179
8
Plant Uptake of
Pharmaceuticals
from Soil
Determined by ELISA
Rudolf J. Schneider
Contents
8.1 Introduction 180
8.2 Background 180
8.2.1 CodepositionofVeterinaryAntibioticswithManuringPractices .180
8.2.2 Pharmaceuticals for Human Use in Soil 181
8.2.3 Fate of Pharmaceuticals in Soil 182
8.2.4 Fate of Pha r mac euticals i n Plants 183
8.2.4.1 Uptake 183
8.2.4.2 Detoxication 184
8.2.5 Analysis of Pharmaceutical Residues in Soil and Plants 184
8.2.5.1 Extraction 184
8.2.5.2 Determination of Antimicrobial Residues 184
8.3 Materials and Methods 185
8.3.1 Sulfonamides and Plants Studied 185
8.3.2 Model Soils 185
8.3.3 Urine 186
8.3.4 Soil Extraction 186
8.3.5 Plant Extraction 187
8.3.6 Immunoassay 187
8.3.7 Laboratory Study: Sorption and Microbial Degradation in Soil 188
8.3.8 Greenhouse Study: Leaching versus Uptake 189
8.4 Results 190
8.4.1 ELISA Measurements and Extraction Recovery 190
8.4.2 Degradation and Sorption 190

um, and potassium, is well established and is the basis of understanding and man-
agi
ng plant production.Sim i
larly, knowledge on the uptake of soil-applied pesticides
is also huge because it is essential for designing and assessing the efcacy of crop
protectionagents.However,studiesonplantuptakeofpharmaceuticalsarelimited
todate.Therefore,thispaperdescribesresultsofexperimentsthatstudyplantuptake
of two sulfonamide antimicrobials: sulfamethazine and sulfamethoxazole—the rst
beingaveterinarydrugandthelatterusedinhumantherapy.Modelexperiments
examining degradation, leaching, and uptake were performed. Concentrations in
water, soil, and plants were analyzed using two selective enzyme immunoassays
(ELISA).
8.2 BACKGROUND
8.2.1 C
ODEPOSITION OF VETERINARY ANTIBIOTICS WITH MANURING PRACTICES
Millions of chemical compounds are present in the atmosphere, mostly of natural ori-
gin,butthousandsofthemareofxenobioticnature.Bywetanddrydeposition,often
adsorbed on aerosol particles and traveling over large distances, they are deposited
onto the soil surface. For most plants the top soil layer is the most important inter
-
fa
cewiththeirenvironment,providingsupportandprotectionoftherootsand,atthe
sametime,water,nutrients,andsomeotherlow-molecularweightcompounds.
Thereareotherintentionaldepositionsofchemicalsubstancesthataretradition
-
al
ly carried out, especially the application of fertilizer or pesticides on agricultural
elds.Theapplicationsofplantresiduessuchasstrawandmulchandofnutrient-rich
residuesfromlivestockbreedingasdungandliquidmanureareinmanyregionsof
the world widely used to fertilize the soil in order to ll up the soil’s nutrient res

8
found
tetracyclines in concentrations between 5 and 870 µg L
–1
in all of 13 studied swine
manure storage tanks. Other studies found concentrations of tetracycline up to 66 g
m
–3
inswinemanurewithadegradationoflessthan5%after7weeks.
5
Sulfonamides
arestableinmanureuntilapplication,too,
9
especially with nonoptimal conditions
(e.g., cold weather, anaerobic conditions),
10,11
buthighertemperaturesandadapta-
tion of the microorganisms enhance degradation.
12
Metabolites (e.g., the glucuro-
nide of chloramphenicol or N-4-acetylsulfamethazine) can be cleaved in manure and
thus transformed back to the active ingredients, such that their concentrations may
increase with time.
13
As manure is usually collected and stored for several weeks,
only compounds that resist degradation for this period will enter the soil compart-
me
ntwhenmanureisappliedtosoil(Figure 8.1).
8.2.2 PHARMACEUTICALS FOR HUMAN USE IN SOIL
Wastewater of human origin may also contain residues of pharmaceuticals, espe-

