within 48 h after injection. These data indicate that the intracellular iron-trafficking mechanism is distinct in different placental cells., This research demonstrates the mechanism of transferrin iron uptake at the subcellular level in mouse placental syncytiotrophoblasts

Historically, neural tube defects (NTD) in Australia affected around 1 in every 1000 births; 42% higher for Aboriginal births. Following firm evidence of the protective effect of periconceptional folate, health promotion activities encouraged increased folate intake through diet and supplements and limited voluntary food fortification. A 30% reduction in NTD was observed in non-Aboriginal but not Aboriginal infants, widening the disparity between Aboriginal and non-Aboriginal rates. Mandatory flour fortification with folic acid in 2009, led to further reductions in overall NTD, and greater reduction among Aboriginal infants, such that rates in Aboriginal and non-Aboriginal infants were similar by 2010–2014. Elimination of this disparity will make a small but important contribution to the Australian government's Closing the Gap initiative to reduce disadvantage among Aboriginal people. Long-term, complete, high-quality surveillance data on NTD have been of great value in monitoring trends in and evaluation of public health interventions for NTD in Australia.

Background Vitamin A (VA) has been demonstrated to be a regulator of adipose tissue (AT) development in adult obese models. However, little is known about the effect of VA on obesity-associated developmental and metabolic conditions in early life. Objectives We aimed to assess the effects of dietary VA supplementation during suckling and postweaning periods on the adiposity and metabolic health of neonatal and weanling rats from mothers consuming a high-fat diet (HFD). Methods Pregnant Sprague-Dawley rats were fed a normal-fat diet (NFD; 25% fat; n = 2) or an HFD (50% fat; n = 2), both with 2.6 mg VA/kg. Upon delivery, half of the rat mothers were switched to diets with supplemented VA at 129 mg/kg, whereas the other half remained at 2.6 mg VA/kg. Four groups of rat pups were designated as NFD, NFD + VA, HFD, and HFD + VA, respectively. At postnatal day (P)14, P25, and P35, pups (n = 4 or 3/group) were killed. Body weight (BW), visceral white AT (WAT) mass, brown AT (BAT) mass, uncoupling protein 1 mRNA expression in BAT, serum glucose, lipids, adipokines, and inflammatory biomarkers, as well as serum and AT redox status were assessed. Results Rat pups in the HFD group exhibited significantly higher BW, WAT mass, and serum glucose and leptin but reduced BAT mass compared with the NFD group. Without affecting the dietary intake, supplementing the HFD with VA significantly reduced the BW and WAT mass of pups but increased the BAT mass, significantly lowered the systemic and WAT oxidative stress, and modulated serum adipokines and lipids to some extent. Conclusions VA supplementation during suckling and postweaning periods attenuated metabolic perturbations caused by excessive fat intake. Supplementing maternal or infant obesogenic diets with VA or establishing a higher RDA of VA for specific populations should be studied further for managing overweight/obesity in early life.

Inconsistent associations between lipids and circulating markers of fat-soluble vitamin and carotenoid status have been reported. The aim of this hypothesis-generating study was to examine the contribution of the LC-MS-based lipidome, characterized by lipid class, carbon count, and the number of unsaturated bonds, to the interindividual variability in circulating concentrations of retinol, carotenoids, 25-hydroxyvitamin D3, α-tocopherol, γ-tocopherol, and phylloquinone in 35 overweight and obese, but healthy men. A sparse partial least-squares method was used to accomplish this aim. Highly abundant phospholipids and triglycerides (TGs) contributed to the interindividual variability in phylloquinone, α-tocopherol, and γ-tocopherol. Interindividual variability in lycopene concentrations was driven by concentrations of low-abundant TG. 25-Hydroxyvitamin D3, retinol, and the other carotenoids were not influenced by lipids. Except for lycopene, evaluation of lipids beyond class does not appear to further explain the interindividual variability in circulating concentrations of fat-soluble vitamins and carotenoids., Lipidomics does not appear to explain the interindividual variability in most circulating concentrations of fat-soluble vitamins and carotenoids beyond the interindividual variability explained by class.

OBJECTIVES: To establish an accurate and reliable method for stabilizing vitamin A (retinol) in Dried blood spots (DBS), quantifying and comparing DBS retinol concentrations with their equivalent plasma retinol levels. METHODS: Antioxidants pretreated on paper combined with vacuum treatment were used to increase retinol stability on DBS. A surrogate matrix of whole blood prepared using mixture of human erythrocytes and 2% BSA in PBS was firstly used in DBS retinol determination based on the fact that retinol is excluded from erythrocytes. RESULTS: DBS retinol was stable during 120 min of air drying and 30 days of room-temperature storage. The method was linear in the concentration range of 0.04–300 μg/mL. Accuracy was calibrated using two National Institute of Standards and Technology (NIST) calibrants generated serum at concentrations of 0.1962 and 0.3948 g/mL, relative errors (RE% values) of 0.07% and 4.95% were found, respectively. Both the between-run (n = 5) and within-run (n = 6) precision (relative standard deviations, RSD%) were below 8.42%. The spiked recoveries at 3 concentrations ranged from 86.48% to 98.13%. A reliable calibration model was first developed to convert DBS retinol concentration to the equivalent plasma retinol concentration. CONCLUSIONS: The validated method can be applied to the nutritional assessment of vitamin A, using the established calibration model, DBS retinol can compare with clinical reference ranges and with studies using serum or plasma samples. FUNDING SOURCES: The National Key Research and Development Program of China (2018YFC1002503); The National Natural Science Foundation of China (Grant No. 81,400,848, 81,701,441); The CAMS Initiative for Innovative Medicine (2016-I2M-1–008); The Beijing municipal program of medical research (Grant No. 2016–04); The National Key Research and Development Program of China (No. 2016YFC1306204).

