In vitro comparative cytotoxic assessment of pristine and carboxylic functionalized multiwalled carbon nanotubes on LN18 cells.

Multiwalled carbon nanotubes (MWCNTs) have been used in biomedical applications due to their ability to enter the cells. Carboxylic functionalization of MWCNT (MWCNT-COOH) is used to mitigate the toxicity of MWCNTs. Our study focuses on comparing the toxicity of MWCNT and MWCNT-COOH on the neuronal cells, LN18. Concentrations of 5, 10, 20, and 40 µg ml-1 were used for the study, and cytotoxicity was determined at 0, 1, 3, 6, 12, 24, and 48 h of incubation. Cell viability was assessed by Trypan Blue, MTT, and Live dead cell assays, and the oxidative stress produced was determined by reactive oxygen species (ROS) and Lipid peroxidation assays. MWCNT-COOH showed higher cell viability than MWCNT for 20 and 40 µg ml-1 at 24 and 48 h. This was also visually observed in the live dead cell imaging. However, at 48 h, the morphology of the cells appeared more stretched for all the concentrations of MWCNT and MWCNT-COOH in comparison to the control. A significant amount of ROS production can also be observed at the same concentration and time. Viability and oxidative stress results together revealed that MWCNT-COOH is less toxic when compared to MWCNT at longer incubation periods and higher concentrations. However, otherwise, the effect of both are comparable. A concentration of 5-10 µg ml-1 is ideal while using MWCNT and MWCNT-COOH as the toxicity is negligible. These findings can further be extended to various functionalizations of MWCNT for wider applications.

Assessment of the Cytotoxicity of Peptide Modifications of Doxorubicin on Tetrahymena pyriformis.

The cytotoxicity of doxorubicin (Dox) and its peptide modifications Z-Gly-Pro-Dox and Boc-Gly-Pro-Dox were studied. Tetrahymena pyriformis was used as a test system, which made it possible, due to the short life cycle and high reproduction rate of ciliates, to trace their response to the effects of toxicants over several generations. It was found that peptide modification of the Dox molecule markedly reduces its cytotoxic and cytostatic effect. The Z-Gly-Pro-Dox modification has less cytotoxic and cytostatic effect compared to Boc-Gly-Pro-Dox. When determining the ability of drugs (at a concentration of 100 µM) to prevent bacterial contamination of samples, it was shown that the smallest degree of overgrowth was recorded in the presence of Dox (OD600nm 81.1). Boc-Gly-Pro-Dox also had a bacteriostatic effect, though less pronounced (OD600nm 93.8). The degree of overgrowth in the presence of Z-Gly-Pro-Dox was close to that of distilled water. The results obtained on ciliates did not contradict the data obtained in similar studies on mice.

A cross-industry collaboration to assess if acute oral toxicity (Q)SAR models are fit-for-purpose for GHS classification and labelling.

This study assesses whether currently available acute oral toxicity (AOT) in silico models, provided by the widely employed Leadscope software, are fit-for-purpose for categorization and labelling of chemicals. As part of this study, a large data set of proprietary and marketed compounds from multiple companies (pharmaceutical, plant protection products, and other chemical industries) was assembled to assess the models' performance. The absolute percentage of correct or more conservative predictions, based on a comparison of experimental and predicted GHS categories, was approximately 95%, after excluding a small percentage of inconclusive (indeterminate or out of domain) predictions. Since the frequency distribution across the experimental categories is skewed towards low toxicity chemicals, a balanced assessment was also performed. Across all compounds which could be assigned to a well-defined experimental category, the average percentage of correct or more conservative predictions was around 80%. These results indicate the potential for reliable and broad application of these models across different industrial sectors. This manuscript describes the evaluation of these models, highlights the importance of an expert review, and provides guidance on the use of AOT models to fulfill testing requirements, GHS classification/labelling, and transportation needs.

Toxicity assessment of zinc sulfate: A commonly used compound.

