Pb 10

Soil pH was measured in solution with 1 g soil sample in 5 mL deionized water and it was determined with a precise pH meter of Sartorius PB10 (Sartorius, Germany). Soil electrical conductivity (EC) was determined with an EC meter (DDS-307, Shanghai Precision Scientific Instruments Co., Ltd., China). Soil moisture was calculated as (fresh weight − dry weight)/dry weight × 100%. Soil bulk density was calculated as the ratio between air-dried soil mass and the soil volume (400 cm³).

Soil moisture content, pH, organic matter (OM), total nitrogen (TN), total phosphorus (TP), and total potassium (TK) were assessed. Moisture was estimated using the oven dry-weight method. Soil pH was measured using a glass electrode pH meter (Sartorius PB-10; Sartorius Scientific Instruments Co., Ltd., Beijing, China) in a suspension of 1 g soil in 5 mL distilled water. OM content was determined by the Walkey and Black method^{49 (link)}. The determination of TN, TP, and TK were carried out in accordance with the national standards of the People’s Republic of China. Briefly, 0.2 g of soil was added to 5 ml of perchloric acid and 5 ml of hydrofluoric acid for low-temperature digestion, followed by determination of TP and TK by the Mo-Sb colorimetric method and the flame photometer method, respectively. TN was extracted from 0.2 g of soil by digestion with 5 ml of concentrated sulfuric acid, and then determined by the Kjeldahl method. According to Wilson^{50 (link)}, heavy metal contaminated soil was analyzed by digestion of 0.5 g of soil with 10 mL of concentrated HNO₃, following the microwave-nitric acid method. The concentrations of Sb, As, Pb, Zn, Cr, and Cd in the soil samples were measured using inductively coupled plasma optical emission spectrometry.

After the RS measurements at the selected sites were concluded, surface soil samples (0–20 cm) were placed in polyethylene boxes and taken to the laboratory. After air-drying at room temperature for at least two weeks, the field-moist soil samples were sieved through a 2-mm nylon sieve to remove the plant roots, coarse debris and sand. The soil bulk density at the soil depth of 0–10 cm was measured using a cutting ring (5 cm in both diameter and depth).
The soil pH was determined in a 1:5 (w/v) soil: water slurry using a pH meter (Sartorius PB-10, Sartorius, Germany). The soluble salt (SS) content was determined by gravimetric method. The microbial biomass carbon (MBC) was determined using the fumigation-extraction method^{62 (link)}. The SOC was analysed by potassium dichromate oxidation titration^{63 (link)}. The total nitrogen (TN) content was measured using an Elemental Analyser (CHNOS Elemental Analyzer, Vario EL, Elementar, Germany). The available phosphorus and available potassium were measured using the Olsen method and flame emission spectrometry, respectively.

All the experimental lambs were slaughtered at 49 days old after a fasting period of 12 h. After the rumen contents were mixed evenly, the rumen pH was measured immediately using an acidity meter (Sartorius PB-10, Sartorius Biotech Inc., Gottingen, Germany), the mixed rumen contents were collected in sterile tubes and stored at-80°C for rumen microbial and fiber degradation rate analysis. The contents were filtered through four layers of sterilized medical gauze, packed in cryovials, and stored in-20°C refrigerator for ruminal fermentation and enzymic activity analysis. After sampling, the rumen was weighed, segments of the ruminal tissue were collected at the location of cranial ventral sac and fixed in 4% paraformaldehyde for morphology measurements.

All 144 soil samples were measured for all physicochemical properties described herein. Soil pH was measured in a solution of 1.00 g soil sample in 5 ml deionized water using a precise pH meter (Sartorius PB10; Sartorius, Germany). Soil electrical conductivity (EC) was determined from the same solution using an EC meter (DDS-307; Shanghai Precision Scientific Instruments Co., Ltd., China). Soil bulk density was calculated as the ratio between the air-dried soil mass and the soil volume (400 cm³, which was fixed by the soil cutting ring). Soil water was calculated as (fresh weight – dry weight)/dry weight × 100%. SOC, N, P and K of the bulk soil and 5 soil fractions (AI, AI+EO, SA, PT and SB) in 12 composite samples were measured. SOC content was determined using the potassium dichromate volumetric method (external heating method). N content was determined using the semi-micro Kjeldahl method. P and K contents were determined using the sodium hydroxide melt method. All the related methods were detailedly described by Bao⁷⁴ .