Following a request from the European Commission to the European Food Safety Authority, the Scientific Panel on Food Additives and Nutrient Sources added to Food (ANS) was asked to provide a scientific opinion on the safety in use of natamycin (E 235) as a food additive. In addition, EFSA should address the issue of antimicrobial resistance to natamycin.
Natamycin (pimaricin) is a fungicide of the polyene macrolide group. According to Directive 95/2/EC, natamycin may be used for the surface treatment of semi-hard and semi-soft cheese and dry, cured sausage at a maximum level of 1 mg/dm2 in the outer 5 mm of the surface, corresponding to 20 mg/kg.
The Scientific Committee for Food (SCF) in 1979 did not establish an Acceptable Daily Intake (ADI) but considered that in relation to the uses of natamycin on cheese and sausages, the database was adequate and did not give rise to safety concern.
The Joint FAO/WHO Expert Committee on Food Additives (JECFA) reviewed the safety of pimaricin (natamycin) in 1968, 1976 and 2002 and assigned an ADI of 0.3 mg/kg body weight (bw)/day.
Information available on the metabolism of natamycin suggests that natamycin is not absorbed to a significant extent from the gastrointestinal tract and is rapidly excreted in faeces either unchanged or as degradation products.
In toxicological studies, the effects observed in animals were a decrease in food intake with a decrease in the rate of body weight gain, gastrointestinal irritation and diarrhoea. Dogs are the most sensitive species to these effects.
Three subchronic toxicity studies with natamycin are available, two in the rat and one in the dog. In the first study, no modifications of haematological and biochemical parameters and organ weights were noted. In the second rat study, decreases of mean food intake and mean body weight have been observed. The No-Observed-Adverse-Effect Level (NOAEL) is considered to be 45 mg/kg bw/day. In the third study, dogs were exposed for 3 months to natamycin. Transient diarrhoea and slight body weight loss have been observed. The NOAEL is considered to be 12 mg/kg bw/day.
Two long-term studies are available, a 2-year chronic toxicity study in the rat and a 2-year chronic toxicity study in the dog. In the rat study, decrease of food intake and reduced growth rate were seen only at the highest dose group. The data showed that the numbers and types of tumours were not significantly different in any of the natamycin-treated groups compared with the untreated control animals. The NOAEL of this study is considered to be 22.4 mg/kg bw/day. In the dog study, the highest dietary concentration induced obesity among the animals. Dietary levels of 6.25 mg/kg bw/day, or less did not affect body weight gain. The NOAEL of this study is considered to be 6.25 mg/kg bw/day.
Natamycin bears a structural alert for genotoxicity since the molecule contains an epoxide ring. However, in the light that:
- the induction of chromosomal aberrations observed in a recent study was accompanied by cytotoxicity,
- there are in vitro studies on mutagenicity in bacteria and mammalian cells and on chromosomal aberrations in mammalian cells which were performed in compliance with GLP and were negative,
- no substance-related neoplastic effects were observed in the long term studies,
the Panel considered that the available data do not raise concern with respect to genotoxicity of natamycin.
In a three-generation study of reproductive toxicity in the rat, at the highest dose, an increased number of fetuses born dead, and a decreased number of animals born alive surviving at 21 days in F1 generation, was described. The NOAEL of this study amounts to 50 mg/kg bw/day.
A developmental toxicity study has been performed in female rats from the second litter of the F1 generation of the three-generation reproductive toxicity study. No adverse effects on nidation or maternal or fetal survival were found. The number of abnormalities seen in the soft or skeletal tissues did not differ from that occurring spontaneously in controls. The NOAEL of this study amounts to 50 mg/kg bw/day. In a rabbit developmental study on mated female Dutch belted rabbits, the maternal mortality rates were 0, 5, 9 and 19% in the 4 treatment groups (0, 5, 15 or 50 mg/kg bw/day), respectively. A significant increase in extra sternebrae was noted in groups treated at 15 and 50 mg/kg bw/day, but was considered as normal variation by the Panel. The NOAEL of this study is considered to be 15 mg/kg bw/day due to maternal toxicity at the higher dose level.
A clinical study in humans performed in 1960 showed that natamycin, used for systemic mycoses, induced nausea, vomiting and diarrhoea. Anorexia, nausea, vomiting and flatulence were observed at different doses in different patients. The Panel considered that this study is too limited to derive a NOAEL.
In 1968, JECFA established an ADI of 0.3 mg/kg bw/day based on these human data. The level causing no toxicological effects in man was estimated to be 200 mg/per/day, equivalent to 3 mg/kg bw/day. Given that this dose was derived from human data, an uncertainty factor equal to 10 has been used to calculate the ADI. In 2002 JECFA confirmed this ADI.
Because of the limitations in the present database on natamycin (design of the animal studies, limited number of animals, lack of a carcinogenicity study) and in view of the inadequate description of the human data, the Panel considered that an ADI could not be established from these data.
The highest potential exposure to natamycin was at the 97.5th percentile below 0.1 mg/kg bw/day for children and below 0.05 mg/kg bw/day for adults, derived from the high level consumption of cheese (assuming solely a rind treatment with natamycin) and dried, cured sausages.
Given that natamycin is very poorly absorbed, the Panel considers that this conservative estimate would provide an adequate margin of safety from the effect level seen from the long-term studies in animals and the human study used by JECFA to establish an ADI. The Panel considered that the proposed use levels of natamycin are not of safety concern if it is only used for the surface treatment of the rind of semi-hard and semi-soft cheese and on the casings of certain sausages.
The Panel noted that natamycin is used in the food industry as an antifungal preservative in cheeses and sausages. Natamycin is a polyene antibiotic. The mechanism of action for polyene antibiotics is binding to sterols (principally ergosterol) in the fungal cell membrane. Bacteria are insensitive to polyene antibiotics because their membrane lacks sterols. Furthermore, induction of natamycin-resistant mutants in yeast is reported to be difficult.The Panel concluded that there was no concern for the induction of antimicrobial resistance