Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

The review of six studies/publications for sodium salts of HEDP and of three studies for other structurally-related homologues gave no indication for a potential of phosphonates to affect fertility in rats, dogs or mice.

Additional information

The substance under registration can chemically be described as a sodium salt of HEDP. Information on reproduction toxicity is not available for this specific sodium salt, but data are available for other sodium salts of HEDP including the salts with the low and high pHs which belong to the same category. Altogether, six studies covering several study types (90-day study, combined chronic tox./carcinogenicity study, dominant-lethal-assay, two-generation reproduction study) are available which provide information on fertility. The tested species were rats, dogs and mice. In the two-generation key study, oestrus cycles but not sperm parameters were assessed. However, no effects on spermatogenesis or testes were observed in the second key study (combined chronic toxicity/carcinogenicity test) or subchronic studies.

In all studies, no effects on reproductive functions were observed up to the highest dose level tested. An overview on all studies and results is given in the table below.

Entry

Test type / species/ author

Klim.

Scope (reproduction parameters)

Deficiencies

Result (Fertility)

1

Key

Two-generation reproduction toxicity study (rat)

 

Nolen & Buehler, 1971

2

Pregnancy rate, number of resorptions, corpora lutea and implantations, number of live/dead fetuses, weight of fetuses/pups/dams; determination of estrus cycles by vaginal smear, feeding from week 5 onwards

Use of 2 instead of three dose levels; Sexual milestones in pups/analytical and confirmation of dose not given; No sperm analyses. Not GLP.

NOAEL = 112 mg/kg bw/d

2

Key

Combined chronic toxicity / carcinogenicity study (rat)

 

Huntingdon Research, 1979

2

Organ weights and histology of testes, seminal vesicles, prostate, uterus, mammary (gland);

Spermatogenesis, atrophy, dilation of tubules, periarteritis

Only 40 instead of 50 animals were used. Not GLP.

NOAEL >= 384 mg/kg/day in male rats and >= 493 mg/kg/day in female rats.

3

Supp

90-day study oral toxicity study (rat),

 

Huntingdon Research, 1977

2

No observable effect, with respect to the testes, ovaries and associated organs

Comparable to OECD, not GLP

NOAEL >= 817 mg/kg bw/d (m) or >= 1000 mg/kg bw/d (f)

4

Supp

Dom.- lethal- test (mouse),

 

Pieper K., 1974

2

Conception rates, total implant averages, fetal death averages, resorption percentage, mutagenic indices

Postive control substance was not included in this study (not necessarily required). Not GLP.

Not toxic to reproduction in doses up to 1000 mg/kg bw/d, (given 5 times, on 5 consecutive days)

5

Supp

90-day oral toxicity study (dog),

 

Industrial Biotest, 1975

2

Organ weights of gonads, histopathology of gonads (males: testes and epididymides), prostate gland and uterus

 

Pre-GLP/-OECD.

NOAEL 1746 mg active acid/kg bw/d (males) and 1620 mg active acid/kg bw/d (females)

6

Supp

90-day oral toxicity study (rat)

 

Marias, A., 1975

2

Organ weights of gonads, microscopic examination of tissues (gonads, seminal vesicles, uterus, prostate gland)

 

Not GLP

NOAEL >= 900 mg/kg bw/d

 

Further information on fertility is available from reproduction toxicity studies conducted with the structurally-related homologue phosphonates DTPMP-H (CAS 15827-60-8, entry 7), ATMP-H (CAS 6419-19-8, entry 8) and HEBMP-xNa (CAS 22036-78-8, entry 9). Those studies support in a weight-of-evidence approach the hypothesis that phosphonates do not induce specific effects on fertility. An overview on these data obtained with structurally related phosphonates is given in the table below.

 

Entry

Test type / species / CAS

Klim.

Scope (reproduction parameters)

Deficiencies

Result (Fertility)

7

WOE

One generation study (rat)

2

Organ weights and histology of testes, seminal vesicles, prostate, uterus, mammary (gland);

Spermatogenesis, atrophy, dilation of tubules, periarteritis

No analytical confirmation of exposure, no pre-mating exposure, no assessment of oestrus cycle or sperm analyses, no evaluation of sexual milestones in pup. Males treated only as F1 and F2 generation.

NOAEL (P) >= 882 mg/kg bw/d (m) and >= 936 mg/kg bw/d (f)

NOAEL (F1, F2)

= 294 mg/kg bw/d (m) and 312 mg/kg bw/d (f)

8

WOE

Three-generation reproduction study (rat)

2

Organ weights and histopathology of gonads

No assessment of estrus cycle, sperm parameters, sexual milestones, no analytical confirmation of exposure levels. Not GLP.