ofwhichhavescatteredsettlementinruralareasorinalpineregions,startedopting
for complementing “decentralized wastewater treatment” pursuing “ecological sani-
tation” (see also: Ecological
sanitationisbasedonsourceseparationoffecesfromurine(e.g.,in“no-mix”toilets
andwaterlessurinals)sothataftersomebasictreatmenttoeliminatefecalpathogens
(often simple storage is sufcient) the solid components can be composted together
with organic garbage and the urine used separately. These low-cost and low-mainte-
na
ncesystemsareespeciallyinterestingfordevelopingcountrieswheremorethan
80%ofsewageisdischargedwithoutanytreatment,wastingthenutrientsnitrogen
(nitrate, ammonia), phosphate, and potassium, which can be used in sustainable agri-
cu
lturelikeorganicfarming,closingnutrientcycleswhileprotectingsurfacewater
and groundwater from eutrophication.
It is clear that human urine may contain residues of pharmaceuticals and that a
sanitation concept including urine recycling will lead to an input of these compounds
intothetopsoillayerofagriculturaleldsorwilllimittheusabilityoftheurine.
The experiments described in this contribution were carried out with urine collected
inaurinetankfromasmallclosed-loopsanitationproject(WaterMillMuseum
“Lambertsmühle,” Burscheid, Germany) where acceptance, efciency, storage, and
fertilization studies have been undertaken.
14
8.2.3 FATE OF PHARMACEUTICALS IN SOIL
Reviews on the fate of veterinary antibiotics,
15
uoroquinolones,
16
and sulfonamides
17
appeared recently. It is also discussed in Chapter 5 of this book. Compounds that

additional factor.
24
Kinetics also have to be taken into account.
25
With some com-
pounds(e.g.,virginiamycin)soilmetabolitesformthathavetobeaccountedfor.
26
© 2008 by Taylor & Francis Group, LLC
Plant Uptake of Pharmaceuticals from Soil 183
Nondegraded residues in the top soil layers can be dislocated by surface run-
offorinterowandpreferentialow.
27
Usuallythegoverningprocesswithpolar
compoundsisleaching.Insandysoilstetracyclinescanleachtolowerlevels,buttoa
higherextentthishappenswithsulfonamides,eventuallyevenintogroundwater.
28,29
Sulfamethoxazole is relatively mobile in soil and has been detected in groundwater
(upto0.47µgL
–1
).
8
For another sulfonamide, sulfachlorpyridazine, sorption coef-
cientsinsoilhavebeendeterminedthatimplypreferentialleachingwiththedrain-
age
water.
30
Thesulfonamidesulfapyridinehasbeenfoundtoadsorbstrongerin
moist soils than in dry ones.
31
Inalysimeterstudyithasbeenshownthatabreak-

–1
.Rootsofcornandsorghumaccumulatedmuch
more active ingredient than the shoots. Similar results were obtained from rye, car-
ro
t,corn,sorghum,andpeaineldtrials.
37
Enrooxacin was also accumulated in
µg g
–1
amounts.
38
Plantuptakeisgovernedbymanyfactors,andithasbeenstudiedwithmany
compounds other than pharmaceuticals because of its potential for phytoremedia-
tion.
39
Even genotype can be important to assess the uptake of contaminants.
40
Roots
suchascarrotsandpotatotuberscantakeupveryhighamountsofpollutants.
41
Mycorrhizationisalsoanimportantfactorashasbeenshownfortheuptakeofatra-
zine.
42
Even desorption-resistant organic compounds can be taken up into the roots,
to some extent, from sediments, which means that compound properties sometimes
do not modulate uptake and that root sorption may be the dominant mechanism
vs.translocationintheplantfornonpolarcompounds.
43
The class of xenobiotics
whose uptake has been intensively studied is pesticides, especially herbicides that

