1-Propanaminium, 3-amino-N-(carboxymethyl)-N,N-dimethyl-, N-C8-10 (even numbered) acyl derivs., hydroxides, inner salts

Toxicity to reproduction

Administrative data

Endpoint:

fertility, other

Remarks:

Repeated Dose 28-Day Oral Toxicity in Rodents

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2015-08-21 to 2015-10-16

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015-08-21 to 2015-10-16

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))

Version / remarks:

2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted 3 October 2008

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 27-29 days
- Weight at study initiation: 92-100 g for males and 80-95 g for females at arrival
- Fasting period before study: no
- Housing: up to 5 of one sex to a cage, in clear polisulphone solid bottomed cages; nesting material was provided inside suitable bedding bag and changed at least twice a week
- Diet (e.g. ad libitum): powdered laboratory rodent diet (4 RF 21, Mucedola S.r.l., Via G. Galilei, 4, 20019, Settimo Milanese (MI), Italy), ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: approx. 3 wk

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±2
- Humidity (%): 55±15
- Air changes (per hr): 15 - 20
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

oral: gavage

Vehicle:

other: purified water (softened water by reverse osmosis)

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
The required amount of test item was dissolved in the vehicle. The formulations were prepared daily at concentrations of 10, 30 and 50 mg/mL. Concentrations were calculated and expressed in terms of test item corrected against the declared purity.

VEHICLE
- Amount of vehicle (if gavage): 10 mL/kg bw/d

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The proposed formulation procedure for the test item was checked in the range from 10 to 100 mg/mL by chemical analysis (concentration) and a 28 hour and 8 day stability at room temperature was verified in the same range, to confirm that the method was suitable and stability was satisfactory.
Final results for all levels were within the acceptability limits for concentration (90-110%).
Samples of the formulations prepared on Weeks 1 and 4 were analysed to check the concentration. Results of the analyses were within the acceptability
limits for concentration of solutions (90-110%)

Duration of treatment / exposure:

4 weeks + recovery period of 2 weeks

Frequency of treatment:

daily

Remarks:

Doses / Concentrations:
0, 100, 300, 500 mg/kg bw/d
Basis:
actual ingested

No. of animals per sex per dose:

5 male and 5 female; control and high dose groups included 5 additional animals per sex to be sacrificed after 2 weeks of recovery

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: based on results with similar substances
- Rationale for animal assignment (if not random): computerised stratified randomisation to give approximately equal initial group mean body weights
- Rationale for selecting satellite groups: to assess recovery from any treatment-related effects
- Post-exposure recovery period in satellite groups: 2 weeks

Positive control:

n.a.

Observations and examinations performed and frequency:

MORTALITY: Yes
- twice daily

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once daily, approximately 1 hour after dosing

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once before commencement of treatment and at least once per week from the start of treatment
- parameters: gait and posture, reactivity to handling, presence of clonic or tonic movements, stereotypies or bizarre behaviour and effects on the autonomic nervous system (e.g. lachrymation, piloerection, unusual respiratory pattern)

BODY WEIGHT: Yes
- Time schedule for examinations: once weekly starting from the day of allocation to treatment group and just prior to necropsy

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

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 (if drinking water study): No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at the end of Week 4 of treatment, prior to necropsy / at the end of Week 2 of the recovery period
- Anaesthetic used for blood collection: Yes (isofluorane)
- Animals fasted: Yes
- How many animals: all surviving animals
- Parameters: Haematocrit, Haemoglobin, Red blood cell count, Reticulocyte count, Mean red blood cell volume, Mean corpuscular haemoglobin, Mean corpuscular haemoglobin concentration, White blood cell count, Differential leucocyte count (Neutrophils, Lymphocytes, Eosinophils, Basophils, Monocytes, Large unstained cells), Platelets, Prothrombin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the end of Week 4 of treatment, prior to necropsy / at the end of Week 2 of the recovery period
- Animals fasted: Yes
- How many animals: all surviving animals
- Parameters: Alkaline phosphatase, Alanine aminotransferase, Aspartate aminotransferase, Gamma glutamyltransferase, Urea, Creatinine, Glucose, Triglycerides, Bile acids, Phosphorus, Total bilirubin, Total cholesterol, Total protein, Albumin, Globulin, A/G Ratio, Sodium, Potassium, Calcium, Chloride

