NMR spectra (δ, ppm; J, Hz) were recorded on a Bruker Avance III-400 instrument (400.0 MHz for 1H and 101 MHz for 13C) using inverse broadband probe with ATM module (5 mm BBO-1H Z-GRD) or Bruker Avance III-400 instrument with broadband PRODIGY cryoprobe with ATM module (5 mm CPBBO BB-1H/19F/D Z-GRD). The NMR experiments were performed in DMSO-d6 or CDCl3 and referenced to the solvent signal (δ 2.50 and 39.70, respectively, 7.26 and 77.16). Mass spectra were measured on a LTQ Orbitrap XL (Thermo Fisher Scientific) using electrospray ionization (ESI). Column chromatography was performed on silica gel 60 (Fluka) and thin-layer chromatography (TLC) on silica gel 60 F254 foils (Merck). Solvents were evaporated at 2 kPa and bath temperature of 30–60 °C; the compounds were dried at 13 Pa and 50 °C. For all the tested compounds satisfactory elemental analysis was obtained supporting >95% purity as also visible in the NMR spectra. Optical rotation was measured on polarimeter Autopol IV (Rudolph Research Analytical) at 589 nm wavelength in chloroform or DMSO. UPLC samples were measured on Waters UPLC H-Class Core System (column Waters Acquity UPLC BEH C18 1.7 μm, 2.1 mm × 100 mm), Waters Acquity UPLC PDA detector, mass spectrometer Waters SQD2, and MassLynx mass spectrometry software. For reverse-phase flash column chromatography, C-18 RediSep Rf columns (Teledyne ISCO) were used.
Acquity uplc pda detector
Manufactured by Waters Corporation
Sourced in United States
The Acquity UPLC PDA detector is a high-performance liquid chromatography (HPLC) detector that utilizes a photodiode array (PDA) to measure the absorbance of analytes across a wide range of wavelengths. It is designed to provide sensitive and accurate detection for a variety of applications in analytical chemistry and life sciences research.
Automatically generated - may contain errors
Lab products found in correlation
Acquity uplc system, by Waters Corporation (3 mentions)
Ltq orbitrap xl, by Thermo Fisher Scientific (2 mentions)
Silica gel 60 f254 foils, by Merck Group (2 mentions)
Waters sqd2, by Waters Corporation (2 mentions)
Masslynx nt software v4, by Waters Corporation (2 mentions)
Avance 3 400, by Bruker (1 mentions)
Waters acquity uplc beh c18, by Waters Corporation (1 mentions)
Autopol 4, by Rudolph Research Analytical (1 mentions)
Acyclovir, by Fujifilm (1 mentions)
Avance 3 hd 500 mhz, by Bruker (1 mentions)
Acquity uplc h class system, by Waters Corporation (1 mentions)
Beh c18 column, by Waters Corporation (1 mentions)
Aquity uplc beh c18 column, by Waters Corporation (1 mentions)
Synapt g2 si q tof, by Waters Corporation (1 mentions)
Acquity uplc 1 class, by Waters Corporation (1 mentions)
Acquity uplc beh c18, by Waters Corporation (1 mentions)
Acquity uplc beh, by Waters Corporation (1 mentions)
8 protocols using acquity uplc pda detector
1
NMR and Mass Spectrometry Analysis of Organic Compounds
2
Vitamin C Quantification in Mice Plasma
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As described previously [23 (link), 35 (link)], vitamin C concentrations in WT or Svct1 KO mice plasma were determined using liquid chromatography-photodiode array (LC-PDA) analysis with an ACQUITY UPLC® PDA Detector (Waters, Milford, MA, USA) coupled with an ACQUITY UPLC System (Waters). After deproteinization with equal volumes of 10% (w/v) metaphosphate solution containing 1 mM EDTA, each sample was centrifuged at 20,000 × g at 4°C for 10 min,the obtained supernatant was diluted with equal volumes of 25 mM phosphate buffer (pH 2.1) containing 60 μM acyclovir (FUJIFILM Wako Pure Chemical) as an internal control and then separated on a CAPCELL PAK ADME S3 column (maintained at 50°C; 3 μm, 2.1 × 100 mm; Osaka Soda, Osaka, Japan) with a gradient mobile phase (0–4 min: 2% B; 4–5 min: 2–98% B; 5–9 min: 98% B; 9–10 min: 98–2% B; 10–12 min: 2% B) of 25 mM phosphate buffer (pH 2.1) (A) and methanol (B) at a flow rate of 300 μL/min. Vitamin C and acyclovir were measured at 243 nm in the PDA spectrum. Calibration curves for the analyte were generated using a series of murine plasma spiked with L-ascorbic acid (the reduced form of vitamin C) standard solutions. Peaks were analyzed using MassLynx NT software v4.1 (Waters).
