Registration Dossier

Toxicological information

Endpoint summary

Currently viewing:

Administrative data

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records
Reference
Endpoint:
two-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study. For read-across justification see Section 13.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)
Deviations:
yes
Remarks:
(proportion Male/female during mating 1/2; oestrus cycle and sperm parameters not analysed)
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Charles River CD rats
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: -P/F0: 5-6 wks; -F1: 22 days
- Housing: single (besides mating and lactation period)
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 16 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-23
- Humidity (%): 25-60
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
volume of 10 ml/kg
Details on mating procedure:
- M/F ratio per cage: 1/2
- Length of cohabitation: maximum of 15 days
- Proof of pregnancy: vaginal plug in vaginal smear referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged: individually
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
- P/F0: from study initiation to the end of the generation
- F1: at 22 days of age and continued throughout the generation (Premating (Premating exposure period: F0 and F1, a minimum of 80 days) and throughout mating, gestation and lactation)
Frequency of treatment:
daily, 7 days/week
Details on study schedule:
P/F0 rats were bred twice to produce the F1a and F1b litters. At weaning, ten males and 20 females from each dose group of the F1b litters were randomly selected to become parents of the F2a and F2b litters. A minimum of nine additional male and eight additional female pups per group were randomly selected to be used as replacements for animals dying prior to initiation of the generation.
Remarks:
Doses / Concentrations:
0, 20, 55, and 150 mg/kg
Basis:
nominal conc.
No. of animals per sex per dose:
10 males, 20 females
Control animals:
yes, concurrent vehicle
Parental animals: Observations and examinations:
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice a day


BODY WEIGHT: Yes
- Time schedule for examinations: weekly (additionally, female body weights were measured at intervals during gestation and lactation)


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes


OTHER: male and female fertility, mean gestation length
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of 10 pups/litter (5/sex/litter as nearly as possible); excess pups were killed and discarded.


PARAMETERS EXAMINED
The following parameters were examined in [P/F0 and F1] offspring: number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies


GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities (besides organ weights); possible cause of death was determined for pups born or found dead.
Postmortem examinations (parental animals):
SACRIFICE
- all P/F0 parents


GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations.


HISTOPATHOLOGY / ORGAN WEIGHTS
These tissues were prepared for microscopic examination and weighed, respectively: adrenal, colon, heart, ileum, duodenum, kidney, liver, lung, bone marrow (sternum), brain, eye, mesenteric lymph node, mammary gland, pituitary, pancreas, salivary gland, skin, ovary, spleen, stomach, testis (with epididymis), thyroid (with a section of trachea and oesophagus), spinal cord (cervical), urinary bladder, prostate, uterus, all other gross lesions including tissue masses
Postmortem examinations (offspring):
SACRIFICE
- all F1 parents
- The F1 offspring not selected as parental animals and all F2 offspring were sacrificed



GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.


HISTOPATHOLOGY / ORGAN WEIGTHS
These tissues were prepared for microscopic examination and weighed, respectively: adrenal, colon, heart, ileum, duodenum, kidney, liver, lung, bone marrow (sternum), brain, eye, mesenteric lymph node, mammary gland, pituitary, pancreas, salivary gland, skin, ovary, spleen, stomach, testis (with epididymis), thyroid (with a section of trachea and oesophagus), spinal cord (cervical), urinary bladder, prostate, uterus, all other gross lesions including tissue masses
Statistics:
All statistical analyses compared the treatment groups with the control group, with a level of significance at p<0 .05. Significance at p<0 .01 was also indicated. 1. Parental Body Weights: The parental body weights by sex were analysed by one-way analysis of variance, Bartlett's test for homogeneity of variances, and the appropriate t-test (for equal or unequal variances). Significant differences were determined using Dunnett's multiple comparison tables. Analyses for the P/F0 generation were conducted at one week prior to the F1a mating (week 11) and at termination of the generation (week 32). Analyses for the F1 generation were conducted at the first week of the generation (week 30), one week prior to the F2a mating (week 41) and at the termination of the generation (week 61). 2 . Male and Female Fertility: Male and female fertility indices were compared using the Chisquare test criterion with Yates' correction for 2 x 2 contingency tables and/or Fisher's exact probability test to judge levels of significant differences. 3. Pup Survival indices: The proportion of live pups at birth per total number born and the survival indices at lactation days 4, 7, 14 and 21 were compared by the Mann-Whitney U-test to judge significant differences. 4. Litter Size and Pup Body Weights: The mean number of liveborn pups per litter and mean body weights of pups were analysed by one-way analysis of variance, Bartlett's test for homogeneity of variances, and the appropriate t- test (for equal or unequal variances). Significant differences were determined using Dunnett's multiple comparison tables. 5. Organ Weights: Absolute and relative organ weights (P/F0, F1, F2a and F2b) were compared by analysis of variance (one-way classification), Bartlett's test for homogeneity of variances and the appropriate t-test (for equal or unequal variances) using Dunnett's multiple comparison tables to judge significance of differences.
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS): With the exception of a few cases of respiratory rales, the clinical appearance and behaviour of all treated animals were not remarkably different from the controls. Mortality of adult males and females was 0 to 10% in the low and mid dose group and 65 to 70% in the high dose group.


BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS): P/F0 body weights in the high-dose group were significantly reduced by week 11 (males: -14.1%; females: -10.4%) in both sexes and this reduction increased as the generation progressed (males: -19.7% on study week 32; females: -18%). The mid-dose group means were low but not statistically significantly different from the controls, except at the end of the generation in the females. No effects in the low dose group in either sex during both generations. No treatment-related effects on food consumption were observed in any group during the study (a slight decrease in mean female food consumption (both g/rat/day and g/kg/day) was observed throughout the P/F0 generation in the 20 mg/kg/day group, but this was considered as not treatment-related.


GROSS PATHOLOGY (PARENTAL ANIMALS)/HISTOPATHOLOGY (PARENTAL ANIMALS): Macroscopic and microscopic examination of tissues from P/F0 adults revealed compound-related changes in the kidneys and bladder of rats from all dose groups. However, renal cortical necrosis, present in 60% of the males and in 15% of the females from this group, was only observed in the high-dose group.


OTHER FINDINGS (PARENTAL ANIMALS): Inflammatory changes were observed in stomachs of rats from all groups. Since these changes could arise by a variety of mechanisms, it was not possible to conclude they were directly related to maleic anhydride. Fertility was significantly reduced in the treatment groups at several time points; however, neither a dose-related reduction nor a pattern within a generation suggested a treatment-related effect (male and female fertility indices were reduced in the 20 and 150 mg/kg/day groups in the F1a mating when compared with the control group. The relation of these findings to treatment is doubtful since the F1b mating fertility indices were generally comparable at all treatment levels.)
Key result
Dose descriptor:
NOAEL
Remarks:
(fertility)
Effect level:
55 other: mg/kg/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No effects; highest dose tested without complete maternal mortality
Remarks on result:
other: Generation: P/F1 (migrated information)
Dose descriptor:
LOAEL
Remarks:
(systemic)
Effect level:
20 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: P: Macroscopic and microscopic compound-related changes in the kidneys and bladder ; mortality 60-70% in the high dose group; F1: increased absolute kidney weights of adult females
Remarks on result:
other: Generation: P/F1 (migrated information)
Dose descriptor:
LOEL
Remarks:
(local)
Effect level:
20 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: inflammatory changes in stomachs (it was not possible to conclude they were directly related to maleic anhydride)
VIABILITY (OFFSPRING)/CLINICAL SIGNS (OFFSPRING):
Respiratory rales were observed and the incidence and severity appeared to increase with dose. These rats often vigorously resisted handling at the time of dosing. Some animals were difficult to treat, and gavage-related deaths were observed in F1 adults. More deaths were observed in the F1 generation than in the P/F0 generation. Most of these deaths were attributed to gavage related injuries. If these deaths are omitted than a 0 to 10% mortality was observed in both the low and the mid dose groups and significant mortality was only observed in the high dose group. Due to 100% mortality among the high-dose F1 females by study week 42, the remaining high-dose males were terminated by study week 44.


