Chloramide

Carcinogenicity

Description of key information

Two carcinogenic studies - with rat and mice

Key value for chemical safety assessment

Carcinogenicity: via oral route

Link to relevant study records

Reference

Endpoint:

carcinogenicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 29 november 1984 To 7 december 1986

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 451 (Carcinogenicity Studies)

Deviations:

yes

Remarks:

Temperature and relative humidity were deviated. Individual weights of animals at the start of the test were not clearly reported. No histopathological examination of lacrimal glan, peripheral nerve and spinal cord were performed.

GLP compliance:

yes

Species:

mouse

Strain:

B6C3F1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Frederick Cancer Research Facility (Frederick, MD)
- Age at study initiation: 5 to 6 weeks of age
- Weight at study initiation: no data
- Fasting period before study: no
- Housing: one per cage
- Diet : ad libitum
- Water : ad libitum
- Acclimation period: 12 to 15 days

ENVIRONMENTAL CONDITIONS
- Temperature : 15 - 28 °C
- Humidity : 21 -73 %
- Air changes : 10 changes/hour
- Photoperiod: 12 hrs dark / 12 hrs light

Route of administration:

oral: drinking water

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Dose formulations for chloraminated water were prepared from a stock solution of buffered sodium hypochlorite. This stock solution was prepared by bubbling chlorine gas into charcoal filtered, deionized water until the solution obtained a deep greenish-yellow color. The concentration of available atomic chlorine in the solution was determined by titration with 0.0028 N ferrous ammonium sulfate (FAS) using N,N-diethyl-p-phenyldiamine (DPD) as an indicator and adding an equivalent weight of sodium hydroxide. The solution was then buffered to approximately pH 9 with bicarbonate-carbonate buffer solution. The buffers were 0.045 M in carbonate.
Chloraminated dose formulations were prepared by mixing the appropriate volume of the buffered sodium hypochlorite stock solution with sodium chloride and bicarbonate-carbonate buffer solutions, and adding the resulting solution to a dilute ammonium hydroxide solution to generate monochloramine. The final solution was 0.022 M in carbonate and 0.035 M in sodium at pH 9. Dose concentrations were expressed as ppm of chloramine. Monochloramine and dichloramine concentrations were then confirmed by titration with FAS using potassium iodide as an indicator.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

For chloraminated water, dose levels of approximately 50, 100, and 200 ppm chloramine retained 95 % to 97 % of the original concentration after 24 hours and 91 % to 94 % after 48 hours.

Duration of treatment / exposure:

2 years

Frequency of treatment:

daily

Post exposure period:

No

Remarks:

Doses / Concentrations:
0, 50, 100, and 200 ppm
Basis:
nominal in water

Remarks:

Doses / Concentrations:
0, 4.9, 8.4, 14.9 mg/kg per day (male)
Basis:
other: Estimate of dose delivered as measured by water consumption - average for more than 52 weeks.

Remarks:

Doses / Concentrations:
0, 4.1, 7.7, 15.6 mg/kg per day (female)
Basis:
other: Estimate of dose delivered as measured by water consumption - average for more than 52 weeks.

No. of animals per sex per dose:

50 males and 50 females
Interim Evaluations: 10 males and 10 females

Control animals:

yes

Details on study design:

- Dose selection rationale:
The doses for the 2-year studies were based on data from unpublished studies of chloramine conducted at the Gulf South Research Institue (GSRI), New Iberia, Louisiana.
- Study design:
Groups of 70 animals of each sex were administered the tested substance in drinking water for 103 to 104 weeks. Control animals received buffered deionized water only. Ten animals per dose group were predesignated for interim evaluation (necropsy, organ weights, histopathology, and hematology) during week 15. Another 10 animals per dose group were predesignated for evaluation at 66 weeks.

Positive control:

None

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: No

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: once per week for the first 13 weeks of the studies and once per month thereafter

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: daily

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: 15-week and 66-week
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: 10

CLINICAL CHEMISTRY: No

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Other examinations:

No

Statistics:

Survival, Calculation of incidence, Analysis of tumor incidence were statistically analysed. Details are given in the field "Any other information on materials and methods incl. tables".

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

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

effects observed, treatment-related

Description (incidence and severity):

A dose-related decrease in water consumption by both sexes was evident in the first week and continued throughout the studies.