3
Synthesis and Characterization of Thymine Derivatives
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Thymine
derivative5 and 2-MeNHA
derivative1 were synthesized using protocols published
previously.13 (link),68 (link)NMR spectra (δ, ppm; J, Hz) were measured on a Bruker Avance III HD 500 MHz instrument
equipped with a cryoprobe (500.0 MHz for 1H and 126 MHz
for 13C) in hexadeuterated dimethyl sulfoxide and referenced
to the solvent signal (δ 2.50 and 39.70). Mass spectra were
measured on an LTQ Orbitrap XL (Thermo Fisher Scientific) using electrospray
ionization. Column chromatography was performed on silica gel 60 (Fluka)
and thin-layer chromatography on silica gel 60 F254 foils (Merck).
Solvents were evaporated at 2 kPa and bath temperatures of 30–60
°C; the compounds were dried at 13 Pa and 50 °C. UPLC samples
were measured on the Waters UPLC H-Class Core System (column Waters
Acquity UPLC BEH C18 1.7 μm, 2.1 × 100 mm), Waters Acquity
UPLC PDA detector, mass spectrometer Waters SQD2, and MassLynx Mass
Spectrometry Software. For reverse-phase flash column chromatography,
C-18 RediSep Rf column Teledyne ISCO was used.
derivative
derivative
previously.13 (link),68 (link)NMR spectra (δ, ppm; J, Hz) were measured on a Bruker Avance III HD 500 MHz instrument
equipped with a cryoprobe (500.0 MHz for 1H and 126 MHz
for 13C) in hexadeuterated dimethyl sulfoxide and referenced
to the solvent signal (δ 2.50 and 39.70). Mass spectra were
measured on an LTQ Orbitrap XL (Thermo Fisher Scientific) using electrospray
ionization. Column chromatography was performed on silica gel 60 (Fluka)
and thin-layer chromatography on silica gel 60 F254 foils (Merck).
Solvents were evaporated at 2 kPa and bath temperatures of 30–60
°C; the compounds were dried at 13 Pa and 50 °C. UPLC samples
were measured on the Waters UPLC H-Class Core System (column Waters
Acquity UPLC BEH C18 1.7 μm, 2.1 × 100 mm), Waters Acquity
UPLC PDA detector, mass spectrometer Waters SQD2, and MassLynx Mass
Spectrometry Software. For reverse-phase flash column chromatography,
C-18 RediSep Rf column Teledyne ISCO was used.
4
UPLC Analysis of Protein Hydrolysates
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The
hydrolysates were analyzed on an H-class Acquity UPLC system
(Waters, Milford, MA) equipped with a BEH C18 column (1.7 μm,
2.1 × 100 mm, Waters), connected to an Acquity UPLC PDA detector
(Waters). The disulfide bridges were reduced by incubating the samples
at protein concentrations of 0.5% for 2 h with 100 mM dithiothreitol
(DTT) in 50 mM Tris-HCl buffer at pH 8.0. The reduced samples were
further diluted to protein concentrations of 0.1% (w/v) and centrifuged
(10 min, 14 000g, 20 °C) before injection
(4 μL).
hydrolysates were analyzed on an H-class Acquity UPLC system
(Waters, Milford, MA) equipped with a BEH C18 column (1.7 μm,
2.1 × 100 mm, Waters), connected to an Acquity UPLC PDA detector
(Waters). The disulfide bridges were reduced by incubating the samples
at protein concentrations of 0.5% for 2 h with 100 mM dithiothreitol
(DTT) in 50 mM Tris-HCl buffer at pH 8.0. The reduced samples were
further diluted to protein concentrations of 0.1% (w/v) and centrifuged
(10 min, 14 000g, 20 °C) before injection
(4 μL).
5
Quantitative Vitamin C Analysis in Biofluids
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Vitamin C levels in plasma, urine, CSF, and tissues were measured by LC-PDA analysis according to a previous report (Kondo et al., 2006 (link)), with some modifications.
In brief, all samples were analyzed on an ACQUITY UPLC PDA Detector (Waters, Milford, MA, USA) coupled with an ACQUITY UPLC System (Waters). A volume of 5 μL of each sample was injected into a CAPCELL PAK ADME S3 column (3 μm, 2.1 × 100 mm; Osaka Soda, Osaka, Japan) and separated. Elution was conducted using a gradient mobile phase (0–4 min: 2% B; 4–5 min: 2–98% B; 5–9 min: 98% B; 9–10 min: 98–2% B; 10–12 min: 2% B) of 25 mM phosphate buffer (pH 2.1) (A) and methanol (B) at a flow rate of 300 μL/min. The column was maintained at 50°C. During the separation, the PDA spectrum was obtained, and 243 nm was chosen for measurement of VC and acyclovir. Calibration curves for the analyte were generated from a series of murine plasma spiked with standard solutions of VC (L-ascorbic acid). Peak analyses were conducted using MassLynx NT software v4.1 (Waters).
