Cyclohexyl methacrylate

Endpoint summary

Description of key information

Based on physicochemical properties of the test item and toxicokinetic studies performed with structural analogue substances, CHMA is most likely absorbed after oral and dermal exposure as well as via inhalation route. Once absorbed, the test item is rapidly hydrolysed within the body whether in local tissues, the blood or ultimately within the liver by non-specific esterases to methacrylic acid and the corresponding alcohol cyclohexanol. The primary metabolites methacrylic acid as well as the corresponding alcohol is cleared rapidly in all cases. On the basis of the rapid metabolism and short half-lives a systemic accumulation of the esters and their metabolites is not expected.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Additional information

Cyclohexyl methacrylate (CHMA) is comparable to the lower methacrylate esters, of which there are extensive data available for the C1 methyl ester (MMA) and this has been reviewed in the EU Risk Assessment (2002). Sufficient data is available to confirm applicability of this data across all members of the C1-C8 methacrylate category and, in the case of C2 to C8 esters, this has been reviewed and confirmed in the OECD SIAR (2009).

Absorption

Oral route

 According to ECHA Guidance R.7c, the smaller the molecule the more easily it may be taken up via oral route (ECHA, 2014). Oral absorption is favoured for CHMA as their molecular weight is below 500 g/mol. CHMA is favourable absorbed by passive diffusion due to their moderate log Pow values (between -1 and 4). In addition, clinical signs or changes in clinical pathology parameters were observed after single or repeated oral dosing indicating oral absorption.

Dermal route

Due to water solubility between 100-10000 mg/L and a log Pow value between 1 and 4 uptake of cyclohexyl methacrylate ester is likely after dermal exposure.

In addition, in vitro skin absorption studies in human skin indicate that the structural analogue methyl methacrylate (MMA) can be absorbed through human skin, absorption being enhanced under occluded conditions. However, only a very small amount of the applied dose (0.56 %) penetrated the skin under unoccluded conditions (presumably due to evaporation of the ester from the skin surface (CEFIC, 1993)). Jones (2002) studied the permeability of separated rat and human skin to alkyl methacrylate esters as part of a PhD thesis on development of a Physiologically Based Pharmacokinetic (PBPK) model to predict the pharmacokinetics and toxicity of methacrylate esters. Data on MMA and further alkyl methacrylates as n-butyl methacrylate (n-BMA) and 2-ethylhexyl methacrylate (2-EHMA) indicate that rat skin is more permeable to the absorption of these esters than human skin and there is a steep decline in the rate of penetration across the category from MMA to the larger ester, 2-EHMA. The rate of penetration of ethyl methacrylate (EMA) was predicted to be between that of MMA and n-BMA and that of iso-butyl methacrylate (i-BMA) comparable to n-BMA.

 

Table 1: Summary of the results for the peak rates of absorption of alkyl methacrylate estersthrough rat & human epidermis(adapted from Jones, 2002)

	Rat epidermis			Human epidermis
Ester	Peak rate of absorption (μg/cm²/hr) ±SEM	Period of peak absorption rate (hours)	% age of applied dose absorbed over x hours	Peak rate of absorption (μg/cm²/hr) ±SEM	Period of peak absorption rate (hours)	% age of applied dose absorbed over x hours
MMA	5888±223	2-8	46 % / 16h	453±44.5	4-24	10 % / 24h
EMA	4421	-	-	253	-	-
i-BMA	1418	-	-	80	-	-
n-BMA	1540±69	0-6	18 % / 24h	76.7±9.8	0-24	2 % / 24h
2-EHMA	234±4.8	0-30	7.8 % / 30h	22.7.7±3.7	3-24	0.6 % / 24h

Key: The values in normal type were obtained experimentally, whilst those in italics, are predicted values based on statistical analysis (single exponential fit) of the experimental data

 

The in vitro measured data of Jones is very comparable to the predictions made later by Heylings who used a QSPeR model for whole human skin based on that described by Potts and Guy (1992) to predict the dermal penetration rate of a large number of methacrylate esters, including members of the lower alkyl category. Quantitatively the trend of decreased skin penetration rate across the C1-C8 methacrylate category was confirmed.

 

Table 2: QSAR prediction of absorption of methacrylate esters through whole human skin (extract from Heylings, 2013)

Substance	Molecular Weight	Log P	Predicted Flux (μg/cm2/h)	Relative Dermal Absorption
MMA	100.12	1.38	64.422	Moderate
EMA	114.14	1.87	36.132	Moderate
iBMA	142.2	2.95	14.271	Moderate
n-BMA	142.2	3.03	12.458	Moderate
2-EHMA	198.3	4.95	1.126	Low

 

Based on comparable physicochemical properties and in accordance to ECHA Guidance R.7c, the CHMA can be categorized between n-BMA and 2-EHMA and thus dermal uptake is anticipated to be low to moderate. However, results of the acute dermal toxicity study indicating no systemic signs.

