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EC number: 289-904-6 | CAS number: 90045-43-5 Extractives and their physically modified derivatives such as tinctures, concretes, absolutes, essential oils, oleoresins, terpenes, terpene-free fractions, distillates, residues, etc., obtained from Citrus paradisi M., Rutaceae.
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
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Endpoint summary
Administrative data
Description of key information
Key value for chemical safety assessment
Skin sensitisation
Link to relevant study records
- Endpoint:
- skin sensitisation: in vivo (non-LLNA)
- Type of information:
- experimental study
- Adequacy of study:
- supporting study
- Study period:
- June 20, 1995 - July 28, 1995
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: The study was performed in accordance with internal standard operating procedures, and internally audited by a Quality Assurance Unit.
- Reason / purpose for cross-reference:
- reference to same study
- Reason / purpose for cross-reference:
- reference to other study
- Qualifier:
- according to guideline
- Guideline:
- other: method of Ritz and Buehler 1980
- Deviations:
- yes
- Principles of method if other than guideline:
- The potential of Grapefruit oil to produce delayed contact hypersensitivity in guinea pigs was evaluated using an adaptation of the method of Ritz and Buehler (Ritz HL and Buehler EV. Current Concepts in Cutaneous Toxicity, ed. Drill VA and Lazar T. (Academic Press, 1980) pp 25-40).
- GLP compliance:
- yes
- Type of study:
- Buehler test
- Species:
- guinea pig
- Strain:
- Hartley
- Sex:
- male/female
- Details on test animals and environmental conditions:
- TEST ANIMALS
- Source: Harlan Sprague Dawley, inc.
- Weight at study initiation: 405 - 478 grams at start of the primary irritation phase; 352 - 487 grams at start of the induction phase.
- Housing: individually housed in wire mesh suspension cages
- Diet: Teklad Guinea Pig Diet ad libitum
- Water: ad libitum
- Acclimation period: at least four days
ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12-hour light / 12-hour dark cycle - Route:
- epicutaneous, occlusive
- Vehicle:
- other: diethyl phthalate
- Concentration / amount:
- primary irritation phase: undiluted, 50%, 25%, 10%, 5%, 2.5%, 1%, 0.5%
induction phase: 25%
challenge phase: 5%, 1.5%, 0.5% - Route:
- epicutaneous, occlusive
- Vehicle:
- other: diethyl phthalate
- Concentration / amount:
- primary irritation phase: undiluted, 50%, 25%, 10%, 5%, 2.5%, 1%, 0.5%
induction phase: 25%
challenge phase: 5%, 1.5%, 0.5% - No. of animals per dose:
- primary irritation phase: 8 animals
main test: 20 test animals, 10 vehicle control animals - Details on study design:
- RANGE FINDING TESTS: Irritation screening
- Concentrations: undiluted, 50%, 25%, 10%, 5%, 2.5%, 1%, 0.5%
- Site: back
MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 3
- Exposure period: three weeks
- Test groups: 25% w/v formulation in diethyl phthalate
- Control group: undiluted diethyl phthalate
- Site: left shoulder
- Frequency of applications: interval between exposures varied from 6 to 9 days
- Duration: 6 hours (approximately)
- Concentrations: 25% test substance
B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: two weeks after last induction exposure
- Exposure period: 6 hours
- Test groups: same 20 test animals as during induction period
- Control group: same 10 control animals as during induction period
- Site: previously untreated test sites
- Concentrations: 5%, 1.5%, and 0.5% test substance in diethyl phthalate for both the test animals and the control animals; and undiluted diethyl phthalate for the control animals
- Evaluation (hr after challenge): the day following primary challenge exposure, evaluation was repeated the following day. - Challenge controls:
- Animals receiving undiluted diethyl phthalate during the induction period and different concentrations of test substance during the challenge period.
- Positive control substance(s):
- no
- Positive control results:
- not applicable
Historical positive control data were added to the study report. - Reading:
- 1st reading
- Hours after challenge:
- 24
- Group:
- negative control
- Dose level:
- 5 %
- No. with + reactions:
- 9
- Total no. in group:
- 10
- Clinical observations:
- 9 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 5 %. No with. + reactions: 9.0. Total no. in groups: 10.0. Clinical observations: 9 animals showed slight, patchy erythema.
