title: Two novel real time cell-based assays quantify beta-blocker and NSAID specific effects in effluents of municipal wastewater treatment plants.
abstract: Pharmaceuticals, such as beta-blockers, nonsteroidal anti-inflammatory drugs (NSAIDs) as well as their metabolites are introduced into the water cycle via municipal wastewater treatment plant (WWTP) effluents in all industrialized countries. As the amino acid sequences of the biological target molecules of these pharmaceuticals - the beta-1 adrenergic receptor for beta-blockers and the cyclooxygenase for NSAIDs - are phylogenetically conserved among vertebrates it is reasonable that wildlife vertebrates including fish physiologically respond in a similar way to them as documented in humans. Consequently, beta-blockers and NSAIDs both exhibit their effects according to their mode of action on one hand, but on the other hand that may lead to unwanted side effects in non-target species. To determine whether residuals of beta-1 adrenergic receptor antagonists and cyclooxygenase inhibitors may pose a risk to aquatic organisms, one has to know the extent to which such organisms respond to the total of active compounds, their metabolites and transformation products with the same modes of action. To cope with this demand, two cell-based assays were developed, by which the total beta-blocker and cyclooxygenase inhibitory activity can be assessed in a given wastewater or surface water extract in real time. The measured activity is quantified as metoprolol equivalents (MetEQ) of the lead substance metoprolol in the beta-blocker assay, and diclofenac equivalents (DicEQ) in the NSAID assay. Even though MetEQs and DicEQs were found to surpass the concentration of the respective lead substances (metoprolol, diclofenac), as determined by chemical analysis by a factor of two to three, this difference was shown to be reasonably explained by the presence and action of additional active compounds with the same mode of action in the test samples. Thus, both in vitro assays were proven to integrate effectively over beta-blocker and NSAID activities in WWTP effluents in a very sensitive and extremely rapid manner.
authors.affiliation: Steinbeis-Innovationszentrum Zellkulturtechnik, c/o University of Applied Sciences Mannheim, Paul-Wittsack-Str. 10, D-68163 Mannheim, Germany.
authors.forename: Kevin
authors.initials: K
authors.lastname: Bernhard
authors.affiliation: Steinbeis-Innovationszentrum Zellkulturtechnik, c/o University of Applied Sciences Mannheim, Paul-Wittsack-Str. 10, D-68163 Mannheim, Germany.
authors.forename: Cordula
authors.initials: C
authors.lastname: Stahl
authors.affiliation: Steinbeis-Innovationszentrum Zellkulturtechnik, c/o University of Applied Sciences Mannheim, Paul-Wittsack-Str. 10, D-68163 Mannheim, Germany.
authors.forename: Regina
authors.initials: R
authors.lastname: Martens
authors.affiliation: University of Tübingen, Animal Physiological Ecology, Auf der Morgenstelle 5, D-72072 Tübingen, Germany.
authors.forename: Heinz-R
authors.initials: HR
authors.lastname: Köhler
authors.affiliation: University of Tübingen, Animal Physiological Ecology, Auf der Morgenstelle 5, D-72072 Tübingen, Germany; Steinbeis Transfer Centre for Ecotoxicology and Ecophysiology, Blumenstr. 13, D-72108 Rottenburg, Germany.
authors.forename: Rita
authors.initials: R
authors.lastname: Triebskorn
authors.affiliation: TZW: DVGW - Technologiezentrum Wasser, Karlsruher Straße 84, D-76139 Karlsruhe, Germany.
authors.forename: Marco
authors.initials: M
authors.lastname: Scheurer
authors.affiliation: Steinbeis-Innovationszentrum Zellkulturtechnik, c/o University of Applied Sciences Mannheim, Paul-Wittsack-Str. 10, D-68163 Mannheim, Germany. Electronic address: manfred.frey@stw.de.
authors.forename: Manfred
authors.initials: M
authors.lastname: Frey
mesh_terms: D000319:Adrenergic beta-Antagonists; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal; D001681:Biological Assay; D006801:Humans; D062065:Wastewater / Q000737:chemistry*; D014874:Water Pollutants, Chemical
chemical_list: D000319:Adrenergic beta-Antagonists; D000894:Anti-Inflammatory Agents, Non-Steroidal; D062065:Wastewater; D014874:Water Pollutants, Chemical
keywords: Beta-blocker effects; In vitro assay; Mode of action-based biotest; NSAID effects; Pharmaceuticals; Wastewater treatment plant effluent