Gas chromatography–mass spectrometry (GC–MS) measurement of relative metabolite levels and isotopic enrichment was performed as described28 (link), using an Agilent 7890B gas chromatograph equipped with a 30-m DB-35ms and a 5-m Duraguard capillary column (Agilent) for separation of derivatized metabolites, and an Agilent 5977B MSD system (Agilent) for measurement of metabolites. Briefly, dried metabolite extracts were derivatized with equal amounts of methoxylamine (20 mg ml−1 in pyridine) and MSTFA or MTBSTFA before injection into the GC–MS system. Measurements were carried out in either full scan mode or selected ion mode30 (link). Processing of chromatograms and the calculation of mass isotopomer distributions as well as relative quantification of metabolites were performed using the Metabolite Detector software31 (link).
Agilent 5977b msd system
Manufactured by Agilent Technologies
Sourced in United States
The Agilent 5977B MSD system is a gas chromatography-mass spectrometry (GC-MS) instrument designed for sensitive and reliable chemical analysis. It provides high-performance mass selective detection for a wide range of applications, including environmental analysis, food testing, and pharmaceutical research.
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Lab products found in correlation
7890b gas chromatograph, by Agilent Technologies (1 mentions)
30 m db 35ms, by Agilent Technologies (1 mentions)
Agilent 7890b gas chromatograph, by Agilent Technologies (1 mentions)
Agilent 7890b gas chromatography system, by Agilent Technologies (1 mentions)
Agilent 7890b gc system, by Agilent Technologies (1 mentions)
4 protocols using agilent 5977b msd system
1
GC-MS Analysis of Metabolite Levels
2
GC-MS Metabolite Profiling Protocol
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GC-MS measurement was performed as described (20 (link)), using an Agilent 7890B gas chromatograph equipped with a 30 m DB-35ms and 5 m Duraguard capillary column (Agilent, Santa Clara, California) for separation of derivatized metabolites, and an Agilent 5977B MSD system (Agilent) for measurement of metabolites, followed with data processing using the Metabolite Detector software (21 (link)).
3
Metabolomic Analysis of Microglia Samples
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We collected microglia from each group, treated the microglia with a mixed reagent of methanol/water = 4:1, and then lysed the cells with an ultrasonic cell disrupter. The samples were then analyzed by GC‒MS+LC‒MS according to the corresponding procedures.
Samples were analyzed in an Agilent 7890B gas chromatography system and an Agilent 5977B MSD system (Agilent Technologies Inc., CA, USA). HP-5MS quartz capillary columns were used to separate derivatives. Nitrogen was passed through the column at a flow rate of 1 mL/min. The temperature of the injector was controlled at 260°C. The initial oven temperature was 60°C, maintained for 30 s, gradually heated at a rate of 8°C/min, and finally maintained at 305°C for 5 min. The MS quadrupole and ion source were set to 150 and 230°C, respectively, with an impact energy of 70 eV. Finally, the generated raw data were analyzed by Progenesis QI v3.0 (Nonlinear Dynamics, Newcastle, UK) software and imported into the KEGG database, the Human Metabolome Database (HMDB), Lipidmaps (v2.3), and the EMDB2.0 database for characterization to obtain the results of Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, heatmap, and volcano map analyses regarding each group of differential metabolites.
Samples were analyzed in an Agilent 7890B gas chromatography system and an Agilent 5977B MSD system (Agilent Technologies Inc., CA, USA). HP-5MS quartz capillary columns were used to separate derivatives. Nitrogen was passed through the column at a flow rate of 1 mL/min. The temperature of the injector was controlled at 260°C. The initial oven temperature was 60°C, maintained for 30 s, gradually heated at a rate of 8°C/min, and finally maintained at 305°C for 5 min. The MS quadrupole and ion source were set to 150 and 230°C, respectively, with an impact energy of 70 eV. Finally, the generated raw data were analyzed by Progenesis QI v3.0 (Nonlinear Dynamics, Newcastle, UK) software and imported into the KEGG database, the Human Metabolome Database (HMDB), Lipidmaps (v2.3), and the EMDB2.0 database for characterization to obtain the results of Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, heatmap, and volcano map analyses regarding each group of differential metabolites.
4
GC-MS Analysis of Derivatized Samples
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The derived samples were analyzed using the Agilent 7890B GC system coupled with an Agilent 5977B MSD system (Agilent Technologies Inc., Santa Clara, CA, USA). An HP-5MS fused-silica capillary column (Agilent J&W Science, Folsom, CA, USA) was used to separate the derivatives. The carrier gas was helium (>99.999%) and the flow rate was set at 1 mL/min. The injector temperature was maintained at 260 °C. The initial oven temperature was 60 °C, which was maintained for 0.5 min, increased to 125 °C at a rate of 8 °C/min, increased to 210 °C at a rate of 5 °C/min, increased to 270 °C at a rate of 10 °C/min, increased to 305 °C at a rate of 20 °C/min and finally maintained at 305 °C for 5 min. The temperatures of the MS quadrupole and the ion source (electron incidence) were set to 150 and 230 °C, respectively. The collision energy was 70 eV. The pooled samples were injected at regular intervals throughout the analysis, to provide a dataset that can be used to evaluate repeatability.
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