New perspectives on the regulation of iron absorption via cellular zinc concentrations in humans.

Iron deficiency is the most prevalent nutritional deficiency, affecting more than 30% of the total world's population. It is a major public health problem in many countries around the world. Over the years various methods have been used with an effort to try and control iron-deficiency anemia. However, there has only been a marginal reduction in the global prevalence of anemia. Why is this so? Iron and zinc are essential trace elements for humans. These metals influence the transport and absorption of one another across the enterocytes and hepatocytes, due to similar ionic properties. This paper describes the structure and roles of major iron and zinc transport proteins, clarifies iron-zinc interactions at these sites, and provides a model for the mechanism of these interactions both at the local and systemic level. This review provides evidence that much of the massive extent of iron deficiency anemia in the world may be due to an underlying deficiency of zinc. It explains the reasons for predominance of cellular zinc status in determination of iron/zinc interactions and for the first time thoroughly explains mechanisms by which zinc brings about these changes.

Supplemental dietary inulin of variable chain lengths alters intestinal bacterial populations in young pigs.

Previously, we showed that supplementation of diets with short-chain inulin (P95), long-chain inulin (HP), and a 50:50 mixture of both (Synergy 1) improved body iron status and altered expression of the genes involved in iron homeostasis and inflammation in young pigs. However, the effects of these 3 types of inulin on intestinal bacteria remain unknown. Applying terminal restriction fragment length polymorphism analysis, we determined the abundances of luminal and adherent bacterial populations from 6 segments of the small and large intestines of pigs (n = 4 for each group) fed an iron-deficient basal diet (BD) or the BD supplemented with 4% of P95, Synergy 1, or HP for 5 wk. Compared with BD, all 3 types of inulin enhanced (P < 0.05) the abundance of beneficial bifidobacteria and lactobacilli in the microbiota adherent to intestinal mucus of various gut segments of pigs. These changes were seen as proximal as in the jejunum with P95 but did not appear until the distal ileum or cecum with HP. Similar effects of inulin on bacterial populations in the lumen contents were found. Meanwhile, all 3 types of inulin suppressed the less desirable bacteria Clostridium spp. and members of the Enterobacteriaceae in the lumen and mucosa of various gut segments. Our findings suggest that the ability of dietary inulin to alter intestinal bacterial populations may partially account for its iron bioavailability-promoting effect and possibly other health benefits.

Biofortified black beans in a maize and bean diet provide more bioavailable iron to piglets than standard black beans.

Our objective was to compare the capacities of biofortified and standard black beans (Phaseolus vulgaris L.) to deliver iron (Fe) for hemoglobin (Hb) synthesis. Two lines of black beans, one standard and the other biofortified (high) in Fe (71 and 106 microg Fe/g, respectively), were used. Maize-based diets containing the beans were formulated to meet the nutrient requirements for swine except for Fe (Fe concentrations in the 2 diets were 42.9 +/- 1.2 and 54.6 +/- 0.9 mg/kg). At birth, pigs were injected with 50 mg of Fe as Fe dextran. At age 28 d, pigs were allocated to the experimental diets (n = 10). They were fed 2 times per day for 5 wk and given free access to water at all times. Body weights and Hb concentrations were measured weekly. Hb repletion efficiencies (means +/- SEM) did not differ between groups and, after 5 wk, were 20.8 +/- 2.1% for the standard Fe group and 20.9 +/- 2.1% for the high Fe group. Final total body Hb Fe contents did not differ between the standard [539 +/- 39 mg (9.7 +/- 0.7 micromol)] and high Fe [592 +/- 28 mg (10.6 +/- 0.5 micromol)] bean groups (P = 0.15). The increase in total body Hb Fe over the 5-wk feeding period was greater in the high Fe bean group [429 +/- 24 mg (7.7 +/- 0.4 micromol)] than in the standard Fe bean group [361 +/- 23 mg (6.4 +/- 0.4 micromol)] (P = 0.034). We conclude that the biofortified beans are a promising vehicle for increasing intakes of bioavailable Fe in human populations that consume beans as a dietary staple.

Supplemental dietary inulin influences expression of iron and inflammation related genes in young pigs.

