Dioxin/PCB crisis in feed/food - BioDetection Systems

6 th BioDetectors Meeting 2012Dioxin/PCB crisis in feed/food –A decade of DR CALUX testingPeter A. BehnischBioDetection SystemsScience Park, Amsterdam1

Outline of Presentation• Current situation in Europe• German Dioxin Crisis 2011• Chile Food/Feed monitoring 2008• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Slovak Republic Food/Feed monitoring• Norway Fish monitoring school project• Taiwan feed and food monitoirng• BDS Client survey 2005• Other food monitoring programs using DR CALUX2

Latest dioxin crisis• 2010 – Maize from Ukraine (Nederlands)• 2011 – eggs (Belgium/Netherlands)• 2011 – eggs and milk (Italy)• 2011 – Biodiesel used as feed led to 25%polluted market eggs (Germany)• 2011 – Sugar molasses (Germany)• 2012 – Red colorants (The Netherlands)• 2012- Nearly weekly egg recalls in Germany by KAT3

New EC guidelines 252/2012, 277/2012 and 278/2012for screening for dioxins/PCBs by DR CALUX• Cell based screening tests can now report samples for dioxins anddioxin-like PCBs as compliant or suspected to be non-compliant(suspected)• New WHO-TEFs are more close to DR CALUX-REPs, lowering theamount of false negative for DR CALUX in case of high PCBcontaminated samples• False-compliant rate with respect to maximum levels is below 5%(before it was 1%)• Repeatability RSD below 20%• 20 confirmed results per matrix group demanded for evaluation of falsenegative rate• Suitable cut-off level at 2/3rd of maxiumum level4

New EC 277 and 278/2012 guidelines:Cut-off levels for DR CALUXMatrixOld Total-TEQOld cutoffNewDioxinNewTotal-TEQPoultry 4 1.5 1.75 3 1.2Bovine 4.5 2.25 2.5 4 1.7Fish 8 3 3.5 6.5 2.3Milk 6 2.25 2.5 5.5 1.7Eggs 6 2.25 2.5 5 1.7Plant oils 1.5 0.56 0.75 1.25 0.5New Cut-OffBabyfood no no 0.1 0.2 0.075

BDS DR CALUX vs HRGC/HRMS –170 comparisons: < 1% false negativ and < 8% false positiveMatrix N False negative False positiveegg 18 0 3fish 14 0 0Poultry meat 9 0 1Ruminant meat 6 0 0Pig meat 14 0 1fish meal 8 0 0Fish oil 15 0 1milk 33 0 3Feed, plant origin 18 0 4Feed, vegetable oil 7 0 16

Z-SCOREProficiency test performance BDS (2004-2011)21.5Orebro univeristy, round 1European Commission, Joint Research CentreBICS 2005Goeyens, Belgium10.50-0.5-10 5 10 15 20 25 30 35 40 45 50Norwegian Institute of PublicHealth in Oslo, NorwayOrebro univeristy,round 2-1.5-2Dutch National Institutefor Coastal and MarineManagementQuasimeme round 487

Acceptance by International Quality systems• MVO Code of dioxins: 100% of batches for coconut,thermic treated oils and oils direct used as animalfeed have to be tested by dioxins/PCBs• Private laboratories have also to report in case ofhigher dioxin/PCB levels• Increase quality product labels on EU market suchas IKB (eggs), QS (feed) German Meat AssociationVDF since Jan 2011, or GMP Plus8

Modernisation by High-Through Put Robot (HTPS)40 samples/hourdosing on 96 wellplates9

High-Through Put Robot (HTPS)40 samples/hourdioxin and separatePCB-TEQmeasurement10

Outline of Presentation• Compliance with EU regulations• German Dioxin Crisis 2011• Chile Food/Feed monitoring 2008• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Slovak Republic Food/Feed monitoring• Norway Fish monitoring school project• BDS Client survey 2005• Other food monitoring programs using DR CALUX11

German Dioxin Crisis 201112

Situation in January 2011• 4700 farms at first closed – issue taken up byinternational media• 70% of Germans doubted, if their breakfast eggs aredioxin-free• Ca. 70% lower revenues for farmers/industry – roughcalculations talked from 20 Mio Euro/week damage• Most local testing needed > 10 days = lack of localtesting capacity• Screening via DR CALUX offers time and cost saving• More than 95% of the ca 1800 samples fromGermany tested as express analysis in 2-3 days13

