Chlorine dioxide

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

No data

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Justification for type of information:

Considering the metabolism pathway of chlorine dioxyde which is likely to undergo rapid redox reactions within biological tissues rather than to be absorbed as parent compound, this study can be used for the assessment of ClO2.

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

publication

Title:

Developmental toxicity of sodium chlorite in the rabbit.

Author:

Harrington, R.M.; Romano, R.R. and Irvine, L.

Year:

1995

Bibliographic source:

Journal of the American College of Toxicology, 14(2):108-118

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

EPA OPP 83-3 (Prenatal Developmental Toxicity Study)

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Sodium chlorite

EC Number:

231-836-6

EC Name:

Sodium chlorite

Cas Number:

7758-19-2

Molecular formula:

ClHO2.Na

IUPAC Name:

sodium chlorite

Details on test material:

- Name of test material (as cited in study report): Sodium chlorite (NaClO2)
- Analytical purity: 80.6 %
- Impurities: Sodium chlorite (NaCl, 10.4 %)
Sodium chlorate (NaClO3, 2.0 %)
Water (H2O, 5.0 %)
The remaining 2 % of the test material consisted of sodium hydroxyde (NaOH), sodium carbonate (Na2CO3), and sodium sulfate (Na2SO4).
- Source: Olin Corporation (Niagara Falls, NY, U.S.A.)

Test animals

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS: Time-mated females
- Source: Interfauna UK Ltd. (Huntington, Cambridge, UK)
- Age at study initiation: 4–5 months old
- Weight at study initiation: 3–4 kg (at mating)
- Housing: individually, in grid bottomed metal cages suspended over paper lined trays
- Diet: SQC Rabbit Standard (Special Diets Services Limited, Witham, Essex, U.K.) ad libitum
- Water: tap water (purified by reverse osmosis) ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature: 18-22 °C
- Humidity: 42-59 %
- Air changes: no data
- Photoperiod: 12 hrs dark / 12 hrs light

Administration / exposure

Route of administration:

oral: drinking water

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: the test article was formulated for dosing by dissolution in the drinking water. Separate formulations were prepared daily for each dose level.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analysis of samples of the formulations used in this study were assessed for concentration on the first and last days of dosing to determine accuracy of preparation.

Details on mating procedure:

- Impregnation procedure: purchased timed pregnant

Duration of treatment / exposure:

Duration of treatment: Days 7-19 (post mating)

Frequency of treatment:

Continuously

Duration of test:

Until day 28 of pregnancy

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 200, 600 and 1200 mg/L
Basis:
nominal in water

No. of animals per sex per dose:

16 females/dose. One additional female was allocated to dose groups 200 and 600 ppm on day 7 of pregnancy as there was one female in each of these group showing poor clinical condition which may have necessitated sacrifice.

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: the dose levels were selected following examination of results of a preliminary maximum tolerated dose study in non-mated female rabbits. The dose levels were selected to cover a range between one expected to elicit minimal maternal effects and a no observable effect level.
- Rationale for animal assignment: randomisation procedure based on bodyweight

Examinations

Maternal examinations:

CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily from day 3 of pregnancy until necropsy.

BODY WEIGHT: Yes
- Time schedule for examinations: day 0, daily from day 3 to day 22 and on days 25 and 28 of pregnancy.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- The amount of food was recorded every 2 days from day 3 to day 27 and over 1 day from day 27 to day 28 of pregnancy.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
Water consumption was measured daily from day 3 to day 22 of pregnancy.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 28
- Organs examined: major organs

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of live and dead implantations: Yes

Fetal examinations:

GENERAL
Number of live foetuses, number of dead foetuses, foetal weight, external abnormalities.

SKELETAL
The bones were identified and examined for normality with respect to shape, size and the extent of ossification.

SOFT TISSUE
Foetuses were briefly fixed in alcohol prior to being skinned and dissected. The brain, eyes, palate and major organs and blood vessels in the thorax and abdomen were examined. The sex of the foetuses, as assessed from the appearance of the internal genitalia were recorded.

