Analyses were conducted using an Agilent 6890 GC system coupled to an Agilent 5975 inert quadrupole mass spectrometer (Agilent, Santa Clara, CA, US) equipped with a Gerstel Thermo Desorption System (TDS2) and a CIS-4 PTV inlet Cooling Injector System (Gerstel, Müllheim an der Ruhr, Germany). The desorption temperature program was as follows: the temperature was kept at 35 °C for 0.1 min, and then ramped at 60 °C/min to 220 °C and held for 5 min. The temperature of the CIS-4 PTV injector, with Tenax TA inlet liner, was held at −35 °C using liquid nitrogen for the total desorption time, and was then raised to 260 °C at a rate of 10 °C/s and held for 4 min. The solvent vent mode was used to transfer the sample to the analytical column. A 50 m × 0.25 mm J&W CPWax-57CB column and a film thickness of 0.20 µm (Agilent, Santa Clara, CA, USA) were used; the carrier gas was He at a 1 mL/min flow rate. The oven temperature program was as follows: 35 °C for 4 min and then raised to 220 °C at 2.5 °C/min (held for 15 min). The quadrupole, source and transfer line temperatures were maintained at 150 °C, 230 °C and 280 °C, respectively. The electron ionisation mass spectra were recorded in the full-scan mode at 70 eV with the electron energy in the range of 29 to 300 m/z.
Agilent 6890 gc system
Manufactured by Agilent Technologies
Sourced in United States, Italy
The Agilent 6890 GC system is a gas chromatography instrument designed for precise, reliable, and efficient separation and analysis of complex sample mixtures. It features advanced hardware and software components to provide high-quality data and optimal performance.
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Lab products found in correlation
Db 17ms capillary column, by Agilent Technologies (2 mentions)
Agilent 5973n msd, by Agilent Technologies (2 mentions)
Db 5 capillary column, by Agilent Technologies (2 mentions)
Fluostar omega, by BMG Labtech (2 mentions)
Hp 5ms, by Agilent Technologies (2 mentions)
Methoxyamine hydrochloride, by Merck Group (1 mentions)
Db 5ms, by Agilent Technologies (1 mentions)
Mstfa, by Merck Group (1 mentions)
Pegasus 4d time of flight mass spectrometer, by Leco (1 mentions)
Agilent 5975 ms, by Agilent Technologies (1 mentions)
5973n mass selective detector, by Agilent Technologies (1 mentions)
J w db 5ms ultra inert, by Agilent Technologies (1 mentions)
5975b mass spectrometer, by Agilent Technologies (1 mentions)
5973n msd, by Agilent Technologies (1 mentions)
Ascorbic acid assay kit, by Abcam (1 mentions)
Optima 1ms column, by Macherey-Nagel (1 mentions)
Agilent 5973 network mass selective detector, by Agilent Technologies (1 mentions)
Agilent 5973 ms, by Agilent Technologies (1 mentions)
Db 5 fused silica capillary column, by Agilent Technologies (1 mentions)
21 protocols using agilent 6890 gc system
1
Comprehensive GC-MS Analysis of Volatile Compounds
2
Fatty Acid Profiling by GC-MS
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The total concentrations of palmitic acid (16:0), stearic acid (18:0), myristic acid (14:0), behenic acid (22:0), arachidic acid (20:0), gondoic (20:1), oleic (18:1), and linoleic (18:2) were determined from tissues by gas chromatography–mass spectrometry73 (link). A known quantity of tissue was hydrolyzed and extracted after adding a known amount of heptadecanoic acid (17:0). Fatty acids were analyzed as their trimethylsilyl derivatives under electron impact ionization mode using an Agilent 5973N-MSD equipped with an Agilent 6890 GC system and a DB17-MS capillary column (30 m × 0.25-mm internal diameter × 0.25-μm film thickness).
3
Lipid and Fatty Acid Analysis of Whole Fish
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Whole fish (5 fish per tank, 15 per dietary group) were analyzed for lipid content and fatty acid composition. In toto fish were minced, homogenized (homogenizer MZ 4110, DCG Eltronic, Monza, Italy), freeze-dried (Edwards EF4, Crawley, Sussex, England) and lipid extraction was carried out on lyophilized powders following a microwave-assisted extraction (MAE)55 (link),56 (link). Fatty acid methyl esters (FAMEs) were prepared according to Truzzi et al. (2017)56 (link), using the methyl ester of nonadecanoic acid (19:0; Dr. Ehrenstorfer GmbH, Augsburg, Germany) as internal standard. FAMEs were determined by an Agilent-6890 GC System (Milano, Italy) coupled to an Agilent-5973N quadrupole Mass Selective Detector (MSD) (Milano, Italy). A CPS ANALITICA CC-wax-MS (30 m × 0.25 mm ID, 0.25 μm film thickness) capillary column was used to separate FAMEs. Instrumental conditions for the studied matrices were set up, according to Truzzi et al. (2017)56 (link). For each sample, at least three runs were performed on the GC-MS. The precision of the proposed method evaluated as in Truzzi et al. (2014)57 (link) and the limits of detection (LOD) and quantification (LOQ) calculated as in Truzzi et al. (2014)58 (link), were as in Zarantoniello et al. (2018)11 (link).