We have previously shown improved hemoglobin (Hb) repletion efficiency by supplementing a 50:50 mixture of short (P95) and long-chain (HP) inulin (Synergy 1, BENEO-Orafti) into a corn-soybean meal-basal diet (BD) for young pigs. In this study, weanling pigs (5 or 6 wk old) were fed the BD or the BD + 4% of P95, HP, or Synergy 1 (50:50 mixtures of HP and P95) for 5-7 wk. Blood Hb concentrations of pigs were measured weekly and digesta samples were collected at the end of the trial. In a replicate experiment, total RNA was isolated from the liver and mucosa of duodenum, ileum, cecum, and colon of all pigs at the end of the trial. Relative mRNA expression of 27 genes, including iron and inflammation-related genes, was quantified using real-time quantitative-PCR. Although all 3 types of inulin resulted in similar improvements (P < 0.05) in blood Hb concentration and liver ferritin protein amount, neither type of inulin was detectable in the digesta of cecum or colon. Supplemental inulin enhanced the expression of iron-storing protein genes but decreased that of inflammation-related genes. Such effects were more pronounced (P < 0.05) in the mucosa of the lower than the upper gut and were seen on 7 genes in liver. In conclusion, all 3 types of inulin shared similar efficacy and possibly similar modes of action in improving dietary iron utilization by young pigs. Suppressing inflammation-induced genes that can negatively influence iron metabolism might help explain the benefit of inulin.

Effects of ascorbic acid, phytic acid and tannic acid on iron bioavailability from reconstituted ferritin measured by an in vitro digestion-Caco-2 cell model.

The effects of ascorbic acid (AA), phytate and tannic acid (TA) on Fe bioavailability from Fe supplied as reconstituted ferritin were compared with FeSO4 using an in vitro digestion-Caco-2 cell model. Horse spleen apoferritin was chemically reconstituted into an animal-type ferritin (HSF) and a plant-type ferritin (P-HSF) according to the typical ratios of Fe:P found in these molecules. In the presence of AA (Fe:AA molar ratio of 1:20), significantly more Fe was absorbed from FeSO4 (about 303 %), HSF (about 454 %) and P-HSF (about 371 %) when compared with ferrous sulfate or ferritin without AA. Phytic acid (PA; Fe:PA molar ratio of 1:20) significantly reduced Fe bioavailability from FeSO4 (about 86 %), HSF (about 82 %) and P-HSF (about 93 %) relative to FeSO4 and the ferritin controls. Treatment with TA (Fe:TA molar ratio of 1:1) significantly decreased Fe bioavailability (about 97 %) from both FeSO4 and the ferritin samples. AA was able to partially reverse the negative effect of PA (Fe:PA:AA molar ratio of 1:20:20) on Fe bioavailability but did not reverse the inhibiting effect of TA (Fe:TA:AA molar ratio of 1:1:20) on Fe bioavailability from ferritin and FeSO4. Overall, there were no significant differences in bioavailable Fe between P-HSF, HSF or FeSO4. Furthermore, the addition of AA (a known promoter) or the inhibitors, PA and TA, or both, did not result in significant differences in bioavailable Fe from ferritin relative to FeSO4. The results suggest that Fe in the reconstituted ferritin molecule is easily released during in vitro digestion and interacts with known promoters and inhibitors.

Relationships between faecal phytate and mineral excretion depend on dietary phytate and age.

We investigated the adverse effect of phytate on mineral absorption and the effect of dietary phytate and age on the relationship between faecal phytate and faecal mineral excretion. Fourteen young women (aged 19-24 years) and fourteen elderly women (64-75 years) were studied for two metabolic periods (MP). In MP1, the subjects consumed a controlled high-phytate (HP) diet for 10 d; in MP2, they were on a low-phytate (LP) diet for 10 d. In each period, diet samples and complete faecal samples for 5 d were collected to analyse phytate and mineral contents. Mineral concentrations in diet and faeces were measured by inductively coupled plasma-atomic emission spectrometry. Linear regression analysis was used to examine the associations between faecal phytate and mineral excretion. The degradation rate of dietary phytate was about 77% for young women, which was significantly lower than that of elderly women (86%) (P < 0.05). Faecal phytate excretion was positively correlated with mineral excretion (Ca, P, Fe and Zn) in both the HP and LP diet groups in young women (P < 0.05). The linear relationship tended to be greater during the LP diet period compared with the HP diet period in young women. However, no association was found between phytate excretion and mineral excretion in elderly women. In summary, undegraded dietary phytate (10-20%) had a negative effect on mineral absorption in young women, and the relationship between faecal phytate and mineral excretion was affected by both dietary phytate and age.

Iron bioavailability from maize and beans: a comparison of human measurements with Caco-2 cell and algorithm predictions.

