DNJ concentrations in colon tissue (tumor or normal) were determined using HILIC MS/MS.(17 (link)) In brief, a 1:10 dilution of colorectal cancer or normal tissue homogenate (500 µl), 100 µl of 0.1 µg/ml miglitol (internal standard) and 600 µl of acetonitrile were mixed by sonicating for 1 min and vortexing for 30 s. After centrifugation at 8,000 × g for 10 min at 4°C (CAX-370), the supernatant was collected. A 5-µl aliquot of the resulting extract was subjected to HILIC-MS/MS using a Shimadzu liquid chromatograph and a 4500 tandem mass spectrometer (AB Sciex, Tokyo, Japan). Under positive ion electrospray ionization conditions, MS/MS parameters were optimized with DNJ and miglitol. Samples (5 µl each) were separated on a HILIC column (TSK gel Amide-80, 4.6 mm × 150 mm; Tosoh, Tokyo, Japan), eluted with a mixture of acetonitrile and water (675:325, v/v) containing 6.5 mM ammonium formate (pH 5.5) at a flow rate of 0.2 ml/min and a temperature of 40°C. Post-column, DNJ was detected by HILIC-MS/MS with multiple reaction monitoring for transition of the parent ion to the product ion. DNJ concentrations were calculated using a calibration curve.
Liquid chromatograph
Manufactured by AB Sciex
Sourced in Japan
A liquid chromatograph is an analytical instrument used to separate, identify, and quantify components in a liquid sample. It operates by passing the sample through a column filled with a stationary phase, where the components are separated based on their interactions with the stationary phase and the mobile phase that is pumped through the column. This process allows for the identification and measurement of individual components within the sample.
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Lab products found in correlation
2 protocols using liquid chromatograph
1
Quantifying DNJ in Colorectal Cancer
2
LC-MS/MS Quantification of Dapsone
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Plasma samples were quantified for dapsone content using a validated, liquid chromatography/tandem mass spectrometry (LC‐MS/MS) analytical method that included a Shimadzu (Columbia, MD) liquid chromatograph and AB Sciex (Foster City, CA) 5500 QTRAP tandem mass spectrometer. Unknown plasma samples (100 μL) were transferred into a 96‐well plate with glass inserts to which 100 μL sulfamethizole (internal standard, IS) was added. The plate was removed and vortex mixed for 5 min. Then 500 μL of 80:20 ACN:MeOH was transferred to each insert, after which the samples were vortex mixed for 10 min then centrifuged for 10 min, 4°C at 3,300g. Next, 500 μL of the supernatant was transferred to a well plate with clean inserts and evaporated to dryness using a Labconco Centrivap. The dried samples were reconstituted with 100 μL of 50:50 MeOH:DIW and analyzed by reverse phase LC‐MS/MS. Dapsone and IS were quantitated using positive electrospray ionization (+ESI) combined with multiple reaction monitoring (MRM) for the respective precursor→product ion combinations of 249.0→155.9 m/z for dapsone and 271.0→155.9 m/z for IS. The standard curve was linear (r2 = 0.9942) and ranged from 5 ng/mL to 1,800 ng/mL.
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