Technical Note 70
Data Reporting in Chromeleon
1. IntroductIon
One of the most important functions of any Chroma-tography application is the reporting of processed data. If it is not possible to use the application to report data in accordance with laboratory requirements, it becomes necessary to transcribe or export data to other software applications. This process is time consuming and error prone, and is one of the major bottlenecks in transform-ing chromatography data into usable information.
The Chromeleon® Chromatography Management
System (CMS) provides powerful reporting capabilities that allow:
• Report templates: quick and simple generation of templates that can be applied easily to standard chromatography methods, independent of the num-ber of injections and peaks.
• Advanced reporting options: including summaries, trend plots and advanced charting.
• User-defined formulas: easy creation of formu-las such as the mathematical operators SUM and AVERAGE, and the logical operators IF and OR.
(Formula entry is analogous to Microsoft® Excel.)
• Improved compliance with regulatory rules such as 21 CFR Part 11.
These capabilities enable creation of reports that automatically calculate and display the required data and statistics, without the need to use additional software ap-plications. As such, the Chromeleon CMS allows users to concentrate on the science, not on result calculation, improving laboratory productivity.
2. tradItIonal data reportIng WorkfloW
In many laboratories the traditional workflow for generating a final report is as follows:
1. Integrate all chromatograms.
2. Export or transcribe peak results (such as peak area) into a third-party software package (such as Micro-soft Excel) for calculation.
3. Enter relevant values such as standard weights and sample weights into the third-party software application.
4. Evaluate calculated results against specifications. 5. Export or transcribe the calculated results into a
Certificate of Analysis (C of A) or similar document. 6. Print final report.
Potential drawbacks of this procedure include: • It can cause delays from data acquisition to creation
of the final result.
• Multiple steps must be repeated if chromatograms need to be reintegrated.
• Any data transcriptions require error checking and, if errors are found, repetition of steps 2–4 and 5.
2.1. Estimated Data Reporting Time
In this example, there are approximately 20 injec-tions, each containing five peaks. The total time required to obtain a result can be calculated using the following assumptions.
Therefore, the total time required to generate the final result is 137 minutes (8200 seconds).
2.2. Annual Data Reporting Time
This example assumes that an HPLC system runs one, 20-injection sequence each working day. With 250 working days per year, total data processing time per system is 24 days. Reducing this time can signifi-cantly improve laboratory productivity.
3. data reportIng WorkfloW In cHroMeleon
Chromeleon’s reporting features enable you to perform all chromatography calculations directly within the application, without the need to export data to third-party software. Chromeleon’s reporting tool supports flexible formats for any report type, including “Certifi-cates of Analysis.” The final results can be printed or
exported in Adobe® Acrobat® Portable Document Format
(PDF) for circulation.
With all this functionality, Chromeleon provides a much simpler reporting workflow:
1. Integrate all chromatograms.
2. Enter relevant values such as standard weights and sample weights into Chromeleon.
3. Evaluate calculated results by checking against specification.
4. Print or export (as PDF) final report.
This workflow still requires review of the output. If changes are needed, the new results can be printed or exported as PDF files as soon as the chromatograms are reintegrated. No re-transcription or re-export to a third-party application is required.
In a typical lab, the integration will probably need to be repeated at least once. This means that steps 1, 2, 4 and 5 must be repeated, adding 48 minutes, 20 seconds to the total time. Also, assuming at least a 5% error rate in transcription, corrections add 100 seconds to the total time (5% from step 2 and 5% from step 5). The total time for steps 1 to 5 is now 102 minutes (6100 seconds).
It is assumed that steps 5 and 6 are started only after steps 1 to 4 are completed. Assuming another 5% transcription error here, total time increases by another 100 seconds.
