MODES OF O2TRANSPORT

 Oxygen is transported from lungs to the tissues in two

methods:
 physically dissolved in the plasma, and,
 in combination with hemoglobin (Hb) in the red cell.
 Approximately 98% of the oxygen is transported in combination
with hemoglobin and the remaining 2% in the physically
dissolved form.
In Physically Dissolved Form
 In the physically dissolved form in plasma, only about 2% of oxygen is
transported. The amount of physically dissolved oxygen in the blood can
be predicted from Henry’s law.
 Henry’s law states that the amount of gas that dissolves in a liquid at a
given temperature is proportional to the partial pressure of the gas.
 Thus, the quantity of dissolved oxygen in arterial blood is calculated from
the following equation:
 Dissolved O2 (ml/dl) = oxygen solubility  partial pressure of O2 in arterial

blood (PaO2).

 = 0.003 (ml /dl of blood per mm Hg)  95 mm Hg (normal PaO2 is 95 mm

Hg)
 = 0.3 ml/dl of blood
 Thus, at PaO2 of 95 mm Hg, the dissolved O2 is 0.3 ml/dl.

That means, in a normal healthy adult, 0.3 ml of O2 is transported in

dissolved form in 100 ml of blood (Application Box 70.1).
 As cardiac output is 5 liters /min at rest, oxygen transported in dissolved
form at rest is about 15 ml/min.
 Therefore, oxygen transported in dissolved form alone is grossly
inadequate to meet the oxygen requirement of the body, which is about
250 ml/min at rest.
In Combination with Hemoglobin
 More than 98% of O2 is transported in combination with Hb.
 This becomes possible due to the binding affinity of hemoglobin for
oxygen.
 Hemoglobin that binds with oxygen is called oxyhemoglobin (HbO2) and

the hemoglobin that does not bind with O2 is called deoxyhemoglobin

(deoxy-Hb) or reduced-hemoglobin.

Oxyhemoglobin Formation
 The hemoglobin molecule is a protein made up of four subunits that are
bound together. Each subunit consists of a molecular group known as
heme and a polypeptide attached to the heme. The four polypeptides of a
Hb molecule are combinely known as globin.
◦ Each heme group contains one atom of iron (Fe++) to which oxygen binds
Structure of Hb molecule
 ◦ Since each iron atom can bind one molecule of oxygen, a single Hb molecule
can bind four molecules of oxygen.

◦ Hb binds with oxygen only when the iron is in ferrous (Fe++) state. The Fe++
iron in Hb is oxidized to ferric (Fe+++) iron to form methemoglobin.

◦ Thus, methemoglobin can’t bind oxygen. Methemoglobin formation occurs

under the influence of various compounds like nitrites or sulfonamides.
Methemoglobin is also formed spontaneously.

◦ However, the enzyme methemoglobin reductase is present in red cell that

reduces methemoglobin to Hb. Therefore, normally only about 1.5% of total
Hb is methemoglobin.

◦ Deficiency of methemoglobin reductase (a genetic defect) increases the

methemoglobin concentration and decreases oxygen carrying capacity.
 Oxygen binds rapidly and reversibly to hemoglobin to form

oxyhemoglobin(HbO2): O2 + Hb  HbO2
 R & T STATES OF Hb:During this process of HbO2 formation, the heme remains in

ferrous state. Thus, this process is oxygenation, not oxidation. As there are four

subunits in Hb molecule, Hb reacts rapidly (in less than 0.01 s) with four molecules

of oxygen to form HbO8.

1. When oxygen is not bound with Hb (deoxyhemoglobin) the four subunits of Hb are

tightly bound with each other. This configuration of Hb is called Tense or T state. In this

T configuration, affinity of Hb for oxygen is less.

2. When, first oxygen molecule binds with Hb, the four subunits enter into the relaxed or R

state that exposes more oxygen binding sites. Therefore, in R configuration, affinity of

Hb for oxygen is greatly increased by 200 to 500 times.

3. When PO2 is very less, most of Hb molecules are in T state and have low oxygen

affinity.

