trans-cyclohexane-1,2-dinitrilotetraacetic acid

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1986

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

excretion

Qualifier:

no guideline followed

Principles of method if other than guideline:

Rats were injected with a single dose of [14C]EDTA. The concentration of EDTA in urine and kidney homogenates was determined at several time points.

GLP compliance:

no

Specific details on test material used for the study:

Specific activity (if radiolabelling): 5 µCi/100 mg

Radiolabelling:

yes

Remarks:

14C

Species:

rat

Strain:

other: Charles River

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River
- Weight at study initiation: 200 - 300 g
- Individual metabolism cages: yes
- Diet: NIH-31 rat food ad libitum
- Water: tap water ad libitum

Route of administration:

intraperitoneal

Vehicle:

not specified

Duration and frequency of treatment / exposure:

single treatment

Dose / conc.:

400 mg/kg bw (total dose)

No. of animals per sex per dose / concentration:

18

Control animals:

other: yes, 0.9% saline treated animals

Details on study design:

18 rats were each injected with a single dose of [14C]EDTA and placed in plastic metabolic cages for urine collection. Six were killed at 16, 22, or 28 hr after the injection. One control rat was killed at each time point. Kidneys were removed, weighed and homogenized. Radioactivity in the urine and kidney homogenates was measured by scintillation counting in order to calculate EDTA concentrations.

Details on dosing and sampling:

PHARMACOKINETIC STUDY
- Body fluids sampled: Urine
- Tissues sampled: Kidneys
- Time and frequency of sampling: urine: 16, 22, 28 h
- Method type for identification: Liquid scintillation counting

Statistics:

Student's t-test, ANOVA and Dunnett's test

Details on distribution in tissues:

- During the 22 h period, the renal EDTA concentration was about 80 µg/g kidney homogenate. This equals to 0.1-0.2% of the EDTA injected.

Details on excretion:

- By 22 hr, 80% + / - 3% of the injected dose had been collected in the urine. This amount stayed approximately the same until 28 h after application.

Metabolites identified:

not measured

Conclusions:

Interpretation of results: no bioaccumulation potential based on study results

Reference 2

Endpoint:

basic toxicokinetics in vivo

Type of information:

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

Adequacy of study:

weight of evidence

Study period:

1986

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

The substance Sodium calcium edetate (CAS 62-33-9) is structurally similar to the substance trans-cyclohexane-1,2-dinitrilotetraacetic acid (CAS 13291-61-7), therefore used for the read-across.

Reason / purpose for cross-reference:

read-across source

Objective of study:

excretion

Qualifier:

no guideline followed

Principles of method if other than guideline:

Rats were injected with a single dose of [14C]EDTA. The concentration of EDTA in urine and kidney homogenates was determined at several time points.

GLP compliance:

no

Specific details on test material used for the study:

Specific activity (if radiolabelling): 5 µCi/100 mg

Radiolabelling:

yes

Remarks:

14C

Species:

rat

Strain:

other: Charles River

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River
- Weight at study initiation: 200 - 300 g
- Individual metabolism cages: yes
- Diet: NIH-31 rat food ad libitum
- Water: tap water ad libitum

Route of administration:

intraperitoneal

Vehicle:

not specified

Duration and frequency of treatment / exposure:

single treatment

Dose / conc.:

400 mg/kg bw (total dose)

No. of animals per sex per dose / concentration:

18

Control animals:

other: yes, 0.9% saline treated animals

Details on study design:

18 rats were each injected with a single dose of [14C]EDTA and placed in plastic metabolic cages for urine collection. Six were killed at 16, 22, or 28 hr after the injection. One control rat was killed at each time point. Kidneys were removed, weighed and homogenized. Radioactivity in the urine and kidney homogenates was measured by scintillation counting in order to calculate EDTA concentrations.

Details on dosing and sampling:

PHARMACOKINETIC STUDY
- Body fluids sampled: Urine
- Tissues sampled: Kidneys
- Time and frequency of sampling: urine: 16, 22, 28 h
- Method type for identification: Liquid scintillation counting

Statistics:

Student's t-test, ANOVA and Dunnett's test

Details on distribution in tissues:

- During the 22 h period, the renal EDTA concentration was about 80 µg/g kidney homogenate. This equals to 0.1-0.2% of the EDTA injected.

