1. 1- Porphyrins are cyclic compounds formed by the linkage of 4 pyrrole rings through methenyl bridges (-CH=). 2- Porphyrins differ from each other in the type and arrangement of side chains attached to numbered positions of pyrrole rings: 1,2,3,4,5,6,7, and 8. 3- The side chains which stituted in numbered may be substituted in numbered positions are:  A = Acetate = CH 2 -COOH  P = Propionate = -CH 2 -CH 2 -COOH  M = Methyl = -CH 3  V = Vinyl = -CH=CH 2 4- Structure of heme: a- Heme is a complex of iron (in ferrous state, F ++) and one of porphyrins called: protoporphyrin IX. b- Iron is held in the center of heme molecule by bonds to the 4 nitrogen atoms of pyrrole rings of protoporphyrin.

3. 5- Hemoproteins : a- These are a group of specialized proteins that contain heme as a prosthetic group. They are: 1) Hemoglobin.2) Myoglobin. 3) Respiratory cytochromes. 4) Cytochrome p450. 5) Catalase and peroxidase.6) Tryptophan oxygenase. b- The role of the heme group in each protein is: 1) In hemoglobin and myoglobin: Acts as oxygen carrier. 2) In cytochromes : Acts as an electron carrier. 3) In catalase and peroxidase: Acts as a part of active site of the enzyme that catalyzes the breakdown of hydrogen peroxide (H 2 O 2 ). B- Heme Synthesis: 1- The 2 starting materials are succinyl CoA (derived from citric acid cycle in mitochondria) and glycine. The reactions need ALA synthase enzyme and pyridoxal phosphate as activator for glycine. 2- Then ALA passes from mitochondria to cytoplasm where 2 molecules are condensed to form porphobilinogen : 3- Porphobilinogen is a pyrrole ring to which a propionate (-CH 2 -CH 2 -COOH), and acetate (-CH 2 -COOH) groups are attached. 4 Porphobilinogens are condensa ted to give protoporphyrin as follows:

5. Heme is the prosthetic group of hemoglobin, myoglobin, & cytochromes. Heme is an asymmetric molecule. E.g., note the positions of methyl side chains around the ring system.

15. 4- Then heme formation is completed by incorporation (Fe ++ ) into protoporphyrin as follows:  Comments on heme synthesis: a- Uroporphyrinogen decarboxylase present in cytosol removes carbon dioxide (CO 2 ) from 4 acetate groups (A), converting them into methyl groups (M). b- Coproporphyrinogen oxidase present in mitochondria catalyzes the decarboxylation, oxidation and dehydration of propionate groups (p) of ring I and II converting them into vinyl groups (V): 5- Regulation of heme synthesis: Delta aminolevulinic acid synthase enzyme (ALA synthase)is the key enzyme in heme synthesis. a- It is inhibited by :  Heme itself, by feedback inhibition.  Glucose and steroids. b- It is stimulated by :  Certain drugs as phenobarbital and iron.

16. C- Porphyrias : 1- These are a group of diseases resulting from a deficiency of one of the enzymes needed for heme synthesis. 2- Porphyrias lead to disturbance. in heme synthesis and causes: a- Anaemia: due to decrease-production of heme. b- Abdominal pain and neuropsychatric symptoms due to toxic ­effect of the accumulated porphyrin intermediates. c- Photosensitivity: Some porphyrin derivatives when exposed to light react with molecular oxygen to form oxygen radicals, which cause skin damage. 3- Porphyrias are either hereditary or aquired (caused by environmental poisons as lead). 4- All hereditary porphyrias are autosomal dominant except congenital erythropoietic porphyria which is autosomal recessive. 5- Classification: a- Liver and bone marrow are the organs where heme synthesis occurs. b- According to the site of enzyme deficiency, porphyrias can be classified into hepatic (liver), erythropoietic (bone marrow) and erythrohepatic (liver and bone marrow).

