3000 rs uhplc

Biochemical assays were analysed on a Bruker MaXis II ESI-Q-TOF-MS connected to a Dionex 3000 RS UHPLC fitted with an ACE C₄−300 RP column (100 × 2.1 mm, 5 μm, 30 °C), controlled using Bruker Otof control 4.0. The column was eluted with a linear gradient of 5–100% MeCN containing 0.1% formic acid over 30 min. The mass spectrometer was operated in positive ion mode with a scan range of 200–3000 m/z. Source conditions were: end plate offset at −500 V; capillary at −4500 V; nebuliser gas (N₂) at 1.8 bar; dry gas (N₂) at 9.0 L min⁻¹; dry temperature at 200 °C. Ion transfer conditions were: ion funnel RF at 400 Vpp; multiple RF at 200 Vpp; quadrupole low mass at 200 m/z; collision RF at 2000 Vpp; transfer time at 110.0 µs; pre-pulse storage time at 10.0 µs. All spectra were analysed using Bruker DataAnalysis 4.4. Measured masses for all species are displayed in Table S2 and S3.

After 24 h at 16°C, followed by 12 h at 4°C, the pH of 50 mL fermentation cultures was adjusted to pH 10 with formic acid. After centrifugation, 2 μL of the supernatant was analyzed directly with LC‐MS (column: Agilent Zorbax Eclipse RP‐C18, 100 × 2.1 mm, 1.8 μm). A Bruker MaXis Impact Q‐TOF mass spectrometer coupled with a Dionex 3000RS UHPLC was used. Mobile phases consist of A, water with 0.1% formic acid, and B, acetonitrile with 0.1% formic acid. After the initial 5 min of isocratic run at 5% B, a gradient of 5B–100% B in 18 min, then isocratic at 100% B for 5 min was employed with flow rate at 0.2 mL/min; UV was set at 210 nm. The mass spectrometer was operated in electrospray positive mode with a scan range of 50–3000 m/z. Source conditions are: end plate offset at −500 V; capillary at −4500 V; nebulizer gas (N2) at 1.4 bar; dry gas (N₂) at 8 L/min; dry temperature at 180°C. Ion transfer conditions as, ion funnel 1 RF at 200 Vpp; ion funnel 2 RF at 200 Vpp, hexapole RF at 200 Vpp; quadruple ion energy at 5 ev, quadrupole low mass set at 55 m/z; collision energy at 5.0 ev; collision RF ramping from 800 to 1500 Vpp; transfer time set from 100 to 155 μsec; pre‐Pulse storage time set at 5 μsec. Calibration was done with sodium formate (10 m mol/L) through a loop injection of 20 μL of standard solution at the beginning of each run.