Fluazinam impairs oxidative phosphorylation and induces hyper/hypo-activity in a dose specific manner in zebrafish larvae.

Fluazinam is a pyridinamine fungicide that induces oxidative stress and mitochondrial damage in cells, and it has been reported to be neurotoxic. To characterize the biological effects of fluazinam, we assessed mitochondrial bioenergetics, dopamine system expression, and behavior of early life staged zebrafish (0.01 µM-0.5 µM). Fluazinam at environmentally-relevant levels did not induce sub-lethal effects in larvae, but at the LC50 (0.5 µM), fluazinam decreased basal and ATP-linked respiration significantly in embryos. As mitochondria are directly related to redox homeostasis and apoptosis, the expression of genes related to oxidative stress and apoptosis were measured. Superoxide dismutase 2 (sod2), heat stock protein 70 (hsp70), bcl2-associated X protein (bax), and caspase 9 (casp9) mRNA levels were up-regulated by 0.5 µM fluazinam. Taken together, there was evidence for mitochondrial dysfunction and oxidative damage at the highest concentration of fluazinam (0.5 µM) tested. As there are reports for fluazinam-induced neurotoxicity in dopamine synthesizing cells, transcriptional targets in the dopamine system were assessed in the zebrafish. Tyrosine hydroxylase 1 (th1) and dopamine receptor 2a (drd2a) mRNA levels were decreased by 0.5 µM fluazinam, suggesting that this fungicide may affect the dopaminergic system. To further assess the potential for fluazinam-mediated neuromodulation, the dark photokinesis response was assessed in larvae following exposure. Larvae exposed to 0.1 µM fluazinam showed hyperactivity, while larvae exposed to 0.2 and 0.3 µM showed hypo-activity. This study demonstrates that fluazinam disrupts mitochondrial bioenergetics in zebrafish, inducing an oxidative stress response, and aberrant behaviors in larvae that are dose dependent.

Comparison of Toxicities to Vibrio fischeri and Fish Based on Discrimination of Excess Toxicity from Baseline Level.

Investigations on the relationship of toxicities between species play an important role in the understanding of toxic mechanisms to environmental organisms. In this paper, the toxicity data of 949 chemicals to fish and 1470 chemicals to V. fischeri were used to investigate the modes of action (MOAs) between species. The results show that although there is a positive interspecies correlation, the relationship is poor. Analysis on the excess toxicity calculated from toxic ratios (TR) shows that many chemicals have close toxicities and share the same MOAs between the two species. Linear relationships between the toxicities and octanol/water partition coefficient (log KOW) for baseline and less inert compounds indicate that the internal critical concentrations (CBRs) approach a constant both to fish and V. fischeri for neutral hydrophobic compounds. These compounds share the same toxic mechanisms and bio-uptake processes between species. On the other hand, some hydrophilic compounds exhibit different toxic effects with greatly different log TR values between V. fischeri and fish species. These hydrophilic compounds were identified as reactive MOAs to V. fischeri, but not to fish. The interspecies correlation is improved by adding a hydrophobic descriptor into the correlation equation. This indicates that the differences in the toxic ratios between fish and V. fischeri for these hydrophilic compounds can be partly attributed to the differences of bioconcentration between the two species, rather than the differences of reactivity with the target macromolecules. These hydrophilic compounds may more easily pass through the cell membrane of V. fischeri than the gill and skin of fish, react with the target macromolecules and exhibit excess toxicity. The compounds with log KOW > 7 exhibiting very low toxicity (log TR < -1) to both species indicate that the bioconcentration potential of a chemical plays a very important role in the identification of excess toxicity and MOAs.

Discrimination of excess toxicity from baseline level for ionizable compounds: Effect of pH.

The toxic effect can be affected by pH in water through affecting the degree of ionization of ionizable compounds. Wrong classification of mode of action can be made from the apparent toxicities. In this paper, the toxicity data of 61 compounds to Daphnia magna determined at three pH values were used to investigate the effect of pH on the discrimination of excess toxicity. The results show that the apparent toxicities are significantly less than the baseline level. Analysis on the effect of pH on bioconcentration factor (BCF) shows that the log BCF values are significantly over-estimated for the strongly ionizable compounds, leading to the apparent toxicities greatly less than the baseline toxicities and the toxic ratios greatly less than zero. A theoretical equation between the apparent toxicities and pH has been developed basing on the critical body residue (CBR). The apparent toxicities are non-linearly related to pH, but linearly to fraction of unionized form. The determined apparent toxicities are well fitted with the toxicities predicted by the equation. The toxicities in the unionized form calculated from the equation are close to, or greater than the baseline level for almost all the strongly ionizable compounds, which are very different from the apparent toxicities. The studied ionizable compounds can be either classified as baseline, less inert or reactive compounds in D. magna toxicity. Some ionizable compounds do not exhibit excess toxicity at a certain pH, due not to their poor reactivity with target molecules, but because of the ionization in water.

