P 16513

Acute Toxicity: inhalation

Administrative data

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 November 2022 to 02 December 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Report date:

2023

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Version / remarks:

OECD Guidelines for Testing of Chemicals No. 403 – Traditional Protocol “Acute Inhalation Toxicity” adopted on 07 September 2009

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

traditional method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Strontium 4-chloro-2-[2-hydroxy-6-sulfonatonaphthalen-1-yl)diazenyl]benzoate

Cas Number:

474814-88-5

Molecular formula:

C17H9ClN2O6SSr

IUPAC Name:

Strontium 4-chloro-2-[2-hydroxy-6-sulfonatonaphthalen-1-yl)diazenyl]benzoate

Test material form:

solid: nanoform, no surface treatment

Details on test material:

red pigment

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: In-house bred
- Females: nulliparous and non-pregnant: yes
- Age at study initiation: 9 weeks
- Weight at study initiation: Males: 182.12 g to 192.56 g; Females: 168.08 g to 177.72 g

- Housing: Three animals per sex were housed in a standard polycarbonate cage (size: L 430 × B 280 × H 210 mm) with stainless steel mesh top grill having facilities for holding pelleted feed and drinking water in water bottle fitted with stainless steel sipper tube
- Diet (e.g. ad libitum): Altromin maintenance diet for rats and mice (manufactured by Altromin Spezialfutter GmbH & Co. KG)
- Water (e.g. ad libitum): Deep bore-well water passed through Reverse osmosis unit
- Acclimation period: 11 November 2022 to 17 November 2022

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.9°C to 22.6°C
- Humidity (%): 43% to 66%
- Air changes (per hr): 12 to 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours fluorescent light and 12 hours dark cycle

IN-LIFE DATES: From: 11 November 2022 To: 02 December 2022

Administration / exposure

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

clean air

Mass median aerodynamic diameter (MMAD):

> 2.87 - <= 3.27 µm

Geometric standard deviation (GSD):

> 2.63 - <= 2.72

Remark on MMAD/GSD:

within prescribed range as per guideline

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: nose-only dynamic inhalation exposure system
- Exposure chamber volume: 0.76
- Method of holding animals in test chamber: Restariner
- Source and rate of air (airflow): compressed air and 20 lt/min
- Method of conditioning air: filtered air
- System of generating particulates/aerosols: Rotating Brush Generator (Palas RBG 1000 - supplied by Palas GmbH)
- Method of particle size determination: Gravimetric method
- Treatment of exhaust air: NaOH
- Temperature, humidity, pressure in air chamber: 22.1°C to 22.4°C, 54.5% to 57.1%, 60 psi

TEST ATMOSPHERE
- Samples taken from breathing zone: yes
- Time needed for equilibrium of exposure concentration before animal exposure:
t95 (min) = 3× Chamber Volume / Chamber airflow
= 3× 0.76 L / 20 L/min
= 0.11 minutes

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

> 0 - <= 240 min

No. of animals per sex per dose:

Limit Test - 3 Males and 3 Females

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: once daily for clinical signs and twice daily for mortality and weighing day 1, 3, 7 and 14
- Necropsy of survivors performed: yes

Results and discussion

Effect levels

Key result

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.04 mg/L air

Based on:

test mat.

Mortality:

No Mortality

Clinical signs:

other: No treatment related clinical signs and mortality were observed at the mean maximum concentration of 5.04 mg/L of air

Body weight:

No treatment related changes were observed in body weight and percent change in body weight with respect to day 1 at the mean maximum concentration of 5.04 mg/L of air. However, all animals showed slight decrease in body weight on day 2 due to exposure and increased in body weight from day 4 onwards

Gross pathology:

No treatment related gross pathological findings were observed at the maximum concentration of 5.04 mg/L of air

Any other information on results incl. tables

• CLINICAL SIGNS AND MORTALITY RECORD

Group  & Concentration (mg/L of air)

Animal No.

