Scientific Opinion on the Safety and Efficacy of thaumatin for all animal species 1

SCIENTIFIC OPINION Scientific Opinion on the Safety and Efficacy of thaumatin for all animal species 1 EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) 2,3 ABSTRACT European Food Safety Authority (EFSA), Parma, Italy Thaumatins are a mixture of proteins first isolated from the katemfe fruit (Thaumatococcus daniellii Benth) of West Africa. The two principal proteins found in thaumatin (thaumatin I and II) have been sequenced. Thaumatin has previously been assessed for safety for consumers by both JECFA and the Scientific Committee for Food, neither body proposing an ADI. It is currently authorised for use as a sweetener and flavour enhancer in the European Union. Thaumatin at a recommended use range of 1 to 5 mg/kg complete feed is safe for all animal species with a considerable margin of safety. With respect to this margin of safety thaumatin can be administered simultaneously via feed or water for drinking. Thaumatins are highly digestible proteins and no residues in edible tissues/ products are expected. Consequently there are no concerns for the consumer safety. The MSDS provided by the manufacturer identifies as potential hazards irritation to the eyes and sensitisation following inhalation. Sensitisation and irritation to the skin are not addressed. The precautions for handling the product proposed by the manufacturer (respiratory, hand, eye and skin protection) are considered sufficient to ensure user safety. Thaumatins occur in nature, albeit with a narrow distribution, amongst plant species. Its use in animal nutrition is not expected to affect the environment as it is not excreted due to its essentially complete digestion. Thaumatin is used in food as a sweetener/flavour modifier. Since its function in feed is essentially the same as that in food, no further demonstration of efficacy is considered necessary. European Food Safety Authority, 2011 KEY WORDS Thaumatin, sensory additive, protein, sweetener, flavour enhancer, safety, efficacy 1 On request from the European Commission Question No EFSA-Q-2010-01223, adopted on 06 September 2011. 2 Panel members: Gabriele Aquilina, Georges Bories, Andrew Chesson, Pier Sandro Cocconcelli, Joop de Knecht, Noël Albert Dierick, Mikolaj Antoni Gralak, Jürgen Gropp, Ingrid Halle, Christer Hogstrand, Reinhard Kroker, Lubomir Leng, Secundino López Puente, Anne-Katrine Lundebye Haldorsen, Alberto Mantovani, Giovanna Martelli, Miklós Mézes, Derek Renshaw, Maria Saarela, Kristen Sejrsen and Johannes Westendorf. Correspondence: FEEDAP@efsa.europa.eu 3 Acknowledgement: The Panel wishes to thank the members of the Working Group on Feed Flavourings, including Paul Brantom, Joaquim Brufau, Gérard Pascal and Guido Rychen for the preparatory work on this scientific opinion. Suggested citation: EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP); Scientific Opinion on the Safety and Efficacy of thaumatin for all animal species.. [10 pp.] doi:10.2903/j.efsa.2011.2354. Available online: www.efsa.europa.eu/efsajournal European Food Safety Authority, 2011

SUMMARY Following a request from the European Commission, the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety for the target animals, consumers of animal products, users and the environment and the efficacy of the natural sweetener thaumatin for all animal species. The thaumatins are a mixture of proteins first isolated from the katemfe fruit (Thaumatococcus daniellii Benth) of West Africa. Commercial extraction of thaumatin from the fruit began in the 1970s and the sequencing of the two principal proteins found in thaumatin (thaumatin I and thaumatin II) was completed in 1990. It has been previously assessed for safety for consumers by both JECFA and the Scientific Committee for Food, neither body proposing an ADI. It is currently authorised for use as a sweetener or flavour enhancer in the European Union and in many other countries. Thaumatin at a recommended use range of 1 to 5 mg/kg complete feed is safe for all animal species with a considerable margin of safety. Consequently, thaumatin can be administered simultaneously via feed or water for drinking. Thaumatins are highly digestible proteins and no residues in edible tissues/ products are expected. Consequently there are no concerns for the consumer safety. The MSDS provided by the manufacturer identifies as potential hazards irritation to the eyes and sensitisation following inhalation. Sensitisation and irritation to the skin are not addressed. The precautions for handling the product proposed by the manufacturer (respiratory, hand, eye and skin protection) are considered sufficient to ensure user safety. Thaumatins occur in nature, albeit with a narrow distribution, amongst plant species. Its use in animal nutrition is not expected to affect the environment since it is not excreted as such due to its essentially complete digestion. Thaumatin is used in food as a sweetener/flavour modifier. Since its function in feed is essentially the same as that in food, no further demonstration of efficacy is considered necessary. 2