Effect of exposure routes on the relationships of lethal toxicity to rats from oral, intravenous, intraperitoneal and intramuscular routes.

The lethal toxicity values (log 1/LD(50)) of 527 aliphatic and aromatic compounds in oral, intravenous, intramuscular and intraperitoneal routes were used to investigate the relationships of log 1/LD(50) from different exposure routes. Regression analysis shows that the log 1/LD(50) values are well correlated between intravenous and intraperitoneal or intramuscular injections. However, the correlations between oral and intravenous or intraperitoneal routes are relatively poor. Comparison of the average residuals indicates that intravenous injection is the most sensitive exposure route and oral administration is the least sensitive exposure route. This is attributed to the difference in kinetic process of toxicity testing. The toxic effect of a chemical can be similar or significantly different between exposure routes, depending on the absorption rates of chemicals into blood. Inclusion of hydrophobic parameter and fractions of ionic forms can improve the correlations between intravenous and intraperitoneal or oral routes, but not between intraperitoneal and oral routes. This is due to the differences of absorption rate in different exposure environments from different routes. Several factors, such as experimental uncertainty, metabolism and toxic kinetics, can affect the correlations between intravenous and intraperitoneal or oral routes.

Investigation on modes of toxic action to rats based on aliphatic and aromatic compounds and comparison with fish toxicity based on exposure routes.

The modes of toxic action (MOAs) play an important role in the assessment of the ecotoxicity of organic pollutants. However, few studies have been reported on the MOAs in rat toxicity. In this paper, the toxic contributions of functional groups in 1255 aromatic compounds were calculated from regression and were then compared with the toxic contributions in aliphatic compounds. The results show that some functional groups have same toxic contributions both in aromatic and aliphatic compounds, but some have not. To investigate the MOAs in rat toxicity, the distribution of toxic ratio (TR) was examined for well-known baseline and less inert compounds and thresholds of log TR=0.3 and 0.5 were used to classify baseline, less inert and reactive compounds. The results showed that some compounds identified as baseline compounds in fish toxicity were also classified as baseline compounds in rat toxicity. Except for phenols and anilines which were identified as less inert compounds in fish toxicity, aromatic compounds with functional groups such as ether, nitrile, nitrophenol, isocyanatoe and chloro were identified as less inert chemicals in rat toxicity. Reactive compounds identified in fish toxicity exhibit greater toxicity to rats. These compounds can undergo nucleophilic substitution, acylation and Schiff base formation with biological macromolecules. The critical body residues (CBRs) calculated from absorption and bioconcentration show that log 1/CBRs in rat toxicity are not equal to that in fish for some compounds. It suggests that the exposure route can affect the identification of MOAs between these two species for these compounds.

Relationship between lethal toxicity in oral administration and injection to mice: effect of exposure routes.

The lethal toxicity (LD50) in oral administration, intravenous, intraperitoneal, intramuscular and subcutaneous injections were used to investigate relationships of log 1/LD50 from different exposure routes. Regression analysis showed that log 1/LD50 in oral route was related to the toxicity in injection route. This relationship in lethality between the two routes is apparently due to the same mechanisms of the compounds to the same species. However, the scatter in the correlation curve indicates that exposure route is an important factor that influences the relationship. Some compounds with low intestinal absorption exhibit much less toxicity in oral administration than that in the injection route. A systemic bias of log 1/LD50 between oral and injection routes indicates that tissue distribution of compounds between blood and target site is a very rapid process, leading to log 1/LD50 in injection greater than those in oral administration. Although compounds can be metabolized in the body both from oral and injection routes, first-pass metabolism occurs in oral route but not in injection route. This will result in decrease of toxicity in oral route for most compounds as compared with injection route. In addition, experimental uncertainty, differences in gender, and species can also affect relationships of log1/LD50 between exposure routes.

