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EC number: 700-823-1 | CAS number: 55514-22-2
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
![](https://poisoncentres.echa.europa.eu/o/diss-blank-theme/images/factsheets/A-REACH/factsheet/print_toxicological-information.png)
Toxicity to reproduction
Administrative data
- Endpoint:
- screening for reproductive / developmental toxicity
- Remarks:
- based on test type (migrated information)
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 2012
- Reliability:
- 1 (reliable without restriction)
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 012
- Report date:
- 2012
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Limit test:
- no
Test material
- Reference substance name:
- 3-[4-(5,6-diphenyl-1,2,4-triazin-3-yl)phenyl]-5,6-diphenyl-1,2,4-triazine
- EC Number:
- 700-823-1
- Cas Number:
- 55514-22-2
- Molecular formula:
- C36H24N6
- IUPAC Name:
- 3-[4-(5,6-diphenyl-1,2,4-triazin-3-yl)phenyl]-5,6-diphenyl-1,2,4-triazine
- Details on test material:
- - Name of test material (as cited in study report): WP30
- Substance type: mono-constituent substance
- Physical state: powder
- Analytical purity: 99.8 %
- Impurities (identity and concentrations): unknown impurity with MW 380: 0.19 %, unknown impurity with MW 558: 0.29 %,
- Purity test date: July 7th 2011, expiry date January 7th 2012
- Lot/batch No.: LP140
- Stability under test conditions: stable
- Storage condition of test material: at room temperature in a dry and well ventilated place
Constituent 1
Test animals
- Species:
- rat
- Strain:
- Sprague-Dawley
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- Number: 130 Sprague-Dawley rats (65 males and 65 females) were received at CIT on 01 September 2011.
Strain and sanitary status: Crl CD® (SD) IGS BR, Caesarian Obtained, Barrier Sustained-Virus Antibody Free (COBS-VAF®).
Breeder: Charles River Laboratories France, L’Arbresle, France.
Age/Weight: at the beginning of the treatment period, the males were approximately 10 weeks old and weighed approximately 400 g (range 348 g to 451 g). The females were approximately 9 weeks old and weighed approximately 220 g (range 180 g to 246 g). The males and the females were sexually mature and were not siblings. The females were previously unmated.
Acclimation: the animals were acclimated for a period of 5 days before the beginning of the treatment period. A larger number of animals than necessary were acclimated to permit selection and/or replacement of individuals.
Allocation to study: during the acclimation period, the required number of animals (64 males and 64 females) was selected according to body weight and clinical condition. They were then allocated to groups (by sex) according to a computerized stratification procedure bases on body weight, so that the average body weight of each group was similar (these data are not shown).
Identification: each animal was individually identified by an ear tattoo (unique CIT identity number.
Environmental conditions:
From arrival at CIT, the animals were housed in a barriered rodent unit. The animal room conditions were set as follows: . temperature : 22 ± 2°C, . relative humidity : 50 ± 20%, light/dark cycle : 12h/12h (7:00 - 19:00), . ventilation : about 12 cycles/hour of filtered, non-recycled air.
The temperature and relative humidity were recorded continuously. The animal room was disinfected before the arrival of the animals and cleaned regularly thereafter.
Housing:
The animals were individually housed, except during pairing, in polycarbonate cages (UAR,
43.0 cm x 21.5 cm x 18.0 cm) with stainless steel lids and containing autoclaved sawdust (SICSA, Alfortville, France). Toward the end of gestation and during lactation with their litter, autoclaved wood shavings (SICSA, Alfortville, France) was provided as nesting material, a few days before delivery and during the lactation period. Each cage contained an object (Nylabone) for enrichment of the environment for the rats.
The cages were placed in numerical order on the racks.
Food and water:
The animals had free access to SSNIFF R/M-H pelleted maintenance diet, batch No. 9615507 (SSNIFF Spezialdiäten GmbH, Soest, Germany), which was distributed weekly. The animals had free access to bottles containing tap water (filtered with a 0.22 µm filter).
Contaminant analyses:
The batches of diet, sawdust and wood shavings are analyzed by the suppliers for composition and contaminant levels.
Bacterial and chemical analyses of water are performed regularly by external laboratories. These analyses include the detection of possible contaminants (pesticides and heavy metals).