NOAEL >= 275 mg/kg bw/d (m) and >= 310 mg/kg bw/d (f)

9

WOE

OECD 422 (rat)

1

Conception rates, total implant averages, fetal death averages, resorption percentage, mutagenic indices

None

NOEL >= 1000 mg/kg bw/d

 

Justification for grouping:

See CSR Annex I and II or IUCLID section 13.



Effects on developmental toxicity

Description of key information

No guideline pre-natal developmental toxicity studies are available for sodium salts of HEDP. However, a combined two-generation study of reproductive toxicity and teratogenicity in rats as well as a prenatal developmental toxicity study in rabbits was published in the peer reviewed literature (Nolen & Buehler, 1971).  In addition, pre-natal studies according to OECD guideline 414 are available for several related phosphonic acid compounds from the ATMP, HEBMP, and DTPMP categories which indicate that these substances are not teratogenic following oral treatments in rats and mice between 312 and >= 1000 mg/kg bw/day during pregnancy. In conclusion, all available data have revealed no adverse effects on development in any of the tested species (rat, rabbit, mouse). On the basis of the key study with the most sensitive species (rabbit), and in consideration of the overall weight of evidence, a developmental NOAEL of >= 100 mg/kg bw/day appears appropriate for HEDP and its salts.

Additional information

The two key studies with HEDP salts which were published in peer-reviewed literature (Nolen & Buehler, 1971) include data on two species (rats, rabbits) and two different study designs (two-generation study, teratogenicity). In general, the tests meet the scientific principles of the corresponding OECD guidelines No. 416 and No. 414. The deviations relevant for the assessment of developmental toxicity have been examined. Though only two doses were tested instead of three, the dose selection can be considered appropriate to evaluate the teratogenic potential, as maternal toxic effects were induced at the highest dose. In the developmental toxicity study with rabbits, the dosing took place from day 2-16 instead of day 6-18. This time span was considered best practice at the time the test was performed. In spite of deviations from the guidelines, adequate information is provided in the two key studies to conclude that HEDP and its salts do not produce any adverse on developmental toxicity in two species (rat, rabbit). Details of the two studies are discussed below.

 

In a pre- OECD combined two-generation study of reproductive toxicity and teratogenicity, five groups of 22 female and 22 male weanling Charles-River rats were given the disodium salt of HEDP (disodium etidronate) at a dietary concentration of 0, 0.1 or 0.5% (equivalent to a dose of 0, 112 or 447 mg/kg bw per day), either continuously or only on days 6–15 of gestation for two generations. Reproductive endpoints and offspring parameters were analyzed in the F1a, F1b, and F2a litters. The third litter of the F1 generation (F1c) and the second litter of the second generation (F2b) were used in teratological examinations. During the teratology phase, half of the animals in each group were sacrificed at day 13 and the others at day 21 of gestation. Body-weight gains were similar for all groups in both generations, and the overall conception rate was 90%, indicating that the compound did not interfere with spermatogenesis or ovulation. In the first generation, at the highest dose, the number of pubs born in the first litter (F1a) was reduced and there was an increase in stillborn pups in the second litter (F1b). The rate of mortality in pups after birth was low and the weights of pups at day 4 and at weaning were the same. Teratological examination of the third litter (F1c) showed no differences in resorptions or implantations in females sacrificed at day 13 and no differences in live fetuses, corpora lutea, or implantation at 21 days. At day 21, however, significant resorptions were reported in the controls. In the second generation, the first litters (F2a) were smaller than the litters in the first generation, but there were no other differences in reproductive parameters. During the teratology phase, no differences in corpora lutea, implantations, or resorptions were noted in rats sacrificed at 13 days. In continually fed rats sacrificed at 21 days, the number of implantations was reduced and corpora lutea formation was depressed at the highest dose. A decrease in the number of live fetuses at the highest dose, significant only in rats fed during gestation, was also observed. The incidence of defective pups was similar to that in control animals.

The NOAEL for maternal toxicity was set at 112 mg/kg bw per day (Nolen & Buehler, 1971). The study authors concluded that disodium etidronate was not teratogenic in rats at either dose tested (NOAEL developmental toxicity: >= 447 mg/kg bw per day).

