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Diss Factsheets

Administrative data

Description of key information

Oral NOAEL (Rat): ≥ 1056 mg/kg (1.28 mL/kg) (similar to OECD TG 408)

 

Inhalation NOAEC (Rat): 690 ppm (3950 mg/m3) (similar to OECD TG 413)

 

Dermal NOAEL (Rat): ≥ 495 mg/kg (similar to OECD TG 410)

Key value for chemical safety assessment

Toxic effect type:
dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
sub-chronic toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1984
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Basic data given:comparable to guidelines/standards.
Justification for type of information:
The justification for read across is provided as an attachment in IUCLID Section 13.
Reason / purpose for cross-reference:
read-across: supporting information
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
- Concentration in vehicle: constant volume dosage of 5 ml/kg bw
Duration of treatment / exposure:
30 days
Frequency of treatment:
daily
Remarks:
Doses / Concentrations:
0.14 ml/kg/day (~116 mg/kg bw)
Basis:
other: nominal
Remarks:
Doses / Concentrations:
0.42 ml/kg/day (~347 mg/kg bw)
Basis:
other: nominal
Remarks:
Doses / Concentrations:
1.28 ml/kg/day (~1056 mg/kg bw)
Basis:
other: nominal
No. of animals per sex per dose:
5 female/5 male
Control animals:
yes, concurrent vehicle
Sacrifice and pathology:
GROSS PATHOLOGY: Yes, organs examined include kidneys and livers.
HISTOPATHOLOGY: Yes, organs examined include kidneys.
Other examinations:
Clinical chemistry- including plasma glucose
hematology - including lymphocyte and platelet counts, cell volume, hemoglobin concentration, and erythrocyte counts;
Urinanalysis - including protein concentrations
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
no effects observed
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
effects observed, treatment-related
Clinical biochemistry findings:
effects observed, treatment-related
Urinalysis findings:
effects observed, treatment-related
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
not examined
Details on results:
CLINICAL SIGNS AND MORTALITY
The majority of animals in the 0.42 and 1.48 ml/kg/day groups showed salivation and/or brown facial staining from day 4 onwards, as did three animals in the 0.14 ml/kg/day group. Salivation was normally for a short period, and the staining resolved within 24 hrs.

HAEMATOLOGY
Males rats in the 1.28 ml/kg/day group showed higher lymphocyte and platelet numbers, and slightly lower packed cell volume, hemoglobin concentration and erythrocyte counts.

CLINICAL CHEMISTRY
Plasma glucose levels of rats in the 1.28 ml/kg/day group were lower than controls.

URINALYSIS
Urinary protein concentrations were higher in all male rats in the two higher dose groups, and in 2 males in the lowest dose group.

ORGAN WEIGHTS
Male rats showed a dosage related increase in liver and kidney weights. Female rats only showed higher liver weight at the highest dose level.

GROSS PATHOLOGY
One male rat in the 1.28 ml/kg/day dose group had occasional cystic spaces in the parenchyma of the left kidney.

HISTOPATHOLOGY: NON-NEOPLASTIC
The changes in the kidneys were a slight degeneration of the cells lining the proximal tubules in all treatment groups. There was tubular cell degeneration, tubular dilation with intratubular protein and regeneration. These changes were only found in three males in the low dose groups, and four males each in the medium and high dose groups.
Dose descriptor:
LOAEL
Effect level:
0.14 other: ml/kg/day
Sex:
male
Basis for effect level:
other: This type of renal pathology is specific to male rats due to an alpha2u-globulin-mediated process that is not relevant to humans.
Key result
Dose descriptor:
NOAEL
Effect level:
1.28 other: ml/kg/day
Sex:
female
Basis for effect level:
other: 1056 mg/kg bw
Critical effects observed:
not specified
Conclusions:
The LOAEL for male rats was 0.14 ml/kg/day based on renal damage. This type of renal damage is specific to male rats, and is not relevant to humans. The NOAEL for female rats was 1.28 ml/kg/day.
Executive summary:

The LOAEL for male rats was 0.14 ml/kg/day based on renal damage. This type of renal damage is specific to male rats, and is not relevant to humans. The NOAEL for female rats was 1.28 ml/kg/day. 

 

This study examined the oral 30 -day subchronic toxicity of hydrocarbons, C11-C14, n-alkanes, isoalkanes, cyclics, aromatics, 2-25% to rats. Groups of 5 rats of each sex were given doses of 0.14 (116 mg/kg), 0.42 (347 mg/kg), or 1.28 (1056 mg/kg) mL/kg of test substance in corn oil for 30 days. Animals were examined for clinical signs, mortality, body weight, food consumption, water consumption, and food conversion. After sacrifice clinical chemistry, hematology, clinical chemistry, urinalysis, organ weights, histopathology, and gross pathology were examined. There was no mortality during the experiment. Renal damage was observed in male rats at all dose levels. This type of renal pathology is specific to male rats due to a alpha2u-globulin-mediated process that is not relevant to humans. Female rats exhibited adaptive liver changes at the highest dosage. The LOAEL for male rats was 0.14 ml/kg/day based on renal damage. The female NOAEL was 1.28 (1056 mg/kg) mL/kg.

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study planned
Study period:
Will be completed in the timescale as indicated in the ECHA decision letter.
Justification for type of information:
This testing proposal has been presented in the lead registrant dossier for this substance submitted to ECHA in 2019. This is a source dossier where the overall approach should be seen in the context of a category of 108 different substances, where the substances are hydrocarbon solvents covering a carbon number range of C5-C20, based on alkane constituents and a range from approximately C8-C18 for aromatic constituents. The basis for this test proposal is set out in detail in the document ‘Hydrocarbon Solvents Test Proposals, Test Plans and Read-Across Strategy for Human Health Endpoints’, which is attached to this endpoint study record and in Section 13.2 of the IUCLID dossier.

TESTING PROPOSAL ON VERTEBRATE ANIMALS
[Please provide information for all of the points below. The information should be specific to the endpoint for which testing is proposed. Note that for testing proposals addressing testing on vertebrate animals under the REACH Regulation this document will be published on the ECHA website along with the third party consultation on the testing proposal(s).]

