Absorption Of Hemoglobin Iron Research Articles

The Journal of clinical Investigation, Volume 41, 1962: iron absorption. IV. The absorption of hemoglobin iron.

The absorption of radioiron in rabbit hemoglobin, hemin, and ferritin has been compared to that of ferrous salt in healthy volunteers and in subjects with iron-deficiency anemia. At a dosage level of 5 mg elemental iron, hemoglobin iron was as well or better absorbed than ferrous salts in the normal subject. Absorption of hemoglobin iron increased less, however, in the iron-depleted or iron-deficient subject. In contrast to the absorption of ferrous salts, that of hemoglobin iron was not decreased by food or by phytate nor increased by ascorbic acid. The absorption of hemin iron was also not decreased by food. Iron absorbed from hemoglobin appeared in the plasma later than that from ferrous salts, but was found to be similarly dialyzable at acid pH with EDTA. These findings suggest that iron in heme complexes is absorbed as a porphyrin complex without conversion to the free ionized form. It is further apparent that there is less effective mucosal regulation of absorption of iron in this form. Finally, the present hypothesis of iron absorption based on the behavior of iron salts is not adequate for all types of food iron.

The effect of cysteine and 2,4-dinitrophenol on heme and nonheme absorption in a rat intestinal model

Previous studies have showed that purified heme iron forms insoluble polymers that are poorly absorbed. The presence of peptides and of amino acids maintaining heme iron in a soluble form could improve its bioavailability. The digestive uptake and transfer of a concentrated hydrolysate of heme peptides (HPH) and of iron gluconate (Gluc) at 100 μM were compared in vitro in a Ussing chamber. The effects of an enhancing amino acid (L-cysteine) on the uptake and transfer of both forms were assessed. An inhibitor of the oxidative phosphorylation (2,4-dinitrophenol; DNP) was used to differentiate the active and passive mechanisms of the absorption. The mucosal uptake (%Tot) and enterocyte transfer (%S) of the two sources of iron did not differ. DNP significantly reduced %Tot and %S of both forms. Cysteine significantly enhanced %Tot and %S of HPH and Gluc, partly corrected the inhibition exerted by DNP on %Tot of HPH and %S of both forms, and fully restored %Tot of Gluc. In presence of peptides produced by globin hydrolysis, the absorption of hemoglobin iron was efficient; it was mostly energy dependent and, therefore, should have occurred by a regulated transcellular pathway. Cysteine enhanced the passive uptake of iron and the passive processes involved in the enterocyte transfer of the common pool made of both sources (heme and nonheme) of iron. These results showed that heme iron can be purified and concentrated without impairing its digestive absorption, provided it remains in presence of peptides or amino acids.

The cadmium effect on iron absorption

Test solutions of cadmium and labeled iron salts, soluble complexes of diferric transferrin, or hemoglobin iron were introduced orally or were injected into tied-off jejunal segments in rat. Cadmium reduced the absorption of iron salts to about half in both normal and iron-deficient rats. Hemoglobin iron absorption was enhanced, indicating that the processing of this form or iron and its release from mucosa to blood was intact. A greater reduction in iron absorption occurred in iron-deficient rats when transferrin iron was injected into gut loops. Mucosal radioiron content in animals given cadmium with either iron salts or transferrin iron was increased. The primary effect of cadmium was on intracellular processing of iron salts and transferrin iron. The major portion of cadmium taken up by the mucosa of normal animals was bound to ferritin, and the effect of cadmium within the mucosal cell may be reduced thus.

Ultrastructural cytochemistry and radioautography of hemoglobin—iron absorption

The subcellular route of hemoglobin-iron absorption by canine intestinal epithelial cells was investigated with cytochemical and radioautographic methods and compared to that observed for inorganic iron absorption. A solution of rabbit [ 59Fe]hemoglobin or an inorganic-iron solution was injected into closed duodenal loops of beagle dogs and mucosal biopsies were obtained 15, 60, and 120 min thereafter. Mucosal retention of radioactivity in dogs given hemoglobin iron was 6.9–10.2% after 120 min, and portal venous blood radioactivity was confined to the inorganic iron fraction. In the same animals diaminobenzidine (DAB)-reactive heme was visualized in microendocytic caveolae at the bases of microvilli and in membrane-bound tubulovesicular structures and granules of the apical cytoplasm. DAB-reactive heme was not identified in the lateral intercellular space or the basal extracellular space. In the same specimens, acid ferrocyanide stained ferric iron in a few apical cytoplasmic vesicles as well as along the outer lateral and basal plasmalemma but failed to identify inorganic iron in microvilli. Both the microvilli and the lateral plasmalemma appeared stained in specimens from animals given inorganic iron. In radioautographic specimens of animals given [ 59Fe]hemoglobin, silver grains were most frequently associated with the apical cytoplasm and the lateral plasmalemma. We conclude that at least one mechanism of hemoglobin-iron absorption involves endocytosis and degradation of hemoglobin in membrane-bound organelles with subsequent conversion of its iron to an inorganic form, which is then transported to the lateral intercellular space where subsequent processing is similar to that observed for absorbed inorganic iron.

