Diisopentyl ether

Endpoint summary

Description of key information

The test substance was found to be not irritating or corrosive to the skin and only slightly irritating to the eye.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was performed according to the OECD guideline No.439 (July 2010) and in compliance with the GLP guidelines.

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Deviations:

no

GLP compliance:

yes

Species:

other: In vitro human skin model

Strain:

other: EpiDerm model

Details on test animals or test system and environmental conditions:

TEST MATERIAL
- Test material: commercially available Epi-200-SIT-Kit
- Source: MatTek Corporation Ashland USA (first experiment), MatTek In Vitro Life Science Laboratories, Bratislava (Second experiment)
- Acclimation period: On the day of receipt the tissues were transferred to 12-well plates and preincubated with prewarmed Maintenance Medium for 24 hours at 37°C.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 37.1 – 37.4°C
- Humidity (%): 88 – 89% containing 5 +/- 0.5% CO2 in air
- Photoperiod (hrs dark / hrs light): dark

Type of coverage:

other: In vitro human skin model

Preparation of test site:

other: In vitro human skin model

Vehicle:

unchanged (no vehicle)

Controls:

other: other: Negative control: Phosphate buffered saline; positive control: 5% Sodium dodecyl sulphate in deionised H2O.

Amount / concentration applied:

Three tissues of the human skin model EpiDerm were treated with 30 µl diisopentyl ether.

Duration of treatment / exposure:

The skin tissues were exposed to diisopentyl ether for 60 minutes at 37°C.

Observation period:

Not applicable

Number of animals:

Not applicable (in vitro study)

Details on study design:

The purpose of this study was to determine skin irritation potential of diisopentyl ether using the in-vitro human epidermal model (EpiDerm). The test consists of a topical exposure of the neat test item to a human reconstructed epidermis model followed by a cell viability test. Cell viability is measured by dehydrogenase conversion of MTT (3-[4,5-dimethyl thiazole 2-yl]2,5-diphenyl-tetrazoliumbromide), present in cell mitochondria, into a blue formazan salt that is quantitatively meaured after extraction from tissues. The percent reduction of cell viability in comparison of untreated negative controls is used to predict skin irritation potential .

Protocol:
1- Pre-Tests: first, it was tested whether the test item is compatible with the EpiDerm test system. No reaction with reagens or materials was observed and therefore this protocol was considered suitable for the determination of the skin irritation potential of diisopentyl ether.
2- Pre-incubation: Skin tissues were pre-incubated to test their viability.
3- Treatment: Pre-incubated tissues were treated with buffer (negative control), SDS solution (positive control) or diisopentyl ether. Hereafter the tissues were incubated for 60 minutes. After the incubation period, the tissues were rinsed, dried and then transferred into a new plate with fresh asay medium. After a total post-incubation period of 42 hours, the cytotoxic effect was measured by determination of the relative formazan production (photometric absorption at 570 nm). The photometric absorption of the negative controls is considered as 100%. The mean of the three replicates of test item and positive control, formazan production is calculated as % photometric absorption compared with the negative control. If the formazan production is < 50% in comparison to the control, the test item is considered as irritant. If >50%, the test item is considered as non-irritant.
Two independant experiments were performed.

Irritation / corrosion parameter:

other: EpiDerm Model

Remarks:

Mean of three tissues

Run / experiment:

1

Value:

ca. 1.517

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Irritation / corrosion parameter:

other: Epiderm Model / Mean of three tissues

Run / experiment:

2

Value:

ca. 2.237

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

After treatment, the absorbance values in the negative controls were within the required acceptability criterion. The positive control showed clear irritating effects. Variation within tissues was acceptable (<18%) for negative control, positive control and test item. In the first experiment the relative aabsorbance values were increased to 148.2% after the treatment with diisopentyl ether. In the second experiment, the relative absorbance values were increased to 151.8% after the treatment with the test item. Both values are well above the threshold for irritation potential (50%) but lie far above the value of the negative control, indicating an increased viability effect of the test item on the skin. It is clear that this effect is not caused by a direct interaction from the test substance with MTT since in the pre-test it was concluded that no interaction takes place.

Mean absorption values, Exp. 1

	Negative control	Diisopentyl ether	Positive control
Tissue 1	1.017	1.531	0.115
Tissue 2	1.024	1.518	0.111
Tissue 3	1.031	1.503	0.117
Mean of three tissues	1.024	1.517	0.114
Relative SD of the three tissues	0.7%	0.9%	2.7%

Mean absorption values, Exp. 2

	Negative control	Diisopentyl ether	Positive control
Tissue 1	1.493	2.226	0.079
Tissue 2	1.466	2.241	0.079
Tissue 3	1.463	2.245	0.095
Mean of three tissues	1.474	2.237	0.084
Relative SD of the three tissues	1.1%	0.4%	11.0%

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Diisopentyl ether is considered as not irritant in the Human Skin Model Test.

Executive summary:

A GLP compliant in-vitro study was performed according to OECD Guideline 439 in order to evaluate the potential of diisopentyl ether to evoke skin irritation in a human skin model. Two valid, independant experiments were performed. Three tissues of the human skin model EpiDerm were treated with diisopentyl ether for 60 minutes. After a post-incubation period, determination of the cytotoxic (irritancy) effect was performed. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT at the end of the treatment. DPBS-buffer was used as negative control, 5% SDS-solution was used as postive control.

After treatment, the absorbance values in the negative controls were within the acceptability criterion. The positive control showed clear irritating effects. In the first experiment the relative absorbance values were increased to 148.2% after the treatment with diisopentyl ether. In the second experiment, the relative absorbance values were increased to 151.8% after the treatment with the test item. Both values are well above the threshold for irritation potential (50%) and therefore it can be concluded that diisopentyl ether is not a skin irritant in the Human Skin Model Test.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline study, no restrictions, fully adequate for assessment.