Evaluation of toxicity data to green algae and relationship with hydrophobicity.

The quality of the biological activity data is of great importance for the development of algal quantitative structure-activity relationship (QSAR) models. However, a number of algal QSAR models in the literature were developed based on toxicity data without considering the response endpoints, exposure periods and species sensitivity. In this paper, 2323 algal toxicity data (log 1/EC50) in different toxicity response endpoints for 1081 compounds to 26 algal species within different exposure periods (14 and 15 min; 24, 48, 72, 96, 168 and 192 h) were used to evaluate the quality of the toxicity data to green algae. Analysis of 72 h toxicity to algae showed that the closed test had the same sensitivity as the open test for most of the test compounds, but a significant difference was observed for a few compounds. The overall average difference for all compounds ranges from 0.15 to 0.43 log units between toxicity endpoints (yield­growth rate). The relationships between exposure periods of 24, 48, 72 and 96 h indicated that 48 h exposure period is the most sensitive for algal growth inhibition test, and its sensitivity is 0.25 log units greater than 72 and 96 h exposure periods, respectively. Interspecies relationships showed that some algal species have very close sensitivity (e.g. Pseudokirchneriella subcapitata and Chlorella pyrenoidosa or Chlorella vulgaris and Scenedesmus obliquus, respectively), whereas some species have significantly different sensitivity (e.g. P. subcapitata and S. obliquus). Relationships between toxicity and hydrophobicity demonstrated that no difference was observed for non-polar narcotics within different exposure periods (24, 48, 72, and 96 h) or response variables (yield and growth rate). For polar narcotics, in contrast, algal toxicity is dependent on algal species and is related to the response variables and exposure period. We cannot expect significant QSAR models between algal toxicity and descriptors without considering species sensitivity, exposure periods and response endpoints.

Discrimination of excess toxicity from narcotic effect: influence of species sensitivity and bioconcentration on the classification of modes of action.

The toxicity data of 2624 chemicals to fish, Daphniamagna, Tetrahymenapyriformis and Vibriofischeri were used to investigate the effects of species sensitivity and bioconcentration on excess toxicity. The results showed that 47 chemical classes were identified as having the same modes of action (MOAs) to all four species, but more than half of the classes were identified as having different MOAs. Difference in chemical MOAs is one of the reasons resulting in the difference in toxic effect to these four species. Other important reasons are the difference in sensitivity and bioconcentration of species. Among the four species, V. fischeri has the most compounds identified as reactive MOA. This may be due to some compounds can be easily absorbed into the bacteria, react with the DNA or proteins, disrupt the normal function of the cell and exhibit significantly greater toxicity to the bacteria. On the other hand, the skin and lipid content of aqueous organisms can strongly inhibit the bio-uptake for some reactive compounds, resulting in a less toxic effect than expected. D. magna is the most sensitive species and T. pyriformis is the least sensitive species of the four species. For a comparison of interspecies toxicity, we need to use the same reference threshold of excess toxicity. However, some reactive compounds may be identified as baseline or less inert compounds for low sensitive species from the threshold developed from high sensitive species. The difference in the discrimination of excess toxicity to different species is not only because of the difference in MOAs for some compounds, but also due to the difference in sensitivity and bioconcentration.

Investigation on the relationship between bioconcentration factor and distribution coefficient based on class-based compounds: The factors that affect bioconcentration.

Bioconcentration factor (BCF) is one of the most important parameters in the assessment of the potential hazard of new compounds in aquatic ecosystems. However, the factors that influence the estimation of BCFs for a large variety of chemicals have not been systemically investigated in the literature. In this paper, a large BCF data set containing 1088 nonionic and ionic organic compounds was used to study the relationship between BCF and molecular descriptors and influencing factors. Step-by-step analysis on the class-based compounds showed that nonlinear Gaussian and Sigmoid equations could well describe relationships between logBCF and distribution coefficient for the compounds over a wide range of structures and chloro or/and bromo substituted aromatics, respectively. The quality of fit from the nonlinear models is better than the BCFBAF method from the Epi Suite program for the class-based compounds. Systemic prediction deviations have been observed for some types of compounds. The reasons for systemic deviations for these compounds can be attributed to the difference in bioconcentration mechanism for hydrophilic compounds, transformation for hydroxyphenols and three-membered rings, physical barrier for long chain and large polycyclic compounds, difference in determining methods of BCF (kinetic and steady-state), bioavailability for highly hydrophobic compounds and accuracy of BCF measurements for compounds with extremely high or low BCFs. These factors are important and should be considered in any reliable bioconcentration prediction.

