Transcript Liver Function Tests (LFTs) Measurement of Serum Bilirubin (Total

T.A. Bahiya Osrah
Bilirubin

 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
Bilirubin Structure:

Bilirubin consists of
four open chain
pyrrols, unlike heme
which consists of four
rings pyrrols called
(porphyrin).
Bilirubin
Heme
Types of bilirubin in
serum

 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
Bilirubin Production

 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.
 hemoglobin destroyed to the heme + globin
amino acid
 Iron is removed from the heme molecule,
 porphyrin ring is opened to form bilirubin
Bilirubin Transportation

 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
Specific carrier
mechanism
Conjugation of bilirubin and
secretion into bile

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
Further metabolic processes are
occurred in intestine and kidney .
Further metabolism of bilirubin
in the gut

 In the intestine:
Bilirubin diglucuronides
Bacteria
Glucouronic acid
+
un-conjugatedbilirubin
Urobilinogen
Further metabolism of
bilirubin in the gut

Reabsorptio
nInto the blood
Increased level in the blood
Jaundice
90%
10%
Jaundice

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
Pre-hepatic
Hepatic
Post-hepatic
SUMMARY

Pre-hepatic (before bile is made
in the liver)

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
Hepatic (the problem arises
within the liver)

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.
Post-hepatic (after bile has been
made in the liver)

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 claycolored. 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
Physiologic jaundice of
the newborn

 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.
Summery

Pre-hepatic jaundice

Hepatic jaundice

Due to increase in RBCs
breakdown due to
hemolytic anemia.

Cause
Type of Bil
Conformational
test

The rate of RBCs
degradation and
hemoglobin production
more than ability of liver to
convert it to conjugated
form.
Post-hepatic jaundice
Due to liver cell

damage or due to
cancer or
cirrhosis
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”.
ID.Bil > D.Bil
High D.Bil, ID.Bil,
and T.Bil
High D.Bil
K+ (high)
Hematology: CBC, low Hb
ALT, AST
ALP (high)
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
Kit pamphlet

The Lab practice

Method principle:
Serum sample composed of :
direct + indirect bilirubin = total bilirubin
Diazotized
sulfanilic
acid
1 min
Methanol
Accelerate the reaction
Azobilirubin
(Purple colored compound)
5 min
Prodedure

Calculations

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
Diagnosis

 First:
 Second
Compare your test result with the expected value
Summery

Pre-hepatic jaundice

Hepatic jaundice

Due to increase in RBCs
breakdown due to
hemolytic anemia.

Cause
Type of Bil
Conformational
test

The rate of RBCs
degradation and
hemoglobin production
more than ability of liver to
convert it to conjugated
form.
Post-hepatic jaundice
Due to liver cell

damage or due to
cancer or
cirrhosis
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”.
ID.Bil > D.Bil
High D.Bil, ID.Bil,
and T.Bil
High D.Bil
K+ (high)
Hematology: CBC, low Hb
ALT, AST
ALP (high)
Diagnosis

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