Registration Dossier

Data are available to assess the acute oral and dermal toxicity of the target substance succinic anhydride.

Succinic anhydride was of low toxicity as indicated by the combined LD50 (male/female) of 1794.9 mg/kg bw, when tested in an acute oral toxicity study comparable to OECD TG 401. When tested for acute dermal toxicity, succinic anhydride did not show signs of toxicity or death up to the limit dose of 2000 mg/kg bw in a study similar to OECD TG 402.

Acute inhalation testing is not required. The results of the granulometry indicate the L10 value for particle size was 377 µm and the L50 was greater than 1100 µm.
On the basis of these results it is clear that the particles present in the batch of succinic anhydride are not in the respirable size range for rats or humans and so inhalation exposure is not likely. Given the results can be predicted from the physical-chemical properties and that there are already studies available from two relevant exposure routes, an inhalation toxicity study cannot be justified because of animal welfare reasons.

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1981-03-11 to 1981-04-14

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Version / remarks:

adopted 12 May 1981

Deviations:

no

Principles of method if other than guideline:

standard protocol for determination of acute median lethal oral dose.

GLP compliance:

yes (incl. QA statement)

Test type:

standard acute method

Limit test:

no

Specific details on test material used for the study:

Succinic anhydride appeared as white solid flakes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

After an acclimation period of at least 7 days, animals were assigned to groups of two males and two females at five dose levels for the preliminary study and five males and five females at six dose levels for the principal study. Animals were individually housed in wire mesh bottom cages in environment controlled rooms.

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on oral exposure:

Animals were fasted overnight prior to receiving a single oral dose of the test article at five dosing levels. The test article was administered at a constant concentration and the volume of dosing solution did not exceed 5 mL per animal, where possible.

Doses:

Preliminary study: 50, 139, 387, 1078 and 3000 mg/kg; Principal study: 1214, 1500, 1854, 2291, 2832, and 3500 mg/kg

No. of animals per sex per dose:

Preliminary study: 2 males and 2 females per dose; Principal study: 5 males and 5 females per dose

Control animals:

no

Details on study design:

All animals on the main study were observed for at least 14 days or until all signs of reversible toxicity subsided, whichever occurred later. Animals were observed three times a day on the day of dosing and twice daily for the remainder of the study. All gross or visible toxic or pharmacological effects were recorded. Body weights were recorded initially and on days 8 and 15 or at death. All animals that died and all animals sacrificed at termination were subject to gross necropsy.

Statistics:

No details but graphical representations indicate probit analysis used to determine the median lethal dose

Preliminary study:

The preliminary dose range -finding study gave mortality results at dose levels of 50 (0/4); 139 (0/4); 387 (0/4); 1078 (0/4) or 3000 (4/4) mg/kg bw

Sex:

male

Dose descriptor:

LD50

Effect level:

2 157.2 mg/kg bw

95% CL:

1 722.2 - 2 758.6

Sex:

female

Dose descriptor:

LD50

Effect level:

1 510.5 mg/kg bw

95% CL:

1 086.9 - 1 816.7

Sex:

male/female

Dose descriptor:

LD50

Effect level:

1 794.9 mg/kg bw

95% CL:

1 505 - 2 071.5

Mortality:

For male rats, cumulative mortality at dose levels of 1214, 1500, 1854, 2291, 2832, and 3500 mg/kg was 0/5, 1/5, 2/5, 3/5, 3/5 and 5/5, respectively. For female rats, cumulative mortality at the same dose levels was 1/5, 3/5, 3/5, 5/5, 5/5 and 5/5, respectively. (see Table 1 in section "Any other information on results incl. tables" for details on mortality data)

Clinical signs:

other: In males at dose levels of 1500 mg/kg and higher, decreased activity and death was seen. Soft stools were reported for males dosed at 2291 mg/kg and ataxia was observed for males at the two highest dose levels. In females, decreased activity and death was

Gross pathology:

Black pylorus in stomach and intestines containing a blood-like substance were seen in males at doses of 2291 mg/kg and above. At the highest dose, green areas on the lungs were seen in males at necropsy. In females, black stomach pyloric and intestines containing a blood-like substance were seen at doses of 1854 mg/kg and higher. At necropsy, green areas on the lungs were seen in females dosed at 2291 and 3500 mg/kg.

