Separation of isomeric glycans by ion mobility spectrometry – the impact of fluorescent labelling

Manz, Christian; Grabarics, Márkó; Hoberg, Friederike; Pugini, Michele; Stuckmann, Alexandra; Struwe, Weston B.; Pagel, Kevin

doi:10.1039/c9an00937j

Cited by 30 publications

(31 citation statements)

References 49 publications

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“…To create the dextran calibration ladder, a 1:1 mix of dextran 5000 and dextran 1000 was dissolved in 20 μL water. Afterwards, both the released glycans and the dextran mix were labeled with procainamide according to established protocols [ 7 , 14 ]. The labeled glycans were purified with the Discovery Glycan SPE tubes according to vendor’s instruction, dried via SpeedVac (Thermo Fisher Scientific, Waltham, USA) and further suspended in 50 μL water before storing them in the HPLC autosampler at 4 °C.…”

Section: Methodsmentioning

confidence: 99%

“…Due to its importance, monitoring of N -glycosylation is a crucial requirement in the characterization of therapeutic antibodies [ 2 ] and also gained increased attention to identify potential biomarkers for cancer and other diseases [ 3 – 5 ]. The characterization of N -glycans usually requires the cleavage of the glycans from the protein and a subsequent reducing end modification with a fluorescent label [ 6 , 7 ]. The following analysis is often based on liquid chromatography (LC) in combination with fluorescence (FLD) or mass spectrometry (MS) detection [ 8 , 9 ].…”

Section: Introductionmentioning

confidence: 99%

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Dextran as internal calibrant for N-glycan analysis by liquid chromatography coupled to ion mobility-mass spectrometry

et al. 2022

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LC–MS is one of the most important tools for the comprehensive characterization of N-glycans. Despite many efforts to speed up glycan analysis via optimized sample preparation (e.g., faster enzyme digestion in combination with instant or rapid labeling dyes), a major bottleneck remains the rather long measurement times of HILIC chromatography. Further complication arises from the necessity to concomitantly calibrate with an external standard to allow for accurate retention times and the conversion into more robust GU values. Here we demonstrate the use of an internal calibration strategy for HILIC chromatography to speed up glycan analysis. By reducing the number of utilized dextran oligosaccharides, the calibrant can be spiked directly into the sample such that external calibration runs are no longer required. The minimized dextran ladder shows accurate GU calibration with a minor deviation of well below 1% and can be applied without modifications in sample preparation or data processing. We further demonstrate the simultaneous use of the minimized dextran ladder as calibrant for the estimation of CCS values in traveling wave ion mobility spectrometry. In both cases, the minimized dextran ladder enables the measurement of calibrant and sample in a single HPLC run without losing information or accuracy. Graphical abstract

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Dextran as internal calibrant for N-glycan analysis by liquid chromatography coupled to ion mobility-mass spectrometry

et al. 2022

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“…IMS, often hyphenated with front-end analytical techniques of chromatography and coupled with MS, has been widely known for its application on isomers/isobars separation. In cases of isomeric compounds that are not separated in the chromatographic dimension and not distinguished by MS, ion mobility offers an additional dimension of separation − while amplifying peak capacity . IMS-based methodologies in the field of food authenticity are still unexploited, with a limited number of studies being reported so far. − These studies are mainly focused on targeted or semitargeted approaches, with untargeted implementation being hampered by the lack of integrated workflows to deal with such complex and large data sets.…”

Section: Introductionmentioning

confidence: 99%

Trapped Ion Mobility Incorporated in LC–HRMS Workflows as an Integral Analytical Platform of High Sensitivity: Targeted and Untargeted 4D-Metabolomics in Extra Virgin Olive Oil

Drakopoulou

Damalas

Baessmann

et al. 2021

J. Agric. Food Chem.

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Trapped ion mobility spectrometry (TIMS) is a promising technique for the separation of isomers based on their mobility. In the present work, TIMS coupled to liquid chromatography (LC) and high-resolution mass spectrometry (HRMS) was applied as a comprehensive analytical platform to address authenticity challenges, focusing on extra virgin olive oil (EVOO). Isomers detected in EVOO's phenolic fraction, classified into secoiridoids group, were successfully separated. Thanks to parallel accumulation serial fragmentation (PASEF) acquisition mode, high-quality spectra were obtained, facilitating identification. Moreover, a four-dimensional (4D) untargeted metabolomics approach was implemented to evaluate EVOO's global profile in cases of both variety and geographical origin discrimination. Potential authenticity markers, attributed to isomers, were successfully identified through the proposed workflow that incorporates ion mobility information along with LC−HRMS analytical evidence (i.e., mass accuracy, retention time, isotopic pattern, MS/MS fragmentation). Our study establishes LC−TIMS−HRMS in food authenticity and highlights mobility-enhanced metabolomics in four dimensions.

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“…Moreover, the molecular features of polysaccharide digests sometimes are not obviously different for the recognition and differentiation of polysaccharide species. 16,17 Therefore, a simple, effective, and efficient analytical method that can produce fingerprint spectra with characteristic signals is needed for polysaccharides' quality control and product evaluation. Direct analysis of intact polysaccharide using MS is difficult because there are no suitable ionization methods.…”

Section: Introductionmentioning

confidence: 99%

Rapid fingerprint analysis for herbal polysaccharides using direct analysis in real‐time ionization mass spectrometry

Wang

Jiang

Liu

et al. 2021

Rapid Comm Mass Spectrometry

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Herbal polysaccharides have various potential medicinal values.Development of reliable analytical method for the fingerprint analysis of polysaccharides is critical for their quality assessment, origin identification, and authenticity evaluation.Methods: Mechanochemical extraction (MCE) was used to extract polysaccharide components from different herbal species. Intact polysaccharides were then directly analyzed by direct analysis in real-time mass spectrometry (DART-MS). Standard addition method with isotope-labeled internal standard was used to quantify polysaccharide amounts directly from liquid extract. Multivariate data analysis was further conducted for species classification.Results: The intact and large polysaccharides were decomposed into small fragment ions less than m/z 350 instantaneously using DART ion source. Different polysaccharides showed distinguished fingerprint DART-MS spectra using both individual and mixed herbal species. The liquid supernatant from MCE was validated to be used as direct sample for DART-MS analysis. Quantitation was successfully achieved for polysaccharide contents in Dendrobium officinale from different locations.Conclusions: A rapid fingerprint protocol in combination of MCE and DART-MS for herbal polysaccharides was developed. The whole process could be accomplished within a few minutes, from raw materials to final spectra, without requirements of pre-digestion and additional sample purification.

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Separation of isomeric glycans by ion mobility spectrometry – the impact of fluorescent labelling

Abstract: Bloodgroup oligosaccharides have been derivatized with labels common in HPLC and evaluated regarding their ion mobility behaviour.

Cited by 30 publications

References 49 publications

Dextran as internal calibrant for N-glycan analysis by liquid chromatography coupled to ion mobility-mass spectrometry

Dextran as internal calibrant for N-glycan analysis by liquid chromatography coupled to ion mobility-mass spectrometry

Trapped Ion Mobility Incorporated in LC–HRMS Workflows as an Integral Analytical Platform of High Sensitivity: Targeted and Untargeted 4D-Metabolomics in Extra Virgin Olive Oil

Rapid fingerprint analysis for herbal polysaccharides using direct analysis in real‐time ionization mass spectrometry

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