The amount (in milliliters) of O₂ that dissolves in plasma is determined by the P_AO₂ to which the plasma is exposed (Henry’s law). The relationship between PO₂ and dissolved O₂ is linear (i.e., doubling or tripling PO₂ doubles or triples the amount of dissolved O₂) (Figure 8-1). Chapter 7 explained that the solubility coefficient of O₂ is 0.0244 mL O₂/mL plasma/atmosphere of pressure (atm). This means a PO₂ of 760 mm Hg causes 0.0244 mL of O₂ to dissolve in 1 mL of plasma at a body temperature of 37° C (Figure 8-2, A). It also means that a PO₂ of only 1 mm Hg causes 0.00003 mL of O₂ to dissolve in 1 mL of plasma, as shown in Figure 8-2, B (0.0244/760 = 0.00003). By convention, blood O₂ content units are expressed in milliliters of O₂ per 100 mL of blood, or mL/dL. (The unit mL/dL has replaced the outdated term vol%.) To express O₂ content in its proper units, the amount of O₂ that dissolves in 100 mL of plasma for each millimeter of mercury (mm Hg) of PO₂ is 0.003 mL/dL (0.00003 mL O₂/mL plasma/mm Hg × 100 mL plasma = 0.003 mL/dL/mm Hg, as shown in Figure 8-2, C). At a normal arterial partial pressure of oxygen (PaO₂) of 100 mm Hg, 0.3 mL of O₂ dissolves in 100 mL of plasma, or 0.3 mL/dL (PO₂ of 100 mm Hg × 0.003 mL/dL/mm Hg = 0.3 mL/dL dissolved O₂, as Figure 8-2, D, shows). The 0.003 factor can be used to calculate dissolved plasma O₂ content at any PO₂ as follows:

Figure 8-1 Relationship between PO₂ and dissolved oxygen in the blood plasma (dissolved O₂ in mL/dL = PO₂ × 0.003).

Figure 8-2 Solubility coefficient of oxygen in blood plasma at 37° C. A, At 1 atm (760 mm Hg) of oxygen pressure, 0.0244 mL of oxygen dissolves in 1 mL of plasma. B, 1/760 of this amount dissolves in 1 mL of plasma at a PO₂ of 1 mm Hg. C, 100 times the amount in B dissolves in 100 mL of plasma. D, At a normal arterial PO₂ of 100 mm Hg, 100 times the amount of oxygen dissolved in C dissolves in 100 mL of plasma; normal dissolved oxygen content in arterial blood is about 0.3 mL/dL.

PO2×0.003=mL/dL dissolvedO2

PO₂ in the blood plasma is sometimes called the blood’s O₂ tension, which comes from the concept that dissolved gases have an “escaping tendency.” That is, if the PO₂ of a gas to which the plasma is exposed suddenly decreased, dissolved O₂ would immediately diffuse out of the plasma into the gaseous phase. Although it is correct to think of PO₂ as a force that dissolves O₂ in the plasma, it is also correct to think of dissolved O₂ as creating a force as it tries to escape to the gaseous phase—a force responsible for the plasma PO₂. The plasma PO₂ is extremely important in determining the adequacy of blood and tissue oxygenation because it determines the direction and rate of O₂ diffusion in the lungs and body tissues. Plasma PO₂ is intimately related to how O₂ binds with hemoglobin, as described later in this chapter.

CONCEPT QUESTION 8-1

What is the PO₂ of normal plasma with a dissolved O₂ content of 1.8 mL/dL?

Oxygen Combined with Hemoglobin

Most O₂ in the blood is bound to hemoglobin inside the erythrocyte. Without hemoglobin, blood could not carry enough O₂ to meet minimal tissue oxygenation requirements.

