Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

Toxicity to reproduction, screening:

source substance Zinc bis(O,O-diisobutyl dithiophosphate) (CAS 68457 -79 -4): GLP, according to OECD Guideline 422, Klimisch 1, rat, oral (gavage), NOEL = 160 mg/kg bw/day

source substance Zinc bis[O,O-bis(2-ethylhexyl)] bis(dithiophosphate) (CAS 4259 -15 -8): GLP, according to OECD Guideline 421, Klimisch 1, rat, oral (gavage), NOAEL = 30 mg/kg bw/day

Based on the available data and based on the performed studies for CSA as key value the lowest value of all category members was choosen. Differences in the values of each category-member are obvious, but this is in range with the category approach, where nevertheless the members of one category may have different values, because all results show, that no substance has to be classified according to Regulation (EC) No 1272/2008.

Toxicity to reproduction, ext. one-gen repro-tox:

experimental study planned (based on read-across)

Link to relevant study records

Referenceopen allclose all

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
weight of evidence
Justification for type of information:
REPORTING FORMAT FOR THE CATEGORY APPROACH
Please refer also to the read-across statement attached in section 13

1. HYPOTHESIS FOR THE CATEGORY APPROACH (ENDPOINT LEVEL)
The target and the source substances are structurally similar substances that share the common organometallic core structure consisting of a central zinc metal bonded to four alkyldithiophosphate esters (ligands) by coordinate covalent bonds -Zn[(S2P(OR)2]2. Structural variations between the target and the source substances are related only to the alkyl (R) groups of the alkyldithiophosphate ligands. The substances in this category give thus rise to an (identical) common compound Phosphorodithioic acid moiety that can be released by the breakage of ester bonds and dissociation from the Zinc complex to which the organism would be exposed if the target substance was tested in the toxicity studies. Exposure to the parent compounds (non-transformed constituents) and to the counter alkyl alcohols, possibly released by hydrolysis of P-O bonds – non-common compounds – would not influence the prediction of the (eco)toxicological properties because they are considered to have the same biological targets and to cause the same type of effects through a common underlying mechanism due to the same functional groups (zinc cation, phosphorodithioic cation and aliphatic alcohol anionic moieties). The impurities of the target and the source substances are not expected to impact the prediction because they are identical or, if slightly structural different, belong to the same class of compounds with the same functional groups and their percentages are very low.

2. CATEGORY APPROACH JUSTIFICATION (ENDPOINT LEVEL)
Since the main constituents of the target substance are structurally similar to the constituents of the source substances with the same functional groups and the alkyl chain lengths of phosphoroditioate moieties are in the range of the established ZDDP category (C3-C12), the same mode of toxicological action is expected for the target and the source substances. The constituents of the target substance do not possess functional groups associated with other modes of action or toxicity effects. Toxicokinetic behavior of the constituents of the target substance is expected to be essentially the same as that of the source substance. Based on the results of the oral repeated dose toxicity studies with the source substances and other ZDDP category members, it is evident that the structural dissimilarities – the chain lengths of the alkyl rests – did not result in different strength of the toxicity effects. Even if absorption through GI tract of the category members with shorter alkyl rests and theoretically higher absorption rates (due to the increased water solubility and decreased molecular weights favouring absorption) is higher, their intrinsic properties are essentially the same as the findings in the toxicity studies are very similar. The impurities of the target substance are considered not to contribute to the toxicity effects because they are also structurally similar to the impurities of the source substances and consist of substances of simple structure without specific mode of action. Furthermore, their amounts are very low. Therefore, it is predicted that the target substance would possess the same toxicity potential by repeated oral exposure as the source substance.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
not specified
Description (incidence):
not specified
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
not specified
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
One male in the 160 mg/kg/day group was euthanized in extremis on study day 17; a gross observation of a thickened stomach was noted at necropsy. Clinical findings noted for this male approximately 1 hour following dose administration on the day of euthanasia consisted of yellow material on various body surfaces, clear material around the mouth, unkempt appearance, decreased defecation, and labored respiration. Microscopically, this male was noted with inflammation, edema, and ulceration in the non-glandular stomach, erosion and inflammation in the trachea, and lymphoid depletion in the thymus, spleen, and lymph nodes (mesenteric, mandibular, and axillary). The lesions in the non glandular stomach were considered test item-related and may have contributed to the moribund state of this male. All other animals in all dosage groups survived to the scheduled necropsies.
Test item related clinical findings were noted in the 160 mg/kg/day group males and females and included rales, decreased, shallow, and/or labored respiration and salivation related findings. These findings were noted at the daily examinations, at the time of dosing, and/or approximately 1 hour following dose administration primarily during the treatment period. However, because of their sporadic occurrence, these cardio pulmonary findings were considered to be incidental and secondary to the nature of the test item and the route of administration.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
Mean body weights, body weight changes, and food consumption were unaffected by test item administration in the 10, 40, and 160 mg/kg/day groups throughout the treatment and post-treatment periods.

FUNCTIONAL OBSERVATIONAL BATTERY (FOB) AND LOCOMOTOR ACTIVITY (PARENTAL ANIMALS)
No test item-related effects were noted during the FOB or locomotor activity evaluations at any dosage level.

CLINICAL PATHOLOGY (PARENTAL ANIMALS)
There were no test substance-related effects on serum chemistry, hematology, or coagulation parameters in the 10, 40, and 160 mg/kg/day groups.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
Male and female mating and fertility, male copulation and female conception indices, mean number of days between pairing and coitus, gestation length, and the process of parturition were unaffected by test item administration at all dosage levels.

ORGAN WEIGHTS (PARENTAL ANIMALS)
There were no test item-related alterations in final body weight or organ weights at any dosage level. Significant (p<0.05) differences were observed when the control and high dose group males were compared at the recovery (post-treatment) necropsy and consisted of lower mean kidney weight relative to body weight, higher mean spleen weight relative to brain weight, higher mean left testis weight relative to brain weight, and higher mean right testis weights (absolute and relative to brain weight). There was no case where all 3 measures (absolute, relative to body weight, and relative to brain weight) were statistically significant. Thus, since the absolute weights and weights relative to body or brain weight were discordant, these organ weight changes were considered to be spurious.

GROSS PATHOLOGY (PARENTAL ANIMALS)
One male (no. 61739) in the 160 mg/kg/day group was euthanized in extremis on study day 17. The thickened stomach noted macroscopically for this male was related to test item administration. There were no other test item related internal findings observed for either sex at any dosage level at the scheduled necropsies. Macroscopic findings observed in the test item groups occurred infrequently and/or in a manner that was not dose related.

HISTOPATHOLOGY (PARENTAL ANIMALS)
Test item-related histologic observations of epithelial hyperplasia, hyperkeratosis, and inflammation of the non-glandular stomach, typically at the limiting ridge, but sometimes more widespread, were noted in the 160 mg/kg/day group males and females.
Dose descriptor:
NOEL
Remarks:
portal-of-entry
Effect level:
40 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
Remarks on result:
other: Localised injury to the nonglandular portion of the stomach.
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Effect level:
160 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
Remarks on result:
other:
Remarks:
No effects observed at any dose level.
Dose descriptor:
NOEL
Remarks:
reproductive
Effect level:
160 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
reproductive performance
Remarks on result:
other: No effects observed at any dose level.
Critical effects observed:
no
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not specified
VIABILITY (OFFSPRING)
Mean numbers of corpora lutea and unaccounted-for sites, mean number of pups born, live litter size, the percentage of males at birth, and postnatal survival in the 10, 40, and 160 mg/kg/day groups were similar to the control group values.

CLINICAL SIGNS (OFFSPRING)
The general physical condition of all F1 pups in this study were unaffected by test item administration. No test item-related clinical findings were noted for the F1 pups.

BODY WEIGHT (OFFSPRING)
Mean male and female pup body weights and body weight gains in the 10, 40, and 160 mg/kg/day groups were unaffected by test item administration during PND 1-4. No statistically significant differences from the control group were noted.

GROSS PATHOLOGY (OFFSPRING)
There were no remarkable macroscopic findings in the F1 pups at the scheduled necropsy at any dosage level.
Dose descriptor:
NOEL
Remarks:
neonatal toxicity
Generation:
F1
Effect level:
160 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
viability
Remarks on result:
other: No effects observed at any dose level.
Critical effects observed:
no
Reproductive effects observed:
no
Conclusions:
Under the conditions of this screening study, no test material-related effects on reproductive performance, gestation length, parturition, reproductive organs, or neurobehavioral parameters were noted at any dosage level. Based on these results, a dosage level of 160 mg/kg/day was considered to be the no-observed-effect level (NOEL) for reproductive toxicity. Test item-related moribundity, clinical findings, and microscopic findings in the non glandular portion of the stomach, characterized by epithelial hyperplasia, hyperkeratosis, and inflammation, were observed in the 160 mg/kg/day group. The injury to the nonglandular portion of the stomach was localized and considered to be irritation from test item portal-of-entry effects. Based on these results, the NOEL for portal-of-entry effects was considered to be 40 mg/kg/day, and excluding the histologic injury to the nonglandular stomach, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day.
Executive summary:

In a guideline repeated dose and reproduction / developmental screening study (OECD 422) conducted according to Good Laboratory Practices, WIL Research Labs (2010) evaluated the potential toxic effects ofphosphorodithioic acid, mixed O,O-bis(iso-Bu and pentyl) esters, zinc salts when administered to rats. This study was designed to evaluate the toxic effects, including neurobehavioral effects, of the test material to parental animals and to evaluate the potential to effect male and female reproductive performance such as gonadal function, mating behavior, conception, parturition, and early postnatal development. The test material was administered orally by gavage once daily to 3 groups of Crl:CD(SD) rats at levels of 10, 40 or 160 mg/kg/day. The low- and mid-dose groups each consisted of 10 rats/sex and the high-dose group consisted of 15 rats/sex. A concurrent control group of 15 rats/sex received the vehicle, mineral oil USP, on a comparable regimen. Ten males/group selected for pairing were dosed for 14 days prior to mating through 1 day prior to euthanasia for a total of 28 doses. Ten females/group selected for pairing were dosed for 14 days prior to mating through lactation day 3 for a total of 40-52 doses; females that failed to deliver were dosed through the day prior to euthanasia (post-mating day 25) for a total of 40 doses. The extra 5 males and 5 females in the control and high-dose groups were not used for mating and were treated beginning on study day 0; following 28 doses for the males and 40 doses for the females, these animals were assigned to the post‑treatment period and remained on study for a 14-day non-dosing period.

