The phenolic compounds were extracted from semolina, olive paste flour (OPF), spaghetti CTRL and spaghetti enriched with 10% OPF in two separated fractions: soluble free and bound, following the method reported by Mattilla [20 (link)] without acid hydrolysis. The difference between total and free polyphenols represented the bound phenolic compounds. HPLC-DAD analysis was performed using the Agilent 1260 infinity system, equipped with a 1260 binary pump, 1260 HiP Degasser, 1260 TCC Thermostat, 1260 Diode Array Detector and Agilent Open Lab Chem Station Rev C.01.05 (35) software. The UV–visible absorption chromatogram was detected at 280 nm, 325 nm and 360 nm. The separation was performed by gradient elution on a 4.6 × 250 mm reversed phase Luna C-18 (5 μm) column (Phenomenex Torrance, California, USA). The elution was performed using methanol (eluent A) and water/acetic acid 95:5 (eluent B). The gradient profile was: 85–60% B (0–25 min), 60% B (25–30 min), 60–37% B (30–45 min), 37% B (45–47 min), 37–0% B (47–52 min). The flow rate was 1 mL/min and the injection volume was 25 μL. Phenolics were identified and quantified by the retention time, spectra and response factors of the pure standards.
1260 binary pump
Manufactured by Agilent Technologies
Sourced in United States
The 1260 binary pump is a high-performance liquid chromatography (HPLC) pump that delivers precise and stable flow rates of mobile phases for analytical applications. It features two independent solvent delivery systems for binary gradients, allowing for the accurate blending of two solvents. The pump operates at a maximum pressure of 600 bar and can deliver flow rates ranging from 0.001 to 10 mL/min, making it suitable for a variety of HPLC methods.
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Lab products found in correlation
1260 infinity system, by Agilent Technologies (1 mentions)
1260 hip degasser, by Agilent Technologies (1 mentions)
1260 diode array detector, by Agilent Technologies (1 mentions)
Luna c 18 5 μm column, by Phenomenex (1 mentions)
1260 dad hs, by Agilent Technologies (1 mentions)
6120 single quadrupole lc ms system, by Agilent Technologies (1 mentions)
1260 infinity 2 lc system, by Agilent Technologies (1 mentions)
Poroshell 120 ec c18, by Agilent Technologies (1 mentions)
1200 isocratic pump, by Agilent Technologies (1 mentions)
Hss t3 c18 column, by Waters Corporation (1 mentions)
6540 q tof, by Agilent Technologies (1 mentions)
Optilab t rex refractometer, by Wyatt Technology (1 mentions)
Superdex 200 increase 10 300 gl column, by GE Healthcare (1 mentions)
6230 time of flight mass spectrometer, by Agilent Technologies (1 mentions)
6 protocols using 1260 binary pump
1
Phenolic Compounds Extraction and Quantification in Semolina and Spaghetti
2
Liquid Chromatography-Mass Spectrometry of cGAMP
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Liquid chromatography-mass spectrometry (LC-MS) with an electrospray ionization (ESI) interface was used to verify the production of cGAMP. The Agilent Technologies 1260 Infinity II LC System was coupled to Agilent Technologies 6120 Single Quadrupole LC/MS System (Agilent Technologies Inc., Santa Clara, CA, USA) and further equipped with a Diode Array Detector (1260 DAD HS, Agilent Technologies Inc., Santa Clara, CA, USA), Multicolumn Thermostat (1260 MCT, Agilent Technologies Inc., Santa Clara, CA, USA), Degasser (1260 Degasser, Agilent Technologies Inc., Santa Clara, CA, USA), Multisampler (1260 Multisampler), Binary Pump (1260 Binary Pump, Agilent Technologies Inc., Santa Clara, CA, USA) and an Agilent Poroshell 120 EC-C18 (4.6 × 100 mm, 2.7 μm particle size) column. Sample volumes of 5 μL were injected and separated by a gradient of mobile phase A (0.1 % formic acid) and mobile phase B (acetonitrile) (0 min 5% A, 95% B; 7 min 95% A, 5% B; 9 min 5% A, 95% B; 14 min 5% A, 95% B). The flow rate was set to 1 mL min−1 at a column temperature of 40 °C and a detection wavelength of 254 nm. Measured masses ranged from 100 to 1000 m/z. The gas chamber was heated to 350 °C with a gas flow of 12 L min−1. The capillary voltage was set to 3 kV and the nebulizer pressure to 35 psi.
