Synthesis and application of BODIPY-based fluorescent labeling
tag for oligosaccharide and N-linked glycan analysis by
high-performance liquid chromatography with fluorescence
detection

J 2024

Synthesis and application of BODIPY-based fluorescent labeling tag for oligosaccharide and N-linked glycan analysis by high-performance liquid chromatography with fluorescence detection

SMOLKOVÁ, Denisa, Michal GREGUŠ, Hubert VESELÝ, Richard CMELIK, Hana PÍŽOVÁ et. al.

Basic information

Original name

Synthesis and application of BODIPY-based fluorescent labeling tag for oligosaccharide and N-linked glycan analysis by high-performance liquid chromatography with fluorescence detection

Authors

SMOLKOVÁ, Denisa (703 Slovakia, belonging to the institution), Michal GREGUŠ (703 Slovakia, belonging to the institution), Hubert VESELÝ (203 Czech Republic, belonging to the institution), Richard CMELIK (203 Czech Republic), Hana PÍŽOVÁ (203 Czech Republic, belonging to the institution), Pavel BOBÁĽ (703 Slovakia, belonging to the institution) and Jana LAVICKA (203 Czech Republic)

Edition

Analytica Chimica Acta, Amsterdam, Elsevier Science publishers, 2024, 0003-2670

Other information

Language

English

Type of outcome

Článek v odborném periodiku

Field of Study

10400 1.4 Chemical sciences

Country of publisher

Netherlands

Confidentiality degree

není předmětem státního či obchodního tajemství

References:

URL

Impact factor

Impact factor: 6.200 in 2022

Organization unit

Faculty of Pharmacy

DOI

http://dx.doi.org/10.1016/j.aca.2023.342032

UT WoS

001166247500001

Keywords in English

Oligosaccharides; Glycans; Labeling; BODIPY; Liquid chromatography; Fluorescence

Abstract

V originále

Background: Glycosylation analysis is still challenging, not only because of the extreme structure complexity and conjugation diversity of glycans but also because of instrumental aspects such as the sensitivity limits of analyses. Therefore, glycan analysis by chromatographic methods is very often combined with fluorescence detection in addition to MS. The majority of fluorescent labeling employed before LC separation is based on 2-aminobenzamide, which has several disadvantages such as low labeling yield, poor fluorescence properties, and MS ionization efficiency. Therefore, even after several decades of development of new labels, there is still a need for new labeling tags with improved characteristics. Results: We present the application of a newly synthesized fluorescent label designed for oligosaccharide and glycan analysis by high-performance liquid chromatography with fluorescence detection (HPLC/FLD). The novel hydrazide derivative of dipyrrometheneboron difluoride (BODIPY) was synthesized from 2,4-dimethylpyrrole, methyl succinyl chloride, and boron trifluoride etherate followed by a reaction with hydrazine. The synthesized label was characterized by several analytical methods including NMR, UV/Vis and fluorescence spectroscopy, and mass spectrometry. The labeling reaction via hydrazone formation chemistry was optimized by labeling of maltooligosaccharide standards. The analysis of maltohexaose labeled by BODIPY-hydrazide followed by HPLC/ FLD analysis provided the limit of detection in the low tens of femtomole. The presented method based on fluorescence detection is at least 30 times more sensitive than the standard approach employing labeling by 2aminobenzamide. In addition, the labeling method by BODIPY-hydrazide was used for N -linked glycan profiling of several glycoproteins (ribonuclease B, immunoglobulin G) by RP-HPLC/FLD as well as HILIC/FLD analysis. Significance: This work represents the design, synthesis, and application of a new fluorescent label based on the BODIPY core and hydrazone formation chemistry for oligosaccharide and glycan analysis by HPLC/FLD. The proposed approach significantly improved the oligosaccharide and glycan analysis in comparison to the commonly used procedure employing 2-aminobenzamide.

Links

GA22-00236S, research and development project

Name: Vývoj nových fluorescenčních značek pro elektroforetické analýzy glykoproteinů asociovaných s rakovinou prsu (Acronym: 22-00236S)

Investor: Czech Science Foundation

MUNI/A/1232/2021, interní kód MU

Name: Design a syntéza nových fluoroforů pro detekci biomolekul

Investor: Masaryk University

Citovat

SMOLKOVÁ, Denisa, Michal GREGUŠ, Hubert VESELÝ, Richard CMELIK, Hana PÍŽOVÁ, Pavel BOBÁĽ and Jana LAVICKA. Synthesis and application of BODIPY-based fluorescent labeling tag for oligosaccharide and N-linked glycan analysis by high-performance liquid chromatography with fluorescence detection. Analytica Chimica Acta. Amsterdam: Elsevier Science publishers, 2024, vol. 1285, No 342032, p. 1-8. ISSN 0003-2670. Available from: https://dx.doi.org/10.1016/j.aca.2023.342032.

