Beyond Fourier Transform Infrared Spectroscopy: External Cavity Quantum Cascade Laser-Based Mid-infrared Transmission Spectroscopy of Proteins in the Amide I and Amide II Region

In this work, we present a setup for mid-IR measurements of the protein amide I and amide II bands in aqueous solution. Employing a latest generation external cavity-quantum cascade laser (EC-QCL) at room temperature in pulsed operation mode allowed implementing a high optical path length of 31 μm t...

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Bibliographic Details
Published in:Analytical chemistry (Washington) 2018-06, Vol.90 (11), p.7072-7079
Main Authors: Schwaighofer, Andreas, Montemurro, Milagros, Freitag, Stephan, Kristament, Christian, Culzoni, María J, Lendl, Bernhard
Format: Article
Language:English
Subjects:
Analytical chemistry
Aqueous solutions
Chemistry
Chemometrics
Data processing
Fourier transforms
Infrared imaging systems
Infrared lasers
Infrared spectrometers
Infrared spectroscopy
Laser applications
Mathematical analysis
Matrix methods
Noise levels
Optical paths
Proteins
Quantum cascade lasers
Robustness (mathematics)
Spectra
Spectrometers
Spectrum analysis
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Summary:In this work, we present a setup for mid-IR measurements of the protein amide I and amide II bands in aqueous solution. Employing a latest generation external cavity-quantum cascade laser (EC-QCL) at room temperature in pulsed operation mode allowed implementing a high optical path length of 31 μm that ensures robust sample handling. By application of a data processing routine, which removes occasionally deviating EC-QCL scans, the noise level could be lowered by a factor of 4. The thereby accomplished signal-to-noise ratio is better by a factor of approximately 2 compared to research-grade Fourier transform infrared (FT-IR) spectrometers at equal acquisition times. Employing this setup, characteristic spectral features of three representative proteins with different secondary structures could be measured at concentrations as low as 1 mg mL–1. Mathematical evaluation of the spectral overlap confirms excellent agreement of the quantum cascade laser infrared spectroscropy (QCL-IR) transmission measurements with protein spectra acquired by FT-IR spectroscopy. The presented setup combines performance surpassing FT-IR spectroscopy with large applicable optical paths and coverage of the relevant spectral range for protein analysis. This holds high potential for future EC-QCL-based protein studies, including the investigation of dynamic secondary structure changes and chemometrics-based protein quantification in complex matrices.
ISSN:0003-2700
1520-6882
DOI:10.1021/acs.analchem.8b01632