MestreNova Quick Guide
(for processing and analyzing 1H and 13C 1D spectra)
By Monika Ivancic and Paul White
Updated 7/19/2012
Opening data files:
Use ctrl-O or
to bring up a GUI that will help you navigate and open the data file.
For Varian data (Hermes, 500s, 600) navigate thru the folders until you find ‘fid’.
For Bruker AC data (Homer, Athena) just click on the file name that you saved it
on the spectrometer (on R:\ac300\homer or Athena).
Alternatively find your data in the Windows (or Mac) folder, click-hold and drag
onto the MNova icon. This opens MNova and your data in it.
A note about settings:
Paper size: Go to File -> Page setup. Change Page size to say Letter (8.5 X 11 inches)
(will avoid any issues you may have later when printing; check this every time you install
a new version)
Other settings may be manipulated by R-clicking on the spectrum background and going
to
Properties. Here you may change the background grid and tick marks, font sizes on peak
picks, integrals, and you may change your spectrum title, etc.
A note about cursor modes:
In MNova you use the cursor to process and manipulate your data. You get the cursor
into the different modes by clicking on particular icons OR by using keyboard shortcuts, which
are faster once you remember them. To get back to ‘normal’ or ‘base mode’ hit Esc.
Working up 1H 1D data:
MNova automatically FTs your data. If you would like to see your fid, you may do so
under the
icon.
Window function (mathematical function called ‘apodization’ applied to raw fid): click
the
icon to get a list of functions. Use a ‘matched filter’ for 1H 1Ds, which is an exponential
that is the inverse of the acquisition time (on Homer/Athena AQ = 2.7 sec; on Hermes at = 4
sec). Other functions may be used as well. To see what they do to your spectrum make sure the
‘interactive’ box is checked in the lower right of the GUI. (Mnova applies the apodization that
has been defined on the spectrometer, although not for Homer/Athena data.)
is useful in 1H 1Ds, because it adds digital resolution to the data (ie. you
Zero filling
can easier observe fine splitting). MNova automatically does one zero fill. You can find the
icon by clicking on the Fourier Transformation
options. In the GUI that appears,
you have the option of changing Spectrum size, if you want more zero fills (Original size is the
number of points in the raw data). However, don’t use more than two zero fills.
Phase correction: MNova does an automatic phase correction, but if you increase the
intensity of your spectrum (
or scroll up on middle mouse) you may notice it’s not perfect.
To get to Manual Phase correction, click on the
icon (under
instructions in the GUI. The shortcut for this mode is ‘Shift + P’.
options) and follow the
Baseline correction: this must be done for integrations to be correct. Click on the arrow
by
and then
to get an interactive GUI for applying the correction. A dark blue line
will show how the polynomial chosen will fit your baseline. Use either ‘Bernstein polynomial
Fit’ or ‘Polynomial Fit’ and you may change the polynomial to better fit your baseline. In some
cases, Whitaker baseline correction works well (ie. for very heterogeneous mixtures that may
have large aggregates), but be careful since it will remove broad lines.
Referencing: click on
to reference TMS or any other solvent peak (L shortcut).
The GUI that pops up allows you to annotate the peak with any text.
Peak picking: Click on the arrow by
threshold
to see the different options. The manual
option (K shortcut) is nice, because it allows you to select groups of peaks with
different thresholds. The peak by peak
option (ctrl-K) is needed if you have shoulder peaks
or ‘hidden peaks’ that were not selected in any of the automatic options.
Integrations: The arrow by
will show you the options, although the best is Manual
Integration
(I shortcut), which allows you to define the integration regions yourself. The
first integral that you define, will automatically be normalized to 1.0. To change this, R-click on
the integral that you want normalized and Edit Integral. In the GUI change the normalized value
and click on ‘Apply to All’ at bottom.
Multiplet analysis: The arrow by
shows that you may either do this automatically
on the entire spectrum or you may choose the multiplet regions yourself. The analysis works
better if you choose the multiplets by clicking on
(J shortcut) and selecting multiplets one
by one. When you select a multiplet, make sure the horizontal line that appears is below your
peaks that you want analyzed (or close to the baseline).
Manipulating integrations and multiplets: move the cursor over the integral or over the
multiplet definition box and R-click. You may manually Edit Multiplet features or delete certain
multiplets. You may change your integration normalization values from here also.
Working up 13C 1D data:
Open
the 13C 1D spectrum, and the FT is automatically displayed. Just like in the
1H 1D, you may view the FID, or just go ahead and check processing.
Window function or apodization: Under
use an exponential between 1Hz and 3Hz
(13C 1Ds are quite noisy, this will kill the noise in the latter part of the FID).
Zero filling: In general this is not needed, since in 13C 1D spectra we typically don’t look
for small splitting. MNova automatically ZFs data, you may go to
and choose
undo this (Spectrum size equals Original size for no ZFs), or just leave as is.
to
Phase correction: MNova does a good job here for most 13C 1Ds. If you’d like to correct
the phase (ie. if you see some dips below baseline), go to
and follow instructions.
to reference the solvent peak (L shortcut). The GUI that
Referencing: click on
pops up allows you to annotate the peak with any text.
Peak picking: Use the K shortcut or go to
and then
to define different
thresholds for different parts of the spectrum. If you would like a report for publication of these
peaks, click on
Report Peaks, which spits a peak list onto your spectrum. Copy Peaks
will allow you to copy and paste into a Word document. You may also go to View -> Tables ->
Peaks and check out what options you have here.
Manipulating your spectrum
Increasing and decreasing intensity is easily done with the middle mouse ‘wheel’.
Scrolling it up (away from you) increases the spectrum intensity, while scrolling it down
(towards you) decreases intensity. You may use
and
icons in the toolbar as well.
