Maleic anhydride

Endpoint summary

Description of key information

The potential of maleic anhydride to induce skin and respiratory sensitisation was tested in suitable in vivo test methods. Based on the results, the target substance can be considered a skin and respiratory sensitiser.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

yes

Remarks:

(no positve controls, mice strain BALB/c used)

GLP compliance:

not specified

Type of study:

mouse local lymph node assay (LLNA)

Species:

mouse

Strain:

Balb/c

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan Seralab, Bicester, Oxfordshire, UK
- Age at study initiation: 6-12 weeks old
- Housing: 10 per cage (acclimation period), 4-5 per cage (during treatment)
- Diet: ad libitum
- Water: ad libitum


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +/- 2
- Humidity (%): 55 +/- 10
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

acetone/olive oil (4:1 v/v)

Concentration:

0, 0.1,0.25, 0.5, 1, 2.5% (w/v)

No. of animals per dose:

4

Details on study design:

MAIN STUDY
Groups of mice (n= 4) were exposed topically on the dorsum of both ears to 25 µL of various concentrations of acid anhydrides or to vehicle (acetone:olive oil, AOO) alone, daily for three consecutive days. Five days after the initiation of exposure all mice were injected intravenously via the tail vein with 20 µCi of [3H] methyl thymidine in 250 µL of phosphate-buffered saline (PBS). Five hours later, mice were killed, and the draining auricular lymph nodes were excised and pooled for each experimental group. A single cell suspension of LNC was prepared by gentle mechanical disaggregation through 200-mesh stainless-steel gauze. Cells were washed twice with PBS and precipitated in 5% trichloroacetic acid (TCA) at 4 °C overnight. Pellets were then resuspended in 1 mL of 5% TCA and transferred to 10 mL of scintillation fluid. Incorporation of 3HTdR was measured by ß-scintillation counting as disintegrations per min (dpm) per node for each experimental group. In each case a stimulation index (SI) relative to the concurrent vehicle-treated control was derived. The estimated concentration of chemical required to induce SI= 3 relative to concurrent vehicle-treated controls i.e. the EC3 value was derived by linear interpolation.

Key result

Parameter:

EC3

Value:

0.16

Parameter:

SI

Value:

1.91

Test group / Remarks:

0.1% w/v

Parameter:

SI

Value:

4.86

Test group / Remarks:

0.25 % w/v

Parameter:

SI

Value:

6.32

Test group / Remarks:

0.5% w/v

Parameter:

SI

Value:

14

Test group / Remarks:

1% w/v

Parameter:

SI

Value:

15.98

Test group / Remarks:

2.5% w/v

Cellular proliferation data / Observations:

For detailed results please refer to Table 1 in box "Any other information on results incl. tables".

Table 1: Local lymph node assay responses to maleic anhydride

Concentrations (% w/v)	DPM per node	SI
0	245	1
0.1	467	1.91
0.25	1181	4.86
0.5	1548	6.32
1	3431	14
2.5	3915	15.98
5	ND
10	ND

ND = not determined

Interpretation of results:

Category 1A (indication of significant skin sensitising potential) based on GHS criteria

Conclusions:

In this study under the given conditions the EC3 was calculated to be 0.16% w/v = 0.016 M. Therefore, the test item in this test can be considered to be a skin sensitizer category 1A according to the CLP Regulation (EC) 1272/2008.

Executive summary:

In a dermal sensitization study conducted equivalent to OECD Guideline 429 young adult Balb/c mice (4 females per dose) were treated with 0.1, 0.25, 0.5, 1 and 2.5% maleic anhydride in acetone/olive oil (4:1, v/v). Five days after the initiation of exposure all mice were injected intravenously via the tail vein with 20 µCi of [3H] methyl thymidine in 250 µL of phosphate-buffered saline (PBS). Five hours later, mice were killed, and the draining auricular lymph nodes were excised and pooled for each experimental group. Incorporation of 3HTdR was measured by ß-scintillation counting as disintegrations per min (dpm) per node. Furthermore, in each case a stimulation index (SI) was derived.