glutathione, a process intensively studied with herbicides.
55,56
Glutathione conjugates
are the transport forms of many xenobiotics; compartmentation is potentially the
nalpurposeofthisprocess.
57
Meanwhile,thispathwayhasalsobeenfoundforthe
detoxication of chlortetracycline in corn
58
;Chapter9ofthisbookisdedicatedto
that mechanism.
8.2.5 ANALYSIS OF PHARMACEUTICAL RESIDUES IN SOIL AND PLANTS
8.2.5.1 Extraction
Sulfonamides are highly water soluble (sulfamethazine: 1500 mg L
–1
, sulfamethoxa-
zole: 610 mg L
–1
), allowing efcient extraction using methanol or methanol/water
mixtures as extraction solvents. On the other hand, it is well known that methanol
coextractsahugevarietyoforganicsoilconstituents,andthisisalsotrueforplants.
liquid chromatography with tandem mass spectrometry (LC-MS/MS) may therefore
behampered,especiallybygivingrisetosuppressionduringionization,whichleads
to underestimations of the concentrations in the extracts.
Methanol has a relatively low bowling point of ca. 64°C, but with the method
of pressurized uid extraction,
59
whichcanbeperformedwithcommerciallyavail-
able accelerated solvent extraction (ASE) equipment, organic solvents can be used
attemperaturesupto200°Cbyholding,forexample,apressureof20MPainhigh

determinationoftetracyclinewithELISAkitsbyAgaandcoworkers
66
and our study
on the determination of sulfamethazine in soil extracts
13
using the antibodies pre-
paredbyFráneketal.
67
8.3 MATERIALS AND METHODS
All chemicals were used at least in analytical grade. Pharmaceutical standards sul-
famethazine (sulfadimidin) and sulfamethoxazole for the construction of calibration
curves were prepared from reference substances obtained from Riedel-de Haën and
were of 99.7% and 99.9% purity, respectively. The expression “ultrapure water” in
thetextreferstowaterdeionizedbyreverseosmosisandsubsequentpassagethrough
aMilli-Q
®
water purication system. Methanol was high performance liquid chro-
matography (HPLC) grade.
8.3.1 SULFONAMIDES AND PLANTS STUDIED
Sulfamethazine (SMZ, in Europe frequently named “sulphadimidine”) and sulfa-
methoxazole (SMX) have been chosen from the sulfonamide antimicrobials because
much data on their properties and behavior exist. The chemical structures of these
compounds are show in Figure 8.2. pK
a
values are 2.4 and 7.4 and 1.8 and 6.0 for
sulfamethazine and sulfamethoxazole, respectively.
61
Sulfamethazine is a frequently
usedveterinaryantibiotic,andinapreviousstudyithasbeendetectedinsoilincon-
ce

NH
2
R
N
O
CH
3
R =
Sulfamethoxazole (SMX)
(hum.)
FIGURE 8.2 Chemical structures of the compounds studied.
© 2008 by Taylor & Francis Group, LLC
186 Fate of Pharmaceuticals in the Environment and in Water Treatment Systems
Crop Science Corporation facilities at Burscheid. All soils had been taken from the
topsoillayeratleast1yearbeforetheexperiments,airdriedandsievedthrougha
sievewith5-mmporewidth,exceptsoilCwhichwasusedfreshly.Table 8.1 pro-
vides soil data.
8.3.3 URINE
Urine was collected from the model project on ecological sanitation described in
Section8.2.2andwasbetween1and3monthsofage;pHwas9.Urinespikingwas
performedatIWWWaterCenter,Mülheim/Ruhr,Germany.Eightotherpharmaceu-
ticals were present in the spiking solution but were not analyzed in this project. Their
effect on the course of the plant experiment and on the analytical accuracy has been
determined to be negligible in separate experiments (data not shown).
8.3.4 SOIL EXTRACTION
Soilsamples(equivalentto10.0gdrymass)weresubjectedtoASEusinganASE
®
200 Extractor (Dionex). The 11-mL extraction cells were used, with the frits of the
cellheadsprotectedagainstcloggingbyaglassberlterandca.1gofdiatoma-
ceous earth (Isolute

byafactorof100andthissamplewasstoredinbrown10-mLglassvialswithplastic
lidandaTeon-coatedrubberlinerintherefrigeratorat4°Cuntilmeasurement.
Inordertodeterminetherecoveriesofsoilextraction,apreliminaryminiatur
-
iz
edincubationexperimentwith10gofsoilandtwolowspikinglevels(8and16
µg kg
–1
equivalent to ca. 1 kg ha
–1
application rate) was used. The soil was put in a
10-mLglassvial,1.6mLofurine(atypical“applicationrate”)wereadded,andthe
openvialsincubatedfor4weeksat20°Cand80%relativeairhumidityinthedark.
Thesoilwaskeptat14%ofitsmaximumwaterholdingcapacity.Eachvariantwas
runinduplicate.
8.3.5 PLANT EXTRACTION
The plant biomass was deep frozen immediately after harvest. The water contents
of the frozen grass were not determined or accounted for. However, in preliminary
experimentsitwasshownthatitdidnotexceeded5%oftheoriginalmass.For
extraction,thedeep-frozenplantswereputinaporcelainmortar,coveredcom
-
pl
etelybyliquidnitrogen,andmanuallygroundwithaporcelainpestletoobtaina
homogeneoussample.Thissamplecouldbeeasilysubdividedintosmallerportions
after evaporation of the nitrogen.
About5gofthehomogenatewereweighedintoashort100-mLglassmeasur
-
ingcylinder.Then50.0mLofpurewaterwereadded,andthemixturewashomog-
en
izedbyarotarymixerathighspeed(Ultra-Turrax

