N-(2-ethylhexyl)-8,9,10-trinorborn-5-ene-2,3-dicarboximide

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Remarks:

Flow-through conditions

Type of information:

experimental study

Adequacy of study:

key study

Study period:

7 April 1990 to 4 March 1991

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EPA OPP 72-1 (Fish Acute Toxicity Test)

GLP compliance:

yes

Specific details on test material used for the study:

MGK 264, Code No. 209-90

Analytical monitoring:

yes

Details on sampling:

Test solutions of MGK 264 were analytically quantified by ABC personnel at ABC Laboratories. This was done at 0- and 96-hours of the study. Analysis of the test water samples for MGK 264 was accomplished based upon a method developed at ABC Labs, based upon information provided by a representative of MGK. Preparation of the water samples and the determination of MGK 264 were performed using the following procedure.
Fifty milliliters of aquatic test water were collected from each aquaria and placed in 70 ml culture tubes. Fortifications were prepared ranging from 0.147 to 7.33 mg/L. Ten mL of toluene with 0.02% polyethylene glycol (PEG) were added to each tube. The samples were shaken vigorously for approximately 1 minute. The phases were allowed to separate and the toluene with 0.02% PEG was transferred to 15 mL culture tubes. Dilutions were then prepared using toluene with 0.02% PEG such that the concentration of the sample would fall within the standard curve range.

Vehicle:

yes

Remarks:

dimethylformamide (DMF)

Details on test solutions:

A diluter stock solution (40,000 mg/L) was prepared by dissolving 10.765 g of MGK 264 in 250 ml of dimethylformamide (DMF) or a similar ratio. The injector volume was adjusted to deliver 0.2 mL of stock to 2 liters of dilution water. The resulting nominal concentration regime of MGK 264 was 0.25, 0.50, 1.0, 2.0 and 4.0 mg/L. The solvent control chamber received an aliquot of 0.1 mL of DMF which was approximately equivalent to that received by the highest test concentration.

A primary stock standard of MGK 264 at a concentration of 1.93 mg/ml in acetone was prepared on February 16, 1990 and was stored at room temperature. Subsequent dilutions were prepared in toluene with 0.02% polyethylene glycol (PEG) for LC chromatography standards and in acetone for the preparation of quality control fortification samples.

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

The 140 rainbow trout (lot #6789) used for this test were obtained from Mt. Lassen Trout Farms in Red Bluff, California. The test fish were received as eyed eggs on November 30, 1989. After hatching, the fish were reared and maintained in well water and were fed newly hatched brine shrimp or a commercially available fish food daily. Approximately six weeks before the test, the fish were placed in a "living stream". The living stream is an enclosed system in which water is recirculated through the system. The fish were acclimated to the soft blended water and approximate test temperature in this system. The laboratory environment was maintained on a 16-hour daylight photoperiod. Fish culture techniques were basically those described by Brauhn and Schoettger (2). A daily record of fish observations during the holding period is included in Appendix I.

48h before the initiation of the test, fish were removed from the culture tank and placed in the temperature acclimation unit. During this time, the fish were held without food. At test initiation, rainbow trout were distributed to the test chambers in an impartial manner. The rainbow trout (Oncorhynchus mykiss) used as the control group during this study had a mean weight of 2.38 (± 0.43) g and a mean standard length of 55 (± 3) mm when measured at the end of the test. The biomass loading was 0.24 g/L/day. The control group was believed to be representative of all groups.

Test type:

flow-through

Water media type:

other: Soft Blended Water

Limit test:

no

Total exposure duration:

96 h

Post exposure observation period:

N/A

Hardness:

44-46 mg/L (as CaC03)

Test temperature:

11.5 to 13°C

pH:

7.7 to 7.9

Dissolved oxygen:

83 and 86%

Salinity:

Not Measured

Conductivity:

Conductivity 100-110 µMhos/cm

Nominal and measured concentrations:

The mean measured concentrations were 0.20, 0.41, 0.83, 1.7 and 4.1 mg/L. These results represented 80 to 103% of the nominal test concentrations of 0.25, 0.50, 1.0, 2.0 and 4.0 mg/L.

