The LC system consisted of a Prominence LC-20A series (Shimadzu Corp., Kyoto, Japan). Each of the BcS derivatives was purified using the following LC conditions: mobile phase, water containing 0.1% trifluoroacetic acid/acetonitrile; linear gradient of 5–21% acetonitrile in 12 min, hold at 21% acetonitrile for 3 min, followed by equilibration in 5% acetonitrile for 15 min before the next injection. The flow rate was 4 mL/min, and the column used was a 250 × 10 mm i.d., 5 μm, Inertsil ODS-3 (GL Sciences Inc., Tokyo, Japan). UV detection was set at 260 nm. The column oven was held at 40 °C. The retention times of MeBcS, DahMeBcS, PyBcS, cytomycin, and 7, were 7.8, 8.9, 9.3, 8.9, and 6.2 min, respectively. The derivatives were obtained as trifluoroacetate salts. In order to remove trifluoroacetate, each compound was subjected to the LC system [column, 250 × 10 mm i.d., 5 μm, InertSustain AQ-C18 (GL Sciences Inc.); isocratic elution of 5% acetonitrile in water; flow rate, 4.0 mL/min; detection at 260 nm]. The stock solution (50 mM) was prepared by dissolving each derivative in water.
Inertsustain aq c18
Manufactured by GL Sciences
Sourced in Japan
The InertSustain AQ-C18 is a high-performance liquid chromatography (HPLC) column designed for the analysis of a wide range of polar and non-polar analytes. The column features a silica-based stationary phase with a chemically bonded C18 alkyl ligand, providing a stable and reproducible separation performance. This column is suitable for use in aqueous mobile phase conditions.
Automatically generated - may contain errors
Lab products found in correlation
Inertsil ods 3, by GL Sciences (1 mentions)
Prominence lc 20a series, by Shimadzu (1 mentions)
Hplc system, by Jasco (1 mentions)
Inertsil ods 4v, by GL Sciences (1 mentions)
L 7400 uv detector, by Hitachi (1 mentions)
F 1050 fluorescence spectrophotometer, by Hitachi (1 mentions)
Model 234 autoinjector, by Gilson (1 mentions)
L 7100 pump, by Hitachi (1 mentions)
Lcms 8060, by Shimadzu (1 mentions)
Pu 2085, by Jasco (1 mentions)
Mx 2080 32, by Jasco (1 mentions)
Tristar 2 surface area and porosity analyzer, by Micromeritics (1 mentions)
Lc netii adc interface box, by Jasco (1 mentions)
Agilent 1100 series system, by Agilent Technologies (1 mentions)
L anserine nitrate, by Fujifilm (1 mentions)
7 protocols using inertsustain aq c18
1
HPLC Purification of Benzacridine Derivatives
2
HPLC Analysis of SBD-Thiols
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HPLC analysis was performed on Jasco HPLC systems (Tokyo, Japan). SBD-thiols were separated on Inertsil ODS-4V (250 × 3.0 mm I.D., 5 μm) or InertSustain AQ-C18 (250 × 3.0 mm I.D., 5 μm) (GL Sciences, Tokyo, Japan). 100 mM citric buffer (pH 3.0)/MeOH was used as mobile phase and flow rate was 0.3 mL/min. The column temperature was 40 °C. Fluorescence detection was performed with excitation and emission wavelengths of 375 and 510 nm, respectively. Gain of the fluorescence detector was programmed as follows, 0–15.5 min: 1000, 15.5–21 min: 10, and 21–30 min: 100.
3
HPLC Analysis of Flavin Cofactors
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Flavin mononucleotide (FMN), flavin adenine dinucleotide (FAD), and riboflavin (RF) in the perfusate of HMP were measured using a high-performance liquid chromatography (HPLC) method. The sample (perfusate) was deproteinized by mixing it with pure methanol, followed by centrifugation and filtration. An aliquot of the resulting deproteinized supernatant was directly injected into the HPLC system. The HPLC system consisted of a GASTORR BG-42 degasser (FLOM Co., Tokyo, Japan), L-7100 pump (Hitachi High-Tech Corporation, Tokyo, Japan), model 234 autoinjector (GILSON Inc., Middleton, WI, USA), ATC-10 column oven (Eicom, Kyoto, Japan), L-7400 UV detector (Hitachi), NOD-10 UV detector (Eicom), and F-1050 fluorescence spectrophotometer (Hitachi) with C18 column for reverse phase HPLC analysis InertSustain AQ-C18 (5 µm) and a guard column E (GL Science, Tokyo, Japan). The HPLC conditions were as follows: ODS column (EICOMPAK SC5-ODS; 3 µm, 150 × 4.6 mm), column oven (40 °C), UV–Vis detector (254 nm), fluorescence detector (excitation 445 nm, emission 530 nm), mobile phase (A. methanol, B. acetic acid buffer. A/B = 35/65, v/v), where the acetic acid buffer was a mixture of 4 M sodium acetate (20 mL) and 50% acetic acid (10 mL) in 1 L of deionized water; flow (0.7 mL/min), and injection volume (25 µL). FMN, FAD, and RF contents were expressed as mmol/L.
