ABSTRACT
A liquid-chromatographic-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the determination of GDC-0879 and its ketone metabolite (M1) in dog plasma to support preclinical toxicokinetic evaluation. The method consisted of solid phase extraction for sample preparation and LC-MS/MS analysis in positive ion mode using electrospray ionization for analysis. D(4)-GDC-0879 and (13)C(2)-D(2)-M1 were used as internal standards. A quadratic regression (weighted 1/concentration(2)) was used to fit calibration curves over the concentration range of 1-1000 ng/ml for both GDC-0879 and M1. The accuracy (%bias) at the lower limit of quantitation (LLOQ) was 12.0% and 2.0% for GDC-0879 and M1, respectively. The precision (%CV) for samples at the LLOQ was 11.3% and 2.6% for GDC-0879 and M1, respectively. For quality control samples at 3.00, 400 and 800 ng/ml, the between run %CV was ≤3.9% for GDC-0879 and ≤2.4% for M1. Between run %bias ranged from 4.6 to 12.0% for GDC-0879 and from -0.8 to 2.7% for M1. GDC-0879 and M1 were stable in dog plasma for at least 44 days at -70 °C.
Subject(s)
Antineoplastic Agents/blood , Chromatography, Liquid , Indenes/blood , Protein Kinase Inhibitors/blood , Pyrazoles/blood , Solid Phase Extraction , Tandem Mass Spectrometry , Animals , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/toxicity , Biotransformation , Calibration , Chromatography, Liquid/standards , Dogs , Drug Stability , Female , Indenes/administration & dosage , Indenes/pharmacokinetics , Indenes/toxicity , Infusions, Intravenous , Ketones/blood , Limit of Detection , Male , Protein Kinase Inhibitors/administration & dosage , Protein Kinase Inhibitors/pharmacokinetics , Protein Kinase Inhibitors/toxicity , Pyrazoles/administration & dosage , Pyrazoles/pharmacokinetics , Pyrazoles/toxicity , Reference Standards , Regression Analysis , Reproducibility of Results , Solid Phase Extraction/standards , Spectrometry, Mass, Electrospray Ionization , Tandem Mass Spectrometry/standards , raf Kinases/antagonists & inhibitorsABSTRACT
The experimental and computational mechanistic details of the Pd(OAc)(2)/TEA-catalyzed aerobic alcohol oxidation system are disclosed. Measurement of various kinetic isotope effects and the activation parameters as well as rate law derivation support rate-limiting deprotonation of the palladium-coordinated alcohol. Rate-limiting deprotonation of the alcohol is contrary to the majority of related kinetic studies for Pd-catalyzed aerobic oxidation of alcohols, which propose rate-limiting beta-hydride elimination. This difference in the rate-limiting step is supported by the computational model, which predicts the activation energy for deprotonation is 3 kcal/mol higher than the activation energy for beta-hydride elimination. The computational features of the similar Pd(OAc)(2)/pyridine system were also elucidated. Details of the study illustrate that the use of TEA results in an active catalyst that has only one ligand bound to the Pd, resulting in a significant lowering of the activation energy for beta-hydride elimination and, therefore, a catalyst that is active at room temperature.
