ARTICLE IN PRESS

0147-6513$ - se

doi101016jec

$Funding so

commission fro

2006) Part of

Norwegian Res

and manageme

in the Norwegi

The work pe

performed in ac

protection of anCorrespondE-mail addr

Please cite th

Ecotoxicol E

Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]]

wwwelseviercomlocateecoenv

Environmental assessment of Norwegian priority pharmaceuticalsbased on the EMEA guideline$

Merete Grunga Torsten Kallqvista Solveig SakshaugbSvetlana Skurtveitb Kevin V Thomasa

aNIVA (Norwegian Institute for Water Research) Gaustadalleen 21 N-0349 Oslo NorwaybNorwegian Institute of Public Health PO Box 4404 Nydalen N-0403 Oslo Norway

Received 23 August 2007 received in revised form 3 October 2007 accepted 21 October 2007

Abstract

An environmental risk assessment of eleven pharmaceuticals according to the guideline recommended by the European Medicines

Evaluation Agency (EMEA) was performed Cefuroxime ciprofloxacin cyclophosphamide diclofenac ethinylestradiol ibuprofen

metoprolol paracetamol sulfamethoxazole tetracycline and trimethoprim were selected for assessment by the Norwegian Pollution

Control Authority Predicted environmental concentrations (PECs) were calculated according to both the EMEA guideline and a

conventional model for comparison and ranged from 00002 to 45mgL Available acute and chronic toxicity data were collected from the

literature although no data were available for cyclophosphamide Toxicity tests showed cyclophosphamide to have relatively low acute

toxicity with an EC50 for Pseudokirchneriella subcapitata 4100mgL and a Daphnia magna reproduction NOEC of 56mgL These and

the literature data were used to derive predicted no effect concentrations (PNEC) Risk quotients (PECPNEC) were then calculated for

all 11 pharmaceutical compounds Risk quotients greater than 1 were obtained for ciprofloxacin diclofenac ethinylestradiol

sulfamethoxazole and tetracycline according to the EMEA guideline Measured environmental concentrations (MECs) confirmed that

the release of ciprofloxacin from wastewater treatment works may potentially be of environmental concern in Norway

r 2007 Elsevier Inc All rights reserved

Keywords Pharmaceuticals Environmental risk assessment (ERA) EMEA guidelines Cyclophosphamide Algal growth inhibition D magna

reproduction

1 Introduction

The sales and use of pharmaceuticals have graduallyincreased over time worldwide In Norway the annualgrowth measured in number of defined daily doses (DDDs)has varied from 1 to 8 in the period 1993ndash2006

e front matter r 2007 Elsevier Inc All rights reserved

oenv200710015

urces Part of this study was performed by NIVA on

m Norwegian Pollution Control Authority (TA 2216

this study was performed as part of a grant from the

earch Council to NIVA (Project number 172526 Fate risk

nt of pharmaceuticals and personal care products (PPCPs)

an sewage system)

rformed in this study with experimental organisms was

cordance with national and international guidelines for the

imal welfare

ing author Fax +4722 18 52 00

ess meretegrungnivano (M Grung)

is article as Grung M et al Environmental assessment o

nviron Saf (2007) doi101016jecoenv200710015

(Norwegian Institute of Public Health 2007) Pharmaceu-ticals over the past years have become an increasing pointof environmental concern Wastewater treatment plants(WTPs) have been identified as a major environmentalsource of these compounds As a consequence variablequantities of pharmaceuticals can reach surface watersgroundwaters and sediments resulting in concentrationsranging from nanograms to micrograms per liter (Kum-merer 2001) Pharmaceuticals can be degraded in theenvironment by biotic andor abiotic processes but maycause persistent exposure due to their continuous infusioninto aquatic media via sewage treatment plant (STP)effluents (Castiglioni et al 2006 Vieno et al 2007) Due tothe large amounts of pharmaceuticals released into theenvironment and their intrinsic properties to causebiological effects the risk they present to the environmentcannot be ignored (Fent et al 2006)

f Norwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESSM Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]]2

Procedures for conducting environmental risk assess-ment (ERA) on pharmaceuticals are in effect in bothEurope and United States (CHMP 2006 US Food andDrug Administration (USFDA) 1998) The Committee forMedicinal Products for Human Use (CHMP) of theEuropean Medicines Evaluation Agency (EMEA) haspublished guidelines for ERA which came into effect onthe 1st December 2006 An ERA is required for all newmarketing authorisation applications for medicinal pro-ducts An evaluation of the environmental impact shouldalso be made if there is an increase in the environmentalexposure eg a new indication may result in a significantincrease in the extent of the use In essence these guidelinesfollow the general principle of the ERA procedures asapplied to existing and new conventional chemicals inEurope (EU TGD) (European Commission 2003)

The EMEA guideline (CHMP 2006) describes how toevaluate the potential risks of the medicinal product to theenvironment The guideline is focused only on theenvironmental risks associated with the use of medicinalproducts not arising from storage disposal synthesis ormanufacture of medicinal products The guideline de-scribes a stepwise tiered procedure for ERA The Phase I isa pre-screening assessment aiming at a first estimation ofexposure The Phase I has an action limit of 001 mgL andif the PEC of surface water is below this limit it is assumedthat the compound is unlikely to represent a risk for theenvironment However in some cases the action limit maynot be applicable for example regarding endocrinedisrupting compounds If the PEC is equal to or above001 mgL then a Phase II environmental fate and effectanalysis should be performed Phase II is further dividedinto an initial prediction of risk based on a dataset ofaquatic toxicology and fate (Phase II Tier A) If necessaryan extended risk assessment is performed (Phase II Tier B)where the objective is a substance and compartment-specific refinement based on an extended dataset onemission fate and effects

In Norway little work has been performed on theoccurrence of pharmaceuticals in the environment orthe environmental risk associated with their occurrence inthe aquatic environment The occurrence of severalpharmaceutical products in Norway has been investigatedand reported (Thomas et al in press 2007) Theoccurrence of selective serotonin reuptake inhibitors hasbeen investigated in a study performed in Tromsoslash Norway(Vasskog et al 2006) Samples were taken from threedifferent STPs and a pump station two of the STPs wereserving the University hospital and its psychiatric depart-ment More recently the occurrence of pharmaceuticals ineffluents from two hospitals and in the influent and effluentof the servicing STP in Oslo was investigated (Thomaset al in press) For Norwayrsquos neighbouring countries ERAof 27 pharmaceutical ingredients and five excipients used inpharmaceutical products have been performed in Sweden(Carlsson et al 2006a b) Nine pharmaceutical ingredientswere identified as dangerous to the aquatic environment

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

but only estradiol and ethinylestradiol were considered tobe associated with possible aquatic environmental risks Ofthe excipients docusate sodium was identified as a possiblerisk for sediment-dwelling organisms In Finland theoccurrence of eight pharmaceuticals was assessed in theraw and treated sewage of 12 STPs (Vieno et al 2007)In Norway ERA of pharmaceuticals has not yet been

performed but the Norwegian Pollution Control Author-ity has performed a preliminary prioritisation of pharma-ceutical compounds to be further investigated (NorwegianPollution Control Authority 2005) Since Norway hasexcellent sources regarding the volume of pharmaceuticalssales we wanted to compare the PEC estimation accordingto EMEA guidelines (CHMP 2006) and a conventionalmodel based on wholesales figures on pharmaceuticalsBased on existing environmental data from the literatureas well as toxicity studies reported in this study riskquotients for eleven pharmaceuticals could be obtainedThe main objective of this study was to assess theenvironmental risk associated with 11 selected pharmaceu-tical compounds

