Registration Dossier
Registration Dossier
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EC number: 201-162-7 | CAS number: 78-96-6
Endpoint summary
Administrative data
Key value for chemical safety assessment
Effects on fertility
Description of key information
An OECD443 with MIPA is currently being conducted. In a weight-of-evidence approach it is so far concluded, that MIPA does not induce effects on fertility: In a combined repeated dose oral toxicity study with a reproduction/developmental screening study (OECD422) in rats exposed to the hydrochloride salt of MIPA by gavage, a NOAEL for reproductive performance and fertility of 1000 mg/kg bw/day (equivalent to 673 mg/kg bw/day MIPA, the highest dose tested) was established for parental males and females. In an oral one-generation study with the structurally similar TIPA in rats according to FDA guidelines, the NOAEL for the parental generation as well as the off-spring was reported to be the highest dose tested 7500 ppm TIPA in males and females (equivalent to a 239 mg/kg bw/d intake of MIPA in males and 275 mg/kg bw/d in females).
Link to relevant study records
- Endpoint:
- extended one-generation reproductive toxicity - basic test design (Cohorts 1A, and 1B without extension)
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 1988
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- ADAPTATION ACCORDING TO REACH ANNEX XI, section 1 - see justification attached to IUCLID section 13.2.
- Justification for WoE: RDT and Toxicity to reproduction - Qualifier:
- according to guideline
- Guideline:
- other: U.S. FDA Toxicological Principles for the Safety Assessment of Direct Food Additives and Color Additives Used in Food - 21 CFR 314.50(d)(2)
- GLP compliance:
- yes
- Limit test:
- no
- Justification for study design:
- A ccording to REACH Annex XI, section 1 and in line with the ECHA Guidance R.7a: Endpoint specific guidance (v6.0, July 2017) and R.6 QSARs and grouping of chemicals (May 2008), a testing does not appear scientifically necessary, because there is sufficient weight of evidence from available toxicological data for TIPA and structural similar substances (DIPA and MIPA), leading to the conclusion, that TIPA does not have an adverse effect on reproduction (fertility) and the available data are adequate to support a robust risk assessment.
- see justification attached to IUCLID section 13
SPECIFICATION OF STUDY DESIGN FOR ONE-GENERATION REPRODUCTION TOXICITY STUDY WITH JUSTIFICATIONS:
- Premating exposure duration for parental (P0) animals : 5 weeks
- Basis for dose level selection : selected after consultation with the FDA (for further details please refer to 'Details on study design' below)
- Exclusion of extension of Cohort 1B
- Termination time for F2 : no F2 generation was produced
- Exclusion of developmental neurotoxicity Cohorts 2A and 2B
- Exclusion of developmental immunotoxicity Cohort 3
- Route of administration : oral by feeding - Specific details on test material used for the study:
- Purity: 98.76%
- Species:
- other:
- Strain:
- Sprague-Dawley
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River Breeding Laboratories, Kingston, New York
- Age: (P) 22 days old upon arrival and 42 days old at study start
- Weight at study initiation: (P) Males: 189.6-193.4 g; Females: 133.0-138.3 g
- Housing:
- during acclimation: 3 animals/cage, sexes separate, in stainless steel, wire-mesh cages suspended above Upjohn Deotized Animal Cage Boards (DACB) or R2 Reemay-backed cage boards
- after grouping: individually housed in stainless steel, wire-mesh cages suspended above Upjohn DACB or R2 Reemay-backed cage boards
- during the premating phase and throughout the study except during mating: housed on separate cage racks
- Diet: ad libitum, Irradiated Purina Certified Rodent Chow #5002 (IPCRC) diet
- Water: ad libitum, tap water
- Acclimation period: 14 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23±2
- Humidity (%): 50±10
- Air changes (per hr): not specified
- Photoperiod: 12-hour light/12-hour dark
IN-LIFE DATES: From: 03-09-1987 To: 09-08-1987 - Route of administration:
- oral: feed
- Vehicle:
- water
- Details on exposure:
- PREPARATION OF DOSING SOLUTIONS:
DIET PREPARATION
- TIPA was dried with Drierite drying agent under vacuum for several days. All compound was stored in containers which were purged with nitrogen after opening.
- Rate of preparation of diet: prepared weekly
- Mixing appropriate amounts with: distilled water (200 mL) and mixed on a magnetic stirrer until dissolved; added to Irradiated Purina Certified Rodent Chow (IPCRC) diet afterwards
- Storage temperature of food: refrigerated until used - Details on mating procedure:
- - M/F ratio per cage: 1/1
- Length of cohabitation: maximum of seven days for the first approach; seven days for the second approach
- Proof of pregnancy: vaginal plug referred to as day 0 of pregnancy
- After 7 days of unsuccessful pairing replacement of first male by another male with proven fertility.
- Further matings after two unsuccessful attempts: no
- After successful mating each pregnant female was caged: individually in the polypropylene cages at the end of this period
Female rats were observed at least twice daily for signs of delivery starting several days prior to expected parturition. - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Methanol was added to each diet sample and TIPA was extracted by sonication, filtered, evaporated under nitrogen, and derivatized with Pyridine and N-Trimethylsilylimidazole (TMSI). The reaction was completed by warming the solutions to 80°C for 10 (0, 500 ppm) or 20 minutes (2000, 7500 ppm). Recovery efficiencies were determined at the 500 ppm level by adding 5 mL of a 1000 µg/mL TIPA stock solution to control diet and at the 2000 and 7500 ppm levels by adding 20.3 and 75.9 mg H-16,648, respectively, to 10.0 g control diet. Extraction and preparation of the recovery samples were performed as described above. Calibration curves were created using standarrd solutions of TIPA or TEA. Peak area ratios were calculated (TIPA to TEA) and used for quantitation by linear regression analysis. The dietary concentration of TIPA was determined by multiple injection with a Hewlett-Packard 5880A gas chromatograph.
- Duration of treatment / exposure:
- The parental generation was fed TIPA in diet for five weeks.
After five weeks of feeding, they were mated to produce offspring which were fed diets containing TIPA for 90 days after weaning. - Frequency of treatment:
- Daily
- Dose / conc.:
- 500 ppm (nominal)
- Remarks:
- in diet (equivalent to a calculated 15.6 and 17.2 mg/kg bw/day intake of MIPA for males and females of the F1 generation, respectively)
- Dose / conc.:
- 2 000 ppm (nominal)
- Remarks:
- in diet (equivalent to a calculated 63 and 71 mg/kg bw/day intake of MIPA for males and females of the F1 generation, respectively)
- Dose / conc.:
- 7 500 ppm (nominal)
- Remarks:
- in diet (equivalent to calculated 239 and 275 mg/kg bw/day intake of MIPA for males and females of the F1 generation, respectively)
- No. of animals per sex per dose:
- 25
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- - Dose selection rationale:
Dose levels for this study were selected after consultation with the FDA. Dose levels were based upon acute and subchronic studies in rats conducted with TIPA in drinking water. On an acute basis, the single dose oral LD50 of TIPA in rats was 5,994 mg/kg bw. When rats were given 140 to 1,350 mg TIPA/kg bw/day in drinking water for 30 days, 1,350 mg/kg bw/day resulted in growth reduction and decreased food consumption. A dose level of 260 mg/kg bw/day produced unspecified histopathologic changes in some rats. No effects were seen at 140 mg/kg bw/day. In a 90-day study in rats, dose levels of 770 mg TIPA/kg bw/day in drinking water produced kidney effects consisting of dilation of Bowman's capsule and convoluted tubules, as well as marked cloudy swelling of liver parenchyma. A level of 220 mg/kg bw/day produced unspecified pathological changes in some animals. A concentration of 110 mg/kg bw/day for 90 days revealed no microscopic changes.
In the present study, the intermediate- and high-dose levels (2,000 and 7500 ppm in diet) were expected to produce mean daily intakes in rats of about 200 and 700 mg/kg bw/day, respectively. The intermediate-concentration level was selected to be similar to one at which pathological changes were seen in some rats on a previous study (220 mg/kg bw/day); the high-concentration level was selected to be comparable to a level at which pathological changes were seen in all rats (770 mg/kg bw/day).
- Fasting period before blood sampling for clinical biochemistry:
Two days prior to collection of blood samples for clinical evaluation, ten rats randomly selected from each group were placed in metabolism cages. The day before blood collection, these rats were fasted for approximately 16 hours. Urine was collected from each rat during this period. At the conclusion of this period, blood samples were collected from the orbital sinus of each rat while the rat was under light carbon dioxide anesthesia. - Positive control:
- None
- Parental animals: Observations and examinations:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: conducted at least once daily throughout the study
- Cage-site examinations to detect moribund or dead rats and abnormal behavior and appearance among rats
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: during the premating phase of the study
- each rat was individually handled at each weighing and carefully examined for abnormal behavior and appearance
BODY WEIGHT: Yes
- Time schedule for examinations: once a week during the five-week premating phase of the study
FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each test group determined during the premating phase: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes - Oestrous cyclicity (parental animals):
- not examined
- Sperm parameters (parental animals):
- Parameters examined in F1 male parental generations:
- testis weight - Litter observations:
- STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of 8 pups/litter (4/sex/litter as nearly as possible); excess pups were killed and discarded.
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
- number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities.
GROSS EXAMINATION OF DEAD PUPS:
- no - Postmortem examinations (parental animals):
- None
- Postmortem examinations (offspring):
- SACRIFICE
- The F1 offspring were sacrificed at 111 (21 days p.p. weaning + 90 days feeding phase) days of age.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:
HISTOPATHOLOGY / ORGAN WEIGTHS
- The following tissues indicated were prepared for microscopic examination and weighed, respectively:
Bone marrow, Lymph nodes (mandibular and mesenteric), Spleen, Aorta (thoracic), Heart, Salivary glands, Esophagus, Stomach, Liver (two sections), Pancreas, Small intestine (duodenum, jejunum, and ileum), Large intestine (cecum, colon, and rectum), Kidneys, Bladder, Pituitary, Thyroid - Parathyroid, Adrenals; Males: Prostate, Testes, Epididymides, Seminal vesicles, Females: Mammary gland, Ovaries, Uterus, Vagina; Brain (three coronal sections), Spinal cord, Peripheral nerve (sciatic), Bone (femur and sternum), Eyes, Exorbital lacrimal glands, Harderian glands, All gross lesions - Statistics:
- For the P premating, gestation, and lactation, and F1 feeding phases, body weights, body weight gains, clinical laboratory measurements, and organ weights were analyzed by a one-way analysis of variance. When the test for differences among test group means (F test) was significant, pairwise comparisons between test and control groups were made with the Dunnett's test. Incidence of clinical observations was evaluated by the Fisher's Exact test with a Bonferroni correction and the Cochran-Armitage test for trend. Homogeneity of variances of organ weights and clinical laboratory data were analyzed with the Bartlett's test (alpha = 0.005). When the results of Bartlett's test were significant (variance was not homogeneous), the Mann-Whitney U test was employed instead of Dunnett's test for comparison of means (alpha = 0.05). Comparisons of offspring numbers and weights were made with the Mann-Whitney U test and Jonckheere's test for trend. Incidences of clinical observations in offspring were evaluated by Fisher's Exact test with a Bonferroni correction and the Cochran-Armitage test for trend. Indices of reproductive performance were also evaluated by Fisher's Exact test and the Cochran-Armitage test for trend.
- Reproductive indices:
- Further details given in other information.
Mating index, fertility index, gestation index - Offspring viability indices:
- Further details given in other information.
Percent pups born alive, viability index, lactation index, litter survival, average number of pups/litter - Clinical signs:
- no effects observed
- Description (incidence and severity):
- In the parent rats, there were no significant differences between groups in clinical observations during the premating period or in maternal rats during gestation or lactation.
- Mortality:
- no mortality observed
- Description (incidence):
- No rats died in the premating period.
- Body weight and weight changes:
- no effects observed
- Description (incidence and severity):
- In the parent rats, there were no significant differences between groups in body weights and body weight gains during the premating period or in maternal rats during gestation or lactation.
- Food consumption and compound intake (if feeding study):
- no effects observed
- Description (incidence and severity):
- Food consumption and food efficiency of parental rats were similar between groups. Variability in the values over time and between sexes was related to the normal differences in body weight and food consumption relative to age and sex.
- Food efficiency:
- 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
- Histopathological findings: non-neoplastic:
- no effects observed
- Histopathological findings: neoplastic:
- not examined
- Other effects:
- not examined
- Reproductive function: oestrous cycle:
- not specified
- Reproductive function: sperm measures:
- not specified
- Reproductive performance:
- no effects observed
- Description (incidence and severity):
- There were no differences attributed to administration of TIPA in gestation length, the number of litters produced (fertility index), or any other indices of reproductive performance.
- Key result
- Dose descriptor:
- NOEL
- Effect level:
- 7 500 ppm (nominal)
- Based on:
- test mat.
- Remarks:
- equivalent to 609 mg/kg bw/day for males; 700 mg/kg bw/day for females
- Sex:
- male/female
- Remarks on result:
- other: No treatment-related effects were observed up to the highest dose tested (602 mg/kg bw/day for males; 693 mg/kg bw/day for females)
- Key result
- Critical effects observed:
- no
- Clinical signs:
- no effects observed
- Description (incidence and severity):
- No treatment-related effects on F1 pups from any TIPA-treated groups were observed in clinical signs.
In the 90-day feeding phase of the F1 rats, there were no TIPA-related changes in clinical signs. - Mortality / viability:
- no mortality observed
- Description (incidence and severity):
- No treatment-related effects on F1 pups from any TIPA-treated groups were observed in the number born or survival.
- Body weight and weight changes:
- no effects observed
- Description (incidence and severity):
- No treatment-related, significant effects on F1 pups from any TIPA-treated groups were observed in body weights.
In the 90-day feeding phase of the F1 rats, there were no TIPA-related changes in body weights or body weight gains. - Food consumption and compound intake (if feeding study):
- no effects observed
- Description (incidence and severity):
- In the 90-day feeding phase of the F1 rats, there were no TIPA-related changes in food consumption. The mean daily intake of TIPA over the entire 90-day feeding period was 0, 39.7, 160, and 609 mg/kg bw/day. For females at the same dose group the mean daily intake ws 0, 43.7,182, and 700 mg/kg bw/day over the same period.
- Food efficiency:
- not examined
- Ophthalmological findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- No changes in the ophthalmological examinations conducted on F1 rats were considered compound-related or biologically significant.
- Haematological findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- No changes in haematology on F1 rats was considered to be biologically significant or compound related.
- Clinical biochemistry findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- No changes in the clinical chemistry evaluations conducted on F1 rats were considered compound-related or biologically significant.
- Urinalysis findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- No changes in the urine analyses conducted on F1 rats were considered compound-related or biologically significant.
- Sexual maturation:
- no effects observed
- Organ weight findings including organ / body weight ratios:
- effects observed, non-treatment-related
- Description (incidence and severity):
- No changes in the organ weights conducted on F1 rats were considered compound-related or biologically significant.
- Gross pathological findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- No changes in the pathological evaluations conducted on F1 rats were considered compound-related or biologically significant.
- Histopathological findings:
- effects observed, non-treatment-related
- Other effects:
- not examined
- Behaviour (functional findings):
- not examined
- Developmental immunotoxicity:
- not examined
- Key result
- Dose descriptor:
- NOEL
- Generation:
- F1
- Effect level:
- 7 500 ppm (nominal)
- Based on:
- test mat.
- Remarks:
- (609 mg/kg bw/day for males; 700 mg/kg bw/day for females)
- Sex:
- male/female
- Remarks on result:
- other: No treatment-related effects were observed up to the highest dose tested (609 mg/kg bw/day for males; 700 mg/kg bw/day for females)
- Key result
- Critical effects observed:
- no
- Key result
- Reproductive effects observed:
- no
- Conclusions:
- The two highest levels produced mean daily intake comparable to those at which pathological changes of the kidneys and liver were reported in a drinking water study by another laboratory. However, in the present study no effects were seen at 7,500 ppm. The no-observable-effect level (NOEL) for this study was 7,500 ppm since no effects were observed at any dietary concentration in parental rats or in offspring rats prior to or after weaning.
- Executive summary:
The reported feeding study in Crl:CDBR rats describes a combination of a sub-chronic and in utero study, i.e. F1 offspring rats were fed triisopropanolamine (TIPA) for 90-days after exposure prior to weaning / during pregnancy.
