Formaldehyde

Repeated dose toxicity: oral

Administrative data

Endpoint:

chronic toxicity: oral

Remarks:

combined repeated dose and carcinogenicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

no analytical control of concentrations in drinking water; however, instability of the formaldehyde in water is not expected; a study on analysis of stability in water is in preparation and results will be submitted after termination of the study

Data source

Reference

Reference 1

Reference Type:

publication

Title:

Two-years drinking-water study of formaldehyde in rats

Author:

Til HP, Woutersen R, Feron V, Hollanders V, Falke H, Clary J

Year:

1989

Bibliographic source:

Fd Chem Toxic 27: 77-87

Reference 2

Reference Type:

study report

Title:

Unnamed

Year:

1988

Report Date:

1988

Materials and methods

Test guideline

Qualifier:

equivalent or similar to

Guideline:

OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)

Deviations:

no

GLP compliance:

yes

Remarks:

(TNO)

Limit test:

no

Test material

Reference

Name:

Unnamed

Type:

Constituent

Test material form:

gas

Specific details on test material used for the study:

- Lot/Batch number
UN2213 (Celanese Chemicals, USA)
- Specification: Paraformaldehyde prills 95% plus 5% water

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals and environmental conditions:

- Source: TNO central institute for the breeding of laboratory animals, Zeist, Netherlands
- Age/weight at study initiation: 5 weeks old / no data on body weight
- Acclimatization period 9 days;
- Housing condition:
rats kept 5 per cage; diet and tap water ad libitum; temperature 20-24°C, rel. air humidity 40-70%, 12 h light/12 h dark.

Administration / exposure

Route of administration:

oral: drinking water

Vehicle:

unchanged (no vehicle)

Details on oral exposure:

Fresh solutions prepared once weekly. Glass bottles filled daily and cleaned once weekly.

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

Up to 105 weeks (interim sacrifice after 12 or 18 months)

Frequency of treatment:

Drinking water daily ad libitum

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

5 mg/kg bw/day (nominal)

Remarks:

1.2 mg/kg bw/day measured dose levels in males (mean)
1.8 mg/kg bw/day measured dose levels in females (mean)

Doses / concentrations 2

Dose / conc.:

25 mg/kg bw/day (nominal)

Remarks:

15 mg/kg bw/day measured dose levels in males (mean)
21 mg/kg bw/day measured dose levels in females (mean)

Doses / concentrations 3

Dose / conc.:

125 mg/kg bw/day (nominal)

Remarks:

82 mg/kg bw/day measured dose levels in males (mean)
109 mg/kg bw/day measured dose levels in females (mean)

No. of animals per sex per dose:

70 m and 70 f per dose; subgroups of 10 rats/sex/dose killed 12 or 18 months after start of exposure.

Control animals:

yes, concurrent vehicle

Details on study design:

Concentrations adapted in weekly intervals to changing body weights up to week 52. Average concentration: 0, 20, 260, or 1900 mg/L (0, 0.002, 0.026, or 0.19% in water).

Positive control:

no

Examinations

Observations and examinations performed and frequency:

- Observations: Clinical symptoms, mortality, food and water intake, body weight (once weekly), haematology, clinical chemistry, urinalysis.

- Clinical signs
At least once daily each rat was observed for clinical signs.
- Mortality
See clinical signs.
- Body weight
Measured once weekly in the 1st 12 weeks starting day 0. Thereafter once every 4 weeks.
- Food consumption
Calculated for each treatment week during the 1st 12 weeks; thereafter over 2-week periods every 3 months .
- Water consumption
Calculated for each treatment week.
- Ophthalmoscopic examination
observations of cornea, conjunctivae, sclera, iris and fundus oculi made in all rats of the control and top-dose groups at week 0, 52 end 103
- Haematology
In week 26 and 103 blood samples of 10 rats per dose per sex were taken for analysis of haemoglobin concentration, erythrocyte and leukocyte counts, packed cell volume; and throbocytes. Glucose (fasting overnight) was examined in whole blood samples at week 27, 52, 78, and 104.
- Clinical Chemisty
Blood samples of 10 rats per dose per sex collected at week 28, 53, 79, and 105 were analysed for alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, total protein, albumin, total bilirubin, urea, creatinine, cholesterol, gamma-glutamyl transferase, inorganic phosphate, calcium, chloride, sodium, potassium.
- Urinalysis
In week 27, 52, 78, 104 rats (10 rats per dose per sex) were deprived of water for 24 h and last 16 h urine collected. Urine volume and density determined. In pooled urine samples collected at week 27 and 104 semi-quantitative observations of protein, glucose, occult blood, ketones, urobillinogen and billirubin were made. Sediment was examined via light microscopy.

