Reaction mass of oxydiethylene dibenzoate and oxydipropyl dibenzoate

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09 September 1997 - 13 September 1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted in accordance with EC, OECD, and US EPA test guidelines, and in compliance with GLP.

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Deviations:

no

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OTS 797.1400 (Fish Acute Toxicity Test)

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

Lot #: 56625030
Expiry date: 19 December 1997
Composition of sample:
Diethylene glycol dibenzoate, CAS # 120-55-8, 89.9%
Diethlene glycol monobenzoate, CAS # 20587-61-5, 5.85%
Dipropylene glycol dibenzoate, CAS #, 27138-31-44, 1.5%
Ethylene glycol dibenzoate, CAS # 94-49-5, 0.19%
Triethylene glycol dibenzoate, CAS # 120-56-9, 0.15%
Unidentified component, CAS # unknown, 0.54%
Unidentified component, CAS # unknown, 0.10%
Unidentified component, CAS # unknown, 0.10%

Analytical monitoring:

yes

Details on sampling:

- Nominal Loading Rate Concentrations: 0 (control), 0.06, 0.3, 1.5, 7.5, and 37.5 mg/L.
- Sampling method: Duplicate samples (100 mL) of fresh and expired media were taken from the control and each test concentration daily. Fresh samples were taken at 0, 24, 48, and 72 hours and were obtained from the pooled solutions. Expired samples were taken at 24, 48, 72, and 96 hours; duplicate samples of expired media were taken from each replicate vessel.
- Sample storage conditions before analysis: Not specified.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: The method of preparation was selected following advice given in ECETOC 1996, Monograph No. 26 as the test material has poor water solubility.
Test material was added by accurate pipette to duplicate 20L glass vessels of dilution water and stirred for at least 18 hours. Once stirred, the solutions were left to stand for one hour prior to removing the water accomodated fraction by peristaltic pump. Approximately 18 L of the solution was removed from a mid water position, and the duplicates pooled to give the exposure solution.

- Evidence of undissolved material (e.g. precipitate, surface film, etc):
When preparing the exposure medium after the WAF had been stirred and left to stand, it was possible to see globules of test material at the bottom ofthe vessels. The compound was also visible as a film on the surface of the water. In exposure vessels expired solutions were noted to be cloudy after 24 hours.

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: Rainbow Trout
- Source: Parkwood Trout Farm, Kent, UK
- Age at study initiation (mean and range, SD): Not reported
- Length at study initiation (length definition, mean, range and SD): Not specified. Control animals were 4.4 cm (mean of control tank 1), S.D. 0.3 and 4.5 cm (mean of control tank 2), S.D. 0.6 at the end of the exposure period.
- Weight at study initiation (mean and range, SD): Not specified. Control animals were 1.0g (mean of control tank 1), S.D. 0.2 and 1.1g (mean of control tank 2), S.D. 0.5 at the end of the exposure period.
- Feeding during test
- No feeding during the 24 hour period before the start of the exposure period, or during the exposure period.

ACCLIMATION
- Acclimation period: 34 days (from 06 August - 09 September)
- Acclimation conditions (same as test or not): Yes
- Type and amount of food: Fed daily to repletion with commercial fry pellets.
- Feeding frequency: Daily.
- Health during acclimation (any mortality observed): Fewer that 1% died in the 7 days before the start of the test.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

Between 65 and 70 mgCaCO3/L.

Test temperature:

15±1°C.

pH:

Fresh solutions: 7.4 - 7.6
Expired solutions: 7.4 - 7.6

Dissolved oxygen:

Fresh solutions: 8.7 - 9.4 mgO2/L
Expired solutions: 9.2 - 9.5 mgO2/L

Nominal and measured concentrations:

Nominal Loading Rate Concentrations: 0 (control) 0.06, 0.3, 1.5, 7.5, and 37.5 mg/L
Mean Measured Diethylene Glycol Dibenzoate Concentrations: 0 (control), 0.047, 0.144, 0.491, 2.176, and 15.35 mg/L:
Measured concentrations for diethylene glycol dibenzoate and diethlene glycol monobenzoate are provided in Table 4.