Situation in January/February 2011• More than 15 countries banned German meat• German Meat Association (VDF) in cooperation withGerman Authorities (BMELV) and the Russian ImportAuthorities accepted more than 40 labs for theGerman meat exports – only 4 non-Germanlaboratories• BDS only accepted non-German lab using screeningtechnology DR CALUX• Regarding acceptance of ISO 17025 accredited labsplease notice that according EC/764/2008 all reportsof such accredited labs have to be accepted in all ECmember states• BDS ISO 17025 accredited since 2005 and newapproved in mid 2011 until 201414

German pig meat > 97,6% negative;less than 2.5 % false positive by DR CALUXPCB-TEQPCDD/F-TEQTotal-TEQSamplecompliantPCDD/FSamplecompliant TOTALSamplecompliantPCDD/FSamplecompliantTOTAL- 25%- 25%- 50%- 50%PigMeat; NPigMeat;Mean496 496 502 99,8% 97,6 94% 89%0,23 0,21 0,43PigMeat;Range0,1-1,10,1-0,770,1-1,4716

PCDD/D/dl-PCB-TEQ byDR CALUXDR CALUX analysis for PCB + PCDD/F-TEQ (# 502 samples):German pig meat DistributionTotal-PCDD/F/dl-PCB-TEQ by DR CALUX for German pigmeat in Jan/Feb 2011 in pg PCDD/F/dl-PCB-TEQ/g fat1,601,401,201,000,800,600,400,200,00EC guided level 1.5 ng PCDD/F/PCB-TEQ/kg fat0 100 200 300 400 500 600Numbers of samples17

PCDD/F-TEQ by DR CALUXDR CALUX analysis for PCDD/Fs (# 496 samples):German pig meat DistributionDistribution PCDD/F-TEQ by DR CALUX for German pig meat in Jan/Feb 20110,80EC guided level 1.0 ng PCDD/F-TEQ/kg fat0,700,600,500,400,300,200,100,000 100 200 300 400 500 600Number of tested samples18

German pig liver > 97% negative;less than 3 % false positive by DR CALUXPCB-TEQPCDD/F-TEQTotal-TEQSamplecompliantPCDD/FSamplecompliantTOTALSamplecompliantPCDD/FSamplecompliantTOTAL- 25%- 25%- 50%- 50%Pig Liver;NPig Liver;Mean32 37 34 99% 97% 93% 90%0,80 0,78 1,57Pig Liver;Range0,1-2,10,1-3,40,1-5,219

German poultry meat > 98% negative;less than 2 % false positive by DR CALUXPCB-TEQPCDD/F-TEQTotal-TEQSamplecompliantPCDD/FSamplecompliantTOTALSamplecompliantPCDD/FSamplecompliantTOTAL- 25%- 25%- 50%- 50%ChickenMeat; NChickenMeat;Mean201 201 207 99,5% 98% 93% 88%0,74 0,56 1,3ChickenMeat;Range0,3-3,30,3 –1,70,6-3,820

PCDD/F/dl-PCB-TEQ byDR CALUX in ngTEQ/kg fatDR CALUX analysis for PCDD/F/PCB-TEQ (# 207 samples):German poultry meat DistributionPCDD/F/dl-PCB-TEQ by DR CALUX for Germanpoultry meat in Jan/Feb 2011EC guided level 4 ng PCDD/F/PCB-TEQ/kg fat432100 50 100 150 200 250Number of samples21

PCDD/F-TEQ by DRCALUX in ng TEQ/kgfatDR CALUX analysis for PCDD/F-TEQ (# 201 samples):German poultry meat DistributionPCDD/F-TEQ by DR CALUX of German poultry inJan/Feb 2011 (cut-off 1,4 pg PCDD/F-TEQ/gr fat)2EC guided level 2 ng PCDD/F-TEQ/kg fat1,510,500 50 100 150 200 250Number of samples22