Sacrifice and pathology:

Survivors of two subgroups (each 10 rats per dose per sex) were killed at week 53 and at week 79. Survivors of 50 rats per dose per sex were killed at week 105.
- Organ Weights
Determined of adrenals, brain, liver, heart, kidneys, spleen, testes, pituitary, thyroid, ovaries.

- Gross pathology
Necropsy of rats in all 4 main and sub-groups performed after sacrifice.

- Histopathology
All organs listed in OECD453 were examined in control and high dose groups. Liver, lungs, stomach and nose were examined also in low and mid dose groups. Furthermore, adrenals, kidney, spleen, testes, thyroid, ovaries, pituitary and mammary gland of rats killed at week 105 of low and mid dose group were examined.
Tissues of rats that were found dead or killed moribund were also preserved if autolysis was not advanced.

Statistics:

Suitable statistical methods were used (one-way analysis of covariance; Dunnett's test; Mann-Whitney U-test and Fishers exact test), the level of significance was p=0.05.

Results and discussion

Results of examinations

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

There was no toxicologically significant difference in mortality between treatment groups and controls.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

There was no toxicologically significant difference in mortality between treatment groups and controls.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

- 5 & 25 mg/kg bw: No effects
- 125 mg/kg bw: in males significant decrease from week 1 onwards and in females from week 24 onwards.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

5 & 25 mg/kg bw: no effects
125 mg/kg bw: In males (in females less pronounce) significantly decreased food intake.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

effects observed, treatment-related

Description (incidence and severity):

5 & 25 mg/kg bw: no effects
125 mg/kg bw:In males & females water consumption significantly decreased (ca. 40% reduction).

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

At week 27 and 79 slight (no details given on statistical significance) changes were seen in clinical chemistry of mid and high dose rats. But no significant effects were noted at week 53 and 105. Therefore, these effects were not considered to be of toxicological relevance.

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

A significant increase in urine density and decreased volume in the high dose group in m&f at week 27 and in m at week 52 was reported. These effects were due to reduced water intake.
Mean urinary pH was increased in m at 5 and 25 mg/kg bw in week 27 and 78 and was decreased in females at 125 mg/kg bw in week 27, 52, and 78. Occult blood was increased in males of all treatment groups at week 27 and in females at 25 and 125 mg/kg bw in week 104. A clear dose-effect relationship was not given suggesting no toxicological significance.

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

5 & 25 mg/kg bw: no effects
125 mg/kg bw: relative brain weight was increased in m & f, rel. testis weight increased in males and relative kidney weight increased in females. These increases are considered to be a result of decreased body weight.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Treatment related changes were detected only in the stomach. The limiting ridge of the forestomach was thickened and irregular mucosal thickenings in the forestomach and/or the glandular stomach. Of high dose males and females. Occasional these alterations were also seen in other groups including controls.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

No effects were detected except lesions of the forestomach and glandular stomach (see Table below) and renal changes in week 105 in high dose groups.
Lesions of the forestomach and glandular stomach:
a)Focal papillary epithelial hyperplasia and focal hyperkeratosis in the forestomach
b) chronic atrophic gastritis, focal ulcerations and glandular hyperplasia in the glandular stomach

Renal changes:
The incidence and degree of renal papillary necrosis was significantly increased in males and females in week 105 (not in week 53 and 79). These changes were located at the tip of the papilla and were characterized by patchy necrosis of interstitial cells, capillaries and loops of Henle.

Histopathological findings: neoplastic:

no effects observed

Description (incidence and severity):

Tumour incidences
There was no treatment related increase in any tumour type. In contrast, the total number of tumours and the number of tumour-bearing rats was lower in high dose males compared to controls.

