Ames Test Research Articles

Building a risk matrix for the safety assessment of wood derived biochars

Biochar is recognized as an efficient amendment and soil improver. However, environmental and quality assessments are needed to ensure the sustainability of its use in agriculture. This work considers the biochar's chemical-physical characterization and its potential phyto- and geno-toxicity, assessed with germination and Ames tests, obtaining valuable information for a safe field application. Three biochar types, obtained from gasification at different temperatures of green biomasses from the Tuscan-Emilian Apennines (in Italy), were compared through a broad chemical, physical and biological evaluation. The results obtained showed the relevance of temperature in determining the chemical and morphological properties of biochar, which was shown with several analytical techniques such as the elemental composition, water holding capacity, ash content, but also with FTIR and X-ray spectroscopies. These techniques showed the presence of different relevant surface aliphatic and aromatic groups. The procedures for evaluating the potential toxicity using seeds germination and Ames genotoxicity assay highlights that biochar does not cause detrimental effects when it enters in contact with soil, micro- and macro-organisms, and plants. The genotoxicity test provided a new highlight in evaluating biochar environmental safety.

Comparison of PAC and MOAH for understanding the carcinogenic and developmental toxicity potential of mineral oils

The carcinogenicity and developmental toxicity of unrefined mineral oil is related to its 3–7 ring polycyclic aromatic compounds (PAC) content. Therefore, refining operations focus on the targeted removal PAC from mineral oil that may contain aromatics of low toxicological concern. There are thus, two types of aromatic substances in mineral oil: hazardous and non-hazardous. The first type consists of 3–7 ring PAC which may be naked (unsubstituted) or lowly alkylated. The second type or non-hazardous consists of 1–7 ring aromatics with high degree of alkylation or lack of bay or fjord regions. Although these are toxicologically different, they may both elute in the same fraction when using chromatography. To understand how these two aromatic types are related we have assessed the entire mineral oil refinement process by measuring total mineral oil aromatic hydrocarbons (MOAH) content by chromatography next to regulatory hazard tests which focus on 3–7 ring PAC. MOAH content is positively correlated to its molecular weight resulting in aromatic content bias for high viscosity substances. Hazard to 3–7 ring PAC is best controlled by the validated IP346 or modified Ames test. We explain the concept of high vs low alkylation by shortly reviewing new data on alkylated PAC.

Hydromorphone impurity 2,2-bishydromorphone does not exert mutagenic and clastogenic properties via in silico QSAR prediction and in vitro Ames and micronucleus test

The opioid agonist hydromorphone is indicated for the management of severe acute and chronic pain given that alternate treatments are insufficient. While the genotoxicity profile of hydromorphone is well investigated, little is known about the genotoxic potential of its impurities. In this study, 2,2-bishydromorphone was tested in silico and in vitro for both its mutagenic potential in an Ames test performed with Salmonella typhimurium and Escherichia coli tester strains up to a maximum concentration of 5 mg per plate in the absence and presence of metabolic activation. Furthermore, it was tested for its ability to induce micronuclei in TK6 cells in a micronucleus test up to a maximum concentration of 500 µg/mL with or without an exogenous metabolic activation system. 2,2-Bishydromorphone did not reveal any potential for inducing mutagenicity or clastogenicity under the conditions of the respective tests and is therefore considered non-mutagenic and non-clastogenic/aneugenic in vitro. These results are in line with negative in silico quantitative structure-activity relationship (QSAR) prediction for 2,2-bishydromorphone mutagenicity and clastogenicity and provide evidence of good correlation of in silico and in vitro data. Conclusively, these studies add important new clinically relevant information on the safety of hydromorphone as the impurity of 2,2-bishydromorphone is proven to be non-mutagenic and non-clastogenic.

Effects of Particle Size and Surface Charge on Mutagenicity and Chicken Embryonic Toxicity of New Silver Nanoclusters.

