Raising Confidence in the Analysis of Environmental...

Copyright 2012 Bruker Corporation. All Rights Reserved.

C h e m i c a l a n d A p p l i e d M a r k e t s

Raising Confidence in the Analysis of Environmental

Samples with Tandem Mass Spectrometry

Bruker Daltonics Chemical and Applied Markets

Fremont, CA Ed George



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Inorganic Contaminants



Agenda

• The Process and Power of Multiple Reaction Monitoring (MRM)

• Key Applications for Tandem MS in Environmental Monitoring

• Pyrethroid Insecticides by EI and NCI with Tandem Mass Spectrometry

• PBDE Flame Retardants in Sewage and Ocean Sediments

• EPA Methods 8270 and 525: Raising Confidence in Results with MRM

• Analysis of PCPPs by LC/MS/MS

4



Environmental Monitoring: The Instrument Challenge

• Performance - Must meet performance requirements

• Sensitivity to meet regulatory limits

• Reality: Labs want to exceed those requirements

• Reliability – Must be designed for Production Labs

• Data quality must be impeccable

• Consumer safety and profitability is at stake with every sample

• Robustness – Withstand the Abuse of complex samples

• QuEChERs/8270 sludge extracts

• Hundreds of samples before required maintenance

• Ion source in LC-MS and GC-MS require reduced maintenance



Power of MRM in Environmental Matrices

(mainlib) Parathion

10 20 30 40 50 60 70 80 90 100 110 120 130 140 150 160 170 180 190 200 210 220 230 240 250 260 270 280 290 3000

50

100

15

29

3945

65

75

81

97109

125

139

150

155

172

186

201

218

235

246

263

275

291

N

O

OO

P

O

SO

Parathion-Ethyl

Precursor ion

m/z 291

MRM Transition

Product ion

m/z 109



Q1

Q3

q2

291

291

Parathion

Matrix Interference

Isobaric Interferences: Parathion Example

109



8 © Copyright 2011 Bruker Corporation. All Rights Reserved.

Demeton-S-methyl

Trifluralin

Diazinon

Heptachlor epoxide

Paclobutrazol

Endosulfan-sulfate

Iprodione Halfenprox

Full Scan Scan range: 80Da to 400Da

Pumpkin matrix with 1ppb pesticide spike

SIM @ 263 Da

? ? ?

Extracted @ 263 Da, search for halfenprox

MRM 263 > 235 Da



Pyrethroid Insecticides by GC/MS/MS



Pyrethroid Facts • Pyrethroids are used in household

insecticides/repellents

• Are synthetic compounds based on structure of natural pyrethrins produced by the flowers of pyrethrums (Chrysanthemum cinerariaefolium and C. coccineum).

• Pyrethroids are toxic to fish and other aquatic organisms

• At extremely low levels, such as 2 parts per trillion, pyrethroids are lethal to mayflies, gadflies, and invertebrates that constitute the base of many aquatic and terrestrial food chains.

• Pyrethroids have been found to be unaffected by secondary treatment systems at municipal wastewater treatment facilities in California. They appear in the effluent, usually at levels lethal to invertebrates

• Therefore very low levels of detection are required in complex sediment and waste water effluent extracts



Pyrethroids- EPA and Other Methods

• 8270 is GC/MS full scan and/or SIM- no MSMS!

• 1660 is HPLC with UV Detection- Good luck!

• 1699 is HRGCMS method- Expensive!

• USGS method uses GC/MS and GC/MS/MS (EI)- Good and addresses some analytes that poorly dissociate in MRM

• GC/MS/MS in both EI and CI MRM modes were analyzed with the Bruker SCION for a specific group of pyrethroids

• Calibration and Instrument DLs determined followed by analysis of sewage and sediment sample extracts



Analytical Conditions - GC

Analytical Conditions

Parameter Setting

Column BR-5ms, 15 m x 0.25 mm, 0.25 µm

Gas Flow Helium, 1 mL/min

Column Oven 60 ºC(hold 2min)150 at 50 ºC/min(hold 1 min) 320

at 10ºC/min (hold 3 min)

Injection 2 µL, pulsed splitless, 250ºC



Analytical Conditions – MS- EI


Parameter Setting

Ion Source Temperature 250ºC

Transferline Temperature 280ºC

Collision Gas Argon (1.7 mTorr)

