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Toxicological information

Acute Toxicity: dermal

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Administrative data

Endpoint:
acute toxicity: dermal
Type of information:
experimental study
Adequacy of study:
key study
Study period:
11 March - 22 April 1998
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP/Guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1998
Report Date:
1990

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 402 (Acute Dermal Toxicity)
Deviations:
not specified
Qualifier:
according to
Guideline:
EU Method B.3 (Acute Toxicity (Dermal))
Deviations:
not specified
GLP compliance:
yes
Test type:
standard acute method
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
BTMAC-60%, a 60% aqueous solution, was used. No additional information provided.

Test animals

Species:
rabbit
Strain:
New Zealand White
Sex:
male/female
Details on test animals and environmental conditions:
Albino rabbit, New Zealand White, (SPF-Quality), from Broekman Institute, Someren, the Netherlands were used. Young adult animals were used with a weight range of 1.9-2.6 kg. Body weight variation did not exceed +/- 20% of the sex mean.

Conditions
Air-conditioned room with approximately 15 air changes per hour and the environment controlled with optimal conditions considered as being a temperature of 21C and a relative humidity of 50%. Fluctuations from these optimal conditions were noted, but were considered not to have affected study integrity. Lighting was 12 hours artificial fluorescent light and 12 hours dark per day.

Accommodation
Individually housed in labelled cages with perforated floors (Scanbur, Denmark) and equipped with an automatic drinking system (ITL, Bergen, the Netherlands). The acclimatisation period was at least 5 days before start of treatment under laboratory conditions.

Diet
Standard laboratory rabbit diet (LKK-20, pellet diameter 4 mm, Hope Farms, Woerden, the Netherlands) approx. 100 g. per day. Certificates of analysis were examined and then retained in the NOTOX archives.
In addition, hay (dried and compressed; BMI, Helmond, the Netherlands) was provided once a week.

Water
Free access to tap-water, diluted with decalcified water. Certificates of quarterly analysis were examined and then retained in the NOTOX archives.

Administration / exposure

Type of coverage:
semiocclusive
Vehicle:
unchanged (no vehicle)
Details on dermal exposure:
One day before exposure (day -1) an area of approximately 5 x 7 cm on the back of the animal was clipped.

The test formulation was applied in an area of approx. 10% of the total body surface, i.e., approximately 150 cm2. The test substance was held in contact with the skin with a dressing, consisting of a Metalline patch (10 x 15 cm) mounted on Medical tape, which was held in place with Micropore tape and subsequently Coban elastic bandage.

During the exposure period the dressing of the animals was checked periodically.
Duration of exposure:
24 hours, after which dressings were removed and residual test substance removed using a tissue moistened with tap water.
Doses:
2000 mg/kg (5 ml/kg) body weight
650 mg/kg (5 ml/kg) body weight
400 mg/kg (5 ml/kg) body weight
200 mg/kg (5 ml/kg) body weight
No. of animals per sex per dose:
3 males and 3 females/dose level
Control animals:
not specified
Details on study design:
After animals were dosed with the test material, the dressings were removed after 24 hours.

Observations
Animals were examined twice daily for mortality/viability. The time of death was recorded as precisely as possible. Body weights were recorded on days 1 (pre-administration), 8 and 15 and at death (if found dead after day 1).

At periodic intervals on the day of dosing (day 1) and once daily thereafter, until day 15 the animals were examined for clinical signs. The time of onset, degree and duration were recorded and the symptoms graded according to fixed scales:
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).

All animals killed for humane reasons and animals surviving to the end of the observation period (day 15) were sacrificed by intravenous injection with an overdose of pentobarbital (Euthensate). All animals assigned to the study were subjected to necropsy and descriptions of all internal macroscopic abnormalities recorded.
Statistics:
The LD50 value, the associated 95% confidence interval and the slope were calculated using Probit Analysis (SAS Technical Report: P-179, Additional SAS/STAT procedures, Release 6.03, Chapter 4, the PROBIT procedure).