Other findings:

No data

Table 1. Mortality data

	Time of Death
Dose level (mg/kg)	Day 1	Day 2	Day 3	Day 8	Day 15	Cumulative mortality
Males
1214	0	0	0	0	0	0/5
1500	0	1	0	0	0	1/5
1854	1	0	1	0	0	2/5
2291	0	3	0	0	0	3/5
2832	3	0	0	0	0	3/5
3500	5	--	--	--	--	5/5
Females
1214	1	0	0	0	0	1/5
1500	0	3	0	0	0	3/5
1854	1	2	0	0	0	3/5
2291	3	2	--	--	--	5/5
2832	5	--	--	--	--	5/5
3500	5	--	--	--	--	5/5

Table 2. Summary of body weights

	Body weights (mean + SD)
Dose level (mg/kg)	Initial*	8 days	15 days	at death
Males
1214#	188.8 + 14.9	254.4 + 11.4	281.6 + 12.5	--
1500	319.8 + 23.3	346.3 + 40.4	365.3 + 38.5	294.0
1854	322.8 + 8.3	348.0 + 19.1	372.7 + 16.3	307.0 + 35.4
2291	324.8 + 24.5	359.5 + 12.0	386.0 + 5.7	308.7 + 12.1
2832	318.0 + 16.1	364.5 + 12.0	385.5 + 13.4	312.7 + 19.4
3500	306.0 + 20.4	--	--	306.0 + 20.4
Females
1214#	191.2 + 6.4	226.0 + 12.3	227.0 + 12.3	188.0
1500	222.0 + 15.7	239.0 + 7.1	249.0 + 9.9	208.7 + 17.2
1854	214.4 + 9.3	247.0 + 15.6	255.0 + 24.0	210.0 + 11.1
2291	230.4 + 28.2	--	--	227.2 + 25.2
2832	225.6 + 8.6	--	--	225.6 + 8.6
3500	231.2 + 19.8	--	--	231.2 + 19.8
*Fasted body weight # Dosed on a different day

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

The results of this study indicate that succinic anhydride is of low acute oral toxicity when tested according to OECD test guideline 401. The combined LD50 for males and females is 1794.9 mg/kg bw. This result would indicate classification for acute toxicity Cat.4, H302 (Harmful if swallowed) according to the CLP criteria as set out in Regulation (EC) 1272/2008 may be appropriate.

Executive summary:

In an acute oral toxicity study performed according to OECD TG 401, groups of 5 fasted Sprague-Dawley rats/sex were given a single oral dose of succinic anhydride by gavage at doses of 1214, 1500, 1854, 2291, 2832 and 3500 mg/kg bw and observed for at least 14 days.

For male rats, cumulative mortality at dose levels of 1214, 1500, 1854, 2291, 2832, and 3500 mg/kg was 0/5, 1/5, 2/5, 3/5, 3/5 and 5/5, respectively. For female rats, cumulative mortality at the same dose levels was 1/5, 3/5, 3/5, 5/5, 5/5 and 5/5, respectively.

The oral LD50 value was 2157.2 mg/kg bw for males and 1510.5 mg/kg bw for females. The combined oral LD50 was 1794.9 mg/kg bw.

Clinical signs included decreased activity in males at dose levels of 1500 mg/kg and higher. Soft stools were reported for males dosed at 2291 mg/kg and ataxia was observed for males at the two highest dose levels.

In females, decreased activity was observed at all dose levels. Soft stools were observed in females dosed at 1214 mg/kg and ataxia was observed at doses of 1500 mg/kg and higher.

Black pylorus in stomach and intestines containing a blood-like substance were seen in males at doses of 2291 mg/kg and above and in females at doses of 1854 mg/kg and higher.

At the highest dose in males, green areas on the lungs were seen at necropsy. In females, these effects were observed at doses levels of 2291 and 3500 mg/kg.

Succinic anhydride is of low toxicity based on the combined LD50 of 1794.9 mg/kg bw. Classification to Category 4 for acute oral toxicity is warranted according to the CLP criteria as set out in Regulation (EC) 1272/2008.