BACKGROUND: An in vitro digestion and Caco-2 cell model may predict iron bioavailability to humans; however, direct comparisons are lacking. OBJECTIVE: The objective was to test the differences in iron bioavailability between 2 maize varieties and 2 bean varieties (white beans and colored beans) by comparing human, Caco-2, and algorithm results. DESIGN: Two randomized, 2 x 2 factorial experiments compared women's iron absorption from 2 maize varieties (ACR and TZB; n = 26) and 2 bean varieties (great northern and pinto; n = 13), each fed with and without ascorbic acid (AA) from orange juice. Nonheme iron bioavailability was determined from 2-wk retention of extrinsic radioiron tracers and was compared with Caco-2 cell and algorithm results from identical meals. RESULTS: Without AA supplementation, women absorbed only about 2% of the iron from the maize or bean meals. The results were unaffected by the variety of either maize or beans. Adding AA (15-20 molar ratios of AA:iron) roughly tripled the iron absorption (P < 0.0001) from all test meals. Although the Caco-2 model predicted a slightly improved bioavailability of iron from ACR maize than from TZB maize (P < 0.05), it accurately predicted relative iron absorption from the maize meals. However, the Caco-2 model inaccurately predicted both a considerable difference between bean varieties (P < 0.0001) and a strong interaction between bean varieties and enhancement by AA (P < 0.0001). The algorithm method was more qualitatively than quantitatively useful and requires further development to accurately predict the influence of polyphenols on iron absorption. CONCLUSIONS: Caco-2 predictions confirmed human iron absorption results for maize meals but not for bean meals, and algorithm predictions were only qualitatively predictive.

Screening of iron bioavailability patterns in eight bean (Phaseolus vulgaris L.) genotypes using the Caco-2 cell in vitro model.

The common bean ( Phaseolus vulgaris) is an important staple plant food in the diets of people of Latin America, East Africa,and other regions of the developing world. It is also a major source of dietary iron. The primary goal of this research was to use an in vitro digestion/Caco-2 model to study iron bioavailability in eight genotypes (three Mesoamerican and five Andean) that represent the diversity of grain types in this crop. Complementing this goal, we measured the distribution of both iron and phytate in different bean grain tissues (cotyledon, seed coats, and embryos). Seed coats were confirmed to be the exclusive tissue containing polyphenols. The removal of the seed coat and associated polyphenols improved Caco-2 iron bioavailability, and significant differences were observed between genotypes. The addition of ascorbate enhanced iron bioavailability and exposed additional differences in Fe availability among the genotypes. These results indicate that iron accumulation and in vitro iron bioavailability vary among bean genotypes and that polyphenols had greater inhibitory effects on Caco-2 iron bioavailability as compared to phytate.

Kaempferol in red and pinto bean seed (Phaseolus vulgaris L.) coats inhibits iron bioavailability using an in vitro digestion/human Caco-2 cell model.

Four different colored beans (white, red, pinto, and black beans) were investigated for factors affecting iron bioavailability using an in vitro digestion/human Caco-2 cell model. Iron bioavailability from whole beans, dehulled beans, and their hulls was determined. The results show that white beans contained higher levels of bioavailable iron compared to red, pinto, and black beans. These differences in bioavailable iron were not due to bean-iron and bean-phytate concentrations. Flavonoids in the colored bean hulls were found to be contributing to the low bioavailability of iron in the non-white colored beans. White bean hulls contained no detectable flavonoids but did contain an unknown factor that may promote iron bioavailability. The flavonoids, kaempferol and astragalin (kaempferol-3-O-glucoside), were identified in red and pinto bean hulls via HPLC and MS. Some unidentified anthocyanins were also detected in the black bean hulls but not in the other colored bean hulls. Kaempferol, but not astragalin, was shown to inhibit iron bioavailability. Treating in vitro bean digests with 40, 100, 200, 300, 400, 500, and 1000 microM kaempferol significantly inhibited iron bioavailability (e.g., 15.5% at 40 microM and 62.8% at 1000 microM) in a concentration-dependent fashion. Thus, seed coat kaempferol was identified as a potent inhibitory factor affecting iron bioavailability in the red and pinto beans studied. Results comparing the inhibitory effects of kaempferol, quercitrin, and astragalin on iron bioavailability suggest that the 3',4'-dihydroxy group on the B-ring in flavonoids contributes to the lower iron bioavailability.

Potential for improving bioavailable zinc in wheat grain (Triticum species) through plant breeding.