Step Process Time Taken
1 Integrate and check chromatogram 30 seconds per injection
Total = 600 seconds
2 Transcribe peak areas to 10 seconds per peak area third-party application Total = 1000 seconds 3 Enter values such as standard 10 seconds per sample weights and sample weights Total = 200 seconds 4 Evaluate calculated results by 15 seconds per injection checking against specifications Total = 300 seconds 5 Evaluate calculated results 10 seconds per peak area by reviewing transcription Total = 1000 seconds Total: 3100 seconds (51 min 40 s) 5 Transcribe results to C of A 10 seconds per peak result
Total = 1000 seconds
6 Review C of A transcription 10 seconds per peak result
Total = 1000 seconds
Total: 2000 seconds (33 min 20 s)
Table 1. Estimated Data Reporting Time for Traditional Workflow
3.1. Estimated Data Reporting Time
Using the same sequence as section 2.1, the fol-lowing data reporting time can be calculated for the Chromeleon workflow.
4. poWerful data reportIng WItH cHroMeleon
How can Chromeleon offer these kinds of benefits? The standard installation provides several report tem-plates designed to report most types of data, including chromatograms, integration reports and calibration curves as well as standard results such as % w/w, % Label Strength. Chromeleon can compute more than 300 different chromatography values, such as peak area or number of theoretical plates, speeding creation of reports that display the information needed by the labo-ratory. Approximately 90% of standard chromatography values and results can be automatically reported with the default Chromeleon installation.
In addition, Chromeleon supports user-defined formulas, similar to those in Microsoft Excel, making Chromeleon an all-in-one, fully integrated data process-ing and reportprocess-ing application.
5. report defInItIon fIle exaMples
Chromeleon reports, like Excel workbooks, may contain several worksheets that display different types of data and results. All Chromeleon installations come with a set of standardized reports that contain sheets such as Integration, Calibration, Peak Analysis, Sum-mary tables, etc.
This following sections provide examples of addi-tional reports that can be used to save time and increase productivity.
5.1. Summary Table
Each default Chromeleon report contains a sum-mary sheet. This is particularly valuable as it displays an instant summary of results over a series of injections. It can be easily modified to show only those injections that are relevant to the user, for example, only standards or only unknowns.
The example below summarizes a sequence contain-ing 10 samples. Each sample was injected in duplicate and contained five peaks. The summary requirements were to compare the average amount (in mg/ml) against the peak specification, and to report whether the result passed or failed.
Step Process Time Taken
1 Integrate and check chromatogram 30 seconds per injection
Total = 600 seconds
2 Enter values such as standard 10 seconds per sample weights and sample weights Total = 200 seconds 3 Evaluate calculated results 5 seconds per injection by checking against specification Total = 100 seconds 4 Print or export (as a PDF) 0 seconds everything is final results (e,g, C of A) performed automatically
by Chromeleon
Total: 900 seconds (15 min)
Table 1. Estimated Data Reporting Time for Chromeleon
Assuming that the sequence will need reintegrated one time, adding 700 seconds, the total time required to generate the final result is 27 minutes (1600 seconds). Compare this to 137 minutes for the workflow in section 2.1.
3.2 Annual Data Reporting Time
Assuming, as per section 2.2, that each HPLC sys-tem runs 250 sequences per year, the total annual data reporting time is 4.7 days. Compared with the traditional data workflow, that is a saving of 19.3 days—an 80% reduction in the total time.
Simple productivity calculations such as these show the tangible business benefits to implementation of a Chromeleon Chromatography Data System.
• Improved data integrity: Even with double-checking, errors in transcribed data may not be detected. This problem is eliminated when working with Chrome-leon reports.
• Time and cost savings: Clear reductions in reporting time.
• Workflow improvements: Automation of repeti-tive and mundane tasks and software that is more enjoyable to work with increase motivation and free resources to focus on science, not reporting.
Figure 1 shows the summary. It automatically cal-culates the average value for each set of duplicates, and compares the values against a specification.
A summary table is automatically generated for each peak in the processing method. This report format will work for any method, with any number of peaks and any peak names.
