Abstract
Non-targeted screening (NTS) with reversed phase liquid chromatography electrospray ionization high resolution mass spectrometry (LC/ESI/HRMS) is increasingly employed as an alternative to targeted analysis; however, it is not possible to quantify all compounds found in a sample with analytical standards. As an alternative, semi-quantification strategies are, or at least should be, used to estimate the concentrations of the unknown compounds before final decision making. All steps in the analytical chain, from sample preparation to ionization conditions and data processing can influence the signals obtained, and thus the estimated concentrations. Therefore, each step needs to be considered carefully. Generally, less is more when it comes to choosing sample preparation as well as chromatographic and ionization conditions in NTS. By combining the positive and negative ionization mode, the performance of NTS can be improved, since different compounds ionize better in one or the other mode. Furthermore, NTS gives opportunities for retrospective analysis. In this tutorial, strategies for semi-quantification are described, sources potentially decreasing the signals are identified and possibilities to improve NTS are discussed. Additionally, examples of retrospective analysis are presented. Finally, we present a checklist for carrying out semi-quantitative NTS.
Highlights
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We suggest doing as little sample preparation as possible, both to prevent losing compounds in the process and to avoid poor recoveries of compounds
Best results are achieved when using different mobile phases for ESI+ and ESI-: suggested mobile phases are presented in Section 2.2 and in Table 3, together with a generic gradient
Summary
A general workflow for non-targeted screening (NTS), as described by Hollender et al [1], includes representative sampling followed by enrichment suitable for the sample matrix. The step is peak detection and grouping of peaks related to the same molecular structure, and comparison of sample peaks with peaks from compounds present in blanks [1,2,3] This step can often be semi-automated, using either the vendor, third-party or open-source software (e.g., Thermo ScientificTM Compound DiscovererTM software, [4] envipy [5] from EAWAG or MZmine [6]). A measurement that is closely related to the ionization efficiency of a compound is the response factor (RF), the ratio of the detected peak area and the concentration of the compound (Equation (1)) This ratio is important in some of the semi-quantification strategies presented below. Pieke et al [17] and Espinosa et al [18]
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