Fatty acids, C16-18 (even numbered), aluminum salts

Endpoint summary

Description of key information

Oral, rat: LD50: > 2000 mg/kg bw 
Inhalation, rat: LC50: > 5.15 mg/L 

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

08 - 25 Jan 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP - Guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, München, Germany

Test type:

acute toxic class method

Limit test:

yes

Species:

rat

Strain:

other: WISTAR Crl: WI (Han)

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Sulzfeld, Germany
- Age at study initiation: 8 - 9 weeks
- Weight at study initiation: step 1: 156 - 160 g; step 2: 137 - 145 g
- Fasting period before study: animals were fasted 16 - 19 h prior to administration (access to water was permitted) until 4 h after dosing.
- Housing: animals were kept in groups in IVC cages, type III, polysulphone cages on Altromin saw fibre bedding (lot No. 160812)
- Diet: Altromin 1234 maintenance diet for rats and mice (lot No. 0636), ad libitum
- Water: tap water (municipal residue control) sulphur acidified to a pH of approx. 2.8
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 10
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: From: 3 Jan To: 25 Jan 2013

Route of administration:

oral: gavage

Vehicle:

cotton seed oil

Details on oral exposure:

VEHICLE
- Amount of vehicle (if gavage): 10 mL/kg bw
- Justification for choice of vehicle: the vehicle was chosen due to it´s known non-toxic properties and the insolubility of the test substance in water.
- Lot/batch no. (if required): MKBJ0602V


MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

DOSAGE PREPARATION (if unusual):
The test substance was ground in a mortar prior to dissolving in cotton seed oil. Homogeneity of the suspension was maintained by vortexing during the administration procedure.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: according to the toxic class method

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

3 (step 1)
3 (step 2)

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: animals were observed several times on the day of dosing and daily thereafter and individual body weights were determined on Day 1 prior to treatment and on Day 8 and 15.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weight, gross pathology, other: changes in skin and fur, eyes and mucous membranes, respiratory, circulatory, autonomic and central nervous system and behaviour pattern. Special emphasis was directed on tremor, convulsions, salivation, diarrhoea, lethargy, sleep and coma.

Sex:

female

Dose descriptor:

LD50

Effect level:

>= 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality occured during the study period.

Clinical signs:

other: No clinical signs of toxicity were observed up to the end of the 14-day observation period.

Gross pathology:

Necropsy revealed no substance-related findings despite acute injection of blood vessels in the abdominal region caused by euthanasia injection.

Other findings:

No other finfings are reported in the study report.

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

The available information comprises an adequate and reliable study (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex VII, 8.5, of Regulation (EC) No 1907/2006.

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

24 Jan - 28 Oct 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

GLP-Guideline study (Initially, rats were exposed to the limit dose of 5 mg/L according to the traditional protocol described in OECD 403. Based on the mortality determined in males, further testing at 1 mg/L was performed. Additionally, a second experiment was conducted including 5 male rats exposed to 5 mg/L to verify the determined LC50 value in the first experiment).

Qualifier:

according to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Deviations:

no

Remarks:

Initially, 5 mg/L were tested as limit dose according to the traditional protocol. Based on the mortality determined in males, further testing at 1 mg/L was performed followed by a second exposure to 5 mg/L of male rats.

GLP compliance:

yes

Test type:

standard acute method

Limit test:

yes

Species:

rat

Strain:

other: Sprague-Dawley derived, albino

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories, Inc.
- Age at study initiation: first experiment, 5 mg/L: 9 - 11 weeks, 1 mg/L: 9 - 11 weeks; second experiment, 5 mg/L: 8 - 9 weeks
- Weight at study initiation: first experiment, 5 mg/L: 253 - 272 g (m), 190 - 220 g (f), 1 mg/L: 284 - 306 g (m); second experiment, 5 mg/L: 241 - 275 g (m)
- Housing: animals were housed singly in suspended stainless steel cages with mesh floors including enrichment (e.g. toys).
- Diet: Harlan Teklad Global 16% Protein Rodent Diet® #2016, ad libitum (except during exposure)
- Water: filtered tap water, ad libitum (except during exposure)
- Acclimation period: at least 8 days (first experiment: 8 or 23 days; second experiment: 12 days)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 23
- Humidity (%): 38 - 69
- Air changes (per hr): 12- 13
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: first experiment (5 mg/L): From: 24 Jan To: 7 Feb 2013
first experiment (1 mg/L): From: 7 Jun To: 21 Jun 2013
second experiment (5 mg/L): From: 11 Oct To: 28 Oct 2014