5.2. Dissolution Summary
Chromeleon provides advanced charting options. This enables data such as dissolution data to be dis-played graphically.
In this example, three batches of a compound were tested in a single dissolution bath of six vessels, that is, one batch per two vessels. The report shows the disso-lution profile of each vessel, and the average profile of each batch.
5.3. Related Impurities Testing
In related impurities testing, a duplicate injection of a sample is typically performed. All unidentified peaks are then integrated and the average result (generally percent area) for each peak is reported. Due to the nature of related impurities testing, the following complications generally arise:
• The number of peaks that will be detected is un-known.
• Peaks can be detected in one duplicate but not in another, as they are close to the limit of detection. These complications can make it difficult to auto-mate the calculation. However, it is possible to do this using Chromeleon, using user-defined formulas.
As can be seen in Figure 3, the average percent area has been calculated automatically. In this case, if the percent area is below 0.05%, it is reported as <RL (less than the reporting limit). The report will calculate results correctly, independently of the number of peaks. It can also easily be extended to cover methods that re-quire triplicate injections, or duplicate injections of two sample preparations (four injections in total). This report is one of the major time savers for laboratories.
The figures in section 3.1 show that data reporting times increase significantly as the number of reported peaks increases. This is not the case with Chromeleon. Companies using these reporting options have realized time savings of up to 98%, mainly because the data reporting step is fully automated.
6. IMproved regulatory coMplIance
If third-party applications such as Microsoft Excel are used to perform calculations, any such applications must be validated, taking valuable laboratory time. Us-ing Chromeleon to calculate the final results minimizes validation time.
Standard results, such as peak area, require no vali-dation as these are tested during the software develop-ment stage (Design Qualification) at Dionex, and during
Figure 1. Summary of five peaks from 20 injections (10 samples with duplicate injection).
Figure 2. Dissolution profiles using Chromeleon charting options.
Figure 3. Summary of related impurities for a duplicate injection.
installation at your site (Operational Qualification). Any calculations performed with “user-defined formulas” should be validated. For example, if a user-defined for-mula generates a Pass/Fail statement, the forfor-mula must be validated to show that it produces the correct state-ment for the data value generated. Chromeleon contains the following tools that make this process easier: • Display and print formula entered (see Figure 4) • Tracks history of changes made to formula
(Fig-ure 5)
Figure 4. Displaying user-defined formulas.
Figure 5. Chromeleon History Trail tracks all changes made to reports, including changes made to user-defined formulas.
7. suMMary of BenefIts
Chromeleon provides a suite of powerful report-ing options that make it possible to fully automate the complex calculations commonly used in today’s labo-ratories—yet impossible to complete in other Chroma-tography Data Systems. Chromeleon removes the need to calculate results using third-party applications such as Microsoft Excel or to manually transcribe results into separate files.
Our examples showed that by using Chromeleon to eliminate these steps it is possible to reduce general data reporting times by 80%, and in the case of specialized reporting requirements, such as related impurities, by as much as 98%.
Since Chromeleon removes the need to validate third-party software applications, validation costs are also reduced. Regulatory compliance is improved, through Chromeleon’s ability to track all changes made to report definition files, even down to the level of changes to user-defined formulas.
Chromeleon’s flexible reporting tools make it possible to generate results in almost any conceivable format, enabling different departments to work as best suits them.
By automating data calculation and reporting Chromeleon not only saves time and increases produc-tivity, it allows scientists to concentrate on the most important laboratory task: the science.
AppEnDIcES – TEchnIcAl InfoRmATIon
a1. report defInItIon fIles
The way in which data is printed (or electronically signed) is defined in a Report Definition File (RDF). These are similar in look and feel to Microsoft Excel spreadsheets. Figure 6 shows the general structure of a report definition file.
Figure 6. Main components of a sheet in a report definition file.
