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Effects on fertility

Description of key information

In line with ECHA Final Decision number CCH-D-2114495836-29-01/F an extended one-generation reproductive toxicity study in rats, according to OECD Test Guideline 443, has been planned for the Category member DTPMP (5-7Na). The deadline for submitting the requested information is 22 May 2023.

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

In line with ECHA Final Decision number CCH-D-2114495836-29-01/F an extended one-generation reproductive toxicity study in rats, according to OECD Test Guideline 443, has been planned for the Category member DTPMP (5-7Na). The deadline for submitting the requested information is 22 May 2023.

The reproductive toxicity study with DTPMP-H in the rat (BioDynamics Inc., 1979) was completed prior to the establishment of GLP and OECD guidelines. Many endpoints considered relevant to an assessment of reproductive toxicity thus are missing from the study (e.g., oestrous cyclicity and sperm parameters, pubertal assessment, anogenital distance, thyroid hormone levels). Nevertheless, the study includes a large number of females per parental generation and the production of two generations of rats. Therefore, the study provides data relevant to an assessment of developmental and reproductive toxicity. It should also be noted that, although titled as a one-generation study, the F1 animals actually were mated to produce two litters (F2a/F2b); thus, the study is more appropriately considered to be a two-generation reproductive study.

The test substance was administered in the diet to Long-Evans rats at dietary concentrations of 0, 300, 1000, and 3000 ppm beginning at the initiation of mating through the production of two generations (F1 and F2a/F2b). Test substance administration to the P0 generation females (20/group) was initiated at the onset of gestation and continued throughout the ensuing gestation and lactation periods of F1. Then, F1 rats (n=10 males and 20 females/ group) were first mated approximately 80 days after weaning, with a 14-day rest period after weaning of the first litter (F2a) before mating for production of the second litter (F2b). Rats of the F2a litters were sacrificed at weaning (postnatal day [PND] 21). Of relevance to the reproductive system, the testes and ovaries of F1 parental animals were weighed, and the following organs were preserved and evaluated histopathologically in 5 F1 adults/sex in the control and high dose groups: ovaries, testes, uterus and prostate.

No treatment-related effects were observed on parental (P0 and F1) survival, clinical signs, or food consumption. No treatment-related gross findings were reported. Both absolute and relative organ weights were comparable across the groups and no treatment-related histopathologic changes were observed in F1 animals. Although F1 female adult body weights were slightly, but not statistically, lower during lactation of the F2b litters, body weight change was generally comparable across groups and similar differences were not observed during the F2a lactation period or for P0 animals during lactation of the F1 pups. Thus, it is unlikely that this finding was related to treatment.

Most reproductive and litter parameters (mating and fertility indices, gestation length, litter size, pup sex ratio) were comparable across the groups with some minor exceptions, as detailed below.

For the F1 litter, at 3000 ppm, the mean number of live pups was slightly, but not statistically significantly, lower compared to control; the live birth index was significantly reduced at this exposure level as a result. However, differences across groups in F1 pup survival showed no clear relationship with dose and similar findings regarding live litter size were not observed for the F2a or F2b litters.

F1 pup body weights at birth were non-statistically significantly lower than control at 3000 ppm; F2a pup weights were lower at 1000 and 3000 ppm (statistically significant at 3000 ppm only). However, in both cases, pup weights during weaning were comparable across groups and a similar finding was not seen for the F2b pups.

F1 adult animals at 3000 ppm exhibited a slightly lower rate of pregnancy compared to controls for generation of the F2a litters; however, the difference was not statistically significant, nor seen for the P0 animals or repeated with generation of the F2b litters. Additionally, F1 gestation length was reduced at 1000 ppm only for generation of the F2a litters, but not for generation of the F2b litters. Because the change in pregnancy rate was not statistically significant, and due to the lack of consistency across generations and litters, these findings are thus unlikely to have been treatment related.

In summary, the reproductive study of DTPMP-H failed to demonstrate any consistent changes indicative of a treatment-related effect. Thus, this study is supportive of there being no adverse effect of DTPMP treatment on development or reproduction at doses that did not cause toxicity in parental animals. Therefore, the reproductive and systemic NOAELs were concluded to be at least 3000 ppm, equivalent to 294 mg/kg bw/day for males and 312 mg/kg bw/day for females.

