Investigations into bioavailability and micelle-assisted spectroscopic quantification of brodifacoum in natural samples†

The ability of a pseudo-surfactant, aqueous humic acid (HA), to significantly enhance the solubility of hydrophobic brodifacoum (BDF) – a second-generation anticoagulant – and to enable micelle assisted spectroscopic quantification of BDF is explored. The solubilization efficiency of aqueous HA for BDF was determined to be 2.3000 ± 0.0010 mg ppm⁻¹. The potential impact of the increased bioavailability of BDF on human health was monitored by studying the interaction of BDF with Bovine Serum Albumin (BSA) protein. There is a strong affinity between BDF and BSA, as evidenced by the binding constant (K) and the number of binding sites (n), which were found to be 391 000 ± 3.03 L mol⁻¹ and 1, respectively. Additionally, fluorescence lifetime studies were conducted that showed that BDF interacted with the hydrophobic domain IIA of BSA. These findings underscore the need for a simple and sensitive analytical technique for effectively quantifying BDF. A micelle assisted fluorescence-based quantification tool for BDF is introduced, demonstrating good fidelity and excellent analytical figures of merits. The micellar system is characterized using Dynamic Light Scattering studies and confocal microscopy (size ∼270 nm). The number of molecules per micelle is calculated using Poisson's distribution, and it is found to be no more than one (<0.2%). Encouragingly, in this study, micelle encapsulation was found to significantly enhance the fluorescence of BDF, allowing for its quantification at the nanomolar level. The analytical figures of merit such as the limit of detection (LOD) and limit of quantification (LOQ), were found to be 62 nM and 208 nM, respectively, with very good linearity (R² = 0.994). The fidelity of the micelle assisted fluorescence quantification method was verified with high performance liquid chromatography using spiked natural water samples of varying HA content and the percentage recovery and Root Mean Square Error of Prediction (RMSEP) value were found to be about 100 and 0.1001, respectively. The low RMSEP indicates the potential utility of micelle assisted fluorescence quantification in developing sensitive assays for detecting BDF.