Registration Dossier

Administrative data

Endpoint:
acute toxicity: dermal
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Justification for type of information:
JUSTIFICATION FOR DATA WAIVING
According to the later amendment of REACH, i.e. Commission Regulation (EU) 2016/863 of 31 May 2016, testing by the dermal route does not need to be conducted if:
— the substance does not meet the criteria for classification as acute toxicity or STOT SE by the oral route and
— no systemic effects have been observed in in vivo studies with dermal exposure (e.g. skin irritation, skin sensitisation) or, in the absence of an in vivo study by the oral route, no systemic effects after dermal exposure are predicted on the basis of non-testing approaches (e.g. read across, QSAR studies).
The oral LD50 in rats is >2000 mg/kg, no animal died at 2000 mg/kg, no symptoms of poisoning were observed in males and females. No clinical signs were noted at both skin irritation and sensitization testing. In the latter, no symptoms of local skin irritation at the ears of the animals and no signs of systemic toxicity were observed during the study period.
Further, due to the high LogPow = 6.3, absorption through the skin may be diminished, as outlined in the subchapter “Toxicokinetics”, and the possibly notable effects are in general considered to be of a lesser extent compared to the ones revealed in oral application testing. So any testing via the dermal route is not considered to provide additional, valuable information.
Hence, testing can be waived.
Cross-referenceopen allclose all
Reason / purpose:
data waiving: supporting information
Reference
Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1990-11-08 - 1991-02-08
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted under GLP according to EU method B.1 and OECD guideline 401 on the registered substance itself. The method is to be considered scientifically reasonable with negligible deficiencies in documentation.
Reference:
Composition 0
Qualifier:
according to
Guideline:
EU Method B.1 (Acute Toxicity (Oral))
Version / remarks:
as set out in 84/449/EWG (1984-09-19)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 401 (Acute Oral Toxicity)
Version / remarks:
adopted 1987-02-24
Deviations:
no
GLP compliance:
yes
Test type:
standard acute method
Limit test:
yes
Test material information:
Composition 1
Species:
rat
Strain:
other: Wistar Bor: WISW (SPF Cpb)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Breeder Winkelmann, Borchen
- Age at study initiation: 8 weeks (males), 10 weeks (females)
- Weight at study initiation: average 185 g (males), 166 g (females), weight variation not exceeding ±20%
- Fasting period before study: yes, 16h prior to application
- Housing: In groups of five in Makrolon cages type III on low-dusting wood pellets
- Diet (e.g. ad libitum): "fixed-formula" standard diet Altromin® 1324 Pellets (Altromin GmbH und Co KG, Lage) ad libitum
- Water (e.g. ad libitum): tap water ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2 °C
- Humidity (%): 50 ± 10%
- Air changes (per hr): ca. 10/h
- Photoperiod (hrs dark / hrs light): 12 / 12
Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on oral exposure:
VEHICLE
- Concentration in vehicle: 200 mg/ml
- Amount of vehicle (if gavage): 10 ml/kg bw
Doses:
2000 mg/kg
No. of animals per sex per dose:
5 / sex / dose
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: several times on application day, otherwise twice a day for clinical symptoms, weighing on day 0, 7 and 14
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight
Key result
Sex:
male/female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: No animal died during study duration.
Key result
Sex:
male/female
Dose descriptor:
LD0
Effect level:
>= 2 000 mg/kg bw
Based on:
test mat.
Remarks on result:
other: No animal died during study duration.
Mortality:
No animal died during the 14-day observation period.
Clinical signs:
After single gavage of 2000 mg/kg bw no symptoms of poisoning were observed in males and females.
Body weight:
There were no effects on growth of males and females.
Gross pathology:
All animals killed after 14 days were without pathological findings.
Interpretation of results:
other: EU-GHS criteria not met
Conclusions:
The study was conducted under GLP according to EU method B.1 and OECD guideline 401 on the registered substance itself. The method is to be considered scientifically reasonable with negligible deficiencies in documentation. Hence, the results can be considered as sufficiently reliable to assess the acute oral toxicity in rats. The determined LD50 value is >2000 mg/kg bw, the LD0 ≥ 2000 mg/kg, as none of the animals died after gavage of 2000 mg/kg bw. The result is suitable to determine the classification of Phenol, 2,2'-methylenebis-(6-cyclohexyl-4-methyl). According to Regulation (EC) No. 1272/2008, the substance does not need to be classified as acute toxic cat. IV or higher.
Executive summary:

In an acute oral toxicity study under GLP according to EU method B.1 and OECD guideline 401, groups of fasted 8-10 weeks old Wistar Bor: WISW (SPF Cpb) rats (5/sex) were given a single oral dose of Phenol, 2,2'-methylenebis-(6-cyclohexyl-4-methyl) in arachis oil at a limit dose 2000 mg/kg bw and observed for 14 days.

