Testing Acute Toxicity in the Embryo of Zebrafish, Brachydanio rerio, as an Alternative to the Acute Fish Test: Preliminary Results

Acute toxicity testing in fish is a standard method used in estimating the influences of chemicals on aquatic vertebrates. The ecotoxicological data obtained from acute toxicity tests in fish are, however, not sufficiently reliable to justify the continued use of this test. Fertilised eggs of zebrafish ( Brachydanio rerio) were used to test the acute toxicity of chemicals. They were chosen because the development of B. rerio has been studied extensively and information already exists concerning the normal development of this species. The following parameters of the development of B. rerio were observed: coagulation of the egg, gastrulation, number of somites, movement, development of organ…

Comparison of testing acute toxicity on embryo of zebrafish, Brachydanio rerio and RTG-2 cytotoxicity as possible alternatives to the acute fish test

Abstract The present study compares two possible alternative methods to replace the acute fish test. Fertilized eggs of zebrafish, Brachydanio rerio were used to investigate the acute toxicity of chemicals. Different toxicological endpoints such as coagulation of the eggs, development of gastrulation, number of somites, development of organs, circulation, heartbeat, otolithanlage and pigmentation have been examined during the embryonic development of zebrafish within the first 48 h. Differences to the normal development were measured by these endpoints. Chemicals have been examined in a preliminary approach with respect to their cytotoxicity on the RTG-2 cells (Rainbow Trout Gonad). The vit…

The metabolism of phenol and substituted phenols in zebra fish.

1. The metabolism of five phenols in zebra fish was studied after uptake from the medium. The results showed no qualitative differences to other Cyprinid fish species, only the oxidation rate seemed to be lower. 2. Phenyl glucuronide, phenyl sulphate, and quinol sulphate were identified as metabolites of phenol. 3. Identified metabolites of 2-cresol were 2-cresyl glucuronide, 2-cresyl sulphate, and 2-hydroxybenzoic acid in trace amounts. 4. Only the glucuronide and sulphate conjugates were detected as metabolites of 4-nitrophenol, 4-chlorophenol, and pentachlorophenol.

Effect of substituted phenols on transaminase activity in the fish,Leuciscus idus melanotus L.

Postgraduale Weiterbildung mit dem zertifizierten Abschluss Fachökotoxikologin/e GDCh/SETAC

Influence of bacteria on the metabolite pattern of phenol in zebra fish (Brachydanio rerio)

Summary An investigation was performed on the potential influence of bacteria in the medium on the metabolism of phenol in zebra fish. The results obtained in vitro and in vivo show that bacteria may influence biotransformation studies of xenobiotics by fish in different manner. First, it was shown that phenol conjugates were deconjugated. Hydrolysis of phenyl glucuronide was observed both in vitro and in vivo. Deconjugation of phenyl sulphate was found to a minor extent merely under in vitro conditions. Second, the xenobiotic itself, in this case phenol, was degradated to CO2. The proportion did not lead to a significant removal of the test compound of the fish. Zusammenfassung Einflus von…

UV-Filtersubstanzen in Wasser und Fischen

Zur Erfassung der UV-Filtersubstanzen wird eine Methode vorgestellt, die die simultane Bestimmung dieser Verbindungen und von Organochlorverbindungen wie PCB und DDT in unterschiedlichen Fischkompartimenten bis in den ng/kg-Bereich ermoglicht. Die Nachweisgrenzen liegen zwischen 40 und 90 ng/kg Filet und die Wiederfindungsraten bewegen sich zwischen 78 und 104%. Die Belastung von Wasser und Fischen des Meerfelder Maares, Eifel, mit UV-Filtersubstanzen wurde 1991 und 1993 exemplarisch untersucht. Insgesamt konnten in den Fischen des Meerfelder Maares 6 UV-Filtersubstanzen identifiziert und quantifiziert werden. Die im Sommer 1991 gefangenen Barsche waren mit 2,0 mg/kg Fett (Summe nachgewiese…

Toxicokinetics of xenobiotics in zebrafish—Comparison between tap and river water

Abstract 1. 1. Uptake and elimination of lindane, 3,4-dichloroaniline, phenol and 4-nitrophenol by the zebrafish Brachydanio rerio were investigated in tap water and in water of the river Rhine. 2. 2. The differences in bioconcentration of chemicals between the two water types did not exceed a factor of 2.5. 3. 3. Elimination kinetics were comparable in tap and river water. 4. 4. It can be concluded that water of the river Rhine does not influence the kinetics of the investigated xenobiotics.