The specifications of each sheet are as follows:
Sheet Name Any characters can be used. Maximum length is 31 characters.
Sheet Size 256 columns × 16,384 rows
Cell Size Maximum number of characters per cell is 255.
Organizing data into separate sheets makes evaluat-ing and interpretevaluat-ing large amounts of data much easier. It also provides more flexibility when printing and exporting. In a typical example, one sheet might display the chromatogram and results sample by sample, while another sheet displays the calibration data and curves for the sequence. It is also possible create sheets that display a summary of the results. In fact, the number of sheets that be created within a report definition file is limited only by your computer memory.
a2. report defInItIon fIle oBjects
Table 3 shows the types of Chromatographic objects that can be inserted within each sheet of a report defini-tion file. These objects can be used in any combinadefini-tion.
Each of these objects is described in more detail below.
A2.1. chromatogram
The chromatogram offers a number of different dis-play and formatting options. In essence every single part of the chromatogram can be customized to show data that is relevant to the analysis (see Figure 7).
Object Description
Chromatogram Freely movable and scaleable representation of the chromatogram
Calibration Plot Freely movable and scaleable representation of the calibration plot
Spectra Plot Freely movable and scaleable representation of the spectra plot, showing a 2D
representation of the UV spectra from any point in the chromatogram.
Mass Spectra Plot Freely movable and scaleable representation of the mass spectrometry plot.
3D Field Plot Freely movable and scaleable representation of the 3D plot.
Trend Plot Freely movable and scaleable representation of the trend plot, showing any chromatogram result against either injection time or sample number.
Fraction Tray Plot Freely movable and scaleable representation of the fraction tray.
Inject Tray Plot Freely movable and scaleable representation of the inject tray.
Chart Freely movable and scaleable charting object. Chromeleon Report Variable Display of a single result or value associated
with the data.
Chromeleon Report Table Display of a result or value set associated with the data.
Free Text A free-text entry.
User-Defined Formula A user-defined formula similar to the formulas available in Microsoft Excel.
Figure 7. Examples of different chromatogram options. (A) Peak tangents and peak width displays. (B) Baseline zoom to view smaller peaks or to check integration. (C) Overlaid chromatograms. (D) Fraction collection display—fractions are marked by green shading.
A
D c
B
A2.2 calibration plot
The calibration plot shows the individual calibration points and the best fit line for a specific peak. It can also show the calibration confidence levels (see Figure 8).
A2.3 Spectra plot
The spectra plot shows a UV spectrum from any point in the chromatogram and is only available for data acquired using a Diode Array Detector (see Figure 9).
Figure 8. Examples of calibration plot display options.(A) Default calibration plot. (B) Calibration plot with 95% confidence limits. Limits of 95, 98, 99, 99.7, 99.8, 99.9, and 99.99 can be shown.
Figure 9. Examples of spectra plot display options. (A) Standard spectra plot showing the UV spectrum at the peak apex. (B) Standard spectra plot showing the UV spectrum at the peak apex. (C) Spectra plot and first derivative plot. (D) Spectra plot overlaid with spectral library hit plot.
A
D c
B
Figure 10. Display options for 3D field plots. (A) ISO plot. This is a contour plot of the 3D field. (B) 3D plot. The 3D field is displayed from a user-defined perspective.
Figure 11. Display options for Mass Spectra Plots. (A) Profile mode. This provides the true mass response for each peak. (B) Centroid mode. This bunches the mass response to provide single m/z values for the peak. This example shows an overlay of three spectra from the same peak: 10% peak height at the front, peak apex, and 10% peak height at the tail.
A2.4. 3D field plot
The 3D field plot shows all data from a diode array acquisition, that is, the signal from all acquired wave-lengths at all retention times. As such, this plot has two main display options as seen in Figure 10.
A2.5. mass Spectra plot
The mass spectra plot shows spectra from any point in the chromatogram using data acquired from a mass spectrometer (see Figure 11).
