BIOLOGIC

BIOLOGIC OXIDATION
OXIDATION
ENERGI
(ATP)
Source of ATP :
1. Oxidative Phosphorylation.
2. Glycolysis.
3. Krebs Cycle.

Ox-red reactions
 O2 accept single electron
 Oxidation-reduction potential

• Oxidation : the removal of electrons

• Reduction : the gain of electrons
• Redox potential (E0’) : the free energy change
is proportionate to the tendency of reactants
to donate or accept electrons
• Redox potential of a system (Eo) is compared
with the potential of the hidrogen electrode
• Biologic systems  E0’ expressed at PH 7
and electrode potential of H : – 0,42 volts
 System E’O volts
H+/H­2 -0.42
NAD+/ NADH -0.32
Lipoate; ox / red -0.29
Acetoacetate/ 3 – -0.27
hydroxybutyrate -0.19
Pyruvate/ lactate -0.17
Oxaloacetate/ malate +0.03
Fumarate/ succcinate +0.08
Cytochrome b; Fe3+/Fe2+ +0.10
Ubiquinone; ox/red +0.22
Cytocrome c1; Fe3+/Fe2+ +0.29
Cytocrome a; Fe3+/Fe2+ +0.82
Oxygen/ water
 Enzymes in ox-red
• Called oxidoreductases (class I),
classified into 4 groups:
- oxidases
- dehydrogenases
- hydroperoxidases
- oxygenases
 Oxidases
• Catalyzing the removal hydrogen and using
oxygen as a acceptor  form water or
hydrogen peroxide
• Some oxidases contain copper and others
are flavoproteins
• Cytochrome oxidase ( cyt.a.a3 ):
heme protein contain Cu
terminal component of respiratory chain
contain two molecules of heme as
prosthetic group and Cu
 Oxidases

• Flavoprotein enzyms contain FMN or FAD as

prosthetic groups
• FMN and FAD are formed in body from riboflavin
• They are tightly bound to their apoenzymes 
but not covalently
• Exampels: L-amino acid oxidase (in kidney),
xanthine oxidase (in intestinal, kidney, liver),
aldehyde oxidase (in liver) and glucose
oxidase (in fungus)
AH2 / 2 O2
1 AH2 O2
(Red)

OXIDASE OXIDASE

A H2O A H2O2
(Ox)
A B

Oxidation of a metabolite catalyzed by an oxidase (A)

forming H2O, (B) forming H2O2
 Dehydrogenases
• Can not use oxygen as a hydrogen acceptor
• Performing two main functions:
1. transfer hydrogen in a coupled
oxidation reduction reaction
specific for their substrates, but utilize
common coenzymes
useful in enabling oxidative process to
occur in the absence of oxygen
2. components in respiratory chain 
transfer electron from
substrate to oxygen
Dehydrogenases link NAD
• Using NAD+ or NADP+ as a coenzyme
• These coenzyme are formed in body from
niacin
- freely and reversibly dissociate from their
apoenzymes
- NAD linked D-ase: oxidative pathways of
metabolism (glycolysis, kreb’s cycle,
respiratory chain)
- NADP linked D-ase: characteristically in
reductive synthesis (fatty acid synthesis,
steroid synthesis and PMP-shunt)
Dehydrogenases link riboflavin
• Using FMN and FAD as a coenzyme
- more tightly bound to their apoenzymes
- most of them are concerned with electron
transport in / to resp chain
- NADH D-ase  carrier of electrons
between NADH and components of higher
redox potential
- succinate D-ase, acyl Co-A D-ase, glycerol
3 P D-ase  transfer electrons directly from
substrate to resp. chain
 Cytochromes as dehydrogenase

• Classified as dehydrogenases, except for

cytochrome oxidase
- as carriers of electrons from flavoproteins to
cytochrome oxidase in the resp chain
- exampels: cyt b, c1, c, a, a3 (resp chain) and
cyt P 450, b5 (endoplasmic reticulum)
AH2 Carrier BH2
(Red) (Ox) (Red)