Though there are many toxicological studies on metal nanoparticles (NPs), it remains difficult to explain discrepancies observed between studies, largely due to the lack of positive controls and disconnection between physicochemical properties of nanomaterials with their toxicities at feasible exposures in a specified test system. In this study, we investigated effects of particle size and surface charge on in vitro mutagenic response and in vivo embryonic toxicity for newly synthesized silver nanoclusters (AgNCs) at human or environmental relevant exposure and compared the new findings with one of the most common nanoscale particles, titanium dioxide NPs (TiO2 NPs as a positive control). We hypothesized that the interaction of the test system and physicochemical properties of nanomaterials are critical in determining their toxicities at concentrations relevant with human or environmental exposures. We assessed the mutagenicity of the AgNCs (around 2 nm) and two sizes of TiO2 NPs (i.e., small: 5–15 nm, big: 30–50 nm) using a Salmonella reverse mutation assay (Ames test). The smallest size of AgNCs showed the highest mutagenic activity with the Salmonella strain TA100 in the absence and presence of the S9 mixture, because the AgNCs maintained the nano-size scale in the Ames test, compared with two other NPs. For TiO2 NPs, the size effect was interfered by the agglomeration of TiO2 NPs in media and the generation of oxidative stress from the NPs. The embryonic toxicity and the liver oxidative stress were evaluated using a chicken embryo model at three doses (0.03, 0.33, and 3.3 μg/g egg), with adverse effects on chicken embryonic development in both sizes of TiO2 NPs. The non-monotonic response was determined for developmental toxicity for the tested NPs. Our data on AgNCs was different from previous findings on AgNPs. The chicken embryo results showed some size dependency of nanomaterials, but they were more well correlated with lipid peroxidation (malondialdehyde) in chicken fetal livers. A different level of agglomeration of TiO2 NPs and AgNCs was observed in the assay media of Ames and chicken embryo tests. These results suggest that the test nanotoxicities are greatly impacted by the experimental conditions and the nanoparticle’s size and surface charge.

Safety evaluation of mutagenicity, genotoxicity, and cytotoxicity of Lactobacillus spp. isolates as probiotic candidates.

BackgroundProbiotics promote a healthy balance of gut bacteria and have many beneficial effects on human digestive physiology. Although, few side effects of probiotics have been reported. This study aimed to assess the safety of five probiotic candidate Lactobacillus strains isolated from healthy individuals by examining mutagenicity, genotoxicity, and oral toxic effects.MethodsFive selected candidate probiotic (SCPs) strains were evaluated for genotoxicity (Ames test with Salmonella typhimurium), in vitro mammalian chromosome aberration test and an in vivo mouse micronucleus assay on peripheral blood of mice. To evaluate the oral dose toxicity, BALB/c mice models were treated repeatedly (2000, 1000, and 500 mg/kg body weight /day) for 28‐days.ResultsThe Ames test performed for two S. typhimurium strains TA 98 and TA100 (both in the absence and in the presence of S‐9 metabolic activation system) did not show an increase in reverse mutation because of exposure to the SCPs in any of the doses (5.0, 2.5, 1.25, 0.625, and 0.3125 mg/plate). There was no genotoxicity in the SCPs treatment in the vitro chromosome aberration assay with Chinese hamster ovary cells (CHO‐K1). In addition, none of the tested strains increased the frequency of micronucleated reticulocytes in reticulocytes, the SCPs with the studied doses caused no substantial variation in the experimental groups compared to the negative control group (p > 0.05). SCPs were not acutely toxic when administered to male and female BALB/c mice by single gavage at (2000, 1000, and 500 mg/kg b.w/day) with no mortality or clinical signs, change in body weight or macroscopic abnormalities were observed in this dose range.ConclusionAs a result, SCPs did not induce mutagenic potential in vitro with bacterial reverse mutation, clastogenicity, and in vivo tests in the ranges of concentrations evaluated in our study.

Light-dependent enhancement of sulfadiazine detoxification and mineralization by non-photosynthetic methanotrophs

Co-metabolism and photodegradation are two approaches for remediating trace organic compounds (TOrCs), however, interactions between the two with regards to TOrCs degradation have not been elucidated. In this study, sulfadiazine (SDZ) was chosen as a representative TOrC and Methylocystis bryophila as a typical strain. Under light conditions, about 80.6% of SDZ was removed by M. bryophila, but only 7.6% or 28.9% of SDZ was eliminated by either individual photodegradation or by co-metabolism. The SDZ stimulated more extracellular organic matter (EOM) production by M. bryophila. The enhanced SDZ degradation was attributed to indirect photolysis caused by the excited triplet state of EOM (3EOM*) and co-metabolism. The UPLC-QTOF-MS analysis showed that due to co-metabolism, the pyrimidine ring was broken and could further be oxidized into smaller molecules under light conditions, such as formic and acetic acids. The SDZ mineralization ratio increased from 9.9% under the co-metabolic condition alone to 36.5% under co-metabolism coupled with photodegradation. The Ames tests confirmed that the SDZ degradation products by co-metabolism were mutagenic, however, their toxicity was ameliorated by light during co-metabolism. In conclusion, light plays a crucial role in co-metabolic processes of TOrCs.

