Re-evaluation of shellac (E 904) as a food additive and a new application on the extension of use of shellac (E 904) in dietary foods for special medical purposes.

The present opinion deals with the re-evaluation of shellac (E 904) when used as a food additive and with the new application on the extension of use of shellac (E 904) in dietary foods for special medical purposes. The Panel derived an acceptable daily intake (ADI) of 4 mg/kg body weight (bw) per day for wax-free shellac (E 904) produced by physical decolouring, based on a NOAEL of 400 mg/kg bw per day and applying an uncertainty factor of 100. The Panel concluded that the ADI of 4 mg/kg bw per day should be considered temporary for wax-free shellac (E 904) produced by chemical bleaching, while new data are generated on the identity and levels of the organochlorine impurities in E 904. This ADI is not applicable for wax-containing shellac as a food additive. For several age groups, the ADI was exceeded at the 95th percentile in the non-brand-loyal exposure assessment scenario and maximum level exposure assessment scenario. Considering the low exceedance and the fact that both the exposure estimation and the toxicological evaluation of shellac were conservative, the panel concluded that the calculated exceedance of the ADI does not indicate a safety concern. The Panel recommended to the European Commission separating specifications for E 904 depending on the manufacturing process, chemical bleaching and physical decolouring, because they result in different impurities; revising the definition of the food additive to include a description of each manufacturing process; deleting information on wax-containing shellac from the EU specifications; revising the acid value for wax-free shellac produced by chemical bleaching; lowering the maximum limit for lead; to consider introducing limits for other toxic elements potentially present in shellac; including a maximum limit for chloroform and total inorganic chloride in the EU specification for shellac produced by chemical bleaching.

Flavouring Group Evaluation 80, Revision 2 (FGE.80Rev2): Consideration of alicyclic, alicyclic-fused and aromatic-fused ring lactones evaluated by the JECFA (61st and 82nd meetings) structurally related to an aromatic lactone evaluated in FGE.27.

The EFSA Panel on Food Additives and Flavourings was requested to evaluate 14 flavouring substances assigned to the Flavouring Group Evaluation 80 (FGE.80), using the Procedure as outlined in the Commission Regulation (EC) No 1565/2000. Thirteen substances have already been considered in FGE.80 and its revision and in FGE.96 [FL-no: 10.005, 10.024, 10.025, 10.050, 10.061, 10.069, 10.070, 10.072, 10.169, 13.009, 13.012, 13.161 and 16.055]. The remaining flavouring substance 3a,4,5,7a-tetrahydro-3,6-dimethylbenzofuran-2(3H)-one [FL-no: 10.057] has been cleared with respect to genotoxicity in FGE.217Rev3 and it is considered in this revision 2 of FGE.80. The substance [FL-no: 10.057] was evaluated through a stepwise approach that integrates information on the structure-activity relationships, intake from current uses, threshold of toxicological concern (TTC) and available data on metabolism and toxicity. The Panel concluded that [FL-no: 10.057] does not give rise to safety concerns at its levels of dietary intake, when estimated on the basis of the 'Maximised Survey-derived Daily Intake' (MSDI) approach. Besides the safety assessment of the flavouring substance, the specifications for the material of commerce have also been considered and the information provided was complete for [FL-no: 10.057]. However, for the flavouring substance [FL-no: 10.057] in the present revision and for eight substances evaluated in previous revisions, the 'modified Theoretical Added Maximum Daily Intakes' (mTAMDIs) values are above the TTC for their structural class (III). For four substances previously evaluated in FGE.80Rev1 and in FGE.96, use levels are still needed to calculate the mTAMDI estimates. Therefore, in total for 13 flavouring substances, data on uses and use levels should be provided to finalise their safety evaluations. For [FL-no: 10.050, 10.069 and 13.161], information on the composition of stereoisomeric mixtures is needed.

Safety of soy leghemoglobin from genetically modified Komagataella phaffii as a food additive.

