Normativa UEAbr. 24, 2013
UE: autoridad alimentaria reevalúa uso de antocianinas como aditivo alimentario
The Panel concluded that the currently available toxicological database was inadequate to establish a numerical ADI for anthocyanins. For anthocyanins extracted from edible fruits and vegetables by aqueous processes, changes in composition would not be expected.
The Panel concluded that provided exposure from use as a food additive was comparable to that from the diet the underlying conclusion in the 1975 SCF opinion that such food additives derived from natural sources would still apply. The majority of data are on aqueous grape skin extract (GSKE) and blackcurrant extracts and the Panel considers that exposures estimated from current uses and use levels these extracts are unlikely to be of safety concern. The Panel recommends that the specifications for E 163 should be modified to reflect this conclusion. For anthocyanins extracted from other sources and/or using non-aqueous extraction methods the absence of characterisation does not allow verification that this conclusion in the 1975 SCF opinion could be applied. The Panel noted that for some extracts it had proven possible to assess a group based on toxicological and compositional data on representative samples across the range of extracts. The Panel concluded that refined exposure estimates of anthocyanins used as a food additive were much higher than dietary intakes and that these did not include intakes from colouring foods. Therefore the Panel would recommend that appropriate characterisation and toxicological data should be required to permit a further re-evaluation of anthocyanins including comparative data on anthocyanins (E 163) produced by aqueous extraction.
Following a request from the European Commission to the European Food Safety Authority (EFSA), the Scientific Panel on Food Additives and Nutrient Sources added to Food (ANS) was asked to provide a scientific opinion re-evaluating the safety of anthocyanins (E 163) when used as a food additive.
The Panel was not provided with a newly submitted dossier and based its evaluation on previous evaluations, additional literature that became available since then and the data available following a public call for data. The Panel noted that not all original studies on which previous evaluations were based were available for re-evaluation by the Panel.
Anthocyanins (E 163) are authorised as food additives in the EU, and have been previously evaluated by JECFA in 1982 and the SCF in 1975. JECFA has established an ADI of 2.5 mg/kg bw/day for anthocyanins from grape skin, while the SCF has not derived an ADI for anthocyanins.
Anthocyanins represent a very large group of water-soluble plant pigments. Anthocyanins are obtained by extraction from the natural strains of vegetables and edible fruits.
Anthocyanins are distributed in various fruits, and several fruit extracts are used as synonyms for anthocyanins. The extracts mentioned most often are grape skin extract (GSKE) (containing glucosides of the anthocyanins peonidin, malvidin, delphinidin and petunidin) and blackcurrant extract (containing the colouring matters cyanidin 3-rutinoside, delphinidin 3-rutinoside, cyanidin 3-glucoside and delphinidin 3-glucoside). However, in the EU specifications it is not indicated which fruits or vegetables can be used to obtain the food additive anthocyanins. In addition, the composition and identity of anthocyanins which may be present in the food additive E 163 is not specified. These compounds are normally present in food as glycosides (anthocyanins).
No JECFA specifications are available for specific anthocyanins, only for GSKE and blackcurrant extract, which contain more than one anthocyanins. The specifications of the European Commission for anthocyanins and of JECFA for GSKE and blackcurrant extract differ. Limits for mercury and cadmium are included in the EU specifications, but not in those of JECFA
Limited data on stability, reaction and fate in food were available. No formal method for the analysis of anthocyanins in food appears to have been adopted.
Studies on the toxicokinetics and toxicological properties of anthocyanins have mainly used fruit extracts, which contain several anthocyanins. Therefore, based on these studies, conclusions cannot be drawn for specific anthocyanins, but may be made for anthocyanins in general. Since anthocyanins used as the food additive E 163 are poorly defined, it is not clear whether the substances used in the various studies are relevant for assessment of the specific E 163 anthocyanins.
Studies in rats have revealed that the amount of absorption was low (< 2 %). After oral administration, a maximum plasma concentration of anthocyanins was reached after 15-120 minutes, depending on the aglycone and sugar moieties of the anthocyanins. Higher absorption levels (up to 37 %) have been reported in in situ experiments in anesthetised rats.
In rats as well as pigs, anthocyanins can be methylated or conjugated with glucuronic acid or sulphate and aglycone, have also been reported. However, delphinidin anthocyanins were not metabolized to any measurable extent.
