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ATMOSPHERIC PRESSURE IONIZATION MASS SPECTROMETRY FOR GC (APGC-MS/MS): AN ENABLING TECHNOLOGY FOR DETECTION OF CONTAMINANTS OF CONCERN Michael S. Young, Lauren Mullin and Jeremy C. Shia Waters Corporation, 5 Technology Drive, Milford, MA USA michael_s_young@waters.com

INTRODUCTION Gas chromatography coupled with mass-spectrometry (GC-MS) has long been the workhorse for environmental analysis. Fragmentation using traditional electron-ionization (EI) provides a wealth of structural information. Substantial archived library information is available to aid the analyst with unknown identification. Tandem quadrupole (MS/MS) and quadrupole time of flight systems (qTof/MS) offer better sensitivity and selectivity compared with single quadrupole systems. Although these techniques have been successfully used for GC-MS using EI, alternative methods of ionization may be more suitable for trace level quantitative and qualitative analysis. Over the last few decades, LC-MS methods have become a very important environmental analysis tool using electrospray ionization (ESI) or atmospheric pressure chemical ionization (APCI). An important advantage to these ionization modes is that a molecular ion or protonated molecular ion can usually be produced at very high abundance and is typically well preserved. With EI, this is not usually the case; one must choose among many lower abundance fragment ions for transmission to the second quadrupole or Tof. Thus, higher sensitivity and selectivity are possible with ESI or APCI. Atmospheric pressure ionization mass spectrometry for GC (APGC) provides mass-spectrometric data comparable to APCI data used for LC-MS. As in tandem LC-MS, cone-voltage can be optimized to produce mainly molecular ion (for quantitative analysis) or in-source fragmentation (for qualitative analysis). As in tandem LC-MS, collision cell energy can be optimized to produce fragments best suited for quantitation. Another advantage is that the same mass-spectrometer can be used for both LC and GC applications. In this presentation we will discuss some applications of this new APGC technology including dioxins, PCBs, brominated flame retardants (BFRs) and pesticides. The instruments used for this poster are presented below.

WHAT IS APGC ? Waters APGC is an optional ion source for Xevo and

SYNAPT systems that provides highly sensitive GC-MS/MS performance

APGC ionization is soft and molecular ions are more readily detected compared with electron impact ionization (EI)

Fragmentation can be induced (CID) to provide information for structural elucidation

It is very easy to swap between APGC-MS and UPLC-MS in a matter of minutes– No venting required– LC-MS and GC-MS can be performed using the same mass-

spectrometer

APGC analysis can usually be performed at higher flowrates, up to 3 mL/min Helium, to achieve faster chromatography compared with EI

Mass Analyser

Corona dischargeat needle creates plasma

N2 make-up gas deliveredthrough transfer line from GC oven

N2 meets GC eluent flow at transfer line tip

Analyte molecules are ionized after GC elution and directed to the mass analyser

HOW DOES APGC WORK ?

Corona discharge needle

Charge Transfer

“Dry” source conditions Better for relatively non-polar compounds

Proton Transfer

With water present as modifier Better for relatively polar compounds

Corona discharge needle

N2+●

N2e-

2e-

2N2

N4+● M+●

M

M+●

M

N2+●

N4+●

H2O

H2O+●

H2O

H3O+●

+OH●

MH+

M

APGC IONIZATION PROCESSES

BROMINATED FLAME RETARDENTS PESTICIDES IN DRIED TEA The pesticide residue analysis presented below was performed using APGC interfaced to a Xevo TQ-S tandem quadrupole mass spectrometer operated in MRM mode. The proton transfer process was utilized for ionization. After QuEChERS extraction, an aliquot was subjected to an SPE cartridge cleanup (below left, Sep-Pak PSA/carbon). The cleanup obtained is shown below right.

DIOXINS

Name RT MRM 1 MRM 2BDE 28 6.86 409.8 > 169.1 409.8 > 328.9BDE 66 8.09 486.71 > 247.9 486.71 > 407.8BDE 47 7.88 486.71 > 247.95 486.71 > 407.8BDE 85 8.61 563.6 > 403.7 563.6 > 405.7BDE 99 8.85 563.6 > 403.7 563.6 > 405.7BDE 100 9.17 563.6 > 403.7 563.6 > 405.7Octa BDE 10.82 643.5 > 483.6 643.5 > 485.7BDE 153 9.46 644.5 > 565.5 644.5 > 405.6BDE 154 9.75 644.5 > 565.5 644.5 > 405.6