Antioxidant capacity and toxicologic potential of Ibervillea sonorae plant

The main objective of this study was to evaluate the antioxidant capacity and possible toxicological potential of the infusions prepared from Ibervillea sonorae. For this, infusions were prepared from the root of I. sonorae, using the skin or the pulp. The antioxidant capacity of the infusions was evaluated through the ABTS and DPPH tests. While, the Artemia salina model, the MTT test using Vero and MCF-7 cells, and the Ames test were used to evaluate the possible toxicological potential. The results showed that I. sonorae peels were more lethal (60-80%) than pulp (<50%). Interestingly, I. sonorae pulp showed greater toxicity on Vero cells (IC50 = 222 µg / mL) than infusions made using the skins (IC50 = 379 µg / mL). Infusions of I. sonorae, either in the form of peel or pulp, showed high mutagenicity at 1000 and 500 µg/mL. The ingestion of infusions prepared from the I. sonorae plant could cause serious damage to health. DOI: https://doi.org/10.54167/tecnociencia.v16i2.903

Optimizing machine-learning models for mutagenicity prediction through better feature selection.

Assessing a compound's mutagenicity using machine learning is an important activity in the drug discovery and development process. Traditional methods of mutagenicity detection, such as Ames test, are expensive and time and labor intensive. In this context, in silico methods that predict a compound mutagenicity with high accuracy are important. Recently, machine-learning (ML) models are increasingly being proposed to improve the accuracy of mutagenicity prediction. While these models are used in practice, there is further scope to improve the accuracy of these models. We hypothesize that choosing the right features to train the model can further lead to better accuracy. We systematically consider and evaluate a combination of novel structural and molecular features which have the maximal impact on the accuracy of models. We rigorously evaluate these features against multiple classification models (from classical ML models to deep neural network models). The performance of the models was assessed using 5- and 10-fold cross-validation and we show that our approach using the molecule structure, molecular properties, and structural alerts as feature sets successfully outperform the state-of-the-art methods for mutagenicity prediction for the Hansen et al. benchmark dataset with an area under the receiver operating characteristic curve of 0.93. More importantly, our framework shows how combining features could benefit model accuracy improvements.

Comparative evaluation of three methylene dianiline isomers in the bacterial reverse mutation assay, the in vitro gene mutation test, and the in vitro chromosomal aberration test.

4,4'-MDA is classified as a genotoxic carcinogen based on numerous in vitro and animal data. The consequential assumption that a safe threshold does not exist is not only applied to 4,4'-MDA but also to its structural isomers and impurities 2,2'- and 2,4'-MDA in the absence of substance-specific data. This constitutes a problem in human risk assessments for all three substances as the inherent risks of 2,2'- and 2,4'-MDA and their contribution as impurities to that of 4,4'-MDA are essentially unknown. A comparative in vitro genotoxicity dataset consisting of the bacterial reverse mutation (Ames) test and the chromosomal aberration test in human lymphocytes (both performed according to the current OECD Guidelines) was generated for all three isomers. Furthermore, an in vitro gene mutation test in Chinese hamster ovary (CHO) cells (HPRT locus assay) was conducted with 2,4'-MDA. The results indicate differences regarding the genotoxic mechanism and potential, respectively, between the three structures and suggest that the no-threshold assumption for 4,4'-MDA may not be appropriate for 2,2'- and 2,4'-MDA.

Genotoxicity and 28-day repeated dose oral toxicity study of garlic essential oil in mice

