Safety evaluation of 5-hydroxytryptophan and S-(2-aminoethyl)isothiouronium bromide hydrobromide on rodent lungs.

OBJECTIVES: During the past few decades, various compounds have been researched for their potential as radioprotectants, and many of them were found to be safe and effective in several preclinical models. However, many of these compounds were found to have serious adverse effects when evaluated in clinical settings, thereby making them unsuitable for human applications. 5-hydroxytryptophan (5-HTP) and S-(2-aminoethyl) isothiouronium bromide hydrobromide (AET) act in a synergistic fashion to promote radioprotection. The present study primarily emphasizes the safety of fixed dose of 5-HTP + AET in the lungs of C57BL/6 mice, a well-known model used in drug safety studies. MATERIALS AND METHODS: Post-administration of the combination of HTP+AET at specific time points, blood and bronchoalveolar lavage fluid (BALF) were collected for the analysis of inflammatory and oxidative stress markers of the lungs. Thereafter, the mice were sacrificed and the lungs were dissected out, weighed, and fixed in formalin for histopathological studies. RESULTS: The inflammatory biomarkers: tumor necrosis factor-alpha and interleukin-10 and oxidative stress biomarkers: 8-isoprostane and 8-hydroxy-2'-deoxyguanosine were found to have normal levels in blood and BALF in both control and treatment groups, which was further supported by normal histological findings. In addition, other endpoints such as food and water intake were found to be within normal limits. CONCLUSION: The present safety study reflects that the combination has no adverse effects on the lungs of the experimental mouse. Further, evaluation in higher mammals including nonhuman primates is essential prior to validation of the safety of the combination in humans.

Low concentration of bromide ions improves sulfadiazine phytoremoval and attenuates its phytotoxicity.

Sulfonamide antibiotics (SAs) are ubiquitous in surface water and soil environments, raising considerable concerns about their risk and removal. However, the impacts of various bromide ion (Br-) concentrations on the phytotoxicity, uptake and fate of SAs in plant growth and physiological metabolism of plants have not been well understood. Our research demonstrated that low concentrations of Br- (0.1, 0.5 mM) promoted the uptake and degradation of sulfadiazine (SDZ) in wheat and attenuated the phytotoxicity of SDZ. Additionally, we proposed a degradation pathway and identified the brominated product of SDZ (SDZBr), which attenuated the dihydrofolate synthesis inhibition by SDZ. The primary mechanism was that Br- reduced the level of reactive oxygen radicals (ROS) and alleviated oxidative damage. The production of SDZBr and the high consumption of H2O2 suggest the potential generation of reactive bromine species, contributing to the degradation of the electron-rich SDZ and thus reducing its toxicity. Moreover, metabolome analysis of wheat roots indicated that low concentrations of Br- stimulated the production of indoleacetic acid under SDZ stress, promoting growth and enhancing the uptake and degradation of SDZ. Conversely, high Br- (1 mM) concentration produced a deleterious effect. These findings provide valuable insights into the mechanisms of antibiotic removal, suggesting a potentially novel approach to plant-based antibiotic remediation.

Toxicity comparison of three imidazolium bromide ionic liquids to soil microorganisms.

Ionic liquids (ILs) are extensively used in several chemistry fields. And research about the effects of ILs on soil microbes is needed. In this study, brown soil was exposed to 1-butyl-3-methylimidazolium bromide ([C4mim]Br), 1-hexyl-3-methylimidazolium bromide ([C6mim]Br) and 1-decyl-3-methylimidazolium bromide ([C10mim]Br). The toxicities of the three ILs are evaluated by measuring the soil culturable microbial number, enzyme activity, microbial diversity and, abundance of the ammonia monooxygenase (amoA) genes of ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA). Results showed that all tested ILs caused a decrease in culturable microbial abundance. Tested ILs exposure inhibit urease activity and promote acid phosphatase and ß-glucosidase activities. Tested ILs reduced soil microbial diversity and the abundances of AOB-amoA and AOA-amoA genes significantly. After a comparison of the integrated biomarker response (IBR) index, the toxicities of tested ILs to soil microorganisms were as follows: [C10mim]Br > [C6mim]Br > [C4mim]Br. Among all collected biomarkers, the abundance of the AOA-amoA gene was the most sensitive one and was easily affected after ILs exposure.