No contaminants were present in the diet, drinking water, sawdust or wood shavings at levels which could be expected to interfere with, or prejudice, the outcome of the study.
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- CMC (carboxymethyl cellulose)
- Remarks:
- 0.5 % in water
- Details on exposure:
- PREPARATION OF DOSING SOLUTIONS:
VEHICLE
- Justification for use and choice of vehicle: validated analytical method demonstrating stability and homogeneity of the test item in the vehicle
- vehicle: 0.5% aqueous methylcellulose solution prepared using: . purified water, obtained by reverse osmosis - Details on mating procedure:
- Mating Females were paired with males from the same dose-level group. One female was placed with one male, in the latter's cage, during the night. Sibling pairings was avoided. Confirmation of mating was made in the morning by checking for the presence of a vaginal plug or for sperm in a vaginal lavage. The day of confirmed mating was designated day 0 p.c.. Each female was placed with the same male until mating occurs or 14 days have elapsed. Any pairs with no evidence of mating after 14 days was separated and the female was placed for a further 7-day period with a different male from the same dose-level group who had already mated. The pre-coital time was calculated for each female. Satellite animals were not mated and were allocated for toxicokinetic investigations only. The estrous cycle stage was determined from a fresh vaginal lavage (stained with methylene blue), each morning during the pairing period, until the females were mated.
- Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Chemical analysis of the dosage forms
1 Principle and validation of the method
A High Performance Liquid Chromatography with UV detection (HPLC/UV) analytical method for the determination of WP30 in dosage form samples was used.
The validation of the analytical method was conducted in CIT/Study No. 37878 VAA. The analytical method was validated for dosage forms ranging from 10 to 100 mg/mL in aqueous solution of methylcellulose at 0.5% (w/w).
2 Composition of an analytical sequence Each analytical sequence consisted of at least: . a blank sample (diluent only), . ten standard samples at nominal concentration, prepared from two independent standard solutions, . study samples prepared from aliquots of the dosage forms.
The standard samples bracketed the dosage form samples. The blank sample was checked for the absence of chromatographic interference.
3 Analytical sequence suitability rules The acceptance of the analytical sequence depended on the precision of the standard samples and
on the agreement of the standard sample results. Acceptance criteria are defined in CIT Standard Operating Procedures (SOPs).
The acceptance criteria include: . precision of the standard solutions [%RSD (Relative Standard Deviation) must pass defined acceptance criteria],
agreement of the standard samples (the mean response factor of standard samples prepared from standard solution #1 must agree with the mean response factor of standard samples prepared from standard solution #2).
4 Study sample re-analysis rules No re-analysis was performed since the acceptance criteria were met for each series.
5 Determination of dosage form homogeneity and stability The test item dosage forms were prepared weekly (according to CIT/Study No. 37879 AHS, dosage
form stability/homogeneity was confirmed from 10 to 100 mg/mL for a 9-day period) and were maintained at +4°C and protected from light prior to use and delivered in brown flasks.
The homogeneity of each test item dosage form prepared for use was determined in weeks 1 and 5
6 Determination of WP30 concentration in dosage forms The concentration of the test item in samples of each control and test item dosage forms prepared for use were determined in weeks 1, 3 and 5 . Before day 1 and whenever possible for the other formulations, these analyses were performed prior to administration of the dosage forms to the animals.
Acceptance criterion: . measured concentration = nominal concentration ± 15%. - Duration of treatment / exposure:
- The dosage forms were administered daily according to the following schedule: . in the males: -2 weeks before pairing,
-during the pairing period (3 weeks), -until sacrifice (at least 5 weeks in total),
. in the females: -2 weeks before pairing, -during the pairing period (3 weeks), -during gestation, -during lactation until day 5 p.p. inclusive, -until sacrifice for females with no delivery - Frequency of treatment:
- Daily
Doses / concentrations
- Remarks:
- Doses / Concentrations:
100, 300 and 1000 mg/kg.day
Basis:
analytical conc.
- No. of animals per sex per dose:
- 10 males and 10 females per dose
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- In addition, three groups of eight males and eight females received also the test item for 5 weeks at dose-levels of 100, 300 or 1000 mg/kg/day for blood plasma concentration measurements on study day 1 and at the end of the treatment period (study week 5). The dosing volume was 10 mL/kg/day.