OBJECTIVES: Selenoprotein V (SELENOV) contains a thioredoxin-like fold and a conserved CxxU motif with a potential redox function. Three experiments were performed to assess its in vivo and in vitro roles and mechanisms in coping with different oxidant insults. METHODS: In Expt.1, SELENOV knockout (KO) and wildtype (WT) mice (male, 8-wk old) were given an IP injection of saline, diquat (DQ, 12.5 mg/kg), or acetaminophen (APAP, 300 mg/kg) (n = 10), and killed 5 h after the injection to collect liver and blood. In Expt. 2, primary hepatocytes were isolated from the 2 genotypes, cultured in complete Williams's medium E, and treated with DQ (0, 0.25 and 0.75 mM) and APAP (0, 1, 3, and 6 mM) for 12 h. In Expt. 3, 293 T cells were transfected with a control plasmid (GFP) or the plasmid containing Selenov gene (full length, OE) and treated with APAP (0, 1, 2, and 4 mM) for 24 h or H(2)O(2) (0.1, 0.2, and 0.4 mM) for 12 h. RESULTS: In Expt. 1, the DQ and APAP injections caused greater (P

OBJECTIVES: Magnesium (Mg), and green tea and rhodiola extract supplementation have, in isolation, been shown to improve subjective stress perception and mood responses to acute stress. The combined capacity of these ingredients to confer protective effects during exposure to stress has yet to be evaluated. We tested the hypothesis that a combination of Mg (with B vitamins) + green tea + rhodiola would improve physiological and subjective responses to stress exposure in adults compared to placebo and Mg + B vitamins + green tea or rhodiola in isolation. METHODS: A double blind, randomised, placebo controlled, parallel group design was employed. 100 moderately stressed (DASS 13–25) adults (mean age = 25.07, SD = 0.65) received oral supplementation of either (i) Mg + B vitamins + green tea + rhodiola; (ii) Mg + B vitamins + rhodiola; (iii) Mg + B vitamins + green tea; or (iv) placebo. After supplementation participants were exposed to the Trier Social Stress Test. Subjective stress (Stress and Arousal Checklist; SACL), mood (Profile of Mood States; POMS) and salivary cortisol responses were collected over 8 hours to ascertain the effects of supplementation on stress responsivity and recovery. RESULTS: Analyses demonstrated the superiority of the combined treatment vs the ingredients in isolation and placebo. The combined treatment significantly attenuated subjective stress (SACL), and tension and total mood disturbance (POMS) ratings after acute stress exposure (all P

OBJECTIVES: We sought to describe the relationship between serum magnesium, inflammation, and chronic pain in adult primary care patient. METHODS: We sought to describe this relationship utilizing a cross-sectional analysis of medical records from 5639 adults (mean age 59 years; 42% men) seen in The University of Vermont Medical Center primary care clinics between 2015 and 2018. Patients with at least one serum magnesium level, C-reactive protein (CRP, a measure of inflammation) level, and chronic pain score (self-reported visual analogue scale) were included. RESULTS: Univariate analysis confirmed the relationship between serum magnesium and chronic pain (−0.31 points/mg/dL; 95% CI −0.47, −0.14; P

OBJECTIVES: Anemia has been a severe public health problem in India for decades, owing to its multifactorial etiology. Large surveys investigating causal factors of anemia in both male and female adolescents 10–19 years have not been available, thus our understanding of optimal interventions has been limited. Using data from a recent national micronutrient survey, our aims were to 1) describe the prevalence of anemia and micronutrient deficiencies (MNDs) in Indian adolescents and 2) examine risk factors of anemia in this population. METHODS: Data were from India's Comprehensive National Nutrition Survey (CNNS, 2016–18). Analyses were run separately for females (F; n = 3966) and males (M; n = 3944) aged 10–19 years. CNNS used a multi-stage, stratified, probability proportion to size cluster sampling design. Prevalence of anemia and micronutrient deficiencies were estimated based on age- and gender-specific WHO cutoffs, using weights for biomarker data. We examined predictors covering socio-demography (age, sex, residence, religion, caste, schooling status, parental education), nutrition (diets, anthropometry, micronutrient status), hygiene, sanitation and access to school-based services. Multivariable logistic regression models were used to examine associations between these factors and anemia. RESULTS: Forty % of females and 18% of males were anemic. The prevalence of anemia was higher in adolescents aged 15–19 years (F: 48%, M: 18%) than those aged 10–14 years (F: 32%, M: 17%). Deficiencies of iron (F: 31%, M: 12%), vitamin B(12) (F: 27%, M: 35%), folate (F: 34%, M: 39%), vitamin A (16% in both), vitamin D (F: 35%, M: 14%) and zinc (F: 28% girls, M: 35%) were also common. Iron deficiency was the strongest predictor of anemia (adjusted OR and 95% CI for F: 4.23, 2.99–5.99; M: 4.12, 2.78–6.12). Among females, other risk factors were being older (1.53, 1.09–2.15) and belonging to a disadvantaged caste (1.86, 1.12–3.09). Among males, other risk factors included being older (1.47, 1.02–2.13), being short for age (1.61, 1.12–2.31) and no mobile or internet access (1.59, 1.16–2.19). CONCLUSIONS: Anemia and MNDs are highly prevalent in Indian adolescents. Iron deficiency is a major biological risk factor for anemia as are a range of social determinants. Program design and targeting should fully account for the range of risk factors. FUNDING SOURCES: UNICEF, POSHAN.

