Iron tissue storage and hemoglobin levels of deficient rats repleted with iron bound to the caseinophosphopeptide 1-25 of beta-casein.

Caseinophosphopeptides (CPP) issued from enzyme digestion of caseins bind cations and keep them soluble in the digestive tract. They could be used as ligands to improve iron (Fe) bioavailability. Fe-deficient young rats were repleted with Fe (40 or 200 mg/kg of diet) bound either to the beta-CN (1-25) of beta-casein or to whole beta-casein or as FeSO(4). A control pair-fed group was given 200 mg of Fe (FeSO(4))/kg of diet for 6 weeks. After repletion, hemoglobin concentration of the control group was reached only by the ) animals fed 200 mg of Fe/kg; beta-CN (1-25) bound Fe (40 and 200 mg) produced higher Fe liver and spleen stores than FeSO(4). Binding Fe to the whole, nonhydrolyzed beta-casein gave results intermediate between the other experimental groups. Binding Fe to phosphoserine residues of low molecular weight CPP improved its ability to cure anemia and to restore iron tissue stores, as compared to Fe bound to the whole casein and to inorganic salts.     

Sensomics mapping and identification of the key bitter metabolites in Gouda cheese

Application of a sensomics approach on the water-soluble extract of a matured Gouda cheese including gel permeation chromatography, ultrafiltration, solid phase extraction, preparative RP-HPLC, and HILIC combined with analytical sensory tools enabled the comprehensive mapping of bitter-tasting metabolites. LC-MS-TOF and LC-MS/MS, independent synthesis, and sensory analysis revealed the identification of a total of 16 bitter peptides formed by proteolysis of caseins. Eleven previously unreported bitter peptides were aligned to beta-casein, among which 6 peptides were released from the sequence beta-CN(57-69) of the N terminus of beta-casein and 2 peptides originated from the C-terminal sequence beta-CN(198-206). The other peptides were liberated from miscellaneous regions of beta-casein, namely, beta-CN(22-28), beta-CN(74-86), beta-CN(74-77), and beta-CN(135-138), respectively. Six peptides were found to originate from alpha(s1)-casein and were shown to have the sequences alpha(s1)-CN(11-14), alpha(s1)-CN(56-60), alpha(s1)-CN(70/71-74), alpha(s1)-CN(110/111-114), and alpha(s1)-CN(135-136). Sensory evaluation of the purified, synthesized peptides revealed that 12 of these peptides showed pronounced bitter taste with recognition thresholds between 0.05 and 6.0 mmol/L. Among these peptides, the decapeptide YPFPGPIHNS exhibited a caffeine-like bitter taste quality at the lowest threshold concentration of 0.05 mmol/L.     

Proteolytic activities of suparen and rennilase on buffalo, cow, and goat whole casein and beta-casein.

The proteolytic specificity and activity of Mucor miehei protease (Rennilase) and Endothia parasitica protease (Suparen) on buffalo, cow, and goat whole casein and beta-casein (CN) were studied by analyzing the degradation products. The results suggest that Rennilase hydrolyzes casein of the three species in a manner similar to that of chymosin, resulting in the formation of alpha(s1)-I and beta-I, -II, -III as initial degradation fragments of alpha(s1)- and beta-CN. alpha(s1)-I was also the initial breakdown product of alpha(s1)-CN by Suparen. Contrary to Rennilase, Suparen showed a higher affinity toward beta-CN and hydrolyzes beta-CN, giving rise to degradation products characterized by mobility lower than that of beta-CN. Increasing NaCl concentration (>3%) reduced the proteolysis of beta-CN of the three species by Rennilase but not by Suparen. The hydrolysis of alpha(s1)-CN and alpha(s1)-I by the two enzymes was enhanced in the presence of NaCl.     

