Environmental analysis can be extremely challenging due to the low detection levels for toxic contaminants specified by legislation, particularly in drinking water, and the complexity of matrices encountered. Consequently highly selective and sensitive detection methods are required. This presentation provides an introduction to tandem quadrupole mass spectrometry and describes the use of high sensitivity tandem quadrupole mass spectrometry for the analysis of various environmental contaminants including pesticides, endocrine disruptors and polyfluorinated compounds such as PFOS.

1. ©2015 Waters Corporation 1
Analysis of Environmental Contaminants using High
Performance Quantitative LC-MS/MS

2. ©2015 Waters Corporation 2
Overview of the presentation
 Background environmental contaminants
– EU legislation overview
– POPS (dioxins; PCBs; PFCs; pesticides), endocrine disruptors
 Environmental LC-MS/MS analysis – the challenges?
 Why use mass spectrometry?
– Waters MS tandem quadrupole solutions
 Multi-residue pesticide analysis in drinking water
 Direct quantification of acidic herbicides in water
 Strategies for the analysis of Eds & POPs
 Perfluorinated compounds (PFCs)
 Summary

3. ©2015 Waters Corporation 3
Transformation
products
Drinking water
(targeted
analysis)
Perfluorinated
compounds
(PFCs)
Environmental applications
Marine
studies
Environmental
water
Unknown Analysis
Dioxins
POPs
Endocrine
disrupting
compounds

4. ©2015 Waters Corporation 4
Legislation – contamination &
environmental factors
 Regulations on the use of specific chemical substances, e.g. those
used in farming or in certain production or food processing techniques
– Measures are taken to limit pollution of water or air, or from exposure to
radioactivity
EU
– Drinking Water Directive (98/83/EC)
– EU Marine Strategy Framework Directive (2008)
– EU Water framework Directive (2010)
– Community strategy for endocrine disruptors SEC (2011) 1001
 EU “criteria based” approach adopted
– Water Framework Directive methods must be certified according to EN
ISO/IEC-17025
USA
– Drinking Water Regulation
– WHO Guidelines Drinking water quality (2011, 4th edition)
 USA “EPA approved methods of analysis”
http://water.epa.gov/scitech/drinkingwater/labcert/analyticalmethods.cfm#approved

5. ©2015 Waters Corporation 5
Inhalation
Ingestion
POPs
How do pollutants get into the
environment?

6. ©2015 Waters Corporation 6
Why are Persistent Organic Pollutants
(POPs) a problem?
 Persistence
These compounds do not easily break down into simpler, less harmful forms. They remain and
continue to build up in our environment.
 Toxicity
Dioxins have been proven to be highly toxic in both animal and human
studies e.g. chloracne, cancer, learning difficulties, cognitive impairment,
 Health Impacts
High exposures to dioxins have been directly related to cancer. Low level, prolonged exposure
has been linked to a host of effects including hormonal and immune system dysfunctions, birth
defects, miscarriages & learning disabilities.
 Bioaccumulative
Dioxins are stored in fat tissue and continue to accumulate throughout
our lifetime. The body has difficulty metabolizing dioxins, especially
laterally substituted congeners.

7. ©2015 Waters Corporation 7
Environmental LC-MS/MS – challenges?
Can I detect all of the
target compounds and
what about unknowns?
Do my results comply with
the regulatory
requirements?
Detection
capability
Confirmation
Productivity Ease of use
Accessibility
Can my staff use
the technology?
Cal 10 inj1 before 10x dilution
Time
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00 5.50 6.00 6.50 7.00 7.50 8.00
%
0
100
Sample throughput
Can I analyse enough
compounds/samples?

8. ©2015 Waters Corporation 8
Why use Mass Spectrometry (MS)?
 Mass spectrometers offer a number of advantages;
– Measure specific analytes in the presence of thousands of others,
simultaneously (multi-residue analysis)
– Measure trace quantities of analytes (femtomolar concentrations)
– Provide structural information about an analyte
– And can do all of these things at the same time
 “The 3 S’s”
– Selectivity; differentiating between the components in a sample
– Specificity; measuring only analytes in the presence of many other
compounds
– Sensitivity; detection of femtomolar concentrations

