Extract obtained from defatted powder of Theobroma cacao (Malvaceae) by extraction with water and ethanol

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Not a guideline study and report does not state whether performed according to GLP. However, data appear well documented and scientifically acceptable.

Data source

Reference

Reference Type:

publication

Title:

Exhaled ethanol and acetaldehyde in human subjects exposed to low levels of ethanol

Author:

Tardif R, Liu L & Raizenne M

Year:

2004

Bibliographic source:

Inhalation Toxicology 16, 203-207

Materials and methods

Objective of study:

other: To determine the concentration of ethanol, and its metabolite acetaldehyde, in the alveolar air of a small number of volunteers exposed (at rest) to short-term low ethanol vapour concentrations.

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Measurement of ethanol, and its metabolite acetaldehyde, in alveolar air samples following exposure by volunteers (at rest) to short-term low ethanol vapour concentrations.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

Ethanol

EC Number:

200-578-6

EC Name:

Ethanol

Cas Number:

64-17-5

Molecular formula:

C2H6O

IUPAC Name:

ethanol

Details on test material:

Ethanol (95%) was used to generate ethanol vapour in the exposure chamber

Radiolabelling:

no

Test animals

Species:

human

Sex:

male/female

Details on test animals or test system and environmental conditions:

Five healthy, non-smoking, Caucasian adults (2 women, 3 men) aged 25-55 years. The volunteers were asked to abstain from consuming alcohol for at least 24 hr before the exposures.

Administration / exposure

Route of administration:

inhalation: vapour

Vehicle:

unchanged (no vehicle)

Details on exposure:

The exposure scheme consisted of three inhalation exposure sessions where the five volunteers were exposed simultaneously for "6 consecutive hours (08:30-14:30)" at rest. During this time subjects "were allowed to leave the inhalation chamber for 5 min after the first 3 h; they had a light lunch between 11:30 and 12:30" [presumably within the exposure chamber]. Exposure sessions were separated by intervals of at least 48 hours.

Duration and frequency of treatment / exposure:

Six consecutive hours of exposure, at each of three exposure concentrations, separated by at least 48 hours.

Doses / concentrations

Remarks:

Doses / Concentrations:
Approximately 26, 102 and 991 ppm

No. of animals per sex per dose / concentration:

2 female and 3 male volunteers per concentration

Control animals:

yes, concurrent no treatment

Positive control reference chemical:

No

Details on dosing and sampling:

Ethanol exposures were carried out in a dynamic 18 m3 chamber, in which purified air was introduced from the ceiling at a rate of 4 m3/min. Humidity (about 40%) and temperature (about 22 deg C) were controlled. Ethanol concentrations were produced by evaporation of liquid ethanol and the dilution of ethanol vapours in purified air. Chamber ethanol concentrations were monitored using flame ionization gas chromatography (at 5 minute intervals) and infrared spectrophotometry (continuously).

Following a period of breath holding and subsequent forced expiration, exhaled alveolar air samples were taken using a three-way valve. The first part of exhaled air was discarded in order to collect only the last portion of air (alveolar air) in the sampling bags. Approximately 2L of alveolar air (end-exhaled) was collected in 3L sample bags. Two samples (one for determining ethanol concentrations and the other for acetaldehyde) were provided by each volunteer at each time point. The time points covered exposure in the chamber (after 2, 4 and 6 hours) and after exposure out of the chamber (6 hr 20 mins and 6 hr 30 mins).

Ethanol concentrations were measured by flame ionization gas chromatography (limit of quantification was set at 2 ppm) within 30 minutes after sampling at the five time points. Acetaldehyde concentrations were assessed, within 30 minutes after sampling at the five time points, following derivatization with 2-hydroxymethylpiperidine and measurement using flame ionization gas chromatography (with a limit of quantification of about 0.025 ppm) .

Results and discussion

Toxicokinetic / pharmacokinetic studies

Details on absorption:

The study indicated that approximately 70 - 80% of inhaled ethanol is absorbed.

