The quantitative assay was performed on an Agilent 1260 series HPLC system equipped with an ELSD (Agilent Technologies, Palo Alto CA, USA), which was controlled by Agilent ChemStation software (B.04.03-SP1). The separation of analytes was conducted on a Waters XBridge HILIC column (4.6 × 150 mm, 3.5 μm) with a flow rate of 1.0 mL/min at 30°C. The mobile phases were acetonitrile (A) and water (B) with a gradient elution of 88% A at 0–1 min, 88–78% A at 1–10 min, 78–65% A at 10–20 min, 65–88% A at 20–20.1 min, and 88% A at 20.1–35 min. The injection volume was 5 μL. The drift tube temperature of ELSD was 50°C and the nitrogen cumulative flow rate was 1.0 mL/min.
Xbridge hilic column
Manufactured by Waters Corporation
Sourced in United States
The XBridge HILIC column is a high-performance liquid chromatography (HPLC) column designed for the separation and analysis of polar, hydrophilic compounds. It utilizes hydrophilic interaction liquid chromatography (HILIC) principles to enable the effective separation of these analytes.
Automatically generated - may contain errors
Lab products found in correlation
Prominence hplc system, by Shimadzu (2 mentions)
Agilent 1260 series hplc system, by Agilent Technologies (1 mentions)
Chemstation, by Agilent Technologies (1 mentions)
0.45 μm membrane filter, by Merck Group (1 mentions)
Lc 20ad pump, by Shimadzu (1 mentions)
L 7612, by Merck Group (1 mentions)
Spd 20a, by Shimadzu (1 mentions)
Hplc system, by Beckman Coulter (1 mentions)
Krudkatcher ultra in line filter, by Phenomenex (1 mentions)
6 protocols using xbridge hilic column
1
Quantitative Analysis of Analytes via HPLC-ELSD
2
HPLC Analysis Using Prominence System
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Shimadzu Prominence HPLC system (Shimadzu Corp., Kyoto, Japan) with an LC-20 AD pump and SPD-20A UV–Vis detector was used.
Waters XBridge™ HILIC column (100 mm × 4.6 mm, 3.5 μm particle size) (Ireland) was utilized to perform the proposed study. Mobile phase was degassed using Merck L-7612 solvent degasser.
Adjusting pH through the work was performed by Consort NV P-901 pH Meter (Belgium).
0.45 μm membrane filter (Millipore, Ireland) was utilized to filter MP.
3
HILIC-Based Metabolite Separation and Detection
4
HILIC-MS Analysis of Biomolecules
5
HILIC-MS/MS Quantification of MMAE
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Chromatographic separation was performed using an XBridge HILIC column (3.0 mm × 50 mm, i.d. 5 μm; Waters, Milford, MA, USA) with an injection volume of 20 μL. Gradient elution was performed with 95%–50% mobile phase B (0.01% FA and 2 mM NH4Ac in 98% ACN), whereas mobile phase A consisted of 0.01% formic acid (FA) and 2 mM NH4Ac in 50% MeOH. The gradient started at 95% B and linearly decreased to 50% B over 2.2 min. It was maintained at 50% for 0.8 min before returning to 95% (over 0.01 min), and reconditioned for 1 min at a flow rate of 0.5 mL/min at 35 °C.
A Triple Quad 5500 mass spectrometer equipped with an electrospray ionization (ESI) source operated in the positive ion model was used. The collision energy (CE) for MMAE and the innate standard D8-MMAE were 38 and 37 V, respectively. The declustering potential (DP) for MMAE and D8-MMAE was 180 V. The collision cell exit potential (CXP) was 40 V, dwell time 100 ms, ion source gas 1 (GS1) 45 psi, curtain gas (CUR) 40 psi, collision gas (CAD) 6 psi, entrance potential (EP) 10 V, source temperature 400 °C, and IS 5500. Multiple-reaction monitoring (MRM) was performed to quantify MMAE and the innate standard. The transition m/z 718.6/686.5 was detected for MMAE, and m/z 726.6/694.6 was used to detect the innate standard.
A Triple Quad 5500 mass spectrometer equipped with an electrospray ionization (ESI) source operated in the positive ion model was used. The collision energy (CE) for MMAE and the innate standard D8-MMAE were 38 and 37 V, respectively. The declustering potential (DP) for MMAE and D8-MMAE was 180 V. The collision cell exit potential (CXP) was 40 V, dwell time 100 ms, ion source gas 1 (GS1) 45 psi, curtain gas (CUR) 40 psi, collision gas (CAD) 6 psi, entrance potential (EP) 10 V, source temperature 400 °C, and IS 5500. Multiple-reaction monitoring (MRM) was performed to quantify MMAE and the innate standard. The transition m/z 718.6/686.5 was detected for MMAE, and m/z 726.6/694.6 was used to detect the innate standard.
6
HILIC Separation of Compounds
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Waters XBridge™ HILIC column (100 mm × 4.6 mm, 3.5 μm particle size) was used throughout the work. The wavelength was adjusted at 260 nm using a flow rate of 1.0 mL/min. MP composition was ACN: 0.1% triethylamine buffer (TEA) in ratio of (90:10, v/v), respectively. The pH was adjusted at 5.0 using 0.2 M orthophosphoric acid then filtered using a 0.45 μm membrane filter. The separation was performed at room temperature.
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