Dietary glycation compounds - implications for human health.

The term "glycation compounds" comprises a wide range of structurally diverse compounds that are formed endogenously and in food via the Maillard reaction, a chemical reaction between reducing sugars and amino acids. Glycation compounds produced endogenously are considered to contribute to a range of diseases. This has led to the hypothesis that glycation compounds present in food may also cause adverse effects and thus pose a nutritional risk to human health. In this work, the Senate Commission on Food Safety (SKLM) of the German Research Foundation (DFG) summarized data on formation, occurrence, exposure and toxicity of glycation compounds (Part A) and systematically assessed potential associations between dietary intake of defined glycation compounds and disease, including allergy, diabetes, cardiovascular and renal disease, gut/gastrotoxicity, brain/cognitive impairment and cancer (Part B). A systematic search in Pubmed (Medline), Scopus and Web of Science using a combination of keywords defining individual glycation compounds and relevant disease patterns linked to the subject area of food, nutrition and diet retrieved 253 original publications relevant to the research question. Of these, only 192 were found to comply with previously defined quality criteria and were thus considered suitable to assess potential health risks of dietary glycation compounds. For each adverse health effect considered in this assessment, however, only limited numbers of human, animal and in vitro studies were identified. While studies in humans were often limited due to small cohort size, short study duration, and confounders, experimental studies in animals that allow for controlled exposure to individual glycation compounds provided some evidence for impaired glucose tolerance, insulin resistance, cardiovascular effects and renal injury in response to oral exposure to dicarbonyl compounds, albeit at dose levels by far exceeding estimated human exposures. The overall database was generally inconsistent or inconclusive. Based on this systematic review, the SKLM concludes that there is at present no convincing evidence for a causal association between dietary intake of glycation compounds and adverse health effects.

