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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
sub-chronic toxicity: oral
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

Data source

Reference
Reference Type:
publication
Title:
Ninety-day toxicity study of sodium monochloroacetate in Sprague-Dawley rats
Author:
Daniel FB, Robinson M, Stober JA, Page NP, Olson GR
Year:
1991
Bibliographic source:
Toxicol. 67, 171-185

Materials and methods

Test guidelineopen allclose all
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Qualifier:
equivalent or similar to guideline
Guideline:
EU Method B.26 (Sub-Chronic Oral Toxicity Test: Repeated Dose 90-Day Oral Toxicity Study in Rodents)
Qualifier:
equivalent or similar to guideline
Guideline:
EPA OPPTS 870.3100 (90-Day Oral Toxicity in Rodents)
Principles of method if other than guideline:
Compliant to current OECD 408 guideline except for: no platelet counts and blood clotting potential, sodium, potassium, urea, total protein and albumin, and only two enzymes indicative of hepatocellular effects measured. No functional observation, arena or ophthalmoscopic tests included. No epididymides and uterus weight included. No spinal cord and eyes examined histopathologically.
GLP compliance:
not specified
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Sodium chloroacetate
EC Number:
223-498-3
EC Name:
Sodium chloroacetate
Cas Number:
3926-62-3
Molecular formula:
C2H2ClNaO2
IUPAC Name:
sodium 2-chloroacetate
Details on test material:
- Name of test material (as cited in study report): sodium monochloroacetate
- Impurities (identity and concentrations): no impurities detected (analyzed by gas chromoatography and mass spectrometry
No further details specified.

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories (Kingston, USA)
- Age at study initiation: approximately 12 weeks
- Weight at study initiation: males: 471-493 grams (mean), females: 275-288 grams (mean)
- Fasting period before study: no
- Housing: 2 per cage in elevated wire mesh cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: approximately 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-22
- Humidity (%): 40-60
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: Not specified

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: distilled water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
Monochloroacetic acid (MCA) was diluted in distilled water. The dosing solutions of sodium monochloroacetate (SMCA) were prepared weekly from a stock solution with dilution to the appropriate concentration with distilled water. The solutions were adjusted with 1.0 N sodium hydroxide in order to achieve a pH of 7.0-7.5.

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Stability tests (gas chromatography) determined that the MCA in these preparations was stable at least for 10 days.
No details provided on accuracy and homogeneity of preparations.
Duration of treatment / exposure:
90 days
Frequency of treatment:
daily
Doses / concentrationsopen allclose all
Dose / conc.:
15 mg/kg bw/day (actual dose received)
Dose / conc.:
30 mg/kg bw/day (actual dose received)
Dose / conc.:
60 mg/kg bw/day (actual dose received)
Dose / conc.:
120 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Not specified, but likely based on LD50 values of MCA.
Positive control:
Not applicable.

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS/DETAILED CLINICAL OBSERVATIONS: All rats were observed for physiological and behavioral responses and for mortality
or morbidity.

OPHTHALMOSCOPY: no

BODY WEIGHT: Body weights were recorded at the initiation of dosing, and twice weekly throughout the study. A final body weight, following an 18 h fast, was recorded at necropsy.

FOOD AND WATER CONSUMPTION: Water and food consumption was determined at the beginning, middle and end of each week.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: end of treatment
- Anaesthetic used for blood collection: Yes (identity): pentobarbital (60 mg/kg i.p.)
- Animals fasted: Yes
- How many animals: all surviving animals
- Parameters checked: red blood cell count (RBC), white blood cell count (WBC), hemoglobin, hematocrit, reticulocyte count, and mean corpuscular volume (MCV). A differential leukocyte count was performed for segmented neutrophils, lymphocytes, monocytes and eosinophils.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood:end of treatment
- Animals fasted: Yes
- How many animals: all surviving animals
- Parameters checked : blood urea nitrogen (BUN), blood calcium (BCAL), inorganic phosphorous (PO4), creatinine, total cholesterol, glucose, lactate dehydrogenase (LDH), alanine aminotransferase (AL T), and aspartate aminotransferase (AST).

URINALYSIS: No.