 

Oral LD50> 2000 mg/kg bw

Oral LD0≥ 2000 mg/kg bw

None of the animals died during the test or showed any signs of toxicity

 

Phenol, 2,2'-methylenebis-(6-cyclohexyl-4-methyl) is of low toxicity based on the LD50 and does not need to be classified as acute toxic cat. IV or higher.

Reason / purpose:
data waiving: supporting information
Reference
Endpoint:
skin irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Conducted prior to GLP implementation similar to OECD 404 on the registered substance itself with some deficiencies in documentation and performance, i.e. only two animals were used instead of three, substance was applied on the ear, not the trunk, evaluation criteria not defined.
Reference:
Composition 0
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
Deviations:
not applicable
GLP compliance:
no
Remarks:
test conducted prior to GLP implementation
Test material information:
Composition 1
Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
TEST ANIMALS
- Housing: individually
Type of coverage:
semiocclusive
Preparation of test site:
not specified
Vehicle:
unchanged (no vehicle)
Controls:
no
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 500 mg
Duration of treatment / exposure:
24 h
Observation period:
7 days
Number of animals:
2
Details on study design:
TEST SITE
- Area of exposure: inner face of the ear
- Type of wrap if used: band aid

REMOVAL OF TEST SUBSTANCE
- Washing (if done): with water and soap/vegetable oil
- Time after start of exposure: 24 h
Irritation parameter:
other: irritating properties
Basis:
mean
Remarks on result:
other: not irritating
Interpretation of results:
not irritating
Conclusions:
The study was conducted prior to GLP implementation similar to OECD 404 on the registered substance itself with some deficiencies in documentation and performance. Nevertheless, with the given properties of the test item, the methodological deficiencies are unlikely to have any influence on the results. The given information on test performance allows the conclusion that the method was scientifically reasonable performed. Also, the result (not irritating) is confirmed by the other available studies on skin and in vivo eye irritation, clearing any doubts regarding the reliability of the result. Hence, Phenol, 2,2'-methylenebis-(6-cyclohexyl-4-methyl) does not need to be regarded as dermal irritant.
Executive summary:

In a primary dermal irritation study similar to OECD 404, two New Zealand White rabbits were dermally exposed to 500 mg of unchanged Phenol, 2,2'-methylenebis-(6-cyclohexyl-4-methyl) for 24 hours to the inner side of the ear.  Animals then were observed for 7 days. In this study, Phenol, 2,2'-methylenebis-(6-cyclohexyl-4-methyl) is not a dermal irritant.

 

