Bilirubin Metabolism
Dr Raghuveer Choudhary
Associate professor
Department of physiology
Dr s.n.medical college jodhpur
Introduction
• Bilirubin is the orange-yellow pigment derived from
senescent red blood cells.
• It is a toxic waste product in the body.
• It is extracted and biotransformed mainly in the liver, and
excreted in bile and urine.
• It is a bile pigment
• Elevations in serum and urine bilirubin levels are normally
associated with Jaundice.
Erythrocytes become “old” as they lose their flexibility
and become pikilocytes (spherical), increasingly rigid
and fragile. Once the cell become fragile, they easily
destruct during passage through tight circulation
spots, especially in spleen, where the intra-capillary
space is about 3 micron as compared to 8 micron of
cell size
RBCs useful life span is 100 to 120 days,After
which they become trapped and fragment in smaller
circulatory channels, particularly in those of the spleen.
For this reason, the spleen is sometimes called the “red
blood cell graveyard.”
Dying erythrocytes are engulfed and destroyed by
macrophages.
Formation of Bilirubin
• Primary site of synthesis:SPLEEN: The Graveyard
of Red Blood Cells
• Secondary site of synthesis:-
LIVER & BONE MARROW
 An average person
produces about 4
mg/kg of bilirubin
per day.
 The daily bilirubin
production from all
sources in man
averages from 250
to 300 mg.
TOTAL BILIRUBIN
85%
HEMOGLOBIN
FROM SENESCENT
RBC’S DESTROYED IN
RETICULOENDOTHELIAL
CELLS OF
LIVER, SPLEEN &
BONE MARROW
15%
RBC PRECURSORS
DESTROYED IN THE
BONE MARROW
CATABOLISM OF
HEME-CONTAINING
PROTEINS (MYOGLOBIN,
CYTOCHROMES &
PEROXIDASES)
Extravascular Pathway for RBC Destruction
(Liver, Bone marrow,
& Spleen)
Phagocytosis & Lysis
Hemoglobin
Globin
Heme
Amino acids
Fe2+
Amino acid pool
Recycled
Bilirubin
Excreted
Within cells of the RE system, heme degraded to bilirubin in a two-step process
.
• a. The porphyrin ring is opened and the
iron atom is removed by the action of
• heme oxygenase to produce the greencolored intermediate biliverdin.
• b. Subsequent reduction converts
biliverdin to bilirubin, which has a
redorange color.
Pathophysiology
RBCs
Breakdown
Hemoglobin Produces
& Breakdown
Heme
Heme
Oxygenase
Biliverdin
Biliverdin
Reductase
Bilirubin
• The globin is recycled or converted into amino acids, which
in turn are recycled or catabolized as required.
• Heme is oxidized, with the heme porphyrin ring being
opened by the endoplasmic reticulum enzyme, heme
oxygenase.
• The oxidation occurs on a specific carbon producing
equimolar amounts of the biliverdin, iron , and carbon
monoxide (CO). This is the only reaction in the body that is
known to produce CO.
• Most of the CO is excreted through the lungs, with the
result that the CO content of expired air is a direct measure
of the activity of heme oxygenase in an individual.
In the first reaction, a
b r i d g i n g m e t h yl e n e
group is cleaved by
heme oxygenase to
form Linear Biliverdin
from Cyclic Heme
m o l e c u l e .
Oxidation
Heme Oxygenase
Fe 2+ is released from
the ring in this process.
I
III
IV
II
Heme Oxygenase
I
IV
Fe2+
III
C
NADPH
II
O2
O2
IV
III
II
Biliverdin
I
H
NADPH
Bilirubin
•
I
III
IV
II
In the next reaction, a second
bridging methylene (between
rings III and IV) is reduced by
biliverdin reductase,
producing bilirubin.
Reduction
I
III
Biliverdin Reductase
IV
II
• biliverdin causing a change in the color of the molecule from
blue-green (biliverdin) to yellow-red (bilirubin).