BODY WEIGHT (OFFSPRING):
- Pup weight at birth: significantly reduced (p<0 .01) in the 150 mg/kg/day group in the F1a litters (-14%). This effect was not observed in the F1b litters.
- Pub weight during lactation: at the 150 mg/kg/day level, inhibition of pup body weight was observed throughout lactation (with statistical significance on days 7, 14 and 21 (-21%) in the F1a litters and after day 7 of lactation in the F1b litters), but there was no consistent pattern between litters in a generation or between generations that suggested a treatment-related effect).
- Parental (F1) body weight: F1 generation showed a similar pattern of depressed weight gain; however, only the F1 males in the high-dose group had significant body weight depression at 30 weeks.
- Mean maternal body weight gain (F1) during gestation in the 20 and 55 mg/kg/day groups (no 150 mg/kg treatment group due to the described mortalities of the female rats) was comparable to the control group during the F2a and F2b matings. The 20 mg/kg/day group exhibited mean maternal body weight gain in excess of the control group during lactation in the F2a mating and mean loss in maternal body weight during lactation in the F2b mating (-24%). Slight reduction of mean maternal body weight gain and no gain in mean maternal body weight were observed during lactation in the F2a and F2b matings respectively, at the 55 mg/kg/day level.


ORGAN WEIGHTS (OFFSPRING): In the F1 generation, the absolute kidney weights of adult females in the low- and mid-dose groups were significantly increased to 108 and to 111%, respectively, of the control value.


GROSS PATHOLOGY (OFFSPRING)/HISTOPATHOLOGY (OFFSPRING):
- F1: Low incidence of mineralization (2/12 males; 3/21 females) and nephrosis (2/12 males ; 5/21 females) in the kidney of animals receiving 150 mg/kg/day. There were no microscopic changes in these kidneys.
- F2: Microscopic examination of tissues from pups in the F2 litters revealed no treatment-related changes. Therefore, treatment up to 55 mg/kg/day for two generations had no adverse effect on pups.

OTHER FINDINGS (OFFSPRING): No treatment-related effects were observed on indices of fertility for males and females in the F1 generation. No adverse effects on litter size and on pup survival were observed at doses up to 150 mg/kg/day in the F1 litters, or at 55 mg/kg/day in the F2 litters.
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
150 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Overall effects
Dose descriptor:
NOAEL
Generation:
F2
Effect level:
55 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Overall effects
Reproductive effects observed:
not specified
Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
108 mg/kg bw/day
Study duration:
chronic
Species:
rat
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

The critical effect of trimellitic anhydride is respiratory sensitisation. This effect has been demonstrated in numerous animal studies at very low exposure concentrations and is also well known as an effect of worker exposure. Safe use of the substance therefore involves minimising worker exposure through the use of engineering controls and personal protective equipment. The sensitising properties of the substance are such that only relatively low exposure concentrations can be used in repeated inhalation studies. Repeated dose oral toxicity studies performed with trimellitic anhydride and the hydrolysis product (trimellitic acid) show very low toxicity, with no evidence of systemic toxicity (including effects on tissues of the reproductive system) at dose levels of up to and including 1000 mg/kg bw/day and findings limited to local effects on the caecum. The substance is therefore clearly of very low systemic toxicity, whereas local effects are seen following inhalation exposure to very low concentrations.

Data for structural homologues of trimellitic anhydride do not suggest the potential for reproduction toxicity. A multi-generation study on maleic anhydride resulted in a NOAEL of 55 mg/kg/day for effects on fertility (corrected to 108 mg/kg/day considering the molecular weight (Mw) differences between the registered substance (Mw 192.13 g/mol) and maleic anhydride (Mw 98.06 g/mol) and bearing in mind that the anhydride group represents the single reactive and toxic functional moiety for both substances), the highest dose tested without complete maternal toxicity. Long-term toxicity tests with another structural analogue, phthalic anhydride, give no indication of effects on reproductive organs to suggest a concern for effects on reproduction. Reprotoxicity screening tests on the structural analogues 4 -methyl hexahydrophthalic anhydride (4 -MHHPA) and methyl tetrahydrophthalic anhydride (MTHPA) again gave no indication of a concern for effects on reproduction.

Considering all the data available, there is no indication that cyclic anhydrides such as TMA may pose a risk with regard to reproductive toxicity and further tests would not be in line with concerns for animal welfare and unnecessary animal experimentation.


Short description of key information:
The available data for structural homologues of trimellitic anhydride indicate neither potential for teratogenic effects nor for reproduction toxicity in different species. These data allow a scientific validated evaluation of the respective endpoints and further tests would not be in line with concerns for animal welfare and unnecessary animal experimentation.

