2-methoxy-4-propylphenol

Endpoint summary

Link to relevant study record(s)

Reference

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

disregarded due to major methodological deficiencies

Study period:

1983

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

significant methodological deficiencies

Remarks:

This study precedes guideline setting. It is well documented but some methodological aspects make it not reliable: the low temperature (21°C) used in this experiment, the fact that amount within the epidermis was not determined, and the potential loss of test substance through its determination by extraction process rather than radiolabelling may have contributed to the very low penetration rate reported. The integrity of the skin was not checked but in any case a loss of integrity would overestimate the penetration rate.It is therefore considered that the absolute penetration level may not be accurate.

Qualifier:

no guideline followed

Principles of method if other than guideline:

In an in vitro skin absorption study, penetration of the test material through excised human epidermis was examined.

GLP compliance:

not specified

Radiolabelling:

no

Species:

other: Human cadaver skin

Doses:

0.2 mL

Details on in vitro test system (if applicable):

SKIN PREPARATION
- Source of skin: Human cadaver
- Ethical approval if human skin: No data
- Preparative technique: Human lower abdominal skin was excised from a cadaver during autopsy and kept at -20 °C and thawed prior to examination. Subcutaneous tissue was removed using Cooper's scissors. Epidermis was separated from dermis by a modification of Baumberger's method (1942). Full thickness skin was placed dermis side down on a metal plate heated to 60 °C for a period of 10 minutes. The epidermis was separated from the dermis using forceps.
- Storage conditions: -20 °C

APPLICATION OF TEST MATERIAL.
- The glass chamber used in this study is described as follows: the upper surface of the epidermis was fixed to the lowest part of a glass tube (a), using adhesive. The glass tube (a) was then placed inside one arm of the U-shaped glass-chamber. Another glass tube (b) was withdrawn from the other side of the glass-chamber, and approximately 5 mL of saline was poured into the chamber until it came into complete contact with the bottom of the epidermis. The glass tube (b) was then replaced into the chamber.
- 0.2 mL of test material was applied to the top of the epidermis attached to the glass tube (Ea) by using a micropipette.
- The top of the glass tube (Ea) was covered with parafilm in order to avoid evaporation of the test sample.
- The chamber was kept in a thermostatically controlled cabinet (HAM-40 type, Seiwa Riko Company) at 21 °C and 55 % relative humidity for 72 h.

MEASUREMENT OF PENETRATION
- Seventy-two hours after application of test material, the glass tube (a) was removed. The saline from the chamber was poured into a test tube. The chamber and the bottom of the epidermis attached to the glass tube (a) were both washed 3 times with saline, poured into the same test tubethe measurement of penetration was performed. This saline (approximately 10 mL) was then poured into a 100mL flask.
- 10 mL of saturated salt water and 25 mL of ether were added and mixed vigorously.
- The compound was extracted in ether and. A An additional 25 mL of ether was added to the water fraction remaining after extraction, in order to insure complete extraction.
- The resultant 50 mL of ether, after the two extractions, was dehydrated by adding approximately 2 g of anhydrous Na2SO4. In order to remove the anhydrous Na2SO4, the ether fraction was filtered with filter paper and then condensed to 1 mL of ether by using a Kderna-Danish condenser.
- Then 2 µL of the condensed sample was injected into a Shimazu GC-6A gas chromatograph under following specifications:Detector: Flame ionization detector Column: 10 % FEAP on Gasport H60-80 mesh 2 mm 1.0 x 2.0 m (glass) Column temperature: 150-250 °C Injection temperature: 280 °C Detector temperature: 280 °CSensitivity: 10^3 x 32 – 10^4 x 16 mVΩ Carrier gas: N2 at 30 mL/minute
- The peak area on the gas chromatograms was compared with that of a standard sample, in which the concentration of the material tested was known.
- The amount of the test material which penetrated the human epidermis was calculated by the following formula:Amount of test material which penetrated human epidermis (µg) = (Peak area of test sample/ peak area of standard sample) x (concentration of test material in standard sample (µg/mL)) x (volume of test sample (mL))
- Percentage penetration was calculated using following formula:Percentage penetration = [Amount penetration (µg) / (200 µL x specific gravity x 10^3)] x 100 - The experiment was repeated 6 times.