Anthocyanins are excreted unchanged and as metabolites. Those with either a di- or tri-saccharide attached to them were primarily excreted unchanged in the urine. Plasma clearance rates are influenced by both the aglycone (delphinidin > cyanidin > petunidin = peonidin > malvidin) and the sugar moiety (galactoside > glucoside > arabinoside). Urinary excretion in rats and pigs is only 0.04-0.58 % of the ingested amount.
Limited studies in humans showed that only a small portion of orally ingested anthocyanins was absorbed (<1 %). Maximum plasma levels were reached within 2 hours of consumption. In humans, glucuronic acid conjugates, sulphate conjugates and methylated metabolites were found in both plasma and urine, together with oxidized derivatives. About 68 % of absorbed anthocyanins was reported to be metabolized, and excretion occurred mainly as a monoglucuronide.
The majority of anthocyanins ingested are excreted in the faeces. The elimination of plasma anthocyanins appeared to follow first-order kinetics in humans.
In guinea pigs and dogs, no short-term or subchronic toxic effects were observed at anthocyanins doses up to 3 g/kg and 15 % of grape-skin extract respectively in the diet. In addition, in rats fed an unspecified anthocyanins extract at levels up to 6 g/day or grape seed extract (GSE) or GSKE at dietary levels up to 2.5 % (1780 mg/kg bw/day in males and 2150 mg/kg bw/day in females) for a period of 90 days, no relevant treatment-related adverse effects were observed. It is not possible to convert the reported NOAEL for GSE and GSKE into a NOAEL for anthocyanins since the anthocyanins contents of GSE and GSKE were not further defined. In this case, the value reported by JECFA of 3 % anthocyanins content in GSKE (and assuming the same level in GSE), would result in a dose equivalent to 53 mg anthocyanins/kg bw/day for males and 64 mg anthocyanin/ kg bw/day for females.
In a 2-generation reproduction study with anthocyanins from GSKE, no effects were observed on reproductive performance or pup viability at dietary levels up to 15 % (equivalent to 225 mg anthocyanins/kg bw/day based on an assumed 3 % anthocyanins content in GSKE) anthocyanins. However, in both the F1 and F2 rats, body weight was reduced in the 15 % group. This was also observed in the F2 pups in the 7.5 % (equivalent to 112 mg anthocyanins/kg bw/day based on an assumed 3 % anthocyanins content in GSKE) group; however, this was marginal and related to a reduced food intake. In addition, a decrease in organ weights of the liver, adrenal and thyroid (without histological effects) occurred in the 15 % group of the F1 rats. The ADI established by JECFA was based on this study with 7.5 % GSKE in the diet (equivalent to 7500 mg/kg bw) considered as the NOAEL. Since GSKE contains approximately 3 % anthocyanins, this level was correlated to a NOAEL of 225 mg/kg bw/day for anthocyanins. This level was converted into an estimated ADI of 0-2.5 mg/kg bw/day for anthocyanins.
There are no indications that anthocyanins glycosides from currants, blueberries or elderberries induce developmental effects in rats, mice or rabbits at dose levels up to 9 g/kg bw.
Several anthocyanidins and anthocyanins (cyanidin, delphinidin, GSE and GSKE were negative in bacterial mutagenicity tests, with and without metabolic activation. Also in vitro Comet assays in mammalian cells did not result in increased DNA strand breaks when exposed to 0.1-100 μg/mL (GSE) or 1-10 μM (delphinidin, malvidin, pelargonidin and peonidin). However, in another study, doses of ≥ 50 μM (~ 17.5 mg/mL), delphinidin, cyanidin, malvidin, pelargonidin and peonidin did induce a slight, but significant increase in strand breaks in HT29 cells. Pelargonidin (doses ≤ 2 μM) was found non-genotoxic in a micronucleus test in HL-60 cells.
Overall, in most in vitro assays anthocyanins, tested at low doses, were not genotoxic. Some evidence of genotoxicity was provided by a single in vitro study using pure anthocyanidins. However in vivo a negative guideline bone marrow micronucleus test at a limit dose was considered to exclude in vivo genotoxicity of GSE and GSKE.
Due to a lack of data, no conclusion can be drawn with respect to long-term toxicity or carcinogenicity of anthocyanins.
