Thomas Rizzo - Academia.edu (original) (raw)

Papers by Thomas Rizzo

Research paper thumbnail of State-Resolved Gas-Surface Reactivity of Methane in the Symmetric C-H Stretch Vibration on Ni(100)

Physical Review Letters, Jun 22, 2005

The state-resolved reactivity of CH 4 in its totally symmetric C-H stretch vibration (1) has been... more The state-resolved reactivity of CH 4 in its totally symmetric C-H stretch vibration (1) has been measured on a Ni(100) surface. Methane molecules were accelerated to kinetic energies of 49 and 63:5 kJ=mol in a molecular beam and vibrationally excited to 1 by stimulated Raman pumping before surface impact at normal incidence. The reactivity of the symmetric-stretch excited CH 4 is about an order of magnitude higher than that of methane excited to the antisymmetric stretch (3) reported by Juurlink et al. [Phys. Rev. Lett. 83, 868 (1999)] and is similar to that we have previously observed for the excitation of the first overtone (2 3). The difference between the state-resolved reactivity for 1 and 3 is consistent with predictions of a vibrationally adiabatic model of the methane reaction dynamics and indicates that statistical models cannot correctly describe the chemisorption of CH 4 on nickel.

Research paper thumbnail of Efficient stimulated Raman pumping for quantum state resolved surface reactivity measurements

Review of Scientific Instruments, May 1, 2006

We describe the use of stimulated Raman pumping in a molecular beam to perform quantum state reso... more We describe the use of stimulated Raman pumping in a molecular beam to perform quantum state resolved gas-surface reactivity measurements for molecules prepared in totally symmetric vibrational states. Vibrational states of homonuclear diatomics as well as totally symmetric vibrations of polyatomic molecules cannot be prepared by direct infrared excitation but are accessible through stimulated Raman pumping by two laser fields when the difference between the incident laser frequencies matches the vibration. We generate a suitable resonant pair of high-energy pump and Stokes laser beams in an injection seeded Raman amplifier filled with the sample gas and equipped with internal gas recirculation. The ability to partially saturate the Raman pumping process in the molecular beam is used to quantify the fraction of vibrationally excited molecules in the irradiated volume, which is needed for quantitative reactivity measurements. We illustrate the method with state resolved reactivity measurements for CH4, prepared in its symmetric C–H stretch vibration on a Ni(100) single crystal surface.

Research paper thumbnail of Identification of human milk oligosaccharide positional isomers by combining IMS-CID-IMS and cryogenic IR spectroscopy

The Analyst

Cryogenic infrared spectra of CID fragments separated by IMS are used to identify human milk olig... more Cryogenic infrared spectra of CID fragments separated by IMS are used to identify human milk oligiosaccharide isomers.

Research paper thumbnail of UV spectroscopic studies on cold host-guest complexes in 22-pole ion trap

Research paper thumbnail of Toward High-Throughput Cryogenic IR Fingerprinting of Mobility-Separated Glycan Isomers

ACS Measurement Science Au, 2021

Infrared (IR) spectroscopy is a powerful tool used to infer detailed structural information on mo... more Infrared (IR) spectroscopy is a powerful tool used to infer detailed structural information on molecules, often in conjunction with quantum-chemical calculations. When applied to cryogenically cooled ions, IR spectra provide unique fingerprints that can be used for biomolecular identification. This is particularly important in the analysis of isomeric biopolymers, which are difficult to distinguish using mass spectrometry. However, IR spectroscopy typically requires laser systems that need substantial user attention and measurement times of tens of minutes, which limits its analytical utility. We report here the development of a new high-throughput instrument that combines ultrahigh-resolution ionmobility spectrometry with cryogenic IR spectroscopy and mass spectrometry, and we apply it to the analysis of isomeric glycans. The ion mobility step, which is based on structures for lossless ion manipulations (SLIM), separates glycan isomers, and an IR fingerprint spectrum identifies them. An innovative cryogenic ion trap allows multiplexing the acquisition of analyte IR fingerprints following mobility separation, and using a turn-key IR laser, we can obtain spectra and identify isomeric species in less than a minute. This work demonstrates the potential of IR fingerprinting methods to impact the analysis of isomeric biomolecules and more specifically glycans.

