2.4 Soil physicochemical properties
The air-dried soil samples were sieved through soil sieves with 2 mm, 1 mm and 0.25 mm aperture, and four groups of soil samples were obtained (>2 mm, 2-1 mm, 1-0.25 mm, <0.25 mm) (Muruganandam, Israel , & Robarge, 2009), use an electronic balance to weigh the weight of each particle size. The soil structural stability was evaluated by the mean weight diameter (MWD) of water-stable aggregate (Zhao, Chen, Hu , & Li, 2017). The content of easily extractable glomalin-related soil protein (EE-GRSP) and total glomalin-related soil protein (T-GRSP) was determined by improving the assay method according to Janos et al. (2008). The EE-GRSP was extracted from 0.5 g soil with 4 ml of 20 mmol L-1 of sodium citrate (pH=7.0) and autoclaved at 121℃ for 30 min. The T-GRSP was extracted from 0.5 g soil with 4 ml of 50 mmol L-1 of sodium citrate (pH=8.0) and autoclaved at 121℃ for 60 min.
The supernatant was collected by centrifugation at 10000 rpm for 10 min at 4℃. The T-GRSP extraction was performed 6 times until the solution was straw-colored ((Gao, Zhou, Ling, Hu , & Chen, 2017). An spectrophotometer (595 nm) was used to determine the GRSP content using bovine serum protein as the standard product.
The rest air-dried soil samples were used to determine soil pH by pH meter (FiveEasy FE20) on a mixture of water to soil ratio of 2.5:1. TC (total carbon), TOC (total organic carbon), and total N (TN) were determined by the analyzer (multi N/C 2100 TOC/TN Analyze, Germany, Analytikjene). Soil ammonium N (NH4+-N) and nitrate N (NO3--N) were determined by continuous flow analyzer (BRAN+LUEBBEAA3, Germany), and the extract solution was 2 mol·L-1 KCl, and soil total phosphorus (TP) and available phosphorus (AP) were determined by the molybdenum antimony anti-colorimetric method.