Identifying and quantifying attributes that help predict rates of heterotrophic soil respiration is a key issue. Similarly, assessing the temperature sensitivity (Q10) of soil C is critical to establishing if increases in Mean Annual Temperature will serve to further increase atmospheric CO2. Using organic soils from three sub-alpine communities that differ significantly in structure, species composition and productivity, we measured the respiratory quotient (RQ = rates of CO2 efflux/rates of O2 uptake) and temperature sensitivity of heterotrophic respiration during long-term (120 days) incubation. As a directly measurable parameter, RQ is free of empirical assumptions and provides an additional tool that can be used in conjunction with constants derived from fitted Arrhenius or exponential equations, to help understand shifts in microbial use of C substrates and how changes in vegetation might affect soil processes. Q10 did not change significantly over the course of a 120-day incubation for any of our studied soils. RQs varied with vegetation type and were consistently lower in grassland soils than woodland soils. RQs also varied during long-term incubations and declined consistently with time for grassland soils. RQs declined towards the end of the 120-day incubation for woodland soils. The generally low Ea for these soils from sub-alpine vegetation types in Australia, and the fairly rapid decline in RQ during incubation, suggest the likely greater temperature sensitivity of recalcitrant C relative to labile C could provide a strong positive feedback to increases in Mean Annual Temperature.