OBJECTIVES: The adolescent period is crucial for optimizing future bone health because during these years bone accumulates rapidly accounting for up to half of adult peak bone mass (PBM). Calcium intake during this stage is critical for adequate bone mineralization but this essential nutrient, the major constituent of hydroxyapatite, is usually inadequate in the diets of US adolescents. A strategy to maximize bone mineralization is to increase calcium absorption. This could be achieved by soluble corn fiber (SCF) supplementation, which fermentation through bacteria in the lower intestine produces short-chain fatty acids and increase calcium absorption. However, there are no studies determining the long-term effects of SCF on bone mass in children. The main aim of the MetA-Bone Trial is to determine the effect of one-year SCF supplementation compared to placebo on bone mass in children with low habitual calcium intake. We hypothesize that SCF supplementation will result in a higher Bone Mineral Content. We will also determine the effects of SCF supplementation on bone biomarkers; we hypothesize that supplementation with SCF will result in higher levels of bone formation and lower bone resorption biomarkers. METHODS: A total of 240 healthy children (10–13 years), with usual low dietary calcium, will be randomized to four experimental groups for 1 year: (1) SCF (12 g/d); (2) SCF (12 g/d) + 600 mg/d of calcium; (3) Placebo (maltodextrin); and (4) Placebo + 600 mg/d of calcium. The supplements will be mixed with a flavored powder beverage to dilute in water and participants will be instructed to drink this twice per day. Bone mass will be measured using dual energy x-ray absorptiometry (DXA) at baseline, 6 and 12 months. Serum bone biomarkers will be measured at baseline and at 12 months. RESULTS: NA CONCLUSIONS: If supplementing diets with SCF lead to higher bone mass during adolescence, this could help achieve the genetic potential for PBM and to start adult life with stronger bones. Incorporating SCF into diets may be a cost-effective intervention for bone health than increasing dairy products consumption, particularly in adolescents. FUNDING SOURCES: Eunice Kennedy Shriver National Institute of Child Health & Human Development of the National Institutes of Health.

OBJECTIVES: Different chemical forms of selenium (Se) regulate distinct sets of genes and have different metabolic effects. We previously showed that mixtures of different Se compounds can have greater inhibitory effects on prostate cancer cell viability than individual chemical forms. Further, we showed that blends of different Se forms can be optimized using response surface methodology. Previous investigators showed that single forms of Se can potentiate the effects of chemotherapeutic and other agents. We hypothesized that optimized mixtures of different forms of Se would be more effective than individual Se compounds in enhancing the efficacy of Se-drug combinations. Our objective was to provide proof of principle for this hypothesis. METHODS: Response surface methodology was used to optimize a blend of methylseleninic acid (MSA), selenite (SEL) and Nano Se to achieve maximum reduction of cell viability in LNCaP prostate cancer cells. Apalutamide (APA), Enzalutamide (ENZ), Docetaxel (DOC), and Mitoxantrone (MIT) were each combined with a single Se compound, or with the optimized blend. Cell viability assays were done using Alamar Blue. RESULTS: The optimized mix of Se forms, determined using response surface methodology, was a 50:50 blend of MSA and SEL, that surprisingly did not include Nano Se. All medium supplemented with Se compounds contained 10 μM Se. MIT treatment alone reduced cell viability >90% and showed no further reductions due to added Se. Addition of either form of Se to APA, DOC, or ENZ significantly decreased cell viability compared to treatment with the drug alone (P values: 0.008 - 0.041). Similarly, in 5 out of 6 cases, combination of the optimized blend of Se compounds with the drug resulted in further decreases in cell viability, compared to the mix of drug plus one chemical form of Se (P values: 0.013 –

OBJECTIVES: We compared a high-fiber bean-rich diet with a low-carbohydrate diet in a randomized controlled trial to assess their effects on body weight, blood lipids and micronutrient adequacy. METHODS: One hundred and seventy-three women and men with a mean body mass index of approximately 36 kg · m(−2) were randomized to a high-fiber bean-rich diet that achieved mean (SD) fiber intakes of 35.5 (18.6) g · day(−1) for women and 42.5 (30.3) g · day(−1) for men, or a low-carbohydrate diet (

