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Description of key information

OTNE is absorbed via the oral route and dermal route for 86 and 15%, respectively, based on experimental in vivo toxico-kinetic information using radiolabel. Using the precautionary principle for the inhalation route 100% inhalation absorption will be selected. The radiolabelled substance is fully metabolised into more hydrophilic metabolites, as is shown in HPLC chromatograms and it is reasoned that these are (partly) glucuronic conjugates of the substance. The DT50 of the substance after oral exposure is 34.3 hours (20 mg/kg bw), after dermal exposure it is 70 (55 mg/kg bw) and 40 (550 mg/kg bw) hours after application, respectively. Via oral exposure the main the main excretion pathways are via urine and bile. Via dermal exposure the urine is the main excretion pathway.

Key value for chemical safety assessment

Bioaccumulation potential:
low bioaccumulation potential
Absorption rate - oral (%):
86
Absorption rate - dermal (%):
15
Absorption rate - inhalation (%):
100

Additional information

The toxico-kinetic assessment of OTNE for humans and air breathing mammals

In this toxico-kinetic assessment the experimental and theoretical consideration of the fate of OTNE in humans and mammals will be presented. There is oral and dermal kinetic information available from several kinetic and repeated dose toxicity studies. First the overall toxico-kinetic assessment is presented and thereafter the executive summaries of the experimental studies.

Introduction

The toxico-kinetic assessment of OTNE, with its constituents and impurities (it is a reaction mass) is presented below.

Structure description: The test material OTNE has three main constituents which all have a double hexyl-ring, two methyl groups are attached to one C atom in the first ring. The second ring has one C atom with one methyl group and one C atom with a methyl group and a ketone-methyl group. Also the impurities have a similar structure. There is one impurity where the right ring is open instead of closed. The double bond is either between the two rings or on the left or right ring.

Relevance of information derived from radiolabelled OTNE: The toxico-kinetic experiments are carried out with radiolabelled OTNE. In view of OTNE being a multi-constituent it is considered appropriate to derive the kinetic values on this radioactivity because the assessment then includes all constituents and impurities. It is also considered conservative because also metabolites and residual carbons will be included in this assessment.

Physico-chemical properties: The substance is a liquid with a molecular weight of 234.4, a volatility of (0.233 Pa), a water solubility of (2.68 mg/l) and a log Kow of 5.65.

Absorption:

Oral: the absorption of OTNE was studied by the NTP (2014) using [14C]OTNE (β-isomer) in male F344 rats following gavage administration of 20 mg/kg bw, resulting in an oral absorption of 86%, which will be forwarded to the risk characterisation.

Skin: Key information is an in an in vivo skin absorption study with male F344 rats following a single dermal application of 55 or 550 mg/kg bw [14C]β-OTNE in ethanol to covered sites after 24, 48, or 96 hours. The dermal absorption of 15% is used (derived from 550 mg/kg bw) because the recovery was highest for this dose. The information from the supporting in vitro study is in line with this 15% value showing 16.5% skin absorption, but with a lower recovery compared to the in vivo study. Therefore the in vivo study is selected as key and 15% dermal absorption is selected for the risk characterisation.

Lungs: Absorption via the lungsis also indicated based on the physico-chemical properties. Though the inhalation exposure route is considered to be of minor relevance, because of its low volatility (0.233 Pa), the potential for lung absorption is assessed. The blood/air (BA) partition coefficient is thought to be relevant for humans and mammals because when the substance enters the airways the first partitioning step is between blood and air. Buist et al. 2012 have developed BA model for humans using the most important and readily available parameters:

Log PBA = 6.96 – 1.04 Log (VP) – 0.533 Log (Kow) – 0.00495 MW.

For OTNE the B/A partition coefficient would result in:

Log P (BA) = 6.96 – 1.04*Log (0.233) – 0.533*5.65 – 0.00495*234.4 = 3.44

This means that the substance has a tendency to go from air into the blood. It should, however, be noted that this regression line is only valid for substances which have a vapour pressure > 100 Pa. From these calculations we would expect absorption via inhalation to be 100%.

Distribution

In the oral in vivo toxico-kinetic studies in rats (NTP, 2014) it is shown that the oral DT50 is 34.3 hours (at 20 mg/kg bw). Tmax via oral exposure is at 8 hours and Cmax is almost 1 mg/l blood after 20 mg/kg bw exposure, (843 ng/g tissue) again based on all radioactivity.