Considering the implication of endogenous glycation compounds in aging and disease, dietary exposure via consumption of an "AGE (advanced glycation end product) rich diet" is increasingly suggested to pose a potential health risk. However, studies attempting to assess an association between dietary glycation compounds and adverse health effects frequently suffer from insufficient chemical analysis of glycation compounds, including inadequate structural characterization and limited quantitative data. The Senate Commission on Food Safety (SKLM) of the German Research Foundation (DFG) previously defined quality criteria for studies designed to assess the effects of dietary glycation compounds on human health. The aim of the present work is to summarize data on formation, occurrence, exposure and toxicity of glycation compounds (Part A) and to systematically evaluate if the currently available scientific database allows for a conclusive assessment of potential health effects of defined glycation compounds (Part B).The term "glycation compounds" comprises a wide range of structurally diverse compounds that derive from the Maillard reaction, a chemical reaction between reducing carbohydrates and amino compounds that occurs during food processing. In the first stage of the Maillard reaction, reducing sugars such as glucose and fructose react for instance with the ε-amino group of lysine, which is most abundant in food ("glycation" of lysine). Subsequently, these primary reaction products undergo Amadori rearrangement to yield products (ARP) such as fructosyllysine (FL) from glucose and also Heyns rearrangement products (HRPs) such as glucosyl- and mannosyllysine from fructose. While ARPs are rapidly formed during food processing, they are not stable and undergo degradation reactions, predominantly to 1,2-dicarbonyl compounds such as glyoxal (GO), methylglyoxal (MGO) and 3-deoxyglucosone (3-DG), which are highly reactive. The last stage of the Maillard reaction is characterized predominantly by the reaction of these dicarbonyl compounds with nucleophilic groups of proteins. The side-chains of lysine and arginine residues as well as the N-termini of proteins are important reaction sites. Carboxyalkylated amino acids such as N-ε-carboxymethyllysine (CML) and N-ε-carboxyethyllysine (CEL) result from reaction of the ε-amino group of lysine with the dicarbonyl compounds GO and MGO. Dicarbonyl compounds with C5 or C6 chains can form cyclic pyrrole derivatives at the ε-amino group of lysine. The most important example for this reaction is pyrraline, which is formed from reaction of 3-DG and lysine. The reaction of dicarbonyl compounds with the guanidino group of arginine mainly leads to hydroimidazolones, of which the MGO-derived hydroimidazolone 1 (MG-H1) is best described in food systems.ARPs are the most abundant glycation products found in food. Up to 55% of the lysine residues in food may be modified to ARPs at the side-chain. Food items particularly rich in ARPs include bread, rusk, biscuits, chocolate, and powdered infant formulas. Exposure estimates range between 0.6­1.6 mg/kg body weight (bw), although exposure may be as high as 14.3 mg/kg bw in individuals consuming foods with extreme ARP concentrations. Foods particularly rich in dicarbonyl compounds include heat-treated or long-term stored items rich in reducing sugars such as jams, alternative sweeteners, soft drinks, honey, candies, cookies, and vinegars, especially balsamico-type vinegars. The main contributors to the daily intake of MGO, GO, and 3-DG are coffee and bread. Dietary exposure to dicarbonyl compounds has been estimated to range between 0.02­0.29 mg/kg bw/d for MGO, 0.04­0.16 mg/kg bw/d for GO, 0.14­2.3 mg/kg bw/d for 3-DG, and 0.08­0.13 mg/kg bw/d for 3-deoxygalactosone (3-DGal). Dietary intake of 5-hydroxymethylfurfural (HMF), which can be formed from 3-DG, is estimated to range between 0.0001­0.9 mg/kg bw/d. Exposure estimates for individual glycated amino acids range from 0.03­0.35 mg/kg bw/d for CML, 0.02­0.04 mg/kg bw/d for CEL and 0.19­0.41 mg/kg bw/d for MG-H1. From a model diet consisting of 1 L milk, 500 g bakery products and 400 mL coffee, an intake of pyrraline corresponding to 0.36 mg/kg bw/d for a 70 kg person was estimated.Quantitative analysis of individual glycation compounds or their metabolites in tissues or body fluids as well as their reaction products with amino acids, proteins or DNA may serve to monitor exposure to glycation compounds. However, since glycation compounds are also formed endogenously, these biomarkers reflect the totality of the exposure, making it inherently difficult to define the body burden due to dietary intake against the background of endogenous formation.Information on the toxicokinetics and toxicity of glycation compounds is scarce and mostly limited to the reactive dicarbonyl compounds GO, MGO, 3-DG, HMF, and individual glycated amino acids such as CML and CEL. Acute toxicity of dicarbonyl compounds is low to moderate. There are some data to suggest that rapid detoxification of dicarbonyls in the gastrointestinal tract and liver may limit their oral bioavailability. Biotransformation of GO and MGO occurs predominantly via the glutathione (GSH)-dependent glyoxalase system, and to a lesser extent via glutathione-independent aldo-keto-reductases, which are also responsible for biotransformation of 3-DG. GO, MGO and 3-DG readily react with DNA bases in vitro, giving rise to DNA adducts. There is clear evidence for genotoxicity of GO, MGO and 3-DG. Repeated dose toxicity studies on GO consistently reported reduced body weight gain concomitant with reduced food and water consumption but did not identify compound related changes in clinical chemistry and hematology or histopathological lesions. There is also no evidence for systemic carcinogenicity of GO and MGO based on the available studies. However, initiation/promotion studies indicate that oral exposure to GO may exhibit genotoxic and tumor promoting activity locally in the gastrointestinal tract. From a 2-year chronic toxicity and carcinogenicity study in rats, a NOAEL for systemic toxicity of GO administered via drinking water of 25 mg/kg bw was reported based on reduced body weight and erosions/ulcer in the glandular stomach. Other non-neoplastic and neoplastic lesions were not observed. Acute toxicity of HMF is also low. From a 90-day repeated dose toxicity study in mice, a NOAEL of 94 mg/kg bw was derived based on cytoplasmic alterations of proximal tubule epithelial cells of the kidney. HMF was mostly negative in in vitro genotoxicity tests, although positive findings for mutagenicity were obtained under conditions that promote formation of the chemically reactive sulfuric acid ester 5-sulfoxymethylfurfural. There is some evidence of carcinogenic activity of HMF in female B6C3F1 mice based on increased incidences of hepatocellular adenoma, but not in male mice and rats of both sexes. Although data on oral bioavailability of glycated amino acids are mostly limited to CML, it appears that glycated amino acids may be absorbed from the gastrointestinal tract after oral exposure to their free and protein bound form. Glycated amino acids that are not absorbed in the intestine may be subject to metabolism by the gut microbiome. Glycated amino acids present in the systemic circulation are rapidly eliminated via the urine. Acute oral toxicity of CML is low. Studies in mice and rats reported changes in clinical chemistry parameters indicative of impaired renal and hepatic function. However, these changes were not dose-related and not supported by histopathological evaluation.Previous risk assessments of individual glycation compounds did not identify a health concern at estimated human exposures (GO, HMF) but also noted the lack of data to draw firm conclusions on health risks associated with exposure to MGO.To identify potential associations between dietary intake of defined glycation compounds and disease a systematic review was carried out according to the Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) model, applying the quality criteria previously defined by the SKLM. Using a combination of keywords defining individual glycation compounds and relevant disease patterns linked to the subject area of food, nutrition and diet, a systematic search in Pubmed (Medline), Scopus and Web of Science was performed. Although the present systematic review identified numerous studies that investigated an association between an "AGE-rich