Hemoglobin allows whole blood (plasma plus cellular components) to carry about 20 mL of O₂ per 100 mL of blood at a normal PaO₂ of 100 mm Hg. This is about 67 times more than the capacity of plasma alone when PaO₂ is 100 mm Hg ([20 mL/dL]/[0.3 mL/dL] = 66.6). A normal cardiac output (Q˙) of 5 L per minute delivers about 1000 mL of O₂ to the body’s tissues each minute. This is more than enough O₂ to meet the resting requirement of tissues, which is only 250 mL per minute. This is shown as follows:

1. mL O2/L blood=20mLO2/100 mL blood×10 dL/L=200 mLO2/L

2. O2DEL=5 L/minbloodflow× 200mLO2/L=1000mLO2/min

Oxygen delivery (O₂ DEL) rate is normally about four times the resting requirement of the body tissues. From another perspective, the tissues normally extract only one fourth of the O₂ present in the blood they receive. The remaining three fourths (750 mL per minute) is an available reserve for increased metabolic demands.

Hemoglobin Molecule

Hemoglobin is a large, complex protein molecule found inside the erythrocyte. It has a molecular weight of about 64,500 and accounts for about one third of the red blood cell volume. Because hemoglobin exists inside the red blood cell, high concentrations can be carried without affecting the blood’s oncotic pressure.

As its name suggests, hemoglobin consists of heme (an iron-containing pigment) and globin (a protein). Heme is an organic molecule consisting of four symmetrically linked pyrrole rings, with a ferrous iron ion (Fe⁺⁺) at its center (Figure 8-3, A-C). A pyrrole is an organic molecule organized in a ringlike structure. In addition to carbon atoms, a nitrogen atom helps form the ring. Pyrroles are building blocks for respiratory pigments that take up O₂. The four pyrrole rings are linked with methylene bridges (see Figure 8-3, B) to form a porphyrin molecule, the basis for hemoglobin’s respiratory pigment. Porphyrin molecules readily form covalent bonds with metals. In this case, Fe⁺⁺, which has six available sites for covalent bonding, bonds with four porphyrin molecules, leaving two unused bonding sites (see Figure 8-3, C). One of these sites bonds with a portion of the globin molecule.

Figure 8-3 Hemoglobin molecule. Pyrrole rings (A) linked by methylene bridges form porphyrin molecules (B), which form covalent bonds with iron to form heme (C). The two remaining bonding sites on the Fe⁺⁺ ion bond with a globin polypeptide chain and an oxygen molecule. Each of the four polypeptide chains of the globin molecule bind with a heme group to form the hemoglobin molecule (D).

The globin molecule is a complex protein consisting of four linked amino acid chains; normal adult hemoglobin (HbA) has two alpha (α) chains, each composed of 141 amino acids, and two beta (β) chains, each composed of 146 amino acids. These amino acid chains are called polypeptides. Each polypeptide chain bonds with one heme molecule at one of the two remaining sites of the Fe⁺⁺ ion. A hemoglobin molecule consists of four heme groups, each bonded with and enfolded in one of the globin molecule’s four polypeptide chains (see Figure 8-3, D). Each of the four heme groups has one remaining bonding site on the Fe⁺⁺ ion. This sixth site binds rapidly and reversibly with an O₂ molecule, but iron remains in the nonoxidized, ferrous state (Fe⁺⁺). Because each of the four polypeptide-heme combinations can bind one molecule of O₂, a single hemoglobin molecule can bind four O₂ molecules. The hemoglobin molecule is called a tetramer because it contains four polypeptide subunits.

By themselves, heme, iron, and globin cannot take up O₂; all three must be combined chemically with ferrous iron in a precise spatial arrangement before O₂ can be taken up from the plasma. The precise amino acid sequence of each polypeptide chain is important for normal O₂ uptake. A variation in this sequence changes how readily hemoglobin binds O₂ (i.e., it changes the affinity of hemoglobin for O₂). Variations of HbA are discussed later in this chapter.

Hemoglobin Combined with Oxygen

Combined with O₂, hemoglobin is called oxyhemoglobin (HbO₂). O₂ does not normally oxidize hemoglobin; rather, it oxygenates hemoglobin. Unoxygenated hemoglobin is called deoxyhemoglobin (Hb). The term “reduced” hemoglobin, which is sometimes used, is chemically incorrect because iron remains in the ferrous (Fe⁺⁺) state.

