Vanillin

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:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study seems to be well conducted.

GLP compliance:

not specified

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

Pimephales promelas

Details on test organisms:

TEST ORGANISM
- Common name: Pimephales promelas
- Age at study initiation (mean and range, SD): 4 to 8 weeks
- Length at study initiation (length definition, mean, range and SD): 11 to 31 mm
- Feeding during test: no data

ACCLIMATION
- Acclimation period: 48 hours
- Acclimation conditions (same as test or not): in flowing water

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test temperature:

18 - 22°C

Dissolved oxygen:

< 4 mg/L

Details on test conditions:

Duplicate tests with 10 fishes at each concentration, in 3 litres cylindrical glass battery jars containing 2 litres of reconstituted water.

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

112 - 121 mg/L

Sublethal observations / clinical signs:

 
           test 1 (mg/l)    test 2 (mg/l) 
LC50-96H=     121               112 
LC50-72H=     121               112  
LC50-48H=     123               116 
LC50-24H=     131               125

(test 1 and 2: acclimated in Lake Superior water)
(test 3: acclimated in reconstituted water)

          test 1 (mg/l)   test 2 (mg/l)   test 3 (mg/l) 
LC50-96H=     88              121             116 
LC50-72H=     88              121             116 
LC50-48H=     97              121             116 
LC50-24H=    100              127             125 
(test 1 and 2: acclimated in Lake Superior water) 
(test 3: acclimated in reconstituted water)

Reference 2

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: The test conditions meet the standardized guideline requirements.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

not specified

GLP compliance:

no

Analytical monitoring:

yes

Details on sampling:

- Concentrations: all tested concentrations were analyzed
- Sampling method: all test exposure chambers were sampled at approximately mid-depth at 0 and 96H. and one of each duplicate exposure chamber at 24, 48 and 72H.
- Sample storage conditions before analysis: all samples were analyzed immediately or adequately preserved for later analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Preparation of a 5g/l of vanillin solution by dilution in test medium.
- Controls: blank controls have been performed
- Eluate: no data
- Differential loading: no data
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): not used
- Evidence of undissolved material: no data

Test organisms (species):

Pimephales promelas

Details on test organisms:

- Common name: Fathead minnow
- Source: From US EPA Environmental Research Laboratory- Duluth or University of Winsconsin Superior Campus.
- Age at study initiation: 29 days
- Weight at study initiation (mean +/- SD): 0.131 +/- 0.032 g
- Length at study initiation (mean +/- SD): From head to the beginning of the tail: 20 +/- 1.7 mm
- Method of breeding: adults were held at 25°C in flowing water with a 16Hr light-controlled photoperiod abd fed with frozen adult brine shrimp (Artemia sp.). They were provided with asbestos pipe as spawning substrate. Eggs were spawned and fertilized on the undersides of these substrates. The embryos were tended by the male parent until eye-up at which time the substrates with embryos attached were removed to another 25°C bath where hatching occurred. Fry of similar (within 24Hr) age were transferred to flow-through rearing tanks at 25°C with a 16 Hr light photoperiod. Fry were freshly hatched brine shrimp nauplii to excess three times daily until 24Hr before a test (twice on most week-end days) and a dry flake ration (Tetramin) during their first week. Fish of approximately 30 days old were used in the toxicity tests.
- Feeding during test: not fed during tests

ACCLIMATION
- Acclimation period: no

QUARANTINE: no data

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

51 mg/L

Test temperature:

23.9 +/- 0.73°C (SD)

pH:

7.16 +/- 0.32 (SD)

Dissolved oxygen:

7.2 +/- 0.93 mg/L (SD)

Nominal and measured concentrations:

Nominal: 0, 34, 56, 94, 157 and 261 mg/L
Mean measured : <0.2, 19.3, 31.6, 55.2, 90, 174 mg/l (corrected for recovery)

Details on test conditions:

TEST SYSTEM
- Test vessel: 6.3 L glass open tank without aeration
- Type of flow-through (e.g. peristaltic or proportional diluter):cycling proportional diluters
- Renewal rate of test solution (frequency/flow rate): 5.7 volumes per day
- No. of organisms per vessel: 25
- No. of vessels per concentration: 2
- No. of vessels per control: 2
- Biomass loading rate: 0.52 g/L