- Reading:
- 1st reading
- Hours after challenge:
- 24
- Group:
- negative control
- Dose level:
- 1.5 %
- No. with + reactions:
- 9
- Total no. in group:
- 10
- Clinical observations:
- 9 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 1.5 %. No with. + reactions: 9.0. Total no. in groups: 10.0. Clinical observations: 9 animals showed slight, patchy erythema.
- Reading:
- 1st reading
- Hours after challenge:
- 24
- Group:
- negative control
- Dose level:
- 0.5 %
- No. with + reactions:
- 9
- Total no. in group:
- 10
- Clinical observations:
- 9 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 0.5 %. No with. + reactions: 9.0. Total no. in groups: 10.0. Clinical observations: 9 animals showed slight, patchy erythema.
- Reading:
- 1st reading
- Hours after challenge:
- 24
- Group:
- negative control
- Dose level:
- 0
- No. with + reactions:
- 8
- Total no. in group:
- 10
- Clinical observations:
- 8 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 0. No with. + reactions: 8.0. Total no. in groups: 10.0. Clinical observations: 8 animals showed slight, patchy erythema.
- Reading:
- 1st reading
- Hours after challenge:
- 24
- Group:
- test chemical
- Dose level:
- 5 %
- No. with + reactions:
- 17
- Total no. in group:
- 19
- Clinical observations:
- 17 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 5 %. No with. + reactions: 17.0. Total no. in groups: 19.0. Clinical observations: 17 animals showed slight, patchy erythema.
- Reading:
- 1st reading
- Hours after challenge:
- 24
- Group:
- test chemical
- Dose level:
- 1.5 %
- No. with + reactions:
- 18
- Total no. in group:
- 19
- Clinical observations:
- 18 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 1.5 %. No with. + reactions: 18.0. Total no. in groups: 19.0. Clinical observations: 18 animals showed slight, patchy erythema.
- Reading:
- 1st reading
- Hours after challenge:
- 24
- Group:
- test chemical
- Dose level:
- 0.5 %
- No. with + reactions:
- 11
- Total no. in group:
- 19
- Clinical observations:
- 11 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 0.5 %. No with. + reactions: 11.0. Total no. in groups: 19.0. Clinical observations: 11 animals showed slight, patchy erythema.
- Reading:
- 2nd reading
- Hours after challenge:
- 48
- Group:
- negative control
- Dose level:
- 5.0 %
- No. with + reactions:
- 8
- Total no. in group:
- 10
- Clinical observations:
- 8 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 5.0 %. No with. + reactions: 8.0. Total no. in groups: 10.0. Clinical observations: 8 animals showed slight, patchy erythema.
- Reading:
- 2nd reading
- Hours after challenge:
- 48
- Group:
- negative control
- Dose level:
- 1.5 %
- No. with + reactions:
- 6
- Total no. in group:
- 10
- Clinical observations:
- 6 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 1.5 %. No with. + reactions: 6.0. Total no. in groups: 10.0. Clinical observations: 6 animals showed slight, patchy erythema.
- Reading:
- 2nd reading
- Hours after challenge:
- 48
- Group:
- negative control
- Dose level:
- 0.5 %
- No. with + reactions:
- 6
- Total no. in group:
- 10
- Clinical observations:
- 6 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 0.5 %. No with. + reactions: 6.0. Total no. in groups: 10.0. Clinical observations: 6 animals showed slight, patchy erythema.
- Reading:
- 2nd reading
- Hours after challenge:
- 48
- Group:
- negative control
- Dose level:
- 0
- No. with + reactions:
- 5
- Total no. in group:
- 10
- Clinical observations:
- 5 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 0. No with. + reactions: 5.0. Total no. in groups: 10.0. Clinical observations: 5 animals showed slight, patchy erythema.
- Reading:
- 2nd reading
- Hours after challenge:
- 48
- Group:
- test chemical
- Dose level:
- 5 %
- No. with + reactions:
- 10
- Total no. in group:
- 19
- Clinical observations:
- 10 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 5 %. No with. + reactions: 10.0. Total no. in groups: 19.0. Clinical observations: 10 animals showed slight, patchy erythema.
- Reading:
- 2nd reading
- Hours after challenge:
- 48
- Group:
- test chemical
- Dose level:
- 1.5 %
- No. with + reactions:
- 10
- Total no. in group:
- 19
- Clinical observations:
- 10 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 1.5 %. No with. + reactions: 10.0. Total no. in groups: 19.0. Clinical observations: 10 animals showed slight, patchy erythema.
