Table of Contents
Indirect calorimetry, or respirometry, estimates energy conversion in organisms by measuring gas exchange with the environment instead of directly measuring heat. From these gas data, heat production and thus energy expenditure can be calculated.
Basic Principle of Indirect Calorimetry
In aerobic metabolism, nutrients are oxidized using oxygen and converted to carbon dioxide and water. The overall idea is:
- Oxygen uptake ($\mathrm{O_2}$ consumption) and carbon dioxide release ($\mathrm{CO_2}$ production) are measured.
- The amounts of $\mathrm{O_2}$ and $\mathrm{CO_2}$ used or formed depend on which substrates (carbohydrates, fats, proteins) are being oxidized.
- For known substrates, the energy released per liter of $\mathrm{O_2}$ consumed is fairly constant and can be used to calculate total energy expenditure.
Instead of measuring heat flow (as in direct calorimetry), indirect calorimetry infers heat production from the chemical changes in respiratory gases.
Respiratory Quotient (RQ)
A central concept in respirometry is the respiratory quotient (RQ). It is defined as
$$
\mathrm{RQ} = \frac{\text{CO}_2\ \text{produced}}{\text{O}_2\ \text{consumed}}
$$
measured over the same time period, usually in molar or volume units (e.g. liters per minute).
Because different nutrient classes have characteristic oxidation equations, they have characteristic RQ values.
RQ for Carbohydrate Oxidation
As a simplified example, consider the oxidation of the monosaccharide glucose:
$$
\mathrm{C_6H_{12}O_6 + 6\ O_2 \rightarrow 6\ CO_2 + 6\ H_2O}
$$
Here:
- $\mathrm{O_2}$ consumed: 6 mol
- $\mathrm{CO_2}$ produced: 6 mol
So:
$$
\mathrm{RQ_{carbohydrate} = \frac{6}{6} = 1.0}
$$
An RQ near 1.0 indicates that mainly carbohydrates are being oxidized.
RQ for Fat Oxidation
A typical fatty acid, such as palmitic acid ($\mathrm{C_{16}H_{32}O_2}$), is oxidized according to a simplified equation:
$$
\mathrm{C_{16}H_{32}O_2 + 23\ O_2 \rightarrow 16\ CO_2 + 16\ H_2O}
$$
Here:
- $\mathrm{O_2}$ consumed: 23 mol
- $\mathrm{CO_2}$ produced: 16 mol
So:
$$
\mathrm{RQ_{fat} = \frac{16}{23} \approx 0.7}
$$
An RQ near 0.7 indicates predominant fat oxidation.
RQ for Protein Oxidation
Proteins are chemically diverse, and their catabolism involves deamination and further transformations. Average protein oxidation yields an overall RQ around:
$$
\mathrm{RQ_{protein} \approx 0.8}
$$
However, exact values are more variable and more complex to determine, so protein is often treated separately or approximated.
Mixed Diet and RQ
In real organisms, a mixture of carbohydrates, fats, and proteins is usually oxidized at the same time. The measured RQ is then a weighted average:
- RQ close to 1.0: metabolism dominated by carbohydrates.
- RQ around 0.85: mixed diet, typical for resting humans after a normal meal.
- RQ near 0.7: metabolism dominated by fats (e.g. fasting, prolonged low-intensity exercise).
Thus, RQ not only helps calculate energy expenditure but also provides information about which substrates are being used.
From Gas Exchange to Energy Expenditure
Because the combustion (oxidation) of nutrients in the body follows fixed chemical relationships, energy release per liter of $\mathrm{O_2}$ consumed can be estimated. For a mixed human diet, commonly used approximate values are:
- At RQ = 1.0 (pure carbohydrate): about 21.1 kJ per liter $\mathrm{O_2}$ (≈ 5.05 kcal/L).
- At RQ = 0.7 (pure fat): about 19.6 kJ per liter $\mathrm{O_2}$ (≈ 4.69 kcal/L).
- At RQ ≈ 0.82–0.85 (mixed): roughly 20.2 kJ per liter $\mathrm{O_2}$ (≈ 4.8 kcal/L).
In practice, once $\mathrm{O_2}$ consumption and RQ (or $\mathrm{CO_2}$ production) are known, one uses tables or equations that relate RQ to energy released per liter of $\mathrm{O_2}$ to calculate heat production.
Calculating Energy Expenditure (Conceptual)
- Measure $\mathrm{O_2}$ consumption per unit time, e.g. $\mathrm{\dot V O_2}$ in $\mathrm{L/min}$.
- Measure $\mathrm{CO_2}$ production per unit time, e.g. $\mathrm{\dot V CO_2}$ in $\mathrm{L/min}$.
- Compute RQ: $\mathrm{RQ = \dot V CO_2 / \dot V O_2}$.
- Use RQ to look up (or compute) the energy equivalent of 1 L $\mathrm{O_2}$ (kJ/L or kcal/L).
- Multiply:
$$\text{Energy per time} = \mathrm{\dot V O_2} \times \text{energy equivalent of } 1\ \text{L } \mathrm{O_2}$$
This gives energy expenditure per minute. It can be converted to per hour or per day.
Methods of Measuring Gas Exchange
Indirect calorimetry can be carried out at different scales: from whole organisms (e.g. humans, small mammals) to isolated tissues or small animals.
Open-Circuit Respirometry
In open-circuit systems, the organism breathes room air (or a defined gas mixture), and the composition of inhaled and exhaled air is compared.
Typical elements:
- A mouthpiece, mask, or hood system for humans, or a chamber with inlet/outlet for small animals.
- Flow meters to measure the volume of air passing per unit time.
