Registration Dossier

Administrative data

Description of key information

In a repeated dose toxicity study the by oral route, the test item was administered by drinking water to rats (10 animals/sex/dose) at doses of 0; 100; 500 and 2500 ppm during 5-weeks. Under the test conditions, the test item Reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda induced only a slight reduction in body weight gain in males at the highest dose tested (2500 ppm). In the absence of other test item-related changes, this variation was considered of no toxicological relevance. The NOAEL was therefore considered to be 2500 ppm (i.e. 317.05 and 335.22 mg/kg b.w./day in males and females, respectively). No classification as STOT-RE is thus required according to the Regulation (EC) 1272/2008 (CLP) and as harmful after repeated dose exposure according to the Directive 67/548/EEC.
In a repeated dose inhalation toxicity study, groups of Wistar rats (5/sex) were exposed by inhalation route to the test substance as dust aerosol for 6 hours per day on 5 days per week for 4 weeks (28-day test) to head/nose at concentrations of 0, 0.8, 8, 80 mg/m³ (nominal concentrations) or 0, 0.86, 8.1, 82 mg/m³ (analytical concentrations). Based on the results of this study, 82 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) systemic based on the absence of toxicologically significant sytemic effects; 0,86 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) local and 8.1 mg/m³ of test item was established as the Lowest Observed Adverse Effect Level (LOAEL) local based on effects on the lung weight. Therefore, reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda is not classified for repeated dose toxicity according to the classification criteria of the Regulation (EC) 1272/2008 (CLP) and of the Directive 67/548/EEC.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
no data available
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: The study is sufficiently documented. Few deviations from the current guidelines were noted.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
Deviations:
yes
Remarks:
5-week treatment period instead of 4. Histopathology was conducted only on a selection of organs collected at necropsy. No analytical verifications of the doses administered.
GLP compliance:
no
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source:SPF. BOR: WISW (SPF CPB) of the breeder Winkelmann (Borchen)
- Age at study initiation: the rats were 5-6 weeks old
- Weight at study initiation: males: a mean body weight of 105 g, and female a mean body weight of 96 g.
- Fasting period before study: no
- Housing: The animals were placed in a traditional way each cage Makrolon Type II (and Gönnert SPIEGEL, 1961) on wood pellets free of dust.
- Diet (e.g. ad libitum): During the test, the animals received during the weekly cage change their weekly amount of food in the form of granules costs (Altromin 1324 Manufacturer: Altrogge GmbH, Lage).
- Water (e.g. ad libitum): The animals had free access to deionized drinking water, which was diluted test substancel.
- Acclimation period: the animals were acclimated for a period of 12 days before the beginning of the treatment period.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 50 ± 10%
- Air changes (per hr): no
- Photoperiod (hrs dark / hrs light): rhythm day / night 12 hours (artificial light from 6 to 18 pm CET)

IN-LIFE DATES: no data available
Route of administration:
oral: drinking water
Vehicle:
water
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS:
The test substance was diluted once a week in deionized water. To dissolve the deposits present at the bottom, it was heated to approx. 50 ° C. The control subjects received only de-ionized water.

VEHICLE
- de-ionized water
- Concentration in vehicle: 100, 500, 2500 ppm
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
5-weeks
Frequency of treatment:
not applicable
Remarks:
Doses / Concentrations:
100, 500, 2500 ppm
Basis:
nominal in water
No. of animals per sex per dose:
10 males and 10 females rats
Control animals:
yes
Details on study design:
ERKANTOL BXG was diluted once a week in deionized water in the following concentrations: 0 (control), 100, 500 and 2500 ppm by reference to the amount of 22% of test substance.
It was not advisable to use tap water, because with high concentrations it presented a phenomenon of turbidity with precipitation.
The concentrations were chosen based on the results of a pre-test only for a period of one week.
For each dose, the test group consisted of 10 male rats and 10 female rats.
The test substance was diluted once a week in deionized water. To dissolve the deposits present at the bottom, it was heated to approx. 50 ° C. The control subjects received only de-ionized water.
At the beginning of the test, the male and female rats were sorted into two weight groups (light, heavy) and divided into groups corresponding doses based on a random list. Finally, the animals were numbered consecutively.
Positive control:
not applicable
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: 2 times a day on weekdays and once a day on weekends and holidays

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: 2 times a day on weekdays and once a day on weekends and holidays

BODY WEIGHT: Yes
- Time schedule for examinations: at the beginning of the test and weekly

FOOD CONSUMPTION: Yes
The daily food intake was determined by weighing again the amount of uneaten food.

WATER CONSUMPTION: Yes
- Time schedule for examinations: The daily intake of water was determined by measuring the amount of new non-drinking water.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: during the 4th week of test
- Anaesthetic used for blood collection: Yes , ether
- Animals fasted: Yes
- How many animals: 5 males and 5 females of each group
- Erythrocyte count and leukocyte count: counting by Coulter Counter, number of platelets: measurement by TOA PL 100 system, hemoglobin concentration and mean cell volume (MCV): Measurement with a Coulter counter, hematocrit, mean corpuscular hemoglobin (MCH) and mean corpuscular hemoglobin concentration (MCHC): Calculation with a Coulter counter, blood count based on smear (modified Wright staining method) thromboplastin time test method according HEENE Hepato-Quick (1974).