© 2008 by Taylor & Francis Group, LLC
188 Fate of Pharmaceuticals in the Environment and in Water Treatment Systems
nomatrixeffectshavebeenobservedpreviously(datanotshown).Thereweresome
concerns as to whether the urine components in the leachate would interfere and affect
theaccuracyoftheELISAmeasurements.Thereforeanexperimentwascarriedout
on recoveries of spiked urine concentrations, including dilutions of the samples.
8.3.7 LABORATORY STUDY: SORPTION AND MICROBIAL DEGRADATION IN SOIL
In an incubation experiment of the active ingredients with different soils the pro-
cesses of sorption and degradation compete with each other. This study was per-
fo
rmed in a controlled environment in the climate chamber. After preliminary
miniaturized recovery experiments it was concluded that soil moisture could be
decisive, and therefore the four soils were studied at three different moisture levels.
The experiments were performed in “microcosms,” 250-mL Erlenmeyer asks with
anarrowneckthatwereclosedbyalidformedofaluminumfoil.Theaskswere
lledwith100.0goftherespectivesoils.Themaximumwater-holdingcapacitywas
determined with the bulk soils, and 40, 60, and 90% of that value were maintained
readjusting the weight with some drops of ultrapure water. Incorporation of the com
-
pounds into the soil was achieved by spiking puried sea sand with a methanolic
solution of the substances and letting the solvent evaporate to yield a nal mass
concentration of ca. 1000 mg kg
–1
(actual concentrations obtained were 996 mg kg
–1
SMZ and 1003 mg kg
–1
SMX). After homogeneous incorporation of 1.00 g of the
seasandintothe100.0gofsoiltheresultingsoilconcentrationwas10mgkg
–1

–1
%
Cross-Reactivity
%
anti-sulfamethazine 5 0.005 0.1 21 SMX <0.1%
anti-sulfamethoxazole 10 0.01 0.1 17 SMZ <0.1%
© 2008 by Taylor & Francis Group, LLC
Plant Uptake of Pharmaceuticals from Soil 189
8.3.8 GREENHOUSE STUDY: LEACHING VERSUS UPTAKE
In this study the concurrent processes of leaching out of the root zone and plant
uptake from the root zone were examined. Degradation, which also takes place,
could not be accounted for directly in this study. In this experiment urine, spiked
withthetwosulfonamides,wasappliedasafertilizeronItalianryegrass.Theveg-
et
ationexperimentwascarriedoutinagreenhouse.Duetolimitedcapacityonly
threesoils(A,C,andD)couldbeused.Thepotswerelocatedonacarriageon
rails and could be exposed to direct sunlight and wind, while it was also possible
toprotectthemfromheavyrainfall.Incontrasttothelaboratoryexperiments,the
containers could not be kept on a constant moisture level. Water demand is higher in
pot experiments than in the open land because the containers surrounding the soil
areexposedtohigherambientairtemperaturesandsoilirradiationgivesrisetoan
increased evapotranspiration from the soil surface.
The plants were grown in “Mitscherlich” containers. These containers are made
of sheet iron coated with white enamel, which prevents sorptive processes. The con-
ta
iners also possess a lid on the bottom that retains the soil while allowing leachate
todrainintoacollectiondishbelow.Mitscherlichcontainershaveasurfaceof0.03
m
2
andhold10Lofsoil;eachsoilwaslledintothepotstoanalweightof6.00