Prior to weaning, these F1 rats were exposed to TIPA in utero in maternal milk during lactation, and in any diet they consumed. The P rats had been continuously fed their group's assigned concentration of TIPA for five weeks prior to mating and during mating, gestation and lactation. Twenty-five rats/sex/group were used for the P generation. Among offspring of the P rats, twenty/sex/group were selected to continue as the F1 generation. Both generations were fed diets that contained 0, 500, 2,000, or 7,500 ppm of TIPA.
Clinical signs, mortality, and body weights of parental rats were recorded throughout the premating period and for females continued to be recorded during the gestation and lactation periods. Food consumption and mean daily intake of TIPA were determined during the premating period for both sexes and for females during gestation. After production of the F1 generation was complete, P rats were sacrificed and discarded without pathological examination. Clinical signs, counts, mortality, and group body weights of F1 offspring were recorded prior to weaning.
At weaning, 20 randomly selected F1 rats/sex/group were selected to continue on their respective treatment group's diet (one/sex/litter when possible). The remainder were sacrificed and discarded without pathological examination. Clinical observations, mortality, body weights, and food consumption of the F1 offspring were recorded during a 90-day feeding phase. F1 offspring were given ophthalmological examinations at the beginning and end of the 90-day feeding period. Clinical chemistry and urine analyses were conducted after approximately 45 and 90 days of feeding.
At the end of the 90-day feeding period, all surviving rats were sacrificed and given a gross pathological examination. Tissues from the control and high concentration groups were examined microscopically. Lungs, liver, kidneys, and organs with lesions in the low- and intermediate-concentration groups were also examined microscopically.
In the parent rats, there were no significant differences between groups in clinical observations, body weights, and weight gains during the premating period or in maternal rats during gestation or lactation. No parental rats died during the premating, mating, gestation, or lactation period. Food consumption and food efficiency of parental rats were similar between groups.
There were no differences attributed to administration of TIPA in gestation length, the number of litters produced (fertility index), or any other indices of reproductive performance.
No treatment-related effects on F1 pups from any TIPA-treated groups were observed in the number born, survival, body weights, or clinical signs.In the 90-day feeding phase of the F1 rats, there were no TIPA-related changes in clinical signs, body weights, body weight gains, or food consumption. One death occurred in the high-concentration group, but is not considered compound related. The mean daily intake of TIPA over the entire
90-day feeding period was 0, 39.7, 160, and 609 mg/kg/day (equivalent to a 15.6, 63 and 239 mg/kg bw/d intake of MIPA) for the control, low-, intermediate-, and high-concentration male groups, respectively. For females at the same concentrations the mean daily intake was 0, 43.7, 182, and 700 mg/kg/day (equivalent to a 17.2, 71 and 275 mg/kg bw/d intake of MIPA) over the same period. No changes in the clinical chemistry evaluations, urine analyses, ophthalmological examinations, organ weights, or pathological evaluations conducted on F1 rats were considered compound-related or biologically significant.
The no-observable-effect-level (NOEL) on this study was 7,500 ppm, the highest level tested.
- Endpoint:
- screening for reproductive / developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- ADAPTATION ACCORDING TO REACH ANNEX XI, section 1 - see justification attached to IUCLID section 13.2.
- Justification for WoE: RDT and Toxicity to reproduction - Reason / purpose for cross-reference:
- reference to same study
- Reason / purpose for cross-reference:
- reference to same study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
- Version / remarks:
- 22 March 1996
- Deviations:
- no
- GLP compliance:
- yes
- Limit test:
- no
- Specific details on test material used for the study:
- SOURCE OF TEST MATERIAL
- Source and batch No.of test material: BASF and BA 1449
- Purity test date: 04 Jan 2006
- concetration of given stock solution:65.4 g isopropanolamine hydrochloride per 100 g aqueous solution
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature (freezer for reanalysis)
- Stability under test conditions: confirmed by reanalysis
- Solubility and stability of the test substance in the solvent/vehicle: stability of aq. solution confirmed by reanalysis
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: test material was weighed up and dissolved in tap water
FORM AS APPLIED IN THE TEST: solution - Species:
- rat
- Strain:
- Wistar
- Remarks:
- Crl:WI(Han)
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- - Source: Charles River Laboratories, Research Models and Services, Germany Gmbh
- Age at study initiation: 11-13 weeks
- Weight at study initiation: 267.4-309.0 g (males) and 186.7-217.7 g (females)
- Number of animals: 96 (12 per sex per dose group)
- Housing: Individually in type DK III stainless steel wire mesh cages supplied by Becker & Co., Castrop-Rauxel, Germany (floor area of about 800 cm2), with the following exceptions: for the overnight mating the females were put into the cages of the males
- Diet: Ground Kliba maintenance diet mouse/rat “GLP” (Provimi Kliba SA, Kaiseraugst, Switzerland), ad libitum
- Water: ad libitum from drinking bottles
- Acclimation period: 6 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12/12 - Route of administration:
- oral: gavage
- Vehicle:
- water
- Remarks:
- tap water
- Details on exposure:
- TEST ITEM
- Dosing solution: pH 6.0 - 7.5
VEHICLE
- Amount of vehicle: 10 mL/kg bw - Details on mating procedure:
- - M/F ratio per cage: 1:1
- Length of cohabitation: maximum of 2 weeks
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged: in Makrolon type M III cages (floor area about 800 cm2)from day 18 p.c. until day 4 p.p., supplied with nesting material (cellulose wadding) toward the end of pregnancy
- Females were allowed to litter and rear their pups until day 4 after parturition - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- - The solutions were analyzed twice and determined to be 0, 1.61, 5.11, 15.73 g/100 mL using a content 68.9 g/100 g, which was determined by potentiometric titration and 0, 1.62, 4.67, 15.60 g/100 mL using 69.0 g/100 g, also determined by potentiometric titration
- Duration of treatment / exposure:
- 38 days (males), 45 days (females)
- Frequency of treatment:
- daily
- Details on study schedule:
- Premating exposure period:
- Male: 13 days
- Female: 13 days - Dose / conc.:
- 100 mg/kg bw/day (nominal)
- Remarks:
- equivalent to 67 mg/kg bw/day Isopropanolamine
- Dose / conc.:
- 300 mg/kg bw/day (nominal)
- Remarks:
- equivalent to 202 mg/kg bw/day Isopropanolamine
- Dose / conc.:
- 1 000 mg/kg bw/day (nominal)
- Remarks:
- equivalent to 673 mg/kg bw/day Isopropanolamine
- No. of animals per sex per dose:
- 12
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- - Dose selection rationale: based on range finding study
- Positive control:
- None
- Parental animals: Observations and examinations:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily (littering and lactation behaviour evaluated in the mornings
- Cage side observations included any signs of morbidity, pertinent behavioral changes and signs of overt toxicity, documented for each animal
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: prior to the administration period and thereafter at weekly intervals
- Parameters: abnormal behavior during “handling”, fur, skin, salivation, nose discharge, lacrimation, pupil size, posture, palpebral closure, respiration, tremors, convulsions, abnormal movements, impairment of gait, activity/arousal level, feces (appearance/consistency), urine, other findings
BODY WEIGHT: Yes
- Time schedule for examinations:once a week at the same time of the day (in the morning);
- Time schedule exceptions: during the mating period the parental females were weighed on the day of positive evidence day 0 p.c. and on days 7, 14 and 20 p.c.; females with litter: day day 0 p.p. and on day 4 p.p.; females without a litter: weekly
FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- once a week (in a period of 7 days) for male and female parental animals
- exceptions: not determined during the mating period; F0 females with evidence of sperm were determined on days 0, 7, 14 and 20 p.c.; F0 females, which gave birth to a litter were determined on days 0 and 4 p.c.
- not determined in females without positive evidence of sperm and females without litter
HAEMATOLOGY and CLINICAL CHEMISTRY:
- hematology (5 animals/sex/group) with EDTA-K3 as anticoagulant: Leukocytes, erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelets, differential blood count, reticulocytes
- Prothrombin time (Hepato Quick's test)
- Clinical chemistry (5 animals/sex/group): alanine aminotransferase, aspartate aminotrasferase, alkaline phosphatase, gamma-glutamyltransferase, sodium, potassium, chloride, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, cholesterol, magnesium
URINANALYSIS:
- on day 38 (males) and 45 (females) volume, color, turbidity, pH, protein, glucose, ketones, urobilinogen, bilirubin, blood, specific gravity, sediment
- Parameters: volume, color, turbidity, pH, protein, glucose, ketones, urobilinogen, bilirubin, blood, specific gravity (urine refractometer), sediment (microscopically)
Functional observation battery (FOB) and motor activity measurement:
- on study day 36 in first 5 male/group and on study day 43 in first 5 female/group - Sperm parameters (parental animals):
- Parameters examined in F0 male parental generations: testes weight, epididymis weight
- Litter observations:
- STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no
PARAMETERS EXAMINED
The following parameters were examined in F1 offspring: number and sex of pups, stillbirths, live births, postnatal mortality, weight gain (PND 1-4), presence of gross anomalies, anogenital distance (AGD)
GROSS EXAMINATION OF DEAD PUPS: yes, for external and internal abnormalities
ASSESSMENT OF DEVELOPMENTAL NEUROTOXICITY: no
ASSESSMENT OF DEVELOPMENTAL IMMUNOTOXICITY: no - Postmortem examinations (parental animals):
- SACRIFICE
- Male animals: after blood from 5 F0 males per group was sampled, on study day 38 necropsy of all male animals was performed
- Female animals: after blood from 5 F0 females per group was sampled, on study day 45 necropsy of all female animals was performed
GROSS NECROPSY
- Gross necropsy was not further specified.
HISTOPATHOLOGY / ORGAN WEIGHTS
- The tissues indicated in Table 1 were prepared for microscopic examination.
- The animals/organs were weighed: anesthetized animals, liver, kidneys, adrenal glands, testes, epididymides, seminal vesicle, prostate, ovaries, uterus, thymus, spleen, brain, heart - Postmortem examinations (offspring):
- - Organs examined at necropsy (F1 pups): all surviving pups, all stillborn pups and those pups that died before schedule
SACRIFICE
- The F1 offsprings were sacrificed on day 4 post partum.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows:
GROSS NECROPSY
- Gross necropsy consisted of external and internal (eviscerated) examinations including and their organs were assessed macroscopically. - Statistics:
- Due to limitations of characters, details for statistics are given as tables (2, 3, 4) under part 'other information'.
CLINICAL EXAMINATIONS
- DUNNETT-test (two-sided) for the hypothesis of equal means
- Pairwise comparison of each dose group with the control group using FISHER'S EXACT test for the hypothesis of equal proportions
- Pairwise comparison of each dose group with the control group using the WILCOXON-test (onesided) for the hypothesis of equal medians
- Non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using Wilcoxon-test (twosided) for the equal medians
CLINICAL PATHOLOGY
- Non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pair-wise comparison of each dose group with the control group was performed using Wilcoxon-test (two-sided) for the equal medians
- Pair-wise comparison of each dose group with the control group using FISHER's exact test for the hypothesis of equal proportions
HISTOPATHOLOGY
- Means and standard deviations AND KRUSKAL-WALLIS and WILCOXON test - Reproductive indices:
- MALE MATING AND FERTILITY INDICES
Male mating index (%) = (number of males with confirmed mating* / number of males placed with females) x 100
* defined by a female with vaginal sperm or with implants in utero
Male fertility index (%) = (number of males proving their fertility* / number of males placed with females) x 100
* defined by a female with implants in utero
FEMALE REPRODUCTION AND DELIVERY DATA
Female mating index (%) = (number of females mated* / number of females placed with males) x 100
* defined as the number of females with vaginal sperm or with implants in utero
Female fertility index (%) = (number of females pregnant* / number of females mated**) x 100
* defined as the number of females with implants in utero
** defined as the number of females with vaginal sperm or with implants in utero
Gestation index (%) = (number of females with live pups on the day of birth / number of females pregnant*) x100
* defined as the number of females with implants in utero
Live birth index (%) = (number of liveborn pups at birth / total number of pups born) x 100
Post implantation loss (%) = ([number of implantations - number of pups delivered] / number of implantations) x 100 - Offspring viability indices:
- Viability index (%) = (number of live pups on day 4 after birth / number of liveborn pups on the day of birth) x 100
Sex ratio = (number of live male or female pups on day 0/4 / number of live male and female pups on day 0/4) x 100 - Clinical signs:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - no test substance-related or spontaneous clinical findings observed in the male and female animals of 100 mg/kg body weight/day test group
- salivation after treatment was seen in all high dose male animals (1000 mg/kg body weight/day) during study weeks 1-5, finding persisted in the respective males only for a few minutes after daily gavage dosing
- urine discoloration was observed in all male animals of the high dose group during the whole study period (week 0-5) and in all mid dose male animals (300 mg/kg body weight/day) during study weeks 1-5 and females, for all high dose females during the whole study period (week 0 - 6) and in all mid dose females during study weeks 1-6
- 7 out of 12 female animals of test group 3 showed salivation after treatment during study weeks 1-6, finding persisted in the respective females only for a few minutes after daily gavage dosing
- detailed clinical observations on study days 0, 7, 14, 21, 28, 35 and additionally day 42 for female animals did not reveal any additional, substance-induced abnormalities in the male and female animals of the test groups 0-3 (0, 100, 300 and 1000 mg/kg body weight/day)
The clinical findings, i.e. transient salivation and discolored urine are considered to be substance-induced, but are without any toxicological relevance and are not assessed as being adverse.
It is assumed that the temporary salivation was induced by the bad taste of the test substance (small droplets of the test substance solution might be adjacent on the tip of the gavage) or minor local affection of the upper digestive tract (without showing a morphological correlate at pathology). The urine discoloration is possibly related to a chemical reaction of the test substance or its metabolites with the bedding or with components of the air. - Mortality:
- no mortality observed
- Description (incidence):
- There were no substance-related or spontaneous mortalities in any of the groups.
- Body weight and weight changes:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Body weights and body weight gains of all male and female (+ during the gestation, lactation period and after weaning) animals of all substance-treated groups (100, 300 and 1000 mg/kg body weight/day) were comparable to that of the concurrent control group during the whole study taking normal biological variability into account.
The statistically significantly increased body weight gains of the low and mid dose females (100 and 300 mg/kg body weight/day) during premating weeks 1-2 are considered to be spontaneous in nature and not to be adverse. - Food consumption and compound intake (if feeding study):
- no effects observed
- Description (incidence and severity):
- There were no effects observed for food consumption compared to the control group.
- Haematological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- - slightly, but statistically significantly decreased hemoglobin and hematocrit values in the peripheral blood of high dose males
- no treatment related effects were seen in the other hematology parameters of males and in the hematology investigations of females - Clinical biochemistry findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - differences in serum enzyme activities were not evident at any dose level in either males or females
- blood chemistry results showed statistically significantly increased urea concentrations in the serum of high dose males and significantly higher albumin levels in the serum of high dose females
- marginally increased urea concentrations were also seen in the high dose females
- the increase in females was not sufficient to be statistically identified
- no treatment related changes were found in the other blood chemistry parameters - Urinalysis findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - decreased amounts of urine with increased specific gravity in all animals, more pronounced in females than in males
- no treatment related effects were seen in the other urine parameters examined - Behaviour (functional findings):
- effects observed, non-treatment-related
- Description (incidence and severity):
- There were no test substance-related or spontaneous findings in the male and female animals of test groups 1-3 during FOB observations.
There were statistically significantly increased motor activity observed in males which were without a clear relation to dosing. This effect has no toxicological relevance because the values from the substance-treated males fit to the historical control range (mean value: 245.1; range: 207.6–299.5) of male rats of this strain at a comparable age.
There were no statistically significant or clearly dose-dependent deviations concerning the overall motor activity (summation of all intervals) in the females.
A substance-induced origin for this finding is excluded with certainty because of its scattered occurrence without any relation to dosing.
Thus, the motor activity of the substance-treated males and females did not show any toxicological relevant deviations. - Immunological findings:
- not examined
- Organ weight findings including organ / body weight ratios:
- effects observed, non-treatment-related
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- Further details to histopathological findings are given in the table 7.
- liver: males of the top dose group revealed a diffuse hepatocellular hypertrophy
- reproductive organs: no histopathological findings observed - Other effects:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Clinical observations during gestation / lactation:
- discolored urine was recorded for all high and mid dose females (1000 and 300 mg/kg body weight/day) during the whole gestation and lactation period
- in 9 out of 12 high dose females salivation after treatment was noted during gestation
- in 10 out of 12 high dose females salivation after treatment was noted during lactation
No test substance-related clinical findings occurred in the female animals of test group 1
(100 mg/kg body weight/day) during the lactation period.