- Positive control:
- None
Examinations
- Parental animals: Observations and examinations:
- CAGE SIDE OBSERVATIONS: Yes
Morbidity and mortality Each animal was checked for mortality and morbidity once a day before the treatment period and at least twice a day during the treatment period, including weekends and public holidays. From arrival, each animal was observed once a day as part of routine examinations. From the start of the treatment period, each animal was observed once a day, at approximately the same time, for the recording of clinical signs.
DETAILED CLINICAL OBSERVATIONS: Yes
Detailed clinical observations (principal animals) Detailed clinical examinations were performed on all animals outside the home cage, in a standard arena, once before the beginning of the treatment period and then once a week until the end of the study. Observations included (but were not limited to) changes in the skin, fur, eyes, mucous membranes, occurrence of secretions and excretions and autonomic activity (e.g. lachrymation, piloerection, pupil size, unusual respiratory pattern). Changes in gait, posture and response to handling as well
as the presence of clonic or tonic movements, stereotypes (e.g. excessive grooming, repetitive circling) or bizarre behavior (e.g. self-mutilation, walking backwards) were also recorded.
BODY WEIGHT: Yes
The body weight of each male (principal and satellite animals) was recorded on the first day of treatment (day 1), then once a week until sacrifice. The body weight of each female was recorded on the first day of treatment (day 1), then once a week until mating or until sacrifice (principal and satellite animals) and on days 0, 7, 14 and 20 post-coitum and on days 1 and 5 p.p. (principal animals).
The body weight of each pup was recorded on days 1 and 5 p.p..
OTHER:
1 Functional Observation Battery (principal animals) The first five males and the first five females to deliver from each group were evaluated once at
the end of the treatment period. For the males, this was conducted after about 5 weeks of treatment and for females, on day 5 p.p. after sacrifice of the pups.
This included a detailed clinical examination, measurement of reactivity to manipulation or to
different stimuli and motor activity.
The animals were randomized in order to ensure "blind" evaluation.
All animals were observed in the cage, in the hand and in the standard arena.
The following parameters were assessed and graded:
. "touch escape" or ease of removal from the cage,
. in the hand: fur appearance, salivation, lachrymation, piloerection, exophthalmos, reactivity to handling, pupil size (presence of myosis or mydriasis),
. in the standard arena (2-minute recording): grooming, palpebral closure, defecation, urination, tremors, twitches, tonic and clonic convulsions, gait, arousal (hypo- and hyper-activity), posture, stereotypy, behavior, breathing, ataxia and hypotonia.
2 Reactivity to manipulation or to different stimuli (principal animals) The following parameter measurements, reflexes and responses were recorded: . touch response, . forelimb grip strength, . pupillary reflex, . visual stimulus response, . auditory startle reflex, . tail pinch response, . righting reflex,
. landing foot splay, . at the end of observation: rectal temperature.
3 Motor activity (principal animals) Finally, motor activity of all animals was measured once by automated infra-red sensor equipment over a 60-minute period.
Values for rearing are not presented in the report for males W29621 (1M), W26953 (3M) and female W27028 (1F)
4 Food consumption (principal animals) The quantity of food consumed by each male was measured once a week, over a 7-day period, from the first day of treatment until the start of the pairing period. The quantity of food consumed by each female was measured once a week, over a 7-day period, from the first day of treatment until the start of the pairing period, during pregnancy at the
intervals days 0-7, 7-14 and 14-20 p.c. and during lactation for interval day 1-5 p.p.. During the pairing period, the food consumption was measured for neither males nor females.
Food intake per animal and per day was calculated by noting the difference between the food given and that in the food-hopper the next time. - Oestrous cyclicity (parental animals):
- The estrous cycle stage was determined from a fresh vaginal lavage (stained with methylene blue), each morning during the pairing period, until the females were mated.
- Sperm parameters (parental animals):
- Microscopic examination post mortem
- Litter observations:
- The total litter sizes and numbers of pups of each sex were recorded after birth. The pups were observed daily for clinical signs of toxicity and pup body weights were recorded on days 1 and 5 p.p
- Postmortem examinations (parental animals):
- The males were sacrificed during study week 6. Body weights and selected organs weights were recorded and a complete macroscopic post-mortem examination performed, with particular attention paid to the reproductive organs. A microscopic examination was also conducted on selected organs from the first five males in the control group and the high-dose group. Microscopic examination was conducted on all macroscopic lesions from all groups.