Dose:

0.2 mL

Parameter:

percentage

Absorption:

1.364 %

Remarks on result:

other: 72 h

Remarks:

penetration through excised human epidermis

Conversion factor human vs. animal skin:

Not applicable

Percentage penetration:

Penetration of 0.2 mL of test material through excised human epidermis after 72 h was found to be 1.364 ± 0.109 %.

 

Solubility in water and octanol:

Test material was freely soluble in octanol. Test material exhibited very slight solubility in water (5 to 10 ppm).

A high correlation between water solubility and percentage penetration was observed (r = 0.900, t = 12.040).

Conclusions:

Under the test condition, penetration of test material through excised human epidermis after 72 h was found to be 1.364 ± 0.109 %. Also, high correlation between water solubility and percentage penetration through human epidermis of test material was observed. However due to significant methodological deficiencies, it is considered that the absolute penetration level may not be accurate.

Executive summary:

In an in vitro skin absorption study, penetration of test material through human epidermis was examined.  0.2 mL of test material was applied to the epidermis attached to the glass tube in a thermostatically controlled cabinet at 21 °C and 55 % relative humidity for 72 h. Seventy-two hours after application of test material, the measurement of penetration was performed by gas chromatography. the solubility of test material in water and octanol was also studied.

Penetration of test material through excised human epidermis after 72 h was found to be 1.364 ± 0.109%. Test material was freely soluble in octanol. Test material exhibited very slight solubility in water (5-10 ppm). A high correlation between water solubility and percentage penetration was observed.

This study precedes guideline setting. It is well documented but some methodological aspects make it not reliable: the low temperature (21°C) used in this experiment, the fact that amount within the epidermis was not determined, and the potential loss of test substance through its determination by extraction process rather than radiolabelling may have contributed to the very low penetration rate reported. The integrity of the skin was not checked but in any case a loss of integrity would overestimate the penetration rate.It is therefore considered that the absolute penetration level may not be accurate.

Description of key information

The available evidence suggests that the substance is bioavailable via the oral and dermal route. Systemic absorption of this substance via inhalation route is expected but to a limited extent. The substance is expected to be mainly excreted in urine.

Key value for chemical safety assessment

Bioaccumulation potential:

low bioaccumulation potential

Additional information

In accordance with the section 8.1.1 of Annex VIII of Regulation (EC) No 1907/2006 (REACH), the toxicokinetic profile of the substance (i.e. absorption, distribution, metabolism and elimination) was derived from the relevant available information collated in the dossier. The physical chemical characteristics of the substance, the results obtained from acute, repeated-dose, and reproductive toxicity studies on the substance or on related substances (see Section 13 for read-across justification) were used to predict its toxicokinetic behaviour.

The JECFA evaluation (JECFA, 2004) was also used to support this toxicokinetic assessment.

Physical-chemical properties:

The substance is a mono-constituent, having a relatively low molecular weight (MW) of 166.22 g/mol. The substance is water soluble liquid (1.1 g/L) and is moderately lipophilic based on the octanol/water partition coefficient (Log Kow = 2.8). The substance has low volatility according to its vapour pressure (3 Pa at 25°C).

Absorption:

Oral/GI absorption

The physical chemical characteristics described above suggest that the substance is absorbed in the gastro-intestinal tract by passive diffusion.Water-soluble substances will readily dissolve into the gastrointestinal fluids. Absorption of very hydrophilic substances by passive diffusion may be limited by the rate at which the substance partitions out of the gastrointestinal fluid. However, since the molecular weight is low (less than 200), the substance may pass through aqueous pores or be carried through the epithelial barrier by the bulk passage of water. This hypothesis is supported by the systemic effects in the oral toxicity data, as summarized below:

- In an acute oral gavage toxicity study, mortality was observed at high dose levels (≥ 1730 mg/kg bw; LD50 = 2600 mg/kg bw).

- The 90-day repeated dose toxicity performed with the source substance (trans-Isoeugenol, see Section 13 for read-across justification) gave a NOAEL of 300 mg/kg bw/day based on increased kidney weights in females (eq. to 304 mg/kg bw/day for the target substance after MW correction).

