Tetrahydrofuran

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14 July 1987 to 23 October 1987

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Although no specific test guideline was identified, the study was conducted according to valid and internationally recognized test procedures that followed recognized GLP standards.

Data source

Reference

Reference 1

Reference Type:

publication

Title:

Unnamed

Year:

1992

Reference 2

Reference Type:

study report

Title:

Unnamed

Year:

1988

Report date:

1988

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Principles of method if other than guideline:

Although no specific testing guideline was identified, the study was conducted generally according to the method specified in OECD Guideline 414. Pregnant Sprague-Dawley female rats were exposed to vapor concentrations of tetrahydrofuran of 0, 600, 1800 or 5000 ppm for 6 hours/day, 7 days/week on days of gestation (DG) 6-19. Developmental evaluations were performed on pregnant rats euthanized on DG 20.

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Tetrahydrofuran

EC Number:

203-726-8

EC Name:

Tetrahydrofuran

Cas Number:

109-99-9

Molecular formula:

C4H8O

IUPAC Name:

tetrahydrofuran

Details on test material:

- Name of test material: tetrahydrofuran- Supplier: ChemCentral, Kansas City, MO, USA- Lot No.: BNW 51437-15- Analytical purity: 99.3 to 100.2% (bulk chemical, gas chromatography); analyzed on 2/6/87, 3/19/87, 5/1/87 and 9/9/87- Storage conditions: under nitrogen at 65 deg F in grounded containers- Stability under test conditions: tetrahydrofuran was monitored during exposures with mean measured concentrations in the exposure chambers within 2% of target concentrations. Tetrahydrofuran was found to be stable in the generator reservoir and the exposure chambers- Peroxide content: tetrahydrofuran was analyzed once/month after opening for peroxide formation and was found to contain less than 100 ppm at each analysis. Chamber atmospheres were analyzed for organic peroxides (p-hydroxyphenylacetic acid-coupling reaction) and levels were below the limit of detection.- Degradation products: chamber atmospheres were also tested for the presence of potential breakdown products including furan, isopropanol, and butyraldehyde. None of these were detected at concentrations in excess of 0.1%.

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

- Source: Charles River, Raleigh, NC- Age at study initiation: 14 weeks- Fasting period before study: none- Housing: stainless steel wire racks equipped with automatic watering system (5-6 rats/cage)- Diet: pelleted NIH-07 (Ziegler Bros. Inc., Gardners, PA) ad libitum except during 6-hour exposure periods- Water (ad libitum): ad libitum throughout the study including exposure periods- Acclimation period: quarantined 20 days prior to the start of exposuresENVIRONMENTAL CONDITIONS- Temperature (°F): during quarantine, 72 +/- 3; during exposures, 75 +/- 3- Humidity (%): during quarantine, 50 +/- 15; during exposures, 55 +/- 15- Chamber airflows: 12 to 18 CFM (average in all chambers, 14.0 to 15.4 CFM)- Photoperiod (hrs dark / hrs light): 12/12In-Life Dates: 11 September 1987 (mating) to 23 October 1987 (fetal exams)

Administration / exposure

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

other: conditioned air

Details on exposure:

Exposure Chambers:Batelle-designed inhalation exposure chambers (Harford Systems; Lab Products Inc., Aberdeen, MD) were employed. The 2-3 cubic meter stainless-steel chambers (1.7 cubic meters active mixing volume) contained three levels of caging. The cage units accommodated individual animal cages, feed troughs, and automatic watering systems.Vapor Generation/Exposure System:Tetrahydrofuran was pumped from a bulk storage reservoir at a steady rate by a liquid micrometering pump into a rotating flask (100 rpm) partially immersed in a hot water bath at 175 deg F. Vapor from the flask was carried by a metered stream of nitrogen. The vapor entered a short distribution manifold from which individual delivery lines carried metered amounts of the vapor to each exposure chamber. Target vapor concentrations were achieved by diluting with filtered air immediately before entering the chambers.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Vapor Concentration Monitoring:Vapor concentrations were monitored during animal exposures at approximately 30-minute intervals by gas chromatography (Hewlett-Packard 5840 GC) using a 1/8 in. o.d. X 1.0 foot nickel column packed with 1% SP-1000 on 60/80 mesh Carbopack B operated at 145 deg C. An 8-port value allowed measurements of concentrations of the test material in the three exposure chambers, the control chamber, the holding chamber, the exposure room, an on-line standard, and nitrogen blank. A 400 ppm tetrahydrofuran certified standard in nitrogen (MG Industries Scientific Gases, Los Angeles, CA) was used to check instrument drift throughout the exposure day.The precision of the on-line concentration monitoring was estimated from 17 consecutive measurements of the 400-ppm on-line THF standard. A 0.26% coefficient of variation was observed. The limit of detection of tetrahydrofuran was 0.04 ppm.Vapor Concentration Uniformity:The uniformity of chamber atmosphere vapor concentrations were measured prior to the study and once during the study. All chambers were acceptable with percentage relative standard deviations of

Details on mating procedure:

- Impregnation procedure: co-housed with male rats - Ratio F/M per cage: 2 or 3 females/male - Length of cohabitation: overnight - Mating was conducted for 5 consecutive nights to obtain approximately 33 pregnant animals/group.- Proof of pregnancy: confirmed by a sperm-positive vaginal smear, referred to as day 0 of pregnancy (0 DG)

Duration of treatment / exposure:

14 consecutive days (6-19 DG for mated rats)

Frequency of treatment:

6 hours/day

Duration of test:

20 days

No. of animals per sex per dose:

10 virgin (unmated) females and approximately 33 positively mated females

Control animals:

yes

Details on study design:

Virgin female rats were included to provide a reference for any effects that the state of pregnancy may have had on the toxic response. These animals were exposed concurrently with mated animals.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS:Animals were observed daily for mortality, morbidity and signs of toxicity. The date and time of death or euthanasia of moribund animals was recorded and the animals necropsied.DETAILED CLINICAL OBSERVATIONS: No dataBODY WEIGHT: Yes- Time schedule for examinations: Mated female rats were weighed on day of gestation (DG) 0, 6, 10, 14, 17 and 20; virgin rats were weighed prior to the start of exposures and on exposure days 1, 5 and 10 and at euthanization.POST-MORTEM EXAMINATIONS: YesRats were killed by CO2 asphyxiation, weighed and examined grossly for signs of maternal toxicity.

Ovaries and uterine content:

Both ovaries from each female were saved for sectioning and quantitative follicle counts. Apparently nongravid uteri from positively mated females were stained with 10% ammonium sulfide to detect possible implantation sites. The number, position and status of implants were recorded for each gravid uterus. Placentas were examined and discarded unless abnormal.

Fetal examinations:

- External examinations: Live fetuses were weighed and examined for gross defects.- Soft tissue examinations: The sex of live fetuses was determined by internal examinations of the gonads after injection of Nembutal(TM) (sodium pentobarbital). Fifty percent of live fetuses and any fetuses with gross external abnormalities were examined for visceral defects by dissection of fresh tissue.- Skeletal examinations: All fetal carcasses, with and without heads, were prepared for skeletal staining. Cartilage as well as ossified bone were visualized by double-staining using Alcian bue and alizarin red S.- Head examinations: The heads of 50% of the live fetuses were removed and placed in Bouin’s fixative. After fixation, the heads were serially sectioned with a razor blade and examined for soft-tissue craniofacial abnormalities.

Statistics:

All means and standard deviations for animal data were calculated. Mean body weights (as a mean of litter means for fetal data) analyzed by an analysis of variance (ANOVA) model for unbalanced data. Response variables, either body weights or the arcsin transformations of proportional incidence data, were analyzed against the class variable, treatment, in a one-way ANOVA model. A Tukey’s t-test (two-tailed) was used to assess statistically significant differences between the control and exposed groups. If appropriate, a dose-response relationship was determined by means of an orthogonal trend test (Winer, 1971). In the case of proportional data this test was performed on transformed variables. The litter was used as the basis for analysis of fetal variables.

