Fatty acid compositions (oils) of the samples were determined by Gas Chromatography-Ion Trap Mass Spectrometry (GC/IT-MS), Thermo Scientific ITQ 900. Briefly, the FAME was analyzed by injecting 1 μL (30 : 1 split ratio) into GC-MS. The fatty acids were identified and quantified using a GC (Trace GC Ultra, Thermo Scientific) equipped with a capillary column (TR-FAME, 30 m × 0.25 mm, 0.25 μm film thickness) and an MS (ITQ 900, Thermo Scientific) attached to it. For separation of fatty acids, the oven temperature program was set as follows: 1 min initial hold at 50°C, temperature raised from 50 to 150°C at the rate of 20°C per min followed by a hold of 15 min at 150°C, temperature raised from 150 to 240°C at the rate of 20°C per min, and a final hold of 2 min at 240°C. Helium was used as a carrier gas with column flow rate of 1.0 mL per min. The transfer line and ion source temperatures were 250 and 220°C, respectively. The MS conditions were as follows: ionization voltage: 70 eV, range of 40–500 m/z, and the scan time equal to the GC run time. The individual constituents showed by GC were identified and quantified by comparing the retention times and peak areas to those of standards (ME-14-KT and ME-19-KT, SUPELCO Analytical) and by using the NIST Library (version 2.0, 2008).
Itq 900
Manufactured by Thermo Fisher Scientific
Sourced in United States
The ITQ 900 is a high-performance ion trap mass spectrometer designed for a wide range of analytical applications. It features a compact and robust design, advanced ion optics, and a high-resolution mass analyzer. The ITQ 900 provides accurate mass measurements and sensitive detection of a variety of analytes.
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Lab products found in correlation
Trace gc ultra, by Thermo Fisher Scientific (7 mentions)
Thermo trace gc ultra, by Thermo Fisher Scientific (1 mentions)
Dionex ultimate 3000 rslc system, by Thermo Fisher Scientific (1 mentions)
Db 5 capillary column, by Agilent Technologies (1 mentions)
Pva sil column, by YMC (1 mentions)
Trace gc ultra itq 900, by Thermo Fisher Scientific (1 mentions)
Factorfour capillary column vf 5 ms, by Agilent Technologies (1 mentions)
Triplus autosampler, by Thermo Fisher Scientific (1 mentions)
Db ffap, by Agilent Technologies (1 mentions)
Luna silica, by Phenomenex (1 mentions)
Ltq orbitrap xl, by Thermo Fisher Scientific (1 mentions)
Agilent 1100 hplc, by Agilent Technologies (1 mentions)
12 protocols using itq 900
1
Fatty Acid Composition Analysis by GC-MS
2
Kinetic Analysis of P450 119 Mutants
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The kinetics constants of P450 119 and its mutants were determined with thioanisole 1 and sulfoxide 2 as substrate. The reaction mixture consisted of 0.5 μM CYP119 mutant, variable concentrations of substrate and TBHP (10‐fold concentration of substrate). Phosphate buffer (pH 7.5) was added to the final volume at 100 μL. The reaction mixtures were incubated at 35 °C for 15 second (for the substrate thioanisole) or 30 second (for the substrate phenyl methyl sulfoxide) in a closed glass vials and stopped by addition of CH2Cl2 (100 μL). The products were analyzed by gas chromatography (Thermo Scientific ITQ900) on a DB‐1MS column (30 m ×0.25 mm inner diameter). The purified sulfoxide 2 and sulfone 3 were used as authentic standards to identify the retention time and to prepare calibration curve. The retention times found with this condition were ca. 11.2 min for sulfoxide and ca. 4.5 min for sulfone. The initial rates of formation of sulfoxide and sulfone were estimated at various concentrations of the substrate and analyzed by double‐reciprocal plot method to evaluate the kinetic parameters Km and kcat.
