Commission Regulation (EC) No 765/2002 of 3 May 2002 on the collection of samples and the adoption of certain detailed rules in connection with physical checks on boneless beef cuts qualifying for export refunds
Date: 2002-05-03
Institutions: European Commission
Subjects: beef, boned meat, customs inspection, export refund, food inspection, labelling

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COMMISSION REGULATION (EC) No 765/2002

of 3 May 2002

on the collection of samples and the adoption of certain detailed rules in connection with physical
checks on boneless beef cuts qualifying for export refunds

THE COMMISSION OF THE EUROPEAN COMMUNITIES,

Having regard to the Treaty establishing the European
Community,

Having regard to Council Regulation (EC) No 1254/1999 of 17
May 1999 on the common organisation of the market in beef
and veal (1), as last amended by Commission Regulation (EC)
No 2345/2001 (2), and in particular Article 33(12) thereof,

Having regard to Council Regulation (EEC) No 386/90 of 12
February 1990 on the monitoring carried out at the time of
export of agricultural products receiving refunds or other
amounts (3), as amended by Regulation (EC) No 163/94 (4), and
in particular Article 6 thereof,

Whereas:

(1)

(2)

(3)

In accordance with Article 33 of Regulation (EC) No
1254/1999, the difference between the prices for the
products listed in Article 1 of that Regulation on the
world market and within the Community can be
covered by an export refund. For agricultural products,
the arrangements governing the system are set out in
Commission Regulation (EC) No 800/1999 of 15 April
the
1999 laying down common detailed rules for
application of the system of export refunds on agricul-
tural products (5), as last amended by Regulation (EC) No
2299/2001 (6).

Sector 5 of Annex I to Commission Regulation (EEC) No
3846/87 of 17 December 1987 establishing an agricul-
tural product nomenclature for export refunds (7), as last
amended by Regulation (EC) No 488/2002 (8), provides,
inter alia,
that refunds are to be granted on certain
boneless cuts on condition they have a minimum lean
bovine meat content and, in the case of cuts of adult
male bovine animals, are individually wrapped.

Commission Regulation (EC) No 2221/95 (9), as last
amended by Regulation (EC) No 2655/1999 (10),
lays
down detailed rules as regards physical checks carried
out at the time of export of agricultural products quali-

(1) OJ L 160, 26.6.1999, p. 1.
(2) OJ L 315, 1.12.2001, p. 29.
(3) OJ L 42, 16.2.1990, p. 6.
(4) OJ L 24, 29.1.1994, p. 2.
(5) OJ L 102, 17.4.1999, p. 11.
(6) OJ L 308, 27.11.2001, p. 19.
(7) OJ L 366, 24.12.1987, p. 1.
(8) OJ L 76, 19.3.2002, p. 11.
(9) OJ L 224, 21.9.1995, p. 13.
(10) OJ L 325, 17.12.1999, p. 12.

fying for refunds. Special conditions regarding beef are
laid down in Commission Regulation (EC) No 2457/97
of 10 December 1997 on the collection of samples in
connection with physical checks of boneless beef cuts
qualifying for export refunds (11).

(4)

(5)

(6)

Provision should be made for checks on the provenance/
origin of boneless cuts of adult male bovine animals and
the methodology to be applied for those checks, with a
view to standardisation, and to setting appropriate
penalties in the event of failure to comply with the
condition regarding provenance/origin. Regulation (EC)
No 2457/97 should, in addition, be updated in order to
take account of changes to the nomenclature applicable
to export refunds on agricultural products provided for
in Regulation (EEC) No 3846/87, as amended by Regula-
tion (EC) No 2556/2001 (12).

In the interests of clarity, Regulation (EC) No 2457/97
should therefore be repealed and replaced.

The measures provided for in this Regulation are in
accordance with the opinion of
the Management
Committee for Beef and Veal,

HAS ADOPTED THIS REGULATION:

Article 1

This Regulation shall apply in the event of physical
1.
checks on the nature and characteristics of products within the
meaning of Article 2(a) of Regulation (EEC) No 386/90
regarding:

(a) compliance with the obligation on the individual wrapping
of boneless cuts covered by the following product codes:

— 0201 30 00 9100,

— 0201 30 00 9120;

(b) the origin of boneless cuts of adult male bovine animals

covered by the following product codes:

— 0201 30 00 9100,

— 0201 30 00 9120;

(11) OJ L 340, 11.12.1997, p. 29.
(12) OJ L 348, 31.12.2001, p. 1.

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(c) the minimum average lean meat content of boneless cuts

covered by the following product codes:
— 0201 30 00 9100,
— 0201 30 00 9120,
— 0201 30 00 9060,
— 0202 30 90 9200.