9. ©2015 Waters Corporation 9
Tandem quadrupole - instrument
anatomy

10. ©2015 Waters Corporation 10
Tandem quadrupole MS
Versatility and power
 Tandem quadrupole instruments offer many modes of operation
 Each of the quadrupoles can operate in Scanning or Static Modes
– Selected Ion Recording (SIR)
– Precursor Ion Scanning (PAR)
– Constant Neutral Loss (CNL)
– Product Ion Scanning (DAU)
– Multiple Reaction Monitoring (MRM) or Selective Reaction Monitoring
(SRM)
 4-5 orders linear dynamic response
 Switch between positive and negative ion modes in the same
chromatographic experiment
 Meet the legislative requirements for analyte confirmation
 Easy to use & robust measurements

11. ©2015 Waters Corporation 11
Single Ion Recording (SIR)
MS1
Static (m/z 821.5)
Precursor(s)
Q1 Static
m/z 821
Collision
Cell (OFF)
Static (m/z 821.5)
Quadrupole 2
Precursor(s)Precursor(s)

12. ©2015 Waters Corporation 12
SIR

13. ©2015 Waters Corporation 13
Multiple Reaction
Monitoring (MRM),
m/z 821 >768
Analyte Injected in Matrix
Hypothetical Example
0.50 1.00 1.50
Time0
100
%
0
100
%
0.50 1.00 1.50
Time0
100
%
0
100
%
3ng / L 30ng / L
Selected Ion
Recording (SIR),
m/z 821

14. ©2015 Waters Corporation 14
Product Ion Spectrum
790 795 800 805 810 815 820 825 830 835 840 845 850
m/z0
100
%
821.5
810.5
822.5
826.5
827.5
200 250 300 350 400 450 500 550 600 650 700 750 800 850 900
m/z0
100
%
768
576
558548
718
750
786
821
Q1 Static
m/z 821
Q2
Scanning

15. ©2015 Waters Corporation 15
Multiple Reaction Monitoring (MRM)
Quadrupole 1
Collision
Cell
Static (m/z 821.5) Static (m/z 768.5)
Ar (2.5 – 3.0e-3mBar)
Precursor(s)
Product(s)
Quadrupole 1

16. ©2015 Waters Corporation 16
MRM

17. ©2015 Waters Corporation 17
Multiple Reaction
Monitoring (MRM),
m/z 821 >768
Analyte Injected in Matrix
Hypothetical Example
0.50 1.00 1.50
Time0
100
%
0
100
%
0.50 1.00 1.50
Time0
100
%
0
100
%
3ng / L 30ng / L
Selected Ion
Recording (SIR),
m/z 821

18. ©2015 Waters Corporation 18
Solutions for Targeted Screening
Analysis
Xevo TQD
• Entry level
• UPLC compatibility
• Multi-residue methods
Xevo TQ-S micro
• Increased sensitivity
• Increased speed
• PICs
• RADAR
• Universal source
Xevo TQ-S
• Ultimate sensitivity

19. ©2015 Waters Corporation 19
The Universal Source – configurations
and options

20. ©2015 Waters Corporation 20
MS Adoption:
UPLC-Xevo TQD
Intermediate:
UPLC- Xevo TQ-S micro
Max efficiency:
UPLC-Xevo TQ-S MS
Less Sample Prep More
Solutions for Targeted Screening
The need for sample preparation?

21. ©2015 Waters Corporation 21
Ultra-trace analysis
Waters MS solutions
Ultra trace analysis
How can Xevo TQ-S help?
Xevo TQ-S
performance
enables:
Reduced
volumes
during
sample prep
and analysis
Proactively
monitor matrix
levels during
analysis
Improved
reproducibility
due to
increased
signal
intensity
Better quality
results,
minimal user
intervention

22. ©2015 Waters Corporation 22
• Automatically sets up instrument,
• Optimises MRMs and performs system suitability
IntelliStart
• Methods generated automatically by QuanPedia,
• Clear flagging when data is out of regulatory tolerance
• Reduced risk of errors
TargetLynx
• Long term system monitoring, Lab management tool
TrendPlot
• Generates optimised methods, time scheduled MRMs
Quanpedia
• Real time QC, Intelligent ‘on the fly’ decision making
QCMonitor
Waters quantitative workflow and
software solutions

23. ©2015 Waters Corporation 23
Multi-residue pesticide analysis in drinking
water
in accordance with 98/83/EC