Under the conditions of this screening study, no test material-related effects on reproductive performance, gestation length, parturition, reproductive organs, or neurobehavioral parameters were noted at any dosage level. Based on these results, a dosage level of 160 mg/kg/day was considered to be the no-observed-effect level (NOEL) for reproductive toxicity. Test item-related moribundity, clinical findings, and microscopic findings in the non glandular portion of the stomach, characterized by epithelial hyperplasia, hyperkeratosis, and inflammation, were observed in the 160 mg/kg/day group. The injury to the nonglandular portion of the stomach was localized and considered to be irritation from test item portal-of-entry effects. Based on these results, the NOEL for portal-of-entry effects was considered to be 40 mg/kg/day, and excluding the histologic injury to the nonglandular stomach, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day.

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
weight of evidence
Justification for type of information:
REPORTING FORMAT FOR THE CATEGORY APPROACH
Please refer also to the read-across statement attached in section 13

1. HYPOTHESIS FOR THE CATEGORY APPROACH (ENDPOINT LEVEL)
The target and the source substances are structurally similar substances that share the common organometallic core structure consisting of a central zinc metal bonded to four alkyldithiophosphate esters (ligands) by coordinate covalent bonds -Zn[(S2P(OR)2]2. Structural variations between the target and the source substances are related only to the alkyl (R) groups of the alkyldithiophosphate ligands. The substances in this category give thus rise to an (identical) common compound Phosphorodithioic acid moiety that can be released by the breakage of ester bonds and dissociation from the Zinc complex to which the organism would be exposed if the target substance was tested in the toxicity studies. Exposure to the parent compounds (non-transformed constituents) and to the counter alkyl alcohols, possibly released by hydrolysis of P-O bonds – non-common compounds – would not influence the prediction of the (eco)toxicological properties because they are considered to have the same biological targets and to cause the same type of effects through a common underlying mechanism due to the same functional groups (zinc cation, phosphorodithioic cation and aliphatic alcohol anionic moieties). The impurities of the target and the source substances are not expected to impact the prediction because they are identical or, if slightly structural different, belong to the same class of compounds with the same functional groups and their percentages are very low.

2. CATEGORY APPROACH JUSTIFICATION (ENDPOINT LEVEL)
Since the main constituents of the target substance are structurally similar to the constituents of the source substances with the same functional groups and the alkyl chain lengths of phosphoroditioate moieties are in the range of the established ZDDP category (C3-C12), the same mode of toxicological action is expected for the target and the source substances. The constituents of the target substance do not possess functional groups associated with other modes of action or toxicity effects. Toxicokinetic behavior of the constituents of the target substance is expected to be essentially the same as that of the source substance. Based on the results of the oral repeated dose toxicity studies with the source substances and other ZDDP category members, it is evident that the structural dissimilarities – the chain lengths of the alkyl rests – did not result in different strength of the toxicity effects. Even if absorption through GI tract of the category members with shorter alkyl rests and theoretically higher absorption rates (due to the increased water solubility and decreased molecular weights favouring absorption) is higher, their intrinsic properties are essentially the same as the findings in the toxicity studies are very similar. The impurities of the target substance are considered not to contribute to the toxicity effects because they are also structurally similar to the impurities of the source substances and consist of substances of simple structure without specific mode of action. Furthermore, their amounts are very low. Therefore, it is predicted that the target substance would possess the same toxicity potential by repeated oral exposure as the source substance.
Clinical signs:
effects observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
not examined
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
effects observed, treatment-related
-CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
Five animals in the 200 mg/kg/day group died prior to the scheduled necropsies. Male nos. 22066 and 22084 in this group died on study days 12 and 19, respectively. Female nos. 22112, 22127 and 22115 in this group died on study days 8, 27 and 39, respectively. Clinical signs noted for at least two of these animals at the daily examinations, at the time of dosing and/or I-hour post dosing on the day of death or the day prior to death were staining, matting or material (brown, tan, yellow, red and/or clear) on various body surfaces, respiratory distress (rales, gasping, labored respiration), hunched appearance and yellow mucoid diarrhea. Single occurrences of unkempt appearance, body cool to touch and aggressiveness were also noted in these animals prior to death. Two females in the 100 mg/kg/day group (nos. 22142 and 22147) and one female in the 200 mg/kg/day group (no. 22130) were euthanized on lactation days 1 or 2 due to total litter loss. Clinical signs observed in these females prior to euthanization were similar to those noted in the animals that died. These findings included staining, matting or material (brown, red, tan and/or clear) on various body surfaces, respiratory distress (rales, gasping) and yellow mucoid diarrhea. In addition, female no. 22147 in the 100 mg/kg/day group exhibited signs of dystocia on lactation day 0, and female no. 22130 in the 200 mg/kg/day group exhibited abnormal nesting behavior on lactation day 1.
All other animals survived to the scheduled necropsies. The predominant clinical signs observed in these animals consisted of the following. Salivation was noted frequently in the 100 and 200 mg/kg/day group males and females at the time of dosing and/or one hour following dosing. Brown staining on various body surfaces was observed frequently one hour following dosing in males and females at dose levels of 100 and 200 mg/kg/day. Other clinical signs apparently related to ZDDP administration were intermittent occurrences of respiratory distress (rales, gasping) and excreta-related findings (mucoid feces, mucoid diarhea, diarhea and/or soft stool) in the 100 and 200 mg/kg/day group males and females. No clinical signs that could be related to compound administration were observed in the 30 mg/kg/day group males and females.

-BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
a. WEEKLY
Mean weekly body weights in the 200 mg/kg/day group males were comparable to the control group values at weeks 0-4. None of the differences were statistically significant. Mean body weight gains for the 200 mg/kg/day group males during weeks 0-1 and 1-2 were slightly reduced, though not statistically significant, when compared to the control group values. A significantly reduced (p<0.01) mean body weight gain for the 200 mg/kg/day group males occurred during weeks 2-3. Mean body weight gain for the high dose group males continued to be reduced (not significant) during weeks 3-4 in comparison to the control group values. Mean cumulative body weight gain for the 200 mg/kg/day group males from weeks 0 to 4 was significantly reduced (p<0.05) in comparison to the control group value.
Mean weekly body weights, body weight gains and cumulative body weight gains for the 200 mg/kg/day group females prior to breeding (weeks 0-2) were similar to the control group values. None of the differences were statistically significant.
At dose levels of 30 and 100 mg/kg/day, mean weekly body weights for the males were comparable to the control group values at weeks 0, 1, 2, 3 and 4. Mean body weight gains for the 30 and 100 mg/kg/day group males during weeks 0-1, 1-2 and 2-3 were comparable to the control group values. During weeks 3-4, the 100 mg/kg/day group males had a significantly reduced @< 0.05) mean body weight gain. This reduction was due to male no. 22062 which lost 14 g of body weight during weeks 3-4. A similar decrease was not observed in the 200 mg/kg/day group males and no relationship to treatment was evident. Mean body weight gain for the 30 mg/kg/day group males during weeks 3-4 was comparable to the control group value.
Mean weekly body weights for the 30 and 100 mg/kg/day group females were increased prior to breeding (weeks 0, 1 and 2) in comparison to the control group values. The difference between the control group and the 100 mg/kg/day group females at week 2 was statistically significant (p<0.05). Mean cumulative body weight gain for the 100 mg/kg/day group females was significantly increased (p <0.05) during weeks 0 to 2 when compared to the control group value; this increase was not considered to be related to treatment. Mean body weight gains for the 30 -and 100 mg/kg/day group females prior to breeding (weeks 0-1 and 1-2) were similar to the control group values. None of the differences were statistically significant.
b. GESTATION
Mean gestation body weights and body weight gains in the 30, 100 and 200 mg/kg/day groups were similar to or slightly greater than the control group values. No relationship to treatment was apparent.
c. LACTATION
Mean body weights in the 30, 100 and 200 mg/kg/day groups were similar to the control group values on lactation days 1 and 4. None of the differences were statistically significant. Mean body weight gains in these groups were slightly reduced, though not statistically significant, when compared to the control group value. No relationship to treatment was evident.

-TEST SUBSTANCE INTAKE (PARENTAL ANIMALS)
The test material was administrated via oral gavage.

-REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
Estrous cycle regularity was not examined.

- REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
No data available.

-REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
Reproductive performance was not adversely affected by compound administration at dose levels of 30, 100 and 200 mg/kg/day. Male mating indices were. 91.7%, 91.7%, 91.7% and 81.8% and female mating indices were 100%, 91.7%, 100% and 90.9% for the control, 30, 100 and 200 mg/kg/day groups, respectively. Males which did not sire a litter numbered 1, 2, 2 and 4 in the same respective dose groups. In these same dose groups, 0, 1, 1 and 2 females, respectively, had evidence of mating but did not deliver; all females which had evidence of mating and did not deliver were determined to be nongravid at the post mortem examination, with the following exceptions. Female no. 22115 had evidence of mating but died prior to the expected parturition; this female was determined to be gravid at the post mortem examination. Female no. 22127 in the 200 mg/kg/day group died on gestation day 5, too early for pregnancy confirmation, and was handled as nongravid for statistical purposes. Female no. 22096 in the 30 mg/kg/day group had no evidence of mating and was nongravid.
Fertility indices for males were 91.7%, 83.356, 83.3% and 81.8% and for females were 100%, 83.3%, 91.7% and 81.8% in the control, 30, 100 and 200 mg/kg/day groups, respectively. Although the male and female fertility indices in the 200 mg/kg/day group (81.8%) were slightly lower than the control group values (91.7% and 100.0%, respectively), they were within the range of values in the WIL reproductive historical control data (66.7%-100.0% and 64.0%- 100.0%,respectively). These slight numerical differences were not statistically significant and represented only 1 or 2 fewer successful matings out of a total of 11 or 12 males and females used for mating in the control and high dose groups. Thus, these slightly reduced fertility indices for males and females in the 200 mg/kg/day group were not clearly a treatment related effect.
The mean numbers of days between pairing and coitus in the treated groups were similar to the control group values.

-ORGAN WEIGHTS (PARENTAL ANIMALS)
No adverse effects were apparent on brain, liver, kidney, testes, ovary or pituitary weights in the treated group males and females when evaluated on an absolute basis and relative to final body weight. None of the differences between the treated groups and control group values were significant.

- GROSS PATHOLOGY (PARENTAL ANIMALS)
1. F0 UNSCHEDULED DEATHS
Two males (nos. 22066 and 22084) in the 200 mg/kg/day group died on study days 12 and 19, respectively. Male no. 22066 had a distended stomach. Male no. 22084 was internally normal. Three females (nos. 22112, 22127 and 22115) in the 200 mg/kg/day group died on study days 8, 27 and 39, respectively. Female no. 22112 had a reddened pituitary and yellow contents in the intestinal tract. Female no. 22127 was nongravid (died gestation day 5) and internally normal. Female no. 22115 died on gestation day 21 and had two late resorptions (mummified) and 14 normally developing implantations in utero. A distended and gas filled cecum and stomach and dark red contents or areas in the jejunum and stomach were also noted for this female at the necropsy examination.

2. F0 FEMALE SPOST-MATING DAY 25
One female in each of the 30 and 100 mg/kg/day groups (nos. 22105 and 22113, respectively) had evidence of mating but did not deliver and were necropsied on gestation day 25. Both females were nongravid. Female no. 22105 had a hemorrhagic thymus gland and female no. 22113 was internally normal.

3. F0 FEMALES25 DAYS FOLLOWING THE BREEDING PWOD
Female nos. 22096 and 22118 in the 30 and 200 mg/kg/day groups, respectively, had no evidence of mating and were necropsied 25 days following the conclusion of the breeding period. These females were nongravid and internally normal.

4. F0 FEMALES WITH TOTAL LITTER LOSS
Two females in the 100 mg/kg/day pup (nos. 22142 and 22147) and one female in the 200 mg/kg/day pup (no. 22130) were euthanized in extremis due to total litter loss and were necropsied within 24 hours. Female no. 22142 was internally normal and female no. 22147 had a distended, gas filled and thickened cecum. Female no. 22130 had dark red contents in the stomach.

5. F0 FEMALES LACTATION DAY 4
At the scheduled necropsy on lactation day 4, all females in the control, 30, 100 and 200 mg/kg/day groups were internally normal.
The calculated differences between the number of pups born and the number of implantation sites counted at the scheduled necropsy in the treated groups were comparable to the control group values. None of the differences were statistically significant.

6. F0 MALE SCHEDULED NECROPSY
At the scheduled necropsy of F, males, no compound-related internal findings were observed at any dose level. Male no. 22067 in the 30 mg/kg/day group had dark red lungs and red foamy contents in the trachea. AU other animals were internally normal.

-HISTOPATHOLOGY (PARENTAL ANIMALS)
No microscopic lesions attributed to ZDDP administration were observed in any tissues upon histopathological examination. The frequency of lesions observed in the treated group males and females was similar to that observed for the control group or the findings were noted in single animals. Microscopic lesions observed in the females which had total litter loss, such as hyperplasia and hypertrophy of the uterus vessels and/or cytoplasmic vacuolation of the vagina, were not considered to be related to treatment.
Microscopic examination of gross lesions noted at the scheduled necropsy did not reveal any effects of ZDDP administration.
Dose descriptor:
NOAEL
Effect level:
30 mg/kg bw/day
Sex:
male/female
Basis for effect level:
clinical signs
mortality
Clinical signs:
effects observed, treatment-related
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
effects observed, treatment-related
Histopathological findings:
not examined
VIABILITY (OFFSPRING)
Viability indices on lactation days 1 and 4 remained above 99% in the control and 30 mg/kg/day groups. Viability indices for the 100 mg/kg/day group at lactation days 1 and 4 were decreased (85.9% and 84.4%, respectively), when compared to the values observed in the WIL reproductive historical control data (96.9% and 96.0%, respectively). Both of these values were significantly decreased (p<0.01) when compared to the control group values. Viability indices at lactation days 1 and 4 for the 200 mg/kg/day group were 98.2% and 86.2%, respectively. The lactation day 4 value was significantly lower (p<0.01) than the control group value. The decreased viability indices in the 100 and 200 mg/kg/day groups were due primarily to the females in these groups which had total litter loss.

CLINICAL SIGNS (OFFSPRING)
On lactation day 0 and post-natal days 1-4, pups were found dead in the control, 30, 100 and 200 mg/kg/day groups. Two pups in each of the 100 and 200 mg/kg/day groups were missing and presumed cannibalized. Total litter loss was noted for female nos. 22142 (12 pups) and 22147 (15 pups) in the 100 mg/kg/day group and female no. 22130 (14 pups) in the 200 mg/kg/day group.

BODY WEIGHT (OFFSPRING):
No adverse effects on mean pup weights were apparent on lactation days 1 and 4 at dose levels of 30, 100 and 200 mg/kg/day. None of the differences between the treated groups and control group values were statistically significant.

SEXUAL MATURATION (OFFSPRING)
No data available.

ORGAN WEIGHTS (OFFSPRING)
No data available.

GROSS PATHOLOGY (OFFSPRING)
An increased incidence of pups with no milk in the stomach was noted in the 100 mg/kg/day group. The only other remarkable necropsy finding for pups found dead during the post-natal period was a heart and great vessel anomaly for pup no. 22097-01 in the 200 mg/kg/day group. This malformation consisted of the pulmonary arteries arising from the ascending aorta, the brachiocephalic trunk and left carotid arising from the pulmonary trunk, an absent ductus arteriosis and a small opening in the anterior portion of the septum. Thirteen pups in the 200 mg/kg/day group were noted with body cool to touch. Other observations on the general physical condition of the F,pups in the treated groups occurred at a low incidence, similarly in the control group and/or in a non dose-related manner.

At the post-natal day 4 scheduled necropsy, internal findings (with the exception of presence of milk in the stomach) were noted for 1, 1 and 3 pups in the 30, 100 and 200 mg/kg/day groups, respectively. Pup no. 22106-04 in the 30 mg/kg/day group had unilateral microphthalmia. Pup no. 22148-08 in the 100 mg/kg/day group had a major blood vessel variation, consisting of the right carotid and right subclavian arising independently from the aortic arch. Pup no. 22123-12 in the high dose group had situs inversus. A filamentous tail was noted for pup no. 22146-16 in the 200 mg/kg/day group. The spleen was attached to the left kidney and multiple white foci on the pancreas were noted for pup no. 22097-06 in the 200 mg/kg/day group. No other remarkable internal findings were noted at any dose level at the scheduled necropsy.

HISTOPATHOLOGY (OFFSPRING)
No data available.
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
30 mg/kg bw/day
Sex:
male/female
Basis for effect level:
mortality
Reproductive effects observed:
not specified

F0 (Parental Generation)

Two males and three females in the high dose group (200 mg/kg/day) died prior to scheduled necropsy (test days 12, 19, 8, 27 and 39). These deaths were considered treatment related. Two females in the mid dose group (100 mg/kg/day) and one female in the high dose group were euthanized on lactation days 1 or 2 due to total litter loss. Five of these animals exhibited gastric irritation upon necropsy. All other animals survived to their scheduled sacrifice.

Clinical signs noted in the found dead or sacrificed animals included staining, matting of fur, respiratory distress, hunched appearance and mucoid diarrhea. Clinical findings noted for the surviving mid and high dose males and females included post dosing salivation, brown staining, respiratory distress and diarrhea. No treatment related clinical findings were observed in the low dose (30 mg/kg/day) animals.

 

Fertility indices (%) for the high dose males and females were slightly lower then control as follows:

                 Control            30 mg/kg       100 mg/kg         300 mg/kg

Males           91.7                  83.3                83.3                   81.8

Females      100                    83.3                91.7                   81.8

These values were within the range of the test facility historical control data (64-100%). In addition, differences from control were not statistically significant and represent only 1 or 2 fewer successful matings out of 11 or 12 males or females used for mating in the control and high dose groups. A microscopic examination of the reproductive organs of these animals did not reveal any treatment-related effects. The Study Director concluded that the low and mid dose groups were unaffected and that these data did not clearly reflect a treatment related effect in the high dose group. Other reproductive parameters (mating indices, days between pairing and coitus, gestation length and parturition) were unremarkable in all treated groups.

 

The premating (weeks 1-4) mean body weight gain of the high dose males was statistically significantly reduced compared to control. The mean body weights of the low and mid dose males and all treated female groups were unremarkable during the premating period. Gestation and lactation body weights were unremarkable in all treated groups. Food consumption data were unremarkable in all treated groups (males and females) during the premating, gestation and lactation periods. With the exception of the gastric irritation noted above in several unscheduled deaths, the macroscopic data were unremarkable. Absolute and relative (to body weight) organ weight data as well as the microscopic examination data of the F0 males and females were unremarkable. There were no treatment-related findings evident in any of these data.