2 Materials and methods

21 Compounds investigated

The compounds investigated are listed in Table 1 All pharmaceuticals

marketed in Norway are classified according to the Anatomical

Therapeutic Chemical Classification (ATC) system (WHO Collaborating

Centre for Drug Statistics Methodology) The corresponding ATC codes

to the chosen compounds are included in Table 1

22 Chemicals

Cyclophosphamide monohydrate (purity 997) was obtained from

Fluka (Schnelldorf Germany) and a primary stock solution (3 gL) was

prepared by dissolving the chemical in test medium All solvents used were

from JT Baker (Deventer The Netherlands ultra resi-analyzed) and were

designed for organic residue analysis

23 Growth inhibition test with Pseudokirchneriella subcapitata

The growth inhibition test was carried out according to the Organisa-

tion for Economic Cooperation and Development Test Guideline 201

(OECD 2006) with P subcapitata as the test organism The strain used

was NIVA strain CHL 1 from the culture collection of the Norwegian

Institute for Water Research (Oslo Norway) A concentration series of

cyclophosphamide (nominal concentrations 32 56 10 18 32 56 and

100mgL) was obtained by dilution of a stock solution in the growth

medium Three replicates for each concentration and six control replicates

were prepared Flat-bottom 100ml glass flasks with 50ml of test solution

were used as test vessels The flasks were first soaked with an aliquot of the

test solution which was discarded before the flasks were filled All vessels

were inoculated with 5 106 cellsmL of P subcapitata from an

exponentially growing culture in the same growth medium The vessels

were incubated on a reciprocating table illuminated by cool-white

fluorescent tubes providing 75mMm2s of photosynthetic active radiation

The incubation temperature was 2371 1C The cell density in the cultures

was monitored by daily counting with a Coulter Multisizer M3 (Beckman

Coulter Miami FL USA) electronic particle counter The test was

terminated after 72 h The specific growth rate of P subcapitata in each

replicate culture was calculated from the logarithmic increase in cell

f Norwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESS

Table 1

Compounds analysed including the corresponding ATC codes defined daily dose and total sales in Norway given in number of DDD100 inhabitantsday

Compound ATC codes DDD (mg) DDD100 inhabitantsday

Cefuroxime J01DC02 3000 0013

Ciprofloxacin J01MA02 1000 (tabl)500 (inj) 0052a

Cyclophosphamide L01AA01 00010b

Diclofenac M01AB05 100 077c

Ethinylestradiol G03Ad and G03HB01 d 45

Ibuprofen M01AE01 1200 14e

Metoprolol C07AB02 150 22

Paracetamol N02BE01 3000 33f

Sulfamethoxazole J01EE01 480g 0032

Tetracycline J01AA07 1000 0064

Trimethoprim J01EA01 400 010h

aSales of eye drops (ATC code S01AX13) where no DDD is assigned is included by using the DDD for the oral formulation of ciprofloxacinbNo DDD assigned sales is given in number of grams of cyclophosphamide100 inhabitantsyearcSales of topical gel and eye drops (ATC code D11AX18 and S01BC03) where no DDD is assigned is included by using the DDD for the oral

formulationdIncludes the following ATC codes G03AA07 G03AA09G03AA12 G03AA13 G03AB03 and G03AB04 The DDD is 1 tablet for packages

containing 28 tablets and 075 tablet for packages containing 21 tabletseSales of creams (ATC code M02AA13) where no DDD is assigned is included in the figures by using the DDD for the oral formulation of ibuprofenfSales of fixed combination products containing paracetamol is included (ATC code N02AA59 and N02AC54) by using the same DDD as for plain

paracetamol productsgDDD is given in unit dose corresponding to a DDD of 480mg sufamethoxazolehSales of trimethoprim in combination with sulfamethoxazole (ATC code J01EE01) is included by using the same DDD as for plain trimethoprim

products

M Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]] 3

density in the intervals from 0 to 72 h using the following equation

m frac14lnethNnTHORN lnethN0THORN

tn t0

where N0 is the cell concentration at t0 (106 L1) N1 the cell concentration

at t1 (106 L1) N2 the cell concentration at t2 (106 L1) Nn the cell

concentration at time tn (106 L1) t1 the time of first measurement (hours

from start) and tn the time of nth measurement (hours from start)

The exposure concentrations of cyclophosphamide were verified by

analysis of unfiltered test solutions at two concentration levels at the start

and end of the test

24 Daphnia magna reproduction test

The effect of cyclophosphamide on the parthogenetic reproduction of

D magna was investigated according to the OECD Test Guideline 211

(OECD 1998) Daphnia magna clone A (Baird et al 1991) was used as the

test organism A culture of this strain was kept in the laboratory in the

medium Elendt M7 (OECD 1998) and was fed with the green alga

P subcapitata Juvenile Daphnia organisms (age 24 h) were isolated from

the stock culture and exposed individually to five levels of cyclopho-

sphamide (nominal concentrations 10 18 32 56 and 100mgL) The test

solutions were prepared by dilution of stock solution (600mgL in dilution

water) of cyclophosphamide to M7 medium Erlenmeyer flasks (volume

100ml) were used as test vessels These flasks were soaked with an aliquot

of the test solution which was discarded before the flasks were filled with

50ml of test solution Ten Daphnia organisms were exposed individually at

each concentration of cyclophosphamide and in the control Every 2ndash3

days the test animals were transferred to fresh test solutions The animals

were fed daily with P subcapitata The feeding ratio was increased

gradually from approximately 01mg organic carbon (C)Daphniad in the

beginning of the test to 02mg CDaphniad after 10 days The number of

offspring produced by each parent animal was recorded daily until the test

was terminated after 21 days The cumulative number of offspring per

parent animal at each treatment was compared to the control and

exposure concentrations were verified by analysis of fresh and used

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

unfiltered test solutions at two concentration levels on two occasions

during the test pH was measured in fresh and used test media at all media

renewals

25 Chemical analysis of cyclophosphamide

The analyses of water samples were performed by Norwegian Institute

for Air Research and the method is described in detail in Thomas et al

(in press) Briefly the water samples were passed through SPE-columns

(Oasis HLB column 200mg) Cyclophosphamide was eluted by

methanol and acetone Analyses were performed by LC-MS (Waters

Alliance HPLC coupled to Micomass Quattro Premier triple quadrupole

mass spectrometer) with gradient elution by water and acetonitrile both

added 0075 formic acid The detector was operated in positive ion

mode and the electrospray source parameters were optimised for the

analyte

26 Calculation of results and statistical treatment

The growth rate of P subcapitata was calculated relative to the control

values and plotted against the logarithmic concentrations of cyclopho-

sphamide A log-logistic model (Hill 1910) was fitted to the data points

using the software REGTOX EV63 (Eric Vindimian Paris France

httpericvindimian9onlinefr) The 50 effect concentration (EC50)

was derived from the model The NOEC defined as the highest

concentration that did not significantly affect the measured parameters

when compared to the control cultures and the LOEC defined as the

lowest concentration causing a significant effect were determined for

growth rate inhibition of P subcapitata as well as for reproductive output

of D magna Normality of data was checked by a KolmogorovndashSmirnov

test before further statistical analysis Significant differences between

normally distributed groups of equal variance (Levenersquos test) were

determined by one-way analysis of variance followed by Dunnettrsquos

post-test whereas differences between groups of unequal variance were

determined by a KruskalndashWallis test followed by Dunnrsquos post-test

Differences were considered to be significant at pp005

f Norwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESSM Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]]4