TABLE OF CONTENTS Table of contents... 3 Background... 4 Terms of reference... 4 Assessment... 6 1. Introduction... 6 2. Characterisation... 6 2.1. Manufacturing process... 7 2.2. Stability... 7 2.3. Conditions of use... 7 2.4. Evaluation of the analytical methods by the European Union Reference Laboratory (EURL)7 3. Safety... 7 3.1. Safety for the target species... 7 3.1.1.1. Conclusions on the safety for the target species... 8 3.2. Safety for the consumer... 8 3.3. Safety for the user... 8 3.4. Safety for the environment... 8 4. Efficacy... 8 5. Post-market monitoring... 8 Conclusions and recommendations... 9 Documentation provided to EFSA... 9 References... 9 Appendix... 10 3

BACKGROUND Regulation (EC) No 1831/2003 4 establishes the rules governing the Community authorisation of additives for use in animal nutrition. In particular, Article 4(1) of that Regulation lays down that any person seeking authorisation for a feed additive or for a new use of a feed additive shall submit an application in accordance with Article 7. In particular Article 10(2) of that Regulation also specifies that for existing products within the meaning of Article 10(1), an application shall be submitted in accordance with Article 7, at the latest one year before the expiry date of the authorisation given pursuant to Directive 70/524/EEC for additives with a limited authorisation period, and within a maximum of seven years after the entry into force of this Regulation for additives authorised without time limit or pursuant to Directive 82/471/EEC. The European Commission received a request from the Feed Flavourings Authorisation Consortium European Economic Interest Grouping (FFAC EEIG) 5 for authorisation of thaumatin, to be used as a feed additive for all animal species (category: sensory additives; functional group: flavourings) under the conditions mentioned in Table 1. According to Article 7(1) of Regulation (EC) No 1831/2003, the Commission forwarded the application to the European Food Safety Authority (EFSA) as an application under Article 4(1) (authorisation of a feed additive or new use of a feed additive) and under Article 10(2) (re-evaluation of an authorised feed additive). EFSA received directly from the applicant the technical dossier in support of this application. 6 According to Article 8 of that Regulation, EFSA, after verifying the particulars and documents submitted by the applicant, shall undertake an assessment in order to determine whether the feed additive complies with the conditions laid down in Article 5. The particulars and documents in support of the application were considered valid by EFSA as of 30 November 2010. The additive is a sweetener for use in foodstuffs (Council Directive 1994/35 on sweeteners for use in foodstuffs). 7 The additive is listed in the Feed Additives Register (Regulation (EC) No 1831/2003) on the basis of the notification procedure and has not been previously assessed by EFSA for this purpose. Thaumatin has been previously assessed by the Scientific Committee for Food (SCF) (EC, 1985 and 1989) and JECFA (1986) as sweetening agent. TERMS OF REFERENCE According to Article 8 of Regulation (EC) No 1831/2003, EFSA shall determine whether the feed additive complies with the conditions laid down in Article 5. EFSA shall deliver an opinion on the safety for the target animal(s), consumer, user and the environment and the efficacy of the active substances listed in the Appendix to Table 1, when used under the conditions described in Table 1. 4 OJ L 268, 18.10.2003, p.29 5 Feed Flavourings Authorisation Consortium European Economic Interest Grouping (FFAC EEIG), Avenue Louise 130A, 1050, Brussels, Belgium 6 EFSA Dossier reference: FAD-2010-0138 7 OJ L 237, 10.9.1994, p. 3 12 4

Table 1: Description and conditions of use of the additive as proposed by the applicant Additive Registration number/ec No/No (if appropriate) Category(ies) of additive Functional group(s) of additive Chemically defined flavouring - Thaumatin - 2. Sensory additives b) flavouring compounds Thaumatin Composition, description Chemical formula Polypeptide of 207 amino acids Description Purity criteria (if appropriate) NLT 15.1 % nitrogen on the dried basis equivalent to NLT 93 % Proteins (N x 6.2) Method of analysis (if appropriate) Protein determination as nitrogen 8 Trade name (if appropriate) - Name of the holder of authorisation (if appropriate) - Species or category of animal All species and categories Conditions of use Minimum content Maximum content Withdrawal period Maximum Age mg /kg of active substance / kg of complete (if appropriate) feeding-stuff or water for drinking - - - - Other provisions and additional requirements for the labelling Specific conditions or restrictions - for use (if appropriate) Specific conditions or restrictions All feedingstuffs and water for drinking, as part of a premixture only for handling (if appropriate) Post-market monitoring - (if appropriate) Specific conditions for use in complementary feedingstuffs - (if appropriate) Marker residue Maximum Residue Limit (MRL) (if appropriate) Species or category of Target tissue(s) or Maximum content in animal food products tissues - - - - 8 Available on the webpage of the CRL 5

