Mutagenicity In Ames Test Research Articles

Biophysical characterization of the complexation of C60 fullerene with doxorubicin in a prokaryotic model

By means of small‐angle neutron scattering and ames mutagenicity test it was shown that C60 fullerene may act as an interceptor of the antibiotic doxorubicin by formation of a hetero‐complex with this drug. This result has been predicted based on previously developed model of chemico‐biological interaction between DNA‐binding drugs and aromatic interceptor molecules.

Mutagenicity Assessment of Drinking Water in Combination with Flavored Black Tea Bags: a Cross Sectional Study in Tehran.

Diseases related to water impurities may present as major public health burdens. The present study aimed to assess the mutagenicity of drinking water from different zones of Tehran, and evaluate possible health risks through making tea with tea bags, by Ames mutagenicity test using TA 100, TA 98 and YG1029 strains. For this purpose, 450 water samples were collected over the period of July to December 2014 from 5 different zones of Tehran. Except for one sample, no mutagenic potential was detected during these two seasons and the MI scores were almost normal (≤ 1-1.6) in TA 100, TA 98 and YG1029 strains. Although no mutagenic effects were considered in TA 98 and TA 100 in the test samples of our three evaluated tea bag brands, one sample from a local company showed mutagenic effects in the YG1029 strain (MI=1.7-1.9 and 2) after prolonged (10-15 min.) steeping. Despite the mild mutagenic effect discovered for one of the brand, this cross sectional study showed relative safety of water samples and black tea bags in Tehran. According to the sensitivity of YG1029 to the mutagenic potential of water and black tea, even without metabolic activation by s9 fraction, this metabolizer strain could be considered as sensitive and applicable to food samples for quantitative analysis of mutagens.

Preliminary safety assessment of C-8 xylitol monoester and xylitol phosphate esters.

Most of the cosmetic compounds with preservative properties available in the market pose some risks concerning safety, such as the possibility of causing sensitization. Due to the fact that there are few options, the proper development of new molecules with this purpose is needed. Xylitol is a natural sugar, and the antimicrobial properties of xylitol-derived compounds have already been described in the literature. C-8 xylitol monoester and xylitol phosphate esters may be useful for the development of skincare products. As an initial screen for safety of chemicals, the combination of in silico methods and in vitro testing can aid in prioritizing resources in toxicological investigations while reducing the ethical and monetary costs that are related to animal and human testing. This study was designed to evaluate the safety of C-8 xylitol monoester and xylitol phosphate esters regarding carcinogenicity, mutagenicity, skin and eye irritation/corrosion and sensitization through alternative methods. For the initial safety assessment, quantitative structure-activity relationship methodology was used. The prediction of the parameters carcinogenicity/mutagenicity, skin and eye irritation/corrosion and sensitization was generated from the chemical structure. The analysis also comprised physical-chemical properties, Cramer rules, threshold of toxicological concern and Michael reaction. In silico results of candidate molecules were compared to 19 compounds with preservative properties that are available in the market. Additionally, in vitro tests (Ames test for mutagenicity, cytotoxicity and phototoxicity tests and hen's egg test--chorioallantoic membrane for irritation) were performed to complement the evaluation. In silico evaluation of both molecules presented no structural alerts related to eye and skin irritation, corrosion and sensitization, but some alerts for micronucleus and carcinogenicity were detected. However, by comparison, C-8 xylitol monoester, xylitol phosphate esters showed similar or better results than the compounds available in the market. Concerning experimental data, phototoxicity and mutagenicity results were negative. As expected for compounds with preservative activity, xylitol-derived substances presented positive result in cytotoxicity test. In hen's egg test, both molecules were irritants. Our results suggested that xylitol-derived compounds appear to be suitable candidates for preservative systems in cosmetics.

Tracing nitrogenous disinfection byproducts after medium pressure UV water treatment by stable isotope labeling and high resolution mass spectrometry.