diet" and adverse health effects, only a subset of studies was found to comply with the quality criteria defined by the SKLM and was thus considered suitable to assess potential health risks of dietary glycation compounds.For each adverse health effect considered in this assessment, only limited numbers of human studies were identified. Although studies in humans offer the advantage of investigating effects at relevant human exposures, these studies did not provide compelling evidence for adverse effects of dietary glycation compounds. Animal studies identified in this systematic review provide some evidence for induction of impaired glucose tolerance, insulin resistance, cardiovascular effects and renal injury in response to oral exposure to GO and MGO as representatives of dicarbonyl compounds. Only limited evidence points to a link between high intake of glycated amino acids and metabolic disorders. However, these effects were typically reported to occur at dose levels that exceed human dietary exposure, often by several orders of magnitude. Unfortunately, most studies employed only one dose level, precluding characterization of dose-response and derivation of a point of departure for riskassessment. While in vitro studies provide some evidence for a potential mechanistic link between individual glycation compounds and presumed adverse health effects, the clinical and toxicological relevance of the in vitro findings is often limited by the use of high concentrations of glycation compounds that by far exceed human dietary exposure and by insufficient evidence for corresponding adverse effects in vivo. A key question that has not been adequately considered in most studies investigating systemic effects of glycation compounds is the extent of oral bioavailability of dietary glycation compounds, including the form in which MRPs may be taken up (e.g. free vs. peptide bound glycated amino acids). Understanding how much dietary glycation compounds really add to the significant endogenous background is critical to appraise the relevance of dietary MRPs for human health.While it appears mechanistically plausible that glycation of dietary allergens may affect their allergenic potential, the currently available data do not support the hypothesis that dietary glycation compounds may increase the risk for diet-induced allergies. There are no human studies addressing the immunological effects of dietary AGEs. Accordingly, there are no data on whether dietary AGEs promote the development of allergies, nor whether existing allergies are enhanced or attenuated. In numerous in vitro studies, the IgG/E binding ability of antigens and therefore their allergenic potential has been predominantly reported to be reduced by glycation. However, some in vitro studies showed that glycated proteins bind to receptors of immunological cells, and thus may have promoting effects on immune response and inflammation.Although experimental data from animal studies provide some evidence that high doses of individual glycation compounds such as MGO and protein-bound CML may produce certain adverse health effects, including diabetogenic, cardiovascular, metabolic and renal effects, the doses required to achieve these effects by far exceed human dietary exposures. Of note, in the only long-term study identified, a high dose of MGO administered via drinking water to mice for 18 months had no adverse effects on the kidneys, cardiovascular system, or development of diabetes.Experimental data from animal studies provide evidence that high doses of defined glycation compounds such as MGO or protein-bound CML may affect glucose homeostasis. However, the doses required to produce these effects markedly exceed human dietary exposure. Results from human studies are inconclusive: Three short-term intervention studies suggested that diets rich in AGEs may impair glucose homeostasis, whereas one recent intervention study and two observational studies failed to show such an effect.For the cardiovascular system, there is some evidence from in vitro and in vivo studies that high concentrations of MRPs, well above the dietary exposure of humans, may enhance inflammation in the cardiovascular system, induce endothelial damage, increase blood pressure and increase the risk of thrombosis. Only a limited number of human intervention studies investigated potential effects of short-term exposure and longer-term effects of glycation compounds on the cardiovascular system, and yielded inconsistent results. The few observational studies available either found no association between dietary MRP intake and cardiovascular function or even reported beneficial effects. Therefore, currently no definitive conclusion on potential acute and chronic effects of dietary MRPs on inflammation and cardiovascular function can be drawn. However, there is currently also no convincing evidence that potential adverse effects on the cardiovascular system are triggered by dietary MRP intake.Furthermore, human studies did not provide evidence for an adverse effect of dietary MRPs on kidney function. In animal studies with high levels of oral intake, MGO was reported to cause structural and functional effects in the kidney. Several studies show that the concentration of modified proteins and amino acids, such as CML, increases significantly in kidney tissue after oral intake. One study showed a negative effect of a high-temperature-treated diet containing increased CML concentrations on kidney structure integrity and impaired glomerular filtration. The causative relationship of accumulation of dietary MRPs and a functional decline of the kidneys, however, needs further confirmation.With regard to gut health, there is some evidence for alterations in gut microflora composition and the production of individual short-chain fatty acids (SCFAs) upon dietary exposure to glycation compounds. However, this has not been linked to adverse health effects in humans and may rather reflect adaptation of the gut microbiota to changing nutrients. In particular, a human observational study and several animal studies did not find a correlation between the intake of glycation compounds and increased intestinal inflammation. In animal studies, positive effects of glycation compounds on gut tissue damage and dysbiosis during colitis were described.Considering clear evidence for DNA reactivity and genotoxicity of the dicarbonyl compounds GO, MGO and 3-DG, it is plausible to suspect that dicarbonyl compounds may induce mutations and cancer. Although there is some evidence for tumor promoting activity of GO locally in the gastrointestinal tract, the only guideline-compatible chronic rodent bioassays reported erosions and ulcer in the glandular stomach but no treatment-related neoplastic lesions. A recent multinational cohort study with focus on CEL, CML, and MG-H1 found no evidence to support the hypothesis that dietary AGEs are linked to cancer risk.Evidence for an association between human exposure to dietary glycation compounds and detrimental effects on the brain and on cognitive performance is far from being compelling. No human studies fully complying with the defined quality criteria were identified. A few experimental studies reported neuroinflammation and cognitive impairment following dietary MRP exposure, but these can be considered indicative at best and do not support firm conclusions for human health. In addition to utilizing exceedingly high dosages of individual agents like CML, harsh processing conditions causing a multitude of major process-related changes do not allow to convincingly reconcile effects observed with measured/supposed contents of free and protein-bound CML alone.Overall, although dietary glycation compounds have been claimed to contribute to a wide range of adverse health effects, the present critical evaluation of the literature allows the conclusion that the available data are insufficient, inadequate or inconclusive and do not compellingly support the hypothesis of human health risks being related to the presence of glycation compounds in food. The study limitations detailed above, together with the fact that a large number of studies did not comply with the defined quality criteria and therefore had to be excluded highlight the importance of performing adequately designed human or animal studies to inform scientifically reliable health risk assessment.To achieve this, high quality, dependable scientific cooperation within various disciplines is pivotal.

