Sodium [[α,α'-(ethylenediimino)bis[2-hydroxybenzene-1-acetato]](4-)]ferrate(1-)

Endpoint summary

Description of key information

In a key subacute 28-day dermal toxicity study conducted with a source substance Fe(Na)EDDHA, the NOEL was established at 100 mg/kg bw/day based on slight effects on the liver and skin and due to increased adrenal weight noted at the high dose level of 1000 mg/kg bw/day.

No data on repeated inhalation exposure are required. Exposure by the inhalation route is considered to be negligible.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1997-01-30 to 1998-06-18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

other: Sprague-Dawley derived; Tif:RAIf (SPF); hybrids of RII/1 x RII/2

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Animal Production, Novartis Pharma AG, 4332 Stein, Switzerland
- Age at start of acclimation period: approximately 5 weeks old
- Weight range at acclimation period: 149.5-185.8 g (males), 134.7-165.6 g (females)
- Housing: in groups of 5 animals, in macrolon cages type 4 with wire mesh tops and standardised granulated soft wood bedding
- Diet: pelleted, certified standard diet (Nafag No. 8900 FOR GLP), provided ad libitum (exception: food was withheld overnight prior to blood collection)
- Water: tap water, ad libitum
- Acclimation period: 11 days

ENVIRONMENTAL CONDITIONS
- Temperature: 22 ± 2 °C
- Humidity: 55 ± 10 %
- Air changes: 16-20 air changes/hour
- Photoperiod: 12 hours dark / 12 hours light

Route of administration:

oral: gavage

Vehicle:

other: 0.5 % CMC and 0.1 % Tween 80 in distilled water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
Suspensions of the test article in the vehicle at the appropriate concentrations were freshly prepared every day immediately prior to the dosing of the animals and administered within about 2 hours.

VEHICLE
- Justification for use and choice of vehicle: standard procedure
- Amount of vehicle: 10 mL/kg bw of suspension were applied

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

1.Analyses of the test article in the vehicle were performed in a previously conducted study (941102) at the analytical laboratories of RCC Umweltchemie AG, 4452 Itingen / Switzerland. In the tested range (0.1 - 100 mg/mL), homogeneous distribution of the test article in the vehicle was recorded. The content of the test article in the vehicle was in agreement with the nominal concentrations. CGA 65047 F (A-5787 A) was stable in the vehicle over a period of 4 hours at room temperature.
2. Control analyses of the test article in the vehicle were carried out at all dose levels on samples collected once per experimental weeks 1, 4, 8 and 13. Samples were analysed by RCC Umweltchemie AG, 4452 Itingen, Switzerland (RCC Project Number 651688). The analytical procedure for content determination yielded values close to the targeted concentrations and within the limits of acceptance.

Duration of treatment / exposure:

90 days

Frequency of treatment:

once per day, 7 times per week

Remarks:

Doses / Concentrations:
5, 50 and 200 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

Main Experiment: 10 animals
Recovery: 10 animals (control and high dose group only)

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: dose levels were selected based on the results of a preceding 28-day dose range finding toxicity study, project no. 941102 (CIBA-GEIGY, 1996):
The treatment with CGA 65047 F (A-5787 A) at doses of 0, 50, 2 00 and 1000 mg/kg body weight by daily gavage resulted in impaired body weight development of rats treated at 200 and 1000 mg/kg body weight and correspondent lower food intake. Animals treated at 1000 mg/kg body weight consumed markedly more water. All animals did not show signs of overt toxicity and survived to scheduled sacrifice. Laboratory investigations revealed an anemia at 200 and 1000 mg/kg body weight without erythropoietic response. At the 1000 and 200 mg/kg dose levels, the kidney was revealed as target organ by histopathological examination, by blood chemistry data and by organ weight evaluation. In addition, organ weight evaluation showed changes in relative weights of heart (males at 200 and males and females at 1000 mg/kg), adrenals (males at 50, 200 and 1000 mg/kg), and spleen (females at 1000 mg/kg). However, in the absence of corroborative findings, the experimental relevance of these findings was considered equivocal.
- Rationale for animal assignment (if not random): computer-generated random numbers
- Rationale for selecting satellite groups: to investigate reversibility of the effects.
- Post-exposure recovery period in satellite groups: 4 weeks

Positive control:

no

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS:
- Time schedule: twice daily for mortality and once daily for general observations.

