Screening for Hundreds of Pesticides in Fruits and ... · Screening for Hundreds of Pesticides in...

Screening for Hundreds of Pesticides in Fruits and Vegetables using the Agilent 7200 GC/Q-TOF with a new Exact Mass Pesticide Library

Philip L. Wylie, Ph.D.

Agilent Technologies

Wilmington, DE

Phil_wylie@agilent.com

March 2, 2015

ASMS

1

7200 Quadrupole Time of Flight

What Can the GC/Q-TOF Do?

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TOF mode (full spectrum acquisition) • High resolution full spectral acquisition • Accurate mass measurements • Fast acquisition of full spectra

Q-TOF mode (MS/MS) • Full spectrum of Product Ions (SIM-TOF)

- With high resolution and accurate mass

• Selectivity beyond any “triple quad” MRM method - Confirm TOF spectrum without interferences

• High sensitivity structural elucidation tool

Ideal tool for solving complex analytical problems

7200B Performance Specifications

Resolving Power: >12.5K at m/z 272 • 13K to 15K typical

Mass Accuracy: <3 ppm for 7200B • <2 ppm typical

MS Sensitivity <250 fg of OFN derived statistically

Dynamic Range: > 3 orders of magnitude • 4 orders with Dual Gain

TOF Mass range: 20-3000 u

MS/MS Mass range: 20-1050 u

Spectral Rate: 1-50 Spectra/sec

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Internal Reference Mass (IRM) Compound

1,3,5-Triazine, 2,4,6-tris(pentafluoroethyl)-, [C9F15N3]*+ = 434.9847

Accurate mass EI spectrum [M]+ = 434.9847 Measured mass Accurate Mass Mass Accuracy ppm

434.9839 434.9847 -1.8391

434.9839

Tune TOF Mass Axis as Part of a Sequence

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In sequence table: - Choose “Keyword” for Type - Choose “MassCal” for Keyword Mass calibration takes only 90 seconds

I usually do not use the Internal Reference Mass because it is not needed.

Camphor Accurate Mass – Internal Reference Mass (IRM) Turned Off

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Measured M*+ =

152.1197

Calculated M*+ = 152.1196 Difference = 0.88 ppm

Where is the GC/Q-TOF being used?

Natural Products

Metabolomics

Food Safety and Testing

Flavor and Fragrances

Forensics and Toxicology

Environmental

Energy and Petrochemical

Screening Pesticides with a GC-Q/TOF

• Accurate Mass provides added compound selectivity - Higher resolution provides added interference selectivity • TOF mode : Always full spectral data, Non-targeted acquisition • ‘Unlimited’ number of compounds can be screened (search

accurate mass compound database for identification) • Sensitivity is the same regardless of number of compounds

screened • MS/MS (Q/TOF Mode) further assists compound

identification

Pesticide screening GC-Q/TOF

About 300 pesticides listed with names beginning with “C”

Enhanced Selectivity using Accurate Mass

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Full Scale

200,000,000

200,000

5000

230.9737 +/- 10 ppm

383.9871 +/- 10 ppm

Ethion @ 5 ppb in Ginseng Extract Extracted Ions = 230.9737 and 383.9871 +/- 10 ppm

PCDL : Personal Compound Database & Library with ~750 Pesticides

• Name • Mol. Formula • Mono-isotopic exact mass • Locked RT • CAS # • Exact mass spectrum

All Ions data processing - Method setup (1) Formula Source

Pesticide screening GC-Q/TOF

• All Ions always uses RT

• Specify PCDL

All Ions data processing - Method setup (2) Formula matching

Pesticide screening GC-Q/TOF

• Set mass window for fragment EICs

All Ions data processing - Method setup (3) Fragment Identification

Pesticide screening GC-Q/TOF

• Select minimum number of qualified fragments required to identify compound

• Co-elution score is the major function to qualify compounds based on similarity of extracted ion peak shapes & RTs

• Select number of fragment ions to be evaluated

Vinclozolin: 10 ppb in Catfish Extract

Vinclozolin: 10 ppb in Catfish Extract

Target Compound List And Results

1 2

3 4

5

Compound Identification Results

Extracted Ion Chromatograms

Coelution Plot

Molecular Ion / Isotope Ratio Results

Spectrum & Extracted Ion Plots

Vinclozolin 10 ppb: EICs and Coelution Plot

Fenpropathrin in Catfish: Is it really there?

Fenpropathrin in Catfish Extract: M+* Isotope Ratios are not Perfect

Fenpropathrin in Catfish Extract: Retention Time and Mass Accuracy are Very Good

Δ RT = 0.008 min Mass Accuracy = 2.45 ppm

Hexachlorobenzene 10 ppb in Catfish Extract

Kadethrin: Review shows that this is a false Positive

Library Spectrum

Analysis of spiked fruits and vegetables Use organically grown produce EN QuEChERS extraction 2 spiking levels – 10 ppb and 100 ppb Compare two different GC methods – 20 min vs 40 min Optimize “All Ions” setpoints See how many pesticides can be found Measure TOF mass accuracy

March 2, 2015

ASMS

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Backflush, RTL GC Methods Employed 40 Minute

Constant Flow Method

20 Minute Constant

Flow Method

Columns 2 x 15m 5m + 15m

Back flush type Mid-point ‘Mid’-point

# Spiked Pesticides in PCDL

100 108

Inlet

MS

Aux

Column #1 15m

Column #2 15m

PCT

Mid-point Back flush Inlet

MS

Aux

Column #1 5m

Column #2 15m

PCT

‘Mid’-point Back flush

Matrix

Ave.

% Identified

(5X15) Of 108 Pest.

95.4 86.1 90.7 96.3 90.7 93.5 92.1

% Identified

(15X15) of 100 Pest.

96 91 94 98 89 94 93.7

Pesticides Identified at 10 ng/mL Spiking Level

Matrix

Ave.

% Identified

(5X15) Of 108 Pest.

100 100 96.3 99.1 98.1 96.3 98.3

% Identified

(15X15) of 100 Pest.

100 99 97 100 100 98 99

Pesticides Identified at 100 ng/mL Spiking Level

Mass Accuracy in Matrix– A function of S/N

Measured S/N Average difference between Exact Mass and Measured Mass (ppm)

< 10 6.7

> 10 < 100 1.8

> 100 1.2

Broccoli extract spiked at 100 ng/mL

Pesticide Screening by GC/Q-TOF Summary

Pesticide screening GC-Q/TOF

• Continuing trend towards comprehensive pesticide screening methods using accurate mass instruments

• Innovative data mining tools for identification of trace contaminants in complex matrices

• All Ions integrated workflow for enhanced qualification of suspected compounds by additional fragment ions in qualitative screening

- Similar Workflow as on LC- QTOF • Easy customizable and generation of user PCDL Libraries • Personal compound database library contains > 700 pesticides

with GC/MS accurate mass MS spectra and locked retention times

• Seamless integration of compound identification and MH Quantitation for easy and quick batch review