Research paper thumbnail of The Structure of the Protonated Serine Octamer

Journal of the American Chemical Society, Jan 20, 2018

The amino acid serine has long been known to form a protonated "magic-number" cluster c... more The amino acid serine has long been known to form a protonated "magic-number" cluster containing eight monomer units that shows an unusually high abundance in mass spectra and has a remarkable homochiral preference. Despite many experimental and theoretical studies, there is no consensus on a SerH structure that is in agreement with all experimental observations. Here, we present the structure of SerH determined by a combination of infrared spectroscopy and ab initio molecular dynamics simulations. The three-dimensional structure that we determine is ∼25 kcal mol more stable than the previous most stable published structure and explains both the homochiral preference and the experimentally observed facile replacement of two serine units.

Research paper thumbnail of Cryogenic Ion Spectroscopy for Identification of Monosaccharide Anomers

The Journal of Physical Chemistry A, 2019

We combine conformer-selective, cryogenic infrared spectroscopy, quantum mechanical computations,... more We combine conformer-selective, cryogenic infrared spectroscopy, quantum mechanical computations, and 18 O substitution at the reducing end to determine the structural preferences of protonated glucosamine in the gas phase. Cryogenic infrared-infrared (IR-IR) double resonance spectroscopy of helium-tagged protonated glucosamine provides vibrational fingerprints of individual conformers, and 18 O isotopic labeling facilitates the match with computed structures and provides a selective probe of the anomeric hydroxyl. This is key for using vibrational spectroscopy for glycan analysis and determining the generality of anomeric memory during glycosidic bond cleavage.

Research paper thumbnail of UV and IR spectroscopic studies of alkali metal ion-crown ether complexes in the gas phase and on gold surface

Gordon Research Conferences –Molecular & Ionic Clusters, from April 27 to May 2, 2014, Renaissanc... more Gordon Research Conferences –Molecular & Ionic Clusters, from April 27 to May 2, 2014, Renaissance Tuscany Il Ciocco, in Lucca (Barga), Italy

Research paper thumbnail of How General Is Anomeric Retention during Collision-Induced Dissociation of Glycans?

Journal of the American Chemical Society, 2020

Despite the essential role that glycans play in many biological processes, their isomeric complex... more Despite the essential role that glycans play in many biological processes, their isomeric complexity makes their structural determination particularly challenging. Tandem mass spectrometry has played a central role in glycan analysis, and recent work has shown that fragments generated by collision-induced dissociation (CID) of disaccharides can retain the anomeric configuration of the glycosidic bond. If this result proves to be general, it would provide a powerful new tool for glycan sequencing. In this work, we use messenger-tagging infrared (IR) spectroscopy to investigate the generality of anomer retention in CID by exploring different fragmentation channels in glycans of increasing complexity. Our results demonstrate that anomericity seems to be retained irrespective of fragment size and branching.

Research paper thumbnail of Using SLIM-Based IMS-IMS Together with Cryogenic Infrared Spectroscopy for Glycan Analysis

Analytical Chemistry, 2020

The isomeric heterogeneity of glycans poses a great challenge for their analysis. While combining... more The isomeric heterogeneity of glycans poses a great challenge for their analysis. While combining ion mobility spectrometry (IMS) with tandem mass spectrometry is a powerful means for identifying and characterizing glycans, it has difficulty distinguishing the subtlest differences between isomers. Cryogenic infrared spectroscopy provides an additional dimension for glycan identification that is extremely sensitive to their structure. Our approach to glycan analysis combines ultrahigh-resolution IMS-IMS using structures for lossless ion manipulation (SLIM) with cryogenic infrared spectroscopy. We present here the design of a SLIM board containing a series of on-board traps in which we perform collision-induced dissociation (CID) at pressures in the millibar range. We characterize the on-board CID process by comparing the fragments generated from a pentapeptide to those obtained on a commercial tandem mass spectrometer. We then apply our new technique to study the mobility and vibrational spectra of CID fragments from two human milk oligosaccharides. Comparison of both the fragment drift times and IR spectra with those of suitable reference compounds allows us to identify their specific isomeric form, including the anomericity of the glycosidic linkage, demonstrating the power of this tool for glycan analysis.