A "whole-body" radioassay procedure was used to assess retention and absorption by rats of Zn in mature kernels of whole grain wheat harvested from 28 genotypes (Triticum spp.) grown in nutrient solution supplied with 2 microM ZnSO4 radiolabeled with 65Zn. Grain-Zn concentration differed among genotypes and ranged from 33 to 149 microg g(-1) of dry weight (DW); similarly, grain-Fe concentration varied approximately 4-fold, from 80 to 368 microg g(-1) of DW. Concentrations of Zn and Fe in the grain were positively correlated. Therefore, selecting genotypes high in grain-Zn also tends to increase grain-Fe concentration. Concentrations of myo-inositolhexaphosphate (phytate) in the wheat grain varied from 8.6 to 26.1 micromol g(-1) of DW. Grain intrinsically labeled with 65Zn was incorporated into test meals fed to Zn-depleted rats. All rats readily ate the test meals, so that Zn intake varied directly with grain-Zn concentration. As determined by the percentage of 65Zn absorbed from the test meal, the bioavailability to rats of Zn in the wheat genotypes ranged from about 60 to 82%. The amount of bioavailable Zn (micrograms) in the grain was positively correlated to the amount of Zn accumulated in the grain. There was a significant negative correlation between grain-phytate levels and percentage of Zn absorbed from the wheat grain, but the effect was not large. These results demonstrate that concentrations of Zn in whole-wheat grain, as well as amounts of bioavailable Zn in the grain, can be increased significantly by using traditional plant-breeding programs to select genotypes with high grain-Zn levels. Increasing the amount of Zn in wheat grain through plant-breeding contrivances may contribute significantly to improving the Zn status of individuals dependent on whole grain wheat as a staple food.

Agriculture: the real nexus for enhancing bioavailable micronutrients in food crops.

Human existence requires that agriculture provide at least 50 nutrients (e.g., vitamins, minerals, trace elements, amino acids, essential fatty acids) in amounts needed to meet metabolic demands during all seasons. If national food systems do not meet these demands, mortality and morbidity rates increase, worker productivity declines, livelihoods are diminished and societies suffer. Today, many food systems within the developing world cannot meet the nutritional needs of the societies they support mostly due to farming systems that cannot produce enough micronutrients to meet human needs throughout the year. Nutrition transitions are also occurring in many rapidly developing countries that are causing chronic disease (e.g., cancer, heart disease, stroke, diabetes, and osteoporosis) rates to increase substantially. These global developments point to the need to explicitly link agricultural technologies to human health. This paper reviews some ways in which agriculture can contribute significantly to reducing micronutrient malnutrition globally. It concludes that it is imperative that close linkages be forged between the agriculture, nutrition and health arenas in order to find sustainable solutions to micronutrient malnutrition with agriculture becoming the primary intervention tool to use in this fight.

Biotechnology, biofortification, and global health.

Deficiencies of micronutrients such as iron, zinc, and vitamin A afflict over three billion people (more than 50% of the world's population), most of them women, infants, and children in resource-poor families in the developing world. This global crisis in nutritional health is the result of dysfunctional food systems that do not consistently supply enough of these essential nutrients to meet the nutritional requirements of high-risk groups. Deficiencies of micronutrients result in increased morbidity and mortality rates, lost worker productivity, stagnated national development, permanent impairment of cognitive development in infants and children, and large economic costs and suffering to those societies affected. Because agricultural systems are the primary source of all micronutrients for all people, changes in agricultural policies and systems must be made that will ensure consistent and adequate supplies of all essential nutrients to all people. Additionally, the nutrition and health sectors must turn to agricultural interventions as a primary tool in their efforts to eliminate malnutrition from the world if they want to ensure sustainability. Biotechnological advances show great promise for improving the output of bioavailable micronutrients from agricultural systems that feed the poor. This paper reviews some of these opportunities and discusses the questions and concerns that should be raised when these technologies are used to improve the micronutrient status of vast numbers of people who are dependent on staple food crops for their sustenance. Further, important issues surrounding micronutrient bioavailability and plant food factors that affect it are discussed.

Transport interactions between cadmium and zinc in roots of bread and durum wheat seedlings.