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: during the 4th week of test
- Animals fasted: Yes
- How many animals: 5 males and 5 females of each group. Due to the suspicious nature of the results, in the fifth week of the test, we measured the concentration of inorganic phosphate on the 5 male rats and the remaining five female rats from each group.
- Enzyme activity in plasma: Alkaline phosphates (ALP), Glutamate-Oxaloacetate-Transaminase (GOT) and glutamate-pyruvate-transaminase (GPT), as recommended by the Deutsche Gesellschaft für Klinische Chemie (1972)
- Substrates in plasma: Creatinine according FABINY and ERTINGSHAUSEN (1971); Urea and BERGMEYER by Gutmann (1974); Glucose by SCHMIDT (1961); Cholesterol in TRINDER (1969); Bilirubin according WAHLEFELD et al. (1972); total albumin according WEICHSELBAUM (1946).
- Electrolytes in serum:Sodium, potassium and calcium determination by flame photometry; Chloride according LANGE LANGE und BORNER (1972);
Inorganic phosphate according DALY and ERTINGSHAUSEN (1972).

URINALYSIS: Yes
- Time schedule for collection of urine: during the 4th week of test. The urine was collected for periods of approx. 16 hours.
- Animals fasted: Yes
- Urinalysis: Semi-quantitative: Glucose, blood, albumin, bilirubin, ketones and pH using sticks Bili-Labstix; Urobilinogènes with Urobilistix, microscopic examinations of sediment after centrifugation of the urine sample; Quantitative: volumes over time; Albumin according RICHTERICH (1968) Specific gravity by digital density meter (DMA 45, Paar Heraeus soc.).

NEUROBEHAVIOURAL EXAMINATION: No data
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
Autopsy of rats died during the test:
The rats died during the treatment period were dissected and subjected to macroscopic evaluation. The organs and tissues listed below were treated with Bouin's solution.

Autopsy of rats killed after the test:
All surviving rats were anesthetized with diethyl ether and killed by exsanguination. During the autopsy, the animals were submitted to a pathologico-anatomical examination. The following organs were weighed: brain, heart, testis, lung, liver, spleen, adrenal glands and kidneys.
Preparations of the following organs of all subjects in all groups were treated in a Bouin's solution: aorta, eyes, intestines (duodenum, jejunum, ileum, cecum, colon, rectum), femoral block with skeletal muscle and sciatic nerve, brain, bladder, heart, testis, pituitary gland, salivary gland, lung, lymph nodes (mesenteric and cervical), stomach, spleen, epididymis, adrenal glands, esophagus, ovary, pancreas, prostate, seminal vesicle, the thyroid with parathyroid glands, sternum, thymus, trachea and uterus and all macroscopically visible changes. The liver and kidneys of 5 males and 5 females in each group were also preserved in Bouin's solution. Pieces of liver of these subjects were also preserved by fixation with formalin-calcium. The liver, kidneys and abdominal adipose tissue of other animals were frozen for possible future evaluation.

HISTOPATHOLOGY: Yes
Histopathological examination of the bones (femur, sternum with marrow), heart, liver, spleen, adrenal glands, kidneys and thyroid with parathyroid glands was performed for 5 male rats and five female rats of groups 0 and 2500 ppm.
Other examinations:
no other examination
Statistics:
- Calculation of the arithmetic mean value of each group, the standard deviation , the confidence limits for the confidence level 95% and 99%.
- Comparison of values ​​for the test item-treated and the control groups performed with the significance test (U-test) (MANN, WHITNEY and Wilcoxon (significance thresholds 5% and 1%)).
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Body weight was not affected by the test item in male given 100 and 500 ppm and in females whatever the dose level. Mean body weight gain was slightly delayed in males of group 2 500 ppm (not statistically significant).
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not examined
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:
Food and water intake, general behaviour and mortality were not affected by treatment with the test item. Body weight gain was slightly reduced only in male rats given 2500 ppm (up to 6% v.s. control, not statistically significant).
Up to 2500 ppm, hematological, plasma chemistry and urinary parameters were not affected by the test item.
No test item-related effects were reported at necropsy or during histopathological examinations up to the highest dose level.

