PowerPointprsentation

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The Toxicology Forum 2005 ANNUAL EUROPEAN
MEETING November 8-10, Hilton Brussels Hotel
Retrospect on Scientific Approaches by the EU
Scientific Committee on Food and by Denmark in
the Safety Assessment of Food Enzymes
Ib Knudsen, DVM Chief Adviser in Food Safety and
Toxicology Danish Institute for Food and
Veterinary Research
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Retrospect on Scientific Approaches by the EU
Scientific Committee on Food and by Denmark in
the Safety Assessment of Food Enzymes

• Overview of the paper
• Scientific approaches 1972-1991
• Opinion of the Scientific Committee for Food
  expressed April 1991
• The October 1991 claim from the European
  Commission that Denmark violates the EU threaty
  regarding imported enzymes
• The situation until today

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Scientific approaches by JECFA from 1972-1982 in
reports no.15,18, 21, 22, and summarised in the
26th report
1

• The toxicological evaluation of enzyme
  preparations used in food is based on five
  classes (1)
• Enzymes obtained from edible tissues commonly
  used as food Regarded as food.
• Enzymes obtained from edible portions of plants
  Regarded as food.
• Enzymes derived from microorganisms that are
  traditionally accepted as constituents of foods
  or are normally used in preparation of foods
  Regarded as foods.

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Scientific approaches by JECFA from 1972-1982 in
reports no.15,18, 21, 22, and summarised in the
26th report
1

• The toxicological evaluation of enzyme
  preparations used in food is based on five
  classes (2)
• 4 Enzymes derived from non-pathogenic
  micro-organisms commonly found as contaminants of
  foods Not considered as foods. Establish
  chemical and microbial specifications. Conduct
  short-term toxicity studies. Establish ADI.
• 5 Enzymes derived from microorganisms that are
  less well known. Chemical and microbiological
  specifications required. More extensive tox.
  studies, including a long-term study in a rodent.

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Scientific approaches by JECFA from 1972-1982 in
reports no.15,18, 21, 22, and summarised in the
26th report
1

• Specifications for identity and purity (1)
• A comprehensive description of the main enzymatic
  activitiy or activities, incl. Enzyme Commission
  numbers, if any.
• A list of subsidiary enzymatic activities
• A clear description of the source
• A list of non-enzymatic substances derived from
  the source material(s), with limits where
  appropriate

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Scientific approaches by JECFA from 1972-1982 in
reports no.15,18, 21, 22, and summarised in the
26th report
1

• Specifications for identity and purity (2)
• A list of added co-factors, with limits where
  appropriate
• A list of carriers and deluents, with limits
  where appropriate
• A list of preservatives present from manufacture
  or deliberately added, with limits where
  appropriate

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The Danish Notification System in the 1980ies
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• The Notifier to file a notification scheme where
  the enzymes generally are classified according to
  the JECFA principles
• Enzymes derived from microbiel non-pathogenic
  food contaminants and from less known, but
  non-pathogenic microorganisms considered as one
  class, unlike JECFA
• For each preparation of the latter type a 28-day
  or 90-day study in rats as well as 2 short term
  mutagenicity tests (gene and chromosome
  mutations) were requested.
• For immobilised enzymes a less stringent
  requirement for tox data was applied

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Main elements of the Danish Notification System
in the 1980ies (1)
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• Notifiers name
• Producers name
• Product name
• Product usage (Food categories)
• Purpose
• Classification according to IUPAC or trivial name
• Strenght of the enzyme product
• Amount to be used in the food
• Source of enzyme
• Production procedure

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Main elements of the Danish Notification System
in the 1980ies (2)
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• Strenght and type of
• Enzyme activities
• Additives/technical aids
• Non-active components
• Absense of production organism
• Analytical data
• Like JECFA
• Toxicological studies, case-by-case
• Other information, e.g. usage in other countries

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The Rationale behind Danish Notification System
in the 1980ies
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• The products tested in the toxicological tests
  should be identical to or as close as possible to
  those used in the food production
• Toxiclogical concerns reduced, if
• Only very little enzyme preparation is in the
  food
• Enzyme activity is well defined and targeted
• Enzyme preparation is well purified
• Specific microbiological identification tags is
  available for the product
• In 1989, the Danish demands for toxicological
  tests were higher than in most other western
  countries.