In the dermal in vivo toxico-kinetic study in rats (NTP, 2014) the DT50 values are 70 and 40 hours, for the low and high dose of 55 and 550 mg/kg bw, respectively. Tmax via dermal exposure is reached within 24 hours and Cmax is 0.9 mg/l at the low dose (55 mg/kg bw). OTNE is distributed in all organs blood/tissue concentrations. Via oral dosing blood/tissue concentration after 24 hours are 1:10 in liver, bladder and pancreas. In adipose tissue it is 1:3, in testes it is 1: 0.5, in brain it is 1: 0.2. In all other organs the distribution is roughly 1:1 based on radioactivity. In the in vivo dermal studies the substance was distributed as follows: 1:17 in skin; 1:14 in bladder; 1:7 in kidney, liver, pancreas (roughly) also in adipose tissue; in brain it is 1:0.5. In all other tissues it is 1:1 based on radioactivity.

Adipose tissue: This shows that in adipose tissue the distribution is 1:3 and 1:7 after 24 hours via oral and dermal route respectively showing that it is limitedly distributed in body fat. In addition, in the NTP studies no parent OTNE could be found. The determination of the peaks showed that the likely metabolites were glucuronic conjugates of OTNE, indicating full metabolism of OTNE.

Milk transfer: In a RIFM study (2001) the transfer of OTNE to milk and placenta is studied during and after pregnancy using radiolabelled OTNE. The parent compound, OTNE, was not detected in extracted milk samples at both dose levels but radioactivity. Exposure to 2 and 20 mg/kg bw resulted roughly in 0.2 and 2 ppm radioactivity in the milk. From the HPLC chromatograms of the milk samples it can be seen that the metabolites have lower retention times and therefore are more hydrophilic . This indicates that radioactivity detected is related to metabolites of OTNE. In a Chinese study OTNE has been detected at low levels in mother’s milk (3.9 ng/g lipid ca 0.175 ug/l milk assuming 3.5% fat in milk) (Yin et al., 2016). The exposure in this paper is related to exposure via cosmetics.

Whole body autoradiographs: OTNE related radioactivity has been measured in whole-body autoradiographs showed barely detectable radioactivity in placenta and foetuses at 4 and 24 hours post dosing, indicating negligible transfer of parent OTNE or metabolites across the placenta. Highest levels of radioactivity were observed in the content of the small intestine and in preputial glands, followed by stomach, liver, large intestinal contents, thyroid, and bladder.

Metabolism for humans and air breathing mammals

OTNE is fully metabolized as no parent OTNE is detected in several in vivo toxico-kinetic studies. The substance is extensively conjugated as is measured in male rats using bile cannulated rats. The alpha-2u hydrocarbon nephropathy seen in the kidneys show that the substance is partly metabolized in the lysosomes. Due to the low pH in the lysosomes, the ketone becomes reduced to a secondary alcohol which is then conjugated with alpha-2u globulin and transported to the kidneys.

There is no possibility of the formation of a gamma-diketone, because OTNE has no ethyl groups at the side of the ketone functional groups. This gamma-diketone is the likely metabolite of AETT (Acetyl ethyl tetramethyl tetralin: Cas no 88-29-9) causing neurotoxicity at low doses (see neurotoxicity section).

Excretion for humans and air breathing mammals

The in vivo oral toxico-kinetic study shows the majority of the administered dose was excreted in urine (28%) and faeces (39%) after 48 hours following gavage administration of 20 mg/kg. Approximately 73% of the 20 mg/kg dose was eliminated in bile in 48 hours confirming that the faecal excretion was not due to poor absorption but biliary excretion. Additionally, approximately 80% of the dose in the bile was excreted within the first 4 hours, while the faecal excretion continued through 48 hours suggesting some enterohepatic recirculation of OTNE. The in vivo dermal kinetic study shows that OTNE is mainly excreted via the urine.

Discussion for humans and air breathing mammals

Key information on bioaccumulation including air breathing organisms:

According to the criteria in the PBT guidance on air breathing organisms (2017) OTNE fulfils the screening criteria for concern for bioaccumulation in air breathing organisms: log Kow (5.6: > 2) and log Koa (6.9: > 5). The concern for air-breathing organisms is relevant for non-metabolising substances, which are not excreted via kidneys and for which the ventilation rate between air and blood is lower than for fish. This type of bioaccumulation is not relevant for metabolizing substances as Gobas et al. explicitly mentions (2020, figure 6, D, assessing oxygen containing substances). The half-life in fish in the BCF test is 1.2 days. OTNE is an oxygen (ketone) substance with a hydrocarbon unsaturated backbone with two-ring structures. The ketone is expected to be reduced to an alcohol because glucuronidation is the key excretion pathway and this acid can only conjugate with an alcohol bond. This transformation is expected to occur in all (air-breathing) organisms. Glucuronic acid has a low log Kow (<-1) and also OTNE-glucuronidated will have the acidic group and therefore this log Kow < -1. This glucuronate will be excreted via the kidneys. In addition, in mammals (rat) the DT50 was 1.4 days after oral doses of 20 mg/kg bw showing absence of bioaccumulating in fish and in air-breathing organisms. This means that OTNE and its degradants or metabolites are not a concern for air-breathing organisms.

Other criteria indicating not a concern for air breathing organisms are met: 1) extensive metabolisation is seen in all studies (no parent substance detected, anticipated glucuronic metabolites are found using radiolabel); 2) low distribution in body fat such as mother’s milk and adipose tissue (based on radiolabel and therefore concerning metabolites and residual carbon) and; 3) low DT50 < 1.4 and 3 days (based on radiolabel from in vivo oral and dermal studies respectively); is supported with the experimental DT50 in fish being 1.2 days and a BCF value: 391.

Key information on absorptionwhich will be brought forward to the risk characterisation: The substance is readily absorbed orally up to 86% and via inhalation expected to be up to 100%, based on the available human toxicological information and physico-chemical parameters. Based on the key in vivo dermal absorption study the absorption via this route is 15%.

In view of the available experimental in vivo toxico-kinetic information the IGHRC (2006) document of the HSE and mentioned in the ECHA Guidance Chapter R.8 will be followed only for the inhalation route.

Oral to inhalation extrapolation: Though OTNE is not a volatile liquid some inhalation exposure will be calculated. OTNE is not corrosive to skin and eye and the systemic effect will overrule the effects at the site of contact. In the absence of inhalation absorption data it is most precautionary that 100% of the inhaled vapour is bioavailable. For the oral absorption 86% will be used and 15% for the dermal route when using route to route extrapolation.

Oral to dermal extrapolation: The experimental oral and dermal absorption values will be used: 86 and 15%, respectively.

Conclusion:

OTNE is absorbed via the oral route and dermal route for 86 and 15%, respectively, based on experimental in vivo toxico-kinetic information using radiolabel. Using the precautionary principle for the inhalation route 100% inhalation absorption will be selected. The radiolabelled substance is fully metabolised into more hydrophilic metabolites, as is shown in HPLC chromatograms and it is reasoned that these are (partly) glucuronic conjugates of the substance. The DT50 of the substance after oral exposure is 34.3 hours (20 mg/kg bw), after dermal exposure it is 70 (55 mg/kg bw) and 40 (550 mg/kg bw) hours after application, respectively. Via oral exposure the main the main excretion pathways are via urine and bile. Via dermal exposure the urine is the main excretion pathway.

References

- Buist, H. E., Wit-Bos de, L., Bouwman, T., Vaes, W. H. J., 2012, Predicting blood: air partition coefficient using basis physico-chemical properties, Regul. Toxicol. Pharmacol., 62, 23-28.

Gobas, F.A.P.C, Lee, Y-S, Lo, J.C., Parkerton, T., Letinsky, D., 2020, A toxicokinetic framework and analysis tool for interpreting Organisation for Economic Co-operation and Development Guideline 305 Dietary Bioaccumulation test, Env. Toxicol. Chem., 39, 171-188.

- Martinez, M. N., And Amidon, G. L., 2002, Mechanistic approach to understanding the factors affecting drug absorption: a review of fundament, J. Clinical Pharmacol., 42, 620-643.

- IGHRC, 2006, Guidelines on route to route extrapolation of toxicity data when assessing health risks of chemicals, http: //ieh. cranfield. ac. uk/ighrc/cr12[1]. Pdf

- NTP, 2014, Waidyanatha, S., and Ryan, K, Disposition of fragrance ingredient [14C]1-(1,2,3,4,5,6,7,8-octahydro-2,3,8,8-tetramethyl-2-naphthalenyl)ethanone in male Fisher rats following oral administration and dermal application Xenobiotica  Vol. 44 , Iss. 8, 749-756.

- Yin, J., Wang, H., Li, J., Wu, Y.,  Shao, B., 2016, Occurrence of synthetic musks in human breast milk samples from 12 provinces in China,Food Additives & Contaminants: Part A,Volume 33 (7), Pages 1219-1227