NON-CONFIDENTIAL NAME OF SUBSTANCE:
- Name of the substance on which testing is proposed to be carried out : Hydrocarbons, C16-C20, n-alkanes, isoalkanes, cyclics, aromatics (2-30%) (EC# 919-006-8)
- Name of the substance for which the testing proposal will be used [if different from tested substance] : Not different

CONSIDERATIONS THAT THE GENERAL ADAPTATION POSSIBILITIES OF ANNEX XI OF THE REACH REGULATION ARE NOT ADEQUATE TO GENERATE THE NECESSARY INFORMATION [please address all points below]:
- Available GLP studies : There are no OECD Guideline 408 studies available on this substance to evaluate the sub-chronic toxicity endpoint. These data only exist for structural analogues.
- Available non-GLP studies : There are no ‘non-GLP’ studies available for this substance to evaluate the sub-chronic toxicity endpoint. These data only exist for for structural analogues.
- Historical human data: No human data exist for this substance to evaluate sub-chronic toxicity hazard.
- (Q)SAR : There are no recognised (Q)SAR methods available for reliable prediction of sub-chronic toxicity.
- In vitro methods : There are no in vitro methods currently accepted by Regulatory Authorities for the reliable prediction of sub-chronic toxicity.
- Weight of evidence : Currently there are insufficient data available to develop a robust weight of evidence approach for the sub-chronic toxicity.
- Grouping and read-across : This test proposal maybe used to help develop a category approach for a wider range of hydrocarbons.
- Substance-tailored exposure driven testing [if applicable] : Insufficient data available
- Approaches in addition to above [if applicable]: None applicable
- Other reasons [if applicable]: None identified

According to the adaption principles of Annex XI of the REACH Regulation, there are no existing sub-chronic toxicity data (mammalian and human) for this substance, or for similar compositions, which could subsequently be used for read-across. Consequently, a weight of evidence approach is not sufficient to address this endpoint.

Both in-vitro and in-silico approaches are not considered to be appropriate, as there are no suitable methods which are considered to be valid and reliable to address the requirements for the Sub-Chronic Toxicity (90-Days) toxicity endpoint.

CONSIDERATIONS THAT THE SPECIFIC ADAPTATION POSSIBILITIES OF ANNEXES VI TO X (AND COLUMN 2 THEREOF) OF THE REACH REGULATION ARE NOT ADEQUATE TO GENERATE THE NECESSARY INFORMATION:
- There are no short-term toxicity studies (28-days) on this substance
- There are no chronic toxicity studies on this substance

FURTHER INFORMATION ON TESTING PROPOSAL IN ADDITION TO INFORMATION PROVIDED IN THE MATERIALS AND METHODS SECTION:
- Details on study design / methodology proposed [if relevant]: See 'Materials and Methods' Section for further information.
Qualifier:
according to guideline
Guideline:
OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)
GLP compliance:
yes (incl. QA statement)
Species:
rat
Sex:
male/female
Route of administration:
oral: unspecified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 056 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
1 key, subchronic study and 1 supporting, subchronic study from structural analogues available for assesment

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records

Referenceopen allclose all

Endpoint:
sub-chronic toxicity: inhalation
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1980
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Similar or equivalent to OECD TG 413.
Justification for type of information:
A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13.
Reason / purpose for cross-reference:
read-across: supporting information
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Shell Toxicology Laboratory (Tunstall) Breeding Unit
- Age at study initiation: 10-13 weeks
- Weight at study initiation: male mean weight: 396-398, female mean weight: 244-245
- Fasting period before study: food removed during exposure
- Housing: three per sex in hanging aluminum cages with stainless steel mesh bases 14 x 10 x 7 in, with two layers of cages for a total of twelve cages per exposure chamber
- Diet (e.g. ad libitum): LAD 1, Spillers Spratts Ltd., replenished daily after exposure
- Water (e.g. ad libitum): tap water ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17-22
- Humidity (%): 32-61
Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Vehicle:
other: unchanged (no vehicle)
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: 1 m³ aluminum exposure chamber
- Method of holding animals in test chamber: cages
- Source and rate of air: laboratory air
- Method of conditioning air: dust filters
- System of generating particulates/aerosols: Solvent was evaporated into the air stream using micrometering pumps and vaporizers. Vaporizers were quartz tubes heated to a surface temperature required for complete evaporation of the solvent.
- Temperature, humidity, pressure in air chamber: 17-22°C, 32-61%
- Air flow rate: 2.0 ± 0.03 m³/min
- Air change rate:
- Method of particle size determination:
- Treatment of exhaust air: Air was exhausted into the laboratory exhaust which exited on the roof of the laboratory.

TEST ATMOSPHERE
- Brief description of analytical method used: total hydrocarbon analyser fitted with a flame-ionization detector

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Total hydrocarbon analyser fitted with a flame-ionization detector.
Duration of treatment / exposure:
6 hours/day
Frequency of treatment:
5 days/week for 13 weeks
Remarks:
Doses / Concentrations:
1293 ppm (7400 mg/m3)
Basis:
analytical conc.
Remarks:
Doses / Concentrations:
690 ppm (3950 mg/m3)
Basis:
analytical conc.
Remarks:
Doses / Concentrations:
345 ppm (1975 mg/m3)
Basis:
analytical conc.
No. of animals per sex per dose:
18 per sex
Control animals:
yes, concurrent no treatment
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations: general health and behaviour

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

FOOD CONSUMPTION: Yes
- Time schedule for examinations: weekly

WATER CONSUMPTION: Yes
- Time schedule for examinations: weekly

HAEMATOLOGY: Yes
- Time schedule for collection of blood: end of experiment
- How many animals: all animals
- Parameters checked: erythrocyte count, mean cell volume, hemoglobin, leucocyte count, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, hematocrit, red cell fragilities, reticulocyte count, prothrombin time, kaolin-cephalin coagulation time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: end of experiment
- How many animals: all animals
- Parameters checked: total protein, urea nitrogen, alkaline phosphatase, aspartate amino transferase, alanine animo transferase, sodium, potassium, chloride, albumin, bilirubin
- Other: Estimations of blood glucose were made after 10 weeks of exposure using samples taken from the tail vein.

Sacrifice and pathology:
GROSS PATHOLOGY: Yes, all animals were examined and the following organs weighed: brain, liver, heart, spleen, kidneys, and testes.

HISTOPATHOLOGY: Yes, the following tissues of the high and medium exposure animals and the control animals were examined: mammary gland, mesenteric lymph node, pancreas, stomach, intestine at 5 levels, caecum, spleen, liver, adrenals, kidneys, ovaries or testes, uterus or prostate, seminal vesicles, urinary bladder, thyroid, trachea, heart, lungs, nasal cavity, thymus, eye and lachrymal glands, salivary glands, brain, spinal cord, pituitary, tongue, sciatic nerves, muscle, knee joint and femur, and macroscopic lesions. The kidneys of low exposure males were also examined.
Statistics:
Body and organ weights were analysed using covariance analysis, with initial body weight as the covariance. Means were adjusted if a significant covariance was found. Organs weights were also analysed using terminal body weights as the covariance. Clinical chemistry and hematological parameters were analysed using analysis of variance. Differences between treatment groups and controls were analysed using Williams t-test. Dunnett's test was used if a monotonic dose response could not be assumed.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY
There were no deaths during the experiment. High exposure animals were lethargic when examined 30 minutes after exposure. Animals were fully recovered by the next morning.

