Mesitylene

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records

Reference

Endpoint:

multi-generation reproductive toxicity

Remarks:

based on test type (migrated information)

Type of information:

migrated information: read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Near guideline study. Published in peer reviewed literature. Minor restrictions in design and/or reporting but otherwise adequate for assessment.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)

Deviations:

yes

Remarks:

Pre 2001 Guideline.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

other: Charles River COBS CD

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Portage, MI, USA
- Age at study initiation: F0, 8-9 weeks old; F1, 5-7 weeks old; F2, 5-7 weeks old; F3, 22 days old.
- Weight at study initiation: not reported
- Fasting period before study: none
- Housing: individually housed in wire mesh cages throughout the study, except during mating period when males and females co-habited (1:1).
- Housing during parturition : between gestational day (GD) 20 and lactational day (LD) 4 , pregnant dams were individually housed in plastic cages with wood chips supplied for nesting materials.
- Diet: Purina Certified Rodent Chow No. 5002 ad libitum except during exposures.
- Water : provided ad libitum - no other details
- Acclimation period: parents - 2-3 weeks


ENVIRONMENTAL CONDITIONS
- Temperature (°C): not reported
- Humidity (%): not reported
- Air changes (per hr): not reported
- Photoperiod (hrs dark / hrs light): not reported


IN-LIFE DATES: From: To: not reported

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

unchanged (no vehicle)

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 16 cubic metre glass and stainless steel chambers
- Method of holding animals in test chamber: not specified
- Source and rate of air: chamber air supplied by HVAC system, separate from general laboratory air handling system
- Method of conditioning air: HVAC system, particulate-filtered and controlled for temperature and humidity.
- System of generating particulates/aerosols: test sample was vapourised with nitogen through a heated glass column. Vapours diluted with chamber ventilation air to achieve desired exposure levels
- Temperature, humidity, pressure in air chamber: monitored every 30 minutes
- Air flow rate: not reported
- Air change rate: not reported
- Treatment of exhaust air: not reported

TEST ATMOSPHERE
- Brief description of analytical method used: gas-phase infrared spectrometer
- Samples taken from breathing zone: not specified

Details on mating procedure:

30 males and females per group were co-housed (1:1) for a 2 week mating period from each of the 3 generations.
F2 parents : due to toxicity, all surviving animals in the high dose group were mated, to produce the F3 generation.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Exposure levels were monitored with a gas-phase infrared spectrophotometer at hourly intervals throughout the study. The accuracy of this method was confirmed by the use of vapour standards. Additionally, the composition (on a weight percentage basis) of the test material within the exposure chamber was determined by gas chromatography during the first exposure week.

Duration of treatment / exposure:

6 hours/day

Frequency of treatment:

F0 males : 6 hours per day, 5 days per week for 10 weeks before mating and 5 days/week during the 2 week mating period.
F0 females: 6 hours per day, 5 days per week for 10 weeks before mating, 5 days per week during mating, daily during preganancy (gestational day 0 to 20) and daily from postnatal day 5 (LD5) until weaning (LD 21).
F1 males : no exposure after weaning until 5-7 weeks of age then 6 hours per day, 5 days per week for 10 weeks before mating and 5 days/week during the 2 week mating period.
F1 females: no exposure after weaning until 5-7 weeks of age then 6 hours per day, 5 days per week for 10 weeks before mating, 5 days/week during mating, daily during pregnancy (gestational day 0 to 20) and daily from postnatal day 5 (LD5) until weaning (LD 21).
F2 males : From postnatal day 22, 6 hours per day, 5 days per week for 10 weeks before mating and 5 days/week during the 2 week mating period.
F2 females: From postnatal day 22, 6 hours per day, 5 days per week for 10 weeks before mating, 5 days/week during mating, daily during pregnancy (gestational day 0 to 20) and daily from postnatal day 5 (LD5) until weaning (LD 21).

