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In-line Fourier-transform infrared spectroscopy as a versatile process analytical technology for preparative protein chromatography

Großhans, Steffen 1; Rüdt, Matthias 1; Sanden, Adrian 1; Brestrich, Nina 1; Morgenstern, Josefine 1; Heissler, Stefan 2; Hubbuch, Jürgen ORCID iD icon 1
1 Institut für Bio- und Lebensmitteltechnik (BLT), Karlsruher Institut für Technologie (KIT)
2 Institut für Funktionelle Grenzflächen (IFG), Karlsruher Institut für Technologie (KIT)


Fourier-transform infrared spectroscopy (FTIR) is a well-established spectroscopic method in the analysis of small molecules and protein secondary structure. However, FTIR is not commonly applied for in-line monitoring of protein chromatography. Here, the potential of in-line FTIR as a process analytical technology (PAT) in downstream processing was investigated in three case studies addressing the limits of currently applied spectroscopic PAT methods. A first case study exploited the secondary structural differences of monoclonal antibodies (mAbs) and lysozyme to selectively quantify the two proteins with partial least squares regression (PLS) giving root mean square errors of cross validation (RMSECV) of 2.42 g/l and 1.67 g/l, respectively. The corresponding Q$^{2}$ values are 0.92 and, respectively, 0.99, indicating robust models in the calibration range. Second, a process separating lysozyme and PEGylated lysozyme species was monitored giving an estimate of the PEGylation degree of currently eluting species with RMSECV of 2.35 g/l for lysozyme and 1.24 g/l for PEG with Q$^{2}$ of 0.96 and 0.94, respectively. Finally, Triton X-100 was added to a feed of lysozyme as a typical process-related impurity. ... mehr

Verlagsausgabe §
DOI: 10.5445/IR/1000083430
Veröffentlicht am 06.01.2022
DOI: 10.1016/j.chroma.2018.03.005
Zitationen: 48
Web of Science
Zitationen: 44
Zitationen: 48
Cover der Publikation
Zugehörige Institution(en) am KIT Institut für Bio- und Lebensmitteltechnik (BLT)
Institut für Funktionelle Grenzflächen (IFG)
Publikationstyp Zeitschriftenaufsatz
Publikationsmonat/-jahr 04.2018
Sprache Englisch
Identifikator ISSN: 0021-9673, 1873-3778
KITopen-ID: 1000083430
HGF-Programm 47.02.06 (POF III, LK 01) Zellpopul.auf Biofunk.Oberflächen IFG
Erschienen in Journal of chromatography / A
Verlag Elsevier
Band 1547
Seiten 37–44
Projektinformation nextBioPharmDSP (EU, H2020, 635557)
Vorab online veröffentlicht am 05.03.2018
Schlagwörter Chromatography, Proteins, Process analytical technology (PAT), Fourier-transform infrared spectroscopy (FTIR), Downstream processing
Nachgewiesen in Scopus
Web of Science
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