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Toxicological information

Acute Toxicity: inhalation

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Administrative data

Endpoint:
acute toxicity: inhalation
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2004

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 403 (Acute Inhalation Toxicity)
Deviations:
no
GLP compliance:
yes
Test type:
standard acute method
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Test compound: Thioglycolic acid
CAS no.: 68-11-1
Source: Bruno Bock Chemische Fabrik GmbH & Co KG
Batch: 9162
Purity: > 99%
Storage: Room temperature / darkness
Stability: Certified for the duration of this study

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
Test animals:
- Species: Rat Strain: Wistar
- Age: ca. 2 months old
- Weight at dosing: ca. 200 g
- Source: Harlan-Winkelmann GmbH, Borchen (Germany)
- Acclimation period: at least 5 days
- Diet: Kliba 3883, ad libitum
- Water: Tap water, ad libitum
- Housing: Animals were housed singly in Makrolon type IIIH cages

Environmental conditions:
- Temperature: 22 +/- 2°C
- Humidity: 40-60%
- Air changes: ca. 10 air changes/h
- Photoperiod: 12h/12h

Administration / exposure

Route of administration:
inhalation: aerosol
Type of inhalation exposure:
nose only
Vehicle:
other: unchanged (no vehicle)
Details on inhalation exposure:
Aerosol Generation and Exposure Technique:
- Mode of exposure:  Animals were exposed to the aerosolized test substance in Plexiglas  exposure tubes applying a directed-flow nose-only exposure principle. 
- Aerosol generation:  Under dynamic conditions the aqueous solution of the test article was  nebulized into a baffle (pre-separator) from which the substance was  conveyed into the intake of the cylindrical inhalation chamber. The test  article was nebulized using a binary nozzle with conditioned compressed  air. The solution was fed into the nozzle by a calibrated, digitally  controlled piston, equipped with a glass syringe.  Inhalation Chamber: The stainless steel directed-flow nose-only  inhalation chamber has the following dimensions: diameter = 30 cm  (two-chamber system), height = 28 cm (internal volume = about 7 L). 
- Inhalation chamber steady-state concentration:  The test atmosphere generation conditions provide an adequate number of  air exchanges per hour (ca. 129 x, continuous generation of test  atmosphere). Under such test conditions steady state is attained within  the first two minutes of exposure (t99% = 4.6 x chamber volume/flow rate;  McFarland, 1976). The ratio between the air supplied and exhausted was  chosen so that approximately 80-90% of the supplied air is removed via  the exhaust system. The remainder provides adequate dead-space  ventilation for the exposure tubes. At each exposure port a minimal air  flow rate of 0.75 l/min was provided. The test atmosphere can by no means  be diluted by bias-air-flows. The inhalation chamber was operated in a  well ventilated chemical fume hood.
- Air flows:  During the exposure period air flows were monitored continuously and, if  necessary, readjusted to the conditions required. 

Inhalation Chamber Temperature and Humidity:
Temperature and humidity measurements were made using a computerized  system. The values were recorded at intervals of 5 min.

Analysis of the Test Atmosphere:
- Nominal concentration:  The nominal concentration was calculated from the ratio of the total  quantity of test substance discharged during the exposure period and the  total throughput of air through the inhalation chamber.
- Gravimetric evaluation:  The test-substance concentration was determined by gravimetric analysis.  Filters were weighed immediately after cessation of sampling.  
- Analytical evaluation: Several samples were taken by bubbling the  inhalation chamber atmosphere through gas bubblers containing an aqueous  solution (0.04%) of phosphoric acid. The analytes were determined by  High-Performance Chromatography (HPLC).

Characterization of Aerodynamic Particle-Size Distribution:
The samples for the analysis of the particle-size distribution were also  taken in the vicinity of the breathing zone. During each exposure two  samples were taken. The particle-size distribution was analyzed using a BERNER-TYPE AERAS  low-pressure critical orifice cascade impactor (Hauke, Gmunden, Austria). 
Analytical verification of test atmosphere concentrations:
yes
Duration of exposure:
4 h
Concentrations:
0, 284.1, 837.2, 1441 and 3629 mg/m3
No. of animals per sex per dose:
5
Control animals:
yes
Details on study design:
Body Weights:
Body weights were measured before exposure, on days 3 and 7, and weekly  thereafter. 

Clinical Signs:
The appearance and behaviour of each rat were examined carefully several  times on the day of exposure and at least once daily thereafter.

Rectal Temperatures:
The rectal temperatures were measured shortly after cessation of exposure  (approximately within ½ hour after the end of exposure) using a digital  thermometer with a rectal probe for rats.