BODY WEIGHT AND WEIGHT GAIN
Males in the medium and high exposure groups had body weights significantly lower than controls. High exposure females also had body weights significantly lower than controls.

FOOD CONSUMPTION
No significant differences in food consumption were observed.

WATER CONSUMPTION
Exposed animals showed significant increase in water consumption, particularly animals in the high exposure group.

HAEMATOLOGY
Male PCV, erythrocyte count, mean cell volume, and mean corpuscular hemoglobin were significantly different from controls at all exposure levels. In females, the number of white cells, and mean cell volume were increased in the high exposure group.

CLINICAL CHEMISTRY
Alkaline phosphatase in both sexes, male aspartate amino transferase, and female albumin levels were significantly higher in the high exposure group. Female total protein was also significantly increased in the medium and high exposure groups.

ORGAN WEIGHTS
Male kidney weights at all exposure levels, and spleen weights in the medium and high exposure levels were significantly increased. Female kidney weights at the medium and high exposure levels, and liver weights at all exposure levels were also increased. However, there were no lesions in these organs found during the histopathology examination. These changes in femals were likely hyperfunctional adaptations of the organs rather than a toxic effect.

GROSS PATHOLOGY
Males in the medium and high exposure groups showed a low incidence of splenic enlargement, renal pallor, and hepatic darkening.

HISTOPATHOLOGY: NON-NEOPLASTIC
Kidneys - Male rats in all exposure group showed multiple, hyaline intracytoplasmic, inclusion droplets in the epithelium of the proximal convoluted tubules of their kidneys. This change did not seem to be dose related. These males also showed increase in the number and size of lysosomes in the cytoplasm of the proximal convoluted tubules. Exposure males also had more frequent focal tubular basophilia. Three males in the high exposure group also showed focal tubular dilatation and inspissated debris in the tubular laminae.

Spleen - Males in the medium and high exposure groups showed accelerated erythropoietic activity and increased hemosiderin deposition. Females in the high exposure group showed increased hemosiderin deposition, and mild extramedullary hematopoiesis.

Lungs - No treatment related effects were noted.
Key result
Dose descriptor:
LOAEC
Effect level:
345 ppm
Sex:
male
Basis for effect level:
other: 1975 mg/m3; increased kidney weights as a result of an alpha2u-globulin-mediated process that is not regarded as relevant to humans
Key result
Dose descriptor:
NOAEC
Effect level:
690 ppm
Sex:
female
Basis for effect level:
other: 3950 mg/m3
Critical effects observed:
not specified

Mean Body Weights of Male Rats (g)

Week

Control

345 ppm

690 ppm

1293 ppm

Standard Deviation of a Single Observation

0

397

396

396

398

24.9

1

422

422

396

396

20.9 (cage effect)

2

435

434

400

402

21.7

3

444

441

407

407

26.0

4

450

444

423

413

25.5

5

455

452

432

423

25.4

6

464

461

443

431

25.0

7

471

473

449

437

34.2 (cage effect)

8

480

479

460

445

28.2

9

486

486

466

448

30.9

10

494

490

469

453

35.6

11

495

491

478

458

34.9

12

502

495

481

466

36.7

13

512

503

491

473

38.4

Mean Body Weights of Female Rats (g)

Week

Control

345 ppm

690 ppm

1293 ppm

Standard Deviation of a Single Observation

0

244

245

244

245

14.2

1

249

253

252

245

6.2

2

256

261

257

248

9.0

3

264

264

263

252

10.1

4

264

269

266

254

13.5 (cage effect)

5

266

271

267

261

12.9 (cage effect)

6

269

274

269

261

11.4

7

274

278

273

264

11.5

8

275

277

274

263

12.5

9

275

277

272

266

13.8 (cage effect)

10

274

278

273

264

11.3

11

275

279

276

267

12.0

12

280

284

280

271

12.1

13

286

291

289

273

13.3
Conclusions:
The 90-day LOAEC for male rats was 345 ppm (inhalation). This value is based on increased kidney weights as a result of an alpha2u-globulin-mediated process that is not regarded as relevant to humans. The LOAEC was established at 1293 ppm (7400 mg/m3) due to a significant body weight reduction. No other effects were noted. The NOAEC for female rats was 690 ppm (3950 mg/m3).
Executive summary:

This study evaluated the subchronic toxicity of low aromatic white spirits to rats when exposed via inhalation. Groups of 18 rats per sex were exposed to 345, 690, or 1293 ppm of test substance for 6 hrs/day, 5 days/week, for 13 weeks. The highest concentration, 1293 ppm, was near the saturation point for test substance vapor. Rats were observed for clinical signs, mortality, food consumption, water consumption, and body weight. At the end of the exposure period, the animals were sacrificed, and clinical chemistry, hematology, gross pathology, and histopathology parameters were examined. Male rats at all exposure levels had degenerative effects of the as a result of an alpha2u-globulin-mediated process that is not regarded as relevant to humans. The LOAEC was established at 1293 ppm (7400 mg/m3) due to a significant body weight reduction. No other effects were noted. The NOAEC for female rats was 690 ppm (3950 mg/m3).  

Endpoint:
sub-chronic toxicity: inhalation
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
1979
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study.
Justification for type of information:
A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13.
Reason / purpose for cross-reference:
read-across: supporting information
Qualifier:
according to guideline
Guideline:
OECD Guideline 413 (Subchronic Inhalation Toxicity: 90-Day Study)
GLP compliance:
no
Limit test:
yes
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories
- Age at study initiation: 4 weeks males, 5 weeks females
- Weight at study initiation: males 142-214 g, females 140-189 g
- Housing: elevated stainless steel wire mesh cages, individually outside of chamber, pairs within exposure chamber
- Diet (e.g. ad libitum): Purina Laboratory Chow, ad libitum except during exposure
- Water (e.g. ad libitum): ad libitum except during exposure
- Acclimation period:

IN-LIFE DATES: From: April 17, 1978 To: July 12, 1978
Route of administration:
inhalation: vapour
Type of inhalation exposure:
whole body
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: stainless steel and glass chamber, 760 l
- Method of conditioning air: Test substance was metered using a syringe pump driven 50 cc Tomac glass syringe from a 500 ml Erlenmeyer flask into a heated flask and flash evaporated. Clean air was passed through this flask to pick up vapor. The test atmosphere was then fed into the chamber air inlet line where it was diluted to the desired concentration.
- Temperature, humidity, pressure in air chamber:
- Air flow rate: 134 lpm
- Air change rate: 7.46 min, with a 99% equilibration time of 34.3 min.