For more discussion on the findings in this study please see the Expert Review of Developmental and Reproductive Toxicity Data for ATMP, DTPMP & EDTMP (DeSesso, 2021) attached to IUCLID Section 13.


Effects on developmental toxicity

Description of key information

In the prenatal developmental toxicity study with DTPMP-7Na (Monsanto, 1982) clear maternal toxicity (approx. 30% decrease in body weight gain, soft stools) was noted in pregnant Sprague-Dawley rats given 2000 mg/kg bw/day DTPMP-7Na (expressed as active acid) on gestation days 6-19, therefore the NOAEL for maternal toxicity was concluded to be 1000 mg/kg bw/day. The NOAEL for developmental toxicity was concluded to be 2000 mg/kg bw/day (active acid) based on no adverse treatment-related developmental effects at the highest dose level tested. Although completed prior to the establishment of OECD guidelines, this study generally meets current guideline recommendations with the exception that the highest dose tested exceeded the limit dose of 1000 mg/kg/day. Additionally, both the in-life portion of the study (done at Monsanto) as well as the foetal examinations (done at WIL Research Laboratories) were conducted according to GLP.

In line with ECHA Final Decision number CCH-D-2114495836-29-01/F a prenatal developmental toxicity study in second species (rabbit), according to OECD Test Guideline 414, has been planned for the Category member DTPMP (5-7Na). The deadline for submitting the requested information is 22 May 2023.

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
21.05.1980 to 09.06.1980
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
No record of gravid uterine weight, number of corpora lutea not recorded, no analytical confirmation of dosing solutions.
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River
- Age at study initiation: No data
- Weight at study initiation: 180-200 g
- Fasting period before study: No data
- Housing: Individually in suspended stainless steel mesh cages.
- Diet (e.g. ad libitum): Ad libitum
- Water (e.g. ad libitum): Ad libitum
- Acclimation period: None, animals were received mated and allocated to cages.


ENVIRONMENTAL CONDITIONS
- Temperature (°F): 72 ±2
- Humidity (%): 40-60
- Air changes (per hr): No data
- Photoperiod (hrs dark / hrs light): 12/12


IN-LIFE DATES: From: 26.05.1980 To: 09.06.1980
Route of administration:
oral: gavage
Vehicle:
other: Aqueous solution
Details on exposure:
No data
Analytical verification of doses or concentrations:
no
Details on mating procedure:
No data. Animals were received on gestational day 1, having already been mated.
Duration of treatment / exposure:
GD 6 - 19
Frequency of treatment:
daily
Duration of test:
20 days
Dose / conc.:
500 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
Dose / conc.:
2 000 mg/kg bw/day
No. of animals per sex per dose:
25
Control animals:
other: 0.9% (w/v) aqueous NaCl
Details on study design:
- Dose selection rationale: No data
Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily for visible toxic effects.


DETAILED CLINICAL OBSERVATIONS: No


BODY WEIGHT: Yes
- Time schedule for examinations: Gestational days 3, 6, 8, 10, 13, 15, 17 and 20.


