Interactive Charts: A Powerful New Tool for Understanding Chromatography Data
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1 Interactive Charts: A Powerful New Tool for Understanding Chromatography Data Jay Lorch and Ryo Komatsuzaki; Thermo Fisher Scientific, Sunnyvale, CA, USA
2 Abstract Most laboratories are constantly under pressure to increase productivity, which can be defined in terms of results delivered per unit time. Advances in methodology, instrumentation, and automation have significantly reduced analysis times, often resulting in much greater volumes of data generated every day. This has created a compelling need for fast, efficient data management tools that can help analysts keep up with the data deluge. A new interactive charting capability provided in a modern chromatography data system facilitates rapid visual screening of results, enabling analysts to quickly gain an overview of trends and patterns, and to spot anomalies or other specific items of interest. Single-click switching to different chart types or source data, combined with instantaneous redraw, allows large volumes of data to be scanned very quickly. Dynamic links between the charted values and their source data provide an interactive experience in which analysts can instantly access details behind the data points, choose different data sets to plot, and gain understanding of the correlations between different aspects of their data. This solution provides advantages over traditional tools for reviewing data, such as tabulated results or externally-generated charts. Various applications of interactive charting for data management and chromatographic troubleshooting are shown. Coping with the Data Deluge Most chromatographers today need to manage much more data than was typical just a few years ago. Demands for higher productivity and higher confidence in results, combined with faster instrumental methods, have significantly increased the average daily volume of data, making data management more challenging than ever. To keep up, chromatographers need to adopt new tools that enable faster analysis of larger data sets. There simply is not enough time available to scrutinize every chromatogram anymore, so analysts need efficient ways to view the big picture, to spot significant items within the larger data set, and to quickly analyze specific items that need closer examination. A natural and effective way to address the challenge is to present data graphically. Graphical representations allow large amounts of data to be presented in a compact format, and they convey information quickly by taking advantage of the human brain s fast visual processing (which is much faster than the language processing required to interpret text and numbers). Graphics also facilitate comparison, and can make anomalies easier to detect. An example of these principles is the MiniPlots feature of the Thermo Scientific Dionex Chromeleon Chromatography Data System software (Figure 1). By presenting a detailed miniature thumbnail image of the chromatogram corresponding to each injection in a list, the software enables the user to rapidly scan through dozens of injections, and immediately spot any that deviate from the others (different number of peaks, different peak sizes, sloping background signal, and so on). MiniPlots enable analysts to spot-check raw data many times faster than was possible before, when chromatograms could only be accessed one by one. FIGURE 1. MiniPlots provide an instant visual summary of the raw data corresponding to each injection in an Injection Sequence or Query Result. Large numbers of chromatograms can be quickly surveyed and compared, and even subtle differences can be easily spotted (note Injection #16). The MiniPlots can be resized in both dimensions to show more details of complex chromatograms, or to fit more MiniPlots into the viewable area. Any chromatogram of interest can be immediately investigated by double-clicking on its MiniPlot, which displays the chromatogram in a larger Chromatography Studio window that facilitates close examination. 2 Interactive Charts: A Powerful New Tool for Understanding Chromatography Data
3 For result data, the Chromeleon 7.1 Chromatography Studio provides a rich set of interactive charting features (Figure 2). The charts can use any reportable parameter as either a dependent or independent variable, and can display the parameters across a single injection or across a range of injections. Mouse-based zooming is supported, making examination of details fast and easy. Many different chart types are available, including line, column, pie, scatter, area, bubble, and more. Multiple Interactive Charts can be configured in a workbook, and later accessed with a single click on the corresponding tab page in the workbook. Charts can also be included in reports (as in Figure 6). These result charts do not have the interactive selection and zooming features, but they offer all the same formatting capabilities as Interactive Charts, and also update dynamically when data are changed. Among the reportable parameters that can be included in Interactive Charts and Result Charts are: Hundreds of common chromatography variables (such as amount, area, height, retention