UV spectra were measured on a UV-2550 UV-visible spectrophotometer (Shimadzu, Shimane-ken, Japan). IR spectra were recorded on a 380 FT-IR spectrometer (Thermo Nicolet, Waltham, MA, USA). The optical rotations were measured on an AutoPol IV automatic polarimeter (Rudolph Research, Wilmington, MA, USA) at room temperature. 1D and 2D NMR data were recorded on a 600 MHz instrument (Varian, Palo Alto, CA, USA) with TMS as internal standard. HRESIMS data were acquired using a Triple TOF 6600 mass spectrometer (AB Sciex, Framingham, MA, USA). Semi-preparative HPLC separations were performed on a Chromaster system (Hitachi, Tokyo, Japan) consisting of a 5110 pump, 5210 autosampler, 5310 column oven, 5430 diode array detector and a Phenomenex Luna C18 column (250 × 10 mm, S-5 μm), all operated using EZChrom Elite software. All solvents were of ACS or HPLC grade, and were obtained from Tansoole (Shanghai, China) and Sigma-Aldrich (St. Louis, MO, USA), respectively. Silica gel (300–400 mesh), C18 reverse-phased Silica gel (150-200 mesh, Merck, Darmstadt, German), and MCI gel (CHP20P, 75–150 μM, Mitsubishi Chemical Industries Ltd., Tokyo, Japan) were used for column chromatography (CC), and pre-coated Silica gel GF254 plates (Qingdao Marine Chemical Plant, Qingdao, China) were used for TLC.
Chromaster system
Manufactured by Hitachi
Sourced in Japan
The Chromaster system is a high-performance liquid chromatography (HPLC) instrument manufactured by Hitachi. It is designed to perform precise separation, identification, and quantification of chemical components in complex samples. The Chromaster system incorporates advanced technology to ensure reliable and consistent analytical results.
Automatically generated - may contain errors
Lab products found in correlation
Tripletof 6600 mass spectrometer, by AB Sciex (2 mentions)
C18 reverse phased silica gel, by Merck Group (2 mentions)
Silica gel, by Qingdao Marine Chemical (2 mentions)
Avance 3 600 spectrometer, by Bruker (2 mentions)
Precoated silica gel gf254 plates, by Qingdao Marine Chemical (2 mentions)
Silica gel, by Merck Group (1 mentions)
600 mhz instrument, by Agilent Technologies (1 mentions)
Uv 2550 uv visible spectrophotometer, by Shimadzu (1 mentions)
Luna c18 column, by Hitachi (1 mentions)
Mightysil rp 18 gp, by Kanto Chemical (1 mentions)
Tensor 27 ftir spectrometer, by Bruker (1 mentions)
6540 qtof mass spectrometer, by Agilent Technologies (1 mentions)
Av 600 mhz spectrometer, by Bruker (1 mentions)
Sephadex lh 20, by GE Healthcare (1 mentions)
1200 series hplc system, by Agilent Technologies (1 mentions)
1290 uplc 6540 q tof mass instrument, by Agilent Technologies (1 mentions)
Mci gel, by Mitsubishi Chemical Corporation (1 mentions)
Chromolith highresolution rp 18 column, by Merck Group (1 mentions)
Agilent 6430 triple quadrupole mass spectrometry, by Agilent Technologies (1 mentions)
Agilent1260 infinity apparatus, by Agilent Technologies (1 mentions)
15 protocols using chromaster system
1
Spectroscopic Characterization of Compounds
2
Quantification of 734THIF and 784THIF in HepG2 Cells
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HepG2 cells were seeded in a 12-well plate (5 × 105/well) for 24 h and then cultured with 40 μM 734THIF or 784THIF for various periods under normoxic and hypoxic conditions in FBS-free medium. We collected HepG2 cells under normoxic (6 h and 12 h), hypoxic (6 h), post-hypoxic (6 h of normoxia followed by 6 h of hypoxia), and pre-hypoxic conditions (6 h of hypoxia followed by 6 h of normoxia). Harvested cells were lysed with 0.5% SDS, washed with methanol, and sonicated for 30 min. The levels of 734THIF and 784THIF in all cell samples passed through a 0.45-μm filter was determined using HPLC. The HPLC system (Chromaster system, Hitachi, Tokyo, Japan) contained a 5410 ultraviolet (UV) detector, 5210 auto sampler, 5110 pump, and a column (250 × 4.6 mm, 5 μm; Mightysil RP-18 GP, Kanto Chemical Co., Inc., Tokyo, Japan) was used to elute the compound. The mobile phase contained 10 mM KH2PO4 with acetonitrile (65:35, v/v; pH 2.8). The detection wavelength was 262 nm and the flow rate and injection volume were 1 ml/min and 20 μL, respectively. A standard curve of 734THIF or 784THIF generated using HPLC was used to calculate the concentration of each sample.
