2,4-dinitroanisole

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

March 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: USEPA 821-R-02-013

Version / remarks:

4th

Deviations:

not specified

GLP compliance:

not specified

Remarks:

Study conducted to a high scientific standard and findings presented in a comprehensive test report.

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Holston Army Ammunition Plant, Lot #BAE10H281-008
- Expiration date of the lot/batch: not specified
- Purity test date: not specified

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: 4°C until analysis

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: dissolution in freswater during 48h using a magnetic stirring bar

Analytical monitoring:

yes

Details on sampling:

- Concentrations (%): 100, 50, 25, 12.5 and 6.25
- Nominal concentrations ranged from 0 - 25 mg/L
- Sample storage conditions before analysis: 4°C

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Dissolution during 48h in freshwater using a magnetic stirring bar
- Eluate: Freshwater
- Controls: Negative control
- Concentration of vehicle in test medium: analytical measurement
- Evidence of undissolved material: no undissolved material

Test organisms (species):

Pimephales promelas

Details on test organisms:

TEST ORGANISM
- Common name: Pimephales promelas
- Strain: not specified
- Source: Aquatic Biosystems (Fort Collins, Colorado)
- Age at study initiation: not specified
- Length at study initiation: not specified
- Weight at study initiation: not specified
- Method of breeding: not specified
- Maintenance of the brood fish: not specified

ACCLIMATION
- Acclimation period: not specified
- Acclimation conditions: not specified
- Type and amount of food during acclimation: not specified
- Feeding frequency during acclimation: not specified
- Health during acclimation (any mortality observed): not specified

FEEDING DURING TEST
- Food type: Artemia sp. nauplii
- Amount: ration
- Frequency: 2 times per day

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Hardness:

80 mg/L CaCO3

Test temperature:

23.4 - 24.5

pH:

6.85 - 7.99

Dissolved oxygen:

7.3 - 8.0 mg/L

Salinity:

not specified

Conductivity:

275 - 318 μS/cm

Nominal and measured concentrations:

- Nominal Concentrations (mg/L): 0, 6, 12, 25, 50 and 100
- Measured Concentrations (mg/L): 0, 7.1, 13, 28, 74 and 120

Details on test conditions:

TEST SYSTEM
- Test vessel: not specified
- Material, size, headspace, fill volume: not specified
- Aeration: not specified
- Renewal rate of test solution (frequency/flow rate): 80% per day
- No. of organisms per vessel: not specified
- No. of vessels per concentration (replicates): 5
- No. of vessels per control (replicates): 5

OTHER TEST CONDITIONS
- Adjustment of pH: not specified
- Photoperiod: not specified
- Light intensity: not specified

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Test concentrations mg/L): 0, 6.25, 12.5, 25, 50, 100

Reference substance (positive control):

yes

Remarks:

KCl

Key result

Duration:

48 h

Dose descriptor:

LC50

Effect conc.:

>= 33 - <= 41 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

13 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Key result

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

28 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

none

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- LC50: 48h-LC50: 0.78 mg/L

Reported statistics and error estimates:

Survival data were arcsine square-root transformed prior to statistical analysis. Data normality (Kolmogorov–Smirnov test), homogeneity (Levene’s test), and treatment differences were compared to the reference (one way ANOVA and Dunn’s or Dunnett’s Methods) and statistical significance was determined at α = 0.05 using SigmaStat software (SPSS, Chicago, Illinois, USA). When normality was not achieved, the nonparametric Kruskal–Wallis one-way ANOVA on ranks was applied. The lethal median concentration producing 50% mortality (LC50) and inhibition concentrations (IC50, IC25) for sublethal endpoints were determined by the trimmed Spearman–Karber method (Toxcalc®, Version 5.0, Tidepool Scientific Software, McKinleyville, California). The LC50 and IC25 values are presented with 95% confidence limits in parentheses. No observableeffect concentrations (NOEC) and lowest observable effect concentrations (LOEC) were also calculated using ToxCalc v5.0 and confirmed by one-way ANOVA and Dunn’s or Dunnett’s Methods. Maximum allowable concentrations(MATC), also known as the chronic value (ChV), were calculated as the geometric mean of the NOEC and LOEC. The acute-to-chronic ratio (ACR) was calculated as the 48-hr LC50 divided by the MATC.

Sublethal observations / clinical signs:

Nominal (mg/L)	Measured test initiation (mg/L)	Measured test termination (mg/L)	Mean survival
0	<0.01	<0.01	100±0
6	7.1	6.6	100±0
12	13	14	100±0
25	28	28	75±6
50	74	74	0±0
100	120	120	0±0

Validity criteria fulfilled:

yes

Conclusions:

The 48-hr acute toxicity of DNAN was found to 33 mg/L for the Pimephales promelas.

Executive summary:

Acute and chronic aquatic toxicity of DNAN was investigated using standard fish (Pimephales promelas) models. Chemical analysis of test water indicated that DNAN concentrations were relatively stable during the bioassays. Acute toxicity presented a 48-hr lethal median concentrations (LC50) of 33 mg/L DNAN. Chronic toxicity tests indicated that fish survival (7-day LC50 = 10 mg/L) was significantly more sensitive to DNAN relative to the invertebrate (no significant impact on survival at 24 mg/L). However, the reproduction endpoint for the invertebrate was significantly more sensitive to DNAN than survival. When assessing the most sensitive chronic endpoints, the two test species indicated similar chronic toxicity, with lowest observable adverse impacts ranging from 10 to 12 mg/L DNAN and median effects on sublethal endpoints (growth, reproduction) ranging from 11 to 15 mg/L DNAN. Chronic no-effect concentrations ranged from approximately 6 to 8 mg/L DNAN.

Description of key information

The 48-hr acute toxicity of DNAN (LC50) was found to be 33 mg/L for the Pimephales promelas.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

33 mg/L

Additional information

A determination of the acute toxicity to fish of DNAN was conducted (U.S. Army Engineer Research and Development Center, ERDC/EL TR-13-2, 2013). Tests were conducted in accordance with standard guidance (U.S. Environmental Protection Agency (USEPA) 2002a, EPA-821-R-02-012).

Groups of Pimephales promelas were initially exposed to a nominal concentration range of 0 - 100 mg/L DNAN in MHRW, using a 90% dilution series (100, 10, 1, 0.1, 0.01%). Based on range-finding results, definitive acute toxicity tests were conducted in the nominal concentration range of 0 – 100 mg/L DNAN, using a 50% dilution series (100, 50, 25, 13, 6 and 0 %). The measurement endpoint was survival after 48-hr exposure.

It was concluded that the 48-hr LC50 for Pimephales promelas ranged from 33 – 41 mg/L.