Elucidation of chromatographic peak shifts in complex samples using a chemometrical approach

Pedro F M Sousa,K Magnus Åberg,Angela De Waard

doi:10.1007/s00216-018-1173-9

Abstract

Chromatographic retention time peak shifts between consecutive analyses is a well-known fact yet not fully understood. Algorithms have been developed to align peaks between runs, but with no specific studies considering the causes of peak shifts. Here, designed experiments reveal chromatographic shift patterns for a complex peptide mixture that are attributable to the temperature and pH of the mobile phase. These results demonstrate that peak shifts are highly structured and are to a high degree explained by underlying differences in physico-chemical parameters of the chromatographic system and also provide experimental support for the alignment algorithm called the generalized fuzzy Hough transform which exploits this fact. It can be expected that the development of alignment algorithms enters a new phase resulting in increasingly accurate alignment by considering the latent structure of the peak shifts.

Highlights

Chromatographic retention mechanisms have been studied and modeled since the 1970s
In the field of quantitative structure–retention relationships (QSRR), the focus has been on predicting parameters, such as log k-values, log P values, log D-values, and retention factor ratios, which are based on molecular descriptors [1,2,3,4]
The research focused on solving the correspondence problem has only aimed to make sure the data are properly aligned for statistical analysis, and several retention time alignment algorithms have been reported in literature, such as nearest-neighbor clustering [13, 14], binning [15, 16], and warping [17,18,19,20]

Summary

Introduction

Chromatographic retention mechanisms have been studied and modeled since the 1970s. In the field of quantitative structure–retention relationships (QSRR), the focus has been on predicting parameters, such as log k-values, log P values, log D-values, and retention factor ratios, which are based on molecular descriptors [1,2,3,4]. This work fuses alignment methodology with retention time modeling to improve the understanding of chromatographic retention in complex systems For this purpose, a liquid chromatography coupled with mass spectrometry (LC/MS) method for analyzing a complex mixture was setup using an experimental design, with temperature of the column and pH of the mobile phase as variables provoking retention time shifts. Along with the theoretical peptides, other species may occur in the mixture, such as possible miscleavages and post-translational modifications in the protein This moderate sample complexity provides a good model for this study, as the peptide elution is well distributed along the chromatogram and the experimental design conditions tested, i.e., pH of the mobile phase and column temperature, influence the retention times with different patterns depending on the peptide. The ASCA algorithm for MATLAB was obtained from literature [24]

Results and discussion

Conclusions

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