2.
The description of the products referred to in paragraph
1 shall be that contained in the nomenclature of agricultural
products for export refunds set out in Sector 5 of Annex I to
Regulation (EEC) No 3846/87.

Article 2

1.
For the purposes of physical checks, samples shall consist
of two full cartons collected from two different parts of the
consignment. The first carton shall be intended for the authori-
ties responsible for the checks while the second shall be kept in
reserve under the supervision of the customs authorities.

The quantity of products covered by one of the following

2.
declarations shall be deemed to comprise a consignment:
(a) a declaration as referred to in Article 5(1) of Regulation

(EC) No 800/1999;

(b) a declaration as referred to in Article 26(1) of Regulation
in the situation referred to in that

(EC) No 800/1999,
paragraph, as regards storage only.

Article 3

For the purposes of checks on compliance with the obligation
referred to in Article 1(1)(a),
the customs authorities shall
check that each cut in the first carton of the sample referred to
in Article 2 is individually wrapped and that each package does
not contain more than one cut. If this is not the case, the same
checks shall be conducted on the second carton.

Where, taking the two cartons together, at most one cut turns
out not to have been wrapped individually or one package to
the other conditions
contain more than one cut and all
regarding the refund are met, the consignment shall be deemed
not to be irregular. If these conditions are not observed, an
irregularity shall be deemed to have occurred.

Where an irregularity is established, the refund payable in
respect of the consignment shall be calculated on the basis of
the corrected weight. The corrected weight shall be obtained by
reducing the declared net weight by a percentage corre-

sponding to the weight of cuts failing to comply in relation to
the total net weight of the sample.

Article 4

For the purposes of checks on the condition regarding origin
referred to in Article 1(1)(b), the sample for analysis shall
consist of one or two cuts collected at random from the first
carton of the sample referred to in Article 2. Where analysis
shows that meat other than beef
from adult male bovine
animals is present, no refund shall be granted.

The checks shall be carried out in accordance with the meth-
odology described in the Annex hereto.

Without prejudice to further checks decided on where an irreg-
ularity is suspected, the checks shall be carried out at random
covering all export operations on not less than one third of
operations selected for the physical check.

Article 5

For the purpose of checks on compliance with the condition
referred to in Article 1(1)(c), the whole contents of the first
carton of the sample referred to in Article 2 shall be minced to
form a homogeneous mixture. Where the lean meat content of
the sample is below the minimum laid down, the contents of
the second carton shall be examined in the same way. Where
the average for the two cartons is below the average lean meat
content laid down, no refund shall be granted.

Article 6

In accordance with Article 68 and without prejudice to Article
78 of Council Regulation (EEC) No 2913/92 (1), samples shall
be collected and checked as provided for in this Regulation at
the time declarations, as referred to in Article 2(2), that have
been accepted are verified.

Regulation (EC) No 2457/97 is hereby repealed.

Article 7

Article 8

This Regulation shall enter into force on 1 July 2002.

It shall apply to operations covered by a declaration as referred
to in Article 2(2) covered with effect from 1 July 2002.

This Regulation shall be binding in its entirety and directly applicable in all Member States.

Done at Brussels, 3 May 2002.

For the Commission

Franz FISCHLER

Member of the Commission

(1) OJ L 302, 19.10.1992, p. 1.

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ANNEX

ANALYSIS OF THE ORIGIN OF CERTAIN BONELESS BEEF CUTS OF ADULT MALE BOVINE ANIMALS

Methodology to be applied for beef sexing

The methodology to be applied is based on polymerase chain reaction (PCR) and comprises sampling, extraction of DNA,
PCR and gel electrophoresis.

1. Sampling and sub-sampling

The sub-sample shall be dissected from a core part (inside) of the provided meat piece, by using a sterile (*) knife
(scalpel or similar). This sample shall then be ground by using a micro grinder or cut into smaller pieces to assure for a
reasonable extraction efficiency.

Samples have to be prepared in a different working place than the one used to perform PCR. The handling of material
has to be performed in an environment which can be cleaned easily, preferably on a clean bench, to ensure that no
cross contamination with other samples occurs.

Sterile (*) blades, scalpels or similar tools are to be used for preparation of the meat sample.

2. Extraction and purification of DNA

DNA extraction and purification has to be accomplished by either using conventional procedures (1), ready-to-use kits
(based on the principle: solubilisation of the meat sample in lysis buffer which contain surfactants, detergents and
proteinase K, application of the solubilised sample to a DNA-binding resin, removal of non-DNA compounds by
repeated washing steps, and finally elution of purified DNA in water or low-salt buffer), or extracting the DNA in
sodium hydroxide solution (2).