Bioavailability of iron from wheat aegilops derivatives selected for high grain iron and protein contents

A coupled in vitro digestion/Caco-2 model was employed to assess iron bioavailability from wheat Aegilops derivatives selected for high iron and protein contents. The iron content in wheat genotypes used in this study correlated to a great extent with both protein (r = 0.80) and phytate (r = 0.68) contents. The iron bioavailability was based on Caco-2 cell ferritin formation from cooked digests of these derivatives (relative to WL711 control) and correlated positively with dialyzable iron (r = 0.63) and total iron content (r = 0.38) but not with the phytate content. The apparently decreased phytate/iron molar ratios, however, correlated negatively (r = -0.42) with the iron bioavailability, justifying the utilization of these parameters in biofortification programs. Iron bioavailability in the derivatives increased up to 1.5-fold, corresponding to a 1.5-2.2-fold increase observed in iron content over control. These data suggest that biofortification for iron proportionately leading to higher iron bioavailability will be the most feasible and cost-effective approach to combat micronutrient deficiency.     

Effect of Liming on the Plant Availability and Distribution of Zinc and Copper among Soil Fractions

Abstract

Information on the redistribution of applied micronutrients into different fractions as a result of lime application is important to predict plant accumulation of nutrients and to select appropriate chemical extraction procedures for evaluation of micronutrient availability. The present work was carried out to study the influence of liming on the availability and redistribution of zinc (Zn) and copper (Cu) among soil fractions. Additionally, the effect of liming was evaluated on the recovery of these micronutrients by different chemical extractants (Mehlich‐1, Mehlich‐3, and diethylenetriaminepentaacetate (DTPA), which were correlated with Zn and Cu concentrations in corn (Zea mays L.) plants and soil fractions (exchangeable, organic matter, amorphous iron oxides, and crystalline iron oxides). The results showed that Zn added to soil samples that did not receive lime was retained mainly in the exchangeable and organic matter fractions. The liming resulted in distribution of Zn into iron oxides and as a result decreased the plant accumulation of Zn. Mehlich‐3 was the most efficient extractant to predict the plant accumulation of Zn in the acid soils, whereas DTPA was the most efficient in the limed soils. The oxide crystalline fraction was the major fraction responsible for retaining Cu in the soils. However, Cu added to soil was distributed mainly into organic matter. Mehlich‐3 was the most suitable extractant for predicting the bioavailability of Cu in limed or unlimed soils.     

Immunochemical evaluation of bovine beta-casein and its 1-28 phosphopeptide in cheese during ripening

Polyclonal antibodies raised against the plasmin-released 1-28 phosphopeptide from bovine beta-casein [i.e., beta-CN(f1-28)4P] specifically recognized the tryptic beta-casein 1-25 and 2-25 peptides, whatever the degree of phosphorylation, but were unresponsive to the shortened beta-casein 16-22 phosphopeptide. These antibodies were able to recognize the parent bovine beta-casein as well as the homologous water buffalo protein, but they could not detect the homologous counterparts from ovine and caprine milks. Such antibodies were used in competitive enzyme-linked immunosorbent assays to monitor the plasmin-mediated release of the 1-28 phosphopeptide from beta-casein and to evaluate the residual native beta-casein in bovine cheese sampled during ripening. Applications of these polyclonal antibodies are suggested mainly for estimating the age of hard cheeses and, possibly, for tracing the presence of bovine casein in fresh ovine and caprine cheeses.     

Relationship between MALDI-TOF analysis of beta-CN f193-209 concentration and sensory evaluation of bitterness intensity of aged cheddar cheese

An internal standard method was previously developed to measure the concentration of a synthetic bitter peptide, beta-CN f193-209, by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. The objective of this study was to evaluate the relationship between beta-CN f193-209 concentration in an aqueous extract of aged Cheddar cheese and bitterness intensity of the cheese. Concentrations of beta-CN f193-209 in cheese extracts were determined by MALDI-TOF at 0, 120, 180, and 270 days. Trained panels evaluated the bitterness intensity of the cheeses at 180 and 270 days. Correlation coefficients between MALDI and sensory data at 180 and 270 days were 0.803 and 0.554, respectively. The decreased correlation may be due to the presence of other bitter peptides more responsible for bitterness at longer aging or the production of compounds that mask bitterness intensity.     

