Graphics And Printing
The GRAPHICS MENU allows you to control the appearance of the graph, including selection of Data files, labels, colors, scaling, etc. This section is interactive, but settings made here will carry over into graphs produced by automatic processing. Changes made to the graph options can be made permanent by saving the REPORT.INI file.
The MATH OPERATIONS section allows you to manipulate the data in various ways (e.g., baseline subtraction).
PRINTING options provide flexibility for setting up printers and selectively printing all or parts of reports.
GRAPHING DATA
Begin by taking the GRAPH DATA choice from the Graphics Menu:
The GRAPH OPTIONS window opens, allowing you to select the options controlling the appearance of the graph:
Each Data file in RAM will be listed in the dialog box. Simply click on the check box to request that one or more of the Data files appear in the graph. A drop-down listbox allows selection of a unique color for each Data file.
Color selection also is provided for other aspects of the graph: you may select whether or not the calculated baseline is drawn underneath peaks, whether or not peak start and stop markers are included, and the colors for these options. Background and border colors also are provided.
The Show Color Assignment in Graphic Display checkbox allows the name for each data set to be printed below the graph, in the appropriate color, when more than one data set is shown on the same graph (i.e., when SPLIT SCREEN is turned off).
Depending on the hardware used during data collection, the graph might already be scaled in appropriate detector units. With external detectors, however, the graph might be scaled in arbitrary Y-units. The units of the Y-scale are purely esthetic: they have no effect on quantitation. However, you may change the units and the axis label to whatever seems appropriate. Press the MATH button, and change FULL SCALE Y and LABEL Y UNITS on the Math Operations screen to something suitable for the detector that was used. (If you find yourself doing this very often, you can change the scaling parameters in Control so data from external detectors are saved with the appropriate information.
Press the OK button and the graph will be drawn. The size of the graph on the screen (and printer) may be adjusted by dragging its borders in or out with the mouse.
The size of the peaks within the graph may be scaled in several ways. For temporary rescaling that only affects the current Data file, see CURSOR TYPES. For rescaling that affects all the Data files to be processed, use the GRAPH SCALING options in the GRAPH DATA OPTIONS dialog box as follows:
AUTOMATIC SCALING is the simplest method of formatting the graph. In general terms, automatic scaling fits the length of the Data file into the X (time) axis, offsets the baseline about 15% above the bottom of the graph, and puts the highest peak near the top of the graph. This generally produces an acceptable graph:
If the baseline is very negative and offscale, uncheck the Neg.Limit-10% box. The box is usually checked, to prevent large negative excursions of the baseline (e.g., the injection disturbance) from affecting scaling.
There are two drawbacks to automatic scaling. First, since each graph is separately scaled, it's hard to do visual comparisons of several Data files processed in a batch: as each file is processed and graphed, it will be maximized to fit the graph. (NOTE: peak heights, areas, and calculated amounts from the report are unaffected by scaling. These numbers may be used for comparing Data files.)
The second drawback to automatic scaling is that the scaling algorithms cannot distinguish between the void-volume response and a peak. At sensitive gains, the detector may show a full-scale deflection in response to the injection, while the true peaks may be quite small. Automatic scaling will fit the void-volume response to the graph, and consequently the peaks may be tiny:
FIXED SCALING allows you to specify the upper and lower limits for the Y (and less commonly, the X) scale. The figure below shows an improved graph of the same Data, achieved by using fixed scaling. The fixed scale is retained in RAM, so every subsequent Data file will be graphed on the same scale. The fixed scale can be saved for subsequent use by saving the REPORT.INI file.
To use fixed scaling, first load the Data file with the largest peaks from your series of runs (this is generally the high standard). Process it with automatic scaling, then observe the graph. Use the Y-scale on the automatically scaled graph to estimate the appropriate upper and lower limits for the fixed scale. Then select GRAPH DATA. Click on the Fixed scale option, and enter the Min. and Max. Y-axis values. Click on OK to see the results, and if necessary, further refine the upper and lower limits. The X-axis can be scaled using the Left and Right time boxes.