 

 

F1 Litter Data

Pup body weights, live litter size and sex ratios were unremarkable. No treatment related effects were evident. An increased number of dead pups was noted in the mid dose group on day 0 of lactation. Pup viability indices in the mid dose (lactation days 1 and 4) and high dose (lactation day 4) groups were reduced. This was attributed to total litter loss by three females. Increased pup deaths were observed in the mid and high dose groups during the post-natal period. An increased incidences of pups without milk in the stomach was noted in the mid dose group. No treatment related effects were evident in the necropsy data of these found dead pups or in the necropsy data from scheduled pup necropsies.

 

Chemical analysis of dosing suspensions confirmed that they were homogeneous and of appropriate concentration throughout the study.

Conclusions:
Parental (F0) toxicity was exhibited at dose levels of 100 (mortality, clinical findings) and 200 mg/kg/day (mortality, clinical signs, reduced body weight gain, gastric irritation). A slightly reduced fertility index was also observed at 200 mg/kg/day. No F0 toxicity was observed at 30 mg/kg. Neonatal (F1) toxicity (mortality) was observed at 100 and 200 mg/kg/day. No F1 toxicity was observed at 30 mg/kg. Based on these findings the Study Director concluded that the NOAEL for both parental and neonatal toxicity was 30 mg/kg/day.
Executive summary:

This screening study was designed to determine the potential adverse effects of Zinc O,O-diethylhexyldithiophosphate (ZDDP) on male and female reproduction in rats. The test material was administered orally by gavage to three groups of 12 F0 male and 12 F0 female Sprague Dawiey Crl:CD®BR rats. Dosage levels were 30, 100 and 200 mg/kg/day. For comparative purposes, a control pup of identical design was concurrently dosed with Mazola®corn oil on a comparable regimen. A dose volume of 5 ml/kg was used in all dose groups. All F0 animals were dosed for a minimum of 14 days prior to mating and through the day of necropsy for each F, animal. All animals were observed twice daily for appearance and behavior. Body weights were recorded weekly for both sexes prior to mating; maternal body weights were also recorded on gestation days 0, 7, 14 and 20 as well as lactation days 1 and 4. Food consumption was measured for corresponding intervals prior to mating, during gestation and during lactation. All of the surviving F0 females were allowed to deliver and rear their pups to lactation day 4. The offspring were also potentially exposed in utero and through nursing during lactation days 1-4 until euthanization on post-natal day 4. The surviving F, dams were necropsied on lactation day 4, following at least 43 days of dosing. The surviving F0 males were necropsied after the breeding period, following 28 days of dosing. F0 females with total litter loss were necropsied within 24 hours. F0 females which failed to deliver were necropsied on post-mating day 25 (evidence of mating) or 25 days following the breeding period (no evidence of mating). Organ weights were collected (all F0 treated groups) and microscopic examinations were conducted (control and high dose groups and all parental animals not surviving to the scheduled necropsies).

Slightly reduced fertility indices for males and females in the 200 mg/kg/day group (81.8% compared to control values of 91.7% and 100%, respectively) were not clearly related to treatment. Fertility indices in the 30 and 100 mg/kg/day groups were unaffected by treatment. Other reproductive parameters (mating, days between pairing and coitus, gestation and parturition) were unaffected by treatment at all dose Levels.

Two males and three females in the 200 mg/kg/day group died prior to the scheduled necropsies. Two and one females in the 100 and 200 mg/kg/day groups, respectively, were euthanized due to total litter loss prior to the scheduled necropsy. Five of these animals had internal changes consistent with gastric irritation. All other animals survived to the scheduled necropsies. The predominant clinical findings noted for surviving males and females in the 100 and 200 mg/kg/day groups at the time of dosing and/or 1-hour post dosing were salivation and brown staining on various body surfaces. Other treatment-related clinical signs observed for the 100 and 200 mg/kg/day group males and females were respiratory distress (rales, gasping) and excreta-related findings (mucoid feces, mucoid diarrhea, diarrhea and/or soft stool). No clinical signs related to ZDDP administration were observed in the 30 mg/kg/day group males and females.

Reduced mean body weight gains occurred in the 200 mg/kg/day group males during weeks 0-1, 1-2, 2-3 and 3-4. Weekly body weight gains in the 30 and 100 mg/kg/day group males and the 30, 100 and 200 mg/kg/day group females were not adversely affected by treatment. Gestation and lactation body weight gains in the 30, 100 and 200 mg/kg/day group females were not adversely affected by treatment. Food consumption in the 30, 100 and 200 mg/kg/day group males and females was unaffected by treatment.

No compound-related internal findings were noted in the surviving animals in the treated groups at the necropsy and microscopic examinations. Microscopic examination of gross lesions noted at the scheduled necropsies did not reveal any adverse effects of ZDDP administration. No adverse effects were apparent on brain, liver, kidney, testes, ovary or pituitary weights in the treated F,, males and females.

An increased number of dead pups on lactation day 0 was noted in the 100 mg/kg/day group. Pup viability indices for the 100 mg/kg/day group (lactation days 1 and 4) and the 200 mg/kg/day group (lactation day 4) were reduced (due to the three females with total litter loss). Increased numbers of pups in the 100 and 200 mg/kg/day groups were found dead during the post-natal period. The predominant clinical observation noted for the 200 mg/kg/day pups was body cool to touch. F, pup sex ratios were not adversely affected by compound administration at any dose level. Necropsy findings did not reveal a cause of death for the F, pups that died or indicate a relationship to treatment for the F, pups that were euthanized on post-natal day 4.

 

In conclusion, parental toxicity was exhibited at dose levels of 100 and 200 mg/kg/day by mortality and clinical signs. Parental toxicity was also exhibited at the 200 mg/kg/day dose level by inhibition of body weight gain in males and signs of gastric irritation. No parental toxicity was observed at the 30 mg/kg/day dose level. Slightly reduced fertility indices were observed at the 200 mg/kg/day dose level. Reproductive performance (fertility, mating, days between pairing and coitus, gestation and parturition) was unaffected by treatment at the 30 and 100 mg/kg/day dose levels. Neonatal toxicity (mortality) in the F1 generation was observed at the 100 and 200 mg/kg/day dose levels. Neonatal toxicity was also noted at the 200 mg/kg/day dose level by clinical signs. No neonatal toxicity was observed at a dose level of 30 mg/kg/day. Based on the results of this study, a dose level of 30 mg/kg/day was considered to be the NOAEL (no observable adverse effect level) for parental and neonatal toxicity.

Endpoint:
extended one-generation reproductive toxicity - basic test design (Cohorts 1A, and 1B without extension)
Type of information:
experimental study planned (based on read-across)
Remarks:
testing proposed with Phosphorodithioic acid, mixed O,O-bis(iso-Bu and pentyl) esters, zinc salts (EC 270-608-0)
Study period:
Investigation to begin as soon as possible after completion of prenatal development toxicity study
Justification for type of information:
The extended one-generation reproductive toxicity study according to OECD guideline 443 proposed for the source substance CAS 68457-79-4 is used as a source for read-across for the target substance (CAS 10147-77-7).

REPORTING FORMAT FOR THE CATEGORY APPROACH
Please refer also to the read-across statement attached in section 13

1. HYPOTHESIS FOR THE CATEGORY APPROACH (ENDPOINT LEVEL)
The target and the source substances are structurally similar substances that share the common organometallic core structure consisting of a central zinc metal bonded to four alkyldithiophosphate esters (ligands) by coordinate covalent bonds -Zn[(S2P(OR)2]2. Structural variations between the target and the source substances are related only to the alkyl (R) groups of the alkyldithiophosphate ligands. The substances in this category give thus rise to an (identical) common compound Phosphorodithioic acid moiety that can be released by the breakage of ester bonds and dissociation from the Zinc complex to which the organism would be exposed if the target substance was tested in the toxicity studies. Exposure to the parent compounds (non-transformed constituents) and to the counter alkyl alcohols, possibly released by hydrolysis of P-O bonds – non-common compounds – would not influence the prediction of the toxicological properties because they are considered to have the same biological targets and to cause the same type of effects through a common underlying mechanism due to the same functional groups (zinc cation, phosphorodithioic cation and aliphatic alcohol anionic moieties). The impurities of the target and the source substances are not expected to impact the prediction, because they are identical or, if slightly structural different, belong to the same class of compounds with the same functional groups and their percentages are very low.

2. CATEGORY APPROACH JUSTIFICATION (ENDPOINT LEVEL)
The results of the two source substances indicate that local irritation of the gastrointestinal tract is the primary effect of the substances in the ZDDP category. This is also in line with the skin and eye irritating properties of the substances.
Since the main constituents of the target substance are structurally similar to the constituents of the source substances with the same functional groups and the alkyl chain lengths of phosphorodithioate moieties are in the range of the established ZDDP category (C3-C12), the same mode of toxicological action is expected for the target and the source substances. The constituents of the target substance do not possess functional groups associated with other modes of action or toxicity effects. The toxicokinetic behaviour of the constituents of the target substance is expected to be essentially the same as that of the source substance. Based on the results of the oral repeated dose toxicity studies with the source substances and other ZDDP category members, it is evident that the structural dissimilarities – the chain lengths of the alkyl rests – did not result in different strength of the toxicity effects. Even if absorption through GI tract of the category members with shorter alkyl rests is higher (due to the increased water solubility and decreased molecular weights favouring absorption), their intrinsic properties are essentially the same as the findings in the toxicity studies are very similar. Therefore, the extended one-generation reproductive toxicity study proposed for the source substance will generate adequate data to fulfil the data requirements for the target substance. The impurities of the target substance are considered not to contribute to the toxicity effects because they are also structurally similar to the impurities of the source substances and consist of substances of simple structure without specific mode of action. Furthermore, their amounts are very low. Therefore, it is predicted that the target substance (CAS 10147-77-7) would possess the same reproductive and developmental toxicity potential by repeated oral exposure as the source substance (CAS 68457-79-4).