Antioxidant mechanisms of caseinophosphopeptides and casein hydrolysates and their application in ground beef

Caseinophosphopeptides (CPP) and casein hydrolysates have been shown to bind prooxidant metals such as iron, but their effectiveness as metal chelators to inhibit lipid oxidation in foods has still not been fully investigated. Thus, the antioxidant activity of CPP and casein hydrolysates was studied in phosphatidylcholine liposome model systems. CPP (< 1.0 mg/mL) and casein hydrolysates (0.3-1.7 mg/mL) were effective inhibitors of TBARS development when oxidation was promoted by ferric/ascorbate. High amounts of CPP (> 1.0 mg/mL) were prooxidant, whereas casein hydrolysates were observed to be only antioxidative. In the presence of peroxyl radicals, casein hydrolysates were more effective scavengers than enriched CPP (3-15 mM). In cooked ground beef, TBARS formation was inhibited 75, 39, and 17% by 0.5% enriched CPP, casein hydrolysates, and low molecular weight casein hydrolysates, respectively, after 4 days of storage. The results show that CPP and casein hydrolysates are promising sources of natural antioxidants for foods.     

Effect of Iron Source on Color and Appearance of Micronutrient‐Fortified Corn Flour Tortillas

Iron deficiency anemia is a widespread occurrence. Consequently, iron is commonly added in cereal fortification programs. However, many iron sources cause undesirable sensory changes, especially color changes, in the food being fortified. This study evaluated the effect of different iron sources on CIE L*a*b* color values and sensory color perception in fortified corn tortillas. Corn masa flour was fortified with micronutrient premix containing vitamins, zinc, and one of eight iron compounds. Iron sources included ferrous fumarate (F), ferrous sulfate (S), ferric orthophosphate (OP), ferrous lactate (L), ferrous gluconate (G), ferric pyrophosphate (PP), sodium iron (III)‐EDTA, and A‐131 electrolytic iron (E), with addition levels adjusted based on bioavailability. Control (Ct) samples were prepared with all micronutrients except iron. All iron‐fortified tortillas had lower L* values and were significantly darker than control tortillas. Based on instrumental color values and Mexican regulatory recommendations, five treatments were selected for further testing. A difference‐from‐control sensory test was conducted comparing PP, E, OP, F, and S with Ct tortillas. Sensory rankings were C t > E = PP > OP > F > S. A‐131 electrolytic iron is recommended for fortification of corn tortillas due to minimal effect on color and significantly lower cost than other iron sources evaluated.     

Silicon alleviates the toxicity of cadmium and zinc for maize (Zea mays L.) grown on a contaminated soil

Although silicon (Si) is not an essential element, it presents a close relationship with the alleviation of heavy‐metal toxicity to plants. This work was carried out to evaluate the effects of Si application to soil on the amelioration of metal stress to maize grown on a contaminated soil amended with Si (0, 50, 100, 150, and 200 mg kg^–1) as calcium silicate (CaSiO₃). Additionally, the cadmium (Cd) and zinc (Zn) bioavailability as well as their distribution into soil fractions was also studied. The results showed that adding Si to a Cd‐ and Zn‐contaminated soil effectively diminished the metal stress and resulted in biomass increase in comparison to metal‐contaminated soil not treated with Si. This relied on Cd and Zn immobilization in soil rather than on the increase of soil pH driven by calcium silicate application. Silicon altered the Cd and Zn distribution in soil fractions, decreasing the most bioavailable pools and increasing the allocation of metals into more stable fractions such as organic matter and crystalline iron oxides.     

Fermentation and lactic acid addition enhance iron bioavailability of maize

Maize is one of the most important cereal crops for human consumption, yet it is of concern due to its low iron bioavailability. The objective of this study was to determine the effects of processing on iron bioavailability in common maize products and elucidate better processing techniques for enhancing iron bioavailability. Maize products were processed to represent different processing techniques: heating (porridge), fermentation (ogi), nixtamalization (tortillas), and decortication (arepas). Iron and phytate contents were evaluated. Iron bioavailability was assessed using the Caco-2 cell model. Phytate content of maize products was significantly reduced by decortication (25.6%, p = 0.003) and nixtamalization (15%, p = 0.03), and iron content was reduced by decortication (29.1%, p = 0.002). The relative bioavailability (RBA, compared to 100% bioavailability of porridge with FeSO4) of ogi was significantly higher than that of other products when fortified with FeSO4 (p < 0.001) or reduced iron (p < 0.001). Addition of lactic acid (6 mg/g of maize) significantly increased iron solubility and increased bioavailability by about 2-fold (p < 0.01), especially in tortillas. The consumer panel results showed that lactic acid addition does not significantly affect the organoleptic characteristics of tortillas and arepas (p = 0.166 and 0.831, respectively). The results suggest that fermentation, or the addition of small amounts of lactic acid to unfermented maize products, may significantly improve iron bioavailability. Lactic acid addition may be more feasible than the addition of highly bioavailable but expensive fortificants. This approach may be a novel means to increase the iron bioavailability of maize products to reduce the incidence of iron deficiency anemia.     

