A versatile extraction and purification method for ivermectin detection through HPLC and LC/MS in feed, water, feces, soil, and sewage

Ivermectin is an effective and safe broad-spectrum anthelmintic drug widely used in veterinary medicine belonging to the macrocyclic lactone family. Pharmaceuticals are of increasing concern as environmental contaminants, they represent a risk, even at low concentrations, for human health, contributing to the diffusion of antimicrobial resistance, and for aquatic and terrestrial ecosystems. The European Commission defined various actions to address this problem linked to drug production and their introduction into the environment by the livestock industry. Environmental residues of ivermectin, if present, may be a risk to non-target organisms, among which fish and some arthropods, which die when in contact even with the lowest doses of the drug. High-performance liquid chromatography with diode array detector (HPLC-DAD) analysis is frequently applied to detect ivermectin residues in feed and environmental samples, but the detectable concentration range is limited to the magnitude of ppm unless the derivatization techniques are applied during the extraction and purification steps. Mass-spectrometry-based techniques allow to detect the ivermectin residues in the ppb-ppt range, improving both the sensitivity and accuracy of the method. Therefore, the purpose of this study was to adapt the HPLC-DAD extraction and purification method to the mass-spectrometry analysis to evaluate ivermectin residues in feed and environmental samples. We adapted an existing analysis method 2 designed for high-performance liquid chromatography with diode array detector analysis to mass spectrometry detection to evaluate residues of ivermectin in feed, water, feces, soil, and sewage. Doramectin was used as a process standard. The extraction is performed using methanol. The purification implies active alumina, a solid phase extraction technique through C-18 and then silica columns. For soil samples, the steps with columns were not necessary. Samples could also be concentrated by loading a larger volume of sample into the columns and by resuming the dry eluate with a smaller volume. The results were interpreted using a calibration curve, diluted 1:100 from the original method for HPLC, (p < .001) with intercept values (>0.999) indicating excellent linearity. The limit of detection was 150 PPT in the extract. The limit of quantification was 300 PPT. This method is easy, inexpensive, precise, and versatile. The extracts obtained with the original extraction and purification method were useful also for the LC/MS/MS analysis after the substitution of the mobile phase, used also to resume the dry shot. It is useful for analysis in all the matrixes examined and it will be useful for future research in ecotoxicology.