Justification for selection of Effect on fertility via oral route:
Best documented study meeting the requirements for a multi-generation study of potential effects on fertility

Effects on developmental toxicity

Description of key information
One research study report is available.  The study report used pregnant Sprague-Dawley rats and Hartley guinea pigs exposed to the test substance Trimellitic Anhydride via whole body inhalation.  The test animals were exposed for 6 hours a day during the period of major organogenesis.  
Link to relevant study records

Referenceopen allclose all

Endpoint:
developmental toxicity
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well-documented publication and study report which meets basic scientific principles. Reliability adopted from OECD SIDS Draft. For read-across justification see Section 13.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
(administered volume in the control and high dose group is higher (1.4 ml/100 g bw) than the advised maximum volume in the guideline (0.4 ml/100 g bw))
GLP compliance:
no
Species:
rat
Strain:
other: Charles River CD rats
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: 12 weeks
- Housing: individually housed, except during mating
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 10 days


Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
VEHICLE
- Concentration in vehicle: 1% (w/v)
- Amount of vehicle (if gavage): 0.3-1.4 ml/100g bw
Details on mating procedure:
- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1/1
- Proof of pregnancy: vaginal plug in vaginal smear referred to as day 0 of pregnancy
- Any other deviations from standard protocol:
Duration of treatment / exposure:
from gestation day 6 through day 15
Frequency of treatment:
daily
Duration of test:
day 20 of gestation
No. of animals per sex per dose:
25
Control animals:
yes, concurrent vehicle
Maternal examinations:
DETAILED CLINICAL OBSERVATIONS: Yes


BODY WEIGHT: Yes
- Time schedule for examinations: gestation days 0, 6, 9, 12, 15, and 20

Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: foetal swellings, number of viable and non-viable foetuses
Fetal examinations:
- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [1/3 per litter ]
- Skeletal examinations: Yes: [2/3 per litter ]
Statistics:
All statistical analyses compared the treatment groups with the control group, with the level of significance at p<0.05. Male to female foetal sex ratio and number of litters with anomalies were compared using the Chi-square test criterion with Yates correction for 2 x 2 contingency tables and/or Fisher's exact probability test as described by Siegel to Judge significance of differences. The proportion of late resorbed foetuses and postimplantation losses were compared by the Mann-Whitney U-test as described by Siegel and Weil to judge significance of differences. Mean number of corpora lutes, total implantations and viable foetuses were compared by analysis of variance (one-way classification), Bartlett's test for homogeneity of variances and the appropriate t-test (for equal or unequal variances) as described by Steel and Torrie using Dunnett's multiple comparison table to judge significance of differences. Fetal body weights were compared by analysis of variance (hierarchal classification) and t-test as described by Steel and Torri using Dunnett's multiple comparison tables to judge significance of differences.
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
Respiratory involvement and red nasal discharge were observed in all dosage groups. The incidence of these was higher in the treated groups, though not in a dose-related pattern. No treatment-related deaths (one rat in each dosage group died during the first part of treatment; the cause of death was not determined) nor abnormal behaviour was observed in any of the maleic anhydride treated groups. Mean body weight gain was reduced in the 30 mg/kg/day dosage group for the first three days of treatment. There was a slight mean body weight loss in the 90 and 140 mg/kg/day dosage groups for the first three days of treatment. These reductions in weight gains resulted in reduced mean body weight gains over the entire treatment period in all treatment groups compared to the control (however, mean weight of all groups was within 5% of control on days 15 and 20. No biologically meaningful differences in the mean number of viable foetuses, implantations, post implantation losses, corpora lutea, or in the male to female sex ratio between any of the maleic anhydride treated groups and the control group. The general appearance and behaviour of rats were not affected by treatment.
Dose descriptor:
NOAEL
Effect level:
>= 140 mg/kg bw/day
Basis for effect level:
other: maternal toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Mean foetal body weights were lower in the treatment groups than in the control group. This was not considered compound related due to the unusually high mean foetal body weight in this concurrent control group (mean: 4 g). External evaluation, internal examination, and skeletal observations of foetuses from all three treatment groups showed no anomalies in foetal development which could be attributed to maleic anhydride (slight increase in foetal malformations in the 30 (2/23 litters) and 140 mg/kg/day dosage group (3/21 litters) when compared to the control group (1/23 litters) is considered due to random occurrence due to the variety of abnormalities observed).
Dose descriptor:
NOAEL
Effect level:
>= 140 mg/kg bw/day
Basis for effect level:
other: teratogenicity
Dose descriptor:
NOAEL
Effect level:
>= 140 mg/kg bw/day
Basis for effect level:
other: fetotoxicity
Abnormalities:
not specified
Developmental effects observed:
not specified
Conclusions:
It is concluded that the treatment with maleic anhydride up to 140 mg/kg/day did not produce a teratogenic response in CD rats.
Executive summary:

It is concluded that the treatment with maleic anhydride up to 140 mg/kg/day did not produce a teratogenic response in CD rats.