DETAILED CLINICAL OBSERVATIONS:
- Time schedule: at least weekly

BODY WEIGHT:
- Time schedule: once weekly

FOOD CONSUMPTION:
- Food consumption was determined cagewise on a weekly basis and mean daily diet consumption calculated as g food/kg bw/day.

WATER CONSUMPTION
- Time schedule for examinations: weekly, determined cagewise

OPHTHALMOSCOPIC EXAMINATION:
- Time schedule for examinations: during the acclimation period (all animals), at the end of the treatment and recovery periods (control and high dose group only)
- Dose groups that were examined: all animals during the acclimation period, only high dose and control group animals on day 87 (treatment period) and day 115 (recovery period)

HAEMATOLOGY:
- Time schedule for collection of blood: at the end of the treatment and recovery periods
- Anaesthetic used for blood collection: ether
- Animals fasted: food was withheld overnight prior to blood sampling
- How many animals: all animals
- Parameters examined: erythrocyte count, haematocrit, mean corpuscular volume, red cell volume distribution width, haemoglobin concentration, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, haemoglobin concentration distribution width, prothrombin time, methaemoglobin concentration and leukocyte, neutrophil, eosinophil, basophil, lymphocyte, monocyte, large unstained cells, reticulocyte and thrombocyte counts

CLINICAL CHEMISTRY:
- Time schedule for collection of blood: at the end of the treatment and recovery periods
- Animals fasted: food was withheld overnight prior to blood sampling
- How many animals: all animals
- Parameters examined: glucose, urea, creatinine, total bilirubin, total protein, albumin, globulin, cholesterol, triglycerides, sodium, potassium, calcium, chloride and inorganic phosphorus concentration, and A/G ratio and aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase and gamma-glutamyl transpeptidase activity

URINALYSIS:
- Time schedule for collection of urine: at the end of the treatment and recovery periods
- Metabolism cages were used for collection of urine
- Animals fasted: food was withheld overnight prior to blood sampling
- Parameters examined: urine volume, relative density, pH-value, urine colour and protein, glucose, ketone, bilirubin, erythrocyte, leukocyte and urobilinogen content

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY:
The following organs or tissues were examined microscopically:
spleen, mesenteric lymph node, axillary lymph node, femur with joint, bone marrow (femur), trachea, lung, heart, aorta, submandibular salivary gland, liver, pancreas, oesophagus, stomach, small intestine (duodenum, ileum, jejunum), large intestine (caecum, colon, rectum), kidney, urinary bladder, testis, epididymis, uterus, vagina, ovary, adrenal gland, thyroid with parathyroid gland, thymus, peripheral nerve (sciatic nerve), brain (including medulla, pons, cerebral and cerebellar cortex) and any organ with gross lesions.

Other examinations:

ORGAN WEIGHT
The weights of the following organs were determined at necropsy:
brain, heart, liver, kidneys, adrenals, thymus, ovaries or testes, spleen and thyroid

Statistics:

Each treated group was compared to the control group either by Lepage's or Wilcoxon's two-sample test and tested for increasing or decreasing trends from control up to the respective dose group by Jonckheere's test for ordered alternatives.

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Food efficiency:

not examined

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

no effects observed

Ophthalmological findings:

no effects observed

Haematological findings:

effects observed, treatment-related

Clinical biochemistry findings:

effects observed, treatment-related

Urinalysis findings:

effects observed, treatment-related

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not specified

Details on results:

CLINICAL SIGNS AND MORTALITY
During this study neither changes to the behavior nor relevant clinical signs were observed.
There was no mortality during this study which was attributed to the treatment with the test article.
In the third treatment week one female animal (no. 86) died. However, there were no indications of a testarticle-related occurrence.

