Reaction mass of sodium hydrogen N,N-dibutyl-10-(sulphonatooxy)octadecanamidate and sodium sulphooctadecanoate

Endpoint summary

Description of key information

In the highest dose reversible and/or adaptive and/or secondary changes were detected in body weight, liver and thyroid gland in an oral gavage study in rats.

 

A subchronic oral toxicity study in the rat according to OECD No. 408 is on-going. After finalization of this test, this section will be updated accordingly.

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:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2002-02-04 to 2002-07-09

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

other: HanBrl:WIST (SPF)

Details on species / strain selection:

Recognized by the international guidelines as the recommended test system.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: RCC Ltd, Biotechnology & Animal Breeding Division, CH-4414 Fuellinsdorf / Switzerland
- Age at study initiation: 6 weeks
- Weight at study initiation: Males: 107 - 130 g; Females: 82 - 119 g
- Fasting period before study: no
- Housing: Makrolon type-4 cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: yes

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 45 – 65
- Photoperiod (hrs dark / hrs light): 12 hours fluorescent light / 12 hours dark

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

bidistilled water

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Concentration, homogeneity and stability (after 2 hours and 7 days) of the dose formulations were determined in samples taken after commencement of treatment. Concentration of the dose formulations was determined in samples taken during weeks 2, 3, and 4 of treatment. The analyses were performed according to a HPLC analytical method.

Duration of treatment / exposure:

28 days

Frequency of treatment:

daily

Dose / conc.:

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

Remarks:

corresponding to dose levels of ca. 76 mg/kg body weight/day of the REACH registration substance

Dose / conc.:

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

Remarks:

corresponding to dose level of ca. 304 mg/kg body weight/day of the REACH registration substance

Dose / conc.:

3 290 mg/kg bw/day (actual dose received)

Remarks:

corresponding to dose level of ca. 1520 mg/kg body weight/day of the REACH registration substance

No. of animals per sex per dose:

5

Control animals:

yes, concurrent vehicle

Details on study design:

- Rationale for animal assignment: random
- Rationale for selecting satellite groups: satellite groups to determine reversibility of possible effect
- Post-exposure recovery period in satellite groups: control and high dose

Positive control:

no

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations checked: state of health or behavior, any reaction to treatment


DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: during pretest and once weekly thereafter

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION
- Food consumption for each animal determined as g food/kg body weight / day: yes


OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: week 4 (main & satellite group) ; week 6 (satellite groups)
- Anaesthetic used for blood collection: Yes (isoflurane)
- Animals fasted: Yes
- How many animals: all

CLINICAL CHEMISTRY: yes
- Time schedule for collection of blood: week 4 (main & satellite groups); week 6 (satellite groups)
- Animals fasted: Yes
- How many animals: all

URINALYSIS: Yes
- Time schedule for collection of urine: week 4 (main & satellite groups); week 6 (satellite groups)
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: end of treatment
- Dose groups that were examined: all
- Battery of functions tested: sensory activity / grip strength / motor activity / landing foot splay

Sacrifice and pathology:

GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes

Statistics:

Body weight data were analyzed using Dunnett's multiple comparison procedure implemented in the RCC Tox system for comparing treated groups with a control group. Analyses of organ weights, functional observations and clinical laboratory were carried out using the statistical routines contained in the NOVATOX system. Quantitative data were analyzed either using parametric or non-parametric statistical tests following a pre-test for uniformity of the within group variances based upon the Bartlett's test of homogeneity of variances. ln the case of a non-significant Bartlett's test (p>0.05) a one-way analysis of variance (ANOVA) were carried out. lf the overall test of differences between the groups is significant (p<0.05), comparisons were made between the control group and each of the treatment groups using Dunnett's multiple comparison test. ln cases where Bartlett's test is significant (p<0.05) the Kruskal-Wallis non-parametric test of the differences between the groups were carried out. lf this test is significant (p<0.05), comparisons were made between the control group and each of the treatment groups using Dunn's multiple comparison test. Ordinal data were analyzed using the non-parametric Kruskal-Wallis test. lf this test is significant (p<0.05), comparisons were made between the control group and each of the treatment groups using Dunn's multiple comparison test (Dunn, 1964). All statistical tests are two-sided: testing for an overall difference rather than a difference in only one direction.

Clinical signs:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

effects observed, treatment-related

Clinical biochemistry findings:

effects observed, treatment-related

Endocrine findings:

not examined

Urinalysis findings:

effects observed, treatment-related

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

effects observed, treatment-related

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

effects observed, treatment-related

Details on results:

Mortality
One male and one female both treated at the high dose (3290 mg/kg/bw/day were found dead on day 13 of the treatment period. Both animals presented with moderate squamous cell hyperplasia of the fore stomach and moderate atrophy of the splenic white pulp. ln both cases, the cause of death was not evident. Two males of the control and mid dose groups died shortly after blood collection at the end of the study.