The EFSA Panel on Food Additive and Flavourings (FAF Panel) provides a scientific opinion on the safety of soy leghemoglobin from genetically modified Komagataella phaffii as a food additive in accordance with Regulation (EC) No 1331/2008. The proposed food additive, LegH Prep, is intended to be used as a colour in meat analogue products. The yeast Komagataella phaffii strain MXY0541 has been genetically modified to produce soy leghemoglobin; the safety of the genetic modification is under assessment by the EFSA GMO Panel (EFSA-GMO-NL-2019-162). The amount of haem iron provided by soy leghemoglobin from its proposed uses in meat analogue products is comparable to that provided by similar amounts of different types of meat. The exposure to iron from the proposed food additive, both at the mean and 95th percentile exposure, will be below the 'safe levels of intake' established by the NDA Panel for all population groups. Considering that the components of the proposed food additive will be digested to small peptide, amino acids and haem B; the recipient (non GM) strain qualifies for qualified presumption of safety status; no genotoxicity concern has been identified and no adverse effects have been identified at the highest dose tested in the available toxicological studies, the Panel concluded that there was no need to set a numerical acceptable daily intake (ADI) and that the food additive does not raise a safety concern at the proposed use in food category 12.9 and maximum use level. The Panel concluded that the use of soy leghemoglobin from genetically modified Komagataella phaffii MXY0541 as a new food additive does not raise a safety concern at the proposed use and use level. This safety evaluation of the proposed food additive remains provisional subject to the ongoing safety assessment of the genetic modification of the production strain by the GMO Panel (EFSA-GMO-NL-2019-162).

Flavouring Group Evaluation 413 (FGE.413): Naringenin.

The EFSA Panel on Food Additives and Flavourings (FAF) was requested to evaluate the safety of naringenin [FL-no: 16.132] as a new flavouring substance, in accordance with Regulation (EC) No 1331/2008. No other substances with sufficient structural similarity have been identified in existing FGEs that could be used to support a read-across approach. The information provided on the manufacturing process, the composition and the stability of [FL-no: 16.132] was considered sufficient. From studies carried out with naringenin, the Panel concluded that there is no concern with respect to genotoxicity. The use of naringenin as a flavouring substance at added portions exposure technique (APET) exposure levels is unlikely to pose a risk for drug interaction. For the toxicological evaluation of naringenin, the Panel requested an extended one-generation toxicity study on naringenin, in line with the requirements of the Procedure and to investigate the consequence of a possible endocrine-disrupting activity. The Panel considered that changes in thymus weight, litter size, post-implantation loss and a consistent reduced pup weight in the high-dose F2 generation could not be dismissed and selected therefore, the mid-dose of 1320 mg/kg body weight (bw) per day for the parental males as the no observed adverse effect level (NOAEL) of the study. The exposure estimates for [FL-no: 16.132] (31,500 and 50,000 µg/person per day for children and adults, respectively) were above the threshold of toxicological of concern (TTC) for its structural class (III). Using the NOAEL of 1320 mg/kg bw per day at step A4 of the procedure, margins of exposure (MoE) of 1590 and 630 could be calculated for adults and children, respectively. Based on the calculated MoEs, the Panel concluded that the use of naringenin as a flavouring substance does not raise a safety concern.

Re-evaluation of guar gum (E 412) as a food additive in foods for infants below 16 weeks of age and follow-up of its re-evaluation as food additive for uses in foods for all population groups.

Guar gum (E 412) was re-evaluated in 2017 by the former EFSA Panel on Food Additives and Nutrient sources added to Food (ANS). As a follow-up to this assessment, the Panel on Food Additives and Flavourings (FAF) was requested to assess the safety of guar gum (E 412) for its uses as food additive in food for infants below 16 weeks of age belonging to food categories 13.1.1 (Infant formulae) and 13.1.5.1 (Dietary foods for infants for special medical purposes and special formulae for infants). In addition, the FAF Panel was requested to address the issues already identified during the re-evaluation of the food additive when used in food for the general population. The process involved the publication of a call for data to allow the interested business operators to provide the requested information to complete the risk assessment. In the response to EFSA requests, one IBO stated that E 412 is not used in food categories 13.1.1 and 13.1.5.1, but it is present in products under food category 13.1.5.2. The Panel concluded that the submitted data are not sufficient to support the safe use of guar gum (E 412) in food for infants (below and above 16 weeks of age) and young children under FC 13.1.1, 13.1.5.1 and 13.1.5.2. Additionally, the Panel concluded that the technical data provided by the IBO support further amendments of the specifications for E 412 laid down in Commission Regulation (EU) No 231/2012.