Endpoint:
developmental toxicity
Type of information:
other: Research study
Adequacy of study:
weight of evidence
Study period:
Not documented
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: The study is a research investigation, the methods are scientifically acceptable and the study appears well conducted.
Qualifier:
no guideline followed
Principles of method if other than guideline:
Teratological evaluation of TMA in rats and guinea pigs
GLP compliance:
no
Limit test:
yes
Species:
other: rat and guinea pig
Strain:
other: Sprague Dawley rats and Hartley guinea pigs
Details on test animals and environmental conditions:
The animals were first generation offspring (rats born in-house) from Caesarean derived Sprague-Dawley rats (Charles River Breeding Laboratories, MI) and Hartley guinea pigs (Murphy Breeding Laboratories, IN) were used for this study.
Animals (that had successfully mated) were housed individually in polycarbonate shoe-box cages with corn cob bedding. Animals were transferred to stainless steel wire mesh cages for the 6 hour inhalation exposure periods.
Purina Rodent Chow 5001 or Guinea Pig Chow 5025 (Ralston Purina) and reverse-osmosis purified water were provided ad libitum except during exposure.
The animal rooms were maintained at 22°C and 40% relative humidity. Fluorescent lighting was provided on a 12 hour light/dark cycle.
Route of administration:
inhalation
Type of inhalation exposure (if applicable):
whole body
Vehicle:
unchanged (no vehicle)
Details on exposure:
The animals were split into two groups, the first group were exposed via inhalation to 500 µg/m³ (nominal), while the other group was exposed to filtered air only.
TMA flakes were ground to micron-sized particles using a jet-mill. Respirable-sized particles of TMA were generated in a modified TSI fluidised bed aerosol generator and blown by means of a transvector jet into a 1 m³ inhalation chamber. A minimum flow of 12-15 air changes per hour was maintained in the chamber.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
A Lambda 5 UV/VIS spectrophotometer was used to determine chamber concentrations. The TMA exposure chamber was sampled 2-5 times per day, and the filtered air control chamber was sampled once per day. Samples were collected on glass fibre filters, and extracted with acetonitrile in water. Particle size was determined weekly during the study using a Portable Continuous Aerosol Monitor. The average particle size was 2.73-2.85 microns, and 99.99% of the particles were less than 10 microns.
Details on mating procedure:
The animals were mated two females to one male. Animals were checked daily to determine if mating had occurred; mating was confirmed by the presence of a vaginal plug or sperm positive vaginal smear. The day mating was confirmed was designated as day 0.
Duration of treatment / exposure:
10 exposure days (gestation days 6 through 15)
Frequency of treatment:
6 hours/day for 10 consecutive days (gestation days 6 through 15)
Duration of test:
Rats were sacrificed on gestation day 20 or after weaning. Guinea pigs were sacrificed on gestation day 62 or after weaning.
No. of animals per sex per dose:
27 female rats per dose
15 guinea pigs per dose
Control animals:
yes, sham-exposed
Details on study design:
Approximately one half of the animals in the treated and control groups were sacrificed one day prior to parturition and the offspring were used for teratologic investigation. In the case of the rats sacrifice was gestation day 20, whereas in the guinea pigs it was gestation day 62. The remaining dams were allowed a natural parturition and their offspring were used to evaluate the response (IgG antibody and grossly observed lesions) to a challenge exposure. Dams allowed a natural parturition were sacrificed at the time of neonatal challenge. At the time of challenge, the offspring were subdivided into non-challenge and challenge groups.
Maternal examinations:
Rats were weighed on gestation days 0, 6, 11, 16 and 20. Rats that were allowed a natural parturition were weighed weekly after gestation day 20. Guinea pigs were weighed on gestation days 0, 6, 11, 16, 20, 26 and weekly thereafter.
A gross examination of all maternal organs was performed at sacrifice.
Ovaries and uterine content:
All dams sacrificed prior to parturition underwent a caesarean section. The uterus was removed from each dam, trimmed of excess adherent fat and weighed. The foetuses were removed and examined (see below). The uterus was inspected for tissue resorptions and foetal deaths
Fetal examinations:
Offspring from both species of dams allowed a natural parturition were weighed on day 0 (day of parturition), 5 (day of culling), 7, 14 and at weaning on day 21. Following weaning the offspring were weighed once every two weeks.

