CO2 cultivated algal cells were harvested and freeze-dried after aerated cultivation for 7 days. Lipids were extracted from 40 mg dried algal powder using a chloroform–methanol protocol according to Wu et al. [9 (link)]. The extracts were combined after three extractions and then evaporated under high purity nitrogen. The gravimetric means was used for quantification of total lipids until extracts maintained at a constant weight. Lipids were methylated and then fatty acid methyl esters were analyzed by gas chromatography according to the protocol described by Liu et al. [51 (link)]. Quantification of fatty acids was carried out with an Agilent 7890A instrument equipped with a DB-FFAP capillary column (30 m × 0.25 mm). Fatty acids were identified by comparison of their retention times with those of standards (Sigma). Nonadecanoic acid (19:0) was used as an internal standard.
Db ffap capillary column
Manufactured by Agilent Technologies
Sourced in United States, Japan, Germany, China
The DB-FFAP capillary column is a stationary phase designed for the separation and analysis of polar compounds in gas chromatography. It features a liquid phase of polyethylene glycol and is suitable for a wide range of applications involving the separation of polar analytes.
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Lab products found in correlation
5975c inert msd quadrupole mass spectrometer, by Agilent Technologies (3 mentions)
Msd chemstation, by Agilent Technologies (3 mentions)
Chemstation software, by Agilent Technologies (2 mentions)
2 ethylbutyric acid, by Merck Group (2 mentions)
7890a gas chromatography system, by Agilent Technologies (2 mentions)
7697a headspace automatic injector, by Agilent Technologies (2 mentions)
Agilent 7820a gc system, by Agilent Technologies (2 mentions)
Gc 2010 plus, by Shimadzu (2 mentions)
Agilent 7890a, by Agilent Technologies (2 mentions)
Gc 2014c, by Shimadzu (2 mentions)
Cg 2010, by Shimadzu (2 mentions)
Hp 7890a, by Agilent Technologies (2 mentions)
Isolongifolene, by Merck Group (2 mentions)
Gc 7890a, by Agilent Technologies (2 mentions)
Agilent 7890b gc system, by Agilent Technologies (1 mentions)
Aoc 20i autoinjector, by Shimadzu (1 mentions)
Tsq 8000 evo mass spectrometer, by Thermo Fisher Scientific (1 mentions)
Triplus rsh autosampler, by Thermo Fisher Scientific (1 mentions)
3800 gas chromatograph, by Agilent Technologies (1 mentions)
Primacsslc, by Skalar (1 mentions)
61 protocols using db ffap capillary column
1
Algal Lipid Extraction and Fatty Acid Analysis
2
Soybean Seed Fatty Acid Profiling
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Fatty acid profiles (16:0, 18:0, 18:1, 18:2, and 18:3) in soybean seeds were expressed as a percentage of the total fatty acids of seed based on gas chromatography data obtained using an Agilent series 7890A capillary gas chromatograph equipped with a flame ionization detector at 250ºC (Agilent Technologies Inc., Wilmington, DE, USA) for fatty acid methyl esters in extracted oil. The oil was extracted by placing crushed seeds in a 1.5 mL solution of chloroform, hexane, and methanol (8:5:2, v/v/v) for ~12 hours. A 100 µL derivatization solvent was mixed with 75 µl of a methylation reagent (0.25 M methanolic sodium methoxide:petroleum ether:ethyl ether [1:5:2, v/v/v]). The samples were diluted with hexane to approximately 1 mL. Five fatty acids were separated on a DB-FFAP capillary column (30 m × 0.25 mm, 0.25 µm, Agilent Technologies Inc., Wilmington, DE, USA). Standard fatty acid mixtures (Fame #16, RESTEK) were used as calibration reference standards.
3
Quantification of Fecal Short-Chain Fatty Acids
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The levels of SCFAs in feces were measured by gas chromatography (GC) as previously reported.45 (link) Briefly, 100.0 mg feces stored at −80°C were diluted with 1 mL vortex-mixed in Milli-Q water for 2 min, incubated at room temperature for 10 min, and centrifuged at 14,000 rpm for 20 min. The impurities in supernatants were removed by centrifugating twice at 14,000 rpm for 20 min, and 2-ethylbutyric acid (Sigma, MO, USA) was added as an internal standard. Then, the supernatants were transferred to into a fresh 2 ml glass vial for GC analysis. GC analysis was performed using an Agilent 7890 N gas chromatograph system (Agilent Technologies Inc., CA, USA) coupled with DB-FFAP capillary column (30 m × 0.25 mm×0.25 µm; Agilent). A flame ionization detector was used for identification and quantification of SCFAs in each sample. The data were acquired using the ChemStation software (Agilent). The concentration (in mM range) of SCFAs (acetic acid, propionic acid, isobutyric acid, butyrate acid, isovaleric acid, valeric acid, and hexanoic acid) were calculated using the linear regression equations (R2 ≥ 0.99) from the corresponding standard curves.
4
Fecal SCFA Quantification Protocol
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Fecal samples were collected individually following the Standard Operating Procedure (SOP) of self-collection (http://www.microbiome-standards.org ). Fecal SCFAs concentrations were extracted and determined according to a method from a previous study (Zhang et al., 2020 (link)). 100 mg dried fecal powder was resuspended in 1 mL of distilled water and vortexed for 2 min. Then, 600 μL of the supernatant of the fecal slurry was acidified with 20% (v/v) H2SO4. After vortexing for 1 min, 500 μL of n-butanol was added to the mixture and vortexed for 2 min. The supernatant was filtered through a 0.22 μm filter membrane for sample injection. Agilent 7890B GC system (Agilent Technologies, Santa Clara, CA) equipped with a DB-FFAP capillary column (30 m × 0.32 mm i.d., 0.25 μm film thickness, Agilent, Santa Clara, USA) was used for component separation and detection of the fecal SCFAs concentrations.