Research paper thumbnail of Combining Cryogenic Infrared Spectroscopy with Selective Enzymatic Cleavage for Determining Glycan Primary Structure

Analytical Chemistry, 2020

Given the biological relevance and intrinsic structural complexity of glycans, increasing efforts... more Given the biological relevance and intrinsic structural complexity of glycans, increasing efforts are being directed toward developing a general glycan database that includes information from different analytical methods. As recently demonstrated, cryogenic infrared (IR) spectroscopy is a promising technique for glycan analysis, as it provides unique vibrational fingerprints of specific glycan isomer ions. One of the main goals of a glycan database is the identification and detailed characterization of unknown species. In this work, we combine enzymatic digestion with cryogenic IR-spectroscopy and demonstrate how it can be used for glycan identification. We measured the IR-spectra of a series of cationic glycan standards of increasing complexity and compared them with spectra of the same species after enzymatic cleavage of larger glycans. We show that the cryogenic IR spectra of the cleaved glycans are highly structured and virtually identical to those of standards after both single and multiple cleavages. Our results suggest that the combination of these methods represents a potentially powerful and specific approach for the characterization of unknown glycans.

Research paper thumbnail of Efficient stimulated Raman pumping for quantum state resolved surface reactivity measurements

Review of Scientific Instruments, 2006

We describe the use of stimulated Raman pumping in a molecular beam to perform quantum state reso... more We describe the use of stimulated Raman pumping in a molecular beam to perform quantum state resolved gas-surface reactivity measurements for molecules prepared in totally symmetric vibrational states. Vibrational states of homonuclear diatomics as well as totally symmetric vibrations of polyatomic molecules cannot be prepared by direct infrared excitation but are accessible through stimulated Raman pumping by two laser fields

Research paper thumbnail of Identifying Mixtures of Isomeric Human Milk Oligosaccharides by the Decomposition of IR Spectral Fingerprints

The analysis of glycans presents a significant challenge that arises from their isomeric heteroge... more The analysis of glycans presents a significant challenge that arises from their isomeric heterogeneity. While high-resolution ion mobility spectrometry (IMS) has shown the ability to resolve subtly different glycan isomers, their unambiguous assignment remains difficult. Here, we demonstrate an infrared (IR) spectroscopic approach for identifying isomers in a glycan mixture. To display the feasibility of this approach, we have constructed a small database of cryogenic spectra of five lacto-Nfucopentaose (LNFP) and six disaccharide isomers and demonstrated that in the cases where they cannot be separated by IMS, we can use a cryogenic IR spectrum to identify the isomeric components of a mixture.

Research paper thumbnail of Infrared Spectroscopy as a Probe of Electronic Energy Transfer

The Journal of Physical Chemistry Letters, 2018

Research paper thumbnail of Cryogenic Vibrational Spectroscopy Provides Unique Fingerprints for Glycan Identification

Journal of The American Society for Mass Spectrometry, 2017

The structural characterization of glycans by mass spectrometry is particularly challenging. This... more The structural characterization of glycans by mass spectrometry is particularly challenging. This is because of the high degree of isomerism in which glycans of the same mass can differ in their stereochemistry, attachment points, and degree of branching. Here we show that the addition of cryogenic vibrational spectroscopy to mass and mobility measurements allows one to uniquely identify and characterize these complex biopolymers. We investigate six disaccharide isomers that differ in their stereochemistry, attachment point of the glycosidic bond, and monosaccharide content, and demonstrate that we can identify each one unambiguously. Even disaccharides that differ by a single stereogenic center or in the monosaccharide sequence order show distinct vibrational fingerprints that would clearly allow their identification in a mixture, which is not possible by ion mobility spectrometry/mass spectrometry alone. Moreover, this technique can be applied to larger glycans, which we demonstrate by distinguishing isomeric branched and linear pentasaccharides. The creation of a database containing mass, collision cross section, and vibrational fingerprint measurements for glycan standards should allow unambiguous identification and characterization of these biopolymers in mixtures, providing an enabling technology for all fields of glycoscience.