Reason / purpose:
data waiving: supporting information
Reference
Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2015-11-02 - 2016-02-04
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Well-documented OECD GLP guideline study without deviations on the registered substance itself.
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
OECD Guidelines for Testing of Chemicals, Updated Guideline 429: Skin Sensitisation: Local Lymph Node Assay (adopted 22 July 2010)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Version / remarks:
Commission Regulation (EC) No. 440/2008, B.42: “Skin Sensitisation: Local Lymph Node Assay”, dated 06 July 2012
Deviations:
not specified
GLP compliance:
yes (incl. certificate)
Remarks:
Hess. Ministerium für Umwelt, Klimaschutz, Landwirtschaft und Verbraucherschutz, Mainzer Straße 80, D65189 Wiesbaden
Type of study:
mouse local lymphnode assay (LLNA)
Test material information:
Composition 1
Species:
mouse
Strain:
other: CBA/CaOlaHsd
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Envigo RMS B.V., Inc, Postbus 6174, 5960 AD Horst / The Netherlands
- Age at study initiation: 8 - 9 weeks
- Weight at study initiation: 17.5 - 21.6 g
- Housing: Conventionally, each dose group (5 mice/group) in one cage, Cage Type: Makrolon Type II (pre-test) / III (main study), with wire mesh top, Bedding: granulated soft wood bedding
- Diet (e.g. ad libitum): 2018C Teklad Global 18% protein rodent diet (certified), ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2 °C
- Humidity (%): approx. 45-65%
- Photoperiod (hrs dark / hrs light): 12/12 (artificial light 6.00 a.m. - 6.00 p.m.)
Vehicle:
acetone/olive oil (4:1 v/v)
Remarks:
Acetone: Purity: 99.98%; Olive oil: highly refined, low acidity
Concentration:
0, 10%, 25%, 50%
No. of animals per dose:
5
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: A solubility experiment was performed according to the recommendations given by OECD 429. The highest test item concentration, which can be technically used, was a 50% solution in acetone/olive oil (4+1, v/v). Vortexing was used to formulate the test item.
- Irritation: To determine the highest non-irritant test concentration that at the same time did not induce signs of systemic toxicity, a pre-test was performed in two animals and stated in raw data and report. Two mice were treated by (epidermal) topical application to the dorsal surface of each ear with test item concentrations of 25 and 50% once daily each on three consecutive days. Prior to the first application of the test item and before sacrifice the body weight was determined. Clinical signs were recorded at least once daily. Eventual signs of local irritation were documented and a score was used to grade a possible erythema of the ear skin. Furthermore, prior to the first application of the test item (day 1), on day 3 and before sacrifice (day 6) the ear thickness was determined using a micrometer. Additionally, for both animals, the ears were punched after sacrifice (day 6) at the apical area using a biopsy punch (Ø 8 mm corresponding to 0.5 cm²) and were immediately pooled per animal and weighed using an analytical balance. Eventual ear irritation was considered to be excessive if an erythema of the ear skin of a score value ≥3 was observed at any observation time and/or if an increase in ear thickness of ≥25% was recorded on day 3 or day 6.
At the tested concentrations the animals did not show any signs of systemic toxicity. On day 2 and 3, the animal treated with 50% test item concentration showed a very slight erythema of the ear skin (Score 1). The animal treated with 25% test item concentration did not show any signs of local skin irritation.
Thus, the test item in the main study was assayed at 10, 25, and 50% (w/w). The highest concentration tested was the highest level that could be achieved whilst avoiding systemic toxicity and excessive local skin irritation as confirmed in the pre-experiment.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response:
A test item is regarded as a sensitiser in the LLNA if the following criteria are fulfilled:
• First, that exposure to at least one concentration of the test item resulted in an incorporation of ³HTdR at least 3-fold or greater than that recorded in control mice, as indicated by the Stimulation Index.
• Second, that the data are compatible with a conventional dose response, although allowance must be made (especially at high topical concentrations) for either local toxicity or immunological suppression.

TREATMENT PREPARATION AND ADMINISTRATION:
Test Item Preparation
The test item was placed into an appropriate container on a tared balance and acetone/olive oil (4+1, v/v) was added.
The different test item concentrations were prepared individually.
The preparations were made freshly before each dosing occasion.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
Statistical Analysis
The mean values and standard deviations were calculated in the body weight tables and for the DPM values (group mean DPM ± standard deviation).
The Dean-Dixon-Test and the Grubb’s test were used for identification of possible outliers (performed with validated program R Script Outlier.Rnw). No outlier value was detected.
Positive control results:
S.I. values per Test item concentration % in acetone:olive oil (4+1, v/v):
0%: 1.00
5%: 1.62
10%: 4.23
25%: 17.56
Parameter:
SI
Remarks on result:
other: Vehicle Control Group (acetone/olive oil (4+1, v/v)): 1.00 10% : 1.32 25% : 1.10 50% : 1.20
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: Vehicle Control Group (acetone/olive oil (4+1, v/v)): 1458.5 10% : 1928.7 25% : 1611.1 50% : 1744.1
Interpretation of results:
GHS criteria not met
Conclusions:
The study was conducted under GLP according to OECD guideline 429 on the registered substance itself. The method is to be considered scientifically reasonable with no deficiencies in documentation or any deviations, the validity criteria are fulfilled, positive and negative controls gave the appropriate response. Hence, the results can be considered as reliable to assess the sensitizing potential of the test item in mice. The test item did not induce signs of general toxicity.
A test item is regarded as a sensitiser in the LLNA if the exposure to one or more test concentration resulted in a 3-fold or greater increase in incorporation of 3HTdR compared with concurrent controls, as indicated by the Stimulation Index (S.I.). The estimated concentration of test item required to produce a S.I. of 3 is referred to as the EC3 value.
In this study Stimulation Indices of 1.32, 1.10, and 1.20 were determined with the test item at concentrations of 10, 25, and 50% (w/w) in acetone/olive oil (4+1, v/v).
So, the test item was identified as a non-sensitizing agent in the Local Lymph Node Assay. Hence, no classification as skin sensitizer is triggered.
Executive summary:

In the study according to OECD 429 under GLP the test item formulated in acetone/olive oil (4+1, v/v) was assessed for its possible skin sensitising potential.

For this purpose a local lymph node assay was performed using test item concentrations of 10, 25, and 50% (w/w). The highest concentration tested was the highest concentration that could be achieved whilst avoiding systemic toxicity and excessive local skin irritation as confirmed by a pre-experiment.

The animals did not show any signs of systemic toxicity or local skin irritation during the course of the study and no cases of mortality were observed.

In this study Stimulation Indices (S.I.) of 1.32, 1.10, and 1.20 were determined with the test item at concentrations of 10, 25, and 50% (w/w) in acetone/olive oil (4+1, v/v), respectively.

The test item was not a skin sensitiser under the test conditions of this study.

Reason / purpose:
data waiving: supporting information
Reference
Endpoint:
partition coefficient
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
no GLP
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 117 (Partition Coefficient (n-octanol / water), HPLC Method)
Deviations:
no
GLP compliance:
no
Type of method:
HPLC method
Partition coefficient type:
octanol-water
Test material information:
Composition 1
Analytical method:
high-performance liquid chromatography
Key result
Type:
log Pow
Partition coefficient:
6.3
Remarks on result:
other: Temperature and pH were not specified in the study.
Conclusions:
The log Pow of 2,2'-methylenebis[6-cyclohexyl-p-cresol] was determined in an OECD 117 study with acceptable restrictions. The reverse phase HPLC method enables partition coefficients to be estimated in the log Pow range between 0 and 6, but can be expanded to cover the log Pow range between 6 and 10 in exceptional cases. The log Pow was determined as 6.3, and so falls into applicability of the employed method.
Executive summary:

The log Pow of 2,2'-methylenebis[6-cyclohexyl-p-cresol] was determined using reverse phase HPLC according to OECD guideline 117, with acceptable restrictions, to be 6.3.

Reason / purpose:
data waiving: supporting information
Reference
Endpoint:
basic toxicokinetics, other
Remarks:
Expert statement
Type of information:
other: Expert statement
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: No GLP-conform guideline study, but scientifically valid expert statement based i.a. on studies assessed with Klimisch 1 or 2
Reference:
Composition 0
Objective of study:
absorption
distribution
excretion
metabolism
toxicokinetics
Qualifier:
no guideline required
Principles of method if other than guideline:
An extensive assessment of the toxicokinetic behaviour of 2,2'-methylenebis[6-cyclohexyl-p-cresol] was performed, taking into account the chemical structure, the available physico-chemical and toxicological data.
GLP compliance:
no
Test material information:
Composition 1
Radiolabelling:
other: not applicable
Species:
other: not applicable
Strain:
other: not applicable
Details on test animals and environmental conditions:
not applicable
Route of administration:
other: All relevant routes of administration are discussed in the expert statement.
Vehicle:
other: not applicable
Details on exposure:
not applicable
Duration and frequency of treatment / exposure:
not applicable
Remarks:
not applicable
Control animals:
other: not applicable
Positive control:
not applicable
Details on study design:
not applicable
Details on dosing and sampling:
not applicable
Statistics:
not applicable
Type:
absorption
Results:
The relevant absorption rates were estimated to: Oral absorption: approx. 50% Dermal absorption: approx. 10% Inhalative absorption: approx. 20%
Type:
distribution
Results:
The absolute systemic bioavailablility is rather low and expected to be more extensive in fat tissues than in other tissues.
Type:
metabolism
Results:
Aliphatic hydroxylation was identified as the mode of action during Phase-I-metabolism, and subsequent conjugation is expected.
Type:
excretion
Results:
Excretion of the parent compound will occur via the gastrointestinal tract (unabsorbed material) and the bile, and it could be subject to enterohepatic recycling. For metabolites elimination of the Phase 2 conjugated metabolites via the bile is expected.
Details on absorption:
Absorption
In general, absorption of a chemical is possible, if the substance crosses biological membranes. In case where no transport mechanisms are involved, this process requires a substance to be soluble, both in lipid and in water, and is also dependent on its molecular weight (substances with molecular weights below 500 are favourable for absorption). Generally, the absorption of chemicals which are surfactants or irritants may be enhanced, because of damage to cell membranes. However, since 2,2’-methylenebis[6-cyclohexyl-p-cresol] was found to be non-irritating to the skin, the possibility of an enhanced absorption due to damaged cell membranes can be excluded.
Due to the lack of experimental absorption data, the following physico-chemical parameters of 2,2’-methylenebis[6-cyclohexyl-p-cresol] will be taken into account when discussing its absorption into the body:
- Molecular weight = 392.57 g/mol
- Water solubility = 0.034 mg/L at 20 °C
- Partition Coefficient Log Pow = 6.3 at 23°C
- Vapour pressure = 9.48exp(-12) kPa at 20°C, 1.23exp(-9) kPa at 50°C, and 5.18exp(-9) kPa at 60°C
- Boiling point: Decomposition starts above 200 °C, the peak for decomposition temperature is 362.67 °C
- Particle size: 0% respirable fraction (<4 µm), 0.06% thoracic fraction (4-10 µm), 40.11% inhalable fraction (10-100 µm)