A Carrier-H2 B
(Ox) (Red) (Ox)
DEHYDROGENASE DEHYDROGENASE
SPECIFIC FOR A SPECIFIC FOR B

Oxidation of a metabolite catalyzed by coupled

dehydrogenases
 Hydroperoxidases

• Using hydrogen peroxide or an organic

peroxide as substrate
• Two type: - peroxidase
- catalase
• Protecting against harmful peroxides
• Peroxides  generate free radicals 
disrupt membranes and cause cancer and
atherosclerosis
• Peroxidases
Reducing peroxides using various electron
acceptors (ascorbate, quinones, cyt c):
H2O2 + AH2  2H2O + A
Founding in milk, leukocytes, platelets,
erythrocytes and other tissues involved in
eicosanoid metabolism
Glutathione peroxidase, containing selenium
 destruction of H2O2 and lipid
hydroperoxidases  protecting membrane
lipids and Hb
• Catalase
Using hydrogen peroxide as electron donor and
electron acceptor:
2 H2O2  2H2O + O2
In addition to possessing peroxidase activity, it
is able to use one of H2O2 as a substrate
(electron donor) and another of H2O2 as an
oxidant (electron acceptor)
Founding in blood, bone marrow, mucous
membranes, kidney and liver
 A’H2 A’

AH2
A

O2 H2O2 CATALASE 2H2

OXIDASE
O

H2O2 O2

Role of catalase in the destruction of hydrogen peroxie

 Oxygenases
• Catalyzing the direct transfer and
incorporation of oxygen into a substrate
• Divided into two subgroups:
1. Dioxygenases / oxygen transferase
Incorporating both atoms of oxygen into
substrate: A + O2  AO2
2. Monooxygenases
Mixed function oxidases and
hydroxylases  incorporate only 1 atom
of oxygen into substrate, the other oxygen
is reduced to water
 MONOOXYGENASES

• Need an additional electron donor /

cosubstrate ( Z ):
A-H + O2 + ZH2  A-OH + H2O + Z
• Cytochromes P450 are monooxygenases (as
cosubstrate )  important for
detoxification of many drugs and for
hydroxylation of steroids
• NADH and NADPH donate reducing
equivalents for the reduction of cyt P450
 Cytochrome P 450
• Mitochondrial cyt P450 systems in
steroidogenic tissues  biosynthesis of
steroid hormones from cholesterol
• Mitochondrial cyt P450 systems in kidney
 metabolism of vitamin D
• Mitochondrial cyt P450 systems in liver 
biosynthesis of bile acid
 Superoxide free radicals (O2-)
• Generated from transfer of a single
electron to O2
• It is formed  reduced flavin, are
reoxidized univalently by molecular oxygen
• Superoxide dismutase  in aerobic
organisms  removal O2- ,  the reaction:
O2- + O2- + 2H+  H2O2 + O2
• Superoxide can reduce oxidized cyt c:
O2- + cyt c (Fe3+)  O2 + cyt c (Fe2+)
• Exposure to an atmosphere of 100%
oxygen causes an adaptive increase in
superoxide dismutase
 OXIDATIVE
OXIDATIVE
PHOSPHORYLATION
PHOSPHORYLATION
 Oxidative phosphorylation
• Oxidative reaction Coupled by
phosphorylation to the generation of high
energy intermediate (ATP or other high
phosphagen)
• Oxidative phosphorylation at resp chain
level  via NAD D-ases form 3 mol ATP
and via flavoprotein D-ases form 2 mol ATP
• Phosphorylations at the substrate level 
captured smaller energy  eg:a) High
energy phosphates are captured in kreb’s
cycle during the conversion of succinyl Co-
A to succinate. And b) in glycolytic
reactions on cytoplasmic.
 Respiratory chain
• Enzyme complexes in mitochondria 
collects and transports reducing equivalents
 directing them to final reaction with
oxygen  form water and ATP
• Reducing equivalents flow through from
redox potential negative to positive
• There are 4 enzyme complexes:
- NADH-Q dehydrogenase / I
- Succinate-Q dehydrogenase / II
- Cytochromes dehydrogenase / III
- Cytochrome oxidase / IV
DEHYDROGENASE DEHYDROGENASE DEHYDROGENASE OXIDASE