Route of administration:

inhalation: aerosol

Type of inhalation exposure:

nose only

Vehicle:

air

Details on inhalation exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: Nose-Only Inhalation Chamber, ADG Developments LTD
- Exposure chamber volume: 6.7 L (5.0 mg/L exposure) or 28 L (1.0 mg/L exposure)
- Method of holding animals in test chamber: polycarbonate holding tubes sealed to the chamber with an “O” ring during exposure. The base unit terminated the chamber with a 0.5- inch diameter tube for discharged air.
- Source and rate of air: filtered and compressed air (air compressor: Powerex Model: SES050822). Additionally, compressed and mixed air from the air compressor was introduced into the chamber to enable uniform distribution of the test atmosphere by creating a vortex at the chamber inlet. Compressed airflow was measured with a Mass Flow Meter (Omega, Model #FMA-5613) or Mass Flow Controller (Omega, Model #FMA-5541). Chamber airflow was monitored throughout each exposure period and recorded periodically. Each exposure was conducted under slightly negative pressure.
- System of generating particulates/aerosols: Dust Generator (Fluid Energy Jet Mill, Serial # J2724E) with an Accurate Series 100 Dry Material Feeder Apparatus. Compressed generator/mixing air was supplied to the dust generator at 40/30 psi, respectively. The aerosolised dust was then fed directly into the chamber through the dust outlet assembly.
- Method of particle size determination: eight-stage 1 ACFM Anderson Ambient Particle Sizing Sampler. Samples were withdrawn from the breathing zone of the animals at two intervals. The aerodynamic mass median diameter and geometric standard deviation were determined graphically using two-cycle logarithmic probit axes.
- Treatment of exhaust air: filtered
- Temperature and humidity in chamber: first experiment, 5 mg/L: 17 - 19 °C, 18 - 21%, 1 mg/L: 21 - 22 °C, 56 - 61%; second experiment, 5 mg/L: 19 - 21 °C, 44 - 49%

TEST ATMOSPHERE
- Brief description of analytical method used: indirect: gravimetric analysis. Samples were withdrawn at six intervals from the breathing zone of the animals during each exposure. Samples were collected using 37 mm glass fiber filters (GF/B Whatman™) in a filter holder attached by ¼ inch Tygon® tubing to a vacuum pump (GAST, Model #1531-107B-G557X or Westech, Model #9803-88). Filter papers were weighed before and after collection to determine the mass collected. This value was divided by the total volume of air sampled to determine the chamber concentration. Sample airflows were measured using a Mass Flow Controller (Aalborg, Model #GFC-17).
- Samples taken from breathing zone: yes

VEHICLE
- Composition of vehicle: air
- Concentration of test material in vehicle (actual concentration): first experiment: 1.03 mg/L and 5.15 mg/L; second experiment: 5.17 mg/L

TEST ATMOSPHERE (if not tabulated)
- Particle size distribution: please refer to Table 2
- MMAD (Mass median aerodynamic diameter) / GSD (Geometric st. dev.): please refer to Table 4

REMARK ON TEST CONCENTRATIONS/STUDY DESIGN
According to OECD 403, a limit test was initially conducted with a start concentration of 5 mg/L as the test article is known or expected to be virtually non-toxic. Based on the mortality determined in the test group of males, further testing at 1 mg/L was performed in male rats as the reason for mortality remained unclear especially as the test item does not seem to have intrinsic properties leading to toxicity . With the intention of collecting additional data and to verifiy the LC50, additional 5 males were exposed to 5.0 mg/L in a second experiment.