Qualifier:
according to
Guideline:
OECD Guideline 443 (Extended One-Generation Reproductive Toxicity Study)
Justification for study design:
It is proposed to use an integrated testing strategy (ITS) and to perform the tests in sequence. Data obtained will inform the protocols of the subsequent tests and the testing plan will be modified as necessary.
Species:
rat
Sex:
male/female
Route of administration:
oral: feed
Details on exposure:
Oral administration of the test item is relevant because physicochemical properties mean systemic uptake via the oral route is more likely than via the dermal or inhalation route. However, absorption is expected to be slow/inefficient via the gastrointestinal tract and previous studies have demonstrated local irritant properties at the site of entry. It is therefore proposed that administration of the substance to animals via dietary admixture is the most likely to achieve maximum exposure without causing undue pain and suffering
Details on study design:
The basic protocol design of the OECD 443 (cohorts 1A and 1B, without extension to the second generation) will be performed. In addition, the ECHA specific modifications of the OECD 443 will be incorporated into the protocol design so that the premating dosing period will be for 10 weeks and the dose levels will be set according to general toxicity.

Physicochemical properties mean that category substances are predicted to be absorbed very slowly via the dermal route and no significant uptake is expected. With respect to the inhalation route, category substances have a vapour pressure of < 0.01 Pa at 25 °C and, under normal conditions of handling and use, absorption of test material in the form of vapours, gases or mists via the respiratory tract is not expected to be relevant or significant.
Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
30 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
High due to guideline studies.
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

Toxicity to reproduction, screening:

source substance Zinc bis(O,O-diisobutyl dithiophosphate) (CAS 68457 -79 -4)

In a guideline repeated dose and reproduction / developmental screening study (OECD 422) conducted according to GLP, the potential toxic effects of phosphorodithioic acid, mixed O,O-bis(iso-Bu and pentyl) esters, zinc salts in rats were evaluated. This study was designed to evaluate the toxic effects, including neurobehavioral effects, of the test material to parental animals and to evaluate the potential to effect male and female reproductive performance such as gonadal function, mating behavior, conception, parturition, and early postnatal development. The test material was administered orally by gavage once daily to 3 groups of Crl:CD(SD) rats at levels of 10, 40 or 160 mg/kg/day. The low- and mid-dose groups each consisted of 10 rats/sex and the high-dose group consisted of 15 rats/sex. A concurrent control group of 15 rats/sex received the vehicle, mineral oil USP, on a comparable regimen. Ten males/group selected for pairing were dosed for 14 days prior to mating through 1 day prior to euthanasia for a total of 28 doses. Ten females/group selected for pairing were dosed for 14 days prior to mating through lactation day 3 for a total of 40-52 doses; females that failed to deliver were dosed through the day prior to euthanasia (post-mating day 25) for a total of 40 doses. The extra 5 males and 5 females in the control and high-dose groups were not used for mating and were treated beginning on study day 0; following 28 doses for the males and 40 doses for the females, these animals were assigned to the post‑treatment period and remained on study for a 14-day non-dosing period.

Under the conditions of this screening study, no test material-related effects on reproductive performance, gestation length, parturition, reproductive organs, or neurobehavioral parameters were noted at any dosage level. Based on these results, a dosage level of 160 mg/kg/day was considered to be the no-observed-effect level (NOEL) for reproductive toxicity. Test item-related moribundity, clinical findings, and microscopic findings in the non glandular portion of the stomach, characterized by epithelial hyperplasia, hyperkeratosis, and inflammation, were observed in the 160 mg/kg/day group. The injury to the nonglandular portion of the stomach was localized and considered to be irritation from test item portal-of-entry effects. Based on these results, the NOEL for portal-of-entry effects was considered to be 40 mg/kg/day, and excluding the histologic injury to the nonglandular stomach, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day.

source substance Zinc bis[O,O-bis(2-ethylhexyl)] bis(dithiophosphate) (CAS 4259 -15 -8)

This screening study according to GLP and OECD guideline 421 was designed to determine the potential adverse effects of Zinc O,O-diethylhexyldithiophosphate (ZDDP) on male and female reproduction in rats. The test material was administered orally by gavage to three groups of 12 F0 male and 12 F0 female Sprague Dawiey Crl:CD®BR rats. Dosage levels were 30, 100 and 200 mg/kg/day. For comparative purposes, a control pup of identical design was concurrently dosed with Mazola®corn oil on a comparable regimen. A dose volume of 5 ml/kg was used in all dose groups. All F0 animals were dosed for a minimum of 14 days prior to mating and through the day of necropsy for each F, animal. All animals were observed twice daily for appearance and behavior. Body weights were recorded weekly for both sexes prior to mating; maternal body weights were also recorded on gestation days 0, 7, 14 and 20 as well as lactation days 1 and 4. Food consumption was measured for corresponding intervals prior to mating, during gestation and during lactation. All of the surviving F0 females were allowed to deliver and rear their pups to lactation day 4. The offspring were also potentially exposed in utero and through nursing during lactation days 1-4 until euthanization on post-natal day 4. The surviving F, dams were necropsied on lactation day 4, following at least 43 days of dosing. The surviving F0 males were necropsied after the breeding period, following 28 days of dosing. F0 females with total litter loss were necropsied within 24 hours. F0 females which failed to deliver were necropsied on post-mating day 25 (evidence of mating) or 25 days following the breeding period (no evidence of mating). Organ weights were collected (all F0 treated groups) and microscopic examinations were conducted (control and high dose groups and all parental animals not surviving to the scheduled necropsies).

In conclusion, parental toxicity was exhibited at dose levels of 100 and 200 mg/kg/day by mortality and clinical signs. Parental toxicity was also exhibited at the 200 mg/kg/day dose level by inhibition of body weight gain in males and signs of gastric irritation. No parental toxicity was observed at the 30 mg/kg/day dose level. Slightly reduced fertility indices were observed at the 200 mg/kg/day dose level. Reproductive performance (fertility, mating, days between pairing and coitus, gestation and parturition) was unaffected by treatment at the 30 and 100 mg/kg/day dose levels. Neonatal toxicity (mortality) in the F1 generation was observed at the 100 and 200 mg/kg/day dose levels. Neonatal toxicity was also noted at the 200 mg/kg/day dose level by clinical signs. No neonatal toxicity was observed at a dose level of 30 mg/kg/day. Based on the results of this study, a dose level of 30 mg/kg/day was considered to be the NOAEL (no observable adverse effect level) for parental and neonatal toxicity.

Effects on developmental toxicity

Description of key information

Developmental toxicity / teratogenicity, screening:

source substance Zinc bis(O,O-diisobutyl dithiophosphate) (CAS 68457 -79 -4): GLP, according to OECD Guideline 422, Klimisch 1, rat, oral (gavage), NOAEL = 160 mg/kg bw/day

source substance Zinc bis[O,O-bis(2-ethylhexyl)] bis(dithiophosphate) (CAS 4259 -15 -8): GLP, according to OECD Guideline 421, Klimisch 1, rat, oral (gavage), NOAEL = 30 mg/kg bw/day

Based on the available data and based on the performed studies for CSA as key value the lowest value of all category members was choosen. Differences in the values of each category-member are obvious, but this is in range with the category approach, where nevertheless the members of one category may have different values, because all results show, that no substance has to be classified according to Regulation (EC) No 1272/2008.

Toxicity to reproduction, ext. one-gen repro-tox:

experimental study planned (based on read-across)

Link to relevant study records

Referenceopen allclose all

Endpoint:
developmental toxicity
Remarks:
screening
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
weight of evidence
Justification for type of information:
REPORTING FORMAT FOR THE CATEGORY APPROACH
Please refer also to the read-across statement attached in section 13

1. HYPOTHESIS FOR THE CATEGORY APPROACH (ENDPOINT LEVEL)
The target and the source substances are structurally similar substances that share the common organometallic core structure consisting of a central zinc metal bonded to four alkyldithiophosphate esters (ligands) by coordinate covalent bonds -Zn[(S2P(OR)2]2. Structural variations between the target and the source substances are related only to the alkyl (R) groups of the alkyldithiophosphate ligands. The substances in this category give thus rise to an (identical) common compound Phosphorodithioic acid moiety that can be released by the breakage of ester bonds and dissociation from the Zinc complex to which the organism would be exposed if the target substance was tested in the toxicity studies. Exposure to the parent compounds (non-transformed constituents) and to the counter alkyl alcohols, possibly released by hydrolysis of P-O bonds – non-common compounds – would not influence the prediction of the (eco)toxicological properties because they are considered to have the same biological targets and to cause the same type of effects through a common underlying mechanism due to the same functional groups (zinc cation, phosphorodithioic cation and aliphatic alcohol anionic moieties). The impurities of the target and the source substances are not expected to impact the prediction because they are identical or, if slightly structural different, belong to the same class of compounds with the same functional groups and their percentages are very low.