Measurement type Dynamic MRM

Scan Times Calculated by CBS



Analytical Conditions – MS- NCI


Parameter Setting




Reagent Gas Ammonia

CI Gas Pressure 15 psi





EI MRM Transitions Compound Name Q1

Mass Q3 Mass

Collision Energy

trans-Permethrin 183 128 -20



Prallethrin 123 81 -15

Prallethrin 123 79 -15

Fipronil Sulfide 351 255 -20

Esfenvalerate-B 225 91 -25



Allethrin 123 81 -15

Allethrin 123 95 -10

Fipronil-sulfone 383 255 -20

Fipronil-sulfone 383 335 -10

Danitol 181 152 -25

Danitol 181 127 -30

Bifenthrin 181 115 -40



Cyfluthrin isomers 206 151 -15



Cypermethrin isomers

181 127 -30

Cypermethrin isomers

181 152 -20

Deltamethrin 253 172 -10



Esfenvalerate-A 225 91 -25



Fipronil 367 178 -40



cis-Permethrin 183 128 -20



Piperonylbutoxide 176 131 -10



Cyhalothrin 208 181 -10

Cyhalothrin 181 152 -20



NCI MRM Transitions

Compound Name Q1 Mass Q3 Mass Collision Energy

cis-Permethrin 207.2 35.3 10

cis-Permethrin 209.2 37.2 10

trans-Permethrin 207.2 35.2 10

trans-Permethrin 209.2 37.2 10

Fipronyl sulfide 384 314.9 20

Fipronyl sulfide 384 248 25

Fipronyl sulfide 384 244 25

Esfenvalerate-B 211.2 35.2 20

Esfenvalerate-B 211.2 167.2 10

Fipronyl sulfone 415.9 282.9 25

Fipronyl sulfone 415.9 243.9 30

Bifenthrin 386 205 5

Bifenthrin 386 161 15

Cypermethrin isomers 207.2 35.2 20

Cypermethrin isomers 171.2 35.2 20

Deltamethrin 296.9 79 2

Deltamethrin 296.9 81 2

Deltamethrin 81.4 81.4 1

Deltamethrin 79.4 79.4 1

Allethrin 167.4 167.4 1

Danitol 141.4 141.4 1

Cyfluthrin isomers 207.2 35.2 20

Cyfluthrin isomers 171.2 35.2 20

Esfenvalerate-A 211.2 167.2 10

Esfenvalerate-A 211.2 35.2 20

Pallethrin 167.4 167.4 1



Good Isomer Separation on 15 M column

EI

CI



EI-MRM Calibration 0.1-50 ppb Compound Name Corr. Avg. RRF % RSD

Fipronil Sulfide 0.9993 0.076 5.0

Allethrin 0.9998 0.020 14.2

Prallethrin 0.9989 0.019 14.2

Fipronil-sulfone 0.9992 0.050 4.6

Fipronil 0.9989 0.026 13.5

Piperonylbutoxide 0.9992 0.158 5.7

Bifenthrin 0.9992 9.58 7.5

Danitol 0.9996 0.943 8.0

Cyhalothrin 0.9995 1.41 6.2

cis-Permethrin 0.9998 1.54 6.1

trans-Permethrin 0.9994 0.675 6.3

Cyfluthrin isomers 0.9997 0.604 13.4

Cypermethrin isomers 0.9996 0.896 10.6

Esfenvalerate-A 0.9999 0.352 9.9

Esfenvalerate-B 0.9999 0.584 4.7

Deltamethrin 0.9996 0.710 8.2

Mean 0.9995 1.10 8.7



EI-MRM IDLs

Parameter Rep 1 Rep 2 Rep 3 Rep 4 Rep 5 Rep 6 Rep 7 mean sd Calculated

IDL Target

concentration % RSD

Fipronil Sulfide 0.078 0.08 0.077 0.074 0.075 0.075 0.076 0.076429 0.00207 0.01 0.1 2.7

Allethrin 0.525 0.574 0.576 0.584 0.608 0.565 0.498 0.561429 0.03744 0.12 0.5 6.7

Prallethrin 0.953 1.018 1.084 1.077 0.93 1.011 1.127 1.028571 0.07183 0.23 1 7.0

Fipronil-sulfone 0.098 0.088 0.074 0.107 0.091 0.089 0.074 0.088714 0.01197 0.03 0.1 13.5