Advanced oxidation processes are important barriers for organic micropollutants (e.g., pharmaceuticals, pesticides) in (drinking) water treatment. Studies indicate that medium pressure (MP) UV/H2O2 treatment leads to a positive response in Ames mutagenicity tests, which is then removed after granulated activated carbon (GAC) filtration. The formed potentially mutagenic substances were hitherto not identified and may result from the reaction of photolysis products of nitrate with (photolysis products of) natural organic material (NOM). In this study we present an innovative approach to trace the formation of disinfection byproducts (DBPs) of MP UV water treatment, based on stable isotope labeled nitrate combined with high resolution mass spectrometry. It was shown that after MP UV treatment of artificial water containing NOM and nitrate, multiple nitrogen containing substances were formed. In total 84 N-DBPs were detected at individual concentrations between 1 to 135 ng/L bentazon-d6 equivalents, with a summed concentration of 1.2 μg/L bentazon-d6 equivalents. The chemical structures of three byproducts were confirmed. Screening for the 84 N-DBPs in water samples from a full-scale drinking water treatment plant based on MP UV/H2O2 treatment showed that 22 of the N-DBPs found in artificial water were also detected in real water samples.

The JaCVAM international validation study on the in vivo comet assay: Selection of test chemicals

The Japanese Center for the Validation of Alternative Methods (JaCVAM) sponsored an international prevalidation and validation study of the in vivo rat alkaline pH comet assay. The main objective of the study was to assess the sensitivity and specificity of the assay for correctly identifying genotoxic carcinogens, as compared with the traditional rat liver unscheduled DNA synthesis assay. Based on existing carcinogenicity and genotoxicity data and chemical class information, 90 chemicals were identified as primary candidates for use in the validation study. From these 90 chemicals, 46 secondary candidates and then 40 final chemicals were selected based on a sufficiency of carcinogenic and genotoxic data, differences in chemical class or genotoxic or carcinogenic mode of action (MOA), availability, price, and ease of handling. These 40 chemicals included 19 genotoxic carcinogens, 6 genotoxic non-carcinogens, 7 non-genotoxic carcinogens and 8 non-genotoxic non-carcinogens. “Genotoxicity” was defined as positive in the Ames mutagenicity test or in one of the standard in vivo genotoxicity tests (primarily the erythrocyte micronucleus assay). These chemicals covered various chemicals classes, MOAs, and genotoxicity profiles and were considered to be suitable for the purpose of the validation study. General principles of chemical selection for validation studies are discussed.

Antimicrobial activity, toxicity and selectivity index of two biflavonoids and a flavone isolated from Podocarpus henkelii (Podocarpaceae) leaves.

BackgroundDifferent parts of Podocarpus henkelii have been used in many cultures around the world to treat ailments such as cholera, stomach diseases, rheumatism, cancer, canine distemper in dogs and gall sickness in cattle. The aim of this study was to evaluate the biological activity and toxicity of isolated compounds from Podocarpus henkelii after an earlier study indicated a promising activity in crude extracts against viral pathogens of veterinary importance.MethodsThe antibacterial and antifungal activity of two biflavonoids 7, 4’, 7”, 4”’-tetramethoxy amentoflavone (TMA), isoginkgetin (IGG) and podocarpus flavone–A (PFA) isolated from the leaves of Podocarpus henkelii were determined using a serial microplate dilution method with tetrazolium violet as growth indicator. The cytotoxicity of compounds TMA and IGG were determined on different cell types using a tetrazolium-based colorimetric cellular assay (MTT). The Ames test was used to determine their mutagenic activities.ResultsTMA had reasonable antifungal activity against Aspergillus fumigatus (MIC = 30 μg/ml). IGG had a wide spectrum of activity against four bacterial and two fungal pathogens with much higher selectivity index values obtained for A. fumigatus and Cryptococcus neoformans (SI > 30). PFA had a broad spectrum of activity against Enterococcus faecalis and Pseudomonas aeruginosa (SI > 15) and less activity against the two fungal pathogens. In both the cytotoxicity assays and Ames mutagenicity test using Salmonella typhimurium strains TA98 and TA100, TMA and IGG had no deleterious effect on the different cell types and did not induce mutations in the Ames test.ConclusionAlthough the antimicrobial activities of the isolated compounds were not that exciting, the compounds had no cytotoxic activity at the highest concentration (1000 μg/ml) tested against all three cell lines. IGG was the most active against E. coli, S. aureus, A. fumigatus and C. neoformans, exhibiting both antibacterial and antifungal activity with good selectivity index values. PFA had a broad spectrum of activity against E. faecalis and P. aeruginosa. The two compounds isolated had low toxicity and no genotoxic activity in the Ames test.