4. When PO2 is very high, the Hb molecules are in R state and have high oxygen affinity
 Conversion of Hb molecule from T state to R state. As O2 is added, salt bridges are
successively broken and finally 2-3,BPG is expelled.
T (taught) conformation of deoxy-Hb is changed to relaxed (R) conformation of Oxy-Hb

5. Thus, the quantity of oxyhemoglobin formation is a function of partial

pressure of oxygen in the blood.

• In pulmonary capillaries, where PO2 is high, the reaction favours to form more

oxyhemoglobin, and in tissue capillaries, where PO2 is low, the reaction favours

formation of deoxyhemoglobin that helps oxygen to be unloaded from hemoglobin

and becomes available to the cells.
O2 Carrying Capacity of Hb
 Each gram of hemoglobin binds with 1.34 ml of oxygen. The maximum
amount of oxygen that can be carried by hemoglobin is called the
oxygen carrying capacity.
 In a healthy individual, the oxygen carrying capacity of arterial blood is
about 20 ml of O2 per 100 ml of blood, considering the hemoglobin
concentration of 15 g% (1.34 ml  15 g = 20.1 ml O2/dl blood).
 OXYGEN CONTENTof Hb (HbO2 content) is the amount of oxygen
actually bound to hemoglobin, whereas OXYGEN CAPACITY of Hb
(HbO2 capacity) is the amount of oxygen that can potentially bind with
Hb.
 The percentage saturation of hemoglobin with oxygen (SO2) is the ratio
of the quantity of oxygen actually bound (oxygen content) to the quantity
that can be potentially bound (oxygen capacity) multiplied by 100.
Oxygen-Hemoglobin Dissociation Curve:
 Oxygen-hemoglobin (Oxy-Hb) equilibrium curve (dissociation or
association curve) explains the relationship between partial pressure of
oxygen (PO2) in blood with oxygen saturation of Hb. Oxy-Hb equilibrium

curve is an S-shaped curve over the range of PO2 from 0 to 100 mm Hg

 The sigmoid shape of the curve results from hemoglobin affinity for oxygen
at various PO2 levels.

 As PO2 rises, the Hb saturation progressively increases.

 However, the saturation is not linear with increase in PO2 for which the
curve becomes sigmoid shaped.
 The S-shaped oxyhemoglobin equilibrium curve enables oxygen to saturate
hemoglobin under high partial pressures in the lungs and to give up large
amounts of oxygen with small changes in PO2 at the tissue level.
 Oxygen-hemoglobin dissociation curve
Note at PO2 of 27 mm Hg, Hb saturation of oxygen is 50% (P50)
 The curve is divided into two major phases: the steep phase
and the plateau phase.
The Steep Phase
 The curve has a steep slopebetween PO2 of 10 and 60 mm Hg.
During this phase of the curve, combination of oxygen with Hb
increases very rapidly as the PO2 increases from 10 to 60 mm Hg.

Oxygen saturation of Hb is about 90% at PO2 60 mm Hg.

Significance of Steep Phase

 In the steep phase, oxygen saturation of Hb is very high.
 Less increase in PO2 leads to greater percentage saturation of Hb

and therefore, facilitates oxygen loading.

 Also, change in steep portion of the curve in reverse direction (that is,
from 60 to 10 mm Hg) causes unloading of oxygen in the tissues.
 A small decrease in PO2 in the tissue results inunloading of large
amount of oxygento the tissues.
 The steep phase allows large quantities of oxygen to be released from
hemoglobin in the tissue capillaries where a lower capillary PO 2 prevails.
This is especially achieved by shifting the curve to right (shift occurs
mainly on the steep phase) by increased H+ or by increased CO2.

The Plateau Phase

 The curve begins to plateau at PO2 around 60 mm Hg and flattens at PO2
of 70 mmHg
 The increase in PO2 above 60 mm Hg produces only a small increase
in oxygen binding.
 Increase in PO2 from 60 to 100 mm Hg in the plateau region of the curve
illustrates that oxygen saturation and content remain apparently
constant over a wide alteration in alveolar PO2.