The four heme groups of the hemoglobin molecule take up and release their O₂ molecules in succession in a process known as cooperative O₂ binding.¹ The binding of an O₂ molecule to one heme group induces a structural change in the shape of the hemoglobin molecule, which increases the affinity of the next heme subunit for O₂. Binding of each O₂ molecule makes the remaining bonds occur with successively greater speed and ease. Similarly, the release of the first O₂ molecule facilitates the release of each remaining molecule. This cooperative binding mechanism is an efficient way to take up O₂ when PO₂ is increased (in the alveoli) and to release O₂ when PO₂ is decreased (in the body’s tissues). Cooperative binding means that a hemoglobin molecule either is bound to four O₂ molecules or is bound to none. In other words, the hemoglobin molecule either is fully oxygenated or is fully deoxygenated.

The change in shape of the hemoglobin molecule that occurs as it binds and releases O₂ causes it to reflect and absorb light differently when it is oxygenated than when it is deoxygenated. This phenomenon is responsible for the bright red color of oxygenated hemoglobin and the deep purple color of deoxyhemoglobin. This difference in light absorption and reflection makes it possible to measure the amount of oxygenated hemoglobin present in a blood sample through a process known as spectrophotometry or, as it is clinically known, oximetry.

Hemoglobin Saturation and Oxygen Partial Pressure

Hemoglobin is 100% saturated with O₂ if every hemoglobin molecule is combined with four O₂ molecules. Saturation is expressed as the percentage of hemoglobin molecules in the blood that are oxygenated. Normally, at sea level about 97.5% of the hemoglobin molecules in arterial blood are oxygenated; the remaining 2.5% are unoxygenated. This means that arterial oxygen saturation (SaO₂) is about 97.5% at a normal PaO₂ of 100 mm Hg. Only 75% of the hemoglobin molecules in mixed venous blood are oxygenated. That is, mixed venous oxygen saturation (Sv¯O₂) is normally 75%, which corresponds to a partial pressure of oxygen in mixed venous blood (Pv¯O₂) of about 40 mm Hg.

Saturation can be thought of as a percentage of hemoglobin’s capacity for O₂. An SaO₂ of 90% means the hemoglobin in the blood is carrying 90% of its O₂ capacity. Mixed venous blood hemoglobin normally carries only 75% of its O₂ capacity. It is important to understand that hemoglobin’s O₂ saturation percentage is not a measure of blood O₂ content, nor is it relevant to the concentration of hemoglobin in the blood. In other words, 100% saturation does not automatically imply that blood O₂ content is normal or that hemoglobin concentration is normal. If hemoglobin concentration is low (as it is in anemia), blood O₂ content is low even if the hemoglobin present is 100% saturated with O₂. Likewise, low saturation does not automatically mean that blood O₂ content is below normal, although this is generally the meaning. For example, hemoglobin concentration may be abnormally high (polycythemia) in individuals who have chronic hypoxia, which makes it possible for blood O₂ content to be normal even though hemoglobin saturation percentage is below normal.

Hemoglobin Capacity for Oxygen

When hemoglobin is 100% saturated with O₂, each gram (g) of hemoglobin carries 1.34 mL of O₂. The normal concentration of hemoglobin in the blood ranges from 12 to 15 g per 100 mL blood (12 to 15 g/dL). Assuming a hemoglobin concentration of 15 g/dL, hemoglobin’s O₂-carrying capacity (100% saturation) is as follows: 15 × 1.34 = 20.1 mL/dL. Mixed venous blood

CLINICAL FOCUS 8-1 Clinical Measurement of Blood Oxygen Saturation

The measurement of the oxygen saturation of hemoglobin in the clinical setting involves a process called spectrophotometry. Spectrophotometry measures light wavelengths. All substances absorb and emit a unique spectrum of light wavelengths, known as an absorption spectrum. Each form of hemoglobin (e.g., HbO₂, Hb, and carboxyhemoglobin [HbCO]) has its own absorption spectrum. A spectrophotometer specifically designed to measure the absorption spectra of various hemoglobin forms is called an oximeter.