Clinical Signs:
In one single female of the high dose group on day 13 only (piloerection, hunched posture, and respiratory sounds)
As two deaths as well as the only clinical findings were observed on day 13, it is likely that these findings were due to difficulties with administration on that day (misgavage) and not connected to the test substance.


Body weights
Slightly lower body weights and markedly reduced body weight gain were noted in males of the high dose group. The body weight development of the remaining animals was unaffected.


Food consumption
Transient reduction in mean food consumption was noted during the first week in rats of the high dose group. Food consumption in both sexes was markedly increased during the first week of the recovery period. All other animals were unaffected.


Hematology:
At the end of the treatment period, males and females of high dose group had a slight leukocytosis associated with neutrophilia and slightly higher monocyte counts. ln addition changes to red blood cell parameters including a lower mean value for hemoglobin concentration with a lower MCHC value and higher red cell volume distribution width (RDW) and hemoglobin concentration distribution width (HDW) indicating anisocytosis and anisochromia of red blood cells. The shift in the reticulocyte maturation index to immature cells (lower value for L-Reti and higher value for H-Reti) recorded for females of group 4 (high dose group) points to a stimulated erythropoietic activity.
All these changes to red and white blood cell parameters were reversible within the two weeks recovery period.


Clinical Chemistry:
Treatment-related changes in males were confined to the high dose group and included a higher mean value for creatinine, a lower globulin level with a higher albumin to globulin ratio, higher values for triglycerides and phospholipids, lower values for sodium and potassium, a higher phosphate level, and increased activities for aspartate aminotransferase and alanine aminotransferase.
In females, treatment related findings included lower protein and albumin levels for groups 3 and 4 (mid and high dose group), and higher plasma glucose, cholesterol, triglyceride, phospholipid and phosphate levels for group 4 animals.
Except for the lower albumin and higher triglyceride levels in females all the changes were at least partially reversible within the recovery period.


Urinalysis
At the end of the treatment period, males of group 4 excreted lower quantities of more concentrated urine than did the controls. The urine of high dose males was contaminated with feces. This resulted in yellow-brown or brown discoloration and turbidity, and a higher content of erythrocytes and leukocytes. The contamination prevented a detailed examination of urine sediment in high dose males.
In contrast, females of group 4 excreted higher quantities of less concentrated urine. Furthermore, the urine had a slightly higher pH-value, and minimally higher erythrocyte content.
At the end of the recovery period, values for all parameters in previously treated animals were similar to those of the controls.


Organ weight:
After 4 weeks
Treatment-related effects on absolute organ weights included decreased carcass (13 %) and heart weights (16 %) in males of the high dose group, and increased liver weight (33 %) in females treated of the high dose group. The relative weights of brain (12 %), liver (28 %) and adrenals (28 %) were increased in males of the high dose group, and the relative weight of liver was increased (43 %) in females of the high dose group.
After 6 weeks
After the 2-week recovery period there were no relevant differences in absolute and relative organ weights between the high-dosed animals and controls.


MACROSCOPIC FINDINGS
There were no test item-related macroscopic findings.


MICROSCOPIC FINDINGS
Forestomach: Squamous hyperplasia of the epithelium of the forestomach was observed in 1/5 males of the mid dose group and 5/5 males and females of the high dose group. After the recovery period, this finding was observed in 2/5 males and 4/5 females of the high dose group. This change was graded as minimal in the male at 200 mg/kg/bw/day, slight to moderate in males at 1000 mg/kg/bw/day, and slight to moderate in females at 1000 mg/kg/bw/day. After recovery period the lesion was scored as minimal in the affected animals indicating that this finding was partially reversible during this period. ln addition, the forestomach of 2/5 males of the high dose group presented with erosion of moderate or marked degree. This finding was not observed after the recovery period.

Liver: Centrilobular hepatocellular hypertrophy was observed in 3/5 males of the low, 3/5 males of the mid, 4/5 males and 4/5 females of the high dose group. The change was graded as minimal in males of the low and mid dose group and minimal to slight in males and females of the high dose group. This finding was not observed after the recovery period.

Spleen: Extra-medullary hematopoiesis was decreased in incidence and grading in males of the high dose group.
After recovery period, the incidence was similar in control and treated males, but the average grading was still decreased in the treated group.
Thyroid gland: Follicular cell hypertrophy was increased in incidence and grading in females of the high dose group. After the recovery period, the incidence and grading of this change was similar in control and treated groups.