Investigation on baseline toxicity to rats based on aliphatic compounds and comparison with toxicity to fish: effect of exposure routes on toxicity.

The aim of this paper was to investigate baseline toxicity to rats and effect of exposure routes on toxicity in rats and fish. In this paper, 1588 industrial chemicals were selected to investigate baseline toxicity to rats. The results showed that rat toxicity varies around a constant for classified compounds or homologues. The toxic contributions of substituted functional groups have been calculated and alkanes were used as baseline toxicity. The toxic contributions, equal to toxic ratios (TR), show that small changes in chemical structure can result in different toxic effect in rat toxicity. However, this situation has not been observed in fish toxicity because the threshold of excess toxicity (e.g. log TR=1) was too high to distinguish differences in toxicity. Very close critical body residues (CBRs) calculated from percentage of absorption and bioconcentration factors indicate that most of aliphatic chemicals may share the same modes of toxic action between rat and fish species. The high estimation error of bioconcentration factor calculated from computer programs for some compounds suggests that classification of excess toxicity should be based on the CBRs, rather than the TR because the TR is closely related to the exposure routes.

The discrimination of excess toxicity from baseline effect: effect of bioconcentration.

Toxic ratio TR is a valuable tool in the discrimination of excess toxicity from baseline effect. Although some authors realized that internal effect concentration or critical body residual (CBR) calculated from bioconcentration factor (BCF) should be used in the TR, the effect of BCF on the discrimination of excess toxicity from baseline effect has not been investigated. In this paper, 951 acute toxicity data to fish (LC50) and 1088 BCFs were used to investigate the relationship between TR and BCF. The results showed that some compounds identified as reactive compounds exhibit excess toxicity, but some do not. BCF is closely related to TR and can significantly affect the TR value. The real excess toxicity which is used to identify reactive chemicals from baseline should be based on the toxic ratio of internal effect concentrations, rather than on the ratio of external effect concentrations, TR. The use of LC50 alone to determine TR can result in errors in TR because toxicokinetics (as estimated by the BCF) are ignored. The foundation in the discrimination of excess toxicity from baseline effect is based on the linear relationship between log BCF and hydrophobicity expressed as log KOW. However, log BCF is not linearly related with log KOW for all the compounds. The BCFs with log KOW >7 or <0 are either overestimated or underestimated by the linear baseline BCF model. Parallel lines are observed from calculated log CBR values for baseline and less inert compounds. The log BCF values overestimated or underestimated by log KOW from the baseline BCF model can result in mis-prediction and mis-classification among baseline, less inert and reactive compounds.

Linear and non-linear relationships between soil sorption and hydrophobicity: model, validation and influencing factors.

The hydrophobic parameter represented by the octanol/water partition coefficient (logP) is commonly used to predict the soil sorption coefficient (K(oc)). However, a simple non-linear relationship between logK(oc) and logP has not been reported in the literature. In the present paper, soil sorption data for 701 compounds was investigated. The results show that logK(oc) is linearly related to logP for compounds with logP in the range of 0.5-7.5 and non-linearly related to logP for the compounds in a wide range of logP. A non-linear model has been developed between logK(oc) and logP for a wide range of compounds in the training set. This model was validated in terms of average error (AE), average absolute error (AAE) and root-mean squared error (RMSE) by using an external test set with 107 compounds. Nearly the same predictive capacity was observed in comparison with existing models. However, this non-linear model is simple, and uses only one parameter. The best model developed in this paper is a non-linear model with six correction factors for six specific classes of compounds. This model can well predict logK(oc) for 701 diverse compounds with AAE = 0.37. The reasons for systemic deviations in these groups may be attributed to the difference of sorption mechanism for hydrophilic/polar compounds, low solubility for highly hydrophobic compounds, hydrolysis of esters in solution, volatilization for volatile compounds and highly experimental errors for compounds with extremely high or low sorption coefficients.

Interspecies correlations of toxicity to eight aquatic organisms: theoretical considerations.

