Journal
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
Volume 856, Issue 1-2, Pages 261-266Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jchromb.2007.06.006
Keywords
5-MI; indirubin; cyclin-dependent kinases; liquid chromatography; tandem mass spectrometry
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A sensitive and rapid LC-MS/MS assay for the quantitative determination of 5-methylindirubine (5-MI) in murine plasma is described. A 50-mu L-murine plasma aliquot was spiked with an internal standard, indirubine-3-monoxime (IMO), and extracted with 1.25 mL diethyl ether. Dried extracts were reconstituted in methanol-water (8:2, v/v) and 10 mu L-volumes were injected onto the HPLC system. Separation was achieved on a Gemini C18 column (150 mm x 2.1 mm ID, particle size 5 mu m) using an alkaline eluent (10 mM ammonium hydroxide-methanol (5:95, v/v)). Detection was performed by negative ion electrospray followed by tandem mass spectrometry. The assay quantifies 5-MI in a range from I to 500 ng/mL using 50 mu L of murine EDTA plasma samples. Validation results demonstrate that 5-MI concentrations can be accurately and precisely quantified in murine plasma. This assay is used to support pre-clinical pharmacologic studies with 5-MI. (C) 2007 Elsevier B.V. All rights reserved.
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