L-Aspartic acid, N,N'-1,2-ethanediylbis-, sodium salt (1:3)

Carcinogenicity

Description of key information

No long-term studies assessing the carcinogenic potential of trisodium EDDS, EDDS acid or its other simple salts have been identified.

In good-quality NCI studies, conducted according to a protocol similar to OECD Guideline 451, trisodium ethylenediaminetetraacetate (EDTA) showed no evidence of carcinogenic potential when fed to rats and mice in the diet for 2 years at up to 7500 ppm (about 375 and 1125 mg/kg bw/day, respectively) (NCI, 1977).

Based on their structural similarity, it is expected that trisodium EDDS is also unlikely to induce tumorigenicity following long-term oral exposure at comparable doses.

Key value for chemical safety assessment

Carcinogenicity: via oral route

Link to relevant study records

Reference

Reference 1

Endpoint:

carcinogenicity: oral

Type of information:

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

Adequacy of study:

weight of evidence

Study period:

no data

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 451 (Carcinogenicity Studies)

Deviations:

yes

Remarks:

only two doses (three are indicated in guideline) and only 20 controls/sex

Principles of method if other than guideline:

Study conducted for the National Cancer Institute using a protocol similar to OECD Guideline 451, on related material

GLP compliance:

no

Remarks:

prior to GLP

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Wilmington, Massachusetts, USA
- Age at study initiation: 6 weeks
- Weight at study initiation: 19-22 g
- Fasting period before study: no data
- Housing: 5 animals of same sex in polycarbonate cages
- Diet (e.g. ad libitum): conventional, ad libitum
- Water (e.g. ad libitum): "acidulated" water (pH 2.5), ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-25
- Humidity (%): 45-55
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 16/8

Route of administration:

oral: feed

Details on exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): twice weekly
- Mixing appropriate amounts with (Type of food): meal
- Storage temperature of food: 4oC

Details on analytical verification of doses or concentrations:

"FDA methods" taking samples from the bottom and two wings of the blender and again after storage at room temperature for 12 days. Recoveries were at least 90% of the theoretical values.

Duration of treatment / exposure:

103 weeks

Frequency of treatment:

ad libitum

Post exposure period:

1 week

Dose / conc.:

3 750 mg/kg bw/day (nominal)

Dose / conc.:

7 500 mg/kg bw/day (nominal)

Dose / conc.:

563 mg/kg bw/day (actual dose received)

Dose / conc.:

1 125 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

50/sex/dose
Controls: 20/sex

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: based on a 7-week dietary study at up to 21,600 ppm
- Rationale for animal assignment (if not random): to provide the same total animal weight per cage

Positive control:

none

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly for first four weeks, then monthly

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data

OPHTHALMOSCOPIC EXAMINATION: no

HAEMATOLOGY: no


CLINICAL CHEMISTRY: no

URINALYSIS: no

NEUROBEHAVIOURAL EXAMINATION: no

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes. All major tissues, organs and gross lesions. Rountinely examined were: skin, lymph nodes, mammary gland, salivary gland, bone marrow, trachea, lungs and bronchi, heart, thyroid, parathyroids, oesophagus, stomach, small intestine, large intestine, liver, gallbladder, pancreas, spleen, kidneys, adrenals, urinary bladder, prostate, uterus, testis, ovary, brain and pituitary. Occasionally other tissues were examined.

Other examinations:

none

Statistics:

Cox's and Tarone's tests for differences in survival between the groups. The Fisher exact test was used to compare the tumour incidence between groups and the Armtage and Cochran test was used for linear trend in proportions, with continuity correction. Bonferroni inequality was used to compare several treated groups with a control group.

Clinical signs:

no effects observed

Description (incidence and severity):

There were no treatment-related clinical signs of toxicity.

Mortality:

no mortality observed

Description (incidence):

No statistically significant or dose-related reduction in survival was reported when compered to controls for either sex.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Body weight gain was depressed in the high-dose males throughout most of the study. In treated females a small, dose-related decrease in body weight gain was observed.