TEST ATMOSPHERE
- Brief description of analytical method used: Miran IA Ambient Air Analyser IR analyzed at 3.4 microns. Samples were drawn at 1, 3, and 5 hrs of exposure. Charcoal trapped vapor samples were also analyzed using GC.
- Samples taken from breathing zone: yes

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Miran IA Ambient Air Analyser IR analyzed at 3.4 microns. Samples were drawn at 1, 3, and 5 hrs of exposure. Charcoal trapped vapor samples were also analyzed using GC.
Duration of treatment / exposure:
6 hrs/day
Frequency of treatment:
5 days/week for 12 weeks
Remarks:
Doses / Concentrations:
100 ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
300 ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
103 ppm
Basis:
analytical conc.
Remarks:
Doses / Concentrations:
294 ppm
Basis:
analytical conc.
No. of animals per sex per dose:
35 per sex/per dose
Control animals:
yes, concurrent no treatment
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Weekly

BODY WEIGHT: Yes
- Time schedule for examinations: weekly starting five days prior to exposure

HAEMATOLOGY: Yes
- Time schedule for collection of blood: day 0, 4 weeks, 8 weeks, and 12 weeks
- How many animals: 10 animals per sex
- Parameters examined: hemoglobin, hematocrit, erythrocyte count, clotting time, total and differential leucocytes

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: day 0, 4 weeks, 8 weeks, and 12 weeks
- How many animals: 10 animals per sex
- Parameters examined: blood urea nitrogen, serum glutamic pyruvic transaminase, alkaline phosphatase
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
10 animals per sex per group were sacrificed at 4, 8, and 12 weeks (all survivors) by exsanguination with anesthesia. The adrenals, brain, gonads, kidneys, liver, and lungs were weighed.

HISTOPATHOLOGY: Yes
The organs of the sacrificed animals from groups I and III were examined histopathologically. The following organs were examined: adrenals, bone marrow, brain, eye, gonad, heart, colon, duodenum, ileum, kidneys, liver, lung, lymph node, mammary gland, pancreas, pituitary, salivary gland, skeletal muscle, skin, spinal cord, spleen, stomach, thyroid, trachea, urinary bladder, uterus or prostate, gross lesions, tissue masses.

Statistics:
hematology, and clinical chemistry: Snedecor, GS, and Cochran, WG, Statistical Methods. 6th ed., Iowa State University Press, Ames, 1967, 104-106, 114-119.

body weight, organ weight, and organ/body weight ratios: Dunnett, CW, J. Am. Stat. Assn. 50: 1096-1121, 1955, and Biometrics 20: 482, September, 1964.
Clinical signs:
effects observed, treatment-related
Mortality:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY
There was no mortality during the study. Females in both exposure groups had yellow staining of the ano-genital fur, which was possibly treatment related. Several animals in all groups exhibited dry rales, mucoid nasal discharge, and red nasal discharge. There was not a clear treatment related pattern. A few animals in all groups exhibited moist rales, chromodacryorrhea, excessive lacrimation, excessive salivation, alopecia, and brown staining of the ano-genital area. There was singular observations of a film covered eye and labored breathing in the high exposure group, and also one observation of loose stool in this group.

BODY WEIGHT AND WEIGHT GAIN
Male body weights were significantly increased at week 2 in both exposure groups. Females in the 300 ppm exposure group had significantly decreased body weights at day 0, week 1, and week 3. These weight differences were not considered to be treatment related.

HAEMATOLOGY
A significant decrease in hematocrit levels was seen in males exposed to 300 ppm at week 8 and week 12. Females in the 100 ppm group had decreased hematocrit values at week 4, and week 8, and females in the 300 ppm group had decreased values at week 4 only. Hemoglobin values were decreased in 100 ppm and 300 ppm exposed females at week 4, and 300 ppm exposed females had significantly decreased mean red blood cell counts at week 4. None of these findings appeared to be biologically significant. Males in the 300 ppm exposure group had elevated mean total leucocyte values at week 12. This was possibly treatment related.

CLINICAL CHEMISTRY
A significant increase in blood urea was seen in both groups of exposed males, indicating a possible treatment related effect. The males in the 100 ppm group had decreased mean glucose level at week 12. Females in the 300 ppm exposure group had decreased mean serum glutamic pyruvic transaminase level in week 4. These effects did not show a treatment related pattern, or indicate an abnormal condition, so they were not considered significant.

ORGAN WEIGHTS
Male kidney weight and kidney/body weight ratios were significantly elevated in the 100 ppm group at the week 4 sacrifice. At the week 12 sacrifice, males in both exposure groups had significantly elevated kidney/body weight ratios, and males in the 300 ppm group also had significanlty elevated kidney weights. At the week 8 sacrifice, females in the 300 ppm exposure group also had elevated kidney weights, and kidney/body weight ratios. These effects did not follow a clear dose related pattern.

HISTOPATHOLOGY
No treatment related effects were observed.
Key result
Dose descriptor:
NOAEC
Effect level:
>= 300 ppm
Sex:
female
Basis for effect level:
other: Lack of adverse treatment-related effects observed at the highest concentration tested
Key result
Dose descriptor:
LOAEC
Effect level:
100 ppm
Sex:
male
Basis for effect level:
other: increased kidney weights as a result of an alpha2u-globulin-mediated process that is not regarded as relevant to humans
Critical effects observed:
not specified

Mean Body Weight of Male Rats

Week

Control

100 ppm

300 ppm

0

Mean (g)

187

185

185

Standard deviation (g)

11

14

12

Number of Animals

35

35

35

1

Mean (g)

240

241

238

Standard deviation (g)

12

17

15

Number of Animals

35

35

35

2

Mean (g)

268

285

283

Standard deviation (g)

17

20

19

Number of Animals

35

35

35

3

Mean (g)

324

323

326

Standard deviation (g)

17

24

25

Number of Animals

35

35

35

4

Mean (g)

336

338

347

Standard deviation (g)

22

28

27

Number of Animals

35

35

35

5

Mean (g)

379

368

387

Standard deviation (g)

26

36

36

Number of Animals

25

24

25

6

Mean (g)

408

400

413

Standard deviation (g)

29

35

39

Number of Animals

25

25

25

7

Mean (g)

430

420

435

Standard deviation (g)

32

36

43

Number of Animals

25

25

25

8

Mean (g)

447

434

451

Standard deviation (g)

24

40

49

Number of Animals

25

25

25

9

Mean (g)

468

460

487

Standard deviation (g)

40

40

56

Number of Animals

15

15

15

10

Mean (g)

481

470

499

Standard deviation (g)

42

41

58

Number of Animals

15

15

15

11

Mean (g)

494

484

516

Standard deviation (g)

45

42

61

Number of Animals

15

15

15

12

Mean (g)

495

491

520

Standard deviation (g)