Absorption from the gastrointestinal tract
In the small intestine absorption occurs mainly via passive diffusion or lipophilic compounds may form micelles and be taken into the lymphatic system. Additionally, metabolism can occur by gut microflora or by enzymes in the gastrointestinal mucosa. However, the absorption of highly lipophilic substances (Log Pow of 4 or above) may be limited by the inability of such substances to dissolve into gastrointestinal fluids and hence make contact with the mucosal surface. The absorption of such substances will be enhanced if they undergo micellular solubilisation by bile salts. Substances absorbed as micelles enter the circulation via the lymphatic system, bypassing the liver. Consequently, immediate Cytochrome P450 metabolism is less important here as for substances which directly enter the hepatic system via the portal vein.
According to ECHA’s guidance R.7c [ECHA 2008], it is stated that the smaller the molecule the more easily it may be taken up. Molecular weights below 500 are favourable for absorption. With a molecular weight of 392.57 g/mol, absorption in general can be considered as possible. However, substances with high Log Pow values, i.e. 6.3 in this case, are less favourable for absorption by passive diffusion, which diminishes the ability of 2,2’-methylenebis[6-cyclohexyl-p-cresol] to be absorbed. Only water-soluble substances will readily dissolve into the gastrointestinal fluids and hence be available for absorption. The substance of interest though has a very low water solubility, i.e. 0.034 mg/L at 20°C. This data indicates that the substance is practically insoluble in water, and so it can be assumed that the solved amount of the substance in e.g. the upper GI tract will be very limited, clearly indicative for a poor absorption. Additionally, the substance is non-toxic, i.e. the available acute 50% lethal doses are above the classification limit of 2000 mg/kg bw, i.e. > 2000 mg/kg bw in rats, no deaths or signs of toxicity occurred, so the actually determined values are LD0 values of ≥ 2000 mg/kg resp. ≥ 2500 mg/kg. Absorption to a certain extent occurred, as in the available OECD 422 study a NOAEL of 100 mg/kg bw/d was determined, but this value is in general not too low, which furthermore indicates either a relative non-toxicity of the substance or its poor absorption.
2,2’-methylenebis[6-cyclohexyl-p-cresol] is not biodegradable, so a preliminary metabolism by gut bacteria is less likely, so additional metabolites prior to absorption do also not have to be regarded.
Based on the available data, an absorption rate of 50% or less can be estimated, predominantly due to the insolubility of the substance in water and the high logPow.