5
Fatty Acid Composition Analysis
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Samples of 2.0 mg of fat were accurately weighed and dissolved in 1.5 mL of methanol; then, 60 μL of methyl acetate was added, and it was shaken and allowed to stand for 10 min. Then we added 100 μL of 27 mg/mL sodium methoxide solution and allowed it to stand for 20 min. This was followed by 60 μL of saturated oxalic acid, then shaken well and allowed to stand for 2 minutes in a −20 °C refrigerator [34 (link)]. Finally, the fatty acid composition was determined by gas chromatography (DB-FFAP capillary column, Agilent Company, Santa Clara, CA, USA).
6
Plasma Fatty Acid Profiling Protocol
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Plasma obtained from the non-fasting blood samples was stored at –70°C, thawed once to obtain a 100 µl aliquot that was refrozen and shipped by airfreight to Rockville, MD, USA, and then thawed for final analyses(42 ). Plasma FA were extracted using transmethylation of lipids with acetyl chloride and methanol(43 ,44 ). Chromatographic separation of the FA methyl esters was achieved via a fast gas chromatography 6890 Plus LAN system (Agilent Technologies) coupled with a fused-silica, narrow bored DB-FFAP capillary column (Agilent 127–32H2, 15m × 0·1 mm I.D. × 0·1 mm film thickness). Assays were carried out during 2009–2010 with the measurement of twenty-two FA, eleven of which were PUFA.
7
GC-MS Analysis of Short-Chain Fatty Acids
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An Agilent Technologies 7890A gas chromatography system coupled to an Agilent Technologies 5975c inert MSD quadrupole mass spectrometer (Agilent Technologies, Germany) and equipped with an Agilent Technologies 7697A Headspace automatic injector (Agilent Technologies, Germany) and an DB-FFAP capillary column (30 m × 0.25 mm i.d., 0.25 μm film thickness, Agilent Technologies, Germany) was used to perform an analysis of SCFAs from fecal samples. The headspace was maintained at 80°C with an incubation time of 30 min. The samples (1 mL) were injected in a splitless mode into the column at a temperature of 80°C. Helium was used as the carrier gas at a constant flow rate of 1 mL/min through the column. The initial oven temperature was 50°C, which was maintained for 1 min and then raised to 200°C at a rate of 10°C/min. The temperatures of the ion source and injector were 250°C. The mass detector system was operated in an electron impact (EI) mode with an ionization energy of 70 eV. The data of ions monitored were collected from m/z 33 to 200. A qualitative analysis was performed by the National Institute of Standards and Technology (NIST 11) MS library.
8
IBA and isobutanol quantification by GC-FID
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Concentrations of IBA and isobutanol were analysed by GC–FID. The GC system is a GC-2010 with an AOC-20 S auto sampler and AOC-20i auto injector (Shimadzu). The column used was a DB-FFAP capillary column (60-m length, 0.32-mm diameter and 1-μm film thickness; Agilent Technologies; Santa Clara, CA, USA). The GC oven temperature was held at 225 °C, and the FID was held at 330 °C. The injection volume was 0.5 μl, injected at a 15:1 split ratio. Helium was used as the carrier gas. 1-Pentanol was used as an internal standard. Retention times from samples were compared with external standards. All samples contained 100 mg l−1 of 1-pentanol which served as an internal standard.
9
Quantifying Short-Chain Fatty Acids in Fecal Samples
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The short-chain fatty acid (SCFA) analysis was used by a 7890A gas chromatography system (Agilent Technologies, Germany) coupled to a 5975c inert MSD quadrupole mass spectrometer, a 7697A headspace automatic injector, and a DB-FFAP capillary column (30 mm × 0.25 mm i.d., 0.25 μm film thickness). A portion of the 0.1 g homogenized fecal sample was mixed in 1 mL of 6% H3PO4 solution by ultrasound for 3 min and was then placed in an automatic headspace sampler at 80°C for 30 min. Subsequently, the samples (1 ml) were injected into the column at 80°C. Helium gas was used as a carrier for samples to pass through the column at a constant flow rate of 1 mL/min. The oven was preheated at 50°C for 1 min; then heated up to 200°C at the rate of 10°C/min. The temperature of the ion source and injector was set to 250°C. The mass detector system was operated in an electron impact (EI) mode with ionization energy of 70 eV. The data of ions monitored were collected from m/z 33 to 200. The short-chain fatty acids were quantified by the MS library of the National Institute of Standards and Technology (NIST 11).
10
SCFA Quantification in Intestinal Contents
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The concentrations of SCFA (acetate, propionate, and butyrate) in intestinal contents were detected using a Fuli 9720 gas chromatograph (Wu et al., 2017; (link)Liu et al., 2021) . A 500-μL sample from the intestinal contents (50 mg) treated with the HALO-F100 stool treatment instrument (Suzhou HALO Bio-Technology Co. Ltd.) was mixed with 100 μL of crotonic acid monophosphate solution. The sample was then centrifuged at (3,500 × g) for 3 min at 4°C, and the supernatant was taken through a 0.22-μm drainage membrane. The extracted sample was stored in a 2-mL gas-phase bottle and finally injected into a GC system equipped with an Agilent DB-FFAP capillary column (30 m × 0.25 mm i.d. × 0.25 μm) and detected by a flame ionization detector at 250°C. The parameters were set as follows: injection volume of 1 μL, split ratio of 5:1, N 2 as the carrier gas, and flow rate of 2.5 mL/min.
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