An oximeter passes the light of specific wavelengths through a blood sample to a photodetector on the opposite side. The photodetector senses the light’s intensity and converts it to a proportional electrical current. When light enters the blood sample, certain wavelengths may be absorbed, transmitted, or reflected. The greater the optical density or opaqueness of the blood sample, the less light can be transmitted through it. The oximeter differentiates between HbO₂ and deoxyhemoglobin based on their different absorption spectra and quantifies their concentrations. Oximeters that use only two light wavelengths can detect only Hb and HbO₂; they cannot detect other forms of hemoglobin such as HbCO. An instrument called a CO-oximeter distinguishes among all forms of hemoglobin and displays their relative percentages. CO-oximeters are used in blood gas laboratories with blood gas analyzers, allowing SaO₂ to be included in the blood gas report. Pulse oximeters are compact portable devices that pass two wavelengths of light through an extremity such as a finger or toe, sensing the absorption spectrum of arterial blood during the systolic pulse. These oximeters are convenient because they do not require blood sampling, but their accuracy is limited in the presence of abnormal hemoglobin forms.

(75% saturated) carries 15.1 mL/dL of O₂ (0.75 × 20.1), whereas arterial blood (97.5% saturated at PO₂ of 100 mm Hg) carries about 19.7 mL/dL of O₂ (0.975 × 20.1).

CONCEPT QUESTION 8-2

What is the O₂ content (mL/dL) of an anemic person’s blood while breathing 100% O₂ (hemoglobin concentration 10 g/dL, PaO₂ 550 mm Hg, and SaO₂ 100%)?

Oxyhemoglobin Equilibrium Curve

The blood plasma PO₂ determines hemoglobin’s saturation with O₂. The HbO₂ equilibrium curve (Figure 8-4), also known as the HbO₂ dissociation curve, shows the relationship between plasma PO₂ (horizontal axis) and the percentage of hemoglobin saturated with oxygen (SO₂) (vertical axis). Because SO₂ and O₂ content are directly related (as described in the foregoing section), either one can be plotted on the vertical axis of the curve (see Figure 8-4).

Figure 8-4 Oxyhemoglobin (HbO₂) equilibrium curve. P₅₀ is the PO₂ at which 50% of the hemoglobin is saturated with oxygen.

The HbO₂ equilibrium curve can be constructed by exposing whole blood (which in Figure 8-4 contains 15 g Hb per 100 mL blood) to gas mixtures with successively higher PO₂ values. The blood pH, PCO₂, and temperature are held constant throughout the process. When hemoglobin is 100% saturated, each gram carries 1.34 mL of O₂. Thus, 100% saturation corresponds to a hemoglobin O₂ content of 20.1 mL/dL (15 g/dL × 1.34 mL O₂/g = 20.1 mL/dL); 50% saturation corresponds to half this content, or 10.05 mL/dL.

In contrast to the relationship between dissolved O₂ content and PO₂ (see Figure 8-1), the relationship between PO₂ and hemoglobin saturation or O₂ content is nonlinear. The sigmoid shape of the HbO₂ curve (see Figure 8-4) is caused by the hemoglobin molecule’s change in O₂ affinity as each O₂ molecule binds in succession with heme. The middle steep portion of the curve reflects the rapid loading or unloading of O₂ molecules after hemoglobin’s binding or release of the first O₂ molecule. Small PO₂ changes cause large blood O₂ content changes in the middle steep portion of the curve (20 to 60 mm Hg). In contrast, large PO₂ changes cause small to minimal changes in O₂ content at the extreme flatter ends of the curve, especially the flat, right end (60 to 100 mm Hg).

CONCEPT QUESTION 8-3

Is the gain in blood O₂ content when PO₂ increases from 40 mm Hg to 60 mm Hg greater than, equal to, or less than the O₂ content gain when PO₂ increases from 100 mm Hg to 600 mm Hg?

Physiological Advantages of the Oxyhemoglobin Equilibrium Curve Shape

The shape of the HbO₂ equilibrium curve has important physiological consequences. As shown in Figure 8-5, the upper flat section between PO₂ values of 60 mm Hg and 100 mm Hg can be thought of as the association part of the curve because O₂ uptake in the lung normally occurs in this PO₂ range. The lower part of the curve (PO₂ <60 mm Hg) can be thought of as the dissociation part of the curve because O₂ release to the tissues occurs at these lower PO₂ values.

Figure 8-5 Association and dissociation parts of the HbO₂ curve.

