Rapid Analysis of Perfluorinated Compounds Using Triple Quadrupole LC/MS/MS Brahm Prakash Tairo Ogura and William Lipps Shimadzu Scientific Instruments, Inc., Columbia MD Promise not to call on anyone. “How many can differentiate between chromatographic resolution and mass resolution?” “How many here are very comfortable with their understanding of MRM, or an MRM transition on an LCMS TQ?” “How many here understand why a TQ is more sensitive than a SQ even though ion intensity goes down?” “How many would enjoy a debate on whether ESI is solution phase chemistry or gas phase chemistry?” “How many have extensive TQ operation experience?” “How many have run PFCs on a TQ?”

EPA Method 537 is an older method, uses SPE and does not take advantage of the power of modern LCMSMS

ASTM D7979 Extraction procedure eliminates SPE step 5 ml Sample Surrogate 5 ml MeOH EPA Method 537 is a solid phase extraction method. In this method you collect sample in a 250 milliliter bottle. Weigh the bottle. Add surrogate to the bottle then pass through the SPE cartridge at 10 milliliters per minute (This will take 25 minutes at minimum). Then rinse the bottle with reagent water and pass that through the SPE cartridge. Then add 4 milliliters of methanol to the sample bottle, rinse the sides, and pass the methanol dropwise through the SPE cartridge. This will take about one hour. Place the extract in a 60 C bath and evaporate (under nitrogen) the methanol extract to dryness. Add 1 milliliter of 96% methanol in reagent water. Add internal standard and analyze. Reweigh the bottle empty to calculate sample volume.

ASTM D7979 Extraction procedure LCMS-8060 10 µL Acetic Acid EPA Method 537 is a solid phase extraction method. In this method you collect sample in a 250 milliliter bottle. Weigh the bottle. Add surrogate to the bottle then pass through the SPE cartridge at 10 milliliters per minute (This will take 25 minutes at minimum). Then rinse the bottle with reagent water and pass that through the SPE cartridge. Then add 4 milliliters of methanol to the sample bottle, rinse the sides, and pass the methanol dropwise through the SPE cartridge. This will take about one hour. Place the extract in a 60 C bath and evaporate (under nitrogen) the methanol extract to dryness. Add 1 milliliter of 96% methanol in reagent water. Add internal standard and analyze. Reweigh the bottle empty to calculate sample volume.

In Water, Sludge, Influent, Effluent and Wastewater Method Validation: Direct Inject Method ASTM D7979 - Standard Test Method for Determination of Perfluorinated Compounds by LCMS – TQ8060 In Water, Sludge, Influent, Effluent and Wastewater 500ng/L, high calibration point (XRODS-3x50x2.4_PhenylHexyl 2.1x100x3 u) Chromatogram Obtained under optimized conditions – only half of the target compounds were spiked, awaiting other standards to arrive. Chromatogram Obtained under optimized conditions

Glass Vial Poly Propylene Standard Stability Study GLASS Vials Vs Poly Propylene Vials- 10% Methanol 90% water Glass Vial Poly Propylene

Poly Propylene Vials Glass Vials Standard Stability Study GLASS Vials Vs Poly Propylene Vials- 30% Methanol 70% water Poly Propylene Vials Glass Vials

Glass Vials Poly Propylene Vials Standard Stability Study GLASS Vials Vs Poly Propylene Vials- 50% Methanol 50% water Glass Vials Poly Propylene Vials

Glass Vials Poly Propylene Vials Standard Stability Study GLASS Vials Vs Poly Propylene Vials- 70% Methanol 30% water Glass Vials Poly Propylene Vials

Glass Vials Poly Propylene Vials Standard Stability Study GLASS Vials Vs Poly Propylene Vials - 90% Methanol 10% water Glass Vials Poly Propylene Vials

Sample Vials - Vortex Effect PFCs Could Float on the top of Sample Vials and be not injected… resulting in poor recoveries or could go as non detect. Same Vial After Vortex Before Vortex

Optimization of Interface Temperature, Desolvation line (DL) Temperature and heat Block Temperature IF100_DL150_HB250 oC

