1. Qassim Univ., College of Medicine The Hemopoietic and Immune Systems Phase II, Year II Iron metabolism Dr. Tarek A. Salem Biochemistry

2. Objectives List the different sources of iron Describe its absorption and the role of vitamin C in this process. Describe how iron is transported and stored in the body Outline the excretion of iron List different biochemical investigations evaluating iron state in blood and iron deficiency anemia. Demonstrate the estimation of iron and total iron binding capacity in serum Mention the principle of estimation of iron and total iron binding capacity in serum Mention the diagnostic importance of serum iron estimation.

3. Iron Iron (Fe) is one of the trace element, M. Wt. 56 May be present as: – Ferrous (2+)reduced – Ferric (3+) oxidised Fe +++ + e -  Fe ++ Redox states allows activity passing electrons around body Redox change required for iron metabolism

4. Distribution of Iron Total body iron of an adult (70 kg) = 3 – 5g Body iron is distributed as follows: Non heme iron compoundsHeme iron compounds -Ferritin (10 -15%) -Hemosiderin (9%) - Transferrin (<1%) - Catalase (<1%) - Hemoglobin (65-80% of body iron) - Myoglobin (10%) - Cytochrome C (<1%) - Peroxidase (<1%) Other iron compounds (<5%) is mainly present bound to proteins.

5. Heme structure Heme

6. Distribution of Iron According to its tissue (location), it is distributed into: 1- Hemoglobin iron. 2- Tissues iron: - Fixed tissues iron: e.g. Myoglobin, cytochrom C, catalase and peroxidase enzyme. - Labile tissues iron: stored iron in the form of ferritin and small amount of hemosidrin. 3- Plasma iron: transport iron (trasferrin).

7. Iron function Oxygen carriers – haemoglobin Oxygen storage – Myoglobin Energy Production – Cytochromes (oxidative phosphorylation)

8. Iron sources Plant sourceAnimal source -Wheat - Date - Molasses -Nuts -Spinach - Meat - Liver - Heart - Kidney - Spleen - Egg yolk - Shellfish

9. Daily requirement Infants: 10-15 mg Children: … 1-3 yrs: 15 mg … 4-10 yrs: 10 mg Older children and adults (Males): …11-18 yrs: 18 mg … After 19 yrs: 10 mg Females: … 11-50 yrs: 15-18 mg During pregnancy or lactation: 20 mg After 51 yrs: 12-15 mg

10. Iron absorption ferrireductase enzyme enterocytes in the proximal duodenum. 1.Iron is present in the diet as Fe 3+ state. It is reduced to Fe 2+ by a ferrireductase enzyme present on the surface of enterocytes in the proximal duodenum. intestinal mucosal 2.The absorbed Fe 2+ iron are oxidized again inside intestinal mucosal cells to Fe 3+, which then, deliver into mitochondria and combine with apoferritin to form ferritin; or to apotransferrin to form transferrin depending upon the iron metabolism in the body. 3.Ferritin is the chief storage form of iron.

11. Factors affecting iron absorption 1.Requirements of body: An increase rate of erythropoiesis (e.g. after hemorrhage) increase the need of iron absorption. 2.Vitamin C: helps reduction of ferric to ferrous facilitating iron absorption. 3.Gastric HCl: helps absorption by formation of soluble ferrous chelates. 4.Some molecules in diet as tannic acid, oxalate and phosphate forming insoluble complexes with iron, inhibiting its absorption.

12. Iron Transport Trasferrin (Tf) is a plasma protein for Iron transport. It is a glycoprotein, synthesized in the liver. It transports iron (2 mol of Fe 3+ ) in the circulation to sites where iron is required. Total iron binding capacity (TIBC): represents the capacity of transferrin to bind iron (300µg of iron/dl). Normally, about 30% (one third) of the available sites on the transferrin molecule are filled.

13. Transferrin Saturation Normal iron Normal transferrin Saturation 30% High iron Low transferrin Saturation 80% Transferrin Iron IRON OVERLOAD NORMAL IRON STATUS

14. Excretion of iron The high binding capacity of iron to macromolecules, leads to absence of free iron salts, thus this metal is not lost via usual excretory routes. This amount represents 1-2 mg/day in stools but also in sweet, hairs and menstrual blood in females. Regulation of the total body iron: the total body iron is regulated at the level of absorption: in iron deficiency the absorbed iron increase while in iron overload, the absorption decreases.

15. Excretion Urine: 0.5-1.5 mg Faeces: 0.3-0.75 mg Bile and sweat: 0.5-1 mg Menstrual flow: 1-3 mg – Faeces - unabsorbed food iron – Pregnancy - iron to the fetus – Sweat, hair loss, nail clippings Iron content regulation of the body By absorption only, not by excretion

16. Different biochemical investigations evaluating iron state Serum iron (65-180 µg/dL men). (50-170 µg/dL women) (100-250 µg/dL baby) (50-120 µg/dL children) Serum transferrin saturation: It is the ratio of serum iron and total-iron binding capacity (measured as %) (15-50% male) (12-45% female) Total iron binding capacity (TIBC) (240-450 µg/dL). Unsaturated iron binding capacity (UIBC): is calculated by substracting the serum iron from TIBC

17. Iron deficiency anemia It may result from: 1.Inadequate intake of iron 2. Lack of iron absorption 3.Excessive blood loss 4.Inhibition of incorporation of iron into hemoglobin (lead poising). Lab investigations: -Serum iron is low -Transferrin is high (liver produce more Tf to maximize use of little iron) -Transferrin saturation is low ©

18. Iron Overload 1- Primary (Hereditary) hemochromatosis: It is results in increase iron absorption in the small intestine. In this case, total body iron exceeds 15 g. The term “hemosidrosis” is generally used to indicate the pathological effect of iron accumulation in any organ, which mainly occurs in the form of hemosidrin. Subcutaneous tissues: giving the bronze coloration. Pancreas: producing diabetes. Brain: causing mental retardation Increase serum iron. Increase TIBC. Decrease in UIBC. Marked increase in serum ferritin

19. Iron Overload 2- Secondary Hemochromatosis: It may result from: -Repeated blood transfusions. -Severe chronic hemolysis. -Increased absorption of iron from intestine. -Excessive administration of injectable iron. -Excessive dietary iron.

20. Estimation of Serum iron Principle: Serum Fe 3+ -transferrin complex is dissociated by addition of an acetic buffer containing hydroxylamine which reduces the Fe 3+ to Fe 2+. Then, the chromogenic agent (PDTS), forms a highly colored Fe 2+ -complex that is measured at 565 nm. The intensity of color is directly proportional to the conc. of iron. Serum Iron (µg/dl) = Reading of Test X Conc. of standard (µg/dl) Reading of standard

21. Estimation of total iron binding capacity (TIBC) Principle: First, unsaturated iron binding capacity (UIBC) is determined by adding Fe 2+ to serum, so that they bind to unsaturated iron binding sites on transferrin. The excess Fe 2+ are reacted with PDTS to form the color complex which is measured. The difference between the amount of Fe 2+ added to the amount of Fe 2+ measured represents unsaturated iron binding capacity.

22. Calculation The intensity of color is directly proportional to the UBIC. Calculate TIBC by adding serum iron to the UBIC © Serum TIBC= Serum iron (µg/dL) + Serum UIBC (µg/dL) UBIC (µg/dl) = Reading of Test X Conc. of standard (µg/dl) Reading of standard