2.3.3. Setting and Monitoring of O2 and CO2 in Incubation

To determine the effect of oxygen on the volatile metabolism, anaerobic conditions were created by flushing the conical flasks with pure nitrogen gas (Figure 2a). The headspace gas sample, 1 mL, was tested to confirm the conditions by using GC-thermal conductivity detector (TCD). A stainless column packed with active charcoal (4 m × 3 mm) was used for oxygen and a stainless column with active aluminum (1 m × 1 mm) was used for carbon dioxide analysis. The flow rate of carrier nitrogen was 10 mL min−1, and oven temperature was 50 ◦C.

Headspace gas samples during incubation were tested periodically to confirm that there were no severe anaerobic conditions, O2 > 15 kPa and CO2 < 5 kPa when using air as the incubation gas.
