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Repeated dose toxicity: inhalation

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Administrative data

Endpoint:
chronic toxicity: inhalation
Remarks:
combined repeated dose and carcinogenicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
No data
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted according to scientifically acceptable protocol for such endpoint, with results found in general agreement with other similar studies. Nevertheless, this study contains some deviations: change of doses after 1 month of treatment, only 2 doses tested and infection by Mycoplasma pulmonis.
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
Unnamed
Year:
1986
Reference Type:
publication
Title:
Unnamed
Year:
1977

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 452 (Chronic Toxicity Studies)
Deviations:
yes
Remarks:
change of doses after 1 month of treatment, no results at 3 months of treatment, few parameters not examined
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): vinylidene chloride
- Physical state: liquid
- Analytical purity: 99 % (see table 1 for details)
- Source: The Dow Chemical Company
- stabilized with hydroquinone monomethylether

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Spartan Research Animals, Haslett, Michigan
- Age at study initiation: 6-7 weeks
- Housing: 2-3/cage
- Diet: Purina Laboratory Chow, Ralston Purina Co., St. Louis, Mo. ad libitum
- Water: ad libitum (expect during exposure)

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: unchanged (no vehicle)
Remarks on MMAD:
MMAD / GSD: test substance was a vapour; MMAD and GSD not relevant.
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 3.7 m3 stainless steel chambers
- Temperature, humidity, pressure in chamber: ambient conditions
- System of generating particulates/aerosols: glass vaporization flask maintained at approximately 30 °C

TEST ATMOSPHERE
- Brief description of analytical method used: vapor phase infrared spectrometry (Perkin-Elmer 12A; path length = 10 m and wavelength = 12.6 µm)
- Samples taken from breathing zone: yes 2-5 times during the 6-hr daily exposure period
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Vapor phase infrared spectrometry (Perkin-Elmer 12A; path length = 10 m and wavelength = 12.6 µm). Chamber concentrations were determined by interpolating from a standard curve using peak height measurements. Standards were prepared by injecting a known volume of vinylidene chloride into a bag made of Saran resin which was filled with a measured amount of filtered, compressed air.
Details of test atmospheres: see table 2
Duration of treatment / exposure:
1, 6, 12 and 18 months
Frequency of treatment:
6 h/day, 5 days/week
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
100 mg/m3
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
300 mg/m3
Basis:
nominal conc.
No. of animals per sex per dose:
1 month period: 4M/4F for control, 40 and 160 mg/m3
6 months period: 5M/5F for control, 100 and 300 mg/m3
12 months period: 5M/5F for control, 100 and 300 mg/m3
18 months period: 86M/86F for control, 85M/85F for 100 mg/m3 and 86M/86F for 300 mg/m3
Control animals:
yes, concurrent no treatment
Details on study design:
During the first 5 weeks, exposure levels were 40 and 160 mg/m3. As no effects were found on 4 killed rats/sex/group at 4 weeks of exposure, exposure levels were raised to 100 and 300 mg/m3 at the end of 5 weeks.
At 6 and 12 months of exposure, 5 rats/sex/group were also sacrificied.
86 rats/sex/group remained after the interim kills
Positive control:
No data

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

BODY WEIGHT: Yes
Individual body weights were recorded weekly for the first month of exposure, biweekly during the second month of exposure, and a minimum of monthly thereafter. Body weights of animals fasted overnight were measured at the time of scheduled sacrifices.

HAEMATOLOGY: Yes
Blood was collected for routine hematology from the tail vein of five animals/sex/exposure group shortly before the 6- and 12-month interim sacrifices and 10 animals/sex/exposure group at the terminal sacrifice.
Haematology parameters: red blood cell count, hemoglobin concentration, packed cell volume and total and differential white cell count.

CLINICAL CHEMISTRY: Yes
blood was collected from the cervical vessel during the necropsy procedure at the interim sacrifices and on 5 animals/sex/exposure group at the terminal sacrifice
Parameters measured: serum glutamic pyruvic transaminase (SGPT), blood urea nitrogen (BUN) and alkaline phosphatase (AP). Blood glucose was also determined at the 12-month interim sacrifice, repeated for confirmation at 18 months on blood obtained by orbital sinus puncture on 5 animals/ sex/exposure group, and on all remaining animals at the terminal sacrifice.