Dose descriptor:
NOAEL
Effect level:
2 500 ppm
Based on:
test mat.
Sex:
male/female
Remarks on result:
other: no effects observed
Dose descriptor:
NOAEL
Effect level:
317.05 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Remarks on result:
other: no effects observed
Dose descriptor:
NOAEL
Effect level:
335.22 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Remarks on result:
other: no effects observed
Critical effects observed:
not specified

Table: Results

 

DOSE ppm

0

100

500

2500

MALE

FOOD INTAKE

g/animal/day

 

 

20

 

 

20

 

 

20

 

 

20

FEMALE

FOOD INTAKE

g/animal/day

 

 

15

 

 

15

 

 

16

 

 

16

 

DOSE ppm

0

100

500

2500

MALE

WATER CONSUMPTION

ml/animal/day

 

 

 

26

 

 

 

24

 

 

 

24

 

 

 

24

MALE

TEST COMPOUND INTAKE

mg/animal/day

mg/kg b.w./day

 

 

 

 

 

 

 

 

2.36

11.99

 

 

 

 11.87

61.36

 

 

 

59.88

317.05

FEMALE

WATER CONSUMPTION

ml/animal/day

 

 

 

22

 

 

 

20

 

 

 

18

 

 

 

19

FEMALE

TEST COMPOUND INTAKE

mg/animal/day

mg/kg b.w./day

 

 

 

 

 

 

 

 

2.05

14.57

 

 

 

 9.15

64.78

 

 

 

 46.72

335.22

Haematology

4 weeks

Dose

ppm

LEU

GIGA/L

ERY

TERA/L

HGB

G/L

MCV

FL

THROM

GIGA/L

THP

SEC

HCT

L/L

MCH

PG/E

MCHC

G/L

Male

0

100

500

2500

 

10.2

10.6

9.8

10.3

 

7.43

7.62

7.52

7.69

 

154

152

153

155

 

61

60

62

61

 

1048

1098

1174

1120

 

30.5

30.4

31.4

29.3

 

0.46

0.46

0.47ns

0.47**

 

20.7

19.9

20.3

20.2

 

338

330**

328*

330ns

Female

0

100

500

2500

 

8.1

7.7

7.0

7.3

 

7.43

7.29

7.45

7.42

 

151

146

151

151

 

62

61

61

61

 

1098

1027

1079

1094

 

28.7

29.4

30.5

29.2

 

0.46

0.44

0.46

0.45

 

20.3

20.0

20.2

20.4

 

330

329

331

332

 

Differential blood count (%)

4 weeks

Dose

ppm

BASO

EOSIN

IPC

JGL

STAB

SEGM

LYM

MONO

PLAS

Male

0

100

500

2500

 

0.0

0.0

0.0

0.0

 

0.2

0.4

0.4

0.2

 

0.0

0.0

0.0

0.0

 

0.0

0.0

0.0

0.0

 

10.2

8.8

7.6**

7.0*

 

89.6

90.8

92.0*

92.8ns

 

0.0

0.0

0.0

0.0

 

0.0

0.0

0.0

0.0

Female

0

100

500

2500

 

0.0

0.0

0.0

0.0

 

0.8

0.4

0.8

0.4

 

0.0

0.0

0.0

0.0

 

0.0

0.0

0.0

0.0

 

6.0

5.8

8.8

8.2

 

93.2

93.8

90.4

91.4

 

0.0

0.0

0.0

0.0

 

0.0

0.0

0.0

0.0

 

Clinical chemistry

4 weeks

Dose

ppm

ALP

U/L

GOT

U/L

GPT

U/L

BILI

µMOL/L

PROT/P

G/L

UREA

MMOL/L

CREA

µMOL/L

CHOL

MMOL/L

GLUC

MMOL/L

AN.PH

MMOL/L

NA

MMOL/L

 

K

MMOL/L

CA

MMOL/L

CL

MMOL/L

Male

0

100

500

2500

 

497

437**

430ns

587

 

63.1

47.8

44.8*

46.5

 

54.1

50.8

57.3

49.2

 

2.1

2.1

2.4

2.4

 

54.7

53.5

54.6

56.3

 

7.72

7.36

7.72

8.36

 

40

48

59

43

 

1.98

2.03

1.91

1.91

 

5.49

5.51

5.33

5.46

 

2.48

2.39

2.21**

2.06**

 

140

139

139

143

 

5.0

5.2

4.9

4.8

 

2.75

2.78

2.67

2.76

 

102

101

103

103

Female

0

100

500

2500

 

316

321

305

337

 

43.4

48.5

42.3

52.0

 

45.7

51.6

52.6

45.5

 

2.0

2.2

2.2

1.9

 

57.6

54.6

54.4

53.0

 

8.40

8.19

7.37*

9.27

 

54

42

47

45

 

2.02

1.71*

1.72

1.72

 

5.22

5.33

5.66

5.18

 

2.03

1.87

1.71*

1.57**

 

141

140

140

141

 

4.4

4.5

4.6

4.2

 

2.73

2.73

2.68

2.67

 

102

103

103

104

Quantitative urinalyses

4 weeks

Dose

ppm

PROTU

MG/16H

VOL

ML

SP.GR.

G/L

Male

0

100

500

2500

 

12.35

9.46

7.71

6.48**

 

35

21

13**

6**

 

1004

1007

1013

1030**

Female

0

100

500

2500

 

2.27

1.22ns

1.35*

1.23*

 

11

5

4*

4*

 

1010

1024

1034**

1037

Absolute organ weights (mg)

Dose

ppm

BODY-W

(G)

HEART

LUNG

LIV

SPLE

KIDN

ADRE

TEST

BRAIN

Male

0

100

500

2500

 

242

243

240

230

 

783

765

756

711

 

1032

1040

1075

988

 

9837

10474

10414

9747

 

484

482

492

474

 

1638

1785*

1682

1714

 

39

40

39

41

 

2915

2921

2991

2994

 

1660

1653

1667

1633

Female

0

100

500

2500

 

173

162

166

162

 

592

590

588

570

 

868

806

838

792*

 

7516

6952

7245

7341

 

408

379

366

373

 

1237

1218

1313

1330

 

58

60

57

60

 

 

1617

1652

1658

1652

Relative organ weights (mg/100g)

Dose

ppm

BODY-W

(G)

HEART

LUNG

LIV

SPLE

KIDN

ADRE

TEST

BRAIN

Male

0

100

500

2500

 

242

243

240

230

 

324

315

317

309

 

427

428

450

430

 

4063

4298

4328

4232

 

201

198

207

205

 

679

734*

700

746*

 

16

17

17

18

 

1213

1204

1256

1305

 

690

683

702

712

Female

0

100

500

2500

 

173

162

166

162

 

342

366

357

352

 

504

500

507

488

 

4350

4291

4369

4514

 

236

232

222

229

 

717

756

796**

819**

 

33

37

35

37

 

 

940

1027*

1008

1023**

Ns : not significant

Significant differences compared to the control group are marked with an asterisk (*) p ≤ 0.05 by two asterisks (**) for p ≤ 0.01.

Conclusions:
Under the test conditions, the test item Reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda induced only a slight reduction of body weight gain in males at the dose of 2500 ppm. In the absence of other test item-related changes, this slight variation is considered of no toxicological relevance. The NOAEL was therefore considered to be 2500 ppm (equivalent to 317.05 and 335.22 mg/kg b.w./day in males and females, respectively). Therefore no classification as STOT-RE is required according to the Regulation (EC) 1272/2008 (CLP) and as harmful after repeated dose exposure according to the Directive 67/548/EEC.
Executive summary:

In a repeated dose toxicity study by the oral route, the test item was administered in drinking water to rats (10 animals/sex/dose) at doses of 0; 100; 500 and 2500 ppm during 5 weeks.

Food and water intake, general behaviour and mortality were not affected by treatment. Body weight gain was slightly reduced only in male rats administered 2500 ppm (not statistically significant) but this variation was considered of no toxicological relevance owing to its minimal magnitude (maximum 6% v.s. control) and absence of associated test item-related variations. Up to 2500 ppm, hematological, plasma chemistry and urinary parameters were not affected by the test item. No test item-related effects were reported at necropsy or during histopathological examinations up to the highest dose level.

Based on the results of this study, 2500 ppm of test item, which corresponds to 317.05 and 335.22 mg/kg b.w./day in males and females, respectively, was established as the No Observed Adverse Effect Level (NOAEL). Therefore, reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda is not classified for repeated dose toxicity according to the classification criteria of the Regulation (EC) 1272/2008 (CLP) and of the Directive 67/548/EEC.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
317.05 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Klimisch 2

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
no data available
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: The study is sufficiently documented. OECD Guideline study; analytical purity of test substance: 68,3%.
Qualifier:
according to
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Dr. K Thomae GmbH
- Age at study initiation: about 7 weeks
- Weight at study initiation: males: mean = 230 g; females: mean = 177 g
- Housing: single
- Diet: ad libitum during post exposure observation period
- Water: ad libitumduring post exposure observation period

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
other: Inhalation: dust aerosol
Type of inhalation exposure:
nose/head only
Vehicle:
air
Remarks on MMAD:
MMAD / GSD: 0.8 mg/m³: 3.3 µm
8 mg/m³: 3.3 µm
80 mg/m³: 3.1 µm
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: aerodynamic exposure apparatus (INA 60, Volume ca. 90 L, BASF AG)
- Method of holding animals in test chamber: The rats were restrained in exposure tubes (glass tubes), their snouts projected into the inhalation chamber and they thus inhaled the dust aerosol.
- System of generating particulates/aerosols: brush generator
- Temperature, humidity, pressure in air chamber: 22°C, 50% humidity
- Method of particle size determination: cascade imapactor

TEST ATMOSPHERE
- Brief description of analytical method used: Spectrophotometric analysis
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
no data available
Duration of treatment / exposure:
6 hours per day for 28 days
Frequency of treatment:
5 days per week
Remarks:
Doses / Concentrations:
0, 0.8, 8, 80 mg/m³
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0, 0.86, 8.1, 82 mg/m³
Basis:
analytical conc.
No. of animals per sex per dose:
5
groups of female rats (5 animals) were hold without further exposure for an about 4-week post exposure
Control animals:
yes, concurrent vehicle
Details on study design:
no data available
Positive control:
no data available
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations: mortality, behavior and state of health