2. CATEGORY APPROACH JUSTIFICATION (ENDPOINT LEVEL)
Since the main constituents of the target substance are structurally similar to the constituents of the source substances with the same functional groups and the alkyl chain lengths of phosphoroditioate moieties are in the range of the established ZDDP category (C3-C12), the same mode of toxicological action is expected for the target and the source substances. The constituents of the target substance do not possess functional groups associated with other modes of action or toxicity effects. Toxicokinetic behavior of the constituents of the target substance is expected to be essentially the same as that of the source substance. Based on the results of the oral repeated dose toxicity studies with the source substances and other ZDDP category members, it is evident that the structural dissimilarities – the chain lengths of the alkyl rests – did not result in different strength of the toxicity effects. Even if absorption through GI tract of the category members with shorter alkyl rests and theoretically higher absorption rates (due to the increased water solubility and decreased molecular weights favouring absorption) is higher, their intrinsic properties are essentially the same as the findings in the toxicity studies are very similar. The impurities of the target substance are considered not to contribute to the toxicity effects because they are also structurally similar to the impurities of the source substances and consist of substances of simple structure without specific mode of action. Furthermore, their amounts are very low. Therefore, it is predicted that the target substance would possess the same toxicity potential by repeated oral exposure as the source substance.
Clinical signs:
no effects observed
Details on maternal toxic effects:
Maternal toxic effects:yes. Remark: portal-of-entry

Details on maternal toxic effects:
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
One male in the 160 mg/kg/day group was euthanized in extremis on study day 17; a gross observation of a thickened stomach was noted at necropsy. Clinical findings noted for this male approximately 1 hour following dose administration on the day of euthanasia consisted of yellow material on various body surfaces, clear material around the mouth, unkempt appearance, decreased defecation, and labored respiration. Microscopically, this male was noted with inflammation, edema, and ulceration in the non-glandular stomach, erosion and inflammation in the trachea, and lymphoid depletion in the thymus, spleen, and lymph nodes (mesenteric, mandibular, and axillary). The lesions in the non glandular stomach were considered test item-related and may have contributed to the moribund state of this male. All other animals in all dosage groups survived to the scheduled necropsies. Test item related clinical findings were noted in the 160 mg/kg/day group males and females and included rales, decreased, shallow, and/or labored respiration and salivation related findings. These findings were noted at the daily examinations, at the time of dosing, and/or approximately 1 hour following dose administration primarily during the treatment period. However, because of their sporadic occurrence, these cardio pulmonary findings were considered to be incidental and secondary to the nature of the test item and the route of administration.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
Mean body weights, body weight changes, and food consumption were unaffected by test item administration in the 10, 40, and 160 mg/kg/day groups throughout the treatment and post-treatment periods.

FUNCTIONAL OBSERVATIONAL BATTERY (FOB) AND LOCOMOTOR ACTIVITY (PARENTAL ANIMALS)
No test item-related effects were noted during the FOB or locomotor activity evaluations at any dosage level.

CLINICAL PATHOLOGY (PARENTAL ANIMALS)
There were no test substance-related effects on serum chemistry, hematology, or coagulation parameters in the 10, 40, and 160 mg/kg/day groups.

ORGAN WEIGHTS (PARENTAL ANIMALS)
There were no test item-related alterations in final body weight or organ weights at any dosage level. Significant (p<0.05) differences were observed when the control and high dose group males were compared at the recovery (post-treatment) necropsy and consisted of lower mean kidney weight relative to body weight, higher mean spleen weight relative to brain weight, higher mean left testis weight relative to brain weight, and higher mean right testis weights (absolute and relative to brain weight). There was no case where all 3 measures (absolute, relative to body weight, and relative to brain weight) were statistically significant. Thus, since the absolute weights and weights relative to body or brain weight were discordant, these organ weight changes were considered to be spurious.

GROSS PATHOLOGY (PARENTAL ANIMALS)
One male (no. 61739) in the 160 mg/kg/day group was euthanized in extremis on study day 17. The thickened stomach noted macroscopically for this male was related to test item administration. There were no other test item related internal findings observed for either sex at any dosage level at the scheduled necropsies. Macroscopic findings observed in the test item groups occurred infrequently and/or in a manner that was not dose related.

HISTOPATHOLOGY (PARENTAL ANIMALS)
Test item-related histologic observations of epithelial hyperplasia, hyperkeratosis, and inflammation of the non-glandular stomach, typically at the limiting ridge, but sometimes more widespread, were noted in the 160 mg/kg/day group males and females.
Dose descriptor:
NOEL
Remarks:
portal-of-entry
Effect level:
40 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Remarks on result:
other: localised injury to the nonglandular portion of the stomach
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Effect level:
160 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Remarks on result:
other: no effects observed at any dose level
Abnormalities:
no effects observed
Remarks on result:
not measured/tested
Remarks:
only screening test
Abnormalities:
no effects observed
Developmental effects observed:
no

VIABILITY (OFFSPRING): Mean numbers of corpora lutea and unaccounted-for sites, mean number of pups born, live litter size, the percentage of males at birth, and postnatal survival in the 10, 40, and 160 mg/kg/day groups were similar to the control group values.

CLINICAL SIGNS (OFFSPRING): The general physical condition of all F1 pups in this study were unaffected by test item administration. No test item-related clinical findings were noted for the F1 pups.

BODY WEIGHT (OFFSPRING): Mean male and female pup body weights and body weight gains in the 10, 40, and 160 mg/kg/day groups were unaffected by test item administration during PND 1-4. No statistically significant differences from the control group were noted.

GROSS PATHOLOGY (OFFSPRING): There were no remarkable macroscopic findings in the F1 pups at the scheduled necropsy at any dosage level.

Conclusions:
Test item-related moribundity, clinical findings, and microscopic findings in the non glandular portion of the stomach, characterized by epithelial hyperplasia, hyperkeratosis, and inflammation, were observed in the 160 mg/kg/day group. The injury to the nonglandular portion of the stomach was localized and considered to be irritation from test item portal-of-entry effects. Based on these results, the NOEL for portal-of-entry effects was considered to be 40 mg/kg/day, and excluding the histologic injury to the nonglandular stomach, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day. In the absence of effects on the general physical condition of the F1 pups, the NOEL for neonatal toxicity was 160 mg/kg/day.
Executive summary:

In a guideline repeated dose and reproduction / developmental screening study (OECD 422) conducted according to Good Laboratory Practices, WIL Research Labs (2010) evaluated the potential toxic effects of phosphorodithioic acid, mixed O,O-bis(iso-Bu and pentyl) esters, zinc salts when administered to rats. This study was designed to evaluate the toxic effects, including neurobehavioral effects, of the test material to parental animals and to evaluate the potential to effect male and female reproductive performance such as gonadal function, mating behavior, conception, parturition, and early postnatal development. The test material was administered orally by gavage once daily to 3 groups of Crl:CD(SD) rats at levels of 10, 40 or 160 mg/kg/day. The low- and mid-dose groups each consisted of 10 rats/sex and the high-dose group consisted of 15 rats/sex. A concurrent control group of 15 rats/sex received the vehicle, mineral oil USP, on a comparable regimen. Ten males/group selected for pairing were dosed for 14 days prior to mating through 1 day prior to euthanasia for a total of 28 doses. Ten females/group selected for pairing were dosed for 14 days prior to mating through lactation day 3 for a total of 40-52 doses; females that failed to deliver were dosed through the day prior to euthanasia (post-mating day 25) for a total of 40 doses. The extra 5 males and 5 females in the control and high-dose groups were not used for mating and were treated beginning on study day 0; following 28 doses for the males and 40 doses for the females, these animals were assigned to the post‑treatment period and remained on study for a 14-day non-dosing period.

Test item-related moribundity, clinical findings, and microscopic findings in the non glandular portion of the stomach, characterized by epithelial hyperplasia, hyperkeratosis, and inflammation, were observed in the 160 mg/kg/day group. The injury to the nonglandular portion of the stomach was localized and considered to be irritation from test item portal-of-entry effects. Based on these results, the NOEL for portal-of-entry effects was considered to be 40 mg/kg/day, and excluding the histologic injury to the nonglandular stomach, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day. In the absence of effects on the general physical condition of the F1 pups, the NOEL for neonatal toxicity was 160 mg/kg/day.

Endpoint:
developmental toxicity
Type of information:
read-across based on grouping of substances (category approach)
Adequacy of study:
weight of evidence
Justification for type of information:
REPORTING FORMAT FOR THE CATEGORY APPROACH
Please refer also to the read-across statement attached in section 13

1. HYPOTHESIS FOR THE CATEGORY APPROACH (ENDPOINT LEVEL)
The target and the source substances are structurally similar substances that share the common organometallic core structure consisting of a central zinc metal bonded to four alkyldithiophosphate esters (ligands) by coordinate covalent bonds -Zn[(S2P(OR)2]2. Structural variations between the target and the source substances are related only to the alkyl (R) groups of the alkyldithiophosphate ligands. The substances in this category give thus rise to an (identical) common compound Phosphorodithioic acid moiety that can be released by the breakage of ester bonds and dissociation from the Zinc complex to which the organism would be exposed if the target substance was tested in the toxicity studies. Exposure to the parent compounds (non-transformed constituents) and to the counter alkyl alcohols, possibly released by hydrolysis of P-O bonds – non-common compounds – would not influence the prediction of the (eco)toxicological properties because they are considered to have the same biological targets and to cause the same type of effects through a common underlying mechanism due to the same functional groups (zinc cation, phosphorodithioic cation and aliphatic alcohol anionic moieties). The impurities of the target and the source substances are not expected to impact the prediction because they are identical or, if slightly structural different, belong to the same class of compounds with the same functional groups and their percentages are very low.