Optimization of Interface Temperature, Desolvation line (DL) Temperature and heat Block Temperature IF200_DL150_HB250 oC

Optimization of Interface Temperature, Desolvation line (DL) Temperature and heat Block Temperature IF300_DL150_HB250

IF300_DL100_HB200 : Optimized Conditions Optimization of Interface Temperature, Desolvation line (DL) Temperature and heat Block Temperature IF300_DL100_HB200 : Optimized Conditions

Optimization of Interface Temperature, Desolvation line (DL) Temperature and heat Block Temperature IF400_DL300_HB400

Target List Including Newly Added by Office of Superfund Remediation and Technology Innovation (OSRTI)

In Water, Sludge, Influent, Effluent and Wastewater Direct Inject Method ASTM D7979 - Standard Test Method for Determination of Perfluorinated Compounds by LCMS – TQ8060 : Chromatogram at 100 ppt Using Optimized Conditions In Water, Sludge, Influent, Effluent and Wastewater 100ng/L, high calibration point (XRODS-3x50x2.4_PhenylHexyl 2.1x100x3 u)

Direct Inject Method ASTM D7979 - Standard Test Method for Determination of Perfluorinated Compounds by LCMS – TQ8060 : Chromatogram at 1.0 ppt Using Optimized Conditions 1.0 ng/L, high calibration point (XRODS-3x50x2.4_PhenylHexyl 2.1x100x3 u)

ASTM D7979 by 10 uL Direct Injection at 40 ppt Using LCMS– TQ8060 MFPBA at 40 ppt Level Concentration 1 5 ppt 2 10 ppt 3 20 ppt 4 40 ppt 5 60 ppt 6 80 ppt 7 100 ppt 8 150 ppt 9 200 ppt 0.2 0.5 1.0 2.0 5.0 10.0 25.0 50.0 75.0 100.0 Calibration Curves Calibration curves were produced in the range of 5 ng/L to 200 ng/L for the PFCs. All curves had a regression coefficient higher than 0.9900. Curves for a selection of target compounds are plotted

ASTM D7979 by 10 uL Direct Injection at 40 ppt Using LCMS– TQ8060 PFPeS at 40 ppt Level Concentration 1 5 ppt 2 10 ppt 3 20 ppt 4 40 ppt 5 60 ppt 6 80 ppt 7 100 ppt 8 150 ppt 9 200 ppt 0.2 0.5 1.0 2.0 5.0 10.0 25.0 50.0 75.0 100.0 Calibration Curves Calibration curves were produced in the range of 5 ng/L to 200 ng/L for the PFCs. All curves had a regression coefficient higher than 0.9900. Curves for a selection of target compounds are plotted

ASTM D7979 by 10 uL Direct Injection at 40 ppt Using LCMS– TQ8060 PFNA at 40 ppt Level Concentration 1 5 ppt 2 10 ppt 3 20 ppt 4 40 ppt 5 60 ppt 6 80 ppt 7 100 ppt 8 150 ppt 9 200 ppt 0.2 0.5 1.0 2.0 5.0 10.0 25.0 50.0 75.0 100.0 Calibration Curves Calibration curves were produced in the range of 5 ng/L to 200 ng/L for the PFCs. All curves had a regression coefficient higher than 0.9900. Curves for a selection of target compounds are plotted

ASTM D7979 by 10 uL Direct Injection at 40 ppt Using LCMS– TQ8060 PFTRE at 40 ppt Level Concentration 1 5 ppt 2 10 ppt 3 20 ppt 4 40 ppt 5 60 ppt 6 80 ppt 7 100 ppt 8 150 ppt 9 200 ppt 0.2 0.5 1.0 2.0 5.0 10.0 25.0 50.0 75.0 100.0 Calibration Curves Calibration curves were produced in the range of 5 ng/L to 200 ng/L for the PFCs. All curves had a regression coefficient higher than 0.9900. Curves for a selection of target compounds are plotted

