CO2 artefact can distort estimate of microbial basal respiration rate in closed chambers

Soil microbial basal respiration is a proposed biological indicator of soil health and a key parameter in studying microbial carbon cycling. It is commonly quantified in closed-chamber incubations by measuring the increase in CO₂ concentration in the vial headspace over time compared to a known background. Assuming a linear CO₂ increase solely caused by microbial activity, respiration rate estimates derived between 1 and 24 hours should compare, but differences have been observed previously.

To investigate how and why estimates of microbial respiration rate vary with incubation duration and amount of soil, gas samples were collected at 12 time points over a 24-hour period for ten soils covering two texture categories and a gradient of organic carbon content.

Microbial respiration rate was on average 3.4-fold higher after 1 hour than after 24 hours. The apparent decline in microbial respiration over time was related to a violation of the assumption that the sample CO₂ concentration at the beginning of the incubation equals the assumed background in soil-free blanks. Follow-up experiments indicated that the dissolution of CO₂ in the soil solution during the pre-incubation can cause an initial peak in emissions at the start of the incubation (i.e. CO₂ artefact) through shifts in chemical equilibria caused by the method itself, which can be misinterpreted as high initial respiration rates.

Over time, the contribution of the method’s artefact decreases. Factors like soil moisture, amount of soil incubated, microbial activity rate, and chamber closure timing affect the artefact's magnitude. Optimising incubation duration and headspace-to-soil ratio (e.g. 24-hour incubation at 22 mL vial : 1 g soil) can mitigate the effect of the CO₂ artefact and produce unbiased estimates of microbial respiration rates.