Significance of Plateau Phase

 There are two significances of plateau phase:

1. In different situations, like at high altitude or in pulmonary diseases in

which a moderate hypoxia (decrease in PO2 from 95 to 60 mm Hg)
occurs, the total amount of oxygen carried by Hb decreases only by
5−10%, since Hb saturation is about 90% at PO2 of 60 mm Hg.

Thus, plateau in the curve provides a safety factor through

which even a significant decrease in lung function can allow normal
saturation of Hb.
2. Oxygen saturation and content remain fairly constant inspite of wide
fluctuations in alveolar PO2(PAO2).

◦ For example, if PAO2 rises from 100 to 120 mm Hg, hemoglobin saturation
increases only slightly (97 to 98%).
◦ This is the reason why oxygen content cannot be raised appreciably by
hyperventilation or by breathing 100% oxygen because Hb is already completely
saturated with oxygen at PO2 of 100 mm Hg. This is true only for normal people
at sea level.

◦ If a person has low arterial PO2 due to lung disease or for his ascension to
high altitude, hyperventilation or breathing 100% oxygen increases Hb
saturation with oxygen as they have more deoxy-Hb initially.
The P50
 The P50 is the level of PO2 at which 50% of the hemoglobin is
saturated with oxygen.
1. P50 assesses the binding affinity of hemoglobin for oxygen.

2. In adults at sea level, the normal P50 occurs at a PO2 of 27 mm Hg.

3. Alteration in the P50 value has a greater impact on the steep phase
of the curve.
4. If the P50 is high, it signifies the decrease in affinity of Hb for
oxygen, which is seen in right-shift of the oxygen-hemoglobin
equilibrium curve.
5. Conversely, if P50 is low, it signifies shift of the curve to the left, in
which affinity of Hb for oxygen is more
Effect of P50 on Oxy-Hb dissociation curve.
Factors Affecting Hb-Binding Affinity with Oxygen

 Several factors influence the affinity of hemoglobin for oxygen.

 The important factors are temperature, arterial PCO2, arterial pH

and 2,3-DPG. A rise in PCO2, a fall in pH, and a rise in temperature

all shift the curve to the right.
 The effect of carbon dioxide and hydrogen ions on the affinity of
hemoglobin for oxygen is known as the Bohr effect.
 A shift of the oxyhemoglobin equilibrium curve to the right is
physiologically advantageousat the tissue level, as affinity of O2

for Hb is lowered (increased P50).

 A rightward shift enhances the unloading of oxygen for a given PO2 in
the tissue, and a leftward shift increases the affinity of hemoglobin for
oxygen, thereby lowering the ability to release oxygen to the tissues.
 A simple way to remember the functional importance of these shifts is that
an exercising muscle is warm and acidic and produces large amounts of
carbon dioxide (high PCO2), all of which favor the unloading of more
oxygen to metabolically active muscles.

Temperature
 Increase in temperature shifts the Oxy-Hb dissociation curve to right
and decrease in temperature shifts the curve to the left.
 In other words, high temperature decreases the affinity of Hb for oxygen.
 This helps in release of oxygen in metabolically active tissues in which
temperature is more.
Effect of temperature on Oxy-Hb dissociation curve
pH
 Alteration in blood pH shifts the Oxy-Hb dissociation curve. Christian Bohr
and Neils Bohr in 1904 demonstrated that respiratory acidosis shifts oxy-
Hb dissociation curve to right. Since then, the decrease in oxygen affinity
in acidosis is known as Bohr effect.
 A decrease in pH shifts the curve to right and increase in pH shifts the
curve to left.
 When blood passes through capillaries, CO2 enters red cell that
decreases intracellular pH and shifts the Oxy-Hb dissociation curve to
right.
 It has been noted that under normal physiological conditions, binding of
about 0.7 mole of H+ causes Hb to release 1 mole of oxygen.
 Thus, when blood passes through tissue capillaries, the acidic
environmentfacilitates release of oxygen from Hb into the tissues.
Effect of pH on Oxy-Hb dissociation curve
Carbon Dioxide
 During cellular metabolism CO2 is released into circulation that increases
generation of H+ and decreases pH. This shifts the curve to right. This helps in
release of oxygen from Hb.
◦ The shift to right in acidosis (Bohr effect) is partly due to effect of decrease in
pH and partly to the direct effect of CO2 on Hb.