The foetuses were removed from the uteri of sacrificed dams. The foetuses were counted, sexed, weighed and given a gross external examination.

One half of each rat's litter was decapitated and utilized for visceral examination. The heads were fixed in Bouins solution and examined separately from the body. A modified Staples' technique was used to examine the thoracic and abdominal viscera, while the heads were serially sectioned and the brain development examined using a modified Wilson's technique. The remaining half of each litter was processed for skeletal evaluation. The same procedures were used for the guinea pigs, however, because of the smaller litter size, all pups underwent a visceral examination and the torso of the decapitated pups were also processed for visceral evaluation.

Rat foetuses were fixed in 70% ethanol for a minimum of one week, gutted, stained with Alizarin Red in 2% aqueous KOH solution for 48 hours and then cleared in glycerin. Guinea pig foetuses were fixed in 70% ethanol for a minimum of one week and then switched to 90% ethanol for an additional week. Prior to staining (with 4% aqueous KOH Alizarin Red solution) and clearing (with glycerin), guinea pig foetuses were macerated for 48 hours in 4% aqueous KOH solution.
Statistics:
A T-test was performed on all data. When sex was a factor, a 2-way analysis of variance (ANOVA) was performed. Maternal body weight data were log-transformed and the data analysed by a multivariate analysis of variance for repeated measures. A mean litter weight was used and not individual foetal or pup weights as litter weights were dependent on the litter size. A minimum significance level of p<0.05 was used in all comparisons. All statistical analyses were performed on a Zenith X-200 PC/AT using SYSTAT.
Indices:
Indices were not calculated.
Historical control data:
No information available.
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
All rats sacrificed on gestation day 20 had lung foci. The mean number of lung foci was 115.9 with the range between 38 and 196. Rats sacrificed following weaning had lower numbers of foci, and only 5 were classed as having a positive response (10 lung foci or more). Rats sacrificed following weaning had a mean of 9.5 lung foci with the range between 0 and 35. The number of external haemorrhagic foci was significantly higher in the TMA-exposed rats relative to control rats at both time points. The number of lung foci in the filtered air control group ranged from 0 - 2 at both sacrifice timepoints.
Two of the seven guinea pig dams sacrificed on gestation day 62 had lung foci (one had 25 foci and the other had 43 foci). None of the guinea pig dams sacrificed following weaning had a positive response with respect to lung foci.
TMA-specific IgG antibody levels were significantly increased at the time of caesarean section and following parturition in the TMA-exposed rats compared to the controls.
Much lower levels of IgG antibody were detected in the guinea pig dams compared to the rat dams at the time of caesarean section and following parturition. Antibody levels were significantly higher than controls at both time points.
Dose descriptor:
LOAEC
Effect level:
500 other: ug/m³ air (nominal)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEC
Based on:
test mat.
Basis for effect level:
other: Overall effects
Remarks on result:
not determinable due to adverse toxic effects at highest dose / concentration tested
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects
Dose descriptor:
NOAEC
Effect level:
500 other: ug/m³ air (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Overall effects
Remarks on result:
other:
Remarks:
No effects seen at the highest exposure concentration
Abnormalities:
not specified
Developmental effects observed:
not specified

No significant differences were detected in the gestating body weights or body weight gains between treated and control dams for either species. No significant differences were detected in gravid uterus weights or in corrected body weights (full-term body weight minus gravid uterus weight) between treated and control dams of either species. No significant differences were detected in the lactating bodyweights between treated and control dams for either species.

No significant differences in body weights were detected between the treated and control foetuses of either species at the time of Caesarean section, parturition, and/or following weaning. No significant differences in litter size or litter viability were detected between the control or treated groups of either species. No significant variations or malformations were observed in the gross external appearance, viscera, skeletal system, or in the development of the brain for either species. None of the foetuses of either species selected for visceral examination had lung foci.

