PPCP QTRAP 5500 TripleTOF 5600 - Pittcon Web Archives

Screening and Quantitation of Targeted and
Non-targeted Environmental Pollutants in
Water Samples
André Schreiber, Rolf Kern, Chris Borton, Nadia Pace
AB SCIEX, Concord, Ontario (Canada)

Outline
− Different LC-MS/MS screening strategies for PPCP
– Pharmaceuticals
– Personal care products
– Pesticides
– Drugs of abuse
– …
− Experimental conditions
− Screening for PPCP in water samples
– Comprehensive screening using multi-MRM methods
– Direct injection of water with LOD < 10ppt
– Compound identification using MRM ratios and MS/MS spectra
– General unknown screening
2 © 2011 AB SCIEX

Quantitation and Screening Strategies
− Multi-Target Quantitation
– AB SCIEX TripleQuad and QTRAP ® systems
– 2 Multiple Reaction Monitoring transitions per compound
– MRM ratio for identification [1]
– Scheduled MRM for extended screening [3-4]
– Information Dependent Acquisition (IDA) MRM survey to trigger
QTRAP ® MS/MS with library searching [2]
− General Unknown Screening
– AB SCIEX TripleTOF system
– IDA: high resolution accurate mass MS and MS/MS
– Statistical data analysis for unknown identification [5-6]
[1] Christopher Borton et al.: at ASMS conference (2006) Seattle, WA
[2] Jim Krol et al.: at AOAC annual meeting (2007) Anaheim, CA
[3] André Schreiber et al.: at WCTOW (2009) Winnipeg, MA
[4] Rolf Kern et al.: at Pittcon (2009) Chicago, IL
[6] André Schreiber et al.: at ASMS (2010) Salt Lake City, UT
[5] André Schreiber et al.: at WQTC (2010) Savannah, GA
3 © 2011 AB SCIEX

Experimental
− Sample preparation
– Direct injection of drinking and surface water samples
− UHPLC
– Shimadzu UFLC XR system
– Phenomenex LUNA 2.5u C18(2)-HST 100x3mm and Kinetex C18
2.6u 100x2.1mm column
– Fast gradient of water/acetonitrile +0.1% formic acid
− MS/MS
– Turbo V source with Electrospray Ionization (ESI) probe
– QTRAP ® 5500 LC/MS/MS system
– MRM with Scheduled MRM algorithm for quantitation and
MS/MS for compound identification based on library searching
– TripleTOF 5600 LC/MS/MS system
– TOF-MS (100ms) and TOF-MS/MS (10x 50ms) for unknown ID
4 © 2011 AB SCIEX

Information Dependent Acquisition (IDA)
Scheduled MRM survey
intensity threshold
Threshold
EPI
297
159
69 173
81 141
109
115
176
201
255
Library Search
5 © 2011 AB SCIEX

LC-MS/MS Screening –Direct Injection of Water
0.1μg/L
− 158 Scheduled MRM transitions to screen for PPCP
− Compound selection based on EPA 1694
− Injection of 100μL into QTRAP ® 5500 LC/MS/MS system
→ No time consuming extraction
→ Single LC setup for increased sample throughput
6 © 2011 AB SCIEX

Calibration Line (Atrazine 10 ng/L – 10 µg/L)
10ppt #1 10ppt #2 10ppt #3
Calibration for Atrazine 1: y = 4.86373e5 x + 721.61304 (r = 0.99798) (weighting: 1 / x^2)
4.5e6
4.0e6
3.5e6
3.0e6
2.5e6
2.0e6
1.5e6
1.0e6
5.0e5
0.0e0
0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5 9.0 9.5
Concentration (ug/L)
7 © 2011 AB SCIEX

Direct Injection of Farmland Drain Sample
LOD of PPCP

Direct Injection of Sample from Urban Area
200 ng/L Cotinine
LOD of PPCP

MRM Ratio Identification (10 ng/L Metolachlor)
10ppt #1 10ppt #2 10ppt #3
± 20%
10 © 2011 AB SCIEX

Trapping and Scanning in a QTRAP ® (Q3)
(1) Trapping
Exit lens
(2) Scanning
Entrance lens
11 © 2011 AB SCIEX

LC/MS/MS Screening –Farmland Drain Sample
Scheduled MRM – EPI
Sulfadimethoxazine
FIT 83.7%
Atrazine
FIT 98.3%
Propazine
FIT = 98.8%
Metolachlor
FIT=91.8%
12 © 2011 AB SCIEX

Benzoylecgonine – a Marker for Cocaine
Consumption
H 3
O
C
O
OH
O
N CH 3
Metabolism
O
N CH 3
O
O
O
Cocaine
Benzoylecgonine (BE)
13 © 2011 AB SCIEX