Details on excretion:

The concentrations of unchanged ethanol measured in the alveolar air samples taken at 6h increased proportionally from 7.5 ppm to 243 ppm, with increasing exposure concentration (see table 1 for details), and a steady state was observed after 2h of exposure. Following a subsequent 30 minutes outside of the exposure chamber (i.e. sampled at 6h 30 mins), alveolar air ethanol concentrations had dropped from 243 ppm to 24 ppm in those individuals exposed to 991 ppm of ethanol for 6h.

Similarly, the concentrations of acetaldehyde measured in the alveolar air increased proportionally from 0.07 ppm to 1.06 ppm with increasing exposure concentration (see table 2 for details), and a steady state was also observed after 2h of exposure. Following a subsequent 30 minutes outside of the exposure chamber (i.e. sampled at 6h 30 mins), acetaldehyde concentrations had dropped from 1.06 ppm to 0.047 ppm in those individuals exposed to 991 ppm of ethanol for 6 h.

The background level of ethanol measured in 3 subjects not exposed to ethanol was about 4.6 ppm (no further details given on control group)

Metabolite characterisation studies

Metabolites identified:

yes

Details on metabolites:

Acetaldehyde was measured in the expired alveolar air of volunteers inhaling ethanol over 6h (see excretion section above and Table 2 for details).

Any other information on results incl. tables

TABLE 1

Concentrations of unchanged ethanol in alveolar air of volunteers exposed to 3 levels of ethanol during 6 hours

	Ethanol in alveolar air (ppm)
Ethanol in air (ppm)	2 h^a	4 h	6 h	6 h 20 min	6 h 30 min
25.9 + 0.34^b	7.5 + 1.1	7.7 + 0.6	7.5 + 1.3	ND^c	ND
101.7 + 2.1	25.0 + 7.6	23.5 + 1.4	25.32 + 5.7	4.0 + 3.1	ND
990.8 + 10.8	232.5 + 48.5	229.5 + 18.2	243.0 + 20.9	ND	24.3 + 19.6

^aTime of sampling following the start of exposures.

^bMean + SD of concentrations in the inhalation chamber measured at 5-min intervals.

^cND, not determined.

TABLE 2

Concentrations of acetaldehyde in alveolar air of volunteers exposed to 3 levels of ethanol during 6 h

	Acetaldehyde in alveolar air (ppm)
Ethanol in air (ppm)	2 h^a	4 h	6 h	6 h 20 min	6 h 30 min
25.9 + 0.34^b	0.06 + 0.03	0.05 + 0.03	0.07 + 0.01	ND^c	ND
101.7 + 2.1	0.15 + 0.08	0.18 + 0.10	0.20 + 0.08	0.021 + 0.01	ND
990.8 + 10.8	1.11 + 0.23	1.19 + 0.27	1.06 + 0.18	ND	0.047 + 0.01

^aTime of sampling following the start of exposures.

^bMean + SD of concentrations in the inhalation chamber measured at 5-min intervals.

^cND, not determined.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
The present study indicates that about 20-30% of inhaled ethanol following low level exposure is exhaled unchanged in alveolar air, which is indicative that approximately 70-80% of inhaled ethanol is absorbed. Low level inhalation of ethanol resulted in measurable amounts of acetaldehyde in alveolar air. For all exposure concentrations, the results show that the concentration of ethanol and acetaldehyde in exhaled alveloar air increased proportionally, and reached a steady state after at least 2h of continuous exposure. Overall there was a significant correlation between ethanol exposure and ethanol and acetaldehyde concentration in alveolar air. The ratios between acetaldehyde and ethanol in alveolar air after 4h of exposure to 26, 102, or 991 ppm ethanol were 0.005, 0.008, and 0.006, respectively.

Executive summary:

A study was designed to determine the concentration of ethanol, and its metabolite acetaldehyde, in the alveolar air of five volunteers exposed (at rest) to short-term low ethanol vapour concentrations. The volunteers were exposed for 6 hours, on three separate occasions, to approximately 26, 102 or 991 ppm ethanol, and exhaled alveolar air samples were taken for analysis. Low level inhalation of ethanol resulted in measurable amounts of acetaldehyde in alveolar air. The study indicated that approximately 70 - 80% of inhaled ethanol is absorbed.