Absorption from the respiratory tract
Concerning absorption in the respiratory tract, any gas, vapour or other substances inhaled as respirable dust (i.e. particle size ≤ 15µm) has to be sufficiently lipophilic to cross the alveolar and capillary membranes (moderate Log Pow values between 0-4 are favourable for absorption). The rate of systemic uptake of very hydrophilic gases or vapours may be limited by the rate at which they partition out of the aqueous fluids (mucus) lining the respiratory tract and into the blood. Such substances may be transported out of the lungs with the mucus and swallowed or pass across the respiratory epithelium via aqueous membrane pores. Lipophilic substances (Log Pow >0) have the potential to be absorbed directly across the respiratory tract epithelium. Any lipophilic compound may be taken up by micellular solubilisation but this mechanism may be of particular importance for highly lipophilic compounds (Log Pow >4), particularly those that are poorly soluble in water (1 mg/L or less) that would otherwise be poorly absorbed [ECHA, 2008].
2,2’-methylenebis[6-cyclohexyl-p-cresol] has a very low vapour pressure (9.48exp(-12) kPa at 20°C, 1.23exp(-9) kPa at 50°C, and 5.18exp(-9) kPa at 60°C) and decomposes without boiling above 200°C, clearly showing that the inhalative absorption as a gas does not have to be regarded. According to its particle size distribution, 0% of the particles belong to the respirable fraction (<4 µm), only 0.06% to the thoracic fraction (4-10 µm), and 40.11% of the particles are inhalable (10-100 µm). Those boundary values are taken from the recent version of ECHA’s Guidance document [ECHA, 2015], the former version gave the following information: In humans, particles with aerodynamic diameters below 100 μm have the potential to be inhaled. Particles with aerodynamic diameters below 50 μm may reach the thoracic region and those below 15 μm the alveolar region of the respiratory tract [ECHA, 2008]. Based on the particle size distribution, only 40% of the applied dose would reach the body via inhalation, all other particles are too large to be bioavailable. Further, most of the particles are subject to nasal clearance, a minor portion to tracheobronchial clearance, and none of the particles are able to reach the alveoli. As a consequence, there is a very small fraction potentially reaching the alveolar region of the respiratory tract beyond the bronchi, where no mechanical excretion mechanism as the ciliary movements is available. So, the potential absorption can only be regarded theoretically.
For absorption of deposited material similar criteria as for GI absorption can be applied. In general, either a prolonged exposure due to deposition and subsequent absorption or immediate absorption by micellular solubilisation has to be assumed. The latter mechanism may be of particular importance for highly lipophilic compounds (LogPow >4), particularly those that are poorly soluble in water (1 mg/l or less) and is hence the only relevant here. To be readily soluble in blood, a gas, vapour or dust must be soluble in water and increasing water solubility would increase the amount absorbed per breath. However, the gas, vapour or dust must also be sufficiently lipophilic to cross the alveolar and capillary membranes. Therefore, a moderate log P value (between -1 and 4) would be favourable for absorption. Generally, liquids, solids in solution and water-soluble dusts would readily diffuse/dissolve into the mucus lining the respiratory tract. Hence, with a logPow of 6.3 and a virtually non-existing water solubility of 0.34 mg/L, the absorption of this fraction which may not be subjected to ciliary clearance can be considered as very limited by the rate at which the substance partitions out of the fluid in the lung surface.
Hence, as only a very minor portion of the particles may reach the part of the lungs where no clearance mechanisms such as ciliary movements exist and a prolonged high exposure can be excluded, and further, micellular solubilisation is the only reasonable uptake mechanism, the inhalative absorption of the substance can be considered as minor. In theory, if absorption was not limited by the particle size, a similar absorption rate as via the oral route, i.e. 50%, could be assumed. Due to the physical limitations however, an inhalative absorption of approx. 20% can be assumed as a worst case.
Due to ciliary movements in the upper respiratory tract, the secretion of possibly unsolved, deposed particles with a subsequent swallowing and hence oral exposure after inhalation has to be additionally regarded.

