Detect and identify peaks
One of the most common and basic workflows in chromatography is the peak detection and subsequent identification of the components. Here, we exemplify a workflow for the detection and identification process for gas chromatographic data acquired with a low resolution mass spectrometer. On many occasions also quantitative or semi-quantitative information is demanded, which is mostly attained by peak integration. Hence, it is included in the workflow here as well. Note, that in OpenChrom the peak detection and integration are kept as two separate processing steps. This allows more flexibility for recombining algorithms.
The workflow of peak detection, integration and identification can be done manually for every chromatogram or batch-wise on e.g. hundreds of measurements with the integrated batch processor.
1.When you start up OpenChrom and select the „MSD perspective“ your first step is to select a chromatogram to process, e.g. via the Chromatogram File Explorer (MSD) (A). Note, that folders are accessed by clicking the triangles and the actual measurement by double-clicking the file name. Find out about other options to open data files here.
2. The measurement selected via the Chromatogram File Explorer (MSD) (B) is displayed in the Chromatogram Overview (MSD) (C) and, once double-clicked, in the Chromatogram Editor. The default mass spectrum displayed in the view Scan (Nominal) (D) corresponds to the first scan of the chromatogram. The mass spectra can also be displayed as numerical values (export e.g. to excel simply by marking and copying) (E).
3. Before you start processing the chromatograms you are recommended to check the settings for the peak detection, integration and identification. You can access these via the Preferences dialog located under the menu tab „Window“. For instance, when a search of the NIST library is intended, the NIST MS Search software and the OpenChrom NIST-DB connector have to be installed and the path to the „nistms$.exe“ in the MSSEARCH folder has to be set. Read here how to install plug-ins.
5. After you pressed automated "First Derivative" peak detection the perspective (set of views) changes to "Chromatogram Peaks" (if you allowed perspective changes). A peak list will be displayed (G) and it can be noticed that the peak areas are still zero. This is the case because detection and integration are kept as separate processing steps. In the Chromatogram Editor the detected peaks are marked with small grey triangles/arrows above the apexes (H). The Peak (MSD) view (I) shows a close-up of the peak selected in the peak list. The view Peak Mass Spectrum displays the mass spectrum extracted for the selected peak (J). In case of the "First Derivative" and the Manual Peak Detector it corresponds to the mass spectrum at the scan of detected maximum and in the case the AMDIS connector was used the deconvoluted mass spectrum is displayed.
6. Note, that if you zoom into the chromatogram in the Chromatogram Editor, the peak list is also updated, only displaying the details of the peaks within the selected retention time window. A peak selected in the peak list (K) is highlighted in the Chromatogram Editor (darker shaded red) (L). Also the scans the peak consists of are displayed. A peak can also be directly selected in the Chromatogram Editor by pressing Ctrl + double-click left mouse button. If you are interested in details on the peak model calculations you can open the view "Peak Values List" (M). Note, you have to be aware that if you perform processing steps while you have zoomed into the chromatogram, the processing will only affect the current retention time window.
8. If you are interested in quantitative measures for the peaks you can get the peak areas or, alternatively, peak heights ("PeakMax Integrator") calculated. For calculation of peak areas currently only the widely used integration according to the trapezoidal rule is available ("Integrator Trapezoid"). Note, that the "Chromatogram Integrator" calculates the total peak area of the chromatogram - if the background has been previously detected with one of the available background detectors, it is distinguished between total background and peak area. The "Sumarea Chromatogram Integrator" does the respective calculation for each m/z separately.
9. The peak areas are now displayed in the Peak List (MSD) together with numerous details including peak width and signal-to-noise (S/N) ratio (settings for the S/N calculation can be adjusted via the preferences). Via the peak list entries can be deleted or copied to clipboard for direct export to e.g. Excel (N). You can select one or multiple entries and delete/copy by right mouse-click. If further peaks are detected via manual detection they are added automatically to the list.
10. If you have the NIST Mass Spectral Library software installed (not older than NIST 08) you can install our NIST-DB connector via the marketplace and perform peak identification one by one or batchwise for all detected peaks. Note, that if you want to identify any signal in your chromatogram which has not been detected as a peak, e.g. components found in shoulders and revealed after background subtraction, you can use the option "Identify Selected Mass Spectrum".
11. After you trigger off peak identification the process may take a while depending on the amount of peaks detected in your current chromatogram. To review the results by default the perspective may change to "Chromatogram Peaks" (if you allowed perspective changes and this perspective is not already opened), but in case you need to open the perspective manually you can use the perspective switcher.
12. The identification results are listed in the table Peak Targets (MSD) (O). The results are displayed for each peak as you go through the Peak List (MSD). The number of results depend on the constraining parameters set by the user in the preferences, that is maximum number of results and threshold for match quality. Additionally the parameters directly adjusted in the NIST software apply. Unfortunately, the results exportable from NIST Mass Spectral Library software do not include the respective mass spectra and structural formulas. But OpenChrom enables saving and loading the mass spectral information saved in data files used by the NIST and AMDIS software (*.msp and *.msl files).