Porphyrins

& Bile Pigments

FY WIDODO

University of Wijaya Kusuma Surabaya Medical Faculty

Department of Biochemistry
 PORFIRIN
• Cyclic compounds formed by the linkage
of four pyrrole rings through methyne (—
HC =) bridges
• Formation of complexes with metal ions
bound to the nitrogen atom of the pyrrole
rings
Examples of Some Important Human Hemoproteins
Protein Function
Hemoglobin Transport of oxygen in blood
Myoglobin Storage of oxygen in muscle
Cytochrome c Involvement in electron transport chain
Cytochrome P450 Hydroxylation of xenobiotics
Catalase Degradation of hydrogen peroxide
Tryptophan pyrrolase Oxidation of tryptophan
Cytochrome
 A (acetate)
P (propionate)
M (methyl)

Addition of iron to protoporphyrin

to form heme. V (vinyl) = —CHCH2.
 Formation of Heme
• The two starting materials are succinyl-CoA and glycine
• Pyridoxal phosphate  "activate" glycine
• Condensation reaction between succinyl-CoA and glycine  a-amino-
b-ketoadipic acid d-aminolevulinate (ALA)
decarboxilation

• Enzyme: ALA synthase MITOCHONDRIA

Two molecules of ALA are condensed by the enzyme ALA

dehydratase to form two molecules of water and one of
porphobilinogen (PBG)
ALA dehydratase is a zinc-containing enzyme and is
sensitive to inhibition by lead, as can occur in lead
poisoning.
CYTOSOL
• The formation of a cyclic tetrapyrrole—ie, a porphyrin—occurs by
condensation of four molecules of PBG

• These four molecules condense in a head-to-tail manner to form a

linear tetrapyrrole, hydroxymethylbilane (HMB)

• Enzyme: uroporphyrinogen I synthase = porphobilinogen (PBG)

deaminase = HMB synthase

cyclizes spontaneously  uroporphyrinogen I

• HMB
uroporphyrinogen III synthase  uroporphyrinogen III

• These compounds are colorless (porphyrinogens)  

 auto-oxidized  porphyrins (colored).
• uroporphyrinogen I  uroporphyrinogen decarboxylase
 coproporphyrinogen I
• uroporphyrinogen III  coproporphyrinogen III
Acetate (A)  methyle (M) : decarboxylated by
uroporphyrinogen decarboxylase
cytosol

coproporphyrinogen III  coproporphyrinogen oxidase

protoporphyrinogen III
• protoporphyrinogen III  protoporphyrinogen oxidase 
protoporphyrin III
• Requires molecular oxygen.
• Protoporphyrin III + Fe2+  Heme
Enzyme: ferrochelatase (heme synthase)
mitochondria
Decarboxylation of uroporphyrinogens to coproporphyrinogens in
cytosol. (A, acetyl; M, methyl; P, propionyl.)

Heme biosynthesis occurs in most mammalian cells with the exception of

mature erythrocytes, which do not contain mitochondria.
However, approximately 85% of heme synthesis occurs in erythroid
precursor cellsin the bone marrow and the majority of the remainder in
hepatocytes.
 Regulation in Biosynthesis of Heme
• ALA synthase occurs in both hepatic (ALAS1) and erythroid (ALAS2)
forms
• The rate-limiting enzyme: ALAS1  feedback inhibition
Heme (-)  ALAS1 Heme (+) ALAS1

• Aporepressor molecule + Heme  negative regulator of the synthesis

of ALAS1

• Drugs (eg, barbiturates, griseofulvin)  increase in ALAS1.

Most of these drugs are metabolized by a system in the liver that
utilizes cytochrome P450

• Glucose can prevent derepression of ALAS1 in liver as can the

administration of hematin (an oxidized form of heme).
 PORPHYRIAS
• Disorders due to abnormalities in the pathway of biosynthesis of
heme  genetic or acquired
 Enzyme Involved Type, Class, Major Signs and Results of Laboratory
Symptoms Tests

ALA synthase (erythroid X-linked sideroblastic Anemia Red cell counts and
form) anemia3 (erythro-poietic) hemoglobin decreased

ALA dehydratase ALA dehydratase defi- Abdominal pain, Urinary ALA and
ciency (hepatic) neuropsychiatric coproporphyrin III
symptoms increased
Uroporphyrinogen I Acute intermittent Abdominal pain, Urinary ALA and PBG
synthase porphyria (hepatic) neuropsychiatric increased
symptoms
Uroporphyrinogen III Congenital erythropoietic No photosensitivity Urinary, fecal, and red cell
synthase (erythropoietic) uroporphyrin I increased