27 Estimation of PECs according to EMEA guideline

The calculation of the predicted environmental concentration (PEC)

according to EMEA guidelines Phase I is restricted to the aquatic

compartment and was determined by the use of the following formula

PECsurface waterethmgL1THORN frac14DOSEai Fpen

WASTEWinhab DILUTION

where PECsurfacewater is the predicted environmental concentration for

surface water DOSEai the maiximum daily dose consumed per

inhabitant Fpen the market penetration factor of active ingredient

WASTEWinhab the volume of wastewater generated per inhabitant and

DILUTION is the dilution of effluent in recipient

For all of the parameters except DOSEai a recommended default value

is included in the guidelines The DOSEai is the highest recommended

dose and the nationally approved and recommended dose can be retrieved

from eg summary of product characteristics (SPCs) or drug textbooks

The highest recommended dose for each compound approved in Norway

was retrieved from the SPCs database available from the website of the

Norwegian Medicines Agency (Norwegian Medicines Agency)

Fpen the percentage of market penetration has a default value of 001

assuming that 1 of the population are treated daily with the drug The

volume of wastewater generated per inhabitant per day (WASTEWinhab)

was set to 200L inh1 d1 and the default value for dilution factor is 10

These default values given in the guidelines were used in our calculation

28 Estimation of PEC according to conventional model

The PEC in water can also be calculated according to the following

equation (Sebastine and Wakeman 2003)

PECethConventionalTHORNsurface waterethgL1THORN frac14

A eth1 R=100THORN

365 PV D

where A is the predicted amount used per year in the relevant geographic

area (kg) R the removal rate (due to loss by adsorption to sludge particles

by volatilisation by hydrolysis by biodegradation or other specific

naturally occurring processes P the number of inhabitants of the

geographic area considered V the volume of wastewater per capita and

day (m3) normally between 015 and 03m3 in EU and D is the dilution of

waste water by surface water flow (average factor 10)

Information about extent of drug use was retrieved from the Drug

Wholesale Statistics Database available at the Norwegian Institute of

Public Health This database includes the total sales of pharmaceuticals in

Norway from all wholesalers in Norway to pharmacies hospitals and non-

pharmacy outlets Data on wholesales of pharmaceuticals were retrieved

for the year 2005 The following variables were retrieved from the

database number of DDDs DDD value for each ATC code number of

packages package size Based on the sales volume in DDDs and the DDD

value the total amounts of the various compound studied were calculated

It was assumed that the entire amount of the pharmaceuticals sold was

consumed and that the amount was evenly distributed over the year and

Table 2

Calculated endpoints in the P subcapitata growth inhibition test (mean value

Concentration (mgL) Growth rate (d1)

Mean St d of control

0 (control) 203 001 100

32 208 001 102

56 210 002 103

10 208 003 102

18 206 002 101

32 210 002 103

56 210 002 103

100 204 002 100

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

throughout the population The volume of wastewater per capita was

assumed to be 200L and a dilution factor of 10 was used Further on to

assess the worst-case scenario drug metabolism in humans was

disregarded and the WTW removal rate was set to 0

3 Results

After an extensive literature research to establish thetoxicity of the selected compounds to the EMEA selectedbioassays no ecotoxicity data were available for cyclopho-sphamide The effect of cyclophosphamide on algal growthinhibition and D magna reproduction was thereforeinvestigated

31 Effects on growth of P subcapitata and reproduction of

D magna by cyclophosphamide

The growth of P subcapitata in the control cultures wasexponential throughout the 72 h exposure period with amean specific growth rate of 203 d1 The mean coefficientof variation for section-by-section growth rates (days 0ndash11ndash2 and 2ndash3) was 62 in the control cultures and the pH-increase was 01 pH unit Thus all validity criteria for thetest were fulfilled (OECD 2006) At all tested concentra-tions of cyclophosphamide (32ndash100mgL) the meanspecific growth rates were slightly higher than in thecontrols as shown in Table 2 and Fig 1 Since no inhibitionof the growth was observed within the concentration rangeof cyclophosphamide tested the effect concentrations(EC50 and NOEC) can only be expressed as 4100mgLFor the D magna reproduction test all tested animals

survived in the controls and at all test concentration except10mgL where one of the animals died after 13 days Thefirst broods of offspring appeared after 10 days in onereplicate at 10mgL and the two replicates at 18mgLAll surviving animals had produced the first brood after12 days Aborted eggs were observed in one animal at56mgL This animal did not produce any live offspringduring the test The number of live offspring produced byeach adult is shown in Fig 2 The average production ofoffspring was lower than respective controls at all testedconcentrations The statistical analysis showed howeverthat the reproductive output was significantly less than the

s of replicates)

Area under growth curve (106 cellsh l1)

Mean St d of control

40419 1527 100

47659 1162 118

52573 3397 130

48900 3808 121

44582 2371 110

50914 2546 126

51196 2255 127

42259 2091 105

f Norwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESSM Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]] 5

control only at 100mgL Thus the test yields a LOEC frac14100mgL and NOEC frac14 56mgL

32 Estimation of predicted environmental concentrations

(PECs)

The PECs for the suite of pharmaceuticals wereestimated according to the EMEA guidelines The basicphysico-chemical properties and the EMEA Phase I datarequired as well as the calculated PECs are presented in

1

10

100

1000

10000

0 20 40 60 80

Hours

10

6 c

ells

l

10

18

32

56

100

Control

mgL

Fig 1 Growth curves (P subcapitata) for control cultures and various

concentrations of cyclophosamide (mgL) (mean values of replicates)

0

20

40

60

80

100

120

140

Control 10 mgL 18 mgL

No of off

springa

nim

al

Fig 2 Response plot for effect of cyclophosphamide

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

Table 3 The PECs estimated according to the conventionalmodel are given in the table for comparisonFor all of the compounds except ethinylestradiol the

PEC was above the action limit of 001 mgL indicated inthe ERA Ethinylestradiol has endocrine disrupting prop-erties and therefore must undergo further evaluationaccording to the EMEA guidelines Diclofenac has alogKow445 and therefore it is recommended that it isscreened for persistence bioaccumulation and toxicity(PBT) according to the EU TGD Diclofenac has beenreported to photodegrade quite rapidly but the photo-transformation products has been reported to have hightoxicity potential (Schmitt-Jansen et al 2007) Howeverpursuit of the PBT of diclofenac is beyond the scope of thisproject and was not further pursued

33 Phase II assessment tier A (EMEA)

Since all the compounds had to be further evaluated arefinement was necessary for all substances This refine-ment takes into consideration the removal of the pharma-ceutical substances through adsorption and degradationA literature search was performed regarding the physico-chemical tests and aquatic effects studies recommended byEMEA The results are shown in Table 4 For all theselected pharmaceutical substances a reported value for theready biodegradability or removal rate in STP system wasavailable Since removal rates are evaluated as a goodmethod in the TGD the results of such studies wereused The results for paracetamol and ibuprofen allindicate that these compounds are readily biodegradablewith no persistence Regarding diclofenac two reportson its elimination in STPs show that the removal rate isquite low 39 in one case and 22 in another Thebiodegradability of diclofenac is 93 when performed insoil For the rest of the compounds the results show thatthe biodegradation of these compounds is relatively low

32 mgL 56 mgL 100 mgL

on the number of offspring per parent D magna

f Norwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESS

Table 3

Estimation of PECs according to the EMEA guidelines and a conventional model

Substance DOSEai(mg) LogKow pKa PEC EMEA(mgL) PECConv (mgL) PECEMEAPECConv

Cefuroxime 9000 45 0029 1579

Ciprofloxacin 1500 04 64 75 027 28

Cyclophosphamide 500a 063 25 000026 9747

Diclofenac 150 451 42 075 048 16

Ethinylestradiol 0035 415 000018 000057 03

Ibuprofen 2400 397 44 12 80 15

Metoprolol 200 188 96 10 17 06

Paracetamol 3000 046 95 15 42 04

Sulfamethoxazole 2400 089 12 0067 180

Tetracycline 1000 119 33 50 031 16

Trimethoprim 480 091 712 24 016 15

aCyclophosphamide is given as intermittent courses and is normally not administered daily

M Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]]6

The sorption behaviour (particion coefficient (Kd) oradsorption coefficient (Koc)) of all compounds apart fromcyclophosphamide has been reported All compounds withthe exception of tetracycline exhibited a low affinity fororganic carbon Since tetracycline is also below the logKoc

limit of 4 the environmental assessment of the pharma-ceutical compounds in terrestrial systems is therefore notnecessary according to the EMEA guidelines The EMEAguideline further recommends that compounds that are notreadily biodegradable should be investigated in a water-sediment study Data were only available for the twosubstances which are regarded as readily biodegradableparacetamol and ibuprofen The transformation of com-pounds in sediment systems was beyond the scope of thisproject and was therefore not further pursued Howeverthis does demonstrate that environmental data for certainpharmaceutical compounds even high use generic phar-maceuticals are not available

In Table 5 PEC values as well as refined PEC values arepresented In addition to the PECEMEA and PECconv (alsoincluded in Table 3) refined PECs correcting for adsorp-tion to particles and biodegradation are shown PECads is arefinement based upon adsorption to suspended particlesThe value for suspended solids used to calculate the PECads

is a value determined in the Oslo area (Vogelsang et al2006) The resulting refined PECs are based upondegradation of the compounds The PECs are presentedas maximum and minimum in order to account for thevariability in the available degradation rates for thesecompounds It is the refined PECs that have been used inthe final assessment

34 Calculation of predicted no-effect concentrations

(PNEC) and risk quotients

Acute and chronic toxicity effects were collected fromthe literature The results are presented in Tables 6 and 7respectively Chronic algal data were available for mostcompounds The effect of the substances on D magna

reproduction was also available (Table 7) Evaluation ofthe toxicity studies and the resulting risk quotients are

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

presented in Table 8 The PNECsurfacewater have beencalculated from the lowest NOEC or EC value presented inTables 6 and 7 using the appropriate assessment factor(AF) as recommended by the TGD The AFs are presentedalongside the PNECs in order to present the level ofconfidence in the available data The calculated riskquotients are also presented in Table 8 The derived riskquotients for the following substances are 41

f N

Ciprofloxacin

Diclofenac

Ethinylestradiol

Sulfamethoxazole

Tetracycline

The derived risk quotients based on PEC estimations bythe conventional model are also given in Table 8 forcomparison

4 Discussion

41 Estimation of PECs

An accurate estimate of the extent of drug exposure in apopulation is difficult in most countries as precise statisticsoften are lacking (Jones et al 2002 Stuer-Lauridsen et al2000) Often the statistics include information on prescribeddrugs only (ie does not include over the counter medicinesor in-hospital use of pharmaceuticals are lacking) Inaddition when a new medicinal drugs that has not been onthe market an assumption of the future sale has to be madewhen performing an ERA for the product A combination ofthese factors is probably the reason why EMEA hasrecommended to initially calculate the PECs independentof consumption statistics but rather to calculate based onthe maximum daily dose and a fixed penetration factor of1 of all drugs This penetration factor is based on a widerange of individual market penetration factors based onGerman consumption in 2001 (CHMP 2006)According to the regulation in Norway all wholesalers in

Norway should report sales volume of pharmaceuticals to

orwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESS

Table

4

Available

physico-chem

icalproperties

andbiodegradationstudies

Substance

Adsorptionusingabatchequilibrium

method

ReadyBiodegradabilitytest

Transform

ationsedim

ent

system

s

Respirationinbibitiontest

Kdabs

Reference

Koc

Reference

transform

ation

orDT50(d)

Reference

DT50(d)

Reference

IC50(m

gL1)

Reference

Cefuroxim

e109ndash119

GlaxoSmithKline

(2004)

28

GlaxoSmithKline(2004)

4100

Kummerer

etal(2004)

1

Alexyet

al(2004)

4100

GlaxoSmithKline(2004)

Ciprofloxacin

430

Tolls(2001)

61000a

Tolls(2001)

0

Kummerer

etal(2000)

008

Tolls(2001)

417

Halling-Sorensenet

al

(2000)

16ndash25

Halling-Sorensenet

al(2000)

Cyclophosphamide

0

Steger-H

artmannet

al(1997)

Negligible

0

Kummerer

etal(1996)

Diclofenac

4ndash10

Kreuziget

al(2003)

2310

Drillia

etal(2005)

93

Kreuziget

al(2003)

1645

bDrillia

etal(2005)

39

cPaxeus(2004)

045d

Drillia

etal(2005)

22

cBendzet

al(2005)

Ethinylestradiol

40

Mastrupet

al(2005)

691

Clara

etal(2004)

20

Ternes

etal(1999)

1

Mastrup

etal(2005)

Ibuprofen

6ndash64

Kreuziget

al(2003)

10ndash20

Jones

etal(2006)

oLOQ

28d

Loffler

etal(2005)

17ndash9

Loffler

etal(2005)

324

Jones

etal(2006)

490

cPaxeus(2004)

453

10ndash60

EuropeanCommissionmdash

EuropeanChem

icalsBureau

(2000a)

o6

Loffler

etal

(2005)

Metoprolol

96ndash376

e0

cBendzet

al(2005)

3ndash8

Jones

etal(2006)

2803e

o10

cPaxeus(2004)

Negligible

Paracetamol

36ndash45

Kreuziget

al(2003)

Low

persist

Loffler

etal(2005)

1ndash10

Jones

etal(2006)

62

Jones

etal(2006)

57

Henschel

etal(1997)

64ndash53

Loffler

etal(2005)

99

EuropeanCommissionmdash

EuropeanChem

icalsBureau

(2000b)

31

Loffler

etal

(2005)

Sulfamethoxazole

376

Drillia

etal(2005)

0

Al-Ahmadet

al(1999)

4100

Kummerer

etal(2004)

023

Drillia

etal(2005)

530

Drillia

etal(2005)

4

Alexyet

al(2004)

256

Tolls(2001)

Tetracycline

8400

Kim

etal(2005)

0ndash62

Vaclavik

etal(2004)

Tim

ecurve

Vaclavik

etal(2004)

1140ndash1620

Sithole

andGuy(1987)

6059

Tolls(2001)

2

Alexyet

al(2004)

1ndash10

Kummerer

etal(2004)

Trimethoprim

22ndash41d

Halling-Sorensenet

al(2000)

76

Halling-Sorensenet

al

(2000)

4

Alexyet

al(2004)

4100

Kummerer

etal(2004)

aSoil

bHighorganic

content

cRem

ovalrate

STP

dLow

organic

content

eData

forpropranolol

M Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]] 7

Please cite this article as Grung M et al Environmental assessment of Norwegian priority pharmaceuticals based on the EMEA guideline

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

ARTICLE IN PRESS

Table 5

Refined PEC according to Phase II guidelines and determined PECPNEC ratios

Compound PEC EMEA

(mlL)PEC Conv

(mgL)Kd

(Lkg)

Refined PECads Biodeg EMEA Conv

EMEA Conv Max

()

Min

()

Refined

PEC min

Refined

PEC max

Refined

PEC min

Refined

PEC max

Cefuroxime 45 0029 45 0029 28 0 32 45 0021 0029

Ciprofloxacin 75 027 417 7 025 0 0 70 70 025 025

Cyclophosphamide 25 000026 3 000026 0 0 25 25 000026 000026

Diclofenac 075 048 165 1 046 93 22 0051 057 0032 036

Ethinylestradiol 0000175 000057 691 0 000051 20 0 000013 000016 000041 000051

Ibuprofen 12 80 453 11 74 90 10 11 10 074 67

Metoprolol 1 17 37 1 17 10 0 089 099 15 17

Paracetamol 15 42 36 15 42 99 57 015 64 042 18

Sulfamethoxazole 12 0067 38 12 0066 4 0 11 12 0063 0066

Tetracycline 5 031 8400 2 013 62 0 078 21 0049 013

Trimethoprim 24 016 76 2 016 4 0 227 237 015 016

PECAds Refinement made based upon partition coefficient (Kd) and assuming a suspended solid concentration of 17mgL (based on Vogelsang et al

