ANNEX IICRITERIA FOR SAMPLE PREPARATION AND FOR METHODS OF ANALYSIS USED FOR THE OFFICIAL CONTROL OF THE LEVELS OF MYCOTOXINS IN FOODSTUFFS
4.METHOD OF ANALYSIS TO BE USED BY THE LABORATORY AND LABORATORY CONTROL REQUIREMENTS
4.1.Definitions
A number of the most commonly used definitions that the laboratory shall be required to use are the following:
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Repeatability, the value below which the absolute difference between two single test results obtained under repeatability conditions, namely same sample, same operator, same apparatus, same laboratory, and short interval of time may be expected to lie within a specific probability (typically 95 %) and hence r = 2,8 × sr.
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Standard deviation, calculated from results generated under repeatability conditions.
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Relative standard deviation, calculated from results generated under repeatability conditions [(sr / 
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Reproducibility, the value below which the absolute difference between single test results obtained under reproducibility conditions, namely on identical material obtained by operators in different laboratories, using the standardised test method may be expected to lie within a certain probability (typically 95 %); R = 2,8 × sR.
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Standard deviation, calculated from results under reproducibility conditions.
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Relative standard deviation calculated from results generated under reproducibility conditions [(sR / 
4.2.General requirements
Methods of analysis used for food control purposes shall comply with the provisions of items 1 and 2 of Annex III to Regulation (EC) No 882/2004.
4.3.Specific requirements
4.3.1.Performance criteria
Where no specific methods for the determination of mycotoxin levels in foodstuffs are required by Community legislation, laboratories may select any method provided the selected method meets the following criteria:
Performance criteria for aflatoxins
| Note: | |||
| |||
| Criterion | Concentration Range | Recommended Value | Maximum permitted Value |
|---|---|---|---|
| Blanks | All | Negligible | — |
| Recovery — Aflatoxin M1 | 0,01-0,05 μg/kg | 60 to 120 % | |
| > 0,05 μg/kg | 70 to 110 % | ||
| Recovery — Aflatoxins B1, B2, G1, G2 | < 1,0 μg/kg | 50 to 120 % | |
| 1-10 μg/kg | 70 to 110 % | ||
| > 10 μg/kg | 80 to 110 % | ||
| Precision RSDR | All | As derived from Horwitz Equation | 2 × value derived from Horwitz Equation |
| Precision RSDr may be calculated as 0,66 times Precision RSDR at the concentration of interest. | |||
Performance criteria for ochratoxin A
| Level μg/kg | Ochratoxin A | ||
|---|---|---|---|
| RSDr % | RSDR % | Recovery % | |
| < 1 | ≤ 40 | ≤ 60 | 50 to 120 |
| 1-10 | ≤ 20 | ≤ 30 | 70 to 110 |
Performance criteria for patulin
| Level μg/kg | Patulin | ||
|---|---|---|---|
| RSDr % | RSDR % | Recovery % | |
| < 20 | ≤ 30 | ≤ 40 | 50 to 120 |
| 20-50 | ≤ 20 | ≤ 30 | 70 to 105 |
| > 50 | ≤ 15 | ≤ 25 | 75 to 105 |
Performance criteria for deoxynivalenol
| Level μg/kg | Deoxynivalenol | ||
|---|---|---|---|
| RSDr % | RSDR % | Recovery % | |
| > 100-≤ 500 | ≤ 20 | ≤ 40 | 60 to 110 |
| > 500 | ≤ 20 | ≤ 40 | 70 to 120 |
Performance criteria for zearalenone
| Level μg/kg | Zearalenone | ||
|---|---|---|---|
| RSDr % | RSDR % | Recovery % | |
| ≤ 50 | ≤ 40 | ≤ 50 | 60 to 120 |
| > 50 | ≤ 25 | ≤ 40 | 70 to 120 |
Performance criteria for Fumonisin B1 and B2
| Level μg/kg | Fumonisin B1 or B2 | ||
|---|---|---|---|
| RSDr % | RSDR % | Recovery % | |
| ≤ 500 | ≤ 30 | ≤ 60 | 60 to 120 |
| > 500 | ≤ 20 | ≤ 30 | 70 to 110 |
Performance criteria for T-2 and HT-2 toxin
| Level μg/kg | T-2 toxin | ||
|---|---|---|---|
| RSDr % | RSDR % | Recovery % | |
| 50-250 | ≤ 40 | ≤ 60 | 60 to 130 |
| > 250 | ≤ 30 | ≤ 50 | 60 to 130 |
| Level μg/kg | HT-2 toxin | ||
|---|---|---|---|
| RSDr % | RSDR % | Recovery % | |
| 100-200 | ≤ 40 | ≤ 60 | 60 to 130 |
| > 200 | ≤ 30 | ≤ 50 | 60 to 130 |
Notes to the performance criteria for the mycotoxins
The detection limits of the methods used are not stated as the precision values are given at the concentrations of interest
The precision values are calculated from the Horwitz equation, i.e.:
RSDR = 2(1-0,5logC)
where:
This is a generalised precision equation which has been found to be independent of analyte and matrix but solely dependent on concentration for most routine methods of analysis.
