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Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
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 available
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.
Radiolabelling:
no
Species:
dog
Strain:
other: Mongrel
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- 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
Dose / conc.:
280 other: mg/kg/6h
Remarks:
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 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.



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

- Statistically significant decrease in the tissue levels of Zn (small intestine, hair and skin) and Mn (hair) when compared to controls

- Statistically significant increase in the concentration of Cu in the kidney when compared to controls

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
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 available
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
Radiolabelling:
yes
Remarks:
14C
Species:
rat
Strain:
other: CD
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 other: mg/kg bw
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, weight 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 absorption:
100%
Details on distribution in tissues:
- During the 22 h period, the renal EDTA concentration was about 80 µg/g kidney homogenate. This equals 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
Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
3 (not reliable)
Rationale for reliability incl. deficiencies:
unsuitable test system
Objective of study:
excretion
Qualifier:
no guideline available
Principles of method if other than guideline:
At regular intervals up to 32 h after a single dose of 95 mg Na2EDTA the gastrointestinal tract was removed and the stomach, small intestine and colon subjected to thorough washing before determining the EDTA concentrations of each of these sections.
GLP compliance:
no
Radiolabelling:
not specified
Species:
rat
Strain:
not specified
Sex:
not specified
Route of administration:
oral: gavage
Vehicle:
not specified
Details on exposure:
no data
Duration and frequency of treatment / exposure:
no data
Dose / conc.:
95 other: mg/animal
Remarks:
ca. 475 mg/kg bw
No. of animals per sex per dose / concentration:
no data
Control animals:
not specified
Details on study design:
no data
Details on dosing and sampling:
no data
Statistics:
no data
Details on absorption:
All EDTA had passed through the stomach within 12 h and 93% of the dose was recovered in the colon after 32 h, which demonstrated a poor absorption from GI tract.
Details on distribution in tissues:
The contents of the small intestine and urine reached a maximum about 4 h after dosage.
Details on excretion:
Urinary excretion over the period of 32 h cumulated to 6% of the dose.
Details on metabolites:
no data

- When adults received daily doses of 95 mg Na2EDTA for 7 days the percentage of daily excretion in urine and feces was similar.

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
3 (not reliable)
Rationale for reliability incl. deficiencies:
other: Inconsistent results.
Objective of study:
excretion
Principles of method if other than guideline:
No data
GLP compliance:
no
Radiolabelling:
not specified
Species:
rat
Strain:
Wistar
Sex:
not specified
Route of administration:
oral: feed
Vehicle:
not specified
Details on exposure:
no data
Duration and frequency of treatment / exposure:
no data
Dose / conc.:
300 mg/kg bw/day
Remarks:
original data 0.5% of the diet, conversion according to EU risk assessment
Dose / conc.:
600 mg/kg bw/day
Remarks:
original data:1.0% of the diet, conversion according to EU risk assessment
Dose / conc.:
3 000 mg/kg bw/day
Remarks:
original data 5.0% of the diet, conversion according to EU risk assessment
No. of animals per sex per dose / concentration:
no data
Control animals:
not specified
Details on study design:
no data
Details on dosing and sampling:
no data
Statistics:
no data
Details on absorption:
no data
Details on distribution in tissues:
no data
Details on excretion:
- 82.2% of the ingested 0.5% in diet dose was excreted in urine and feces thereof 99.4% were recovered from feces and 0.6% in urine.
- 44.5% of the ingested 1.0% in diet dose was excreted in urine and feces thereof 98.2% were recovered from feces and 1.8% in urine.
- 45.4% of the ingested 5.0% in diet dose was excreted in urine and feces thereof 97.5% were recovered from feces and 2.5% in urine.
Details on metabolites:
no data
Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
documentation insufficient for assessment
Objective of study:
excretion
Qualifier:
no guideline available
Principles of method if other than guideline:
48 h after administration of Na2EDTA and Na2CaEDTA the percentage of administered dose in feces and urine was determined.
GLP compliance:
no
Radiolabelling:
no
Species:
rat
Strain:
not specified
Sex:
not specified
Details on test animals or test system and environmental conditions:
- weanlings and adults were tested
Route of administration:
oral: gavage
Vehicle:
not specified
Duration and frequency of treatment / exposure:
single exposure
Dose / conc.:
600 other: mg/kg bw
Remarks:
equivalent to 1% EDTA in diet (corresponding to 600 mg/kg bw, conversion according to EU risk assessment)
No. of animals per sex per dose / concentration:
no data
Control animals:
no
Details on excretion:
within 48 h:
WEANLINGS
Na2EDTA: 8.5% in urine; 88.6% in feces

ADULTS
Na2EDTA: 5.3% in urine; 89.9% in feces
Na2CaEDTA: 4.6% in urine, 88.5% in feces