3
Spectroscopic Characterization of Compounds
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IR spectra were measured on a Bruker Tensor 27 FTIR spectrometer using KBr disks. NMR spectra were performed with a Bruker AV600 MHz spectrometer with TMS as an internal standard. HRMS–ESI data were obtained through an Agilent 1290 UPLC/6540 Q-TOF mass instrument. Column chromatography (CC) was performed using silica gel (30–400 mesh, Qingdao Marine Chemical Inc., China), Sephadex LH-20 (25–100 µm, Pharmacia Biotech Ltd., Sweden), and MCI gel (75–150 µm, Mitsubishi Chemical Corporation, Tokyo, Japan). Semipreparative HPLC was conducted on a HITACHI Chromaster system equipped with a DAD detector, a YMC-Triart C18 column (250 × 10 mm, i.d. 5 μm), and a flow rate of 3.0 mL/min. LC–MS was performed using an Agilent 1200 series HPLC system coupled to an Agilent q TOF 6540 mass spectrometer with a YMC-Triart C18 column (250 × 4.6 mm, i.d. 5 μm) and a flow rate of 1.0 mL/min.
4
Validated HPLC Analysis of Pharmaceutical Drugs
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All HPLC analyses were conducted using a Chromaster system (Hitachi High-Tech Science Corporation, Tokyo, Japan). The system consisted of a 5310 column oven, a 5210 autosampler, a 5110 pump, and a 5430 diode-array detector. The RP-HPLC separation of each drug (PHT, LTG, VCM, CBZ, or VRCZ) was performed at 40 °C (default temperature) on a Chromolith HighResolution RP-18 column (100 mm × 4.6 mm i.d.; Merck, Darmstadt, Germany) equipped with a guard column (5 mm × 4.6 mm i.d.). All HPLC analyses were performed in isocratic or gradient elution mode using a mixture of two mobile-phase solvents: A (CH3CN) and B (10 mM acetate buffer, pH 5.0). For example, an analyte containing CBZ (injection volume: 20 µL) was simply eluted with a solvent mixture (A:B = 40:60, v/v) at a flow rate of 2.0 mL/min over 3 min, and the CBZ in the eluate was detected using UV absorption at 280 nm. Details of the optimized HPLC conditions of all five drugs are listed in Tables S1 and S2 in the Supplementary data.
5
NMR and LC-MS Analysis of Compounds
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1D
and 2D (DQF-COSY, HSQC, HMBC, and NOESY) NMR spectra were recorded
on Bruker AVANCE III 600 spectrometers at 274 K. Chemical shifts were
reported with reference to the respective solvent peaks and the residual
solvent peaks (δH 3.31 and δC 49.0
for CD3OD). The NMR spectra were processed using the MestReNova
6.1.1 program. The Agilent series 1290 HPLC system equipped with Agilent
6430 triple quadrupole mass spectrometry was used for the quantitative
LC–MS analysis. Analytical HPLC was performed on an Agilent
1260 Infinity apparatus with a diode array detector. Preparative HPLC
was performed on a Hitachi Chromaster System.
and 2D (DQF-COSY, HSQC, HMBC, and NOESY) NMR spectra were recorded
on Bruker AVANCE III 600 spectrometers at 274 K. Chemical shifts were
reported with reference to the respective solvent peaks and the residual
solvent peaks (δH 3.31 and δC 49.0
for CD3OD). The NMR spectra were processed using the MestReNova
6.1.1 program. The Agilent series 1290 HPLC system equipped with Agilent
6430 triple quadrupole mass spectrometry was used for the quantitative
LC–MS analysis. Analytical HPLC was performed on an Agilent
1260 Infinity apparatus with a diode array detector. Preparative HPLC
was performed on a Hitachi Chromaster System.
6
Multimodal Analytical Techniques for Biomolecular Characterization
7
Spectroscopic Characterization of Natural Products
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IR spectra were recorded on a Nicolet 380 FT-IR spectrometer. The optical rotations were measured on an Auto Pol IV automatic polarimeter (Rudolph Research, Flanders, NJ, USA) at room temperature. 1D and 2D NMR data were recorded on a Varian 400 MHz instrument with TMS as internal standard. HR-ESI-MS data were acquired using a Triple TOF 6600 mass spectrometer (AB Sciex, Framingham, MA, USA). Semi-preparative HPLC separations were performed on a Hitachi Chromaster system consisting of a 5110 pump, 5210 autosampler, 5310 column oven, 5430 diode array detector, and a Phenomenex Luna C18 column (250 × 10 mm, S-5 μm), all operated using EZChrom Elite software. All solvents were of ACS or HPLC grade, and were obtained from Tansoole (Shanghai, China), Sigma-Aldrich (St. Louis, MO, USA), respectively. Silica gel (300−400 mesh), C18 reverse-phased silica gel (150–200 mesh, Merck, city, country), and MCI gel (CHP20P, 75−150 μM, Mitsubishi Chemical Industries Ltd., Tokyo, Japan) were used for column chromatography (CC), and pre-coated silica gel GF254 plates (Qingdao Marine Chemical Plant, Qingdao, China) were used for TLC.