Absorption after dermal exposure
In order to cross the skin, a compound must first penetrate into the stratum corneum and may subsequently reach the epidermis, the dermis and the vascular network. The stratum corneum provides its greatest barrier function against hydrophilic compounds, whereas the epidermis is most resistant to penetration by highly lipophilic compounds. Substances with a molecular weight below 100 are favourable for penetration through the skin and substances above 500 are normally not able to penetrate. The substance must be sufficiently soluble in water to partition from the stratum corneum into the epidermis. Therefore, if the water solubility is below 1 mg/L, dermal uptake is likely to be low. Additionally, Log Pow values between 1 and 4 favour dermal absorption.
Above 4, the rate of penetration may be limited by the rate of transfer between the stratum corneum and the epidermis, but uptake into the stratum corneum will be high. Above 6, the rate of transfer between the stratum corneum and the epidermis will be slow and will limit absorption across the skin. Uptake into the stratum corneum itself may be slow. Moreover vapours of substances with vapour pressures below 100 Pa are likely to be well absorbed and the amount absorbed dermally is most likely more than 10% and less than 100 % of the amount that would be absorbed by inhalation. If the substance is a skin irritant or corrosive, damage to the skin surface may enhance penetration. During the whole absorption process into the skin, the compound can be subject to biotransformation.
In case of 2,2’-methylenebis[6-cyclohexyl-p-cresol], an evaporation after skin contact does not need to be regarded due to the high decomposition temperature and low vapour pressure, and hence it can be assumed that the substance will remain on the skin until mechanical removal. Furthermore, since the substance is not a skin irritant, additional absorption-enhancing effects can be disregarded, too.
With a molecular weight of 392.57 g/mol in theory a high absorption via the skin and hence a default dermal absorption rate of 100% [ECHA, 2008] could be assumed.
However, a log Pow of 6.3 at 23 °C and the fact that the substance is with 0.034 mg/l virtually insoluble in water indicate the opposite and are considered to be more important and so, dermal uptake is likely to be low. In addition, dry particulates will have to dissolve into the surface moisture of the skin before uptake can begin [ECHA, 2008]. Therefore, a limited dermal absorption rate can be assumed, most likely about 10%.
Details on distribution in tissues:
Distribution
In general, it can be stated that the smaller the molecule, the wider is its distribution. A lipophilic molecule (Log Pow >0) is likely to distribute into cells and the intracellular concentration may be higher than extracellular concentration particularly in fatty tissues. It is not possible to foresee protein binding, which can limit the amount of a substance available for distribution. ToxTree modelling revealed protein binding alerts, both a Michael acceptor and for nucleophilic aliphatic substitution, but the extent however is unknown. Furthermore, if a substance undergoes extensive first-pass metabolism, predictions made on the basis of the physico-chemical characteristics of the parent substance may not be applicable.
In case of 2,2’-methylenebis[6-cyclohexyl-p-cresol], no quantitative data is available for distribution patterns. Taking into account its medium molecular weight of 392.57 g/mol, its lipophilicity and poor water solubility, the absolute systemic bioavailability is rather low and expected to be more extensive in fat tissues than in other tissues, especially when absorption occurred while bypassing the liver.
After oral exposure, the first target will be the gastrointestinal tract, where the substance and possibly bacterial metabolites will be absorbed in small quantities and transferred via the blood stream to the liver. After first pass metabolism, the substance will be further distributed via the bloodstream. Here, especially the kidneys due to their filter function and the heart due to its enormous need for nutrients and consequently large blood flow through coronary arteries will be affected. In general, the estimated metabolites are slightly more hydrophilic due to the hydroxyl groups, which is however expected to be of minor importance given the very high lowPow of the parent compound, influences are considered to be only minor. Since the solubility and hence absorption via the GI tract of the parent compound is rather limited, an excessive formation of metabolites is unlikely and so is a peak exposure to the metabolites. After absorption of the parent compound via other routes, only a subsequent metabolism has to be taken into account, also leading to no relevant peak exposure.
Additionally, accumulation of the parent compound in all directly and indirectly (general supply via bloodstream) involved organs has to be considered. Furthermore, the affection of the lymphatic system via micellar uptake must also be taken into consideration.
Details on excretion:
Excretion
In general, the major routes of excretion for substances from the systemic circulation are the urine and/or the faeces (via bile and directly from the gastrointestinal mucosa). For non-polar volatile substances and metabolites exhaled air is an important route of excretion. Substances that are excreted favourable in the urine tend to be water-soluble and of low molecular weight (below 300 in the rat) and be ionized at the pH of urine. Most will have been filtered out of the blood by the kidneys though a small amount may enter the urine directly by passive diffusion and there is the potential for reabsorption into the systemic circulation across the tubular epithelium. Substances that are excreted in the bile tend to be amphipathic (containing both polar and nonpolar regions), hydrophobic/strongly polar and have higher molecular weights and pass through the intestines before they are excreted in the faeces and as a result may undergo enterohepatic recycling which will prolong their biological half-life. This is particularly a problem for conjugated molecules that are hydrolysed by gastrointestinal bacteria to form smaller more lipid soluble molecules that can then be reabsorbed from the GI tract. Those substances less likely to recirculate are substances having strong polarity and high molecular weight of their own accord. Other substances excreted in the faeces are those that have diffused out of the systemic circulation into the GIT directly, substances which have been removed from the gastrointestinal mucosa by efflux mechanisms and non-absorbed substances that have been ingested or inhaled and subsequently swallowed. Non-ionized and lipid soluble molecules may be excreted in the saliva (where they may be swallowed again) or in the sweat. Highly lipophilic substances that have penetrated the stratum corneum but not penetrated the viable epidermis may be sloughed off with or without metabolism with skin cells.
For 2,2’-methylenebis[6-cyclohexyl-p-cresol] no data is available regarding its elimination. Concerning the above mentioned behaviour predicted for its metabolic fate, it is unlikely that the parent substance will be excreted unchanged. However, if unchanged excretion is assumed, based on the chemical structure of 2,2’-methylenebis[6-cyclohexyl-p-cresol], its molecular weight and its non-existent water solubility, it is unlikely to be excreted via the urine. The excretion, if any, of the parent compound will occur via the gastrointestinal tract (unabsorbed material) and the bile (small amounts of unchanged compound), and it could be subject to enterohepatic recycling.
Regarding the metabolites of 2,2’-methylenebis[6-cyclohexyl-p-cresol], a elimination of the Phase 2 conjugated metabolites, via the bile is expected.
Metabolites identified:
yes
Details on metabolites:
For details, see attached file. Metabolites of the substance are estimated to be formed via Aliphatic hydroxylation.