BODY WEIGHT AND WEIGHT GAIN
Decreased mean body weights were noted from weeks 5 and 8 onwards at 200 mg/kg bw/day in male and female rats, respectively. The mean body weight gains were decreased by the end of the treatment period in animals treated with 200 mg/kg bw/day. During the recovery period, body weight gain in animals previously treated with 200 mg/kg bw/day was higher as compared to controls.
The body weight development in the other treated groups was not influenced by treatment.

FOOD CONSUMPTION
Food consumption and food consumption ratios were decreased at 200 mg/kg bw/day in both sexes (decreased food consumption of 13% in males and of 8% in females, compared to the controls). During the recovery period food intake improved.
The mean food consumption of the other treated groups was not influenced by treatment.

WATER CONSUMPTION
The mean water consumption was not influenced by treatment.

OPHTHALMOSCOPIC EXAMINATION
The examinations of the eyes (lids and surrounds, conjunctiva, pupillary reflex, cornea, sclera, anterior chamber, lens, vitreous, and fundus) towards treatment end (day 87) and towards recovery end (day 115) did not reveal any treatment-related findings.

HAEMATOLOGY
At the end of the treatment period, a normochromic anaemia with lower erythrocyte count, haemoglobin concentration and haematocrit was observed in males and females at 200 mg/kg bw/day and males at 50 mg/kg bw/day. A higher reticulocyte count associated with higher MCV and MCH values and reduced white blood cell, basophil, lymphocyte and monocyte counts were confined to males at 200 mg/kg bw/day. A higher platelet count was recorded for males at 50 and 200 mg/kg bw/day and a higher prothrombin activity was recorded for males and females at 200 mg/kg bw/day. Evidence of reversibility for all the above parameters was apparent after the recovery period.

CLINICAL CHEMISTRY
Several clinical chemistry parameters including creatinine, urea, protein, globulin, cholesterol and sodium concentration were increased at 50 and/or 200 mg/kg bw/day. Lower potassium levels were noted in males at 200 mg/kg bw/day. All values were similar to the control group values after the 4-week recovery period.

URINALYSIS
Excretion of more acidic red-brown (males) or yellow-brown (females) discoloured urine was observed at 200 mg/kg bw/day. More acidic urine was also excreted by males at 50 mg/kg bw/day. By the end of the recovery period, the colour and pH of the urine excreted by male and females previously treated at 200 mg/kg bw/day was similar to that of control group animals.

ORGAN WEIGHTS
The mean carcass weights were decreased at the end of the treatment period at 200 mg/kg bw/day. Males at 200 mg/kg bw/day showed an elevated mean heart to body weight ratio which still was higher than the concurrent control value at the end of the recovery period.

GROSS PATHOLOGY and HISTOPATHOLOGY
There were no test item related findings.

Dose descriptor:

NOAEL

Effect level:

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

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: haematology parameters (anaemia) The NOAEL was estimated based on the LOAEL of 50 mg/kg bw/day applying an assessment factor of 5.

Critical effects observed:

not specified

CHEMICAL ANALYSIS OF DOSE FORMULATIONS

The calculated overall mean contents of the test item in the dose formulations used for the low, mid and high dose groups were 96.9, 104.2 and 101.0 % of the nominal concentrations, respectively (RCC project no. 651688). In a previous 28 -day dose range finding study with the same test item and vehicle, the stability and homogeneity of dose formulations were within the limits of acceptance (RCC project no. 393322).

Estimation of NOAEL:

A more realistic NOAEL was estimated from the LOAEL of 50 mg/kg bw/day applying an assessment factor of 5. This method is applicable and scientifically justified for this test, as only slight adverse effects were observed at 50 mg/kg bw/day (haematology parameters - anaemia). To take the relatively large concentration gap between 5 (clear NOEL) and 50 mg/kg bw/day into account, 5 mg/kg bw/day was not taken as NOAEL, but extrapolated form the LOAEL. The guidance on information requirements and CSA, R.8 (ECHA, 2008 -2010) recommends a factor between 3 and 10 for extrapolation from LOAEL to NOAEL. An assessment factor of 5 seems to be approprate and conservative enough for the current study as only slight effects were observed at the LOAEL of 50 mg/kg bw/day. Consequently a NOAEL of 10 mg/kg bw/day is calculated for this study and used for DNEL and PNEC(oral) derivation.