of SMX, respectively.
In order to produce signicant amounts of leachate, the soils were over-saturated
to about 110% of the maximum water-holding capacity twice a week. Certainly these
drying/saturation cycles would lead to different microbial populations than in closed
container experiments. The drying/saturation cycles are therefore part of a “worst-
case” scenario for leaching. After each of these seven intense rainfall events leachate
© 2008 by Taylor & Francis Group, LLC
190 Fate of Pharmaceuticals in the Environment and in Water Treatment Systems
wasformed(40to960mL,dependentonthesoil),sampledfromthecollectingdish,
weighed, and aliquoted.
Attheendoftheexperimentthewholesoilvolumewassievedthrougha2-mm
sieveandreducedtoa200-glaboratorysample.Theplantswereharvestedattwo
times of the experiment; this means that after a rst cut, a second cut was obtained
from the same plants that grew without a signal of further growth inhibition. Soil
extractionbyASEwasperformedasdescribedabove,theleachatewasdilutedbya
factorof100to10,000,soilextractsbyafactorof1to100(SMZ)and1to10,000
(SMX), respectively.
8.4 RESULTS
8.4.1 ELISA M
EASUREMENTS AND EXTRACTION RECOVERY
Table 8.3showstheresultsoftherecoveryexperimentsofspikedsulfonamidesin
urine stored for 4 weeks measured with the sulfamethazine ELISA. It can be seen that
in direct urine samples an overestimation of the concentrations was observed, giving
recoveryratesofmorethan200%.Itcanalsobeseenthattheselectivityoftheserum
forsulfamethazineinthepresenceofequalconcentrationsofsulfamethoxazoleis
sufcient(“recovery”of2%orless).Whileadilutionofthesamplesbyafactorof10
doesnotsolvetheproblem,adilutionstartingfrom1:100andhighercancircumvent
matrix effects of urine and leads to acceptable recovery rates (mean 75%).
Thesoilextractionexperimentgaveanaveragerecoveryof84%forresidues
agedinasoilfor4weeks.Itwasalsoconcludedthatwiththeseminiaturizedexperi

*
mean of two sets of triplicates from two different runs
© 2008 by Taylor & Francis Group, LLC
Plant Uptake of Pharmaceuticals from Soil 191
theconcentrationsofsulfamethazinewasobserved.SolelyinsoilC,whichisthe
freshlytakensoilthathadnotbeendriedbefore,adeclinewasfoundtoalevelof
20%oftheinitialconcentrationafter3months.InthesandysoilDbothdegrada
-
ti
on and sorption are lowest. Hence, any concentration higher than 50% of the initial
concentration after 3 months can be attributed to high desorption of the residues
from this sandy soil. Even with soil C, it is reasonable to assume that not all 80%
of the compound lost has been degraded; i.e., only about 50% degraded while 30%
isstronglyboundtotheorganicmatterofthesoil
.Thisb ound fraction possesses a
lowerbioavailabilityandwillbemoreresistanttodegradation.
With sulfamethoxazole the situation is similar (Figure 8.4). There is no clear
correlation between the degradation rate and the moisture level. However, the high
-
es
tmoisturelevelappearstoleadtoreduceddegradation.With90%watersaturation
anaerobic conditions can be assumed in the soil, at least temporarily. Under these
circumstances sulfamethoxazole seems to be especially difcult to degrade. In the
fresheldsoilCdegradationdecreasesthelevelstoabout15%aswellasintheallu
-
vi
al soil B. In the sandy soil D concentrations were determined that are within the
error tolerance close to the initial concentration. Sulfamethoxazole does not appear
to degrade in sandy soil D. Only the drier variants show some decay. With the incu
-