Differentiated and exponentially growing HL60 cells exhibit different sensitivity to some genotoxic agents in the comet assay.

The aim of this study was to investigate the effect of the cell differentiation status on the sensitivity to genotoxic insults. For this, we utilized the comet assay to test the DNA damage after treatment with 5 different substances with different mechanism of action in human promyelocytic HL60 cells with or without cell differentiation. A 4-hour MMS treatment induced a significant and concentration-dependent increase in DNA damage for both differentiated and undifferentiated cells, but the difference in sensitivity was only significant at the highest concentration. A 4-hour doxorubicin treatment did not induce DNA damage in differentiated HL60 cells, while it did in undifferentiated cells with its highest tested concentration. A one-hour etoposide treatment caused significant increase in DNA damage concentration dependently in both cell variants. This DNA damage was significantly higher in undifferentiated HL60 cells with several tested concentrations of etoposide. The treatment with the oxidizing substances hydrogen peroxide and potassium bromate yielded significant DNA damage induction in both undifferentiated and differentiated cells with no difference according to the differentiation status. Doxorubicin and etoposide are known to inhibit topoisomerase II. The activity of this enzyme has been shown to be higher in undifferentiated actively proliferating cells than in differentiated cells. This may be of relevance when exposures to topoisomerase-inhibiting compounds or the genotoxicity of compounds with unknown mechanism of action are assessed in routine testing.

Potential impurities in drug substances: Compound-specific toxicology limits for 20 synthetic reagents and by-products, and a class-specific toxicology limit for alkyl bromides.

This paper provides compound-specific toxicology limits for 20 widely used synthetic reagents and common by-products that are potential impurities in drug substances. In addition, a 15 µg/day class-specific limit was developed for monofunctional alkyl bromides, aligning this with the class-specific limit previously defined for monofunctional alkyl chlorides. Both the compound- and class-specific toxicology limits assume a lifetime chronic exposure for the general population (including sensitive subpopulations) by all routes of exposure for pharmaceuticals. Inhalation-specific toxicology limits were also derived for acrolein, formaldehyde, and methyl bromide because of their localized toxicity via that route. Mode of action was an important consideration for a compound-specific toxicology limit. Acceptable intake (AI) calculations for certain mutagenic carcinogens assumed a linear dose-response for tumor induction, and permissible daily exposure (PDE) determination assumed a non-linear dose-response. Several compounds evaluated have been previously incorrectly assumed to be mutagenic, or to be mutagenic carcinogens, but the evidence reported here for such compounds indicates a lack of mutagenicity, and a non-mutagenic mode of action for tumor induction. For non-mutagens with insufficient data to develop a toxicology limit, the ICH Q3A qualification thresholds are recommended. The compound- and class-specific toxicology limits described here may be adjusted for an individual drug substance based on treatment duration, dosing schedule, severity of the disease and therapeutic indication.

The toxicity of ionic liquid 1-decylpyridinium bromide to the algae Scenedesmus obliquus: Growth inhibition, phototoxicity, and oxidative stress.

Although ionic liquids (ILs) are unlikely to act as air contaminants, their high solubility and slow degradation make them a potential threat to the aquatic environment. The IL 1-decylpyridinium bromide ([DPy]Br) is a common type of pyridine IL, which has varied applications such as in extraction, separation, and catalytic synthesis. Herein, the toxicity of [DPy]Br to S. obliquus is determined. Growth was inhibited by high-concentration [DPy]Br, whereas it had a hormetic effect at low concentrations. The IC50-96h was approximately 0.06mg/L. The cell membrane permeability of S. obliquus increased with [DPy]Br concentration, indicating that [DPy]Br can cause damage to the algae cell structure. Chlorophyll content decreased at high [DPy]Br concentration; chlorophyll fluorescence parameters, such as the maximum effective quantum yield of PSII (Fv/Fm), potential activity of PSII (Fv/F0), yield of the photochemical quantum [Y(II)], and the non-photochemical quenching coefficient (NPQ) were affected, suggesting that [DPy]Br can damage PSII. The ROS fluorescent images revealed that the morphology of cells changed gradually from fusiform to round. High ROS levels were observed with high concentrations of [DPy]Br, indicating that [DPy]Br induced oxidative stress on S. obliquus. The SOD and CAT activities increased when the concentration was lower than IC50, whereas they decreased when the concentration was higher than IC50. The relative ROS content was significantly correlated with growth inhibition rate, cell membrane permeability, chlorophyll content, and SOD and CAT activities. The increase of ROS content in algal cells is an important toxicological mechanism of [DPy]Br to S. obliquus.