Peptides isolated from in vitro digests of milk enhance iron uptake by caco-2 cells

Milk proteins, during digestion, produce a range of biologically active peptides. Among those are peptides that may enhance iron absorption. The objective of this project was to investigate the effect of isolated milk peptides on iron uptake. Cow's milk, 0% fat, was subjected to a modified in vitro digestion process. The milk digest was further fractionated by gel filtration. All eluted fractions as well as beta-casein synthetic peptides (a tripeptide and a hexapeptide) were subsequently tested for effects on iron uptake with Caco-2 cell monolayers. Fractions of milk digests obtained through Sephadex G-25 gel filtration had a significant enhancing effect on iron uptake in Caco-2 cells compared to nonfractionated milk digests. Two fractions (P = 0) and the hexapeptide (P < 0.0001) enhanced iron uptake by up to 3-fold, whereas others and the tripeptide had no effect. These results suggest that selected peptides produced during the in vitro digestion of milk may enhance iron absorption; however, it remains to be demonstrated whether this effect may be nutritionally significant.     

Iron bioavailability of hemoglobin from soy root nodules using a Caco-2 cell culture model

Heme iron has been identified in many plant sources-most commonly in the root nodules of leguminous plants, such as soy. Our objective was to test the effectiveness of soy root nodule (SRN) and purified soy hemoglobin (LHb) in improving iron bioavailability using an in vitro Caco-2 cell model, with ferritin response as the bioavailability index. We assessed bioavailability of iron from LHb (either partially purified (LHbA) or purified (LHbD)) with and without food matrix and compared it with that from bovine hemoglobin (BHb), ferrous sulfate (FeSO4), or SRN. Bioavailability of each treatment was normalized to 100% of the FeSO4 treatment. When iron sources were tested alone (100 ug iron/mL), ferritin synthesis by LHbD and BHb were 19% (P > 0.05) and 113% (P < 0.001) higher than FeSO4, respectively. However, when iron sources were used for fortification of maize tortillas (50 ppm), LHbA and BHb showed similar bioavailability, being 27% (P < 0.05) and 33% (P < 0.05) higher than FeSO4. Heat treatment had no effect on heme iron but had a significant reduction on FeSO4 bioavailability. Adding heme (LHbA) iron with nonheme (FeSO4) had no enhancement on nonheme iron absorption. Our data suggest that heme iron from plant sources may be a novel value-added product that can provide highly bioavailable iron as a food fortificant.     

Genetic modification of low phytic acid 1-1 maize to enhance iron content and bioavailability

High phytate content in staple food crops is a major barrier to successful iron biofortification. We have exploited the low phytic acid 1-1 (lpa1-1) mutant of maize to generate transgenic plants with up-to 70 μg/g seed iron through the endosperm-specific overexpression of soybean ferritin, resulting in more than 2-fold improvement in iron bioavailability. The levels of bioavailable seed iron achieved in this study greatly exceed any achieved thus far and closely approach values estimated to have a nutritional impact on target populations. Gene expression studies reveal a large induction of the YS1 transporter in leaves and severe repression of an iron acquisition gene DMAS1 in roots, suggesting significant alterations in the iron homeostatic mechanisms in transgenic lpa1-1. Furthermore, preliminary tests show that the high-iron lpa1-1 seeds have higher germination rates and seedling vigor when compared to those of the nontransgenic seeds, which may help improve their value to plant breeders.     