Fipronil 0.514 0.524 0.499 0.551 0.501 0.486 0.449 0.503429 0.03186 0.10 0.5 6.3

Piperonylbutoxide 0.092 0.09 0.094 0.083 0.097 0.094 0.081 0.090143 0.00598 0.02 0.1 6.6

Bifenthrin 0.087 0.09 0.081 0.094 0.084 0.075 0.096 0.086714 0.00739 0.02 0.1 8.5

Danitol 0.075 0.083 0.099 0.091 0.081 0.069 0.085 0.083286 0.00989 0.03 0.1 11.9

Cyhalothrin 0.1 0.091 0.1 0.081 0.09 0.08 0.115 0.093857 0.01227 0.04 0.1 13.1

cis-Permethrin 0.081 0.091 0.1 0.071 0.084 0.074 0.121 0.088857 0.01726 0.05 0.1 19.4

trans-Permethrin 0.519 0.662 0.413 0.555 0.503 0.649 0.492 0.541857 0.0887 0.28 0.5 16.4

Cyfluthrin isomers 1.078 1.101 1.183 1.178 1.222 1.314 1.302 1.196857 0.09062 0.28 1 7.6

Cypermethrin isomers 0.524 0.552 0.553 0.476 0.553 0.481 0.406 0.506429 0.05535 0.17 0.5 10.9

Esfenvalerate-A 0.459 0.593 0.528 0.455 0.622 0.565 0.426 0.521143 0.07593 0.24 0.5 14.6

Esfenvalerate-B 0.101 0.1 0.108 0.1 0.083 0.066 0.123 0.097286 0.0182 0.06 0.1 18.7

Deltamethrin 0.489 0.525 0.556 0.482 0.503 0.548 0.491 0.513429 0.02983 0.09 0.5 5.8

Mean 0.11 10.6



NCI-MRM Calibration 0.1-50 ppb

Compound Name Corr. Avg. RRF % RSD

Fipronyl sulfide 0.9987 54.5 14.1

Allethrin 0.9983 12.7 11.2

Pallethrin 0.9988 8.38 8.0

Fipronyl sulfone 0.9995 2.86 5.3

Fipronil 0.9982 4.83 17.2

Bifenthrin 0.9963 14.1 12.5

Danitol 0.9991 17.0 9.3

Cyhalothrin 0.9997 0.12 2.3

cis-Permethrin 0.9994 0.12 11.0

trans-Permethrin 0.9990 0.01 9.6

Cyfluthrin isomers 0.9988 0.10 11.2

Cypermethrin isomers 0.9991 0.08 6.5

Esfenvalerate-A 0.9997 2.08 7.4

Esfenvalerate-B 0.9997 3.34 9.5

Deltamethrin 0.9997 7.01 11.9

Average 0.9989 8.49 9.8



NCI-MRM Low Calibration 1-5000 ppt

Compound Name Corr. Avg. Resp % RSD

Fipronyl sulfide 0.9995 2202 18.8

Fipronyl sulfone 0.9997 139 27.2

Bifenthrin 0.9998 590 14.7

Danitol 0.9998 801 21.9

Esfenvalerate-A 0.9998 1166 8.0

Esfenvalerate-B 0.9995 1993 18.6

Deltamethrin 0.9985 4881 17.9

Average 0.9995 1682 18.1



NCI-MRM IDLs


IDL Target

concentration % RSD

Fipronil Sulfide 0.112 0.106 0.112 0.112 0.106 0.102 0.108 0.11 0.0039 0.01 0.1 3.6

Allethrin 0.104 0.123 0.116 0.115 0.111 0.102 0.116 0.11 0.00737 0.02 0.1 6.6

Prallethrin 0.082 0.092 0.092 0.097 0.094 0.084 0.092 0.09 0.00541 0.02 0.1 6.0

Fipronil-sulfone 0.105 0.101 0.104 0.107 0.098 0.096 0.103 0.10 0.00392 0.01 0.1 3.8

