Semisynthetic Meal Research Articles

Comparison between in vitro and in vivo methods to screen iron bioavailability

An enhanced in vitro method to screen iron bioavailability (Fe-BA) was compared with an in vivo human method. Two recipes, previously evaluated in vivo for Fe-BA, were replicated and fed to an in vitro system. The first, a semi-synthetic meal, tested the effect of ascorbic acid on iron absorption. The second, a corn-bean meal, assessed in three experiments, the effect of Na2EDTA plus ferrous fumarate and the effect of FeSO4 and NaFeEDTA fortificants on iron absorption. For the first recipe, the in vitro and in vivo results were highly correlated (r = 0.9765; P = 0.0008). For the second recipe, the in vitro method displayed a similar trend to the in vivo results vis-à-vis ferrous sulfate and NaFeEDTA, but not ferrous fumarate plus Na2EDTA. This method is appropriate for rapid screening of a large number of samples prior to human evaluations; it uses less samples, reagent and supplies than a previous in vitro method.

Reactivity of Chromium(III) Nutritional Supplements in Biological Media: An X-Ray Absorption Spectroscopic Study

Chromium(III) nutritional supplements are widely used due to their purported ability to enhance glucose metabolism, despite growing evidence on low activity and the potential genotoxicity of these compounds. Reactivities of Cr(III) complexes used in nutritional formulations, including [Cr3O(OCOEt)6(OH2)3](+) (A), [Cr(pic)3] (pic=2-pyridinecarboxylato(-) (B), and trans-[CrCl2(OH2)4](+) (CrCl3.6H2O; C), in a range of natural and simulated biological media (artificial digestion systems, blood and its components, cell culture media, and intact L6 rat skeletal muscle cells) were studied by X-ray absorption near-edge structure (XANES) spectroscopy. The XANES spectroscopic data were processed by multiple linear-regression analyses with the use of a library of model Cr(III) compounds, and the results were corroborated by the results of X-ray absorption fine structure spectroscopy and electrospray mass spectrometry. Complexes A and B underwent extensive ligand-exchange reactions under conditions of combined gastric and intestinal digestion (in the presence of a semisynthetic meal, 3 h at 310 K), as well as in blood serum and in a cell culture medium (1-24 h at 310 K), with the formation of Cr(III) complexes with hydroxo and amino acid/protein ligands. Reactions of compounds A-C with cultured muscle cells led to similar ligand-exchange products, with at least part of Cr(III) bound to the surface of the cells. The reactions of B with serum greatly enhanced its propensity to be converted to Cr(VI) by biological oxidants (H2O2 or glucose oxidase system), which is proposed to be a major cause of both the insulin-enhancing activity and toxicity of Cr(III) compounds (Mulyani, I.; Levina, A.; Lay, P. A. Angew. Chem. Int. Ed. 2004, 43, 4504-4507). This finding enhances the current concern over the safety of consumption of large doses of Cr(III) supplements, particularly [Cr(pic)3].

Investigation of an in vitro model for predicting the effect of food components on calcium availability from meals

The availability of calcium from semi-synthetic meals and model food systems was studied by an in vitro method using equilibrium dialysis after simulating gastric digestion. Twelve different food components (phytate, oxalate, wheat fibre- and barley fibre-extract, D-sorbitol, xylitol, galactitol, casein, three different casein phosphopeptide preparations, and lactose) were added to the semi-synthetic meal and their effect on the relative index of availability of 45Ca was studied. Phytate, oxalate, wheat fibre-extract and barley fibre-extract, and, surprisingly, casein had a negative effect on availability of calcium. The most pronounced effect was observed for the addition of phytate. The other dietary factors had no effect on calcium availability. Therefore, this rapid and inexpensive in vitro model could be a useful screening method for factors affecting calcium absorption, especially inhibitory factors, in future human calcium bioavailability studies.