Mass SpectrometryInstrument: Waters Xevo TQ-SMode: API positive (APGC)Corona: 2.2 µA Source Temperature: 150o C Probe Temperature: 450o CCone Gas: 170 L/hrAux Gas: 250 L/hrNebulizer Gas: 4.0 BarCollison Gas (Argon): 0.18 mL/min

Gas ChromatographyInstrument: Agilent 7890 Column: J&W DB% MS 30 m x 0.25 mm x 0.25 µm Injection Volume: 2 µL splitlessFlowrate: 2.0 mL/min helium (constant flow) Temperature Program: 80°C initial, hold for 0.5 min, 12°C/ min to 300°C and hold for 10 min

Acephate 50 5.18 183.8>94.8 (10,20) 183.8>142.8 (10,10) 66 (9) 59 (9)Bifenthrin 100 15.97 242.8>122.9 (20,10) 242.8>154.9 (20,10) 91 (8) 71 (10)Bitertanol 100 17.63 337.9>98.8 (20,10) 337.9>268.9 (20,10) 86 (9) 78 (13)Carfentrazone 20 14.86 411.7>276 (20,30) 411.7>301.8 (20,30) 101 (17) 93 (11)Chlorpyrifos methyl 100 10.96 321.6>124.7 (35, 20) 321.6>289.6 (35,10) 63 (14) 76 (11)Chlorfenapyr 50000 14.05 408.7>270.8 (20,20) 408.7>378.7 (20,10) 98 (10) 93 (12)Cyfluthrin 100 18.81 433.7>126.8 (15,30) 433.7>190.8 (15,10) 101 (6) 91 (20)Cypermethrin 500 18.67 415.8>126.8(25,25) 415.8>190.8 (25,10) 78 (16) 89 (18)Dicofol 20000 16.06 352.6>281.7 (20,20) 352.6>316.6 (20,10) 65 (67) 88 (2)Diazinon 50 9.99 304.9>168.9 (20,20) 304.9>276.9 (20,10) 98 (16) 79 (9)Dichlorvos 20 5.17 220.8>108.9 (20, 10) 220.8>144.8 (20, 10) 87 (9) 77 (17)Deltamethrin 5000 20.42 505.6>252.7 (20,20) 505.6>280.7 (20,10) 63 (47) 88 (23)Ethion 3000 14.44 384.6>142.7 (10,20) 384.6>170.8 (10,10) 90 (10) 90 (13)Etoxazole 15000 16.13 359.9>140.8 (35,30) 359.9>303.8(35,20) 77 (14) 71 (11) Endosulfan 30000 13.17 406.5>252.6 (10,20) 406.5>288.6(10,10) 145 (7) 101 (18)Fenpropathrin 2000 16.11 349.9>96.8 (25,30) 349.9>124.8(25, 10) 94 (7) 81 (10) Fenvalerate 50 19.54 419.8>124.8 (10,40) 419.8>166.8(10,10) 72 (14) 86 (16)l-Cyhalothrin 1000 16.76 449.8>196.8 (15,20) 449.8>224.8(15,10) 61 (20) 40 (11) Malathion 500 11.67 330.8>126.8 (15, 10) 330.8>210.8(15,20) 57 (33) 97 (30)Monocrotophos 50 8.88 223.8>97.8 (25, 10) 223.8>126.8(25, 10) 85 (6) 74 (17)Propargite 5000 15.17 230.8>80.8 (20, 20) 230.8>162.8(20,10) 94 (21) 107 (28)Propetamphos 100 (US) 9.78 281.9>137.8 (10,20) 281.9>194.8(10,10) 101 (16) 88 (5)Pyriproxyfen 50 16.67 321.9>95.8 (10,20) 321.9>184.8(10,20) 89 (10) 80 (10)Phenothrin 50 16.45 350.9>182.8 (20,40) 350.9>248.8(20,20) 69 (22) 72 (7)Phosalone 50 16.62 367.7>124.8 (15,20) 367.7>181.8(15,20) 80 (8) 71 (11)Resmethrin 200 15.46 338.9>170.9 (25,10) 338.9>292.9(25,10) 36 (22) 53 (16)Trifluralin 50 8.76 335.9 > 235.8 (30,10) 335.9>251.8(30, 20) 96 (14) 124 (24)

% Recovery (n=6) Pesticide MRL ppb (EU) RT min MRM m/z (Cone V, Collision eV) @ 10, 100 ppb (% RSD)

COMPARISON EI VS APGC Fragmentation of Endosulfan

EI spectrum (NIST)