17. c- The following table gives summary of the major findings of porphyrias : Hepatic porphyrias:  Acute intermittent porphyria  Porphyria cutanea tarda  Hereditary copro­ porphyria  Variegate porphyria Uroporphyrinogen I synthase Uroporphyrinogen decarboxylase Coproporphyrin oxidase Protoporphyrinogen oxidase Abdominal pain Neuropsychatric. Photosensitivity Abdominal pain Neuropsychatric Photosensitivity Abdominal pain Neuropsychatric Photosensitivity Erythropoietic porphyrias :  Congenital eryth­ropoietic porphy­ria Uroporphyrinogen III synthase Photosensitivity Erythrohepatic porphyrias:  Protoporphyria FerrochelatasePhotosensitivity

18. A- Hemoglobin is found only in the red blood cells. B- Its main function is to transport oxygen from the lungs to the capillaries of the tissues and carbon dioxide (CO 2 ) from the tissues to the lungs. C- Structure of hemoglobin: 1- Hemoglobin is conjugated protein which consists of specialized protein called: globin that is tightly bound to 4 heme molecules. 2- Globin is a protein with four peptide chains joined together by noncovalent bonds (tetramer). 3- Several different kinds of hemoglobin are normally found in human. They vary in the primary structure of the peptide chains of globin. These are HbA, HbA 2, HbF, HbA 1. a- Hemoglobin A : 1) It is the major hemoglobin in adults (97%). 2) Its globin comprises 4 polypeptide chains:  Two α - chains (141 amino acids).  Two β - chains (146 amino acids).  The globin is abbreviated α 2 β 2.

19. b- Hemoglobin A 2 : 1) It accounts about 2% of adult human hemoglobin. 2) Its globin consists of 2 α - chains and 2 delta chains (δ) : α 2 δ 2 c- Fetal hemoglobin (HbF) : 1) This is hemoglobin present in the fetus during intrautrine fetal life. 2) It consists of 2 α - chains and 2 gamma chains (α 2 γ 2 ) 3 Hemoglobin F accounts about 1% of adult human hemoglobin. d- Hemoglobin Al (Glycated hemoglobin) : 1) Hemoglobin A, reacts non enzymatically with glucose to form a derivative known as glycated hemoglobin or HbA 1c. 2) Normally the concentration of HbA 1c is very low (5-8%) but in diabetes mellitus, where blood sugar levels may be high, the concentration of HbA 1c may reach 12% or more of the total hemoglobin. E- hemoglobinopathies: 1- These are a group of diseases caused by either abnormal globin formation or synthesis of insufficient quantities of normal hemoglobin. 2- Many disorders are present, but here, two of them discussed: a- Sickle cell anaemia: 1) The blood cells of these patients contain abnormal hemoglobin call­ed hemoglobin S (HbS).

20. 2) A molecule of HbS contains 2 nor­mal α - chains and 2 mutant β ­chains in which glutamate at. po­sition six has been replaced with valine. 3) Glutamate is polar while valine is nonpolar. This single error makes hemoglobin S less soluble especially in its deoxygenated form. This will lead to:  The molecules of HbS aggregate to form fibers that deform red cells into a crescent or sickle shape.  Hemolysis of RBSickling of cells will block the flow of blood in small capi­ llaries leading to hypoxia, pain and death of cells supplied by these capillaries.  Cs. 4) Two types of sickle cell anaemia are present:  Homozygous recessive disorder: occurs in individuals who have two mutant genes coding for synthesis of 1­chains (one gene from father and the other from mother).  Heterozygous disorder:(sickle cell trait) occurs in individuals having one normal gene and one sickle cell gene. Usually patients with sickle cell trait do not show clinical symptoms except if they exposed to very low oxygen tension. b- Thalassemia: 1) Are anaemias characterized by reduced synthesis of eith­er alpha (α - tha1assemias) or beta (β- thalassemia) chains of hemoglobin. 2) The causes are most often due to gene deletions. 3) Thalassemia may be either homozygous with severe anaemia or heterozygous (thalassemia trait) with no clinical symptoms.