Quantitation of Benzoylecgonine in 70 Water
Samples from Different Geographies and Types
0.20
Berlin
0.18
0.15
BE Concentration (ppt)
0.13
0.10
0.08
0.05
0.03
Standard 0.1µg/L
Zermatt
Creeks
Drinking water
Canoe Lake (Algonquin)
Lake Zürich
Lakes
Dresden
Philadelphia
Santa Fe
Rio
Rom
Vienna
Toronto
Winnipeg
0.00
Sample
Rivers
14 © 2011 AB SCIEX

Atrazine – a Persistent Organic Pollutant (POP)
Cl
N
N
H 3
CH 3
http://toxipedia.org/display/toxipedia/Atrazine+-+History+and+Uses
C
NH N NH
Atrazine
CH 3
15 © 2011 AB SCIEX

Quantitation of Atrazine in 70 Water Samples
from Different Geographies and Types
0.140
(Highest # of samples)
(Only 2 samples)
0.120
Standard 0.1µg/L
(Only 2 samples)
0.100
Atrazine Concentration (ppt)
0.080
0.060
0.040
Creeks
Drinking water
(Atrazine is banned in EU.)
Lakes
0.020
0.000
16 © 2011 AB SCIEX
Sample
Rivers

Caffeine – A Marker for Human Activity (Coffee
Consumption and Pain Medication)
O
CH 3
H 3 C
N
N
CH 3
O
N
N
Caffeine
17 © 2011 AB SCIEX

Quantitation of Caffeine in 70 Water Samples
from Different Geographies and Types
40.0
35.0
Holy water from a church in Roma
30.0
Philadelphia
Caffeine Concentration (ppt)
25.0
20.0
Mountain hut Switzerland
Zürich
Berlin
Rio
15.0
10.0
5.0
Creeks
Vienna
Lakes
Roma
Vienna
0.0
Drinking water
18 © 2011 AB SCIEX
Sample
Rivers

General Unknown Screening
–The Challenge of Finding the Unexpected –
19 © 2011 AB SCIEX

General Unknown Screening
–The Challenge of Finding the Unexpected –
BPC (Lake Zürich)
20 © 2011 AB SCIEX

Comparison of 70 Water Samples
1. Unknown Screening (PCA)
Samples from wilderness areas
with extensive recreational use
Drinking water produced from a
river bank filtrate during flood
Samples from wilderness areas
with extensive recreational use
Increasing urbanity of samples
Drinking water produced from a
river bank filtrate during flood
Increasing urbanity of samples
21 © 2011 AB SCIEX

Comparison of 70 Water Samples
1. Unknown Screening (PCA)
Samples from wilderness areas
with extensive recreational use
Drinking water produced from a
river bank filtrate during flood
?
?
Increasing urbanity of samples
22 © 2011 AB SCIEX

Empirical Formula Calculation / Library Search
O
H 3 C
C +
H 3 C
DEET
O
N
H 3 C
C + NH
CH 3
CH 3
C 12 H 17 NO+H +
192amu
OH
Unknown spectrum
N CH 2
C 8 H 7 O +
119amu
C 8 H 8 N +
118amu
CH
1 st hit
2
– 96% (DEET)
C 5 H 9 NO+H +
100amu
CH + CH +
C 7 H 7
+
91amu
+
C 6 H 5
+
65amu
C 5 H 5
+
65amu
O
N CH 3
C 3 H 5 NO+H +
72amu
2 nd hit – 25%
23 © 2011 AB SCIEX

Comparison of 70 Water Samples
1. Unknown Screening (PCA)
2. Identification
Samples from wilderness areas
with extensive recreational use
DEET
Drinking water produced from a
river bank filtrate during flood
Increasing urbanity of samples
3. Quantitation
2.6µg/L
2.4µg/L
3.3µg/L
2.5µg/L
3.6µg/L
24 © 2011 AB SCIEX

Summary
− LC/MS/MS is a powerful tool for the analysis of
Pharmaceuticals and Personal Care Products in
environmental samples
– The combination of high resolution LC separation and high
sensitivity MS/MS is the most powerful tool to screen and quantify
targeted compounds.
– Direct injection of water samples
– MS/MS fragmentation required to identify compounds (MRM ratio
or full scan MS/MS with library searching)
– High resolution and accurate mass MS and MS/MS to identify
unexpected compounds
– Statistical data analysis to find unexpected compounds
25 © 2011 AB SCIEX

Trademarks/Licensing
− The authors like to thank colleagues of AB SCIEX for collecting water
samples all over the world during their vacation or while attending
conferences.
− For Research Use Only. Not for use in diagnostic procedures.
− The trademarks mentioned herein are the property of AB Sciex Pte. Ltd. or
their respective owners. AB SCIEX is being used under license.
− © 2011 AB SCIEX.
26 © 2011 AB SCIEX

Thank you for Listening !
27 © 2011 AB SCIEX