Statistics:

Analysis of variance; Student's t test; Kruskal-Wallis test; Dunn's multiple comparison test.

Indices:

No data

Historical control data:

No data

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
Two rabbits were sacrificed in extremis during the study because of a general deterioratio of clinical condition that was not considered to be related to treatment. One of the rabbits was from the control group and the other was from the 600 ppm group. There were no mortalities at the highest dose level of 1200 ppm. The only clinical observation was a dose-related increase in the incidence of rabbits with reduced fecal production during the dosing period, which coincided with the observed reduction in food consumption.
Because of palatability, there were significant decreases in water consumption. At 200 ppm, water consumption was significantly reduced only during days 16-20 of pregnancy. Intake of NaClO2 averaged 13.0 mg/kg bw/d (equivalent to 9.7 mg/kg bw/d ClO2-) during the treatment period. At 600 ppm, there was a significant decrease in water consumption for the entire dosing period. The mean NaClO2 intake at 600 ppm during the dosing period was 35.5 mg/kg bw/d (equivalent to 2.5 mg/kg bw/d ClO2-). At 1200 ppm, mean water consumption was reduced by > 50 % at the onset of dosing. Water consumption increased by remained significantly lower than that of the control group throughout the entire dosing period. The average NaClO2 intake at 1200 ppm was 53.1 mg/kg bw/d (equivalent to 39.6 mg/kg bw/d ClO2-).
There was no significant reduction of food consumption at 200 ppm. At 600 ppm, ther was a significant reduction in food consumption during days 7-11 of pregnancy (p < 0.001). For the remainder of pregnancy, food consumption was generally similar to that of the control group. At 1200 ppm, food consumption was significantly (p < 0.001) lower than that in the control group for the remainder of the dosing period; food consumption increased the levels greater than that for the control group.
There were no statistically significant changes in absolute body weights in any treatment group throughout the study. At 200 and 600 ppm, there were no significant differences from the controls in body weight gain. At 1200 ppm, animals lost a significant amount of weight at the onset of dosing (days 7-11). This was only a transient loss since these animals gained more weight during the remainder of the pregnancy that did the control group.
There were no treatment-related abnormalities observed at maternal necropsy. Pregnancy incidence was slightly lower than expected in the 200- and 600-ppm groups but was within expected limits in the control and the 1200-ppm groups.

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

1 200 mg/L drinking water

Basis for effect level:

other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
The mean numbers of corpora lutea, implantations, and live fetuses were similar in all groups. The sex distribution of the live fetuses was similar in all groups with near equal proportions of male and female fetuses.
Mean fetal weight was similar between the control group and the group treated at 200 ppm. Fetal weights were marginally lower than control values at 600 and 1200 ppm. Since fetal weights at 1200 ppm were greater that at 600 ppm, these slightly lower mean fetal weights may not have been related to treatment. Mean gravid uterus weight was similar in the control group and the group treated at 600 and 1200 ppm. It was slightly higher in the group treated at 200 ppm where there were slightly higher mean numbers of live fetuses in utero. Differences from the control group were not statistically significant.
At fetal examination, the major and minor external/visceral abnormalities observed were of a type and incidence that occur spontaneously in this strain of rabbit. There were no dose-related abnormalities. Most of the major and minor visceral abnormalities were associated with the aortic arch, a particularly labile region during organogenesis in this strain of rabbit. The minor aortic arch abnormalities were mainly either additional minor blood vessels or abnormal origin of the common carotid artery. The major aortic arch abnormalities involved severe dilations or constrictions of the great vessels.
There were no treatment-related skeletal abnormalities. The higher incidence of major skeletal abnormalities in the 600 ppm in comparison with the control was considered coincidental. The abnormalities were of a type that occur spontaneously in this strain of rabbit, and there were no major skeletal abnormalities at the highest dose tested, which achieved a higher NaClO2 intake throughout the exposure period. At 600 and 1200 ppm there was an increase in both minor skeletal abnormalities and skeletal variants related to incomplete ossification of certain bones such as the pubis and sternebrae. This was not unexpected since the fetuses in these groups were slightly lower in body weight. The incidences of minor skeletal abnormalities and skeletal variants at 200 ppm was similar to that of the controls.

