Tsq 8000 triple quadrupole mass spectrometer

All extracts from entire tomato leaves and leaf tissue fractions were analyzed by combined gas chromatography–mass spectrometry (GC–MS) using a TRACE 1310 gas chromatograph system linked to a TSQ 8000 Triple Quadrupole mass spectrometer (Thermo Fisher Scientific, Waltham, MA, United States). Two μl of each sample was injected under a spitless mode, volatilized at 220°C, and then separated on a TraceGOLD TG-5MS GC column (30 m length, 0.25 mm I.D., and 0.25 μm film; Thermo Fisher Scientific, Pittsburgh, PA, United States). The initial column temperature was held at 40°C for 3 min and then ramped at 5°C/min to 120°C, 10°C/min to 180°C, and 20°C/min to 300°C which was maintained for 2 min. The helium carrier gas flow was 1.3 ml/min. All samples were analyzed using the total ion chromatogram (TIC) mode. Individual terpene compounds were identified by comparing their mass spectra (15–300 m/z) with those deposited in the NIST/EPA/NIH Mass Spectral Library (NIST11; National Institute of Standards and Technology NIST, Scientific Instrument Services, Inc., NJ, United States),² as well as those reported previously (Sallaud et al., 2009 (link)). Terpenes identified in the leaf and leaf tissue extracts were quantified using a previously determined average response factor for sesquiterpenes (Wang et al., 2020 (link)) in combination with the internal naphthalene standard.

Thawed serum smaples (450 μl) were spiked with ice cold methanol (1350 μl). After centrifugation of the mixture at 12000 rpm g for 15 min at 4 °C, the supernatant fraction was collected and divided into two parts: one (300ìl) for liquid chromatography- mass spectrometry (LC-MS) analysis and one (150 ìl) for gas chromatography -mass spectrometer (GC-MS) analysis after derivatization with BSTFA (containing 1% TMCS). Serum metabolic profiling analysis by LC-MS was performed as our previous study^{46 (link)}. Briefly, LC-MS analysis was performed on a Thermo Scientific Q Exactive hybrid quadrupole-orbitrap mass spectrometer coupled with a UPLC Ultimate 3000 system (Thermo Fisher Scientific, Bremen, Germany) equipped a heated electrospray source (HESI), at both positive and negative ion modes. LC-MS analysis was performed on a Thermo TRACE 1310 gas chromatograph system coupled with a Thermo TSQ 8000 Triple Quadrupole mass spectrometer. The quality control (QC) was pooled with same volume from each sample to ensure the reproducibility and stability druing the whole procedure.

Gas chromatography–mass spectrometry (GC-MS) untargeted analysis was used to find differences in potential metabolites between tumors and adjacent normal tissues in 40 OSCC patients. Untargeted analysis was performed by Trace 1310 Gas Chromato-graph equipped with an AS 1310 auto sampler, connected to a TSQ 8000 triple quadrupole mass spectrometer (Thermo Scientific, Waltham, MA, USA). Materials and specific steps for GC–MS analysis were based on the previously described method [18 (link)].

Each 50 μL aliquot of plasma was mixed with 200 μL methanol containing 50 μg 1,2-¹³C₂-myristic acid as an internal standard.The mixture was vortexed for 3 min and centrifuged at 20,000 × g for 10 min. One hundred microliters of supernatant were dried in a SpeedVac sample concentrator at 45 °C for 2 h. The dried aliquots were combined with 30 μL methoxyamine hydrochloride in pyridine (10 mg/mL), then vortexed for 3 min and shaken at 30 °C for 90 min with the Thermo Mixer C (Eppendorf, Hamburg, Germany). Thirty microliters of BSTFA containing 1% TMCS were added to the sample and shaken at 37 °C for 30 min. The mixture was then transferred to a sampler vial with a glass insert and subjected to GC–MS analysis. QC samples were prepared by pooling aliquots of all the plasma samples and were processed with the same procedure as that for the experiment samples. During analyses of the sample sequence, one quality control sample was run after every 10 injections.
Analysis was performed on a Trace 1310 Gas Chromatograph equipped with an AS 1310 autosampler connected to a TSQ 8000 triple quadrupole mass spectrometer (Thermo Scientific, Waltham, MA, USA) as described previously by Xie et al.^{55 (link)}. To detect and eliminate retention time shifts, the standard n-alkane mixture (C8–C40) was injected into the GC/MS during analysis of each batch of samples.

GC-MS analysis was performed using a Trace 1310 gas chromatograph coupled with TSQ 8000 triple quadrupole mass spectrometer (Thermo Fisher Scientific Inc., San Jose, CA, USA) equipped with an HP-5MS capillary column (30 m × 0.25 mm, 0.25 μm) (Agilent Technologies, Santa Clara, CA, USA). Ultrapure helium was used as the carrier gas with a flow rate of 1.2 mL/min. The temperatures of injector, MS transfer line, and ion source were set as 250 °C, 230 °C, and 230 °C, respectively. The injection volume was 1 μL, which was split in the ratio of 20:1. The oven temperature was programmed to hold initially at 60 °C for 2 min, then increase from 60 °C to 140 °C at 15 °C/min followed by a hold at 140 °C for 2 min, then increase from 140 °C to 180 °C at 5 °C/min followed by a hold at 180 °C for 3 min, and then increase from 180 °C to 250 °C at 10 °C/min with a final hold at 250 °C for 10 min. MS was performed in full scan mode with the mass range of m/z 100 to 700, using an electron impact ion source operated at 70 eV.

Cells constitutively grown in 2 mM or 4 mM glutamine were grown to 80% confluence. Media was removed and replaced with fresh media containing 100% of either 2 mM or 4 mM universally labeled ¹³C₅-glutamine for two hours at 37 °C prior to harvesting samples in 70% ethanol heated to 70 °C. As an internal standard, norvaline (1 µg norvaline/mL sample) was added to each sample, vortexed, and incubated at 95 °C for 5 min. Samples were cooled on ice for 5 min and centrifuged at 18,000× g for 5 min at room temperature. Cell pellets were analyzed for protein content with a bicinchoninic acid (BCA) assay (ThermoFisher, Waltham, MA, USA). Supernatants were dried and derivatized with methoxylamine hydrochloride in pyridine and prepared with N-tert-butyldimethylsilyl-N-methyltrifluoroacetamide with 1% (wt/wt) tert-butyldimethylchlorosilane for analysis with gas chromatography-mass spectrometry (Thermo TSQ 8000 triple quadrupole mass spectrometer coupled with a Thermo Trace 1310 gas chromatography) [44 (link)].

Sample extracts were analyzed with a Thermo Scientific TSQ‐8000 triple quadrupole mass spectrometer and a Trace GC 1300 gas chromatograph, which was equipped with a TriPlus AI 1310 autosampler (Thermo Fisher Scientific, San Jose, CA). Chromatographic separation was achieved with a J&W HP‐5MS capillary column (30 m × 0.25 mm, 0.25 μm film thickness). Helium (99.999% purity) was used as the carrier gas and maintained at a constant flow rate of 1.0 ml min⁻¹. A sample volume of 1.0 μl was injected in the splitless mode with an ion source temperature of 300°C. The transfer line to tandem MS was maintained at 280°C. The column temperature was programmed from 70.0°C (held for 1.0 min) to 200°C at a rate of 15°C min⁻¹, follow by 200°C to 260°C (held for 10 min) at a rate of 5 °C⋅min⁻¹. The tandem MS was operated in the multiple reaction monitoring (MRM) mode with an electron energy of 70 eV and an emission current of 30 μA. Argon gas (Ar) was chosen as collision gas with a pressure of 1.5 mTorr. The full‐scan mode with the mixed standard solution at 1.0 μg⋅ml⁻¹ was used to obtain the retention time (tR) and to select the most intense ions as optimal precursor ions.