The 2′-FL and xylose concentrations were analyzed using high-performance liquid chromatography (HPLC) system (Agilent Technologies 1260 Series) equipped with a Rezex ROA Organic Acid H + (8%) Column (Phenomenex, Torrance, CA, USA). The column and the refractive index detector temperature were set at 50 °C, and the mobile phase was 10 mM H2SO4 at a flow rate of 0.5 mL/min at 40 °C. The sucrose and lactose concentrations were analyzed using the HPLC system (Agilent Technologies 1260 Series) equipped with an XBridge BEH Amide Column (Waters, Milford, MA, USA). The column and the refractive index detector temperature were set at 35 °C, and the mobile phase was 75% acetonitrile at a flow rate of 1.0 mL/min at 35 °C. The liquid chromatography/mass spectrometry (LC/MS) matrix-assisted laser desorption/ionization time-of-flight system (Waters) was used to identify 2′-FL according to a previous study [22 (link)], and the mass range was 100–700 m/z. The relative transcription levels of the regulated genes were determined by quantitative real-time polymerase chain reaction (qRT-PCR) as described previously [23 (link)], and the rpsJ gene was used as the internal standard [24 (link)]. All experiments were independently carried out at least thrice.
Rezex roa organic acid h 8 column
Manufactured by Phenomenex
Sourced in United States, Germany, United Kingdom, Japan, Switzerland
The Rezex ROA-Organic Acid H+ (8%) column is a high-performance liquid chromatography (HPLC) column designed for the separation and analysis of organic acids. The column features a stationary phase composed of a styrene-divinylbenzene copolymer with a sulfonic acid functionality, which allows for the effective separation of various organic acids. The column operates at a nominal particle size of 8 micrometers and a pore size of 300 angstroms, providing efficient chromatographic performance.
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Lab products found in correlation
Agilent 1100 series, by Agilent Technologies (8 mentions)
Rezex roa organic acid h 8 guard column, by Phenomenex (6 mentions)
1200 series, by Agilent Technologies (5 mentions)
Agilent 1200 series, by Agilent Technologies (4 mentions)
1100 series ri detector, by Agilent Technologies (3 mentions)
Whatman, by Cytiva (3 mentions)
E2695 separation module, by Phenomenex (3 mentions)
Nylon filter, by Merck Group (2 mentions)
Biomate 5, by Thermo Fisher Scientific (2 mentions)
1260 series, by Agilent Technologies (2 mentions)
Vecta spin tubes, by Cytiva (2 mentions)
Membrane of 0.2 μm, by Cytiva (2 mentions)
Rezex rpm monosaccharide pb 2 8 column, by Phenomenex (2 mentions)
Guard column, by Phenomenex (2 mentions)
Rid 10a refractive index detector, by Shimadzu (2 mentions)
Nexera hplc system, by Shimadzu (2 mentions)
Diluphotometer, by Implen (2 mentions)
0.45 μm polypropylene membrane, by Avantor (2 mentions)
Software galaxie chromatography, by Agilent Technologies (2 mentions)
Spectroquant test kit, by Merck Group (2 mentions)
77 protocols using rezex roa organic acid h 8 column
1
Quantification of Carbohydrates and Gene Expression in Biological Samples
2
HPLC Quantification of Wort and Beer Compounds
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The sugars, organic acids, glycerol, and ethanol of wort and beer samples were quantitated by an HPLC-refractive index detection (RID) system (1260 Infinity II; Agilent, Santa Clara, CA, USA) equipped with a Rezex ROA-organic acid H+ (8%) column (300 by 7.8 mm; Phenomenex, Torrance, CA, USA).
Sample preparation and analysis were carried out as previously described (61 (link)) with a few modifications. Fermentation samples were centrifuged at 15,000 rpm for 5 min, followed by filtration of supernatant through a 0.45-μm nylon membrane filter. The filtrate obtained was loaded for sugar, organic acid, glycerol, and ethanol analysis using a mobile phase of 0.005 N H2SO4 at 60°C with a flow rate of 0.5 ml/min for 35 min. Serial standard solutions of compounds of interest were prepared and analyzed to establish standard calibration curves.
Sample preparation and analysis were carried out as previously described (61 (link)) with a few modifications. Fermentation samples were centrifuged at 15,000 rpm for 5 min, followed by filtration of supernatant through a 0.45-μm nylon membrane filter. The filtrate obtained was loaded for sugar, organic acid, glycerol, and ethanol analysis using a mobile phase of 0.005 N H2SO4 at 60°C with a flow rate of 0.5 ml/min for 35 min. Serial standard solutions of compounds of interest were prepared and analyzed to establish standard calibration curves.
3
Acetic Acid Quantification in Tea and Kombucha
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Samples of tea and kombucha at 1, 7 and 14 days of fermentation were filtered through nylon filters (0.45 μm, diam. 25 mm; Sigma-Aldrich, Poznań, Poland). Acetic acid (AA) was analysed by high-performance liquid chromatography (HPLC) using a 1200 series HPLC connected to a 1100 series RI detector (Agilent Technologies, Santa Clara, CA, USA) with a Rezex ROA-Organic Acid H+ (8%) column (Phenomenex, Torrance, CA, USA). The column was eluted with a degassed mobile phase containing 5 mM H2SO4, pH 2.25 at 60 °C with a flow rate of 0.5 mL/min for 30 min per sample [19 , 20 (link)]. The results are shown in mg acetic acid/L.
