REGUL ATIONS 
COMMISSION REGUL ATION (EU) 2017/771 
of 3 May 2017 
amending Regulation (EC) No 152/2009 as regards the methods for the deter mination of the levels 
of dioxins and poly chlor inated biphen yls 
(Text with EEA relevance) 
THE EUR OPEAN COMMISSION, 
Having regar d to the Treaty on the Functioning of the European Union, 
Having regard to Regulation (EC) No 882/2004 of the European Parliament and of the Council of 29 Apr il 2004 on 
official controls perf ormed to ensure the verification of compliance with feed and food law, animal health and animal 
welf are rules (1), and in particular Article 11(4) thereof, 
Whereas: 
(1)  Commission Regulation (EC) No 152/2009 (2) includes methods for the deter mination of the levels of 
poly chlor inated dibenzo-p-dio xins (PCDDs), poly chlor inated dibenzofurans (PCDFs), dioxin-like poly chlor inated 
biphen yls (PCBs) and non-dio xin-like PCBs in feed. 
(2)  EU reference laborat ory for dioxins and PCBs in feed and food has provided evidence that analytical results for 
dioxins and PCBs in certain cases are not reliable when the performance criteria provided for in Part B of 
Annex V to Regulation (EC) No 152/2009 are not applied by laboratories performin g the analysis of sam ples 
take n by feed business operators in accordance with Regulation (EC) No 183/2005 of the European Parliament 
and of the Council (3). It is theref ore appropr iate to make the application of the performa nce criteria for the 
analysis of sam ples oblig atory . 
(3)  Given that the method of using a decision limit to ensure, with a certain probability , that an analytical result is 
above the maximum level, as provided for in Commission Decision 2002/657/EC (4), is no longer applied for the 
analysis of dioxins, furans and PCBs in feed, it is appropr iate to delet e that method and to keep only the method 
of the expanded uncer tainty using the coverag e factor of 2, which gives a level of conf idence of appro ximat ely 
95 %. 
(4)  Guidance documents for the measurement uncer tainty and for the estimation of the Limit of Detection (LOD) 
and Limit of Quantification (LOQ) have been developed. It is appropr iate to make referen ce to them. 
(5)  In line with the repor ting requirements for bioanalytical screening methods provided for in Part B of Annex V to 
Regulation (EC) No 152/2009, it is appropr iate to provide also for physico-che mical methods, to be used for 
screening, specifi c repor ting requirements in Chapt er II of that Part. 
(6)  Given that the analysis of dioxins, dioxin-like PCBs and non-dio xin-like PCBs are in most cases deter mined 
together , it is appropr iate to align the performa nce criteria for the non-dio xin-lik e PCBs provided for in point 3.3 
of Chapt er III of Part B of Annex V to Regulation (EC) No 152/2009 to the performa nce criteria for dioxins and 
dioxin-like PCBs. This is a simplification without substantial changes in practice as for non-dio xin-like PCBs the 
relative intensity of qualif ier ions compared to target ions is > 50 %. 4.5.2017 L 115/22 Official Jour nal of the European Union EN     
(1)OJ L 165, 30.4.2004, p. 1. 
(2)Commission Regulation (EC) No 152/2009 of 27 Januar y 2009 laying down the methods of samplin g and analysis for the official 
control of feed (OJ L 54, 26.2.2009, p. 1). 
(3)Regulation (EC) No 183/2005 of the European Parliament and of the Council of 12 Januar y 2005 laying down requirements for feed 
hygiene (OJ L 35, 8.2.2005, p. 1). 
(4)Commission Decision 2002/657/EC of 14 Aug ust 2002 implementing Council Directive 96/23/EC concer ning the performance of 
analytical methods and the interpretation of results (OJ L 221, 17.8.2002, p. 8). (7)  Follo wing exper ience gained, it is appropr iate to adapt some technical specifi cations such as reco veries of 
isotope-labelled standards provided for in points 7.3 and 7.5 of Chapt er III of Part B of Annex V to Regulation 
(EC) No 152/2009. 
(8)  Further more, there are several other minor modifications proposed to the current provisions to impro ve the 
consiste ncy in the terminology used, requir ing replacing the whole Part B of Annex V to Regulation (EC) 
No 152/2009 to maintain the readability of the text. 
(9)  Regulation (EC) No 152/2009 should theref ore be amended according ly. 
(10)  The measures provided for in this Regulation are in accordance with the opinion of the Standing Committ ee on 
the Food Chain and Animal Health, 
HAS ADOPTED THIS REGUL ATION: 
Article 1 
Part B of Annex V to Regulation (EC) No 152/2009 is amended in accordance with the Annex to this Regulation. 
Article 2 
This Regulation shall enter into force on the twentieth day following that of its publication in the Official Journal of the 
European Union. 
This Regulation shall be binding in its entirety and directly applicable in all Member State s. 
Done at Brussels, 3 May 2017. 
For the Commission 
The Presid ent 
Jean-Claude JUNCKER  4.5.2017 L 115/23 Official Jour nal of the European Union EN     ANNEX 
In Annex V to Regulation (EC) No 152/2009, Part B ‘DETERMINA TION OF THE LEVELS OF DIO XINS (PCDD/PCDF) 
AND PCBs’ is replaced by the following:  
‘B. DETERMINA TION OF THE LEVELS OF DIO XINS (PCDD/PCDF) AND PCBs 
CHAPTER I 
Methods of sampling and inter pretation of analytical results 
1.  Scope and definitions 
The samples intended for the official control of the levels of poly chlor inated dibenzo-p-dio xins (PCDDs), 
poly chlor inated dibenzofurans (PCDFs), dioxin-like poly chlor inated biphen yls (PCBs) (1) and non dioxin-like 
PCBs in feed shall be take n in accordance with the provisions of Annex I. The quantitative requirements in 
relation to the control of substances or products unif ormly distr ibut ed throughout the feed as provided for 
in point 5.1. of Annex I shall be applied. Aggreg ate samples thus obtained shall be considered representative 
for the lots or sublots from which they are take n. Complian ce with maximum levels laid down by Directive 
2002/32/EC shall be established on the basis of the levels deter mined in the laboratory samples. 
For the purposes of this Part B, the definitions laid down in Annex I to Commission Decision 
2002/657/EC (2) shall apply . 4.5.2017 L 115/24 Official Jour nal of the European Union EN     
(1)Table of TEF (= toxic equivalency factors) for PCDDs, PCDFs and dioxin-lik e PCBs: WHO- TEFs for human risk assessment based on 
the conclusions of the World Health Org anization (WHO) — Intern ational Programme on Chemical Safety (IPCS) exper t meeting 
whic h was held in Geneva in June 2005 (Mar tin van den Berg et al., The 2005 World Health Org anization Re-evaluation of Human 
and Mammalian Toxic Equivalency Fact ors for Dioxins and Dioxin-like Compo unds. Toxicological Sciences 93(2), 223–241 (2006)). 
Cong ener TEF value Cong ener TEF value 
Dibenzo-p-dio xins (“PCDDs”) and Dibenzo-p- 
furans '(“PCDFs”)  “Dio xin-like” PCBs 
Non-or tho PCBs + Mono-or tho PCBs 
2,3,7,8- TCDD  1    
1,2,3,7,8-P eCDD  1 Non-or tho PCBs   
1,2,3,4,7,8-HxCDD  0,1 PCB 77  0,0001 
1,2,3,6,7,8-HxCDD  0,1 PCB 81  0,0003 
1,2,3,7,8,9-HxCDD  0,1 PCB 126  0,1 
1,2,3,4,6,7,8-HpCDD  0,01 PCB 169  0,03 
OCDD  0,0003 Mono-or tho PCBs   
2,3,7,8- TCDF  0,1 PCB 105  0,00003 
1,2,3,7,8-P eCDF  0,03 PCB 114  0,00003 
2,3,4,7,8-P eCDF  0,3 PCB 118  0,00003 
1,2,3,4,7,8-HxCDF  0,1 PCB 123  0,00003 
1,2,3,6,7,8-HxCDF  0,1 PCB 156  0,00003 
1,2,3,7,8,9-HxCDF  0,1 PCB 157  0,00003 
2,3,4,6,7,8-HxCDF  0,1 PCB 167  0,00003 
1,2,3,4,6,7,8-HpCDF  0,01 PCB 189  0,00003 
1,2,3,4,7,8,9-HpCDF  0,01    
OCDF  0,0003     
Abbreviations used: “T” = tetra; “Pe” = penta; “Hx” = hexa; “Hp” = hepta; “O” = octa; “CDD” = chlorodibenzodio xin; “CDF” = chlorodiben ­
zofuran; “CB” = chlorobiphen yl. 