ASSESSMENT 1. Introduction Thaumatin is a low-calorie sweetener said to be approximately 10,000-times as sweet as sucrose on a molar basis. However, the nature of the sweet taste, particularly its persistence, differs from that of sugar and, as a consequence, it is more commonly used as a flavour modifier. The thaumatins were first found as a mixture of proteins isolated from the katemfe fruit (Thaumatococcus daniellii Benth) of West Africa. Commercial extraction of thaumatin from the fruit began in the 1970s and the sequencing of the two principal proteins found in thaumatin (thaumatin I and thaumatin II) was completed in 1990. Thaumatin is an authorized sweetener according to Council Directive 94/35/EC 9 on sweeteners for use in foodstuffs and an authorized flavour enhancer in chewing-gum, beverages and desserts under the Miscellaneous Additives Directive (European Parliament and Council Directive No 95/2/EC of 20 February 1995 on food additives other than colours and sweeteners) 95/2/EC. 10 Thaumatin is also approved in Europe as a flavour preparation under Council Directive 88/388/EC (Council Directive 88/388/EEC of 22 June 1988 on the approximation of the laws of the Member States relating to flavourings for use in foodstuffs and to source materials for their production). 11 When used as a sweetener, maximum levels need to be observed in given food categories but, when used as a flavour enhancer, thaumatin can be used without restriction. Similar approvals exist in Switzerland, Canada, Israel, Mexico, Japan, Hong Kong, Korea, Singapore, Australia, New Zealand and South Africa. It is classified as GRAS (Generally Recognized as Safe) by the Food and Drug Administration in the USA. A consortium of companies (FFAC) supplying flavours to the feed industry has requested authorisation for the use of thaumatin as an additive to feed and water for drinking for all animal species. Regulation (EC) No 1831/2003 allows substances already approved for use in human food to be assessed with a more limited procedure than for other feed additives. However use of such an abbreviated procedure is always subject to the condition that the food safety assessment is relevant to the use in feed. 2. Characterisation Thaumatin (CAS No. 53850-34-3) is a mixture of two proteins (thaumatin I 22,209 and thaumatin II 22,293 daltons), extracted from the arils of Thaumatococcus daniellii. Analysis of six batches showed a mean nitrogen content of 15.8 % which using a conversion factor of 6.2 gives a mean protein content of 98.0 %. This does not comply with Commission Directive 2008/60/EC (specific purity criteria concerning sweeteners for use in foodstuffs) which specifies a minimum nitrogen content of 16 % on a Dry Matter (DM) basis and using a conversion factor of 5.8 to estimate the protein content. Other specifications of the Directive are not more than 3 % carbohydrate and not more than 2 % sulphated ash. The analysed carbohydrate content was around 0.12 % and sulphated ashes 0.42 % based on three batches. Limits set for chemical impurities on a dry matter basis are lead ( 3 mg/kg), aluminium ( 100 mg/kg) and arsenic ( 3 mg/kg) and for microbial contaminants total anaerobes (<10 3 ) and E. coli (non detectable in 1 g). These specifications are those set by the European Union (Commission Directive 2008/60/EC) 12 and comply with those of JECFA (2006). 13 Data from six batches of the 9 OJ L 237, 10.9.1994, p. 3 12 10 OJ L 61, 18.3.1995, p. 1 40 11 OJ L 184, 15.7.1988, p. 61 66 12 OJ L 158 18.06.2008, p.17-40 13 Technical Dossier/ Section II/ Annex 8 6

