Chapter BIOCHEMISTRY
OF BLOOD
Piao jinhua
English name: joy
BIOMEDICAL IMPORTANCE

The study of its constituents has been of central
importance in the development of biochemistry
and clinical biochemistry.

Changes in the amounts of various plasma
proteins occur in many diseases

Alterations of the activities of certain enzymes
found in plasma are of diagnostic use in a
number of pathologic conditions.
Section 1
Plasma proteins
Ⅰ.The composition of blood

Ⅰ）The formed
element—the
cellular components
Red blood cells (Erythrocytes)
White blood cells (Leukocytes)
Platelets ( thrombocytes )

Ⅱ）The liquid
element
plasma
Plasma: The blood fraction
obtained after removal of the
cellular components.
NPN
The nitrogen content of substances other
than protein in blood, tissues, and waste
materials.
urea
Serum: Separation of the protein
fibrinogen from plasma yields the
blood fraction.
 Serum
is generally obtained by
allowing the blood to clot.
 fibrinogen
fibrin
Ⅱ.The functions of blood
Major functions of blood
 (1)
Respiration—transport of oxygen
from the lungs to the tissues and of
CO2 from the tissues to the lungs
 (2)
Nutrition—transport of absorbed
food materials
Major functions of blood
Excretion—transport of metabolic
waste to the kidneys, lungs, skin, and
intestines for removal
 (3)
Maintenance of the normal acidbase balance in the body
 (4)
Major functions of blood
 (5)
Regulation of water balance
through the effects of blood on the
exchange of water between the
circulating fluid and the tissue fluid
 (6)
Regulation of body temperature
by the distribution of body heat
Major functions of blood
 (7)
Defense against infection by
the white blood cells and
circulating antibodies
 (8)
Transport of hormones and
regulation of metabolism
Major functions of blood
 (9)
Transport of metabolites
 (10)
Coagulation
Ⅲ.Plasma contains a complex
mixture of proteins
The proteins of the plasma
simple proteins
total protein
7.0-7.5 g/100ml
conjugated proteins
glycoproteins
lipoproteins
The proteins of the plasma are
actually a complex mixture
 eg:Thousands
of antibodies are
present in human plasma, though
the amount of any one antibody
is usually quite low under normal
circumstances.
The relative dimensions and molecular masses
of some of the most important plasma proteins
The separation of individual
proteins
 salting-out
methods
 Using solvents or electrolytes (or both) to
remove different protein fractions in
accordance with their solubility
characteristics.
 sodium sulfate or ammonium sulfate
by the use of varying concentrations
of sodium or ammonium sulfate



proteins of the plasma
Fibrinogen
albumin
globulins
The method of analyzing plasma
proteins
 electrophoresis
 An
electrochemical process in which
macromolecules or colloidal particles
with a net electric charge migrate in a
solution under the influence of an
electric current.
 cellulose acetate is widely used as a
supporting medium
Technique of cellulose acetate
zone electrophoresis




A: A small amount of serum or
other fluid is applied to a cellulose
acetate strip.
B: Electrophoresis of sample in
electrolyte buffer is performed.
C: Separated protein bands are
visualized in characteristic positions
after being stained.
D: Densitometer scanning from
cellulose acetate strip converts
bands to characteristic peaks of
albumin, α1-globulin, α2-globulin,
β-globulin, and γ-globulin.
Ⅲ.1
The concentration of protein in
plasma is important in
determining the distribution of
fluid between blood and tissues
In arterioles
 hydrostatic
pressure 37 mmHg
 interstitial (tissue) pressure 1 mmHg
 The osmotic pressure (oncotic pressure
exerted by the plasma proteins ) 25
mmHg
a net outward force of about 11 mm Hg drives
fluid out into the interstitial spaces
In venules
 the
hydrostatic pressure is about 17
mmHg
 interstitial (tissue) pressure 1 mmHg
 The osmotic pressure (oncotic pressure)
25 mmHg
a net force of about 9 mm Hg attracts water
back into the circulation
Edema
 If
the concentration of plasma proteins is
markedly diminished (eg, due to severe
protein malnutrition),
 fluid is not attracted back into the
intravascular compartment and accumulates
in the extravascular tissue spaces, a condition
known as edema
 Edema has many causes; protein deficiency
is one of them.
Ⅲ.2
Plasma proteins have been
studied extensively
 Considerable
information is available
about the biosynthesis, turnover, structure,
and functions of the major plasma
proteins.
 Alterations of their amounts and of their
metabolism in many disease states have
also been investigated.
 many of the genes for plasma proteins
have been cloned and their structures
determined.
Many of the preparations have been
used in the study of the plasma proteins.
 The
preparation of antibodies specific for the
individual plasma proteins
allowing the precipitation and isolation of
pure proteins from the complex mixture
present in tissues or plasma.
 the use of isotopes
this has made possible the determination of
their pathways of biosynthesis and of their
turnover rates in plasma.
generalizations have emerged from
studies of plasma proteins
1. Most plasma proteins are synthesized in the liver
 the whole-animal level (hepatectomy)