◦ CO2 combines with unprotonated amino groups on Hb (Hb-NH2) to form

carbamino groups. The formation of carbamino group causes negative shift in
the charge on one amino acid side chain that changes the conformation of Hb.
This results in decrease in oxygen affinity of Hb.
◦ In metabolically active tissues, metabolic vasodilation increases blood flow to
the tissue increasing the oxygen transport to the area.
◦ In the area, already persisting local hypercapnia and acidosis shift the curve to
the right (decrease oxygen affinity for Hb) that facilitates oxygen delivery to
the tissue
2,3-DPG
 Red blood cells lack mitochondria. Therefore, anerobic glycolysis occurs in
red cells. During glycolysis, large quantities of 2,3-diphosphoglycerate
(2,3-DPG), an organic phosphate compound is produced as the metabolic
intermediary. This compound is called 2,3-biphosphoglycerate (2,3-BPG).
Thus, 2,3-DPG level is much higher in red cells than in other cells.

◦ 2,3-DPG significantly affects affinity of hemoglobin for oxygen.

◦ In red cells, concentration of 2,3-DPG is almost same as that of Hb. In
fact, 2,3-DPG binds to Hb in 1:1 stoichiometry, interacting with a
central cavity formed by two β chains.
◦ 2,3-DPG has about 3.5 negative charges that interact with eight
positively charged amino acids in the central cavity. This destabilizes
the interaction of Hb with oxygen.
 ◦ Thus, binding of 2,3-DPG with Hb shifts the Oxy-Hb dissociation curve to
right and facilitates the release of oxygen.
◦ At the tissue level, an increase in 2,3-DPG facilitates unloading of oxygen
from the red cell.
◦ Hypoxia stimulates glycolysis resulting in increased production of 2,3-DPG.
◦ Red cell 2,3-DPG increases in anemia, exercise and hypoxic conditions like
high altitude, chronic lung disease etc. Therefore, in these conditions, Oxy-Hb
dissociation curve shifts to right

Factors that affect 2,3-DPG in Red Cells:

1. pH: Acidosis inhibits red cell glycolysis and therefore decreases 2,3-DPG
concentration.

2. Type of Hb: The γ chains of fetal Hb have less avidity for 2,3-DPG than β
chains of adult Hb. Therefore, fetal Hb has higher oxygen affinity. This
provides an advantage to fetus to extract oxygen from maternal blood in
Effect of 2,3-diphosphoglycerate (2,3-DPG) on Oxy-Hb dissociation curve
3. Fetal Hb shifts Oxy-Hb dissociation curve to left.

4. Hormones: Growth hormone, thyroxine and testosterone stimulate the

synthesis of 2,3-DPG.

5. Altitude: At high altitude, 2,3-DPG concentration increases

substantially in red cells. This increases the availability of oxygen to
the tissues.