Both findings are not assessed as adverse or toxic effects for reasons described under overall clinical findings above.
One sperm positive control female (No. 102), one sperm positive low dose female (No. 123) and one sperm positive mid dose female (No. 136) did not deliver any F1 pups. - Reproductive function: oestrous cycle:
- not examined
- Reproductive function: sperm measures:
- no effects observed
- Description (incidence and severity):
- - mean duration until sperm was detected (day 0 p.c.) amounted to 3.2 / 3.2 / 2.7 and 2.5 days (0, 100, 300 and 1000 mg/kg body weight/day)
These values reflect the normal range of biological variation inherent in the strain used in this study. - Reproductive performance:
- no effects observed
- Description (incidence and severity):
- Male and female fertility indices are given in table 8 and 9. Mating was confirmed for all F0 males placed with females.
- not generating F1 pups: one control male (No. 2 mated with female No. 102), one low dose male (No. 23 mated with female No. 123 – 100 mg/kg body weight/day) and one mid dose male (No. 36 mated with female No. 136 – 300 mg/kg body weight/day)
- female rats not becoming pregnant (but sperm positive): control female No. 102 (mated with male No. 2), low dose female No. 123 (mated with male No. 23) and mid dose female No. 136 (mated with male No. 36)
- not pregnant females failed to show a morphological correlate for the apparent infertility for females Nos. 102 (control) and 123 (low dose)
- mid dose female No. 136 had a minimal diffuse inflammation of the uterus, which may well account for the observed infertility (see also Pathology)
The fertility indices for male and female varied between 92% (0, 100 and 300 mg/kg body weight/day) and 100% (1000 mg/kg body weight/day). These values reflect the normal range of biological variation inherent in the strain of rats used for this study. All respective values were within the range of the historical control data and do not show any relation to dosing. - Dose descriptor:
- NOAEL
- Remarks:
- for systemic toxicity
- Effect level:
- 300 mg/kg bw/day
- Based on:
- test mat.
- Remarks:
- equivalent to 202 mg/kg bw/day Isopropanolamine
- Sex:
- male
- Basis for effect level:
- haematology
- Dose descriptor:
- NOAEL
- Remarks:
- for systemic toxicity
- Effect level:
- 1 000 mg/kg bw/day
- Based on:
- test mat.
- Remarks:
- equivalent to 673 mg/kg bw/day Isopropanolamine
- Sex:
- female
- Remarks on result:
- other: No effects observed for female animals.
- Dose descriptor:
- NOAEL
- Remarks:
- for reproductive performance and fertility
- Effect level:
- 1 000 mg/kg bw/day
- Based on:
- test mat.
- Remarks:
- equivalent to 673 mg/kg bw/day Isopropanolamine
- Sex:
- male/female
- Remarks on result:
- other: no substance-related effects on fertility indices observed
- Critical effects observed:
- no
- Clinical signs:
- no effects observed
- Description (incidence and severity):
- - numbers of runts: 0 /0 /0 /2 in test groups 0-3 (0, 100, 300 and 1000 mg/kg body weight/day)
This finding is fully in the range of the biological variation inherent in the strain of rats used for this study.
- no adverse clinical signs up to scheduled sacrifice on day 4 post partum - Mortality / viability:
- no mortality observed
- Description (incidence and severity):
- - viability index between days 0 - 4 p.p. varied between 98% (control) and 100% (test group 2)
- no compound related changes - Body weight and weight changes:
- no effects observed
- Description (incidence and severity):
- - 1000 mg/kg body weight/day: mean b.w. data were statistically significantly increased on post partum day 1 (males, females and males + females) and day 4 (males and males + females)
- 100 and 300 mg/kg body weight/day: mean b.w. data were comparable to the concurrent control values
- mean pup body weight changes in all test groups: did not show any statistically significant differences - Sexual maturation:
- not examined
- Organ weight findings including organ / body weight ratios:
- not examined
- Gross pathological findings:
- no effects observed
- Histopathological findings:
- not examined
- Other effects:
- no effects observed
- Description (incidence and severity):
- Sex ratio:
- sex distribution and sex ratios of live F1 pups on the day of birth and 4 days post partum did not show biologically relevant differences between controls and test groups - Dose descriptor:
- NOAEL
- Generation:
- F1
- Effect level:
- 1 000 mg/kg bw/day
- Based on:
- test mat.
- Remarks:
- equivalent to 673 mg/kg bw/day Isopropanolamine
- Sex:
- male/female
- Remarks on result:
- other: No test substance-induced signs of developmental toxicity were noted in the progeny of the F0 parents up to and including 1000 mg/kg body weight/day.
- Critical effects observed:
- no
- Reproductive effects observed:
- no
- Conclusions:
- - The NOAEL for reproductive performance and fertility is 1000 mg/kg bw/day (equivalent to 673 mg/kg bw/day Isopropanolamine) for the F0 parental rats
- The NOAEL for general, systemic toxicity is 300 mg/kg body weight/day (equivalent to 202 mg/kg bw/day Isopropanolamine) for the F0 parental males based on some indications of a mild anemic process and 1000 mg/kg body weight/day for the F0 parental females - Executive summary:
The hydrochloride salt of Isopropanolamine was administered orally via gavage to groups of 12 male and 12 female Wistar rats (F0 animals) at doses of 100 (673 mg/kg bw/day Isopropanolamine), 300 (202
mg/kg bw/day Isopropanolamine) and 1000 mg/kg of body weight/day (673 mg/kg bw/day Isopropanolamine) in order to detect possible effects of the test substance on the integrity and performance of the reproductive system of both sexes. Furthermore, it was intended to obtain information about the general toxicological profile including target organs and no observed adverse effect level (NOAEL) after repeated oral administration, according to the OECD 422 guideline. Control animals were dosed daily with the vehicle (tap water).
The duration of treatment covered a 2 week premating period and mating period in both sexes, approximately 2 weeks post-mating in males, and the entire gestation period and 4 days of lactation in females.
F0 animals were mated 13 days after the beginning of treatment to produce a litter (F1 generation pups). Mating pairs were from the same dose group. Mating was discontinued as soon as sperm was detected in the vaginal smear, mating was discontinued. F0 animals were examined for their reproductive performance including determination of the number of implantation sites and the calculation of postimplantation loss for all F0 females.
A detailed clinical observation was performed in all animals before test substance administration and thereafter at weekly intervals. Food consumption of the F0 parents was determined regularly during premating and after the mating period and during the gestation and lactation periods in dams. In general, body weights of F0 animals were determined once a week; however, during gestation and lactation, F0 females were weighed on days 0, 7, 14 and 20 post coitum, on the parturition day and day 4 post partum. Pups were sexed on day 0 post partum and weighed one day after birth and on day 4 post partum. Their viability was recorded twice daily on each workday or only in the morning on Saturday and Sunday. All pups were sacrificed with CO2 on day 4 post partum and examined macroscopically for external and visceral findings.
Near the end of the administration period a functional observational battery was performed and motor activity was measured in 5 randomly selected parental males and females per group, respectively. Blood was sampled from 5 randomly selected parental males and 5 parental females per group for hematological and clinical chemistry examination. All surviving F0 parental animals were sacrificed by decapitation, under CO2 anesthesia, and were assessed by gross pathology. Organ weights were recorded and a histopathological examination was performed.
The following test substance-related, adverse effects/findings were noted:
- 1000 mg/kg body weight/day:
F0 parental animals: statistically significantly reduced hemoglobin and hematocrit values (i.e. indications of a mild anemic process) in the F0 males
F1 pups: no adverse effects/findings
- 100 and 300 mg/kg body weight/day:
F0 parental animals and F1 pups: no adverse effects/findings
Thus, under the conditions of this reproduction/developmental toxicity screening test the NOAEL (no observed adverse effect level) for reproductive performance and fertility is 1000 mg/kg body weight/day
(673 mg/kg bw/day Isopropanolamine) for the F0 parental rats. The NOAEL for general, systemic toxicity of the test substance is 300 mg/kg body weight/day (202 mg/kg bw/day Isopropanolamine) for the F0 parental males based on some indications for a mild anemic process.
Referenceopen allclose all
Table 1: Mean absolute organ weights (g) from male F1 rats
GROUP |
CONC.(PPM) |
LIVER |
KIDNEYS |
HEART |
ADRENALS |
BRAIN |
TESTES |
I-1 |
0 |
20.327 (2.457) |
3.679 (0.388) |
1.584 (0.120) |
0.066 (0.011) |
2.129 (0.191) |
3.513 (0.231) |
III-1 |
500 |
21.192 (2.372) |
3.821 (0.478) |
1.577 (0.148) |
0.059 (0.012) |
2.114 (0.133) |
3.484 (0.231) |
V-1 |
2000 |
22.022 (3.142) |
3.900 (0.391) |
1.688 (0.175) |
0.066 (0.012) |
2.200 (0.151) |
3.617 (0.315) |
VII-1 |
7500 |
21.743 (3.262) |
3.924 (0.498) |
1.626 (0.167) |
0.067 (0.019) |
2.181 (0. 143) |
3.529 (0.417) |
Table 2: Mean relative organ weights (% of body weight) from male F1 rats
GROUP |
CONC.(PPM) |
LIVER |
KIDNEYS |
HEART |
ADRENALS |
BRAIN |
TESTES |
I-1 |
0 |
3.7428 (.2441) |
0.6792 (.0524) |
0.2932 (.0235) |
0.0122 (.0019) |
0.3958 (.0501) |
0.6518 (.0642) |
III-1 |
500 |
3.8119 (.3235) |
0.6874 (.0705) |
0.2841 (.0237) |
0.0106 (.0021) |
0.3818 (.0326) |
0.6284 (.0426) |
V-1 |
2000 |
3.7627 (.3125) |
0.6715 (.0746) |
0.2898 (.0224) |
0.0114 (.0021) |
0.3794 (.0381) |
0.6251 (.0822) |
VII-1 |
7500 |
3.8580 (.3470) |
0.6987 (.0640) |
0.2902 (.0264) |
0.0119 (.0035) |
0.3902 (.0327) |
0.6302 (.0722) |
Table 3: Mean absolute organ weights (g) from female F1 rats
GROUP |
CONC. (PPM) |
LIVER |
KIDNEYS |
HEART |
ADRENALS |
BRAIN |
II-1 |
0 |
9.623(1.428) |
2.212 (0.232) |
1.023 (0.125) |
0.076 (0.010) |
1.995 (0.088) |
IV-1 |
500 |
9.586(1.455) |
2.167 (0.266) |
1.029(0.112) |
0.073 (0.012) |
1.984 (0.064) |
VI-1 |
2000 |
9.979(1.485) |
2.301 (0.201) |
1.073 (0.097) |
0.078 (0.014) |
1.969 (0.075) |
VIII-1 |
7500 |
9.596(1.242) |
2.255 (0.216) |
1.027 (O.097) |
0.075 (0.014) |
1.930 (0.185) |
Table 4: Mean absolute organ weights (% of body weight) from female F1 rats
GROUP |
CONC. (PPM) |
LIVER |
KIDNEYS |
HEART |
ADRENALS |
BRAIN |
II-1 |
0 |
3.3253 (.2382) |
0.7685 (.0638) |
0.3552 (.0349) |
0.0265 (.0048) |
0.6982(.0758) |
IV-1 |
500 |
3.3637(.2409) |
0.7627 (.0521) |
0.3631 (.0300) |
0.0256 (.0031) |
0.7051 (.0715) |
VI-1 |
2000 |
3.2420(.3182) |
0.7539 (.0889) |
0.3503 (.0284) |
0.0255 (.0045) |
0.6458(.0654) |
VIII-1 |
7500 |
3.4011 (.2820) |
0.8009 (.0562) |
0.3659 (.0381) |
0.0269 (.0053) |
0.6888 (.0824) |
Standard deviation in paranthesis
+ - Significantly different (P<0.05) from control group by LSD (least standard deviation)
# - Significantly different (P<0.05) from control group LSD and Dunnet's test
No treatment-related effects were noted in hematology investigations of females and serum enzyme examinations of both sexes.
The most prominent findings in clinical pathology were the reduced amounts of urine with subsequently increased specific gravity excreted by treated animals of either sex. The decreased urinary volume and the increased urinary specific gravity, however, are not considered as markers of kidney toxicity or an impairment of renal function. These findings are regarded non-renal in nature and are possibly due to decreased water intake.
A reduction in water intake could also well account for the slightly increased urea and albumin levels of the high dose males and/or females.
It is concluded that the changes in urinalysis and in blood chemistry examination are not caused by a direct toxic effect of the test compound and are not adverse in nature. This assessment is supported by the fact, that absolute and relative kidney weights as well as gross and histopathological evaluations of this organ did not give any indications for substance-induced alterations.
The livers of female animals of the top dose did not reveal any histological findings. Nevertheless, the increase in liver weight in the top dose females is thought to be substance-related.
These liver findings are regarded to be non-adverse in nature, but are considered to mirror adaptive responses to the test substance administration. Moreover, liver enzymes in these rats remained unaffected (see clinical biochemistry), but showed the usual range of biological variation.
The fertility indices observed in the study reflect the normal range of biological variation inherent in the strain of rats used for this study as all respective values were within the range of the historical control data. Furthermore, gross and histopathological examinations of these males failed to show a relevant morphological correlate for the apparently impaired fertility (see also Pathology).
Table 5: Mean absolute weights of statistically significant changes in organs.
Absolute Weights |
Male |
Female |
||||
Group |
1 |
2 |
3 |
1 |
2 |
3 |
Liver |
+3% |
-3% |
+22%* |
-4% |
-4% |
+17%* |
Brain |
- |
- |
- |
0% |
+4%* |
-1% |
Adrenal Glands |
+4% |
+4% |
+19%* |
- |
- |
- |
Thymus |
- |
- |
- |
0% |
+28%* |
+26%* |
*values were statistically significant different, P = 0.05 |
||||||
Table 6: Relative absolute weights of statistically significant changes in organs.
Relative Weights |
Male |
Female |
||||
Group |
1 |
2 |
3 |
1 |
2 |
3 |
Liver |
+3% |
+1% |
+23%* |
-2% |
0% |
+16%* |
Brain |
- |
- |
- |
+2% |
+8% |
-2% |
Adrenal Glands |
0% |
+5% |
+15%* |
- |
- |
- |
Thymus |
- |
- |
- |
+3% |
+33%* |
+25%** |
Kidneys |
- |
- |
- |
+3% |
+9%* |
+4% |
*values were statistically significant different, P <= 0.001, (**p= 0.05) |
||||||
Table 7: Histopathological findings in the liver
Liver |
Male animals |
Female animals |
||||||
Group |
0 |
1 |
2 |
3 |
0 |
1 |
2 |
3 |
Organs examined |
12 |
12 |
12 |
12 |
12 |
|
|
12 |
Hypertrophy, diffuse |
0 |
0 |
0 |
9 |
0 |
|
|
0 |
Table 8: Male fertility indices (F0 males) in %:
|
Test group 0 (0 mg/kg bw/day) |
Test group 1 (100 mg/kg bw/day) |
Test group 2 (300 mg/kg bw/day) |
Test group 3 (1000 mg/kg bw/day) |
concerning F1 litters |
92 |
92 |
92 |
100 |
Table 9: Female fertility indices (F0 females) in %:
|
Test group 0 (0 mg/kg bw/day) |
Test group 1 (100 mg/kg bw/day) |
Test group 2 (300 mg/kg bw/day) |
Test group 3 (1000 mg/kg bw/day) |
concerning F1 litters |
92 |
92 |
92 |
100 |
Effect on fertility: via oral route
- Endpoint conclusion:
- no adverse effect observed
Effect on fertility: via inhalation route
- Endpoint conclusion:
- no study available
Effect on fertility: via dermal route
- Endpoint conclusion:
- no study available
Additional information
An EOGRTS with MIPA is currently ongoing. Based on the available reliable data, it is concluded in a weight-of-evidence approach, that MIPA does not induce effects on fertility (see IUCLID section 13.2: "Justification_WoE_RDT_Reprotox_Oct2022").
