Titanium dioxide

Long-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

fish short-term toxicity test on embryo and sac-fry stages

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: Shortcomings include: no quantification of titanium concentration of test solutions; test medium, loading rate and other test parameter are not described; no information on validity criteria & low number of fertilised eggs per treatment & control.

Reason / purpose:

reference to same study

Qualifier:

according to

Guideline:

OECD Guideline 212 (Fish, Short-term Toxicity Test on Embryo and Sac-Fry Stages)

Deviations:

yes

Remarks:

: several test conditions were not met, including temperature

GLP compliance:

not specified

Remarks:

not specified in the publication

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

Direct addition of test material into test vessel (dispersion): yes via stock solution
Sonication/stirring of stock solution: 30 min sonication and subsequent agitation in darkness for 24 h prior to preparation of test solutions
Stirring of test solution: preparation under agitation and further sonication for 15 min prior test
Solvents: no
Filtration or other separation of test material from test solution: no
Appearance of solution: no information

test medium: fish medium

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebra fish
- Strain: wild type

METHOD FOR PREPARATION AND COLLECTION OF FERTILIZED EGGS
- Numbers of parental fish (i.e. of females used to provide required number of eggs): no data
- Method of collection of fertilised eggs: zebrafish embryos were obtained by natural mating
- Subsequent handling of eggs: embryos were maintained in a dark incubator in fish culture water at 28.5 °C

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

8 d

Hardness:

no data

Test temperature:

28.5 °C

pH:

no data

Dissolved oxygen:

no data

Salinity:

no data

Nominal and measured concentrations:

nominal concentration: 0, 10, 100, and 1000 mg n-TiO2/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 6-well plate
- Fill volume: 5 mL
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 1

OTHER TEST CONDITIONS
- Photoperiod: test was performend in dark

Reference substance (positive control):

not specified

Duration:

8 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

; 3 different n-TiO2 materials (each individually tested as dispersion)

Basis for effect:

mortality

Remarks:

(embryonic/larval survival)

Duration:

8 d

Dose descriptor:

LOEC

Effect conc.:

1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

; 3 different n-TiO2 materials (each individually tested as dispersion)

Basis for effect:

length

Remarks:

p < 0.05

Details on results:

Survival of developing zebrafish embryos in controls and exposed to n-TiO2 was not significantly different. Nanomaterial aggregates of TiO2 were marginally toxic to 8 dpf zebrafish embryos, affecting their body length at 1 g/L. A NOEC should not be derived from these two experiments since the separation factor should not exceed 3.2 according to OECD 212. No other morphological effects were observed in 8 dpf exposed embryos.

Validity criteria fulfilled:

not specified

Conclusions:

Three different titanium dioxide nanomaterials did not significantly affect the survival of zebrafish embryos in an early life stage toxicity test (OECD TG 212), and a NOEC (8d) of ≥ 1000 mg n-TiO2/L (as dispersion) was determined for the nanomaterials P25, NM-103, and NM-104. Further, morphological effects were not observed beside some marginal effects on body length at the highest test concentration (1000 mg n-TiO2/L as dispersion). A NOEC should not be derived from these two experiments since the separation factor should not exceed 3.2 according to OECD 212.

Reference 2

Endpoint:

fish short-term toxicity test on embryo and sac-fry stages

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: Russian study, no quantification of titanium concentration of test solutions, little information on test performance

Reason / purpose:

reference to same study

Qualifier:

no guideline followed

Principles of method if other than guideline:

A 115-d early-life stage toxicity test conducted with TiO2 with concentrations up to 3 mg/L. Survival and growth of Coregonus autumnalis migratorius G was observed.

GLP compliance:

no

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

Direct addition of test material into test vessel (dispersion): most likely
Sonication/shaking or stirring of stock or test solution (duration): no information
Solvents: no
Filtration or other separation of test material from test solution: no
Appearance of solution: no information

Test organisms (species):

other: Coregonus autumnalis migratorius G.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

115 d

Test temperature:

4.4°C +/- 0.6°C

pH:

7.6-7.8

Dissolved oxygen:

11-12 mg/L

Salinity:

0.1%

Nominal and measured concentrations:

nominal: 0 (control), 1, 2, 3 mg/L
suspension tested

Details on test conditions:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water

Reference substance (positive control):

not specified

Duration:

115 d

Dose descriptor:

NOEC

Effect conc.:

>= 3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

TiO2 as dispersion

Basis for effect:

mortality

Remarks:

, length and weight (p<0.05)

Validity criteria fulfilled:

not specified

Conclusions:

In a 115-d early-life stage toxicity test, TiO2 concentrations up to 3 mg/L (as dispersion) did not affect survival and growth of Coregonus autumnalis migratorius G. A stimulation of growth (weight and length) was observed.