(2)Commission Decision 2002/657/EC of 14 Aug ust 2002 implementing Council Directive 96/23/EC concer ning the performance of 
analytical methods and inter pretation of results (OJ L 221, 17.8.2002, p. 8). In addition to those definit ions, the following definit ions shall apply for the purpose of this Part B: 
“Screening methods” means methods used for selection of those sam ples with levels of PCDD/Fs and dioxin- 
like PCBs that excee d the maximum levels or the action thresholds. They shall allow a cost-effective high 
sample-th roughput, thus increasing the chance to disco ver new incidents with high exposure and health 
risks to consumers. Screening methods shall be based on bioanalytical or GC-MS methods. Results from 
samples excee ding the cut-off value used to check compl iance with the maximum level shall be verified by 
a full re-analysis from the original samp le using a conf irmatory method. 
“Confi rmatory methods” means methods that provide full or complementary information enabling the 
PCDD/Fs and dioxin-like PCBs to be identif ied and quantifi ed unequiv ocally at the maximum or in case of 
need at the action threshold. Such methods utilize gas chromat ography /high resolution mass spectrometr y 
(GC-HRMS) or gas chromatograph y/tandem mass spectrometr y (GC-MS/MS). 
2.  Compliance of the lot or sublot with the maximum level 
2.1.  As regards non-dioxin -like PCBs 
The lot or sublot compl ies with the maximum level if the analytical result for the sum of PCB 28, PCB 52, 
PCB 101, PCB 138, PCB 153 and PCB 180 (hereaf ter refer red to as non-dio xin-like PCBs) does not excee d 
the maximum level laid down by Directive 2002/32/EC, taking into account the expanded measurement 
uncer tainty (1). The lot or sublot does not compl y with the maximum level as laid down by Directive 
2002/32/EC, if the mean of two upper -bound (2) analytical results obtained from duplicat e analysis (3), 
taking into account the expanded measurement uncer tainty , excee ds the maximum level beyond reasonable 
doubt, i.e. the analysed concentration after deduction of the expanded measurement uncer tainty is used to 
assess compl iance. 
The expanded measurement uncer tainty is calculat ed using a coverag e factor of 2 whic h gives a level of 
conf idence of appro ximately 95 %. A lot or sublot is non-com pliant if the mean of the measured values 
minus the expanded uncer tainty of the mean is above the maximum level. 
The rules, mentioned in the paragraphs above under this point, shall apply for the analytical result obtained 
on the sample for official control. In case of analysis for defe nce or referen ce purposes, the national rules 
shall apply . 
2.2.  As regards PCDD/Fs and dioxin-like PCBs 
The lot or sublot compl ies with the maximum level if the result of a sing le analysis 
—  performed by a screening method with a false-compliant rate belo w 5 %, indicates that the level does 
not exceed the respective maximum level of PCDD/Fs and the sum of PCDD/Fs and dioxin-lik e PCBs laid 
down by Directive 2002/32/EC, 
—  performed by a confir matory method, does not exceed the respective maximum level of PCDD/Fs and 
the sum of PCDD/Fs and dioxin-like PCBs laid down by Directive 2002/32/EC, taking into account the 
expanded measurement uncer tainty . 4.5.2017 L 115/25 Official Jour nal of the European Union EN     
(1)The principles descr ibed in the “Guidance Document on Measurement Uncer tainty for Laboratories perfor ming PCDD/F and PCB 
Analysis using Isotope Dilution Mass Spectrometr y” (http://ec.europa.eu/f ood/safety/animal-f eed_en) shall be followed when applicable. 
(2)The concept of “upper -bound” requires using the limit of quantification for the contr ibution of each non-quantified congener . The 
concept of “lower-bound” requires using zero for the contr ibution of each non-quantified cong ener . The concept of “medium-bound” 
requires using half of the limit of quantific ation calculating the contr ibution of each non-quantif ied cong ener . 
(3)Duplicat e analysis: Separate analysis of the analyt es of interest using a second aliquot of the same homog enized sample. In general, the 
requirements for duplicate analysis as provided for in Annex II, Chap ter C, point 3 apply . However , for methods with the use 
of 13C-labelled inter nal standard for the relevant analyt es, the duplicate analysis is only necessar y if the result of the first determination is 
not compliant. The duplicate analysis is necessar y to exclude the possibility of internal cross-contamination or an accidental mix-up of 
samples. In case the analysis is perfor med in the course of a contamination incident, confir mation by duplicate analysis may be omitted 
in case the samples selected for analysis are through traceability linked to the contamination incident and the level found is significantly 
above the maximum level. For screening assa ys a cut-off value shall be established for decisions on sample compl iance with the 
respective maximum levels set for either PCDD/Fs, or for the sum of PCDD/Fs and dioxin-like PCBs. 
The lot or sublot does not comply with the maximum level as laid down by Directive 2002/32/EC if the 
mean of two upper -bound (1) analytical results obtained from duplicate analysis (2) using a conf irmat ory 
method, taking into account the expanded measurement uncer tainty , exceed s the maximum level beyond 
reasonable doubt, i.e. the analysed concentration after deduction of the expanded measurement uncer tainty 
is used to assess compl iance. 
The expanded measurement uncer tainty is calculat ed using a coverag e factor of 2 whic h gives a level of 
conf idence of appro ximately 95 %. A lot or sublot is non-com pliant if the mean of the measured values 
minus the expanded uncer tainty of the mean is above the maximum level. 
The sum of the estimate d expanded uncer tainties of the separate analytical results of PCDD/Fs and dioxin- 
like PCBs shall be used for the sum of PCDD/Fs and dioxin-lik e PCBs. 
The rules, mentioned in the paragraphs above under this point, shall apply for the analytical result obtained 
on the sample for official control. In case of analysis for defe nce or referen ce purposes, the national rules 
shall apply . 
3.  Results exceeding action thresholds as laid down in Annex II to Directiv e 2002/32/EC 
Action thresholds serve as a tool for the selection of samp les in those cases where it is necessar y to identify 
a source of contamination and to take measures for its reduction or elimination. Screening methods shall 
establish the appropr iate cut-off values for selection of those sample s. Where signif icant efforts are 
necessar y to identify a source and to reduce or eliminate the contamination, it is appropr iate to confir m 
exceed ance of the action thresholds by duplicat e analysis using a conf irmat ory method and taking into 
account the expanded measurement uncer tainty (3). 
CHAPTER II 
Sample prepar ation and requir ements for met hods of analy sis used in offical control of the levels of 
dioxins (PCDD/Fs) and dioxin-like PCBs in feed 
1.  Field of application 
The requirements set out in this Chapt er shall be applied where feed is analysed for the official control of 
the levels of 2,3,7,8-substituted PCDD/Fs and dioxin-like PCBs and as regar ds sample preparation and 
analytical requirements for other regulator y purposes, whic h includes the controls perf ormed by the feed 
business operat or to ensure compl iance with the provisions of Regulation (EC) No 183/2005 of the 
European Parliament and of the Council (4). 4.5.2017 L 115/26 Official Jour nal of the European Union EN     
(1)The concept of “upper -bound” requires using the limit of quantificat ion for the contr ibution of each non-quantified congener to the 
Toxic Equivalent (TEQ). The concept of “lower-bound” requires using zero for the contr ibution of each non-quantified cong ener to the 
TEQ. The concept of “medium-bound” requires using half of the limit of quantification calculating the contr ibution of each non- 
quantified cong ener to the TEQ. 
