Palm oil, epoxidized

Eye irritation

Administrative data

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:

August 10, 1981 - August 24, 1981

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

This study meets the criteria laid out in OECD Guideline 405. This read-across is based on the hypothesis that source and target substances have similar toxicological properties because of their structural similarities and they are assumed to have similar toxicokinetic profiles i.e. they are expected to be metabolised in a similar fashion. The target substance Epoxidized Palm Oil (EPO) and source substance Epoxidized Soybean Oil (ESBO) are derived respectively from their raw materials, palm oil and soybean oil, and are of variable composition consisting of fatty acid triglycerides. Both EPO and ESBO are organic UVCB sub-type 1: substances of biological nature that have been modified in chemical processing. Both are manufactured by the reaction of the respective oils with an epoxidizing agent (50-60% hydrogen peroxide at 60-75°C). The olefinic bonds of the oils are converted to epoxy oxirane groups. As Palm oil has lower unsaturated bonds than Soybean Oil, less of the epoxidizing agent hydrogen peroxide is required and thus EPO has a lower epoxidized adduct content than ESBO and is therefore expected to be less chemically reactive. The % epoxidation in ESBO is 6-8% while the % epoxidation in EPO is 2.5-3.5%. The target substance (EPO) and source substance (ESBO) have a structurally similar backbone which is an epoxidized triglyceride structure derived from one glycerol molecule and three fatty acid molecules. Therefore, the source and the target substances share structural similarities with common functional groups and side chains varying in their length and the amount of epoxide groups. The target substance contains 8 fatty acids with the largest components being C16:0 palmitic acid (44%), C18:1 oleic acid (39.2%) and C18:2 linoleic acid (10.1%). The source substance contains 5 fatty acids with the largest components being C16:0 palmitic acid (11.3%), C18:1 oleic acid (39.2%) and C18:2 linoleic acid (55.8%). ESBO does not contain lauric (C12), myristic (C14) and arachidic (C20) acids while they are present in very low amounts in EPO (0.2, 1.1 and 0.3% respectively). Stearic acid (C18:0) is present in both substances at similar levels (4.5% in EPO and 3.4% in ESBO) while α-linolenic acid (C18:3) is present at 0.4% in EPO and 6.4% in ESBO. So, the main component of the triglyceride structure of both EPO and ESBO is C16 (44%; 11.3%) and C18 (54.2%; 88.7%). The data gap for the target substance EPO is an in vivo eye irritation study (Annex VIII, 8.2.1). No reliable data on in vivo eye irritation of EPO is available. Therefore, read-across from an existing in vivo eye irritation study of the source substance, ESBO, is considered as an appropriate adaptation to the standard information requirements of Annex VIII, 8.2.1 of the REACH Regulation for the target substance, in accordance with the provisions of Annex XI, 1.5 of the REACH Regulation.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1981

Report date:

1981

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Principles of method if other than guideline:

Procedure used was that described in the Proposed Guideline of the United States Environmental Protection Agency (EPA) S 163.81 - 4 "Primary eye irritation study", Federal Register Vol. 43, No. 163, August 2, 1978 that were in draft at time of study conduct and subsequently adopted.

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

ESBO

IUPAC Name:

ESBO

Constituent 2

Reference substance name:

Epoxidised Soybean Oil

IUPAC Name:

Epoxidised Soybean Oil

Constituent 3

Reference substance name:

Soybean oil, epoxidized

EC Number:

232-391-0

EC Name:

Soybean oil, epoxidized

Cas Number:

8013-07-8

IUPAC Name:

8013-07-8

Details on test material:

- Name of test material (as cited in study report): TK 11'278
- Physical state: Liquid
- Lot/batch No.: prod. Oct. 80

Test animals / tissue source

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

The test was performed on 3 male and 6 female New Zealand White rabbits bred and raised on the premises weighing 2 to 3 kgs. The animals were housed individually in metal cages, numbered by ear tags, were kept at constant room temperature of 22 ± 2°C, at a relative humidity of 55 ± 10% and on a 12 hour light cycle day. The animals had ad libitum access to standard rabbit food and water.

Prior to treatment they were adapted to the laboratory for a minimum of 4 days. Only rabbits with normal opthalmic findings were used for these tests.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

other: the untreated eye of each rabbit served as the inherent control

Amount / concentration applied:

0.1 mL

Duration of treatment / exposure:

In three of the nine rabbits approximately 30 seconds after treatment the treated eye was flushed with 10 mL of physiological saline.

Observation period (in vivo):

14 days

Number of animals or in vitro replicates:

9 - three rinsed and six not rinsed after installation

Details on study design:

0.1 mL of undiluted test material was instilled into the conjunctival sac of the left eye of the rabbits and the lids were gently closed for a few seconds. The right eye was not treated and served as an untreated control. In three of the nine rabbits approximately 30 seconds after treatment the treated eye was flushed with 10 ml of physiological saline. The eye irritation was appraised with a slit-lamp on day 1, 2, 3, 4, 7, 10 and 14 and was scored for each individual rabbit.

As the test compound is not soluble in water, an assessment of pH was not performed prior to dosing.

Results and discussion

In vivo

Results

Results 1

Basis:

mean

Remarks:

Kay and Callendra system

Time point:

other: 24 hours

Score:

4.7

Max. score:

20

Reversibility:

fully reversible within: 10 days

Results 2

Basis:

mean

Remarks:

Kay and Callendra system

Time point:

other: 24 hours

Score:

3.3

Max. score:

20

Reversibility:

other: after 7 days only one of the three animals had slight conjunctival redness

Irritant / corrosive response data:

The ocular reactions were assessed by the Kay and Callendra system, from which a PII was derived. For classification purposes the results were re-calculated using a standard Draize score deriving a mean for the reactions over 24-72 hours.
On this basis the mean results for the rinsed eyes (scores for six rabbits) were -
corneal opacity = 0.0
Iris response = 0.0
Conjunctival redness = 1.06
Conjunctival swelling/chemosis = 0.61
and for the rinsed eyes the mean values for 24-72 hours were:
corneal opacity = 0.0
Iris response = 0.0
Conjunctival redness = 0.89
Conjunctival swelling/chemosis = 0.56.
No corneal or iridial changes were evident for any of the nine rabbits. Conjunctival irritation did not exceed diffuse crimson coloration and had resolved in all unrinsed rabbits by Day 10. reactions persisted for one of the rinsed eyes to Day 14. Conjunctival chemosis had largely resolved by Day 4 for all nine rabbits although slight reactions persisted for one, unrinsed, eye to Day 7.

Other effects:

No symptoms of systemic intoxication were observed throughout the whole test period.

Any other information on results incl. tables

Table: 1 Calculation of the primary eye irritation index

Time after exposure days	Mean reaction score
	Unrinsed eyes (A)			Rinsed eyes (B)
	Cornea	iris	conjunctiva	cornea	iris	conjunctiva
1	0	0	4.7	0	0	3.3
2	0	0	3.3	0	0	2.7
3	0	0	2	0	0	2.7
4	0	0	1.3	0	0	1.3
7	0	0	1	0	0	0.7
10	0	0	0	0	0	0.7
14	0	0	0	0	0	0.7

The ocular reactions were assessed by the Kay and Callendra system, from which a PII was derived. For classification purposes the results were re-calculated using a standard Draize score deriving a mean for the reactions over 24-72 hours. On this basis the mean results for the rinsed eyes (scores for six rabbits) were - corneal opacity = 0.0 Iris response = 0.0 Conjunctival redness = 1.06 Conjunctival swelling/chemosis = 0.61 and for the rinsed eyes the mean values for 24-72 hours were: corneal opacity = 0.0 Iris response = 0.0 Conjunctival redness = 0.89 Conjunctival swelling/chemosis = 0.56. No corneal or iridial changes were evident for any of the nine rabbits. Conjunctival irritation did not exceed diffuse crimson coloration and had resolved in all unrinsed rabbits by Day 10. reactions persisted for one of the rinsed eyes to Day 14. Conjunctival chemosis had largely resolved by Day 4 for all nine rabbits although slight reactions persisted for one, unrinsed, eye to Day 7.

Read-Across Justification: Full report is attached in study summary

3 Analogue approach justification

3.1. Physicochemical properties

Physicochemical data shows that the physicochemical properties of the target and source substances are similar as outlined in the data matrix (Table 3). Both are liquids and the structural differences in the side chains do not significantly influence the physicochemical properties of both substances, i.e. vapour pressure, water solubility and partition co-efficient (log Pow). Both substances are highly insoluble in water; <0.01 mg/L at 30°C for EPO and <0.02 µg/L at 20°C (calculated) for ESBO. Neither of the substances is volatile, with a vapour pressure of 0.5 kPa at 25°C for EPO and 8.4 x 10-8 Pa at 25°C for ESBO. Both substances are highly lipophilic; mean Log Pow >6.2 at 25°C for ESBO and log Pow >10 (calculated) for EPO.

3.2. Toxicokinetics

No specific experimental data on absorption, distribution, metabolism or excretion is available for the source or target substance. Read-across was performed for all human health toxicity endpoints to Epoxidised Soybean Oil (ESBO, CAS No. 8013-07-8). An OECD SIDS report is available that concluded on a proposed metabolic pathway for epoxidised fatty acid esters, including ESBO (OECD, 2006). The toxicokinetic analysis is based on physicochemical data from EPO, read-across ESBO data from in vivo animal models and the OECD SIDS report in the literature (OECD, 2006).

Physicochemical data

The molecular weight of EPO and ESBO is > 500 g/mol and is not in the range for favourable oral absorption (<500 g/mol). The calculated log Pow of EPO (>10) and mean Log Pow >6.2 at 25°C for ESBO indicate they are highly lipophilic and water solubility (<0.01 mg/L at 30°C) for EPO and <0.02 µg/L at 20°C (calculated) for ESBO indicates they are both insoluble in water. These characteristics will not facilitate transport of EPO or ESBO via passive diffusion. Based on its high lipophilicity, absorption of EPO and ESBO via the lymphatic system through micellular solubilisation by bile salts is likely, similar to other vegetable oils. Insolubility in water of both EPO and ESBO indicates low dermal uptake while the high log Pow values for both are an indication for a high uptake into the stratum corneum but little or no penetration into the lower layers of the epidermis and dermis. Overall, the physical state, molecular weight, calculated log Pow and water insolubility indicate that dermal absorption of EPO and ESBO is unlikely. Due to the low vapour pressure of EPO (0.5 kPa at 25°C) and ESBO (8.4 x 10-8 Pa at 25°C) and physical state (liquid), exposure via the inhalation route of both is expected to be negligible. Based on the information available for the analogue ESBO in the OECD SIDS report (OECD, 2006; see ‘Other data in the literature’), during metabolism, breakdown products are produced that are more water soluble than the parent substance i.e. free fatty acids, so it is expected that any EPO metabolites will be excreted in the urine.

Other data in the literature

The OECD produced a report on Epoxidised Oils and Derivatives in 2006 (OECD, 2006), which included ESBO. The OECD SIDS concluded that epoxidised fatty acid esters, such as ESBO and therefore we assume EPO, produce metabolic products with similar primary constituents as other vegetable oils and are assumed to have similar metabolic pathways e.g. breakdown in the gastrointestinal tract by esterases (pancreatic lipase) to epoxidised fatty acids and glycerol which enter the normal nutritional pools (JECFA, 1974). Pancreatic lipase works at the oil/water interface since triglycerides are insoluble. During metabolism in the GI tract, pancreatic lipase preferentially hydrolyses triglycerides to release the free fatty acids from the SN-1 and SN-3 (terminal) positions of the glycerol backbone. The other products of metabolism are mono- and di-glycerides (OECD, 2006). The EFSA Panel on Contaminants in the Food Chain agreed with this assessment for ESBO in 2011 (EFSA, 2011). Overall, the proposed metabolic pathway for ESBO is enzymatic breakdown to epoxidised fatty acids and glycerol; a similar pathway is predicted for EPO. Based on the information available for the analogue ESBO in the OECD SIDS report, during metabolism, breakdown products are produced that are more water soluble than the parent substance i.e. free fatty acids, so it is expected that any EPO metabolites will be excreted in the urine.

Available in vivo toxicological data

The in vivo read-across data from ESBO indicate no adverse effects if oral absorption occurs (acute oral LD50 of >5,000 mg/kg (3), 2 year combined chronic/carcinogenicity toxicity study

NOEL (male) of 1000 mg/kg bw/day and NOEL (female) of 1400 mg/kg bw/day (13), pre-natal developmental toxicity maternal/developmental NOAEL of 1000 mg/kg bw/day (14). The in vivo read-across data from ESBO indicates is poorly absorbed via the dermal route (slightly irritating in the in vivo skin irritation study in rabbits (6) and non-sensitising in Guinea pig maximization test (8)). Any significant dermal absorption is unlikely.

3.3. Comparison of data from human health endpoints

3.3.1 Toxicity data of the target and source substances

There is no existing human health toxicity data for the target substance, EPO. As is presented in the data matrix (Table 3), the acute oral (LD 50 (male/female) >5,000 mg/kg bw) and acute dermal toxicity data (LD50 >20mL/kg bw) shows very low toxicity for the source chemical, ESBO, in rats and rabbits. The source chemical is slightly irritating to the skin in rabbits. The source substance is not a skin sensitizer in the guinea pig maximization test. In the in vitro bacterial reverse mutation study (Ames test), in vitro chromosomal aberration study and in vitro gene mutation study in mammalian cells, the source substance ESBO was negative in the presence and absence of metabolic activation. The source substance ESBO is not genotoxic. In a 2 year combined chronic toxicity/carcinogenicity study in rats for 104 weeks, a NOEL value of 1000 mg/kg bw/day (male) and 1400 mg/kg bw/day (female) was derived. In a pre-natal developmental toxicity study in rats, the NOAEL (maternotoxic, embryofetal) was 1000 mg/kg bw/day with no adverse effects noted. In accordance with Column 2 of ANNEX IX of the REACH Regulation, a two generation reproductive toxicity study does not need not to be conducted as the existing read across combined chronic toxicity/carcinogenicity study from ESBO is available and does not indicate clear adverse effects on reproductive organs or tissues.

The data gap for the target substance EPO is an in vivo eye irritation study (Annex VIII, 8.2.1). No reliable data on in vivo eye irritation of EPO is available. Therefore, read-across from an existing in vivo eye irritation study of the source substance, ESBO, is considered as an appropriate adaptation to the standard information requirements of Annex VIII, 8.2.1 of the REACH Regulation for the target substance, in accordance with the provisions of Annex XI, 1.5 of the REACH Regulation. The read-across oral acute toxicity study (RL2) was according to a test guideline that was equivalent or similar to OECD 405. In a primary eye irritation study, 0.1 mL of ESBO (undiluted) was instilled into the conjunctival sac of the left eye of 3 male and 6 female New Zealand white rabbits. In three of the nine rabbits approximately 30 seconds after treatment, the treated eye was flushed with 10 mL of physiological saline. Animals were then observed for 14 days. Irritation was scored by the method of Kay and Callendra from which a PII was derived. For classification purposes the results were re-calculated using a standard Draize score deriving a mean for the reactions over 24-72 hours.In unrinsed eyes, no irritation of cornea and iris was observed at any reading. Irritation was seen only in the conjunctiva with a maximum mean a score of 4.7 after 24 hours. After 10 days all irritation had disappeared. In rinsed eyes, no irritation of cornea and iris was observed at any reading. Irritation was seen only in the conjunctiva with a maximum mean score of 3.3 after 24 hours. After 7 days onwards only one of the three animals had a slight conjunctival redness. Overall, ESBO was found to cause slight irritation (conjunctiva) when applied to the rabbit eye mucosa. EPO is predicted to have a similar result.

3.3.2 Effect of structural differences between target and source chemical

The target substance consists of 8 fatty acids with the largest components being C16:0 palmitic acid (44%), C18:1 oleic acid (39.2%) and C18:2 linoleic acid (10.1%). The source substance consists of 5 fatty acids with the largest components being C16:0 palmitic acid (11.3%), C18:1 oleic acid (39.2%) and C18:2 linoleic acid (55.8%). ESBO does not contain lauric, myristic and arachidic acids while they are present in very low amounts in EPO (0.2, 1.1 and 0.3% respectively). Stearic acid is present in both substances at similar levels (4.5% in EPO and 3.4% in ESBO) while α-linolenic acid is present at 0.4% in EPO and 6.4% in ESBO. So, the main component of the triglyceride structure of both EPO and ESBO is C16 (44%; 11.3%) and C18 (54.2%; 88.7%). When these epoxidized fatty acid products are released during metabolism (see Section 3.2) they are not expected to have any adverse effects in the body as all are naturally occurring fatty acids and they are all ‘Not Classified’ in the ECHA Classification and Labelling (C&L) inventory (checked 24-08-15). The only exception is α-linolenic acid which is indicated as a skin sensitizer but this is not relevant to ESBO and EPO as the former is negative in the guinea pig maximization test and EPO is also predicted to be negative also. The main structural difference is that palm oil has lower unsaturated bonds than soybean oil therefore less of the epoxidizing agent hydrogen peroxide is required to produce the epoxidized derivatives. EPO has a lower epoxidized adduct content than ESBO and is therefore expected to be less chemically reactive. The % epoxidation in ESBO is 6-8% while the % epoxidation in EPO is 2.5-3.5%.

3.3.3 Classification and labelling

According to the ECHA Classification and Labelling (C&L) inventory, the source substance, ESBO, is ‘Not Classified’ (647 notifiers, joint entry; checked 24-08-15). The target substance, EPO, is not listed in the C&L inventory (checked 24-08-15). Based on the read-across eye irritation study presented for the source substance, EPO does not need to be classified for eye irritation/damage when the criteria outlined in Annex I of 1272/2008/EC are applied.

4. Conclusion

The structural similarities between the source and the target substances and estimated similar toxicokinetics presented above support the read-across hypothesis. The structural differences between the target and source substance are not expected to have an impact on the prediction. Adequate, reliable and available scientific information indicates that using the source substance for read across to the target substance is acceptable.

Therefore, based on the considerations above, it can be concluded that the eye irritation study conducted in rabbits with ESBO is likely to predict the eye irritation of EPO and is considered as adequate to fulfill the information requirement of Annex VIII, 8.2.1

Applicant's summary and conclusion

Interpretation of results:

slightly irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the conditions of this experiement, the test material was found to cause slight conjunctival irritiation when applied to the rabbit eye mucosa:
Unrinsed eyes:
No irritation of cornea and iris was observed at any reading. Irritation was seen only in the conjunctiva with a maximum mean acore of 4.7 after 24 hours. After 10 days all irritation had disappeared.

Rinsed eyes:
No irritation of cornea and iris was observed at any reading. Irritation was seen only in the conjunctiva with a maximum mean score of 3.3 after 24 hours. After 7 days onwards only one of the three animals had a slight conjunctival redness. No symptoms of systemic intoxication were observed throughout the whole test period. According to Directive 67/548/EEC, no classification is warranted. According to Regulation (EC) No. 1272/2008, no classification is warranted.

Executive summary:

The test was performed on three male and six female New Zealand White rabbits. The test material was found to cause the following irritiation when applied to the rabbit eye mucosa: In the unrinsed eyes no irritation of cornea and iris was observed at any reading. Irritation was seen only in the conjunctiva with a maximum mean acore of 4.7 after 24 hours. After 10 days all irritation had disappeared. In the rinsed eyes, no irritation of cornea and iris was observed at any reading. Irritation was seen only in the conjunctiva with a maximum mean score of 3.3 after 24 hours. After 7 days onwards only one of the three animals had a slight conjunctival redness. No symptoms of systemic intoxication were observed throughout the whole test period.

Reactions were re-assessed using EU classification criteria and responses were confirmed as not exceeding the EU thresholds for triggering classification of an ocular irritant.