Because zinc sulfate (ZnSO4) is widely used in many fields such as biomedicine, electronics, and chemistry, it is important to evaluate its toxic effects. In this study, the cyto-genotoxic effects of ZnSO4 on meristematic cells in the root tip of Allium cepa L. were investigated. After calculating the effective concentration (EC50 = 70 ppm) of ZnSO4, A. cepa root tip cells were suspended for 24, 48, 72, and 96 h in solutions of 35 ppm (EC50/2), 70 ppm (EC50), and 140 ppm (EC50 × 2) concentrations. Using the counts of dividing cells, the mitotic index (MI) was calculated. Chromosome aberration index (CAI) was determined from percentages of abnormal cells. When the obtained data were statistically evaluated, it was determined that all application concentrations caused a significant decrease in MI and an increase in CAI compared to the control group (distilled water). It was concluded that increased ZnSO4 dose concentrations and exposure times caused cytotoxicity and genotoxicity in the root cells of A. cepa L.

Biogenic Silver Nanoparticles: Assessment of Their Cytotoxicity, Genotoxicity and Study of Capping Proteins.

The development of nanotechnology in the last two decades has led to the use of silver nanoparticles (AgNPs) in various biomedical applications, including antimicrobial, anti-inflammatory, and anticancer therapies. However, the potential of the medical application of AgNPs depends on the safety of their use. In this work, we assessed the in vitro cytotoxicity and genotoxicity of silver nanoparticles and identified biomolecules covering AgNPs synthesized from actinobacterial strain SH11. The cytotoxicity of AgNPs against MCF-7 human breast cancer cell line and murine macrophage cell line RAW 264.7 was studied by MTT assay, cell LDH (lactate dehydrogenase) release, and the measurement of ROS (reactive oxygen species) level while genotoxicity in Salmonella typhimurium cells was testing using the Ames test. The in vitro analysis showed that the tested nanoparticles demonstrated dose-dependent cytotoxicity against RAW 264.6 macrophages and MCF-7 breast cancer cells. Moreover, biosynthesized AgNPs did not show a mutagenic effect of S. typhimurium. The analyses and identification of biomolecules present on the surface of silver nanoparticles showed that they were associated with proteins. The SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) analysis revealed the presence of 34 and 43 kDa protein bands. The identification of proteins performed by using LC-MS/MS (liquid chromatography with tandem mass spectrometry) demonstrated their highest homology to bacterial porins. Capping biomolecules of natural origin may be involved in the synthesis process of AgNPs or may be responsible for their stabilization. Moreover, the presence of natural proteins on the surface of bionanoparticles eliminates the postproduction steps of capping which is necessary for chemical synthesis to obtain the stable nanostructures required for application in medicine.

Assessment of cyto- and genotoxic effects of Cesium-133 in Vicia faba using single-cell gel electrophoresis and random amplified polymorphic DNA assays.

The aim of this study was to evaluate the ecotoxic effect of high concentration cesium (Cs) exposure on plant root growth and its toxicological mechanism. The radicle of broad bean (Vicia faba) was selected as experimental material. The cytotoxic and genotoxic effects of plants exposed to different Cs levels (0.19-1.5 mM) for 48 h were evaluated using scanning electron microscopy (SEM), X-ray fluorescence (XRF) analysis, single-cell gel electrophoresis (SCGE) and random amplified polymorphic DNA (RAPD) assays. The results showed that radicle elongation decreased clearly after 48 h of exposure treatment with different concentrations of Cs solution. The root cell structure was obviously damaged in the Cs treatment groups (0.19-1.5 mM). At a Cs concentration of 1.5 mM, the percentages of viable non-apoptotic cells, viable apoptotic cells, non-viable apoptotic cells, and non-viable cells were 40.09%, 20.67%, 28.73%, and 10.52%, respectively. SCGE showed DNA damage in radicle cells 48 h after Cs exposure. Compared with the control group, the percentage of tail DNA in Cs exposed group (0.38-1.5 mM) increased by 0.56-1.12 times (P < 0.05). RAPD results showed that the genomic stability of V. faba radicles decreased by 4.44%-15.56%. This study confirmed that high concentration Cs exposure had cytotoxicity and genotoxicity effects on plants.

Assessment of allelopathic, cytotoxic, genotoxic and antigenotoxic potential of Smilax brasiliensis Sprengel leaves.