 

A preparatory study inrabbits (Nolen and Buehler, 1971) found that doses of 500 mg/kg bw/day in water by intubation induced maternal death, so the doses were reduced to 250 mg/kg bw/day in four surviving animals for the rest of the study. The results show that during pregnancy rabbits are more sensitive than rats towards maternal toxicity and that - based on a limited number of remaining maternal animals - a dose of 250 mg/kg bw/day is a potential NOAEL for developmental toxicity in rabbits. In the subsequent feed study no evidence of maternal or fetal effects was apparent when rabbits were fed diets designed to deliver the equivalent of 25, 50 or 100 mg disodium etidronate/kg bw/day from day 2 through to day 16 of pregnancy (Nolen and Buehler, 1971). There were no apparent treatment-related effects on pregnancy rates, numbers of corpora lutea, implantations, resorptions or live fetuses and their weights, in either of the studies in rabbits. Nor was there a treatment-related increase in teratogenic effects observed in either study. a reduction in fetal weights with HEDP at a dose of 100 mg/kg bw per day administered by gavage, was attributed by the study authors to slightly larger litters. The combined results from the preparatory study and the main study indicate that the NOAEL for maternal toxicity in the rabbit is at least 100 mg disodium etidronate/kg bw/day. The NOAEL for developmental toxicity was concluded to be >= 100 mg/kg bw/day on the basis of 886 rabbit fetuses examined in the main study.

 

Two publications on sodium salts of HEDP (King, 1967 and Eguchi, 1982) are not considered for the derivation of a NOAEL for developmental toxicity because of missing background information and methodological deficiencies. However, the information given in those publications was evaluated within a weight-of-evidence approach.

 

Further evidence for the assessment of developmental toxicity is provided by a study with only few data available on the potential impact of the parent etidronic acid on fetal development (Monsanto plc 1966). 20 female Long-Evans rats per dosing group were tested according to the FDA guidelines for reproduction studies for safety evaluation of drugs for human use. They were given 16.5, 110 or 330 mg/kg bw/d by gavage, 2 times daily half of the total daily dose, during days 6 -15 of gestation. For all test material concentrations no effect was observed, therefore a NOAEL => 330 mg/kg bw/d was concluded.

 

In a non-standard dietary teratology study (King, 1967), only available as a brief summary, the NOAEC for HEDP was at least 0.5% HEDP in diet in rats.

 

Eguchi et al. (1982) reported significantly reduced body weights of the live fetuses and a disturbance of bone development in female mice administered HEDP by subcutaneous injection at 200 mg/kg bw/d on gestation days 11-17.

Having regard to likely (non medical) routes of human exposure subcutaneous injection is not an appropriate route of administration. The effects might be very well triggered by maternal toxic effects although an examination of maternal toxicity seemed not to be part of the Eguchi study. Based on the assumption that gastrointestinal absorption of orally administered HEDP would occur at a rate of <= 10% (see summary of toxicokinetics), a subcutaneous dose of 200 mg/kg bw would correspond to an exaggerated oral dose of ca. 2000 mg/kg bw, which is much higher than the expected maternal toxic dose for mice. Additionally, no first pass effect in the liver occurs after subcutaneous administration and differences in the disposition of HEDP are likely related to the uncommon route of administration. Nevertheless, the described effects can not be excluded at exaggerated doses, and would not be in contrast to the proposed mode of action for general systemic effects at high doses, namely the complexation of essential metals leading e.g. to calcium and iron deficiency. Another explanation for the reduced body weights of the fetuses would be the higher number of fetuses counted in the dose group (7.4 per dam) compared to the control group (5.2 per dam). Last but not least, single treatment level, use of a non preferred rodent species, low number of animals (control: 6, treatment group 7), and generally poor reporting of methods and results impose limitations on the overall suitability of the study for a quantitative risk assessment.

  

Several other results from guideline developmental toxicity studies in rats and mice performed using structurally-related phosphonic acid analogues (ATMP and salts; HEBMP salt, DTPMP and its salts) are included in the dossier as part of the weight of evidence approach. The related phosphonic acid compounds from the ATMP, HEBMP, and DTPMP categories are not teratogenic following oral treatments between 312 and >= 1000 mg/kg bw/day during pregnancy.

 

Based upon these considerations, a developmental NOAEL of >= 100 mg/kg bw/day appears appropriate for HEDP and its salts.

Justification for grouping:

See CSR Annex I and II or IUCLID section 13.

Justification for classification or non-classification

Based on the available data, no classification is required for the reproductive and developmental toxicity of HEDP sodium salt according to Regulation (EC) No 1272/2008.