Registration Dossier

Administrative data

Description of key information

Skin: The test substance caused only very slight irritation in the rabbit, fully reversible within 8 days.
Eye: The test substance showed eye irritating potential in the EpiOcular in vitro test as well as in vivo with rabbits.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records
Reference
Endpoint:
skin irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1967
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Non GLP, study predating OECD guideline, but scientifically acceptable
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
Deviations:
not specified
Principles of method if other than guideline:
BASF-Test
GLP compliance:
no
Species:
rabbit
Strain:
Vienna White
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Weight at study initiation: 3.0 kg
Type of coverage:
occlusive
Preparation of test site:
not specified
Vehicle:
unchanged (no vehicle)
Controls:
not specified
Amount / concentration applied:
no data
Duration of treatment / exposure:
1, 5, 15 min
20 h
Observation period:
8 days
Number of animals:
2 males
Details on study design:
TEST SITE
- Area of exposure: Back (2.5 x 2.5 cm) and ear

SCORING SYSTEM: Draize scoring system. Results originally reported according to BASF scoring system and transformed to scores according to OECD guideline 404.
Irritation parameter:
erythema score
Basis:
animal #1
Time point:
other: 24/48 h
Score:
1.5
Max. score:
4
Reversibility:
fully reversible within: 8 d
Remarks on result:
other: application time 20 h
Irritation parameter:
erythema score
Basis:
animal #2
Time point:
other: 24/48 h
Score:
1.5
Max. score:
4
Reversibility:
fully reversible within: 8 d
Remarks on result:
other: application time 20 h
Irritation parameter:
edema score
Basis:
animal #1
Time point:
other: 24/48 h
Score:
0
Max. score:
4
Remarks on result:
other: application time 20 h
Irritation parameter:
edema score
Basis:
animal #2
Time point:
other: 24/48 h
Score:
0
Max. score:
4
Remarks on result:
other: application time 20 h
Irritant / corrosive response data:
No signs of irritation were observed in two animals after 1, 5 and 15 min exposure on the back. Slight redness was observed in both animals after 24 and 48 h (ear and back) when treated for 20 h. The irritation was reversible within 8 days.
Other effects:
none
Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
Classification: not irritating
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: Modern guideline study conducted according to GLP.
Qualifier:
no guideline available
Principles of method if other than guideline:
There are no official national or international guidelines for the EpiOcularTM test yet; however, the study was performed according to the methods described in the following publications:

MatTek Corporation, Ashland, MA 01721, USA: EpiOcularTM human cell construct: Procedure details, Version 3.1a of February 10, 2010.

Harbell J.W. et al. (2009): COLIPA Program on Optimization of Existing In Vitro Eye Irritation Assays for Entry into Formal Validation: Technology

Transfer and Intra/Inter Laboratory Evaluation of EpiOcular Assay for Chemicals. Poster # 378, Society of Toxicology, March 2009.

In addition the study follows the testing strategy for determination of eye irritation/corrosion as given in the following OECD guideline:
OECD Guideline for Testing of Chemicals No. 405, April 24, 2002 (“Acute Eye Irritation/Corrosion”)
GLP compliance:
yes (incl. QA statement)
Species:
other: non-keratinized tissue construct composed of normal human derived epidermal keratinozytes
Vehicle:
unchanged (no vehicle)
Controls:
other: negative and positive control
Duration of treatment / exposure:
undiluted liquid test substance was applied covering the whole tissue surface
Observation period (in vivo):
after application, the tissues were placed into the incubator until the total exposure time of 30 minutes was completed
Irritation parameter:
other: optical density [wavelength 570 nm]
Basis:
mean
Time point:
other: after 30 minutes exposure
Score:
0.326
Remarks on result:
other: test substance
Irritation parameter:
other: optical density [wavelength 570 nm]
Basis:
mean
Time point:
other: after 30 minutes exposure
Score:
0.534
Remarks on result:
other: positive control
Irritation parameter:
other: optical density [wavelength 570 nm]
Basis:
mean
Time point:
other: after 30 minutes exposure
Score:
1.856
Remarks on result:
other: negative control
Irritation parameter:
other: tissue viability [% of negative control]
Basis:
mean
Time point:
other: after 30 minutes exposure
Score:
18
Remarks on result:
other: test substance
Irritation parameter:
other: tissue viability [% of negative control]
Time point:
other: after 30 minutes exposure
Score:
29
Remarks on result:
other: positive control
Irritation parameter:
other: tissue viability [% of negative control]
Time point:
other: after 30 minutes exposure
Score:
100
Remarks on result:
other: negative control
Conclusions:
Based on the observed results and applying the evaluation criteria it was concluded, that 2-Morpholinoethanol shows an eye irritation potential in the EpiOcular™ eye irritation test under the test conditions chosen.
Executive summary:

The potential of 2-Morpholinoethanol to cause ocular irritation was assessed by a single topical application of 50 µL of the undiluted test substance to a reconstructed three dimensional human cornea model (EpiOcular™). Two EpiOcular™ tissue samples were incubated with the test substance for 30 minutes followed by a 2-hours post-incubation period. Tissue destruction was determined by measuring the metabolic activity of the tissue after exposure/post-incubation using a colorimetric test. The reduction of mitochondrial dehydrogenase activity, measured by reduced formazan production after incubation with a tetrazolium salt (MTT) was chosen as endpoint. The formazan production of the test-substance treated epidermal tissues is compared to that of negative control tissues. The quotient of the values indicates the relative tissue viability. The EpiOcular™ eye irritation test showed the following results: The test substance is able to reduce MTT directly. Therefore an additional MTT reduction control was introduced. However, this ability of direct MTT reduction did not impair the study result as demonstrated by the concurrently performed exposure of control tissues inactivated by freezing. The mean viability of the test-substance treated tissues was 18 %. Based on the observed results it was concluded, that 2-Morpholinoethanol shows an eye irritation potential in the EpiOcular™ eye irritation test under the test conditions chosen. The test method does not yet allow for the evaluation of serious eye damage. The result does not exclude a serious eye irritation potential of the test substance. For final assignment of a risk phrase at present, results from another study would be needed (see 63V0386/11A583).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irritating)

Additional information

Skin:

In a primary dermal irritation study Vienna White rabbits (2 males) were dermally exposed to undiluted 2-morpholinoethanol for 1, 5, 15 min and 20 hours. Animals then were observed for 8 days. Irritation was scored after 24 and 48 h. No signs of irritation were observed in two animals after 1, 5 and 15 min exposure on the back. Slight redness was observed in both animals after 24 h (ear and back) when treated for 20 h. The irritation was reversible within 8 days (BASF, 1968).

Test article 2 -morpholinoethanol (0.5 mL) was applied to shaved skin sites (intact and abraded, occluded) of 6 young adult New Zealand White rabbits weighing 2 to 3 kg. Dermal scoring of erythema and edema on both skin sites at 24 and 72 hours, and calculation of the primary irritation index, was according to Draize (1959). Study duration was 4 days. Draize Primary Irritation Index was determined to be 0.66 of 8.0. Thus the substance was considered to be minimally irritating. Slight to well-defined erythema was observed only at the 24 h observations. Very slight edema was observed at the 72 hour observations. All scores returned to normal by the day 4 observations (Papciak, 1990).

Eye:

The potential of the test substance to cause ocular irritation was assessed by a single topical application of 50 µL of the undiluted test substance to a reconstructed three dimensional human cornea model (EpiOcular™). Tissue destruction was determined by measuring the metabolic activity of the tissue after exposure/post-incubation using a colorimetric test. The reduction of mitochondrial dehydrogenase activity, measured by reduced formazan production after incubation with a tetrazolium salt (MTT) was chosen as endpoint. The formazan production of the test-substance treated epidermal tissues is compared to that of negative control tissues. The quotient of the values indicates the relative tissue viability. The mean viability of the test-substance treated tissues was 18 %. Based on the observed results it was concluded, that 2-Morpholinoethanol shows an eye irritation potential in the EpiOcular™ eye irritation test under the test conditions chosen. Since the test method does not yet allow for the evaluation of serious eye damage, an additional in vitro study according to OECD 437 (BCOP) was performed.

Therefore, the potential of the test substance to cause serious damage to the eyes was assessed by a single topical application of 750 µL of the undiluted test substance to theepithelial surface of isolated bovine corneas. Three corneas were treated with the test substance for 10 minutes followed by a 2-hours post-incubation period. Corneal opacity was measured quantitatively as the amount of light transmission through the cornea. Permeability was measured quantitatively as the amount of sodium fluorescein dye that passes across the full thickness of the cornea. Both measurements were used to calculate an In Vitro Irritancy Score of the test substancerelative to the control corneas.

Based on the observed results it was concluded, that 2-Morpholinoethanol does not cause serious eye damagein the Bovine Corneal Opacity and Permeability Test (BCOP Test) under the test conditions chosen.

To conclude, taking the results of these two in vitro studies into consideration, 2-Morpholinoethanol shows an eye irritating potential.

In a primary eye irritation study which support the findings of the in vitro studies, 0.05 mL of undiluted 2-morpholinoethanol was instilled into the conjunctival sac of the left eye of Vienna white rabbits (1/sex). Animals then were observed for 8 days. Irritation was scored after 24/48/72 h and 8 days.

In both animals tested a slight to severe redness and edema was observed one hour and 24 hours after treatment. 24 hours after treatment a slight opacity was noted in both animals. All effects were reversible within 8 days after treatment. Mean scores (24/48/72 h) were cornea = 1, iris = 0, conjunctiviae = 1.33, chemosis = 0.33.

In conclusion, the test substance was classified as reversible irritating to the eye (BASF, 1968).

In contrast, the test article 2-morpholinoethanol (0 .1 mL) was administered directly to one eye of 6 young adult New Zealand White rabbits, weighing 2 to 3 kg. Ocular scoring of corneal, iridial, and conjuctival lesions with calculation of an ocular irritation toxicity score was according to Draize et al. (1944). Study duration was 7 days. The Draize ocular irritation score was 5.2 of 110 at 24 hours post treatment. The substance was considered minimally irritating. Slight ocular irritation was observed through 72 hours. All scores returned to normal by day 7. Due to methological deficiencies and non-sufficient documentation, the study results were regarded as not reliable (Papciak, 1990).

Additionally, in a very old, non guideline study the substance was rated eye irritating. In this primary eye irritation study, 0.005 mL of undiluted 2-morpholinoethanol was instilled into the conjunctival sac of albino rabbits for 18-24 hours. Irritation was scored by the method of Smyth and Carpenter (1944). In the study the test substance was rated grade 5 of max. 10, corresponding to severe eye damage. Due to the limitations of this study the data was not taken into account for hazard assessment (Carpenter, 1946).


Effects on eye irritation: irritating

Justification for classification or non-classification

Based on the results obtained in skin irritation studies with rabbits (only slight irritation, reversible within 8 days) the test substance has not to be classified with respect to skin irritation according to Regulation (EC) No 1272/2008 (CLP, GHS).

Based on the results obtained in in vitro tests and in an eye irritation study with rabbits, the test substance has to be classified with respect to eye irritation according to Regulation (EC) No 1272/2008 (CLP, GHS): cat. 2, H319