A hydrogen bonding based SERS method for direct label-free L-hydroxyproline detection.

[Display omitted] • This paper presents a sensitive and label-free SERS method that requires no pre-derivatization and adjustment of pH for L-Hyp detection. • The mechanism of this work provides inspiration for detecting molecules with small Raman cross-section. • The proposed SERS method has a low LOD of 8.18 ng/mL in aqueous solution and 0.13 μg/mL in milk. The detection of non-protein nitrogen adulterants is a major challenge in dairy testing. As a marker molecule of animal hydrolyzed protein, the presence of non-edible L-hydroxyproline (L-Hyp) molecules can be used to identify low-quality milk containing components of animal hydrolyzed protein. However, it is still difficult to detect L-Hyp directly in milk. The Ag@COF-COOH substrate in this paper can be used to realize label-free L-Hyp sensitive detection based on the hydrogen bond transition mechanism. To explore the mechanism, the binding sites of hydrogen bond interaction have been verified experimentally and computationally, and the charge transfer process was also explained in terms of HOMO/LOMO energy level. In conclusion, the quantitative models for L-Hyp in an aqueous environment and in milk were developed. The limit of detection (LOD) of L-Hyp in an aqueous environment could reach 8.18 ng/mL, with R2 of 0.982. The linear range of quantitative detection in milk was 0.5–1000 μg/mL and the LOD was as low as 0.13 μg/mL. In this work, a hydrogen bond interaction based Surface-enhanced Raman spectroscopy (SERS) method for the label-free detection of L-Hyp was proposed, which complemented the application of SERS technology in the detection of dairy products. [ABSTRACT FROM AUTHOR]