Biotic and abiotic degradation studies of organic contaminants course pdf/pdf/Manuela...
Transcript of Biotic and abiotic degradation studies of organic contaminants course pdf/pdf/Manuela...
1
Biotic and abiotic degradation studies
of organic contaminants
Manuela Peschka
Ankara, 8th October 2007
Outline
• Organic pollutants
Selected compounds
• Simulation of abiotic/biotic degradation
Fixed bed bioreactor
Bottle test
Suntest
• Analysis of micopollutants and identification of transformation products
Sample pretreatment
Analytical Instrumentation
• Results:
Case study I: barbiturates
Case study I: photodegradation of bentazone
Case study II: biodegradation of a new fluorosurfactant
Case study IV: biotransformation of clotrimazole
2
pesticides
surfactants
pharmaceuticals
OrganicMicropollutants
HN SO2
N
O
bentazone
F
F
F
OS
O
O
O
10-(trifluoromethoxy) decane-1-sulfonate
C NH
N OO
O
R3
R1
R2
barbiturates
abiotic degradation
biodegradation
monitoring
OrganicMicropollutants
NN
Cl
clotrimazole
Degradation Tests:
3
sampling
10 L storage tank
aerator
fixed bed
pump
Fixed Bed Bioreaktor
5 L storage tankaerator
Bottle Test
4
quartz tubes
xenon arc lampE = h·ν
Suntest – simulation of abiotic photodegradation
Sample Pretreatment:
5
200mbar
sorbent
OAIS® HLB Reversed Phase material with HHydrophilic – LLipophilic BBalanceA = 810 m2/gpore diameter = 80 ÅV = 1,3 cm3/gparticle diameter = 60, 30, 25, 15 und 5 µm
NO
NO
hydrophilicmonomer
lipophilicmonomer
1 atm1 atm
6
SO3-
N
O
SO3-
O NH2
OH
OAIS® MCX
MMixed-Mode CCation eXXchange and Reversed Phase1,0 meq/g SO3H Ion exchange capacity
OAIS® MAX – Phasenchemie
MMixed-Mode AAnion eXXchange and Reversed Phase
COO-
CH3O
S
R+
NO
CH2
N+ CH3H3C
C4H9
7
A B
– typical solvents:
• methanol
– methanol with acid
– methanol with base
• acetonitrile
• acetic acid
• THF
• acetone
• dichlormethan, chloroform
• hexane
increasing elution potential
Elution (for non polar SPE)
Analysis
Analytical Instrumentation:
8
signalconvertion
vacuum system
result
injector ion source detectormassmass analyzeranalyzer
10-5 – 10-8 Torr
General setup of a mass spectrometer
chromatographic separation
GC-MS
9
Advantage: utilization of mass spectra librariesstrong fragmention gives structural information
Disadvantage: single MSEI only in combination with GC
LC-MS
10
Advantage: - analysis of polar, thermolabile compounds
- very robust for routine analysis
Disadvantage: - no spectral libraries
- matrix effects
Quadrupole Time of Flight Mass Spectrometer
UezvmEkin **²** ==21
Uezm **²** =⎟⎠⎞
⎜⎝⎛
21
21
²²
*)**(Lt
Uezm 2=
L = ca. 2m
11
Advantage: - high resolution, e.i. determination of up to the
fourth dezimal (e.g. 255.0432 amu)
- determination of the elemental composition
- MS²
Disadvantage: - not sensitive
QqLIT
Advantage: - structure elucidation of unknown compounds
- high sensitivity
Disadvantage: ?
- user can select various modi, e.g. Q1MI, MRM, product ion scan
- enhanced modi: Q3 acts as trap resulting in higher sensitivity
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Ion Ion chromatographychromatography
CH3 CH3
S SO
O-
O OO
O-
A+
A+
E+
E+
flow direction
stationary phase
-determination of inorganic paramter, e.g. SO42-, Cl-, F-
- mineralization of a molecule
GC-MSLC-MS
QqTOF
QqLIT
monitoringmonitoring
structurestructureelucidationelucidation
(IC)
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Case studies:
2,08
0,21 0,
49
0,28
0,09 0,12 0,30 0,39
0,30 0,40 0,51
0,98
2,48
1,41
0,86
4,55
0,31
0,29
0,28
0,19
0,22 0,27
0,88
0,10
0,10
1,5
0,3
0,6
5,45,3
0
1
2
3
4
5
2004
-04-
02
2004
-04-
12
2004
-04-
30
200
4-05
-30
2004
-09-
30
2005
-02-
02
2005
-02
mix
2005
-03-
28
2005
-04-
08
2005
-04-
09
2005
-04-
10
2005
-04-
11
2005
-04-
12
2005
-05-
07
2005
-07-
01
2005
-07-
02
2005
-07-
03
2005
-11-
13
conc
. [µg
/L] pentobarbital
