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(within past 24 months)
No relevant financial relationship(s) to disclose.
Abstract
Introduction: For toxicological screening, methods not only need to be highly specific and sensitive but capable of identifying and quantifying a broad range of analytes in complex matrices. In this work a generic data independent acquisition (DIA) method was developed to detect and quantify a panel of drugs of abuse in human plasma using a high resolution accurate mass LC-MS/MS QTOF system. The method was designed as a generic data acquisition approach to extend the capability of LC-MS/MS analysis in routine clinical pathology for both targeted and untargeted data analysis.
Objectives: Develop a high resolution accurate mass LC-MS/MS QTOF system capable of generating targeted and untargeted data analysis for toxicological screening.
Methods: Human plasma (BioIVT) was prepared using a QuEChERS protocol modified for clinical applications (Dulaurent et al. 2016) and spiked with a drugs of abuse panel creating calibration curves from 5 to 200ng/mL. Samples were separated using a biphenyl column binary gradient method over 10.5 minutes and analysed by an LC-MS/MS Q-TOF. All data was acquired using external mass calibration. MS acquisition scan range was m/z 100-500. Nineteen consecutive MS/MS scans were created isolating 20 Da ion width starting from m/z 120-140 through to 480-500; measurement mass range m/z 40-500, each MS/MS scan lasting 25 msec. Collision energy scanning (0-30V) enabled detection of precursor and product ions in MS/MS scans.
Results: The DIA-MS/MS method developed on a Q-TOF platform was designed to capture precursor and product ion spectra for all ionizing analytes providing options for targeted quantitation at either the MS or MS/MS level and retrospective untargeted analysis. This approach differs from a conventional targeted approach such as high resolution MRM which is designed to be highly specific for a panel of analytes.
In this work, 41 drugs of abuse compounds were detected and quantified at the lowest calibration standard (5ng/mL) to an accuracy of 85-115% with linear and quadratic calibration curves up to 200 ng/mL for all targets using precursor DIA-MS/MS data (as each mass scan used a collision energy from 0-30V both precursor and product ion were present in the DIA-MS/MS data). As a generic method this DIA-MS/MS acquisition method requires no other modification for the analysis of other small molecule compound classes such as antidepressants, immunosuppressants, multi-panel analytes.
= Discovery stage.
= Translation stage.
= Clinically available.
