Bis(2-chloroethyl) ether

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

distribution

Qualifier:

no guideline followed

Principles of method if other than guideline:

- Principle of test: Inhalation exposure to radiolabelled DCEE in a closed all-glass exposure system
- Short description of test conditions: A single dose corresponding to 0.25 mCi/animal (final uptake) was given to three rats. The rats were sacrificed after 24h.
- Parameters analysed / observed: distribution in organs

GLP compliance:

no

Remarks:

pre-GLP

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Name (as cited): 2,2'-Dichlorodiethylether (BCEE)
- Purity: 99%
- Source of test material: Merck-Schuchard

RADIOLABELLED TEST MATERIAL
- Name (as cited): [1-14C]2,2'-Dichlorodiethylether
- Radiochemical purity: no specified
- Source of test material: NEN Chemicals
- Specific activity: 1.42 mCi/mmol

Radiolabelling:

yes

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Ivanovas, Kissleg, FRG
- Age at study initiation: not specified
- Weight at study initiation: 200g
- Diet: ad libitum
- Water: ad libitum

Route of administration:

inhalation: vapour

Vehicle:

unchanged (no vehicle)

Duration and frequency of treatment / exposure:

At the start of the experiment the glass chamber, where the three rats were housed, was filled with vapor of radiolabelled BCEE. After 18 h exposure > 95% of the administered BCEE was taken up by the animals. Uptake was 0.25 mCi/animal.

Dose / conc.:

125.9 mg/kg bw (total dose)

Remarks:

Calculated based on the specific activity of radiolabelled BCEE (1.42 mCi/mmol), the uptake of test animals (0.25 mCi/animal) and the animal weight (200 g).

No. of animals per sex per dose / concentration:

3 animals

Control animals:

no

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY
- Test chamber: closed all-glass exposure system
- Dose: 0.25 mCi/animal
- Time of sampling: 24 h post-exposure
- Tissues sampled: Liver, kidney, spleen, lung, small intestine and muscle tissues
- Sample storage: - 80°C
- Parameter measured: irreversible binding of radioactive metabolites of [1-14C]BCEE to tissue proteins
- Other: The decline of BCEE vapor in the inhalation chamber was measured by gas chromatography. After 18 h exposure >95% of the administered BCEE was taken up by the animals.

Type:

distribution

Results:

Irreversibly protein bound radiolabelled BCEE was mainly found in the liver. Smaller amounts were found in kidney, small intetsine, lung and spleen.

The relative distribution of irreversible protein binding in different rat tissues after exposure of the animals to [¹⁴C]BCEE as observed in this study is presented in the table below:

Tissue	% of radioactivity irreversibly bound per g tissue protein (n = 4)
Liver	0.32 +/- 0.016
Lung	0.07 +/- 0.015
Spleen	0.06 +/- 0.014
Kidney	0.17 +/- 0.024
Small intestine	0.12 +/- 0.017
Muscle	0.01 +/- 0.003

Conclusions:

3 male rats were exposed to radiolabelled BCCE by inhalation (dose: approc. 125.9 mg/kg BCEE). After 18 h exposure > 95% of the administered BCEE was taken up by the animals. Radiolabbed BCEE irreversibly bound to protein was mainly found in the rat liver. Smaller amounts were found in kidney, small intestine, lung and spleen.

Executive summary:

The objective of this study was to evaluate the role of vinyl chloride metabolites (namely chloroethylene oxyde (CEO) an chloroacetaldehyde (CAA)) in the carcinogenicity of vinyl chloride (VC). 2,2' dichlorodiethyl ether (BCEE) was studied as a metabolite precusor of CAA. Following exposure to vapour of radiolabelled VC and BCEE, the distribution of radioactiviy in various tissues following covalent binding to protein was studied. The carcinogenic potential of exposure to these substance was evaluated by the occurrence of nucleic acid (RNA and DNA) alkylation and the potency of the two chemicals to induce preneoplastic ATPase-deficient foci in rat liver.

After 18 h exposure > 95% of the administered BCEE was taken up by the animals. Both chemicals displayed similar distribution in rat tissues. Radiolabbed BCEE irreversibly bound to protein was mainly found in the rat liver. Smaller amounts were found in kidney, small intestine, lung and spleen.

Reference 2

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Objective of study:

excretion

metabolism

Qualifier:

equivalent or similar to guideline

Guideline:

EU Method B.36 (Toxicokinetics)

GLP compliance:

no

Remarks:

pre-GLP

Specific details on test material used for the study:

TEST MATERIAL:
- Names (as cited): Bis(2-chloroethyl)ether (BCEE)
- Source of test material: Eastman No. 886 - Commercial grade
- Purity: 98.5% (checked by GC-FID and GC/MS)

Radiolabelling:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Inc.
- Weight at study initiation: 280-330g

Route of administration:

oral: gavage

Vehicle:

corn oil

Duration and frequency of treatment / exposure:

Single dose

Dose / conc.:

40 mg/kg bw (total dose)

Remarks:

single dose

No. of animals per sex per dose / concentration:

This study was performed on male rat only at a single dose. 7 rats were used.