time, peak width, retention factor) Instrument parameters (such as background signal, pressure, temperature) and other data recorded in Audit Trails Any number of custom user-defined variables (with correspondence to sequences, injections, or components) Basic mathematical formulas can be applied to any of the above, providing even more flexibility (Figure 3). This broad flexibility allows the great majority of chromatography-related charting needs to be satisfied directly by the chromatography software. FIGURE 2. In the Chromeleon 7.1 Chromatography Studio, an Interactive Chart (lower right) instantly plots results for any selected variable(s) across a single injection, or across a range of injections. Each chart is dynamically linked with the other panes in the Studio: when an item is selected in one pane, all the linked panes instantly update to show the data related to the selection. Thus, selecting a data point in the chart highlights the corresponding injection, displays the corresponding chromatogram with the relevant peak indicated, highlights the corresponding result in the Interactive Result Tables, and so forth. Similarly, selecting a different injection, a different peak, or a different result instantly shows an updated Interactive Chart. FIGURE 3. Chartable parameters include hundreds of common chromatography variables that are automatically calculated by Chromeleon Chromatography Data System software. Many of these variables can be customized using built-in options (such as those shown here for Signal-to-Noise Ratio). Numerous instrument parameters can be charted using values from the Audit Trail category. Still more flexibility is provided by the ability to create custom user-defined variables with correspondence of one value per Sequence, per Injection, or per Component. Any of these variables can be further customized using simple mathematical formulas (note the 10* in the Formula field). Thermo Scientific Poster Note PN70361_e 11/12S 3
4 Advantages of Integrated Charts over Externally Created Charts Using charts to present and interpret chromatography data is certainly not a new idea; scientists have used commercial off-the-shelf software for many years to analyze results from chromatography data systems. What is new is the complete, seamless integration of a charting capability into the chromatography software. This offers many advantages over charts created using external programs: Integrated Charts No extra software required Single, integrated, consistent user interface One-click charting Instantaneous dynamic updates Chart always consistent with source data Instant drill-down to source data and metadata Externally Created Charts Extra software must be purchased, learned and maintain Second user interface for charting, with many additional functions not related to the purpose Export/import steps required, consuming more time and creating risk of error Updates require data to be re-exported and reimported Results can be changed without chart reflecting the updates Locating source data and metadata is a tedious manual process, with no guaranty of success External charting software may offer additional capabilities not available with integrated charts, such as macro/scripting features and facilities for merging data from sources other than the chromatography data system. Nevertheless, for a large proportion of chromatography charting applications, integrated charts offer all needed capabilities, and their advantages over externally created charts are compelling. Sample Applications of Interactive Charts Trending and Control Charting Figure 2 shows a typical example of a trend plot, in this case showing peak retention time vs injection name for a series of injections of a mixed standard interspersed with water blanks. Horizontal lines on the plot define a band of ±2 standard deviation units (2σ) around the mean value, making outliers easy to spot. Statistical overlay options include mean, linear trend line, 1σ, 2σ, and 3σ. Control charts can be easily created by defining the target value, upper limit, and lower limit for each charted variable (Figure 4). The values can be defined using static values, or formulas incorporating any variables from Chromeleon Chromatography Data System software s extensive selection. Results outside the acceptability band are easily recognized, and can be immediately investigated by simply clicking on the stray result. FIGURE 4. Any chromatography variable can be tracked in a control chart. The target value, upper limit, and lower limit can be set individually for each component using static values, or using formulas involving any chromatography variables (for example, a ratio to a different component s result). In this example, a column-style chart is being used to track peak areas for three compounds of interest in a series of standards injected to test an experimental electrode. The chart clearly shows low values for all three compounds in injection #16 (corresponding to the blue chromatogram in the overlay), suggesting an injection problem such as an air bubble. However, the chart also reveals a subtler anomaly in injection #4: the area of the H peak is above its upper limit, while the A2 and T2 peaks are close to their averages.. 4 Interactive Charts: A Powerful New Tool for Understanding Chromatography Data
