Transcript Bilirubin
Liver Function profile (LFT) Excretion functions (Bilirubin measurement) Khadija Balubaid KAU-Faculty of Science- Biochemistry department Clinical biochemistry lab (BIOC 416) 2013 Liver Function Test (LFT) profile Integrity of liver cells Execratory function Synthetic function Clinical Lab tests Liver enzymes AST, ALT, GGT, ALP Bilirubin, ALP Proteins TP, Alb, A/G ratio Definition of bilirubin Bilirubin is the water insoluble breakdown product of normal heme catabolism It’s a yellow pigment present in bile ( a fluid made by the liver) , urine and feces . Heme is found in hemoglobin, a principal component of RBCs [Heme: iron + organic compound “porphyrin”]. Heme source in body: 80% from hemoglobin 20% other hemo-protein: cytochrome, myoglobin) Heme and bilirubin Heme four pyrrols rings connected together to form (porphyrin). Bilirubin consists of open chain of four pyrrols-like rings Hemoglobin degrading and bilirubin formation Spleen Plasma Hemoglobin globin Protein and a.a pool Heme iron Bilirubin Liver Conjugation process Iron pool Binds with albumin Bilirubin Metabolism 1. Unconjugation process : RBCs are phagocytized in the spleen. Hemoglobin is catabolized into amino acids, iron and heme. Heme ring is broken open and converted to unconjugated ( indirect ) bilirubin. This unconjugated bilirubin is not soluble in water, due to intramolecular hydrogen bonding. It is then bound to albumin and sent to the liver. 2. Congugation process: In liver: Bilirubin is conjugated with Glucouronic acid to produce bilirubin diglucuronides, which is water soluble and readily transported to bile. and thus out into the small intestine. Blilirubin + Glucouronic acid "water insoluble" UDP-glucuronyl transferase bilirubin diglucuronides "water soluble" "Conjugated BIL" Bile Then conjugated bilirubin is excreted in bile through bile duct to help in food digestion (mainly fat). The excess amount transferred to intestine to be excreted in urine and stool. However 95% of the secreted bile is reabsorbed by the small intestine. This bile is then resecreted by the liver into the small intestine. This process is known as enterohepatic circulation About half of the conjugated bilirubin remaining in the large intestine (about 5% of what was originally secreted) is metabolised by colonic bacteria to form urobilinogen , which may be further oxidized to urobilin and stercobilin . Urobilin, stercobilin and their degradation products give feces its brown color.[ Elevated levels of bilirubin in blood and urine indicate certain diseases. Direct bilirubin: is conjugated (water soluble bilirubin) in aqueous solution it reacts rapidly with reagent (direct reacting). Indirect bilirubin: is unconjugated (water insoluble bilirubin) because it is less soluble in it reacts more slowly with reagent (reaction carried out in methanol). - in this case both conjugated and unconjugated bilirubin are measured given total bilirubin. Unconjugated will calculated by subtracting direct from total and so called indirect. Total bilirubin = D+ ID • Knowing the level of each type of bilirubin has diagnostic important. • It is a medical term describes the elevation of bilirubin in blood result in yellow color of skin and sclera. • Other symptoms include nausea, vomiting, dark-colored urine andTypes of Jaundice: • fatigue. • according to the cause of jaundice it is classified to three main types: Pre-hepatic jaundice Hepatic jaundice Post-hepatic (most common type) haemolytic jaundice Pre-hepatic jaundice Due to increase in RBCs breakdown due to hemolytic anemia. Causes The rate of RBCs lysis and bilirubin production more than ability of liver to convert it to the conjugated form Occur in: Erythroblastosis fetalis Hemolytic anemia hepato-cellular jaundice Hepatic jaundice Due to liver cell damage (cancer, cirrhosis or hepatitis) Conjugation of bilirubin decreased (ID.Bil. ). obstructive jaundice Post-hepatic jaundice Due to obstruction of bile duct which prevents passage of bilirubin into intestine. Blilirubin that is conjugated is D.Bil will back to liver and then to circulation not efficiently secreted into elevating its level in blood bile but leaks to blood (D.Bil. and urine. ) Occur in: Occur in : Cirrhosis (scarring of