Combined approach for high‐throughput preparation and analysis of plasma samples from exposure studies

Abstract

In drug discovery today, drug exposure is determined in preclinical efficacy and safety studies and drug effects are related to measured concentrations rather than to the administered dose. This leads to a strong increase in the number of bioanalytical samples, demanding the development of higher throughput methods to cope with the increased workload. Here, a combined approach is described for the high-throughput preparation and liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis of drug levels in plasma samples from the preclinical efficacy and safety studies, i.e. exposure studies. Appropriate pharmacokinetic (PK) compartmental models were fitted to data from PK screening studies in the rat, which were subsequently used to simulate the expected plasma concentrations of the respective exposure studies. Information on the estimated drug concentrations was used to dilute the samples to appropriate concentration levels. A Tecan Genesis RSP liquid handling system was utilized to perform automated plasma sample preparation including serial dilution of standard solutions, dilution of plasma samples, addition of internal standard solution and precipitation with acetonitrile. This robotic sample preparation process permitted two studies of 1-96 samples each to be run simultaneously. To ensure the performance of this method the accuracy and precision for diazepam were examined. Two novel drugs were used to illustrate the suggested approach. In conclusion, our method for sample preparation of exposure samples, based on the combined use of PK simulations, a liquid handling system and a fast LC/MS/MS method, increased the throughput more than three times and minimized the errors, while maintaining the required accuracy and precision.

Amide Hydrolysis of a Novel Chemical Series of Microsomal Prostaglandin E Synthase-1 Inhibitors Induces Kidney Toxicity in the Rat

“…Liquid chromatography mass spectrometry (LC-MS) analysis for quantifications was performed with a Micromass Quattro Micro triple quadrupole (Waters Corporation, Milford, MA), as previously described elsewhere (Briem et al, 2007) with the exception of analysis of the M1 metabolite. Total concentration of M1 in plasma was determined by LC-MS/MS using electrospray in the negative ionization mode.…”

“…All TK values are presented as mean values from at least two animals. Low-dose PK studies for dose setting in toxicity studies were performed as described elsewhere (Briem et al, 2007).…”

From Known Knowns to Known Unknowns: Predicting in Vivo Drug Metabolites

“…On some occasions, more comprehensive sample preparation methods, such as solid-phase extraction or liquid extraction, are used in addition to protein precipitation. All the main techniques used, namely protein precipitation, solid-phase extraction and liquid-liquid extraction, are now also available in well-plate format for higher throughput and can be easily automated using liquid-handling systems [84,85]. Most HPLC autosamplers are also capable of injecting directly from the well plate, without the need for sample transfer into traditional autosampler vials.…”

Full utilization of a mass spectrometer using on‐demand sharing with multiple LC units

“…Various implementations of staggered parallel systems have been reported following the first implementation of this system in which two LCs were used . The potential of staggered parallel approach for high throughput analyses has been demonstrated for a number of applications, including absorption, distribution, metabolism and excretion screening, pharmacokinetic studies, high‐throughput enzymatic assays and, most recently, proteomics analysis using capillary LCs …”