Flavouring group evaluation 419 (FGE.419): 2-methyl-1-(2-(5-(p-tolyl)-1H-imidazol-2-yl)piperidin-1-yl)butan-1-one.

The EFSA Panel on Food Additives and Flavourings (FAF) was requested to evaluate the safety of 2-methyl-1-(2-(5-(p-tolyl)-1H-imidazol-2-yl)piperidin-1-yl)butan-1-one [FL-no: 16.134] as a new flavouring substance, in accordance with Regulation (EC) No 1331/2008. The substance has not been reported to occur naturally and is chemically synthesised. In food, it is intended to be used as a flavouring substance only in chewing gum. The chronic dietary exposure to [FL-no: 16.134] was estimated to be 45 µg/person per day for a 60-kg adult and 28.4 µg/person per day for a 15-kg 3-year-old child. [FL-no: 16.134] did not show genotoxicity in a bacterial reverse mutation test and an in vitro mammalian cell micronucleus assay. Based on the submitted toxicokinetic and metabolism data, it can be predicted that the flavouring substance is metabolised to innocuous products only. The Panel derived a lower confidence limit of the benchmark dose (BMDL) of 0.71 mg/kg bw per day for a 20% increase in the relative thyroid (including parathyroid) weight observed in a 90-day toxicity study in rats. Based on this BMDL, adequate margins of exposure of 887 and 374 could be calculated for adults and children, respectively. The Panel concluded that there is no safety concern for [FL-no: 16.134], when used as a flavouring substance at the estimated level of dietary exposure, based on the intended use and use levels as specified in Appendix B. The Panel further concluded that the combined exposure to [FL-no: 16.134] from its use as a food flavouring substance and from its presence in toothpaste and mouthwash is also not of safety concern.

Role of albumin in the metabolism and excretion of ochratoxin A.

Ochratoxin A (OTA) is known to be strongly bound to serum albumin, but it remains unknown how albumin affects its metabolism and kinetics. To close this gap, we used a mouse model, where heterozygous albumin deletion reduces serum albumin to concentrations similar to hypoalbuminemic patients and completely eliminates albumin by a homozygous knockout. OTA and its potential metabolites (OTα, 4-OH-OTA, 7'-OH-OTA, OTHQ, OP-OTA, OTB-GSH, OTB-NAC, OTB) were time-dependently analyzed in plasma, bile, and urine by LC-MS/MS and were compared to previously published hepatotoxicity and nephrotoxicity data. Homozygous albumin deletion strongly accelerated plasma clearance as well as biliary and urinary excretion of the parent compound and its hydroxylation products. Decreasing albumin in mice by the heterozygous and even more by the homozygous knockout leads to an increase in the parent compound in urine which corresponded to increased nephrotoxicity. The role of albumin in OTA-induced hepatotoxicity is more complex, since heterozygous but not homozygous nor wild-type mice showed a strong biliary increase in the toxic open lactone OP-OTA. Correspondingly, OTA-induced hepatotoxicity was higher in heterozygous than in wild-type and homozygous animals. We present evidence that albumin-mediated retention of OTA in hepatocytes is required for formation of the toxic OP-OTA, while complete albumin elimination leads to rapid biliary clearance of OTA from hepatocytes with less formation of OP-OTA. In conclusion, albumin has a strong influence on metabolism and toxicity of OTA. In hypoalbuminemia, the parent OTA is associated with increased nephrotoxicity and the open lactone with increased hepatotoxicity.

Integrated data from intravital imaging and HPLC-MS/MS analysis reveal large interspecies differences in AFB1 metabolism in mice and rats.