isolated perfused liver preparation
 by use of
liver slices

liver homogenates

in vitro translation systems ( using preparations of
mRNA extracted from liver )
 γ-globulins are synthesized in plasma cells

generalizations have emerged from
studies of plasma proteins
 2.
Plasma proteins are generally
synthesized on membrane-bound
polyribosomes:

traverse the major secretory route in the cell
 rough
endoplasmic membrane → smooth
endoplasmic membrane → Golgi apparatus →
secretory vesicles → plasma
 most
plasma proteins are synthesized as
preproteins
 subjected to various posttranslational
modifications as they travel through the cell.
(proteolysis, glycosylation, phosphorylation,
etc)
 Transit times through the hepatocyte from the
site of synthesis to the plasma vary from 30
minutes to several hours or more for
individual proteins.
generalizations have emerged from
studies of plasma proteins
3. Almost all plasma proteins are glycoproteins
 they contain either N- or O- linked oligosaccharide
chains, or both.
 Removal of terminal sialic acid residues from
certain plasma proteins (eg, ceruloplasmin) by
exposure to neuraminidase can markedly shorten
their half-lives in plasma.
 Albumin does not contain sugar residues.

generalizations have emerged from
studies of plasma proteins
 4.
Many plasma proteins exhibit
polymorphism
 A polymorphism
is a mendelian or monogenic
trait that exists in the population in at least two
phenotypes, neither of which is rare (neither of
which occurs with frequency of less than 12%).
 The ABO blood group substances
Human plasma proteins that exhibit
polymorphism include
 α1-antitrypsin
Haptoglobin
 Transferrin
 ceruloplasmin,
 immunoglobulins

 The
polymorphic forms of these
proteins can be distinguished by
different procedures
various types of electrophoresis
isoelectric focusing
 each form may show a characteristic
migration.
generalizations have emerged from
studies of plasma proteins
5. Each plasma protein has a characteristic
half-life in the circulation:
The half-lives of albumin and haptoglobin in
normal healthy adults are approximately 20 and
5 days, respectively.
 In certain diseases, the half-life of a protein
may be markedly altered.
Such as regional ileitis (Crohn disease), the
half-life of injected iodinated albumin in these
subjects may be reduced to as little as 1 day.

The half-life of a plasma protein can be determined
labeling the isolated pure protein with 131I under mild,
nondenaturing conditions
isotope unites covalently bound with
tyrosine residues in the protein.
 The labeled protein is freed of unbound 131I
and its specific activity (disintegrations per
minute per milligram of protein)
determined.

method

A known amount of the radioactive protein is injected
into a normal adult subject.

samples of blood are taken at various time intervals
for determinations of radioactivity.

The values for radioactivity are plotted against time,
and the half-life of the protein can be calculated from
the resulting graph ( the time for the radioactivity to
decline from its peak value to one-half of its peak
value ).

discounting the times for the injected protein to
equilibrate (mix ) in the blood and in the extravascular
spaces.
generalizations have emerged from
studies of plasma proteins
6. The levels of certain proteins in plasma
increase during acute inflammatory states
or secondary to certain types of tissue
damage
 acute phase proteins :
 C-reactive protein (CRP), α1-antitrypsin,
haptoglobin, α1-acid glycoprotein, and
fibrinogen.


The elevations of the levels of these
proteins vary from as little as 50% to as
much as 1000-fold in the case of CRP.
Their levels are also usually elevated
during chronic inflammatory states and in
patients with cancer.

These proteins are believed to play a role
in the body's response to inflammation.
For example:
C-reactive protein can stimulate the
classical complement pathway
 α1-antitrypsin can neutralize certain
proteases released during the acute
inflammatory state.
 Interleukin 1 (IL-1) is the principal
stimulator of the synthesis of the majority
of acute phase reactants by hepatocytes.

Some functions of plasma proteins.

Table 13-2 summarizes the functions of
many of the plasma proteins.

Various other protein hormones circulate
in the blood but are not usually designated
as plasma proteins.

Similarly, ferritin is also found in plasma in
small amounts, but it too is not usually
characterized as a plasma protein.
Ⅲ.3
Albumin is the major protein in
human plasma
Albumin
Molecular mass : 69 kDa

plasma :40%

3.4-4.7 g / dL



(60% of the total plasma protein)
extracellular space. 60%
The liver produces about 12 g of
albumin per day

representing :
25% of total hepatic protein synthesis
half its secreted protein.
Albumin is initially synthesized as a
preproprotein

Its signal peptide is removed as it passes into
the cisternae of the rough endoplasmic
reticulum

a hexapeptide at the resulting amino terminal is
subsequently cleaved off farther along the
secretory pathway.
The synthesis of albumin is
depressed in a variety of diseases
particularly diseases of the liver.
 The plasma of patients with liver disease
often shows a decrease in the ratio of
albumin to globulins (decreased albumin/
globulin ratio).
 The synthesis of albumin decreases
relatively early in conditions of protein
malnutrition, such as kwashiorkor.