Elimination and distribution of different substituted phenols by frog (Rana temporaria) and crayfish (Astacus leptodactylus)

Ökotoxikologie. Eine junge Wissenschaft mit hohem Anspruch

Kinetic studies on the elimination of different substituted phenols by goldfish (Carassius auratus)

Contributions to the analysis of organic xenobiotics in fish

For the residue-analysis of fish samples, a method has been developed to separate the medium polar triazines and acetamides together with lipophilic xenobiotics (organochlorine pesticides, PCBs and octachlorostyrene) from fish tissue. The detection limits of the analytes are mostly within the ppt-range (ng/kg fillet) and vary between 40 and 1050 ng/kg fillet of fish. Their recoveries range from 75 to 108%, spiked at 1.5 μg/kg. The substances are extracted with a mixture of petroleum ether/ethylacetate (2:1) using a soxhlet apparatus. Subsequently, the lipids in the sample extracts are reduced to 0.4% by gel chromatography. A 5 g silica gel clean-up separates the analytes according to their …

Metabolism of chlorobenzene and hexachlorobenzene by the zebra fish, Brachydanio rerio.

It is now becoming evident that in many cases metabolites have a greater toxic potential than the parent xenobiotic exposed. For this reason, the knowledge of biotransformation pathways of xenobiotics plays a substantial role in environmental monitoring programs. The zebra fish (Brachydanio rerio, Hamilton-Buchanan) has received increased attention as a model species for freshwater fish in environmental surveillance programs to evaluate potential health and impacts of anthropogenic chemicals. The metabolism of phenol and various substituted phenols in zebra fish has recently been studied. The purpose of the present study was to examine the metabolism of chlorobenzene and hexachlorobenzene (…

Consequences of 3,4-dichloroaniline to guppy populations (Poecilia reticulata): computer simulation and experimental validation

Abstract In order to evaluate the effects of 3,4-dichloroaniline (3,4-DCA), which were detected in a life-cycle test with guppies, on the population level, laboratory guppy populations were examined with regard to regulative parameters. Among the endpoints investigated in life-cycle tests, fecundity was shown to be a sensitive parameter. In laboratory guppy populations, however, intraspecific predation on newborn was the main regulator. Based on these data, a computer simulation programme was used to predict the consequences of 3,4-DCA to guppy populations. A population exposure experiment was conducted to validate the simulation results. The effects detected in the life-cycle test (LOEC: 2…

Effect of 3,4-dichloroaniline on the early life stages of the zebrafish (Brachydanio rerio): results of a comparative laboratory study.

An early life-stage (ELS) test was conducted with the zebrafish and 3,4-dichloroaniline (3,4-DCA) in eight laboratories. Based on the results from all eight laboratories, an LOEC of 200 micrograms 3,4-dichloroaniline/liter applies for the early life stages of the zebrafish. Effects observed were reduction of the survival rate and malformations. If the 100 micrograms/liter concentration additionally tested by one laboratory is included in the assessment, an LOEC of 100 micrograms 3,4-DCA/liter is obtained for the survival rate and increase in length. The NOEC is 20 micrograms/liter. The present results of a comparative laboratory study with the zebrafish show that a 42-day ELS test can be co…

Bioconcentration, metabolism and toxicity of substituted anilines in the zebrafish (Brachydanio rerio).

The LC50 (96 h) values and the bioconcentration factors (BCF) for nine anilines (aniline; 2-, 3-, 4-chloroaniline; 2-, 3-, 4-nitroaniline; 2,4- and 3,4-dichloroaniline) in the zebrafish (Brachydanio rerio) were determined. Biotransformation products of anilines in the zebrafish were analyzed by HPLC. The aim of the investigations was to find relationships between accumulation/elimination/metabolism and toxicity on the one hand and between chemical structure and biotransformation on the other. We found a good correlation of log BCF and of log Pow with log LC50. This concurs with the assumption that the internal dose determines the toxicological effect. All anilines investigated, with the exc…

Toxicity of lindane, atrazine, and deltamethrin to early life stages of zebrafish (Brachydanio rerio).

Fertilized eggs of zebrafish were exposed under flow-through conditions to several concentrations of the following pesticides: lindane 40, 80, 110, 130, and 150 micrograms/liter; atrazine 300, 1300, and 9100 micrograms/liter; deltamethrin 0.5, 0.8, and 1.2 micrograms/liter. Hatching, abnormalities in development (external deformations, edema, etc.), and mortality were recorded over a period of 35 days. At the end of the experiment, the body lengths of the fish were measured. Survival of juvenile fish after 35 days was reduced by increasing concentrations of all xenobiotics tested: lindane enhanced the mortality from 110 micrograms/liter and atrazine from 1300 micrograms/liter, and deltameth…

Comparative diazinon toxicity in guppy and zebra fish: Different role of oxidative metabolism

Guppies pretreated with 1.1 μmol/L diazinon (0.5 times the LC50 with the guppy) for 7 d were less susceptible to the toxicity of the compound. The LC50 in these fish was about five times higher than that in nonpretreated animals. The lethal body burden and the slope of the concentration-response curve of diazinon in pretreated guppies were similar to those in zebra fish. The 7-d pretreatment of zebra fish with 12 μmol/L diazinon (0.5 times the LC50 with the zebra fish) did not affect the susceptibility of this species to diazinon. The pretreatment of guppies resulted in a strong inhibition of the formation of diazoxon and pyrimidinol during incubations of diazinon with the hepatic postmitoc…