Effect of 1-methyl-3-hexylimidazolium bromide on zebrafish (Danio rerio).

Room-temperature ionic liquids, generally referred to ionic liquids (ILs), are "green solvents". Antioxidant responses and DNA damage in zebrafish livers exposed to 1-methyl-3-hexylimidazolium bromide ([C6mim]Br) were evaluated at various doses (5-40 mg/L) for a 28-day IL-exposure. A significant decrease of superoxide dismutase (SOD) activity was exhibited, and catalase (CAT) was inhibited at the highest dose (40 mg/L). Reactive oxygen species (ROS) levels were significantly promoted at most exposure interval times except for the dose of 5 mg/L on day 21 in male and days 21 and 28 in female. Malonaldehyde (MDA) contents remarkable increased exposed to [C6mim]Br. Besides, a notable increase was exhibited, which indicated an inducement of DNA damage with respect to control groups. Thus, we believed that [C6mim]Br causes oxidative stress and DNA damage in zebrafish. Gender differences were insignificant in almost all the tested biomarkers, thus, male and female zebrafish could be mixed at a ratio of 1:1 in the future evaluation. The present study may also provide basic toxicology information for IL evaluation to aquatic organisms.

Comparative mammalian cell cytotoxicity of wastewater with elevated bromide and iodide after chlorination, chloramination, or ozonation.

Recycling wastewater is becoming more common as communities around the world try to better control their water resources against an increased frequency of either prolonged droughts or intense flooding. For communities in coastal areas, wastewaters may contain elevated levels of bromide (Br-) and iodide (I-) from seawater intrusion or high mineral content of source waters. Disinfection of such wastewater is mandatory to prevent the spread of pathogens, however little is known about the toxicity of wastewater after disinfection in the presence of Br- and I-. In this study we compared the induction of chronic cytotoxicity in mammalian cells in samples of municipal secondary wastewater effluent amended with elevated levels of Br-/I- after disinfection by chlorine, chloramines or ozone to identify which disinfection process generated wastewater with the lowest level of adverse biological response. Chlorination increased mammalian cell cytotoxicity by 5 times as compared to non-disinfected controls. Chloramination produced disinfected wastewater that expressed 6.3 times more cytotoxicity than the non-disinfected controls and was 1.3 times more cytotoxic than the chlorinated samples. Ozonation produced wastewater with cytotoxicity comparable to the non-disinfected controls and was at least 4 times less cytotoxic than the chlorine disinfected wastewaters. These results indicate that compared to chlorination and chloramination, ozonation of wastewater with high Br-/I- levels yielded the lowest mammalian cell cytotoxicity, suggesting its potential as a more favorable method to disinfect wastewater with minimizing the biological toxicity in mind.

Effect of increasing bromide concentration on toxicity in treated drinking water.

Research is increasingly indicating the potential chronic health effects of brominated disinfection by-products (DBPs). This is likely to increase with elevated bromide concentrations resulting from the impacts of climate change, projected to include extended periods of drought and the sudden onset of water quality changes. This will demand more rigorous monitoring throughout distribution systems and improved water quality management at water treatment plants (WTPs). In this work the impact of increased bromide concentration on formation of DBPs following conventional treatment and chlorination was assessed for two water sources. Bioanalytical tests were utilised to determine cytotoxicity of the water post disinfection. Coagulation was shown to significantly reduce the cytotoxicity of the water, indicating that removal of natural organic matter DBP precursors continues to be an important factor in drinking water treatment. Most toxic species appear to form within the first half hour following disinfectant addition. Increasing bromide concentration across the two waters was shown to increase the formation of trihalomethanes and shifted the haloacetic acid species distribution from chlorinated to those with greater bromine substitution. This correlated with increasing cytotoxicity. This work demonstrates the challenges faced by WTPs and the possible effects increasing levels of bromide in source waters could have on public health.

