Oxygen moves into our cells and CO₂ moves out by the process of diffusion. In Chapter 6 we learned that there must be a difference in concentration of the diffusing substance for overall diffusion to take place. Outside the cell must have a higher concentration of O₂ and a lower concentration of CO₂ than inside the cell. High concentrations of O₂ and low concentrations of CO₂ are the conditions found in the air around us and, to a lesser extent, in the alveolar air. It is the business of the circulation to bring these conditions close to the individual cells.

Blood is a liquid tissue (plasma) containing formed elements (cells). The red blood cells (RBC, erythrocytes) play an important part in the transport of O₂ to and CO₂ away from the tissues. RBC should strictly be called corpuscles, because they contain no nucleus, but the term cell is in general use.

This exchange of gases between the cells and blood at our tissues is a repeat of the exchange between air and blood at the lungs, and results in the differences in composition between venous and arterial blood shown in Table 8.1. Although the RBCs play an important role in this carriage and exchange (carrying the majority of O₂ and processing CO₂ at both lungs and tissues), the gases must first enter into simple solution in the plasma before being carried or processed by the RBCs.

Table 8.1

Some values for normal blood

Oxygen in the blood is mainly carried in loose combination with haemoglobin (Hb) within the RBC. Carbon dioxide is carried partly in solution, partly in combination with proteins (particularly Hb), but mainly as bicarbonate in the plasma. The carrier mechanisms in blood have evolved so that the uptake or loss of O₂ promotes the loss or uptake of CO₂ and vice versa, a most useful arrangement.

The addition of CO₂ to the blood at the tissues would cause a dangerously large change in acidity if the blood did not contain efficient buffering systems, in particular protein, bicarbonate and phosphate, which take up hydrogen ions added to the blood and release them when the blood becomes more alkaline, thereby buffering (resisting) changes in acidity.

Oxygen transport

Haemoglobin (Hb)

Because we are large and complicated animals most of the cells of our bodies are far removed from the atmosphere. Oxygen has therefore to be carried from the lungs to these cells in the blood. All gases dissolve in water to a greater or lesser extent, but O₂ dissolves only to the extent of 3 mL L⁻¹ in the watery plasma leaving the lungs.

While exercising vigorously we may need up to 3 L of O₂ per minute. This implies that if O₂ was only carried to the tissues in simple solution we would need a blood flow of 1000 L min⁻¹ to supply our bodies with O₂. Olympic athletes can increase the output of their hearts to about 30 L min⁻¹ which you can see is still several hundred times too little to supply their tissues with O₂. The answer to this problem is that, like all other vertebrates, we have evolved a carrier molecule in the blood which picks up and then releases a great deal of O₂. In us this molecule is haemoglobin (Hb).

Case 8.1 Carriage of gases by the blood and acid/base balance: 1

Carbon monoxide poisoning – a failure of oxygen transport

Mr Jones is a pensioner who lives alone. One winter’s evening, he turned on his gas fire and settled down to watch the television. The gas fire was very old indeed, and had never been serviced, as far as Mr Jones could remember. As the evening wore on, Mr Jones became increasingly drowsy. Eventually, he fell deeply asleep. Later that evening, Mr Jones’ daughter called round to see him. She found her father unconscious on the sofa and called an ambulance. On his arrival at the Accident and Emergency department an arterial blood sample was taken from Mr Jones for analysis of blood gas tensions. The sample revealed that his arterial blood gases were close to normal. However, his carboxyhaemoglobin level was measured at 52%, confirming the diagnosis of carbon monoxide (CO) poisoning.

He was treated with high concentrations of oxygen and was closely monitored on the high-dependency ward. Gradually, he regained consciousness as the CO was displaced from his blood by oxygen and the effects of the alcohol he had consumed began to wear off. The carboxyhaemoglobin levels in his blood were measured later that day and were found to be 12%.

In this chapter, we will consider:

1. Why carbon monoxide is poisonous.

2. Symptoms of carbon monoxide poisoning.

3. The treatment of carbon monoxide poisoning.

Haemoglobin has remarkable O₂-carrying properties which are related to its molecular structure (Fig. 8.1).

