I want to measure metabolism rates in Tardigrades
Metabolic rates will be assessed immediately following their exit from cryptobiosis (an animal dormant state).
I know that oxygen levels is the usual way to measure metabolism, but tardigrades are extremely small and hence oxygen level differences will be minute. Is there any other way to characterize metabolic rates, or is there a way to make the oxygen method viable?
Metabolism in other small organisms, such as bacteria, are measured by taking the metabolism of a very large population of organisms, and then dividing that by the count of organisms in order to obtain the metabolic rate of a given individual.
In this paper on the metabolism of E. coli, they looked at the various input and output chemicals. Glucose, biomass, oxygen, and carbon dioxide were all observed. Observation of multiple variables also helps in the reduction of errors by using the stoichiometric balance to make sure that no errors have occurred in the measurements.
The methodology section of the paper had the following techniques:
Continuous cultivations were performed at 28 °C, at a stirring speed of 800 r.p.m., and the pH was controlled at pH 6·6 with 2·5 mol NaOH l−1. The 2-l bioreactor (model SGI 7F-Set2; Setric) was equipped with standard measuring and control units (temperature, pH, dissolved oxygen, stirrer speed). A constant aeration rate of 1·33 v.v.m. was achieved by a mass flow controller (Brooks). Continuous cultivation was performed for at least eight residence times at a given dilution rate; steady-state conditions were verified by constant optical density and oxygen and carbon dioxide transfer rates. Except for glucose, all other substrates were in excess. The concentration of dissolved oxygen was always maintained above 20 % of air saturation. Analytical techniques.
Culture liquid was withdrawn from the bioreactor into an ice-cooled beaker. Cell dry mass was determined gravimetrically after centrifugation of a known sample volume. The pellet was washed twice with 0·9 % w/v NaCl and dried at 40 °C under vacuum for 48 h. Elemental biomass composition was analysed with a PE 2400 II elemental analyser (Perkin Elmer). Glucose was measured enzymically by a YSI glucose analyser (model 2700; Yellow Springs Instrument Co.). Organic acids, i.e. acetic acid, formic acid, lactic acid and pyruvic acid, were analysed by HPLC using an HPX-87H Aminex ion-exclusion column (Bio-Rad). The column, connected to a UV detector (λ=210 nm) and RI detector, was eluted at 50 °C with 5 mmol H2SO4 l−1 at a flow rate of 0·5 ml min−1.
If the tardigrades are grown in solution, an identical methodology can be used to measure respiration rate.
