Culture broth of B. methylotrophicum in medium supplemented with corn steep liquor (CSL) or yeast extract (YE) was sampled at 0 h, 36 h (middle exponential phase), 60 h (later exponential phase), and 84 h (stationary phase), respectively. After the centrifugation at 13,000 g for 10 min, the supernatant was taken and filtered. 200 μL of supernatant was lyophilized under low temperature under low temperature (− 60 °C). The dried samples were derivatized at 40 °C for 160 min with 50 μL methoxamine hydrochloride (20 mg/mL in pyridine) and 80 μL N-methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA). Derivatized samples were subsequently analyzed using a TRACE 1310 GC system (Thermo Fisher Scientific, USA) combined with ISQ 7000 MS system (Thermo Fisher Scientific, USA), which was equipped with a fused-silica capillary column (30 m × 0.25 mm i.d., 0.25 μm DB-5MS stationary phase, J&W Scientific, Folsom, CA). Four replicates were performed for each sample.
Trace 1310 gc system
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Trace 1310 GC system is a high-performance gas chromatography instrument designed for comprehensive analysis of complex samples. It features a modular design, advanced electronic pneumatic controls, and intuitive software for precise and efficient separation, identification, and quantification of analytes.
Automatically generated - may contain errors
Lab products found in correlation
Q exactive gc orbitrap hrms detector, by Thermo Fisher Scientific (2 mentions)
Tracegold tg 5ms column, by Thermo Fisher Scientific (2 mentions)
Db5 ms stationary phase, by Agilent Technologies (1 mentions)
Allegra x 15r centrifuge, by Beckman Coulter (1 mentions)
Advanced ir vortex mixer, by VELP Scientifica (1 mentions)
Unity xr, by Markes International (1 mentions)
J w db 5ms, by Agilent Technologies (1 mentions)
Triplus rsh autosampler, by Thermo Fisher Scientific (1 mentions)
Q exactive orbitrap, by Thermo Fisher Scientific (1 mentions)
6 protocols using trace 1310 gc system
1
Metabolic Profiling of B. methylotrophicum
2
GC-HRMS Analysis Using Thermo Fisher Scientific Instruments
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A Trace 1310 GC system equipped with a TraceGOLD TG-5MS column (30 m, 0.25 mm, 0.25 µm), coupled to a Q-Exactive GC Orbitrap HRMS detector, all from Thermo Fisher Scientific (Waltham, MA, USA), were used for GC-HRMS analysis.
A Conterm laboratory oven, from J.P. Selecta (Barcelona, Spain); an Advanced IR vortex mixer, from VELP Scientifica (Usmate Velate, Italy); and an Allegra X-15R centrifuge, from Beckman Coulter (Indianapolis, IN, USA), were also used.
A Conterm laboratory oven, from J.P. Selecta (Barcelona, Spain); an Advanced IR vortex mixer, from VELP Scientifica (Usmate Velate, Italy); and an Allegra X-15R centrifuge, from Beckman Coulter (Indianapolis, IN, USA), were also used.
3
Volatile Organic Compound Profiling
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The VOC extraction protocol was carried out using a previously validated method[21 ]. Extraction of VOCs from TD tubes was performed using a thermal desorption unit (Unity™-XR, Markes International Ltd, Llantrisant, United Kingdom). The collected TD tubes were heated from 10°C to 250°C in split flow mode to release the VOCs collected inside. The VOC profiling was then analyzed by gas chromatography (GC) using the Thermo Scientific TRACE1310 GC system (Waltham, MA, United States), which is equipped with an HP-PLOT U gas chromatography column of 30 m length × 0.32 mm of inner diameter × 10 µm of film thickness (Agilent Technologies, Santa Clara, CA, United States). The GC column was initially programmed to heat at 40°C for 2 min and ramp to 130°C at the rate of 10°C/min for 10 min. Helium was used as the carrier gas at a flow rate of 1 mL/min during the VOC separation. The extracted VOCs then flowed through a transfer line at 130°C to final extraction using a FAIMS system (Owlstone Medical Lonestar VOC Analyzer FAIMS system, Cambridge, United Kingdom) (Figure 1B ). After final extraction, VOCs extracted from each participant’s group were shown in the form of chromatograms (Figure 1C ).
4
GC-Orbitrap HRMS Analysis of Compounds
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A Trace 1310 GC system equipped with a TraceGOLD TG-5MS column (30 m, 0.25 mm, 0.25 µm), coupled to a Q-Exactive GC Orbitrap HRMS detector, all from Thermo Fisher Scientific (Waltham, MA, USA), were used. The injection volume was 1 µL (splitless mode) and the inlet was set at 280 °C. The GC operated in constant flow mode at 1.2 mL min−1 of helium as carrier gas, using the following oven temperature program: 40 °C, held for 5 min; 5 °C min−1 up to 315 °C, held for 10 min. The MS transfer line was set at 300 °C. The EI ion source operated at 70 eV, and the ion source temperature was set at 250 °C. The acquisition was performed in full scan mode with a resolving power of 60,000 FWHM and a mass range from 40 to 500 m/z.
To perform retention index (RI) calculation, standard n-alkane mixtures, C8-C20 and C10-C40 (all even), were injected with the same conditions.
To perform retention index (RI) calculation, standard n-alkane mixtures, C8-C20 and C10-C40 (all even), were injected with the same conditions.
5
GC-MS Analysis of Organic Compounds
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A TRACE™ 1310 GC system was the chromatographic equipment with a TriPlus™ RSH autosampler (Thermo Scientific) and a J&W DB-5ms non-polar column (30 m × 0.25 mm × 0.25 μm) from Agilent Technologies, coupled to a Q-Exactive Orbitrap (Thermo Fisher Scientific) mass spectrometer. The injection volume was 1 µL. For chromatographic conditions, initial oven temperature was 60 °C (hold 2 min) and it was increased at 6 °C/min rate to 220°C (hold 2 min). Finally, it was raised to 280 °C with a 20 °C/min rate (hold 4 min). The total running time was 37 min. For MS conditions, full scan in positive mode was used (30–450 m/z range) with a 70-eV positive electron ionization (EI). The resolution was 70,000 FWHM, and an AGC value was set 106. Helium was used as carrier gas with a constant flow rate of 1 mL/min.
6
GC-MS Analysis of Plasma Metabolites
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Plasma samples were derivatized and analyzed by gas chromatography-mass spectrometry (GC/MS) using TRACE 1310 GC system connected to TSQ8000 triple quad (Thermo Fisher Scientific, Bremen, Germany) as described previously (Marczak et al., 2021 (link)). The identified compounds were quantified by applying extracted ion chromatograms (EIC) based on selected m/z values and retention times corresponding to specific compounds. The unique masses were chosen based on NIST library spectra and EIC peaks were integrated to measure their areas. The total ion current was measured by integrating all TIC peaks and used for compounds normalization.
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