Chapter II: Hemoglobin fortification of food. Absorption of hemoglobin and non‐heme iron.

AbstractA simplified dose response study showed no significant reduction of absorption when the dose was increased from 0.9 to 4.2 mg hemoglobin iron, where absorption was about 20 per cent, but a significant reduction to 11 per cent when dose was increased from 4.2 to 19 mg hemoglobin iron. Nor was any significant absorption inhibition found when 3.5 mg hemoglobin iron were given together with an oatmeal porridge test meal. In contrast, this meal reduced non‐heme, ferrous iron absorption from 16.7 to 5.5 per cent.When given on a fasting stomach, no significant difference was found between absorption of hemoglobin and non‐heme iron at 3.5 mg dose levels. Numerically, hemoglobin iron was slightly superior.Meat products, expected to facilitate absorption of non‐heme iron were also studied, either given alone or together with a test meal. Mean absorptions of 3.5‐10 mg hemoglobin iron in different meals varied between 14.7 and 18.9 per cent, as compared to mean absorptions of non‐heme iron between 5.5 and 7.9 per cent.In contrast, hemoglobin iron absorption was reduced to only 0.8 to 7.6 per cent when baked products, such as bread or liver‐paté were studied. Even so, however, mean hemoglobin iron absorptions were 2 to 3 times higher than those of non‐heme iron.The cause of the absorption inhibition in bread was studied. No correlation could be found to the content of bran. No significant inhibition was found when unbaked, sour leaven dough was eaten. However, when hemoglobin was heated to baking temperature for baking times, absorption was reduced as in bread. Hemoglobin breakdown during prolonged heating is thus suggested.No significant difference was found between hemoglobin capsules and hemoglobin tablets.The present results suggest that hemoglobin fortification can contribute substantial amounts of absorbed iron. A single meal containing about 5 mg hemoglobin iron seems to provide about 0.8 mg of absorbed iron. This alone corresponds to the total iron requirements of a woman with a loss of 30 ml blood per menstruation.

Absorption of hemoglobin and hemoglobin iron in alcohol-induced liver injury.

A major source of iron in man’s diet is found in the form of hemoglobin. Previous investigations of the absorption of hemoglobin iron and of hemoglobin itself were performed with radio-labelled, pure hemoglobin. There was evidence of the absorption of hemoglobin iron as well as heme, but not of hemoglobin. As hemoglobin will be ingested together with other proteins, it was decided to use unlabelled hemoglobin prepared from crude red cell sediment without chromatography or other purification. By means of polyacrylamide-gel disc electrophoresis, immunoelectrophoresis, immunofluorescence and gel filtration, it was shown that, in rats treated with ethanol, hemoglobin can be absorbed and transported to the liver as a hemoglobin-haptoglobin complex.

Effect of Phenobarbital on the Absorption of Inorganic and Hemoglobin Iron in the Rat

Previous studies have suggested that iron mucosal protein complexes, possibly related to a carrier substance, are involved in intestinal iron transport. Since there is evidence indicating that phenobarbital induces an increase in intestinal microsomal proteins and enzymes, it was postulated that phenobarbital might affect iron absorption by increasing the synthesis of a carrier protein for iron. This investigation describes studies carried out in the rat to determine the effect of phenobarbital on the absorption of inorganic and hemoglobin iron labeled with Fe 59 . In vivo the absorption of Fe 59 SO 4 was significantly increased in phenobarbital-treated rats compared to controls (20.4 ± 5.7 versus 9.8 ± 4.2%; P P P 0.001). In vitro, phenobarbital treatment resulted in a significant increase in mucosal transfer of Fe 59 SO 4 by everted duodenal gut sacs compared to controls (51.3 ± 8.8. versus 28.0 ± 5.9 mμmoles; P 59 SO 4 by jejunal gut sacs. It is postulated that the phenobarbital-mediated increase in absorption of inorganic iron is related to increased synthesis of a carrier substance that influences iron transport. Further, it is suggested that the increased absorption of hemoglobin iron resulting from phenobarbital treatment is due, at least in part, to increased mucosal heme-splitting enzyme acitivity.

