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Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information
The potential effects on the reproductive organs of male (testes, prostate, epididymides and/or seminal vesicles) and female (ovaries, uterus, mammary glands and/or vagina) rats were evaluated in three 1-month repeated dose toxicity studies with Diisopropylamine (DIPA) performed by the oral, dermal and inhalation routes (see repeated dose toxicity section). No effects on the reproductive organs were observed histologically with DIPA up to 150 mg/kg bw/day by the oral and dermal routes (Verschuere, 1991; Naylor, 1987) and 2000 mg/m3 by inhalation (Val Roloff, 1987), indicating that no effect on fertility can be assumed. 
Effect on fertility: via oral route
Endpoint conclusion:
no study available
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information
Short description of key information:
No data was generated on Diisopropylamine (DIPA) for the effects on fertility.

Effects on developmental toxicity

Description of key information
No data was generated on Diisopropylamine (DIPA) with respect of developmental toxicity. Therefore, read-across to an analogous substance, dimethylamine (CAS No. 124-40-3) is suggested. A developmental study was performed on dimethylamine hydrochloride in rat. In this study, a NOAEL for developmental toxicity has been derived at 1000 mg/kg.
Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study 414
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Limit test:
no
Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
Time-mated Wistar rats (Crl:WI[Han]) were supplied by Charles River Laboratories, Research Models and Services, Germany GmbH at an age of about 10-15 weeks. Only animals free from clinical signs of disease were used for the investigations.
The animals were paired by the breeder and supplied on GD 0 (= detection of vaginal plug/sperm). After arrival, they were randomly allocated to the test groups by withdrawal from the transport box at random and placed in to a random distribution of groups. After randomization the rats were identified uniquely by ear tattoo.
Reason for species selection: The Crl:WI(Han) strain was selected since extensive experience is available on Wistar rats. This specific strain has been proven to be sensitive to substances with a teratogenic potential.

Acclimatization period: from arrival to GD 6. (so from Day GD 0 to GD 6)

Housing: singly from GD 0-20 in type M III Makrolon cages supplied by BECKER & CO., Castrop-Rauxel, Germany (floor area about 800 cm²).
Bedding: Lignocel FS 14 fibres, dustfree bedding, supplied by SSNIFF, Soest, Germany
Enrichment: wooden gnawing blocks (Typ NGM E-022, supplied by Abedd® Lab. and Vet. Service GmbH, Vienna, Austria).

Accomodation: in fully air-conditioned rooms (central air conditioning)
Temperature: 20-24°C
relative humidity: 30-70%.
Air change rate: 10 times per hour.
The light cycle rhythm was 12 hours light from 6:00 a.m. to 6:00 p.m. and 12 hours darkness from 6:00 p.m. to 6:00 a.m.

Before the study started, the animal room was completely disinfected using a disinfector ("AUTEX" fully automatic, formalin-ammonia-based terminal disinfection). In general, each week the walls and the floor were cleaned with water containing about 0.5% Mikro-Quat (supplied by Ecolab Deutschland GmbH, Hanau, Germany).
Food: ground Kliba maintenance diet mouse/rat “GLP” supplied by PROVIMI KLIBA SA (Kaiseraugst, Switzerland).
Food: available ad libitum throughout the study (from the day of supply to the day of necropsy),
Drinking water: available from water bottles ad libitum

The food used in the study was assayed for chemical and for microbiological contaminants.
The drinking water was regularly assayed for chemical contaminants.
Bedding and the enrichment were regularly assayed for contaminants (chlorinated hydrocarbons and heavy metals).

Based on the pregnant animals the body weight on day 0 varied between 142.5-191.3 g.
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
The oral route was selected since this has proven to be suitable for the detection of a toxicological hazard.
A standard dose volume of 10 mL/kg body weight was used for each group.
The calculation of the volume administered was based on the most recent individual body weight.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Tested by various analyses:
• the stability of the test substance solutions was demonstrated over a period of 10 days at room temperature
• The results of the analyses of the test substance solutions in drinking water confirmed the correctness of the prepared concentrations. Generally, the analytical values of the samples corresponded to the expected values within the limits of the analytical method, i.e. were above 90% and below 110% of the nominal concentrations (see PART III; Supplement); except two deviant values in test groups 1 and 2. One deviant value in the samples from the study beginning (test group 2 = 88.8%) and one deviant value in the samples from the end of the study (test group 1 = 80.5%) were considered as outliers with respect to precision and accuracy of the analytical method. As no effects of toxicological concern were observed even at 1000 mg/kg bw/d, these minor deviations had virtually no effect on the quality of the study
Details on mating procedure:
The animals were paired by the breeder ("time-mated") and supplied on GD 0 (= detecion of vaginal plugs/sperm).
Duration of treatment / exposure:
administration from gestational day (GD) 6 through GD 19 = from implantation to one day prior to the expected day of parturition
Frequency of treatment:
Once daily
Duration of test:
On GD 20 all surviving females were sacrificed.
Remarks:
Doses / Concentrations:
0, 100, 300, 1000 mg/kg b.w.
Basis:
nominal in water
No. of animals per sex per dose:
25 females per dose group
Control animals:
yes, concurrent vehicle
Maternal examinations:

