Plasma levels of non-steroidal anti-inflammatory drugs (NSAIDs) and - PowerPoint PPT Presentation

Plasma levels of non-steroidal anti-inflammatory drugs (NSAIDs) and gemfibrozil in fish exposed to treated sewage effluents: Assessing the risk of discharges to aquatic environments Jeffrey N. Brown † , Nicklas Paxéus*, Lars Förlin ‡ and D.G. Joakim Larsson † † Institute for Neuroscience and Physiology, the Sahlgrenska Academy at Göteborg University, Sweden. Email: jeffrey.brown@fysiologi.gu.se * Environmental Chemistry, Gryaab AB, Göteborg. ‡ Department of Zoology, Göteborg University.

Introduction PHARMACEUTICALS IN AQUATIC ENVIRONMENTS • Widespread detection in surface waters • Effects on fish at environmentally relevant concentrations • Environmental Risk Assessment (ERA) required for all new pharmaceuticals in EU

Fish Plasma Model * Fish exposure via water Human therapeutic plasma concentration QSPR calculates giving known effects bioconcentration (H T PC) Fish steady state plasma concentration (F SS PC) [ ] [ ] Effect Ratio (ER) = The lower the ER, the higher the risk for a specific target interaction * Huggett, et al. (2003) Human Ecol. Risk Assess. (9) 1789-99

Fish Plasma Model * Fish exposure via water Human therapeutic plasma concentration QSPR calculates giving known effects bioconcentration (H T PC) Fish steady state plasma concentration (F SS PC) Log BCF plasma = 0.73 log K ow – 0.88 Fitzsimmons et al. (2001) Aquatic Toxicol. (55) 23-34. [ ] [ ] Non-polar PCBs, chloroethanes + benzene Effect Ratio (ER) = • Not yet well validated for pharmaceuticals (often ionic) The lower the ER, the higher the risk for a specific target • Complex environmental exposures? interaction

Aims 1. To test the bioconcentration QSPR of the Fish Plasma Model using fish exposed to diluted sewage effluents. 2. Use the Fish Plasma Model to rank the risks of target interactions occurring in fish.

Non-steroidal anti-inflammatory drugs (NSAIDs) Ibuprofen • 0.1 – 2 μ g/l in treated log K ow = 3.80 sewage effluents pK a = 4.40 Diclofenac • Non-ionised state log K ow = 4.02 pK a = 4.15 lipophilic enough to Naproxen bioconcentrate log K ow = 3.10 pK a = 4.20 • Reported effects in Ketoprofen fish log K ow = 3.00 pK a = 4.60

Gemfibrozil • Blood lipid lowering log K ow = 4.80 agent pK a = 4.80 • ~ 1 μ g/l in treated sewage effluents • In aquarium studies bioconcentrates into goldfish blood plasma • Effects on fish observed

Exposure Sites 1. GRYAAB STP Göteborg ~ 600,000 people, some trade wastes, activated sludge, chemical P and biological N removal

Exposure Sites 2. Gråbo STP ~ 4000 people, activated sludge

Exposure Sites 3. Spenshult Rheumatic Hospital STP 180 beds + staff, activated sludge higher pharmaceutical load?

Methods Fish Plasma • Juvenile rainbow trout (70g) exposed 2-3 weeks, sacrificed, blood taken, centrifuged & plasma frozen • SPE column extraction, derivatisation & GC-MS Water samples • Sampling every 2-3 days • SPE extraction, derivatisation & GC-MS

Results Qualitative static aquarium exposure (2 days) Pharmaceutical Nominal Measured exposure level plasma level BCF (ng/ml) (ng/ml) 520 4680 9 Ibuprofen 1000 3640 4 Naproxen Diclofenac 520 3440 7 490 60 0.1 Ketoprofen Gemfibrozil 510 32070 63

Sewage exposures - pharmaceutical concentrations (ng/ml, mean ± SD) Pharmaceutical Exposure Site Gryaab Gråbo Spenshult 0.0045 (0.0027) 0.014 (0.013) 0.048 (0.029) Ibuprofen Naproxen 0.25 (0.17) 0.68 (0.33) 0.043 (0.016) 2.32 (0.48) 0.64 (0.28) 0.28 (0.08) Diclofenac Ketoprofen 0.28 (0.18) 0.10 (0.04) 0.23 (0.08) 1.05 (0.25) 0.54 (0.23) 1-5* Gemfibrozil * Interfering long-chain fatty acid prevented full quantification