21. F- Abnormal derivatives of hemoglobin: 1- Methemoglobin (Met-Hb) : a- It is oxidized hemoglobin in which the ferrous ions (Fe ++ ) of hemoglobin has been oxidized to the ferric state (Fe +++ ). b- Oxidation is caused by some drugs,H 2 O 2 and number of free radicals. C- Met-Hb binds oxygen irreversibly and is unable to act as an oxygen carrier. d- If present in high concentration, met-Hb will lead to hypoxia and cyanosis. 2- Carboxyhemoglobin (COHb) : a- It is hemoglobin combining with carbon monoxide (CO). b- Carbon monoxide combines at the same position in the Hb molecule as O 2, with affinity about 200 times greater than O 2. c- Concenirafion of COHb above 40% usually result in uncon­-sciousness, and may be fatal. 3- Sulfhemoglobin (S-Hb) : a- It is hemoglobin combining with sulfur. b- It results from exposure of hemoglobin to the toxic effects of certain drugs as sulfonamides. c- S-Hb produces anoxia and cyanosis because it can not act as oxygen carrier. 4- Hematin: a- It is hemoglobin without iron (i.e. protoporphyrin combi­ning,with globin). b- It may be formed following intravascular hemolysis.

22. G- Hemoglobin catabolism: 1- The average life span of the red blood cells is 120 days. 2- At the end of that time, they are removed from circulation by the cells of reticuloendothelial (RE) system mostly present in liver, spleen and bone marrow, where they are hemolyzed (extravascular hemolysis) and hemoglobin comes out. 3- Globin. molecule is hydrolyzed into free amino acids. 4- Formation of bilirubin: a- The heme ring is catabolized by the microsomal heme oxyge­nase enzymes of the RE cells. a- The heme ring is catabolized by the microsomal heme oxyge­nase enzymes of the RE cells. b- In this reaction (which needs O 2 and NADPH),iron (Fe ++ ) is removed for re- use. The remaining of heme ring is cleaved between pyrrole rings number I and II to form. Biliverdin (green pigment) and carbori monoxide (CO). c- Biliverdin is then reducec into bilirubin (golden yellow) in a reaction requires biliverdin reductase enzyme. 5- Transport of bilirubin in the plasma:  Bilirubin is nonpolar, and is insoluble in plasma. There­ fore it binds by noncovalent bonds to plasma albumin. This form is called: unconjugated or indirect bilirubin.

23. 6- Uptake of bilirubin by the liver: a- Bilirubin dissociates from the carrier albumin molecule and enters a hepatocytes. b- Bilirubin is conjugated with one or two molecules of gluc­uronic acid (the acid form of glucose) to form bilirubin monoglucuronide and bilirubin diglucuronide.This form is called conjugated or direct bilirubin. This reaction needs UDP-lucuronyltrahsferase enzyme: 7- Secretion of bilirubin into bile:  Bilirubin diglucuronide is actively transported against concentration gradient into the bile canaliculi and then into the bile. 8- Van den Bergh reaction : a- This is a reaction between bilirubin and Ehrlich diazo reagent giving a reddish purple compound. b- Conjugated bilirubin reacts directly with the reagent. Thus it may called: direct bilirubin. c- Unconjugated bilirubin does not react with the reagent directly except after addition of alcohol. Thus it may be called: indirect bilirubin.

24. 1- Present normally in plasma. 2- Attached noncovalently to albtnnin. 3- Has high molecular weight andcan not be filtered through the kidney. 4- Nonpolar, insoluble in plasma and can cross brain barrier in neonates causing brain da­mage. 5- Gives indirect Van den Bergh reaction. 1- Present normally in bile. 2- Conjugated to glucuronic acid. 3- Has small molecular weight and if present in plasma can be filtered through the kidney. 4- Polar, soluble in plasma and can not cross brain barrier. 5- Gives direct Van den Bergh re­action. A- Normal plasma bilirubin level is up to 1 mg/dl (17.1 umol/L). B- Hyperbilirubinemia results when plasma bilirubin exceeds 1 mg/dl. C- Hyperbilirubinemia may be due to increase conjugated and/or un­conjugated bilirubin(s).