Outline of Presentation• Compliance with EU regulations• German Dioxin Crisis 2011• Kuwait Monitoring of imported feed/food 2009• Chile Food/Feed monitoring 2008• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Slovak Republic Food/Feed monitoring• Norway Fish monitoring school project• BDS Client survey 200523

ng TEQ/kg productTotal DR CALUX ® distribution for FEED samples,2009 Asprocer Program, Chile21.81.60% above max EU PCDD/F-PCB limitmax EU limit PCDD/F-PCB = 1.5 ng TEQ/kg product1.41.210.80.60.40% above max EU PCDD/F limitmax EU limit PCDD/F = 0.75 ng TEQ/kg product75% max EU PCDD/F limit0.2Santiago, Chile, 22 March 201000 10 20 30 40 50 60 70 80 90 100# of analyses24

ng TEQ/kg productTotal DR CALUX ® distribution for MINERAL samples,2009 Asprocer Program, Chile338 ng TEQ/kg productmineral clay2.5zinc oxide21.55.4% above max EU PCDD/F-PCB limitmax EU limit PCDD/F-PCB = 1.5 ng TEQ/kg productzinc oxidezinc oxidemineral clayzinc oxide17.6% above max EU PCDD/F limitmax EU limit PCDD/F = 0.75 ng TEQ/kg productmineral clay0.575% max EU PCDD/F limit00 10 20 30 40 50 60 70 80 90 100# of analysesSantiago, Chile, 22 March 201025

ng TEQ/kg productTotal DR CALUX ® distribution for FISH OIL samples,2009 Asprocer Program, Chile25200% above max EU PCDD/F-PCB limitmax EU limit PCDD/F-PCB = 24 ng TEQ/kg product15100% above max EU PCDD/F limitmax EU limit PCDD/F = 6 ng TEQ/kg product575% max EU PCDD/F limit00 2 4 6 8 10# of analysesSantiago, Chile, 22 March 201026

ng TEQ/kg productTotal DR CALUX ® distribution for ANIMAL OIL samples,2009 Asprocer Program, Chile43.530% above max EU PCDD/F-PCB limitmax EU limit PCDD/F-PCB = 3 ng TEQ/kg product2.520% above max EU PCDD/F limitmax EU limit PCDD/F = 2 ng TEQ/kg product1.575% max EU PCDD/F limit10.500 2 4 6 8 10 12# of analysesSantiago, Chile, 22 March 201027

ng TEQ/kg productTotal DR CALUX ® distribution for VEGATABLE OIL samples,2009 Asprocer Program, Chile2.521.50% above max EU PCDD/F-PCB limitmax EU limit PCDD/F-PCB = 1.5 ng TEQ/kg product118% above max EU PCDD/F limitmax EU limit PCDD/F = 0.75 ng TEQ/kg product0.575% max EU PCDD/F limit00 5 10 15 20# of analysesSantiago, Chile, 22 March 201028

Outline of Presentation• Compliance with EU regulations• German Dioxin Crisis 2011• Chile Food/Feed monitoring 2008• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Slovak Republic Food/Feed monitoring• Norway Fish monitoring school project• BDS Client survey 2005• Other food monitoring programs using DR CALUX29

TEQ (pg/g fat)Dutch Dioxin crisis 2004:Clay effecting potato peeling used a animal feed.total DR CALUX TEQ distribution for milk samples, 2004 Dutch clay crisis1098761.3 % above 6 pg TEQ/g54325.8 % above 3 pg TEQ/g0 50 100 150 200 250 300 3501060 % below 1.5 pg TEQ/greported TEQ (sorted from lowest to highest)30

Outline of Presentation• International food/feed crisis• Compliance with EU regulations• Chemical TEQ vs. DR CALUX TEQ – a few examples• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Slovak Republic Food/Feed monitoring• Norway Fish monitoring school project• Other Monitoring Programs31

Australia 2006Soils Recycled Organics & Remediation Technologies Unit,Sydney, AustraliaDioxins: A fishy business;the analysis of dioxins in seafood from Sydney Harbour32

Sydney Harbour; preliminary results33

Time line rapid implementationWeektraining CALUXchain of custodyoptimalisation lab11 12 13 1415 16QA/QC data set1825 3545total TEQ testing12 2432 5070PCB TEQ testing1224325070evaluation accreditationfish samples done1224325070analiysis done42739914518534

Sydney harbour:Comparison of DR CALUX ® and HR-GCMS1286432DR CALUX ®(log scale)1684210.50.51248163264128HRGCMS(log scale)35

