Dibismuth trioxide

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to microorganisms

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

disregarded due to major methodological deficiencies

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: Insufficient information on metrhods and results reported. Only nominal concentrations reported without information on solubility of the test substance. Not useful for read-across.

Justification for type of information:

Read-across is considered justified based on the structural similarity between bismuth potassium citrate and bismuth oxide. Both subdtance contain bismuth, the moiety of toxicological concern.

Qualifier:

no guideline followed

Principles of method if other than guideline:

24h toxicity (mortality, endocytosis and growth inhibition) study with Tetrahymena pyriformis

GLP compliance:

not specified

Analytical monitoring:

not specified

Vehicle:

no

Details on test solutions:

Aqueous stock solutions were made up at pH 7, since bismuth salts precipitate below pH 5.5

Test organisms (species):

Tetrahymena pyriformis

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

24 h

Test temperature:

28°C

pH:

6.8-7.0

Nominal and measured concentrations:

Eight nominal added concentrations between 0.21 and 313.5 mg Bi/L

Details on test conditions:

Test organisms were grown axenically in 2% proteose peptone enriched with 0.1% yeast extract and inorganic salts, pH 6.8. The 100 mL cultures in 500 mL Fernbach flasks were aerated and agitated.
For growth experiments, 100 mL cell cultures at a density of 40,000 cells/mL were divided into 2-4 40 mL subscultures. One served as control and Bi was added to the others to make different Bi concentrations.
The endocytic capacity was tested by a 10-min exposure of the cells to a suspension of carmine particles in the same medium as the cells.

Reference substance (positive control):

no

Duration:

24 h

Dose descriptor:

other: LC25

Effect conc.:

104.5 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Bi

Basis for effect:

other: mortality

Duration:

24 h

Dose descriptor:

other: LC55

Effect conc.:

209 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Bi

Basis for effect:

other: Mortality

Duration:

24 h

Dose descriptor:

other: LC78

Effect conc.:

313.5 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Bi

Basis for effect:

other: Mortality

Duration:

24 h

Dose descriptor:

NOEC

Effect conc.:

10.4 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Bi

Basis for effect:

other: Endocytosis

Duration:

24 h

Dose descriptor:

NOEC

Effect conc.:

20.9 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Bi

Basis for effect:

other: Endocytosis

Details on results:

inhibition of growth: Data only allow qualitative estimate of effects based on interpretation of graphs

Reported statistics and error estimates:

NOEC for endocytosis = highest concentration with <10% inhibition.

Validity criteria fulfilled:

not specified

Conclusions:

Only nominal concentrations reported without information on solubility of the test substance; study is disregarded for read-across.

Reference 2

Endpoint:

toxicity to microorganisms

Type of information:

experimental study

Adequacy of study:

disregarded due to major methodological deficiencies

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: Data only allow rough estimate of effects based on interpretation of graphs; E. coli is not relevant for assessment of toxicity to STP.

Qualifier:

no guideline followed

Principles of method if other than guideline:

48h toxicity (growth inhibition) study with E. coli

GLP compliance:

not specified

Analytical monitoring:

not specified

Vehicle:

no

Details on test solutions:

ceramic powder suspended in sterile physiological saline.

Test organisms (species):

Escherichia coli

Details on inoculum:

The test bacteria were cultured in Brain Heart Infusion broth at 310 K for 24 h on a reciprocal shaker. The culture was then suspended in sterile physiological saline to yield a final bacterial concentration of approx. 1000 CFU/mL.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

pH:

11.3

Nominal and measured concentrations:

not reported

Details on test conditions:

Modified plate agar was used as the growth medium. Both bacterial suspension and the ceramic powder slurry were pippeted into the agar. Incubation at 308 K for 48h.

The endpoint is the time required to reach a threshold concentration of 10 E+7 viable organisms/mL.

Reference substance (positive control):

no

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Bi

Basis for effect:

growth inhibition

Details on results:

Data only allow rough estimate of effects based on interpretation of graphs

Reported statistics and error estimates:

no information on statistics is reported.

Validity criteria fulfilled:

no

Conclusions:

Data only allow rough estimate of effects based on interpretation of graphs; study is disregarded.

Reference 3

Endpoint:

activated sludge respiration inhibition testing

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

29 April to 6 May 2010

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP-conform study according to OECD 209 guideline. Rated as Klimisch 2 because it is a read-across study.

Justification for type of information:

Please see read-across justification attached below.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

yes

Remarks:

In the reference test, the respiration rates at the two highest treatment levels were assessed between 7.69 O2/L and 9.12 O2/L, which is outside the range of 6.5 mg O2/L to 2.5 mg O2/L that is recommended in the guideline.

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
- Melting point: > 500 degrees C
- Vapour pressure: 3.4 x 10-5 Pa at 20 degrees C
- Water solubility (under test conditions): 900.7 mg/L at 20 degrees C

Analytical monitoring:

yes

Details on sampling:

Samples from test solutions were taken to determine the actual test item concentrations in comparison to the nominally applied concentrations.
At start of exposure, samples were taken from each test vessel after preparation of the test solutions before adding the microbial inoculum.
At the end of exposure, samples were taken after the oxygen measurements. Appropriate volumes of the test solutions taken at the end of the exposure period were filtered using folded paper filters (pore size: 4 to 7 µm) to remove the microbial inoculum. The filtrates, and the homogenised solutions from the start of exposure were sent to the analytical test site for determination of the test concentrations. The retained solids were stored in the freezer as a reserve until potential further processing.