Research paper thumbnail of Unravelling the structures of sodiated β-cyclodextrin and its fragments

Physical Chemistry Chemical Physics

Experiment and theory unravel the structure and formation mechanism of β-cyclodextrin fragment ions.

Research paper thumbnail of A new approach for identifying positional isomers of glycans cleaved from monoclonal antibodies

The Analyst

We apply high-resolution ion mobility combined with cryogenic infrared spectroscopy to distinguis... more We apply high-resolution ion mobility combined with cryogenic infrared spectroscopy to distinguish isomeric N-glycans with different terminal galactose positions.

Research paper thumbnail of Analyzing glycans cleaved from a biotherapeutic protein using ultrahigh-resolution ion mobility spectrometry together with cryogenic ion spectroscopy

The Analyst

Glycans covalently attached to protein biotherapeutics have a significant impact on their biologi... more Glycans covalently attached to protein biotherapeutics have a significant impact on their biological activity, clearance, and safety. As a result, glycosylation is categorized as a critical quality attribute that needs an adequate analytical approach to guarantee product quality. However, the isomeric complexity and branched structure of glycans makes their analysis a significant challenge. In this work, we propose a multidimensional approach for monitoring released glycans that combines ultrahigh-resolution ion mobility spectrometry (IMS) and cryogenic vibrational spectroscopy, and we demonstrate this technique by characterizing four N-glycans cleaved from the therapeutic fusion protein etanercept that range in abundance from 1% to 22% of the total N-glycan content. The recorded vibrational spectra exhibit wellresolved transitions that can be used as a fingerprint to identify a particular glycan. This work represents an important advance in the analysis of N-linked glycans cleaved from biopharmaceutical proteins that could eventually be used as tool for monitoring biopharmaceutical glycoforms. † Electronic supplementary information (ESI) available. See

Research paper thumbnail of ChemInform Abstract: IR Spectrum of tert.-Butyl Hydroperoxide Excited to the 4νOH Vibrational Overtone Level

Research paper thumbnail of Separation and Identification of Glycan Anomers Using Ultrahigh-Resolution Ion-Mobility Spectrometry and Cryogenic Ion Spectroscopy

Journal of The American Society for Mass Spectrometry

The analysis of carbohydrates, or glycans, is challenging for established structuresensitive gas-... more The analysis of carbohydrates, or glycans, is challenging for established structuresensitive gas-phase methods. The multitude of possible stereo-, regio-, and structural isomers makes them substantially more complex to analyze than DNA or proteins, and no one method is currently able to fully resolve them. While the combination of tandem mass spectrometry (MS) and ion-mobility spectrometry (IMS) have made important inroads in glycan analysis, in many cases, this approach is still not able to identify the precise isomeric form. To advance the techniques available for glycan analysis, we employ two important innovations. First, we perform ultrahigh-resolution mobility separation using structures for lossless ion manipulations (SLIM) for isomer separation and pre-selection. We then complement this IMS-MS stage with a cryogenic IR spectroscopic dimension since a glycan's vibrational spectrum provides a fingerprint that is extremely sensitive to the precise isomeric form. Using this unique approach in conjunction with oxygen-18 isotopic labeling, we show on a range of disaccharides how the two α and β anomers that every reducing glycan adopts in solution can be readily separated by mobility and identified based on their IR spectra. In addition to highlighting the power of our technique to detect minute differences in the structure of isomeric carbohydrates, these results provide the means to determine if and when anomericity is retained during collision-induced dissociation (CID) of larger glycans.

