| LC–MS sample conditions | Tris–HCl buffer, 50 mM, pH 8.0; 3-hydroxyanthranilic acid 1 mM; ATP 1 mM; MgCl₂ 10 mM; NatL2 1 µM; NatAM 2 µM; 30 °C and 1000 rpm for 30 min; stopped at 99 °C for 5 min |
|---|---|
| LC–MS (positive-ion mode) | Agilent 1260 Infinity II HPLC system coupled to an Agilent 6120 Quadrupole mass spectrometer; Nucleoshell RP 18 ec column, 100 × 2 mm, 2.7 µm, at 40 °C; water and methanol, each containing 0.1 % (v/v) formic acid; 0.3 mL min⁻¹; 2 % methanol from 0 to 3 min, 2–95 % methanol from 3 to 12 min, 95 % methanol from 12 to 14 min, 95–2 % methanol from 14 to 16 min, and 2 % methanol from 16 to 20 min; capillary voltage 3 kV; N₂ drying gas at 350 °C and 12 L min⁻¹; nebulizer pressure 35 psig |
| LC–MS chromatogram assignments | 3-hydroxyanthranilic acid (3-HAA), EIC at m/z 154.1; shunt amide product (3), EIC at m/z 289.1; benzoxazole (2), EIC at m/z 271.1; the identity of the benzoxazole 2 was confirmed by comparison of the retention time and mass spectrum with a reference standard generated in our previous study |
Julio Vieyto-Nuñez, Joel Mieres-Perez, Markus Nett, Lea Winand, Elsa Sanchez-Garcia. “Regulation of the enzymatic activity of a condensing amidohydrolase by organic solvents.” ChemRxiv (). https://doi.org/10.26434/chemrxiv.15007629/v1