In brief, all samples were analyzed on an ACQUITY UPLC PDA Detector (Waters, Milford, MA, USA) coupled with an ACQUITY UPLC System (Waters). A volume of 5 μL of each sample was injected into a CAPCELL PAK ADME S3 column (3 μm, 2.1 × 100 mm; Osaka Soda, Osaka, Japan) and separated. Elution was conducted using a gradient mobile phase (0–4 min: 2% B; 4–5 min: 2–98% B; 5–9 min: 98% B; 9–10 min: 98–2% B; 10–12 min: 2% B) of 25 mM phosphate buffer (pH 2.1) (A) and methanol (B) at a flow rate of 300 μL/min. The column was maintained at 50°C. During the separation, the PDA spectrum was obtained, and 243 nm was chosen for measurement of VC and acyclovir. Calibration curves for the analyte were generated from a series of murine plasma spiked with standard solutions of VC (L-ascorbic acid). Peak analyses were conducted using MassLynx NT software v4.1 (Waters).
6
Quantification of Naproxen Levels in Gastrocnemius Tissue
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At the end of treatment, animals were sacrificed and gastrocnemius samples were removed, immediately frozen in liquid nitrogen and stored at −80 °C until analyzed. Then, each sample was homogenized in a mortar in the presence of liquid nitrogen and three volumes of ACN were added. Thereafter, samples were vortex-mixed and centrifuged for 10 min at 4 °C (3200 g). The supernatant was transferred to a clean tube and analyzed by LC-PDA for naproxen level quantification. Data were reported as ng of naproxen in mg of tissue.
Liquid chromatography was performed on ACQUITY UPLC system (Waters Corp., Milford, MA) with autosampler and column oven enabling temperature control of analytical column. AQUITY UPLC BEH C18 column (2.1x50mm, 1.7 μm; Waters Corp., Milford, MA) was employed. The column temperature was maintained at 40 °C. A 1.7-min linear gradient from 60 to 0 % of mobile phase A (formic acid 0.1 %) was used at the flow rate of 0.5 ml/min, and mobile phase B was methanol containing 0.1 % of formic acid.
PDA detection was carried out on ACQUITY UPLC PDA detector (Waters Corp., Milford, MA). Wavelength of 230 nm was used to monitor naproxen levels in the experiment.
Liquid chromatography was performed on ACQUITY UPLC system (Waters Corp., Milford, MA) with autosampler and column oven enabling temperature control of analytical column. AQUITY UPLC BEH C18 column (2.1x50mm, 1.7 μm; Waters Corp., Milford, MA) was employed. The column temperature was maintained at 40 °C. A 1.7-min linear gradient from 60 to 0 % of mobile phase A (formic acid 0.1 %) was used at the flow rate of 0.5 ml/min, and mobile phase B was methanol containing 0.1 % of formic acid.
PDA detection was carried out on ACQUITY UPLC PDA detector (Waters Corp., Milford, MA). Wavelength of 230 nm was used to monitor naproxen levels in the experiment.
7
Identification of PHNQs via UPLC-PDA-ESI-HRMS
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The identification of the PHNQs in the extract was carried out using UPLC-PDA-ESI-HRMS. The UPLC-PDA-ESI-HRMS analyses were performed by the Unitech COSPECT Mass Spectrometry facility (University of < city>Milan < /city>, Italy). The UPLC equipment was Acquity UPLC I Class (Waters, Milford, MA, USA), and the following conditions were used: column: ACQUITY UPLC BEH C18 (50 × 2.1 mm, 1.7 um) (Waters, USA), column temperature: 34°C, and eluents were A: water + 0.1% formic acid and B: acetonitrile + 0.1% formic acid. The flow rate was set at 0.5 ml min−1 and the linear gradient elution was set: START: 95% A: 5% B; 5.15 min: 60% A: 40% B; 6 min: 10% A: 90% B; 6.30 min: 10% A: 90% B; 6.60 min: 95% A: 5% B; 10 min: 95% A: 5% B. The sample temperature was kept constant at 20°C and the injection volume was 4 μl. The detector was an Acquity UPLC PDA Detector (Waters, Milford, MA, USA) working in a wavelength range of 210–500 nm. The HRMS detector was a Synapt G2-Si QTof (Waters, USA). The operative parameters are given in the following: ESI Ionization mode, negative ionization polarity, full scan range 50–1,200 m/z, leucine enkephalin was the lock mass compound. The used software was MassLynxTM v4.2 (Waters, Milford, MA, USA).
8
UPLC-PDA Analysis of Compounds
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Analyses were performed using an ACQUITY UPLC H-Class Core System (Waters) with an ACQUITY UPLC PDA Detector, according to Gontijo et al. [33] , in the Faculdade de Farmácia, UFMG, equipped with a reversed-phase column ACQUITY UPLC BEH (1.7 μm, 50 × 2 mm i.d.; Waters). The mobile phase consisted of water 0.1 % formic acid (solvent A) and acetonitrile 0.1 % formic acid (solvent B). The elution protocol was 0-13 min, linear gradient from 5 to 95 % B. The flow rate was 0.3 mL min -1 , and the sample injection volume was 4.0 μL. The UV spectra were registered from 200 to 400 nm.
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