Background and aimGarlic essential oil (GEO) isolated from Garlic (Allium sativum L.) exerts biological activities in disease prevention, particularly in metabolic and liver diseases, and is used for a dietary therapy for centuries. However, due to the side effects associated with the excessive consumption of GEO, there is a need to evaluate the safety of the GEO. Experimental procedureAmes test using five Salmonella typhimurium strains (TA98, TA100, TA102, TA1535, and TA1537) and Chinese hamster ovary (CHO–K1) cells with or without metabolic activation (S9 system), and mammalian erythrocyte micronucleus test were used to assess the genotoxicity and clastogenic effects of GEO. A repeated dose of GEO (15, 25, and 50 mg/kg body weight, p.o.) were administrated to ICR mice for 28 days to ascertain the subacute toxicity of GEO. Results and conclusionsThe results of the Ames test with or without S9 system indicated that GEO did not induce mutagenicity nor have clastogenic effects in CHO–K1 cells with or without S9 activation. Furthermore, GEO did not affect the ratio of immature to total erythrocytes or the number of micronuclei in immature erythrocytes of ICR mice after 24 and 48 h. In a 28-day oral toxicity assessment, GEO (15, 25, and 50 mg/kg body weight, p.o.)-fed ICR mice exhibited normal behaviors, mortality, body weight, daily intake, hematology, clinical biochemistry, and organ weight. GEO shows no genotoxicity, and the no-observed-adverse-effect level (NOAEL) for GEO is considered to be greater than 50 mg/kg bw/day orally for 28 days in mice.

Anti‐hypertensive effect of BPM4 on spontaneously hypertensive rats that antagonizes Dopamine βeta hydroxylase

BackgroundHypertension, which is described as chronically increased blood pressure above 140/90 mmHg, has emerged as a starring cause of mortality and morbidity worldwide. It is a leading risk factor for major cardiovascular events (such as heart attack and stroke), chronic kidney disease, heart failure, cognitive impairment and dementia. The dire consequences of uncontrolled hypertension indicate that controlling it will have a direct preventive effect on the vital organs. Despite the availability of a large number of classical anti‐hypertensive drugs it still affects humans adversely and poses a global disease burden thus necessitating new drugs. A non‐conventional target whose inhibition can regulate hypertension is Dopamine βeta hydroxylase (DBH). This enzyme, which plays a significant role in the catecholamine biosynthetic pathway, utilizes the body’s blood pressure (BP) lowering mechanism and thus can combat resistant hypertension.MethodologyIn the study, we have screened thousands of small molecules against the active site of DBH (PDB ID:4ZEL). Hits identified in silico were assessed for their inhibitory potential and strength of binding (Kd) in vitro. The ex vivo toxicity parameters were evaluated to get rid of toxic molecules before going for in vivostudies. Selected leads were screened in spontaneously hypertensive rats (SHRs) to evaluate their efficacy in reducing blood pressure as well as their ability to improve other cardiac parameters. Thereafter, acute as well as genotoxicity study was performed to minimize the rate of attrition in later phases of clinical development.ResultsOur best lead molecule “BPM4” inhibited DBH efficiently with an IC50 value of 18 μM and dissociation constant of 22 nM. BPM4 when administered through IP at a dose of 10 mg/kg BW significantly reduced BP with a p‐value of 0.001 as compared to the untreated group. Additionally, a significant reduction was observed in IVSs, IVSd, LVPWs and LVPWd, whereas increment was observed in internal diameter in the case of end systole and diastole (LVIDs and LVIDd). A high oral lethal dose (LD50) > 2000 mg/kg was observed in the acute toxicity test. Ames test showed no positive mutagenic effect +‐S9. No statistically significant decrease in % mitotic index nor any aberrant cells were observed in lymphocytes and bone marrow of rodents.ConclusionBPM4 (at 10 mg/kg BW) statistically reduced the BP as well as improved the left ventricular hypertrophy parameters in SHRS. The area of fibrosis was also decreased post treatment (cardiac fibrosis disrupts the normal myocardial structure and has a direct impact on heart function and survival). Subsequently, the compound successfully passed the three battery genotoxicity tests as it did not cause any structural damage to DNA nor did it induce any ploidy changes, holds scope for being a potent anti‐hypertensive drug candidate.

Cytotoxicity and genotoxicity evaluations of oleic acid and conjugated linoleic acid

Background and Aims: Oleic acid (OLA) and conjugated linoleic acid (CLA) occur in dairy products and meats and are also widespread at lower levels in many other foodstuffs. It is known that OLA and CLA are very bioactive compounds with substantially anti-carcinogenic effects. The objective of this study was to evaluate the cytotoxic potentials of OLA and CLA which were tested against cancerous and non-cancerous cell lines and to determine their genotoxicity. Methods: The cytotoxic activities of OLA and CLA against to cancer cell lines (U-87-MG, A549, MCF-7, CaCo-2, HeLa and PC3) and a control cell line (HEK293) were assessed by MTT assay. Ames MPFtm mutagenicity assay on 4 strains (TA98, TA100, TA 1535 and TA 1537) of Salmonella typhimurium was used for genotoxicity determination. Results: CLA showed cytotoxic activity on PC-3 cells, while OLA was created on A549 and PC-3 cell lines with the IC50 of 20 nM and 15 nM, respectively. No cytotoxic activity was observed on MCF-7, HeLa, U-87-MG, and CaCo-2 cells with the administered doses of OLA and CLA. It has been proved that OLA and CLA are characterized by a high cytotoxic activity towards cancer cells, as observed in the cell line test. There was no evidence for a mutagenic effect of OLA and CLA in the Ames test, with or without metabolic activation (S9) against Salmonella typhimurium strains. Conclusion: These in vitro test results indicate that these fatty acids can be considered a beneficial dietary supplement for enhancing anti-cancer therapy.

