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.

Risk assessment of nitrites for the Austrian adult population with probabilistic modelling of the dietary exposure.

Nitrites are present in the food chain as naturally occurring species or contaminants. Additionally, sodium and potassium nitrites are authorised food additives. Nitrites exert acute toxicity through methemoglobinemia or cardiovascular effects, chronic toxicity associated with endocrine, reproductive and developmental effects and have been classified as probable gastric carcinogens. Ingestion of food and water are the main sources of human exposure. This study comprises a tiered risk assessment of nitrites for the Austrian adult population, along with the identification of the food categories most contributing to their intake. The dietary exposure, based on Austrian occurrence and consumption data, was modelled with the Monte Carlo simulation method. In an additional scenario, data gaps were addressed with the usage of occurrence data published by the European Food Safety Authority and from the available literature to account for the exposure from all sources. Risk estimates regarding only the exposure to nitrite additives and to contaminated water indicate low level of concern. However, when exposure from all sources is considered, the estimated exposure is elevated and exceeds the Acceptable Daily Intake for high consumers. Mean exposure attributed to the use of nitrites as additives accounts for only a very small proportion of the total intake.

Incorporating Exogenous and Endogenous Exposures into Dietary Risk Assessment of Nitrates and Nitrites in Vegetables: A Probabilistic Integrated Toxicokinetic Modeling Approach.

This study aimed to estimate the dietary risk of nitrates and nitrites in vegetables based on internal dose in a probabilistic manner by integrating exogenous exposure based on measured concentrations in vegetables with endogenous exposure using a toxicokinetic (TK) model. We optimized and validated a previous TK model and incorporated Monte Carlo simulations to account for variability across different age populations for predicting internal dose. High levels of nitrates were detected in leafy vegetables (from 545 ± 274 to 1641 ± 873 mg/kg). Nitrite contents of vegetables were generally low (from 1.26 ± 1.40 to 8.20 ± 14.1 mg/kg). The dietary risk was found to be different based on internal versus external dose, suggesting that it is critical to include endogenous nitrite formation into risk assessment. Nitrate and nitrite exposure from vegetables is unlikely to result in appreciable risks for most populations but may be a potential risk for preschoolers.

Enzyme-Based Ultrasensitive Electrochemical Biosensors for Rapid Assessment of Nitrite Toxicity: Recent Advances and Perspectives.

In the present era of rapid international globalization and industrialization, intensive use of nitrite as a fertilizing agent in agriculture, preservative, dyeing agent, food additive and as corrosion inhibitor in industrial sectors is adversely effecting environment, natural habitats and human health. The issue of toxicity and carcinogenicity due to excessive ingestion of nitrites via the dietary intake has led to an imminent need for its efficient real-time monitoring in situ. Nitrite detection employing electrochemical biosensors has been gaining high credibility in the field of clinical research. Nitrite biosensors have emerged as an outstanding choice for portable point of care testing of nitrite quantification owing to the excellent properties, such as rapidity, miniaturization, ultra-low limits of detection, multiplexing and enhanced detection sensitivity. The article is enclosed with an interesting outlook on latest emerging trends in the development of nitrite biosensors utilizing nanomaterials, such as metal nanoparticles, carbon nanotubes, metal oxide nanoparticles, nanocomposites, polymers and biomaterials. The present review embarks on the highlights relevant to the nitrite quantification in real samples, then proceeds with a meticulous description of the most pertinent electrochemical nitrite biosensors, which have been proposed by adopting diverse materials and strategies of fabrication and finally end with the achievements and future outlook signifying the application of these nanoengineered biosensors for environmental surveillance and human safety.

Mitigation of nitrite toxicity by increased salinity is associated with multiple physiological responses: A case study using an economically important model species, the juvenile obscure puffer (Takifugu obscurus).