The stimulation indices (SI) were 1.91 (0.1%), 4.86 (0.25%), 6.32 (0.5%), 14.00 (1%) and 15.98 (2.5%). Based on the results, the extrapolated EC3 value was determined to be 0.16%. Thus, maleic anhydride must be considered as a dermal sensitizer and classification as Skin Sens 1A, H317 according to the criteria of the CLP Regulation (EC) 1272/2008 is warranted.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

In a dermal sensitization study conducted equivalent to OECD Guideline 429 young adult Balb/c mice (4 females per dose) were treated with 0.1, 0.25, 0.5, 1 and 2.5% maleic anhydride in acetone/olive oil (4:1, v/v). Five days after the initiation of exposure all mice were injected intravenously via the tail vein with 20 µCi of [3H] methyl thymidine in 250 µL of phosphate-buffered saline (PBS). Five hours later, mice were killed, and the draining auricular lymph nodes were excised and pooled for each experimental group. Incorporation of 3HTdR was measured by ß-scintillation counting as disintegrations per min (dpm) per node. Furthermore, in each case a stimulation index (SI) was derived. The stimulation indices (SI) were 1.91 (0.1%), 4.86 (0.25%), 6.32 (0.5%), 14.00 (1%) and 15.98 (2.5%). Based on the results, the extrapolated EC3 value was determined to be 0.16%.Thus, maleic anhydride is a dermal sensitizer and classification as Skin Sens 1A, H317 according to the criteria of the CLP Regulation (EC) 1272/2008 is warranted. Supporting evidence for the potential of maleic anhydride to induce skin sensitisation was derived from data published in the OECD SIDS for maleic anhydride. In this study, a dermal sensitization study (according to OECD guideline 406 and EPA OPPTS 870.2600) with maleic anhydride in mineral oil 20Crl: (HA)BR guinea pigs were tested using the method of Buehler. The guinea pigs were exposed with 5% (w/v) maleic anhydride in mineral oil during the three-application induction phase. The maleic anhydride produced very faint to moderate erythema reactions in all 20 test animals (eight animals with scores of 0.5, and 12 animals with scores of 1.0 to 2.0). Pinpoint areas of subcutaneous haemorrhaging were also observed in two of these animals. None of the vehicle control animals reacted to the initial challenge application of the test material. None of the test or vehicle control animals reacted to the vehicle control material. All reactions to maleic anhydride in the test group (scores of 0.5 to 2.0) exceeded the highest reaction in the vehicle control group (score of 0.0). In this study maleic anhydride is a dermal sensitizer. Furthermore, maleic anhydride is generally considered as skin sensitizer in humans and was positively tested in patch and prick tests.

Respiratory sensitisation

Link to relevant study records

Reference

Endpoint:

respiratory sensitisation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Qualifier:

no guideline followed

Principles of method if other than guideline:

To determine the sensitising potential of Maleic anhydride, rats were exposed to a maleic anhydride aerosol 6 hours/day for five days. Following a 3-week rest period, the animals were challenged for 6 hours. One group was not challenged (i.e., nonexposed/nonchallenged control).

GLP compliance:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Route of induction exposure:

inhalation

Route of challenge exposure:

inhalation

Vehicle:

not specified

Concentration:

500 µg/m³ (induction and challenge period)

No. of animals per dose:

10/sex

Details on study design:

A. INDUCTION EXPOSURE
- No. of exposures: 5
- Exposure period: 6 hrs
- Frequency of applications: every day
- Duration: 5 days
- Concentrations: 500 µg/m³


B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 26
- Exposure period: 6 hrs
- Concentrations: 500 µg/m³

Positive control substance(s):

none

Negative control substance(s):

other: unexposed group

Results:

The maleic anhydride-exposed/maleic anhydride-challenged animals had small, but statistically significant (p< 0.05), increases in maleic anhydride-specific serum IgG antibody compared to the controls (challenged and nonchallenged; females higher than males). Two rats of the MA-exposed/nonchallenged group had more than 10 lung foci (i.e., positive response); however, mean values for lung foci, weight, and volume were not significantly different from control values. Microscopic lung lesions were minimal and provided no evidence of pulmonary sensitization. The serum IgG antibody levels of the maleic anhydride-exposed group were significantly increased over those of the MA-exposed and challenged males and the challenged and nonchallenged control males.