butalbitalsecobarbitalphenobarbital
time [h]0 2 13 21
conc
entra
tion
[ng/
L]
0
500
1000
1500
2000 aprobarbitalpentobarbitalbutalbitalsecobarbitalhexobarbitalphenobarbital
Fixed Bed BioreactorExposion to sunlightHydrolysis (pH 2 – 12)
groundwater in Berlin
Barbiturates
C NH
N OO
O
R3
R1
R2
14
Bentazone
HN SO2
N
O
• one of the most applied herbicides in the Ebro river delta • 31 µg/L in application time from May to August (mean value) (Ebro 0.27 µg/L)• comparable high concentrations river waters of the Tiber region in Italy
• not biodegradeable in a fixed bed bioreactor• passes through membrane bioractor• present in ground- and surface water
quartz tubes
xenon arc lampE = h·ν
0
500
1000
1500
2000
2500
3000
3500
0 2 4 7 9 11 13 14 16 18 20 23
Time [h]
Are
a (b
enta
zone
)
0
20
40
60
80
100
120
140
Are
a (S
O3-
bent
azon
e)
bentazoneSO3-bentazone
0
500
1000
1500
2000
2500
3000
3500
4000
4500
0 2 4 7 9 11 13 14 16 18 20 23
Time [h]
Are
a (b
enta
zone
)
0
50
100
150
200
250
Are
a (S
O3-
bent
azon
e)
bentazoneSO3-bentazone
0
500
1000
1500
2000
2500
3000
3500
4000
4500
5000
0 2 4 7 9 11 13 14 16 18 20 23
Time [h]
Are
a (b
enta
zone
)
0
20
40
60
80
100
120
140
160
180
200
Are
a (S
O3-
bent
azon
e)
bentazoneSO3-bentazone
B - moderately hard water
A – distilled water
C – moderately hard water plus 5 µg/mL humic acids
bentazone
0
500
1000
1500
2000
2500
3000
3500
4000
4500
0 2 4 7 9 11 13 14 16 18 20 23
Time [h]
Are
a (b
enta
zone
)
bentazone
0
500
1000
1500
2000
2500
3000
3500
4000
4500
5000
0 2 4 7 9 11 13 14 16 18 20 23
Time [h]
Are
a (b
enta
zone
)
bentazone
0
500
1000
1500
2000
2500
3000
3500
0 2 4 7 9 11 13 14 16 18 20 23
Time [h]
Are
a (b
enta
zone
)
bentazone
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observed mass(m/z)
elementalcompositionactual mass
∆ mDa
255.0432 ±0.0013 C10H11N2O4S
m/z = 255.0440 - 0.8
197.0044C7H5N2O3S
m/z = 197.0021 2.3
191.0826
C10H11N2O2m/z = 191.0821 0.5
132.0320C7H4N2O
m/z = 132.0324 - 0.4
HN SO2
N
O
OH
NH
HN
O
SO
OHO
observed mass (m/z)
elemental composition actual mass
∆ mDa
257.0584 ± 0.0034
C10H13N2O4S m/z = 257.0596 - 1.2
177.1024 ± 0.0003
C10H13N2O m/z = 177.1028 - 0.4
92.0497 C6H6N m/z = 92.0500 - 0.3
present in drainage channels in the Ebro delta
HN SO2
N
O
0
5
10
15
20
25
30
35
40
1999 2000 2001 2002 2003 2004 2005
An
zah
l d
er
Pu
blikati
on
en
(P
rim
ärl
itera
tur)
Fluorosurfactants
F
F
F
OS
O
O
O
properties:
- persistent to hydrolysis, oxidation, reduction and biodegradation- accumulate in liver, kidneys, mussel tissue and bind to blood proteins
blood
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10-(trifluormethoxy)decane-1-sulfonate
β = 100 mg/LV = 5 L surface waterpH 7T = room temperature (20°C)
Biodegradation in a Biodegradation in a FixedFixed BedBed BioreaktorBioreaktor
AnalyticalAnalytical methodologiesmethodologies
Ion Ion ChromatographyChromatographyF-, SO4
2-, TOCQQ--ToFToF
high resolutionmass spectrometry
QQ--TrapTrapMS³-experiments
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massmass spectrumspectrum of 10of 10--(trifluormethoxy)(trifluormethoxy)--
decanedecane--11--sulfonatesulfonate
F
F
F
O
S
O
O
O
m/z = 85
m/z = 80
F3CO
(CH2)10
SO3
[M-H]-
m/z = 305
SO42-
OSO3
F3C
F3C O O
F3C O COO
F3C O COOH
F3C O COOH
oxidation
beta-oxidation
- CO2
-CO2
F3C OH
- HF
CO2 F-
O
F F
H
H
SO42-
oxygenase
SO3-
(CH2)x(CH2)x
OF3C
O
- F-
HO SO3
HOOC SO3
HOOC SO3
- CO2
SO42-
beta-oxidation-CO2
18
MonitoringMonitoring of TOC, of TOC, FF-- and and SOSO4422--
0
5
10
15
20
25
30
35
40
0 10 20 30 40Time [d]
c (F
- , S
O4
2- ,
TO
C)
[mg
/L]
0,00
0,20
0,40
0,60
0,80
1,00
1,20
c/co
(Tri
flu
orm
eth
oxyd
eca
nsu
lfo
nat)
TOC F- SO42- primary degradation
metabolites (m/z = 85)
OSO3
F3C
after 28 days:
- primary degradation: 100%- > 90 % SO4
2- - concentrationtheoret.- 58 % F- - concentrationtheoret.