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The Working Programme of the Scientific Committee
for Food for 1989-1992
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• 2.1 Additives
• To date enzymes have not been dealt with. Of
  immidiate priority are those four enzymes which
  are considered to be direct food additives. It
  may, however, be appropriate to start to look at
  enzymes used in food production in general

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Guidelines for the presentation of data on food
enzymes (SCF opinion expressed 11 April 1991)
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• Safety issues involved in the use of enzyme
  preparations
• Toxicological properties of the enzyme
  preparation
• Quantity of enzyme consumed
• Allergies and irritations caused by enzyme
  activity in the final product
• Unintended reaction products in the food caused
  by enzymatic reactions in the final foodstuffs
• Safety of the source organism (microbials)

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Technical information to be supplied for the
evaluation of an enzyme preparation to be used in
foodstuffs (SCF 1991) (1)
2

• Active components
• The principal enzyme activities
• The activity of the enzyme preparations
• A list of subsidiary enzymatic activities
• Source materials
• Animal sources
• Plant sources
• Microbial sources
• Genetically modified organisms

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Technical information to be supplied for the
evaluation of an enzyme preparation to be used in
foodstuffs (SCF 1991) (2)
2

• Manufacturing process
• The method of manufacture
• The purification procedure
• Carriers and other additives and ingredients
• Information on carriers, diluents, excipients,
  supports and other additives and ingredients
• For immobilized enzyme preparations, the carriers
  and immobilization agents should be acceptable
• Statement of percentage Total Organic Solids (TOS)

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Technical information to be supplied for the
evaluation of an enzyme preparation to be used in
foodstuffs (SCF 1991) (3)
2

• Usage
• Technological function of the enzyme
• Types of foodstuffs
• Maximum amount of enzyme preparation to be used
• Stability and fate in the food
• Amount of enzyme preparation in the final food
• Main reaction products and possible reaction
  products not considered normal constituents of
  the diet
• Possible effects on nutrients

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Technical information to be supplied for the
evaluation of an enzyme preparation to be used in
foodstuffs (SCF 1991) (4)
2

• Hygiene
• Enzyme preparations are to be produced in
  accordance with good food manufacturing practice.
  Stock cultures to be tested for purity
• Addition of the enzyme preparation to a food must
  not cause increase in total microbial count

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Technical information to be supplied for the
evaluation of an enzyme preparation to be used in
foodstuffs (SCF 1991) (5)
2

• Contaminants
• Should not contain toxicologically significant
  amounts of heavy metals
• No pathogenic microorganisms should be detectable
• Coliforms not more than 30 per gram
• Total viable count not more than 10²-10³ per gram
  as determined by suitable method

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Technical information to be supplied for the
evaluation of an enzyme preparation to be used in
foodstuffs (SCF 1991) (6)
2

• Contaminants (continued)
• Tests shall be performed to ensure that viable
  cells from the microbial source organism are not
  present in the final product
• Enzyme preparations may not contain any
  antibiotic activity
• Enzyme preparations may not contain detectable
  amounts of toxins

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Documentation for safety in use to be supplied
for the evaluation of an enzyme preparation to be
used in foodstuffs (SCF 1991) (1)
2

• Basic toxicological requirements
• For enzymes derived from edible parts of animals
  and plants no toxicological tests are normally
  required
• For enzyme preparations derived from
  microorganisms the following tests are normally
  required
• 90-day oral toxicity test in a rodent species
• Two short-term tests
• A test for gene-mutations in bacteria
• A test for chromosomal aberrations (preferably in
  vitro)

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Documentation for safety in use to be supplied
for the evaluation of an enzyme preparation to be
used in foodstuffs (SCF 1991) (2)
2

• Exemptions from the basic toxicological
  requirements
• If one enzyme preparation from a specific strain
  has been throroughly tested and the manufacturing
  process does not differ significantly for other
  enzymes from the same strain, the full testing
  battery may be waived.
• If the microorganism used in the production
• Has a long history of safe use, and
• Belongs to a species about which it has been
  documentet that no toxins are produced, and
• The actual strain used is of well documented
  origin

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Documentation for safety in use to be supplied
for the evaluation of an enzyme preparation to be
used in foodstuffs (SCF 1991) (3)
2