Reference 3

Endpoint:

fish, juvenile growth test

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: Russian study, no quantification of titanium concentration of test solutions, little information on test performance

Reason / purpose:

reference to same study

Qualifier:

no guideline followed

Principles of method if other than guideline:

A 30-d juvenile fish growth test conducted with TiO2 with concentrations up to 3 mg/L. Survival and growth of Coregonus autumnalis migratorius G was observed.

GLP compliance:

no

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

Direct addition of test material into test vessel (dispersion): most likely
Sonication/shaking or stirring of stock or test solution (duration): no information
Solvents: no
Filtration or other separation of test material from test solution: no
Appearance of solution: no information

Test organisms (species):

other: Coregonus autumnalis migratorius G.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

30 d

Test temperature:

5.8°C (4-6°C)

pH:

7.6-7.8

Dissolved oxygen:

11-12 mg/L

Salinity:

0.1%

Nominal and measured concentrations:

nominal: 0 (control), 1, 2, 3 mg/L
suspension tested

Details on test conditions:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water

Reference substance (positive control):

not specified

Duration:

30 d

Dose descriptor:

NOEC

Effect conc.:

>= 3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

TiO2 as dispersion

Basis for effect:

mortality

Remarks:

, length and weight (p<0.05)

Validity criteria fulfilled:

not specified

Conclusions:

In a 30-d juvenile fish growth test, TiO2 concentrations up to 3 mg/L (as dispersion) did not affect survival and growth of Coregonus autumnalis migratorius G. A stimulation of growth (weight) was observed.

Reference 4

Endpoint:

adult fish: sub(lethal) effects

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: Russian study, no quantification of titanium concentration of test solutions, little information on test performance given available

Reason / purpose:

reference to same study

Qualifier:

no guideline followed

Principles of method if other than guideline:

A 28-d adult fish growth test conducted with TiO2 with concentrations up to 1000 mg/L. Survival and growth of Phoxinus phoxinus was observed.

GLP compliance:

no

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

Direct addition of test material into test vessel (dispersion): most likely
Sonication/shaking or stirring of stock or test solution (duration): no information
Solvents: no
Filtration or other separation of test material from test solution: no
Appearance of solution: no information

Test organisms (species):

Phoxinus phoxinus

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

28 d

Test temperature:

10.8°C (6-15 °C)

pH:

7.6-7.8

Dissolved oxygen:

11-12 mg/L

Salinity:

0.1%

Nominal and measured concentrations:

nominal: 0 (control), 100, 500, 1000 mg/L
suspension tested

Details on test conditions:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water

Reference substance (positive control):

not specified

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

TiO2 as dispersion

Basis for effect:

mortality

Remarks:

, length and weight

Validity criteria fulfilled:

not specified

Conclusions:

In a 28-d adult fish growth test, TiO2 concentrations up to 1000 mg/L (as dispersion) did not affect survival and growth of Phoxinus phoxinus.

Reference 5

Endpoint:

fish, juvenile growth test

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: Russian study, no quantification of titanium concentration of test solutions, little information on test performance given available

Reason / purpose:

reference to same study

Qualifier:

no guideline followed

Principles of method if other than guideline:

A 30-d adult fish growth test conducted with TiO2 with concentrations up to 1000 mg/L. Survival and growth of Phoxinus phoxinus was observed.

GLP compliance:

no

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

Direct addition of test material into test vessel (dispersion): most likely
Sonication/shaking or stirring of stock or test solution (duration): no information
Solvents: no
Filtration or other separation of test material from test solution: no
Appearance of solution: no information

Test organisms (species):

Phoxinus phoxinus

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

30 d

Test temperature:

12°C (10-15 °C)

pH:

7.6-7.8

Dissolved oxygen:

11-12 mg/L

Salinity:

0.1%

Nominal and measured concentrations:

nominal: 0 (control), 100, 500, 1000 mg/L
suspension tested

Details on test conditions:

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: tap water

Reference substance (positive control):

not specified

Duration:

30 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

TiO2 as dispersion

Basis for effect:

mortality

Remarks:

, length

Duration:

30 d

Dose descriptor:

NOEC

Effect conc.:

>= 500 - < 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

TiO2 as dispersion

Basis for effect:

weight

Duration:

30 d

Dose descriptor:

LOEC

Effect conc.:

1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

TiO2 as dispersion

Basis for effect:

weight

Remarks:

(p 0.05)

Validity criteria fulfilled:

not specified

Conclusions:

In a 30-d juvenile fish growth test, TiO2 concentrations up to 1000 mg/L (as dispersion) did not affect survival and growth (length) of Phoxinus phoxinus. However, weight of fish iat TO2 concentrations at 1000 mg/L was significantly smaller than the weight of control fish.