F3C O O
F3C O COO
F3C O COOH
F3C O COOH
oxidation
beta-oxidation
- CO2
-CO2
F3C OH
- HF
CO2, F-
O
F F
H
H
SO42-
oxygenase
SO3-
(CH2)x(CH2)x
OF3C
O
?
19
0,E+00
1,E+04
2,E+04
3,E+04
4,E+04
5,E+04
6,E+04
0 5 10 15 20 25 30
Time [d]
are
a
SO3-
(CH2)x(CH2)x
O
F3C
O
0
5
10
15
20
25
30
35
40
0 10 20 30 40Time [d]
c (F
- , S
O4
2- ,
TO
C)
[mg
/L]
0,00
0,20
0,40
0,60
0,80
1,00
1,20
F- SO42-
20
[M-H]-
* *
*
* * **
C11H18F3SO5-* 319.0833
SO3-
(CH2)x(CH2)x
O
F3C
O
F
F
F
OS
O
O
OO
-CO
SO3
oxidation in omega-position!
MSMS²² MSMS³³
85
233
205
233
CO
SO3
[M-H]-
319233
21
OSO3
F3C
oxygenase
SO3-
(CH2)x(CH2)x
OF3C
O
- F-
HO SO3
HOOC SO3
HOOC SO3
- CO2
SO42-
beta-oxidation-CO2
C6H11SO5-195.0333
SO3OOC
SO3HOOC
97.0058
195.0341
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F3C O O
F3C O COO
F3C O COOH
F3C O COOH
oxidation
beta-oxidation
- CO2
-CO2
F3C OH
- HF
CO2, F-
O
F F
H
H
SO42-
OSO3
F3C
> 90%fast
oxygenase
SO3-
(CH2)x(CH2)x
OF3C
O
<10%slow
- F-
HO SO3
HOOC SO3
HOOC SO3
- CO2
SO42-
beta-Oxidation-CO2
494bp [°C]
141 – 145mp [°C]
3.31*10-7Vp [Pa]
0.49Swater [mg/L]
4.1logKOW
344.84MW [g/mol]
NN
Cl
Mode of action:Inhibitor of ergosterolsynthesis resulting in the damage of cell membrane.
23
approx. 2 ng/L
False positive signal in next four measured samples(solvent - acetone)!
0
1
2
3
4
5
6
0,00 2,00 4,00 6,00 8,00 10,0 12,0 14,0pH
log
KO
W
N
N
ClNH
N
Cl
Log KLog KOWOW versusversus pHpH
24
• Acidification to pH 2pH 2 (stirring 30 min) of samples before filtration• SPE-Material Oasis HLB 3cc (60 mg), Waters• conditioning: 1 x 2 mL n-hexan, 3 x 2 mL methanol
5 x 2 mL groundwater (pH 2pH 2) • SPE: 100 ng Atrazin D5 (internal standard)
flow approx. 20 mL/min• drying: 45 min (N2)• elution: 4 x 1 mL acetone• evaporation: to 150 µL• external standard: 100 ng Fluazifop-buthyl (ESTD) and fill up to
200µL final volume with acetone
adapted SPE-Method
Adsorption hinders standardized biodegradation tests
FBBR
0
0,2
0,4
0,6
0,8
1
1,2
0 2 4 6 8 10 12 14t [d]
c/c0
two hours after FBBR passage
25
OHphase I
CRz.B. glucoronic acid
SH
phase II
reaction with „electrophiles“
reaction with „nucleophiles“
5 L storage tankaerator
1 1 5 11 50 60 68 68 68 98
0,00E+00
5,00E+05
1,00E+06
1,50E+06
2,00E+06
2,50E+06
3,00E+06
3,50E+06
Are
a
Time [d]
CLT-metabolite clotrimazole
26
478.1478.1
EPI
- Cl
Cl
Cl
NH3
O
HO
NH3
O
O
NN
Cl
HONH3
O
O
86.1
118.1
165.1
199.1
242.2
277.1
478.1
27
SummarySummary::
bentazone
newfluorosurfactant
barbituratesclotrimazole
FateFate& &
DegradationDegradation
no biotic nor abioticdegradation⇒ persist in theaquatic enironmentfor several years
no bioticdegradation butundergoesphotolysis⇒ degradable in sunlight exessablecompartments
undergoes biodegradation⇒ potential substitute for the harmfulfluorosurfactants presently used
biotransformed⇒ furtherinvestigations
AcknowledgementAcknowledgement::
• AQUATERRA (Project number 505428 GOCE)
• Spanish Ministry of Education and Science
(Project number CTM2005-25168-E)
• Merck KGaA
ThankThank youyou forfor youryour attentionattention!!