ASTM D7979 by 10 uL Direct Injection at 40 ppt Using LCMS– TQ8060 PFTreA at 40 ppt Level Concentration 1 5 ppt 2 10 ppt 3 20 ppt 4 40 ppt 5 60 ppt 6 80 ppt 7 100 ppt 8 150 ppt 9 200 ppt 0.2 0.5 1.0 2.0 5.0 10.0 25.0 50.0 75.0 100.0 Calibration Curves Calibration curves were produced in the range of 5 ng/L to 200 ng/L for the PFCs. All curves had a regression coefficient higher than 0.9900. Curves for a selection of target compounds are plotted

MRL’s and MDL’s for ASTM D7979- Shimadzu LCMS-TQ8060 Minimum Reporting Level (ng/L) n=8 Compound Ret. Time Fortified Conc. (ng/L) MDL (ng/L) Calibration Range (ng/L) PFBA 5 4.1 5 - 200 MPFBA 5.0 PFPeA 0.9 M5PFPeA 0.6 4-2 FTS 1.7 M4-2 FTS 1.2 PFHxA 1.3 M4PFHxA 1.1 PFBS 1.5 M3PFBS FHUEA 2.6 FHEA 100 32.5 100 - 4000 PFHpA 1.4 M4PFHpA 0.7 PFPeS 6-2 FTS 2.5 M6-2 FTS 1.8 PFOA 5.1 M8PFOA

MRL’s and MDL’s for ASTM D7979- LCMS-TQ8060 Minimum Reporting Level (ng/L) n=8 Compound Ret. Time Fortified Conc. (ng/L) MDL (ng/L) Calibration Range (ng/L) FhpPA 5 9.4 5 -200 FOEA 100 48.3 100-4000 FOUEA 1.6 5 - 200 PFHxS 1.5 M3PFHxS 1.7 PFNA M9PFNA 8-2 FTS 3.2 M8-2 FTS 1.8 PFHpS N-MeFOSAA 3.6 MN-MeFOSAA 5.4 PFDA 2.3 M6PFDA 1.1 FDEA 35.5 100 - 4000 N-EtFOSAA 5.3 MN-EtFOSAA 4.2 PFOS 3.0 M8PFOS

MRL’s and MDL’s for ASTM D7979- LCMS-TQ8060 Minimum Reporting Level (ng/L) n=8 Compound Ret. Time Fortified Conc. (ng/L) MDL (ng/L) Calibration Range (ng/L) PFUnA 5 2.9 5 - 200 M7PFUnA 1.5 PFNS 1.3 PFDoA 2.2 MPFDoA 0.8 FOSA 0.6 M8FOSA 1.6 PFDS 2.1 PFTriA 1.1 PFTreA M2PFTreA 0.7

Method Detection Limit and Reporting Ranges ASTM Vs Shimadzu Shimadzu LCMSMS-TQ8060 Analyte MDL(ng/L) Ranges MDL(ng/L) Ranges (ng/L PFTreA 1.2 10 – 400 1.1 5–200 PFTriA 0.7 10 – 400 1.1 5-200 PFDoA 1.2 10 – 400 2.1 5-200 PFUnA 1.2 10- 400 2.9 5-200 PFDA 1.4 10- 400 2.3 5-200 PFDS 2.2 10 – 400 2.0 5-200 PFOS 2.2 10 – 400 3.0 5-200 PFNA 1.1 10 – 400 1.5 5-200 PFNS 1.4 10 – 400 1.2 5-200 PFOA 1.7 10 – 400 5.1 5-200 PFHpS 2.5 10 – 400 1.6 5-200

Method Detection Limit and Reporting Ranges ASTM Vs Shimadzu Shimadzu LCMSMS-TQ8060 Analyte MDL(ng/L) Ranges (ng/L) MDL (ng/L) Ranges (ng/L) PFHxS 1.2 10 – 400 1.4 5-200 PFHpA 1.0 10 – 400 1.3 5-200 PFHxA 2.0 10-400 1.3 5-200 PFBS 0.8 10 – 400 1.4 5-200 PFPeS 1.3 10 – 400 1.1 5-200 PFPeA 4.6 10 – 400 0.9 5-200 PFBA 0.8 50 - 2000 4.1 50-2000 FOSA 1.6 50 – 2000 0.6 50-2000 4:2 FTS 1.5 10 – 400 1.7 5-200 6:2 FTS 1.6 10 – 400 2.5 5-200 8:2 FTS 2.7 10 – 400 3.2 5-200

