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
- Institución
- Consejo Nacional de Investigaciones Científicas y Técnicas
- OAI Identificador
- oai:ri.conicet.gov.ar:11336/217029
Ver los metadatos del registro completo
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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 Técnicas2025-09-03T09:50:41Zoai: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:34982025-09-03 09:50:42.251CONICET 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 |
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1842269048281235456 |
score |
13.13397 |