Sewage exposures - Fish steady state plasma concentrations (F SS PC) (ng/ml, mean ± SD) Pharmaceutical Exposure Site Gryaab Gråbo Spenshult 84 (62) <mdl <mdl Ibuprofen Naproxen 14 (9) <mdl <mdl 12 (14) <mdl <mdl Diclofenac Ketoprofen <mdl <mdl <mdl 210 (400) <mdl 8.0 (5.5) Gemfibrozil

Sewage exposures - Fish steady state plasma concentrations (F SS PC) (ng/ml, mean ± SD) Pharmaceutical Exposure Site Gryaab Gråbo Spenshult 84 (62) <mdl <mdl Ibuprofen Naproxen 14 (9) <mdl <mdl 12 (14) <mdl <mdl Diclofenac Ketoprofen <mdl <mdl <mdl 210 (400) <mdl 8.0 (5.5) Gemfibrozil Temperature o C 12 2 14

Sewage exposures - Fish steady state plasma concentrations (F SS PC) (ng/ml, mean ± SD) Pharmaceutical Exposure Site Gryaab Gråbo Spenshult 84 (62) <mdl <mdl Ibuprofen Naproxen 14 (9) <mdl <mdl 12 (14) <mdl <mdl Diclofenac Ketoprofen <mdl <mdl <mdl 210 (400) <mdl 8.0 (5.5) Gemfibrozil Temperature ( o C) 12 2 14 pH 7.0 6.4 7.1

Sewage exposures - Fish steady state plasma concentrations (F SS PC) (ng/ml, mean ± SD) Pharmaceutical Exposure Site Gryaab Gråbo Spenshult 84 (62) <mdl <mdl Ibuprofen Naproxen 14 (9) <mdl <mdl 12 (14) <mdl <mdl Diclofenac Ketoprofen <mdl <mdl <mdl 210 (400) <mdl 8.0 (5.5) Gemfibrozil Temperature ( o C) 12 2 14 pH 7.0 6.4 7.1 Exposure duration 2, 8, 16 days 2, 8, 16, 23 14 days days

Site specific differences in drug bioconcentration • Temperature • pH • Exposure duration • Particles/colloids in natural stream dilution water limiting bioavailability at Gråbo and Spenshult? • Gryaab – Surfactants enhancing bioavailability? – Factors reducing active excretion/metabolism?

Fish Plasma Model QSPR Evaluation - Bioconcentration Factors 100000 Model Bioconcentration Gryaab 10000 Factor (BCF) Gråbo 1000 Spenshult 100 10 1 Naproxen Diclofenac Ibuprofen Ketoprofen Gemfibrozil Max. estimated BCF based on plasma concentration set at 3 ng/ml detection limit

Fish Plasma Model Evaluation - Bioconcentration Factors • Measured BCFs generally ≤ modelled BCFs – QSPR does not take into account special characteristics of pharmaceuticals – Active excretion/metabolism may lower plasma concentrations – Drug bioavailability may be significantly lower through partitioning to colloids or particles • Model greatly underestimated ibuprofen BCF at Gryaab

Ranking of risk of target interactions [ ] [ ] Effects Ratio (ER) = Lower ER → higher risk of target interaction Model ranking Diclofenac > gemfibrozil > ibuprofen > naproxen - ketoprofen Measured ranking = model ranking Diclofenac > gemfibrozil > ibuprofen > naproxen - ketoprofen

Measured v Modelled Effects Ratio at Gryaab 100000 Model Effects Ratio (ER) Gryaab 10000 1000 100 10 1 Naproxen Diclofenac Ketoprofen Ibuprofen Gemfibrozil Minimum estimated ER based on 3 ng/ml detection limit

Conclusions 1. Pharmaceuticals bioconcentrated from treated sewage effluents into fish blood 2. Major bioconcentration differences between sites shows the importance of field studies 3. Diclofenac appears to present the highest risk of pharmacological effects occurring in fish 4. Fish Plasma Model looks promising with refinement of the QSPR and further validation

Acknowledgements Lars Förlin Joakim Larsson Jari Parkkonen Linda Samuelsson Lina Gunarsson Linda Augustsson • Spenshult Rheumatic Hospital • Gråbo STP Nicklas Paxéus

Acknowledgements You!

Risk of target interactions 10000 Gryaab Effects Ratio (ER) Gråbo 1000 Spenshult 100 10 1 Naproxen Diclofenac Ketoprofen Ibuprofen Gemfibrozil Minimum estimated ER based on 3 ng/ml detection limit

Human therapeutic plasma concentrations Pharmaceutical H T PC (ng/ml) Ibuprofen 15,000 Naproxen 20,000 Diclofenac 420 Ketoprofen 5,000 Gemfibrozil 15,000