FOOD CONSUMPTION: No


WATER CONSUMPTION: No


POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: Gross examination of all animals that included examination of external surfaces and thoracic and abdominal cavities.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: yes
Examinations included:
- Gravid uterus weight: No
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late 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: No data
Statistics:
Comparison of body weights between treatment and control groups was performed using Dunnett's test. Counted data (corpora lutea, implants, resorptions, live and dead fetuses) and data expressed as percentage were analysed, when appropriate with the Mann-Whitney U test. Response data (pregnancy rates, number of litters with post-implantation loss, and fetuses or litters with abnormalities and variants) were analysed, when appropriate, with Fisher's exact test and the chi-square test.
Indices:
None
Historical control data:
No data
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
9/25 dams had soft stools in the 2000 mg/kg bw/day group beginning on gestation day 14 (9th day of the treatment) and persisted through to gestation day 17. This finding was not observed in the other treated groups or the controls.
Mortality:
no mortality observed
Description (incidence):
No deaths occurred prior to the scheduled sacrifice.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
The only statistically significant (P<0.01) effect observed was lower body weight gain (mean value approximately 68% of the control mean) between gestation day 6 and 20 for dams in the 2000 mg/kg bw/day group (33.6/27.6/27.5/22.9). However, terminal body weights were not statistically significantly affected (mean terminal body weights with uterine contents by dose: 360.1/368.1/357.0/346.1 and mean terminal body weights without uterine contents: 265.8/264.5/260.1/257.2).
Food consumption and compound intake (if feeding study):
no effects observed
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:
no effects observed
Gross pathological findings:
no effects observed
Description (incidence and severity):
There were no treatment-related findings in the gross necropsy.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
There were no significant differences between any of the treatment groups and the control group in the number of pre- or postimplantation losses.
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
There were no significant differences between any of the treatment groups and the control group in the resorptions.
Early or late resorptions:
no effects observed
Description (incidence and severity):
There were no significant differences between any of the treatment groups and the control group in the resorptions.
Dead fetuses:
no effects observed
Description (incidence and severity):
There were no significant differences between any of the treatment groups and the control group in the mean number of live fetuses.
Changes in pregnancy duration:
no effects observed
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
There was no effect on pregnancy rate.
Other effects:
no effects observed
Description (incidence and severity):
There was no effect on the mean number of corpora lutea.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
act. ingr.
Remarks:
active acid
Basis for effect level:
body weight and weight gain
Abnormalities:
effects observed, treatment-related
Localisation:
other: body weight gain
Fetal body weight changes:
no effects observed
Description (incidence and severity):
There were no effects on fetal body weight.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
There were no significant differences between any of the treatment groups and the control group in the mean number of live fetuses.
Changes in sex ratio:
no effects observed
Description (incidence and severity):
There were no effects on sex ratios (males/litter: 5.6/5.8/5.9/4.8. Females/litter: 5.9/6.5/5.4/6.4).
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
not examined
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
There were no treatment-related gross external malformations present.
Subcutaneous haematomas were present in controls and all treated groups, however the incidence increased at 500 mg/kg bw/day (P<0.05) and was considered unrelated to treatment by the study authors (no dose relationship was present). The occurrence of one female fetus that was hydrocephalic and one female with gastroschisis in the 1000 mg/kg bw/day group was considered by the study authors to be spontaneous. The hydrocephalic female also exhibited a developmental variation (underdeveloped renal papilla) that was not considered to be related to treatment.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
Skeletal examination revealed single incidences of dwarfism (one female fetus of 500 mg/kg bw/day group) and fused sternebrae (one female fetus of the 2000 mg/kg bw/day group), which were considered spontaneous. Two fetuses from different litters in the 2000 mg/kg bw/day group and one fetus of the 1000 mg/kg bw/day group had vertebral anomalies (missing, reduced or fused vertebral arches). Although the incidence of these anomalies was not statistically significant compared with the control group, the rare spontaneous occurrence of such anomalies and the pattern of incidence indicated that they might have been treatment-related. Various skeletal variations occurred, but none showed a clear dose-response and so were considered spontaneous. Review of these results for the REACH assessment concluded that the effects should not be considered adverse as they were not statistically significant and were only observed in the presence of maternal toxicity. The limit dose for a prenatal developmental toxicity study conducted to OECD 414 is 1000 mg/kg bw/day, therefore the high dose used in this old study was double the limit dose. When findings up to the limit dose are considered, vertebral anomalies were observed in only one fetus. Therefore, this finding is not considered adverse.
Visceral malformations:
no effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
>= 2 000 mg/kg bw/day
Based on:
act. ingr.
Remarks:
active acid
Sex:
male/female
Basis for effect level:
other: No adverse developmental effects observed.
Abnormalities:
no effects observed
Developmental effects observed:
no
Conclusions:
In the prenatal developmental toxicity study with DTPMP-7Na, conducted according to a protocol similar to OECD Test Guideline 414 and in compliance with GLP, clear maternal toxicity (approx. 30% decrease in body weight gain, soft stools) was noted in pregnant Sprague-Dawley rats given 2000 mg/kg bw/day DTPMP-7Na (expressed as active acid) on gestation days 6-19, therefore the NOAEL for maternal toxicity was concluded to be 1000 mg/kg bw/day. The NOAEL for developmental toxicity was concluded to be 2000 mg/kg bw/day (active acid) based on no adverse developmental effects.
Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
2 000 mg/kg bw/day
Study duration:
subacute
Species:
rat
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 the prenatal developmental toxicity study with DTPMP-7Na (Monsanto, 1982) mated female Sprague-Dawley rats (25/dose) were given daily oral gavage doses of 500, 1000, 2000 mg/kg bw/day DTPMP-7Na in 0.9% sodium chloride (expressed as active acid) during gestation days 6 to 19. Control animals were given 0.9% sodium chloride solution. On day 20 all surviving animals were sacrificed and foetuses were examined. Although completed prior to the establishment of OECD guidelines, this study generally meets current guideline recommendations with the exception that the highest dose tested exceeded the limit dose of 1000 mg/kg bw/day. Additionally, both the in-life portion of the study (conducted at Monsanto) as well as the foetal examinations (conducted at WIL Research Laboratories) were conducted according to GLP.

