Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography

Autores
Akhgar, Christopher K.; Ebner, Julian; Alcaraz, Mirta R.; Kopp, Julian; Goicoechea, Hector Casimiro; Spadiut, Oliver; Schwaighofer, Andreas; Lendl, Bernhard
Año de publicación
2022
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
An external-cavity quantum cascade laser (EC-QCL)-based flow-through mid-infrared (IR) spectrometer was placed in line with a preparative size exclusion chromatography system to demonstrate real-time analysis of protein elutions with strongly overlapping chromatographic peaks. Two different case studies involving three and four model proteins were performed under typical lab-scale purification conditions. The large optical path length (25 μm), high signal-to-noise ratios, and wide spectral coverage (1350 to 1750 cm-1) of the QCL-IR spectrometer allow for robust spectra acquisition across both the amide I and II bands. Chemometric analysis by self-modeling mixture analysis and multivariate curve resolution enabled accurate quantitation and structural fingerprinting across the protein elution transient. The acquired concentration profiles were found to be in excellent agreement with the off-line high-performance liquid chromatography reference analytics performed on the collected effluent fractions. These results demonstrate that QCL-IR detectors can be used effectively for in-line, real-time analysis of protein elutions, providing critical quality attribute data that are typically only accessible through time-consuming and resource-intensive off-line methods.
Fil: Akhgar, Christopher K.. Vienna University of Technology; Austria
Fil: Ebner, Julian. Vienna University of Technology; Austria
Fil: Alcaraz, Mirta R.. Institute Of Chemical Technologies And Analytics; Austria
Fil: Kopp, Julian. Vienna University of Technology; Austria
Fil: Goicoechea, Hector Casimiro. Universidad Nacional del Litoral; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
Fil: Spadiut, Oliver. Vienna University of Technology; Austria
Fil: Schwaighofer, Andreas. Vienna University of Technology; Austria
Fil: Lendl, Bernhard. Vienna University of Technology; Austria
Materia
Amides
Size exclusion chromatography
Chemometrics
,Chromatography
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/217029
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/217029
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion ChromatographyAkhgar, Christopher K.Ebner, JulianAlcaraz, Mirta R.Kopp, JulianGoicoechea, Hector CasimiroSpadiut, OliverSchwaighofer, AndreasLendl, BernhardAmidesSize exclusion chromatographyChemometrics,Chromatographyhttps://purl.org/becyt/ford/1.4https://purl.org/becyt/ford/1An external-cavity quantum cascade laser (EC-QCL)-based flow-through mid-infrared (IR) spectrometer was placed in line with a preparative size exclusion chromatography system to demonstrate real-time analysis of protein elutions with strongly overlapping chromatographic peaks. Two different case studies involving three and four model proteins were performed under typical lab-scale purification conditions. The large optical path length (25 μm), high signal-to-noise ratios, and wide spectral coverage (1350 to 1750 cm-1) of the QCL-IR spectrometer allow for robust spectra acquisition across both the amide I and II bands. Chemometric analysis by self-modeling mixture analysis and multivariate curve resolution enabled accurate quantitation and structural fingerprinting across the protein elution transient. The acquired concentration profiles were found to be in excellent agreement with the off-line high-performance liquid chromatography reference analytics performed on the collected effluent fractions. These results demonstrate that QCL-IR detectors can be used effectively for in-line, real-time analysis of protein elutions, providing critical quality attribute data that are typically only accessible through time-consuming and resource-intensive off-line methods.Fil: Akhgar, Christopher K.. Vienna University of Technology; AustriaFil: Ebner, Julian. Vienna University of Technology; AustriaFil: Alcaraz, Mirta R.. Institute Of Chemical Technologies And Analytics; AustriaFil: Kopp, Julian. Vienna University of Technology; AustriaFil: Goicoechea, Hector Casimiro. Universidad Nacional del Litoral; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Spadiut, Oliver. Vienna University of Technology; AustriaFil: Schwaighofer, Andreas. Vienna University of Technology; AustriaFil: Lendl, Bernhard. Vienna University of Technology; AustriaAmerican Chemical Society2022-08info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/217029Akhgar, Christopher K.; Ebner, Julian; Alcaraz, Mirta R.; Kopp, Julian; Goicoechea, Hector Casimiro; et al.; Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography; American Chemical Society; Analytical Chemistry; 94; 32; 8-2022; 11192-112000003-2700CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://pubs.acs.org/doi/10.1021/acs.analchem.2c01542info:eu-repo/semantics/altIdentifier/doi/10.1021/acs.analchem.2c01542info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:46:01Zoai:ri.conicet.gov.ar:11336/217029instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 09:46:01.952CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography
title Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography
spellingShingle Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography
Akhgar, Christopher K.
Amides
Size exclusion chromatography
Chemometrics
,Chromatography
title_short Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography
title_full Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography
title_fullStr Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography
title_full_unstemmed Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography
title_sort Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography
dc.creator.none.fl_str_mv Akhgar, Christopher K.
