Evolution in the Sample Preparation for the Measurement of POPs in Biological Matrices
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Evolution in the Sample Preparation for the Measurement of POPs in Biological Matrices. J.-F. Focant. MS Laboratory Organic and Biological Anal. Chem. Human Exposure?. Non-occupational exposure (adults) > 90% due to food Invasive unpopular human biomonitoring

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Evolution in the Sample Preparation for the Measurement of POPs in Biological Matrices

J.-F. Focant

MS Laboratory

Organic and Biological Anal. Chem.


Human Exposure? POPs in Biological Matrices

  • Non-occupational exposure (adults)

  • > 90% due to food

  • Invasive unpopular human biomonitoring

    • analyses of foodstuffs (feed, …)

  • Analyticaly challenging…


  • A ‘Good’ Kick… POPs in Biological Matrices

    Coût : 400 106 € (0.21% PIB)

    1 gouvernement

    Time axis

    Focant and De Pauw, Les Dioxines. La Recherche323 (1999) 84.

    Cost : 1 government and 1-2 billions €…

    7 106 chickens & 50,000 pigs slaughtered


    EU Commission Strategy (food-feed) POPs in Biological Matrices

    • Maximum levels set as ALARA

    • MRL strategy(SANCO/11248/2011 & 11515/2011)

    • Uniform sampling and analysis criteria

    • Comprehensive & reliable data

      Robust tools needed…


    Analytical Criteria POPs in Biological Matrices

    • Commission Regulation 1883/2006

    • Commission Regulation 152/2009

    • EPA1613B

    • prEN16215 (feed)

      • PBMS (different modules)

      • Module B: Extraction

      • Module C: Clean-up

    Manual

    or

    automated


    Today’s Capacity POPs in Biological Matrices

    • D+1 Fat matrices (regular input)

    • D+3 Feed matrices (regular input)

    • D+5 All matrices (irregular input)


    Automated Solution POPs in Biological Matrices


    prEN16215 POPs in Biological MatricesModule B:

    Extraction


    Parallel PLE POPs in Biological Matrices

    Extraction,

    solventreduction and exchange


    Automated PLE POPs in Biological Matrices

    70-85 min.

    + 5-25 min wash time = 75-110 min cycle time


    Control software POPs in Biological Matrices


    Traceability of Events POPs in Biological Matrices

    P(psi)

    T(C)

    1800

    150

    1500

    125

    Heat & Pressurize Cool & Depressurize

    0

    0

    27 37 45

    Time

    (min.)

    Pressure: Chan. 1

    Chan. 2

    Chan. 3

    Temperature: Chan. 1

    Chan. 2

    Chan. 3


    1-6 POPs in Biological Matriceslines system


    Natural Feed QC POPs in Biological Matrices


    Online Solvent Reduction POPs in Biological Matrices

    • 6 lines system

    • N2 flow

    • T prog.

    • No water bath

    • Solvent exchange

    • Dustfilter in


    PLE Properties POPs in Biological Matrices

    • Stand-alone automated parallel system

    • Up to 6 samples at a time (modular)

    • 1.8h / 6 samples (cycle time)

    • Wide bore plumbing (no pump cavitation)

    • PLE is an ‘old’ and proven concept

    • Robust and versatile (various cell sizes)

    • Easily usable under QA/QC requirements

    • Good throughput / cost effectiveness ratio


    prEN16215 POPs in Biological MatricesModule C:

    Clean-up


    Parallel Clean-up and Fractionation POPs in Biological Matrices


    1) Hexane 90ml POPs in Biological Matrices

    2) 50:50 100ml

    Waste

    3) Hexane 10ml

    PCBs

    Waste

    4) Toluene

    Extract in hexane (20 ml)

    Silica

    Alumina

    PCDD/Fs & NO-PCBs

    Carbon


    Automated Clean-up POPs in Biological Matrices

    75 min.

    + 15 min wash time = 90 min cycle time


    Manual Smith POPs in Biological Matriceset al. setup


    1-6 POPs in Biological Matriceslines system


    Previous generation POPs in Biological Matrices

    Current generation


    Previous POPs in Biological Matrices

    generation

    Current

    generation

    Previous

    generation

    Current

    generation


    Tracability by Lot # POPs in Biological Matrices

    • Good stability

    • Good lot-to-lot reproducibility

    • ‘PCB-free’ , ‘PBDE-free’ columns…


    Natural Spray Dried Milk POPs in Biological Matrices

    Experiment

    CR values

    10.5±2.4

    26.2±0.4

    4.8±0.4

    BCR-607 (n=3)

    (PCDD/Fs + cPCBs in ppt, other PCBs in ppb)

    RSDs < 20% Recovery rates inside EU Directive range


    QC Serum Samples POPs in Biological Matrices

    (n=4)

    (PCDD/Fs + cPCBs in ppt, other PCBs in ppb)

    RSDs < 10% Recovery rates in the high 60% range


    Online Solvent Reduction POPs in Biological Matrices

    • 100 mL Hx/DCM to 500 µL

    • 80 mL toluene to 500 µL

    • Nitrogen flow

    • 30°C, heating metal block


    Sample Clean-up Properties POPs in Biological Matrices

    • Stand-alone automated parallel system

    • Up to 6 samples at a time (modular)

    • 1.5h / 6 samples (cycle time)

    • Disposable Si, Al, and C

    • 20 year old concept, many revisions since then

    • Robust and versatile (various column sizes)

    • Easily usable under QA/QC requirements

    • Good throughput / cost effectiveness ratio


    Global Solution POPs in Biological Matrices


    The Integrated Approach POPs in Biological Matrices

    13C spiking

    Solvent exchange

    Lipid

    Determination

    Keeper

    Several fractions

    PTV-LVI

    PLE

    Keep it open

    SR-1

    Clean-up & Fractionation

    SR-2

    GC-MS


    Time Scale POPs in Biological Matrices

    • Condition the clean-up system

    • Fill PLE cell

    • Heat and presurize

    • Cool, nitrogen purge to evaporator

    • Reduce volume (on the go)

    • Load on silica

    • Elute silica with hexane

    • Elute alumina with 50:50 (PCBs)

    • Back-elute carbon with toluene (D/Fs)

    25 min

    25 min

    per cycle

    30 min

    40

    min

    Total time : 2 to 3h per sets of 6 samples in parallel


    Same-Day Testing Capability POPs in Biological Matrices

    • Sample preparation for PLE 0.5 h

    • Parallel PLE (2 cycles) 1.5 h

    • First solvent reduction 0.5 h

    • Parallel Clean-up and fractionation 2 h

    • Second solvent reduction 1.5 h

    • Std addition and transfer to GC 0.1 h

    • PTVLVI-GC-IDHRMS 1 h

    • QA/QC check and reporting 0.5 h

    • TOTAL ~ 8 h


    The missing link (2009)... POPs in Biological Matrices

    PLE-Vap-PrepLC-GCxGC-PTVLVI-IDEIHRMS


    The missing link (2012)... POPs in Biological Matrices

    Std addition & injection on

    TriPlusTM Autosampler


    What Next for Tomorrow ? POPs in Biological Matrices

    • Dioxin testing is getting faster and more reliable

    • Confirmatory approach status changed…

    • Confirmatory not too far from screening…

    • EU strategy to be revisited ?

    • Focus on ‘better’ use of CALUX


    Not to Forget… POPs in Biological Matrices

    • Dioxin testing remains a task for chemists…

    • Keeping blank levels under control is crucial…

    • ‘Simplifying’ the MS side is one thing, keep

    • performing state-of-the-art sample preparation is

    • another one…

    • Fast and reliable does not mean trivial…


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