Details on study schedule:

- Age at start of 10 week exposure period: F0, 8-9 weeks; F1 , 5-7 weeks; F2, 22 days.
- Random selection of parents from F1 generation (30/sex/group) was done approximately 1 week after all had completed weaning on LD21.
The study was extended to a 3rd generation but exposure of the F2 animals used to produce the F3 generation began immediately after weaning (LD 22). Due to concerns about toxicity, 40 animals/sex/group were initally randomised and exposed. 30 animals/sex/group were then randomly selected for mating in the control, 100 and 500ppm groups. Due to the very high number of deaths in the first week at 1500ppm, all surviving animals were mated to produce the F3 pups.
- Age at mating of the mated animals in the study: F0 ,18-19 weeks; F1 , 15-17 weeks; F2, 14 weeks .

Remarks:

Doses / Concentrations:
0, 100, 500 and 1500 ppm
Basis:
other: Target concentrations

Remarks:

Doses / Concentrations:
0, 103 ± 2.1,495 ± 8.0, 1480 ± 20.5 ppm
Basis:
analytical conc.

No. of animals per sex per dose:

Exposure groups: 3 groups plus control group.

F0: 30 males caged with 30 females/group;
F1: 30 males caged with 30 females /group;
F2: 30 males caged with 30 females in control, low and mid dose groups, all survivors (4 males and 6 females) mated in high dose group.

Control animals:

yes, sham-exposed

Details on study design:

- Dose selection rationale: dose levels based on previous developmental and reproductive toxicity studies in rats of Aromatol, a branded product conforming to the specifications of High Flash Aromatic Naphtha.

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly

BODY WEIGHT: Yes
- Time schedule for examinations: recorded weekly until confirmation of mating.
Females were also weighed on gestational days 0, 7, 14 and 21 and lactational days 0, 7, 14 and 21.

FOOD CONSUMPTION
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No
Food consumption was measured weekly except during mating, gestation and lactation.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No
- Time schedule for examinations:


OTHER:

Oestrous cyclicity (parental animals):

No information

Sperm parameters (parental animals):

No information

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of 8 pups/litter (4/sex/litter as nearly as possible); excess pups were killed and discarded.

PARAMETERS EXAMINED
The following parameters were examined in F1 / F2 / F3 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain.

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities; possible cause of death was not determined for pups born or found dead.

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals at the end of the mating period.
- Maternal animals: All surviving animals after weaning.

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

HISTOPATHOLOGY / ORGAN WEIGHTS
The following organs were weighed : epididymides, lung, ovary, testis, prostate/seminal vesicle and uterus/vagina.
The following organs and tisses were submittted : epididymides, lung, ovary, pituitary, prostate, seminal vesicle, testis, uterus, vagina and regional lymph nodes and were examined microscopically in the control and high dose groups only. Any masses and gross lesions in any groups were examined .

Postmortem examinations (offspring):

SACRIFICE
- The F1 and F2 offspring not selected as parental animals were sacrificed at 4 days of age.
- These animals and any that died during the lactation period were necropsied and examined for anomalies.

GROSS NECROPSY
- Gross necropsy for anomalies - no further details

Statistics:

Fertility indices and male to female sex ratios were compared using the Chi-square test criterion with Yates' correction for 2x2 contingency tables. Parental body weights, maternal body weight changes and organ weights were compared by analysis of variance (one-way), Bartlett's test for homogeneity of variance and the appropriate t-test (for equal or unequal variance) using Dunnett's multiple comparison tables. Mean numbers of liveborn pups per litter and mean pup weight were compared by analysis of variance and the appropriate t-test.

Reproductive indices:

Female mating index, female conception index, female gestation index, male fertility index, cohabitation time required for mating, litter size at birth.

Offspring viability indices:

Gestation survival index, postnatal survival index for days 4 and 21.

