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Diss Factsheets
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EC number: 203-396-5 | CAS number: 106-42-3
- 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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Skin sensitisation
Administrative data
- Endpoint:
- skin sensitisation: in vivo (LLNA)
- Type of information:
- experimental study
- Adequacy of study:
- supporting study
- Study period:
- Not reported
- Reliability:
- 3 (not reliable)
- Rationale for reliability incl. deficiencies:
- other: see 'Remark'
- Remarks:
- GLP status not known, published in peer reviewed literature. Method considered not to be valid following review by the European Centre for the Validation of Alternative Methods (Basketter et al, 2008). Overall, the result obtained by Ehling et al. for mixed xylenes is considered unreliable.
Data source
Referenceopen allclose all
- Reference Type:
- publication
- Title:
- An European inter-laboratory validation of alternative endpoints of the murine local lymph node assay 2nd round
- Author:
- Ehling G et al
- Year:
- 2 005
- Bibliographic source:
- Toxicology 212 69-79
- Reference Type:
- publication
- Title:
- An evaluation of performance standards and nonradioactive endpoints for the Local Lymph Node Assay - the Report and Recommendations of ECVAM Workshop 65
- Author:
- Basketter, D, Cockshott, A, Corsini, E, Gerberick, GF, Idehara, K, Kimber, I, van Loveren, H, Matheson, J, Mehling, A, Omori, T, Rovida, C, Sozu, T, Takeyoshi, M and Casati, S.
- Year:
- 2 008
- Bibliographic source:
- ATLA 36, 243-257
- Reference Type:
- review article or handbook
- Title:
- Unnamed
- Year:
- 2 010
Materials and methods
Test guideline
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
- Deviations:
- yes
- Remarks:
- Considered a non-standard, unvalidated methodology by ECVAM Workshop (e.g. based on mouse stain used, choice of vehicle, time of cell collection, magnitude of discriminating SI; determination of [3H]TdR incorporation by lymph node cells in vitro)
- Principles of method if other than guideline:
- Ear-draining lymph node (LN) weight and cell counts were used to assess LN cell proliferation instead of [3H]TdR incorporation. In addition, the acute inflammatory skin reaction was measured by ear weight determination of circular biopsies of the ears to identify skin irritation properties of the test item.
- GLP compliance:
- not specified
- Type of study:
- mouse local lymph node assay (LLNA)
Test material
- Reference substance name:
- Xylene
- EC Number:
- 215-535-7
- EC Name:
- Xylene
- Cas Number:
- 1330-20-7
- Molecular formula:
- C8H10
- IUPAC Name:
- xylene
- Details on test material:
- - Name of test material (as cited in study report): Xylene
- Analytical purity: 97.5%
Constituent 1
In vivo test system
Test animals
- Species:
- mouse
- Strain:
- Balb/c
- Sex:
- not specified
- Details on test animals and environmental conditions:
- BALB/c mice from local breeder were used. The age of the animals was between 7 and 12 weeks. The animals were kept under standard conditions and had free access to food and water.
Study design: in vivo (LLNA)
- Vehicle:
- other: DAE433 (DAE433 = 40% dimetyhlacetamide, 30% acetone, 30% ethanol by volume
- Concentration:
- 10, 30 and 100%
- No. of animals per dose:
- 6
- Details on study design:
- Dorsal side of both ears treated with 25 µL/ear of the xylene solutions on days 1 to 3. Lymph node weight, lymph node cell counts and topical irritation were determined on day 4, 24 h after the last treatment. Lymph node (LN) weights obtained from LN pairs of individual animals. For the determination of individual LN cell counts, single-cell suspensions from LN pairs of individual animals prepared by mechanical tissue disaggregation. The cells were counted using automated equipment. To assess irritant potential, an ear punch (8 mm diameter) was taken 24 h after last exposure and weighed.
[3H]TdR incorporation by lymph node cells was determined in vitro in other animals sacrificed 4 days following the last topical application (day 6). The auricular lymph nodes (LN) were excised, weighed and pooled for each animal, and suspended in 5 mL RPMI-1640 supplemented with 5% heat inactivated foetal calf serum, 100 U/mL penicillin and 100 mg/mL streptomycin. Single cell suspensions were prepared under aseptic condition. Cells washed twice in standard medium then resuspended in 1 mL standard medium with 10% FCS. Cells counted (Coulter Counter) and cultured at a concentration of 1 × 10^7 cells/mL. When necessary, cell suspensions from several animals were pooled to obtain the concentration required. Cell suspensions seeded in triplicate into micro titre plates. Cells cultured with 10 mL [3H]TdR (3.7 MBq/mL) for 24 h at 37°C in a humidified atmosphere of 5% CO2 in air. [3H]TdR incorporation determined by liquid scintillation counting. [3H]TdR incorporation is expressed per animal.