Bifenthrin 0.113 0.11 0.116 0.114 0.107 0.1 0.11 0.11 0.00532 0.02 0.1 4.8

Danitol 0.113 0.102 0.111 0.112 0.108 0.106 0.119 0.11 0.00546 0.02 0.1 5.0

Cyhalothrin 0.106 0.106 0.109 0.095 0.091 0.088 0.112 0.10 0.00949 0.03 0.1 9.4

cis-Permethrin 0.127 0.119 0.126 0.124 0.118 0.108 0.128 0.12 0.00707 0.02 0.1 5.8

trans-Permethrin 0.437 0.618 0.654 0.58 0.588 0.757 0.608 0.61 0.09558 0.30 0.5 15.8

Cyfluthrin isomers 0.144 0.133 0.155 0.141 0.14 0.129 0.136 0.14 0.00844 0.03 0.1 6.0

Cypermethrin isomers 0.101 0.087 0.096 0.108 0.099 0.09 0.092 0.10 0.0072 0.02 0.1 7.5

Esfenvalerate-A 0.11 0.105 0.111 0.106 0.103 0.098 0.107 0.11 0.00439 0.01 0.1 4.1

Esfenvalerate-B 0.111 0.109 0.114 0.111 0.106 0.1 0.108 0.11 0.0045 0.01 0.1 4.2

Deltamethrin 0.105 0.103 0.098 0.098 0.094 0.109 0.103 0.10 0.00506 0.02 0.1 5.0

Fipronil 0.103 0.098 0.102 0.104 0.098 0.104 0.109 0.10 0.00382 0.01 0.1 3.7

Mean 0.04 6.1



NCI-MRM Low-Level IDLs (Units in ppt)


IDL Target

concentration %

RSD

Fipronil Sulfide 8.317 9.167 9.236 8.823 9.644 9.392 9.437 9.1 0.44596 1.4 10 4.9

Fipronil-sulfone 7.156 7.928 11.07 7.112 7.369 9.277 7.097 8.1 1.5072 4.7 10 18.5

Bifenthrin 11.55 12.21 10.55 10.81 11.17 12.4 11.83 11.5 0.69677 2.2 10 6.1

Danitol 8.442 8.304 9.949 10.84 8.639 7.802 7.844 8.8 1.13907 3.6 10 12.9

Esfenvalerate-A 10.21 11.29 11.18 10.69 11.2 11.69 11.3 11.1 0.4827 1.5 10 4.4

Esfenvalerate-B 10.42 10.57 10.97 11.32 10.84 11.24 10.82 10.9 0.32765 1.0 10 3.0

Deltamethrin 8.408 6.26 6.48 8.945 7.064 8.161 7.217 7.5 1.01632 3.2 10 13.5

Fipronil 9.032 9.28 11.94 11.35 10.17 11 10.08 10.4 1.07353 3.4 10 10.3

Mean 2.6 9.2



NCI Sensitivity Excellent for Select Cmpds

1 ppt Fipronyl Sulfide 10 ppt Bifenthrin



NCI-MRM: Excellent vs. Matrix

Ocean Sediment Spike

Sewage Effluent Spike

Cal Standard



EI-MRM: Matrix effects results

Ocean Sediment

Sewage Effluent

Cal Standard



EI-MRM: Matrix Closer Look

Ocean Sediment

Ocean Sediment

Sewage

Sewage

Cal standard

Cal standard



NCI: Allethrin/Pallethrin

Ocean Sediment

Sewage

Cal Standard

Poorly dissociates in MS/MS (m/z 167) however ‘parent to parent’ with Ar in cell cleans up the matrix nicely



EI-MRM: Allethrin/Pallethrin Extreme matrix interference at MRM transitions studied!

Ocean Sediment

Sewage

Cal Std



Deltamethrin- Both Techniques

EI-MS/MS NCI-MS/MS

Ocean Sediment

Sewage

Cal Standard



Pyrethroid Analysis- Lessons Learned

• MS/MS will provide good confidence in your data, at extreme low levels, however…. • Consider your test matrix and added discrimination

using NCI, especially approaching sub ppb levels and very dirty extracts

• Poorly dissociated cmpds can still benefit ‘parent-to-parent ‘ transitions vs. SIM alone!

• Not all cmpds will work well by NCI, look at structure and optimize conditions

• Watch out for ‘matrix enhanced chromatographic response’ high bias NOT due to detection technique but rather matrix shielding of active sites in injection and chromatographic pathway



Brominated Flame Retardants by GC/MS/MS



PBDE Facts

• Used as flame retardants in a variety of products

• Tend to bio-accumulate in tissues, breast milk

• Shown to be related to increased infertility rates in humans at levels that are normally found in US households

• Studies in Canada have found significant concentrations of PBDEs in common foods such as salmon, ground beef, butter, and cheese

• PBDEs have also been found at high levels in indoor dust, sewage sludge, and effluents from wastewater treatment plants. Increasing PBDE levels have been detected in the blood of marine mammals such as harbor seals.