OBJECTIVES: To assess the following relations in women of reproductive age: 1) food security (FS) and iron status, 2) iron status and depressive symptoms (DS), 3) FS and DS and if iron status mediates this relation. METHODS: From the NHANES 2005–2010 cycles, we included 2507 non-pregnant women (20–44 yrs) with complete data for iron biomarkers, DS (Patient Health Questionnaire-9), FS (Food Security Survey Module), BMI, and sociodemographic variables. Ferritin (Ft), transferrin receptor (TfR), body iron (BI) and hemoglobin (Hb) were used to classify iron sufficient (IS), ID, and ID anemic (IDA). Logistic regression models were run (entire sample and within poverty income ratio (PIR) categories). A mediation model was also run. Age, race/ethnicity, marital status, total number of people in the household, PIR, education level, health insurance, parity, BMI, welfare status, and antidepressant use were potential covariates. RESULTS: The prevalence of ID was 43% (Ft), 2% (TfR), 37% (BI) and of anemia, 9% (Hb); IDA ranged from 19–77%. In adjusted models: 1) those with food insecurity (FIS) were 2.58 times (95% CI: 1.53–4.35) more likely to be ID (TfR); those in the lowest PIR and with FIS were 1.46 (95% CI: 1.06–2.01) (Ft), 4.00 (1.80–8.88) (TfR), and 1.43 (1.06–1.93) (BI) times more likely to be ID; 2) ID in the lowest PIR were 2.84 (95% CI:1.24–6.51) times more likely to have higher DS; 3) those with low FS had increased risk for moderate and severe DS (2.17 (95% CI: 1.28–3.68) and 3.44 (95% CI: 1.55–7.63)), and for those with very low FS, the risk for mild, moderate, and severe DS was 2.26 (95% CI: 1.20–4.24), 3.10 (95% CI: 1.67–5.76), and 9.01 (95% CI: 4.87–16.64), respectively. Iron status did not mediate the relation between FS and DS. CONCLUSIONS: Women with FIS are at increased risk of ID and DS compared to FS women, and ID women are at increased risk of DS compared to IS women. However, iron status did not mediate the relation between FS and DS. These risks are higher in those with the lowest PIR. FUNDING SOURCES: Internal Penn State funds.

OBJECTIVES: Transferrin, Tf, the protein that transports iron from the blood to the tissues via endocytosis, is believed to also transport chromium(III), Cr(III). Recently, the presence of multiple conformations was suggested by spectroscopic studies. The objective of this work is confirm whether various conformers of Cr(III)(2)-Tf exist and their potential significance for Cr(III) transport. METHODS: Cr(III) was added to apoTf in a buffered solution at pH 7.4 containing 25 mM bicarbonate at 37 °C. After time intervals, ultraviolet spectra were collected, or aliquots were removed and frozen for analysis by electron paramagnetic resonance (EPR) spectroscopy, which can distinguish free Cr(III) and Cr(III) bound to the two metal binding sites of Tf. To model the acidification of the endosome that triggers release of metal ions from Tf, the Cr(III)(2)-Tf conformer solutions were acidified by the addition of hydrochloric acid to pH 4.5 or 5.5. At time intervals after acidification, samples were again analyzed by ultraviolet and EPR spectroscopies. RESULTS: A combination of electronic and EPR studies reveal that the addition of Cr(III) to apoTf at near neutral pH in the presence of 25 mM bicarbonate results in the rapid binding of two Cr(III) accompanied and then followed by a series of conformation changes in Cr(III)(2)-Tf. These multiple conformations give rise to different spectroscopic properties and upon acidification different rates of Cr(III) release. CONCLUSIONS: The conformer of Cr(III)(2)-Tf used in most previous studies and giving rise to EPR features at g ∼ 5.1, 5.4, and 5.6 forms too slowly to be physiologically relevant; however, two previously unknown conformers of Cr(III)(2)-Tf, giving rise to an EPR feature at g ∼2 and at g ∼ 5.4, respectively, were identified. The latter of these conformers has a lifespan similar to the turnover time of transferrin and releases Cr(III) rapidly, suggesting it is probably the most physiologically significant conformer of Cr(III)(2)-Tf. FUNDING SOURCES: The University of Alabama College of Arts and Sciences Research Award.

OBJECTIVES: We sought to determine if, compared to placebo, vitamin D supplementation reduces risk of incident cancer and precancerous lesions in a cohort of overweight/obese adults with prediabetes. METHODS: The D2d cancer outcomes study (D2dCA) is an ancillary study of the Vitamin D and type 2 diabetes (D2d) study. Participants were randomized to 4000 IU/day of vitamin D(3) or placebo. At scheduled study visits (4 times/year), cancer and precancer outcomes were assessed with questionnaires. For all reported events (including possible cancer or pre-cancerous lesions, all colonoscopies, and prostate, breast, skin biopsies), clinical data were adjudicated by an oncologist who was unaware of treatment assignment. Cox proportional hazard models compared the hazard ratio (HR) of incident cancer and incident precancers between the two groups. RESULTS: Over a median follow-up of 2.9 years, among 2385 participants (mean age 60 years; mean BMI 32 kg/m(2)), there were 89 cases of incident cancer, 46 cases in the vitamin D arm and 43 cases in the placebo arm. The HR (95% CI) of incident cancer for vitamin D vs. placebo was 1.07 (0.70, 1.62). 241 participants had incident precancers, the majority of whom (239 participants) had colorectal adenomatous polyps. The HR (95% CI) of polyps for vitamin D vs. placebo was 0.83 (0.64, 1.07). Among the 259 participants who had a screening colonoscopy, the HR (95% CI) for polyps for vitamin D vs. placebo was 0.76 (0.49, 1.17). CONCLUSIONS: In D2dCA, there was no significant difference in risk of incident cancer or risk of colon polyps between vitamin D and placebo. This ancillary study was not designed or powered for these outcomes, but it provides important data that can be combined with other trials to further our insight about the role of vitamin D in cancer risk. FUNDING SOURCES: NIDDK and ODS.

