Automated online liquid chromatographic/mass spectrometric metabolic study for prodrug stability
Fran Lai, , and Siamak Cyrus Khojasteh-Bakht Drug Metabolism and Pharmacokinetics, Genentech Inc., 1 DNA Way, M/S 70, South San Francisco, CA 94080, USA Received 4 June 2004; accepted 13 October 2004. Available online 11 November 2004.
Abstract In vitro metabolic stability studies are performed routinely in drug discovery to determine the rate of metabolism as well as the metabolic fate of compounds. These studies are labor intensive, involving incubation of the compound with a biological matrix, sampling at various time points, stopping the reaction, and sample preparation for analysis. All of these steps involve manual pipetting in the conventional method. An automated method for in vitro metabolism studies is reported here. The method reduces the time and manual labor required and has other advantages, such as better reproducibility and unattended operation. This method utilizes an autosampler custom configured with cooling and incubation capabilities. The autosampler is programmed to directly inject incubation samples at set time points onto an online extraction column. The extracted sample then enters an analytical column for separation and ultimately the mass spectrometer for detection. The injection has the dual function of stopping the reaction and starting the analysis on the LC-MS. This method was used for the metabolic stability study of a prodrug in plasma and liver S9 fractions of five different species. The stability data from the automated method were similar to those obtained using the conventional method. The potential for this method to increase throughput of metabolic stability studies in drug discovery is demonstrated.
Increasing throughput of parallel on-line extraction liquid chromatography/electrospray ionization tandem mass spectrometry system for GLP quantitative bioanalysis in drug development
RAPID COMMUNICATIONS IN MASS SPECTROMETRY 2004; 18: 285-292
Showchien Hsieh 1/*, Tom Tobien2, Kevin Koch1 and John Dunn1 1GlaxoSmithKline, Inc., Research Triangle Park, NC 27709, USA 2Leap Technologies, P.O. Box 969, Carrboro, NC 27510, USA Received 12 September 2003; Revised 18 November 2003; Accepted 18 November 2003
Abstract An approach is described with turbulent flow on-line extraction liquid chromatography/electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) for GLP quantitative bioanalysis of a drug candidate. Two systems were built in-house with standard laboratory parts and equipments. One system consisted of one gradient HPLC pump, one isocratic pump, one ten-port valve, two turbulent flow columns, one analytical column, one autosampler and one mass spectrometer. Using this system, an injection-to-injection cycle time of 0.8 min was achieved. By adding an additional valve, another analytical column and an isocratic pump, the injection-to-injection cycle time decreased to 0.4 min. Validation results from the two systems showed that precision and accuracy were acceptable for GLP quantitative analyses. The system was utilized to support sample bioanalysis of a drug candidate in a first-time in-human clinical trial.
Full article available upon request.
Integrated high capacity solid phase extraction-MS/MS system for pharmaceutical profiling in drug discovery
Journal of Pharmaceutical and Biomedical Analysis 34 (2004)1-9 Edward H. Kerns a,, Teresa Kleintop a, David Little b, Thomas Tobien c, Larry Mallis d,LiDia, Mei Hu a, Yan Hong a, Oliver J. McConnell d a Wyeth Research, CN8000, Princeton, NJ 08543-8000, USA b Micromass, Manchester, UK c LEAP Technologies, Carrboro, NC, USA d Wyeth Research, Collegeville, PA, USA Received 16 September 2002; received in revised form 20 March 2003; accepted 1May2003
Abstract A method is described for use in analysis of samples from pharmaceutical profiling of early drug discovery compounds. The method consists of a high capacity autosampler which injects samples into one of two solid phase extraction columns operated in parallel for alternating trapping, washing and elution into a tandem quadrupole mass spectrometer operated in multiple reaction monitoring (MRM) MS/MS mode. A primary method, which is useful for 80-90% of compounds, and a secondary method, which is useful for a majority of the remaining compounds, are described. No analytical HPLC column is used and the analysis rate is approximately 50 samples/h. Specificity is obtained using MRM analysis. Application of the method for high capacity analysis of metabolic stability samples is described.
Full article available upon request.
Using the PAL Autosampler as a Syringe Pump for MS Tuning
LEAP Application Note
Revision: 1.0 Date: 06-21-2004 Author: Thomas Tobien
Objective/Abstract
The PAL autosampler injection syringe is utilized for analyte infusion during mass spectrometer tuning. Infusion occurs either into an LC mobile phase stream (Option A) or directly into the LC interface of a mass spectrometer (Option B). After installation of a large volume syringe, a local firmware method and job is created that controls analyte infusion.
