The power of energy‐resolved tandem mass spectrometry experiments for resolution of isomers: the case of drug plasma stability investigation of multidrug resistance inhibitors

Abstract

RATIONALE: A series of drug plasma stability experiments were carried out to evaluate the bioavailability of three multidrug resistance inhibitors. The studied compounds are positional isomers; therefore, a chromatographic separation or taking advantage of specific collisionally activated decomposition pathways, obtained by tandem mass spectrometry (MS/MS) experiments, is necessary in order to resolve them. METHODS: A method was developed for quantitative determination of the analytes in plasma using liquid chromatography (LC) coupled with a triple quadrupole mass spectrometer operating in MS/MS mode. Different collisional approaches were employed based on the potentiality of a triple quadrupole system. Aside from the classical product ion spectroscopy, energy-resolved MS/MS experiments and a post-processing mathematical algorithm tool (LEDA) were used to distinguish among different kinds of inhibitors present in the sample batch. RESULTS: The developed LC/MS/MS method showed precision between 1.8-7.9%, accuracy ranging from 92.8 to 99.9% and limit of detection (LOD) values in the range 1.0-1.4 ng mL À1 for all the analytes. The evaluation of matrix effects demonstrated that the sample preparation procedure did not affect the ionization efficiency or recovery (matrix effects and recovery larger than 88%). Finally, the LEDA tool was able to differentiate among the isomers, ensuring their proper monitoring. CONCLUSIONS: The proposed LC/MS/MS method was suitable for evaluating the stability of the analytes in plasma samples, although small concentration variations occurred. Furthermore, the investigation on the energetics of fragmentation pathways allowed the better product ions and optimal abundance ratios to be selected for LEDA application into a multi-component analysis.

Resolution of co-eluting isomers of anti-inflammatory drugs conjugated to carbonic anhydrase inhibitors from plasma in liquid chromatography by energy-resolved tandem mass spectrometry

“…In order to evidence possible different intrinsic molecular stability of the analytes, the study of fragment ion yields present at different CEs (breakdown curves) was performed. To achieve valid and well reproducible fragmentation pattern, the collision gas pressure was critically evaluated, based on the results obtained and discussed in a previous paper 26 .…”

“…The Pi scan spectra were acquired in the m/z range from 50 to 650, scan time 600 ms; argon was used as collisional gas and the collision energy (CE) was increased stepwise in the range 5–40 V. A series of energy resolved tandem mass spectrometry (ERMS) experiments were performed to study the fragmentation of molecular species of each analyte and build its breakdown curves 26 , 27 . The ERMS experiments were carried out by introducing working solution 1 of each analyte, via syringe pump at 10 µL min −1 ; the protonated molecule was isolated and the abundance of Pis were monitored.…”

“…This algorithm (linear equations of deconvolution analysis [LEDA]) consists in the application of matrix of linear regression equations to different experimental data. In our case, the experimental data used were the abundance ratios of product vs. precursor ions selected during the MS/MS method set-up 26 , 27 . In this way, it was possible to resolve the MS/MS spectra assigning the correct signal to the isomer also for chromatographically unresolved peaks.…”

Synthesis and analytical characterization of new thiazol-2-(3H)-ones as human neutrophil elastase (HNE) inhibitors

“…The breakdown curves data were obtained by introducing 1.0 µg/mL of each analyte via syringe pump at 10 µL/min; the protonated molecule was isolated and the abundance of product ions were monitored. The values of the relative intensities of MS/MS spectra used to build the breakdown curves were the mean of 50 scans for each collision energy and the collision gas pressure was critically evaluated in order to achieve valid and reproducible fragmentation patterns on the basis of the results obtained [29]. …”

Determination of coeluted isomers in wine samples by application of MS/MS deconvolution analysis

“…In our case, the experimental data used were the abundance ratios of product versus precursor ions selected during the MS/MS method set-up on the triple quadrupole MS system. [18][19][20] In this way, it was possible to resolve the MS/MS spectra and assign the correct signal to the isomer, even given chromatographically unresolved peaks. MilliQ water 18 MΩ cm was obtained using the Millipore Simplicity Water Purification System (Milan, Italy).…”