Nitrite is a common pollutant in water and is highly toxic to aquatic animals. To reveal the mechanism of salinity in attenuating nitrite toxicity to fish, we measured the physiological responses of juvenile Takifugu obscurus exposed to nitrite concentrations (0, 10, 20, 50, and 100 mg/L) under different salinity levels (0, 10, and 20 ppt) for 96 h. Salinity increased the survival rates of juvenile T. obscurus exposed to nitrite. Changes in key hematological parameters, antioxidant system, malondialdehyde, Na+/K+-ATPase, and HSP70 indicated that nitrite induced considerable damage to juveniles; salinity mitigated the harmful effects. This finding reflects similar changing trends in both antioxidants and their gene expressions among different tissues. We applied an overall index, an integrated biomarker response (IBR), that increased under high-nitrite condition but recovered to the normal levels under salinity treatment. Analysis of the selected detection indices and IBR values showed that the overall mitigating effect of salinity on nitrite toxicity seems to be at sub-cellular level and associated with complicated physiological responses.

Safety assessment of the aqueous extract of the flowers of Nymphaea lotus Linn (Nymphaeaceae): Acute, neuro- and subchronic oral toxicity studies in albinos Wistar rats.

Background Nymphaea lotus Linn (N. lotus) is a medicinal plant widely used in Cameroon popular medicine, to treat neuropsychiatric conditions, male sexual disorders or as food supplement. However, scientific data on the pharmacotoxic profile of this plant are not available. The safety of N. lotus was assessed in acute, neuro- and subchronic toxicity studies by following the OECD guidelines. Effectively, no data have been published until now in regard to its safety on the nervous system. Methods Aqueous extract of N. lotus at doses of 200, 400 and 600 mg/kg body weight (BW) was evaluated for nitrites contents and orally administered to rats daily for 28 days (5 male, 5 female per group). The control group received distilled water (10 mL/kg) and a satellite group was used to observe reversal effects. Neurotoxicity of the plant was determined using open field test for motor coordination, ataxia and gait analysis. Clinical signs and state of livelihood were recorded during the 24 h, then for 28 days of treatments. At the end of 28-day period, animals were anesthetized and decapitated. The whole brain was homogenized for neurobiochemical analysis. Blood samples were collected with or without anticoagulant for hematological examinations and serum analysis. Specimens of liver, kidney, testis, ovaries, and brain were fixed in 10 % formalin and processed for histopathological examinations. Results Our findings indicate dose-dependent elevation of nitrites contents in the flowers aqueous extract of N. lotus. Acute toxicity study revealed no signs of toxicity neither at the dose 2,000 mg/kg nor at 5,000 mg/kg. Thus the LD50 value of aqueous extract of N. lotus flowers is superior to 5,000 mg/kg. The repeated administration of N. lotus during 28 days, induced no signs of neurobehavioral changes in male, but female rats exhibited dose-dependent response in the open field test, suggesting sex and dose-relative psychotropic effects of N. lotus. The evaluation of neurobiochemistry revealed consistent rise of brain cholesterol by 44.05 %; 158.10 % and 147.62 % respectively in male rats treated with the doses of 200, 400 and 600 mg/kg. In female rats, these levels were significantly increased (p<0.001) only at the dose of 600 mg/kg compared to control. This trend persisted after 14 days withdrawal. Brain potassium and calcium concentrations were increased in all rats compared to their respective control receiving distilled water, suggesting transmembrane current stabilizing properties of brain cells by our extract. Further, serum biochemical analysis demonstrated that 28-day administration of N. lotus flowers increased depending on the dose and sex, the levels of serum urea, proteins, creatinine and bilirubin and reduced γ-glutamyltransferase (GGT) and alkaline phosphatase (ALP) activities. These results suggest liver alterations that are endowed by lower liver relative weight and histology damages observed in female rats treated with the dose of 600 mg/kg of our extract. We also observed a rise in the low-density lipoprotein (LDL) fraction and AI of male rats undergoing N. lotus treatment. In female rats, the latter remains unaltered, confirming the dose- and sex-dependent response of our extract. The levels of white blood cells (WBC) and granulocytes were higher in male irrespective to their control, revealing stimulatory properties of the male hematopoietic system. Such variations (sex- and dose-dependent) are without biological relevance for the majority of the biochemical parameters evaluated, indicating a wide margin of safety for the traditional use of N. lotus. The alkaloids, nitrites and phytosterols contained in N. lotus flowers extract may probably account for its neuroprotective, anti-oxidant, and immunoboosting properties. Conclusions N. lotus do not possesses neurotoxicity but is able to induce behavioral changes in rats. Therefore, the application of this plant as either drug or supplementary food should be carefully considered.

