title: Determination of nutlin-3a in murine plasma by liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS).
abstract: A sensitive and precise LC-ESI-MS/MS method for determination of nutlin-3a in murine plasma using ketoconazole as an internal standard was developed and validated. Plasma nutlin-3a samples were prepared by either a simple protein precipitation (PP) for the high concentration range (10-20,000ng/mL) or by liquid-liquid extraction (LLE) for the low concentration range (0.25-300ng/mL). Nutlin-3a and ketoconazole were separated on a modified C18 analytical column (4microm, 75mmx2mm) with an isocratic mobile phase (acetonitrile/5mM HCOONH(4)=70/30, v/v). The retention times of nutlin-3a and ketoconazole were 1.14 and 1.45min. Detection was achieved by a tandem MS system, monitoring m/z 582/99 and m/z 532/82 for nutlin-3a and ketoconazole, respectively. The PP method was linear in a range of 10-20,000ng/mL (R(2)>or=0.993) and the LLE method was linear in a range of 0.25-300ng/mL (R(2)>or=0.992). The mean recoveries for PP and LLE were 24% and 78%, respectively. Within-day and between-day precisions were <or=4.5% for PP and were <or=4.9% for LLE. Within-day and between-day accuracies (% error) ranged from 4.8 to -7.9 for PP, and from -0.2 to -8.4 for LLE. The two extraction methods produced equivalent results, allowing use of both within the same study. This method has been applied to the measurement of nutlin-3a concentrations in murine plasma samples obtained from a preclinical pharmacokinetic study.
authors.affiliation: Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
authors.forename: Feng
authors.initials: F
authors.lastname: Bai
authors.forename: Fangyi
authors.initials: F
authors.lastname: Zhu
authors.forename: Michael
authors.initials: M
authors.lastname: Tagen
authors.forename: Laura
authors.initials: L
authors.lastname: Miller
authors.forename: Thandranese S
authors.initials: TS
authors.lastname: Owens
authors.forename: Jeremy
authors.initials: J
authors.lastname: Mallari
authors.forename: Edward
authors.initials: E
authors.lastname: Derrick
authors.forename: Fan
authors.initials: F
authors.lastname: Zhang
authors.forename: Clinton F
authors.initials: CF
authors.lastname: Stewart
mesh_terms: D000284:Administration, Oral; D000818:Animals; D000970:Antineoplastic Agents / Q000008:administration & dosage / Q000097:blood* / Q000493:pharmacokinetics; D002021:Buffers; D011232:Chemical Precipitation; D002853:Chromatography, Liquid / Q000592:standards; D006863:Hydrogen-Ion Concentration; D007093:Imidazoles / Q000008:administration & dosage / Q000097:blood* / Q000493:pharmacokinetics; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D010879:Piperazines / Q000008:administration & dosage / Q000097:blood* / Q000493:pharmacokinetics; D012015:Reference Standards; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000592:standards; D053719:Tandem Mass Spectrometry / Q000592:standards
chemical_list: D000970:Antineoplastic Agents; D002021:Buffers; D007093:Imidazoles; D010879:Piperazines; C482205:nutlin 3