Smilax brasiliensis (Smilacaceae) is a native Brazilian plant found in the Cerrado biome and commonly used in folk medicine. The aim of this study was to evaluate the allelopathic, cytotoxic, genotoxic, and antigenotoxic potential of extract and fractions of Smilax brasiliensis leaves. Quercetin and rutin isomers were observed in the subfractions. The dichloromethane fraction (1000 µg/mL) decreased lettuce (Lactuca sativa) seed vigor, while and ethyl acetate and hydromethanol fractions (1000 µg/mL) affected the germination, and quercetin and rutin affected the vigor and germination of onion seeds. The extract, fractions, quercetin, and rutin inhibited or promoted lettuce hypocotyl and radicle growth. The extract and fractions inhibited onion hypocotyl growth at all concentrations. With regards to radicle growth, the results were diversified: growth was either inhibited or promoted. Rutin and quercetin inhibited onion hypocotyl and radicle growth at all concentrations. The extract and fractions of Smilax brasiliensis, rutin, and quercetin did not cause cytotoxic effect evaluated by mitotic index. The extract and fractions showed genotoxic effects. Quercetin and rutin did not cause genotoxic effects. On the other hand, the extract and fractions showed antigenotoxic effects at all tested concentrations, where they were able to revert chromosomal abnormalities caused by glyphosate. However, additional studies are required to evaluate the possible use of the S. brasiliensis leaf methanol extract and fractions as natural sources of bioherbicides.

Assessment of the cytotoxic and genotoxic potential of pillar[5]arene derivatives by Allium cepa roots and Drosophila melanogaster haemocytes.

In this study pillar[5]arene (P5) and a quinoline-functionalized pillar[5]arene (P5-6Q) which is used for detecting radioactive element, gas adsorption and toxic ions were synthesized. These materials were characterized by Nuclear Magnetic Resonance (NMR), Fourier Transform Infrared (FTIR), elemental analysis, melting point, Mass Spectroscopy, Scanning Electron Microscopy (SEM) and Zeta Potential. The cytotoxic and genotoxic potential of P5 and P5-6Q at distinct concentrations of 12.5, 25, 50, and 100 µg/mL were also investigated by Allium ana-telophase and comet assays on Allium cepa roots and Drosophila melanogaster haemocytes. P5 and P5-6Q showed dose dependent cytotoxic effect by decreasing mitotic index (MI) and genotoxic effect by increasing chromosomal aberrations (CAs such as disturbed anaphase-telophase, polyploidy, stickiness, chromosome laggards and bridges) and DNA damage at the exposed concentrations. These changes in P5-6Q were lower than P5. Further research is necessary to clarify the cytotoxic and genotoxic action mechanisms of P5 and P5-6Q at molecular levels.

Pathogenesis of local necrosis induced by Naja atra venom: Assessment of the neutralization ability of Taiwanese freeze-dried neurotoxic antivenom in animal models.

Naja atra envenomation is one of the most significant clinical snakebite concerns in Taiwan. Taiwanese freeze-dried neurotoxic antivenom (FNAV) is currently used clinically for the treatment of cobra snakebite, and has been shown to limit the mortality of cobra envenomation to less than 1%. However, more than half of victims (60%) require surgery because of local tissue necrosis, a major problem in patients with cobra envenomation. Although the importance of evaluating the neutralizing effect of FNAV on this pathology is recognized, whether FNAV is able to prevent the local necrosis extension induced by N. atra venom has not been investigated in detail. Cytotoxins (CTXs) are considered as the major components of N. atra venom that cause necrosis. In the current study, we isolated CTXs from whole cobra venom and used both whole venom and purified CTXs to develop animal models for assessing the neutralization potential of FNAV against venom necrotizing activity. Local necrotic lesions were successfully produced in mice using CTXs in place of whole N. atra venom. FNAV was able to rescue mice from a subcutaneously injected lethal dose of cobra venom; however, it was unable to prevent CTX-induced dermo-necrosis. Furthermore, using the minimal necrosis dose (MND) of CTXs and venom proteome data, we found a dose of whole N. atra venom suitable for FNAV and developed a workable protocol for inducing local necrosis in rodent models that successfully imitated the clinical circumstance of cobra envenoming. This information provides a more comprehensive understanding of the pathophysiology of N. atra envenomation, and serves as a guide for improving current antivenom strategies and advancing clinical snakebite management in Taiwan.

Stochastic dynamics and survival analysis of a cell population model with random perturbations.