DETAILED CLINICAL OBSERVATIONS: No

OPHTHALMOSCOPIC EXAMINATION: No

BODY WEIGHT: Yes
- Time schedule for examinations: each week

HAEMATOLOGY: Yes
- Time schedule for collection of blood: after 28 day exposure period and after post-exposure observation period
- Anaesthetic used for blood collection: No data
- Animals fasted: No
- How many animals: all
- Parameters checked: leukocytes, erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelets, differential blood count, thromboplastin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: after 28 day exposure period and after post-exposure observation period
- Animals fasted: No
- How many animals: all
- Parameters checked: alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, sodium, potassium, chloride, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, cholesterol

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Other examinations:
None
Statistics:
Clinical examinations
Means and standard deviation were calculated for the variables (body weight and body weight gain/body weight change) of the animais of each test group for the statistical evaluation of the study. Statistical relevance was established using methods of ANOVA and DUNNETT.
Clinical chemistry and hematology
Mean and standard deviation were calculated for each test group and tabulated together with the individual values.
Except of the differential blood count, a non-parametric one-way analysis of variance is done via the Kruskal-Wallis-h-test. If the resulting p-value is equal or less than 0.05 a pairwise comparison of each dose group with the control group was carried out. This comparison is done using the Mann-Whitney-U-test for the hypotheses of equal medians.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
statistically significant retarded body weight change in male rats
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
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:
effects observed, treatment-related
Histopathological findings: neoplastic:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY:
80 mg/m³: no substance related effects
8 mg/m³: no substance related effects
0.8 mg/m³: no substance related effects
No mortalities were recorded throughout the exposure period.

BODY WEIGHT AND WEIGHT GAIN:
80 mg/m³: statistically significant retarded body weight change in male rats
8 mg/m³: no substance related effects
0.8 mg/m³: no substance related effects

HAEMATOLOGY:
all groups: no substance related effects

CLINICAL CHEMISTRY:
80 mg/m³: no substance related effects; 8 mg/m³: no substance related effects; 0.8 mg/m³: no substance related effects

ORGAN WEIGHTS:
80 mg/m³: increased absolute and relative lung weights (females and males); in the post exposure observation group a trend towards increased absolute and relative lung weights were still seen
8 mg/m³: increased absolute and relative lung weights which was still apparent in the post exposure observation group
0.8 mg/m³: no substance related effects

GROSS PATHOLOGY:
80 mg/m³: no substance related effects; 8 mg/m³: no substance related effects; 0.8 mg/m³: no substance related effects

HISTOPATHOLOGY:
80 mg/m³: lungs: hypertrophy of goblet cells (male and females), interstitial pneumonitis (increased in grading; males and females), increased connective tissue content (males and females)
nasal cavity (level 1): focal metaplasia of the respiratory epithelium (males and females)
post exposure observation period: increased connective tissue content lungs
8 mg/m³: no substance related effects
0.8 mg/m³: no substance related effects
Dose descriptor:
NOAEL
Remarks:
local
Effect level:
0.86 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
(equivalent to the substance as registered)
Sex:
male/female
Basis for effect level:
other: based on the absence of local effects
Dose descriptor:
LOAEL
Remarks:
local
Effect level:
8.1 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
(equivalent to the substance as registered)
Sex:
male/female
Basis for effect level:
other: based on the effects on lung weight
Dose descriptor:
NOAEL
Remarks:
systemic
Effect level:
82 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
(equivalent to the substance as registered)
Sex:
male/female
Basis for effect level:
other: based on the absence of toxicologically significant systemic effects
Critical effects observed:
not specified
Conclusions:
In a repeated dose inhalation toxicity study conducted according to OECD Guideline 412 (1990), groups of male and female Wistar rats (5/sex) were exposed by the inhalation route to the test substance as dust aerosol for 6 hours per day on 5 days per week for 4 weeks (28-day test) to head/nose at concentrations of 0, 0.8, 8, 80 mg/m³ (nominal concentrations) or 0, 0.86, 8.1, 82 mg/m³ (analytical concentrations). Based on the results of this study, 82 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) systemic based on the absence of toxicologically significant systemic effects; 0,86 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) local and 8.1 mg/m³ of test item was established as the Lowest Observed Adverse Effect Level (LOAEL) local based on the effects on lung weight. Therefore, reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda is not classified for repeated dose toxicity according to the classification criteria of the Regulation (EC) 1272/2008 (CLP) and of the Directive 67/548/EEC.
Executive summary:

In a repeated dose inhalation toxicity study according to OECD Guideline 412 (1990), groups of male and female Wistar rats (5/sex) were exposed by the inhalation route to the test substance as dust aerosol for 6 hours per day on 5 days per week for 4 weeks (28-day test) to head/nose at concentrations of 0, 0.8, 8, 80 mg/m³ (nominal concentrations) or 0, 0.86, 8.1, 82 mg/m³ (analytical concentrations). Groups of 5 female rats were hold without further exposure for an about 4-week post-exposure observation period to obtain informations on persistency, progression or regression of effects.