Concentration in Soil … mg kg
–1
FIGURE 8.3 Mean residual concentration of sulfamethazine in the four soils (up left: A; up
right: B; down left: C; down right: D); Shading: soil moisture levels 30% (black), 60% (gray),
90%(white);Samplingintervalsrepresent3,6,9,and12weeksafterspiking.Theinitial
concentration was 10.0 mg kg
–1
; Error bars represent one standard deviation.
© 2008 by Taylor & Francis Group, LLC
192 Fate of Pharmaceuticals in the Environment and in Water Treatment Systems
rst harvest could be performed. The grass continued growing at the same growth
rateand2weekslaterasecondcutwasobtained.
8.4.3.2 Leaching
Bysampling1dayafterevery“heavyrain”event,withleachateconcentrationsof
up to 30 mg L
–1
and determination of amount of leachate together with the respec-
tive concentrations, the leached loads of sulfonamides could be calculated. For sul-
fa
methazinethedataarepresentedinFigure 8.5 for t
hethreesoils.Itcanbeseen
thatthemaincarry-outofthecompoundtakesplaceaftertherstraineventwitha
loadof3tomorethan6mg.Milligramamountswerealsoleachedonthefollowing
days,thesandysoilgivingthelargesttailing.Theloamysiltshowsthelowestpeak
concentration.
8.4.3.3
Plant Uptake
Figure 8.6showstheresiduesoffreesulfonamidesfoundinItalianryegrassgrown
inspikedpotexperiments.Thecontrolshadmassconcentrationsbelow0.05mg
kg

4.5
6.0
7.5
9.0
Sampling Time … Weeks after Spiking
3 6 9 12 3 6 9 12
36912 36912
Concentration in Soil … mg kg
–1
FIGURE 8.4 Mean residual concentration of sulfamethoxazole in the four soils. See legend
of Figure 8.3.
© 2008 by Taylor & Francis Group, LLC
Plant Uptake of Pharmaceuticals from Soil 193
8.5 CONCLUSION
Itwasfoundthatthesulfonamidesstudiedinfoursoilswerenotdegradedtohalfof
the initial concentration after 3 months. Furthermore, it is possible that the observed
decreaseinconcentrationswasnotduetometabolismbutratherduetopoorextrac-
tion recovery resulting from the strong binding of the compounds to the soil matrix
(mineralsandorganicmatter).Degradationseemstobehigherunderaerobiccon-
ditionswhenmicrobialactivityishigh.Thiswasobviouswhenfreshsoilresulted
in higher degradation of the sulfonamides while anaerobic conditions (high water
saturation of soils) resulted in lower degradation rates.
FIGURE 8.6 Residual concentrations (measured in triplicate) in Italian ryegrass grown on
three different soils spiked with 10.0 mg kg
–1
of the sulfonamides sulfamethazine and sulfa-
methoxazole. Left: grass blades harvested after initial growth; right: concentrations in grass
grown after the rst cut. Means of three replicates (pots); bars: standard deviations; Soils as
in Table 8.1.
© 2008 by Taylor & Francis Group, LLC

194 Fate of Pharmaceuticals in the Environment and in Water Treatment Systems
Apart from the two sulfonamides for which the concentrations were determined
byELISAandthatwereappliedwithaloadof10mgkg
–1
(dry mass), which repre-
sentsanapplicationrateofca.20kgha
–1
, there were eight other compounds present
inthespikingsolutiontogiveatotalsoilconcentrationof60mgkg
–1
.Thiswasto
present a worst-case scenario.
Leaching is an important pathway for dissipation of sulfonamide residues in
soil. Polar compounds such as sulfamethazine are preferentially translocated, and
leachate concentrations reach the milligram-per-liter range. Most of the sulfon-
am
idesweretransportedwiththerstsimulatedrainfallevents,whichissimilar
to the behavior of polar pesticides that often drain to lower levels and even down to
groundwaterbypreferentialow.
For a rough estimation, the total load in the leachate was calculated by summing
upthedailyloads.Thisgivesanestimateofca.6mgfortheloamysiltsoilA,ca.
10mgforthefreshsoilC,andca.12mgforthesandysoilD,withitsloworganic
carboncontent,respectively.Fromthe60mgappliedtothecontainerthisis10,16,
and20%,respectively.
It was found that the polar sulfonamides can be taken up into forage grass. The
plant concentrations were in the milligram-per-kilogram range as in some refer
-
enceswithhighamountstakenupfromhydroponics.
9,38
On an average 200 to 500 g

