As the year 2011 winds down to a hopefully very-festive close, we wish you all the best for this holiday season and trust that 2012 will be a productive and intellectually-stimulating year. We plan to continue sending out notices to keep you apprised of technology happenings in the field of chemometrics in general and the integration of chemometric methods into instrument systems in particular.
We will focus on one particular application of chemometrics as a brief overview of relevant technology. In this issue, we outline the integration of retention time alignment such that the correction of chromatographic peak position can be done as part of a method. The technical underpinnings of the algorithm in LineUp come from the literature describing Correlation Optimized Warping, or COW, which was brought into active research by the Danish Technical University a decade ago. But, having an algorithm and using it as part of a routine process in a lab or on-line are very different topics; we focus here on the effective integration of alignment into a chromatographic instrument method.
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Automating Chromatographic Alignment
The LineUp utility that provides software-based alignment for chromatographic data has a friendly user-interface to present both alignment parameters and a view of the target and sample traces, before and after alignment. With this interface you can set up a batch of runs as well to handle a whole collection of chrom
atograms that are in need of alignment.
On the other hand, you may want to align data as soon as acquired. We have worked with chromatographic vendors to learn how best to automate the alignment process. Solutions vary with the vendor; here are snapshots of how this can be done for several of the more prevalent chromatography systems.
The EZChrom software, now owned by Agilent, allows you to specify that a user program be run immediately after data acquisition. In the Advance Methods section of the EZChrom software, you specify that you will export an AIA trace (*.CDF); choose the folder where the exported file will be stored. Then, in the Files tab, under After Export, choose our custom utility, called EZCRM2LU.exe. The command line will need a few parameters; these and other specifics can be found in the
LineUp User Guide, p. 4-6.
When the EZChrom method, which can be called by a sequence, runs, data is acquired, a CDF file is exported, then EZCRM2LU is invoked, LineUp is called, and a new, aligned CDF file is created. Afterwards, if you need to integrate and post-process the aligned chromatograms, simply create an EZChrom sequence in which you load all of the aligned CDF files, then Analyze.
The process for automating alignment with TotalChrom, from Perkin Elmer, is much like that for EZChrom. First, you set the Report Format options by instructing the software to Create AIA Metafile. Then, in the Method editor, under Process > User Programs, you choose our utility called TCRM2LU.exe and provide the appropriate
command line parameters (see p. 4-8 of the LineUp User Guide).
When a TotalChrom sequence is started, a chromatogram is acquired, a CDF file is exported, TCRM2LU is invoked which calls LineUp, and, finally, an aligned CDF file is created. To post-process the aligned file, it (they) must first be converted to RAW format. This is done with the Convert utility provided as part of the TotalChrom package.
In ChromPerfect, automation has been carried to the next level, in which the software calls LineUp directly, with no need for user programs to be invoked. In the Method editor, there is a tab that allows specification of the alignment
parameters, including the target file name.
In the Processing tab, you tell the method to Perform Alignment, then specify that the method will produce an AIA file and/or produce a bound file. The produced CDF file can be used subsequently in Pirouette or InStep while the bound file can be used for additional processing in ChromPerfect.
Chromatographic technology has improved considerably over the years, yet the need for alignment remains, particularly when chemometrics is applied to whole chromatograms, the current trend. Our alignment tool, together with your chromatography system, can insure that you will always have at hand the best chromatographic result.