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BILE PIGMENT OR
BILIRUBIN
METABOLISM

Haemoglobin
•Haemoglobin is a conjugated protein
made up of four subunits

•Each subunit contains a haem group and

a polypeptide chain

•The haem group is made up of

porphyrin and a ferrous ion

•Haemoglobin can reversibly

combine with oxygen, and transports
oxygen in the body
3/1/2018 Rajendra Dev Bhatt

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Catabolism of Haemoglobin
1.Generation of Bilirubin

 When life-span of RBCs is over, they are broken

down in reticulo-endothelial system
 Haem and globin are separated.
 Globin is broken down into amino acids.
 The iron liberated from heme is reutilized.
 The porphyrin ring is broken down in reticuloendothelial
(RE) cells of liver, spleen and bone marrow to bile
pigments, mainly bilirubin

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1. Generation of Bilirubin Cont…

 About 6 g of Hb is broken down per day, from which

about 250 mg of bilirubin is formed
 From myoglobin and other heme containing proteins,
another 50 mg of bilirubin is formed.
 Approximately 35 mg of bilirubin is formed from 1 g
of Hb.
 A total of 300 mg of bilirubin is formed everyday; of
which 80% is from destruction of old RBCs, 10% from
ineffective erythropoiesis and the rest 10% from
degradation of myoglobin and other heme containing
proteins.

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1. Generation of Bilirubin Cont…

 Heme is degraded primarily by a microsomal
enzyme system; heme oxygenase. It requires
molecular oxygen and NADPH
 The oxygenase enzyme specifically catalyzes the
cleavage of the alpha methenyl bridge, which is
linking the pyrrole rings I and II.
 The linear tetrapyrrole formed is biliverdin which is
green in color.
 In mammals it is further reduced to bilirubin, a red-
yellow pigment, by an NADPH dependent biliverdin
Reductase.

1. Generation of Bilirubin Cont…

In the first reaction, a

bridging methylene group
i s c l e av ed b y hem e
oxygenase to form Linear
Biliverdin from Cyclic
Heme molecule.
Fe 2+ is released from the
ring in this process.

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1. Generation of Bilirubin Cont…

RBCs Breakdown

Hemoglobin Produces & Breakdown

Heme
Heme
Oxygenase Biliverdin
Biliverdin
Reductase
Bilirubin

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2. Transport to Liver
 The bilirubin formed in the reticuloendothelial cells is
insoluble in water.
 The lipophilic bilirubin is therefore transported in plasma bound to
albumin.
 One molecule of albumin can bind 2 molecules of bilirubin.
 Albumin binds bilirubin in loose combination. So when present in
excess, bilirubin can easily dissociate from albumin.
 The binding sites for bilirubin on albumin can be occupied by
aspirin, penicillin, etc. Such drugs can, therefore, displace bilirubin
from albumin. Hence, care should be taken while administering such
drugs to newborn babies to avoid kernicterus.
 When the albumin–bilirubin complex reaches the sinusoidal surface
of the liver, the bilirubin is taken up.
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2. Transport to Liver

Bilirubin is not very water-soluble, so most of it is carried to the liver bound to albumin.

3. Conjugation in Liver
 Inside the liver cell, the bilirubin is conjugated
with glucuronic acid, to make it water soluble.
 The first carbon of glucuronic acid is combined with the
carboxyl group of the propionic acid side chains of the
bilirubin molecule. About 80% molecules are in the
diglucuronide form, while 20% are monoglucuronides.
 Drugs like primaquine, novobiocin, chloramphenicol,
androgens and pregnanediol may interfere in this
conjugation process and may cause jaundice.

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3. Conjugation in Liver

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Unconjugated bilirubin:
Bilirubin that are not conjugated with gluconic
acid , also called hemobilirubin, indirect
bilirubin.

conjugated bilirubin:
Bilirubin that are conjugated with gluconic acid,
also called hepatic bilirubin, direct bilirubin.

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Unconjugated VS Conjugated Bilirubin

FEATURE Unconjugated Conjugated

Normal serum level More Less (less than 0.2mg/dl)

Water solubility Absent Present

Affinity to lipids (alcohol Present Absent

solubilty)
Serum albumin binding High Low
Van den Bergh reaction Indirect (Total minus Direct
direct)
Reanal excretion Absent Present

Affinity to brain tissue Present (kernicterus) Absent

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4. Excretion of Bilirubin to Bile

 The water soluble conjugated bilirubin is excreted
into the bile by an active process and this
occurs against a concentration gradient.
 This is the rate limiting step in the catabolism of
heme. It is induced by phenobarbitone.
 Excretion of conjugated bilirubin into bile is mediated
by an ATP binding cassette protein which is called
Multispecific organic anion transporter (MOAT),
located in the plasma membrane of the biliary
canaliculi.

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4. Excretion of Bilirubin to Bile Cont..

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5.Fate of Conjugated Bilirubin in Intestine

 The conjugated bilirubin reaches the intestine through the bile.
 In the small intestine, conjugated bilirubin is poorly reabsorbed
and 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 (44 Hydrogen)
c) Further reduction of UBG leads to formation of stercobilinogen
(SBG,48 hydrogen)
 The SBG is mostly excreted through feces (250-300 mg/day).
 Upto 20 % of urobilinogen produced daily is reabsorbed from the intestine
& enters the entero-hepatic circulation.
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5.Fate of Conjugated Bilirubin in Intestine

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6. Final Excretion
UBG and SBG are both colorless compounds
but are oxidized to colored products, urobilin
(42 hydrogen) or stercobilin (46 hydrogen)
respectively by atmospheric oxidation.
Black color is seen in constipation. If intestinal
flora is decreased by prolonged administration
of antibiotics, bilirubin is not reduced to
bilinogens, and in the large gut, it is re-oxidized
by O2 to form biliverdin. Then green tinged feces
is seen, especially in children.
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BLOOD
Stercobilin
CELLS Urobilin
excreted in feces
Hemoglobin excreted
in urine
Globin
Urobilinogen
Heme
O2 formed by bacteria KIDNEY
reabsorbed
Heme oxygenase INTESTINE into blood
CO

Biliverdin via bile duct to intestines

NADPH
Biliverdin Bilirubin diglucuronide
reductase (water-soluble)

NADP+ 2 UDP-glucuronic acid

Bilirubin Bilirubin
(water-insoluble) LIVER
(water-insoluble) via blood
unconjugated to the liver

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Summary of bilirubin metabolism

Old red cells are major source of hemeproteins

Breakdown of heme to bilirubin occur in macrophage of reticuloendithelial

system ( tissue macrophages, spleen and liver).

Unconjugated bilirubin is transported through blood ( complex to albumin) to liver.

Bilirubin is taken into liver and conjugate with glucuronic acid.

Bile is secreted into intestine where glucuronic acid is removed and the resulting
bilirubin is converted to urobilinogen.

A portion of urobilinogen is reabsorbed into blood, where it is converted to the

yellow urobilin and excreted by kidneys.

Urobilinogen is oxidized by intestinal bacteria to the brown stercobilin.

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