 

Inhalation

Considering the low vapour pressure (< 0.5 kPa) and the resulting low volatility of CHMA exposure as vapour is limited. However, absorption via inhalation is possible as absorption following ingestion did also occur. Generally, liquids are able to readily dissolve into the mucus lining the respiratory tracts. Lipophilic substances (log Pow > 0) have the potential to be absorbed directly across the respiratory tract epithelium by passive diffusion. The acute inhalation study and repeated dose inhalation study conducted with structural analogue substances (MMA, n-BMA) suggested a low potential for toxicity. Additionally, after inhalation exposure to rats, 10 to 20 % of MMA is deposited in the upper respiratory tract where it is metabolized by non-specific esterases to the acid, MAA (Morris, 1992)).

Metabolism/Distribution

Studies confirmed that all lower alkyl-methacrylate esters are rapidly hydrolysed by ubiquitous carboxylesterases to methacrylic acid and the corresponding alcohol (Table 3, adapted from Jones; 2002).The carboxylesterases are a group of non-specific enzymes that are widely distributed throughout the body and are known to show high activity within many tissues and organs, including the liver, blood, GI tract, nasal epithelium and skin (Satoh & Hosokawa, 1998; Junge & Krish, 1975; Bogdanffy et al., 1987; Frederick et al., 1994). Activities of local tissue esterases of the nasal epithelial cells appear to be lower in man than in rodents (Green, 1996 later published as Mainwaring, 2001). Those organs and tissues that play an important role and/or contribute substantially to the primary metabolism of the short-chain, volatile, alkyl-methacrylate esters are the tissues at the primary point of exposure, namely the nasal epithelia and the skin, and systemically, the liver and blood.

First pass (local) hydrolysis of the parent ester has been shown to be significant for all routes of exposure. For example, no parent ester can be measured systemically following skin exposure to EMA and larger esters, as the lower rate of absorption for these esters is within the metabolic capacity of the skin (Jones, 2002). Parent ester will also be effectively hydrolysed within the G.I. tract and within the tissues of the upper respiratory tract (particularly the olfactory tissue). Systemically absorbed parent ester will be effectively removed during the first pass through the liver resulting in their relatively rapid elimination from the body.

Table 3: Rate Constants for ester hydrolysis by rat-liver microsomes and predicted systemic fate kinetics following i.v. administration

Ester	Rat liver microsomes (100mg ml-1) Vmax               Km    (nM min-1 mg-1)      (mM)		CL (%LBF)	T50% (min)	Cmax (MAA) (mg L-1)	Tmax (MAA) (min)
MMA	445.8	164.3	98.8%	4.4	14.7	1.7
EMA	699.2	106.2	99.5%	4.5	12.0	1.8
i-BMA	832.9	127.4	99.5%	11.6	7.4	1.6
n-BMA	875.7	77.3	99.7%	7.8	7.9	1.8
HMA	376.4	34.4	99.7%	18.5	5.9	1.2
2EHMA	393.0	17.7	99.9%	23.8	5.0	1.2
OMA	224.8	11.0	99.9%	27.2	5.0	1.2

HMA – hexyl methacrylate; OMA – octyl methacrylate. Fate kinetics determined using the “well-stirred” model; CL%LBF – Clearance as percentage removed from liver blood flow i.e. first pass clearance; T50% - time taken for 50% of parent ester to have been eliminated from the body; Cmax – maximum concentration of MAA in circulating blood; Tmax – time in minutes to peak MAA concentration in blood “Jones, 2002”.

 

Ester hydrolysis is considered to be the major metabolic pathway for alkyl-methacrylate esters, with GSH conjugation only playing a minor role in their metabolism, and then possibly only when very high tissue concentrations are achieved. (McCarthy and Witz, 1991; Elovaara et al., 1983). In addition, data with n-BMA indicate that GSH conjugation is negligible (McCarthy et a., 1994). Hence, ester hydrolysis is considered to be the only significant metabolic pathway for n-BMA.

The same metabolism can be assumed for CHMA which is primarily hydrolysed to methacrylic acid and its corresponding alcohol cyclohexanol. The

hydrolysis of CHMA, n-BMA, t-BMA and MMA were determined in liver S9 fraction and plasma from rats by determining the degradation of methacrylates and forming of methacrylic acid. Both parameters were also examined in blood after treatment of rats with MMA. In general, the degradation rate in liver S9 fraction decreased with increasing chain length and degree of branching. The degradation rates in plasma were slower in plasma in comparison to liver S9 fraction. However, hydrolysis was independent regarding chain length and degree of branching. Degradation of MMA was comparable in plasma and blood (Roos, 2015).

An in vitro metabolism study consisting of two assays was performed to investigate the metabolic stability of cyclohexyl methacrylate in subcellular fraction of liver of rats and humans (BASF, 2020).

The cyclohexyl methacrylate was incubated in duplicates at a nominal concentration of 500 µM at 37°C with rat liver S9 ranging from 0 (t=0) up to 30 min (first and second run) and with human liver S9 ranging from 0 (t=0) up to 240 and 120 min (first and second run, respectively).