Antioxidant, antimutagenic, and In vivo buccal mucosa cancer preventive effects of Fructus Malvae

Fructus Malvae is functional food known for antioxidant, anti-mutagenic, and in vivo anticancer effects. Fructus Malvae extracts demonstrated high antioxidant activities in DPPH and hydroxyl radical scavenging activity assays. In an Ames mutagenicity test, Fructus Malvae exhibited antimutagenicty in association with MNNG (Nmethyl-N’-nitro-N-nitrosoguanidine) in Salmonella Typhimurium TA100 cells. U14 squamous carcinoma cells were injected into the buccal mucosa of KM (Kunming) mice. The wound at the injection site was smeared with a Fructus Malvae solution, which was also administered to mice by gavage. Tumor volumes were reduced and tissue section analysis of buccal mucosa cancer and cervical lymph node tissues showed anti-cancer effects in Fructus Malvae treated groups. Fructus Malvae possesses good antioxidant and antimutagenic activities and exerts a preventive effect against buccal mucosa cancer in vivo.

Mutagenicity of Edible Palm Oil on the Ghanaian Market before and after Repeated Heating

Red palm oil produced in Ghana largely by village folks has never been tested for its mutagenic potential. The study aimed at determining the mutagenicity of high-energy heated red palm oil (RRPO) and refined, bleached imported palm oil (PO) on the Ghanaian market. Samples of RRPO and PO were 1× and 5× heated for 10 min at 180 °C with a cooling period of 5 h in-between. Unheated, together with heated samples, were tested for mutagenicity using Salmonella typhimurium TA 98 and TA 100 tester stains. Unheated PO was negative for the Ames mutagenicity test with TA 98 strain. However, 1× and 5× heated PO were mutagenic (P = 0.05, each). Testing PO, using TA 100 strain was negative. RRPO was mutagenic with TA 98 strain for heated oils (P = 0.05, each). Assays with TA 100 strain showed highly significant mutations (P = 0.001, each) that increased with increasing heating frequency. PO 1× and 5× heated samples caused significant frameshift mutation in the S. typhimurium TA 98 strain. RRPO caused highly significant point and frameshift mutations in heated samples. Furthermore, unheated RRPO mutagenic potential has serious health implications.

Experimental Testing of Quantum Mechanical Predictions of Mutagenicity: Aminopyrazoles

A computational method for predicting the likelihood of aromatic amines being active in the Ames test for mutagenicity was trialed on a set of aminopyrazoles. A virtual array of compounds was generated from the available sets of hydrazines and α-cyanoaldehydes (or ketones) and quantum mechanical calculations used to compute a probability of being active in the Ames test. The compounds selected for synthesis and testing were not based on the predictions and so spanned the range of predicted probabilities. The subsequently generated results of the Ames test were in good correspondence with the predictions and confirm this approach as a useful means of predicting likely mutagenic risk.

Nanosilica coating for bonding improvements to zirconia

Resin bonding to zirconia cannot be established from standard methods that are currently utilized in conventional silica-based dental ceramics. The solution–gelatin (sol–gel) process is a well developed silica-coating technique used to modify the surface of nonsilica-based ceramics. Here, we use this technique to improve resin bonding to zirconia, which we compared to zirconia surfaces treated with alumina sandblasting and tribochemical silica coating. We used the shear bond strength test to examine the effect of the various coatings on the short-term resin bonding of zirconia. Furthermore, we employed field emission scanning electron microscopy, energy-dispersive X-ray spectroscopy, atomic force microscopy, and Fourier transform infrared spectroscopy to characterize the zirconia surfaces. Water–mist spraying was used to evaluate the durability of the coatings. To evaluate the biological safety of the experimental sol–gel silica coating, we conducted an in vitro Salmonella typhimurium reverse mutation assay (Ames mutagenicity test), cytotoxicity tests, and in vivo oral mucous membrane irritation tests. When compared to the conventional tribochemical silica coating, the experimental sol–gel silica coating provided the same shear bond strength, higher silicon contents, and better durability. Moreover, we observed no apparent mutagenicity, cytotoxicity, or irritation in this study. Therefore, the sol–gel technique represents a promising method for producing silica coatings on zirconia.