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

1 200 mg/L drinking water

Basis for effect level:

other: embryotoxicity

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

Table 7.8.1/1: Mean maternal body weight and body weight gain of rabbits exposed to NaClO2 during pregnancy

NaClO2 concentration (ppm)		Control	200	600	1200
n		13	13	12	14
Body weight (mg) ± SD on Day 0 of pregancy		3.59 ± 0.26	3.61 ± 0.24	3.53 ± 0.25	3.61 ± 0.21
Body weight gain (kg) ± SD on indicated day of pregnancy	0-7	0.10 ± 0.13	0.05 ± 0.11	0.06 ± 0.13	0.09 ± 0.11
	7-11	0.09 ± 0.10	0.05 ± 0.11	0.02 ± 0.08	-0.16 ± 0.11*
	11-15	0.14 ± 0.06	0.16 ± 0.04	0.15 ± 0.08	0.20 ± 0.11
	15-19	0.06 ± 0.09	0.04 ± 0.09	0.05 ± 0.05	0.08 ± 0.05
	19-22	0.05 ± 0.03	0.04 ± 0.05	0.03 ± 0.04	0.13 ± 0.07*
	22-26	0.14 ± 0.06	0.16 ± 0.08	0.16 ± 0.07	0.14 ± 0.05

* significantly different from control p< 0.01: Student's test  

Table 7.8.1/2: Reproductive parameters in rabbits exposed to NaClO₂during pregnancy

NaClO2 concentration (ppm)	Control	200	600	1200
No. in group on Day 28	15	17	16	16
No. pregnant on Day 28	13	13	12	14
Mean no. of corpora lutea ± SD	11.9 ± 2.3	11.9 ± 1.9	12.8 ± 2.3	12.1 ± 2.7
Mean no. of implantation ± SD	10.7 ± 2.2	10.8 ± 1.8	10.7 ± 2.4	10.1 ± 2.2
Mean no. of live fetuses ± SD	8.5 ± 2.9	9.6 ± 1.9	9.0 ± 2.6	8.9 ± 2.4
Mean preimplantation loss (%)	10.4	8.4	16.1	14.9
Mean postimplantation loss (%)	21.3	11.1	15.4	12.6
Sex ratio (M:F)	55:45	41:59	48:52	52:48

 

Table 7.8.1/3: Mean fetal and gravid uterus weights in rabbits exposed to NaClO2 during pregnancy

NaClO2 concentration (ppm)	Control	200	600	1200
n	13	13	12	14
Mean male fetuses weight (g)	35.2 ± 4.9	36.7 ± 4.7	32.2 ± 4.7	33.5 ± 3.4
Mean female fetuses weight (g)	35.1 ± 4.2	35.3 ± 3.6	33.1 ± 3.2*	33.4 ± 3.4
Mean fetuses weight (g)	35.0 ± 4.2	35.8 ± 3.7	33.1 ± 2.6	33.2 ± 3.1
Gravid uterus weight (g)	494.5 ± 132.5	553.7 ± 69.7	480.4 ± 112.1	471.2 ± 105.5

* n = 11 for this value

Table 7.8.1/4: Incidences of fetal abnormalities in rabbits exposed to NaClO2 during pregnancy