- Reading:
- 2nd reading
- Hours after challenge:
- 48
- Group:
- test chemical
- Dose level:
- 0.5 %
- No. with + reactions:
- 8
- Total no. in group:
- 19
- Clinical observations:
- 8 animals showed slight, patchy erythema
- Remarks on result:
- other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 0.5 %. No with. + reactions: 8.0. Total no. in groups: 19.0. Clinical observations: 8 animals showed slight, patchy erythema.
- Interpretation of results:
- other: not sensitising
- Remarks:
- Based on CLP criteria
- Conclusions:
- The test substance grapefruit oil showed no sensitising properties in guinea pigs under the conditions of this test.
- Executive summary:
The potential of grapefruit oil (25% w/v in diethyl phtalate), to produce delayed contact hypersensitiviy in guinea pigs was evaluated using an adaptation of the method of Ritz and Buehler. Following primary challenge using grapefruit oil as 5%, 1.5%, and 0.5% w/v formulations in diethyl phtalate, responses in the test group were compared to those of the vehicle control group. The incidence of test animals exhibiting a positive response to levels of 5%, 1.5%, and 0.5% test substance was 0 of 19 for all concentrations. These data indicate that sensitisation to grapefruit oil had not been induced. Based on the EU criteria outlined in 1272/2008/EC and 67/548/EEC, grapefruit oil 25% is not a sensitiser.
- Endpoint:
- skin sensitisation: in vivo (LLNA)
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- From May 07 to 18, 2004
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: see 'Remark'
- Remarks:
- GLP study conducted in compliance with OECD guideline 429 with minor deviations: no certificate of analysis of the test substance. Due to the read-across purpose it was given a Klimisch 2 rating, in accordance with the ECHA Practical guide #6 on the reporting of read-across in IUCLID.
- Justification for type of information:
- The justification for read across is provided in the attached background material of the chapter summary.
- Reason / purpose for cross-reference:
- reference to same study
- Reason / purpose for cross-reference:
- reference to other study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
- Deviations:
- yes
- Remarks:
- no certificate of analysis of the test substance
- Principles of method if other than guideline:
- Not applicable
- GLP compliance:
- yes
- Type of study:
- mouse local lymph node assay (LLNA)
- Species:
- mouse
- Strain:
- other: CBA/Ca
- Sex:
- female
- Details on test animals and environmental conditions:
- TEST ANIMALS
- Source: Charles River UK Ltd, UK
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 17.7-20 g
- Housing: Housed in a group of 4 mice (maximum) per cage
- Diet (e.g. ad libitum): Diet (RM1) (Special Diet Services Limited, Witham, UK), ad libitum
- Water (e.g. ad libitum): Mains water, ad libitum
- Acclimation period: 5 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 °C
- Humidity (%): 30-70%
- Air changes (per hour): 15/hour (minimum)
- Photoperiod (hours dark / hours light): 12 hours dark / 12 hours light - Vehicle:
- other: ethanol/diethyl phthalate (3: 1 v/v)
- Concentration:
- 0, 10, 25, 50, 75 or 100% v/v in ethanol/diethyl phthalate (3: 1 v/v)
- No. of animals per dose:
- Four females
- Details on study design:
- MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Local Lymph Node Assay
- Criteria used to consider a positive response: One or more concentrations of the test substance should elicit a 3-fold or greater increase in isotope incorporation relative to the vehicle control group.