(2)In general, the requirements for duplicate analysis as provided for in Annex II, Chapt er C, point 2 apply . However , for confir mat ory 
methods with the use of 13C-labelled internal standard for the relevant analytes, the duplicate analysis is only necessar y if the result of the 
first deter mination is not compliant. The duplicate analysis is necessar y to exclud e the possibility of internal cross-contamination or an 
accidental mix-up of samples. In case the analysis is perf ormed in the course of a contamination incident, confir mation by duplicate 
analysis may be omitted in case the samples selected for analysis are through traceability linked to the contamination incident and the 
level found is signifi cantly above the maximum level. 
(3)Identical explanation and requirements for duplicate analysis for control of action thresholds as in footnote 2 above for maximum levels. 
(4)Regulation (EC) No 183/2005 of the European Parliament and of the Council of 12 Januar y 2005 laying down requirements for feed 
hygiene (OJ L 35, 8.2.2005, p. 1). Monitoring for the presence of PCDD/Fs and dioxin-lik e PCBs in feed may be performe d with two different 
types of analytical methods: 
(a)  Screening methods 
The goal of screening methods is to select those sam ples with levels of PCDD/Fs and dioxin-like PCBs 
that exceed the maximum levels or the action thresholds. Screening methods shall ensure cost-effe ctive 
high sample-throughput, thus increasing the chance to disco ver new incidents with high exposure and 
health risks of consumers. Their application shall aim to avoid false-compliant results. They may 
compris e bioanalytical and GC-MS methods. 
Screening methods compar e the analytical result with a cut-off value, providing a yes/no-decision over 
the possible exceed ance of the maximum level or action threshold. The concentration of PCDD/Fs and 
the sum of PCDD/Fs and dioxin-like PCBs in samples suspected to be non-com pliant with the maximum 
level shall be determi ned or confir med by a conf irmatory method. 
In addition, screening methods may give an indication of the levels of PCDD/Fs and dioxin-like PCBs 
present in the sample. In case of application of bioanalytical screening methods the result is expressed 
as Bioanalytical Equivalents (BEQ), whereas in case of application of physico-chemical GC-MS methods 
it is expressed as Toxic Equivalents (TEQ). The numer ically indicated results of screening methods are 
suitable for demonstrating compl iance or suspecte d noncompliance or exceedance of action thresholds 
and give an indication of the range of levels in case of follow-up by conf irmatory methods. They are 
not suitable for purposes such as evaluation of backg round levels, estimation of intake, following of 
time trends in levels or re-evaluation of action thresholds and maximum levels. 
(b)  Confir mator y methods 
Confi rmat ory methods allow the unequivocal identif ication and quantif ication of PCDD/Fs and dioxin- 
like PCBs present in a sam ple and provide full information on cong ener level. Theref ore, those methods 
allow the control of maximum levels and action thresholds, including the conf irmation of results 
obtained by screening methods. Further more, results may be used for other purposes such as deter mina ­
tion of low backg round levels in feed monitori ng, following of time trends, exposure assessment and 
building of a database for possible re-evaluation of action thresholds and maximum levels. They are also 
impor tant for establishing congener patt erns in order to identify the source of a possible contamination. 
Such methods utilise GC-HRMS. For conf irming compliance or non-com pliance with the maximum 
level, also GC-MS/MS can be used. 
2.  Backg round 
For calculation of TEQ concentrations, the concentrations of the individual substances in a given sample 
shall be multiplied by their respective Toxic Equivalency Fact or (TEF) (see footnote 1 of Chapt er I) and 
subsequently summed to give the total concentration of dioxin-like compo unds expressed as TEQs. 
For the purposes of this Part B, the accept ed specifi c limit of quantifi cation of an individual cong ener 
means the lowest content of the analyt e that can be measured with reasonable statistical certainty , fulfilling 
the identif ication criteria as descr ibed in internati onally recognised standards, for example, in standard EN 
16215:2012 (Animal feed — Determination of dioxins and dioxin-like PCBs by GC-HRMS and of indicator 
PCBs by GC-HRMS) and/or in EPA methods 1613 and 1668 as revised. 
The limit of quantification of an individual cong ener may be identif ied as 
(a)  the concentration of an analyte in the extract of a sample whic h produces an instr umental response at 
two differ ent ions to be monitor ed with a S/N (signal/noise) ratio of 3:1 for the less intensi ve raw data 
signal; or 4.5.2017 L 115/27 Official Jour nal of the European Union EN     (b)  if for technical reasons the signal-to-no ise calculation does not provide reliable results, the lowest 
concentration point on a calibration curve that gives an acceptable (≤ 30 %) and consist ent (measured 
at least at the start and at the end of an analytical series of sam ples) deviation to the average relative 
response factor calculated for all points on the calibration curve in each series of samples . The limit of 
quantifi cation (LOQ) is calculated from the lowest concentration point taking into account the reco very 
of internal standards and sample intak e. 
Bioanalytical screening methods will not give results at the cong ener level but merely an indication (1) of 
the TEQ level, expressed in BEQ to ackno wledge the fact that not all compo unds present in a sample 
extract that produce a response in the test may fulfill or meet all requirements of the TEQ-pr inciple. 
Screening and confir matory methods may only be applied for control of a certain matr ix if the methods are 
sensitive enough to detect levels reliably at the action threshold or maximum level. 
3.  Quality assurance requirements 
3.1.  Measures shall be taken to avoid cross-contamination at each stage of the sampli ng and analysis procedure. 
3.2.  The sam ples shall be stored and transpor ted in glass, aluminum, polyprop ylene or polyet hylene containers 
suitable for storag e without any influence on the levels of PCDD/Fs and dioxin-lik e PCBs in the sam ples. 
Traces of paper dust shall be remo ved from the sample container . 
3.3.  The sam ple storag e and transpor tation shall be performe d in a way that maintains the integrit y of the feed 
sample. 
3.4.  Insof ar as relevant, each laboratory sam ple shall be finely grinded and mixe d thoroughly using a process 
that has been demonstrate d to achi eve compl ete homogenisatio n (for examp le, ground to pass a 1 mm 
sieve). Samp les shall be dried before grinding if the moisture cont ent is too high. 
3.5.  Control of reagents, glassware and equipment for possible influence of TEQ- or BEQ-based results shall be 
carried out. 
3.6.  A blank analysis shall be performe d by carrying out the entire analytical procedure omitting only the 
sample. 
3.7.  For bioanalytical methods, all glassware and solvents used in analysis shall be tested to be free of 
compo unds that interfere with the detection of targe t compo unds in the working rang e. Glassware shall be 
rinsed with solvents or heat ed at temperatures suitable to remo ve traces of PCDD/Fs, dioxin-like compo unds 
and interfering compounds from its surface. 
3.8.  Sample quantity used for the extraction shall be sufficient to fulfill the requirements with respect to 
a sufficiently low working rang e including the concentrations of maximum levels or action threshold. 
3.9. The specif ic sam ple preparation procedures used for the products under consideration shall follow internat­
ionally accept ed guidelines. 
4.  Requirements for laborator ies 
4.1.  In accordance with the provisions of Regulation (EC) No 882/2004, laboratories shall be accredit ed by 
a recognised body operating in accordance with ISO Guide 58 to ensure that they are applying analytical 
quality assurance. Laboratori es shall be accredited following the EN ISO/IEC 17025 standard. The principles 
as descr ibed in the Technical Guidelines for the estimation of measurement uncer tainty and limits of quanti ­
fication for PCDD/F and PCB analysis shall be followed when applicable (2) 4.5.2017 L 115/28 Official Jour nal of the European Union EN     
(1)Bioanalytical methods are not specific to those cong eners included in the TEF-scheme. Other structurally relate d AhR -active compounds 
may be present in the sample extract whic h contr ibut e to the overall response. Theref ore, bioanalytical results cannot be an estimat e but 
rather an indication of the TEQ level in the sample. 