WATER PARAMETERS
- Source/preparation of dilution water: Lake Superior water
- Total organic carbon: no data
- Particulate matter: total non-filterable residue: <1 mg/l
- Metals: Al: 1-26, Cd: <0.1, Cr: 2-20, Co: <0.5; Cu: 0.3-3.2; Fe: 2-83; Pb: 7-20; Mn: 0.2-11.5; Ni: <0.5; Zn: 1.0-2.7 µg/l
- Pesticides: no data
- Chlorine: 1.2-1.3 mg/l
- Alkalinity: no data
- Ca/mg ratio: Ca: 13.0-14.7 mg/l; Mg: 2.9-3.6 mg/l
- Conductivity: 78-86 µmhos/cm
- Culture medium different from test medium: no
- Intervals of water quality measurement:hardness, alkalinity, pH, at least once during each test; water temperature: daily in each exposure chamber; dissolved oxygen: in high, medium, low and control exposure chambers at least three times (0,24, 96H) during a test if surviving fish existed in that chamber.

OTHER TEST CONDITIONS
- Adjustment of pH: no data
- Photoperiod: no data
- Light intensity: no data

EFFECT PARAMETERS MEASURED: during exposure to toxicants the fish were routinely observed for behavioural responses (effects) and deaths. Death was defined as the cessation of opercular movements and the inability to respond when prodded. Dead fish were removed and recorded at 3, 6, 12, 24, 48, 72 and 96h.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.65
- Nominal test concentrations: 0, 34, 56, 94, 157, 261 mg/l

Reference substance (positive control):

not specified

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

57 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: (53-61.1 mg/L)

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

57 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

behaviour

Remarks on result:

other: (53.0-61.3 mg/L)

Details on results:

Results were calculated on the basis of the mean measured exposure concentrations of toxicants for the duration of the test.
Initial measured values were 50 to 60% lower than the nominal concentrations, but were stable during the 96H of exposure.

- Behavioural abnormalities: Affected fish stopped schooling, became hypoactive, swam at the surface, and lost equilibrium prior to death.
- Mortality of control: None
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No details

Reported statistics and error estimates:

Duplicates were combined resulting in a single LC50 and EC50 estimate. Estimation of the E/LC50 was made using the "trimmed Spearman-Karber" method. Confidence limits (95%) were calculated.

Sublethal observations / clinical signs:

Cumulative mortality (initial number of fish = 25 per replicate; 2  replicates)
_______________________________________________________
         Control      A        B         C          D          E       LC50 (CL 95%)
************************************************************************** *****
24          0           0        0           0         12         50      69.3 (91.3-101.2)
48          0           0        0         10         48         50      62.1 (56.9-68.6)
72          0           0        0         20         50         50      57.0 (53.0-61.3)
96          0           0        0         20          50        50      57.0 (53.0-61.3)
_______________________________________________________

Cumulative behaviour effects (initial number of fish = 25 per replicate;  2 replicates)
_______________________________________________________
         Control      A        B         C          D          E       EC50 (CL 95%)
************************************************************************** *****
24          0            0        0           0         12        50      94.5 (88.8-101.0)
48          0            0        0         10         48        50      62.1 (56.9-68.6)
72          0            0        0         20         50        50      57.0 (53.0-61.3)
96          0            0        0         20         50        50      57.0 (53.0-61.3)
_______________________________________________________

Validity criteria fulfilled:

yes

Remarks:

No death within controls, O2 content throughout the test: 7.2+/-0.93 mg/l

Conclusions:

Harmful for Pimephales promelas.

Executive summary:

In an acute toxicity study (Brooke, 1984), Pimephales promelas were exposed to vanillin at nominal concentrations of 0 to 262 mg/L, under flow-through conditions.

The LC50 (96h) was 57 mg/L (CI: 53 - 61.3 mg/L), based on mean measured concentrations for the duration of the test.

Vanillin is harmful for Pimephales promelas in the tested conditions .

This study is reliable with restrictions and satisfies the guideline requirements.

