Lignin phenols in soils were extracted and quanti ed using the alkaline copper oxide (CuO) oxidation method (Feng and Simpson, 2007) . Brie y, aired-dried soils containing at least ~ 5 mg organic carbon were mixed with 0.5 g CuO, 100 mg of ammonium iron (II) sulfate hexahydrate [Fe (NH 4 ) 2 (SO 4 ) 2 ⋅6H 2 O],
50 mg glucose (co-solvent), and 15 ml of nitrogen-purged 2 M NaOH solution in Te on-lined bombs. All bombs were ushed with N 2 in the headspace for 10 min and heated at 170°C for 2 h with constant stirring. After the heating, the bombs were cooled overnight. Then, 0.5 ml internal standard 1 (ethyl vanillin, 50µg/ml) was added to the oxidation products and mixed by stirring for 5 min. The oxidation products mixed with internal standards were transferred to a 50 ml centrifuge tube. The supernatant was transferred into a new centrifuge tube, after centrifugation at 3000 rpm for 15 min. The liquid was acidi ed with 6M HCl to a nal pH of 1.8-2.2, which was then incubated in the dark for 1 h at room temperature to precipitate humic acid. The supernatant was transferred to a 100 ml brown volumetric ask after centrifugation at 4000 rpm for 30 min. The precipitate was washed with HCl (v:v = 1:1000) and centrifuged at 4000 rpm for 30 min, then the supernatant was pooled with the supernatant from the last step. The combined supernatant was bound to a C 18 column and the column was dried by N 2 -blowing.
The bound product was eluted several times with ethyl acetate (0.5 ml each time, about 4.5 ml total). Then 0.5 ml of the internal standard 2 was added (Phenylacetic acid, 50µg/ml) to the collected eluent. After drying by N 2 -blowing, the product was derivatized by adding 50 µL of pyridine and 100 µL of derivatization reagent (N, O-bis-(trimethylsilyl) tri uoroacetamide) and incubated at 60°C, 3 h.
After cooled to room temperature, the derivatized products were analyzed by GC-MS equipped with HP-1 column (30 m × 0.25 mm × 0.25 µm) and a ame ionization detector (FID). The carrier gas was high purity N 2 (99.999%) operated with constant ow mode (1.5 ml min - 1 for N2, 40 ml min - 1 for H2, and 450 ml min - 1 for air). The inlet temperature was 300°C, the injection volume was 1 µl, a spilt injection was used (spilt ratio of 10:1), and the detector temperature was 300°C. The temperature program for the column was as follows: the initial column temperature was 100°C, and increased to 140°C at a rate of 8°C min - 1 , then increased to 170°C at a rate of 4°C min - 1 and hold for 5 min, nally increased to 300°C at a rate of 10°C min - 1 and held for 4 min. The concentrations of lignin phenols were quanti ed by comparing the peak intensity with the surrogate standards. Finally, the vanillyl phenols (V; vanillin, acetovanillone, vanillic acid), syringyl phenols (S; syringaldehyde, acetosyringone, syringic acid), and cinnamyl phenols (C; p-coumaric acid, ferulic acid) were summarized to represent the total lignin contents in the soil. The concentration of lignin phenols was normalized to SOC content to re ect its contribution to SOC.
50 mg glucose (co-solvent), and 15 ml of nitrogen-purged 2 M NaOH solution in Te on-lined bombs. All bombs were ushed with N 2 in the headspace for 10 min and heated at 170°C for 2 h with constant stirring. After the heating, the bombs were cooled overnight. Then, 0.5 ml internal standard 1 (ethyl vanillin, 50µg/ml) was added to the oxidation products and mixed by stirring for 5 min. The oxidation products mixed with internal standards were transferred to a 50 ml centrifuge tube. The supernatant was transferred into a new centrifuge tube, after centrifugation at 3000 rpm for 15 min. The liquid was acidi ed with 6M HCl to a nal pH of 1.8-2.2, which was then incubated in the dark for 1 h at room temperature to precipitate humic acid. The supernatant was transferred to a 100 ml brown volumetric ask after centrifugation at 4000 rpm for 30 min. The precipitate was washed with HCl (v:v = 1:1000) and centrifuged at 4000 rpm for 30 min, then the supernatant was pooled with the supernatant from the last step. The combined supernatant was bound to a C 18 column and the column was dried by N 2 -blowing.
The bound product was eluted several times with ethyl acetate (0.5 ml each time, about 4.5 ml total). Then 0.5 ml of the internal standard 2 was added (Phenylacetic acid, 50µg/ml) to the collected eluent. After drying by N 2 -blowing, the product was derivatized by adding 50 µL of pyridine and 100 µL of derivatization reagent (N, O-bis-(trimethylsilyl) tri uoroacetamide) and incubated at 60°C, 3 h.
After cooled to room temperature, the derivatized products were analyzed by GC-MS equipped with HP-1 column (30 m × 0.25 mm × 0.25 µm) and a ame ionization detector (FID). The carrier gas was high purity N 2 (99.999%) operated with constant ow mode (1.5 ml min - 1 for N2, 40 ml min - 1 for H2, and 450 ml min - 1 for air). The inlet temperature was 300°C, the injection volume was 1 µl, a spilt injection was used (spilt ratio of 10:1), and the detector temperature was 300°C. The temperature program for the column was as follows: the initial column temperature was 100°C, and increased to 140°C at a rate of 8°C min - 1 , then increased to 170°C at a rate of 4°C min - 1 and hold for 5 min, nally increased to 300°C at a rate of 10°C min - 1 and held for 4 min. The concentrations of lignin phenols were quanti ed by comparing the peak intensity with the surrogate standards. Finally, the vanillyl phenols (V; vanillin, acetovanillone, vanillic acid), syringyl phenols (S; syringaldehyde, acetosyringone, syringic acid), and cinnamyl phenols (C; p-coumaric acid, ferulic acid) were summarized to represent the total lignin contents in the soil. The concentration of lignin phenols was normalized to SOC content to re ect its contribution to SOC.