Large interspecies differences between rats and mice concerning the hepatotoxicity and carcinogenicity of aflatoxin B1 (AFB1) are known, with mice being more resistant. However, a comprehensive interspecies comparison including subcellular liver tissue compartments has not yet been performed. In this study, we performed spatio-temporal intravital analysis of AFB1 kinetics in the livers of anesthetized mice and rats. This was supported by time-dependent analysis of the parent compound as well as metabolites and adducts in blood, urine, and bile of both species by HPLC-MS/MS. The integrated data from intravital imaging and HPLC-MS/MS analysis revealed major interspecies differences between rats and mice: (1) AFB1-associated fluorescence persisted much longer in the nuclei of rat than mouse hepatocytes; (2) in the sinusoidal blood, AFB1-associated fluorescence was rapidly cleared in mice, while a time-dependent increase was observed in rats in the first three hours after injection followed by a plateau that lasted until the end of the observation period of six hours; (3) this coincided with a far stronger increase of AFB1-lysine adducts in the blood of rats compared to mice; (4) the AFB1-guanine adduct was detected at much higher concentrations in bile and urine of rats than mice. In both species, the AFB1-glutathione conjugate was efficiently excreted via bile, where it reached concentrations at least three orders of magnitude higher compared to blood. In conclusion, major differences between mice and rats were observed, concerning the nuclear persistence, formation of AFB1-lysine adducts, and the AFB1-guanine adducts.

Follow-up of the re-evaluation of quillaia extract (E 999) as a food additive and safety of the proposed extension of uses.

Quillaia extract (E 999) was re-evaluated in 2019 by the EFSA Panel on Food Additives and Flavourings (FAF). EFSA derived an acceptable daily intake (ADI) of 3 mg saponins/kg bw per day for E 999. Following a European Commission call for data to submit data to fill the data gaps, the present follow-up opinion assesses data provided by interested business operators (IBOs) to support an amendment of the EU specifications for E 999. Additionally, this opinion deals with the assessment of the proposed extension of use for E 999 in food supplements supplied in a solid and liquid form, excluding food supplements for infants and young children and, as a carrier in botanical nutrients. The Panel concluded that the proposed extension of use, if authorised, could result in an exceedance of the ADI at the maximum of the ranges of the mean for children, adolescents and the elderly, and for all populations at the 95th percentile. An additional proposed extension of use for E 999 to be used as a carrier for glazing agents on entire fresh fruits and vegetables has been received. Since no information on the proposed use levels of E 999 on a saponins content basis has been provided by this applicant, the Panel was not able to evaluate the safety of this extension of use. Considering the technical data submitted, the Panel recommended some modifications of the existing EU specifications for E 999, mainly to lower the limits for lead, mercury and arsenic and to include a maximum limit for cadmium and for calcium oxalate. The Panel also recommended that the limits would be expressed on a saponins basis. The Panel proposed to revise the definition of E 999 to better describe the composition in a qualitative way.

Aflatoxin M1 Analysis in Urine of Mill Workers in Bangladesh: A Pilot Study.

Presence of aflatoxin B1 (AFB1) in food and feed is a serious problem, especially in developing countries. Human exposure to this carcinogenic mycotoxin can occur through dietary intake, but also through inhalation or dermal contact when handling and processing AFB1-contaminated crops. A suitable biomarker of AFB1 exposure by all routes is the occurrence of its hydroxylated metabolite aflatoxin M1 (AFM1) in urine. To assess mycotoxin exposure in mill workers in Bangladesh, we analyzed AFM1 levels in urine samples of this population group who may encounter both dietary and occupational AFB1 exposure. In this pilot study, a total of 76 participants (51 mill workers and 25 controls) were enrolled from the Sylhet region of Bangladesh. Urine samples were collected from people who worked in rice, wheat, maize and spice mills and from controls with no occupational contact to these materials. A questionnaire was used to collect information on basic characteristics and normal food habits of all participants. Levels of AFM1 in the urine samples were determined by a competitive enzyme linked immunosorbent assay. AFM1 was detected in 96.1% of mill workers' urine samples with a range of LOD (40) of 217.7 pg/mL and also in 92% of control subject's urine samples with a range of LOD of 307.0 pg/mL). The mean level of AFM1 in mill workers' urine (106.5 ± 35.0 pg/mL) was slightly lower than that of the control group (123.3 ± 52.4 pg/mL), whilst the mean AFM1 urinary level adjusted for creatinine was higher in mill workers (142.1 ± 126.1 pg/mg crea) than in the control group (98.5 ± 71.2 pg/mg crea). Yet, these differences in biomarker levels were not statistically significant. Slightly different mean urinary AFM1 levels were observed between maize mill, spice mill, rice mill, and wheat mill workers, yet biomarker values are based on a small number of individuals in these subgroups. No significant correlations were found between the study subjects' urine AFM1 levels and their consumption of some staple food items, except for a significant correlation observed between urinary biomarker levels and consumption of groundnuts. In conclusion, this pilot study revealed the frequent presence of AFM1 in the urine of mill workers in Bangladesh and those of concurrent controls with dietary AFB1 exposure only. The absence of a statistical difference in mean biomarker levels for workers and controls suggests that in the specific setting, no extra occupational exposure occurred. Yet, the high prevalence of non-negligible AFM1 levels in the collected urines encourage further studies in Bangladesh regarding aflatoxin exposure.