Mature human albumin
 consists :
one polypeptide chain of 585 amino acids
and contains 17 disulfide bonds, three
domains
 Albumin has an ellipsoidal shape, it does
not increase the viscosity of the plasma
function of albumin
1.albumin is responsible for 75-80% of the
osmotic pressure of human plasma
 because of : low molecular mass

high concentration


2.bind various ligands : free fatty acids (FFA),
calcium, certain steroid hormones, bilirubin, and
some of the plasma tryptophan.
function of albumin

3. play an important role in transport of
copper in the human body.

4. A variety of drugs are bound to
albumin :sulfonamides, penicillin G, and
aspirin
Ⅲ.4 Haptoglobin
Haptoglobin (Hp)
plasma glycoprotein
 Human haptoglobin exists in three
polymorphic forms, known as Hp 1-1, Hp 2-1,
and Hp 2-2.
 The amount of haptoglobin in human plasma
ranges from 40 to 180 mg of hemoglobinbinding capacity per deciliter.

Haptoglobin binds
extracorpuscular hemoglobin,
preventing free hemoglobin from
entering the kidney
Haptoglobin (Hp) binds extracorpuscular
hemoglobin (Hb) in a tight noncovalent complex
(Hb-Hp).
 the Hb-Hp complex is too large to pass through
the glomerulus of kidneys.

The levels of haptoglobin in human
plasma vary and are of some
diagnostic use.
 Low
levels of haptoglobin: hemolytic
anemias
 Haptoglobin is an acute phase protein,
and its plasma level is elevated in a
variety of inflammatory states.
Certain other plasma proteins bind
heme
 Hemopexin
( β1-globulin) binds free
heme.
 Albumin bind some metheme (ferric
heme) to form methemalbumin
Section 2
Red blood cells
The mature erythrocytes are
devoid of
nucleus
intracellular organelles
I. Metabolic characteristics of
mature erythrocytes*
 1.it
can not synthesize nucleic
acid and proteins
 2. can not obtain energy by
oxidative phosphorylation
 3. glucose metabolism are :
glycolysis
the pentose-phosphate pathway
Ⅰ）Glycolysis
 utilizes
2ATP molecules
 produces 4ATP molecules
(substrate level
phosphorylation )
 net gain of 2ATP.
The ATP is used to

maintain the correct ion balance
 to
protect hemoglobin against
oxidative denaturation;
 to
protect against the formation of
methemoglobin
 synthesize
NAD+ and glutathion.
Ⅱ）The pathway of
2,3-diphosphoglycerate
(2,3-DPG)
1. Formation of 2,3-DPG
2. The role of 2,3DPG*
2,3-DPG can combine with
hemoglobin, causing a decrease
in affinity for oxygen.
Under conditions of oxygen lack

The released oxygen can be used by the
tissues.
Ⅲ）The role of pentose
phosphate pathway
 produce
the NADPH
 NADPH is essential for
the regeneration of
reduced glutathione
from oxidized
glutathione
Glutathione
The role of glutathione
 1.
The role in the destruction of
H2O2 in erythrocytes
 2. Reduction of methemoglobin
 3. Genetic abnormality-deficiency of
glucose-6-phosphate
dehydrogenase.
hemolytic anemia
Ⅱ.
Biosynthesis of
heme
Ⅰ）Precursors
Succinyl-coenzyme
A
Glycine
2+
Fe
Ⅱ）Synthetic
pathway
Synthetic
location:
mitochondria
cytoplasm
Synthetic process*
1. The formation of ALA (δaminolevulinic acid )
2. The formation of
porphobilinogen (PBG)
3. The formation of
uroporphyrinogen Ⅲ (UPGⅢ)
coproporphyrinogen Ⅲ (CPG) Ⅲ
4. The formation of heme.
1) ALA synthesis
 Synthetic
location:
mitochondria
 essential cofactor :
pyridoxal phosphate
(vitamin B-6)
Synthesis of ALA
rate-limiting reaction
ALA synthetase is the rate-limiting
enzyme
2) Production of
porphobilinogen(PBG)
 Synthetic
location:
cytoplasm
enzyme : ALA dehydratase SH
susceptible to heavy metals,
especially lead.
3)The formation of
uroporphyrinogen Ⅲ (UPGⅢ)
and coproporphyrinogen Ⅲ
(CPG Ⅲ )
Production of uroporphyrinogen
III
 Synthetic
location:
cytoplasm
the substrates :
four molecules of
porphobilinogen
Production of
coproporphyrinogen III
 Synthetic
location:
cytoplasm
 coproporphyrinogen
the mitochondria.
III back to
4)The formation of
heme
mitochondria
 incorporation of ferrums iron
into protoporphyrin
 reaction is catalyzed by
heme synthase or
ferrochelatase.
Heme biosynthesis:
in most mammalian cells except mature
erythrocytes.
85% in erythroid precursor cells in the
bone marrow
in hepatocytes.
Regulation of the synthesis of
heme
 ALA
synthase
Heme can markedly inhibite :
the synthesis ALA synthetase
the activity of ALA synthetase
A feedback mechanism controls heme
synthesis.
 some hormones and drugs also can result
in a marked increase of ALA synthetase
