Evidence suggests that biochar is among ideal strategies for climate change mitigation and sustainable agriculture. However, the effects of soil aging on the physicochemical characteristics of biochar and nitrous oxide (N2O) production remain elusive. We set up a microcosm experiment with two greenhouse vegetable production (GVP) (alkaline and acid) soils by using the 15N tracing technique and quantitative polymerase chain reaction (qPCR) to investigate the mechanisms of N2O production as affected by fresh (FB) and aged biochar (AB) amendment. The results showed that AB increased the specific surface area, organic C, ammonium sorption capacity and cation exchange capacity, whereas decreased the pore size and pH relative to the FB. Results also demonstrated that FB effectively decreased N2O emissions from both soils while it enhanced the abundance of nirK and nosZI genes in alkaline soil and reduced the abundance of ammonia-oxidizing bacteria (AOB) amoA and increased nirK and nosZII genes in acid soil. In contrast, AB significantly stimulated nitrification and denitrification in both soils and thus significantly increased the N2O emissions by 43-78%. Furthermore, AB induced increases in ammonia-oxidizing archaeal (AOA) amoA and nirK gene abundances in alkaline soil and fungal nirK gene abundances in acid soil. These results suggest that AB may not be suitable for the mitigation of soil N2O emissions in GVP soils thus improving our understanding of the potential mechanism of biochar in N2O emissions.