Quantitation of PFAS using MALDI-TOF MSBrooke Dilmetz
Per and polyfluoroalkyl substances• PFOS and PFOA manufactured from early 1950’s – present1
• Highly persistent and bioaccumulative
University of Adelaide 2
PFOS (Perfluorooctanesulfonate)
PFOA (Perfluorooctanoate)
1 Oliaei et al. (2013). Environ Sci Pollut Res
Per and polyfluoroalkyl substances• PFOS and PFOA manufactured from early 1950’s – present1
• Germany, Italy and China• Highly persistent and bioaccumulative
University of Adelaide 3
PFOS (Perfluorooctanesulfonate)
PFOA (Perfluorooctanoate)
- Evidence that PFAS:• Disrupt normal endocrine activity;• Reduce immune function;• Cause adverse effects on multiple organs, including the liver and
pancreas; • Cause developmental problems in rodent offspring exposed in the
womb.2,3
2Lindstrom AB, Strynar MJ, Libelo EL. (2011). Environ Sci Technol
3White SS, Fenton SE, Hines EP. (2011). J Steroid Biochem
Measuring PFAS: LCMS • Liquid chromatography coupled tandem mass spectrometry
University of Adelaide 4
1. Zheo et al. (2011). Mircochem.2. Shimadzu, Application note C813. Adelaide Proteomics Centre, 20174. Cao et al. (2011). Talanta5. Food standards Australia, 2017
Drinking water quality guideline5
PFOS PFOA
0.07µg/L 0.56µg/L
Advantages• Sensitive/Selective • Low detection (LOD 0.03 – 3.0 ng/L1,2)
Disadvantages• Cost ($200.00-400.00/sample3)• Time - column conditioning,
- carry-over• Contamination of fittings and tubing4
University of Adelaide 5
• Typical analytes are macromolecules (proteins, polymers)• Limited for the detection of small molecules (M.W. <500 Da)• Analyte co-deposited with matrix
PFAS analysis by MALDI-TOF MS
Advantages• High throughput MS analysis:
– typically 2-10 sec/spot
• Able to analyse multiple samples:– 1536 spot plate format (800µm)
• No carry-over:– Straight-forward target washing
• Low Cost ($5-10 per sample)
University of Adelaide 6Technical Note # TN-41, Bruker Daltonics GmbH
Disadvantages: small molecule analysis
• Quantitation is difficult, due to:o Matrix interference for small
molecules o Spot-spot variability
PFAS analysis by MALDI-TOF MS
University of Adelaide 7
PFAS analysis by MALDI-TOF MS
Project Aims
To create a screening platform for the detection of PFAS by MALDI-TOF MS
University of Adelaide 8
1. Determine suitable matrices for PFAS detection 2. Determine LODs/ LOQs for various PFAS
3. Quantify of PFAS spiked into water samples4. Apply methodology to contaminated water sample
MALDI Matrix Properties
Name Other Names Applications
α-Cyano-4-hydroxycinnamic acid CHCA Peptides and proteins
2,5-Dihydroxybenzoic acid 2,5-DHB Peptides, phosphoprotein digests, glycoprotein digests, proteins, glycans
2,5-Dihydroxyacetophenone 2,5-DHAP Proteins
Sinnapinic acid SA Proteins, lipids
3-Hydroxypicolinic acid 3-HPA Oligonucleotides
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• Strong UV light absorption at the wavelength of the laser (355nm)• Typically low molecular weight acid• Different matrices for different analyte classes – ionisation threshold
Matrices for PFAS Detection
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Cao et al. (2011) Paper
TMGN Replicate 1
TMGN Replicate 2
TMGN Replicate 3
TMGN Replicate 4
TMGN Replicate 5
Matrices for PFAS Detection
University of Adelaide 11
Cao et al. (2011) Paper
Matrices used for detection of PFAS
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Matrix Advantages Disadvantages
CHCA Homogenouscrystallisation
Interfering matrix ion peaks
Novel Proton Sponge
Less interfering