Field studies have shown that the addition of Zn to Cd-containing soils can help reduce accumulation of Cd in crop plants. To understand the mechanisms involved, this study used 109Cd and 65Zn to examine the transport interactions of Zn and Cd at the root cell plasma membrane of bread wheat (Triticum aestivum L.) and durum wheat (Triticum turgidum L. var. durum). Results showed that Cd2+ uptake was inhibited by Zn2+ and Zn2+ uptake was inhibited by Cd2+. Concentration-dependent uptake of both Cd2+ and Zn2+ consisted of a combination of linear binding by cell walls and saturable, Michaelis-Menten influx across the plasma membrane. Saturable influx data from experiments with and without 10 micro m concentrations of the corresponding inhibiting ion were converted to double reciprocal plots. The results revealed a competitive interaction between Cd2+ and Zn2+, confirming that Cd2+ and Zn2+ share a common transport system at the root cell plasma membrane in both bread and durum wheat. The study suggests that breeding or agronomic strategies that aim to decrease Cd uptake or increase Zn uptake must take into account the potential accompanying change in transport of the competing ion.

Reversed-phase liquid chromatographic determination of phytometallophores from Strategy II Fe-uptake species by 9-fluorenylmethyl chloroformate fluorescence.

An HPLC method to quantitate phytometallophores (phytosiderophores) exuded from roots of barley (Hordeum vulgare L.) growing in nutrient solution culture was developed. 9-Fluorenylmethyl chloroformate (FMOC) derivatives of phytometallophores were separated on a C18 reverse-phase column using a sodium acetate (pH 7.2) and acetonitrile-methanol gradient over 20 min followed by fluorescence detection. Detection limits ranged from 15 to 370 pmol depending on the particular phytometallophore. The effectiveness of this method was demonstrated using the response of barley seedlings to Fe-sufficient and Fe-deficient nutrient solution conditions. Phytometallophores collected in root washings of Fe-deficient barley seedlings increased with plant age while phytometallophore release from Fe-adequate roots was negligible.

Mother-child participation in conversation about the past: relationships to preschoolers' theory of mind.

Forty 3.5 to 4.5-year-olds discussed 3 past events with their mothers and completed a set of theory of mind tasks indexing their ability to reason about conflicting mental representations and their understanding of knowledge. Semipartial correlations and analyses of covariance showed that children's theory of mind scores were related to their participation in memory conversations, independent of age and linguistic skill. The frequency with which mothers provided new information was related to children's theory of mind scores, although mothers' direct replies to children were generally unrelated to children's understanding of mind. This research takes an important step toward examining the relevance of theory of mind skills to real-world, social interaction. The results have implications for explaining the emergence of autobiographical memory.

Young children's understanding of conflicting mental representation predicts suggestibility.

This study examined the relation between developmental suggestibility effects and preschoolers' emerging ability to reason about conflicting mental representations (CMRs). Three- to 5-year-olds listened to a story accompanied by pictures. Following a 4-min delay, children answered straightforward and misleading questions about the story. One week later, their memory for the story was assessed. Children also completed tasks indexing their ability to reason about CMRs. Stepwise regression analyses revealed that suggestibility was negatively related to performance on CMR tasks. This finding remained significant after controlling for age, children's level of initial encoding of the event, and their ability to retrieve event details when not misled. An integration is proposed between children's theory of mind and source monitoring that may help to explain early developmental changes in suggestibility.

Comparison of iron bioavailability from 15 rice genotypes: studies using an in vitro digestion/caco-2 cell culture model.

An in vitro digestion/Caco-2 model was used to compare iron bioavailability from 15 selected Fe-dense and normal genotypes of unpolished rice from the International Rice Research Institute. Iron uptake was determined using Caco-2 cell ferritin formation in response to exposure to a digest of the cooked rice. Iron bioavailabilities from all rice genotypes were ranked as a percent relative to a control variety (Nishiki). Iron concentration in the rice samples ranged from 14 to 39 microg/g. No correlation was observed between Fe uptake and grain-Fe concentration. Furthermore, phytic acid levels were not correlated with Fe bioavailability. Genotypes with low Fe bioavailability (Tong Lan Mo Mi, Zuchein, Heibao, and Xua Bue Nuo) were noticeably more brown to purple in color. The results suggest that certain unknown compounds related to rice grain color may be a major factor limiting Fe bioavailability from unpolished rice.

Iron and zinc absorption from two bean (Phaseolus vulgaris L.) genotypes in young women.