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: glass aquaria, approximate dimensions 25 x 46 x 25 cm, filled to a depth of 17 cm (18 liters of dilution water or test solution).
- Aeration: Supplementary aeration was provided via narrow bore glass tubes.
- Renewal rate of test solution (frequency/flow rate): Semi-static renewal - media were renewed every 24 hours.
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates):2
- No. of vessels per control (replicates):2
- Biomass loading rate: 0.58 g bodyweight/litre

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Laboratory tap water was filtered, dechlorinated and softened by passage through an Elga water purification system.
- Total organic carbon: 1.1 mg/L
- Chlorine: 0.05 to 0.14 mg/L
- Alkalinity: 84 mg/L (as CaCO3)
- Ca/mg ratio: 53/3.5 (=15.1)
- Conductivity: 440 µS/cm
- Culture medium different from test medium: No

OTHER TEST CONDITIONS
- Adjustment of pH: None
- Photoperiod: 16h light : 8h dark

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
Mortality, defined as absence of respiratory movement, and absence of response to physical stimulation of the caudal peduncle. Observations were taken at 3, 6, 24, 48, 72, and 96 hours.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 5-fold
- Range finding study
- Test concentrations: A preliminary study was performed with initial loadings of test substance ranging from 0.1 to 100 mg/L.
- Results used to determine the conditions for the definitive study: Yes

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

1.12 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% CI 0.60, 1.64 mg/L

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

2.9 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% CL 2.2 - 3.8

Duration:

96 h

Dose descriptor:

NOELR

Effect conc.:

0.06 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Behavioural abnormalities: lethargy and loss of equilibrium seem at 0.3 mg/L and above, moribundity seen at 7.5 mg/L and above.
- Other biological observations: Increased pigmentationin the control and 0.3 mg/L levels.
- Mortality of control: One fish was found dead after 48 hours.
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values:
When preparing the exposure medium after the WAF had been stirred and left to stand, it was possible to see globules of test material at the bottom of the vessels. The compound was also visible as a film on the surface of the water. In exposure vessels expired solutions were noted to be cloudy after 24 hours.
The analytical method used to confirm the concentrations of test material in solution determined concentrations of diethylene glycol monobenzoate (DEGMB) as well as diethylene glycol dibenzoate (DEGDB); the concentration of DEGDB was found to be lower after 24 hours in expired samples than in the freshly prepared samples whereas the concentrations of DEGMB had increased. This observation suggests that the monobenzoate was a degradation product of the dibenzoate.

Reported statistics and error estimates:

96 hour LC50 = 1.12 mg/L (95% CI 0.60, 1.64 mg/L) based upon mean measured diethlyene glycol dibenzoate concentrations. The original study report provided LL50 and NOELR results based upon nominal loading rates only. The key result LC50 was calculated post study based upon the mean measured diethylene glycol dibenzoate concentrations reported in the study. The LC50 was calculated using the Probit Model within the TOSTAT version 3.4 statistical program.

Validity criteria fulfilled:

yes

Remarks:

Environmental parameters remained within acceptable limits throughout the duration of the study.

Conclusions:

The 96 h LC50 value for DEGDB with rainbow trout is 1.12 mg/L (95% CI 0.60, 1.64 mg/L) based upon the mean measured concentrations of diethylene glycol dibenzoate.

Executive summary:

A study was conducted to determine the acute toxicity of the test material DEGDB to Rainbow Trout (Oncorhynchus mykiss). The study was conducted according to EC, OECD, and US EPA test guidelines, and in compliance with GLP. Groups of 20 fish (duplicate groups of ten fish per loading level) were exposed to nominal loading rates of 0.06, 0.3, 1.5, 7.5, and 37.5 mg/L for 96 hours, in a semi-static test. The original test reported the 96 h LL50 value for DEGDB with rainbow trout was 2.9 mg/L and the no-observed effect loading rate (NOELR) for DEGDB with rainbow trout was 0.06 mg/L. The study did include analytical measurements of the exposure solutions for both the parent substance diethylene glycol dibenzoate (DEGDB) and the breakdown product diethylene glycol monobenzoate. Subsequent to the study the mean measured diethlene glycol dibenzoate exposure concentrations were calculated as 0.047, 0.144, 0.491, 2.176, and 15.35 mg/L based upon the study report data. The 96 hour LC50 was determined to be 1.12 mg/L (95% CI 0.60, 1.64 mg/L) based upon the mean measured diethlene glycol dibenzoate exposure concentrations.

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

26 October 1999 - 30 October 1999

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted in accordance with EC, OECD, and US EPA test guidelines, and in compliance with GLP.

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Deviations:

yes

Remarks:

Not thought to have effected the integrity of the study.

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

yes

Remarks:

Not thought to have effected the integrity of the study.

Qualifier:

according to guideline

Guideline:

EPA OTS 797.1400 (Fish Acute Toxicity Test)

Deviations:

yes

Remarks:

Not thought to have effected the integrity of the study.