The flat association part of the HbO₂ curve provides a considerable safety margin in that blood PO₂ can decrease from 100 mm Hg to 60 mm Hg (Figure 8-6) and cause only a small reduction in blood O₂ content; SO₂ decreases only 7.5% (97.5% to 90%), corresponding to a reduction in arterial oxygen content (CaO₂) of only 1.5 mL/dL (19.6 mL/dL to 18.1 mL/dL). PaO₂ can decrease considerably without reducing the blood O₂ content significantly, as long as this occurs on the flat part of the HbO₂ curve; this is diagnostically important because a change in PaO₂ reflects oxygenation problems much sooner than a change in SaO₂. If the HbO₂ curve were linear rather than sigmoid, a reduction in PO₂ from 100 mm Hg to 60 mm Hg would greatly decrease SO₂ and O₂ content (Figure 8-7).

Figure 8-6 Significance of the HbO₂ curve shape. A 40-mm Hg decrease in PO₂ from 100 mm Hg to 60 mm Hg causes a 7.5% reduction in hemoglobin saturation (*pink area*). The same 40-mm Hg decrease in PO₂ from 60 mm Hg to 20 mm Hg causes a 60% reduction in hemoglobin saturation (*green area*).

Figure 8-7 Hypothetical linear HbO₂ curve versus the actual S-shaped curve. If the curve were linear (straight line from 0 to 100 mm Hg), a PO₂ of 60 mm Hg would produce a HbO₂ saturation of 60 mm Hg and an oxygen content of 12 mL/dL instead of the actual saturation of 90% and oxygen content of 18 mL/dL.

Increases in PO₂ above 60 mm Hg do not add much oxygen to the blood; this is especially true for PO₂ values greater than 100 mm Hg, where the HbO₂ curve is virtually flat. Likewise, maximal hyperventilation is ineffective in increasing SO₂ and O₂ content in healthy individuals; for example, extreme hyperventilation with room air at best might lower alveolar pressure of carbon dioxide (P_ACO₂) to about 15 mm Hg. According to the alveolar gas equation (see Chapter 7), this would produce a maximally attainable PO₂ in the alveolus of about 130 mm Hg and a slightly lower PO₂ in the arterial blood because of normal shunt. Even at a normal PaO₂ of 100 mm Hg when breathing room air, hemoglobin is already 97.5% saturated with O₂; technically, it is fully saturated at a PO₂ of about 250 mm Hg², although for all practical purposes, one can consider hemoglobin to be 100% saturated at a PO₂ of 130 mm Hg. Even increasing the PaO₂ to 600 mm Hg by breathing 100% O₂ increases SaO₂ by only 2.5%, from 97.5% to 100%. This increase adds only 0.5 mL/dL of O₂ to the blood (from 19.6 mL/dL to 20.1 mL/dL). The steep dissociation part of the HbO₂ curve is especially suited for releasing O₂ to the tissues, where the PO₂ is between 10 mm Hg and 40 mm Hg. A reduction in PO₂ from 60 mm Hg to 20 mm Hg causes SO₂ to decrease from 90% to about 32% (see Figure 8-6), corresponding to a reduction in CaO₂ of about 11.7 mL/dL (from 18.1 mL/dL to 6.4 mL/dL). Thus, hemoglobin releases about 11.7 mL/dL of O₂ into the plasma; this increases the plasma PO₂, which provides the pressure gradient for O₂ to diffuse into the tissues.

Pv¯O₂ reflects the average PO₂ of all body tissues and is normally about 40 mm Hg. A PO₂ of 40 mm Hg corresponds with about 75% Hb saturation with O₂. The body’s tissues extract about 25% of the arterial O₂ content (about 5 mL/dL) under resting conditions. During heavy exercise, average tissue PO₂ may decrease to 20 mm Hg or less, and the tissues may extract 70% or more of the CaO₂ (Figure 8-8). Coupled with this greater O₂ extraction rate, cardiac output increases considerably—six or seven times the resting level in trained athletes—which can increase O₂ delivery to the tissues 20-fold.¹

Figure 8-8 Oxygen unloading at rest and during exercise. Decreases in tissue PO₂ values occur on the steep part of the curve and cause greater oxygen unloading. (From Thibodeau GA, Patton KT: *Anatomy & physiology,* ed 3, St Louis, 1996, Mosby. Network Graphics.)

