3,5,5-trimethylhexanoic acid

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:

Study performed from 06 Nov 1995 to 10 Nov 1995 (in-life test period).

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

yes

Remarks:

fish size smaller

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Samples were collected at each test concentration of new test solution at each renewal (day 0, 1 , 2, 3) and of the old test solution on day 1 and at termination.
- Sampling method: Samples were collected with no headspace in volatile organic analysis vials
- Sample storage conditions before analysis: refrigerated

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Individual water accommodated fractions (WAF) were prepared by adding the appropriate amount of the test substance to 19.5 L of dilution water in a 20-L glass carboy. The mixtures were stirred for ca. 24 h on a magnetic stirrer (=< 10 % vortex) at 22 +-2 °C. Each vessel was sealed with a neoprene stopper and completely covered with an opaque covering. After stirring the mixtures were adjusted to the pH of the dilution water and cooled to test temperature.
The WAF was removed by glass tube siphon from the bottom of the vessel.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: rainbow trout
- Source: Thomas Fish Company, Anderson CA, USA
- Age at study initiation (mean and range, SD): ca. 5 weeks
- Length at study termination (length definition, mean, range and SD): 3.3 cm (sd = 0.2) (fish smaller than required by protocol in order to maintain sufficient oxygen level in closed test vessel)
- Weight at study termination (mean and range, SD): 0.269 g (sd = 0.038)
- Feeding during test: none


ACCLIMATION
- Acclimation period: at least 7 d
- Acclimation conditions (same as test or not): yes
- Type and amount of food: Salmon Starter/Crumble (Zeigler Bros., Inc., Gardners, PA, USA); Tetramin (That Fish Place, Lancaster, PA, USA)
- Feeding frequency: no data
- Health during acclimation (any mortality observed): no mortality occurred during holding period


QUARANTINE
- Duration: at least 12 d
- Health/mortality: not treated for disease or parasites; no mortality occurred during holding period

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

15 °C (sd = 0.04)

pH:

7.0-8.3

Dissolved oxygen:

>= 6.0 mg/L

Nominal and measured concentrations:

nominal/measured (mg/L): 0/n.d. (control), 62/44.5-79.8, 96/53.5-182, 148/88.5-167, 228/230-267 and 350/265

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: 4-L glass aspirator bottles, no headspace, fill volume: 4.5 L; sealed with foil covered neoprene stoppers
- Aeration: none
- Renewal rate of test solution (frequency/flow rate): daily (80 %)
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3
- Biomass loading rate: 0.30 g/L


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: groundwater, carbon-filtered, reverse osmosis dialyzed
- Alkalinity: 55/61 mg/L as CaCO3
- Hardness: 174/160 mg/L as CaCO3
- pH 7.5/7.9
- Disssolved oxygen: 9.0/10.2
- TOC: 0.458/0.739 ppm (sample date: 12 Oct 1995/20 Nov 1995)
- Conductivity: 340/350 µmhos
- Intervals of water quality measurement: data from two analyses (sample date: 03 Nov and 09 Nov 1995); monitored semi-annuallyfor priority pollutants, un-ionized ammonia, total suspended solids, herbicides, residual chlorine and annually for bacterial properties


OTHER TEST CONDITIONS
- Adjustment of pH: yes
- Photoperiod: 19 h light : 5 h dark
- Light intensity: 528-538 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- mortality, abnormal behaviour, appearance at 3, 24, 48, 72 and 96 h
- water quality measurements (pH, dissolved oxygen, temperature): performed on new test solutions on day 0, 1, 2 and 3; performed on old test solutions on day 1, 2, 3 and at termination

TEST CONCENTRATIONS
- Spacing factor for test concentrations:
- Justification for using less concentrations than requested by guideline:
- Range finding study: 48-h exposure
- Test concentrations: 0 (control), 0.25, 2.5 and 25 mg/L
- Results used to determine the conditions for the definitive study: no data

Reference substance (positive control):

not specified

Duration:

96 h

Dose descriptor:

LC0

Effect conc.:

108 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

122 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95 % CI: 116-129 mg/L

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

123 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95 % CI: 116-130 mg/L

Duration:

96 h

Dose descriptor:

LC100

Effect conc.:

248 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

act. ingr.