Screening study for reproduction/developmental toxicity (OECD 422) in rats
In a combined repeated dose oral toxicity study with a reproduction/developmental screening study (according to OECD guideline 422 and under GLP), Wistar rats (12/sex/dose) were administerd the hydrochloride salt of MIPA (CAS no. 7780 -04 -3) at 0, 100, 300 or 1000 mg/kg bw/day (equivalent to 67, 202 and 673 mg/kg bw/day MIPA) by oral gavage. The duration of treatment covered a 2-week premating period and mating period in both sexes, approximately 2 weeks post-mating in males, and the entire gestation period and 4 days of lactation in females, resulting in a total of 38 and 45 exposure days for males and females, respectively. Clinical pathology examinations revealed slightly, but statistically significant reduced hemoglobin and hematocrit values, which are indicative of a mild anemic process, in high dose males. Other (minor) clinical and pathological findings appear to be incidental and not dose-related. Based on the mild anemic process in males, the NOAEL for systemic toxicity is 300 mg/kg bw/day. The fertility indices for male and female varied between 92% (0, 100 and 300 mg/kg body weight/day) and 100% (1000 mg/kg body weight/day). These values reflect the normal range of biological variation inherent in the strain of rats used for this study. All respective values were within the range of the historical control data and do not show any relation to dosing.
No adverse effects were found on reproductive and fertility parameters, thus a NOAEL of 1000 mg/kg bw/day (equivalent to 673 mg/kg bw/day MIPA, the highest dose tested) was established for effects on fertility in males and females.
Oral one generation study in rats
In a one-generation study according to FDA guidelines, Sprague-Dawley rats (25/sex/dose) were administered TIPA in the diet at 0, 500, 2000 or 7500 ppm (equivalent to a 0, 15.6, 63 or 239 mg/kg bw/d intake of MIPA in males or 0, 17.2, 71 or 275 mg/kg bw/d in females) for 5 weeks prior to mating as well as during mating, gestation and lactation. Offspring (20/sex/dose) were also administered the same doses for 90 days after weaning. Prior to weaning, these rats were exposed to TIPA in utero during pregnancy of the P rats, in maternal milk during lactation, and in any diet they consumed prior to weaning. Ophthalmological, clinical chemistry, haematology and urinalysis examinations were conducted. Histopathological examination was conducted on controls and high-dose animals for all organs and on lungs, liver, kidneys and organs with lesions at the low and intermediate doses. No adverse clinical, histological, or reproductive effects (mating index, fertility index, gestation index, gestation length, percent pups born alive, viability index, lactation index, litter survival, average number ofpups/litter, litter size) were noted.
The NOAEL was reported to be 609 mg/kg bw/d TIPA (equivalent to a 239 mg/kg bw/d intake of MIPA in males or 275 mg/kg bw/d in females) for males and 700 mg/kg bw/d TIPA for females, the highest doses tested.
Effects on developmental toxicity
Description of key information
Maternal and developmental NOAEL in rats = 1000 mg/kg bw/d (OECD414, gavage).
Furthermore, no adverse effects were observed in the OECD414 studies in rats with the structural analogues DIPA and TIPA up to the limit dose. Supportingly, in an OECD422 study in rats exposed to MIPA HCl salt by gavage, a NOAEL for developmental toxicity of 1000 mg/kg bw/d (equivalent to 673 mg/kg bw/d MIPA) was established. Based on the mild anemic process in males, the NOAEL for systemic toxicity is 300 mg/kg bw/day.
Developmental toxicity data for analogon TIPA in the second species rabbit (OECD414 with Picloram TIPA salt and sub-chronic dose range finder with TIPA) also revealed no adverse developmental effects but showed, that there is a MTD (maximal tolerable dose) for the dams below the limit dose.
Link to relevant study records
- Endpoint:
- developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- ADAPTATION ACCORDING TO REACH ANNEX XI, section 1 - see justifications attached to IUCLID section 13.2.
- Justification WoE_RDT_Reprotox_Oct2022
- Justification RA_CAS 122-20-2_Dev.tox.2ndspecies_Nov2021 - Reason / purpose for cross-reference:
- reference to same study
- Reason / purpose for cross-reference:
- reference to same study
- Qualifier:
- according to guideline
- Guideline:
- other: OECD Test Guideline 422 - Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxicity Screening Test
- Version / remarks:
- 22 March 1996
- Deviations:
- no
- GLP compliance:
- yes
- Limit test:
- no
- Specific details on test material used for the study:
- SOURCE OF TEST MATERIAL
- Source and batch No.of test material: BASF and BA 1449
- Purity test date: 04 Jan 2006
- concetration of given stock solution:65.4 g isopropanolamine hydrochloride per 100 g aqueous solution
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature (freezer for reanalysis)
- Stability under test conditions: confirmed by reanalysis
- Solubility and stability of the test substance in the solvent/vehicle: stability of aq. solution confirmed by reanalysis
TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: test material was weighed up and dissolved in tap water
FORM AS APPLIED IN THE TEST: solution - Species:
- rat
- Strain:
- Wistar
- Remarks:
- Crl:WI(Han)
- Details on test animals or test system and environmental conditions:
- - Source: Charles River Laboratories, Research Models and Services, Germany Gmbh
- Age at study initiation: 11-13 weeks
- Weight at study initiation: 267.4-309.0 g (males) and 186.7-217.7 g (females)
- Number of animals: 96 (12 per sex per dose group)
- Housing: Individually in type DK III stainless steel wire mesh cages supplied by Becker & Co., Castrop-Rauxel, Germany (floor area of about 800 cm2), with the following exceptions: for the overnight mating the females were put into the cages of the males
- Diet: Ground Kliba maintenance diet mouse/rat “GLP” (Provimi Kliba SA, Kaiseraugst, Switzerland), ad libitum
- Water: ad libitum from drinking bottles
- Acclimation period: 6 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12/12 - Route of administration:
- oral: gavage
- Vehicle:
- water
- Remarks:
- tap water
- Details on exposure:
- TEST ITEM
- Dosing solution: pH 6.0 - 7.5
VEHICLE
- Amount of vehicle: 10 mL/kg bw - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- - The solutions were analyzed twice and determined to be 0, 1.61, 5.11, 15.73 g/100 mL using a content 68.9 g/100 g, which was determined by potentiometric titration and 0, 1.62, 4.67, 15.60 g/100 mL using 69.0 g/100 g, also determined by potentiometric titration
- Details on mating procedure:
- - Impregnation procedure: cohoused
- M/F ratio per cage: 1:1
- Length of cohabitation: maximum of 2 weeks
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy and the following day "day 1" post coitum
- After successful mating each pregnant female was caged: in Makrolon type M III cages (floor area about 800 cm2)from day 18 p.c. until day 4 p.p., supplied with nesting material (cellulose wadding) toward the end of pregnancy
- Females were allowed to litter and rear their pups until day 4 after parturition - Duration of treatment / exposure:
- 38 days (males), 45 days (females)
- Frequency of treatment:
- daily
- Duration of test:
- 39-47 days
- Dose / conc.:
- 100 mg/kg bw/day (nominal)
- Remarks:
- equivalent to 67 mg/kg bw/day Isopropanolamine
- Dose / conc.:
- 300 mg/kg bw/day (nominal)
- Remarks:
- equivalent to 202 mg/kg bw/day Isopropanolamine
- Dose / conc.:
- 1 000 mg/kg bw/day (nominal)
- Remarks:
- equivalent to 673 mg/kg bw/day Isopropanolamine
- No. of animals per sex per dose:
- 12
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- - Dose selection rationale: based on range finding study
- Maternal examinations:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily (littering and lactation behaviour evaluated in the mornings
- Cage side observations included any signs of morbidity, pertinent behavioral changes and signs of overt toxicity, documented for each animal
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: prior to the administration period and thereafter at weekly intervals
- Parameters: abnormal behavior during “handling”, fur, skin, salivation, nose discharge, lacrimation, pupil size, posture, palpebral closure, respiration, tremors, convulsions, abnormal movements, impairment of gait, activity/arousal level, feces (appearance/consistency), urine, other findings
BODY WEIGHT: Yes
- Time schedule for examinations:once a week at the same time of the day (in the morning);
- Time schedule exceptions: during the mating period the parental females were weighed on the day of positive evidence day 0 p.c. and on days 7, 14 and 20 p.c.; females with litter: day day 0 p.p. and on day 4 p.p.; females without a litter: weekly
FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- once a week (in a period of 7 days) for male and female parental animals
- exceptions: not determined during the mating period; F0 females with evidence of sperm were determined on days 0, 7, 14 and 20 p.c.; F0 females, which gave birth to a litter were determined on days 0 and 4 p.c.
- not determined in females without positive evidence of sperm and females without litter
POST-MORTEM EXAMINATIONS: Yes
SACRIFICE
- Male animals: after blood from 5 F0 males per group was sampled, on study day 38 necropsy of all male animals was performed
- Female animals: after blood from 5 F0 females per group was sampled, on study day 45 necropsy of all female animals was performed
GROSS NECROPSY
- Gross necropsy was not further specified.
HISTOPATHOLOGY / ORGAN WEIGHTS
- The tissues indicated in Table 1 were prepared for microscopic examination.
- The animals/organs were weighed: anesthetized animals, liver, kidneys, adrenal glands, testes, epididymides, seminal vesicle, prostate, ovaries, uterus, thymus, spleen, brain, heart
OTHER:
HAEMATOLOGY and CLINICAL CHEMISTRY:
- hematology (5 animals/sex/group) with EDTA-K3 as anticoagulant: Leukocytes, erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelets, differential blood count, reticulocytes
- Prothrombin time (Hepato Quick's test)
- Clinical chemistry (5 animals/sex/group): alanine aminotransferase, aspartate aminotrasferase, alkaline phosphatase, gamma-glutamyltransferase, sodium, potassium, chloride, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, cholesterol, magnesium - Ovaries and uterine content:
- The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: No
- Number of implantations: Yes
- Number of early resorptions: No
- Number of late resorptions: No - Fetal examinations:
- For all surviving pups (after sacrifice on day 4 post partum), all stillborn pups and those pups that died before schedule:
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: all per litter
- Skeletal examinations: No
- Head examinations: No - Statistics:
- Due to limitations of characters, details for statistics are given as tables (2, 3, 4) under part 'other information'.
CLINICAL EXAMINATIONS
- DUNNETT-test (two-sided) for the hypothesis of equal means
- Pairwise comparison of each dose group with the control group using FISHER'S EXACT test for the hypothesis of equal proportions
- Pairwise comparison of each dose group with the control group using the WILCOXON-test (onesided) for the hypothesis of equal medians
- Non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using Wilcoxon-test (twosided) for the equal medians
CLINICAL PATHOLOGY
- Non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). If the resulting p-value was equal or less than 0.05, a pair-wise comparison of each dose group with the control group was performed using Wilcoxon-test (two-sided) for the equal medians
- Pair-wise comparison of each dose group with the control group using FISHER's exact test for the hypothesis of equal proportions
HISTOPATHOLOGY
- Means and standard deviations AND KRUSKAL-WALLIS and WILCOXON test - Indices:
- FEMALE REPRODUCTION AND DELIVERY DATA
Gestation index (%) = (number of females with live pups on the day of birth / number of females pregnant*) x100
* defined as the number of females with implants in utero
Live birth index (%) = (number of liveborn pups at birth / total number of pups born) x 100
Post implantation loss (%) = ([number of implantations - number of pups delivered] / number of implantations) x 100
OFFSPRING VIABILITY INDICES
Viability index (%) = (number of live pups on day 4 after birth / number of liveborn pups on the day of birth) x 100
Sex ratio = (number of live male or female pups on day 0/4 / number of live male and female pups on day 0/4) x 100 - Historical control data:
- There are historical control data from various studies with the same species and strain available in-house.
The included paramters are among others: mean maternal body weights during gestation and lactation, reproduction and litter indices as indicated before, pup weights and pup necropsy data and motor activity observations. - Clinical signs:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - no test substance-related or spontaneous clinical findings observed in the male and female animals of 100 mg/kg body weight/day test group
- salivation after treatment was seen in all high dose male animals (1000 mg/kg body weight/day) during study weeks 1-5, finding persisted in the respective males only for a few minutes after daily gavage dosing
- urine discoloration was observed in all male animals of the high dose group during the whole study period (week 0-5) and in all mid dose male animals (300 mg/kg body weight/day) during study weeks 1-5 and females, for all high dose females during the whole study period (week 0 - 6) and in all mid dose females during study weeks 1-6
- 7 out of 12 female animals of test group 3 showed salivation after treatment during study weeks 1-6, finding persisted in the respective females only for a few minutes after daily gavage dosing
- detailed clinical observations on study days 0, 7, 14, 21, 28, 35 and additionally day 42 for female animals did not reveal any additional, substance-induced abnormalities in the male and female animals of the test groups 0-3 (0, 100, 300 and 1000 mg/kg body weight/day)
The clinical findings, i.e. transient salivation and discolored urine are considered to be substance-induced, but are without any toxicological relevance and are not assessed as being adverse.
It is assumed that the temporary salivation was induced by the bad taste of the test substance (small droplets of the test substance solution might be adjacent on the tip of the gavage) or minor local affection of the upper digestive tract (without showing a morphological correlate at pathology). The urine discoloration is possibly related to a chemical reaction of the test substance or its metabolites with the bedding or with components of the air. - Mortality:
- no mortality observed
- Description (incidence):
- There were no substance-related or spontaneous mortalities in any of the groups.
- Body weight and weight changes:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Body weights and body weight gains of all male and female (+ during the gestation, lactation period and after weaning) animals of all substance-treated groups (100, 300 and 1000 mg/kg body weight/day) were comparable to that of the concurrent control group during the whole study taking normal biological variability into account.
The statistically significantly increased body weight gains of the low and mid dose females (100 and 300 mg/kg body weight/day) during premating weeks 1-2 are considered to be spontaneous in nature and not to be adverse. - Food consumption and compound intake (if feeding study):
- no effects observed
- Description (incidence and severity):
- There were no effects observed for food consumption compared to the control group.
- Haematological findings:
- effects observed, treatment-related
- Description (incidence and severity):
- - slightly, but statistically significantly decreased hemoglobin and hematocrit values in the peripheral blood of high dose males
- no treatment related effects were seen in the other hematology parameters of males and in the hematology investigations of females - Clinical biochemistry findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - differences in serum enzyme activities were not evident at any dose level in either males or females
- blood chemistry results showed statistically significantly increased urea concentrations in the serum of high dose males and significantly higher albumin levels in the serum of high dose females
- marginally increased urea concentrations were also seen in the high dose females
- the increase in females was not sufficient to be statistically identified
- no treatment related changes were found in the other blood chemistry parameters - Urinalysis findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - decreased amounts of urine with increased specific gravity in all animals, more pronounced in females than in males
- no treatment related effects were seen in the other urine parameters examined - Behaviour (functional findings):
- effects observed, non-treatment-related
- Description (incidence and severity):
- There were no test substance-related or spontaneous findings in the male and female animals of test groups 1-3 during FOB observations.
There were statistically significantly increased motor activity observed in males which were without a clear relation to dosing. This effect has no toxicological relevance because the values from the substance-treated males fit to the historical control range (mean value: 245.1; range: 207.6–299.5) of male rats of this strain at a comparable age.
There were no statistically significant or clearly dose-dependent deviations concerning the overall motor activity (summation of all intervals) in the females.
A substance-induced origin for this finding is excluded with certainty because of its scattered occurrence without any relation to dosing.
Thus, the motor activity of the substance-treated males and females did not show any toxicological relevant deviations. - Immunological findings:
- not examined
- Organ weight findings including organ / body weight ratios:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Detailed information on changes (absolute and relative) of organ weights are given in the table 5 and 6 under any other information.
ABSOLUTE WEIGHT EFFECTS
- statistically increased brain weight of the mid dose group of female animals
This effect is thought to be incidental in nature, as no relevant microscopic findings were noted and no dose response relationship was observed.
- statistically increased thymus weights of females of the mid and top dose group
This effect is also thought to be incidental as no microscopic findings were noted.
RELATIVE WEIGHT EFFECTS
- statistically significant increased brain and kidney weights of females of the mid dose group
This effect was considered incidental in nature, as there was no dose-response relationship and no relevant microscopic findings were noted.
- statistically increased thymus weights of females of the mid and top dose group
This effect is also thought to be incidental as no microscopic findings were noted. - Gross pathological findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - additional statistically significant intergroup differences in the results of clinical pathology testing
- these deviations were marginal, incidental or inconsistent, when compared with the other sex, and/or lack dose-response relationship
These findings were considered to be of no toxicological significance. - Neuropathological findings:
- not examined
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- Further details to histopathological findings are given in the table 7.