- Gas analyzers that continuously or periodically measure $\mathrm{O_2}$ and $\mathrm{CO_2}$ fractions in inspired and expired air.
Basic idea:
- Know the flow rate of air (e.g. liters per minute).
- Measure the difference between inhaled and exhaled $\mathrm{O_2}$ concentrations (and similarly for $\mathrm{CO_2}$).
- From flow and concentration differences, calculate volumes of $\mathrm{O_2}$ consumed and $\mathrm{CO_2}$ produced per unit time.
Because the organism is not sealed, the system is technically simpler and more commonly used in larger animals and humans.
Closed-Circuit Respirometry
In closed-circuit systems, the organism breathes from a defined, closed gas volume.
Key features:
- The subject is connected to a limited gas reservoir.
- $\mathrm{CO_2}$ is absorbed chemically (e.g. by soda lime) so its concentration does not accumulate dangerously.
- The decline in $\mathrm{O_2}$ in the system, or the volume of $\mathrm{O_2}$ that must be added to keep concentration constant, is measured.
From the change in $\mathrm{O_2}$ content over time, $\mathrm{O_2}$ consumption can be determined. Closed systems are often used in smaller animals or in controlled laboratory experiments but are less common for humans due to comfort and safety considerations.
Whole-Body vs. Local Respirometry
- Whole-body respirometry: measures gas exchange of the entire organism, providing total metabolic energy expenditure.
- Local (tissue or organ) respirometry: small preparations (e.g. isolated muscle fibers) are placed in sealed chambers, and the small changes in gas composition are measured to estimate local metabolism.
Whole-body methods are typically used in physiology, sports science, and clinical medicine, whereas local methods are common in experimental cell and tissue studies.
Assumptions and Limitations
Indirect calorimetry does not measure heat directly, so its applicability depends on certain assumptions.
Requirement for Aerobic Metabolism
Indirect calorimetry relies on the link between oxygen use, carbon dioxide production, and energy release during oxidative metabolism.
- Under purely aerobic conditions, the relationship between gas exchange and heat production is well-defined.
- During strong anaerobic metabolism (e.g. intense short-term exercise), part of the energy comes from pathways that do not immediately consume $\mathrm{O_2}$ (e.g. glycolysis with lactate formation). Then, $\mathrm{O_2}$ consumption during the measurement period may not match the instant energy expenditure.
Therefore, indirect calorimetry is most accurate when the organism is in a metabolic steady state dominated by aerobic processes.
Nitrogen and Protein Metabolism
When proteins are substantially oxidized, nitrogen is removed and excreted (e.g. as urea in urine). Because nitrogen is not exchanged via the lungs in the same way as $\mathrm{O_2}$ and $\mathrm{CO_2}$:
- Protein oxidation complicates the direct use of $\mathrm{O_2}$ and $\mathrm{CO_2}$ for exact energy calculations.
- In detailed measurements, nitrogen excretion (often urinary nitrogen) may be measured to correct for protein metabolism.
In many practical applications, especially short-term measurements in humans, protein contribution to energy expenditure is relatively small and can be approximated.
RQ Versus Respiratory Exchange Ratio (RER)
In theory, RQ refers strictly to cellular metabolism (moles of $\mathrm{CO_2}$ produced / moles of $\mathrm{O_2}$ consumed at the tissue level). Indirect calorimetry often measures the respiratory exchange ratio (RER):
$$
\mathrm{RER = \frac{V_{CO_2\ (expired)}}{V_{O_2\ (inspired)}}}
$$
Under steady-state conditions, RER ≈ RQ, and the distinction is unimportant. However:
- Non-steady conditions (e.g. hyperventilation, changes in body bicarbonate stores) can cause RER to deviate from true tissue RQ.
- For example, during intense exercise, RER can exceed 1.0 due to extra $\mathrm{CO_2}$ from buffering of lactic acid, even though no substrate has an RQ > 1.0.
This must be considered when interpreting RER measurements.
Effects of Environment and Equipment
Several sources of error can influence measurements:
- Leaks in the system or inaccurate flow measurements.
- Changes in water vapor in inhaled and exhaled air.
- Temperature and pressure effects on gas volume (often corrected by standardizing to STPD: standard temperature and pressure, dry).
Proper calibration and correction are essential for accurate data.
Applications of Indirect Calorimetry
Because it is noninvasive and relatively practical, respirometry is widely used to study metabolism.
Determining the Metabolic Rate
Indirect calorimetry is a cornerstone for determining:
- Basal metabolic rate (BMR) under standardized resting conditions.
- Resting metabolic rate (RMR) in more typical but still low-activity conditions.
- Energy expenditure during physical activity or exercise at different intensities.
These concepts tie into broader topics of metabolic rate and performance metabolism, treated elsewhere.
Substrate Utilization
Because RQ/RER indicates which substrates are being oxidized, indirect calorimetry can be used to estimate:
- Fraction of energy derived from fats vs. carbohydrates at various exercise intensities.
- Changes in substrate use during fasting, feeding, or after training.
- The effects of diet composition on fuel selection during rest and activity.
This is important in sports physiology, nutrition, and clinical metabolism.
Clinical and Ecological Uses
In medicine:
- Monitoring energy needs of critically ill patients to individualize nutritional support.
- Detecting abnormalities in metabolism (e.g. unusually high or low energy expenditure).
In ecology and animal physiology:
- Measuring field metabolic rates of animals (with adapted methods) to understand energy budgets and adaptations.
- Comparing metabolic strategies of species in different environments.
In all these contexts, indirect calorimetry provides a window into how much energy an organism uses and how that energy is derived from different nutrient sources, without directly measuring heat production.