Basis for effect:

mortality (fish)

Reported statistics and error estimates:

96-h LC50 and LL 50 were calculated using the trimmed Spearman-Kärber method.

Sublethal observations / clinical signs:

Mortalities:

Test conc.	Mortalities (n) after x days
measured (mg/L)	3 h	1	2	3	4
0	0	0	0	0	0
67.2	0	0	0	0	0
108	0	0	0	0	0
131	0	0	0	0	14
248	0	15	15	15	15
265	0	15	15	15	15

Mortality increased upruptly in a narrow concentration range on day 4 of exposure (steep concentration-response curve).

Observations:

Test conc.	Observations after x days
measured (mg/L)	3 h	1	2	3	4
0
67.2
108					DP(2)
131				DP(1)	S(1)
248	L(15), DP(15)
265	L(15), DP(15)

Legend:

L	lethargic (number of affected fish)
DP	dark pigmentation (number of affected fish)
S	swimming on side (number of affected fish)

Validity criteria fulfilled:

yes

Conclusions:

Based on mean measured concentrations, the 96-h LC50 was 122 mg/L.

Executive summary:

3,5,5 -trimethylhexanoic acid was tested on rainbow trout (Oncorhynchus mykiss) in a 96 -h semi-static acute study according to OECD TG 203. Three replicates were set up per concentration and control. Five fish were introduced in each replicate. The test concentrations were analytically verified by GC-FID at each renewal (new test solution: day 0, 1, 2, 3; old test solution: day 1 and at termination). Nominal (and mean measured for each replicate) concentrations were as follows: 0 (0) mg/L, 62 (67.2) mg/L, 96 (108) mg/L, 148 (131) mg/L, 228 (248) mg/L, and 350 (265) mg/L. No signs of distress were observed in the two lowest test concentrations (LC0 = 108 mg/L). Mortalities occurred in the three highest test concentrations. Based on mean measured concentrations, the 96-h LC50 was 122 mg/L.

Description of key information

Rainbow trout (O. mykiss): 96-h LC50 = 122 mg/L (OECD 203, Exxon Chemical Europe, 1996)

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

122 mg/L

Additional information

Reliable results on acute fish toxicity are available for two fish species (cold and warm water):

Endpoint	Organism group	Species	Value [mg/L]	Reliability	Source
96-h LC50	Fish	Rainbow trout (Oncorhynchus mykiss)	122	1 (key study)	Davi (1996)
96-h LC50	Fish	Pumpkinseed (Lepomis gibbosus)	190	2 (supp. study)	Hoechst AG (1979)
96-h LC50	Fish	Rainbow trout (Oncorhynchus mykiss)	160	2	Markert & Weigand (1979)

All available information indicates that acute toxicity to fish is beyond 100 mg/L. Hence 3,5,5 -trimethylhexanoic acid can be regarded as not acutely toxic to fish.

The study with the cold water species O. mykiss was selected as key study since it was the most sensitive. In contrast to the the other two reliable studies the test concentrations were verified by chemical analysis.

In this study, 3,5,5 -trimethylhexanoic acid was tested on rainbow trout (Oncorhynchus mykiss) in a 96 -h semi-static acute study according to OECD TG 203. Three replicates were set up per concentration and control. Five fish were introduced in each replicate. The test concentrations were analytically verified by GC-FID at each renewal (new test solution: day 0, 1, 2, 3; old test solution: day 1 and at termination). Nominal (and mean measured for each replicate) concentrations were as follows: 0 (0) mg/L, 62 (67.2) mg/L, 96 (108) mg/L, 148 (131) mg/L, 228 (248) mg/L, and 350 (265) mg/L. pH values were in the neutral range for all concentration levels applied. No signs of distress were observed in the two lowest test concentrations (LC0 = 108 mg/L). Mortalities occurred in the three highest test concentrations. Based on mean measured concentrations, the 96-h LC50 was 122 mg/L.