We consider a model based on the logistic equation and linear kinetics to study the effect of toxicants with various initial concentrations on a cell population. To account for parameter uncertainties, in our model the coefficients of the linear and the quadratic terms of the logistic equation are affected by noise. We show that the stochastic model has a unique positive solution and we find conditions for extinction and persistence of the cell population. In case of persistence we find the stationary distribution. The analytical results are confirmed by Monte Carlo simulations.

Algorithm of Molecular and Biological Assessment of the Mechanisms of Sensitivity to Drug Toxicity by the Example of Cyclophosphamide.

Comparative study of the liver, blood, and spleen of DBA/2JSto and BALB/cJLacSto mice sensitive and resistant to acute toxicity of the cyclophosphamide allowed us to reveal basic toxicity biomarkers of this antitumor and immunosuppressive agent. Obtained results can be used for the development of an algorithm for evaluation of toxic effects of drugs and food components.

Is hyperthermic intraperitoneal chemotherapy (HIPEC) safe for healthcare workers?

BACKGROUND: During hyperthermic intraperitoneal chemotherapy (HIPEC), caregivers are exposed by different routes to cytotoxic drugs. This review proposes an overview of the safety of HIPEC by assessing existing data on protection procedures, biological and non-biological samples. Based on these data, relevant good practices, eventual irrelevant overprotection procedures and missing data to implement adapted protections are highlighted. MATERIALS AND METHODS: Data were extracted from a systematic review of literature from 1980 till 2016: number and type of surgical procedure, healthcare professionals present, protective equipment, samples, pre-analytical method and analytical method. RESULTS AND DISCUSSION: Only 55 HIPEC procedures have been evaluated. The majority of antineoplastic drugs used have all required characteristics to penetrate the organism and are recognized as very dangerous. Moreover, a great heterogeneity in protective equipment used, either individual or collective is observed. Environmental contamination occurs during HIPEC, especially for all surfaces in the operating room. Compounds penetration into caregivers lungs cannot be excluded. Priority remains to prove professionals contamination by focusing on biological samples. Biological material is rarely sampled or samples are not necessarily adapted. CONCLUSION: Repeated blood tests should be preferred using appropriate sampling schedules and validated sensitive analytical methods. Furthermore, there is a great need of new biological indicators to monitor caregivers exposure. During hyperthermic intraperitoneal chemotherapy (HIPEC), healthcare workers are exposed by different routes to cytotoxic drugs. There are currently few available occupational exposure data and environmental monitoring and biomonitoring must be improved in order to ensure optimal protection against antineoplastic drugs.

A comparative assessment of cytotoxicity of commonly used agricultural insecticides to human and insect cells.

The cytotoxic potential of 13 commonly used agricultural insecticides was examined using cell-based systems with three human HepG2, Hek293, HeLa cells and three insect Tn5B1-4, Sf-21, and Drosophila S2 cells. Data showed that (1) an enhancement of some insecticides (e.g. pyrethroids) on cells proliferation; (2) an inhibition of some insecticides on cells viability; (3) various levels of susceptibility of different cells to the same insecticide; and (4) the cell type dependent sensitivity to different insecticides. The degree of cytotoxicity of insecticides on human cells was significantly lower than that on insect cells (P<0.05). Methomyl, even 20µg/ml, showed little cytotoxicity at 24h exposure whereas emamectin benzoate possessed the strongest cytotoxic potential in a dose-dependent fashion. The results revealed comparable cytotoxic property of agricultural insecticides against intact cells.

In Vitro Toxicological Assessment of Cobalt Ferrite Nanoparticles in Several Mammalian Cell Types.