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:

effects observed, treatment-related

Reproductive function: oestrous cycle:

not specified

Reproductive function: sperm measures:

not specified

Reproductive performance:

effects observed, treatment-related

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
F0 females - at 1500ppm, 7 females died or were killed - 3 prior to mating, 3 during gestation and 1 during lactation.
F1 males & females - at 1500ppm ataxia was seen in 18 males and 23 females and reduced motor activity was seen in 11 males and 8 females.
F1 females - at 1500ppm, 6 females died - 3 during exposure, 1 during gestation , 1 during delivery and 1 on day 2 of lactation. One female died in each of the other 3 groups.
F2 males & females - exposures were initiated on day 22 and most animals at 1500ppm (36/40 males and 34/40 females) died within the first week.
BODY WEIGHT (PARENTAL ANIMALS)
F0 males - body weight gain was significantly reduced at 500ppm (by (5-7%) and at 1500ppm (by 14-16%).
F0 females- body weight gain was significantly reduced at 500ppm (by (5-7%) and at 1500ppm (by 5-7%).
F1 males & females - mean body weights were less than controls (by 17% and 12%, respectively) at the start of exposure. During exposure, bodyweight gain was reduced by 5-7% in males at 500ppm, 21-25% in males at 1500ppm and 9-14% in females at 1500ppm.
F2 males & females - at 1500ppm, body weights of survivors were 40% & 35% (males/females) below controls in week 4 and remained 31-38% / 21-30% (males /females) below controls for the remainder of the exposure period.
F2 males& females - at 500 and 100ppm, body weights were initially lower than controls and remained at similar levels throughout the exposure period.
FOOD CONSUMPTION (PARENTAL ANIMALS)
There were no effects on food consumption at any exposure level in any of the generations.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
Male fertility was statistically significantly lower (p< 0.05) at the 1500ppm group in the F1 generation only.

Key result

Dose descriptor:

NOAEC

Effect level:

500 ppm (nominal)

Sex:

male/female

Basis for effect level:

other: Based on severe systemic toxicity at higher dose level. No consistent evidence of reproductive toxicity in the presence of systemic toxicity.

Remarks on result:

other: Generation: F0, F1, F2, F3 (migrated information)

Key result

Dose descriptor:

NOAEC

Effect level:

ca. 500 ppm (nominal)

Sex:

male/female

Basis for effect level:

other: Based on severe systemic toxicity at higher dose level. No consistent evidence of reproductive toxicity in the presence of systemic toxicity.

Clinical signs:

no effects observed

Mortality / viability:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Sexual maturation:

not specified

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

not examined

Histopathological findings:

not examined

VIABILITY (OFFSPRING)
F2 - the mean number of live offspring/litter and number of live births/number delivered were reduced in the 1500ppm group but this was associated with unconfirmed matings. Where mating was not confirmed, exposure of the dams continued until delivery.
The fraction of liveborn offspring was slightly but significantly reduced in the high dose group but survival throughout the weaning period was similar to controls.
F3 - there was little evidence of reproductive toxicity even at 1500ppm, even though there were only 6 litters for evaluation.

CLINICAL SIGNS (OFFSPRING)
No effects.

BODY WEIGHT (OFFSPRING)
F2 - mean body weights were lower in 1500ppm group from LD7 throughout the rest of the weaning period.
F3- mean bodyweights at 1500ppm were significantly lower at birth but there were no significant differences at LD4. The reduced birth weight may have been a result of the small size of the dams.

Key result

Dose descriptor:

NOAEC

Generation:

F1

Effect level:

ca. 500 ppm (nominal)

Sex:

male/female

Basis for effect level:

other: Based on severe systemic toxicity at higher dose level. No consistent evidence of reproductive toxicity in the presence of systemic toxicity.

Key result

Dose descriptor:

NOAEC

Generation:

F2

Effect level:

ca. 500 ppm (nominal)

Sex:

male/female

Basis for effect level:

other: Based on severe systemic toxicity at higher dose level. No consistent evidence of reproductive toxicity in the presence of systemic toxicity.

Reproductive effects observed:

not specified

Conclusions:

Exposure to analysed concentrations of 1500 ppm of High Flash Aromatic Naphtha caused toxicity but no consistent evidence of reproductive toxicity.

Executive summary:

In the first generation parental body weights were significantly reduced at 1500 ppm but reproductive parameters were unaffected. There were no differences in litter size, mean birth weight, or postnatal survival. Once maternal exposures were reinitiated (at day 4L), offspring in the 1500 ppm group gained weight more slowly than controls.

