Diethyl bis(2-hydroxyethyl)aminomethylphosphonate

Endpoint summary

Description of key information

NOAEL ≥ 500 mg/kg bw/day (highest dose tested) (eq. OECD 408, Key, Rel.2)

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

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

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Version / remarks:

adopted in 1981

GLP compliance:

not specified

Limit test:

no

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source of test material: Stauffer Chemical Company, Westport, USA

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Stability under test conditions: stable at room temperature

Species:

rat

Strain:

Sprague-Dawley

Details on species / strain selection:

Not reported

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Inc., Kingston, USA
- Age at study initiation: 6 weeks
- Housing: individually in suspended polycarbonate cages (20.3 x 17.8 x 17.8 cm) with hardwood chip bedding
- Diet: Purina Certified Rodent Chow 5002 ad libitum (except when fasted overnight prior to taking blood samples)
- Water: ad libitum
- Acclimation period: 2 weeks

Route of administration:

oral: gavage

Details on route of administration:

Not reported

Vehicle:

corn oil

Details on oral exposure:

- Dose volume: 5 mL/kg bw

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

No details reported

Duration of treatment / exposure:

13 weeks (control and test groups)

Frequency of treatment:

once daily, 7 days/week

Dose / conc.:

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

Dose / conc.:

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

Dose / conc.:

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

No. of animals per sex per dose:

22 rats per sex per dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: not reported
- Rationale for animal assignment (if not random): random
- Fasting period before blood sampling for clinical biochemistry: none
- Dose range finding studies: not reported

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: weekly

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Not specified
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION AND COMPOUND INTAKE: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: during quarantine, at study midpoint and study termination
- Animals fasted: Yes
- How many animals: 20 animals (10 rats per sex) for midpoint analysis; all 176 animals (88 rats per sex) during quarantine and at study termination
- Complete hematology profiles were determined

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at study midpoint and study termination
- Animals fasted: Yes
- How many animals: 20 animals (10 rats per sex) for midpoint analysis; all 176 animals (88 rats per sex) during quarantine and at study termination
- Complete clinical chemistry profiles (incl. plasma and erythrocyte cholinesterase assays) were determined

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

IMMUNOLOGY: No

OTHER:
-

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Other examinations:

Brain acetylcholinesterase activity was measured for 10 rats in each group. Absolute and relative organ weights of liver, kidneys, heart, gonads, brain and adrenals of all animals were determined.

Statistics:

Body weight, food consumption, absolute and relative organ weights, hematology and clinical chemistry were analyzed statistically by a one-way analysis of variance and Dunnett's t-test. Results between groups were considered statistically significant when p < 0.05.

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

Dehydration, diarrhea, skin abrasions, minor hair loss, chromodacryorrhea and chromorhinorrhea were observed. The incidences of these symptoms were similar in control and treated rats.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

Necropsy findings showed deaths to be dosing accidents related to gavage administration.

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):

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

100 mg/kg bw: relative kidney and liver weights were significantly increased in females
500 mg/kg bw: absolute and relative kidney and liver weights were increased in males and females
See table 1 in "any other information on results incl. tables" for details

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

100 mg/kg bw: hepatocellular hypertrophy (3/17 males and 1/20 females), increased eosinophilia of centrilobular hepatocytes (1/17 males)
500 mg/kg bw: hepatocellular hypertrophy (10/21 males and 6/16 females); increased eosinophilia in centrilobular hepatocytes (12/21 males); hepatocellular hypertrophy (48% of males and 38% of females); 40% increase in cross-sectional area of male livers compared to controls (see table 2 in "any other information on results incl. tables" for details)

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Details on results:

No treatment-related mortality and few signs of toxicity were noted during the study. The test material did not inhibit plasma, erythrocyte, or brain cholinesterase activities and treatment-related necropsy and microscopic alterations were restricted to the liver. Increased liver weights, hepatocellular hypertrophy, and eosinophilia of centrilobular hepatocytes were evident in 100 mg/kg females and in both sexes at 500 mg/kg. Morphometric analysis revealed a 40% increase in cross-sectional area of individual hepatocytes in 500 mg/kg males compared to controls. Morphologic evidence of hepatic necrosis or clinical evidence of liver dysfunction was not observed.

Key result

Dose descriptor:

NOAEL

Effect level:

>= 500 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Dose descriptor:

LOAEL

Effect level:

> 500 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Remarks on result:

not determinable due to absence of adverse toxic effects

Key result

Critical effects observed:

no

Table 1: Effect of test material on absolute and relative kidney and liver weights

		Liver weight*		Kidney weight*
Test material conc. (mg/kg bw)	Sex	Abs. (g)	Rel.	Abs. (g)	Rel.
0	M	16.05 ± 2.32	3.02 ± 0.30	2.89 ± 0.24	0.55 ± 0.05
	F	7.57 ± 0.96	2.70 ± 0.19	1.62 ± 0.18	0.60 ± 0.06
20	M	15.84 ± 2.07	2.95 ± 0.30	2.94 ± 0.29	0.55 ± 0.05
	F	7.41 ± 0.83	2.76 ± 0.16	1.68 ± 0.15	0.63 ± 0.06
100	M	16.19 ± 1.91	3.05 ± 0.31	3.05 ± 0.35	0.58 ± 0.06
	F	8.01 ± 1.01	2.93 ± 0.31**	1.84 ± 0.18	0.67 ± 0.08**
500	M	18.45 ± 2.99**	3.44 ± 0.32**	3.26 ± 0.45**	0.61 ± 0.07
	F	9.41 ± 1.80**	3.47 ± 0.24**	2.09 ± 0.24**	0.78 ± 0.07**