Carrier Carrier Carrier

AH2 1 2 3 H2O
(Red) (Ox) (Red) (Ox)

Carrier-H2 Carrier Carrier-H2

A 2 / 2 O2
1
1 3
(Ox) (Ox)
(Red) (Red)

Oxidation of a metabolite by dehydrogenases and finally

by an oxidase in a respiratory chain
 AH2 NAD+ FpH2 2Fe3+ H2O

Substrate Flavoprotein Cytochrome

s
A NADH Fp 2Fe2+ 1
/2O

H+ H+ 2H
+ 2H+ 2

Transport of reducing equivalents through the respiratory

chain
 Mitochondrial
• Powerhouses of the cell  most of energy
captured takes place inside it
• Outer membrane  permeable to most
metabolites, contain various enzym (acyl Co-
A synthetase, glycerolphosphate
acyltransferase )
• Inner membrane  selectively permeable
• Matrix  contain phospholipid cardiolipin
together with enzymes of resp chain
• Intermembrane space has similar
composition with cytoplasmic and contain
adenylyl kinase and creatine kinase
 A B MATRIX
F1 subunits
OUTER
Phosphorylating F0 subunits
MEMBRANE
B complexes
INNER
MEMBRANE

MATRIX

Sonication
Cristae

INNER
MEMBRANE
OUTER
MEMBRANE
Submitochondrial particel
Formed from fragments of
the inner membrance
 Respiratory chain

• Not all substrates are linked to resp chain

through NAD-D-ase
• Co-Q (ubiquinone)  mobile component,
collects reducing equivalents from
flavoprotein complexes and passes them on
to cytochrome b (the lowest redox pot)
• Cytochrome oxidase has a very high
affinity for oxygen  resp chain to
function at maximum rate until tissue
depleted of O2 irreversible reaction
 Proline Succin
3-Hydroxyacyl- ate
CoA Cholin
3-Hydroxybutyrate e
Pyruvat Glutamate Fp
e Malate (FAD)
Isocitrate FeS

Fp Fp Cyt aa3
Lipoate Cyt b Cyt c1 Cyt O2
(FAD) NAD (FMN) Q
FeS c Cu
FeS
FeS
- ETF
Fp FeS : Iron-sulfur
Ketoglutarate (FAD)
(FAD) protein
FeS ETF : Electron-
Fp transferring
(FAD) flavoprotein
Fp : Flavoprotein
Acyl-CoA Q : Ubiquinone
Sarcosine Cyt : Cytochrome
Glycerol 3-phosphate Dimethylglycin
 Resp chain & oxd phos inhibitors
• Inhibitors of resp chain
1. Blocking electrons transfer from Fe-S to
co-Q , ie: barbiturates , pierisidin-A ,
rotenon , carboxine ,
succinate D’ase competitive inhibitor:
malonate
2. Blocking electrons transfer from cty b to
cyt c, ie: dimercaprol , antimycin A
3. Inhibitors of cytochrome oxidase: H2S , CO
and CN
 Resp chain & oxd phos inhibitors

• Inhibitors of oxidative phosphorylation, ie:

oligomycine, atractyloside
• Un-couplers (dissociate oxidation in resp
chain from phosphorylation)  respiration
to become uncontrolled, ie: dinitrophenol,
dinitrochressol, pentachlorophenol, chloro
carbonyl cyanide phenilhydrazon (cccp)
 FAD
Succinate
FeS
H2S
BAL CO
Antimycin A CN -