Control animals:

no

Details on study design:

Immediately following administration of the dose, animals were placed in Delmar-Roth metabolism cages equipped with all glass urine-feces separators. The urine and feces traps were immersed in an ice bath.
Laboratory air was drawn by vacuum through the metabolism cages at a rate of 400 ml/min. The air was passed through a charcoal trap before entering
the cage. Unaltered BCEE from the expired air was collected with a trap consisting of a 500-ml threenecked round-bottom flask fitted with a Liebig-type
condenser (bottom) and a Graham-type condenser (top) in series. The condensers were cooled with recirculating water at 5°C.
In pre-trials, the efficiency of the system was tested. Recovery reached 95.5+/-1.5% over a 8hr period and 93.8+/-2.8% over a 36 hr period.

Details on dosing and sampling:

- Dosing:
A single dosage at 40 mg/kg bw was administered by gavage; BCEE was administered with corn oil as a vehicle.

- Air samples:
Two samples were taken from the expired air trap. the first sample corresponded to the air expelled in the frst 8 hr following dosage. The second sample was obtained at 48 hr representing collection between 8 and 48 hr.

- Urine samples:
Urine samples were collected at 4, 8, 20, 32, and 48 hr and were stored at -60°C until analysis by GC/MS

Metabolites identified:

yes

Details on metabolites:

Two metabolites were identified:
- Thioglycolic acid (TGDA)
- 2-chloroethyl β-D-glucosiduronic acid (CEGA)

- Analysis of expelled air:

Unaltered BCEE was below the limit of quantification in 0-8 hr air sample. If the minimum amount allowing determination by the GC analysis is considered as the upper limit of BCEE expired unaltered following dosing, no more than 1.6% of the dose was expired unaltered over the first 8 hr post exposure. No BCEE was identified in the air sample taken at 48 hr.

- Analysis of urine samples:

Thioglycolic acid (TGDA) was identified as the main metabolite in the rat urine. Quantitative analysis by GC gave an average recovery for 48 hr of 33.0 +/-11.8 mg TGDA/kg following a single oral dose of 40 mg/kg BCEE. 2-chloroethyl β-D-glucosiduronic acid (CEGA) was identified as another metabolite of BCEE.

Conclusions:

Following a single peroral administration of 40 mg/kg BCEE, less than 2 % of the coupound is expired unaltered. After 48h, the compund appears to be mainly excreted through the urine as TGDA. This suggests that BCEE is rapidely absored in the body and excreted as the urinary metabolite TDGA.

Executive summary:

In this published study, seven male rats were given a single oral dose (40 mg/kg) of bis(2-chloroethyl)ether (BCEE). Less than 2 % of the dose was recovered from the expired air as the unaltered parent compound during an 8 hr collection period (no BCEE was detected after 48 hr).

Urine samples representing 48 hr of collection were analyzed by GC/MS after derivitization using methyl ester/trimethylsilyl ether. Two metabolites were identified in this fraction : thiodiglycolic acid (TDGA) and 2-chloroethyl β-D-glucosiduronic acid (CEGA).

Quantitative analysis for TDGA gave an average (seven rats) yield for 48 hr of 33 +/-11.8 mg TDGA/kg from a single dose of 40 mg/kg BCEE. This suggests that BCEE was absored in the body and excreted as the urinary metabolite TDGA. However, as BCEE was not radiolabeled and the exact stoichiometric relationship between BCEE and TGDA is unknown, the proportion of the parent compound converted to TDGA could not be evaluated.

Reference 3

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Objective of study:

distribution

excretion

metabolism

Qualifier:

equivalent or similar to guideline

Guideline:

EU Method B.36 (Toxicokinetics)

GLP compliance:

no

Remarks:

pre-GLP

Specific details on test material used for the study:

TEST MATERIAL:
- Names (as cited): Bis(2-chloroethyl)ether (BCIE)
- Purity: 98.5% (determined by GC/MS)
- Impurity: 1,1,2,2-tetrachloroethane (tentative identification)
- Source of test material: Eastman Organic Chemicals

RADIOLABELLED TEST SUBSTANCE:
- Source of test material: New England Nuclear
- Radiochemical purity: > 98.5% (supplier information)
- Specific activity: 1.34 mCi/mmole
- Locations of the label: Bis(1-14C-2-chloroethyl)ethe

Radiolabelling:

yes

Species:

rat

Strain:

CD-1

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Inc.
- Age at study initiation:
- Weight at study initiation: 180-240 g
- Housing: fasted 24 hr prior to dosing
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: not specified

Route of administration:

oral: gavage

Vehicle:

corn oil

Duration and frequency of treatment / exposure:

Single dose

Dose / conc.:

40 mg/kg bw (total dose)

Remarks:

Single dose equivalent to 1-1.4 µCi-14C

No. of animals per sex per dose / concentration:

This study was performed on male rat only at a single dose. 7 rats were used.