5 Column Performance Charting One of the preconfigured charts provided in Chromeleon 7.1 Chromatography Data System software is the Column Performance Chart, which plots the efficiency, asymmetry, and resolution for all the peaks in a chromatogram. These common measures of separation effectiveness can be used to make a quick assessment of how well a chromatographic column is performing, and can be used to diagnose common problems such as a poorly packed stationary phase (Figure 5). FIGURE 5. The Column Performance chart on the left shows a typical pattern for monovalent species in a separation using ion-exchange chromatography. Peak efficiency (plate count) is lowest for the first-eluted compound, which is most affected by extracolumn dead volume, then efficiency increases and then plateaus for subsequent compounds. In the chart on the right, the efficiency of analytes decreases as retention time increases, suggesting that the column bed has a packing problem or very slow mass transport.. Component Ratio Profiling In many applications, knowing the ratio of sample components can be more important than knowing their absolute concentrations. For example, ratios of various sugars, organic acids, or other components are often used in the food industry to distinguish between authentic ingredients and cheap substitutes. Pie charts (Figure 6) are very useful for showing relative proportions of sample components at a glance, and can help food chemists quickly spot adulterated ingredients. FIGURE 6. Any number of result charts can be incorporated into a report template, facilitating comparisons. In this example, pie charts provide a quick visual profile of the ratios of major antioxidants in juices. Pomegranate juice adulterated with grape juice (bottom) shows different anthocyanin ratios than pure pomegranate juice (top) note relative sizes of green D3 and red C3,5 sectors. Thermo Scientific Poster Note PN70361_e 11/12S 5
6 Estimation of Extracolumn Volume A creative use of the Interactive Chart feature is the estimation of the volume between injector and detector that is not within the chromatography column. Too much of this extracolumn volume can cause excessive peak broadening and interfere with separation. The estimation technique involves injecting a homologous series of compounds under isocratic conditions that fully resolve the peaks while maintaining low k values and small peak elution volumes. A plot of the square of the peak width vs the square of the retention time is constructed, and fitted with a linear regression (Figure 8). The extracolumn volume can be estimated as V ec = f * y o, where f is the flow rate and y o is the y-intercept of the plot, and the column efficiency can be estimated from the reciprocal of the slope. This technique has many limitations and relies on many assumptions, so discretion must be exercised when using it, but it can be a useful tool for getting a sense of the performance of a system and of a chromatography column. FIGURE 8. Extracolumn dead volume can be estimated using the intercept of a plot of the square of the peak width vs the square of the retention time for an isocratic separation of a homologous series of peaks with low k and comparable efficiency. In this example, the squares were calculated in the Interactive Results sheet (see area highlighted in black), and plotted against each other in an Interactive Chart. The legend of the linear regression overlay gives the slope and intercept of a line fit to the data points. Conclusion The interactive charting capabilities integrated into Chromeleon 7.1 Chromatography Data System software provide chemists with powerful new tools for analyzing chromatography data more efficiently. Instant graphing of results makes it easy for analysts to observe trends and spot anomalies. Dynamic interactive links with chromatograms, tabulated results, and other displays of interrelated data invite exploration and allow rapid drill-down int source data. With Interactive Charts, chromatographers can interpret results much more quickly, and better cope with the ever-increasing volumes of data they need to manage. Acknowledgements The authors would like to thank Petr Jandik, Jun Cheng, Chris Pohl, Pranathi Perati, Bob Stathers, and Mark Tracy for providing data used in the examples shown here. 6 Interactive Charts: A Powerful New Tool for Understanding Chromatography Data
7 Thermo Fisher Scientific Inc. All rights reserved. All trademarks are the property of Thermo Fisher Scientific Inc. and its subsidiaries. This information is presented as an example of the capabilities of Thermo Fisher Scientific Inc. products. It is not intended to encourage use of these products in any manners that might infringe the intellectual property rights of others. Specifications, terms and pricing are subject to change. Not all products are available in all countries. Please consult your local sales representative for details. Australia Austria Belgium Brazil China Denmark France Germany India Italy Japan Korea Netherlands Singapore Sweden Switzerland Taiwan UK/Ireland USA and Canada PN70361_E 07/16S
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