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

1 794.9 mg/kg bw

Quality of whole database:

guideline study

Reference

Endpoint:: acute toxicity: inhalation
Data waiving:: study scientifically not necessary / other information available
Justification for data waiving:: the study does not need to be conducted because exposure of humans via inhalation is not likely taking into account the vapour pressure of the substance and/or the possibility of exposure to aerosols, particles or droplets of an inhalable size

Endpoint conclusion:

no study available

Reference

Endpoint:

acute toxicity: dermal

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010-03-16 to 2010-04-28

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 402 (Acute Dermal Toxicity)

Version / remarks:

24 February 1987

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.3 (Acute Toxicity (Dermal))

Version / remarks:

30 May 2008

Deviations:

no

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Specific details on test material used for the study:

- Appearance: white powder/ colourless solid
- Name of the test material used in the report: Succinic anhydride (trade name Bernsteinsäureanhydrid)
- Batch no.: LEBA5A3070
- Purity: 99.5%
- Storage temperature: room temperature
- Expiry date: 30 March 2010

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
Appropriate amounts of the test substance are ground to a fine dust (if necessary) and then administered undiluted on a cellulose patch after moistening with corn oil to allow an optimal contact to the skin.

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

The animals were male and female Crl:CD(SD) rats, obtained from Charles River Deutschland GmbH. Females were nulliparous and non-pregnant. The males were approximately 8 weeks old and the females were approximately 12 weeks old at the time of administration. The rats were acclimatised for at least 7 days. A health inspection was performed prior to commencement of the study to ensure that the animals were in good health. Special attention was paid to the skin, which was confirmed to be intact and free from any abnormality. Room temperature ranged from 19.96 to 20.99°C. Relative humidity ranged from 45.08 to 58.78%. There were 12 air changes per hour, and artificial light was provided on a 12 hour light/dark cycle. The rats were housed singly in Makrolon type III cages with wire mesh lids. Autoclaved Aspen wood chips were provided as bedding. Nibbling wood bricks and nesting material from the same material and source as the bedding material were also provided. Ssniff R/M-H maintenance diet for rats and mice was provided ad libitum (Ssniff Spezialdiäten GmbH, Germany). Tap water was provided ad libitum. Individuals were identified with felt-tipped pen on the tails.

Type of coverage:

semiocclusive

Vehicle:

corn oil

Details on dermal exposure:

A single dermal administration was performed by applying the test substance to an area of least 10% of the estimated body surface. The test site was located on the dorsal thoracic region. An area of 6.5 cm x 8 cm (52 cm²) was marked. The hair of the dorsal trunk was clipped with an electric hair clipper one day before test substance application. Test substance amounts were calculated and weighed for each rat using individual body weights determined on the day of administration. A cellulose patch (Pehazell, Hartmann AG) with the correct amount of test substance on the surface and soaked in corn oil (to get optimal contact with the skin) was applied to the test site and held in place with non-irritating tape (Blenderm 3M). The test site was covered by a semi-occlusive dressing (Fixomull Stretch). At the end of the exposure period the dressing, the tape and the patch were removed. Residual test substance was wiped off using wet cellulose tissue, if necessary.

Duration of exposure:

24 hours

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

5 rats/sex

Control animals:

not required

Details on study design:

Observations were performed 0-0.5, >0.5-1, >1-2, >2-4 and >4-6 hours after administration, then at least once daily for a total of 2 weeks. Observations included but were not limited to changes in skin, fur, eyes, the occurrence of secretions and excretions, autonomic activity, changes in gait, posture and the presence of convulsions. Body weights were determined immediately prior to administration, then 7 and 14 days after administration. All animals were sacrificed at the end of the observation period by CO2 inhalation and subjected to a necropsy including a gross pathological examination.

Statistics:

Formal statistical analysis was not required.

Preliminary study:

Not applicable.

Key result

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No deaths occurred during the study.

Clinical signs:

other: No clinical signs were observed in any animal during the observation period.

Gross pathology:

No abnormalities were detected at necropsy.

Other findings:

3/5 males and all females exhibited erythema and eschar formation at the application site. These local effects were observed 1 day after administration lasting until a maximum of 7 days post administration.