4
GC-MS Analysis of Derivatized Metabolites
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The sample derivatization method is the same as previously described7 (link). The extracts were added with 40 µL pyridine solution of 20 mg/mL methoxyamine hydrochloride (Sigma-Aldrich), vortexed for 30 s and incubated at 30 °C and 130 rpm for 90 min in a gas bath shaker, followed by addition of 40 µL MSTFA (Sigma-Aldrich) with 1% chlorotrimethylsilane (TMCS, Sigma-Aldrich) and incubated at 37 °C for further 30 min. The derivatized samples were stored at 4 °C before detection.
The samples were analyzed with an Agilent 6890 GC system coupled with a Pegasus 4D time-of-flight mass spectrometer (Leco Corporation, St Joseph, MI, USA). The column was DB-5 MS (30 m × 250 µm i.d., 0.25 µm, Agilent J&W Scientific, Folsom, CA, USA), with an oven program set at 70 °C for 1 min, increased to 280 °C with a ramp rate of 5 °C/min holding for 15 min. Samples (1 μL) were injected with no split. The temperature of the inlet, interface and ion source was set as 250, 250, 220 °C, respectively. The solvent acquisition delay was 300 s. The mass spectrometer was operated in full scan mode (m/z 50–800) with an acquisition rate of 10 spectra/s. Electron impact ionization was set at 70 eV7 (link),52 (link).
The samples were analyzed with an Agilent 6890 GC system coupled with a Pegasus 4D time-of-flight mass spectrometer (Leco Corporation, St Joseph, MI, USA). The column was DB-5 MS (30 m × 250 µm i.d., 0.25 µm, Agilent J&W Scientific, Folsom, CA, USA), with an oven program set at 70 °C for 1 min, increased to 280 °C with a ramp rate of 5 °C/min holding for 15 min. Samples (1 μL) were injected with no split. The temperature of the inlet, interface and ion source was set as 250, 250, 220 °C, respectively. The solvent acquisition delay was 300 s. The mass spectrometer was operated in full scan mode (m/z 50–800) with an acquisition rate of 10 spectra/s. Electron impact ionization was set at 70 eV7 (link),52 (link).
5
Volatile Compounds Analysis by GC-MS
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The analysis of volatile compounds was conducted using the Agilent 6890 GC System equipped with the Agilent 5975 MS (Agilent Technologies, Santa Clara, CA, USA) and fitted with the DB-5 capillary column (30 m × 0.25 mm ID, 0.25 μm film thickness; J&W Scientific, Folsom, CA, USA). Ultrahigh purity helium (≥99.999%) was employed as the carrier gas with a constant flow rate of 1 mL·min−1, and 1 μl extraction of sample was injected at 250°C in splitless mode. The oven temperature was programmed at 40°C for 2 min and then ramped to 220°C at a rate of 6 °C·min−1 and held at 220°C for 5 min. The mass spectrometer was operated in the electron impact ionization mode at a voltage of 70 eV and ion source temperature at 230°C. Mass spectra were taken over an m/z range of 30–400. Retention indices were calculated after analyzing C8–C20n-alkane series under the same chromatographic conditions.
6
GC-MS Analysis of Honeysuckle Seed Oil
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The fatty acid composition in blue honeysuckle seed oil was analyzed using GC–MS. To increase the volatility of the seed oil, pre-column derivatization was used. In a 10 mL screw-cap glass tube, 30 µL of seed oil was dissolved in a mixed solution of n-hexane and benzene (1:1, v/v) and gently shaken. Then, 2 mL of 0.5 mol/L KOH in methanol was added, and the mixture was allowed to stand for 30 min. Subsequently, 5 mL of distilled water was added to separate the organic phase solution (n-hexane), causing it to rise to the upper layer of the tube. The top layer solution was diluted 20 times and analyzed by GC–MS on an Agilent 6890 GC system equipped with a 5973 N mass selective detector (Agilent Technologies Inc., Wilmington, DE, USA) using a DB-5 capillary column (60 m × 0.25 mm id, 0.25 µm film thickness, J&W Scientific, CA, USA). The GC injection port temperature was 250 °C, and helium was used as the carrier gas at a flow rate of 1.0 mL/min. The injection volume was 1.0 µL, and the injection split ratio was 1:10. The oven temperature was started at 180 °C and held for 5 min, and then ramped to 240 °C at 3 °C/min and held for 8 min. The total program time was 32 min. The ion source temperature was 230 °C. The MS detection was operated at 70 eV with a scan range of 50–550 m/z.