Results and discussion

Effect levels
Sex:
male/female
Dose descriptor:
LD50
Effect level:
510 mg/kg bw
Based on:
dissolved
Remarks:
60% aqueous solution
Mortality:
See Table 1 below. The decedents were found dead on days 1 and 2. One male and two females dosed at 2000 mg/kg were killed in extremis on day 1.
Clinical signs:
Lethargy, flat posture, quick and slow breathing, (excessive) salivation, pale skin and diarrhea was noted among the animals that died.

Lethargy was noted in the surviving animals of the 650 and 400 mg/kg dose group on day 1 only.

Scales were seen in the treated skin-area among the animals treated at 400 mg/kg between days 5 and 8.
Body weight:
The changes noted in body weight gain in surviving males and females were within the range expected for rabbits used in this type of study and were therefore considered not indicative of toxicity.
Gross pathology:
Macroscopic post mortem examination of the animals that died or were killed in extremis during the study revealed hemorrhages or dark red foci in the thymus, thickening of the fundus in the stomach, irregular surface of the lungs and hemorrhagic fluid in the abdominal cavity. Dark red discoloration of the subcutis of the treated skin area was found in two females dosed at 2000 mg/kg. Macroscopic post mortem examination of the surviving animals at termination did not reveal any abnormalities.
Other findings:
No additional information available.

Any other information on results incl. tables

Initially, a group of three males and three females was dosed at 200 mg/kg body weight. Additional groups of 3 males and 3 females were selected in chronological order 2000, 650 and 400 mg/kg body weight.

Table 1 Mortality

 Dose level  Males  Females  Sexes Combined
 2000 mg/kg  3/3  3/3  6/6
 650 mg/kg  2/3  3/3  5/6
 400 mg/kg  0/3  1/3  1/6
 200 mg/kg  0/3  0/3  0/6

Applicant's summary and conclusion

Interpretation of results:
harmful
Remarks:
Migrated information Criteria used for interpretation of results: other: Guidelines in Commission Directive 93/21/EEC and Accord European Relatif ADR)
Conclusions:
The dermal LD50 values of BTMAC-60% in rabbits were calculated to be 633 mg/kg body weight for the males, 410 mg/kg body weight for the females and 510 mg/kg body weight for the sexes combined. Due to the mortality distribution, no fiducial limits could be determined for the LD50 values for males and females.
Executive summary:

Assessment of acute dermal toxicity with BTMAC-60% in the rabbit was carried out based on the guidelines described in: EC Commission Directive 92/69/EEC, Part B.3, "Acute Toxicity Dermal" and OECD # 402, "Acute Dermal Toxicity".

BTMAC-60% was administered to three rabbits of each sex by dermal application at 200, 400, 650 or 2000 mg/kg body weight for 24 hours. Animals were subjected to daily observations and weekly determinations of body weight. Macroscopic examination was performed on the day of death or after terminal sacrifice (day 15).

The incidence of mortality from low to high dose level was 0/3, 0/3, 2/3, 3/3 for males and 0/3, 1/3, 3/3, 3/3 for females. the decedents were found dead on days 1 and 2. One male and two females dosed at 2000 mg/kg were killed in extremis on day 1.

Lethargy, flat posture, quick and slow breathing, (excessive) salivation, pale skin and diarrhea was noted among the animals that died. Lethargy was noted in the surviving animals of the 650 and 400 mg/kg dose group on day 1 only. No clinical signs were noted in the animals dosed at 200 mg/kg.

Scales were seen in the treated skin-area among the animals treated at 400 mg/kg between days 5 and 8.

The changes noted in body weight pain in surviving males and females were considered to be normal.

Macroscopic post mortem examination of the animals that died or were killed in extremis during the study revealed hemorrhages or dark red foci in the thymus, thickening of the fundus in the stomach, irregular surface of the lungs and hemorrhagic fluid in the abdominal cavity. Dark red discoloration of hte subcutis of the treated skin area was found in two females dosed at 2000 mg/kg. Macroscopic post mortem examination of the surviving animals at termination did not reveal any abnormalities.

The dermal LD50 values of BTMAC-60% in rabbits were calculated to be 633 mg/kg body weight for the males, 410 mg/kg body weight for the females and 510 mg/kg body weight for the sexes combined. Due to the mortality distribution, no fiducial limits could be determined for the LD50 values for males and females.