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: once during Week 4 of treatment and once during Week 2 of recovery
- Dose groups that were examined: all groups
- Battery of functions tested: sensory activity / grip strength / motor activity

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (see table)
HISTOPATHOLOGY: Yes (see table)

Other examinations:

The testes and epididymides of main group animals were cut at 2-3 µm thickness and stained with Periodic Acid Schiff (PAS). The morphological evaluation of the seminiferous epithelium (staging of spermatogenic cycle) was performed initially in all animals in the control and high dose groups dying during the treatment period or killed at the end of the 4 weeks of treatment.
Moreover, the histopathological evaluation of the oestrous cycle in the uterus/ cervix and vagina from all animals in the control and high dose groups of the
main phase was performed.

Statistics:

Standard deviations were calculated as considered appropriate. For continuous variables the significance of the differences amongst groups was assessed by analysis of variance. Differences between each treated group and the control group were assessed by Dunnett’s test using a pooled error variance. The homogeneity of the data was verified by Bartlett’s test before Dunnett’s test.
If the data were found to be inhomogeneous a Modified t test (Cochran and Cox) was applied.
The mean values, standard deviations and statistical analysis were calculated from the actual values in the computer without rounding off.

Clinical signs:

no effects observed

Description (incidence and severity):

No clinical signs were seen during the study.
No toxicological relevant changes were observed at the weekly detailed clinical signs.
Statistically significant decreases in rearing were seen in males receiving 300 and 500 mg/kg bw/day during Weeks 3 and 4 of treatment, increases were seen in females receiving 100 and 300 mg/kg bw/day during Week 3 and in females receiving 500 mg/kg bw/day during Week 4 of treatment, when compared to controls. Since the direction of changes was opposite in the two sexes, the above mentioned changes were considered incidental and not treatment related.

Mortality:

no mortality observed

Description (incidence):

No mortality occurred during the study.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

No differences in body weight were seen between treated and control animals during the study.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

No differences in food consumption were seen between treated and control groups during the study.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Description (incidence and severity):

Dosing phase
Lymphocytosis and/or monocytosis were recorded in two females dosed at 300 mg/kg bw/day and 3 females receiving 500 mg/kg bw/day. When compared with mean control data, individual changes were between 61% and 93% for lymphocytes and 137% and 150% for monocytes.
The statistically significant increase of eosinophils observed in females dosed at 300 mg/kg bw/day was not dose-related, therefore considered incidental.

Recovery phase
After the recovery period, one female showed lymphocytosis and neutrophilia. When compared with mean control data, the individual change was of 49% and 3.7 fold, respectively. Concerning the other treated animals, lymphocytes showed complete reversibility and monocytes data were comparable with controls, even though values were similar to those observed during the dosing phase.
The statistically significant differences recorded between control and treated animals (reticulocytes and lymphocytes relative count in males, mean corpuscular haemoglobin in females) were not observed during the dosing phase, therefore considered unrelated to treatment.
No changes in coagulation parameter were seen in control and treated animals.

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

Chloride and sodium showed a statistically significant decrease in females dosed at 300 and/or 500 mg/kg bw/day, when compared to controls. Due to the minimal severity (up to 2%), these findings were considered of no toxicological relevance.
The statistically significant changes recorded in animals dosed at 100 and/or 300 mg/kg bw/day (bilirubin, bile acids and inorganic phosphorus in males; urea in females) were not dose-related, therefore considered unrelated to treatment.

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Description (incidence and severity):

No relevant differenecs between treated and control groups were seen in landing foot splay.
Changes in grip strength were considered incidental and not treatment-related, since the direction of changes was opposite in the two sexes: decreases in males and increases were seen in females, receiving 500 mg/kg bw/day, when compared to controls.
No differences between treated and control groups were evident at the motor activity measurements.