A B
Figure 12. Display options for trend plots. (A) Trend plot with mean and standard deviation calculated from data values. (B) Trend plot with mean and standard deviation manually defined. This is generally used as a control chart.
A2.6. Trend plot
The trend plot in Chromeleon can display data in two main ways.
• As a trend of data against injection time or sample number, with the mean and standard deviations calculated based on the values in the trend.
• As a trend of data against injection time or sample number, with a user-defined mean and standard deviation. This can be used as a control chart, for example.
Figure 12 shows an example of the main two display options.
A2.7. Inject Tray plot
The inject tray plot shows a graphical representation of the injection tray. It is available in two main modes, the plot of a standard autosampler tray and the plot of an injection tray for a fraction collection instrument. Figure 13 shows an example of the two display modes.
Figure 13. Display options for inject tray plots. (A) Inject tray plot for a standard autosampler tray. The active sample appears in green. (B) Inject tray plot for a fraction collection instrument. The inject tray is shown in the main part of the plot. The position of the tray within the fraction collection instrument is a darker shade of gray. All other settings are as for view (A).
A B
A2.8. fraction Tray plot
The fraction tray plot shows the location of fractions collected. It is linked to the inject tray plot. That is, if the inject tray plot shows that the active sample is position 7, the fraction tray plot shows all fractions collected from this injection. As such, there is only one display mode, as shown in Figure 14.
Figure 14. Display option for fraction tray plots. A. Fraction Tray Plot. A. This plot displays the location of all fractions collected. The areas marked in yellow show the location of fractions col-lected from the active sample. This plot also shows the position of the fraction tray within the fraction collection instrument (in a darker shade of grey).
Figure 15. Using a bar chart to display instrument utilization data.
A2.9. chart
The charting option provides more advanced chart-ing than is available through the trend plot. For example, Figure 15 shows a bar chart of instrument usage data.
Table 5 shows all available chart types.
Type 2D 3D Area Bar Line Step Combination Pie Horizontal Bar Cluster Bar na Hi Lo na Gantt Bubble na Doughnut na Contour na Surface na XY (Scatter) na XYZ (Scatter) na Polar na Radar na
A2.10. chromeleon Report Variables
A Chromeleon report variable shows a single Chromeleon value or result, such as sequence name or sample name. There are other 300 variables available from several different categories such as Peak Results
(area, height, etc.) and Peak Calibration (slope, R2, etc.).
Numeric variables can be modified using four math-ematical operators: plus, minus, multiply and divide. They can be combined with absolute values (Figure 16) or with other variables.
Figure 16. Dividing the Relative Retention Time variable by 100 to generate an absolute value, rather than a percentage value.
A2.11. chromeleon Report Table
A Chromeleon report table displays a collection of chromatography results. For example, an integration table for a chromatogram will display variables such as peak area and height for all peaks. The two main table types are:
• Integration table. This displays the peak results for each chromatogram.
• Summary table. This displays results for each injec-tion.
Note: Summary tables are particularly useful since they evaluate data for an entire sequence in a single table, saving time that would be required to review each individual integration report. In addition, summary tables can be filtered to contain only appropriate results. The filter can include or exclude samples according to their type, name, comments, or other fields, then sort the results as required. Figure 17 shows an example filter and sort settings for a summary table.
Figure 17. Defining the data set to be shown in a summary table.
A2.12. User-Defined formula
Chromeleon supports user-defined formulas that are analogous to those used in Microsoft Excel, to perform calculations that are not included as standard Chrome-leon calculations. Table 6 shows the available functions.
For example, if it is necessary to display the maxi-mum value from a set of results the “MAX” function can be used. Detailed explanations for each function are available in the Chromeleon online help system. In all cases they operate in the same way as the equivalent functions in Microsoft Excel.