NaClO2 concentration (ppm)	Control	200	600	1200
Total no. of fetuses (litters) examined	111 (13)	125 (13)	108 (12)	124 (14)
External and visceral examination : - No. with minor abnormalities only - Mean % with minor abnormalities only - No. of major abnormalities - Mean % with major abnormalities	33 (9) 28.6 2 (1) 1.5	29 (9) 22.8 1 (1) 0.5	36 (11) 32.2 4 (3) 6.6	32 (11) 26.8 3 (3) 2.9
Skeletal examination : - No. with minor abnormalities only - Mean % with minor abnormalities only - No. of major abnormalities - Mean % with major abnormalities - No. with any major abnormalities - Mean % with any major abnormalities	10 (4) 7.7 0 (0) 0.0 2 (1) 1.5	9 (7) 6.3 1 (1) 0.8 2 (2) 1.3	12 (6) 14.2 4 (4) 5.4* 5 (4) 7.5	18 (10) 13.9 0 (0) 0.0 3 (3) 2.9

Major abnormalities:

- 0 ppm x aortic arch abnormalities

- 200 ppm x aortic arch abnormalities, 1 x scoliosis

- 600 ppm x spinal dysmorphogenesis, 1 x spina bifida, 1 + microphtalmia, 1 x fused ribs

- 1200 ppm 1 x cleft plate, 1 x aortic arch abnormalities, 1 x umbilical hernia

* Significantly different from the control: p < 0.01: Dunn’s multiple comparison test

Applicant's summary and conclusion

Conclusions:

The NOAEL for both maternal and developmental effects in this study was determined to be 1200 mg/L, the highest dose tested corresponding to 44 mg chlorite/kg bw/day.

Executive summary:

In a developmental toxicity study conducted similarly to EPA OPP 83-3 guideline, Sodium chlorite (80.58 % pure) was administered to 16 or 17 females New Zealand White rabbits/dose in drinking water at dose levels of 0, 200, 600 and 1200 mg/L (0, 9.7, 26.5 and 39.6 mg chlorite/kg bw/day) from days 7 through 19 of gestation. Dams were sacrificed on gestation day 28.

Oral administration of sodium chlorite during organogenesis at 600 and 1200 mg/L elicited dose-related reductions in maternal water with consequent reductions in maternal food consumption, production of fecal pellets and body weight gain.

Despite the maternal effects there was no evidence of embryo-lethality or teratogenicity. Although the number and mean percentage of major external and visceral and skeletal abnormalities were increased in the 600 and 1200 mg/L groups (external/visceral: 6.6 and 2.9%, respectively, vs. 1.5% in controls; skeletal: 5.4 and 0%, respectively, vs. 0% in controls), the authors did not consider these to be treatment-related adverse effects. Mean fetal weights in the 600 and 1200 mg/L groups were slightly decreased (<9%, relative to controls). In the 600 and 1200 mg/L groups, the incidence of minor skeletal abnormalities (13.9 and 14.2% for the 600 and 1200 mg/L groups, respectively, vs. 7.7% in controls) and skeletal variants related to incomplete fetal bone ossification was higher than for controls. The authors state in their discussion that these alterations in fetal body weight and delayed ossification indicate embryonic growth retardation. Decreases in maternal food and water consumption and body weight gain may be responsible, at least in part, for some of the fetal effects.

Sodium chlorite was not considered to have any effect on maternal toxicity or reproductive development.

The NOAEL for both maternal and developmental effects in this study was determined to be 1200 mg/L, the highest dose tested.

Considering the metabolism pathway of chlorine dioxyde which is likely to undergo rapid redox reactions within biological tissues rather than to be absorbed as parent compound, this study can be used for the assessment of ClO₂.

However correction of doses should be done using the metabolism percentage of ClO₂into ClO_{2 -}(Abdel Rhaman, 1980a). Using a percentage of 11%, adjusted doses are the following: 0; 88.1; 240; 360 mg/kg bw/d.

Therefore the NOAEL for both maternal and developmental toxicity of ClO₂ in this study is 360 mg/kg bw/d. As ClO₂has corrosive properties at doses as low as 40 mg/kg bw (Tos, 1996), effects that will be observed will be linked to corrosive properties.