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes:
The necropsy included gross examination of the external surfaces, all orifices, external surface of the brain, cervical tissues, all organs,
and the cranial, thoracic, abdominal, and pelvic cavities. Tissues from selected organs, including the brain (with the stem), liver, spleen, lung with the lower half of the trachea, thymus, kidneys, adrenal glands, heart and gonads of each animal were weighed and grossly examined at necropsy.

HISTOPATHOLOGY: Yes:
In addition to tissues grossly examined at necropsy, skin, mandibular and mesenteric lymph nodes, mammary gland, thigh muscle, sciatic nerve, sternebrae, thymus, salivary gland, tongue, esophagus, stomach, duodenum, jejunum, ileum, colon, cecum, rectum, pancreas, urinary bladder, seminal vesicles, prostate, uterus, nasal cavity and turbinates, pituitary, preputial or clitoral gland, Zymbal's gland, aorta, thyroid, parathyroids and any gross lesions.
Other examinations:
None.
Statistics:
The standard statistical procedure was a one-factor analysis of variance (ANOVA) to analyze normally-distributed measures for a dose-related response (or effect). Males and females were considered separately in all statistical analyses and the parameters analyzed were: body weights, organ weights, organ-to-body weight ratios, water and food consumption, hematology, and clinical chemistry measurements. The differences between the control and treatment groups were analyzed pairwise using the Student t-test at an adjusted significance level to control the false positive rate (overall alpha = 0.05). Due to the high variability of some of the clinical chemistry measures, a non parametric analysis of variance, procedure i.e. the Kruskal-Wallis test, was also employed to determine differences among the dose groups and to pairwise compare each dose group to the control group. The incidence of microscopic lesions were tested for a dose related trend using the exact test for trend, which is a generalization of Fisher's Exact Test. Pairwise comparisons of each dose group vs. the control (one-tail test) were made using Fisher's Exact Test. Analysis of gross pathology diagnosis were limited to descriptive statistics.

Results and discussion

Results of examinations

Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
not examined
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:
CLINICAL SIGNS AND MORTALITY
Thirteen (13) rats died during the study; all but two were in the high-dose group. The two deaths in the lower dose groups were both males, one at 15 and another at 60 mglkg per day. None of the control rats died during the experiment. It is apparent that 120 mg/kg per day is acutely toxic as over half of the deaths (7/11; 4 male and 3 female) occurred within the first 3 days of treatment. The other 4 deaths in the 120 mg/kg per day male group occurred between the 14th and 90th days of treatment. This may indicate a form of tolerance to continued treatment.
No details on clinical signs given.

FOOD AND WATER INTAKE
No significant (P< 0.05) differences from controls were found in food or water consumption in any group of either sex. For the 120 mg/kg per day dose food consumption in the males was lowered and water consumption was higher for both sexes but the data was not statistically evaluated due to the small number of survivors.

HAEMATOLOGY (no statistical analysis performed in the male 120 mg/kg per day group due to excess mortality)
Statistically significant increases were observed in WBC at dose levels of 30 mg/kg per day and higher and % segmented neutrophils at 60 mg/kg per day and higher in females; the latter finding was not dose-related. The percentage monocytes was significantly decreased in females of the 15 mg/kg group only, and increased in males at 15, 30 and 60 mg/kg per day; however, no dose-response was observed.

CLINICAL CHEMISTRY
Statistically significant differences (no statistical analysis performed in the male 120 mg/kg per day group due to excess mortality) were observed in the following clinical chemistry parameters:
(a) increased BUN levels in females at 120 mg/kg per day and in males at 15 and 30 mg/kg per day but not in males at 60 mg/kg bw, thus a lack of dose-response
(b) increased creatinine levels in females at 30 and 60 mg/kg per day but not at 120 mg/kg bw, thus a lack of dose-response
(c) increased creatinine levels in males at 15, 30 and 60 mg/kg bw; however, without a dose-response
(d) increased PO4 in females at 120 mg/kg per day
(e) increases in BCAL in males at 15 and 30 mg/kg per day, but not at 60 mg/kg bw, thus a dose-response is lacking
(f) increases in serum ALT in females at 120 mg/kg per day and in males at 15 and 30 mg/kg per day, but not at 60 mg/kg bw, thus a dose-response is missing.
(g) Increases in AST in females at 120 mg/kg per day
Overall, increases were seen in females of the 120 mg/kg bw group and changes in males were lacking a dose-response relationship. Increases in males of the 120 mg/kg bw group were not statistically evaluated as only 2 males were left.