6. PO2: Hypoxia increases 2,3-DPG production.

7. Procedure of storage of blood: The 2,3-DPG concentration in red

cells of blood stored in blood bank decreases. Therefore, transfusion of
stored blood decreases oxygen transport, especially when transfused
into hypoxic patients. However, if blood is stored in citrate-dextrose-
phosphate solution this effect is less than when stored in acid-citrate-
dextrose solution
Myoglobin
 Myoglobin is the Hb pigment in muscle.
 Though it is similar to Hb in structure, myoglobin binds one molecule of O 2
per mole.
 It shifts Oxy-Hb dissociation curve to left and the curve becomes
hyperbolic (loses sigmoid shape) (Fig. 70.8).
 This helps in picking up of O2 from blood.
 Myoglobin is more in regularly exercising muscles especially that are
trained in isometric exercise. This helps the muscle to draw oxygen from
myoglobin, especially when blood flow ceases for a longer duration during
sustained muscle contraction.
 Myoglobin is also believed to facilitate transfer of O2from blood to
mitochondria.
Effects of myoglobin and fetal hemoglobin (Hb) on Oxy-Hb dissociation curve
Effect of Carbon Monoxide (CO)
 The binding affinity of CO with Hb is 210 times more than oxygen.
Moreover, CO interferes with oxygen transport by competing for the
same binding sites on hemoglobin.
 CO binds to hemoglobin to form carboxyhemoglobin (HbCO).
 When the blood is 60% saturated with CO (carboxyhemoglobin) the
oxygen content is reduced to less than 10 ml/dl.
 As the partial pressure of CO approaches 1 mm Hg, Hb is fully
saturated with CO. However, arterial PO2 may be normal as oxygen
diffusion gradient remains normal.
 But,oxygen content is greatly reduced as it can’t bind to Hb. This
grossly decreases the oxygen carrying capacity.
 Moreover, CO also shifts the Oxy-Hb dissociation curve to the left, which
decreases oxygen release to the tissues.
 Therefore, severe tissue hypoxia occurs in CO poisoning, which is fatal if not
treated immediately (for details refer ‘Hypoxia’).
 Normally, in healthy individuals, CO occupies 1–2% of Hb binding sites.
 However in chronic cigarette smokers, traffic personnel and in residents of
crowded traffic areas, CO concentration increases in plasma and CO occupies
about 10% of Hb.
Factors that Shift Oxy-Hb Dissociation Curve

Factors that shift the curve to right:

 Increased temperature
 Decreased pH
 Increased PCO2
 Increased 2,3-DPG
 Hypoxia
 Shift of the curve to right increases P50 (decreased affinity
of Hb for oxygen), which facilitates oxygen release.
Factors that shift the curve to left:
 Decreased temperature
 Increased pH
 Decreased PCO2
 Decreased 2,3-DPG
 Fetal Hb
 Carbon monoxide
 Shift of the curve to left decreases P50 (increases affinity of
Hb for oxygen) that facilitates oxygen uptake and prevents
oxygen release.
O2 DELIVERY TO TISSUE

Diffusion of O2 from Capillaries to Tissue Fluid

 When the arterial bloodreaches the peripheral tissues, diffusion of
oxygen occurs from the peripheral
 Capillary blood into the tissue fluid.
 This occurs along the concentration gradient of oxygen from area of
higher level to lower level. The PO2 in systemic capillary blood is95

mm Hg, whereas PO2 in the interstitial fluidthat surrounds the tissue

cells is about40 mmHg.
 Thus, there is a great pressure gradient due to considerably large
pressure differenceof O2 between the systemic capillary blood and the
tissue fluid that causes oxygento diffuse rapidly fromthe capillary blood
into the interstitial fluid.
 Therefore, PO2 falls rapidly in the capillaryto about 40 mm Hg.

Thus, the PO2 of the blood leaving the tissuecapillaries (PO2 at

venous end of capillary) and entering the systemic veins is 40 mm
Hg
 From tissue fluid, oxygen rapidly enters the cell as in the intracellular
fluid thePO2is about 23 mmHg (ranges between 7 to 40 mmHg), as
oxygen is constantly utilized by cell. Especially in metabolically
active tissues, this gradient is more as mitochondrial oxygen
utilization is more.
 Diffusion of oxygen to tissues depend on two important factors:
i)rate of blood flow to the tissue and ii) the rate of tissue metabolism.
Oxygen delivery is more if the blood flow is more and the metabolic
activity is more
Delivery of oxygen from capillary blood to tissue.
Bohr Effect
 Any factor that shifts Oxy-Hb dissociation curve to right,decreases
affinity of oxygen for Hb.
1. At the tissue level, CO2 enters the blood and shifts the curve to right. This
helps in unloading of oxygen from Hb and facilitates tissue oxygenation.
This phenomenon is called Bohr Effect as was initially described by Christian
Bohr (1855-1911), who has showed experimentally the influence of CO2
tension on blood binding of oxygen.

2. This occurs mainly due to decreased pH in the tissue.

3. Increased temperature of blood in the tissue also contributes.

4. In tissue, acidification of blood decreases affinity of O 2 to Hb, as deoxygenated

Hb binds H+more actively than that of oxyhemoglobin.

