A respirometer is a device used to measure the rate of gas exchange in a living system, typically determining the rate of cellular respiration in small organisms or seeds. The apparatus works by enclosing biological material in a sealed chamber and monitoring the change in gas volume over time. During aerobic respiration, oxygen is consumed and carbon dioxide is produced, so the net change in gas volume indicates the rate of oxygen uptake. This measurement relies on the careful preparation and reading of a liquid marker, often called a gas bubble or manometric fluid, within a calibrated tube.
Preparing the Apparatus for Measurement
Before any measurement can be taken, the respirometer system must be sealed to ensure the only gas volume change recorded is due to metabolic activity. This involves checking all connections, such as stoppers and tubing, for leaks. A leak-check can be performed by briefly manipulating the manometric fluid with a syringe and observing whether the fluid level remains static.
A carbon dioxide absorbent, such as potassium hydroxide (KOH) solution or soda lime, is placed within the chamber. This strong base chemically absorbs all the carbon dioxide produced by the respiring material. By removing the produced carbon dioxide, the net change in gas volume within the chamber is solely due to oxygen consumption, simplifying the interpretation of the fluid’s movement.
Temperature control is essential for accurate measurement because gases expand and contract with changes in temperature. To maintain a constant temperature, the respirometer is often submerged in a thermostatically controlled water bath. This thermal stability prevents fluctuations that would cause the gas volume to change independently of the respiration rate. Once sealed and placed in the water bath, the apparatus must equilibrate for 5 to 10 minutes to ensure all components reach a stable temperature before starting the experiment.
Measuring Fluid Displacement
Reading the gas bubble or manometric fluid involves documenting its change in position against a calibrated scale. The first step involves setting the initial position of the fluid meniscus to a known starting point, often the zero mark on the capillary tube or pipette. This starting point is achieved by using an attached syringe or a screw clip to adjust the internal pressure and push the fluid to the desired position.
After the initial reading is established, the measurement period begins, and the fluid moves toward the respirometer chamber as oxygen is consumed and internal pressure decreases. After a set time interval, the final position of the meniscus is read against the calibrated scale. The difference between the initial and final readings gives the total distance the fluid has traveled, which corresponds to the volume of oxygen consumed.
The observer must avoid parallax error to ensure accuracy when reading the final position. Parallax occurs when the observer’s eye is not level with the meniscus, leading to an inaccurate reading against the scale. Therefore, the reading must always be taken at eye level, focusing on the lowest point of the concave curve formed by the fluid meniscus. This careful reading of the distance traveled is the foundation for calculating the organism’s respiration rate.
Converting Readings to Volume
The distance the fluid travels is a linear measurement that must be converted into a volume of gas consumed. This conversion uses the formula for the volume of a cylinder, since the capillary tube is cylindrical in shape. The volume of oxygen consumed is calculated using the formula \(V = \pi r^2 h\), where \(r\) is the internal radius of the capillary tube and \(h\) is the distance the fluid moved.
The internal radius, \(r\), is a fixed value determined by the specific respirometer apparatus. The calculated volume, usually expressed in units like cubic millimeters (\(\text{mm}^3\)) or cubic centimeters (\(\text{cm}^3\)), represents the total oxygen consumed during the measured time interval. Dividing this volume by the time interval yields the rate of oxygen consumption, which is the organism’s respiration rate.
A control respirometer is necessary to account for fluctuations in temperature or atmospheric pressure despite the water bath. This control is set up identically to the experimental respirometer but contains non-respiring material. The distance moved by the fluid in the control is then subtracted from the distance moved in the experimental setup, ensuring the final calculated volume is exclusively due to the organism’s biological activity.