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J Chromatogr A. 2017 Aug 18;1511:1-8. doi: 10.1016/j.chroma.2017.06.076. Epub 2017 Jul 3.

Sample-independent approach to normalize two-dimensional data for orthogonality evaluation using whole separation space scaling.

Author information

1
Laboratory of Metabolomics, Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacký University Olomouc, Hněvotínská 5, 779 00 Olomouc, Czech Republic; Department of Clinical Chemistry, University Hospital Olomouc, I. P. Pavlova 6, 779 00 Olomouc, Czech Republic.
2
Laboratory of Metabolomics, Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacký University Olomouc, Hněvotínská 5, 779 00 Olomouc, Czech Republic; Department of Clinical Chemistry, University Hospital Olomouc, I. P. Pavlova 6, 779 00 Olomouc, Czech Republic. Electronic address: david.friedecky@gmail.com.
3
School of Mechanical Engineering, Georgia Institute of Technology, 801 Ferst Dr. NW, 30313 Atlanta, United States.

Abstract

Orthogonality is a key parameter that is used to evaluate the separation power of chromatography-based two-dimensional systems. It is necessary to scale the separation data before the assessment of the orthogonality. Current scaling approaches are sample-dependent, and the extent of the retention space that is converted into a normalized retention space is set according to the retention times of the first and last analytes contained in a unique sample to elute. The presence or absence of a highly retained analyte in a sample can thus significantly influence the amount of information (in terms of the total amount of separation space) contained in the normalized retention space considered for the calculation of the orthogonality. We propose a Whole Separation Space Scaling (WOSEL) approach that accounts for the whole separation space delineated by the analytical method, and not the sample. This approach enables an orthogonality-based evaluation of the efficiency of the analytical system that is independent of the sample selected. The WOSEL method was compared to two currently used orthogonality approaches through the evaluation of in silico-generated chromatograms and real separations of human biofluids and petroleum samples. WOSEL exhibits sample-to-sample stability values of 3.8% on real samples, compared to 7.0% and 10.1% for the two other methods, respectively. Using real analyses, we also demonstrate that some previously developed approaches can provide misleading conclusions on the overall orthogonality of a two-dimensional chromatographic system.

KEYWORDS:

2D separations; Normalization; Orthogonality; Sample-independency; Scaling; Separation efficacy

PMID:
28693825
DOI:
10.1016/j.chroma.2017.06.076
[Indexed for MEDLINE]

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