Uroporphyrinogen Porphyria cutanea tarda Photosensitivity Urinary uroporphyrin I

decarboxylase (hepatic) increased

Coproporphyrinogen Hereditary Photosensitivity, abdo- Urinary ALA, PBG, and

oxidase coproporphyria (hepatic) minal pain, neuropsy- coproporphyrin III and fecal
chiatric symptoms coproporphyrin III increase
Protoporphyrinogen Variegate porphyria Photosensitivity, abdo- Urinary ALA, PBG, and
oxidase (hepatic) (OMIM 176200) minal pain, neuropsy- coproporphyrin III and fecal
chiatric symptoms protoporphyrin IX increased
Ferrochelatase Protoporphyria Photosensitivity Fecal and red cell
(erythropoietic) (OMIM protoporphyrin IX increased
177000)
• In general, the porphyrias described are inherited in an autosomal
dominant manner, with the exception of congenital erythropoietic
porphyria, which is inherited in a recessive mode
• The signs and symptoms of porphyria result from either a deficiency of
metabolic products beyond the enzymatic block or from an
accumulation of metabolites behind the block.
• If the enzyme lesion occurs early in the pathway, clinically, patients
complain of abdominal pain and neuropsychiatric symptoms  relate
to elevated levels of ALA or PBG or to a deficiency of heme
• If the enzyme blocks later in the pathway result in the accumulation of
the porphyrinogens, their oxidation products, the corresponding
porphyrin derivatives, cause photosensitivity, a reaction to visible light
of about 400 nm The porphyrins, when exposed to light of this
wavelength, are thought to become "excited" and then react with
molecular oxygen to form oxygen radicals  injure lysosomes and
other organelles  release their degradative enzymes  skin
damage, including scarring.
• The porphyrias can be classified on the basis of the organs or cells
that are most affected: erythropoietic, hepatic & erythrohepatic
• Barbiturates, griseofulvin  cytochrome P450  heme  ALAS1
 porphyria
• Diagnosis: clinical and family history, the physical examination, and
appropriate laboratory tests
• Treatment: - symptomatic
- to avoid drugs that cause induction of cytochrome P450.
- glucose,hematin  may repress ALAS1
- photosensitivity : b-carotene  free radicals
Sunscreens
 Katabolisme Heme
• 1–2 x 108 erythrocytes are destroyed per hour  in 1 day turns over
approximately 6 g of hemoglobin
• globin  amino acids  reused; iron pool
• Enzyme: complex enzyme system called heme oxygenase
 METABOLISME BILIRUBIN

PENGAMBILAN BILIRUBIN OLEH HATI

• Bilirubin hanya sedikit larut dalam plasma & terikat dengan albumin
• Obat / antibiotika  kompetisi untuk berikatan dg albumin
• LIVER: Bilirubin dilepas dari albumin  diambil pada permukaan
sinusoid hapatosit  Sistem Transport Berfasilitas (facilitated
transport system = carrier-mediated saturable system)  masuk ke
sel hati berikatan dengan cytosolic protein (ligandin, protein Y).
Ikatan ini juga menjaga agar bilirubin tidak kembali ke aliran darah
lagi.
Aktivitas sistem ini menurun pada keadaan patologis
Biliverdin

Bilirubin
KONJUGASI BILIRUBIN
• LIVER: Bilirubin yang non-polar  polar
• Konjugasi dengan GLUKORONAT  Bilirubin diglukoronida
(conjugated, "direct-reacting" bilirubin)  polar

• Enzyme: glucuronosyltransferase (dlm retikulum endoplasma)

• Donor glukuronosil: UDP-glucuronic acid
• Enzim dapat diinduksi oleh Phenobarbital
• Bilirubin diglukoronida diekskresi melalui faeces
• when exist abnormally in human plasma (eg, in obstructive jaundice),
they are predominantly monoglucuronides
• Obstructive jaundice  bilirubin conjugates (predominan
monoglukuronida)

Bilirubin diglukoronida
METABOLISME BILIRUBIN DALAM USUS
Diglukoronida
• Bilirubindiglukoronida Bilirubin
b-glukoronidase Bakteri usus

• Bilirubin direduksi oleh flora usus  UROBILINOGEN (tak berwarna)

