Tadayoshi Shigeoka

Cytotoxicity of 109 chemicals to goldfish GFS cells and relationships with 1-octanol/water partition coefficients

Abstract The midpoint cytotoxicity (NR50 values) of 109 chemicals including alcohols, aromatics, phenols and pesticides to goldfish scale GFS cells were compared with the fish acute toxicities and with the 1 - octanol/water partition coefficient (Pow). The NR50 values were significantly correlated to the available acute toxicity data for fathead minnow (n=31, r=0.958, s=0.480, F=325) and guppy (n=29, r=0.957, s=0.482, F=294). The Pow gave a good correlation in simple linear regression analysis for all compounds excluding 8 compounds that the NR50 values could not be obtained for their low solubility in culture medium. Log 1 NR 50 (mM) = (0.87±0.08) Log Pow − (2.17±0.24) ( n =101, r =0.914, s =0.587, F =505) For 29 narcotic type of compounds, the regression analysis revealed a high correlation between the NR50 values and Pow, and appeared to give a baseline cytotoxicity. Log 1 NR 50 (mM) = (0.77±0.06) Log Pow − (2.48±0.12) ( n =29, r =0.984, s =0.252, F =817) And also, the regression analyses between NR50 and Pow for some types of compounds were discussed. These results suggest that NR assay is useful for prediction of acute toxicity to fish and studies of quantitative structure-activity relationships (QSARs).

Embryonic development assay with Daphnia magna: application to toxicity of aniline derivatives.

An assay system using Daphnia magna embryos was applied to investigate the adverse effects of aniline derivatives. The data were compared with our previous data for chlorophenols. This new assay provides useful information to evaluate the toxicity of chemicals and the differences in sensitivity between the life stages. The effects of 15 aniline derivatives on embryonic development of D. magna embryos were determined. At the start of exposure, 2-6-h old eggs (between stages 1 and 2, round in shape, diameter approx. 400 microm), were used. In control and solvent control groups, embryonic development from an egg to a free-swimming animal proceeded completely within 3 days with more than 90% hatchability. Median effective concentrations (EC50s) to reduce the numbers hatched were determined and gross morphological abnormalities of hatched animals recorded. Anilines induced no obvious morphological abnormalities and no developmental delay although premature deaths occurred. However, they affected the number of embryos hatching in a dose-dependent manner. In addition, this embryo assay was more sensitive to aniline derivatives (except for aniline) than acute juveniles immobilization assay. Ratios of 48-h EC50 (juvenile)/3-day EC50 (embryo) for eight anilines were greater than 5.0. Particularly, the ratios of 4-methyl-, 4-ethyl- and 3-methylaniline were 77, 23 and 11, respectively. EC50s for embryos and juveniles were poorly correlated (r = 0.41). This indicated that the sensitivities of the two life stages were different to the effects of anilines. EC50s were poorly correlated (r = -0.097) with the log Kow (1-octanol/water partition coefficient). These results were compared with previous results for phenols.

In vitro cytotoxicity of 45 pesticides to goldfish GF-scale (GFS) cells

Abstract The cytotoxicities of 45 pesticides including 18 organophosphates and 7 carbamates to GF-Scale (GFS) cells, derived from the scale of goldfish ( Carassius auratus ), were determined with the 24-hr neutral red assay. The assay revealed that 13 pesticides were classified into the highly cytotoxic group (NR50 ≦ 10 mg/L), while 17 pesticides were into the highly cytotoxic group (NR50 ≦ 10 mg/L), while 17 pesticides were moderate (10

Relationship between molar refraction and n-octanol/water partition coefficient

The n-octanol/water partition coefficient (Pow) for organic chemicals which do not form the hydrogen bond was considered on the basis of Londons dispersion force, because the dispersion force is the main component of van der Waals-type attractive forces between solute and solvent molecules. The dispersion force is a function of both the ionization potential and the molar refraction. As the variation of the ionization potential is small in comparison with that of the molar refraction, the latter is the most important factor to determine the value of Pow. Thus the relationship between Pow and the molar refraction can be established. The relationship was confirmed for some halogenated alkanes, alkenes and aromatics, and polycyclic aromatic hydrocarbons by analysis of observed values of Pow and calculated values of the molar refraction. Estimation of Pow from the molar refraction is advantageous for the preliminary hazard assessment of new chemicals, because direct measurement of Pow is laborious and time consuming for lack of information about analytical methods.

Cytotoxicity of chlorophenols to goldfish GFS cells with the MTT and LDH assays

The cytotoxicity of phenol and 14 chlorophenols (CPs) were examined in goldfish scale (GFS) cells, a fibroblastic cell line derived from goldfish scales. MTT reduction and intracellular LDH activities were examined as cytotoxicity indicators. Cell densities at inoculation were 1.0 x 10(5) cells/ml in the MTT assay and 0.5 x 10(5) or 0.1 x 10(5) cells/ml in the LDH assay. No change in the midpoint cytotoxicity concentration (IC(50)) was observed in either assay at the same order of cell density (0.5-1.0 x 10(5) cells/ml). However, at low chemical concentrations, the LDH assay was more sensitive than the MTT assay. In addition, for many CPs, the LDH assay at 0.1 x 10(5) cells/ml was more sensitive than the MTT and LDH assays at the higher cell density levels. The correlation coefficients (r) for all IC(50) determinations were greater than 0.97. Good correlations between these assays and the neutral red (NR) assay were also observed (r > 0.97). The 1-octanol/water partition coefficient (log P) gave good correlation coefficients (r > 0.90) with IC(50) values of both the MTT and LDH assays in simple linear regression analyses. The study demonstrates that both the MTT and LDH cytotoxicity assays with GFS cells, together with the NR assay, can be applied to screen the acute toxicities of chemicals.

