Epoch 2ns microplate spectrophotometer

Total phenolic content determination by Folin–Ciocalteu assay, was performed according to Swain and Hillis [49 (link)] with a few modifications. Briefly, 10 µL of the SCG extracts were mixed with 230 µL of distilled water and 10 µL of Folin–Ciocalteu reagent in a 96-well microplate. The mixture was incubated at 25 °C for 3 min. Lastly, 25 µL of 4 N Na₂CO₃ was added. The mixture was incubated for 2 h in the dark at 25 °C. After incubation, the absorbance at 750 nm was registered using an EPOCH 2NS microplate spectrophotometer (BioTek Instruments, Inc., Winooski, VT, USA). The calculations were completed using a gallic acid standard curve (from 0 to 0.4 mg/mL) and the results were expressed as milligrams of gallic acid equivalents per 100 g of dry sample (mg GAE/100 g). Measurements were made in triplicate (n = 3).

DPPH radical scavenging activity assay was performed according to Brand-Williams et al. [51 (link)], with slight modifications. Briefly, 20 µL of the SCG extracts were mixed with 280 μL of the DPPH reagent (100 µM in ethanol). After 30 min of incubation, the absorbance was recorded at 540 nm in an EPOCH 2NS microplate spectrophotometer (Biotek Instruments, Inc., Winooski, VT, USA). The results were calculated from a Trolox standard curve (from 0 to 400 µM) and were expressed as micromoles of Trolox equivalent per 100 g of dry sample (µmol TE/100 g). Measurements were made in triplicate (n = 3).

Haloferax volcanii strains were grown in Hv-YPC medium (Dyall-Smith, 2008 ) at 45°C with an optimal concentration of 18% salt water (2.46 M NaCl, 88 mM MgCl₂, 85 mM MgSO₄, 56 mM KCl, 12 mM Tris-HCl, pH 7.5). For low and high salt stress, salt concentration was adjusted to 15% and 23%, respectively, without modifying the ion ratios. While standard conditions (18% salt water) correspond to 15% NaCl, low salt (15% salt water) corresponds to 10.8% NaCl and high salt (23% salt water) corresponds to 19.2% NaCl. Growth of cultures was monitored in 96 well microtiter plates using an Epoch2 NS Microplate Spectrophotometer (BioTek Instruments, Bad Friedrichshall, Germany). Strains were precultured in Hv-YPC medium to OD_650nm of 0.4–0.7, after dilution to an OD_650nm of 0.05 they were transferred to microtiter plates. Cultures were incubated aerobically with orbital shaking at 45°C and OD_650nm was measured every 30 min. Outer wells were filled with salt water as evaporation barriers (Liao et al., 2016 (link)). The growth curves represent the average of at least three biological replicates.