ABS ACOS ACOSH ADDRESS
AND ASC ASIN ASINH
ATAN ATAN2 ATANH AVERAGE
CEILING CHAR CHOOSE CLEAN
CODE COLUMN COLUMNS CONCATENATE
COS COSH COUNT COUNTA
COUNTIF DATE DATEVALUE DAY DAYS360 DBCS ERRORTYPE EVEN
EXACT EXP FACT FALSE
FIND FINDB FIXED FLOOR
HLOOKUP HOUR IF INDEX
INDIRECT INT ISBLANK ISERR
ISERROR ISLOGICAL ISNA ISNONTEXT ISNUMBER ISREF ISTEXT LEFT
LEFTB LEN LENB LN
LOG LOG10 LOOKUP LOWER
MATCH MAX MID MIDB
MIN MINUTE MOD MONTH
N NA NOT NOW
ODD OFFSET OR PI
PRODUCT PROPER RAND REPLACE
REPLACEB REPT RIGHT RIGHTB
ROUND ROUNDDOWN ROUNDUP ROW
ROWS SEARCH SEARCHB SECOND
SIGN SIN SINH SQRT
STDEV STDEVP SUBSTITUTE SUM
SUMIF SUMSQ T TAN
TANH TEXT TIME TIMEVALUE
TODAY TRIM TRUE TRUNC
TYPE UPPER VALUE VAR
VARP VLOOKUP WEEKEND YEAR
Table 6. Functions a3. autorepeat areas
Autorepeat areas expand the power of the Chrome-leon reports. Any area of a sheet can be defined as an autorepeat area to be printed repeatedly for each set of channel, sample or peak information.
For example, a summary table might report values for a specific peak in all samples. If there is more than one peak, it can be time consuming to create a new sum-mary table for each peak. Instead, the sumsum-mary table can be defined as an autorepeat area that automatically reprints for each peak in the method.
The following autorepeat area options are available.
A3.1. print and Repeat channels
This option applies if more than one data channel is acquired, for example, one channel at 230 nm and one channel at 254 nm. Chromeleon can autorepeat an area for all channels, or for specific channels. Figure 18 shows an automatically-generated two-page chromato-gram and integration report for a sample with two data channels.
A3.2. Repeat Samples
This option prints information about different samples on the same page. For example, it can print all chromatograms of the same replicates on the same page. (See Figure 19.)
Figure 18. Automatically repeated chromatogram and integration report for a sample with two data channels.
Figure 20. Automatically repeated calibration plot for each peak in the chromatogram.
A3.3. print and Repeat peaks
This option applies if there is more than one peak in the chromatogram. Chromeleon can repeat an area of the report for all peaks, or for specific peaks. Figure 20 shows an automatically-generated report that shows the calibration plot for all peaks, with the associated calibra-tion data.
A3.4. Benefits of the Autorepeat function
The autorepeat area function makes it possible to create a single report template that will be applicable to almost all methods. It does not matter if the method uses one data channel or several, or analyzes only one compound or several. The report will automatically ad-just based on the data collected to show all the relevant results.
Other benefits are:
• Reduced administrative costs: create and apply a single template for multiple methods.
• Reduced error: ensure that all data is reported cor-rectly.
• Easier review (users become quickly used to locat-ing important data settlocat-ings)
a4. prInt settIngs
Each sheet in a report (e.g. integration sheet, sum-mary sheet, etc.) has different printing requirements. For example, an integration report is generally printed for each chromatogram, whereas a summary report is printed only once. Chromeleon makes it simple to define when each sheet of a report should be printed. The main settings are:
Figure 21. Printing conditions for sheets.
“Print under certain conditions” provides several the options shown in Figure 21.
These options provide very fine control over the conditions in which a sheet is printed. For example, a calibration sheet might print only for standards, or a Summary Sheet might print only for the last sample in the sequence.
Do not print Used if sheet shows data for information only
Print for every sample Generally used for sheets such as an integration report
Print under certain conditions Generally used for sheets such as a summary report.
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