Utilizing Internal Standard Responses to Assess Risk on Reporting Bioanalytical Results from Hemolyzed Samples.

Bioanalytical analysis of toxicokinetic and pharmacokinetic samples is an integral part of small molecule drugs development and liquid chromatography-tandem mass spectrometry (LC-MS/MS) has been the technique of choice. One important consideration is the matrix effect, in which ionization of the analytes of interest is affected by the presence of co-eluting interfering components present in the sample matrix. Hemolysis, which results in additional endogenous components being released from the lysed red blood cells, may cause additional matrix interferences. The effects of the degree of hemolysis on the accuracy and precision of the method and the reported sample concentrations from hemolyzed study samples have drawn increasing attention in recent years, especially in cases where the sample concentrations are critical for pharmacokinetic calculation. Currently, there is no established procedure to objectively assess the risk of reporting potentially inaccurate bioanalytical results from hemolyzed study samples. In this work, we evaluated the effect of different degrees of hemolysis on the internal standard peak area, accuracy, and precision of the analyses of BMS-906024 and its metabolite, BMS-911557, in human plasma by LC-MS/MS. In addition, we proposed the strategy of using the peak area of the stable isotope-labeled internal standard (SIL-IS) from the LC-MS/MS measurement as the surrogate marker for risk assessment. Samples with peak areas outside of the pre-defined acceptance criteria, e.g., less than 50% or more than 150% of the average IS response in study samples, plasma standards, and QC samples when SIL-IS is used, are flagged out for further investigation.