Sodium hydrogen [N,N-bis[2-[bis(carboxymethyl)amino]ethyl]glycinato(5-)]ferrate(2-)

Endpoint summary

Description of key information

The oral LD50 value is in excess of 2000 mg/kg bw; the 4-h LC50 value is in excess of 5.08 g/m3. No acute dermal toxicity studies are available but based on read across (see also section 13), acute dermal toxicity following exposure to DTPA-FeHNa is not expected.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

October 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Well performed and reported GLP study

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

Source: Charles River Deutschland, Sulzfeld, Germany.
Number of animals: 6 Females (nulliparous and non-pregnant). Each dose group consisted of 3 animals.
Age and body weight: young adult animals (approx. 8 weeks old) were selected. Body weight variation did not exceed +/- 20% of the sex mean.
Identification: ear- and tailmark

Conditions: Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, approximately 15 room air changes/hour, and a 12-hour light/12-hour dark cycle. Any variations to these conditions were recorded in the raw data and were considered to have had no effect on the outcome of the study.

Accommodation: Group housing of 3 animals per cage in labeled Makrolon cages (MIV type; height 18 cm.) containing sterilized sawdust as bedding material (Litalabo, S.P.P.S., Argenteuil, France) and paper as cage-enrichment (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom).
Acclimatization period was at least 5 days before start of treatment under laboratory conditions.

Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).

Water: Free access to tap water.

Diet, water, bedding and cage enrichment evaluation for contaminants and/or nutrients was performed according to facility standard procedures.

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

Rationale: the vehicle was selected based on trial formulations performed at WIL Research Europe and on test substance data supplied by the sponsor.
Preparation: the formulations (w/w) were prepared within 4 hours prior to dosing. The (test substance) formulations were protected from light using amber coloured glassware. Homogeneity was accomplished to a visually acceptable level. No correction was made for purity of the test substance.
Method: oral gavage, using plastic feeding tubes.
Fasting: animals were deprived of food overnight prior to dosing and until 3-4 hours after administration of the test substance. Water was available ad libitum.
Frequency: single dosage on Day 1.
Dose level (volume): 2000 mg/kg (20 mL/kg) body weight.

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

6

Control animals:

no

Details on study design:

Mortality/Viability: twice daily.
Body weights: Days 1 (pre-administration), 8 and 15.
Clinical signs: at periodic intervals on the day of dosing (Day 1) and once daily thereafter, until Day 15. The symptoms were graded according to fixed scales and the time of onset, degree and duration were recorded:
Maximum grade 4: grading slight (1) to very severe (4)
Maximum grade 3: grading slight (1) to severe (3)
Maximum grade 1: presence is scored (1).
Necropsy: at the end of the observation period, all animals were sacrificed by oxygen/carbon dioxide procedure and subjected to necropsy. Descriptions of all internal macroscopic abnormalities were recorded.

Statistics:

Not needed, limit test

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occurred.

Clinical signs:

other: Piloerection and/or hunched posture was noted among most animals on Day 1. In addition, brown staining was observed on the back of three animals on Days 2-4.

Gross pathology:

Macroscopic examination did not reveal any toxicologically relevant abnormalities.
Pelvic dilation found in three animals at macroscopic post mortem examination is commonly seen among rats of this age and strain and was therefore considered not toxicologically significant.

Other findings:

None

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The oral LD50 was in excess of 2000 mg/kg bw.

Executive summary:

The study was carried out based on the guidelines described in: OECD No.423 (2001) "Acute Oral Toxicity, Acute Toxic Class Method"; Commission Regulation (EC) No 440/2008, B1 tris: "Acute Oral Toxicity, Acute Toxic Class Method"; EPA, OPPTS 870.1100 (2002), "Acute Oral Toxicity"

JMAFF guidelines (2011) including the most recent partial revisions.

DTPA-FeHNa was administered by oral gavage to two subsequent groups of three female Wistar rats at 2000 mg/kg body weight. Animals were subjected to daily observations and weekly determination of body weight. Macroscopic examination was performed after terminal sacrifice (Day 15).

No mortality occurred. Piloerection and/or hunched posture was noted among most animals on Day 1. In addition, brown staining was observed on the back of three animals on Days 2-4. The body weight gain shown by the animals over the study period was considered to be normal. Macroscopic examination did not reveal any toxicologically relevant abnormalities.

The oral LD50value of DTPA-FeHNa in Wistar rats was established to exceed 2000 mg/kg body weight. According to the OECD 423 test guideline, the LD50 cut-off value was considered to exceed 5000 mg/kg body weight.

Based on these results, DTPA-FeHNa does not have to be classified and has no obligatory labelling requirement for acute oral toxicity according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2011) andRegulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

One well performed study; result comparable to other metal chelates.