Comparative chemical analysis, mutagenicity, and genotoxicity of Petroleum refinery wastewater and its contaminated river using prokaryotic and eukaryotic assays.

Concern on the toxicity of final wastewater generated by the petroleum refining industry has increased in recent years due to the potential health threats associated with their release into the waterways. This study determined the mutagenic and genotoxic potential of petroleum refinery wastewater and a receiving river using the Ames fluctuation test onSalmonella typhimuriumstrains TA100 and TA98, SOS chromotest onEscherichia coliPQ37, and piscine peripheral micronucleus (MN) assay. Analyses of the physicochemical parameters, heavy metal, and organic contents of the samples were also performed. Ames test result showed that the two tested samples were mutagenic with TA100 strain as the more responsive strain for both the refinery wastewater and the river sample in terms of the calculated mutagenic index. A similar result was obtained in the SOS chromotest; however, the E. coli PQ37 system recorded a slightly higher sensitivity for detecting genotoxins than the Salmonella assay in the two samples. MN data showed induction of a concentration-dependent significant (p < 0.05) increase in the frequency of MN by both samples when compared with the negative control. Generally, the refinery wastewater induced the highest mutagenicity and genotoxicity compared to the river sample in the three assays used. Haemoglobin, platelets, red blood cells, mean corpuscular volume, total white blood cells, heterophils, haematocrit, and eosinophils reduced significantly with increased lymphocytes, basophils, mean corpuscular haemoglobin, and mean corpuscular haemoglobin concentration in fishes exposed to both samples. Total petroleum hydrocarbon, benzene, toluene, phenol index, polycyclic aromatic hydrocarbons, cadmium, mercury, nickel, lead, and vanadium contents analysed in the samples were believed to be responsible for the observed genotoxicity and mutagenicity. The findings of this study revealed that petroleum refinery wastewater is a potential mutagenic and genotoxic risk to the environment.

Computational studies, antimicrobial activity, inhibition of biofilm production, and safety profile of the cadmium complex of 1,10‐phenanthroline and cyanoguanidine

AbstractWe have previously determined the crystal structure of the metal complex [Cd(1,10‐phenanthroline)2(SO4)H2O]cyanoguanidine•5H2O that was characterized by spectroscopic measurements. The antibacterial activity was also determined by the agar diffusion test (zones of inhibition). In the present work, calculations based on density functional theory were performed to determine the contributions of the two different tautomers of cyanoguanidine to the vibrational IR and Raman experimental spectra. Moreover, the antimicrobial (antibacterial and antifungal) activity of the complex and cadmium ion was assessed and the minimum inhibitory concentration (MIC) was determined. The complexation increased the antimicrobial activity against Pseudomonas aeruginosaATCC 27853 and inhibited the biofilm production of this pathogen. Antimicrobial studies were coupled with the evaluation of safety, using the Artemia salinaand Ames test. At the concentrations tested, the cadmium complex behaved as a non‐mutagenic and non‐toxic compound. Therefore, the interaction of cadmium (II) with N and O‐containing ligands may provide a promising strategy for the development of novel and safe drugs with antimicrobial activity.