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY
The survival rates of mice receiving chloraminated water were not significantly different from those of controls. There were no clinical findings attributable to the consumption of chlorinated water.

BODY WEIGHT AND WEIGHT GAIN
There was a dose-relatd decrease in mean body weights of dosed male and female mice throughout most of ths studies. Mean body weights of high-dose males were 10 % to 22 % lower than those of controls after week 37, and body weights of high-dose female mice were 10 % to 35 % lower than controls after week 8. Mean body weights of high-dose male and female mice were 91 % and 84 % of the control values at 15 weeks. At the 66-week evaluation, the body weights of high-dose female mice were similar to controls, but the body weights of mid- and high-dose male mice were lower than controls.

FOOD CONSUMPTION
Feed consumption by dosed male mice was similar to controls throughout the studies. In females, mean feed consumption was similar in all treatment groups except the high-dose group, in which feed consumption was slghtly lower than in the other groups.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)
Administration of chloraminated water resulted in a dose-related decrease in water consumption by both sexes that was evident in the first week and continued throughout the studies. Within dose groups, estimated ingestion of chloramine by males was lower than that by females the first year, but higher than that by females the second year, except for the high-dose group.

HAEMATOLOGY
There were no alterations in hematologic parameters attributable to the consumption of chloraminated water at either the 15-week or 66-week interim evaluation.

ORGAN WEIGHTS
Although there were some statistically significant differences in organ weights and organ-weight-to-body-weight ratios between dosed and control groups, the differences were not biologically significant. Decreases in liver weights and increases in brain- or kidney-weight-to-body-weight ratios observed in high-dose mice at 15 or 66 weeks were related to the lower body weights in these groups.
GROSS PATHOLOGY

HISTOPATHOLOGY: NON-NEOPLASTIC
There were no nonneoplastic lesions in the mice in these 2-year studies that were clearly attributable to the consumption of chloraminated water.

HISTOPATHOLOGY: NEOPLASTIC (if applicable)
There were no neoplastic lesions in the mice in these 2-year studies that were clearly attributable to the consumption of chloraminated water.

Relevance of carcinogenic effects / potential:

There were no increases in the incidence of neoplasms or nonneoplastic lesions that could be clearly attributed to the consumption of chloraminated water.

Conclusions:

Under the conditions of these 2-year drinking water studies, there was no evidence of carcinogenic activity of chloramine water in male or female B6C3F1 mice receiving 50, 100, or 200 ppm.

Executive summary:

In these 2 -year studies, groups of 70 B6C3F1 mice of each sex were administered 50, 100, or 200 ppm chloramine in buffered, charcoal-filtered, deionized drinking water for 103 to 104 weeks. Control animals received buffered deionized water only. Ten animals per dose group were predesignated for interim evaluation (necropsy, organ weights, histopathology, and hematology) during week 15. Another 10 animals per dose group were redesignated for evaluation at 66 weeks. Survival at 2 years of mice receiving chloraminated water was similar to that of the controls. Mean body weights of dosed mice were lower than those of their respective controls groups. There was a dose-related decrease in water consumption by mice. Water consumption by high-dose mice was 42 % lower than controls for males and 40 % lower than controls for females. There were no neoplasms or nonneoplastic lesions in male or female mice that were clearly associated with the consumption of chloraminated water. Under the conditions of these 2-year drinking water studies, there was no evidence of carcinogenic activity of chloraminated water in male or female B6C3F1 mice receiving 50, 100, or 200 ppm.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Study duration:

chronic

Species:

other: mouse and rat

Quality of whole database:

These studies are GLP compliant and have klimish score 2.

Carcinogenicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Carcinogenicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Chloraminated drinking-water (predominantly in the form of monochloramine) was tested for carcinogenicity by oral administration in female and male mice and rats without demonstrating clear evidence of carcinogenic activity (NTP study). Therefore, no classification is proposed.

