The nitrogenase activity was quantified by measuring the reduction of acetylene into ethylene using gas chromatography-flame ionization detection (GC-FID). Two independent LORE1 lines were inoculated with M. loti MAFF303099-DsRed, as described previously, and assayed at 21 dpi. Five biological replicates were analyzed. A single replicate comprised two plants with nodulated roots in a 25 mL glass tube with 500 μL of FAB medium and sealed with a rubber stopper. Subsequently, 1 mL of air was extracted and replaced with 1 mL of acetylene. Per each replicate the gas mixture was sampled at 0, 20, 40, 60, and 80 min while keeping the plants at 28 °C in a water bath. At each time point, 1 mL of the mixture was injected into a GC 2010 Pro (Shimadzu). The activity was calculated as nanomoles of ethylene per hour using a linear regression.
Gc 2010 pro
Manufactured by Shimadzu
Sourced in Japan
The Shimadzu GC-2010 Pro is a gas chromatograph designed for analytical separation and quantification of chemical compounds. It features dual injection ports, dual detectors, and advanced temperature control capabilities to enable precise and reliable analysis. The GC-2010 Pro is a core laboratory instrument suitable for a wide range of applications in the chemical, environmental, and life science industries.
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Lab products found in correlation
Sh rt 2560, by Shimadzu (3 mentions)
Cp sil 5 cb column, by Shimadzu (1 mentions)
5973 msd mass spectrometer, by Agilent Technologies (1 mentions)
L 8900 amino acid analyzer, by Hitachi (1 mentions)
Acetophenone, by Merck Group (1 mentions)
ε caprolactone, by Merck Group (1 mentions)
Gc solution software, by Shimadzu (1 mentions)
Gc 2014, by Shimadzu (1 mentions)
Supelco 37 component fame mix, by Merck Group (1 mentions)
Tg 5ms capillary column, by Thermo Fisher Scientific (1 mentions)
Tq8050, by Shimadzu (1 mentions)
Cary 50 uv vis, by Agilent Technologies (1 mentions)
Rtx 5ms capillary column, by Restek (1 mentions)
Aoc 201, by Shimadzu (1 mentions)
Cary 60 spectrophotometer, by Agilent Technologies (1 mentions)
Tcs sp8 sted 3x confocal laser scanning microscope, by Leica (1 mentions)
Ultima 4 x ray diffractometer, by Rigaku (1 mentions)
Jem 2010, by JEOL (1 mentions)
Sdta851e, by Mettler Toledo (1 mentions)
16 protocols using gc 2010 pro
1
Quantifying Nitrogenase Activity in LORE1 Mutants
2
Dietary Fat Sources and Encapsulation in Piglet Nutrition
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Three experimental diets based on corn and soybean meal are shown in Table S1 : soybean oil diet (SBO, control group, 6.0% soybean oil), palm oil diet (PO, 6.0% palm oil), and encapsulated palm oil diet (EPO, 7.5% encapsulated palm oil). All the diets meet the nutrient requirements of 7 to 11 kg piglets [11 ]. Soybean oil was obtained from Cargill Grain and Oil Co., Ltd. (Dongguan, China). Palm oil was purchased from Yihai Kerry Grain & Oil Ltd. (Guangzhou, China). The same source of palm oil was encapsulated using 8.5% dried casein and 11.5% whey powder. The three ingredients were stirred at 60 °C for 30 min to uniformly mixed them in liquid form, pasteurized at 65 °C for 30 min, and homogenized at a pressure of 250 kg/cm2 before spray drying (EYELA Spray Drier SD-1000, Tokyo Rikakikai Co., Ltd., Tokyo, Japan). The inlet temperature of the drier was 180 °C and the outlet temperature was 80 °C. The size of the dried fat powder particles ranged from 30 to 50 μm. The fatty acid composition of the fat sources used in the experiment was analyzed using gas chromatography (GC-2010 Pro, SHIMADZU Corporation, Kyoto, Japan) and is shown in Table S2 .
3
Lipid and Fatty Acid Profiling of Muscle Tissue
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The muscle total lipid content was extracted using the chloroform–methanol method. The fatty acid composition was assayed using a gas chromatograph method (Shimadzu GC-2010 Pro), as previously described [4 (link)]. A flame ionization detector and fused silica capillary column (SH-RT-2560, 100 m × 0.25 mm × 0.20 μm, Shimadzu, Japan; dicyano-propyl-polysiloxane as stationary phase) were used. The results are expressed as the percentages of each fatty acid with respect to the total fatty acids.