Research paper thumbnail of State-Resolved Gas-Surface Reactivity of Methane in the Symmetric C-H Stretch Vibration on Ni(100)

Physical Review Letters, Jun 22, 2005

The state-resolved reactivity of CH 4 in its totally symmetric C-H stretch vibration (1) has been... more The state-resolved reactivity of CH 4 in its totally symmetric C-H stretch vibration (1) has been measured on a Ni(100) surface. Methane molecules were accelerated to kinetic energies of 49 and 63:5 kJ=mol in a molecular beam and vibrationally excited to 1 by stimulated Raman pumping before surface impact at normal incidence. The reactivity of the symmetric-stretch excited CH 4 is about an order of magnitude higher than that of methane excited to the antisymmetric stretch (3) reported by Juurlink et al. [Phys. Rev. Lett. 83, 868 (1999)] and is similar to that we have previously observed for the excitation of the first overtone (2 3). The difference between the state-resolved reactivity for 1 and 3 is consistent with predictions of a vibrationally adiabatic model of the methane reaction dynamics and indicates that statistical models cannot correctly describe the chemisorption of CH 4 on nickel.

Research paper thumbnail of Efficient stimulated Raman pumping for quantum state resolved surface reactivity measurements

Review of Scientific Instruments, May 1, 2006

We describe the use of stimulated Raman pumping in a molecular beam to perform quantum state reso... more We describe the use of stimulated Raman pumping in a molecular beam to perform quantum state resolved gas-surface reactivity measurements for molecules prepared in totally symmetric vibrational states. Vibrational states of homonuclear diatomics as well as totally symmetric vibrations of polyatomic molecules cannot be prepared by direct infrared excitation but are accessible through stimulated Raman pumping by two laser fields when the difference between the incident laser frequencies matches the vibration. We generate a suitable resonant pair of high-energy pump and Stokes laser beams in an injection seeded Raman amplifier filled with the sample gas and equipped with internal gas recirculation. The ability to partially saturate the Raman pumping process in the molecular beam is used to quantify the fraction of vibrationally excited molecules in the irradiated volume, which is needed for quantitative reactivity measurements. We illustrate the method with state resolved reactivity measurements for CH4, prepared in its symmetric C–H stretch vibration on a Ni(100) single crystal surface.

Research paper thumbnail of Identification of human milk oligosaccharide positional isomers by combining IMS-CID-IMS and cryogenic IR spectroscopy

The Analyst

Cryogenic infrared spectra of CID fragments separated by IMS are used to identify human milk olig... more Cryogenic infrared spectra of CID fragments separated by IMS are used to identify human milk oligiosaccharide isomers.

Research paper thumbnail of UV spectroscopic studies on cold host-guest complexes in 22-pole ion trap

Research paper thumbnail of Toward High-Throughput Cryogenic IR Fingerprinting of Mobility-Separated Glycan Isomers

ACS Measurement Science Au, 2021

Infrared (IR) spectroscopy is a powerful tool used to infer detailed structural information on mo... more Infrared (IR) spectroscopy is a powerful tool used to infer detailed structural information on molecules, often in conjunction with quantum-chemical calculations. When applied to cryogenically cooled ions, IR spectra provide unique fingerprints that can be used for biomolecular identification. This is particularly important in the analysis of isomeric biopolymers, which are difficult to distinguish using mass spectrometry. However, IR spectroscopy typically requires laser systems that need substantial user attention and measurement times of tens of minutes, which limits its analytical utility. We report here the development of a new high-throughput instrument that combines ultrahigh-resolution ionmobility spectrometry with cryogenic IR spectroscopy and mass spectrometry, and we apply it to the analysis of isomeric glycans. The ion mobility step, which is based on structures for lossless ion manipulations (SLIM), separates glycan isomers, and an IR fingerprint spectrum identifies them. An innovative cryogenic ion trap allows multiplexing the acquisition of analyte IR fingerprints following mobility separation, and using a turn-key IR laser, we can obtain spectra and identify isomeric species in less than a minute. This work demonstrates the potential of IR fingerprinting methods to impact the analysis of isomeric biomolecules and more specifically glycans.