Fig. 8.1 The structure of haemoglobin. Each of the four globin chains (the wormlike structures in the figure) is made up of a spiral of just over 100 amino acids. Each chain is attached at one point to an iron-containing haem group. Each haem group can carry a molecule of O₂, so each haemoglobin molecule has four ‘hooks’, each of which can carry one O₂.

Each haemoglobin molecule consists of a protein (globin) and haem (protoporphyrin and ferrous iron). The globin is made up of four polypeptide chains, each carrying a haem group. This structure is repeated four times in each molecule, which means there are four sites, each capable of carrying one O₂ on each Hb molecule. This explains many of the properties of Hb, as we will see in a moment. Each of the four chains can vary in a way which will vary the O₂-carrying properties of the blood. The chains are described chemically as α or β, depending on their structure. The types of polypeptides that make them up give rise to the various forms of normal and abnormal Hb.

Adult Hb consists of two α and two β chains, with 141 and 146 amino acid residues per chain, respectively. Each Hb molecule thus has 574 amino acids and four haems, which gives the molecule a weight of about 64 500. Fetal Hb is slightly, but importantly, different. Men have about 150 g L⁻¹ Hb in their blood, women about 130 g L⁻¹.

Oxygen combination with haemoglobin

This reversible reaction can be summarized as follows:

(Equation 8.3)

which will be driven to the right (to HbO₂) by increased Po₂ and to the left by low Po₂. The Hb of this equation is deoxyhaemoglobin – often, and incorrectly, referred to as ‘reduced haemoglobin’, despite the fact that the Hb is not chemically reduced. The HbO₂ in this equation is oxyhaemoglobin, and by the same token that Hb is not chemically oxidized, the combination between Hb and O₂ is oxygenation, a much looser combination than oxidation.

Case 8.1 Carriage of gases by the blood and acid/base balance: 2

Why carbon monoxide is poisonous

Carbon monoxide (CO) is a colourless, odourless, tasteless gas formed from the incomplete combustion of organic substances. Poisoning with CO may be the result of inhalation of fumes from a faulty heating system, or the inhalation of smoke from house fires.

CO produces its toxic effect by causing hypoxia in peripheral tissues. It does this by interfering with the transport of oxygen by haemoglobin, and also by interfering with cellular respiration.

CO binds to haemoglobin with an affinity about 250 times that of oxygen, to form carboxyhaemoglobin. Carboxyhae-moglobin cannot carry oxygen, and so the amount of haemoglobin available for the carriage of oxygen is reduced. This means that even if the partial pressure of oxygen in arterial blood is normal and therefore the oxygen saturation of the available haemoglobin is maintained, the oxygen content of the blood is reduced because the amount of haemoglobin available to carry oxygen is reduced. Furthermore, in the presence of carboxyhaemoglobin the oxygen–haemoglobin dissociation curve is distorted and shifted to the left. As a result of this, oxygen tends to remain bound to haemoglobin in the peripheral circulation rather than being released into the tissues. In other words, less oxygen is carried in the circulation, and of the oxygen that is carried, much remains bound to haemoglobin and is not available for tissue respiration.

In addition to these effects on oxygen carriage by the blood, CO also interferes with electron transport in mitochondria and with other cellular processes. Overall, then, its effect is to reduce oxygen delivery to peripheral tissues, and also to reduce the ability of these tissues to use what oxygen they obtain. Cellular ATP levels fall and vital cellular functions begin to fail. In severe cases of poisoning this may lead to impairment of the respiratory and cardiovascular systems, which exacerbates oxygen transport even more.

Each of the four haem groups of the Hb molecule represents a site for combination with O₂. It might be more correct to consider each haemoglobin molecule as Hb4, with which association or dissociation with O₂ takes place in four steps, in which case Equation 8.3 should be written:

(Equation 8.4)

It is conceptually useful to consider each Hb molecule as having only four ‘hooks’. On each hook can hang one O₂.

