Développement de techniques analytiques pour la détermination des agents anti-infectieux dans les eaux environnementales

Anti-infectives are used to treat or prevent infections in humans, animals, insects and plants. The occurrence of traces of these substances in wastewaters, natural waters and even drinking water has caused concern among the scientific community especially because of their biological activiy. The goal of this research was to study the occurrence of anti-infectives in contaminated environmental waters (wastewaters, natural waters, and drinking water) and to develop new analytical methods able to quantitate and confirm their presence in these matrices. A meta-analysis on the occurrence of anti-infectives in contaminated environmental waters demonstrated that at least 68 parent compounds and 10 transformation products have been quantified to date. Environmental concentrations vary between 0.1 ng/L and 1 mg/L depending on the compound, the matrix and the source of contamination. According to this study, detrimental effects of anti-infectives on aquatic biota are possible and these substances could also affect indirectly human health because of their possible contribution to the dissemination of antibiotic resistance in bacteria. Preliminary tests on the development of a method of determination of anti-infectives in wastewaters showed the main difficulties to overcome during solid-phase extraction (SPE) as well as the importance of the detector selectivity. A novel method of determination of anti-infectives was described using off-line tandem SPE and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Six target anti-infectives (sulfamethoxazole, trimethoprim, ciprofloxacin, levofloxacin, clarithromycin and azithromycine) were quantitated at concentrations between 39 and 276 ng/L in samples of influent and effluent collected from a primary and physico-chemical wastewater treatement plant. Reported effluent concentrations indicate that the mean mass of these substances discharged daily in the St. Lawrence River was ~ 2 kg. In order to reduce total analysis time and simplify sample preparation, a new on-line SPE-LC-MS/MS was presented. This method used a column-switching technique to preconcentrate 1.00 mL of sample in an on-line SPE column. Method analytical performance allowed the quantitation of six anti-infectives in municipal wastewaters and limits of detection were of the same magnitude (13-60 ng/L) than methods based in offline SPE. Next, the application of turbulent flow chromatography on-line SPE columns for the preconcentration of six anti-infectives in wastewaters was explored. Results showed that these columns are an interesting alternative to traditional on-line SPE columns. Finally, in order to allow analysis of anti-infectives in surface and drinking water, we developed an on-line SPE-LC-MS/MS method using large-volume injections (10 mL). Breakthrough volumes of several on-line SPE columns were estimated and the column having the best retention, Strata-X, was chosen. Method detection and confirmation limits were between 1 and 6 ng/L. Analysis of real samples indicated that the concentration of the three target anti-infectives (sulfamethoxazole, trimethoprim and clarithromycin) was lower that the method detection limits. Accurate mass measurement by time-of-flight mass spectrometry and product ion spectra obtained by a reversed-energy ramp in a triple quadrupole mass spectrometer were explored as alternative confirmation methods.

Keyphrases: Anti-infectives, Pharmaceuticals, Drinking water, Surface water, Solid-phase extraction, Tandem mass spectrometry, Liquid chromatography, Antibiotics, Wastewaters, Trace analysis