Research paper thumbnail of The Structure of the Protonated Serine Octamer

Journal of the American Chemical Society, Jan 20, 2018

The amino acid serine has long been known to form a protonated "magic-number" cluster c... more The amino acid serine has long been known to form a protonated "magic-number" cluster containing eight monomer units that shows an unusually high abundance in mass spectra and has a remarkable homochiral preference. Despite many experimental and theoretical studies, there is no consensus on a SerH structure that is in agreement with all experimental observations. Here, we present the structure of SerH determined by a combination of infrared spectroscopy and ab initio molecular dynamics simulations. The three-dimensional structure that we determine is ∼25 kcal mol more stable than the previous most stable published structure and explains both the homochiral preference and the experimentally observed facile replacement of two serine units.

Research paper thumbnail of Cryogenic Ion Spectroscopy for Identification of Monosaccharide Anomers

The Journal of Physical Chemistry A, 2019

We combine conformer-selective, cryogenic infrared spectroscopy, quantum mechanical computations,... more We combine conformer-selective, cryogenic infrared spectroscopy, quantum mechanical computations, and 18 O substitution at the reducing end to determine the structural preferences of protonated glucosamine in the gas phase. Cryogenic infrared-infrared (IR-IR) double resonance spectroscopy of helium-tagged protonated glucosamine provides vibrational fingerprints of individual conformers, and 18 O isotopic labeling facilitates the match with computed structures and provides a selective probe of the anomeric hydroxyl. This is key for using vibrational spectroscopy for glycan analysis and determining the generality of anomeric memory during glycosidic bond cleavage.

Research paper thumbnail of UV and IR spectroscopic studies of alkali metal ion-crown ether complexes in the gas phase and on gold surface

Gordon Research Conferences –Molecular & Ionic Clusters, from April 27 to May 2, 2014, Renaissanc... more Gordon Research Conferences –Molecular & Ionic Clusters, from April 27 to May 2, 2014, Renaissance Tuscany Il Ciocco, in Lucca (Barga), Italy

Research paper thumbnail of How General Is Anomeric Retention during Collision-Induced Dissociation of Glycans?

Journal of the American Chemical Society, 2020

Despite the essential role that glycans play in many biological processes, their isomeric complex... more Despite the essential role that glycans play in many biological processes, their isomeric complexity makes their structural determination particularly challenging. Tandem mass spectrometry has played a central role in glycan analysis, and recent work has shown that fragments generated by collision-induced dissociation (CID) of disaccharides can retain the anomeric configuration of the glycosidic bond. If this result proves to be general, it would provide a powerful new tool for glycan sequencing. In this work, we use messenger-tagging infrared (IR) spectroscopy to investigate the generality of anomer retention in CID by exploring different fragmentation channels in glycans of increasing complexity. Our results demonstrate that anomericity seems to be retained irrespective of fragment size and branching.

Research paper thumbnail of Using SLIM-Based IMS-IMS Together with Cryogenic Infrared Spectroscopy for Glycan Analysis

Analytical Chemistry, 2020

The isomeric heterogeneity of glycans poses a great challenge for their analysis. While combining... more The isomeric heterogeneity of glycans poses a great challenge for their analysis. While combining ion mobility spectrometry (IMS) with tandem mass spectrometry is a powerful means for identifying and characterizing glycans, it has difficulty distinguishing the subtlest differences between isomers. Cryogenic infrared spectroscopy provides an additional dimension for glycan identification that is extremely sensitive to their structure. Our approach to glycan analysis combines ultrahigh-resolution IMS-IMS using structures for lossless ion manipulation (SLIM) with cryogenic infrared spectroscopy. We present here the design of a SLIM board containing a series of on-board traps in which we perform collision-induced dissociation (CID) at pressures in the millibar range. We characterize the on-board CID process by comparing the fragments generated from a pentapeptide to those obtained on a commercial tandem mass spectrometer. We then apply our new technique to study the mobility and vibrational spectra of CID fragments from two human milk oligosaccharides. Comparison of both the fragment drift times and IR spectra with those of suitable reference compounds allows us to identify their specific isomeric form, including the anomericity of the glycosidic linkage, demonstrating the power of this tool for glycan analysis.

