Tissues of overnight (14 h) fasted mice were homogenized and extracted twice with CHCl3/MeOH/H2O (2:1:0.6, v v−1v−1) containing 1-heptadecanoyl-rac-glycerol (C17:0-MG, Avanti Lipids, Alabaster, AL, USA) as an internal standard. The lipid-containing organic phase was dried and monoglycerides (MGs) were isolated by solid phase extraction using silica gel columns. Fractions were obtained by consecutive elution with 99/1 and 90/10 CHCl3/MeOH (v v−1). The latter fraction containing MGs was used for 2-AG quantification using an AQUITY-UPLC (Waters, Manchester, UK) equipped with a BEH-C18-column (2.1 × 150 mm, 1.7 μm; Waters) coupled to a SYNAPT™ G1 qTOF high-definition mass spectrometer (Waters) equipped with an electrospray ionization sources. Quantifier ions were MNa + (m/z 401) for 2-AG and MNa + (m/z 367) for C17:0-MG respectively. Concentrations of 0.5–100 pmol·μL−1 2-AG were used for the calibration curve together with C17:0-MG.
Synapt g1 qtof high definition mass spectrometer
Manufactured by Waters Corporation
Sourced in United Kingdom
The SYNAPT™ G1 qTOF high-definition mass spectrometer is a laboratory instrument designed for high-resolution mass analysis. It utilizes quadrupole time-of-flight (qTOF) technology to provide accurate mass measurements and detailed structural information about molecular samples.
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2 protocols using synapt g1 qtof high definition mass spectrometer
1
Quantification of 2-Arachidonoylglycerol in Mouse Tissues
2
UHPLC-qTOF MS Analysis of Hydromethanolic Extracts
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The hydromethanolic extracts were analyzed on an ultrahigh-performance liquid chromatography-quadrupole time-of-flight MS instrument (UHPLC-qTOF SYNAPT G1 system, Waters Corporation, Manchester, UK) with an Acquity HSS T3 C18 column (150 mm × 2.1 mm with particle size of 1.7 μm) (Waters, Milford, MA, USA). A binary solvent gradient [8 ] was followed to separate extracts. The chromatographic effluents were analyzed utilizing a SYNAPT G1 q-TOF high definition mass spectrometer with electrospray ionization (ESI) (Waters Corporation, Manchester, UK). To obtain fragmentation information of the ionized analytes, fragmentation experiments (MSE) were performed by ramping the collision energy from 15 to 60 eV [61 (link)]. An in-source collision-induced dissociation (ISCID) method was used to generate data for the annotation of closely related positional isomers of CGAs [8 ].
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