Non-steady-state equilibrium model for the preliminary prediction of the fate of chemicals in the environment.

Prediction of the fate of chemicals in the environment is essential to the assessment of the potential hazard of chemicals. The fate of a chemical in the environment depends mainly upon (1) transfer processes between environmental compartments due to physicochemical properties, (2) transformation processes in each compartment, and (3) characteristics of the environment. Many equilibrium or kinetic models have been proposed to predict the fate of chemicals in the environment. Though kinetic models are useful in predicting the concentration-time profile of a chemical at the non-steady state, they are rather complicated. In this work is proposed a simple and useful equilibrium model for prediction of the mass and concentration distribution fraction, mean residence time, and concentration-time profile of a chemical released into the environment, an advantage of our model. For the validation of this model, the results were compared with data from Neelys pond experiment and field data monitored by the Japanese Environment Agency. The results show that the equilibrium model is valuable for preliminary prediction of the fate of chemicals that are priority chemicals for hazard assessment in the environment.

Mutational spectra of benzo[a]pyrene and MeIQx in rpsL transgenic zebrafish embryos

To evaluate the rpsL transgenic zebrafish (Brachydanio rerio) mutation assay, we treated the embryos with benzo[a]pyrene (B[a]P) (10 microg/ml) or 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) (300 microg/ml) for 16h and determined the mutation spectra. These treatments were previously reported to induce mutant frequencies that were 4.3 and 2.4 times the control value, respectively. In the B[a]P-treated group, half of the mutations were single base substitutions, 74% of which occurred at G:C base pairs. Among G:C base pair substitutions, G:C to T:A and G: C to C:G transversions were predominant, suggesting that B[a]P induced mutations in zebrafish embryos by mechanisms previously described in mammalian tissues. In the MeIQx-treated group, about 60% of the mutations were deletions. Some specific mutations were found, but the compound primarily amplified the background mutation level; improvement in the conditions of treatment may be required for elucidating MeIQx-mutagenesis in this system. This study showed that transgenic zebrafish may be a useful tool for detecting mutagens in aquatic environments and for elucidating mutagenic mechanisms.

Evaluation of aquatic environmental fate of 2,4,6-trichlorophenol with a mathematical model

Abstract A mathematical model, Simplified Approach for Fate Evaluation of Chemicals in Aquatic Systems (SAFECAS), was applied to evaluate fate of 2,4,6-trichlorophenol(TCP) in three types of aquatic environments; a river, an oligotrophic lake, and an eutrophic lake. Since phenols are dissociative substances, equilibrium constants and rate constants for transfer and transformation processes were measured or estimated as a function of pH. Results suggest that most of TCP is distributed to the water phase in aquatic environments, and is almost removed by the advective outflow in the river and by the photodegradation in lakes.

Multi‐phase non‐steady state equilibrium model for evaluation of environmental fate of organic chemicals

A simple mathematical fate model, Multi‐Phase Non‐Steady State Equilibrium Model (MNSEM) is proposed to evaluate distribution, persistence, and concentrations of chemicals in a model environment consisting of air, water, soil and sediment phases. The model is applied to evaluation of environmental fate and concentration of trichloroethylene and 1,4‐dichlorobenzene under generic conditions representative of Japan. Evaluated chemical concentrations in air are within a factor of 3 of average values in Japanese atmosphere, and evaluated concentrations in water, sediment, or fish are greater than an order of magnitude below detection limits in real environments, so that evaluated concentrations are in reasonable agreement with environmental measurement data in Japan. Although MNSEM is not a model for site‐specific evaluation of environmental fate, results suggested that this model is an adequate method to aid in evaluation of fate of chemicals under generic environment conditions. Evaluated concentration‐profi...

Cytotoxicity of anilines and aldehydes to goldfish GFS cells and relationships with 1-octanol/water partition coefficients

Abstract The cytotoxicites of 13 anilines and 10 aldehydes to goldfish scale GFS cells were determined with the neutral red assay. The sequence of cytotoxicity was based on the concentration of chemicals that reduced uptake of neutral red by 50 % (NR50 values). Among a series of aniline derivatives, the parent aniline molecule was the least cytotoxic, with potency increasing with the progressive incorporation of chlorine atoms or with the alkyl chain length into the aromatic ring structure. Among a series of aldehyde derivatives, methanal (formaldehyde) and propanal, which have less than two carbon atoms of the alkyl chains, were more cytotoxic than predicted from the alkyl chain length. The in vitro cytotoxicity of these chemicals except for methanal was found to be significantly correlated to their in vivo acute toxicity to guppies. Moreover, NR50 values were significantly correlated with 1-octanol/water partition coefficients (Pow) excluding the lower aliphatic aldehydes.