PBDEs- Method Considerations

• EPA 1614A is HRGCMS- Expensive!

• EPA 8081 is GC with ECD Detection- Background!

• EPA 527 is GCMS method- No MSMS!

• USGS method uses GC/MS and GC/MS/MS (EI)

• GC/MS/MS in EI MRM mode was analyzed with the Bruker SCION for a specific group of PBDEs

• Calibration and Instrument DLs determined followed by analysis of sewage and sediment sample extracts



Analytical Conditions - GC


Parameter Setting

Column BR-5ms, 15 m x 0.25 mm, 0.25 µm

Gas Flow Helium, 1 mL/min

Column Oven 60 ºC(hold 2min)150 at 50 ºC/min(hold 1 min) 325

at 10ºC/min (hold 8 min)

Injection 2 µL, pulsed splitless, 290ºC



Analytical Conditions – MS- EI


Parameter Setting








MRM Transitions

Compound Name Q1 Mass

Q3 Mass

Collision Energy

BDE-71 486 326 -25

BDE-71 326 138 -45

SS-BDE-209-C13 970.5 811 -20

SS-BDE-209-C13 811 651 -45

BDE-154 644 484 -20

BDE-154 484 377 -45

BDE-209 960.5 799 -20

BDE-209 799 639.5 -45

BDE-153 644 484 -20

BDE-153 484 377 -45

BDE-47 486 326 -25

BDE-47 326 138 -45

BDE-28 406 246 -20

BDE-28 246 139 -30

SS-BDE-4FHBE 662 502 -25

BDE-66 486 326 -25

BDE-66 326 138 -45

BDE-138 644 484 -20

BDE-138 484 377 -45

IS-BDE-4FTBE 504 344 -20

BDE-100 564 404 -20

BDE-100 404 137 -45

BDE-183 722 562 -25

BDE-183 562 455 -45

BDE-49 486 326 -25

BDE-49 326 138 -45

BDE-99 564 404 -20

BDE-99 404 137 -45

BDE-17 406 246 -20

BDE-17 246 139 -30

BDE-190 722 562 -25

BDE-190 562 455 -45

BDE-85 564 404 -20

BDE-85 404 137 -45



TIC MRM Chromatogram- 50 ppb std



MRM Calibration 0.1-50 ppb Compound Name Corr. Avg. RRF % RSD

BDE-17 0.9997 2.04 6.7

BDE-28 0.9986 5.26 5.0

BDE-49 0.9990 2.64 8.7

BDE-71 0.9985 2.70 8.0

BDE-47 0.9989 2.48 5.6

BDE-66 0.9988 1.86 6.3

BDE-100 0.9988 1.00 4.9

BDE-99 0.9989 0.87 8.0

BDE-85 0.9960 0.72 6.4

BDE-154 0.9990 0.55 7.8

BDE-153 0.9985 0.40 5.8

SS-BDE-4FHBE 0.9977 0.21 11.0

BDE-138 0.9985 0.39 8.7

BDE-183 0.9983 0.31 17.2

BDE-190 0.9965 0.28 15.0

SS-BDE-209-C13 0.9981 0.07 17.5

BDE-209 0.9989 0.08 11.2

Mean 0.9984 1.29 9.1



MRM IDLs


IDL Target

concentration % RSD

BDE-17 0.102 0.104 0.116 0.099 0.1 0.097 0.11 0.10 0.00676 0.02 0.1 6.5

BDE-28 0.089 0.086 0.091 0.093 0.088 0.088 0.081 0.09 0.00383 0.01 0.1 4.4

BDE-49 0.095 0.114 0.105 0.095 0.101 0.09 0.09 0.10 0.00873 0.03 0.1 8.9

BDE-71 0.07 0.069 0.068 0.071 0.07 0.069 0.068 0.07 0.00111 0.01 0.1 1.6

BDE-47 0.094 0.096 0.096 0.098 0.093 0.086 0.085 0.09 0.00509 0.02 0.1 5.5

BDE-66 0.101 0.076 0.073 0.083 0.072 0.074 0.074 0.08 0.01036 0.03 0.1 13.1

BDE-100 0.081 0.096 0.075 0.091 0.083 0.099 0.071 0.09 0.01056 0.03 0.1 12.4

BDE-99 0.1 0.103 0.083 0.101 0.104 0.085 0.104 0.10 0.00912 0.03 0.1 9.4

BDE-85 0.105 0.119 0.108 0.121 0.104 0.099 0.099 0.11 0.00891 0.03 0.1 8.3

BDE-154 0.097 0.088 0.103 0.086 0.097 0.094 0.085 0.09 0.00672 0.02 0.1 7.2

BDE-153 0.113 0.106 0.113 0.09 0.081 0.09 0.092 0.10 0.01269 0.04 0.1 13.0

SS-BDE-4FHBE 0.114 0.093 0.104 0.096 0.082 0.078 0.106 0.10 0.01302 0.04 0.1 13.5

BDE-138 0.459 0.434 0.33 0.424 0.36 0.377 0.384 0.40 0.04537 0.14 0.5 11.5