 

In the second generation parental body weight gain was again reduced at 1500 ppm and, male fertility was lower. Litter size, birth weight, and postnatal survival appeared to be reduced in the 1500 ppm exposure group although this was affected by the inclusion of several litters exposed beyond GD20, until just prior to birth. In these litters, the frequency of live births and mean birth weight was reduced and survival early in lactation was poor. Litters from dams exposed to 1500 ppm to GD20 pup survival was 92.5% compared with 98.7% in controls.

 

For the third generation, exposure of each rat was continuous (i.e. there was no exposure-free period between weaning and commencement of exposure as in previous generations). In the 1500 ppm group, 88% of the rats died. There was no evidence of fertility or reproductive effects although only 6 litters were available for evaluation at 1500 ppm. Birth weights were lower in the 1500 ppm group although mean body weight at LD4 was not different from control. The toxicity of 1500 ppm in this generation precludes definitive evaluation of the potential for reproductive effects and clarification of the results obtained in the previous generation.

Effect on fertility: via oral route

Endpoint conclusion:

no study available

Effect on fertility: via inhalation route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

2 500 mg/m³

Study duration:

subchronic

Species:

rat

Quality of whole database:

The available data provide information that is adequate for the purpose of hazard assessment

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

A three-generation reproduction/fertility study (McKee et al., 1990) using C9 aromatic naphtha (ASTM D-3734, Type 1) as the test substance, was conducted prior to current guidelines. Thirty rats/sex/dose level) were exposed via inhalation to 0, 100, 500, or 1500 ppm for 6 hours/day for 5 days/week for 10-12 weeks prior to mating and during the 2-week mating period. The males were exposed thereafter until termination but the F0 and selected F1 generation female rats were exposed until day 20 of gestation and then from postnatal day 5 until weaning at day 21. Exposure of the selected F1 and F2 offspring commenced approximately 1 week after all pups were weaned and from the day of weaning for selected F3 offspring.

 

The highest concentration of 1500 ppm was associated with the premature death of several females:3 F0 females died or were killed prior to mating, 3 during gestation and 1 during lactation; 3 F1 females died prior to mating, 1 during gestation, 1 during delivery and 1 on day 2 of lactation. Exposure of the F2 animals was continuous from postnatal day 5 and 88% died within the first week of exposure post-weaning, although a few did survive the duration of the study. The exposure concentration of 1500 ppm was considered to be in excess of the maximum tolerated dose for rats and therefore inappropriate for the evaluation of reproductive toxicity especially for the F2/F3 generations.

 

For the F0 generation,a dose-related reduction in body weight gain was observed in males and females exposed to 500 or 1500 ppm but reproductive parameters were unaffected. There were no differences in litter size, mean birth weight, or postnatal survival but the F1 offspring exposed to 1500 ppm group gained weight more slowly than controls.

 

For the selected F1 generation,a dose-related reduction in body weight gain was observed in males and females exposed to 500 or 1500 ppm; the effect at 1500 ppm was greater than seen previously in the F0 generation and clinical observations of ataxia and reduced motor activity were also observed indicating a more severe response to exposure. The fertility of the selected F1 males was reduced and the lack of detection of mating in approximately one third of the females resulted in exposure continuing until just prior to delivery. This extended exposure is considered by the authors to have caused reductions in the frequency of live births, mean birth weight and neonatal survival. However, evidence of reduced neonatal survival was also observed in litters whose mothers were exposed to 1500 ppm until day 20 of gestation.

 

For the selected F2 generation the severity of the toxicity due to 1500 ppm precludes meaningful evaluation of the reproductive data and, as a consequence, the significance of the observation of reduced male fertility and neonatal survival in the F1 generation exposed to this concentration cannot be put into context. Given the increased severity of the effect of 1500 ppm on the parental F1 generation compared to the F0 generation, the possibility of an effect of 1500 ppm on male fertility and neonatal survival cannot be dismissed.