Integrated in silico approaches for the prediction of Ames test mutagenicity

The bacterial reverse mutation assay (Ames test) is a biological assay used to assess the mutagenic potential of chemical compounds. In this paper approaches for the development of an in silico mutagenicity screening tool are described. Three individual in silico models, which cover both structure activity relationship methods (SARs) and quantitative structure activity relationship methods (QSARs), were built using three different modelling techniques: (1) an in-house alert model: which uses SAR approach where alerts are generated based on experts judgements; (2) a kNN approach (k-Nearest Neighbours), which is a QSAR model where a prediction is given based on outcomes of its k chemical neighbours; (3) a naive Bayesian model (NB), which is another QSAR model, where a prediction is derived using a Bayesian formula through preselected identified informative chemical features (e.g., physico-chemical, structural descriptors). These in silico models, were compared against two well-known alert models (DEREK and ToxTree) and also against three different consensus approaches (Categorical Bayesian Integration Approach (CBI), Partial Least Squares Discriminate Analysis (PLS-DA) and simple majority vote approach). By applying these integration methods on the validation sets it was shown that both integration models (PLS-DA and CBI) achieved better performance than any of the individual models or consensus obtained by simple majority rule. In conclusion, the recommendation of this paper is that when obtaining consensus predictions for Ames mutagenicity, approaches like PLS-DA or CBI should be the first choice for the integration as compared to a simple majority vote approach.

Computational toxicology using OpenTox & Bioclipse

Computational methods, e.g., OECD QSAR Toolbox and ToxPredict, are increasingly used to support chemicals toxicity assessment in academic settings, industry and governments. The in silico-based assessments complement experimental approaches and have potential to fill the knowledge gaps needed to broadly assess chemical hazards. We present here the interoperable Bioclipse–OpenTox platform as a novel alternative made freely and openly available.The interactive Bioclipse software is combined with remote computational toxicity prediction provided by services in the OpenTox network from various European institutes and SMEs. These online services apply machine learning methods for integration of a number of end points, e.g., Ames mutagenicity test in salmonella, Caco-2 cell model permeability and micronucleus assay in rodents. Coupling of such data to chemical structure assessments lead to prediction of site(s) for metabolism (using SMARTCyp), biodegradation (START), and toxicity mode prediction (Verhaar scheme). The OpenTox platform thus unifies how the services are accessed whereas the Bioclipse software provides the easy-to-use interface for interactively studying the toxic part of molecules. Additional predictive methods that are made accessible via the OpenTox network are automatically discovered by Bioclipse and models can be improved over time without any need to reinstall Bioclipse itself.

An illusion of hormesis in the Ames test: Statistical significance is not equivalent to biological significance

A recent report (Calabrese et al., Mutat. Res. 726 (2011) 91–97) concluded that an analysis of Ames test mutagenicity data provides evidence of hormesis in mutagenicity dose–response relationships. An examination of the data used in this study and the conclusions regarding hormesis reveal a number of concerns regarding the analyses and possible misinterpretations of the Salmonella data. The claim of hormesis is based on test data from the National Toxicology Program using Salmonella strain TA100. Approximately half of the chemicals regarded as hormetic, and the majority of the specific dose–responses identified as hormetic, were actually nonmutagenic. We conclude that the data provide no evidence of hormetic effects. The Ames test is an excellent measure of bacterial mutagenicity, but the numbers of revertant (mutant) colonies on the plate are the result of a complex interaction between mutagenicity and toxicity, which renders the test inappropriate for demonstrating hormesis in bacterial mutagenicity experiments.