43

43

62

Number of Animals

15

15

15

Mean Body Weight of Female Rats

Week

Control

100 ppm

300 ppm

0

Mean (g)

163

162

158

Standard deviation (g)

10

10

9

Number of Animals

35

35

35

1

Mean (g)

188

186

181

Standard deviation (g)

11

12

10

Number of Animals

35

35

35

2

Mean (g)

202

203

199

Standard deviation (g)

12

14

13

Number of Animals

35

35

35

3

Mean (g)

226

223

219

Standard deviation (g)

14

16

13

Number of Animals

35

33

35

4

Mean (g)

230

229

224

Standard deviation (g)

15

20

15

Number of Animals

35

35

35

5

Mean (g)

253

251

248

Standard deviation (g)

17

21

17

Number of Animals

25

25

25

6

Mean (g)

265

263

258

Standard deviation (g)

19

22

17

Number of Animals

25

25

25

7

Mean (g)

272

270

270

Standard deviation (g)

19

23

19

Number of Animals

25

25

25

8

Mean (g)

279

277

275

Standard deviation (g)

20

24

18

Number of Animals

25

25

25

9

Mean (g)

289

285

284

Standard deviation (g)

23

28

22

Number of Animals

15

15

15

10

Mean (g)

292

289

285

Standard deviation (g)

26

30

20

Number of Animals

15

15

15

11

Mean (g)

297

291

289

Standard deviation (g)

26

31

23

Number of Animals

15

15

15

12

Mean (g)

298

291

289

Standard deviation (g)

24

31

24

Number of Animals

15

15

15
Conclusions:
For male rats, the LOAEC was 100 ppm via inhalation. This value is based on kidney effects due to a alpha2u-globulin-mediated process that is not regarded as relevant to humans that are not relevant to humans. For female rats, the NOAEC was 300 ppm. These results do not warrant classification under either the Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC or under the new Regulation (EC) 1272/2008 on classification, labeling and packaging of substances and mixtures (CLP).
Executive summary:

This study examined the subchronic toxicity of MRD-78-25 to rats via inhalation. Groups of 35 rats per sex were exposed to 0, 100, or 300 ppm of test substance vapors. Exposure was 6 hrs/day, 5 days/week, for 12 weeks. 10 rats/sex from each group were sacrificed at week 4 and week 8. Animals were observed for clinical signs daily, and weighed weekly. At the end of the study, all surviving animals were sacrificed. After sacrifice, hematological, clinical chemistry, and histopathological parameters were examined. There was no treatment related mortality during the study, and no treatment related body weight effects. For male rats, the LOAEC was 100 ppm via inhalation. This value is based on kidney effects due to a alpha2u-globulin-mediated process that is not regarded as relevant to humans that are not relevant to humans. For female rats, the NOAEC was 300 ppm. These results do not warrant classification under either the Directive 67/518/EEC for dangerous substances and Directive 1999/45/EC or under the new Regulation (EC) 1272/2008 on classification, labeling and packaging of substances and mixtures (CLP).

Endpoint conclusion
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEC
3 950 mg/m³
Study duration:
subchronic
Species:
rat
Quality of whole database:
1 supporting, short-term study and 2 key, subchronic studies available from structural analogues

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records
Reference
Endpoint:
sub-chronic toxicity: dermal
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
1997
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: According to or similar to guideline study OECD 411: GLP.
Justification for type of information:
The justification for read across is provided as an attachment in IUCLID Section 13.
Reason / purpose for cross-reference:
read-across: supporting information
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 411 (Subchronic Dermal Toxicity: 90-Day Study)
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
Groups of 12 male and 12 female, individually housed, Sprague-Dawley rats aged 7-9 weeks were used. The males weighed 198-328 g and the females weighed 156-249 g at the initiation of the study.
Type of coverage:
semiocclusive
Vehicle:
other: unknown
Details on exposure:
Test material was applied at concentrations of 20, 40 or 60% (v/v) at a rate of 1 ml/kg/day to the shorn intrascapular region of the rats. This was equivalent to doses of test material of 165, 330 or 495 mg/kg/day. Dosing was continued daily for five consecutive days each week, five days a week for 13 weeks. In addition a group of 12 male and 12 female rats of similar age were administered mineral oil at a dose rate of 1 ml/kg/day; these animals served as vehicle controls. An additional 12 rats/sex/group in the vehicle controls and high dose group were maintained for a 4-week recovery period following dosing for 13 weeks. All animals were fitted with collars to prevent ingestion and these were removed six hours after dosing and any residual test or control material was wiped from the skin. Animals were observed for clinical signs prior to dosing and 1, 6 and 24 hours after the first dose. Subsequently, observations were made prior to each dose being applied. Prior to the administration of each dose, the treated skin site was evaluated for dermal irritation using the Draize scoring method. Body weights were recorded prior to the first dose and weekly thereafter. An ophthalmic examination was conducted on each rat prior to application of the first dose and again prior to sacrifice at the end of the study. During the week prior to the first dose, each rat was subjected to a functional observation battery (FOB). The FOB was conducted again 1 and 24 hours after the first dose and at 7 and 14 days. During the study, the FOB, motor activity and startle response testing was conducted on all rats at weeks 4, 8 and 12. [The details of the FOB, the test for startle response test and the test for motor activity are given in detail in the laboratory report but are not included here]. At the 14 week necropsy, blood samples were collected from 12 animals/sex/group and at the week 18 necropsy from the recovery rats (vehicle and high dose groups).
Duration of treatment / exposure:
This was equivalent to doses of test material of 165, 330 or 495 mg/kg/day. Dosing was continued daily for five consecutive days each week, five days a week for 13 weeks.
Frequency of treatment:
This was equivalent to doses of test material of 165, 330 or 495 mg/kg/day. Dosing was continued daily for five consecutive days each week, five days a week for 13 weeks.
Remarks:
Doses / Concentrations:
0, 165, 330 or 495 mg/kg/day
Basis:
nominal per unit body weight
No. of animals per sex per dose:
Groups of 12 male and 12 female per dose
Control animals:
yes, concurrent vehicle
Details on study design:
Groups of 12 male and 12 female, individually housed, Sprague-Dawley rats aged 7-9 weeks were used. The males weighed 198-328 g and the females weighed 156-249 g at the initiation of the study.
Test material was applied at concentrations of 20, 40 or 60% (v/v) at a rate of 1 ml/kg/day to the shorn intrascapular region of the rats. This was equivalent to doses of test material of 165, 330 or 495 mg/kg/day. Dosing was continued daily for five consecutive days each week, five days a week for 13 weeks. In addition a group of 12 male and 12 female rats of similar age were administered mineral oil at a dose rate of 1 ml/kg/day; these animals served as vehicle controls. An additional 12 rats/sex/group in the vehicle controls and high dose group were maintained for a 4-week recovery period following dosing for 13 weeks. All animals were fitted with collars to prevent ingestion and these were removed six hours after dosing and any residual test or control material was wiped from the skin. Animals were observed for clinical signs prior to dosing and 1, 6 and 24 hours after the first dose. Subsequently, observations were made prior to each dose being applied. Prior to the administration of each dose, the treated skin site was evaluated for dermal irritation using the Draize scoring method. Body weights were recorded prior to the first dose and weekly thereafter. An ophthalmic examination was conducted on each rat prior to application of the first dose and again prior to sacrifice at the end of the study. During the week prior to the first dose, each rat was subjected to a functional observation battery (FOB). The FOB was conducted again 1 and 24 hours after the first dose and at 7 and 14 days. During the study, the FOB, motor activity and startle response testing was conducted on all rats at weeks 4, 8 and 12. [The details of the FOB, the test for startle response test and the test for motor activity are given in detail in the laboratory report but are not included here]. At the 14 week necropsy, blood samples were collected from 12 animals/sex/group and at the week 18 necropsy from the recovery rats (vehicle and high dose groups).

