There are few methods available for broadly assessing microbial community metabolism directly within a groundwater environment. In this study, hydrogen consumption rates were estimated from in situ injection/withdrawal tests conducted in two geochemically varying, contaminated aquifers as an approach towards developing such a method. The hydrogen consumption first-order rates varied from 0.002 nM h−1 for an uncontaminated, aerobic site to 2.5 nM h−1 for a contaminated site where sulfate reduction was a predominant process. The method could accommodate the over three orders of magnitude range in rates that existed between subsurface sites. In a denitrifying zone, the hydrogen consumption rate (0.02 nM h−1) was immediately abolished in the presence of air or an antibiotic mixture, suggesting that such measurements may also be sensitive to the effects of environmental perturbations on field microbial activities. Comparable laboratory determinations with sediment slurries exhibited hydrogen consumption kinetics that differed substantially from the field estimates. Because anaerobic degradation of organic matter relies on the rapid consumption of hydrogen and subsequent maintenance at low levels, such in situ measures of hydrogen turnover can serve as a key indicator of the functioning of microbial food webs and may be more reliable than laboratory determinations.