The haem and the globin of each molecule are held in a fixed relationship to each other by links (salt bridges) between the polypeptide chains. In each of the steps in Equation 8.4, when a molecule of O₂ binds to the iron atom in each haem the molecular shape is distorted, making the attachment of the next O₂ molecule easier. This distortion is called an allosteric effect and, together with the fact that there are only four ‘hooks’ for O₂ per molecule, explains the sigmoid shape of the graph obtained when we plot percentage saturation of Hb by O₂ against Po₂ (Fig. 8.2). This S-shaped curve is called the oxyhaemoglobin dissociation curve, and it is so important to our understanding of the transport of O₂ that a description of how it is obtained is well worthwhile.

Fig. 8.2 The oxyhaemoglobin dissociation curve. This curve is obtained by exposing blood to a number of partial pressures of O₂ and measuring how much of its total O₂ carrying capacity is occupied. This can be expressed in two ways. The saturation curve (A) represents the percentage of the total number of ‘hooks’ for O₂ that are occupied (the 50% occupancy, P₅₀, is shown). This curve gives us no idea of the amount of O₂ being carried, which depends on the amount of Hb present and is shown by the content curve (B). That content depends on the amount of Hb as shown by the lower curve, which represents anaemic blood. Both this and the normal curve are equally saturated at different Po₂, as shown by their identical P₅₀, but the amount they are carrying is very different. Because temperature and pH affect the properties of Hb we must state that these curves were obtained at pH 7.4 and 37°C. The important loading and unloading conditions for blood are from arterial blood at (a) and into mixed venous blood at (). Note that the slope of the curve is very different at these two points.

Obtaining a dissociation curve

If you take, say, five test tubes of blood and expose each of them to a different partial pressure of O₂ (say 0, 2, 4, 8, 16 kPa O₂, as in Fig. 8.2), in each tube a different percentage of haemoglobin will be converted to oxyhaemoglobin, depending on the partial pressure it has been exposed to. Each sample will have a different colour because oxyhaemoglobin is brighter red than haemoglobin (arterial blood is red; venous blood is purple). An instrument called a spectrophotometer can use this colour to measure what percentage of the Hb has been converted to HbO₂, and so we can plot a graph of percentage saturation (percentage of the O₂-carrying ‘hooks’ occupied) against the Po₂ to which that particular sample of blood was exposed (Fig. 8.2A). We have talked about each Hb molecule having four hooks, each of which can carry one O₂ molecule. This might suggest that blood can only be 25% (one hook), 50% (two hooks) 75% (three hooks occupied) or 100% (four hooks). This is true for each individual molecule, but would ignore the fact that even a drop of blood contains millions of Hb molecules, any one of which can be carrying from zero to four O₂ molecules.

Properties of the oxyhaemoglobin dissociation curve

When 100% saturated (all ‘hooks’ occupied), 1 g of Hb carries about 2 mg – 1.36 ml – of O₂ at normal body temperature. Therefore, 1 L of your blood, containing 150 g of Hb, can transport 200 ml of O₂ as HbO₂. Comparing this with the 3 ml carried in simple solution gives some idea of the advantage of having Hb in our blood.

The carriage of O₂ in our blood is not quite as simple as hanging O₂ molecules on hooks, like coats on a stand: evolution has refined this already efficient process even further. To understand these refinements requires the definition of four terms:

• Oxygen tension (Po₂; kPa). We have met this term before, but revision of its meaning might be useful. Oxygen tension is sometimes called the partial pressure of O₂ in solution. The difference in Po₂ between two sites determines the rate and direction of diffusion of O₂. This is because the partial pressures correspond to the concentrations in solution (Henry’s Law). Thus dissolved O₂ will diffuse down its concentration (partial pressure) gradient. The Po₂ of active skeletal muscle may be as low as 1 kPa. Arterial blood supplying that muscle has a Po₂ of about 13 kPa, and this large pressure difference ‘pushes’ O₂ strongly into the tissues.