Vehicle:

no

Details on test solutions:

The test solutions were prepared without using a stock solution.
The test item concentration levels were prepared by stirring corresponding amounts of the test item in 284 mL of deionised water at room temperature in the dark for seven days before adding the synthetic sewage feed and the microbial inoculum. A magnetic stirrer was used which was set to 300 rpm in order to avoid settling of particles.
The controls were prepared by combining 284 mL of temperature adapted dilution medium (deionised water) with 16 mL of the synthetic sewage feed and adding the microbial inoculum.

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

Activated sludge was obtained from the sewage treatment works in frankfurt/Main Niederrad, Germany, treating predominantly domestic sewage. Activated sludge was washed with reconstituted water (according to OECD guideline) and stored for 4 days at 20 ± 2 °C. During storage 50 mL synthetic sewage feed were added per litre activated sludge and per day. The activated sludge was stirred and aerated during storage.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Hardness:

257.2 mg CaCO3/L

Test temperature:

20.3-20.6°C

pH:

6.2

Dissolved oxygen:

No data

Salinity:

Not applicable

Nominal and measured concentrations:

nominal added test item concentrations: 0-4.1-12.3-41-123-411 mg/L
nominal added bismuth concentrations: 0-3.0-9.0-30-90-300 mg V/L
measured dissolved bismuth concentrations: <0.3 mg/L, except for highest dose: 14.1 mg Bi/L.

Details on test conditions:

Storage time of the activated sludge before use: 3 days
Test vessels: 1000 mL glass beakers without cover
Amount of test mixture per test vessel: 500 mL
Volume of microbial inoculum per test mixture: 200 mL
Volume of synthetic sewage feed per test mixture: 16 mL
Mixed liquor suspended solids level in the microbial inoculum: 3.78 g/L
Mixed liquor suspended solids level in the final test mixtures: 1.51 g/L
Number of test concentrations: 5 plus control
Number of replicates per test item concentration: 2 plus one for chemical analysis
Number of replicates in the control: 2 plus one for chemical analysis
Aeration of test vessels: 40 to 50 litre of air per hour (ambient air, oil-free air-compressor)
Effect parameter measured: inhibition of respiration.

Reference substance (positive control):

yes

Remarks:

3,5-Dichlorophenol

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

>= 300 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Bi

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other: with inhibition around 17%

Duration:

3 h

Dose descriptor:

EC10

Effect conc.:

175.4 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Bi

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other: 95% CL: 127.3-209.1 mg Bi/L

Duration:

3 h

Dose descriptor:

other: EC20

Effect conc.:

341.2 mg/L

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

Bi

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other: 95% CL: 292.4-434.1 mg Bi/L

Details on results:

A concentration-related effect was observed only in the highest test item concentration level, where respiration compared to the controls was inhibited by approximately 17%. Since this inhibition was not statistically significant at p ≤ 0.05, EC50 values were not calculated.
3h-EC10 for respiration rate in activated sludge inoculum: 175.4 mg Bi/L (95%CL: 127.3-209.1)
3h-EC20 for respiration rate in activated sludge inoculum: 341.2 mg V/L (95%CL: 292.4-434.1)

The measured test concentrations throughout the test were outside the range of ± 20% (average recoveries of the test concentrations <10%) of the nominal test concentrations. Therefore the calculations of the biological effect concentrations were expressed based on the average recovery of the largest dose tested (4.7%):
3h-NOEC for respiration rate in activated sludge inoculum: ≥14.1 mg Bi/L
3h-EC10 for respiration rate in activated sludge inoculum: 8.2 mg Bi/L (95%CL: 6.0-9.8)
3h-EC20 for respiration rate in activated sludge inoculum: 16.0 mg V/L (95%CL: 13.7-20.4)

Results with reference substance (positive control):

In the reference test 4 concentration levels (1.8, 5.9, 18.8, 60 mg/L) were tested. The reference test was performed according to ECT's SOP A 16.1. A 3h-EC50 value of 8.5 mg/L was obtained. The EC50 value is within the accepted range of 5 to 30 mg/L as required by the test guideline.

Reported statistics and error estimates:

Welch-t test for inhomogeneous variances with Bonferroni-Holm adjustment was used to detect significant differences from the control (p ≤ 0.05)

Validity criteria fulfilled:

yes

Conclusions:

GLP-conform study according to OECD 209 guideline. A 3h-EC10 value for activated sludge respiration inhibition of 175.4 mg Bi/L is obtained and selected for the effects assessment.

Executive summary:

GLP study according to OECD 209 guideline with related substance, bismuth subnitrate.

A 3h-EC10 value for activated sludge respiration inhibition of 175.4 mg Bi/L is obtained and selected for the effects assessment.

Key value for chemical safety assessment

EC50 for microorganisms:

175.4 mg/L

Additional information

The key study is a GLP-conform study according to OECD 209 guideline on the effect of bismuth subnitrate (Bi₅O(OH)₉(NO₃)₄) on the activated sludge respiration inhibition after seven days equilibration of the test substance in deionised water (Egeler et al., 2010). A concentration-related effect was observed only in the highest concentration level (300 mg Bi/L), where respiration compared to the controls was inhibited by approximately 17%. Since this inhibition was not statistically significant at p ≤ 0.05, the NOEC is ≥ 300 mg Bi/L. An EC10 of 175.4 mg Bi/L was calculated based on nominal concentrations.

The measured dissolved Bi concentrations were <0.3 mg Bi/L, except for highest dose, where an average concentration of 14.1 mg Bi/L was measured (corresponding to 4.7% of the total nominal concentration). The NOEC and EC10 expressed as dissolved Bi concentration (based on recovery of 4.7%) are ≥14.1 and 8.2 mg Bi/L.