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Integration

 

NUTS Help

Integration Overview

NUTS includes a manual integration subroutine and an automatic integration function. This Overview is a description of the former. It is recommended that users read through this section, as the manual tools are often useful to supplement the automatic method. A summary of subcommands active in the integration subroutine is given below. Show me how to integrate.

The currently displayed region can be integrated with the menu selection Process / Integrate, with the ID command from the NUTS base level or with the Zoom subcommand I. After entering the integrate subroutine, the displayed region will have the standard NMR integration line. The first time the integration routine is entered, the integral extends over the entire spectrum and is automatically scaled such that the integral will be full scale at the end of the display. Any changes which are made in scaling or by defining sub-integrals are retained when the integration routine is exited and re-entered. On entering the integration routine, the menu choices change to display commands which are active. All commands can be accessed using the menus, with the exception of those which involve using the mouse. The commands are also available as single-letter keyboard commands.

Adjusting the integral scale — The scroll bar on the left can be used to adjust the vertical scale of the displayed integral. If the integral display goes off the top of the display, it will wrap around to the bottom of the display.  Note that changing the vertical scale of the integral trace does not change the integral values.

While in the ID routine, the Z subcommand brings up another scroll bar on the left to adjust the beginning level of the integral. A single <Enter> exits this offset adjustment routine.

Plotting can be done from within the integration routine (P subcommand, or File/Plot menu selection).  Whether plotted from within the integration routine, or from the NUTS base level, the plotted spectrum will have the integral plotted on it as it appears on the screen.

The ID routine is exited with the <Enter> key.

Defining separate sub-integrals — Certain of the integral routine commands are initiated by clicking the left mouse button to display a vertical cursor. For example, the integral trace can be broken into sub-integral regions of interest using this approach. Click the left mouse button once to produce the vertical cursor, click a second time at the beginning of the region to be integrated, and click a third time at the end of the region of interest to complete the sub-integral. Repeat as needed. Once a single sub-integral has been defined, the integral is only displayed for defined sub-integrals. To toggle between this display and the entire integral, type F. The sub-integrals are not lost on exiting the integration subroutine.

Setting integral values — The value of a chosen sub-integral can be set to a user-defined value. Place the cursor on one of the sub-integrals (by clicking once with the left mouse button) and type V, which brings up a dialog box in which you can set the value of the selected sub-integral. Once a sub-integral value has been defined, each sub-integral is labeled with its value. The integral values remain unchanged until a sub-integral is explicitly set to a new value, regardless of changes in the vertical scale of either the integral trace or the spectrum. Setting the value of a sub-integral to zero removes all labels.

The integral labels can be displayed vertically, which reduces overlap of the labels.  This can be set in the nuts.ini file, or while in the integration subroutine.  Integration subcommand V displays the labels vertically, subcommand H displays the labels horizontally.  Note that the V subcommand is also used to set the value of a sub-integral, but that command is only invoked when a vertical cursor is displayed and overlaps the chosen sub-integral, so there is no conflict.

By default, the label appears just below the end of each sub-integral trace. The labels can be moved to the top of the screen by typing M. This is a toggle command, so typing it again moves the labels back to below the end of the trace.

The integral font can be set with FI or by choosing Set Fonts from the Edit menu. (The user must exit the Integration subroutine to change the font.) Sometimes sub-integrals are so close together that the labels overlap, making them hard to read. To remedy this, the individual labels can be moved up or down slightly. Three positions are defined and are chosen with the keys 1, 2 and 3, as follows. Place the cursor on the sub-integrals whose label is to be moved (by clicking the left mouse button once). Typing 1 moves the label up slightly from its initial position. Typing 3 moves the label down slightly from its initial position. Typing 2 returns the label to its initial default position.  Show me how to adjust label positions.

Removing sub-integrals — Single sub-integrals can be deleted by placing the cursor on a sub-integral (by clicking once with the left mouse button) and typing D. At any time in the integral subroutine, typing C will clear the set of sub-integrals and display the entire integral.  Subcommand L will delete the last-created sub-integral.

Saving integrals — The positions of all sub-integrals and their values are saved with the spectrum, so that when the spectrum is recalled at a later time, these integrals can be displayed without having to be redefined manually. This information is saved in a file “tailer”, meaning it is appended to the end of the file. See the description of file tailers for more information.  An example of the integration information contained in the tailer is:

INTEGRALS
REL_VALUE = 3.755704e-006
START_PT  START_PPM END_PT    END_PPM         VALUE   LABEL POSITION
   809     7.560      951      6.636         5.593325    1
  1540     2.801     1623      2.261         2.000000    1
  1722     1.616     1767      1.323         1.923050    1
  1769     1.310     1848      0.796         3.017987    1
END_INTEGRALS

The integral values listed in the VALUE column are the result of scaling all integrals by setting the second integral to 2.  Absolute comparison of integral values between different spectra is possible using the “REL_VALUE” parameter, which relates the absolute area to the chosen scaled value.