(2)“Guidance Document on Measurement Uncer tainty for Laboratories perfor ming PCDD/F and PCB Analysis using Isotope Dilution Mass 
Spectrometr y” (http://ec.europa.eu/f ood/safety/animal-f eed_en), “Guidance Document on the Estimation of LOD and LOQ for 
Measurements in the Field of Contaminants in Feed and Food” (http://ec.europa.eu/f ood/safety/anim al-feed_en). 4.2.  Laboratory proficiency shall be proven by the continuous successful participation in inter -laborat ory studies 
for the determination of PCDD/Fs and dioxin-like PCBs in relevant feed matr ices and concentration rang es. 
4.3.  Laboratori es applying screening methods for the routine control of sample s shall establish a close 
cooperation with laborat ories applying the conf irmatory method, both for quality control and confir mation 
of the analytical result of suspecte d samp les. 
5.  Basic requirements to be met by analytical procedure for dioxins (PCDD/Fs) and dioxin-like PCBs 
5.1.  Low working range and limits of quantif ication 
For PCDD/Fs, detectable quantities shall be in the upper femtogram (10– 15g) rang e because of extreme 
toxicity of some of these compounds. For most PCB congeners a limit of quantif ication in the nanogram 
(10– 9g) range is already sufficient. For the measurement of the more toxic dioxin-like PCB cong eners (in 
particular non-or tho-substitut ed congeners), the lower end of the working rang e shall reac h the low 
picogram (10– 12g) levels. For all other PCB cong eners a limit of quantification in the nanogram (10– 9g) 
range is suffi cient. 
5.2.  High selectivity (specif icity) 
5.2.1.  A distinction is required between PCDD/Fs and dioxin-lik e PCBs and a multitude of other , coextract ed and 
possibly interfe ring compounds present at concentrations up to several orders of magnitude higher than 
those of the analytes of inter est. For GC-MS methods, a differ entiation among various congeners is required, 
such as between toxic (for exam ple, the seventeen 2,3,7,8-substituted PCDD/Fs, and twelve dioxin-like PCBs) 
and other cong eners. 
5.2.2.  Bioanalytical methods shall be able to detec t the target compo unds as the sum of PCDD/Fs, and/or dioxin- 
like PCBs. Sample clean-up shall aim at remo ving compounds causing false non-compliant results or 
compo unds that may decrease the response, causing false compl iant results. 
5.3.  High accur acy (trueness and precision, bioassay apparen t recover y) 
5.3.1.  For GC-MS methods, the deter mination shall provide a valid estimate of the true concentration in a sample. 
High accuracy is required to avoid the rejection of a sample analysis result on the basis of poor reliability of 
the determined TEQ level. Accuracy is expressed as trueness (differ ence between the mean value measured 
for an analyt e in a certified mat erial and its certified value, expressed as a percentage of this value) and 
precision (RSDR relative standard deviation calculat ed from results generated under reproducibility 
conditions). 
5.3.2.  For bioanalytical methods, the bioassa y apparent reco very shall be determi ned. Bioassa y apparent reco very 
means the BEQ level calculat ed from the TCDD or PCB 126 calibration curve corrected for the blank and 
then divided by the TEQ level determined by the conf irmatory method. It aims at correcting factors like the 
loss of PCDD/Fs and dioxin-like compo unds during the extraction and clean-up step s, co-extracte d 
compo unds increasing or decreasing the response (agonistic and antagonistic effects), the quality of the 
curve fit, or diffe rences between the TEF values and the Relative Poten cy (REP) values. The bioassa y 
apparent reco very is calculated from suitable reference samples with representative congener patte rns 
around the level of interest. 
5.4.  Validation in the range of maximum level and gener al quality contro l measures 
5.4.1.  Laboratori es shall demonstrate the performa nce of a method in the range of the maximum level, for 
example, 0,5x, 1x and 2x the maximum level with an acceptable coefficient of variation for repeated 
analysis, during the validation procedure and during routine analysis. 4.5.2017 L 115/29 Official Jour nal of the European Union EN     5.4.2.  Regular blank controls and spiking exper iments or analysis of control samples (prefera bly, if available, 
certified reference mate rial) shall be performe d as internal quality control measures. Quality control char ts 
for blank controls, spiking exper iments or analysis of control samples shall be recorded and check ed to 
make sure the analytical performa nce is in accordance with the requirements. 
5.5.  Limit of quantif ication 
5.5.1. For a bioanalytical screening method, the establishment of the limit of quantifi cation (LOQ) is not an indis ­
pensable requirement but the method shall prove that it can diffe rentiate between the blank and the cut-off 
value. When providing a BEQ level, a repor ting level shall be established to deal with samples showing 
a response belo w this level. The repor ting level shall be demonstrated to be differ ent from procedure blank 
samples at least by a factor of three, with a response belo w the working range. It shall theref ore be 
calculat ed from sam ples containing the target compo unds around the required minimum level, and not 
from an S/N ratio or an assa y blank. 
5.5.2.  The LOQ for a conf irmatory method shall be about one fifth of the maximum level. 
5.6.  Analytical criteria 
For reliable results from conf irmatory or screening methods, the following criteria shall be met in the rang e 
of the maximum level for the TEQ or BEQ value, respectively , whether determined as total TEQ or total 
BEQ (as the sum of PCDD/Fs and dioxin-like PCBs) or separately for PCDD/Fs and dioxin-like PCBs:  
Screening with bioanaly ­
tical or physico-chemical 
methods Confir matory methods 
False-compliant rate(*) < 5 %  
Trueness  – 20 % to + 20 % 
Repeatability (RSDr) < 20 %  
Interm ediat e precision (RSDR) < 25 % < 15 % 
(*)  With respect to the maximum levels.  
5.7.  Specif ic requiremen ts for screening methods 
5.7.1.  Both GC-MS and bioanalytical methods may be used for screening. For GC-MS methods the requirements 
laid down in point 6 shall be met. For cell based bioanalytical methods specifi c requirements are laid down 
in point 7. 
5.7.2.  Laboratori es applying screening methods for the routine control of sample s shall establish a close 
cooperation with laboratori es applying the conf irmat ory method. 
5.7.3.  Performan ce verification of the screening method is required during routine analysis, by analytical quality 
control and on-going method validation. There shall be a continuous programme for the control of 
compl iant results. 4.5.2017 L 115/30 Official Jour nal of the European Union EN     5.7.4.  Check on possible suppression of the cell response and cytotoxicity: 
20 % of the sample extracts shall be measured in routine screening without and with 2,3,7,8- TCDD added 
corresponding to the maximum level or action threshold, to chec k if the response is possibly suppressed by 
interfe ring substances present in the sam ple extract. The measured concentration of the spik ed sample shall 
be compared to the sum of the concentration of the unspiked extract plus the spiking concentration. If this 
measured concentration is more than 25 % lower than the calculat ed (sum) concentration, this is an 
indication of pote ntial signal suppression and the respective sample shall be submitted to GC-HRMS 
conf irmatory analysis. Results shall be monitore d in quality control charts. 
5.7.5.  Quality control on compliant samples: 
Appro ximat ely 2 to 10 % of the compl iant sam ples, depending on sample matr ix and laborat ory 
exper ience, shall be confir med by GC/HRMS. 
