RESUMEN
A high-throughput bioanalytical method using automated sample transferring, automated liquid-liquid back extraction and liquid chromatography-tandem mass spectrometry was developed in a GLP regulated environment for the determination of ABT-202 in human plasma. Samples of 0.30 ml were transferred into 96-well plate using an automatic liquid handler. Automated liquid-liquid extraction (LLE) was carried out on a 96-channel programmable liquid handling workstation using methyl tert-butyl ether as the extraction solvent. A dual-HPLC with single mass spectrometer configuration was utilized to provide a reliable and routine means to increase sample throughput. The standard curve range was 0.38-95.02 ng/ml. There was no interference from endogenous components in the blank plasma tested. The accuracy (% bias) at the lower limit of quantitation (LLOQ) was 7.7% and the precision (% CV) for samples at the LLOQ was 4.7%. The inter-day % CV and % bias of the quality control samples were < or = 6.8 and < or = 7.6%, respectively. Coefficients of determination, a measure of linearity, ranged from 0.994 to 0.997. The method was accurate and reproducible and was successfully applied to generate plasma concentration-time profiles for human subjects after low oral doses of the compound.
Asunto(s)
Cromatografía Liquida/métodos , Espectrometría de Masas/métodos , Agonistas Nicotínicos/sangre , Piridinas/sangre , Administración Oral , Automatización , Calibración , Humanos , Nicotina/sangre , Agonistas Nicotínicos/farmacocinética , Estándares de Referencia , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Manejo de EspecímenesRESUMEN
A mecA-containing Staphylococcus aureus strain was grown in the presence of high concentrations of D-serine, D-threonine, and D-phenylalanine. These growth conditions resulted in the replacement of the carboxyl-terminal (fifth) D-alanine residue of peptidoglycan stem peptides with the D-amino acid present in the growth medium and a reduced ability to grow in the presence of methicillin. The most dramatic effect was seen with D-serine. With 32 mM D-serine, strains that had been able to grow in the presence of 800 micro g of methicillin per ml were only able to grow in the presence of less than 50 micro g/ml. The results also suggest that in S. aureus vancomycin resistance mediated through the incorporation of precursors not terminating in D-alanyl-D-alanine would be mutually exclusive with expression of mecA-mediated methicillin resistance.