3-methyl-1,3-butanediol-1-ethylate

Endpoint summary

Description of key information

Skin Sensitisation

An in vivo skin sensitisation study with 3-methyl-1,3-butanediol acetate (IPD-AC) is available. A local lymph node assay (LLNA) in the mouse has been conducted in accordance with OECD 429. Neat test substance and dilutions of 50% or 25% were added to the ears (25 µL per ear) of 4 animals. A control groups was also included. The results show that the test substance is not sensitising to skin. Therefore, according to Annex I for Regulation (EC) 1272/2008 the active ingredient, IPD-AC has no obligatory labelling requirement for skin sensitisation and is unclassified for this endpoint.

 

Respiratory sensitisation

There are currently no validated animal tests that deal specifically with respiratory tract sensitisation, therefore this endpoint was not investigated.

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

Study period:

2015-11-11 to 2016-05-31

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2010

GLP compliance:

yes (incl. QA statement)

Type of study:

mouse local lymph node assay (LLNA)

Specific details on test material used for the study:

Batch number: IPDAC-63313
Purity: 96.0%
Expiry date: 30 March 2016
Storage conditions: Room temperature in the dark, under nitrogen

Species:

mouse

Strain:

CBA/Ca

Sex:

female

Details on test animals and environmental conditions:

Acclimatisation period: 5 days
Weight at start of study: 15-23 g
Age at start of study: 8-12 weeks
Temperature: 19-25°C
Relative humidity: 30-70%
Air exchanges: 15 changes/hour
Photoperiod: 12 hour light/dark cycle
Food: 2014C Teklad Global Rodent diet; ad libitum
Water: Mains tap water; ad libitum
Caging: Suspended solid floor polypropylene cage

Vehicle:

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

Concentration:

Undiluted (100%), 50% or 25%

No. of animals per dose:

Four

Details on study design:

PRE-SCREEN TESTS:
One mouse was treated by daily application of 25 µL of undiluted test item to the dorsal surface of each ear for three consecutive days (Days 1, 2 and 3). Local skin irritation was recorded daily on Days 1 to 6. The ear thickness was measured pre-dose on Day 1, post dose on Day 3 and again on Day 6.Change in ear thickness was determined and a change of =>25% was considered to be excessive irritation.

MAIN STUDY
Groups of four mice were treated with undiluted test item, 50% or 25% concentrations in acetone:olive oil (4:1), or the vehicle alone. The mice were treated with by daily application of 25 µL of test substance or control to the dorsal surface of each ear for three consecutive days (Days 1, 2 and 3). The substance was administered by micropipette and spread over the dorsal surface of the ear using the tip of the pipette.
Five days following the first topical application (Day 6), mice were injected via the tail vein with 250 µL phosphate buffered saline contained 3H-methyl thymidine (3HTdR; 80 µCi/mL; specific activity 2.0 Ci/mmol) giving each mouse a total of 20 µCi.
Clinical observations were made twice daily on Days 1 to 3 and daily on Days 4 to 6. The body weight was recorded on Day 1 prior to dosing and on Day 6 prior to termination.
Five hours after administration of 3HTdR, the animals were euthanised. The draining auricular lymph node from the mice were excised and pooled for each group. Phosphate buffered saline (PBS) (1 mL) was added to each pooled sample. A single cell suspension was prepared by mechanical disaggregation through a 200-mesh gauze. The cells were rinsed through the gauze with 4 mL of PBS into a petri dish. The lymph nodes were transferred to a centrifuge tube and the petri dish rinsed with 5 mL of PBS which was added to the tube. The cells were centrifuged at 190 g for 10 minutes and resuspended in 10 mL of PBS and repelleted in 3 mL of 5% trichloroacetic acid.

After approximately 18 hours incubation at approximately 4°C, the precipitates were recovered by centrifugation at 450 g for 10 minutes, resuspended in 1 mL of TCA and transferred to 10 mL of scintillation fluid. The incorporation of 3HTdR was measured by ß-scintillation counting. After approximately 20 minutes in the dark, the samples were shaken and the number of radioactive disintegrations per minutes were measured.

DATA EVALUATION:
The proliferation of the lymph nodes was expressed as the number of radioactive disintegrations per minutes per lymph node (disintegrations per minutes/node) and as the ratio of 3HTdR incorporations into lymph node cells of test nodes relative to that recorded for the control nodes (Stimulation Index).
A test substance is considered to be sensitising if at least one concentration has a threefold or greater increase in 3HTdR incorporation compared to control values.
A test substance is considered to be not sensitising if there is no threefold or greater increase in 3HTdR incorporation compared to control values.

Positive control substance(s):

not specified

Statistics:

None

Positive control results:

Positive control, a-hexylcinnamaldehyde was tested previously in a separate study with the same vehicle and shown to be a sensitisier in the assay system (SI was 6.08).

Parameter:

SI

Value:

1.44

Test group / Remarks:

25%

Remarks on result:

other: No indication of skin sensitisation

Parameter:

SI

Value:

1.6

Test group / Remarks:

50%

Remarks on result:

other: No indication of skin sensitisation

Parameter:

SI

Value:

1.02

Test group / Remarks:

100%

Remarks on result:

other: No indication of skin sensitisation

Disintegrations per minute, disintegrations per minute/node and Stimulation Index

Concentration (%v/v) in acetone:olive oil (4:1)	Disintegrations per minute	Disintegrations per minute/node	Stimulation Index	Result
Vehicle control	2513.87	314.23	N/A	N/A
25	3628.08	453.51	1.44	Negative
50	4013.47	501.68	1.60	Negative
100	2561.86	320.23	1.02	Negative

Interpretation of results:

GHS criteria not met

Conclusions:

IPD-AC is considered not to be a skin sensitisier under the conditions of this study.

Executive summary:

A skin sensitisation study has been conducted in mice using the local lymph node assay in accordance with OCED test guideline 429 and in compliance with GLP. Groups of four mice were each treated with 50 µL (25 µL per ear) of 25%, 50% or 100% of IPD-AC. A control group of four mice were also tested with the vehicle control, acetone:olive oil at 4:1. The Stimulation Index (SI) was expressed as the mean radioactive incorporation for each treatment group divided by the mean radioactive incorporation of the vehicle control. The SI values for all the treatment groups were less than 3, indicating that the test substance is not sensitising to skin under the conditions of this study.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Comparison with the CLP criteria

In conclusion, the LLNA concludes that IPD-AC is devoid of skin sensitisation potential. Therefore, according to Annex I for Regulation (EC) 1272/2008 the active ingredient, IPD-AC has no obligatory labelling requirement for skin sensitisation and is unclassified for this endpoint.