Reference 3

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: The test conditions meet the standardized guideline requirements.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

not specified

GLP compliance:

no

Analytical monitoring:

yes

Details on sampling:

- Concentrations: all tested concentrations were analyzed
- Sampling method: all test exposure chambers were sampled at approximately mid-depth at 0 and 96H. and one of each duplicate exposure chamber at 24, 48 and 72H.
- Sample storage conditions before analysis: all samples were analyzed immediately or adequately preserved for later analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Preparation of a 5g/l of vanillin solution by dilution in test medium.
- Controls: blank controls have been performed
- Eluate: no data
- Differential loading: no data
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): not used
- Evidence of undissolved material: no data

Test organisms (species):

Pimephales promelas

Details on test organisms:

TEST ORGANISM
- Common name: Fathead minnow
- Source: From US EPA Environmental Research Laboratory- Duluth or University of WinsconsinSuperior Campus.
- Age at study initiation: 31 days
- Weight at study initiation (mean +/- SD): 0.055 +/- 0.018 g
- Length at study initiation (mean +/- SD): From head to the beginning of the tail: 17.3 +/- 2.02 mm
- Method of breeding: adults were held at 25°C in flowing water with a 16Hr light-controlled photoperiod abd fed with frozen adult brine shrimp (Artemia sp.). They were provided with asbestos pipe as spawning substrate. Eggs were spawned and fertilized on the undersides of these substrates. The embryos were tended by the male parent until eye-up at which time the substrates with embryos attached were removed to another 25°C bath where hatching occurred. Fry of similar (within 24Hr) age were transferred to flow-through rearing tanks at 25°C with a 16 Hr light photoperiod. Fry were freshly hatched brine shrimp nauplii to excess three times daily until 24Hr before a test (twice on most week-end days) and a dry flake ration (Tetramin) during their first week. Fish of approximately 30 days old were used in the toxicity tests.
- Feeding during test: not fed during tests

ACCLIMATION
- Acclimation period: no

QUARANTINE: no data

Test type:

flow-through

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

53.5 +/- 1 mg/l (SD)

Test temperature:

24.7 +/- 0.72 (SD)

pH:

7.00 +/- 0.09 (SD)

Dissolved oxygen:

6.2 +/- 0.37 (SD)

Nominal and measured concentrations:

Nominal: 0, 23, 38, 64, 107, 178 mg/l
Mean measured conc, corrected for recovery: <0.2, 10.8, 19.1, 42.6, 64.75, 135 mg/l

Details on test conditions:

TEST SYSTEM
- Test vessel: 5.5 L glass open tank without aeration
- Type of flow-through (e.g. peristaltic or proportional diluter):cycling proportional diluters
- Renewal rate of test solution (frequency/flow rate): 13.1 volumes per day
- No. of organisms per vessel: 25
- No. of vessels per concentration: 2
- No. of vessels per control: 2
- Biomass loading rate: 0.2 g/L

WATER PARAMETERS
- Source/preparation of dilution water: Lake Superior water
- Total organic carbon: no data
- Particulate matter: total non-filterable residue: <1 mg/l
- Metals: Al: 1-26, Cd: <0.1, Cr: 2-20, Co: <0.5; Cu: 0.3-3.2; Fe: 2-83; Pb: 7-20; Mn: 0.2-11.5; Ni: <0.5; Zn: 1.0-2.7 µg/l
- Pesticides: no data
- Chlorine: 1.2-1.3 mg/l
- Alkalinity: no data
- Ca/mg ratio: Ca: 13.0-14.7 mg/l; Mg: 2.9-3.6 mg/l
- Conductivity: 78-86 µmhos/cm
- Culture medium different from test medium: no
- Intervals of water quality measurement:hardness, alkalinity, pH, at least once during each test; water temperature: daily in each exposure chamber; dissolved oxygen: in high, medium, low and control exposure chambers at least three times (0,24, 96H) during a test if surviving fish existed in that chamber.

OTHER TEST CONDITIONS
- Adjustment of pH: no data
- Photoperiod: no data
- Light intensity: no data

EFFECT PARAMETERS MEASURED: during exposure to toxicants the fish were routinely observed for behavioural responses (effects) and deaths. Death was defined as the cessation of opercular movements and the inability to respond when prodded. Dead fish were removed and recorded at 3, 6, 12, 24, 48, 72 and 96h.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.66
- Nominal test concentrations: 0, 23, 38, 64, 107, 178 mg/l

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

123 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: (104-146)

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

123 mg/L

Nominal / measured:

meas. (TWA)

Conc. based on:

test mat.