BDE-183 0.114 0.116 0.102 0.12 0.113 0.115 0.12 0.11 0.00607 0.02 0.1 5.3

BDE-190 0.463 0.464 0.413 0.4 0.396 0.365 0.359 0.41 0.04211 0.13 0.5 10.3

SS-BDE-209-C13 4.783 4.333 4.539 4.108 4.215 4.706 4.665 4.5 0.26166 0.82 5 5.8

BDE-209 4.068 4.282 3.915 4.117 3.912 4.324 4.276 4.1 0.1731 0.54 5 4.2

Mean 0.11 8.3



TIC MRM in Matrix-1

Ocean Sediment

Sewage extract

CAL Standard



TIC MRM in Matrix-2

Ocean Sediment

CAL Standard

Sewage Extract



TIC MRM in Matrix-3

Ocean Sediment

Sewage Extract

CAL standard

BDE-209



PBDE Analysis by GC/MS/MS

• MS/MS provides excellent discrimination against the matrices studied to provide good confidence in your data

• Be aware however of ‘matrix enhanced chromatographic response’

• BDE-209 sensitivity can be improved with thinner phase and even shorter column to reduce elution temperature by minimizing thermal degradation



EPA Methods 8270 and 525: Raising Confidence in Results with Tandem Mass Spectrometry



Method 8270: Want Lower Detection Levels?

• Method is for a wide variety of compounds in complex matrices

• Contractors and labs may want lower reporting limits than method was validated for

• Labs can use SIM, but what if isobaric interferences in heavy matrix?

• MS/MS can give added specificity and confidence in results at low levels of detection



8270 Test Sludge Extract

PCP injection 1

PCP injection after 35

Left- Blank Extract; Right-Sludge Extract



Sludge Sample Injection: Full Scan

TIC of highly contaminated soil extract from EPA 8270. The sample was spiked with 100 pg per component for precision testing by Tandem Mass Spectrometry



MS/MS to Find Needles in the Haystack

MS/MS on SCION in soil extract, 15-20 pg injected on column – pulsed-split injection

MS/MS on same Extract Previous Slide



Method Precision Examples: 100 Injections at 100 ppb in Sludge Extract

1-Chloronaphthalene

%RSD = 5.7%

0

10000

20000

30000

40000

50000

60000

70000

0 20 40 60 80 100 120

Injection Number

Raw

Peak A

rea

Series1

2,4-Dichlorophenol

%RSD = 4.7%

0

1000

2000

3000

4000

5000

6000

7000

8000

9000

10000

0 20 40 60 80 100 120

Injection Number

Raw

Peak A

rea

Series1

4-Bromophenoxy Ether

%RSD = 6.6%

0

2000

4000

6000

8000

10000

12000

14000

16000

18000

20000

0 20 40 60 80 100 120

Injection Number

Raw

Peak A

rea

Series1



Method 525: Drinking Water (DW) Method

• Method covers wide variety of pesticides and industrial pollutants

• Current version written as Full Scan with SIM target analytes for lower reporting limits

• Method applies to finished DW; however, often used to test source waters as in past method version (525.2) (ie. Surface and Ground Waters)

• Method concentrates analytes (and matrix) by SPE and inject 1-2 uL into GC/MS

• MS/MS can give better specificity and confidence in results at low levels of detection



Tandem MS and SIM in STD Look the Same…

B

A

TIC of over one hundred compounds in EPA 525.2 in MS/MS (A) and SIM (B) acquisition modes at concentration of 500 ng/mL in a pure solvent.