tolerantimmunoassayscouldbeavaluabletoolinperformingteststhatmightbe
necessary for the registration of chemicals or for gathering data on chemicals in
the European Registration Evaluation, Authorization, and Restriction of Chemicals
(REACH) process.
© 2008 by Taylor & Francis Group, LLC
Plant Uptake of Pharmaceuticals from Soil 195
ACKNOWLEDGMENTS
ThisworkwascarriedoutattheInstituteofCropScienceandResourceConserva-
tion (INRES), Department of Plant Nutrition of the Faculty of Agriculture of the
UniversityofBonn.IwouldliketothankMrs.MelanieMuchaforvaluablelabora-
to
ry assistance, Mohamed Hashim for help in the lab, and Dr. Thorsten Christian
for developing the anti-sulfamethoxazole antibodies. Supply of anti-sulfamethazine
antibodies by Dr. Milan Fránek, Veterinary Research Institute, Brno, Czech Repub-
li
c,isgratefullyacknowledged.PartofthisworkwassupportedbytheMinistry
ofEnvironment,NatureConservation,AgricultureandConsumerProtectionofthe
Federal State of North Rhine-Westphalia, Germany. The IWW Water Center at Mül-
hei
m/Ruhr,Germany,isthankedforpreparingspikedseasandsamples.Thanksto
Dr.JoachimClemens(INRES)forcoordinatingtheproject.
REFERENCES
1. Langhammer, J.P., Büning-Pfaue, H., Winkelmann, J., and Körner, E., Chemotherapeu-
tical residues and resistance in post-partum sows during herd treatment [in German],
Tierärztl. Umsch.,
4
3, 375, 1988.
2. Hirsch, R., Ternes, T., Haberer, K., and Kratz, K L., Occurrence of antibiotics in the
aquatic environment, Sci. Total Environ.,
2

me
thoxineusedinintensivefarming(Panicum miliaceum, Pisum sativum and Zea
mays)
, Agric. Ecosys. Environ., 5
2, 103, 1995.
10. Richardson, M.L. and Bowron, J.M., The fate of pharmaceutical chemicals in the
aquaticenvironment—areview,J. Pharm. Pharmacol.,
3
7, 1, 1985.
11. Gavalchin, J. and Katz, S.E., The persistence of faecal-borne antibiotics in soil,
J.
AOAC Int., 7
7: 481–485, 1994.
12. Wang, Q Q., Bradford, S.A., Zheng, W., and Yates, S.R., Sulfadimethoxine degrada-
ti
on kinetics in manure as affected by initial concentration, moisture, and temperature,
J. Environ. Qual.,
3
5, 2162, 2006.
13. Christian, T., Schneider, R.J., Färber, H., Skutlarek, D., and Goldbach, H.E., Deter-
mi
nation of antibiotic residues in manure, soil and surface waters, Acta Hydrochim.
Hydrobiol.,
3
1, 36, 2003.
© 2008 by Taylor & Francis Group, LLC
196 Fate of Pharmaceuticals in the Environment and in Water Treatment Systems
14. Otterpohl, R., Options for alternative types of sewerage and treatment systems directed
to improvement of the overall performance, Water Sci. Technol., 45,149,2002.
15. Sarmah,A.K.,Meyer,M.T.,andBoxall,A.B.A.,Aglobalperspectiveontheuse,sales,

27. Addison, J.B., Antibiotics in sediments and run-off waters from feedlots, Res. Rev., 92,
1, 1984.
28. Hamscher, G., Pawelzick, H.T., Höper, H., and Nau, H., Different behavior of tetracy-
clines and sulfonamides in sandy soils after repeated fertilization with liquid manure,
Environ. Toxicol. Chem., 24,861,2005.
29. Hamscher, G., Sczesny, S., Höper, H., and Nau, H., Determination of persistent tet-
racycline residues in soil fertilized with liquid manure by HPLC-ESI-MS-MS, Anal.
Chem., 74, 1509, 2002.
30. Boxall, A.B.A., Blackwell, P., Cavallo, R., Kay, P., and Tolls, P., The sorption and trans-
portofasulphonamideantibioticinsoilsystems,Toxicol. Lett., 131, 19, 2002.
31. Thiele, S., Adsorption of the antibiotic pharmaceutical compound sulfapyridine by a long-
term differently fertilized loess Chernozem, J. Plant Nutr. Soil. Sci., 163, 589, 2000.
32
. Wehrhan, A., Kasteel, R., Simunek, J., Groeneweg, J., and Vereecken, H., Transport
of sulfadiazine in soil columns—Experiments and modelling approaches, J. Contam.
Hydrol., 8
9, 107, 2007.
33. Jjemba, P.K., The potential impact of veterinary and human therapeutic agents in
manure and biosolids on plants grown on arable land: a review, Agric. Ecosyst. Envi-
ron., 93:267,2002.
34. Batchelder, A.R., Chlortetracycline and oxytetracycline effects on plant growth and
developmentinliquidcultures,J. Environ. Qual., 10,515,1981.
© 2008 by Taylor & Francis Group, LLC
Plant Uptake of Pharmaceuticals from Soil 197
35. Batchelder, A.R., Chlortetracycline and oxytetracycline effects on plant growth and
developmentinsoilsystems,J. Environ. Qual., 1
1, 675, 1982.
36. King,L.D.,Saey,L.M.Jr.,andSpears,J.W.,Agreenhousestudyontheresponseofcorn
(Zea mays L.) t
omanurefrombeefcattlefedantibiotics,Agric. Wastes, 8, 185, 1983.