25. D- In hyperbilirubinemia, bilirubin accumulates in blood and when it reachs 3 mg/dl, it diffuses into the tissues which become yellow. This condition is called: jaundice or icterus. E- Clinically, jaundice can be detected by yellow colouration of skin, sclera and mucous membranes. F- Types of hyperbilirubinemia: It can be classified into prehepatic, hepatic and post hepatic. It can be classified into prehepatic, hepatic and post hepatic. 1- Prehepatic: (Hemolytic jaundice) : a- Hyperbilirubinemia, mostiy of unconjugated type occurs in all forms of hemolytic anaemias i.e. excessive destruction of RBCs inside blood vessels. b- It is due to increase the amount of plasma unconjugated bilirubin more than the capacity of the liver that can deal with. c- Biochemical changes: 1) Increased production of bilirubin leads to increased production of urobilinogen which appears in urine in large amounts. 2) No bilirubin appears in urine. (So that the combination of increased urobilinogen and absence of bilirubin in urine is suggestive of hemolytic jaundice).

26. 2- Hepatic: (hepatocellular jaundice) : a- It is due to liver cells damage by cirrhosis, infective hepatitis or toxins. b- Usually there is an associated obstruction of some biliary /canaliculi. c- Hyperbilirubinemia is thus a mixture of unconjugated and conjugated types. d- Biochemical changes: 1) Urobilinogen appears in normal trace amounts. 2) Bilirubin also appears in urine. e- Abnormalities at hepatpcellular level may be due to conge­nital causes (discussed later) as defect of transport of bilirubin into the cell, defective conjugation or defective excretion into the bile canalculi. 3- Posthepatic (cholestatic jaundice) : a- Cholestasis (stopagge of bile flow) may be due to mechanical obstruction of biliary tree by gallstone in common bile duct or carcinoma of the head of the pancreas or carcinoma of the biliary tree. b- Hyperbilirubinemia is mostly of conjugated type. c- Biochemical changes: 1) Urobilinogen is absent in urine. 2) Urobilinogen is absent in stool giving clay colour stool. 3) Bilirubin appears in urine.

27. d- In obstructive jaundice, bilirubin together with bile salts return to blood. Increased bile salts in blood leads to : 1) Itching because bile salts are irritant to sensory nerves. 2) Bradycardia because bile salts are to cardiac muscles. lab results in normal subjects and patients with 3 different causes of jaundice: Normal Indirect : 0.2-0.1 Direct : 0.0 – 0.2 Total : 0.2 – 1.2 0-3Absent30-300 Hemolytic anaemia Elevation of indirect IncreasedAbsentIncreased Hepatitis Elevation of indirect and indirect PresentDecreased Obstructive jaundice Elevation of indirect AbsentPresentAbsent

28. Physiologic jaundice of neonates: 1- This is a transient condition occurs in some newborn infants especially if they are premature. 2- It results from: a) At birth, liver contains very little UDP-glucuronyltransferase enzyme, which is important for conjugation of bilirubin. b) Accerelated hemolysis of RBCs. 3- This leads to increased unconjugated bilirubin which lasts 2 - 3 days in full term infants and and jaundice about 6 days in premature infants. 4- If unconjugated bilirubin exceeds the concentration which can be tightly bound to plasma albumin (20-25 mg/dl), free bilirubin can pass blood-brain barrier, causing kernicterus (toxic encephalopathy) which can cause mental retardation. 5- Kernicterus develops because the excess bilirubin is soluble in the lipid of the basal ganglia of the brain. 6- Treatment : Neonatal jaundice is treated by phenobarbital and exposure of jaundiced baby to visible light (phototherapy) as bilirubin is broken down in light.

29. H- Congenital hyperbilirubinemia: 1- Gilbert's disease: a- It is asymptomatic unconjugated hyperbilirubinemia. b- Bilirubin concentration is usually less than 3 mg/dl. c- It is due to a defect in the uptake of bilirubin by the liver parenchymal cells and a mild deficiency of UDP-glucuronyltra-­nsferase. 2. Crigler - Najjar syndrome: a- It is severe unconjugated hyperbilirubinemia. b- It occurs rarely in neonates, leading to kernicterus and often to early death. c- Bilirubin concentration is usually exceeds 20 mg/dl. d- It is due to marked reduction in the UDP-glucuronyltransferase. 3. Dubin - Johnson syndrome: a- It is a conjugated hyperbilirubinemia which occurs during adult life. b- It is due to defect in the hepatic secretion of conjugated bilirubin into the bile. I- Neonatal/jaundice: These are a group of diseases including phys­iologic and congenintal jaundice and other hemolytic diseases as Rh incompatibility.