1. T.A. Bahiya Osrah

2.   Bilirubin is the product of heme degradation  (80% hemoglobin, 20% other hemo-protein as cytochrome, myoglobin).  Elevated levels of bilirubin in blood and urine indicate certain diseases. Bilirubin

3.   Bilirubin Structure: Bilirubin consists of four open chain pyrrols, unlike heme which consists of four rings pyrrols called (porphyrin). Bilirubin Heme

4.   Indirect bilirubin: is unconjugated or water insoluble, it is called indirect because it reacts slowly, so it indicates indirect reaching to reagent.  Direct bilirubin: is conjugated or water soluble it is called direct because it reacts faster, so it indicates direct reaching to reagent.  Note: Total bilirubin = D+ ID  Knowing the level of each type of bilirubin has diagnostic important Types of bilirubin in serum

5.   After approximately 120 days in the circulation, red blood cells are taken up and degraded by the reticuloendothelial (RE) system, particularly in the liver, spleen and in the bone marrow.  hemo globin destroyed to the heme + globin amino acid  Iron is removed from the heme molecule,  porphyrin ring is opened to form bilirubin Bilirubin Production

6.   Bilirubin is insoluble in water and is carried in plasma bound to albumin  On reaching the liver, the bilirubin is taken into the hepatocyte by specific carrier mechanism Bilirubin Transportation Specific carrier mechanism

7.  In the liver: Glucouronic acid + un-conjugated bilirubin (water insoluble) UDP-glucuronyltransferase Bilirubin diglucuronides (water soluble) Bilirubin diglucuronides are water soluble and readily transported into bile Conjugation of bilirubin and secretion into bile Further metabolic processes are occurred in intestine and kidney.

8.   In the intestine: Bilirubin diglucuronides Bacteria Glucouronic acid + un-conjugatedbilirubin Further metabolism of bilirubin in the gut Urobilinogen

9.  Further metabolism of bilirubin in the gut Reabsorptio n Into the blood 90% 10% Increased level in the blood Jaundice

10.  Jaundice: Is a term used in clinical medicine to describe a condition in which the skin and sclera appear yellow caused by increased amounts of bilirubin in the blood Classification of the causes of Jaundice: 1. Prehepatic jaundice 2. Hepatic jaundice 3. Posthepatic jaundice

11.  SUMMARY Pre-hepatic Hepatic Post-hepatic

12.  Jaundice in these cases is caused by rapid increase in the breakdown and destruction of the red blood cells (hemolysis), overwhelming the liver's ability to adequately remove the increased levels of bilirubin from the blood. Examples of conditions with increased breakdown of red blood cells include:  Malaria  sickle cell crisis  Thalassemia  glucose-6-phosphate dehydrogenase deficiency (G6PD)  drugs or other toxins  autoimmune disorders Pre-hepatic (before bile is made in the liver)

13.  Jaundice in these cases is caused by the liver's inability to properly metabolize and excrete bilirubin. Results from: Impaired cellular uptake. Defective conjugation. Abnormal secretion of bilirubin by the liver cell. Hepatic (the problem arises within the liver)

14.  Jaundice in these cases, also termed obstructive jaundice, is caused by conditions which interrupt the normal drainage of conjugated bilirubin in the form of bile from the liver into the intestines. This may due to: gallstones in the bile ducts, tumor  Rise in the serum conjugated bilirubin level and stool becomes clay- colored. Why? Because of the normal drainage interruption of conjugated bilirubin in the form of bile from the liver into the intestines urine urobilinogen levels got decreased therefore the secretion of sterocobilin resulted to a clay-colored stool Post-hepatic (after bile has been made in the liver)

15.   High bilirubin levels are common in newborns age (1-3 days old).  It is happened because:  breaking down the excess RBCs  because the newborn’s liver is not fully mature, it is unable to process the extra bilirubin, leads to elevate its level in blood and other body tissues.  Usually newborn is treated by phototherapy which breakdown bilirubin (ID<<<<D) and convert it to the photo isomer form which is more soluble.  Very high bilirubin is danger and toxic. It may cause brain damage and affect on muscles, eyes and even death. Physiologic jaundice of the newborn

16.  Summery Post-hepatic jaundiceHepatic jaundicePre-hepatic jaundice  Due to the obstruction of hepatobiliary duct.  D.Bil is formed in liver but can’t pass to bile, so it accumulates in liver and transferred to blood “instead of bile”.  Due to liver cell damage or due to cancer or cirrhosis  Due to increase in RBCs breakdown due to hemolytic anemia.  The rate of RBCs degradation and hemoglobin production more than ability of liver to convert it to conjugated form. Cause High D.Bil High D.Bil, ID.Bil, and T.Bil ID.Bil > D.Bil Type of Bil ALP (high)ALT, AST K+ (high) Hematology: CBC, low Hb Conformational test

17.  Calculations The absorbance of bilirubin equivalent standard represents: 1.Direct bilirubin=2.5 mg/dl 2.Total bilirubin= 5 mg/dl 3.Direct bilirubin after 1min= (abs test- abs test blank/abs std )* 2.5 4.Total bilirubin after 5 min= (abs test- abs test blank/abs std )* 5 5.To convert mg/dl into µmol/l, multiply the final results by 17.1 The Lab practice

18.  Kit pamphlet

19.  The Lab practice Method principle: Serum sample composed of : direct + indirect bilirubin = total bilirubin Diazotized sulfanilic acid Azobilirubin ( Purple colored compound ) 1 min Methanol Accelerate the reaction 5 min

20.  Prodedure

21.  Calculations

22.  The Lab practice Calculations: The absorbance of bilirubin equivalent standard represents: 1.Direct bilirubin=2.5 mg/dl 2.Total bilirubin= 5 mg/dl 3.Direct bilirubin after 1min= (abs test- abs test blank/abs std )* 2.5 4.Total bilirubin after 5 min= (abs test- abs test blank/abs std )* 5 5.To convert mg/dl into µmol/l, multiply the final results by 17.1

23.   First:  Second Compare your test result with the expected value Diagnosis

24.  Summery Post-hepatic jaundiceHepatic jaundicePre-hepatic jaundice  Due to the obstruction of hepatobiliary duct.  D.Bil is formed in liver but can’t pass to bile, so it accumulates in liver and transferred to blood “instead of bile”.  Due to liver cell damage or due to cancer or cirrhosis  Due to increase in RBCs breakdown due to hemolytic anemia.  The rate of RBCs degradation and hemoglobin production more than ability of liver to convert it to conjugated form. Cause High D.Bil High D.Bil, ID.Bil, and T.Bil ID.Bil > D.Bil Type of Bil ALP (high)ALT, AST K+ (high) Hematology: CBC, low Hb Conformational test

25.  The patient sex is not important here Normal Range in the kit : Direct UP TO 0.5 mg/dl Total UP TO 1 mg/dl 1. The test value above the normal range: Abnormal value Possible disorders are:  ID= T – D,,,Then say  If ID>> D it is pre-hepatic jaundice due to increase in RBCs breakdown due to hemolytic anemia. We need to send the sample to the hematology lab and for CBC  If all high and Id is Not >> D it is hepatic jaundice (liver disease) we need to do ALT and AST tests  Direct is the only one high then it is Post-Hepatic jaundice due to the obstruction of hepatobiliary duct 2. No abnormal value here below the normal range (NO decrease case) 3. Within the range : NORMAL value Diagnosis