Laser-based mid-IR spectroscopy as a rapid analytical technique for the quantification of human milk proteins

Human milk (HM) provides essential nutrients for infant growth and development. When own mother's milk is unavailable, donor human milk (DHM) is used and typically pasteurized, which may alter protein integrity. This study presents the use of external cavity-quantum cascade laser (EC-QCL) mid-infrared spectroscopy, as a rapid, non-destructive technique, to quantify both individual and total proteins in 276 HM and DHM samples. The method showed high accuracy (≥84% explained variance) and strong correlation with reference techniques. Pasteurization significantly reduced α-lactalbumin (24%) and lactoferrin (14%), while casein and total protein declined slightly (3%). Protein levels decreased notably during early lactation, then stabilized. In the cohort of the present study, no statistically significant differences in protein content were observed between preterm and term milk. EC-QCL spectroscopy also differentiated raw and pasteurized DHM. These results support EC-QCL as a reliable tool for clinical use and quality control, with implications for improving processing methods to preserve the nutritional and immunological integrity of DHM.