OBJECTIVES: Choline is a critical nutrient for fetal development. Pregnancy studies showed that most women have choline intakes below the adequate intake (AI) level of 450 mg/d. Research on plasma free choline as an indicator of dietary choline intake showed conflicting results to date. We sought to compare plasma free choline concentration between women with different choline intakes and to explore the association between plasma free choline and dietary choline intake in early (EP) and late pregnancy (LP). METHODS: This study included data and non-fasting plasma samples from pregnant women enrolled in the Alberta Pregnancy Outcomes and Nutrition (APrON) cohort study. EP (20 weeks of gestation) dietary choline intake was estimated using a 24-hr recall. Two categories of dietary choline intake were created: 1) low choline (LCI), i.e., choline intake in 1st quartile (Q) in EP, with these women having choline intake in 1st or 2nd Q in LP (n = 61); 2) high choline intake (HCI), i.e., choline intake in 4th Q in EP and in 3rd or 4th Q in LP (n = 46). Linear mixed-effects models were used to explore the association between plasma free choline and dietary choline intake across EP and LP, after adjustment for maternal age, ethnicity and weeks of gestation. RESULTS: Median (IQR) maternal age was 32 (30–35) y, and 80% were Caucasian. LCI was 101 (86–109) and 109 (93–127) mg/day in EP and LP, respectively, and HCI was 251 (223–286) and 212 (177–274) mg/day. Plasma free choline (μmol/L) did not differ between LCI and HCI at EP [LCI: 10.6 (9.03, 12.9); HCI: 11.7 (10.2, 13.8)] and LP [LCI: 11.7 (10.6, 12.7); HCI: 12.7 (10.7, 15.8)] (P > 0.05, Wilcoxon rank-sum test). Per 10 mg of choline intake, plasma free choline increased by 0.34 (95%CI 0.12, 0.56) in those with LCI, and 0.18 (95%CI 0.050, 0.31) in women with HCI, across EP and LP after adjustment. CONCLUSIONS: In this subgroup of pregnant women, plasma free choline concentration did not reflect differences in dietary choline intake in EP or LP. This may be explained by an overall low choline intake (

OBJECTIVES: Metabolomic profiling is used to identify biological pathways and biomarkers for cardiovascular disease (CVD) and diabetes. Thus, metabolomics could be used to identify metabolic effects and demonstrate short-term efficacy of interventions designed to reduce longer-term risk of these conditions. Low potassium (K) intake has been linked to high blood pressure and increased CVD and diabetes risk. The effects of increasing K intake on metabolomic measures related to CVD and diabetes risk are not known. In this ancillary study, we tested the hypothesis that potassium chloride (KCl) supplementation would be associated with improvements in metabolomic biomarkers, such as branched-chain amino acids (BCAA) which are associated with CVD and diabetes risk. METHODS: We performed targeted mass-spectrometry-based metabolomic profiling of 60 metabolites on baseline and 12-week (end-of-study) plasma samples from 26 African-American participants with prediabetes randomized to KCl supplements vs. placebo. Principal component analysis (PCA) was used for dimensionality reduction. Univariate and multivariable analyses were used to assess differences between the two intervention arms in the changes in metabolomic factor scores and individual metabolites. RESULTS: In univariate comparisons, compared to placebo, a PCA factor composed of long-chain acylcarnitines (LCA) increased in the KCl arm (P = 0.02). In multivariable models adjusted for baseline factor score, age, and sex, this association with the LCA factor was no longer significant; but those taking KCl had significant reductions in the BCAA factor (P = 0.004) and in valine levels (P = 0.02). CONCLUSIONS: These results suggest that KCl supplementation may be associated with improvements in BCAA metabolism. Further studies among a larger population and with longer-term follow-up are warranted to verify these results and to determine if KCl supplementation may be an effective intervention for CVD and diabetes prevention. FUNDING SOURCES: NIH/Duke CTSA.

OBJECTIVES: The two objectives were 1) to evaluate the solubility of two iron casein complexes (ICCs) under a condition mimicking gastric pH, 2) to evaluate the impact of ascorbic acid (AA) on the in vitro iron absorption of ICCs after incorporation in reconstituted whole milk powder. METHODS: The in vitro solubility was determined over time after addition of diluted HCl (pH 1.7), ultracentrifugation and measurement of iron appearing in the supernatant by ICP-OES (n = 2). The impact of AA on iron uptake from the Fe compounds in reconstituted milk was determined using the in vitro digestion coupled with the Caco-2 cell model and the measurement of ferritin/total protein produced by the cells (n = 3). The molar ratio of AA to iron of 2 to 1 recommended by the WHO for iron absorption optimization has been tested with an iron level corresponding to 3.3 mg Fe/serving of milk. Ferrous sulfate (FeSO(4)), the reference compound for iron bioavailability and micronized ferric pyrophosphate (FePP), main salt used for milk fortification were used as references. RESULTS: The dissolution test showed a rapid solubilization of iron from the ICCs i.e., >75 ± 19.3% at 5 min and >89 ± 0.3% at 90 min. The kinetics of soluble iron from the complexes were like that from FeSO(4). The solubility of FePP was only 37.6 ± 4.7% at 90 min. Without AA, the iron uptake from FeSO(4) was lower than expected translating into a relative in vitro bioavailability (iRBA) of FePP and of the two ICCs to FeSO(4) of 66, 169 and 215%. This might be explained by a rapid conversion of soluble iron from FeSO(4) into Fe(3+) and insoluble iron hydroxide when the pH increased from 2 to >7 during in vitro digestion. However, with the addition of AA in the milk, iron uptake by the cells was found to be increased to levels of 341.8 ± 8.9, 124 ± 12.2, 403.1 ± 117.8 and 362.9 ± 36.9 ng ferritin/mg protein for FeSO(4), FePP and the two ICCs respectively. This translates into iRBAs to FeSO(4) of 36% for FePP and of 118 and 106% for the two ICCs. CONCLUSIONS: The solubility and the demonstrated impact of AA on Fe uptake suggest that ICCs are absorbed to a similar amount as FeSO(4) and thus provide an excellent source of Fe. FUNDING SOURCES: Société des Produits Nestlé, NPTC Konolfingen, Switzerland.