Details on excretion:

- By 22 hr, 80% + / - 3% of the injected dose had been collected in the urine. This amount stayed approximately the same until 28 h after application.

Metabolites identified:

not measured

Conclusions:

Interpretation of results: no bioaccumulation potential based on study results

Reference 3

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1967

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

significant methodological deficiencies

Objective of study:

excretion

Qualifier:

no guideline followed

Principles of method if other than guideline:

Rats got ip applications of 14[C] labeled CaNa2EDTA for 10 consecutive days. Additionally one control group was treated with comparable volumes of normal saline and another group with diethylenetriaminepentaacetate (DTPA). Urine and feces were collected daily and the radioactivity was determined. At the end of the experiment the kidneys were analysed for histologic changes.

GLP compliance:

no

Radiolabelling:

yes

Remarks:

14C

Species:

rat

Strain:

other: Long-Evans and Sprangue-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight: 150-450 g
- Individual metabolism cages: yes
- Diet: ad libitum
- Water: ad libitum

Route of administration:

intraperitoneal

Vehicle:

not specified

Details on exposure:

DOSAGE PREPARATION
The radioactive material was diluted and mixed with “cold” salt. The total activity injected per day ranged from 1.1 to 8.8 x l0^6 dpm.

Duration and frequency of treatment / exposure:

10 days. Administered daily

Remarks:

300-436 mg/kg bw/day

No. of animals per sex per dose / concentration:

no data

Control animals:

yes

other: administration of comparable volume of normal saline

Details on study design:

Urine and feces were collected daily. One of the kidneys was taken for counting at the time of sacrifice. Urines were diluted with water and the feces and kidneys were homogenized in a dilution of ammonium hydroxide. Aliquots were counted in a scintillation counter.

Details on dosing and sampling:

PHARMACOKINETIC STUDY
- Body fluids sampled: urine, feces
- Time and frequency of sampling: daily
- Tissues sampled: Kidneys; 24 h after the last injection

Details on excretion:

- 66.4 - 92.3% of the total dose injected was recovered in urine within 24 h after application
- usually less than 5% of the dose was recovered in feces
- in 3 rats fecal excretion amounted to 8, 16.3 and 23.8% of the dose. (In any individual animal the occurrence of a very high fecal excretion on a given day was invariably accompanied by a low urinary excretion and was considered to be due to intestinal puncture incident to the intraperitoneal injection)
- 24 h after the last application the kidneys showed less than 0.1% of the activity injected.
- DTPA showed the same excretion patterns

Metabolites identified:

not measured

Histological changes in the kidneys were grade 4 in the rat that received the largest dose of CaNa2EDTA, grade 2 -3 in 2 of the rats that received CaNa3DTPA and grade 1 -2 in the remaining 2 animals in each group.

Grade 1: small, clear, randomly dispersed, well demarcated vacuoles in the cytoplasm of the epithelium of the proximal tubule

Grade 2: increased number of vacuoles, but the cytoplasm remained clairly visible

Grade 3: larger vacuoles replaced the cytoplasm but did not displace the nucleus

Grade 4: extrusion of cell contents through the disrupted apical border and cell nuclei sometimes displaced

Conclusions:

No bioaccumulation potential based on study results

Reference 4

Endpoint:

basic toxicokinetics in vivo

Type of information:

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

Adequacy of study:

weight of evidence

Study period:

1967

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

significant methodological deficiencies

Justification for type of information:

The substance Sodium calcium edetate (CAS 62-33-9) is structurally similar to the substance trans-cyclohexane-1,2-dinitrilotetraacetic acid (CAS 13291-61-7), therefore used for the read-across.

Reason / purpose for cross-reference:

read-across source

Objective of study:

excretion

Qualifier:

no guideline followed

Principles of method if other than guideline:

Rats got ip applications of 14[C] labeled CaNa2EDTA for 10 consecutive days. Additionally one control group was treated with comparable volumes of normal saline and another group with diethylenetriaminepentaacetate (DTPA). Urine and feces were collected daily and the radioactivity was determined. At the end of the experiment the kidneys were analysed for histologic changes.