ORGAN WEIGHTS
There were no apparent dose-response related differences between treated and control groups in the absolute weights of the brain, gonads, heart, kidneys, lungs, spleen, thymus and adrenal glands for either sex. A significant increase in liver and kidney weights for the 120 mg/kg per day female group was recorded.
Statistically significant increases were seen in the relative liver and kidney weight in females at 60 and 120 mg/kg per day, and in males at 60 mg/kg bw per day. Statistical analyses were not performed in the 120 mg/kg male group because of the poor survival; the mean relative liver and kidney weight of the two rats left was higher than at 60 mg/kg per day.

GROSS PATHOLOGY
Reddened lungs (possibly a postmortem effect) in animals that died early (days 1-3) in the study. Pale, yellow-colored livers were seen in male rats at all dose levels and in female rats at the two highest dose levels. This change was particularly evident in the high-dose male group as well as in two animals that died at 18 and 53 days. In contrast, only 2 out of the 7 surviving females at 120 mg/kg per day showed these liver colorations. The 3 females that died before day 3 were not examined.

HISTOPATHOLOGY: NON-NEOPLASTIC
Increases were observed in the incidences of lesions in the kidneys, liver, heart and spleen in treated rats compared to controls. Due to low survival rates, the animals in the high dose group (120 mg/kg per day) were excluded from the statistical analyses of the microscopic lesions data.
- Chronic nephropathy was only seen in male rats, controls included, but was statistically significantly increased at 60 mg/kg per day.
- Hepatocytic vacuolated foci (lipidosis) at the base of the liver lobes and in the subcapsular region were observed in male rats. The trend analysis indicated a significant increase in the incidence of hepatocytic vacuolated foci with increasing dose (P = 0.013) in male rats, but no significant increase was found at any individual dose level (P > 0.05). Some other liver lesions (periportal fatty changes and necrosis) were also observed in males of the high-dose group that died during the experiment. Since the livers of the animals that died early in the study were not routinely examined, the true incidence of liver lesions in the high-dose group is not known.
- Chronic inflammation of the heart, a commonly seen incidental lesion in rats at this age, was seen in all male and female groups, controls included, and no statistically significant increase in the incidences was observed.
- Inflammation of lung interstitium and perivascular areas was seen in both treated and control male and female rats and was considered to be an incidental finding not related to SMCA. Congestion and hemorrhage was consistently noted in males of the 60 and 120 mg/kg bw groups that were found dead during the course of the experiment.
- The incidence of splenic pigments, brown and localized in splenic macrophages, was significantly increased in males of the 60 mg/kg group, and was also present in the 2 surviving males of the 120 mg/kg group.

Effect levels

Dose descriptor:
NOAEL
Effect level:
30 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Based on increased relative liver and kidney weight and increased incidence of histopathological changes in these organs

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

See also attached background material on tables with results.

Applicant's summary and conclusion

Conclusions:
Rats were exposed to SMCA for 90-days by oral gavage; the NOAEL was set at 30 mg/kg bw.
Executive summary:

Male and female Sprague-Dawley rats were administered the sodium salt of monochloroacetic acid (SMCA) by oral gavage for a period of 90 consecutive days. Dosage levels of 15, 30, 60 or 120 mg/kg per day were employed. SMCA clearly induced toxicity in both females and males, with the greatest severity in the male animals. Both the liver and kidneys were identified as target organs. At 120 mg/kg per day, 30% of females and 80% of the males died, most within the first 2 days of treatment. Hemorrhagic and congested lungs (possibly a postmortem change) were seen in the early deaths (1 -3 days) whereas liver lesions were observed in later deaths. In addition, there was nephrotoxicity as evidenced by elevated creatinine, blood calcium (BCAL), and blood urea nitrogen (BUN) levels. Hepatotoxicity was indicated by increases in the serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Both liver and kidneys showed statistically significant increased organ-to-body weight ratios, in females at 60 and 120 mg/kg bw, and in males at 60 mg/kg bw (the two animals of the 120 mg/kg bw group left, also showed a higher mean relative weight) . Microscopic examination revealed a significant increase in chronic renal nephropathy and increased splenic pigmentation at 60 mg/kg per day in the males. Based on the observation of toxicity at all treatment levels, the NOAEL was set at 30 mg/kg bw.