Registration Dossier

Administrative data

Endpoint:
toxicity to other aquatic vertebrates
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
No data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: No guideline study used but comparable to national guidelines/standards

Data source

Reference
Reference Type:
publication
Title:
Evaluation of the developmental toxicity of trichloroethylene and detoxification metabolites using Xenopus
Author:
Fort DJ
Year:
1993
Bibliographic source:
Teratog., Carcinog., and Mutag. 13: 35-45

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
The Frog Embryo Teratogenesis Assay-Xenopus (FETAX) is a 96 h, whole-embryo, static-renewal assay using embryos of the South African clawed frog, Xenopus laevis. Because Xenopus embryos lack most major metabolic detoxification pathways through 4 days of development, the utility and versatility of FETAX has been significantly enhanced by the development, validation, and optimization of a complementary exogenous metabolic activation system (MAS).
GLP compliance:
no

Test material

Constituent 1
Reference substance name:
Oxalic acid
EC Number:
205-634-3
EC Name:
Oxalic acid
Cas Number:
144-62-7
IUPAC Name:
oxalic acid
Details on test material:
oxalic acid was purchased from the Sigma Chemical Company, St. Louis, Missouri.
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
No data

Sampling and analysis

Analytical monitoring:
no
Details on sampling:
No data

Test solutions

Vehicle:
no
Details on test solutions:
All solutions were changed every 24 h of the 4-day test, dead embryos removed, and fresh solutions added.

Test organisms

Test organisms (species):
Xenopus laevis
Details on test organisms:
TEST ORGANISM
- Common name: frog
- Scientific name: Xenopus laevis
- Age at study initiation (mean and range, SD): embryo

Study design

Test type:
other: static renewal
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Post exposure observation period:
No data

Test conditions

Hardness:
No data
Test temperature:
23±1°C
pH:
7
Dissolved oxygen:
No data
Salinity:
No data
Nominal and measured concentrations:
nominal
Details on test conditions:
TEST SYSTEM
- Test vessel: covered 60 mm plastic Petri dishes (Fisher Scientific, Houston, Texas)
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: Each treatment vessel contained a total of 8 ml of solution.
- No. of organisms per vessel: Four separate dishes of 20 embryos each were exposed to FETAX solution and designated FETAX solution controls.
Tests conducted with the MAS or inhibited MAS were also performed in duplicate with 20 embryos exposed per replicate concentration. Embryos were cultured at 23 -!-1.0°C.
- No. of vessels per concentration (replicates): Ten to sixteen concentrations were tested in duplicate.

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: The metabolites were each dissolved in an appropriate volume of FETAX solution, a reconstituted water media suitable for the culture of developing Xenopus embryos. Each metabolically activated treatment received 0.4 units/dish of N-nitrosodimethylamine activity, a NADPH generating system, and a penicillin-streptomycin mixture to control bacterial contamination.

OTHER TEST CONDITIONS
- Adjustment of pH: The pH of each of the stock solutions was adjusted to 7.0 with NaOH.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Following 96 h of exposure (stage 46 embryos), embryos were fixed in 0.7% formalin (pH 7.0), and the number of live malformed embryos were ascertained using a dissecting microscope.

TEST CONCENTRATIONS
- Test concentrations: For each treatment, 8 to 14 concentrations were tested. Controls, including FETAX solution, 1% v/v DMSO, uninhibited MAS (with and without DMSO), each inhibited MAS (with and without DMSO), activated acetylhydrazide (FETAX reference proteratogen), and unactivated toxicant, were tested simultaneously with each experiment. All control treatments received antibiotics, as well. One range-finding and two definitive concentration-response experiments were conducted with and without the MAS or inhibited MAS.
All solutions were changed every 24 h of the 4-day test, dead embryos removed, and fresh solutions added.
Reference substance (positive control):
not specified

Results and discussion

Effect concentrationsopen allclose all
Duration:
96 h
Dose descriptor:
LC50
Effect conc.:
5 330 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: 5030-5610
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
4 020 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
morphology
Remarks:
developmental toxicity
Remarks on result:
other: 3700-4400
Duration:
96 h
Dose descriptor:
other: Teratogenic index (ratio 96 h LC50 to 96 h EC50)
Effect conc.:
1.3
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: mortality and malformation
Remarks on result:
other: 1.1-1.7
Duration:
96 h
Dose descriptor:
other: MCIG (minimum concentrations to inhibit growth
Effect conc.:
4 000 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
morphology
Remarks:
growth
Details on results:
Control Results
In each of the experiments conducted, the FETAX solution, 1 % v/v DMSO, and MAS or MAS with 1 % v/v DMSO, control mortality and malformation rates were -5%. Mortality and malformation rates for the inhibited MAS failed to exceed 6%. Embryos exposed to 3.0 g/l acetylhydrazide and the intact MAS exhibited mortality and deformity rates of =90% and 100%, respectively.
Results with reference substance (positive control):
No data
Reported statistics and error estimates:
Litchfield-Wilcoxon probit analysis was used to determine the 96 h median lethal (LC50) and teratogenic (EC50) concentrations of two pooled definitive concentration-response tests. The 95% confidence intervals were calculated as well. A teratogenic index (TI) was calculated by taking the ratio of the 96 h LC50 value to the 96 h EC50 (malformation) value as a means of assessing teratogenic potential. Ninety-five percent fiducial intervals for the TI values were calculated by the method of Finney, Head-tail length of surviving embryos was measured as an index of growth using an IBM-AT compatible computer and Sigma Scan (Jandel Scientific, Corte Madra, CA) digitizing software. The length data were then used to calculate a minimum concentration to inhibit growth (MCIG) value for each experiment using the t-test (P <0.05).

Any other information on results incl. tables

Terata Induced in Xenopus by Exposure to Trichloroacetic Acid, Dichloroacetic Acid, Trichloroethanol, and Oxalic Acid

     Terata induced      
 Treatment  Concentration (mg/l) Malformation   n % Responding 
 Control b Gut miscoilingVisceral edemaMuscular kinking cSkeletal kinking  640640640640  2.41.01.40.8
 Oxalic acid > 3000> 5000> 7000  Gut miscoilingAbnormal mouth developmentMicroencephalyMicroophthalmia  80808080  2550100100 

a Defines concentration range of teratogenicity.

b Embryos exposed to FETAX solution alone. Pooled results from each test performed in study.

c Termmuscular kinking, caused most likely by abnormal somite development, is used to differentiate from that finding gtermed skeletal kinking, in which the spine (including the notocord) is affected.

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The 96 h LC50 was 5330 mg/l and the 96 h EC50 was 4020 mg/l
Executive summary:

Frog (Xenopus laevis) embryos were exposed to oxalic acid according to no specific guideline but in a Frog Embryo Teratogenesis Assay-Xenopus (FETAX). A 96 h LC50 of 5330 mg/l was determined. The 96 h EC50 was 4020 mg/l.