Details on test conditions:

A proportional diluter system described by Mount and Brungs and a Hamilton Model 420 syringe dispenser were used for the intermittent introduction of MGK 264 test solutions and diluent water into each test chamber. Five exposure concentrations, solvent control and dilution (soft blended) water control comprised the test design. Twenty fish were added to each concentration, solvent control and dilution water control chamber at test initiation. Chemical characteristics, typical of soft blended water, are provided in Table 1 of the attached report.
The diluter delivered 1 liter of test solution or control water to the test vessels at an average rate of 8.4 times per hour over the course of the study. This flow rate resulted in 201 liters of water flowing through the 30 liter aquaria per day and represented 6.7 tank volume replacements in each chamber everyday. The test aquaria were immersed in a circulating water bath that was thermostatically held at 12 ± 1°C.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

> 0.83 - < 1.7 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: LC50 was determined to be 1.4 mg/L with 95% confidence limits of 0.83 and 1.7 mg/L

Key result

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

0.41 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

other: lack of mortality or behavioral/sublethal effects

Details on results:

The flow-through toxicity test concentrations of MGK 264 to rainbow trout (Oncorhynchus mykiss) were 0.25, 0.50, 1.0, 2.0 and 4.0 mg/L. The mean measured concentrations from the 0- and 96-hour analyses were 0.20, 0.41, 0.83, 1.7 and 4.1 mg/L. The test material appeared to be stable in the flow through test system. The 96 hour LC50 for rainbow trout exposed to MGK 264 was 1.4 mg/L with 95% confidence limits of 0.83 and 1.7 mg/L. The 96 hour no-effect concentration for this study was 0.41 mg/L. An oily surface film was observed in the mixing cell and the 1.7 and 4.1 mg/L test solutions. Since the measured concentrations were 85 and 103% of the nominal values, this surface film should not affect the integrity of the study.

Results with reference substance (positive control):

Not Applicable

Reported statistics and error estimates:

The 96 hour LC50 for rainbow trout exposed to MGK 264 was 1.4 mg/L with 95% confidence limits of 0.83 and 1.7 mg/L.
The 96 hour no-effect concentration for this study was 0.41 mg/L.
96-hour slope of dose-response line = 6.6

Validity criteria fulfilled:

not specified

Conclusions:

The flow-through toxicity test concentrations of MGK 264 to rainbow trout (Oncorhynchus mykiss) were 0.25, 0.50, 1.0, 2.0 and 4.0 mg/L. The mean measured concentrations from the 0- and 96-hour analyses were 0.20, 0.41, 0.83, 1.7 and 4.1 mg/L. The test material appeared to be stable in the flow through test system. The 96 hour LC50 for rainbow trout exposed to MGK 264 was 1.4 mg/L with 95% confidence limits of 0.83 and 1.7 mg/L. The 96 hour no-effect concentration for this study was 0.41 mg/L.

Executive summary:

A 96-hour flow-through toxicity study of MGK 264 to rainbow trout (Oncorhynchus mykiss) was conducted for McLaughlin Gormley King Company from April 7, 1990 to April 11, 1990. Twenty fish per test concentration plus soft blended water control and solvent control were used in a nominal dosing regime of 0.25, 0.50, 1.0, 2.0 and 4.0 mg/L. Analytical measurements of MGK 264 were made at 0 hours and 96 hours. The measured concentrations averaged 0.20, 0.41, 0.83, 1.7 and 4.1 mg/L, respectively.

Water chemistry measurements of dissolved oxygen, pH and temperature were consistent throughout the term of the experiment and were within the protocol specifications.

 

From the data collected during this study, a 96 -hour LC50 (with 95% confidence limits) was calculated to be 1.4 mg/L (0.83 and 1.7 mg/L) based on the mean measured concentrations of MGK 264. The slope of the 96 -hour dose-response line was 6.6. Complete mortality occurred in the 4.1 mg/L test concentration within 24 hours. Eighty percent mortality was also elicited by the 1.7 mg/L test concentration by 96 hours. Sublethal/behavioral responses (e.g., surfacing, labored respiration, dark discoloration, erratic swimming, flared gills, fish on bottom of test chamber, quiescence and loss of equilibrium) were noted among the fish in the 0.83 and 1.7 mg/L test levels. As determined by this study, a 96 -hour no-effect concentration of MGK 264 toxicity to rainbow trout was 0.41 mg/L based on a lack of sublethal responses at this concentration.

Description of key information

The LC50 concentration for MGK 264 which is considered to be the most suitable for use in the CSA is from the rainbow trout flow-through study and was determined to be 1.4 mg/L. The flow-through study has been selected over the static study for setting the end point value to be used in the CSA based on concerns over adsorption / bioaccumulation of the test item due to the high Koc / Kow.  Studies conducted under flow-through conditions where test solutions are renewed continuously during the study help to maintain the test concentration during the duration of the study, these studies are preferable over those studies conducted under static conditions, when the test material has known test material properties which could lead to reduced test solution concentration due to adsorption processes for example. This approach is consistent with the approach taken by the USEPA evaluation of MGK 264.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

1.4 mg/L