No mortality and no clinical signs were reported throughout the study whatever the dose level. Body weight gain was reduced in high dose males only. There were no test item-related effects on hematology and clinical chemistry parameters at any dose levels. No macroscopic findings were reported at necropsy. An increase of absolute and relative lung weights in male animals was observed at 82 mg/m³ and in female animals at 8.1 and 82 mg/m³. Histopathologically, goblet cell hyperplasia was reported in the bronchi of both sexes at 82 mg/m³. Furthermore, the grade of severity of pneumonitis that occurred also in control animals was slightly increased in the animals at 82 mg/m³, as was there a slight increase in the content of connective tissue of the lungs. In the nasal cavity, focal metaplasia of the respiratory epithelium to stratifying epithelium was noted in the animals at 82 mg/m³ at the transition of the stratifying epithelium to respiratory epithelium. After a 4-week recovery period (females only), the lungs of the animals at 82 mg/m³ showed only a slight reparative increase of mature connective tissue. Focal metaplasia in the nasal cavity was evident in only 2 animals at 82 mg/m³.

Based on the results of this study, 82 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) systemic based on the absence of toxicologically significant systemic effects; 0,86 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) local and 8.1 mg/m³ of test item was established as the Lowest Observed Adverse Effect Level (LOAEL) local based on the effects on lung weight.

Therefore, reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda is not classified for repeated dose toxicity according to the classification criteria of the Regulation (EC) 1272/2008 (CLP) and of the Directive 67/548/EEC.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
82 mg/m³
Study duration:
subacute
Species:
rat
Quality of whole database:
Klimisch 2

Repeated dose toxicity: inhalation - local effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
no data available
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: The study is sufficiently documented. OECD Guideline study; analytical purity of test substance: 68,3%.
Qualifier:
according to
Guideline:
OECD Guideline 412 (Subacute Inhalation Toxicity: 28-Day Study)
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Dr. K Thomae GmbH
- Age at study initiation: about 7 weeks
- Weight at study initiation: males: mean = 230 g; females: mean = 177 g
- Housing: single
- Diet: ad libitum during post exposure observation period
- Water: ad libitumduring post exposure observation period

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
other: Inhalation: dust aerosol
Type of inhalation exposure:
nose/head only
Vehicle:
air
Remarks on MMAD:
MMAD / GSD: 0.8 mg/m³: 3.3 µm
8 mg/m³: 3.3 µm
80 mg/m³: 3.1 µm
Details on inhalation exposure:
GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: aerodynamic exposure apparatus (INA 60, Volume ca. 90 L, BASF AG)
- Method of holding animals in test chamber: The rats were restrained in exposure tubes (glass tubes), their snouts projected into the inhalation chamber and they thus inhaled the dust aerosol.
- System of generating particulates/aerosols: brush generator
- Temperature, humidity, pressure in air chamber: 22°C, 50% humidity
- Method of particle size determination: cascade imapactor

TEST ATMOSPHERE
- Brief description of analytical method used: Spectrophotometric analysis
- Samples taken from breathing zone: yes
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
no data available
Duration of treatment / exposure:
6 hours per day for 28 days
Frequency of treatment:
5 days per week
Remarks:
Doses / Concentrations:
0, 0.8, 8, 80 mg/m³
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0, 0.86, 8.1, 82 mg/m³
Basis:
analytical conc.
No. of animals per sex per dose:
5
groups of female rats (5 animals) were hold without further exposure for an about 4-week post exposure
Control animals:
yes, concurrent vehicle
Details on study design:
no data available
Positive control:
no data available
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations: mortality, behavior and state of health

DETAILED CLINICAL OBSERVATIONS: No

OPHTHALMOSCOPIC EXAMINATION: No

BODY WEIGHT: Yes
- Time schedule for examinations: each week

HAEMATOLOGY: Yes
- Time schedule for collection of blood: after 28 day exposure period and after post-exposure observation period
- Anaesthetic used for blood collection: No data
- Animals fasted: No
- How many animals: all
- Parameters checked: leukocytes, erythrocytes, hemoglobin, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, platelets, differential blood count, thromboplastin time

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: after 28 day exposure period and after post-exposure observation period
- Animals fasted: No
- How many animals: all
- Parameters checked: alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, sodium, potassium, chloride, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, cholesterol