Analytical verification of test atmosphere concentrations:

yes

Remarks:

gravimetric analyses of samples withdrawn from the breathing zone at 6 intervals during the exposure

Duration of exposure:

>= 240 min

Remarks on duration:

the exposure period was extended beyond 4 h to allow the chamber to reach equilibrium (T 99) (please refer to Table 3).

Concentrations:

first experiment, 5 mg/L: 5.15 ± 0.13 mg/L (analytical concentration), nominal chamber concentration: 19.28 mg/L;
first experiment, 1 mg/L: 1.03 ± 0.02 mg/L (analytical concentration), nominal chamber concentration: 5.29 mg/L;
second experiment, 5 mg/L: 5.17 ± 0.31 mg/L (analytical concentration), nominal chamber concentration: 21.14 mg/L;

No. of animals per sex per dose:

first experiment, 5 mg/L: 5 males and females;
first experiment, 1 mg/L: 5 males;
second experiment, 5 mg/L: 5 males;

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: animals were observed daily, including gross evaluation of skin and fur, eyes and mucous membranes, respiratory, circulatory, autonomic and central nervous systems, somatomotor activity and behaviour pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhea, and coma. Individual body weights were determined prior to exposure and on Day 1, 3, 7 and 14.
- Necropsy of survivors performed: yes (tissues and organs of the thoracic and abdominal cavities)
- Necropsy of deceased performed: yes (tissues and organs of the thoracic and abdominal cavities)

Statistics:

Means and standard deviations of the examined parameter were calculated.

Sex:

female

Dose descriptor:

LC50

Effect level:

> 5.15 mg/L air (analytical)

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

other: 0/5 females died until the end of the observation period

Sex:

male

Dose descriptor:

LC50

Effect level:

> 5.17 mg/L air (analytical)

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

other: 3/10 males died until the end of the observation period (based on a weight-of-evidence approach of the first and second experiment, in which 3/5 and 0/5 males died, respectively)

Mortality:

In the first experiment, no mortality was observed in the female group whereas 3/5 males were found dead at chamber removal after exposure to 5.15 mg/L. In the subsequent exposure to 1 mg/L, no further male died. In the second experiment no male died until the end of the observation period. In summary of both experiments following a weight-of-evidence approach, 0/5 females and 3/10 males died following acute exposure to 5 mg/L test substance following inhalation.

Clinical signs:

other: In the first experiment, deceased males (3/5) were found dead at chamber removal and hence, no findings on clinical symptoms were reported. Surviving males exhibited irregular respiration lasting until 1.5 h after chamber removal (2/2 males), dry rales at

Body weight:

In the first experiment, exposure to 5 mg/L induced body weight loss in all animals through Day 3 but most animals surpassed their initial body weight by Day 7 and gained body weight through Day 14. For the remaining animals, the initial body weight was surpassed until the end of the observation period (please refer to Table 6). Males exposed to 1 mg/L test substance lost or failed to gain body weight by Day 1 and/or Day 3. All animals surpassed their initial body weight by Day 7 and gained body weight through Day 14.
The second exposure to 5 mg/L induced body weight loss in all males which reached or surpassed their initial body weight latest by Day 7.

Gross pathology:

Gross necropsy of deceased males revealed light coloring of the lungs (3/3 males) and distended stomachs (2/3 males). Surviving animals of the first and second experiment (expsoure to 1 and 5 mg/L) did not reveal gross abnormalities.

Table 2: Particle size distribution

 

