Xevo g2 s mass spectrometer

The rhIL-12 samples were denatured by 10mM DTT (Sigma, St. Louis, MO, USA) and deglycosylated by PNGase F (New England Biolabs, Beijing, China), then analyzed by the Acquity UPLC system connected online to a Xevo G2-S mass spectrometer (Waters Corporation, Milford, MA, USA). The column was a Waters BEH300 C4 column (2.1 mm × 50 mm, 1.7 μm particle). The flow rate was 0.2 mL/min using a gradient from 5% to 50% Solvent B (Solvent B being 0.1% formic acid in acetonitrile, Solvent A being 0.1% formic acid in water) in 7 min at a column temperature of 35 °C. The scan range of the mass spectrometric was m/z 500–3000. Data were acquired and processed using UNIFI 1.6 (Waters Corporation, Milford, MA, USA).

Optical rotations were measured on a Jasco P-2000 polarimeter (Jasco, Easton, MD, USA). IR spectra were recorded on a Perkin–Elmer FT-IR Spectrum Two spectrometer (Perkin Elmer, Boston, MA, USA). ¹H and ¹³C NMR spectra were recorded on an Agilent 500 (Agilent Technologies, Santa Clara, CA, USA) or on a Bruker 500 spectrometer (Bruker, Billerica, MA, USA), using CD₃OD as solvent. Chemical shifts were referenced using the solvent signals at δ_H 3.30 and δ_C 49.0. COSY, HSQC, HMBC, and NOESY experiments were performed using standard Agilent or Bruker pulse sequences. High-resolution mass spectra (HRESIMS) were obtained on a Waters XEVO G2-S Mass spectrometer (Waters, Milford, MA, USA). Column chromatography was carried out on Merck Silica gel 60 (70–230 mesh) (Merck, Darmstadt, Germany). SPE separations were performed on Supelco DSC18 cartridges (Supelco, Bellefonte, PA, USA). HPLC separations were performed on a LaChrom-Hitachi apparatus (Merck, Darmstadt, Germany) using a differential refractometer RI-71. Luna Si (2) (250 × 4.6 mm, 5 μm) (Phenomenex, Torrance, CA, USA) and Luna Si (2) (250 × 10 mm, 5 μm) (Phenomenex, Torrance, CA, USA) columns were used for separations in normal phase. All solvents were of HPLC grade.

The purified AlgM4 (0.25 mg/mL) enzyme was mixed with an equal volume of 1.0% (w/v) SA, polyM, or polyG; incubated in a water bath at 30 °C for 6 h; and then concentrated in vacuo. After high-speed centrifugation, the supernatants were collected and filtered through filter membranes with a 0.22 µm pore size. The filtrates were analyzed using liquid chromatography (LC)-MS. The equipment used in the analysis was a UPLC-QTOF-MS/MS system (Waters Corporation, Milford, MA, USA), which consisted of a UPLC I-Class instrument (Waters Corporation, Singapore) and a XEVO G2-S mass spectrometer (Waters Corporation, Milford, MA, USA). The operating conditions of LC were as follows: ACQUITY UPLC HSS T3 C18 column (2.1 mm × 100 mm, 1.8 μm, Waters Corporation, Milford, MA, USA); gradient elution with 0.1% formic acid-water (A) and formic acid-acetonitrile (B); flow rate, 0.5 mL/min; column temperature, 35 °C; analytic time, 10 min; injection volume, 1.0 μL. The MS conditions were as follows: ion scan mode, negative-ion ESI mode; scan range, 100–1700 Da; ion source temperature, 100 °C; desolvation gas temperature, 400 °C; desolvation gas flow rate, 1000 L/h; capillary voltage, 2.5 kV; cone voltage, 40 V; low collision energy, 6 V; high collision energy, 35–50 V; data acquisition software, MassLynx 4.1 SCN 884 (Waters Corporation, Milford, MA, USA); data acquisition mode, MSE.