3
Liver Metabolite Extraction and Analysis
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Pieces of whole liver were minced, weighed into individual wells, then homogenization buffer [63 (link)] was added to those corresponding wells. Control tissue homogenate was made by adding homogenization buffer at a 9 to 1 ratio to weighed amount of mouse untreated liver. Aliquots and appropriate amounts of calibration standards were added into wells. Internal standard and approximately 0.25cm3 granite beads were added and then extracted via a liquid-liquid extraction with ammonium hydroxide and phenol: chloroform: isoamyl alcohol (25:24:1). The aqueous layer was then further processed via solid phase extraction utilizing a Phenyl plate. Eluates had a final pass through a protein precipitation plate before dry down under nitrogen at 50°C. Dried samples were reconstituted in water containing 100 μM EDTA. These samples were injected into an Agilent LCMS instrument consisting of a 1260 binary pump, a 1200 isocratic pump, a column oven, an auto sampler, and a 6130 single quadrupole mass spectrometer for analysis (Agilent, Wilmington, DE, USA).
4
Organic Acids Quantification by LC-QTOF-MS
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LC/MS analysis for organic acids was performed using an Agilent Technologies 6540 Q-TOF equipped with a Jet Spray ESI source operated in negative ionization mode at a 3 Hz scan rate in high resolution mode across mass range 50 to 1,700 m/z. The source parameters were 150°C drying 76 gas, 12 L/min drying gas, nebulizer pressure of 45 psi, sheath gas temperature 325°C, sheath gas flow 12 L/min, capillary at 2,000 V, nozzle at 0 V, fragmentor 90 V, skimmer 35 V, octopole RF 750 V. Five microliter of each sample was injected using an HTC autosampler into a 2 μL loop supplied by an Agilent 1260 binary pump. The column was a 2.1 mm × 100 mm Waters HSS-T3 C18 column with 1.8 μm packing. Buffer A was 20 mmol/L ammonium formate pH 2.9; buffer B was acetonitrile. Gradient separation began with 0.5% buffer B at 0.4 mL/min, held until 2.5 minutes, then buffer B was increased to 99.5% at 4 minutes. The stream was switched from the LC/MS to waste, and for regeneration, flow was increased to 0.5 mL/min at 5 minutes and 1 mL/min at 5.5 minutes, held until 8.5 minutes, and then decreased to 0.5 mL/min at 9 minutes. The column was re-equilibrated by a shift to 0.5% buffer B at 10.5 minutes, flow as decreased to 0.4 mL/min at 13 minutes, and held until 14 minutes. Other compounds were analyzed by ion-paired reverse phase analysis in negative mode on a 6550 Q-TOF.
5
Multi-Angle Light Scattering Analysis
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MALS was performed as per the previous report [55 ]. Superdex 200 Increase 10/300 GL column (GE Healthcare) was pre-equilibrated overnight at 0.1 ml/minute flow rate with buffer E (20 mM HEPES pH 8.0, 200 mM NaCl, 1 mM β-ME). For each analysis, 200 μl of a protein sample at 2.0 mg/ml concentration was loaded onto the column at 0.6 ml/minute flow rate using a 1260 binary pump (Agilent Technologies). The scattered light was measured with a miniDAWN TREOS II light scatterer, (Wyatt Technologies), and the refractive index was measured with an Optilab T-rEX refractometer, (Wyatt Technologies). Data analysis was performed with ASTRA 7.3.2.21 (Wyatt Technologies) [56 (link)].
6
UHPLC/TOF-MS Analysis of AC-X-A Samples
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Ultra high performance liquid chromatography coupled with time–of–flight mass spectrometry (UHPLC/TOF–MS) experiments were performed using an Agilent 1260 binary pump in low dwell volume mode, an Agilent column oven heated to 45°C, and an Agilent 6230 Time of Flight Mass Spectrometer with an electrospray ionization (ESI) source. Experimentally, 1 µL of the AC–X–A sample dissolved in 50% aq. MeOH was injected onto a 1.8 mm particle size, 50×2.3 mm I.D., Zorbax RRHT column. Each sample was subjected to a gradient from 10: 1 to 1: 10 H2O: CH3CN over 4 min with a 1.5 min hold at 1: 10 H2O: CH3CN before a 3 min re–equilibration. The flow rate was maintained at 0.8 mL/min. Formic acid, 200 µL/L, was added to both the H2O and the CH3CN. The TOF–MS was run in positive high–resolution 4 GHz mode with a detector range from 100 m/z to 1700 m/z. The ESI source was operated with a desolvation temperature of 350°C and a drying gas flow rate of 11 L/min. The fragmentor voltage was held at 135 V while the capillary voltage was ramped from 2500–3000 V.
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