Response of Oreochromis niloticus (Teleostei: Cichlidae) exposed to a guanitoxin-producing cyanobacterial strain using multiple biomarkers

Changes in environmental conditions in aquatic ecosystems caused by anthropic actions can modify the composition of primary producers, promoting the excessive proliferation of cyanobacteria. These organisms can form cyanobacterial blooms, which directly affect aquatic life. The present study investigated the mutagenicity of the cyanobacterium Sphaerospermopsis torques-reginae (strain ITEP-024), guanitoxin-producing (natural organophosphate), and sublethal effects on fish in relevant environment concentrations. For this, the Ames test (Salmonella/microsome) was performed as a mutagenic assay for extracts of the ITEP-024 strain. Specimens of Oreochromis niloticus (Teleostei: Cichlidae) were subjected to acute 96 h exposure to different concentrations of aqueous extract of the strain: C = control group; T1 = 31.25 mg/L; T2 = 62.5 mg/L; T3 = 125 mg/L; and T4 = 250 mg/L. Genotoxic, biochemical, osmoregulatory, and physiologic biomarkers were analyzed. Our results showed that the cyanobacterium had a weak mutagenic response for the TA102 strain of Salmonella with and without metabolic activation by S9. Strains TA98 and TA100 were not affected. Fish from treatments T3 and T4 showed changes in oxidative stress (CAT, SOD, and GST enzymes), inhibition of the enzyme acetylcholinesterase activity, micronucleus formation, and osmoregulatory disorders. No guanitoxin accumulation was detected in the different tissues of O. niloticus by LC-MS/MS. Our results showed unprecedented mutagenicity data of the guanitoxin-producing cyanobacteria by the Ames test and biochemical, osmoregulatory, and genotoxic disorders in fish, providing efficient aquatic contamination biomarkers. Despite the great concern related to the presence of guanitoxin in blooms in freshwater ecosystems, its concentration is not yet regulated, and thus there is no monitoring agenda in current legislation.

Identification of potential human beta-secretase 1 inhibitors by hierarchical virtual screening and molecular dynamics

Alzheimer's disease (AD) is a neurodegenerative pathology responsible for 70% of dementia cases worldwide. Despite its relevance, the few drugs available for the treatment of this disease offer only symptomatic relief, with limited efficacy and serious adverse effects. The most accepted hypothesis about the pathogenesis involves the aggregation and deposition of β-amyloid peptides, mainly in the cerebral cortex and hippocampus, through the catalytic action of beta-secretase 1 (BACE-1), making this enzyme a promising target for the development of new drugs. In order to prioritize candidates for BACE-1 inhibitors, a hierarchical virtual screening by pharmacophore model and molecular docking was performed against the 216,833 molecules contained in several databases. Our previously built pharmacophore model was used for the first filtering step, which resulted in the selection of 399 molecules. The remaining molecules were filtered through molecular docking with GOLD 5.4.0. In this step, molecules with scoring values ​​greater than the mean plus standard deviation were evaluated for commercial availability and absence of asymmetric centers. Four molecules were selected and evaluated for mutagenic potential by the AMES test with the help of the pkCSM server. Finally, they were tested against the descriptors on Lipinski and Veber rules, and ZINC01589617 (QFIT = 56.52/Score = 44.95) satisfied all requirements, being subjected to molecular dynamics simulations (t = 100 ns) in order to obtain robust data on the mode of bonding and profile of intermolecular interactions. Those in silico strategies demonstrated that ZINC01589617 is a potential candidate for biological tests. Communicated by Ramaswamy H. Sarma

Safety evaluation of carbon tetrafluoride as an inert hyperbaric breathing gas in Sprague-Dawley rats

Carbon tetrafluoride (CF4) is an inert gas with higher molecular weight and lower water solubility than commonly used hyperbaric breathing gases. These inert gas properties decrease time required to decompress and avoid decompression sickness after deep dives. To assess CF4 toxicity, Sprague-Dawley rats were exposed to 8 atm absolute (ATA) air (10 males, 10 females) or 8 ATA 79% CF4/21% O2 (25 males, 25 females). Exposures were 30 min daily for 5 days. Rat behavior was normal throughout the testing period. There were no gross or microscopic pathology abnormalities following repeat dose exposure. Male body weight trends were similar between groups. Female body weight trends were 0.5 ± 0.8% day−1 for hyperbaric air exposure and − 0.2 ± 0.8% day−1 for hyperbaric CF4 exposure (P = 0.01) but remained within literature cited norms. Organ weights and hematologic indices remained within or near literature normal ranges. Clinical chemistry panels showed no signs of toxicity in renal or hepatic biomarkers. Polychromatic erythrocyte micronucleus frequency showed no chromosomal damage. Comet assay showed no DNA damage in lung tissue. Females exposed to CF4 had 2.5 times greater percent tail DNA in liver tissue than controls (P = 0.009). However this result remained within the normal range of local negative controls. A bacterial reverse mutation assay with exposure to 1 ATA 79% CF4/21% O2 for 72 h was nonmutagenic in four strains of Salmonella typhimurium and one strain of Escherichia coli. Overall, there was no evidence that CF4 caused organ toxicity or genetic toxicity.

