1,3,4,6,7,8-hexahydro-4,6,6,7,8,8-hexamethylindeno[5,6-c]pyran

Endpoint summary

Description of key information

Oral absorption: 50%
Dermal absorption: 5.2% based on available human dermal absorption study
Inhalation absorption: 100%

Key value for chemical safety assessment

Bioaccumulation potential:

low bioaccumulation potential

Absorption rate - oral (%):

50

Absorption rate - dermal (%):

5.2

Absorption rate - inhalation (%):

100

Additional information

Introduction:The test material HHCB (Cas no. 1222-05-5 ) is a polycyclic musk, a hexyl ring with an ether bond, with a benzyl and a pentyl ring attached to it. On the pentyl ring methyl groups are attached to each C atom. The substance is a highly viscous liquid with a molecular weight of 258.4 that does not preclude absorption. The test material is not likely to hydrolyse and has a low volatility (0.0727 Pa).

Absorption:

Oral: The results of the 90-day repeat oral dose (dietary) and oral (gavage) developmental toxicity study show that the substance is being absorbed by the gastro-intestinal tract following oral administration because some effects on blood chemistry were seen, although considered of minor toxicological significance and likely reflecting slight nutritional effects. The relatively low molecular weight and the moderate octanol/water partition coefficient (Log Kow 5.3 and water solubility (1.65 mg/l) would allow absorption through the gut. According to Martinez and Amidon (2002) the optimal log Kow for oral absorption falls within a range of 2-7. This shows that HHCB is likely to be absorbed orally and therefore the oral absorption is expected to be > 50%. 

Skin: The substance is not irritating to skin and eye and not a skin sensitizer. Based on the physico-chemical characteristics of the substance, being a liquid, its molecular weight (MW 258.4, log Kow (5.3) and water solubility (1.65 mg/l), indicate that (some) dermal absorption is likely to occur. The optimal MW and log Kow for dermal absorption is < 100 and in the range of 1-4, respectively (ECHA guidance, 7.12, Table R.7.12-3). HHCB is just outside this range optimal range and therefore the skin absorption is not expected to exceed 50%.

Several experimental studies are available to determine dermal absorption. In the key study, the dermal absorption (non-GLP, but with QA statement) of HHCB was determined over a 24-hr period according to the methodology of the SCCNFP. Radiolabeled HHCB (uniformly labelled in the aromatic ring – radiochemical purity 99.3%) was applied in 1% solution in ethanol (96% v/v) to human epidermal membranes (prepared from female breast or abdominal skin and assayed for integrity with tritiated water) supported on a piece of filter paper (for strength) in glass diffusion cells (n=12). The area of the membrane available for absorption was approximately 1 cm2and the average applied dose was 20±0.2 µL/cm2. The amount of material absorbed into the receptor phase, 6% Volpo N20 (to enhance solubility) in pH 7.4 phosphate buffered saline, after 24 hr was 0.40±0.06% of the applied dose. The majority of applied HHCB (81±2% of the applied dose) was found in the 24-hr surface wipe and donor chamber wash plus wipe. The stratum corneum tape strips contained 5.8±0.8% of the applied dose and the remaining stratum corneum plus epidermis 4.5±0.6% of the applied dose. Levels of HHCB in the remaining stratum corneum plus epidermis, filter paper (on which the epidermis samples rested) and permeated HHCB were combined to produce a total absorbed dose value of 5.2±0.6% of the applied dose. Overall recovery of radioactivity was 92±0.8% (Green and Brain, 2001).

From a supporting study (not summarised), it appeared that under similar conditions about 2.4% of an applied dose might evaporate. Because this study used a 24-hr application and because of the limitations of the human simulated exposure study (primarily the small number of subjects) this figure is used as a conservative estimate for absorption of HHCB via human skin in the risk characterization.

Two other supporting studies on dermal absorption are available. In an in vivo human study, a 6 hr unoccluded exposure with 0.4% HHCB in 70% ethanol, intended to simulate a typically high exposure from the use of alcohol-based products such as perfumes or eaux de toilette, resulted in considerable absorption into the skin (approximately 20%). However, most of the material in this skin reservoir was not absorbed but was recovered from dressings over the site of exposure over a 120 hr period presumably from reverse diffusion and/or desquamation. Based on amounts excreted, primarily in the urine, approximately 0.1% was actually absorbed under the conditions of this experiment. In addition, because the application was unoccluded about 22% of the applied dose may evaporate under the conditions of the test, which may explain that about 14% of the radioactivity was not recovered in the in vivo human dermal absorption study (Hawkins et al., 1996b, Ford et al., 1999). An in vivo study in rats supports the assumption that a good indication of the amount absorbed is the amount excreted (Hawkins, 1995).

A similar picture was seen in vivo with rats where the material was applied for 6 hr under occlusion in 70% alcohol. Here again, a reservoir in the skin of about 10% of the applied dose was formed after the six-hr application with about 5% of this reservoir being lost presumably from reverse diffusion and/or desquamation to the dressing 120 hr after dose removal. Based on the amount remaining in the tissues, including that at the site of dosing, at sacrifice (2.7%) and the amount excreted (13%) almost all (11.6%) of which was in the faeces, a total absorption under the conditions of this experiment of ~ 16% can be concluded. The principal differences from the human study were the much larger absorption as a result of the application under occlusion and the well-known fact that rat skin is more permeable than human. For the risk characterisation, a value of 16% will be used to estimate the dermal absorption via rat skin. This value also includes the continued absorption from the dermal reservoir at 120 hr (2% of the dose) (Hawkins, 1995, Ford, 1999).