See attached expert statement.

Conclusions:
The present expert statement covers all relevant toxicokinetic parameters to assess the behaviour of 2,2’-methylenebis[6-cyclohexyl-p-cresol] in the body, the available information is sufficient to enable one to perform a proper risk assessment. Hence, no further information needs to be gathered and further studies can be omitted due to animal welfare. In conclusion, the substance has a low potential for bioaccumulation in its non-metabolized or metabolized form.
Executive summary:

In order to assess the toxicokinetic behaviour of 2,2’-methylenebis[6-cyclohexyl-p-cresol], the available toxicological and physico-chemical data were evaluated.

The substance is expected to be moderately absorbed via the oral route. An oral absorption rate of 50% or less was estimated due to its very low water solubility and high logPow. Due to its particle size distribution, the inhalation of particles practically does not have to be regarded, however in theory it has a potential for absorption via respiratory tract epithelium by micellular solubilisation. As a consequence, due to physical limitations, a inhalative absorption rate of 20% can be estimated. Taking into account its poor water solubility, a diminished dermal absorption rate can be assumed, approx. 10%.

So in summary, the absorption rates may be estimated to:

-      Absorption via oral route: 50%

-      Absorption via inhalative route: 20%

-      Absorption via dermal route: 10%

The absolute systemic bioavailablility is rather low and expected to be more extensive in fat tissues than in other tissues. The estimated Cytochrome P450 metabolites of the substance are formed via aliphatic hydroxylation, and need to be regarded, too, as either a first-pass metabolism or a metabolism by other tissues is expected to occur. The introduction of single hydroxyl groups into the otherwise unchanged molecule, is not expected to alter its ADME behaviour substantially given the magnitude of its logPow.

Due to the high logPow and the very low water solubility of 2,2’-methylenebis[6-cyclohexyl-p-cresol], a certain potential for accumulation in the body is indicated. Nevertheless, a prolonged, extensive accumulation is not expected.

It is unlikely that the parent substance will be excreted unchanged. However, if unchanged excretion is assumed, it is unlikely to be excreted via the urine. The excretion, if any, of the parent compound will occur via the gastrointestinal tract (unabsorbed material) and the bile (small amounts of unchanged compound), and it could be subject to enterohepatic recycling. Regarding the metabolites of 2,2’-methylenebis[6-cyclohexyl-p-cresol], a elimination of the Phase 2 conjugated metabolites, via the bile is expected.

In conclusion, 2,2’-methylenebis[6-cyclohexyl-p-cresol] has a not very high potential for bioaccumulation, and will be excreted after metabolism.

Data source

Materials and methods

Results and discussion

Applicant's summary and conclusion