5.7.6.  Determination of false-compliant rates from quality control data: 
The rate of false-compliant results from screening of samples belo w and above the maximum level or the 
action threshold shall be determined. Actual false-com pliant rates shall be belo w 5 %. When a minimum of 
20 conf irmed results per matr ix/matr ix group is available from the quality control of compliant samples , 
conclusions on the false compliant rate shall be drawn from this database. The results from samples 
analysed in ring trials or during contamination incidents, covering a concentration rang e up to for example 
2x the maximum level (ML), may also be included in the minimum of 20 results for evaluation of the false- 
compl iant rate. The samples shall cover most frequent cong ener patte rns, representing various sources. 
Although screening assa ys shall preferentially aim to detect samples exceeding the action threshold, the 
criterion for determining false-com pliant rates is the maximum level, taking into account the expanded 
measurement uncer tainty of the conf irmat ory method. 
5.7.7.  Potential non-com pliant samples from screening shall alwa ys be verified by a full re-analysis of the original 
sample by a conf irmat ory method of analysis. These samples may also be used to evaluate the rate of false 
non-com pliant results. For screening methods, the rate of false non-com pliant results shall be the fraction of 
results conf irmed to be compl iant from confir matory analysis, while in previous screening the sam ple has 
been declared to be poten tially non-com pliant. Evaluation of the advantage s of the screening method shall 
be based on comparison of false-non-compliant sam ples with the total number of samples check ed. This 
rate shall be low enough to make the use of a screening tool advantageous. 
5.7.8.  Under validation conditions, bioanalytical methods shall provide a valid indication of the TEQ level, 
calculat ed and expressed as BEQ. 
Also for bioanalytical methods carried out under repeated conditions, the intra-laboratory RSDr would 
typically be smaller than under reproducibility conditions(RSDR) 
6.  SPECIFIC requirements for GC-MS methods to be complied with for screening or conf irmatory 
pur poses 
6.1.  Accep table differences betw een upper -bound and lower-bound WHO- TEQ results 
The differ ence between upper -bound level and lower-bound level shall not exceed 20 % for conf irmation of 
exceed ance of maximum level or in case of need of action thresholds. 4.5.2017 L 115/31 Official Jour nal of the European Union EN     6.2.  Contro l of recoveries 
6.2.1.  Addition of 13C-labelled 2,3,7,8-ch lorine-substitut ed intern al PCDD/F standards and of 13C-labelled intern al 
dioxin-like PCB standards shall be carried out at the very beginning of the analytical method e.g. prior to 
extraction in order to validat e the analytical procedure. At least one cong ener for each of the tetra- to octa- 
chlor inated homologous groups for PCDD/Fs and at least one congener for each of the homologous groups 
for dioxin-lik e PCBs shall be added (alter natively , at least one congener for each mass spectrometr ic select ed 
ion recording function used for monitori ng PCDD/Fs and dioxin-like PCBs). In the case of conf irmat ory 
methods, all 17 13C-labelled 2,3,7,8-substituted internal PCDD/F standards and all 12 13C-labelled intern al 
dioxin-like PCB standards shall be used. 
6.2.2.  Relative response factors shall also be deter mined for those cong eners for whic h no 13C-labelled analogue is 
added by using appropr iate calibration solutions. 
6.2.3.  For feed of plant origin and feed of animal origin containing less than 10 % fat, the addition of the intern al 
standards shall be mandat ory prior to extraction. For feed of animal origin containing more than 10 % fat, 
the internal standards shall be added either before or after fat extraction. An appropr iate validation of the 
extraction effic iency shall be carried out, depending on the stage at whic h internal standards are introduced. 
6.2.4.  Prior to GC-MS analysis, 1 or 2 reco very (surrogat e) standard(s) shall be added. 
6.2.5.  Control of reco very is required. For conf irmat ory methods, the reco veries of the individual intern al 
standards shall be in the rang e of 60 to 120 %. Lower or higher reco veries for individual congeners, in 
particular for some hepta - and octa- chlor inated dibenzo-p-dio xins and dibenzofurans, shall be acceptable 
on the condition that their contr ibution to the TEQ value does not exceed 10 % of the tota l TEQ value 
(based on sum of PCDD/F and dioxin-like PCBs). For GC-MS screening methods, the reco veries shall be in 
the range of 30 to 140 %. 
6.3.  Removal of interfer ing substances 
—  Separation of PCDD/Fs from interf ering chlor inated compo unds such as non-dio xin-like PCBs and 
chlor inated diphen yl ethers shall be carried out by suitable chromat ographic techniques (prefer ably with 
a florisil, alumina and/or carbon column). 
—  Gas-c hromatographic separation of isomers shall be < 25 % peak to peak between 1,2,3,4,7,8-HxCDF 
and 1,2,3,6,7,8-HxCDF . 
6.4.  Calibr ation with standar d curve 
The rang e of the calibration curve shall cover the relevant rang e of maximum level or action thresholds. 
6.5.  Specif ic criteria for confirmator y methods 
—  For GC-HRMS: 
In HRMS, the resolution shall typically be greater than or equal to 10 000 for the entire mass range at 
10 % valley . 
Fulfilment of further identification and conf irmation criteria as descr ibed in internatio nally recognised 
standards, for example, in standard EN 16215:2012 (Animal feed — Determination of dioxins and 
dioxin-like PCBs by GC-HRMS and of indicator PCBs by GC-HRMS) and/or in EPA methods 1613 
and 1668 as revised. 4.5.2017 L 115/32 Official Jour nal of the European Union EN     —  For GC-MS/MS: 
Monitoring of at least 2 specifi c precursor ions, each with one specifi c corresponding transition product 
ion for all labelled and unlabelled analytes in the scope of analysis. 
Maximum permitted tolerance of relative ion intensi ties of ± 15 % for selected transition product ions in 
comparison to calculated or measured values (averag e from calibration standards), applying identical 
MS/MS conditions, in particular collision energy and collision gas pressure, for each transition of an 
analyte . 
Resolution for each quadr upole to be set equal to or better than unit mass resolution (unit mass 
resolution: suffic ient resolution to separate two peaks one mass unit apar t) in order to minimise 
possible interferences on the analytes of interest. 
Fulfilment of the further criteria as descr ibed in internationally recognised standards, for example, in 
standard EN 16215:2012 (Animal feed — Determination of dioxins and dioxin-like PCBs by GC-HRMS 
and of indicator PCBs by GC-HRMS) and/or in EPA methods 1613 and 1668 as revised, excep t the 
oblig ation to use GC-HRMS. 
7.  Specif ic requirements for bioanalytical methods 
Bioanalytical methods are methods based on the use of biological principles like cell-based assa ys, recep tor- 
assa ys or immunoassa ys. This point 7 establishes requirements for bioanalytical methods in general. 
A screening method in principle classif ies a sample as compl iant or suspect ed to be non-complian t. For 
this, the calculated BEQ level is compared to the cut-off value (see point 7.3). Sam ples belo w the cut-off 
value are declared compl iant, sam ples equal or above the cut-off value are suspected to be non-com pliant, 
requir ing analysis by a conf irmatory method. In practice, a BEQ level corresponding to two- thirds of the 
maximum level may serve as cut-off value provided that a false-compliant rate belo w 5 % and an acceptable 
rate for false non-com pliant results are ensured. With separate maximum levels for PCDD/Fs and for the 
sum of PCDD/Fs and dioxin-like PCBs, checkin g compl iance of samples without fractionation requires 
appropr iate bioassa y cut-off values for PCDD/Fs. For checking of samples exceeding the action thresholds, 
an appropr iate percentage of the respective action threshold shall suit as cut-off value. 
If an indicative level is expressed in BEQs, samp le results shall be in the working rang e and shall exceed the 
repor ting limit (see points 7.1.1 and 7.1.6). 
7.1.  Eval uation of the test respo nse 
7.1.1.  General requirements 
—  When calculating the concentrations from a TCDD calibration curve, values at the higher end of the 
curve will show a high variation (high coeffi cient of variation (CV)). The working range is the area 
where this CV is smaller than 15 %. The lower end of the working rang e (repor ting limit) shall be set at 
least by a factor of three above the procedure blanks. The upper end of the working range is usually 
represent ed by the EC70 value (70 % of maximal effective concentration), but lower if the CV is higher 
than 15 % in this range. The working range shall be established during validation. Cut-off values (see 
point 7.3) shall be well within the working range . 