Research paper thumbnail of Combining Cryogenic Infrared Spectroscopy with Selective Enzymatic Cleavage for Determining Glycan Primary Structure

Analytical Chemistry, 2020

Given the biological relevance and intrinsic structural complexity of glycans, increasing efforts... more Given the biological relevance and intrinsic structural complexity of glycans, increasing efforts are being directed toward developing a general glycan database that includes information from different analytical methods. As recently demonstrated, cryogenic infrared (IR) spectroscopy is a promising technique for glycan analysis, as it provides unique vibrational fingerprints of specific glycan isomer ions. One of the main goals of a glycan database is the identification and detailed characterization of unknown species. In this work, we combine enzymatic digestion with cryogenic IR-spectroscopy and demonstrate how it can be used for glycan identification. We measured the IR-spectra of a series of cationic glycan standards of increasing complexity and compared them with spectra of the same species after enzymatic cleavage of larger glycans. We show that the cryogenic IR spectra of the cleaved glycans are highly structured and virtually identical to those of standards after both single and multiple cleavages. Our results suggest that the combination of these methods represents a potentially powerful and specific approach for the characterization of unknown glycans.

Research paper thumbnail of Efficient stimulated Raman pumping for quantum state resolved surface reactivity measurements

Review of Scientific Instruments, 2006

We describe the use of stimulated Raman pumping in a molecular beam to perform quantum state reso... more We describe the use of stimulated Raman pumping in a molecular beam to perform quantum state resolved gas-surface reactivity measurements for molecules prepared in totally symmetric vibrational states. Vibrational states of homonuclear diatomics as well as totally symmetric vibrations of polyatomic molecules cannot be prepared by direct infrared excitation but are accessible through stimulated Raman pumping by two laser fields

Research paper thumbnail of Identifying Mixtures of Isomeric Human Milk Oligosaccharides by the Decomposition of IR Spectral Fingerprints

The analysis of glycans presents a significant challenge that arises from their isomeric heteroge... more The analysis of glycans presents a significant challenge that arises from their isomeric heterogeneity. While high-resolution ion mobility spectrometry (IMS) has shown the ability to resolve subtly different glycan isomers, their unambiguous assignment remains difficult. Here, we demonstrate an infrared (IR) spectroscopic approach for identifying isomers in a glycan mixture. To display the feasibility of this approach, we have constructed a small database of cryogenic spectra of five lacto-Nfucopentaose (LNFP) and six disaccharide isomers and demonstrated that in the cases where they cannot be separated by IMS, we can use a cryogenic IR spectrum to identify the isomeric components of a mixture.

Research paper thumbnail of Infrared Spectroscopy as a Probe of Electronic Energy Transfer

The Journal of Physical Chemistry Letters, 2018

Research paper thumbnail of Cryogenic Vibrational Spectroscopy Provides Unique Fingerprints for Glycan Identification

Journal of The American Society for Mass Spectrometry, 2017

The structural characterization of glycans by mass spectrometry is particularly challenging. This... more The structural characterization of glycans by mass spectrometry is particularly challenging. This is because of the high degree of isomerism in which glycans of the same mass can differ in their stereochemistry, attachment points, and degree of branching. Here we show that the addition of cryogenic vibrational spectroscopy to mass and mobility measurements allows one to uniquely identify and characterize these complex biopolymers. We investigate six disaccharide isomers that differ in their stereochemistry, attachment point of the glycosidic bond, and monosaccharide content, and demonstrate that we can identify each one unambiguously. Even disaccharides that differ by a single stereogenic center or in the monosaccharide sequence order show distinct vibrational fingerprints that would clearly allow their identification in a mixture, which is not possible by ion mobility spectrometry/mass spectrometry alone. Moreover, this technique can be applied to larger glycans, which we demonstrate by distinguishing isomeric branched and linear pentasaccharides. The creation of a database containing mass, collision cross section, and vibrational fingerprint measurements for glycan standards should allow unambiguous identification and characterization of these biopolymers in mixtures, providing an enabling technology for all fields of glycoscience.