• Exemptions from the basic toxicological
  requirements (continued)
• When a mutant is substituted for the original
  strain of microorganism used, a modified less
  comprehensive test procedure may be appropriate.
• In the combination with immobilized enzyme
  preparations
• With introduction of well specified,
  non-toxin-producing genetically engineered source
  organisms
• There may also be circumstances where additional
  testing are needed

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (1)
3

• The 21 October 1991 the European Commission
  opened the case against Denmark with the view
  that Denmark violated the EU threaty regarding
  sales of food enzymes imported from Germany,
  because requests for additional data is only
  allowed
• If it can be demonstrated to be necessary in
  order to fulfill unavoidable demands, such as
  protection of public health
• If it keeps proportionate to the desired target
• If the instruments chosen to achieve this target
  is those of least disturbance to the free trade
  between the Member States

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (2)
3

• The 12 July 1993 after several exchanges of
  notes - the European Commission requested Denmark
  within two months to withdraw its demand for
  toxicological studies on enzyme preparations due
  to violation of the EU Treaty, Article 30 ff.

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (3)
3

• The Danish reply of 12 October 1993 underlines
  that
• Articles 30 and 36 applies since there are no
  harmonised EU regulation
• The step taken (request for 90-day studies) is
  not an un-proportional measure
• Denmark accepts 90-day studies performed in the
  other Member States
• The Danish Authorities requests a meeting with
  the Commission to explain the Danish regulation
  and Danish point-of-views

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (4)
3

• The Commission accepts the idea about a meeting
  in letter of 18 March 1994
• The meeting takes place 5 July 1994
• The Danish Authorities writes a new letter to the
  Commission 16 September 1994
• Thanking for the meeting
• Promising to re-write the Danish regulation so it
  become totally clear that studies performed in
  other Member States are fully acceptable
• Request a new meeting with the Commission if the
  Commission does not drop the case

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (5)
3

• Meeting between Commissioner Bangeman and Danish
  secretary of Commerce 14 September 1995
• The Danish regulation complies with SCF
  guidelines from 1991
• 90-day studies only requested on case-by-case
• Letter of 19 Sept. 1995 from the Danish Secretary
  of Health to Commissioner Monti
• Proposing a Danish scientific report to explain
  the background for the Danish request for
  toxicological data
• Proposing another meeting

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (6)
3

• Reply of 27 Nov. 1995 from Commissioner Monti
• ..the Commission attaches,inter alia, great
  importance to the fact that enzymatic
  preparations in question so-called processing
  aids have for years been legally produced and
  used in the European foodstuffs industry
• Consequently, the Commission had no other choice
  that to decide on 4th April 1995 to bring the
  case before the Court of Justice of the European
  Communities. Although its Legal Service has
  prepared the application for that purpose, it has
  not yet been filed with the Court pending the
  comments from DG III and DG XV.
• In the ligth of the foregoing, You will
  appreciate that I regret that you still insist on
  the necessity and proportionality of the
  contested measure. Moreover, given the advanced
  stage of the preparation of the Court
  proceedings, it is difficult for the Commission
  to grant any further delay.

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (7)
3

• Reply of 21 Dec. 1995 from The Danish Secretary
  of Health, Yvonne Herløv Andersen
• I am glad that you agree, that we together can
  find a solution to the problem, and that we as
  soon as possible arrange a meeting at the
  technical level
• I am sure that it then is possible to discuss
  changes in the Danish regulation on food
  enzymes.
• In my letter to you of 19 Sept 1995 I suggested
  that the Danish National Food Agency should
  develop a scientific report addressing the Danish
  requirements for marketing of food enzymes to be
  given to the Scientific Committee for Food
• I attach this report

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (8)
3

• The report
• On the need for toxicological testing of
  microbial food enzymes
• Discussion paper prepared by
• Inger Thorup, Jørgen Schlundt and
• Torben Hallas-Møller
• Copenhagen, November 1995

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (9)
3

• Background considerations (1)
• There are conflicting reports in the literature
  on the exact identification of microorganisms,
  especially fungi,
• The capabilities of all known microorganisms,
  including the potential source organisms, to
  produce toxic metabolites are highly influenced
  by the environmental conditions,
• There are often conflicting information on the
  metabolic profile of so-called identical
  microorganisms reported in the literature,