Nanoparticles have been widely used in various fields due to the superior physicochemical properties and functions. As a result, human exposure to nanoparticles increases dramatically. Previous researches have shown that nanoparticles could travel through the respiratory, digestive system, or skin into the blood and then to the secondary organs such as the brain, heart, and liver. Besides, the nanoparticle toxicity is controversial and dependent on the sensitivity of the cell type, route of exposure, and condition, as well as their characteristics. Similarly, cobalt ferrite nanoparticles (CoFe2O4-NPs) have been used in different industrial fields, and have also various application possibilities in medical and biomedical fields. CoFe2O4-NPs induce toxic responses in various organisms such as human, mice, and algae. However, there is a serious deficit of information concerning their effects on human health and the environment. We aimed to investigate the toxic effects of CoFe2O4-NPs on liver (HepG2), colon (Caco-2), lung (A549), and neuron (SH-SY5Y) cells, which reflect different exposure routes in vitro, by using various toxicological endpoints. The cytotoxicity, genotoxicity, oxidative damage, and apoptosis induction of CoFe2O4-NPs (39 ± 17 nm) were evaluated. After 24 h, the nanoparticles decreased cell viability at ≤100 µg/mL, while increasing viability at >100 µg/mL. CoFe2O4-NPs induced DNA and oxidative damage with increased malondialdehyde (MDA) and 8-hydroxy deoxyguanosine (8-OHdG) levels and decreased glutathione (GSH) levels with no change in protein carbonyl (PC) levels. CoFe2O4-NPs had apoptotic effect in HepG2 and Caco-2 cells in a concentration-dependent manner and necrotic effects on SH-SY5Y and A549 cells. Consequently, the adverse effects of CoFe2O4-NPs should raise concern about their safety in consumer products.

Artemia salina as a new index for assessment of acute cytotoxicity during co-composting of sewage sludge and lignocellulose waste.

Considering the necessity to constantly monitor the safety of use of sewage sludge, we have focused on evaluating the toxicity of raw sludge and sludge treated by co-composting with date palm waste using an in vitro assessment of cytotoxicity based on Artemia salina larvae as a simple new sensitive and reliable routine test. The efficiency of co-composting in decreasing sludge toxicity was evaluated in terms of cytotoxicity abatement reaching 100% by the second month of composting for mixture A (1/3 sludge+2/3 date palm waste) and the third month for mixture B (1/2 sludge+1/2 date palm waste). Cytotoxicity abatement was confirmed by the increase of germination index, which reached over 100% with positive correlation for lettuce (R(2)=0.81 and 0.86) and for turnip (R(2)=0.87 and 0.74) for mixtures A and B respectively. A strong correlation between the proposed cytotoxicity test and the evolution of regulatory physical-chemical approaches was found, (R(2)=0.88 and 0.89) for NH4(+)/NO3(-) and (R(2)=0.80 and 0.88) for C/N respectively for mixture A and B. These findings allow the inexpensive bioassay reported to be used as a highly sensitive test to determine the cytotoxicity and maturity of composts.

In vivo micronucleus test in the assessment of cytogenotoxicity of landfill leachates in three animal models from various ecological habitats.

The in vivo micronucleus (MN) test, a standard test for the genotoxicity screening of xenobiotics, was used to evaluate the cytotoxic and genotoxic activities of landfill leachates in Clarias gariepinus, Coturnix coturnix japonica and Rattus norvegicus. These organisms were exposed to various sub-lethal concentrations (1-50%) of Olusosun and Aba Eku landfill leachates. At post exposure, peripheral erythrocytes from catfish and quail, and bone marrow cells of quail and rat were subjected to MN analysis following standard protocols. The leachates induced significant increase in MN formation and total nuclear abnormalities (NAs) in the peripheral erythrocytes of catfish and quail. NAs occurred in the order; BN > BL > LB > NT in the catfish and BN > BudN > TLN > TN in quail. There was significant increase in MN formation in the bone marrow cells of quail, and micronucleated polychromatic erythrocytes and micronucleated normochromatic erythrocytes formation in the bone marrow of rats. The concentration dependent significant (p < 0.05) decrease in the PCE/NCE ratio in the bone marrow of the leachate treated rats suggest alterations in the bone marrow cell proliferation, leading to the suppression of immature erythrocytes (PCE). MN induction showed positive corrections with leachate concentrations in the test organisms; and it increased with exposure duration in the catfish. Indiscriminate disposal of solid waste generates leachates containing multiple xenobiotics that are capable of increasing genomic instability among vertebrates inhabiting various ecological habitats.

Differential gene transcription, biochemical responses, and cytotoxicity assessment in Pacific oyster Crassostrea gigas exposed to ibuprofen.