2
006.
44
. Khan, S.U. and Marriage, P.B., Residues of atrazine and its metabolites in an orchard
soil and their uptake by oat plants, J. Agric. Food Chem., 2
5, 1408, 1977.
45. Balke,N.E.andPrice,T.P.,Relationshipoflipophilicitytoinuxandefuxoftriazine
herbicides in oat roots, Pestic. Biochem. Physiol., 3
0, 228, 1988.
46. Colinas, C., Ingham, E., and Molina, R., Population responses of target and nontarget
forest soil organisms to selected biocides, Soil Biol. Biochem., 2
6, 41, 1994.
47. Halling-Sørensen, B., Sengelov, G., and Tjornelund, J., Toxicity of tetracyclines and tet-
ra
cycline degradation products to environmentally relevant bacteria, including selected
tetracycline-resistant bacteria, Arch. Environ. Contam. Toxicol., 4
2, 263, 2002.
48. Chander, Y., Kumar, K., Goyal, S.M., and Gupta, S.C., Antibacterial activity of soil-
bound antibiotics, J. Environ. Qual., 3
4, 1952, 2005.
49. Opalinski, K.W., Dmowska, E., Makulec, G., Mierzejewska, E., Petrov, P., Pier-
zy
nowski,S.G.,andWojewoda,D.,Thereverseofthemedal:feedadditivesinthe
environment, J. Animal Feed Sci.,
7
,35,1998.
50. Migliore, L., Civitareale, C., Cozzolino, S., Casoria, P., Brambilla, G., and Gaudio,
L., Laboratory models to evaluate phytotoxicity of sulphadimethoxine on terrestrial
plants, Chemosphere, 3
7, 2957, 1998.

Environ. Sci. Tech-
nol., 41, 1450, 2007.
5
9. Zhu, Y., Yanagihara, K., Guo, F.M., and Li, Q.X., Pressurized uid extraction for quan
-
t
i
tative recovery of acetanilide and nitrogen heterocyclic herbicides in soil,
J. Agric.
Food Chem., 48, 4097, 2000.
60. Obana,H.,Kikuchi,K.,Okihashi,M.,andHori,S.,Determinationoforganophospho
-
r
u
s pesticides in foods using an accelerated solvent extraction system,
Analyst, 1
22,
2
17, 1997.
61. Stoob, K., Singer, H.P., Stettler, S., Hartmann, N., Mueller, S.R., and Stamm, C.H.,
Exhaustive extraction of sulfonamide antibiotics from aged agricultural soils using
pressurized liquid extraction,
J. Chrom. A, 1
128, 1, 2006.
62. Miao, X S. and Metcalfe, C.D., Determination of carbamazepine and its metabolites
in aqueous samples using liquid chromatography-electrospray tandem mass spectrom
-
et
ry.
Anal. Chem.,

and Buxton, H.T., Pharmaceuticals, hormones, and other organic wastewater contami
-
nantsinU.S.streams,1999–2000:anationalreconnaissance,
Environ. Sci. Technol.,
3
6
, 1202, 2002.
69. Scherer, H.W., Goldbach, H.E., and Clemens, J., Potassium dynamics in the soil and
yield formation in a long-term eld experiment,
Plant Soil Environ.,
4
9, 531, 2003.
70. Schneider, C., Schöler, H.F., and Schneider, R.J., A novel enzyme-linked immunosor
-
b
e
nt assay for ethynylestradiol using a long-chain biotinylated EE2 derivative,
Steroids,
6
9
,245,2004.
© 2008 by Taylor & Francis Group, LLC