OBJECTIVES: To biosynthesize a stable-isotope labeled [(13)C]-cyanocobalamin and to measure its bioavailability in humans. METHODS: Salmonella enterica was precultured in Luria-Bertani medium and single-cell colonies were grown in No carbon E medium, supplemented with [(13)C]-ethanolamine as a sole-carbon source along with other precursors (dicyanocobinamide and dimethylbenzimidazole) and incubated for 48 h at 30°C and centrifuged. The pellets were resuspended in methanol/sodium cyanide and incubated overnight and extracts were purified by HPLC. The peaks corresponding to standard cyanocobalamin were collected and characterized by MALDI-MS. After animal testing, healthy young subjects were orally administered with [(13)C]-cyanocobalamin in a protocol that included hourly blood sampling for 12 h post-dose, and daily sampling for the next 5 days, to characterize its kinetics at a higher dose than the daily requirement (∼10x, n = 4, 2 males and females) and its appearance was modeled using a two-compartment model with early and late absorption phases. The same model was applied for lower dose at daily requirement (∼2.5 μg, n = 11 males), with sampling for 12 h post-dose. In addition, the effect of co-ingested food (n = 1, male) and the effect of parenteral replenishment of existing body vitamin B(12) stores by intramuscular injection of 1 mg hydroxocobalamin, (n = 3, males) on bioavailability was also studied. The appearance of tracer in plasma was quantified by Q-Exactive-Orbitrap-MS. RESULTS: Biosynthesized [(13)C]-cyanocobalamin was labeled with up to 7–(13)[C] atoms and at a daily requirement dose (∼2.5 μg), its mean bioavailability was 50.4 ± 8.2%, but this was also dependent on the existing body store of vitamin B(12). Two-weeks after replenishing the body stores, bioavailability increased by 1.5-fold (from 45.6 ± 0.02% to 67.8 ± 0.06%). At a higher dose (∼20 μg), its bioavailability was 6.0 ± 1.2% but the absolute amount absorbed was similar to that with the low dose. CONCLUSIONS: A novel stable isotope-based vitamin B(12) bioavailability assessment is described at low and high doses. This has implications toward rationalizing dosages used in clinical supplementation and public health fortification programs. FUNDING SOURCES: Department of Biotechnology, India (to SD and AVK) and by the Wellcome Trust/DBT India Alliance Margdarshi Fellowship [No. IA/M/14/1/501,681] awarded to AVK.

OBJECTIVES: The aim of the current study was to assay the content of vitamin E in stinging nettle (Urtica simensis) METHODS: Urtica simensis type of stinging nettle is an indigenous wild plant which is widely growing in different parts of Ethiopia. Samples of leaves were collected from Chacha, one of the central highlands in Ethiopia and portioned into sun dried, shade dried and lyophilized groups. For comparison, samples of leaves were also taken from spinach. Vitamin E family such as tocopherols ((α, β, γ, and λ) and tocotrienols (α, β, γ, and λ) were determined using high performance liquid chromatography (HPLC) at department of Food Biofunctionality, University of Hohenheim, Stuttgart, Germany. RESULTS: The results showed that the total tocols of stinging nettle in sun-dried, shade dried, and lyophilized groups were 14.1 ± 1.1 mg, 13.8 ± 1.1 mg and 16.9 ± 1.2 mg per 100 g, respectively. In spinach, this value was 3.04 ± 0.7 mg/100 g. Of all vitamin E family, α- tocopherol was the maximum and identified in shade dried group (16.5 ± 1.2 mg/100 g). As compared to stinging nettle, spinach contained very small amount of α- tocopherol (1.7 ± 0.5 mg/100 g). CONCLUSIONS: In conclusion, Urtica simensis type of stinging nettle contains considerable amount of tocols and can serve as potential resource of vitamin E. Further research is warranted on the nutritional and medicinal values of Urtica simensis stinging nettle. FUNDING SOURCES: None.

OBJECTIVES: Hepcidin is a negative regulator of extracellular iron availability and dietary iron absorption. Previous studies have shown that hepcidin increases with prolonged endurance exercise, but not in those with low iron stores (serum ferritin 0.05 for all). CONCLUSIONS: The post-exercise hepcidin response occurs in male, but not female, collegiate distance runners with low iron stores. These findings indicate that declines in iron status in female distance runners are independent of exercise-induced hepcidin. FUNDING SOURCES: Intramural funds to S.R.H.