Reference 6

Endpoint:

fish short-term toxicity test on embryo and sac-fry stages

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

other: Shortcomings include: no quantification of titanium concentration of test solutions; test medium, loading rate and other test parameter are not described; no information on validity criteria & low number of fertilised eggs per treatment & control.

Reason / purpose:

reference to same study

Qualifier:

according to

Guideline:

OECD Guideline 212 (Fish, Short-term Toxicity Test on Embryo and Sac-Fry Stages)

Deviations:

yes

Remarks:

: several test conditions were not met, including temperature

GLP compliance:

not specified

Remarks:

not specified in the publication

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

Direct addition of test material into test vessel (dispersion): yes via stock solution
Sonication/stirring of stock solution: 30 min sonication and subsequent agitation in darkness for 24 h prior to preparation of test solutions
Stirring of test solution: preparation under agitation and further sonication for 15 min prior test
Solvents: no
Filtration or other separation of test material from test solution: no
Appearance of solution: no information

test medium: fish medium

Test organisms (species):

Danio rerio (previous name: Brachydanio rerio)

Details on test organisms:

TEST ORGANISM
- Common name: zebra fish
- Strain: wild type

METHOD FOR PREPARATION AND COLLECTION OF FERTILIZED EGGS
- Numbers of parental fish (i.e. of females used to provide required number of eggs): no data
- Method of collection of fertilised eggs: zebrafish embryos were obtained by natural mating
- Subsequent handling of eggs: embryos were maintained in a dark incubator in fish culture water at 28.5 °C

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

8 d

Hardness:

no data

Test temperature:

28.5 °C

pH:

no data

Dissolved oxygen:

no data

Salinity:

no data

Nominal and measured concentrations:

nominal concentration: 0, 10, 100, and 1000 mg TiO2/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 6-well plate
- Fill volume: 5 mL
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 1

OTHER TEST CONDITIONS
- Photoperiod: test was performend in dark

Reference substance (positive control):

not specified

Duration:

8 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

TiO2 as dispersion

Basis for effect:

mortality

Remarks:

(embryonic/larval survival)

Duration:

8 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

TiO2 as dispersion

Basis for effect:

length

Remarks:

p < 0.05

Details on results:

Survival of developing zebrafish embryos in controls and exposed to microsized TiO2 was not significantly different. Morphological effects were not observed in 8 dpf exposed embryos.

Validity criteria fulfilled:

not specified

Conclusions:

Microsized TiO2 did not significantly affect the survival and length of zebrafish embryos in an early life stage toxicity test (OECD TG 212), and a NOEC (8d) of ≥ 1000 mg TiO2/L (as dispersion) was determined. Further, morphological effects were not observed up to the highest test concentration (1000 mg TiO2/L as dispersion).

Description of key information

Results of several studies of the toxicity to early life-stages of different fish species indicate that dispersed microsized TiO2 is not chronically toxic to freshwater fish up to at least 500 mg/L (NOEC ≥ 500- 1000 mg/L) whereas dispersed nanosized TiO2 is not chronically toxic up to 100 mg/L (NOEC ≥ 100 - < 1000 mg/L). Considering the low solubility of microsized TiO2, it is further concluded that microsized TiO2 is not chronically toxic up to its solubility limit.

Additional information

Microsized TiO2:

Studies investigating the toxicity of microsized TiO2 to early life-stages and adults of freshwater fish including Danio rerio, Phoxinus phoxinus, and Coregonus atuumnalis migratorius demonstrate that microsized TiO2 is not chronically toxic to freshwater fish up to its solubility limit and dispersed TiO2 concentration of 500 mg/L. In a 115 d early-life stage toxicity test with embryos and in a 30 d juvenile fish growth test by Beim et al (1982) with Coregonus autumnalis migratorius, dispersed microsized TiO2 concentrations up to 3 mg/L did not negatively affect survival and growth of exposed fish (115 d NOEC and 30 d NOEC: ≥ 3 mg/L (nominal)). To the contrary, a stimulation of growth, weight and length, was observed (Beim et al. 1982). In a 28 d fish growth test by Beim et al (1982) with adult Phoxinus phoxinus, dispersed microsized TiO2 concentrations up to 1000 mg/L (nominal) did not affect survival and growth of exposed fish (28 d NOEC: ≥ 1000 mg/L (nominal)). Faria et al. (2014) also observed that dispersed microsized TiO2 did not significantly affect the survival and length of zebrafish embryos in an early life-stage toxicity test following OECD TG 212. Morphological effects were also not observed up to the highest test concentration and the determined 8 d NOEC amounts to ≥ 1000 mg TiO2/L (dispersed, nominal).