Conclusions:

NOEL of 5 mg/kg bw was established in this study (for males and females). NOAEL of 10 mg/kg bw was derived based on LOAEL of 50 mg/kg bw.

Executive summary:

In a subchronic toxicity study (Novartis Crop Protection AG, 1998), FeNaEDDHA in 0.5 % CMC and 0.1 % Tween 80 in distilled water was administered to 10 Sprague-Dawley derived rats/sex/dose level by oral gavage (10 mL/kg bw) at dose levels of 5, 50 or 200 mg/kg bw/day for a period of 90 days. Male and female animals of the concurrent control group were treated with the vehicle only. Additional 10 animals/sex of the high dose and control group were kept on control diet for a 4 -week recovery period before sacrifice. Treatment with the test item resulted in lower food intake and impaired body weight development of rats treated at 200 mg/kg bw/day. Reversible effects on the red blood cell (normochromic anaemia) and white blood cell parameters, and higher values of platelets and prothrombin activity were noted at 50 and/or 200 mg/kg bw/day. In addition, there were changes of blood chemistry and urine parameters concerning the liver and kidneys. The body weight relative heart weight was increased in males at 200 mg/kg bw/day. Under the conditions of this study, the NOAEL for FeNaEDDHA when administered by daily oral gavage for three months was 10 mg/kg bw/day (estimated from the LOAEL).

This subchronic toxicity study in the rat is acceptable and satisfies the guideline requirement for a subchronic oral study (OECD 408).

Estimation of NOAEL:

A more realistic NOAEL was estimated from the LOAEL of 50 mg/kg bw/day applying an assessment factor of 5. This method is applicable and scientifically justified for this test, as only slight adverse effects were observed at 50 mg/kg bw/day (haematology parameters - anaemia). To take the relatively large concentration gap between 5 (clear NOEL) and 50 mg/kg bw/day into account, 5 mg/kg bw/day was not taken as NOAEL, but extrapolated form the LOAEL. The guidance on information requirements and CSA, R.8 (ECHA, 2008 -2010) recommends a factor between 3 and 10 for extrapolation from LOAEL to NOAEL. An assessment factor of 5 seems to be approprate and conservative enough for the current study as only slight effects were observed at the LOAEL of 50 mg/kg bw/day. Consequently a NOAEL of 10 mg/kg bw/day is calculated for this study.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

10 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

There is no deficiencies in the available data set. The data on structurally related substances is reliable and consisitent.

System:

haematopoietic

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

100 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

The dermal study available meets required adequacy criteria and herewith demonstrating a good quality of data base

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

Study duration:

subacute

Species:

rat

Quality of whole database:

The dermal study available meets required adequacy criteria and herewith demonstrating a good quality of data base

Additional information

No repeated dose toxicity studies are available for the target substance o-o-EDDHA (CAS 16455 -61 -1). The data on the structurally related substance Fe(Na)EDDHA (CAS 84539 -55 -9) has been used to assess toxicity potential of o-o-EDDHA (CAS 16455 -61 -1) at repeated exposures.

Oral route

The subchronic toxicity of Fe(Na)EDDHA (CAS 84539 -55 -9) by oral route was investigated in rats (Novartis Crop Protection AG, 1998). The test item was administered to 10 Sprague-Dawley derived rats/sex/dose level by oral gavage at 5, 50 or 200 mg/kg bw/day for 90 days. A concurrent control group was treated with the vehicle only. Additional 10 animals/sex of the high dose and control group were kept on control diet for a 4 -week recovery period before sacrifice. Treatment with the test item resulted in lower food intake and impaired body weight development at 200 mg/kg bw/day. Reversible effects on red blood cell (normochromic anaemia) and white blood cell parameters, and higher values of platelets and prothrombin activity were noted at 50 and/or 200 mg/kg bw/day. In addition, changes of blood chemistry and urine parameters concerning the liver and kidneys were noted. The body weight relative heart weight was increased in males at 200 mg/kg bw/day. Under the conditions of this study, the NOAEL for Fe(Na)EDDHA when administered daily by oral gavage for three months was 10 mg/kg bw/day (estimated from the LOAEL).

