2.4 Soil property analysis
Soil MBC and MBN were determined using the chloroform fumigation extraction method (Wu et al., 1990; Jenkinson et al., 2004). One subsample (20 g) was directly extracted with K2SO4 solution (60 mL; 0.5 M); another (also 20 g) was first fumigated with ethanol-free chloroform in the dark for 24 h and then extracted with K2SO4solution (60 mL; 0.5 M). The extracted C in the fumigated and unfumigated samples was measured after acidification using the Shimadzu TOC-VWP analyzer (Shimadzu), and inorganic N (NO3- and NH4+) was extracted with 2 M KCl and analyzed using a continuous-flow analyzer (Fiastar 5000; Foss Tecator AB, Höganäs, Sweden). The TN of the soil samples was determined using an elemental analyzer (MACRO cube, Elementar, Germany).
MBC and MBN were calculated as the difference in C and N contents between fumigated and non-fumigated sample extracts, adjusted by a proportionality coefficient (kEC = 0.45, kEN = 0.54) to account for the extraction efficiency. The 0.5 M K2SO4-extractable C from the unfumigated sample was considered as DOC (Vance et al., 1987).
The leaching water samples collected for NO3-(NO3--L) and NH4+(NH4+-L) were analyzed using a continuous-flow analyzer (Fiastar 5000; Foss Tecator AB, Höganäs, Sweden). Leaching water DOC (DOC-L) was measured using the Shimadzu TOC-VWP analyzer (Shimadzu).
The metabolic quotient (qCO2) and microbial carbon-use efficiency (CUE) were calculated using previously published methods (Anderson and Domsch, 1993; Sinsabaugh et al., 2013). qCO2 was defined as the ratio of C in respired CO2 and soil microbial biomass (qCO2 = CO2-C/MBC) and the CUE was defined as the amount of C in the microbial biomass relative to the sum of the CO2-C and MBC [CUE = MBC/(MBC + CO2-C)].