- liver: males of the top dose group revealed a diffuse hepatocellular hypertrophy
- reproductive organs: no histopathological findings observed - Other effects:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Clinical observations during gestation / lactation:
- discolored urine was recorded for all high and mid dose females (1000 and 300 mg/kg body weight/day) during the whole gestation and lactation period
- in 9 out of 12 high dose females salivation after treatment was noted during gestation
- in 10 out of 12 high dose females salivation after treatment was noted during lactation
No test substance-related clinical findings occurred in the female animals of test group 1 (100 mg/kg body weight/day) during the lactation period.
Both findings are not assessed as adverse or toxic effects for reasons described under overall clinical findings above. - Number of abortions:
- not specified
- Pre- and post-implantation loss:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - postimplantation loss in the high dose group slightly exceeded the historical control range, however, the mean number of F1 pups delivered per dam remained unaffected (12.9 / 11.6 / 10.7 and 11.5 pups/dam at 0, 100, 300 and 1000 mg/kg body weight/day)
- no statistically significant differences between the groups for the postimplantation loss - Dead fetuses:
- no effects observed
- Description (incidence and severity):
- - rate of liveborn pups: live birth indices ranging between 99% (high and low dose) and 100% (control and mid dose)
- rate of stillborn pups was comparable between the groups - Changes in number of pregnant:
- no effects observed
- Description (incidence and severity):
- - gestation index reached 100% in all test groups
- Other effects:
- no effects observed
- Description (incidence and severity):
- - mean number of implantation sites was comparable between all test groups including the controls and did not show any relation to dosing (13.6 / 12.4 / 11.8 and 13.2 implants/dam in test groups 0 - 3 (0, 100, 300 and 1000 mg/kg body weight/day))
- Details on maternal toxic effects:
- Hematology determinations in high dose males revealed slightly, but statistically significantly reduced hemoglobin and hematocrit values, which are indicative of a mild anemic process. This is considered as an adverse substance-induced effect.
No treatment-related effects were noted in hematology investigations of females and serum enzyme examinations of both sexes.
The most prominent findings in clinical pathology were the reduced amounts of urine with subsequently increased specific gravity excreted by treated animals of either sex. The decreased urinary volume and the increased urinary specific gravity, however, are not considered as markers of kidney toxicity or an impairment of renal function. These findings are regarded non-renal in nature and are possibly due to decreased water intake.
A reduction in water intake could also well account for the slightly increased urea and albumin levels of the high dose males and/or females.
It is concluded that the changes in urinalysis and in blood chemistry examination are not caused by a direct toxic effect of the test compound and are not adverse in nature. This assessment is supported by the fact, that absolute and relative kidney weights as well as gross and histopathological evaluations of this organ did not give any indications for substance-induced alterations.
The livers of female animals of the top dose did not reveal any histological findings. Nevertheless, the increase in liver weight in the top dose females is thought to be substance-related.
These liver findings are regarded to be non-adverse in nature, but are considered to mirror adaptive responses to the test substance administration. Moreover, liver enzymes in these rats remained unaffected (see clinical biochemistry), but showed the usual range of biological variation.
Considering the unaffected implantation and litter size and the absence of any statistically significant differences between the groups for the postimplantation loss, no test substance-induced intrauterine embryo-/fetolethality is assumed. The rate of liveborn pups was also not affected by the test substance administration as indicated by live birth indices ranging between 99% (high and low dose) and 100% (control and mid dose). Moreover, the rate of stillborn pups was comparable between the groups.
Thus, the administration of the Hydrochloride salt of Isopropanolamine did not adversely affect reproduction and delivery of the F0 females. - Dose descriptor:
- NOAEL
- Remarks:
- for systemic toxicity
- Effect level:
- 300 mg/kg bw/day (nominal)
- Based on:
- test mat.
- Remarks:
- equivalent to 202 mg/kg bw/day Isopropanolamine
- Basis for effect level:
- haematology
- Dose descriptor:
- NOAEL
- Remarks:
- for systemic toxicity
- Effect level:
- 1 000 mg/kg bw/day (nominal)
- Based on:
- test mat.
- Remarks:
- equivalent to 673 mg/kg bw/day Isopropanolamine
- Remarks on result:
- other: No effects observed for female animals.
- Dose descriptor:
- NOAEL
- Remarks:
- for developmental toxicity
- Effect level:
- 1 000 mg/kg bw/day (nominal)
- Based on:
- test mat.
- Remarks:
- equivalent to 673 mg/kg bw/day Isopropanolamine
- Remarks on result:
- other: No effects observed for maternal developmental toxicity
- Abnormalities:
- no effects observed
- Fetal body weight changes:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - mean pup body weights in test group 3 (1000 mg/kg body weight/day) were statistically significantly increased on post partum day 1 (males, females and males + females) and day 4 (males and males + females)
- mean pup body weights in test groups 1 and 2; 100 and 300 mg/kg body weight/day were comparable to the concurrent control values
- mean pup body weight changes in test groups 1, 2 and 3; 100, 300 and 1000 mg/kg body weight/day did not show any statistically significant differences
- body weight and body weight changes: spontaneous in nature, are not of toxicological relevance and are not considered as adverse effects - Reduction in number of live offspring:
- no effects observed
- Description (incidence and severity):
- - mean number of delivered pups per dam and the rate of liveborn and stillborn pups were evenly distributed among the groups
- Changes in sex ratio:
- no effects observed
- Description (incidence and severity):
- - sex distribution and sex ratios of live F1 pups on the day of birth and 4 days post partum did not show biologically relevant differences between controls and test groups
- Changes in litter size and weights:
- no effects observed
- Description (incidence and severity):
- - unaffected litter size between the groups
- Changes in postnatal survival:
- no effects observed
- Description (incidence and severity):
- - viability index as indicator for pup mortality between days 0 - 4 p.p. varied between 98% (control) and 100% (test group 2)
- no compound related changes - External malformations:
- no effects observed
- Description (incidence and severity):
- - numbers of runts were 0 /0 /0 /2 in test groups 0-3 (0, 100, 300 and 1000 mg/kg body weight/day), fully in the range of the biological variation inherent in the strain of rats
- scattered occurrence without a clear relation to dosing of post mortem autolysis - Visceral malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - very few findings as situs inversus, misshapen spleen, infarct of liver, empty stomach, and dilated renal pelvis were of scattered occurrence without a clear relation to dosing
- substance-induced origin for these findings is excluded due to existence in the historical control data at comparable or even higher incidences - Details on embryotoxic / teratogenic effects:
- No test substance-induced signs of developmental toxicity were noted in the progeny of the F0 parents up to and including 1000 mg/kg body weight/day. The number of delivered F1 pups/litter, their postnatal survival and their body weights remained unaffected by the test substance. Clinical and/or gross necropsy examinations of the F1 pups revealed only findings, which were considered to be spontaneous in nature.
- Dose descriptor:
- NOAEL
- Effect level:
- 1 000 mg/kg bw/day (nominal)
- Based on:
- test mat.
- Remarks:
- equivalent to 673 mg/kg bw/day Isopropanolamine
- Sex:
- male/female
- Remarks on result:
- other: No effects observed for F1 animals.
- Abnormalities:
- no effects observed
- Developmental effects observed:
- no
- Conclusions:
- - Only the findings of decreased hemoglobin and hematocrit at 1000 mg/kg bw/day (equivalent to 673 mg/kg bw/day Isopropanolamine) in males are considered compound-related
- The other clinical and pathological findings appear to be incidental and not dose-related.
- The NOAEL for developmental toxicity in the F1 progeny is 1000 mg/kg bw/day. - Executive summary:
The hydrochloride salt of Isopropanolamine was administered orally via gavage to groups of 12 male and 12 female Wistar rats (F0 animals) at doses of 100 (673 mg/kg bw/day Isopropanolamine), 300 (202
mg/kg bw/day Isopropanolamine) and 1000 mg/kg of body weight/day (673 mg/kg bw/day Isopropanolamine) in order to detect possible effects of the test substance on the integrity and performance of the reproductive system of both sexes. Furthermore, it was intended to obtain information about the general toxicological profile including target organs and no observed adverse effect level (NOAEL) after repeated oral administration, according to the OECD 422 guideline. Control animals were dosed daily with the vehicle (tap water).
The duration of treatment covered a 2 week premating period and mating period in both sexes, approximately 2 weeks post-mating in males, and the entire gestation period and 4 days of lactation in females.
F0 animals were mated 13 days after the beginning of treatment to produce a litter (F1 generation pups). Mating pairs were from the same dose group. Mating was discontinued as soon as sperm was detected in the vaginal smear, mating was discontinued. F0 animals were examined for their reproductive performance including determination of the number of implantation sites and the calculation of postimplantation loss for all F0 females.
A detailed clinical observation was performed in all animals before test substance administration and thereafter at weekly intervals. Food consumption of the F0 parents was determined regularly during premating and after the mating period and during the gestation and lactation periods in dams. In general, body weights of F0 animals were determined once a week; however, during gestation and lactation, F0 females were weighed on days 0, 7, 14 and 20 post coitum, on the parturition day and day 4 post partum. Pups were sexed on day 0 post partum and weighed one day after birth and on day 4 post partum. Their viabilitys recorded twice daily on each workday or only in the morning on Saturday and Sunday. All pups were sacrificed with CO2 on day 4 post partum and examined macroscopically for external and visceral findings.
Near the end of the administration period a functional observational battery was performed and motor activity was measured in 5 randomly selected parental males and females per group, respectively. Blood was sampled from 5 randomly selected parental males and 5 parental females per group for hematological and clinical chemistry examination. All surviving F0 parental animals were sacrificed by decapitation, under CO2 anesthesia, and were assessed by gross pathology. Organ weights were recorded and a histopathological examination was performed.
The following test substance-related, adverse effects/findings were noted:
- 1000 mg/kg body weight/day:
F0 parental animals: statistically significantly reduced hemoglobin and hematocrit values (i.e. indications of a mild anemic process) in the F0 males
F1 pups: no adverse effects/findings
- 100 and 300 mg/kg body weight/day:
F0 parental animals and F1 pups: no adverse effects/findings
Thus, under the conditions of this reproduction/developmental toxicity screening test the NOAEL (no observed adverse effect level) for reproductive performance and fertility is 1000 mg/kg body weight/day
(673 mg/kg bw/day Isopropanolamine) for the F0 parental rats. The NOAEL for general, systemic toxicity of the test substance is 300 mg/kg body weight/day (202 mg/kg bw/day Isopropanolamine) for the F0 parental males based on some indications for a mild anemic process.
The NOAEL for parental females (F0) was found to be 1000 mg/kg body weight/day (673 mg/kg bw/day Isopropanolamine).
The NOAEL for developmental toxicity was 1000 mg/kg body weight/day (673 mg/kg bw/day Isopropanolamine).
- Endpoint:
- developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- Apr - Sep 2022
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 414 (Prenatal Developmental Toxicity Study)
- Version / remarks:
- June 2018
- GLP compliance:
- yes (incl. QA statement)
- Specific details on test material used for the study:
- No specific handling conditions required. The required amount of test item was weighed and dissolved in Elix water (w/w). The pH of the formulation was lowered to pH 7.9-8.1 using HCl. Formulations were prepared at least weekly, filled out in daily portions and stored in the refrigerator. Dosing formulations were removed from the refrigerator at least 30 minutes before dosing and were stirred at room temperature.
- Species:
- rat
- Strain:
- Wistar
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Females (if applicable): nulliparous, arrived on Day 0 or 1 post-coitum
- Age at study initiation: between 10-15 weeks old
- Weight at study initiation: 178 - 281 g females
- Fasting period before study: -
- Housing: Polycarbonate cages (Makrolon type MIII, height 18 cm) containing sterilized wooden fibers as bedding material (Lignocel S 8 15, JRS J.Rettenmaier & Söhne GmbH + CO. KG, Rosenberg, Germany) and enrichment and nesting material (paper and aspen wooden sticks), equipped with water bottles. Animals will be individually housed.
- Diet (e.g. ad libitum): pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest,
Germany) ad libitum
- Water (e.g. ad libitum): yes. Municipal tap water
- Acclimation period: at least 5 days prior to the commencement of dosing
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 43-54% (target: 40-70%)
- Light Cycle: 12-hours light and 12-hours dark
- Ventilation: At least 10 air changes per hour.
IN-LIFE DATES: Initiation of Dosing: 25 Apr 2022, completion of In-life: 13 May 2022 - Route of administration:
- oral: gavage
- Vehicle:
- water
- Details on exposure:
- The dose volume for each animal were based on the most recent body weight measurement. The doses were given using a plastic feeding tube. The dose formulations were stirred continuously during dosing. Dose pot identification via Provantis was used as additional check to verify the dosing procedure according to Standard Operating Procedures.
PREPARATION OF DOSING SOLUTIONS:
- Formulations (w/w) homogenized to visually acceptable levels and pH set to 7.9-8.1 using 37% HCl (Merck, Darmstadt, Germany)
- Frequency of Preparation: At least weekly, filled out in daily portions and stored in the refrigerator.
- Storage Conditions Set to Maintain: 4°C
The dosing formulations will be removed from the refrigerator and stirred at room temperature for at least 30 minutes before dosing and dosed within 24 hours after removal from the refrigerator. Dose formulation samples will be collected for analysis. - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Concentration and Homogeneity Analysis and Stability Analysis were performed using a validated analytical procedure (Test Facility Study No. 20316908).
- Details on mating procedure:
- time-mated.
Untreated females were mated at the Supplier and were at Day 0 or 1 post-coitum on arrival at the Test Facility (Day 0 post-coitum is the day of successful mating). - Duration of treatment / exposure:
- Day 6 to Day 20 post-coitum.
- Frequency of treatment:
- Once daily
- Dose / conc.:
- 0 mg/kg bw/day (nominal)
- Dose / conc.:
- 100 mg/kg bw/day (nominal)
- Dose / conc.:
- 300 mg/kg bw/day (nominal)
- Dose / conc.:
- 1 000 mg/kg bw/day (nominal)
- No. of animals per sex per dose:
- 22
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- - Dose selection rationale:
The dose levels were selected based on the results of the Dose Range Finder (Test Facility Study No. 20316909) and in an attempt to produce graded responses to the test material. Recorded test material-related changes in the DRF were considered not dose-limiting for the subsequent Main study. Therefore, selected dose levels for the Main study were 100, 300 and 1000 mg/kg bw/d. - Maternal examinations:
- CAGE SIDE OBSERVATIONS: Yes
- At least once daily (immediately after dosing) starting on Day 6 post coitum up to and including the day prior to necropsy.
DETAILED CLINICAL OBSERVATIONS: Yes
- On Days 2, 6, 15 and 21 post coitum
BODY WEIGHT: Yes
- On Days 2, 6, 9, 12, 15, 18 and 21 post-coitum.
FOOD CONSUMPTION: Yes
- Over Days 2-6, 6-9, 9-12, 12-15, 15-18 and 18-21 post coitum.
WATER CONSUMPTION: Yes
- Regular basis throughout the study.
POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 21 post-coitum
- Necropsy (external examination, e.g. of ovaries and uterine horns, thyroid glands), Collection of Gross Lesions, Organ Weights (uteri, thyroid gland), Histology and Histopathology (thyroid gland) - Ovaries and uterine content:
- yes
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: number and distribution of live and dead fetuses; sex of each fetus based on the anogenital distance, if possible. - Blood sampling:
- At day 21 post coitum, no fasting, for thyroid hormone measurement: Triiodothyronine (T3), Thyroxine (T4), Thyroid-Stimulating Hormone (TSH).
- Fetal examinations:
- External Examination, Body Weight, External Sex Determination, AGD Measurement
External, visceral, and skeletal findings were recorded as developmental variations (alterations in anatomic structure that are considered to have no significant biological effect on animal health or body conformity and/or represent slight deviations from normal) or malformations (those structural anomalies that alter general body conformity, disrupt or interfere with normal body function, or may be incompatible with life).
- External examinations: Yes: [all per litter], Each viable fetus was sexed, examined in detail to detect macroscopic visible abnormalities and their weight was determined. The anogenital distance (AGD) was measured for all viable fetuses. The AGD was normalized to the cube root of the fetal body weight.