• The latter catabolic changes in the structure of tetrapyrroles
are responsible for the progressive changes in color of a
hematoma, or bruise, in which the damaged tissue changes
its color from an initial dark blue to a red-yellow and finally
to a yellow color before all the pigment is transported out of
the affected tissue.
• Peripherally arising bilirubin is transported to the liver in
association with albumin, where the remaining catabolic
reactions take place.
Bilirubin is not very water-soluble, so most of it is carried to the liver bound to
albumin.
In cells of the liver, bilirubin undergoes modification to
increase its water solubility so that it can be excreted more
easily.
a.Bilirubin is conjugated to two molecules
of glucuronic acid, creating bilirubin
diglucuronide.
b. Bilirubin diglucuronide is transported
out of the hepatocytes into the bile
canaliculi and is thus excreted in bile.
In Blood
• The bilirubin synthesized in
spleen, liver & bone marrow
is unconjugated bilirubin.
• It is hydrophobic in nature so
it is transported to the liver
as a complex with the
plasma protein, albumin.
Unconjugated bilirubin
– Lipid soluble
– : limits excretion
– 1 gm albumin binds 8.5
mg bilirubin
– Fatty acids & drugs can
displace bilirubin
– Indirect positive reaction
in van den Bergh test
Role of Blood Proteins in the
Metabolism of Bilirubin
1. Albumin
Dissolved in Blood
Blood
Liver
Ligandin
Ligandin
(-) charge
(-) charge
Ligandin Prevents bilirubin from
going back to plasma
In Endoplasmic Reticulum
In the microsomes of the endoplasmic reticulum,
unconjugated bilirubin is converted to water soluble
mono- or di- conjugates by sequential covalent
coupling with glucuronic acid.
Bilirubin is conjugated in
a two step process to
form bilirubin mono- &
di- glucuronide
Conjugation with Glucoronates
BILIRUBIN DIGLUCORONIDE
Excretion of Bilirubin
In the Intestine
•
In the small intestine, conjugated bilirubins are poorly
reabsorbed, but are partly hydrolyzed back to unconjugated
bilirubin by catalytic action of bacterial ß-glucuronidases.
•
In the distal ileum and colon, anaerobic flora mediate further
catabolism of bile pigments:
a) hydrolysis of conjugated bilirubin to unconjugated bilirubin by
bacterial β-glucuronidases;
b) multistep hydrogenation (reduction) of unconjugated bilirubin to
form colorless urobilinogens; and
c) oxidation of unconjugated bilirubin to brown colored
mesobilifuscins.
• Urobilinogens is a collective
term for a group of 3
tetrapyrroles;
– Stercobilinogen (6H)
– Mesobilinogen (8H)&,
– Urobilinogen (12H)
• Upto 20 % of urobilinogen
produced daily is reabsorbed
from the intestine & enters the
entero-hepatic circulation.
Urobilinogen Structure
• Most of the reabsorbed urobilinogen is taken up by the liver
& is re-excreted in the bile.
• A small fraction (2 % - 5 %) enters the general circulation &
appears in the urine.
• In the lower intestinal tract, the 3 urobilinogens
spontaneously oxidize to produce the corresponding bile
pigments;
– Stercobilin
– Mesobilin &
– Urobilin;
which are orange-brown in color and are the major
pigments of stool.
JAUNDICE
Clinical Significance
Hyperbilirubinemia & Types of Jaundice
• Hyperbilirubinemia : Increased plasma concentrations
of bilirubin (> 3 mg/dl) occurs when there is an
imbalance between its production and excretion.
•
Recognized clinically as jaundice.
• Also known as icterus, a yellow discoloration of the skin,
sclerae and mucous membrane.
• Jaundice becomes clinically evident when the serum
bilirubin level exceeds 2.5mg/dL.