A total of 16 rat pups (exposed in utero) were challenged with TMA. Two male and 3 female rat pups had a positive response of 10 or more lung foci. One male had 7 foci (higher than the concurrent control values). None of the non-challenged pups that were exposed in utero exhibited a positive lung foci response (10 or more). Also, none of the non-challenged filtered air controls had a positive lung foci response. The number of foci was significantly higher in the in utero exposed challenged offspring compared to both the filtered air challenged/non-challenged controls, and also compared to in utero exposed non-challenged pups.

Sixteen in utero exposed rats were challenged as adults with TMA. No treatment-related foci were observed in any of the adult challenged or non-challenged rats. Guinea pigs were not challenged as adults because no response was observed at the time of neonatal challenge.

All TMA in utero exposed rat and guinea pig foetuses had detectable TMA-specific antibody levels. The antibody levels for the majority of the guinea pig foetuses were much lower than rat foetuses. Almost all male and female in utero exposed rat neonates had TMA-specific antibodies. The antibody levels were similar for the TMA in utero exposed challenged and nonchallenged neonates. None of the filtered air controls had antibodies to TMA. None of the guinea pig pups (challenged, non-challenged, TMA-exposed or filtered air controls) sacrificed at approximately 4 weeks of age had TMA-specific antibody. One challenged rat had a barely perceptible level of TMA-specific IgG antibody.

Conclusions:
Under the conditions of this study, there were no treatment-related effects observed in maternal, foetal or offspring body weights or litter viability in the rat or guinea pig. It can therefore be concluded that there were no teratogenic effects of Trimellitic Anhydride observed in either species.
Executive summary:

Trimellitic Anhydride was administered to 27 pregnant rats and 14 pregnant guinea pigs via inhalation at a concentration of 500 µg/m³. A group of similar size was exposed to filtered air and served as a sham control. During the period of major organogenesis, the species were exposed for 6 hours per day. In the rat, the period of major organogenesis was gestation days 6 through 15 and for the guinea pig, it was gestation days 6 through 26. Half of each species per group were used for teratological evaluation while the remaining animals were allowed a natural parturition. Offspring of both species were challenged with a single 6-hour Trimellitic Anhydride exposure.

No treatment-related effects were observed on maternal, foetal or offspring body weights. In addition to this, there were no teratogenic effects or effects on litter viability in either species. No teratogenic effects were seen in either species. Lung foci and TA-specific IgG antibody were observed in dams of both species. TMA-specific IgG antibody was detected in serum from in utero exposed rat and guinea pig foetuses. TMA-specific IgG antibody was found in only the neonatal rats. Lung foci response, of challenged neonatal rat offspring, was variable. Foci were observed in only the challenged offspring whose mothers had not completely recovered from the original TMA exposure. No treatment-related lung foci were observed in neonatal guinea pigs or adult rat offspring.

It can be concluded that exposure to trimellitic anhydride at a concentration of 500 µg/m³ in air during the period of major organogenesis was not teratogenic to rats and guinea pigs.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
274 mg/kg bw/day
Study duration:
subacute
Species:
rat
Effect on developmental toxicity: via inhalation route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
500 µg/m³
Study duration:
subacute
Species:
rat
Effect on developmental toxicity: via dermal route
Endpoint conclusion:
no study available
Additional information

Trimellitic Anhydride (TMA) was administered to 27 pregnant rats and 14 pregnant guinea pigs via inhalation at a concentration of 500 µg/m³ (Ryan, 1988). A group of similar size was exposed to filtered air and served as a sham control. During the period of major organogenesis, the species were exposed for 6 hours per day. In the rat, the period of major organogenesis was gestation days 6 through 15 and for the guinea pig, it was gestation days 6 through 26. Half of each species per group were used for teratological evaluation while the remaining animals were allowed a natural parturition. Offspring of both species were challenged with a single 6-hour Trimellitic Anhydride exposure. No treatment-related effects were observed in maternal, foetal or offspring body weights. In addition to this, there were no teratogenic effects or effects on litter viability in either species. No teratogenic effects were seen in either species. Lung foci and TA-specific IgG antibody were observed in dams of both species. TMA-specific IgG antibody was detected in serum fromin uteroexposed rat and guinea pig foetuses. TMA-specific IgG antibody was found in only the neonatal rats. Lung foci response, of challenged neonatal rat offspring, was variable. Foci were observed in only the challenged offspring whose mothers had not completely recovered from the original TMA exposure. No treatment-related lung foci were observed in neonatal guinea pigs or adult rat offspring. It can be concluded that exposure to trimellitic anhydride at a concentration of 500 µg/m³ in air during the period of major organogenesis was not teratogenic to rats and guinea pigs. Although the value of the study is limited by the relatively low exposure concentration, it clearly shows an absence of developmental toxicity in the presence of maternal toxicity.