product show compliance with these limits for aluminium and microbial contaminants but no data were shown for lead and arsenic. 2.1. Manufacturing process Thaumatin is produced by aqueous extraction of the homogenised arils of Thaumatococcus daniellii. Extracts are membrane filtered or dialysed to remove low molecular weight material and then the proteins are absorbed on an ion-exchange resin and eluted. The recovered extract is further purified by dialysis, filtered and freeze-dried. 2.2. Stability The manufacturer indicates a shelf life of at least 36 months when stored in air tight containers and kept at a cool temperature. Short term stability of the additive in water for drinking is indicated by the documented use of the product in a variety of beverages (soft drinks) (Etheridge, 1994). 2.3. Conditions of use The applicant proposes the use of thaumatin in feed at a dose of 1 to 5 mg/kg complete feedingstuffs and in water for drinking for all animal species. No specific proposals are made for doses to be used in water for drinking. No withdrawal period is specified. 2.4. Evaluation of the analytical methods by the European Union Reference Laboratory (EURL) EFSA has verified the EURL report as it relates to the methods used for the control of thaumatin in animal feed. The Executive Summary of the EURL report can be found in the Appendix. 3. Safety 3.1. Safety for the target species The approach to safety assessment for target species takes account of the use levels in animal feed, relative to those used in human food. To avoid unnecessary testing the data used in consumer safety assessment are used to provide an indication of safety for the target species. Any difference in metabolism of the substance is also considered in this evaluation along with any evidence for specific toxicity which might be relevant to the proposed target species. Compared with an estimated human exposure of 2 mg/person/day (equivalent to 93 µg/kg metabolic body weight (= kg 0.75 )/day) 14 the calculated intake by the target species (salmonids 118, piglets 526 and dairy cows 777 µg/kg 0.75 ) from the proposed maximum feed concentration of 5 mg exceeds that of humans resulting from use in food. As a consequence, safety for the target species at the feed concentration supplied cannot be derived from the risk assessment for food use. As an alternative the maximum feed concentration which can be considered as safe for the target animal is derived from the lowest NOAEL found for thaumatin. This NOAEL was identified by JECFA (2006) to be 10 g/kg bw/day in dogs (the highest dose tested). The maximum safe intake of thaumatin is therefore 100 mg/kg bodyweight/day. Table 2 shows the maximum safe feed concentrations of thaumatin for the target species. 14 See also Guidance for the preparation of dossiers for sensory additives 7

Table 2: Maximum safe feed concentrations for thaumatin in target species Target animal Default settings Maximum safe intake of thaumatin Body weight (kg) Feed intake (g/d) mg/d mg/kg feed Salmonids 2 40 200 5000 Veal calves (milk replacer) 100 2000 10000 5000 Cattle for fattening 400 8000 40000 5000 Pigs for fattening 100 3000 10000 3333 Sows 200 6000 20000 3333 Dairy Cows 650 20000 65000 3250 Turkeys for fattening 12 400 1200 3000 Piglets 20 1000 2000 2000 Chickens for fattening 2 120 200 1667 Laying hens 2 120 200 1667 3.1.1.1. Conclusions on the safety for the target species The FEEDAP Panel considers the proposed range of use levels of 1 to 5 mg thaumatin/kg feed as safe for all animal species with a considerable margin of safety. This margin of safety would allow the simultaneous administration of thaumatin in feed and water for drinking. 3.2. Safety for the consumer JECFA (1986) and SCF (EC, 1985, 1989) evaluated thaumatin as a sweetener and an ADI was not specified ( not allocated ). The amino acid sequence of the protein is known and there is no indication of the presence of unusual amino acid side-chains, or atypical peptide linkages or end-groups. Thaumatin, in vitro, can be broken down to the same extent as ovo-albumin, and the in vivo nitrogen digestibility of both compounds appears comparable. No antibodies to thaumatin were detected in either rats or man after prolonged oral administration (JECFA, 1986). Consequently no residues in animal tissues or products are to be expected and the use of thaumatin in animal nutrition is considered safe for the consumer. 3.3. Safety for the user No data on the safety for the user was provided. The Material Safety Data Sheet (MSDS) provided by the manufacturer identifies as potential hazards irritation to the eyes and sensitisation following inhalation. The precautions for handling the product proposed by the manufacturer (respiratory, hand, eye and skin protection) are considered sufficient to ensure user safety. 3.4. Safety for the environment Thaumatin occurs in nature, albeit with a narrow distribution, amongst plant species. Its use in animal nutrition is not expected to affect the environment since it is not excreted as such due to its essentially complete digestion. 4. Efficacy Thaumatin is used in food as a sweetener and flavour enhancer. Since its function in feed is essentially the same as that in food, no further demonstration of efficacy is necessary. 5. Post-market monitoring The FEEDAP Panel considers that there is no need for specific requirements for a post-market monitoring plan other than those established in the Feed Hygiene Regulation 15 and Good Manufacturing Practice. 15 OJ L 35, 8.2.2005, p. 1. 8