3
Herbal Extract Purification and GC-MS Analysis
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The herbal extract was dissolved in methanol (MeOH), then purified by solid-phase extraction using the QuEChERS method. For further analysis, GC (Thermo Trace GC Ultra, USA) and ITQ900 (Thermo, Waltham, MA, USA) were conducted. A TG-SQC capillary column (30 m × 0.25 mm × 0.25 μm) was utilized for the GCMS analysis. Helium (99.999%), a carrier gas, was set at a constant flow rate of 1 mL/min. The sample solution (1 μL) was injected in a split ratio of 10:1. The temperature of the injector and the ion source were set at 250 and 230 °C, respectively. The temperature program of the oven was set at 70 °C (isothermal for 2 min) and increased up to 280 °C with an increasing speed of 15 °C/min, ending with a 10 min isothermal at 280 °C. MS data were at 70 eV, a scanning interval time of 0.5 s, and for fragments from 50 to 650 Da. The compounds were identified via comparison with reported compounds using compounds data of the Mass Spectra Library (NIST 17.L and Wiley).
4
Phospholipid Separation and Fatty Acid Analysis
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Phospholipid separation by polar headgroup was performed on a Thermo Fisher Dionex UltiMate-3000 RSLC system. The separation of lipids was performed on a PVA-Sil column (150 × 2.1 mm I.D., 120 A) from YMC Europe GmbH thermostated at 35 °C. Chromatographic method was adapted from Ramos et al. [48 (link)]. For fatty acid quantification, phospholipids were digested and methylated using the one pot procedure described in [49 (link)]. Methylated fatty acids were analyzed in TraceGC Ultra coupled to an ITQ900 from Thermo Fisher equipped with an Agilent DB-5 capillary column. Further details are given in SI.
5
GC/MS Analysis of HACs Intermediates
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The intermediate products in the HDH of HACs were determined by GC/MS (Thermo Scientific ITQ 900) with a column of TR-5MS (30 m × 0.25 mm × 0.25 μm). The composition of the reaction/product mixture was analyzed by gas chromatography (Agilent 7890A), employing a flame ionization detector (FID) and a column of DB-1701 (30 m × 0.32 mm × 0.25 μm). Prior to analysis, the basic solution samples were neutralized with dilute CH3COOH (ca. 0.2 mol L−1). The detection limit of GC analysis for HACs was 1 pg mL−1. Taking 4-chlorophenol as a representative reactant, the conversion is defined as:
where C4-chlorophenol, 0 and C4-chlorophenol, t represent the initial concentration of 4-chlorophenol and the value at time t, respectively. And conversion rate is given as:
where C4-chlorophenol represents total amount of substance of 4-chlorophenol. The conversion rate is the average one within reaction time.
where C4-chlorophenol, 0 and C4-chlorophenol, t represent the initial concentration of 4-chlorophenol and the value at time t, respectively. And conversion rate is given as:
where C4-chlorophenol represents total amount of substance of 4-chlorophenol. The conversion rate is the average one within reaction time.
6
GC-MS Analysis of Sulfur Compounds
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Analysis was run on a Thermo Scientific ITQ-900 system (Trace GC Ultra/ITQ 900, Thermo, Milano, Italy) coupled with an ionic trap and fitted with a TG-XLB-MS capillary column with 0.25 μm film thickness, 30 m length, and 0.25 mm inner diameter, using helium as gas carrier with a flow rate of 1 mL/minute. The operating conditions were as follows: injector port temperature 250°C, split ratio 20:20, detector temperature 270°C, and program temperature from 40°C to 100°C with 4°C/minute, from 100°C to 150°C with 4°C/minute and from 150°C to 270°C with 10°C/minute.
The MS conditions were as follows: ionization voltage 70 eV, ion source temperature 200°C, scan range m/z 50–450. The qualitative identification of the sulfur compounds was based on computer matching with Wiley9 library and by comparison with data in the literature.
The MS conditions were as follows: ionization voltage 70 eV, ion source temperature 200°C, scan range m/z 50–450. The qualitative identification of the sulfur compounds was based on computer matching with Wiley9 library and by comparison with data in the literature.
7
GC-MS Analysis of Gut Samples
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Gas chromatographic
analysis was performed in a Trace GC ULTRA gas chromatograph coupled
to an ion trap mass spectrometer detector ITQ900 (Thermo Fisher Scientific),
using a Factor Four capillary column VF-5MS 30 m × 0.25 mm ID,
with 0.25 μm of film thickness (Agilent Technologies, Tokyo,
Japan). The column temperature was set to 50 °C during 1 min
and programmed to reach 310 °C at a rate of 10 °C per minute.