Controversy on health-based guidance values for bisphenol A-the need of criteria for studies that serve as a basis for risk assessment.

Since 2006, the responsible regulatory bodies have proposed five health-based guidance values (HBGV) for bisphenol A (BPA) that differ by a factor of 250,000. This range of HBGVs covers a considerable part of the range from highly toxic to relatively non-toxic substances. As such heterogeneity of regulatory opinions is a challenge not only for scientific risk assessment but also for all stakeholders, the Senate Commission on Food Safety (SKLM) of the German Research Foundation (DFG) analyzed the reasons for the current discrepancy and used this example to suggest improvements for the process of HBGV recommendations. A key aspect for deriving a HBGV is the selection of appropriate studies that allow the identification of a point of departure (PoD) for risk assessment. In the case of BPA, the HBGV derived in the 2023 EFSA assessment was based on a study that reported an increase of Th17 cells in mice with a benchmark dose lower bound (BMDL40) of 0.53 µg/kg bw/day. However, this study does not comply with several criteria that are important for scientific risk assessment: (1) the selected end-point, Th17 cell frequency in the spleen of mice, is insufficiently understood with respect to health outcomes. (2) It is unclear, by which mechanism BPA may cause an increase in Th17 cell frequency. (3) It is unknown, if an increase of Th17 cell frequency in rodents is comparably observed in humans. (4) Toxicokinetics were not addressed. (5) Neither the raw data nor the experimental protocols are available. A further particularly important criterion (6) is independent data confirmation which is not available in the present case. Previous studies using other readouts did not observe immune-related adverse effects such as inflammation, even at doses orders of magnitude higher than in the Th17 cell-based study. The SKLM not only provides here key criteria for the use of such studies, but also suggests that the use of such a "checklist" requires a careful and comprehensive scientific judgement of each item. It is concluded that the Th17 cell-based study data do not represent an adequate basis for risk assessment of BPA.

Commentary of the SKLM to the EFSA opinion on risk assessment of N-nitrosamines in food.

Dietary exposure to N-nitrosamines has recently been assessed by the European Food Safety Authority (EFSA) to result in margins of exposure that are conceived to indicate concern with respect to human health risk. However, evidence from more than half a century of international research shows that N-nitroso compounds (NOC) can also be formed endogenously. In this commentary of the Senate Commission on Food Safety (SKLM) of the German Research Foundation (DFG), the complex metabolic and physiological biokinetics network of nitrate, nitrite and reactive nitrogen species is discussed with emphasis on its influence on endogenous NOC formation. Pioneering approaches to monitor endogenous NOC have been based on steady-state levels of N-nitrosodimethylamine (NDMA) in human blood and on DNA adduct levels in blood cells. Further NOC have not been considered yet to a comparable extent, although their generation from endogenous or exogenous precursors is to be expected. The evidence available to date indicates that endogenous NDMA exposure could exceed dietary exposure by about 2-3 orders of magnitude. These findings require consolidation by refined toxicokinetics and DNA adduct monitoring data to achieve a credible and comprehensive human health risk assessment.