The second assay was performed with the inclusion of perdeuterated methacrylic acid (D6-methacrylic acid) as an internal standard in the incubations. The application of the internal standard allowed to refer the analytics of methacrylic acid to the internal standard and therewith calculate an individual recovery for each incubate / analytical sample.

In the first assay cyclohexyl methacrylatewas already degraded completely after 5 min in the in vitro metabolism system of rats, whereas more or less complete degradation in the human system was observed after an incubation period of 240 min. For the assessment of these data, it may be taken into consideration that the metabolic turn over value of the positive control substrate testosterone parallels in principle this species dependency and demonstrates the common trend of a generally lower activity of xenobiotic metabolizing enzymes in human versus rat in vitro systems.The in vitro data demonstrate that cyclohexyl methacrylate is metabolized in liver S9 fraction of rats and humans. Detected amounts of methacrylic acid in the incubates are able to proof hydrolysis of the tested ester in principle. The quantified amounts of this hydrolysis product however are stoichiometrically lower than the degraded amounts of cyclohexyl methacrylate, indicating that the formed methacrylic acid might be further metabolized.

In the second assay cyclohexyl methacrylate was also metabolized in liver S9 fraction of both species under the chosen incubation conditions. Whereas in the applied in vitro metabolism system of rats the test substance was degraded completely after 30 min, degradation in the human system was observed to about 95 % after 120 min. As in the first assay, the quantified amounts of methacrylic acid were stoichiometrically lower than the degraded amounts of cyclohexyl methacrylate. However, the internal standard D6-methacrylic aciddecreased also with increasing incubation time. It may be assumed that this degradation is due to metabolism of the compound in the liver S9 incubations. When the loss of D6-methyacrylic acid is calculated and added to the amounts of detected methacrylic acid in the respective liver S9 incubations, the corrected amount (in % of formed methacrylic acid) correlated generally to the degradation of the test substrate cyclohexyl methacrylate. Although the hydrolysis product of cyclohexyl methacrylate is detected stoichiometrically underproportionally to the degraded cyclohexyl methacrylate, the correction by the loss of the internal standard D6-methyacrylic acid indicates that this may be due to further degradation of the acid. Consequently, hydrolysis is considered as major metabolic pathway for the ester cyclohexyl methacrylate in rats and humans.

Subsequent metabolism of the primary metabolites within the body and excretion

Again, taken from the OECD SIAR: Methacrylic acid and the corresponding alcohol are subsequently cleared predominantly via the liver (valine pathway and the TCA (TriCarboxylic Acid) cycle, respectively).

There are no studies which specifically address the metabolism of exogenously applied methacrylic acid. However it is generally accepted that methacrylic acid-coenzyme-A is a naturally occurring intermediate of the valine pathway. Methacrylic acid-CoA is rapidly converted into (S)-3-hydroxyisobutyryl-CoA by the enzyme enoyl-CoA-hydratase. This pathway joins the citrate cycle, carbon dioxide, and water being the final products (Rawn, 1983; Shimomura et al., 1994; Boehringer, 1992).”

In terms of the corresponding alcohol metabolite for these esters, they are all subsequently rapidly metabolised, primarily in the liver, by oxidative pathways involving aldehyde dehydrogenase (ALDH), alcohol dehydrogenase (ADH), cytochrome P450 (CYP2E1), and catalase enzymes to the corresponding aldehyde and acid before ultimately being converted to CO2. In the case of the more volatile alcohols a significant portion is excreted un-metabolised in urine and/or exhaled air.

The toxicokinetic and metabolism of cyclohexanol (CH) were studied in 8 human volunteers (4/sex) exposed to 236 mg/m³ CH for 8 h. The retention in the respiratory tract and the total absorbed dose were determined and the urine which was collected over a 72h-period was analysed for Cyclohexanol (CH), 1,2- and 1,4 -CH-diol (free and conjugated).

Barely 1 % of the absorbed dose (8.46 ± 1.85 mmol) was excreted as CH (1 %), whereas the major metabolite was 1,2-CH-diol (19.1 ± 3.8 %) followed by 1,4-CH-diol (8.4 ± 1.4 %). The excretion of CH in urine peaked at the end of exposure; thereafter, CH decayed rapidly with the half-life being estimated as 1.5 h reflecting a rapid clearance of CH which to a large extent is due to its rapid oxidization to CH-diols. The excretion curves of 1,2- and 1,4-CH-diol reached the maximum at a few hours post exposure with elimination half-lives being 14.3 ± 1.2 h and 18 ± 2.5 h, respectively.

The rate-limiting step in the elimination of CH (14-18 h) thus appears to be associated with the elimination, rather than formation, of CH-diols. Whereas the 1,2 CH-diol metabolite is excreted conjugated to glucoronic acid, the 1,4 CH-diol conjugate is eliminated freely.