Control animals:

no

Details on study design:

Rat were housed individually in all glass metabolism cages fitted with urine-feces separator traps packed in ice for the course of an experiment. A charcoal trap was placed at the inlet of the metabolism cages and a glass impinger trap containing 100 ml of 5N ethanolamine in methoxyethanol (EA-ME) was placed in series with the outlet. Room air was drawn through the system at a rate of 400 ml/min.

No provision was made for separating and trapping unchanged chloroether, because previous experiments have shown that less than 2% of an administered dose of either compound is excreted unchanged.

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine, faeces, blood, organs
- Time and frequency of sampling: 4, 8, 20, 32 and 48 hr after dosing (except organs which were collected at the end of the experiment)

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled (delete / add / specify): urine
- Time and frequency of sampling: 4, 8, 20, 32 and 48 hr after dosing
- From how many animals: 7
- Method type for identification: GC-MS

Details on distribution in tissues:

see section "Any other information on results incl. tables"

Details on excretion:

The elimination half-life, corresponding to the time where the sum of urinary radiolabelled carbon and expired radiolabbeled CO2 equals 50% of the administered dose, was 12 hours.

Key result

Toxicokinetic parameters:

half-life 1st: 12 hr

Remarks:

excretion

Metabolites identified:

yes

Remarks:

urinary metabolites

Details on metabolites:

3 metabolites identified by GC-MS:
- Thiodiglycolic acid (TDGA) identified from an auhtentic standard
- 2-chloroethoxyacetic acid (CEAA) deduced from the mass spectrum
- N-acetyl-S-[2-(2-chloroethoxy)ethyl]-L-cysteine (ACEEC) deduced from the mass spectrum

** Recovery of administered ¹⁴C activity recorder over 48 hr following single peroral doses of bis(1-¹⁴C-2-chloroethyl)ether (40 mg/kg, 7 rats):

Expired as CO2 (%)	Urine (%)	Feces (%)	Organs and tissues (%)	Cage wash (%)	Total recovery (%)
11.5 +/- 5.6	64.7 +/- 14.8	2.4 +/- 1.3	2.3 +/- 1.2	< 0.5	80.9 +/- 16.9

** Percentage of administered ¹⁴C activity recorder in blood and selected organs of the rats 48 hr after the administration of a single peroral dose of bis(1-¹⁴C-2-chloroethyl)ether (40 mg/kg, 7 rats). Blood, muscle, liver and kidney were the only organs that had ¹⁴C levels exceeding 0.05% of the dose at 48 hr:

Blood (% of dose)	Liver (% of dose)	Kidney  (% of dose)	Muscle  (% of dose)
0.49 +/- 1.28	0.19 +/- 0.13	0.56 +/- 0.072	0.963 ++- 1.49

Conclusions:

The excretion half-life of [1-14C]BCEE was 12 hr. Expired 14CO2 accounted for 11.5 +/- 5.6(SD) % of the dose, urinary 14C accounted for 64.7 +/- 14.8%, and 2.4 +/- 1.3% was found in the faeces. BCEE is thus befficiently metabolised and rapidely excreted. Thiodiglycolic acid (TDGA) was found to be the main (75%) urinary metabolite of BCEE. Other urinary metabolites included 2-chloroethoxyacetic acid (CEAA) (5%) and N-acetyl-S-[2-(2-chloroethoxy)ethyl]-L-cysteine (ACEEC) (7%).

Executive summary:

In this study, seven male rats were given single peroral doses of bis(1-¹⁴C-2-chloroethyl)ether ([1-¹⁴C]BCEE) (40 mg/kg). Excretion of ¹⁴CO₂ and urinary ¹⁴C was followed for 48 hr. The excretion half-life of [1-¹⁴C]BCEE was 12 hr. Expired ¹⁴CO₂ accounted for 11.5 +/- 5.6(SD) % of the dose, urinary ¹⁴C accounted for 64.7 +/- 14.8%, and 2.4 +/- 1.3% was found in the faeces.

Thiodiglycolic acid (TDGA) accounted for roughly 75% of the total urinary ¹⁴C collected after dosing, corresponding to 48% of administered [1-¹⁴C]BCEE. This shows that major portion of a single peroral dose of BCEE undergoes cleavage at the ether linkage to give thiodiglycolic acid (TDGA) as a final urinary metabolite. A highly reactive intermediate product from cleavage of BCEE is suggested by the fact that no unconjugated cleavage products were found in the urine. Muller et al. (1979) have reported that the reactive intermediate metabolite of BCEE is the same as that of VC. The fact that BCEE and VC, a known hepatocarcinogen, are converted to a common reactive intermediate supports the bioassay evidence which suggests that BCEE is a carcinogen.