matrix peaks
Heterogeneous crystallisation
TMGN Homogenouscrystallisation
Interfering matrix ion peaks
• Matrix-free applications• NALDI• Monoliths
CHCA for PFOS
Novel Matrix for PFOA Novel Matrix for PFHxS
Quantitation of PFAS by MALDI-TOF MSPFOS -Matrix: CHCA
y = 5.1488x ‐ 0.4286R² = 0.99980
20
40
60
0 2 4 6 8 10 12
Average Intensity
Ra
tio
Concentration PFOS (ng/uL)
PFOA -Matrix: Novel proton sponge
y = 2.3366x ‐ 0.4994R² = 0.988
0
5
10
15
20
25
30
0 2 4 6 8 10Average Intensity
Ratio
Concentration of PFOA (ng/µL)
PFHxS -Matrix: Novel proton sponge
y = 0.7298x ‐ 0.0295R² = 0.9997
0
2
4
6
8
10
0 2 4 6 8 10Average Intensity
Ratio
Concentration PFHxS (ng/µL)
• Above detection limits of drinking water
• All 3 compounds show linearity – (0.1-10ng/µL)
MC
Slide 13
MC2 You need to have the ccalculated concentration hereMark Condina, 7/09/2017
Workflow for extraction of samples
University of Adelaide 14
SPE of contaminated water sample
University of Adelaide 15
• 10 replicates per concentration• No heavy labelled standard
PFOS Evaluation
SPEContaminated
Water Concentration
(µg/L)
LC-MSContaminated
Water Concentration
(µg/L)
% Difference
1278.69 1170 8.5
y = 16357x + 9423.6R² = 0.9862
0
50000
100000
150000
200000
250000
0 2 4 6 8 10 12
Average Intensity
Concentration of PFOS (ng/uL)
PFOS Calibration Contaminated Water (After SPE)
Matrix free: NALDI application• Nanostructured laser desorption ionisation • Disposable target • Low mass organic molecules (e.g. 100-2000 m.w.) ion target• Surface deposited layer of inorganic nanostructures
University of Adelaide 16
Daniels, H., Dikler, S. Technical Note 22, Bruker Daltonics, and NanoSys Inc.
Quantitation of PFOS by NALDI-TOF
• Able to achieve linearity for same concentration range as with matrix• Matrix: NALDI target
y = 1.3584x + 0.0997R² = 0.9912
0.00
2.00
4.00
6.00
8.00
10.00
12.00
14.00
16.00
18.00
0 2 4 6 8 10 12
Average Intensity
Ratio
Concentration of PFOS (ng/µL)
MC
Slide 17
MC2 You need to have the ccalculated concentration hereMark Condina, 7/09/2017
PFC evaluation by NALDI-TOF MS: screening• PFOS Threshold limit: 0.07µg/L
University of Adelaide 18
‐Matrix free allows lower levels of detection for PFOS-Will be used to determine LOQ
498.915
0
500
1000
1500
2000
2500
Inte
ns. [
a.u.
]
498.5 498.6 498.7 498.8 498.9 499.0 499.1 499.2m/z
Conclusions• Quantitate using MALDI-TOF MS
– Possible to achieve quant without labelled standard (higher variance)– Able to detect down to required thresholds using matrix free platform
• MALDI-TOF MS offers high-throughput assessment of samples after SPE enrichment– Able to screen 1000’s samples low cost
• More comprehensive sampling of potentially contaminated areas• Able to determine which samples to be analysed by LCMS (if required)
University of Adelaide 19
Future work• Improve LOQ and LOD for MALDI and determine for NALDI • Improve MALDI-TOF MS acquisition
– Lower acquisition time per spot
• Evaluate alternative enrichment strategies for PFAS– Porous monolithic material for enrichment
• Investigate other matrix-free applications:– Porous monolithic surfaces – Laser desorption/ionisation (LDI)
• Poly(butyl methacrylate-co-ethylenedimethacrylate) monolith• Poly(styrene-co-divinyl benzene) monolith
University of Adelaide 20
Acknowledgements
Prof. Peter HoffmannDr. Mark CondinaDr. Dario ArruaDr. Lyron Winderbaum Silvana Napoli
Contact information: [email protected]
Yin Ying HoParul MittalChristopher Cursaro Matthew BriggsMitchell AcklandChao ZhangNoor Alia LokmanCarly Gregor
Dr. Richard Stewart