Additional information

Two reliable carcinogenic studies with rats and mice are available:

Mouse

 

Groups of 70 male and 70 female B6C3F1 mice, 5–6 weeks of age, were given chloraminated, deionized, charcoal-filtered drinking-water¹that contained 50, 100 or 200 ppm [mg/L] monochloramine expressed as available atomic chlorine or deionized, charcoalfiltered drinking-water (controls). The dose levels of chloramine were based on a previous study that indicated that 200 ppm was the maximum concentration that mice will drink. Interim sacrifices of 9–10 mice each were carried out at 14 or 15 and 66 weeks, leaving 50–51 mice of each sex that were killed after 103–104 weeks of exposure. At 2 years, survival of mice that received chloraminated drinking-water was not significantly different from that of controls. Within the first week of chloramine treatment, a dose-related decrease in drinking-water consumption was observed in both males and females, which continued throughout the study. There was also a dose-related decrease in the body weights of males and females administered chloramine, which occurred earlier and to a greater extent in females than in males. The chloraminated water decreased the body weight of female mice by approximately 10% (50 ppm), 15% (100 ppm) and 30% (200 ppm). No neoplastic or non-neoplastic lesions associated with the consumption of chloraminated drinking-water were observed in male or female mice (National Toxicology Program, 1992).

 

Rat

 

Groups of 70 male and 70 female Fischer 344/N rats, 5–6 weeks of age, were given chloraminated, deionized, charcoal-filtered drinking-water¹that contained 50, 100 or 200 ppm [mg/mL] monochloramine expressed as available atomic chlorine or deionized, charcoal-filtered drinking-water (controls). The dose levels of chloramine were based on a previous study that indicated that 200 ppm was the maximum concentration that rats will drink. Interim sacrifices of 9–10 rats each were carried out at 15 and 66 weeks, leaving 50–51 rats of each sex that were killed after 103–104 weeks of exposure. Survival of male and female rats that received chloraminated drinking-water was not significantly different from that of controls except for the low-dose male group in which survival was greater. Within the first week of chloramine administration, a dose-related decrease in drinking-water consumption by males and females was observed, which remained reduced throughout the study. Chloraminated water did not affect the body weight of female and male rats until weeks 97 and 101, at which time a 10% reduction relative to controls occurred in the high-dose female and male rats, respectively. The incidence of mononuclear-cell leukaemia was higher in female rats that received the high dose of chloramine than in controls: 8/50 (16%), 11/50 (22%), 15/50 (30%) and 16/50 (32%) for the 0-, 50-, 100- and 200-ppm chloramine-treated groups, respectively. When adjusted for intercurrent mortality (Kaplan-Meier estimated tumour incidence), the rate of leukaemia was 20.8, 29.0, 39.9 and 41.4% for the 0-, 50-, 100- and 200-ppm chloramine-treated groups, respectively. The results were analysed by the life table test (Haseman, 1984) for an overall dose–response trend, resulting in a p-value of 0.02. This was followed by pairwise comparison between the controls and the treated groups resulting in p-values of 0.280, 0.077 and 0.036 for the 50-, 100- and 200-ppm chloramine-treated groups, respectively. The authors reported that the incidence of leukaemia in the concurrent control group was lower than that in untreated historical controls (170/680 [25%], with a range of 14–36%). There was a marginal increase in spleenic histiocytic lymphoid hyperplasia in females (controls, 3/50; 50 ppm, 4/50; 100 ppm, 2/50; and 200 ppm, 6/50). Since this apparent increase lacked a dose–response relationship and was only marginally significant, it was not considered by the authors to be related to the chloraminated water. There were no other neoplastic or non-neoplastic lesions associated with the consumption of chloraminated drinking-water. The report concluded that there was no evidence for the carcinogenic activity of chloraminated drinking-water in male rats and equivocal evidence in female rats based on an increased incidence of mononuclear-cell leukaemia (National Toxicology Program, 1992).

Based on the studies described above, Monochloramine has been considered by IARC as not classifiable as to its carcinogenicity to humans (Group 3).

¹The chloramine formulations were prepared from buffered sodium hypochlorite (pH 9) stock solutions to which dilute ammonium hydroxide was added. The sodium hypochlorite stock solution was prepared by bubbling chlorine gas into charcoal-filtered, deionized water and was raised to pH 9 with sodium bicarbonate/carbonate.

Justification for selection of carcinogenicity via oral route endpoint:

Selected studies tested Chloraminated drinking-water for carcinogenicity in female and male mice and rats (Key studies).