Case 8.1 Carriage of gases by the blood and acid/base balance: 3

Signs and symptoms and diagnosis of carbon monoxide poisoning

On arrival at the Accident and Emergency department, Mr Jones was more awake, but was drowsy, with slurred speech. An arterial blood sample was taken for analysis of blood gas pressures. The sample revealed that the partial pressure of carbon dioxide in Mr Jones’ arterial blood was high, suggesting that he was hypoventilating. The partial pressure of oxygen in his blood was slightly higher than normal because he was breathing oxygen. However, his carboxyhaemoglobin level was measured at 52%, confirming the diagnosis of carbon monoxide poisoning.

Measurement of carboxyhaemoglobin levels is the key to diagnosing CO poisoning. The carboxyhaemoglobin is usually expressed as a percentage of total haemoglobin. In city dwellers up to 5% carboxyhaemoglobin is normal, and levels of up to 10% are very often found in smokers. In cases of CO poisoning levels of over 20% may be associated with symptoms, and levels in excess of 60% often result in coma. However, although measuring carboxyhaemoglobin levels is useful in diagnosing CO poisoning, it is often difficult to relate these measurements to the patient’s clinical condition. This is because once the patient is removed from the source of CO blood levels fall very quickly, and the clinical picture is thought to be more closely related to peak carboxyhaemoglobin levels.

The signs and symptoms of CO poisoning are all related to the tissue hypoxia that it causes. Many organs, but particularly the brain, the heart and the lungs, can be affected.

In mild cases of CO poisoning patients develop a frontal headache that may be associated with drowsiness or agitation and confusion, particularly in the elderly. Often these symptoms are associated with nausea or vomiting. In more severe cases of poisoning, patients may lose consciousness or start to fit.

CO may also affect the heart and cause ECG abnormalities, and is recognized as a cause of cardiac failure and myocardial infarction. Its effects on the lungs include hyperventilation and pulmonary oedema.

It may affect the nervous system, producing hemiplegia or peripheral nerve damage, and recovery from poisoning may be complicated by long-term psychiatric problems such as personality changes and memory loss, which are thought to be a consequence of hypoxic brain damage.

Classically, patients with CO poisoning are said to have pink skin and mucosae owing to the presence of carboxyhaemoglobin, which has a bright red colour. This is not a reliable sign, and in severe cases of CO poisoning associated with a failing circulation it may not be apparent.

Summary 1

• O₂ is used as a proton (H⁺) acceptor in oxidative metabolism.

• The presence of haemoglobin improves the O₂ carrying capacity of blood 100 times.

• There are four sites on the haemoglobin molecule, each of which carries one O₂ molecule.

• Each site has a different affinity for O₂ which causes the O₂–haemoglobin dissociation curve to have an S-shape.

• Haemoglobin content (Hb, g L⁻¹). It is Hb that has the ‘hooks’ that carry the O₂. The number of ‘hooks’ determines the maximum O₂-carrying capacity per mL of blood. If blood has only 50% (say) of the normal amount of Hb (it is anaemic), it will only have 50% of the normal number of ‘hooks’, and even when fully saturated with O₂ it will only be able to carry 100 mL rather than 200 mL of O₂.

• Haemoglobin saturation (%). This is the percentage of the total number of ‘hooks’ that are in fact occupied. It is nothing to do with the number of ‘hooks’ present. The number present may be increased (polycythaemia), normal or reduced (anaemia). Measurement of Hb saturation is technically simple using the spectrophotometer as described (Obtaining a dissociation curve, p. 102) and gives useful information for clinical assessment as 100% saturation of arterial blood implies the lungs are doing a good job of gas exchange. However, other measurements, particularly Po₂ and Hb content, are necessary to provide a complete picture.

Students sometimes find it helpful to think of saturation as the ‘appetite’ haemoglobin has for O₂. If haemoglobin finds itself in a Po₂ where its saturation should be high (say 10 kPa in Fig. 8.2) it is ‘hungry’ and will readily accept O₂ until it is appropriately saturated, ‘full’. At low Po₂ (say 2 kPa in Fig. 8.2) it is not so hungry; in fact, it is overstuffed for these conditions and vomits off its excess oxygen.