• Several types of Jaundice:
– Hemolytic
– Hepatocellular
– Obstructive
• Symptoms:
– Yellow discoloration of the skin, sclerae and mucous
membranes
– Itching (pruritus) due to deposits of bile salts on the skin
– Stool becomes light in color
– Urine becomes deep orange and foamy
Different Causes of Jaundice
•
•
•
•
Excessive Production of Bilirubin
Reduced Hepatocyte Uptake
Impaired Bilirubin conjugation
Impaired Bile Flow
Classification
Jaundice
Pre-hepatic
Hepatic
Post-Hepatic
Prehepatic (hemolytic) jaundice
• Results from excess production
of bilirubin (beyond the livers
ability to conjugate it) following
hemolysis
• Excess RBC lysis is commonly
the result of autoimmune
disease; hemolytic disease of the
newborn (Rh- or ABOincompatibility); structurally
abnormal RBCs (Sickle cell
disease); or breakdown of
extravasated blood
• High plasma concentrations of
unconjugated bilirubin (normal
concentration ~0.5 mg/dL)
Hepatic jaundice
• Impaired uptake, conjugation,
or secretion of bilirubin
• Reflects a generalized liver
(hepatocyte) dysfunction
• In this case,
hyperbilirubinemia is usually
accompanied by other
abnormalities in biochemical
markers of liver function
• Hemolytic jaundice arises as a
consequence of excessive destruction
of RBCs.
• – This overloads the capacity of the RE
system to metabolize heme.
• – Failure to conjugate bilirubin to
glucuronic acid causes accumulation of
bilirubin in the unconjugated form in the
blood.
• Hepatocellular jaundice arises from
liver disease, either inherited or
acquired.
• – Liver dysfunction impairs conjugation of
bilirubin.
• – Consequently, unconjugated bilirubin
spills over into the blood.
• –In addition, urobilinogen is elevated in the
urine.
Posthepatic(Obstructive) jaundice
• Caused by an obstruction of
the biliary tree.
• Plasma bilirubin is conjugated,
and other biliary metabolites,
such as bile acids accumulate
in the plasma.
• Characterized by pale colored
stools (absence of fecal
bilirubin or urobilin), and dark
urine (increased conjugated
bilirubin).
• In a complete obstruction,
urobilin is absent from the
urine.
• • Obstructive jaundice, as the name implies,
is caused by blockage of the bile duct by a
gallstone or a
• tumor (usually of the head of the pancreas).
• – This prevents passage of bile into the intestine
and consequently conjugated bilirubin builds up
in the blood.
• – Patients with this condition suffer severe
abdominal pain associated with the
obstruction (if due togallstone) and their feces
are gray in color due to lack of stercobilin.
Pre-hepatic
Hepatic
Post hepatic
cause
Excessive break down
Of RBC’s
Malaria,HS
Gilbert Syndrome
Infective
Liver Damage
Bile Duct Obstruction
Serum Bilirubin
unconjugated
Both conj+unconj.
conjugated
Urine bilirubin
Absent
Achloric jaundice
Bilirubinemia +
Deep yellow urine
As in hepatic jaundice
++
Absent(-)
Because of increased
stercobilinogen
Decreases
Because of decreased
stercobilinogen
Fecal
stercobilinogen
20-250mg/day
Fecal fat 5-6%
Markedly increased
Dark brown stool
Reduced
Pale coloured stool
Absent
clay colored stool
normal
Increased 40-50%
Bulky,pale greasy foul
smelling faeces
As hepatic jaundice
Liver functions
normal
Impaired SGOT/SGPT
Normal
Alkaline phosphatase++
Vonden burg test
Indirect+
biphasic
Direct+
Urine urobilinogen Increases
Neonatal Jaundice
• Common, particularly in premature infants.
• Transient (resolves in the first 10 days).
• Due to immaturity of the enzymes involved in bilirubin
conjugation.
• High levels of unconjugated bilirubin are toxic to the
newborn – due to its hydrophobicity it can cross the
blood-brain barrier and cause a type of mental
retardation known as kernicterus
• If bilirubin levels are judged to be too high, then
phototherapy with UV light is used to convert it to a
water soluble, non-toxic form.