OBJECTIVES: Assessment of erythrocyte linoleic: dihomo-γ-linolenic acid (LA: DGLA) ratio as a biomarker of Zn status, and structure and function of cecal microbiota following the consumption of a Zn-biofortified wheat diet. METHODS: The efficacy of the LA: DGLA ratio to predict the Zn status of subjects consuming a wheat-based diet, a representative diet of the target Zn-deficient populations, was evaluated in-vivo (Gallus gallus). Two groups (n = 15) were fed two different diets, a “high-Zn” diet (46.5 ppm Zn) and a “low-Zn” diet (32.8 ppm Zn), for 6 weeks. Dietary Zn-intake, body-weight, serum zinc, intestinal microbiome, expression of zinc related proteins, and the erythrocyte fatty acid profile were assessed (LA: DGLA ratio). RESULTS: Serum and tissues Zn concentrations were greater in the high-Zn group (P

OBJECTIVES: The trace element selenium (Se) is known mainly for its antioxidant properties and is critical for proper brain function. The role of Se in regulating energy metabolism, and the sexually dimorphic nature of Se functions, however, are underappreciated, and warrant increased attention. Recent work in our lab has highlighted the importance of Se utilization in hypothalamic regulation of energy metabolism. Dietary Se is incorporated into selenoproteins in the form of the unique amino acid selenocysteine (Sec). The objective of this study was to assess the role of selenoproteins in Agouti-related peptide (Agrp)-positive neurons, an orexigenic sub-population of the hypothalamus. METHODS: We generated mice with Agrp-Cre-driven deletion of selenocysteine tRNA (Trsp-Agrp KO mice), which is essential for Sec incorporation into selenoproteins, thus ablating selenoprotein synthesis in Agrp-positive neurons. The metabolic phenotype of Trsp-Agrp KO mice challenged with a high-fat diet was characterized via glucose tolerance test (i.p. injection) and the use of analytical chambers to measure food intake and respiratory metabolism. Prior to sacrifice, mice were challenged with leptin (i.p. injection) to assess neuronal leptin responsivity via immunohistochemistry and western blot. Brown adipose tissue (BAT) morphology and thermogenic protein expression were also analyzed. RESULTS: Female Trsp-Agrp KO mice displayed resistance to diet-induced obesity, which was accompanied by improved glucose tolerance and elevated energy expenditure levels without changes in food intake. Female Trsp-Agrp KO mice also had greater leptin sensitivity and showed signs of elevated BAT thermogenesis. Male Trsp-Agrp KO mice displayed no changes in metabolic phenotype. CONCLUSIONS: Loss of selenoproteins in Agrp-positive neurons of the hypothalamus promotes energy expenditure and reduces diet-induced obesity in a sexually dimorphic manner, leading to resistance to a high-fat diet in females. FUNDING SOURCES: This work was funded by grant support from the National Institute of Diabetes and Digestive and Kidney Diseases (MJB) and Ola HAWAII, a grant from the National Institute on Minority Health and Health Disparities.

OBJECTIVES: Developing countries rely on poorly bioavailable plant-based sources of iron (i.e., FeSO(4)), leading to iron deficiency anemia. Heme iron is more bioavailable, but predominantly found in red meat. Iron chlorophyllin (IC) utilizes the porphyrin ring of plant-based chlorophyll to bind iron. IC has previously been shown to survive digestion, and to deliver iron to Caco-2 cells (measured as increased ferritin levels). However, the dose-response of IC treatment has not been assessed. We hypothesized that increasing IC concentrations would increase Caco-2 cell iron in a dose dependent manner. We also hypothesized that novel IC metabolites would be observed following in vitro digestion and incubation with Caco-2 cells. METHODS: In vitro digestion was performed using FeSO(4) and hemoglobin as positive controls, deionized water as a negative control, and IC as the treatment (n = 3). Two doses (low and high) of both FeSO(4) and hemoglobin were tested, in addition to four doses of IC (i.e., 2, 8, 34, and 81 ppm iron). Gastric and intestinal phases of digestion were mimicked, and digested chyme was centrifuged and filtered before incubation with differentiated Caco-2 human intestinal cells for 4 h. Cell were harvested, and iron concentrations in the chyme, micelles and harvested cells were tested using furnace atomic absorption spectrometry at 248.3 nm, and concentrations compared using one-way ANOVA, followed by Tukey's post-hoc test (P

OBJECTIVES: To evaluate the association between depression and consumption of retinol, vitamins C and E, selenium and zinc in participants of the Brazilian Longitudinal Study of Adult Health (ELSA-Brasil) baseline data. METHODS: This study is part of the ELSA-Brasil, a population-based cohort study with 15,105 civil servants aged 35–74 years. In this analysis, we included cross-sectional data from the baseline examination conducted from August 2008 to December 2010. Antioxidants consumption was measure by The Food Frequency Questionnaire (FFQ) using the Nutrition Data System for Research (NDSR) software. Depression disorder was accessed by a Clinical Interview Schedule Revised (CIS-R). The use of antidepressant drugs was recorded during a face-to-face systematic interview about use of continuous medication in the prior two weeks. Logistic regression models were built comparing two groups: DEP1 - no depression by CIS-R and no current antidepressants use, and DEP2 - depression by CIS-R or current use of antidepressant. Model 1 yield crude values and model 2 adjusted by age, sex and total energy. These analyses were carried out using the software Stata and P

