| Instrument |
Q Exactive hybrid Q-Orbitrap-high resolution mass spectrometry (Q-Orbitrap-HRMS) (Thermo Fisher Scientific) |
| Instrument type |
UPLC-ESI-Q-Orbitrap |
| Ionization |
ESI |
| Ion Mode |
Positive and Negative |
| Description |
- Compound Extraction
In brief, the freeze-dried tissues were ground using a tissue grinder (catalog No. 05010997; Shanghai BiHeng Biotechnology Company Limited, Shanghai, China) for 2 min at 55 Hz. 150 mg of powder was weighed and mixed with 1.5 ml 70% aqueous methanol (containing 1 mg/L capsaicin as the internal standard). The samples were vortexed every 10 min for 3 times and extracted overnight at 4 °C. Following centrifugation at 12000 g for 15 min, all the supernatants were filtered using a 0.22 μm filter and transferred to a liquid phase bottle for subsequent analysis.
- Liquid chromatography (LC)-mass spectrometry (MS) analysis
Samples were analyzed on a Waters Acquity Ultra Performance Liquid Chromatography system equipped with an ACQUITY UPLC BEH C18 column (1.7 μm, 2.1 mm × 100 mm) coupled with Q Exactive hybrid Q-Orbitrap-high resolution mass spectrometry (Q-Orbitrap-HRMS) (Thermo Fisher Scientific).
|
| Solvent |
70% aqueous methanol |
| Column |
ACQUITY UPLC BEH C18 column (1.7 μm, 2.1 mm × 100 mm) |
| Total separation time |
30 min |
| Mobile phase |
(A) water with 0.04% acetic acid and (B) acetonitrile with 0.04% acetic acid |
| Gradient |
95:5 A/B at 0 min,5:95 A/B at 20.0 min, 5:95 A/B at 24.0 min, 95:5 A/B at 24.1 min, and 95:5 A/B at 30.0 min. |
| Flow rate |
0.25 ml/min |
| CollisionEnergy |
27.50(15eV,40eV) |