Acetaldehyde as a Food Flavoring Substance: Aspects of Risk Assessment.

The Senate Commission on Food Safety (SKLM) of the German Research Foundation (DFG) has reviewed the currently available data in order to assess the health risks associated with the use of acetaldehyde as a flavoring substance in foods. Acetaldehyde is genotoxic in vitro. Following oral intake of ethanol or inhalation exposure to acetaldehyde, systemic genotoxic effects of acetaldehyde in vivo cannot be ruled out (induction of DNA adducts and micronuclei). At present, the key question of whether acetaldehyde is genotoxic and mutagenic in vivo after oral exposure cannot be answered conclusively. There is also insufficient data on human exposure. Consequently, it is currently not possible to reliably assess the health risk associated with the use of acetaldehyde as a flavoring substance. However, considering the genotoxic potential of acetaldehyde as well as numerous data gaps that need to be filled to allow a comprehensive risk assessment, the SKLM considers that the use of acetaldehyde as a flavoring may pose a safety concern. For reasons of precautionary consumer protection, the SKLM recommends that the scientific base for approval of the intentional addition of acetaldehyde to foods as a flavoring substance should be reassessed.

FEMA GRAS assessment of natural flavor complexes: Sage oil, Orris Root Extract and Tagetes Oil and related flavoring ingredients.

In recent years, the Expert Panel of the Flavor and Extract Manufacturers Association (FEMA) has conducted a program to re-evaluate the safety of natural flavor complexes (NFCs) used as flavor ingredients. This publication, twelfth in the series, details the re-evaluation of NFCs whose constituent profiles are characterized by alicyclic or linear ketones. In its re-evaluation, the Expert Panel applies a scientific constituent-based procedure for the safety evaluation of NFCs in commerce using a congeneric group approach. Estimated intakes of each congeneric group of the NFC are evaluated using the well-established and conservative Threshold of Toxicological Concern (TTC) approach. In addition, studies on the toxicity and genotoxicity of members of the congeneric groups and the NFCs under evaluation are reviewed. The scope of the safety evaluation of the NFCs contained herein does not include added use in dietary supplements or any products other than food. Thirteen (13) NFCs derived from the Boronia, Cinnamomum, Thuja, Ruta, Salvia, Tagetes, Hyssopus, Iris, Perilla and Artemisia genera are affirmed as generally recognized as safe (GRAS) under conditions of their intended use as flavor ingredients based on an evaluation of each NFC and the constituents and congeneric groups therein.

FEMA GRAS assessment of natural flavor complexes: Lemongrass oil, chamomile oils, citronella oil and related flavoring ingredients.

In 2015, the Expert Panel of the Flavor and Extract Manufacturers Association (FEMA) initiated a program for the re-evaluation of the safety of over 250 natural flavor complexes (NFCs) used as flavor ingredients. This publication, eleventh in the series, evaluates the safety of NFCs characterized by primary alcohol, aldehyde, carboxylic acid, ester and lactone constituents derived from terpenoid biosynthetic pathways and/or lipid metabolism. The scientific-based evaluation procedure published in 2005 and updated in 2018 that relies on a complete constituent characterization of the NFC and organization of the constituents into congeneric groups. The safety of the NFCs is evaluated using the threshold of toxicological concern (TTC) concept in addition to data on estimated intake, metabolism and toxicology of members of the congeneric groups and for the NFC under evaluation. The scope of the safety evaluation does not include added use in dietary supplements or any products other than food. Twenty-three NFCs, derived from the Hibiscus, Melissa, Ricinus, Anthemis, Matricaria, Cymbopogon, Saussurea, Spartium, Pelargonium, Levisticum, Rosa, Santalum, Viola, Cryptocarya and Litsea genera were affirmed as generally recognized as safe (GRAS) under their conditions of intended use as flavor ingredients based on an evaluation of each NFC and the constituents and congeneric groups therein.

FEMA GRAS assessment of natural flavor complexes: Allspice, anise, fennel-derived and related flavoring ingredients.