Key result

Dose descriptor:

NOAEL

Effect level:

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

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: overall effects body weight; food consumption; haematology; clinical chemistry; urinalysis; organ weights; histopathology

Key result

Critical effects observed:

not specified

Conclusions:

Based on the results obtained in this study, 658 mg/kg body weight/day of the test material was established as the no-observed-adverse-effect-level (NOAEL) in males and females.

Executive summary:

The purpose of this oral toxicity study was to assess the cumulative toxicity of the test item when administered daily to rats by gavage for a period of 28 days according to OECD TG 407. The reversibility of treatment-related changes was assessed after a treatment-free 14-day recovery period. In this sub-acute toxicity study, the test item was administered daily by oral gavage to SPF-bred Wistar rats (HanBrl: WIST) of both sexes at applied doses of 164.5, 658, and 3290 mg/kg body weight/day of the test substance (corresponding to dose levels of ca. 76, 304 and 1520 mg/kg body weight/day of the REACH registration substance) for a period of 28 days. A control group (0 mg/kg body weight/day) was treated similarly with the vehicle, bidistilled water, only. The groups comprised 5 animals per sex which were sacrificed after 4 weeks of treatment. Additional 5 rats per sex and group were used at 0 and 3290 mg/kg body weight/day. These animals were treated for 28 days and then allowed a 14-day treatment-free recovery period after which they were sacrificed. General and detailed clinical signs, food consumption and body weights were recorded periodically during pretest, the treatment and recovery periods. Functional observational battery and locomotor activies were performed during week 4. At the end of the dosing and the treatment-free recovery periods, blood samples were withdrawn for hematology and plasma chemistry analyses. Urine samples were collected for urinalysis. All animals were killed, necropsied and examined post mortem. Histological examinations were performed on organs and tissues from all control and high-dosed animals, and, in addition, on liver, mesenteric lymph node, spleen, stomach and thyroid gland from low and mid-dosed animals. Based on the results obtained in this study, 658 mg/kg body weigh/day (group 3) of the test material was established as the no-observed-adverse-effect-level (NOAEL) in males and females.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

658 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

reliable with restriction, guideline study under GLP, limited study duration

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

The purpose of this oral toxicity study was to assess the cumulative toxicity of the test item when administered daily to rats by gavage for a period of 28 days according to OECD TG 407. The study was performed with a commercial formulation containing 46.2% (w/w) of the REACH registration substance. The water content of the actual test item was 57.6% (w/w). For the substance subject to REACH registration a water content of 3.8% (w/w) was analytically determined (please refer IUCLID section 1.2). Therefore, results for the actual test item were recalculated to obtain effect concentrations that correspond to the substance to be registered according to REACH taking into account its specific water content.

 

The reversibility of treatment-related changes was assessed after a treatment-free 14-day recovery period. In this sub-acute toxicity study, the test item was administered daily by oral gavage to SPF-bred Wistar rats (HanBrl: WIST) of both sexes at applied doses of 164.5, 658, and 3290 mg/kg body weight/day of the test item (corresponding to dose levels of ca. 76, 304 and 1520 mg/kg body weight/day of the REACH registration substance) for a period of 28 days. A control group (0 mg/kg body weight/day) was treated similarly with the vehicle, bidistilled water, only. The groups comprised 5 animals per sex which were sacrificed after 4 weeks of treatment. Additional 5 rats per sex and group were used at 0 and 3290 mg/kg body weight/day. These animals were treated for 28 days and then allowed a 14-day treatment-free recovery period after which they were sacrificed. General and detailed clinical signs, food consumption and body weights were recorded periodically during pretest, the treatment and recovery periods. Functional observational battery and locomotor activies were performed during week 4. At the end of the dosing and the treatment-free recovery periods, blood samples were withdrawn for hematology and plasma chemistry analyses. Urine samples were collected for urinalysis. All animals were killed, necropsied and examined post mortem. Histological examinations were performed on organs and tissues from all control and high-dosed animals, and, in addition, on liver, mesenteric lymph node, spleen, stomach and thyroid gland from low and mid-dosed animals. Based on the results obtained in this study, 658 mg/kg body weigh/day (group 3) of the test material was established as the no-observed-adverse-effect-level (NOAEL) in males and females.

A subchronic oral toxicity study in the rat according to OECD No. 408 is on-going. After finalization of this test, this section will be updated accordingly.

Justification for classification or non-classification

Not classified

There was no relevant finding in the subacute repeated dose toxicity study that could have triggered a classification. All findings were slight and of reversible and/or adaptive nature.