- Soft tissue examinations: Yes: [half per litter] The sex of all fetuses was confirmed by internal examination and approximately one-half of the fetuses (live and dead) in each litter (all groups) were examined for visceral anomalies by dissection in the fresh (non-fixed) state. The thoracic and abdominal cavities were opened and dissected, including the heart and major vessels. Fetal kidneys were examined and graded for renal papillae development. The heads were removed from this one-half of the fetuses in each litter and placed in Bouin's solution for soft-tissue examination. After examination, the tissues without variations or malformations were discarded. Tissues with variations or malformations were stored in 10% formalin. Any remaining tissues (from the fetuses used for fresh visceral examination) were discarded. The carcasses were processed and stained (see described below) but were not examined in first instance.
- Skeletal examinations: Yes: [half per litter] All fetuses were eviscerated, followed by fixation in 96% aqueous ethanol, and maceration in potassium hydroxide. Thereafter, they were stained with Alizarin Red S. Subsequently, skeletal examination was done for one-half of the fetuses (i.e. the fetuses with heads). All specimens were archived in glycerin with bronopol as preservative. A few bones were not available for skeletal examination because they were accidentally damaged or lost during processing. The missing bones were listed in the raw data; evaluation by the fetal pathologist and Study Director determined there was no influence on the outcome of the individual or overall skeletal examinations, or on the integrity of the study as a whole.
- Head examinations: Yes: [half per litter], see soft tissue examination
- Anogenital distance of all live rodent pups: yes, normalized to the cube root of the fetal BW. - Statistics:
- Any data collected during the (Pre)treatment Period are tabulated, summarized or statistically analyzed. All statistical analyses were performed within the respective study phase, unless otherwise noted. Numerical data collected on scheduled occasions were summarized and statistically analyzed as indicated below according to sex and occasion or by litter.
Means, standard deviations (or % coefficient of variation or standard error, when deemed appropriate), ratio, percentages, numbers, and/or incidences were reported as appropriate by dataset. - Indices:
- Pregnancy Rate (%): No. of pregnant females x 100 / No. of mated females
Male Fetuses (%): No. of male fetuses x 100 / No. of fetuses
Female Fetuses (%): No. of female fetuses x 100 / No. of fetuses
Pre-Implantation Loss (%): No. of corpora lutea – No. of implantations x 100 / No. of corpora lutea
Post-Implantation Loss (%): No. of implantations – No. of live fetuses x 100 / No. of implantations
Litter % of Fetuses with Abnormalities: No. of fetuses in litter with a given finding x 100 / No. of fetuses in litter examined - Historical control data:
- Historical data on thyroid hormone analyses and fetal pathology are available (see attached document below)
- Clinical signs:
- no effects observed
- Mortality:
- no mortality observed
- Body weight and weight changes:
- no effects observed
- Food consumption and compound intake (if feeding study):
- no effects observed
- Water consumption and compound intake (if drinking water study):
- not examined
- Ophthalmological findings:
- not examined
- Haematological findings:
- not specified
- Description (incidence and severity):
- Serum levels of TSH were comparable to control values. Lower mean serum levels of total T4 at all dose levels were not statistically significant at 300 mg/kg bw/d and thus not dose-dependent. A statistically significant lower mean serum level of T3 was recorded at 1000 mg/kg bw/d. All mean values of T3 and T4 remained within the historical control data (see attachments below). Thus, this was considered as non-adverse.
- Clinical biochemistry findings:
- not examined
- Endocrine findings:
- not specified
- Description (incidence and severity):
- Serum levels of TSH were not statistically significantly different compared to control values. Lower mean serum levels of total T4 were noted at all dose levels (0.87, 0.89 and 0.85x of control at 100, 300 and 1000 mg/kg bw/d, respectively, not statistically significant at 300 mg/kg bw/d) and thus not dose-dependent. A statistically significant lower mean serum level of T3 (0.78x of control) was recorded at 1000 mg/kg bw/d. All mean values of T3 and T4 remained within the historical control data. As there were no weight changes and microscopic observations in the thyroid glands, this was considered as non-adverse.
(see attachments below) - Urinalysis findings:
- not examined
- Behaviour (functional findings):
- not examined
- Immunological findings:
- not examined
- Organ weight findings including organ / body weight ratios:
- no effects observed
- Description (incidence and severity):
- There were no test material-related alterations in thyroid gland weights.
- Gross pathological findings:
- no effects observed
- Neuropathological findings:
- not examined
- Histopathological findings: non-neoplastic:
- no effects observed
- Description (incidence and severity):
- There were no test material-related microscopic observations in the thyroid glands.
- Histopathological findings: neoplastic:
- not examined
- Other effects:
- no effects observed
- Details on results:
- (see attached tables below)
- Number of abortions:
- no effects observed
- Pre- and post-implantation loss:
- no effects observed
- Total litter losses by resorption:
- no effects observed
- Early or late resorptions:
- no effects observed
- Dead fetuses:
- no effects observed
- Changes in pregnancy duration:
- no effects observed
- Changes in number of pregnant:
- no effects observed
- Description (incidence and severity):
- A total of five females were non-gravid: Nos. 15 and 17 (control), No. 35 (100 mg/kg bw/d), No. 45 (300 mg/kg bw/d) and No. 82 (1000 mg/kg bw/d). Therefore, the number of females with viable litters available for evaluation was 20, 21, 21 and 21 in the control, 100, 300 and 1000 mg/kg bw/d groups, respectively.
- Other effects:
- no effects observed
- Details on maternal toxic effects:
- (see attached tables below)
- Dose descriptor:
- NOAEL
- Effect level:
- 1 000 mg/kg bw/day
- Based on:
- test mat.
- Remarks on result:
- not determinable due to absence of adverse toxic effects
- Abnormalities:
- no effects observed
- Fetal body weight changes:
- no effects observed
- Reduction in number of live offspring:
- no effects observed
- Changes in sex ratio:
- no effects observed
- Changes in litter size and weights:
- no effects observed
- Anogenital distance of all rodent fetuses:
- no effects observed
- Changes in postnatal survival:
- not examined
- External malformations:
- no effects observed
- Skeletal malformations:
- no effects observed
- Visceral malformations:
- no effects observed
- Other effects:
- no effects observed
- Details on embryotoxic / teratogenic effects:
- (see attached tables below)
- Dose descriptor:
- NOAEL
- Effect level:
- 1 000 mg/kg bw/day
- Based on:
- test mat.
- Sex:
- male/female
- Remarks on result:
- not determinable due to absence of adverse toxic effects
- Abnormalities:
- no effects observed
- Developmental effects observed:
- no
- Conclusions:
- No maternal and developmental toxicity was observed up to the highest dose level tested.
- Executive summary:
In a prenatal developmental toxicity study according to OECD test guideline 414, time-mated female Wistar Han rats were treated with 1-aminopropan-2-ol from Days 6 to 20 post coitum, inclusive, by daily oral gavage at dose levels of 100, 300 and 1000 mg/kg bw/d. The rats of the control group received the vehicle, Water (Elix), alone. No maternal and developmental toxicity was observed up to the highest dose level tested. Lower serum T4 levels were measured for all treatment groups, without statistical significance in the mid dose and additionally a lower serum T3 level was recorded at 1000 mg/kg bw/d. All group means remained within the available historical control data range. Further, no test material related changes in either serum TSH levels, thyroid gland weight or morphology were observed. Thus, the observed changes in serum T3 and T4 were considered not to represent an adverse effect of the test material.
In conclusion, the following maternal and developmental No Observed Adverse Effect Levels (NOAELs) for 1-aminopropan-2-ol were established: Maternal NOAEL: at least 1000 mg/kg bw/day. Developmental NOAEL: at least 1000 mg/kg bw/day.
- Endpoint:
- developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: Acceptable, well-documented publication which meets basic scientific prnciples.
- Justification for type of information:
- ADAPTATION ACCORDING TO REACH ANNEX XI, section 1 - see justifications attached to IUCLID section 13.2.
- Justification WoE_RDT_Reprotox_Oct2022
- Justification RA_CAS 122-20-2_Dev.tox.2ndspecies_Nov2021 - Qualifier:
- equivalent or similar to guideline
- Guideline:
- OECD Guideline 414 (Prenatal Developmental Toxicity Study)
- Principles of method if other than guideline:
- Groups of 25 time-mated female Sprague–Dawley rats were given 0, 100, 300, or 1000 mg DIPA/kg/day on gestation days 6 through 20 by oral gavage.
- GLP compliance:
- yes
- Limit test:
- no
- Species:
- rat
- Strain:
- other: CRL:CD(SD)
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River Laboratories Inc. (Kingston, New York and Raleigh, North Carolina)
- Diet: LabDiet #5002 Certified Rodent Diet (PMI Nutrition International, St. Louis, MO) ad libitum
- Water: tap water ad libitum
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-25 °C
- Humidity (%): 45-60 %
- Air changes (per hr): 12-15
- Photoperiod (hrs dark / hrs light): 12/12 - Route of administration:
- oral: gavage
- Vehicle:
- water
- Analytical verification of doses or concentrations:
- yes
- Details on mating procedure:
- - Impregnation procedure: purchased timed pregnant
- Duration of treatment / exposure:
- Gestation days 6-20
- Frequency of treatment:
- daily
- Duration of test:
- 21 days
- Remarks:
- Doses / Concentrations:
0, 100, 300 or 1000 mg/kg bw/day
Basis:
nominal conc. - No. of animals per sex per dose:
- 25 (females only)
- Control animals:
- yes, concurrent vehicle
- Maternal examinations:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
BODY WEIGHT: Yes
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 21
- Organs examined: The weights of the kidneys, liver and gravid uteri were obtained and the kidneys, liver and all gross lesions were preserved in 10% NBF. - Ovaries and uterine content:
- The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of resorptions: Yes - Fetal examinations:
- - External examinations: Yes: all per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: Yes: half per litter - Statistics:
- Means and standard deviations were calculated for all continuous data. These parameters were first examined for equality of variance using Bartlett’s test (a = 0.01; Winer, 1971). If the results of the Bartlett’s test were significant, the data were transformed in an attempt to obtain equality of variances. The order of transformations used was the common log, the inverse and the square root with the best fit used for subsequent statistical testing. Maternal body weight, weight gain, feed consumption, organ weights and fetal body weights were analyzed using parametric or non-parametric ANOVA followed by Dunnett's or Wilcoxon's test for comparison to controls; pre- and post-implantation loss were evaluated using a Censored Wilcoxon's test; Corpora lutea, implantations and litter size were analyzed using non-parametric ANOVA follewed by Wilcoxon's test; pregnancy rates were evaluated using Fisher exact probability test; and fetal sex ratios were analyzed using a binomial distribution test. As statistical interactions (i.e., time by dose or sex by dose) were identified in several of the tests, the alpha levels for interaction terms were set a priori at 0.01–0.10, with Bonferroni’s correction. The alpha level for comparison of individual dose groups to controls was set a priori at 0.05.
- Details on maternal toxic effects:
- Maternal toxic effects:no effects
- Dose descriptor:
- NOAEL
- Effect level:
- 1 000 mg/kg bw/day (nominal)
- Basis for effect level:
- other: maternal toxicity
- Details on embryotoxic / teratogenic effects:
- Embryotoxic / teratogenic effects:no effects
- Dose descriptor:
- NOAEL
- Effect level:
- 1 000 mg/kg bw/day (nominal)
- Basis for effect level:
- other: fetotoxicity
- Abnormalities:
- not specified
- Developmental effects observed:
- not specified
- Endpoint:
- developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Study period:
- 27 Jan 1994 - 21 Feb 1994
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- ADAPTATION ACCORDING TO REACH ANNEX XI, section 1 - see justifications attached to IUCLID section 13.2.
- Justification WoE_RDT_Reprotox_Oct2022
- Justification RA_CAS 122-20-2_Dev.tox.2ndspecies_Nov2021 - Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 414 (Prenatal Developmental Toxicity Study)
- GLP compliance:
- yes
- Limit test:
- no
- Specific details on test material used for the study:
- - Name of test material (as cited in study report): Triisopropanolamine
- Analytical purity: 92 %
- Lot/batch No.: 10-4852
- Storage condition of test material: Room temperature, under nitrogen atmosphere - Species:
- rat
- Strain:
- Wistar
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: THOMAE, Biberach an der Riss, FRG
- Age at study initiation: 77-89 days
- Weight at study initiation: 242 g (mean)
- Housing: singly in type DK III stainless steel wire mesh cage.
- Diet: ground Kliba 343 feed, Klingenthalmuehle AG, Kaiseraugst, Switzerland, ad libitum
- Water: tap water, ad libitum
- Acclimation period: at least 5 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12/12
- Route of administration:
- oral: gavage
- Vehicle:
- water
- Remarks:
- double distilled
- Details on exposure:
- Doses of 0, 100, 400 and 1000 mg/kg bw/day were administered in a volume of 10 mL/kg, at a concentration of 0, 1000, 4000 and 10000 mg/100mL, respectively.
- Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Test substance: purity and stability were analyzed by gas chromatography (GC); and homogeneity was proven visually.
Test solutions: analysis of stability (for 3 hrs) in double distilled water was carried out in a range-finding study. Concentrations were analyzed twice during the study by GC. Test solutions were freshly prepared on the day of dosing. - Details on mating procedure:
- - Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1/1
- Length of cohabitation: overnight
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy - Duration of treatment / exposure:
- on day 6 through day 15 post coitum (p.c.)
(Sacrifice on day 20 p.c.) - Frequency of treatment:
- once a day during the period of major organogenesis (day 6 to day 15 p.c.)
- Duration of test:
- 20 days
- Dose / conc.:
- 100 mg/kg bw/day (nominal)
- Dose / conc.:
- 400 mg/kg bw/day (nominal)
- Dose / conc.:
- 1 000 mg/kg bw/day (nominal)
- No. of animals per sex per dose:
- 25
- Control animals:
- yes, concurrent vehicle
- Maternal examinations:
- CAGE SIDE OBSERVATIONS
- Time schedule: once a day and more often when clinical signs of toxicity were elicited. Mortality was check twice a day on workdays and once a day during weekends and public holidays.
BODY WEIGHT and FOOD CONSUMPTION
- Time schedule for examinations: days 0 (only bw), 1, 3, 6, 8, 10, 13, 15, 17, 20 p.c.
POST-MORTEM EXAMINATIONS
- Sacrifice on gestation day # 20
- Organs examined: uterus and ovaries after gross pathology
OTHER: The correct body weight gain was calculated after terminal sacrifice. - Ovaries and uterine content:
- The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: furthermore, calculations of conception rate and pre- and postimplantation losses were carried out - Fetal examinations:
- - External examinations: Yes: all per litter: fetus was weighed, sexed, examined macroscopically for external findings, and viability, condition of the placentae, umbilical cords, fetal membranes and fluids were examined.
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: No - Statistics:
- The data were evaluated statistically using the computer systems of the Department of Toxicology of BASF AG. The Dunnett-test was used for a simultaneous comparison of several dose groups with the control. The hypothesis of equal means was tested. This test was performed two-sided and was used for the statistical evaluation of the following parameters: food consumption, body weight, body weight change, corrected body weight gain (net maternal body weight change), weight of the uterus before it was opened, number of corpora lutea, number of implantations, number of resorptions and number of live fetuses, proportion of preimplantation loss, postimplantation loss, resorptions and live fetuses in each litter, litter mean fetal body weight and litter mean placental weight. Fisher's Exact Test was used for a pairwise comparison of each dose group with the control for the hypothesis of equal proportions. This test was performed one-sided and was used for female mortality, females pregnant at terminal sacrifice and the number of litters with fetal findings. The Wilcoxon-Test was used for a comparison of each dose group with the control for the hypothesis of equal medians. This test was performed one-sided and was used for the proportion of fetuses with malformations, variations, retardations and/or unclassified observations in each litter. If the results of these tests were significant, labels (* for p< 0.05, ** for p< 0.01) were printed in the summary tables.
- Clinical signs:
- no effects observed
- Dermal irritation (if dermal study):
- not examined
- Mortality:
- no mortality observed
- Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- There were no statistically significant differences between the controls and the substance-treated dams concerning mean body weights. At the beginning of the treatment period (days 6-8 p.c.), however, the dams of the highest dose group (1,000 mg/kg bw/day) showed a statistically significantly reduced body weight gain (only about 36% of the weight gain of the concurrent control group; see table in remarks on results). The corrected body weight gain (terminal body weight on day 20 p.c. minus weight of the uterus before it was opened minus body weight on day 6 p.c.) was statistically significantly lower in the high dose group (about 87% of the value of the concurrent control group), which is related to the test substance administration (see table in remarks on results).
- Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- In the 1000 mg/kg group statistically significantly decreased food consumption at the beginning of the treatment period (days 6-10 p.c.; about 13% (days 6 to 8 p.c.) or about 16% (days 8 to 10 p.c.) lower than the values of the concurrent control group.
