Systematic evaluation of the root cause of non‐linearity in liquid chromatography/tandem mass spectrometry bioanalytical assays and strategy to predict and extend the linear standard curve range

Abstract

In this work, the root cause of the non-linear behavior of the standard curve when using a SIL-IS was investigated and identified. Based on the findings, an improved multiple SRM channels approach was proposed and successfully applied to obtain a linear dynamic range of five orders of magnitude for one test compound. This approach may work particularly well for LC/MS/MS bioanalytical assay of dried blood spot (DBS) samples, for which a direct dilution is cumbersome.

Comparison of different linear calibration approaches for LC–MS bioanalysis

“…Thirdly, according to a recent research by Yuan et al [31], there exists a thresh-hold value specific to each type of mass spectrometer. As long as the absolute response (peak height) at the ULOQ is below this value, linearity should be good independent of the compounds tested.…”

“…Non-linearity (quadratic) response often exists in LC-MS bioanalysis because of saturation, formation of dimer/multimer/cluster ions, and cross signal contribution from an analyte to its internal standard, particularly when the concentration range is wide [18,31]. Since linear regression is usually favoured due to its robustness or owing to the preference of regulatory agencies (at least generally perceived as such) [8,[31][32][33], many would try to fit non-linear data linearly.…”

“…Since linear regression is usually favoured due to its robustness or owing to the preference of regulatory agencies (at least generally perceived as such) [8,[31][32][33], many would try to fit non-linear data linearly. To mimic this scenario, the following simulations were performed.…”

Nitrated fatty acids: synthesis and measurement

“…Determinations based on LC-MS data usually have a dynamic range of 3 orders of magnitude, which can be extended on the upper end using less sensitive multiple reaction monitoring (MRM) transitions [32]. The proposed standard curve spans over 5 orders of magnitude to accommodate quantifications using instruments with higher or lower sensitivity.…”

Increasing the linear dynamic range in LC–MS: is it valid to use a less abundant isotopologue?

“…This is particularly problematic in multi‐analyte methods where the desirable inter‐analyte concentration ranges may span several orders of magnitude. In modern LC–MS instruments, the reason for non‐linearity at higher concentrations is often due to detector saturation …”