Research paper thumbnail of Unravelling the structures of sodiated β-cyclodextrin and its fragments

Physical Chemistry Chemical Physics

Experiment and theory unravel the structure and formation mechanism of β-cyclodextrin fragment ions.

Research paper thumbnail of A new approach for identifying positional isomers of glycans cleaved from monoclonal antibodies

The Analyst

We apply high-resolution ion mobility combined with cryogenic infrared spectroscopy to distinguis... more We apply high-resolution ion mobility combined with cryogenic infrared spectroscopy to distinguish isomeric N-glycans with different terminal galactose positions.

Research paper thumbnail of Analyzing glycans cleaved from a biotherapeutic protein using ultrahigh-resolution ion mobility spectrometry together with cryogenic ion spectroscopy

The Analyst

Glycans covalently attached to protein biotherapeutics have a significant impact on their biologi... more Glycans covalently attached to protein biotherapeutics have a significant impact on their biological activity, clearance, and safety. As a result, glycosylation is categorized as a critical quality attribute that needs an adequate analytical approach to guarantee product quality. However, the isomeric complexity and branched structure of glycans makes their analysis a significant challenge. In this work, we propose a multidimensional approach for monitoring released glycans that combines ultrahigh-resolution ion mobility spectrometry (IMS) and cryogenic vibrational spectroscopy, and we demonstrate this technique by characterizing four N-glycans cleaved from the therapeutic fusion protein etanercept that range in abundance from 1% to 22% of the total N-glycan content. The recorded vibrational spectra exhibit wellresolved transitions that can be used as a fingerprint to identify a particular glycan. This work represents an important advance in the analysis of N-linked glycans cleaved from biopharmaceutical proteins that could eventually be used as tool for monitoring biopharmaceutical glycoforms. † Electronic supplementary information (ESI) available. See

Research paper thumbnail of ChemInform Abstract: IR Spectrum of tert.-Butyl Hydroperoxide Excited to the 4νOH Vibrational Overtone Level

Research paper thumbnail of Separation and Identification of Glycan Anomers Using Ultrahigh-Resolution Ion-Mobility Spectrometry and Cryogenic Ion Spectroscopy

Journal of The American Society for Mass Spectrometry

The analysis of carbohydrates, or glycans, is challenging for established structuresensitive gas-... more The analysis of carbohydrates, or glycans, is challenging for established structuresensitive gas-phase methods. The multitude of possible stereo-, regio-, and structural isomers makes them substantially more complex to analyze than DNA or proteins, and no one method is currently able to fully resolve them. While the combination of tandem mass spectrometry (MS) and ion-mobility spectrometry (IMS) have made important inroads in glycan analysis, in many cases, this approach is still not able to identify the precise isomeric form. To advance the techniques available for glycan analysis, we employ two important innovations. First, we perform ultrahigh-resolution mobility separation using structures for lossless ion manipulations (SLIM) for isomer separation and pre-selection. We then complement this IMS-MS stage with a cryogenic IR spectroscopic dimension since a glycan's vibrational spectrum provides a fingerprint that is extremely sensitive to the precise isomeric form. Using this unique approach in conjunction with oxygen-18 isotopic labeling, we show on a range of disaccharides how the two α and β anomers that every reducing glycan adopts in solution can be readily separated by mobility and identified based on their IR spectra. In addition to highlighting the power of our technique to detect minute differences in the structure of isomeric carbohydrates, these results provide the means to determine if and when anomericity is retained during collision-induced dissociation (CID) of larger glycans.