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

No relevant differences were seen in terminal body weights of treated animals respect to controls.
No toxicological relevance was attributed to the statistically significant increase in absolute (13%) and/or relative (9-10%) kidneys weight seen in females treated at 300 and/or 500 mg/kg/day not to the statistically significant increase in relative (up to 7%) kidneys weight seen in males treated at 300 and 500 mg/kg/day, when compared to controls, since these increases were slight and no changes were detected at the histopathological evaluation.
No other differences were seen in treated groups when compared to controls.

Gross pathological findings:

no effects observed

Description (incidence and severity):

No relevant changes were noted in treated animals. The sporadic changes observed in few treated animals could be considered incidental.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

No treatment-related changes were noted. All observed changes in organs and tissues are considered as incidental-age related findings, characteristically seen in untreated Sprague Dawley rats of the same age or comparable with control animals. Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted.
Normal physiology of the oestrous cycle (oestrous, metestrous, diestrous and proestrous) was noted in control and treated females. The morphological changes seen were normal when compared to each “oestrous phase” in the ovaries, uterus/cervix and vagina.

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Key result

Dose descriptor:

NOAEL

Effect level:

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

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: no treatment-related adverse effects up to and including the highest tested dose level

Key result

Critical effects observed:

no

Conclusions:

On the basis of the results obtained in this study, the dose level of 500 mg/kg bw/day was considered the NOAEL.

Executive summary:

In a subacute toxicity study according to OECD guideline 407 (2008) and EU method B.7 (2008) C8-10 Alkylamidopropyl betaine (34.65% a.i.) was administered to 5Hsd: Sprague Dawley SDrats/sex/dose in purified water by gavage at dose levels of 0 (control), 100, 300 and 500 mg/kg bw/day for 28 consecutive days. Control and high dose groups included 5 additional animals per sex to be sacrificed after 2 weeks of recovery.

No mortality occurred. No clinical signs and no changes were observed at the weekly detailed clinical observations. Neurotoxicity assessment did not reveal any treatment-related effects. No changes on body weight and food consumption were noted.

The lymphocytosis and monocytosis seen in single females dosed at 300 and/or 500 mg/kg bw/day showed reversibility at the end of the recovery period or comparability to control data. No toxicological relevant effects in coagulation and clinical chemistry parameters were observed. No differences were reported in terminal body weights and organ weights between treated and control animals and no treatment-related changes were noted at macroscopic and microscopic observations.

Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted.

Normal physiology of the oestrous cycle (oestrous, metestrous, diestrous and proestrous) was noted in control and treated females. The morphological changes seen were normal when compared to each “oestrous phase” in the ovaries, uterus/cervix and vagina.

On the basis of the results obtained in this study, the dose level of 500 mg/kg bw/day was considered the NOAEL (No Observed Adverse Effect Level).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))

Version / remarks:

2008

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2008

Deviations:

no

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2-[(3-decanamidopropyl)dimethylazaniumyl]acetate 2-[dimethyl(3-octanamidopropyl)azaniumyl]acetate

EC Number:

944-170-2

Molecular formula:

not applicable, UVCB substance

IUPAC Name:

2-[(3-decanamidopropyl)dimethylazaniumyl]acetate 2-[dimethyl(3-octanamidopropyl)azaniumyl]acetate

Test material form:

solid - liquid: aqueous solution

Details on test material:

- Name of test material (as cited in study report): 1-Propanaminium, 3-amino-N-(carboxymethyl)-N,N-dimethyl-, N-(C8-10 acyl) derivs., hydroxides, inner salts
- Analytical purity: 34.65% active ingredient (betain)

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 27-29 days
- Weight at study initiation: 92-100 g for males and 80-95 g for females at arrival
- Fasting period before study: no
- Housing: up to 5 of one sex to a cage, in clear polisulphone solid bottomed cages; nesting material was provided inside suitable bedding bag and changed at least twice a week
- Diet (e.g. ad libitum): powdered laboratory rodent diet (4 RF 21, Mucedola S.r.l., Via G. Galilei, 4, 20019, Settimo Milanese (MI), Italy), ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: approx. 3 wk

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±2
- Humidity (%): 55±15
- Air changes (per hr): 15 - 20
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: purified water (softened water by reverse osmosis)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
The required amount of test item was dissolved in the vehicle. The formulations were prepared daily at concentrations of 10, 30 and 50 mg/mL. Concentrations were calculated and expressed in terms of test item corrected against the declared purity.