3% of the EDTA was removed from the surface of metabolism cage and 5% from intubation syringe
Metabolites identified:
no
Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
documentation insufficient for assessment
Objective of study:
excretion
Qualifier:
no guideline available
Principles of method if other than guideline:
Single doses of Na2EDTA were administered and the amount excreted in urine determined.
GLP compliance:
no
Radiolabelling:
not specified
Species:
rat
Strain:
not specified
Sex:
not specified
Route of administration:
oral: gavage
Vehicle:
not specified
Details on exposure:
no data
Duration and frequency of treatment / exposure:
no data
Dose / conc.:
237.5 mg/kg bw (total dose)
Remarks:
47.5 mg/animal, corresponding to 237.5 mg/kg bw assuming a body weight of 200 g
Dose / conc.:
475 mg/kg bw (total dose)
Remarks:
95.0 mg/animal, corresponding to 475.0 mg/kg bw assuming a body weight of 200 g
Dose / conc.:
712.5 mg/kg bw (total dose)
Remarks:
142.5 mg/animal, corresponding to 712.5 mg/kg bw assuming a body weight of 200 g
No. of animals per sex per dose / concentration:
no data
Control animals:
not specified
Details on study design:
no data
Details on dosing and sampling:
no data
Statistics:
no data
Details on absorption:
no data
Details on distribution in tissues:
no data
Details on excretion:
The amount of EDTA excreted in urine was directly proportional to the amount ingested. The peak urinary excretion was after 4 h. The authors concluded that due to the linear relationship EDTA is most likely absorbed by diffusion.
Details on metabolites:
no data
Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
documentation insufficient for assessment
Objective of study:
absorption
excretion
metabolism
toxicokinetics
Qualifier:
no guideline available
Principles of method if other than guideline:
Metabolism studies were conducted in rat. First distribution and balance studies were carried out with male animals. The animals were housed individually in metabolism cages for the separate collection of expired CO2, urine and feces. In a second series of experiments detailed studies on blood and urine were conducted on female animals. Studies were made after intraperitoneal, intravenous, intramuscular, and oral administration (intubation).
GLP compliance:
no
Radiolabelling:
yes
Remarks:
14C
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Weight at study initiation: 200 - 250 g
Route of administration:
other: ip, iv, im and oral (gavage)
Vehicle:
water
Duration and frequency of treatment / exposure:
single dose
Dose / conc.:
50 mg/kg bw (total dose)
Remarks:
additionally, one study was performed using 250 mg/kg bw for ip application
No. of animals per sex per dose / concentration:
no data
Control animals:
no
Details on absorption:
Oral studies:
2-18% (most of the values were in the range of 2-4%)
Details on distribution in tissues:
- no organ retained more than 0.5% of the dose
Details on excretion:
ORAL
- 2-18% of the dose was excreted in the urine
- 80-95% of the dose was excreted via feces within 24 h
- less than 0.1% of the dose was exhaled as CO2

PARENTERAL ADMINISTRATION
- 95 - 98% of the dose appeared in the urine within 6 h (iv: 96.09%; ip: 98.67%; im: 95.35%)
- less than 0.1% of the dose was exhaled as CO2
- the dose size 250 mg/kg bw vs 50 mg/kg bw had no effect on the excretion kinetics
(see table 1)
Key result
Toxicokinetic parameters:
half-life 1st: 32 min (half-live urine; im application)
Key result
Toxicokinetic parameters:
half-life 1st: 31 min (half-live urine; iv application)
Key result
Toxicokinetic parameters:
half-life 1st: 35 min (half-live urine; ip application)
Toxicokinetic parameters:
half-life 1st: no data (half-live urine; oral)
Details on metabolites:
Chromatography provided evidence that EDTA is not metabolized but passes the body unchanged.

While the half-life determine in urine was 31 to 35 min after parenteral application, a 50% drop of activity in plasma took 35- 50 min.

Table 1: Distribution of 14C activity following administration of Ca C14 -EDTA to rats

time after ip administration oral administration
Tissue sample 1.5 h 6 h 24 h 24 h
Urine 85.2 95.7 94.6 10.3
Feces 0.5 2.0 3.6 88.3
Expired CO2 0.05 0.08
Skin 2.3 0.8 0.3 0.16
Kidney 1.4 0.3 0.3 0.04
GI tract 0.2 0.2 0.7 0.45
Liver 0.5 0.2 0.05 0.18
Skeleton 0.3 0.1 0.08
Muscle 0.1 0.3 0.36
Blood 1.7 0.1 0.01 0.04
Remains 8.1 0.03 0.01 0.03

Description of key information

In rat as well as in human studies CaNa2EDTA and Na2EDTA is poorly absorbed from GI tract and the absorbed part is rapidly excreted via urine, therefore one can rule out the danger of bioaccumulation. However, administration of EDTA salts results in an increased excretion of necessary ions like Zn, Mn or Ca.

Key value for chemical safety assessment

Additional information

No studies on the toxicokinetics of diammonium dihydrogen ethylenediamine or other ammonium salts of EDTA are available. However, several studies have been performed using CaNa2EDTA or Na2EDTA. According to the dissociation equilibrium of EDTA salts, administration of different salts will result in the formation of various anionic species of EDTA and the respective cationic ions in dependence on the intestinal pH value.