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
HISTOPATHOLOGY: Yes
Other examinations:
None
Statistics:
Clinical examinations
Means and standard deviation were calculated for the variables (body weight and body weight gain/body weight change) of the animais of each test group for the statistical evaluation of the study. Statistical relevance was established using methods of ANOVA and DUNNETT.
Clinical chemistry and hematology
Mean and standard deviation were calculated for each test group and tabulated together with the individual values.
Except of the differential blood count, a non-parametric one-way analysis of variance is done via the Kruskal-Wallis-h-test. If the resulting p-value is equal or less than 0.05 a pairwise comparison of each dose group with the control group was carried out. This comparison is done using the Mann-Whitney-U-test for the hypotheses of equal medians.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
statistically significant retarded body weight change in male rats
Food consumption and compound intake (if feeding study):
not specified
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
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:
effects observed, treatment-related
Histopathological findings: neoplastic:
effects observed, treatment-related
Details on results:
CLINICAL SIGNS AND MORTALITY:
80 mg/m³: no substance related effects
8 mg/m³: no substance related effects
0.8 mg/m³: no substance related effects
No mortalities were recorded throughout the exposure period.

BODY WEIGHT AND WEIGHT GAIN:
80 mg/m³: statistically significant retarded body weight change in male rats
8 mg/m³: no substance related effects
0.8 mg/m³: no substance related effects

HAEMATOLOGY:
all groups: no substance related effects

CLINICAL CHEMISTRY:
80 mg/m³: no substance related effects; 8 mg/m³: no substance related effects; 0.8 mg/m³: no substance related effects

ORGAN WEIGHTS:
80 mg/m³: increased absolute and relative lung weights (females and males); in the post exposure observation group a trend towards increased absolute and relative lung weights were still seen
8 mg/m³: increased absolute and relative lung weights which was still apparent in the post exposure observation group
0.8 mg/m³: no substance related effects

GROSS PATHOLOGY:
80 mg/m³: no substance related effects; 8 mg/m³: no substance related effects; 0.8 mg/m³: no substance related effects

HISTOPATHOLOGY:
80 mg/m³: lungs: hypertrophy of goblet cells (male and females), interstitial pneumonitis (increased in grading; males and females), increased connective tissue content (males and females)
nasal cavity (level 1): focal metaplasia of the respiratory epithelium (males and females)
post exposure observation period: increased connective tissue content lungs
8 mg/m³: no substance related effects
0.8 mg/m³: no substance related effects
Dose descriptor:
NOAEL
Remarks:
local
Effect level:
0.86 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
(equivalent to the substance as registered)
Sex:
male/female
Basis for effect level:
other: based on the absence of local effects
Dose descriptor:
LOAEL
Remarks:
local
Effect level:
8.1 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
(equivalent to the substance as registered)
Sex:
male/female
Basis for effect level:
other: based on the effects on lung weight
Dose descriptor:
NOAEL
Remarks:
systemic
Effect level:
82 mg/m³ air (analytical)
Based on:
test mat.
Remarks:
(equivalent to the substance as registered)
Sex:
male/female
Basis for effect level:
other: based on the absence of toxicologically significant systemic effects
Critical effects observed:
not specified
Conclusions:
In a repeated dose inhalation toxicity study conducted according to OECD Guideline 412 (1990), groups of male and female Wistar rats (5/sex) were exposed by the inhalation route to the test substance as dust aerosol for 6 hours per day on 5 days per week for 4 weeks (28-day test) to head/nose at concentrations of 0, 0.8, 8, 80 mg/m³ (nominal concentrations) or 0, 0.86, 8.1, 82 mg/m³ (analytical concentrations). Based on the results of this study, 82 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) systemic based on the absence of toxicologically significant systemic effects; 0,86 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) local and 8.1 mg/m³ of test item was established as the Lowest Observed Adverse Effect Level (LOAEL) local based on the effects on lung weight. Therefore, reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda is not classified for repeated dose toxicity according to the classification criteria of the Regulation (EC) 1272/2008 (CLP) and of the Directive 67/548/EEC.
Executive summary:

In a repeated dose inhalation toxicity study according to OECD Guideline 412 (1990), groups of male and female Wistar rats (5/sex) were exposed by the inhalation route to the test substance as dust aerosol for 6 hours per day on 5 days per week for 4 weeks (28-day test) to head/nose at concentrations of 0, 0.8, 8, 80 mg/m³ (nominal concentrations) or 0, 0.86, 8.1, 82 mg/m³ (analytical concentrations). Groups of 5 female rats were hold without further exposure for an about 4-week post-exposure observation period to obtain informations on persistency, progression or regression of effects.

No mortality and no clinical signs were reported throughout the study whatever the dose level. Body weight gain was reduced in high dose males only. There were no test item-related effects on hematology and clinical chemistry parameters at any dose levels. No macroscopic findings were reported at necropsy. An increase of absolute and relative lung weights in male animals was observed at 82 mg/m³ and in female animals at 8.1 and 82 mg/m³. Histopathologically, goblet cell hyperplasia was reported in the bronchi of both sexes at 82 mg/m³. Furthermore, the grade of severity of pneumonitis that occurred also in control animals was slightly increased in the animals at 82 mg/m³, as was there a slight increase in the content of connective tissue of the lungs. In the nasal cavity, focal metaplasia of the respiratory epithelium to stratifying epithelium was noted in the animals at 82 mg/m³ at the transition of the stratifying epithelium to respiratory epithelium. After a 4-week recovery period (females only), the lungs of the animals at 82 mg/m³ showed only a slight reparative increase of mature connective tissue. Focal metaplasia in the nasal cavity was evident in only 2 animals at 82 mg/m³.

