Xcalibur 2 2 sp1

After the chromatographic separations, using a Thermo Scientific™ Q Exactive™ hybrid quadrupole Orbitrap mass spectrometer equipped with a HESI-II probe, we could perform data acquisition. The positive and negative HESI-II spray voltages were 3.7 and 3.5 kV, respectively. The heated capillary temperature was set to 320°C. The sheath gas pressure was 30 psi. The auxiliary gas setting was 10 psi, and the heated vaporizer temperature was 300°C. Both the sheath gas and the auxiliary gas were nitrogen. The collision gas was also nitrogen at a pressure of 1.5 mTorr. The parameters of the full mass scan were as follows: a resolution of 70,000, an auto gain control target that was under 1 × 10⁶, a maximum isolation time of 50 ms, and an m/z range of 50–1,500. The LC-MS system was controlled using Xcalibur 2.2 SP1.48 software (Thermo Fisher Scientific), and data were collected and processed with the same software. All data that was obtained using both positive and negative ion modes were processed using the Progenesis QI data analysis software (Non-linear Dynamics, Newcastle, UK). The ranges of automatic peak picking for the C18 and HILIC assays were between 1 and 19 min and between 1 and 12 min, respectively. Next, the adduct ions of each “feature” (m/z, tR) were deconvoluted, and these features were identified in the human metabolome database (HMDB) and lipid maps.

Then the components of the TGQZD were analyzed by liquid chromatography/mass spectrometry (LC/MS)/MS instrument (Thermo Fisher). Briefly, DIONEX Ultimate 3,000 (Thermo Fisher) ultrahigh-performance liquid chromatography and Thermo Hypersil Gold C18 column (1.7 μm × 2.1 mm × 100 mm) were used to analyze sample. The mobile phase consisted of A (water, 2 mmoL/L ammonium formate, and 0.1% formic acid, v/v) and B (acetonitrile) with gradient elution. The gradient conditions for C18 separation was as follows: 100% A and 0% B, initial; 70% A and 30% B, 2 min; 30% A and 70% B, 9 min; 5% A and 95% B, 11 min; 0% A and 100% B, 12 min; 0% A and 100% B, 14 min; 100% A and 0% B, 14.1 min, 14 min; 100% A and 0% B, 16 min. The flow rate was 300 μl/min and the injection volume was 1 µl. The column temperature was maintained at 45°C. Mass spectrometry analysis was performed by the Q Exactive mass spectrometer (Thermo Fisher). The voltage of positive and negative ion source were 3.7 and 3.5 kv, respectively, and the heated vaporizer temperature was maintained at 320°C. Data were collected and processed by the Xcalibur 2.2 SP1.48 software (Thermo Fisher).

A Thermo Scientific™ Q Exactive™ Hybrid Quadrupole-Orbitrap Mass Spectrometer equipped with a HESI-II probe was employed. The positive and negative HESI-II spray voltages were 3.7 and 3.5 kV, respectively, the heated capillary temperature was 320°C, the sheath gas pressure was 30 psi, the auxiliary gas setting was 10 psi, and the heated vaporizer temperature was 300°C. Both the sheath gas and the auxiliary gas were nitrogen. The collision gas was also nitrogen at a pressure of 1.5 mTorr. The parameters of the full mass scan were as follows: a resolution of 70,000, an auto gain control target under 1 × 10⁶, a maximum isolation time of 50 ms, and an m/z range 50–1500. The LC-MS system was controlled using Xcalibur 2.2 SP1.48 software (Thermo Fisher Scientific, Waltham, MA, USA), and data were collected and processed with the same software.