GLP compliance:

no

Radiolabelling:

yes

Remarks:

14C

Species:

rat

Strain:

other: Long-Evans and Sprangue-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight: 150-450 g
- Individual metabolism cages: yes
- Diet: ad libitum
- Water: ad libitum

Route of administration:

intraperitoneal

Vehicle:

not specified

Details on exposure:

DOSAGE PREPARATION
The radioactive material was diluted and mixed with “cold” salt. The total activity injected per day ranged from 1.1 to 8.8 x l0^6 dpm.

Duration and frequency of treatment / exposure:

10 days. Administered daily

Remarks:

300-436 mg/kg bw/day

No. of animals per sex per dose / concentration:

no data

Control animals:

yes

other: administration of comparable volume of normal saline

Details on study design:

Urine and feces were collected daily. One of the kidneys was taken for counting at the time of sacrifice. Urines were diluted with water and the feces and kidneys were homogenized in a dilution of ammonium hydroxide. Aliquots were counted in a scintillation counter.

Details on dosing and sampling:

PHARMACOKINETIC STUDY
- Body fluids sampled: urine, feces
- Time and frequency of sampling: daily
- Tissues sampled: Kidneys; 24 h after the last injection

Details on excretion:

- 66.4 - 92.3% of the total dose injected was recovered in urine within 24 h after application
- usually less than 5% of the dose was recovered in feces
- in 3 rats fecal excretion amounted to 8, 16.3 and 23.8% of the dose. (In any individual animal the occurrence of a very high fecal excretion on a given day was invariably accompanied by a low urinary excretion and was considered to be due to intestinal puncture incident to the intraperitoneal injection)
- 24 h after the last application the kidneys showed less than 0.1% of the activity injected.
- DTPA showed the same excretion patterns

Metabolites identified:

not measured

Histological changes in the kidneys were grade 4 in the rat that received the largest dose of CaNa2EDTA, grade 2 -3 in 2 of the rats that received CaNa3DTPA and grade 1 -2 in the remaining 2 animals in each group.

Grade 1: small, clear, randomly dispersed, well demarcated vacuoles in the cytoplasm of the epithelium of the proximal tubule

Grade 2: increased number of vacuoles, but the cytoplasm remained clairly visible

Grade 3: larger vacuoles replaced the cytoplasm but did not displace the nucleus

Grade 4: extrusion of cell contents through the disrupted apical border and cell nuclei sometimes displaced

Conclusions:

No bioaccumulation potential based on study results

Reference 5

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1992

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

other: Effects of CaNa2EDTA on the metabolism of Zn, Cu, Mn

Qualifier:

no guideline followed

Principles of method if other than guideline:

The effect of calcium disodium ethylenediaminetetraacetate (CaNa2EDTA) on the metabolism of Zn, Cu and Mn was investigated in mongrel female dogs. Dogs received either CaNa2EDTA subcutaneously or 0.9% NaCl (controls). Urine was collected every 6 h. Tissue samples were obtained from liver, kidney, duodenum, muscle, hair, skin and bone post exsanguination.

GLP compliance:

no

Specific details on test material used for the study:

Preparation: disodium EDTA was mixed with equimolar quantities of CaCO3, in water and heated with constant stirring until excess CO2 was expelled and the final solution was adjusted to pH 7.4 with NaOH

Species:

dog

Strain:

other: Mongrel

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight: 20-30 Kg
- Source: Canine Control Center
- Diet: Purina Dog Chow (concentration of Zn, Cu or Mn was 1.25; 0.16; or 0.98 mg/g dry weight, respectively)
- Water: ad libitum
- Acclimation period: 3-4 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 25°C

Route of administration:

subcutaneous

Vehicle:

water

Duration and frequency of treatment / exposure:

54 h

Remarks:

280 mg/kg/6 h (original data: 0.75 mmol/kg/6 h)

No. of animals per sex per dose / concentration:

5

Control animals:

other: 0.9% sterile NaCl

Details on study design:

Site of injection was infiltrated with procaine hydrochloride prior to chelate administration.
Concentrations of Zn, Cu and Mn in urine and tissue were determined by atomic absorption spectrophotometry.

