[No public or meaningful name is available]

Acute Toxicity: dermal

Administrative data

Endpoint:

acute toxicity: dermal

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

data waiving: supporting information

Reference

Given the high false positive rates of the NRU assay, overall low acute oral toxicity indication from studies on main constituents, the test substance, ‘mono- and di- C16-18 PSE and C16 -18 AE10 PSE’, can be considered to have a low acute oral toxicity potential with LD50 value >2000 mg/kg bw. This is further supported by the low bioavailability potential of the test substance (based on high MW and low water solubility).

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Quality of whole database:

Good quality studies

Endpoint conclusion:

no adverse effect observed

Endpoint conclusion:

no adverse effect observed

Oral:

For the acute oral toxicity endpoint, an acute screening study (OECD 129) test is available with the test substance, ‘mono- and di- C16 -18 PSE and C16 -18 AE10 PSE’. Thein vitroNeutral Red Uptake (NRU) cytotoxicity screening study according to the ECHA R.7a Guidance, cannot be used as a stand-alone test, but could be used within a WoE approach to adapt the standard information requirements for acute oral toxicity. Therefore, the acute oral toxicity endpoint assessment has been based on the acute oral screening study available on the test substance along with supporting studies available for substances representative of the main constituents, which can be categorised as phosphate esters (PSE), ethoxylated phosphate ester (AE PSE) and free ethoxylated alcohol (AE). As, representative studies are not available for the constituent, AE PSE, the endpoint assessment has been based on representative studies available on PSE and AE only, under the assumption that AE PSE is likely to hydrolyse to AE and PSE. The results are presented below:

Acute screening study with the test substance: An in vitro study was conducted to determine the acute toxicity potential of test substance, ‘mono- and di C16 -18 PSE and C16 -18 AE10 PSE’, using cytotoxicity based on Neutral Red Uptake Method, according to OECD Guideline 129, in compliance with GLP. The study was assessedin vitrousing XCellR8’s internally validated Human Cell-Based Screen (Non-Regulatory Method). The test uses cultured human dermal fibroblasts in animal product-free culture, Neutral Red Uptake (NRU) method and a prediction model, based on the GHS classification system for acute toxicity. After a 24 h ± 1 h exposure of 8 concentrations of test substance in cell culture medium of Human Dermal Fibroblasts neonatal (HDFn), cytotoxicity was evaluated. Using a prediction model, determined previously, the IC50 value is converted to a corresponding GHS classification for oral acute toxicity. The percentage of viability for each concentration was calculated and normalised to viability results of the negative control (untreated cells) arbitrarily set to 100%. The IC50 (i.e. the concentration at which a decrease in cell viability of 50% was observed) was calculated as being 119.2 µg/mL in the Range Finding Experiment (RFE); 289.9 µg/mL in the main experiment 1 (ME1); 96.8 µg/mL in ME2 and 77.7 µg/mL in ME3. Based on the study results (IC50: 77.7 to 289.9 µg/mL), the study author concluded, the test substance could fall in potential EU CLP category 4 (LD50: 300 to 2000 mg/kg bw) (XCellR8, 2017). However, it is known that thein vitroNRU cytotoxicity assay has a high false positive rate and, therefore, positive results cannot be readily used in a meaningful way in characterising the acutely toxic substances.

Constituent PSE - read across studies:

Study 1:A study was conducted to determine the acute oral toxicity of the read across substance, mono- and di- C16 PSE, K+ (purity: ca. 85%), according to the OECD Guideline 401, standard acute method, in compliance with GLP. Five male and 5 female Fü-albino SPF rats were randomly selected for an acute oral toxicity study. Fasted rats were given a single dose of the test substance suspended in SSV (Standard Suspended Vehicle) by gavage at a dose level of 5000 mg/kg bw. They were observed for 15 d for toxic signs, mortality and body weight changes. All rats were examined for gross lesions. No compound-related deaths occurred. No compound-related incompatibility reactions were observed. No compound-related effect on body weight development appeared. No compound-related gross or microscopic lesions were observed. The LD50 was determined at >5000 mg/kg bw (i.e., equivalent to 4250 mg a.i./kg bw) (XXXX, 1987).

Study 2:A study was conducted to determine the acute toxicity of the read across substance, di- C16 PSE (purity: 100%), according to the notification no. 118 of the Pharmaceuticals Affairs Bureau, 15 Feb 1984, Toxicity Test Guideline (similar to OECD guideline 401). Groups of fasted, 4 to 6 weeks old CFY (Sprague-Dawley origin) rats, 10/sex were given a single oral dose of test substance in distilled water at doses of 0 (control) and 16 g/kg bw and observed for 14 d. No mortality occurred. Piloerection was observed in all animals in the treated group, however, the animals had recovered on Day 3. No effects on body weight were observed. Terminal necropsy findings were found to be normal. Under the study conditions, the oral LD50 in rats for test substance was determined to be >16000 mg/kg bw (Huntingdon, 1985).