2. CATEGORY APPROACH JUSTIFICATION (ENDPOINT LEVEL)
Since the main constituents of the target substance are structurally similar to the constituents of the source substances with the same functional groups and the alkyl chain lengths of phosphoroditioate moieties are in the range of the established ZDDP category (C3-C12), the same mode of toxicological action is expected for the target and the source substances. The constituents of the target substance do not possess functional groups associated with other modes of action or toxicity effects. Toxicokinetic behavior of the constituents of the target substance is expected to be essentially the same as that of the source substance. Based on the results of the oral repeated dose toxicity studies with the source substances and other ZDDP category members, it is evident that the structural dissimilarities – the chain lengths of the alkyl rests – did not result in different strength of the toxicity effects. Even if absorption through GI tract of the category members with shorter alkyl rests and theoretically higher absorption rates (due to the increased water solubility and decreased molecular weights favouring absorption) is higher, their intrinsic properties are essentially the same as the findings in the toxicity studies are very similar. The impurities of the target substance are considered not to contribute to the toxicity effects because they are also structurally similar to the impurities of the source substances and consist of substances of simple structure without specific mode of action. Furthermore, their amounts are very low. Therefore, it is predicted that the target substance would possess the same toxicity potential by repeated oral exposure as the source substance.
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
Parental toxicity was exhibited at dose levels of 100 and 200 mg/kg/day by mortality and clinical signs. Parental toxicity was also exhibited at the 200 mg/kg/day dose level by inhibition of body weight gain in males and signs of gastric irritation. No parental toxicity was observed at the 30 mg/kg/day dose level.
Dose descriptor:
NOAEL
Effect level:
30 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: developmental toxicity
Abnormalities:
no effects observed
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Neonatal toxicity (mortality) in the F1 generation was observed at the 100 and 200 mg/kg/day dose levels. Neonatal toxicity was also noted at the 200 mg/kg/day dose level by clinical signs. No neonatal toxicity was observed at a dose level of 30 mg/kg/day.
Dose descriptor:
NOAEL
Effect level:
30 mg/kg bw/day (actual dose received)
Based on:
test mat.
Remarks:
maternal dose
Sex:
male/female
Basis for effect level:
other: development toxicity
Remarks on result:
other: no mortality or clinical signs observed at this dose level
Abnormalities:
no effects observed
Developmental effects observed:
no

F0 (Parental Generation)

Two males and three females in the high dose group (200 mg/kg/day) died prior to scheduled necropsy (test days 12, 19, 8, 27 and 39). These deaths were considered treatment related. Two females in the mid dose group (100 mg/kg/day) and one female in the high dose group were euthanized on lactation days 1 or 2 due to total litter loss. Five of these animals exhibited gastric irritation upon necropsy. All other animals survived to their scheduled sacrifice.

Clinical signs noted in the Found dead or sacrificed animals included staining, matting of fur, respiratory distress, hunched appearance and mucoid diarrhea. Clinical findings noted for the surviving mid and high dose males and females included post dosing salivation, brown staining, respiratory distress and diarrhea. No treatment related clinical findings were observed in the low dose (30 mg/kg/day) animals.

 

Fertility indices (%) for the high dose males and females were slightly lower then control as follows:

                 Control            30 mg/kg       100 mg/kg         300 mg/kg

Males           91.7                  83.3                83.3                   81.8

Females      100                    83.3                91.7                   81.8

These values were within the range of the test facility historical control data (64-100%). In addition, differences from control were not statistically significant and represent only 1 or 2 fewer successful matings out of 11 or 12 males or females used for mating in the control and high dose groups. A microscopic examination of the reproductive organs of these animals did not reveal any treatment-related effects. The Study Director concluded that the low and mid dose groups were unaffected and that these data did not clearly reflect a treatment related effect in the high dose group. Other reproductive parameters (mating indices, days between pairing and coitus, gestation length and parturition) were unremarkable in all treated groups.

 

The premating (weeks 1-4) mean body weight gain of the high dose males was statistically significantly reduced compared to control. The mean body weights of the low and mid dose males and all treated female groups were unremarkable during the premating period. Gestation and lactation body weights were unremarkable in all treated groups. Food consumption data were unremarkable in all treated groups (males and females) during the premating, gestation and lactation periods. With the exception of the gastric irritation noted above in several unscheduled deaths, the macroscopic data were unremarkable. Absolute and relative (to body weight) organ weight data as well as the microscopic examination data of the F0 males and females were unremarkable. There were no treatment-related findings evident in any of these data.

 

 

F1 Litter Data

Pup body weights, live litter size and sex ratios were unremarkable. No treatment related effects were evident. An increased number of dead pups was noted in the mid dose group on day 0 of lactation. Pup viability indices in the mid dose (lactation days 1 and 4) and high dose (lactation day 4) groups were reduced. This was attributed to total litter loss by three females. Increased pup deaths were observed in the mid and high dose groups during the post-natal period. An increased incidences of pups without milk in the stomach was noted in the mid dose group. No treatment related effects were evident in the necropsy data of these found dead pups or in the necropsy data from scheduled pup necropsies.

 

Chemical analysis of dosing suspensions confirmed that they were homogeneous and of appropriate concentration throughout the study.

Conclusions:
Parental (F0) toxicity was exhibited at dose levels of 100 (mortality, clinical findings) and 200 mg/kg/day (mortality, clinical signs, reduced body weight gain, gastric irritation). A slightly reduced fertility index was also observed at 200 mg/kg/day. No F0 toxicity was observed at 30 mg/kg. Neonatal (F1) toxicity (mortality) was observed at 100 and 200 mg/kg/day. No F1 toxicity was observed at 30 mg/kg. Based on these findings the Study Director concluded that the NOAEL for both parental and neonatal toxicity was 30 mg/kg/day.
Executive summary:

This screening study according to GLP and OECD guideline 421 was designed to determine the potential adverse effects of Zinc O,O-diethylhexyldithiophosphate (ZDDP) on male and female reproduction in rats. The test material was administered orally by gavage to three groups of 12 F0 male and 12 F0 female Sprague Dawiey Crl:CD®BR rats. Dosage levels were 30, 100 and 200 mg/kg/day. For comparative purposes, a control pup of identical design was concurrently dosed with Mazola®corn oil on a comparable regimen. A dose volume of 5 ml/kg was used in all dose groups. All F0 animals were dosed for a minimum of 14 days prior to mating and through the day of necropsy for each F, animal. AU animals were observed twice daily for appearance and behavior. Body weights were recorded weekly for both sexes prior to mating; maternal body weights were also recorded on gestation days 0, 7, 14 and 20 as well as lactation days 1 and 4. Food consumption was measured for corresponding intervals prior to mating, during gestation and during lactation. AU of the surviving F0 females were allowed to deliver and rear their pups to lactation day 4. The offspring were also potentially exposed in urero and through nursing during lactation days 1-4 until euthanization on post-natal day 4. The surviving F, dams were necropsied on lactation day 4, following at least 43 days of dosing. The surviving F0 males were necropsied after the breeding period, following 28 days of dosing. F0 females with total litter loss were necropsied within 24 hours. F0 females which failed to deliver were necropsied on post-mating day 25 (evidence of mating) or 25 days following the breeding period (no evidence of mating). Organ weights were collected (all F0 treated groups) and microscopic examinations were conducted (control and high dose groups and all parental animals not surviving to the scheduled necropsies).

Slightly reduced fertility indices for males and females in the 200 mg/kg/day group (81.8% compared to control values of 91.7% and 100%, respectively) were not clearly related to treatment. Fertility indices in the 30 and 100 mg/kg/day groups were unaffected by treatment. Other reproductive parameters (mating, days between pairing and coitus, gestation and parturition) were unaffected by treatment at all dose Levels.

Two males and three females in the 200 mg/kg/day group died prior to the scheduled necropsies. Two and one females in the 100 and 200 mg/kg/day groups, respectively, were euthanized due to total litter loss prior to the scheduled necropsy. Five of these animals had internal changes consistent with gastric irritation. All other animals survived to the scheduled necropsies. The predominant clinical findings noted for surviving males and females in the 100 and 200 mg/kg/day groups at the time of dosing and/or 1-hour post dosing were salivation and brown staining on various body surfaces. Other treatment-related clinical signs observed for the 100 and 200 mg/kg/day group males and females were respiratory distress (rales, gasping) and excreta-related findings (mucoid feces, mucoid diarrhea, diarrhea and/or soft stool). No clinical signs related to ZDDP administration were observed in the 30 mg/kg/day group males and females.

Reduced mean body weight gains occurred in the 200 mg/kg/day group males during weeks 0-1, 1-2, 2-3 and 3-4. Weekly body weight gains in the 30 and 100 mg/kg/day group males and the 30, 100 and 200 mg/kg/day group females were not adversely affected by treatment. Gestation and lactation body weight gains in the 30, 100 and 200 mg/kg/day group females were not adversely affected by treatment. Food consumption in the 30, 100 and 200 mg/kg/day group males and females was unaffected by treatment.

No compound-related internal findings were noted in the surviving animals in the treated groups at the necropsy and microscopic examinations. Microscopic examination of gross lesions noted at the scheduled necropsies did not reveal any adverse effects of ZDDP administration. No adverse effects were apparent on brain, liver, kidney, testes, ovary or pituitary weights in the treated F,, males and females.

An increased number of dead pups on lactation day 0 was noted in the 100 mg/kg/day group. Pup viability indices for the 100 mg/kg/day group (lactation days 1 and 4) and the 200 mg/kg/day group (lactation day 4) were reduced (due to the three females with total litter loss). Increased numbers of pups in the 100 and 200 mg/kg/day groups were found dead during the post-natal period. The predominant clinical observation noted for the 200 mg/kg/day pups was body cool to touch. F, pup sex ratios were not adversely affected by compound administration at any dose level. Necropsy findings did not reveal a cause of death for the F, pups that died or indicate a relationship to treatment for the F, pups that were euthanized on post-natal day 4.

 

In conclusion, parental toxicity was exhibited at dose levels of 100 and 200 mg/kg/day by mortality and clinical signs. Parental toxicity was also exhibited at the 200 mg/kg/day dose level by inhibition of body weight gain in males and signs of gastric irritation. No parental toxicity was observed at the 30 mg/kg/day dose level. Slightly reduced fertility indices were observed at the 200 mg/kg/day dose level. Reproductive performance (fertility, mating, days between pairing and coitus, gestation and parturition) was unaffected by treatment at the 30 and 100 mg/kg/day dose levels. Neonatal toxicity (mortality) in the F1 generation was observed at the 100 and 200 mg/kg/day dose levels. Neonatal toxicity was also noted at the 200 mg/kg/day dose level by clinical signs. No neonatal toxicity was observed at a dose level of 30 mg/kg/day. Based on the results of this study, a dose level of 30 mg/kg/day was considered to be the NOAEL (no observable adverse effect level) for parental and neonatal toxicity.