URINALYSIS: Yes
- Time schedule for collection of urine: urine samples from 10 animals/sex/exposure group shortly before the 6- and 12-month sacrifices and at the terminal sacrifice.
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters: determinations of specific gravity, pH, glucose, protein, ketones, occult blood, bilirubin and, at the terminal sacrifice only, urobilinogen. Specific gravity measurements were made using a T.S. Meter. All other measurements were made using Bililabstix.
Sacrifice and pathology:
A complet gross pathologic examination was performed on all animals. Rats alive were anesthetized with methoxyflurane and sacrificed by exsanguination.
The following tissues were removed and placed in neutral buffered 10 % Formalin: accessory sex glands, adipose tissue, adrenals, aorta, bone marrow (sternal), brain, epididymes, esophagus, heart, intestines (large and small), kidneys, liver, lungs, lymph nodes (mesenteric, mediastinal), mammy gland, nasal turbinates, ovaries, oviduct, pancreas, parathyroid, peripheral nerve, pituitary gland, prostate, salivary glands, skeletal muscle, skin, spleen, spinal cord, sternum, stomach, testes, thymus, thyroid gland, trachea, urinary bladder, uterus, and any gross lesion or mass. Eyes of rats which died or were culled from the study were preserved in Formalin. Eyes of a portion of the rats from the interim or terminal sacrifices were fixed in Zenker's fixative with the remainder in Formalin.
Other examinations:
No data
Statistics:
Body weight data, clinical chemistry, hematology, urinalysis (specific gravity), and organ weight data: analysis of variance and Dunnett's test (Steel and Torrie, 1960).
Data for cumulative mortality, incidence of gross and microscopic pathological findings, and tumor incidence: Fischer's exact test (Siegel, 1956).
Level of significance : 0.05

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
no effects observed
Details on results:
Intercurrent murine pneumonia was observed among animals of both sexes during the last 6 months of the study
No eye or nasal irritation or changes in appearance

MORTALITY (Table 3):
Statistically significant non-dose-related mortality increases among female rats at 100 mg/m3 during the 15th to 22nd months
Trend towards increased cumulative percentage of mortality among female rats at 300 mg/m3 during the 14th to the 24th months (statistically significant at months 15, 17 and 21)

BODY WEIGHTS (Table 4):
Mean body weights of male rats at 100 mg/m3 significantly lower than control for the first 13 months of exposure and remained lower but not systematically significant.
Mean body weights of male rats at 300 mg/m3 significantly lower from 6th to 12th months. General trend toward decreased body weights from the 17th to 24th months
Mean body weights of female rats exposed to vinylidene chloride generally significantly greater than control for the first 6 months.

HEMATOLOGY:
no effects

CLINICAL CHEMISTRY:
blood glucose was significantly lower only for the male rats exposed to 300 mg/m3 at the 12th month (Mean±SD: 134.6±9.6 vs. 102.8±7.0 for the control and the 300 mg/m3 groups, respectively)

URINALYSIS:
no effects

ORGAN WEIGHTS:
Significant changes only at the 12th month:
- Male rats exposed to vinylidene chloride had significantly lower absolute liver weights (Mean±SD: 20.57±1.51, 15.28±2.10 and 15.13±2.06 for the control, the 100 mg/m3 and the 300 mg/m3 groups, respectively) with a concomitant decrease in fasted body weights (Mean±SD: 660±44, 576±48 and 563±41 for the control, 100 mg/m3 and 300 mg/m3 groups, respectively).
- Female rats exposed to vinylidene chloride had significantly increased absolute kidney weights (Mean±SD: 2.09±0.10, 2.44±0.26 and 2.60±0.18 for control, 100 mg/m3 and 300 mg/m3 groups, respectively)

PATHOLOGY:
Incidence of lesions of the respiratory tissues due to infection by Mycoplasma pulmonis was lower in the control than in the groups exposed to vinylidene chloride.
Changes related to the treatment in the liver: minimal midzonal hepatocellular fatty change in both male and female at the 6th and 12th months and in female rats dying during the 18-month exposure period (Table 5). However, no such incidence was observed at the terminal sacrifice or among the animals of the postexposure period.

No increase in any tumor type except a significant increase in mammary adenocarcinomas in the females exposed to 100 mg/m3 (Animals with mammary gland carcinoma : 2/84, 7/86* and 4/84 for control, 100 mg/m3 and 300 mg/m3 groups, respectively; *Significantly different from control by Fisher's exact probability test). Significantly decreased incidence of several tumor types in vinylidene chloride exposed rats: pituitary adenomas in male and female rats at 100 and in male rats at 300 mg/m3, pancreatic islet cell adenomas and thyroid adenocarcinomas in male rats at 300 mg/m3. Taking into account the (lack of ) consistency of the observed differences in tumor numbers, and the historical data for the mammary adenomacarcinomas, none of these observations were judged to be attributable to exposure to 1,1-dichloroethene.