Method Detection Limit and Reporting Ranges ASTM Vs Shimadzu Shimadzu LCMSMS-TQ8060 Analyte MDL(ng/L) Ranges (ng/L) MDL (ng/L) Ranges (ng/L) FHEA 92.9 50 – 2000 32.5 50-2000 FOEA 106.8 50 – 2000 48.2 50-2000 FDEA 47.2 50 – 2000 35.4 50-2000 FOUEA 2.3 10 – 400 1.6 5-200 FPpPA 3.3 10 – 400 9.3 5-200 FHUEA 1.5 10 – 400 1.5 5-200

Comparison MDL Data EPA M537 Vs Shimadzu Results Obtained using LCMS-TQ8060 Compound EPA 537 Spike (ng/L) Sample Extract vol250 mL EPA M537 MDL (ng/L) LCMRL(ng/L) Shimadzu LCMS-8060 Spike (ng/L) LCMS-8060 DL (ng/L) PFBA 9.1 5 4.1 PFPeA 0.9 PFHxA 5.0 0.5 2.9 1.5 PFBS 1.6 3.7 1.4 PFHpA 3.1 3.8 1.3 PFOA 4.6 1.7 5.1 PFNA 4.8 0.7 5.5 PFDA 2.3 PFOS 9.6 6.5 3.0 PFUnA 5.4 2.8 6.9 PFDoA 1.1 3.5 2.1 PFTriA 2.2 PFTreA 4.7 Qwing to the high sensitivity of the LCMS-TQ8060 system these low ng/L levels were easily obtained for all compounds

Precision & Accuracy for ASTM D7979 Using LCMS-TQ8060 Precision & Accuracy at 10ng/L (n=8) Compound Mean Concentration (ng/L) % Recovery % RSD PFBA 16 164 10.5 MPFBA 11 111 9.0 PFPeA 10 104 4.7 M5PFPeA 101 3.1 4-2 FTS 103 3.4 M4-2 FTS 9 93 3.5 PFHxA 105 6.9 M4PFHxA 102 2.1 PFBS 9.6 M3PFBS 106 4.8 FHUEA 112 5.5 FHEA 211 PFHpA 7.5 M4PFHpA 2.7

Precision & Accuracy for ASTM D7979 Using Shimadzu LCMS-TQ8060 Precision & Accuracy at 20ng/L (n=8) Compound Mean Concentration (ng/L) % Recovery % RSD PFBA 22.4 112 6.6 MPFBA 17.3 86 10.2 PFPeA 20.1 101 2.9 M5PFPeA 20.0 100 1.4 4-2 FTS 20.3 102 3.2 M4-2 FTS 18.4 92 3.0 PFHxA 20.2 3.9 M4PFHxA 2.3 PFBS 10.4 M3PFBS 19.6 98 4.1 FHUEA 21.6 108 5.6 FHEA 397.5 99 5.3 PFHpA 20.5 M4PFHpA 19.7

Summary and Conclusions Under Optimized conditions-The Shimadzu LCMS-TQ8060 will exceed and meet performance criteria specified for EPA Method 537 Results demonstrated that high-sensitivity analysis with high repeatability is possible with Shimadzu’s triple quadrupole instruments The method presented here used an GL Science InertSustain Phenylhexyl 3um 2.1x100mm C/N 5020-89127 analytical column and a gradient that was designed to increase method throughput , while still providing sufficient chromatographic resolution. Lower MRLS for most PFCs is possible using advanced state-of-the art technologies Recommendation Due to high-sensitivity performance of the LCMS-TQ8060, it is recommended for use for methods allowing direct injections of PFCs, ASTM D7979

Study Continues …on Method Development for PFCs Thank You for attending… For questions and to know more details about the Method conditions please contact Brahm Prakash brprakash@shimadzu.com Phone:410-910-0903