OBJECTIVES: High blood methylmalonic acid (MMA) and homocysteine (tHcy) and low serum B-12 and holotranscobalamin (holoTC) concentrations indicate vitamin B-12 deficiency. The diagnosis is often contradictory when using any test independently. A mathematical model including all 4 biomarkers, named combined indicator of vitamin B-12 status (cB(12)), has been proposed. Our goal was to describe cB(12) in US adults and estimate the prevalence of low or transitional vitamin B-12 status compared to conventional single biomarkers. METHODS: We assessed cB(12) in persons ≥20 y participating in NHANES from 1999 to 2004 from 3 biomarkers: serum vitamin B-12, MMA, and plasma tHcy. RESULTS: The following groups had significantly lower cB12: persons ≥70 y compared with younger age groups, males compared with females, non-Hispanic whites compared with non-Hispanic blacks and Mexican Americans, and non-users of vitamin B-12 containing supplements compared with users. Shorter fasting times and impaired renal function also resulted in lower cB12. There was a strong significant association of cB12 with serum vitamin B-12 (Spearman r = 0.75), MMA (r = −0.70), and tHcy concentrations (r = −0.59). The prevalence of vitamin B-12 deficiency varied with the biomarker and cutoff used: 2.2% and 13% for serum vitamin B-12 13 μmol/L. Using the proposed cB12 cutoff of < −0.5, 2.7% of adults had “low or transitional vitamin B-12 status”. CONCLUSIONS: Based on the new combined indicator using 3 biomarkers a larger portion of US adults had adequate vitamin B-12 status (97%) compared to conventional single biomarkers (between 85% and 94%), likely indicating the higher specificity of cB(12). It is unclear whether the availability of holoTC data would change these findings. FUNDING SOURCES: This work was supported by direct appropriations from US Congress.

OBJECTIVES: We hypothesized that meal fat co-administered with fat-soluble α-tocopherol (vitamin E) facilitates vitamin E absorption and increases relative bioavailability in the immediate post-prandial phase, when compared with non-fat meals. We tested this hypothesis using deuterated oral α-tocopherol co-administered with breakfast containing meal fat (40% group) and without meal fat (0% group) in hospitalized healthy women. We also evaluated the role of subsequent meals in modulating the vitamin E relative bioavailability by fasting patients for 12 h following a breakfast meal with 0% fat (0% fat-fasting group). We compared area under the curve (AUC) for oral d3-α-tocopherols %enrichment at 0–4 h, 0–12 h and 0–24 h. METHODS: Custom-synthesized deuterated d3-α-tocopherol was co-administered with breakfast meal with and without fat, with subsequent serial sampling. Enrolled subjects were healthy women hospitalized for 5–6 days at the NIH Clinical Research Center. RESULTS: The AUC0–4 h for 40% fat group was more than twice 0% fat group (6.4 ± 1.8 vs 2.3 ± 0.7). This difference was erased following ingestion of meals containing 30% fat at 4 h and 8 h post-dosing, with AUC0–12 h for 40% and 0% fat groups (96.8 ± 10.2 vs 107.4 ± 8.) and AUC0–24 h (254 ± 16.3 vs 301.8 ± 19.7). To evaluate the effect of fasting, we compared 0% fat group with 0% fat-fast groups. At 4 h and 8 h post-dosing, the 0% fat group received meals with 30% fat, while 0% fat-fast group remained in fasting state. We therefore predicted and found significantly higher AUC0–12 h in the 0% fat group compared with the 0% fat-fast (107.4 ± 8.1 vs 41.1 ± 5.2 P

OBJECTIVES: To assess the practicality and assumptions of the high Fe concentration approach for bean Fe biofortification in east Africa. METHODS: A collection of 76 marketplace samples (East Africa Marketplace Collection; EAMC) were assembled, consisting of multiple color classes from locations in Uganda, Rwanda, Democratic Republic of Congo, Burundi, Ethiopia, Kenya and Tanzania. In addition, because market samples can be a mixture of seed varieties within a market class, 170 samples from the Africa CIAT Collection were also assembled that represent beans of common markets in the region. Iron concentration was measured via inductively coupled plasma emission spectroscopy (ICP-ES) and Fe bioavailability via an established Caco-2 cell bioassay. Using these measures, the following assumptions of high Fe bean Fe biofortification approach were assessed: 1) the average Fe concentration in beans currently consumed in east Africa is approximately 50 μg/g (dry weight), 2) a 40 μg/g increase (target value 90 μg/g) can be sustained through traditional breeding, and 3) iron bioavailability from the biofortified bean will not decrease substantially to negate the increase in Fe concentration. RESULTS: The average EAMC bean Fe concentration was 72 μg/g, ranging from 52–93 μg/g, with a couple of outlying varieties at 105 μg/g (MAC9) and 129 μg/g (MAC49). The Africa CIAT collection averaged 67.5 μg/g with a range of 51–90 μg/g. The 18 biofortified varieties within the EAMC averaged 73 μg/g (range of 55–94 μg/g), which is essentially equal to the overall mean (70 μg/g) and range (54–93 μg/g) of the non-biofortified bean varieties in the EAMC. Using a Caco-2 cell bioassay to measure Fe bioavailability of the EAMC, the biofortified varieties did not deliver any additional Fe relative to non-biofortified varieties. CONCLUSIONS: The results indicate that the assumptions of the high Fe bean breeding approach are not met in the typical east African market place. Furthermore, based on the Fe content and bioavailability data collected from this study, the biofortified bean varieties from these markets are providing no additional dietary Fe. An alternative approach for bean Fe biofortification, such as enhanced Fe bioavailability should be pursued. FUNDING SOURCES: USDA.