Estimation of NOAEL:

A more realistic NOAEL was estimated from the LOAEL of 50 mg/kg bw/day applying an assessment factor of 5. This method is applicable and scientifically justified for this test, as only slight adverse effects were observed at 50 mg/kg bw/day (haematology parameters - anaemia). To take the relatively large concentration gap between 5 (clear NOEL) and 50 mg/kg bw/day into account, 5 mg/kg bw/day was not taken as NOAEL, but extrapolated from the LOAEL. The guidance on information requirements and CSA, R.8 (ECHA, 2008 -2010) recommends a factor between 3 and 10 for extrapolation from LOAEL to NOAEL. An assessment factor of 5 seems to be appropriate and conservative enough for the current study as only slight effects were observed at the LOAEL of 50 mg/kg bw/day. Consequently a NOAEL of 10 mg/kg bw/day is calculated for this study.

In a dose range-finding subacute oral toxicity study (CIBA-GEIGY Limited, 1996a), Fe(Na)EDDHA was administered to 5 Sprague-Dawley derived rats/sex/dose level by oral gavage at 50, 200 or 1000 mg/kg bw/day for 28 days. A concurrent control group was treated with the vehicle only. Treatment with the test item resulted in impaired body weight development at 200 and 1000 mg/kg bw/day and correspondend lower food intake. Anaemia without erythropoietic response was noted at 200 and 1000 mg/kg bw/day. At the same dose levels, the kidney was revealed as target organ by microscopical examination, by blood chemistry data evaluation and by organ weight evaluation. In addition, body weight relative organ weight changes were noted in the heart, adrenals and spleen. However, the relevance of these findings was considered as equivocal. This study was used as scientific basis for dose level selection for the above-mentioned 90 -day repeated dose oral toxicity study in the rat.

Dermal route

In a repeated dose dermal toxicity study (CIBA-GEIGY Limited, 1996b), Fe(Na)EDDHA was administered to the skin of 5 Sprague-Dawley derived rats/sex/dose level at 10, 100 or 1000 mg/kg bw/day for 28 days (5 days/week). A concurrent control group was treated with the vehicle only. Dermal treatment with the test item resulted in no mortality, no relevant clinical signs, and no changes in food consumption, no effects on haematology and clinical chemistry parameters and no gross findings. A transient slight body weight loss was noted in females at 1000 mg/kg bw/day during the first week of treatment. There was an increase in adrenal weight in males at 1000 mg/kg bw/day. Microscopically, the skin application sites of females at 1000 mg/kg bw/day revealed epidermal hyperkeratosis associated with an increased severity of acanthosis. In 2/5 males at 1000 mg/kg bw/day centrilobular hypertrophy of hepatocytes was noted. Based on the slight effects on the liver and skin and due to the increased adrenal weight noted at 1000 mg/kg bw/day, the NOEL was established at 100 mg/kg bw/day.

Inhalation route

In accordance with column 2 of REACH Annex IX, the test repeated dose toxicity after inhalation (required in section 8.6) does not need to be conducted as repeated dose toxicity studies for oral and dermal application are available. In addition, the substance showed only very low toxicity after acute inhalation exposure with an LD50 of greater than 4200 mg/m³ in rats.

Justification for classification or non-classification

Based on the results of the available repeated dose toxicity studies conducted with the source substance Fe(Na)EDDHA (CAS 84539 -55 -9), the criteria for STOT-RE are not fulfilled as no damage to organs was observed. Thus, the target substance o,o-Fe(Na)EDDHA is not subject to classification and labelling according to the Regulation No 1272/2008 (CLP).