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

no effects observed

Description (incidence and severity):

No significant differences were observed between the groups

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

There were no statistically significant differences between the groups

Histopathological findings: neoplastic:

no effects observed

Description (incidence and severity):

A variety of tumours were found in both treated and control groups, all of which had previously been observed as spontaneous lesions in the mouse in previous studies. A dose-related increase in the incidence of lung tumours in the male mice (11, 18 and 26% in controls, low-dose and high-dose, respectively) was reported, but this was not statistically significant and was in the range of the historical controls. An increased incidence of tumours of the pituitary gland in females and liver in males was observed, but again these were not statistically significant. All other tumours occurred at similar frequency in treated and control animals. The report states "In the judgement of the pathologist, the nature, incidence, and severity of the lesions observed in this study provide no clear evidence of carcinogenic effect of EDTA in mice."

Relevance of carcinogenic effects / potential:

There were no statistically significant treatment-related increases in tumour incidence as compared to controls.

Key result

Dose descriptor:

NOAEL

Effect level:

ca. 7 500 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Remarks:

No statistically significant increase in tumour incidence was detected between the treated and control groups

Dose descriptor:

NOAEL

Effect level:

ca. 3 750 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

Conclusions:

In a good-quality NCI study, conducted according to a protocol similar to OECD Guideline 451, trisodium ethylenediaminetetraacetate (EDTA) showed no clear evidence of carcinogenic potential when fed to mice at up to 7500 ppm (about 1125 mg/kg bw/day) in a 2-year dietary study. Based on their structural similarity, it is expected that trisodium EDDS is also unlikely to induce tumorigenicity following long-term oral exposure.

Executive summary:

In a good-quality NCI study, conducted according to a protocol similar to OECD Guideline 451, trisodium ethylenediaminetetraacetate (EDTA) was assessed for carcinogenic potential in a 2-year oral study in B6C3F1 mice.

 

Groups of 50 mice of each sex were fed 3750 or 7500 ppm (about 563 or 1125 mg/kg bw/day) for 2 years in the diet; groups of 20 control animals of each sex received the untreated diet. Animals were observed twice daily for clinical signs of toxicity and mortalities throughout the study. Body weights were recorded weekly (for the first month) and monthly thereafter. At necropsy all major organs and tissues, plus any gross lesions, were examined microscopically.

 

No signs of overt toxicity were observed; body weight gain was decreased in high-dose males and a small, dose-related decrease was observed in females when compared to the control groups. Survival was similar in all groups. At necropsy, there were no statistically significant differences in the incidence of tumours or non-neoplastic changes between treated animals and controls.

 

In conclusion, trisodium EDTA showed no evidence of carcinogenic potential when fed to mice at up to 7500 ppm (about 1125 mg/kg bw/day) in a 2-year dietary study. Based on the structural similarity of the two ethylenediamines, it is expected that trisodium EDDS is also unlikely to induce neoplasia following long-term oral exposure.

Reference 2

Endpoint:

carcinogenicity: oral

Type of information:

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

Adequacy of study:

weight of evidence

Study period:

no data

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 451 (Carcinogenicity Studies)

Deviations:

yes

Remarks:

only two doses (three are indicated in guideline) and only 20 controls/sex

Principles of method if other than guideline:

Study conducted for the National Cancer Institute using a protocol similar to OECD Guideline 451, on related material

GLP compliance:

no

Remarks:

prior to GLP

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: AR Schmidt, Madison, Wisconsin, USA
- Age at study initiation: 6 weeks
- Weight at study initiation: 85-110 g
- Fasting period before study: no
- Housing: 4 of same sex in polycarbonate cages
- Diet (e.g. ad libitum): conventional, ad libitum
- Water (e.g. ad libitum): "acidulated" water (pH 2.5), ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-25
- Humidity (%): 45-55
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 16/8

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): three times a week
- Mixing appropriate amounts with (Type of food): meal
- Storage temperature of food: 4oC

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

"FDA methods" taking samples from the bottom and two wings of the blender and again after storage at room temperature for 12 days. Recoveries were at least 90% of the theoretical values.