Endpoint:
developmental toxicity
Type of information:
experimental study planned (based on read-across)
Remarks:
testing proposed with Phosphorodithioic acid, mixed O,O-bis(iso-Bu and pentyl) esters, zinc salts (EC 270-608-0)
Study period:
Investigation to begin as soon as possible after completion of prenatal development toxicity study
Justification for type of information:
The prenatal developmental toxicity study according to OECD guideline 414 proposed for the source substance CAS 68457-79-4 is used as a source for read-across for the target substance (CAS 10147-77-7).

REPORTING FORMAT FOR THE CATEGORY APPROACH
Please refer also to the read-across statement attached in section 13

1. HYPOTHESIS FOR THE CATEGORY APPROACH (ENDPOINT LEVEL)
The target and the source substances are structurally similar substances that share the common organometallic core structure consisting of a central zinc metal bonded to four alkyldithiophosphate esters (ligands) by coordinate covalent bonds -Zn[(S2P(OR)2]2. Structural variations between the target and the source substances are related only to the alkyl (R) groups of the alkyldithiophosphate ligands. The substances in this category give thus rise to an (identical) common compound Phosphorodithioic acid moiety that can be released by the breakage of ester bonds and dissociation from the Zinc complex to which the organism would be exposed if the target substance was tested in the toxicity studies. Exposure to the parent compounds (non-transformed constituents) and to the counter alkyl alcohols, possibly released by hydrolysis of P-O bonds – non-common compounds – would not influence the prediction of the toxicological properties because they are considered to have the same biological targets and to cause the same type of effects through a common underlying mechanism due to the same functional groups (zinc cation, phosphorodithioic cation and aliphatic alcohol anionic moieties). The impurities of the target and the source substances are not expected to impact the prediction, because they are identical or, if slightly structural different, belong to the same class of compounds with the same functional groups and their percentages are very low.

2. CATEGORY APPROACH JUSTIFICATION (ENDPOINT LEVEL)
The results of the two source substances indicate that local irritation of the gastrointestinal tract is the primary effect of the substances in the ZDDP category. This is also in line with the skin and eye irritating properties of the substances.
Since the main constituents of the target substance are structurally similar to the constituents of the source substances with the same functional groups and the alkyl chain lengths of phosphorodithioate moieties are in the range of the established ZDDP category (C3-C12), the same mode of toxicological action is expected for the target and the source substances. The constituents of the target substance do not possess functional groups associated with other modes of action or toxicity effects. The toxicokinetic behaviour of the constituents of the target substance is expected to be essentially the same as that of the source substance. Based on the results of the oral repeated dose toxicity studies with the source substances and other ZDDP category members, it is evident that the structural dissimilarities – the chain lengths of the alkyl rests – did not result in different strength of the toxicity effects. Even if absorption through GI tract of the category members with shorter alkyl rests is higher (due to the increased water solubility and decreased molecular weights favouring absorption), their intrinsic properties are essentially the same as the findings in the toxicity studies are very similar. Therefore, the prenatal developmental toxicity study proposed for the source substance will generate adequate data to fulfil the data requirements for the target substance. The impurities of the target substance are considered not to contribute to the toxicity effects because they are also structurally similar to the impurities of the source substances and consist of substances of simple structure without specific mode of action. Furthermore, their amounts are very low. Therefore, it is predicted that the target substance (CAS 10147-77-7) would possess the same reproductive and developmental toxicity potential by repeated oral exposure as the source substance (CAS 68457-79-4).
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Species:
rat
Route of administration:
oral: feed
Details on exposure:
Oral administration of the test item is relevant because physicochemical properties mean systemic uptake via the oral route is more likely than via the dermal or inhalation routes. However, absorption is expected to be slow/inefficient via the gastrointestinal tract and previous studies have demonstrated local irritant properties at the site of entry. It is therefore proposed that administration of the substance to animals via dietary admixture is the most likely to achieve maximum exposure without causing undue pain and suffering.
Details on study design:
Physicochemical properties mean that category substances are predicted to be absorbed very slowly via the dermal route and no significant uptake is expected. With respect to the inhalation route, category substances have a vapour pressure of < 0.01 Pa at 25 °C and, under normal conditions of handling and use, absorption of test material in the form of vapours, gases or mists via the respiratory tract is not expected to be relevant or significant.
Abnormalities:
not specified
Developmental effects observed:
not specified
Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
30 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
High due to guideline studies
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

Developmental toxicity / teratogenicity, screening:

source substance Zinc bis(O,O-diisobutyl dithiophosphate) (CAS 68457 -79 -4)

In a guideline repeated dose and reproduction / developmental screening study (OECD 422) conducted according to Good Laboratory Practices, WIL Research Labs (2010) evaluated the potential toxic effects of phosphorodithioic acid, mixed O,O-bis(iso-Bu and pentyl) esters, zinc salts when administered to rats. This study was designed to evaluate the toxic effects, including neurobehavioral effects, of the test material to parental animals and to evaluate the potential to effect male and female reproductive performance such as gonadal function, mating behavior, conception, parturition, and early postnatal development. The test material was administered orally by gavage once daily to 3 groups of Crl:CD(SD) rats at levels of 10, 40 or 160 mg/kg/day. The low- and mid-dose groups each consisted of 10 rats/sex and the high-dose group consisted of 15 rats/sex. A concurrent control group of 15 rats/sex received the vehicle, mineral oil USP, on a comparable regimen. Ten males/group selected for pairing were dosed for 14 days prior to mating through 1 day prior to euthanasia for a total of 28 doses. Ten females/group selected for pairing were dosed for 14 days prior to mating through lactation day 3 for a total of 40-52 doses; females that failed to deliver were dosed through the day prior to euthanasia (post-mating day 25) for a total of 40 doses. The extra 5 males and 5 females in the control and high-dose groups were not used for mating and were treated beginning on study day 0; following 28 doses for the males and 40 doses for the females, these animals were assigned to the post‑treatment period and remained on study for a 14-day non-dosing period.

Test item-related moribundity, clinical findings, and microscopic findings in the non glandular portion of the stomach, characterized by epithelial hyperplasia, hyperkeratosis, and inflammation, were observed in the 160 mg/kg/day group. The injury to the nonglandular portion of the stomach was localized and considered to be irritation from test item portal-of-entry effects. Based on these results, the NOEL for portal-of-entry effects was considered to be 40 mg/kg/day, and excluding the histologic injury to the nonglandular stomach, the no-observed-adverse-effect level (NOAEL) for systemic toxicity was considered to be 160 mg/kg/day. In the absence of effects on the general physical condition of the F1 pups, the NOEL for neonatal toxicity was 160 mg/kg/day.

source substance Zinc bis[O,O-bis(2-ethylhexyl)] bis(dithiophosphate) (CAS 4259 -15 -8)

This screening study according to GLP and OECD guideline 421 was designed to determine the potential adverse effects of Zinc O,O-diethylhexyldithiophosphate(ZDDP) on male and female reproduction in rats. The test material was administered orally by gavage to three groups of 12 F0 male and 12 F0 female Sprague Dawiey Crl:CD®BR rats. Dosage levels were 30, 100 and 200 mg/kg/day. For comparative purposes, a control pup of identical design was concurrently dosed with Mazola®corn oil on a comparable regimen. A dose volume of 5 ml/kg was used in all dose groups. All F0 animals were dosed for a minimum of 14 days prior to mating and through the day of necropsy for each F, animal. All animals were observed twice daily for appearance and behavior. Body weights were recorded weekly for both sexes prior to mating; maternal body weights were also recorded on gestation days 0, 7, 14 and 20 as well as lactation days 1 and 4. Food consumption was measured for corresponding intervals prior to mating, during gestation and during lactation. All of the surviving F0 females were allowed to deliver and rear their pups to lactation day 4. The offspring were also potentially exposed in utero and through nursing during lactation days 1-4 until euthanization on post-natal day 4. The surviving F, dams were necropsied on lactation day 4, following at least 43 days of dosing. The surviving F0 males were necropsied after the breeding period, following 28 days of dosing. F0 females with total litter loss were necropsied within 24 hours. F0 females which failed to deliver were necropsied on post-mating day 25 (evidence of mating) or 25 days following the breeding period (no evidence of mating). Organ weights were collected (all F0 treated groups) and microscopic examinations were conducted (control and high dose groups and all parental animals not surviving to the scheduled necropsies).

In conclusion, parental toxicity was exhibited at dose levels of 100 and 200 mg/kg/day by mortality and clinical signs. Parental toxicity was also exhibited at the 200 mg/kg/day dose level by inhibition of body weight gain in males and signs of gastric irritation. No parental toxicity was observed at the 30 mg/kg/day dose level. Slightly reduced fertility indices were observed at the 200 mg/kg/day dose level. Reproductive performance (fertility, mating, days between pairing and coitus, gestation and parturition) was unaffected by treatment at the 30 and 100 mg/kg/day dose levels. Neonatal toxicity (mortality) in the F1 generation was observed at the 100 and 200 mg/kg/day dose levels. Neonatal toxicity was also noted at the 200 mg/kg/day dose level by clinical signs. No neonatal toxicity was observed at a dose level of 30 mg/kg/day. Based on the results of this study, a dose level of 30 mg/kg/day was considered to be the NOAEL (no observable adverse effect level) for parental and neonatal toxicity.

Justification for classification or non-classification

In accordance to Regulation (EC) No 1272/2008 classification of this substance is not required for reproductive toxicity occurring at maternally toxic doses.