The LABEL PEAKS WITH option allows the peaks to be labeled with their retention time, number (sequential number from the integration report or name (from the Standards file). These options are only available when the Data file has been processed. Unprocessed data may be graphed but not labeled.
FORMATTING OPTIONS
The formatting options include the following choices for controlling the appearance of the graph:
FILL PEAKS colors in the area under each peak. Peaks will be filled with the line color selected for that Data file. If several Data files are graphed simultaneously, only the most recently processed Data file will have filled peaks.
OVERVIEW WINDOW enables a second window which always shows the entire chromatogram. When a section of the main graph has been enlarged with the Zoom Cursor, the enlarged section is indicated on the Overview Window by a rectangle. The relative sizes of the Overview Window and the main graph can be adjusted by dragging on the border between them with the mouse.
SPLIT SCREEN divides the screen horizontally so that Data files will graph separately rather than overlap.
GRID DOTS overlays a grid on the graph, to facilitate visual estimations of peak height and retention time. For more accurate estimates, enlarge the peak of interest with the Zoom Cursor so the grid switches to a finer scale.
These options can be turned on or off by clicking on their names in the Graphics Menu:
The figure below shows a graph with Split Screen, Overview Window, Fill Peaks and Grid Dots turned on:
INTERACTIVE CURSORS
Five cursor types allow interactive manipulation of the graph. These are ZOOM, SHIFT, SCALE, PEAK MARK and EVENT. The cursor type may be selected from the Graphics Menu:
Alternatively, if CURSOR INFO is checked in the Graphics Menu, cursor type may be chosen from a selector box that appears near the graph:
Each cursor type has a distinctive shape when positioned on the graph. In addition to allowing cursor selection, this box provides a continuous display of the cursor's position on the X and Y axes. To select a cursor type, click on the appropriate button in the selector box.
ZOOM CURSOR. Parts of the graph may be enlarged using the Zoom Cursor, which is the default cursor type. You can select the Zoom Cursor, if necessary, by clicking on its button. To use the zoom feature, simply click and hold the left mouse button on a section of the graph, drag the cursor to another section (thus making a rectangle) and release the button. The rectangle can be drawn either on the main graph or on the Overview Window, but only the main graph shows the enlargement:
Notice that the Overview Window retains the original graph, with a rectangle delineating the enlarged area. This rectangle can be dragged along the Overview Window to enlarge other sections of the graph. To return from an enlarged graph to the previous view, position the cursor on the main graph and click the right mouse button.
SHIFT CURSOR. The Shift Cursor may be used to move the peaks of the most recently processed chromatogram to the right or left, or to line up peaks on two chromatograms. A shift to the right or left actually changes the retention times of the data in RAM. To use the Shift Cursor, first select it by clicking on its button. Then click and hold the left mouse button anywhere on the graph. Drag the cursor to make a line in the desired direction of movement, then release the mouse button. If the shift affects the time axis, a warning message will appear, indicating that the data must now be reintegrated. Reintegration is needed only if you wish to analyze the modified data; it is not needed for replotting and visual examination. Click on OK and the new graph will be drawn. A shift cannot be easily undone. If this becomes necessary, reload the original Data file.
SCALE CURSOR. The Scale Cursor allows the peaks in the most recently processed chromatogram to be stretched or shrunk in any direction. Select this cursor by clicking on its button. To rescale the chromatogram, click and hold the left mouse button anywhere on the graph, then drag a line in the desired direction. The length of this line is proportional to the amount of enlargement or reduction. Lines drawn up and to the right enlarge, while those drawn down and to the left reduce. When the mouse button is released the new graph will be drawn. If the scale operation occurs along the time axis, a warning that the data must be reintegrated appears. Reintegration is needed only if you wish to analyze the modified data; it is not needed for replotting and visual examination. The most common use of the Scale Cursor is to enlarge the peaks prior to printing the graph. A change in scale is not easily undone. If this becomes necessary, reload the original Data file.