Food consumption and body weights of the animals checked regularly ( GD 0, 1, 3, 6, 8, 10, 13, 15, 17, 19 and 20).
Mortality: a check was made twice a day on working days or once a day on Saturdays, Sundays or on public holidays (GD 0-20).
Clinical symptoms: a cage-side examination was conducted at least once daily for any signs of morbidity, pertinent behavioral changes and signs of overt toxicity. If such signs occurred, the animals were examined several times daily (GD 0-20).
gross pathology (including weight determinations of the unopened uterus and the placentae).

Sacrifice of the animals on GD 20, then necropsied and assessed by gross pathology.
The uteri and the ovaries were removed and the following data were recorded:
- Weight of the unopened uterus*
- Number of corpora lutea
- Number and distribution of implantation sites classified as:
• live fetuses
• dead implantations:
a) early resorptions (only decidual or placental tissues visible or according to SALEWSKI (Salewski, 1964) from uteri from apparently non pregnant animals and the empty uterus horn in the case of single horn pregnancy)
b) late resorptions (embryonic or fetal tissue in addition to placental tissue visible)
c) dead fetuses (hypoxemic fetuses which did not breathe spontaneously after the uterus had been opened)
Ovaries and uterine content:
For each dam, corpora lutea were counted and number and distribution of implantation sites (differentiated by resorptions, live and dead fetuses) were determined
Fetal examinations:
Examinations of the fetuses after dissection from the uterus At necropsy each fetus was weighed, sexed, and external tissues and all orifices were examined macroscopically. The sex was determined by observing the distance between the anus and the base of the genital tubercle and was later confirmed by internal examination, in all fetuses designated for soft tissue examination. If there were discrepancies between the "external" and the "internal" sex of a fetus, the fetus was finally sexed according to the internal sex.

Furthermore, the viability of the fetuses and the condition of placentae, umbilical cords, fetal membranes, and fluids were examined. Individual placental weights were recorded. Thereafter, the fetuses were sacrificed by subcutaneous injection of a pentobarbital (Narcoren®; dose: 0.1 mL/fetus). After these examinations, approximately one half of the fetuses per dam were eviscerated, skinned and placed in ethanol, the other half was placed in Harrison’s fluid for fixation.

Soft tissue examination of the fetuses
The fetuses fixed in Harrison’s fluid were examined for any visceral findings according to the method of BARROW and TAYLOR (Barrow and Taylor, 1969). After this examination these fetuses were discarded.

Skeletal examination of the fetuses
The skeletons of the fetuses fixed in ethanol were stained according to a modified method of KIMMEL and TRAMMELL (Kimmel and Trammell, 1981). Thereafter, the skeletons of these fetuses were examined under a stereomicroscope. After this examination the stained fetal skeletons were retained individually.

Classifications based on terms and definitions proposed by CHAHOUD et al. and SOLECKI et al. (Chahoud et al., 1999; Solecki et al., 2001; Solecki et al., 2003):
- Malformation
A permanent structural change that is likely to adversely affect the survival or health.
- Variation
A change that occurs also in fetuses of control animals and is unlikely to adversely affect the survival or health. This includes delays in growth or morphogenesis that has otherwise followed a normal pattern of development.