Re-evaluation of erythritol (E 968) as a food additive.

This opinion addresses the re-evaluation of erythritol (E 968) as food additive and an application for its exemption from the laxative warning label requirement as established under Regulation (EU) No 1169/2011. Erythritol is a polyol obtained by fermentation with Moniliella pollinis BC or Moniliella megachiliensis KW3-6, followed by purifications and drying. Erythritol is readily and dose-dependently absorbed in humans and can be metabolised to erythronate to a small extent. Erythritol is then excreted unchanged in the urine. It does not raise concerns regarding genotoxicity. The dataset evaluated consisted of human interventional studies. The Panel considered that erythritol has the potential to cause diarrhoea in humans, which was considered adverse because its potential association with electrolyte and water imbalance. The lower bound of the range of no observed adverse effect levels (NOAELs) for diarrhoea of 0.5 g/kg body weight (bw) was identified as reference point. The Panel considered appropriate to set a numerical acceptable daily intake (ADI) at the level of the reference point. An ADI of 0.5 g/kg bw per day was considered by the Panel to be protective for the immediate laxative effect as well as potential chronic effects, secondary to diarrhoea. The highest mean and 95th percentile chronic exposure was in children (742 mg/kg bw per day) and adolescents (1532 mg/kg bw per day). Acute exposure was maximally 3531 mg/kg bw per meal for children at the 99th percentile. Overall, the Panel considered both dietary exposure assessments an overestimation. The Panel concluded that the exposure estimates for both acute and chronic dietary exposure to erythritol (E 968) were above the ADI, indicating that individuals with high intake may be at risk of experiencing adverse effects after single and repeated exposure. Concerning the new application, the Panel concluded that the available data do not support the proposal for exemption.

Safety evaluation of the food additive steviol glycosides, predominantly Rebaudioside M, produced by fermentation using Yarrowia lipolyticaVRM.

The EFSA Panel on Food Additive and Flavourings (FAF Panel) provides a scientific opinion on the safety of a new process to produce steviol glycosides by fermentation of simple sugars using a genetically modified strain of Yarrowia lipolytica (named Y. lipolytica VRM). The manufacturing process may result in impurities different from those that may be present in the other steviol glycosides E 960a-d, therefore the Panel concluded that separate specifications are required for the food additive produced as described in the current application. Viable cells and DNA from the production strain are not present in the final product. The Panel considered that the demonstration of the absence of kaurenoic acid in the proposed food additive, using a method with a limit of detection (LOD) of 0.3 mg/kg, is adequate to dispel the concerns for potential genotoxicity. Given that all steviol glycosides follow the same metabolic pathways, the Panel considered that the current steviol glycosides would fall within the same group of substances. Therefore, the Panel considered that the already existing data on rebaudioside M and structurally related steviol glycosides are sufficient, and a similar metabolic fate and toxicity is expected for the food additive. The results from the bacterial reverse mutation assay and the in vitro micronucleus assay were negative and indicated absence of genotoxicity from the food additive. The existing acceptable daily intake (ADI) of 4 mg/kg body weight (bw) per day, expressed as steviol equivalents, was considered to be applicable to the proposed food additive. The Panel concluded that there is no safety concern for steviol glycosides, predominantly Rebaudioside M, produced by fermentation using Y. lipolytica VRM, to be used as a food additive at the proposed uses and use levels.

