G2 xs qtof

Chromatographic separation was performed by a Waters ACQUITY I-Class UPLC system equipped with an ACQUITY UPLC HSS T3 column (2.1 × 100 mm, 1.8 μm, Waters). Parameters were set as follows: column temperature 40°C; flow rate of 0.4 ml/min; injection volume of 4 μl; mobile phase A was H₂O containing 0.1% FA and B was ACN. The gradient elution condition was referenced by previous study (He et al., 2021 (link)): 0–1.00 min, 0% B; 1.01–4.00 min, 0–35% B; 4.01–15.50 min, 35–95% B; 15.51–18.00 min, maintaining at 95% B; 18.01–23.00 min, back to 0% B. MS detection and data acquisition were performed by a Q-TOF MS (Waters, Xevo, G2-XS QTof) in both positive and negative ion modes. MS parameters were set as follows: m/z range at 50–700 Da; capillary voltage at + 3.0 kV or −2.5 kV for positive or negative ion mode respectively; sampling cone at 40 V; source temperature at 110°C; desolvation temperature at 450°C; cone gas at 50 L/h, desolvation gas at 600 L/h. MassLynx (version 4.1, Waters) was used for data acquisition.

The screening and identification of RSV was performed with a Waters Acquity I Class UPLC system with the Xevo G2-XS QTof equipped with an ACQUITY UPLC BEH C18 column (Water, Milford, CT, USA) (1.7 µm, 2.1 × 50 mm i.d.). The chromatographic conditions included 0.01% formic acid in water as solvent A, and 100% acetonitrile as solvent B. The established elution gradient was 1% B (1 min), 1 to 20% B (10 min), 20 to 25% B (10 min), 35 to 50% B (5 min), and a column rebalancing flow rate of 0.25 mL min⁻¹. The analysis was carried out with an electrospray ionization (ESI) source, within a mass range of m/z 100 to 1200 Da, negative mode with a capillary voltage of 0.5 kV, cone gas flow of 30 L h⁻¹, desolvation gas flow of 900 L h⁻¹, source temperature of 140 °C, and a desolvation temperature of 450 °C, with a sample cone and source offset of 40 and 80, respectively. MS/MS experiments were carried out in conjunction with collision energy (CE) ramps: low CE off and high CE from 20 to 30 eV. All data were processed using UNIFI TM v1.8 software under the Mass Lynx NT 4.1 (Waters, Milford, USA) operating interface.

The headspace concentrations were measured at equilibrium as independent triplicates with a G2-XS Q-TOF high-definition mass spectrometer (XEVO, Waters) coupled to a patented Venturi interface [16] . An automated PAL system sampled 5 mL of headspace air with a 5 mL headspace syringe and injected this into the mass spectrometer. Mass spectra were collected in centroid mode over the range m/z 20-400 every 1 s. APCI-MS was performed in positive ionization mode with a cone voltage of 4.0 kV, source temperature of 105°C, heated sample transfer line temperature of 130°C and auxiliary gas flow of 600 Lh⁻¹. Lock spray (on-the-fly mass calibration) was used to apply a mass correction to measured m/z values during the analysis. All the signal intensities were corrected for the background addition. The relative headspace concentration (RHC) was calculated as: $\text{RHC}\text{\%}=\frac{\text{Peak area}{}_{\text{flavored dispersion}}}{\text{Peak area}{}_{\text{flavor in water}}}⁎100\text{\%}$

Nuclear magnetic resonance (¹H and ¹³C NMR) spectra were collected using Bruker ARX400 Nuclear Magnetic Resonance. The chemical shift was relative to tetramethylsilane as the internal standard. Resonance patterns were reported with the notation of s (singlet), d (doublet), t (triplet), q (quartet), and m (multiplet). High-resolution mass spectrum was measured on Waters G2-XS QTOF in a positive ion mode. High-performance liquid chromatogram spectrum was recorded on Agilent 1290II. UV–Visible absorption spectra were obtained using Perkin Elmer Lambda 35. Steady-state PL spectra and excitation spectra were measured using Hitachi F-4600. The delayed PL spectra and lifetimes were measured on an Edinburgh FLSP 980 spectrophotometer equipped with a xenon arc lamp (Xe 900) and microsecond flash-lamp (μF900), respectively. The PLQYs were measured on an Edinburgh FLSP 980 spectrophotometer equipped with an integrating sphere. The variable temperature emission spectra were measured on OLYMPUS BX53M with the constant excitation wavelength of 360 nm under nitrogen atmosphere. Single-crystal x-ray diffraction experiments were carried out using a Bruker D8 QUEST x-ray single crystal diffractometer. Luminous patterns of information encryption and display under different UV excitation wavelengths were taken by cell phone in professional mode.

Samples were extracted using methanol:water:chloroform as previously described with slight modifications^{9 (link)}. Briefly, untargeted extractions were reconstituted with 400 µl tributylamine with no dilutions for analysis. Analysis was performed using a Xevo G2-XS QTOF attached to a Waters UPLC (Waters, Milford, MA) with negative-mode electrospray ionization run in MS^E continuum mode. LC phases, gradient rates, and columns were used as previously published^{9 (link)}. For untargeted acyl-CoA analysis, MS^E continuum data was processed with Progenesis QI (Waters) to align features, peaks, deconvolute, and annotate metabolite peaks. Metabolite annotations were scored based on a mass error < 12 ppm to Human metabolome Database entries^{85 (link)}, isotopic distribution similarity, and theoretical fragmentation comparisons to MS^E high-energy mass spectra using the MetFrag option with each metric contributing a max of 20 points towards a max score of 60. Raw signals for each compound abundances were normalized to a correction factor calculated using a median and mean absolute deviation approach by Progenesis QI. Significance was determined by a one-way ANOVA adjusted for multiple comparisons with a Dunnett’s post-hoc test.

The reaction was performed in 200 mM potassium phosphate buffer (pH 8.0) containing 2.5 mM vinyl acetate, 0.5 mM cyanidin-3-O-glucoside, and 0.5 mg/mL purified enzymes. The reactions were incubated at 40 °C for 10 min. An ultrahigh-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) system (G2-XS QTof, Waters Milford MA USA) was used to analyze the molecular mass of the product. Three microliters of the solution was injected into the UPLC column (2.1 × 100 mm ACQUITY UPLC BEH C18 column containing 1.7 μm particles) with a flow rate of 0.3 mL/min. Mobile phase A was water containing 0.5% formic acid, while mobile phase B was acetonitrile. The gradient program of the mobile phase was as follows: 0–1 min, 5% B; 1–5 min, 5–90% B; 5–6 min, 90% B; 6–6.1 min, 90–5% B; 6.1–8 min, 5% B. UPLC-ESI-MS/MS was performed with a collision energy of 20 eV employing an electrospray source in positive ion mode with a selected mass range of 50–600 m/z. The ionization parameters included the following: capillary voltage of 3.0 kV, cone voltage of 40 V, source temperature of 550 °C, and dissolution gas temperature of 400 °C. Data acquisition and processing were performed employing Masslynx 4.1 (Waters, Milford, MA, USA) software package.