Get on the Analyzer “FAST TRACK”

Pesticide and Environmental Pollutant GC/MS/MS Multi-residue Analyzer 2.0

… Identify Analytes in Complex Matrices … Obtain Analytical Results on DAY ONE!

ASTS 2012

ASTS 2012

Representative Sample

Extraction

General Workflow: Screen Confirm and Quantify

Clean-up

QuEChERS

S C Q

C16 H19 N3 P Cl C15 H25 O P S Cl C18 H21 O P Cl

LC/QTOF or GC/QTOF – for unknown compounds

S

Exact Mass Database Search

Molecular Formula Generation

Q C Another injection for MS/MS (QQQ or QTOF) 3

1

LC/QQQ MRM – for known targets S

C

Q

3 1

GC/QQQ MRM – for known targets S

C

Q

3 1

GC/MS (PTV) SIM/Scan – for known and unknown Final Report

S

C

Q

Deconvolution

(+backflush)

2

1

QuEChERS Procedure

3/20/2012

2

*

Add homogenized sample to tube Add 2 ceramic homogenizers (see below)*

Add ACN, shake

Add extraction packet of salts

Shake, Shake, shake

Add Internal standards, shake

Centrifuge, partitioning and extraction

Transfer an aliquot to the d-SPE tube

Vortex

Centrifuge

Transfer aliquot to vial for analysis

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2 3

4

3

5 3

6

8

7

9

10 11

The Agilent Analyzer Concept

ASTS 2012

Factory Configuration and Chemical Test with Agilent Specialist

On-site Application Set-up

CE Setup HW I&F

Customer Responsible

Aftermarket

Factory Checkout

Individual Components produced

Individual Components Tested (OFN)

System assembled

Appl. method developed on system

System checkout on appl. mix

Run 1st customer sample

Method optimization or customization

On-going maintenance & service support

Factory Setup & Checkout

Consulting or Field Services

Aftersales Support

CE Specialist Analyzer I&F

Hardware Only Sale

Analyzer Sale

Analyzer Added Value

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Technical Features: Pesticide and Environmental Pollutant GC/MS/MS Analyzer 2.0

• Allow simple system-to-system method transfer and column replacement

Retention Time Locking

• Large-volume injection enhances trace-level detection Multimode Inlet (MMI)

• Improve uptime and shorten analysis times Capillary Flow

Technology (CFT) and Backflush

• Comprehensive and optimized MRM database for 1000+ compounds

Comprehensive MS/MS MRM Database

• Factory setup and check out on pesticide mixture Pre-config. and factory

setup / tested

New

ASTS 2012 5

RTL GC-MS (/MS) Analysis

ASTS 2012

Load method, single injection to re-lock RT on customer site, and acquisition method is set-up!

Trifluralin Expected RT = 5.8185 min

System at Agilent Lab

System at Customer Lab after re-lock

5.8177 min

5.8176 min

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Why Use Backflush?

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Increased Productivity:

• Selectivity of MS/MS eliminates “dirty matrix”

• LOD requirements necessitates users inject larger sample volumes

• Many late eluting peaks are not “chromatographically ideal” and leave a

residue throughout the column

• Heavy matrices contaminate the source faster – performance is LOST!

Backflush ensures the highest data quality with GC/MS/MS! The trade-off is 10-20% decrease in sensitivity.

ASTS 2012

Milk Extract Analysis

5 10 15 20 25 30 35 40 45 50 55 60 65 70

Run stopped at 42 min and back flushed at 280oC for 7 min

It took additional 33 min and column to 320oC to remove these high boilers.

Blank run after back flushing

min

Column is clean.

Once set, backflush timing will work for ALL high boilers. - heaviest compounds elute first

8 ASTS 2012

Without Backflush: A Serious Problem

Overlay of two chromatograms of a blank extract injected BEFORE (A) and AFTER (B) three injections without back flush

Abundance

4.00 6.00 8.00 10.00 12.00 14.00 16.00 18.00 20.00 0 2000000 4000000 6000000 8000000 1e+07

1.2e+07 1.4e+07 1.6e+07 1.8e+07 2e+07 2.2e+07 2.4e+07 2.6e+07 2.8e+07 3e+07 3.2e+07 3.4e+07 3.6e+07 3.8e+07 4e+07 4.2e+07 4.4e+07 4.6e+07

Time

A: TIC: lettuce_blank.D\data.ms

B: TIC: lettuce_blank3.D\data.ms

B

A

Data provided by user in Almeria, Spain

After 3 sample injections, background significantly higher with increased chemical noise in every spectrum