Sex

Day 1

Days

During Exposure

Post exposure

1 hr*

2 hrs*

3 hrs*

4 hrs*

30-40 min

1 hr*

2

3

4

5

6

7

8

9

10

11

12

13

14

15

Limit Test & 5.04

Rh7087

M

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh7088

M

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh7089

M

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh7090

F

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh7091

F

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

Rh7092

F

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

N

              *: ± 10 minutes; N: Normal; M: Male; F: Female; min: minute; hr(s): hour(s)

                    

• BODY WEIGHT (g) AND PERCENT CHANGE IN BODY WEIGHT (%) WITH RESPECT TO DAY 1

Group &    Concentration (mg/L of air)	Animal No.	Sex	Body Weight (g) on Days						Percent Change in Body Weight with Respect to Day 1
			1^#	2	4	8	15		1-2	1-4	1-8	1-15
Limit Test & 5.04	Rh7087	M	207.86	204.36	209.23	222.34	246.58		-1.68	0.66	6.97	18.63
	Rh7088	M	211.49	209.04	213.69	228.58	253.46		-1.16	1.04	8.08	19.84
	Rh7089	M	218.65	214.98	219.05	233.63	256.61		-1.68	0.18	6.85	17.36
	Mean		212.67	209.46	213.99	228.18	252.22		-1.51	0.63	7.30	18.61
	(±) SD		5.49	5.32	4.92	5.66	5.13		0.30	0.43	0.68	1.24
	n		3	3	3	3	3		3	3	3	3
	Rh7090	F	187.09	186.15	188.13	197.18	214.78		-0.50	0.56	5.39	14.80
	Rh7091	F	194.92	192.21	195.06	206.09	225.19		-1.39	0.07	5.73	15.53
	Rh7092	F	196.73	194.70	197.51	208.44	223.28		-1.03	0.40	5.95	13.50
	Mean		192.91	191.02	193.57	203.90	221.08		-0.97	0.34	5.69	14.61
	(±) SD		5.12	4.40	4.87	5.94	5.54		0.45	0.25	0.28	1.03
	n		3	3	3	3	3		3	3	3	3

#: Prior to exposure; M: Male; F: Female; SD: Standard Deviation; n; Number of animals

• BREATHING ZONE CONCENTRATION (ACTUAL TEST ITEM CONCENTRATION)

 Technical Pre-test:

Sl. No.	Feed rate (mm/hour)	Rotation (rpm)	Initial weight (mg) (a)	Final weight (mg) (b)	Difference (mg) (c) = (b) – (a)	Air Flow Rate (L/min) (d)	Time (min) (e)	BZC (mg/L of air)	Mean BZC (mg/L of Air)
1	30	600	353.94	361.57	7.63	1.52	1	5.02	5.04
2	30	600	348.21	355.89	7.68	1.52	1	5.05

 Limit Test:

Sl. No.	Feed rate (mm/hour)	Rotation (rpm)	Initial weight (mg) (a)	Final weight (mg) (b)	Difference (mg) (c) = (b) – (a)	Air Flow Rate (L/min) (d)	Time (min) (e)	BZC (mg/L of air)	Mean BZC (mg/L of Air)
1	30	600	351.91	359.56	7.65	1.52	1	5.03	5.04
2	30	600	347.31	354.97	7.66	1.52	1	5.04
3	30	600	348.36	356.05	7.69	1.52	1	5.06

   BZC: Breathing Zone Concentration; Sampled volume: 1.52 L/min; Sampling time: 1 minute

 

BZC (f) =	Mass of test item collected on the filter paper (c)
	Volume of air passed through the filter paper (d) X time (e)

 

• CHAMBER (EXPOSURE) CONDITIONS

Technical Pre-test:

Concentration (mg/L of air)	SL. No.	Feed rate (mm/hour)	Rotation (rpm)	Temperature (°C)	Relative Humidity (%)	Oxygen Concentration (%)	Carbon dioxide Concentration (ppm)	Air Flow (L/min)
5.04	1	30	600	22.4	56.1	20.1	615	20
	2	30	600	22.2	57.2	20.3	618	20

 

Limit Test:

Concentration (mg/L of air)	SL. No.	Feed rate (mm/hour)	Rotation (rpm)	Temperature (°C)	Relative Humidity (%)	Oxygen Concentration (%)	Carbon dioxide Concentration (ppm)	Air Flow (L/min)
5.04	1	30	600	22.3	57.1	20.2	614	20
	2	30	600	22.1	56.2	20.1	612	20
	3	30	600	22.4	55.8	20.3	615	20
	4	30	600	22.2	54.5	20.4	618	20