TREATMENT PREPARATION AND ADMINISTRATION: 25 µL of test material at concentrations of 0 (vehicle control), 10, 25, 50, 75 or 100% v/v in ethanol/diethyl phthalate (3: 1 v/v) were applied to the dorsal surface of each ear on Days 1, 2 and 3. On Day 6, 250 μL phosphate buffered saline (PBS) containing 20 μCi of 3H-methyl thymidine (specific gravity: 2.0 Ci/mmol) was injected into the tail vein of each experimental mouse. Five hours later, all mice were killed by inhalation of halothane vapour followed by cervical dislocation and the draining auricular lymph node of each ear was excised into PBS. A single cell suspension of lymph node cells (LNC) was prepared by gentle separation through 200-mesh stainless steel gauze. LNC were washed thrice with 10 mL of PBS and precipitated with 5% trichloroacetic acid (TCA) at 4 °C. Pellets were re-suspended in 1 mL TCA and transferred to scintillation vials containing 10 mL of scintillation fluid (Optiphase) for 3H-counting. The number of radioactive disintegrations per minute (DPM) was measured using a Packard Tri-Carb Liquid Scintillation Counter. - Positive control substance(s):
- hexyl cinnamic aldehyde (CAS No 101-86-0)
- Statistics:
- No data
- Positive control results:
- Mean DPM
- Vehicle (acetone/olive oil 4:1 v/v) = 2263
- Test material (5 % w/v) = 5496
- Test material (10 % w/v) = 7571
- Test material (25 % w/v) = 24752
Stimulation Index
- Test material (5 % w/v) = 2.4
- Test material (10 % w/v) = 3.3
- Test material (25 % w/v) = 10.9 - Parameter:
- other: disintegrations per minute (DPM)
- Remarks on result:
- other: - Vehicle = 2511 - Test material (10% v/v) = 3319 - Test material (25% v/v) = 8554 - Test material (50% v/v) = 9916 - Test material (75% v/v) = 22063 - Test material (100% v/v) = 16259
- Key result
- Parameter:
- SI
- Value:
- 1.3
- Test group / Remarks:
- 10% v/v
- Key result
- Parameter:
- SI
- Value:
- 3.4
- Test group / Remarks:
- 25% v/v
- Key result
- Parameter:
- SI
- Value:
- 4
- Test group / Remarks:
- 50% v/v
- Interpretation of results:
- other: Sensitising category 1
- Remarks:
- Based on CLP criteria
- Conclusions:
- Based on the results of the study for read-across substance D-limonene, Grapefruit oil "May cause sensitisation by skin contact", according to the criteria of Annex VI to the Directive 67/548/EEC and ‘Category 1’ according to the CLP Regulation (EC) N° (1272-2008).
- Executive summary:
In a local lymph node assay performed in CBA/Ca strain mice according to OECD guideline 429 and in compliance with GLP, groups of mice (4 females/dose) were applied with 25 µL of d-limonene at concentrations of 0 (vehicle control), 10, 25, 50, 75 or 100% v/v in ethanol/diethyl phthalate (3: 1 v/v) to the dorsal surface of each ear for three consecutive days. On Day 6, all animals were injected with 3H-methyl thymidine and after five hours the draining (auricular) lymph nodes were excised and measured for radioactivity expressed as number of disintegrations per minute (DPM). Historic data of hexylcinnamaldehyde (5, 10 and 25 % w/v) in acetone/olive oil (4:1 v/v) was used as the data for positive control group.
Mean DPM for 0, 10, 25, 50, 75 or 100% d-limonene were observed to be 2511, 3319, 8554, 9916, 22063 or 16259 dpm, respectively. Stimulation index for 10, 25, 50, 75 or 100% d-limonene were calculated to be 1.3, 3.4, 4.0, 8.8 or 6.5, respectively. The estimated concentration giving rise to a 3 fold increase in lymphocyte proliferation (EC3) was 22% v/v (5500 µg/cm2). No increase in visual levels of irritancy to the ear skin was observed during the study.
Under the test conditions, d-limonene is classified as ‘R43 May cause sensitisation by skin contact’, according to the criteria of Annex VI to the Directive 67/548/EEC and ‘Category 1’ according to the CLP Regulation (EC) N° (1272-2008).
- Endpoint:
- skin sensitisation: in vivo (LLNA)
- Type of information:
- read-across from supporting substance (structural analogue or surrogate)
- Adequacy of study:
- key study
- Rationale for reliability incl. deficiencies:
- other: see 'Remark'
- Remarks:
- GLP study conducted in compliance with OECD guideline 429 with minor deviations: no certificate of analysis of the test substance. Due to the read-across purpose it was given a Klimisch 2 rating, in accordance with the ECHA Practical guide #6 on the reporting of read-across in IUCLID.
- Justification for type of information:
- The read across justification is presented in the document attached to this record.