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

July-August 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Well performed and reported GLP study

Qualifier:

according to guideline

Guideline:

OECD Guideline 436 (Acute Inhalation Toxicity: Acute Toxic Class Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

yes

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

On the exposure day, the rats were about 11 weeks old
Their body weight variation did not exceed ± 20% of the mean weight for each sex.
Mean body weight just before exposure on day 0 was 352 and 235 g for males and females, respectively.
Duration of the acclimatization period: 26 days.

Housing: under conventional conditions in one room separated by sex.
Temperature and relative humidity in the animal room: 22 ± 2°C and at 45-65%, respectively.
Occasionally, the relative humidity briefly (about two hours or less) exceeded 65% after cleaning activities or filling up drinking bottles in the animal room (highest value recorded: 95.5%). In addition, the relative humidity was higher than 65% during three longer periods due to unknown
causes (these periods lasted about 4, 10 or 16 hours and occurred on 18-20 August 2012; the highest value recorded was 85.6%).
Lighting: artificial (fluorescent tubes) with a sequence of 12 hours light and 12 hours dark.
Number of air changes: about 10 per hour.

Caging: except during exposure, the animals were housed in groups of three, separated by sex , in macrolon cages (type 4S) with a bedding of wood shavings (Lignocel, Type ¾ from Rettenmaier, Rosenberg, Germany) and a wooden block (from ABEDD, Austria) and strips of paper (Enviro-dri obtained through Lillico, Betchworth, England) as environmental enrichment.
During exposure, the animals were housed individually in the exposure unit.
Immediately after exposure, the animals were returned to their home cages.

Feed and drinking water were provided ad libitum from the arrival of the animals until the end of the study, except during exposure when they had no access to feed or water. The animals received a cereal-based (closed formula) rodent diet.

Route of administration:

inhalation: dust

Type of inhalation exposure:

nose/head only

Vehicle:

air

Details on inhalation exposure:

The animals were exposed to the test atmosphere in a nose-only inhalation chamber. The column had a volume of about 50 litres and consisted of a top assembly with one mixing chamber, two rodent tube sections and at the bottom the base assembly with the exhaust port. Empty ports were used for test atmosphere sampling (for analysis of the actual concentration and particle size distribution) and measurement of oxygen concentration, temperature and relative humidity. The animals were secured in plastic animal holders, positioned radially through the outer cylinder around the central column. By securing a positive pressure in the central column and a slightly negative pressure in the outer cylinder which encloses the entire animal holder, dilution of test atmosphere by air leaking from the animal’s thorax to the nose was avoided. The unit was illuminated externally by normal laboratory fluorescent tube lighting.The total air flow through the unit was at least 1 litre/min for each rat. The air entering the unit was controlled at 22 ± 2˚C and the relative humidity was maintained between 30 and 70%.

The test material was milled using a ball mill fitted with a zirconium beaker and 100 zirconium balls with a diameter of 10 mm. Milling of test material for exposure of the animals was conducted on 9 August 2012.

The inhalation equipment was designed to expose rats to a continuous supply of fresh test atmosphere. The milled test material was aerosolized using a turntable dust feeder and an eductor. The eductor was supplied with humidified compressed air and operated at a pressure of 1.0 kg/cm2. The total air flow consists of the flow of the air driving the eductor and the air flow added by suction. Since the ratio of air pressure driving the eductor and the total flow is constant, the driving air pressure was recorded at regular intervals during exposure (approximately each half hour) and used to monitor the total air flow. At the driving air pressure used, the total air flow was determined to be 87.6 L/min. The test atmosphere was introduced at the top of the exposure chamber, directed downward and led to the noses of the animals. At the bottom of the unit the test atmosphere was exhausted.

The animals were placed in the exposure unit after stabilization of the test atmosphere (T95 was about 2 minutes). The period between the start of the generation of the test atmosphere and the start of exposure of the animals was 56 minutes.

The actual concentration of the test material in the test atmosphere was measured by gravimetric analysis. Representative test atmosphere samples were obtained from the animals’ breathing zone by passing approximately 2.5 L test atmosphere at 5 L/min through fibre glass filters (Sartorius, 13400-47). Filters were weighed before sampling, loaded with a sample of test atmosphere, and weighed again. The actual concentration was calculated by dividing the amount of test material present on the filter by the volume of the sample taken. A total number of 12 samples were taken
during exposure of the animals.
The nominal concentration was determined by dividing the total amount of test material used (by weight) by the total volume of air passed through the exposure unit. The generation efficiency was calculated from the actual and the nominal concentration (efficiency = actual concentration as percentage of nominal concentration).
Particle size distribution measurements were carried out by means of a 10-stage cascade impactor (Sierra instruments, Carmel Valley, California, USA). During exposure of the animals, two measurements were made. The Mass Median Aerodynamic Diameter (MMAD) and geometric standard deviation (gsd) were calculated.