 

On the basis of the results of this study, the NOAEC for reproductive toxicity is 500 ppm (equivalent to 2500 mg/m³ according to Firth, 2008) although an effect of the higher concentration of 1500 ppm on reproduction has not been clearly established due to the overt toxicity seen. The NOAEC for systemic toxicity is 100 ppm (equivalent to 500 mg/m³ according to Firth, 2008).  The test material contained a mixture of aromatics with similar molecular weight to trimethylbenzene and are assumed to have a similar toxicology profile therefore no adjustment is made to the NOAEC to account for the specific trimethylbenzene content.

Short description of key information:

In a 3-generation study using the inhalation route of exposure, a NOAEC of 2500 mg/m3 for reproductive toxicity was determined and a NOAEC of 500 mg/m3 for systemic toxicity.  

Justification for selection of Effect on fertility via inhalation route:

Results from a 3-generation reproduction study indicate a NOAEC of 500 mg/m3 for systemic (parental) toxicity and a NOAEC of 2500 mg/m3 for reproductive toxicity.

Effects on developmental toxicity

Description of key information

Two inhalation prenatal developmental toxicity studies have been considered, one using rats and the other mice.  The most recent study conducted was reported.  It was well conducted to current guidelines and used the rat as the test species; a NOAEC of 1470 mg/m3 was determined for maternal and developmental toxicity.

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP status unknown, guideline study, published in peer reviewed literature, no restrictions, fully adequate for assessment.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: IFFA CREDO Breeding Laboratories (Saint-Germain-sur-l'Arbresle, France)
- Details: nulliparous females
- Weight at study initiation: 180-200 g
- Fasting period before study: none
- Housing: Mated females were individually housed in clear polycarbonate cages with stainless-steel wire lids and corn cob granules as bedding
- Diet: Food pellets (UAR Alimentation Villemoisson, France) ad libitum except during exposure
- Water: filtered tap water ad libitum except during exposure
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 21±2°C
- Humidity: 50±5%
- Air changes (per hr): Not reported
- Photoperiod: 12hrs dark / 12hrs light

IN-LIFE DATES: Not reported

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

other: air

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 200 L glass/stainless steel inhalation chambers
- Chambers were maintained at a negative pressure of no more than 3 mm water. The concentrations were adjusted by varying the airflow passing through the fritted of the bubbler.
- System of generating test atmosphere: vapours were generated by passing an additional airflow rate through the fritted disk of a heated bubbler containing the test chemical. The vaporized compound was carried out into the main air inlet pipe of the exposure chambers.
- Temperature, humidity, pressure in air chamber: Temperature 23±0.6°C; relative humidity 57±5%.
- Air flow rate: Dynamic/adjustable laminar air flow (5-10 m3/h).

TEST ATMOSPHERE
- Brief description of analytical method used: Actual concentrations measured once daily by collecting atmosphere samples through glass tubes packed with activated charcoal.
- Mesitylene was desorbed with carbon disulfide and analysed by a gas chromatograph using ethylbenzene as internal standard.
- Samples were chromatographed on a column full of PEG 20 M and 2m length, which was maintained at a temperature of 100°C. Another gas chromatograph was used for the continuous monitoring of the levels of exposure; this latter chromatograph was equipped with a flame ionization detector and an automatic gas-sampling valve. Concentration measurements were performed at regular intervals (0.5 min).
- The presence of liquid particles was evaluated at the highest concentration generated (i.e. 1200 ppm), due to mesitylene having a low vapour pressure (2.31 mm Hg at 22°C).
- Airborne particles were measured with an optical counter of particles with a lower detection limit of 0.75 µm.
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The actual concentrations (±SD) (from charcoal tubes) were 101 (±4), 299 (±10), 599 (±11), and 1194 (±29) ppm of mesitylene. Intra-day variations were <4.3%. No difference in particles counts was observed between the clean filtered air (control) and vapour-laden air in the exposure chamber

Details on mating procedure:

- Impregnation procedure: co-housed
- M/F ratio per cage: 1 male : 2-3 females
- Length of cohabitation: overnight until evidence of mating
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy

Duration of treatment / exposure:

15 days

Frequency of treatment:

6 h/day, on days 6 through 20 of gestation

Duration of test:

Following acclimatisation and mating, the study lasted from day 0-21 of gestation

Remarks:

Doses / Concentrations:
0, 100, 300, 600 and 1200 ppm
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
0, 492, 1470, 2950 and 5900 mg/m3
Basis:
nominal conc.
based on 1 ppm = approx 4.92 mg/m3 (Korsak et al, 1997)

Remarks:

Doses / Concentrations:
101±4, 299±10, 599±11 and 1194±29 ppm
Basis:
analytical conc.