• If necessary, exchange blood transfusion is used to remove
excess bilirubin
• Phenobarbital is oftentimes administered to Mom prior to
an induced labor of a premature infant – crosses the
placenta and induces the synthesis of UDP glucuronyl
transferase
• Jaundice within the first 24 hrs of life or which takes longer
then 10 days to resolve is usually pathological and needs to
be further investigated
Phototherapy
•Phototherapy is usually not needed
unless the bilirubin levels rise very
quickly or go above 16-20 mg/dl in
healthy, full term babies.
During phototherapy, the
treatment of choice for
jaundice, babies are placed
under blue lights that convert
the bilirubin into compounds
that can be eliminated from
the body.
•
Bilirubin Toxicity - Kernicterus
• Kernicterus or brain encephalopathy refers to the yellow
staining of the deep nuclei (i.e., the kernel) of the brain
namely, the basal ganglia.
• It is a form of permanent brain damage caused by
excessive jaundice.
• The concentration of bilirubin in serum is so high that it can
move out of the blood into brain tissue by crossing the fetal
blood-brain barrier.
• This condition develops in newborns with prolonged
jaundice due to:
– Polycythemia
– Rh incompatibility between mother & fetus
Inherited Disorders of Bilirubin
Metabolism
•
•
•
•
•
•
Gilbert’s Syndrome
Crigler-Najjar (Type I)
Crigler-Najjar (Type II)
Lucey-Driscoll
Dubin-Johnson
Rotor’s Syndrome
Algorithm for differentiating the familial causes of
Hyperbilirubinemia
Isolated increased serum bilirubin
Ruling out of hemolysis, subsequent fractionation of the bilirubin
Conjugated
Possibility of the
following syndromes:
• Dublin-Johnson
• Rotor
Unconjugated
Possibility of following syndromes
based on the bilirubin concentration:
• Gilbert’s - <3 mg/dl
• Crigler-Najjar (Type I) - >25 mg/dl
• Crigler-Najjar (Type II) - 5 to 20 mg/dl
• Lucey-Driscoll - Transiently ~ 5 mg/dl
Crigler-Najjar Syndrome (Type I)
• Crigler-Najjar Syndrome (Type I) is a rare genetic disorder
caused by complete absence of UDP-glucuronyltransferase
and manifested by very high levels of unconjugated
bilirubin.
• It is inherited as an autosomal recessive trait.
• Most patients die of severe brain damage caused by
kernicterus within the first year of life.
• Early liver transplantation is the only effective therapy.
Crigler-Najjar Syndrome (Type II)
• This is a rare autosomal dominant disorder.
• It is characterized by partial deficiency of UDPglucuronyltransferase.
• Unconjugated bilirubin is usually 5 – 20 mg/dl.
• Unlike Crigler-Najjar Type I, Type II responds
dramatically to Phenobarbital & a normal life can be
expected.
Gilbert’s Syndrome
• Gilbert’s syndrome is also called as familial non-hemolytic
non-obstructive jaundice.
•
mild unconjugated Hyperbilirubinemia.
• It affects 3% – 5% of the population. It is often misdiagnosed
as chronic Hepatitis.
• The concentration of Bilirubin in serum fluctuates between 1.5
& 3 mg/dl.
• In this condition the activity of hepatic glucuronyltransferase is
low as a result of mutation in the bilirubin-UDPglucuronyltransferase gene(UGT1A1).
Dubin-Johnson Syndrome
• It is a benign, autosomal recessive
condition characterized by jaundice
with predominantly elevated
conjugated bilirubin and a minor
elevation of unconjugated bilirubin.
• Excretion of various conjugated
anions and bilirubin into bile is
impaired, reflecting the underlying
defect in canalicular excretion.
• The Liver has a characteristic
greenish black appearance and liver
biopsy reveals a dark brown melaninlike pigment in hepatocytes and
kupffer cells.
Rotor’s Syndrome
• It is another form of conjugated hyperbilirubinemia.
• It is similar to dubin-johnson syndrome but without
pigmentation in liver.