Sci Rep. 2026 Jul 5;16(1):20571. doi: 10.1038/s41598-026-60367-3.
ABSTRACT
A validated HPTLC densitometric approach was employed for the simultaneous determination of Cefazolin (CFZ), Sulfadimidine (SDD), and Marbofloxacin (MFC) residues in milk. The method utilizes a mobile phase composed of ethyl acetate, methanol, and triethylamine in a 50:30:1 v/v/v ratio, with detection carried out at 270 nm. The chromatographic conditions were carefully optimized, and the retardation factor (Rf) values for CFZ, MFC, and SDD were 0.29, 0.58, and 0.73, respectively. Sample preparation included protein precipitation with acetonitrile, followed by dispersive solid-phase extraction (dSPE) using Enhanced Matrix Removal-Lipid (EMR-L) tubes to remove interfering lipids. Validation in accordance with ICH Q2(R2) guidelines demonstrated excellent linearity for all analytes over the concentration range of 0.001-0.09 µg/band. Accuracy was confirmed by recovery values between 98.19% and 99.15%. The proposed method was successfully applied to the determination of the investigated drug residues in real cattle milk samples following the withdrawal period, showing no statistically significant differences in accuracy and precision compared to previously reported HPLC method. Furthermore, sustainability evaluation using multiple greenness assessment tools (Eco-scale score of 77, AGSA score of 75%, BAGI score of 77.5, and RGB12 score of 85.2) demonstrated that the method is excellent green.
PMID:42402651 | DOI:10.1038/s41598-026-60367-3