Moreover, the term "unclassified observation" was used for those fetal findings, which could not be classified as malformations or variations.
All fetal findings were listed in tables according to these classifications.
Statistics:
DUNNETT-test (twosided)
FISHER'S EXACT test (one-sided)
WILCOXON-test (onesided)
Indices:
The conception rate (in %) was calculated according to the following formula:
(number of pregnant animals x 100)/number of fertilized animals
The preimplantation loss (in %) was calculated according to the following formula:
((number of corpora lutea – number of implantations) x 100) / number of corpora lutea
The postimplantation loss (in %) was calculated according to the following formula:
((number of implantations – number of live fetuses) x 100)) / number of implantations
Historical control data:
yes
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
There were no test substance-related mortalities in any of the female animals in any of the groups.
The mean food consumption of the high-dose dams (1000 mg/kg bw/d) was statistically significantly reduced between GD 6 to 8 (14% below control) and GD 8 to 10 (12% below control; see Fig. 4.2.1.3.1.). However, on the following days the food consumption of the high-dose rats became comparable to control. The average food consumption of the highdose dams during the treatment phase (GD 6-19) was less than 4% below the control value. The impaired food consumption is considered to be related to treatment. The food consumption of the females of test groups 1 and 2 (100 and 300 mg/kg bw/d) was unaffected and did not show any statistically significant or biologically relevant differences in comparison to the controls.
Mean body weight and mean body weight gain of low-, mid- and high-dose animals (100; 300 and 1000 mg/kg bw/d) were similar to those of the concurrent controls. All differences observed in these groups during the pretreatment and the treatment period were without biological relevance and reflected the normal variation inherent in the strain of rats used in the present experiment.
The mean gravid uterus weight of the dams of test groups 1; 2 and 3 (100; 300 and 1000 mg/kg bw/d) were comparable to the control and not affected by treatment.
At necropsy, no test substance-related findings were observed in the dams of test groups 0 - 3 (0; 100; 300 and 1000 mg/kg bw/d). One animal of test group 2 showed a diaphragmatic hernia (No. 56). This observation was not considered to be associated to the test compound

The conception rate reached 96% in the control group as well as in test group 1 (100 mg/kg bw/d), 92% in test group 2 (300 mg/kg bw/d) and 100% in test group 3 (1000 mg/kg bw/d). As all presumed pregnant rats had implantation sites at necropsy, a sufficient number of dams were available for the purpose of the study.
No test substance-related and/or biologically relevant differences with regard to conception rate, mean number of corpora lutea, implantation sites, pre- and postimplantation loss and resorptions (total, early and late) were observed

Test group 3 (1000 mg/kg bw/d)
Dams
• Salivation after treatment in 25 dams
• Statistically significant impairment of food consumption between GD 6-8 and GD 8-10.

All animals of the low-, mid- and high-dose groups showed yellowish discolored urine what has been considered to be treatment-related. Discolored urine occurred from GD 8 onwards and persisted until the end of the study. This urine discoloration was a sign of systemic availability of the test substance rather than being an adverse effect. It happened most likely due to the excreted test compound or its metabolite(s).
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Basis for effect level:
other: developmental toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects. Remark: up to 1000 mg/kg bw /day

Details on embryotoxic / teratogenic effects:
The sex distribution of the fetuses in test groups 1-3 (100; 300 and 1000 mg/kg bw/d) was comparable to the control fetuses. Observable differences were without biological relevance.
The mean placental weights of dose group 1 and 2 (100 and 300 mg/kg bw/d) were similar to the corresponding control. The mean placental weight of the male fetuses of dose group 3 (1000 mg/kg bw/d) was statistically significantly reduced (about 19% below the concurrent control value) but clearly in the range of the historical control data (PART III, Supplement). Therefore, the observed differences were not considered to be biologically relevant and without relation to dosing.
The mean fetal weights were not influenced by the test substance administration and did not show any biologically relevant differences between the test substance-treated groups and the control. The observable differences between the groups reflect the usual fluctuation for this parameter.
External malformations were recorded for one fetus in the control, for four fetuses in the lowdose group (100 mg/kg bw/d) and for four fetuses in the mid-dose group; (300 mg/kg bw/d; Tab. 4.3.2.1.1). Since two low-dose (100 mg/kg bw/d) fetuses of the same litter (dam no. 35) showed gastroschisis and two mid-dose (300 mg/kg bw/d) fetuses of the same litter (dam No. 58) had a menigocele it is reasonable to consider a spontaneous background in single animals rather than a test substance-induced effect. Furthermore, these findings did not show a relation to dosing (Tab. 4.3.2.1.2.). All other findings were incidental or can be found in the historical control data (PART III, Supplement).
No external variations were observed.
No unclassified external observations were recorded.

Soft tissue malformations were recorded for three fetuses in the low-dose group (100 mg/kg bw/d) and for three fetuses in the mid-dose group; (300 mg/kg bw/d; Tab. 4.3.3.1.1.). Since the low- (100 mg/kg bw/d) and the mid-dose (300 mg/kg bw/d) fetuses belonged to the same litters (dam no. 35 and dam no. 58, respectively) and with regard to the findings of the external observation, it is reasonable to consider a spontaneous background in single animals rather than a test substance-induced effect due to a missing dose-response relationship (Tab. 4.3.3.1.2.).