The following hematological and clinical chemical parameters were measured.
HEMATOLOGY:
Erythrocyte count
Hemoglobin
Hematocrit
Mean corpuscular volume
Mean corpuscular hemoglobin
Mean corpuscular hemoglobin concentration
Platelet count
Reticulocyte count
Total leukocyte count
Differential leukocyte count
Morphological examination of erythrocytes and platelets Coagulation determinations (prothrombin time & activated partial thromboplastin time) were also carried out on six animals from each group at week 14 and from the recovery groups at the week 18 necropsy.

CLINICAL CHEMISTRY
Blood urea nitrogen,
Creatinine,
Serum aspartate aminotransferase,
Serum alanine aminotransferase,
Alkaline phosphatase,
Lactate dehydrogenase,
Sorbitol dehydrogenase,
Gamma glutamyl transferase,
Creatinine kinase,
Serum glucose
Total, direct and indirect bilirubin
Total protein
Albumin
Calcium
Phosphorus
Sodium
Potassium
Chloride
A complete necropsy was performed on six rats/sex/group following 13 weeks dosing, and on 6 rats/sex/group of the recovery animals (high dose and controls) at week 18. A limited necropsy was performed on the remaining six animals and their organs were not weighed (see below). Each full necropsy included an examination of the external surface of the body, all orifices, cranial, thoracic, abdominal and pelvic cavities and their contents. Gross observations were recorded and the following organs were weighed: Adrenal glands, brain, epididymides, heart, kidneys, liver, ovaries, prostate, spleen, testes, thymus and uterus.

The following tissues were collected, processed and then examined microscopically.
Adrenal glands,
Nose (nasal cavity & turbinates),
Animal identification,
Ovaries,
Bone marrow (from sternum),
Oviducts,
Brain,
Pancreas,
Epididymides,
Parathyroid glands,
Esophagus,
Pituitary gland,
Exorbital lacrimal glands,
Prostate Eyes with optic nerve,
Salivary glands,
Femur (incl. articular surface),
Seminal vesicles,
Gross lesions Skin (application site),
Harderian gland,
Skin (inguinal),
Heart and aorta,
Spinal cord (3 levels),
Intestine (3 levels),
Spleen,
Kidneys,
Stomach,
Larynx and pharynx,
Testes,
Liver,
Thymus,
Lungs with mainstream bronchi,
Thyroid gland,
Lymph nodes (mandibular/mesenteric),
Urinary bladder,
Mammary glands with adjacent skin,
Uterus Muscle (thigh),
Vagina Nerve (sciatic).

The remaining six rats of each group were anesthetized with an intraperitoneal injection of Pentothal ® and transcardially perfused in-situ using 10% neutral-buffered formalin and given a limited necropsy. For these rats, no organs were weighed and the following tissues were collected: Head/skull, Sural nerve, Brain, Tibial nerve, Spinal cord, Gross lesions, Sciatic nerve.

The following tissues were examined microscopically in these animals: Brain (forebrain, cerebrum, midbrain, cerebellum, pons and medulla obligata), Gasserian ganglia, Dorsal root ganglia, Dorsal and ventral root fibers, Sural nerve, Tibial nerve, Spinal cord (cervical and lumbar areas), and Sciatic nerve.
Observations and examinations performed and frequency:
Groups of 12 male and 12 female, individually housed, Sprague-Dawley rats aged 7-9 weeks were used. The males weighed 198-328 g and the females weighed 156-249 g at the initiation of the study.
Test material was applied at concentrations of 20, 40 or 60% (v/v) at a rate of 1 ml/kg/day to the shorn intrascapular region of the rats. This was equivalent to doses of test material of 165, 330 or 495 mg/kg/day. Dosing was continued daily for five consecutive days each week, five days a week for 13 weeks. In addition a group of 12 male and 12 female rats of similar age were administered mineral oil at a dose rate of 1 ml/kg/day; these animals served as vehicle controls. An additional 12 rats/sex/group in the vehicle controls and high dose group were maintained for a 4-week recovery period following dosing for 13 weeks. All animals were fitted with collars to prevent ingestion and these were removed six hours after dosing and any residual test or control material was wiped from the skin. Animals were observed for clinical signs prior to dosing and 1, 6 and 24 hours after the first dose. Subsequently, observations were made prior to each dose being applied. Prior to the administration of each dose, the treated skin site was evaluated for dermal irritation using the Draize scoring method. Body weights were recorded prior to the first dose and weekly thereafter. An ophthalmic examination was conducted on each rat prior to application of the first dose and again prior to sacrifice at the end of the study. During the week prior to the first dose, each rat was subjected to a functional observation battery (FOB). The FOB was conducted again 1 and 24 hours after the first dose and at 7 and 14 days. During the study, the FOB, motor activity and startle response testing was conducted on all rats at weeks 4, 8 and 12. [The details of the FOB, the test for startle response test and the test for motor activity are given in detail in the laboratory report but are not included here].
Sacrifice and pathology:
At the 14 week necropsy, blood samples were collected from 12 animals/sex/group and at the week 18 necropsy from the recovery rats (vehicle and high dose groups).

The following hematological and clinical chemical parameters were measured.
HEMATOLOGY:
Erythrocyte count
Hemoglobin
Hematocrit
Mean corpuscular volume
Mean corpuscular hemoglobin
Mean corpuscular hemoglobin concentration
Platelet count
Reticulocyte count
Total leukocyte count
Differential leukocyte count
Morphological examination of erythrocytes and platelets Coagulation determinations (prothrombin time & activated partial thromboplastin time) were also carried out on six animals from each group at week 14 and from the recovery groups at the week 18 necropsy.