—  Standard solutions and sam ple extracts shall be tested in triplicate or at least in duplicat e. When using 
duplicat es, a standard solution or a control extract tested in four to six wells divided over the plate shall 
produce a response or concentration (only possible in the working range ) based on a CV < 15 %. 4.5.2017 L 115/33 Official Jour nal of the European Union EN     7.1.2.  Calibration 
7.1.2.1.  Calibration with standard curve 
—  Levels in samples shall be estimat ed by compari son of the test response with a calibration curve of 
TCDD (or PCB 126 or a PCDD/PCDF/dio xin-like PCB standard mixture) to calculate the BEQ level in the 
extract and subsequently in the sample. 
—Calibration curves shall contain 8 to 12 concentrations (at least in duplicat es), with enough concen ­
trations in the lower part of the curve (working rang e). Special attention shall be paid to the quality of 
the curve-fit in the working range . As such, the R2 value is of little or no value in estimating the 
goodness of fit in non-linear regression. A better fit shall be achi eved by minimising the diffe rence 
between calculated and obser ved levels in the working rang e of the curve, for exam ple by minimising 
the sum of squared residuals. 
—  The estimated level in the sam ple extract shall be subsequently corrected for the BEQ level calculat ed for 
a matr ix or solvent blank sam ple (to account for impur ities from solvents and chemicals used), and the 
apparent reco very (calculated from the BEQ level of suitable reference sample s with representative 
cong ener patter ns around the maximum level or action threshold). To perform a reco very correction, 
the apparent reco very shall be within the required range (see point 7.1.4). Reference samples used for 
reco very correction shall compl y with the requirements laid down in point 7.2. 
7.1.2.2.  Calibration with referen ce samples 
Alte rnatively , a calibration curve prepared from at least four reference sam ples (see point 7.2.4): one matr ix 
blank, plus three referen ce samples at 0,5x, 1x and 2x the maximum level or action threshold may be used, 
eliminating the need to correct for blank and reco very if matr ix proper ties of the referen ce sam ples match 
those of the unkno wn samples. In this case, the test response corresponding to two-thirds of the maximum 
level (see point 7.3) may be calculated directly from these sam ples and used as cut-off value. For checkin g 
of samples exceeding the action thresholds, an appropr iate percentage of these action thresholds shall suit 
as cut-off value. 
7.1.3.  Separat e determ ination of PCDD/Fs and dio xin-lik e PCBs 
Extracts may be split into fractions containing PCDD/Fs and dioxin-like PCBs, allowing a separate indication 
of PCDD/Fs and dioxin-like PCB TEQ levels (in BEQ). A PCB 126 standard calibration curve shall preferen ­
tially be used to evaluate results for the fraction containing dioxin-like PCBs. 
7.1.4.  Bioassa y apparent reco ver ies 
The “bioassa y apparent reco very” shall be calculat ed from suitable reference sam ples with representative 
cong ener patter ns around the maximum level or action threshold and expressed as percentage of the BEQ 
level in compari son to the TEQ level. Depending on the type of assa y and TEFs (1) used, the differences 
between TEF and REP factors for dioxin-like PCBs can cause low apparent reco veries for dioxin-like PCBs in 
compari son to PCDD/Fs. Theref ore, if a separate determination of PCDD/Fs and dioxin-like PCBs is 
performe d, bioassa y apparent reco veries shall be: for dioxin-lik e PCBs 20 % to 60 %, for PCDD/Fs 50 % 
to 130 % (range s apply for the TCDD calibration curve). As the contr ibution of dioxin-like PCBs to the sum 
of PCDD/Fs and dioxin-like PCBs can vary between different matr ices and samples , bioassa y apparent 
reco veries for the sum of PCDD/Fs and dioxin-like PCBs reflect these rang es and shall be between 30 % and 
130 %. Any implication of substantially revised TEF values for the Union legislation for PCDD/Fs and 
dioxin-like PCBs requires the revision of these rang es. 4.5.2017 L 115/34 Official Jour nal of the European Union EN     
(1)Current requirements are based on the TEFs published in: M. Van den Berg et al, Toxicol Sci 93 (2), 223–241 (2006). 7.1.5.  Control of reco ver ies for clean-up 
The loss of compo unds during the clean-up shall be check ed during validation. A blank sample spiked with 
a mixture of the diffe rent cong eners shall be submitted to clean-up (at least n = 3) and the reco very and 
variability check ed by a confir matory method. The reco very shall be within 60 % to 120 % especially for 
cong eners contr ibuting more than 10 % to the TEQ-level in various mixtures. 
7.1.6.  Repor ting limit 
When repor ting BEQ levels, a repor ting limit shall be determi ned from relevant matr ix sam ples involving 
typical cong ener patt erns, but not from the calibration curve of the standards due to low precision in the 
lower rang e of the curve. Effects from extraction and clean-up shall be taken into account. The repor ting 
limit shall be set at least by a factor of three above the procedure blanks. 
7.2.  Use of reference samples 
7.2.1.  Reference samples shall represent sample matr ix, congener patt erns and concentration rang es for PCDD/Fs 
and dioxin-like PCBs around the maximum level or action threshold. 
7.2.2.  A matr ix blank, and where it is not possible, a procedure blank, and a reference sam ple at the maximum 
level or action threshold shall be included in each test series. These samples shall be extract ed and tested at 
the same time under identical conditions. The referen ce sam ple shall show a clearly elevat ed response in 
compari son to the blank sam ple, thus ensur ing the suitability of the test. Those samples may be used for 
blank and reco very corrections. 
7.2.3.  Reference samples chosen to perform a reco very correction shall be representative for the test sam ples, 
meaning that cong ener patter ns may not lead to an underestimation of levels. 
7.2.4.  Extra reference samples at e.g. 0,5x and 2x the maximum level or action threshold may be included to 
demonstrate the proper performa nce of the test in the range of interest for the control of the maximum 
level or action threshold. Combined, these samples may be used for calculating the BEQ levels in test 
samples (see point 7.1.2.2). 
7.3.  Deter mination of cut-of f values 
The relationship between bioanalytical results in BEQ and results from the conf irmatory method in TEQ 
shall be established, for example by matr ix-matc hed calibration exper iments, involving reference sam ples 
spik ed at 0, 0,5x, 1x and 2x the ML, with 6 repetitions on each level (n = 24). Correction factors (blank and 
reco very) may be estimated from this relationship but shall be check ed in accordance with point 7.2.2. 
Cut-off values shall be established for decisions over sam ple compliance with maximum levels or for the 
control of action thresholds, if relevant, with the respective maximum levels or action threshold set for 
either PCDD/Fs and dioxin-like PCBs alone, or for the sum of PCDD/Fs and dioxin-like PCBs. They are 
represente d by the lower end-point of the distr ibution of bioanalytical results (cor recte d for blank and 
reco very) corresponding to the decision limit of the confir matory method based on a 95 % level of 
conf idence, implying a false-com pliant rate < 5 %, and on a RSDR < 25 %. The decision limit of the 
conf irmatory method is the maximum level, taking into account the expanded measurement uncer tainty . 4.5.2017 L 115/35 Official Jour nal of the European Union EN     The cut-off value (in BEQ) may be calculated in accordance with one of the approaches set out in 
points 7.3.1, 7.3.2 and 7.3.3. (see Figure 1). 