Genotoxicity, acute and subchronic toxicity evaluation of fermented Morinda officinalis

Morinda officinalis has diverse pharmacological effects and has the potential to be used as functional food and medicine. Fermentation is traditionally used to process Morinda officinalis. However, the toxicological profile of fermented Morinda officinalis (FMO) is not reported. In the present study, the toxicological characteristics of FMO were assessed for the first time. FMO did not show any genotoxicity based on the Ames test, mammalian erythrocyte micronucleus test, and mouse primary spermatocyte chromosome aberration test. FMO administered by gavage in mice and rats at a dose of 20 g/kg BW did not induce death or toxicity based on acute study, indicating that FMO could be regarded as non-toxic at the tested dose. In the 90-day subchronic toxicity study, rats fed with FMO at the maximum dose of 8 g/kg BW did not affect mortalities, BW, food consumption, organ weights, hematology, serum biochemistry, or urinalysis. The no observed adverse effect level of FMO in both sexes was not less than 8 g/kg BW/day based on subchronic toxicity. The obtained results support the safe use of FMO as functional food and medicine.

2-{5-[(Z,2Z)-2-Chloro-3-(4-nitrophenyl)-2-propenylidene]-4-oxo-2-thioxothiazolidin-3-yl}-3-methylbutanoic Acid as a Potential Anti-Breast Cancer Molecule.

It was established that the synthesis of hybrid molecules containing a thiazolidinone and a (2Z)-2-chloro-3-(4-nitrophenyl)prop-2-ene structural fragments is an effective approach for the design of potential anticancer agents. Given the results of the previous SAR-analysis, the aim of the study was to synthesize a novel 4-thiazolidinone derivative Les-3331 and investigate its molecular mechanism of action in MCF-7 and MDA-MB-231 breast cancer cells. The cytotoxic properties and antiproliferative potential of Les-3331 were determined. The effect of the tested compound on apoptosis induction and mitochondrial membrane potential was checked by flow cytometry. ELISA was used to determine caspase-8 and caspase-9, LC3A, LC3B, Beclin-1, and topoisomerase II concentration. Additionally, PAMPA, in silico or in vitro prediction of metabolism, CYP3A4/2D6 inhibition, and an Ames test were performed. Les-3331 possesses high cytotoxic and antiproliferative activity in MCF-7 and MDA-MB-231 breast cancer cells. Its molecular mechanism of action is associated with apoptosis induction, decreased mitochondrial membrane potential, and increased caspase-9 and caspase-8 concentrations. Les-3331 decreased LC3A, LC3B, and Beclin-1 concentration in tested cell lines. Topoisomerase II concentration was also lowered. The most probable metabolic pathways and no DDIs risk of Les-3331 were confirmed in in vitro assays. Our studies confirmed that a novel 4-thiazolidinone derivative represents promising anti-breast cancer activity.

Safety Assessment of Lactiplantibacillus plantarum TWK10 Based on Whole-Genome Sequencing, Phenotypic, and Oral Toxicity Analysis.

Lactiplantibacillus plantarum TWK10 (TWK10), isolated from Taiwanese pickled cabbage, has been demonstrated to exert beneficial probiotic effects in both mice and humans. Here, we comprehensively assessed the safety of TWK10 using both in vivo and in vitro approaches, including whole-genome sequence analysis, an assessment of hemolytic activity, and performing an antimicrobial susceptibility test, the Ames bacterial reverse mutation assay, the chromosomal aberration test, a rodent peripheral blood micronucleus test, and the 28-day subacute oral toxicity assay. The results showed that there was no significant increase in the incidence of reverse mutations or chromosomal aberrations following exposure to TWK10. Moreover, no significant changes were detected either in the number of reticulocytes or the incidence of micronuclei in ICR mice, and no subacute toxicity was recorded in SD rats at the oral TWK10 dosage of 2000 mg/kg body weight/day repeated for 28 days. Additionally, TWK10 exhibited no hemolytic activity and was susceptible to all the antibiotics tested, except kanamycin. However, no antimicrobial resistance genes, virulence factors, or genes involved in biogenic amine synthesis were found in the genome of TWK10. Our findings demonstrated that TWK10 has high potential of being safe for human consumption as a probiotic.