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

作者：Schwaighofer Andreas; Montemurro Milagros; Freitag Stephan; Kristament Christian; Culzoni Maria J; Lendl Bernhard^*

来源：Analytical Chemistry, 2018, 90(11): 7072-7079.

DOI：10.1021/acs.analchem.8601632

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 pm 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 ECQCL-based protein studies, including the investigation of dynamic secondary structure changes and chemometrics-based protein quantification in complex matrices.

• 出版日期2018-6-5