One gram of fresh weight of wild type cells or elo2Δelo3Δ double deletion mutant cells complemented by individual cotton KCSs were ground to a fine powder with a mortar and pestle. Fatty acid were extracted by sulfuric acid: methanol (1:40, v/v) and derivatized by heating to 85 °C for 2 h (Browse et al. 1986 ). After cooling to room temperature, fatty acid methyl esters were extracted three times with hexane, dried under nitrogen gas and then concentrated to 0.5 mL. C17:0 fatty acid (heptadecanoic acid; Sigma‐Aldrich), which does not exist in plants, was added before extraction to monitor sample recovery and also for quantitative analysis. 1 µL of each sample was analyzed on the Agilent 6890N GC system with the DB‐225MS column (J&W) that operated with hydrogen carrier gas and a splitless inlet (injection temperature, 280 °C), according to the manufacturer's instructions. All fatty acids and hydroxylated fatty acids were identified by the Agilent 6890N GC system coupled to an HP 5973 mass selective detector using both the National Institute of Standards, and Technology and Wiley databases.
Agilent 6890n gc system
Manufactured by Agilent Technologies
Sourced in United States
The Agilent 6890N GC system is a gas chromatography instrument designed for the separation, identification, and quantification of chemical compounds. It features advanced electronics, a high-performance oven, and precise temperature control to deliver reliable and reproducible analytical results.
Automatically generated - may contain errors
Lab products found in correlation
Hp innowax fsc column, by Agilent Technologies (3 mentions)
C17 0 fatty acid, by Merck Group (2 mentions)
Agilent 5975 gc msd system, by Agilent Technologies (2 mentions)
Hp 5ms, by Agilent Technologies (1 mentions)
Agilent 7683 series auto sampler, by Agilent Technologies (1 mentions)
Chemstation data analysis software, by Agilent Technologies (1 mentions)
Butylated hydroxytoluene, by Merck Group (1 mentions)
Biotek microplate reader, by Agilent Technologies (1 mentions)
Lc 20 system, by Shimadzu (1 mentions)
3200 qtrap mass spectrometer, by AB Sciex (1 mentions)
Luteolin 7 glucoside, by Merck Group (1 mentions)
L ascorbic acid, by Merck Group (1 mentions)
Gallic acid, by Merck Group (1 mentions)
Chlorogenic acid, by Merck Group (1 mentions)
Agilent 5975 mass selective detector, by Agilent Technologies (1 mentions)
Butyric, by Merck Group (1 mentions)
Isovaleric, by Merck Group (1 mentions)
Valeric, by Merck Group (1 mentions)
Hp 5975 mass selective detector, by Agilent Technologies (1 mentions)
5975 gc msd system, by Agilent Technologies (1 mentions)
17 protocols using agilent 6890n gc system
1
Quantitative Fatty Acid Analysis in Plants
2
GC-MS Analysis of Organic Compounds
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GCMS analysis was performed on a Agilent 6890 N GC System fitted with an HP5-MS (J & W Scientific) capillary column (30 m x 0.25 mm i.d. x 0.25 µm film thickness; maximum temperature, 250ºC) and coupled to a Agilent 5975 MS. Ultra-high purity helium was used as carrier gas at a constant flow of 1,0 mL.min -1 . The ionizing energy was 70 eV. The sample was prepared in ethyl acetate, and the injected sample volume was 1 µL with a split ratio of 30:1. The injector operated in 250ºC and detector 300ºC. The oven temperature program was 110ºC, and accelerated at a rate of 5°C.min -1 , 280°C (26 min). The compounds were identified by comparing the spectra obtained with mass spectrum library NIST 2011 and data reported in the literature (Adams, 2007) .
3
Optimized Urinary Organic Acids Analysis
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The extraction and derivatization protocol for urinary organic acids was optimized based on a previously reported method (17 (link)). Derivatized extracts (e.g., organic acids) were injected by an Agilent 7683 Series autosampler (Agilent Technologies, Palo Alto, CA, USA) followed by the analysis employing Agilent 6890N GC system coupled with electron impact (EI) ionization mode 5973N mass selective detector (Agilent Technologies, Palo Alto, CA, USA). Raw GC-MS data (“.D” file format) were first transformed into CDF format by the ChemStation Data Analysis software (Agilent Technologies, Palo Alto, CA, USA) prior to data pretreatment. Identification and quantification of metabolites was firstly processed and analyzed automatically by a web-based software called GC-AutoFit (http://gcms.wishartlab.com/ ) and results were further confirmed manually following the method as previously described (18 (link)). Further details on the organic acid extraction, derivatization, separation, and GC-MS data processing of urine samples were elaborated previously (19 (link)).
4
Cecal Fatty Acid Analysis Protocol
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Fatty acid analysis was conducted using an Agilent 6890N GC system (Agilent Technologies, PA, United States) and performed as previously described (Zhao et al., 2006 (link); Kang et al., 2017 (link)). Briefly, cecal contents from each mouse were weighed and homogenized in 1 mL deionized water for 3 min. The pH value of the suspension was adjusted to 2–3, and the suspension was subsequently transferred into a polypropylene tube and centrifuged for 20 min at 3,000 g, yielding a clear supernatant. 2-ethylbutyric acid (TEBA) was used as the internal standard and added into the supernatant at a final concentration of 1 mM.
