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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: Study was not run according to guideline; however, it was conducted appropriately. Limited tissues were analyzed; No analytical information was available regarding test substance used.
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

Reference
Reference Type:
publication
Title:
Lack of effect on rat testicular organogenesis after in utero exposure to 3-monochloropropane-1,2-diol (3-MCPD)
Author:
El Ramy R, Elhkim MO, Poul M, Forest MG, Leduque P, and Le Magueresse-Battistoni B
Year:
2006
Bibliographic source:
Reproductive Toxicology 22 (2006) 485-492

Materials and methods

Objective of study:
absorption
distribution
metabolism
toxicokinetics
Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
The levels of 3-MCPD and its main metabolite, beta-chlorolactic acid, were assayed in fetal tissues and dam plasma following a single oral dose of 25 mg/kg BW of 3-MCPD to three pregnant Sprague-Dawley rats per sampling time.
GLP compliance:
not specified

Test material

Constituent 1
Chemical structure
Reference substance name:
3-chloropropane-1,2-diol
EC Number:
202-492-4
EC Name:
3-chloropropane-1,2-diol
Cas Number:
96-24-2
Molecular formula:
C3H7ClO2
IUPAC Name:
3-chloropropane-1,2-diol
Constituent 2
Reference substance name:
3-monochloropropane-1,2-diol
IUPAC Name:
3-monochloropropane-1,2-diol
Details on test material:
- Name of test material (as cited in study report): 3-monochloropropane-1,2-diol (3-MCPD)
- Obtained from Sigma-Aldrich Chemical Co., St. Quentin-Fallavier, France
Radiolabelling:
no

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Elevage Janvier (le Genest, France)
- Age at study initiation: 12 weeks
- Weight at study initiation: no data
- Fasting period before study: no data
- Housing: plastic cages in air conditioned room
- Diet (e.g. ad libitum): commercial food ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 h light, 12 h dark

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: distilled water
Duration and frequency of treatment / exposure:
Single oral exposure on day 14.5 postcoitum
Doses / concentrations
Remarks:
Doses / Concentrations:
25 mg/kg bw
No. of animals per sex per dose / concentration:
3 pregnant females per sampling time
Control animals:
yes, concurrent vehicle
Positive control reference chemical:
The method was validated using plasma supplemented with 2 - 80 ug/g or mL of 3-MCPD or beta-chlorolactic acid.
Details on study design:
- Dose selection rationale: Based on preliminary experiments, doses of 3-MCPD greater than 50 mg/kg induced an excess of maternal and fetal toxicity. Therefore, an exposure of 25 mg/kg BW was chosen for the current study.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: plasma (3 dams per sampling time) and fetal tissue. Dam blood was removed and the plasma was collected by centrifugation (600 × g for 15 min at 4 deg C). The placental/fetal units were removed from the uterus and the fetuses were separated from the placenta, sampled, and weighed. Plasma and tissues were stored at −80 deg C.
- Time and frequency of sampling: 0.5, 1, 2, 3, 5, and 8 hours post-exposure
- Other: To characterize the distribution kinetics of 3-MCPD and beta-chlorolactic acid, the data were estimated by a non-compartmental analysis using WinNonlin 4.01 pharmacokinetic software (Pharsight, Mountain View, CA). This software provided parameters including the maximum concentration (Cmax), the time to reach maximum concentration (Tmax), the area under the concentration–time curve during the period of observation (AUC0−t) and the mean residence time (MRT0−t). The fetus-to-plasma partition coefficient was determined by the ratio AUCfetus,0−t/AUCplasma,0−t. The density of rat plasma was considered equal to 1 g/mL.


METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled: plasma (3 dams per sampling time) and fetal tissue. Dam blood was removed and the plasma was collected by centrifugation (600 × g for 15 min at 4 deg C). The placental/fetal units were removed from the uterus and the fetuses were separated from the placenta, sampled, and weighed. Plasma and tissues were stored at −80 deg C.
- Time and frequency of sampling: 0.5, 1, 2, 3, 5, and 8 hours post-exposure
- From how many animals: 3 animals per sampling time
- Method type(s) for identification: Gas chromatography analysis of 3-MCPD and beta-chlorolactic acid was performed. Approximately 100 mg of fetal tissue or 100 uL of dam plasma was extracted using 800 or 300 uL acetonitrile, respectively. After centrifugation (5 min, 20,000 × g, at 4 deg C), the supernatant was collected and evaporated to dryness at 60 deg C under a gentle stream of nitrogen. One-hundred microlitres of HMDS/TMCS/pyridine (3/1/9) mixture was added to the sample extracts. The mixture was quickly sonicated and heated at 50 deg C for 10 min. After cooling at room temperature for 5 min, 200 uL of hexane and 200 uL of the internal standard (hexadecane, 0.1 mg/mL in hexane) were added, vortexed, and the mixture was centrifuged for 2 min at 20,000 × g at 4 deg C. A 2 uL aliquot of the supernatant was injected into the gas chromatograph for analysis of 3-MCPD and beta-chlorolactic acid. Gas chromatography analysis was carried out on a Hewlet-Packard 5890 equipped with a FID detector and an Ultra-2 (Hewlet–Packard) fused-silica capillary column (25 m, 0.2 mm i.d., 0.33 um film thickness). Ultra-high purity helium was used as the carrier gas at a constant flow rate of 0.9 mL/min. A split–splitless injection system operating in the split mode (3 mL/min) with a quartz 4 mm i.d. deactivated injector liner, packed with silica beads, was used. Operating conditions for gas chromatography were: column temperature 0 min at 80 deg C, 10 deg C/min–200 deg C, 40 deg C/min–280 deg C hold for 5 min, injector temperature 250 deg C and detector temperature 300 deg C. For each run, two replicates of fetal tissues or plasma spiked with 5 and 20 ug/g or mL of 3-MCPD and beta-chlorolactic acid were used. The average retention times for 3-MCPD and beta-chlorolactic acid derivatives were 6.6 and 7.4 min, respectively. The method was validated using untreated rat fetal tissue or plasma supplemented with 2–80 ug/g or mL of 3-MCPD or beta-chlorolactic acid.
- Limits of detection and quantification: For 3-MCPD, the limit of quantification of the method was 2 ug/g.
Statistics:
The data are presented as the mean +/- S.D. Statistical analysis of data was performed by ANOVA, and pairwise comparisons were made using the Dunnett’s test. A p-value of 0.05 or less was determined to be statistically significant.