2006)

M Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]]8

the Norwegian Institute of Public Health The NorwegianPrescription Database (NorPD) was established in Norwayin 2004 The NorPD includes information of all prescrip-tions on medicines dispensed in a pharmacy in Norway(Furu et al 2005) Therefore Norway has detailedinformation regarding sales and use of pharmaceuticalsIn Table 1 the penetration factor for the elevencompounds in this report can be seen as the DDD100personsday represents the percentage of the Norwegianpopulation that are being treated daily with thesecompounds The sales figures indicate that Fpen is above1 in Norway for 4 of the compounds (ethinylestradiolibuprofen metoprolol and paracetamol) It can also beconcluded that that the use of some of the compounds arelimited eg cefuroxime and cyclophosphamide

The conventional model (Sebastine and Wakeman 2003)of estimating the PEC may be more closely linked to thetrue emission numbers assuming that sold pharmaceuticalcompounds are actually used From the ERA in Sweden in2004 (Swedish Medical Products Agency 2004) dataindicate that 90 of pharmaceuticals sold in Sweden areconsumed so this seems to be a fair assumption From thecomparison of the two estimated PECs it can be seen thatthe PECEMEA for a number of pharmaceuticals is quitesimilar to PECconv however for others there are significantdifferences For example the PECEMEA provided lowerestimations than the PECconv for paracetamol ibuprofenand ethinylestradiol These three compounds are frequentlyused pharmaceuticals with a market penetration of morethan 1 For the less frequently used pharmaceuticals thePECEMEA is higher than the PECconv This is particularlytrue for the anticancer agent cyclophosphamide which iscertainly not used by 1 of the population The differencebetween the two PECs for the antibacterial cefuroxime isalso large However for the estimation of a new drug onthe market the use of a market penetration of 1 seems

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

reasonable and will give an indication of which PEC levelto expect after introduction onto the marketIn both models of PEC estimation the daily wastewater

per capita was set to 200L Data from 2003 (StatisticsNorway 2005) show that the actual use of wastewater inNorway was slightly higher 216L However the difference issmall the pharmaceuticals will be more diluted by the higherwater consumption than predicted so the estimated PECswill be slightly too high In this study we have used onlystatistics including sales of medicines for use in humans Thesale of veterinary medicines only approved for use in animalsis not included This is the case for trimethoprim andconsequently the PECs estimated for this compound will beunderestimated Estimates of PECs both according to theEMEA guidelines and the conventional model are conserva-tive and probably do not reflect realistic environmentalconcentrations The numbers should therefore only be used asrough estimates and should always be followed by measuredenvironmental concentrations (MECs)

42 PNEC values

A lack of data on toxicity is the primary obstacle forassessing the environmental risk of pharmaceuticals Whilestandard acute toxicity tests play an important role in atiered approach to ERA the lack of chronic toxicity datafor many pharmaceutical compounds is a major hindranceto their effective risk assessment Because of their low butpersistent occurrence pharmaceuticals will most likelyhave chronic rather than acute toxic effects It is likelythat the lack of chronic toxicity is due to the recentemergence of the problem of pharmaceuticals in theenvironment and the large number of pharmaceuticalsubstances used by societyComparing acute toxicity (Table 6) and chronic toxicity

(Table 7) it is evident that more compounds have been

f Norwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESS

Table 6

Acute toxicity effects on algae invertebrates and fish

Algae Invertebrate Fish

EC50 Species Reference EC50 Species Reference LC50 Species Reference

Cefuroxime 491 P subcapitata GlaxoSmithKline

(2004)

41000 D magna GlaxoSmithKline

(2004)

4120 O mykiss GlaxoSmithKline

(2004)

Ciprofloxacin 0005 M aeruginosa Halling-Sorensen et

al (2000)

460 D magna Halling-Sorensen et

al (2000)

4100 B rerio Halling-Sorensen et

al (2000)

297 P subcapitata Halling-Sorensen et

al (2000)

0017 M aeruginosa Robinson et al

(2005)

0203 L minor Robinson et al

(2005)

187 P subcapitata Robinson et al

(2005)

Cyclophosphamide 4100 P subcapitata This report 4100 D magna This report

Diclofenac 72 D subspicatus Cleuvers (2004) 68 D magna Cleuvers (2004)

16 P subcapitata Ferrari et al (2004) 41 T

platyurus

Nalecz-Jawecki and

Persoone (2006)

19 C meneghiniana Ferrari et al (2004) 80 D magna Han et al (2006)

145 S leopoliensis Ferrari et al (2004) 22 D magna Ferrari et al (2004)

23 C dubia Ferrari et al (2004)

Ethinylestradiol 4 D magna Goto and Hiromi

(2003)

Ibuprofen 4 Lemna Pomati et al (2004)

342 D subspicatus Cleuvers (2004) 101 D magna Cleuvers (2004)

Metoprolol 320 Lemna Cleuvers (2005) 775 T patyurus Nalecz-Jawecki and

Persoone (2006)

4100 Medaka Nalecz-Jawecki and

Persoone (2006)

79 D subspicatus Cleuvers (2005) 639 D magna Nalecz-Jawecki and

Persoone (2006)

88 C dubia Nalecz-Jawecki and

Persoone (2006)

64 D magna Dzialowski et al

(2006)

Paracetamol 134 S subcapitata Henschel et al (1997) 50 D magna Henschel et al

(1997)

378 B rerio Henschel et al

(1997)

92 D magna (Kuhn et al 1989)

Sulfamethoxazole 0146 P subcapitata Ferrari et al (2004) 4100 D magna Ferrari et al (2004) 41000 B rerio Isidori et al (2005)

24 C meneghiniana Ferrari et al (2004) 4100 C dubia Ferrari et al (2004)

0027 S leopoliensis Ferrari et al (2004) 26 B

caliciflorus

Isidori et al (2005)

057 P subcapitata Isidori et al (2005) 155 C dubia Isidori et al (2005)

25 D magna Isidori et al (2005)

Tetracycline 16 N closterium Peterson et al (1993) 4340 D magna Wollenberger et al

(2000)

220 S namaycush Cunningham et al

(2006)

22 S capricornutum Halling-Sorensen

(2000)

009 M aeruginosa Halling-Sorensen

(2000)

Trimethoprim 110 P subcapitata Halling-Sorensen et

al (2000)

123 D magna Halling-Sorensen et

al (2000)

4100 B rerio Halling-Sorensen et

al (2000)

112 M aeruginosa Halling-Sorensen et

al (2000)

16 R salina Lutzhoft et al (1999)

Numbers in bold indicate lowest value

M Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]] 9

investigated by the former tests For many of the selectedcompounds in this study chronic toxicity data are absentespecially for fish An exception is ethinylestradiol anddiclofenac for which several studies have shown its high

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

toxicity to fish (Table 7) Long-term data are based on sexratio for ethinylestradiol and histopathology and cyto-pathology for diclofenac Hence the use of the NOECfrom these studies must be regarded as conservative and

f Norwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESS

Table 7

Chronic toxicity effects and sublethal effects on algae invertebrates and fish

Alga Daphnia Fish

NOEC Species Reference NOEC

EC10

Species Reference NOEC Species Reference

Cefuroxime 91 P subcapitata GlaxoSmithKline (2004)