Constituent AE - read across study:

A study was conducted to determine the acute oral toxicity of the read across substance, C16 -18 AE (1- 2.5EO) (purity: 100%), in Wistar rats, according to the OECD Guideline 401, standard acute method. The undiluted test substance was administered to groups of five fasted male and female albino rats at a limit dose of 10 gm/kg bw by oral gavage. Animals were observed for 14 d post-dosing. No deaths occurred at the end of the test. No effects were observed in gross pathology and no treatment related changes were observed on body weight. The only clinical sign of toxicity was piloerection. Under the study conditions, the LD50 of the read across substance was determined to be >10000 mg/kg bw (Mϋrmann, 1986).

Further, a HERA 2009 review report on AEs indicated low to moderate order of acute oral toxicity in the rat with LD50 values ranging between 600 to more than 10000 mg/kg bw. The structure of the test compound influenced acute toxicity determined by the relative number of ethoxy units, whereas, carbon chain length was not correlated with the acute oral toxicity. The degree of ethoxylation of the AE appeared to be the only factor found to be of relevance in acute oral toxicity with the compounds with ethoxylate chains between 5 and 14 being more toxic by oral consumption than those with less than 4 or more than 21 ethoxy units. For example, LD50 values for C9-11 AE with 2.5EO ranged from 2.7 to 10 g/kg bw compared to 1.2 to 2.7 g/kg bw for C9-11 AE8. The same trend was observed for C12-14 and C13-15 AE LD50 values; C12-14AE3 (LD50 9.35 g/kg bw) was less acutely toxic than C12-14AE10 (LD50 2.82 g/kg bw), and C13-15AE4 (LD50 > 5g/kg bw) was less acutely toxic than C13-15AE11 (LD50 2.45 g/kg bw). Clinical findings observed in the test animals after treatment were indicative of gastrointestinal irritation such as ulcerations of the stomach, pilo-erection, diarrhoea and lethargy and may be linked with administration of a bolus dose, in particular in cases where the test substance was administered undiluted. The study which resulted in the lowest LD50 value of 600 mg/kg bw for males and 500 mg/kg bw for females were determined for C15-16AE10. It was noted that this study was not in conducted in compliance with OECD guidelines and GLP regulations. C15-16AE10 was administered to four rats of each sex as a 19% w/v solution in water. Diarrhea and lethargy were observed at the highest dose of 1.5 g/kg bw within 24 h. These signs had subsided in surviving animals within 48 h. On the 9th day, two females were underweight; these animals subsequently died on Days 12 and 15. Necropsies at study termination were not conducted.

Overall, given the high false positive rates of the NRU assay, low acute oral toxicity indication from studies on the main constituents, together with the low bioavailability potential of the test substance (based on high MW and low water solubility), the test substance can be considered to have a low acute oral toxicity potential with LD50 value >2000 mg/kg bw.

Dermal:

As per Annex VIII (8.5.3), the acute dermal toxicity testing is not needed as the substance does not meet the criteria for classification for acute toxicity and STOT SE for the oral route. This is also supported by the absence of any systemic effects in the in vivo skin sensitisation studies available for a substance representative of the main constituent, ‘mono- and di- C16 PSE, K+ and H3PO4’ as well as C16-18 AE (1-2.5EO). Moreover, given the physico-chemical properties of the test substance, the dermal LD50 value is less likely (due to lower absorption potential of dermal route) to be lower than oral LD50 or the oral doses showing clinical signs. Hence, testing via dermal route will less likely result in any additional hazard identification and testing is therefore considered unnecessary.

Inhalation:

The substance is a solid block with a low vapour pressure at room temperature. Due to its physical state and physical chemical properties it is unlikely that this substance will form inhalable dust, mist or fumes during normal processing and use conditions. In case inhalable forms of the substance are created under particular conditions (e.g. spraying, elevated temperature/pressure), appropriate risk management measures such as closed systems, exhaust ventilation or wearing of respirators are implemented to control exposure. Under such conditions, the risk to humans following inhalation exposure can be considered minimal and further testing involving vertebrate animals may be omitted, in accordance with Annex XI (1.2) of the REACH regulation.

Overall based on the available weight of evidence, the test substance, 'mono- and di- C16 -18 PSE and C16 -18 AE10 PSE', is concluded not to warrant classification for acute oral or dermal toxicity, according to the EU CLP criteria (Regulation 1272/2008/EC).

Cross-reference 2

Reason / purpose for cross-reference:

data waiving: supporting information

Reference

Based on the available weight of evidence from studies on substances representative of the main constituents, the test substance, 'mono- and di- C16-18 PSE and C16-18 AE10 PSE', is considered to be non-sensitising to the skin.

