2.3. Sampling and chemical analyses
Soil samples were collected from each plot before the experiment and after the rice had been harvested to analyze the basic soil properties including pH, SOC, TN, NH4+-N, NO3-N, AP, AK, permanganate oxidizes carbon (POXC), and other available medium and trace elements. Soil samples were collected for determining NH4+-N and NO3-N contents at the seeding, tillering, elongation, earing, flowering, and maturation stages, respectively. Quadruplicate soil samples from a depth of 0−20 cm for each plot were collected and mixed thoroughly. The soil samples were air-dried at room temperature and then sieved to < 2 mm and/or < 0.15 mm after removing the residues and plant roots. Soil pH was measured with a pH meter (PH-21, Sartorius, Goettingen, Germany) in a 1:2.5 mass: volume soil/water suspension (Thomas, 1996). SOC content was determined using the potassium dichromatic oxidation titration method (Walkley & Black, 1934). TN content was determined using the Kjeldahl method (Pansu & Gautheyrou, 2006). AP was extracted with sodium bicarbonate solution and then was measured by the molybdenum antimony blue colorimetric method (Olsen, 1954). AK was extracted with 1 N ammonium acetate and then was determined by flame spectrometry (Jackson, 1958). POXC was measured using the colorimetric method after oxidating by 0.02 M KMnO4 (Blair, Lefroy, & Lisle, 1995). Available medium and trace elements were extracted with DTPA-CaCl2-TEA solution and then were measured using an ICP-OES (ICAP7000, ThermoFisher, USA) (Jackson, 1958). Soil NH4+-N and NO3-N were extracted with 2 M KCl and then were measured using a flow analyzer (San++System, SAKLAR, Netherlands). The shoot and/or grain of rice and wheat were also collected at seeding, tillering, elongation, earing, flowering, and maturation stages, respectively. The oven-dried and ground shoot and grain were digested with sulfuric acid-hydrogen peroxide, and the N content was determined by the Kjeldahl method. The grain yield and the grain moisture were measured during harvesting. Grain yield was adjusted to standard moisture of 15% for rice and 12% for wheat. The N agronomic efficiency (NAE, kg kg−1 N) was calculated as follow:
\begin{equation} \begin{matrix}\text{NAE}=\frac{Y_{N}-Y_{0}}{F_{N}}\#\left(\ AUTONUM\ \ \backslash*\ Arabic\ \right)\\ \end{matrix}\nonumber \\ \end{equation}
where \(\mathrm{Y}_{\mathrm{0}}\) and \(\mathrm{Y}_{\mathrm{N}}\) (kg ha−1) are the dry grain yields without N fertilizer and with N treatments respectively, \(\mathrm{F}_{\mathrm{N}}\) (kg N ha−1) is the N fertilization rate.