Effect of myofibrillar muscle proteins on the in vitro bioavailability of non-haem iron

It is well established that the bioavailability of non-haem iron from foods is enhanced by the presence of meat. However, the nature of the promoter in meat has not yet been characterised. The present study was designed to compare the effects of the myofibrillar protein fractions on the bioavailability of non-haem iron in an attempt to identify the ‘meat factor’. Rabbit skeletal muscle was fractionated and whole muscle, myofibrillar protein, myosin and actin were isolated. Myosin was subjected to selective proteolysis with chymotrypsin and the heavy meromyosin, light meromyosin, rod region and head region were prepared. Protein fractions (1 g) were incorporated into 100 g semi-synthetic liquid meal and the in vitro dialysability of iron was determined. Egg albumin was used as a reference protein. When compared with egg albumin, all protein fractions significantly enhanced iron dialysability, except for light meromyosin which was inhibitory. Myosin had a greater enhancing effect than actin and, within myosin, the enhancing effect was greatest for the heavy meromyosin fraction. The enhancement appeared to coincide with the known distribution of cysteine residues in the myofibrillar proteins. The presence of the sulphydryl blocking agent, N-ethylmaleimide (NEM), in meals containing myosin reduced iron dialysability in a dose-related manner, but NEM had only a small effect in meals containing actin. Meanwhile, incorporation of cysteine into meals containing actin increased iron dialysability. The present results suggest that the enhancement of non-haem iron dialysability by meat is associated with myosin, in particular, with the heavy meromyosin region. Peptide fractions rich in cysteine residues, probably constitutes the ‘meat factor’.

Effects of pH and chelating agents on iron binding by dietary fiber: implications for iron availability

Iron binding by isolated dietary fiber in the presence of iron ligands was measured. Iron-ligand solutions were mixed with isolated fiber and incubated. Bound and polymerized iron were estimated by centrifugation and ultrafiltration, respectively. Results showed that iron binding varied greatly with ligand used and with pH. Based on these studies, it was predicted that ligands would influence iron availability in the order ascorbate < citrate = nitrolotriacetic acid < EDTA. When added to a complex meal containing beef, bread, green beans, and milk, iron availability (measured by an in vitro method) was little affected by fiber, citrate, and ascorbate, but enhanced by nitrolotriacetic acid and EDTA. When added to a semisynthetic meal, fiber, ascorbate, citrate, nitrolotriacetic acid, and EDTA increased iron availability. It appears that the effect of dietary fiber on iron availability is greatly dependent on the ligands present in the meal.

In Vitro Estimation of Iron Availability in Meals Containing Soy Products

Effects on relative nonheme iron availability of soy protein products incorporated into meals were evaluated. An in vitro method was used to estimate nonheme iron availability. Incorporation of soy isolate into a semisynthetic meal reduced estimated iron availability (dialyzable iron) compared to an egg white control, but this effect was reversed by baking the soy isolate. Ascorbic acid was not as effective in increasing estimated iron availability in soy-containing meals as in an egg white control meal. When 34 individual soy products were substituted for egg white in a semisynthetic meal, iron availability ranged from 3.23 to 0.00% dialyzable iron. Iron availability appears to be inversely related to the protein content of the soy products. Soy isolates, as a group, had lower relative iron availability than the group of soy flours evaluated. The amount of soy product protein incorporated into a standard meal was negatively correlated with relative iron availability. The majority of meals containing soy products had lower estimated iron availabilities than meals that did not contain soy.iron iron availability in vitro iron absorption soy protein

The inhibitory effect of soy products on nonheme iron absorption in man

Radioiron absorption studies were performed in male volunteer subjects to determine the effect on nonheme iron absorption of various semipurified proteins. When egg albumen and casein were substituted in protein-equivalent quantities in a semisynthetic meal, similar mean absorptions of 2.5 and 2.7% were observed. In contrast, isolated soy protein reduced absorption sharply, to an average of 0.5%. When egg albumen in the semisynthetic meal was replaced with full fat soy flour, textured soy flour, and isolated soy protein, absorption fell from 5.5 to 1.0, 1.9, and 0.4%, respectively, indicating an inhibitory effect by a wide range of soy products. The effect of substituting textured soy flour for meat in a meal containing a hamburger, french fries, and a milkshake was also evaluated. With 3:1 and 2.1 ratios of meat to unhydrated textured soy flour, absorption decreased by 61 and 53%, respectively. The soy products tested in this study have a pronounced inhibitory effect on the absorption of nonheme iron.

Food iron absorption in human subjects V. Effects of the major dietary constituents of a semisynthetic meal

Studies were performed in adult volunteer subjects to determine the effect on nonheme iron absorption of protein, carbohydrate, and fat. These constituents were administered as egg albumin, dextrimaltose, and corn oil, respectively, in a semisynthetic meal containing 700 kcal and 4.1 mg iron. Because any one of these ingredients are unpalatable when administered alone, their effect was determined by serially deleting or doubling their content in the basal semisynthetic meal. With both approaches, carbohydrate and fat had little influence whereas egg albumin had a significant inhibitory effect on the absorption of nonheme iron.