Results and discussion

Preliminary studies:
No data
Main ADME resultsopen allclose all
Type:
absorption
Results:
3-MCPD and beta-chlorolactic acid were detected in maternal plasma and fetal tissues at all sampling times between 30 min and 5 h after administration
Type:
distribution
Results:
3-MCPD and beta-chlorolactic acid were detected in maternal plasma and fetal tissues at all sampling times between 30 min and 5 h after administration
Type:
metabolism
Results:
Beta-chlorolactic acid was detected in maternal plasma and fetal tissues at all sampling times between 30 min and 5 h after administration of 3-MCPD

Toxicokinetic / pharmacokinetic studies

Details on absorption:
3-MCPD and beta-chlorolactic acid were detected in maternal plasma and fetal tissues at all sampling times between 30 min and 5 h after administration (see table below). However, 5 h after administration, the plasma and tissue levels of 3-MCPD were under the LOQ (2 ug/g).
Details on distribution in tissues:
3-MCPD and beta-chlorolactic acid were detected in maternal plasma and fetal tissues at all sampling times between 30 min and 5 h after administration (see table below). However, 5 h after administration, the plasma and tissue levels of 3-MCPD were under the LOQ (2 ug/g).
Transfer into organs
Test no.:
#1
Transfer type:
blood/placenta barrier
Observation:
distinct transfer
Details on excretion:
Not measured.
Toxicokinetic parametersopen allclose all
Test no.:
#1
Toxicokinetic parameters:
Cmax: 16.4 ug/g in dam plasma at 30 min after administration. (3-MCPD)
Test no.:
#1
Toxicokinetic parameters:
Cmax: 14.3 ug/g in fetal tissue at 1 h after administration. (3-MCPD)
Test no.:
#1
Toxicokinetic parameters:
Cmax: 10.0 ug/g in dam plasma at 1 h after administration. (beta-chlorolactic acid)
Test no.:
#1
Toxicokinetic parameters:
Cmax: 9.5 ug/g in fetal tissue at 2 h after administration. (beta-chlorolactic acid)
Test no.:
#1
Toxicokinetic parameters:
other: Mean partition coefficients were 0.93 and 1.08 for 3-MCPD and beta-chlorolactic acid, respectively.
Test no.:
#1
Toxicokinetic parameters:
other: MRT0-3h of 3-MCPD was 1.26 h in plasma and 1.31 h in fetus.
Test no.:
#1
Toxicokinetic parameters:
other: MRT0-5h of beta-chlorolactic acid was 2.06 h in plasma and 2.32 h in fetus.

Metabolite characterisation studies

Metabolites identified:
yes
Details on metabolites:
Beta-chlorolactic acid was detected in maternal plasma and fetal tissues at all sampling times between 30 min and 5 h after administration of 3-MCPD (see table below).

Any other information on results incl. tables

3-MCPD and Beta-chlorolactic acid concentrations in fetal tissues and dam plasma (mean +/- S.D.; LOQ = 2 ug/g or mL)

3-MCPD

Beta-chlorolactic acid

 

 

Sampling time (h)

Fetus (ug/g)

Dam plasma (ug/mL)

Fetus (ug/g)

Dam plasma (ug/mL)

0.5

13.2 +/- 1.2

16.5 +/- 0.6

5.1 +/- 0.9

7.5 +/- 1.2

1

14.3 +/- 2.7

15.1 +/- 1.6

8.8 +/- 1.2

10.0 +/- 0.3

2

7.8 +/- 1.2

7.5 +/- 0.1

9.6 +/- 1.9

8.5 +/- 0.7

3

4.0 +/- 1.0

4.4 +/- 0.8

7.7 +/- 1.8

5.8 +/- 0.9

5

< LOQ

< LOQ

3.3 +/- 0.5

2.1 +/- 0.4

8

< LOQ

< LOQ

< LOQ

< LOQ

     

 

 

Applicant's summary and conclusion

Conclusions:
3-MCPD and beta-chlorolactic acid (the main metabolite of 3-MCPD) were found to readily cross the placental barrier and diffuse throughout the fetal tissues.