Lesser metabolites of BCEE were 2-chloroethoxyacetic acid (CEAA) (5%) produced by oxidation of BCEE, and N-acetyl-S-[2-(2-chloroethoxy)ethyl]-L-cysteine (ACEEC) (7%) produced by direct substitution. Two compounds which have been previously identified as urinary metabolites of BCEE were not identified in this work: S-carboxymethyl-L-cysteine (CMC) and 2-chloroethyl-beta-D-glucopyrano-siduronic acid (CEG). CMC is thought to be an intermediate in the pathways leadin to TGDA. The authors suggest that CMC was not identified due to the low dosage used in this study (40 mg/kg bw). CEG was not quantitated in previous studies and was not found in any of the urine samples collected in this study. It is not considered as a major metabolite.

Reference 4

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

metabolism

Qualifier:

no guideline followed

Principles of method if other than guideline:

- Principle of test: GC-MS investigation of urinary metabolites following intraperitoneal administration of Bis(chloroethyl) ether to rats

GLP compliance:

no

Remarks:

pre-GLP

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source of test material: E. Merck AG, Darmstadt (F.R.G.)
- Purity: 99%

Radiolabelling:

no

Species:

rat

Strain:

other: SPF Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: AF/Han Strata of the Zentralinstitut fiir Versuchstierzucht, Hannover-Linden
- Age at study initiation: not specified
- Weight at study initiation: 200 g

Route of administration:

intraperitoneal

Vehicle:

unchanged (no vehicle)

Dose / conc.:

100 mg/kg bw (total dose)

Remarks:

Single dose

No. of animals per sex per dose / concentration:

Not specified

Control animals:

not specified

Details on dosing and sampling:

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled: urine
- Time and frequency of sampling: 24h period following administration
- Method type(s) for identification: factionation by ion-echange column chromatography, identification using GC-MS

Metabolites identified:

yes

Remarks:

urinary metabolites

Conclusions:

Following the administration of a single intraperitoneal dose of Bis(2-chloroethyl) ether (BCEE) at 100 mg/kg bw, dimethylester of thiodiglycollic acid and the N-trifluoro-acetyl-n-butylester of S-(carboxymethyl)-L-cysteine were identified in the urine of treated rats. The authors alos suggest that the breakdown of the compound may be initiated in vivo by an enzyme-catalyzed step and/or gastric acid hydrolysis, leading to the production of chloroacetaldehyde, a known mutagenic agent.

Executive summary:

In this published study, male rats (number of animals not specified) were given a single intraperitoneal dose of Bis(2-chloroethyl) ether (BCEE) at 100 mg/kg bw. The objective of this study was to better understand the role of BCEE metabolites in the carcinogenicity and mutagenicity of the substance.

Dimethylester of thiodiglycollic acid and the N-trifluoro-acetyl-n-butylester of S-(carboxymethyl)-L-cysteine were identified in the urine of treated rats. Metabolites were identified based on their mass fragmentation pattern as compared to authentic standard substances.

From previous studies, the authors found that BCEE has a much weaker alkylating activity than α-halo ethers, which suggest that the breakdown of the compound may be initiated in vivo by an enzyme-catalyzed step. Gastric acid hydrolysis could also induce O-dealkylation leading to generating chlorothanol. Chloroethanol could then be transformed in chloroacetaldehyde, a known mutagenic agent.

Reference 5

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

excretion

Qualifier:

no guideline followed

GLP compliance:

no

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and of test material: Merck Co. (D-6100 Darmstadt, FRG)
- Purity: 99%

Radiolabelling:

no

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Zentralinstitut für Versuehstiere (D-3000 Hannover, FRG)
- Strain: strain/AF/Han
- Age at study initiation: not specified
- Weight at study initiation: 200-250g
- Housing: Makrolon cages
- Diet: ad libitum
- Water: ad libitum

Route of administration:

intraperitoneal

Vehicle:

other: Mazola oil

Dose / conc.:

5.7 mg/kg bw (total dose)

Remarks:

(i.e. 40 µmole/kg bw)

Dose / conc.:

22.9 mg/kg bw (total dose)

Remarks:

(i.e. 160 µmole/kg bw)

No. of animals per sex per dose / concentration:

5

Control animals:

not specified

Details on study design:

Metabolism of 2,2'-DDE, chloracetaldehyde and chloroethanol was studied in male Wistar rats after i.p. injections of the compounds. Each compound was given to a group of 5 rats at the dosages of 40 and 160 p.mole (in Mazola oil)/kg body weight. Each animal was housed in an individual metabolic cage and 24-h urine samples collected.