The total air flow, temperature and the relative humidity of the test atmosphere were recorded eight times during exposure (about twice per hour). The driving air pressure of the eductor was used to monitor the total air flow. The temperature and relative humidity were measured by means of a RH/T device (TESTO 635, TESTO GmbH & Co, Lenzkirch, Schwarzwald, Germany). The oxygen concentration was checked once during exposure (Oxygen analyser type PMA-10, M&C Products Analysentechnik GmbH, Ratingen-Lintorf, Germany). The carbon dioxide concentration was calculated.

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

4 h

Concentrations:

5.08 +/- 0.20 g/m3 (mean ± sd; n=12)

No. of animals per sex per dose:

3 males and 3 females

Control animals:

no

Details on study design:

On the exposure day, the animals were observed for clinical signs just before exposure, four times during exposure (about once per hour, starting ten minutes after initiation of treatment), and twice after exposure. During the observation period, each animal was observed daily in the morning hours by cage-side observations and, if necessary, handled to detect signs of toxicity. On working days, all cages were checked again in the afternoon. In weekends only one check per day was carried out. During exposure, attention was directed to breathing abnormalities and restlessness. All abnormalities, signs of ill health, and reactions to treatment were recorded.

The body weight of each animal was recorded on days -1, 0 (just before exposure), 1, 3, 7 and 14 (prior to necropsy). The weight changes over adjacent days and over the 14-day observation period were calculated from the body weight data.

At the end of the 14-day observation period, the animals were sacrificed by exsanguination from the abdominal aorta under pentobarbital anaesthesia. At necropsy, abdominal and thoracic organs were examined in situ for gross pathological changes, with particular attention to any changes in the respiratory tract. After the post-mortem examination, the carcasses were discarded.

Statistics:

Not applicable (limit test)

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.08 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Mortality:

All animals survived until scheduled termination.

Clinical signs:

other: Clinical observations during exposure revealed clear restlessness and decreased breathing rate in all animals. These symptoms occurred from about one hour after initiation of exposure and aggravated in the course of the exposure period. Clinical signs aft

Body weight:

One day after exposure, all males and one female had lost weight, which may partly be explained by the restraint during exposure. Thereafter, body weight developed normally, except in the female that initially lost weight (this female hardly grew after exposure).

Gross pathology:

Macroscopic abnormalities observed at necropsy were limited to a white patch on the left lung lobe in one male and a swollen uterus and cervix in one female.

Other findings:

No

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The 4-h LC50 is in excess of 5.08 mg/L.

Executive summary:

To examine the acute inhalation toxicity of DTPA-FeNaH, three male and three female rats were exposed for four hours to a target limit concentration of 5 g/m3 DTPA-FeNaH. Thereafter, the animals were kept for an observation period of 14 days. To detect adverse effects, clinical observations were made during and after exposure, body weight was measured before exposure (day 0) and at days 1, 3, 7 and 14, and macroscopic examination was conducted at the end of the observation period. The actual concentration of the test material during exposure was 5.08 ± 0.20 g/m3 (mean ± sd; n=12) based on gravimetric analysis of the test atmosphere. The mass median aerodynamic diameter (MMAD) of the aerosol was 4.00 and 3.54 μm (duplicate measurements) and the distribution of particle sizes had a geometric standard deviation (gsd) of 2.04 and 2.03, respectively. All animals survived until scheduled termination. Clinical observations during exposure revealed clear restlessness and decreased breathing rate in all animals. These symptoms occurred from about one hour after initiation of exposure and aggravated in the course of the exposure period. Clinical signs after exposure were limited to blepharospasm (in one male shortly after exposure) and piloerection (in two males and one female shortly after exposure, and in all animals about two hours after exposure). One day after exposure, all males and one female had lost weight, which may partly be explained by the restraint during exposure. Thereafter, body weight developed normally, except in the female that initially lost weight (this female hardly grew after exposure). Macroscopic abnormalities observed at necropsy were limited to a white patch on the left lung lobe in one male and a swollen uterus and cervix in one female. Conclusion Since none of the animals died during the 14-day observation period following a 4- hour exposure to DTPA-FeNaH, it was concluded that the 4-hour LC50 in rats is above 5.08 g/m3.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Quality of whole database:

One well performed study; result comparable to other metal chelates.

Acute toxicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Justification for selection of acute toxicity – oral endpoint

Well performed and reported GLP study

Justification for selection of acute toxicity – inhalation endpoint

Well performed and reported GLP study

Justification for classification or non-classification

No classification needed for acute oral and inhalation exposure as the oral LD50 value is in excess of 2000 mg/kg bw, and the 4-h LC50 value is in excess of 5.08 g/m3. Based on the expec ted absence of acute dermal toxicity, no classification needed for acute dermal toxicity.