The subject dealt with in this thesis, is connected with the fact that the iron content of the human diet in the Western countries is thought to be low. Iron retention in man (and in the rat) is determined not only by the iron levels in the diet, but also by the food composition. Different foods or their components (egg, buttermilk, milk, bread, coconut fat, maize oil and beef fat) were studied in vitro and in vivo under comparable conditions to examine whether they had an inhibiting effect on the iron retention from the gut.
The definitions of iron retention and iron absorption are given on page 10, while different factors influencing iron retention are set out in an extensive literature survey.
To examine in vitro whether the foodstuffs bound ferrous or ferric ions, a weak solution of radioactive ferrous or ferric citrate was brought into contact with each of them in a seated bag of semi permeable material. Unlike large molecules (M>12,000) and hydrophobe fats, small molecules like ferrous and ferric ions are not stopped by this material. The radioactivity measured outside the enclosed space was used to determine the fixative capacity for ferrous or ferric ions of the particular food. By using radioactive iron it was possible to measure quickly and with great accuracy very small amounts of ferrous or ferric ions. The basic idea behind the experiments made in vitro was that a strong fixation between iron and the substrate could impede the iron retention.
Both long and short trials were done on animals. The long trials were used to see whether the food composition, without changing the iron content of the diet, affected the iron status of the rats. The iron status of four groups of rats was ascertained by taking 59
Fe iron retention measurements at the beginning and end of a six-month period. After concluding the experiments, the iron content of liver, spleen, heart, kidney, pancreas and duodenum were determined. Both these groups of results were used to assess the effect of the several additives to the rat diets. This data could be collected about the long term effect of the foodstuffs and the physiological response of the animals to the imposed feed situation.
To determine the direct effect of the different foodstuffs on iron retention, a specific food and a radioactive ferric citrate solution were introduced into the stomach of the rat. The cannula with the food was followed immediately by the cannula with the radioactive solution. The iron retention was determined by measuring the animal radioactivity immediately after the insertion of the two stomach tubes and again after two weeks. The period between the measurements was long enough to allow the non-retained iron to leave the body. Radioactivity of the rats was always measured with a whole body counter.
It was hoped to complete the project by using human volunteers, but because radioactive tracers would have been used, the idea was cancelled (see page 5). With the exception of egg, there was little correlation between experiments made in vitro and in vivo. In vitro, egg fixed ferric ions; in vivo, egg added to the diet, reduced iron levels in the rat. In the short trial, the use of both raw and hard boiled egg clearly inhibited iron retention.
In vitro, milk proteins did not fix ferrous or ferric ions. Trials in vivo showed that after 6-7 months on the same solid diet, bodies of rats drinking milk contained less iron than those of rats drinking water. It was found, though less obviously than with egg, that milk also had a directly diminishing effect on iron retention.
When 20 % dried whole meal bread was added to the control diet, no effect on the amount of body iron was seen. No distinction was observed between the animals eating the diets with additions of ferric citrate and hemoglobin. All diets contained 30 ppm iron. The conclusion was that under the experimental conditions the rats utilized ferric citrate or hemoglobin in a similar way.
Rats eating the low-fat control diet with the addition of 20% coconut fat, maize oil or beef fat had a specific distribution of stored iron over the liver and the spleen. Also the type of fat affected the iron levels of the rats. The fats used differed in the proportions of the fatty acids. In coconut fat at least 75 % of the fatty acid was saturated with 14 or less carbon atoms in the molecule. In maize oil more than 80 % of the fatty acid was monoenoic, dienoic or trienoic and in beef fat half the fatty acid was saturated with 16 or 18 carbon atoms in the molecule. In contrast to the control, the addition of 20 % maize oil to the diet lowered iron levels in the rats, while beef fat raised them. Coconut fat was intermediate. The experiments in vitro did not fix ferrous or ferric ions. In the short experiments coconut fat, maize oil or beef fat had no noticeable effect on iron retention.