Basis for effect:

behaviour

Remarks on result:

other: (104-146)

Details on results:

Results were calculated on the basis of the mean measured exposure concentrations of toxicants for the duration of the test.
Initial measured values were 50 to 60% lower than the nominal concentrations, but were stable during the 96H of exposure.

- Behavioural abnormalities: Affected fish stopped lost equilibrium prior to death.
- Observations on body length and weight:no data
- Other biological observations:
- Mortality of control: None
- Any observations that might cause a difference between measured and nominal values: No details

Reported statistics and error estimates:

Duplicates were combined resulting in a single LC50 and EC50 estimate. Estimation of the E/LC50 was made using the "trimmed Spearman-Karber" method. Confidence limits (95%) were calculated.

Sublethal observations / clinical signs:

Cumulative mortality (initial number of fish = 25 per replicate; 2  replicates)
_______________________________________________________
         Control      A        B         C          D          E       LC50 (CL 95%)
************************************************************************** *****
24          0           0        0          0          0           6       177 (169-189)
48          0           0        0          0          0          12      153 (151-157)
72          0           0        0          0          1          27      123 (104-146)
96          0           0        0          0          1          27      123 (104-146)
_______________________________________________________

Cumulative behaviour effects (initial number of fish = 25 per replicate;  2 replicates)
_______________________________________________________
         Control      A        B         C          D          E       LC50 (CL 95%)
************************************************************************** *****
24          0           0        0          0          0           6       177 (169-189)
48          0           0        0          0          0          12      153 (151-157)
72          0           0        0          0          1          27      123 (104-146)
96          0           0        0          0          1          27      123 (104-146)
_______________________________________________________

Validity criteria fulfilled:

yes

Remarks:

No death within controls, O2 content throughout the test: 6.2+/-0.37 mg/l

Conclusions:

This test, even if conducted in strictly the same conditions as in Brooke et al. (1984), referenced as ST tox fish V2 1984BROO, gives higher LC50 (96h) : 123 mg/L, leading to consider Vanillin as not harmful to fish Pimephales promelas.
No analysis of this discrepancy was discussed by the authors.

Executive summary:

In an acute toxicity study (Brooke et al, 1984b), Pimephales promelas were exposed to vanillin at nominal concentrations of 0 to 178 mg/L, under flow-through conditions.

The LC50 (96h) was 123 mg/L (CI: 104 -146 mg/L), based on mean measured concentrations for the duration of the test.

In the conditions of this test, Vanillin is not harmful for Pimephales promelas .

This study is reliable with restrictions and satisfies the guideline requirements.

Reference 4

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

other: Not enough data for assessment

Principles of method if other than guideline:

no data

GLP compliance:

not specified

Analytical monitoring:

not specified

Test organisms (species):

Oryzias latipes

Details on test organisms:

The fish were taken from a stream near the laboratory of the authors.
Mean body-weight and length were about 0.18g and 2.3 cm.

Test type:

static

Water media type:

freshwater

Hardness:

no data

Test temperature:

20-27°C

pH:

test solutions were adjusted to pH7 prior to test

Dissolved oxygen:

No data

Salinity:

No data

Nominal and measured concentrations:

No data

Details on test conditions:

The toxicity was evaluated at 20-27°C using 1-liter glass-beaker containing 10 fish. 
Dilution was made with aerated deionized water. 
The LC50  concentration was determined after 24 hr.

Reference substance (positive control):

not specified

Duration:

24 h

Dose descriptor:

LC50

Effect conc.:

117 mg/L

Description of key information

Brooke et al. (1984 and 1984b) were selected as key studies. Conditions of both of these studies were the same: Pimephales promelas were exposed to vanillin  under flow-through conditions. The 96H-LC50 was 57 mg/L in the first study and 123 mg/L in the second, based on measured concentrations. No discussion about the differing results was provided by the authors. A geometric mean of these 96H-LC50 was calculated at 83,7 mg/L.

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

83.7 mg/L

Additional information

Another 96H-LC50 value on the same fish was in the same range : 112 to 121 mg/L. A 24H-LC50 on Oryzias latipes has been reported at 117 mg/L. Both of these publications have been selected as supporting studies.

Vanillin is considered to be harmful to fish.