59Fe 2+- and hemoglobin- 59Fe absorption in human beings do not require gastric juice or intrinsic factor

The intestinal 59Fe absorption from 10-μmole (=0.56 mg) 59Fe 2+ and hemoglobin- 59Fe as well as from 5.0-mg hemoglobin- 59Fe doses was measured with the 59Fe-absorption whole-body retention test in normal volunteers and in vitamin-B 12-treated pernicious anemia patients with gastric atrophy and absolute intrinsic-factor deficiency. A completely normal 59Fe absorption from ferrous iron and hemoglobin iron at the 0.56-mg iron level was observed in all the patients without gastric juice. At the 5.0-mg iron dose level hemoglobin- 59Fe was even better (2-fold) absorbed in patients with absolute intrinsic-factor deficiency and a purified hog intrinsic-factor concentrate of known potency did not influence the already doubled hemoglobin- 59Fe absorption. Depleted iron stores in prelatent iron deficiency resulted in a 4-fold increase of 59Fe 2+ absorption and a 50% increase of hemoglobin- 59Fe absorption also in patients without gastric juice and without intrinsic factor. Gastric juice or a gastric intrinsic factor are therefore not required for intestinal ferrous iron or hemoglobin-iron absorption in humans.

Intestinal Absorption of Hemoglobin Iron

The enhanced absorption of hemoglobin iron in iron deficiency was studied in iron-deficient and iron-loaded rats. Both groups were given 59Fe-labeled hemoglobin by gastric intubation and the radioactivity of each rat was measured in a whole body counter at 3 hr and at 9 days. In addition, 59Fe-labeled hemoglobin was instilled into closed duodenal loops of iron-deficient and iron-loaded rats. Three hours later radioactivity was measured in the duodenal washings and in the carcass and duodenum separately using a whole body counter. Radioactivity of hemoglobin and of heme in the washings were measured after separation by charcoal adsorption. Heme and radioiron split from heme were measured separately in homogenates of duodenum using a nonheme iron extraction method. The results indicated that: (1) iron-deficient rats absorb up to 20 times more iron from hemoglobin than do iron-loaded rats; (2) deferioxamine did not influence absorption of hemoglobin iron, indicating that iron split from hemoglobin intraluminally does not contribute significantly to absorption; (3) during the 3-hr closed loop study intraluminal splitting of heme from hemoglobin was the same in both groups, 2 times more heme entered iron deficient mucosa compared with the iron loaded, and, most importantly, 3 times as much iron was transported to carcass by iron deficient mucosa; (4) little, if any, heme was transported as such to the carcass. These results suggest a mucosal mechanism for absorption of hemoglobin iron as well as ionic iron and that the mechanism is enhanced in iron deficiency. The final step in absorption of hemoglobin iron, namely the transport of Fe++ from mucosal cell to plasma, may be rate limiting as it is for absorption of elemental iron. This suggests a common site for regulation of absorption of both elemental and hemoglobin iron.

Absorption of Hemoglobin Iron: The Role of Xanthine Oxidase in the Intestinal Heme-Splitting Reaction

AbstractHeme from ingested hemoglobin—59Fe is taken into the epithelial cell of the small intestinal mucosa of the dog and the 59Fe subsequently appears in the plasma bound to transferrin. A substance was demonstrated in homogenates of the mucosa which releases iron from a hemoglobin substrate in vitro. Thus: (1) The addition of catalase to the mucosal homogenate reduces the "heme-splitting" reaction. In contrast, sodium azide, a catalase inhibitor, potentiates the reaction. This suggests that a peroxide generating system participates in the "heme-splitting" reaction. (2) Xanthine oxidase, an enzyme present in the intestinal epithelial cell, produces H2O2 by oxidation of its substrate. The addition of allopurinol, a xanthine oxidase inhibitor, to the intestinal mucosal homogenate diminishes the "heme-splitting" reaction. (3) Fractionation of the 50,000 Gm. supernatant of the mucosal homogenate on a G-200 Sephadex column shows the "heme-splitting" activity to have the same elution volume as xanthine oxidase, indicating a similar molecular weight. (4) The addition of a mucosal homogenate to a xanthine substrate results in the production of uric acid. These data suggest that xanthine oxidase in the intestinal epithelial cell is important in the release of iron from absorbed heme. The enzyme mediates the "heme-splitting" reaction by the generation of peroxides which, in turn, oxidize the alpha-methene bridge of the heme ring releasing iron and forming biliverdin.