• Sebagian kecil urobilinogen diabsorbsi & diresekresi melalui liver (SIKLUS
ENTEROHEPATIK)
• Sebagian besar urobilinogen  oksidasi  UROBILIN  faeces
• Urobilinogen dlm urine  abnormal
Diagrammatic representation of the three
major processes (uptake, conjugation,
and secretion) involved in the transfer of
bilirubin from blood to bile. Certain proteins
of hepatocytes, such as ligandin (a member
of the glutathione S-transferase family of
enzymes) and Y protein, bind intracellular
bilirubin and may prevent its efflux into the
blood stream. The process affected in a
number of conditions causing jaundice
is also shown.
HIPERBILIRUBINEMIA
• Bilirubin dlm darah >1 mg/dl.
Penyebab: - produksi bilirubin h tidak sebanding dg ekskresi hati
- kegagalan ekskresi hati karena:
- kerusakan hati
- obstruksi saluran ekskresi
• Bilirubin masuk ke jaringan  KUNING / ICTERUS / JAUNDICE
• Ehrlich’s test dari van den Bergh:
- reagen diazo + Bilirubin + etanol  senyawa Azo (ungu-merah)
INDIRECT REACTION  BILI UNCONJUGATED (FREE)
- reagen diazo + Bilirubin (tanpa etanol)  senyawa Azo
DIRECT REACTION  BILI CONJUGATED
• Retention hyperbilirubinemia: unconjugated hh
Regurgitation hyperbilirubinemia : conjugated hh
• Bili unconjugated dpt menembus blood-brain barrier  encephalopathy
 Kern icterus
Jaundice
UNCONJUGATED HYPERBILIRUBINEMIA
A. Hemolytic Anemias
Biasanya ringan (< 4 mg/dL; < 68.4 mmol/L)  kapasitas hati besar
utk “mengelola” bilirubin (uptake, konjugasi, ekskresi)

B. Neonatal Physiologic Jaundice

- Hemolisis > metabolisme bilirubin dlm hati
- aktivitas / sintesis UDP-glukoronosil transferase K
- Dpt menyebabkan Kern icterus
- Tx: Phenobarbital +
C. Crigler-Najjar Syndrome, Type I
- = Congenital nonhemolytic jaundice
- Autosomal resesif; mutasi pada gen pengkode enzim Bilirubin-
UGT  UDP-glukoronosil transferase K
- Bilirubin bisa sd > 20 mg/dL  bilirubin tdk bisa dikonjugasikan
- Klinis: ikterus kongenital berat  fatal dalam 15 bulan
- Tx: , liver transplant. Phenobarbital tdk menolong.
D. Crigler-Najjar Syndrome, Type II
- Mutasi pada gen pengkode enzim Bilirubin-UGT, tdk seberat
tipe I, bilirubin serum tdk sampai 20 mg/dL
- Dapat di Tx dg Phenobarbital dosisi tinggi
E. Gilbert Syndrome
- Mutasi pada gen pengkode enzim Bilirubin-UGT, sering pada pria
- Klinis tdk berat, aktivitas enzim masih ada sekitar 30 %
F. Toxic Hyperbilirubinemia
- chloroform, arsphenamine, CCl4, acetaminophen, virus hepatitis,
chirrhosis, jamur beracun  kerusakan parenkim hati
CONJUGATED HYPERBILIRUBINEMIA
A. OBSTRUKSI SALURAN EMPEDU
- Penyebab: baru empedu, ca. caput pankreas.
- Bilirubin diglukoronida tidak bisa diekskresi  regurgitasi ke vena-
vena di liver dan saluran limfe  bilirubin masuk ke aliran darah dan
urine (choluric jaundice)
- Cholestatic Jaundice: extrhepatic obstructive jaundice
B. DUBIN-JOHNSON SYNDROME
- Autosomal resesif, mutasi gen MRP-2
- Hepatosit pada area centrilobular mengandung pigmen hitam yang
abnormal yang merupakan derivat epinephrine.
C. ROTOR SYNDROME
- Hiperbilirubinemia terkonjugasi yang kronis, histopatologi hepar tetap
normal
- Etiologi: tdk tahu
Diagrammatic representation of some major causes of jaundice

Prehepatic indicates events in

the blood stream, the major
cause would be various forms
of hemolytic anemia
Hepatic signifies events in the
liver, such as the various types
of hepatitis or other forms of
liver disease (eg. cancer)
Posthepatic refers to evens in
the billiary tree, the major
causes of posthepatic jaundice
are obstruction of the common
bile duct by a gallstone (billiary
calculus) or bya cancer of the
head of pancreas
 Laboratory Results in Normal Patients and Patients with
Three Different Causes of Jaundice
Condition Serum Bilirubin Urine Urine Fecal
Urobilinogen Bilirubin Urobilinogen

Normal Direct: 0.1–0.4 0–4 mg/24 h Absent 40–280 mg/24 h

mg/dL
Indirect: 0.2–0.7
mg/dL
Hemolytic Indirect h Increased Absent Increased
anemia
Hepatitis Direct and Decreased if Present if Decreased
indirect h micro- micro-
obstruction is obstruction
present occurs

Obstructive Direct h Absent Present Trace to absent

jaundice