7.3.1.  Use of the lower band of the 95 % prediction interval at the decision limit of the conf irmat ory method: 
Cut-off value ¼BEQ DL − s y,x�t α,f¼m − 2ffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi ffi
1=nþ1=m þðxi − xÞ2=Q xxq
with: 
BEQDL  BEQ corresponding to the decision limit of the confir matory method, being the maximum 
level taking into account the expanded measurement uncer tainty 
sy,x  residual standard deviation 
t α,f = m – 2  student factor (α = 5 %, f = degrees of freedom, sing le-sided) 
m  total number of calibration points (index j) 
n  number of repetitions on each level 
xi  sample concentration (in TEQ) of calibration point i determined by a confir matory method 
x  mean of the concentrations (in TEQ) of all calibration sam ples 
Qxx¼Xm
j¼1ðxi − xÞ2  square sum parameter , i = index for calibration point i 
7.3.2.  Calculation from bioanalytical results (cor rected for blank and reco very) of multiple analyses of sam ples 
(n ≥ 6) contaminated at the decision limit of the conf irmat ory method, as the lower endpoint of the data 
distr ibution at the corresponding mean BEQ value: 
Cut-off value = BEQDL – 1,64 × SDR 
with: 
SDR  standard deviation of bioassa y results at BEQDL, measured under within-laboratory reproducibility 
conditions 
7.3.3.  Calculation as mean value of bioanalytical results (in BEQ, corrected for blank and reco very) from multiple 
analysis of samples (n ≥ 6) contaminate d at two- thirds of the maximum level or action threshold, based on 
the obser vation that this level will be around the cut-off value deter mined under point 7.3.1 or point 7.3.2: 
Calculation of cut-off values based on a 95 % level of conf idence implying a false-com pliant rate < 5 %, and 
a RSDR < 25 %:  
(1) from the lower band of the 95 % prediction interval at the decision limit of the conf irmatory method.  
(2) from multiple analysis of samples (n ≥ 6) contaminated at the decision limit of the conf irmatory 
method as the lower end-point of the data distr ibution (represented in the figure by a bell-shaped curve) 
at the corresponding mean BEQ value. 4.5.2017 L 115/36 Official Jour nal of the European Union EN     Figure 1 
7.3.4.  Restr ictions to cut-off values 
BEQ-based cut-off values calculated from the RSDR achi eved during validation using a limited number of 
samples with diffe rent matr ix/congener patt erns may be higher than the TEQ-based maximum levels or 
action thresholds due to a better precision than attainable in routine when an unkno wn spectr um of 
possible congener patter ns has to be controlled. In such cases, cut-off values shall be calculated from an 
RSDR = 25 %, or two- thirds of the maximum level or action threshold shall be pref erred. 
7.4.  Performance character istics 
7.4.1.  Since no internal standards can be used in bioanalytical methods, tests on the repeatability of bioanalytical 
methods shall be carried out to obtain information on the standard deviation within and between test 
series. Repeatability shall be belo w 20 % and intra-laboratory reproducibility shall be belo w 25 %. This shall 
be based on the calculat ed levels in BEQ after blank and reco very correction. 
7.4.2.  As part of the validation process, the test shall be shown to discr iminate between a blank sam ple and 
a level at the cut-off value, allowing the identification of samples above the corresponding cut-off value (see 
point 7.1.2). 
7.4.3.  Target compounds, possible interferences and maximum tolerable blank levels shall be defined. 
7.4.4.  The percent standard deviation in the response or concentration calculat ed from the response (only possible 
in working rang e) of a triplicate determination of a sample extract may not be above 15 %. 
7.4.5.  The uncor rected results of the reference sample (s) expressed in BEQ (blank and at the maximum level or 
action threshold) shall be used for evaluation of the perf ormance of the bioanalytical method over 
a constant time period. 
7.4.6.  Quality control charts for procedure blanks and each type of reference sample shall be recorded and 
check ed to mak e sure the analytical performa nce is in accordance with the requirements, in particular for 
the procedure blanks with regar d to the request ed minimum difference to the lower end of the working 
range and for the reference samples with regard to within-laboratory reproducibility . Procedure blanks shall 
be controlled in a manner to avoid false-compliant results when subtracted . 4.5.2017 L 115/37 Official Jour nal of the European Union EN     7.4.7.  The results from the confir matory methods of suspect ed samples and 2 to 10 % of the compliant samples 
(minimum of 20 samples per matr ix) shall be collect ed and used to evaluate the performa nce of the 
screening method and the relationship between BEQ and TEQ. This database may be used for the re- 
evaluation of cut-off values applicable to routine samples for the validat ed matr ices. 
7.4.8.  Successful method performa nce may also be demonstrated by participation in ring trials. The results from 
samples analysed in ring trials, covering a concentration rang e up to e.g. 2 × maximum level, may be 
included in the evaluation of the false-compliant rate, if a laboratory is able to demonstrate its successful 
performa nce. The samples shall cover most frequent cong ener patte rns, representing various sources. 
7.4.9.  Dur ing incidents, the cut-off values may be re-evaluat ed, reflecting the specifi c matr ix and congener 
patter ns of this sing le incident. 
8.  Repor ting of the results 
8.1.  Confir mator y methods 
8.1.1.  The analytical results shall contain the levels of the individual PCDD/F and dioxin-lik e PCB congeners and 
TEQ-values shall be repor ted as lower-bound, upper -bound and medium-bound in order to include 
a maximum of information in the repor ting of the results and thereby enabling the interpr etation of the 
results according to specifi c requirements. 
8.1.2.  The repor t shall include the method used for extraction of PCDD/Fs and dioxin-like PCBs. 
8.1.3.  The reco veries of the individual internal standards shall be made available in case the reco veries are outside 
the range refer red to in point 6.2.5, in case the maximum level is exceeded (in this case, the reco veries for 
one of the two duplicat e analysis) and in other cases upon request. 
8.1.4.  As the expanded measurement uncer tainty is to be take n into account when deciding about the compl iance 
of a sample, this paramete r shall be made available. Thus, analytical results shall be repor ted as 
x +/– U whereby x is the analytical result and U is the expanded measurement uncer tainty using a coverag e 
factor of 2 whic h gives a level of confidence of appro ximately 95 %. In the case of a separate determi nation 
of PCDD/Fs and dioxin-like -PCBs, the sum of the estimate d expanded uncer tainty of the separate analytical 
results of PCDD/Fs and dioxin-like PCBs shall be used for the sum of PCDD/Fs and dioxin-like PCBs. 
8.1.5.  The results shall be expressed in the same units and with at least the same number of significant figures as 
the maximum levels laid down by Directive 2002/32/EC 
8.2.  Bioanalytical screening methods 
8.2.1.  The result of the screening shall be expressed as “com pliant” or “suspected to be non-com pliant” 
(“suspected ”). 
8.2.2.  In addition, an indicative result for PCDD/Fs and/or dioxin-like PCBs expressed in BEQ, and not TEQ, may 
be given. 
8.2.3.  Samples with a response belo w the repor ting limit shall be expressed as “lower than the repor ting limit”. 
Samples with a response above the working range shall be repor ted as “exceeding the working rang e” and 
the level corresponding to the upper end of the working range shall be given in BEQ. 
8.2.4.  For each type of samp le matr ix, the repor t shall mention the maximum level or action threshold on whic h 
the evaluation is based. 
8.2.5.  The repor t shall mention the type of the test applied, the basic test principle and the kind of calibration. 4.5.2017 L 115/38 Official Jour nal of the European Union EN     8.2.6.  The repor t shall include the method used for extraction of PCDD/Fs and dioxin-like PCBs. 
8.2.7.  In case of samp les suspecte d to be non-com pliant, the repor t needs to include a note on the action to be 
taken. The concentration of PCDD/Fs and the sum of PCDD/Fs and dioxin-like PCBs in those samples with 
elevated levels has to be determined/confir med by a confir matory method. 
8.2.8.  Non-complia nt results shall only be repor ted from conf irmatory analysis. 
8.3.  Physico -chemical screeni ng methods 
8.3.1.  The result of the screening shall be expressed as “com pliant” or “suspected to be non-com pliant” 
(“suspected ”). 