7
Fatty Acid Profiling of Zebrafish Larvae
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Experimental diets and F1 zebrafish larvae samples were analyzed (in triplicate) for fatty acid composition according to Zarantoniello et al. [8 (link)]. Briefly, samples were minced and homogenized (homogenizer MZ 4110, DCG Eltronic, Monza, Italy), and larvae were also freeze-dried (Edwards EF4, Crawley, Sussex, UK). Aliquots of 200 mg of each sample (three aliquots per sample) were added with 100 μL of Internal Standard (methyl ester of nonadecanoic acid, 99.6%, Dr. Ehrenstorfer GmbH, Augsburg, Germany), and lipid extraction was carried out on lyophilized powders following a microwave-assisted extraction (MAE) [54 (link)]. All lipid extracts were evaporated under laminar flow inert gas (N2) until constant weight and re-suspended in 0.5 mL of n-epthane. Fatty acid methyl esters (FAMEs) were prepared according to Canonico et al. [55 (link)] and were determined using an Agilent-6890 GC System (Milano, Italy) coupled to an Agilent-5973 N quadrupole Mass Selective Detector (MSD) (Milano, Italy) and separated through a CPS ANALITICA CC-wax-MS (30 m × 0.25 mm ID, 0.25 μm film thickness) capillary column [56 (link)]. For each analyzed aliquot of sample, at least three runs were performed on the GCMS.
8
Quantification of S-Nitrosoglutathione and Ascorbic Acid
9
Metabolite Extraction and GC-MS Analysis
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50 mg of liver sample was added to 150 μl of physiological saline for homogenate. Then 500 μl methanol were added, followed by 1 min of vortex mixing and 10 min of standing at -20 ℃ for protein precipitation. After centrifugation at 12000 g for 10 min at 4 ℃, 300 μl of the supernatant was transferred to an autosampler vial and blown to dryness with nitrogen. The residue was dissolved in 80 μl of methoxyamine (15 mg/ml) and the methoximation reaction was carried out for 90 min of shaking at 30 ℃, then 50 μl of BSTFA containing 1% TMCS was added for another 1 h of thimethylsilylation at 70℃. At last, a 1 μl of aliquot of the solution was injected into an Agilent 6890 GC system coupled with 5975B mass spectrometer (Agilent technologies, USA) for analysis.
Chromatographic separation was carried out on a capillary column (Agilent J&W DB-5ms Ultra Inert, 30 m * 0.25 mm * 0.25 μm) using programmed temperature showed inTable 1 . Parameters in mass spectrometer were as follows: scan range, m/z 30–550; temperature of injection, interface and source, 280 ℃, 260 ℃ and 230 ℃.
Chromatographic separation was carried out on a capillary column (Agilent J&W DB-5ms Ultra Inert, 30 m * 0.25 mm * 0.25 μm) using programmed temperature showed in
10
Fatty Acid Profiling of Fish and Diets
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The experimental diets and fish deprived of the viscera (15 per dietary group; n = 3) were analyzed for fatty acid composition. Samples were minced and homogenized (homogenizer MZ 4,110, DCG Eltronic, Monza, Italy), and freeze-dried (Edwards EF4, Crawley, Sussex, England). Lipid extraction was carried out with the Folch method (1957)52 (link) for experimental diets and with Microwave-Assisted Extraction (MAE) for fish53 (link). All lipid extracts were evaporated under laminar flow inert gas (N2) until constant weight and re-suspended in 0.5 ml of n-epthane. Fatty acid methyl esters (FAMEs) were prepared according to Canonico et al. (2016)54 using methyl ester of nonadecanoic acid (19:0; Dr. Ehrenstorfer GmbH, Augsburg, Germany) as internal standard. FAMEs were determined by an Agilent-6890 GC System (Milano, Italy) coupled to an Agilent-5973 N quadrupole Mass Selective Detector (MS) (Milano, Italy). A CPS ANALITICA CC-wax-MS (30 m × 0.25 mm ID, 0.25 μm film thickness) capillary column was used to separate FAMEs. Instrumental conditions for the studied matrices were set up, according to Truzzi et al., 201855 (link). For each analysed sample, at least three runs were performed on the GCMS.
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