Bile acids analysis was performed by UPLC-MS chromatography in an AQUITY I-Class (Waters Corp., Milford, MA) connected to a Xevo-G2 QTof MS detector. Separation was run on an ACQUITY UPLC BEH C18 1.7 mm column (2.1 × 100 mm, Waters Corp.) using water and acetonitrile as mobile phases, both containing a 0.1% of formic acid. Mass spectroscopy detection was performed in full-scan negative mode (100 to 1,200 Da). Concentration of bile acid was determined based on standard curves with QuanLynx software (Waters Corp.). Bile acids were extracted using the following methodologies and using chenodeoxycholic acid-d4 (CDCA-d4 ) as internal standard: lyophilized ileal digesta samples were homogenized in absence of solvent on a TissueLyzer II (QIAGEN, Hilden, Germany) and 20 mg of homogenate were extracted with 840 µL of H2 O: ACN (1:1) including internal standard, respectively. After homogenization, the mixture was centrifuged (15,000 g × 10 min, 4°C) and the supernatant diluted in H2 O: ACN (1:50) for UPLC-MS analysis. Plasma proteins were precipitated by addition of 200 µL of ACN with 5 µL internal standard to 50 µL of plasma. After centrifugation, supernatants were directly analyzed by UPLC analysis.
Aquity 1 class
Manufactured by Waters Corporation
The AQUITY I-Class is a high-performance liquid chromatography (HPLC) system designed for analytical laboratory applications. It provides precise and reliable separation and detection of chemical compounds in complex samples. The AQUITY I-Class features advanced technology to deliver consistent and accurate results, supporting a wide range of analytical workflows.
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Lab products found in correlation
2 protocols using aquity 1 class
1
Bile Acids Quantification by UPLC-MS
2
Bisphenol Quantification in Cell Culture Supernatants
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The supernatant of the cell cultures (approximately 800 µl) was transferred into 1.5 ml centrifuge tubes and stored at −18°C until analysis at the Environmental Agency Austria. For sample preparation, the supernatants were centrifuged (10 min, 17 000 rpm) to separate the solid matrix from the liquid phase. In total, 200 µl of the supernatant were spiked with an isotope labeled standard (BPA Ring d8) and analyzed using a Ultra High Performance Liquid Chromatography system coupled to triple quadrupole tandem mass spectrometry (Waters Aquity I Class and Waters Xevo TQ-S) in negative electrospray ionization mode. Chromatographic separation was performed on a reversed-phase column. Detection of the different bisphenols was conducted with typical fragment ions ([M–H]) for each compound in the multiple reaction mode. Quantification was performed using internal (for bisphenol A) and external calibration curves ranging from 0.5 ng/ml to 100 ng/ml. Higher concentrated supernatants were diluted with HPLC water to reach suitable concentrations. To check for matrix effects all dilution levels of the two cell lines were spiked with the bisphenols and recovery rates of this spiking-trial were considered in the calculation of the results.
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