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Dual-color bioluminescent bioreporter for forensic analysis: evidence of androgenic and anti-androgenic activity of illicit drugs

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Abstract

Bioassays represent promising complementary techniques to conventional analytical approaches used in doping analysis to detect illicit drugs like anabolic-androgenic steroids (AAS). The fact that all AAS share a common mechanism of action via the human androgen receptor (hAR) enables the use of bioassays, relying on the activation of hAR as antidoping screening tools. Previously, we developed a dual-color bioreporter based on yeast cells engineered to express hAR and androgen response elements driving the expression of the bioluminescent (BL) reporter protein Photinus pyralis luciferase. A second reporter protein, the red-emitting luciferase PpyRE8, was introduced in the bioreporter as internal viability control. Here, we report the first forensic application of a straightforward, accurate, and cost-effective bioassay, relying on spectral resolution of the two BL signals, in 96-microwell format. The bioreporter responds to dihydrotestosterone as reference androgen in a concentration-dependent manner from 0.08 to 1,000 nM with intra- and inter-assay variation coefficients of 11.4 % and 13.1 %, respectively. We also demonstrated the suitability of this dual-color bioreporter to assess (anti)-androgenic activity of pure AAS, mixtures of AAS, and other illicit drugs provided by the Scientific Police. Significant anti-androgenic activity was observed in samples labeled as marijuana and hashish, containing Δ9-tetrahydrocannabinol as major constituent.

Evaluation of (anti)-androgenic activity of seized drugs with the dual-color bioluminescent bioreporter

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Acknowledgments

The authors gratefully acknowledge the financial support provided by the Italian Ministry of University and Research MIUR with the project FIRB 2008 “Development of bioremediation procedures and novel detection biotechnologies for Endocrine Disruptors” (prot. RBFR08SZTR). We are very grateful to Prof. Bruce Branchini for providing the cDNA encoding for PpyRE 8 and Dr. Jorma Lampinen for the opportunity to test the Varioskan® Flash instrument. We thank Marco Zanella and Ernesto Urbani (forensic chemists, Gabinetto Regionale di Polizia Scientifica, Bologna) for contributions in seized steroids analysis.

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Authors and Affiliations

  1. INBB, Istituto Nazionale di Biostrutture e Biosistemi, Viale delle Medaglie d’Oro 305, 00136, Rome, Italy

    Luca Cevenini, Elisa Michelini & Aldo Roda

  2. Department of Chemistry “G. Ciamician”, University of Bologna, Via Selmi 2, 40126, Bologna, Italy

    Elisa Michelini, Massimo Guardigli & Aldo Roda

  3. Gabinetto Regionale di Polizia Scientifica per l’Emilia Romagna, Polizia di Stato, Via Volto Santo, 3, 40123, Bologna, Italy

    Marcello D’Elia

Authors
  1. Luca Cevenini
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  2. Elisa Michelini
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  3. Marcello D’Elia
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  4. Massimo Guardigli
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  5. Aldo Roda
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Corresponding author

Correspondence to Elisa Michelini.

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Published in the special issue Analytical Science in Italy with guest editor Aldo Roda.

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Cevenini, L., Michelini, E., D’Elia, M. et al. Dual-color bioluminescent bioreporter for forensic analysis: evidence of androgenic and anti-androgenic activity of illicit drugs. Anal Bioanal Chem 405, 1035–1045 (2013). https://doi.org/10.1007/s00216-012-6416-6

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  • DOI: https://doi.org/10.1007/s00216-012-6416-6

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