VEHICLE
- Amount of vehicle (if gavage): 10 mL/kg bw/d

Details on mating procedure:

animals were not mated

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The proposed formulation procedure for the test item was checked in the range from 10 to 100 mg/mL by chemical analysis (concentration) and a 28 hour and 8 day stability at room temperature was verified in the same range, to confirm that the method was suitable and stability was satisfactory.
Final results for all levels were within the acceptability limits for concentration (90-110%).
Samples of the formulations prepared on Weeks 1 and 4 were analysed to check the concentration. Results of the analyses were within the acceptability
limits for concentration of solutions (90-110%)

Duration of treatment / exposure:

4 weeks + recovery period of 2 weeks

Frequency of treatment:

daily

Details on study schedule:

-animals were not mated

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

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

Doses / concentrations 2

Dose / conc.:

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

Doses / concentrations 3

Dose / conc.:

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

Doses / concentrations 4

Dose / conc.:

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

No. of animals per sex per dose:

5 male and 5 female; control and high dose groups included 5 additional animals per sex to be sacrificed after 2 weeks of recovery

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: based on results with similar substances
- Rationale for animal assignment (if not random): computerised stratified randomisation to give approximately equal initial group mean body weights
- Rationale for selecting satellite groups: to assess recovery from any treatment-related effects
- Post-exposure recovery period in satellite groups: 2 weeks

Positive control:

n.a.

Examinations

Parental animals: Observations and examinations:

MORTALITY: Yes
- twice daily

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: once daily, approximately 1 hour after dosing

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once before commencement of treatment and at least once per week from the start of treatment
- parameters: gait and posture, reactivity to handling, presence of clonic or tonic movements, stereotypies or bizarre behaviour and effects on the autonomic nervous system (e.g. lachrymation, piloerection, unusual respiratory pattern)

BODY WEIGHT: Yes
- Time schedule for examinations: once weekly starting from the day of allocation to treatment group and just prior to necropsy

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): No

OTHER: NEUROBEHAVIOURAL EXAMINATION, CLINICAL CHEMISTRY, HAEMATOLOGY
- described in more detail in section "Repeated dose toxicity"

Oestrous cyclicity (parental animals):

The histopathological evaluation of the oestrous cycle in the uterus/ cervix and vagina from all animals in the control and high dose groups of the
main phase was performed.

Sperm parameters (parental animals):

The testes and epididymides of main group animals were cut at 2-3 µm thickness and stained with Periodic Acid Schiff (PAS). The morphological evaluation of the seminiferous epithelium (staging of spermatogenic cycle) was performed initially in all animals in the control and high dose groups dying during the treatment period or killed at the end of the 4 weeks of treatment.

Litter observations:

n.a.

Postmortem examinations (parental animals):

GROSS PATHOLOGY: Yes (see table)
HISTOPATHOLOGY: Yes (see table)

Postmortem examinations (offspring):

n.a.

Statistics:

Standard deviations were calculated as considered appropriate. For continuous variables the significance of the differences amongst groups was assessed by analysis of variance. Differences between each treated group and the control group were assessed by Dunnett’s test using a pooled error variance. The homogeneity of the data was verified by Bartlett’s test before Dunnett’s test.
If the data were found to be inhomogeneous a Modified t test (Cochran and Cox) was applied.
The mean values, standard deviations and statistical analysis were calculated from the actual values in the computer without rounding off.

Reproductive indices:

n.a.

Offspring viability indices:

n.a.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Description (incidence and severity):

No clinical signs were seen during the study.
No toxicological relevant changes were observed at the weekly detailed clinical signs.
Statistically significant decreases in rearing were seen in males receiving 300 and 500 mg/kg bw/day during Weeks 3 and 4 of treatment, increases were seen in females receiving 100 and 300 mg/kg bw/day during Week 3 and in females receiving 500 mg/kg bw/day during Week 4 of treatment, when compared to controls. Since the direction of changes was opposite in the two sexes, the above mentioned changes were considered incidental and not treatment related.