Pharmaceuticals, such as anti-inflammatory nonsteroidal drugs, are frequently detected in aquatic ecosystems. Studies about the effects of these substances in nontarget organisms, such as bivalves, are relevant. The aim of this study was to evaluate the effects on antioxidant status caused by ibuprofen (IBU) in oysters Crassostrea gigas exposed for 1, 4, and 7 days at concentrations 1 and 100 µg L(-1). Levels of IBU in tissues of oysters, as well as cell viability of hemocytes, were measured. The transcription of cytochrome P450 genes (CYP2AU2, CYP356A1, CYP3071A1, CYP30C1), glutathione S-transferase isoforms (GST-ω-like and GST-π-like), cyclooxygenase-like (COX-like), fatty acid binding protein-like (FABP-like), caspase-like, heat shock protein-like (HSP70-like), catalase-like (CAT-like), and the activity of catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione S-transferase (GST) were also evaluated in the gills of oysters. The highest levels of IBU were observed in animals exposed to 100 µg L(-1). A significant upregulation of CYP2AU1, CYP356A1, CYP3071A1, GST-ω-like, GST-π-like, COX-like, and FABP-like was observed in oysters exposed to IBU under different experimental conditions. Oysters exposed to 1 µg L(-1) for 7 days showed a significantly higher transcription of CYP2AU2, CYP356A1, CYP3071A1, GST-ω-like, and GST-π-like but lower GR activity. In conclusion, C. gigas exposed to environmentally relevant concentrations of IBU (1 µg L(-1)) exhibited increased transcription of certain genes and alterations on antioxidant and auxiliary enzymes, which could, in the the long term, cause damages to exposed organisms.

Cytotoxic 1,5-diaryl-3-oxo-1,5-pentadienes: an assessment and comparison of membrane permeability using Caco-2 and MDCK monolayers.

A number of cytotoxic conjugated unsaturated ketones were screened for their membrane permeability characteristics using Caco-2 and MDCK cells with the view of finding promising leads for in vivo evaluations. 3b-e and 4a-b demonstrated high permeability characteristics. In particular, 4a emerged as a promising lead which showed excellent apparent permeability (P(app): 54.70) and efflux ratio (ER: 0.15) values. In general, the relative apparent permeabilities of these enones are similar in both bioassays.

In situ electrochemical assessment of cytotoxicity of chlorophenols in MCF-7 and HeLa cells.

An in situ electrochemical method was used to assess the cytotoxicity of chlorophenols using human breast cancer (MCF-7) and cervical carcinoma (HeLa) cells as models. On treatment with different chlorophenols, the electrochemical responses of the selected cells, resulting from the oxidation of guanine and xanthine in the cytoplasm, indicated the cell viability. In addition, the in situ in vitro electrochemical method was further compared with the traditional MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays. Although similar cytotoxicity data were obtained from both methods, the effective concentrations of chlorophenols that inhibited 50% cell growth (EC50 values) from the electrochemical method were only slightly lower than those from the MTT assay. These results indicate that the in situ in vitro electrochemical method paves a simple, rapid, strongly responsive, and label-free way to the cytotoxicity assessment of different chlorophenol pollutants.

Cytotoxic effect of linear alkylbenzene sulfonate on human intestinal Caco-2 cells: associated biomarkers for risk assessment.

Linear alkylbenzene sulfonate (LAS) is a synthetic anionic surfactant widely present in the environment due to its intensive production and use in the detergency field. Admitting that current procedure of risk assessment has limits in providing realistic risk assessment data and predicting the cumulative effect of the toxicant mixtures, the incorporation of information regarding the mode of action and cell response mechanism seems to be a potential solution to overcome these limits. In this regard, we investigated in this study the LAS cytotoxicity on human intestinal Caco-2 cells, trying to unveil the protein actors implicated in the cell response using proteomics approach in order to give a better understanding of the toxicological effect and allow the identification of appropriate biomarkers reflecting the mode of action associated with LAS. As results, we demonstrated that LAS induces a time- and dose-dependent cytotoxicity in Caco-2 cells accompanied by an induction of oxidative stress followed by an excessive increase of intracellular calcium level. Proteomics approach helped in discovering three informative biomarkers of effect associated with LAS cytotoxic effect, reported for the first time: calreticulin, thioredoxin, and heat shock cognate 71 (HSP7C), confirmed by real-time PCR and western blot analysis. These biomarkers could serve for more reliable future risk assessment studies that consider the toxicants mode of action in order to help in the prediction of potential cumulative effects of environmentally coexisting contaminants.