The FEMA Expert Panel program to re-evaluate the safety of natural flavor complexes (NFCs) used as flavoring ingredients in food has resulted in the publication of an updated constituent-based procedure as well as publications on the safety evaluation of many botanical-derived NFCs. This publication, ninth in the series and related to the ninth publication, describes the affirmation of the generally recognized as safe (GRAS) status for NFCs with propenylhydroxybenzene and allylalkoxybenzene constituents under their conditions of intended use as flavoring ingredients added to food. The Panel's procedure applies the threshold of toxicological concern (TTC) concept and evaluates relevant data on absorption, metabolism, genotoxic potential and toxicology for the NFCs themselves and their respective constituent congeneric groups. For NFCs containing allylalkoxybenzene constituent(s) with suspected genotoxic potential, the estimated intake of the individual constituent is compared to the TTC for compounds with structural alerts for genotoxicity and if exceeded, a margin of exposure is calculated using BMDL10 values derived from benchmark dose analyses using Bayesian model averaging, as presented in the tenth article of the series. Safety evaluations for NFCs derived from allspice, anise seed, star anise, sweet fennel seed and pimento leaves were conducted and their GRAS status was affirmed for use as flavoring ingredients. The scope of the safety evaluation contained herein does not include added use in dietary supplements or any products other than food.

FEMA GRAS assessment of derivatives of basil, nutmeg, parsley, tarragon and related allylalkoxybenzene-containing natural flavor complexes.

In 2015, the Expert Panel of the Flavor and Extract Manufacturers Association (FEMA) initiated a program for the re-evaluation of the safety of over 250 natural flavor complexes (NFCs) used as flavoring ingredients in food. In this publication, tenth in the series, NFCs containing a high percentage of at least one naturally occurring allylalkoxybenzene constituent with a suspected concern for genotoxicity and/or carcinogenicity are evaluated. In a related paper, ninth in the series, NFCs containing anethole and/or eugenol and relatively low percentages of these allylalkoxybenzenes are evaluated. The Panel applies the threshold of toxicological concern (TTC) concept and evaluates relevant toxicology data on the NFCs and their respective constituent congeneric groups. For NFCs containing allylalkoxybenzene constituent(s), the estimated intake of the constituent is compared to the TTC for compounds with structural alerts for genotoxicity and when exceeded, a margin of exposure (MOE) is calculated. BMDL10 values are derived from benchmark dose analyses using Bayesian model averaging for safrole, estragole and methyl eugenol using EPA's BMDS software version 3.2. BMDL10 values for myristicin, elemicin and parsley apiole were estimated by read-across using relative potency factors. Margins of safety for each constituent congeneric group and MOEs for each allylalkoxybenzene constituent for each NFC were determined that indicate no safety concern. The scope of the safety evaluation contained herein does not include added use in dietary supplements or any products other than food. Ten NFCs, derived from basil, estragon (tarragon), mace, nutmeg, parsley and Canadian snakeroot were determined or affirmed as generally recognized as safe (GRAS) under their conditions of intended use as flavor ingredients based on an evaluation of each NFC and the constituents and congeneric groups therein.

FEMA GRAS assessment of natural flavor complexes: Asafetida oil, garlic oil and onion oil.

The Expert Panel of the Flavor and Extract Manufacturers Association (FEMA) applies its procedure for the safety evaluation of natural flavor complexes (NFCs) to re-evaluate the safety of Asafetida Oil (Ferula assa-foetida L.) FEMA 2108, Garlic Oil (Allium sativum L.) FEMA 2503 and Onion Oil (Allium cepa L.) FEMA 2817 for use as flavoring in food. This safety evaluation is part of a series of evaluations of NFCs for use as flavoring ingredients conducted by the Expert Panel that applies a scientific procedure published in 2005 and updated in 2018. Using a group approach that relies on a complete chemical characterization of the NFC intended for commerce, the constituents of each NFC are organized into well-defined congeneric groups and the estimated intake of each constituent congeneric group is evaluated using the conservative threshold of toxicological concern (TTC) concept. Data on the metabolism, genotoxic potential and toxicology for each constituent congeneric group are reviewed as well as studies on each NFC. Based on the safety evaluation, Asafetida Oil (Ferula assa-foetida L.), Garlic Oil (Allium sativum L.) and Onion Oil (Allium cepa L.) were affirmed as generally recognized as safe (GRASa) under their conditions of intended use as flavor ingredients.

Evaluation of the genotoxic potential of acrylamide: Arguments for the derivation of a tolerable daily intake (TDI value).

This opinion of the Senate Commission on Food Safety (SKLM) of the German Research Foundation (Deutsche Forschungsgemeinschaft, DFG) presents arguments for an updated risk assessment of diet-related exposure to acrylamide (AA), based on a critical review of scientific evidence relevant to low dose exposure. The SKLM arrives at the conclusion that as long as an appropriate exposure limit for AA is not exceeded, genotoxic effects resulting in carcinogenicity are unlikely to occur. Based on the totality of the evidence, the SKLM considers it scientifically justified to derive a tolerable daily intake (TDI) as a health-based guidance value.