- Reason / purpose for cross-reference:
- read-across source
- Statistics:
- No data
- Positive control results:
- Mean DPM
- Vehicle (acetone/olive oil 4:1 v/v) = 2263
- Test material (5 % w/v) = 5496
- Test material (10 % w/v) = 7571
- Test material (25 % w/v) = 24752
Stimulation Index
- Test material (5 % w/v) = 2.4
- Test material (10 % w/v) = 3.3
- Test material (25 % w/v) = 10.9 - Parameter:
- other: disintegrations per minute (DPM)
- Remarks on result:
- other: - Vehicle = 2511 - Test material (10% v/v) = 3319 - Test material (25% v/v) = 8554 - Test material (50% v/v) = 9916 - Test material (75% v/v) = 22063 - Test material (100% v/v) = 16259
- Key result
- Parameter:
- SI
- Value:
- 1.3
- Test group / Remarks:
- 10% v/v
- Key result
- Parameter:
- SI
- Value:
- 3.4
- Test group / Remarks:
- 25% v/v
- Key result
- Parameter:
- SI
- Value:
- 4
- Test group / Remarks:
- 50% v/v
- Interpretation of results:
- other: Sensitising Category 1
- Remarks:
- Based on CLP criteria
- Conclusions:
- Under the test conditions, d-limonene is classified as ‘R43 May cause sensitisation by skin contact’, according to the criteria of Annex VI to the Directive 67/548/EEC and ‘Category 1’ according to the CLP Regulation (EC) N° (1272-2008).
- Executive summary:
In a local lymph node assay performed in CBA/Ca strain mice according to OECD guideline 429 and in compliance with GLP, groups of mice (4 females/dose) were applied with 25 µL of d-limonene at concentrations of 0 (vehicle control), 10, 25, 50, 75 or 100% v/v in ethanol/diethyl phthalate (3: 1 v/v) to the dorsal surface of each ear for three consecutive days. On Day 6, all animals were injected with 3H-methyl thymidine and after five hours the draining (auricular) lymph nodes were excised and measured for radioactivity expressed as number of disintegrations per minute (DPM). Historic data of hexylcinnamaldehyde (5, 10 and 25 % w/v) in acetone/olive oil (4:1 v/v) was used as the data for positive control group.
Mean DPM for 0, 10, 25, 50, 75 or 100% d-limonene were observed to be 2511, 3319, 8554, 9916, 22063 or 16259 dpm, respectively. Stimulation index for 10, 25, 50, 75 or 100% d-limonene were calculated to be 1.3, 3.4, 4.0, 8.8 or 6.5, respectively. The estimated concentration giving rise to a 3 fold increase in lymphocyte proliferation (EC3) was 22% v/v (5500 µg/cm2). No increase in visual levels of irritancy to the ear skin was observed during the study.
Under the test conditions, d-limonene is classified as ‘R43 May cause sensitisation by skin contact’, according to the criteria of Annex VI to the Directive 67/548/EEC and ‘Category 1’ according to the CLP Regulation (EC) N° (1272-2008).
- Endpoint:
- skin sensitisation: in vitro
- Data waiving:
- study scientifically not necessary / other information available
- Justification for data waiving:
- an in vitro skin sensitisation study does not need to be conducted because adequate data from an in vivo skin sensitisation study are available
Referenceopen allclose all
No control animals exhibited grades greater than 0.5 (slight, patchy erythema). A test animal must therefore at least exhibit a grade 1 at 48 hours to be considered positive (According to the selection criteria defined in the 'interpretation of the results').
- Dermal reactions: Application of the test substance did not result in an increase in visual levels of irritancy to the skin on or around the ear area for the duration of the study.
Table 1 - Skin sensitisation potential of d-limonene in ethanol/diethyl phthalate (3:1 v/v)
Concentration of test substance (% v/v) |
Number of lymph nodes assayed |
Disintegrations per min (dpm) |
dpm per lymph node |
Test:control ratio |
0 (vehicle only) |
8 |
2511 |
314 |
Not applicable |
10 |
8 |
3319 |
415 |
1.3 |
25 |
8 |
8554 |
1069 |
3.4 |
50 |
8 |
9916 |
1240 |
4.0 |
75 |
8 |
22063 |
2758 |
8.8 |
100 |
8 |
16259 |
2032 |
6.5 |
EC3 |
Estimated to be 22% (5500 µg/cm2) |
Table 2 - Skin sensitisation potential of the positive control substance (hexylcinnamaldehyde)
Concentration of hexylcinnamaldehyde (% w/v) |
Number of lymph nodes assayed |
Disintegrations per min (dpm) |
dpm per lymph node |
Test:control ratio |
0 (vehicle only) |
8 |
2263 |
283 |
Not applicable |
5 |
8 |
5496 |
687 |
2.4 |
10 |
8 |
7571 |
946 |
3.3 |
25 |
8 |
24752 |
3094 |
10.9 |
- Dermal reactions: Application of the test substance did not result in an increase in visual levels of irritancy to the skin on or around the ear area for the duration of the study.