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

15 mg/kg bw/day (actual dose received)

Sex:

male

Basis for effect level:

other: concentration in drinking water: 0.026%

2

Dose descriptor:

NOAEL

Effect level:

21 mg/kg bw/day (actual dose received)

Sex:

female

Basis for effect level:

other: concentration in drinking water: 0.026%

3

Dose descriptor:

LOAEL

Effect level:

82 mg/kg bw/day (actual dose received)

Sex:

male

Basis for effect level:

other: local effects in the stomach; concentration in drinking water: 0.19%

4

Dose descriptor:

LOAEL

Effect level:

109 mg/kg bw/day (actual dose received)

Sex:

female

Basis for effect level:

other: Local effects in the stomach; concentration in drinking water: 0.19%

Target system / organ toxicity

Critical effects observed:

yes

Lowest effective dose / conc.:

82 mg/kg bw/day (actual dose received)

System:

gastrointestinal tract

Organ:

stomach

Treatment related:

yes

Dose response relationship:

not specified

Relevant for humans:

not specified

Any other information on results incl. tables

Incidence of lesions in the forestomach and glandular stomach of rats
Type of lesion	Males, dose in mg/kg bw/d				Females, dose in mg/kg bw/d
	0	5	25	125	0	5	25	125
	Week 53
Number of rats examined	9	10	10	10	10	10	10	9
Forestomach
Focal papillary epithelial hyperplasia	0	0	0	7	0	0	0	5
Glandular stomach
Chronic atrophic gastritis	0	0	0	10***	0	0	0	9***
Focal ulceration	0	0	0	3	0	0	0	1
	Week 79
Number of rats examined	10	10	10	10	10	9	10	9
Forestomach
Focal papillary epithelial hyperplasia	2	1	1	8	1	0	1	9
Glandular stomach
Chronic atrophic gastritis	0	0	0	10***	0	0	0	10***
Focal ulceration	0	0	0	2	0	0	0	0
	Week 105
Number of rats examined	47	45	44	47	48	49	47	48
Forestomach
Focal papillary epithelial hyperplasia	1	2	1	45***	1	0	2	45***
Focal hyperkeratosis	2	6	4	24***	3	5	3	33***
Focal ulceration	1	1	1	8	0	0	2	5
Glandular stomach
Chronic atrophic gastritis	0	0	0	46***	0	0	0	48***
Focal ulceration	0	0	0	11***	0	0	0	10***
Glandular hyperplasia	0	1	0	20***	0	0	0	13***
*** : p<0.001

Applicant's summary and conclusion

Conclusions:

Oral exposure via the drinking water induced local effects in the stomach of rats at a concentration of 0.19% corresponding to 82 mg/kg bw/d in males and 109 mg/kg bw/d in females; the NOAEC is 0.026% corresponding to 15 mg/kg bw/d in males and 21 mg/kg bw/d in females.

Executive summary:

The study is comparable to OECD Guideline 453.

In a combined chronic toxicity/carcinogenicity study 70 male and 70 female Wistar rats per dose (subgroups of 10 rats/sex/dose killed 12 or 18 months after start of exposure) received via the drinking water 0, 1.2, 15, 82 mg/kg bw/day (males) or 0, 1.8, 21, 109 mg/kg bw/day (females) for 105 weeks (concentration: 0, 20, 260, or 1900 mg/L or 0, 0.002, 0.026, 0.19%).At the high dose body weight gain was decreased in males & females and food and water consumption decreased. Other parameters (except pathology) were not altered.Pathological alterations in the kidney like the renal papillary necrosis detected in high dose males and females is discussed as an effect of the reduced water intake and is indirectly treatment related. Treatment related lesions were detected in the forestomach (focal papillary epithelial hyperplasia, ulceration and hyperkeratosis) and the glandular stomach (chronic atrophic gastritis, ulceration and hyperplasia) of males and females in the high dose group. No gastric tumours were induced. This study did not provide any evidence of carcinogenicity in rats after oral administration of formaldehyde (see Section 7.7).

Conclusion: Oral exposure via the drinking water induced local effects in the stomach of rats at a concentration of 0.19% corresponding to 82 mg/kg bw/d in males and 109 mg/kg bw/d in females; the NOAEC is 0.026% corresponding to 15 mg/kg bw/d in males and 21 mg/kg bw/d in females.