[Aquatic ecological risk assessment of microcystins and nitrogen pollution based on species sensitivity distribution].

Species sensitivity distributions (SSDs) methods in both forward and reverse modes were used to evaluate the ecological risk and determine the contaminant concentration threshold for the protection of aquatic species and ecological quality. In this study, the existing toxicity data of freshwater organisms were fitted to SSD functions to estimate the hazardous concentrations for 5% of the species (HC5) for microcystins, ammonia and nitrite, and the ecological risk of their mixtures. The potentially affected fractions (PAFs) of various concentrations of microcystins, ammonia and nitrite were also calculated. Results showed that microcystins exhibited a higher ecological risk than ammonia and nitrite. The HC5 value for microcystins exposure was 19.22 microg x L(-1) whereas the HC5 values for ammonia and nitrite exposure were 6583.94 microg x L(-1) and 334.33 microg x L(-1), respectively. The sensitivity of freshwater organisms varied with exposed concentrations of microcystins, ammonia and nitrite. Crustaceans were more sensitive than fishes to microcystins, and less sensitive than fishes to nitrite when the concentrations of microcystins and nitrite were below 125.04 microg x L(-1) and 2989.40 microg x L(-1), respectively, and vice versa when exposed to higher concentrations of microcystins and nitrite. No significant difference was observed for the sensitivities of fishes and crustaceans exposed to ammonia. In studies with selected lakes in China, our results showed that the ecological risk in both Tai and Hongfeng lakes exceeded the permissible HC5 threshold, and the multiple substance potentially affected fractions (msPAFs) of microcystins, ammonia and nitrite were 2.6%-5.6%, indicating that the ecological risk of their mixtures was more threatening than each individual contaminant being investigated.

Monitoring of nitrites and nitrates levels in leafy vegetables (spinach and lettuce): a contribution to risk assessment.

BACKGROUND: Nitrites and nitrates are compounds considered harmful to humans and the major part of the daily intake of nitrates in foodstuffs is related to vegetable consumption. In this work, 150 leafy vegetables samples (75 spinach and 75 lettuce) were analysed in order to assess the levels of nitrites and nitrates. The analyses were carried out by a validated ion chromatography method and the samples with nitrate concentrations higher than legal limits and/or with quantifiable concentrations of nitrites were confirmed by an alternative ion chromatography method. RESULTS: Nitrate levels higher than legal limits were detected both in spinach (four samples) and in lettuce (five samples). Nitrite residues were registered both at low concentrations--lower than 28.5 mg kg⁻¹ (12 spinach samples)--and at high concentrations, up to 197.5 mg kg⁻¹ (three spinach and one lettuce sample). CONCLUSION: Considering the non-negligible percentage of 'not-compliant' samples for nitrates (6.0%), control is needed. Moreover, it is possible to suggest the introduction in the Communities Regulations of a 'maximum admissible level' for nitrites in leafy vegetables.

Assessment of health hazard associated with nitrogen compounds in water.

This paper presents results of analyses of water in the river Mala Panew in South West Poland flowing through a rural area with some chemical industry developed. The aims of the work were to investigate the pollutants level in the river, compare the obtained results with obligatory drinking water standards and determine possible health effects when using the river as a source for drinking water production. Attention was given to nitrogen compounds as nitrate(V) ions (NO(3-)) and nitrite(III) ions (NO(2-)), mostly of anthropogenic origin, were detected in the monitored water. The average concentrations of NO(3-) and NO(2-) were 3.54 and 0.286 mg/dm(3), respectively. The chances for non-carcinogenic effects, namely methemoglobinemia, resulting from possible exposure to the examined chemicals were determined based on the analytical and toxicological data. As infants are the sub-population most susceptible to nitrate-induced methemoglobinemia, the assessment was limited to children aged 0-3 years. The determined values expressed by hazard quotient (HQ) and hazard index (HI) indicate that the water pollutants and their concentrations do not exceed unity; however, in the case of infants, the other nitrate sources should be controlled.

Ecological risk assessment for aquatic species exposed to contaminants in Keelung River, Taiwan.