Indices:

The following parameters, expressed as mean +/- SE, when appropriate, were computed from data for all pregnant animals and their litters:- Number of dead maternal animals, animals removed from the study and reason for removal- Summary of maternal toxicity, including incidences of changes detected during clinical observations- Number and percentage of pregnant animals- Maternal body weights- Weights of gravid uterus- Extragestational weight and weight gain- Number of implantation sites/litter- Number of litters with live fetuses- Number and percent of live fetuses/litter- Body weight of live fetuses/litter- Body weight of male and female fetuses/litter- Sex ratio of fetuses/litter- Number and percent of early and late resorptions/litter- Number and percent of non-live/litter (early and late resorptions and dead fetuses)- Listings of malformations and variations observed in fetuses/litters- Number and percent of malformed fetuses- Number and percent of litters with malformed fetuses

Historical control data:

Contemporary control data (based on N=80 litters) for Sprague-Dawley rats was available in addition to the unexposed (0 ppm) control animals:Maternal Wt (18 DG): 404.8 +/- 29.0Gravid Uterine Wt: 79.6 +/- 18.7Extragestational Weight Gain: 48.2 +/- 15.0Implants: 15.7 +/- 2.9Live Fetuses: 14.7 +/- 3.3 (92.5 +/- 9.0%)Early Resorptions: 0.9 +/- 1.1 (6.4 +/- 8.6%)Late Resorptions: 0.2 +/- 0.5 (1.1 +/- 3.3%)Dead Fetuses: 0.0 +/- 0.0 (0%)Total Intrauterine Death: 1.1 +/- 1.1 (7.5 +/- 9.0%)All Fetuses ( Weight, g): 3.55 +/- 0.33 male (g): 3.64 +/- 0.39 female (g): 3.45 +/- 0.32

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:yesDetails on maternal toxic effects:There were no maternal deaths; however, alopecia was present in 12 exposed rats (5 at 600 ppm; 3 at 1800 ppm; 4 at 5000 ppm). There was no alopecia noted in controls.Maternal body weight gains were reduced in exposed rats at the 5000 ppm dose level. Mean body weights on DG 6, 10, 14, 17 and 20 for control and 5000 ppm treated rats were as follows:- Controls (N=31): 306.4 +/- 18.5; 324.7 +/- 20.0; 344/4 +/- 21.7; 388.9 +/- 23.3; 405.3 +/- 29.0- 5000 ppm (N=32): 299.1 +/- 26.2; 302.8 +/- 24.0*; 324.0 +/- 27.0*; 349.8 +/- 29.4*; 380.8 +/- 32.8** significantly different from control groups, p<0.05Uterine weights and extragestational weight gains (EGWG: body weight at sacrifice - uterine weight - DG0 weight) were reduced in the 5000 ppm exposed rats but the decreases were not statistically significant:- Uterine Wts: 76.8 +/- 21.1 (control); 66.5 +/- 15.1 (5000 ppm)- EGWG: 48.5 +/- 14.7 (control); 38.7 +/- 16.2 (5000 ppm)

Effect levels (maternal animals)

1

Dose descriptor:

NOAEL

Effect level:

1 800 ppm (nominal)

Basis for effect level:

other: maternal toxicity

2

Dose descriptor:

NOAEL

Effect level:

1 800 ppm (nominal)

Basis for effect level:

other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yesDetails on embryotoxic / teratogenic effects:Each exposure group consisted of 32 to 33 sperm-positive female rats and 10 virgin female rats. Exposures had no effect on the number of implantations, the mean percent of live pups/litter, or the mean percent of resorptions/litter with respect to the control or to contemporary control data. The ratio of males to females in the litters was not affected. Refer to Table 1.Fetal body weights (as means of litter means) were significantly reduced for the 5000 ppm exposure group when compared to controls:- All fetuses (g): 3.5 +/- 0.3 (control); 2.9 +/- 0.2 (5000 ppm, p<0.05)- Males (g): 3.6 +/- 0.4 (control); 3.0 +/- 0.3 (5000 ppm, p<0.05)- Females (g): 3.5 +/- 0.2 (control); 2.8 +/- 0.2 (5000 ppm, p<0.05)A trend test indicated a significant correlation between fetal weight reduction and increasing exposure concentration, but was due to the abrupt drop in fetal weight between the 1800 and 5000 ppm exposure groups and did not represent a gradual decline in increasing exposure concentrations.Fetal Malformations:The incidence of fetal malformations was not significantly affected following gestational exposures to tetrahydrofuran. However, several malformations were observed at low incidences in the 5000 ppm group that were not present either in the 0 ppm controls or contemporary controls: anal atresia, cleft palate, vestigal tail, and vertebral agenesis. In all cases, the incidence rate of these latter malformations, as a percent of total fetuses examined, was less than 0.4% in all cases and represented only one or two fetuses of the total number examined.

Effect levels (fetuses)

Remarks on result:

other: see results above

Remarks:

see results above

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

Table 1: Reproductive Measures for Rats Exposed to Tetrahydrofuran

	Tetrahydrofuran exposure conc. (ppm)
	0	600	1800	5000
Number of
Sperm-positive females	33	32	32	32
Pregnant	31	28	29	32
Implantations/dam	15.5±3.7	16.7±3.0	15.4±2.3	15.7±3.2
Live fetuses/litter	14.5±4.2	15.4±3.3	14.5±2.5	14.8±3.4
Resorptions/litter
Total	1.1±1.2	1.3±3.6	0.9±1.4	0.9±1.0
Early	1.0±1.2	1.3±3.6	0.7±1.2	0.8±1.0
Late	0.1±0.4	0.0±0.2	0.2±0.5	0.1±0.3
Litters with resorptions	18	12	13	18
Percent of
Pregnant	94	88	91	100
Live fetuses/litter	91.6±11.1	93.3±15.1	94.2±8.5	93.8±10.4
Resorptions/litter
Total	8.4±11.1	6.7±15.1	5.8±8.5	6.2±10.4
Early	7.7±10.9	6.4±15.2	4.2±7.4	5.8±10.4
Late	0.7±2.9	0.2±1.2	1.6±3.8	0.4±1.5
Litters with resorptions	55	43	45	56
Male fetuses	46.9±13.6	48.4±15.2	48.0±16.6	49.4±16.3

Applicant's summary and conclusion

Conclusions:

Results of this study indicate that exposure of Sprague-Dawley rats to 0, 600, 1800 or 5000 ppm of tetrahydrofuran did not result in selective toxicity to the offspring. The only overt sign of maternal toxicity observed was a significant reduction in body weight for the 5000 pm group, which was due to a reduced rate of gain from 6 to 11 DG. No treatment-correlated reductions in virgin females were observed. The only observed fetal effect was a significant reduction in fetal body weight for the 5000 ppm group. There were no treatment-related increases in the incidence of intrauterine mortality, or in the frequency of fetal malformations or variations. The no-observed-adverse-effect level (NOAEL) for maternal and developmental toxicity in the rat was 1800 ppm.

Executive summary:

Pregnant and virgin female Sprague-Dawley rats were exposed to 0, 600, 1800 or 5000 ppm of tetrahydrofuran vapors whole-body for 6 hours/day, 7 days/week on days of gestation (DG) 6 to 19. Pregnant rats at the 5000 ppm exposure concentration displayed decreased body weights, however, virgin female rats were not similarly affected indicating that pregnancy was a factor in the toxicological response. Mean gravid uterine weights and extra-gestational weight gains were reduced at the 5000 ppm exposure concentration relative to the control, but the differences were not significant. There were no effects on the percentage of live rat fetuses/litter or on fetal sex ratios. Fetal body weights were reduced at the highest exposure concentration. No statistically significant increases in fetal malformations were observed, but several malformations were observed at low incidences (< 0.4% in all cases) that were not present in either the concurrent control or contemporary (historical) control animals. These included anal atresia, cleft palate, vestigal tail, and vertebral agenesis. The NOAEL for both maternal and developmental toxicity in the rat was 1800 ppm.