MassFinder Analysing your own data

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MassFinder 4: Tutorial

This is the eighth lesson of the MassFinder 4 tutorial. You will learn how to use your own data files, how to employ the unique retention index feature and how to configure MassFinder to perfectly suit your requirements.

If you are unsure whether your data files are compatible with MassFinder, please read data file compatibility.

Contents

Step 1: Open dialog

Start MassFinder and call the open dialog. Select the appropriate data file format for your system and use the explorer tree on the left side to navigate to your data file folder.

Image:Tutorial-ButtonAddFavorite.pngYou may add this folder to your favorite folders by clicking on the add folder button. Favorite folders make accessing several different data file locations easy. MassFinder will remember your last data file folder automatically anyway.

Please note that MassFinder files (*.mfg) will be displayed regardless which data file format is selected. This ensures that you can both access your native data files as well as your processed MassFinder files at the same time.

Please be assured that MassFinder will never modify a native data file. MassFinder can only write to *.mfg files and will not overwrite your native data files.

Step 2: Open a data file

Please now open one of your native data files. Navigate and analyse the GC/MS in order to check whether MassFinder imported your files correctly and both the chromatographic profile as well as the mass spectra are authentic and identical to your native mass spectrometer's acquisition software.

Hint: Since we have not yet set-up the retention index table for your system, MassFinder will not find proper library hits if the retention index feature is activated. If you want MassFinder to return sensible library hits, you need to deactivate MassFinder's retention index feature by clicking on the 'RI1' in the status bar or by pressing the key 'r'. The RI1 icon in the status bar should be greyed now.

Step 3: Measuring an alkane pattern

In order to automatically calculate correct retention indices for your data files, MassFinder needs to know the retention times of the corresponding n-alkanes. Please read our Retention index guide to learn about the concept of retention indices.

In summary, please note the following aspects:

  • You probably already have a default method and standard temperature program for your samples. If not, you need to define a default method now.
  • Determine the relevant retention time range of your target compounds and estimate the necessary range of n-alkanes. For monoterpene and sesquiterpene analytes C8 - C20 is a sensible choice; highly polar, oxygenated sesquiterpenes and diterpenes might require an upper limit of C26. Try to avoid higher n-alkanes than elute in reasonable time with your default method.
  • Purchase a commercial standard mixture of n-alkanes covering your estimated range. (Examples: Fluka #04070 with C8-C20 for approx. Euro 23,00 or Restek #31214 with C10-C25 for approx. Euro 38,00).
  • Measure this standard alkane mixture using your default method.
Image:RetentionIndexGuide-AlkanePattern.png

Naturally, MassFinder supports an unlimited number of methods and temperature programs. However, it is reasonable and practical to limit the number of methods to the necessary minimum.

Please note that MassFinder's retention indices are measured on a DB-1 stationary phase (100% polydimethylsiloxane) and are not compatible with other stationary phases. If you want to use MassFinder's library and retention indices you need to use the same stationary phase. Using other stationary phases will lead to deviations between your and our retention indices. Naturally, you can update the library values, create a new library or convert your existing proprietary library to be used with MassFinder, so that you become independent from our choice of stationary phase.

Step 4: Opening the alkane data file

Please open the data file with the n-alkane pattern.

Please note that MassFinder does not display any retention indices in the mass spectra's upper right corner, but only the retention times. This is because MassFinder does not yet know the retention table belonging to your GC/MS data.

Please mark all alkane peaks. It does not matter whether you mark them as unknown or identified peaks. If the peak threshold is appropriately adjusted, you may even simply click the auto-report button and MassFinder will mark all peaks as unknown (hollow blue arrow). Please make sure that all and only alkane peaks are marked. Manually unmark all signals that do not belong to the alkane pattern if necessary.

Step 5: Entering the retention index table

Image:Tutorial-ButtonOptions.png Please open the options dialog and select the retention indices tab sheet. Please note that a freshly installed MassFinder version only has one retention table called "N/A" with no retention times stored at all.

Image:Tutorial-ButtonEditName.png Please click on the edit name button and enter a short description for the new retention table, e.g. corresponding with your method name. If you use multiple stationary phase, we suggest to include an abbreviation of your stationary phase in the name of the retention table. Many customers include a reference to the date of the alkane pattern. Please note that this name will be shown in the open dialog selection box. You may edit this name anytime without problems. Example: "DB1-081012" for an alkane pattern measured on 12th October 2008 on a DB1 column with your default temperature program.