Outline of Presentation• International food/feed crisis• Compliance with EU regulations• Chemical TEQ vs. DR CALUX TEQ – a few examples• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Slovak Republic Food/Feed monitoring• Norway Fish monitoring school project• Other food monitoring programs using DR CALUX36

total DR CALUX TEQ/max EC total TEQ leveltotal DR CALUX TEQ/max EC total TEQ levelNational Monitoring Programfrom State Veterinary and Food Service of Slovak RepublicRatio:Total DR CALUX TEQ vs. accepted European Total-TEQ1.25Bovine Meat1.25Pork Meat1.001.000.750.750.500.500.250.250.000 2 4 6 8 100.000 2 4 6 8 10 12LocationLocationCALUX Exchange 2006 Presentation37

total DR CALUX TEQ/max EC total TEQ leveltotal DR CALUX TEQ/max EC total TEQ levelNational Monitoring Programfrom State Veterinary and Food Service of Slovak RepublicRatio:Total DR CALUX TEQ vs. accepted European Total-TEQ1.25Egg1.25Milk1.001.000.750.750.500.500.250.250.000 2 4 6 8 10 12 140.000 2 4 6 8 10 12LocationLocationCALUX Exchange 2006 Presentation38

total DR CALUX TEQ/max EC total TEQ leveltotal DR CALUX TEQ/max EC total TEQ levelNational Monitoring Programfrom State Veterinary and Food Service of Slovak RepublicRatio:Total DR CALUX TEQ vs. accepted European Total-TEQ1.25Fish1.25Poultry1.000.750.50Location 10:Ratio 3.4 due to formerPCB production1.000.750.500.250.250.000 5 10 15 200.000 5 10LocationLocationCALUX Exchange 2006 Presentation39

Outline of Presentation• International food/feed crisis• Compliance with EU regulations• Chemical TEQ vs. DR CALUX TEQ – a few examples• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Czech Food/Feed monitoring• Norway Fish monitoring school project• Taiwan Feed/Food Monitoring• Other food monitoring programs using DR CALUX40

School students workingas environmental scientists-”Global POP”Dioxins in fish with BDS CALUXEldbjørg Sofie Heimstad 1 , Gaute Grønstøl 2 , Karl TorsteinHetland 3 , Javier Martinez Alarcon 1 , Charlotta Rylander 1,4and Espen Mariussen 1,51Norwegian Institute for Air Research (NILU)2University of Bergen, Bergen.3Norwegian Centre for Science Education4Institute of Community Medicine, University of Tromsø5Norwegian Defence Research Establishment (FFI)41

Global POP – a school projectKids participate with field work anddata publishing in a research projectInvestigate dioxins and dioxin-likecompounds (CALUX) in fish commonfor consumptionSchools from Arctic areas will beencouraged to participate. TEQ will bedetermined by NILU in fish that werecaught by the studentsInvestigate dioxins and dioxin-likecompounds (CALUX) in fish commonfor consumption42

Norwegian Fish Monitoring Project; Polar Bear43

Salvelinus alpinusAnguilla anguillaGadus morhuaPleuronectes platessaSebastes marinusPollachius virensSalmo truttaPollachius pollachiusPerca fluviatilisCyprinus caprioCoregonus lavaretusPlatichtys flesusScomber scombusLota lotaMelanogramus aeglefinusMolva molvaLeuciscus cephalusEsox luciusRutilus rutilusBlicca bjoerknaLimanda limandaMugil cephalusSparus aurataAcanthopagrus australisAcanthopagrus butcheriWithout datasheetGlobal POP50454035302520151050NumberNumberGadus morhua, cod.Perca fluviatilis, European perch.Salmo trutta, trout.44

EU wide Fish Monitoring from 54 schools45

DL-PCBs+PCDD/FsPCDD/FsMay 2009:Countries: 11Schools: 51Fish samples:19246

TEQ(pg/g)07'199607'200407'200807'201207'203708'33308'33708'34708'33908'34308'36108'36508'36908'37308'38808'39208'39608'40008'48408'50708'54608'62108'64508'35608'65908'74808'75208'75608'76008'80508'80908'81908'87708'106608'107008'127508'139008'139408'156608'1596Low average level of all samples: 0,42 (pg TEQ/g)2,001,501,00AverageTEQ0,500,00Sample number47

