Abstract
A high-throughput semi-automated procedure for simultaneously stability evaluation of multiple compounds in plasma using direct single column high-performance liquid chromatography (HPLC) combined with tandem mass spectrometry (MS/MS) was developed to eliminate the laborious procedures that are traditionally used for stability studies. Untreated human, monkey, mouse and rat plasma samples containing ten drug components were directly injected into a mixed-functional column that provided both protein removal and chromatographic functionality. Ten test compounds were simultaneously assayed using a tandem mass spectrometer in the positive ion mode using multiple reaction monitoring (MRM). Plasma samples containing ten test compounds were placed in a thermostatic autosampler and then sequentially monitored in one analytical procedure. The time between each injection was set about 7 minutes. The peak responses of the test compounds in individual plasma samples were repeatedly determined every 28 minutes. Drug stability in plasma was indicated by the change of the mass chromatographic peak areas for the test compounds and was observed to be a function of animal species, incubation time and incubation temperature. The potential for matrix ionization suppression on the direct single column HPLC-MS/MS system was also investigated using the post-column infusion technique. The proposed cassette assay procedure provides an analytical throughput ten times greater than the single component approach for the evaluation of drug stability in plasma without compromising data quality.
Highlights
The stability of new chemical entities (NCE’s) in plasma is a concern in the drug discovery area [1,2,3]
We investigated a cassette assay procedure combined with the direct high-performance liquid chromatography (HPLC)-MS/MS approach for an even higher-throughput screen-type assay to simultaneously measure drug stability of multiple drug candidates in several plasma types
The advancement of genomics techniques and high speed chemical synthesis results in large volume of routine samples created from pharmacokinetic (PK) and drug metabolism (DM) research in drug discovery
Summary
The stability of new chemical entities (NCE’s) in plasma is a concern in the drug discovery area [1,2,3]. Drug stability in plasma has been measured using tedious sample preparation procedures including sequential plasma extraction at each incubation time point [4]. This labor-intensive approach is not suitable for the evaluation of a large number of bioactive compounds, which are generated by medicinal chemists in the modern pharmaceutical industry. The tremendous influx of many potent new chemical entities (NCE’s) produced weekly in today’s pharmaceutical industry demands an even higher throughput procedure for plasma stability experiments
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