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Description (incidence):

No mortality occurred during the study.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

No differences in body weight were seen between treated and control animals during the study.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

No differences in food consumption were seen between treated and control groups during the study.

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

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

No treatment-related changes were noted. All observed changes in organs and tissues are considered as incidental-age related findings, characteristically seen in untreated Sprague Dawley rats of the same age or comparable with control animals.

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

no effects observed

Description (incidence and severity):

Normal physiology of the oestrous cycle (oestrous, metestrous, diestrous and proestrous) was noted in control and treated females. The morphological changes seen were normal when compared to each “oestrous phase” in the ovaries, uterus/cervix and vagina.

Reproductive function: sperm measures:

no effects observed

Description (incidence and severity):

Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted.

Reproductive performance:

not examined

Effect levels (P0)

1

Key result

Dose descriptor:

NOEL

Remarks:

fertility parameters

Effect level:

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

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: no treatment-related effects up to and including the highest tested dose level

2

Dose descriptor:

NOAEL

Remarks:

general toxicity

Effect level:

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

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: no treatment-related adverse effects up to and including the highest tested dose level

Target system / organ toxicity (P0)

Key result

Critical effects observed:

no

Results: P1 (second parental generation)

General toxicity (P1)

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality:

not examined

Body weight and weight changes:

not examined

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Reproductive function / performance (P1)

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

not examined

Results: F1 generation

General toxicity (F1)

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality / viability:

not examined

Body weight and weight changes:

not examined

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Histopathological findings:

not examined

Other effects:

not examined

Developmental neurotoxicity (F1)

Behaviour (functional findings):

not examined

Developmental immunotoxicity (F1)

Developmental immunotoxicity:

not examined

Results: F2 generation

General toxicity (F2)

Clinical signs:

not examined

Dermal irritation (if dermal study):

not examined

Mortality / viability:

not examined

Body weight and weight changes:

not examined

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Histopathological findings:

not examined

Other effects:

not examined

Developmental neurotoxicity (F2)

Behaviour (functional findings):

not examined

Developmental immunotoxicity (F2)

Developmental immunotoxicity:

not examined

Overall reproductive toxicity

Key result

Reproductive effects observed:

no

Applicant's summary and conclusion

Conclusions:

On the basis of the results obtained in this study, the dose level of 500 mg/kg bw/day was considered the NOAEL for general toxicity and the NOEL for fertility parameters.

Executive summary:

In a subacute toxicity study according to OECD guideline 407 (2008) and EU method B.7 (2008) C8-10 Alkylamidopropyl betaine (34.65% a.i.) was administered to 5Hsd: Sprague Dawley SDrats/sex/dose in purified water by gavage at dose levels of 0 (control), 100, 300 and 500 mg/kg bw/day for 28 consecutive days. Control and high dose groups included 5 additional animals per sex to be sacrificed after 2 weeks of recovery.

No mortality occurred. No clinical signs and no changes were observed at the weekly detailed clinical observations. Neurotoxicity assessment did not reveal any treatment-related effects. No changes on body weight and food consumption were noted.

The lymphocytosis and monocytosis seen in single females dosed at 300 and/or 500 mg/kg bw/day showed reversibility at the end of the recovery period or comparability to control data. No toxicological relevant effects in coagulation and clinical chemistry parameters were observed. No differences were reported in terminal body weights and organ weights between treated and control animals and no treatment-related changes were noted at macroscopic and microscopic observations.

Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted.

Normal physiology of the oestrous cycle (oestrous, metestrous, diestrous and proestrous) was noted in control and treated females. The morphological changes seen were normal when compared to each “oestrous phase” in the ovaries, uterus/cervix and vagina.

On the basis of the results obtained in this study, the dose level of 500 mg/kg bw/day was considered the NOAEL (No Observed Adverse Effect Level).

The NOEL for fertility parameters is 500 mg/kg bw/d.