River fish higher than coastal fishN Norway: n=27,S Norway n=4348

Outline of Presentation• Compliance with EU regulations• German Dioxin Crisis 2011• Chile Food/Feed monitoring 2008• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Slovak Republic Food/Feed monitoring• Norway Fish monitoring school project• Taiwan Monitoring of feed and food• BDS Client survey 2005• Other food monitoring programs using DR CALUX49

TACTRI/COA in Taiwan: Egg and Fish monitoringSample Nr of samples Fat % Total-TEQEggs 31 10 2.0 (1.3-4.8)Eel-Feed 6 9 0,4 (0.2-0.5)Seabass-Feed 9 11 0.9 (0.3-1.7)Tilapia-Feed 8 8 0.3 (0.2-0.5)Duck eggs 8 15 1.8 (1.4-2.4)Salted duck eggs 16 46 1,2 (0.6-2.8)50

Outline of Presentation• Compliance with EU regulations• German Dioxin Crisis 2011• Chile Food/Feed monitoring 2008• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Slovak Republic Food/Feed monitoring• Norway Fish monitoring school project• Taiwan Monitoring of feed and food• BDS Client survey 2005• Other food monitoring programs using DR CALUX51

BDS clients survey in 2006:Dioxin/PCB-TEQ by DR CALUX ® TEQin food samples (n=490)52

BDS clients survey in 2006:Dioxin/PCB-TEQ by DR CALUX ® TEQin feed samples (n=100)53

Outline of Presentation• Compliance with EU regulations• German Dioxin Crisis 2011• Chile Food/Feed monitoring 2008• Dutch Clay/Potato peeling crisis 2004• Sydney harbor fish screening 2005• Slovak Republic Food/Feed monitoring• Norway Fish monitoring school project• Taiwan Feed/Food Monitoring• BDS Client survey 2005• Other monitoring programs using DR CALUX54

Other Monitoring Programs using DR CALUX ®I. Ireland 2008 – used PCB electricity transformator oil in foodrecycling contaminated more than 80% of pig meat export Intensive monitoring program incl. DR CALUX ® .II. Italian „Mozarella Crisis“ in 2008 Intensive monitoring program of local farmer association incl.DR CALUX ® III. E.g. Israel, Mauretania, Chile, Thailand, China, Korea,Spain…. are currently running monitoring programs via DRCALUX55

Mother milk in Hong Kong and ChinaComparison DR CALUX-Total-TEQ (BDS) and HRGC/HRMS WHO-Total-TEQ (WHO Reference lab) for pooled breast milksamples (pg/g fat) from Hui et al. Chemosphere 69, 1287 (2007)181614DR CALUXWHO Lab (TEF1997)WHO Lab (TEF2005)1210864201. HongKong (eversmokers,n=25; age 26.5)2. HongKong (highdairy productintake,16;31)3. HongKong (highseafoodintake, 17;30.5)4. HongKong (lowdairy ANDlow seafoodintake, 32;31.4)5. HongKong (highdairy ORhigh seafoodintake, 28;32.6)6. MainlandChina (lowdairy ANDlow seafoodintake, 21;26.6)7. MainlandChina (highdairy ORhigh seafoodintake, 34;28.1)8. ChinaImmigrant(2–6 yearsstay in HongKong, 22;27.6)9. ChinaImmigrant (7years stay inHong Kongor above, 23;30.1)10. Overseas(1–10 yearsof overseasstay, 18;32.1)11. Overseas(11 years ofoverseasstay orabove, 10;25.7)Overall (246;30)56

Take home message• The DR CALUX cell-based screening method is used for feed/foodtesting since more than 15 years….and evaluated in many laboratory inmany countries• The new EC guidelines open now via the qualitative screeningapproach an easier and faster approach for the reporting of compliantor suspected samples• BDS is further focusing to decrease the costs for the PCDD/F/PCB-TEQ analysis (under ISO 17025 mode) via easier clean-up methods andRobotics!• The results of these studies shows again that the DR CALUX ® bioassayfor screening of PCDD/Fs and dioxin-like PCBs in feed and food is animportant device to identify the few percentage of the EU limitexceeding samples among the bulk of the compliant samples….!!!57