Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2006-12-20 to 2007-01-29
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2007
Report Date:
2007

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 bis (Acute Oral Toxicity - Fixed Dose Procedure)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Statement of GLP Compliance No. G 024 (Slovak National Accreditation Service); Statement of GLP Compliance No. 4/2006/DPL.
Test type:
fixed dose procedure
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
other: water solution
Details on test material:
- Name of test material (as cited in study report): Gu (II) IDHA
- Substance type: chelate
- Physical state: solid (odourless, blue microgranules)
- Analytical purity: Cu2+ (9.95 %)
- Impurities (identity and concentrations): 4.41 % of N -NO3 (% m/m)
- Purity test date: CU-005/906/P from 2006-09-14
- Lot/batch No.: 906/P/Cu
Data concerning the identification, purity and durability of the analysed material are the responsibility of the Mandator.

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Institute of Occupational Medicine in Łódź, Poland
- Age at study initiation: Pilot study: 9 week (dose of 2,000 mg/kg bw); 11 week (dose of 300 mg/kg bw). Main experiment: 11 weeks (average body weight of 179 g).
- Weight at study initiation: Pilot study: 168 g (dose of 2,000 mg/kg bw); 198 g (female; dose of 300 mg/kg bw). Main experiment: average body weight of 179 g.
- Fasting period before study: yes. The day before the experiment was due to commence, some 18 hours before administration of the analysed material, the animals were deprived of feed, being left with only water. Feed was made available again 3 hours after administration of the analysed substance.
- Housing: The animals were kept in plastic cages with metal wire covers, with the following dimensions (length x width x height): 58 x 37 x 21 cm.
During the experiment, the animals were kept in cages individually (initial study) and in groups of four (main study).
- Diet (e.g. ad libitum): ad libitum (standard granulated "Murigran" laboratory feed, manufactured by Wytwórnia Koncentratów i Mieszanek Paszowych AGROPOL of Motycz
- Water (e.g. ad libitum): ad libitum (tap water)
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-21
- Humidity (%): 45-71
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES:
The analysed substance was administered to the rats on the following days: 20.12.2006. (1 female – initial experiment, dose of 2,000 mg/kg of body mass), 04.01.2007. (1 female – initial experiment, dose of 300 mg/kg of body mass), 09.01.2007. (4 females – experiment proper, dose of 300 mg/kg of body mass). The experiment was terminated on the following days: 20.12.2006. (female that died during the initial experiment, dose of 2,000 mg/kg of body mass), 18.01.2007. (female – initial experiment, dose of 300 mg/kg of body mass), 19.01.2007 (female that died during the experiment proper, dose of 300 mg/kg of body mass), 23.01.2007. (3 females – experiment proper, dose of 300 mg/kg of body mass), respectively.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
VEHICLE
- Concentration in vehicle: 1 ml of the water solution of the analysed substance contained: 400 mg of the substance (dose of 2,000 mg/kg of body mass); 60 mg of the substance (dose of 300 mg/kg of body mass).
- Amount of vehicle (if gavage): 0.5 mL per 100 g of the body weight.

- Rationale for the selection of the starting dose: During the initial experiment, one female received a dose of 2,000 mg/kg of body mass of the analysed substance. Since this female died, the analysed substance was then administered to another female, in a dose of 300 mg/kg of body mass. On the basis of the initial experiment, during the experiment proper the analysed substance was administered to four successive females in a dose of 300 mg/kg of body mass.
Doses:
Initial experiment: 2000 mg/kg bw, 300 mg/kg bw;
Main strudy: 300 mg/kg bw
No. of animals per sex per dose:
Initial experiment: 2000 mg/kg bw (one animal), 300 mg/kg bw (one animal);
Main strudy: four animals
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: An assessment of the general condition of the animals, i.e. observation of all animals in terms of incidence and mortality, was performed twice daily throughout the 14-day duration of the experiment. Detailed clinical observations were performed on the day of administration of the analysed substance (day 0), 10, 30 and 60 minutes after administration, and subsequently every hour over a period of 5 hours from the time of administration. On successive days of the 14-day period of the experiment – once daily.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight.