ASTS 2012 9

With Backflush: Less Spectral Noise (Background) and Consistent Retention Times

Stable retention times and baseline with reduced chemical noise

4.00 6.00 8.00 10.00 12.00 14.00 16.00 18.00 20.00 0

2000000 4000000 6000000 8000000 1e+07

1.2e+07 1.4e+07 1.6e+07 1.8e+07 2e+07

2.2e+07 2.4e+07 2.6e+07 2.8e+07 3e+07

3.2e+07 3.4e+07 3.6e+07 3.8e+07 4e+07

4.2e+07 4.4e+07 4.6e+07

Time

Abundance

TIC: lettuce_10_ppb.D\data.ms TIC: lettuce_100_ppb.D\data.ms TIC: lettuce_5_ppb.D\data.ms

Overlay of three chromatograms of lettuce extract run with 2 min of back flush

Data provided by user in Almeria, Spain

ASTS 2012 10

But… How will this Analyzer work with

my list of target pesticides?

11 ASTS 2012

Agilent Provides the Most Comprehensive and Flexible MRM Database

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1. Contains >8500 optimized MRMs for 1074 pesticides and environmental pollutants

2. Characteristics for each compound include CAS, MW, Molecular Formula, RT in CF and CP BF modes, etc. Relative intensity of each transition

3. Tools and macros for quick method customization Requires <5 Min. to build acquisition method Tutorial videos on database CD for education and training

ASTS 2012

Important!

Classes of Compounds in the Database

Compound Classification Total Number Pesticides (fungicides, herbicides, insecticides, rodenticides and others) 675 Breakdown Products 42 Deuterated Compounds 6 Polybrominated Diphenyl Ether (PBDE) 4 Polybrominated Biphenyl (PBB) 1 Polychlorinated Biphenyl (PCB) 209 Polycyclic Aromatic Hydrocarbon (PAH) 26 Phthalates 17 Additional Semi-volatile Pollutants 94

13 ASTS 2012

Why Do You Need 8 Transitions for Each Compound? -- Top Two Transitions of Methamidophos (10 pg) in Three Matrices

small matrix interference

Spinach

strong matrix interference (overlapping peak gives

inaccurate quantitation result)

Pear Orange

medium matrix interference (overlapping peak gives

incorrect ion ratio)

141 -> 95

95 -> 79

Alternative Transitions Help Minimize Matrix Interferences and Improve Quantitation Result

14 ASTS 2012

Successful Use of Two Alternate Methamidophos Transitions to Minimize Matrix Interference

95 -> 64

141 -> 80

Pear Orange

minimum matrix interference minimum matrix interference

Recommendation: Evaluate chemical noise of an analyte’s multiple transitions in different matrices to select the most appropriate transitions in a particular matrix.

Database’s feature of 8 MRM transitions for each analyte makes this task easy!

15 ASTS 2012

16 ASTS 2012

Pesticides Analysis Case Studies

Case Study 1

Background: A major Chinese independent lab required a single injection GC/MS/MS method to analyze 323 pesticides in vegetables

Instrument Time for method

development Chemicals Required End Result

Other Vendor’s System 32 weeks of hard work 323 chemical

standards Poor results, not happy,

switched to Agilent

Agilent Pesticide 1.0 2 weeks, to add cmpds and adjust method

34 compounds not in 0501 Database Happy!

Agilent Pesticide 2.0 3 minutes

(NOT weeks, NOT days, NOT hours)

None Love it !!!

To build the database Agilent performed >3000 runs, so you don’t have to!

17 ASTS 2012

18 ASTS 2012

Case Study 2: Endosulfan Sulfate (?) in Organic Tea Extract

3.3 ppb, Equiv : 0.016 mg/Kg in Tea Tea EU MRL : Sum Endo a+b+sulfate = 30 mg/kg

Std 5 ppb Reagent Blank Organic Tea A

• Single Qualifier Transition • Ion Ratio outside 80-120%

confidence band • Unconfirmed

19 ASTS 2012

Case Study 2: Optimized MRM Transitions for Endosulfan Sulfate

• Alter method to include next highest transitions

• Rerun analysis

20 ASTS 2012

Endosulfan Sulfate () in Organic Tea Extract – Using 3 Qualifier Transitions

• Rerun of data • Ion Ratio within 80-120%

confidence band • CONFIRMED!!

Organic Tea A • Demonstrates the value of multiple optimized transitions in the MRM Database.

• Not just to avoid matrix interferences but also for additional confirmation!

Analyzer Benefits

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1. Reduces end user investment for method development and start-up

2. Deployment of cutting edge Agilent technology such as RTL, CFT and MRM Database increases productivity

3. Ongoing support for hardware PLUS application performance

Agilent Analyzers offers laboratories the quickest return on their capital investment!

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Thank you for your attention!