There were no deaths prior to scheduled sacrifice. Findings consistent with maternal toxicity were noted at the highest dose tested (2000 mg/kg bw/day), including soft stools beginning GD 14, and a reduction in maternal body weight gain of approximately 32%; however, terminal body weights were only marginally different from control (3%-4%). No treatment-related lesions were detected at gross necropsy of the dams of any treatment group. All groups generally showed low pregnancy rates, but since mating occurred prior to dosing, this finding is not related to treatment. No significant treatment-related effects were observed on pregnancy or litter parameters, although foetal weights at the high dose were approximately 7% lower than control. No external malformations were observed at the foetal examinations. A single foetus with hydrocephalus and a single foetus with gastroschisis were reported at the middle dose of 1000 mg/kg bw/day; gastroschisis (in which the intestines protrude from the abdominal cavity) is a finding typically observed at external examination, so its reporting as a visceral finding only is unusual. Importantly, no visceral findings were observed at 2000 mg/kg bw/day. Few skeletal anomalies were observed in the study; these findings occurred at single instances and/or without dose-response, and thus, their relation to treatment is unlikely. Among the skeletal observations are vertebral anomalies in one and two foetuses at 1000 and 2000 mg/kg bw/day, respectively, the incidence of which was not statistically significant and the effect was only observed in the presence of maternal toxicity. These findings involve minor reductions in ossification, extra ossification centres and bony attachments between bones that are likely transient. Few foetal variations were reported and those of the skeletal system primarily related to the degree of bone ossification (data not shown). None of the variations were observed with a dose-response with the exception of unossified sternebrae #5 and/or #6, which was increased in foetal incidence at the high dose. In summary, these data show that no treatment-related developmental toxicity occurred at DTPMP doses above the limit dose of 1000 mg/kg bw/day (2000 mg/kg bw/day). The NOAEL for maternal toxicity was concluded to be 1000 mg/kg bw/day based on the lower body weight gain, while the NOAEL for developmental toxicity was concluded to be 2000 mg/kg bw/day (active acid) based on no adverse treatment-related developmental effects.

For more discussion on the findings in this study please see the Expert Review of Developmental and Reproductive Toxicity Data for ATMP, DTPMP & EDTMP (DeSesso, 2021) attached to IUCLID Section 13.

Information is also available on DTPMP-H from a one-generation reproductive toxicity study (Biodynamics, 1979b), conducted prior to OECD Test Guidelines and GLP, in which DTPMP-H was administered continuously via the diet to Long-Evans rats at concentrations of 300, 1000 and 3000 ppm through one complete generation. Test substance administration to the F0 generation females (20/group) was initiated at the onset of gestation and continued throughout the ensuing gestation and lactation periods. Administration then continued to the F1 generation animals (10 males and 20 females/group) through a growth period and mating, gestation and lactation period for two successive litters. In the F0 generation, no treatment-related effects in the low and mid dose groups were evident. In the high dose group, females delivered litters containing fewer live pups and more dead pups (not statistically significant) resulting in a lower live birth index. Pups also had a lower weight at birth (not statistically significant). No other treatment-related effects were observed. In the F1 generation, no treatment-related effects were evident in the low dose group. No such effects were observed in the second litters and no other treatment-related effects were observed during the remainder of the study. Gross pathological examination of five adult F1 generation males and females of the control and high dose groups did not reveal any abnormal findings. The NOAEL was concluded to be 3000 ppm, equivalent to 294 mg/kg bw/day for males and 312 mg/kg bw/day for females.

Justification for classification or non-classification

Based on the available data, no classification for reproductive and developmental toxicity is required for DTPMP (5-7Na) according to Regulation (EC) No 1272/2008.

Additional information