Fetuses
• No test substance-related adverse effects Test group 2 (300 mg/kg bw/d)
• No test substance-related adverse effects on dams, gestational parameters or fetuses Test group 1 (100 mg/kg bw/d)
• No test substance-related adverse effects on dams, gestational parameters or fetuses
Abnormalities:
not specified
Developmental effects observed:
not specified

Malformations of the fetuses: cleft palate, gastroschisis, exencephaly, mandibular micrognathia, malrotated limb (bilateral hindlimb) meningocele.

Soft tissue malformations:

Fetus with multiple visceral malformations: short intestine, large uterus horns (reaching to the middle of the kidneys), enlarged ovaries, persistent truncus arteriosus, heart: muscular ventricular septum defect, anophthalmia (bilateral),

Fetus with multiple visceral malformations: absent subclavian, persistent truncus arteriosus, heart: muscular ventricular septum defect, anophthalmia (bilateral)

anophthalmia (left)

Soft tissue variation:

Four soft tissue variations were detected, i.e. dilated cerebral ventricle, short innominate and uni- or bilateral dilation of renal pelvis and ureter. Dilated cerebral ventricle and short innominate occurred only in the male fetus 35-07 of the low-dose group and the male fetus 58-05 of the mid-dose group in addition to external, soft tissue and skeletal malformations. Uni- or bilateral dilation of renal pelvis and ureter were seen in several fetuses of all test groups including the control. These findings were considered to be incidental because they did not show a relation to dosing. (Tab. 4.3.3.2.1.). In addition, they can be found in the historical control data (PART III, Supplement) in comparable or even higher incidences.

Fetal skeletal malformations:

Skeletal malformations were noted in fetuses of test groups 1 and 2 (100 and 300 mg/kg bw/d; Tab. 4.3.4.1.1.). No dose-response relationship was observed (Tab. 4.3.4.1.2.). Based on the rate of affected fetuses per litter, the incidence of skeletal malformations was comparable to the historical control data (PART III, Supplement).

Fetal skeletal variations:

For all test groups, skeletal variations of different bone structures were observed, with or without effects on corresponding cartilages. The observed skeletal variations were related to several parts of fetal skeletons and appeared without a relation to dosing (Tab. 4.3.4.2.1.). Based on the rate of affected fetuses per litter, the incidence of skeletal variations was comparable to the historical control data (PART III, Supplement).

Fetal skeletal unclassified cartilage observation:

Two isolated cartilage findings without impact on the respective bone structures, which were designated as unclassified cartilage observations, were noted in all test groups or only in test group 2. These cartilage findings, i.e. bipartite processus xiphoideus and notched manubrium were related to the sternum. An association to the test substance is not assumed because the incidences of both observations were within the historical control range (PART III, Supplement).

Conclusions:
Thus, the oral administration of Dimethylamine hydrochloride to pregnant Wistar rats had no effect on morphology of offspring at any dose level tested (100; 300 and 1000 mg/kg bw/d). The recorded incidences did not suggest a treatment-relationship, but reflected the usual biological variation inherent in the strain of rats used for this experiment.
In conclusion, the no observed adverse effect level (NOAEL) for maternal toxicity is 300 mg/kg bw/d based on decreased food consumption and salivation after treatment in the highdose dams (1000 mg/kg bw/d). The no observed adverse effect level (NOAEL) for prenatal developmental toxicity is 1000 mg/kg bw/d because there was no evidence of an adverse effect of the test compound on fetal morphology.
Executive summary:

In an OECD 414 and GLP study, dimethylamine hydrochloride was administered by gavage at dose levels of 100, 300 and 1000 mg/kg bw/d to pregnant Wistar rats from implantation (GD 6) to one day prior to the expected day of parturition (GD 19). Dimethylamine hydrochloride did not cause any mortality. Test substance-related relevant clinical effects were only seen in the high-dose dams (1000 mg/kg bw/d), i.e. salivation after treatment and decreased food consumption, although the latter did not affect body weight, body weight gain, net body weight gain and uterus weight. At necropsy, no test substance related findings were noted in any of the dams. The temporary salivation was likely to be induced by the taste of the test substance or by local irritation of the upper digestive tract. It was not considered to be a sign of systemic toxicity. All animals of the low-, mid- and high-dose groups showed yellowish discoloured urine which occurred from GD 8 onwards and persisted until the end of the study. The urine discoloration was a sign of systemic availability of the test substance and happened most likely due to the excreted test compound or its metabolite(s). This finding has been considered to be treatment-related but was not assessed as an adverse effect. No differences of toxicological relevance between the control and the dose groups were determined for reproductive parameters such as conception rate, mean number of corpora lutea, mean number of implantations, pre- and postimplantation losses, live fetuses and fetal sex ratio. Examination of the fetuses revealed incidental fetal external, soft tissue and skeletal malformations in individual litters of the low- and the mid-dose groups as well as the control. Since malformations only occurred in one litter of the low- and one litter of the mid-dose group, it is reasonable to consider a spontaneous background in single animals rather than a test substance-induced effect. A consistent pattern and a dose-response relationship were missing. Thus, a test substance-related effect on ontogeny is not assumed. No external variation was noted. Four soft tissue and a broad range of skeletal variations occurred in every test group including the control. All of these variations are documented at a comparable frequency in the historical control data. A spontaneous origin is also assumed for the unclassified cartilage observations, which were recorded for fetuses of all dose-groups including the control. Character as well as distribution of all of these findings did not suggest a relation to treatment. In summary, there was no evidence of an adverse effect of Dimethylamine hydrochloride on fetal morphology at any dose level tested.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Species:
rat
Quality of whole database:
Read across to dimethylamine
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 study conducted according to OECD TG 414 (Prenatal Developmental Toxicity Study), groups of 25 time-mated female Wistar rats per group were administered dimethylamine hydrochloride (DMA HCl; CAS Number 506-59-2), a structural analogue to diisopropylamine, by oral gavage at doses of 100, 300 and 1000 mg/kg bw from gestation day (GD) 6 through 19 (BASF SE, 2009a). There were no deaths. Clinical signs at 1000 mg/kg bw/d included salivation after treatment and decreased food consumption. All animals of the low-, mid- and high-dose groups showed yellowish discolored urine which occurred from GD 8 onwards and persisted until the end of the study. The urine discoloration was a sign of systemic availability of the test substance and happened most likely due to the excreted test compound or its metabolite(s). This finding has been considered to be treatment-related but was not assessed as an adverse effect. There were no effects on body weight, body weight gain, net body weight gain and uterus weight. At necropsy, there were no gross findings. There was no effect on reproductive parameters (conception rate, mean number of corpora lutea, mean number of implantations, pre- and post-implantation losses, live fetuses and fetal sex ratio). However, placental weight at 1000 mg/kg bw/day was 19% lower than controls. Examination of the fetuses revealed incidental fetal external, soft tissue and skeletal malformations in individual litters of the low- and the mid-dose groups as well as the control. Since malformations only occurred in one litter of the low- and one litter of the mid-dose group, it is reasonable to consider a spontaneous background in single animals rather than a test substance-induced effect. A consistent pattern and a dose-response relationship were missing. No external variation was noted. Four soft tissue variations and a broad range of skeletal variations occurred in every test group including the control. All of these variations were documented at a comparable frequency in the historical control data. There was no evidence of an adverse effect of DMA HCl on fetal morphology at any dose level tested. The NOAEL (maternal toxicity) was 300 mg/kg bw/d, based on salivation after treatment at 1000 mg/kg bw/d. The NOAEL (developmental toxicity) was 1000 mg/kg bw/day as there was no evidence of an adverse effect of the test compound on fetal morphology.

Justification for selection of Effect on developmental toxicity: via oral route:
Key study

Toxicity to reproduction: other studies

Additional information

The histological effects of Diisopropylamine (DIPA) on the reproductive organs of male and female rats were evaluated in:

- a repeated oral toxicity performed according to OECD N°407 guideline (Repeated Dose 28-Day Oral Toxicity in Rodents) (Verschuere, 1991). Groups of 15 male and 15 female Sprague Dawley rats were administered orally once daily for 4 weeks with diisopropylamine at dose levels of 0, 15, 50, and 150 mg/kg bw/d,

- a repeated dermal toxicity performed according to a protocol similar to OECD TG #410 guideline. Groups of 10 male and 10 female Charles River CD(SD)BR rats received an open dermal exposure to 0, 15, 50, and 150 mg/kg bw/d of undiluted Diisopropylamine five times a week for 1 month,

- a repeated inhalation toxicity performed following a protocol similar to OECD N°412 guideline (Repeated Dose 28/14-Day inhalation Toxicity in Rodents).Groups of 15 male and 15 female Sprague-Dawley rats were exposed to 100, 600 and 2000 mg/m3 Diisopropylamine for 6 h per day, 5 days a week for 1 month,

no treatment-related effects were observed on mammary tissue, uterine cervix, ovaries, uterine tubes, vagina, prostate, testes, seminal vesicles and/or epididymides.

Justification for classification or non-classification

According to CLP criteria, no classification is warranted for reprotoxicity.