Registration Dossier

Administrative data

Description of key information

Acute oral toxicity (based on read across from Iso cyclo geraniol): LD50 > 2000 mg/kg bw.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records
acute toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Results derived from a valid read across, with adequate and reliable documentation / justification.
Justification for type of information:
The read across justification is presented in the Acute Toxicity Endpoint summary. The corresponding documentation file is also attached there.
Reason / purpose:
read-across source
Key result
Dose descriptor:
Effect level:
> 2 000 mg/kg bw
Based on:
other: read across information
Remarks on result:
LD50 includes conversion from the analogue towards Citrolate
Interpretation of results:
other: Not classified, criteria not met
according to EU CLP Regulation (EC) No. 1272/2008 and its amendments.
The acute oral LD50 of Citrolate is considered to be 2600 mg/kg bw, based on read across information from Floralol. Based on this result Citrolate should be classified for acute oral toxicity, Category 5 according to GHS.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The acute oral toxicity result is of sufficient quality and adequate for this dossier.

Additional information

First the experimental information from Iso cyclo geraniol is presented being used for read-across to Citrolate and thereafter the read across justification.

Iso cyclo geraniol and its acute oral toxicity

Acute oral toxicity: In this study, 6 rats (3 males and 3 females) were administered with the substance at dose levels of 2000 mg/kg bw. The rats showed no mortality.Lethargy, flat posture, hunched posture and/or uncoordinated movements were noted among the animals on day 1. Hunched posture was seen in three males on day 2. No effects on bodyweight gain were observed and no abnormalities were found at macroscopic post mortem examination of the animals. Based on the results in this study, the acute oral LD50 for the substance in male and female rats was determined to be >2000 mg/kg bw and the substance is not considered acutely toxic.

The acute oral toxicity of Citrolate (CAS no 67634-26-8) using read across from Iso cyclo geraniol (Cas no68527-77-5).

1. Introduction and hypothesis for the analogue approach

The chemical structure of Citrolate contains a cyclohexene ring with a methyl acetate group at position 1 and methyl groups at positions 2 and 4. For this substance no acute oral toxicity data are available.

In accordance with Article 13 of REACH,lacking information should be generated whenever possible by means other than vertebrate animal tests, i.e. applying alternative methods such as in vitro tests, QSARs, grouping and read-across. For assessing the acute oral toxicity of Citrolate the analogue approach is selected because acute oral toxicity information on one closely related analogue is available.

Hypothesis:Citrolatehas a similar acute oral toxicity profile compared to Iso cyclo geraniol resulting in a similar LD50. Iso cyclo geraniol can be used for read across because the Citrolate-ester will metabolise into its alcohol, which is closely related to Iso cyclo geraniol, and Acetic acid.

Available experimental information: For Iso cyclo geraniol acute oral toxicity in rats is available using the OECD TG 401 method (Kl 1), resulting in an LD50 > 2000 mg/kg bw.The LD50 for Acetic acid (Cas no 64-19-7) is > 2000 mg/kg bw as presented on the ECHA dissemination site.

2. Target chemical and source chemical(s)

Chemical structures of Citrolate and Iso cyclo geraniol are shown in the data matrix, including physico-chemical properties and toxicological information, thought relevant for acute oral toxicity.

3. Purity / Impurities

Citrolate is a multi-constituent. Its two constituents are stereo-isomers and indicate the same acute oral toxicity potential. Impurities of Citrolate are below 10% and are not influencing the acute oral toxicity.

4. Analogue approach justification

According to Annex XI 1.5 read across can be used to replace testing when the similarity can be based on a common backbone and a common functional group. It can also be used when the analogues used will have the same or similar metabolites. When using read across the result derived should be applicable for C&L and/or risk assessment and it should be presented with adequate and reliable documentation.

Analogue selection:Two potential analogues were identified in the OECD QSAR toolbox by using structural similarity with Tanimoto Atom pairs 60% and with structural similarity Tanimoto 60% atom centred fragments. In addition, one isomer was found in RIFM database. These three analogues are esters on which no acute oral toxicity information was available.

In view of Citrolate being an ester, it will be cleaved by carboxylesterases in the gut (and liver) and result in an analogue similar to the alcohol of Citrolate. The alcohol of Citrolate is called Floralol (CAS no 67634-17-7) and this substance has acute oral toxicity information available but the LD50 could not be well derived because only 2 animals of the same sex were used which is not in accordance with OECD TG 401 (see also uncertainty section). Therefore Iso cyclo geraniol is used as an analogue.

Structural similarities and differences:Citrolate andIso cyclo geraniolhave the same backbone: a cyclohexene ring to which methyl groups at positions 2 and 4 are attached. The functional group of Citrolate is a methyl- acetate on position 1, while Iso cyclo geraniol has a methyl alcohol at this position. This latter substance also has an additional methyl group on the ring.