But in Surface Water Extracts…

Actual surface water source sample submitted for Method 525 analysis. Many water treatment plants are required to monitor source waters for regulated and unregulated contaminants.

FULL SCAN



Chromatograms Showing d-BHC in Surface Water Extract

MRM mode in solvent

MRM mode In matrix

SIM mode in solvent

SIM mode in matrix



Chromatograms Showing Triademefon in Surface Water Extract

SIM mode in solvent

MRM mode in solvent

SIM mode in matrix

MRM mode In matrix



SIM vs. MS/MS Recovery Comparisons

Note: Spiked in the matrix at 50 ng/mL (n=5)

Spiked in SW measured by SIM

Spiked SW measured by MRM

Peak Name RT (min) Measured %RSD Measured %RSD

Chlorprophan 16.26 1199. 6024 64.51 1.35

a-BHC 16.76 1014. 6.52 44.38 7.45

b-BHC 17.35 135. 7.35 46.74 5.94

g-BHC 17.56 313. 5.30 45.19 4.09

d-BHC 18.20 ND ND 45.11 3.88

Metribuzin 18.76 266. 5.69 66.27 1.81

MGK-264 21.06 75. 6.50 54.76 4.96

Endosulfan II 24.16 104. 17.74 51.34 6.37

Average 365. 7.45 59.50 3.84



Conclusions

• Full Scan and SIM are techniques required in Methods 8270 and 525

• However, lower reporting limits and heavy matrix challenge this requirement

• Tandem MS provides lower detection limits and confidence in results, eliminating isobaric interferences and potential erroneous reporting (high-biased)



Analysis of Trace PPCPs in Water Using The EVOQ™ TQ LC-MS/MS System with OLE



Bruker EVOQ™ With OLE for PPCPs in Water

• PPCPs refers to a broad group of chemical substances used for health or cosmetic

reasons, or used by agribusiness to boost growth or health of livestock

• PPCPs= Pharmaceuticals and Personal Care Product

• Wastewater from households and industrial sources contains multiple PPCPs of various combinations, concentrations and relative toxicologies

• PPCPs are bioactive, they do not dissolve easily or evaporate at normal temperatures or pressures, so can easily enter the soil and aquatic environments via sewage, biosolids and irrigation

• Evidence suggests PPCPs are linked to some ecological damage such as the delayed development in fish and delayed metamorphosis in frogs

• Conventional methods of PPCP detection requires the pre-concentration of large volume water samples and tedious solid phase extraction (SPE) clean up to achieve the low ng/L (ppt) level detection

• This study demonstrates how using UHPLC with an integrated on-line extraction (OLE) option coupled to LC-MS/MS can be used for the detection of PPCPs

• The on-line extraction module enables convenient method-driven on-line sample cleanup or sample pre-concentration



Bruker EVOQ™ With OLE for PPCPs in Water Experimental Conditions

Chromatography parameters (Advance UHPLC OLE)

Trap column YMC-Pack ODS-AQ, 3µm,

35 mm x 2.0 mm I.D.

Column temperature 40°C

Injection volume 400 µL

Flow rate 400 µL/min

Solvent A 2 mM ammonium

formate, 0.1% FA in water

Solvent B 2 mM ammonium

formate, 0.1% FA in MeOH

Solvent C 2 mM ammonium

formate, 0.1% FA in water

Gradient conditions 0.0 min, 10% B

0.2 min, 10% B

0.8 min, 25% B

8.0 min, 95% B

9.0 min, 95% B

9.1 min, 10% B

12.0 min, 10% B

Mass spectrometer parameters (EVOQ Elite)

HV 4000 V

Cone gas flow 15 units

Cone gas temperature 300°C

Heated probe gas flow 40 units

Heated probe temperature 450°C

Nebulizer gas flow 50 units

Exhaust gas On

Q2 pressure 1.5 mTorr (Argon)

Sample Preparation: • Water sample acidified with

0.1% FA and filtered (0.45um)