Salivary nitrate/nitrite and acetaldehyde in humans: potential combination effects in the upper gastrointestinal tract and possible consequences for the in vivo formation of N-nitroso compounds-a hypothesis.

Subsequent to the dietary uptake of nitrate/nitrite in combination with acetaldehyde/ethanol, combination effects resulting from the sustained endogenous exposure to nitrite and acetaldehyde may be expected. This may imply locoregional effects in the upper gastrointestinal tract as well as systemic effects, such as a potential influence on endogenous formation of N-nitroso compounds (NOC). Salivary concentrations of the individual components nitrate and nitrite and acetaldehyde are known to rise after ingestion, absorption and systemic distribution, thereby reflecting their respective plasma kinetics and parallel secretion through the salivary glands as well as the microbial/enzymatic metabolism in the oral cavity. Salivary excretion may also occur with certain drug molecules and food constituents and their metabolites. Therefore, putative combination effects in the oral cavity and the upper digestive tract may occur, but this has remained largely unexplored up to now. In this Guest Editorial, published evidence on exposure levels and biokinetics of nitrate/nitrite/NOx, NOC and acetaldehyde in the organism is reviewed and knowledge gaps concerning combination effects are identified. Research is suggested to be initiated to study the related unresolved issues.

The BlueScreen HC assay to predict the genotoxic potential of fragrance materials.

BlueScreen HC is a mammalian cell-based assay for measuring the genotoxicity and cytotoxicity of chemical compounds and mixtures. The BlueScreen HC assay has been utilized at the Research Institute for Fragrance Materials in a safety assessment program as a screening tool to prioritize fragrance materials for higher-tier testing, as supporting evidence when using a read-across approach, and as evidence to adjust the threshold of toxicological concern. Predictive values for the BlueScreen HC assay were evaluated based on the ability of the assay to predict the outcome of in vitro and in vivo mutagenicity and chromosomal damage genotoxicity assays. A set of 371 fragrance materials was assessed in the BlueScreen HC assay along with existing or newly generated in vitro and in vivo genotoxicity data. Based on a weight-of-evidence approach, the majority of materials in the data set were deemed negative and concluded not to have the potential to be genotoxic, while only a small proportion of materials were determined to show genotoxic effects in these assays. Analysis of the data set showed a combination of high positive agreement but low negative agreement between BlueScreen HC results, in vitro regulatory genotoxicity assays, and higher-tier test results. The BlueScreen HC assay did not generate any false negatives, thereby providing robustness when utilizing it as a high-throughput screening tool to evaluate the large inventory of fragrance materials. From the perspective of protecting public health, it is desirable to have no or minimal false negatives, as a false-negative result may incorrectly indicate the lack of a genotoxicity hazard. However, the assay did have a high percentage of false-positive results, resulting in poor positive predictivity of the in vitro genotoxicity test battery outcome. Overall, the assay generated 100% negative predictivity and 3.9% positive predictivity. In addition to the data set of 371 fragrance materials, 30 natural complex substances were evaluated for BlueScreen HC, Ames, and in vitro micronucleus assay, and a good correlation in all three assays was observed. Overall, while a positive result may have to be further investigated, these findings suggest that the BlueScreen HC assay can be a valuable screening tool to detect the genotoxic potential of fragrance materials and mixtures.

Assessment of the genotoxic potential of mintlactone.

Mintlactone (chemical name 3,6-dimethyl-5,6,7,7a-tetrahydro-1-benzofuran-2(4H)-one, CAS Number 13341-72-5) is a fragrance and flavor ingredient with reported uses in many different cosmetics, personal care, and household products. In order to evaluate the genotoxic potential of mintlactone, in vitro and in vivo genotoxicity tests were conducted. Results from bacterial mutagenicity tests varied across different batches of differing purity with positive results observed in TA98 only. An in vivo comet assay was also considered to be positive in livers of female mice but negative in male mice. In contrast, in vitro and in vivo micronucleus tests, as well as 3D skin comet/micronucleus tests, were negative, indicating no chromosomal or DNA damage. The underlying causes for these contradictory results are not clear. It appears that the purity and/or stability of the test material may be an issue. In the absence of dependable scientific information on the purity and/or storage stability of mintlactone, its safety for use as a fragrance ingredient cannot be substantiated.

FEMA GRAS assessment of natural flavor complexes: Eucalyptus oil and other cyclic ether-containing flavoring ingredients.