4
Quantification of Extracellular Lactic Acid
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Supernatant samples of cultures were subjected to the D-/L-lactic acid (Rapid) assay (Megazyme), according to manufacturer protocol, adjusted for use in a 96-well plate as described earlier [10 (link)]. Occasionally, the extracellular lactic acid concentration was determined by HPLC analysis. HPLC samples were prepared by treating 500-μL supernatant samples with 50 μL of 35% perchloric acid (Merck), with incubation on ice for 10 minutes and the addition of 27 μL of 7 M KOH (Merck). After vortexing, the precipitate was removed by centrifugation for 5 minutes at 12,000 rpm and subsequent filtering (Sartorius Stedim Biotech, Minisart SRP4, 0.45 μm). Separation of organic acids was achieved by application of a 20-μL aliquot on a Rezex ROA-Organic Acid H+ (8%) column (Phenomenex), coupled to a refractive index detector (Jasco, RI-1530), using a flow of 0.5 mL/min and a column temperature of 45°C. The concentration was determined by comparison of the peak with known amounts of lactic acid (Megazyme).
5
Volatile Fatty Acid Analysis of Rumen Fluid
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Rumen fluid samples were analyzed for volatile fatty acids by diluting rumen fluid 1:1 in 5 mM H2SO4. The mixture was homogenized then centrifuged at 22,000 g for 7 min. The supernatant was then put through two 0.22 μm filters. The VFA concentration in rumen fluid of cows was measured using a high performance liquid chromatograph (Shimadzu, Kyoto, Japan) equipped with a degasser, a LC-20AT pump, a SIL-20A autosampler, a CTO-20A column oven and an SPD-M20A diode array detector. The mobile phase used was 5 mM sulfuric acid and VFAs were separated on a Rezex ROA-Organic Acid H+ (8%) column (300 × 7.8 mm) from Phenomenex. The conditions for liquid chromatography were as follows: samples were eluted into 5 mM sulfuric acid and 1.0 μL of sample was injected into the oven at 67°C at a rate of 0.850 mL/min. The absorption spectra of the compounds was recorded between 190 and 500 nm. Data was processed using LabSolutions CS software (Shimadzu).
6
Quantitative HPLC Analysis of Sugars and Organic Acids
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Sugars and organic acid content were determined using an Agilent 1100 series HPLC system, equipped with a diode array detector (DAD) and a refractometer index (RI) detector (Model 132; Gilson, Villiers-le-Bel, France), as previously described by Woznicki et al. [28 (link)]. The methanolic extracts (20 μL) were injected in a randomized order, and separation was performed on a Rezex ROA-Organic acid H+ (8%) column (300 × 7.8 mm; Phenomenex, Torrance, CA, USA) at 45 °C with mobile phase 7.2 mmol/L H2SO4 run at a flow rate of 0.5 mL/min. The detection of the sugars was performed with a RI detector and the organic acid detection was performed with DAD at 210 nm. Identification and quantification was performed according to the method described by Amundsen, et al. [24 (link)], and results were presented on a equivalent g/100 g fresh weight (fw) basis.
7
Quantifying 2-oxoglutarate in cultures
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2-oxoglutarate levels were measured as reported earlier [36 (link)]. Briefly, after measuring the ethylene in the headspace, the glass vials were de-capped and culture centrifuged for 10 min at 12,000 rpm and the supernatant subsequently filtered (Sartorius Stedim Biotech, Minisart SRP4, 0.45 μm). Separation of organic acids was achieved by application of a 20-μL aliquot on a Rezex ROA-Organic Acid H+ (8%) column (Phenomenex), coupled to a refractive index detector (Jasco, RI-1530), using a flow of 0.5 mL/min and a column temperature of 45 °C.
8
Measuring Sucrose and Monosaccharides
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Optical density was determined with a Libra S11 photometer (Biochrom) at λ = 600 nm. Samples were diluted to be in the linear range of the photometer. Sucrose concentrations and corresponding monomeric glucose and fructose were determined with a photometric assay kit from r-biopharm (Art. No. 10716260035). For measuring of glucose concentration, a second method was applied using an Agilent 1260 series HPLC and a Phenomenex REZEX ROA-Organic Acid H+ (8%) column (300 mm × 7.8 mm × 8 µm) at 65 °C with a RID detector and an isocratic gradient of 0.05 mM H2SO4 for 45 min.
9
Organic Acids Quantification by HPLC
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The collected fermentation broth was treated with 6 M HCl, heated at 80 °C for 30 min, and then filtered through a 0.22 μm filter membrane to obtain the supernatant samples. Organic acids were quantified by high-performance liquid chromatography using a standard high-performance liquid chromatography (HPLC) device (Agilent 1100 Series, Agilent Technologies Inc., Santa Clara, CA, USA) equipped with a Rezex ROA organic acid H+(8%) column (300 by 7.8 mm, 8 m; Phenomenex) and a Rezex ROA organic acid H+(8%) guard column (50 by 7.8 mm). The samples were analyzed at 35 °C with 0.5 mM H2SO4 as the mobile phase and a flow of 0.6 mL/min at a wavelength of 210 nm, according to a method described previously.
10
Quantifying Bacterial Metabolites by HPLC
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One milliliter of bacterial culture was centrifuged, and the supernatant was used for high‐performance liquid chromatography (HPLC) analysis. Metabolites were quantified using an HPLC system equipped with a model SPD‐M20A diode array detector model (Shimadzu, Kyoto, Japan) and a Rezex ROA‐Organic acid H+ (8%) column (Phenomenex, Torrance, CA, USA). The analytical conditions were as follows: eluent, 0.0025 N sulfuric acid; flow rate, 0.5 ml/min; detection, ultraviolet (UV) 210 nm; and column temperature 55°C. Succinate, propionate, and acetate were used as standards.
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