Evaluating C18 stationary phases with a positively charged surface for proteomic LC-MS applications using mobile phase acidified with reduced formic acid concentration

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00216208%3A11160%2F24%3A10486531" target="_blank" >RIV/00216208:11160/24:10486531 - isvavai.cz</a>

Výsledek na webu

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

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1016/j.chroma.2024.465142" target="_blank" >10.1016/j.chroma.2024.465142</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Evaluating C18 stationary phases with a positively charged surface for proteomic LC-MS applications using mobile phase acidified with reduced formic acid concentration

Popis výsledku v původním jazyce

We have recently demonstrated the ability of a C18 stationary phase with a positively charged surface (PCS-C18) to provide superior chromatographic separation of peptides using mobile phase acidified with a mere 0.01 % formic acid, significantly improving MS sensitivity. Here, we examined three columns packed with different PCS-C18 phases using the MS-favorable mobile phase acidified with low formic acid concentrations to establish the impact of separation performance and better MS sensitivity on peptide identifications. The surface charge interaction was evaluated using the retention of nitrate. The highest interaction was observed for the AdvanceBio Peptide Plus column. A surface charge-dependent shift in the retention time of peptides was confirmed with a change in formic acid concentration in the mobile phase. The separation performance of the columns with MS-favorable mobile phase acidified with low concentrations of formic acid was evaluated using well-characterized peptides. The loading capacity was assessed using a basic peptide with three lysine residues. Good chromatographic peak shapes and high loading capacity were observed for the Acquity Premier CSH C18 column, even when using a mobile phase acidified with 0.01 % formic acid. The extent of improvement in peptide identification when using reduced formic acid concentration was evaluated by analyzing the tryptic digest of trastuzumab and tryptic digest of whole bacteria cell lysate. Each column provided improved MS signal intensity and peptide identification when using the mobile phase with 0.01 % formic acid. The ability of the Acquity Premier CSH C18 column to provide better separation of peptides, even with a reduced formic acid concentration in the mobile phase, boosted MS signal intensity by 65 % and increased the number of identified peptides from the bacterial sample by 19 %. Our study confirms that significant improvement in the proteomic outputs can be achieved without additional costs only by tailoring the chemistry of the stationary phase to the composition of the mobile phase. Our results can help researchers understand the retention mechanism of peptides on the PCS-C18 stationary phases using low-ionic strength mobile phases and, more importantly, select the best-suited stationary phases for their LC-MS proteomic applications.

Název v anglickém jazyce

Evaluating C18 stationary phases with a positively charged surface for proteomic LC-MS applications using mobile phase acidified with reduced formic acid concentration

Popis výsledku anglicky

We have recently demonstrated the ability of a C18 stationary phase with a positively charged surface (PCS-C18) to provide superior chromatographic separation of peptides using mobile phase acidified with a mere 0.01 % formic acid, significantly improving MS sensitivity. Here, we examined three columns packed with different PCS-C18 phases using the MS-favorable mobile phase acidified with low formic acid concentrations to establish the impact of separation performance and better MS sensitivity on peptide identifications. The surface charge interaction was evaluated using the retention of nitrate. The highest interaction was observed for the AdvanceBio Peptide Plus column. A surface charge-dependent shift in the retention time of peptides was confirmed with a change in formic acid concentration in the mobile phase. The separation performance of the columns with MS-favorable mobile phase acidified with low concentrations of formic acid was evaluated using well-characterized peptides. The loading capacity was assessed using a basic peptide with three lysine residues. Good chromatographic peak shapes and high loading capacity were observed for the Acquity Premier CSH C18 column, even when using a mobile phase acidified with 0.01 % formic acid. The extent of improvement in peptide identification when using reduced formic acid concentration was evaluated by analyzing the tryptic digest of trastuzumab and tryptic digest of whole bacteria cell lysate. Each column provided improved MS signal intensity and peptide identification when using the mobile phase with 0.01 % formic acid. The ability of the Acquity Premier CSH C18 column to provide better separation of peptides, even with a reduced formic acid concentration in the mobile phase, boosted MS signal intensity by 65 % and increased the number of identified peptides from the bacterial sample by 19 %. Our study confirms that significant improvement in the proteomic outputs can be achieved without additional costs only by tailoring the chemistry of the stationary phase to the composition of the mobile phase. Our results can help researchers understand the retention mechanism of peptides on the PCS-C18 stationary phases using low-ionic strength mobile phases and, more importantly, select the best-suited stationary phases for their LC-MS proteomic applications.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

30104 - Pharmacology and pharmacy

Projekt

—

Návaznosti

S - Specificky vyzkum na vysokych skolach<br>I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2024

Kód důvěrnosti údajů

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

Název periodika

Journal of Chromatography A

ISSN

0021-9673

e-ISSN

1873-3778

Svazek periodika

1730

Číslo periodika v rámci svazku

August

Stát vydavatele periodika

NL - Nizozemsko

Počet stran výsledku

9

Strana od-do

465142

Kód UT WoS článku

001271294900001

EID výsledku v databázi Scopus

2-s2.0-85198279627