Extrinsic and intrinsic iron and zinc labels were used to test iron and zinc absorption from two bean (Phaseolus vulgaris) genotypes, containing normal (common beans, CB) or higher (HFeZnB) iron and zinc concentrations, fed in single meals to young women with low iron reserves. The women were divided into two groups, with one receiving a CB test meal (n = 12) and the other, an HFeZnB test meal (n = 11). The beans were intrinsically labeled hydroponically with (55)Fe (CB and HFeZnB) and with (70)Zn (HFeZnB). Concentrations of zinc and iron were 98 and 65% higher, respectively, in HFeZnB as compared to CB, but phytic acid contents were similar. Extrinsic labels were (59)Fe (CB and HFeZnB), (67)Zn (CB), and (68)Zn (HFeZnB). Iron and zinc percent absorption levels were calculated from radio-iron activity in red blood cells and from urinary excretion of zinc isotopes. Intrinsic and extrinsic iron absorption measures were highly correlated (R (2) = 0.986) (average extrinsic/intrinsic ratio was 1.00). Iron absorption was low (geometric mean < 2%) in both bean types, and total iron absorbed was not different between types. Intrinsic zinc absorption from the HFeZn beans was higher than extrinsic absorption (15.2% vs 13.4%, p < 0.05) (average extrinsic/intrinsic was 0.90). The correlation between intrinsic and extrinsic zinc measures was not as high as that for iron (R (2) = 0.719). Percent zinc absorption levels were similar in both bean types, but total extrinsic zinc absorbed was 90% higher (p < 0.05) from the HFeZnB meal. Thus, the less expensive and time-consuming extrinsic labeling may be used to screen various varieties of beans for iron bioavailability in humans, but it underestimates zinc absorption by approximately 10%. Selective breeding for high-zinc bean genotypes may improve zinc status. However, high-iron genotypes appear to have little effect on iron status when fed alone in single meals to women with low iron reserves.

Bioavailability of iron from spinach using an in vitro/human Caco-2 cell bioassay model.

Spinach (Spinacia oleracea) cv Whitney was tested for iron bioavailabilty using an in vitro human intestinal cell culture ferritin bioassay technique previously developed. Spinach was cultured in a growth chamber for 33 days, harvested, and freeze-dried. Total iron in the samples was an average of 71 micrograms/g dry weight. Spinach was digested in vitro (pepsin and 0.1 M HCl followed by pancreatin and 0.1 M NaHCO3) with and without the addition of supplemental ascorbic acid. Caco-2 cell cultures were used to determine iron bioavailability from the spinach mixtures. Production of the iron-binding protein ferritin in the Caco-2 cells showed the supplemental ascorbic acid doubled bioavailability of iron from spinach. The data show fresh spinach is a poor source of iron, and emphasize the importance of evaluation of whole meals rather than single food items. The data support the usefulness of the in vitro/Caco-2 cell ferritin bioassay model for prescreening of space flight diets for bioavailable iron.

Evaluation of metallothionein formation as a proxy for zinc absorption in an in vitro digestion/Caco-2 cell culture model.

Caco-2 cell metallothionein (MT) formation was studied to determine if MT could be used as a proxy for zinc (Zn) absorption in a cell culture model. The MT intracellular concentration was determined using a cadmium/hemoglobin affinity assay. The cellular Zn uptake was determined by acid digests (5% HNO(3)) using inductively-coupled argon-plasma emission spectroscopy. The effect of phytic acid (PA) on cellular Zn and MT concentrations was also studied. Cells were treated with a media containing 0, 2, 5, 10, 25, 50, 75 µmol L(-1) Zn (ZnCl(2)). The effect of varying the Zn:PA molar ratios (1:0, 1:1, 1:5, 1:10, 1:20) on the Zn uptake and MT formation was determined. The results showed a positive linear correlation between Zn-media concentrations and cellular Zn uptake, and MT formation was observed. Zn and MT concentrations in the cells treated with increasing levels of Zn (>25 µmol L(-1) Zn) were elevated. The Zn and MT concentrations in the cells incubated with Zn (when <10 µmol L(-1)) were similar to the untreated cells. PA significantly lowered the cellular Zn and MT concentrations. When the Zn:PA molar ratios were >1:5, cellular MT concentrations were no different to untreated cells. When a combined in vitro digestion/cell model was used, the cellular MT concentrations in white or red beans and fish samples were no different to the cell baseline. This study suggests that measurements of cellular Zn and MT concentrations have some limitations (<10 µmol L(-1) Zn). PA was observed to be a potent inhibitor of Zn uptake. Under the conditions of this in vitro model, Caco-2 cell monolayers are not useful for evaluating the Zn availability from foods.