Influence of bromide on the performance of the amphipod Hyalella azteca in reconstituted waters.

Poor performance of the amphipod Hyalella azteca has been observed in exposures using reconstituted waters. Previous studies have reported success in H. azteca water-only exposures with the addition of relatively high concentrations of bromide. The present study evaluated the influence of lower environmentally representative concentrations of bromide on the response of H. azteca in 42-d water-only exposures. Improved performance of H. azteca was observed in reconstituted waters with >0.02 mg Br/L. Environ Toxicol Chem 2016;35:2425-2429. Published 2016 Wiley Periodicals Inc. on behalf of SETAC. This article is a US Government work and, as such, is in the public domain in the United States of America.

Phytotoxicity of imidazolium-based ILs with different anions in soil on Vicia faba seedlings and the influence of anions on toxicity.

To evaluate the toxic effects of ionic liquids (ILs) in soil on plants at the molecular and cellular levels and to assess the influence of anions on IL toxicity, the toxic effects of 1-decyl-3-methylimidazolium chloride ([Demim]Cl), 1-decyl-3-methylimidazolium bromide ([Demim]Br) and 1-decyl-3-methylimidazolium nitrate ([Demim]NO3) in soil on Vicia faba (V. faba) seedlings were studied for the first time. Our results show that these ILs had little impact on the growth of V. faba seedlings at 1, 5 and 25 mg kg(-1); however, the shoot length, root length, dry weight and pigment contents of the seedlings were significantly affected at 50 mg kg(-1). Furthermore, the EC50 values for effects of [Demim]Cl, [Demim]Br and [Demim]NO3 on the shoot length, root length and dry weight induced were consistent, indicating that the anion may have little influence on IL toxicity. ROS levels were also significantly enhanced at 50 mg kg(-1), resulting in cellular lipid peroxidation, DNA damage and oxidative damage.

Toxic effects of ionic liquid 1-octyl-3-methylimidazolium bromide on the antioxidant defense system of freshwater planarian, Dugesia japonica.

The activities of antioxidant enzymes and the levels of glutathione (GSH) and malondialdehyde (MDA) were determined when freshwater planarian Dugesia japonica was exposed to different concentrations of 1-octyl-3-methylimidazolium bromide ([C8mim]Br) for one, three, and five days. The results showed that superoxide dismutase (SOD) activity began to increase in all treated groups after three days of exposure, while catalase (CAT) activity was inhibited after the first day, but increased notably on the fifth day except for the lowest concentration group. The activity of glutathione peroxidase (GPX) was induced from the first day of exposure and increased significantly after five days in all treated groups. During the experiment, the levels of intracellular GSH in all treated groups were higher than that of the control group. Changes in MDA suggest that [C8mim]Br is toxic to D japonica and may result in lipid peroxidation in planarian. Our results also indicate that GPX as well as GSH seem to be more sensitive biomarkers of oxidative stress compared with SOD and CAT.

Effects of the ionic liquid 1-hexyl-3-methylimidazolium bromide on root gravitropism in Arabidopsis seedlings.

The toxic effects of ionic liquids (ILs) have attracted increasing attention in recent years. However, the knowledge about the toxic effects of ILs on tropism in organisms remains quite limited. In this study, the effects of 1-hexyl-3-methylimidazolium bromide [C6mim]Br on root gravitropism were evaluated using Arabidopsis seedlings. Our results showed that the root growth and gravity response were significantly inhibited with increasing IL concentration. [C6mim]Br treatment affected the amount and distribution pattern of amyloplasts in root cap compared with controls. The auxin distribution marked with DR5rev::VENUS was altered in IL-treated seedlings. The signal intensity and gene expression of auxin efflux carriers PIN2 and PIN3 were obviously decreased by IL stress. Moreover, as consequences in response to gravity stimulus, the asymmetric DR5 signals in control root apex were impaired by IL treatment. The predominant PIN2 signals along the lower flank of root and PIN3 polarization in columella cells were also significantly reduced in seedlings exposed to IL. Our results suggest that the ionic liquid [C6mim]Br affects the amount and distribution of amyloplasts and disturbs the deployment of PIN2 and PIN3, thus impairing auxin flows in response to gravity stimulus and causing deficient root gravitropism in Arabidopsis seedlings.