Results and discussion

Preliminary study:
Following the single administration of the analysed substance at the dose of 2,000 mg/kg of body weight to one female (initial experiment), on the same day there were observed changes in posture, the way of walking, kinetic activity and reactivity. The female died within the second hour of administration.
Following the administration of the analysed substance at the dose of 300 mg/kg of body weight to one female (initial experiment), no symptoms of toxicity were observed during the 14-day period of observation. The animal survived the 14-day period of observation.
Effect levels
Sex:
female
Dose descriptor:
LD50
Effect level:
> 300 - < 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: based on mortality, clinical signs and findings at necropsy at 2000 mg/kg bw
Mortality:
One female died between the ninth and tenth day of observation. The remaining females survived the 14-day observation period.
Clinical signs:
Following the single administration of the analysed substance at the dose of 300 mg/kg bw to four successive females (main experiment), no symptoms of toxicity were observed during the 14-day period of observation for three of the animals. On day seven and eight, one female displayed changes in posture, the way of walking, kinetic activity, which were accompanied by horripilation, quickened breathing, and a mucoid secretion from the nostrils. On the ninth day of observation, the female additionally displayed changes in reactivity, respiratory murmurs, and blood-coloured secretion from the nostrils. In all probability, these changes were not connected with the analysed material. The female died between the ninth and tenth day of observation. The remaining females survived the 14-day observation period.
Body weight:
No body weight increase disorders were observed. The exception was one female, which body weight decreased during the first week of the experiment.
Gross pathology:
The females that died (initial and main studies) and the remaining females that survived following the 14-day period of observation underwent autopsies and macroscopic studies. The macroscopic study of the female that died having received a dose of 2,000 mg/kg bw disclosed hepatic hyperaemia, as well as pale and enlarged lungs. No changes were found during the macroscopic study in the female that died having received a dose of 300 mg/kg bw (the animal was devoured by the other animals present in the cage). Macroscopic studies did not find any pathological changes in the remaining animals.

Any other information on results incl. tables

Table 1. Clinical signs.

Dose(mg/kg of body mass)

Day following administration

Number of live animals

Rat no.

1*

2

3

4

5

2,000

0

1

2

3

4

5

6

7

8

9

10

11

12

13

14

1

0

0

0

0

0

0

0

0

0

0

0

0

0

0

OK, P

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

-

300

0

1

2

3

4

5

6

7

8

9

10

11

12

13

14

5

5

5

5

5

5

5

5

5

5

4

4

4

4

4

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

BZ

OK

OK

OK

P

-

-

-

-

* females from the initial experiment

BZ = without change

OK = clinical symptoms observed

P = died

Table 2. Body weights

Dose

mg/kg of body mass

Rat no.

Day of experiment

Difference

14 – 0

0

7

14

2,000

1*

168

-

-

-

300

1*

2

3

4

5

198

179

174

189

174

233

206

189

219

155

241

197

196

221

-

43

18

22

32

-

* females from the initial experiment

Applicant's summary and conclusion

Interpretation of results:
Toxicity Category IV
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Based on the mortality, clinical signs and findings at necropsy in one animal dosed by 2000 mg/kg bw, LD50 is considered to be between 300 - 2000 mg/kg bw that corresponds to Cat. 4 in accordance with the ECHA guidance on the Application of the CLP Criteria (2013).
Executive summary:

A study was conducted to test oral toxicity potential of Cu (II) IDHA in rats. Following the single administration of the analysed substance at dose level of 2000 mg/kg bw to a single female Wistar rat symptoms of toxicity were observed within the first hour of administration. The female died within the second hour of administration of the analysed material. Following the single administration of the analysed substance at dose level of 300 mg/kg bw to a single female, no symptoms of toxicity were observed during the 14-day period of observation. The female survived the 14-day period of observation. Based on this result, the test substance at the dose level of 300 mg/kg bw was administered to four successive females. No symptoms of toxicity were observed during the 14-day period of observation for three of the animals. In one animal, symptoms of toxicity were observed from the seventh day of observation. In all probability, these changes were not connected with the analysed material. The female died on the tenth day of observation. The remaining females survived the 14-day observation period.

The females that died (initial and main studies) and the remaining females that survived the 14-day period of observation underwent autopsies and macroscopic studies.

The macroscopic study of the female that died having received the dose of 2000 mg/kg bw disclosed hepatic hyperaemia, as well as pale and enlarged lungs. No changes were found during the macroscopic study in the female that died having received the dose of 300 mg/kg bw (the animal was devoured by the other animals present in the cage). Macroscopic studies did not find any pathological changes in the remaining animals.