Effect of Temperature, Fuming Time, and Distance from Source on the Polycyclic Aromatic Compound (PAC) Content of Bitumen Fumes: Assessment Using a Laboratory Fuming Protocol in Combination with a Sensitive Method for Determining Relative Aggregate Levels of PACs in Complex Mixtures

A laboratory fuming procedure was used in combination with a previously reported variation of the Ames Test (the Nitration Assay) to investigate the relationship between temperature, time at temperature, and distance from source, on the relative PAC content of fumes generated from paving bitumens. The test method, whose endpoint is mutagenic potency (Nitration Mutagenicity Index, NMI), takes advantage of large amplifications of Ames Test mutagenicity resulting from nitro-derivatization of PACs. NMIs were similar and quite low for fumes generated at 110°C, but increased exponentially up to 160°C. NMIs increased linearly with fuming time up to 40 min, after which they remained constant. With some initial fluctuation, they also remained constant with increasing distance of the condenser from the bitumen surface up to 100 cm. NMI increased with temperature at a faster rate than fume weight suggesting that the increases in NMI were related to both quantitative and qualitative changes in the PAC content of the fumes.

Comparative Evaluation of in Silico Systems for Ames Test Mutagenicity Prediction: Scope and Limitations

The predictive power of four commonly used in silico tools for mutagenicity prediction (DEREK, Toxtree, MC4PC, and Leadscope MA) was evaluated in a comparative manner using a large, high-quality data set, comprising both public and proprietary data (F. Hoffmann-La Roche) from 9,681 compounds tested in the Ames assay. Satisfactory performance statistics were observed on public data (accuracy, 66.4-75.4%; sensitivity, 65.2-85.2%; specificity, 53.1-82.9%), whereas a significant deterioration of sensitivity was observed in the Roche data (accuracy, 73.1-85.5%; sensitivity, 17.4-43.4%; specificity, 77.5-93.9%). As a general tendency, expert systems showed higher sensitivity and lower specificity when compared to QSAR-based tools, which displayed the opposite behavior. Possible reasons for the performance differences between the public and Roche data, relating to the experimentally inactive to active compound ratio and the different coverage of chemical space, are thoroughly discussed. Examples of peculiar chemical classes enriched in false negative or false positive predictions are given, and the results of the combined use of the prediction systems are described.

Application of cyclodextrins in environmental bioassays for soil

Based on our former experience on contaminant solubilisation and mobilisation in the process of soil remediation we used cyclodextrins as additives in environmental bioassays, for improving solubility and bioavailability of the contaminant in soil and as a consequence sensitivity of the bioassay. In this article we introduce the findings on the application of RAMEB (randomly methylated β-cyclodextrin) for testing PCP (pentachlorophenol) in soil, in three bioassays: bacterial luminescence-inhibition test with Vibrio fischeri, protozoon growth inhibition test with Tetrahymena pyriformis, and Ames mutagenicity test. We applied RAMEB which has a high solubilising capacity on many typical soil contaminant and PCP, because contradictory results were published for its toxicity and mutagenicity. The RAMEB-aided Ames test, gave a sudden and expressed increase in the mutagenicity of PCP, however, Ames mutagenicity was negative without cyclodextrin (CD). Based on these results we tried to apply RAMEB for increasing sensitivity of other bioassays, such as acute toxicity tests with different test organisms. According to our results the effect of RAMEB on bioavailability and toxic effect depends not only on the K ow value (octanol–water partition coefficient) of the chemical substances, but also on the test organism, the water-content of the test-matrix and the applied concentration of RAMEB, as well as its ratio to PCP. We collected all the characteristics of the bioassays applied for PCP and some other contaminants and showed the measured effect data in comparison with each other. We found that in the complex system of soil and soil suspension, used in the bioassays, the interactions between soil solid, water and gaseous phases, as well as between the test organism and RAMEB result in K ow dependent partition of the contaminant between solid and water phases of soil, RAMEB, and the test organism. The conclusion is that RAMEB undoubtedly has an influence on the fate and behaviour of the contaminant in soil and soil suspensions, and the direction of the RAMEB-induced changes depends on the effective concentration of the RAMEB in the bioassay, the time of contact, the type of test organism, and the characteristics of the RAMEB–contaminant complex. In those cases, when RAMEB increased the effect of a contaminated environmental sample, this CD-induced increase can be considered as a “realistic worse case” situation, which can be very useful in risk assessment, resulting in a moderate overestimate in the value of environmental risk.