the liver) Biliary stricture Hepatitis Gilbert's disease Cancer of the pancreas or gallbladder Gallstones Transfusion reaction Type of Bil. ID.Bil > D.Bil D.Bil, ID.Bil, T.Bil all (High) D.Bil (High) Conformational test K+ ( High) Hematology: CBC (low Hb) ALT, AST (High) ALP ( High) • High bilirubin levels is common in newborns age (1-3 days). • After birth the newborns breaking down the excess RBCs they are born with and, because the newborn’s liver is not fully mature, (unable to process the extra bilirubin) leads to elevate its level in blood and other body tissues. • This situation usually resolves itself within a few days SO, WHAT TYPE OF JUNDUCE IS THIS ??? Note:Your child's doctor must consider the following when deciding whether your baby's bilirubin levels are too high: How fast the level has been rising Whether the baby was born early How old the baby is New born jaundice treatment: • Usually newborn is treated by phototherapy which breakdown bilirubin (IDD) and convert it to the photo isomer form which is more soluble. Bilirubin Toxicity : Very high bilirubin is danger and toxic it may cause brain damage effect on muscles, eyes and Leading to death Bilirubin measurement How to Prepare for the Test You should not eat or drink for at least 4 hours before the test. Your health care provider may instruct you to stop taking drugs that affect the test. Many drugs may change the bilirubin levels in your blood. Make sure your doctor knows which medications you are taking. Tell your doctor if you have allergy Why the Test is Performed Large amounts of bilirubin in the blood can lead to jaundice. Jaundice is a yellow color in the skin, mucus membranes, or eyes. Jaundice is the most common reason to check bilirubin levels. Most newborns have some jaundice. The doctor or nurse will often check the newborn's bilirubin level. See: Newborn jaundice The test may also be done in older infants, children, and adults who develop jaundice. A bilirubin test will also be done if your doctor thinks you may have liver or gallbladder problems Procedure Measuring serum bilirubin level Principle: Sulphanalic acid + NaNO3 DSA + Bilirubin “D” Bilirubin “ID”+ DSA + accelerator diazotized sulphanalic acid (DSA) Azobilirubin “purple” (methanol ) Total bil. Kit components Sulfanalic acid reagent Sodium nitrate reagent Methanol reagent Bilirubinequavalent standard (5mg/dl T.bil; 2.5 mg/dl D.bil) Procedure: Equivalent Standard Test blank Test Sulfanilic acid 1.4ml 1.4 ml 1.4 ml NaNO3 25ml - 25ml 25ml - dis. H2O Mix, stand for 1 min Sample 100 µl (Standard) 100 µl (serum) 100 µl (serum) - Read the Abs after 1 min at 540nm, (Blank dis. H2O) Use this to calculate D.bil Methanol 1.5 ml 1.5 ml 1.5 ml - Mix by inversion, stand 5 min or more - Read Abs. at 540nm (Blank dis. H2O) Use this to calculate T.bil Pour Bilirubinequavelant standard in clean cuvette read Abs. at 540nm, (blank dis. H O) Calculations • Conc. of Bilirubin equavelant is 5mg/dl for T.bil, and 2.5 mg/dl for D.bil D.Bil: Abs (test) - Abs (test blank) X 2.5 mg/dl Abs of bilirubin equivalent T.Bil: Abs (test) - Abs (test blank) X Abs of Bilirubin equivalent • To convert to mmol/L multiply by 17.1 5 mg/dl Normal Results It is normal to have some bilirubin in your blood. Normal levels are: Direct (also called conjugated) bilirubin: 0 to 0.5 mg/dL Total bilirubin: 0.3 to 1.9 mg/dL Note: mg/dL = milligrams per deciliter Normal value ranges may vary slightly among different laboratories. Case Study This 48-year-old man had complained of abdominal pain and intermittent fever for 3 months. The pain was usually felt in the right upper quadrant. His appetite was not good and lost his body weight apparently. He once suffered from hepatitis ten years ago. Physical examination revealed the patient who appeared chronically ill with icteric sclera. The liver was enlarged to 2 cm below the costal margin with tenderness, and the spleen was enlarged to 3 cm below costal margin Laboratory data: Hb 9 mg/dL, WBC 8.0×109/L, PC 90×109/L, ALT 1020U/L, AST 800U/L,ALP 255U/L, TP 55g/L, A 25g/L, Y 40%, TBI 55umol/L, DBI 38umol/L, URO (++), UBI (+) Question: What is your diagnosis for this patient? Which kind of jaundice this patient has? How to evaluate this patients liver function?