Lungs: Absorption via the lungs is also indicated based on these physico-chemical properties. Though the inhalation exposure route is thought minor, because of its low volatility (0.0727 Pa), the octanol/water partition coefficient (Log Kow 5.3), indicates that inhalation absorption is possible. The blood/air (BA) partition coefficient is another partition coefficient indicating lung absorption. 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 HHCB the B/A partition coefficient would result in:

Log P (BA) = 6.96 – 1.04 Log (0.0727) – 0.533 5.3 – 0.00495 258.4 = 4.04

This means that the substance has a high 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. Despite HHCB being somewhat out of the applicability domain and the exact B/A may not be fully correct, it can be seen that the substance will be readily absorbed via the inhalation route and will be close to 100%.

Distribution:The fairly low water solubility of the test substance would limit distribution in the body via the water channels. The log Kow would suggest that the substance would pass through the biological cell membrane. Due to the expected metabolisation the substance as such would limitedly accumulate in the body fatwhich is confirmed in a bioaccumulation study in fish resulting in a BCB of 1600.

Metabolism: The metabolisation of the substance is assessed using OECD Toolbox 3 liver metabolism simulator. The formed metabolites are mainly:

1) the hexylring with the ether bond will be oxidised and form additional ketone or acid groups;

2) the hexylring with the ether bond will open and alcohols may be formed;

3) the methyl groups attached to the pentyl ring will be oxidized into alcohols, ketones and/or acids.

HHCB metabolites according to QSAR toolbox rat liver simulator

 

  

Fig. 1 The theoretical metabolisation of HHCB can occur via hydroxylation of the methyl groups of the pentyl ring and/or an acid group can be formed at C4 as predicted by the OECD Toolbox 3.0 liver metabolism simulator.

 

These metabolites are expected to be more water soluble, have a lower Log Kow values and will therefore be more easily excreted. 

Excretion:The intravenous studies in rats and the pig showed that HHCB is rapidly distributed and is excreted primarily in the faeces by the rat as was seen in the dermal study (~ 68% of total excretion as opposed to ~90% after dermal exposure) but in the pig the principle route of exposure is in the urine. In neither of these studies was any evidence of accumulation seen. However, clearance from the fat was considerably slower than from other tissues. It is noteworthy that in neither of these studies was any of the urinary radioactivity present shown to be present as unmetabolised HHCB, however the faeces, which is the major excretion route of the rat, was not analysed for metabolites or parent (Hawkins, 1997a, Hawkins, 1997b).

An oral study with pregnant and later lactating rats shows that orally dosed HHCB and HHCB metabolites can end up in the milk. The levels seen in the milk of the lactating dams can aid in the interpretation of the study (see Table 4.14 above) where neonate rats were exposed to HHCB and its metabolites through nursing. HHCB is also found in human milk at levels up to 1316 μg/kg fat (equivalent to 48 μg/kg whole milk based on a measured fat content of 3.67%) and in adipose tissue at levels ranging from 12 – 189 μg/kg fat (Hawkins et al., 1996a).

Discussion:The substance is expected to be readily absorbed, orally and via inhalation, based on the human toxicological information and physico-chemical parameters. For dermal absorption of HHCB in rats and humans, values of 16 and 5.2% are taken forward to the risk characterization, based on experimental information.

The IGHRC (2006) document of the HSE and mentioned in the ECHA guidance Chapter 8 will be followed to derive the final absorption values for the risk characterisation.

Oral to dermal extrapolation:There are adequate data via the oral route and the critical toxic effect is related to systemic effects and therefore route to route extrapolation is applicable. The toxicity of the substance will be due to the parent compound but also to its metabolites. The overriding principle will be to avoid situations where the extrapolation of data would underestimate toxicity resulting from human exposure to a chemical by the route to route extrapolation. HHCB is not expected to be detoxified in the gut because it is hydrolytically stable. Though some first pass effect via the liver may occur the toxicity via the dermal route will not be underestimated because absorption will be slower (as has been shown experimentally) and the compound will also pass the liver. Using the asymmetric handling of uncertainty the oral absorption will be considered 50% (though likely to be higher) and the dermal absorption will be based on the in vitro experimental study: 5.2% for humans.

Oral to inhalation extrapolation:Though HHCB is not a volatile liquid some inhalation exposure will be calculated. HHCB is not a corrosive for skin and eye and systemic effects will overrule the effects at the site of contact. In the absence of bioavailability data it is most precautionary that 100% of the inhaled vapour is bioavailable. For inhalation absorption 100% will be used for route to route extrapolation, because this will be precautionary for the inhalation route.

Conclusion:HHCB is expected to be readily absorbed via the oral and inhalation route based on toxicity and physico-chemical data. Using the precautionary principle for route to route extrapolation the final absorption percentages derived are: 50% oral absorption, and 100% inhalation absorption. For dermal absorption of HHCB in rats and humans, values of 16 and 5.2% are taken forward to the risk characterization, based on experimental information.

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. 

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

 

(Summary in line with EU-RAR, 2008)