The benefits of mixed-mode chromatography columns for separation of peptides and protein digests

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00216208%3A11310%2F23%3A10470986" target="_blank" >RIV/00216208:11310/23:10470986 - isvavai.cz</a>

Výsledek na webu

<a href="https://verso.is.cuni.cz/pub/verso.fpl?fname=obd_publikace_handle&handle=VQbxkS7061" target="_blank" >https://verso.is.cuni.cz/pub/verso.fpl?fname=obd_publikace_handle&handle=VQbxkS7061</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1007/s00706-023-03088-x" target="_blank" >10.1007/s00706-023-03088-x</a>

Jazyk výsledku

angličtina

Název v původním jazyce

The benefits of mixed-mode chromatography columns for separation of peptides and protein digests

Popis výsledku v původním jazyce

In this work, the evaluation and comparison of mixed-mode chromatography and reversed-phase chromatography for separation of peptides and protein digests have been performed. The effects of pH of aqueous part of mobile phase as well as the effects of organic modifier on retention, resolution, and peak shape were investigated on several columns including three mixed-mode columns possessing reversed-phase/anion-exchange mechanism, two reversed-phase octadecyl columns, and one column with mixed-mode reversed-phase/anion-exchange character only in defined pH range. The set of peptides varying in their polarity, length, amino acid sequence, and charge state, namely dipeptides, N-blocked dipeptides, and oligopeptides, was selected to describe the chromatographic behavior under different conditions properly. These measurements showed the potential of mixed-mode chromatography columns for analysis of differently charged peptides in a single run. The applicability of the tested conditions has been verified by the analysis of cytochrome C digested fragments. Two types of samples were analyzed and compared, i.e., commercial cytochrome C digested standard and cytochrome C digested via trypsin spin columns. The obtained results point to the necessity of using mass spectrometry detection because of large number of unknown peaks in cytochrome C digested standard, probably originating from chymotryptic and miscleavage activities.

Název v anglickém jazyce

The benefits of mixed-mode chromatography columns for separation of peptides and protein digests

Popis výsledku anglicky

In this work, the evaluation and comparison of mixed-mode chromatography and reversed-phase chromatography for separation of peptides and protein digests have been performed. The effects of pH of aqueous part of mobile phase as well as the effects of organic modifier on retention, resolution, and peak shape were investigated on several columns including three mixed-mode columns possessing reversed-phase/anion-exchange mechanism, two reversed-phase octadecyl columns, and one column with mixed-mode reversed-phase/anion-exchange character only in defined pH range. The set of peptides varying in their polarity, length, amino acid sequence, and charge state, namely dipeptides, N-blocked dipeptides, and oligopeptides, was selected to describe the chromatographic behavior under different conditions properly. These measurements showed the potential of mixed-mode chromatography columns for analysis of differently charged peptides in a single run. The applicability of the tested conditions has been verified by the analysis of cytochrome C digested fragments. Two types of samples were analyzed and compared, i.e., commercial cytochrome C digested standard and cytochrome C digested via trypsin spin columns. The obtained results point to the necessity of using mass spectrometry detection because of large number of unknown peaks in cytochrome C digested standard, probably originating from chymotryptic and miscleavage activities.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10403 - Physical chemistry

Projekt

<a href="/cs/project/GA20-19655S" target="_blank" >GA20-19655S: Strategie pro on-line štěpení proteinů s následnou separací v mix-mode chromatografii</a><br>

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2023

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Monatshefte für Chemie - Chemical Monthly

ISSN

0026-9247

e-ISSN

1434-4475

Svazek periodika

154

Číslo periodika v rámci svazku

9

Stát vydavatele periodika

AT - Rakouská republika

Počet stran výsledku

10

Strana od-do

993-1002

Kód UT WoS článku

001004557200007

EID výsledku v databázi Scopus

2-s2.0-85161436504