Please click the accept times of marked peaks button to transfer the retention times of your currently marked peaks into the curent retention table. The Assign retention table dialog allows to enter the number of the first alkane, i.e. the identity of the first marked peaks. For example, if you have measured the alkanes C8 to C20 and marked all alkane peaks, you would enter "8" as first alkane. Click OK to accept this setting and to return to the options dialog. Please note that MassFinder has entered all retention times in the appropriate cells of the retention table. Please observe how MassFinder re-colors the lines from grey to black text for the lines that create a proper retention table, i.e. increasing retention times without gaps in between.

Please click the recalculate current GC/MS button. MassFinder will now convert all retention times of your current GC/MS run into retention indices. Each scan will now have the correct retention index associated. This enables you to check the validity of your retention indices and you will easily be able to check whether everything went fine with the retention table setup. You should execute this recalculation each time you update your alkane pattern. Please do not recalculate any other GC/MS runs besides the alkane pattern used to setup the retention table.

Now close the options dialog and navigate to the alkane peaks. Check whether each alkane peak now shows the correct retention index, e.g. C8 with RI 800 up to C20 with RI 2000. Please note that GC/MS scans earlier than the first alkane peak or later than the last alkane peak still do not show any RI value. You need to use an alkane pattern range large enough to cover your relevant time range. There is no problem to let the beginning and end of runs uncovered if you do not need this retention time ranges.

Step 6: Opening a sample data file

If you want to use MassFinder's Terpenoids Library, then please open any data file containing common terpene peaks whose identity you know for sure. Please check that the options dialog displays the freshly entered retention table in the appropriate combo box (lower right corner).

Navigate to peaks you know and compare their calculated retention indices with the library values. Check whether MassFinder displays the correct identification as one of the first library hits.

Please remember to switch on the RI feature by clicking on the RI1 icon in the status bar or by pressing the 'r' key. The RI1 icon should be black.

If you ever happen to open a native data file with the wrong retention table, just re-open it with the correct retention table.

Step 7: Practical considerations of using retention indices

Measuring alkane patterns

Modern gas chromatographs will reproduce retention times over several weeks. Please watch your retention indices of common compounds and be alerted by sudden deviations.

Please re-measure the alkane pattern and update the retention table each time whenever you notice unusual deviations and whenever you modify your gas chromatography system, i.e. by changing or shortening the column, adjusting the carrier gas pressure, installing pre-columns and so on. We recommend to re-measure your alkane pattern at least once a month.

Storing retention indices

MassFinder calculates retention indices for each scan of your native GC/MS run by employing the retention table selected at the moment of opening. When saving a GC/MS run to a MassFinder *.mfg file, the retention indices are stored in the *.mfg file. When opening a *.mfg file, MassFinder reads the retention indices from the file and does not apply the open dialog's selected retention table.

The advantage of this method is that *.mfg files contain retention indices forever, no matter whether you later re-measure or loose the alkane pattern. Just ensure to use the correct and up-to-date retention table at the moment of opening a native data file. If you open all your data files soon after acquisition using the correct retention table and save them as *.mfg files, then you do not need to keep track of your changing retention tables. There is no need to remember when and how often you updated your retention table, if you open and save all acquired files.

Step 8: Setting auto-report thresholds

The auto-report feature (yellow flash button) uses a peak rejection threshold based on the integral area, i.e. MassFinder detect peaks, integrates them and then decides whether to keep them by comparing the calculated integral with the area threshold.

Please open one typical native data file and navigate to a small peak that just represents the lower limit of peaks you want MassFinder to include in the auto-report, i.e. all peaks with an area equal or larger than this peak will be included and all peaks smaller than this peak will be excluded.

Now open the options dialog, select the configuration tab sheet and locate the auto-report panel. Please click the calculator button beside the area threshold (peak reject) edit box. MassFinder will automatically enter the current peak's area into the area threshold field. Alternatively, you could look up the integral value in the result table and enter any value of your choice manually.

Please note that you may modify this threshold as often as you like. If you use the same type of data files and fairly similar samples, then you will not need to change this value very often.

Please note that MassFinder distinguishes between GC/MS and GC data. You may enter different thresholds for these types of data.

Further information

This was the last lesson of the MassFinder 4 tutorial. You are now able to analyse your data and to use most of MassFinder's features.

The MassFinder Manual will explain all options and features of each dialog in detail. We recommend to continue with reading options and configuration and printing. Again, if any problems arise, please do not hesitate to contact us!

Please note that the options dialog features two hyperlinks in the lower left corner which allow to access this online manual and to look for updates of MassFinder.


Please feel free to email us your feedback regarding this tutorial. Also we gladly accept suggestions for further tutorial lessons or any other details you missed in this tutorial.

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