No. of animals per sex per dose:

24-25 females (17-24 pregnant)

Control animals:

yes, sham-exposed

Details on study design:

- Dose selection rationale: Based on results from:
(i) McKee et al, 1990. 3-generation reproduction study , rats exposed to vapours of a C9 aromatic hydrocarbon mixture containing significant proportions of trimethylbenzenes (corresponding to c.a. 8.7, 42, and 124 ppm mesitylene). No effects on male and female fertility. Offspring growth affected at the mid and high exposure concentrations. Mice also exposed to same C9 fraction (i.e. 8.6, 42, or 127 ppm mesitylene). The high concentration caused maternal toxicity and evidence of developmental toxicity.

(ii) Ungvary et al, 1983. A decrease in the body weight of male foetuses and a foetal ossification delay were reported following inhalation exposure of pregnant rats to a C9 aromatic hydrocarbon mixture (containing approximately 18-36 ppm of mesitylene) from day 7 to 15 of gestation.

(iii) Lehotzky et al, 1985. In a behavioural developmental toxicity study, rats were exposed to 600, 1000, or 2000 mg/m3 aromatol (a mixture of methyl-ethyl benzenes and trimethylbenzenes). No significant effects were observed in dams and offspring.

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Not reported

DETAILED CLINICAL OBSERVATIONS: No data
BODY WEIGHT: Yes
- Time schedule for examinations: gestation days 0, 6, 13 and 21

FOOD CONSUMPTION: Yes
- Time schedule: gestation day periods 6-13 and 13-21

WATER CONSUMPTION: No

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: Uterus

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

Fetal examinations:

- External examinations: Yes: all per litter (including the oral cavity)
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter

Statistics:

Number of corpora lutea, implantation sites, live foetuses, body weights: one-way analysis of variance, followed by Dunnett's test if differences were found.
Post-implantation loss, dead foetuses, resorptions, alterations among litters: Kruskal-Wallis test followed by the Mann-Whitney test where appropriate.
Pregnancy rate, incidences of foetal alterations: Fisher's test, where applicable, least-squares analysis was carried out. The litter was used as the basis for the analysis of foetal variables.

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
- Mortality: No deaths
- Clinical signs: No treatment-related effects
- Bodyweight: Maternal body weight gain significantly reduced during the second half of exposure at 300 ppm (1470 mg/m3), and throughout the whole exposure period at 600 and 1200 ppm (5900 mg/m3). When body weight gains were corrected for gravid uterine weights, weight gain over the exposure period was significantly reduced at 600 ppm (2950 mg/m3), and weight loss occurred at 1200 ppm (5900 mg/m3).
- Food consumption: Significant decrease throughout exposure at 300 ppm and above.
- The mean numbers of implantation sites and of live foetuses, and the incidences of post-implantation loss and resorptions were comparable among groups

Dose descriptor:

NOAEC

Effect level:

492 mg/m³ air (nominal)

Basis for effect level:

other: maternal toxicity

Dose descriptor:

NOAEC

Effect level:

1 470 mg/m³ air (nominal)

Basis for effect level:

other: developmental toxicity

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
- Foetal body weight: There was a concentration-related decrease which achieved statistical significance at 600 ppm in males only, and at 1200 ppm in males, females and males and females combined. Mean bodyweights were 5-7% less than control at 600 ppm and 12% less than control at 1200 ppm (2950 and 5900 mg/m3 respectively).
- Malformations: Few malformations were observed. One control foetus showed multiple skeletal malformations. Diaphragmatic hernia was seen in one foetus of the 100 ppm group and in one foetus of the 600 ppm (2950 mg/m3) group. Common external and visceral variations were found, but were considered not to be treatment-related. The incidence of foetuses with incomplete sternebral ossification was slightly, but not significantly, elevated at 1200 ppm (5900 mg/m3).