Dams were sacrificed on day 6 p.p.. Body weights and selected organs weights were recorded and a complete macroscopic examination was performed, with particular attention paid to the reproductive organs. A microscopic examination was then conducted on selected organs from the first five females to deliver in the control group and the high-dose group and on any macroscopic lesions from all groups. - Postmortem examinations (offspring):
- Pups, including those found dead before study termination, were also submitted for a macroscopic post-mortem examination.
- Statistics:
- The following statistical tests were done:
-For hematology, biochemistry and urinanalysis parameters, the normality of data distribution was checked. This was followed by assessment of homogenity of variance or by logarithmic transformation of values
For Organ weights and depending of normaity of each group, a non-parametric (Kruska-Wallis followed by a Dunn test ) or a parametric (one way analysis of variance followed by a Dunnett test) approach was selected.
The other data were compared by one-way analysis of variances and Dunnett test or by Fisher exact probability test. - Reproductive indices:
- The total litter size and numbers of pups of each sex were recorded as soon as possible after birth.
Any gross malformations in pups were noted. The litters were observed daily in order to note the number of live, dead and cannibalized pups - Offspring viability indices:
- The litters were observed daily in order to note the number of live, dead and cannibalized pups
Results and discussion
Results: P0 (first parental generation)
General toxicity (P0)
- Clinical signs:
- no effects observed
- 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:
- no effects observed
- Other effects:
- not examined
- Description (incidence and severity):
- Test substance intake: not relevant for a gavage study
Reproductive function / performance (P0)
- Reproductive function: oestrous cycle:
- no effects observed
- Reproductive function: sperm measures:
- no effects observed
- Reproductive performance:
- no effects observed
Details on results (P0)
Effect levels (P0)
- Dose descriptor:
- NOEL
- Effect level:
- > 1 000 mg/kg bw/day (actual dose received)
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: No effect observed
Results: F1 generation
General toxicity (F1)
- 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 examined
Details on results (F1)
Overall reproductive toxicity
- Reproductive effects observed:
- not specified
Applicant's summary and conclusion
- Conclusions:
- Based on the experimental conditions of this study: . the No Observed Effect Level (NOEL) for parental toxicity was considered to be 1000 mg/kg/day, . the NOEL for reproductive performance (mating and fertility) was considered to be 1000 mg/kg/day, . the NOEL for toxic effects on progeny was considered to be 1000 mg/kg/day.
- Executive summary:
The objective of this study was to evaluate the potential toxic effects of the test item, WP30, following daily oral (gavage) administration to male and female rats from before mating, during mating and, for the females, throughout gestation until day 5post-partum (p.p.)inclusive. This study provides initial information on possible health hazards (including neurological and immunological effects) likely to arise from repeated exposure over a limited period of time. It can also indicate effects on male and female reproductive performance, such as gonadal function, mating behavior, conception, development of the conceptus and parturition.
MethodsThree groups of 10 male and 10 female Sprague-Dawley rats received the test item, WP30, daily, by oral (gavage) administration, before mating (2 weeks), during mating (3 weeks at least for the males and up to 3 weeks for the females) and, for the females, throughout gestation and until day 5p.p., at dose-levels of 100, 300 or 1000 mg/kg/day. A group of 10 males and 10 females received the vehicle control, 0.5% (w/w) aqueous methylcellulose, under the same experimental conditions. In addition, three groups of eight males and eight females received also the test item for 5 weeks at dose-levels of 100, 300 or 1000 mg/kg/day for blood plasma concentration measurements on study day 1 and at the end of the treatment period (study week 5). The dosing volume was 10 mL/kg/day.