4
Proximate and Fatty Acid Analysis of Fish Tissues
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The proximate composition analysis of fish tissues was conducted according to the methods of the Association of Official Analytical Chemists [50 ]. Briefly, the protein content: Kjeldahl method by measuring total nitrogen (N× 6.25); total lipid: Soxhlet method (petroleum ether extraction); moisture content: oven-drying at 105 °C to constant weight; ash: incineration at 550 °C.
The fatty acid compositions of all tissues (muscle, liver and intestine) were analyzed through gas chromatography (GC-2010 pro, Shimadzu, Japan). Briefly, fatty acids in lyophilized tissue samples were esterified with KOH-methanol and HCL-methanol in 72 °C water bath. Fatty acid methyl esters were extracted with hexane and then analyzed through gas chromatography using a flame ionization detector and a fused silica capillary column (SH-RT-2560, 100 m × 0.25 mm × 0.20 μm). The column temperature increase was programmed: 150 °C to 200 °C at 15 °C min-1; then from 200 °C to 250 °C at 2 °C min-1. The injector and detector temperatures were 250 °C. Fatty acid results were expressed as percentage of each fatty acid with respect to total fatty acids (% TFA).
The fatty acid compositions of all tissues (muscle, liver and intestine) were analyzed through gas chromatography (GC-2010 pro, Shimadzu, Japan). Briefly, fatty acids in lyophilized tissue samples were esterified with KOH-methanol and HCL-methanol in 72 °C water bath. Fatty acid methyl esters were extracted with hexane and then analyzed through gas chromatography using a flame ionization detector and a fused silica capillary column (SH-RT-2560, 100 m × 0.25 mm × 0.20 μm). The column temperature increase was programmed: 150 °C to 200 °C at 15 °C min-1; then from 200 °C to 250 °C at 2 °C min-1. The injector and detector temperatures were 250 °C. Fatty acid results were expressed as percentage of each fatty acid with respect to total fatty acids (% TFA).
5
Gas Chromatography-Mass Spectrometry Analysis
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The analyses were perform using a Thermo Finnigan Trace gas chromatograph interfaced to a Thermo Finnigan Automass mass spectrometer (Thermo Fisher Scientific, Waltham, MA, USA, 70 eV) operated in fullscan or SIM mode with a fused silica capillary column (HP-5% phenylmethylpolysiloxane 30 m × 0.25 mm, 0.25 µm film thickness) and helium as carrier gas. Line transfer and source were held, respectively, at 280 °C and 220 °C. The GC oven temperature program was from 60 °C, at 5 °C·min−1 up to 300 °C, and the temperature was finally held for 10 min at 300 °C. The temperatures of the injector and the detector were 250 and 300 °C, respectively.
Py-GC/MS experiments used a EGA-PY 3030D Pyrolyzer (Frontier Lab, Japan) connected to a gas chromatography instrument (GC2010 Pro, Shimadzu, Kyoto, Japan) with capillary column (SLB-5% phenylmethylsiloxan 30 m × 0.25 mm, 0.25 µm film thickness) and coupled with mass spectrometry (Ultra QP 2010). The GC and MS conditions were the same as for GC-MS analysis.
Py-GC/MS experiments used a EGA-PY 3030D Pyrolyzer (Frontier Lab, Japan) connected to a gas chromatography instrument (GC2010 Pro, Shimadzu, Kyoto, Japan) with capillary column (SLB-5% phenylmethylsiloxan 30 m × 0.25 mm, 0.25 µm film thickness) and coupled with mass spectrometry (Ultra QP 2010). The GC and MS conditions were the same as for GC-MS analysis.
6
Fatty Acid Profiling of Biological Samples
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The fatty-acid compositions of oils, diets, muscles, and livers were analyzed with gas chromatography, as previously described [39 (link)]. Lipids in the samples were firstly extracted with the chloroform methanol method. Fatty acids in the extracted oil were then saponified and methylated with KOH-methanol and HCL-methanol. Gas chromatograph (GC-2010 pro, Shimadzu, Kyoto, Japan) equipped with a quartz capillary column (SH-RT-2560, 100 m × 0.25 mm × 0.20 um) and a flame ionization detector was used in the analysis. The results are expressed as percentage of each fatty acid relative to the total fatty acids (%TFA).