Beim et al. (1982) performed a 30 d fish growth test with juvenile Phoxinus phoxinus and microsized TiO2 concentrations of up to 1000 mg/L (dispersed, nominal) did not affect survival and growth (length) of Phoxinus phoxinus whereas the weight of fish exposed to microsized TiO2 concentrations of 1000 mg/L were significantly lower than the weight of control fish, resulting in a 30 d NOEC of ≥ 500- < 1000 mg/L (nominal).

One further supporting study additionally demonstrates that microsized TiO2 materials are not chronically toxic to freshwater fish: In a study by Shaw et al. (2016), the titanium dioxide bulk material NM-100 (148.3 ± 109.7 nm) did not affect embryo survival, hatching rate and the time to hatch of Danio rerio embryos at the highest test concentration of 160 mg/L (as dispersion) in a fish early-life stage toxicity test (OECD TG 210) after 4 d and 6 d of exposure. Further, an effect on larvae length, muscle block width and yolk sac volume could not be observed after 6 d of exposure (NOEC ≥ 160 mg/L (nominal)). Due to the shorter exposure period of 6 d instead of 30 d as recommended in OECD 210, this study is considered supporting.

However, all chronic fish experiments were performed at concentrations several magnitudes above the solubility limit of microsized TiO2. Transformation/dissolution data of different microsized TiO2 materials indicate a low solubility in environmental media as dissolved Ti concentrations after 28 d were below the respective LOD/LOQ (< 0.11 / < 0.34 µg Ti/L). In sum, it is concluded that microsized TiO2 is not chronically toxic to fish up to concentrations of 1000 mg/L dispersed TiO2 and its solubility limit in freshwater.

Marine data do not exist.

Nanosized TiO2:

Three different titanium dioxide nanomaterials P25, NM-103, and NM-104 did not significantly affect the survival of zebrafish embryos in an early life-stage toxicity test (OECD TG 212) and 8 d NOEC values of ≥ 1000 mg n-TiO2/L (dispersed, nominal) were derived by Faria et al (2014). Furthermore, morphological effects were not observed with the exception of marginal effects on body length at the highest test concentration of 1000 mg n-TiO2/L but not at 100 mg n-TiO2/L (dispersed). A NOEC should nevertheless not be derived from the study by Faria et al (2014) since the separation factor should not exceed 3.2 according to OECD 212. Further, according to OECD 212, concentrations higher than 100 mg/L should not to be tested. Three supporting studies, including one fish early-life stage toxicity test (Bar-Ilan et al. 2013), one short-term toxicity test on fish embryo and sac fry stages (Bar-Ilan et al. 2012), and one prolonged fish toxicity test (Clemente et al. 2015), additionally indicate that nanosized TiO2 is not chronically toxic to embryos and juveniles of Danio rerio and Piaracus mesopotamicus since unbounded NOEC values ranged from ≥ 100 to ≥ 10,000 mg/L dispersed n-TiO2. The latter studies are considered as supporting studies since validity criteria were not provided (Bar-Ilan et al. 2013, Clemente et al. 2015), validity criteria were not met (Bar-Ilan et al. 2013), and/or tests were performed with too much biomass (Clemente et al. 2015).

One further supporting study additionally demonstrates that nanosized TiO2 materials are not chronically toxic to freshwater fish: The titanium dioxide nanomaterial NM-105 (20.6 ± 3.9 nm) did not affect embryo survival and hatching rate of Danio rerio at the highest test concentration of 160 mg/L (as dispersion) in an early-life stage fish toxicity test (OECD TG 210) after 4 d and 6 d of exposure. No effect on larvae length, muscle block width and yolk sac volume could be observed after 6 d of exposure (NOEC ≥ 160 mg/L (nominal)). Although a delay of hatching could be observed at the highest tested nano-TiO2 concentration (6 d LOEC: 160 mg/L; 6 d NOEC: 80 mg/L (nominal)), effects occurred at nano-TiO2 concentrations exceeding the recommended limit test concentration of 10 mg/L according to OECD 210. Hence, nano-TiO2 is considered to be not chronically toxic to Danio rerio embryos. Due to the shorter exposure period of 6 d instead of 30 d, as recommended in OECD 210, this study is considered supporting.

Finally, based on reliable chronic toxicity studies it is concluded that nanosized TiO2 dispersions are not chronically toxic to freshwater fish up to 100 mg/L.

Marine data do not exist.

Based on a weight of evidence approach, dispersed microsized TiO2 is not chronically toxic to freshwater fish up to 1000 mg/L. Considering the low solubility of microsized TiO2, it is further concluded that microsized TiO2 is not toxic up to its solubility limit. Dispersions of nanosized TiO2 are not chronically toxic to freshwater fish up to 100 mg/L (NOEC > 100 mg/L).