In whatever salt EDTA is administered it is likely to chelate metal ions in vivo. Therefore the studies using CaNa2EDTA have been used as read-across. It can be assumed that the systemic absorption of EDTA anions from the intestinal tract is low. The obtained data can be used to predict that dermal absorption should be even lower. Additionally, absorbed EDTA does not undergo any biotransformation and is excreted unchanged.

In toxicokinetics studies on humans as well as rats the CaNa2 and Na salts of EDTA are poorly absorbed from the gastrointestinal tract (2 -18% in rats; less than 5% in humans) CaNa2EDTA does not penetrate the skin, only 0.001% were absorbed within 24 h of administration. Intravenously applied EDTA is rapidly excreted in urine (humans 50% within the first hour 98% within 24 h; rats: 95%- 98% within 6 h).

The ammonium ions which are intestinally absorbed, are converted to urea in the liver, and subsequently excreted in urine (within 6 hours) it can also be utilized to form amino acids and proteins. The ratio of ammonium ion to neutral ammonia is about 100 in blood of normal pH range (WHO, 1986)

Discussion on bioaccumulation potential result:

In a study conducted by Foreman et al. (1953) 50 mg/kg bw of 14C labeled calcium salt of EDTA were administered to rats orally (gavage), intraperitoneally, intravenously or intramuscularly. After oral application calcium EDTA was poorly absorbed from gastrointestinal tract (2 - 18% within 24 h). Most of the administered dose was excreted by feces 80 - 95% and much less in urine (2 -18%). After parenteral application 95 - 98% (iv: 96.09%; ip: 98.67%; im: 95.35%) of the radioactivity was excreted in urine within 6 h after application, while less than 0.1% was exhaled as CO2. None of the tissues contained at this time point more than 0.5% of the radioactivity administered.

Two additional studies on the toxicokinetics of CaNa2EDTA after i.p. application are available. In one study rats got 10 injections of 300 - 436 mg/kg bw/day 14 C labeled CaNa2EDTA. 66 - 92% of the administered dose were recovered in urine while generally less than 5% were excreted by feces. 24 h after the last injection kidneys showed less than 0.1% of the radioactivity (Doolan, 1967). In the other study, 18 rats got a single i. p. application of 400 mg/kg bw 14[C] CaNa2EDTA. Within 22 h 80% of the radioactivity were excreted in urine, while the concentration in kidney homogenate was approximately 0.1 - 0.2% during this time period (Miller, 1986).

The effects of sc application of CaNa2EDTA on Zn, Cu and Mn metabolism were investigated in female dogs. CaNa2EDTA was applied with a dose of 280 mg/kg bw/ every 6 hours for 54 h. Urine was collected every 6 h and the Zn, Cu and Mn content analysed. CaNa2EDTA application increased the urinary excretion of Zn, Cu and Mn significantly (Ibim 1992).

In addition in poorly documented studies by Yang (1964) the toxicokinetics of Na2EDTA were analysed in rats. After gavage application of 47.5, 95 and 142.5 mg/kg bw the amount of EDTA ingested was proportional to the amount of urinary excretion with a peak excretion 4 h after application. After gavage administration of 400 mg/kg bw to weanling and adult rats roughly 90% of the dose were recovered in feces, while only 5.3 % (adults) - 8.6 % (weanlings) were recovered in urine within 48 h. It was therefore assumed that most of the orally applied EDTA is not absorbed. After a single gavage application of ca. 475 mg/kg bw to rats, the gastrointestinal tract was removed in intervals up to 32 h and the EDTA content analyzed. All EDTA passed through the stomach within 12 h and 93% of the dose was recovered in the colon after 32 h, which demonstrated a poor absorption from GI tract. The EDTA contents of the small intestine and urine reached a maximum about 4 h after dosage. Urinary excretion over the period of 32 h cumulated to 6% of the dose. In additional study Yang stated that of a dietary dose of 300, 600 and 3000 mg/kg bw 82%, 44% and 45% could be recovered in urine and feces. However, it is unclear were the residual percentage of Na2EDTA remained.

Foreman and Trujillo (1954) studied the toxicokinetics of 14C-CaNa2EDTA in young, healthy male volunteers. 4.2 mg/person were applied i.v. or i.m. 50% of the dose was excreted in urine within 1 h (i.v.) or 2.5 h (i.m.). Within 24 h >98% of the dose were excreted in urine after both applications. The half-live blood clearance was 1 h 5 min (i.v.) or 1.5 h (i.m.) respectively. Additionally Foreman and Trujillo administered 14C-CaNa2EDTA orally at a dose of 1.5 mg/person. CaNa2EDTA was poorly absorbed from gastrointestinal tract. Within 72 h 91% of the dose was excreted in feces and 4.2% in urine. No test substance could be detected in blood.

Discussion on absorption rate:

In a study on young, healthy, male volunteers Foreman and Trujillo (1954) investigated the dermal absorption of CaNa2EDTA. 3 mg of a mixture of 14C labeled and unlabeled substance was prepared in water soluble base. The past was applied over an area of 100 cm2 for 24 h under occlusive conditions. In one study Na salt was used instead of Ca salt. The maximum activity in the urine was 0.001% of the administered dose.