Sample preparation for the untargeted analysis of the serum samples by GC-MS was based on the method published by Trimigno et al. [38 ] using a GC-MS 7890B/MS5977A (Agilent Technologies, Santa Clara, CA, USA) with a CombiPAL autosampler (CTC-Analytics AG, Zwingen, Switzerland). After deconvolution, features from subjects that only appeared in less than three samples were eliminated (n = 4828 remaining features). A second manual integration was conducted on 54 selected features that showed significant differences between treatment group responses to the interventions (see Statistical Analyses). The areas were normalized with the isotopically-labelled fructose. The features demonstrating a significant treatment effect were searched in EI-Mass Spectral Library (NIST 2017, Gaithersburg, MD 20899–6410, USA), masslib-library (www.masslib.com ). The molecules demonstrating sufficient potential for identification were acquired from Sigma-Aldrich (Buchs, Switzerland) and analyzed by GC-MS.
Combipal autosampler
Manufactured by CTC Analytics
Sourced in Switzerland, United States, Germany, Belgium
The CombiPAL autosampler is a versatile and reliable laboratory instrument designed for automated sample handling and preparation. It is capable of performing various tasks, including liquid, gas, and headspace sampling, as well as sample injection into analytical instruments such as gas chromatographs (GCs) and liquid chromatographs (LCs).
Automatically generated - may contain errors
Lab products found in correlation
Amber glass vial, by Avantor (3 mentions)
J w db 1701, by Agilent Technologies (2 mentions)
Pegasus 2 tof mass spectrometer, by Leco (2 mentions)
J w db ffap column, by Agilent Technologies (2 mentions)
Cis 4 injector, by Gerstel (2 mentions)
Db 5 capillary column, by CTC Analytics (2 mentions)
Polyacrylate fibre 85 m, by Merck Group (2 mentions)
7890a gc system, by Agilent Technologies (2 mentions)
5975c ms detector, by Agilent Technologies (2 mentions)
Spme fiber, by Merck Group (2 mentions)
Pentadecane, by Merck Group (1 mentions)
Db 5ms column, by Agilent Technologies (1 mentions)
Magnetic crimp caps, by Gerstel (1 mentions)
S monovette, by Sarstedt (1 mentions)
Lithium heparin, by Sarstedt (1 mentions)
Agilent 6890n gas chromatograph, by Agilent Technologies (1 mentions)
Pegasus 3 time of flight mass spectrometer, by Leco (1 mentions)
Gc msd chemstation software, by Agilent Technologies (1 mentions)
Dvb car pdms fibre, by Merck Group (1 mentions)
Millex gp, by Merck Group (1 mentions)
42 protocols using combipal autosampler
1
Untargeted serum GC-MS analysis protocol
2
Quantification of ent-kaurene in R. toruloides
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Following growth of engineered R. toruloides cultures, the dodecane phase was sampled and diluted 1:40 into dodecane containing 40 mg/L pentadecane (Sigma-Aldrich, 76510), and analyzed by gas chromatography—mass spectrometry (GC–MS) using an Agilent 6890 Plus gas chromatograph (Agilent Technologies, G1530A, Santa Clara, CA) connected to an Agilent 5973 Network mass spectrometer (Agilent Technologies, G1099A). 1 µL of each sample was injected by a CombiPal autosampler (CTC Analytics, MXY 02-00B, Zwingen, Switzerland). Analytes were separated on a DB-5MS column (30 m long, 0.25 mm internal diameter, 0.25 μm film thickness, Agilent Technologies, 122-5532) using the following oven parameters: hold for 0.5 min at an initial temperature of 100 °C, followed by a temperature ramp of 30 °C/min to 250 °C, a ramp of 10 °C/min to 270 °C, and a ramp of 30 °C/min to 300 °C. The mass spectrometer was operated in selected ion mode, with target ions (m/z) of 70, 85, 139 and 154. A standard curve was generated by running ent-kaurene standards at concentration range of 5–80 µg/mL. Analysis was performed using Enhanced ChemStation (Agilent Technologies, MSD Chemstation E.02.00.493) with ent-kaurene peak areas normalized to peak areas for pentadecane. The ent-kaurene standard was a gift from Dr. Joe Chappell, University of Kentucky, Lexington, KY.
3
Quantifying Prilocaine Metabolite in Blood
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The main metabolite of prilocaine, o-toluidine, was determined in the headspace over arterial blood by means of HS-SPME-GC-MS measurements. Therefore, full blood was withdrawn in Li-Heparin-tubes (S-Monovette®, 2.7 mL, Lithium-Heparin, Sarstedt AG & Co. KG, Nürnbrecht, Germany), and 1 mL was transferred into 20 mL headspace vials filled with 3 mL phosphate buffer to prevent coagulation. The vials were closed with Teflon-coated rubber septa in combination with magnetic crimp caps (Gerstel GmbH & Co. KG (Mülheim/Ruhr, Germany). Preconcentration was performed by means of Carboxen/polydimethylsiloxane (CAR/PDMS)-SPME-fibres (75 µm, SIGMA, Bellefonte, PA, USA) using a CombiPAL autosampler (CTC analytics AG, Zwingen, Switzerland) with 3 min equilibration time at 42 °C and 7 min adsorption time. For the separation and detection of o-toluidine desorbed from the SPME fibre, a GC-MS system (Agilent 7980A/5975C inert XL MSD, Santa Clara, CA, USA) was used as described before [27 (link)]. Quantification was performed via 5-point calibration in the range of 66.3 to 1060.5 ppbV (0.32–5.07 ug/mL). Two replicates were analysed per concentration level. Relative standard deviation was 4.89% at 530.2 ppbV (n = 4) and calculated value was 99.0% (±4.83%) of spiked value. The limit of detection was determined according to standard practices from the analysis of blank samples (n = 10) (see Table 1 ).
4
Headspace Fingerprinting of Food Volatiles
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Samples were thawed overnight in the cooling room (4 °C). 2.5 g thawed sample and 2.5 ml saturated NaCl solution were mixed in a 10 ml amber glass vial (10 ml, VWR International, Radnor, PA, USA). The vials were tightly closed using screw-caps with silicon septum seal (Grace, Columbia, MD), homogenized and transferred to the cooling tray of the autosampler which was maintained at 10 °C. Headspace fingerprinting was conducted on a GC system (6890N, Keysight Technologies, Diegem, Belgium) coupled to a mass selective detector (MSD) (5973N, Keysight Technologies, Diegem, Belgium) and equipped with a CombiPAL autosampler (CTC analytics, Zwingen, Switzerland) . Targeting detection of as many as possible volatiles in a particular food extract, an HS-SPME-GC-MS method of analysis was optimized beforehand. In the selected method, the samples were incubated at 40 °C for 20 min under To minimize the phenomenon of fiber degradation, a new fiber was used for each treatment condition (thermal and HPHT). Per treatment condition, during analysis, the samples were randomized as a function of storage time. Possible fiber degradation was carefully monitored by analysis of a reference sample (blanched carrot samples), every 10 injections. Per storage time, each sample was analyzed six times.
5
Metabolomic Analysis of Leaf Samples
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Frozen leaf samples harvested after 4 h illumination were ground in 2 ml microcentrifuge tubes using stainless steel beads and a Retsch Mixer Mill MM 400 (Retsch, Haan, Germany) under cryogenic conditions. From these samples, metabolites were extracted, derivatized, and analyzed via gas chromatography–time-of-flight mass spectrometry (GC-TOF-MS) analyses as described before (Lisec et al., 2006 (link)). The GC-TOF-MS system consisted of a CTC CombiPAL autosampler (CTC Analytics, Zwingen, Switzerland), an Agilent 6890N gas chromatograph (Agilent Technologies, Santa Clara, CA, USA) and a LECO Pegasus III time-of-flight mass spectrometer running in EI+ mode (Leco Instruments, St. Joseph, MI, USA). Metabolites were identified by comparison with database entries of authentic standards (Kopka et al., 2005 (link); Schauer et al., 2005 (link)) using TagFinder software (Luedemann et al., 2012 ). The peak intensity of a representative fragment was normalized with that of the internal standard ribitol and sample fresh weight (FW) and referred to as relative abundance. The parameters used for the peak annotation are listed in Supplementary Table S1 according to Fernie et al. (2011) (link).
6
GC-MS Analysis of Organic Compounds
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An Agilent GC/MS system (7890-5975C, Agilent Technologies, Santa Clara, CA, USA) was used in combination with a Combi-PAL autosampler (CTC Analytics, Zwingen, Switzerland) for the analysis of all samples. A 1 μL aliquot of sample was injected in splitless mode into the GC and separated on a fused-silica capillary column HP-5MS (30 m, 0.25 mm id, 0.25 μm thickness, Agilent J and W Scientific, Folsom, CA, USA). The injector temperature was set at 250 °C. High-purity helium was used as carrier gas at a constant flow rate of 1 mL/min. The column temperature was initially kept at 80 °C for 3 min, ramped to 320 °C at 10 °C/min, and then held for 10 min. The MS quadrupole temperature was set at 150 °C and the ion source temperature at 230 °C. Ions were generated by electronic impact (EI) at 70 eV. Masses were acquired from m/z 50 to 500. The filament was turned on after a solvent delay time of 6 min. The retention time of diphenylamine (the internal standard) was set to 11.7 min with the retention time locking mode. GC/MSD ChemStation software (Agilent Technologies, Santa Clara, CA, USA) was used for acquisition of the MS data. The measured mass spectra were compared with the National Institute of Standards and Technology (NIST) mass spectra library using the ChemStation software. Peaks with mass spectra similarity index greater than 70% were assigned compound names.
7
Headspace SPME-GC Analysis of Olive Oil VOCs
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The extraction of VOC from virgin olive oil samples was performed according to Vichi et al. (2003) by means of a Combi-pal autosampler (CTC Analytics, Zwingen, Switzerland) configured for HS-SPME. First, 2 g of olive oil sample was placed into a 10 mL glass vial fitted with a polytetrafluoroethylene (PTFE)/silicone septum (Scharlab, Barcelona, Spain)). Then, the vial was kept at 40 °C under agitation for 10 min, for sample conditioning. After that, a divinylbenzene/carboxen/polymethylsiloxane (DVB/CAR/PDMS) fibre (2 cm length, 50/30 thickness) from Supelco (Bellefonte, PA, USA) was exposed for 30 min to the sample headspace to extract VOC. Finally, the analytes were desorbed by placing the fibre in the gas chromatograph injector port (260 °C) with an SPME injector sleeve (0.75 mm ID) for 10 min, maintaining it for the first 5 min in split-less mode.
8
Toluene concentration analysis via GC-MS
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Toluene concentrations were measured via headspace analysis on a Trace DSQ GC/MS (Thermo Electron, Germany) equipped with a Combi PAL autosampler (CTC Analytics, Switzerland) as previously described (Avramov et al., 2013 (link)). A DB5-MS capillary column (Agilent Technologies, Germany) and helium as carrier gas at a flow rate of 1 ml min−1 were used. Briefly, 1 ml of slurry was taken with glass syringe from each serum bottle and transferred to a 2 ml vial. Ethylbenzene was added with a glass syringe to a final concentration of 2.3 mg l−1 as internal standard, and then the vial was immediately capped. Standard vials containing different concentrations of toluene (0, 0.25, 0.5, 0.75, 1.0, and 1.5 mM) but with the same amount of internal standard were prepared in the same way. Both the sample and standard vials were shaken for 17 min at 70°C to reach toluene liquid-headspace equilibrium. Then, 100 μl of headspace gas was injected into the GC-MS. Toluene concentrations were calculated according to the calibration standard.
Liquid samples filtered through a 0.45 μm filter (Millex-GP; Merck Milipore, Germany) were used for nitrate and nitrite quantification with ion chromatography using a coupled Dionex ICS 1100 system, equipped with an AS4A 4 × 250 mm and a CS12 A 4 × 250 mm column for anions and cations, respectively.
Liquid samples filtered through a 0.45 μm filter (Millex-GP; Merck Milipore, Germany) were used for nitrate and nitrite quantification with ion chromatography using a coupled Dionex ICS 1100 system, equipped with an AS4A 4 × 250 mm and a CS12 A 4 × 250 mm column for anions and cations, respectively.
9
Serum Metabolite Profiling by GC-TOF/MS
10
GC×GC-TOFMS method for comprehensive analysis
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A 6890 GC (Agilent) was equipped with Combi PAL autosampler (CTC Analytics), a CIS 4 injector (Gerstel, Mülheim an der Ruhr, Germany), and a J&W DB-FFAP column, 30 m × 0.25 mm i.d., 0.25 µm film thickness (Agilent) used as first column. The carrier gas was helium at a constant flow of 2.0 mL/min. The injection volume was 1 µL. The initial oven temperature of 40 °C was held for 2 min, followed by a gradient of 6 °C/min to a final temperature of 230 °C. The end of the column was connected to a second column, J&W DB-1701, 2.7 m × 0.18 mm i.d., 0.18 µm film thickness (Agilent). A liquid nitrogen-cooled dual stage quadjet thermal modulator was installed at the beginning of the second column and operated with a modulation period of 4 s. The major part of the second column was installed in a secondary oven mounted inside the primary GC oven. The initial oven temperature of the secondary oven was 45 °C and was held for 2 min, followed by a gradient of 6 °C/min to a final temperature of 240 °C. The end of the column was connected to a Pegasus II TOF mass spectrometer (Leco, Mönchengladbach, Germany) via a heated (250 °C) transfer line. Mass spectra were generated in the electron ionisation (EI) mode at 70 eV with a scan range of m/z 35 -350 and a scan rate of 100 spectra/s. For data analysis, the GC Image (Lincoln, NE, USA) software was used.
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