Dose descriptor:

NOAEC

Effect level:

5 900 mg/m³ air (nominal)

Basis for effect level:

other: teratogenicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Conclusions:

The NOAEC for maternal toxicity was 100 ppm (492 mg/m3) and the NOAEC for developmental toxicity was 300 ppm (1470 mg/m3).

Executive summary:

Pregnant rats were exposed whole body to vapours of mesitylene (0, 100, 300, 600 and 1200 ppm; 0, 492, 1470, 2950 and 5900 mg/m³) 6 h/day, on gestational days 6 through 20. A significant decrease in maternal body weight gain and food consumption was observed at concentrations of 300 ppm mesitylene or greater. Foetal toxicity, expressed as significant reduction in foetal body weight, occurred at 600 and 1200 ppm mesitylene. There was no evidence of embryolethal or teratogenic effects following inhalation. In summary, the NOAEC for maternal toxicity was 100 ppm and the NOAEC for developmental toxicity was 300 ppm.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

1 470 mg/m³

Quality of whole database:

The available data provide information that is adequate for the purpose of hazard assessment

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

A well conducted developmental toxicity study to current guidelines has evaluated the exposure of rats to 1,3,5-trimethylbenzene by inhalation (Saillenfait et al., 2005). Female rats were exposed to 0, 100, 300, 600 or 1200 ppm for 6 hours/day on days 6 – 20 inclusive, of gestation (where day 0 was the day of confirmation of mating and day 21 the day of termination). Reduced maternal body weight gain and food consumption were associated with exposure to 600 ppm and to 1200 ppm; the NOAEC was 300 ppm. Foetal body weights were also reduced following maternal exposure to 600 or 1200 ppm 1,3,5-trimethylbenzene but there was no evidence for an effect of treatment on foetal development and no evidence of teratogenicity. The reduction in foetal body weight was seen in the presence of an effect of 1,3,5-trimethylbenzene on maternal body weight gain. Thus, the NOAEC of 1,3,5 -trimethylbenzene for maternal and developmental toxicity in the rat was 300 ppm (1470 mg/m³).

An earlier developmental toxicity study (McKee et al., 1990) using C9 aromatic naphtha (ASTM D-3734, Type 1) as the test substance, was conducted prior to current guidelines. This study used mice as the experimental model and females were exposed to 0, 100, 500 or 1500 ppm for 6 hours/day on days 6 - 15 inclusive, of gestation (where day 0 was the day of confirmation of mating and day 18 the day of termination).

The exposure concentration of 1500 ppm was found to be in excess of the maximum tolerated dose for mice and 44% maternal lethality was caused; this concentration of 1,2,4-trimethylbenzene was therefore considered inappropriate for the evaluation of developmental toxicity in mice.

Maternal body weight gain was reduced during exposure to 500 ppm on gestation days 6 to 15 inclusive; foetal body weight was also reduced. However, there was no evidence for an effect of the test substance on foetal development and no evidence of teratogenicity. Thus, the NOAEC ofC9 aromaticnaphtha (ASTM D-3734, Type 1) for maternal and developmental toxicity in the mouse was 100 ppm (equivalent to 500 mg/m³according to Firth, 2008).  The test material contained a mixture of aromatics with similar molecular weight to trimethylbenzene and are assumed to have a similar toxicology profile therefore no adjustment is made to the NOAEC to account for the specific trimethylbenzene content.

As 1,3,5-trimethylbenzene is structurally similar to C9 aromatic naphtha (ASTM D-3734, Type 1) and is considered to have similar toxicity the results of these two studies can be used to confirm a highest NOAEC of 300 ppm /1470 mg/m³.

Justification for selection of Effect on developmental toxicity: via inhalation route:

Studies in rats and mice indicate that 1,2,4-trimethylbenzene is not selectively toxic to the foetus

Justification for classification or non-classification

1,3,5-Trimethylbenzene is not toxic to reproduction and has no effect on fertility or development. Consequently, it does not warrant classification under Regulation (EC) No 1272/2008.