Based on the results of this study, 82 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) systemic based on the absence of toxicologically significant systemic effects; 0,86 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) local and 8.1 mg/m³ of test item was established as the Lowest Observed Adverse Effect Level (LOAEL) local based on the effects on lung weight.

Therefore, reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda is not classified for repeated dose toxicity according to the classification criteria of the Regulation (EC) 1272/2008 (CLP) and of the Directive 67/548/EEC.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEC
0.86 mg/m³
Study duration:
subacute
Species:
rat
Quality of whole database:
Klimisch 2

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Repeated dose toxicity: oral

One study (Klimish score 2) was available on the substance and was used as a key study:

In a repeated dose toxicity study by the oral route, the test item was administered in drinking water to rats (10 animals/sex/dose) at doses of 0; 100; 500 and 2500 ppm during 5 weeks.

Food and water intake, general behaviour and mortality were not affected by treatment. Body weight gain was slightly reduced only in male rats administered 2500 ppm (not statistically significant) but this variation was considered of no toxicological relevance owing to its minimal magnitude (maximum 6% v.s. control) and absence of associated test item-related variations. Up to 2500 ppm, hematological, plasma chemistry and urinary parameters were not affected by the test item. No test item-related effects were reported at necropsy or during histopathological examinations up to the highest dose level.

Based on the results of this study, 2500 ppm of test item, which corresponds to 317.05 and 335.22 mg/kg b.w./day in males and females, respectively, was established as the No Observed Adverse Effect Level (NOAEL). Therefore, reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda is not classified for repeated dose toxicity according to the classification criteria of the Regulation (EC) 1272/2008 (CLP) and of the Directive 67/548/EEC.

Repeated dose toxicity: inhalation

One study (Klimish score 2) was available on the substance and was used as a key study:

In a repeated dose inhalation toxicity study according to OECD Guideline 412 (1990), groups of male and female Wistar rats (5/sex) were exposed by the inhalation route to the test substance as dust aerosol for 6 hours per day on 5 days per week for 4 weeks (28-day test) to head/nose at concentrations of 0, 0.8, 8, 80 mg/m³ (nominal concentrations) or 0, 0.86, 8.1, 82 mg/m³ (analytical concentrations). Groups of 5 female rats were hold without further exposure for an about 4-week post-exposure observation period to obtain informations on persistency, progression or regression of effects.

No mortality and no clinical signs were reported throughout the study whatever the dose level. Body weight gain was reduced in high dose males only. There were no test item-related effects on hematology and clinical chemistry parameters at any dose levels. No macroscopic findings were reported at necropsy. An increase of absolute and relative lung weights in male animals was observed at 82 mg/m³ and in female animals at 8.1 and 82 mg/m³. Histopathologically, goblet cell hyperplasia was reported in the bronchi of both sexes at 82 mg/m³. Furthermore, the grade of severity of pneumonitis that occurred also in control animals was slightly increased in the animals at 82 mg/m³, as was there a slight increase in the content of connective tissue of the lungs. In the nasal cavity, focal metaplasia of the respiratory epithelium to stratifying epithelium was noted in the animals at 82 mg/m³ at the transition of the stratifying epithelium to respiratory epithelium. After a 4-week recovery period (females only), the lungs of the animals at 82 mg/m³ showed only a slight reparative increase of mature connective tissue. Focal metaplasia in the nasal cavity was evident in only 2 animals at 82 mg/m³.

Based on the results of this study, 82 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) systemicbased on the absence of toxicologically significant systemic effects; 0,86 mg/m³ of test item was established as the No Observed Adverse Effect Level (NOAEL) local and 8.1 mg/m³ of test item was established as the Lowest Observed Adverse Effect Level (LOAEL) local based on theeffects on lung weight.

Therefore, reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda is not classified for repeated dose toxicity according to the classification criteria of the Regulation (EC) 1272/2008 (CLP) and of the Directive 67/548/EEC.


Justification for selection of repeated dose toxicity via oral route - systemic effects endpoint:
The selected study is the only study available for the oral route.

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
The selected study is the only study available for the inhalation route.

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:
The selected study is the only study available for the inhalation route.

Justification for classification or non-classification

Harmonized classification:

No harmonized classification is available according to the Regulation (EC) No 1272/2008 including ATP3.

Self classification:

Reaction product of naphthalene, butanol, sulfonated and neutralized by caustic soda is not classified for repeated dose toxicity according to the Regulation (EC) 1272/2008 (CLP) and to the Directive 67/548/EEC.