Positive control results:

not available

Negative control results:

Accumulation of alveolar macrophages was seen histologically in one control rat and no maleic anhydride-exposed rats.

The analytical time-weighted averaged concentration of maleic anhydride was 500 and 317 µg/m³, for the induction and challenge phases, respectively.

Interpretation of results:

Category 1 (respiratory sensitising) based on GHS criteria

Conclusions:

Based on the results of this study, maleic anhydride is considered to be a respiratory sensitiser. In accordance with ATP13 to CLP regulation 1272/2008 classification as Resp. Sens. 1, H334 is warranted.

Executive summary:

In a respiratory sensitisation study, Sprague- Dawley rats (n=10/sex) were exposed to a particulate aerosol target concentration of 0 or 500 µg/m³ maleic anhydride, 6 hours/day for five days. Following a 3-week rest period, the animals were challenged with 500 µg/m³ for 6 hours. The analytical time weighted averaged concentration of maleic anhydride was 500 and 317 µg/m³, for the induction and challenge phases, respectively. The maleic anhydride-exposed/maleic anhydride-challenged animals had small, but statistically significant (p< 0.05), increases in maleic anhydride-specific serum IgG antibody compared to the controls (challenged and nonchallenged; females higher than males). However, other prominent features of respiratory sensitization reactions in this rat model (such as increased numbers of external hemorrhagic lung foci, increased lung weight and volume, and extensive lung pathology) were not evident.

Based on the results of this study, maleic anhydride is considered to be a respiratory sensitiser. In accordance with ATP13 to CLP regulation 1272/2008 classification as Resp. Sens. 1, H334 is warranted.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Additional information:

In a respiratory sensitisation study, Sprague- Dawley rats (n=10/sex) were exposed to a particulate aerosol target concentration of 0 or 500 µg/m³ maleic anhydride, 6 hours/day for five days. Following a 3-week rest period, the animals were challenged with 500 µg/m³ for 6 hours. The analytical time weighted averaged concentration of maleic anhydride was 500 and 317 µg/m³, for the induction and challenge phases, respectively. The maleic anhydride-exposed/maleic anhydride-challenged animals had small, but statistically significant (p< 0.05), increases in maleic anhydride-specific serum IgG antibody compared to the controls (challenged and nonchallenged; females higher than males). However, other prominent features of respiratory sensitization reactions in this rat model (such as increased numbers of external hemorrhagic lung foci, increased lung weight and volume, and extensive lung pathology) were not evident. Based on the results of this study, maleic anhydride is considered to be a respiratory sensitiser. Furthermore, several publications described occupational asthma due to maleic anhydride (Lee et al. 1991; Gannon et al. 1992; Graneek et al. 1986, Val. 4). When all subjects were included and all three acid anhydrides were taken into account there was no consistent evidence for an exposure-response relation, but with the analysis restricted to a factory where only trimellitic anhydride (TMA) was in use, there was an increased prevalence of sensitisation to acid anhydrides and work related respiratory symptoms with increasing full shift exposure. This relation was apparent within the current occupational exposure standard of 40 μg/m³ and was not modified significantly by smoking or atopy.

Justification for classification or non-classification

Based on the available data and in accordance with ATP13 to CLP regulation 1272/2008 classification as Skin Sens 1A, H317 and Resp. Sens. 1, H334 is warranted.