- 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:
- no effects observed
- Neuropathological findings:
- not examined
- Histopathological findings: non-neoplastic:
- no effects observed
- Histopathological findings: neoplastic:
- not examined
- Other effects:
- not examined
- Number of abortions:
- no effects observed
- Pre- and post-implantation loss:
- no effects observed
- Total litter losses by resorption:
- no effects observed
- Early or late resorptions:
- no effects observed
- Dead fetuses:
- no effects observed
- Changes in pregnancy duration:
- not examined
- Description (incidence and severity):
- Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed - Changes in number of pregnant:
- no effects observed
- Other effects:
- not examined
- Details on maternal toxic effects:
- There were no substance-related and/or biologically relevant differences between the groups in conception rate, in the mean number of corpora lutea and implantation sites or in the values calculated for the pre- and the postimplantation losses, the number of resorptions and viable fetuses. The differences evident are considered to be incidental and within the normal range of deviations for animals of this strain and age.
- Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 400 mg/kg bw/day
- Based on:
- test mat.
- Basis for effect level:
- body weight and weight gain
- food consumption and compound intake
- Key result
- Abnormalities:
- no effects observed
- Fetal body weight changes:
- no effects observed
- Description (incidence and severity):
- Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed - Reduction in number of live offspring:
- no effects observed
- Changes in sex ratio:
- no effects observed
- Changes in litter size and weights:
- no effects observed
- Changes in postnatal survival:
- no effects observed
- External malformations:
- no effects observed
- Skeletal malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- The few statistically significant differences on embryo-/fetotoxicity, which occurred were exclusively related to fetal skeletal variations and retardations. These findings consisted of: an increased rate of affected fetuses/litter and an increased litter incidence with shortened 13th rib(s) at 100, 400 and 1,000 mg/kg bw/day, respectively; a consequently increased rate of low and intermediate dose fetuses/litter with total skeletal variations; an increased litter incidence of incompletely ossified or smaller sternebra(e) in the 400 mg/kg body weight group. These findings are considered to be spontaneous in nature because no dose-response relationship is given and/or the respective values are fully in the historical control range.
- Visceral malformations:
- no effects observed
- Other effects:
- not examined
- Key result
- Remarks on result:
- not determinable due to absence of adverse toxic effects
- Key result
- Abnormalities:
- no effects observed
- Key result
- Developmental effects observed:
- no
- Endpoint:
- developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- weight of evidence
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- ADAPTATION ACCORDING TO REACH ANNEX XI, section 1 - see justifications attached to IUCLID section 13.2.
- Justification WoE_RDT_Reprotox_Feb2019
- Justification RA_CAS 122-20-2_Dev.tox.2ndspecies_Nov2018 - Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 414 (Prenatal Developmental Toxicity Study)
- Version / remarks:
- 1981
- Qualifier:
- according to guideline
- Guideline:
- EPA OPP 83-3 (Prenatal Developmental Toxicity Study)
- GLP compliance:
- yes
- Limit test:
- no
- Specific details on test material used for the study:
- - Test material name: 4-Amino-3.5,6-trichloropicolinate, Triisopropanolamine Salt
- Species:
- rabbit
- Strain:
- New Zealand White
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Hazleton Research Products, Inc. (Kalamazoo, MI)
- Age at study initiation: 6 - 8 weeks
- Weight at study initiation: 3250 – 4250 g
- Housing: individually in cages with wire floors
- Diet: 8 oz/day, Certified Laboratory Rabbit Chow No. 5322 (Purina Mills, Inc., St. Louis, MO)
- Water: ad libitum, municipal tap water
- Acclimation period: at least 2 weeks - Route of administration:
- oral: gavage
- Vehicle:
- water
- Remarks:
- deionized
- Details on exposure:
- VEHICLE
- Amount of vehicle (if gavage): 2 mL/kg bw - Analytical verification of doses or concentrations:
- yes
- Details on mating procedure:
- - Impregnation procedure: artificial insemination
- Duration of treatment / exposure:
- Days 7 - 19 of gestation
- Frequency of treatment:
- daily
- Duration of test:
- Until day 28 of gestation
- Dose / conc.:
- 54 mg/kg bw/day (nominal)
- Remarks:
- Phase II
- Dose / conc.:
- 180 mg/kg bw/day (nominal)
- Remarks:
- Phase I & II
- Dose / conc.:
- 538 mg/kg bw/day (nominal)
- Remarks:
- Phase I & II
- Dose / conc.:
- 1 000 mg/kg bw/day (nominal)
- Remarks:
- Phase I & II
- No. of animals per sex per dose:
- 18
- Control animals:
- yes, concurrent vehicle
- Maternal examinations:
- CAGE SIDE OBSERVATIONS:
Animals were observed daily. Any animal which showed indications of early termination of pregnancy was submitted for a complete necropsy.
BODY WEIGHT:
Body weights were recorded on Days 0, daily during the dosing period, and on Days 20 and 28 of gestation
FOOD CONSUMPTION: only in Phase II study
Feed consumption data was collected daily throughout the test period to better interpret the results obtained from body weight, body weight gain and fecal output data.
POST-MORTEM EXAMINATIONS:
- Sacrifice on gestation day # 28
- Examinations: a limited gross pathologic examination (necropsy) was performed on all animals and liver with gallbladder, kidneys and gravid uterine weights were recorded. Any obvious structural or gross pathologic alterations noted in the adult were recorded. Sections of liver with gallbladder, kidneys and gross lesions were preserved in neutral, phosphate-buffered 10% formalin. Since there were no gross pathologic observations considered. treatment-related. in the dams at any dose level, tissues were not examined histopathologically. - Ovaries and uterine content:
- The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: position of fetuses in utero and position of resorption sites - Fetal examinations:
- - External examinations: Yes: The sex and body weight of each fetus was determined. Any gross external alterations were recorded.
- Soft tissue examinations: Yes: All fetuses were examined by dissection under a low power stereomicroscope for evidence of visceral alterations. This examination also included a fresh examination of the brain.
- Skeletal examinations: Yes: All fetuses were then preserved in alcohol, eviscerated, cleared and stained with alizarin red-S and examined for skeletal alterations.
- Head examinations: Yes - Statistics:
- Descriptive statistics (means and standard deviations) were calculated for feed consumption. Maternal body weights, body weight gains, organ weights (absolute and relative) and fetal body weights were evaluated by Bartlett's test for equality of variances. Based on the outcome of Bartlett's test, a parametric or nonparametric analysis of variance (ANOVA) was performed. If the ANOVA was significant, analysis by Dunnett's test or the Wilcoxon Rank-Sum test with Bonferroni's correction was performed, respectively. Statistical evaluation of the frequency of pre-implantation loss, resorptions and fetal alterations among litters and the fetal population was performed using a censored Wilcoxon test with Bonferroni's correction. The number of corpora lutea and implants, and litter size were evaluated using a nonparametric ANOVA followed by the Wilcoxon Rank-Sum test with Bonferroni's correction. Pregnancy rates were analyzed using the Fisher exact probability test. Fetal sex ratios were analyzed using a binomial distribution test. Statistical outliers were identified using a sequential method, but were not excluded unless justified by sound scientific reasons unrelated to treatment.
- Clinical signs:
- effects observed, treatment-related
- Description (incidence and severity):
- - Phase I: The only treatment-related in-life alteration observed was an increase in the incidence of decreased feces in rabbits given 538 and 1000 mg/kg bw/day. This alteration, though transient, lasting 1-4 days, was considered to be a treatment-related response since it was accompanied by a corresponding decrease in body weight gain.
- Phase II: Dose-related increases in the incidence of decreased feces were observed in treated rabbits during the treatment period. Soft feces was also common among the treated rabbits. Perineal soiling was seen in 7 out of 18 high-dose rabbits. - Dermal irritation (if dermal study):
- not examined
- Mortality:
- no mortality observed
- Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- - Phase I: There were no significant treatment-related effects on body weights. Dose-related statistically significant decreases in body weight gain were observed in dams given 180, 538 and 1000 mg/kg bw/day during Days 7 through 10 of gestation. The body weight gain of dams given 1000 mg/kg bw/day was also statistically decreased during Days 7-20. A compensatory increase in body weight gain following the exposure period (Days 20 through 28) was observed in dams given 180, 538 and 1000 mg/kg bw/day. These compensatory increases in body weight gain were statistically significant in dams given 538 and 1000 mg/kg bw/day and resulted in equal weight gains for all groups of rabbits over the test period (Days 0 through 28).
- Phase II: Body weights of the high-dose dams were decreased throughout most of the dosing period, with statistically significant decreases occurring on Days 10, 16 and 20. The decreased weights were consistent with and reflective of the decreased feed consumption noted in these animals during this time period. No significant effect on body weights was observed in dams given 54, 180 or 538 mg/kg bw/day. Dose-related statistically significant decreases in body weight gains were observed in dams given 180,538 and 1000 mg/kg bw/day at the start of the dosing period (Days 7-10). The decrease in body weight gain in the 180 mg/kg/day dose group dams was not considered to be of biological significance since body weight gain after Day 10 of gestation increased and was higher than the controls for the rest of the test period (Days 10-28). The 538 and 1000 mg/kg bw/day dose group dams had, in general, lower body weight gain during the later part of the dosing period, resulting in a statistically decreased body weight gain during Days 7-20. A statistically significant compensatory increase in body weight gain was seen in the 538 and 1000 mg/kg/ day dose group rabbits following the exposure period (Days 20-28). The changes in body weight gain paralleled the trends in feed consumption. - Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- - Phase I: Although feed consumption was not recorded in Phase I of the study, the decreased body weight gain in all treated rabbits during Days 7 through 10 and decreased fecal output at the 538 and 1000 mg/kg bw/day dose levels indicate a reduction in feed consumption at the start of the dosing period in these rabbits.
- Phase II: There was a dose-related decrease in feed consumption in dams given 538 and 1000 mg/kg bw/day during the dosing period. Feed consumption was also slightly decreased during Days 7-10 in dams given 180 mg/kg bw/day. No significant effect on feed consumption was observed in dams given 54 mg/kg/day. Feed consumption of all treated groups of rabbits was comparable to the control during the post-dosing period. - 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:
- no effects observed
- Description (incidence and severity):
- - Phase I & II: There were no significant treatment-related effects on absolute or relative liver or kidney weights at any dose level.
- Gross pathological findings:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - Phase I: Gross pathologic examination revealed no remarkable alterations other than a pale liver, decreased ingesta and blood in the urine.
- Neuropathological findings:
- not examined
- Histopathological findings: non-neoplastic:
- not examined
- Histopathological findings: neoplastic:
- not examined
- Other effects:
- not examined
- Number of abortions:
- effects observed, treatment-related
- Description (incidence and severity):
- - Phase II: Two rabbits from the 1000 mg/kg bw/day dose group aborted their fetuses and were sent for a complete necropsy.
- Pre- and post-implantation loss:
- no effects observed
- Total litter losses by resorption:
- no effects observed
- Early or late resorptions:
- effects observed, treatment-related
- Description (incidence and severity):
- - Phase I: The resorption rates of dams given 180, 538 or 1000 mg/kg/day were increased relative to the control, with statistically significant increases in dams given 180 or 1000 mg/kg/day. The apparent increases in resorption rates of dams given the test substance were not considered to be treatment-related since, 1) the resorption rates for all dose groups were within the range of historical controls and 2) the resorption rate of the concurrent controls was unusually low and fell below the range of historical control.
- Phase II: No effects attributed to treatment were observed in the reproductive parameters made at the time of necropsy. - Dead fetuses:
- no effects observed
- Changes in pregnancy duration:
- effects observed, treatment-related
- Description (incidence and severity):
- - Phase I: One rabbit from the 1000 mg/kg bw/day dose group showed signs of early termination of pregnancy and was sent for a complete necropsy on gestation Day 22. This rabbit incurred a severe weight loss (673 grams) during the treatment period (Days 7 through 20) which may have contributed to the early delivery of the fetuses.
- Phase II: One rabbit from the 1000 mg/kg bw/day dose group started delivering just prior to necropsy. - Changes in number of pregnant:
- not examined
- Other effects:
- not examined
- Dose descriptor:
- NOAEL
- Effect level:
- 180 mg/kg bw/day
- Based on:
- test mat.
- Basis for effect level:
- body weight and weight gain
- clinical signs
- number of abortions
- Abnormalities:
- no effects observed
- Fetal body weight changes:
- no effects observed
- Reduction in number of live offspring:
- no effects observed
- Changes in sex ratio:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - Phase I: The distribution of male and female fetuses within the 180 mg/kg/day dose group was statistically different from that of a binomial distribution. This deviation from a binomial distribution was considered to be a random event in view of the lack of an effect on the sex ratio in fetuses from dams given 538 or 1000 mg/kg/day.
-Phase II: The distribution of male and female fetuses in the 538 mg/kg/day dose group was significantly different from that of a binomial distribution. This deviation was not seen at the high dose and hence, was considered to be a random event and not due to treatment. - Changes in litter size and weights:
- not examined
- Changes in postnatal survival:
- not examined
- External malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - Phase II: In the control group a cleft palate was observed one fetus.
- Skeletal malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - Phase I: One fetus in the high dose group had a hemivertebrae.
- Phase II: In the 54 mg/kg bw/day dose group one fetus had a thoracic hemivertebra and an extra lumbar rib.
In the 180 mg/kg bw/day dose group one fetus had a thoracic hemivertebra.
In the high dose group a cervical and a thoracic hemivertebrae and a fused rib was seen one fetus. Another fetus had fused caudal vertebra. - Visceral malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- - Phase I: There was a slightly higher incidence of retroesophageal right subclavian artery in rabbits given 1000 mg/kg bw/day, suggestive of a treatment-related effect. However, this malformation has been noted in low numbers in the historical control rabbits used in the test laboratory. There were no statistically or biologically significant treatment-related differences in the incidence of any fetal variation or malformation, or in the number of malformed fetuses in any dose group.
In the control dose group, 7 fetuses had single malformations. Three fetuses had single malformations consisting of an omphalocele, missing gall bladder or ectopic kidneys. Four fetuses had a missing caudal lobe of the lung.
The 180 mg/kg bw/day dose group had 12 fetuses with malformations. Of the 12 fetuses, 11 had incidences of a missing caudal lobe of the lung. The twelfth fetus had a missing caudal lobe of the lung and a retroesophageal right subclavian artery arising from the descending aorta.
In the 538 mg/kg/ day dose group, there were 6 fetuses that were malformed. Among the malformed fetuses, three had a missing caudal lobe of the lung. The remaining three fetuses had single malformations consisting of either an omphalocele, retroesophageal right subclavian artery or an enlarged right atrium.
The high dose group had 10 fetuses with malformations. Two fetuses had retroesophageal right subclavian arteries, one of which arose from the descending aorta. A third fetus had a missing caudal lung lobe, missing cartilaginous rings in the trachea and a persistent right fourth aortic arch. A fourth fetus had a missing caudal lobe of the lung. The remaining 6 fetuses had a missing caudal lobe of the lung.
- Phase II: A total of 3,5,3,9 and 9 fetuses were malformed in the 0, 54, 180, 538 and 1000 mg/kg bw/day dose groups, respectively. However, each malformation within a dose group occurred at a low frequency or was within the historical control range observed in the test laboratory. The following visceral malformations were observed:
In two fetuses of the control group a caudal lobe of the lung was missing.
The 54 mg/kg bw/day dose group had 4 fetuses that were malformed. They each had a single malformation consisting of missing caudal lobe of the lung, retroesophageal aortic arch, ectopic kidney or persistent truncus arteriosus with no connection to the right ventricle.
In the 180 mg/kg bw/day dose group, three malformed fetuses. One fetus had a missing caudal lobe of the lung. The remaining two fetuses had retroesophageal right subclavian arteries.
Of the 8 fetuses that were malformed in the 538 mg/kg bw/day dose group, five had a missing caudal lobe of the lung. A sixth fetus had an omphalocele. A retroesophageal right subclavian artery was seen in the seventh fetus. A eighth fetus had a diaphragmatic hernia and hypoplastic lungs.
The high dose group had nine fetuses that were malformed. A missing caudal lobe of the lung was seen in six fetuses. A seventh fetus had an omphalocele with secondary constriction of the liver, liver hemorrhage and a missing caudal lobe of the lung. - Other effects:
- no effects observed
- Details on embryotoxic / teratogenic effects:
- All fetal the malformations and variations within the treated groups were either not dose-related, occurred at low frequencies, or fell within the range of New Zealand White rabbit historical control data generated in this laboratory.