Toxico-kinetics:Absorption: Citrolate and Iso cyclo geraniol have similar absorption potential based on the similarity in chemical structure. Citrolate is an ester and the analogue is an alcohol resulting in some expected differences in physico-chemical properties: the water solubility of Citrolate is somewhat lower and the log Kow being somewhat higher than for Iso cyclo geraniol. For both substances the physico- chemical values indicate readily absorption from the intestinal tract.

Metabolisation: Citrolate will fully metabolise into Floralol and Acetic acid due to activity of carboxylesterases in the gut and liver (Belsito et al., 2008, Yamada et al., 2013 and Wu et al., 2010). This is also presented in the OECD Toolbox rat liver simulator (version Thereafter, Citrolate will follow a similar metabolic pathway as Iso cyclo geraniol. The metabolisation of Citrolate is shown in Figure 1 but instead of presenting Floralol here Iso cyclo geraniol is presented. 

Figure 1: Metabolisation of Citrolate (in the gut and in the liver) into a closely related alcohol (Iso cyclo geraniol) and Acetic acid. The arrow indicates the additional methyl group of Iso cyclo geraniol compared to the direct alcohol of Citrolate being Floralol.


A similar metabolisation scheme is also described by Wu et al, 2010 for terpinyl-propionate (also a para-methyl-cyclohexene ring but with a propyl-ester) who are presenting this as a justified read across. Acetic acid is a natural component in the body, which will be consumed in the Krebs cycle and will therefore not be considered further.

Uncertainty of the prediction:The hydrolysis of esters in the gut to the respective alcohol is well known and referenced (see above) and therefore the closely related Iso cyclo geraniol can be used for read across. Floralol (67634-17-7)being the first metabolite for Citrolate (with one methyl group less) showed an LD50 value between 2000-5000 mg/kg for all animals. The LD50 for females could not correctly be derived because one out of 2 females died (but not the 2 males) at 2000 mg/kg bw and therefore Iso cyclo geraniol is used. Using either information, Iso cyclo geraniol or Floralol, the acute oral toxicity of Citrolate will be > 2000 mg/kg bw after conversion for molecular weight:LD50 would be > 2364 mg/kg bw (2000 mg/kg bw Iso cyclo geraniol / 154 W of Iso cyclo geraniol * 182 MW of Citrolate).

5. Data matrix

The relevant information on physico-chemical properties and toxicological characteristics are presented in the Data matrix at the end of this document.

6. Conclusions per endpoint for C&L and/or risk assessment

When using read across the result derived should be applicable for C&L and/or risk assessment, cover an exposure period duration comparable to the corresponding method and be presented with adequate and reliable documentation.

The acute oral toxicity data of Iso cyclo geraniol can be used for Citrolate because the systemic exposure of Citrolate will be via its alcohol. The LD50 of Iso cyclo geraniol is > 2000 mg/kg bw and therefore the LD50 for Citrolate will be > 2000 mg/kg bw too.

Final conclusion on hazard and C&L

Citrolate has an LD50 > 2000 mg/kg bw and does not need to be classified for acute oral toxicity this will be used for classification and labelling and the risk assessment



Belsito, D., Bickers, D., Bruze, M., Calow, P., Greim, H., Hanifin, J.M., Rogers, A.E., Saurat, J.H., Sipes, I.G., Tagami, H., 2008, A toxicologic and dermatologic assessment of cyclic acetates when used as fragrance ingredients, Food and Chemical Toxicology 46, Suppl 12:S1-27.


Wu, S., Blackburn, K., Amburgery, J., Jaworska, J., and Federle, T., 2010, A framework for using structural, reactivity, metabolic and physico-chemical similarity to evaluate the suitability of analogs for SAR-based toxicological assessments, Regul. Toxicol. Pharmacol., 56, 67-81.


Yamada, T., Tanaka, Y., Hasegawa, R., Sakuratani, Y., Yamada, J., Kamata, E., Ono, A., Hirose., A., Yamazoe, Y., Mekenyan, O., Hayashi, M., 2013, A category approach to predicting the repeated-dose hepatotoxicity of allyl esters, Reg. Toxicol. Pharmacol, 65, 189-195.

Data matrix for Citrolate using read across from Floralol for assessment of acute oral toxicity properties.

Name of substance


2,4-dimethylcyclohex-3-en-1-yl)methyl acetate

Iso cyclo geraniol


Chemical structure

Empirical formula



Cas no.



EC no.



REACH registration

Registered for 2018

Registered for 2018

Mol weight




IFF measured data

IFF measured data




Melting point (oC)



Vapour pressure (Pa)



Water solubility




Log Kow



Human health



Acute oral toxicity

(mg/kg bw)

Read across from Iso cyclo geraniol

LD50 > 2000



Justification for classification or non-classification

Based on the results presented above Citrolate does not need to be classified for acute oral toxicity according to EU CLP Regulation (EC) No. 1272/2008 and its amendments.