Ebner, Julian
Alcaraz, Mirta R.
Kopp, Julian
Goicoechea, Hector Casimiro
Spadiut, Oliver
Schwaighofer, Andreas
Lendl, Bernhard
author Akhgar, Christopher K.
author_facet Akhgar, Christopher K.
Ebner, Julian
Alcaraz, Mirta R.
Kopp, Julian
Goicoechea, Hector Casimiro
Spadiut, Oliver
Schwaighofer, Andreas
Lendl, Bernhard
author_role author
author2 Ebner, Julian
Alcaraz, Mirta R.
Kopp, Julian
Goicoechea, Hector Casimiro
Spadiut, Oliver
Schwaighofer, Andreas
Lendl, Bernhard
author2_role author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Amides
Size exclusion chromatography
Chemometrics
,Chromatography
topic Amides
Size exclusion chromatography
Chemometrics
,Chromatography
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.4
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv An external-cavity quantum cascade laser (EC-QCL)-based flow-through mid-infrared (IR) spectrometer was placed in line with a preparative size exclusion chromatography system to demonstrate real-time analysis of protein elutions with strongly overlapping chromatographic peaks. Two different case studies involving three and four model proteins were performed under typical lab-scale purification conditions. The large optical path length (25 μm), high signal-to-noise ratios, and wide spectral coverage (1350 to 1750 cm-1) of the QCL-IR spectrometer allow for robust spectra acquisition across both the amide I and II bands. Chemometric analysis by self-modeling mixture analysis and multivariate curve resolution enabled accurate quantitation and structural fingerprinting across the protein elution transient. The acquired concentration profiles were found to be in excellent agreement with the off-line high-performance liquid chromatography reference analytics performed on the collected effluent fractions. These results demonstrate that QCL-IR detectors can be used effectively for in-line, real-time analysis of protein elutions, providing critical quality attribute data that are typically only accessible through time-consuming and resource-intensive off-line methods.
Fil: Akhgar, Christopher K.. Vienna University of Technology; Austria
Fil: Ebner, Julian. Vienna University of Technology; Austria
Fil: Alcaraz, Mirta R.. Institute Of Chemical Technologies And Analytics; Austria
Fil: Kopp, Julian. Vienna University of Technology; Austria
Fil: Goicoechea, Hector Casimiro. Universidad Nacional del Litoral; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina
Fil: Spadiut, Oliver. Vienna University of Technology; Austria
Fil: Schwaighofer, Andreas. Vienna University of Technology; Austria
Fil: Lendl, Bernhard. Vienna University of Technology; Austria
description An external-cavity quantum cascade laser (EC-QCL)-based flow-through mid-infrared (IR) spectrometer was placed in line with a preparative size exclusion chromatography system to demonstrate real-time analysis of protein elutions with strongly overlapping chromatographic peaks. Two different case studies involving three and four model proteins were performed under typical lab-scale purification conditions. The large optical path length (25 μm), high signal-to-noise ratios, and wide spectral coverage (1350 to 1750 cm-1) of the QCL-IR spectrometer allow for robust spectra acquisition across both the amide I and II bands. Chemometric analysis by self-modeling mixture analysis and multivariate curve resolution enabled accurate quantitation and structural fingerprinting across the protein elution transient. The acquired concentration profiles were found to be in excellent agreement with the off-line high-performance liquid chromatography reference analytics performed on the collected effluent fractions. These results demonstrate that QCL-IR detectors can be used effectively for in-line, real-time analysis of protein elutions, providing critical quality attribute data that are typically only accessible through time-consuming and resource-intensive off-line methods.
publishDate 2022
dc.date.none.fl_str_mv 2022-08
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/217029
Akhgar, Christopher K.; Ebner, Julian; Alcaraz, Mirta R.; Kopp, Julian; Goicoechea, Hector Casimiro; et al.; Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography; American Chemical Society; Analytical Chemistry; 94; 32; 8-2022; 11192-11200
0003-2700
CONICET Digital
CONICET
url http://hdl.handle.net/11336/217029
identifier_str_mv Akhgar, Christopher K.; Ebner, Julian; Alcaraz, Mirta R.; Kopp, Julian; Goicoechea, Hector Casimiro; et al.; Application of Quantum Cascade Laser-Infrared Spectroscopy and Chemometrics for In-Line Discrimination of Coeluting Proteins from Preparative Size Exclusion Chromatography; American Chemical Society; Analytical Chemistry; 94; 32; 8-2022; 11192-11200
0003-2700
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/https://pubs.acs.org/doi/10.1021/acs.analchem.2c01542
info:eu-repo/semantics/altIdentifier/doi/10.1021/acs.analchem.2c01542
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
dc.publisher.none.fl_str_mv American Chemical Society
publisher.none.fl_str_mv American Chemical Society
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
_version_ 1844613438499717120
score 13.070432

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