Based on the hepatocellular fatty changes in the midzonal region of hepatic lobule, the NOAEL in male rats in this study is 300 mg/m3 (the highest exposure tested). The NOAEL in female rats in this study is 100 mg/m3; the LOAEL is 300 mg.

Effect levels

Dose descriptor:
LOAEC
Effect level:
100 mg/m³ air (nominal)
Sex:
male/female
Basis for effect level:
other: Reversible minimal midzonal hepatocellular fatty change

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Table 3: Selected cumulative number (percentage) of dead female rats

Exposure level (mg/m3)

Months on study

0

100

300

Number of rats

84

86

84

6

1 (1)

1 (1)

1 (1)

8

1 (1)

1(1)

1 (1)

9

1 (1)

3 (4)

1(1)

11

2 (2)

6 (7)

1(1)

12

2  (2)

6 (7)

5 (6)

13

4 (5)

9 (11)

6 (7)

14

4 (5)

10 (12)

7 (8)

15

5 (6)

18 (21) *

13 (16) *

16

8 (10)

19 (22) *

16 (19)

17

9 (11)

19 (22) *

18 (21) *

18

16 (19)

29 (34) *

20 (24)

19

23 (27)

37 (43) *

29 (35)

20

30 (36)

49 (57) *

41 (49)

21

39 (46)

55 (64) *

51 (61) *

22

46 (55)

59 (69) *

54 (64)

23

56 (67)

65 (76)

61(73)

24

64 (76)

72 (84)

68 (81)

* Statistically different from controls by Fischer's exact probability test

Table 4: Mean body weights of male and female rats at selected times

Exposure level (mg/m3)

Months on study

0

100

300

Males

0

276

276

283 *

1

399

386 *

396

3

489

469 *

488

6

562

529 *

549

9

598

555 *

565 *

12

639

593 *

586 *

15

638

623

646

18

627

586 *

618

21

607

582

563 *

24

587

550

530

Females

0

210

212

203 *

1

259

259

264

3

294

301 *

309 *

6

333

343 *

342 *

9

340

348

348

12

368

377

363

15

386

391

402

18

396

419

426

21

443

412

449

24

427

463

439

* Statistically different from control by analysis of variance and Dunnett's test

Table 5: Incidence of midzonal hepatocellular fatty change in male and female rats

No. of animals with fatty

change/no. of animals examined

Exposure level (mg/m3)

 Time of examination

0

100

300

 At scheduled sacrifice

      6-month interim

0/5 M

1/5 M

4/5 M

0/5 F

2/5 F

4/5 F

      12-month interim

0/5 M

3/5 M

5/5 M

0/5 F

5/5 F

5/5 F

      24-month terminal

1/13 M

0/13 M

0/8 M

1/19 F

0/11 F

0/16 F

 At death or following culling

0/27 M

0/25 M

1/27 M

      During 18-month exposure period

0/16 F

6/29 F

7/20 F

      During 6-month postexposure period, but not including

0/46 M

1/47 M

0/51M

           24-month terminal sacrifice

0/49 F

0/46 F

1/48 F

Applicant's summary and conclusion

Conclusions:
Based on the hepatocellular fatty changes in the midzonal region of hepatic lobule, the NOAEL in male rats in this study is 300 mg/m3 (the highest exposure tested). The NOAEL in female rats in this study is 100 mg/m3; the LOAEL is 300 mg.
Executive summary:

Male and female Sprague-Dawley rats were exposed to 1,1-dichloroethene by inhalation at 0, 100 and 300 mg/m3, 6 h/day, 5 days/week for exposure period of up to 18 months, followed by a 6-month recovery period. No exposure related changes in mortality, appearance and demeanor, body weight, clinical chemistry determinations, hematologic evaluations, and urinalysis were observed. Minimal hepatocellular fatty change in the midzonal region of the hepatic lobule was observed in both male and female rats after 6 and 12 months of exposure at both exposure levels. After 18 months of exposure, the midzonal hepatocellular fatty change was only observed in the female rats exposed to 150 mg/m3. After a 6 month post-inhalation period, the hepatic fatty change was no longer discernable. Accordingly the NOAEL for male rats in this study is 300 mg/m3(the highest exposure concentration), the NOAEL for female rats is 100 mg/m3, the LOAEL is 300 mg/m3.