Interspecies correlations allow the prediction of toxicity to a number of other species. However, little attention has been paid to the theoretical considerations of the interspecies relationship based on the differences of bio-uptake and toxic mechanism between species. This study examines the interspecies correlations of toxicity between species of Vibrio fischeri, river bacteria, algae, Daphnia magna, carp, Tetrahymena pyriformis, fathead minnow and guppy based on the theoretical background. The results show that there are good interspecies correlations between marine bacterium and fresh water bacteria or fish and fish. It is suggested that compounds share the same bio-uptake and toxic mechanism of action between the species. On the other hand, poor interspecies relationships were found between toxicities to algae and T. pyriformis or D. magna. It is suggested that compounds have different toxic mechanisms of action between these species. Interspecies relationships can be improved by inclusion of the octanol/water partition coefficient or the energy of the lowest unoccupied molecular orbital. They reflect the difference of bio-uptake or toxic mechanism of action between species for organic compounds. Benzoic acids show very different toxicity contributions to the three species, V. fischeri, D. magna and carp. They can be easily absorbed into the unicellular bacteria, V. fischeri. On the contrary, the skin and lipid content of multicellular organisms, such as D. magna and fish, can strongly inhibit the bio-uptake for ionizable compounds, which results in the different toxic effect between V. fischeri and D. magna or carp. Good correlation coefficients were observed between toxicities to V. fischeri and D. magna or fishes by inclusion of hydrophobic and ionization parameters. V. fischeri or D. magna can serve as a surrogate of fish toxicity for hydrophobic and ionizable compounds studied. Toxic mechanisms of action are discussed based on the theoretical background of the interspecies correlation.

Classification of toxicity of phenols to Tetrahymena pyriformis and subsequent derivation of QSARs from hydrophobic, ionization and electronic parameters.

Phenolic compounds were classified into different groups based on the structure and functional groups of the phenol. Quantitative structure-activity relationship (QSAR) analysis was performed between the toxicity and octanol/water partition coefficient (logP) for these groups. The results showed that the toxicity of non-ionisable phenols is dependent on their hydrophobicity. Poor relationships were found between the toxicity and logP for ionisable compounds, and the use of methods based on logP to predict the toxicity of ionisable compounds can result in considerable errors. Ionized and unionized forms have different contributions to toxicity; the unionized form plays a more important role than the ionized form because the toxicity of organic acids and phenols decreases as the pH increases. In order to investigate the effect of ionization, the fraction of ionized and unionized forms of phenols at different pH values were calculated from the pK(a) values, and a corrected distribution partition coefficient (D(T)) was derived from QSAR analysis for ionisable compounds. The prediction of toxicity of non-reactive ionisable compounds was improved remarkably by using the D(T) parameter. Ionization not only affects the bio-uptake of ionisable compounds, but interaction with the receptor micromolecule can also depend on the electronic situation, which is also related to the ionization. Stepwise regression showed that the reactivity of ionisable phenols was strongly correlated with the fraction of negatively charged form (F(-)). Interpretable QSAR equations with good statistical fits were developed from hydrophobic, ionization and electronic parameters for 207 phenols.

Pancreatic injuries during laparoscopic urologic surgery.

OBJECTIVES: To review our experience with pancreatic injury (PI) associated with urologic laparoscopy. PI associated with urologic laparoscopy is uncommon. METHODS: From January 1999 to April 2004, 890 laparoscopic operations for upper urinary tract pathologic findings were performed, of which 574 were left-sided procedures. We reviewed the medical records for recognized intraoperative or postoperatively documented PI and recorded the patient demographic data and intraoperative and postoperative management. RESULTS: Four PIs were identified, all associated with left-sided procedures. The overall rate of PI was 0.44%. Two injuries occurred during left radical nephrectomy (2.1%) and two during left adrenalectomy (8.6%). The mean tumor size was 6 +/- 3.4 cm. The diagnosis was made intraoperatively in 1 patient, postoperative midepigastric pain with elevated serum amylase levels was the clinical presentation in 2 patients, and the incidental finding of pancreatic tissue in the final pathologic specimen confirmed the injury in the fourth patient. A prolonged pancreatic fistula developed only in 1 case. Three patients were treated conservatively and computed tomography-guided drainage of a pancreatic collection was required in one. The mean hospital stay was 18 days (range 4 to 57). CONCLUSIONS: PI is uncommon but can result in significant morbidity. Intraoperative detection and repair can minimize sequelae. Management with conservative measures and percutaneous drainage is usually successful, although a prolonged postoperative course can be expected.