APGC spectrum

The most abundant molecular ion from the isotopic cluster was used as precursor ion for generation of MRMs for quantitation. The molecular-ion cluster is too weak in the EI spectrum for quantitative use

Dilute 1 mL of QuEChERS extract with 10 mL 3:1 acetone/toluene

Place 200 mg MgSO4 atop cartridge frit

Pass diluted extract through Sep-Pak SPE cartridge* and collect

*500 mg GCB/500 mg PSA

Rinse cartridge with 2 mL 3:1 acetone/toluene and collect

Evaporate to ~ 0.5 mL

Add 2 mL toluene

Evaporate to 0.5 mL

no SPE with SPE

SPE Cleanup for GC-MS

Results

2,2′,4,4′,5-pentabromodiphenyl ether (BDE 99) detected on the surface of a computer keyboard

Dioxin analysis in food and the environment has required the use of GC with high resolution mass-spectrometry (GC-HRMS). This is traditionally performed using expensive magnetic-sector instruments. However, the recent introduction of tandem GC-MS systems, such as the APGC-MS/MS system used in this poster presentation, allows for sensitive and selective data equivalent to GC-HRMS at much lower cost. A recent EU directive addresses this issue: “In addition to the gas chromatography/high resolution mass spectrometry (GC-HRMS), technical progress and developments have shown that also gas chromatography/tandem mass spectrometry (GC-MS/MS) can be used as a confirmatory method for checking compliance with the maximum level (ML). Regulation (EU) No 252/2012 should therefore be replaced by a new Regulation providing for the use of gas chromatography/tandem mass spectrometry (GC-MS/MS) as an appropriate confirmatory method for checking compliance with the maximum level” (EU COMMISSION REGULATION No 589/2014).

1368-TCDD

1379-TCDD1379-TCDD

1368-TCDD

1378-TCDD1378-TCDD

1478-TCDD

1478-TCDD

1234-TCDD

1234-TCDD

2378-TCDD

2378-TCDD

GC-HRMS GC-MS/MS

The chromatograms shown at right compare GC-HRMS with APGC-MS/MS for analysis of a 2 pg/mL calibration standard (2 fg on column)

PBDE detected on a computer keyboard This PBDE (polybrominated diphenyl ether) analysis presented below was performed using APGC interfaced to a Xevo TQ-S tandem quadrupole mass spectrometer operated in MRM mode. The MRM transitions chosen for this analysis are shown in the table below.

PBDEs in whale blubber The PBDE analysis shown below was obtained with APGC using the SYNAPT G2-Si mass spectrometer in Tof mode. The total ion-current chromatogram is shown in grey; selected-ion chromatograms are superimposed for tri-, tetra-, penta– and hexabromo diphenyl ether congeners detected in the sample.

Time2.50 5.00 7.50 10.00 12.50 15.00 17.50 20.00 22.50 25.00 27.50 30.00 32.50 35.00 37.50

%

0

100

01312014_DIOXIN_016 Sm (Mn, 1x2) 1: TOF MS AP+ 403.8 0.0500Da

9.33e4

1.7 ppm

2.3 ppm0.4 ppm

1.4 ppm

CONCLUSIONS

Theoretical Spectrum

Spectrum from blubber sample extracted ion chromatogram

C12H7Br3O Theoretical Spectrum

Spectrum from blubber sample extracted ion chromatogram

C12H6Br4O

Spectra obtained from APGC-MS/Tof; note that congeners identified by retention time or by characteristic polybrominated ion clusters are confirmed by exact mass match with theoretical spectra

APGC-MS/MS system with Waters Xevo TQ-S mass spectrometer. Tandem quadrupole mass-spectrometry offers the highest sensitivity for targeted analysis of known compounds. Note the Acquity UPLC I-Class system on the left; conversion between UPLC and APGC is a matter of minutes without venting the MS.

APGC-MS/MS system with Waters Synapt G2-Si mass spectrometer for APGC-Tof/MS and APGC-QTof/MS with target enhancement (Tof-MRM). Tof mass-spectrometry offers the highest degree of qualitative information for identification of unknowns and very good sensitivity. Using the Tof-MRM mode, sensitivities approaching tandem MS/MS can be achieved.

APGC is a robust and sensitive technique well suited for the identification and determination of a wide variety of compounds of emerging concern

With APGC (unlike EI) the molecular ion is typically well preserved resulting in higher sensitivity and selectivity

APGC-MS/MS offers a less expensive and well suited alternative to GC-HRMS (EI) for determination of dioxins and related compounds at ultra-trace levels

Xevo and Synapt instruments can be used for both LC and GC applications with a simple and rapid conversion