CLINICAL CHEMISTRY
Blood urea nitrogen,
Creatinine,
Serum aspartate aminotransferase,
Serum alanine aminotransferase,
Alkaline phosphatase,
Lactate dehydrogenase,
Sorbitol dehydrogenase,
Gamma glutamyl transferase,
Creatinine kinase,
Serum glucose
Total, direct and indirect bilirubin
Total protein
Albumin
Calcium
Phosphorus
Sodium
Potassium
Chloride
A complete necropsy was performed on six rats/sex/group following 13 weeks dosing, and on 6 rats/sex/group of the recovery animals (high dose and controls) at week 18. A limited necropsy was performed on the remaining six animals and their organs were not weighed (see below). Each full necropsy included an examination of the external surface of the body, all orifices, cranial, thoracic, abdominal and pelvic cavities and their contents. Gross observations were recorded and the following organs were weighed: Adrenal glands, brain, epididymides, heart, kidneys, liver, ovaries, prostate, spleen, testes, thymus and uterus.

The following tissues were collected, processed and then examined microscopically.
Adrenal glands,
Nose (nasal cavity & turbinates),
Ovaries,
Bone marrow (from sternum),
Oviducts,
Brain,
Pancreas,
Epididymides,
Parathyroid glands,
Esophagus,
Pituitary gland,
Exorbital lacrimal glands,
Prostate Eyes with optic nerve,
Salivary glands,
Femur (incl. articular surface),
Seminal vesicles,
Gross lesions Skin (application site),
Harderian gland,
Skin (inguinal),
Heart and aorta,
Spinal cord (3 levels),
Intestine (3 levels),
Spleen,
Kidneys,
Stomach,
Larynx and pharynx,
Testes,
Liver,
Thymus,
Lungs with mainstream bronchi,
Thyroid gland,
Lymph nodes (mandibular/mesenteric),
Urinary bladder,
Mammary glands with adjacent skin,
Uterus Muscle (thigh),
Vagina Nerve (sciatic).

The remaining six rats of each group were anesthetized with an intraperitoneal injection of Pentothal ® and transcardially perfused in-situ using 10% neutral-buffered formalin and given a limited necropsy. For these rats, no organs were weighed and the following tissues were collected: Head/skull, Sural nerve, Brain, Tibial nerve, Spinal cord, Gross lesions, Sciatic nerve.

The following tissues were examined microscopically in these animals: Brain (forebrain, cerebrum, midbrain, cerebellum, pons and medulla obligata), Gasserian ganglia, Dorsal root ganglia, Dorsal and ventral root fibers, Sural nerve, Tibial nerve, Spinal cord (cervical and lumbar areas), and Sciatic nerve.
Statistics:
Statistics Normally-distributed in-life data (parametic) were analyzed for test substance effects by analysis of variance and pairwise comparisons made between groups using Dunnett's test. Nonparametric data (nonhomogenous as determined by Bartlett's test) were analyzed using a modified t-test. Statistical significance was reported at the P < 0.05 level. Statistical analyses of neurobehavior data (FOB and motor activity) are described in the results section.
Clinical signs:
no effects observed
Dermal irritation:
effects observed, treatment-related
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
There were no test substance-related effects on survival, clinical observations (apart from skin irritation), neurobehavioral signs or ophthalmological findings. The only clinical observations during the study were related to skin irritation at the application site. There was a generally dose-related increase in the incidence and severity of erythema, edema, epidermal scaling, scab formation, thickening of the skin and ulceration at the treated site. Males seemed to be more sensitive than females. The FOB screen did not demonstrate any substance-related effects. The areas monitored were: behavioral parameters, including autonomic, muscle tone and equilibrium, sensorimotor responses, central nervous system. In addition the test substance had little effect on motor activity or startle response. Growth rates were unaffected by treatment. At necropsy no substance-related observations were made for males in any group. In the females there was a suggestion of a possible treatment- related effect which occurred in 7 rats across all groups and consisted of skin crusts or ulceration at the site of application of test material. Hematological and serum clinical parameters were unaffected by treatment. The only organ weight effects noted were an increase in spleen/body weight and spleen/brain weight ratios in the high dose group females at the 13 week necropsy and an increase in absolute spleen weight in the same dose group females after the 4 weeks recovery period. Since there were no associated microscopic or clinical chemical findings, these differences were not considered to be of biological relevance. There were no treatment-related microscopic changes in the tissues examined with the exception of the findings in the skin. The skin observations were minimal in nature with a severity score less than 1 on a 1 [low] to 4 [severe] scale. The findings included acanthosis, ulceration, parakeratosis, chronic active inflammation and hyperkeratosis. The males were affected at all doses, however, the effects indicated very little irritation. Recovery group animals revealed complete recovery in the females and minimal hyperkeratosis in the high dose group males. No effects were found in the animals subjected to a detailed neuropathological examination.
Key result
Dose descriptor:
NOAEL
Remarks:
Systemic
Effect level:
> 495 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: No systemic or neurological effects were noted in the study
Critical effects observed:
not specified
Conclusions:
There were no systemic or neurological effects noted at any of the tested doses. The systemic NOAEL was >495 mg/kg/day.
Executive summary:

Test material was applied at concentrations of 165, 330 or 495 mg/kg/day. Dosing was continued daily for five consecutive days each week, five days a week for 13 weeks. In addition a group of 12 male and 12 female rats of similar age was administered mineral oil as vehicle controls and an additional high dose group was maintained for a 4-week recovery period following dosing for 13 weeks. At the 14 week necropsy, blood samples were collected from 12 animals/sex/group and at the week 18 necropsy from the recovery rats (vehicle and high dose groups). There were no systemic or neurological effects noted at any of the tested doses. The systemic NOAEL was >495 mg/kg/day.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
495 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Quality of whole database:
1 supporting, short-term study and 1 key, subchronic study from structural analogues available for assessment

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

No oral, inhalation, or dermal repeat dose toxicity data is available for Hydrocarbons, C16-C20, n-alkanes, isoalkanes, cyclics, aromatics (2-30%). However, data is available for structural analogues, Hydrocarbons, C9-C12, n-alkanes, isoalkanes, cyclics, aromatics (2 -25%), Hydrocarbons, C11-C14, n-alkanes, isoalkanes, cyclics, aromatics (2-25%) and Kerosene. Petroleum substances of similar carbon number and aromatic content, principally kerosene and jet fuel, are typically in the range of C9-C16. These substances also contain similar types of molecules in similar proportions to those in C14-C20 aliphatic [2-30% Aromatics] Hydrocarbon solvents. In general, hydrocarbon solvents are more highly refined than petroleum substances. Accordingly, the petroleum substances typically represent a “worse case” with respect to hydrocarbon solvents and can be used for read across on that basis. This data is read across to based on analogue read across and a discussion and report on the read across strategy is provided as an attachment in IUCLID Section 13.