Necropsy:
All surviving rats were sacrificed at the end of the observation period  and were subject to a gross-pathological examination.
Statistics:
Necropsy findings: pair-wise Fisher test after the R x C chi-squared  test. The Fisher test was only performed if differences occurred between  groups in the R x C chi-squared test or if a frequency value of < 5 was  calculated.  Body weights: one-way ANOVA (vide infra). Physiological data: ANOVA procedure (vide infra). Initial LC50 calculation by the author used moving average Interpolation according to  Scharper et al. (1994). LC50 was recalculated using a probit standard model.

Results and discussion

Effect levelsopen allclose all
Sex:
male/female
Dose descriptor:
LC50
Effect level:
1 388 mg/m³ air (analytical)
Based on:
test mat.
95% CL:
1 000 - 2 080
Exp. duration:
4 h
Remarks on result:
other: calculated with probit standard model
Sex:
male
Dose descriptor:
LC50
Effect level:
1 981 mg/m³ air (analytical)
Based on:
test mat.
95% CL:
921 - 11 700
Exp. duration:
4 h
Remarks on result:
other: calculated with probit standard model
Sex:
female
Dose descriptor:
LC50
Effect level:
1 094 mg/m³ air (analytical)
Based on:
test mat.
95% CL:
98 - 2 130
Exp. duration:
4 h
Remarks on result:
other: calculated with probit standard model
Mortality:
Mortality occurred at exposure concentrations equal to and exceeding 1441  mg/m³. In the range of the LC50-concentration, female rats appeared to be  more susceptible when compared to male rats. Mortality occurred up to the  first postexposure day.
Clinical signs:
Group 1: All rats tolerated the exposure without specific signs.
Group 2/males: Nose: reddened.
Group 3/males: bradypnea, labored breathing patterns, irregular breathing patterns, nose: reddened, nose: red encrustations, nostrils: red encrustations, piloerection, hair-coat ungroomed, motility reduced.
Group 4/males: Bradypnea, labored breathing patterns, nose: reddened, nose: red encrustations, muzzle: red encrustations, piloerection, hair-coat ungroomed, motility reduced, limp, paralysis (all legs), high-legged gait.
Group 5/males: Bradypnea, labored breathing patterns, nose: reddened, nose: red encrustations, muzzle: red encrustations, piloerection, motility reduced, limp, high-legged gait, flaccidity (all limbs).
Group 2/females: Bradypnea, labored breathing patterns, nose: reddened, nostrils: red encrustations, piloerection, hair-coat ungroomed, cyanosis, motility reduced, limp, flaccidity (all limbs) , posture squatted.
Group 3/females:Bradypnea, labored breathing patterns, irregular breathing patterns, tachypnea, nasal discharge (serous), nose: reddened, nose: red encrustations, muzzle: red encrustations, nostrils: red encrustations, stridor, piloerection, hair-coat ungroomed, cyanosis, emaciation, motility reduced, limp, apathy, tremor (hind limbs), paralysis (all legs), flaccidity (all limbs) , prostration, corneal opacity.
Group 4/females: Bradypnea, labored breathing patterns, nose: reddened, pilo-erection, motility reduced, limp, high-legged gait, flaccidity (all limbs).
Group 5/females: Bradypnea, labored breathing patterns, nose: reddened, nose: gray discolorations, nose: red encrustations, nostrils: red encrustations, piloerection, motility reduced, limp, high-legged gait, flaccidity (all limbs) , tremor (entire body).
Body weight:
Comparisons between the control and exposure group revealed transient changes in body weights at 837.2 and 1441 mg/m3 (see attached figure).
Gross pathology:
Animals succumbed during the observation period:  Nose: red; lung: less collapsed; dark-red marbled stomach: bloated, red  mucosa; intestines: bloated, reddish-mucous content, mucosa reddened;  liver: light coloured; spleen: light coloured; kidneys: bilaterally light  coloured; incisors: white discolouration; oral cavity: filled with  bedding material; oesophagus: filled with bedding material. Animals sacrificed at the end of the observation period: The macroscopic  findings in the exposure groups showed a low incidence of discolourated  lungs.
Other findings:
Reflex measurements
A battery of reflex measurements was made on the first post-exposure day. In comparison to the rats of the control group, male rats of group 4 and some female rats in groups 2-3 exhibited changes in the reflex behavior suggestive impaired reflexes in the first post-exposure day.

Rectal temperature
Mean values are shown in Table 2. Statistical comparisons between the control and the exposure groups 3-5 revealed significantly decreased the body temperatures shortly after cessation of exposure.