5
Analytical Techniques for Compound Characterization
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The GC–FID analyses were carried out with capillary GC using an Agilent 6890N GC system (Agilent, Santa Clara, CA, USA), and the GC/MS analyses were performed on an Agilent 5975 GC–MSD system (Agilent, Santa Clara, CA, USA). An HP-Innowax FSC column (60 m × 0.25 mm, 0.25 μm film thickness, Agilent, Wilmington, DE, USA) was used for the analyses. The HPLC chromatographic separations were carried out using Shimadzu LC 20 System (Shimadzu, Tokyo, Japan). The mass spectra were recorded with AB Sciex 3200 Q TRAP mass spectrometer (AB Sciex, Toronto, Canada). GL Science Inertsil ODS 250 × 4.6 mm, 5 μm i.d. particle size, analytical column (GL Sciences, Tokyo, Japan) was used for the HPLC analyses. The turbidity of the standardized microbial sample solutions was measured using McFarland densitometer (Biosan McFarland Densitometer, Model Den-1B, Riga, Latvia). Antioxidant activity absorbances were recorded with a Biotek microplate reader (BioTek, Winooski, Vermont, USA). Chlorogenic acid, luteolin 7-glucoside, gallic acid, butylated hydroxytoluene (BHT), and L-ascorbic acid were purchased from Sigma-Aldrich (St. Louis, MO, USA).
6
Metabolite Analysis by GC-MS
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Five-gram subsamples were added to 10 ml ddH2O, placed in an ultrasonic cleaner (AS30600B; Autoscience, Tianjin, China) for 30 min, and centrifuged at 8,000 × g for 10 min. After filtering using an 0.2-μm-pore-size filter, the filtrate was used to analyze metabolite concentrations (27 (link)). Metabolites were detected using gas chromatography-mass spectrometry (Agilent 6890N GC system and Agilent 5975 mass selective detector; Agilent, Santa Clara, CA) with condition details based on a previous study (65 (link)).
7
Quantification of Fecal Short-Chain Fatty Acids
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Fecal samples collected at the end of the study were processed in duplicates for SCFA analyses according to previously published protocol.48 (link) To assess fecal SCFA concentration, samples were freeze-dried and then pulverized into powder. Approximately, 150 mg fecal powder was mixed with 250 μL hydrochloric acid, 45 μL internal standard (1 mM 2-ethyl butyric acid in 12% formic acid), followed by 2 extractions with 1 mL diethyl ether. An aliquot of the organic extract was transferred into glass vials for gas chromatographic analysis using Agilent 6890N GC system with a flame ionizable detector and an automatic liquid sampler (Agilent Technologies, Santa Clara, CA). Sample concentrations were determined using a 5-point calibration curve, with each standard containing the SCFAs, acetic, propionic, butyric, valeric, isovaleric, and isobutyric acids (Sigma-Aldrich).
8
Fatty acid profiling of GhBZR3-silenced plants
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Fatty acid profiling in GhBZR3‐silenced plants was performed as previously described (Qin et al., 2007 (link)). The 5‐DPA fibers were obtained from 10 μm BZR treatment after 6 h or 12 h and immersed in chloroform/methanol (2:1, v/v) for 1 min to remove surface waxes. Then, the samples were ground to powder in liquid nitrogen and extracted using 2.5% H2SO4 (v/v) in methanol. A known quantity of heptadecanoic acid (C17:0) was added into the extraction mixture to identify percentages for the quantification of fatty acids. Then, hexane was used for the extraction of fatty acid methyl esters. The Agilent 6890 N GC system (Agilent, https://www.agilent.com) with an HP 5975 mass selective detector (Agilent) coupled to a gas chromatography mass spectrometer (GC‐MS) system was used for fatty acid detection and measurement, as described previously (Liu et al., 2015 (link)).
9
GC Analysis of Separated Compounds
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The GC analysis was carried out using an Agilent 6890N GC system (Agilent Technologies, Santa Clara, CA, USA). FID detector temperature was 300 °C. To obtain the same elution order as with GC–MS, simultaneous auto-injection was done on a duplicate of the same column applying the same operational conditions. Relative percentage amounts of the separated compounds were calculated from FID chromatograms. The analysis results are given in Table 2 .
10
Quantitative GC-FID and GC-MS Analysis of Essential Oil Constituents
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The Agilent
6890N GC system was used. Simultaneous automatic injection was carried
out using the same conditions in two identical columns [HP-Innowax
FSC column (60 m × 0.25 mm, 0.25
μm film thickness, Agilent, Walt & Jennings Scientific,
Wilmington, Delaware)] in the Agilent 5975 GC/MSD system. Relative
percentages (%) of the compounds were calculated using the FID chromatograms.
For idenfication and characterization in-house “Baser Library
of Essential Oil Constituents” and various GC/MS Libraries
such as MassFinder 3 Library, where authentic samples or the relative
retention index (RRI) of n-alkanes were also considered,29 (link) see, for details,Figures S1 and S2 .
6890N GC system was used. Simultaneous automatic injection was carried
out using the same conditions in two identical columns [HP-Innowax
FSC column (60 m × 0.25 mm, 0.25
μm film thickness, Agilent, Walt & Jennings Scientific,
Wilmington, Delaware)] in the Agilent 5975 GC/MSD system. Relative
percentages (%) of the compounds were calculated using the FID chromatograms.
For idenfication and characterization in-house “Baser Library
of Essential Oil Constituents” and various GC/MS Libraries
such as MassFinder 3 Library, where authentic samples or the relative
retention index (RRI) of n-alkanes were also considered,29 (link) see, for details,
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