- In the 2-year carcinogenicity study performed with the source substance (trans-Isoeugenol, see Section 13 for read-across justification), a NOAEL of 150 mg/kg bw was determined for non-neoplastic effects, based on the adverse effects on the kidney in the female mice (eq. to 152 mg/kg bw/day for the target substance after MW correction). The renal neoplastic-like effects were considered equivocal.

- In the two-generation study performed with the source substance (trans-Isoeugenol, see Section 13 for read-across justification), a NOAEL for fertility higher than 700 mg/kg bw/day was determined, i.e. the highest concentration tested in the study (eq. to 709 mg/kg bw/day for the target substance after MW correction). A NOAEL development higher than 230 mg/kg bw/day and a LOAEL of 700 mg/kg bw/day were determined (eq. to 233 and 709 mg/kg bw/day for the target substance after MW correction), as a worst-case, based on decreased male and female F2 pup weights.

Taken together, the observation of systemic effects indicates the oral bioavailability of the substance and/or its metabolites. In light of these data, and the lack of specific information on oral absorption, the substance was assumed to be 100% bioavailable by oral route for the purposes of human health risk assessment.

Dermal absorption

Regarding dermal absorption, systemic absorption by the dermal route is expected to be moderate to high based on the Log Kow and the water solubility values. Only few systemic effects were observed in the acute dermal toxicity study. This may be explained by a combination of a low toxicity and the absence of dermal absorption.

An in vitro skin absorption study indicated that the percutaneous absorption level of the substance was not significant (Jimbo, 1983). Following 72 hours exposure, 1.364 ± 0.109 % of the applied dose (mean ± standard error) had permeated into the receptor phase. However this result is not considered as accurate due to the poor reliability of this study (Klimisch score = 3) which is strengthened by the different dermal absorption percentage reported by Shen et al. (2014) for other substances mentioned in Jimbo’s publication:

Isoeugenol 0.489 ± 0.029 % vs 38.4%;

Methyl eugenol 0.511 ± 0.038 % vs 49.7%;

Dihydroeugenol 1.364 ± 0.109 % vs 22.6%.

In light of these data, the substance was conservatively assumed to be 100% bioavailable by dermal route for the purposes of human health risk assessment.

Respiratory absorption

The potential for inhalation toxicity was not evaluated in vivo.

The vapour pressure of the substance (Vp = 3 Pa at 25°C) indicated a low volatility and inhalability and therefore no exposure by inhalation is anticipated. Thus, at ambient temperature, no respiratory absorption is expected under normal use and handling of the substance.

However, when used as a vapour in aerosol, the substance is expected to be directly absorbed across the respiratory tract epithelium by passive diffusion.

In light of these data, and the lack of specific information on respiratory absorption, the substance was conservatively assumed to be 100% bioavailable by inhalation for the purposes of human health risk assessment.

Distribution:

Any material that is absorbed will be distributed via the blood to the liver, and other organs and tissues. The water solubility of the substance would allow 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 metabolization, the substance as such would not accumulate in the body fat.

Metabolism:

As reported in the JECFA assessment, Isoeugenol derivatives containing a phenolic OH group are rapidly absorbed from the gastrointestinal tract and are metabolized principally in the liver via conjugation of the phenolic hydroxy group with sulfate or glucuronic acid. The conjugates are subsequently excreted, primarily in the urine.

Excretion:

The registered substance, having a molecular weight lower than 300 g/mol, is expected to be mainly excreted in urine and no more than 5-10% may be excreted in bile. Any substance that is not absorbed from the gastro-intestinal tract, following oral ingestion, will be excreted in the faeces.

 

References:

- Fischer IU, von Unruh GE, Dengler HJ. The metabolism of eugenol in man. Xenobiotica. 1990 Feb;20(2):209-22.

- JECFA (2004). Safety evaluation of certain food additives and contaminants. Prepared by the Sixty-first meeting of the Joint FAO/WHO Expert Committee on Food Additives (JECFA). WHO FOOD ADDITIVES SERIES: 52.

- Shen Lie, Kromidas Lambros, Schultz Terry and Bhatia Sneha. An in silico skin absorption model for fragrance materials. Food and Chemical Toxicology 74 (2014) 164 -176.