Any other information on results incl. tables

Table 1: Generation and characterization of chamber atmosphere (aerosolization of the neat test article) - Mean values

Group 1

Group 2

Group 3

Group 4

Group 5

Target Conc. (mg/m³)

Control (air)

250

1200

1500

5000

Nominal concentration (mg/m³)a

0

1408

5632

11440

35200

Gravimetric Conc. (mg/m³)

--

196.6

837.4

1542

3865

Analytical concentration (mg/m³)

--

284.1

837.2

1441

3629

Gravimetric:analytical (%)

--

69

100

»100

»100

Recovery (%)

--

20

15

13

10

Test article supply (ml/min)

--

16

64

130

400

Inlet Air Flow (l/min)

15

15

15

15

15

Exhaust Air Flow (l/min)

13

13

13

13

13

Temperature (mean,oC)

21.8

22.7

22.6

22.6

22.6

Rel. Humidity (mean, %)

< 6.5

< 5.4

< 5

< 5.2

< 5.3

MMAD (µm)
GSD

--
--

2.48
2.20

2.85
2.15

2.77
2.01

3.07
2.01

Aerosol Mass < 3 µm (%)

--

59.8

53

54.6

49

Mass recovered (mg/m³)

--

136.7

799.2

1310

3926

MMAD = Mass Median Aerodynamic Diameter, GSD = Geometric Standard Deviation; -- = not applicable. Recovery: Analytical concentration relative to nominal concentration. a) For calculation a specific density D204= 1.32 g/cm³ was used.

Table 2: Summary of acute inhalation toxicity - 4 hour exposure to aerosolized test substance

N
Group
/sex

Actual

Concentration (mg/m³)

Toxicological Result

Onset and Duration of Signs

Onset of Mortality

Rectal Temperature (°C)

1 / m

0

0 / 0 / 5

--

--

37.6

2 / m

284.1

0 / 5 / 5

0d (5h)

--

37.3

3 / m

837.2

0 / 5 / 5

0d – 2d

--

35.3

4 / m

1440.9

1 / 5 / 5

0d–4d,7d–14d

1d

31.1*

5 / m

3628.8

5 / 4 / 5

0d

0d, 1d

28.7**

1 / f

0

0 / 0 / 5

--

--

38.4

2 / f

284.1

0 / 5 / 5

0d (5h) – 3d

--

37.5

3 / f

837.2

0 / 5 / 5

0d – 12d

--

36.9*

4 / f

1440.9

5 / 5 / 5

0d

1d

35.2**

5 / f

3628.8

5 / 5 / 5

0d

1d

28.8**

N = group assignment, m = males, f = females, * = p < 0.05, ** = p < 0.01

Values given in the 'Toxicological results' column are:

    1st = number of dead animals.

    2nd = number of animals with signs after cessation of exposure.

    3rd = number of animals exposed. 

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
Based on the calculations obtained by the Moving Average Interpolation procedure, the approximate combined LC50 for males and females is 1388 mg/m³.
Executive summary:

A study on the acute inhalation toxicity of THIOGLYCOLIC ACID on rats has been conducted in accordance with OECD Guideline No. 403, EU Directive 92/69/EEC, OPPTS 870.1300 and Japan MAFF, Notification No. 12 Nousan-8147. Four groups of rats were nose-only expo­sed to a mean aerosol concentrations of 284, 837, 1441, and 3629 mg/m³ air. The liquid aerosol generated (aerosolization of the undiluted test article) was respirable to rats and all

concentration data represent actual concentrations of the test substance in the rats breathing zone.Internationally recognized recommendations such as of SOT (1992) were fulfilled, in regard to the respirability of the aerosol generated, i.e. the MMAD was <4 µm (MMAD 2.5-3.1 µm, GSD»2.1).

Mortality occurred at exposure concentrations equal to and exceeding 1441 mg/m³. In the range of the LC50-concentration, female rats appeared to be more susceptible when compared to male rats. Based on the calculations obtained by the probit standard modele, the approximate LC50for males and females is 1891 and 1094 mg/m³, respectively, and the combined LC50 is 1388 mg/m3 . Mortality occurred up to the first postexposure day. The clinical signs observed included the following findings: bradypnea, labored breathing patterns, irregular breathing patterns, tachypnea, nasal discharge (serous), nose: reddened, nose: gray discolorations, nose: red encrustations, muzzle: red encrustations, nostrils: red encrustations, stridor, piloerection, hair-coat ungroomed, cyanosis, emaciation, motility reduced, limp, apathy, tremor (hind limbs), paralysis (all legs), high-legged gait, flaccidity (all limbs) , posture squatted, prostration, corneal opacity, tremor (entire body), reduced reflexes, decreased body weights, and hypothermia. The duration of clinical signs was governed by respiratory distress and behavioral abnormalities. At the end of the postexposure period most rats appeared to be clinically normal. Necropsy findings were suggestive that lung damage was causally related to mortality. .