The Panel noted that the specification for anthocyanins and the information available on their manufacture do not allow identification of the specific anthocyanins nor their overall composition in the material used as the food additive E 163. The Panel noted that there is no information on the range of anthocyanins composition within the food additive and it is therefore not possible to determine the extent to which the available toxicological data are relevant.
Furthermore, the Panel noted that the variety of sources for anthocyanins and lack of information on manufacturing process do not allow identification and quantification of minor components. The Panel considered that minor components could be present at different ratios than in the normal consumption of foods and might therefore exert different biological effects. The Panel also noted that the importance of data covering the range of compositions arising from different manufacturing methods was illustrated in the opinion of the Panel on Food Additives, Flavourings, Processing Aids and Materials in Contact with Food (AFC) on Rosemary extracts (EFSA, 2008b). The recent opinion of the Scientific Committee entitled ‘Guidance on Safety assessment of botanicals* and botanical preparations** intended for use as ingredients in food supplements’ (EFSA, 2009) highlights principles which should be considered in the risk assessment of any botanical preparations including extracts. The Panel considered that the absence of characterisation does not allow verification of the underlying presumption of safety on food additives derived from natural sources in the 1975 SCF opinion.
In a refined exposure estimation performed with the EFSA Comprehensive European Food Consumption Database, using the maximum reported use levels, the mean exposure to anthocyanins (E 163) range from 1.5 to 4.0 mg/kg bw/day for toddlers (high level exposure 3.2-6.9 mg/kg bw/day), from 1.5 to 4.7 mg/kg bw/day for children (high level exposure 2.7-7.8 mg/kg bw/day) and from 1.0 to 2.5 mg/kg bw/day for adolescents (high level exposure 1.6-3.9 mg/kg bw/day). The mean estimated exposures for adults and the elderly are respectively from 0.7 to 1.9 mg/kg bw/day (high level exposure 1.1-3.8 mg/kg bw/day) and from 0.5 to 1.1 mg/kg bw/day (high level exposure 0.9-2.3 mg/kg bw/day). Compared to these exposure estimates, the estimated exposure to anthocyanins from the regular diet is very low (at the mean about 0.1 mg/kg bw/day for adults and 0.3 mg/kg bw/day for children, values at the 97th percentiles are 0.6 mg/kg bw/day and 2.1 mg/kg bw/day respectively).
The Panel concluded that the currently available toxicological database was inadequate to establish a numerical ADI for anthocyanins.
For anthocyanins extracted from edible fruits and vegetables by aqueous processes, the Panel would not expect of changes in composition. The Panel concluded in principle that provided exposure from use of food colours was comparable to that from the diet the underlying conclusion of safety on food additives derived from natural sources in the 1975 SCF opinion would still apply.
Using a weight of evidence evaluation of toxicological and general exposure data, the Panel concluded that aqueous grape skin and blackcurrant extracts are unlikely to be of safety concern. The Panel recommends that the specifications for E 163 should be modified to reflect the conclusions on these two sources.
With the exception of aqueous grape skin and blackcurrant extracts, the Panel considered that the absence of characterisation does not allow verification of the applicability of the conclusion of safety of food additives derived from natural sources in the 1975 SCF opinion for anthocyanins extracted from other sources and/or using non-aqueous extraction methods.
The Panel concluded that the following information is required to permit an adequate risk evaluation for derivation of an ADI for anthocyanins (E 163) as food additive;
– Definition of the sources from which it is extracted
– Method of extraction
– Qualitative and quantitative chemical characterisation of the extracts including minor components
Data on toxicokinetics, subchronic toxicity, genotoxicity, reproductive and developmental toxicity and chronic toxicity/carcinogenicity for an appropriate number of extracts covering the range of sources and current manufacturing methods for each source.
The Panel considered that this data package would also need to include comparative data on anthocyanins (E 163) produced by aqueous extraction. The Panel noted that using target read across it has proven possible to perform a risk assessment for a group of extracts (e.g. rosemary extracts) based on compositional and toxicological data on representative samples. This approach was currently not applicable on anthocyanins.
The Panel concluded that refined exposure estimates of anthocyanins from their use as a food additive (E 163) were much higher than estimated intakes from the regular diet and that these did not include intakes from colouring foods. The general principle about safety of food additives derived from natural sources does not apply.
The Panel recommends that appropriate chemical characterisation and toxicological data are required to permit a further re-evaluation of anthocyanins.