- Sex:
- male/female
- Remarks on result:
- not determinable due to absence of adverse toxic effects
- Abnormalities:
- effects observed, non-treatment-related
- Localisation:
- visceral/soft tissue: respiratory system
- Developmental effects observed:
- yes
- Lowest effective dose / conc.:
- 538 mg/kg bw/day
- Treatment related:
- yes
- Relation to maternal toxicity:
- developmental effects as a secondary non-specific consequence of maternal toxicity effects
- Dose response relationship:
- yes
- Relevant for humans:
- no
- Endpoint:
- developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- supporting study
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- test procedure in accordance with national standard methods with acceptable restrictions
- Justification for type of information:
- ADAPTATION ACCORDING TO REACH ANNEX XI, section 1 - see justifications attached to IUCLID section 13.2.
- Justification WoE_RDT_Reprotox_Feb2019
- Justification RA_CAS 122-20-2_Dev.tox.2ndspecies_Nov2018 - Principles of method if other than guideline:
- Female onn-pregnant rabbits were administered the test substance orally by gavage for 21 days, after blood sampling all animals will be sacrificed and examined.
- GLP compliance:
- yes
- Species:
- rabbit
- Strain:
- New Zealand White
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH / Charles River Laboratories, France
- Housing: single
- Diet (e.g. ad libitum): Pelleted Kliba maintenance diet rabbit and guinea pig “GLP”, Provimi Kliba SA, Kaiseraugst/Switzerland; ad libitum
- Water (e.g. ad libitum): drinking water, ad libitum
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-22
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12 - Route of administration:
- oral: gavage
- Vehicle:
- water
- Details on exposure:
- Dose: 1000 mg/kg bw
- Duration of treatment / exposure:
- 21 days
- Frequency of treatment:
- daily
- No. of animals per sex per dose:
- Control group: 3 animals
Test group (1000 mg/kg bw): 5 animals (due to the premature dead of 2 animals (1 found dead and 1 sacrificed after gavage error) from test group 1 (1000 mg/kg bw/d) 2 animals will be added) - Control animals:
- yes, concurrent no treatment
- Details on study design:
- The test substance was administered to the animals orally by gavage for 21 days. For this purpose, disposable syringes (e.g. B. Braun Melsungen AG, Melsungen, Germany) and suitable gavage tubes (e.g. Nelaton soft rubber tubes, Willy Ruesch GmbH, Kernen, Germany) were used. During the study, all animals were observed for any clinically abnormal signs. One day after the last administration blood samples were obtained from all surviving animals from the ear veins. After blood sampling all animals were sacrificed and examined.
- Maternal examinations:
- Mortality: A check for moribund and dead animals was made twice daily from Mondays to Fridays and once daily on Saturdays, Sundays and public holidays.
Clinical signs: A cageside examination was conducted at least once daily for any signs of morbidity, pertinent behavioral changes and/or signs of overt toxicity. During the administration period all animals were checked daily for any abnormal clinically signs before the administration as well as within 5 hours after the administration. Abnormalities and changes were documented for each animal.
Food consumption: Food consumption was recorded daily for days -4 - 0 and 0 - 21.
Body weight: Body weights was recorded on day 0, 2, 4, 6, 9, 11, 14, 16, 20 and 21.
Post-Mortem Examinations:The surviving animals were sacrificed by an intravenous injection of pentobarbital (dose: 2 mL/animal). After exsanguination the animals were necropsied and examined according to 3.12. Animals which died intercurrently or were sacrificed in a moribund state were necropsied in the same way as soon as possible after their death (with the exception of the organ weights).
Clinical Pathology: Blood samples were taken from the ear veins. The following parameters were examined: Hematology: Leukocytes, Erythrocytes, Hemoglobin, Hematocrit, mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), platelets, differential blood count, reticulocytes. Clinical chemistry: Alanine aminotransferase, Aspartate aminotransferase, Alkaline phosphatase,serum γ-glutamyl transferase, Inorg. phosphate, Calcium, Urea, Creatinine, Glucose, total bilirubin, total protein, Albumin, Globulins, Triglycerides, Cholesterol.
Pathology:
Necropsy: The sacrificed animals were necropsied and assessed by gross pathology. All animals which died intercurrently or were sacrificed in a moribund state were necropsied as soon as possible after their death and assessed by gross pathology.
Organ weights: The following weights were determined in all does sacrificed on schedule: Adrenal glands, kidneys, liver and spleen. The final body weights (ToxLims-System) will be transferred to the ACOPAT-System to calculate the relative organ weights.
Organ / tissue fixation: The following organs or tissues were fixed in 4% buffered formaldehyde solution: All gross lesions, adrenal glands, kidneys, liver and spleen. - Statistics:
- Dunnett-test (two-sided)
Kruskal-Wallis and Wilcoxon test - Details on results:
- The test substance was administered dissolved in drinking water for 21 days to groups of 3 – 5 female New Zealand White rabbits, via daily gavage. The dose levels were 0 and 1000 mg/kg body weight/day.
In the dose group (1000 mg/kg bw/d) all 5 females had reduced food consumption and lost body weight throughout the study. These animals also had reduced feces or diarrhea. Two rabbits died on the 3rd and 5th treatment day, respectively. The deceased animals showed no other pre-terminal clinical signs of toxicity, nor were there any specific necropsy findings.
Referenceopen allclose all
- 1000 mg/kg body weight/day, F1 pups: no adverse effects/findings
- 100 and 300 mg/kg body weight/day, F1 pups: no adverse effects/findings
Mean maternal body weight change during gestation (grams):
| 0 mg/kg bw/d(n = 25) | 100 mg/kg bw/d(n = 24) | 400 mg/kg bw/d(n = 25) | 1000 mg/kg bw/d(n = 23) | ||
| Days 0 - 1 | 4.3±3.76 | 3.7±3.60 | 4.3±3.41 | 4.3±2.89 | |
| Days 1 - 3 | 11.7±3.56 | 11.9±3.41 | 11.7±3.77 | 11.8±2.95 | |
| Days 3 - 6 | 13.0±4.56 | 12.1±4.40 | 13.1±4.37 | 12.5±3.39 | |
| Days 6 - 8 | 7.5±3.90 | 7.4±4.21 | 5.0±4.26 | 2.7±3.93** | |
| Days 8 - 10 | 9.9±4.59 | 9.4±5.15 | 11.4±4.43 | 8.0±3.72 | |
| Days 10 - 13 | 17.5±3.57 | 17.1±4.92 | 19.1±5.42 | 19.0±4.98 | |
| Days 13 - 15 | 11.7±4.51 | 11.9±4.15 | 12.5±4.36 | 13.7±4.62 | |
| Days 15 - 17 | 25.4±6.47 | 25.8±5.23 | 25.3±4.38 | 26.6±3.60 | |
| Days 17 - 20 | 50.3±10.57 | 49.2±6.64 | 51.8±7.58 | 52.6±5.56 |
**: p ≤ 0.01; Dunnett-test.
Mean gravid uterine Weights and net maternal body weight change (grams):
| 0 mg/kg bw/d(n = 25) | 100 mg/kg bw/d(n = 24) | 400 mg/kg bw/d(n = 25) | 1000 mg/kg bw/d(n = 23) | ||
| Gravid uterus | 78.3±19.22 | 80.3±10.49 | 82.5±13.18 | 84.4±10.16 | |
| Carcass | 317.7±22.09 | 310.8±20.66 | 314.3±22.22 | 306.2±13.17 | |
| Net weight change from day 6 | 44.1±8.54 | 40.4±7.52 | 42.6±7.06 | 38.2±6.72* |
*: p ≤ 0.05; Dunnett-test.
Under the conditions of this full-scale prenatal toxicity study, the administration of Triisopropanolamine to pregnant female Wistar rats during organogenesis elicited overt signs of maternal toxicity at 1000 mg/kg body weight/day, while 100 or 400 mg/kg body weight/day were tolerated by the dams without any substance-induced findings. The gestational parameters were not influenced by the test substance administration in any of the three test groups.
No signs of embryo-/fetotoxicity, especially no substance-induced indications of teratogenicity were noted up to and including the dose of 1,000 mg/kg body weight/day.
Therefore, the no observed adverse effect level (NOAEL) for the dams in this full-scale prenatal toxicity study is 400 mg/kg body weight/day, while it is 1000 mg/kg body weight/day for the fetal organism.
Given the unusually low resorption rate observed in the concurrent control rabbits and the historical control incidence of retroesophageal right subclavian artery, the apparent increased incidence of embryonal/fetal resorptions and retroesophageal right subclavian artery observed at the high dose were interpreted to be unrelated to treatment. However, the fact that these parameters were elevated above the concurrent controls raised some concern regarding the cause of the apparent increases. Therefore, in order to improve the confidence in the conclusion that the apparent increases in resorption rates and retroesophageal right subclavian arteries were unrelated to treatment, the study was repeated (Phase II).
Female non-pregnant rabbits were exposed to the test substance.
Effect on developmental toxicity: via oral route
- Endpoint conclusion:
- no adverse effect observed
Effect on developmental toxicity: via inhalation route
- Endpoint conclusion:
- no study available
Effect on developmental toxicity: via dermal route
- Endpoint conclusion:
- no study available
Additional information
In a prenatal developmental toxicity study according to OECD414, time-mated female Wistar Han rats were treated with 1-aminopropan-2-ol from Days 6 to 20 post coitum, inclusive, by daily oral gavage at dose levels of 100, 300 and 1000 mg/kg bw/d (2022). The rats of the control group received the vehicle, Water (Elix), alone.
Chemical analyses of formulations were conducted once during the study to assess accuracy and homogeneity. Formulation analyses confirmed that formulations of test material in Water (Elix) were prepared accurately and homogenously. The following parameters and end points were evaluated in this study for the F0-generation: mortality/moribundity, clinical signs, body weights, food consumption, thyroid hormone levels (triiodothyronine (T3), thyroxine (T4), thyroid-stimulating hormone (TSH)), organ weights (thyroid gland), macroscopic examination, microscopic examination (thyroid gland) and uterine contents (including corpora lutea, implantation sites, pre- and postimplantation loss, and number of live and dead fetuses). In addition, the following parameters were determined for the F1-generation: fetal body weights, sex ratio, anogenital distance, external, visceral and skeletal malformations and developmental variations.
Non-adverse changes in serum levels of the thyroid hormones T3 and T4 were observed in this study. These comprised a statistically signifanct lower T3 concentration at 1000 mg/kg bw/d and lower T4 concentrations starting at 100 mg/kg bw/d, not dose-dependent and without statistical significance in the mid dose. All group means remained within the available historical control data range. No test material-related changes were noted in any of the remaining maternal parameters investigated in this study (i.e., mortality/moribundity, clinical appearance, body weight, food consumption, thyroid-stimulating hormone (TSH), thyroid gland weights, macroscopic evaluation, microscopic evaluation of the thyroid gland, and uterine contents including corpora lutea, implantation sites and pre- and postimplantation loss).
No developmental toxicity was observed in the 100, 300 and 1000 mg/kg bw/d groups. No test material-related changes were noted in any of the developmental parameters investigated in this study (i.e,. litter size, sex ratio, fetal body weights, anogenital distance, external, visceral and skeletal malformations and developmental variations).
In conclusion, the maternal and developmental NOAELs for 1-aminopropan-2-ol were at least 1000 mg/kg bw/d.
In a weight-of-evidence approach, the read-across to TIPA (CAS 122-20-3) and Picloram TIPA salt (CAS 6753-47-5) is used to adapt the REACH Annex X standard information requirement for a pre-natal developmental toxicity study in a 2nd species (rabbit), which is lacking for the target substance MIPA (CAS no. 78-96-6). Supportingly, reproductive toxicity data in rats with MIPA HCl salt and the structural analogons DIPA (CAS 110-97-4) and TIPA (CAS 122-20-3) are included. The justification is prepared according to REACH Annex XI, section 1.5 and in line with the ECHA Guidance R.7a: Endpoint specific guidance (v6.0, July 2017), R.6 QSARs and grouping of chemicals [May 2008] and ECHA´s Read-Across Assessment Framework (RAAF, 2017).
(Justification documents are in IUCLID section 13.2: "Justification_WoE_RDT_Reprotox_Oct2022" and "Justification_RA_Dev.tox_2ndspecies_Nov2021").
A teratogenicity study in rabbits with Picloram TIPA salt was conducted according to OECD414 and GLP compliant [Dow, 1992; RL1]. In Phase I, groups of 18 inseminated New Zealand White rabbits were administered the test substance by oral gavage on GD7-19 at targeted dose levels of 0, 180, 538 or 1000 mg/kg bw/d. Phase II of this study was conducted as a repeat of Phase I to determine whether the increased resorption rate and certain fetal observations in Phase I were reproducible and/or attributed to treatment. In Phase II, an additional dose group of 54 mg/kg bw/d was added at the low end to establish a no-observed-effect level. On GD28, all surviving animals were necropsied and liver, kidneys and gravid uterine weights were recorded. The ovaries were examined for corpora lutea and the uterus was examined for implantations and resorptions. Fetuses were removed, weighed, sexed and examined for external, visceral and skeletal alterations. Maternal toxicity was observed in rabbits given 538 or 1000 mg/kg bw/d, as evidenced by decreased feed consumption accompanied by higher incidences of decreased feces, soft feces, perineal soiling and decreased body weight gain (Phase I and II). In addition, an increased incidence of high-dose dams aborting (one in Phase I and two in Phase II) was also interpreted to represent maternal toxicity. No other adverse maternal or embryonal/fetal effects attributed to treatment were observed at any dose level. The test substance was not teratogenic at any dose level tested. In conclusion, the maternal no-observed-adverse-effect level (NOAEL) was 180 mg/kg bw/d. Corrected for molecular weight this is equivalent to 79.56 mg/kg bw/d for TIPA. The no-observed-effect level (NOEL) for embryonal/fetal toxicity or teratogenicity of the test substance was 1000 mg/kg/d, the highest dose level tested; equivalent to 440 mg TIPA/kg bw/d.
The limit dose for an OECD414 study that needs to be tested is 1000 mg/kg bw/d. Based on this there might be remaining uncertainties for hazard characterisation. To further clarify these uncertainties, a short‐term repeated dose study was performed with New Zealand White rabbits [BASF SE, 2016; RL2; not added to table 1 for reasons of clarity]. TIPA was administered dissolved in drinking water for 21 days to groups of 3‐5 female New Zealand White rabbits, via daily gavage. Animals were exposed to either 0 or 1000 mg/kg bw/d. In the dose group (1000 mg/kg bw/d) all 5 females showed reduced food consumption and lost body weight throughout the study. These animals also had reduced faeces or diarrhoea. Two rabbits died on the 3rd and 5th treatment day, respectively. The deceased animals showed no other preterminal clinical signs of toxicity, nor were there any specific necropsy findings. In conclusion, if additional testing would be performed with TIPA, the dose‐selection for this study would be in the range of 500 mg/kg bw/d (half of the lethal dose), which is comparable to the converted dose of 440 mg TIPA/kg bw/d from the OECD414 rabbit study with Picloram TIPA salt (where no developmental toxic effects were observed).
In an OECD422 study in rats with a 65.4% solution of MIPA HCl salt (CAS 7780-04-3) [BASF, 2008; RL1], the dose levels were 100, 300, and 1000 mg/kg bw/d; equal to 67, 202 and 673 mg/kg bw/d organic MIPA. Only slightly reduced hemoglobin and hematocrit values were observed in the high dose male rats after 38 days of exposure, indicating a mild anemic process. No toxicity effects were observed in female rats after 45 days of exposure at all dose levels nor in the F1 pups and no developmental and fertility effects were observed. Based on the mild effects in the high dose male rats, the NOAEL for general, systemic toxicity of the test substance was set at the mid dose level (i.e. 202 mg/kg bw/d expressed as organic MIPA).
Supportingly, in OECD414 studies in rats with the analogons TIPA (CAS 122-20-3) and DIPA (110-97-4) the maternal and develpmental NOAELs were 1000 mg/kg bw/d, respectively.
Thus, based on the available reliable data, it is concluded, that MIPA induce neither pre-natal development toxicity nor teratogenicity.
Mode of Action Analysis / Human Relevance Framework
No information available.
Justification for classification or non-classification
The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. As a result the substance is not considered to be classified for reproductive toxicity under Regulation (EC) No. 1272/2008.
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