Creating an integral list — A list of all currently defined sub-integrals and their values can be placed into the clipboard for pasting into a document, for editing, printing or transfer to another application, such as a spreadsheet.  The list looks like:

NUMBER          FROM           TO        VALUE
   1          7.56 PPM      6.64 PPM      5.59
   2          2.81 PPM      2.27 PPM      2.00
   3          1.62 PPM      1.33 PPM      1.92
   4          1.32 PPM      0.80 PPM      3.02

This is accomplished with the Integration sub-command T. All sub-integrals are included in the list, whether or not they are currently visible on the screen. The T command orders the sub-integral regions from largest to smallest chemical shift, so the list comes out in order, regardless of the order in which the sub-integrals were defined. Once T has been typed, the list can be pasted into the Windows Notepad or any word processing program or text editor.  From the NUTS base program level, after exiting Integration, the IL command is another way to place the list of integrals into the clipboard.  Note that if changes are made to the integrals, the information in the clipboard is not automatically updated – it is necessary to do this manually.  Any text contents of the clipboard can be displayed on the NUTS screen by typing Ctrl-B (or CB), which is a toggle, so typing it again will turn off display of the text.  This can also be done from the View menu.

The integral list can be created using either spaces or tabs to separate the columns.  With space-separated columns, when a fixed size font is used, the columns line up. (Best for pasting into Notepad).  Tab-separated columns are best for pasting into a spreadsheet program.  The choice is determined by a setting in the nuts.ini file, and can be toggled between the 2 options with the TB command.

Flattening the integral — If the integral is not “flat” enough, the region must have its baseline adjusted. NUTS provides several baseline adjustment options. Type <Enter> to leave the Integration routine, correct the baseline with one of the options described in the section on baseline correction. One more baseline correction option is available within the Integration routine, which lets the user correct DC offset and tilt while viewing the effect on the integral trace in real time. From within the integration routine, typing B enters this baseline correction routine. Press and hold the left mouse button. Moving the mouse left and right performs a zero-order correction (DC level). The same operation with the right mouse button performs a first-order (tilt) correction. Type <Enter> to keep the correction or Q to quit, both of which exit the baseline correction routine to Integration. The correction is performed only on the currently displayed region. This function applies a correction to the spectrum, rather than just adjusting the appearance of the integral trace. Because the ends of the region are altered, this operation may have an effect on adjacent regions. Show me baseline correction within Integration.

The easiest way to integrate an entire spectrum is to integrate expanded regions of the spectrum separately, in order to be able to see spectral details. Expand a region using the Zoom routine, enter the integrate routine and define the sub-integrals. Then either pan to another spectral region using the scroll bar at the bottom of the spectrum or exit the integration subroutine (with <Enter>) and select another expanded region. On re-entering the integration subroutine, the previously defined sub-integrals are still there and defining additional sub-integrals adds to those previously defined. After all sections are integrated, plot limits can be selected, the integrals displayed (using Ctrl-I) and the fully integrated spectrum can be plotted.

N.B. If any of the options which involve baseline correction of only the currently displayed region are used, other regions of the spectrum are altered, and their integral values will change. It is preferable to baseline correct the entire spectrum with FB before beginning integration.

The alternative is to integrate each section of the spectrum separately, copying the integral list to the Notepad for each section, and using the Notepad to build a complete integral list of the entire spectrum. While in the integration routine, copy the current region’s integral information to the clipboard (T) and paste into the Notepad. Then clear the sub-integrals (C) before moving on to the next region. This is because whenever T is typed, all sub-integral information is copied to the clipboard, not just those in the currently displayed region. If baseline correction is applied to the current region, it will change the baseline (and therefore the integral values) in other parts of the spectrum. Clearing sub-integrals does not affect the integral scaling.

Subcommands:

Left Mouse Button Click Displays a vertical cursor. Used to define sub-integrals and to select a sub-integral.
V When typed while vertical cursor is displayed, brings up a dialog box allowing the user to sets the value of the sub-integral at the cursor location. If the value is set to zero, the integral values are not displayed.
C Clears the sub-integrals, displaying one integral line for the entire spectrum.
D Deletes the selected sub-integral in the sub-integral list at the cursor location.
L  Deletes the last created sub-integral.
Q Removes the cursor and aborts a sub-integral selection process.
T Transfers the current sub-integral list to the clipboard for pasting into Notepad or other document.
R Read integral information from file “tailer”. The integral information must have previously been saved with the file (see TA command).
M Toggles location of the printed sub-integral label among the end of the integral line, the top of the page and the bottom of the page. Applies to both screen display and plot.
1, 2 & 3 Moves location of the printed sub-integral label at the current cursor location to one of 3 vertical positions. Used to remove overlap of the labels of closely spaced integrals. Applies to both screen display and plot.
V Displays sub-integral labels vertically
H Displays sub-integral labels horizontally
Z Brings up another left scroll bar to adjust the beginning level of the integral line. An <ENTER> removes the second left scroll bar. Another <ENTER> exits the Integration sub-routine.
F Toggles between showing the sub-integrals and the full spectrum integral.
B Starts an integral/baseline correction process to provide a zero order and first order correction to the integral display. A BF or FB command before entering the integration routine is recommended to get a flat integral. If these don’t work, the B sub-command of the ID routine allows use of mouse movements to correct the integral display to make it flat. If the B sub-command is typed while displaying sub-integrals, the system is toggled to full integral display ( See F sub-command ). Within this baseline correction routine, the baseline is adjusted as follows:
Left Mouse Button Down Mouse movement left and right does zero order integral correction.
Right Mouse Button Down Mouse movement left and right does first order integral correction.
Q Aborts the process and ignores corrections made.
<Enter> Applies the baseline correction and exits the B sub-command.
Show me how to use the B routine.

Unlike other integral correction routines, this operation changes the baseline of the data, rather than just tilting the integral trace. The changes are permanent, not limited to the integral display. Because the integral is so sensitive to subtle baseline imperfections, this can be a very useful way to correct the baseline.

AI — Automatic Integration

This command displays an integral of the currently displayed region of the spectrum, with the integral broken into integral regions, and labels each region with its relative value. This command is available from the Tools menu. It is advisable to correct the baseline (with FB) before attempting integration.

When this command is executed while the entire spectrum is being displayed, any previously defined integral regions and scaling are overridden, whether they were created with AI or manually within the Integral Display (ID) subroutine.  If AI is executed while displaying an expanded region of the spectrum, only the displayed peaks are integrated.  The new integrals are added to any previously existing integrals, without changing the previously established scaling.  This allows a spectrum to be integrated piecemeal.

NUTS determines which peaks to integrate, and where to break the integral in the following manner. The spectrum is divided into 256 equal segments (with the requirement that each segment contain at least 16 points; if not, fewer segments are chosen). A region must contain a peak exceeding the Minimum Height (MH) value are integrated.  Continuous segments with no peaks above MH are ignored.  The RM value can be used to change the sensitivity by which AI determines which segments have peaks.

Starting from the left end of the spectrum, the first segment found to contain a peak will set the start of an integral region. Each subsequent segment will be included in that integral region if it contains a peak. When a segment is encountered which does not contain a peak, the integral region is ended. This procedure continues for the rest of the spectrum.

The smallest integral so defined is assigned a value of 1 and all other regions scaled accordingly. Following automatic integration, it is possible to enter the Integral Display subroutine and make changes, such as deleting unwanted integral regions, defining new regions or rescaling by setting the value of a chosen region. See the description of the ID routine, above, for details.

Display of the integral trace can be toggled on and off by typing Ctrl-I or selecting Show Integrals from the View menu. This works whether integral regions have been defined by either the automatic or manual methods. The commands AN and AF can also be used to turn On and Off the integral display, respectively. The latter 2 commands can be used in Links and macros. Ctrl-I is active at all program levels, including within subroutines. To use Ctrl-I in a Link or macro, use ^I.

In the non-2-letter command mode, the AI command will accept some arguments:

AI segments n – sets the number of segments the spectrum is divided into to look for segments which contain peaks. The default is 256 segments.

AI show on – turns on a displaying mode showing segments identified as having peaks.

AI show off – turns off the above displaying mode.

AI show – toggles the above displaying mode.

 

IL — Integral List

Transfers a list of sub-integral values to the clipboard, from which it can be pasted into the Notepad or other document for editing and printing. This is equivalent to the sub-command T within the Integration subroutine. All sub-integrals are included in the list, whether or not they are in the currently displayed region.

Control-I — Toggle on/off integral display

This is the same as the Display Integral option under the View menu. This can also be accomplished with the commands AN and AF, which turn integral display on and off, respectively. To include this in a Link or macro, use ” ^I “.

Note that the Tab key executes a Ctrl-I, and so performs the same function.

AN — Turn on integral display

Displays the integral trace. If integral regions were previously defined, either automatically (with AI) or manually in the ID subroutine, these regions will be displayed. Otherwise, the integral trace will encompass the entire spectrum. The integral trace can be turned off by typing AF. This operation can also be performed by selecting Show Integrals from the View menu or by typing Ctrl-I, both of which function as a toggle.

AF — Turn off integral display

Removes display of the integral trace. The integral trace can be turned on by typing AN. This operation can also be performed by selecting Show Integrals from the View menu or by typing Ctrl-I, both of which function as a toggle.

 

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Command Index

 


Last updated: 5/31/03.