Interpretation of results:

not classified

Conclusions:

The acute dermal LD50 of succinic anhydride is greater than 2000 mg/kg bw in rats. No classification is warranted according to the CLP criteria as set out in Regulation (EC) 1272/2008.

Executive summary:

In an acute dermal toxicity study performed according to OECD TG 302, a group 5 young adult Sprague-Dawley rats/sex were dermally exposed to succinic anhydride to the clipped dorsal skin corresponding to 10% of the estimated body surface area at the limit dose of 2000 mg/kg bw. The test site was covered by a semi-occlusive dressing, held in place for 24 hours. Animals were then observed for 14 days.

No mortality occurred during the study period. There were no effects on body weight; all animals gained weight during the observation period. No clinical signs of systemic toxicity were observed, however local irritation at the test site (erythema and eschar formation) was noted 1 to 7 days after administration. No abnormalities were detected at necropsy.

Based on these results, it was concluded that the acute dermal LD50 of succinic anhydride is greater than 2000 mg/kg bw in rats. Therefore, no classification is warranted according to the CLP criteria as set out in Regulation (EC) 1272/2008.

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Quality of whole database:

guideline study

In an acute oral toxicity study performed according to OECD TG 401, groups of 5 fasted Sprague-Dawley rats/sex were given a single oral dose of succinic anhydride by gavage at doses of 1214, 1500, 1854, 2291, 2832 and 3500 mg/kg bw and observed for at least 14 days.

For male rats, cumulative mortality at dose levels of 1214, 1500, 1854, 2291, 2832, and 3500 mg/kg was 0/5, 1/5, 2/5, 3/5, 3/5 and 5/5, respectively. For female rats, cumulative mortality at the same dose levels was 1/5, 3/5, 3/5, 5/5, 5/5 and 5/5, respectively.

The oral LD50 value was 2157.2 mg/kg bw for males and 1510.5 mg/kg bw for females. The combined oral LD50 was 1794.9 mg/kg bw.

Clinical signs included decreased activity in males at dose levels of 1500 mg/kg and higher. Soft stools were reported for males dosed at 2291 mg/kg and ataxia was observed for males at the two highest dose levels.

In females, decreased activity was observed at all dose levels. Soft stools were observed in females dosed at 1214 mg/kg and ataxia was observed at doses of 1500 mg/kg and higher.

Black pylorus in stomach and intestines containing a blood-like substance were seen in males at doses of 2291 mg/kg and above and in females at doses of 1854 mg/kg and higher.

At the highest dose in males, green areas on the lungs were seen at necropsy. In females, these effects were observed at doses levels of 2291 and 3500 mg/kg.

Succinic anhydride is of low toxicity based on the combined LD50 of 1794.9 mg/kg bw.

In an acute dermal toxicity study performed according to OECD TG 302, a group 5 young adult Sprague-Dawley rats/sex were dermally exposed to succinic anhydride to the clipped dorsal skin corresponding to 10% of the estimated body surface area at the limit dose of 2000 mg/kg bw. The test site was covered by a semi-occlusive dressing, held in place for 24 hours. Animals were then observed for 14 days.

No mortality occurred during the study period. There were no effects on body weight; all animals gained weight during the observation period. No clinical signs of systemic toxicity were observed, however local irritation at the test site (erythema and eschar formation) was noted 1 to 7 days after administration. No abnormalities were detected at necropsy.

Based on these results, it was concluded that the acute dermal LD50 of succinic anhydride is greater than 2000 mg/kg bw in rats.

Acute inhalation testing is not required. The results of the granulometry indicate the L10 value for particle size was 377 µm and the L50 was greater than 1100 µm. On the basis of these results it is clear that the particles present in the batch of succinic anhydride are not in the respirable size range for rats or humans and so inhalation exposure is not likely. Given the results can be predicted from the physical-chemical properties and that there are already studies available from two relevant exposure routes, an inhalation toxicity study cannot be justified because of animal welfare reasons.

Based on the available data and in accordance with Annex VI to CLP Regulation 1272/2008 succinic anhydride classification as Acute Tox 4, H302 is warranted.