describe and carry out investigations, using simple respirometers, to determine the RQ of germinating seeds or small invertebrates (e.g. blowfly larvae)
Energy – Determining the Respiratory Quotient (RQ) with Simple Respirometers
1. Introduction
Cellular respiration releases energy from organic substrates. The ratio of carbon dioxide produced to oxygen consumed, known as the respiratory quotient (RQ), provides insight into which macromolecule (carbohydrate, protein or lipid) is being metabolised.
2. The Respiratory Quotient (RQ)
The RQ is defined as:
\$\text{RQ} = \frac{\text{Volume of } \mathrm{CO2} \text{ produced}}{\text{Volume of } \mathrm{O2} \text{ consumed}}\$
Carbohydrate oxidation: RQ ≈ 1.0
Protein oxidation: RQ ≈ 0.8
Lipid oxidation: RQ ≈ 0.7
3. Simple Respirometer – Principle and Design
A simple respirometer measures the change in gas volume inside a sealed chamber as the organism consumes O₂ and produces CO₂. The change is recorded by a water‑filled manometer or a graduated syringe.
Suggested diagram: Schematic of a simple respirometer showing the sealed chamber, water‑filled manometer, and inlet/outlet tubing.
3.1 Components
Chamber – airtight container (e.g., 100 mL graduated cylinder).
Manometer – U‑tube filled with water (or coloured water) to detect volume changes.
Stopcock – to seal the system after loading the organism.
Thermometer – to monitor temperature (respiration rate is temperature‑dependent).
Timer – for fixed observation periods.
4. Preparing the Test Organisms
4.1 Germinating Seeds
Soak seeds (e.g., wheat or barley) in distilled water for 12 h.
Place 5–10 seeds on moist filter paper in a Petri dish and keep at 25 °C for 48 h until radicle emergence.
Transfer the germinated seeds to the respirometer chamber using a fine brush.
4.2 Small Invertebrates (e.g., Blowfly Larvae)
Collect active third‑instar larvae and rinse briefly in distilled water.
Weigh 0.5–1.0 g of larvae (record mass).
Place the larvae in the chamber with a small amount of moist filter paper to prevent desiccation.
5. Experimental Procedure
Fill the manometer with water and ensure no air bubbles are trapped.
Insert the sealed chamber into a water bath set at the desired temperature (usually 25 °C).
Introduce the test organism into the chamber, then quickly close the stopcock to seal the system.
Record the initial water level (baseline) in the manometer.
Allow the organism to respire for a fixed period (e.g., 30 min). Record the water level at regular intervals (every 5 min).
After the observation period, remove the organism and note the final water level.
Repeat the experiment at least three times for each organism to obtain reliable data.
6. Data Collection and Presentation
Convert the change in water level (Δh) to gas volume change (ΔV) using the cross‑sectional area (A) of the manometer tube:
\$\Delta V = A \times \Delta h\$
Trial
Initial level (cm)
Final level (cm)
Δh (cm)
ΔV (mL)
O₂ consumed (mL)
CO₂ produced (mL)
RQ
1
0.0
2.3
2.3
1.15
0.92
0.23
0.25
2
0.0
2.5
2.5
1.25
1.00
0.25
0.25
3
0.0
2.4
2.4
1.20
0.96
0.24
0.25
7. Calculations
Determine the total change in gas volume (ΔV) from the manometer reading.
Assuming a 1:1 stoichiometry of O₂ consumption to CO₂ production in the sealed system, split ΔV into O₂ and CO₂ components:
O₂ consumed = (ΔV × fraction of O₂ taken up)
CO₂ produced = (ΔV × fraction of CO₂ released)
In practice, the volume change is the net result of O₂ loss minus CO₂ gain. For a simple respirometer, the net change equals O₂ consumed – CO₂ produced. Rearranging:
\$\text{O}2\text{ consumed} = \frac{\Delta V + \text{CO}2\text{ produced}}{1}\$
If the system is calibrated with a known gas mixture, the fractions can be derived; otherwise, use the assumption that the change is dominated by O₂ uptake for short periods.
Average the RQ values and compare with theoretical values to infer the primary substrate being metabolised.
8. Safety and Precautions
Handle live organisms humanely; minimise stress.
Use gloves when handling water‑filled manometers to avoid slipping.
Ensure the stopcock is fully closed before starting the timer to prevent gas leaks.
Maintain a constant temperature; fluctuations affect respiration rates.
9. Evaluation of the Method
Advantages
Low‑cost and easy to construct.
Suitable for small organisms where larger respirometers are impractical.
Limitations
Assumes that all volume change is due to O₂/CO₂ exchange; water vapour and temperature changes can introduce error.
Calibration is required for accurate separation of O₂ and CO₂ contributions.
Short observation periods may give low signal‑to‑noise ratios.
Possible Improvements
Incorporate a CO₂‑absorbing solution (e.g., NaOH) in a side arm to isolate O₂ consumption.
Use a digital pressure sensor to increase measurement precision.
Conduct experiments at multiple temperatures to generate Arrhenius plots.
10. Conclusion
By using a simple respirometer, students can quantitatively determine the respiratory quotient of germinating seeds or small invertebrates. The measured RQ provides insight into the metabolic substrate being utilised during early development, linking the concepts of energy metabolism to observable experimental data.