The methodology used in this paper was reviewed as part of an ECVAM Workshop on “An Evaluation of Performance Standards and Non-radioactive Endpoints for the Local Lymph Node assay”, and the discussion and recommendations reported by Basketter et al. (2008). The Workshop noted that the method used by Ehling et al. (2005) for measurement of cell numbers and ear thickness deviated from the standard method in terms of the strain of mouse used (BALB/c (inbred) or NMRI (outbred) versus CBA/Ca or CBA/J (inbred) in the Guideline); excision of lymph nodes on day 4 (rather than on day 6, as stated in Guideline); use of dimethylacetone/acetone/ethanol (4:3:3) as vehicle (versus vehicles / vehicle validation requirements of the Guideline); and a cut-off at SI = 1.4 or above (versus a cut-off of SI=3 or above in the Guideline). Together, these were concluded (by the ECVAM Workshop) to represent a MAJOR change in methodology that required further validation.
The use of [3H]TdR incorporation by lymph node cells in vitro (as opposed to incorporation of [3H]TdR following i.v. injection in vivo was not discussed but appears to represent a change versus the Guideline.
Basketter and Kimber (2010) also commented that the findings of Ehling et al. (2005) for mixed xylenes can only be considered of limited relevance since the assay used was not considered to be valid following review by ECVAM. - Positive control substance(s):
- hexyl cinnamic aldehyde (CAS No 101-86-0)
- Statistics:
- Measured parameters compared using non-parametric Kruskal-Wallis test as the global test, and the non-parametric two-group Wilcoxon-Mann-Whitney rank test for all two-group comparisons. Because of power considerations the non-parametric Steel test was not applied for the comparison of the three concentration groups with the control group.
Results and discussion
- Positive control results:
- A satisfactory response was obtained with the positive control substance (alpha-hexylcinnamaldehyde, assessed blind at all 11 laboratories) however no quantitative results were included in the publication.
In vivo (LLNA)
Resultsopen allclose all
- Parameter:
- SI
- Remarks on result:
- other: Not reported
- Parameter:
- other: disintegrations per minute (DPM)
- Remarks on result:
- other: Not reported
Any other information on results incl. tables
Treatment with mixed xylene was associated with a statistically significant increase in ear-draining lymph node weight and cell count (indicative of a sensitisation response according to criteria applied by the study authors) in 7 of the 9 laboratories involved in the trial; and an increase in ear weight (indicative of irritation according to criteria applied by the study authors) in 3 of 9 laboratories.
Although not discussed in any detail, one laboratory also determined 3[H]TdR incorporation by lymph node cells in vitro and obtained a "positive" result.
Overall, mixed xylene was classified as an allergen in 8 of the 9 laboratories participating in the trial.
Applicant's summary and conclusion
- Interpretation of results:
- ambiguous
- Conclusions:
- Mixed xylene produced a statistically significant increase in ear-draining lymph node weight and cell count (indicative of a sensitisation response according to criteria used by the study authors) in 7 of the 9 laboratories involved in the trial; and an increase in ear weight (indicative of irritation) in 3 of 9 laboratories. The study authors concluded that mixed xylene was an allergen, however the findings are of limited relevance because the assay was not considered valid following review by the European Centre for the Validation of Alternative methods (Basketter et al., 2008; Basketter and Kimber, 2010).
- Executive summary:
The sensitisation potential of mixed xylenes was assessed in a modified local lymph node assay where increased ear-draining lymph node weight and cell counts were used to quantify lymph node cell proliferation by 9 laboratories participating in a ring-trial. [3H]TdR incorporation in vitro by lymph node cells was also assessed in a single laboratory (assay not conducted by other laboratories). Acute skin inflammation (a potential confounder that can lead to "false positive" results) was also determined based on changes in ear tissue weight. Treatment with mixed xylenes was associated with a statistically significant increase in ear-draining lymph node weight and cell count in 7 of the 9 laboratories, and an increase in ear tissue weight in 3 of 9 laboratories. An increased (but not quantified) increase in 3[H]TdR incorporation by lymph node cells was also reported by one laboratory. Based on these findings, mixed xylene was considered an allergen by a majority of laboratories participating in the trial. However the methodology used was evaluated by an ECVAM Workshop (reported by Basketter et al., 2008) which concluded the assay methods deviated from the Guideline in terms of the strain of mouse, the choice of vehicle, the time of lymph node/lymph node cell collection, and the magnitude of the SI used to differentiate positive and negative samples. Overall, the ECVAM Workshop concluded that these differences represented a major change in methodology that required further validation. The use of [3H]TdR incorporation by lymph node cells in vitro (as opposed to incorporation of [3H]TdR following i.v. injection in vivo, as required by the Guideline) was not discussed by the ECVAM Workshop but is another change relative to the Guideline. Basketter and Kimber (2010) also noted that these findings for mixed xylenes can only be considered of limited relevance given methodological deviations from the Guideline. It is concluded that this publication does not provide convincing evidence that mixed xylenes possesses a potential to induce or elicit skin sensitisation.
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