Ciprofloxacin

Cyclophosphamide 4100 P subcapitata This study 56 D magna This study

Diclofenac 49 D subspicatus Cleuvers (2004) 4 D rerio Ferrari et al (2004)

10 P subcapitata Ferrari et al (2004) 10 D magna Han et al (2006) 05 D rerio Hallare et al (2004)

10 C menheginiana Ferrari et al (2004) 1 C dubia Ferrari et al (2004) 00015 S trutta Hoeger et al (2005)

10 S leopoliensis Ferrari et al (2004) 0005 O mykiss Schwaiger et al (2004)

o0001 O mykiss Triebskorn et al (2004)

Ethinylestradiol 002 D magna Goto and Hiromi (2003) 0000001 D rerio Orn et al (2003)

Ibuprofen o0001 Lemna Pomati et al (2004)

41 Synechocystis sp Pomati et al (2004) 20 D magna Han et al (2006)

103 D subspicatus Cleuvers (2004) 102 Pcarinatus Pounds et al (2004)

Paracetamol

Metoprolol 31 D magna Dzialowski et al (2006)

Sulfamethoxazole 001 L gibba Brain et al (2004)

009 P subcapitata Ferrari et al (2004) 021a C dubia Isidori et al (2005)

125 C meneghiniana Ferrari et al (2004) 963a B caliciflorus Isidori et al (2005)

00059 S leopolensis Ferrari et al (2004) 025 C dubia Ferrari et al (2004)

Tetracycline 294 D magna Wollenberger et al (2000)

Trimethoprim

Numbers in bold indicate lowest valueaChronic EC50 value

Table 8

Calculated risk quotients based on PEC estimations by EMEA and refined by adsorption to particles and biodegradation

Lowest NOEC

EC50 (mgL)

AF PNEC (mgL) EMEA PECPNEC Conventional PECPNEC

Min Max Min Max

Cefuroxime 91 1000 91 036 049 000024 000033

Ciprofloxacin 0005 1000 0005 1401 1401 52 52

Cyclophosphamide 56 50 1120 00022 00022 24E07 24E07

Diclofenac 0001 10 01 051 57 034 38

Ethinylestradiol 100E06 10 00001 13 16 43 54

Ibuprofen 1 50 20 0056 050 0039 035

Metoprolol 31 100 31 0029 0032 0051 0057

Paracetamol 92 1000 92 0016 070 0048 20

Sulfamethoxazole 00059 50 0118 97 101 057 059

Tetracycline 009 1000 009 87 23 057 15

Trimethoprim 16 1000 16 014 015 0010 0010

AF Assessment Factor

M Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]]10

needs further evaluation As a consequence diclofenac andethinylestradiol are the only compounds with an assess-ment factor of 10 (Table 8)

Regarding the effects of ibuprofen we have not used thelowest observed NOEC of 1 mgL due to the fact thatPomati et al (2004) did not replace the medium daily andthat the result is substantially lower than other toxicitystudies with the same organism (Brain et al 2004Kummerer 2001)

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

43 Comparison of PEC with MEC

The effluent concentrations of the selected pharmaceu-ticals have recently been measured in an STP near Oslo(Thomas et al in press) The measured effluent concentra-tions (MECs) can be used to generate further refinedquasi-MECs in which we have greater confidence Com-parison of the refined PEC with quasi-MECs provides anindication of how reliable the PEC determinations

f Norwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESS

Table 9

Comparison of median and maximal quasi-measured environmental

concentrations (Quasi-MEC) with predicted no-effect concentrations

(PNEC)

Pharmaceutical Quasi-MECa (ngL) PNEC (ngL) Quasi-MECPNEC

Median Maximal Median Maximal

Cefuroxime nd nd 91 000 ndash ndash

Ciprofloxacin 4 74 5 08 15

Cyclophosphamide nd nd 1 120 000 ndash ndash

Diclofenac 26 37 100 020 028

Ethinylestradiol nd nd 01 ndash ndash

Ibuprofen 4 62 20 000 2 104 31 103

Metoprolol 65 77 31 000 2 103 25 103

Paracetamol 3 432 9200 32 104 005

Sulfamethoxazole 16 21 118 014 018

Tetracycline nd nd 90 ndash ndash

Trimethoprim 68 126 16 000 43 104 79 104

Numbers in bold indicate risk quotient 41

nd compound not detected in any final effluent sample collected

ndash No risk quotient calculated due to insufficient dataaEffluent concentration divided by a dilution factor of 10

M Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]] 11

including refinements are The concentrations of theselected compounds were measured in influent and effluentto an Oslo STP (VEAS) during a 7-week period from 9thAugust 2006 to 20th September 2006 The measuredmedian effluent values were converted into quasi-MECsusing a dilution factor of 10 (Table 9) Keeping in mindthat this is one local measurement in comparison to PECsestimated for the country as such this is still useful

The quasi-MECs are as expected considerably lowerthan the PECs (estimated by both EMEA guideline andliterature model) None of the compounds has a quasi-MEC 4PEC which indicates that the PEC models are veryconservative and precautionary However some cautionmust be used when assessing the data since they aremedian values for one STP only (VEAS Oslo Norway)For ciprofloxacin the maximal quasi-MEC had a riskquotient 41 and also the median quasi-MEC was only alittle higher than the PNEC so this compound should befurther evaluated in later ERAs and monitoring campaignsin Norway Three of the compounds could not be detectedin the effluents and for cefuroxime and cyclophosphamidethe limit of detection are well below the PNEC value Sothese compounds probably do not represent a problem inthe environment However for ethinylestradiol the limit ofdetection is comparable to the PNEC and consequentlythis compound should also be monitored more closely inthe future

5 Conclusions

P

E

The EMEA guideline provides a useful tool to assess therisks associated with pharmaceuticals entering theaquatic environment through STPs

PECs calculated according to the EMEA guidelines are

conservative and precautionary even following refine-

lease cite this article as Grung M et al Environmental assessment of N

cotoxicol Environ Saf (2007) doi101016jecoenv200710015

ment The absence of good reliable and consistent fatedata makes PEC refinement difficult This is veryimportant when the resultant risk quotient is 1 Itmay be more suitable in Norway to use a more specificapproach due to the availability of data regarding salesand use of pharmaceuticals

MECs will always provide better estimates for the

calculation of environmental occurrence and thereforewe recommend the use of MECs where possible inpharmaceutical ERA

The major weakness in any ERA of pharmaceutical is

the PNEC PNECs based on mortality even whenexposure is chronic may not be sufficient to protect theaquatic environment Exposure in the environment islikely to be chronic and the effects sub-lethal Werecommend that better strategies are developed to dealwith this

Basic ecotoxicity data are not available for many

pharmaceuticals (eg cyclophosphamide) The use ofbasic D magna and algae data provide data for simpleERA but are not sufficient for safe-guarding againstlong-term sub-lethal effects since at present the effectsare not known

Based on the EMEA guidelines ciprofloxacin diclofe-

nac ethinylestradiol sulfamethoxazole and tetracyclinehave a risk coefficients 41 For ciprofloxacin themaximal quasi-MEC was larger than the PNEC andtherefore this compound may pose a threat to theaquatic environment The substance should thereforebe monitored in the future

For many of the selected compounds in this study

chronic toxicity data were absent especially for fishThis lack of data is a major hindrance to performingERA of pharmaceuticals We recommend furthersupplementation of chronic toxicity data for pharma-ceuticals in the open literature to facilitate a betterassessment of the risk these compounds pose to theenvironment

Acknowledgments

Part of this study was performed by NIVA oncommission from Norwegian Pollution Control Authority(TA 22162006) Part of this study was performed as partof a grant from the Norwegian Research Council to NIVA(Project number 172526 Fate risk and management ofpharmaceuticals and personal care products (PPCPs) in theNorwegian sewage system)