The control of the successful extraction by gel electrophoresis is recommended, but not mandatory.

Validation aspect: for each batch of samples to be extracted, one extraction control (i.e. no meat) shall be performed in
parallel to prove the integrity of the applied procedure.

3. Polymerase chain reaction (PCR)

Principle: the principle of PCR is a three-step procedure (denaturation, annealing of primers, extension) which has to
be repeated for some 25 to 40 times (number of ‘cycles’
in the method). The reagents (reaction buffer, MgCl
2,
deoxynucleotides, primers, heat-stable DNA-polymerase, sterile water) are mixed together according to the developed
method, leading the ‘mastermix’. Dedicated pipettes shall be used for the preparation of the mastermix. This mastermix
is then added to the DNA template (extracted DNA). The reaction is performed in a thermo-cycler. After completion,
the PCR products are analysed by gel electrophoresis or stored at 4 °C or at – 20 °C.

The recommended (3) method to be applied, referring to the template, has to either amplify a sequence within the
amelogenin locus (homologous gene) or within the ZFX/Y region (allele-specific PCR).

Specific primers for these two types of methods are:

Amelogenin forward: 5'-CAGCCAAACCTCCCTCTGC-3'
Amelogenin reverse: 5'-CCCGCTTGGTCTTGTCTGTTGC-3'

(Ennis, S., and Gallagher, T.F. (1994) Anim. Genet. 25: 425-427)

Amelogenin forward: 5'-AAATTCTCTCACAGTCCAAG-3'
Amelogenin reverse: 5'-CAACAGGTAATTTTCCTTTAG-3'

(Chen, C.M., Hu, C.L., Wang, C.H., Hung, C.M., Wu, H.K., Choo, K.B., and Cheng, W.T.K. (1999) Mol. Reprod. Dev. 54, 209-214)

ZFX (allele-specific), forward: 5'-GACAGCTGAACAAGTGTTACTG-3'
ZFX (allele-specific), reverse: 5'-AATGTCACACTTGAATCGCATC-3'
ZFY (allele-specific), forward: 5'-GAAGGCCTTCGAATGTGATAAC-3'
ZFY (allele-specific), reverse: 5'-CTGACAAAAGGTGGCGATTTCA-3'

(Kirkpatrick, B.W., and Monson, R.L. (1993) J. Reprod. Fertil. 98: 335-340)

ZFX forward: 5'-AGCTGAACAAGGGTTACTG-3'
ZFY forward: 5'-CAAGCTTACCAGCAAGTCA-3'
ZFX/Y reverse: 5'-CCAGTATGGATTCGCATGT-3'

(Zinovieva, N., Palma, G., Müller, M., and Brem G. (1995) Theriogenology 43: 265)

(*) Not contaminated with DNA.
(1) As described in: Sambrook, J., Fritsch, E.F., and Maniatis, T. (eds) in: Molecular Cloning, Cold Spring Harbour Laboratory Press (1989).
(2) As described in: Elphinestone, J.G., Hennessey, J., Wilson, J.K., and Stead, D.E. (1996) Bulletin OEPP/EPPO 26, 663-678.
(3) PCR methods other than the one recommended are subject to approval by an officially designated reference laboratory.

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PCR mastermixes have to be prepared on a clean bench, which is decontaminated after work using detergents and UV
light.

— Method development: possible alterations to published methods might be required, such as the exact mastermix
composition (e.g. MgCl
2 concentration, primer concentration), the amount of template DNA used, and an adapted
temperature program (temperatures, holding times). The appearance of non-specific amplification products shall
lead to optimisation (e.g. annealing temperature, MgCl
2 concentration, primer concentrations etc.) to guarantee the
accuracy of the results.

— Validation aspects: the method to be applied in routine analysis has to be properly validated. The analysis of the
following controls has to be included in a set of samples: extraction control (no meat), negative PCR control, and
reference samples (male and female beef but also one sample of meat other than beef). Moreover, revalidation shall
be applied if key components in the procedure are changed, such as DNA polymerase (different supplier or
product) or primers (new lot).

— Good laboratory practice is deemed indispensable such as proper cleaning and decontamination of the working place

and instruments used, aliquoting of primers, dedicated use of pipettes etc.

4. Analysis of amplicons by gel electrophoresis

The PCR fragments (amplicons) obtained are to be analysed by gel electrophoresis. Either agarose gels stained with
ethidium bromide or polyacrylamide gels which are silver stained after completion of the electrophoretic separation
are applicable. An appropriate molecular weight marker on the gel to determine the approximate size of the obtained
amplicons has to be used.

5. Documentation

The obtained results have to be properly documented (gel image, description of results, noting of any non-expected
result).