An ecological risk assessment was conducted for Keelung River in northern Taiwan. The objective of this study was to assess the risk to fish, aquatic insects, and benthic macroinvertebrates associated with chemical-of-potential-concern (COPC) in the river and to rank ecological risk for these chemicals. The protection of at least 95% of the species 90% of the time from acute and chronic COPC exposures was the defined assessment endpoint. Nine inorganic and organic contaminants were selected to evaluate the impact to aquatic community in the Keelung River. The quotient method served as screen level estimation of risk. The Aquatic Ecological Risk Assessment model was used to analyze exposure and ecological effects and to estimate community level risk. The logarithmic regression model between probability and lethal concentration was established. The combined risks of multiple chemicals were evaluated under assumption of additive risk. The results indicated that zinc and copper pose higher risk among metals. Ammonia, copper, and zinc posed virtually all of the risk, while organic COPCs posed a negligible risk. Potential ecological risk from ammonia exposure was greatest. The probability of more than 5% of the species being affected by acute or chronic toxicity of COPCs is about 100%. In average (50% of the time), 99% of the species would be affected by acute toxicity of COPCs, and about all the species would be affected by chronic toxicity of COPCs. Uncertainties in this assessment were associated with variability in ecosystem stressors, exposure data, ecological effect data, and risk characterization.

Acid-induced structural modifications of unsaturated Fatty acids and phenolic olive oil constituents by nitrite ions: a chemical assessment.

The structural modifications of the unsaturated fatty acid components of triglycerides in extra virgin olive oil (EVOO) following exposure to nitrite ions in acidic media were determined by two-dimensional (2D) NMR spectroscopy, aided by (15)N labeling and GC analysis, allowing investigation of the matrix without fractionation steps. In the presence of excess nitrite ions in a 1% sulfuric acid/oil biphasic system, extensive double bond isomerization of the oleic/linoleic acid components of triglycerides was observed associated with nitration/oxidation processes. Structurally modified species were identified as E/Z-nitroalkene, 1,2-nitrohydroxy, and 3-nitro-1-alkene(1,5-diene) derivatives based on (1)H, (13)C, and (15)N 2D NMR analysis in comparison with model compounds. Minor constituents of EVOO, including phenolic compounds and tocopherols, were also substantially modified by nitrite-derived nitrating species, even under milder reaction conditions relevant to those occurring in the gastric compartments. Novel nitrated derivatives of tyrosol, hydroxytyrosol, and oleuropein (6-8) were identified by LC/MS analysis of the polar fraction of EVOO and by comparison with synthetic samples. Overall, these results provide the first systematic description at the chemical level of the consequences of exposing EVOO to nitrite ions at acidic pH and offer an improved basis for further investigations in the field of toxic nitrosation/nitration reactions and dietary antinitrosating agents.

[Toxicological and hygienic assessment of nitrates in food products]. / Toksiko-gigienicheskaia otsenka nitratov v pishchevykh produktakh.

The toxicological and hygienic estimation of the contents of nitrates in the basic vegetable cultures (beet, carrots, potato etc.) is given. Is shown, that nitrates, contained in the increased amounts in the aforesaid cultures are not indifferent for human and animals organism. At a unbalanced diet of cattle use of forages with the increased contents of nitrates can result in occurrence them in livestock products. The hygienic regulation of permissible concentrations nitrates in some products should be carried out with an allowance fo structure of nutrition of the population, contents nitrates in drinking water, and also other factors capable to influence on formation of nitrate body burden on the population and to aggravate effect of nitrates action.

[Assessment of the degree of carcinogenicity of small doses of nitrites]. / Otsenka stepeni kantserogennoi opasnosti malykh doz nitritov.

Chronic experiments on CBA and C57B1 mice and acute experiments on CBA mice established: (a) carcinogenic effect of sodium nitrite given continuously with drinking water (0.1; 1.0 and 10.0 maximum allowable concentration) in combination with morpholine fed with bread, and (b) endogenous synthesis of nitrosomorpholine as a result of simultaneous intragastric administration of same doses of sodium nitrite and morpholine. Also, nitrosomorpholine and N-nitrosodimethylamine synthesis was observed in vitro following addition of low-dose sodium nitrite, morpholine and amidopyrine to human gastric juice. Carcinogenic hazard associated with low-dose nitrite consumption in humans is discussed.