Cytotoxicity of 1-dodecyl-3-methylimidazolium bromide on HepG2 cells.

We evaluated the cytotoxicity of 1-dodecyl-3-methylimidazo-lium bromide ([C12mim][Br]) on HepG2 cells and its influence on plasma membrane permeability. The results showed that [C12mim][Br] inhibited HepG2 cell growth and decreased cell viability in a concentration-depen-dent manner. The results also revealed that [C12mim][Br] exposure induced apoptosis in [C12mim][Br]-treated HepG2 cells. In addition, the results showed that [C12mim][Br] increased membrane permeability in HepG2 cells. These results suggest that plasma membrane permeability may be responsible for apoptosis induced by [C12mim][Br] in HepG2 cells.

Mechanistic Study on the Formation of Cl-/Br-/I-Trihalomethanes during Chlorination/Chloramination Combined with a Theoretical Cytotoxicity Evaluation.

Chlorination followed by chloramination can be used to mitigate the formation of potentially toxic iodinated disinfection byproducts (I-DBPs) while controlling the formation of regulated chloro-bromo-DBPs (Cl-/Br-DBPs). Water samples containing dissolved organic matter (DOM) isolates were subjected to 3 disinfection scenarios: NH2Cl, prechlorination followed by ammonia addition, and HOCl alone. A theoretical cytotoxicity evaluation was carried out based on the trihalomethanes (THMs) formed. This study demonstrates that the presence of bromide not only enhances the yield and rate of iodate formation, it also increases the formation of brominated I-THM precursors. A shift in the speciation from CHCl2I to the more toxic CHBr2I, as well as increased iodine incorporation in THMs, was observed in the presence of bromide. For low bromide concentrations, a decrease in I-THM formation and theoretical cytotoxicity was achieved only for high prechlorination times, while for high bromide concentrations, a short prechlorination time enabled the full conversion of iodide to iodate. For low DOM concentrations or DOM with low reactivity, Br-/I-THMs were preferentially formed for short prechlorination times, inducing high cytotoxicity. However, for high chlorine exposures, the cytotoxicity induced by the formation of regulated THMs might outweigh the benefit of I-THM mitigation. For high DOM concentrations or DOM with higher reactivity, mixed I-THMs were formed together with high concentrations of regulated THMs. In this case, based on the cytotoxicity of the THMs formed, the use of NH2Cl is recommended.

Toxic impact of bromide and iodide on drinking water disinfected with chlorine or chloramines.

Disinfectants inactivate pathogens in source water; however, they also react with organic matter and bromide/iodide to form disinfection byproducts (DBPs). Although only a few DBP classes have been systematically analyzed for toxicity, iodinated and brominated DBPs tend to be the most toxic. The objectives of this research were (1) to determine if monochloramine (NH2Cl) disinfection generated drinking water with less toxicity than water disinfected with free chlorine (HOCl) and (2) to determine the impact of added bromide and iodide in conjunction with HOCl or NH2Cl disinfection on mammalian cell cytotoxicity and genomic DNA damage induction. Water disinfected with chlorine was less cytotoxic but more genotoxic than water disinfected with chloramine. For both disinfectants, the addition of Br(-) and I(-) increased cytotoxicity and genotoxicity with a greater response observed with NH2Cl disinfection. Both cytotoxicity and genotoxicity were highly correlated with TOBr and TOI. However, toxicity was weakly and inversely correlated with TOCl. Thus, the forcing agents for cytotoxicity and genotoxicity were the generation of brominated and iodinated DBPs rather than the formation of chlorinated DBPs. Disinfection practices need careful consideration especially when using source waters containing elevated bromide and iodide.

Response to copper bromide exposure in Vicia sativa L. seeds: analysis of genotoxicity, nucleolar activity and mineral profile.