References

Al-Ahmad A Daschner F Kummerer K 1999 Biodegradability of

cefotiam ciprofloxacin meropenem penicillin G and sulfamethox-

azole and inhibition of waste water bacteria Arch Environ

Contamin Toxicol 37 158ndash163

Alexy R Kumpel T Kummerer K 2004 Assessment of degradation

of 18 antibiotics in the Closed Bottle Test Chemosphere 57 505ndash512

orwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESSM Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]]12

Baird DJ Barber I Soares AM Calow P 1991 An early life-stage

test with Daphnia magna Straus an alternative to the 21-day chronic

test Ecotoxicol Environ Saf 22 1ndash7

Bendz D Paxeus NA Ginn TR Loge FJ 2005 Occurrence and

fate of pharmaceutically active compounds in the environment a case

study Hoje River in Sweden J Hazard Mater 122 195ndash204

Brain RA Johnson DJ Richards SM Sanderson H Sibley PK

Solomon KR 2004 Effects of 25 pharmaceutical compounds to

Lemna gibba using a seven-day static-renewal test Environ Toxicol

Chem 23 371ndash382

Carlsson C Johansson AK Alvan G Bergman K Kuhler T

2006a Are pharmaceuticals potent environmental pollutants Part I

environmental risk assessments of selected active pharmaceutical

ingredients Sci Total Environ 364 67ndash87

Carlsson C Johansson AK Alvan G Bergman K Kuhler T

2006b Are pharmaceuticals potent environmental pollutants Part II

environmental risk assessments of selected pharmaceutical excipients

Sci Total Environ 364 88ndash95

Castiglioni S Bagnati R Fanelli R Pomati F Calamari D

Zuccato E 2006 Removal of pharmaceuticals in sewage treatment

plants in Italy Environ Sci Technol 40 357ndash363

Clara M Strenn B Saracevic E Kreuzinger N 2004 Adsorption of

bisphenol-A 17 beta-estradiole and 17 alpha-ethinylestradiole to

sewage sludge Chemosphere 56 843ndash851

Cleuvers M 2004 Mixture toxicity of the anti-inflammatory drugs

diclofenac ibuprofen naproxen and acetylsalicylic acid Ecotoxicol

Environ Safe 59 309ndash315

Cleuvers M 2005 Initial risk assessment for three b-blockers found in

the aquatic environment Chemosphere 59 199ndash205

Committee for Medicinal Products for Human Use (CHMP) 2006

European Medicines Agency Pre-Authorisation Evaluation of Medi-

cines for Human Use (Doc Ref EMEACHMPSWP444700)

Guideline on the environmental risk assessment of medicinal products

for human use 12pp

Cunningham VL Buzby M Hutchinson T Mastrocco F Parke N

Roden N 2006 Effects of human pharmaceuticals on aquatic life

next steps Environ Sci Technol 40 3456ndash3462

Drillia P Stamatelatou K Lyberatos G 2005 Fate and mobility of

pharmaceuticals in solid matrices Chemosphere 60 1034ndash1044

Dzialowski E Turner P Brooks B 2006 Physiological and

reproductive effects of beta adrenergic receptor antagonists in

Daphnia magna Arch Environ Contamin Toxicol 50 503ndash510

European CommissionmdashEuropean Chemicals Bureau 2000a IUCLID

dataset ibuprofen 45pp

European CommissionmdashEuropean Chemicals Bureau 2000b IUCLID

dataset paracetamol 76pp

European Commission 2003 Technical Guidance Document on Risk

AssessmentmdashCommission Regulation (EC) No 148894 on Risk

Assessment for existing substances 337pp

Fent K Weston AA Caminada D 2006 Ecotoxicology of human

pharmaceuticals Aquat Toxicol 76 122ndash159

Ferrari B Mons R Vollat B Fraysse B Paxeus N Lo Giudice R

Pollio A Garric J 2004 Environmental risk assessment of six

human pharmaceuticals Are the current environmental risk assess-

ment procedures sufficient for the protection of the aquatic environ-

ment Environ Toxicol Chem 23 1344ndash1354

Furu K Stroslashm H Roslashnning M Skurtveit S Engeland A Tverdal

A 2005 The Norwegian prescription database (NorPD)mdasha new

register for pharmacoepidemiologic research covering a whole nation

Pharmacoepidemiol Drug Safe 14 S49

GlaxoSmithKline 2004 Safety data sheet cefuroxime 7pp

Goto T Hiromi J 2003 Toxicity of 17alpha-ethynylestradiol and

norethindrone constituents of an oral contraceptive pill to the

swimming and reproduction of cladoceran Daphnia magna with special

reference to their synergetic effect Mar Pollut Bull 47 139ndash142

Hallare AV Kohler HR Triebskorn R 2004 Development toxicity

and stress protein responses in zebrafish embryos after exposure to

diclofenac and its solvents DMSO Chemosphere 56 659ndash666

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

Halling-Sorensen B 2000 Algal toxicity of antibacterial agents used in

intensive farming Chemosphere 40 731ndash739

Halling-Sorensen B Lutzhoft H Andersen H Ingerslev F 2000

Environmental risk assessment of antibiotics comparison of mecilli-

nam trimethoprim and ciprofloxacin J Antimicrob Chemother 46

53ndash58

Han G Hur H Kim S 2006 Ecotoxicological risk of pharmaceuticals

from wastewater treatment plants in Korea Occurrence and toxicity to

Daphnia magna Environ Toxicol Chem 25 265ndash271

Henschel KP Wenzel A Diedrich M Fliedner A 1997 Environ-

mental hazard assessment of pharmaceuticals Regul Toxicol

Pharmacol 25 220ndash225

Hill AV 1910 The possible effects of aggregation of the molecules

of hemoglobin on its dissociation curves J Physiol (Lond) 40

IVndashVII

Hoeger B Kollner B Dietrich DR Hitzfeld B 2005 Water-borne

diclofenac affects kidney and gill integrity and selected immune

parameters in brown trout (Salmo trutta f fario) Aquat Toxicol 75

53ndash64

Isidori M Lavorgna M Nardelli A Pascarella L Parrella A 2005

Toxic and genotoxic evaluation of six antibiotics on non-target

organisms Sci Total Environ 346 87ndash98

Jones OA Voulvoulis N Lester JN 2002 Aquatic environmental

assessment of the top 25 English prescription pharmaceuticals Water

Res 36 5013ndash5022

Jones OA Voulvoulis N Lester JN 2006 Partitioning behavior of

five pharmaceutical compounds to activated sludge and river sediment

Arch Environ Contam Toxicol 50 297ndash305

Kim S Eichhorn P Jensen J Weber A Aga D 2005 Removal of

antibiotics in wastewater Effect of hydraulic and solid retention times

on the fate of tetracycline in the activated sludge process Environ Sci

Technol 39 5816ndash5823

Kreuzig R Kullmer C Matthies B Holtge S Dieckmann H 2003

Fate and behaviour of pharmaceutical residues in soils Fresen

Environ Bull 12 550ndash558

Kuhn R Pattard M Pernak K Winter A 1989 Results of the

harmful effects of selected water pollutants (anilines phenols

aliphatic-compounds) to Daphnia-Magna Water Research 23

495ndash499

Kummerer K 2001 Pharmaceuticals in the EnvironmentmdashSources

Fate Effects and Risks Springer Berlin

Kummerer K StegerHartmann T Baranyai A Burhaus I 1996

Examination of the biodegradation of the antineoplastics cyclopho-

sphamide and ifosfamide with the closed bottle test (OECD 301 D)