Details on dosing and sampling:

- Body fluids sampled: urine; every 6 h
- Tissues sampled: liver, kidney, upper duodenum (upper small intestine), muscle (thigh), bone (femur), hair and skin.
- All samples were stores at -20 °C prior to processing

Statistics:

Analysis of variance (ANOVA) and Duncan’s new multiple range test were used to test for differences between groups. Differences between means were tested by two-way unpaired Student’s t-test.

Details on excretion:

- CaNa2EDTA to dogs caused a significant increase in urine flow when compared to controls

URINARY EXCRETION OF ZN
- significant increase in the excretion of Zn when compared to control (cumulative excretion within 54 h: 24.92 + /- 0.346 mg compared to 1.58 + /- 0.24 mg in the control)
- Maximal excretion of Zn occurred between 6 and 12 h of treatment (17% of total Zn)

URINARY EXCRETION OF CU
CaNa2EDTA treatment resulted in a significant increase in the excretion of Cu when compared to control (cumulative excretion within 54 h: 0.922 + /- 0.24 mg compared to 0.12 + /- 0.03 mg in the control)

URINARY EXCRETION OF MN
CaNa2EDTA treatment resulted in a significant increase in the excretion of Mn when compared to control (cumulative excretion within 54 h: 0.582 + /- 0.166 mg compared to 0.042 + /- 0.01 mg in the control)

Metabolites identified:

no

Tissue levels of Zn, Cu and Mn:

significant decrease in tissue levels of Zn (small intestine, hair and skin) and Mn (hair) when compared to controls ; significant increase of concentration of Cu in kidney when compared to controls.

Conclusions:

The present study was designed to assess the effects of CaNa2EDTA on the metabolism of Zn, Cu and Mn in healthy mongrel dogs. The results show that parental administration of CaNa2EDTA to dogs results in a significant increase in the urinary excretion of Zn, Cu and Mn and in a decrease in Zn levels in duodenum. These data suggest that processes which require Zn could be significantly affected by CaEDTA treatment in the dog. Essential elements like Zn, Cu and Mn which have a higher affinity for EDTA will displace Ca preferentially from Ca-chelate and their corresponding chelates will be excreted in urine.
It was also observed that CaNa2EDTA caused a significant decrease in the levels of Zn and Mn in hair and bone; this suggests that these tissues could be a site of chelate action. The increase of Cu (kidneys) and Zn (kidneys and muscle) was probably associated, in part, with mobilization of these elements from storage tissues followed by redistribution.
It was concluded that the use of CaNa2EDTA for the management of heavy metal poisoning in dogs could adversely affect the metabolism of Zn, Cu and Mn.

Executive summary:

The results of the study show that parental administration of CaNa2EDTA to dogs results in a significant increase in the urinary excretion of Zn, Cu and Mn, decrease in Zn levels in duodenum, skin and hair, decrease in Mn levels in hair and increase in Cu levels in kidneys.

Reference 6

Endpoint:

basic toxicokinetics in vivo

Type of information:

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

Adequacy of study:

weight of evidence

Study period:

1992

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Justification for type of information:

The substance Sodium calcium edetate (CAS 62-33-9) is structurally similar to the substance trans-cyclohexane-1,2-dinitrilotetraacetic acid (CAS 13291-61-7), therefore used for the read-across.

Reason / purpose for cross-reference:

read-across source

Objective of study:

other: Effects of CaNa2EDTA on the metabolism of Zn, Cu, Mn

Qualifier:

no guideline followed

Principles of method if other than guideline:

The effect of calcium disodium ethylenediaminetetraacetate (CaNa2EDTA) on the metabolism of Zn, Cu and Mn was investigated in mongrel female dogs. Dogs received either CaNa2EDTA subcutaneously or 0.9% NaCl (controls). Urine was collected every 6 h. Tissue samples were obtained from liver, kidney, duodenum, muscle, hair, skin and bone post exsanguination.