Table 1 - Skin sensitisation potential of d-limonene in ethanol/diethyl phthalate (3:1 v/v)
Concentration of test substance (% v/v) |
Number of lymph nodes assayed |
Disintegrations per min (dpm) |
dpm per lymph node |
Test:control ratio |
0 (vehicle only) |
8 |
2511 |
314 |
Not applicable |
10 |
8 |
3319 |
415 |
1.3 |
25 |
8 |
8554 |
1069 |
3.4 |
50 |
8 |
9916 |
1240 |
4.0 |
75 |
8 |
22063 |
2758 |
8.8 |
100 |
8 |
16259 |
2032 |
6.5 |
EC3 |
Estimated to be 22% (5500 µg/cm2) |
Table 2 - Skin sensitisation potential of the positive control substance (hexylcinnamaldehyde)
Concentration of hexylcinnamaldehyde (% w/v) |
Number of lymph nodes assayed |
Disintegrations per min (dpm) |
dpm per lymph node |
Test:control ratio |
0 (vehicle only) |
8 |
2263 |
283 |
Not applicable |
5 |
8 |
5496 |
687 |
2.4 |
10 |
8 |
7571 |
946 |
3.3 |
25 |
8 |
24752 |
3094 |
10.9 |
Endpoint conclusion
- Endpoint conclusion:
- adverse effect observed (sensitising)
- Additional information:
The key study is a local lymph node assay performed with major constituent limonene which was read across to grapefruit oil. The study was performed in CBA/Ca strain mice according to OECD guideline 429 and in compliance with GLP.
Mean DPM for 0, 10, 25, 50, 75 or 100% d-limonene were observed to be 2511, 3319, 8554, 9916, 22063 or 16259 dpm, respectively. Stimulation index for 10, 25, 50, 75 or 100% d-limonene were calculated to be 1.3, 3.4, 4.0, 8.8 or 6.5, respectively. The estimated concentration giving rise to a 3 fold increase in lymphocyte proliferation (EC3) was 22% v/v (5500 µg/cm2). No increase in visual levels of irritancy to the ear skin was observed during the study.
Under the test conditions, d-limonene is classified as ‘R43 May cause sensitisation by skin contact’, according to the criteria of Annex VI to the Directive 67/548/EEC and ‘Category 1’ according to the CLP Regulation (EC) N° (1272-2008).
Three supporting studies on the skin sensitising potential of grapefruit oil were available for the dossier, a human repeated insult patch test, a human maximisation study and a delayed contact hypersensitivity study in guinea pigs.
In the repeated insult patch test (included in 7.10.4), the sensitising properties of grapefruit oil were investigated in humans. 106 subjects completed the study, none of them showed sensitisation.
In the maximisation test (included in 7.10.4), exposure to grapefruit oil also did not induce sensitisation in 24 healthy adult males.
In the delayed contact hypersensitivity study (Buehler Technique), the potential of grapefruit oil to induce skin sensitisation was investigated. After the challenge with several concentrations of test substance, none of the guinea pigs showed sensitisation compared to the controls.
Based on the results of the supporting studies, it can be concluded that the substance graperfruit oil is not a sensitiser. However, as the major constituent of grapefruit oil is limonene, which is classified for skin sensitisation, grapefruit oil will be classified for skin sensitisation.
Migrated from Short description of key information:
Skin sensitisation::
Read across based on d-limonene content: sensitising
Justification for classification or non-classification
Based on the skin sensitising properties of the major constituent of grapefruit oil, d-limonene, grapefruit oil has to be labelled as skin sensitiser in accordance with the EU criteria outlined in Annex VI of 67/548/EEC and Annex I of 1272/2008/EC.
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.
Reproduction or further distribution of this information may be subject to copyright protection. Use of the information without obtaining the permission from the owner(s) of the respective information might violate the rights of the owner.
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