@article{2380465,
   author = {Smolková, Denisa and Greguš, Michal and Veselý, Hubert and Cmelik, Richard and Pížová, Hana and Bobáľ, Pavel and Lavicka, Jana},
   article_location = {Amsterdam},
   article_number = {342032},
   doi = {http://dx.doi.org/10.1016/j.aca.2023.342032},
   keywords = {Oligosaccharides; Glycans; Labeling; BODIPY; Liquid chromatography; Fluorescence},
   language = {eng},
   issn = {0003-2670},
   journal = {Analytica Chimica Acta},
   title = {Synthesis and application of BODIPY-based fluorescent labeling tag for oligosaccharide and N-linked glycan analysis by high-performance liquid chromatography with fluorescence detection},
   url = {https://pdf.sciencedirectassets.com/271374/1-s2.0-S0003267023X00464/1-s2.0-S0003267023012539/main.pdf?X-Amz-Security-Token=IQoJb3JpZ2luX2VjEFkaCXVzLWVhc3QtMSJHMEUCIQCfgifaC8tRpXFB%2F4tVDJ4386lwIOmxH38Q0ElbRrTFTQIgaEba7F3qlS8%2ByHAwZVa4RfNShZVFB%2B12U%2BX},
   volume = {1285},
   year = {2024}
}

TY  - JOUR
ID  - 2380465
AU  - Smolková, Denisa - Greguš, Michal - Veselý, Hubert - Cmelik, Richard - Pížová, Hana - Bobáľ, Pavel - Lavicka, Jana
PY  - 2024
TI  - Synthesis and application of BODIPY-based fluorescent labeling tag for oligosaccharide and N-linked glycan analysis by high-performance liquid chromatography with fluorescence detection
JF  - Analytica Chimica Acta
VL  - 1285
IS  - 342032
SP  - 1-8
EP  - 1-8
PB  - Elsevier Science publishers
SN  - 00032670
KW  - Oligosaccharides
KW  - Glycans
KW  - Labeling
KW  - BODIPY
KW  - Liquid chromatography
KW  - Fluorescence
UR  - https://pdf.sciencedirectassets.com/271374/1-s2.0-S0003267023X00464/1-s2.0-S0003267023012539/main.pdf?X-Amz-Security-Token=IQoJb3JpZ2luX2VjEFkaCXVzLWVhc3QtMSJHMEUCIQCfgifaC8tRpXFB%2F4tVDJ4386lwIOmxH38Q0ElbRrTFTQIgaEba7F3qlS8%2ByHAwZVa4RfNShZVFB%2B12U%2BX
N2  - Background: Glycosylation analysis is still challenging, not only because of the extreme structure complexity and conjugation diversity of glycans but also because of instrumental aspects such as the sensitivity limits of analyses. Therefore, glycan analysis by chromatographic methods is very often combined with fluorescence detection in addition to MS. The majority of fluorescent labeling employed before LC separation is based on 2-aminobenzamide, which has several disadvantages such as low labeling yield, poor fluorescence properties, and MS ionization efficiency. Therefore, even after several decades of development of new labels, there is still a need for new labeling tags with improved characteristics. Results: We present the application of a newly synthesized fluorescent label designed for oligosaccharide and glycan analysis by high-performance liquid chromatography with fluorescence detection (HPLC/FLD). The novel hydrazide derivative of dipyrrometheneboron difluoride (BODIPY) was synthesized from 2,4-dimethylpyrrole, methyl succinyl chloride, and boron trifluoride etherate followed by a reaction with hydrazine. The synthesized label was characterized by several analytical methods including NMR, UV/Vis and fluorescence spectroscopy, and mass spectrometry. The labeling reaction via hydrazone formation chemistry was optimized by labeling of maltooligosaccharide standards. The analysis of maltohexaose labeled by BODIPY-hydrazide followed by HPLC/ FLD analysis provided the limit of detection in the low tens of femtomole. The presented method based on fluorescence detection is at least 30 times more sensitive than the standard approach employing labeling by 2aminobenzamide. In addition, the labeling method by BODIPY-hydrazide was used for N -linked glycan profiling of several glycoproteins (ribonuclease B, immunoglobulin G) by RP-HPLC/FLD as well as HILIC/FLD analysis. Significance: This work represents the design, synthesis, and application of a new fluorescent label based on the BODIPY core and hydrazone formation chemistry for oligosaccharide and glycan analysis by HPLC/FLD. The proposed approach significantly improved the oligosaccharide and glycan analysis in comparison to the commonly used procedure employing 2-aminobenzamide.
ER  -

SMOLKOVÁ, Denisa, Michal GREGUŠ, Hubert VESELÝ, Richard CMELIK, Hana PÍŽOVÁ, Pavel BOBÁĽ and Jana LAVICKA. Synthesis and application of BODIPY-based fluorescent labeling tag for oligosaccharide and N-linked glycan analysis by high-performance liquid chromatography with fluorescence detection. \textit{Analytica Chimica Acta}. Amsterdam: Elsevier Science publishers, 2024, vol.~1285, No~342032, p.~1-8. ISSN~0003-2670. Available from: https://dx.doi.org/10.1016/j.aca.2023.342032.

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