24. ©2015 Waters Corporation 24
Prepare sample
• Dechlorinate
• Transfer to sample
vial
Analyse 100 µL
aliquot
• UPLC separation
• Xevo TQ-S
detection
Review results
• Achieve required
limits
• Reproducible ppq
detection
Direct injection of drinking water
Within the EU Drinking Water Directive (DWD) sets quality standards for drinking water
quality at the tap (microbiological, chemical and organoleptic parameters
• 0.1μg/L (100 ppt) for each individual pesticide in drinking water
• 0.5μg/L (500 ppt) total Pesticides (the sum of all the substances detected in a sample)
• stricter separate health based standards for 4 organochlorine pesticides which are no longer
permitted to be used
Drinking water analysis

25. ©2015 Waters Corporation 25
Time
4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 7.00
%
0
100
4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 7.00
%
0
100
4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 7.00
%
0
100
4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 7.00
%
0
100
How does StepWave™ help?
Time
4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 7.00
%
0
100
4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 7.00
%
0
100
4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 7.00
%
0
100
4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 7.00
%
0
100
Enhanced sensitivity
with StepWaveTM
ion
optics
Linuron
Azinphos-
methyl
Atrazine
Metosulam
Without
StepWaveTM ion
optics
100µL drinking water, 81 compounds, 10 x below EU limits (10 ppt)

26. ©2015 Waters Corporation 26
Ultra trace analysis
Direct injection of drinking water
200 pg/L (200ppq) pesticides in drinking water
1.8 ng/L (1.8 ppt) total pesticides
Acetamiprid
223>126
223>56
Omethoate 214>125
214>183
Thiabendizole
202>175
202>131
Simetryn
214>124
214>96
Hexazinone
253>171
253>71
Terbuthiuron
229>172
229>116
Metalaxyl 280>220
280>192
Pirimiphos -
methyl
306>164
306>108
Chlorpyriphos 350>198
350>97
Detection capability

27. ©2015 Waters Corporation 27
Measurement repeatability
Pesticide classes:
Summary of
repeatability @ 100ppt
Ametryn 1.39
Terbutryn 1.96
Cyanazine 1.26
Atrazine 1.46
Simetryn 1.78
Spiroxamine 1.85
Kresoxim Methyl 4.29
Azoxystrobin 2.19
Dimethomorph 4.14
Pyraclostrobin 4.18
Chlortoluron 0.59
Siduron 1.24
Monuron 1.56
Monolinuron 1.09
Diuron 1.24
Dicrotophos 0.94
Heptenophos 1.47
Mevinphos 2.34
Tetrachlorvinphos 2.47
Chlorfenvinphos 3.67
Omethoate 1.22
Demeton S Methyl 1.50
Azinphos Methyl 2.48
Dimethoate 2.27
Ethoprophos 2.31
2.04Mean
Class Compound %RSD (n=32)
Triazine herbicides
Organothiophosphorous pesticides
Fungicides
Phenylurea herbicides
Organophosphorous pesticides

28. ©2015 Waters Corporation 28
TrendPlot
TrendPlot report – reproducibility @100 ppt (EU limit)
%RSD 0.83Fenuron
%RSD 0.78Simazine
%RSD 0.71Flumeturon

29. ©2015 Waters Corporation 29
River water
•A relatively simple matrix?
Total Organic Carbon content (TOC) can commonly
be between 0.5 to 10 ppm
•If 30 compounds are present at 100 ppt (EU limit) this is 3 ppb
of detected compounds
If we have 5 ppm TOC, and detect 3 ppb of
targeted compounds, what else is in the sample?
•Food is a much more complex sample
MRM is targeted, we see only what we tell the
instrument to show us
•How can we tell what is also present?
Monitoring the matrix effects in
river water

30. ©2015 Waters Corporation 30
 Rapid electronics allow instrument to
switch between MRM and full scan (FS
takes 100ms)
– No loss of MRM data quality
– Added information can be gained from
full scan
o Monitor matrix
o Assess for possibility of matrix effects
o Added information for sample prep
method development
o Search for significant non-targeted
compounds
RADAR (Xevo TQ-S & Xevo TQ-S micro)
Simultaneous full scan & MRM
MRM
Full
scan81 compound multi-residue
method, using RADAR

31. ©2015 Waters Corporation 31
100 ng/L (100 ppt) pesticides in river water
RADAR view – what else is in the
sample?