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine
- Time and frequency of sampling: collected for 24 h

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled: urine
- Time and frequency of sampling: collected for 24 h
- From how many animals: 5/group
- Method type for identification: GC-FID

Metabolites identified:

yes

Details on metabolites:

- TdGA: thiodiglycollic acid
- HEMA: hydroxyethyl mercapturic acid

The urinary excretion of TdGA and HEMA in male Wistar rats following i.p. injection of 2,2'-DDE, 2-chloroethanol or 2-chloroacetaldehyde diethyl acetal (5 rats/chemical) was as follow:

Substance	BCEE		2 -chloroacetaldehyde diethyl acetal		2-chloroethanol
Dose (µmole)	40	160	40	160	40	160
TdGA (µmole)	19.3 +/- 6.9	42.3 +/- 15.3	26.3 +/- 11.7	39.5 +/- 17.3	11.8 +/- 1.7	22.8 +/- 6.5
HEMA (µmole)	2.5 +/- 0.9	3.4 +/- 1.3	0.36 +/- 0.12	0.91 +/- 0.24	0.06	0.06

Urinary excretion of HEMA was appreciably higher among the BCEE treated animals than among those treated with 2-Chloroacetaldehyde diethyl acetal or 2-chloroethanol. However, it remained lower than exretion of HEMA observed following inhalation exposure to vinyl chloride.

Conclusions:

5 male Wistar rats were given a single intraperitoneal dosing with Bis(2-chloroethyl)ether (BCEE),  2-chloroacetaldehyde diethyl acetal and  2-chloroethanol (doses: 40 or 160 µmole). BCEE was found to be mainly mletabolized as TdGA and to a lesser extent as HEMA. HEMA is considered to be produced through detoxification of chloroethylene oxide. The fact that exposure to BCEE lead to a lesser production of HEMA compared to exposure to vinyl chloride could be the sign that BCEE is not efficiently metabolised as chloroethylene oxide, and may explain its lower mutagenic and carcinogenic potential.

Executive summary:

In this published study, 5 male Wistar rats were given a single intraperitoneal dosing with Bis(2-chloroethyl)ether (BCEE),  2-chloroacetaldehyde diethyl acetal and  2-chloroethanol (doses: 40 or 160 µmole).

The results suggest dissimilar routes of metabolism among different chloroacetaldehyde producers in urine. While treatment of animals with vinyl chloride resulted in a higher urinary excretion of HEMA than TdGA, an inverse phenomenon was observed with the other compounds including BCEE. The authors suggest that this might be due to the production of chloroethylene oxide during the oxidative metabolism of vinyl chloride and that HEMA is derived from primary metabolites that cannot be produced by other chloroacetaldehyde producers. The production of HEMA from chloroethylene oxide is considered to represent a detoxification mechansim. The fact that exposure to BCEE lead to a lesser production of HEMA compared to exposure to vinyl chloride could be the sign that BCEE is not efficiently metabolised as chloroethylene oxide, and may explain its lower mutagenic and carcinogenic potential.

Reference 6

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

excretion

Qualifier:

no guideline followed

Principles of method if other than guideline:

- Principle of test: Inhalation experiment

GLP compliance:

not specified

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and of test material: Merck Co. (D-6100 Darmstadt, FRG)
- Purity: 99%

Radiolabelling:

no

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Zentralinstitut für Versuehstiere (D-3000 Hannover, FRG)
- Strain: strain/AF/Han
- Age at study initiation: not specified
- Weight at study initiation: 200-250g
- Housing: Makrolon cages
- Diet: ad libitum
- Water: ad libitum

Route of administration:

inhalation: vapour

Vehicle:

not specified

Details on exposure:

TYPE OF INHALATION EXPOSURE: nose only

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: inhalation chamber designed for nose-exposure

Duration and frequency of treatment / exposure:

8 h exposure in an inhalation chamber designed for nose-exposure of rats.

Dose / conc.:

10 ppm

Dose / conc.:

50 ppm

Dose / conc.:

100 ppm

Dose / conc.:

500 ppm

No. of animals per sex per dose / concentration:

8

Control animals:

not specified

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine
- Time and frequency of sampling: collected for 24 h

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled: urine
- Time and frequency of sampling: collected for 24 h
- From how many animals: 8/group
- Method type for identification: GC-FID

Metabolites identified:

yes

Remarks:

thiodiglycollic acid (TdGA)

Urinary excretion of thiodiglycollic acid (TdGA) in male Wistar rats following 8-h inhalation of Bis(2-chloroethyl)ether (BCEE):

Concentration of BCEE over the 8 h inhalation exposure (ppm)	Excretion of TdGA in urine over the 24 h (µmole)
0	0.37 +/- 0.13
10	0.71 +/- 0.20
50	1.36 +/- 1.00
100	1.47 +/- 0.40
500	1.93 +/- 0.27

The amounts of TdGA excreted in urine within 24 h of exposure to various doses of BCEE was a dose-dependent phenomenon (saturating relationship).