GLP compliance:

no

Specific details on test material used for the study:

Preparation: disodium EDTA was mixed with equimolar quantities of CaCO3, in water and heated with constant stirring until excess CO2 was expelled and the final solution was adjusted to pH 7.4 with NaOH

Species:

dog

Strain:

other: Mongrel

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Weight: 20-30 Kg
- Source: Canine Control Center
- Diet: Purina Dog Chow (concentration of Zn, Cu or Mn was 1.25; 0.16; or 0.98 mg/g dry weight, respectively)
- Water: ad libitum
- Acclimation period: 3-4 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 25°C

Route of administration:

subcutaneous

Vehicle:

water

Duration and frequency of treatment / exposure:

54 h

Remarks:

280 mg/kg/6 h (original data: 0.75 mmol/kg/6 h)

No. of animals per sex per dose / concentration:

5

Control animals:

other: 0.9% sterile NaCl

Details on study design:

Site of injection was infiltrated with procaine hydrochloride prior to chelate administration.
Concentrations of Zn, Cu and Mn in urine and tissue were determined by atomic absorption spectrophotometry.

Details on dosing and sampling:

- Body fluids sampled: urine; every 6 h
- Tissues sampled: liver, kidney, upper duodenum (upper small intestine), muscle (thigh), bone (femur), hair and skin.
- All samples were stores at -20 °C prior to processing

Statistics:

Analysis of variance (ANOVA) and Duncan’s new multiple range test were used to test for differences between groups. Differences between means were tested by two-way unpaired Student’s t-test.

Details on excretion:

- CaNa2EDTA to dogs caused a significant increase in urine flow when compared to controls

URINARY EXCRETION OF ZN
- significant increase in the excretion of Zn when compared to control (cumulative excretion within 54 h: 24.92 + /- 0.346 mg compared to 1.58 + /- 0.24 mg in the control)
- Maximal excretion of Zn occurred between 6 and 12 h of treatment (17% of total Zn)

URINARY EXCRETION OF CU
CaNa2EDTA treatment resulted in a significant increase in the excretion of Cu when compared to control (cumulative excretion within 54 h: 0.922 + /- 0.24 mg compared to 0.12 + /- 0.03 mg in the control)

URINARY EXCRETION OF MN
CaNa2EDTA treatment resulted in a significant increase in the excretion of Mn when compared to control (cumulative excretion within 54 h: 0.582 + /- 0.166 mg compared to 0.042 + /- 0.01 mg in the control)

Metabolites identified:

no

Tissue levels of Zn, Cu and Mn:

significant decrease in tissue levels of Zn (small intestine, hair and skin) and Mn (hair) when compared to controls ; significant increase of concentration of Cu in kidney when compared to controls.

Conclusions:

The present study was designed to assess the effects of CaNa2EDTA on the metabolism of Zn, Cu and Mn in healthy mongrel dogs. The results show that parental administration of CaNa2EDTA to dogs results in a significant increase in the urinary excretion of Zn, Cu and Mn and in a decrease in Zn levels in duodenum. These data suggest that processes which require Zn could be significantly affected by CaEDTA treatment in the dog. Essential elements like Zn, Cu and Mn which have a higher affinity for EDTA will displace Ca preferentially from Ca-chelate and their corresponding chelates will be excreted in urine.
It was also observed that CaNa2EDTA caused a significant decrease in the levels of Zn and Mn in hair and bone; this suggests that these tissues could be a site of chelate action. The increase of Cu (kidneys) and Zn (kidneys and muscle) was probably associated, in part, with mobilization of these elements from storage tissues followed by redistribution.
It was concluded that the use of CaNa2EDTA for the management of heavy metal poisoning in dogs could adversely affect the metabolism of Zn, Cu and Mn.

Executive summary:

The results of the study show that parental administration of CaNa2EDTA to dogs results in a significant increase in the urinary excretion of Zn, Cu and Mn, decrease in Zn levels in duodenum, skin and hair, decrease in Mn levels in hair and increase in Cu levels in kidneys.

Description of key information

No bioaccumulation potential based on the results of the studies for Sodium calcium edetate (CAS 62 -33 -9) was determined.

In one study, the parental administration of CaNa2EDTA to dogs resulted in a significant increase in the urinary excretion of Zn, Cu and Mn, decrease in Zn levels in duodenum, skin and hair, decrease in Mn levels in hair and increase in Cu levels in kidneys.

The test item trans-cyclohexane-1,2-dinitrilotetraacetic acid (CAS 13291-61-7), structurally similar to the substance tested, is considered not to have bioaccumulation potential.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential