Petrolatum (petroleum), oxidized, Bu ester

Endpoint summary

Description of key information

SKIN
Not corrosive (in vitro, EpiSkin Model), OECD 431, EU Method B.40, Ágh (2012)
Not irritating (in vivo) (hydrocarbon waxes (petroleum), oxidised), EPA OPPTS 870.2500, Hoff (2002)
EYE
"slightly irritating", (in vitro), ICCVAM Test Method Evaluation Report: Appendix G - ICCVAM recommended HET-CAM Method Protocol (Nov. 2006), Andres (2012)
Not irritating (in vivo) (hydrocarbon waxes (petroleum), oxidised), EPA OPPTS 870.2400, Cerven (2002)

Key value for chemical safety assessment

Skin irritation / corrosion

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

3 December 2001 to 6 December 2001

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, conducted to a valid guideline and was performed under GLP conditions. Since the study was conducted with the read across substance, hydrocarbon waxes (petroleum), oxidised, it has been assigned a reliability score of 2. Read across from this substance is justified on the basis of its similar physical and chemical properties to those of the registered substance. Furthermore, it has a very similar chemical structure; the registered substance is an esterified form of the read across substance.

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2400 (Acute Eye Irritation)

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Millbrook Breeding Lads, Amherst, MA, USA
- Only healthy animals free from ocular damage were selected for this study
- Age at study initiation: approximately 13 weeks
- Weight at study initiation: 3.2 - 3.4 kg
- Housing: Individually in suspended cages
- Diet: Animals fed daily rations of Purina Rabbit Chow, diet # 5321, supplied by Purina Mills, Inc., USA
- Water: as libitum
- Quarantine period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Controlled
- Photoperiod (hrs dark / hrs light): 12 hours light/dark cycle

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 mL/rabbit

Observation period (in vivo):

72 hours

Number of animals or in vitro replicates:

Three female animals

Details on study design:

METHOD
- The test material was placed into the conjunctival sac, formed by gently pulling the lower eyelid away from the eye. The lid was held closed for a second, before the animal was released.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): No.

SCORING SYSTEM: Reactions were assessed according to the Draize scale (1977), which can be seen in table 1.

OBSERVATIONS
-Time points: 1, 24, 48 and 72 hours
- General health was recorded at each time point
- Body weights were recorded before exposure

TOOL USED TO ASSESS SCORE:
- A Mini-Maglite® was during ocular examinations
- Sodium fluorescein dye was used at 24 hours

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

other: 24, 48, 72 hours

Score:

0

Max. score:

80

Remarks on result:

other: No reaction observed

Irritation parameter:

iris score

Basis:

mean

Time point:

other: 24, 48, 72 hours

Score:

0

Max. score:

10

Remarks on result:

other: No reaction observed

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

other: 24, 48, 72 hours

Score:

0.66

Max. score:

20

Reversibility:

fully reversible within: 72 hours

Irritant / corrosive response data:

There was no corneal opacity or iritis noted at any observation period. Conjunctival irritation, noted in 3/3 eyes, cleared within 72 hours.

Other effects:

No abnormal physical signs were noted during the observation period.

Table 2. Results for Ocular Damage

Animal No.	Tissue	Reading	1 hr	24 hrs	48 hrs	72 hrs	Mean Score from 24, 48 and 72 hours
1	Cornea	Opacity	0	0	0	0	0
		Area	0	0	0	0
	Sub-total		0	0	0	0
	Iris		0	0	0	0	0
	Sub-total		0	0	0	0
	Conjunctiva	Redness	2	0	0	0	0
		Chemosis	2	0	0	0	0
		Discharge	2	0	0	0
	Sub-total		12	0	0	0
	Total		12	0	0	0
2	Cornea	Opacity	0	0	0	0	0
		Area	0	0	0	0
	Sub-total		0	0	0	0
	Iris		0	0	0	0	0
	Sub-total		0	0	0	0
	Conjunctiva	Redness	2	1	0	0	0.33
		Chemosis	2	0	0	0	0
		Discharge	2	0	0	0
	Sub-total		12	2	0	0
	Total		12	2	0	0
3	Cornea	Opacity	0	0	0	0	0
		Area	0	0	0	0
	Sub-total		0	0	0	0
	Iris		0	0	0	0	0
	Sub-total		0	0	0	0
	Conjunctiva	Redness	2	1	1	0	0.66
		Chemosis	2	0	0	0	0
		Discharge	2	0	0	0
	Sub-total		12	2	2	0
	Total		12	2	2	0

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the conditions of the study, the test material only elicited slight reactions in any of the animals during the course of the study that meant the test material does not require classification as an eye irritant.

Executive summary:

In a GLP compliant study performed according to the standardised guideline EPA OPPTS 870.2440, the ocular irritation caused by exposure to the test material was assessed in three rabbits. 0.1 mL of the test material was administered into one eye of each of three rabbits, the other eye remained untreated and served as a control. Initially some mild irritation (chemosis and discharge) was observed, however this did not persist past 1 hour. Conjunctival redness was the parameter which persisted past the 1 hour observation time point. Conjunctival redness was assigned an average score of 0.33 according to the Draize scale (1977), this was completely reversible within 72 hours. Under the conditions of the study, the test material only elicited slight reactions in any of the animals during the course of the study that meant the test material does not require classification as an eye irritant.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

SKIN

In the in vitro study, the skin corrosion potential of the test material (butyl ester of different oxidised hydrocarbon fractions) was investigated under GLP conditions and in accordance with standardised guidelines OECD 431 and EU Method B.40 using the EPISKIN Model. During the study, duplicate tissues were treated with the test material for an exposure period of 4 hours. At the end of the exposure period the test material was rinsed with PBS 1x solution. The viability of each e3podermis was assessed by incubating the tissues for 3 hours with MTT solution at 37 °C in 5 % CO2 protected from light. The formazan precipitated and was then extracted using acidified isopropanol and quantified spectrophotometrically. NaCl (9 g/L saline) and glacial acetic acid treated epidermis were used as negative and positive controls, respectively. From each treated tissue, viability was expressed as a % relative to negative control.

The test material did not show significantly reduced cell viability in comparison to the negative control after 4 hours of exposure. Both individual tissue viabilities were above 35 % of the mean negative control value. The average test material treated viability was 103 %. Positive and negative controls showed the expected cell viability values within acceptable limits and so the experiment was considered to be valid. Under the conditions of the study, the test material was considered to be non-corrosive to skin.

 

In the in vivo study, the skin irritation potential of the test material (hydrocarbon waxes (petroleum), oxidised) was investigated under GLP conditions and in accordance with the standardised guideline EPA OPPTS 870.2500. During the study three rabbits were exposed to the test material for a period of 4 hours. The average dermal reaction, taken from observations at 24, 48 and 72, were scored according to the Draize (1977) scale, and recorded. The mean score for erythema was 0.33 while the mean score for edema was 0.11; all observed signs of irritation completely reversed within 48 hours. Under the conditions of the study, the test material did not elicit any reaction in any of the animals during the course of the study that meant the test material required classification as a skin irritant according to Regulation (EC) 1272/2008. Therefore the results have been interpreted as "not classified" according to EU criteria.

 

Both studies were performed under GLP conditions and in accordance with standardised guidelines with a sufficient level of detail to assess the accuracy and reliability of the data. Since the in vitro study was performed on the registered substance it was assigned a relibaility score of 1 according to the criteria of Klimisch. Since the in vivo study was conducted with the read across substance, hydrocarbon waxes (petroleum), oxidised, it was assigned a reliability score of 2.

 

 

EYE

In the in vitro study, the eye irritation potential of the test material (butyl ester of different oxidised hydrocarbon fractions) was investigated under GLP conditions and in accordance with the ICCVAM Test Method Evaluation Report: Appendix G - ICCVAM recommended HET-CAM Method Protocol (Nov. 2006). This study assessed the irritating potential of a test material by detection of damages in blood vessels under the chorionallantoic membrane of incubated chicken eggs (9 days old). Observation time was five minutes at room temperature. Physiological sodium chloride solution was uses as negative control, sodium dodecyl sulphate (1 % solution) and sodium hydroxide (0.1 N solution) were used as positive controls. The positive controls induced a severe irritation on the blood vessels. The negative control showed no irritation. The irritation score for the positive controls lay above the demanded threshold. The test material was tested pure and gave a mean irritation score of 3.55, corresponding to a classification as slightly irritating according to the classification scheme employed in the study.

 

In the in vivo study, the eye irritation potential of the test material (hydrocarbon waxes (petroleum), oxidised) was investigated under GLP conditions and in accordance with the standardised guideline EPA OPPTS 870.2440. During the study 0.1 mL of the test material was administered into one eye of each of three rabbits, the other eye remained untreated and served as a control. Initially some mild irritation (chemosis and discharge) was observed, however this did not persist past 1 hour. Conjunctival redness was the parameter which persisted past the 1 hour observation time point. Conjunctival redness was assigned an average score of 0.33 according to the Draize scale (1977), this was completely reversible within 72 hours. Under the conditions of the study, the test material only elicited slight reactions in any of the animals during the course of the study that meant the test material does not require classification as an eye irritant.

 

Both studies were performed under GLP conditions. The in vivo study was conducted in accordance with standardised guidelines while the in vitro study was performed to sound scientific principles. Both reports contained a sufficient level of detail to assess the accuracy and reliability of the data. While the in vitro study was performed with the registered substance, it was performed to a method currently undergoing validation, the in vivo study was performed with the read across substance, hydrocarbon waxes (petroleum), oxidised. Both studies were therefore assigned a relibaility score of 2 according to the criteria of Klimisch (1997).

The in vivo eye irritation study that is available is considered to represent a worst case scenario since it was conducted with the read across substance (hydrocarbon waxes (petroleum), oxidised) which has an acid number of 24 - 40 whereas the registered substance has a lower acid number of 7 -13 due to the esterification of the hydrocarbon chains.

Justification for selection of skin irritation / corrosion endpoint:
No single study is selected since two key studies are available to address skin irritation and corrosion.

Justification for selection of eye irritation endpoint:
Cerven (2002) was selected as the key study since it is a GLP study conducted to a standardised in vivo guideline. Conversely, the method followed during the in vitro test has not yet been validated.

Justification for classification or non-classification

Skin

In accordance with the criteria for classification and labelling as defined in Regulation (EC) No 1272/2008 (CLP) and Directive 67/548/EEC (DSD), the substance does not require classification for skin irritation and corrosion.

Eye

In accordance with the criteria for classification as defined in Annex I, Regulation 1272/2008 and Directive 67/548/EEC (DSD), the substance does not require classification for eye irritation.