• Oxygen content (mL L⁻¹). We have seen (‘Haemoglobin content’, above) that the amount of oxygen in a litre of arterial blood is limited by the amount of Hb it contains. It also depends on the Po₂ of the air in the lungs driving O₂ into the blood. This underlies the difference between ‘saturation’ and ‘content’, which is very important to understand. An analogy that might help is to consider the RBC as a cloakroom used to store coats. The number of coats (O₂ molecules) that can be stored depends on the number of hooks (Hb molecules) present. The number that are actually stored (O₂ content), up to the theoretical maximum when all hooks are occupied (100% saturation), depends on the size of the cloakroom (amount of Hb) and the pressure from customers wishing to leave their coats (Po₂).

Case 8.1 Carriage of gases by the blood and acid/base balance: 4

Treatment of carbon monoxide poisoning

After the diagnosis of carbon monoxide poisoning had been made Mr Jones was treated with high concentrations of oxygen (60% inspired oxygen) and was closely monitored on the high-dependency ward. Gradually, he regained consciousness as the carbon monoxide was displaced from his blood by oxygen and the effects of the alcohol he had consumed began to wear off. The carboxyhaemoglobin levels in his blood were measured later that day and were found to be 12%.

Oxygen is the treatment of CO poisoning. By administering high concentrations of oxygen, carbon monoxide is dissociated from carboxyhaemoglobin, producing haemoglobin, which is then free to combine with oxygen. In some centres CO poisoning is treated with hyperbaric oxygen: in other words, the patient is placed in a pressurized chamber and breathes 100% oxygen. Under these circumstances, the partial pressure of oxygen is very high because the ambient pressure is high. The high partial pressure of oxygen leads to an even more rapid removal of CO. It is also possible that an improvement in oxygenation occurs because the administration of a high partial pressure of oxygen results in an increase in the amount of dissolved oxygen in the blood, possibly to a clinically significant level. The use of hyperbaric oxygen remains controversial, however, and it is only useful if it is quickly available, which is not the case in most centres.

So, if everything else is normal:

• O₂ content in blood depends on the amount of Hb present and Po₂.

• O₂ saturation depends only on oxygen partial pressure (Po₂).

The shape of the curve (Fig. 8.2)

The oxyhaemoglobin dissociation curve (Fig. 8.2) can express the relationship between Po₂ and saturation, which is independent of blood Hb content, or Po₂ and O₂ content which is not. In terms of content the curve is displaced downwards in anaemia (where Hb content is low).

Whether expressing the relationship between Po₂ and saturation or content, the curves in Figure 8.2 have the same characteristic shape, which has an important influence on function. The major function of Hb is to load with O₂ at the lungs and unload at the tissues. This function is carried out at the flat loading region at the top of the curve and at the steep unloading region. The difference in slope of the curve at these two points has the following consequences:

• Loading region (used at the lungs). Above about 10 kPa Hb cannot take up much more O₂: it is saturated, because most of the molecules of Hb are carrying their full complement of four O₂ molecules and this number cannot be exceeded however high the Po₂. Alveolar ventilation can decrease by up to 25% or increase indefinitely without affecting O₂ content significantly. O₂ tension varies, however. The evolutionary advantage of this is that normal activities such as talking, sighing, coughing, etc. do not greatly alter the amount of O₂ per litre of blood leaving the lungs for the tissues.

• Unloading region (used at the tissues). Blood in the capillaries of active tissues finds itself in an environment of low Po₂. Oxygen diffuses from blood to tissues, and even a small fall in blood Po₂ causes a large unloading of O₂, i.e. the blood is working on the steep part of the HbO₂ dissociation curve. If it stays in the tissue long enough blood Po₂ will equilibrate with tissue Po₂. If the blood is anaemic (low Hb content), however, removal of even a small amount of O₂ causes a large fall in Po₂ because there is little O₂ in the blood to begin with. A situation is quickly reached where there is little possibility of further supply to the tissues and a reduced Po₂ to drive it in. Thus anaemia can cause tissue hypoxia even though arterial blood has normal Po₂ and Hb saturation.