 

Data on structural analogues of the substance do not suggest a concern for developmental toxicity. The developmental toxicity of maleic anhydride has been assessed in a rat study very similar to the current OECD 414 guideline. The substance was administered at doses of 0, 30, 90, 140 mg/kg bw/day to female rats (25 per dose) for 20 days during gestation. The following observations were made: Observed maternal toxicity included respiratory involvement and red nasal discharge in all dosage groups. The incidence of these was higher in the treated groups, though not in a dose-related pattern. No treatment-related deaths (one rat in each dosage group died during the first part of treatment; the cause of death was not determined) nor abnormal behaviour was observed in any of the maleic anhydride treated groups. Mean body weight gain was reduced in the 30 mg/kg bw/day dosage group for the first three days of treatment. There was a slight mean body weight loss in the 90 and 140 mg/kg bw/day dosage groups for the first three days of treatment. These reductions in weight gains resulted in reduced mean body weight gains over the entire treatment period in all treatment groups compared to the control (however, mean weight of all groups was within 5% of control on days 15 and 20. No biologically meaningful differences in the mean number of viable foetuses, implantations, post implantation losses, corpora lutea, or in the male to female sex ratio between any of the maleic anhydride treated groups and the control. The general appearance and behaviour of rats were not affected by treatment. No embryotoxic effects were observed and it was concluded that treatment with maleic anhydride up to 140 mg/kg bw/day did not produce a teratogenic response in rats. This results in a corrected NOAEL of 274 mg/kg/day considering the molecular weight (Mw) differences between the registered substance (Mw 192.13 g/mol) and maleic anhydride (Mw 98.06 g/mol) and bearing in mind that the anhydride group represents the single reactive and toxic functional moiety for both substances. Developmental toxicity of another structural analogue, phthalic acid, was also assessed in rats in a method similar to OECD 414. Pregnant rats were given phthalic acid in the diet at a dose of 0 (control), 1.25 % (1021 mg/kg bw), 2.5 % (1763 mg/kg bw) or 5.0 % (2981 mg/kg bw). The rats receive the compound daily during the gestation day 7 – 16. The pregnant rats were observed daily for evidence of clinical signs of toxicity. The pregnant rats were sacrificed on day 20 of pregnancy. The peritoneal cavity and uterus were opened and the numbers of live and dead foetuses and resorptions were counted. The live foetuses removed from the uterus were sexed, weighed and inspected for external malformations and malformations within the oral cavity. In the higher dose group significant decreases in the weight of foetuses and number of ossification centres of the caudal vertebrae were found, indicative for a mal-supply of the offspring due to pronounced maternal toxicity. As the NOAEL for teratogenicity was already well above the limit value of the respective OECD guideline and effects were noted only at very high doses linked with marked maternal toxicity, it can be concluded that there is no indication for direct teratogenic effects of the substance in this study. The determined NOAEL was found to be 1000 mg/kg bw/day for maternal toxicity and a NOAEL of 1700 mg/kg bw/day for teratogenicity was derived, corrected to 2205 mg/kg/day considering the molecular weight (Mw) differences between the registered substance (Mw 192.13 g/mol) and phthalic anhydride (Mw 148.12 g/mol).

 

Considering all the data available, there is no indication that cyclic anhydrides such as TMA may pose a risk with regard to reproductive toxicity and further tests would not be in line with concerns for animal welfare and unnecessary animal experimentation. Respiratory sensitisation is seen at exposure levels orders of magnitude lower than the exposure concentrations used in the studies described and this effect of exposure will drive the human risk assessment. It can be concluded that, as occupational exposure to the substance will be limited by its potential to cause respiratory sensitisation, there is no potential for developmental toxicity.


Justification for selection of Effect on developmental toxicity: via oral route:
Best documented study meeting the requirements for a developmental toxicity study

Justification for classification or non-classification

Classification is not justified based on the available information on the substance itself and from the lack of findings in structurally related substances.