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

In absence of skin sensitisation study with the test substance, the endpoint can be assessed based on studies for substances representative of the main constituents, which can be categorised as phosphate esters (PSE), ethoxylated phosphate ester (AE PSE) and/or free ethoxylated alcohol (AE). As, representative studies are not available for the constituent, AE PSE, the endpoint assessment has been based on representative studies available on PSE and AE only, under the assumption that AE PSE is likely to hydrolyse to AE and PSE. This is further supported with the HRIPT availanle for the read across substance, 'mono- and di- C18-unsatd. PSE and C18-unsatd. AE5 PSE'. The results are presented below:

Constituent: PSE - read across study:

A study was conducted to determine the skin sensitisation potential of the read across substance, ‘mono- and di- C16 PSE, K+ and H3PO4’ (98.5%) according to OECD Guideline 429 and US EPA OPPTS 870.2600 (LLNA), in compliance with GLP. The read across substance concentrations selected for the main study were based on the results of a pre-screen test. In the main study, four experimental groups of five female CBA/J mice were treated with read across substance concentrations of 2.5, 5, 10 or 25% w/w for three consecutive days, by topical application on the ears. Five vehicle control animals were similarly treated, but with the vehicle alone (Liquid petrolatum). A positive control group with a-hexylcinnamaldehyde (HCA - 50%) was also included in the experiment. Five days after the last exposure, all animals were injected with 5-bromo-2’-deoxy-uridine (BrdU) and the draining (auricular) lymph nodes were then isolated and pooled for each animal. Cells were fixed using 70% ethanol and used for the measurement of the cell number and the BrdU determination (percentage of proliferating cells in “S” phase). Flow cytometry was conducted for analysis and stimulation index (SI) was recorded. Mortality/viability, body weights, clinical signs, ear size and irritation (and other local effects) were recorded as well. SI values were similar among the control and test groups and were below 3. Three concentrations (i.e.2.5, 5 and 25%) induced ear swelling. No mortality occurred and no clinical signs of systemic toxicity were observed in the animals of the main study. Body weights and body weight gain of experimental animals remained in the same range as controls over the study period. Under the study conditions, the read across substance, was considered to be non-sensitising to the skin (MBRL, 2005).

Constituent: AE - read across study:

A study was conducted to determine the skin sensitivity potential of the read across substance, C16 -18 AE (1 -2.5 EO) (purity not specified), using Buehler test method, according to OECD Guideline 406, in compliance with GLP. In this study 20 female guinea pigs were induced by an epicutaneous occlusive dressing with 100% test substance (in maize oil) for 6 h on Day 0, 7 and 14. Two weeks after the last induction animals were challenged by epicutaneous occlusive exposure for 6 h to 100% test substance (in maize oil). 24 and 48 h after patch removal the application site was assessed for signs of local irritation or sensitisation. No dermal reactions were observed in any test animal at any time point. Under the study conditions, the read across substance was considered to be non-sensitising to the skin (Eisele, 1995).

Further, a HERA 2009 review report on AEs, reported that overwhelming majority of available guinea pig studies in which AEs were tested for skin sensitisation properties demonstrated the absence of skin sensitisation potential with both the Magnusson and Kligman and Buehler protocol. Only one study following the Magnusson and Kligman protocol indicated a weak sensitisation potential of selected AE. No follow-up work was conducted to further investigate the relevance of the observation. However, for structurally similar products the sensitisation reaction was not seen and therefore it was considered that the observed reactions may have been confounded with irritation reactions.

HRIPT with read across substance

A study was conducted to determine the skin sensitivity potential of the read across substance, ‘mono- and di- C18-unsatd. PSE and C18-unsatd. AE5 PSE’, according to the Human Repeated Insult Patch Test (HRIPT). In the induction phase, a 10% test substance preparation (in distilled water; pH 6.49) was applied epicutaneously to 48 healthy volunteers for 1 to 4 d under a semi-occlusive conditions. After a rest period of ca. 2 weeks, the substance was applied under the same conditions to a new site as a challenge. At the end of 24 h, the occluded patch was removed and the site was read for immediate response. Follow-up readings were made 72 h after challenge. Subject #12 developed moderate erythema and mild edema during Day 1 and 2 of induction, barely perceptible response on Day 3 and mild erythema on Day 4. However, two weeks later the subject #12 was challenged with the test substance and there was no response indicating that the initial response was not clinically relevant. Subject # 13 developed spotty erythema on Days 2, 3, and 7. Two weeks later subject #13 was challenged and 24 h later they developed moderate erythema and mild edema. By 72 h post-challenge only spotty erythema was noted (Subject # 13). This pattern of skin reactivity is suggestive of a pre-existing hypersensitivity. No evidence of induced allergic contact sensitisation was recorded in any other volunteers. Under the study conditions, the read across substance did not indicate a potential for dermal irritation or allergic contact sensitization in human volunteers at 10% test concentrations (Croda, 2004). Based on the results of the read across study, a similar absenc of skin sensitisation in HRIPT can be expected for the test substance, 'mono- and di- C16-18 PSE and C16-18 AE10 PSE'.

Overall, based on the available weight of evidence from studies on the main constituents, the test substance, ‘mono- and di- C16 -18 PSE and C16-18 AE10 PSE’, is considered to be non-sensitising to skin.

Endpoint conclusion:

no study available

Based on the available weight of evidence from studies on substances representative of the main constituents, the test substance, 'mono- and di- C16-18 PSE and C16-18 AE10 PSE' is concluded not to warrant classification for skin sensitisation, according to the EU CLP criteria (Regulation 1272/2008/EC).