Recent advancements of EC-QCL based mid-IR transmission spectroscopy of proteins and application to analysis of bovine milk1

Abstract

BACKGROUND: High emission powers of external cavity-quantum cascade laser (EC-QCL) light sources allow to employ significantly larger path lengths for infrared (IR) transmission measurements compared to conventional Fourier-transform infrared (FTIR) measurements employing thermal emitters. OBJECTIVE: An EC-QCL based IR transmission setup is presented as a viable alternative for analysis of proteins in both, academic protein structure studies as well as in process analytical applications. Here, the application of EC-QCL based IR transmission spectroscopy is introduced for i) monitoring of the protein secondary structure and ii) rapid screening of the thermal history of commercial milk samples without prior sample preparation. METHODS: Proteins present in milk were measured by QCL-IR and FTIR spectroscopy and spectra were compared. Dynamic conformational changes were followed by QCL-IR spectroscopy after chemical denaturation. Sixteen commercial milk samples were surveyed by QCL-IR spectroscopy and classified according to the experienced heat load during processing. RESULTS: The 4-5 times higher applicable transmission path length (38 μm for QCL-IR vs. 8 μm for FTIR measurements) allows robust measurements of the protein amide I band in aqueous solutions. It was shown that IR spectra of the protein amide I band acquired by EC-QCL transmission spectroscopy are comparable to FTIR spectra and the acquired spectra were employed for the study of conformational changes in protein standard solutions. Furthermore, a classification analysis of commercial bovine milk samples based on their thermal history was accomplished. CONCLUSIONS: The potential application of EC-QCL IR spectroscopy was demonstrated as a tool for following conformational changes of the secondary protein structure as well as for fast screening to estimate the heat load applied to commercial milk.

pH titration of β-lactoglobulin monitored by laser-based Mid-IR transmission spectroscopy coupled to chemometric analysis

“…EC-QCL-based IR transmission measurements have been successfully accomplished for the analysis of protein secondary structure [13][14][15]. Furthermore, the feasibility of protein discrimination and quantitation in commercial bovine milk samples has been demonstrated by QCL-IR spectroscopy and evaluation of the amide I band using partial least squares (PLS) modelling [16][17][18][19]. Recently, an EC-QCL-based IR transmission setup was introduced for the analysis of the protein amide I and II regions, which favourably competes with FTIR spectroscopy regarding the signal to noise ratio at equal data acquisition times [20].…”

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

“…13 Ten years ago, a new type of these mid-IR lasers, external-cavity QCL (EC-QCLs) became commercially available, which combine high emission powers with spectral tuning ranges of several hundred wavenumbers. The high available emission powers enabled to employ 4-5 times larger path lengths for transmission 2 measurements of glucose [14][15][16] and proteins [17][18][19][20][21][22] than achievable with conventional FT-IR spectrometers. Furthermore, EC-QCLs were employed for broadband IR transmission measurements to study protein dynamics by fast kinetic experiments at high temporal resolution.…”

“…Ten years ago, a new type of these mid-IR lasers, external-cavity QCLs (EC-QCLs) became commercially available, which combine high emission powers with spectral tuning ranges of several hundred wavenumbers. The high available emission powers enabled to employ 4–5 times larger path lengths for transmission measurements of glucose − and proteins − than achievable with conventional FT-IR spectrometers.…”

Application of Vibrational Spectroscopy and Imaging to Point-of-Care Medicine: A Review

“…This is a somewhat underdeveloped area in the field of spectroscopy but offers rich opportunity and critical need as subtle changes in the spectra are increasing analyzed by complex mathematical algorithms and instruments with exceptional performance to measure very small changes are becoming increasingly available. 169,[211][212][213] Chemometric Techniques for Point-of-Care Applications…”