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Chromatography with PowerLab

Posted: 15 Apr 2002     Print Version  Bookmark and Share



AN104A 1 February 1998 Chromatography with PowerLab All types of chromatography and capillary electrophoresis data can be recorded with PowerLab and Chart and then processed with Peaks. Paul Duckworth, ADInstruments. Pty Ltd. Introduction PowerLab is as easy to connect and operate as a traditional strip chart recorder but your data will be displayed and stored on a computer. All chromatographs with recorder or analogue integrator outputs can be connected. Which PowerLab? Usually a PowerLab/e or PowerLab/s is chosen (depending on the number of detectors). Note however that the PowerLab/400e hardware is capable of 16 bit resolution but the /200 is limited to 12 bits (1 part in 4096 or 0.024%). Currently Chart and Scope software only utilise 12 bits, but a future software upgrade will enable the PowerLab to achieve true 16 bit resolution (1 part in 65536 or 0.0015%). The high performance PowerLab/4s and /8s are also upgradable to 16 bits when software allows. Remember, however, that the signal/noise ratio of most detectors is less than 16 bits and often less than 12 bits. Sensitivity Some chromatographs offer both a chart recorder output (usually 0-1 V or 0-10 V full scale) and an integrator output (usually 0-10 mV full scale). If you have a choice, always use the higher gain chart recorder output as this will minimise noise. Each channel of a PowerLab unit has twelve gain settings from 10 V to 2 mV, full scale, in 10:5:2 steps. It is possible to split the incoming signal between two (or more) of the the PowerLab channels and to record the data at more than one gain setting. On the higher gain channel large peaks will simply be truncated while small peaks that are lost in the baseline of the low gain channel will be easily visible. If required, the data analysis program Peaks can combine data collected at different gains to give a single trace of enhanced dynamic range. Sampling Rate This will depend on whether you are using GG, HPLC, Capillary Electrophoresis, etc., and also on your particular method. PowerLab units are capable of sampling between 12 points per hour to 1000 points per second direct to hard disk or to RAM. (Up to 10000 points per second with a PowerLab/4s or /8s). Most chromatography requires sampling between 10 points per second to 1 point every few seconds - well within the capability of PowerLab. As a general rule try to have a minimum of 10 points across the width of the narrowest peak i.e. if your narrowest peak is 8 seconds wide the you should collect your data at a rate of 48 or more points per minute. This would be achieved with a Chart setting of 100 points per minute. This will provide well-defined peak shapes and give accurate integration. For chromatography it is recommended that when using Chart you select data buffering to hard disk, Figure 3. This ensures that Chart's automatic failsafe system is on, and that even in the event of a power failure you will lose at most the last 2 minutes of data. Integration Online integration is possible with Chart. The results appear on a second channel as an integral trace - similar to an integrating pen recorder. If your detector does not have a "zero" control you may need to consider passing the signal though a PowerLab GP Amp (general purpose amplifier) so that a DC offset can be applied to zero the signal. In most cases integration can be best achieved with a data analysis program, for example Peaks. Peaks software can accept Chart or ASCII text files, and performs smoothing, baseline subtraction, integration, one point calibration, application of response factors, and use of calibration files. Specialist data processing can be done within the program Igor which offers very extensive data analysis features, and very high quality output with full control over line thickness, axis tick marks, etc. ApplicationNote Physical Science ADInstruments AN104A 2 February 1998 Units Conversion The signal from some detectors (eg. UV-visible photometric detectors) can be calibrated in "native units" with the Units Conversion feature of Chart. For example if your HPLC photometric detector outputs 1 V for an absorbance reading of 2 then enter the appropriate values in the Units Conversion dialogue, Figure 3. Multiple Runs Recording can be initiated in Chart from an external trigger signal or by simply clicking the start button, Figure 2. Unlimited number of chromatograms can be stored in the one file, and each one is automatically time stamped. Annotation The Notebook feature allows relevant observations and conditions to be recorded in the data file. The Comments facility lets the user add brief notes during or after a run, say to identify a particular peak, or the injection point. There is no need to use a separate laboratory notebook. Data Display A selection of the main chromatograph can be expanded in the Zoom Window, Figure 2. A movable cursor gives interactive readout of signal and time values. Data can be stored as a compact Chart file, cut and pasted via the Clipboard, or saved as an ASCII text file. Peaks is ideal for analysing data with Chart on a PowerLab system. A report table can be prepared that contains peak areas and heights, normalised areas (or heights), data is easily expanded, and can be printed on a laser printer, Figure 4. Peaks can combine data collected at different gains on different PowerLab channels. In this way the resulting chromatogram can have a greatly enhanced dynamic range. Figure 1. Gas chromatogram of the pyrolysis gasses from a coal sample, recorded at 4 samples per second. The peaks are due to hydrocarbons of increasing molecular weight. The pointer is tracked by a cursor on the trace. A graticle is provided on the screen for easy estimates of coordinates Time and signal readings for the trace cursor are provided The time axis has been shrunk by 20:1 TheYaxiscanbe shiftedandstretched A slider bar lets you scan through the data The start/stop button commences and halts data acquisition Different colours can be selected for the trace and the graticle AN104A 3 February 1998 A variety of functions can be selected for display. Units Conversion allows you to alter the units on the Y axis. Figure 2. Gas chromatogram shown in Figure 1 with a selected area expanded in the Zoom Window to show the fine detail. Computed Inputs include the ability to do on line integration as well as many other functions. Figure 3. Some of the menus and features of Chart. It is usual to record chromatography data directly to the hard disk. AN104A 4 February 1998 Figure 4. Chromatogram in Figure 1 imported into Peaks. The baseline has been corrected, the chromatogram trimmed and peaks numbered and their areas and heights determined and displayed in the Report Window, at left. Peak areas have also been reported normalised to the largest peak. Many other options are available. Data in the Report Window can be copied to spreadsheet and word processor documents for the easy generation of reports. Trademarks MacLab and PowerLab are registered trademarks, and Chart and Scope are trademarks, of ADInstruments Pty Ltd. Other trademarks are the properties of their respective owners. Addresses International ADInstruments Pty Ltd Unit 6, 4 Gladstone Road Castle Hill, NSW 2154 AUSTRALIA Phone:+61 (2) 9899 5455 Fax:+61 (2) 9899 5847 Email:[email protected] Web: North America ADInstruments 1949 Landings Dr Mountain View CA 94043 U.S.A. Phone:+1 (650) 965 9292 Fax: +1 (650) 965 9293 Email: [email protected] Europe ADInstruments Ltd Grove House Grove Road, Hastings East Sussex, TN35 4JS UNITED KINGDOM Phone: +44 (1424) 424 342 Fax: +44 (1424) 460 303 Email:enquiries Japan ADInstruments Japan Inc. Okajima Bldg 2-10-1 Iwamoto-cho Chiyoda-ku, 101 Tokyo JAPAN Phone:+81 (3) 5820 7556 Fax:+81 (3) 3861 7022 Email:[email protected] Your local distributor: Copyright. All rights reserved

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