Vitamin C, the common cold, and iron absorption

A sizable segment of the population was found to be taking large quantities of vitamin C to reduce the number or severity of upper respiratory infections. To determine the affect of this supplementation on iron balance, multiple radioiron absorption tests were performed in 63 male subjects. The increase in iron absorption from a semisynthetic meal was directly proportional to the amount of ascorbic acid added over a range of 25 to 1,000 mg. The ratio of iron absorption with/without ascorbic acid at these two extremes was 1.65 and 9.57, respectively. The relative increase was substantially less when the test meal contained meat. A large dose of vitamin C taken with breakfast did not effect iron absorption from the noon or evening meal. A telephone survey of 100 individuals revealed that 67 were taking supplemental ascorbic acid in doses ranging as high as 2 g daily. The average intake of supplemental ascrobic acid in this population was 280 mg daily. If taken only with breakfast, this level of supplementation would produce a nearly 2-fold increase in the amount of iron absorbed daily. If taken in divided doses with each mean, the increase in iron absorption would be more than 3-fold.

Food iron absorption in human subjects IV. The effects of calcium and phosphate salts on the absorption of nonheme iron

The influence of physiological levels of calcium and phosphorus on the absorption of nonheme iron from a semisynthetic meal was evaluated. Each of the 34 participating subjects received two to four test meals. In three studies where both calcium and phosphate were added, absorption of nonheme iron was reduced to 27 to 47% of that absorbed when no salts were added. However, with the single addition of either calcium or phosphate to the test meals no significantly inhibiting effect was observed. As the absorption of nonheme iron was significantly reduced only with the combined addition of calcium and phosphate, it is suggested that a calcium-phosphate-iron complex forms which inhibits iron absorption.

Food iron absorption in human subjects. III. Comparison of the effect of animal proteins on nonheme iron absorption

The ability of various animal proteins to enhance the absorption of dietary nonheme iron was evaluated by performing multiple radioiron absorption measurements in 70 volunteer subjects. Protein equivalent substitutions of nine animal foods were made in two basic test meals. The first was a standard meal of high iron availability (mean absorption, 8.3%) containing beef muscle as the animal protein. The second was a semisynthetic meal of low iron availability (mean absorption, 1.4%) containing ovalbumin as the protein source. Two categories of animal protein were defined. Substitution of beef, lamb, pork, liver, fish, and chicken for the egg ovalbumin in the sannisynthetic meal resulted in a significant, 2-fold to 4-fold increase in iron absorption whereas no increase was observed with milk, cheese, or egg. Reciprocal findings were obtained when these foods were substituted for the beef contained in the standard meal. All sources of animal proteins are not equivalent in their effect on nonheme iron absorption.

Food iron absorption in man II. The effect of EDTA on absorption of dietary non-heme iron

One hundred and eighty iron absorption tests were performed in 45 normal men to determine the effect of EDTA on the absorption of dietary non-heme iron. The addition of 50 mg EDTA to test meals containing 4.1 mg iron reduced absorption by approximately one-half from meals of both high (standard meal) and low (semisynthetic meal) iron availability. Studies employing dual radioiron labels demonstrated complete isotopic exchange of ferric EDTA with dietary non-heme iron. Further studies were carried out to determine the decrease in food iron absorption at varying levels of EDTA. At a 1:1 molar ratio of EDTA to iron, absorption of non-heme iron was reduced to 72% and at a 2:1 molar ratio, to 50% of absorption without EDTA. These levels of EDTA are within the range believed to be present in the United States diet.

Food iron absorption. I. Use of semisynthetic diet to study absorption of nonheme iron

Recent studies have established the validity of employing an extrinsic radioiron tag to measure the absorption of nonheme iron from a complex meal. In the present study, extrinsic tagging was used to measure absorption of nonheme iron from a standard meal chosen as representative of a typical American meal, and from a semisynthetic meal having the same total chemical composition. The latter was designed so that the major dietary components could be systematically altered to determine their separate effects on food iron absorption. Absorption from the standard meal in 32 healthy women averaged 10.0% as compared with a mean absorption of 1.8% from semisynthetic meal. Most but not all of this fivefold difference in absorption could be explained by the enhancing effect of meat in the standard meal. The low availability of iron from the semisynthetic meal will be of particular value in studying factors which enhance food iron absorption, whereas the standard meal is suitable for studying factors which reduce availability of food iron.