title: Rapid determination of tacrine and other drug metabolites in microsomal incubate by newly developed targeting algorithm on UHPLC/TOFMS.
abstract: A rapid simultaneous determination method for in vitro Cytochrome P450 (CYP) activity assay of 1,2,3,4-tetrahydroacridin-9-amine (tacrine) metabolites using ultra high performance liquid chromatography (UHPLC) coupled with computer-assisted in-source collision induced dissociation (CID) monitoring was investigated. In general, enzyme inhibition assays require quantitative analysis of incubates with drugs using various concentrations of substrates/inhibitors. The assay of CYP isozyme inhibition is an important informational step in the drug discovery process and, with the many substrates listed by the FDA, high-throughput qualitative and quantitative analyses are desired. Based on sub-2-micron packing material with reversed phase chromatography combined with a single liquid chromatography/time-of-flight mass spectrometry (LC/TOFMS), a less than 1min analysis time is presented for two additional drugs. We successfully determined seven of the eight potential isomer metabolites for the drug tacrine in 2.5min using a 2mm internal diameterx100mm length column and applying in-source CID with our newly developed chromatographic peak deconvolution technique. Although two of the peaks were heavily fused at a peak width of less than 300ms, we could clearly identify these peaks by monitoring the chromatographic intensity difference of their fragment peaks on the mass spectrum.
authors.affiliation: JEOL Analytical Science, LLC, 5843 Stoddard Rd, Suite 2, Modesto, CA 95356, United States. kris.cloos@gmail.com
authors.forename: Kristie C
authors.initials: KC
authors.lastname: Cloos
authors.forename: Toshinobu
authors.initials: T
authors.lastname: Hondo
mesh_terms: D000465:Algorithms; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000191:economics / Q000379:methods*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D013058:Mass Spectrometry / Q000191:economics / Q000379:methods*; D008862:Microsomes, Liver / Q000737:chemistry*; D018697:Nootropic Agents / Q000032:analysis* / Q000378:metabolism; D013619:Tacrine / Q000032:analysis* / Q000378:metabolism; D013997:Time Factors
chemical_list: D018697:Nootropic Agents; D013619:Tacrine; D003577:Cytochrome P-450 Enzyme System