8.3.2.  For each type of samp le matr ix, the repor t shall mention the maximum level or action threshold on whic h 
the evaluation is based. 
8.3.3.  In addition, levels for individual PCDD/F and/or dioxin-like PCB cong eners and TEQ-values repor ted as 
lower-bound, upper -bound and medium-bound may be given. The results shall be expressed in the same 
units and with at least the same number of significant figures as the maximum levels laid down by 
Directive 2002/32/EC. 
8.3.4.  The reco veries of the individual internal standards shall be made available in case the reco veries are outside 
the range refer red to in point 6.2.5, in case the maximum level is exceeded (in this case, the reco veries for 
one of the two duplicat e analysis) and in other cases upon request. 
8.3.5.  The repor t shall mention the GC-MS method applied. 
8.3.6.  The repor t shall include the method used for extraction of PCDD/Fs and dioxin-like PCBs. 
8.3.7.  In case of samp les suspecte d to be non-com pliant, the repor t needs to include a note on the action to be 
taken. The concentration of PCDD/Fs and the sum of PCDD/Fs and dioxin-like PCBs in those samples with 
elevated levels has to be determined/confir med by a confir matory method. 
8.3.8.  Non-complia nce can only be decided after confir matory analysis. 
CHAPTER III 
Sample prepar ation and requir ements for met hods of analy sis used in offical control of the levels of 
non dioxin-like PCBs in feed 
1.  Field of application 
The requirements set out in this Chapt er shall be applied where feed is analysed for the official control of 
the levels of non-dio xin-like PCBs and as regar ds sample preparation and analytical requirements for other 
regulatory purposes, which includes the controls performed by the feed business operator to ensure 
compl iance with the provisions of Regulation (EC) No 183/2005. 
2.  Applicable detection methods 
Gas chromat ography/Electron Cap ture Detection (GC-ECD), GC-LRMS, GC-MS/MS, GC-HRMS or equivalent 
methods. 4.5.2017 L 115/39 Official Jour nal of the European Union EN     3.  Identif ication and conf irmation of analytes of interes t 
3.1.  Relative retention time in relation to internal standards or reference standards (acceptable deviation of 
+/– 0,25 %). 
3.2.  Gas chromat ographic separation of the non-dio xin-like PCBs from interfering substances, especially co- 
eluting PCBs, in particular if levels of samples are in the range of legal limits and non-com pliance is to be 
conf irmed (1). 
3.3.  Requirements for GC-MS techniques 
Monitoring of at least the following number of molecular ions or charact eristic ions from the molecular 
cluster: 
(a)  two specifi c ions for HRMS; 
(b)  three specific ions for LRMS; 
(c)  two specifi c precursor ions, each with one specifi c corresponding transition product ion for for MS-MS. 
Maximum permitted tolerances for abundance ratios for selected mass fragments: 
Relative deviation of abundance ratio of selected mass fragments from theoretical abundance or calibration 
standard for target ion (most abundant ion monitored) and qualifi er ion(s): ± 15 % 
3.4.  Requirements for GC-ECD techniques 
Results exceeding the maximum level shall be confir med with two GC columns with stationar y phases of 
differ ent polar ity. 
4.  Demonstration of perfor mance of method 
The perf ormance of the method shall be validat ed in the range of the maximum level (0,5 to 2 times the 
maximum level) with an accepta ble coefficient of variation for repeated analysis (see requirements for 
interm ediate precision in point (9). 
5.  Limit of quantif ication 
The sum of the LOQs (2) of non-dio xin-like PCBs shall not be higher than one-third of the maximum 
level (3). 
6.  Quality control 
Regular blank controls, analysis of spik ed sam ples, quality control sam ples, participation in inter-laborat ory 
studies on relevant matr ices. 
7.  Control of reco veries 
7.1.  Suitable internal standards with physico-chem ical proper ties compar able to analytes of interest shall be 
used. 4.5.2017 L 115/40 Official Jour nal of the European Union EN     
(1)Cong eners often found to co-elute are for exam ple PCB 28/31, PCB 52/69 and PCB 138/163/164. For GC-MS also possible interferences 
from fragments of higher chlorinated cong eners shall be considered. 
(2)The principles as descr ibed in the “Guidance Document on the Estimation of LOD and LOQ for Measurements in the Field of 
Contaminants in Feed and Food” (http://ec.europa.eu/f ood/safety/animal-f eed_en) shall be followed when applicable. 
(3)It is highly recommendable to have a lower contr ibution of the reagent blank level to the level of a contaminant in a sample. It is in the re­
sponsibility of the laborato ry to control the variation of blank levels, in particular , if the blank levels are subtracted. 7.2.  Addition of internal standards: 
Addition to products (before extraction and clean-up process). 
7.3.  Requirements for methods using all six isotope -labelled non-dio xin-like PCB cong eners 
(a)  results shall be corrected for reco veries of internal standards; 
(b)  reco veries of isotope -labelled internal standards shall be between 60 and 120 %; 
(c)  lower or higher reco veries for individual cong eners with a contr ibution to the sum of non-dio xin-like 
PCBs belo w 10 % are acceptable. 
7.4.  Requirements for methods using not all six isotope -labelled internal standards or other internal standards: 
(a)  reco very of internal standard(s) shall be controlled for ever y sample ; 
(b)  reco veries of internal standard(s) shall be between 60 and 120 %; 
(c)  results shall be corrected for reco veries of internal standards. 
7.5.  The reco veries of unlabelled cong eners shall be check ed by spik ed sam ples or quality control samples with 
concentrations in the range of the maximum level. Reco veries for these cong eners shall be considered 
acceptable, if they are between 60 and 120 %. 
8.  Requirements for laborator ies 
In accordance with the provisions of Regulation (EC) No 882/2004, laboratories shall be accredit ed by 
a recognised body operating in accordance with ISO Guide 58 to ensure that they are applying analytical 
quality assurance. Laboratories shall be accredited following the EN ISO/IEC 17025 standard. In addition, 
the principles as descr ibed in Techn ical Guidelines for the estimation of measurement uncer tainty and limits 
of quantifi cation for PCB analysis shall be followed when applicable (1). 
9.  Perfor mance character istics: criter ia for the sum of non-dio xin-like PCBs at the maximum level  
Isotope dilution mass spec ­
trometr y(1) Other techniques 
Trueness – 20 to + 20 % – 30 to + 30 % 
Interm ediat e precision (RSD %) ≤ 15 % ≤ 20 % 
Diffe rence between upper and lower-bound calcula ­
tion ≤ 20 % ≤ 20 % 
(1)  Use of all six 13C-labelled analogues as inter nal standards required.  
10.  Repor ting of the results 
10.1.  The analytical results shall contain the levels of the individual non-dio xin-like PCBs and the sum of those 
PCB congenersrepor ted as lower-bound, upper -bound and medium-bound in order to include a maximum 
of information in the repor ting of the results and thereby enabling the interpretation of the results 
according to specif ic requirements. 4.5.2017 L 115/41 Official Jour nal of the European Union EN     
(1)Current requirements are based on the TEFs published in: M. Van den Berg et al, Toxicol Sci 93(2), 223–241 (2006). 10.2.  The repor t shall include the method used for the extraction of PCBs. 
10.3.  The reco veries of the individual internal standards shall be made available in case the reco veries are outside 
the range refer red to in point 7, in case the maximum level is excee ded and in other cases upon request. 
10.4.  As the expanded measurement uncer tainty is to be take n into account when deciding about the compl iance 
of a sample, that parameter shall also be made available. Thus, analytical results shall be repor ted as 
x +/– U whereby x is the analytical result and U is the expanded measurement uncer tainty using a coverag e 
factor of 2 which gives a level of confidence of appro ximat ely 95 %. 
10.5.  The results shall be expressed in the same units and with at least the same number of significant figures as 
the maximum levels laid down by Directive 2002/32/EC.’  4.5.2017 L 115/42 Official Jour nal of the European Union EN