8
HPLC Analysis of Lavender and Lavandin Extracts
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Qualitative and quantitative analyses of the lavender and lavandin extracts were performed using HPLC-DAD-UV/VIS. Separation of phenolic compounds and coumarins was achieved using the Hitachi Chromaster system (Tokyo, Japan) with a Purospher STAR RP-18e column (5 μm particle size, 250 mm × 4.6 mm, Merck) at 30 °C. The autosampler temperature was set at 20°C, and the sample injection volume was 20 μL. The mobile phase consisted of 0.1% (v/v) formic acid in water (A) and 0.1% (v/v) formic acid in acetonitrile (B) at a flow rate of 1 mL/min. The gradient conditions were as follows: 10–20% B (0–35 min), 20–35% B (35–60 min), 35–10% B (60–60.1 min), 10% B (60.1–70 min). The freeze-dried extracts (40 mg) were dissolved in 2 mL of 50% ethanol (ultrasound-assisted extracts and macerates) or water (decoctions). The chromatograms were recorded at 250, 280 and 330 nm. The peaks were identified by comparing their retention times and UV-Vis spectra with standards.
9
Kinetic and Mechanistic Analysis of α-Glucosidase
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The sample was prepared at different concentrations (0 mg, 0.2 mg, 0.4 mg and 0.8 mg) in Tris-HCl buffer and added to the α-glucosidase solution following the conditions of the AAG activity assay as described above. Then, 50 µL of pNPG at different concentrations (0.625 mM to 20 mM) was added to the mixture. After incubation, the optical density values of the final solutions were obtained at 410 nm and the reaction kinetics were investigated using the Lineweaver–Burk plot analysis [10 (link)].
HPLC analysis was also used to test the AAG mechanism. A mixture (1 mL) of the sample and α-glucosidase was incubated at 37 °C for 180 min. Every 30 min, 100 µL was collected for further measurements, including HPLC analysis and AAG activity. HPLC analysis was performed on a Hitachi Chromaster system (Column, KW802.5; mobile phase, 0.3 M NaCl in 20 mM Tris buffer (pH 7.0; sample volume, 20 µL; flow rate, 1.0 mL/min; detector, UV 280 nm, Taipei, Taiwan).
HPLC analysis was also used to test the AAG mechanism. A mixture (1 mL) of the sample and α-glucosidase was incubated at 37 °C for 180 min. Every 30 min, 100 µL was collected for further measurements, including HPLC analysis and AAG activity. HPLC analysis was performed on a Hitachi Chromaster system (Column, KW802.5; mobile phase, 0.3 M NaCl in 20 mM Tris buffer (pH 7.0; sample volume, 20 µL; flow rate, 1.0 mL/min; detector, UV 280 nm, Taipei, Taiwan).
10
Determination of 5-Fluorouracil by HPLC
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Preliminary 5‑FU determination conditions were developed using a Dionex system consisting of a pump type 7580, Jetstream II Plus (WO Industrial Electronics, Vienna, Austria) thermostat, DAD-detector UVD 340S. A pre-column Phenomenex C‑18 4 mm × 3 mm and column Phenomenex Luna C‑18 25 cm × 4.6 mm (particle size 5 µm) were used. The analytical wavelength was 266 nm according to the literature [38 (link)]. The mobile phase was composed of solvent A (H2O + 0.1% TFA) and solvent B (acetonitrile (ACN) + 0.1% TFA), and the gradient conditions were described in Table S1 (Supplementary Materials) .
Column temperature was 35 °C, the injection volume was 20 µL, and the flow rate was 1.0 mL/min. The described method was moved to a Hitachi Chromaster system consisting of a pump type 5160, thermostat 5310, DAD-detector UVD 5430, and autosampler model 5260. The method was validated, and the concentrations of 5‑FU were determined.
Column temperature was 35 °C, the injection volume was 20 µL, and the flow rate was 1.0 mL/min. The described method was moved to a Hitachi Chromaster system consisting of a pump type 5160, thermostat 5310, DAD-detector UVD 5430, and autosampler model 5260. The method was validated, and the concentrations of 5‑FU were determined.
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