To identify the position of the steep part of the oxyhaemoglobin dissociation curve (usually to see if it is being oxygenated properly) the Po₂ at which 50% of the Hb is saturated is measured: this is called the P₅₀ and is about 3.2 kPa for normal adult human arterial blood.

Displacement of the oxyhaemoglobin dissociation curve (Fig. 8.3)

The evolution of Hb with a dissociation curve of the shape described has been a wonderful advantage to the species that possess it. Even more wonderful is the displacement of this curve along the Po₂ axis of the graph that takes place with each circuit of the blood between lungs and tissue and between tissue and lungs. This displacement of the dissociation curve represents cyclic changes in the properties of Hb which make it an even more efficient carrier of O₂.

Fig. 8.3 Shifts of the dissociation curve. Conditions in terms of [H⁺], CO₂, temperature and 2,3-diphosphoglycerate are very different at the sites of loading (the lungs) and unloading (the tissues) of Hb with O₂. These differences shift the dissociation curve and improve its carrying properties. For example, the effect of increased H⁺ and CO₂ at the tissues is to shift the curve to the right (this is called the Bohr shift), and this steepens the functional dissociation curve to that shown as a dotted line.

We have seen (Fig. 8.2) that abnormal amounts of Hb in the blood will displace the O₂ content curve vertically but will not affect the saturation curve. We will now look at factors that displace the curve horizontally and the way in which this improves the supply of O₂ to the tissues.

• Hydrogen ion concentration. Increased [H⁺] (decreased pH, increased acidity) displaces the curve to the right (Fig. 8.3). This Bohr shift is due to H⁺ acting on the Hb molecule to decrease its affinity for O₂. This does not affect the loading region of the curve because it is horizontal, and so movement to left or right does not produce a change in saturation.

The steep unloading region of the curve is a different matter. In metabolizing tissues the release of acids or of CO₂ (which increases [H⁺]) shifts the curve to the right. This has two major consequences, the first fairly obvious, the second less so:

1. Take a vertical line at some Po₂ on the steep part of the curve, say 4 kPa in Figure 8.3. If the curve moves to the right the saturation appropriate for that Po₂ will fall. The Hb has less ‘appetite’ for O₂ and it ‘vomits off’ the excess (see Haemoglobin saturation, p. 104, above). This is clearly an advantage, liberating O₂ to diffuse down the concentration gradient to the tissues.

2. What is not so immediately obvious but equally important is revealed if you take a horizontal line, at, say 50% saturation. When the curve moves to the right the Po₂ appropriate for that saturation increases! This increases the partial pressure gradient driving O₂ into the tissues.

These effects of acidity are so powerful that a decrease of 0.2 pH units can increase O₂ release by 25% at low Po₂.

• Carbon dioxide. In addition to its acid properties, which are dealt with above, CO₂ reacts with Hb to form carbamino Hb. This also moves the curve to the right. If hypercapnia (increased Pco₂) persists for several hours, with chronic acidosis, red cell 2,3,diphosphoglycerate (DPG, see below) is decreased, shifting the curve back to the left.

• Temperature. A decrease in temperature shifts the curve to the left. Blood therefore gives up its O₂ less readily in cold tissues, and blood leaving them may be well oxygenated because of this effect. Also, cold will reduce the metabolic demand for O₂. For this reason, children playing in the snow have pink ears and noses when their vasoconstricted skin might have been expected to turn blue. This effect is not very important in the lungs because the air in them is so well warmed. It is important, however, in patients made hypothermic during open heart surgery. In these patients, even if arterial Po₂ is low the Hb is relatively well saturated and the patient does not look hypoxic.

• 2,3-Diphosphoglycerate (DPG). In most cells under anaerobic conditions 1,3-diphosphoglycerate (1,3-DPG) is converted to 3-phosphoglycerate, with the release of energy which is stored in the form of adenosine triphosphate (ATP).

In red cells, however, 1,3-DPG is converted to 2,3-DPG without the release of energy (Fig. 8.4), an apparently pointless metabolic reaction.