Safety evaluation of synthesised DNA oligonucleotides as a food additive.

The EFSA Panel on Food Additives and Flavourings (FAF) was requested to evaluate the safety of synthesised DNA oligonucleotides as a new food additive, in accordance with Regulation (EC) No 1331/2008. Considering that the additional information requested by the Panel during the risk assessment was not provided by the applicant, the assessment was concluded on the basis of the sole information available in the application. The proposed food additive consists of purified synthetic DNA sequences intended to be used for traceability purposes, alone or combined with carriers. Information provided by the applicant on the identity, characterisation and production process of the proposed food additive was considered insufficient. The Panel considered that the product specifications as proposed by the applicant do not adequately define and characterise the proposed food additive. The applicant proposed for the food additive the maximum use levels of 0.001 mg/kg for a variety of food categories. The food additive was also proposed as a Group I additive at a specific maximum level of quantum satis. The applicant did not provide exposure estimates according to the EFSA ANS Panel guidance (2012). No biological or toxicological data were provided by the applicant for the proposed food additive. Considering the inadequate information available and the uncertainty introduced by the proposal at quantum satis, along with the insufficient specifications, the Panel could not conclude on the safety of the food additive as proposed and described by the applicant.

Umbrales para las clases específicas de carcinógenos genotóxicos. nueva estrategia para la categorización de la carcinogenicidad de las sustancias químicas / Thresholds for specific classes of genotoxic carcinogens: a new strategy for carcinogenicity categorisation of chemicals

Cada vez está más claro el principio general de la secuencia de acontecimientos que finalmente conducen al cáncer después de la exposición a carcinógenos genotóxicos. Esto ayuda a conocer los parámetros que influyen en la forma de la curva dosis-efecto para la carcinogénesis, que incluyen activación e inactivación metabólica de carcinógenos, reparación del ADN, control del ciclo celular, proliferación regenerativa, apoptosis, senescencia inducida por oncogenes y control por el sistema inmunitario. Una relación lineal dosis-respuesta sin umbral observable parece ser una descripción conservadora pero adecuada para la actividad carcinógena de muchos carcinógenos genotóxicos, por ejemplo, la aflatoxina B1. Sin embargo, algunos modelos de extrapolación lineal que conectan el riesgo de alto nivel a la intersección en el cero han conducido a predicciones erróneas. En esta revisión se demuestra que el acetato de vinilo es un ejemplo de carcinógeno que actúa a través de un mecanismo de umbral. En los tejidos de contacto, el acetato de vinilo es convertido en ácido acético y acetaldehído. Sólo cuando se alcanzan las concentraciones umbral se activa el mecanismo que finalmente conduce al cáncer, es decir una reducción del pH mayor de 0.15 unidad que conduce a citotoxicidad, daño del ADN y proliferación regenerativa. En esta revisión se presenta un nuevo sistema de categorización de los carcinógenos que tiene en cuenta que pueden actuar por mecanismos de umbral.

The general principle of the sequence of events that finallylead to cancer after exposure to genotoxic carcinogenshas become increasingly clear. This helps to understandthe parameters that influence the shape of the dose effectcurve for carcinogenesis, including metabolic activationand inactivation of carcinogens, DNA repair, cell cyclecontrol, regenerative proliferation, apoptosis, oncogeneinducedsenescence and control by the immune system.A linear dose response relationship with no observablethreshold seems to be a conservative but adequatedescription for the carcinogenic activity of many genotoxiccarcinogens, such as for instance aflatoxin B1. However,some linear extrapolation models connecting the highlevelrisk to the zero intercept have resulted in wrongpredictions. In this review we demonstrate that vinylacetate is an example of a carcinogen acting by athreshold mechanism. In tissues of contact vinyl acetateis converted to acetic acid and acetaldehyde. Only whenthreshold levels are achieved critical steps in themechanism that ultimately leads to cancer become active,namely pH reduction of more than 0.15 units leadingto cytotoxicity, damage to DNA and regenerativeproliferation. In this review we present a new system ofcarcinogen categorisation taking into account thatcarcinogens may act by threshold mechanisms.