Combination of HPLC and SERS detection applied to the analysis of the trace content of amoxicillin in milk

Abstract

The systematic consumption of food contaminated with antibiotics leads to emergence of resistant forms of microorganisms and also causes dysbiosis. Therefore, it is very important to ensure the necessary control of the residues of these contaminants in food by using fast and reliable methods. A simple and low-cost method for determining the trace content of amoxicillin in milk based on surface enhanced Raman spectroscopy (SERS) has been developed. To remove interfering components, a simple and fast "one-tube" method is proposed. Subsequent chromatographic purification makes it possible to almost completely eliminate the matrix effect of the sample, which ensures high accuracy and specificity of the SERS analysis. The conditions for chromatographic separation were selected to avoid the sorption of amoxicillin by free silanol groups of the stationary phase of the chromatographic column (C18), as well as to immediately carry out SERS analysis without intermediate steps. A calibration model was developed using the PLS chemometric algorithm, which further improved the accuracy and reproducibility of the analysis (R2 =0.9957 for the range 0.1–1.0 µM, RMSEC = 0.031 µM, RMSEP = 0.017 µM). This method allows to determine the content of amoxicillin in milk in the range of 1.1–11.0 µM and is characterized by good repeatability (RSD) ≤ 5.5% and a low limit of quantitation = 0.19 µM, which is considerably lower compared to the UV-HPLC method. Verification of the proposed method on samples of UHT (Ultra-High Temperature processed) milk of various fat content (1.8%, 2.5% and 3.2%) showed the recovery value = 100.6–105.7%.