Digiblock digester

The dried plant shoots (leaves, stems and pod walls) and seeds were ground to a powder. Nitrogen content and atom% ¹⁵N in the ground samples were determined using the elemental analyzer (FlashEA 1112, Thermo Scientific, Italy) coupled with the isotope ratio mass spectrometer Delta V Advantage (Thermo Scientific, Dreieich, Germany) and the continuous flow interface ConFlo III following manufacturer’s instructions. To determine Al and nutrient (B, Ca, Co, Cu, Fe, K, Mg, Mn, Mo, Ni, S, P and Zn) contents, the ground shoot samples were digested in a mixture of concentrated HNO₃ and 38% H₂O₂ at 70 °C using DigiBlock digester (LabTech, Sorisole, Italy). Rhizosphere soil samples (80 DAP) were incubated in deionized water (5 g soil + 25 mL water) at continuous shaking for 1 h. Soil samples were centrifuged at 10,000 g for 10 min and supernatants tested for pH_H2O using pH meter F20 (Mettler-Toledo, Schwerzenbach, Switzerland). Then soil supernatants were acidified with concentrated HNO₃ up to 1% to suppress microbial activity. Elemental content of digested plant samples and soil supernatants were determined using an inductively coupled plasma emission spectrometer ICPE-9000 (Shimadzu, Kyoto, Japan) following manufacturer’s instructions.

Concentrations of sodium, potassium and phosphorus in the roots and first and second mature leaves of wheat plants were assessed using an ICPE-9000 inductively coupled plasma emission spectrometer (Shimadzu, Kyoto, Japan) [31 (link)]. For this purpose, plant samples were digested in a mixture of concentrated HNO₃ and 38% H₂O₂ at 70 °C using a DigiBlock digester (LabTech, Sorisole, Italy).

Dried plant shoots were ground into powder. To determine their Cd and nutrient—P, K, Ca, Mg, S, Fe, B, Mn, Co, Cu, Mo, Na, Ni, and Zn—contents, ground shoot samples were digested in a mixture of concentrated HNO₃ and 38% H₂O₂ at 70°C using the DigiBlock digester (LabTech, Sorisole, Italy), and the content of each element measured by an inductively coupled plasma emission spectrometer ICPE-9000 (Shimadzu, Tokyo, Japan). Total nitrogen in shoot samples was determined using a Kjeltec 8200 Auto distillation unit (FOSS Analytical, Hillerød, Denmark). Statistical analysis of the data was performed using the STATISTICA v10 (StatSoft Inc., Tulsa, OK, USA). ANOVA analysis and Fisher’s least significant difference (LSD) test were used to evaluate differences between means.