4
Quantitative Analysis of Peptides by LC-TQMS
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Quantification of the two peptides of interest (see in Results ) was performed by LC-triple quadrupole mass spectrometry (LC-TQMS; LC-MS-8060, Nexela system, Shimadzu, Kyoto, Japan). Samples were injected and separated by reversed-phase chromatography using InertSustain AQ-C18 (150 mm, 2.1 mm ID, 2.0 μm particle size, GL Science, Osaka, Japan) at a flow rate of 400 μL/min. The gradient was obtained by changing the mixing ratio of two eluents: A, 0.1% (v/v) formic acid and B, acetonitrile containing 0.1% (v/v) formic acid. The 23 min gradient separation at a flow rate of 400 μL/min and 40°C was as follows: (i) 5% B for 2 min, (ii) a 15-min gradient to 45% B, (iii) an immediate increase to 95% B, (iv) a 3 min hold at 95% B, (v) an immediate decrease back to 5% B, and (vi) a 3-min hold at 5% B. The parameters of Multiple Reaction Monitoring were set as follows: (i) polarity, positive; transition, 358.15 > 86.05; Q1 Pre-bias: −20 V; collision energy (CE), −30 V; Q3 Pre-bias: −20 V for Leu–Leu–Leu and (ii) polarity, positive; transition, 376.20 > 229.20; Q1 Pre-bias: −20 V; collision energy (CE), −20 V; Q3 Pre-bias: −20 V for Phe–Pro–Leu.
Relative quantification of C19H28O, C17H26N2O3, [Leu/Ile]–[Leu/Ile]–[Leu/Ile]–Pro, and [Leu/Ile]–[Leu/Ile]–[Leu/Ile]–[Leu/Ile]–Pro was performed using extracted ion chromatograms of metabolome profiles obtained by nanoLC-MS.
Relative quantification of C19H28O, C17H26N2O3, [Leu/Ile]–[Leu/Ile]–[Leu/Ile]–Pro, and [Leu/Ile]–[Leu/Ile]–[Leu/Ile]–[Leu/Ile]–Pro was performed using extracted ion chromatograms of metabolome profiles obtained by nanoLC-MS.
5
HPLC Analysis of Nucleobase Sorbent Recovery
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The recovery of each sorbent was evaluated by an HPLC system as follows: a GD-2080-53 degasser, two PU-2085 pumps, an MX-2080-32 dynamic mixer, an AS-4050 autosampler, a CO-2065 Plus column oven, a UV-2070 UV-Vis detector, and an LC-Net II/ADC interface box (Jasco). An InertSustain AQ-C18 (3 μm, 150 × 2.1 mm i.d., GL Sciences) was used for the separation on HPLC. The separation conditions on HPLC were as follows: flow rate, 0.2 mL min -1 ; mobile phase, a 5% methanol solution (for the evaluation of Ura, U, and Ade) or a 10% methanol solution (for the evaluation of A, C, and G); column temperature, 40°C; and injection volume, 10 μL. Elemental analysis was performed by JM11 (J-SCIENCE LAB CO., Ltd.). The specific surface area of each base resin was measured by a Micromeritics TriStar II Surface Area and Porosity Analyzer.
6
Quantifying Imidazole Dipeptides in Meat Samples
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The imidazole dipeptides, anserine and carnosine, were measured by HPLC. Digested meat samples were centrifuged at 7000× g for 5 min. Then, 0.5 mL of the supernatant from each sample was ultra-filtered at 15,000× g for 20 min (Nanosep 10K OMEGA; Pall Corp., New York, NY, USA) to obtain the under-10-kDa fractions. Each fraction was adjusted to 0.5 mL and analyzed with the HPLC Agilent SERIES 1100 system (Agilent Technologies Inc., Santa Clara, CA, USA). For the analysis of anserine and carnosine, the tested solution was injected into a reversed-phase column (InertSustain AQ-C18; GL Sciences Inc., Tokyo, Japan). Elution was performed at 30 °C with 0.2 M ammonium dihydrogenphosphate, 0.1 mM 1-pentanesulfonic acid sodium salt, and 4% acetonitrile solution and adjusted to pH 2.0 with HCl at a flow rate of 0.8 mL min−1. Anserine and carnosine were detected by measuring the absorbance at 220 nm. A solution containing 5.0 mM L -anserine nitrate (Fujifilm Wako Pure Chemical) and 5.0 mM carnosine (β-Alanyl-L -Histidine, Peptide Institute, Inc., Osaka, Japan) was used as the standard.
7
HPLC-PR-CL for N-Nitrosamine Detection
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N-nitrosamine concentrations were determined by HPLC-PR-CL. This method is based on the chemiluminescence reaction between peroxynitrite with luminol. Peroxynitrite is formed by the photochemical reaction of N-nitrosamines with UV irradiation at 254 nm after HPLC separation. The HPLC separation was performed with an InertSustain AQ-C18 (5 μm, 4.6 × 250 mm) (GL Sciences, Tokyo, Japan) with an eluent of 5 mM phosphate buffer and methanol (95:5 v/v). Further details of this method are provided elsewhere (Fujioka et al., 2016; Kodamatani et al., 2016) . A sample HPLC-PR-CL chromatogram of the separation of NDMA, NMOR, NMEA and NPYR is shown in Fig. S2. Each sample from the RO feed was pre-filtered with a 0.45 µm hydrophilic PTFE syringe filter (Filtstar, Starlab Scientific, China). The sample injection volume was from 20 to 200 µL.
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