Vario el 3 chnos elemental analyzer

For the determination of algal tissue C and N contents, 100 mL of cells was centrifuged at 12,000×g for 5 min at 4 °C and then the pellet was lyophilized at −50 °C. The dry algal sample of 0.01 g was used for the determination of C and N by the Elementar vario EL III CHNOS Elemental Analyzer (Elementar, Hanau, Germany).
The carbohydrate composition in wild type and mutant was determined using the modified quantitative saccharification (QS) method reported by the Nation Renewable Energy Laboratory (NREL), USA [43 (link)]. A small amount of dry cell powder (~0.1 g dry weight) was added to 3 mL of 72% (w/w) sulfuric acid and incubated for 60 min at 30 °C for the primary hydrolysis. The hydrolysate was then diluted to 4% (w/w) sulfuric acid and incubated for 20 min at 121 °C (sterilization) as the secondary hydrolysis. The supernatant was neutralized and analyzed by high performance liquid chromatography for the concentrations of reducing sugars present and the total carbohydrates obtained.

The elemental analysis of C, H, and O was carried out using a Vario EL III CHNOS Elemental Analyzer (Elementar, Langenselbold, Germany). The Cu content was determined based on edta titration in the presence of 1-(2-pyridylazo)-2-naphtol as an indicator [32 ]. Analysis for the studied compound [determined/theoretical (%)]: C 48.9/49.8; H 6.0/6.1; O 27.1/26.5; Cu 16.9/17.6. The FT-IR spectra were recorded on a Jasco FT/IR 6200 spectrophotometer (JASCO, Easton, MD, USA), in the form of KBr pellets, in the spectral range 4000–400 cm⁻¹, with resolution 1 cm⁻¹. The thermal decompositions were carried out with a Netzsch STA 449 F1 Jupiter thermoanalyzer (Netzsch-Geratebau GmbH, Selb, Germany) coupled with a Netzsch Aeolos Quadro QMS 403 mass spectrometer (Netzsch-Geratebau GmbH, Selb, Germany). Samples were heated in Al₂O₃ crucibles, in the temperature range 35–1000 °C, with the heating rate 10 °C/min in synthetic air (80% N₂, 20% O₂).

Plants were dried for 72h in an oven at 80 °C and then weighted and subsequently ground into powder in a mixer mill (Retsch MM400, Hann, Germany). The powder was digested with a mixture of nitric acid and hydrogen peroxide using a microwave system (MARS; CEM Corporation, Matthews, NC, USA), which was used to determine total Na contents by atomic emission spectrometry (ICP-AES, Thermo, Waltham, MA, USA). N contents were determined by a Vario EL III CHNOS Elemental Analyzer (Elementar Analysensysteme GmbH, Germany). The NO₃^– concentration was measured as previously described (Jampeetong and Brix, 2009 ). Dried material (5–10mg) was extracted with 10ml of Milli Q water at 80 °C in a water bath for 20min. Then, NO₃^– concentrations were analysed by an AA3 continuous flow-analyser (Seal Analytical, Germany).