4-chlorophthalic anhydride

Short-term toxicity to fish

Administrative data

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2003

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2003

Report date:

2003

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

4-chlorophthalic anhydride

EC Number:

204-251-9

EC Name:

4-chlorophthalic anhydride

Cas Number:

118-45-6

Molecular formula:

C8-H3-Cl-O3

IUPAC Name:

5-chloro-1,3-dihydro-2-benzofuran-1,3-dione

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Start and end of the test from each replicate test vessel

Test solutions

Vehicle:

no

Details on test solutions:

Water used for the acclimation of test organisms and for all toxicity testing was carbon filtered, deionized water collected at T.R. Wilbury Laboratories in Marblehead, Massachusetts. Water was adjusted to a hardness of 40 to 48 mg/L as CaCO3 and stored in polyethylene tanks where it was aerated and recirculated through particle filters, activated carbon, and an ultraviolet sterilizer. A sample of dilution water collected at the start of the definitive test had a hardness of 48 mg/L as CaCO3.

A stock solution of the test substance was prepared at a nominal concentration of 1000 mg/L by bringing 79.9574 g of the test substance (ground to a fine powder in a mortar) to a total volume of 75 liters with dilution water in a 200 liter polyethylene mixing tank. The mixing tank was equipped with a recirculating centrifugal pump and a mechanical mixer. The solution was mixed for approximately 22 hours and the pH of the stock was adjusted to 7.8 using 1 N sodium hydroxide. Appropriate amounts of the stock solution were added directly to dilution water to formulate the test concentrations. Nominal concentrations of the active ingredient were 0 mg/L (control), 150, 250, 400, 600 and 1,000 mg/L.

Test organisms

Test organisms (species):

Pimephales promelas

Details on test organisms:

Juvenile fathead minnow, Pimephales promelas, were procured from a commercial supplier (Thomas Fish Company, Anderson, CA, USA). Prior to testing, the fish were maintained under flow-through conditions in a 220 L polyethylene tank. During acclimation, fish were not treated for disease and they were free of apparent disease, injuries, and abnormalities at the beginning of the test. Fish were fed dry commercial fish food once per day except during the 48 hours preceding the test and they were not fed during the toxicity test. During the 14 day period before the start of the test, the acclimation temperature range was 21.5 to 22.9°C and the dissolved oxygen concentration was always at least 8.1 mg/L. Mortality during the last 48 hours of acclimation was less than 3%. At the conclusion of the test, the control fish had an average wet weight of 0.28 g (loading rate was 0.19 g/L) and they ranged in length from 3.2 to 3.8 cm.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Post exposure observation period:

None

Test conditions

Hardness:

48 mg/L as CaCO3 (at study start)

Test temperature:

21.9 - 22.8°C

pH:

7.2 - 7.7

Dissolved oxygen:

6.7 - 8.9 mg/L

Nominal and measured concentrations:

Nominal: 0 mg/L (control), 150, 250, 400, 600 and 1,000 mg 4-CLPA/L

Details on test conditions:

A range-finding test with 4-CLPA was conducted under static conditions with a control, a solvent control (0.10 mL/L dimethylformamide) and four concentrations of test substance: 0.10, 1.0, 10 and 20 mg/L. After 96 hours there was 100% survival in the control and solvent control and at all tested concentrations. No sublethal effects or insoluble material were observed at any tested concentration.

A second range-finding test with 4-CLPA was conducted under static conditions with a control and two concentrations of test substance: 100 and 1000 mg/L. A stock solution with a nominal concentration of 1000 mg/L was prepared without the use of a solvent and allowed to mix for approximately 23 hours. The pH of the stock was adjusted to 7.0 using 0.1 N sodium hydroxide. After 96 hours there was 100% survival in the control and at all tested concentrations. No sublethal effects or insoluble material were observed at any tested concentration.

The definitive toxicity test was also conducted under static conditions. Twenty fathead minnows were randomly distributed among two replicates of each treatment and control. The test was performed in 20-liter glass aquaria that contained 15 liters of test solution (water depth was approximately 18 cm). Test vessels were randomly arranged in a water bath during the 96 hour test (random numbers tables were used to add test organisms to test vessels, to select the location of each vessel, as well as the assignment of test concentrations). A 16 hour light and 8 hour dark photoperiod was automatically maintained with cool white fluorescent lights that provided a light intensity of approximately 41 footcandles. A 15 minute transition period was provided between dark and light. Aeration was not required to maintain dissolved oxygen concentrations above acceptable levels. The numbers of surviving organisms, the occurrence of sublethal effects and the presence of insoluble material were determined visually and recorded after 0, 24, 48, 72 and 96 hours.

Dissolved oxygen, pH, conductivity, and temperature were measured and recorded daily in each test vessel. The temperature in a control vessel was recorded continuously during the test.

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

> 1 070 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Effect concentrations 2

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

> 1 070 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Effect concentrations 3

Duration:

72 h

Dose descriptor:

LC50

Effect conc.:

> 1 070 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Effect concentrations 4

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 1 070 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Effect concentrations 5

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

1 070 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Details on results:

No insoluble material was observed at any tested concentration during the definitive toxicity test. Nominal concentrations of 4-chlorophthalic anhydride were: 0 mg/L (control), 150, 250, 400, 600 and 1,000 mg/L. Mean measured concentrations were: ND (not detected at or above the limit of quantitation), 164, 261, 422, 617 and 1,070 mg/L. The mean measured concentrations were 103 to 109% of the nominal concentrations. One hundred percent survival occurred and no sublethal effects were observed in the control. No sublethal effects were noted during the definitive toxicity test.

Results with reference substance (positive control):

NA

Reported statistics and error estimates:

Results of the toxicity test could not be interpreted by standard statistical techniques because there was at least 95% survival at all tested concentrations. The no observed effect concentration is the highest tested concentration at which there was no toxicant-related mortality or sublethal effect.

Any other information on results incl. tables

Sublethal observations / clinical signs:

Nominal Concentration of 4-CLPA (mg/L)	Measured Concentration of 4-CLPA (mg/L)
	0 Hour	96 Hours	Mean	% Recovery
Test Media Samples
Control	ND^1	ND	---	---
	ND	ND	ND	---
4-CLPA
150	157	169	164	109
	158	170
250	244	276	261	104
	262	260
400	414	436	422	106
	407	429
600	579	647	617	103
	594				646
1000	1020	1120	1070	107
	1020	1110
Laboratory Control Sample
400	424	388	406	102
Matrix Spike Sample
400	---	389	391	98
	---	392
Blank
0	ND	ND	ND	---
ND = none detected at or above the limit of quantitation of 0.0538 mg/L.

Applicant's summary and conclusion

Conclusions:

The 96-h LC50 value was determined at >1000 mg/L.

Executive summary:

The short-term toxicity to freshwater fish was determined in an OECD 203 study performed in compliance with GLP criteria. In this study, fathead minnows (Pimephales promelas) were exposed to nominal test concentrations of 0, 150, 250, 400, 600 and 1,000 mg 4-CLPA/L. The nominal test concentrations were analytically verified at the start and end of exposure and remained within ±20% of nominal. Therefore, the effect value is expressed as nominal. Fish mortality and general symptoms of toxicity were recorded after 24, 48, 72 and 96 hours exposure. No mortality was observed during the test and no sub lethal effects were noted. Based on these findings the 96-h LC50 value was determined at >1000 mg/L.