Click on
to change the cursor to zoom mode and select the region you want to zoom
in on. Z-shortcut works best: 1st Z is horizontal zoom, 2nd Z is vertical zoom, 3rd Z is in both
dimensions (hit Esc to get your cursor back to normal).
Zooming out may be done by clicking on
icon or using Shift-Z, which puts the
cursor in ‘zoom out’ mode. Click on the spectrum to zoom out.
The Full Spectrum icon
will get the entire spectrum back in horizontal (ppm) and
vertical (intensity) dimensions. The F shortcut will get the entire spectrum back in the horizontal
(ppm) dimension. While the H shortcut or
to the top of the page.
icon will fit the tallest peak in the visible region
Some like to see the entire spectrum, while zooming in on particular regions. Go to View
-> Full View, which brings up a small box with the whole spectrum and the zoomed in part
highlighted in blue. You may click and drag the blue region to different parts of your spectrum.
Helpful for getting to different parts quickly. You may also drag the Full View box around, to
check out how it can get “plugged in” to different parts of the window.
icon and then select the region of your spectrum that
Spectrum insert: E shortcut or
you want the insert of. You may manipulate the insert the same way you would the entire
spectrum (using the above shortcuts), just make sure the insert is selected.
Cutting parts of the spectrum: If you have large baseline regions, you may click on
icon (X shortcut), which puts your cursor in scissor mode and you may cut certain regions out of
view. The scale at the bottom will reflect the cut. Use the V shortcut to get the cursor into
‘restore’ mode and highlight the cut that you want restored.
Annotating your spectrum:
Look under ‘Annotate’ in the main menu (or go to
in the lower left corner) to see
what options you have (lines, arrows, rectangle, ellipse, polygon, text)
Adding text: Go to Annotate -> Text (T shortcut), click where you want to add text and
start typing. The size of the text box will be automatically determined as you type. Once
finished, you may change the size (make sure the text box is selected).
Note: the text box is attached to the peaks, so when you increase peak intensity it may go
off the page (this makes it so that you can annotate specific peaks, but not so great if it is general
info about your spectrum). Bug: If you copy’n’paste the text box from a previous spectrum, it
will be attached to the page (position on the page).
Change annotation features, such as font: Look at icons in lower left to get to different
annotation features. Here you can change the font, font size, super and subscript, etc.
Spectrum title: This automatically gets added in the upper left corner of your spectrum
(except for Bruker AC data). To change this text or to add line breaks (so it’s lower than the top
of the page), R-click on background and go to
Properties. Then under General click on
of the Title. In this GUI you can add line breaks at the beginning or change the text.
Absolute Referencing (using MestreNova 8.0 or higher version)
MestreNova now has a way to reference X-nucleus spectra based on a properly referenced 1H
nucleus spectrum acquired under identical lock conditions. This implementation works for
Bruker AC (Athena) data as well as data acquired with more modern spectrometers. For more
info on this see our NMR facility web site -> User Guides -> Other Guides and Resources ->
Absolute Referencing in NMR: The Unified Scale (a *.pdf document).
To implement this in MNova:
Open your 1H spectrum and process as described above, making sure to properly
reference to TMS or solvent
.
In the same document, open your X-nucleus spectrum and work it up similarly to the 13C
1D instructions on p.3. To reference, click on
and the GUI that pops up lists the nuclei of
the spectra in this document. Make sure that both the 1H and X-nucleus spectra are checked and
click OK. The scale (and peak picks) will shift to a properly referenced scale; you may now
peak pick and do your analysis.
Note: Open up a new document to use absolute referencing for a different compound.
Generating Stack Plots
(by Paul White)
For Arrays
1) Open the array of data collected (from the Varians…soon to be Brukers). The data should be
present already as a stack plot which is great if your phasing is the same for each spectrum but it
usually isn’t due to slight changes in temperature over the period of acquisitions.
2) First, I look at the active spectrum by clicking
drop-down menu on the left and selecting
“Active Spectrum”. Proceed to phase this spectrum which will be applied to all the other spectra
in the array. This will provide a rough phasing so that the subsequent phasing of the individual
spectra will be easier.
3) In order to phase each slice of the array, you must break the array into the individual spectra.
Click “Stack > Extract All Spectra” to perform this operation. Give MNova time as this is an
intensive process, especially if you have >100 slices. After the extraction, you can delete the old
array page as it may distract you later on.
4) Perform the proper phasing for each spectrum.
5) To recombine them into a stack plot, click over to the pages section and hit Ctrl-A (Cmd-A for
Mac) to select all the spectra. Next, hit “Stack > Stack Spectra” and a next page containing your
stacked spectrum should appear.
6) To apply a baseline correction to each spectrum in the array select the page and hit
“Processing > Baseline > Baseline Correction”.
7) Finally, you can integrate each spectrum in the array by selecting one spectrum (preferably
with both reactants and starting material) and use
on the left menu to select “Active
Spectrum”. Integrate the regions of choice and onced finished, you can export these integrals to
a .txt file by clicking “File > Save As”. Under Format you see you can change it to “MestraNova
Integral Regions” or “Script: Integral Table Series” among others containing the word
“integration”. I primarily use Script: Integral Table Series and I open the text file in Excel. For
understanding the text file, see the end of the page.
For Non-Arrayed Kinetics
1) Begin by phasing and referencing each spectrum.
2) Follow directions from step 5-7 above.
Example (red text is my notes):
Proton_01 – this will be your title of your FID
Downfield ppm value Upfield ppm value
Norm. Int.
Proton 1 6.02503
6.14733
Proton 2 3.66153
3.84782
5.68089
5.92122
5.05659
5.17889
Absolute Integration
1
8729.96218
3.10418
27099.39158
0.67411
5884.94555
0.6703
5851.71054