PEAK MARK. The Peak Mark (Mark) Cursor allows interactive adjustment of peak start and stop markers on the most recently processed chromatogram. To move a peak marker, first select the Mark Cursor by clicking on its button. Then simply click and hold the left mouse button on a marker, drag it to another location, and release the mouse button. You must move the cursor to the precise location in the horizontal direction. Vertically, however, it will jump to the baseline when you release the mouse button. A warning that the data must now be reintegrated will appear. Reintegration is needed only if you wish to analyze the modified data; it is not needed for replotting and visual examination.
To remove a pair of peak markers, position the cursor between the start (up arrow) and end (down arrow) markers and press the delete key. To insert a pair of markers, position the cursor above the baseline, near where you want them. Press the insert key to insert the pair of markers, then move them with the mouse. It is helpful to enlarge the chromatogram with the Zoom Cursor before moving baseline markers.
A peak marker may not be moved across another peak marker. For example, an up-arrow (peak start) may not be moved to the right of its down-arrow mate, nor to the left of any previous markers.
Moving the baseline markers does not change either the Data file or the Method. The Data file will have to be reintegrated in order for the modified peak markers to be recognized. If you redetect peaks the original placement of the markers will be restored.
EVENT CURSOR. The Event Cursor allows processing events to be interactively inserted and deleted. Existing events are indicated by letters along the lower margin of the graph:
A corresponding letter over the cursor indicates the type of event to be edited:
| E (e) | The initial state (can delete any event) |
| D | Peak detect on |
| d | Peak detect off |
| N | Negative peak on |
| n | Negative peak off |
| P | Perpendicular drop on |
| p | Perpendicular drop off |
| F (f) | Fix baseline |
| T (t) | New threshold |
To use this feature, first click the Event button. Then press the appropriate key so the letter code for the function of choice appears above the cursor. Position the cursor over the baseline, then press the insert key to insert an event, or the delete key to remove an existing event. The events are inserted or deleted from the Method in RAM, and any changes will appear in the Processing Events dialog box. The Data file must be processed for the changes to be put into effect. Subsequent Data files also will be processed with the changed events. SAVE the Method to make the changes permanent. If it becomes necessary to restore the original Events, do not save the changed Method: instead, reload the original Method.
MATH OPERATIONS
A number of mathematical operations are available through the MATH OPERATIONS section of the Graphics Menu. These include addition and subtraction of Data files, normalization, smoothing, derivatization, etc. Most of these are beyond the scope of this introductory manual, and additional information about them can be found in the Help screens. One feature of more general interest is subtraction, which can be used to remove from the samples any regular and predictable disturbances in the baseline. These disturbances may include slope (e.g., from gradients) or peaks from contaminants or derivatization reagents.
To perform a baseline subtraction, first perform the chromatography and collect the data. You should have a sample run and a relevant blank run. Next load the two runs into data slots in Report. Select the Math Operations section of the Graphics Menu:
Designate the sample Data file as Source 1 and the blank Data file as Source 2. Select subtract (-) as the Operation, and any unoccupied data slot as the Destination Data file. Press OK and the operation will be performed. The new Data file will be called 1Sub2, and can be processed like a typical Data file.
PRINTING
ChromGraph offers various options for printing. For example, via the Report Options section of the Method you can instruct Report to send all output to the printer. Also, various other menu options employ LIST DEVICE boxes which permit you to send that task's output to the printer. Occasionally you may want to print or capture only the screen currently displayed. Options in the PRINT MENU afford that opportunity:
Text on the output screen can be sent either to Windows' Clipboard, for pasting to other Window's programs, or to a printer. First highlight the text by dragging the mouse over the lines of interest. Then click on either COPY TEXT (to send it to Clipboard) or PRINT TEXT (to send it to the printer). To print graphics, select PRINT SCREEN to send the entire screen (text and graphics) to the printer, or PRINT GRAPH to send only the graphics. To send only the graph to Clipboard, press COPY GRAPH. To send the entire screen to Clipboard, press the PRINT SCREEN key on the keyboard.
PRINTER SETUP opens a dialog for choosing among several printers and selecting certain options. This need not be done if a single printer has been configured under the Windows system.
PRINTER FONT opens a dialog for selecting printer font type and size. We have found the True Type (TT) fonts to work well under all conditions.