Sex-specific reaction of liver ultrastructure in zebra fish (Brachydanio rerio) after prolonged sublethal exposure to 4-nitrophenol

Abstract Morphological alterations of the liver of zebra fish ( Brachydanio rerio ) following prolonged exposure to 0.1, 1 and 5 mg · l −1 4-nitrophenol (4-NP) were investigated by means of light and electron microscopy. Based on marked sexual dimorphism in control animals, liver recactions were both sex- and dose-dependent. Whereas at 0.1 mg·1 −1 only minor changes could be revealed, there were numerous structural modifications at 1 mg·1 −1 : Whereas male fish primarily react with a proliferation of smooth endoplasmic reticulum, female fish display a high degree of fenetrastion within cisternate of the rough endoplasmic reticulum. Both sexes exhibit infiltration of macrophages and lymphocy…

Kinetics and metabolism of 14C-lindane and 14C-atrazine in early life stages of zebrafish (Brachydanio Rerio)

Abstract Fertilized eggs, yolk sac larvae, and juveniles of zebrafish were exposed to 14C-lindane and 14C-atrazine under static conditions and the uptake was studied for a period of 48 hours. After 24 hours, a number of animals were transferred into pesticide-free running water and the elimination was examined for another 24 hours. The bioconcentration factors (BCFs) as well as the rate constants for uptake and elimination were calculated. Except for the lindane uptake by eggs and the atrazine uptake by juveniles, the steady state was reached within five hours. For all tested developmental stages the bioconcentration factor of lindane distinctly exceeded 100, whereas it was less than 10 in …

Effects of 3,4-dichloroaniline on fish populations. Comparison between r- and K-strategists: A complete life cycle test with the guppy (Poecilia reticulata)

In order to compare data derived from life cycle tests with zebrafish (Brachydanio rerio Ham.- Buch.), an r-strategist, with data of fish exhibiting a differing strategy of reproduction, a complete life cycle test with the guppy (Poecilia reticulata Peters) was performed with the chemical 3,4-dichloroaniline. The generation of guppies exposed during the whole life span (FI) reacted with more sensitivity than the generation exposed only as adults (Fo): Growth of adult females and reproduction was reduced significantly at 200 Μg/L in Fo, and even at 2 and 20 Μg/L in FI. The survival rates of the early life stages were not influenced at the tested concentrations. There is hardly any difference…

Toxicity of 3,4-dichloroaniline to perch (Perca fluviatilis) in acute and early life stage exposures

Abstract The toxitity of 3,4-dichloroaniline to perch was investigated in order to compare the sensitivity of crucial life stages with common test fish like zebrafish. Although in the acute toxicity test perch were more sensitive by a factor of five, larval sensitivity was in the same range in both species as well as in other egg-laying species. In spite of different water temperature and size, the zebrafish is a good model to predict early life stage toxicity of 3,4-DCA to the compared European freshwater species. Perch larvae are not suited for early life stage tests, as they are sensible towards experimental conditions and perform early cannibalism.

Bioconcentration and metabolism of 3,4-dichloroaniline in different life stages of guppy and zebrafish

Abstract Bioconcentration factors (BCF) and metabolism of 3,4-dichloroaniline in different life stages of guppy (Poecilia reticulata Peters ) and zebrafish (Brachydanio rerio Hamilton-Buchanan ) were investigated. The results prove that embryos of zebrafish, four-day-old yolk sac larvae, 17-day-old larvae and adult zebrafish are able to transform the chemical to 3,4-dichloroacetanilide. Bioconcentration factors in these life stages are between 30 and 42, exept yolk sac larvae showing a BCF of 86. 3,4-Dichloroacetanilide was also detected in extracts of newborn and adult guppies, but they are able to form an additional, yet unknown metabolite. The BCF varies from 34 in six-month-old guppies …

On the relevance of genotoxicity for fish populations II: genotoxic effects in zebrafish (Danio rerio) exposed to 4-nitroquinoline-1-oxide in a complete life-cycle test.

In order to characterize the impact of genotoxic potentials on populations of aquatic organisms in surface waters, zebrafish (Danio rerio) were exposed to the model genotoxicant 4-nitroquinoline-1-oxide (NQO) in a complete life-cycle test. Fish exposed to mean NQO concentrations of 0, 0.1, 0.3, 1.1, and 2.9 microg/l were examined by several genotoxicity assays with different endpoints. Assays included the unscheduled DNA synthesis (UDS) test, the comet assay, the alkaline filter elution, and the micronucleus test. The genotoxicity assays revealed an increasing genotoxicity, ranging from induction of DNA repair (even at the lowest concentration tested) to primary and secondary DNA alteration…