Experiment	Exposure level (target concentration) [mg/L]	Sample No	Stage	Cut-Off diameter (µm)	Cumulative (%)*
1^stexperiment	1	1	0	9.0	91.1
			1	5.8	80.4
			2	4.7	68.5
			3	3.3	45.2
			4	2.1	26.8
			5	1.1	6.0
			6	0.7	3.0
			7	0.4	0.8
			F	0.0	0.0
		2	0	9.0	94.8
			1	5.8	86.7
			2	4.7	78.6
			3	3.3	57.7
			4	2.1	38.3
			5	1.1	18.1
			6	0.7	10.1
			7	0.4	4.8
			F	0.0	0.0
	5	1	0	9.0	88.3
			1	5.8	70.1
			2	4.7	56.0
			3	3.3	36.7
			4	2.1	21.3
			5	1.1	16.4
			6	0.7	13.7
			7	0.4	9.9
			F	0.0	0.0
		2	0	9.0	88.4
			1	5.8	72.0
			2	4.7	57.4
			3	3.3	35.5
			4	2.1	19.8
			5	1.1	13.9
			6	0.7	10.7
			7	0.4	8.8
			F	0.0	0.0
2^ndexperiment	5	1	0	9.0	94.7
			1	5.8	78.9
			2	4.7	70.2
			3	3.3	48.3
			4	2.1	30.9
			5	1.1	10.6
			6	0.7	3.8
			7	0.4	1.9
			F	0.0	0.0
		2	0	9.0	97.1
			1	5.8	87.6
			2	4.7	77.6
			3	3.3	55.8
			4	2.1	35.0
			5	1.1	12.1
			6	0.7	4.5
			7	0.4	2.1
			F	0.0	0.0

*: Percent of particles smaller than the corresponding effective cut-off diameter

Table 3: Chamber concentrations

 

Experiment	Exposure level (target concentration) [mg/L]	Average Total airflow [Lpm]	Total time of exposure (incl. the time until reaching equilibrium) [min]	Nominal concentration [mg/L]	Actual chamber concentration [mg/L]
1^stexperiment	1	46.0	243	5.29	1.03 ± 0.02
	5	46.0	241	19.28	5.15 ± 0.13
2^ndexperiment	5	46.0	241	21.14	5.17 ± 0.31

 

Table 4: Summary of particle size distribution (MMAD)

Experiment	Exposure Level (nominal) [mg/L]	Sample No.	Sampling time [h]	Collection time [min]	MMAD [µm]
1^stexperiment	1	1	1.5	1	3.2 ± 2.41
		2	3	1	2.35 ± 2.42
	Mean				2.78 ± 2.42
	5	1	1.5	1	3.38 ± 3.07
		2	3	1	3.344 ± 2.97
	Mean				3.41 ± 3.02
2^ndexperiment	5	1	1.5	1	2.90 ± 2.37
		2	3	1	2.47 ± 2.24
	Mean				2.69 ± 2.31

 

Table 5: Summary table of toxicological results

 

Experiment	Target concentration [mg/L air]	Toxicological results*	Duration of clinical signs	Time of death	Mortality
	Males
1^s texperiment	1.03	0/5/5	Day 0 - 2	---	0
	5.15	3/2/5	Day 0-2	At chamber removal	3
2^ndexperiment	5.17	0/5/5	Day 0-3	---	0
	Females
1^stexperiment	5.15	0/5/5	Day 0 -5	---	0
	LC50 > 5 mg/L air

                                                                                           

* first number = number of dead animals                                 

 second number = number of animals with clinical signs         

 third number = number of animals used

Table 6: Individual body weights and mortality

Experiment	Target concentration [mg/L air]	Gender	Body weight [g]					Mortality (Day / body weight [g])
			Initial	Day 1	Day 3	Day 7	Day 14
1^stexperiment	1	M	293	291	302	318	341
			284	282	281	294	312
			302	302	301	322	343
			292	290	291	301	315
			306	303	304	322	339
	5	M	268					0 / 260
			253	234	250	264	296
			272					0 / 264
			260					0 / 251
			264	253	270	284	327
	5	F	214	190	207	217	236
			199	185	203	216	230
			220	211	217	212	245
			212	197	212	218	241
			190	183	192	190	210
2^ndexperiment	5	M	275	264	277	306	348
			241	228	234	244	265
			250	239	247	270	298
			242	221	226	242	270
			256	232	242	262	298

                                                                                           

 

 

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

CLP: not classified
DSD: not classified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Quality of whole database:

The available information comprises an adequate and reliable study (Klimisch score 1), and is thus sufficient to fulfil the standard information requirements set out in Annex VIII, 8.5, of Regulation (EC) No 1907/2006.