4.3.2.‘Fitness-for-purpose’ approach
In the case where there are a limited number of fully validated methods of analysis, alternatively, a ‘fitness-for-purpose’ approach, defining a single parameter, a fitness function, to evaluate the acceptability of methods of analysis may be used. A fitness function is an uncertainty function that specifies maximum levels of uncertainty regarded as fit for purpose.
Given the limited number of methods of analysis, fully validated by a collaborative trial, especially for the determination of T-2 and HT-2 toxin, the uncertainty function approach, specifying the maximum acceptable uncertainty, may also be used to assess the suitability (the ‘fitness-for-purpose’) of the method of analysis to be used by the laboratory. The laboratory may use a method which produces results within the maximum standard uncertainty. The maximum standard uncertainty may be calculated using the following formula:
where:
Uf is the maximum standard uncertainty (μg/kg)
LOD is the limit of detection of the method (μg/kg)
α is a constant, numeric factor to be used depending on the value of C. The values to be used are set out in the table hereafter
C is the concentration of interest (μg/kg).
If the analytical method provides results with uncertainty measurements less than the maximum standard uncertainty the method shall be considered being equally suitable to one which meets the performance criteria given in point 4.3.1.
| Table | |
| Numeric values to be used for α as constant in formula set out in this point, depending on the concentration of interest | |
| C (μg/kg) | α |
|---|---|
| ≤ 50 | 0,2 |
| 51-500 | 0,18 |
| 501-1 000 | 0,15 |
| 1 001-10 000 | 0,12 |
| > 10 000 | 0,1 |
4.4.Estimation of measurement uncertainty, recovery calculation and reporting of results(1)
The analytical result must be reported corrected or uncorrected for recovery. The manner of reporting and the level of recovery must be reported. The analytical result corrected for recovery shall be used for controlling compliance.
The analytical result must be reported as x +/– U whereby x is the analytical result and U is the expanded measurement uncertainty.
U is the expanded measurement uncertainty, using a coverage factor of 2 which gives a level of confidence of approximately 95 %.
For food of animal origin, the taking into account of the measurement uncertainty can also be done by establishing the decision limit (CCα) in accordance with Commission Decision 2002/657/EC(2) (point 3.1.2.5. of the Annex — the case of substances with established permitted limit).
The present interpretation rules of the analytical result in view of acceptance or rejection of the lot apply to the analytical result obtained on the sample for official control. In case of analysis for defence or referee purposes, the national rules apply.
4.5.Laboratory quality standards
Laboratory must comply with the provisions of Article 12 of Regulation (EC) No 882/2004 on official controls performed to ensure the verification of compliance with feed and food law, animal health and animal welfare rules(3).
More details on procedures for the estimation of measurement uncertainty and on procedures for assessing recovery can be found in the report ‘Report on the relationship between analytical results, measurement uncertainty, recovery factors and the provisions of EU food and feed legislation’ — http://europa.eu.int/comm/food/food/chemicalsafety/contaminants/report-sampling_analysis_2004_en.pdf
OJ L 221, 17.8.2002, p. 8. Decision as last amended by Decision 2004/25/EC (OJ L 6, 10.1.2004, p. 38).
See also the transitional arrangements provided for in article 18 of Commission Regulation (EC) No 2076/2005 of 5 December 2005 laying down transitional arrangements for the implementation of Regulation (EC) No 853/2004, 854/2004 and 882/2004 of the European Parliament and of the Council and amending Regulations (EC) No 853/2004 and 854/2004 (OJ L 338, 22.12.2005, p. 83).