CONCLUSIONS AND RECOMMENDATIONS CONCLUSIONS Thaumatin at a use range of 1 to 5 mg/kg feed is safe for all animal species with a considerable margin of safety. Therefore thaumatin can be administered simultaneously via feed or water for drinking. Thaumatin is a highly digestible protein and no residues in edible tissues/ products are expected. Consequently there are no concerns for the consumer safety. The MSDS provided by the manufacturer identifies as potential hazards irritation to the eyes and sensitisation following inhalation. Sensitisation and irritation to the skin are not addressed. The precautions for handling the product proposed by the manufacturer (respiratory, hand, eye and skin protection) are considered sufficient to ensure user safety. Thaumatin occurs in nature, albeit with a narrow distribution, amongst plant species. Its use in animal nutrition is not expected to affect the environment since it is not excreted as such due to its essentially complete digestion. Thaumatin is used in food as a sweetener/flavour enhancer. Since its function in feed is essentially the same as that in food, no further demonstration of efficacy is necessary. Recommendations The FEEDAP Panel recommends the use of the provisions of Commission Directive 2008/60/EC as specification for authorisation of thaumatin as a feed additive. DOCUMENTATION PROVIDED TO EFSA 1. Thaumatin for all animal species. September 2010. Submitted by Feed Flavourings Authorisation Consortium European Economic Interest Grouping (FFAC EEIG). 2. Thaumatin for all animal species. Supplementary information. April 2011. Submitted by Feed Flavourings Authorisation Consortium European Economic Interest Grouping (FFAC EEIG). 3. Evaluation report of the European Union Reference Laboratory for Feed Additives on the methods(s) of analysis for Thaumatin. 4. Comments from Member States received through the ScienceNet. REFERENCES EC (European Commission), 1985: Opinion of the Scientific Committee for Food on Sweeteners. Sixteenth series. Sweeteners (Opinion expressed on 14 September 1984). Available from: http://ec.europa.eu/food/fs/sc/scf/reports_en.html EC (European Commission), 1989: Opinion of the Scientific Committee for Food on Coulouring matters, sweeteners, quinine and emulsifiers, stabilizers, thickeners and gelling agents. Twenty-first series Second Addendum to the first Report of the Scientific Committee for Food on certain monomers and other starting substances to be used in the manufacture of plastic materials and articles intended to come into contact with foodstuffs (adopted on 10 December 1987)- first add. 19th series of SCF reports. Available from: http://ec.europa.eu/food/fs/sc/scf/reports_en.html Etheridge, K., 1994. The Sales and Marketing of Talin. In: M. Witty, J.D Higginbotham D.: Thaumatin. CRC Press, pp. 47 59. JECFA, 1986. Evaluation of certain food additives and contaminants. Twenty-ninth report of the Joint FAO/WHO Expert Committee on Food Additives. World Health Organization. Technical Report Series 733. 9

JECFA, 2006. FAO WHO Expert Committee on Food Additives: FAO JECFA monographs. Thaumatin. Specification Monograph 1. APPENDIX Executive Summary of the Evaluation Report of the European Union Reference Laboratory for Feed Additives on the Method(s) of Analysis for Thaumatin In the current application authorisation is sought under articles 4(1) (new use in water) and 10(2) (reevaluation of additives already authorised under Directive 70/524/EC) for Thaumatin under the category "sensory additives", functional group 2(b) "flavouring compounds", according to the classification system of Annex I of Regulation (EC) No 1831/2003. Thaumatin belongs to the Chemically Defined Flavourings 30 (Miscellaneous substances), according to the Annex I of Commission Regulation (EC) No 1565/2000. Authorisation is sought for the use of Thaumatin for all species and categories. According to the Commission Directive 2008/60/EC preparations containing Thaumatin should have a minimum nitrogen content of 15 % related to dry mass, equivalent to a minimum content of 93 % protein. The feed additive is intended to be incorporated only into feedingstuffs or drinking water, in combination with other flavouring substances as constituents of flavouring mixtures. The Applicant suggested no minimum or maximum levels for Thaumatin, but normal contents of the flavouring compound in feedingstuffs range up to from 0.1 to 100 mg/kg. For the identification of Thaumatin in the feed additive, the Applicant proposed the internationally recognised FAO JECFA monograph for Thaumatin in food additives. For the determination of the Thaumatin nitrogen content in the feed additive, the Applicant submitted the Kjeldhal method, described in the above mentioned JECFA monograph. Even though no performance characteristics are provided, the EURL recommends for official control the JECFA monograph based on the Kjeldhal method, for the quantification of the Thaumatin nitrogen content in the feed additive. As no experimental data were provided by the Applicant for the determination of the product in feedingstuffs and water, the EURL could not evaluate nor recommend the method for official control to determine Thaumatin in feedingstuffs and water. Further testing or validation of the methods to be performed through the consortium of National Reference Laboratories as specified by Article 10 (Commission Regulation (EC) No 378/2005) is not considered necessary. 10