Duration of treatment / exposure:

103 weeks

Frequency of treatment:

daily, ad libitum

Post exposure period:

1 week

Dose / conc.:

3 750 ppm (nominal)

Dose / conc.:

7 500 ppm (nominal)

Dose / conc.:

188 mg/kg bw/day (actual dose received)

Dose / conc.:

375 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

50/sex/dose
Control animals: 20/sex

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: based on a 7-week dietary study at up to 21,600 ppm
- Rationale for animal assignment (if not random): to provide the same total animal weight per cage

Positive control:

none

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice daily

BODY WEIGHT: Yes
- Time schedule for examinations: weekly for first four weeks, then monthly

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: no data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: no data

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: no data

OPHTHALMOSCOPIC EXAMINATION: no

HAEMATOLOGY: no

CLINICAL CHEMISTRY: no

URINALYSIS: no

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes. All major tissues, organs and gross lesions. Rountinely examined were: skin, lymph nodes, mammary gland, salivary gland, bone marrow, trachea, lungs and bronchi, heart, thyroid, parathyroids, oesophagus, stomach, small intestine, large intestine, liver, gallbladder, pancreas, spleen, kidneys, adrenals, urinary bladder, prostate, uterus, testis, ovary, brain and pituitary. Occasionally other tissues were examined

Other examinations:

none

Statistics:

Cox's and Tarone's tests for differences in survival between the groups. The Fisher exact test was used to compare the tumour incidence between groups and the Armtage and Cochran test was used for linear trend in proportions, with continuity correction. Bonferroni inequality was used to compare several treated groups with a control group.

Clinical signs:

no effects observed

Description (incidence and severity):

In the last 6 months of the study corneal opacities, ascites and urine stains were seen in both control and treated groups, presumably due to aging. Survival was slightly higher in treated groups compared to the controls.

Mortality:

no mortality observed

Description (incidence):

In the last 6 months of the study corneal opacities, ascites and urine stains were seen in both control and treated groups, presumably due to aging. Survival was slightly higher in treated groups compared to the controls.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Body weight gain was similar in all groups throughout study.

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

no effects observed

Description (incidence and severity):

No significant differences were observed between the groups

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

No neoplasms were detected in the nervous, musculoskeletal, urinary systems or in the organs of special sense. A variety of endocrine tumours were detected, some types of which were only observed in treated animals. However, since these only occurred in low numbers and were frequently reported in historical control data, they were not considered to be treatment-related. Overall, there was no statistically significant increase in the incidence of tumours in the treated animals compared to the controls. The report states "In the judgement of the pathologist, the nature, incidence, and severity of the lesions observed in this study provide no clear evidence of carcinogenic effect in rats."

Histopathological findings: neoplastic:

no effects observed

Description (incidence and severity):

There were no statistically significant differences between the groups.

Relevance of carcinogenic effects / potential:

There were no statistically significant treatment-related increases in tumour incidence compared to the controls.

Key result

Dose descriptor:

NOAEL

Effect level:

ca. 7 500 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Remarks:

No statistically significant increase in tumour incidence was detected between the treated and control groups

Key result

Dose descriptor:

NOAEL

Effect level:

ca. 7 500 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Remarks:

No treatment-related clinical signs of toxicity were observed, average body weights were comparable to those of the controls and survival was similar (or slightly higher) in the treated animals compared to the controls.

Conclusions:

In a good-quality study conducted according to a protocol similar to OECD Guideline 451, trisodium ethylenediaminetetraacetate (EDTA) showed no evidence of carcinogenic potential when fed to rats at up to 7500 ppm (about 375 mg/kg bw/day) in a 2-year dietary study. Based on their structural similarity, it is expected that trisodium EDDS is also unlikely to induce tumorigenicity following long-term oral exposure.

Executive summary:

Trisodium EDTA was assessed for carcinogenic potential in a 2-year dietary study in Fischer 344 rats.

Groups of 50 rats of each sex were fed 3750 or 7500 ppm in the diet (approximately 188 or 375 mg/kg bw/day) for 2 years; groups of 20 control animals of each sex received the untreated diet. Animals were observed twice daily for clinical signs of toxicity and mortalities throughout the study. Body weights were recorded weekly (for the first month) and monthly thereafter. At necropsy all major organs and tissues plus any gross lesions were examined microscopically.