This temperature was maintained during 10 min. A constant flow rate
of 1 mL min–1 of helium as carrier gas was introduced
into the system. Moreover, ionization was performed by electronic
impact (EI) using 70 eV as voltage. The filament was off in the first
6 min of the chromatogram to avoid the signal of a large band from
the solvent and other nonseparated peaks. Finally, full scan mode
in the m/z range 35–650 was
monitored and 1 μL of gut sample was injected in splitless mode.
analysis was performed in a Trace GC ULTRA gas chromatograph coupled
to an ion trap mass spectrometer detector ITQ900 (Thermo Fisher Scientific),
using a Factor Four capillary column VF-5MS 30 m × 0.25 mm ID,
with 0.25 μm of film thickness (Agilent Technologies, Tokyo,
Japan). The column temperature was set to 50 °C during 1 min
and programmed to reach 310 °C at a rate of 10 °C per minute.
This temperature was maintained during 10 min. A constant flow rate
of 1 mL min–1 of helium as carrier gas was introduced
into the system. Moreover, ionization was performed by electronic
impact (EI) using 70 eV as voltage. The filament was off in the first
6 min of the chromatogram to avoid the signal of a large band from
the solvent and other nonseparated peaks. Finally, full scan mode
in the m/z range 35–650 was
monitored and 1 μL of gut sample was injected in splitless mode.
8
GC-MS Analysis of Fatty Acids
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The extract fraction was analyzed on a Trace GC Ultra gas chromatograph coupled to an ion trap mass spectrometer (GC–MS) ITQ 900 (Thermo Fisher Scientific, Waltham, MA). An Agilent HP-5MS fused silica column (30 m 9250 μm 90 25 μm) was used. The mass spectrometer detector operated in mass scanning mode was used for quantification. The chromatographic conditions were as follows: carrier gas, helium (1 0.1 mL/min); injection mode, splitless; injector and detector temperatures, 270 and 250 °C, respectively. The following program was used to analyze specific fatty acids: 70 °C for 35 min; ramp at 10 °C min 1 to 300 °C, and hold 5 min.
9
GC-MS Quantification of γ-Nonalactone
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The GC-MS (CI) analyses for the quantification of 𝛾-nonalactone were performed with a Trace Ultra GC and ITQ 900, which was equipped with a Triplus autosampler from Thermo-Fisher Scientific. The analytical column used was a DB-FFAP (30 m × 0.25 mm, film thickness 0.25 μm; Agilent J&W Scientific). An uncoated, deactivated fused silica capillary (2-5 m × 0.53 mm) was used as precolumn. Another uncoated fused silica capillary (0.3-0.6 m × 0.25 mm) was connected to the end of the analytical main column as a transferline into the ITQ. The carrier gas was helium with a flow rate of 1.0 mL min -1 . The chemical ionization gas was methane with a flow of 2.2 mL min -1 . The mass spectra were recorded at 70 eV in TIC (m/z range of 40-400) and in SIM mode. For SIM the selected quantifiers were 157 m/z for the unlabeled and 161 m/z for the labeled 𝛾-nonalactone. For the temperature program of the oven and the injection volume refer to Section 2.12.1.
10
Purification and Characterization of Natural Compounds
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1 H, 13 C, and 2D NMR spectra were recorded on a Bruker AVANCE III NMR spectrometer (Bruker, Billerica, Massachusetts, US), acquiring 1 H data at 400 MHz and 13 C data at 100 MHz, using standard experiments from Bruker pulse programs library. High-resolution mass spectrometry (HR-MS) was determined using an LTQ Orbitrap XL (Thermo Fisher Scientific, Waltham, Massachusetts, U.S.). The compositions of the essential oil were analyzed by an ITQ 900 mass spectrometer coupled to a TRACE GC Ultra gas chromatography (Thermo Fisher Scientific, Waltham, Massachusetts, U.S.). Methanol (MeOH) extracts were fractionated on silica gel 60 (230-400 mesh ASTM, Merck) and then purified with semi-preparative normal-phase column (luna silica (2), 250 × 10 mm, 5 μm, Phenomenex) on an Agilent 1100 HPLC (Agilent Technologies, Santa Clara, California, U.S.).
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