Extraction and determination of aflatoxins in rich-lipidic matrices using deep eutectic solvent as the extraction medium and UHPLC-FLD analysis

Since their discovery in the 1960s, aflatoxins have presented a significant public health concern and posed a serious challenge to food quality control. Their carcinogenic potential and widespread occurrence in various food matrices, including nuts, maize, rice, and dairy products, have prompted regulatory authorities to establish maximum allowable levels for these contaminants in these commodities. Official methods traditionally rely on methanol/water or acetonitrile/water solutions as extraction media due to the slightly polar nature of aflatoxins. Pistachio samples, in particular, present a complex matrix rich in lipids, requiring a critical sample preparation step before analysis. Usually, to reduce the presence of lipidic interferents from the matrix, a de-fatting step using hexane is mandatory, in the present study it was not necessary. We introduce a novel method utilizing deep eutectic solvents (DES) as the extraction medium. DES, a new class of green solvents formed by combining a hydrogen bonding donor and acceptor in an appropriate molar ratio, yields a clear solution with a melting point significantly lower than that of its constituents. Previous studies have demonstrated the non-toxic, biodegradable nature of most DESs, along with their efficient extraction capabilities for various target compounds. Given the complex nature of extracts obtained from aflatoxin-contaminated pistachios, a subsequent clean-up and concentration step was required. The DES extracts were diluted with water and passed through a Supelco Discovery® DSC-18 solid-phase extraction (SPE) cartridge. Trapped aflatoxins were then eluted with a minimal quantity of methanol and injected into a high-performance liquid chromatography with fluorescence detection (HPLC-FLD) system equipped with a partially porous Supelco Ascentis Express® C18 column. The mobile phase, composed of methanol and water, utilized gradient elution conditions ranging from 30% to 60% MeOH over 7.5 minutes. The flow rate was maintained at 0.45 mL/min, the injection volume was 2 µL, and the column temperature was held at 35°C. This innovative approach shows the feasibility of utilizing DES as an extraction medium for the challenging lipidic matrix of aflatoxin-contaminated pistachios, thereby reducing the reliance on conventional solvents and minimizing process waste.