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

other: in-vitro test

Strain:

other: isolated bovine cornea

Details on test animals or tissues and environmental conditions:

TEST MATERIAL
- Source: slaughterhouse (Muller Fleisch GmbH, Birkenfeld, Germany)
- Acclimation period: the eyes were transported to the test facility in Hank's balanced salt solution (supplemented with 0.01% streptomycin and 0.01% penicillin). Then, the corneas were dissected and incubated in incubation medium at 32 +/- 1°C in an incubation chamber for 1 hour.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 32 +/- 1°C

Vehicle:

unchanged (no vehicle)

Controls:

other: Negative control: physiological sodium chloride solution; positive control: 10% sodium hydroxide solution.

Amount / concentration applied:

An amount of 750 µl diisopentyl ether was applied directly on the corneas in such a way that the cornea was completely covered.

Duration of treatment / exposure:

The corneas were incubated in a horizontal position for 10 minutes at 32°C.

Observation period (in vivo):

Not applicable, the effects on the corneas was determined directly after the exposure period.

Number of animals or in vitro replicates:

Not applicable (in vitro study)

Details on study design:

The aim of this study was to evaluate the ocular irritancy of diisopentyl ether as measured by its ability to induce opacity and increase permeability in an isolated bovine cornea.

PROTOCOL
- For each treatment group (negative control, positive control and test item), three replicates were used.
- The opacity of each cornea was determined before exposure.
- The corneas were exposed for 10 minutes.
- After the incubation the change in opacity for each individual cornea (including the negative control) was determined.
- Following the final opacity measurement, permeability of the cornea to Na-fluorescein was evaluated.

INTERPRETATION
The assay is considered acceptable if:
- The positive control gives an in vitro irritancy score (IVIS) that falls within two standard deviations of the laboratory historical mean value.
- The negative control response has to show an IVIS between 0 and 3.
- The mean opacity and mean permeability values (OD490) were used for each treatment group to calculate an in vitro irritancy score (IVIS)
- A test substance that induces an IVIS ≥ 55.1 is defined as a corrosive or severe irritant.

Irritation parameter:

other: The mean opacity and mean permeability values (OD490) were used for each treatment group to calculate an in vitro score.

Remarks on result:

other: Diisopentyl ether did not induce ocular irritation through both endpoints, resulting in a mean in vitro irritancy score of 0.0080 after 10 minutes of treatment.

Irritant / corrosive response data:

The negative control showed no irritation (mean IVIS was 0.443). The positive control induced a very severe irritation on the cornea, mean IVIS was 195.5. It was therefore concluded that the test conditions were adequate and that the test system functioned properly.
The mean in vitro irritancy score of the test item was 0.008 after 10 minutes of treatment with diisopentyl ether. For this reason it can be concluded that diisopentyl ether is not severe irritant or corrosive in the Bovine Corneal Opacity and Permeability test.

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Diisopentyl ether is not severe irritant or corrosive in the Bovine Corneal Opacity and Permeability test.

Executive summary:

A GLP compliant in-vitro study was performed according to OECD Guideline 437 in order to evaluate the ocular irritation properties of diisopentyl ether on an isolated bovine cornea. The possible ocular irritancy of diisopentyl ether was tested through topical application for approximately 10 minutes. For each treatment group (negative control, positive control and test item), three replicates were used. After removal of the test item and two hours post-incubation, opacity and permeability values were measured.

After treatment, the IVIS (in vitro irritancy score) in the controls is within the acceptability criterion. The positive control showed clear irritating effects. Diisopentyl ether showed no effects on the cornea of the bovine eyes. The IVIS score is 0.008 and therefore it can be concluded that diisopentyl ether is not an eye irritant in the BCOP Test.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Additional information

Skin irritation:

A GLP compliant in-vitro study was performed accoring to OECD Guideline 439 in order to evaluate the potential of diisopentyl ether to evoke skin irriation in a human skin model. Two valid, independant experiments were performed. After treatment, the absorbence values in the negative controls were within the acceptability criterion. The positive control showed cleare irritating effects. In the first and second experiment the relative absorbence values were increased to 148.2% and 151.8%, resp. after the treatment with diisopentyl ether. Both values are well above the threshold for irritation potential (50%) and therefore it can be concluded that diisopentyl ether is not a skin irritant in the Human Skin Model Test.

Eye irritation:

A GLP compliant in-vitro study was performed according to OECD Guideline 437 in order to evaluate the ocular irritation properties of diisopentyl ether on an isolated bovine cornea. Diisopentyl ether showed no effects on the cornea of the bovine eyes and therefore it can be concluded that diisopentyl ether is not an eye irritant in the BCOP Test.

An in-vivo test according to OECD Guideline 405 and GLP Guidelines was performed to confirm the results of the BCOP Test. Single samples of 0.1 mL of diisopentyl ether were instilled into one eye of each of three rabbits. Observations were made 1, 24, 48, 72 hours and 7 days after instillation. Instillation of the test substance resulted in irritation of the conjunctivae, which consisted of redness, chemosis and discharge. The irritation had completely resolved within 72 hours in two animals and within 7 days in the remaining animal. Based on these results, diisopentyl ether is considered to be a slight irritant to the eye.

Effects on eye irritation: slightly irritating

Justification for classification or non-classification

Based on the findings in the skin and eye irritation study the test substance does not need to be classified according to the Directive 67/548/EEC and according to the EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008