*: mean ± standard deviation (n = 16-21)

**: significantly different from mean of untreated control (0 mg/kg bw) for the same sex (p < 0.05)

Relative weight = organ weight/body weight x 100

Table 2: Comparison of cross-sectional area measured for individual liver cells in rats treated with test material and control rats

Treatment	Animal no.	Number of cells measured	Mean cross-sectional area	Group mean± SD
Control (corn oil)	1	50	494.4	424.2 ± 41.8
	2	50	425.8
	3	50	412.4
	4	50	387.0
	5	50	401.4
test material (500 mg/kg)	6	50	587.1	595.8* ± 35.6
	7	50	631.1
	8	50	568.3
	9	50	635.1
	10	50	557.6

*: p < 0.05

Conclusions:

In conclusion, test item treatment resulted in measurable effects upon rat liver and kidney. These effects were indicative of an adaptive rather than a toxic response (NOAEL = 500 mg/kg bw/day; highest dose tested). No evidence of a treatment-related alteration was found in rats given the lowest dose (NOEL = 20 mg/kg bw/day).

Executive summary:

The subchronic oral toxicity was determined using 22 rats/sex/group treated by gavage daily for three months at doses of 0, 20, 100 or 500 mg/kg bw/day.

No treatment-related changes were found with respect to body weights, food consumption, clinical signs or urinalysis. Certain hematological and clinical chemistry changes occurred in mid and/or high dose groups.

The hematology changes included reductions in red blood cell count, hemoglobin concentration and hematocrit, as well as altered total and differential white blood cell counts. Clinical chemistry changes were altered electrolyte (Ca and K) levels and increased albumin and total protein. Although these changes were statistically significant, they were mild and considered to be within the range of normal variation.

Liver changes noted at mid and/or high dose levels were reported. These included very slight hepatocellular hypertrophy (males and females), cytoplasmic eosinophilia of centrolobular hepatocytes (males) and increased relative liver weights (males and females). These changes were considered to be minimal, reversible, and not representative of degenerative processes; this interpretation is supported by the fact that liver function in treated rats appeared normal based on serum enzyme levels. These morphologic findings are compatible with an adaptive response.

Relative kidney weights were increased in high dose males and mid and high dose females. However, no gross or microscopic kidney changes were noted. Based on serum BUN and creatinine levels, kidney function appeared normal in all treated groups. The findings with respect to the kidneys were not considered indicative of degeneration.

 

In conclusion, test item treatment resulted in measurable effects upon rat liver and kidney. These effects were indicative of an adaptive rather than a toxic response (NOAEL = 500 mg/kg bw/day; highest dose tested). No evidence of a treatment-related alteration was found in rats given the lowest dose (NOEL = 20 mg/kg bw/day).

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

500 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Subchronic oral toxicity was determined using 22 rats/sex/group treated by gavage daily for three months at doses of 0, 20, 100 or 500 mg/kg bw/day.

No treatment-related changes were found with respect to body weights, food consumption, clinical signs or urinalysis. Certain hematological and clinical chemistry changes occurred in mid and/or high dose groups.

The hematology changes included reductions in red blood cell count, hemoglobin concentration and hematocrit, as well as altered total and differential white blood cell counts. Clinical chemistry changes were altered electrolyte (Ca and K) levels and increased albumin and total protein. Although these changes were statistically significant, they were mild and considered to be within the range of normal variation.

Liver changes noted at mid and/or high dose levels were reported. These included very slight hepatocellular hypertrophy (males and females), cytoplasmic eosinophilia of centrolobular hepatocytes (males) and increased relative liver weights (males and females). These changes were considered to be minimal, reversible, and not representative of degenerative processes; this interpretation is supported by the fact that liver function in treated rats appeared normal based on serum enzyme levels. These morphologic findings are compatible with an adaptive response.

Relative kidney weights were increased in high dose males and mid and high dose females. However, no gross or microscopic kidney changes were noted. Based on serum BUN and creatinine levels, kidney function appeared normal in all treated groups. The findings with respect to the kidneys were not considered indicative of degeneration.

 

In conclusion, test item treatment resulted in measurable effects upon rat liver and kidney. These effects were indicative of an adaptive rather than a toxic response (NOAEL = 500 mg/kg bw/day; highest dose tested). No evidence of a treatment-related alteration was found in rats given the lowest dose (NOEL = 20 mg/kg bw/day).

Justification for classification or non-classification

Harmonised classification:

The substance has no harmonised classification for repeated dose toxicity according to the Regulation (EC) No. 1272/2008 (CLP). 

 

Self-classification:

Based on the available information, no self-classification is proposed according to the CLP or GHS.