Conclusions:

Following inhalation exposure of 8 male rats to BCEE to various BCEE concentrations (10, 50, 100 and 500 ppm) for 8h, urinary excreted thiodiglycollic acid (TdGA) was monitored for 24h. Excretion of TdGA displayed a saturating dose-dependent relationship with the concentration of BCEE. Metabolism saturation began at BCEE concentrations higher than 50 ppm.

Executive summary:

This publication includes the results from an 8 h inhalation exposure of male Wistar rats at various BCEE concentrations (10, 50, 100 and 500 ppm). Following exposure, urine samples were collected for 24 h and thiodiglycollic acid (TdGA) concentrations in urine was measured. Excretion of TdGA displayed a saturating dose-dependent relationship with the concentration of BCEE, similar to that found after exposure to vinyl chloride. Metabolim saturation began at concentrations higher than 50 ppm.

Reference 7

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

3 (not reliable)

Rationale for reliability incl. deficiencies:

significant methodological deficiencies

Remarks:

only 2 animals were studied, limited information provided in the report

Objective of study:

excretion

Qualifier:

no guideline followed

Principles of method if other than guideline:

Study of the excretion of radiolabelled BCEE following a single administration to two female rhesus monkeys

GLP compliance:

no

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source of test material: not specified
- Purity: not specified

RADIOLABELLED TEST MATERIAL
- Radiochemical purity: not specified
- Specific activity: 0.56 µCi/mL

Radiolabelling:

yes

Species:

monkey

Strain:

other: rhesus

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: not specified
- Weight at study initiation: 8.9 and 9.75 kg, respectively
- Housing:
- Diet: 10-15 pellets of Purina chow per day
- Water: ad libitum

Route of administration:

oral: gavage

Vehicle:

other: see "details on dosing and sampling"

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
4 mL of ethanol was mixed to 4.3 mL Emulphor. 0.39 g of cold BCEE and c.a. 0.02 mL of radiolabelled BCEE (0.56 µCi/mL) were then added. The solution was diluted with 31.7 g of deionised distilled water. Liquid scintillation of this solution established the specific activity to be 0.49 µCi/mL (i.e. 10,906,00 dpm) and 106.27 dpm/µg BCEE.

DIET PREPARATION
- Frequency: single orale exposure to 10 mg/kg bw

Duration and frequency of treatment / exposure:

Single exposure

Dose / conc.:

10 mg/kg bw (total dose)

No. of animals per sex per dose / concentration:

2

Control animals:

no

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY
- Tissues and body fluids sampled: urine, faeces
- Time and frequency of sampling: Urine samples were collected (if available) at 6, 12, 24, 36, 48, 72, 96 h after dosing. Collection flasks were iced over the entire collection time. Feces samples were collected at 24, 48, 72 and 96 h.

Metabolites identified:

not measured

EXCRETION DATA:

Data on cumulative excretion of radiolabelled Bis(2-chloroethyl)ether (BCEE) in urine and feces of the two female rhesus monkeys (following a single oral exposure to 10 mg/kg bw) are presented in the table below:

Time post treatment (hours)	Percent dose (%)
	Female n°1			Female n°96
	Urine	Feces	Urine + feces	Urine	Feces	Urine + feces
6	16.66	-	16.66	-	-	-
12	29.85	-	29.85	0.003	-	0.003
24	43.43	0.22	43.65	43.23	0.04	43.27
36	48.60	-	48.88	43.50	-	43.50
48	51.03	0.64	51.67	61.05	0.09	61.14
72	53.30	0.76	54.06	62.96	0.65	63.61
96	*	1.10	54.40	63.45	1.63	65.08

* The monkey was placed back in the stock cage.

Both monkeys excreted over 40% of the dose in the urine by 24 h and from 53-63% by 72 h post exposure. Very little of the compound was recovered in the feces (total recovery ranged from 1.10 to 1.63% after 96 h).

PHYSIOLOGICAL OBSERVATIONS:

Rhesus n°1 began menstruation 24 hours fater dosing. Also within 24 hours hemorrhagic darkening occurred below her eyes. This symptom was not observed in the other female rhesus monkeys. Blood samples analysis were also performed on rhesus n°1 but the reliability and representativity of the data is limited.

Conclusions:

BCEE was administered to 2 female rhesus monkeys in a single oral dose of 10 mg/kg bw. From 54 to 65% of the dose as 14C was excreted in the urine and 1.10 to 1.63% in the faeces 96 hours post-exposure.

Executive summary:

In this published study, BCEE was administered to 2 female rhesus monkeys in a single oral dose of 10 mg/kg bw. From 54 to 65% of the dose as 14C was excreted in the urine and 1.10 to 1.63% in the faeces 96 hours post-exposure.