Oral:

Hydrocarbons, C11-C14, n-alkanes, isoalkanes, cyclics, aromatics (2-25%)

A key repeat dose sub-chronic oral toxicity study (DHC, 1984a) examined the oral 30 -day subchronic toxicity of Hydrocarbons, C11-C14, n-alkanes, isoalkanes, cyclics, aromatics (2-25%) in rats. Groups of 5 rats of each sex were given doses of 0.14 (116 mg/kg), 0.42 (347 mg/kg), or 1.28 (1056 mg/kg) mL/kg of test substance in corn oil for 30 days. Animals were examined for clinical signs, mortality, body weight, food consumption, water consumption, and food conversion. After sacrifice clinical chemistry, hematology, clinical chemistry, urinalysis, organ weights, histopathology, and gross pathology were examined. There was no mortality during the experiment. Renal damage was observed in male rats at all dose levels. This type of renal pathology is specific to male rats due to a alpha2u-globulin-mediated process that is not relevant to humans. Female rats exhibited adaptive liver changes at the highest dosage. The LOAEL for male rats was 0.14 mL/kg/day based on renal damage (specific to male rats and is not relevant to humans). The female NOAEL was 1.28 (1056 mg/kg) mL/kg.

Kerosene

In a supporting study (Mattie et al. 1995), male rats were given 0, 750, 1500, or 3,000 mg/kg of the test material (JP-8) daily by gavage for 90 days. The test material was administered without a vehicle. Rats were given the last dose on day 90 and sacrificed within 3.8 ± 1.8 hours after dosing. Exposure to high concentrations of JP-8 by the oral route resulted in minimal toxicity except for a decrease in body weight. The NOAEL was determined to be 750 mg/kg/day based on body weight reduction.

Additionally, an OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents) test is proposed for Hydrocarbons, C16-C20, n-alkanes, isoalkanes, cyclics, aromatics (2-30%). This endpoint will be updated subsequent to ECHA’s approval of the testing proposal and availability of data upon completion of the study.

Inhalation:

Hydrocarbons, C9-C12, n-alkanes, isoalkanes, cyclics, aromatics (2-25%)

A key sub-chronic repeated dose study (Shell, 1980a) evaluated the subchronic toxicity of low aromatic white spirits to rats when exposed via inhalation. Groups of 18 rats per sex were exposed to 345, 690, or 1293 ppm of test substance for 6 hrs/day, 5 days/week, for 13 weeks. The highest concentration, 1293 ppm, was near the saturation point for test substance vapor. Rats were observed for clinical signs, mortality, food consumption, water consumption, and body weight. At the end of the exposure period, the animals were sacrificed, and clinical chemistry, hematology, gross pathology, and histopathology parameters were examined. Male rats at all exposure levels had degenerative effects of the kidney as a result of an alpha2u-globulin-mediated process that is not regarded as relevant to humans. The LOAEC was established at 1293 ppm (7400 mg/m3) due to a significant body weight reduction. No other effects were noted. The NOAEC for female rats was 690 ppm (3950 mg/m3).  

A second key sub-chronic repeated dose study (ExxonMobil, 1979a) examined the subchronic toxicity of the test material to rats via inhalation. Groups of 35 rats per sex were exposed to 0, 100, or 300 ppm of test substance vapors. Exposure was 6 hrs/day, 5 days/week, for 12 weeks. 10 rats/sex from each group were sacrificed at week 4 and week 8. Animals were observed for clinical signs daily and weighed weekly. At the end of the study, all surviving animals were sacrificed. After sacrifice, hematological, clinical chemistry, and histopathological parameters were examined. There was no treatment related mortality during the study, and no treatment related body weight effects. For male rats, the LOAEC was 100 ppm via inhalation. This value is based on kidney effects due to a alpha2u-globulin-mediated process that is not regarded as relevant to humans that are not relevant to humans. For female rats, the NOAEC was 300 ppm. 

Kerosene

In a supporting study (API, 1986a), groups of 20 male and 20 female Sprague Dawley rats (aged approximately six weeks) were exposed to a nominal concentration of 25mg/m³ of the test material (hydrodesulfurized kerosene) by inhalation. Exposures were for approximately six hours each day, five days each week for four consecutive weeks. Control groups of 20 male and 20 females were exposed to filtered air. Animals were observed twice daily for overt signs of toxicity and they underwent detailed examination once weekly. Body weights were also recorded weekly. At study termination, the animals were killed and blood samples were taken for the following clinical chemical and hematological investigations. There were no treatment-related effects on clinical condition, growth rate organ or organ body weight ratios or on any of the hematological or clinical chemistry determinations. Furthermore, there were no treatment-related microscopic changes observed in any of the organs examined. The NOAEC was determined to be 25mg/m3, which was the highest concentration tested.

Dermal:

Kerosene

In a key 90-day study (American Petroleum Institute, 1997), test material (Hydrodesulferized kerosene) was applied at concentrations of 165, 330 or 495 mg/kg/day. Dosing was continued daily for five consecutive days each week, five days a week for 13 weeks. In addition a group of 12 male and 12 female rats of similar age was administered mineral oil as vehicle controls and an additional high dose group was maintained for a 4-week recovery period following dosing for 13 weeks. At the 14 week necropsy, blood samples were collected from 12 animals/sex/group and at the week 18 necropsy from the recovery rats (vehicle and high dose groups). There were no systemic or neurological effects noted at any of the tested doses. The systemic NOAEL was ≥495 mg/kg/day.

In a short-term study (American Petroleum Institute, 1985c), test material (Straight run kerosene) was applied to 5 male and 5 female rabbits at doses of 200, 1000 and 2000 mg/kg/day, three times weekly until 12 doses had been applied. Five rabbits of each sex served as sham treated controls. Animals were observed twice daily for clinical signs, body weights recorded weekly, test site examined daily and reactions graded using the Draize scale. At termination, all animals were necropsied. Treatment-related mortality occurred in 1 male and 1 female in the 2000 mg/kg/day group. Treatment-related increases in heart weights were observed in the mid- and high- dose animals, and increased spleen weights were observed in females. The systemic NOAEL was determined to be 1000 mg/kg bw based on the treatment-related deaths noted at 2000 mg/kg/day.

Justification for classification or non-classification

Based on available read across data, Hydrocarbons, C16-C20, n-alkanes, isoalkanes, cyclics, aromatics (2-30%) does not meet the criteria for classification for repeated dose toxicity (STOT-RE) under the new Regulation (EC) 1272/2008 on classification, labeling and packaging of substances and mixtures (CLP).