Principles of method if other than guideline:

Protocol deviations
- The fish were not fed for 24 hours before the start of the test, not 48 hours as specified by the protocol. However, the test vessels were inspected regularly each day for fecal material and this was removed when found. This deviation is not considered to have increased the microbial loading as measured levels of the test substance were maintained between 88 to 104% of their nominal values after 24 hours, when the majority of faecal material was produced.
- Microbial counts were performed at 24, 48 and 72 hours in addition to the 0 and 96 hour counts specified in the protocol, at the request of the sponsor.
- Microbial counts were not made for the control vessels, as this was not considered necessary. Counts from the solvent control and test levels only were deemed sufficient for comparative purposes.
- The flow rate of the diluent was reduced from 280 mL/min as stated in the protocol to 260 mL/min during the definitive test in order to maintain the ratio of solvent to diluent.
- The above deviations were not thought to have affected the integrity of the study.

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0 (control), 0.25, 0.55, 1.2, 2.7, and 5.8 mg/L.
- Sampling method: Samples (50 mL) of media were taken from the control and each test concentration daily for chemical analysis. Samples (10 mL) were also taken for determination of bacterial count Samples were taken at 0, 24, 48, 72 and 96 hours.
- Sample storage conditions before analysis: Not specified.

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Dynamic exposure conditions were used. Dipropylene glycol dibenzoate has poor water solubility and had been shown to rapidly degrade in other aquatic studies. The test substance was dissolved in dimethyl formamide to give a series of stock solutions at 2.5, 5.5, 12, 27 and 58 mg/L DPGDB. The stock solutions were dispensed automatically by dosing apparatus into the flowing test water.
- Eluate: The water used for the study was laboratory tap water filtered dechlorinated and softened by passage through an Elga water purification system
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): dimethyl formamide (DMF)
- Concentration of vehicle in test medium (stock solution and final test solution(s) including control(s)): 100 µL DMF/L
- Evidence of undissolved material (e.g. precipitate, surface film, etc): None

The test and mixing vessels were changed daily to maintain the cleanliness of the test system. The vessels were removed every day and replaced with clean dry tanks. Used tanks were washed with detergent, rinsed with acetone, then acid-washed and rinsed with water. The tanks were allowed to dry before re-use.

Test organisms (species):

Pimephales promelas

Details on test organisms:

TEST ORGANISM
- Common name: Fathead minnow
- Strain: Pimephales promelas
- Source: Aquatic Research Organisms Hampton New Hampshire USA
- Length at study initiation (length definition, mean, range and SD): At this time the fish were measured from the head to the caudle peduncle (standard length). The mean standard length of the sample was 2.7 cm (SD = 0.14 cm) The EC guideline specifies a lower size limit of 3.0 cm total length. In order to confirm that the stock fish were within this limit a second sample of fish from the same stock tank were measured after the definitive test. The mean standard length of this sub sample was 25 cm (SD=0.14 cm) and the mean total length head to tip of caudal fin was 31 cm (SD 0.16 cm). As the mean standard lengths of both batches of fish from the stock tank were similar it can be assumed that the total length of the fish used for the definitive test was greater than 3 cm.
- Weight at study initiation (mean and range, SD): Mean weight was 0.28 g (SD = 0.04 g)
- Feeding during test: No feeding during the 24 hour period before the start of the exposure period, or during the exposure period.

ACCLIMATION
- Acclimation period: 14 days
- Acclimation conditions (same as test or not):Temperature remained within the range 17 to 18°C in the 14 day period immediately before the study and at 18°C in the seven day period before the study.
- Type and amount of food: The fish were fed daily to repletion with Tetramin flakes.
- Feeding frequency: Food was not given during the 24 hour period immediately before exposure or during the exposure period itself
- Health during acclimation (any mortality observed): no

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Post exposure observation period:

None

Hardness:

During acclimation , hardness, measured as mg/L CaCO3 varied between 127 and 162. During the exposure period it varied between 133 and 164 mg/L CaCO3.

Test temperature:

23 to 24°C

pH:

7.9 to 8.1

Dissolved oxygen:

6.8 to 7.9 mg O2 / L

Salinity:

Not applicable

Nominal and measured concentrations:

Nominal concentrations; 0 (control), 0.25, 0.55, 1.2, 2.7 and 5.8 mg/L
Mean measured concentrations; 0.25, 0.53, 1.2, 2.6 and 4.9 mg/L
Expressed as % of nominal concentration: 100, 96, 100, 97, 84%

Due to the nature of the study design the concentrations of dipropylene glycol monobenzoate were not determined in this study

Details on test conditions:

TEST SYSTEM
- Test vessel: glass aquarium
- Type: open 20 litre mixing vessel
- Aeration: No
- Type of flow-through (e.g. peristaltic or proportional diluter): peristaltic pump.
- Renewal rate of test solution (frequency/flow rate): 260 mL/min
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- No. of vessels per vehicle control (replicates): 2
- Biomass loading rate:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Laboratory tap water filtered, dechlorinated and softened by passage through an Elga water purification system.
- Total organic carbon: C 1.1 mg/L
- Chlorine: <0.1 mg/L
- Alkalinity: as CaCO3, 98 mg/L
- Ca/mg ratio: 53/3.2 = 10.19
- Conductivity: 420 µS/cm

OTHER TEST CONDITIONS
- Photoperiod: 16 hours light / 8 hours dark

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : The criteria of death employed in this study were i) absence of respiratory movement and ii) absence of response to physical stimulation of the caudal peduncle. In addition to observations on mortality at 0.25, 3, 6, 24, 48, 72 and 96 hours subjective assessments were made on the incidence and type of sub lethal effects compared with control fish.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2.2-fold
- Range finding study
- Test concentrations: A preliminary study was performed with initial concentrations of test substance of 4.0 and 6.0 mg/L.
- Results used to determine the conditions for the definitive study: Yes

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

3.7 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% CL 3.2 - 4.3

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

1.2 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Behavioural abnormalities: None reported
- Other biological observations: Inflammation of the gills and loss of equilibrium were noticed after two days exposure at 4.9 mg/L, and after 4 days at 2.6 mg/L. Fish exposed to 4.9 mg/L showed a curved body shape caused by muscle rigidity, and lesions were visible one one fish at this level
- Mortality of control: None

Validity criteria fulfilled:

yes

Conclusions:

The 96 hour LC50 value for dipropyleneglycol dibenzoate with fathead minnow was 3.7 mg/L. The "no-observed effect concentration" (NOEC) was 1.2 mg/L
Measured concentrations of DPGDB correlated well with the nominal concentrations.

Executive summary:

A study was performed to determine the acute toxicity of the test material DPGDB to fathead minnows. The study was conducted according to EC, OECD, and US EPA test guidelines, and in compliance with GLP.

Groups of 20 fish (duplicate groups of ten fish per group) were exposed to nominal dose levels 0, (control), 0.25, 0.55, 1.2, 2.7, and 5.8 mg/L DPGDB over a period of 96 hours using a dynamic exposure system.

The 96 h LC50 value for DPGDB with fathead minnow was 3.7 mg/L based upon mean measured concentrations. The "no-observed effect concentration" (NOEC) for DPGDB with fathead minnow was 1.2 mg/L.

Description of key information

DEGDB:

Short-term toxicity to fish:

- Key study, reliability 1, OECD 203, rainbow trout - LC50 (96h) = 1.12 mg/L

- Supporting study, reliability 1, OECD 203, fathead minnow - LC50 (96h) = 1.85 mg/L

DPGDB:

Short-term toxicity to fish:

- Key study, reliability 1, OECD 203, fathead minnow - LC50 (96h) = 3.7 mg/L

- Supporting study, reliability 1, OECD 203, rainbow trout - LC50 (96h) > 3.0 mg/L

- Supporting study, reliability 2, QSAR calculation (EPIWIN-ECOSAR v1.00) - LC50 = 3.94 mg/L

- Disregarded study, reliability 3, OECD 203, fathead minnow - LL50 > 100 mg/L, NOELR = 0.8 mg/L, poor correlation of nominal levels with measured concentrations

- Disregarded study, reliability 3, OECD 203, rainbow trout - LL50 = 0.25 mg/L, NOELR = 0.06 mg/L, poor correlation of nominal levels with measured concentrations

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Dose descriptor:

LC50

Effect concentration:

1.12 mg/L

Marine water fish

Marine water fish

Dose descriptor:

LC50

Effect concentration:

1.12 mg/L

Additional information

This substance is a reaction mass of dipropylene glycol dibenzoate (DPGDB) and diethylene glycol dibenzoate (DEGDB). No testing has been performed on the reaction mass itself but data are available for DPGDB and DEGDB individually. All the studies were performed according to international test guidelines and in compliance with GLP. They are presented below, by component. 

The acute toxicity studies were carried out on freshwater fish species (rainbow trout and fathead minnow).  The components of the reaction mass have quantitatively similar aquatic toxicity properties with all components exhibiting fish 96 hour LC50 values within the same order of magnitude. The most sensitive endpoint result obtained for these components has been used as the key value to evaluate short-term hazard and is highlighted below. A long-term toxicity to fish study of the reaction mass is in progress.

 

The key value used in the chemical safety assessment is:

Short-term aquatic toxicity LC50 = 1.12 mg/L (DEGDB, rainbow trout)