In 2015, the Expert Panel of the Flavor and Extract Manufacturers Association (FEMA) initiated a program for the re-evaluation of the safety of over 250 natural flavor complexes (NFCs) used as flavor ingredients. This publication, the sixth in the series, will summarize the re-evaluation of eight NFCs whose constituent profiles are characterized by significant amounts of eucalyptol and/or other cyclic ethers. This re-evaluation was based on a procedure first published in 2005 and subsequently updated in 2018 that evaluates the safety of naturally occurring mixtures for their intended use as flavoring ingredients. The procedure relies on a complete chemical characterization of the NFC intended for commerce and the organization of its chemical constituents into well-defined congeneric groups. The safety of the NFC is evaluated using the well-established and conservative threshold of toxicological concern (TTC) concept in addition to data on absorption, metabolism and toxicology of the constituents of the congeneric groups and the NFC under evaluation. Eight NFCs derived from the Eucalyptus, Melaleuca, Origanum, Laurus, Rosmarinus and Salvia genera were affirmed as generally recognized as safe (GRAS) under their conditions of intended use as flavor ingredients based on an evaluation of each NFC and the constituents and congeneric groups therein.

FEMA GRAS assessment of natural flavor complexes: Origanum oil, thyme oil and related phenol derivative-containing flavoring ingredients.

In 2015, the Expert Panel of the Flavor and Extract Manufacturers Association (FEMA) initiated a re-evaluation of the safety of over 250 natural flavor complexes (NFCs) used as flavor ingredients, mostly consisting of a variety of essential oils and botanical extracts. This publication, seventh in the series, re-evaluates NFCs with constituent profiles dominated by phenolic derivatives including carvacrol, thymol and related compounds using a constituent-based procedure first published in 2005 and updated in 2018. The procedure is based on the chemical characterization of each NFC as intended for commerce and the estimated intake of the constituent congeneric groups. The procedure applies the threshold of toxicological concern (TTC) concept and evaluates relevant data on absorption, metabolism, genotoxic potential and toxicology of the constituent congeneric groups and the NFC under evaluation. Herein, the FEMA Expert Panel affirmed the generally recognized as safe (GRAS) status of seven phenolic derivative-based NFCs, Origanum Oil (Extractive) (FEMA 2828), Savory Summer Oil (FEMA 3013), Savory Summer Oleoresin (FEMA 3014), Savory Winter Oil (FEMA 3016), Savory Winter Oleoresin (FEMA 3017), Thyme Oil (FEMA 3064) and Thyme White Oil (FEMA 3065) under their conditions of intended use as flavor ingredients.

Contribution to the ongoing discussion on fluoride toxicity.

Since the addition of fluoride to drinking water in the 1940s, there have been frequent and sometimes heated discussions regarding its benefits and risks. In a recently published review, we addressed the question if current exposure levels in Europe represent a risk to human health. This review was discussed in an editorial asking why we did not calculate benchmark doses (BMD) of fluoride neurotoxicity for humans. Here, we address the question, why it is problematic to calculate BMDs based on the currently available data. Briefly, the conclusions of the available studies are not homogeneous, reporting negative as well as positive results; moreover, the positive studies lack control of confounding factors such as the influence of well-known neurotoxicants. We also discuss the limitations of several further epidemiological studies that did not meet the inclusion criteria of our review. Finally, it is important to not only focus on epidemiological studies. Rather, risk analysis should consider all available data, including epidemiological, animal, as well as in vitro studies. Despite remaining uncertainties, the totality of evidence does not support the notion that fluoride should be considered a human developmental neurotoxicant at current exposure levels in European countries.

Comparison of points of departure between subchronic and chronic toxicity studies on food additives, food contaminants and natural food constituents.

It was generally accepted as a default assumption that No-Observed-Adverse-Effect Levels (NOAELs) or Lowest-Observed-Adverse-Effect Levels (LOAELs) in long-term toxicity studies are lower than in short-term ones, i.e. the toxic potency increases with prolonged exposure duration. Recent studies on pesticides and industrial chemicals reported that subacute, subchronic or chronic NOAELs/LOAELs are similar when study design factors are appropriately considered. We investigated whether these findings also apply to certain food constituents. After reviewing subchronic and chronic toxicity studies on more than 100 compounds, a total of 32 compounds could be included in the analysis. Geometric mean (GM) values of subchronic vs. chronic NOAEL or LOAEL ratios ranged from 1.0 to 2.0, with a geometric standard deviation from 2.2 to 4.2, which is consistent with data reported in the literature. While for many of the investigated compounds the ratio is around 1 - suggesting that health-based guidance values could appropriately be derived from subchronic toxicity studies - our study also identified some substances with higher ratios leading to a GM of around 2. The EFSA Scientific Committee suggested to apply an uncertainty factor of 2 to extrapolate from subchronic to chronic studies and, as a precautionary approach, we concur with this suggestion.