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (10)
3

• Background considerations (2)
• The industrial strain improvements often include
  isolation of mutants, which could behave
  differently compared to the parent organisms,
  e.g. leading to the formation of toxins,
• Toxic metabolites migth also emerge in the final
  product due to changes in the manufacturing of
  the enzyme e.g. the purification process,
• Limitations in the analytical procedures

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (11)
3

• Conclusions (3)
• A safety assessment only relying on the
  identification of a source organism on species
  level will give a false impression of safety,
• A minimum battery of safety tests must be
  performed on each enzyme product, preferably be
  defined from batch to batch,
• The request of a 90 days study and 2 mutagenicity
  tests as required in the Danish guidelines as
  well as those of SCF and CoT, is judged by these
  bodies to give reasonable assurrance of safety
• There may be situations, where a 90-day feeding
  study and 2 mutagenicity tests may not be needed

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (12)
3

• Follow-up
• Main issues at a meeting in Bruxelles 27 February
  1996
• The strength of the scientific basis for the
  Danish rules
• Better precision and clarification of the rules
• Letter to the Commission of 29 February 1996
• Appraisal of SCF as the scientific forum to
  settle the scientific issues behind the case
• Willingness to notify re-written Danish rules
• Letter to the Commission of 25 March 1996
• Presentation of the mandate to be given to the
  two independent and unbiased Danish scientific
  experts

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (13)
3

• SAFETY ASPECTS OF MICROBIAL FOOD ENZYMES
• MICROBIOLOGICAL AND TOXICOLOGICAL TESTING OF
  MICROBIAL FOOD ENZYMES/ENZYME PRODUCTS
• FOLKE RASMUSSEN, D.V.M., D.V.Sc.
• Professor Emeritus in Pharmacology and Toxicology
• NIELS SKOVGAARD, D.V.M.
• Professor Emeritus in Food Hygiene and
  Microbiology
• The Royal Veterinary and Agricultural University,
  Copenhagen

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (14)
3

• SAFETY ASPECTS OF MICROBIAL FOOD ENZYMES
• Conclusion
• A safety assessment only relying on the
  identification of a source organism on species
  level alone will give a false impression of
  safety. The same arguments applies to changes in
  the production technology apart from the change
  of species e.g. change of the nutritional
  composition of the broth. Consequently, in
  general a safety documentation has to include
  toxicity tests performed on microbially derived
  enzyme products.
• Issued 22 April, 1996

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The Commission claims that the Danish request for
a 90-day study on certain enzyme preparations
hinders the free trade (EU-Treaty Article 30) (15)
3

• Follow-up
• Letter to the Commission of 6 May 1996
• Revised version of the notification paper
• The scientific report from the two independent
  experts
• Letter from the Danish EU Office of 12 June 1996
• Seems that the Legal Service of the Commission
  will postpone to bringing the enzyme-case to the
  Court
• DG III and DG XV will now examine the Danish
  scientific report
• Letter from the Danish EU Office of 23 September
  1996
• DG XV is according to the Legal Service inclined
  to drop the case, while DG III still has not
  decided what to do.

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The situation until today (1)
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• The proposed changes of 18 September 1998 to
  Directive 95/2/EU on other additives to
  foodstuffs than colors and sweeteners says
• The Commission is obliged as soon as possible to
  examine the problem in relation to enzymes and as
  soon as possible to present a reasonable proposal
  for the Council and the European Parliament

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The situation until today (2)
4

• The Commission White Paper from 1999 states that
  the use of enzymes in general should be
  investigated with the purpose to
• Ensure consumer safety
• Ensure the trade with enzymes
• Regulate their use and
• Develop procedures for risk assessment, use and
  labeling of enzymes and their products

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The situation until today (3)
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• Brussels, 2 February 2005
• WGA/003/03 rev8
• Draft Working Paper for a
• REGULATION OF THE EUROPEAN AND OF THE COUNCIL
• On Food Enzymes
• A written hearing was initiated in February 2005.
  It has been suggested that a Commission proposal
  should be ready for the Parliament and for the
  Council in November/December 2005

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The Toxicology Forum 2005 ANNUAL EUROPEAN
MEETING November 8-10, Hilton Brussels Hotel
Thank you for your attention!
Ib Knudsen, DVM Chief Adviser in Food Safety and
Toxicology Danish Institute for Food and
Veterinary Research