Complex I Complex III Complex IV

NADH FMN, FeS Cyt b, FeS, Cyt c Cyt a Cyt a3 O2

Q
Cyt C1 Cu Cu

Piericidin A
Amobarbital
Rotenone Uncouplers
Uncouplers

Oligomycin
Oligomycin

ADP + P1 ATP ADP + P1 ATP ADP + P1 ATP

 Mechanism of oxidative
phosphorylation
• Mitchell’s chemiosmotic theory:
- energy from oxidation in resp chain 
translocation of H+ (protons) 
electrochemical potential difference in
matrix and intermembrane space 
drive the mechanism of responsible for
the formation of ATP (ATP synthase)
 Mechanism of oxidative
phosphorylation
• Complexes I, III and IV of resp chain is a
proton pump
• Pi + ADP  ATP, by ATP synthase
• ATP synthase is a complex enzyme  consist
of several protein subunits (F1), which
attached to membrane protein complex (F0)
• F1  project into matrix and contain the
phosphorylation mechanism
F0  spans the membrane and forms
the proton channel
 Exchange metabolites at inner
mitochondrial membrane
- Exchange of anions against OH- ions and
cations against H+ ions  for transport of
ionized metabolites
- Freely permeable to uncharged small
molecules  O2 , H2O , CO2 , NH3
monocarboxylic acids (3 hydroxy butyric,
acetoacetic, acetic)
- Long chain fatty acids need carnitine system
- Symport pyruvate - H+
 Exchange metabolites at inner
mitochondrial membrane
• Dicarboxylate and tricarboxylate anions
require specific carrier  linked to
inorganic phosphate (H2PO4- )
• Exchange ATP / ADP by adenine nucleotide
transporter
• Transport of oxaloacetate need
transamination process
Oxidation of extramitochondrial NADH

- NADH cannot penetrate mitochondrial

membrane  produced continuously in
cytosol by 3 phosphoglyceraldehyde D-ase
- Aerobic conditions: not accumulated  be
oxidized by resp chain
- Transfer of reducing equivalents from
cytosol to mitochondrial require substrate
pairs, linked by suitable D-ase
Oxidation of extramitochondrial NADH
- The mechanism:
1. Glycerophosphate shuttle  only 2 mol
ATP are formed per atom oxygen
consumed  present in brain, muscle,
adipose, liver but deficient in heart muscle
2. Malate shuttle  more universal utility
 more complex, due to the
impermeability of mitochondrial membrane
to oxaloacetate
 OUTER INNER
MEMBRANE MEMBRANE

MITOCHONDRION
CYTOSOL

NAD+
Glycerol 3- Glycerol 3-
phosphate phosphate

GLYCEROL-3- GLYCEROL-3- FAD

PHOSPHATE PHOSPHATE
DEHYDROGENASE DEHYDROGENASE
(CYTOSOLIC) (MITHOCONDRIAL)

Dehydroxyacetone Dehydroxyacetone
NADH + phosphate phosphate
H+ FDH2

Respiratory Chain

Glycerophosphate shuttle for transfer of reducing equivalents from the cytosol

into the mitochondrion
 INNER
MAMBRAN
CYTOSOL E MITHOCOND
RION
1
NAD+ Malat Malate NAD+
e

MALATE MALATE
DEHYDROGENASE DEHYDROGENASE
NADH NADH
+H+ Oxaloacetat -KG -KG Oxaloacetat +H+
e e
TRANSAMI TRANSAMI
NASE NASE
Glutamate Asp Asp Glutamate

H+ H+

Malate shuttle for transfer of reducing equivalents from the

cytosol into the mitocondrion. 1. Ketoglutarate transporter, 2.
 Creatine phosphate shuttle
• Facilitating transport of high energy
phosphat from mitochondria in active
tissues
• Isoenzyme of creatine kinase (CKM), in
intermembrane space  catalyzing
transfer ~ P (ATP) to creatine:
~ P(ATP) + creatine  creatine-P ,
transported into cytosol via protein pores
 available for generation of
extramitochondrial ATP
 H

P
N CREATINE H2N
KINASE

C NH C NH

N H3C N

ΔGO’ = 12.6
COO- kJ/mol COO-

Creatine
Creatine
phosphate
 Clinical aspects

• Fatal infantile mitochondrial myopathy and

renal dysfunction  due to severe
diminution / absence of most
oxidoreductase
• MELAS (mitochondrial encephalopathy,
lactic acidosis and stroke)  due to
complex I or complex IV deficiency 
mutation in mt DNA