In silico predictions of Ames test mutagenicity: An integrated approach

In silico predictions of Ames test mutagenicity: An integrated approach

Applicability domain for in silico models to achieve accuracy of experimental measurements

AbstractThe accuracy of in silico models can be inhomogeneous: models can show excellent performance on some chemical subspaces but have low accuracy on others. We show that applicability domain (AD) approaches can differentiate reliable and non‐reliable predictions and identify those with experimental accuracy for both regression and classification models. For reliably predicted molecules, the predicted values can be used instead of experimental measurements. This can halve time and costs of experimental measurements. The developed classification models for AMES mutagenicity test and CYP450 inhibition, which are important drug discovery properties, are publicly available at the online chemical database and modeling environment (OCHEM) site http://qspr.eu Copyright © 2010 John Wiley & Sons, Ltd.

QSAR models to predict mutagenicity of acrylates, methacrylates and [formula omitted], [formula omitted]-unsaturated carbonyl compounds

Objective The purpose of this study is to develop a quantitative structure–activity relationship (QSAR) model that can distinguish mutagenic from non-mutagenic species with α , β -unsaturated carbonyl moiety using two endpoints for this activity – Ames test and mammalian cell gene mutation test – and also to gather information about the molecular features that most contribute to eliminate the mutagenic effects of these chemicals. Methods Two data sets were used for modeling the two mutagenicity endpoints: (1) Ames test and (2) mammalian cells mutagenesis. The first one comprised 220 molecules, while the second one 48 substances, ranging from acrylates, methacrylates to α , β -unsaturated carbonyl compounds. The QSAR models were developed by applying linear discriminant analysis (LDA) along with different sets of descriptors computed using the DRAGON software. Results For both endpoints, there was a concordance of 89% in the prediction and 97% confidentiality by combining the three models for the Ames test mutagenicity. We have also identified several structural alerts to assist the design of new monomers. Significance These individual models and especially their combination are attractive from the point of view of molecular modeling and could be used for the prediction and design of new monomers that do not pose a human health risk.

Mutagenicity of Chinese traditional medicine Semen Armeniacae amarum by two modified Ames tests

BackgroundSemen armeniacae amarum (SAA) is a Chinese traditional medicine and has long been used to control acute lower respiratory tract infection and asthma, as a result of its expectorant and antiasthmatic activities. However, its mutagenicity in vitro and in vivo has not yet been reported. The Ames test for mutagenicity is used worldwide. The histidine contained in biological samples can induce histidine-deficient cells to replicate, which results in more his+ colonies than in negative control cells, therefore false-positive results may be obtained. So, it becomes a prerequisite to exclude the effects of any residual histidine from samples when they are assayed for their mutagenicity. Chinese traditional herbs, such as SAA, are histidine-containing biological sample, need modified Ames tests to assay their in vitro mutagenicity.MethodsThe mutagenicity of SAA was evaluated by the standard and two modified Ames tests. The first modification used the plate incorporation test same as standard Ames teat, but with new negative control systems, in which different amounts of histidine corresponding to different concentrations of SAA was incorporated. When the number of his+ revertants in SAA experiments was compared with that in new negative control, the effect of histidine contained in SAA could be eliminated. The second modification used a liquid suspension test similar to the standard Ames test, except with histidine-rich instead of histidine-limited medium. The aim of this change was to conceal the effect of histidine contained in SAA on the final counting of his+ revertants, and therefore to exclude false-positive results of SAA in the Ames test. Furthermore, the effect of SAA on chromosomal aberration in mammalian bone marrow cells was tested.ResultsThe standard Ames test showed a positive result for mutagenicity of SAA. In contrast, a negative response was obtained with the modified plate incorporation and modified suspension Ames tests. Moreover, no apparent chromosomal aberrations were observed in mammalian bone marrow cells treated with SAA.ConclusionThe standard Ames test was not suitable for evaluating the mutagenicity of SAA, because false-positive result could be resulted by the histidine content in SAA. However, the two modified Ames tests were suitable, because the experimental results proved that the effect of histidine in SAA and therefore the false-positive result were effectively excluded in these two modified Ames tests. This conclusion needs more experimental data to support in the future. Moreover, the experimental results illustrated that SAA had no mutagenicity in vitro and in vivo. This was in agreement with the clinical safety of SAA long-term used in China.