Luna omega ps c18

Ultra-high performance liquid chromatography analysis was performed using a Waters Acquity H-Class UPLC system (Waters Corp, Milford, USA). Amino acids and sulfur contain metabolites separation was performed using a Waters UPLC HSS T3 column (2.1 × 100 mm, 1.8 µm). The mobile phase consisting of water containing 0.1% formic acid (A) and acetonitrile/methanol 50:50 v/v containing 0.1% formic acid (B) was applied with the optimized gradient elution as follows: 100% A at 0–1.5 min, 100–80% A at 1.5–2 min, 80–20% A at 2–2.5 min, 20% A at 2.5–4.5 min, 20–100% A at 4.5–5 min, 100% A at 5–7 min. The flow rate was kept at 0.4 mL/min, and column temperature was maintained at 25 °C.
The separation of organic acids, phosphorylated sugars, secondary metabolites and two amino acids (aspartic acid and glutamic acid) was achieved using a Phenomenex Luna^® Omega PS C18 (100 × 2.1 µm, 1.6 µm) column (Torrance, USA). The mobile phase consisting of water containing 0.5% formic acid (A) and methanol/water 70:30 v/v containing 0.5% formic acid (B) was applied with the optimized gradient elution as follows: 100% A at 0–1 min, 100–20% A at 1–4 min, 20–0% A at 4–6.5 min, 0% A at 6.5–7.5 min, 0–100% A at 7.5–7.9 min, 100% A at 7.9–10 min. The flow rate was kept at 0.3 mL/min, column temperature was maintained at 35 °C. The injection volume for both columns was 10 µL and samples were maintained at 10 °C.

Several adult specimens, collected from Mayotte and Australia, were kept in captivity and regularly milked for their predatory venom. Pooled samples from each geographic location were then fractioned by High Performance Liquid Chromatography, operated on an UltiMate^® 3000 Standard LC system (ThermoFisher Scientific, Waltham, MA, USA). First, crude venom was loaded onto a C₁₈ reversed-phase (RP) HPLC column Luna Omega PS C₁₈, 250 mm × 4.6 mm, 5 µm (Phenomenex, Torrance, CA, USA) and separated using a linear gradient water/acetonitrile (ACN) in 0.1% formic acid. Components were eluted at a flow rate of 1 mL/min and 1-min fractions were automatically collected in a 2 mL 96-well plate with the following gradient: 0–80% of ACN over 80 min for a total run of 105 min, at 40 °C. Venom fractions of interest were freeze-dried and resolubilized in 200 µL of pure water.