Samples were diluted 1:2 with CH3CN and 5 μL of each sample was applied to a HILIC column (Acclaim Mixed-Mode HILIC-1, 3 μm, 2.1 × 150 mm). Metabolites were separated at 30 °C by LC using a DIONEX Ultimate 3000 UPLC system and the following solvents: Solvent A consisting of 5 mM NH4OAc in CH3CN/H2O (5/95, v/v) and solvent B consisting of 5 mM NH4OAc in CH3CN/H2O (95/5, v/v). The LC gradient program was: 100% solvent B for 1 min, followed by a linear decrease to 40% solvent B within 5 min, then maintaining 40% B for 13 min, then returning to 100% B in 1 min and 5 min 100% solvent B for column equilibration before each injection. The flow rate was maintained at 350 μL/min. The eluent was directed to the hESI source of the Q Exactive mass spectrometer (QE-MS) from 1.85 min to 18.0 min after sample injection (Thermo Fisher Scientific Waltham, MA, USA). The scan range was set to 69.0 to 1000 m/z with a resolution of 70,000 and polarity switching (negative and positive ionisation). Peaks corresponding to the calculated metabolites masses taken from an in-house metabolite library (MIM +/− H+ ± 2 mmU) were integrated using TraceFinder software (Thermo Fisher Scientific, Waltham, MA, USA).
Q exactive mass spectrometer qe ms
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Q Exactive-Mass Spectrometer (QE-MS) is a high-resolution mass spectrometry instrument designed for accurate mass measurements and qualitative analysis. The core function of the QE-MS is to ionize, separate, and detect molecular ions based on their mass-to-charge ratio.
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Lab products found in correlation
Xbridge amide column, by Waters Corporation (6 mentions)
Ultimate 3000 uhplc, by Thermo Fisher Scientific (5 mentions)
Xbridge beh c18 column, by Waters Corporation (2 mentions)
Ultimate 3000 uplc system, by Thermo Fisher Scientific (1 mentions)
Tracefinder software, by Thermo Fisher Scientific (1 mentions)
Ultimate 3000 hplc, by Thermo Fisher Scientific (1 mentions)
7 protocols using q exactive mass spectrometer qe ms
1
HILIC-LCMS Metabolite Profiling
2
UHPLC-QE-MS Metabolite Separation and Detection
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Ultimate 3000 UHPLC (Dionex) was coupled to Q Exactive-Mass spectrometer (QE-MS, Thermo Scientific) for metabolite separation and detection. For additional polar metabolite analysis, a hydropHILIC interaction chromatography method (HILIC) with an Xbridge amide column (100 × 2.1 mm i.d., 3.5 μm; Waters) was used for compound separation at room temperature. The mobile phase and gradient information was described previously 37 (link).
3
Metabolomics and Lipidomics Analysis Protocol
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Ultimate 3000 UHPLC (Dionex) was coupled to Q Exactive-Mass spectrometer (QE-MS; Thermo Scientific) for metabolite separation and detection.
For additional polar metabolite analysis, a HILIC with an Xbridge amide column (100 × 2.1 mm internal diamenter [i.d.], 3.5 μm; Waters) was used for compound separation at room temperature. The mobile phase and gradient information were described previously (Liu et al., 2015 (link)).
For lipid analysis, a reverse-phase liquid chromatography method with an Xbridge BEH C18 column (100 × 2.1 mm i.d., 2.5 μm; Waters) at 40°C was used for compound separation. The column was employed with mobile phase A: acetonitrile:water (60:40, v/v) with 0.1% formic acid and 10 mM ammonium formate, and mobile phase B: Isopropanol:acetonitrile (90:10, v/v) with 0.1% formic acid and 10 mM ammonium formate. Linear gradient was: 0 min, 40% B; 1.5 min, 40% B; 5 min, 85% B; 12 min, 97% B; 16 min, 97% B; 16.5 min, 40% B, and 20.5 min, 40% B. Flow rate is 0.2 mL/min.
For additional polar metabolite analysis, a HILIC with an Xbridge amide column (100 × 2.1 mm internal diamenter [i.d.], 3.5 μm; Waters) was used for compound separation at room temperature. The mobile phase and gradient information were described previously (Liu et al., 2015 (link)).
For lipid analysis, a reverse-phase liquid chromatography method with an Xbridge BEH C18 column (100 × 2.1 mm i.d., 2.5 μm; Waters) at 40°C was used for compound separation. The column was employed with mobile phase A: acetonitrile:water (60:40, v/v) with 0.1% formic acid and 10 mM ammonium formate, and mobile phase B: Isopropanol:acetonitrile (90:10, v/v) with 0.1% formic acid and 10 mM ammonium formate. Linear gradient was: 0 min, 40% B; 1.5 min, 40% B; 5 min, 85% B; 12 min, 97% B; 16 min, 97% B; 16.5 min, 40% B, and 20.5 min, 40% B. Flow rate is 0.2 mL/min.
4
Metabolomics and Lipidomics Analysis Protocol
5
Metabolite Profiling by UHPLC-QE-MS
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Ultimate 3000 UHPLC (Dionex) is coupled to Q Exactive-Mass spectrometer (QE-MS, Thermo Scientific) for metabolite profiling. A hydrophilic interaction chromatography method (HILIC) employing an Xbridge amide column (100 × 2.1 mm i.d., 3.5 μm; Waters) is used for polar metabolite separation. Detailed LC method was described previously (Liu et al., 2014b (link)) except that mobile phase A was replaced with water containing 5 mM ammonium acetate (pH 6.8). The QE-MS is equipped with a HESI probe with related parameters set as below: heater temperature, 120°C; sheath gas, 30; auxiliary gas, 10; sweep gas, 3; spray voltage, 3.0 kV for the positive mode and 2.5 kV for the negative mode; capillary temperature, 320°C; S-lens, 55; scan range (m/z): 70 to 900; resolution: 70000; automated gain control (AGC), 3 × 106 ions. Customized mass calibration was performed before data acquisition.
6
UHPLC-QE-MS Metabolite Separation and Detection
7
HPLC-MS Metabolite Separation and Detection
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Ultimate 3000 HPLC (Dionex) with an Xbridge amide column (100 × 2.1 mm i.d., 3.5 μm; Waters) is coupled to Q Exactive-Mass spectrometer (QE-MS, Thermo Scientific) for metabolite separation and detection at room temperature. The mobile phase A reagent is composed of 20 mM ammonium acetate and 15 mM ammonium hydroxide in 3% acetonitrile in HPLC-grade water (pH 9.0), while the mobile phase B reagent is acetonitrile. All solvents are LC-MS grade, purchased from Fischer Scientific. The flow rate used was 0.15 ml/min from 0 to 10 min and 15–20 min, and 0.3 ml/min from 10.5–14.5 min. The linear gradient was as follows: 0 min 85% B; 1.5 min 85% B, 5.5 min 35% B; 10 min 35% B, 10.5 min 25% B, 14.5 min 35% B, 15 min 85% B, and 20 min 85% B.
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