Analysis of iron chelates in commercial iron fertilizers by gel chromatography

A gel chromatographic method for the quality control of iron chelate fertilizers is described. The iron chelates are separated on a column of Sephadex G-10 and the eluates are analysed for iron. Using a sample quantity of 25 mg in a volume of 5 ml water and eluting with 0.15 M soidum chloride solution, a separation was achieved of commercial products of Fe-EDDHA or Fe-EDDHMA. The chromatographic analysis of Fe-EDTA or Fe-DTPA calls for a better resolution. This was obtained by decreasing the sample quantity and eluting with 0.035 M calcium chloride solution of pH 7.0. In this way it is possible to eliminate the interference of iron chelates of moderate stability which can be present in commercial products containing Fe-EDTA or Fe-DTPA.     

Quantitative studies and taste re-engineering experiments toward the decoding of the nonvolatile sensometabolome of Gouda cheese

The first comprehensive quantitative determination of 49 putative taste-active metabolites and mineral salts in 4- and 44-week-ripened Gouda cheese, respectively, has been performed; the ranking of these compounds in their sensory impact based on dose-over-threshold (DoT) factors, followed by the confirmation of their sensory relevance by taste reconstruction and omission experiments enabled the decoding of the nonvolatile sensometabolome of Gouda cheese. The bitterness of the cheese matured for 44 weeks was found to be induced by CaCl2 and MgCl2, as well as various bitter-tasting free amino acids, whereas bitter peptides were found to influence more the bitterness quality rather than the bitter intensity of the cheese. The DoT factors determined for the individual bitter peptides gave strong evidence that their sensory contribution is mainly due to the decapeptide YPFPGPIHNS and the nonapeptides YPFPGPIPN and YPFPGPIHN, assigned to the casein sequences beta-CN(60-69) and beta-CN(60-68), respectively, as well as the tetrapeptide LPQE released from alphas1-CN(11-14). Lactic acid and hydrogen phosphate were identified to play the key role for the sourness of Gouda cheese, whereas umami taste was found to be due to monosodium L-glutamate and sodium lactate. Moreover, saltiness was induced by sodium chloride and sodium phosphate and was demonstrated to be significantly enhanced by L-arginine.     

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.     

Iron and zinc bioavailability in rats fed intrinsically labeled bean and bean-rice infant weaning food products

Beans are the core of the Latin American diet and contain iron and zinc. However, the bioavailability of these trace minerals from beans is low. The objective of this study was to determine if the bioavailability of iron and zinc could be improved with the use of fermentation and germination processing technologies. Black beans native to Costa Rica were grown hydroponically with either radioactive iron or zinc. The influence of fermentation and germination on iron and zinc bioavailability from intrinsically labeled infant weaning food products based on black beans and beans-rice was determined in rats. Mineral bioavailability was determined using whole-body (59)Fe retention for iron, and whole-body (65)Zn retention and incorporation of radiolabel into bone for zinc. Percent absorption of (59)Fe from fermented products ranged between 48.0 and 58.0. Percent absorption of (65)Zn ranged from 57.0 to 64.0. Fermentation did not increase iron bioavailability in rats fed fermented beans without rice. Fermentation of cooked beans significantly increased zinc retention. Germination significantly enhanced iron retention from cooked beans from 46 to 55% and from cooked beans-cooked rice from 34 to 48%. Germination significantly improved zinc absorption and retention from cooked beans without added rice.     

Iron bioavailability in fortified fruit beverages using ferritin synthesis by Caco-2 cells

The bioavailability of iron from fortified fruit beverages was estimated by an in vitro system including enzymatic digestion, iron uptake by Caco-2 cells, and ferritin formation determined via an enzyme immunoassay (ELISA). Thus, the aim of the present study was to assess iron bioavailability as influenced by the presence of known dietary promoter and inhibitory factors in fortified fruit beverages containing iron and/or zinc and/or skimmed milk. No negative effect ( p > 0.05) derived from micronutrient interaction can be ascribed to zinc supplementation on iron availability. Besides, the presence of caseinophosphopeptides derived from casein hydrolysis during digestion may confer enhancing effects on iron absorption in samples with milk added with respect to nonadded samples ( p < 0.05). Therefore, from a nutritional point of view, individuals in need of optimal iron absorption may choose dairy samples to ensure optimal iron bioavailability.