Development of a sensitive E-screen assay for quantitative analysis of estrogenic activity in municipal sewage plant effluents

Development of a sensitive E-screen assay for quantitative analysis of estrogenic activity in... A simplified proliferation test with human estrogen receptor-positive MCF-7 breast cancer cells (E-screen assay) was optimized and validated for the sensitive quantitative determination of total estrogenic activity in effluent samples from municipal sewage plants. After solid phase extraction of 1 l sewage on either 0.2 g polystyrene copolymer (ENV+) or 1 g RP-C18 material and removal of the solvent, analysis of the extracts in the E-screen assay could be performed without any clean-up step. This was even possible with untreated sewage. Parallel extraction of four sewage samples on both different solid phase materials gave comparable quantitative results in the E-screen. A blank sample did not induce cell proliferation. As additive behaviour of the estrogenic response of single compounds was proven for two different mixtures each containing three xenoestrogens, total estrogenic activity in the sewage samples, expressed as 17β-estradiol equivalent concentration (EEQ), could be calculated comparing the EC 50 values of the samples with those of the positive control 17β-estradiol. The detection limit of the E-screen method was 0.05 pmol EEQ/l (0.014 ng EEQ/l), the limit of quantification 0.25–0.5 pmol EEQ/l (0.07–0.14 ng EEQ/l). In total, extracts of nine effluent and one influent sample from five different municipal sewage plants in South Germany were analyzed in the E-screen. All samples strongly induced cell proliferation in a dose-dependent manner which was completely inhibited by coincubation with 5 nM of the estrogen receptor-antagonist ICI 182,780. The proliferative effect relative to the positive control 17β-estradiol (RPE) was between 30 and 101%. 17β-Estradiol equivalent concentrations were between 2.5 and 25 ng/l indicating a significant input of estrogenic substances via sewage treatment plants into rivers. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Science of the Total Environment Elsevier

Development of a sensitive E-screen assay for quantitative analysis of estrogenic activity in municipal sewage plant effluents

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Publisher
Elsevier
Copyright
Copyright © 1999 Elsevier Science B.V.
ISSN
0048-9697
eISSN
1879-1026
DOI
10.1016/S0048-9697(99)80015-1
Publisher site
See Article on Publisher Site

Abstract

A simplified proliferation test with human estrogen receptor-positive MCF-7 breast cancer cells (E-screen assay) was optimized and validated for the sensitive quantitative determination of total estrogenic activity in effluent samples from municipal sewage plants. After solid phase extraction of 1 l sewage on either 0.2 g polystyrene copolymer (ENV+) or 1 g RP-C18 material and removal of the solvent, analysis of the extracts in the E-screen assay could be performed without any clean-up step. This was even possible with untreated sewage. Parallel extraction of four sewage samples on both different solid phase materials gave comparable quantitative results in the E-screen. A blank sample did not induce cell proliferation. As additive behaviour of the estrogenic response of single compounds was proven for two different mixtures each containing three xenoestrogens, total estrogenic activity in the sewage samples, expressed as 17β-estradiol equivalent concentration (EEQ), could be calculated comparing the EC 50 values of the samples with those of the positive control 17β-estradiol. The detection limit of the E-screen method was 0.05 pmol EEQ/l (0.014 ng EEQ/l), the limit of quantification 0.25–0.5 pmol EEQ/l (0.07–0.14 ng EEQ/l). In total, extracts of nine effluent and one influent sample from five different municipal sewage plants in South Germany were analyzed in the E-screen. All samples strongly induced cell proliferation in a dose-dependent manner which was completely inhibited by coincubation with 5 nM of the estrogen receptor-antagonist ICI 182,780. The proliferative effect relative to the positive control 17β-estradiol (RPE) was between 30 and 101%. 17β-Estradiol equivalent concentrations were between 2.5 and 25 ng/l indicating a significant input of estrogenic substances via sewage treatment plants into rivers.

Journal

Science of the Total EnvironmentElsevier

Published: Jan 12, 1999

References

  • A survey of estrogenic activity in United Kingdom inland waters
    Harries, JE; Sheahan, DA; Jobling, S; Matthiessen, P; Neall, P; Routledge, EJ; Rycroft, R; Sumpter, JP; Taylor, T
  • Development of a marker of estrogenic exposure in human serum
    Sonnenschein, C; Soto, A; Fernandez, MF; Olea, N; Olea-Serrano, MF; Ruiz-Lopez, MD
  • Behavior and occurrence of estrogens in municipal sewage treatment plants. I. Investigations in Germany, Canada and Brazil
    Ternes, TA; Stumpf, M; Mueller, J; Haberer, K; Wilken, R-D; Servos, M

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