Bruker EVOQ™ With OLE for PPCPs in Water Target Compounds



Bruker EVOQ™ With OLE for PPCPs in Water Selected PPCPs at 2 ppt



Bruker EVOQ™ With OLE for PPCPs in Water Linearity, 1 ppt – 100 ppt

Trimethoprim

R2=0.999

Carbamazepine

R2=0.999

Naproxen

R2=0.999

Triclosan

R2=0.998

Gemfibrozil

R2=0.997

Sulfamethoxazole

R2=0.999



Bruker EVOQ™ With OLE for PPCPs in Water Summary of PPCPs Calibration Curves

Compound Name

RT (min)

ESI R2

S/N (p/p) 2 ppt

RSD n=10

(5ppt)

Trimethoprim 1.66 + 0.994 389 5.80% Hydroxy Atrazine 1.88 + 0.997 61 9.20%

Thiabendazole 1.93 + 0.996 408 8.70% Ciproxacin 2.03 + 0.999 25 4.90%

Caffeine 2.19 + 0.995 68 8.00% Sildenafil 2.33 + 0.998 132 6.60%

Sulfamethoxazole 2.51 + 1 68 5.00% Cyanazine 4.1 + 0.998 201 8.70% Simazine 4.38 + 1 56 9.40%

Metribuzin 4.39 + 0.999 26 12.50% Hexazinone 4.42 + 0.997 553 4.90% Dapoxetine 4.62 + 0.999 268 6.00% Bentazone 4.69 - 0.999 360 6.20% Ametryn 4.73 + 0.997 404 5.70%

Carboxine 4.8 + 0.999 217 6.20% Carbamazepine 4.85 + 0.999 485 4.50%

Atrazine 5.2 + 0.996 39 6.90% Alpazolam 5.38 + 0.998 178 5.90%

Diuron 5.54 + 0.999 56 3.60% Prometryn 5.56 + 0.994 376 8.40%

2,4-D 5.7 - 1 26 5.60% MCPA 5.81 - 1 56 3.80%

Mecoprop 6.39 - 1 27 2.90% Metolachlor 6.76 + 0.996 177 9.20% Pyriproxifen 8.36 + 1 23 3.50%



Bruker EVOQ™ With OLE for PPCPs in Water Summary of PPCPs in Water Samples

Compound Name Tap Water 1 Tap Water 2 Creek Water

Bottle Water

pg/mL (ppt) Trimethoprim <2 <2 5 <2

Hydroxy Atrazine 4 <2 7 <2 Thiabendazole ND <2 <2 <2

Ciproxacin ND ND ND ND Caffeine ND <2 <2 10 Sildenafil ND ND ND <2

Sulfamethoxazole <2 <2 ND <2 Cyanazine ND ND ND <2 Simazine 3 <2 5 ND

Metribuzin ND ND ND ND Hexazinone 17 3 3 ND Dapoxetine ND ND ND ND Bentazone ND ND ND ND Ametryn ND ND <2 ND

Carboxine ND ND ND ND Carbamazepine <2 <2 <2 ND

Atrazine <2 ND ND ND Alpazolam ND ND ND ND

Diuron 9 <2 6.2 ND Prometryn ND ND ND <2

2,4-D 9 <2 13 <2 MCPA <2 <2 <2 ND

Mecoprop <2 <2 11 2 Metolachlor 22 <2 <2 <2 Pyriproxifen ND <2 ND <2



Bruker EVOQ™ With OLE for PPCPs in Water Summary

• All of the PPCPs studied were detected at 2 ppt or better with an injection of 400 uL water samples, with a linear response range up to 200 or 500 ppt. • Replicate injections with 5 ppt level spiked in tap water

demonstrated excellent robustness

• This new method illustrates how modern UHPLC with

OLE and LC-MS/MS can provide the sensitivity demanded by regulatory bodies, even with challenging ranges of samples at various concentrations

• The technique also presents a more convenient and simpler approach to the analysis of PPCPs compared to traditional solid phase extraction techniques



Conclusions

• The power of Multiple Reaction Monitoring (MRM) in Environmental Samples • Can reduce detection/reporting limits and

provide more confidence in your data

• Enables matrix discrimination in samples against isobaric interferences that will occur with SIM alone

• Other techniques such as NCI as shown in pyrethroids may be required to eliminate matrix interferences

• Keep in mind that your matrix needs some study for optimized MRM transitions and/or ionization techniques for best results

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