Zbl Hyg Umweltmed 198 215ndash225

Kummerer K Al-Ahmad A Mersch-Sundermann V 2000 Biode-

gradability of some antibiotics elimination of the genotoxicity and

affection of wastewater bacteria in a simple test Chemosphere 40

701ndash710

Kummerer K Alexy R Huttig J Scholl A 2004 Standardized tests

fail to assess the effects of antibiotics on environmental bacteria Water

Res 38 2111ndash2116

Loffler D Rombke J Meller M Ternes TA 2005 Environmental

fate of pharmaceuticals in watersediment systems Environ Sci

Technol 39 5209ndash5218

Lutzhoft HCH Halling-Sorensen B Jorgensen SE 1999 Algal

toxicity of antibacterial agents applied in Danish fish farming Arch

Environ Contamin Toxicol 36 1ndash6

Mastrup M Schafer AI Khan SJ 2005 Predicting fate of the

contraceptive pill in wastewater treatment and discharge Water Sci

Technol 52 279ndash286

Nalecz-Jawecki G Persoone G 2006 Toxicity of selected pharmaceu-

ticals to the anostracan crustacean Thamnocephalus platyurusmdash

comparison of sublethal and lethal effect levels with the 1 h

Rapidtoxkit and the 24 h Thamnotoxkit microbiotests Environ Sci

Pollut Res 13 22ndash27

Norwegian Institute of Public Health 2007 Drug Consumption in

Norway 2002ndash2006 httpwwwlegemiddelforbruknoenglishS

f Norwegian priority pharmaceuticals based on the EMEA guideline

ARTICLE IN PRESSM Grung et al Ecotoxicology and Environmental Safety ] (]]]]) ]]]ndash]]] 13

Norwegian Medicines Agency Summary of Product Charactereristics

(SPCs) database httpwwwlegemiddelverketnotemplates

InterPage____14833aspxS

Norwegian Pollution Control Authority 2005 Kartlegging av omsetning

av enkelte miljoslashskadelige stoffer i legemidler og kosmetikkmdashTA-2128

2005 ISBN82-7655-496-2

OECD 1998 Daphnia magna reproduction test OECD Guidelines for the

testing of chemicals Paris

OECD 2006 Freshwater alga and cyanobacteria growth inhibition test

OECD Guidelines for the testing of chemicals Paris

Orn S Holbech H Madsen TH Norrgren L Petersen GI 2003

Gonad development and vitellogenin production in zebrafish (Danio

rerio) exposed to ethinylestradiol and methyltestosterone Aquat

Toxicol 65 397ndash411

Paxeus N 2004 Removal of selected non-steroidal anti-inflammatory

drugs (NSAIDs) gemfibrozil carbamazepine beta-blockers trimetho-

prim and triclosan in conventional wastewater treatment plants in five

EU countries and their discharge to the aquatic environment Water

Sci Technol 50 253ndash260

Peterson S Batley G Scammell M 1993 Tetracycline in antifouling

paints Mar Pollut Bull 26 96ndash100

Pomati F Netting AG Calamari D Neilan BA 2004 Effects of

erythromycin tetracycline and ibuprofen on the growth of Synecho-

cystis sp and Lemna minor Aquat Toxicol 67 387ndash396

Pounds NA MacIean S Webley M Pascoe D Hutchinson T

2004 Ibuprofen evaluation of chronic growth and reproductive effects

in the freshwater pulmonate mollusc Planorbis carinatus SETAC

North America

Robinson AA Belden JB Lydy MJ 2005 Toxicity of fluoroquinolone

antibiotics to aquatic organisms Environ Toxicol Chem 24 423ndash430

Schmitt-Jansen M Bartels P Adler N Altenburger R 2007

Phytotoxicity assessment of diclofenac and its phototransformation

products Anal Bioanal Chem 387 1389ndash1396

Schwaiger J Ferling H Mallow U Wintermayr H Negele RD

2004 Toxic effects of the non-steroidal anti-inflammatory drug

diclofenac Part 1 histopathological alterations and bioaccumulation

in rainbow trout Aquat Toxicol 68 141ndash150

Sebastine I Wakeman R 2003 Consumption and environmental

hazards of pharmaceutical substances in the UK Process Saf Environ

Protect 81 229ndash235

Sithole B Guy R 1987 Models for tetracycline in aquatic environ-

ments 1 Interaction with bentonite clay systems Water Air Soil

Pollut 32 303ndash314

Statistics Norway 2005 Water resources and water pollution 19pp

httpwwwssbnoenglishsubjects01sa_nrmnrm2005kap8-water

pdfS

Steger-Hartmann T Kummerer K Hartmann A 1997 Biological

degradation of cyclophosphamide and its occurrence in sewage water

Ecotoxicol Environ Safe 36 174ndash179

Please cite this article as Grung M et al Environmental assessment o

Ecotoxicol Environ Saf (2007) doi101016jecoenv200710015

Stuer-Lauridsen F Birkved M Hansen LP Lutzhoft HC Halling-

Sorensen B 2000 Environmental risk assessment of human

pharmaceuticals in Denmark after normal therapeutic use Chemo-

sphere 40 783ndash793

Swedish Medical Products Agency 2004 Environmental effects of

pharmaceuticals cosmetics and hygiene products Excerpts from the

Report of Swedish Medical Products Agency regarding the environ-

mental effects of pharmaceuticals 60pp

Ternes TA Kreckel P Mueller J 1999 Behaviour and occurrence

of estrogens in municipal sewage treatment plantsmdashII Aerobic

batch experiments with activated sludge Sci Total Environ 225

91ndash99

Thomas KV Dye C Schlabach M Langford KH in press Source

to sink tracking of selected human pharmaceuticals doi101039

b709745j

Thomas KV Langford KH Grung M Schlabach M Dye C 2007

Occurrence of selected pharmaceuticals in wastewater effluents from

hospitals (Ulleval and Rikshospitalet) and VEAS wastewater treat-

ment works Report Norwegian Pollution Control Authority TA

22462007 ISBN978-82-577-5111-1 36pp

Tolls J 2001 Sorption of veterinary pharmaceuticals in soils a review

Environ Sci Technol 35 3397ndash3406

Triebskorn R Casper H Heyd A Eikemper R Kohler HR

Schwaiger J 2004 Toxic effects of the non-steroidal anti-inflamma-

tory drug diclofenac Part II Cytological effects in liver kidney gills

and intestine of rainbow trout (Oncorhynchus mykiss) Aquat

Toxicol 68 151ndash166

US Food and Drug Administration 1998 R Office of Training and

Communications MD Guidance for industry for submission of an

environmental assessment in human drug applications and supple-

ments CMC6 July Revision 1 39pp

Vaclavik E Halling-Sorensen B Ingerslev F 2004 Evaluation of

manometric respiration tests to assess the effects of veterinary

antibiotics in soil Chemosphere 56 667ndash676

Vasskog T Berger U Samuelsen P Kallenborn R Jensen E 2006

Selective serotonin reuptake inhibitors in sewage influents and effluents

from Tromso Norway J Chromatogr A 1115 187ndash195

Vieno N Tuhkanen T Kronberg L 2007 Elimination of pharma-

ceuticals in sewage treatment plants in Finland Water Res 41

1001

Vogelsang C Grung M Jantsch T Tollefsen K Liltved H 2006

Occurrence and removal of selected organic micropollutants at

mechanical chemical and advanced wastewater treatment plants in

Norway Water Research 40 3559ndash3570

WHO Collaborating Centre for Drug Statistics Methodology http

wwwwhoccnoS

Wollenberger L Halling-Sorensen B Kusk KO 2000 Acute and

chronic toxicity of veterinary antibiotics to Daphnia magna Chemo-

sphere 40 723ndash730

f Norwegian priority pharmaceuticals based on the EMEA guideline