CLINICAL FOCUS 8-2 Sensitivity of Arterial Oxygen Saturation and Arterial Oxygen Partial Pressure to Decreased Blood Oxygen Content

You are called to the emergency department to draw arterial blood for blood gas analysis on a young teenager experiencing a severe asthma attack. Emergency department personnel assure you that the oxygen saturation, as obtained by a pulse oximeter, is “fine” at 90%. You are further informed that in the past 45 minutes the saturation decreased only “slightly” from 95% to 90%. The patient appears to be in significant respiratory distress. Before you draw the blood gas sample, what is your interpretation of the recent decrease in saturation from 95% to 90%?

Discussion

A health care provider’s perspective suddenly changes when it is realized that a “slight” reduction in SaO₂ from 95% to 90% means that the PaO₂ decreased about 25 mm Hg, from 85 mm Hg to 60 mm Hg (see Figure 8-6). Health care personnel tend to be a bit more alarmed by a decrease in PaO₂ of this magnitude. The arterial oxygen content decreased only 5%. However, the PaO₂ of 60 mm Hg is on the “shoulder” of the curve. As the PO₂ falls below this point, sharp decreases in oxygen saturation and content occur for relatively small changes in PO₂. The important clinical point is that as a previously healthy person develops hypoxemia, changes in PaO₂ reflect oxygenation defects much sooner than changes in SaO₂. A decrease in a young person’s PaO₂ from 85 mm Hg to 60 mm Hg indicates a significant change in the lung’s oxygenation ability. The nature of hemoglobin’s combination with oxygen during this decrease provides a protective margin of safety because blood oxygen content does not quickly decrease on the “flat” portion of the O₂-Hb equilibrium curve. At the same time, this protective mechanism masks the severity of the oxygenation defect. Changes in SaO₂ must be interpreted in light of the corresponding PaO₂ values to gain a true appreciation for changes in the lung’s oxygenating ability.

Affinity of Hemoglobin for Oxygen

The O₂ pressure at which hemoglobin is half-saturated (P₅₀) is a measure of hemoglobin’s affinity for O₂. Normally, a PO₂ of 27 mm Hg produces an SO₂ of 50% when the blood temperature is 37° C, pH is 7.40, and PCO₂ is 40 mm Hg (see Figure 8-4). An increased P₅₀ means more than 27 mm Hg of PO₂ is required to saturate 50% of the hemoglobin. In this instance, one can think of Hb as being more “reluctant” to bind O₂, and greater pressure is required to make it do so. An increased P₅₀ means hemoglobin’s affinity for O₂ is decreased. Similarly, a decreased P₅₀ means hemoglobin’s affinity for O₂ is increased, causing it to bind O₂ molecules more aggressively (i.e., in this situation, hemoglobin saturation is >50% at a PO₂ of 27 mm Hg).

In the alveoli where O₂ loading takes place, blood with a decreased P₅₀ (high O₂ affinity) is advantageous. Conversely, high hemoglobin affinity for O₂ is detrimental at the tissue level where O₂ release is required. Ideally, hemoglobin affinity for O₂ changes depending on whether loading or unloading is required.

Oxyhemoglobin Curve Shifts

Factors that change Hb affinity for O₂ shift the HbO₂ curve to the left or to the right. Factors that increase Hb affinity for O₂ cause a leftward curve shift, or a decreased P₅₀; factors that decrease Hb affinity cause a rightward curve shift, or an increased P₅₀ (Table 8-1). Figure 8-9 shows HbO₂ curves for blood passing through pulmonary capillaries (left, blue curve) and through systemic tissue capillaries of heavily exercising muscle (right, red curve). As blood from the lungs travels through the systemic tissue capillaries, the HbO₂ curve shifts rightward from the blue curve position in Figure 8-9 to the red curve position. This right shift reflects hemoglobin’s

TABLE 8-1

Indicators of Hemoglobin Affinity for Oxygen

Increased Affinity	Decreased Affinity
Left HbO₂ curve shift	Right HbO₂ curve shift
Decreased P₅₀ (<27 mm Hg)	Increased P₅₀ (>27 mm Hg)
Greater SO₂ for given PO₂	Decreased SO₂ for given PO₂