Copper bromide (CuBr2) effects on seed germination and plantlet development of Vicia sativa L. are evaluated through mitotic index, chromosome aberrations, nucleolar activity and mineral profile. CuBr2 induces a significant presence of micronuclei, sticky and c-metaphases, anaphase bridges and chromosome breaks. Increased number of nucleoli and scattering of AgNOR proteins from the nucleolus in the nuclear surface at CuBr2 1mM and in the cytoplasm at CuBr2 5mM, goes along with the decrease of root growth. In V. sativa embryo the content of many macro and micronutrients increases up to copper 1mM in agreement with reserve mobilization while at CuBr2 5mM some elements are present in lower amount. We hypothesize that inhibitory effects observed at 5mM are due either to a nutrient shortage or to a direct influence of copper on root cell division, evidenced by low mitotic index, high occurrence of chromosome aberrations and loss of material from the nucleolus.

Oxidative stress and genotoxicity of the ionic liquid 1-octyl-3-methylimidazolium bromide in zebrafish (Danio rerio).

Ionic liquids (ILs) have a great reputation due to their negligible volatility, designability, good stability, and ability to be recycled. They are considered to be "green" solvents and have great promise in many fields. In recent years, the toxicities of ILs have garnered increasing attention as reported by a number of studies. However, previous studies have primarily focused on their lethal toxicities, and data were limited on their toxic effects at nonlethal doses. We performed a study on the toxic effects of 1-octyl-3-methylimidazolium bromide ([Omim]Br) on zebrafish. During a 28-day period, male and female zebrafish were separately exposed to sequential concentrations (0, 5, 10, 20, and 40 mg/L) of [Omim]Br. Fishes were sampled after 7, 14, 21, and 28 days of exposure, and reactive oxygen species (ROS) levels, activities of antioxidant enzymes (superoxide dismutase and catalase), lipid peroxidation (LPO), and DNA damage in fish livers were measured. ROS, LPO, and DNA damage were all induced by the ionic liquid, and antioxidant enzyme activities increased at the beginning and then decreased. These phenomena demonstrate that [Omim]Br can induce oxidative stress and DNA damage in zebrafish.

The role of the anion in the toxicity of imidazolium ionic liquids.

From the environmental protection point of view, the growing interest of ionic liquids in various industrial branches has raised concerns for the toxicity assessment of these compounds. The paper discusses the effect of salts containing the shared 1-ethyl-3-methylimidazolium [EMIM] cation as coupled with five different anions: bromide [Br], nitrate [NO3], p-toluenesulfonate (tosylate) [Ts], dimethylphosphate [dMP] and methanesulfonate [MS] on the growth and development of higher land plants - spring barley and common radish. The experiment was done according to the ISO Standard 11269-2:1995 and the OECD/OCDE Guide 208/2006. As the indications of phytotoxicity, the percentage of sprouts and the level of dry and fresh plant mass were used; in addition, the visual assessment of any signs of damage to the examined plant species, such as growth inhibition and chlorotic changes, was also made. Results of our study has proved the negative impact of ILs on the tested plants and the toxic effect of imidazolium salts was dependent primarily on the applied ionic liquids concentration. The common radish revealed the higher tolerance to the imidazolium as compared to spring barley. The anion type of ionic liquid was crucial for the toxicity against common radish.

Toxicity of ionic liquids prepared from biomaterials.

In search of environmentally-friendly ionic liquids (ILs), 14 were prepared based on the imidazolium, pyridinium and choline cations, with bromide and several amino acids as anions. Good yields were obtained in the synthesis of pyridinium ILs and those prepared from choline and amino acids. Four of the ILs synthesized from choline and the amino acids arginine, glutamine, glutamic acid and cystine are described here for the first time. The toxicity of the synthesized ILs was checked against organisms of various levels of organization: the crustacean Artemia salina; Human cell HeLa (cervical carcinoma); and bacteria with different types of cell wall, Bacillus subtilis and Escherichia coli. The toxicity was observed to depend on both the cation and anion. Choline-amino acid ILs showed a remarkable low toxicity to A. salina and HeLa cell culture, ten times less than imidazolium and pyridinium ILs. None of ionic liquids exhibited marked toxicity to bacteria, and the effect was 2-3 orders of magnitude smaller than that of the antibiotic chloramphenicol.