Animals were checked daily for clinical signs, mortality, and detailed clinical observations were conducted weekly. In males, body weights and food consumption were recorded weekly until mating and then body weight were recorded at designated intervals. In females, body weights and food consumption were recorded weekly until mating and then at designated intervals throughout gestation and lactation. The animals were paired for mating after 2 weeks of treatment and the dams were allowed to litter and rear their progeny until day 5p.p.. The total litter sizes and numbers of pups of each sex were recorded after birth. The pups were observed daily for clinical signs of toxicity and pup body weights were recorded on days 1 and 5p.p.. A Functional Observation Battery including touch response, forelimb grip strength, pupillary reflex, visual stimulus response, auditory startle reflex, tail pinch response, righting reflex, landing foot splay, rectal temperature and motor activity was performed on five males and females per group at the end of the study. Prior to sacrifice, blood samples were also taken from these animals for analysis of hematology, urinalysis and blood biochemistry parameters.
The males were sacrificed during study week 6. Body weights and selected organs weights were recorded and a complete macroscopicpost-mortemexamination performed, with particular attention paid to the reproductive organs. A microscopic examination was also conducted on selected organs from the first five males in the control group and the high-dose group. Microscopic examination was conducted on all macroscopic lesions from all groups.
Dams were sacrificed on day 6p.p..Body weights and selected organs weights were recorded and a complete macroscopic examination was performed, with particular attention paid to the reproductive organs. A microscopic examination was then conducted on selected organs from the first five females to deliver in the control group and the high-dose group and on any macroscopic lesions from all groups.
Pups, including those found dead before study termination, were also submitted for a macroscopicpost-mortemexamination.
Results
The WP30 concentrations in the administered dosage forms were within an acceptable range of variation (± 15%). WP30 was not detected in control samples.
ToxicokineticsOn determination of blood plasma concentration for toxicokinetic calculation, none of the satellite male and female rats had significant blood plasma levels on day 1 or at the end of the treatment period (blood plasma level < 0.500 ng/mL, the limit of quantification), with the exception of two satellite males and five satellite females which had blood plasma levels slightly higher than the limit of quantification on study day 1 or at the end of the treatment period.
Overall, it was considered that there was no significant systemic exposure to the test item.
MortalityThere were no treatment-related deaths.
Clinical signsThere were no treatment-related clinical signs.
Functional Observationand motor activityAmongst the findings noted during the study, none were indicative of a treatment-related effect.
Body weight and food consumptionThere were no treatment-related effects or significant toxicological effects on mean body weight, mean body weight change and mean food consumption neither in males nor in females.
HematologyThere were no effects on mean hematology parameters.
Blood biochemistry and urinalysisThere were no toxicological significant effects on mean blood chemistry parameters and urinalysis.
Pairing, mating and fertilityThere were no treatment-related effects on pairing, mating and fertility parameters. There were no indications of abnormal estrous cycles in any treated females.
Pup observationsThere were no effects on live birth, viability and lactation indexes.
Organ weightsNo organ weight differences between treated and control rats were attributed to treatment with WP30.
Macroscopicpost-mortemexaminationThere were no treatment-related macroscopic findings.
CIT/Study No. 37881 RSR/WP30/Institut de RechercheFabre Sponsor Reference Number: DSPC-11-0267
Microscopic examinationThere were no treatment-related microscopic findings. From a morphological point of view, spermatogenesis in treated male rats was intact: all germ cells were present and interstitial cell cellularity and morphology was within normal range.
Conclusion
The test item, WP30, was administered daily by oral gavage to male and female Sprague-Dawley rats, for 2 weeks before mating, during mating (3 weeks at least for the males and up to 3 weeks for the females), and (for females) throughout gestation and until day 5post-partum, at dose-levels of 100, 300 or 1000 mg/kg/day.
On determination of blood plasma concentration for toxicokinetic calculation, none of the satellite male and female rats had significant blood plasma levels on day 1 or at the end of the treatment period (blood plasma level < 0.500 ng/mL, the limit of quantification), with the exception of two satellite males and five satellite females which had blood plasma levels slightly higher than the limit of quantification on study day 1 or at the end of the treatment period. Overall, it was considered that there was no significant systemic exposure to the test item.
Based on the experimental conditions of this study: . the No Observed Effect Level (NOEL) for parental toxicity was considered to be 1000 mg/kg/day, . the NOEL for reproductive performance (mating and fertility) was considered to be 1000 mg/kg/day, . the NOEL for toxic effects on progeny was considered to be 1000 mg/kg/day.
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