No signs of overt toxicity were observed and body weight gain was similar in both the treated and control groups throughout the study. Survival was similar, or slightly better, in the treated animals than in the controls. At necropsy, there were no statistically significant differences in the incidence of tumours or non-neoplastic changes between treated animals and controls.

In conclusion, trisodium EDTA showed no evidence of carcinogenic potential when fed to rats at up to 7500 ppm (about 375 mg/kg bw/day) in a 2-year dietary study. Based on the structural similarity of the two ethylenediamines, it is expected that trisodium EDDS is also unlikely to induce neoplasia following long-term oral exposure.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

375 mg/kg bw/day

Study duration:

chronic

Species:

rat

Carcinogenicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Carcinogenicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

According to EU CLP regulation, trisodium EDDS would not be classified as carcinogenic based on the data described.

Additional information

No long-term studies assessing the carcinogenic potential of trisodium EDDS, EDDS acid or its other simple salts have been identified.

 

However, in good-quality NCI studies, conducted according to a protocol similar to OECD Guideline 451, trisodium ethylenediaminetetraacetate (EDTA) was assessed for carcinogenic potential in 2-year dietary studies in Fischer 344 rats and B6C3F1 mice. Groups of 50 animals of each sex were fed 3750 or 7500 ppm (approximately 188 or 375 mg/kg bw/day for rats, and 563 and 1125 mg/kg bw/day for mice) for 2 years; groups of 20 control animals of each sex received the untreated diet. Animals were observed twice daily for clinical signs of toxicity and mortalities throughout the study. Body weights were recorded weekly (for the first month) and monthly thereafter. At necropsy all major organs and tissues plus any gross lesions were examined microscopically. In rats, no signs of overt toxicity were observed and body weight gain was similar in both the treated and control groups throughout the study. Survival was similar, or slightly better, in the treated animals than in the controls. In mice, no signs of overt toxicity were observed, but body weight gain was decreased in high-dose males and a small, dose-related decrease was observed in females when compared to the control groups. Survival was similar in all groups. At necropsy, there were no statistically significant differences in the incidence of tumours or non-neoplastic changes between treated animals and their matched controls (NCI, 1977). The final RAR (EU, 2004) also evaluated these studies and concluded that "that there is no concern on carcinogenic properties of EDTA".

A total of 33 Wistar rats (number and sex per group not reported) were divided into 5 groups and dosed with 0, 0.5, 1.0, or 5.0% disodium EDTA in the diet for a period of 2 years (approximately 0, 250, 500, and 2500 mg/kg bw/day, respectively). No gross lesions were observed upon histopathological examination of the internal organs and tissues (which included the heart, liver, pancreas, kidneys, urinary bladder, stomach, small and large intestines, lungs, spleen, ovaries, testes, voluntary muscle, and bone marrow smears). Based on the results of this study, the NOEL was considered to be 2500 mg/kg bw/day disodium EDTA in rats (Yang, 1952). In addition, a NOEL of 250 mg/kg bw/day (the highest tested dose) was seen in Wistar rats (25/sex/group) fed calcium disodium EDTA in the diet for 2 years (Oser et al. 1963).

Based on its structural similarity with EDTA, and its lack of mutagenic activity, it is expected that trisodium EDDS is also unlikely to induce neoplasia following long-term oral exposure at comparable doses.

References (not included elsewhere in IUCLID dossier - need to move to reference list in CSR)

BIBRA (1964). Summaries of toxicological data. Toxicology of EDTA. Food and Cosmetics Toxicology 2, 763-7.

EU (2004). European Union Risk Assessment Report (RAR); edetic acid (EDTA). Vol. 49. European Chemicals Bureau (ECB). Final report available at http://ecb.jrc.ec.europa.eu/DOCUMENTS/Existing-Chemicals/RISK_ASSESSMENT/REPORT/edtareport061.pdf.

Yang S-S (1952). Toxicological investigation of ethylenediaminetetraacetic acid in the rat. Thesis dated May 1952 (cited in BIBRA, 1964).