While this study was conducted on a species closely related to humans and is thus valuable to evaluate how BCEE might impact human health, only two animals were studied and various methodological deficiencies impact the reliability of the results.

Description of key information

Absorption through the oral, inhalation and dermale route:

Bis(2-chloroethyl) ether (hereafter named BCEE) is efficiently absorbed through the oral and inhalation route. Lingg et al. (1982) reported that only 2% of a single dose (40 mg/kg bw) of radiolabelled BCEE was excreted in the faeces. This result indicate that absoprtion across the gastrointestinal tract was essentially complete (i.e. 98%). Similar results were reported in two rhesus mokeys orally exposed to a single dose of BCEE (10 mg/kg bw), where only 1.10 and 1.63% of the dose was recovered in the faeces after 96h (Smith, 1985). Similar gastrointestinal absorption is also assumed in humans. Efficient gastrointestinal absoprtion of BCEE is supported by the small molecular weigth of the substance (143.01 g/mol), the moderate Log Kow (1.12 at 20°C). In rats exposed to vapors of radiolabelled BCEE in a closed all-glass exposure system, >95% of the administered BCEE (125.9 mg/kg bw) was taken up by the animals fater 18 h exposure (Gwinner et al., 1983). While no data is available for humans, effective absorption via the respiratory tract is to be assumed. No studies are available regarding the absorption of BCEE by the dermale route. However, acute dermale toxicity studies in rabbits (Unpublished report from Dow Chemical Co. cited Patty's Industrial Hygiene and Toxicology Vol. IIA, p. 2518) and in Guinea pigs (Smyth & Carpenter, 1948) support that BCEE exposure may occur through the dermal route. Skin absorption in humans is also expected.

Bioaccumulation potential:

Lingg et al. (1979) found that approximately 80% of an oral dose of Bis(2-chloroethyl) ether administered to rats was excreted within 48 hours. Most of the dose (64.7%) was excreted as urinary metabolites, only small amounts excreted in feces (2.4%) or expired air (11.5% with only 1.6% as unaltered Bis(2-chloroethyl) ether). Only c.a. 2% of the dose remained in the body (Lingg et al. 1982). From 54 to 65% of the dose as 14C-BCEE administered to 2 female rhesus monkeys (single oral dose of 10 mg/kg bw) was excreted in the urine and 1.10 to 1.63% in the faeces 96 hours post-exposure (Smith 1985). Following a 8 h inhalation exposure period of male Wistar rats at various BCEE concentrations (10, 50, 100 and 500 ppm), urine samples were collected for 24 h and thiodiglycollic (TdGA) concentrations in urine was measured. The excretion of TdGA displayed a saturating dose-dependent relationship with the conentration of BCEE, similar to that found after exposure to vinyl chloride. This suggests that transient systemic accumulation of BCEE metabolites may occur. Overall, available data indicate that BCEE is effectively excreted, and that it has a low tendency to accumulate in tissues.

These results are supproted by a study on the distribution on BCEE in organs and tissues. Lingg et al. (1982) administered a single oral dose of 14C-BCEE to rats, and measured the radioactive content of tissues 48 hours later. Only a small fraction of the dose (2.3%) was found in organs and tissues, with 0.96% in muscle, 0.56% in kidney, 0.49% in blood, 0.19% in liver, and 0.1% in other tissues. These findings suggest that BCEE is not preferentially accumulated or retained in any one tissue or organ of the body.

Overall, based on available evidences, BCEE is considered to have a low bioaccumulation potential.

Key value for chemical safety assessment

Bioaccumulation potential:

low bioaccumulation potential

Absorption rate - oral (%):

98

Absorption rate - inhalation (%):

95

Additional information

Six published studies includes data on the toxicocinetics, metabolism and distribution of BCEE. Most studies (5) were conducted on rats and one study was conducted on rhesus monkeys. While none of these studies were conducted according to current guidelines, the methdology used is generally acceptable and the obsorption, distribution, metabolims and excretion of BCEE has been fairly well characterised. However, no data is available on the dermal absorption of the compound. The following section synthetise available knowledge on BCEE.

ABSORPTION

1. Gastrointestinal tract:

Lingg et al. (1982) reported that only 2% of a single dose (40 mg/kg bw) of radiolabelled Bis(2-chloroethyl) ether (hereafter named BCEE) was excreted in the faeces, indicating that absoprtion across the gastrointestinal tract was essentially complete. Similar results were reported in two rhesus mokeys orally exposed to a single dose of BCEE (10 mg/kg bw), where only 1.10 and 1.63% of the dose was recovered in the faeces after 96h (Smith, 1985). Similar gastrointestinal absorption is also assumed in humans. Efficient gastrointestinal absoprtion of BCEE is also supported by the small molecular weigth of the substance (143.01 g/mol), the moderate Log Kow (1.12 at 20°C)

2. Respiratory tract:

In rats exposed to vapors of radiolabelled BCEE in a closed all-glass exposure system, >95% of the administered BCEE (125.9 mg/kg bw) was taken up by the animals fater 18 h exposure (Gwinner et al., 1983). While no data is available for humans, effective absorption via the respiratory tract is to be assumed.

