Understanding the change in function of the biological community under different soil conditions is key to effective soil quality monitoring and mitigation of soil degradation. Current measures of biological soil quality suffer from drawbacks with most techniques having high expense, low throughput or a narrow focus on one component of the community. The aim of this study was to assess the use of volatilomics as a method to profile the soil microbial community and compare the technique to phospholipid fatty acid (PLFA) profiling as a measure of biological soil quality. An agricultural grassland soil (Eutric Cambisol) was subjected to a range of stresses in replicate laboratory mesocosms. Treatments included the imposition of hypoxia/anoxia by flooding with freshwater or saltwater in the presence or absence of plant residues. The volatile organic compound (VOC) and PLFA profile of each treatment was then compared to unamended mesocosms. We hypothesized that the VOC fingerprint of soil would be highly responsive to changes in microbial metabolic status/functioning and thus provide a complementary approach to PLFAs for evaluating soil biological health. We also hypothesized that the VOC profile would have greater discriminatory power than PLFAs for determining differences between soil treatments. A headspace solid phase microextraction (HSSPME) method coupled with gas chromatography quadrupole-time of flight mass spectrometry (GC/Q-TOFMS) was used to analyse the broad spectrum of VOCs produced by each soil. Across all soil treatments 514 unique VOC peaks were detected. Overall, VOCs showed greater sensitivity than the PLFA analysis in separating soil quality treatments. Eighteen individual VOCs were identified which were primarily responsible for this separation (e.g. indole, α-ionone, isophorone, 3-octanone, p-cresol, 2-ethyl-phenol). Anaerobic soils amended with residues showed the greatest separation from other treatments, with most of this differentiation associated with ten individual VOCs. The anaerobic soils also showed a significant reduction in the number of VOCs emitted but an increase in total VOC emissions. In conclusion, our findings provide evidence that soil VOCs rapidly respond to changes in soil quality and therefore hold great potential as a novel functionally relevant diagnostic measure of biological soil quality.