Malabsorption of hemoglobin iron in pernicious anemia: correction with intrinsic factor—containing substances

Hemoglobin iron absorption in patients with treated prenicious anemia (PA) and concomitant iron deficiency was low compared to absorption in patients with iron deficiency alone. Crude and purified hog intrinsic factor (IF) concentrates doubled the absorption of hemoglobin iron in these patients as did normal (neutralized depepsinized) human gastric juice. Hemoglobin iron absorption was not significantly enhanced by PA gastric juice. Absorption of heme iron, like that of hemoglobin iron, was enhanced by normal neutralized depepsinized gastric juice. No enhancement of hemoglobin iron absorption by these substances was obtained in the normal or iron-deficient non-PA control subjects. Preincubation of the hog IF concentrate with antisera to IF significantly reduced the enhancement of hemoglobin iron absorption due to the concentrate. In vitro studies suggest that heme complexes with a substance present in IF-containing materials. Whether a gastric glycoprotein similar to IF serves as an intestinal transport factor for heme, similar to transport of vitamin B(12), or whether normal gastric juice acts by another mechanism cannot be determined at this time.

Absorption of hemoglobin iron: the role of a heme-splitting substance in the intestinal mucosa.

The dog behaves like man in his ability to utilize dietary hemoglobin iron and, therefore, is an excellent model in which to study the mechanisms of absorption. Heme is taken up intact into the epithelial cell of the small intestine but the iron appears in the plasma in a nonheme form. A substance is present in mucosal homogenates which is capable of releasing iron from a hemoglobin substrate in vitro. This has a molecular weight greater than 64,000, and appears to behave as an enzyme. There is no difference in the in vitro, effective concentration of the hemesplitting substance in the mucosa of iron-loaded and iron-deficient dogs to explain in vivo changes in iron absorption. However, the rate at which the heme-splitting substance works in vivo appears to be increased by the removal of the nonheme-iron-end product from the epithelial cell to the plasma. Reduction of the heme-iron content within the epithelial cell may then enhance uptake from the lumen. These studies suggest that the labile nonheme-iron content of the intestinal epithelial cell determines its ability to accept heme as well as ionized iron from the lumen.

Absorption of hemoglobin iron.

Absorption of hemoglobin iron.

Absorption of hemoglobin iron by the rat.

SummaryIn the rat the absorption of a test dose of hemoglobin or hemin iron59 was significantly less than a comparable dose of iron59 as FeSO4. That this is of physiologic significance was demonstrated by producing a state of iron deficiency in animals raised on a diet with hemoglobin as the only source of iron, These findings are in contrast to recent observations in man and suggest a difference between species as well as a difference in the epithelial cells”ability to sequester iron as the salt or in the heme ring.

ROLE OF HYDROCHLORIC ACID IN IRON ABSORPTION.

The absorption of 5 mg iron administered in 300 ml water was compared with absorption from 300 ml 0.05 n HCL in 24 subjects with histamine-fast achlorhydria, using a double isotope technique. In ten individuals given ferric chloride the mean absorptions were 2.2% with water and 9.2% in acid; the difference was less marked in the case of the seven subjects to whom ferrous ascorbate was administered (10.9% and 19.3%, respectively). In contrast, acid did not increase the absorption of hemoglobin iron, the mean figures being 13.4% with water and 9.6% with HCL. pernicious anemia; achlorhydria; gastric pH Fe55 and Fe59; radioactive hemoglobin iron Submitted on June 17, 1963

Iron absorption. IV. The absorption of hemoglobin iron.

The absorption of radioiron in rabbit hemoglobin, hemin, and ferritin has been compared to that of ferrous salt in healthy volunteers and in subjects with iron-deficiency anemia. At a dosage level of 5 mg elemental iron, hemoglobin iron was as well or better absorbed than ferrous salts in the normal subject. Absorption of hemoglobin iron increased less, however, in the iron-depleted or iron-deficient subject. In contrast to the absorption of ferrous salts, that of hemoglobin iron was not decreased by food or by phytate nor increased by ascorbic acid. The absorption of hemin iron was also not decreased by food. Iron absorbed from hemoglobin appeared in the plasma later than that from ferrous salts, but was found to be similarly dialyzable at acid pH with EDTA. These findings suggest that iron in heme complexes is absorbed as a porphyrin complex without conversion to the free ionized form. It is further apparent that there is less effective mucosal regulation of absorption of iron in this form. Finally, the present hypothesis of iron absorption based on the behavior of iron salts is not adequate for all types of food iron.