Acute toxicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Acute Toxicity

Oral Route

A GLP-guideline study according to the acute toxic class method (OECD 423) did not determine mortality or any clinical signs of toxicity after oral administration of 2000 mg Fatty acids, C16-18 (even numbered), aluminum salt/kg bw (Lütkenhaus, 2013a). Pathological examinations did not reveal treatment related gross pathological findings. Thus, the LD50 was > 2000 mg/kg bw.

 

Inhalation

Acute toxicity of Fatty acids, C16-18 (even numbered), aluminum salts following inhalation was studied in a GLP-guideline study according to OECD guideline 403 (Durando,

Acute toxicity of Fatty acids, C16-18 (even numbered), aluminum salts following inhalation was studied in a GLP-guideline study according to OECD guideline 403 (Peter Greven, 2013). Initially, 5 Sprague Dawley rats/sex were exposed to a nominal concentration of 5 mg/L of the dust aerosol for 4 hours in a nose-only exposure system. The gravimetric and nominal chamber concentrations were determined to be 5.15 and 19.28 mg/L, respectively. The average mass median aerodynamic diameter (MMAD) was calculated to be 3.41 ± 3.02 µm. No mortality was observed in the female group until the end of the 14 -day observation period. In the male group, 3/5 animals were found dead at chamber removal. Gross necropsy of the deceased revealed light coloring of the lungs (3/3 males) and distended stomachs (2/3 males). Surviving animals exhibited irregular respiration, red nasal discharge and moist or dry rales that resolved in all animals by study Day 5. Body weight loss was observed in all animals but at the end of the observation period, all animals surpassed their initial body weight. No gross abnormalities were noted in surviving animals necropsied at the end of the 14-day observation period. Following the standard protocol according to OECD guideline 403, a lower dose of 1 mg/L was tested subsequently in 5 further male Sprague Dawley rats based on the mortality observed in the male group. The gravimetric and nominal chamber concentrations were 1.03 and 5.29 mg/L, respectively, and a MMAD of 2.78 ± 2.42 µm was calculated. Up to the end of the 14 -day observation period, no male died after exposure to 1 mg/mL. Irregular respiration was observed in all males but they recovered by Day 3 and appeared active and healthy up to the end of the observation period. Body weight loss was observed but all animals surpassed their initial body weight by Day 7.

In addition to the first experiment, a further acute toxicity study following inhalation to the limit dose 5 mg/mL was considered necessary as the reason for mortality seen only in the male group remained unclear especially as Fatty acids, 16-18 (even numbered), aluminum salts seem not to have intrinsic properties leading to toxicity. Therefore, a second experiment including 5 males was conducted with a nominal concentration of 5 mg/L according to the same protocol. Gravimetric and nominal chamber concentrations of 5.17 and 21.14 mg/L, respectively, were determined and a MMAD of 2.69 ± 2.31 was calculated. No mortality was observed in this experiment until the end of the 14 -day observation period. Observed clinical symptoms included irregular respiration, nasal discharge, moist rales and/or hypoactivity. All males appeared active and healthy latest on Day 4. Body weight loss was observed in all males during the first days post exposure, but their initial body weight was surpassed latest by Day 7. No abnormalities were observed in gross pathology.

In conclusion, following a weight-of-evidence approach including both experiments, 0/5 females and 3/10 males died after exposure to > 5 mg/L test substance (nominal) following inhalation. Thus, a LC50 > 5 mg/L was established for male and female rats according to the conducted study.

Justification for selection of acute toxicity – oral endpoint
There is only one study available.

Justification for selection of acute toxicity – inhalation endpoint
There is only one study available.

Justification for selection of acute toxicity – dermal endpoint
Testing by the dermal route is not appropriate in accordance with Column 2 of Annex VIII, Section 8.5, since information on acute toxicity by the oral and inhalation route is provided and no significant rate of dermal absorption is anticipated. Furthermore, the available data on physico-chemical and toxicological properties provide sufficient weight of evidence leading to the conclusion that the substance is not acutely toxic by the dermal route. Thus in vivo testing for acute dermal toxicity does not appear scientifically necessary and shall be omitted in accordance with Annex XI, Section 1.2 of Regulation (EC) No 1907/2006.

Justification for classification or non-classification

The available data on acute oral and inhalation toxicity do not meet the classification criteria according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.