32. ©2015 Waters Corporation 32
PCP XIC (m/z 264.8)
TIC
m/z 264.8 - PCP
RADAR - TIC
Sewage Effluent spiked with
20 ng/L pentachlorophenol (PCP)

33. ©2015 Waters Corporation 33
Strategies for the analysis of endocrine
disruptors &
Persistent organic Pollutants (POPs)

34. ©2015 Waters Corporation 34
Endrocrine disrupting compounds
Natural hormones & synthetic compounds
Time
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
3.13
3.19
3.10
3.09
2.97
2.46
17a-ethinylestradiol
17b- and 17a-estradiols
Estrone
Bisphenol A
Estriol
Endocrine
Disruptors
Challenging LODs
8 minute analysis
17b- and 17-a-estradiols
Estrone
Bisphenol A
Estriol

35. ©2015 Waters Corporation 35
Endrocrine disrupting compounds
Endocrine
Disruptors
10ng/ml matrix matched
standard
Time
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
1.60 1.80 2.00 2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00
%
0
100
3.06
3.133.03
3.01
2.81
2.35
17a-ethinylestradiol
17b- and 17a-estradiols
Estrone
Bisphenol A
Estriol
Compounds
Method
LOD ng/L
200 mL
sample
Estriol 4
Bisphenol A 2
Estrone 0.5
17-estradiol 1
17-ethinyl
estradiol 1.5
17-estradiol 1
17b- and 17-a-estradiols
Estrone
Bisphenol A
Estriol

36. ©2015 Waters Corporation 36
Perfluorinated compounds (PFCs)
perfluoroalkyl sulfonates (PFAS), perfluoroalkyl carboxylates
(PFAC), perfluorohexane sulfonic acid (PFHxS) & perfluorooctane
sulfonic acid (PFOS)

37. ©2015 Waters Corporation 37
Xevo TQ-S
Perfluoronated compounds (PFCs)
 Projects to determine the levels of PFCs in
– Mink Liver
– Fish Liver
– Drinking Water
 PFCs present in the environment after use of fire fighting foams
The production, supply, use and disposal of PFOS in Europe is
controlled by the Persistent Organic Pollutants Regulation (EC)
850/2004
Contaminating groundwater or surface water with firewater that
contains PFOS is not permissible
At present there is no lower concentration limit in the EU POPs
Regulations –any material containing any concentration of PFOS
should be disposed of or recovered by one of the prescribed methods
 Evaluation of UPLC with Xevo TQ-S
– Instrument sensitivity
– Explore RADAR Acquisition
– Reduction of matrix effects via sample dilution

38. ©2015 Waters Corporation 38
PFCs analysis using Xevo TQ-S
Fire fighting foams
Seepage water samples
from fire training activity
after cleaning steps (more
complex sample)
Borehole from lake adjacent to
fire training activity at an
airport in a different location
(less complex sample)

39. ©2015 Waters Corporation 39
Instrument sensitivity PFCs
200fg on column
PFBA
PFPeA
PFHpA
PFHxA
PFOA
PFNA
PFDA
PFDoDA
PFUnDA
PFTrDA
PFBuS
PFHxS
PFOS
PFDS

40. ©2015 Waters Corporation 40
Impact of sensitivity
RADAR data for complex samples
Original Sample
x10 dilution
x100 dilution
acquisitions
allow levels of matrix to be
monitored at the same
time as running MRM
experiments
Other matrix components
can affect both
chromatography AND
ionization (ion suppression)

41. ©2015 Waters Corporation 41
PFHxA
x100
x10
x2.5
decrease
x10.2
decrease
802268
318971
31270
Peak Area
Reducing matrix (dilution)
Original Sample (Seepage sample close to FFF source)
x10 dilution
x100 dilution
Expected RT

42. ©2015 Waters Corporation 42
PFHxA (Peak Area vs Matrix Conc)
0
500000
1000000
1500000
2000000
2500000
3000000
3500000
0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1
Relative Matrix Concentration
PeakArea
Ionsuppression
Undiluted sample
~75%

43. ©2015 Waters Corporation 43
Summary
 Monitoring environmental contamination and the advantages of mass
spectrometry?
– Fundamentals and inside the mass spectrometer
– How does all of this work?
– Multiple Reaction Monitoring, RADAR and new ion source designs
 The application of tandem quadrupole instruments to environmental
analytical applications
– Multi- residue pesticide analysis in drinking water
– Acidic herbicides
– Surface water analysis
– Endocrine disruptors
– POPs and perfluorinated compounds
 Sensitivity is not just about detecting lower concentrations of analytes; it’s
also about being able to dilute out interferences
 Mass Spectrometry has become more accessible and routine than ever
before