3. Skin:

No studies are available regarding the absorption of BCEE by the dermale route. However, acute dermale toxicity studies in rabbits (Unpublished report from Dow Chemical Co. cited Patty's Industrial Hygiene and Toxicology Vol. IIA, p. 2518) and in Guinea pigs (Smyth & Carpenter, 1948) support that BCEE exposure may occur through the dermal route. Skin absorption in humans is also expected.

DISTRIBUTION

Lingg et al. (1982) administered a single oral dose of 14C-BCEE to rats, and measured the radioactive content of tissues 48 hours later. Only a small fraction of the dose (2.3%) was found in organs and tissues, with 0.96% in muscle, 0.56% in kidney, 0.49% in blood, 0.19% in liver, and 0.1% in other tissues. These findings suggest that BCEE is not preferentially accumulated or retained in any one tissue or organ of the body.

METABOLISM

Animal studies indicate that Bis(2-chloroethyl) ether is extensively metabolized, with thiodiglycolic acid (TDGA) being the principal endproduct (Lingg et al. 1979; Lingg et al. 1982; Norpoth et al. 1986). The pathway leading to TDGA probably involves oxidative cleavage of the ether bond to yield chloroacetaldehyde and 2-chloroethanol (Gwinner et al. 1983; Norpoth et al. 1986; Lingg et al. 1979, 1982; Muller and Norpoth 1979). TDGA recovered in urine usually accounts for 50% to 80% of a dose of BCEE (Lingg et al. 1979, 1982). Smaller amounts of BCEE (3% to 5%) are metabolized by oxidation or substitution at a chlorine without ether cleavage, and about 10% is degraded to CO2 (Lingg et al. 1982). Only about 2% of the dose is excreted via the lungs as unchanged parent compound (Lingg et al. 1979).

BCEEhas a much weaker alkylating activity than α-halo ethers, which suggest that the breakdown of the compound may be initiatedin vivoby an enzyme-catalyzed step. Gastric acid hydrolysis could also induce O-dealkylation leading to generating chloroethanol. A highly reactive intermediate product from cleavage of BCEE is suggested by the fact that no unconjugated cleavage products were found in the urine (Lingg et al. 1982). Muller et al. (1979) have reported that the reactive intermediate metabolite of BCEE is the same as that of Vinyl chloride (VC). The fact that BCEE and VC, a known hepatocarcinogen, are converted to a common reactive intermediate supports the bioassay evidence which suggests that BCEE is a carcinogen.

Gwinner et al. (1983) exposed 3 rats to 14C-labelled Bis(2-chloroethyl) ether and monitored the distribution of radioactivity in various tissues follofing covalent binding to protein 24h post exposure. The larger amount was found in the liver (0.32% of administered dose). Irreversibly bound metabolites were also found in the kidnzy (0.17%), the small intestine (0.12%), the lungs (0.07%), the spleen (0.06%) and muscles (0.01%). The presence of protein-bound label in these tissues suggested to the authors that reactive intermediates were formed that led to covalent adducts, but incorporation of label into protein might also have occurred through normal synthetic pathways involving non-toxic breakdown products from Bis(2-chloroethyl) ether. No label was detectable in liver DNA or RNA.

EXCRETION:

Lingg et al. (1979) found that approximately 80% of an oral dose of Bis(2-chloroethyl) ether administered to rats was excreted within 48 hours. Most of the dose (64.7%) was excreted as urinary metabolites, only small amounts excreted in feces (2.4%) or expired air (11.5% with only 1.6% as unaltered Bis(2-chloroethyl) ether). Only c.a. 2% of the dose remained in the body (Lingg et al. 1982).From 54 to 65% of the dose as 14C-BCEE administered to 2 female rhesus monkeys (single oral dose of 10 mg/kg bw) was excreted in the urine and 1.10 to 1.63% in the faeces 96 hours post-exposure (Smith 1985). Following a 8 h inhalation exposure period of male Wistar rats at various BCEE concentrations (10, 50, 100 and 500 ppm), urine samples were collected for 24 h and thiodiglycollic (TdGA) concentrations in urine was measured. The excretion of TdGA displayed a saturating dose-dependent relationship with the conentration of BCEE, similar to that found after exposure to vinyl chloride. This suggests that transient systemic accumulation of BCEE metabolites may occur. Overall, available data indicate that BCEE is effectively excreted, and that it has a low tendency to accumulate in tissues.