title: Identification of P-glycoprotein inhibitors in contaminated freshwater sediments.
abstract: P-glycoprotein (P-gp, ABCB1) is an important part of the multixenobiotic resistance (MXR) defense system in aquatic organisms. The main goal of this study was identification of P-gp inhibitors in contaminated sediments using the effect-directed analysis (EDA) approach. The samples were collected from the Gorjak creek (Zagreb, Croatia), a recipient of wastewater effluents from the pharmaceutical industry. Sediment samples were extracted and fractionated using a two-tiered approach. Resulting nonpolar, medium polar, and polar fractions were tested on the inhibition of P-gp activity using P-gp overexpressing PLHC-1/dox cells and calcein-AM as model substrate. The obtained EC50 values (up to 757 μg/g, expressed in toxicity equivalents of model P-gp inhibitor cyclosporine A) revealed high inhibitory potential of polar fractions of investigated sediments and clearly reflected the impact of pharmaceutical wastewater. P-gp specific ATPase assay and the cytotoxicity modulation experiments with colchicine indicated that most of the observed P-gp inhibition was due to the presence of noncompetitive inhibitors. A detailed chemical analysis by ultrahigh-performance liquid chromatography-quadrupole/time-of-flight mass spectrometry (UPLC-QTOFMS) revealed nonionic surfactants, including alcohol polyethoxylates (LAEOs) and polypropylene glycols (PPGs), as the major components of the most active subfractions. Testing of several LAEO and PPG commercial mixtures confirmed their potential to inhibit the fish P-glycoprotein and modulate toxicity of other xenobiotics present in complex environmental samples.
authors.affiliation: Division for Marine and Environmental Research, Rudjer Boskovic Institute, Zagreb, Croatia.
authors.forename: Roko
authors.initials: R
authors.lastname: Zaja
authors.forename: Senka
authors.initials: S
authors.lastname: Terzić
authors.forename: Ivan
authors.initials: I
authors.lastname: Senta
authors.forename: Jovica
authors.initials: J
authors.lastname: Lončar
authors.forename: Marta
authors.initials: M
authors.lastname: Popović
authors.forename: Marijan
authors.initials: M
authors.lastname: Ahel
authors.forename: Tvrtko
authors.initials: T
authors.lastname: Smital
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors*; D000818:Animals; D045744:Cell Line, Tumor; D005618:Fresh Water; D019015:Geologic Sediments / Q000737:chemistry*; D014874:Water Pollutants, Chemical / Q000032:analysis* / Q000494:pharmacology
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D014874:Water Pollutants, Chemical