5. The PCO2 level in the tissue rises, the curve shifts to right and P 50 rises. Thus,

Bohr’s effect helps in unloading of O2 and loading of CO2.

Importance of Oxygen Saturation and Content

Oxygen Saturation
 The Oxygen saturation is the ratio of the amount of oxygen bound to
Hb to the maximum amount of oxygen that can bind Hb (100%
oxygen capacity). At 100% oxygen capacity, heme groups of Hb are fully
saturated with oxygen.

Oxygen Content
 The oxygen content of blood is the volume of oxygen contained in
unit volume of blood, which includes the oxygen bound to Hb and also
dissolved in plasma. As the dissolved oxygen is negligible, the oxygen
content depends on concentration of Hb and oxygen binding capacity of
Hb
Oxygen Extraction
Oxygen extraction is the amount of oxygen taken up by the tissues from the
blood.
This is an index of oxygen consumption of the tissue. It is better quantified in
terms of OXYGEN EXTRACTION RATIO (OER).
OER is also called as oxygen coefficient ratio, is the amount of oxygen
extracted by the tissue divided by the amount of oxygen delivered.
In metabolically more active tissues like cardiac muscle, OER is as high as 85%
at rest.
APPLIED ASPECTS

Measurement of O2 Saturation of Hb

Pulse Oximetry
 Pulse oximetry is a noninvasive method of measurement of
oxygen saturation of Hb. Oxygen saturation is continuously
measured in hospitalized patients, especially patients in intensive
care unit by this method. Instrument used is the pulse oximeter.
◦ The probe of the oximeter is attached usually to the finger-tip or ear
lobule where the pulsating blood vessels are accessible externally.
Red and infrared light are transmitted through the vascular bed, and
pulsatile, nonpulsatile and total absorbances are calculated .
 ◦ The pulsatile component of absorbance represents the
arterial oxygenated blood and the nonpulsatile component
represents the deoxygenated capillary and venous blood.
Abnormalities of Tissue Oxygenation
Less Oxygenation
 Less oxygenation of tissue could be due to less oxygen in the
atmosphere decreasing oxygen content in blood, decreased
Hb level in blood decreasing oxygen saturation, or decreased
capacity of tissue to utilize oxygen resulting in decreased
oxygen extraction.
Excess Oxygenation & Metabolism
Pulmonary Damage by Free Radicals
 Though tissue oxygenation is essential for life, excess or
inappropriate oxygenation and oxygen metabolismis harmful for
the tissues.
 During synthesis of ATP, molecular oxygen is reduced to form
water in mitochondria.
 The reduction of oxygen is accomplished by addition of four
electrons by the mitochondrial electron transport system.
 However, leak in the electron transfer system allows oxygen to
accept less than four electrons that form free radicals.
 Free radicals cause damage to the tissues. Lung is frequently
damaged by free radicals.
 Pulmonary capillaries are mainly damaged that results in
pulmonary edema.
Reactive Oxygen Species & Antioxidants
 A free radical is an atom or a molecule with an unpaired electron in
its outermost orbit.
 Superoxide radical (O2−) and hydroxyl radical (OH∙) are
commonly produced free radicals in the body. Hydrogen peroxide
also can generate hydroxyl radical.
 Superoxide ion reacts with NO to form peroxynitrite, which is also
a free radical. These free radicals are combinely known as reactive
oxygen species (ROS). ROS cause tissue damage and promote
tissue degeneration. They are sometimes called pro-oxidants, as
they are produced during the process of tissue oxidation.
 However, there are antioxidants in the body that prevent the body from oxidative
damage. Antioxidants are mainly enzymes such as superoxide dismutase,
catalase and peroxidases that neutralize ROS. Imbalance between ROS and
antioxidants by either more production of ROS or decreased formation of
protective enzymes results in oxidative stress (oxidative tissue damage).
 ROS are also produced during inflammations, which occurs mainly due to
respiratory burst of neutrophils. Reperfusion injury deteriorates condition by
generating local oxidative stress