7
Pd-Catalyzed Heterocyclic Synthesis
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All chemicals were obtained from commercial suppliers or prepared according to known synthetic procedures. An additional purification via column chromatography was performed for the heterocyclic reactants, and the other chemicals were used as received. N,N-Dimethylacetamide (>99.9%, HPLC grade) was used as the solvent without further drying steps. The Pd2dba3 precursor was stored under inert, moisture-free atmosphere. GC analysis was performed on a Shimadzu GC-2010 Pro equipped with a CP-Sil 5 CB column and flame ionization detector. The NMR spectra were recorded on a Bruker Avance III HD 400 MHz spectrometer. Column chromatography was performed on 70–230 mesh silica 60 (Merck) as the stationary phase. For TLC analysis, silica gel on TLC Al foils with fluorescent indicator (254 nm) was used. The standard MS spectra were obtained via GC-MS (Agilent 6890 equipped with HP-5MS column and a 5973 MSD mass spectrometer using electron impact ionization). The MS data of the products featuring a high boiling point or limited stability were acquired using a Water's Radian ASAP mass spectrometer.
8
Fatty Acid and Amino Acid Analysis
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The fatty acid compositions of oil, diet, muscle, and liver were analyzed with gas chromatography (GC2010 pro, Shimadzu, Kyoto, Japan) equipped with a flame ionization detector and a quartz capillary column (SH-RT−2560, 100 m × 0.25 mm × 0.20 μm). Lipids were first extracted from the samples using the chloroform methanol method. Fatty acids in the lipid samples were then saponified and methylated with boron trifluoride and KOH-methanol. The fatty acid contents are expressed as % total fatty acids (TFA). More details can be found in our previous publications [51 (link)].
The muscle samples were deproteinized using trichloroacetic acid (6%) and centrifuged at 10,000g at 4°C for 10 min to obtain the supernatant. Amino acid contents were determined using the L-8900 amino acid analyzer (Hitachi, Japan).
The muscle samples were deproteinized using trichloroacetic acid (6%) and centrifuged at 10,000g at 4°C for 10 min to obtain the supernatant. Amino acid contents were determined using the L-8900 amino acid analyzer (Hitachi, Japan).
9
Extraction and GC Analysis of Organic Compounds
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Extraction was done by ethyl acetate with 2 mM acetophenone (Sigma-Aldrich, US) as an internal standard. 300 µl sample with 0.06% (v/v) HCl was extracted twice with 150 µl ethyl acetate and shaken for 15 min with multi-tube vortex (Baxter, US). After centrifugation the ethyl acetate (Lab-Scan Analytical Sciences, Poland) phase was collected and dried with anhydrous MgSO4. The samples were centrifuged and measured with GC-2010 Pro gas chromatograph (Shimadzu, Japan) equipped with a HP-5MS 30 m × 0.25 mm (5%-Phenyl)-methylpolysiloxane column (19091S-133, Agilent) using nitrogen as carrier gas with splitless injection mode. Compounds were separated at 35 °C (hold 3 min), 200 °C (hold 3 min, 10 °C min−1) and 300 °C (hold 3 min, 25 °C min−1). Linear velocity was 11 cm sec−1. Calibration was done by using known amounts of cyclohexanone, ε-caprolactone and acetophenone (Sigma-Aldrich, US).
10
Quantitative HPLC and GC Analysis
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The pH of each sample was measured on-site using a HANNA® pocket pH meter (HANNA, United States). Acetate, lactate, glucose and glycerol were analyzed on a Finnigan Surveyor Plus HPLC fitted with a Biorad Aminex HPX 87H ion exchange column (300 mm × 10 mm), using 5 mM sulphuric acid at a flow rate of 0.6 ml min–1 as eluent. Acids were detected with a PDA detector at 202 nm and glycerol and glucose with a refractive index detector connected in series. Analytes were quantified by external standards. Prior to HPLC analysis, 5 ml of each of the samples were centrifuged (10,000 × g for 5 min) and the supernatant collected for analyses. Ethanol concentrations were determined by gas chromatography (Shimadzu GC-2010 Pro), carrier gas: hydrogen 60 cm.s–1,CPWax 52CB column [30 m (L) × 0.32 mm (ID) × 0.25 μm (Film thickness)]. The flame ionization detector temperature was 260°C and the injection port was 150°C. Injection volume was 1 μL with a split of 1:10. The initial column temperature was 40°C held for 3 min and then ramped to 256°C at 25°C min–1 and held for 1°min. Shimadzu GC Solution software was used for instrument control, data collection and analysis.
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