Our earlier study [J. Chromatogr. A 1218 (2011) 7765] on separation of an oligolysine mixture consisting of chains with 2-8 lysine residues (number of lysine residues, dp=2-8) by ion-pairing reversed-phase chromatography using heptafluorobutyric acid (HFBA) as an ion pairing reagent at fixed mobile phase acetonitrile (ACN) content was extended to isocratic elution conditions with different ACN percentages. The present work explored how manipulating the mobile phase HFBA concentration ([HFBA]) and %-ACN content influences separations of the oligolysine mixture. The closed pairing model was used to analyze variation of the retention factor as a function of [HFBA]. The partition coefficient of the paired peptide decreased with increasing %-ACN. Pairing of HFBA to oligolysine was cooperative, and the effect increased when %-ACN in the mobile phase was lowered. A plot of the partition coefficient as a function of %-ACN for oligolysines varying in dp converged at one ACN content, indicating a critical condition in which components of different dp co-elute.
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Comparison of retention behavior of oligolysine and oligoarginine in ion-pairing chromatography using heptafluorobutyric acid.
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Center for Biocatalysis and Bioprocessing of Macromolecules, Department of Chemical and Biological Engineering, Polytechnic Institute of New York University, Six Metrotech Center, Brooklyn, NY, 11201, USA.
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