Note: 1% Carbon dioxide = 10000 ppm

 

• SUMMARY OF CHAMBER (EXPOSURE) CONDITIONS

 

Technical Pre-Test
Chamber Conditions	Range
Feed Rate (mm/hour) and Speed (RPM)	30 and 600
Temperature (°C)	22.2 to 22.4
Relative humidity (%)	56.1 to 57.2
Oxygen concentration (%)	20.1 to 20.3
Carbon dioxide concentration (ppm)	615 to 618
Air Flow (L/min)*	20
BZC (mg/L)	5.02 to 5.05
MMAD (µm)	3.08 ^#
GSD	2.69 ^#
Limit Test
Feed Rate (mm/hour) and Speed (RPM)	30 and 600
Temperature (°C)	22.1 to 22.4
Relative humidity (%)	54.5 to 57.1
Oxygen concentration (%)	20.1 to 20.4
Carbon dioxide concentration (ppm)	612 to 618
Air Flow (L/min)*	20
BZC (mg/L)	5.03 to 5.06
MMAD (µm)	2.87 to 3.27
GSD	2.63 to 2.72

*: Values were constant throughout the exposure; ^#: Individual value; BZC: Breathing Zone Concentration (Actual Concentration), MMAD: Mass Median Aerodynamic Diameter, GSD: Geometric Standard Deviations.

 

• GROSS PATHOLOGICAL FINDINGS

Group & Concentration (mg/L of air)	Animal No.	Sex	Fate	Gross Pathology Findings
				External	Internal
Limit Test & 5.04	Rh7087	M	TS	NAD	NAD
	Rh7088	M	TS	NAD	NAD
	Rh7089	M	TS	NAD	NAD
	Rh7090	F	TS	NAD	NAD
	Rh7091	F	TS	NAD	NAD
	Rh7092	F	TS	NAD	NAD

  NAD: No Abnormality Detected; M: Male; F: Female; TS: Terminal Sacrifice

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Remarks:

Based on the results of this study the test item has not to be classified for acute inhalation toxicity according to Regulation (EC) No 1272/2008.

Conclusions:

Under the experimental conditions employed and based on the above results of experiment, there were no clinical signs and mortality observed at mean maximum concentration of 5.04 mg/L of air. Hence, the LC50 of the test item is > 5.04 mg/L of air.

Executive summary:

The test item was evaluated for acute inhalation toxicity in Sprague Dawley rats.

The objective of the study was to assess the toxic potential and to determine the LC₅₀ of test item when administered by inhalation route through flow-past nose-only dynamic inhalation equipment for a single 4 hours exposure to rats. Three male and three female rats were used for conducting the acute inhalation toxicity study.

As such test item was used during technical pre-test and limit test to generate the dust aerosols through rotating brush generator. The technical pre-test was carried out without animals. During the technical pre-test, the target concentration i.e.  5.04 mg/L of air was achieved at the feed rate of 30 mm/hour with a rotation of 600 RPM.

During the exposure period, the temperature, relative humidity, oxygen and carbon dioxide concentration of the chamber was found within the required range. The particle size MMAD and GSD values were within the required range at the mean maximum breathing zone concentration (actual concentration) of 5.04 mg/L and it was considered as the limit concentration.

All animals were observed for clinical signs of toxicity and mortality during exposure and post exposure on day 1 and once daily thereafter for clinical signs and twice daily for mortality for 14 days for post exposure period. Individual animal body weight was recorded on day 1 (on the day of exposure) prior to the exposure and on day 2, 4, 8 and 15. All surviving rats were euthanized after 14 days post exposure by intraperitoneal administration of sodium thiopentone and they were subjected to gross necropsy.

No treatment related clinical signs of toxicity and mortalities were observed. Slight decrease in body weight was noted on Day 2 due to exposure. All animals showed an increase in body weight on day 4, 8 and 15.

No treatment related gross pathological findings were observed at the mean maximum concentration of 5.04 mg/L of air.